Fluoride and arsenic exposure affects spatial memory and activates the ERK/CREB signaling pathway in offspring rats.

PubMed

Zhu, Yu-Peng; Xi, Shu-Hua; Li, Ming-Yan; Ding, Ting-Ting; Liu, Nan; Cao, Fu-Yuan; Zeng, Yang; Liu, Xiao-Jing; Tong, Jun-Wang; Jiang, Shou-Fang

2017-03-01

Fluoride and arsenic are inorganic contaminants that occur in the natural environment. Chronic fluoride and/or arsenic exposure can induce developmental neurotoxicity and negatively influence intelligence in children, although the underlying molecular mechanisms are poorly understood. This study explored the effects of fluoride and arsenic exposure in drinking water on spatial learning, memory and key protein expression in the ERK/CREB signaling pathway in hippocampal and cerebral cortex tissue in rat offspring. Pregnant rats were divided into four groups. Control rats drank tap water, while rats in the three exposure groups drank water with sodium fluoride (100mg/L), sodium arsenite (75mg/L), and a sodium fluoride (100mg/L) and sodium arsenite (75mg/L) combination during gestation and lactation. After weaning, rat pups drank the same solution as their mothers. Spatial learning and memory ability of pups at postnatal day 21 (PND21) and postnatal day 42 (PND42) were measured using a Morris water maze. ERK, phospho-ERK (p-ERK), CREB and phospho-CREB (p-CREB) protein expression in the hippocampus and cerebral cortex was detected using Western blot. Compared with the control pups, escape latencies increased in PND42 pups exposed to arsenic and co-exposed to fluoride and arsenic, and the short-term and long-term spatial memory ability declined in pups exposed to fluoride and arsenic, both alone and in combination. Compared with controls, ERK and p-ERK levels decreased in the hippocampus and cerebral cortex in pups exposed to combined fluoride and arsenic. CREB protein expression in the cerebral cortex decreased in pups exposed to fluoride, arsenic, and the fluoride and arsenic combination. p-CREB protein expression in both the hippocampus and cerebral cortex was decreased in pups exposed to fluoride and arsenic in combination compared to the control group. There were negative correlation between the proteins expression and escape latency periods in pups. These data indicate that exposure to fluoride and arsenic in early life stage changes ERK, p-ERK, CREB and p-CREB protein expression in the hippocampus and cerebral cortex of rat offspring at PND21 and PND 42, which may contribute to impaired neurodevelopment following exposure. Copyright © 2017 Elsevier B.V. All rights reserved.

Differential transcriptional activation by human T-cell leukemia virus type 1 Tax mutants is mediated by distinct interactions with CREB binding protein and p300.

PubMed

Bex, F; Yin, M J; Burny, A; Gaynor, R B

1998-04-01

The human T-cell leukemia virus type 1 Tax protein transforms human T lymphocytes, which can lead to the development of adult T-cell leukemia. Tax transformation is related to its ability to activate gene expression via the ATF/CREB and the NF-kappaB pathways. Transcriptional activation of these pathways is mediated by the actions of the related coactivators CREB binding protein (CBP) and p300. In this study, immunocytochemistry and confocal microscopy were used to localize CBP and p300 in cells expressing wild-type Tax or Tax mutants that are able to selectively activate gene expression from either the NF-kappaB or ATF/CREB pathway. Wild-type Tax colocalized with both CBP and p300 in nuclear bodies which also contained ATF-1 and the RelA subunit of NF-kappaB. However, a Tax mutant that selectively activates gene expression from only the ATF/CREB pathway colocalized with CBP but not p300, while a Tax mutant that selectively activates gene expression from only the NF-kappaB pathway colocalized with p300 but not CBP. In vitro and in vivo protein interaction studies indicated that the integrity of two independent domains of Tax delineated by these mutants was involved in the direct interaction of Tax with either CBP or p300. These studies are consistent with a model in which activation of either the NF-kappaB or the ATF/CREB pathway by specific Tax mutants is mediated by distinct interactions with related coactivator proteins.

Hypoxia-induced endothelial NO synthase gene transcriptional activation is mediated through the tax-responsive element in endothelial cells.

PubMed

Min, Jiho; Jin, Yoon-Mi; Moon, Je-Sung; Sung, Min-Sun; Jo, Sangmee Ahn; Jo, Inho

2006-06-01

Although hypoxia is known to induce upregulation of endothelial NO synthase (eNOS) gene expression, the underlying mechanism is largely unclear. In this study, we show that hypoxia increases eNOS gene expression through the binding of phosphorylated cAMP-responsive element binding (CREB) protein (pCREB) to the eNOS gene promoter. Hypoxia (1% O2) increased both eNOS expression and NO production, peaking at 24 hours, in bovine aortic endothelial cells, and these increases were accompanied by increases in pCREB. Treatment with the protein kinase A inhibitor H-89 or transfection with dominant-negative inhibitor of CREB reversed the hypoxia-induced increases in eNOS expression and NO production, with concomitant inhibition of the phosphorylation of CREB induced by hypoxia, suggesting an involvement of protein kinase A/pCREB-mediated pathway. To map the regulatory elements of the eNOS gene responsible for pCREB binding under hypoxia, we constructed an eNOS gene promoter (-1600 to +22 nucleotides) fused with a luciferase reporter gene [pGL2-eNOS(-1600)]. Hypoxia (for 24-hour incubation) increased the promoter activity by 2.36+/-0.18-fold in the bovine aortic endothelial cells transfected with pGL2-eNOS(-1600). However, progressive 5'-deletion from -1600 to -873 completely attenuated the hypoxia-induced increase in promoter activity. Electrophoretic mobility shift, anti-pCREB antibody supershift, and site-specific mutation analyses showed that pCREB is bound to the Tax-responsive element (TRE) site, a cAMP-responsive element-like site, located at -924 to -921 of the eNOS promoter. Our data demonstrate that the interaction between pCREB and the Tax-responsive element site within the eNOS promoter may represent a novel mechanism for the mediation of hypoxia-stimulated eNOS gene expression.

Novel mechanisms and signaling pathways of esophageal ulcer healing: the role of prostaglandin EP2 receptors, cAMP, and pCREB

PubMed Central

Ahluwalia, Amrita; Baatar, Dolgor; Jones, Michael K.

2014-01-01

Clinical studies indicate that prostaglandins of E class (PGEs) may promote healing of tissue injury e.g., gastroduodenal and dermal ulcers. However, the precise roles of PGEs, their E-prostanoid (EP) receptors, signaling pathways including cAMP and cAMP response element-binding protein (CREB), and their relation to VEGF and angiogenesis in the tissue injury healing process remain unknown, forming the rationale for this study. Using an esophageal ulcer model in rats, we demonstrated that esophageal mucosa expresses predominantly EP2 receptors and that esophageal ulceration triggers an increase in expression of the EP2 receptor, activation of CREB (the downstream target of the cAMP signaling), and enhanced VEGF gene expression. Treatment of rats with misoprostol, a PGE1 analog capable of activating EP receptors, enhanced phosphorylation of CREB, stimulated VEGF expression and angiogenesis, and accelerated esophageal ulcer healing. In cultured human esophageal epithelial (HET-1A) cells, misoprostol increased intracellular cAMP levels (by 163-fold), induced phosphorylation of CREB, and stimulated VEGF expression. A cAMP analog (Sp-cAMP) mimicked, whereas an inhibitor of cAMP-dependent protein kinase A (Rp-cAMP) blocked, these effects of misoprostol. These results indicate that the EP2/cAMP/protein kinase A pathway mediates the stimulatory effect of PGEs on angiogenesis essential for tissue injury healing via the induction of CREB activity and VEGF expression. PMID:25059824

Chronic Enhancement of CREB Activity in the Hippocampus Interferes with the Retrieval of Spatial Information

ERIC Educational Resources Information Center

Viosca, Jose; Malleret, Gael; Bourtchouladze, Rusiko; Benito, Eva; Vronskava, Svetlana; Kandel, Eric R.; Barco, Angel

2009-01-01

The activation of cAMP-responsive element-binding protein (CREB)-dependent gene expression is thought to be critical for the formation of different types of long-term memory. To explore the consequences of chronic enhancement of CREB function on spatial memory in mammals, we examined spatial navigation in bitransgenic mice that express in a…

The ubiquitin-proteasome system is necessary for long-term synaptic depression in Aplysia.

PubMed

Fioravante, Diasinou; Liu, Rong-Yu; Byrne, John H

2008-10-08

The neuropeptide Phe-Met-Arg-Phe-NH(2) (FMRFa) can induce transcription-dependent long-term synaptic depression (LTD) in Aplysia sensorimotor synapses. We investigated the role of the ubiquitin-proteasome system and the regulation of one of its components, ubiquitin C-terminal hydrolase (ap-uch), in LTD. LTD was sensitive to presynaptic inhibition of the proteasome and was associated with upregulation of ap-uch mRNA and protein. This upregulation appeared to be mediated by CREB2, which is generally regarded as a transcription repressor. Binding of CREB2 to the promoter region of ap-uch was accompanied by histone hyperacetylation, suggesting that CREB2 cannot only inhibit but also promote gene expression. CREB2 was phosphorylated after FMRFa, and blocking phospho-CREB2 blocked LTD. In addition to changes in the expression of ap-uch, the synaptic vesicle-associated protein synapsin was downregulated in LTD in a proteasome-dependent manner. These results suggest that proteasome-mediated protein degradation is engaged in LTD and that CREB2 may act as a transcription activator under certain conditions.

Tunable regulation of CREB DNA binding activity couples genotoxic stress response and metabolism

PubMed Central

Kim, Sang Hwa; Trinh, Anthony T.; Larsen, Michele Campaigne; Mastrocola, Adam S.; Jefcoate, Colin R.; Bushel, Pierre R.; Tibbetts, Randal S.

2016-01-01

cAMP response element binding protein (CREB) is a key regulator of glucose metabolism and synaptic plasticity that is canonically regulated through recruitment of transcriptional coactivators. Here we show that phosphorylation of CREB on a conserved cluster of Ser residues (the ATM/CK cluster) by the DNA damage-activated protein kinase ataxia-telangiectasia-mutated (ATM) and casein kinase1 (CK1) and casein kinase2 (CK2) positively and negatively regulates CREB-mediated transcription in a signal dependent manner. In response to genotoxic stress, phosphorylation of the ATM/CK cluster inhibited CREB-mediated gene expression, DNA binding activity and chromatin occupancy proportional to the number of modified Ser residues. Paradoxically, substoichiometric, ATM-independent, phosphorylation of the ATM/CK cluster potentiated bursts in CREB-mediated transcription by promoting recruitment of the CREB coactivator, cAMP-regulated transcriptional coactivators (CRTC2). Livers from mice expressing a non-phosphorylatable CREB allele failed to attenuate gluconeogenic genes in response to DNA damage or fully activate the same genes in response to glucagon. We propose that phosphorylation-dependent regulation of DNA binding activity evolved as a tunable mechanism to control CREB transcriptional output and promote metabolic homeostasis in response to rapidly changing environmental conditions. PMID:27431323

The cAMP Response Element Binding protein (CREB) is activated by Insulin-like Growth Factor-1 (IGF-1) and regulates myostatin gene expression in skeletal myoblast

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zuloaga, R.; Fuentes, E.N.; Molina, A.

2013-10-18

Highlights: •IGF-1 induces the activation of CREB via IGF-1R/PI3K/PLC signaling pathway. •Calcium dependent signaling pathways regulate myostatin gene expression. •IGF-1 regulates myostatin gene expression via CREB transcription in skeletal myoblast. -- Abstract: Myostatin, a member of the Transforming Growth Factor beta (TGF-β) superfamily, plays an important role as a negative regulator of skeletal muscle growth and differentiation. We have previously reported that IGF-1 induces a transient myostatin mRNA expression, through the activation of the Nuclear Factor of Activated T cells (NFAT) in an IP{sub 3}/calcium-dependent manner. Here we examined the activation of CREB transcription factor as downstream targets of IGF-1more » during myoblast differentiation and its role as a regulator of myostatin gene expression. In cultured skeletal myoblast, IGF-1 induced the phosphorylation and transcriptional activation of CREB via IGF-1 Receptor/Phosphatidylinositol 3-Kinase (PI3K)/Phospholipase C gamma (PLC γ), signaling pathways. Also, IGF-1 induced calcium-dependent molecules such as Calmodulin Kinase II (CaMK II), Extracellular signal-regulated Kinases (ERK), Protein Kinase C (PKC). Additionally, we examined myostatin mRNA levels and myostatin promoter activity in differentiated myoblasts stimulated with IGF-1. We found a significant increase in mRNA contents of myostatin and its reporter activity after treatment with IGF-1. The expression of myostatin in differentiated myoblast was downregulated by the transfection of siRNA–CREB and by pharmacological inhibitors of the signaling pathways involved in CREB activation. By using pharmacological and genetic approaches together these data demonstrate that IGF-1 regulates the myostatin gene expression via CREB transcription factor during muscle cell differentiation.« less

Computational analysis of human and mouse CREB3L4 Protein

PubMed Central

Velpula, Kiran Kumar; Rehman, Azeem Abdul; Chigurupati, Soumya; Sanam, Ramadevi; Inampudi, Krishna Kishore; Akila, Chandra Sekhar

2012-01-01

CREB3L4 is a member of the CREB/ATF transcription factor family, characterized by their regulation of gene expression through the cAMP-responsive element. Previous studies identified this protein in mice and humans. Whereas CREB3L4 in mice (referred to as Tisp40) is found in the testes and functions in spermatogenesis, human CREB3L4 is primarily detected in the prostate and has been implicated in cancer. We conducted computational analyses to compare the structural homology between murine Tisp40α human CREB3L4. Our results reveal that the primary and secondary structures of the two proteins contain high similarity. Additionally, predicted helical transmembrane structure reveals that the proteins likely have similar structure and function. This study offers preliminary findings that support the translation of mouse Tisp40α findings into human models, based on structural homology. PMID:22829733

Mechanisms of protein kinase C signaling in the modulation of 3',5'-cyclic adenosine monophosphate-mediated steroidogenesis in mouse gonadal cells.

PubMed

Manna, Pulak R; Huhtaniemi, Ilpo T; Stocco, Douglas M

2009-07-01

The protein kinase C (PKC) signaling pathway plays integral roles in the expression of the steroidogenic acute regulatory (StAR) protein that regulates steroid biosynthesis in steroidogenic cells. PKC can modulate the activity of cAMP/protein kinase A signaling involved in steroidogenesis; however, its mechanism remains obscure. In the present study, we demonstrate that activation of the PKC pathway, by phorbol 12-myristate 13-acetate (PMA), was capable of potentiating dibutyryl cAMP [(Bu)(2)cAMP]-stimulated StAR expression, StAR phosphorylation, and progesterone synthesis in both mouse Leydig (MA-10) and granulosa (KK-1) tumor cells. The steroidogenic potential of PMA and (Bu)(2)cAMP was linked with phosphorylation of ERK 1/2; however, inhibition of the latter demonstrated varying effects on steroidogenesis. Transcriptional activation of the StAR gene by PMA and (Bu)(2)cAMP was influenced by several factors, its up-regulation being dependent on phosphorylation of the cAMP response element binding protein (CREB). An oligonucleotide probe containing a CREB/activating transcription factor binding region in the StAR promoter was found to bind nuclear proteins in PMA and (Bu)(2)cAMP-treated MA-10 and KK-1 cells. Chromatin immunoprecipitation studies revealed that the induction of phosphorylated CREB was tightly correlated with in vivo protein-DNA interactions and recruitment of CREB binding protein to the StAR promoter. Ectopic expression of CREB binding protein enhanced CREB-mediated transcription of the StAR gene, an event that was markedly repressed by the adenovirus E1A oncoprotein. Further studies demonstrated that the activation of StAR expression and steroid synthesis by PMA and (Bu)(2)cAMP was associated with expression of the nuclear receptor Nur77, indicating its essential role in hormone-regulated steroidogenesis. Collectively, these findings provide insight into the mechanisms by which PKC modulates cAMP/protein kinase A responsiveness involved in regulating the steroidogenic response in mouse gonadal cells.

Novel mechanism of transcriptional regulation of cell matrix protein through CREB

PubMed Central

Habib, Samy L; Mohan, Sumathy; Liang, Sitai; Li, Baojie; Yadav, Mukesh

2015-01-01

The transcription mechanism(s) of renal cell matrix accumulation in diabetes does not explored. Phosphorylation of the transcription factor cAMP-responsive element binding protein (CREB) significantly increased in cells treated with high glucose (HG) compared to cell grown in normal glucose (NG). Cells pretreated with rapamycin before exposure to HG showed significant decrease phosphorylation of CREB, increase in AMPK activity and decrease protein/mRNA and promoter activity of fibronectin. In addition, cells transfected with siRNA against CREB showed significant increase in AMPK activity, decrease in protein/mRNA and promoter activity of fibronectin. Cells treated with HG showed nuclear localization of p-CREB while pretreated cells with rapamycin reversed HG effect. Moreover, gel shift analysis shows increase binding of CREB to fibronectin promoter in cells treated with HG while cells pretreated with rapamycin reversed the effect of HG. Furthermore, db/db mice treated with rapamycin showed significant increase in AMPK activity, decrease in expression of p-CREB and protein/mRNA of fibronectin. Strong staining of fibronectin and p-CREB was detected in kidney cortex of db/db mice while treated mice with rapamycin reversed hyperglycemia effect. In summary, our data provide a novel mechanism of transcriptional regulation of fibronectin through CREB that may be used as therapeutic approach to prevent diabetes complications. PMID:26115221

CREB expression in the brains of two closely related parasitic wasp species that differ in long-term memory formation.

PubMed

van den Berg, M; Verbaarschot, P; Hontelez, S; Vet, L E M; Dicke, M; Smid, H M

2010-06-01

The cAMP/PKA signalling pathway and transcription factor cAMP response element-binding protein (CREB) play key roles in long-term memory (LTM) formation. We used two closely related parasitic wasp species, Cotesia glomerata and Cotesia rubecula, which were previously shown to be different in LTM formation, and sequenced at least nine different CREB transcripts in both wasp species. The splicing patterns, functional domains and amino acid sequences were similar to those found in the CREB genes of other organisms. The predicted amino acid sequences of the CREB isoforms were identical in both wasp species. Using real-time quantitative PCR we found that two low abundant CREB transcripts are differentially expressed in the two wasps, whereas the expression levels of high abundant transcripts are similar.

Activation of the kinase activity of ATM by retinoic acid is required for CREB-dependent differentiation of neuroblastoma cells.

PubMed

Fernandes, Norvin D; Sun, Yingli; Price, Brendan D

2007-06-01

The ATM protein kinase is mutated in ataxia telangiectasia, a genetic disease characterized by defective DNA repair, neurodegeneration, and growth factor signaling defects. The activity of ATM kinase is activated by DNA damage, and this activation is required for cells to survive genotoxic events. In addition to this well characterized role in DNA repair, we now demonstrate a novel role for ATM in the retinoic acid (RA)-induced differentiation of SH-SY5Y neuroblastoma cells into post-mitotic, neuronal-like cells. RA rapidly activates the activity of ATM kinase, leading to the ATM-dependent phosphorylation of the CREB protein, extrusion of neuritic processes, and differentiation of SH-SY5Y cells into neuronal-like cells. When ATM protein expression was suppressed by short hairpin RNA, the ATM-dependent phosphorylation of CREB was blocked. Furthermore, ATM-negative cells failed to differentiate into neuronal-like cells when exposed to retinoic acid; instead, they underwent cell death. Expression of a constitutively active CREBVP16 construct, or exposure to forskolin to induce CREB phosphorylation, rescued ATM negative cells and restored differentiation. Furthermore, when dominant negative CREB proteins with mutations in either the CREB phosphorylation site (CREBS133A) or the DNA binding domain (KCREB) were introduced into SH-SY5Y cells, retinoic acid-induced differentiation was blocked and the cells underwent cell death. The results demonstrate that ATM is required for the retinoic acid-induced differentiation of SH-SY5Y cells through the ATM dependent-phosphorylation of serine 133 of CREB. These results therefore define a novel mechanism for activation of the activity of ATM kinase by RA, and implicate ATM in the regulation of CREB function during RA-induced differentiation.

Angiotensin II regulates brain (pro)renin receptor expression through activation of cAMP response element-binding protein

PubMed Central

Li, Wencheng; Liu, Jiao; Hammond, Sean L.; Tjalkens, Ronald B.; Saifudeen, Zubaida

2015-01-01

We reported that brain (pro)renin receptor (PRR) expression levels are elevated in DOCA-salt-induced hypertension; however, the underlying mechanism remained unknown. To address whether ANG II type 1 receptor (AT1R) signaling is involved in this regulation, we implanted a DOCA pellet and supplied 0.9% saline as the drinking solution to C57BL/6J mice. Sham pellet-implanted mice that were provided regular drinking water served as controls. Concurrently, mice were intracerebroventricularly infused with the AT1R blocker losartan, angiotensin-converting-enzyme inhibitor captopril, or artificial cerebrospinal fluid for 3 wk. Intracerebroventricular infusion of losartan or captopril attenuated DOCA-salt-induced PRR mRNA elevation in the paraventricular nucleus of the hypothalamus, suggesting a role for ANG II/AT1R signaling in regulating PRR expression during DOCA-salt hypertension. To test which ANG II/AT1R downstream transcription factors were involved in PRR regulation, we treated Neuro-2A cells with ANG II with or without CREB (cAMP response element-binding protein) or AP-1 (activator protein-1) inhibitors, or CREB siRNA. CREB and AP-1 inhibitors, as well as CREB knockdown abolished ANG II-induced increases in PRR levels. ANG II also induced PRR upregulation in primary cultured neurons. Chromatin immunoprecipitation assays revealed that ANG II treatment increased CREB binding to the endogenous PRR promoter in both cultured neurons and hypothalamic tissues of DOCA-salt hypertensive mice. This increase in CREB activity was reversed by AT1R blockade. Collectively, these findings indicate that ANG II acts via AT1R to upregulate PRR expression both in cultured cells and in DOCA-salt hypertensive mice by increasing CREB binding to the PRR promoter. PMID:25994957

The ERK/CREB pathway is involved in the c-Ski expression induced by low TGF-β1 concentrations during primary fibroblast proliferation.

PubMed

Li, Ping; Liu, Ping; Peng, Yan; Zhang, Zhuo-Hang; Li, Xiao-Ming; Xiong, Ren-Ping; Chen, Xing; Zhao, Yan; Ning, Ya-Lei; Yang, Nan; Zhang, Bo; Zhou, Yuan-Guo

2018-06-27

Increasing evidence has suggested that bidirectional regulation of cell proliferation is one important effect of TGF-β1 in wound healing. Increased c-Ski expression plays a role in promoting fibroblast proliferation at low TGF-β1 concentrations, but the mechanism by which low TGF-β1 concentrations regulate c-Ski levels remains unclear. In this study, the proliferation of rat primary fibroblasts was assessed with an ELISA BrdU kit. The mRNA and protein expression and phosphorylation levels of corresponding factors were measured by RT-qPCR, immunohistochemistry or Western blotting. We first found that low TGF-β1 concentrations not only promoted c-Ski mRNA and protein expression in rat primary fibroblasts but also increased the phosphorylation levels of Extracellular Signal-Regulated Kinases (ERK) and cAMP response element binding (CREB) protein. An ERK kinase (mitogen-activated protein kinase kinase, MEK) inhibitor significantly inhibited ERK1/2 phosphorylation levels, markedly reducing c-Ski expression and CREB phosphorylation levels and abrogating the growth-promoting effect of low TGF-β1 concentrations. At the same time, Smad2/3 phosphorylation levels were not significantly changed. Taken together, these results suggest that the increased cell proliferation induced by low TGF-β1 concentrations mediates c-Ski expression potentially through the ERK/CREB pathway rather than through the classic TGF-β1/Smad pathway.

Presenilins regulate neurotrypsin gene expression and neurotrypsin-dependent agrin cleavage via cyclic AMP response element-binding protein (CREB) modulation.

PubMed

Almenar-Queralt, Angels; Kim, Sonia N; Benner, Christopher; Herrera, Cheryl M; Kang, David E; Garcia-Bassets, Ivan; Goldstein, Lawrence S B

2013-12-06

Presenilins, the catalytic components of the γ-secretase complex, are upstream regulators of multiple cellular pathways via regulation of gene transcription. However, the underlying mechanisms and the genes regulated by these pathways are poorly characterized. In this study, we identify Tequila and its mammalian ortholog Prss12 as genes negatively regulated by presenilins in Drosophila larval brains and mouse embryonic fibroblasts, respectively. Prss12 encodes the serine protease neurotrypsin, which cleaves the heparan sulfate proteoglycan agrin. Altered neurotrypsin activity causes serious synaptic and cognitive defects; despite this, the molecular processes regulating neurotrypsin expression and activity are poorly understood. Using γ-secretase drug inhibitors and presenilin mutants in mouse embryonic fibroblasts, we found that a mature γ-secretase complex was required to repress neurotrypsin expression and agrin cleavage. We also determined that PSEN1 endoproteolysis or processing of well known γ-secretase substrates was not essential for this process. At the transcriptional level, PSEN1/2 removal induced cyclic AMP response element-binding protein (CREB)/CREB-binding protein binding, accumulation of activating histone marks at the neurotrypsin promoter, and neurotrypsin transcriptional and functional up-regulation that was dependent on GSK3 activity. Upon PSEN1/2 reintroduction, this active epigenetic state was replaced by a methyl CpG-binding protein 2 (MeCP2)-containing repressive state and reduced neurotrypsin expression. Genome-wide analysis revealed hundreds of other mouse promoters in which CREB binding is similarly modulated by the presence/absence of presenilins. Our study thus identifies Tequila and neurotrypsin as new genes repressed by presenilins and reveals a novel mechanism used by presenilins to modulate CREB signaling based on controlling CREB recruitment.

Presenilins Regulate Neurotrypsin Gene Expression and Neurotrypsin-dependent Agrin Cleavage via Cyclic AMP Response Element-binding Protein (CREB) Modulation*

PubMed Central

Almenar-Queralt, Angels; Kim, Sonia N.; Benner, Christopher; Herrera, Cheryl M.; Kang, David E.; Garcia-Bassets, Ivan; Goldstein, Lawrence S. B.

2013-01-01

Presenilins, the catalytic components of the γ-secretase complex, are upstream regulators of multiple cellular pathways via regulation of gene transcription. However, the underlying mechanisms and the genes regulated by these pathways are poorly characterized. In this study, we identify Tequila and its mammalian ortholog Prss12 as genes negatively regulated by presenilins in Drosophila larval brains and mouse embryonic fibroblasts, respectively. Prss12 encodes the serine protease neurotrypsin, which cleaves the heparan sulfate proteoglycan agrin. Altered neurotrypsin activity causes serious synaptic and cognitive defects; despite this, the molecular processes regulating neurotrypsin expression and activity are poorly understood. Using γ-secretase drug inhibitors and presenilin mutants in mouse embryonic fibroblasts, we found that a mature γ-secretase complex was required to repress neurotrypsin expression and agrin cleavage. We also determined that PSEN1 endoproteolysis or processing of well known γ-secretase substrates was not essential for this process. At the transcriptional level, PSEN1/2 removal induced cyclic AMP response element-binding protein (CREB)/CREB-binding protein binding, accumulation of activating histone marks at the neurotrypsin promoter, and neurotrypsin transcriptional and functional up-regulation that was dependent on GSK3 activity. Upon PSEN1/2 reintroduction, this active epigenetic state was replaced by a methyl CpG-binding protein 2 (MeCP2)-containing repressive state and reduced neurotrypsin expression. Genome-wide analysis revealed hundreds of other mouse promoters in which CREB binding is similarly modulated by the presence/absence of presenilins. Our study thus identifies Tequila and neurotrypsin as new genes repressed by presenilins and reveals a novel mechanism used by presenilins to modulate CREB signaling based on controlling CREB recruitment. PMID:24145027

Small Molecule Inhibition of cAMP Response Element Binding Protein in Human Acute Myeloid Leukemia Cells

PubMed Central

Mitton, Bryan; Chae, Hee-Don; Hsu, Katie; Dutta, Ritika; Aldana-Masangkay, Grace; Ferrari, Roberto; Davis, Kara; Tiu, Bruce C.; Kaul, Arya; Lacayo, Norman; Dahl, Gary; Xie, Fuchun; Li, Bingbing X.; Breese, Marcus R.; Landaw, Elliot M.; Nolan, Garry; Pellegrini, Matteo; Romanov, Sergei; Xiao, Xiangshu; Sakamoto, Kathleen M.

2016-01-01

The transcription factor CREB (cAMP Response Element Binding Protein) is overexpressed in the majority of acute myeloid leukemia (AML) patients, and this is associated with a worse prognosis. Previous work revealed that CREB overexpression augmented AML cell growth, while CREB knockdown disrupted key AML cell functions in vitro. In contrast, CREB knockdown had no effect on long-term hematopoietic stem cell activity in mouse transduction/transplantation assays. Together, these studies position CREB as a promising drug target for AML. To test this concept, a small molecule inhibitor of CREB, XX-650-23, was developed. This molecule blocks a critical interaction between CREB and its required co-activator CBP (CREB Binding Protein), leading to disruption of CREB-driven gene expression. Inhibition of CBP-CREB interaction induced apoptosis and cell cycle arrest in AML cells, and prolonged survival in vivo in mice injected with human AML cells. XX-650-23 had little toxicity on normal human hematopoietic cells and tissues in mice. To understand the mechanism of XX-650-23, we performed RNA-seq, ChIP-seq and Cytometry Time of Flight with human AML cells. Our results demonstrate that small molecule inhibition of CBP-CREB interaction mostly affects apoptotic, cell cycle, and survival pathways, which may represent a novel approach for AML therapy. PMID:27211267

Sodium Benzoate, a Metabolite of Cinnamon and a Food Additive, Upregulates Ciliary Neurotrophic Factor in Astrocytes and Oligodendrocytes.

PubMed

Modi, Khushbu K; Jana, Malabendu; Mondal, Susanta; Pahan, Kalipada

2015-11-01

Ciliary neurotrophic factor (CNTF) is a promyelinating trophic factor that plays an important role in multiple sclerosis (MS). However, mechanisms by which CNTF expression could be increased in the brain are poorly understood. Recently we have discovered anti-inflammatory and immunomodulatory activities of sodium benzoate (NaB), a metabolite of cinnamon and a widely-used food additive. Here, we delineate that NaB is also capable of increasing the mRNA and protein expression of CNTF in primary mouse astrocytes and oligodendrocytes and primary human astrocytes. Accordingly, oral administration of NaB and cinnamon led to the upregulation of astroglial and oligodendroglial CNTF in vivo in mouse brain. Induction of experimental allergic encephalomyelitis, an animal model of MS, reduced the level of CNTF in the brain, which was restored by oral administration of cinnamon. While investigating underlying mechanisms, we observed that NaB induced the activation of protein kinase A (PKA) and H-89, an inhibitor of PKA, abrogated NaB-induced expression of CNTF. The activation of cAMP response element binding (CREB) protein by NaB, the recruitment of CREB and CREB-binding protein to the CNTF promoter by NaB and the abrogation of NaB-induced expression of CNTF in astrocytes by siRNA knockdown of CREB suggest that NaB increases the expression of CNTF via the activation of CREB. These results highlight a novel myelinogenic property of NaB and cinnamon, which may be of benefit for MS and other demyelinating disorders.

Coordinated activation of the secretory pathway during notochord formation in the Xenopus embryo.

PubMed

Tanegashima, Kosuke; Zhao, Hui; Rebbert, Martha L; Dawid, Igor B

2009-11-01

We compared the transcriptome in the developing notochord of Xenopus laevis embryos with that of other embryonic regions. A coordinated and intense activation of a large set of secretory pathway genes was observed in the notochord, but not in notochord precursors in the axial mesoderm at early gastrula stage. The genes encoding Xbp1 and Creb3l2 were also activated in the notochord. These two transcription factors are implicated in the activation of secretory pathway genes during the unfolded protein response, where cells react to the stress of a build-up of unfolded proteins in their endoplasmic reticulum. Xbp1 and Creb3l2 are differentially expressed but not differentially activated in the notochord. Reduction of expression of Xbp1 or Creb3l2 by injection of antisense morpholinos led to strong deficits in notochord but not somitic muscle development. In addition, the expression of some, but not all, genes encoding secretory proteins was inhibited by injection of xbp1 morpholinos. Furthermore, expression of activated forms of Xbp1 or Creb3l2 in animal explants could activate a similar subset of secretory pathway genes. We conclude that coordinated activation of a battery of secretory pathway genes mediated by Xbp1 and Creb/ATF factors is a characteristic and necessary feature of notochord formation.

Toll-like receptor 4-mediated cAMP production up-regulates B-cell activating factor expression in Raw264.7 macrophages.

PubMed

Moon, Eun-Yi; Lee, Yu-Sun; Choi, Wahn Soo; Lee, Mi-Hee

2011-10-15

B-cell activating factor (BAFF) plays a role in the generation and the maintenance of mature B cells. Lipopolysaccharide (LPS) increased BAFF expression through the activation of toll-like receptor 4 (TLR4)-dependent signal transduction. Here, we investigated the mechanism of action on mouse BAFF (mBAFF) expression by cAMP production in Raw264.7 mouse macrophages. mBAFF expression was increased by the treatment with a cAMP analogue, dibutyryl-cAMP which is the activator of protein kinase A (PKA), cAMP effector protein. PKA activation was measured by the phosphorylation of cAMP-response element binding protein (CREB) on serine 133 (S133). cAMP production and CREB (S133) phosphorylation were augmented by LPS-stimulation. While mBAFF promoter activity was enhanced by the co-transfection with pS6-RSV-CREB, it was reduced by siRNA-CREB. PKA inhibitor, H-89, reduced CREB (S133) phosphorylation and mBAFF expression in control and LPS-stimulated macrophages. Another principal cAMP effector protein is cAMP-responsive guanine nucleotide exchange factor (Epac), a Rap GDP exchange factor. Epac was activated by the treatment with 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate (CPT), Epac activator, as judged by the measurement of Rap1 activation. Basal level of mBAFF expression was increased by CPT treatment. LPS-stimulated mBAFF expression was also slightly enhanced by co-treatment with CPT. In addition, dibutyryl-cAMP and CPT enhanced mBAFF expression in bone marrow-derived macrophages (BMDM). With these data, it suggests that the activation of PKA and cAMP/Epac1/Rap1 pathways could be required for basal mBAFF expression, as well as being up-regulated in the TLR4-induced mBAFF expression. Crown Copyright © 2011. Published by Elsevier Inc. All rights reserved.

Requirement of the cyclic adenosine monophosphate response element-binding protein for hepatitis B virus replication.

PubMed

Kim, Bo Kyung; Lim, Seoung Ok; Park, Yun Gyu

2008-08-01

The cyclic adenosine monophosphate-response element (CRE)-transcription factor complex participates in the regulation of viral gene expression and pathologic processes caused by various viruses. The hepatitis B virus (HBV) enhancer I directs liver-specific transcription of viral genes and contains a CRE sequence (HBV-CRE); however, whether the HBV-CRE and CRE-binding protein (CREB) are required for the HBV life cycle remains to be determined. This study was designed to investigate the role of CREB in HBV replication and gene expression. Sequence-comparison analysis of 984 HBVs reported worldwide showed that the HBV-CRE sequence is highly conserved, indicating the possibility that it plays an important role in the HBV life cycle. The binding of CREB to the HBV-CRE site was markedly inhibited by oligonucleotides containing HBV-CRE and consensus CRE sequences in vitro and in vivo. The HBV promoter activity was demonstrated to be dependent upon the transactivation activity of CREB. Treatment with CRE decoy oligonucleotides reduced HBV promoter activity, and this was reversed by CREB overexpression. The levels of viral transcripts, DNA, and antigens were remarkably decreased in response to the overexpression of CREB mutants or treatment with the CRE decoy oligonucleotides, whereas enhancing CREB activity increased the levels of viral transcripts. In addition, introduction of a three-base mutation into the HBV-CRE led to a marked reduction in HBV messenger RNA synthesis. Taken together, our results demonstrate that both replication and gene expression of HBV require a functional CREB and HBV-CRE. We have also demonstrated that CRE decoy oligonucleotides and the overexpression of CREB mutants can effectively block the HBV life cycle, suggesting that interventions against CREB activity could provide a new avenue to treat HBV infection.

Identification of cholinergic and non-cholinergic neurons in the pons expressing phosphorylated cyclic adenosine monophosphate response element-binding protein as a function of rapid eye movement sleep.

PubMed

Datta, S; Siwek, D F; Stack, E C

2009-09-29

Recent studies have shown that in the pedunculopontine tegmental nucleus (PPT), increased neuronal activity and kainate receptor-mediated activation of intracellular protein kinase A (PKA) are important physiological and molecular steps for the generation of rapid eye movement (REM) sleep. In the present study performed on rats, phosphorylated cyclic AMP response element-binding protein (pCREB) immunostaining was used as a marker for increased intracellular PKA activation and as a reflection of increased neuronal activity. To identify whether activated cells were either cholinergic or noncholinergic, the PPT and laterodorsal tegmental nucleus (LDT) cells were immunostained for choline acetyltransferase (ChAT) in combination with pCREB or c-Fos. The results demonstrated that during high rapid eye movement sleep (HR, approximately 27%), significantly higher numbers of cells expressed pCREB and c-Fos in the PPT, of which 95% of pCREB-expressing cells were ChAT-positive. With HR, the numbers of pCREB-positive cells were also significantly higher in the medial pontine reticular formation (mPRF), pontine reticular nucleus oral (PnO), and dorsal subcoeruleus nucleus (SubCD) but very few in the locus coeruleus (LC) and dorsal raphe nucleus (DRN). Conversely, with low rapid eye movement sleep (LR, approximately 2%), the numbers of pCREB expressing cells were very few in the PPT, mPRF, PnO, and SubCD but significantly higher in the LC and DRN. The results of regression analyses revealed significant positive relationships between the total percentages of REM sleep and numbers of ChAT+/pCREB+ (Rsqr=0.98) cells in the PPT and pCREB+ cells in the mPRF (Rsqr=0.88), PnO (Rsqr=0.87), and SubCD (Rsqr=0.84); whereas significantly negative relationships were associated with the pCREB+ cells in the LC (Rsqr=0.70) and DRN (Rsqr=0.60). These results provide evidence supporting the hypothesis that during REM sleep, the PPT cholinergic neurons are active, whereas the LC and DRN neurons are inactive. More importantly, the regression analysis indicated that pCREB activation in approximately 98% of PPT cholinergic neurons, was caused by REM sleep. Moreover the results indicate that during REM sleep, PPT intracellular PKA activation and a transcriptional cascade involving pCREB occur exclusively in the cholinergic neurons.

Hepatic CREB3L3 controls whole-body energy homeostasis and improves obesity and diabetes.

PubMed

Nakagawa, Yoshimi; Satoh, Aoi; Yabe, Sachiko; Furusawa, Mika; Tokushige, Naoko; Tezuka, Hitomi; Mikami, Motoki; Iwata, Wakiko; Shingyouchi, Akiko; Matsuzaka, Takashi; Kiwata, Shiori; Fujimoto, Yuri; Shimizu, Hidehisa; Danno, Hirosuke; Yamamoto, Takashi; Ishii, Kiyoaki; Karasawa, Tadayoshi; Takeuchi, Yoshinori; Iwasaki, Hitoshi; Shimada, Masako; Kawakami, Yasushi; Urayama, Osamu; Sone, Hirohito; Takekoshi, Kazuhiro; Kobayashi, Kazuto; Yatoh, Shigeru; Takahashi, Akimitsu; Yahagi, Naoya; Suzuki, Hiroaki; Yamada, Nobuhiro; Shimano, Hitoshi

2014-12-01

Transcriptional regulation of metabolic genes in the liver is the key to maintaining systemic energy homeostasis during starvation. The membrane-bound transcription factor cAMP-responsive element-binding protein 3-like 3 (CREB3L3) has been reported to be activated during fasting and to regulate triglyceride metabolism. Here, we show that CREB3L3 confers a wide spectrum of metabolic responses to starvation in vivo. Adenoviral and transgenic overexpression of nuclear CREB3L3 induced systemic lipolysis, hepatic ketogenesis, and insulin sensitivity with increased energy expenditure, leading to marked reduction in body weight, plasma lipid levels, and glucose levels. CREB3L3 overexpression activated gene expression levels and plasma levels of antidiabetic hormones, including fibroblast growth factor 21 and IGF-binding protein 2. Amelioration of diabetes by hepatic activation of CREB3L3 was also observed in several types of diabetic obese mice. Nuclear CREB3L3 mutually activates the peroxisome proliferator-activated receptor (PPAR) α promoter in an autoloop fashion and is crucial for the ligand transactivation of PPARα by interacting with its transcriptional regulator, peroxisome proliferator-activated receptor gamma coactivator-1α. CREB3L3 directly and indirectly controls fibroblast growth factor 21 expression and its plasma level, which contributes at least partially to the catabolic effects of CREB3L3 on systemic energy homeostasis in the entire body. Therefore, CREB3L3 is a therapeutic target for obesity and diabetes.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Serk In, E-mail: serkin@korea.edu; The BK21 Plus Program for Biomedical Sciences, Korea University College of Medicine, Seoul; Department of Medicine and Center for Bone Biology, Vanderbilt University School of Medicine, Nashville, TN

The radiation stress induces cytotoxic responses of cell death as well as cytoprotective responses of cell survival. Understanding exact cellular mechanism and signal transduction pathways is important in improving cancer radiotherapy. Increasing evidence suggests that cyclic AMP response element binding protein (CREB)/activating transcription factor (ATF) family proteins act as a survival factor and a signaling molecule in response to stress. We postulated that CREB inhibition via CRE decoy oligonucleotide increases tumor cell sensitization to γ-irradiation-induced cytotoxic stress. In the present study, we demonstrate that CREB phosphorylation and CREB DNA-protein complex formation increased in time- and radiation dose-dependent manners, while theremore » was no significant change in total protein level of CREB. In addition, CREB was phosphorylated in response to γ-irradiation through p38 MAPK pathway. Further investigation revealed that CREB blockade by decoy oligonucleotides functionally inhibited transactivation of CREB, and significantly increased radiosensitivity of multiple human cancer cell lines including TP53- and/or RB-mutated cells with minimal effects on normal cells. We also demonstrate that tumor cells ectopically expressing dominant negative mutant CREB (KCREB) and the cells treated with p38 MAPK inhibitors were more sensitive to γ-irradiation than wild type parental cells or control-treated cells. Taken together, we conclude that CREB protects tumor cells from γ-irradiation, and combination of CREB inhibition plus ionizing radiation will be a promising radiotherapeutic approach. - Highlights: • γ-Irradiation induced CREB phosphorylation and CRE-directed transcription in tumor. • γ-Irradiation-induced transcriptional activation of CREB was via p38 MAPK pathway. • CRE blockade increased radiosensitivity of tumor cells but not of normal cells. • CRE decoy oligonucleotides or p38 MAPK inhibitors can be used as radiosensitizers.« less

CREB activity in dopamine D1 receptor expressing neurons regulates cocaine-induced behavioral effects

PubMed Central

Bilbao, Ainhoa; Rieker, Claus; Cannella, Nazzareno; Parlato, Rosanna; Golda, Slawomir; Piechota, Marcin; Korostynski, Michal; Engblom, David; Przewlocki, Ryszard; Schütz, Günther; Spanagel, Rainer; Parkitna, Jan R.

2014-01-01

It is suggested that striatal cAMP responsive element binding protein (CREB) regulates sensitivity to psychostimulants. To test the cell-specificity of this hypothesis we examined the effects of a dominant-negative CREB protein variant expressed in dopamine receptor D1 (D1R) neurons on cocaine-induced behaviors. A transgenic mouse strain was generated by pronuclear injection of a BAC-derived transgene harboring the A-CREB sequence under the control of the D1R gene promoter. Compared to wild-type, drug-naïve mutants showed moderate alterations in gene expression, especially a reduction in basal levels of activity-regulated transcripts such as Arc and Egr2. The behavioral responses to cocaine were elevated in mutant mice. Locomotor activity after acute treatment, psychomotor sensitization after intermittent drug injections and the conditioned locomotion after saline treatment were increased compared to wild-type littermates. Transgenic mice had significantly higher cocaine conditioned place preference, displayed normal extinction of the conditioned preference, but showed an augmented cocaine-seeking response following priming-induced reinstatement. This enhanced cocaine-seeking response was associated with increased levels of activity-regulated transcripts and prodynorphin. The primary reinforcing effects of cocaine were not altered in the mutant mice as they did not differ from wild-type in cocaine self-administration under a fixed ratio schedule at the training dose. Collectively, our data indicate that expression of a dominant-negative CREB variant exclusively in neurons expressing D1R is sufficient to recapitulate the previously reported behavioral phenotypes associated with virally expressed dominant-negative CREB. PMID:24966820

The transcription factor cyclic adenosine 3',5'-monophosphate response element-binding protein enhances the odonto/osteogenic differentiation of stem cells from the apical papilla.

PubMed

Su, S; Zhu, Y; Li, S; Liang, Y; Zhang, J

2017-09-01

To investigate the role of cAMP response element-binding protein (CREB) in the regulation of odonto/osteogenic differentiation of stem cells from the apical papilla (SCAPs). Stem cells from the apical papilla were obtained from human impacted third molars (n = 15). Isolated SCAPs were transfected with CREB overexpressing/silenced lentivirus. Transfected cells were stained with alizarin red to investigate mineralized nodule formation. The expression of the mineralization-related genes, alkaline phosphatase (ALP), collagen type I (Col I), runt-related transcription factor 2 (RUNX2), osterix (OSX) and osteocalcin (OCN), was determined by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Protein expression of the odontogenic-related marker dentine sialoprotein (DSP) and the osteogenic-related marker RUNX2 was measured by Western blotting analysis. One-way analysis of variance (anova) and Student's t-test were used for statistical analysis (a = 0.05). The overexpression of CREB enhanced mineralized nodule formation and up-regulated (P < 0.05) the mRNA levels of odonto/osteogenic-related markers, including ALP, Col I, RUNX2, OSX and OCN, and also increased (P < 0.05) the protein expression of DSP and RUNX2. In contrast, the silencing of CREB inhibited (P < 0.05) the mineralization capacity of the SCAPs and decreased (P < 0.05) the expression of odonto/osteogenic-related markers. Up-regulation of CREB expression promoted odonto/osteogenic differentiation of SCAPs and provided a potential method for the regeneration of the dentine-pulp complex. © 2016 International Endodontic Journal. Published by John Wiley & Sons Ltd.

Flow-induced protein kinase A–CREB pathway acts via BMP signaling to promote HSC emergence

PubMed Central

Kim, Peter Geon; Nakano, Haruko; Das, Partha P.; Chen, Michael J.; Rowe, R. Grant; Chou, Stephanie S.; Ross, Samantha J.; Sakamoto, Kathleen M.; Zon, Leonard I.; Schlaeger, Thorsten M.; Orkin, Stuart H.; Nakano, Atsushi

2015-01-01

Fluid shear stress promotes the emergence of hematopoietic stem cells (HSCs) in the aorta–gonad–mesonephros (AGM) of the developing mouse embryo. We determined that the AGM is enriched for expression of targets of protein kinase A (PKA)–cAMP response element-binding protein (CREB), a pathway activated by fluid shear stress. By analyzing CREB genomic occupancy from chromatin-immunoprecipitation sequencing (ChIP-seq) data, we identified the bone morphogenetic protein (BMP) pathway as a potential regulator of CREB. By chemical modulation of the PKA–CREB and BMP pathways in isolated AGM VE-cadherin+ cells from mid-gestation embryos, we demonstrate that PKA–CREB regulates hematopoietic engraftment and clonogenicity of hematopoietic progenitors, and is dependent on secreted BMP ligands through the type I BMP receptor. Finally, we observed blunting of this signaling axis using Ncx1-null embryos, which lack a heartbeat and intravascular flow. Collectively, we have identified a novel PKA–CREB–BMP signaling pathway downstream of shear stress that regulates HSC emergence in the AGM via the endothelial-to-hematopoietic transition. PMID:25870201

[Effects of predator stress on the expression of cAMP responsive element binding protein in hippocampus: experiment with rats].

PubMed

Wang, Qing-Song; Wu, Yu-Xian; Wang, Wei-Wen; Xiang, Yang

2007-12-18

To explore the neurobiological basis involved in the pathogenesis of the lasting emotionality and cognitive impairment following severe psychological stress. Ninety-six male Wistar rats were divided randomly into 2 equal groups: group of predator stress (Group PS) put into a cage in the experimental box singly to be exposed to a cat in the box but outside the cage for 23-57 min until tremor, polypnea, and nares flaring appeared for 6 min so as to establish predator stress models, and control group, put into the cage without non-injurious exposure of cat. 1, 12, and 24 hours later 8 rats from each group were killed with the hippocampus taken out. Western blotting was used to detect the protein expressions of cAMP response element-binding protein (CREB), phosphorylated CREB (pCREB) and CREB binding protein (CBP). Twelve hours after the experiment 24 rats from each group were killed with their brains taken out to obtain serial coronary sections. Immunohistochemistry was used to detect the positive immunoreactivities of CREB, pCREB, and CBP. Immunohistochemistry revealed that the absorbance (A) value of CREB-in the tissues of hippocampus and frontal cortex 12h after the cat exposure of Group PS were 0.55 +/- 0.13 and 0.88 +/- 0.20 respectively, both significantly lower than those of the control group (1.78 +/- 0.40 and 1.18 +/- 0.26 respectively, both P < 0.01), the A values of. pCREB in the hippocampus and frontal cortex of Group PS were 1.51 +/- 0.34 and 1.07 +/- 0.24 respectively, both significantly higher than those of the control group (0.47 +/- 0.11 and 0.48 +/- 0.11 respectively, both P < 0.01), and the A values of CBP in the hippocampus and frontal cortex of Group PS were 1.01 +/- 0.23 and 0.81 +/- 0.18 respectively, both significantly higher than those of the control group (0.52 +/- 0.12 and 0.29 +/- 0.07 respectively, both P < 0.01). Western blotting showed that the CREB protein expression levels 1 h and 24 h after the cat exposure of Group PS were 2.82 +/- 0.65 and 5.12 +/- 1.13 respectively, both significantly lower than those of the control group (11.86 +/- 2.47 and 10.56 +/- 2.38 respectively, both P < 0.01), the CBP protein expression levels 1 h and 24 h after the cat exposure of Group PS were 1.77 +/- 0.39 and 2.44 +/- 0.55 respectively, both significantly higher than those of the control group (1.06 +/- 0.24 and 0.86 +/- 0.20 respectively, both P < 0.01), and the pCREB protein expression levels 1 h and 12 h after the cat exposure of Group PS were 2.56 +/- 0.59 and 1.93 +/- 0.41 respectively, both significantly higher than those of the control group (1.04 +/- 0.22 and 0.96 +/- 0.21 respectively, both P < 0.01). The dysfunction of CREB signaling in the central nervous system, especially in the hippocampal formation after predation stress may play an important role in the long-term neuropsychological sequelae following severe stress.

Phospholipase D1 increases Bcl-2 expression during neuronal differentiation of rat neural stem cells.

PubMed

Park, Shin-Young; Ma, Weina; Yoon, Sung Nyo; Kang, Min Jeong; Han, Joong-Soo

2015-01-01

We studied the possible role of phospholipase D1 (PLD1) in the neuronal differentiation, including neurite formation of neural stem cells. PLD1 protein and PLD activity increased during neuronal differentiation. Bcl-2 also increased. Downregulation of PLD1 by transfection with PLD1 siRNA or a dominant-negative form of PLD1 (DN-PLD1) inhibited both neurite outgrowth and Bcl-2 expression. PLD activity was dramatically reduced by a PLCγ (phospholipase Cγ) inhibitor (U73122), a Ca(2+)chelator (BAPTA-AM), and a PKCα (protein kinase Cα) inhibitor (RO320432). Furthermore, treatment with arachidonic acid (AA) which is generated by the action of PLA2 (phospholipase A2) on phosphatidic acid (a PLD1 product), increased the phosphorylation of p38 MAPK and CREB, as well as Bcl-2 expression, indicating that PLA2 is involved in the differentiation process resulting from PLD1 activation. PGE2 (prostaglandin E2), a cyclooxygenase product of AA, also increased during neuronal differentiation. Moreover, treatment with PGE2 increased the phosphorylation of p38 MAPK and CREB, as well as Bcl-2 expression, and this effect was inhibited by a PKA inhibitor (Rp-cAMP). As expected, inhibition of p38 MAPK resulted in loss of CREB activity, and when CREB activity was blocked with CREB siRNA, Bcl-2 production also decreased. We also showed that the EP4 receptor was required for the PKA/p38MAPK/CREB/Bcl-2 pathway. Taken together, these observations indicate that PLD1 is activated by PLCγ/PKCα signaling and stimulate Bcl-2 expression through PLA2/Cox2/EP4/PKA/p38MAPK/CREB during neuronal differentiation of rat neural stem cells.

Type I human T cell leukemia virus tax protein transforms rat fibroblasts through the cyclic adenosine monophosphate response element binding protein/activating transcription factor pathway.

PubMed Central

Smith, M R; Greene, W C

1991-01-01

The Tax oncoprotein of the type I human T cell leukemia virus (HTLV-I) activates transcription of cellular and viral genes through at least two different transcription factor pathways. Tax activates transcription of the c-fos proto-oncogene by a mechanism that appears to involve members of the cAMP response element binding protein (CREB) and activating transcription factor (ATF) family of DNA-binding proteins. Tax also induces the nuclear expression of the NF-kappa B family of rel oncogene-related enhancer-binding proteins. We have investigated the potential role of these CREB/ATF and NF-kappa B/Rel transcription factors in Tax-mediated transformation by analyzing the oncogenic potential of Tax mutants that functionally segregate these two pathways of transactivation. Rat fibroblasts (Rat2) stably expressing either the wild-type Tax protein or a Tax mutant selectively deficient in the ability to induce NF-kappa B/Rel demonstrated marked changes in morphology and growth characteristics including the ability to form tumors in athymic mice. In contrast, Rat2 cells stably expressing a Tax mutant selectively deficient in the ability to activate transcription through CREB/ATF demonstrated no detectable changes in morphology or growth characteristics. These results suggest that transcriptional activation through the CREB/ATF pathway may play an important role in Tax-mediated cellular transformation. Images PMID:1832173

Fat mass and obesity-associated (FTO) protein interacts with CaMKII and modulates the activity of CREB signaling pathway.

PubMed

Lin, Li; Hales, Chadwick M; Garber, Kathryn; Jin, Peng

2014-06-15

Polymorphisms in the fat mass and obesity-associated (FTO) gene have been associated with obesity in humans. FTO is a nuclear protein and its physiological function remains largely unknown, but alterations in its expression in mice influence energy expenditure, food intake and, ultimately, body weight. To understand the molecular functions of FTO, we performed a yeast two-hybrid screen to identify the protein(s) that could directly interact with human FTO protein. Using multiple assays, we demonstrate that FTO interacts with three isoforms of calcium/calmodulin-dependent protein kinase II: α, β and γ, which are protein kinases that phosphorylate a broad range of substrates. This interaction is functional; overexpression of FTO delays the dephosphorylation of cAMP response element-binding protein (CREB) in human neuroblastoma (SK-N-SH) cells, which in turn leads to a dramatic increase in the expression of the CREB targets neuropeptide receptor 1 (NPY1R) and brain-derived neurotrophic factor (BDNF), which already are known to regulate food intake and energy homeostasis. Thus, our results suggest that FTO could modulate obesity by regulating the activity of the CREB signaling pathway. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

The CreB deubiquitinating enzyme does not directly target the CreA repressor protein in Aspergillus nidulans.

PubMed

Alam, Md Ashiqul; Kamlangdee, Niyom; Kelly, Joan M

2017-08-01

Ubiquitination/deubiquitination pathways are now recognized as key components of gene regulatory mechanisms in eukaryotes. The major transcriptional repressor for carbon catabolite repression in Aspergillus nidulans is CreA, and mutational analysis led to the suggestion that a regulatory ubiquitination/deubiquitination pathway is involved. A key unanswered question is if and how this pathway, comprising CreB (deubiquitinating enzyme) and HulA (ubiquitin ligase) and other proteins, is involved in the regulatory mechanism. Previously, missense alleles of creA and creB were analysed for genetic interactions, and here we extended this to complete loss-of-function alleles of creA and creB, and compared morphological and biochemical phenotypes, which confirmed genetic interaction between the genes. We investigated whether CreA, or a protein in a complex with it, is a direct target of the CreB deubiquitination enzyme, using co-purifications of CreA and CreB, first using strains that overexpress the proteins and then using strains that express the proteins from their native promoters. The Phos-tag system was used to show that CreA is a phosphorylated protein, but no ubiquitination was detected using anti-ubiquitin antibodies and Western analysis. These findings were confirmed using mass spectrometry, which confirmed that CreA was differentially phosphorylated but not ubiquitinated. Thus, CreA is not a direct target of CreB, and nor are proteins that form part of a stable complex with CreA a target of CreB. These results open up new questions regarding the molecular mechanism of CreA repressing activity, and how the ubiquitination pathway involving CreB interacts with this regulatory network.

Immunocytochemistry and fluorescence imaging efficiently identify individual neurons with CRISPR/Cas9-mediated gene disruption in primary cortical cultures.

PubMed

Tsunematsu, Hiroto; Uyeda, Akiko; Yamamoto, Nobuhiko; Sugo, Noriyuki

2017-08-01

CRISPR/Cas9 system is a powerful method to investigate the role of genes by introducing a mutation selectively and efficiently to specific genome positions in cell and animal lines. However, in primary neuron cultures, this method is affected by the issue that the effectiveness of CRISPR/Cas9 is different in each neuron. Here, we report an easy, quick and reliable method to identify mutants induced by the CRISPR/Cas9 system at a single neuron level, using immunocytochemistry (ICC) and fluorescence imaging. Dissociated cortical cells were transfected with CRISPR/Cas9 plasmids targeting the transcription factor cAMP-response element binding protein (CREB). Fluorescence ICC with CREB antibody and quantitative analysis of fluorescence intensity demonstrated that CREB expression disappeared in a fraction of the transfected neurons. The downstream FOS expression was also decreased in accordance with suppressed CREB expression. Moreover, dendritic arborization was decreased in the transfected neurons which lacked CREB immunoreactivity. Detection of protein expression is efficient to identify individual postmitotic neurons with CRISPR/Cas9-mediated gene disruption in primary cortical cultures. The present method composed of CRISPR/Cas9 system, ICC and fluorescence imaging is applicable to study the function of various genes at a single-neuron level.

Perturbing NR2B-PSD-95 interaction relieves neuropathic pain by inactivating CaMKII-CREB signaling.

PubMed

Xu, Fangxia; Zhao, Xin; Liu, Lin; Song, Jia; Zhu, Yingjun; Chu, Shuaishuai; Shao, Xueming; Li, Xiuxiu; Ma, Zhengliang; Gu, Xiaoping

2017-09-06

Neuropathic pain is characterized by central sensitization. The interaction between N-methyl-D-aspartate receptors (NMDARs) and postsynaptic density protein-95 (PSD-95) plays a major role in central sensitization. Here, we aimed to investigate the analgesic effect of disruption of the interaction between NMDAR and PSD-95. Chronic dorsal root ganglia compression model rats were used to mimic sciatica. Thermal hyperalgesia and mechanical allodynia were evaluated. The expression of spinal phospho-NR2B, PSD-95, calcium/calmodulin-dependent protein kinase II (CaMKII), and cAMP response element binding protein (CREB) was measured using western blotting. A mimetic peptide Myr-NR2B9c was injected intrathecally to disrupt the interaction between PSD-95 and NR2B and detected by coimmunoprecipitation. Chronic dorsal root ganglia compression surgery induced thermal hyperalgesia and mechanical allodynia, and upregulated pain-related proteins such as phospho-NR2B, PSD-95, CaMKII, and CREB expressions in the spinal cord. Myr-NR2B9c disrupted the interaction between NR2B-containing NMDARs and PSD-95 in the spinal cord. Intrathecal administration of Myr-NR2B9c attenuated neuropathic pain behaviors and downregulated the expressions of phospho-NR2B, PSD-95, CaMKII, and CREB in the spinal cord. The present study indicates that dissociation of NR2B-containing NMDARs from PSD-95 inactivates CaMKII and CREB signaling and relieves pain.

CREB3L1-mediated functional and structural adaptation of the secretory pathway in hormone-stimulated thyroid cells.

PubMed

García, Iris A; Torres Demichelis, Vanina; Viale, Diego L; Di Giusto, Pablo; Ezhova, Yulia; Polishchuk, Roman S; Sampieri, Luciana; Martinez, Hernán; Sztul, Elizabeth; Alvarez, Cecilia

2017-12-15

Many secretory cells increase the synthesis and secretion of cargo proteins in response to specific stimuli. How cells couple increased cargo load with a coordinate rise in secretory capacity to ensure efficient transport is not well understood. We used thyroid cells stimulated with thyrotropin (TSH) to demonstrate a coordinate increase in the production of thyroid-specific cargo proteins and ER-Golgi transport factors, and a parallel expansion of the Golgi complex. TSH also increased expression of the CREB3L1 transcription factor, which alone caused amplified transport factor levels and Golgi enlargement. Furthermore, CREB3L1 potentiated the TSH-induced increase in Golgi volume. A dominant-negative CREB3L1 construct hampered the ability of TSH to induce Golgi expansion, implying that this transcription factor contributes to Golgi expansion. Our findings support a model in which CREB3L1 acts as a downstream effector of TSH to regulate the expression of cargo proteins, and simultaneously increases the synthesis of transport factors and the expansion of the Golgi to synchronize the rise in cargo load with the amplified capacity of the secretory pathway. © 2017. Published by The Company of Biologists Ltd.

Monocyte 15-lipoxygenase gene expression requires ERK1/2 MAPK activity.

PubMed

Bhattacharjee, Ashish; Mulya, Anny; Pal, Srabani; Roy, Biswajit; Feldman, Gerald M; Cathcart, Martha K

2010-11-01

IL-13 induces profound expression of 15-lipoxygenase (15-LO) in primary human monocytes. Our studies have defined the functional IL-13R complex, association of Jaks with the receptor components, and the tyrosine phosphorylation of several Stat molecules in response to IL-13. Furthermore, we identified both p38MAPK and protein kinase Cδ as critical regulators of 15-LO expression. In this study, we report an ERK1/2-dependent signaling cascade that regulates IL-13-mediated 15-LO gene expression. We show the rapid phosphorylation/activation of ERK1/2 upon IL-13 exposure. Our results indicate that Tyk2 kinase is required for the activation of ERK1/2, which is independent of the Jak2, p38MAPK, and protein kinase Cδ pathways, suggesting bifurcating parallel regulatory pathways downstream of the receptor. To investigate the signaling mechanisms associated with the ERK1/2-dependent expression of 15-LO, we explored the involvement of transcription factors, with predicted binding sites in the 15-LO promoter, in this process including Elk1, early growth response-1 (Egr-1), and CREB. Our findings indicate that IL-13 induces Egr-1 nuclear accumulation and CREB serine phosphorylation and that both are markedly attenuated by inhibition of ERK1/2 activity. We further show that ERK1/2 activity is required for both Egr-1 and CREB DNA binding to their cognate sequences identified within the 15-LO promoter. Furthermore, by transfecting monocytes with the decoy oligodeoxyribonucleotides specific for Egr-1 and CREB, we discovered that Egr-1 and CREB are directly involved in regulating 15-LO gene expression. These studies characterize an important regulatory role for ERK1/2 in mediating IL-13-induced monocyte 15-LO expression via the transcription factors Egr-1 and CREB.

Berberine Suppresses Adipocyte Differentiation via Decreasing CREB Transcriptional Activity

PubMed Central

Deng, Ruyuan; Wang, Ning; Zhang, Yuqing; Wang, Yao; Liu, Yun; Li, Fengying; Wang, Xiao; Zhou, Libin

2015-01-01

Berberine, one of the major constituents of Chinese herb Rhizoma coptidis, has been demonstrated to lower blood glucose, blood lipid, and body weight in patients with type 2 diabetes mellitus. The anti-obesity effect of berberine has been attributed to its anti-adipogenic activity. However, the underlying molecular mechanism remains largely unknown. In the present study, we found that berberine significantly suppressed the expressions of CCAAT/enhancer-binding protein (C/EBP)α, peroxisome proliferators-activated receptor γ2 (PPARγ2), and other adipogenic genes in the process of adipogenesis. Berberine decreased cAMP-response element-binding protein (CREB) phosphorylation and C/EBPβ expression at the early stage of 3T3-L1 preadipocyte differentiation. In addition, CREB phosphorylation and C/EBPβ expression induced by 3-isobutyl-1-methylxanthine (IBMX) and forskolin were also attenuated by berberine. The binding activities of cAMP responsive element (CRE) stimulated by IBMX and forskolin were inhibited by berberine. The binding of phosphorylated CREB to the promoter of C/EBPβ was abrogated by berberine after the induction of preadipocyte differentiation. These results suggest that berberine blocks adipogenesis mainly via suppressing CREB activity, which leads to a decrease in C/EBPβ-triggered transcriptional cascades. PMID:25928058

CRE-Mediated Transcription and COX-2 Expression in the Pilocarpine Model of Status Epilepticus

PubMed Central

Lee, Boyoung; Dziema, Heather; Lee, Kyu Hyun; Choi, Yun-Sik; Obrietan, Karl

2007-01-01

Status epilepticus (SE) triggers neuronal death, reactive gliosis and remodeling of synaptic circuitry, thus leading to profound pathological alterations in CNS physiology. These processes are, in part, regulated by the rapid upregulation of both cytotoxic and cytoprotective genes. One pathway that may couple SE to transcriptionally-dependent alterations in CNS physiology is the CREB (cAMP response element-binding protein)/CRE (cAMP response element) cascade. Here, we utilized the pilocarpine model of SE on a mouse strain transgenic for a CRE-reporter construct (β-galactosidase) to begin to characterize how seizure activity regulates the activation state of the CREB/CRE pathway in both glia and neurons of the hippocampus. SE triggered a rapid (4–8 hrs post SE) but transient increase in CRE-mediated gene expression in the neuronal sublayers. In contrast to neurons, SE induced a lasting increase (up to 20 days) in CRE-mediated transcription in both reactive astrocytes and microglia. CRE-mediated gene expression correlated with expression of the pro-inflammatory enzyme cyclooxygenase-2 (COX-2). To examine the role of CREB in SE-induced COX-2 expression, we generated a transgenic mouse strain that expresses A-CREB, a potent repressor of CREB-dependent transcription. In these animals, the capacity of SE to stimulate COX-2 expression was markedly attenuated, indicating that CREB is a key intermediate in SE-induced COX-2 expression. Collectively these data show that SE triggers two waves of CREB-mediated gene expression, a transient wave in neurons and a long-lasting wave in reactive glial cells, and that CREB couples SE to COX-2 expression. PMID:17029965

Aripiprazole Increases the PKA Signalling and Expression of the GABAA Receptor and CREB1 in the Nucleus Accumbens of Rats.

PubMed

Pan, Bo; Lian, Jiamei; Huang, Xu-Feng; Deng, Chao

2016-05-01

The GABAA receptor is implicated in the pathophysiology of schizophrenia and regulated by PKA signalling. Current antipsychotics bind with D2-like receptors, but not the GABAA receptor. The cAMP-responsive element-binding protein 1 (CREB1) is also associated with PKA signalling and may be related to the positive symptoms of schizophrenia. This study investigated the effects of antipsychotics in modulating D2-mediated PKA signalling and its downstream GABAA receptors and CREB1. Rats were treated orally with aripiprazole (0.75 mg/kg, ter in die (t.i.d.)), bifeprunox (0.8 mg/kg, t.i.d.), haloperidol (0.1 mg/kg, t.i.d.) or vehicle for 1 week. The levels of PKA-Cα and p-PKA in the prefrontal cortex (PFC), nucleus accumbens (NAc) and caudate putamen (CPu) were detected by Western blots. The mRNA levels of Gabrb1, Gabrb2, Gabrb3 and Creb1, and their protein expression were measured by qRT-PCR and Western blots, respectively. Aripiprazole elevated the levels of p-PKA and the ratio of p-PKA/PKA in the NAc, but not the PFC and CPu. Correlated with this elevated PKA signalling, aripiprazole elevated the mRNA and protein expression of the GABAA (β-1) receptor and CREB1 in the NAc. While haloperidol elevated the levels of p-PKA and the ratio of p-PKA/PKA in both NAc and CPu, it only tended to increase the expression of the GABAA (β-1) receptor and CREB1 in the NAc, but not the CPu. Bifeprunox had no effects on PKA signalling in these brain regions. These results suggest that aripiprazole has selective effects on upregulating the GABAA (β-1) receptor and CREB1 in the NAc, probably via activating PKA signalling.

Molecular characterization of thyroid hormone-inhibited atrial L-type calcium channel expression: implication for atrial fibrillation in hyperthyroidism.

PubMed

Chen, Wei-Jan; Yeh, Yung-Hsin; Lin, Kwang-Huei; Chang, Gwo-Jyh; Kuo, Chi-Tai

2011-03-01

Atrial fibrillation (AF) is a common complication in hyperthyroidism. Earlier studies demonstrate that thyroid hormone decreases L-type calcium channel (LCC) current expression with resultant shortening of action potential duration (APD), providing a substrate for AF. The aim of this study was to investigate the potential mechanism underlying the regulatory effect of thyroid hormone on LCC. In a hyperthyroid rat model, thyroid hormone (triiodothyronine [T3]) administration down-regulated atrial LCC expression. In vitro, treatment of murine atrial myocytes (HL-1) with T3 decreased the expression of LCC and its current, resulting in abbreviation of APD. Furthermore, T3 inhibited the activation of cyclic AMP response element (CRE)-binding protein (CREB), including phosphorylation at Ser133 and its nuclear translocation. Transient transfection studies in HL-1 cells indicated that T3 reduced LCC promoter activity. Deletion and mutation analysis of the LCC promoter region along with chromatin immunoprecipitation using anti-CREB antibody showed that CRE was essential for T3-mediated LCC gene expression. Transfection of dominant-negative CREB (mutated Ser133) and mutant thyroid hormone receptor (TR, mutated Cys51) abolished the T3-dependent effects, suggesting an association between both transcriptional factors. Co-immunoprecipitation documented an increased binding of TR with CREB after T3 treatment. The transcriptional cross-talk 3 between TR and CREB bound to CRE mediates T3-inhibited CREB activity and LCC expression. Thyroid hormone-induced TR binding of CREB inhibits CREB activity and LCC current expression, which may contribute to AF. These findings provide an important mechanistic insight into hyperthyroidism-induced AF.

Glutaredoxin 1 (GRX1) inhibits oxidative stress and apoptosis of chondrocytes by regulating CREB/HO-1 in osteoarthritis.

PubMed

Sun, Jie; Wei, Xuelei; Lu, Yandong; Cui, Meng; Li, Fangguo; Lu, Jie; Liu, Yunjiao; Zhang, Xi

2017-10-01

GRX1 (glutaredoxin1), a sulfhydryl disulfide oxidoreductase, is involved in many cellular processes, including anti-oxidation, anti-apoptosis, and regulation of cell differentiation. However, the role of GRX1 in the oxidative stress and apoptosis of osteoarthritis chondrocytes remains unclear, prompting the current study. Protein and mRNA expressions were measured by Western blot and RT-qPCR. Oxidative stress was detected by the measurement of MDA and SOD contents. Cells apoptosis were detected by Annexin V-FITC/PI and caspase-3 activity assays. We found that the mRNA and protein expressions of GRX1 were significantly down-regulated in osteoarthritis tissues and cells. GRX1 overexpression increased the mRNA and protein expression of CREB and HO-1. Meanwhile, GRX1 overexpression inhibited oxidative stress and apoptosis in osteoarthritis chondrocytes. Furthermore, we found that GRX1 overexpression regulated HO-1 by increasing CREB, and that HO-1 regulated oxidative stress and apoptosis in osteoarthritis chondrocytes. Thus, GRX1 overexpression constrains oxidative stress and apoptosis in osteoarthritis chondrocytes by regulating CREB/HO-1, providing a novel insight into the molecular mechanism and potential treatment of osteoarthritis. Copyright © 2017. Published by Elsevier Ltd.

Aging impairs transcriptional regulation of vascular endothelial growth factor in human microvascular endothelial cells: implications for angiogenesis and cell survival.

PubMed

Ahluwalia, A; Jones, M K; Szabo, S; Tarnawski, A S

2014-04-01

In some tissues, aging impairs angiogenesis and reduces expression of vascular endothelial growth factor A (VEGF), a fundamental regulator of angiogenesis. We previously examined angiogenesis in aging and young gastric mucosa in vivo and in vitro and showed that an imbalance between expressions of VEGF (pro-angiogenic factor) and endostatin (anti-angiogenic protein) results in an aging-related impairment of angiogenesis in rats. However, the human relevance of these findings, and whether these mechanisms apply to endothelial cells derived from other tissues, is not clear. Since P-STAT3 and P-CREB are transcription factors that, in association with HIF-1α, can activate VEGF gene expression in some cells (e.g., liver cancer cells, vascular smooth muscle cells), we examined the expression of these two proteins in human dermal microvascular endothelial cells (HMVECs) derived from aging and neonatal individuals. We examined and quantified in vitro angiogenesis, expression of VEGF, P-STAT3, P-CREB and importin-α in HMVECs isolated from neonates (neonatal) and a 66 year old subject (aging). We also examined the effects of treatment with exogenous VEGF and endostatin on in vitro angiogenesis in these cells. Endothelial cells isolated from aging individuals had impaired angiogenesis (vs. neonatal endothelial cells) and reduced expression of VEGF mRNA and protein. Aged HMVECs also had reduced importin-α expression, and reduced expression and nuclear translocation of P-STAT3 and P-CREB. Reduced VEGF gene expression in aged HMVECs strongly correlated with the decreased levels of P-STAT3, P-CREB and importin-α in these cells. Our study clearly demonstrates that endothelial cells from aging individuals have impaired angiogenesis and reduced expression of VEGF likely due to impaired nuclear transport of P-STAT3 and P-CREB transcription factors in these cells.

Up-regulation of neurotrophic factors by cinnamon and its metabolite sodium benzoate: therapeutic implications for neurodegenerative disorders.

PubMed

Jana, Arundhati; Modi, Khushbu K; Roy, Avik; Anderson, John A; van Breemen, Richard B; Pahan, Kalipada

2013-06-01

This study underlines the importance of cinnamon, a widely-used food spice and flavoring material, and its metabolite sodium benzoate (NaB), a widely-used food preservative and a FDA-approved drug against urea cycle disorders in humans, in increasing the levels of neurotrophic factors [e.g., brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3)] in the CNS. NaB, but not sodium formate (NaFO), dose-dependently induced the expression of BDNF and NT-3 in primary human neurons and astrocytes. Interestingly, oral administration of ground cinnamon increased the level of NaB in serum and brain and upregulated the levels of these neurotrophic factors in vivo in mouse CNS. Accordingly, oral feeding of NaB, but not NaFO, also increased the level of these neurotrophic factors in vivo in the CNS of mice. NaB induced the activation of protein kinase A (PKA), but not protein kinase C (PKC), and H-89, an inhibitor of PKA, abrogated NaB-induced increase in neurotrophic factors. Furthermore, activation of cAMP response element binding (CREB) protein, but not NF-κB, by NaB, abrogation of NaB-induced expression of neurotrophic factors by siRNA knockdown of CREB and the recruitment of CREB and CREB-binding protein to the BDNF promoter by NaB suggest that NaB exerts its neurotrophic effect through the activation of CREB. Accordingly, cinnamon feeding also increased the activity of PKA and the level of phospho-CREB in vivo in the CNS. These results highlight a novel neutrophic property of cinnamon and its metabolite NaB via PKA - CREB pathway, which may be of benefit for various neurodegenerative disorders.

Resveratrol prevents cognitive deficits induced by chronic unpredictable mild stress: Sirt1/miR-134 signalling pathway regulates CREB/BDNF expression in hippocampus in vivo and in vitro.

PubMed

Shen, Jun; Xu, Linling; Qu, Chujie; Sun, Huimin; Zhang, Junjian

2018-04-30

Chronic unpredictable mild stress (CUMS) leads to neuropsychiatric disorders, such as depression, anxiety and cognitive impairment. Resveratrol is a natural polyphenol existed in polygonum cuspidatum and has been demonstrated to be a potent activator of Sirtuin 1 (Sirt1). Previous studies reported that resveratrol treatment ameliorated CUMS-induced depressive-like behavior and cognitive deficits through upregulating cAMP response element-binding protein (CREB) and brain derived neurotrophic factor (BDNF) expression. However, the upstream signalling pathway mediating CREB/BDNF expression and then exerting a protective role on cognitive function remains unclear. The present study aims to investigate the possible mechanism of resveratrol on CUMS-induced cognitive deficits. Male Sprague Dawley rats were adminstrated resveratrol (40 and 80 mg/kg) every day for 4 consecutive weeks before exposure to CUMS procedure. Morris Water Maze test was used to appraise spatial learing and memory of rats. Sirt1/miR-134 signalling pathway and CREB/BDNF expression in hippocampus of rats were measured. We also explored Sirt1/miR-134 signalling pathway and CREB/BDNF expression in primary cultured hippocampus neurons with resveratrol (25, 50 and 100 μmol/L) treatment. We found that resveratrol treatment prevented spatial learing and memory impairment induced by CUMS. Meanwhile the potential mechanism of resveratrol was associated with increased levels of Sirt1, CREB phosphorylation (p-CREB), CREB, BDNF and decreased levels of miR-134 in vivo and in vitro. In conclusion, our study showed that the neuroprotective effect of resveratrol on CUMS-induced cognitive impairment may rely on activating Sirt1/miR-134 pathway and then upregulating its downstream CREB/BDNF expression in hippocampus. Copyright © 2018 Elsevier B.V. All rights reserved.

17Beta-estradiol signaling and regulation of proliferation and apoptosis of rat Sertoli cells.

PubMed

Royer, Carine; Lucas, Thaís F G; Lazari, Maria F M; Porto, Catarina S

2012-04-01

The aim of the present study was to investigate the intracellular signaling events downstream of the classical estrogen receptors (ESRs) and G protein-coupled estrogen receptor 1 (GPER) involved in regulation of proliferation and apoptosis of rat Sertoli cells, in which we have previously described ESR1, ESR2, and GPER. ESRs play a role in Sertoli cell proliferation, and GPER, but not ESRs, plays a role modulating gene expression involved with apoptosis. The present study shows that 17beta-estradiol (E2) and the GPER-selective agonist G-1 rapidly activate phosphatidylinositol 3-kinase (PIK3)/serine threonine protein kinase (AKT) and cyclic AMP response element-binding (CREB) phosphorylation. E2 and the ESR1-selective agonist 4,4',4″-(4-propyl-(1H)-pyrazole-1,3,5-triyl)trisphenol (PPT) increase the expression of cyclin D1 (CCND1), whereas the ESR2-selective agonist 2,3-bis(4-hydroxyphenyl)-propionitrile (DPN) and G-1 do not change the expression of this protein, suggesting that ESR1 is the upstream receptor regulating Sertoli cell proliferation. E2- or PPT-ESR1, through activation of epidermal growth factor receptor (EGFR)/mitogen-activated protein kinase 3/1 (MAPK3/1) and PIK3 pathways, induces upregulation of CCND1. KG-501, the compound that disrupts the phospho-CREB/CREB binding protein (CBP) complex, does not change E2- or PPT-ESR1-mediated CCND1 expression, suggesting that phospho-CREB/cyclic AMP response element/CBP is not involved in the expression of this protein. E2- or G-1-GPER, through activation of EGFR/MAPK3/1 and PIK3 pathways, may be involved in the upregulation of antiapoptotic proteins BCL2 and BCL2L2. E2- or G-1-GPER/EGFR/MAPK3/1/phospho-CREB decreases BAX expression. Taken together, these results show a differential effect of E2-GPER on the CREB-mediated transcription of proapoptotic and antiapoptotic genes of the same BCL2 gene family. ESR1 and GPER can mediate the rapid E2 actions in the Sertoli cells, which in turn can modulate nuclear transcriptional events important for Sertoli cell function and maintenance of normal testis development and homeostasis. Our findings are important to clarify the role of estrogen in a critical period of testicular development, and to direct further studies, which may contribute to better understanding of the causes of male infertility.

Quercetin-3-O-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranoside suppresses melanin synthesis by augmenting p38 MAPK and CREB signaling pathways and subsequent cAMP down-regulation in murine melanoma cells

PubMed Central

Jung, Hyun Gug; Kim, Han Hyuk; Paul, Souren; Jang, Jae Yoon; Cho, Yong Hun; Kim, Hyeon Jeong; Yu, Jae Myo; Lee, Eun Su; An, Bong Jeun; Kang, Sun Chul; Bang, Byung Ho

2015-01-01

In this study, the effect of purified quercetin-3-O-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranosid (QCGG) on melanogenesis was investigated. QCGG was isolated from the calyx of a traditional Korean medicinal herb, Persimmon (Diospyros kaki). The hypopigmentation effects of QCGG were determined by examination of cellular melanin contents, tyrosinase activity assay, cAMP assay, and Western blotting of α-MSH-stimulated B16F10 mouse melanoma cells. Our results showed that QCGG inhibited both melanin synthesis and tyrosinase activity in a concentration-dependent manner as well as significantly reduced the expression of melanogenic proteins such as microphthalmia-associated transcription factor (MITF), tyrosinase-related protein-1, tyrosinase-related protein-2, and tyrosinase. Moreover, QCGG inhibited intracellular cAMP levels, cAMP response element-binding protein (CREB), and p38 MAPK expression in α-MSH-stimulated B16F10 cells. Taken together, the suppressive effects of QCGG on melanogenesis may involve down-regulation of MITF and its downstream signaling pathway via phosphorylation of p38 MAPK and CREB along with reduced cAMP levels. These results indicate that QCGG reduced melanin synthesis by reducing expression of tyrosine and tyrosine-related proteins via extracellular signal-related protein kinase (ERK) activation, followed by down-regulation of CREB, p38, and MITF. PMID:26586997

Quercetin-3-O-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranoside suppresses melanin synthesis by augmenting p38 MAPK and CREB signaling pathways and subsequent cAMP down-regulation in murine melanoma cells.

PubMed

Jung, Hyun Gug; Kim, Han Hyuk; Paul, Souren; Jang, Jae Yoon; Cho, Yong Hun; Kim, Hyeon Jeong; Yu, Jae Myo; Lee, Eun Su; An, Bong Jeun; Kang, Sun Chul; Bang, Byung Ho

2015-11-01

In this study, the effect of purified quercetin-3-O-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranosid (QCGG) on melanogenesis was investigated. QCGG was isolated from the calyx of a traditional Korean medicinal herb, Persimmon (Diospyros kaki). The hypopigmentation effects of QCGG were determined by examination of cellular melanin contents, tyrosinase activity assay, cAMP assay, and Western blotting of α-MSH-stimulated B16F10 mouse melanoma cells. Our results showed that QCGG inhibited both melanin synthesis and tyrosinase activity in a concentration-dependent manner as well as significantly reduced the expression of melanogenic proteins such as microphthalmia-associated transcription factor (MITF), tyrosinase-related protein-1, tyrosinase-related protein-2, and tyrosinase. Moreover, QCGG inhibited intracellular cAMP levels, cAMP response element-binding protein (CREB), and p38 MAPK expression in α-MSH-stimulated B16F10 cells. Taken together, the suppressive effects of QCGG on melanogenesis may involve down-regulation of MITF and its downstream signaling pathway via phosphorylation of p38 MAPK and CREB along with reduced cAMP levels. These results indicate that QCGG reduced melanin synthesis by reducing expression of tyrosine and tyrosine-related proteins via extracellular signal-related protein kinase (ERK) activation, followed by down-regulation of CREB, p38, and MITF.

Involvement of neuron-derived orphan receptor-1 (NOR-1) in LDL-induced mitogenic stimulus in vascular smooth muscle cells: role of CREB.

PubMed

Rius, Jordi; Martínez-González, José; Crespo, Javier; Badimon, Lina

2004-04-01

Low density lipoproteins (LDLs) modulate the expression of key genes involved in atherogenesis. Recently, we have shown that the transcription factor neuron-derived orphan receptor-1 (NOR-1) is involved in vascular smooth muscle cell (VSMC) proliferation. Our aim was to analyze whether NOR-1 is involved in LDL-induced mitogenic effects in VSMC. LDL induced NOR-1 expression in a time- and dose-dependent manner. Antisense oligonucleotides against NOR-1 inhibit DNA synthesis induced by LDL in VSMCs as efficiently as antisense against the protooncogene c-fos. The upregulation of NOR-1 mRNA levels by LDL involves pertusis-sensitive G protein-coupled receptors, Ca2+ mobilization, protein kinases A (PKA) and C (PKC) activation, and mitogen-activated protein kinase pathways (MAPK) (p44/p42 and p38). LDL promotes cAMP response element binding protein (CREB) activation (phosphorylation in Ser133). In transfection assays a dominant-negative of CREB inhibits NOR-1 promoter activity, while mutation of specific (cAMP response element) CRE sites in the NOR-1 promoter abolishes LDL-induced NOR-1 promoter activity. In VSMCs, LDL-induced mitogenesis involves NOR-1 upregulation through a CREB-dependent mechanism. CREB could play a role in the modulation by LDL of key genes (containing CRE sites) involved in atherogenesis.

Transgenic Mice Expressing a Truncated Form of CREB-Binding Protein (CBP) Exhibit Deficits in Hippocampal Synaptic Plasticity and Memory Storage

ERIC Educational Resources Information Center

Wood, Marcelo A.; Kaplan, Michael P.; Park, Alice; Blanchard, Edward J.; Oliveira, Ana M. M.; Lombardi, Thomas L.; Abel, Ted

2005-01-01

Deletions, translocations, or point mutations in the CREB-binding protein (CBP) gene have been associated with Rubinstein-Taybi Syndrome; a human developmental disorder characterized by retarded growth and reduced mental function. To examine the role of CBP in memory, transgenic mice were generated in which the CaMKII[alpha] promoter drives…

Molecular cellular mechanisms of peptide regulation of melatonin synthesis in pinealocyte culture.

PubMed

Khavinson, V Kh; Linkova, N S; Kvetnoy, I M; Kvetnaia, T V; Polyakova, V O; Korf, H-W

2012-06-01

The effects of epithalone and vilone peptides on the synthesis of melatonin and factors involved in this process, arylalkylamine-N-acetyltransferase (AANAT) enzyme and pCREB transcription protein, were studied in rat pinealocyte culture. Epithalone stimulated AANAT and pCREB synthesis and increased melatonin level in culture medium. Simultaneous addition of norepinephrine and peptides into the culture potentiated the expression of AANAT and pCREB.

Involvement of HTLV-I Tax and CREB in aneuploidy: a bioinformatics approach.

PubMed

de la Fuente, Cynthia; Gupta, Madhur V; Klase, Zachary; Strouss, Katharine; Cahan, Patrick; McCaffery, Timothy; Galante, Anthony; Soteropoulos, Patricia; Pumfery, Anne; Fujii, Masahiro; Kashanchi, Fatah

2006-07-05

Adult T-cell leukemia (ATL) is a complex and multifaceted disease associated with human T-cell leukemia virus type 1 (HTLV-I) infection. Tax, the viral oncoprotein, is considered a major contributor to cell cycle deregulation in HTLV-I transformed cells by either directly disrupting cellular factors (protein-protein interactions) or altering their transcription profile. Tax transactivates these cellular promoters by interacting with transcription factors such as CREB/ATF, NF-kappaB, and SRF. Therefore by examining which factors upregulate a particular set of promoters we may begin to understand how Tax orchestrates leukemia development. We observed that CTLL cells stably expressing wild-type Tax (CTLL/WT) exhibited aneuploidy as compared to a Tax clone deficient for CREB transactivation (CTLL/703). To better understand the contribution of Tax transactivation through the CREB/ATF pathway to the aneuploid phenotype, we performed microarray analysis comparing CTLL/WT to CTLL/703 cells. Promoter analysis of altered genes revealed that a subset of these genes contain CREB/ATF consensus sequences. While these genes had diverse functions, smaller subsets of genes were found to be involved in G2/M phase regulation, in particular kinetochore assembly. Furthermore, we confirmed the presence of CREB, Tax and RNA Polymerase II at the p97Vcp and Sgt1 promoters in vivo through chromatin immunoprecipitation in CTLL/WT cells. These results indicate that the development of aneuploidy in Tax-expressing cells may occur in response to an alteration in the transcription profile, in addition to direct protein interactions.

Long-term Administration of Salicylate-induced Changes in BDNF Expression and CREB Phosphorylation in the Auditory Cortex of Rats

PubMed Central

Yi, Bin; Wu, Cong; Shi, Runjie; Han, Kun; Sheng, Haibin; Li, Bei; Mei, Ling; Wang, Xueling; Huang, Zhiwu; Wu, Hao

2018-01-01

Hypothesis: We investigated whether salicylate induces tinnitus through alteration of the expression levels of brain-derived neurotrophic factor (BDNF), proBDNF, tyrosine kinase receptor B (TrkB), cAMP-responsive element-binding protein (CREB), and phosphorylated CREB (p-CREB) in the auditory cortex (AC). Background: Salicylate medication is frequently used for long-term treatment in clinical settings, but it may cause reversible tinnitus. Salicylate-induced tinnitus is associated with changes related to central auditory neuroplasticity. Our previous studies revealed enhanced neural activity and ultrastructural synaptic changes in the central auditory system after long-term salicylate administration. However, the underlying mechanisms remained unclear. Methods: Salicylate-induced tinnitus-like behavior in rats was confirmed using gap prepulse inhibition of acoustic startle and prepulse inhibition testing, followed by comparison of the expression levels of BDNF, proBDNF, TrkB, CREB, and p-CREB. Synaptic ultrastructure was observed under a transmission electron microscope. Results: BDNF and p-CREB were upregulated along with ultrastructural changes at the synapses in the AC of rats treated chronically with salicylate (p < 0.05, compared with control group). These changes returned to normal after 14 days of recovery (p > 0.05). Conclusion: Long-term administration of salicylate increased BDNF expression and CREB activation, upregulated synaptic efficacy, and changed synaptic ultrastructure in the AC. There may be a relationship between these factors and the mechanism of tinnitus. PMID:29342042

The minimal promoter region of the dense-core vesicle protein IA-2: transcriptional regulation by CREB.

PubMed

Cai, Tao; Hirai, Hiroki; Xu, Huanyu; Notkins, Abner L

2015-06-01

IA-2 is a transmembrane protein found in the dense-core vesicles (DCV) of neuroendocrine cells and one of the major autoantigens in type 1 diabetes. DCV are involved in the secretion of hormones (e.g., insulin) and neurotransmitters. Stimulation of pancreatic β cells with glucose upregulates the expression of IA-2 and an increase in IA-2 results in an increase in the number of DCV. Little is known, however, about the promoter region of IA-2 or the transcriptional factors that regulate the expression of this gene. In the present study, we constructed eight deletion fragments from the upstream region of the IA-2 transcription start site and linked them to a luciferase reporter. By this approach, we have identified a short bp region (-216 to +115) that has strong promoter activity. We also identified a transcription factor, cAMP responsive element-binding protein (CREB), which binds to two CREB-related binding sites located in this region. The binding of CREB to these sites enhanced IA-2 transcription by more than fivefold. We confirmed these findings by site-directed mutagenesis, chromatin immunoprecipitation assays and RNAi inhibition. Based on these findings, we conclude that the PKA pathway is a critical, but not the exclusive signaling pathway involved in IA-2 gene expression.

Hydroalcoholic extract of Rhodiola rosea L. (Crassulaceae) and its hydrolysate inhibit melanogenesis in B16F0 cells by regulating the CREB/MITF/tyrosinase pathway.

PubMed

Chiang, Hsiu-Mei; Chien, Yin-Chih; Wu, Chieh-Hsi; Kuo, Yueh-Hsiung; Wu, Wan-Chen; Pan, Yu-Yun; Su, Yu-Han; Wen, Kuo-Ching

2014-03-01

We investigated the effects of an aqueous alcohol extract of Rhodiola rosea (R. rosea) and its hydrolysate on melanin synthesis and the mechanisms mediating the activity. The ratio of tyrosol to salidroside was 2.3 in hydroalcoholic extract, and 51.0 in hydrolysate. We found that R. rosea extract and its hydrolysate inhibited melanin synthesis and tyrosinase activity in mouse melanoma cells (B16F0 cells). R. rosea extract also inhibited gene and protein expression of melanocortin 1 receptor (MC1R) and inhibited c-AMP response element binding protein (CREB) phosphorylation, suppressed the activation of AKT and glycogen synthase kinase-3 beta (GSK3β), and inhibited the expression of microphthalmia-associated transcription factor (MITF) and tyrosinase-related protein 1 (TRP-1). R. rosea hydrolysate inhibited the phosphorylation of CREB, the activation of AKT and GSK3β, and the expression of MITF and tyrosinase. Our results suggest that R. rosea extract is a novel tyrosinase inhibitor and that it exerts its effects by regulating the CREB/MITF/tyrosinase pathway in B16F0. Further in vivo studies are needed to determine the effectiveness of R. rosea extract as a skin whitening agent. Copyright © 2014 Elsevier Ltd. All rights reserved.

CREB1 regulates glucose transport of glioma cell line U87 by targeting GLUT1.

PubMed

Chen, Jiaying; Zhang, Can; Mi, Yang; Chen, Fuxue; Du, Dongshu

2017-12-01

Glioma is stemmed from the glial cells in the brain, which is accounted for about 45% of all intracranial tumors. The characteristic of glioma is invasive growth, as well as there is no obvious boundary between normal brain tissue and glioma tissue, so it is difficult to resect completely with worst prognosis. The metabolism of glioma is following the Warburg effect. Previous researches have shown that GLUT1, as a glucose transporter carrier, affected the Warburg effect, but the molecular mechanism is not very clear. CREB1 (cAMP responsive element-binding protein1) is involved in various biological processes, and relevant studies confirmed that CREB1 protein regulated the expression of GLUT1, thus mediating glucose transport in cells. Our experiments mainly reveal that the CREB1 could affect glucose transport in glioma cells by regulating the expression of GLUT1, which controlled the metabolism of glioma and affected the progression of glioma.

Monocyte 15-Lipoxygenase Gene Expression Requires ERK1/2 MAPK Activity

PubMed Central

Bhattacharjee, Ashish; Mulya, Anny; Pal, Srabani; Roy, Biswajit; Feldman, Gerald M.; Cathcart, Martha K.

2011-01-01

IL-13 induces profound expression of 15-lipoxygenase (15-LO) in primary human monocytes. Our studies have defined the functional IL-13R complex, association of Jaks with the receptor components, and the tyrosine phosphorylation of several Stat molecules in response to IL-13. Furthermore, we identified both p38MAPK and protein kinase Cδ as critical regulators of 15-LO expression. In this study, we report an ERK1/2-dependent signaling cascade that regulates IL-13–mediated 15-LO gene expression. We show the rapid phosphorylation/activation of ERK1/2 upon IL-13 exposure. Our results indicate that Tyk2 kinase is required for the activation of ERK1/2, which is independent of the Jak2, p38MAPK, and protein kinase Cδ pathways, suggesting bifurcating parallel regulatory pathways downstream of the receptor. To investigate the signaling mechanisms associated with the ERK1/2-dependent expression of 15-LO, we explored the involvement of transcription factors, with predicted binding sites in the 15-LO promoter, in this process including Elk1, early growth response-1 (Egr-1), and CREB. Our findings indicate that IL-13 induces Egr-1 nuclear accumulation and CREB serine phosphorylation and that both are markedly attenuated by inhibition of ERK1/2 activity. We further show that ERK1/2 activity is required for both Egr-1 and CREB DNA binding to their cognate sequences identified within the 15-LO promoter. Furthermore, by transfecting monocytes with the decoy oligodeoxyribonucleotides specific for Egr-1 and CREB, we discovered that Egr-1 and CREB are directly involved in regulating 15-LO gene expression. These studies characterize an important regulatory role for ERK1/2 in mediating IL-13–induced monocyte 15-LO expression via the transcription factors Egr-1 and CREB. PMID:20861348

CRTC2 activation in the suprachiasmatic nucleus, but not paraventricular nucleus, varies in a diurnal fashion and increases with nighttime light exposure.

PubMed

Highland, Julie A; Weiser, Michael J; Hinds, Laura R; Spencer, Robert L

2014-10-01

Entrainment of the intrinsic suprachiasmatic nucleus (SCN) molecular clock to the light-dark cycle depends on photic-driven intracellular signal transduction responses of SCN neurons that converge on cAMP response element-binding protein (CREB)-mediated regulation of gene transcription. Characterization of the CREB coactivator proteins CREB-regulated transcriptional coactivators (CRTCs) has revealed a greater degree of differential activity-dependent modulation of CREB transactivational function than previously appreciated. In confirmation of recent reports, we found an enrichment of crtc2 mRNA and prominent CRTC2 protein expression within the SCN of adult male rats. With use of a hypothalamic organotypic culture preparation for initial CRTC2-reactive antibody characterization, we found that CRTC2 immunoreactivity in hypothalamic neurons shifted from a predominantly cytoplasmic profile under basal culture conditions to a primarily nuclear localization (CRTC2 activation) 30 min after adenylate cyclase stimulation. In adult rat SCN, we found a diurnal variation in CRTC2 activation (peak at zeitgeber time of 4 h and trough at zeitgeber time of 16-20 h) but no variation in the total number of CRTC2-immunoreactive cells. There was no diurnal variation of CRTC2 activation in the hypothalamic paraventricular nucleus, another site of enriched CRTC2 expression. Exposure of rats to light (50 lux) for 30 min during the second half of their dark (nighttime) phase produced CRTC2 activation. We observed in the SCN a parallel change in the expression of a CREB-regulated gene (FOS). In contrast, nighttime light exposure had no effect on CRTC2 activation or FOS expression in the paraventricular nucleus, nor did it affect corticosterone hormone levels. These results suggest that CRTC2 participates in CREB-dependent photic entrainment of SCN function. Copyright © 2014 the American Physiological Society.

Chronic gonadotropin-releasing hormone inhibits activin induction of the ovine follicle-stimulating hormone beta-subunit: involvement of 3',5'-cyclic adenosine monophosphate response element binding protein and nitric oxide synthase type I.

PubMed

Shafiee-Kermani, Farideh; Han, Sang-oh; Miller, William L

2007-07-01

FSH is induced by activin, and this expression is modulated by GnRH through FSHB expression. This report focuses on the inhibitory effect of GnRH on activin-induced FSHB expression. Activin-treated primary murine pituitary cultures robustly express mutant ovine FSHBLuc-DeltaAP1, a luciferase transgene driven by 4.7 kb of ovine FSHB promoter. This promoter lacks two GnRH-inducible activator protein-1 sites, making it easier to observe GnRH-mediated inhibition. Luciferase expression from this transgene was decreased 94% by 100 nM GnRH with a half-time of approximately 4 h in pituitary cultures, and this inhibition was independent of follistatin. Activators of cAMP and protein kinase C like forskolin and phorbol 12-myristate 3-acetate (PMA), respectively, mimicked GnRH action. Kinetic studies of wild-type ovine FSHBLuc in LbetaT2 cells showed continuous induction by activin (4-fold) over 20 h. Most of this induction (78%) was blocked, beginning at 6 h. cAMP response element binding protein (CREB) was implicated in this inhibition because overexpression of its constitutively active mutant mimicked GnRH, and its inhibitor (inducible cAMP early repressor isoform II) reversed the inhibition caused by GnRH, forskolin, or PMA. In addition, GnRH, forskolin, or PMA increased the expression of a CREB-responsive reporter gene, 6xCRE-37PRL-Luc. Inhibition of nitric oxide type I (NOSI) by 7-nitroindazole also reversed GnRH-mediated inhibition by 60%. It is known that GnRH and CREB induce production of NOSI in gonadotropes and neuronal cells, respectively. These data support the concept that chronic GnRH inhibits activin-induced ovine FSHB expression by sequential activation of CREB and NOSI through the cAMP and/or protein kinase C pathways.

YC-1 potentiates cAMP-induced CREB activation and nitric oxide production in alveolar macrophages.

PubMed

Hwang, Tsong-Long; Tang, Ming-Chi; Kuo, Liang-Mou; Chang, Wen-De; Chung, Pei-Jen; Chang, Ya-Wen; Fang, Yao-Ching

2012-04-15

Alveolar macrophages play significant roles in the pathogenesis of several inflammatory lung diseases. Increases in exhaled nitric oxide (NO) are well documented to reflect disease severity in the airway. In this study, we investigated the effect of 3-(5'-hydroxymethyl-2'-furyl)-1-benzyl indazole (YC-1), a known activator of soluble guanylyl cyclase, on prostaglandin (PG)E₁ (a stable PGE₂ analogue) and forskolin (a adenylate cyclase activator) induced NO production and inducible NO synthase (iNOS) expression in rat alveolar macrophages (NR8383). YC-1 did not directly cause NO production or iNOS expression, but drastically potentiated PGE₁- or forskolin-induced NO production and iNOS expression in NR8383 alveolar macrophages. Combination treatment with YC-1 and PGE₁ significantly increased phosphorylation of the cAMP response element-binding protein (CREB), but not nuclear factor (NF)-κB activation. The combined effect on NO production, iNOS expression, and CREB phosphorylation was reversed by a protein kinase (PK)A inhibitor (H89), suggesting that the potentiating functions were mediated through a cAMP/PKA signaling pathway. Consistent with this, cAMP analogues, but not the cGMP analogue, caused NO release, iNOS expression, and CREB activation. YC-1 treatment induced an increase in PGE₁-induced cAMP formation, which occurred through the inhibition of cAMP-specific phosphodiesterase (PDE) activity. Furthermore, the combination of rolipram (an inhibitor of PDE4), but not milronone (an inhibitor of PDE3), and PGE₁ also triggered NO production and iNOS expression. In summary, YC-1 potentiates PGE₁-induced NO production and iNOS expression in alveolar macrophages through inhibition of cAMP PDE activity and activation of the cAMP/PKA/CREB signaling pathway. Copyright © 2012 Elsevier Inc. All rights reserved.

IL-1β Upregulates StAR and Progesterone Production Through the ERK1/2- and p38-Mediated CREB Signaling Pathways in Human Granulosa-Lutein Cells.

PubMed

Dang, Xuan; Zhu, Qinling; He, Yaqiong; Wang, Yuan; Lu, Yao; Li, Xiaoxue; Qi, Jia; Wu, Hasiximuke; Sun, Yun

2017-10-01

The proinflammatory cytokine interleukin-1β (IL-1β) may be involved in several ovulation-associated events, such as protease synthesis, prostaglandin production, and steroidogenesis in granulosa cells. However, the exact effect of IL-1β on progesterone synthesis in granulosa cells and the underlying mechanism remain unclear. By using cultured granulosa-lutein cells collected from women undergoing in vitro fertilization or intracytoplasmic sperm injection, we found that IL-1β upregulated steroidogenic acute regulatory protein (StAR) expression and progesterone synthesis in granulosa-lutein cells, which was comparable with luteinizing hormone effect and could be abolished by an IL-1 receptor antagonist. Moreover, IL-1β activated the phosphorylation of cyclic adenosine monophosphate response element-binding protein (CREB), and knockdown of CREB attenuated the induction of StAR expression and progesterone synthesis by IL-1β in granulosa-lutein cells. Furthermore, IL-1β activated the extracellular signal-regulated kinase (ERK)1/2 and p38 pathways and inhibition of the ERK1/2 and p38 pathways attenuated the IL-1β-induced phosphorylation of CREB, StAR expression, and progesterone synthesis in granulosa-lutein cells. In conclusion, IL-1β could upregulate StAR expression and stimulate progesterone biosynthesis through increase in CREB phosphorylation via activating the ERK1/2 and p38 pathways in human granulosa-lutein cells. Copyright © 2017 Endocrine Society.

Antinociceptive effects of oxymatrine from Sophora flavescens, through regulation of NR2B-containing NMDA receptor-ERK/CREB signaling in a mice model of neuropathic pain.

PubMed

Wang, Haiyan; Li, Yuxiang; Dun, Linglu; Xu, Yaqiong; Jin, Shaojv; Du, Juan; Ma, Lin; Li, Juan; Zhou, Ru; He, Xiaoliang; Sun, Tao; Yu, Jianqiang

2013-08-15

In this study we investigated antinociceptive effects of oxymatrine through regulation of NR2B-containing NMDA receptor-ERK/CREB signaling in a chronic neuropathic pain model induced by chronic constrictive injury (CCI) of the sciatic nerve. The von Frey and plantar tests were performed to assess the degree of mechanical and thermal changes respectively. Immunohistochemistry assay was used to evaluate the expressions of NR2B. Western blotting assay were used to evaluate the expressions of NR2B, tERK, p-ERK, tCREB and p-CREB. The intraperitoneal administration of OMT (160, 80 mg/kg) could prevent the development of mechanical allodynia, thermal hyperalgesia induced by CCI. Intraperitoneal administration of OMT decreased the mean IOD of NR2B in the dorsal horn and expression of NR2B, p-ERK and p-CREB protein. Regulation of NMDA NR2B receptor-ERK/CREB signaling maybe the targets for the antinociceptive effects of OMT on a chronic neuropathic pain model induced by chronic constrictive injury of the sciatic nerve. Copyright © 2013 Elsevier GmbH. All rights reserved.

Stochasticity and bifurcations in a reduced model with interlinked positive and negative feedback loops of CREB1 and CREB2 stimulated by 5-HT.

PubMed

Hao, Lijie; Yang, Zhuoqin; Bi, Yuanhong

2016-04-01

The cyclic AMP (cAMP)-response element-binding protein (CREB) family of transcription factors is crucial in regulating gene expression required for long-term memory (LTM) formation. Upon exposure of sensory neurons to the neurotransmitter serotonin (5-HT), CREB1 is activated via activation of the protein kinase A (PKA) intracellular signaling pathways, and CREB2 as a transcriptional repressor is relieved possibly via phosphorylation of CREB2 by mitogen-activated protein kinase (MAPK). Song et al. [18] proposed a minimal model with only interlinked positive and negative feedback loops of transcriptional regulation by the activator CREB1 and the repressor CREB2. Without considering feedbacks between the CREB proteins, Pettigrew et al. [8] developed a computational model characterizing complex dynamics of biochemical pathways downstream of 5-HT receptors. In this work, to describe more simply the biochemical pathways and gene regulation underlying 5-HT-induced LTM, we add the important extracellular sensitizing stimulus 5-HT as well as the product Ap-uch into the Song's minimal model. We also strive to examine dynamical properties of the gene regulatory network under the changing concentration of the stimulus, [5-HT], cooperating with the varying positive feedback strength in inducing a high state of CREB1 for the establishment of long-term memory. Different dynamics including monostability, bistability and multistability due to coexistence of stable steady states and oscillations is investigated by means of codimension-2 bifurcation analysis. At the different positive feedback strengths, comparative analysis of deterministic and stochastic dynamics reveals that codimension-1 bifurcation with respect to [5-HT] as the parameter can predict diverse stochastic behaviors resulted from the finite number of molecules, and the number of CREB1 molecules more and more preferentially resides near the high steady state with increasing [5-HT], which contributes to long-term memory formation. Copyright © 2016 Elsevier Inc. All rights reserved.

The antidepressant hyperforin increases the phosphorylation of CREB and the expression of TrkB in a tissue-specific manner.

PubMed

Gibon, Julien; Deloulme, Jean-Christophe; Chevallier, Tiphaine; Ladevèze, Elodie; Abrous, Djoher Nora; Bouron, Alexandre

2013-02-01

Hyperforin is one of the main bioactive compounds that underlie the antidepressant actions of the medicinal plant Hypericum perforatum (St. John's wort). However, the effects of a chronic hyperforin treatment on brain cells remains to be fully addressed. The following study was undertaken to further advance our understanding of the biological effects of this plant extract on neurons. Special attention was given to its impact on the brain-derived neurotrophic factor (BDNF) receptor TrkB and on adult hippocampal neurogenesis since they appear central to the mechanisms of action of antidepressants. The consequences of a chronic hyperforin treatment were investigated on cortical neurons in culture and on the brain of adult mice treated for 4 wk with a daily injection (i.p.) of hyperforin (4 mg/kg). Its effects on the expression of the cyclic adenosine monophosphate response element-binding protein (CREB), phospho-CREB (p-CREB), TrkB and phospho-TrkB (p-TrkB) were analysed by Western blot experiments and its impact on adult hippocampal neurogenesis was also investigated. Hyperforin stimulated the expression of TRPC6 channels and TrkB via SKF-96365-sensitive channels controlling a downstream signalling cascade involving Ca(2+), protein kinase A, CREB and p-CREB. In vivo, hyperforin augmented the expression of TrkB in the cortex but not in the hippocampus where hippocampal neurogenesis remained unchanged. In conclusion, this plant extract acts on the cortical BDNF/TrkB pathway leaving adult hippocampal neurogenesis unaffected. This study provides new insights on the neuronal responses controlled by hyperforin. We propose that the cortex is an important brain structure targeted by hyperforin.

Creb1 regulates late stage mammalian lung development via respiratory epithelial and mesenchymal-independent mechanisms

PubMed Central

Antony, N.; McDougall, A. R.; Mantamadiotis, T.; Cole, T. J.; Bird, A. D.

2016-01-01

During mammalian lung development, the morphological transition from respiratory tree branching morphogenesis to a predominantly saccular architecture, capable of air-breathing at birth, is dependent on physical forces as well as molecular signaling by a range of transcription factors including the cAMP response element binding protein 1 (Creb1). Creb1−/− mutant mice exhibit complete neonatal lethality consistent with a lack of lung maturation beyond the branching phase. To further define its role in the developing mouse lung, we deleted Creb1 separately in the respiratory epithelium and mesenchyme. Surprisingly, we found no evidence of a morphological lung defect nor compromised neonatal survival in either conditional Creb1 mutant. Interestingly however, loss of mesenchymal Creb1 on a genetic background lacking the related Crem protein showed normal lung development but poor neonatal survival. To investigate the underlying requirement for Creb1 for normal lung development, Creb1−/− mice were re-examined for defects in both respiratory muscles and glucocorticoid hormone signaling, which are also required for late stage lung maturation. However, these systems appeared normal in Creb1−/− mice. Together our results suggest that the requirement of Creb1 for normal mammalian lung morphogenesis is not dependent upon its expression in lung epithelium or mesenchyme, nor its role in musculoskeletal development. PMID:27150575

CREB and the CRTC co-activators: sensors for hormonal and metabolic signals

PubMed Central

Altarejos, Judith Y.; Montminy, Marc

2014-01-01

The cyclic AMP-responsive element-binding protein (CREB) is phosphorylated in response to a wide variety of signals, yet target gene transcription is only increased in a subset of cases. Recent studies indicate that CREB functions in concert with a family of latent cytoplasmic co-activators called cAMP-regulated transcriptional co-activators (CRTCs), which are activated through dephosphorylation. A dual requirement for CREB phosphorylation and CRTC dephosphorylation is likely to explain how these activator–co-activator cognates discriminate between different stimuli. Following their activation, CREB and CRTCs mediate the effects of fasting and feeding signals on the expression of metabolic programmes in insulin-sensitive tissues. PMID:21346730

Angelica sinensis polysaccharides promotes apoptosis in human breast cancer cells via CREB-regulated caspase-3 activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhou, Wei-Jie; Wang, Sheng; Hu, Zhuang, E-mail: zhuanghu475000@sina.com

Angelica sinensis polysaccharide (ASP) is purified from the fresh roots of Angelica sinensis (AS). This traditional Chinese medicine has been used for thousands of years for treating gynecological diseases and used in functional foods for the prevention and treatment of various diseases, such as inflammation and cancer. The antitumor activity of ASP is related to its biological activities, because it suppresses a variety of pro-proliferative or anti-apoptotic factors that are dramatically expressed in cancer cells of given types. In this study, we show that angelica sinensis polysaccharide induced apoptosis in breast cancer cells of T47D over-expressing the Cyclic AMP responsemore » element binding protein (CREB), inducing apoptosis-related signaling pathway activity. The result also found that ASP caused cell death was linked to caspase activity, accompanied by the loss of mitochondrial membrane potential, cytochrome c release, and Bax translocation from the cytosol to the mitochondria. We found that ASP significantly affected the poly-ADP-ribose polymerase (PARP), Bcl-2 Associated X Protein (Bax), Bcl-2, Bcl-xL and apoptotic protease activating facter-1 (Apaf1) protein expression in a dose- and time-dependent manner. DAPI staining and Flow cytometry were used to analyze apoptosis. The nude mice xenograft model was used to evaluate the antitumor effect of ASP in vivo. ASP has profound antitumor effect on T47D cells, probably by inducing apoptosis through CREB signaling pathway. Thus, these results suggest that ASP would be a promising therapeutic agent for breast cancer. - Highlights: • CREB and Caspase-3 signaling pathways are involved in the ASP induced breast cancer cells apoptosis. • ROCK1/Mlc signaling pathway plays a critical role in this ASP-mediated apoptosis. • Angelica sinensis polysaccharide (ASP) affected the PARP, Bax, Bcl-2, Bcl-xL and Apaf1 protein expression. • The activation of CREB and ROCK1 promotes caspase-3 activation and apoptosis induced by ASP.« less

Glial interleukin-1β upregulates neuronal sodium channel 1.7 in trigeminal ganglion contributing to temporomandibular joint inflammatory hypernociception in rats.

PubMed

Zhang, Peng; Bi, Rui-Yun; Gan, Ye-Hua

2018-04-20

The proinflammatory cytokine interleukin-1β (IL-1β) drives pain by inducing the expression of inflammatory mediators; however, its ability to regulate sodium channel 1.7 (Nav1.7), a key driver of temporomandibular joint (TMJ) hypernociception, remains unknown. IL-1β induces cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2). We previously showed that PGE2 upregulated trigeminal ganglionic Nav1.7 expression. Satellite glial cells (SGCs) involve in inflammatory pain through glial cytokines. Therefore, we explored here in the trigeminal ganglion (TG) whether IL-1β upregulated Nav1.7 expression and whether the IL-1β located in the SGCs upregulated Nav1.7 expression in the neurons contributing to TMJ inflammatory hypernociception. We treated rat TG explants with IL-1β with or without inhibitors, including NS398 for COX-2, PF-04418948 for EP2, and H89 and PKI-(6-22)-amide for protein kinase A (PKA), or with adenylate cyclase agonist forskolin, and used real-time PCR, Western blot, and immunohistofluorescence to determine the expressions or locations of Nav1.7, COX-2, cAMP response element-binding protein (CREB) phosphorylation, and IL-1β. We used chromatin immunoprecipitation to examine CREB binding to the Nav1.7 promoter. Finally, we microinjected IL-1β into the TGs or injected complete Freund's adjuvant into TMJs with or without previous microinjection of fluorocitrate, an inhibitor of SGCs activation, into the TGs, and evaluated nociception and gene expressions. Differences between groups were examined by one-way analysis of variance (ANOVA) or independent samples t test. IL-1β upregulated Nav1.7 mRNA and protein expressions in the TG explants, whereas NS398, PF-04418948, H89, or PKI-(6-22)-amide could all block this upregulation, and forskolin could also upregulate Nav1.7 mRNA and protein expressions. IL-1β enhanced CREB binding to the Nav1.7 promoter. Microinjection of IL-1β into the TGs or TMJ inflammation both induced hypernociception of TMJ region and correspondingly upregulated COX-2, phospho-CREB, and Nav1.7 expressions in the TGs. Moreover, microinjection of fluorocitrate into the TGs completely blocked TMJ inflammation-induced activation of SGCs and the upregulation of IL-1β and COX-2 in the SGCs, and phospho-CREB and Nav1.7 in the neurons and alleviated inflammation-induced TMJ hypernociception. Glial IL-1β upregulated neuronal Nav1.7 expression via the crosstalk between signaling pathways of the glial IL-1β/COX-2/PGE2 and the neuronal EP2/PKA/CREB/Nav1.7 in TG contributing to TMJ inflammatory hypernociception.

DPPC regulates COX-2 expression in monocytes via phosphorylation of CREB

DOE Office of Scientific and Technical Information (OSTI.GOV)

Morris, R.H.K.; Tonks, A.J.; Jones, K.P.

2008-05-23

The major phospholipid in pulmonary surfactant dipalmitoyl phosphatidylcholine (DPPC) has been shown to modulate inflammatory responses. Using human monocytes, this study demonstrates that DPPC significantly increased PGE{sub 2} (P < 0.05) production by 2.5-fold when compared to untreated monocyte controls. Mechanistically, this effect was concomitant with an increase in COX-2 expression which was abrogated in the presence of a COX-2 inhibitor. The regulation of COX-2 expression was independent of NF-{kappa}B activity. Further, DPPC increased the phosphorylation of the cyclic AMP response element binding protein (CREB; an important nuclear transcription factor important in regulating COX-2 expression). In addition, we also showmore » that changing the fatty acid groups of PC (e.g. using L-{alpha}-phosphatidylcholine {beta}-arachidonoyl-{gamma}-palmitoyl (PAPC)) has a profound effect on the regulation of COX-2 expression and CREB activation. This study provides new evidence for the anti-inflammatory activity of DPPC and that this activity is at least in part mediated via CREB activation of COX-2.« less

[Role of cyclic adenosine monophosphate response element binding protein in ventricular pacing induced cardiac electrical remodeling in a canine model].

PubMed

Chen, Xuesi; Chen, Xingxing; Cheng, Junhua; Hong, Jun; Zheng, Cheng; Zhao, Jinglin; Li, Jin; Lin, Jiafeng

2015-04-01

This project is designed to explore the potential role of cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) in cardiac electrical remodeling induced by pacing at different ventricular positions in dogs. An animal model by implanting the pacemakers in beagles was established. According to the different pacing positions, the animals were divided into 4 groups:conditional control group (n=6), left ventricle pacing group (n=6), right ventricle pacing group (n=6) and bi-ventricle pacing group (n=6). Cardiac and electrical remodeling were observed by echocardiography, electrocardiogram and plasma BNP. Myocardial pathology and protein expression of extracellular regulated protein kinases1/2 (ERK1/2), P38 mitogen activated protein kinases (P38 MAPK) and CREB were examined at 4 weeks post pacing. Cardiac structure and plasma BNP level were similar among 4 groups (all P>0.05). Electrocardiogram derived Tp-Te interval was significantly prolonged post pacing (92±11, 91±10, and 79±13 ms vs. 60±12 ms), and the Tp-Te interval in bi-ventricle pacing group was shorter than in left or right ventricle pacing group (P < 0.05). Western blot results showed that the expression of p-ERK1/2 in left ventricular myocardium of left ventricle pacing group, right ventricular myocardium of right ventricle pacing group and bi-ventricular myocardium of bi-ventricle pacing group was 2.7±0.4, 2.4±0.2, 1.7±0.1 and 1.9±0.2, respectively, the expression of p-P38 MAPK was 1.9±0.3, 1.7±0.2, 0.8±0.1 and 1.1±0.1, respectively, and the expression of p-CREB was 2.1±0.2, 2.0±0.2, 2.7±0.4 and 2.6±0.3, respectively. The p-ERK1/2 and p-P38 MAPK expression of bi-ventricle pacing group was lower,but the p-CREB expression was higher compared to the other pacing groups (P < 0.05). Ventricular pacing could induce electrical remodeling evidenced by prolonged Tp-Te interval and increased phosphorylation of ERK1/2 and p38 MAPK and reduced phosphorylation of CREB. Compared with single ventricle pacing, bi-ventricle pacing could attenuate electrical remodeling in this model.

3',5'-cyclic adenosine monophosphate response element binding protein up-regulated cytochrome P450 lanosterol 14alpha-demethylase expression involved in follicle-stimulating hormone-induced mouse oocyte maturation.

PubMed

Ning, Gang; Ouyang, Hong; Wang, Songbo; Chen, Xiufen; Xu, Baoshan; Yang, Jiange; Zhang, Hua; Zhang, Meijia; Xia, Guoliang

2008-07-01

Cytochrome P450 lanosterol 14alpha-demethylase (CYP51) is a key enzyme in sterols and steroids biosynthesis that can induce meiotic resumption in mouse oocytes. The present study investigated the expression mechanism and function of CYP51 during FSH-induced mouse cumulus oocyte complexes (COCs) meiotic resumption. FSH increased cAMP-dependent protein kinase (PKA) RIIbeta level and induced cAMP response element-binding protein (CREB) phosphorylation and CYP51 expression in cumulus cells before oocyte meiotic resumption. Moreover, CYP51 and epidermal growth factor (EGF)-like factor [amphiregulin (AR)] expression were blocked by (2)-naphthol-AS-Ephosphate (KG-501) (a drug interrupting the formation of CREB functional complex). KG-501 and RS21607 (a specific inhibitor of CYP51 activity) inhibited oocyte meiotic resumption, which can be partially rescued by progesterone. These two inhibitors also inhibited FSH-induced MAPK phosphorylation. EGF could rescue the suppression by KG-501 but not RS21607. Furthermore, type II PKA analog pairs, N(6)-monobutyryl-cAMP plus 8-bromo-cAMP, increased PKA RIIbeta level and mimicked the action of FSH, including CREB phosphorylation, AR and CYP51 expression, MAPK activation, and oocyte maturation. All these data suggest that CYP51 plays a critical role in FSH-induced meiotic resumption of mouse oocytes. CYP51 and AR gene expression in cumulus cells are triggered by FSH via a type II PKA/CREB-dependent signal pathway. Our study also implicates that CYP51 activity in cumulus cells participates in EGF receptor signaling-regulated oocyte meiotic resumption.

Association of MMP7 -181A→G Promoter Polymorphism with Gastric Cancer Risk: INFLUENCE OF NICOTINE IN DIFFERENTIAL ALLELE-SPECIFIC TRANSCRIPTION VIA INCREASED PHOSPHORYLATION OF cAMP-RESPONSE ELEMENT-BINDING PROTEIN (CREB).

PubMed

Kesh, Kousik; Subramanian, Lakshmi; Ghosh, Nillu; Gupta, Vinayak; Gupta, Arnab; Bhattacharya, Samir; Mahapatra, Nitish R; Swarnakar, Snehasikta

2015-06-05

Elevated expression of matrix metalloproteinase7 (MMP7) has been demonstrated to play a pivotal role in cancer invasion. The -181A→G (rs11568818) polymorphism in the MMP7 promoter modulates gene expression and possibly affects cancer progression. Here, we evaluated the impact of -181A→G polymorphism on MMP7 promoter activity and its association with gastric cancer risk in eastern Indian case-control cohorts (n = 520). The GG genotype as compared with the AA genotype was predisposed (p = 0.02; odds ratio = 1.9, 95% confidence interval = 1.1-3.3) to gastric cancer risk. Stratification analysis showed that tobacco addiction enhanced gastric cancer risk in GG subjects when compared with AA subjects (p = 0.03, odds ratio = 2.46, and 95% confidence interval = 1.07-5.68). Meta-analysis revealed that tobacco enhanced the risk for cancer more markedly in AG and GG carriers. Activity and expression of MMP7 were significantly higher in GG than in AA carriers. In support, MMP7 promoter-reporter assays showed greater transcriptional activity toward A to G transition under basal/nicotine-induced/cAMP-response element-binding protein (CREB) overexpressed conditions in gastric adenocarcinoma cells. Moreover, nicotine (a major component of tobacco) treatment significantly up-regulated MMP7 expression due to enhanced CREB phosphorylation followed by its nuclear translocation in gastric adenocarcinoma cells. Furthermore, chromatin immunoprecipitation experiments revealed higher binding of phosphorylated CREB with the -181G than the -181A allele. Altogether, specific binding of phosphorylated CREB to the G allele-carrying promoter enhances MMP7 gene expression that is further augmented by nicotine due to increased CREB phosphorylation and thereby increases the risk for gastric cancer. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Transcription factor assisted loading and enhancer dynamics dictate the hepatic fasting response

PubMed Central

Goldstein, Ido; Baek, Songjoon; Presman, Diego M.; Paakinaho, Ville; Swinstead, Erin E.; Hager, Gordon L.

2017-01-01

Fasting elicits transcriptional programs in hepatocytes leading to glucose and ketone production. This transcriptional program is regulated by many transcription factors (TFs). To understand how this complex network regulates the metabolic response to fasting, we aimed at isolating the enhancers and TFs dictating it. Measuring chromatin accessibility revealed that fasting massively reorganizes liver chromatin, exposing numerous fasting-induced enhancers. By utilizing computational methods in combination with dissecting enhancer features and TF cistromes, we implicated four key TFs regulating the fasting response: glucocorticoid receptor (GR), cAMP responsive element binding protein 1 (CREB1), peroxisome proliferator activated receptor alpha (PPARA), and CCAAT/enhancer binding protein beta (CEBPB). These TFs regulate fuel production by two distinctly operating modules, each controlling a separate metabolic pathway. The gluconeogenic module operates through assisted loading, whereby GR doubles the number of sites occupied by CREB1 as well as enhances CREB1 binding intensity and increases accessibility of CREB1 binding sites. Importantly, this GR-assisted CREB1 binding was enhancer-selective and did not affect all CREB1-bound enhancers. Single-molecule tracking revealed that GR increases the number and DNA residence time of a portion of chromatin-bound CREB1 molecules. These events collectively result in rapid synergistic gene expression and higher hepatic glucose production. Conversely, the ketogenic module operates via a GR-induced TF cascade, whereby PPARA levels are increased following GR activation, facilitating gradual enhancer maturation next to PPARA target genes and delayed ketogenic gene expression. Our findings reveal a complex network of enhancers and TFs that dynamically cooperate to restore homeostasis upon fasting. PMID:28031249

Adipose Tissue CLK2 Promotes Energy Expenditure during High-Fat Diet Intermittent Fasting.

PubMed

Hatting, Maximilian; Rines, Amy K; Luo, Chi; Tabata, Mitsuhisa; Sharabi, Kfir; Hall, Jessica A; Verdeguer, Francisco; Trautwein, Christian; Puigserver, Pere

2017-02-07

A promising approach to treating obesity is to increase diet-induced thermogenesis in brown adipose tissue (BAT), but the regulation of this process remains unclear. Here we find that CDC-like kinase 2 (CLK2) is expressed in BAT and upregulated upon refeeding. Mice lacking CLK2 in adipose tissue exhibit exacerbated obesity and decreased energy expenditure during high-fat diet intermittent fasting. Additionally, tissue oxygen consumption and protein levels of UCP1 are reduced in CLK2-deficient BAT. Phosphorylation of CREB, a transcriptional activator of UCP1, is markedly decreased in BAT cells lacking CLK2 due to enhanced CREB dephosphorylation. Mechanistically, CREB dephosphorylation is rescued by the inhibition of PP2A, a phosphatase that targets CREB. Our results suggest that CLK2 is a regulatory component of diet-induced thermogenesis in BAT through increased CREB-dependent expression of UCP1. Copyright © 2017 Elsevier Inc. All rights reserved.

The requirement for enhanced CREB1 expression in consolidation of long-term synaptic facilitation and long-term excitability in sensory neurons of Aplysia

PubMed Central

Liu, Rong-Yu; Cleary, Leonard J.; Byrne, John H.

2011-01-01

Accumulating evidence suggests that the transcriptional activator CREB1 is important for serotonin (5-HT)-induced long-term facilitation (LTF) of the sensorimotor synapse in Aplysia. Moreover, creb1 is among the genes activated by CREB1, suggesting a role for this protein beyond the induction phase of LTF. The time course of the requirement for CREB1 synthesis in the consolidation of long-term facilitation was examined using RNA interference (RNAi) techniques in sensorimotor co-cultures. Injection of CREB1 small-interfering RNA (siRNA) immediately or 10 h after 5-HT treatment blocked LTF when measured at 24 h and 48 h after treatment. In contrast, CREB1 siRNA did not block LTF when injected 16 h after 5-HT treatment. These results demonstrate that creb1 expression must be sustained for a relatively long time in order to support the consolidation of LTF. In addition, LTF is also accompanied by a long-term increase in the excitability (LTE) of sensory neurons (SNs). Because LTE was observed in the isolated SN after 5-HT treatment, this long-term change was intrinsic to that element of the circuit. LTE was blocked when CREB1 siRNA was injected into isolated SNs immediately after 5-HT treatment. These data suggest that 5-HT-induced CREB1 synthesis is required for consolidation of both LTF and LTE. PMID:21543617

Ferulic Acid Administered at Various Time Points Protects against Cerebral Infarction by Activating p38 MAPK/p90RSK/CREB/Bcl-2 Anti-Apoptotic Signaling in the Subacute Phase of Cerebral Ischemia-Reperfusion Injury in Rats.

PubMed

Cheng, Chin-Yi; Tang, Nou-Ying; Kao, Shung-Te; Hsieh, Ching-Liang

2016-01-01

This study aimed to evaluate the effects of ferulic acid (FA) administered at various time points before or after 30 min of middle cerebral artery occlusion (MCAo) followed by 7 d of reperfusion and to examine the involvement of mitogen-activated protein kinase (MAPK) signaling pathways in the cortical penumbra. FA was intravenously administered to rats at a dose of 100 mg/kg 24 h before ischemia (B-FA), 2 h before ischemia (P-FA), immediately after ischemic insult (I-FA), 2 h after reperfusion (R-FA), or 24 h after reperfusion (D-FA). Our study results indicated that P-FA, I-FA, and R-FA effectively reduced cerebral infarct areas and neurological deficits. P-FA, I-FA, and R-FA significantly downregulated glial fibrillary acidic protein (GFAP), mitochondrial Bax, cytochrome c, and cleaved caspase-3 expression, and effectively restored the phospho-p38 MAPK (p-p38 MAPK)/p38 MAPK ratio, phospho-90 kDa ribosomal S6 kinase (p-p90RSK) expression, phospho-Bad (p-Bad) expression, the phospho-cAMP response element-binding protein (p-CREB)/CREB ratio, the cytosolic and mitochondrial Bcl-2/Bax ratios, and the cytosolic Bcl-xL/Bax ratio in the cortical penumbra 7 d after reperfusion. SB203580, a specific inhibitor of p38 MAPK, administered 30 min prior to ischemia abrogated the downregulating effects of I-FA on cerebral infarction, and mitochondrial Bax and cleaved caspase-3 expression, and the upregulating effects of I-FA on the p-p38 MAPK/p38 MAPK ratio, p-p90RSK expression, p-Bad expression, and the p-CREB/CREB, and cytosolic and mitochondrial Bcl-2/Bax ratios. Our study results thus indicate that P-FA, I-FA, and R-FA effectively suppress reactive astrocytosis and exert neuroprotective effects against cerebral infarction by activating p38 MAPK signaling. The regulating effects of P-FA, I-FA, and R-FA on Bax-induced apoptosis result from activation of the p38 MAPK/p90RSK/CREB/Bcl-2 signaling pathway, and eventually contribute to inhibition of the cytochrome c-mediated caspase-3-dependent apoptotic pathway in the cortical penumbra 7 d after reperfusion.

Ferulic Acid Administered at Various Time Points Protects against Cerebral Infarction by Activating p38 MAPK/p90RSK/CREB/Bcl-2 Anti-Apoptotic Signaling in the Subacute Phase of Cerebral Ischemia-Reperfusion Injury in Rats

PubMed Central

Cheng, Chin-Yi; Tang, Nou-Ying; Kao, Shung-Te; Hsieh, Ching-Liang

2016-01-01

Objectives This study aimed to evaluate the effects of ferulic acid (FA) administered at various time points before or after 30 min of middle cerebral artery occlusion (MCAo) followed by 7 d of reperfusion and to examine the involvement of mitogen-activated protein kinase (MAPK) signaling pathways in the cortical penumbra. Methods FA was intravenously administered to rats at a dose of 100 mg/kg 24 h before ischemia (B-FA), 2 h before ischemia (P-FA), immediately after ischemic insult (I-FA), 2 h after reperfusion (R-FA), or 24 h after reperfusion (D-FA). Results Our study results indicated that P-FA, I-FA, and R-FA effectively reduced cerebral infarct areas and neurological deficits. P-FA, I-FA, and R-FA significantly downregulated glial fibrillary acidic protein (GFAP), mitochondrial Bax, cytochrome c, and cleaved caspase-3 expression, and effectively restored the phospho-p38 MAPK (p-p38 MAPK)/p38 MAPK ratio, phospho-90 kDa ribosomal S6 kinase (p-p90RSK) expression, phospho-Bad (p-Bad) expression, the phospho-cAMP response element-binding protein (p-CREB)/CREB ratio, the cytosolic and mitochondrial Bcl-2/Bax ratios, and the cytosolic Bcl-xL/Bax ratio in the cortical penumbra 7 d after reperfusion. SB203580, a specific inhibitor of p38 MAPK, administered 30 min prior to ischemia abrogated the downregulating effects of I-FA on cerebral infarction, and mitochondrial Bax and cleaved caspase-3 expression, and the upregulating effects of I-FA on the p-p38 MAPK/p38 MAPK ratio, p-p90RSK expression, p-Bad expression, and the p-CREB/CREB, and cytosolic and mitochondrial Bcl-2/Bax ratios. Conclusions Our study results thus indicate that P-FA, I-FA, and R-FA effectively suppress reactive astrocytosis and exert neuroprotective effects against cerebral infarction by activating p38 MAPK signaling. The regulating effects of P-FA, I-FA, and R-FA on Bax-induced apoptosis result from activation of the p38 MAPK/p90RSK/CREB/Bcl-2 signaling pathway, and eventually contribute to inhibition of the cytochrome c-mediated caspase-3-dependent apoptotic pathway in the cortical penumbra 7 d after reperfusion. PMID:27187745

Roles of p300 and cyclic adenosine monophosphate response element binding protein in high glucose-induced hypoxia-inducible factor 1α inactivation under hypoxic conditions.

PubMed

Ding, Lingtao; Yang, Minlie; Zhao, Tianlan; Lv, Guozhong

2017-05-01

Given the high prevalence of diabetes and burn injuries worldwide, it is essential to dissect the underlying mechanism of delayed burn wound healing in diabetes patients, especially the high glucose-induced hypoxia-inducible factor 1 (HIF-1)-mediated transcription defects. Human umbilical vein endothelial cells were cultured with low or high concentrations of glucose. HIF-1α-induced vascular endothelial growth factor (VEGF) transcription was measured by luciferase assay. Immunofluorescence staining was carried out to visualize cyclic adenosine monophosphate response element binding protein (CREB) localization. Immunoprecipitation was carried out to characterize the association between HIF-1α/p300/CREB. To test whether p300, CREB or p300+CREB co-overexpression was sufficient to rescue the HIF-1-mediated transcription defect after high glucose exposure, p300, CREB or p300+CREB co-overexpression were engineered, and VEGF expression was quantified. Finally, in vitro angiogenesis assay was carried out to test whether the high glucose-induced angiogenesis defect is rescuable by p300 and CREB co-overexpression. Chronic high glucose treatment resulted in impaired HIF-1-induced VEGF transcription and CREB exclusion from the nucleus. P300 or CREB overexpression alone cannot rescue high glucose-induced HIF-1α transcription defects. In contrast, co-overexpression of p300 and CREB dramatically ameliorated high glucose-induced impairment of HIF-1-mediated VEGF transcription, as well as in vitro angiogenesis. Finally, we showed that co-overexpression of p300 and CREB rectifies the dissociation of HIF-1α-p300-CREB protein complex in chronic high glucose-treated cells. Both p300 and CREB are required for the function integrity of HIF-1α transcription machinery and subsequent angiogenesis, suggesting future studies to improve burn wound healing might be directed to optimization of the interaction between p300, CREB and HIF-1α. © 2016 The Authors. Journal of Diabetes Investigation published by Asian Association for the Study of Diabetes (AASD) and John Wiley & Sons Australia, Ltd.

Neuron-derived orphan receptor 1 transduces survival signals in neuronal cells in response to hypoxia-induced apoptotic insults.

PubMed

Chio, Chung-Ching; Wei, Li; Chen, Tyng Guey; Lin, Chien-Min; Shieh, Ja-Ping; Yeh, Poh-Shiow; Chen, Ruei-Ming

2016-06-01

OBJECT Hypoxia can induce cell death or trigger adaptive mechanisms to guarantee cell survival. Neuron-derived orphan receptor 1 (NOR-1) works as an early-response protein in response to a variety of environmental stresses. In this study, the authors evaluated the roles of NOR-1 in hypoxia-induced neuronal insults. METHODS Neuro-2a cells were exposed to oxygen/glucose deprivation (OGD). Cell viability, cell morphology, cas-pase-3 activity, DNA fragmentation, and cell apoptosis were assayed to determine the mechanisms of OGD-induced neuronal insults. RNA and protein analyses were carried out to evaluate the effects of OGD on expressions of NOR-1, cAMP response element-binding (CREB), and cellular inhibitor of apoptosis protein 2 (cIAP2) genes. Translations of these gene expressions were knocked down using RNA interference. Mice subjected to traumatic brain injury (TBI) and NOR-1 was immunodetected. RESULTS Exposure of neuro-2a cells to OGD decreased cell viability in a time-dependent manner. Additionally, OGD led to cell shrinkage, DNA fragmentation, and cell apoptosis. In parallel, treatment of neuro-2a cells with OGD time dependently increased cellular NOR-1 mRNA and protein expressions. Interestingly, administration of TBI also augmented NOR-1 levels in the impacted regions of mice. As to the mechanism, exposure to OGD increased nuclear levels of the transcription factor CREB protein. Downregulating CREB expression using RNA interference simultaneously inhibited OGD-induced NOR-1 mRNA expression. Also, levels of cIAP2 mRNA and protein in neuro-2a cells were augmented by OGD. After reducing cIAP2 translation, OGD-induced cell death was reduced. Sequentially, application of NOR-1 small interfering RNA to neuro-2a cells significantly inhibited OGD-induced cIAP2 mRNA expression and concurrently alleviated hypoxia-induced alterations in cell viability, caspase-3 activation, DNA damage, and cell apoptosis. CONCLUSIONS This study shows that NOR-1 can transduce survival signals in neuronal cells responsible for hypoxiainduced apoptotic insults through activation of a CREB/cIAP2-dependent mechanism.

CREB- and NF-κB-Regulated CXC Chemokine Gene Expression in Lung Carcinogenesis

PubMed Central

Sun, Hongxia; Chung, Wen-Cheng; Ryu, Seung-Hee; Ju, Zhenlin; Tran, Hai T.; Kim, Edward; Kurie, Jonathan M.; Koo, Ja Seok

2009-01-01

The recognition of the importance of angiogenesis in tumor progression has led to the development of antiangiogenesis as a new strategy for cancer treatment and prevention. By modulating tumor microenvironment and inducing angiogenesis, the proinflammatory cytokine interleukine (IL)-1 β has been reported to promote tumor development. However, the factors mediating IL-1β-induced angiogenesis in non-small cell lung cancer (NSCLC) and the regulation of these angiogenic factors by IL-1β are less clear. Here, we report that IL-1β upregulated an array of proangiogenic CXC chemokine genes in NSCLC cell line A549 and in normal human tracheobronchial epithelium (NHTBE) cells, as determined by microarray analysis. Further analysis revealed that IL-1β induced much higher protein levels of CXC chemokines in NSCLC cells than in NHTBE cells. Conditioned medium from IL-1β treated A549 cells markedly increased endothelial cell migration, which was suppressed by neutralizing antibodies against CXCL5 and CXCR2. We also found that IL-1β-induced CXC chemokine gene overexpression in NSCLC cells was abrogated with the knockdown of CREB or NF-κB. Moreover, the expression of the CXC chemokine genes as well as CREB and NF-κB activities were greatly increased in tumorigenic NSCLC cell line compared with normal, premalignant immortalized or non-tumorigenic cell lines. A disruptor of the interaction between CREB-binding protein (CBP) and transcription factors such as CREB and NF-κB, 2-naphthol-AS-E-phosphate (KG-501), inhibited IL-1β-induced CXC chemokine gene expression and angiogenic activity in NSCLC. We propose that targeting CREB or NF-κB using small molecule inhibitors, such as KG-501, holds promise as a preventive and/or therapeutic approach for NSCLC. PMID:19138976

Clonidine preconditioning improved cerebral ischemia-induced learning and memory deficits in rats via ERK1/2-CREB/ NF-κB-NR2B pathway.

PubMed

Li, Yanli; Yu, Min; Zhao, Bo; Wang, Yan; Zha, Yunhong; Li, Zicheng; Yu, Lingling; Yan, Lingling; Chen, Zhangao; Zhang, Wenjuan; Zeng, Xiaoli; He, Zhi

2018-01-05

Clonidine, a classical α-2 adrenergic agonists, has been shown to antagonize brain damage caused by hypoxia, cerebral ischemia and excitotoxicity and reduce cerebral infarction volume in recent studies. We herein investigate the regulatory effect and possible underlying mechanism of clonidine on learning and memory in rats with cerebral ischemia. The cerebral ischemia rat model was established by right middle cerebral artery occlusion for 2h and reperfusion for 28 days. Drugs were administrated to the rats for consecutive 7 days intraperitoneally and once again on the day of surgery. The learning and memory in rats was assayed by Morris water maze. Moreover, protein expression levels of NMDAR2B (NR2B)/ phosphor - NR2B, ERK1/2/phosphor- ERK1/2, CREB/phosphor-CREB and NF-κB/phosphor-NF-κB in the cortex and hippocampus of the rats were assayed by western blotting. Our results demonstrated that clonidine treatment significantly abrogated the negative effect induced by cerebral ischemia on the learning and memory in the rats. In the Western blotting assay, clonidine treatment led to significant up-regulation of the expression level of NR2B and Phospho-NR2B in the hippocampus of the rats when compared with the cerebral ischemia group. Furthermore, clonidine also significantly decreased the protein expression levels of ERK1/2, Phospho-ERK1/2, CREB, Phospho-CREB and Phospho-NF-κB in the hippocampus of the rats when compared with the cerebral ischemia group. In conclusion, clonidine could improve the learning and memory ability of rats with cerebral ischemia, and NR2B, ERK1/2, CREB, NF-κB were involved in this effect. Copyright © 2017 Elsevier B.V. All rights reserved.

BDNF–ERK–CREB signalling mediates the role of miR-132 in the regulation of the effects of oleanolic acid in male mice

PubMed Central

Yi, Li-Tao; Li, Jing; Liu, Bin-Bin; Luo, Liu; Liu, Qing; Geng, Di

2014-01-01

Background Although previous study has demonstrated that brain-derived neurotrophic factor (BDNF) is involved in the antidepressant-like effect of oleanolic acid, there is little information regarding the details of the molecular mechanism involved in this effect. Methods We used a chronic unpredictable mild stress (CUMS) model to test the antidepressant-like effect of oleanolic acid on depressant-like behaviour, miR-132 expression and synaptic protein expression in the male mouse hippocampus. Furthermore, we explored the possible signalling pathways associated with miR-132 expression that mediate the effect of oleanolic acid on neuronal proliferation. Results The results demonstrated that a 3-week treatment with oleanolic acid ameliorated CUMS-induced anhedonic and anxiogenic behaviours. Furthermore, we found that oleanolic acid led to the BDNF-related phosphorylation and activation of extracellular signal-regulated kinases (ERK) and cyclic adenosine monophosphate response element binding protein (CREB), which was associated with the upregulation of miR-132 and hippocampal neuronal proliferation. Moreover, experiments with an miR-132 antagomir revealed that targeting miR-132 led to inhibition of neuronal proliferation and the postsynaptic density protein 95, but did not affect presynaptic protein synapsin I. Limitations Several other stimuli can also induce CREB phosphorylation in the hippocampus. Thus, regulation of miR-132 may not be restricted to neurotrophic signalling. Conclusion Our results show that oleanolic acid induces the upregulation of miR-132, which serves as an important regulator of neurotrophic actions, mainly through the activation of the hippocampal BDNF–ERK–CREB signalling pathways. PMID:25079084

Conformation of Tax-response elements in the human T-cell leukemia virus type I promoter.

PubMed

Cox, J M; Sloan, L S; Schepartz, A

1995-12-01

HTLV-I Tax is believed to activate viral gene expression by binding bZIP proteins (such as CREB) and increasing their affinities for proviral TRE target sites. Each 21 bp TRE target site contains an imperfect copy of the intrinsically bent CRE target site (the TRE core) surrounded by highly conserved flanking sequences. These flanking sequences are essential for maximal increases in DNA affinity and transactivation, but they are not, apparently, contacted by protein. Here we employ non-denaturing gel electrophoresis to evaluate TRE conformation in the presence and absence of bZIP proteins, and to explore the role of DNA conformation in viral transactivation. Our results show that the TRE-1 flanking sequences modulate the structure and modestly increase the affinity of a CREB bZIP peptide for the TRE-1 core recognition sequence. These flanking sequences are also essential for a maximal increase in stability of the CREB-DNA complex in the presence of Tax. The CRE-like TRE core and the TRE flanking sequences are both essential for formation of stable CREB-TRE-1 and Tax-CREB-TRE-1 complexes. These two DNA segments may have co-evolved into a unique structure capable of recognizing Tax and a bZIP protein.

Induction of Chemoresistance by All-Trans Retinoic Acid via a Noncanonical Signaling in Multiple Myeloma Cells

PubMed Central

Jiang, Kesheng; Huang, Qiaoli; Chen, Yicheng; Qian, Feng

2014-01-01

Despite the successful application of all-trans retinoic acid (ATRA) in multiple myeloma treatment, ATRA-induced chemoresistance in the myeloma patients is very common in clinic. In this study, we evaluated the effect of ATRA on the expression of apurinic endonuclease/redox factor-1 (Ape/Ref-1) in the U266 and RPMI-8226 myeloma cells to explore the chemoresistance mechanism involved. ATRA treatment induced upregulation of Ape/Ref-1 via a noncanonical signaling pathway, leading to enhanced pro-survival activity counteracting melphalan (an alkylating agent). ATRA rapidly activated p38-MSK (mitogen- and stress activated protein kinase) cascade to phosphorylate cAMP response element-binding protein (CREB). Phosphorylated CREB was recruited to the Ape/Ref-1 promoter to evoke the gene expression. The stimulation of ATRA on Ape/Ref-1 expression was attenuated by either p38-MSK inhibitors or overexpression of dominant-negative MSK1 mutants. Moreover, ATRA-mediated Ape/Ref-1 upregulation was correlated with histone modification and activation of CBP/p300, an important cofactors for CREB transcriptional activity. C646, a competitive CBP/p300 inhibitor, abolished the upregulation of Ape/Ref-1 induced by ATRA. Intriguingly, CBP rather than p300 played a dominant role in the expression of Ape/Ref-1. Hence, our study suggests the existence of a noncanonical mechanism involving p38-MSK-CREB cascade and CBP induction to mediate ATRA-induced Ape/Ref-1 expression and acquired chemoresistance in myeloma cells. PMID:24416428

Two CGTCA motifs and a GHF1/Pit1 binding site mediate cAMP-dependent protein kinase A regulation of human growth hormone gene expression in rat anterior pituitary GC cells.

PubMed

Shepard, A R; Zhang, W; Eberhardt, N L

1994-01-21

We established the cis-acting elements which mediate cAMP responsiveness of the human growth hormone (hGH) gene in transiently transfected rat anterior pituitary tumor GC cells. Analysis of the intact hGH gene or hGH 5'-flanking DNA (5'-FR) coupled to the hGh cDNA or chloramphenicol acetyltransferase or luciferase genes, indicated that cAMP primarily stimulated hGH promoter activity. Cotransfection of a protein kinase A inhibitory protein cDNA demonstrated that the cAMP response was mediated by protein kinase A. Mutational analysis of the hGH promoter identified two core cAMP response element motifs (CGTCA) located at nucleotides -187/-183 (distal cAMP response element; dCRE) and -99/-95 (proximal cAMP response element; pCRE) and a pituitary-specific transcription factor (GHF1/Pit1) binding site at nucleotides -123/-112 (dGHF1) which were required for cAMP responsiveness. GHF1 was not a limiting factor, since overexpression of GHF1 in cotransfections increased basal but not forskolin induction levels. Gel shift analyses indicated that similar, ubiquitous, thermostable protein(s) specifically bound the pCRE and dCRE motifs. The CGTCA motif-binding factors were cAMP response element binding protein (CREB)/activating transcription factor-1 (ATF-1)-related, since the DNA-protein complex was competed by unlabeled CREB consensus oligonucleotide, specifically supershifted by antisera to CREB and ATF-1 but not ATF-2, and was bound by purified CREB with the same relative binding affinity (pCRE < dCRE < CREB) and mobility as the GC nuclear extract. UV cross-linking and Southwestern blot analyses revealed multiple DNA-protein interactions of which approximately 100- and approximately 45-kDa proteins were predominant; the approximately 45-kDa protein may represent CREB. These results indicate that CREB/ATF-1-related factors act coordinately with the cell-specific factor GHF1 to mediate cAMP-dependent regulation of hGH-1 gene transcription in anterior pituitary somatotrophs.

CREB-binding protein controls response to cocaine by acetylating histones at the fosB promoter in the mouse striatum

PubMed Central

Levine, Amir A.; Guan, Zhonghui; Barco, Angel; Xu, Shiqin; Kandel, Eric R.; Schwartz, James H.

2005-01-01

Remodeling chromatin is essential for cAMP-regulated gene expression, necessary not only for development but also for memory storage and other enduring mental states. Histone acetylation and deacetylation mediate long-lasting forms of synaptic plasticity in Aplysia as well as cognition in mice. Here, we show that histone acetylation by the cAMP-response element binding protein (CREB)-binding protein (CBP) mediates sensitivity to cocaine by regulating expression of the fosB gene and its splice variant, ΔfosB, a transcription factor previously implicated in addiction. Using the chromatin immunoprecipitation assay with antibodies against histone H4 or CBP, we find that CBP is recruited to the fosB promoter to acetylate histone H4 in response to acute exposure to cocaine. We show that mutant mice that lack one allele of the CBP gene and have normal levels of fosB expression are less sensitive to chronic (10-day) administration of cocaine than are wild-type mice. This decreased sensitivity is correlated with decreased histone acetylation and results in decreased fosB expression and diminished accumulation of ΔfosB. Thus, CBP, which forms part of the promoter complex with CREB, mediates sensitivity to cocaine by acetylating histones. PMID:16380431

Curcumin reverses the effects of chronic stress on behavior, the HPA axis, BDNF expression and phosphorylation of CREB.

PubMed

Xu, Ying; Ku, Baoshan; Tie, Lu; Yao, Haiyan; Jiang, Wengao; Ma, Xing; Li, Xuejun

2006-11-29

Curcuma longa is a major constituent of the traditional Chinese medicine Xiaoyao-san, which has been used to effectively manage stress and depression-related disorders in China. Curcumin is the active component of curcuma longa, and its antidepressant effects were described in our prior studies in mouse models of behavioral despair. We hypothesized that curcumin may also alleviate stress-induced depressive-like behaviors and hypothalamic-pituitary-adrenal (HPA) axis dysfunction. Thus in present study we assessed whether curcumin treatment (2.5, 5 and 10 mg/kg, p.o.) affects behavior in a chronic unpredictable stress model of depression in rats and examined what its molecular targets may be. We found that subjecting animals to the chronic stress protocol for 20days resulted in performance deficits in the shuttle-box task and several physiological effects, such as an abnormal adrenal gland weight to body weight (AG/B) ratio and increased thickness of the adrenal cortex as well as elevated serum corticosterone levels and reduced glucocorticoid receptor (GR) mRNA expression. These changes were reversed by chronic curcumin administration (5 or 10 mg/kg, p.o.). In addition, we also found that the chronic stress procedure induced a down-regulation of brain-derived neurotrophic factor (BDNF) protein levels and reduced the ratio of phosphorylated cAMP response element-binding protein (pCREB) to CREB levels (pCREB/CREB) in the hippocampus and frontal cortex of stressed rats. Furthermore, these stress-induced decreases in BDNF and pCREB/CREB were also blocked by chronic curcumin administration (5 or 10 mg/kg, p.o.). These results provide compelling evidence that the behavioral effects of curcumin in chronically stressed animals, and by extension humans, may be related to their modulating effects on the HPA axis and neurotrophin factor expressions.

Elevated mu-opioid receptor expression in the nucleus of the solitary tract accompanies attenuated withdrawal signs after chronic low dose naltrexone in opiate-dependent rats.

PubMed

Van Bockstaele, E J; Rudoy, C; Mannelli, P; Oropeza, V; Qian, Y

2006-02-15

We previously described a decrease in withdrawal behaviors in opiate-dependent rats that were chronically treated with very low doses of naltrexone in their drinking water. Attenuated expression of withdrawal behaviors correlated with decreased c-Fos expression and intracellular signal transduction elements [protein kinase A regulatory subunit II (PKA) and phosphorylated cAMP response element binding protein (pCREB)] in brainstem noradrenergic nuclei. In this study, to determine whether similar cellular changes occurred in forebrain nuclei associated with drug reward, expressions of PKA and pCREB were analyzed in the ventral tegmental area, frontal cortex, striatum, and amygdala of opiate-treated rats that received low doses of naltrexone in their drinking water. No significant difference in PKA or pCREB was detected in these regions following drug treatment. To examine further the cellular mechanisms in noradrenergic nuclei that could underlie attenuated withdrawal behaviors following low dose naltrexone administration, the nucleus of the solitary tract (NTS) and locus coeruleus (LC) were examined for opioid receptor (OR) protein expression. Results showed a significant increase in muOR expression in the NTS of morphine-dependent rats that received low doses of naltrexone in their drinking water, and increases in muOR expression were also found to be dose dependent. Protein expression of muOR in the LC and deltaOR in either brain region remained unchanged. In conclusion, our previously reported decreases in c-Fos and PKA expression in the NTS following pretreatment with low doses of naltrexone may be partially explained by a greater inhibition of NTS neurons resulting from increased muOR expression in this region.

Erk-Creb pathway suppresses glutathione-S-transferase pi expression under basal and oxidative stress conditions in zebrafish embryos.

PubMed

Hrubik, Jelena; Glisic, Branka; Fa, Svetlana; Pogrmic-Majkic, Kristina; Andric, Nebojsa

2016-01-05

Transcriptional activation of phase II enzymes including glutathione-S-transferase pi class (Gst Pi) is important for redox regulation and defense from xenobiotics. The role of extracellular signal-regulated kinase (Erk) and protein kinase B (Akt) in regulation of Gst Pi expression has been described using adult mammalian cells. Whether these signaling pathways contribute to Gst Pi expression during embryogenesis is unknown. Using zebrafish embryo model, we provide novel evidence that Erk signaling acts as a specific suppressor of gstp1-2 mRNA during early embryogenesis. Addition of Erk inhibitor U0126 enhanced gstp1-2 mRNA expression during transition from blastula to the segmentation stage and from pharyngula until the hatching stage. Basal Erk activity did not affect gstp1-2 expression in tert-butylhydroquinone-exposed embryos. Addition of phorbol 12-myristate 13-acetate increased Erk activity leading to suppression of gstp1-2 mRNA. Activation of cAMP/Creb pathway by forskolin prevented gstp1-2 expression, whereas U0126 suppressed Creb phosphorylation, thus setting up Creb as a proximal transmitter of Erk inhibitory effect. Collectively, these findings suggest that Erk-Creb pathway exerts suppressive effect on gstp1-2 mRNA in a narrow developmental window. This study also provides a novel link between Erk and gstp1-2 expression, setting apart a possible differential regulation of gstp1-2 in adult and embryonic cells. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Dynorphin up-regulation in the dentate granule cell mossy fiber pathway following chronic inhibition of GluN2B-containing NMDAR is associated with increased CREB (Ser 133) phosphorylation, but is independent of BDNF/TrkB signaling pathways.

PubMed

Rittase, W Bradley; Dong, Yu; Barksdale, DaRel; Galdzicki, Zygmunt; Bausch, Suzanne B

2014-05-01

Emerging evidence suggests that neuronal responses to N-methyl-d-aspartate (NMDAR) activation/inactivation are influenced by subunit composition. For example, activation of synaptic NMDAR (comprised of GluN2A>GluN2B) phosphorylates cAMP-response-element-binding protein (CREB) at Ser 133, induces BDNF expression and promotes neuronal survival. Activation of extrasynaptic NMDAR (comprised of GluN2B>GluN2) dephosphorylates CREB (Ser 133), reduces BDNF expression and triggers neuronal death. These results led us to hypothesize that chronic inhibition of GluN2B-containing NMDAR would increase CREB (Ser 133) phosphorylation, increase BDNF levels and subsequently alter downstream dynorphin (DYN) and neuropeptide Y (NPY) expression. We focused on DYN and NPY because these neuropeptides can decrease excitatory neurotransmission and seizure occurrence and we reported previously that seizure-like events are reduced following chronic treatment with GluN2B antagonists. Consistent with our hypothesis, chronic treatment (17-21days) of hippocampal slice cultures with the GluN2B-selective antagonists ifenprodil or Ro25,6981 increased both CREB (Ser 133) phosphorylation and granule cell mossy fiber pathway DYN expression. Similar treatment with the non-subtype-selective NMDAR antagonists d-APV or memantine had no significant effect on either CREB (Ser 133) phosphorylation or DYN expression. In contrast to our hypothesis, BDNF levels were decreased following chronic treatment with Ro25,6981, but not ifenprodil, d-APV or memantine. Blockade of BDNF actions and TrkB activation did not significantly augment hilar DYN expression in vehicle-treated cultures and had no effect in Ro25,6981 treated cultures. These findings suggest that chronic exposure to GluN2B-selective NMDAR antagonists increased DYN expression through a putatively pCREB-dependent, but BDNF/TrkB-independent mechanism. Published by Elsevier Inc.

YC-1 potentiates cAMP-induced CREB activation and nitric oxide production in alveolar macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hwang, Tsong-Long, E-mail: htl@mail.cgu.edu.tw; Chinese Herbal Medicine Research Team, Healthy Aging Research Center, Chang Gung University, Kweishan, Taoyuan, Taiwan; Tang, Ming-Chi

2012-04-15

Alveolar macrophages play significant roles in the pathogenesis of several inflammatory lung diseases. Increases in exhaled nitric oxide (NO) are well documented to reflect disease severity in the airway. In this study, we investigated the effect of 3-(5′-hydroxymethyl-2′-furyl)-1-benzyl indazole (YC-1), a known activator of soluble guanylyl cyclase, on prostaglandin (PG)E{sub 1} (a stable PGE{sub 2} analogue) and forskolin (a adenylate cyclase activator) induced NO production and inducible NO synthase (iNOS) expression in rat alveolar macrophages (NR8383). YC-1 did not directly cause NO production or iNOS expression, but drastically potentiated PGE{sub 1}- or forskolin-induced NO production and iNOS expression in NR8383more » alveolar macrophages. Combination treatment with YC-1 and PGE{sub 1} significantly increased phosphorylation of the cAMP response element-binding protein (CREB), but not nuclear factor (NF)-κB activation. The combined effect on NO production, iNOS expression, and CREB phosphorylation was reversed by a protein kinase (PK)A inhibitor (H89), suggesting that the potentiating functions were mediated through a cAMP/PKA signaling pathway. Consistent with this, cAMP analogues, but not the cGMP analogue, caused NO release, iNOS expression, and CREB activation. YC-1 treatment induced an increase in PGE{sub 1}-induced cAMP formation, which occurred through the inhibition of cAMP-specific phosphodiesterase (PDE) activity. Furthermore, the combination of rolipram (an inhibitor of PDE4), but not milronone (an inhibitor of PDE3), and PGE{sub 1} also triggered NO production and iNOS expression. In summary, YC-1 potentiates PGE{sub 1}-induced NO production and iNOS expression in alveolar macrophages through inhibition of cAMP PDE activity and activation of the cAMP/PKA/CREB signaling pathway. Highlights: ► YC-1 potentiated PGE1-induced iNOS expression in alveolar macrophages. ► The combination of YC-1 and PGE1 increased CREB but not NFκB activation. ► The combined effects were reversed by H89. ► The combination of rolipram and PGE1 triggered NO production and iNOS expression. ► Effect of YC-1 occurred through inhibition of cAMP-specific PDE.« less

Differential contributions of Caenorhabditis elegans histone deacetylases to huntingtin polyglutamine toxicity.

PubMed

Bates, Emily A; Victor, Martin; Jones, Adriana K; Shi, Yang; Hart, Anne C

2006-03-08

Expansion of a polyglutamine tract in the huntingtin protein causes neuronal degeneration and death in Huntington's disease patients, but the molecular mechanisms underlying polyglutamine-mediated cell death remain unclear. Previous studies suggest that expanded polyglutamine tracts alter transcription by sequestering glutamine rich transcriptional regulatory proteins, thereby perturbing their function. We tested this hypothesis in Caenorhabditis elegans neurons expressing a human huntingtin fragment with an expanded polyglutamine tract (Htn-Q150). Loss of function alleles and RNA interference (RNAi) were used to examine contributions of C. elegans cAMP response element-binding protein (CREB), CREB binding protein (CBP), and histone deacetylases (HDACs) to polyglutamine-induced neurodegeneration. Deletion of CREB (crh-1) or loss of one copy of CBP (cbp-1) enhanced polyglutamine toxicity in C. elegans neurons. Loss of function alleles and RNAi were then used to systematically reduce function of each C. elegans HDAC. Generally, knockdown of individual C. elegans HDACs enhanced Htn-Q150 toxicity, but knockdown of C. elegans hda-3 suppressed toxicity. Neuronal expression of hda-3 restored Htn-Q150 toxicity and suggested that C. elegans HDAC3 (HDA-3) acts within neurons to promote degeneration in response to Htn-Q150. Genetic epistasis experiments suggested that HDA-3 and CRH-1 (C. elegans CREB homolog) directly oppose each other in regulating transcription of genes involved in polyglutamine toxicity. hda-3 loss of function failed to suppress increased neurodegeneration in hda-1/+;Htn-Q150 animals, indicating that HDA-1 and HDA-3 have different targets with opposing effects on polyglutamine toxicity. Our results suggest that polyglutamine expansions perturb transcription of CREB/CBP targets and that specific targeting of HDACs will be useful in reducing associated neurodegeneration.

Effects of lentivirus-mediated CREB expression in the dorsolateral striatum: memory enhancement and evidence for competitive and cooperative interactions with the hippocampus.

PubMed

Kathirvelu, Balachandar; Colombo, Paul J

2013-11-01

Neural systems specialized for memory may interact during memory formation or recall, and the results of interactions are important determinants of how systems control behavioral output. In two experiments, we used lentivirus-mediated expression of the transcription factor CREB (LV-CREB) to test if localized manipulations of cellular plasticity influence interactions between the hippocampus and dorsolateral striatum. In Experiment 1, we tested the hypothesis that infusion of LV-CREB in the dorsolateral striatum facilitates memory for response learning, and impairs memory for place learning. LV-CREB in the dorsolateral striatum had no effect on response learning, but impaired place memory; a finding consistent with competition between the striatum and hippocampus. In Experiment 2, we tested the hypothesis that infusion of LV-CREB in the dorsolateral striatum facilitates memory for cue learning, and impairs memory for contextual fear conditioning. LV-CREB in the dorsolateral striatum enhanced memory for cue learning and, in contrast to our prediction, also enhanced memory for contextual fear conditioning, consistent with a cooperative interaction between the striatum and hippocampus. Overall, the current experiments demonstrate that infusion of LV-CREB in the dorsolateral striatum (1) increases levels of CREB protein locally, (2) does not alter acquisition of place, response, cue, or contextual fear conditioning, (3) facilitates memory for cue learning and contextual fear conditioning, and (4) impairs memory for place learning. Taken together, the present results provide evidence that LV-CREB in the dorsolateral striatum can enhance memory formation and cause both competitive and cooperative interactions with the hippocampus. Copyright © 2013 Wiley Periodicals, Inc.

Transcription factor assisted loading and enhancer dynamics dictate the hepatic fasting response.

PubMed

Goldstein, Ido; Baek, Songjoon; Presman, Diego M; Paakinaho, Ville; Swinstead, Erin E; Hager, Gordon L

2017-03-01

Fasting elicits transcriptional programs in hepatocytes leading to glucose and ketone production. This transcriptional program is regulated by many transcription factors (TFs). To understand how this complex network regulates the metabolic response to fasting, we aimed at isolating the enhancers and TFs dictating it. Measuring chromatin accessibility revealed that fasting massively reorganizes liver chromatin, exposing numerous fasting-induced enhancers. By utilizing computational methods in combination with dissecting enhancer features and TF cistromes, we implicated four key TFs regulating the fasting response: glucocorticoid receptor (GR), cAMP responsive element binding protein 1 (CREB1), peroxisome proliferator activated receptor alpha (PPARA), and CCAAT/enhancer binding protein beta (CEBPB). These TFs regulate fuel production by two distinctly operating modules, each controlling a separate metabolic pathway. The gluconeogenic module operates through assisted loading, whereby GR doubles the number of sites occupied by CREB1 as well as enhances CREB1 binding intensity and increases accessibility of CREB1 binding sites. Importantly, this GR-assisted CREB1 binding was enhancer-selective and did not affect all CREB1-bound enhancers. Single-molecule tracking revealed that GR increases the number and DNA residence time of a portion of chromatin-bound CREB1 molecules. These events collectively result in rapid synergistic gene expression and higher hepatic glucose production. Conversely, the ketogenic module operates via a GR-induced TF cascade, whereby PPARA levels are increased following GR activation, facilitating gradual enhancer maturation next to PPARA target genes and delayed ketogenic gene expression. Our findings reveal a complex network of enhancers and TFs that dynamically cooperate to restore homeostasis upon fasting. Published by Cold Spring Harbor Laboratory Press.

GABA-CREB signalling regulates maturation and survival of newly generated neurons in the adult hippocampus

PubMed Central

Jagasia, Ravi; Steib, Kathrin; Englberger, Elisabeth; Herold, Sabine; Faus-Kessler, Theresa; Saxe, Michael; Gage, Fred H.; Song, Hongjun; Lie, D. Chichung

2009-01-01

Survival and integration of new neurons in the hippocampal circuit are rate-limiting steps in adult hippocampal neurogenesis. Neuronal network activity is a major regulator of these processes, yet little is known about the respective downstream signalling pathways. Here, we investigate the role of CREB signalling in adult hippocampal neurogenesis. CREB is activated in new granule neurons during a distinct developmental period. Loss of CREB function in a cell-autonomous fashion impairs dendritic development, decreases the expression of the neurogenic transcription factor NeuroD and of the neuronal microtubule associated protein, DCX, and compromises the survival of newborn neurons. In addition, GABA-mediated excitation regulates CREB activation at early developmental stages. Importantly, developmental defects following loss of GABA-mediated excitation can be compensated by enhanced CREB signalling. These results indicate that CREB signalling is a central pathway in adult hippocampal neurogenesis, regulating the development and survival of new hippocampal neurons downstream of GABA-mediated excitation. PMID:19553437

Suppressing cAMP response element-binding protein transcription shortens the duration of status epilepticus and decreases the number of spontaneous seizures in the pilocarpine model of epilepsy.

PubMed

Zhu, Xinjian; Dubey, Deepti; Bermudez, Camilo; Porter, Brenda E

2015-12-01

Current epilepsy therapies directed at altering the function of neurotransmitter receptors or ion channels, or release of synaptic vesicles fail to prevent seizures in approximately 30% of patients. A better understanding of the molecular mechanism underlying epilepsy is needed to provide new therapeutic targets. The activity of cyclic AMP (cAMP) response element-binding protein (CREB), a major transcription factor promoting CRE-mediated transcription, increases following a prolonged seizure called status epilepticus. It is also increased in the seizure focus of patients with medically intractable focal epilepsy. Herein we explored the effect of acute suppression of CREB activity on status epilepticus and spontaneous seizures in a chronic epilepsy model. Pilocarpine chemoconvulsant was used to induce status epilepticus. To suppress CREB activity, a transgenic mouse line expressing an inducible dominant negative mutant of CREB (CREB(IR) ) with a serine to alanine 133 substitution was used. Status epilepticus and spontaneous seizures of transgenic and wild-type mice were analyzed using video-electroencephalography (EEG) to assess the effect of CREB suppression on seizures. Our findings indicate that activation of CREB(IR) shortens the duration of status epilepticus. The frequency of spontaneous seizures decreased in mice with chronic epilepsy during CREB(IR) induction; however, the duration of the spontaneous seizures was unchanged. Of interest, we found significantly reduced levels of phospho-CREB Ser133 upon activation of CREB(IR) , supporting prior work suggesting that binding to the CRE site is important for CREB phosphorylation. Our results suggest that CRE transcription supports seizure activity both during status epilepticus and in spontaneous seizures. Thus, blocking of CRE transcription is a novel target for the treatment of epilepsy. Wiley Periodicals, Inc. © 2015 International League Against Epilepsy.

Resveratrol ameliorates spatial learning memory impairment induced by Aβ1-42 in rats.

PubMed

Wang, Rui; Zhang, Yu; Li, Jianguo; Zhang, Ce

2017-03-06

β-amyloid (Aβ) deposition is considered partially responsible for cognitive dysfunction in Alzheimer's disease (AD). Recently, resveratrol has been reported to play a potential role as a neuroprotective biofactor by modulating Aβ pathomechanisms, including through anti-neuronal apoptotic, anti-oxidative stress, and anti-neuroinflammatory effects. In addition, SIRT1 has been demonstrated to modulate learning and memory function by regulating the expression of cAMP response binding protein (CREB), which involves in modulating the expression of SIRT1. However, whether resveratrol can alleviate Aβ-induced cognitive dysfunction, whether SIRT1 expression and CREB phosphorylation in the hippocampus are affected by Aβ, and whether resveratrol influences these effects remain unknown. In the present study, we used a hippocampal injection model in rats to investigate the effects of resveratrol on Aβ 1-42 -induced impairment of spatial learning, memory and synaptic plasticity as well as on alterations of SIRT1 expression and CREB phosphorylation. We found that resveratrol significantly reversed the water maze behavioral impairment and the attenuation of long-term potentiation (LTP) in area CA1 that were induced by hippocampal injection of Aβ 1-42 . Interestingly, resveratrol also prevented the Aβ 1-42 -induced reductions in SIRT1 expression and CREB phosphorylation in rat hippocampus. In conclusion, in rats, resveratrol protects neurons against Aβ 1-42 -induced disruption of spatial learning, memory and hippocampal LTP. The mechanisms underlying the neuroprotective effects may involve rescue of SIRT1 expression and CREB phosphorylation. Copyright © 2016. Published by Elsevier Ltd.

The effect of glutamate on ghrelin release in mice.

PubMed

Chacrabati, Rakhi; Gong, Zhi; Ikenoya, Chika; Kondo, Daisuke; Zigman, Jeffrey M; Sakai, Takafumi; Sakata, Ichiro

2017-03-01

Ghrelin is abundantly produced in the stomach. Here, we found that glutamate decreased ghrelin expression and release in ghrelin-producing cells, and decreased levels of food intake and plasma acyl-ghrelin in mice. Treatment with siRNA of G protein-coupled receptor, family C, group 5, member B (GPRC5B) in ghrelin-producing cell lines completely blocked the effect of glutamate-induced ghrelin suppression. In addition, glutamate inhibited ghrelin release via the extracellular signal-regulated kinase (ERK) activity pathway, and stimulated CREB2 mRNA expression in ghrelin-producing cell lines. These results suggest that glutamate inhibits ghrelin release via ERK-CREB2 pathway. These results suggest that the GPRC5B-ERK-CREB2 pathway is involved in the inhibition of ghrelin expression and secretion in ghrelin cells. © 2017 International Federation for Cell Biology.

Cloning of Novel Isoforms of the Human Gli2 Oncogene and Their Activities To Enhance Tax-Dependent Transcription of the Human T-Cell Leukemia Virus Type 1 Genome

PubMed Central

Tanimura, Akira; Dan, Shingo; Yoshida, Mitsuaki

1998-01-01

The expression of human T-cell leukemia virus type 1 (HTLV-1) is activated by interaction of a viral transactivator protein, Tax, and cellular transcription factor, CREB (cyclic AMP response element binding protein), which bind to a 21-bp enhancer in the long terminal repeats (LTR). THP (Tax-helping protein) was previously determined to enhance the transactivation by Tax protein. Here we report novel forms of the human homolog of a member of the Gli oncogene family, Gli2 (also termed Gli2/THP), an extended form of a zinc finger protein, THP, which was described previously. Four possible isoforms (hGli2 α, β, γ, and δ) are formed by combinations of two independent alternative splicings, and all the isoforms could bind to a DNA motif, TRE2S, in the LTR. The longer isoforms, α and β, were abundantly expressed in various cell lines including HTLV-1-infected T-cell lines. Fusion proteins of the hGli2 isoforms with the DNA-binding domain of Gal4 activated transcription when the reporter contained a Gal4-binding site and one copy of the 21-bp sequence, to which CREB binds. This activation was observed only in the presence of Tax. The 21-bp sequence in the reporter was also essential for the activation. These results suggest that simultaneous binding of hGli2 and CREB to the respective sites in the reporter seems to be critical for Tax protein to activate transcription. Consequently, it is probable that the LTR can be regulated by two independent signals through hGli2 and CREB, since the LTR contains the 21-bp and TRE2S sequences in the vicinity. PMID:9557682

Ethanol exposure induces neonatal neurodegeneration by enhancing CB1R Exon1 histone H4K8 acetylation and up-regulating CB1R function causing neurobehavioral abnormalities in adult mice.

PubMed

Subbanna, Shivakumar; Nagre, Nagaraja N; Umapathy, Nagavedi S; Pace, Betty S; Basavarajappa, Balapal S

2014-10-31

Ethanol exposure to rodents during postnatal day 7 (P7), which is comparable to the third trimester of human pregnancy, induces long-term potentiation and memory deficits. However, the molecular mechanisms underlying these deficits are still poorly understood. In the present study, we explored the potential role of epigenetic changes at cannabinoid type 1 (CB1R) exon1 and additional CB1R functions, which could promote memory deficits in animal models of fetal alcohol spectrum disorder. We found that ethanol treatment of P7 mice enhances acetylation of H4 on lysine 8 (H4K8ace) at CB1R exon1, CB1R binding as well as the CB1R agonist-stimulated GTPγS binding in the hippocampus and neocortex, two brain regions that are vulnerable to ethanol at P7 and are important for memory formation and storage, respectively. We also found that ethanol inhibits cyclic adenosine monophosphate response element-binding protein (CREB) phosphorylation and activity-regulated cytoskeleton-associated protein (Arc) expression in neonatal and adult mice. The blockade or genetic deletion of CB1Rs prior to ethanol treatment at P7 rescued CREB phosphorylation and Arc expression. CB1R knockout mice exhibited neither ethanol-induced neurodegeneration nor inhibition of CREB phosphorylation or Arc expression. However, both neonatal and adult mice did exhibit enhanced CREB phosphorylation and Arc protein expression. P7 ethanol-treated adult mice exhibited impaired spatial and social recognition memory, which were prevented by the pharmacological blockade or deletion of CB1Rs at P7. Together, these findings suggest that P7 ethanol treatment induces CB1R expression through epigenetic modification of the CB1R gene, and that the enhanced CB1R function induces pCREB, Arc, spatial, and social memory deficits in adult mice. © The Author 2015. Published by Oxford University Press on behalf of CINP.

Human T-cell leukemia virus type 1 Tax requires direct access to DNA for recruitment of CREB binding protein to the viral promoter.

PubMed

Lenzmeier, B A; Giebler, H A; Nyborg, J K

1998-02-01

Efficient human T-cell leukemia virus type 1 (HTLV-1) replication and viral gene expression are dependent upon the virally encoded oncoprotein Tax. To activate HTLV-1 transcription, Tax interacts with the cellular DNA binding protein cyclic AMP-responsive element binding protein (CREB) and recruits the coactivator CREB binding protein (CBP), forming a nucleoprotein complex on the three viral cyclic AMP-responsive elements (CREs) in the HTLV-1 promoter. Short stretches of dG-dC-rich (GC-rich) DNA, immediately flanking each of the viral CREs, are essential for Tax recruitment of CBP in vitro and Tax transactivation in vivo. Although the importance of the viral CRE-flanking sequences is well established, several studies have failed to identify an interaction between Tax and the DNA. The mechanistic role of the viral CRE-flanking sequences has therefore remained enigmatic. In this study, we used high resolution methidiumpropyl-EDTA iron(II) footprinting to show that Tax extended the CREB footprint into the GC-rich DNA flanking sequences of the viral CRE. The Tax-CREB footprint was enhanced but not extended by the KIX domain of CBP, suggesting that the coactivator increased the stability of the nucleoprotein complex. Conversely, the footprint pattern of CREB on a cellular CRE lacking GC-rich flanking sequences did not change in the presence of Tax or Tax plus KIX. The minor-groove DNA binding drug chromomycin A3 bound to the GC-rich flanking sequences and inhibited the association of Tax and the Tax-CBP complex without affecting CREB binding. Tax specifically cross-linked to the viral CRE in the 5'-flanking sequence, and this cross-link was blocked by chromomycin A3. Together, these data support a model where Tax interacts directly with both CREB and the minor-groove viral CRE-flanking sequences to form a high-affinity binding site for the recruitment of CBP to the HTLV-1 promoter.

Exposure to bisphenol A (BPA) in Wistar rats reduces sperm quality with disruption of ERK signal pathway.

PubMed

Li, Juan; Mao, Rui; Zhou, Qin; Ding, Ling; Tao, Jin; Ran, Mao-Mei; Gao, Er-Sheng; Yuan, Wei; Wang, Jin-Tao; Hou, Li-Fang

2016-01-01

Bisphenol A (BPA) is an estrogenic environmental toxin widely used in the production of plastics and ubiquitous human exposure to this chemical has been proposed to be a potential risk to human health. Exposure to BPA can negatively impact sperm quality. However, the mechanism remains largely unknown. The objectives of this study were to assess the role of BPA on sperm quality and explore the possible mechanisms. The Wistar male rats (aged 28 days) were administered BPA by oral gavage for 28 days at dose of 50, 100 and 200 mg/kg/day; meanwhile, the negative control with corn oil (0 mg/kg/day BPA) and positive control with E2 at the dose of 100 μg/kg/day. The sperm density, sperm activity and sperm survival rate were analyzed byCASA system, and the sperm abnormality rate was analyzed by improved Papanicolaou stained. The protein expression levels of Src/p-Src, ERK1/2, p-ERK1/2 and CREB/p-CREB were detected by Western bolt. The results showed that the body weight gain, testes weight, testis coefficient, sperm density, sperm activity, sperm survival rate and protein expression levels of p-ERK1, p-ERK2 and p-CREB decreased, but the sperm abnormality rate increased with increasing BPA concentrations. There were positive correlations between sperm density, sperm activity and sperm survival rate with protein expression levels of p-ERK1, p-ERK2 and p-CREB, and negative correlations between sperm abnormality rate with the protein expression levels of p-ERK1, p-ERK2 and p-CREB. Results from the structural equation model demonstrated that BPA retained a significant negative effect to p-ERK, whereas p-ERK retained a significant positive effect to sperm quality and acted as the mediate variable. This study provides a novel insight regarding the potential role of p-ERK1 and p-ERK2 protein kinase on reproductive toxicity of BPA. The adverse effects of BPA on adult male sperm quality may be through the induction of the disruption of ERK signal pathway. However, additional research is needed to confirm our findings and to further test the suggested potential mechanisms.

Hijacking of the O-GlcNAcZYME complex by the HTLV-1 Tax oncoprotein facilitates viral transcription

PubMed Central

Waast, Laetitia; Kuo, Mei-Shiue; Mangeney, Marianne; Martella, Christophe; Souidi, Mouloud; Issad, Tarik

2017-01-01

The viral Tax oncoprotein plays a key role in both Human T-cell lymphotropic virus type 1 (HTLV-1)-replication and HTLV-1-associated pathologies, notably adult T-cell leukemia. Tax governs the transcription from the viral 5’LTR, enhancing thereby its own expression, via the recruitment of dimers of phosphorylated CREB to cAMP-response elements located within the U3 region (vCRE). In addition to phosphorylation, CREB is also the target of O-GlcNAcylation, another reversible post-translational modification involved in a wide range of diseases, including cancers. O-GlcNAcylation consists in the addition of O-linked-N-acetylglucosamine (O-GlcNAc) on Serine or Threonine residues, a process controlled by two enzymes: O-GlcNAc transferase (OGT), which transfers O-GlcNAc on proteins, and O-GlcNAcase (OGA), which removes it. In this study, we investigated the status of O-GlcNAcylation enzymes in HTLV-1-transformed T cells. We found that OGA mRNA and protein expression levels are increased in HTLV-1-transformed T cells as compared to control T cell lines while OGT expression is unchanged. However, higher OGA production coincides with a reduction in OGA specific activity, showing that HTLV-1-transformed T cells produce high level of a less active form of OGA. Introducing Tax into HEK-293T cells or Tax-negative HTLV-1-transformed TL-om1 T cells is sufficient to inhibit OGA activity and increase total O-GlcNAcylation, without any change in OGT activity. Furthermore, Tax interacts with the OGT/OGA complex and inhibits the activity of OGT-bound OGA. Pharmacological inhibition of OGA increases CREB O-GlcNAcylation as well as HTLV-1-LTR transactivation by Tax and CREB recruitment to the LTR. Moreover, overexpression of wild-type CREB but not a CREB protein mutated on a previously described O-GlcNAcylation site enhances Tax-mediated LTR transactivation. Finally, both OGT and OGA are recruited to the LTR. These findings reveal the interplay between Tax and the O-GlcNAcylation pathway and identify new key molecular actors involved in the assembly of the Tax-dependent transactivation complex. PMID:28742148

Hijacking of the O-GlcNAcZYME complex by the HTLV-1 Tax oncoprotein facilitates viral transcription.

PubMed

Groussaud, Damien; Khair, Mostafa; Tollenaere, Armelle I; Waast, Laetitia; Kuo, Mei-Shiue; Mangeney, Marianne; Martella, Christophe; Fardini, Yann; Coste, Solène; Souidi, Mouloud; Benit, Laurence; Pique, Claudine; Issad, Tarik

2017-07-01

The viral Tax oncoprotein plays a key role in both Human T-cell lymphotropic virus type 1 (HTLV-1)-replication and HTLV-1-associated pathologies, notably adult T-cell leukemia. Tax governs the transcription from the viral 5'LTR, enhancing thereby its own expression, via the recruitment of dimers of phosphorylated CREB to cAMP-response elements located within the U3 region (vCRE). In addition to phosphorylation, CREB is also the target of O-GlcNAcylation, another reversible post-translational modification involved in a wide range of diseases, including cancers. O-GlcNAcylation consists in the addition of O-linked-N-acetylglucosamine (O-GlcNAc) on Serine or Threonine residues, a process controlled by two enzymes: O-GlcNAc transferase (OGT), which transfers O-GlcNAc on proteins, and O-GlcNAcase (OGA), which removes it. In this study, we investigated the status of O-GlcNAcylation enzymes in HTLV-1-transformed T cells. We found that OGA mRNA and protein expression levels are increased in HTLV-1-transformed T cells as compared to control T cell lines while OGT expression is unchanged. However, higher OGA production coincides with a reduction in OGA specific activity, showing that HTLV-1-transformed T cells produce high level of a less active form of OGA. Introducing Tax into HEK-293T cells or Tax-negative HTLV-1-transformed TL-om1 T cells is sufficient to inhibit OGA activity and increase total O-GlcNAcylation, without any change in OGT activity. Furthermore, Tax interacts with the OGT/OGA complex and inhibits the activity of OGT-bound OGA. Pharmacological inhibition of OGA increases CREB O-GlcNAcylation as well as HTLV-1-LTR transactivation by Tax and CREB recruitment to the LTR. Moreover, overexpression of wild-type CREB but not a CREB protein mutated on a previously described O-GlcNAcylation site enhances Tax-mediated LTR transactivation. Finally, both OGT and OGA are recruited to the LTR. These findings reveal the interplay between Tax and the O-GlcNAcylation pathway and identify new key molecular actors involved in the assembly of the Tax-dependent transactivation complex.

Interconnected network motifs control podocyte morphology and kidney function.

PubMed

Azeloglu, Evren U; Hardy, Simon V; Eungdamrong, Narat John; Chen, Yibang; Jayaraman, Gomathi; Chuang, Peter Y; Fang, Wei; Xiong, Huabao; Neves, Susana R; Jain, Mohit R; Li, Hong; Ma'ayan, Avi; Gordon, Ronald E; He, John Cijiang; Iyengar, Ravi

2014-02-04

Podocytes are kidney cells with specialized morphology that is required for glomerular filtration. Diseases, such as diabetes, or drug exposure that causes disruption of the podocyte foot process morphology results in kidney pathophysiology. Proteomic analysis of glomeruli isolated from rats with puromycin-induced kidney disease and control rats indicated that protein kinase A (PKA), which is activated by adenosine 3',5'-monophosphate (cAMP), is a key regulator of podocyte morphology and function. In podocytes, cAMP signaling activates cAMP response element-binding protein (CREB) to enhance expression of the gene encoding a differentiation marker, synaptopodin, a protein that associates with actin and promotes its bundling. We constructed and experimentally verified a β-adrenergic receptor-driven network with multiple feedback and feedforward motifs that controls CREB activity. To determine how the motifs interacted to regulate gene expression, we mapped multicompartment dynamical models, including information about protein subcellular localization, onto the network topology using Petri net formalisms. These computational analyses indicated that the juxtaposition of multiple feedback and feedforward motifs enabled the prolonged CREB activation necessary for synaptopodin expression and actin bundling. Drug-induced modulation of these motifs in diseased rats led to recovery of normal morphology and physiological function in vivo. Thus, analysis of regulatory motifs using network dynamics can provide insights into pathophysiology that enable predictions for drug intervention strategies to treat kidney disease.

Interconnected Network Motifs Control Podocyte Morphology and Kidney Function

PubMed Central

Azeloglu, Evren U.; Hardy, Simon V.; Eungdamrong, Narat John; Chen, Yibang; Jayaraman, Gomathi; Chuang, Peter Y.; Fang, Wei; Xiong, Huabao; Neves, Susana R.; Jain, Mohit R.; Li, Hong; Ma’ayan, Avi; Gordon, Ronald E.; He, John Cijiang; Iyengar, Ravi

2014-01-01

Podocytes are kidney cells with specialized morphology that is required for glomerular filtration. Diseases, such as diabetes, or drug exposure that causes disruption of the podocyte foot process morphology results in kidney pathophysiology. Proteomic analysis of glomeruli isolated from rats with puromycin-induced kidney disease and control rats indicated that protein kinase A (PKA), which is activated by adenosine 3′,5′-monophosphate (cAMP), is a key regulator of podocyte morphology and function. In podocytes, cAMP signaling activates cAMP response element–binding protein (CREB) to enhance expression of the gene encoding a differentiation marker, synaptopodin, a protein that associates with actin and promotes its bundling. We constructed and experimentally verified a β-adrenergic receptor–driven network with multiple feedback and feedforward motifs that controls CREB activity. To determine how the motifs interacted to regulate gene expression, we mapped multicompartment dynamical models, including information about protein subcellular localization, onto the network topology using Petri net formalisms. These computational analyses indicated that the juxtaposition of multiple feedback and feedforward motifs enabled the prolonged CREB activation necessary for synaptopodin expression and actin bundling. Drug-induced modulation of these motifs in diseased rats led to recovery of normal morphology and physiological function in vivo. Thus, analysis of regulatory motifs using network dynamics can provide insights into pathophysiology that enable predictions for drug intervention strategies to treat kidney disease. PMID:24497609

Neurotrophic Effect of Citrus 5-Hydroxy-3,6,7,8,3′,4′-Hexamethoxyflavone: Promotion of Neurite Outgrowth via cAMP/PKA/CREB Pathway in PC12 Cells

PubMed Central

Lai, Hui-Chi; Wu, Ming-Jiuan; Chen, Pei-Yi; Sheu, Ting-Ting; Chiu, Szu-Ping; Lin, Meng-Han; Ho, Chi-Tang; Yen, Jui-Hung

2011-01-01

5-Hydroxy-3,6,7,8,3′,4′-hexamethoxyflavone (5-OH-HxMF), a hydroxylated polymethoxyflavone, is found exclusively in the Citrus genus, particularly in the peels of sweet orange. In this research, we report the first investigation of the neurotrophic effects and mechanism of 5-OH-HxMF in PC12 pheochromocytoma cells. We found that 5-OH-HxMF can effectively induce PC12 neurite outgrowth accompanied with the expression of neuronal differentiation marker protein growth-associated protein-43(GAP-43). 5-OH-HxMF caused the enhancement of cyclic AMP response element binding protein (CREB) phosphorylation, c-fos gene expression and CRE-mediated transcription, which was inhibited by 2-naphthol AS-E phosphate (KG-501), a specific antagonist for the CREB-CBP complex formation. Moreover, 5-OH-HxMF-induced both CRE transcription activity and neurite outgrowth were inhibited by adenylate cyclase and protein kinase A (PKA) inhibitor, but not MEK1/2, protein kinase C (PKC), phosphatidylinositol 3-kinase (PI3K) or calcium/calmodulin-dependent protein kinase (CaMK) inhibitor. Consistently, 5-OH-HxMF treatment increased the intracellular cAMP level and downstream component, PKA activity. We also found that addition of K252a, a TrKA antagonist, significantly inhibited NGF- but not 5-OH-HxMF-induced neurite outgrowth. These results reveal for the first time that 5-OH-HxMF is an effective neurotrophic agent and its effect is mainly through a cAMP/PKA-dependent, but TrKA-independent, signaling pathway coupling with CRE-mediated gene transcription. A PKC-dependent and CREB-independent pathway was also involved in its neurotrophic action. PMID:22140566

Silymarin suppresses the PGE2 -induced cell migration through inhibition of EP2 activation; G protein-dependent PKA-CREB and G protein-independent Src-STAT3 signal pathways.

PubMed

Woo, Seon Min; Min, Kyoung-Jin; Chae, In Gyeong; Chun, Kyung-Soo; Kwon, Taeg Kyu

2015-03-01

Silymarin has been known as a chemopreventive agent, and possesses multiple anti-cancer activities including induction of apoptosis, inhibition of proliferation and growth, and blockade of migration and invasion. However, whether silymarin could inhibit prostaglandin (PG) E2 -induced renal cell carcinoma (RCC) migration and what are the underlying mechanisms are not well elucidated. Here, we found that silymarin markedly inhibited PGE2 -stimulated migration. PGE2 induced G protein-dependent CREB phosphorylation via protein kinase A (PKA) signaling, and PKA inhibitor (H89) inhibited PGE2 -mediated migration. Silymarin reduced PGE2 -induced CREB phosphorylation and CRE-promoter activity. PGE2 also activated G protien-independent signaling pathways (Src and STAT3) and silymarin reduced PGE2 -induced phosphorylation of Src and STAT3. Inhibitor of Src (Saracatinib) markedly reduced PGE2 -mediated migration. We found that EP2, a PGE2 receptor, is involved in PGE2 -mediated cell migration. Down regulation of EP2 by EP2 siRNA and EP2 antagonist (AH6809) reduced PGE2 -inudced migration. In contrast, EP2 agonist (Butaprost) increased cell migration and silymarin effectively reduced butaprost-mediated cell migration. Moreover, PGE2 increased EP2 expression through activation of positive feedback mechanism, and PGE2 -induced EP2 expression, as well as basal EP2 levels, were reduced in silymarin-treated cells. Taken together, our study demonstrates that silymarin inhibited PGE2 -induced cell migration through inhibition of EP2 signaling pathways (G protein dependent PKA-CREB and G protein-independent Src-STAT3). © 2013 Wiley Periodicals, Inc.

Estrogen receptor ESR1 mediates activation of ERK1/2, CREB, and ELK1 in the corpus of the epididymis.

PubMed

Cavalcanti, Fernanda N; Lucas, Thais F G; Lazari, Maria Fatima M; Porto, Catarina S

2015-06-01

Expression of the estrogen receptor ESR1 is higher in the corpus than it is in the initial segment/caput and cauda of the epididymis. ESR1 immunostaining in the corpus has been localized not only in the nuclei but also in the cytoplasm and apical membrane, which indicates that ESR1 plays a role in membrane-initiated signaling. The present study investigated whether ESR1 mediates the activation of rapid signaling pathways by estradiol (E2) in the epididymis. We investigated the effect of E2 and the ESR1-selective agonist (4,4',4''-(4-propyl-(1H)-pyrazole-1,3,5-triyl)trisphenol (PPT) on the activation of extracellular signal-regulated protein kinases (ERK1/2), CREB protein, and ETS oncogene-related protein (ELK1). Treatment with PPT did not affect ERK1/2 phosphorylation in the cauda, but it rapidly increased ERK1/2 phosphorylation in the initial segment/caput and corpus of the epididymis. PPT also activated CREB and ELK1 in the corpus of the epididymis. The PPT-induced phosphorylation of ERK1/2, CREB, and ELK1 was blocked by the ESR1-selective antagonist MPP and by pretreatment with a non-receptor tyrosine kinase SRC inhibitor, an EGFR kinase inhibitor, an MEK1/2 inhibitor, and a phosphatidylinositol-3-kinase inhibitor. In conclusion, these results indicate that the corpus, which is a region with high expression of the estrogen receptor ESR1, is a major target in the epididymis for the activation of rapid signaling by E2. The sequence of events that follow E2 interaction with ESR1 includes the SRC-mediated transactivation of EGFR and the phosphorylation of ERK1/2, CREB, and ELK1. This rapid estrogen signaling may modulate gene expression in the corpus of the epididymis, and it may play a role in the dynamic microenvironment of the epididymal lumen. © 2015 Society for Endocrinology.

Mechanisms of angiogenesis in a Curculigoside A-treated rat model of cerebral ischemia and reperfusion injury

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhu, Haibo; Institute of Toxicology, Binzhou Medical University, Yantai; He, Jie

Curculigoside A has shown protective effects against rat cortical neuron damage in vivo. However, the molecular mechanisms through which Curculigoside A affords this protection are unclear. In the present study, we sought to elucidate the mechanisms of angiogenesis in rat aortic endothelial cells (RAEC), rat aortic smooth muscle cells (RASMC) as well as a rat model of cerebral ischemia and reperfusion injury following treatment with Curculigoside A. We examined the role of Curculigoside A on RAEC and RASMC proliferation, migration, and tube formation in vitro and in a cerebral ischemia and reperfusion injury rat model. We used the recombinant Dickkopfmore » (DKK)-1 protein, a Wnt/β-catenin inhibitor, and the recombinant WIF-1 protein, a Wnt5a antagonist to determine mechanisms. In addition, we measured leakage of the blood–brain barrier (BBB) and tested for angiogenesis associated proteins. Our data suggest that Curculigoside A induces angiogenesis in vitro by increasing proliferation, migration and tube formation in RAEC and RASMC. The increase in Curculigoside A-induced proliferation and tube formation was counteracted by DKK-1 and WIF-1. Curculigoside A increased expression of VEGF, p-VEGFR, p-CREB, Egr-3, VCAM-1, Ang1 and Tie2 while prohibiting BBB leakage in cerebral ischemia and reperfusion injured rats. However, Cyclosporine A, a CREB inhibitor, reduced the expression of p-CREB, Egr-3, VCAM-1, Ang1 and Tie2. These data suggest that Curculigoside A induces cell proliferation and angiogenesis through the Wnt5a/β-catenin and VEGF/CREB/Egr-3/VCAM-1 signaling axis and promotes maturation and stability of new blood vessels via increasing Ang1 and Tie-2 expression. - Highlights: • Curculigoside A induces cell proliferation through Wnt5a/β-catenin pathway. • Curculigoside A induces angiogenesis via VEGF/CREB/Egr-3/VCAM-1 signaling axis. • Curculigoside A promotes blood vessel maturation via Ang1/Tie2 pathway.« less

Suppression of type I interferon production by porcine epidemic diarrhea virus and degradation of CREB-binding protein by nsp1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Qingzhan; Shi, Kaichuang; Yoo, Dongwan, E-mail: dyoo@illinois.edu

Type I interferons (IFN-α/β) are the major components of the innate immune response of hosts, and in turn many viruses have evolved to modulate the host response during infection. We found that the IFN-β production was significantly suppressed during PEDV infection in cells. To identify viral IFN antagonists and to study their suppressive function, viral coding sequences for the entire structural and nonstructural proteins were cloned and expressed. Of 16 PEDV nonstructural proteins (nsps), nsp1, nsp3, nsp7, nsp14, nsp15 and nsp16 were found to inhibit the IFN-β and IRF3 promoter activities. The sole accessory protein ORF3, structure protein envelope (E),more » membrane (M), and nucleocapsid (N) protein were also shown to inhibit such activities. PEDV nsp1 did not interfere the IRF3 phosphorylation and nuclear translocation but interrupted the enhanceosome assembly of IRF3 and CREB-binding protein (CBP) by degrading CBP. A further study showed that the CBP degradation by nsp1 was proteasome-dependent. Our data demonstrate that PEDV modulates the host innate immune responses by degrading CBP and suppressing ISGs expression. - Highlights: • PEDV modulates the host innate immune system by suppressing the type I interferon production and ISGs expression. • Ten viral proteins were identified as IFN antagonists, and nsp1 was the most potent viral IFN antagonist. • PEDV nsp1 did not interfere the IRF3 phosphorylation and nuclear translocation but interrupted the enhanceosome assembly of IRF3 and CREB-binding protein (CBP). • PEDV nsp1 caused the CBP degradation in the nucleus, which may be the key mechanism for PEDV-mediated IFN downregulation.« less

Regulatory motifs for CREB-binding protein and Nfe2l2 transcription factors in the upstream enhancer of the mitochondrial uncoupling protein 1 gene.

PubMed

Rim, Jong S; Kozak, Leslie P

2002-09-13

Thermogenesis against cold exposure in mammals occurs in brown adipose tissue (BAT) through mitochondrial uncoupling protein (UCP1). Expression of the Ucp1 gene is unique in brown adipocytes and is regulated tightly. The 5'-flanking region of the mouse Ucp1 gene contains cis-acting elements including PPRE, TRE, and four half-site cAMP-responsive elements (CRE) with BAT-specific enhancer elements. In the course of analyzing how these half-site CREs are involved in Ucp1 expression, we found that a DNA regulatory element for NF-E2 overlaps CRE2. Electrophoretic mobility shift assay and competition assays with the CRE2 element indicates that nuclear proteins from BAT, inguinal fat, and retroperitoneal fat tissue interact with the CRE2 motif (CGTCA) in a specific manner. A supershift assay using an antibody against the CRE-binding protein (CREB) shows specific affinity to the complex from CRE2 and nuclear extract of BAT. Additionally, Western blot analysis for phospho-CREB/ATF1 shows an increase in phosphorylation of CREB/ATF1 in HIB-1B cells after norepinephrine treatment. Transient transfection assay using luciferase reporter constructs also indicates that the two half-site CREs are involved in transcriptional regulation of Ucp1 in response to norepinephrine and cAMP. We also show that a second DNA regulatory element for NF-E2 is located upstream of the CRE2 region. This element, which is found in a similar location in the 5'-flanking region of the human and rodent Ucp1 genes, shows specific binding to rat and human NF-E2 by electrophoretic mobility shift assay with nuclear extracts from brown fat. Co-transfections with an Nfe2l2 expression vector and a luciferase reporter construct of the Ucp1 enhancer region provide additional evidence that Nfe2l2 is involved in the regulation of Ucp1 by cAMP-mediated signaling.

PKA-CREB-BDNF signaling pathway mediates propofol-induced long-term learning and memory impairment in hippocampus of rats.

PubMed

Zhong, Yu; Chen, Jing; Li, Li; Qin, Yi; Wei, Yi; Pan, Shining; Jiang, Yage; Chen, Jialin; Xie, Yubo

2018-04-20

Studies have found that propofol can induce widespread neuroapoptosis in developing brains, which leads to cause long-term learning and memory abnormalities. However, the specific cellular and molecular mechanisms underlying propofol-induced neuroapoptosis remain elusive. The aim of the present study was to explore the role of PKA-CREB-BDNF signaling pathway in propofol-induced long-term learning and memory impairment during brain development. Seven-day-old rats were randomly assigned to control, intralipid and three treatment groups (n = 5). Rats in control group received no treatment. Intralipid (10%, 10 mL/kg) for vehicle control and different dosage of propofol for three treatment groups (50, 100 and 200 mg/kg) were administered intraperitoneally. FJB staining, immunohistochemistry analysis for neuronal nuclei antigen and transmission electron microscopy were used to detect neuronal apoptosis and structure changes. MWM test examines the long-term spatial learning and memory impairment. The expression of PKA, pCREB and BDNF was quantified using western blots. Propofol induced significant increase of FJB-positive cells and decrease of PKA, pCREB and BDNF protein levels in the immature brain of P7 rats. Using the MWM test, propofol-treated rats demonstrated long-term spatial learning and memory impairment. Moreover, hippocampal NeuN-positive cell loss, long-lasting ultrastructural abnormalities of the neurons and synapses, and long-term down-regulation of PKA, pCREB and BDNF protein expression in adult hippocampus were also found. Our results indicated that neonatal propofol exposure can significantly result in long-term learning and memory impairment in adulthood. The possible mechanism involved in the propofol-induced neuroapoptosis was related to down-regulation of PKA-CREB-BDNF signaling pathway. Copyright © 2018. Published by Elsevier B.V.

Sodium Phenylbutyrate Enhances Astrocytic Neurotrophin Synthesis via Protein Kinase C (PKC)-mediated Activation of cAMP-response Element-binding Protein (CREB)

PubMed Central

Corbett, Grant T.; Roy, Avik; Pahan, Kalipada

2013-01-01

Neurotrophins, such as brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3), are believed to be genuine molecular mediators of neuronal growth and homeostatic synapse activity. However, levels of these neurotrophic factors decrease in different brain regions of patients with Alzheimer disease (AD). Induction of astrocytic neurotrophin synthesis is a poorly understood phenomenon but represents a plausible therapeutic target because neuronal neurotrophin production is aberrant in AD and other neurodegenerative diseases. Here, we delineate that sodium phenylbutyrate (NaPB), a Food and Drug Administration-approved oral medication for hyperammonemia, induces astrocytic BDNF and NT-3 expression via the protein kinase C (PKC)-cAMP-response element-binding protein (CREB) pathway. NaPB treatment increased the direct association between PKC and CREB followed by phosphorylation of CREB (Ser133) and induction of DNA binding and transcriptional activation of CREB. Up-regulation of markers for synaptic function and plasticity in cultured hippocampal neurons by NaPB-treated astroglial supernatants and its abrogation by anti-TrkB blocking antibody suggest that NaPB-induced astroglial neurotrophins are functionally active. Moreover, oral administration of NaPB increased the levels of BDNF and NT-3 in the CNS and improved spatial learning and memory in a mouse model of AD. Our results highlight a novel neurotrophic property of NaPB that may be used to augment neurotrophins in the CNS and improve synaptic function in disease states such as AD. PMID:23404502

Sodium phenylbutyrate enhances astrocytic neurotrophin synthesis via protein kinase C (PKC)-mediated activation of cAMP-response element-binding protein (CREB): implications for Alzheimer disease therapy.

PubMed

Corbett, Grant T; Roy, Avik; Pahan, Kalipada

2013-03-22

Neurotrophins, such as brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3), are believed to be genuine molecular mediators of neuronal growth and homeostatic synapse activity. However, levels of these neurotrophic factors decrease in different brain regions of patients with Alzheimer disease (AD). Induction of astrocytic neurotrophin synthesis is a poorly understood phenomenon but represents a plausible therapeutic target because neuronal neurotrophin production is aberrant in AD and other neurodegenerative diseases. Here, we delineate that sodium phenylbutyrate (NaPB), a Food and Drug Administration-approved oral medication for hyperammonemia, induces astrocytic BDNF and NT-3 expression via the protein kinase C (PKC)-cAMP-response element-binding protein (CREB) pathway. NaPB treatment increased the direct association between PKC and CREB followed by phosphorylation of CREB (Ser(133)) and induction of DNA binding and transcriptional activation of CREB. Up-regulation of markers for synaptic function and plasticity in cultured hippocampal neurons by NaPB-treated astroglial supernatants and its abrogation by anti-TrkB blocking antibody suggest that NaPB-induced astroglial neurotrophins are functionally active. Moreover, oral administration of NaPB increased the levels of BDNF and NT-3 in the CNS and improved spatial learning and memory in a mouse model of AD. Our results highlight a novel neurotrophic property of NaPB that may be used to augment neurotrophins in the CNS and improve synaptic function in disease states such as AD.

Effects of Carbocysteine and Beclomethasone on Histone Acetylation/Deacetylation Processes in Cigarette Smoke Exposed Bronchial Epithelial Cells.

PubMed

Pace, Elisabetta; Di Vincenzo, Serena; Ferraro, Maria; Siena, Liboria; Chiappara, Giuseppina; Dino, Paola; Vitulo, Patrizio; Bertani, Alessandro; Saibene, Federico; Lanata, Luigi; Gjomarkaj, Mark

2017-10-01

Histone deacetylase expression/activity may control inflammation, cell senescence, and responses to corticosteroids. Cigarette smoke exposure, increasing oxidative stress, may negatively affect deacetylase expression/activity. The effects of cigarette smoke extracts (CSE), carbocysteine, and beclomethasone dipropionate on chromatin remodeling processes in human bronchial epithelial cells are largely unknown. The present study was aimed to assess the effects of cigarette smoke, carbocysteine, and beclomethasone dipropionate on histone deacetylase 3 (HDAC3) expression/activity, N-CoR (nuclear receptor corepressor) expression, histone acetyltransferases (HAT) (p300/CBP) expression, p-CREB and IL-1 m-RNA expression, neutrophil chemotaxis. Increased p-CREB expression was observed in the bronchial epithelium of smokers. CSE increased p-CREB expression and decreased HDAC3 expression and activity and N-CoR m-RNA and protein expression. At the same time, CSE increased the expression of the HAT, p300/CBP. All these events increased acetylation processes within the cells and were associated to increased IL-1 m-RNA expression and neutrophil chemotaxis. The incubation of CSE exposed cells with carbocysteine and beclomethasone counteracted the effects of cigarette smoke on HDAC3 and N-CoR but not on p300/CBP. The increased deacetylation processes due to carbocysteine and beclomethasone dipropionate incubation is associated to reduced p-CREB, IL-1 m-RNA expression, neutrophil chemotaxis. These findings suggest a new role of combination therapy with carbocysteine and beclomethasone dipropionate in restoring deacetylation processes compromised by cigarette smoke exposure. J. Cell. Physiol. 232: 2851-2859, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

CREB-1 and AP-1 transcription factors JunD and Fra-2 regulate bone sialoprotein gene expression in human breast cancer cells.

PubMed

Detry, C; Lamour, V; Castronovo, V; Bellahcène, A

2008-02-01

Bone sialoprotein (BSP) expression is detected in a variety of human osteotropic cancers. High expression of BSP in breast and prostate primary carcinomas is associated with progression and bone metastases development. In this study, we examined the transcriptional regulation of BSP gene expression in MDA-MB-231 and MCF-7 human breast cancer cells compared with Saos-2 human osteoblast-like cells. BSP human promoter deletion analyses delineated a -56/-84 region, which comprises a cAMP response element (CRE) that was sufficient for maximal promoter activity in breast cancer cell lines. We found that the basic fibroblast growth factor response element (FRE) also located in the proximal promoter was a crucial regulator of human BSP promoter activity in Saos-2 but not in breast cancer cells. Promoter activity experiments in combination with DNA mobility shift assays demonstrated that BSP promoter activity is under the control of the CRE element, through CREB-1, JunD and Fra-2 binding, in MDA-MB-231, MCF-7 and in Saos-2 cells. Forskolin, a protein kinase A pathway activator, failed to enhance BSP transcriptional activity suggesting that CRE site behaves as a constitutive rather than an inducible element in these cell lines. Over-expression of JunD and Fra-2 increased BSP promoter activity and upregulated endogenous BSP protein expression in MCF-7 and Saos-2 cells while siRNA-mediated inhibition of both factors expression significantly reduced BSP protein level in MDA-MB-231. Collectively, these data provide with new transcriptional mechanisms, implicating CREB and AP-1 factors, that control BSP gene expression in breast cancer cells.

Induction of cyclooxygenase-2 by ginsenoside Rd via activation of CCAAT-enhancer binding proteins and cyclic AMP response binding protein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jeong, Hye Gwang; Pokharel, Yuba Raj; Han, Eun Hee

2007-07-20

Panax ginseng is a widely used herbal medicine in East Asia and is reported to have a variety of pharmacological effects against cardiovascular diseases and cancers. Here we show a unique effect of ginsenoside Rd (Rd) on cyclooxygenase-2 (COX-2) expression in RAW264.7 macrophages. Rd (100 {mu}g/ml), but not other ginsenosides induced COX-2 and increased prostaglandin E{sub 2} production. Gel shift and Western blot analyses using nuclear fractions revealed that Rd increased both the DNA binding of and the nuclear levels of CCAAT/enhancer binding protein (C/EBP){alpha}/{beta} and cyclic AMP response element binding protein (CREB), but not of p65, in RAW264.7 cells.more » Moreover, Rd increased the luciferase reporter gene activity in cells transfected with a 574-bp mouse COX-2 promoter construct. Site-specific mutation analyses confirmed that Rd-mediated transcriptional activation of COX-2 gene was regulated by C/EBP and CREB. These results provide evidence that Rd activated C/EBP and CREB, and that the activation of C/EBP and CREB appears to be essential for induction of COX-2 in RAW264.7 cells.« less

Extracellular visfatin activates gluconeogenesis in HepG2 cells through the classical PKA/CREB-dependent pathway.

PubMed

Choi, Y J; Choi, S-E; Ha, E S; Kang, Y; Han, S J; Kim, D J; Lee, K W; Kim, H J

2014-04-01

Adipokines reportedly affect hepatic gluconeogenesis, and the adipokine visfatin is known to be related to insulin resistance and type 2 diabetes. However, whether visfatin contributes to hepatic gluconeogenesis remains unclear. Visfatin, also known as nicotinamide phosphoribosyltransferase (NAMPT), modulates sirtuin1 (SIRT1) through the regulation of nicotinamide adenine dinucleotide (NAD). Therefore, we investigated the effect of extracellular visfatin on glucose production in HepG2 cells, and evaluated whether extracellular visfatin affects hepatic gluconeogenesis via an NAD+-SIRT1-dependent pathway. Treatment with visfatin significantly increased glucose production and the mRNA expression and protein levels of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) in HepG2 cells in a time- and concentration-dependent manner. Knockdown of SIRT1 had no remarkable effect on the induction of gluconeogenesis by visfatin. Subsequently, we evaluated if extracellular visfatin stimulates the production of gluconeogenic enzymes through the classical protein kinase A (PKA)/cyclic AMP-responsive element (CRE)-binding protein (CREB)-dependent process. The phosphorylation of CREB and PKA increased significantly in HepG2 cells treated with visfatin. Additionally, knockdown of CREB and PKA inhibited visfatin-induced gluconeogenesis in HepG2 cells. In summary, extracellular visfatin modulates glucose production in HepG2 cells through the PKA/CREB pathway, rather than via SIRT1 signaling. © Georg Thieme Verlag KG Stuttgart · New York.

Epigenetic Regulation of the NR4A Orphan Nuclear Receptor NOR1 By Histone Acetylation

PubMed Central

Zhao, Yue; Nomiyama, Takashi; Findeisen, Hannes M.; Qing, Hua; Aono, Jun; Jones, Karrie L.; Heywood, Elizabeth B.; Bruemmer, Dennis

2014-01-01

The nuclear receptor NOR1 is an immediate-early response gene implicated in the transcriptional control of proliferation. Since the expression level of NOR1 is rapidly induced through cAMP response element binding (CREB) protein-dependent promoter activation, we investigated the contribution of histone acetylation to this transient induction. We demonstrate that NOR1 transcription is induced by histone deacetylase (HDAC) inhibition and by depletion of HDAC1 and HDAC3. HDAC inhibition activated the NOR1 promoter, increased histone acetylation and augmented the recruitment of phosphorylated CREB to the promoter. Furthermore, HDAC inhibition increased Ser133 phosphorylation of CREB and augmented NOR1 protein stability. These data outline previously unrecognized mechanisms of NOR1 regulation and illustrate a key role for histone acetylation in the rapid induction of NOR1. PMID:25451221

NR2B-containing NMDA receptors promote neural progenitor cell proliferation through CaMKIV/CREB pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Mei, E-mail: limeihit@163.com; Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, Nanjing; Zhang, Dong-Qing

2011-08-12

Highlights: {yields} The NR2B component of the NMDARs is important for the NSPC proliferation. {yields} pCaMKIV and pCREB exist in NSPCs. {yields} The CaMKIV/CREB pathway mediates NSPC proliferation. -- Abstract: Accumulating evidence indicates the involvement of N-methyl-D-aspartate receptors (NMDARs) in regulating neural stem/progenitor cell (NSPC) proliferation. Functional properties of NMDARs can be markedly influenced by incorporating the regulatory subunit NR2B. Here, we aim to analyze the effect of NR2B-containing NMDARs on the proliferation of hippocampal NSPCs and to explore the mechanism responsible for this effect. NSPCs were shown to express NMDAR subunits NR1 and NR2B. The NR2B selective antagonist, Romore » 25-6981, prevented the NMDA-induced increase in cell proliferation. Moreover, we demonstrated that the phosphorylation levels of calcium/calmodulin-dependent protein kinase IV (CaMKIV) and cAMP response element binding protein (CREB) were increased by NMDA treatment, whereas Ro 25-6981 decreased them. The role that NR2B-containing NMDARs plays in NSPC proliferation was abolished when CREB phosphorylation was attenuated by CaMKIV silencing. These results suggest that NR2B-containing NMDARs have a positive role in regulating NSPC proliferation, which may be mediated through CaMKIV phosphorylation and subsequent induction of CREB activation.« less

TLR4 induces CREB-mediated IL-6 production via upregulation of F-spondin to promote vascular smooth muscle cell migration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Guan-Lin; Graduate Institutes of Life Sciences, National Defense Medical Center, Taipei, Taiwan; Wu, Jing-Yiing

Toll-like receptor 4 (TLR4) is important in promoting inflammation and vascular smooth muscle cell (VSMC) migration, both of which contribute to atherosclerosis development and progression. But the mechanism underlying the regulation of TLR4 in VSMC migration remains unclear. Stimulation of VSMCs with LPS increased the cellular level of F-spondin which is associated with the regulation of proinflammatory cytokine production. The LPS-induced F-spondin expression depended on TLR4-mediated PI3K/Akt pathway. Suppression of F-spondin level by siRNA inhibited not only F-spondin expression but also LPS-induced phosphorylation of cAMP response element binding protein (CREB) and IL-6 expression, VSMC migration and proliferation as well asmore » MMP9 expression. Moreover, suppression of CREB level by siRNA inhibited TLR4-induced IL-6 production and VSMC migration. Inhibition of F-spondin siRNA on LPS-induced migration was restored by addition of exogenous recombinant mouse IL-6. We conclude that upon ligand binding, TLR4 activates PI3K/Akt signaling to induce F-spondin expression, subsequently control CREB-mediated IL-6 production to promote VSMC migration. These findings provide vital insights into the essential role of F-spondin in VSMC function and will be valuable for developing new therapeutic strategies against atherosclerosis. -- Highlights: •LPS-induced F-spondin expression of VSMCs is via a TLR4/PI3K/Akt signaling. •F-spondin is pivotal for LPS-induced CREB-mediated IL-6 production. •F-spondin is required for LPS-induced VSMC migration and proliferation.« less

17beta-estradiol stimulates the growth of human keratinocytes by inducing cyclin D2 expression.

PubMed

Kanda, Naoko; Watanabe, Shinichi

2004-08-01

Estrogen is reported to prevent age-associated epidermal thinning in the skin. We examined if 17beta-estradiol (E2) may enhance the growth of human keratinocytes, focusing on its effects on the expression of cell cycle-regulatory proteins. E2 enhanced proliferation, bromodeoxyuridine incorporation of keratinocytes, and increased the proportion of cells in the S phase. The E2-induced stimulation of proliferation and bromodeoxyuridine incorporation was suppressed by antisense oligonucleotide against cyclin D2, which induces G1 to S phase progression. E2 increased protein and mRNA levels of cyclin D2, and resultantly enhanced assembly and kinase activities of cyclin D2-cyclin-dependent kinases 4 or 6 complexes. E2 enhanced cyclin D2 promoter activity, and the element homologous to cAMP response element (CRE) on the promoter was responsible for the effect. Cyclin D2 expression was enhanced by antiestrogens, ICI 182,780 and 4-hydroxytamoxifen, and membrane-impermeable bovine serum albumin-conjugated E2, indicating the effects via membrane E2-binding sites. E2 increased the enhancer activity of CRE-like element and the amount of phosphorylated cAMP response element binding protein (CREB) binding this element, and the increases were suppressed by H-89, an inhibitor of cAMP-dependent protein kinase A. H-89 also suppressed E2-induced cyclin D2 expression, proliferation, and bromodeoxyuridine incorporation in keratinocytes. Antisense oligonucleotide against G-protein-coupled receptor GPR30 suppressed the E2-induced increases of phosphorylated CREB, cyclin D2 level, proliferation, and bromodeoxyuridine incorporation in keratinocytes. These results suggest that E2 may stimulate the growth of keratinocytes by inducing cyclin D2 expression via CREB phosphorylation by protein kinase A, dependent on cAMP. These effects of E2 may be mediated via cell surface GPR30.

PPARgamma agonists inhibit TGF-beta-PKA signaling in glomerulosclerosis.

PubMed

Zou, Rong; Xu, Gang; Liu, Xiao-cheng; Han, Min; Jiang, Jing-jing; Huang, Qian; He, Yong; Yao, Ying

2010-01-01

To study the probable mechanisms of the anti-glomerulosclerosis effects induced by peroxisome proliferator-activated receptor gamma (PPARgamma) agonists in rat intraglomerular mesangial cells (MCs). Cells were transfected with the pTAL-PPRE-tk-Luc(+) plasmid and then treated with different concentrations of PPARgamma agonist, either troglitazone or telmisartan, for the indicated times. Promega luciferase assays were subsequently used for the detection of PPARgamma activation. Protein expression levels were assessed by Western blot, and PepTag assays were used for the non-radioactive detection of protein kinase A (PKA) activity. The deposition of alpha-smooth muscle actin (alpha-SMA) and p-cyclic AMP responsive element binding protein (pCREB) were analyzed by confocal laser scanning. Both troglitazone and telmisartan remarkably inhibit the PKA activation and pCREB expression that is stimulated by TGF-beta. The PPARgamma agonists also inhibited alpha-SMA and collagen IV protein expression by blocking PKA activation. PPARgamma ligands effectively suppress the activation of MCs and the accumulation of collagen IV stimulated by TGF-beta in vitro. The renal protection provided by PPARgamma agonists is partly mediated via their blockade of TGF-beta/PKA signaling.

Hypoxia-mediated alterations and their role in the HER-2/neuregulated CREB status and localization

PubMed Central

Steven, André; Leisz, Sandra; Sychra, Katharina; Hiebl, Bernhard; Wickenhauser, Claudia; Mougiakakos, Dimitrios; Kiessling, Rolf; Denkert, Carsten; Seliger, Barbara

2016-01-01

The cAMP-responsive element-binding protein (CREB) is involved in the tumorigenicity of HER-2/neu-overexpressing murine and human tumor cells, but a link between the HER-2/neu-mediated CREB activation, its posttranslational modification and localization and changes in the cellular metabolism, due to an altered (tumor) microenvironment remains to be established. The present study demonstrated that shRNA-mediated silencing of CREB in HER-2/neu-transformed cells resulted in decreased tumor formation, which was associated with reduced angiogenesis, but increased necrotic and hypoxic areas in the tumor. Hypoxia induced pCREBSer133, but not pCREBSer121 expression in HER-2/neu-transformed cells. This was accompanied by upregulation of the hypoxia-inducible genes GLUT1 and VEGF, increased cell migration and matrix metalloproteinase-mediated invasion. Treatment of HER-2/neu+ cells with signal transduction inhibitors targeting in particular HER-2/neu was able to revert hypoxia-controlled CREB activation. In addition to changes in the phosphorylation, hypoxic response of HER-2/neu+ cells caused a transient ubiquitination and SUMOylation as well as a co-localization of nuclear CREB to the mitochondrial matrix. A mitochondrial localization of CREB was also demonstrated in hypoxic areas of HER-2/neu+ mammary carcinoma lesions. This was accompanied by an altered gene expression pattern, activity and metabolism of mitochondria leading to an increased respiratory rate, oxidative phosphorylation and mitochondrial membrane potential and consequently to an enhanced apoptosis and reduced cell viability. These data suggest that the HER-2/neu-mediated CREB activation caused by a hypoxic tumor microenvironment contributes to the neoplastic phenotype of HER-2/neu+ cells at various levels. PMID:27409833

Methamphetamine-induced sensitization differentially alters pCREB and DeltaFosB throughout the limbic circuit of the mammalian brain.

PubMed

McDaid, John; Graham, Martin P; Napier, T Celeste

2006-12-01

Enhancements in behavior that accompany repeated, intermittent administration of abused drugs (sensitization) endure long after drug administration has ceased. Such persistence reflects changes in intracellular signaling cascades and associated gene transcription factors in brain regions that are engaged by abused drugs. This process is not characterized for the most potent psychomotor stimulant, methamphetamine. Using motor behavior as an index of brain state in rats, we verified that five once-daily injections of 2.5 mg/kg methamphetamine induced behavioral sensitization that was demonstrated (expressed) 3 and 14 days later. Using immunoblot procedures, limbic brain regions implicated in behavioral sensitization were assayed for extracellular signal-regulated kinase and its phosphorylated form (pERK/ERK, a signal transduction kinase), cAMP response element binding protein and its phosphorylated form (pCREB/CREB, a constitutively expressed transcriptional regulator), and DeltaFosB (a long-lasting transcription factor). pERK, ERK, and CREB levels were not changed for any region assayed. In the ventral tegmental area, pCREB and DeltaFosB also were not changed. pCREB (activated CREB) was elevated in the frontal cortex at 3 days withdrawal, but not at 14 days. pCREB levels were decreased at 14 days withdrawal in the nucleus accumbens and ventral pallidum. Accumbal and pallidal levels of DeltaFosB were increased at 3 days withdrawal, and this increase persisted to 14 days in the pallidum. Thus, only the ventral pallidum showed changes in molecular processes that consistently correlated with motor sensitization, revealing that this region may be associated with this enduring behavioral phenotype initiated by methamphetamine. The present findings expand our understanding of the neuroanatomical and molecular substrates that may play a role in the persistence of druginduced sensitization.

CREB at the Crossroads of Activity-Dependent Regulation of Nervous System Development and Function.

PubMed

Belgacem, Yesser H; Borodinsky, Laura N

2017-01-01

The central nervous system is a highly plastic network of cells that constantly adjusts its functions to environmental stimuli throughout life. Transcription-dependent mechanisms modify neuronal properties to respond to external stimuli regulating numerous developmental functions, such as cell survival and differentiation, and physiological functions such as learning, memory, and circadian rhythmicity. The discovery and cloning of the cyclic adenosine monophosphate (cAMP) responsive element binding protein (CREB) constituted a big step toward deciphering the molecular mechanisms underlying neuronal plasticity. CREB was first discovered in learning and memory studies as a crucial mediator of activity-dependent changes in target gene expression that in turn impose long-lasting modifications of the structure and function of neurons. In this chapter, we review the molecular and signaling mechanisms of neural activity-dependent recruitment of CREB and its cofactors. We discuss the crosstalk between signaling pathways that imprints diverse spatiotemporal patterns of CREB activation allowing for the integration of a wide variety of stimuli.

Epigenetic regulation of the NR4A orphan nuclear receptor NOR1 by histone acetylation.

PubMed

Zhao, Yue; Nomiyama, Takashi; Findeisen, Hannes M; Qing, Hua; Aono, Jun; Jones, Karrie L; Heywood, Elizabeth B; Bruemmer, Dennis

2014-12-20

The nuclear receptor NOR1 is an immediate-early response gene implicated in the transcriptional control of proliferation. Since the expression level of NOR1 is rapidly induced through cAMP response element binding (CREB) protein-dependent promoter activation, we investigated the contribution of histone acetylation to this transient induction. We demonstrate that NOR1 transcription is induced by histone deacetylase (HDAC) inhibition and by depletion of HDAC1 and HDAC3. HDAC inhibition activated the NOR1 promoter, increased histone acetylation and augmented the recruitment of phosphorylated CREB to the promoter. Furthermore, HDAC inhibition increased Ser133 phosphorylation of CREB and augmented NOR1 protein stability. These data outline previously unrecognized mechanisms of NOR1 regulation and illustrate a key role for histone acetylation in the rapid induction of NOR1. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Hypoxic adaptation engages the CBP/CREST-induced coactivator complex of Creb-HIF-1α in transactivating murine neuroblastic glucose transporter

PubMed Central

Thamotharan, Shanthie; Raychaudhuri, Nupur; Tomi, Masatoshi; Shin, Bo-Chul

2013-01-01

We have shown in vitro a hypoxia-induced time-dependent increase in facilitative glucose transporter isoform 3 (GLUT3) expression in N2A murine neuroblasts. This increase in GLUT3 expression is partially reliant on a transcriptional increase noted in actinomycin D and cycloheximide pretreatment experiments. Transient transfection assays in N2A neuroblasts using murine glut3-luciferase reporter constructs mapped the hypoxia-induced enhancer activities to −857- to −573-bp and −203- to −177-bp regions. Hypoxia-exposed N2A nuclear extracts demonstrated an increase in HIF-1α and p-Creb binding to HRE (−828 to −824 bp) and AP-1 (−187 to −180 bp) cis-elements, respectively, in electromobility shift and supershift assays, which was confirmed by chromatin immunoprecipitation assays. In addition, the interaction of CBP with Creb and HIF-1α and CREST with CBP in hypoxia was detected by coimmunoprecipitation. Furthermore, small interference (si)RNA targeting Creb in these cells decreased endogenous Creb concentrations that reduced by twofold hypoxia-induced glut3 gene transcription. Thus, in N2A neuroblasts, phosphorylated HIF-1α and Creb mediated the hypoxia-induced increase in glut3 transcription. Coactivation by the Ca++-dependent CREST and CBP proteins may enhance cross-talk between p-Creb-AP-1 and HIF-1α/HRE of the glut3 gene. Collectively, these processes can facilitate an adaptive response to hypoxic energy depletion targeted at enhancing glucose transport and minimizing injury while fueling the proliferative potential of neuroblasts. PMID:23321477

Upregulation of suppressor of cytokine signaling 3 in microglia by cinnamic acid.

PubMed

Chakrabarti, Sudipta; Jana, Malabendu; Roy, Avik; Pahan, Kalipada

2018-05-06

Neuroinflammation plays an important role in the pathogenesis of various neurodegenerative diseases including Alzheimer's disease (AD). Suppressor of cytokine signaling 3 (SOCS3) is an anti-inflammatory molecule that suppresses cytokine signaling and inflammatory gene expression in different cells including microglia. However, pathways through which SOCS3 could be upregulated are poorly described. Cinnamic acid is a metabolite of cinnamon, a natural compound that is being widely used all over the world as a spice or flavoring agent. This study delineates the importance of cinnamic acid for the upregulation of SOCS3 in microglia. Cinnamic acid upregulated the expression of SOCS3 mRNA and protein in mouse BV-2 microglial cells in dose- and time-dependent manner. Accordingly, cinnamic acid also increased the level of SOCS3 and suppressed the expression of inducible nitric oxide synthase and proinflammatory cytokines (TNFα, IL-1β and IL-6) in LPS-stimulated BV-2 microglial cells. Similar to BV-2 microglial cells, cinnamic acid also increased the expression of SOCS3 in primary mouse microglia and astrocytes. Presence of cAMP response element in the promoter of socs3 gene, activation of cAMP response element binding (CREB) by cinnamic acid, abrogation of cinnamic acid-mediated upregulation of SOCS3 by siRNA knockdown of CREB, and the recruitment of CREB to the socs3 gene promoter by cinnamic acid suggest that cinnamic acid increases the expression of SOCS3 by CREB. These studies suggest that cinnamic acid upregulates SOCS3 via CREB pathway, which may be of importance in neuroinflammatory and neurodegenerative disorders. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Ibuprofen and lipoic acid conjugate neuroprotective activity is mediated by Ngb/Akt intracellular signaling pathway in Alzheimer's disease rat model.

PubMed

Zara, Susi; De Colli, Marianna; Rapino, Monica; Pacella, Stephanie; Nasuti, Cinzia; Sozio, Piera; Di Stefano, Antonio; Cataldi, Amelia

2013-01-01

Alzheimer's disease (AD) is a frequent form of senile dementia. Neuroglobin (Ngb) has a neuroprotective role and decreases Aβ peptide levels. Ngb, promoting Akt phosphorylation, activates cell survival involving cyclic-nucleotide response element-binding protein (CREB). A new molecule (IBU-LA) was synthetized and administered to an AD rat model to counteract AD progression. The aim of this study was to investigate the IBU-LA-mediated induction of Ngb neuroprotective and antiapoptotic activities. Brain morphology was analyzed through Bielschowsky staining, Aβ(1-40) and Ngb expression by immunohistochemistry. Akt, p-Akt, CREB and p-CREB expression was evaluated by Western blot, apoptosis through cytochrome C/Apaf 1 immunocomplex formation, and TUNEL analysis. Bielschowsky staining and Aβ(1-40) expression show few nerve connections and Aβ(1-40) expression in an Aβ sample, preserved neuronal cells and Aβ(1-40) expression lowering in an IBU sample, mostly in IBU-LA. The Ngb level decreases in Aβ samples, compared to control and IBU-LA samples. p-Akt/Akt and p-CREB/CREB ratios reveal a reduction in Aβ sample, going back to the basal level in control and IBU-LA samples. Cytochrome C/Apaf 1 co-immunoprecipitate occurs and TUNEL-positive nuclei percentage decreases in Aβ sample. Probe test performance shows an increased spatial reference memory in the IBU-LA compared to the Aβ sample; no significant differences were seen between the IBU-LA and IBU samples. This evidence reveals that IBU-LA administration has the capability to maintain a high Ngb level allowing Ngb to perform a neuroprotective and antiapoptotic role, representing a valid tool in the therapeutic strategy of AD progression. Copyright © 2013 S. Karger AG, Basel.

Activation of the ATF2/CREB-PGC-1α pathway by metformin leads to dopaminergic neuroprotection

PubMed Central

Jeong, Ga Ram; Kim, Hyojung; Jo, Minkyung; Lee, Byoung Dae; Lee, Yun Il; Jo, Areum; Park, ChiHu; Kim, Hyein; Seo, Jeongkon; Paek, Sun Ha; Lee, Yun-Song; Choi, Jeong-Yun; Lee, Yunjong; Shin, Joo-Ho

2017-01-01

Progressive dopaminergic neurodegeneration is responsible for the canonical motor deficits in Parkinson's disease (PD). The widely prescribed anti-diabetic medicine metformin is effective in preventing neurodegeneration in animal models; however, despite the significant potential of metformin for treating PD, the therapeutic effects and molecular mechanisms underlying dopaminergic neuroprotection by metformin are largely unknown. In this study, we found that metformin induced substantial proteomic changes, especially in metabolic and mitochondrial pathways in the substantia nigra (SN). Consistent with this data, metformin increased mitochondrial marker proteins in SH-SY5Y neuroblastoma cells. Mitochondrial protein expression by metformin was found to be brain region specific, with metformin increasing mitochondrial proteins in the SN and the striatum, but not the cortex. As a potential upstream regulator of mitochondria gene transcription by metformin, PGC-1α promoter activity was stimulated by metformin via CREB and ATF2 pathways. PGC-1α and phosphorylation of ATF2 and CREB by metformin were selectively increased in the SN and the striatum, but not the cortex. Finally, we showed that metformin protected dopaminergic neurons and improved dopamine-sensitive motor performance in an MPTP-induced PD animal model. Together these results suggest that the metformin-ATF2/CREB-PGC-1α pathway might be promising therapeutic target for PD. PMID:28611284

Role of CREB on heme oxygenase-1 induction in adrenal cells: involvement of the PI3K pathway.

PubMed

Astort, F; Repetto, E M; Rocha-Viegas, L; Mercau, M E; Puch, S Sanchez; Finkielstein, C V; Pecci, A; Cymeryng, C B

2016-08-01

In addition to the well-known function of ACTH as the main regulator of adrenal steroidogenesis, we have previously demonstrated its effect on the transcriptional stimulation of HO-1 expression, a component of the cellular antioxidant defense system. In agreement, we hereby demonstrate that, in adrenocortical Y1 cells, HO-1 induction correlates with a significant prevention of the generation of reactive oxygen species induced by H2O2/Fe(2+) ACTH/cAMP-dependent activation of redox-imbalanced related factors such as NRF2 or NFκB and the participation of MAPKs in this mechanism was, however, discarded based on results with specific inhibitors and reporter plasmids. We suggest the involvement of CREB in HO-1 induction by ACTH/cAMP, as transfection of cells with a dominant-negative isoform of CREB (DN-CREB-M1) decreased, while overexpression of CREB increased HO-1 protein levels. Sequence screening of the murine HO-1 promoter revealed CRE-like sites located at -146 and -37 of the transcription start site and ChIP studies indicated that this region recruits phosphorylated CREB (pCREB) upon cAMP stimulation in Y1 cells. In agreement, H89 (PKA inhibitor) or cotransfection with DN-CREB-M1 prevented the 8Br-cAMP-dependent increase in luciferase activity in cells transfected with pHO-1[-295/+74].LUC. ACTH and cAMP treatment induced the activation of the PI3K/Akt signaling pathway in a PKA-independent mechanism. Inhibition of this pathway prevented the cAMP-dependent increase in HO-1 protein levels and luciferase activity in cells transfected with pHO-1[-295/+74].LUC. Finally, here we show a crosstalk between the cAMP/PKA and PI3K pathways that affects the binding of p-CREB to its cognate element in the murine promoter of the Hmox1 gene. © 2016 Society for Endocrinology.

Interaction between C/EBPbeta and Tax down-regulates human T-cell leukemia virus type I transcription.

PubMed

Hivin, P; Gaudray, G; Devaux, C; Mesnard, J-M

2004-01-20

The human T-cell leukemia virus type I (HTLV-I) Tax protein trans-activates viral transcription through three imperfect tandem repeats of a 21-bp sequence called Tax-responsive element (TxRE). Tax regulates transcription via direct interaction with some members of the activating transcription factor/CRE-binding protein (ATF/CREB) family including CREM, CREB, and CREB-2. By interacting with their ZIP domain, Tax stimulates the binding of these cellular factors to the CRE-like sequence present in the TxREs. Recent observations have shown that CCAAT/enhancer binding protein beta (C/EBPbeta) forms stable complexes on the CRE site in the presence of CREB-2. Given that C/EBPbeta has also been found to interact with Tax, we analyzed the effects of C/EBPbeta on viral Tax-dependent transcription. We show here that C/EBPbeta represses viral transcription and that Tax is no more able to form a stable complex with CREB-2 on the TxRE site in the presence of C/EBPbeta. We also analyzed the physical interactions between Tax and C/EBPbeta and found that the central region of C/EBPbeta, excluding its ZIP domain, is required for direct interaction with Tax. It is the first time that Tax is described to interact with a basic leucine-zipper (bZIP) factor without recognizing its ZIP domain. Although unexpected, this result explains why C/EBPbeta would be unable to form a stable complex with Tax on the TxRE site and could then down-regulate viral transcription. Lastly, we found that C/EBPbeta was able to inhibit Tax expression in vivo from an infectious HTLV-I molecular clone. In conclusion, we propose that during cell activation events, which stimulate the Tax synthesis, C/EBPbeta may down-regulate the level of HTLV-I expression to escape the cytotoxic-T-lymphocyte response.

Huanglian-Jie-Du-Tang Extract Ameliorates Depression-Like Behaviors through BDNF-TrkB-CREB Pathway in Rats with Chronic Unpredictable Stress

PubMed Central

Ye, Yi-Lu; Zhong, Kai; Liu, Dan-Dan; Xu, Jing; Pan, Bei-Bei; Yu, Yue-Ping

2017-01-01

Neuroinflammation is considered as one of the common pathogeneses of depression. Huanglian-Jie-Du-Tang (HJDT) is a traditional Chinese herbal formula. The present study investigates the antidepressant-like effect of HJDT and its possible mechanism in rats. Rats were given HJDT (2, 4, and 8 g/kg, intragastrically), paroxetine (1.8 mg/kg, intragastrically), or an equivalent volume of saline for 42 days. The depression-related behaviors, including sucrose preference test (SPT), open field test (OFT), novel objective recognition task (NORT), and forced swimming test (FST), were detected. 5-Hydroxytryptamine (5-HT) and dopamine (DA) contents, microglial activation, proinflammatory cytokines, and brain derived neurotrophic factor (BDNF), tropomyosin receptor kinases B (TrkB), and cAMP-responsive element binding protein (CREB) expression were investigated. The results indicated HJDT (2 and 4 g/kg) dramatically ameliorated the depression-like behaviors. Also HJDT decreased the number of microglia and the proinflammatory cytokines in hippocampus. Western-blotting analysis displayed HJDT upregulated BDNF, TrkB, and pCREB/CREB expression in hippocampus. Particularly, pCREB DNA activity enhanced with HJDT treatment in hippocampus. But there was no difference in the 5-HT and DA contents with HJDT treatment. In conclusion, it was supposed that HJDT might be a potential Chinese medicine decoction for treating or alleviating complex symptoms of depression through BDNF-TrkB-CREB pathway. PMID:28694833

The membrane trafficking and functionality of the K+-Cl- co-transporter KCC2 is regulated by TGF-β2.

PubMed

Roussa, Eleni; Speer, Jan Manuel; Chudotvorova, Ilona; Khakipoor, Shokoufeh; Smirnov, Sergei; Rivera, Claudio; Krieglstein, Kerstin

2016-09-15

Functional activation of the neuronal K(+)-Cl(-) co-transporter KCC2 (also known as SLC12A5) is a prerequisite for shifting GABAA responses from depolarizing to hyperpolarizing during development. Here, we introduce transforming growth factor β2 (TGF-β2) as a new regulator of KCC2 membrane trafficking and functional activation. TGF-β2 controls membrane trafficking, surface expression and activity of KCC2 in developing and mature mouse primary hippocampal neurons, as determined by immunoblotting, immunofluorescence, biotinylation of surface proteins and KCC2-mediated Cl(-) extrusion. We also identify the signaling pathway from TGF-β2 to cAMP-response-element-binding protein (CREB) and Ras-associated binding protein 11b (Rab11b) as the underlying mechanism for TGF-β2-mediated KCC2 trafficking and functional activation. TGF-β2 increases colocalization and interaction of KCC2 with Rab11b, as determined by 3D stimulated emission depletion (STED) microscopy and co-immunoprecipitation, respectively, induces CREB phosphorylation, and enhances Rab11b gene expression. Loss of function of either CREB1 or Rab11b suppressed TGF-β2-dependent KCC2 trafficking, surface expression and functionality. Thus, TGF-β2 is a new regulatory factor for KCC2 functional activation and membrane trafficking, and a putative indispensable molecular determinant for the developmental shift of GABAergic transmission. © 2016. Published by The Company of Biologists Ltd.

Active CREB1 promotes a malignant TGFβ2 autocrine loop in glioblastoma.

PubMed

Rodón, Laura; Gonzàlez-Juncà, Alba; Inda, María del Mar; Sala-Hojman, Ada; Martínez-Sáez, Elena; Seoane, Joan

2014-10-01

In advanced cancer, including glioblastoma, the TGFβ pathway acts as an oncogenic factor. Some tumors exhibit aberrantly high TGFβ activity, and the mechanisms underlying this phenomenon are not well understood. We have observed that TGFβ can induce TGFβ2, generating an autocrine loop leading to aberrantly high levels of TGFβ2. We identified cAMP-responsive element-binding protein 1 (CREB1) as the critical mediator of the induction of TGFβ2 by TGFβ. CREB1 binds to the TGFB2 gene promoter in cooperation with SMAD3 and is required for TGFβ to activate transcription. Moreover, the PI3K-AKT and RSK pathways regulate the TGFβ2 autocrine loop through CREB1. The levels of CREB1 and active phosphorylated CREB1 correlate with TGFβ2 in glioblastoma. In addition, using patient-derived in vivo models of glioblastoma, we found that CREB1 levels determine the expression of TGFβ2. Our results show that CREB1 can be considered a biomarker to stratify patients for anti-TGFβ treatments and a therapeutic target in glioblastoma. TGFβ is considered a promising therapeutic target, and several clinical trials using TGFβ inhibitors are generating encouraging results. Here, we discerned the molecular mechanisms responsible for the aberrantly high levels of TGFβ2 found in certain tumors, and we propose biomarkers to predict the clinical response to anti-TGFβ therapies. ©2014 American Association for Cancer Research.

CREB1 Genotype Modulates Adaptive Reward-Based Decisions in Humans.

PubMed

Wolf, Claudia; Mohr, Holger; Diekhof, Esther K; Vieker, Henning; Goya-Maldonado, Roberto; Trost, Sarah; Krämer, Bernd; Keil, Maria; Binder, Elisabeth B; Gruber, Oliver

2016-07-01

Cyclic AMP response element-binding protein (CREB) contributes to adaptation of mesocorticolimbic networks by modulating activity-regulated transcription and plasticity in neurons. Activity or expression changes of CREB in the nucleus accumbens (NAc) and orbital frontal cortex (OFC) interact with behavioral changes during reward-motivated learning. However, these findings from animal models have not been evaluated in humans. We tested whether CREB1 genotypes affect reward-motivated decisions and related brain activation, using BOLD fMRI in 224 young and healthy participants. More specifically, participants needed to adapt their decision to either pursue or resist immediate rewards to optimize the reward outcome. We found significant CREB1 genotype effects on choices to pursue increases of the reward outcome and on BOLD signal in the NAc, OFC, insula cortex, cingulate gyrus, hippocampus, amygdala, and precuneus during these decisions in comparison with those decisions avoiding total reward loss. Our results suggest that CREB1 genotype effects in these regions could contribute to individual differences in reward- and associative memory-based decision-making. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Soybean isoflavone ameliorates β-amyloid 1-42-induced learning and memory deficit in rats by protecting synaptic structure and function.

PubMed

Ding, Juan; Xi, Yuan-Di; Zhang, Dan-Di; Zhao, Xia; Liu, Jin-Meng; Li, Chao-Qun; Han, Jing; Xiao, Rong

2013-12-01

This research aims to investigate whether soybean isoflavone (SIF) could alleviate the learning and memory deficit induced by β-amyloid peptides 1-42 (Aβ 1-42) by protecting the synapses of rats. Adult male Wistar rats were randomly allocated to the following groups: (1) control group; (2) Aβ 1-42 group; (3) SIF group; (4) SIF + Aβ 1-42 group (SIF pretreatment group) according to body weight. The 80 mg/kg/day of SIF was administered orally by gavage to the rats in SIF and SIF+Aβ 1-42 groups. Aβ 1-42 was injected into the lateral cerebral ventricle of rats in Aβ 1-42 and SIF+Aβ 1-42 groups. The ability of learning and memory, ultramicrostructure of hippocampal synapses, and expression of synaptic related proteins were investigated. The Morris water maze results showed the escape latency and total distance were decreased in the rats of SIF pretreatment group compared to the rats in Aβ1-42 group. Furthermore, SIF pretreatment could alleviate the synaptic structural damage and antagonize the down-regulation expressions of below proteins induced by Aβ1-42: (1) mRNA and protein of the synaptophysin and postsynaptic density protein 95 (PSD-95); (2) protein of calmodulin (CaM), Ca(2+) /calmodulin-dependent protein kinase II (CaMK II), and cAMP response element binding protein (CREB); (3) phosphorylation levels of CaMK II and CREB (pCAMK II, pCREB). These results suggested that SIF pretreatment could ameliorate the impairment of learning and memory ability in rats induced by Aβ 1-42, and its mechanism might be associated with the protection of synaptic plasticity by improving the synaptic structure and regulating the synaptic related proteins. Copyright © 2013 Wiley Periodicals, Inc.

Splicing factor NSSR1 reduces neuronal injury after mouse transient global cerebral ischemia.

PubMed

Qi, Yao; Li, Ya; Cui, Shi-Chao; Zhao, Jing-Jing; Liu, Xiao-Yan; Ji, Chun-Xia; Sun, Feng-Yan; Xu, Ping; Chen, Xian-Hua

2015-05-01

This study focuses on the function of NSSR1, a splicing factor, in neuronal injury in the ischemic mouse brain using the transient global cerebral ischemic mouse model and the cultured cells treated with oxygen-glucose deprivation (OGD). The results showed that the cerebral ischemia triggers the expression of NSSR1 in hippocampal astrocytes, predominantly the dephosphorylated NSSR1 proteins, and the Exon3 inclusive NCAM-L1 variant and the Exon4 inclusive CREB variant. While in the hippocampus of astrocyte-specific NSSR1 conditional knockdown (cKD) mice, where cerebral ischemia no longer triggers NSSR1 expression in astrocytes, the expression of Exon3 inclusive NCAM-L1 variant and Exon4 inclusive CREB variant were no longer triggered as well. In addition, the injury of hippocampal neurons was more severe in astrocyte-specific NSSR1 cKD mice compared with in wild-type mice after brain ischemia. Of note, the culture media harvested from the astrocytes with overexpression of NSSR1 or the Exon3 inclusive NCAM-L1 variant, or Exon4 inclusive CREB variant were all able to reduce the neuronal injury induced by OGD. The results provide the evidence demonstrating that: (1) Splicing factor NSSR1 is a new factor involved in reducing ischemic injury. (2) Ischemia induces NSSR1 expression in astrocytes, not in neurons. (3) NSSR1-mediated pathway in astrocytes is required for reducing ischemic neuronal injury. (4) NCAM-L1 and CREB are probably mediators in NSSR1-mediated pathway. In conclusion, our results suggest for the first time that NSSR1 may provide a novel mechanism for reducing neuronal injury after ischemia, probably through regulation on alternative splicing of NCAM-L1 and CREB in astrocytes. © 2014 Wiley Periodicals, Inc.

Sulforaphane suppresses lipopolysaccharide-induced cyclooxygenase-2 (COX-2) expression through the modulation of multiple targets in COX-2 gene promoter.

PubMed

Woo, Kyung Jin; Kwon, Taeg Kyu

2007-12-15

Sulforaphane is a natural, biologically active compound extracted from cruciferous vegetables such as broccoli and cabbage. It possesses potent anti-inflammation and anti-cancer properties. The mechanism by which sulforaphane suppresses COX-2 expression remains poorly understood. In the present report, we investigated the effect of sulforaphane on the expression of COX-2 in lipopolysaccharide (LPS)-activated Raw 264.7 cells. Sulforaphane significantly suppressed the LPS-induced COX-2 protein and mRNA expression in a dose-dependent manner. The ability of sulforaphane to suppress the expression of the COX-2 was investigated using luciferase reporters controlled by various cis-elements in COX-2 promoter region. Electrophoretic mobility shift assay (EMSA) verified that NF-kappaB, C/EBP, CREB and AP-1 were identified as responsible for the sulforaphane-mediated COX-2 down-regulation. In addition, we demonstrated the signal transduction pathway of mitogen-activated protein kinase (MAP kinase) in LPS-induced COX-2 expression. Taken together, these results demonstrate that sulforaphane effectively suppressed the LPS-induced COX-2 protein via modulation of multiple core promoter elements (NF-kappaB, C/EBP, CREB and AP-1) in the COX-2 transcriptional regulation. These results will provide new insights into the anti-inflammatory and anti-carcinogenic properties of sulforaphane.

Enhancement of Behavioral Sensitization, Anxiety-Like Behavior, and Hippocampal and Frontal Cortical CREB Levels Following Cocaine Abstinence in Mice Exposed to Cocaine during Adolescence

PubMed Central

Valzachi, Maria Cristina; Teodorov, Elizabeth; Marcourakis, Tania; Bailey, Alexis; Camarini, Rosana

2013-01-01

Adolescence has been linked to greater risk-taking and novelty-seeking behavior and a higher prevalence of drug abuse and risk of relapse. Decreases in cyclic adenosine monophosphate response element binding protein (CREB) and phosphorylated CREB (pCREB) have been reported after repeated cocaine administration in animal models. We compared the behavioral effects of cocaine and abstinence in adolescent and adult mice and investigated possible age-related differences in CREB and pCREB levels. Adolescent and adult male Swiss mice received one daily injection of saline or cocaine (10 mg/kg, i.p.) for 8 days. On day 9, the mice received a saline injection to evaluate possible environmental conditioning. After 9 days of withdrawal, the mice were tested in the elevated plus maze to evaluate anxiety-like behavior. Twelve days after the last saline/cocaine injection, the mice received a challenge injection of either cocaine or saline, and locomotor activity was assessed. One hour after the last injection, the brains were extracted, and CREB and pCREB levels were evaluated using Western blot in the prefrontal cortex (PFC) and hippocampus. The cocaine-pretreated mice during adolescence exhibited a greater magnitude of the expression of behavioral sensitization and greater cocaine withdrawal-induced anxiety-like behavior compared with the control group. Significant increases in CREB levels in the PFC and hippocampus and pCREB in the hippocampus were observed in cocaine-abstinent animals compared with the animals treated with cocaine in adulthood. Interestingly, significant negative correlations were observed between cocaine sensitization and CREB levels in both regions. These results suggest that the behavioral and neurochemical consequences of psychoactive substances in a still-developing nervous system can be more severe than in an already mature nervous system. PMID:24205196

Melanogenesis-inducing effect of cirsimaritin through increases in microphthalmia-associated transcription factor and tyrosinase expression.

PubMed

Kim, Hyo Jung; Kim, Il Soon; Dong, Yin; Lee, Ik-Soo; Kim, Jin Sook; Kim, Jong-Sang; Woo, Je-Tae; Cha, Byung-Yoon

2015-04-20

The melanin-inducing properties of cirsimaritin were investigated in murine B16F10 cells. Cirsimaritin is an active flavone with methoxy groups, which is isolated from the branches of Lithocarpus dealbatus. Tyrosinase activity and melanin content in murine B16F10 melanoma cells were increased by cirsimaritin in a dose-dependent manner. Western blot analysis revealed that tyrosinase, tyrosinase-related protein (TRP) 1, TRP2 protein levels were enhanced after treatment with cirsimaritin for 48 h. Cirsimaritin also upregulated the expression of microphthalmia-associated transcription factor (MITF) after 24 h of treatment. Furthermore, cirsimaritin induced phosphorylation of cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB) in a dose-dependent manner after treatment for 15 min. The cirsimaritin-mediated increase of tyrosinase activity was significantly attenuated by H89, a cAMP-dependent protein kinase A inhibitor. These findings indicate that cirsimaritin stimulates melanogenesis in B16F10 cells by activation of CREB as well as upregulation of MITF and tyrosinase expression, which was activated by cAMP signaling. Finally, the melanogenic effect of cirsimaritin was confirmed in human epidermal melanocytes. These results support the putative application of cirsimaritin in ultraviolet photoprotection and hair coloration treatments.

Osthole Enhances Osteogenesis in Osteoblasts by Elevating Transcription Factor Osterix via cAMP/CREB Signaling In Vitro and In Vivo.

PubMed

Zhang, Zhong-Rong; Leung, Wing Nang; Li, Gang; Kong, Siu Kai; Lu, Xiong; Wong, Yin Mei; Chan, Chun Wai

2017-06-08

Anabolic anti-osteoporotic agents are desirable for treatment and prevention of osteoporosis and fragility fractures. Osthole is a coumarin derivative extracted from the medicinal herbs Cnidium monnieri (L.) Cusson and Angelica pubescens Maxim.f. Osthole has been reported with osteogenic and anti-osteoporotic properties, whereas the underlying mechanism of its benefit still remains unclear. The objective of the present study was to investigate the osteopromotive action of osthole on mouse osteoblastic MC3T3-E1 cells and on mouse femoral fracture repair, and to explore the interaction between osthole-induced osteopromotive effect and cyclic adenosine monophosphate (cAMP) elevating effect. Osthole treatment promoted osteogenesis in osteoblasts by enhancing alkaline phosphatase (ALP) activity and mineralization. Oral gavage of osthole enhanced fracture repair and increased bone strength. Mechanistic study showed osthole triggered the cAMP/CREB pathway through the elevation of the intracellular cAMP level and activation of the phosphorylation of the cAMP response element-binding protein (CREB). Blockage of cAMP/CREB downstream signals with protein kinase A (PKA) inhibitor KT5720 partially suppressed osthole-mediated osteogenesis by inhibiting the elevation of transcription factor, osterix. In conclusion, osthole shows osteopromotive effect on osteoblasts in vitro and in vivo. Osthole-mediated osteogenesis is related to activation of the cAMP/CREB signaling pathway and downstream osterix expression.

Osthole Enhances Osteogenesis in Osteoblasts by Elevating Transcription Factor Osterix via cAMP/CREB Signaling In Vitro and In Vivo

PubMed Central

Zhang, Zhong-Rong; Leung, Wing Nang; Li, Gang; Kong, Siu Kai; Lu, Xiong; Wong, Yin Mei; Chan, Chun Wai

2017-01-01

Anabolic anti-osteoporotic agents are desirable for treatment and prevention of osteoporosis and fragility fractures. Osthole is a coumarin derivative extracted from the medicinal herbs Cnidium monnieri (L.) Cusson and Angelica pubescens Maxim.f. Osthole has been reported with osteogenic and anti-osteoporotic properties, whereas the underlying mechanism of its benefit still remains unclear. The objective of the present study was to investigate the osteopromotive action of osthole on mouse osteoblastic MC3T3-E1 cells and on mouse femoral fracture repair, and to explore the interaction between osthole-induced osteopromotive effect and cyclic adenosine monophosphate (cAMP) elevating effect. Osthole treatment promoted osteogenesis in osteoblasts by enhancing alkaline phosphatase (ALP) activity and mineralization. Oral gavage of osthole enhanced fracture repair and increased bone strength. Mechanistic study showed osthole triggered the cAMP/CREB pathway through the elevation of the intracellular cAMP level and activation of the phosphorylation of the cAMP response element-binding protein (CREB). Blockage of cAMP/CREB downstream signals with protein kinase A (PKA) inhibitor KT5720 partially suppressed osthole-mediated osteogenesis by inhibiting the elevation of transcription factor, osterix. In conclusion, osthole shows osteopromotive effect on osteoblasts in vitro and in vivo. Osthole-mediated osteogenesis is related to activation of the cAMP/CREB signaling pathway and downstream osterix expression. PMID:28629115

Neuroprotective effects of α-iso-cubebene against glutamate-induced damage in the HT22 hippocampal neuronal cell line.

PubMed

Park, Sun Young; Jung, Won Jung; Kang, Jum Soon; Kim, Cheol-Min; Park, Geuntae; Choi, Young-Whan

2015-02-01

Since oxidative stress is critically involved in excitotoxic damage, we sought to determine whether the activation of the transcription factors, cAMP-responsive element binding protein (CREB) and nuclear factor (erythroid-derived 2)-like 2 (Nrf2, also known as NFE2L2), by α-iso-cubebene is involved in its protective effects against glutamate-induced neuronal cell death. Pre-treatment with α-iso-cubebene significantly attenuated glutamate-induced cytotoxicity in mouse hippocampus-derived neuronal cells. α-iso-cubebene also reduced the glutamate-induced generation of reactive oxygen species and calcium influx, thus preventing apoptotic cell death. α-iso-cubebene inhibited glutamate-induced mitochondrial membrane depolarization and, consequently, inhibited the release of the apoptosis-inducing factor from the mitochondria. Immunoblot anlaysis revealed that the phosphorylation of extracellular signal-regulated kinase (ERK) by glutamate was reduced in the presence of α-iso-cubebene. α-iso-cubebene activated protein kinase A (PKA), CREB and Nrf2, which mediate the expression of the antioxidant enzymes, heme oxygenase-1 (HO-1) and NAD(P)H dehydrogenase [quinone] 1 (NQO1), involved in neuroprotection. In addition, α-iso-cubebene induced the expression of antioxidant responsive element and CRE transcriptional activity, thus conferring neuroprotection against glutamate-induced oxidative injury. α-iso-cubebene also induced the expression of Nrf2-dependent genes encoding HO-1 and NQO1. Furthermore, the knockdown of CREB and Nrf2 by small interfering RNA attenuated the neuroprotective effects of α-iso-cubebene. Taken together, our results indicate that α-iso-cubebene protects HT22 cells from glutamate-induced oxidative damage through the activation of Nrf2/HO-1/NQO-1, as well as through the PKA and CREB signaling pathways.

Modulation of cAMP levels by high-fat diet and curcumin and regulatory effects on CD36/FAT scavenger receptor/fatty acids transporter gene expression.

PubMed

Zingg, Jean-Marc; Hasan, Syeda T; Nakagawa, Kiyotaka; Canepa, Elisa; Ricciarelli, Roberta; Villacorta, Luis; Azzi, Angelo; Meydani, Mohsen

2017-01-02

Curcumin, a polyphenol from turmeric (Curcuma longa), reduces inflammation, atherosclerosis, and obesity in several animal studies. In Ldlr -/- mice fed a high-fat diet (HFD), curcumin reduces plasma lipid levels, therefore contributing to a lower accumulation of lipids and to reduced expression of fatty acid transport proteins (CD36/FAT, FABP4/aP2) in peritoneal macrophages. In this study, we analyzed the molecular mechanisms by which curcumin (500, 1000, 1500 mg/kg diet, for 4 months) may influence plasma and tissue lipid levels in Ldlr -/- mice fed an HFD. In liver, HFD significantly suppressed cAMP levels, and curcumin restored almost normal levels. Similar trends were observed in adipose tissues, but not in brain, skeletal muscle, spleen, and kidney. Treatment with curcumin increased phosphorylation of CREB in liver, what may play a role in regulatory effects of curcumin in lipid homeostasis. In cell lines, curcumin increased the level of cAMP, activated the transcription factor CREB and the human CD36 promoter via a sequence containing a consensus CREB response element. Regulatory effects of HFD and Cur on gene expression were observed in liver, less in skeletal muscle and not in brain. Since the cAMP/protein kinase A (PKA)/CREB pathway plays an important role in lipid homeostasis, energy expenditure, and thermogenesis by increasing lipolysis and fatty acid β-oxidation, an increase in cAMP levels induced by curcumin may contribute to its hypolipidemic and anti-atherosclerotic effects. © 2016 BioFactors, 43(1):42-53, 2017. © 2016 International Union of Biochemistry and Molecular Biology.

Fisetin Ameliorated Photodamage by Suppressing the Mitogen-Activated Protein Kinase/Matrix Metalloproteinase Pathway and Nuclear Factor-κB Pathways.

PubMed

Chiang, Hsiu-Mei; Chan, Shih-Yun; Chu, Yin; Wen, Kuo-Ching

2015-05-13

Ultraviolet (UV) irradiation is one of the most important extrinsic factors contributing to skin photodamage. After UV irradiation, a series of signal transductions in the skin will be activated, leading to inflammatory response and photoaged skin. In this study, fisetin, a flavonol that exists in fruits and vegetables, was investigated for its photoprotective effects. The results revealed that 5-25 μM fisetin inhibits cyclooxygenase-2 (COX-2) and matrix metalloproteinase (MMP)-1, MMP-3, MMP-9 expression induced by ultraviolet B (UVB) irradiation in human skin fibroblasts. In addition, fisetin suppressed UVB-induced collagen degradation. With regard to its effect on upper-stream signal transduction, we found that fisetin reduced the expression of ultraviolet (UV)-induced ERK, JNK, and p38 phosphorylation in the mitogen-activated protein kinase (MAP kinase) pathway. Furthermore, fisetin reduced inhibitor κB (IκB) degradation and increased the amount of p65, which is a major subunit of nuclear factor-κB (NF-κB), in cytoplasm. It also suppressed NF-κB translocated to the nucleus and inhibited cAMP response element-binding protein (CREB) Ser-133 phosphorylation level in the phosphoinositide 3-kinase/protein kinase B/CREB (PI3K/AKT/CREB) pathway. Finally, fisetin inhibited UV-induced intracellular reactive oxygen species (ROS), prostaglandin E2 (PGE2), and nitric oxide (NO) generation. The mentioned effects and mechanisms suggest that fisetin can be used in the development of photoprotective agents.

Chronic copper exposure causes spatial memory impairment, selective loss of hippocampal synaptic proteins, and activation of PKR/eIF2α pathway in mice.

PubMed

Ma, Quan; Ying, Ming; Sui, Xiaojing; Zhang, Huimin; Huang, Haiyan; Yang, Linqing; Huang, Xinfeng; Zhuang, Zhixiong; Liu, Jianjun; Yang, Xifei

2015-01-01

Copper is an essential element for human growth and development; however, excessive intake of copper could contribute to neurotoxicity. Here we show that chronic exposure to copper in drinking water impaired spatial memory with simultaneous selective loss of hippocampal pre-synaptic protein synapsin 1, and post-synaptic density protein (PSD)-93/95 in mice. Copper exposure was shown to elevate the levels of nitrotyrosine and 8-hydroxydeoxyguanosine (8-OHdG) in hippocampus, two markers of oxidative stress. Concurrently, we also found that copper exposure activated double stranded RNA-dependent protein kinase (PKR) as evidenced by increased ratio of phosphorylated PKR at Thr451 and total PKR and increased the phosphorylation of its downstream signaling molecule eukaryotic initiation factor 2α (eIF2α) at Ser51 in hippocampus. Consistent with activation of PKR/eIF2α signaling pathway which was shown to mediate synaptic deficit and cognitive impairment, the levels of activating transcription factor 4 (ATF-4), a downstream signaling molecule of eIF2α and a repressor of CREB-mediated gene expression, were significantly increased, while the activity of cAMP response elements binding protein (CREB) was inactivated as suggested by decreased phosphorylation of CREB at Ser133 by copper exposure. In addition, the expression of the pro-apoptotic target molecule C/EBP homology protein (CHOP) of ATF-4 was upregulated and hippocampal neuronal apoptosis was induced by copper exposure. Taken together, we propose that chronic copper exposure might cause spatial memory impairment, selective loss of synaptic proteins, and neuronal apoptosis through the mechanisms involving activation of PKR/eIF2α signaling pathway.

In vitro selection of DNA elements highly responsive to the human T-cell lymphotropic virus type I transcriptional activator, Tax.

PubMed

Paca-Uccaralertkun, S; Zhao, L J; Adya, N; Cross, J V; Cullen, B R; Boros, I M; Giam, C Z

1994-01-01

The human T-cell lymphotropic virus type I (HTLV-I) transactivator, Tax, the ubiquitous transcriptional factor cyclic AMP (cAMP) response element-binding protein (CREB protein), and the 21-bp repeats in the HTLV-I transcriptional enhancer form a ternary nucleoprotein complex (L. J. Zhao and C. Z. Giam, Proc. Natl. Acad. Sci. USA 89:7070-7074, 1992). Using an antibody directed against the COOH-terminal region of Tax along with purified Tax and CREB proteins, we selected DNA elements bound specifically by the Tax-CREB complex in vitro. Two distinct but related groups of sequences containing the cAMP response element (CRE) flanked by long runs of G and C residues in the 5' and 3' regions, respectively, were preferentially recognized by Tax-CREB. In contrast, CREB alone binds only to CRE motifs (GNTGACG[T/C]) without neighboring G- or C-rich sequences. The Tax-CREB-selected sequences bear a striking resemblance to the 5' or 3' two-thirds of the HTLV-I 21-bp repeats and are highly inducible by Tax. Gel electrophoretic mobility shift assays, DNA transfection, and DNase I footprinting analyses indicated that the G- and C-rich sequences flanking the CRE motif are crucial for Tax-CREB-DNA ternary complex assembly and Tax transactivation but are not in direct contact with the Tax-CREB complex. These data show that Tax recruits CREB to form a multiprotein complex that specifically recognizes the viral 21-bp repeats. The expanded DNA binding specificity of Tax-CREB and the obligatory role the ternary Tax-CREB-DNA complex plays in transactivation reveal a novel mechanism for regulating the transcriptional activity of leucine zipper proteins like CREB.

HTLV-1 Tax protein cooperates with Ras in protecting cells from apoptosis.

PubMed

Vajente, Nicola; Trevisan, Roberta; Saggioro, Daniela

2009-02-01

Tax protein of the human T-cell leukemia virus type 1 (HTLV-1) plays a critical role in HTLV-I-correlated diseases through its ability to deregulate the expression of a vast array of cellular genes. We have previously shown that Tax counteracts apoptosis induced by stimuli triggering mitochondria apoptotic pathway, most likely by activating CREB-mediated transcription and affecting the phosphorylation levels of CREB at Ser-133. Here, we report data that indicate the oncoprotein Ras as a possible mediator of Tax-induced apoptosis protection and suggest a possible role of Tax in Ras activation. In addition, using inhibitors of down stream effectors of Ras, we found that ERK signaling is the most relevant for Tax-mediated apoptosis protection. As a whole, our findings provide intriguing evidence of a possible link between Ras signaling and Tax capability to counteract apoptosis and to enhance P-CREB levels, and implicates a potential role for Ras in HTLV-1-induced diseases.

A Positive Autoregulatory BDNF Feedback Loop via C/EBPβ Mediates Hippocampal Memory Consolidation

PubMed Central

Bambah-Mukku, Dhananjay; Travaglia, Alessio; Chen, Dillon Y.; Pollonini, Gabriella

2014-01-01

Little is known about the temporal progression and regulation of the mechanisms underlying memory consolidation. Brain-derived-neurotrophic-factor (BDNF) has been shown to mediate the maintenance of memory consolidation, but the mechanisms of this regulation remain unclear. Using inhibitory avoidance (IA) in rats, here we show that a hippocampal BDNF-positive autoregulatory feedback loop via CCAAT-enhancer binding protein β (C/EBPβ) is necessary to mediate memory consolidation. At training, a very rapid, learning-induced requirement of BDNF accompanied by rapid de novo translation controls the induction of a persistent activation of cAMP-response element binding-protein (CREB) and C/EBPβ expression. The latter, in turn, controls an increase in expression of bdnf exon IV transcripts and BDNF protein, both of which are necessary and, together with the initial BDNF requirement, mediate memory consolidation. The autoregulatory loop terminates by 48 h after training with decreased C/EBPβ and pCREB and increased methyl-CpG binding protein-2, histone-deacetylase-2, and switch-independent-3a binding at the bdnf exon IV promoter. PMID:25209292

Improved α-Amylase Production by Dephosphorylation Mutation of CreD, an Arrestin-Like Protein Required for Glucose-Induced Endocytosis of Maltose Permease and Carbon Catabolite Derepression in Aspergillus oryzae

PubMed Central

Tanaka, Mizuki; Hiramoto, Tetsuya; Tada, Hinako; Shintani, Takahiro

2017-01-01

ABSTRACT Aspergillus oryzae produces copious amount of amylolytic enzymes, and MalP, a major maltose permease, is required for the expression of amylase-encoding genes. The expression of these genes is strongly repressed by carbon catabolite repression (CCR) in the presence of glucose. MalP is transported from the plasma membrane to the vacuole by endocytosis, which requires the homolog of E6-AP carboxyl terminus ubiquitin ligase HulA, an ortholog of yeast Rsp5. In yeast, arrestin-like proteins mediate endocytosis as adaptors of Rsp5 and transporters. In the present study, we examined the involvement of CreD, an arrestin-like protein, in glucose-induced MalP endocytosis and CCR of amylase-encoding genes. Deletion of creD inhibited the glucose-induced endocytosis of MalP, and CreD showed physical interaction with HulA. Phosphorylation of CreD was detected by Western blotting, and two serine residues were determined as the putative phosphorylation sites. However, the phosphorylation state of the serine residues did not regulate MalP endocytosis and its interaction with HulA. Although α-amylase production was significantly repressed by creD deletion, both phosphorylation and dephosphorylation mimics of CreD had a negligible effect on α-amylase activity. Interestingly, dephosphorylation of CreD was required for CCR relief of amylase genes that was triggered by disruption of the deubiquitinating enzyme-encoding gene creB. The α-amylase activity of the creB mutant was 1.6-fold higher than that of the wild type, and the dephosphorylation mimic of CreD further improved the α-amylase activity by 2.6-fold. These results indicate that a combination of the dephosphorylation mutation of CreD and creB disruption increased the production of amylolytic enzymes in A. oryzae. IMPORTANCE In eukaryotes, glucose induces carbon catabolite repression (CCR) and proteolytic degradation of plasma membrane transporters via endocytosis. Glucose-induced endocytosis of transporters is mediated by their ubiquitination, and arrestin-like proteins act as adaptors of transporters and ubiquitin ligases. In this study, we showed that CreD, an arrestin-like protein, is involved in glucose-induced endocytosis of maltose permease and carbon catabolite derepression of amylase gene expression in Aspergillus oryzae. Dephosphorylation of CreD was required for CCR relief triggered by the disruption of creB, which encodes a deubiquitinating enzyme; a combination of the phosphorylation-defective mutation of CreD and creB disruption dramatically improved α-amylase production. This study shows the dual function of an arrestin-like protein and provides a novel approach for improving the production of amylolytic enzymes in A. oryzae. PMID:28455339

Improved α-Amylase Production by Dephosphorylation Mutation of CreD, an Arrestin-Like Protein Required for Glucose-Induced Endocytosis of Maltose Permease and Carbon Catabolite Derepression in Aspergillus oryzae.

PubMed

Tanaka, Mizuki; Hiramoto, Tetsuya; Tada, Hinako; Shintani, Takahiro; Gomi, Katsuya

2017-07-01

Aspergillus oryzae produces copious amount of amylolytic enzymes, and MalP, a major maltose permease, is required for the expression of amylase-encoding genes. The expression of these genes is strongly repressed by carbon catabolite repression (CCR) in the presence of glucose. MalP is transported from the plasma membrane to the vacuole by endocytosis, which requires the homolog of E6-AP carboxyl terminus ubiquitin ligase HulA, an ortholog of yeast Rsp5. In yeast, arrestin-like proteins mediate endocytosis as adaptors of Rsp5 and transporters. In the present study, we examined the involvement of CreD, an arrestin-like protein, in glucose-induced MalP endocytosis and CCR of amylase-encoding genes. Deletion of creD inhibited the glucose-induced endocytosis of MalP, and CreD showed physical interaction with HulA. Phosphorylation of CreD was detected by Western blotting, and two serine residues were determined as the putative phosphorylation sites. However, the phosphorylation state of the serine residues did not regulate MalP endocytosis and its interaction with HulA. Although α-amylase production was significantly repressed by creD deletion, both phosphorylation and dephosphorylation mimics of CreD had a negligible effect on α-amylase activity. Interestingly, dephosphorylation of CreD was required for CCR relief of amylase genes that was triggered by disruption of the deubiquitinating enzyme-encoding gene creB The α-amylase activity of the creB mutant was 1.6-fold higher than that of the wild type, and the dephosphorylation mimic of CreD further improved the α-amylase activity by 2.6-fold. These results indicate that a combination of the dephosphorylation mutation of CreD and creB disruption increased the production of amylolytic enzymes in A. oryzae IMPORTANCE In eukaryotes, glucose induces carbon catabolite repression (CCR) and proteolytic degradation of plasma membrane transporters via endocytosis. Glucose-induced endocytosis of transporters is mediated by their ubiquitination, and arrestin-like proteins act as adaptors of transporters and ubiquitin ligases. In this study, we showed that CreD, an arrestin-like protein, is involved in glucose-induced endocytosis of maltose permease and carbon catabolite derepression of amylase gene expression in Aspergillus oryzae Dephosphorylation of CreD was required for CCR relief triggered by the disruption of creB , which encodes a deubiquitinating enzyme; a combination of the phosphorylation-defective mutation of CreD and creB disruption dramatically improved α-amylase production. This study shows the dual function of an arrestin-like protein and provides a novel approach for improving the production of amylolytic enzymes in A. oryzae . Copyright © 2017 American Society for Microbiology.

Hippocampal Focal Knockout of CBP Affects Specific Histone Modifications, Long-Term Potentiation, and Long-Term Memory

PubMed Central

Barrett, Ruth M; Malvaez, Melissa; Kramar, Eniko; Matheos, Dina P; Arrizon, Abraham; Cabrera, Sara M; Lynch, Gary; Greene, Robert W; Wood, Marcelo A

2011-01-01

To identify the role of the histone acetyltransferase (HAT) CREB-binding protein (CBP) in neurons of the CA1 region of the hippocampus during memory formation, we examine the effects of a focal homozygous knockout of CBP on histone modifications, gene expression, synaptic plasticity, and long-term memory. We show that CBP is critical for the in vivo acetylation of lysines on histones H2B, H3, and H4. CBP's homolog p300 was unable to compensate for the loss of CBP. Neurons lacking CBP maintained phosphorylation of the transcription factor CREB, yet failed to activate CREB:CBP-mediated gene expression. Loss of CBP in dorsal CA1 of the hippocampus resulted in selective impairments to long-term potentiation and long-term memory for contextual fear and object recognition. Together, these results suggest a necessary role for specific chromatin modifications, selectively mediated by CBP in the consolidation of memories. PMID:21508930

Antitumorigenic Effects of ZAKβ, an Alternative Splicing Isoform of ZAK.

PubMed

Lee, Jin-Sun; Lin, Yuh-Yih; Wang, Tsu-Shing; Liu, Jer-Yuh; Lin, Wei-Wen; Yang, Jaw-Ji

2018-02-28

Sterile alpha motif (SAM)- and leucine-zipper-containing kinase (ZAK) plays a role in the regulation of cell cycle progression and oncogenic transformation. The ZAK gene generates two transcript variants, ZAKα and ZAKβ, through alternative splicing. In this study, we identified that ZAKα proteins were upregulated in tumor tissues, whereas ZAKβ proteins were mostly expressed in corresponding normal tissues. The ectopically expressed ZAKβ proteins in cancer cells inhibited cancer cell proliferation as well as anchorage-independent growth. The ZAKβ:ZAKα protein ratio played a role in the regulation of the cyclic adenosine monophosphate (cAMP) signaling pathway, whereas high ZAKβ protein levels led to the activation of cAMP response element binding protein 1 (CREB1) and exerted antitumor properties. Overexpression of ZAKβ or CREB1 cDNAs in cancer cells inhibited anchorage-independent growth and also reduced the levels of cyclooxygenase 2 (Cox2) and β-catenin proteins. Cancer cells treated with doxorubicin (Doxo) resulted in the switching from the expression of ZAKα to ZAKβ and also inhibited cancer cell growth in soft agar, demonstrating that pharmacological drugs could be used to manipulate endogenous reprogramming splicing events and resulting in the activation of endogenous antitumorigenic properties. We showed that the two ZAK transcript variants, ZAKα and ZAKβ, had opposite biological functions in the regulation of tumor cell proliferation in that ZAKβ had powerful antitumor properties and that ZAKα could promote tumor growth.

Niclosamide suppresses acute myeloid leukemia cell proliferation through inhibition of CREB-dependent signaling pathways

PubMed Central

Chae, Hee-Don; Cox, Nick; Dahl, Gary V.; Lacayo, Norman J.; Davis, Kara L.; Capolicchio, Samanta; Smith, Mark; Sakamoto, Kathleen M.

2018-01-01

CREB (cAMP Response Element Binding protein) is a transcription factor that is overexpressed in primary acute myeloid leukemia (AML) cells and associated with a decreased event-free survival and increased risk of relapse. We recently reported a small molecule inhibitor of CREB, XX-650-23, which inhibits CREB activity in AML cells. Structure-activity relationship analysis for chemical compounds with structures similar to XX-650-23 led to the identification of the anthelminthic drug niclosamide as a potent anti-leukemic agent that suppresses cell viability of AML cell lines and primary AML cells without a significant decrease in colony forming activity of normal bone marrow cells. Niclosamide significantly inhibited CREB function and CREB-mediated gene expression in cells, leading to apoptosis and G1/S cell cycle arrest with reduced phosphorylated CREB levels. CREB knockdown protected cells from niclosamide treatment-mediated cytotoxic effects. Furthermore, treatment with a combination of niclosamide and CREB inhibitor XX-650-23 showed an additive anti-proliferative effect, consistent with the hypothesis that niclosamide and XX-650-23 regulate the same targets or pathways to inhibit proliferation and survival of AML cells. Niclosamide significantly inhibited the progression of disease in AML patient-derived xenograft (PDX) mice, and prolonged survival of PDX mice. Niclosamide also showed synergistic effects with chemotherapy drugs to inhibit AML cell proliferation. While chemotherapy antagonized the cytotoxic potential of niclosamide, pretreatment with niclosamide sensitized cells to chemotherapeutic drugs, cytarabine, daunorubicin, and vincristine. Therefore, our results demonstrate niclosamide as a potential drug to treat AML by inducing apoptosis and cell cycle arrest through inhibition of CREB-dependent pathways in AML cells. PMID:29435104

The anthocyanin cyanidin-3-O-β-glucoside, a flavonoid, increases hepatic glutathione synthesis and protects hepatocytes against reactive oxygen species during hyperglycemia: Involvement of a cAMP-PKA-dependent signaling pathway.

PubMed

Zhu, Wei; Jia, Qianju; Wang, Yun; Zhang, Yuhua; Xia, Min

2012-01-15

Enhanced oxidative stress due to high glucose contributes to pathological changes in diabetes-related liver complications. Reducing oxidative stress may alleviate these pathogenic processes. Anthocyanin, a natural antioxidant, has been reported to reduce intracellular reactive oxygen species (ROS) levels but the mechanism of this reduction is not fully understood. The glutathione (GSH) antioxidant system is critical for counteracting oxidative stress-induced intracellular injury. In this study, we evaluated the mechanism of the anthocyanin-mediated regulation of GSH synthesis and reduction in intracellular ROS levels. We observed that treatment of human HepG2 cells with the anthocyanin C3G significantly reduced ROS levels induced by high glucose. C3G incubation increased glutamate-cysteine ligase expression, which in turn mediated the reduction in ROS levels. However, the upregulation of glutamate-cysteine ligase catalytic subunit (Gclc) expression by C3G occurred independent of the Nrf1/2 transcription factors. Notably, the cAMP-response element binding protein (CREB) was identified as the target transcription factor involved in the C3G-mediated upregulation of Gclc expression. C3G increased phosphorylation of CREB through protein kinase A (PKA) activation, which induced a CREB-mediated upregulation of Gclc transcription. In vivo, treatment with C3G increased the GSH synthesis in the liver of diabetic db/db mice through PKA-CREB-dependent induction of Gclc expression. Finally, oxidative stress determined by lipid peroxidation, neutrophil infiltration, and hepatic steatosis was attenuated in C3G-treated db/db mice. Our results demonstrate that the anthocyanin C3G has an effect of activating GSH synthesis through a novel antioxidant defense mechanism against excessive ROS production, contributing to the prevention of hyperglycemia-induced hepatic oxidative damage. Crown Copyright © 2011. Published by Elsevier Inc. All rights reserved.

Topiramate via NMDA, AMPA/kainate, GABAA and Alpha2 receptors and by modulation of CREB/BDNF and Akt/GSK3 signaling pathway exerts neuroprotective effects against methylphenidate-induced neurotoxicity in rats.

PubMed

Motaghinejad, Majid; Motevalian, Manijeh; Fatima, Sulail; Beiranvand, Tabassom; Mozaffari, Shiva

2017-11-01

Chronic abuse of methylphenidate (MPH) often causes neuronal cell death. Topiramate (TPM) carries neuroprotective effects, but its exact mechanism of action remains unclear. In the present study, the role of various doses of TPM and its possible mechanisms, receptors and signaling pathways involved against MPH-induced hippocampal neurodegeneration were evaluated in vivo. Thus, domoic acid (DOM) was used as AMPA/kainate receptor agonist, bicuculline (BIC) as GABA A receptor antagonist, ketamine (KET) as NMDA receptor antagonist, yohimbine (YOH) as α 2 adrenergic receptor antagonist and haloperidol (HAL) was used as dopamine D 2 receptor antagonist. Open field test (OFT) was used to investigate the disturbances in motor activity. Hippocampal neurodegenerative parameters were evaluated. Protein expressions of CREB/BDNF and Akt/GSK3 signaling pathways were also evaluated. Cresyl violet staining was performed to show and confirm the changes in the shape of the cells. TPM (70 and 100 mg/kg) reduced MPH-induced rise in lipid peroxidation, oxidized form of glutathione (GSSG), IL-1β and TNF-α levels, Bax expression and motor activity disturbances. In addition, TPM treatment increased Bcl-2 expression, the level of reduced form of glutathione (GSH) and the levels and activities of superoxide dismutase, glutathione peroxidase and glutathione reductase enzymes. TPM also inhibited MPH-induced hippocampal degeneration. Pretreatment of animals with DOM, BIC, KET and YOH inhibited TPM-induced neuroprotection and increased oxidative stress, neuroinflammation, neuroapoptosis and neurodegeneration while reducing CREB, BDNF and Akt protein expressions. Also pretreatment with DOM, BIC, KET and YOH inhibited TPM-induced decreases in GSK3. It can be concluded that the mentioned receptors by modulation of CREB/BDNF and Akt/GSK3 pathways, are involved in neuroprotection of TPM against MPH-induced neurodegeneration.

Protein Phosphatase-1 Inhibitor-2 Is a Novel Memory Suppressor.

PubMed

Yang, Hongtian; Hou, Hailong; Pahng, Amanda; Gu, Hua; Nairn, Angus C; Tang, Ya-Ping; Colombo, Paul J; Xia, Houhui

2015-11-11

Reversible phosphorylation, a fundamental regulatory mechanism required for many biological processes including memory formation, is coordinated by the opposing actions of protein kinases and phosphatases. Type I protein phosphatase (PP1), in particular, has been shown to constrain learning and memory formation. However, how PP1 might be regulated in memory is still not clear. Our previous work has elucidated that PP1 inhibitor-2 (I-2) is an endogenous regulator of PP1 in hippocampal and cortical neurons (Hou et al., 2013). Contrary to expectation, our studies of contextual fear conditioning and novel object recognition in I-2 heterozygous mice suggest that I-2 is a memory suppressor. In addition, lentiviral knock-down of I-2 in the rat dorsal hippocampus facilitated memory for tasks dependent on the hippocampus. Our data indicate that I-2 suppresses memory formation, probably via negatively regulating the phosphorylation of cAMP/calcium response element-binding protein (CREB) at serine 133 and CREB-mediated gene expression in dorsal hippocampus. Surprisingly, the data from both biochemical and behavioral studies suggest that I-2, despite its assumed action as a PP1 inhibitor, is a positive regulator of PP1 function in memory formation. We found that inhibitor-2 acts as a memory suppressor through its positive functional influence on type I protein phosphatase (PP1), likely resulting in negative regulation of cAMP/calcium response element-binding protein (CREB) and CREB-activated gene expression. Our studies thus provide an interesting example of a molecule with an in vivo function that is opposite to its in vitro function. PP1 plays critical roles in many essential physiological functions such as cell mitosis and glucose metabolism in addition to its known role in memory formation. PP1 pharmacological inhibitors would thus not be able to serve as good therapeutic reagents because of its many targets. However, identification of PP1 inhibitor-2 as a critical contributor to suppression of memory formation by PP1 may provide a novel therapeutic target for memory-related diseases. Copyright © 2015 the authors 0270-6474/15/3515082-06$15.00/0.

Spatial Memory in the Morris Water Maze and Activation of Cyclic AMP Response Element-Binding (CREB) Protein within the Mouse Hippocampus

ERIC Educational Resources Information Center

Porte, Yves; Buhot, Marie Christine; Mons, Nicole E.

2008-01-01

We investigated the spatio-temporal dynamics of learning-induced cAMP response element-binding protein activation/phosphorylation (pCREB) in mice trained in a spatial reference memory task in the water maze. Using immunohistochemistry, we examined pCREB immunoreactivity (pCREB-ir) in hippocampal CA1 and CA3 and related brain structures. During the…

Forskolin increases angiogenesis through the coordinated cross-talk of PKA-dependent VEGF expression and Epac-mediated PI3K/Akt/eNOS signaling.

PubMed

Namkoong, Seung; Kim, Chun-Ki; Cho, Young-Lai; Kim, Ji-Hee; Lee, Hansoo; Ha, Kwon-Soo; Choe, Jongseon; Kim, Pyeung-Hyeun; Won, Moo-Ho; Kwon, Young-Geun; Shim, Eun Bo; Kim, Young-Myeong

2009-06-01

Forskolin, a potent activator of adenylyl cyclases, has been implicated in modulating angiogenesis, but the underlying mechanism has not been clearly elucidated. We investigated the signal mechanism by which forskolin regulates angiogenesis. Forskolin stimulated angiogenesis of human endothelial cells and in vivo neovascularization, which was accompanied by phosphorylation of CREB, ERK, Akt, and endothelial nitric oxide synthase (eNOS) as well as NO production and VEGF expression. Forskolin-induced CREB phosphorylation, VEGF promoter activity, and VEGF expression were blocked by the PKA inhibitor PKI.Moreover, phosphorylation of ERK by forskolin was inhibited by the MEK inhibitor PD98059, but not PKI. The forskolin-induced Akt/eNOS/NO pathway was completely inhibited by the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002, but not significantly suppressed by PKI. These inhibitors and a NOS inhibitor partially inhibited forskolin-induced angiogenesis. The exchange protein directly activated by cAMP (Epac) activator, 8CPT-2Me-cAMP, promoted the Akt/eNOS/NO pathway and ERK phosphorylation,but did not induce CREB phosphorylation and VEGF expression. The angiogenic effect of the Epac activator was diminished by the inhibition of PI3K and MEK, but not by the PKA inhibitor. Small interfering RNA-mediated knockdown of Epac1 suppressed forskolin-induced angiogenesis and phosphorylation of ERK, Akt, and eNOS, but not CREB phosphorylation and VEGF expression. These results suggest that forskolin stimulates angiogenesis through coordinated cross-talk between two distinct pathways, PKA-dependent VEGF expression and Epac-dependent ERKactivation and PI3K/Akt/eNOS/NO signaling.

Tau hyperphosphorylation and P-CREB reduction are involved in acrylamide-induced spatial memory impairment: Suppression by curcumin.

PubMed

Yan, Dandan; Yao, Jianling; Liu, Ying; Zhang, Xing; Wang, Yiqi; Chen, Xiaoyi; Liu, Liegang; Shi, Nian; Yan, Hong

2018-04-26

Acrylamide (ACR) is an axonal toxicant that produces peripheral neuropathy in laboratory animals and humans. Epidemiological study found that diet ACR exposure was associated with a mild cognitive decline in men. However, limited information is available as regards its potential and underlying mechanism to cause memory alterations. Curcumin is a polyphenol with neuroprotective and cognitive-enhancing properties. In this study, we aimed to investigate the mechanism of ACR-induced spatial memory impairment and the beneficial effect of curcumin. ACR exposure at 10 mg/kg/d for 7 weeks caused slight gait abnormality and spatial memory deficits, which was associated with an activation of glial cells, a reduction of phosphorylated cAMP response elements binding protein (P-CREB) and an aggregation of hyperphosphorylated tau including p-tau (Ser 262 ), AT8 (p-tau Ser 202 /Thr 205 ) and PHF1 (p-tau Ser 396/404 ) in the hippocampus and cortex. ACR markedly regulate the expression of glycogen synthase kinase-3β (GSK-3β) and cyclin-dependent kinase-5 (cdk5) to accelerate tau hyperphosphorylation. ACR inhibited the protein phosphatase 2A (PP2A) and lysosomal protease cathepsin D to decrease the p-tau dephosphorylation and degradation. The P-CREB and brain derived neurotrophic factor (BDNF) were significantly decreased by ACR. The upstream signalings of P-CREB, extracellular signal-related kinase (ERK) and Akt were markedly inhibited. The protein kinase RNA-like endoplasmic reticulum kinase (PERK) -eukaryotic initiation factor-2α (eIF2α) - activating transcription factor 4 (ATF4) signaling which negatively regulate memory processes by suppressing CREB was activated by ACR. Curcumin alleviated ACR-induced spatial memory impairment through reversing tau abnormalities and P-CREB reduction in the hippocampus. These results offered deeper insight into the mechanisms of and presented a potential new treatment for ACR-induced neurotoxicity. Copyright © 2018 Elsevier Inc. All rights reserved.

Curcumin Upregulates Antioxidant Defense, Lon Protease, and Heat-Shock Protein 70 Under Hyperglycemic Conditions in Human Hepatoma Cells.

PubMed

Gounden, Shivona; Chuturgoon, Anil

2017-05-01

Sirtuin 3 (SIRT3) regulates mitochondrial antioxidant (AO) defense and improves mitochondrial disorders. Curcumin protects mitochondria; however, the mechanisms need investigation. We postulated that curcumin increases AO defense under hyperglycemic conditions in HepG2 cells through SIRT3-mediated mechanisms. Cell viability was determined in HepG2 cells cultured with 5 mM glucose, 19.9 mM mannitol, vehicle control, 10 mM glucose, and 30 mM glucose in the absence or presence of curcumin for 24 h. SIRT3, nuclear factor-kappa B (NF-κB), heat-shock protein 70 (Hsp70), and Lon protein expressions were determined using western blot. Transcript levels of SIRT3, peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α), cAMP response element-binding protein (CREB), glutathione peroxidase 1 (GPx1), and superoxide dismutase 2 (SOD2) were measured by quantitative polymerase chain reaction. Cell viability, SIRT3 protein expression, transcript levels of SIRT3, PGC-1α, CREB, GPx1, and SOD2 and protein expressions of NF-κB, Lon, and Hsp70 were significantly increased in the curcumin-treated hyperglycemic groups. Since curcumin and SIRT3 both improve mitochondrial function and AO defense, SIRT3 may be involved in the protective effects of curcumin.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cherukuri, Durga Prasad; Chen, Xiao B.O.; Goulet, Anne-Christine

Accumulating evidence indicates that elevated levels of prostaglandin E{sub 2} (PGE{sub 2}) can increase intestinal epithelial cell proliferation, and thus play a role in colorectal tumorigenesis. PGE{sub 2} exerts its effects through four G-protein-coupled PGE receptor (EP) subtypes, named the EP1, EP2, EP3, and EP4. Increased phosphorylation of extracellular regulated kinases (ERK1/2) is required for PGE{sub 2} to stimulate cell proliferation of human colon cancer cells. However, the EP receptor(s) that are involved in this process remain unknown. We provide evidence that L-161,982, a selective EP4 receptor antagonist, completely blocks PGE{sub 2}-induced ERK phosphorylation and cell proliferation of HCA-7 cells.more » In order to identify downstream target genes of ERK1/2 signaling, we found that PGE{sub 2} induces expression of early growth response gene-1 (EGR-1) downstream of ERK1/2 and regulates its expression at the level of transcription. PGE{sub 2} treatment induces phosphorylation of cyclic AMP response element binding protein (CREB) at Ser133 residue and CRE-mediated luciferase activity in HCA-7 cells. Studies with dominant-negative CREB mutant (ACREB) provide clear evidence for the involvement of CREB in PGE{sub 2} driven egr-1 transcription in HCA-7 cells. In conclusion, this study reveals that egr-1 is a target gene of PGE{sub 2} in HCA-7 cells and is regulated via the newly identified EP4/ERK/CREB pathway. Finally our results support the notion that antagonizing EP4 receptors may provide a novel therapeutic approach to the treatment of colon cancer.« less

Involvement of phosphorylated Apis mellifera CREB in gating a honeybee's behavioral response to an external stimulus

PubMed Central

Gehring, Katrin B.; Heufelder, Karin; Feige, Janina; Bauer, Paul; Dyck, Yan; Ehrhardt, Lea; Kühnemund, Johannes; Bergmann, Anja; Göbel, Josefine; Isecke, Marlene

2016-01-01

The transcription factor cAMP-response element-binding protein (CREB) is involved in neuronal plasticity. Phosphorylation activates CREB and an increased level of phosphorylated CREB is regarded as an indicator of CREB-dependent transcriptional activation. In honeybees (Apis mellifera) we recently demonstrated a particular high abundance of the phosphorylated honeybee CREB homolog (pAmCREB) in the central brain and in a subpopulation of mushroom body neurons. We hypothesize that these high pAmCREB levels are related to learning and memory formation. Here, we tested this hypothesis by analyzing brain pAmCREB levels in classically conditioned bees and bees experiencing unpaired presentations of conditioned stimulus (CS) and unconditioned stimulus (US). We demonstrate that both behavioral protocols display differences in memory formation but do not alter the level of pAmCREB in bee brains directly after training. Nevertheless, we report that bees responding to the CS during unpaired stimulus presentations exhibit higher levels of pAmCREB than nonresponding bees. In addition, Trichostatin A, a histone deacetylase inhibitor that is thought to enhance histone acetylation by CREB-binding protein, increases the bees’ CS responsiveness. We conclude that pAmCREB is involved in gating a bee's behavioral response driven by an external stimulus. PMID:27084927

Leptin induces CREB-dependent aromatase activation through COX-2 expression in breast cancer cells.

PubMed

Kim, Hyung Gyun; Jin, Sun Woo; Kim, Yong An; Khanal, Tilak; Lee, Gi Ho; Kim, Se Jong; Rhee, Sang Dal; Chung, Young Chul; Hwang, Young Jung; Jeong, Tae Cheon; Jeong, Hye Gwang

2017-08-01

Leptin plays a key role in the control of adipocyte formation, as well as in the associated regulation of energy intake and expenditure. The goal of this study was to determine if leptin-induced aromatase enhances estrogen production and induces tumor cell growth stimulation. To this end, breast cancer cells were incubated with leptin in the absence or presence of inhibitor pretreatment, and changes in aromatase and cyclooxygenase-2 (COX-2) expression were evaluated at the mRNA and protein levels. Transient transfection assays were performed to examine the aromatase and COX-2 gene promoter activities and immunoblot analysis was used to examine protein expression. Leptin induced aromatase expression, estradiol production, and promoter activity in breast cancer cells. Protein levels of phospho-STAT3, PKA, Akt, ERK, and JNK were increased by leptin. Leptin also significantly increased cAMP levels, cAMP response element (CRE) activation, and CREB phosphorylation. In addition, leptin induced COX-2 expression, promoter activity, and increased the production of prostaglandin E 2 . Finally, a COX-2 inhibitor and aromatase inhibitor suppressed leptin-induced cell proliferation in MCF-7 breast cancer cells. Together, our data show that leptin increased aromatase expression in breast cancer cells, which was correlated with COX-2 upregulation, mediated through CRE activation and cooperation among multiple signaling pathways. Copyright © 2017 Elsevier Ltd. All rights reserved.

Epigenetic modification of miR-10a regulates renal damage by targeting CREB1 in type 2 diabetes mellitus.

PubMed

Shan, Qun; Zheng, Guihong; Zhu, Aihua; Cao, Li; Lu, Jun; Wu, Dongmei; Zhang, ZiFeng; Fan, Shaohua; Sun, Chunhui; Hu, Bin; Zheng, Yuanlin

2016-09-01

Emerging evidence has shown that microRNA-mediated gene expression modulation plays a crucial role in the pathogenesis of type 2 diabetes mellitus, but the novel miRNAs involved in type 2 diabetes and its functional regulatory mechanisms still need to be determined. In this study, we assessed the role of miR-10a in extracellular matrix accumulation in the kidney of diabetic mellitus induced by combining administration of chronic high fat diet (HFD) and low dosage of streptozotocin (STZ, 35mg/kg). Here, we found that HFD/STZ administration decreased the level of microRNA (miR-10a) expression in ICR strain mice. Overexpression of miR-10a alleviated the increased ratio of urine albumin-to-creatinine (ACR) ratio of HFD/STZ mice. In contrast, knockdown of miR-10a increased the ratio of kidney ACR in naïve mice. Furthermore, cAMP response element binding protein 1 (CREB1) was validated as a target of miR-10a in vitro and in vivo. CREB1 and its downstream fibronectin (FN, extracellular matrix) were increased in HFD/STZ-treated mice, which was reversed by kidney miR-10a overexpression. The content of CREB1 and FN was increased by miR-10a knockdown in kidney of naïve mice. Furthermore, histone deacetylase 3 (HDAC3) was revealed to be increased in kidney of HFD/STZ mice, accompanied with the augmentation of ACR ratio and FN level. Knockdown of HDAC3 with siRNA significantly caused the increase of miR-10a, resulting in the decrease in CREB1 and FN expression in kidney of HFD/STZ mice. Contrarily, HDAC3 overexpression mediated by lentivirus decreased miR-10a content, and enhanced ACR value, CREB1 and FN formation in naïve mice. Collectively, these results elucidate that HDAC3/miR-10a/CREB1 serves as a new mechanism underlying kidney injury, providing potential therapeutic targets in type 2 diabetes. Copyright © 2016. Published by Elsevier Inc.

Involvement of BDNF/TrkB and ERK/CREB axes in nitroglycerin-induced rat migraine and effects of estrogen on these signals in the migraine

PubMed Central

Guo, Jiu-Qing; Deng, Hui-Hui; Bo, Xiao

2017-01-01

ABSTRACT Migraine is a highly prevalent headache disorder, especially in women. Brain-derived neurotrophic factor (BDNF) and its receptor tropomyosin receptor kinases (TrkB), as well as extracellular signal-regulated kinase (ERK) and its downstream target c-AMP-responsive element binding protein (CREB) are strongly associated with the transmission of nociceptive information. However, the involvement of these substances in migraine has rarely been examined. In the present study, intraperitoneal injection of nitroglycerin (NTC) successfully induced rat migraine attack, as evidenced by behavioral testing. The location and abundance of these substances in the migraine model were determined by immunohistochemistry, real-time polymerase chain reaction (RT-PCR), western blot and enzyme-linked immunosorbant assays (ELISA). Results showed that BDNF, TrkB, phosphor(p)-ERK and p-CREB were up-regulated in the brain neurons of both male and female rats with NTG-induced migraine compared to non-migraine control, whereas their expression levels were decreased in headache-free intervals of the migraine compared to migraine attacks. Estrogen is an important contributor to migraine. Female ovariectomized rats showed significant reduction in the expression of BDNF, TrkB, p-CREB and p-ERK in both attacks and intervals of NTG-induced migraine, relative to rats that have their ovaries. But, intraperitoneal administration of exogenous estrogen recovered their expression in ovariectomized rats. Collectively, this study unveiled a positive correlation of BDNF/TrkB and ERK/CREB axes in NTG-induced migraine and promoting effects of estrogen on their signals in the migraine. These findings contribute to further understanding the pathogenesis of migraine in the molecular basis. PMID:27875242

Nutraceuticals to promote neuronal plasticity in response to corticosterone-induced stress in human neuroblastoma cells.

PubMed

Gite, Snehal; Ross, R Paul; Kirke, Dara; Guihéneuf, Freddy; Aussant, Justine; Stengel, Dagmar B; Dinan, Timothy G; Cryan, John F; Stanton, Catherine

2018-01-29

To search for novel compounds that will protect neuronal cells under stressed conditions that may help to restore neuronal plasticity. A model of corticosterone (CORT)-induced stress in human neuroblastoma cells (SH-SY5Y) was used to compare the efficacy of 6 crude extracts and 10 pure compounds (6 polyphenols, 2 carotenoids, 1 amino acid analogue, and 1 known antidepressant drug) to increase neuronal plasticity and to decrease cytotoxicity. Astaxanthin (among pure compounds) and phlorotannin extract of Fucus vesiculosus (among crude extracts) showed a maximum increase in cell viability in the presence of excess CORT. BDNF-VI mRNA expression in SH-SY5Y cells was significantly improved by pretreatment with quercetine, astaxanthin, curcumin, fisetin, and resveratrol. Among crude extracts, xanthohumol, phlorotannin extract (Ecklonia cava), petroleum ether extract (Nannochloropsis oculata), and phlorotannin extract (F. vesiculosus) showed a significant increase in BDNF-VI mRNA expression. CREB1 mRNA expression was significantly improved by astaxanthin, β-carotene, curcumin, and fluoxetine whereas none of the crude extracts caused significant improvement. As an adjunct of fluoxetine, phlorotannin extract (F. vesiculosus), β-carotene, and xanthohumol have resulted in significant improvement in BDNF-VI mRNA expression and CREB1 mRNA expression was significantly improved by phlorotannin extract (F. vesiculosus). Significant improvement in mature BDNF protein expression by phlorotannin extract (F. vesiculosus) and β-carotene as an adjunct of fluoxetine confirm their potential to promote neuronal plasticity against CORT-induced stress. The carotenoids, flavonoids, namely quercetine, curcumin, and low molecular weight phlorotannin-enriched extract of F. vesiculosus may serve as potential neuroprotective agents promoting neuronal plasticity in vitro. Graphical abstract: Cascade of events associated with disturbed homeostatic balance of glucocorticoids and impact of phlorotannin extract (F. vesiculosus) and β-carotene in restoring neuronal plasticity. Abbreviation: TrKB, tropomyosin receptor kinase B; P-ERK, phosphorylated extracellular signal-related kinase; PI3K, phosphatidylinositol 3-kinase; Akt, protein kinase B; Ca++/CaMK, calcium/calmodulin-dependent protein kinase; pCREB, phosphorylated cAMP response element-binding protein; CRE, cAMP response elements, CORT, corticosterone; and BDNF; brain-derived neurotrophic factor.

Involvement of Phosphorylated "Apis Mellifera" CREB in Gating a Honeybee's Behavioral Response to an External Stimulus

ERIC Educational Resources Information Center

Gehring, Katrin B.; Heufelder, Karin; Feige, Janina; Bauer, Paul; Dyck, Yan; Ehrhardt, Lea; Kühnemund, Johannes; Bergmann, Anja; Göbel, Josefine; Isecke, Marlene; Eisenhardt, Dorothea

2016-01-01

The transcription factor cAMP-response element-binding protein (CREB) is involved in neuronal plasticity. Phosphorylation activates CREB and an increased level of phosphorylated CREB is regarded as an indicator of CREB-dependent transcriptional activation. In honeybees ("Apis mellifera") we recently demonstrated a particular high…

Atorvastatin inhibits insulin synthesis by inhibiting the Ras/Raf/ERK/CREB pathway in INS-1 cells

PubMed Central

Sun, Hongxi; Li, Yu; Sun, Bei; Hou, Ningning; Yang, Juhong; Zheng, Miaoyan; Xu, Jie; Wang, Jingyu; Zhang, Yi; Zeng, Xianwei; Shan, Chunyan; Chang, Bai; Chen, Liming; Chang, Baocheng

2016-01-01

Abstract Backround: Type 2 diabetes has become a global epidemic disease. Atorvastatin has become a cornerstone in the prevention and treatment of atherosclerosis. However, increasing evidence showed that statins can dose-dependently increase the risk of diabetes mellitus. The mechanism is not clear. Objective: The Ras complex pathway (Ras/Raf/extracellular signal-regulated kinase [ERK]/cAMP response element-binding protein [CREB]) is the major pathway that regulates the gene transcription. Except for the inhibition of cholesterol synthesis by inhibiting the 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-COA) reductase, statins can also downregulate the phosphorylation of a series of downstream substrates including the key proteins of the Ras complex pathway, therefore may inhibit the insulin syntheses in pancreatic beta cells. In our study, we investigated the inhibitory effect and the underlying mechanism of atorvastatin on insulin synthesis in rat islets. Methods: Islets were isolated from Wistar rats and cultured in Roswell Park Memorial Institute (RPMI)-1640 medium. The insulin content in the medium was measured by radioimmunoassay before and after the treatment of 50 μM atorvastatin. Effect of atorvastatin on the expression of insulin message Ribonucleic acid (mRNA) in pancreatic islet beta cells was also detected using quantitative real-time polymerase chain reaction. Western blotting was used to explore the possible role of the Ras complex pathway (Ras/Raf/ERK/CREB) in atorvastatin-inhibited insulin synthesis. The effects of atorvastatin on the binding of nuclear transcription factor p-CREB with CRE in INS-1 cells were examined via chromatin immunoprecipitation assay. Results: Compared with the control group, the insulin level decreased by 27.1% at 24 hours after atorvastatin treatment. Atorvastatin inhibited insulin synthesis by decreasing insulin mRNA expression of pancreatic islet beta cells. The activities of Ras, Raf-1, and p-CREB in the Ras complex pathway were inhibited by 50 μM atorvastatin in INS-1 cells in vitro. Moreover, 50 μM atorvastatin reduced the binding of p-CREB with deoxyribonucleic acid (DNA) in INS-1 cells in vitro. Conclusion: Atorvastatin inhibits insulin synthesis in beta cells by inhibiting the activation of the Ras complex pathway. PMID:27684825

Long-term neurocognitive dysfunction in offspring via NGF/ ERK/CREB signaling pathway caused by ketamine exposure during the second trimester of pregnancy in rats.

PubMed

Li, Yanan; Li, Xinran; Guo, Cen; Li, Lina; Wang, Yuxin; Zhang, Yiming; Chen, Yu; Liu, Wenhan; Gao, Li

2017-05-09

Early life exposure to ketamine caused neurohistopathologic changes and persistent cognitive dysfunction. For this study, a pregnant rat model was developed to investigate neurocognitive effects in the offspring, following ketamine exposure during the second trimester. Pregnant rats on gestational day 14 (equal to midtrimester pregnancy in humans), intravenously received 200 mg/kg ketamine for 3 h. Their behavior was tested (Morris water maze, odor recognition test, and fear conditioning) at postnatal days (P25-30). Furthermore, hippocampal morphology of the offspring (P30) was examined via Nissl staining and hippocampal dendritic spine density was determined via Golgi staining. The hippocampal protein levels of nerve growth factor (NGF), extracellular signal-regulated kinase (ERK), phosphorylated-ERK (p-ERK), cyclic adenosine monophosphate response element-binding (CREB), p-CREB, synaptophysin (SYP), synapsin (SYN), and postsynaptic density-95 (PSD95) were measured via western blot. Additionally, SCH772984 (an ERK inhibitor) was used to evaluate both role and underlying mechanism of the ERK pathway in PC12 cells. We found that ketamine caused long-term neurocognitive dysfunction, reduced the density of the dendritic spin, caused neuronal loss, and down-regulated the expression of NGF, ERK, p-ERK, mitogen, and stress-activated protein kinase (MSK), CREB, p-CREB, SYP, SYN, and PSD95 in the hippocampus. These results suggest that ketamine induced maternal anesthesia during period of the fetal brain development can cause long-term neurocognitive dysfunction in the offspring, which likely happens via inhibition of the NGF-ERK-CREB pathway in the hippocampus. Our results highlight the central role of ERK in neurocognition.

Cyclic AMP Response Element Binding Protein Mediates Pathological Retinal Neovascularization via Modulating DLL4-NOTCH1 Signaling

PubMed Central

Singh, Nikhlesh K.; Kotla, Sivareddy; Kumar, Raj; Rao, Gadiparthi N.

2015-01-01

Retinal neovascularization is the most common cause of moderate to severe vision loss in all age groups. Despite the use of anti-VEGFA therapies, this complication continues to cause blindness, suggesting a role for additional molecules in retinal neovascularization. Besides VEGFA and VEGFB, hypoxia induced VEGFC expression robustly. Based on this finding, we tested the role of VEGFC in pathological retinal angiogenesis. VEGFC induced proliferation, migration, sprouting and tube formation of human retinal microvascular endothelial cells (HRMVECs) and these responses require CREB-mediated DLL4 expression and NOTCH1 activation. Furthermore, down regulation of VEGFC levels substantially reduced tip cell formation and retinal neovascularization in vivo. In addition, we observed that CREB via modulating the DLL4-NOTCH1 signaling mediates VEGFC-induced tip cell formation and retinal neovascularization. In regard to upstream mechanism, we found that down regulation of p38β levels inhibited hypoxia-induced CREB-DLL4-NOTCH1 activation, tip cell formation, sprouting and retinal neovascularization. Based on these findings, it may be suggested that VEGFC besides its role in the regulation of lymphangiogenesis also plays a role in pathological retinal angiogenesis and this effect depends on p38β and CREB-mediated activation of DLL4-NOTCH1 signaling. PMID:26870802

Connexin31.1 deficiency in the mouse impairs object memory and modulates open-field exploration, acetylcholine esterase levels in the striatum, and cAMP response element-binding protein levels in the striatum and piriform cortex.

PubMed

Dere, E; Zheng-Fischhöfer, Q; Viggiano, D; Gironi Carnevale, U A; Ruocco, L A; Zlomuzica, A; Schnichels, M; Willecke, K; Huston, J P; Sadile, A G

2008-05-02

Neuronal gap junctions in the brain, providing intercellular electrotonic signal transfer, have been implicated in physiological and behavioral correlates of learning and memory. In connexin31.1 (Cx31.1) knockout (KO) mice the coding region of the Cx31.1 gene was replaced by a LacZ reporter gene. We investigated the impact of Cx31.1 deficiency on open-field exploration, the behavioral response to an odor, non-selective attention, learning and memory performance, and the levels of memory-related proteins in the hippocampus, striatum and the piriform cortex. In terms of behavior, the deletion of the Cx31.1 coding DNA in the mouse led to increased exploratory behaviors in a novel environment, and impaired one-trial object recognition at all delays tested. Despite strong Cx31.1 expression in the peripheral and central olfactory system, Cx31.1 KO mice exhibited normal behavioral responses to an odor. We found increased levels of acetylcholine esterase (AChE) and cAMP response element-binding protein (CREB) in the striatum of Cx31.1 KO mice. In the piriform cortex the Cx31.1 KO mice had an increased heterogeneity of CREB expression among neurons. In conclusion, gap-junctions featuring the Cx31.1 protein might be involved in open-field exploration as well as object memory and modulate levels of AChE and CREB in the striatum and piriform cortex.

The Role of CREB, SRF, and MEF2 in Activity-Dependent Neuronal Plasticity in the Visual Cortex.

PubMed

Pulimood, Nisha S; Rodrigues, Wandilson Dos Santos; Atkinson, Devon A; Mooney, Sandra M; Medina, Alexandre E

2017-07-12

The transcription factors CREB (cAMP response element binding factor), SRF (serum response factor), and MEF2 (myocyte enhancer factor 2) play critical roles in the mechanisms underlying neuronal plasticity. However, the role of the activation of these transcription factors in the different components of plasticity in vivo is not well known. In this study, we tested the role of CREB, SRF, and MEF2 in ocular dominance plasticity (ODP), a paradigm of activity-dependent neuronal plasticity in the visual cortex. These three proteins bind to the synaptic activity response element (SARE), an enhancer sequence found upstream of many plasticity-related genes (Kawashima et al., 2009; Rodríguez-Tornos et al., 2013), and can act cooperatively to express Arc , a gene required for ODP (McCurry et al., 2010). We used viral-mediated gene transfer to block the transcription function of CREB, SRF, and MEF2 in the visual cortex, and measured visually evoked potentials in awake male and female mice before and after a 7 d monocular deprivation, which allowed us to examine both the depression component (Dc-ODP) and potentiation component (Pc-ODP) of plasticity independently. We found that CREB, SRF, and MEF2 are all required for ODP, but have differential effects on Dc-ODP and Pc-ODP. CREB is necessary for both Dc-ODP and Pc-ODP, whereas SRF and MEF2 are only needed for Dc-ODP. This finding supports previous reports implicating SRF and MEF2 in long-term depression (required for Dc-ODP), and CREB in long-term potentiation (required for Pc-ODP). SIGNIFICANCE STATEMENT Activity-dependent neuronal plasticity is the cellular basis for learning and memory, and it is crucial for the refinement of neuronal circuits during development. Identifying the mechanisms of activity-dependent neuronal plasticity is crucial to finding therapeutic interventions in the myriad of disorders where it is disrupted, such as Fragile X syndrome, Rett syndrome, epilepsy, major depressive disorder, and autism spectrum disorder. Transcription factors are essential nuclear proteins that trigger the expression of gene programs required for long-term functional and structural plasticity changes. Our results elucidate the specific role of the transcription factors CREB, SRF, and MEF2 in the depression and potentiation components of ODP in vivo , therefore better informing future attempts to find therapeutic targets for diseases where activity-dependent plasticity is disrupted. Copyright © 2017 the authors 0270-6474/17/376628-10$15.00/0.

The Role of CREB, SRF, and MEF2 in Activity-Dependent Neuronal Plasticity in the Visual Cortex

PubMed Central

Rodrigues, Wandilson dos Santos; Mooney, Sandra M.

2017-01-01

The transcription factors CREB (cAMP response element binding factor), SRF (serum response factor), and MEF2 (myocyte enhancer factor 2) play critical roles in the mechanisms underlying neuronal plasticity. However, the role of the activation of these transcription factors in the different components of plasticity in vivo is not well known. In this study, we tested the role of CREB, SRF, and MEF2 in ocular dominance plasticity (ODP), a paradigm of activity-dependent neuronal plasticity in the visual cortex. These three proteins bind to the synaptic activity response element (SARE), an enhancer sequence found upstream of many plasticity-related genes (Kawashima et al., 2009; Rodríguez-Tornos et al., 2013), and can act cooperatively to express Arc, a gene required for ODP (McCurry et al., 2010). We used viral-mediated gene transfer to block the transcription function of CREB, SRF, and MEF2 in the visual cortex, and measured visually evoked potentials in awake male and female mice before and after a 7 d monocular deprivation, which allowed us to examine both the depression component (Dc-ODP) and potentiation component (Pc-ODP) of plasticity independently. We found that CREB, SRF, and MEF2 are all required for ODP, but have differential effects on Dc-ODP and Pc-ODP. CREB is necessary for both Dc-ODP and Pc-ODP, whereas SRF and MEF2 are only needed for Dc-ODP. This finding supports previous reports implicating SRF and MEF2 in long-term depression (required for Dc-ODP), and CREB in long-term potentiation (required for Pc-ODP). SIGNIFICANCE STATEMENT Activity-dependent neuronal plasticity is the cellular basis for learning and memory, and it is crucial for the refinement of neuronal circuits during development. Identifying the mechanisms of activity-dependent neuronal plasticity is crucial to finding therapeutic interventions in the myriad of disorders where it is disrupted, such as Fragile X syndrome, Rett syndrome, epilepsy, major depressive disorder, and autism spectrum disorder. Transcription factors are essential nuclear proteins that trigger the expression of gene programs required for long-term functional and structural plasticity changes. Our results elucidate the specific role of the transcription factors CREB, SRF, and MEF2 in the depression and potentiation components of ODP in vivo, therefore better informing future attempts to find therapeutic targets for diseases where activity-dependent plasticity is disrupted. PMID:28607167

Effects of intravenous anesthetics on the phosphorylation of cAMP response element‑binding protein in hippocampal slices of adult mice.

PubMed

Gao, Haiying; Zhang, Lingyu; Chen, Zhenyi; Liu, Shuncui; Zhang, Qinghong; Zhang, Bingxi

2018-04-27

cAMP response‑element binding protein (CREB) functions in hippocampal synaptic plasticity and memory formation. However, it remains unknown whether intravenous anesthetics modulate CREB. The present study aimed to examine the effects of intravenous anesthetics on CREB phosphorylation in the mouse hippocampus. CREB phosphorylation was examined in hippocampal slices with and without pharmacological or intravenous anesthetics via immunoblotting. In a dose‑response experiment, the concentrations of intravenous anesthetics ranged from 10‑9 to 10‑4 mol/l for 1 h. For the time‑response experiment, these slices were incubated with 5x10‑6 mol/l of propofol for 0, 1, 2, 5, 7, 9, 12, 15, 30 and 60 min. In order to examine whether CREB phosphorylation could be recovered following washing out the propofol, the slices were incubated in plain artificial cerebrospinal fluid at different time durations following 5 min incubation with propofol. Propofol, etomidate, ketamine and midazolam inhibited CREB phosphorylation (P<0.05) in a time‑ and dose‑dependent manner. This inhibition was reversible following the removal of propofol, and was rescued by CREB phosphorylation (P<0.05). The decrease in CREB phosphorylation revealed additive effects with 100 µM of chelerythrine and 20 µM of PD‑98059, and the etomidate‑induced decrease in CREB phosphorylation was blocked by 1 mM of NMDA. However, 0.1 µM of phorbol 12‑myristate 13‑acetate, 50 µM of U 73122, 100 µM of carbachol and 10 µM of MK801 were ineffective in the anesthetic‑induced decrease in CREB phosphorylation. Intravenous anesthetics markedly decreased CREB phosphorylation in the mouse hippocampus, which was most likely via the protein kinase C and mitogen activated protein kinase pathways. This suggests that CREB represents a target for anesthetic action in the brain.

Astaxanthin Inhibits Acetaldehyde-Induced Cytotoxicity in SH-SY5Y Cells by Modulating Akt/CREB and p38MAPK/ERK Signaling Pathways.

PubMed

Yan, Tingting; Zhao, Yan; Zhang, Xia; Lin, Xiaotong

2016-03-10

Excessive alcohol consumption can lead to brain tissue damage and cognitive dysfunction. Acetaldehyde, the most toxic metabolite of ethanol, mediates the brain tissue damage and cognitive dysfunction induced by chronic excessive alcohol consumption. In this study, the effect of astaxanthin, a marine bioactive compound, on acetaldehyde-induced cytotoxicity was investigated in SH-SY5Y cells. It was found that astaxanthin protected cells from apoptosis by ameliorating the effect of acetaldehyde on the expression of Bcl-2 family proteins, preventing the reduction of anti-apoptotic protein Bcl-2 and the increase of pro-apoptotic protein Bak induced by acetaldehyde. Further analyses showed that astaxanthin treatment inhibited acetaldehyde-induced reduction of the levels of activated Akt and cyclic AMP-responsive element binding protein (CREB). Astaxanthin treatment also prevented acetaldehyde-induced increase of the level of activated p38 mitogen-activated protein kinase (MAPK) and decrease of the level of activated extracellular signal-regulated kinases (ERKs). Activation of Akt/CREB pathway promotes cell survival and is involved in the upregulation of Bcl-2 gene. P38MAPK plays a critical role in apoptotic events while ERKs mediates the inhibition of apoptosis. Thus, astaxanthin may inhibit acetaldehyde-induced apoptosis through promoting the activation of Akt/CREB and ERKs and blocking the activation of p38MAPK. In addition, astaxanthin treatment suppressed the oxidative stress induced by acetaldehyde and restored the antioxidative capacity of SH-SY5Y cells. Therefore, astaxanthin may protect cells against acetaldehyde-induced cytotoxicity through maintaining redox balance and modulating apoptotic and survival signals. The results suggest that astaxanthin treatment may be beneficial for preventing neurotoxicity associated with acetaldehyde and excessive alcohol consumption.

Cardiac glycoside ouabain induces activation of ATF-1 and StAR expression by interacting with the α4 isoform of the sodium pump in Sertoli cells.

PubMed

Dietze, Raimund; Konrad, Lutz; Shihan, Mazen; Kirch, Ulrike; Scheiner-Bobis, Georgios

2013-03-01

Sertoli cells express α1 and α4 isoforms of the catalytic subunit of Na(+),K(+)-ATPase (sodium pump). Our recent findings demonstrated that interactions of the α4 isoform with cardiotonic steroids (CTS) like ouabain induce signaling cascades that resemble the so-called non-classical testosterone pathway characterized by activation of the c-Src/c-Raf/Erk1/2/CREB signaling cascade. Here we investigate a possible physiological significance of the activated cascade. The results obtained in the current investigation show that the ouabain-induced signaling cascade also leads to the activation of the CREB-related activating transcription factor 1 (ATF-1) in the Sertoli cell line 93RS2 in a concentration- and time-dependent manner, as demonstrated by detection of ATF-1 phosphorylated on Ser63 in western blots. The ouabain-activated ATF-1 protein was found to localize to the cell nuclei. The sodium pump α4 isoform mediates this activation, as it is ablated when cells are incubated with siRNA to the α4 isoform. Ouabain also leads to increased expression of steroidogenic acute regulator (StAR) protein, which has been shown to be a downstream consequence of CREB/ATF-1 activation. Taking into consideration that CTS are most likely produced endogenously, the demonstrated induction of StAR expression by ouabain establishes a link between CTS, the α4 isoform of the sodium pump, and steroidogenesis crucial for male fertility and reproduction. Copyright © 2012 Elsevier B.V. All rights reserved.

Attenuation of pCREB and Egr1 expression in the insular and anterior cingulate cortices associated with enhancement of CFA-evoked mechanical hypersensitivity after repeated forced swim stress.

PubMed

Imbe, Hiroki; Kimura, Akihisa

2017-09-01

The perception and response to pain are severely impacted by exposure to stressors. In some animal models, stress increases pain sensitivity, which is termed stress-induced hyperalgesia (SIH). The insular cortex (IC) and anterior cingulate cortex (ACC), which are typically activated by noxious stimuli, affect pain perception through the descending pain modulatory system. In the present study, we examined the expression of phospho-cAMP response element-binding protein (pCREB) and early growth response 1 (Egr1) in the IC and ACC at 3h (the acute phase of peripheral tissue inflammation) after complete Freund's adjuvant (CFA) injection in naïve rats and rats preconditioned with forced swim stress (FS) to clarify the effect of FS, a stressor, on cortical cell activities in the rats showing SIH induced by FS. The CFA injection into the hindpaw induced mechanical hypersensitivity and increased the expression of the pCREB and Egr1 in the IC and ACC at 3h after the injection. FS (day 1, 10min; days 2-3, 20min) prior to the CFA injection enhanced the CFA-induced mechanical hypersensitivity and attenuated the increase in the expression of pCREB and Egr1 in the IC and ACC. These findings suggested that FS modulates the CFA injection-induced neuroplasticity in the IC and ACC to enhance the mechanical hypersensitivity. These findings are thought to signify stressor-induced dysfunction of the descending pain modulatory system. Copyright © 2017 Elsevier Inc. All rights reserved.

Syk Inhibits the Activity of Protein Kinase A by Phosphorylating Tyrosine 330 of the Catalytic Subunit*

PubMed Central

Yu, Shuai; Huang, He; Iliuk, Anton; Wang, Wen-Horng; Jayasundera, Keerthi B.; Tao, W. Andy; Post, Carol B.; Geahlen, Robert L.

2013-01-01

The Syk protein-tyrosine kinase can have multiple effects on cancer cells, acting in some as a tumor suppressor by inhibiting motility and in others as a tumor promoter by enhancing survival. Phosphoproteomic analyses identified PKA as a Syk-specific substrate. Syk catalyzes the phosphorylation of the catalytic subunit of PKA (PKAc) both in vitro and in cells on Tyr-330. Tyr-330 lies within the adenosine-binding motif in the C-terminal tail of PKAc within a cluster of acidic amino acids (DDYEEEE), which is a characteristic of Syk substrates. The phosphorylation of PKAc on Tyr-330 by Syk strongly inhibits its catalytic activity. Molecular dynamics simulations suggest that this additional negative charge prevents the C-terminal tail from interacting with the substrate and the nucleotide-binding site to stabilize the closed conformation of PKAc, thus preventing catalysis from occurring. Phosphoproteomic analyses and Western blotting studies indicate that Tyr-330 can be phosphorylated in a Syk-dependent manner in MCF7 breast cancer cells and DT40 B cells. The phosphorylation of a downstream substrate of PKAc, cAMP-responsive element-binding protein (CREB), is inhibited in cells expressing Syk but can be rescued by a selective inhibitor of Syk. Modulation of CREB activity alters the expression of the CREB-regulated gene BCL2 and modulates cellular responses to genotoxic agents. Thus, PKA is a novel substrate of Syk, and its phosphorylation on Tyr-330 inhibits its participation in downstream signaling pathways. PMID:23447535

Regulation of human bone sialoprotein gene transcription by platelet-derived growth factor-BB.

PubMed

Mezawa, Masaru; Araki, Shouta; Takai, Hideki; Sasaki, Yoko; Wang, Shuang; Li, Xinyue; Kim, Dong-Soon; Nakayama, Youhei; Ogata, Yorimasa

2009-04-15

Platelet-derived growth factor (PDGF) is produced by mesenchymal cells and released by platelets following aggregation and is synthesized by osteoblasts. In bone, PDGF stimulates proliferation and differentiation of osteoblasts. PDGF also increases bone resorption, most likely by increasing the number of osteoclasts. Bone sialoprotein (BSP) is thought to function in the initial mineralization of bone, selectively expressed by differentiated osteoblast. To determine the molecular mechanisms PDGF regulation of human BSP gene transcription, we have analyzed the effects of PDGF-BB on osteoblast-like Saos2 and ROS17/2.8 cells. PDGF-BB (5 ng/ml) increased BSP mRNA and protein levels at 12 h in Saos2 cells, and induced BSP mRNA expression at 3 h, reached maximal at 12 h in ROS17/2.8 cells. Transient transfection analyses were performed using chimeric constructs of the human BSP gene promoter linked to a luciferase reporter gene. Treatment of Saos2 cells with PDGF-BB (5 ng/ml, 12 h) increased luciferase activities of all constructs between -184LUC to -2672LUC including the human BSP gene promoter. Effects of PDGF-BB abrogated in constructs included 2 bp mutations in the two cAMP response elements (CRE1 and CRE2), activator protein 1(3) (AP1(3)) and shear stress response element 1 (SSRE1). Luciferase activities induced by PDGF-BB were blocked by protein kinase A inhibitor H89 and tyrosine kinase inhibitor herbimycin A. Gel mobility shift analyses showed that PDGF-BB increased binding of CRE1, CRE2, AP1(3) and SSRE1 elements. CRE1- and CRE2-protein complexes were supershifted by CREB1 and phospho-CREB1 antibodies. Notably, AP1(3)-protein complexes were supershifted by c-Fos and JunD, and disrupted by CREB1, phospho-CREB1, c-Jun and Fra2 antibodies. These studies, therefore, demonstrate that PDGF-BB stimulates human BSP transcription by targeting the CRE1, CRE2, AP1(3) and SSRE1 elements in the human BSP gene promoter.

Curcumin confers neuroprotection against alcohol-induced hippocampal neurodegeneration via CREB-BDNF pathway in rats.

PubMed

Motaghinejad, Majid; Motevalian, Manijeh; Fatima, Sulail; Hashemi, Hajar; Gholami, Mina

2017-03-01

Alcohol abuse causes severe damage to the brain neurons. Studies have reported the neuroprotective effects of curcumin against alcohol-induced neurodegeneration. However, the precise mechanism of action remains unclear. Seventy rats were equally divided into 7 groups (10 rats per group). Group 1 received normal saline (0.7ml/rat) and group 2 received alcohol (2g/kg/day) for 21days. Groups 3, 4, 5 and 6 concurrently received alcohol (2g/kg/day) and curcumin (10, 20, 40 and 60mg/kg, respectively) for 21days. Animals in group 7 self- administered alcohol for 21days. Group 8 treated with curcumin (60mg/kg, i.p.) alone for 21days. Open Field Test (OFT) was used to investigate motor activity in rats. Hippocampal oxidative, antioxidative and inflammatory factors were evaluated. Furthermore, brain cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) and brain derived neurotrophic factor (BDNF) levels were studied at gene level by reverse transcriptase polymerase chain reaction (RT-PCR). In addition, protein expression for BDNF, CREB, phosphorylated CREB (CREB-P), Bax and Bcl-2 was determined by western blotting. Voluntary and involuntary administration of alcohol altered motor activity in OFT, and curcumin treatment inhibited this alcohol-induced motor disturbance. Also, alcohol administration augmented lipid peroxidation, mitochondrial oxidized glutathione (GSSG), interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α) and Bax levels in isolated hippocampal tissues. Furthermore, alcohol-induced significant reduction were observed in reduced form of glutathione (GSH), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GR) activities and CREB, BDNF and Bcl-2 levels. Also curcumin alone did not change the behavior and biochemical and molecular parameters. Curcumin can act as a neuroprotective agent against neurodegenerative effects of alcohol abuse, probably via activation of CREB-BDNF signaling pathway. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Effects of oral eicosapentaenoic acid versus docosahexaenoic acid on human peripheral blood mononuclear cell gene expression.

PubMed

Tsunoda, Fumiyoshi; Lamon-Fava, Stefania; Asztalos, Bela F; Iyer, Lakshmanan K; Richardson, Kris; Schaefer, Ernst J

2015-08-01

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have beneficial effects on inflammation and cardiovascular disease (CVD). Our aim was to assess the effect of a six-week supplementation with either olive oil, EPA, or DHA on gene expression in peripheral blood mononuclear cells (PBMC). Subjects were sampled at baseline and six weeks after receiving either: olive oil 6.0 g/day (n = 16), EPA 1.8 g/day (n = 16), or DHA 1.8 g/day (n = 18). PBMC were subjected to gene expression analysis by microarray with key findings confirmed by quantitative real-time polymerase chain reaction (Q-PCR). Plasma phospholipid EPA increased 3 fold in the EPA group, and DHA increased 63% in the DHA group (both p < 0.01), while no effects were observed in the olive oil group. Microarray analysis indicated that EPA but not DHA or olive oil significantly affected the gene expression in the following pathways: 1) interferon signaling, 2) receptor recognition of bacteria and viruses, 3) G protein signaling, glycolysis and glycolytic shunting, 4) S-adenosyl-l-methionine biosynthesis, and 5) cAMP-mediated signaling including cAMP responsive element protein 1 (CREB1), as well as many other individual genes including hypoxia inducible factor 1, α subunit (HIF1A). The findings for CREB1 and HIF1A were confirmed by Q-PCR analysis. Our data indicate that EPA supplementation was associated with significant effects on gene expression involving the interferon pathway as well as down-regulation of CREB1 and HIF1A, which may relate to its beneficial effect on CVD risk reduction. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

A role for neuronal cAMP responsive-element binding (CREB)-1 in brain responses to calorie restriction

PubMed Central

Fusco, Salvatore; Ripoli, Cristian; Podda, Maria Vittoria; Ranieri, Sofia Chiatamone; Leone, Lucia; Toietta, Gabriele; McBurney, Michael W.; Schütz, Günther; Riccio, Antonella; Grassi, Claudio; Galeotti, Tommaso; Pani, Giovambattista

2012-01-01

Calorie restriction delays brain senescence and prevents neurodegeneration, but critical regulators of these beneficial responses other than the NAD+-dependent histone deacetylase Sirtuin-1 (Sirt-1) are unknown. We report that effects of calorie restriction on neuronal plasticity, memory and social behavior are abolished in mice lacking cAMP responsive-element binding (CREB)-1 in the forebrain. Moreover, CREB deficiency drastically reduces the expression of Sirt-1 and the induction of genes relevant to neuronal metabolism and survival in the cortex and hippocampus of dietary-restricted animals. Biochemical studies reveal a complex interplay between CREB and Sirt-1: CREB directly regulates the transcription of the sirtuin in neuronal cells by binding to Sirt-1 chromatin; Sirt-1, in turn, is recruited by CREB to DNA and promotes CREB-dependent expression of target gene peroxisome proliferator-activated receptor-γ coactivator-1α and neuronal NO Synthase. Accordingly, expression of these CREB targets is markedly reduced in the brain of Sirt KO mice that are, like CREB-deficient mice, poorly responsive to calorie restriction. Thus, the above circuitry, modulated by nutrient availability, links energy metabolism with neurotrophin signaling, participates in brain adaptation to nutrient restriction, and is potentially relevant to accelerated brain aging by overnutrition and diabetes. PMID:22190495

Nobiletin enhances differentiation and lipolysis of 3T3-L1 adipocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Saito, Takeshi; Abe, Daigo; Sekiya, Keizo

2007-06-01

Nobiletin is a polymethoxylated flavone found in certain citrus fruits. Here we demonstrate that nobiletin enhance differentiation of 3T3-L1 preadipocytes. Nobiletin dose-dependently increased accumulation of lipid droplets in adipocytes. Quantitative RT-PCR analyses indicated that nobiletin increased the expression of genes critical for acquisition of the adipocyte phenotype. Some of them were known peroxisome proliferator activated receptor {gamma} (PPAR{gamma}) targets and PPAR{gamma} itself, however, nobiletin did not exhibit PPAR{gamma} ligand activity. We observed the expression of CCAAT/enhancer binding protein {beta} (C/EBP{beta}), a transcription factor for PPAR{gamma}, was increased by nobiletin. The activation of cAMP-responsive element binding protein (CREB) and extracellular signal-regulatedmore » kinase (ERK), which play important roles in C/EBP{beta} expression were also potentiated by nobiletin. Furthermore, nobiletin stimulated lipolysis in differentiated adipocytes, which is known to be stimulated by cAMP pathway. These results suggested that nobiletin enhanced both differentiation and lipolysis of adipocyte through activation of signaling cascades mediated by cAMP/CREB.« less

The effect of soybean isoflavone on the dysregulation of NMDA receptor signaling pathway induced by β-amyloid peptides 1-42 in rats.

PubMed

Xi, Yuan-Di; Ding, Juan; Han, Jing; Zhang, Dan-Di; Liu, Jin-Meng; Feng, Ling-Li; Xiao, Rong

2015-05-01

Synaptic damage is the key factor of cognitive impairment. The purpose of this study was to understand the effect of soybean isoflavone (SIF) on synaptic damage induced by β-amyloid peptide 1-42 (Aβ1-42) in rats. Adult male Wistar rats were randomly divided into control, Aβ1-42, SIF, and SIF + Aβ1-42 (SIF pretreatment) groups according to body weight. SIF was treated orally by gavage in SIF and SIF + Aβ1-42 groups. After 14 days pretreatment with SIF or vehicle, Aβ1-42 was injected into the lateral cerebral ventricle of rats in Aβ1-42 and SIF + Aβ1-42 groups using miniosmotic pump. The level of Aβ1-42 and the expression of N-methyl-D-aspartic-acid receptor (NMDAR) were observed by immunohistochemistry. Reverse transcriptase polymerase chain reaction was used to detect the mRNA levels of NMDAR, calmodulin (CaM), calcium/CaM-dependent protein kinase II (CaMKII), cAMP-response element binding protein (CREB), and brain-derived neurotrophic factor (BDNF). The results showed that Aβ1-42 down-regulated mRNA and protein expression of the NR1 and NR2B subunits of NMDAR, SIF pretreatment could reverse these changes. The mRNA expression of CaM, CaMKII, CREB, and BDNF were down-regulated by Aβ1-42, but they were all regulated by SIF pretreatment. These results suggest that SIF pretreatment could antagonize the neuron damage in rats induced by Aβ1-42, and its mechanism might be associated with the NMDA receptor and CaM/CaMKII/CREB/BDNF signaling pathway, which are the synaptic plasticity-related molecules.

Molecular interactions involved in the transactivation of the human T-cell leukemia virus type 1 promoter mediated by Tax and CREB-2 (ATF-4).

PubMed

Gachon, F; Thebault, S; Peleraux, A; Devaux, C; Mesnard, J M

2000-05-01

The human T-cell leukemia virus type 1 (HTLV-1) Tax protein activates viral transcription through three 21-bp repeats located in the U3 region of the HTLV-1 long terminal repeat and called Tax-responsive elements (TxREs). Each TxRE contains nucleotide sequences corresponding to imperfect cyclic AMP response elements (CRE). In this study, we demonstrate that the bZIP transcriptional factor CREB-2 is able to bind in vitro to the TxREs and that CREB-2 binding to each of the 21-bp motifs is enhanced by Tax. We also demonstrate that Tax can weakly interact with CREB-2 bound to a cellular palindromic CRE motif such as that found in the somatostatin promoter. Mutagenesis of Tax and CREB-2 demonstrates that both N- and C-terminal domains of Tax and the C-terminal region of CREB-2 are required for direct interaction between the two proteins. In addition, the Tax mutant M47, defective for HTLV-1 activation, is unable to form in vitro a ternary complex with CREB-2 and TxRE. In agreement with recent results suggesting that Tax can recruit the coactivator CREB-binding protein (CBP) on the HTLV-1 promoter, we provide evidence that Tax, CREB-2, and CBP are capable of cooperating to stimulate viral transcription. Taken together, our data highlight the major role played by CREB-2 in Tax-mediated transactivation.

Molecular Interactions Involved in the Transactivation of the Human T-Cell Leukemia Virus Type 1 Promoter Mediated by Tax and CREB-2 (ATF-4)

PubMed Central

Gachon, Frederic; Thebault, Sabine; Peleraux, Annick; Devaux, Christian; Mesnard, Jean-Michel

2000-01-01

The human T-cell leukemia virus type 1 (HTLV-1) Tax protein activates viral transcription through three 21-bp repeats located in the U3 region of the HTLV-1 long terminal repeat and called Tax-responsive elements (TxREs). Each TxRE contains nucleotide sequences corresponding to imperfect cyclic AMP response elements (CRE). In this study, we demonstrate that the bZIP transcriptional factor CREB-2 is able to bind in vitro to the TxREs and that CREB-2 binding to each of the 21-bp motifs is enhanced by Tax. We also demonstrate that Tax can weakly interact with CREB-2 bound to a cellular palindromic CRE motif such as that found in the somatostatin promoter. Mutagenesis of Tax and CREB-2 demonstrates that both N- and C-terminal domains of Tax and the C-terminal region of CREB-2 are required for direct interaction between the two proteins. In addition, the Tax mutant M47, defective for HTLV-1 activation, is unable to form in vitro a ternary complex with CREB-2 and TxRE. In agreement with recent results suggesting that Tax can recruit the coactivator CREB-binding protein (CBP) on the HTLV-1 promoter, we provide evidence that Tax, CREB-2, and CBP are capable of cooperating to stimulate viral transcription. Taken together, our data highlight the major role played by CREB-2 in Tax-mediated transactivation. PMID:10779337

Steroid Receptor Coactivator-1 Knockdown Decreases Synaptic Plasticity and Impairs Spatial Memory in the Hippocampus of Mice.

PubMed

Bian, Chen; Huang, Yan; Zhu, Haitao; Zhao, Yangang; Zhao, Jikai; Zhang, Jiqiang

2018-05-01

Steroids have been demonstrated to play profound roles in the regulation of hippocampal function by acting on their receptors, which need coactivators for their transcriptional activities. Previous studies have shown that steroid receptor coactivator-1 (SRC-1) is the predominant coactivator in the hippocampus, but its exact role and the underlying mechanisms remain unclear. In this study, we constructed SRC-1 RNA interference (RNAi) lentiviruses, injected them into the hippocampus of male mice, and then examined the changes in the expression of selected synaptic proteins, CA1 synapse density, postsynaptic density (PSD) thickness, and in vivo long-term potentiation (LTP). Spatial learning and memory behavior changes were investigated using the Morris water maze. We then transfected the lentiviruses into cultured hippocampal cells and examined the changes in synaptic protein and phospho-cyclic AMP response element-binding protein (pCREB) expression. The in vivo results showed that SRC-1 knockdown significantly decreased the expression of synaptic proteins and CA1 synapse density as well as PSD thickness; SRC-1 knockdown also significantly impaired in vivo LTP and disrupted spatial learning and memory. The in vitro results showed that while the expression of synaptic proteins was significantly decreased by SRC-1 knockdown, pCREB expression was also significantly decreased. The above results suggest a pivotal role of SRC-1 in the regulation of hippocampal synaptic plasticity and spatial learning and memory, strongly indicating SRC-1 may serve as a novel therapeutic target for hippocampus-dependent memory disorders. Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.

Long-Term Memory for Place Learning Is Facilitated by Expression of cAMP Response Element-Binding Protein in the Dorsal Hippocampus

ERIC Educational Resources Information Center

Brightwell, Jennifer J.; Smith, Clayton A.; Neve, Rachael L.; Colombo, Paul J.

2007-01-01

Extensive research has shown that the hippocampus is necessary for consolidation of long-term spatial memory in rodents. We reported previously that rats using a place strategy to solve a cross maze task showed sustained phosphorylation of hippocampus cyclic AMP response element-binding protein (CREB), a transcription factor implicated in…

A novel regulatory circuit in base excision repair involving AP endonuclease 1, Creb1 and DNA polymerase β

PubMed Central

Pei, De-Sheng; Yang, Xiao-Jie; Liu, Wei; Guikema, Jeroen E. J.; Schrader, Carol E.; Strauss, Phyllis R.

2011-01-01

DNA repair is required to maintain genome stability in stem cells and early embryos. At critical junctures, oxidative damage to DNA requires the base excision repair (BER) pathway. Since early zebrafish embryos lack the major polymerase in BER, DNA polymerase ß, repair proceeds via replicative polymerases, even though there is ample polb mRNA. Here, we report that Polb protein fails to appear at the appropriate time in development when AP endonuclease 1 (Apex), the upstream protein in BER, is knocked down. Because polb contains a Creb1 binding site, we examined whether knockdown of Apex affects creb1. Apex knockdown results in loss of Creb1 and Creb complex members but not Creb1 phosphorylation. This effect is independent of p53. Although both apex and creb1 mRNA rescue Creb1 and Polb after Apex knockdown, Apex is not a co-activator of creb1 transcription. This observation has broad significance, as similar results occur when Apex is inhibited in B cells from apex+/− mice. These results describe a novel regulatory circuit involving Apex, Creb1 and Polb and provide a mechanism for lethality of Apex loss in higher eukaryotes. PMID:21172930

VEGF/Flk1 Signaling Cascade Transactivates Etv2 Gene Expression

PubMed Central

Rasmussen, Tara L.; Shi, Xiaozhong; Wallis, Alicia; Kweon, Junghun; Zirbes, Katie M.; Koyano-Nakagawa, Naoko; Garry, Daniel J.

2012-01-01

Previous reports regarding the genetic hierarchy between Ets related protein 71 (Er71/Etv2) and Flk1 is unclear. In the present study, we pursued a genetic approach to define the molecular cascade between Etv2 and Flk1. Using a transgenic Etv2-EYFP reporter mouse, we examined the expression pattern of Etv2 relative to Flk1 in the early conceptus. Etv2-EYFP was expressed in subset of Flk1 positive cells during primitive streak stages, suggesting that Flk1 is upstream of Etv2 during gastrulation. Analysis of reporter gene expression in Flk1 and Etv2 mutant mice further supports the hypothesis that Flk1 is necessary for Etv2 expression. The frequency of cells expressing Flk1 in Etv2 mutants is only modestly altered (21% decrease), whereas expression of the Etv2-EYFP transgenic reporter was severely reduced in the Flk1 null background. We further demonstrate using transcriptional assays that, in the presence of Flk1, the Etv2 promoter is activated by VEGF, the Flk1 ligand. Pharmacological inhibition studies demonstrate that VEGF mediated activation is dependent on p38 MAPK, which activates Creb. We identify the VEGF response element in the Etv2 promoter and demonstrate that Creb binds to this motif by EMSA and ChIP assays. In summary, we provide new evidence that VEGF activates Etv2 by signaling through Flk1, which activates Creb through the p38 MAPK signaling cascade. PMID:23185546

Stimulation of StAR expression by cAMP is controlled by inhibition of highly inducible SIK1 via CRTC2, a co-activator of CREB.

PubMed

Lee, Jinwoo; Tong, Tiegang; Takemori, Hiroshi; Jefcoate, Colin

2015-06-15

In mouse steroidogenic cells the activation of cholesterol metabolism is mediated by steroidogenic acute regulatory protein (StAR). Here, we visualized a coordinated regulation of StAR transcription, splicing and post-transcriptional processing, which are synchronized by salt inducible kinase (SIK1) and CREB-regulated transcription coactivator (CRTC2). To detect primary RNA (pRNA), spliced primary RNA (Sp-RNA) and mRNA in single cells, we generated probe sets by using fluorescence in situ hybridization (FISH). These methods allowed us to address the nature of StAR gene expression and to visualize protein-nucleic acid interactions through direct detection. We show that SIK1 represses StAR expression in Y1 adrenal and MA10 testis cells through inhibition of processing mediated by CRTC2. Digital image analysis matches qPCR analyses of the total cell culture. Evidence is presented for spatially separate accumulation of StAR pRNA and Sp-RNA at the gene loci in the nucleus. These findings establish that cAMP, SIK and CRTC mediate StAR expression through activation of individual StAR gene loci. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

CYCLIC AMP-DEPENDENT PROTEIN KINASE INDUCTION BY POLYCHLORINATED BIPHENYLS (PCBS) STIMULATES CREB PHOSPHORYLATION VIA A CALCIUM-DEPENDENT, PKC-INDEPENDENT PATHWAY IN CORTICAL NEURONS.

EPA Science Inventory

We have previously demonstrated that the PCB mixture, Aroclor 1254 (A1254), increases the phosphorylated form of CREB (pCREB), the cAMP-responsive element binding protein. This transcription factor is important in nervous system development and plasticity. Phosphorylation
of C...

Key roles of Arf small G proteins and biosynthetic trafficking for animal development.

PubMed

Rodrigues, Francisco F; Harris, Tony J C

2017-04-14

Although biosynthetic trafficking can function constitutively, it also functions specifically for certain developmental processes. These processes require either a large increase to biosynthesis or the biosynthesis and targeted trafficking of specific players. We review the conserved molecular mechanisms that direct biosynthetic trafficking, and discuss how their genetic disruption affects animal development. Specifically, we consider Arf small G proteins, such as Arf1 and Sar1, and their coat effectors, COPI and COPII, and how these proteins promote biosynthetic trafficking for cleavage of the Drosophila embryo, the growth of neuronal dendrites and synapses, extracellular matrix secretion for bone development, lumen development in epithelial tubes, notochord and neural tube development, and ciliogenesis. Specific need for the biosynthetic trafficking system is also evident from conserved CrebA/Creb3-like transcription factors increasing the expression of secretory machinery during several of these developmental processes. Moreover, dysfunctional trafficking leads to a range of developmental syndromes.

A novel synthetic phosphodiesterase 5 inhibitor, KJH-1002, ameliorates scopolamine-induced cognitive impairments in mice by activating the cGMP/CREB signaling pathway and attenuating oxidative damage.

PubMed

Zhang, Lijun; Seo, Jae Hong; Li, Huan; Nam, Ghilsoo; Yang, Hyun Ok

2018-05-30

Inhibition of PDE5 has been demonstrated to improve synaptic plasticity and memory via enhancing of cGMP expression, thus activating the cGMP/CREB signaling pathway. This study aimed to investigate the ameliorating effect of PDE5 inhibitor on scopolamine-induced cognitive dysfunction using memory-related behavioral tests and biochemical assays. After the mice were pretreated with PDE5 inhibitor, amnesia was induced by scopolamine administration. The learning and memory abilities of mice were tested using the Morris water maze test, the Y-maze test, the passive avoidance test and the novel object recognition test in sequence. Expression of memory-related bio-molecules and oxidative stress parameters in brain tissue were measured using western blot and spectrophotometry, respectively. KJH-1002, a novel inhibitor of phosphodiesterase 5 (PDE5), was synthesized (IC 50 of 0.059 ±0.04 nmol·L -1 ), and it markedly improved the memory performance impaired by scopolamine in the behavioral tests, indicating a restoration of cognitive function in the mice. Moreover, KJH-1002 increased the cGMP level in the cortex, the scopolamine-reduced expression of phosphorylated cAMP response element binding protein (CREB), extracellular-regulated kinase 1/2 (ERK 1/2), protein kinase B (Akt) and brain-derived neurotrophic factor (BDNF) in the cortex and hippocampus were reversed by KJH-1002 treatment. In addition, KJH-1002 administration increased the activities of superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GR), and decreased the level of malondialdehyde (MDA). KJH-1002 restored cognitive function in scopolamine-induced amnesia mice by activating the cGMP/CREB signaling pathway and attenuating oxidative stress. The beneficial effect of KJH-1002 on cognition suggests its potential as a therapeutic candidate for Alzheimer's disease. This article is protected by copyright. All rights reserved.

Piper sarmentosum Roxb. produces antidepressant-like effects in rodents, associated with activation of the CREB-BDNF-ERK signaling pathway and reversal of HPA axis hyperactivity.

PubMed

Li, Qing; Qu, Fa-Lin; Gao, Yue; Jiang, Yi-Ping; Rahman, Khalid; Lee, Kuo-Hsiung; Han, Ting; Qin, Lu-Ping

2017-03-06

There are many plants of genus Piper which have been reported to induce antidepressant-like effects, Piper sarmentosum (PS) is one of them. PS is a Chinese herbal medicine and a traditional edible vegetable. In the present study, the antidepressant-like effects of PS extracts and the ethyl acetate fraction of PS extracts (PSY) were assessed using the open field test (OFT), forced swimming test (FST), and tail suspension test (TST) in mice. Furthermore, we applied a 4 consecutive weeks of chronic unpredictable mild stress (CUMS) as a model of depression in rats, followed by a sucrose preference test. Then we examined the possible mechanisms of this action. The activity of the hypothalamic-pituitary-adrenal (HPA) axis was evaluated by detecting the serum corticosterone (CORT) concentrations, and the protein expression levels of brain-derived neurotrophic factor (BDNF), the phosphorylated form CREB and ERK1/2 were detected by qRT-PCR or Western blot. The results showed that PS extracts (100, 200mg/kg) and PSY (12.5, 25, 50mg/kg) treatment produced antidepressant-like effects in mice similar to fluoxetine (20mg/kg), indicated by the reduced immobility time in the FST and TST, while both had no influence on the locomotor activity in the OFT. PSY treatment significantly increased sucrose preference and reduced serum CORT levels in CUMS rats. Moreover, PSY up-regulated BDNF protein levels, and increased CREB and ERK phosphorylation levels in the hippocampus on CUMS rats. These findings suggest that the antidepressant-like effects of PS extracts and PSY are mediated, at least in part, by modulating HPA axis, BDNF, CREB and ERK phosphorylation and expression in the hippocampus. Copyright © 2017 Elsevier Ireland Ltd. All rights reserved.

Genome-wide Functional Analysis of CREB/Long-Term Memory-Dependent Transcription Reveals Distinct Basal and Memory Gene Expression Programs

PubMed Central

Lakhina, Vanisha; Arey, Rachel N.; Kaletsky, Rachel; Kauffman, Amanda; Stein, Geneva; Keyes, William; Xu, Daniel; Murphy, Coleen T.

2014-01-01

SUMMARY Induced CREB activity is a hallmark of long-term memory, but the full repertoire of CREB transcriptional targets required specifically for memory is not known in any system. To obtain a more complete picture of the mechanisms involved in memory, we combined memory training with genome-wide transcriptional analysis of C. elegans CREB mutants. This approach identified 757 significant CREB/memory-induced targets and confirmed the involvement of known memory genes from other organisms, but also suggested new mechanisms and novel components that may be conserved through mammals. CREB mediates distinct basal and memory transcriptional programs at least partially through spatial restriction of CREB activity: basal targets are regulated primarily in nonneuronal tissues, while memory targets are enriched for neuronal expression, emanating from CREB activity in AIM neurons. This suite of novel memory-associated genes will provide a platform for the discovery of orthologous mammalian long-term memory components. PMID:25611510

The cellular transcription factor CREB corresponds to activating transcription factor 47 (ATF-47) and forms complexes with a group of polypeptides related to ATF-43.

PubMed

Hurst, H C; Masson, N; Jones, N C; Lee, K A

1990-12-01

Promoter elements containing the sequence motif CGTCA are important for a variety of inducible responses at the transcriptional level. Multiple cellular factors specifically bind to these elements and are encoded by a multigene family. Among these factors, polypeptides termed activating transcription factor 43 (ATF-43) and ATF-47 have been purified from HeLa cells and a factor referred to as cyclic AMP response element-binding protein (CREB) has been isolated from PC12 cells and rat brain. We demonstrated that CREB and ATF-47 are identical and that CREB and ATF-43 form protein-protein complexes. We also found that the cis requirements for stable DNA binding by ATF-43 and CREB are different. Using antibodies to ATF-43 we have identified a group of polypeptides (ATF-43) in the size range from 40 to 43 kDa. ATF-43 polypeptides are related by their reactivity with anti-ATF-43, DNA-binding specificity, complex formation with CREB, heat stability, and phosphorylation by protein kinase A. Certain cell types vary in their ATF-43 complement, suggesting that CREB activity is modulated in a cell-type-specific manner through interaction with ATF-43. ATF-43 polypeptides do not appear simply to correspond to the gene products of the ATF multigene family, suggesting that the size of the ATF family at the protein level is even larger than predicted from cDNA-cloning studies.

Ketamine administered pregnant rats impair learning and memory in offspring via the CREB pathway.

PubMed

Li, Xinran; Guo, Cen; Li, Yanan; Li, Lina; Wang, Yuxin; Zhang, Yiming; Li, Yue; Chen, Yu; Liu, Wenhan; Gao, Li

2017-05-16

Ketamine has been reported to impair the capacity for learning and memory. This study examined whether these capacities were also altered in the offspring and investigated the role of the CREB signaling pathway in pregnant rats, subjected to ketamine-induced anesthesia. On the 14th day of gestation (P14), female rats were anesthetized for 3 h via intravenous ketamine injection (200 mg/Kg). Morris water maze task, contextual and cued fear conditioning, and olfactory tasks were executed between the 25th to 30th day after birth (B25-30) on rat pups, and rats were sacrificed on B30. Nerve density and dendritic spine density were examined via Nissl's and Golgi staining. Simultaneously, the contents of Ca2+/Calmodulin-Dependent Protein Kinase II (CaMKII), p-CaMKII, CaMKIV, p-CaMKIV, Extracellular Regulated Protein Kinases (ERK), p-ERK, Protein Kinase A (PKA), p-PKA, cAMP-Response Element Binding Protein (CREB), p-CREB, and Brain Derived Neurotrophic Factor (BDNF) were detected in the hippocampus. We pretreated PC12 cells with both PKA inhibitor (H89) and ERK inhibitor (SCH772984), thus detecting levels of ERK, p-ERK, PKA, p-PKA, p-CREB, and BDNF. The results revealed that ketamine impaired the learning ability and spatial as well as conditioned memory in the offspring, and significantly decreased the protein levels of ERK, p-ERK, PKA, p-PKA, p-CREB, and BDNF. We found that ERK and PKA (but not CaMKII or CaMKIV) have the ability to regulate the CREB-BDNF pathway during ketamine-induced anesthesia in pregnant rats. Furthermore, ERK and PKA are mutually compensatory for the regulation of the CREB-BDNF pathway.

Serine 133 Phosphorylation Is Not Required for Hippocampal CREB-Mediated Transcription and Behavior

ERIC Educational Resources Information Center

Brian, Lisa A.; Lee, Bridgin G.; Lelay, John; Kaestner, Klaus H.; Blendy, Julie A.

2015-01-01

The cAMP response element (CRE)-binding protein, CREB, is a transcription factor whose activity in the brain is critical for long-term memory formation. Phosphorylation of Ser133 in the kinase-inducible domain (KID), that in turn leads to the recruitment of the transcriptional coactivator CREB-binding protein (CBP), is thought to mediate the…

Neural Plasticity Associated with Hippocampal PKA-CREB and NMDA Signaling Is Involved in the Antidepressant Effect of Repeated Low Dose of Yueju Pill on Chronic Mouse Model of Learned Helplessness

PubMed Central

Zou, Zhilu; Chen, Yin; Shen, Qinqin; Guo, Xiaoyan; Zhang, Yuxuan

2017-01-01

Yueju pill is a traditional Chinese medicine formulated to treat syndromes of mood disorders. Here, we investigated the therapeutic effect of repeated low dose of Yueju in the animal model mimicking clinical long-term depression condition and the role of neural plasticity associated with PKA- (protein kinase A-) CREB (cAMP response element binding protein) and NMDA (N-methyl-D-aspartate) signaling. We showed that a single low dose of Yueju demonstrated antidepressant effects in tests of tail suspension, forced swim, and novelty-suppressed feeding. A chronic learned helplessness (LH) protocol resulted in a long-term depressive-like condition. Repeated administration of Yueju following chronic LH remarkably alleviated all of depressive-like symptoms measured, whereas conventional antidepressant fluoxetine only showed a minor improvement. In the hippocampus, Yueju and fluoxetine both normalized brain-derived neurotrophic factor (BDNF) and PKA level. Only Yueju, not fluoxetine, rescued the deficits in CREB signaling. The chronic LH upregulated the expression of NMDA receptor subunits NR1, NR2A, and NR2B, which were all attenuated by Yueju. Furthermore, intracerebraventricular administration of NMDA blunted the antidepressant effect of Yueju. These findings supported the antidepressant efficacy of repeated routine low dose of Yueju in a long-term depression model and the critical role of CREB and NMDA signaling. PMID:29075536

Neural Plasticity Associated with Hippocampal PKA-CREB and NMDA Signaling Is Involved in the Antidepressant Effect of Repeated Low Dose of Yueju Pill on Chronic Mouse Model of Learned Helplessness.

PubMed

Zou, Zhilu; Chen, Yin; Shen, Qinqin; Guo, Xiaoyan; Zhang, Yuxuan; Chen, Gang

2017-01-01

Yueju pill is a traditional Chinese medicine formulated to treat syndromes of mood disorders. Here, we investigated the therapeutic effect of repeated low dose of Yueju in the animal model mimicking clinical long-term depression condition and the role of neural plasticity associated with PKA- (protein kinase A-) CREB (cAMP response element binding protein) and NMDA (N-methyl-D-aspartate) signaling. We showed that a single low dose of Yueju demonstrated antidepressant effects in tests of tail suspension, forced swim, and novelty-suppressed feeding. A chronic learned helplessness (LH) protocol resulted in a long-term depressive-like condition. Repeated administration of Yueju following chronic LH remarkably alleviated all of depressive-like symptoms measured, whereas conventional antidepressant fluoxetine only showed a minor improvement. In the hippocampus, Yueju and fluoxetine both normalized brain-derived neurotrophic factor (BDNF) and PKA level. Only Yueju, not fluoxetine, rescued the deficits in CREB signaling. The chronic LH upregulated the expression of NMDA receptor subunits NR1, NR2A, and NR2B, which were all attenuated by Yueju. Furthermore, intracerebraventricular administration of NMDA blunted the antidepressant effect of Yueju. These findings supported the antidepressant efficacy of repeated routine low dose of Yueju in a long-term depression model and the critical role of CREB and NMDA signaling.

Adipocyte iron regulates leptin and food intake

PubMed Central

Gao, Yan; Li, Zhonggang; Gabrielsen, J. Scott; Simcox, Judith A.; Lee, Soh-hyun; Jones, Deborah; Cooksey, Bob; Stoddard, Gregory; Cefalu, William T.; McClain, Donald A.

2015-01-01

Dietary iron supplementation is associated with increased appetite. Here, we investigated the effect of iron on the hormone leptin, which regulates food intake and energy homeostasis. Serum ferritin was negatively associated with serum leptin in a cohort of patients with metabolic syndrome. Moreover, the same inverse correlation was observed in mice fed a high-iron diet. Adipocyte-specific loss of the iron exporter ferroportin resulted in iron loading and decreased leptin, while decreased levels of hepcidin in a murine hereditary hemochromatosis (HH) model increased adipocyte ferroportin expression, decreased adipocyte iron, and increased leptin. Treatment of 3T3-L1 adipocytes with iron decreased leptin mRNA in a dose-dependent manner. We found that iron negatively regulates leptin transcription via cAMP-responsive element binding protein activation (CREB activation) and identified 2 potential CREB-binding sites in the mouse leptin promoter region. Mutation of both sites completely blocked the effect of iron on promoter activity. ChIP analysis revealed that binding of phosphorylated CREB is enriched at these two sites in iron-treated 3T3-L1 adipocytes compared with untreated cells. Consistent with the changes in leptin, dietary iron content was also directly related to food intake, independently of weight. These findings indicate that levels of dietary iron play an important role in regulation of appetite and metabolism through CREB-dependent modulation of leptin expression. PMID:26301810

Regulation of Energy Stores and Feeding by Neuronal and Peripheral CREB Activity in Drosophila

PubMed Central

Iijima, Koichi; Zhao, LiJuan; Shenton, Christopher; Iijima-Ando, Kanae

2009-01-01

The cAMP-responsive transcription factor CREB functions in adipose tissue and liver to regulate glycogen and lipid metabolism in mammals. While Drosophila has a homolog of mammalian CREB, dCREB2, its role in energy metabolism is not fully understood. Using tissue-specific expression of a dominant-negative form of CREB (DN-CREB), we have examined the effect of blocking CREB activity in neurons and in the fat body, the primary energy storage depot with functions of adipose tissue and the liver in flies, on energy balance, stress resistance and feeding behavior. We found that disruption of CREB function in neurons reduced glycogen and lipid stores and increased sensitivity to starvation. Expression of DN-CREB in the fat body also reduced glycogen levels, while it did not affect starvation sensitivity, presumably due to increased lipid levels in these flies. Interestingly, blocking CREB activity in the fat body increased food intake. These flies did not show a significant change in overall body size, suggesting that disruption of CREB activity in the fat body caused an obese-like phenotype. Using a transgenic CRE-luciferase reporter, we further demonstrated that disruption of the adipokinetic hormone receptor, which is functionally related to mammalian glucagon and β-adrenergic signaling, in the fat body reduced CRE-mediated transcription in flies. This study demonstrates that CREB activity in either neuronal or peripheral tissues regulates energy balance in Drosophila, and that the key signaling pathway regulating CREB activity in peripheral tissue is evolutionarily conserved. PMID:20041126

RGS10 exerts a neuroprotective role through the PKA/c-AMP response-element (CREB) pathway in dopaminergic neuron-like cells

PubMed Central

Lee, Jae-Kyung; Chung, Jaegwon; Druey, Kirk M.; Tansey, Malú G.

2012-01-01

Regulator of G-protein signaling-10 (RGS10) is a GTPase activating protein (GAP) for Gαi/q/z subunits that is highly expressed in the immune system and in a broad range of brain regions including the hippocampus, striatum, dorsal raphe, and ventral midbrain. Previously, we reported that RGS10-null mice display increased vulnerability to chronic systemic inflammation-induced degeneration of nigral dopaminergic (DA) neurons. Given that RGS10 is expressed in DA neurons, we investigated the extent to which RGS10 regulates cell survival under conditions of inflammatory stress. Because of the inherent limitations associated with use of primary DA neurons for biochemical analyses, we employed a well-characterized ventral mesencephalon DA neuroblastoma cell line (MN9D) for our studies. We found that stable over-expression of RGS10 rendered them resistant to TNF-induced cytotoxicity; whereas MN9D cells expressing mutant RGS10-S168A (which is resistant to phosphorylation by protein kinase A (PKA) at a serine residue that promotes its nuclear translocation) showed similar sensitivity to TNF as the parental MN9D cells. Using biochemical and pharmacological approaches, we identified protein kinase A (PKA) and the downstream phospho-cAMP response element-binding (CREB) signaling pathway (and ruled out ERK 1/2, JNK, and NFkB) as key mediators of the neuroprotective effect of RGS10 against inflammatory stress. PMID:22564151

NAc Shell Arc/Arg3.1 Protein Mediates Reconsolidation of Morphine CPP by Increased GluR1 Cell Surface Expression: Activation of ERK-Coupled CREB is Required

PubMed Central

Lv, Xiu-Fang; Sun, Lin-Lin; Han, Ji-Sheng

2015-01-01

Background: Relapse into drug abuse evoked by reexposure to the drug-associated context has been a primary problem in the treatment of drug addiction. Disrupting the reconsolidation of drug-related context memory would therefore limit the relapse susceptibility. Methods: Morphine conditioned place preference (CPP) was used to assess activity-regulated cytoskeleton-associated protein (Arc/Arg3.1) and correlative molecule expression in the Nucleus accumbens (NAc) shell during the reconsolidation of morphine CPP. U0126 and Arc/Arg3.1 antisense oligodeoxynucleotide were adapted to evaluate the role and the underlying mechanism of Arc/Arg3.1 during the reconsolidation. Results: The retrieval of morphine CPP in rats specifically increased the Arc/Arg3.1 protein level in the NAc shell, accompanied simultaneously by increases in the phosphorylation of extracellular signal-regulated kinase1/2 (pERK1/2), the phosphorylation of Cyclic Adenosine monophosphate (cAMP) response element-binding (pCREB), and the up-regulation of the membrane α-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptors GluR1 subunit level. Intra-NAc shell infusion U0126, an inhibitor of the Mitogen-activated protein kinase kinase (MEK), prevented the retrieval-induced up-regulation of pERK1/2, pCREB, Arc/Arg3.1, and membrane GluR1 immediately after retrieval of morphine CPP. The effect of disrupting the reconsolidation of morphine CPP by U0126 could last for at least 14 days, and could not be evoked by a priming injection of morphine. Furthermore, the specific knockdown of Arc/Arg3.1 in the NAc shell decreased the membrane GluR1 level, and impaired both the reconsolidation and the reinstatement of morphine CPP. Conclusions: Arc/Arg3.1 in the NAc shell mediates the reconsolidation of morphine-associated context memory via up-regulating the level of membrane of GluR1, for which the local activation of the ERK-CREB signal pathway, as an upstream mechanism of Arc/Arg3.1, is required. PMID:25746394

Rhubarb tannins extract inhibits the expression of aquaporins 2 and 3 in magnesium sulphate-induced diarrhoea model.

PubMed

Liu, Chunfang; Zheng, Yanfang; Xu, Wen; Wang, Hui; Lin, Na

2014-01-01

Tannins, a group of major active components of Chinese rhubarb and widely distributed in nature, have a significant antidiarrhoeal activity. Aquaporins (AQPs) 2 and 3 play important roles in regulating water transfer during diarrhoea. The present study aims to determine the effect of the total tannins extract of rhubarb on aquaporins (AQPs) 2 and 3 in diarrhoea mice and HT-29 cells both induced by magnesium sulphate (MgSO4). Our results showed that rhubarb tannins extract (RTE) significantly decreased the faecal water content in colon and evaluation index of defecation of diarrhoea mice. Interestingly, RTE could markedly reduce the mRNA and protein expression levels of AQPs 2 and 3 in apical and lateral mucosal epithelial cells in the colons of diarrhoea mice and HT-29 cells both induced by MgSO4 in a dose-dependent manner. Furthermore, RTE suppressed the production of cyclic monophosphate- (cAMP-) dependent protein kinase A catalytic subunits α (PKA C-α) and phosphorylated cAMP response element-binding protein (p-CREB, Ser133) in MgSO4-induced HT-29 cells. Our data showed for the first time that RTE inhibit AQPs 2 and 3 expression in vivo and in vitro via downregulating PKA/p-CREB signal pathway, which accounts for the antidiarrhoeal effect of RTE.

Hunting increases phosphorylation of calcium/calmodulin-dependent protein kinase type II in adult barn owls.

PubMed

Nichols, Grant S; DeBello, William M

2015-01-01

Juvenile barn owls readily adapt to prismatic spectacles, whereas adult owls living under standard aviary conditions do not. We previously demonstrated that phosphorylation of the cyclic-AMP response element-binding protein (CREB) provides a readout of the instructive signals that guide plasticity in juveniles. Here we investigated phosphorylation of calcium/calmodulin-dependent protein kinase II (pCaMKII) in both juveniles and adults. In contrast to CREB, we found no differences in pCaMKII expression between prism-wearing and control juveniles within the external nucleus of the inferior colliculus (ICX), the major site of plasticity. For prism-wearing adults that hunted live mice and are capable of adaptation, expression of pCaMKII was increased relative to prism-wearing adults that fed passively on dead mice and are not capable of adaptation. This effect did not bear the hallmarks of instructive information: it was not localized to rostral ICX and did not exhibit a patchy distribution reflecting discrete bimodal stimuli. These data are consistent with a role for CaMKII as a permissive rather than an instructive factor. In addition, the paucity of pCaMKII expression in passively fed adults suggests that the permissive default setting is "off" in adults.

Synergistic Activation of Steroidogenic Acute Regulatory Protein Expression and Steroid Biosynthesis by Retinoids: Involvement of cAMP/PKA Signaling

PubMed Central

Manna, Pulak R.; Slominski, Andrzej T.; King, Steven R.; Stetson, Cloyce L.

2014-01-01

Both retinoic acid receptors (RARs) and retinoid X receptors (RXRs) mediate the action of retinoids that play important roles in reproductive development and function, as well as steroidogenesis. Regulation of steroid biosynthesis is principally mediated by the steroidogenic acute regulatory protein (StAR); however, the modes of action of retinoids in the regulation of steroidogenesis remain obscure. In this study we demonstrate that all-trans retinoic acid (atRA) enhances StAR expression, but not its phosphorylation (P-StAR), and progesterone production in MA-10 mouse Leydig cells. Activation of the protein kinase A (PKA) cascade, by dibutyrl-cAMP or type I/II PKA analogs, markedly increased retinoid-responsive StAR, P-StAR, and steroid levels. Targeted silencing of endogenous RARα and RXRα, with small interfering RNAs, resulted in decreases in 9-cis RA-stimulated StAR and progesterone levels. Truncation of and mutational alterations in the 5′-flanking region of the StAR gene demonstrated the importance of the −254/−1-bp region in retinoid responsiveness. An oligonucleotide probe encompassing an RXR/liver X receptor recognition motif, located within the −254/−1-bp region, specifically bound MA-10 nuclear proteins and in vitro transcribed/translated RXRα and RARα in EMSAs. Transcription of the StAR gene in response to atRA and dibutyrl-cAMP was influenced by several factors, its up-regulation being dependent on phosphorylation of cAMP response-element binding protein (CREB). Chromatin immunoprecipitation studies revealed the association of phosphorylation of CREB, CREB binding protein, RXRα, and RARα to the StAR promoter. Further studies elucidated that hormone-sensitive lipase plays an important role in atRA-mediated regulation of the steroidogenic response that involves liver X receptor signaling. These findings delineate the molecular events by which retinoids influence cAMP/PKA signaling and provide additional and novel insight into the regulation of StAR expression and steroidogenesis in mouse Leydig cells. PMID:24265455

Salicylic acid and aspirin inhibit the activity of RSK2 kinase and repress RSK2-dependent transcription of cyclic AMP response element binding protein- and NF-kappa B-responsive genes.

PubMed

Stevenson, M A; Zhao, M J; Asea, A; Coleman, C N; Calderwood, S K

1999-11-15

Sodium salicylate (NaSal) and other nonsteroidal anti-inflammatory drugs (NSAIDs) coordinately inhibit the activity of NF-kappa B, activate heat shock transcription factor 1 and suppress cytokine gene expression in activated monocytes and macrophages. Because our preliminary studies indicated that these effects could be mimicked by inhibitors of signal transduction, we have studied the effects of NSAIDs on signaling molecules potentially downstream of LPS receptors in activated macrophages. Our findings indicate that ribosomal S6 kinase 2 (RSK2), a 90-kDa ribosomal S6 kinase with a critical role as an effector of the RAS-mitogen-activated protein kinase pathway and a regulator of immediate early gene transcription is a target for inhibition by the NSAIDs. NSAIDs inhibited the activity of purified RSK2 kinase in vitro and of RSK2 in mammalian cells and suppressed the phosphorylation of RSK2 substrates cAMP response element binding protein (CREB) and I-kappa B alpha in vivo. Additionally, NaSal inhibited the phosphorylation by RSK2 of CREB and I-kappa B alpha on residues crucial for their transcriptional activity in vivo and thus repressed CREB and NF-kappa B-dependent transcription. These experiments suggest that RSK2 is a target for NSAIDs in the inhibition of monocyte-specific gene expression and indicate the importance of RSK2 and related kinases in cell regulation, indicating a new area for anti-inflammatory drug discovery.

Involvement of sigma-1 receptor in astrocyte activation induced by methamphetamine via up-regulation of its own expression.

PubMed

Zhang, Yuan; Lv, Xuan; Bai, Ying; Zhu, Xinjian; Wu, Xiaodong; Chao, Jie; Duan, Ming; Buch, Shilpa; Chen, Ling; Yao, Honghong

2015-02-17

Although it has been documented that methamphetamine induces astrocyte activation, the mechanism(s) underlying this effect remain poorly understood. We thus sought to examine the molecular mechanisms involved in methamphetamine-mediated activation of astrocytes with a focus on the role of sigma-1 receptor (σ-1R) in this process. The expression of σ-1R and glial fibrillary acidic protein (GFAP) was examined by reverse transcription PCR (RT-PCR), real-time PCR, Western blot, and immunofluorescent staining; phosphorylation of cell signaling pathways was detected by Western blot analysis. Immunoprecipitation was used to determine the interaction between σ-1R and p-Src. Chromatin immunoprecipitation (ChIP) assay was employed to discern the binding of cAMP-response element-binding protein (CREB) with the promoter of σ-1R. The role of σ-1R in astrocyte activation was further validated in σ-1R knockout (KO) mice by Western blot combined with immunofluorescent staining. Exposure of primary rat astrocytes to methamphetamine increased the expression of σ-1R via the activation of Src, ERK mitogen-activated protein kinase, and downstream CREB pathways. Subsequently, CREB translocated into nucleus and interacted with the promoter of σ-1R resulting in increased expression of σ-1R with a concomitant increase in expression of GFAP. This effect was inhibited in cells treated with the σ-1R antagonist-BD1047, thereby implicating the role of σ-1R in the activation of astrocytes. In vivo relevance of these findings was further corroborated in σ-1R KO mice that were administered methamphetamine. In the methamphetamine administered mice, there was a failure of the drug to induce activation of astrocytes, an effect that was evident in wild-type (WT) mice exposed to methamphetamine. The study presented herein demonstrates that methamphetamine-mediated activation of astrocytes involved up-regulation of σ-1R through a positive-feedback mechanism. Understanding the regulation of σ-1R expression could provide insights into the development of potential therapeutic strategies for astrocyte activation induced by methamphetamine.

The Rewarding and Locomotor-Sensitizing Effects of Repeated Cocaine Administration are Distinct and Separable in Mice

PubMed Central

Riday, Thorfinn T.; Kosofsky, Barry E.; Malanga, C.J.

2011-01-01

Repeated psychostimulant exposure progressively increases their potency to stimulate motor activity in rodents. This behavioral or locomotor sensitization is considered a model for some aspects of drug addiction in humans, particularly drug craving during abstinence. However, the role of increased motor behavior in drug reward remains incompletely understood. Intracranial self-stimulation (ICSS) was measured concurrently with locomotor activity to determine if acute intermittent cocaine administration had distinguishable effects on motor behavior and perception of brain stimulation-reward (BSR) in the same mice. Sensitization is associated with changes in neuronal activity and glutamatergic neurotransmission in brain reward circuitry. Expression of AMPA receptor subunits (GluR1 and GluR2) and CRE binding protein (CREB) was measured in the ventral tegmental area (VTA), dorsolateral striatum (STR) and nucleus accumbens (NAc) before and after a sensitizing regimen of cocaine, with and without ICSS. Repeated cocaine administration sensitized mice to its locomotor stimulating effects but not its ability to potentiate BSR. ICSS increased GluR1 in the VTA but not NAc or STR, demonstrating selective changes in protein expression with electrical stimulation of discrete brain structures. Repeated cocaine reduced GluR1, GluR2 and CREB expression in the NAc, and reductions of GluR1 and GluR2 but not CREB were further enhanced by ICSS. These data suggest that the effects of repeated cocaine exposure on reward and motor processes are dissociable in mice, and that reduction of excitatory neurotransmission in the NAc may predict altered motor function independently from changes in reward perception. PMID:22197517

Epigenetic modification of miR-10a regulates renal damage by targeting CREB1 in type 2 diabetes mellitus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shan, Qun, E-mail: shanp@jsnu.edu.cn; Zheng, Guiho

Emerging evidence has shown that microRNA-mediated gene expression modulation plays a crucial role in the pathogenesis of type 2 diabetes mellitus, but the novel miRNAs involved in type 2 diabetes and its functional regulatory mechanisms still need to be determined. In this study, we assessed the role of miR-10a in extracellular matrix accumulation in the kidney of diabetic mellitus induced by combining administration of chronic high fat diet (HFD) and low dosage of streptozotocin (STZ, 35 mg/kg). Here, we found that HFD/STZ administration decreased the level of microRNA (miR-10a) expression in ICR strain mice. Overexpression of miR-10a alleviated the increasedmore » ratio of urine albumin-to-creatinine (ACR) ratio of HFD/STZ mice. In contrast, knockdown of miR-10a increased the ratio of kidney ACR in naïve mice. Furthermore, cAMP response element binding protein 1 (CREB1) was validated as a target of miR-10a in vitro and in vivo. CREB1 and its downstream fibronectin (FN, extracellular matrix) were increased in HFD/STZ-treated mice, which was reversed by kidney miR-10a overexpression. The content of CREB1 and FN was increased by miR-10a knockdown in kidney of naïve mice. Furthermore, histone deacetylase 3 (HDAC3) was revealed to be increased in kidney of HFD/STZ mice, accompanied with the augmentation of ACR ratio and FN level. Knockdown of HDAC3 with siRNA significantly caused the increase of miR-10a, resulting in the decrease in CREB1 and FN expression in kidney of HFD/STZ mice. Contrarily, HDAC3 overexpression mediated by lentivirus decreased miR-10a content, and enhanced ACR value, CREB1 and FN formation in naïve mice. Collectively, these results elucidate that HDAC3/miR-10a/CREB1 serves as a new mechanism underlying kidney injury, providing potential therapeutic targets in type 2 diabetes. - Highlights: • Diabetes induces the decrease of miR-10a level in the kidney. • MiR-10a overexpression improves kidney damage of diabetes. • MiR-10a targeting CREB1/FN implicates in kidney impairment. • HDAC3 regulates kidney damage by epigenetically modulating miR-10a.« less

Transcriptional regulation of miR-15b by c-Rel and CREB in Japanese encephalitis virus infection

PubMed Central

Zhu, Bibo; Ye, Jing; Ashraf, Usama; Li, Yunchuan; Chen, Huanchun; Song, Yunfeng; Cao, Shengbo

2016-01-01

MicroRNAs (miRNAs) have been well known to play diverse roles in viral infection at the level of posttranscriptional repression. However, much less is understood about the mechanism by which miRNAs are regulated during viral infection. It is likely that both host and virus contain factors to modulate miRNA expression. Here we report the up-regulation of microRNA-15b (miR-15b) in vitro upon infection with Japanese encephalitis virus (JEV). Analysis of miR-15b precursor, pri-miR-15b and pre-miR-15b, suggest that the regulation occurs transcriptionally. Further, we identified the transcriptional regulatory region of miR-15b that contains consensus binding motif for NF-κB subunit c-Rel and cAMP-response element binding protein (CREB), which are known as transcription factor to regulate gene expression. By promoter fusion and mutational analyses, we demonstrated that c-Rel and CREB bind directly to the promoter elements of miR-15b, which are responsible for miR-15b transcription in response to JEV infection. Finally, we showed that pharmacological inhibition of ERK and NF-κB signaling pathway blocked induction of miR-15b in JEV infection, suggesting important roles of ERK and NF-κB pathway in the regulation of miR-15b gene. Therefore, our observations indicate that induced expression of miR-15b is modulated by c-Rel and CREB in response to JEV infection. PMID:26931521

Preventive role of social interaction for cocaine conditioned place preference: correlation with FosB/DeltaFosB and pCREB expression in rat mesocorticolimbic areas

PubMed Central

El Rawas, Rana; Klement, Sabine; Salti, Ahmad; Fritz, Michael; Dechant, Georg; Saria, Alois; Zernig, Gerald

2012-01-01

The worsening of drug abuse by drug-associated social interaction is a well-studied phenomenon. In contrast, the molecular mechanisms of the beneficial effect of social interaction, if offered as a mutually exclusive choice to drugs of abuse, are under-investigated. In a rat place preference conditioning (CPP) paradigm, four 15 min episodes of social interaction with a gender- and weight-matched male early-adult conspecific inhibited cocaine-induced reinstatement of cocaine CPP, a model of relapse. These protective effects of social interaction were paralleled by a reduced activation, as assessed by Zif268 expression, in brain areas known to play pivotal roles in drug-seeking behavior. Here we show that social interaction during extinction of cocaine CPP also reduced cocaine-CPP-stimulated FosB expression in the nucleus accumbens shell and core. In addition, social interaction during cocaine CPP extinction increased pCREB (cAMP response element binding protein) expression in the nucleus accumbens shell and the cingulate cortex area 1 (Cg1). Our results show that FosB and pCREB may be implicated in the protective effect of social interaction against cocaine-induced reinstatement of CPP. Thus, social interaction, if offered in a context that is clearly distinct from the previously drug-associated one, may profoundly inhibit relapse to cocaine addiction. PMID:22403532

Hyperforin and Miquelianin from St. John's Wort Attenuate Gene Expression in Neuronal Cells After Dexamethasone-Induced Stress.

PubMed

Verjee, Sheela; Weston, Anna; Kolb, Christiane; Kalbhenn-Aziz, Heba; Butterweck, Veronika

2018-07-01

Dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis plays an important part in the development of depressive symptoms. In this study, the effects of a commercial St. John's wort extract (STW3-VI), hyperforin, miquelianin, and the selective serotonin reuptake inhibitor citalopram on the expression of genes relevant to HPA axis function were investigated in human neuronal cells. SH-SY5Y cells were treated with STW3-VI (20 µg/mL), hyperforin (1 µM), miquelianin (10 µM), or citalopram (10 µM) in the presence of the glucocorticoid receptor agonist dexamethasone (DEX,10 µM) for 6 h and 48 h, respectively. Quantitative real-time polymerase chain reaction was used to determine the expression of FKBP5 (FK506 binding protein 51), CREB (cAMP responsive element binding protein), GRIK4 (glutamate ionotropic receptor kainate type subunit 4), VEGF (vascular endothelial growth factor), NET (norepinephrine transporter), and ARRB ( β -arrestins), promising biomarkers of antidepressant therapy. Using DEX to mimic stress conditions, it was shown that the gene expression pattern of FKBP5, CREB, GRIK4, VEGF, NET, and ARRB2 in SH-SY5Y cells is time- and treatment-dependent. Most pronounced effects were observed for FKBP5: after 6 h of co-incubation, only STW3-VI could reverse the DEX-induced increase in FKBP5 expression, and after 48 h, citalopram, miquelianin, and hyperforin also reversed the glucocorticoid-induced increase in FKBP5 mRNA expression. The effects observed on FKBP5, CREB, GRIK4, VEGF, NET, and ARRB2 are in good correlation with published data, suggesting that this in vitro model could be used to screen the responsiveness of antidepressants under stress conditions. Georg Thieme Verlag KG Stuttgart · New York.

Developmental Decline in the MicroRNA 199a (miR-199a)/miR-214 Cluster in Human Fetal Lung Promotes Type II Cell Differentiation by Upregulating Key Transcription Factors.

PubMed

Mishra, Ritu; Benlhabib, Houda; Guo, Wei; Lerma Cervantes, Connie B; Mendelson, Carole R

2018-06-01

The major surfactant protein, SP-A (a product of the SFTPA gene), serves as a marker of type II pneumocyte differentiation and surfactant synthesis. SFTPA expression in cultured human fetal lung (HFL) epithelial cells is upregulated by hormones that increase cyclic AMP (cAMP) and activate TTF-1/NKX2.1 and NF-κB. To further define mechanisms for type II cell differentiation and induction of SP-A, we investigated roles of microRNAs (miRNAs). Using microarray to identify differentially expressed miRNAs in HFL epithelial cells during type II cell differentiation in culture, we observed that members of the miRNA 199a (miR-199a)/miR-214 cluster were significantly downregulated during differentiation. Validated and predicted targets of miR-199a-3p/miR-199a-5p and miR-214, which serve roles in type II cell differentiation (COX-2, NF-κB p50/p65, and CREB1), and the CREB1 target, C/EBPβ, were coordinately upregulated. Accordingly, overexpression of miR-199a-5p, miR-199a-3p, or miR-214 mimics in cultured HFL epithelial cells decreased COX-2, NF-κB p50/p65, CREB1, and C/EBPβ proteins, with an associated inhibition of SP-A expression. Interestingly, overexpression of the EMT factor, ZEB1, which declines during cAMP-induced type II cell differentiation, increased pri-miR-199a and reduced the expression of the targets NF-κB/p50 and COX-2. Collectively, these findings suggest that the developmental decline in miR-199a/miR-214 in HFL causes increased expression of critical targets that enhance type II cell differentiation and SP-A expression. Copyright © 2018 American Society for Microbiology.

DA-9801 promotes neurite outgrowth via ERK1/2-CREB pathway in PC12 cells.

PubMed

Won, Jong Hoon; Ahn, Kyong Hoon; Back, Moon Jung; Ha, Hae Chan; Jang, Ji Min; Kim, Ha Hyung; Choi, Sang-Zin; Son, Miwon; Kim, Dae Kyong

2015-01-01

In the present study, we examined the mechanisms underlying the effect of DA-9801 on neurite outgrowth. We found that DA-9801 elicits its effects via the mitogen-activated protein kinase (MEK) extracellular signal-regulated kinase (ERK)1/2-cAMP response element-binding protein (CREB) pathway. DA-9801, an extract from a mixture of Dioscorea japonica and Dioscorea nipponica, was reported to promote neurite outgrowth in PC12 cells. The effects of DA-9801 on cell viability and expression of neuronal markers were evaluated in PC12 cells. To investigate DA-9801 action, specific inhibitors targeting the ERK signaling cascade were used. No cytotoxicity was observed in PC12 cells at DA-9801 concentrations of less than 30 µg/mL. In the presence of nerve growth factor (NGF, 2 ng/mL), DA-9801 promoted neurite outgrowth and increased the relative mRNA levels of neurofilament-L (NF-L), a marker of neuronal differentiation. The Raf-1 inhibitor GW5074 and MEK inhibitor PD98059 significantly attenuated DA-9801-induced neurite outgrowth. Additionally, the MEK1 and MEK2 inhibitor SL327 significantly attenuated the increase in the percentage of neurite-bearing PC12 cells induced by DA-9801 treatment. Conversely, the selective p38 mitogen-activated protein kinase inhibitor SB203580 did not attenuate the DA-9801 treatment-induced increase in the percentage of neurite-bearing PC12 cells. DA-9801 enhanced the phosphorylation of ERK1/2 and CREB in PC12 cells incubated with and without NGF. Pretreatment with PD98059 blocked the DA-9801-induced phosphorylation of ERK1/2 and CREB. In conclusion, DA-9801 induces neurite outgrowth by affecting the ERK1/2-CREB signaling pathway. Insights into the mechanism underlying this effect of DA-9801 may suggest novel potential strategies for the treatment of peripheral neuropathy.

Distinctive Roles for Amygdalar CREB in Reconsolidation and Extinction of Fear Memory

ERIC Educational Resources Information Center

Tronson, Natalie C.; Wiseman, Shari L.; Neve, Rachael L.; Nestler, Eric J.; Olausson, Peter; Taylor, Jane R.

2012-01-01

Cyclic AMP response element binding protein (CREB) plays a critical role in fear memory formation. Here we determined the role of CREB selectively within the amygdala in reconsolidation and extinction of auditory fear. Viral overexpression of the inducible cAMP early repressor (ICER) or the dominant-negative mCREB, specifically within the lateral…

The neurotoxicant PCB-95 by increasing the neuronal transcriptional repressor REST down-regulates caspase-8 and increases Ripk1, Ripk3 and MLKL expression determining necroptotic neuronal death.

PubMed

Guida, Natascia; Laudati, Giusy; Serani, Angelo; Mascolo, Luigi; Molinaro, Pasquale; Montuori, Paolo; Di Renzo, Gianfranco; Canzoniero, Lorella M T; Formisano, Luigi

2017-10-15

Our previous study showed that the environmental neurotoxicant non-dioxin-like polychlorinated biphenyl (PCB)-95 increases RE1-silencing transcription factor (REST) expression, which is related to necrosis, but not apoptosis, of neurons. Meanwhile, necroptosis is a type of a programmed necrosis that is positively regulated by receptor interacting protein kinase 1 (RIPK1), RIPK3 and mixed lineage kinase domain-like (MLKL) and negatively regulated by caspase-8. Here we evaluated whether necroptosis contributes to PCB-95-induced neuronal death through REST up-regulation. Our results demonstrated that in cortical neurons PCB-95 increased RIPK1, RIPK3, and MLKL expression and decreased caspase-8 at the gene and protein level. Furthermore, the RIPK1 inhibitor necrostatin-1 or siRNA-mediated RIPK1, RIPK3 and MLKL expression knockdown significantly reduced PCB-95-induced neuronal death. Intriguingly, PCB-95-induced increases in RIPK1, RIPK3, MLKL expression and decreases in caspase-8 expression were reversed by knockdown of REST expression with a REST-specific siRNA (siREST). Notably, in silico analysis of the rat genome identified a REST consensus sequence in the caspase-8 gene promoter (Casp8-RE1), but not the RIPK1, RIPK3 and MLKL promoters. Interestingly, in PCB-95-treated neurons, REST binding to the Casp8-RE1 sequence increased in parallel with a reduction in its promoter activity, whereas under the same experimental conditions, transfection of siREST or mutation of the Casp8-RE1 sequence blocked PCB-95-induced caspase-8 reduction. Since RIPK1, RIPK3 and MLKL rat genes showed no putative REST binding site, we assessed whether the transcription factor cAMP Responsive Element Binding Protein (CREB), which has a consensus sequence in all three genes, affected neuronal death. In neurons treated with PCB-95, CREB protein expression decreased in parallel with a reduction in binding to the RIPK1, RIPK3 and MLKL gene promoter sequence. Furthermore, CREB overexpression was associated with reduced promoter activity of the RIPK1, RIPK3 and MLKL genes. Collectively, these results indicate that PCB-95 was associated with REST-induced necroptotic cell death by increasing RIPK1, RIPK3 and MLKL expression and reducing caspase-8 levels. In addition, since REST is involved in several neurological disorders, therapies that block REST-induced necroptosis could be a new strategy to revert the neurodetrimental effects associated to its overexpression. Copyright © 2017 Elsevier Inc. All rights reserved.

Phenolic acid intake, delivered via moderate champagne wine consumption, improves spatial working memory via the modulation of hippocampal and cortical protein expression/activation.

PubMed

Corona, Giulia; Vauzour, David; Hercelin, Justine; Williams, Claire M; Spencer, Jeremy P E

2013-11-10

While much data exist for the effects of flavonoid-rich foods on spatial memory in rodents, there are no such data for foods/beverages predominantly containing hydroxycinnamates and phenolic acids. To address this, we investigated the effects of moderate Champagne wine intake, which is rich in these components, on spatial memory and related mechanisms relative to the alcohol- and energy-matched controls. In contrast to the isocaloric and alcohol-matched controls, supplementation with Champagne wine (1.78 ml/kg BW, alcohol 12.5% vol.) for 6 weeks led to an improvement in spatial working memory in aged rodents. Targeted protein arrays indicated that these behavioral effects were paralleled by the differential expression of a number of hippocampal and cortical proteins (relative to the isocaloric control group), including those involved in signal transduction, neuroplasticity, apoptosis, and cell cycle regulation. Western immunoblotting confirmed the differential modulation of brain-derived neurotrophic factor, cAMP response-element-binding protein (CREB), p38, dystrophin, 2',3'-cyclic-nucleotide 3'-phosphodiesterase, mammalian target of rapamycin (mTOR), and Bcl-xL in response to Champagne supplementation compared to the control drink, and the modulation of mTOR, Bcl-xL, and CREB in response to alcohol supplementation. Our data suggest that smaller phenolics such as gallic acid, protocatechuic acid, tyrosol, caftaric acid, and caffeic acid, in addition to flavonoids, are capable of exerting improvements in spatial memory via the modulation in hippocampal signaling and protein expression. Changes in spatial working memory induced by the Champagne supplementation are linked to the effects of absorbed phenolics on cytoskeletal proteins, neurotrophin expression, and the effects of alcohol on the regulation of apoptotic events in the hippocampus and cortex.

The viral protein A238L inhibits TNF-alpha expression through a CBP/p300 transcriptional coactivators pathway.

PubMed

Granja, Aitor G; Nogal, Maria L; Hurtado, Carolina; Del Aguila, Carmen; Carrascosa, Angel L; Salas, María L; Fresno, Manuel; Revilla, Yolanda

2006-01-01

African swine fever virus (ASFV) is able to inhibit TNF-alpha-induced gene expression through the synthesis of A238L protein. This was shown by the use of deletion mutants lacking the A238L gene from the Vero cell-adapted Ba71V ASFV strain and from the virulent isolate E70. To further analyze the molecular mechanism by which the viral gene controls TNF-alpha, we have used Jurkat cells stably transfected with the viral gene to identify the TNF-alpha regulatory elements involved in the induction of the gene after stimulation with PMA and calcium ionophore. We have thus identified the cAMP-responsive element and kappa3 sites on the TNF-alpha promoter as the responsible of the gene activation, and demonstrate that A238L inhibits TNF-alpha expression through these DNA binding sites. This inhibition was partially reverted by overexpression of the transcriptional factors NF-AT, NF-kappaB, and c-Jun. Furthermore, we present evidence that A238L inhibits the activation of TNF-alpha by modulating NF-kappaB, NF-AT, and c-Jun trans activation through a mechanism that involves CREB binding protein/p300 function, because overexpression of these transcriptional coactivators recovers TNF-alpha promoter activity. In addition, we show that A238L is a nuclear protein that binds to the cyclic AMP-responsive element/kappa3 complex, thus displacing the CREB binding protein/p300 coactivators. Taken together, these results establish a novel mechanism in the control of TNF-alpha gene expression by a viral protein that could represent an efficient strategy used by ASFV to evade the innate immune response.

cAMP Response Element-binding Protein (CREB) and Nuclear Factor κB Mediate the Tamoxifen-induced Up-regulation of Glutamate Transporter 1 (GLT-1) in Rat Astrocytes*

PubMed Central

Karki, Pratap; Webb, Anton; Smith, Keisha; Lee, Kyuwon; Son, Deok-Soo; Aschner, Michael; Lee, Eunsook

2013-01-01

Tamoxifen (TX), a selective estrogen receptor modulator, exerts antagonistic effects on breast tissue and is used to treat breast cancer. Recent evidence also suggests that it may act as an agonist in brain tissue. We reported previously that TX enhanced the expression and function of glutamate transporter 1 (GLT-1) in rat astrocytes, an effect that was mediated by TGF-α. To gain further insight into the mechanisms that mediate TX-induced up-regulation of GLT-1 (EAAT2 in humans), we investigated its effect on GLT-1 at the transcriptional level. TX phosphorylated the cAMP response element-binding protein (CREB) and recruited CREB to the GLT-1 promoter consensus site. The effect of TX on astrocytic GLT-1 was attenuated by the inhibition of PKA, the upstream activator of the CREB pathway. In addition, the effect of TX on GLT-1 promoter activity was abolished by the inhibition of the NF-κB pathway. Furthermore, TX recruited the NF-κB subunits p65 and p50 to the NF-κB binding domain of the GLT-1 promoter. Mutation of NF-κB (triple, −583/-282/-251) or CRE (-308) sites on the GLT-1 promoter led to significant repression of the promoter activity, but neither mutant completely abolished the TX-induced GLT-1 promoter activity. Mutation of both the NF-κB (-583/-282/-251) and CRE (-308) sites led to a complete abrogation of the effect of TX on GLT-1 promoter activity. Taken together, our findings establish that TX regulates GLT-1 via the CREB and NF-κB pathways. PMID:23955341

Heterotrimeric G Stimulatory Protein α Subunit Is Required for Intestinal Smooth Muscle Contraction in Mice.

PubMed

Qin, Xiaoteng; Liu, Shangming; Lu, Qiulun; Zhang, Meng; Jiang, Xiuxin; Hu, Sanyuan; Li, Jingxin; Zhang, Cheng; Gao, Jiangang; Zhu, Min-Sheng; Feil, Robert; Li, Huashun; Chen, Min; Weinstein, Lee S; Zhang, Yun; Zhang, Wencheng

2017-04-01

The α subunit of the heterotrimeric G stimulatory protein (Gsa), encoded by the guanine nucleotide binding protein, α-stimulating gene (Gnas, in mice), is expressed ubiquitously and mediates receptor-stimulated production of cyclic adenosine monophosphate and activation of the protein kinase A signaling pathway. We investigated the roles of Gsa in vivo in smooth muscle cells of mice. We performed studies of mice with Cre recombinase-mediated disruption of Gnas in smooth muscle cells (Gsa SMKO and SM22-CreER T2 , induced in adult mice by tamoxifen). Intestinal tissues were collected for histologic, biochemical, molecular, cell biology, and physiology analyses. Intestinal function was assessed in mice using the whole-gut transit time test. We compared gene expression patterns of intestinal smooth muscle from mice with vs without disruption of Gnas. Biopsy specimens from ileum of patients with chronic intestinal pseudo-obstruction and age-matched control biopsies were analyzed by immunohistochemistry. Disruption of Gnas in smooth muscle of mice reduced intestinal motility and led to death within 4 weeks. Tamoxifen-induced disruption of Gnas in adult mice impaired contraction of intestinal smooth muscle and peristalsis. More than 80% of these died within 3 months of tamoxifen exposure, with features of intestinal pseudo-obstruction characterized by chronic intestinal dilation and dysmotility. Gsa deficiency reduced intestinal levels of cyclic adenosine monophosphate and transcriptional activity of the cyclic adenosine monophosphate response element binding protein 1 (CREB1); this resulted in decreased expression of the forkhead box F1 gene (Foxf1) and protein, and contractile proteins, such as myosin heavy chain 11; actin, α2, smooth muscle, aorta; calponin 1; and myosin light chain kinase. We found decreased levels of Gsa, FOXF1, CREB1, and phosphorylated CREB1 proteins in intestinal muscle layers of patients with chronic intestinal pseudo-obstruction, compared with tissues from controls. Gsa is required for intestinal smooth muscle contraction in mice, and its levels are reduced in ileum biopsies of patients with chronic intestinal pseudo-obstruction. Mice with disruption of Gnas might be used to study human chronic intestinal pseudo-obstruction. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Exercise Prevents Memory Impairment Induced by Arsenic Exposure in Mice: Implication of Hippocampal BDNF and CREB

PubMed Central

Yu, Zi-Jiang; Yu, Yan; Xiao, Chao-Lun; Kang, Chao-Sheng; Ge, Guo; Linghu, Yan; Zhu, Jun-De; Li, Yu-Mei; Li, Qiang-Ming; Luo, Shi-Peng; Yang, Dang; Li, Lin; Zhang, Wen-Yan; Tian, Guang

2015-01-01

High concentrations of arsenic, which can be occasionally found in drinking water, have been recognized as a global health problem. Exposure to arsenic can disrupt spatial memory; however, the underlying mechanism remains unclear. In the present study, we tested whether exercise could interfere with the effect of arsenic exposure on the long-term memory (LTM) of object recognition in mice. Arsenic (0, 1, 3, and 10 mg/ kg, i.g.) was administered daily for 12 weeks. We found that arsenic at dosages of 1, 3, and 10 mg/kg decreased body weight and increased the arsenic content in the brain. The object recognition LTM (tested 24 h after training) was disrupted by 3 mg/ kg and 10 mg/ kg, but not 1 mg/ kg arsenic exposure. Swimming exercise also prevented LTM impairment induced by 3 mg/ kg, but not with 10 mg/ kg, of arsenic exposure. The expression of brain-derived neurotrophic factor (BDNF) and phosphorylated cAMP-response element binding protein (pCREB) in the CA1 and dentate gyrus areas (DG) of the dorsal hippocampus were decreased by 3 mg/ kg and 10 mg/ kg, but not by 1 mg/ kg, of arsenic exposure. The decrease in BDNF and pCREB in the CA1 and DG induced by 3 mg/ kg, but not 10 mg/ kg, of arsenic exposure were prevented by swimming exercise. Arsenic exposure did not affect the total CREB expression in the CA1 or DG. Taken together, these results indicated that swimming exercise prevented the impairment of object recognition LTM induced by arsenic exposure, which may be mediated by BDNF and CREB in the dorsal hippocampus. PMID:26368803

Comparative transcriptomics reveals CrebA as a novel regulator of infection tolerance in D. melanogaster

PubMed Central

2018-01-01

Host responses to infection encompass many processes in addition to activation of the immune system, including metabolic adaptations, stress responses, tissue repair, and other reactions. The response to bacterial infection in Drosophila melanogaster has been classically described in studies that focused on the immune response elicited by a small set of largely avirulent microbes. Thus, we have surprisingly limited knowledge of responses to infection that are outside the canonical immune response, of how the response to pathogenic infection differs from that to avirulent bacteria, or even of how generic the response to various microbes is and what regulates that core response. In this study, we addressed these questions by profiling the D. melanogaster transcriptomic response to 10 bacteria that span the spectrum of virulence. We found that each bacterium triggers a unique transcriptional response, with distinct genes making up to one third of the response elicited by highly virulent bacteria. We also identified a core set of 252 genes that are differentially expressed in response to the majority of bacteria tested. Among these, we determined that the transcription factor CrebA is a novel regulator of infection tolerance. Knock-down of CrebA significantly increased mortality from microbial infection without any concomitant change in bacterial number. Upon infection, CrebA is upregulated by both the Toll and Imd pathways in the fat body, where it is required to induce the expression of secretory pathway genes. Loss of CrebA during infection triggered endoplasmic reticulum (ER) stress and activated the unfolded protein response (UPR), which contributed to infection-induced mortality. Altogether, our study reveals essential features of the response to bacterial infection and elucidates the function of a novel regulator of infection tolerance. PMID:29394281

γCaMKII shuttles Ca2+/CaM to the nucleus to trigger CREB phosphorylation and gene expression

PubMed Central

Ma, Huan; Groth, Rachel D.; Cohen, Samuel M.; Emery, John F.; Li, Bo-Xing; Hoedt, Esthelle; Zhang, Guo-An; Neubert, Thomas A.; Tsien, Richard W.

2014-01-01

SUMMARY Activity-dependent CREB phosphorylation and gene expression are critical for long-term neuronal plasticity. Local signaling at CaV1 channels triggers these events but how information is relayed onward to the nucleus remains unclear. Here we report a novel mechanism that mediates long-distance communication within cells: a shuttle that transports Ca2+/calmodulin from the surface membrane to the nucleus. We show that the shuttle protein is γCaMKII, that its phosphorylation at Thr287 by βCaMKII protects the Ca2+/CaM signal, and that CaN triggers its nuclear translocation. Both βCaMKII and CaN act in close proximity to CaV1 channels, supporting their dominance, while γCaMKII operates as a carrier, not as a kinase. Upon arrival within the nucleus, Ca2+/CaM activates CaMKK and its substrate CaMKIV, the CREB kinase. This mechanism resolves longstanding puzzles about CaM/CaMK-dependent signaling to the nucleus. The significance of the mechanism is emphasized by dysregulation of CaV1, γCaMKII, βCaMKII and CaN in multiple neuropsychiatric disorders. PMID:25303525

Myocyte enhancer factor (MEF)-2 plays essential roles in T-cell transformation associated with HTLV-1 infection by stabilizing complex between Tax and CREB.

PubMed

Jain, Pooja; Lavorgna, Alfonso; Sehgal, Mohit; Gao, Linlin; Ginwala, Rashida; Sagar, Divya; Harhaj, Edward W; Khan, Zafar K

2015-02-27

The exact molecular mechanisms regarding HTLV-1 Tax-mediated viral gene expression and CD4 T-cell transformation have yet to be fully delineated. Herein, utilizing virus-infected primary CD4+ T cells and the virus-producing cell line, MT-2, we describe the involvement and regulation of Myocyte enhancer factor-2 (specifically MEF-2A) during the course of HTLV-1 infection and associated disease syndrome. Inhibition of MEF-2 expression by shRNA and its activity by HDAC9 led to reduced viral replication and T-cell transformation in correlation with a heightened expression of MEF-2 in ATL patients. Mechanistically, MEF-2 was recruited to the viral promoter (LTR, long terminal repeat) in the context of chromatin, and constituted Tax/CREB transcriptional complex via direct binding to the HTLV-1 LTR. Furthermore, an increase in MEF-2 expression was observed upon infection in an extent similar to CREB (known Tax-interacting transcription factor), and HATs (p300, CBP, and p/CAF). Confocal imaging confirmed MEF-2 co-localization with Tax and these proteins were also shown to interact by co-immunoprecipitation. MEF-2 stabilization of Tax/CREB complex was confirmed by a novel promoter-binding assay that highlighted the involvement of NFAT (nuclear factor of activated T cells) in this process via Tax-mediated activation of calcineurin (a calcium-dependent serine-threonine phosphatase). MEF-2-integrated signaling pathways (PI3K/Akt, NF-κB, MAPK, JAK/STAT, and TGF-β) were also activated during HTLV-1 infection of primary CD4+ T cells, possibly regulating MEF-2 activity. We demonstrate the involvement of MEF-2 in Tax-mediated LTR activation, viral replication, and T-cell transformation in correlation with its heightened expression in ATL patients through direct binding to DNA within the HTLV-1 LTR.

Determining the Molecular and Genetic Basis for Diabetes in Navy Bottlenose Dolphins (Tursiops truncatus)

DTIC Science & Technology

2015-01-12

thereby reduces hepatic glucose production. 15. SUBJECT TERMS Gluconeogenesis , CREB ZF, Fasting, Diabetes 16. SECURITY CLASSIFICATION OF: a...Dolphin PEPCK transcription increased in the face of increasing cAMP, supporting that this enzyme induces gluconeogenesis during the fasting state...D. Test effects of CREB-ZF over-expression on gluconeogenic gene expression • Dolphin CREB-ZF is a novel, negative regulator of gluconeogenesis

A Genomic Response to Trace Fear Conditioning in the Amygdala of Female Rats After Developmental Exposure to Manganese

EPA Science Inventory

Increases in brain-derived neurotrophic factor (Bdnf), Ca2+/calmodulin-dependent protein kinase II alpha (Camk2a), and cyclic adenosine monophosphate (cAMP) response element binding (Creb1) gene expression have been associated with learning in a variety of different rodent studie...

Gotu Kola (Centella Asiatica) extract enhances phosphorylation of cyclic AMP response element binding protein in neuroblastoma cells expressing amyloid beta peptide.

PubMed

Xu, Yanan; Cao, Zhiming; Khan, Ikhlas; Luo, Yuan

2008-04-01

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that shows cognitive deficits and memory impairment. Extract from the leaves of Gotu Kola (Centella Asiatica) have been used as an alternative medicine for memory improvement in Indian Ayurvedic system of medicine for a long time. Although several studies have revealed its effect in ameliorating the cognitive impairment in rat models of AD and stimulating property on neuronal dendrites of hippocampal region, the molecular mechanism of Gotu Kola on neuroprotection still remains to be elucidated. In this study, we report that phosphorylation of cyclic AMP response element binding protein (CREB) is enhanced in both a neuroblastoma cell line expressing amyloid beta 1-42 (Abeta) and in rat embryonic cortical primary cell culture. In addition, the contribution of two major single components to the enhanced CREB phosphorylatioin was examined. Furthermore, inhibitors were applied in this study revealing that ERK/RSK signaling pathway might mediate this effect of Gotu Kola extract. Taken together, we provide a possible molecular mechanism for memory enhancing property of Gotu Kola extract for the first time.

Genomic Organization and Identification of Promoter Regions for the BDNF Gene in the Pond Turtle Trachemys scripta elegans

PubMed Central

Zheng, Zhaoqing; Keifer, Joyce

2014-01-01

Brain-derived neurotrophic factor (BDNF) is an important regulator of neuronal development and synaptic function. The BDNF gene undergoes significant activity-dependent regulation during learning. Here, we identified the BDNF promoter regions, transcription start sites, and potential regulatory sequences for BDNF exons I–III that may contribute to activity-dependent gene and protein expression in the pond turtle Trachemys scripta elegans (tBDNF). By using transfection of BDNF promoter/luciferase plasmid constructs into human neuroblastoma SHSY5Y cells and mouse embryonic fibroblast NIH3T3 cells, we identified the basal regulatory activity of promoter sequences located upstream of each tBDNF exon, designated as pBDNFI–III. Further, through chromatin immunoprecipitation (ChIP) assays, we detected CREB binding directly to exon I and exon III promoters, while BHLHB2, but not CREB, binds within the exon II promoter. Elucidation of the promoter regions and regulatory protein binding sites in the tBDNF gene is essential for understanding the regulatory mechanisms that control tBDNF gene expression. PMID:24443176

Genomic organization and identification of promoter regions for the BDNF gene in the pond turtle Trachemys scripta elegans.

PubMed

Ambigapathy, Ganesh; Zheng, Zhaoqing; Keifer, Joyce

2014-08-01

Brain-derived neurotrophic factor (BDNF) is an important regulator of neuronal development and synaptic function. The BDNF gene undergoes significant activity-dependent regulation during learning. Here, we identified the BDNF promoter regions, transcription start sites, and potential regulatory sequences for BDNF exons I-III that may contribute to activity-dependent gene and protein expression in the pond turtle Trachemys scripta elegans (tBDNF). By using transfection of BDNF promoter/luciferase plasmid constructs into human neuroblastoma SHSY5Y cells and mouse embryonic fibroblast NIH3T3 cells, we identified the basal regulatory activity of promoter sequences located upstream of each tBDNF exon, designated as pBDNFI-III. Further, through chromatin immunoprecipitation (ChIP) assays, we detected CREB binding directly to exon I and exon III promoters, while BHLHB2, but not CREB, binds within the exon II promoter. Elucidation of the promoter regions and regulatory protein binding sites in the tBDNF gene is essential for understanding the regulatory mechanisms that control tBDNF gene expression.

Mixture of Peanut Skin Extract and Fish Oil Improves Memory in Mice via Modulation of Anti-Oxidative Stress and Regulation of BDNF/ERK/CREB Signaling Pathways

PubMed Central

Xiang, Lan; Cao, Xue-Li; Xing, Tian-Yan; Mori, Daisuke; Tang, Rui-Qi; Li, Jing; Gao, Li-Juan; Qi, Jian-Hua

2016-01-01

Long-term use of fish oil (FO) is known to induce oxidative stress and increase the risk of Alzheimer’s disease in humans. In the present study, peanut skin extract (PSE), which has strong antioxidant capacity, was mixed with FO to reduce its side effects while maintaining its beneficial properties. Twelve-week Institute of Cancer Research (ICR) mice were used to conduct animal behavior tests in order to evaluate the memory-enhancing ability of the mixture of peanut skin extract and fish oil (MPF). MPF significantly increased alternations in the Y-maze and cognitive index in the novel object recognition test. MPF also improved performance in the water maze test. We further sought to understand the mechanisms underlying these effects. A significant decrease in superoxide dismutase (SOD) activity and an increase in malonyldialdehyde (MDA) in plasma were observed in the FO group. The MPF group showed reduced MDA level and increased SOD activity in the plasma, cortex and hippocampus. Furthermore, the gene expression levels of brain-derived neurotrophic factor (BDNF) and cAMP responsive element-binding protein (CREB) in the hippocampus were increased in the MPF group, while phosphorylation of protein kinase B (AKT), extracellular signal-regulated kinase (ERK) and CREB in the hippocampus were enhanced. MPF improves memory in mice via modulation of anti-oxidative stress and activation of BDNF/ERK/CREB signaling pathways. PMID:27136583

Alteration in brain-derived neurotrophic factor (BDNF) after treatment of mice with herbal mixture containing Euphoria longana, Houttuynia cordata and Dioscorea japonica.

PubMed

Jeon, Songhee; Lee, Chia-Hung; Liu, Quan Feng; Kim, Geun Woo; Koo, Byung-Soo; Pak, Sok Cheon

2014-11-28

Literature data indicate that brain-derived neurotrophic factor (BDNF), cyclic-AMP response element-binding protein (CREB) and phospho-CREB (pCREB) may have a place in depression. BDNF belongs to the neurotrophin family that plays an important role in proliferation, survival and differentiation of different cell populations in the mammalian nervous system. The herbal mixture used in the present study consists of Euphoria longana, Houttuynia cordata and Dioscorea japonica. The purpose of the present study was to determine the neuroprotective effect of herbal mixture. We also tested the hypothesis that administration of herbs reverses memory deficits and promotes the protein expression of BDNF in the mouse brain. Mice were randomized into four different treatment groups (n = 10/group). Normal and stress groups received regular lab chow without stress and under stress conditions, respectively, for 3 weeks. The animals in the stress group were immobilized for 4 hours a day for 2 weeks. Different doses of herbal mixture (206 and 618 mg/kg) were administered for 3 weeks to those mice under stress conditions. Mice were analyzed by behavioral tests and immunoblotting examination in the hippocampus and cortex. An additional in vitro investigation was performed to examine whether herbs induce neurotoxicity in a human neuroblastoma cell line, SH-SY5Y cells. No significant toxicity of herbs on human neuroblastoma cells was observed. These herbs demonstrated an inductive effect on the expression of BDNF, pCREB and pAkt. For spatial working memory test, herbal mixture fed mice exhibited an increased level of spontaneous alternation (p < 0.01) compared to those in stress conditions. Moreover, herbal mixture produced highly significant (p < 0.01) reduction in the immobility time in the tail suspension test. Mice in the herbal mixture groups demonstrated lower serum corticosterone concentration than mice in the stress group (p < 0.05). Effects of the oral administration of herbal mixture on protein levels of BDNF in the hippocampi and cortices were significant. Our study showed that herbal mixture administration has antidepressant effects in mice. It is proposed that adverse events such as stress and depression can modulate the expression of molecular players of cellular plasticity in the brain.

Transcriptional activation by MEIS1A in response to protein kinase A signaling requires the transducers of regulated CREB family of CREB co-activators.

PubMed

Goh, Siew-Lee; Looi, Yvonne; Shen, Hui; Fang, Jun; Bodner, Caroline; Houle, Martin; Ng, Andy Cheuk-Him; Screaton, Robert A; Featherstone, Mark

2009-07-10

The transcription factor encoded by the murine ecotropic integration site 1 gene (MEIS1) is a partner of HOX and PBX proteins. It has been implicated in embryonic patterning and leukemia, and causally linked to restless legs syndrome. The MEIS1A C terminus harbors a transcriptional activation domain that is stimulated by protein kinase A (PKA) in a manner dependent on the co-activator of cAMP response element-binding protein (CREB), CREB-binding protein (CBP). We explored the involvement of another mediator of PKA-inducible transcription, namely the CREB co-activators transducers of regulated CREB activity (TORCs). Overexpression of TORC1 or TORC2 bypassed PKA for activation by MEIS1A. Co-immunoprecipitation experiments demonstrated a physical interaction between MEIS1 and TORC2 that is dependent on the MEIS1A C terminus, whereas chromatin immunoprecipitation revealed PKA-inducible recruitment of MEIS1, PBX1, and TORC2 on the MEIS1 target genes Hoxb2 and Meis1. The MEIS1 interaction domain on TORC1 was mapped to the N-terminal coiled-coil region, and TORC1 mutants lacking this domain attenuated the response to PKA on a natural MEIS1A target enhancer. Thus, TORCs physically cooperate with MEIS1 to achieve PKA-inducible transactivation through the MEIS1A C terminus, suggesting a concerted action in developmental and oncogenic processes.

Transcriptional Activation by MEIS1A in Response to Protein Kinase A Signaling Requires the Transducers of Regulated CREB Family of CREB Co-activators*

PubMed Central

Goh, Siew-Lee; Looi, Yvonne; Shen, Hui; Fang, Jun; Bodner, Caroline; Houle, Martin; Ng, Andy Cheuk-Him; Screaton, Robert A.; Featherstone, Mark

2009-01-01

The transcription factor encoded by the murine ecotropic integration site 1 gene (MEIS1) is a partner of HOX and PBX proteins. It has been implicated in embryonic patterning and leukemia, and causally linked to restless legs syndrome. The MEIS1A C terminus harbors a transcriptional activation domain that is stimulated by protein kinase A (PKA) in a manner dependent on the co-activator of cAMP response element-binding protein (CREB), CREB-binding protein (CBP). We explored the involvement of another mediator of PKA-inducible transcription, namely the CREB co-activators transducers of regulated CREB activity (TORCs). Overexpression of TORC1 or TORC2 bypassed PKA for activation by MEIS1A. Co-immunoprecipitation experiments demonstrated a physical interaction between MEIS1 and TORC2 that is dependent on the MEIS1A C terminus, whereas chromatin immunoprecipitation revealed PKA-inducible recruitment of MEIS1, PBX1, and TORC2 on the MEIS1 target genes Hoxb2 and Meis1. The MEIS1 interaction domain on TORC1 was mapped to the N-terminal coiled-coil region, and TORC1 mutants lacking this domain attenuated the response to PKA on a natural MEIS1A target enhancer. Thus, TORCs physically cooperate with MEIS1 to achieve PKA-inducible transactivation through the MEIS1A C terminus, suggesting a concerted action in developmental and oncogenic processes. PMID:19473990

New learning while consolidating memory during sleep is actively blocked by a protein synthesis dependent process

PubMed Central

Levy, Roi; Levitan, David; Susswein, Abraham J

2016-01-01

Brief experiences while a memory is consolidated may capture the consolidation, perhaps producing a maladaptive memory, or may interrupt the consolidation. Since consolidation occurs during sleep, even fleeting experiences when animals are awakened may produce maladaptive long-term memory, or may interrupt consolidation. In a learning paradigm affecting Aplysia feeding, when animals were trained after being awakened from sleep, interactions between new experiences and consolidation were prevented by blocking long-term memory arising from the new experiences. Inhibiting protein synthesis eliminated the block and allowed even a brief, generally ineffective training to produce long-term memory. Memory formation depended on consolidative proteins already expressed before training. After effective training, long term memory required subsequent transcription and translation. Memory formation during the sleep phase was correlated with increased CREB1 transcription, but not CREB2 transcription. Increased C/EBP transcription was a correlate of both effective and ineffective training and of treatments not producing memory. DOI: http://dx.doi.org/10.7554/eLife.17769.001 PMID:27919318

New learning while consolidating memory during sleep is actively blocked by a protein synthesis dependent process.

PubMed

Levy, Roi; Levitan, David; Susswein, Abraham J

2016-12-06

Brief experiences while a memory is consolidated may capture the consolidation, perhaps producing a maladaptive memory, or may interrupt the consolidation. Since consolidation occurs during sleep, even fleeting experiences when animals are awakened may produce maladaptive long-term memory, or may interrupt consolidation. In a learning paradigm affecting Aplysia feeding, when animals were trained after being awakened from sleep, interactions between new experiences and consolidation were prevented by blocking long-term memory arising from the new experiences. Inhibiting protein synthesis eliminated the block and allowed even a brief, generally ineffective training to produce long-term memory. Memory formation depended on consolidative proteins already expressed before training. After effective training, long term memory required subsequent transcription and translation. Memory formation during the sleep phase was correlated with increased CREB1 transcription, but not CREB2 transcription. Increased C/EBP transcription was a correlate of both effective and ineffective training and of treatments not producing memory.

Comparative Studies on Behavioral, Cognitive and Biomolecular Profiling of ICR, C57BL/6 and Its Sub-Strains Suitable for Scopolamine-Induced Amnesic Models

PubMed Central

Karthivashan, Govindarajan; Park, Shin-Young; Kim, Joon-Soo; Cho, Duk-Yeon

2017-01-01

Cognitive impairment and behavioral disparities are the distinctive baseline features to investigate in most animal models of neurodegenerative disease. However, neuronal complications are multifactorial and demand a suitable animal model to investigate their underlying basal mechanisms. By contrast, the numerous existing neurodegenerative studies have utilized various animal strains, leading to factual disparity. Choosing an optimal mouse strain for preliminary assessment of neuronal complications is therefore imperative. In this study, we systematically compared the behavioral, cognitive, cholinergic, and inflammatory impairments of outbred ICR and inbred C57BL/6 mice strains subject to scopolamine-induced amnesia. We then extended this study to the sub-strains C57BL/6N and C57BL/6J, where in addition to the above-mentioned parameters, their endogenous antioxidant levels and cAMP response-element binding protein (CREB)/brain-derived neurotrophic factor (BDNF) protein expression were also evaluated. Compared with the ICR strain, the scopolamine-inflicted C57BL/6 strains exhibited a substantial reduction of spontaneous alternation and an approximately two-fold increase in inflammatory protein expression, compared to the control group. Among the sub-strains, scopolamine-treated C57BL/6N strains exhibited declined step-through latency, elevated acetylcholinesterase (AChE) activity and inflammatory protein expression, associated with reduced endogenous antioxidant levels and p-CREB/BDNF expression, compared to the control and tacrine-treated groups. This indicates that the C57BL/6N strains exhibit significantly enhanced scopolamine-induced neuronal impairment compared to the other evaluated strains. PMID:28792471

Comparative Studies on Behavioral, Cognitive and Biomolecular Profiling of ICR, C57BL/6 and Its Sub-Strains Suitable for Scopolamine-Induced Amnesic Models.

PubMed

Karthivashan, Govindarajan; Park, Shin-Young; Kim, Joon-Soo; Cho, Duk-Yeon; Ganesan, Palanivel; Choi, Dong-Kug

2017-08-09

Cognitive impairment and behavioral disparities are the distinctive baseline features to investigate in most animal models of neurodegenerative disease. However, neuronal complications are multifactorial and demand a suitable animal model to investigate their underlying basal mechanisms. By contrast, the numerous existing neurodegenerative studies have utilized various animal strains, leading to factual disparity. Choosing an optimal mouse strain for preliminary assessment of neuronal complications is therefore imperative. In this study, we systematically compared the behavioral, cognitive, cholinergic, and inflammatory impairments of outbred ICR and inbred C57BL/6 mice strains subject to scopolamine-induced amnesia. We then extended this study to the sub-strains C57BL/6N and C57BL/6J, where in addition to the above-mentioned parameters, their endogenous antioxidant levels and cAMP response-element binding protein (CREB)/brain-derived neurotrophic factor (BDNF) protein expression were also evaluated. Compared with the ICR strain, the scopolamine-inflicted C57BL/6 strains exhibited a substantial reduction of spontaneous alternation and an approximately two-fold increase in inflammatory protein expression, compared to the control group. Among the sub-strains, scopolamine-treated C57BL/6N strains exhibited declined step-through latency, elevated acetylcholinesterase (AChE) activity and inflammatory protein expression, associated with reduced endogenous antioxidant levels and p-CREB/BDNF expression, compared to the control and tacrine-treated groups. This indicates that the C57BL/6N strains exhibit significantly enhanced scopolamine-induced neuronal impairment compared to the other evaluated strains.

Transcription factor CREB is involved in CaSR-mediated cytoskeleton gene expression.

PubMed

Huang, Shuaishuai; Ren, Yu; Wang, Ping; Li, Yanyuan; Wang, Xue; Zhuang, Haihui; Fang, Rong; Wang, Yuduo; Liu, Ningsheng; Hehir, Michael; Zhou, Jeff X

2015-03-01

Our previous studies illustrated that a steady increase of intracellular calcium concentration ([Ca2+]i) was important for maintaining microtubules (MTs) rearrangement in apoptotic cells. However, little is known about the effect of calcium sensing receptor (CaSR)-mediated increase in [Ca2+]i on cytoskeleton gene expression. We examined the impact of taxol or CaSR agonist/antagonist on the regulation of [Ca2+]i concentration, cytoskeleton arrangement, phosphorylated CREB and cytoskeleton gene expressions in HeLa cells with dominant negative plasmid of CREB (PM). This study demonstrated that Gdcl3 (a specific CaSR agonist) evoked a rapid increase of [Ca2+]i, formed a rigid bundle of MTs which surrounded the nucleus and decreased the cytoskeleton gene expressions in HeLa cells. These effects were rescued by addition of NPS2390 (a specific CaSR antagonist). Moreover, CaSR activity affected cytoskeleton gene expression through transcription factor CREB. Histoscores of pCREB immunoreactivity in tissues of cervical adenocarcinoma, renal clear cell carcinoma, and diffuse large B-cell lymphoma were markedly increased compared with non malignant tissue. These data demonstrate, for the first time, that CaSR-mediated increase in [Ca2+]i probably modulate cytoskeleton organization and gene expression via transcription factor. © 2014 Wiley Periodicals, Inc.

Modulation of neuroplasticity pathways and antidepressant-like behavioural responses following the short-term (3 and 7 days) administration of the 5-HT₄ receptor agonist RS67333.

PubMed

Pascual-Brazo, Jesús; Castro, Elena; Díaz, Alvaro; Valdizán, Elsa M; Pilar-Cuéllar, Fuencisla; Vidal, Rebeca; Treceño, Begoña; Pazos, Angel

2012-06-01

It has been recently suggested that activation of 5-HT₄ receptors might exert antidepressant-like effects in rats after 3 d treatment, suggesting a new strategy for developing faster-acting antidepressants. We studied the effects of 3 d and 7 d treatment with the 5-HT₄ receptor partial agonist RS67333 (1.5 mg/kg.d) in behavioural tests of chronic efficacy and on neuroplastic-associated changes, such as adult hippocampal neurogenesis, expression of CREB, BDNF, β-catenin, AKT and 5-HT₄ receptor functionality. RS67333 treatment up-regulated hippocampal cell proliferation, β-catenin expression and pCREB/CREB ratio after 3 d treatment. This short-term treatment also reduced immobility time in the forced swim test (FST), together with a partial reversion of the anhedonic-like state (sucrose consumption after chronic corticosterone). Administration of RS67333 for 7 d resulted in a higher increase in the rate of hippocampal cell proliferation, a significant desensitization of 5-HT₄ receptor-coupled adenylate cyclase activity and a more marked increase in the expression of neuroplasticity-related proteins (BDNF, CREB, AKT): these changes reached the same magnitude as those observed after 3 wk administration of classical antidepressants. Consistently, a positive behavioural response in the novelty suppressed feeding (NSF) test and a complete reversion of the anhedonic-like state (sucrose consumption) were also observed after 7 d treatment. These results support the antidepressant-like profile of RS67333 with a shorter onset of action and suggest that this time period of administration (3-7 d) could be a good approximation to experimentally predict the onset of action of this promising strategy.

BDNF and exercise enhance neuronal DNA repair by stimulating CREB-mediated production of apurinic/apyrimidinic endonuclease 1.

PubMed

Yang, Jenq-Lin; Lin, Yu-Ting; Chuang, Pei-Chin; Bohr, Vilhelm A; Mattson, Mark P

2014-03-01

Brain-derived neurotrophic factor (BDNF) promotes the survival and growth of neurons during brain development and mediates activity-dependent synaptic plasticity and associated learning and memory in the adult. BDNF levels are reduced in brain regions affected in Alzheimer's, Parkinson's, and Huntington's diseases, and elevation of BDNF levels can ameliorate neuronal dysfunction and degeneration in experimental models of these diseases. Because neurons accumulate oxidative lesions in their DNA during normal activity and in neurodegenerative disorders, we determined whether and how BDNF affects the ability of neurons to cope with oxidative DNA damage. We found that BDNF protects cerebral cortical neurons against oxidative DNA damage-induced death by a mechanism involving enhanced DNA repair. BDNF stimulates DNA repair by activating cyclic AMP response element-binding protein (CREB), which, in turn, induces the expression of apurinic/apyrimidinic endonuclease 1 (APE1), a key enzyme in the base excision DNA repair pathway. Suppression of either APE1 or TrkB by RNA interference abolishes the ability of BDNF to protect neurons against oxidized DNA damage-induced death. The ability of BDNF to activate CREB and upregulate APE1 expression is abolished by shRNA of TrkB as well as inhibitors of TrkB, PI3 kinase, and Akt kinase. Voluntary running wheel exercise significantly increases levels of BDNF, activates CREB, and upregulates APE1 in the cerebral cortex and hippocampus of mice, suggesting a novel mechanism whereby exercise may protect neurons from oxidative DNA damage. Our findings reveal a previously unknown ability of BDNF to enhance DNA repair by inducing the expression of the DNA repair enzyme APE1.

Increased pCREB expression and the spontaneous epileptiform activity in a BCNU-treated rat model of cortical dysplasia.

PubMed

Pennacchio, Paolo; Noé, Francesco; Gnatkovsky, Vadym; Moroni, Ramona Frida; Zucca, Ileana; Regondi, Maria Cristina; Inverardi, Francesca; de Curtis, Marco; Frassoni, Carolina

2015-09-01

Cortical dysplasias (CDs) represent a wide range of cortical abnormalities that closely correlate with intractable epilepsy. Rats prenatally exposed to 1-3-bis-chloroethyl-nitrosurea (BCNU) represent an injury-based model that reproduces many histopathologic features of human CD. Previous studies reported in vivo hyperexcitability in this model, but in vivo epileptogenicity has not been confirmed. To determine whether cortical and hippocampal lesions lead to epileptiform discharges and/or seizures in the BCNU model, rats at three different ages (3, 5, and 9 months old) were implanted for long-term video electroencephalographic recording. At the end of the recording session, brain tissue was processed for histologic and immunohistochemical investigation including cAMP response element binding protein (CREB) phosphorylation, as a biomarker of epileptogenicity. BCNU-treated rats showed spontaneous epileptiform activity (67%) in the absence of a second seizure-provoking hit. Such activity originated mainly from one hippocampus and propagated to the ipsilateral neocortex. No epileptiform activity was found in age-matched control rats. The histopathologic investigation revealed that all BCNU rats with epileptiform activity showed neocortical and hippocampal abnormalities; the presence and the severity of these lesions did not correlate consistently with the propensity to generate epileptiform discharges. Epileptiform activity was found only in cortical areas of BCNU-treated rats in which a correlation between brain abnormalities and increased pCREB expression was observed. This study demonstrates the in vivo occurrence of spontaneous epileptiform discharges in the BCNU model and shows that increased pCREB expression can be utilized as a reliable biomarker of epileptogenicity. Wiley Periodicals, Inc. © 2015 International League Against Epilepsy.

Anodal transcranial direct current stimulation boosts synaptic plasticity and memory in mice via epigenetic regulation of Bdnf expression

PubMed Central

Podda, Maria Vittoria; Cocco, Sara; Mastrodonato, Alessia; Fusco, Salvatore; Leone, Lucia; Barbati, Saviana Antonella; Colussi, Claudia; Ripoli, Cristian; Grassi, Claudio

2016-01-01

The effects of transcranial direct current stimulation (tDCS) on brain functions and the underlying molecular mechanisms are yet largely unknown. Here we report that mice subjected to 20-min anodal tDCS exhibited one-week lasting increases in hippocampal LTP, learning and memory. These effects were associated with enhanced: i) acetylation of brain-derived neurotrophic factor (Bdnf) promoter I; ii) expression of Bdnf exons I and IX; iii) Bdnf protein levels. The hippocampi of stimulated mice also exhibited enhanced CREB phosphorylation, pCREB binding to Bdnf promoter I and recruitment of CBP on the same regulatory sequence. Inhibition of acetylation and blockade of TrkB receptors hindered tDCS effects at molecular, electrophysiological and behavioral levels. Collectively, our findings suggest that anodal tDCS increases hippocampal LTP and memory via chromatin remodeling of Bdnf regulatory sequences leading to increased expression of this gene, and support the therapeutic potential of tDCS for brain diseases associated with impaired neuroplasticity. PMID:26908001

Protective effect of total flavones of Abelmoschus manihot L. Medic against poststroke depression injury in mice and its action mechanism.

PubMed

Liu, Mei; Jiang, Qiu-Hong; Hao, Ji-Li; Zhou, Lan-Lan

2009-03-01

Total flavones of Abelmoschus manihot L. Medic (TFA) is the major active component isolated from the traditional Chinese herb Abelmoschus manihot L. Medic. We investigated the protective effect of TFA against poststroke depression (PSD) injury in mice and its action mechanism. A mouse model of PSD was induced by middle cerebral artery occlusion (MACO) 30 min/reperfusion, followed by isolation feeding and chronic unpredictable mild stress for 2 weeks. Treatment groups received TFA at three different doses (160, 80, and 40 mg/kg, p.o.) or fluoxetine (Flu, 2.5 mg/kg, p.o.) daily for 24 days. Change in behavior, brain tissue malondialdehyde (MDA) levels, and the activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were measured. The expression of brain-derived neurotrophic factor (BDNF) was detected by immunohistochemistry, and mRNA expression of BDNF and cAMP response element-binding protein (CREB) analyzed by reverse transcription-polymerase chain reaction (RT-PCR). Treatment with TFA (160, 80, and 40 mg/kg) significantly ameliorated mice escape-directed behavioral impairment induced by PSD, markedly reduced MDA levels, and increased the activity of SOD, GSH-Px close to normal levels. TFA administration also attenuated PSD-induced neuronal death/losses, upregulated expression of BDNF both at mRNA and protein levels, as well as CREB mRNA levels. TFA had a protective effect against PSD injury in mice. Cardioprotection involves the inhibition of lipid peroxidation and upregulation of BDNF-CREB levels in the hippocampus, which may also be important mechanism of its antidepressants. This potential protection makes TFA a promising therapeutic agent for the PSD. (c) 2009 Wiley-Liss, Inc.

Protection of rats spinal cord ischemia-reperfusion injury by inhibition of MiR-497 on inflammation and apoptosis: Possible role in pediatrics.

PubMed

Xu, Meng; Wang, Hai-Feng; Zhang, Ying-Ying; Zhuang, Hui-Wen

2016-07-01

MicroRNAs are extensively included in the pathogenesis and progression of many diseases by inhibiting target gene expression. Recently, studies have demonstrated that microRNA-497 (miR-497) may be implicated in human breast cancer that miR-497 predicts the prognosis of breast cancer patients from the posttranscriptional level. However, the specific function of miR-497 in spinal cord ischemia-reperfusion (IR) injury is far from clear nowadays. The present study was designed to determine the role of miR-497 in spinal cord IR injury and investigate the underlying spinal cord protective mechanism. The rat spinal cord IR injury model was performed by occluding the left anterior descending coronary artery for 30 min, which is then followed by 12h reperfusion. As predicted, miR-497 over-expression markedly decreased the expression of IL-1 receptor associated kinase (IRAK1) and Cyclic AMP response element binding protein (CREB). Moreover, Toll-like receptor 4 (TLR4), nuclear factor-kappa B (NF-κB) and Caspase-3, as miR-497 potential targets were significantly suppressed after miR-497 transfection, then preventing inflammatory cytokines and factors regulating apoptosis. We also found that tumor necrosis factor-a (TNF-α) and interleukin-1beta (IL-1β) activity, pro-apoptotic related genes, such as extracellular regulated protein kinases (ERK), Bcl-2 Associated X Protein (Bax), Bcl-2, Bcl-xL levels were all decreased associated with the down-regulation of IRAK1 and CREB. In conclusion, our data demonstrate that miR-497 could inhibit inflammation and apoptosis of spinal cord IR through its targets, IRAK1 of TLR4 and CREB signaling pathway. Thus, miR-497 may constitute a new therapeutic target for the prevention of spinal cord IR injury. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Rhubarb Tannins Extract Inhibits the Expression of Aquaporins 2 and 3 in Magnesium Sulphate-Induced Diarrhoea Model

PubMed Central

Liu, Chunfang; Zheng, Yanfang; Xu, Wen; Wang, Hui

2014-01-01

Tannins, a group of major active components of Chinese rhubarb and widely distributed in nature, have a significant antidiarrhoeal activity. Aquaporins (AQPs) 2 and 3 play important roles in regulating water transfer during diarrhoea. The present study aims to determine the effect of the total tannins extract of rhubarb on aquaporins (AQPs) 2 and 3 in diarrhoea mice and HT-29 cells both induced by magnesium sulphate (MgSO4). Our results showed that rhubarb tannins extract (RTE) significantly decreased the faecal water content in colon and evaluation index of defecation of diarrhoea mice. Interestingly, RTE could markedly reduce the mRNA and protein expression levels of AQPs 2 and 3 in apical and lateral mucosal epithelial cells in the colons of diarrhoea mice and HT-29 cells both induced by MgSO4 in a dose-dependent manner. Furthermore, RTE suppressed the production of cyclic monophosphate- (cAMP-) dependent protein kinase A catalytic subunits α (PKA C-α) and phosphorylated cAMP response element-binding protein (p-CREB, Ser133) in MgSO4-induced HT-29 cells. Our data showed for the first time that RTE inhibit AQPs 2 and 3 expression in vivo and in vitro via downregulating PKA/p-CREB signal pathway, which accounts for the antidiarrhoeal effect of RTE. PMID:25215286

Nobiletin inhibits human osteosarcoma cells metastasis by blocking ERK and JNK-mediated MMPs expression

PubMed Central

Cheng, Hsin-Lin; Hsieh, Ming-Ju; Yang, Jia-Sin; Lin, Chiao-Wen; Lue, Ko-Haung; Lu, Ko-Hsiu; Yang, Shun-Fa

2016-01-01

Nobiletin, a polymethoxyflavone, has a few pharmacological activities, including anti-inflammation and anti-cancer effects. However, its effect on human osteosarcoma progression remains uninvestigated. Therefore, we examined the effectiveness of nobiletin against cellular metastasis of human osteosarcoma and the underlying mechanisms. Nobiletin, up to 100 μM without cytotoxicity, significantly decreased motility, migration and invasion as well as enzymatic activities, protein levels and mRNA expressions of matrix metalloproteinase (MMP)-2 and MMP-9 in U2OS and HOS cells. In addition to inhibition of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), the inhibitory effect of nobiletin on the DNA-binding activity of the transcription factor nuclear factor-kappa B (NF-κB), cAMP response element-binding protein (CREB), and specificity protein 1 (SP-1) in U2OS and HOS cells. Co-treatment with ERK and JNK inhibitors and nobiletin further reduced U2OS cells migration and invasion. These results indicated that nobiletin inhibits human osteosarcoma U2OS and HOS cells motility, migration and invasion by down-regulating MMP-2 and MMP-9 expressions via ERK and JNK pathways and through the inactivation of downstream NF-κB, CREB, and SP-1. Nobiletin has the potential to serve as an anti-metastatic agent for treating osteosarcoma. PMID:27144433

Upregulation of CREB-mediated transcription enhances both short- and long-term memory.

PubMed

Suzuki, Akinobu; Fukushima, Hotaka; Mukawa, Takuya; Toyoda, Hiroki; Wu, Long-Jun; Zhao, Ming-Gao; Xu, Hui; Shang, Yuze; Endoh, Kengo; Iwamoto, Taku; Mamiya, Nori; Okano, Emiko; Hasegawa, Shunsuke; Mercaldo, Valentina; Zhang, Yue; Maeda, Ryouta; Ohta, Miho; Josselyn, Sheena A; Zhuo, Min; Kida, Satoshi

2011-06-15

Unraveling the mechanisms by which the molecular manipulation of genes of interest enhances cognitive function is important to establish genetic therapies for cognitive disorders. Although CREB is thought to positively regulate formation of long-term memory (LTM), gain-of-function effects of CREB remain poorly understood, especially at the behavioral level. To address this, we generated four lines of transgenic mice expressing dominant active CREB mutants (CREB-Y134F or CREB-DIEDML) in the forebrain that exhibited moderate upregulation of CREB activity. These transgenic lines improved not only LTM but also long-lasting long-term potentiation in the CA1 area in the hippocampus. However, we also observed enhanced short-term memory (STM) in contextual fear-conditioning and social recognition tasks. Enhanced LTM and STM could be dissociated behaviorally in these four lines of transgenic mice, suggesting that the underlying mechanism for enhanced STM and LTM are distinct. LTM enhancement seems to be attributable to the improvement of memory consolidation by the upregulation of CREB transcriptional activity, whereas higher basal levels of BDNF, a CREB target gene, predicted enhanced shorter-term memory. The importance of BDNF in STM was verified by microinfusing BDNF or BDNF inhibitors into the hippocampus of wild-type or transgenic mice. Additionally, increasing BDNF further enhanced LTM in one of the lines of transgenic mice that displayed a normal BDNF level but enhanced LTM, suggesting that upregulation of BDNF and CREB activity cooperatively enhances LTM formation. Our findings suggest that CREB positively regulates memory consolidation and affects memory performance by regulating BDNF expression.

Methamphetamine acutely inhibits voltage-gated calcium channels but chronically up-regulates L-type channels.

PubMed

Andres, Marilou A; Cooke, Ian M; Bellinger, Frederick P; Berry, Marla J; Zaporteza, Maribel M; Rueli, Rachel H; Barayuga, Stephanie M; Chang, Linda

2015-07-01

In neurons, calcium (Ca(2+) ) channels regulate a wide variety of functions ranging from synaptic transmission to gene expression. They also induce neuroplastic changes that alter gene expression following psychostimulant administration. Ca(2+) channel blockers have been considered as potential therapeutic agents for the treatment of methamphetamine (METH) dependence because of their ability to reduce drug craving among METH users. Here, we studied the effects of METH exposure on voltage-gated Ca(2+) channels using SH-SY5Y cells as a model of dopaminergic neurons. We found that METH has different short- and long-term effects. A short-term effect involves immediate (< 5 min) direct inhibition of Ca(2+) ion movements through Ca(2+) channels. Longer exposure to METH (20 min or 48 h) selectively up-regulates the expression of only the CACNA1C gene, thus increasing the number of L-type Ca(2+) channels. This up-regulation of CACNA1C is associated with the expression of the cAMP-responsive element-binding protein (CREB), a known regulator of CACNA1C gene expression, and the MYC gene, which encodes a transcription factor that putatively binds to a site proximal to the CACNA1C gene transcription initiation site. The short-term inhibition of Ca(2+) ion movement and later, the up-regulation of Ca(2+) channel gene expression together suggest the operation of cAMP-responsive element-binding protein- and C-MYC-mediated mechanisms to compensate for Ca(2+) channel inhibition by METH. Increased Ca(2+) current density and subsequent increased intracellular Ca(2+) may contribute to the neurodegeneration accompanying chronic METH abuse. Methamphetamine (METH) exposure has both short- and long-term effects. Acutely, methamphetamine directly inhibits voltage-gated calcium channels. Chronically, neurons compensate by up-regulating the L-type Ca(2+) channel gene, CACNA1C. This compensatory mechanism is mediated by transcription factors C-MYC and CREB, in which CREB is linked to the dopamine D1 receptor signaling pathway. These findings suggest Ca(2+) -mediated neurotoxicity owing to over-expression of calcium channels. © 2015 International Society for Neurochemistry.

The extraction of simple relationships in growth factor-specific multiple-input and multiple-output systems in cell-fate decisions by backward elimination PLS regression.

PubMed

Akimoto, Yuki; Yugi, Katsuyuki; Uda, Shinsuke; Kudo, Takamasa; Komori, Yasunori; Kubota, Hiroyuki; Kuroda, Shinya

2013-01-01

Cells use common signaling molecules for the selective control of downstream gene expression and cell-fate decisions. The relationship between signaling molecules and downstream gene expression and cellular phenotypes is a multiple-input and multiple-output (MIMO) system and is difficult to understand due to its complexity. For example, it has been reported that, in PC12 cells, different types of growth factors activate MAP kinases (MAPKs) including ERK, JNK, and p38, and CREB, for selective protein expression of immediate early genes (IEGs) such as c-FOS, c-JUN, EGR1, JUNB, and FOSB, leading to cell differentiation, proliferation and cell death; however, how multiple-inputs such as MAPKs and CREB regulate multiple-outputs such as expression of the IEGs and cellular phenotypes remains unclear. To address this issue, we employed a statistical method called partial least squares (PLS) regression, which involves a reduction of the dimensionality of the inputs and outputs into latent variables and a linear regression between these latent variables. We measured 1,200 data points for MAPKs and CREB as the inputs and 1,900 data points for IEGs and cellular phenotypes as the outputs, and we constructed the PLS model from these data. The PLS model highlighted the complexity of the MIMO system and growth factor-specific input-output relationships of cell-fate decisions in PC12 cells. Furthermore, to reduce the complexity, we applied a backward elimination method to the PLS regression, in which 60 input variables were reduced to 5 variables, including the phosphorylation of ERK at 10 min, CREB at 5 min and 60 min, AKT at 5 min and JNK at 30 min. The simple PLS model with only 5 input variables demonstrated a predictive ability comparable to that of the full PLS model. The 5 input variables effectively extracted the growth factor-specific simple relationships within the MIMO system in cell-fate decisions in PC12 cells.

CB2 Cannabinoid Receptor Targets Mitogenic Gi Protein–Cyclin D1 Axis in Osteoblasts

PubMed Central

Ofek, Orr; Attar-Namdar, Malka; Kram, Vardit; Dvir-Ginzberg, Mona; Mechoulam, Raphael; Zimmer, Andreas; Frenkel, Baruch; Shohami, Esther; Bab, Itai

2011-01-01

CB2 is a Gi protein–coupled receptor activated by endo- and phytocannabinoids, thus inhibiting stimulated adenylyl cyclase activity. CB2 is expressed in bone cells and Cb2 null mice show a marked age-related bone loss. CB2-specific agonists both attenuate and rescue ovariectomy-induced bone loss. Activation of CB2 stimulates osteoblast proliferation and bone marrow derived colony-forming units osteoblastic. Here we show that selective and nonselective CB2 agonists are mitogenic in MC3T3 E1 and newborn mouse calvarial osteoblastic cultures. The CB2 mitogenic signaling depends critically on the stimulation of Erk1/2 phosphorylation and de novo synthesis of MAP kinase–activated protein kinase 2 (Mapkapk2) mRNA and protein. Further downstream, CB2 activation enhances CREB transcriptional activity and cyclin D1 mRNA expression. The CB2-induced stimulation of CREB and cyclin D1 is inhibitable by pertussis toxin, the MEK-Erk1/2 inhibitors PD098059 and U0126, and Mapkapk2 siRNA. These data demonstrate that in osteoblasts CB2 targets a Gi protein–cyclin D1 mitogenic axis. Erk1/2 phosphorylation and Mapkapk2 protein synthesis are critical intermediates in this axis. © 2011 American Society for Bone and Mineral Research. PMID:20803555

PKC/CREB pathway mediates the expressions of GABAA receptor subunits in cultured hippocampal neurons after low-Mg2+ solution treatment.

PubMed

Wu, Guofeng; Yu, Jinpeng; Wang, Likun; Ren, Siying; Zhang, Yixia

2018-02-01

To investigate the potential effects of the PKC/CREB pathway on the expressions of GABA A receptor subunits α1, γ2, and δ in cultured hippocampal neurons using a model of epilepsy that employed conditions of low magnesium (Mg 2+ ). A total of 108 embryonic rats at the age of 18 embryonic days (E18)prepared from adult female SD rats were used as experimental subjects. Primary rat hippocampal cultures were prepared from the embryonic 18 days rats. The cultured hippocampal neurons were then treated with artificial cerebrospinal fluid containing low Mg 2+ solutions to generate a low Mg 2+ model of epilepsy. The low Mg 2+ stimulation lasted for 3 h and then returned to in maintenance medium for 20 h. The changes of the GABA A receptor subunit α1, γ2, δ were observed by blocking or activating the function of the CREB. The quantification of the GABA A receptor subunit α1, γ2, δ and the CREB were determined by a qRT-PCR and a Western blot method. After the neurons were exposed to a low-Mg 2+ solution for 3 h, GABA A receptor mRNA expression markedly increased compared to the control, and then gradually decreased. In contrast, CREB mRNA levels exhibited a dramatic down-regulation 3 h after terminating low-Mg 2+ treatment, and then peaked at 9 h. Western blot analyses verified that staurosporine suppressed CREB phosphorylation (p-CREB). The mRNA expression of GABA A receptor subunit α1 increased only in the presence of staurosporine, whereas the expressions of subunits γ2 and δ significantly increased in the presence of either KG-501 or staurosporine. Furthermore, phorbol 12-myristate 13-acetate (PMA) decreased the expressions of GABA A subunits α1, γ2, and δ when administered alone. However, the administration of either KG-501 or staurosporine reversed the inhibitory effects of PMA. The PKC/CREB pathway may negatively regulate the expressions of GABA A receptor subunits α1, γ2, and δ in cultured hippocampal neurons in low Mg 2+ model of epilepsy. Copyright © 2017. Published by Elsevier B.V.

Study of the Role of CREB, BDNF, and VGF Neuropeptide in Long Term Antidepressant Activity of Crocin in the Rat Cerebellum

PubMed Central

Razavi, Bibi Marjan; Sadeghi, Mahdieh; Abnous, Khalil; Vahdati Hasani, Faezeh; Hosseinzadeh, Hossein

2017-01-01

Antidepressant activity of crocin, saffron main component, has been established before. Based on previous study, it is suggested that elevation in the levels of BDNF (brain-derived neurotrophic factor), CREB (cAMP response element binding) and VGF neuropeptide could be considered as one probable molecular mechanisms involved in antidepressant activity of long term crocin administration in the rat hippocampus. In this study we further investigated whether the antidepressant activity of crocin in long term administration was associated with alteration in these factors in the rat cerebellum. Crocin (12.5, 25 and 50 mg/kg/day) and imipramine (10 mg/kg/day) were administered interaperitoneally for 21 days to rats. At the end of experiment, animals were sacrificed and cerebellums were dissected. BDNF, VGF, CREB, and phospho-CREB (P-CREB) protein and mRNA levels in the rat cerebellum were evaluated using Western blot and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). In the current study significant increases in mRNA and protein levels of VGF, CREB and (BDNF) in long term crocin treatment were not observed in the rat cerebellum. Although a slight increase was observed in protein level of P-CREB compared to normal saline, but it was not significant. It is concluded that antidepressant activity of crocin might be partially mediated to CREB. Moreover, other factors rather than BDNF and VGF neuropeptides may alter following long term crocin treatment in the cerebellum. To understand the precise mechanism of crocin antidepressant effects in the cerebellum, longer duration of crocin treatment in further studies is recommended. PMID:29552054

Integrative analysis identifies targetable CREB1/FoxA1 transcriptional co-regulation as a predictor of prostate cancer recurrence.

PubMed

Sunkel, Benjamin; Wu, Dayong; Chen, Zhong; Wang, Chiou-Miin; Liu, Xiangtao; Ye, Zhenqing; Horning, Aaron M; Liu, Joseph; Mahalingam, Devalingam; Lopez-Nicora, Horacio; Lin, Chun-Lin; Goodfellow, Paul J; Clinton, Steven K; Jin, Victor X; Chen, Chun-Liang; Huang, Tim H-M; Wang, Qianben

2016-05-19

Identifying prostate cancer-driving transcription factors (TFs) in addition to the androgen receptor promises to improve our ability to effectively diagnose and treat this disease. We employed an integrative genomics analysis of master TFs CREB1 and FoxA1 in androgen-dependent prostate cancer (ADPC) and castration-resistant prostate cancer (CRPC) cell lines, primary prostate cancer tissues and circulating tumor cells (CTCs) to investigate their role in defining prostate cancer gene expression profiles. Combining genome-wide binding site and gene expression profiles we define CREB1 as a critical driver of pro-survival, cell cycle and metabolic transcription programs. We show that CREB1 and FoxA1 co-localize and mutually influence each other's binding to define disease-driving transcription profiles associated with advanced prostate cancer. Gene expression analysis in human prostate cancer samples found that CREB1/FoxA1 target gene panels predict prostate cancer recurrence. Finally, we showed that this signaling pathway is sensitive to compounds that inhibit the transcription co-regulatory factor MED1. These findings not only reveal a novel, global transcriptional co-regulatory function of CREB1 and FoxA1, but also suggest CREB1/FoxA1 signaling is a targetable driver of prostate cancer progression and serves as a biomarker of poor clinical outcomes. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Period1 gates the circadian modulation of memory-relevant signaling in mouse hippocampus by regulating the nuclear shuttling of the CREB kinase pP90RSK.

PubMed

Rawashdeh, Oliver; Jilg, Antje; Maronde, Erik; Fahrenkrug, Jan; Stehle, Jörg H

2016-09-01

Memory performance varies over a 24-h day/night cycle. While the detailed underlying mechanisms are yet unknown, recent evidence suggests that in the mouse hippocampus, rhythmic phosphorylation of mitogen-activated protein kinase (MAPK) and cyclic adenosine monophosphate response element-binding protein (CREB) are central to the circadian (~ 24 h) regulation of learning and memory. We recently identified the clock protein PERIOD1 (PER1) as a vehicle that translates information encoding time of day to hippocampal plasticity. We here elaborate how PER1 may gate the sensitivity of memory-relevant hippocampal signaling pathways. We found that in wild-type mice (WT), spatial learning triggers CREB phosphorylation only during the daytime, and that this effect depends on the presence of PER1. The time-of-day-dependent induction of CREB phosphorylation can be reproduced pharmacologically in acute hippocampal slices prepared from WT mice, but is absent in preparations made from Per1-knockout (Per1(-/-) ) mice. We showed that the PER1-dependent CREB phosphorylation is regulated downstream of MAPK. Stimulation of WT hippocampal neurons triggered the co-translocation of PER1 and the CREB kinase pP90RSK (pMAPK-activated ribosomal S6 kinase) into the nucleus. In hippocampal neurons from Per1(-/-) mice, however, pP90RSK remained perinuclear. A co-immunoprecipitation assay confirmed a high-affinity interaction between PER1 and pP90RSK. Knocking down endogenous PER1 in hippocampal cells inhibited adenylyl cyclase-dependent CREB activation. Taken together, the PER1-dependent modulation of cytoplasmic-to-nuclear signaling in the murine hippocampus provides a molecular explanation for how the circadian system potentially shapes a temporal framework for daytime-dependent memory performance, and adds a novel facet to the versatility of the clock gene protein PER1. We provide evidence that the circadian clock gene Period1 (Per1) regulates CREB phosphorylation in the mouse hippocampus, sculpturing time-of-day-dependent memory formation. This molecular mechanism constitutes the functional link between circadian rhythms and learning efficiency. In hippocampal neurons of wild-type mice, pP90RSK translocates into the nucleus upon stimulation with forskolin (left), whereas in Period1-knockout (Per1(-/-) ) mice (right) the kinase is trapped at the nuclear periphery, unable to efficiently phosphorylate nuclear CREB. Consequently, the presence of PER1 in hippocampal neurons is a prerequisite for the time-of-day-dependent phosphorylation of CREB, as it regulates the shuttling of pP90RSK into the nucleus. Representative immunofluorescence images show a temporal difference in phosphorylated cAMP response element-binding protein (pCREB; green color) levels in all regions of the dorsal hippocampus between a wild-type C3H mouse (WT; left) and a Period1-knockout (Per1(-/-) ; right) mouse. Images were taken 2 h after lights on, thus, when fluctuating levels of pCREB peak in WT mouse hippocampus. Insets show a representative hippocampal neuron, in response to activating cAMP signaling, stained for the neuronal marker NeuN (red), the nuclear marker DAPI (blue) and the activated CREB kinase pP90RSK (green). The image was taken 2 h after light onset (at the peak of the endogenous CREB phosphorylation that fluctuates with time of day). Magnification: 100X, inset 400X. Read the Editorial Highlight for this article on page 650. Cover image for this issue: doi: 10.1111/jnc.13332. © 2016 International Society for Neurochemistry.

Identification of differential pathways in papillary thyroid carcinoma utilizing pathway co-expression analysis.

PubMed

Qiu, Wei-Hai; Chen, Gui-Yan; Cui, Lu; Zhang, Ting-Ming; Wei, Feng; Yang, Yong

2016-01-01

To identify differential pathways between papillary thyroid carcinoma (PTC) patients and normal controls utilizing a novel method which combined pathway with co-expression network. The proposed method included three steps. In the first step, we conducted pretreatments for background pathways and gained representative pathways in PTC. Subsequently, a co-expression network for representative pathways was constructed using empirical Bayes (EB) approach to assign a weight value for each pathway. Finally, random model was extracted to set the thresholds of identifying differential pathways. We obtained 1267 representative pathways and their weight values based on the co-expressed pathway network, and then by meeting the criterion (Weight > 0.0296), 87 differential pathways in total across PTC patients and normal controls were identified. The top three ranked differential pathways were CREB phosphorylation, attachment of GPI anchor to urokinase plasminogen activator receptor (uPAR) and loss of function of SMAD2/3 in cancer. In conclusion, we successfully identified differential pathways (such as CREB phosphorylation, attachment of GPI anchor to uPAR and post-translational modification: synthesis of GPI-anchored proteins) for PTC using the proposed pathway co-expression method, and these pathways might be potential biomarkers for target therapy and detection of PTC.

Inhibition of phosphodiesterase10A attenuates morphine-induced conditioned place preference.

PubMed

Mu, Ying; Ren, Zhaoxiang; Jia, Jia; Gao, Bo; Zheng, Longtai; Wang, Guanghui; Friedman, Eitan; Zhen, Xuechu

2014-09-25

Phosphodiesterase (PDE) 10A is selectively expressed in medium spiny neurons of the striatum. Nucleus accumbens (NAc) is a key region that mediates drug reward and addiction-related behaviors. To investigate the potential role of PDE10A in the reinforcement properties of morphine, we tested the effect of MP-10, a selective inhibitor of PDE10A, on acquisition, expression, and extinction of morphine-induced conditioned place preference (CPP). The results show that 2.5 mg/kg MP-10, administered subcutaneously, significantly inhibited the acquisition of morphine-induced CPP. The same dose of MP-10 alone did not result in the CPP. Moreover, MP-10 did not alter the expression of morphine-induced CPP, but did accelerate the extinction of morphine-induced CPP. Additionally, chronic treatment with 2.5 mg/kg MP-10 decreased expression of phosphorylated CREB (pCREB), activated cAMP response element binding protein, in dorsomedial striatum, in shell of NAc, and in anterior cingulate cortex (ACC) as well as decreased expression of ΔFosB in the shell of NAc and ACC. The results suggest that inhibition of PDE10A may have therapeutic potential in the treatment of opioid addiction.

Control of energy balance by hypothalamic gene circuitry involving two nuclear receptors, neuron-derived orphan receptor 1 and glucocorticoid receptor.

PubMed

Kim, Sun-Gyun; Lee, Bora; Kim, Dae-Hwan; Kim, Juhee; Lee, Seunghee; Lee, Soo-Kyung; Lee, Jae W

2013-10-01

Nuclear receptors (NRs) regulate diverse physiological processes, including the central nervous system control of energy balance. However, the molecular mechanisms for the central actions of NRs in energy balance remain relatively poorly defined. Here we report a hypothalamic gene network involving two NRs, neuron-derived orphan receptor 1 (NOR1) and glucocorticoid receptor (GR), which directs the regulated expression of orexigenic neuropeptides agouti-related peptide (AgRP) and neuropeptide Y (NPY) in response to peripheral signals. Our results suggest that the anorexigenic signal leptin induces NOR1 expression likely via the transcription factor cyclic AMP response element-binding protein (CREB), while the orexigenic signal glucocorticoid mobilizes GR to inhibit NOR1 expression by antagonizing the action of CREB. Also, NOR1 suppresses glucocorticoid-dependent expression of AgRP and NPY. Consistently, relative to wild-type mice, NOR1-null mice showed significantly higher levels of AgRP and NPY and were less responsive to leptin in decreasing the expression of AgRP and NPY. These results identify mutual antagonism between NOR1 and GR to be a key rheostat for peripheral metabolic signals to centrally control energy balance.

Hydrogen-Deuterium Exchange Mass Spectrometry Reveals Calcium Binding Properties and Allosteric Regulation of Downstream Regulatory Element Antagonist Modulator (DREAM).

PubMed

Zhang, Jun; Li, Jing; Craig, Theodore A; Kumar, Rajiv; Gross, Michael L

2017-07-18

Downstream regulatory element antagonist modulator (DREAM) is an EF-hand Ca 2+ -binding protein that also binds to a specific DNA sequence, downstream regulatory elements (DRE), and thereby regulates transcription in a calcium-dependent fashion. DREAM binds to DRE in the absence of Ca 2+ but detaches from DRE under Ca 2+ stimulation, allowing gene expression. The Ca 2+ binding properties of DREAM and the consequences of the binding on protein structure are key to understanding the function of DREAM. Here we describe the application of hydrogen-deuterium exchange mass spectrometry (HDX-MS) and site-directed mutagenesis to investigate the Ca 2+ binding properties and the subsequent conformational changes of full-length DREAM. We demonstrate that all EF-hands undergo large conformation changes upon calcium binding even though the EF-1 hand is not capable of binding to Ca 2+ . Moreover, EF-2 is a lower-affinity site compared to EF-3 and -4 hands. Comparison of HDX profiles between wild-type DREAM and two EF-1 mutated constructs illustrates that the conformational changes in the EF-1 hand are induced by long-range structural interactions. HDX analyses also reveal a conformational change in an N-terminal leucine-charged residue-rich domain (LCD) remote from Ca 2+ -binding EF-hands. This LCD domain is responsible for the direct interaction between DREAM and cAMP response element-binding protein (CREB) and regulates the recruitment of the co-activator, CREB-binding protein. These long-range interactions strongly suggest how conformational changes transmit the Ca 2+ signal to CREB-mediated gene transcription.

CREB trans-activation of disruptor of telomeric silencing-1 mediates forskolin inhibition of CTGF transcription in mesangial cells.

PubMed

Yu, Zhiyuan; Kong, Qun; Kone, Bruce C

2010-03-01

Connective tissue growth factor (CTGF) participates in diverse fibrotic processes including glomerulosclerosis. The adenylyl cyclase agonist forskolin inhibits CTGF expression in mesangial cells by unclear mechanisms. We recently reported that the histone H3K79 methyltransferase disruptor of telomeric silencing-1 (Dot1) suppresses CTGF gene expression in collecting duct cells (J Clin Invest 117: 773-783, 2007) and HEK 293 cells (J Biol Chem In press). In the present study, we characterized the involvement of Dot1 in mediating the inhibitory effect of forskolin on CTGF transcription in mouse mesangial cells. Overexpression of Dot1 or treatment with forskolin dramatically suppressed basal CTGF mRNA levels and CTGF promoter-luciferase activity, while hypermethylating H3K79 in chromatin associated with the CTGF promoter. siRNA knockdown of Dot1 abrogated the inhibitory effect of forskolin on CTGF mRNA expression. Analysis of the Dot1 promoter sequence identified a CREB response element (CRE) at -384/-380. Overexpression of CREB enhanced forskolin-stimulated Dot1 promoter activity. A constitutively active CREB mutant (CREB-VP16) strongly induced Dot1 promoter-luciferase activity, whereas overexpression of CREBdLZ-VP16, which lacks the CREB DNA-binding domain, abolished this activation. Mutation of the -384/-380 CRE resulted in 70% lower levels of Dot1 promoter activity. ChIP assays confirmed CREB binding to the Dot1 promoter in chromatin. We conclude that forskolin stimulates CREB-mediated trans-activation of the Dot1 gene, which leads to hypermethylation of histone H3K79 at the CTGF promoter, and inhibition of CTGF transcription. These data are the first to describe regulation of the Dot1 gene, and disclose a complex network of genetic and epigenetic controls on CTGF transcription.

CaMKII and CaMKIV mediate distinct prosurvival signaling pathways in response to depolarization in neurons

PubMed Central

Bok, Jinwoong; Wang, Qiong; Huang, Jie; Green, Steven H.

2007-01-01

By fusing the CaMKII inhibitory peptide AIP to GFP, we constructed a specific and effective CaMKII inhibitor, GFP-AIP. Expression of GFP-AIP and/or dominant-inhibitory CaMKIV in cultured neonatal rat spiral ganglion neurons (SGNs) shows that CaMKII and CaMKIV act additively and in parallel, to mediate the prosurvival effect of depolarization. Depolarization or expression of constitutively-active CaMKII functionally inactivates Bad, indicating that this is one means by which CaMKII promotes neuronal survival. CaMKIV, but not CaMKII, requires CREB to promote SGN survival, consistent with the exclusively nuclear localization of CaMKIV and indicating that the principal prosurvival function of CaMKIV is activation of CREB. Consistent with this, a constitutively-active CREB construct that provides a high level of CREB activity promotes SGN survival, although low levels of CREB activity did not do so. Also, in apoptotic SGNs, activation of CREB by depolarization is disabled, presumably as part of a cellular commitment to apoptosis. PMID:17651987

A Transcription Factor-Binding Domain of the Coactivator CBP Is Essential for Long-Term Memory and the Expression of Specific Target Genes

ERIC Educational Resources Information Center

Oliveira, Ana M. M.; Brindle, Paul K.; Abel, Ted; Wood, Marcelo A.; Attner, Michelle A.

2006-01-01

Transcriptional activation is a key process required for long-term memory formation. Recently, the transcriptional coactivator CREB-binding protein (CBP) was shown to be critical for hippocampus-dependent long-term memory and hippocampal synaptic plasticity. As a coactivator with intrinsic histone acetyltransferase activity, CBP interacts with…

Analgesic effect of paeoniflorin in rats with neonatal maternal separation-induced visceral hyperalgesia is mediated through adenosine A(1) receptor by inhibiting the extracellular signal-regulated protein kinase (ERK) pathway.

PubMed

Zhang, Xiao-Jun; Chen, Hong-Li; Li, Zhi; Zhang, Hong-Qi; Xu, Hong-Xi; Sung, Joseph J Y; Bian, Zhao-Xiang

2009-11-01

Paeoniflorin (PF), a chief active ingredient in the root of Paeonia lactiflora Pall (family Ranunculaceae), is effective in relieving colorectal distention (CRD)-induced visceral pain in rats with visceral hyperalgesia induced by neonatal maternal separation (NMS). This study aimed at exploring the underlying mechanisms of PF's analgesic effect on CRD-evoked nociceptive signaling in the central nervous system (CNS) and investigating whether the adenosine A(1) receptor is involved in PF's anti-nociception. CRD-induced visceral pain as well as phosphorylated-extracellular signal-regulated protein kinase (p-ERK) and phospho-cAMP response element-binding protein (p-CREB) expression in the CNS structures of NMS rats were suppressed by NMDA receptor antagonist dizocilpine (MK-801) and ERK phosphorylation inhibitor U0126. PF could similarly inhibit CRD-evoked p-ERK and c-Fos expression in laminae I-II of the lumbosacral dorsal horn and anterior cingulate cortex (ACC). PF could also reverse the CRD-evoked increased glutamate concentration by CRD as shown by dynamic microdialysis monitoring in ACC, whereas, DPCPX, an antagonist of adenosine A(1) receptor, significantly blocked the analgesic effect of PF and PF's inhibition on CRD-induced p-ERK and p-CREB expression. These results suggest that PF's analgesic effect is possibly mediated by adenosine A(1) receptor by inhibiting CRD-evoked glutamate release and the NMDA receptor dependent ERK signaling.

Lgr4 Protein Deficiency Induces Ataxia-like Phenotype in Mice and Impairs Long Term Depression at Cerebellar Parallel Fiber-Purkinje Cell Synapses*

PubMed Central

Guan, Xin; Duan, Yanhong; Zeng, Qingwen; Pan, Hongjie; Qian, Yu; Li, Dali; Cao, Xiaohua; Liu, Mingyao

2014-01-01

Cerebellar dysfunction causes ataxia characterized by loss of balance and coordination. Until now, the molecular and neuronal mechanisms of several types of inherited cerebellar ataxia have not been completely clarified. Here, we report that leucine-rich G protein-coupled receptor 4 (Lgr4/Gpr48) is highly expressed in Purkinje cells (PCs) in the cerebellum. Deficiency of Lgr4 leads to an ataxia-like phenotype in mice. Histologically, no obvious morphological changes were observed in the cerebellum of Lgr4 mutant mice. However, the number of PCs was slightly but significantly reduced in Lgr4−/− mice. In addition, in vitro electrophysiological analysis showed an impaired long term depression (LTD) at parallel fiber-PC (PF-PC) synapses in Lgr4−/− mice. Consistently, immunostaining experiments showed that the level of phosphorylated cAMP-responsive element-binding protein (Creb) was significantly decreased in Lgr4−/− PCs. Furthermore, treatment with forskolin, an adenylyl cyclase agonist, rescued phospho-Creb in PCs and reversed the impairment in PF-PC LTD in Lgr4−/− cerebellar slices, indicating that Lgr4 is an upstream regulator of Creb signaling, which is underlying PF-PC LTD. Together, our findings demonstrate for first time an important role for Lgr4 in motor coordination and cerebellar synaptic plasticity and provide a potential therapeutic target for certain types of inherited cerebellar ataxia. PMID:25063812

Irradiation induces regionally specific alterations in pro-inflammatory environments in rat brain

PubMed Central

Lee, Won Hee; Sonntag, William E.; Mitschelen, Matthew; Yan, Han; Lee, Yong Woo

2010-01-01

Purpose Pro-inflammatory environments in the brain have been implicated in the onset and progression of neurological disorders. In the present study, we investigate the hypothesis that brain irradiation induces regionally specific alterations in cytokine gene and protein expression. Materials and methods Four month old F344 × BN rats received either whole brain irradiation with a single dose of 10 Gy γ-rays or sham-irradiation, and were maintained for 4, 8, and 24 h following irradiation. The mRNA and protein expression levels of pro-inflammatory mediators were analysed by real-time reverse transcriptase-polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and immunofluorescence staining. To elucidate the molecular mechanisms of irradiation-induced brain inflammation, effects of irradiation on the DNA-binding activity of pro-inflammatory transcription factors were also examined. Results A significant and marked up-regulation of mRNA and protein expression of pro-inflammatory mediators, including tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and monocyte chemoattractant protein-1 (MCP-1), was observed in hippocampal and cortical regions isolated from irradiated brain. Cytokine expression was regionally specific since TNF-α levels were significantly elevated in cortex compared to hippocampus (57% greater) and IL-1β levels were elevated in hippocampus compared to cortical samples (126% greater). Increases in cytokine levels also were observed after irradiation of mouse BV-2 microglial cells. A series of electrophoretic mobility shift assays (EMSA) demonstrated that irradiation significantly increased activation of activator protein-1 (AP-1), nuclear factor-κB (NF-κB), and cAMP response element-binding protein (CREB). Conclusion The present study demonstrated that whole brain irradiation induces regionally specific pro-inflammatory environments through activation of AP-1, NF-κB, and CREB and overexpression of TNF-α, IL-1β, and MCP-1 in rat brain and may contribute to unique pathways for the radiation-induced impairments in tissue function. PMID:20148699

Resveratrol prevents CA1 neurons against ischemic injury by parallel modulation of both GSK-3β and CREB through PI3-K/Akt pathways.

PubMed

Simão, Fabrício; Matté, Aline; Pagnussat, Aline S; Netto, Carlos A; Salbego, Christianne G

2012-10-01

Accumulating evidence indicates that resveratrol potently protects against cerebral ischemia damage due to its oxygen free radicals scavenging and antioxidant properties. However, cellular mechanisms that may underlie the neuroprotective effects of resveratrol in brain ischemia are not fully understood yet. This study aimed to investigate the potential association between the neuroprotective effect of resveratrol and the apoptosis/survival signaling pathways, in particular the glycogen synthase kinase 3 (GSK-3β) and cAMP response element-binding protein (CREB) through phosphatidylinositol 3-kinase (PI3-K)-dependent pathway. An experimental model of global cerebral ischemia was induced in rats by the four-vessel occlusion method for 10 min and followed by different periods of reperfusion. Nissl staining indicated extensive neuronal death at 7 days after ischemia/reperfusion. Administration of resveratrol by i.p. injections (30 mg/kg) for 7 days before ischemia significantly attenuated neuronal death. Both GSK-3β and CREB appear to play a critical role in resveratrol neuroprotection through the PI3-K/Akt pathway, as resveratrol pretreatment increased the phosphorylation of Akt, GSK-3β and CREB in 1 h in the CA1 hippocampus after ischemia/reperfusion. Furthermore, administration of LY294002, an inhibitor of PI3-K, compromised the neuroprotective effect of resveratrol and decreased the level of p-Akt, p-GSK-3β and p-CREB after ischemic injury. Taken together, the results suggest that resveratrol protects against delayed neuronal death in the hippocampal CA1 by maintaining the pro-survival states of Akt, GSK-3β and CREB pathways. These data suggest that the neuroprotective effect of resveratrol may be mediated through activation of the PI3-K/Akt signaling pathway, subsequently downregulating expression of GSK-3β and CREB, thereby leading to prevention of neuronal death after brain ischemia in rats. © 2012 The Authors. European Journal of Neuroscience © 2012 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.

Early transcutaneous electrical nerve stimulation reduces hyperalgesia and decreases activation of spinal glial cells in mice with neuropathic pain.

PubMed

Matsuo, Hideaki; Uchida, Kenzo; Nakajima, Hideaki; Guerrero, Alexander Rodriguez; Watanabe, Shuji; Takeura, Naoto; Sugita, Daisuke; Shimada, Seiichiro; Nakatsuka, Terumasa; Baba, Hisatoshi

2014-09-01

Although transcutaneous electrical nerve stimulation (TENS) is widely used for the treatment of neuropathic pain, its effectiveness and mechanism of action in reducing neuropathic pain remain uncertain. We investigated the effects of early TENS (starting from the day after surgery) in mice with neuropathic pain, on hyperalgesia, glial cell activation, pain transmission neuron sensitization, expression of proinflammatory cytokines, and opioid receptors in the spinal dorsal horn. Following nerve injury, TENS and behavioral tests were performed every day. Immunohistochemical, immunoblot, and flow cytometric analysis of the lumbar spinal cord were performed after 8 days. Early TENS reduced mechanical and thermal hyperalgesia and decreased the activation of microglia and astrocytes (P<0.05). In contrast, the application of TENS at 1 week (TENS-1w) or 2 weeks (TENS-2w) after injury was ineffective in reducing hyperalgesia (mechanical and thermal) or activation of microglia and astrocytes. Early TENS decreased p-p38 within microglia (P<0.05), the expression levels of protein kinase C (PKC-γ), and phosphorylated anti-phospho-cyclic AMP response element-binding protein (p-CREB) in the superficial spinal dorsal horn neurons (P<0.05), mitogen-activated protein (MAP) kinases, and proinflammatory cytokines, and increased the expression levels of opioid receptors (P<0.05). The results suggested that the application of early TENS relieved hyperalgesia in our mouse model of neuropathic pain by inhibiting glial activation, MAP kinase activation, PKC-γ, and p-CREB expression, and proinflammatory cytokines expression, as well as maintenance of spinal opioid receptors. The findings indicate that TENS treatment is more effective when applied as early after nerve injury as possible. Copyright © 2014 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

Mechanism of repression of the inhibin alpha-subunit gene by inducible 3',5'-cyclic adenosine monophosphate early repressor.

PubMed

Burkart, Anna D; Mukherjee, Abir; Mayo, Kelly E

2006-03-01

The rodent ovary is regulated throughout the reproductive cycle to maintain normal cyclicity. Ovarian follicular development is controlled by changes in gene expression in response to the gonadotropins FSH and LH. The inhibin alpha-subunit gene belongs to a group of genes that is positively regulated by FSH and negatively regulated by LH. Previous studies established an important role for inducible cAMP early repressor (ICER) in repression of alpha-inhibin. These current studies investigate the mechanisms of repression by ICER. It is not clear whether all four ICER isoforms expressed in the ovary can act as repressors of the inhibin alpha-subunit gene. EMSAs demonstrate binding of all isoforms to the inhibin alpha-subunit CRE (cAMP response element), and transfection studies demonstrate that all isoforms can repress the inhibin alpha-subunit gene. Repression by ICER is dependent on its binding to DNA as demonstrated by mutations to ICER's DNA-binding domain. These mutational studies also demonstrate that repression by ICER is not dependent on heterodimerization with CREB (CRE-binding protein). Competitive EMSAs show that ICER effectively competes with CREB for binding to the inhibin alpha CRE in vitro. Chromatin immunoprecipitation assays demonstrate a replacement of CREB dimers bound to the inhibin alpha CRE by ICER dimers in ovarian granulosa cells in response to LH signaling. Thus, there is a temporal association of transcription factors bound to the inhibin alpha-CRE controlling inhibin alpha-subunit gene expression.

Role of olfactory bulb serotonin in olfactory learning in the greater short-nosed fruit bat, Cynopterus sphinx (Chiroptera: Pteropodidae).

PubMed

Ganesh, Ambigapathy; Bogdanowicz, Wieslaw; Haupt, Moritz; Marimuthu, Ganapathy; Rajan, Koilmani Emmanuvel

2010-09-17

The role of olfactory bulb (OB) serotonin [5-hydroxytryptamine (5-HT)] in olfactory learning and memory was tested in the greater short-nosed fruit bat, Cynopterus sphinx (family Pteropodidae). Graded concentrations (25, 40, and 60microg) of 5,7-dihydroxytryptamine (5,7-DHT) or saline were injected into the OB of bats one day before training to the novel odor. In a behavioral test, 5,7-DHT (60microg) injected bats made significantly fewer feeding attempts and bouts when compared to saline-injected bats during learning and in the memory test. Subsequent biochemical analysis showed that 5-HT level was effectively depleted in the OB of 5,7-DHT injected bats. To test odor-induced 5-HT mediated changes in 5-HT receptors and second messenger cascade in the OB, we examined the expression of 5-HT receptors and mitogen-activated protein kinase (MAPK)/Erk cascade after training to the novel odor. We found that odor stimulation up-regulated the expression of 5-HT(1A) receptor, Erk1 and Creb1 mRNA, and phosphorylation of ERK1 and CREB1. Odor stimulation failed to induce expression in 5-HT-depleted bats, which is similar to control bats and significantly low compared to saline-treated bats. Together these data revealed that the level of 5-HT in the OB may regulate olfactory learning and memory in C. sphinx through Erk and CREB.

Lithium promotes DNA stability and survival of ischemic retinal neurocytes by upregulating DNA ligase IV.

PubMed

Yang, Ying; Wu, Nandan; Tian, Sijia; Li, Fan; Hu, Huan; Chen, Pei; Cai, Xiaoxiao; Xu, Lijun; Zhang, Jing; Chen, Zhao; Ge, Jian; Yu, Keming; Zhuang, Jing

2016-11-17

Neurons display genomic fragility and show fragmented DNA in pathological degeneration. A failure to repair DNA breaks may result in cell death or apoptosis. Lithium protects retinal neurocytes following nutrient deprivation or partial nerve crush, but the underlying mechanisms are not well defined. Here we demonstrate that pretreatment with lithium protects retinal neurocytes from ischemia-induced damage and enhances light response in rat retina following ischemia-reperfusion injury. Moreover, we found that DNA nonhomologous end-joining (NHEJ) repair is implicated in this process because in ischemic retinal neurocytes, lithium significantly reduces the number of γ-H2AX foci (well-characterized markers of DNA double-strand breaks in situ) and increases the DNA ligase IV expression level. Furthermore, we also demonstrate that nuclear respiratory factor 1 (Nrf-1) and phosphorylated cyclic AMP-response element binding protein-1 (P-CREB1) bind to ligase IV promoter to cause upregulation of ligase IV in neurocytes. The ischemic upregulation of Nrf-1 and lithium-induced increase of P-CREB1 cooperate to promote transcription of ligase IV. Short hairpin RNAs against Nrf-1 and CREB1 could significantly inhibit the increase in promoter activity and expression of ligase IV observed in the control oligos following lithium treatment in retinal neurocytes. More importantly, ischemic stimulation triggers the expression of ligase IV. Taken together, our results thus reveal a novel mechanism that lithium offers neuroprotection from ischemia-induced damage by enhancing DNA NHEJ repair.

Lithium promotes DNA stability and survival of ischemic retinal neurocytes by upregulating DNA ligase IV

PubMed Central

Yang, Ying; Wu, Nandan; Tian, Sijia; Li, Fan; Hu, Huan; Chen, Pei; Cai, Xiaoxiao; Xu, Lijun; Zhang, Jing; Chen, Zhao; Ge, Jian; Yu, Keming; Zhuang, Jing

2016-01-01

Neurons display genomic fragility and show fragmented DNA in pathological degeneration. A failure to repair DNA breaks may result in cell death or apoptosis. Lithium protects retinal neurocytes following nutrient deprivation or partial nerve crush, but the underlying mechanisms are not well defined. Here we demonstrate that pretreatment with lithium protects retinal neurocytes from ischemia-induced damage and enhances light response in rat retina following ischemia–reperfusion injury. Moreover, we found that DNA nonhomologous end-joining (NHEJ) repair is implicated in this process because in ischemic retinal neurocytes, lithium significantly reduces the number of γ-H2AX foci (well-characterized markers of DNA double-strand breaks in situ) and increases the DNA ligase IV expression level. Furthermore, we also demonstrate that nuclear respiratory factor 1 (Nrf-1) and phosphorylated cyclic AMP-response element binding protein-1 (P-CREB1) bind to ligase IV promoter to cause upregulation of ligase IV in neurocytes. The ischemic upregulation of Nrf-1 and lithium-induced increase of P-CREB1 cooperate to promote transcription of ligase IV. Short hairpin RNAs against Nrf-1 and CREB1 could significantly inhibit the increase in promoter activity and expression of ligase IV observed in the control oligos following lithium treatment in retinal neurocytes. More importantly, ischemic stimulation triggers the expression of ligase IV. Taken together, our results thus reveal a novel mechanism that lithium offers neuroprotection from ischemia-induced damage by enhancing DNA NHEJ repair. PMID:27853172

Functional characterization of the human phosphodiesterase 7A1 promoter.

PubMed Central

Torras-Llort, Mònica; Azorín, Fernando

2003-01-01

In this paper, the human phosphodiesterase 7A1 (h PDE7A1 ) promoter region was identified and functionally characterized. Transient transfection experiments indicated that a 2.9 kb fragment of the h PDE7A1 5'-flanking region, to position -2907, has strong promoter activity in Jurkat T-cells. Deletion analysis showed that the proximal region, up to position -988, contains major cis -regulatory elements of the h PDE7A1 promoter. This minimal promoter region contains a regulatory CpG island which is essential for promoter activity. The CpG island contains three potential cAMP-response-element-binding protein (CREB)-binding sites that, as judged by in vivo dimethyl sulphate (DMS) footprinting, are occupied in Jurkat T-cells. Moreover, over-expression of CREB results in increased promoter activity, but, on the other hand, promoter activity decreases when a dominant-negative form of CREB (KCREB) is over-expressed. In vivo DMS footprinting strongly indicates that other transcription factors, such Ets-2, nuclear factor of activated T-cells 1 (NFAT-1) and nuclear factor kappaB (NF-kappaB), might also contribute to the regulation of h PDE7A1 promoter. Finally, h PDE7A1 promoter was found to be induced by treatment with PMA, but not by treatment with dibutyryl cAMP or forskolin. These results provide insights into the factors and mechanisms that regulate expression of the h PDE7A gene. PMID:12737631

Prenatal caffeine exposure induced a lower level of fetal blood leptin mainly via placental mechanism

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Yi-meng; Luo, Han-wen; Kou, Hao

It's known that blood leptin level is reduced in intrauterine growth retardation (IUGR) fetus, and placental leptin is the major source of fetal blood leptin. This study aimed to investigate the decreased fetal blood leptin level by prenatal caffeine exposure (PCE) and its underlying placental mechanisms. Pregnant Wistar rats were intragastrically administered caffeine (30–120 mg/kg day) from gestational day 9 to 20. The level of fetal serum leptin and the expression of placental leptin-related genes were analyzed. Furthermore, we investigated the molecular mechanism of the reduced placental leptin's expression by treatment with caffeine (0.8–20 μM) in the BeWo cells. Inmore » vivo, PCE significantly decreased fetal serum leptin level in caffeine dose-dependent manner. Meanwhile, placental mRNA expression of adenosine A2a receptor (Adora2a), cAMP-response element binding protein (CREB), a short-type leptin receptor (Ob-Ra) and leptin was reduced in the PCE groups. In vitro, caffeine significantly decreased the mRNA expression of leptin, CREB and ADORA2A in concentration and time-dependent manners. The addition of ADORA2A agonist or adenylyl cyclase (AC) agonist reversed the inhibition of leptin expression induced by caffeine. PCE induced a lower level of fetal blood leptin, which the primary mechanism is that caffeine inhibited antagonized Adora2a and AC activities to decreased cAMP synthesis, thus inhibited the expression of the transcription factor CREB and target gene leptin in the placenta. Meantime, the reduced transportation of maternal leptin by placental Ob-Ra also contributed to the reduced fetal blood leptin. Together, PCE decreased fetal blood leptin mainly via reducing the expression and transportation of leptin in the placenta. - Highlights: • Caffeine reduced fetal blood leptin level. • Caffeine inhibited placental leptin production and transport. • Caffeine down-regulated placental leptin expression via antagonizing ADORA2. • Caffeine inhibited placental leptin transport via decreased OB-Ra expression.« less

Functional Connectivity of Multiple Brain Regions Required for the Consolidation of Social Recognition Memory.

PubMed

Tanimizu, Toshiyuki; Kenney, Justin W; Okano, Emiko; Kadoma, Kazune; Frankland, Paul W; Kida, Satoshi

2017-04-12

Social recognition memory is an essential and basic component of social behavior that is used to discriminate familiar and novel animals/humans. Previous studies have shown the importance of several brain regions for social recognition memories; however, the mechanisms underlying the consolidation of social recognition memory at the molecular and anatomic levels remain unknown. Here, we show a brain network necessary for the generation of social recognition memory in mice. A mouse genetic study showed that cAMP-responsive element-binding protein (CREB)-mediated transcription is required for the formation of social recognition memory. Importantly, significant inductions of the CREB target immediate-early genes c-fos and Arc were observed in the hippocampus (CA1 and CA3 regions), medial prefrontal cortex (mPFC), anterior cingulate cortex (ACC), and amygdala (basolateral region) when social recognition memory was generated. Pharmacological experiments using a microinfusion of the protein synthesis inhibitor anisomycin showed that protein synthesis in these brain regions is required for the consolidation of social recognition memory. These findings suggested that social recognition memory is consolidated through the activation of CREB-mediated gene expression in the hippocampus/mPFC/ACC/amygdala. Network analyses suggested that these four brain regions show functional connectivity with other brain regions and, more importantly, that the hippocampus functions as a hub to integrate brain networks and generate social recognition memory, whereas the ACC and amygdala are important for coordinating brain activity when social interaction is initiated by connecting with other brain regions. We have found that a brain network composed of the hippocampus/mPFC/ACC/amygdala is required for the consolidation of social recognition memory. SIGNIFICANCE STATEMENT Here, we identify brain networks composed of multiple brain regions for the consolidation of social recognition memory. We found that social recognition memory is consolidated through CREB-meditated gene expression in the hippocampus, medial prefrontal cortex, anterior cingulate cortex (ACC), and amygdala. Importantly, network analyses based on c-fos expression suggest that functional connectivity of these four brain regions with other brain regions is increased with time spent in social investigation toward the generation of brain networks to consolidate social recognition memory. Furthermore, our findings suggest that hippocampus functions as a hub to integrate brain networks and generate social recognition memory, whereas ACC and amygdala are important for coordinating brain activity when social interaction is initiated by connecting with other brain regions. Copyright © 2017 the authors 0270-6474/17/374103-14$15.00/0.

Estradiol up-regulates L-type Ca2+ channels via membrane-bound estrogen receptor/phosphoinositide-3-kinase/Akt/cAMP response element-binding protein signaling pathway.

PubMed

Yang, Xiaoyan; Mao, Xiaofang; Xu, Gao; Xing, Shasha; Chattopadhyay, Ansuman; Jin, Si; Salama, Guy

2018-05-01

In long QT syndrome type 2, women are more prone than men to the lethal arrhythmia torsades de pointes. We previously reported that 17β-estradiol (E2) up-regulates L-type Ca 2+ channels and current (I Ca,L ) (∼30%) in rabbit ventricular myocytes by a classic genomic mechanism mediated by estrogen receptor-α (ERα). In long QT syndrome type 2 (I Kr blockade or bradycardia), the higher Ca 2+ influx via I Ca,L causes Ca 2+ overload, spontaneous sarcoplasmic reticulum Ca 2+ release, and reactivation of I Ca,L that triggers early afterdepolarizations and torsades de pointes. The purpose of this study was to investigate the molecular mechanisms whereby E2 up-regulates I Ca,L , which are poorly understood. H9C2 and rat myocytes were incubated with E2 ± ER antagonist, or inhibitors of downstream transcription factors, for 24 hours, followed by western blots of Cav1.2α1C and voltage-clamp measurements of I Ca,L . Incubation of H9C2 cells with E2 (10-100 nM) increased I Ca,L density and Cav1.2α1C expression, which were suppressed by the ER antagonist ICI182,780 (1 μM). Enhanced I Ca,L and Cav1.2α1C expression by E2 was suppressed by inhibitors of phosphoinositide-3-kinase (Pi3K) (30 μM LY294002; P <.05) and Akt (5 μM MK2206) but not of mitogen-activated protein kinase (5 μM U0126) or protein kinase A (1 μM KT5720). E2 incubation increased p-CREB via the Pi3K/Akt pathway, reached a peak in 20 minutes (3-fold), and leveled off to 1.5-fold 24 hours later. Furthermore, a CREB decoy oligonucleotide inhibited E2-induced Cav1.2α1C expression, whereas membrane-impermeable E2 (E2-bovine serum albumin) was equally effective at Cav1.2α1C up-regulation as E2. Estradiol up-regulates Cav1.2α1C and I Ca,L via plasma membrane ER and by activating Pi3K, Akt, and CREB signaling. The promoter regions of the CACNA1C gene (human-rabbit-rat) contain adjacent/overlapping binding sites for p-CREB and ERα, which suggests a synergistic regulation by these pathways. Copyright © 2018 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.

Crosslinking transcription factors to their recognition sequences with PtII complexes

NASA Technical Reports Server (NTRS)

Chu, B. C.; Orgel, L. E.

1992-01-01

We have prepared phosphorothioate-containing cyclic oligodeoxynucleotides that fold into 'dumbbells' containing CRE and TRE sequences, the binding sequences for the CREB and JUN proteins, respectively. Six phosphorothioate residues were introduced into each of the recognition sequences. K2PtCl4 crosslinks CRE to CREB and TRE to JUN. The extent of crosslinking is about eight times greater than that observed with standard oligodeoxynucleotides and amounts to 30-50% of the efficiency of non-covalent association as estimated by gel-shift assays. Crosslinking is reversed by incubation with NaCN. The crosslinking reaction is specific--a dumbbell oligonucleotide with six phosphorothioate groups introduced into the Sp1 recognition sequence could not be crosslinked efficiently to CREB or JUN proteins with K2PtCl4. The binding of TRE to CREB is not strong enough for effective detection by gel-shift assays, but the TRE-CREB complex is crosslinked efficiently by K2PtCl4 and can then readily be detected.

Parathyroid hormone regulation of the human bone sialoprotein gene transcription is mediated through two cAMP response elements.

PubMed

Araki, Shouta; Mezawa, Masaru; Sasaki, Yoko; Yang, Li; Li, Zhengyang; Takai, Hideki; Nakayama, Youhei; Ogata, Yorimasa

2009-03-01

Parathyroid hormone (PTH) regulates serum calcium and inorganic phosphate levels through its actions on kidney and bone. Bone sialoprotein (BSP) is an early marker of osteoblast differentiation and bone metabolism. We here report that two cAMP response elements (CRE) in the human BSP gene promoter are target of PTH. In human osteoblast-like Saos2 cells, PTH (human 1-34 PTH, 10 nM) increased BSP mRNA and protein levels at 3 h. From transient transfection assays, 2- to 2.5-fold increase in transcription by PTH was observed at 3 and 6 h in -184, -211, -428, -868, and -927 luciferase constructs that included the human BSP gene promoter. Effect of PTH was abrogated by 2 bp mutations in either the CRE1 (-79 to -72) or CRE2 (-674 to -667). Luciferase activities induced by PTH were blocked by protein kinase A inhibitor H89 and tyrosine kinase inhibitor herbimycin A. Gel shift analyses showed that PTH increased binding of nuclear proteins to the CRE1 and CRE2 elements. The CRE1-protein and CRE2-protein complexes were disrupted by CRE binding protein 1 (CREB1) antibodies and supershifted by phospho-CREB1 antibody. ChIP assays detected binding of CREB1 and phospho-CREB1 to a chromatin fragment containing CRE1 and CRE2, and increased binding of phospho-CREB1 to the both sites. These studies demonstrate that PTH stimulates human BSP gene transcription by targeting the two CREs in the promoter of the human BSP gene.

The CRTC1-SIK1 Pathway Regulates Entrainment of the Circadian Clock

PubMed Central

Jagannath, Aarti; Butler, Rachel; Godinho, Sofia I.H.; Couch, Yvonne; Brown, Laurence A.; Vasudevan, Sridhar R.; Flanagan, Kevin C.; Anthony, Daniel; Churchill, Grant C.; Wood, Matthew J.A.; Steiner, Guido; Ebeling, Martin; Hossbach, Markus; Wettstein, Joseph G.; Duffield, Giles E.; Gatti, Silvia; Hankins, Mark W.; Foster, Russell G.; Peirson, Stuart N.

2013-01-01

Summary Retinal photoreceptors entrain the circadian system to the solar day. This photic resetting involves cAMP response element binding protein (CREB)-mediated upregulation of Per genes within individual cells of the suprachiasmatic nuclei (SCN). Our detailed understanding of this pathway is poor, and it remains unclear why entrainment to a new time zone takes several days. By analyzing the light-regulated transcriptome of the SCN, we have identified a key role for salt inducible kinase 1 (SIK1) and CREB-regulated transcription coactivator 1 (CRTC1) in clock re-setting. An entrainment stimulus causes CRTC1 to coactivate CREB, inducing the expression of Per1 and Sik1. SIK1 then inhibits further shifts of the clock by phosphorylation and deactivation of CRTC1. Knockdown of Sik1 within the SCN results in increased behavioral phase shifts and rapid re-entrainment following experimental jet lag. Thus SIK1 provides negative feedback, acting to suppress the effects of light on the clock. This pathway provides a potential target for the regulation of circadian rhythms. PMID:23993098

Andrographolide inhibits adipogenesis of 3T3-L1 cells by suppressing C/EBPβ expression and activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Ching-Chu

Andrographolide, a diterpenoid, is the most abundant terpenoid in Andrographis paniculata, a popular Chinese herbal medicine. Andrographolide displays diverse biological activities including hypoglycemia, hypolipidemia, anti-inflammation, and anti-tumorigenesis. Recent evidence indicates that andrographolide displays anti-obesity property by inhibiting lipogenic gene expression, however, the underlying mechanisms remain to be elucidated. In this study, the effects of andrographolide on transcription factor cascade and mitotic clonal expansion in 3T3-L1 preadipocyte differentiation into adipocyte were determined. Andrographolide dose-dependently (0–15 μM) inhibited CCAAT/enhancer-binding protein α (C/EBPα) and C/EBPβ mRNA and protein expression as well as peroxisome proliferator-activated receptor γ (PPARγ) protein level during the adipogenesis ofmore » 3T3-L1 cells. Concomitantly, fatty acid synthase and stearoyl-CoA desaturase expression and lipid accumulation were attenuated by andrographolide. Oil-red O staining further showed that the first 48 h after the initiation of differentiation was critical for andrographolide inhibition of adipocyte formation. Andrographolide inhibited the phosphorylation of PKA and the activation of cAMP response element-binding protein (CREB) in response to a differentiation cocktail, which led to attenuated C/EBPβ expression. In addition, ERK and GSK3β-dependent C/EBPβ phosphorylation was attenuated by andrographolide. Moreover, andrographolide suppressed cyclin A, cyclin E, and CDK2 expression and impaired the progression of mitotic clonal expansion (MCE) by arresting the cell cycle at the Go/G1 phase. Taken together, these results indicate that andrographolide has a potent anti-obesity action by inhibiting PKA-CREB-mediated C/EBPβ expression as well as C/EBPβ transcriptional activity, which halts MCE progression and attenuates C/EBPα and PPARγ expression. - Highlights: • Andrographolide is a diterpenoid phytochemical. • Andrographolide inhibits adipogenesis of 3 T3-L1 adipocytes. • Andrographolide suppresses differentiation cocktail-induced C/EBPβ expression. • Andrographolide attenuates ERK and GSK3β-dependent C/EBPβ activation. • Andrographolide arrests 3 T3-L1 adipocytes at G0/G1 phase.« less

Exercise activates the phosphatidylinositol 3-kinase pathway.

PubMed

Chen, Michael J; Russo-Neustadt, Amelia A

2005-04-27

Physical exercise is known to enhance psychological well-being and coping capacity. Voluntary physical exercise in rats also robustly and rapidly up-regulates hippocampal brain-derived neurotrophic factor (BDNF) mRNA levels, which are potentiated following a regimen of chronic antidepressant treatment. Increased BDNF levels are associated with enhanced activity of cyclic AMP response element binding protein (CREB). So far, relatively little is known about the intracellular signaling mechanisms mediating this effect of exercise. We wished to explore the possibility that exercise and/or antidepressant treatment activate the hippocampal phosphatidylinositol-3 (PI-3) kinase pathway, which mediates cellular survival. In young male Sprague-Dawley rats, we examined the effects of 2 weeks of daily voluntary wheel-running activity and/or tranylcypromine (n = 7 per group) on the levels of the active forms of protein-dependent kinase-1 (PDK-1), PI-3 kinase, phospho-thr308-Akt, phospho-ser473-Akt, and phospho-glycogen synthase kinase-3beta (GSK3beta; inactive form), as well as BDNF, activated CREB, and the phospho-Trk receptor, in the rat hippocampus, and compared these with sedentary saline-treated controls. Immunoblotting analyses revealed that in exercising rats, there was a significant increase in PI-3 kinase expression (4.61 times that of controls, P = 0.0161) and phosphorylation of PDK-1 (2.73 times that of controls, P = 0.0454), thr308-Akt (2.857 times that of controls, P = 0.0082), CREB (60.27 times that of controls, P = 0.05), and Trk (35.3 times that of controls, P < 0.0001) in the hippocampi of exercising animals; BDNF was also increased (3.2 times that of controls), but this was not statistically significant. In rats receiving both exercise and tranylcypromine, BDNF (4.51 times that of controls, P = 0.0068) and PI-3 kinase (4.88 times that of controls, P = 0.0103), and the phospho- forms of Trk (13.67 times that of controls, P = 0.0278), thr308-Akt (3.644 times that of controls, P = 0.0004), GSK-3beta (2.93 times that of controls, P = 0.026), and CREB (88.97 times that of controls, P = 0.0053) were significantly increased. These results suggest that the exercise-induced expression of BDNF is associated with the increased expression of several key intermediates of the PI-3 kinase/Akt pathway, which is known for its role in enhancing neuronal survival.

Hydrocortisone-induced parkin prevents dopaminergic cell death via CREB pathway in Parkinson's disease model.

PubMed

Ham, Sangwoo; Lee, Yun-Il; Jo, Minkyung; Kim, Hyojung; Kang, Hojin; Jo, Areum; Lee, Gum Hwa; Mo, Yun Jeong; Park, Sang Chul; Lee, Yun Song; Shin, Joo-Ho; Lee, Yunjong

2017-04-03

Dysfunctional parkin due to mutations or post-translational modifications contributes to dopaminergic neurodegeneration in Parkinson's disease (PD). Overexpression of parkin provides protection against cellular stresses and prevents dopamine cell loss in several PD animal models. Here we performed an unbiased high-throughput luciferase screening to identify chemicals that can increase parkin expression. Among promising parkin inducers, hydrocortisone possessed the most favorable profiles including parkin induction ability, cell protection ability, and physicochemical property of absorption, distribution, metabolism, and excretion (ADME) without inducing endoplasmic reticulum stress. We found that hydrocortisone-induced parkin expression was accountable for cell protection against oxidative stress. Hydrocortisone-activated parkin expression was mediated by CREB pathway since gRNA to CREB abolished hydrocortisone's ability to induce parkin. Finally, hydrocortisone treatment in mice increased brain parkin levels and prevented 6-hydroxy dopamine induced dopamine cell loss when assessed at 4 days after the toxin's injection. Our results showed that hydrocortisone could stimulate parkin expression via CREB pathway and the induced parkin expression was accountable for its neuroprotective effect. Since glucocorticoid is a physiological hormone, maintaining optimal levels of glucocorticoid might be a potential therapeutic or preventive strategy for Parkinson's disease.

Neuroprotective effects of various doses of topiramate against methylphenidate-induced oxidative stress and inflammation in isolated rat amygdala: the possible role of CREB/BDNF signaling pathway.

PubMed

Motaghinejad, Majid; Motevalian, Manijeh; Falak, Reza; Heidari, Mansour; Sharzad, Mahshid; Kalantari, Elham

2016-12-01

Methylphenidate (MPH) abuse damages brain cells. The neuroprotective effects of topiramate (TPM) have been reported previously, but its exact mechanism of action still remains unclear. This study investigated the in vivo role of various doses of TPM in the protection of rat amygdala cells against methylphenidate-induced oxidative stress and inflammation. Seventy adult male rats were divided into seven groups. Groups 1 and 2 received normal saline (0.7 ml/rat) and MPH (10 mg/kg), respectively, for 21 days. Groups 3, 4, 5, 6, and 7 were concurrently treated with MPH (10 mg/kg) and TPM (10, 30, 50, 70, and 100 mg/kg), respectively, for 21 days. elevated plus maze (EPM) was used to assess motor activity disturbances. In addition, oxidative, antioxidantand inflammatory factors and CREB, Ak1, CAMK4, MAPK3, PKA, BDNF, and c FOS gene levels were measured by RT-PCR, and also, CREB and BDNF protein levels were measured by WB in isolated amygdalae. MPH significantly disturbed motor activity and TPM (70 and 100 mg/kg) neutralized its effects. MPH significantly increased lipid peroxidation, mitochondrial GSSG levels and IL-1β and TNF-α level and CAMK4 gene expression in isolated amygdala cells. In contrast, superoxide dismutase, glutathione peroxidase, and glutathione reductase activities and CREB, BDNF Ak1, MAPK3, PKA, BDNF, and c FOS expression significantly decreased. The various doses of TPM attenuated these effects of MPH. It seems that TPM can be used as a neuroprotective agent and is a good candidate against MPH-induced neurodegeneration.

Vitis labruscana leaf extract ameliorates scopolamine-induced impairments with activation of Akt, ERK and CREB in mice.

PubMed

Pariyar, Ramesh; Yoon, Chi-Su; Svay, Thida; Kim, Dae-Sung; Cho, Hyoung-Kwon; Kim, Sung Yeon; Oh, Hyuncheol; Kim, Youn-Chul; Kim, Jaehyo; Lee, Ho-Sub; Seo, Jungwon

2017-12-01

Grapes are among the most widely consumed plants and are used as a folk medicine. Vitis species have been traditionally used as anti-inflammatory, analgesic, and memory-enhancing agents, but, their biological activities of discarded grape leaves are not completely understood. We investigated the effects of alcoholic aqueous leaf extract of Vitis labruscana (LEVL) in a mouse model of memory impairment and tried to ascertain its mechanism. We also evaluated its effects in SH-SY5Y cells. LEVL (50, 100, and 150 mg/kg) was administered to ICR mice once daily for 7 days. Memory impairment was induced with intraperitoneal scopolamine injections (1 mg/kg) and measured with the Y-maze test and a passive avoidance task. LEVL-induced signaling was evaluated in SH-SY5Y cells and mouse hippocampi. We first identified quercetin-3-O-glucuronide as LEVL's major component. We then showed that LEVL promoted phosphorylation of Akt, extracellular regulated kinase (ERK), and cyclic AMP response element binding protein (CREB) and proliferation of SH-SY5Y cells. Oral LEVL administration (100 mg/kg) for 7 days significantly reversed scopolamine-induced reductions of spontaneous alternation in the Y-maze test and scopolamine-induced shortening of latency times in the passive avoidance task's retention trial. Consistent with the cell experiment results, LEVL restored scopolamine-decreased phosphorylation of Akt, ERK, and CREB and scopolamine-reduced expression of brain-derived neuroprotective factor expression in mouse hippocampi. Our results suggest that LEVL promotes phosphorylation of Akt, ERK, and CREB in the hippocampus and ameliorates scopolamine-induced memory impairment in mice. Copyright © 2017 Elsevier GmbH. All rights reserved.

Fasting launches CRTC to facilitate long-term memory formation in Drosophila.

PubMed

Hirano, Yukinori; Masuda, Tomoko; Naganos, Shintaro; Matsuno, Motomi; Ueno, Kohei; Miyashita, Tomoyuki; Horiuchi, Junjiro; Saitoe, Minoru

2013-01-25

Canonical aversive long-term memory (LTM) formation in Drosophila requires multiple spaced trainings, whereas appetitive LTM can be formed after a single training. Appetitive LTM requires fasting prior to training, which increases motivation for food intake. However, we found that fasting facilitated LTM formation in general; aversive LTM formation also occurred after single-cycle training when mild fasting was applied before training. Both fasting-dependent LTM (fLTM) and spaced training-dependent LTM (spLTM) required protein synthesis and cyclic adenosine monophosphate response element-binding protein (CREB) activity. However, spLTM required CREB activity in two neural populations--mushroom body and DAL neurons--whereas fLTM required CREB activity only in mushroom body neurons. fLTM uses the CREB coactivator CRTC, whereas spLTM uses the coactivator CBP. Thus, flies use distinct LTM machinery depending on their hunger state.

Hydrostatic pressure-dependent changes in cyclic AMP signaling in optic nerve head astrocytes from Caucasian and African American donors

PubMed Central

Chen, Lin; Hernandez, M. Rosario

2009-01-01

Purpose Investigate the effect of hydrostatic pressure (HP) on 3′, 5′-cyclic adenosine monophosphate (cAMP) levels and downstream signaling in cultures of normal optic nerve head (ONH) astrocytes from Caucasian American (CA) and African American (AA) donors. Methods Intracellular cAMP levels were assayed after exposing ONH astrocytes to HP for varying times. Quantitative RT–PCR was used to determine the expression levels of selected cAMP pathway genes in human ONH astrocytes after HP treatment. Western blots were used to measure changes in the phosphorylation state of cAMP response element binding protein (CREB) in astrocytes subjected to HP, ATP, and phosphodiesterase or kinase inhibitors. Results The basal intracellular cAMP level is similar among AA and CA astrocytes. After exposure to HP for 15 min and 30 min in the presence of a phosphodiesterase inhibitor a further increase of intracellular cAMP was observed in AA astrocytes, but not in CA astrocytes. Consistent with activation of the cAMP-dependent protein kinase pathway, CREB phosphorylation (Ser-133) was increased to a greater extent in AA than in CA astrocytes after 3 h of HP. Exposure to elevated HP for 3–6 h differentially altered the expression levels of selected cAMP pathway genes (ADCY3, ADCY9, PTHLH, PDE7B) in AA compared to CA astrocytes. Treatment with ATP increased more CREB phosphorylation in CA than in AA astrocytes, suggesting differential Ca2+ signaling in these populations. Conclusions Activation of the cAMP-dependent signaling pathway by pressure may be an important contributor to increased susceptibility to elevated intraocular pressure and glaucoma in AA, a population at higher risk for the disease. PMID:19710943

Inhibitory Effects of Bangladeshi Medicinal Plant Extracts on Interactions between Transcription Factors and Target DNA Sequences

PubMed Central

Lampronti, Ilaria; Khan, Mahmud T.H.; Borgatti, Monica; Bianchi, Nicoletta

2008-01-01

Several transcription factors (TFs) play crucial roles in governing the expression of different genes involved in the immune response, embryo or cell lineage development, cell apoptosis, cell cycle progression, oncogenesis, repair and fibrosis processes and inflammation. As far as inflammation, TFs playing pivotal roles are nuclear factor kappa B (NF-kB), activator protein (AP-1), signal transducer and activator of transcription (STATs), cAMP response element binding protein (CREB) and GATA-1 factors. All these TFs regulate the expression of pro-inflammatory cytokines and are involved in the pathogenesis of a number of human disorders, particularly those with an inflammatory component. Since several medicinal plants can be employed to produce extracts exhibiting biological effects and because alteration of gene transcription represents a very interesting approach to control the expression of selected genes, this study sought to verify the ability of several extracts derived from Bangladeshi medicinal plants in interfering with molecular interactions between different TFs and specific DNA sequences. We first analyzed the antiproliferative activity of 19 medicinal plants on different human cell lines, including erythroleukemia K562, B lymphoid Raji and T lymphoid Jurkat cell lines. Secondly, we employed the electrophoretic mobility shift assay as a suitable technique for a fast screening of plant extracts altering the binding between NF-kB, AP-1, GATA-1, STAT-3, CREB and the relative target DNA elements. PMID:18830455

DNA cytosine methylation in the bovine leukemia virus promoter is associated with latency in a lymphoma-derived B-cell line: potential involvement of direct inhibition of cAMP-responsive element (CRE)-binding protein/CRE modulator/activation transcription factor binding.

PubMed

Pierard, Valérie; Guiguen, Allan; Colin, Laurence; Wijmeersch, Gaëlle; Vanhulle, Caroline; Van Driessche, Benoît; Dekoninck, Ann; Blazkova, Jana; Cardona, Christelle; Merimi, Makram; Vierendeel, Valérie; Calomme, Claire; Nguyên, Thi Liên-Anh; Nuttinck, Michèle; Twizere, Jean-Claude; Kettmann, Richard; Portetelle, Daniel; Burny, Arsène; Hirsch, Ivan; Rohr, Olivier; Van Lint, Carine

2010-06-18

Bovine leukemia virus (BLV) proviral latency represents a viral strategy to escape the host immune system and allow tumor development. Besides the previously demonstrated role of histone deacetylation in the epigenetic repression of BLV expression, we showed here that BLV promoter activity was induced by several DNA methylation inhibitors (such as 5-aza-2'-deoxycytidine) and that overexpressed DNMT1 and DNMT3A, but not DNMT3B, down-regulated BLV promoter activity. Importantly, cytosine hypermethylation in the 5'-long terminal repeat (LTR) U3 and R regions was associated with true latency in the lymphoma-derived B-cell line L267 but not with defective latency in YR2 cells. Moreover, the virus-encoded transactivator Tax(BLV) decreased DNA methyltransferase expression levels, which could explain the lower level of cytosine methylation observed in the L267(LTaxSN) 5'-LTR compared with the L267 5'-LTR. Interestingly, DNA methylation inhibitors and Tax(BLV) synergistically activated BLV promoter transcriptional activity in a cAMP-responsive element (CRE)-dependent manner. Mechanistically, methylation at the -154 or -129 CpG position (relative to the transcription start site) impaired in vitro binding of CRE-binding protein (CREB) transcription factors to their respective CRE sites. Methylation at -129 CpG alone was sufficient to decrease BLV promoter-driven reporter gene expression by 2-fold. We demonstrated in vivo the recruitment of CREB/CRE modulator (CREM) and to a lesser extent activating transcription factor-1 (ATF-1) to the hypomethylated CRE region of the YR2 5'-LTR, whereas we detected no CREB/CREM/ATF recruitment to the hypermethylated corresponding region in the L267 cells. Altogether, these findings suggest that site-specific DNA methylation of the BLV promoter represses viral transcription by directly inhibiting transcription factor binding, thereby contributing to true proviral latency.

PI3K Activation in Neural Stem Cells Drives Tumorigenesis which can be Ameliorated by Targeting the cAMP Response Element Binding (CREB) Protein.

PubMed

Daniel, Paul M; Filiz, Gulay; Brown, Daniel V; Christie, Michael; Waring, Paul M; Zhang, Yi; Haynes, John M; Pouton, Colin; Flanagan, Dustin; Vincan, Elizabeth; Johns, Terrance G; Montgomery, Karen; Phillips, Wayne A; Mantamadiotis, Theo

2018-04-30

Hyperactivation of PI3K signaling is common in cancers but the precise role of the pathway in glioma biology remains to be determined. Some understanding of PI3K signaling mechanisms in brain cancer comes from studies on neural stem/progenitor cells, where signals transmitted via the PI3K pathway cooperate with other intracellular pathways and downstream transcription factors to regulate critical cell functions. To investigate the role for the PI3K pathway in glioma initiation and development, we generated a mouse model targeting the inducible expression of a PIK3CAH1047A oncogenic mutant and deletion of the PI3K negative regulator, PTEN, to neural stem/progenitor cells (NSPCs). Expression of a Pik3caH1047A was sufficient to generate tumors with oligodendroglial features but simultaneous loss of PTEN was required for the development of invasive, high-grade glioma. Pik3caH1047A-PTEN mutant NSPCs exhibited enhanced neurosphere formation which correlated with increased WNT signaling, while loss of CREB in Pik3caH1047A-Pten mutant tumors led to longer symptom-free survival in mice. Taken together, our findings present a novel mouse model for glioma demonstrating that the PI3K pathway is important for initiation of tumorigenesis and that disruption of downstream CREB signaling attenuates tumor expansion.

MicroRNA-155 Is Required for Mycobacterium bovis BCG-Mediated Apoptosis of Macrophages

PubMed Central

Ghorpade, Devram Sampat; Leyland, Rebecca; Kurowska-Stolarska, Mariola; Patil, Shripad A.

2012-01-01

Pathogenic mycobacteria, including Mycobacterium tuberculosis and Mycobacterium bovis, cause significant morbidity and mortality worldwide. However, the vaccine strain Mycobacterium bovis BCG, unlike virulent strains, triggers extensive apoptosis of infected macrophages, a step necessary for the elicitation of robust protective immunity. We here demonstrate that M. bovis BCG triggers Toll-like receptor 2 (TLR2)-dependent microRNA-155 (miR-155) expression, which involves signaling cross talk among phosphatidylinositol 3-kinase (PI3K), protein kinase Cδ (PKCδ), and mitogen-activated protein kinases (MAPKs) and recruitment of NF-κB and c-ETS to miR-155 promoter. Genetic and signaling perturbations presented the evidence that miR-155 regulates PKA signaling by directly targeting a negative regulator of PKA, protein kinase inhibitor alpha (PKI-α). Enhanced activation of PKA signaling resulted in the generation of PKA C-α; phosphorylation of MSK1, cyclic AMP response element binding protein (CREB), and histone H3; and recruitment of phospho-CREB to the apoptotic gene promoters. The miR-155-triggered activation of caspase-3, BAK1, and cytochrome c translocation involved signaling integration of MAPKs and epigenetic or posttranslational modification of histones or CREB. Importantly, M. bovis BCG infection-induced apoptosis was severely compromised in macrophages derived from miR-155 knockout mice. Gain-of-function and loss-of-function studies validated the requirement of miR-155 for M. bovis BCG's ability to trigger apoptosis. Overall, M. bovis BCG-driven miR-155 dictates cell fate decisions of infected macrophages, strongly implicating a novel role for miR-155 in orchestrating cellular reprogramming during immune responses to mycobacterial infection. PMID:22473996

Estrogen signaling through the G protein-coupled estrogen receptor regulates granulocyte activation in fish.

PubMed

Cabas, Isabel; Rodenas, M Carmen; Abellán, Emilia; Meseguer, José; Mulero, Victoriano; García-Ayala, Alfonsa

2013-11-01

Neutrophils are major participants in innate host responses. It is well known that estrogens have an immune-modulatory role, and some evidence exists that neutrophil physiology can be altered by these molecules. Traditionally, estrogens act via classical nuclear estrogen receptors, but the identification of a G protein-coupled estrogen receptor (GPER), a membrane estrogen receptor that binds estradiol and other estrogens, has opened up the possibility of exploring additional estrogen-mediated effects. However, information on the importance of GPER for immunity, especially, in neutrophils is scant. In this study, we report that gilthead seabream (Sparus aurata L.) acidophilic granulocytes, which are the functional equivalent of mammalian neutrophils, express GPER at both mRNA and protein levels. By using a GPER selective agonist, G1, it was found that GPER activation in vitro slightly reduced the respiratory burst of acidophilic granulocytes and drastically altered the expression profile of several genes encoding major pro- and anti-inflammatory mediators. In addition, GPER signaling in vivo modulated adaptive immunity. Finally, a cAMP analog mimicked the effects of G1 in the induction of the gene coding for PG-endoperoxide synthase 2 and in the induction of CREB phosphorylation, whereas pharmacological inhibition of protein kinase A superinduced PG-endoperoxide synthase 2. Taken together, our results demonstrate for the first time, to our knowledge, that estrogens are able to modulate vertebrate granulocyte functions through a GPER/cAMP/protein kinase A/CREB signaling pathway and could establish therapeutic targets for several immune disorders in which estrogens play a prominent role.

Neuropeptide Trefoil Factor 3 Reverses Depressive-Like Behaviors by Activation of BDNF-ERK-CREB Signaling in Olfactory Bulbectomized Rats.

PubMed

Li, Jiali; Luo, Yixiao; Zhang, Ruoxi; Shi, Haishui; Zhu, Weili; Shi, Jie

2015-11-30

The trefoil factors (TFFs) are a family of three polypeptides, among which TFF1 and TFF3 are widely distributed in the central nervous system. Our previous study indicated that TFF3 was a potential rapid-onset antidepressant as it reversed the depressive-like behaviors induced by acute or chronic mild stress. In order to further identify the antidepressant-like effect of TFF3, we applied an olfactory bulbectomy (OB), a classic animal model of depression, in the present study. To elucidate the mechanism underlying the antidepressant-like activity of TFF3, we tested the role of brain-derived neurotrophic factor (BDNF)-extracellular signal-related kinase (ERK)-cyclic adenosine monophosphate response element binding protein (CREB) signaling in the hippocampus in the process. Chronic systemic administration of TFF3 (0.1 mg/kg, i.p.) for seven days not only produced a significant antidepressant-like efficacy in the OB paradigm, but also restored the expression of BDNF, pERK, and pCREB in the hippocampal CA3. Inhibition of BDNF or extracellular signal-related kinase (ERK) signaling in CA3 blocked the antidepressant-like activity of TFF3 in OB rats. Our findings further confirmed the therapeutic effect of TFF3 against depression and suggested that the normalization of the BDNF-ERK-CREB pathway was involved in the behavioral response of TFF3 for the treatment of depression.

Neuroprotective Effect of Arctigenin via Upregulation of P-CREB in Mouse Primary Neurons and Human SH-SY5Y Neuroblastoma Cells

PubMed Central

Zhang, Nan; Wen, Qingping; Ren, Lu; Liang, Wenbo; Xia, Yang; Zhang, Xiaodan; Zhao, Dan; Sun, Dong; Hu, Yv; Hao, Haiguang; Yan, Yaping; Zhang, Guangxian; Yang, Jingxian; Kang, Tingguo

2013-01-01

Arctigenin (Arc) has been shown to act on scopolamine-induced memory deficit mice and to provide a neuroprotective effect on cultured cortical neurons from glutamate-induced neurodegeneration through mechanisms not completely defined. Here, we investigated the neuroprotective effect of Arc on H89-induced cell damage and its potential mechanisms in mouse cortical neurons and human SH-SY5Y neuroblastoma cells. We found that Arc prevented cell viability loss induced by H89 in human SH-SY5Y cells. Moreover, Arc reduced intracellular beta amyloid (Aβ) production induced by H89 in neurons and human SH-SY5Y cells, and Arc also inhibited the presenilin 1(PS1) protein level in neurons. In addition, neural apoptosis in both types of cells, inhibition of neurite outgrowth in human SH-SY5Y cells and reduction of synaptic marker synaptophysin (SYN) expression in neurons were also observed after H89 exposure. All these effects induced by H89 were markedly reversed by Arc treatment. Arc also significantly attenuated downregulation of the phosphorylation of CREB (p-CREB) induced by H89, which may contribute to the neuroprotective effects of Arc. These results demonstrated that Arc exerted the ability to protect neurons and SH-SY5Y cells against H89-induced cell injury via upregulation of p-CREB. PMID:24025424

Activation of cAMP-dependent signaling pathway induces mouse organic anion transporting polypeptide 2 expression.

PubMed

Chen, Chuan; Cheng, Xingguo; Dieter, Matthew Z; Tanaka, Yuji; Klaassen, Curtis D

2007-04-01

Rodent Oatp2 is a hepatic uptake transporter for such compounds as cardiac glycosides. In the present study, we found that fasting resulted in a 2-fold induction of Oatp2 expression in liver of mice. Because the cAMP-protein kinase A (PKA) signaling pathway is activated during fasting, the role of this pathway in Oatp2 induction during fasting was examined. In Hepa-1c1c7 cells, adenylyl cyclase activator forskolin as well as two cellular membrane-permeable cAMP analogs, dibutyryl cAMP and 8-bromo-cAMP, induced Oatp2 mRNA expression in a time- and dose-dependent manner. These three chemicals induced reporter gene activity in cells transfected with a luciferase reporter gene construct containing a 7.6-kilobase (kb) 5'-flanking region of mouse Oatp2. Transient transfection of cells with 5'-deletion constructs derived from the 7.6-kb Oatp2 promoter reporter gene construct, as well as 7.6-kb constructs in which a consensus cAMP response element (CRE) half-site CGTCA (-1808/-1804 bp) was mutated or deleted, confirms that this CRE site was required for the induction of luciferase activity by forskolin. Luciferase activity driven by the Oatp2 promoter containing this CRE site was induced in cells cotransfected with a plasmid encoding the protein kinase A catalytic subunit. Cotransfection of cells with a plasmid encoding the dominant-negative CRE binding protein (CREB) completely abolished the inducibility of the reporter gene activity by forskolin. In conclusion, induction of Oatp2 expression in liver of fasted mice may be caused by activation of the cAMP-dependent signaling pathway, with the CRE site (-1808/-1804) and CREB being the cis- and trans-acting factors mediating the induction, respectively.

Mechanisms of Gravity-Evoked Neuronal Plasticity

NASA Technical Reports Server (NTRS)

Kalb, Robert

2002-01-01

The grant focuses on a gene we identified called, serum and glucocorticoid- induced kinase (SGK), during a previously funded NASA project. The abundance of SGK messenger RNA (mRNA) and protein is increased in CNS tissues from animals reared in microgravity in comparison with 1G reared animals. In the funded proposal we had three aims: 1) characterize the distribution of SGK mRNA in the developing and adult rat CNS, 2) determine if expression of enzymatically active or inactive forms of SGK in cells influenced cell morphology (neurite growth), and 2) determine if SGK is a CREB kinase - that is, a protein kinase that adds phosphate groups to the transcription factor CREB. Over the past year we have made strong progress in the two most difficult parts of the project, namely specific aims 2 and 3. In specific aim #2 we planned to express a dominant negative or a constitutively active form of SGK in PC12 cells and assay the effects on neurite growth. Several methods are available for examining the effects of a transgene on PC12 neurite growth. Relevant variables include the performance of the assay +/- serum, +/- NGF, substratum for growth, timing between transfection and assay. Over the past 8 months we have customized the assay to enable us to most readily determine the effects of transgene expression on neurite growth. We have also compared the relative utility of transfecting DNA as opposed to protein itself. We are now well positioned to study the effects of SGK on neurite growth. We have also made progress in parallel studies in primary neurons. We have made constructs which will lead to transgene expression in cultures of spinal cord neurons. Co-transfection of a reporter and the SGK constructs can now be performed.

Alterations in phosphorylated cyclic adenosine monophosphate response element of binding protein activity: a pathway for fetal alcohol syndrome-related neurotoxicity.

PubMed

Roberson, Robin; Cameroni, Irene; Toso, Laura; Abebe, Daniel; Bissel, Stephanie; Spong, Catherine Y

2009-02-01

Fetal alcohol syndrome (FAS) is the leading cause of a spectrum of preventable nongenetic learning and behavioral disorders. In adult (FAS) mice, we measured phosphorylated cyclic adenosine monophosphate response element of binding protein (pCREB) staining in hippocampal subregions to evaluate a possible mechanism underlying FAS learning deficits. Pregnant C57BL6/J mice were treated on gestational day 8 with alcohol or control (saline). After learning assessment, the offspring were perfused for immunohistochemistry and brain sections probed using SER 133 pCREB antibody. Relative staining density was assessed using National Institutes of Health Image software. Statistical analysis included analysis of variance with P < .05 considered significant. In all hippocampal subregions, pCREB staining was greater in the control animals than in the alcohol-treated group (P < or = .0001). In utero alcohol exposure decreased pCREB activity in hippocampal subregions of adult mice. The dentate gyrus had the most robust cumulative decrease in pCREB staining, suggesting FAS adult learning deficits may correlate to enhanced dentate gyrus neurodegeneration.

Increases in cAMP, MAPK Activity and CREB Phosphorylation during REM Sleep: Implications for REM Sleep and Memory Consolidation

PubMed Central

Luo, Jie; Phan, Trongha X.; Yang, Yimei; Garelick, Michael G.; Storm, Daniel R.

2013-01-01

The cyclic adenosine monophosphate (cAMP), mitogen-activated protein kinase (MAPK) and cAMP response element-binding protein (CREB) transcriptional pathway is required for consolidation of hippocampus-dependent memory. In mice, this pathway undergoes a circadian oscillation required for memory persistence that reaches a peak during the daytime. Since mice exhibit polyphasic sleep patterns during the day, this suggested the interesting possibility that cAMP, MAPK activity and CREB phosphorylation may be elevated during sleep. Here, we report that cAMP, phospho-p44/42 MAPK and phospho-CREB are higher in rapid eye movement (REM) sleep compared to awake mice but are not elevated in non-rapid eye movement (NREM) sleep. This peak of activity during REM sleep does not occur in mice lacking calmodulin-stimulated adenylyl cyclases, a mouse strain that learns but cannot consolidate hippocampus-dependent memory. We conclude that a preferential increase in cAMP, MAPK activity and CREB phosphorylation during REM sleep may contribute to hippocampus-dependent memory consolidation. PMID:23575844

Fluoxetine increases the activity of the ERK-CREB signal system and alleviates the depressive-like behavior in rats exposed to chronic forced swim stress.

PubMed

Qi, Xiaoli; Lin, Wenjuan; Li, Junfa; Li, Huanhuan; Wang, Weiwen; Wang, Donglin; Sun, Meng

2008-08-01

Our previous research indicates that the extracellular signal-regulated kinase (ERK)-cyclic AMP-responsive-element-binding protein (CREB) signal system may be involved in the molecular mechanism of depression. The present study further investigated the effect of antidepressant fluoxetine on the ERK-CREB signal system and the depressive-like behaviors in rats. Fluoxetine was administrated to either naive rats or stressed rats for 21 days. The results showed that chronic forced swim stress induced depressive-like behaviors and decreased the levels of P-ERK2, P-CREB, ERK1/2 and CREB in hippocampus and prefrontal cortex. Fluoxetine alleviated the depressive-like behaviors and reversed the disruptions of the P-ERK2 and P-CREB in stressed rats. Fluoxetine also exerted mood-elevating effect and increased the levels of the P-ERK2 and P-CREB in naive rats. These results suggest that the ERK-CREB signal system may be the targets of the antidepressant action of fluoxetine and participate in the neuronal mechanism of depression.

CRTC2 is required for β-cell function and proliferation.

PubMed

Eberhard, Chandra E; Fu, Accalia; Reeks, Courtney; Screaton, Robert A

2013-07-01

Previous work in insulinoma cell lines has established that calcineurin plays a critical role in the activation of cAMP-responsive element binding protein (Creb), a key transcription factor required for β-cell function and survival, by dephosphorylating the Creb coactivator Creb-regulated transcription coactivator (Crtc)2 at 2 regulatory sites, Ser171 and Ser275. Here, we report that Crtc2 is essential both for glucose-stimulated insulin secretion and cell survival in the β-cell. Endogenous Crtc2 activation is achieved via increasing glucose levels to the physiological feeding range, indicating that Crtc2 is a sensor that couples ambient glucose concentrations to Creb activity in the β-cell. Immunosuppressant drugs such as cyclosporin A and tacrolimus that target the protein phosphatase calcineurin are commonly administered after organ transplantation. Chronic use is associated with reduced insulin secretion and new onset diabetes, suggestive of pancreatic β-cell dysfunction. Importantly, we show that overexpression of a Crtc2 mutant rendered constitutively active by introduction of nonphosphorylatable alanine residues at Ser171 and Ser275 permits Creb target gene activation under conditions when calcineurin is inhibited. Taken together, these data suggest that promoting Crtc2-Creb activity is required for β-cell function and proliferation and promoting this pathway could ameliorate symptoms of new onset diabetes after transplantation.

C/EBPβ is a transcriptional key regulator of IL-36α in murine macrophages.

PubMed

Nerlich, Andreas; Ruangkiattikul, Nanthapon; Laarmann, Kristin; Janze, Nina; Dittrich-Breiholz, Oliver; Kracht, Michael; Goethe, Ralph

2015-08-01

Interleukin (IL)-36α - one of the novel members of the IL-1 family of cytokines - is a potent regulator of dendritic and T cells and plays an important role in inflammatory processes like experimental skin inflammation in mice and in mouse models for human psoriasis. Here, we demonstrate that C/EBPβ, a transcription factor required for the selective expression of inflammatory genes, is a key activator of the Il36A gene in murine macrophages. RNAi-mediated suppression of C/EBPβ expression in macrophages (C/EBPβ(low) cells) significantly impaired Il36A gene induction following challenge with LPS. Despite the presence of five predicted C/EBP binding sites, luciferase reporter assays demonstrated that C/EBPβ confers responsiveness to LPS primarily through a half-CRE•C/EBP element in the proximal Il36A promoter. Electrophoretic mobility shift assays showed that C/EBPβ but not CREB proteins interact with this critical half-CRE•C/EBP element. In addition, overexpression of C/EBPβ in C/EBPβ(low) cells enhanced the expression of Il36A whereas CREB-1 had no effect. Finally, chromatin immunoprecipitation confirmed that C/EBPβ but neither CREB-1, ATF-2 nor ATF4 is directly recruited to the proximal promoter region of the Il36A gene. Together, these findings demonstrate an essential role of C/EBPβ in the regulation of the Il36A gene via the proximal half-CRE•C/EBP element in response to inflammatory stimuli. Copyright © 2015 Elsevier B.V. All rights reserved.

Essential role of the cAMP-cAMP response-element binding protein pathway in opiate-induced homeostatic adaptations of locus coeruleus neurons.

PubMed

Cao, Jun-Li; Vialou, Vincent F; Lobo, Mary Kay; Robison, Alfred J; Neve, Rachael L; Cooper, Donald C; Nestler, Eric J; Han, Ming-Hu

2010-09-28

Excessive inhibition of brain neurons in primary or slice cultures can induce homeostatic intrinsic plasticity, but the functional role and underlying molecular mechanisms of such plasticity are poorly understood. Here, we developed an ex vivo locus coeruleus (LC) slice culture system and successfully recapitulated the opiate-induced homeostatic adaptation in electrical activity of LC neurons seen in vivo. We investigated the mechanisms underlying this adaptation in LC slice cultures by use of viral-mediated gene transfer and genetic mutant mice. We found that short-term morphine treatment of slice cultures almost completely abolished the firing of LC neurons, whereas chronic morphine treatment increased LC neuronal excitability as revealed during withdrawal. This increased excitability was mediated by direct activation of opioid receptors and up-regulation of the cAMP pathway and accompanied by increased cAMP response-element binding protein (CREB) activity. Overexpression of a dominant negative CREB mutant blocked the increase in LC excitability induced by morphine- or cAMP-pathway activation. Knockdown of CREB in slice cultures from floxed CREB mice similarly decreased LC excitability. Furthermore, the ability of morphine or CREB overexpression to up-regulate LC firing was blocked by knockout of the CREB target adenylyl cyclase 8. Together, these findings provide direct evidence that prolonged exposure to morphine induces homeostatic plasticity intrinsic to LC neurons, involving up-regulation of the cAMP-CREB signaling pathway, which then enhances LC neuronal excitability.

Impact of embryo number and maternal undernutrition around the time of conception on insulin signaling and gluconeogenic factors and microRNAs in the liver of fetal sheep.

PubMed

Lie, Shervi; Morrison, Janna L; Williams-Wyss, Olivia; Suter, Catherine M; Humphreys, David T; Ozanne, Susan E; Zhang, Song; MacLaughlin, Severence M; Kleemann, David O; Walker, Simon K; Roberts, Claire T; McMillen, I Caroline

2014-05-01

This study aimed to determine whether exposure of the oocyte and/or embryo to maternal undernutrition results in the later programming of insulin action in the liver and factors regulating gluconeogenesis. To do this, we collect livers from singleton and twin fetal sheep that were exposed to periconceptional (PCUN; -60 to 7 days) or preimplantation (PIUN; 0-7 days) undernutrition at 136-138 days of gestation (term = 150 days). The mRNA and protein abundance of insulin signaling and gluconeogenic factors were then quantified using qRT-PCR and Western blotting, respectively, and global microRNA expression was quantified using deep sequencing methodology. We found that hepatic PEPCK-C mRNA (P < 0.01) and protein abundance and the protein abundance of IRS-1 (P < 0.01), p110β (P < 0.05), PTEN (P < 0.05), CREB (P < 0.01), and pCREB (Ser(133); P < 0.05) were decreased in the PCUN and PIUN singletons. In contrast, hepatic protein abundance of IRS-1 (P < 0.01), p85 (P < 0.01), p110β (P < 0.001), PTEN (P < 0.01), Akt2 (P < 0.01), p-Akt (Ser(473); P < 0.01), and p-FOXO-1 (Thr24) (P < 0.01) was increased in twins. There was a decrease in PEPCK-C mRNA (P < 0.01) but, paradoxically, an increase in PEPCK-C protein (P < 0.001) in twins. Both PCUN and PIUN altered the hepatic expression of 23 specific microRNAs. We propose that the differential impact of maternal undernutrition in the presence of one or two embryos on mRNAs and proteins involved in the insulin signaling and gluconeogenesis is explained by changes in the expression of a suite of specific candidate microRNAs.

Impact of embryo number and maternal undernutrition around the time of conception on insulin signaling and gluconeogenic factors and microRNAs in the liver of fetal sheep

PubMed Central

Lie, Shervi; Morrison, Janna L.; Williams-Wyss, Olivia; Suter, Catherine M.; Humphreys, David T.; Ozanne, Susan E.; Zhang, Song; MacLaughlin, Severence M.; Kleemann, David O.; Walker, Simon K.; Roberts, Claire T.

2014-01-01

This study aimed to determine whether exposure of the oocyte and/or embryo to maternal undernutrition results in the later programming of insulin action in the liver and factors regulating gluconeogenesis. To do this, we collect livers from singleton and twin fetal sheep that were exposed to periconceptional (PCUN; −60 to 7 days) or preimplantation (PIUN; 0–7 days) undernutrition at 136–138 days of gestation (term = 150 days). The mRNA and protein abundance of insulin signaling and gluconeogenic factors were then quantified using qRT-PCR and Western blotting, respectively, and global microRNA expression was quantified using deep sequencing methodology. We found that hepatic PEPCK-C mRNA (P < 0.01) and protein abundance and the protein abundance of IRS-1 (P < 0.01), p110β (P < 0.05), PTEN (P < 0.05), CREB (P < 0.01), and pCREB (Ser133; P < 0.05) were decreased in the PCUN and PIUN singletons. In contrast, hepatic protein abundance of IRS-1 (P < 0.01), p85 (P < 0.01), p110β (P < 0.001), PTEN (P < 0.01), Akt2 (P < 0.01), p-Akt (Ser473; P < 0.01), and p-FOXO-1 (Thr24) (P < 0.01) was increased in twins. There was a decrease in PEPCK-C mRNA (P < 0.01) but, paradoxically, an increase in PEPCK-C protein (P < 0.001) in twins. Both PCUN and PIUN altered the hepatic expression of 23 specific microRNAs. We propose that the differential impact of maternal undernutrition in the presence of one or two embryos on mRNAs and proteins involved in the insulin signaling and gluconeogenesis is explained by changes in the expression of a suite of specific candidate microRNAs. PMID:24496309

Single nucleotide polymorphism near CREB1, rs7591784, is associated with pretreatment methamphetamine use frequency and outcome of outpatient treatment for methamphetamine use disorder

PubMed Central

Heinzerling, Keith G.; Demirdjian, Levon; Wu, Yingnian; Shoptaw, Steven

2016-01-01

Although stimulant dependence is highly heritable, few studies have examined genetic influences on methamphetamine dependence. We performed a candidate gene study of 52 SNPs and pretreatment methamphetamine use frequency among 263 methamphetamine dependent Hispanic and Non-Hispanic White participants of several methamphetamine outpatient clinical trials in Los Angeles. One SNP, rs7591784 was significantly associated with pretreatment methamphetamine use frequency following Bonferroni correction (p < 0.001) in males but not females. We then examined rs7591784 and methamphetamine urine drug screen results during 12 weeks of outpatient treatment among males with treatment outcome data available (N = 94) and found rs7591784 was significantly associated with methamphetamine use during treatment controlling for pretreatment methamphetamine use. rs7591784 is near CREB1 and in a linkage disequilibrium block with rs2952768, previously shown to influence CREB1 expression. The CREB signaling pathway is involved in gene expression changes related to chronic use of multiple drugs of abuse including methamphetamine and these results suggest that variability in CREB signaling may influence pretreatment frequency of methamphetamine use as well as outcomes of outpatient treatment. Medications targeting the CREB pathway, including phosphodiesterase inhibitors, warrant investigation as pharmacotherapies for methamphetamine use disorders. PMID:26736037

Lethal cutaneous disease in transgenic mice conditionally expressing type I human T cell leukemia virus Tax.

PubMed

Kwon, Hakju; Ogle, Louise; Benitez, Bobby; Bohuslav, Jan; Montano, Mauricio; Felsher, Dean W; Greene, Warner C

2005-10-21

Type I human T cell leukemia virus (HTLV-I) is etiologically linked with adult T cell leukemia, an aggressive and usually fatal expansion of activated CD4+ T lymphocytes that frequently traffic to skin. T cell transformation induced by HTLV-I involves the action of the 40-kDa viral Tax transactivator protein. Tax both stimulates the HTLV-I long terminal repeat and deregulates the expression of select cellular genes by altering the activity of specific host transcription factors, including cyclic AMP-responsive element-binding protein (CREB)/activating transcription factor, NF-kappaB/Rel, and serum response factor. To study initiating events involved in HTLV-I Tax-induced T cell transformation, we generated "Tet-off" transgenic mice conditionally expressing in a lymphocyte-restricted manner (EmuSR alpha promoter-enhancer) either wild-type Tax or mutant forms of Tax that selectively compromise the NF-kappaB (M22) or CREB/activating transcription factor (M47) activation pathways. Wild-type Tax and M47 Tax-expressing mice, but not M22-Tax expressing mice, developed progressive alopecia, hyperkeratosis, and skin lesions containing profuse activated CD4 T cell infiltrates with evidence of deregulated inflammatory cytokine production. In addition, these animals displayed systemic lymphadenopathy and splenomegaly. These findings suggest that Tax-mediated activation of NF-kappaB plays a key role in the development of this aggressive skin disease that shares several features in common with the skin disease occurring during the preleukemic stage in HTLV-I-infected patients. Of note, this skin disease completely resolved when Tax transgene expression was suppressed by administration of doxycycline, emphasizing the key role played by this viral oncoprotein in the observed pathology.

Different requirements for cAMP response element binding protein in positive and negative reinforcing properties of drugs of abuse.

PubMed

Walters, C L; Blendy, J A

2001-12-01

Addiction is a complex process that relies on the ability of an organism to integrate positive and negative properties of drugs of abuse. Therefore, studying the reinforcing as well as aversive components of drugs of abuse in a single model system will enable us to understand the role of final common mediators, such as cAMP response element-binding protein (CREB), in the addiction process. To this end, we analyzed mice with a mutation in the alpha and Delta isoforms of the CREB gene. Previously we have shown that CREB(alphaDelta) mutant mice in a mixed genetic background show attenuated signs of physical dependence, as measured by the classic signs of withdrawal. We have generated a uniform genetically stable F1 hybrid (129SvEv/C57BL/6) mouse line harboring the CREB mutation. We have found the functional activity of CREB in these F1 hybrid mice to be dramatically reduced compared with their wild-type littermates. These mice maintain a reduced withdrawal phenotype after chronic morphine. We are now poised to examine a number of complex behavioral phenotypes related to addiction in a well defined CREB-deficient mouse model. We demonstrate that the aversive properties of morphine are still present in CREB mutant mice despite a reduction of physical withdrawal. On the other hand, these mice do not respond to the reinforcing properties of morphine in a conditioned place preference paradigm. In contrast, CREB mutant mice demonstrate an enhanced response to the reinforcing properties of cocaine compared with their wild-type controls in both conditioned place preference and sensitization behaviors. These data may provide the first paradigm for differential vulnerability to various drugs of abuse.

CRTC2 Is a Coactivator of GR and Couples GR and CREB in the Regulation of Hepatic Gluconeogenesis.

PubMed

Hill, Micah J; Suzuki, Shigeru; Segars, James H; Kino, Tomoshige

2016-01-01

Glucocorticoid hormones play essential roles in the regulation of gluconeogenesis in the liver, an adaptive response that is required for the maintenance of circulating glucose levels during fasting. Glucocorticoids do this by cooperating with glucagon, which is secreted from pancreatic islets to activate the cAMP-signaling pathway in hepatocytes. The cAMP-response element-binding protein (CREB)-regulated transcription coactivator 2 (CRTC2) is a coactivator known to be specific to CREB and plays a central role in the glucagon-mediated activation of gluconeogenesis in the early phase of fasting. We show here that CRTC2 also functions as a coactivator for the glucocorticoid receptor (GR). CRTC2 strongly enhances GR-induced transcriptional activity of glucocorticoid-responsive genes. CRTC2 physically interacts with the ligand-binding domain of the GR through a region spanning amino acids 561-693. Further, CRTC2 is required for the glucocorticoid-associated cooperative mRNA expression of the glucose-6-phosphatase, a rate-limiting enzyme for hepatic gluconeogenesis, by facilitating the attraction of GR and itself to its promoter region already occupied by CREB. CRTC2 is required for the maintenance of blood glucose levels during fasting in mice by enhancing the GR transcriptional activity on both the G6p and phosphoenolpyruvate carboxykinase (Pepck) genes. Finally, CRTC2 modulates the transcriptional activity of the progesterone receptor, indicating that it may influence the transcriptional activity of other steroid/nuclear receptors. Taken together, these results reveal that CRTC2 plays an essential role in the regulation of hepatic gluconeogenesis through coordinated regulation of the glucocorticoid/GR- and glucagon/CREB-signaling pathways on the key genes G6P and PEPCK.

Cell-specific occupancy of an extended repertoire of CREM and CREB binding loci in male germ cells

PubMed Central

2010-01-01

Background CREB and CREM are closely related factors that regulate transcription in response to various stress, metabolic and developmental signals. The CREMτ activator isoform is selectively expressed in haploid spermatids and plays an essential role in murine spermiogenesis. Results We have used chromatin immunoprecipitation coupled to sequencing (ChIP-seq) to map CREM and CREB target loci in round spermatids from adult mouse testis and spermatogonia derived GC1-spg cells respectively. We identify more than 9000 genomic loci most of which are cell-specifically occupied. Despite the fact that round spermatids correspond to a highly specialised differentiated state, our results show that they have a remarkably accessible chromatin environment as CREM occupies more than 6700 target loci corresponding not only to the promoters of genes selectively expressed in spermiogenesis, but also of genes involved in functions specific to other cell types. The expression of only a small subset of these target genes are affected in the round spermatids of CREM knockout animals. We also identify a set of intergenic binding loci some of which are associated with H3K4 trimethylation and elongating RNA polymerase II suggesting the existence of novel CREB and CREM regulated transcripts. Conclusions We demonstrate that CREM and CREB occupy a large number of promoters in highly cell specific manner. This is the first study of CREM target promoters directly in a physiologically relevant tissue in vivo and represents the most comprehensive experimental analysis of CREB/CREM regulatory potential to date. PMID:20920259

Inhibition of JNK by pi class of glutathione S-transferase through PKA/CREB pathway is associated with carnosic acid protection against 6-hydroxydopamine-induced apoptosis.

PubMed

Lin, Chia-Yuan; Fu, Ru-Huei; Chou, Ruey-Hwang; Chen, Jing-Hsien; Wu, Chi-Rei; Chang, Shu-Wei; Tsai, Chia-Wen

2017-05-01

Pi class of glutathione S-transferase (GST) is known to suppress c-Jun N-terminal kinase (JNK)-related apoptosis through protein-protein interactions. Moreover, signaling by PKA/cAMP response element binding protein (CREB) is necessary for GSTP up-regulation. This study explored whether carnosic acid (CA) from rosemary prevents 6-hydroxydopamine (6-OHDA)-induced neurotoxicity by inhibition of JNK through GSTP via PKA/CREB signaling. Results indicated that the GSTP protein was increased in SH-SY5Y cells treated with CA for 18 and 24 h. However, CA had no significant effect on alpha or mu class of GST. Treatment of CA increased the induction of p-PKAα, nuclear p-CREB, and CRE-DNA binding activity. These effects of CA were attenuated in cells pretreated with the PKA inhibitor H89. CA pretreatment suppressed 6-OHDA-induced apoptosis by inhibition of JNK phosphorylation, poly(ADP)-ribose polymerase cleavage, and nuclear condensation. Pretreatment with H89 and GSTP siRNA attenuated the ability of CA to reverse 6-OHDA-induced apoptosis. By use of immunoprecipitation with JNK antibody to examine the interaction of GSTP-JNK with CA, we showed that CA pretreatment increased the immunoprecipitation of GSTP after 6-OHDA treatment, which suggests that CA promoted the interaction between GSTP and JNK. CA prevents 6-OHDA-induced apoptosis via inhibition of JNK by GSTP through the PKA/CREB pathway. Copyright © 2017 Elsevier Ltd. All rights reserved.

Antidepressant-like effects of water extract of Gastrodia elata Blume on neurotrophic regulation in a chronic social defeat stress model.

PubMed

Lin, Yu-En; Chou, Shao-Ting; Lin, Shih-Hang; Lu, Kuan-Hung; Panyod, Suraphan; Lai, Yi-Syuan; Ho, Chi-Tang; Sheen, Lee-Yan

2018-04-06

Gastrodia elata Blume (GE) is a traditional Chinese medicine commonly used to treat dizziness, epilepsy, paralysis and some emotional symptoms in east Asia. We previously showed that the water extract of Gastrodia elata Blume (WGE) possesses anti-depression like effects in a forced swimming test and chronic mild stress model. The aim of this study was to investigate the antidepressant-like effects of WGE and potential mechanisms related to brain-derived neurotrophic factor (BDNF) regulation in mice exposed to chronic social defeat stress (CSDS) model. Fifty C57BL/6 mice were divided into 5 groups as follows: a control (CTL) group, CSDS group, and 3 WGE groups receiving 250, 500 or 1000mg/kg body weight in the CSDS model. Mice were administered WGE for 24 days by oral gavage, and the social defeat stress paradigm began on day 14, except for the control group. A social interaction test was conducted to evaluate the antidepressant-like effects of WGE. Blood samples were collected to measure serum corticosterone levels, and the brain was dissected to investigate the expression of BDNF-related signaling pathway proteins using western blotting. Oral administration of WGE improved depression-like behaviors and stress-induced elevations of corticosterone. Further, WGE increased the protein expression of BDNF and promoted the hippocampal protein phosphorylation ratio of cAMP response element binding protein (CREB) and protein kinase B (Akt). WGE exerts antidepressant-like effects on mice in a CSDS model, likely through activating of the BDNF/CREB/Akt pathway. Therefore, WGE has potential as a supplement or an adjuvant to prevent or treat clinical depressive disorders. Copyright © 2018 Elsevier B.V. All rights reserved.

CREB Overexpression Ameliorates Age-related Behavioral and Biophysical Deficits

NASA Astrophysics Data System (ADS)

Yu, Xiao-Wen

Age-related cognitive deficits are observed in both humans and animals. Yet, the molecular mechanisms underlying these deficits are not yet fully elucidated. In aged animals, a decrease in intrinsic excitability of pyramidal neurons from the CA1 sub-region of hippocampus is believed to contribute to age-related cognitive impairments, but the molecular mechanism(s) that modulate both these factors has yet to be identified. Increasing activity of the transcription factor cAMP response element-binding protein (CREB) in young adult rodents has been shown to facilitate cognition, and increase intrinsic excitability of their neurons. However, how CREB changes with age, and how that impacts cognition in aged animals, is not clear. Therefore, we first systematically characterized age- and training-related changes in CREB levels in dorsal hippocampus. At a remote time point after undergoing behavioral training, levels of total CREB and activated CREB (phosphorylated at S133, pCREB) were measured in both young and aged rats. We found that pCREB, but not total CREB was significantly reduced in dorsal CA1 of aged rats. Importantly, levels of pCREB were found to be positively correlated with short-term spatial memory in both young and aged rats i.e. higher pCREB in dorsal CA1 was associated with better spatial memory. These findings indicate that an age-related deficit in CREB activity may contribute to the development of age-related cognitive deficits. However, it was still unclear if increasing CREB activity would be sufficient to ameliorate age-related cognitive, and biophysical deficits. To address this question, we virally overexpressed CREB in CA1, where we found the age-related deficit. Young and aged rats received control or CREB virus, and underwent water maze training. While control aged animals exhibited deficits in long-term spatial memory, aged animals with CREB overexpression performed at levels comparable to young animals. Concurrently, aged neurons overexpressing CREB had increased excitability. This indicates that overexpression of CREB was sufficient to rescue both the cognitive deficits, and the biophysical dysfunction normally seen in aged animals. Together, the results from this thesis identify CREB as a new mechanism underlying age-related cognitive deficits. This not only furthers our understanding of how cognitive processes change with age, but also suggests that increasing activity of CREB or its downstream transcription targets may be a novel therapeutic for the treatment of age-related cognitive decline.

Rufinamide, an antiepileptic drug, improves cognition and increases neurogenesis in the aged gerbil hippocampal dentate gyrus via increasing expressions of IGF-1, IGF-1R and p-CREB.

PubMed

Chen, Bai Hui; Ahn, Ji Hyeon; Park, Joon Ha; Song, Minah; Kim, Hyunjung; Lee, Tae-Kyeong; Lee, Jae Chul; Kim, Young-Myeong; Hwang, In Koo; Kim, Dae Won; Lee, Choong-Hyun; Yan, Bing Chun; Kang, Il Jun; Won, Moo-Ho

2018-04-25

Rufinamide is a novel antiepileptic drug and commonly used in the treatment of Lennox-Gastaut syndrome. In the present study, we investigated effects of rufinamide on cognitive function using passive avoidance test and neurogenesis in the hippocampal dentate gyrus using Ki-67 (a marker for cell proliferation), doublecortin (DCX, a marker for neuroblast) and BrdU/NeuN (markers for newly generated mature neurons) immunohistochemistry in aged gerbils. Aged gerbils (24-month old) were treated with 1 mg/kg and 3 mg/kg rufinamide for 4 weeks. Treatment with 3 mg/kg rufinamide, not 1 mg/kg rufinamide, significantly improved cognitive function and increased neurogenesis, showing that proliferating cells (Ki-67-immunoreactive cells), differentiating neuroblasts (DCX-immunoreactive neuroblasts) and mature neurons (BrdU/NeuN-immunoreactive cells) in the aged dentate gyrus compared with those in the control group. When we examined its mechanisms, rufinamide significantly increased immunoreactivities of insulin-like growth factor-1 (IGF-1), its receptor (IGF-1R), and phosphorylated cAMP response element binding protein (p-CREB). However, rufinamide did not show any increase in immunoreactivities of brain-derived neurotrophic factor and its receptor. Therefore, our results indicate that rufinamide can improve cognitive function and increase neurogenesis in the hippocampus of the aged gerbil via increasing expressions of IGF-1, IGF-1R and p-CREB. Copyright © 2018 Elsevier B.V. All rights reserved.

Levo-tetrahydropalmatine inhibits the acquisition of ketamine-induced conditioned place preference by regulating the expression of ERK and CREB phosphorylation in rats.

PubMed

Du, Yan; Du, Li; Cao, Jie; Hölscher, Christian; Feng, Yongming; Su, Hongliang; Wang, Yujin; Yun, Ke-Ming

2017-01-15

Levo-tetrahydropalmatine (l-THP) is an alkaloid purified from the Chinese herbs Corydalis and Stephania and has been used in many traditional Chinese herbal preparations for its sedative, analgesic and hypnotic properties. Previous studies demonstrated that l-THP has antagonistic activity on dopamine receptors; thus, it may have potential therapeutic effects on drug abuse. However, whether l-THP affects ketamine-induced conditioned place preference (CPP) remains unclear. Therefore, the present study was designed to evaluate the effects of l-THP on the rewarding behavior of ketamine through CPP. Results revealed that ketamine (5, 10 and 15mg/kg) induced CPP in rats. Furthermore, Ketamine (10mg/kg) promoted the phosphorylation of extracellular-regulated kinase (ERK) and cAMP responsive element binding protein (CREB) in the hippocampus (Hip) and caudate putamen (CPu), but not in the prefrontal cortex (PFc). l-THP (20mg/kg) co-administered with ketamine during conditioning inhibited the acquisition of ketamine-induced CPP in rats. Furthermore, l-THP (20mg/kg) prevented the enhanced phosphorylation of ERK and CREB in CPu and Hip. These results suggest that l-THP has potential therapeutic effects on ketamine-induced CPP. The underlying molecular mechanism may be related to its inhibitory effect on ERK and CREB phosphorylation in Hip and CPu. The present data supports the potential use of l-THP for the treatment of ketamine addiction. Copyright © 2016 Elsevier B.V. All rights reserved.

Serum brain-derived neurotrophic factor and glucocorticoid receptor levels in lymphocytes as markers of antidepressant response in major depressive patients: a pilot study.

PubMed

Rojas, Paulina Soledad; Fritsch, Rosemarie; Rojas, Romina Andrea; Jara, Pablo; Fiedler, Jenny Lucy

2011-09-30

Depressive patients often have altered cortisol secretion, an effect that likely derives from impaired activity of the glucocorticoid receptor (GR), the main regulator of the hypothalamus-pituitary-adrenal (HPA) axis. Glucocorticoids reduce the levels of brain-derived neurotrophic factor (BDNF), a downstream target of antidepressants. Antidepressants promote the transcriptional activity of cyclic adenosine monophosphate (cAMP) response element binding protein (CREB), a regulator of BDNF expression. To identify potential biomarkers for the onset of antidepressant action in depressive patients, GR and phospho-CREB (pCREB) levels in lymphocytes and serum BDNF levels were repeatedly measured during the course of antidepressant treatment. Thirty-four depressed outpatients (10 male and 24 female) were treated with venlafaxine (75mg/day), and individuals exhibiting a 50% reduction in their baseline 17-Item Hamilton Depression Rating Scale score by the 6th week of treatment were considered responders. Responders showed an early improvement in parallel with a rise in BDNF levels during the first two weeks of treatment. Non-responders showed increased GR levels by the third week and reduced serum BDNF by the sixth week of treatment. In contrast, venlafaxine did not affect levels of pCREB. We conclude that levels of BDNF in serum and GR levels in lymphocytes may represent biomarkers that could be used to predict responses to venlafaxine treatment. Copyright © 2011 Elsevier Ltd. All rights reserved.

Pulmonary microRNA profiles identify involvement of Creb1 and Sec14l3 in bronchial epithelial changes in allergic asthma

PubMed Central

Bartel, Sabine; Schulz, Nikola; Alessandrini, Francesca; Schamberger, Andrea C.; Pagel, Philipp; Theis, Fabian J.; Milger, Katrin; Noessner, Elfriede; Stick, Stephen M.; Kicic, Anthony; Eickelberg, Oliver; Freishtat, Robert J.; Krauss-Etschmann, Susanne

2017-01-01

Asthma is highly prevalent, but current therapies cannot influence the chronic course of the disease. It is thus important to understand underlying early molecular events. In this study, we aimed to use microRNAs (miRNAs) - which are critical regulators of signaling cascades - to identify so far uncharacterized asthma pathogenesis pathways. Therefore, deregulation of miRNAs was assessed in whole lungs from mice with ovalbumin (OVA)-induced allergic airway inflammation (AAI). In silico predicted target genes were confirmed in reporter assays and in house-dust-mite (HDM) induced AAI and primary human bronchial epithelial cells (NHBE) cultured at the air-liquid interface. We identified and validated the transcription factor cAMP-responsive element binding protein (Creb1) and its transcriptional co-activators (Crtc1-3) as targets of miR-17, miR-144, and miR-21. Sec14-like 3 (Sec14l3) - a putative target of Creb1 - was down-regulated in both asthma models and in NHBE cells upon IL13 treatment, while it’s expression correlated with ciliated cell development and decreased along with increasing goblet cell metaplasia. Finally, we propose that Creb1/Crtc1-3 and Sec14l3 could be important for early responses of the bronchial epithelium to Th2-stimuli. This study shows that miRNA profiles can be used to identify novel targets that would be overlooked in mRNA based strategies. PMID:28383034

Pulmonary microRNA profiles identify involvement of Creb1 and Sec14l3 in bronchial epithelial changes in allergic asthma.

PubMed

Bartel, Sabine; Schulz, Nikola; Alessandrini, Francesca; Schamberger, Andrea C; Pagel, Philipp; Theis, Fabian J; Milger, Katrin; Noessner, Elfriede; Stick, Stephen M; Kicic, Anthony; Eickelberg, Oliver; Freishtat, Robert J; Krauss-Etschmann, Susanne

2017-04-06

Asthma is highly prevalent, but current therapies cannot influence the chronic course of the disease. It is thus important to understand underlying early molecular events. In this study, we aimed to use microRNAs (miRNAs) - which are critical regulators of signaling cascades - to identify so far uncharacterized asthma pathogenesis pathways. Therefore, deregulation of miRNAs was assessed in whole lungs from mice with ovalbumin (OVA)-induced allergic airway inflammation (AAI). In silico predicted target genes were confirmed in reporter assays and in house-dust-mite (HDM) induced AAI and primary human bronchial epithelial cells (NHBE) cultured at the air-liquid interface. We identified and validated the transcription factor cAMP-responsive element binding protein (Creb1) and its transcriptional co-activators (Crtc1-3) as targets of miR-17, miR-144, and miR-21. Sec14-like 3 (Sec14l3) - a putative target of Creb1 - was down-regulated in both asthma models and in NHBE cells upon IL13 treatment, while it's expression correlated with ciliated cell development and decreased along with increasing goblet cell metaplasia. Finally, we propose that Creb1/Crtc1-3 and Sec14l3 could be important for early responses of the bronchial epithelium to Th2-stimuli. This study shows that miRNA profiles can be used to identify novel targets that would be overlooked in mRNA based strategies.

Emodin from Polygonum multiflorum ameliorates oxidative toxicity in HT22 cells and deficits in photothrombotic ischemia.

PubMed

Ahn, Sung Min; Kim, Ha Neui; Kim, Yu Ri; Choi, Young Whan; Kim, Cheol Min; Shin, Hwa Kyoung; Choi, Byung Tae

2016-07-21

Polygonum multiflorum Thunb. has been used widely in East Asia in treatment of diseases associated with aging. Emodin, an active component from Polygonum multiflorum Thunb., provides benefits for brain disturbances induced by severe cerebral injury. We investigated the neuroprotective effect of emodin from Polygonum multiflorum Thunb. against glutamate-induced oxidative toxicity and cerebral ischemia. For examination of neuroprotective effects of emodin, cell viability, cytotoxicity, flow cytometry, and Western blot were performed in HT22 cells and infarct volume, behavioral tests and Western blot in a mouse model of photothrombotic ischemic stroke. Pretreatment with emodin resulted in significantly reduced glutamate-induced apoptotic cell death in HT22 cells. However, blocking of phosphatidylinositol-3 kinase (PI3K) activity with LY294002 resulted in significantly inhibited cell survival by emodin. Exposure of glutamate-treated cells to emodin induced an increase in the level of Bcl-2 expression, whereas the expression of Bax and active caspase-3 proteins was significantly reduced. In addition, treatment with emodin resulted in increased phosphorylation of Akt and cAMP response element binding protein (CREB), and expression of mature brain-derived neurotrophic factor (BDNF). This expression by emodin was also significantly inhibited by blocking of PI3K activity. In a photothrombotic ischemic stroke model, treatment with emodin resulted in significantly reduced infarct volume and improved motor function. We confirmed the critical role of the expression levels of Bcl-2/Bax, active caspase-3, phosphorylated (p)Akt, p-CREB, and mature BDNF for potent neuroprotective effects of emodin in cerebral ischemia. These results suggest that emodin may afford a significant neuroprotective effect against glutamate-induced apoptosis through activation of the PI3K/Akt signaling pathway, and subsequently enhance behavioral function in cerebral ischemia. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Antioxidative and Anti-Melanogenic Activities of Bamboo Stems (Phyllostachys nigra variety henosis) via PKA/CREB-Mediated MITF Downregulation in B16F10 Melanoma Cells.

PubMed

Choi, Moon-Hee; Jo, Han-Gyo; Yang, Ji Hye; Ki, Sung Hwan; Shin, Hyun-Jae

2018-01-30

Phyllostachys nigra var. henosis, a domestic bamboo species, has been attracting much attention; its bioactive compounds (especially in the leaf) show antioxidant, anti-inflammatory, and anti-obesity activities. Little information is available on the antioxidative and anti-melanogenetic activities of the bioactive compounds in bamboo stems. The anti-melanogenic and antioxidative activities of the EtOAc fraction (PN3) of a P. nigra stem extract were investigated in a cell-free system and in B16F10 melanoma cells. PN3 consisted of a mixture of flavonoids, such as catechin, chlorogenic acid, caffeic acid, and p -coumaric acid. The antioxidant activity (2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS)), and hydroxyl radical scavenging) was evaluated, as well as the inhibition of reactive oxygen species (ROS) produced by the Fenton reaction. PN3 showed in vitro tyrosinase inhibition activity with the half maximal inbihitory concentration (IC 50 ) values of 240 μg/mL, and in vivo cytotoxic concentration ranges > 100 μg/mL. The protein expression levels and mRNA transcription levels of TYR , TRP-1 , and MITF were decreased in a dose-dependent manner by the treatment with PN3. PN3 interfered with the phosphorylation of intracellular protein kinase A (PKA)/cAMP response element-binding protein (CREB), demonstrating potent anti-melanogenic effects. PN3 could inhibit PKA/CREB and the subsequent degradation of microphthalmia-associated transcription factor (MITF), resulting in the suppression of melanogenic enzymes and melanin production, probably because of the presence of flavonoid compounds. These properties make it a candidate as an additive to whitening cosmetics.

Antioxidative and Anti-Melanogenic Activities of Bamboo Stems (Phyllostachys nigra variety henosis) via PKA/CREB-Mediated MITF Downregulation in B16F10 Melanoma Cells

PubMed Central

Choi, Moon-Hee; Jo, Han-Gyo; Yang, Ji Hye; Ki, Sung Hwan

2018-01-01

Phyllostachys nigra var. henosis, a domestic bamboo species, has been attracting much attention; its bioactive compounds (especially in the leaf) show antioxidant, anti-inflammatory, and anti-obesity activities. Little information is available on the antioxidative and anti-melanogenetic activities of the bioactive compounds in bamboo stems. The anti-melanogenic and antioxidative activities of the EtOAc fraction (PN3) of a P. nigra stem extract were investigated in a cell-free system and in B16F10 melanoma cells. PN3 consisted of a mixture of flavonoids, such as catechin, chlorogenic acid, caffeic acid, and p-coumaric acid. The antioxidant activity (2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS)), and hydroxyl radical scavenging) was evaluated, as well as the inhibition of reactive oxygen species (ROS) produced by the Fenton reaction. PN3 showed in vitro tyrosinase inhibition activity with the half maximal inbihitory concentration (IC50) values of 240 μg/mL, and in vivo cytotoxic concentration ranges > 100 μg/mL. The protein expression levels and mRNA transcription levels of TYR, TRP-1, and MITF were decreased in a dose-dependent manner by the treatment with PN3. PN3 interfered with the phosphorylation of intracellular protein kinase A (PKA)/cAMP response element-binding protein (CREB), demonstrating potent anti-melanogenic effects. PN3 could inhibit PKA/CREB and the subsequent degradation of microphthalmia-associated transcription factor (MITF), resulting in the suppression of melanogenic enzymes and melanin production, probably because of the presence of flavonoid compounds. These properties make it a candidate as an additive to whitening cosmetics. PMID:29385729

Intestinal DMBT1 Expression Is Modulated by Crohn’s Disease-Associated IL23R Variants and by a DMBT1 Variant Which Influences Binding of the Transcription Factors CREB1 and ATF-2

PubMed Central

Le Bras, Emmanuelle; Zimmermann, Eva; Olszak, Torsten; Bedynek, Andrea; Göke, Burkhard; Franke, Andre

2013-01-01

Objectives DMBT is an antibacterial pattern recognition and scavenger receptor. In this study, we analyzed the role of DMBT1 single nucleotide polymorphisms (SNPs) regarding inflammatory bowel disease (IBD) susceptibility and examined their functional impact on transcription factor binding and downstream gene expression. Methods Seven SNPs in the DMBT1 gene region were analyzed in 2073 individuals including 818 Crohn’s disease (CD) patients and 972 healthy controls in two independent case-control panels. Comprehensive epistasis analyses for the known CD susceptibility genes NOD2, IL23R and IL27 were performed. The influence of IL23R variants on DMBT1 expression was analyzed. Functional analysis included siRNA transfection, quantitative PCR, western blot, electrophoretic mobility shift and luciferase assays. Results IL-22 induces DMBT1 protein expression in intestinal epithelial cells dependent on STAT3, ATF-2 and CREB1. IL-22 expression-modulating, CD risk-associated IL23R variants influence DMBT1 expression in CD patients and DMBT1 levels are increased in the inflamed intestinal mucosa of CD patients. Several DMBT1 SNPs were associated with CD susceptibility. SNP rs2981804 was most strongly associated with CD in the combined panel (p = 3.0×10−7, OR 1.42; 95% CI 1.24–1.63). All haplotype groups tested showed highly significant associations with CD (including omnibus P-values as low as 6.1×10−18). The most strongly CD risk-associated, non-coding DMBT1 SNP rs2981804 modifies the DNA binding sites for the transcription factors CREB1 and ATF-2 and the respective genomic region comprising rs2981804 is able to act as a transcriptional regulator in vitro. Intestinal DMBT1 expression is decreased in CD patients carrying the rs2981804 CD risk allele. Conclusion We identified novel associations of DMBT1 variants with CD susceptibility and discovered a novel functional role of rs2981804 in regulating DMBT1 expression. Our data suggest an important role of DMBT1 in CD pathogenesis. PMID:24223725

Intestinal DMBT1 expression is modulated by Crohn's disease-associated IL23R variants and by a DMBT1 variant which influences binding of the transcription factors CREB1 and ATF-2.

PubMed

Diegelmann, Julia; Czamara, Darina; Le Bras, Emmanuelle; Zimmermann, Eva; Olszak, Torsten; Bedynek, Andrea; Göke, Burkhard; Franke, Andre; Glas, Jürgen; Brand, Stephan

2013-01-01

DMBT is an antibacterial pattern recognition and scavenger receptor. In this study, we analyzed the role of DMBT1 single nucleotide polymorphisms (SNPs) regarding inflammatory bowel disease (IBD) susceptibility and examined their functional impact on transcription factor binding and downstream gene expression. Seven SNPs in the DMBT1 gene region were analyzed in 2073 individuals including 818 Crohn's disease (CD) patients and 972 healthy controls in two independent case-control panels. Comprehensive epistasis analyses for the known CD susceptibility genes NOD2, IL23R and IL27 were performed. The influence of IL23R variants on DMBT1 expression was analyzed. Functional analysis included siRNA transfection, quantitative PCR, western blot, electrophoretic mobility shift and luciferase assays. IL-22 induces DMBT1 protein expression in intestinal epithelial cells dependent on STAT3, ATF-2 and CREB1. IL-22 expression-modulating, CD risk-associated IL23R variants influence DMBT1 expression in CD patients and DMBT1 levels are increased in the inflamed intestinal mucosa of CD patients. Several DMBT1 SNPs were associated with CD susceptibility. SNP rs2981804 was most strongly associated with CD in the combined panel (p = 3.0 × 10(-7), OR 1.42; 95% CI 1.24-1.63). All haplotype groups tested showed highly significant associations with CD (including omnibus P-values as low as 6.1 × 10(-18)). The most strongly CD risk-associated, non-coding DMBT1 SNP rs2981804 modifies the DNA binding sites for the transcription factors CREB1 and ATF-2 and the respective genomic region comprising rs2981804 is able to act as a transcriptional regulator in vitro. Intestinal DMBT1 expression is decreased in CD patients carrying the rs2981804 CD risk allele. We identified novel associations of DMBT1 variants with CD susceptibility and discovered a novel functional role of rs2981804 in regulating DMBT1 expression. Our data suggest an important role of DMBT1 in CD pathogenesis.

Combined inhibition of glycolysis and AMPK induces synergistic breast cancer cell killing

PubMed Central

Wu, Yong; Sarkissyan, Marianna; Mcghee, Eva; Lee, Sangkyu

2015-01-01

Targeting glycolysis for cancer treatment has been investigated as a therapeutic method but has not offered a feasible chemotherapeutic strategy. Our aim was to examine whether AMP-activated protein kinase (AMPK), a conditional oncogene, rescues the energetic stress and cytotoxicity induced by 2-deoxyglucose (2-DG), a glycolytic inhibitor, and the related mechanisms. Luciferin/luciferase adenosine triphosphate (ATP) determination, Western analysis, qRT-PCR analyses, MTT growth assay, clonogenic assay, and statistical analysis were performed in this study. 2-DG decreased ATP levels and subsequently activated AMPK, which contribute to intracellular ATP recovery in MCF-7 cells thus exhibiting no apparent cytotoxicity. Compound C, an AMPK inhibitor, further potentiates 2-DG-induced decrease in ATP levels and inhibits their recovery. 2-DG, via AMPK activation, stimulated cAMP response element-binding protein (CREB) phosphorylation and activity and promoted nuclear peroxisome proliferator-activated receptor gamma coactivator-1-beta (PGC-1β) and estrogen-related receptor α (ERRα) protein expression, leading to augmented mitochondrial biogenesis and expression of fatty acid oxidation (FAO) genes including PPARα, MCAD, CPT1C, and ACO. This metabolic adaptation elicited by AMPK counteracts the ATP-depleting and cancer cell-killing effect of 2-DG. However, 2-DG in combination with AMPK antagonists or small interfering RNA caused a dramatic increase in cytotoxicity in MCF-7 but not in MCF-10A cells. Similarly, when combined with inhibition of CREB/PGC-1β/ERRα pathway, 2-DG saliently suppressed mitochondrial biogenesis and the expression of FAO genes, depleted ATP production, and enhanced cytotoxicity in cancer cells. Collectively, the combination of 2-DG and AMPK inhibition synergistically enhanced the cytotoxic potential in breast cancer cells with a relative nontoxicity to normal cells and may offer a promising, safe, and effective breast cancer therapeutic strategy. PMID:25975952

β-Agonists Selectively Modulate Proinflammatory Gene Expression in Skeletal Muscle Cells via Non-Canonical Nuclear Crosstalk Mechanisms

PubMed Central

Kolmus, Krzysztof; Van Troys, Marleen; Van Wesemael, Karlien; Ampe, Christophe; Haegeman, Guy; Tavernier, Jan; Gerlo, Sarah

2014-01-01

The proinflammatory cytokine Tumour Necrosis Factor (TNF)-α is implicated in a variety of skeletal muscle pathologies. Here, we have investigated how in vitro cotreatment of skeletal muscle C2C12 cells with β-agonists modulates the TNF-α-induced inflammatory program. We observed that C2C12 myotubes express functional TNF receptor 1 (TNF-R1) and β2-adrenoreceptors (β2-ARs). TNF-α activated the canonical Nuclear Factor-κB (NF-κB) pathway and Mitogen-Activated Protein Kinases (MAPKs), culminating in potent induction of NF-κB-dependent proinflammatory genes. Cotreatment with the β-agonist isoproterenol potentiated the expression of inflammatory mediators, including Interleukin-6 (IL-6) and several chemokines. The enhanced production of chemotactic factors upon TNF-α/isoproterenol cotreatment was also suggested by the results from migrational analysis. Whereas we could not explain our observations by cytoplasmic crosstalk, we found that TNF-R1-and β2-AR-induced signalling cascades cooperate in the nucleus. Using the IL-6 promoter as a model, we demonstrated that TNF-α/isoproterenol cotreatment provoked phosphorylation of histone H3 at serine 10, concomitant with enhanced promoter accessibility and recruitment of the NF-κB p65 subunit, cAMP-response element-binding protein (CREB), CREB-binding protein (CBP) and RNA polymerase II. In summary, we show that β-agonists potentiate TNF-α action, via nuclear crosstalk, that promotes chromatin relaxation at selected gene promoters. Our data warrant further study into the mode of action of β-agonists and urge for caution in their use as therapeutic agents for muscular disorders. PMID:24603712

AMP kinase promotes glioblastoma bioenergetics and tumour growth.

PubMed

Chhipa, Rishi Raj; Fan, Qiang; Anderson, Jane; Muraleedharan, Ranjithmenon; Huang, Yan; Ciraolo, Georgianne; Chen, Xiaoting; Waclaw, Ronald; Chow, Lionel M; Khuchua, Zaza; Kofron, Matthew; Weirauch, Matthew T; Kendler, Ady; McPherson, Christopher; Ratner, Nancy; Nakano, Ichiro; Dasgupta, Nupur; Komurov, Kakajan; Dasgupta, Biplab

2018-06-18

Stress is integral to tumour evolution, and cancer cell survival depends on stress management. We found that cancer-associated stress chronically activates the bioenergetic sensor AMP kinase (AMPK) and, to survive, tumour cells hijack an AMPK-regulated stress response pathway conserved in normal cells. Analysis of The Cancer Genome Atlas data revealed that AMPK isoforms are highly expressed in the lethal human cancer glioblastoma (GBM). We show that AMPK inhibition reduces viability of patient-derived GBM stem cells (GSCs) and tumours. In stressed (exercised) skeletal muscle, AMPK is activated to cooperate with CREB1 (cAMP response element binding protein-1) and promote glucose metabolism. We demonstrate that oncogenic stress chronically activates AMPK in GSCs that coopt the AMPK-CREB1 pathway to coordinate tumour bioenergetics through the transcription factors HIF1α and GABPA. Finally, we show that adult mice tolerate systemic deletion of AMPK, supporting the use of AMPK pharmacological inhibitors in the treatment of GBM.

Reversible inactivation of interpeduncular nucleus impairs memory consolidation and retrieval but not learning in rats: A behavioral and molecular study.

PubMed

Khatami, Leila; Khodagholi, Fariba; Motamedi, Fereshteh

2018-04-16

The Interpedundular nucleus (IPN) is a small midbrain structure located deeply between the two cerebral peduncles. The strategic placement of this nucleus makes it a possible relay between structures involved in the modulation of hippocampal theta rhythm activity. In this study we aimed to investigate how reversible inactivation of IPN could affect the acquisition, consolidation and retrieval phases of memory in passive avoidance (PA) and Morris water maze (MWM) tasks. To support our data, molecular studies were performed in order to detect possible changes in the expression of proteins related to learning and memory in the hippocampus. To address this issue rats' IPN was reversibly inactivated by microinjection of lidocaine hydrochloride (4%). After the behavioral studies, the phosphorylation of CREB and P70, and c-fos expression levels in the hippocampus were determined using western blotting and immunohistochemistry respectively. Our results in the PA and MWM tasks showed that IPN reversible inactivation could impair immediate post training consolidation and retrieval while it had no effect on the acquisition phase. In addition, there was a deficit in the retention of the MWM working memory. Our data showed the ratio of pCREB/CREB, pP70/P70 and c-fos expression in the hippocampus significantly decreased after IPN reversible inactivation. Collectively, the results show that behaviorally defined changes could be due to what happens molecularly in the hippocampus after IPN reversible inactivation. It is concluded that IPN not only makes part of a network involved in the modulation of hippocampal theta rhythm activity, but also is actively engaged in hippocampal memory formation. Copyright © 2018 Elsevier B.V. All rights reserved.

N-3 polyunsaturated fatty acids and 17β-estradiol injection induce antidepressant-like effects through regulation of serotonergic neurotransmission in ovariectomized rats.

PubMed

Jin, Youri; Park, Yongsoon

2015-09-01

Previous studies have suggested that estrogen and n-3 polyunsaturated fatty acids (PUFAs), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), have antidepressant-like effects. The purpose of the present study was to determine the interaction between n-3 PUFAs and estrogen, and their neurotrophic mechanism in rats after the forced swimming test (FST). Rats were fed a modified American Institute of Nutrition 93G diet with 0%, 1% or 2% EPA+DHA relative to the total energy intake during 12 weeks. At 8 weeks, rats were ovariectomized and injected with either 17β-estradiol-3-benzoate (E2) or corn oil during the last 3 weeks. Both n-3 PUFA supplementation and E2 injection increased climbing and decreased immobility during the FST. Serum serotonin concentration was also increased by both n-3 PUFA and E2. N-3 PUFA and E2 decreased hippocampal expressions of interleukin (IL)-6 and tumor necrosis factor-α, and increased cAMP response element binding protein (CREB), phosphorylated CREB and brain-derived neurotrophic factor (BDNF). N-3 PUFA supplementation decreased hippocampal expression of IL-1β only in rats injected with E2. Both n-3 PUFA supplementation and E2 injection increased estrogen receptor (ER)-α in the hippocampus, but ER-β was increased only by E2 injection. Additionally, there was a significant interaction between n-3 PUFA supplementation and E2 injection on the hippocampal expression of pCREB, suggesting membrane-mediated interaction of n-3 PUFAs and E2. In conclusion, both n-3 PUFA and E2 had antidepressant-like effects by regulating serotonergic neurotransmission through BDNF and inflammatory cytokines. Copyright © 2015 Elsevier Inc. All rights reserved.

Sequestration of cAMP response element-binding proteins by transcription factor decoys causes collateral elaboration of regenerating Aplysia motor neuron axons.

PubMed

Dash, P K; Tian, L M; Moore, A N

1998-07-07

Axonal injury increases intracellular Ca2+ and cAMP and has been shown to induce gene expression, which is thought to be a key event for regeneration. Increases in intracellular Ca2+ and/or cAMP can alter gene expression via activation of a family of transcription factors that bind to and modulate the expression of CRE (Ca2+/cAMP response element) sequence-containing genes. We have used Aplysia motor neurons to examine the role of CRE-binding proteins in axonal regeneration after injury. We report that axonal injury increases the binding of proteins to a CRE sequence-containing probe. In addition, Western blot analysis revealed that the level of ApCREB2, a CRE sequence-binding repressor, was enhanced as a result of axonal injury. The sequestration of CRE-binding proteins by microinjection of CRE sequence-containing plasmids enhanced axon collateral formation (both number and length) as compared with control plasmid injections. These findings show that Ca2+/cAMP-mediated gene expression via CRE-binding transcription factors participates in the regeneration of motor neuron axons.

Structure-Based Design of Highly Selective Inhibitors of the CREB Binding Protein Bromodomain.

PubMed

Denny, R Aldrin; Flick, Andrew C; Coe, Jotham; Langille, Jonathan; Basak, Arindrajit; Liu, Shenping; Stock, Ingrid; Sahasrabudhe, Parag; Bonin, Paul; Hay, Duncan A; Brennan, Paul E; Pletcher, Mathew; Jones, Lyn H; Chekler, Eugene L Piatnitski

2017-07-13

Chemical probes are required for preclinical target validation to interrogate novel biological targets and pathways. Selective inhibitors of the CREB binding protein (CREBBP)/EP300 bromodomains are required to facilitate the elucidation of biology associated with these important epigenetic targets. Medicinal chemistry optimization that paid particular attention to physiochemical properties delivered chemical probes with desirable potency, selectivity, and permeability attributes. An important feature of the optimization process was the successful application of rational structure-based drug design to address bromodomain selectivity issues (particularly against the structurally related BRD4 protein).

Noxa/Mcl-1 Balance Regulates Susceptibility of Cells to Camptothecin-Induced Apoptosis1

PubMed Central

Mei, Yide; Xie, Chongwei; Xie, Wei; Tian, Xu; Li, Mei; Wu, Mian

2007-01-01

Although camptothecin (CPT) has been reported to induce apoptosis in various cancer cells, the molecular details of this regulation remain largely unknown. In this study, we demonstrate that BH3-only protein Noxa is upregulated during CPT-induced apoptosis, which is independent of p53. In addition, we show that phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway is responsible for Noxa's induction. Luciferase assay and cAMP response element binding protein (CREB) knockdown experiments further demonstrate that CREB is involved in the transcriptional upregulation of Noxa. Moreover, blocking Noxa expression using specific small interfering ribonucleic acid (siRNA) significantly reduces the apoptosis in response to CPT, indicating that Noxa is an essential mediator for CPT-induced apoptosis. Interestingly, antiapoptotic Mcl-1 was also upregulated through PI3K/Akt signaling pathway upon CPT treatment. Using immunoprecipitation assay, Noxa was found to interact with Mcl-1 in the presence or absence of CPT. Knockdown of Mcl-1 expression by short hairpin ribonucleic acid (shRNA) was shown to potentiate CPT-induced apoptosis. Consistently, ectopic overexpression of Mcl-1 rescued cells from apoptosis induced by CPT. Cells coexpressing Noxa and Mcl-1 at different ratio correlates well with the extent of apoptosis, suggesting that the balance between Noxa and Mcl-1 may determine the susceptibility of HeLa cells to CPT-induced apoptosis. PMID:17971907

The n-butanolic extract of Opuntia ficus-indica var. saboten enhances long-term memory in the passive avoidance task in mice.

PubMed

Kim, Jong Min; Kim, Dong Hyun; Park, Se Jin; Park, Dong Hyun; Jung, Seo Yun; Kim, Hyoung Ja; Lee, Yong Sup; Jin, Changbae; Ryu, Jong Hoon

2010-08-16

Opuntia ficus-indica var. saboten Makino (Cactaceae) is used to treat burns, edema, dyspepsia, and asthma in traditional medicine. The present study investigated the beneficial effects of the n-butanolic extract of O. ficus-indica var. saboten (BOF) on memory performance in mice and attempts to uncover the mechanisms underlying its action. Memory performance was assessed with the passive avoidance task, and western blotting and immunohistochemistry were used to measure changes in protein expression and cell survival. After the oral administration of BOF for 7 days, the latency time in the passive avoidance task was significantly increased relative to vehicle-treated controls (P<0.05). Western blotting revealed that the expression levels of brain-derived neurotrophic factor (BDNF), phosphorylated cAMP response element binding-protein (pCREB), and phosphorylated extracellular signal-regulated kinase (pERK) 1/2 were significantly increased in hippocampal tissue after 7 days of BOF administration (P<0.05). Doublecortin and 5-bromo-2-deoxyuridine immunostaining also revealed that BOF significantly enhanced the survival of immature neurons, but did not affect neuronal cell proliferation in the subgranular zone of the hippocampal dentate gyrus. These results suggest that the subchronic administration of BOF enhances long-term memory, and that this effect is partially mediated by ERK-CREB-BDNF signaling and the survival of immature neurons. Copyright (c) 2010 Elsevier Inc. All rights reserved.

Two natural products, trans-phytol and (22E)-ergosta-6,9,22-triene-3β,5α,8α-triol, inhibit the biosynthesis of estrogen in human ovarian granulosa cells by aromatase (CYP19)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guo, Jiajia; Yuan, Yun; School of Life Science and Engineering, Southwest University of Science and Technology, Mianyang

2014-08-15

Aromatase is the only enzyme in vertebrates to catalyze the biosynthesis of estrogens. Although inhibitors of aromatase have been developed for the treatment of estrogen-dependent breast cancer, the whole-body inhibition of aromatase causes severe adverse effects. Thus, tissue-selective aromatase inhibitors are important for the treatment of estrogen-dependent cancers. In this study, 63 natural products with diverse structures were examined for their effects on estrogen biosynthesis in human ovarian granulosa-like KGN cells. Two compounds—trans-phytol (SA-20) and (22E)-ergosta-6,9,22-triene-3β,5α,8α-triol (SA-48)—were found to potently inhibit estrogen biosynthesis (IC{sub 50}: 1 μM and 0.5 μM, respectively). Both compounds decreased aromatase mRNA and protein expression levelsmore » in KGN cells, but had no effect on the aromatase catalytic activity in aromatase-overexpressing HEK293A cells and recombinant expressed aromatase. The two compounds decreased the expression of aromatase promoter I.3/II. Neither compound affected intracellular cyclic AMP (cAMP) levels, but they inhibited the phosphorylation or protein expression of cAMP response element-binding protein (CREB). The effects of these two compounds on extracellular regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinases (MAPKs), and AKT/phosphoinositide 3-kinase (PI3K) pathway were examined. Inhibition of p38 MAPK could be the mechanism underpinning the actions of these compounds. Our results suggests that natural products structurally similar to SA-20 and SA-48 may be a new source of tissue-selective aromatase modulators, and that p38 MAPK is important in the basal control of aromatase in ovarian granulosa cells. SA-20 and SA-48 warrant further investigation as new pharmaceutical tools for the prevention and treatment of estrogen-dependent cancers. - Highlights: • Two natural products inhibited estrogen biosynthesis in human ovarian granulosa cells. • They inhibited aromatase transcription without affecting its catalytic activity. • They decreased the transcription or protein expression of CREB. • They inhibited p38 MAPK to exert their inhibitory effects on aromatase expression.« less

Overlapping ETS and CRE Motifs (G/CCGGAAGTGACGTCA) Preferentially Bound by GABPα and CREB Proteins

PubMed Central

Chatterjee, Raghunath; Zhao, Jianfei; He, Ximiao; Shlyakhtenko, Andrey; Mann, Ishminder; Waterfall, Joshua J.; Meltzer, Paul; Sathyanarayana, B. K.; FitzGerald, Peter C.; Vinson, Charles

2012-01-01

Previously, we identified 8-bps long DNA sequences (8-mers) that localize in human proximal promoters and grouped them into known transcription factor binding sites (TFBS). We now examine split 8-mers consisting of two 4-mers separated by 1-bp to 30-bps (X4-N1-30-X4) to identify pairs of TFBS that localize in proximal promoters at a precise distance. These include two overlapping TFBS: the ETS⇔ETS motif (C/GCCGGAAGCGGAA) and the ETS⇔CRE motif (C/GCGGAAGTGACGTCAC). The nucleotides in bold are part of both TFBS. Molecular modeling shows that the ETS⇔CRE motif can be bound simultaneously by both the ETS and the B-ZIP domains without protein-protein clashes. The electrophoretic mobility shift assay (EMSA) shows that the ETS protein GABPα and the B-ZIP protein CREB preferentially bind to the ETS⇔CRE motif only when the two TFBS overlap precisely. In contrast, the ETS domain of ETV5 and CREB interfere with each other for binding the ETS⇔CRE. The 11-mer (CGGAAGTGACG), the conserved part of the ETS⇔CRE motif, occurs 226 times in the human genome and 83% are in known regulatory regions. In vivo GABPα and CREB ChIP-seq peaks identified the ETS⇔CRE as the most enriched motif occurring in promoters of genes involved in mRNA processing, cellular catabolic processes, and stress response, suggesting that a specific class of genes is regulated by this composite motif. PMID:23050235

Appetitive Pavlovian conditioned stimuli increase CREB phosphorylation in the nucleus accumbens.

PubMed

Shiflett, Michael W; Mauna, Jocelyn C; Chipman, Amanda M; Peet, Eloise; Thiels, Edda

2009-10-01

The transcription factor cAMP response element-binding protein (CREB) in the nucleus accumbens (NAc) has been shown to regulate an animal's behavioral responsiveness to emotionally salient stimuli, and an increase in CREB phosphorylation in the NAc has been observed during exposure to rewarding stimuli, such as drugs of abuse. Here we show that CREB phosphorylation increases in the NAc also during exposure to cues that an animal has associated with delivery of natural rewards. Adult male Sprague-Dawley rats (rattus norvegicus) were trained to associate an auditory stimulus with delivery of food pellets, and CREB phosphorylation was examined in the striatum following training. We found that repeated tone-food pairings resulted in an increase in CREB phosphorylation in the NAc but not in the adjacent dorsal striatum or in the NAc 3h after the final training session. We further found that the cue itself, as opposed to the food pellets, the training context, or tone-food pairings, was sufficient to increase CREB phosphorylation in the NAc. These results suggest that the processing of primary rewarding stimuli and of environmental cues that predict them triggers similar accumbal signaling mechanisms.

Extinction of aversive memories associated with morphine withdrawal requires ERK-mediated epigenetic regulation of brain-derived neurotrophic factor transcription in the rat ventromedial prefrontal cortex.

PubMed

Wang, Wei-Sheng; Kang, Shuo; Liu, Wen-Tao; Li, Mu; Liu, Yao; Yu, Chuan; Chen, Jie; Chi, Zhi-Qiang; He, Ling; Liu, Jing-Gen

2012-10-03

Recent evidence suggests that histone deacetylase (HDAC) inhibitors facilitate extinction of rewarding memory of drug taking. However, little is known about the role of chromatin modification in the extinction of aversive memory of drug withdrawal. In this study, we used conditioned place aversion (CPA), a highly sensitive model for measuring aversive memory of drug withdrawal, to investigate the role of epigenetic regulation of brain-derived neurotrophic factor (BDNF) gene expression in extinction of aversive memory. We found that CPA extinction training induced an increase in recruiting cAMP response element-binding protein (CREB) to and acetylation of histone H3 at the promoters of BDNF exon I transcript and increased BDNF mRNA and protein expression in the ventromedial prefrontal cortex (vmPFC) of acute morphine-dependent rats and that such epigenetic regulation of BDNF gene transcription could be facilitated or diminished by intra-vmPFC infusion of HDAC inhibitor trichostatin A or extracellular signal-regulated kinase (ERK) inhibitor U0126 (1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene) before extinction training. Correspondingly, disruption of the epigenetic regulation of BDNF gene transcription with U0126 or suppression of BDNF signaling with Trk receptor antagonist K252a or BDNF scavenger tyrosine kinase receptor B (TrkB)-Fc blocked extinction of CPA behavior. We also found that extinction training-induced activation of ERK and CREB and extinction of CPA behavior could be potentiated or suppressed by intra-vmPFC infusion of d-cycloserine, a NMDA receptor partial agonist or aminophosphonopentanoic acid, a NMDA receptor antagonist. We conclude that extinction of aversive memory of morphine withdrawal requires epigenetic regulation of BDNF gene transcription in the vmPFC through activation of the ERK-CREB signaling pathway perhaps in a NMDA receptor-dependent manner.

Inhibitory Effect of Dried Pomegranate Concentration Powder on Melanogenesis in B16F10 Melanoma Cells; Involvement of p38 and PKA Signaling Pathways

PubMed Central

Kang, Su Jin; Choi, Beom Rak; Lee, Eun Kyoung; Kim, Seung Hee; Yi, Hae Yeon; Park, Hye Rim; Song, Chang Hyun; Lee, Young Joon; Ku, Sae Kwang

2015-01-01

Plants rich in antioxidant substances may be useful for preventing skin aging. Pomegranates, containing flavonoids and other polyphenolic compounds, are widely consumed due to their beneficial properties. We examined the underlying mechanisms of dried pomegranate concentrate powder (PCP) on melanin synthesis in B16F10 melanoma cells. The antioxidant effects of PCP were determined by measuring free radical scavenging capacity and transcript levels of antioxidant enzymes. To explore the inhibitory effects of PCP on melanin synthesis, we measured tyrosinase activity and melanin content in α-melanocyte stimulating hormone (α-MSH)-stimulated B16F10 cells. In addition, the levels of tyrosinase-related protein-1 (TRP-1), TRP-2, tyrosinase, and microphthalmia-associated transcription factor (MITF) expression were determined by Western blotting. Changes in the phosphorylation status of protein kinase A (PKA), cAMP response element-binding protein (CREB), mitogen-activated protein kinases (MAPKs), phosphatidylinositol 3-kinase (PI3K), serine/threonine kinase Akt, and glycogen kinase 3β (GSK3β) were also examined. The free radical scavenging activity of PCP increased in a dose-dependent manner. In PCP-treated B16F10 cells, transcript levels of glutathione peroxidase-1 (GPx-1) were increased compared with α-MSH-stimulated cells. In addition, PCP led to the down-regulation of phospho-p38, phospho-PKA, phospho-CREB, phospho-GSK3β, MITF, and TRP-1 compared with α-MSH-stimulated B16F10 cells. We believe this effect may be associated with PCP activity, which leads to the inhibition of melanin production and tyrosinase activity. These results suggest that PCP decreases tyrosinase activity and melanin production via inactivation of the p38 and PKA signaling pathways, and subsequently decreases phosphorylation of CREB, MITF, and melanogenic enzymes. These observations provided new insights on the molecular mechanisms of the skin-whitening property of PCP. PMID:26473849

A New Family of Nuclear Receptor Coregulators That Integrate Nuclear Receptor Signaling through CREB-Binding Protein

PubMed Central

Mahajan, Muktar A.; Samuels, Herbert H.

2000-01-01

We describe the cloning and characterization of a new family of nuclear receptor coregulators (NRCs) which modulate the function of nuclear hormone receptors in a ligand-dependent manner. NRCs are expressed as alternatively spliced isoforms which may exhibit different intrinsic activities and receptor specificities. The NRCs are organized into several modular structures and contain a single functional LXXLL motif which associates with members of the steroid hormone and thyroid hormone/retinoid receptor subfamilies with high affinity. Human NRC (hNRC) harbors a potent N-terminal activation domain (AD1), which is as active as the herpesvirus VP16 activation domain, and a second activation domain (AD2) which overlaps with the receptor-interacting LXXLL region. The C-terminal region of hNRC appears to function as an inhibitory domain which influences the overall transcriptional activity of the protein. Our results suggest that NRC binds to liganded receptors as a dimer and this association leads to a structural change in NRC resulting in activation. hNRC binds CREB-binding protein (CBP) with high affinity in vivo, suggesting that hNRC may be an important functional component of a CBP complex involved in mediating the transcriptional effects of nuclear hormone receptors. PMID:10866662

Persistent improvement in synaptic and cognitive functions in an Alzheimer mouse model after rolipram treatment

PubMed Central

Gong, Bing; Vitolo, Ottavio V.; Trinchese, Fabrizio; Liu, Shumin; Shelanski, Michael; Arancio, Ottavio

2004-01-01

Evidence suggests that Alzheimer disease (AD) begins as a disorder of synaptic function, caused in part by increased levels of amyloid β-peptide 1–42 (Aβ42). Both synaptic and cognitive deficits are reproduced in mice double transgenic for amyloid precursor protein (AA substitution K670N,M671L) and presenilin-1 (AA substitution M146V). Here we demonstrate that brief treatment with the phosphodiesterase 4 inhibitor rolipram ameliorates deficits in both long-term potentiation (LTP) and contextual learning in the double-transgenic mice. Most importantly, this beneficial effect can be extended beyond the duration of the administration. One course of long-term systemic treatment with rolipram improves LTP and basal synaptic transmission as well as working, reference, and associative memory deficits for at least 2 months after the end of the treatment. This protective effect is possibly due to stabilization of synaptic circuitry via alterations in gene expression by activation of the cAMP-dependent protein kinase (PKA)/cAMP regulatory element–binding protein (CREB) signaling pathway that make the synapses more resistant to the insult inflicted by Aβ. Thus, agents that enhance the cAMP/PKA/CREB pathway have potential for the treatment of AD and other diseases associated with elevated Aβ42 levels. PMID:15578094

Repeated forced swim stress enhances CFA-evoked thermal hyperalgesia and affects the expressions of pCREB and c-Fos in the insular cortex.

PubMed

Imbe, H; Kimura, A; Donishi, T; Kaneoke, Y

2014-02-14

Stress affects brain activity and promotes long-term changes in multiple neural systems. Exposure to stressors causes substantial effects on the perception and response to pain. In several animal models, chronic stress produces lasting hyperalgesia. The insular (IC) and anterior cingulate cortices (ACC) are the regions exhibiting most reliable pain-related activity. And the IC and ACC play an important role in pain modulation via the descending pain modulatory system. In the present study we examined the expression of phospho-cAMP response element-binding protein (pCREB) and c-Fos in the IC and ACC after forced swim stress (FS) and complete Freund's adjuvant (CFA) injection to clarify changes in the cerebral cortices that affect the activity of the descending pain modulatory system in the rats with stress-induced hyperalgesia. FS (day 1, 10min; days 2-3, 20min) induced an increase in the expression of pCREB and c-Fos in the anterior IC (AIC). CFA injection into the hindpaw after the FS shows significantly enhanced thermal hyperalgesia and induced a decrease in the expression of c-Fos in the AIC and the posterior IC (PIC). Quantitative image analysis showed that the numbers of c-Fos-immunoreactive neurons in the left AIC and PIC were significantly lower in the FS+CFA group (L AIC, 95.9±6.8; L PIC, 181.9±23.1) than those in the naive group (L AIC, 151.1±19.3, p<0.05; L PIC, 274.2±37.3, p<0.05). These findings suggest a neuroplastic change in the IC after FS, which may be involved in the enhancement of CFA-induced thermal hyperalgesia through dysfunction of the descending pain modulatory system. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

MOLECULAR CHARACTERIZATION OF HTLV-1 TAX INTERACTION WITH THE KIX DOMAIN OF CBP/p300

PubMed Central

Ramírez, Julita A.; Nyborg, Jennifer K.

2007-01-01

Summary The viral oncoprotein Tax mediates transcriptional activation of human T-cell leukemia virus type 1 (HTLV-1). Both Tax and the cellular transcription factor CREB bind to viral cyclic AMP response elements (vCREs) located in the viral promoter. Tax and serine 133 phosphorylated CREB (pCREB) bound to the HTLV-1 promoter facilitate viral transcription via the recruitment of the large cellular coactivators CBP/p300. While the interaction between the phosphorylated kinase inducible domain (pKID) of pCREB and the KIX domain of CBP/p300 has been well-characterized, the molecular interactions between KIX, full-length Tax, and pCREB have not been examined. In this study we biochemically characterized the interaction between Tax and KIX in a physiologically relevant complex containing pCREB and vCRE DNA. Our data show that Tax and pCREB simultaneously and independently bind two distinct surfaces on the KIX domain: Tax binds KIX at the previously-characterized mixed-lineage leukemia (MLL) protein interaction surface while pCREB binds KIX at the pKID-KIX interface. These results provide evidence for a model in which Tax and pCREB bind distinct surfaces of KIX for effective CBP/p300 recruitment to the HTLV-1 promoter. We also show that MLL competes with Tax for KIX binding, suggesting a novel mechanism of Tax oncogenesis in which normal MLL function is disrupted by Tax. PMID:17707401

Unliganded estrogen receptor α stimulates bone sialoprotein gene expression.

PubMed

Takai, Hideki; Matsumura, Hiroyoshi; Matsui, Sari; Kim, Kyung Mi; Mezawa, Masaru; Nakayama, Yohei; Ogata, Yorimasa

2014-04-10

Estrogen is one of the steroid hormones essential for skeletal development. The estrogen receptor (ER) is a transcription factor and a member of the steroid receptor superfamily. There are two different forms of the ER, usually referred to as α and β, each encoded by a separate gene. Hormone-activated ERs form dimers, since the two forms are coexpressed in many cell types. Bone sialoprotein (BSP) is a tissue-specific acidic glycoprotein that is expressed by differentiated osteoblasts, odontoblasts and cementoblasts during the initial formation of mineralized tissue. To determine the molecular basis of the tissue-specific expression of BSP and its regulation by estrogen and the ER, we have analyzed the effects of β-estradiol and ERα on BSP gene transcription. ERα protein levels were increased after ERα overexpression in ROS17/2.8 cells. While BSP mRNA levels were increased by ERα overexpression, the endogenous and overexpressed BSP mRNA levels were not changed by β-estradiol (10(-8)M, 24 h). Luciferase activities of different sized BSP promoter constructs (pLUC3~6) were increased by ERα overexpression, whereas basal and induced luciferase activities by ERα overexpression were not influenced by β-estradiol. Effects of ERα overexpression were abrogated by 2 bp mutations in either the cAMP response element (CRE) or activator protein 1 (AP1)/glucocorticoid response element (GRE). Gel shift analyses showed that ERα overexpression increased binding to the CRE and AP1/GRE elements. Notably, the CRE-protein complexes were disrupted by ERα, CREB and phospho-CREB antibodies. The AP1/GRE-protein complexes were supershifted by the c-Fos antibody. These studies demonstrate that ERα stimulates BSP gene transcription in a ligand-independent manner by targeting the CRE and AP1/GRE elements in the rat BSP gene promoter. Copyright © 2014 Elsevier B.V. All rights reserved.

Involvement of PI3K/Akt/FoxO3a and PKA/CREB Signaling Pathways in the Protective Effect of Fluoxetine Against Corticosterone-Induced Cytotoxicity in PC12 Cells.

PubMed

Zeng, Bingqing; Li, Yiwen; Niu, Bo; Wang, Xinyi; Cheng, Yufang; Zhou, Zhongzhen; You, Tingting; Liu, Yonggang; Wang, Haitao; Xu, Jiangping

2016-08-01

The selective serotonin reuptake inhibitor fluoxetine is neuroprotective in several brain injury models. It is commonly used to treat major depressive disorder and related conditions, but its mechanism of action remains incompletely understood. Activation of the phosphatidylinositol-3-kinase/protein kinase B/forkhead box O3a (PI3K/Akt/FoxO3a) and protein kinase A/cAMP-response element binding protein (PKA/CREB) signaling pathways has been strongly implicated in the pathogenesis of depression and might be the downstream target of fluoxetine. Here, we used PC12 cells exposed to corticosterone (CORT) to study the neuroprotective effects of fluoxetine and the involvement of the PI3K/Akt/FoxO3a and PKA/CREB signaling pathways. Our results show that CORT reduced PC12 cells viability by 70 %, and that fluoxetine showed a concentration-dependent neuroprotective effect. Neuroprotective effects of fluoxetine were abolished by inhibition of PI3K, Akt, and PKA using LY294002, KRX-0401, and H89, respectively. Treatment of PC12 cells with fluoxetine resulted in increased phosphorylation of Akt, FoxO3a, and CREB. Fluoxetine also dose-dependently rescued the phosphorylation levels of Akt, FoxO3a, and CREB, following administration of CORT (from 99 to 110, 56 to 170, 80 to 170 %, respectively). In addition, inhibition of PKA and PI3K/Akt resulted in decreased levels of p-CREB, p-Akt, and p-FoxO3a in the presence of fluoxetine. Furthermore, fluoxetine reversed CORT-induced upregulation of p53-upregulated modulator of apoptosis (Puma) and Bcl-2-interacting mediator of cell death (Bim) via the PI3K/Akt/FoxO3a signaling pathway. H89 treatment reversed the effect of fluoxetine on the mRNA level of brain-derived neurotrophic factor, which was decreased in the presence of CORT. Our data indicate that fluoxetine elicited neuroprotection toward CORT-induced cell death that involves dual regulation from PI3K/Akt/FoxO3a and PKA/CREB pathways.

Role of A-Kinase anchor protein (AKAP4) in growth and survival of ovarian cancer cells.

PubMed

Kumar, Vikash; Jagadish, Nirmala; Suri, Anil

2017-08-08

Ovarian cancer represents one of the most common malignancies among women with very high mortality rate worldwide. A-kinase anchor protein 4 (AKAP4), a unique cancer testis (CT) antigen has been shown to be associated with various malignant properties of cancer cells. However, its involvement in various molecular pathways in ovarian cancer remains unknown. In present investigation, employing gene silencing approach, we examined the role of AKAP4 in cell cycle, apoptosis and epithelial-mesenchymal transition (EMT). Further, we also investigated the effect of ablation of AKAP4 on tumor growth in SCID mice ovarian cancer xenograft mouse model. Our results showed that ablation of AKAP4 resulted in increased reactive oxygen species (ROS) generation, DNA damage, cell cycle arrest and apoptosis in ovarian cancer cells. AKAP4 knockdown lead to degradation of protien kinase A (PKA) which was rescued by proteosome inhibitor MG-132. ROS quencher N-acetyl cysteine (NAC) treatment rescued cell cycle arrest and resumed cell division. Subsequently, increased expression of pro-apoptotic molecules and decreased expression of pro-survival/anti-apoptotic factors was observed. As a result of AKAP4 depletion, DNA damage response proteins p-γH2AX, p-ATM and p21 were upregulated. Also, knockdown of CREB resulted in similar findings. Further, PKA inhibitor (H89) and oxidative stress resulted in similar phenotype of ovarian cancer cells as observed in AKAP4 ablated cells. Collectively, for the first time our data showed the involvement of AKAP4 in PKA degradation and perturbed signaling through PKA-CREB axis in AKAP4 ablated ovarian cancer cells.

Moringa oleifera Seed Extract Alleviates Scopolamine-Induced Learning and Memory Impairment in Mice.

PubMed

Zhou, Juan; Yang, Wu-Shuang; Suo, Da-Qin; Li, Ying; Peng, Lu; Xu, Lan-Xi; Zeng, Kai-Yue; Ren, Tong; Wang, Ying; Zhou, Yu; Zhao, Yun; Yang, Li-Chao; Jin, Xin

2018-01-01

The extract of Moringa oleifera seeds has been shown to possess various pharmacological properties. In the present study, we assessed the neuropharmacological effects of 70% ethanolic M. oleifera seed extract (MSE) on cognitive impairment caused by scopolamine injection in mice using the passive avoidance and Morris water maze (MWM) tests. MSE (250 or 500 mg/kg) was administered to mice by oral gavage for 7 or 14 days, and cognitive impairment was induced by intraperitoneal injection of scopolamine (4 mg/kg) for 1 or 6 days. Mice that received scopolamine alone showed impaired learning and memory retention and considerably decreased cholinergic system reactivity and neurogenesis in the hippocampus. MSE pretreatment significantly ameliorated scopolamine-induced cognitive impairment and enhanced cholinergic system reactivity and neurogenesis in the hippocampus. Additionally, the protein expressions of phosphorylated Akt, ERK1/2, and CREB in the hippocampus were significantly decreased by scopolamine, but these decreases were reversed by MSE treatment. These results suggest that MSE-induced ameliorative cognitive effects are mediated by enhancement of the cholinergic neurotransmission system and neurogenesis via activation of the Akt, ERK1/2, and CREB signaling pathways. These findings suggest that MSE could be a potent neuropharmacological drug against amnesia, and its mechanism might be modulation of cholinergic activity via the Akt, ERK1/2, and CREB signaling pathways.

Phosphodiesterase 10A inhibition attenuates sleep deprivation-induced deficits in long-term fear memory.

PubMed

Guo, Lengqiu; Guo, Zhuangli; Luo, Xiaoqing; Liang, Rui; Yang, Shui; Ren, Haigang; Wang, Guanghui; Zhen, Xuechu

2016-12-02

Sleep, particularly rapid eye movement (REM) sleep, is implicated in the consolidation of emotional memories. In the present study, we investigated the protective effects of a phosphodiesterase 10A (PDE10A) inhibitor MP-10 on deficits in long-term fear memory induced by REM sleep deprivation (REM-SD). REM-SD caused deficits in long-term fear memory, however, MP-10 administration ameliorated the deleterious effects of REM-SD on long term fear memory. Brain-derived neurotropic factor (BDNF) and phosphorylated cAMP response element-binding protein (pCREB) were altered in specific brain regions associated with learning and memory in REM-SD rats. Accordingly, REM-SD caused a significant decrease of pCREB in hippocampus and striatum and a significant decrease of BDNF in the hippocampus, striatum and amygdala, however, MP-10 reversed the effects of REM-SD in a dose-dependent manner. Our findings suggest that REM-SD disrupts the consolidation of long-term fear memory and that administration of MP-10 protects the REM-SD-induced deficits in fear memory, which may be due to the MP-10-induced expression of BDNF in the hippocampus, striatum and amygdala, and phosphorylation of CREB in the hippocampus and striatum. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

β1-adrenergic receptors activate two distinct signaling pathways in striatal neurons

PubMed Central

Meitzen, John; Luoma, Jessie I.; Stern, Christopher M.; Mermelstein, Paul G.

2010-01-01

Monoamine action in the dorsal striatum and nucleus accumbens plays essential roles in striatal physiology. Although research often focuses on dopamine and its receptors, norepinephrine and adrenergic receptors are also crucial in regulating striatal function. While noradrenergic neurotransmission has been identified in the striatum, little is known regarding the signaling pathways activated by β-adrenergic receptors in this brain region. Using cultured striatal neurons, we characterized a novel signaling pathway by which activation of β1-adrenergic receptors leads to the rapid phosphorylation of cAMP Response Element Binding Protein (CREB), a transcription-factor implicated as a molecular switch underlying long-term changes in brain function. Norepinephrine-mediated CREB phosphorylation requires β1-adrenergic receptor stimulation of a receptor tyrosine kinase, ultimately leading to the activation of a Ras/Raf/MEK/MAPK/MSK signaling pathway. Activation of β1-adrenergic receptors also induces CRE-dependent transcription and increased c-fos expression. In addition, stimulation of β1-adrenergic receptors produces cAMP production, but surprisingly, β1-adrenergic receptor activation of adenylyl cyclase was not functionally linked to rapid CREB phosphorylation. These findings demonstrate that activation of β1-adrenergic receptors on striatal neurons can stimulate two distinct signaling pathways. These adrenergic actions can produce long-term changes in gene expression, as well as rapidly modulate cellular physiology. By elucidating the mechanisms by which norepinephrine and β1-adrenergic receptor activation affects striatal physiology, we provide the means to more fully understand the role of monoamines in modulating striatal function, specifically how norepinephrine and β1-adrenergic receptors may affect striatal physiology. PMID:21143600

Glucagon-Like Peptide-1 Regulates Cholecystokinin Production in β-Cells to Protect From Apoptosis.

PubMed

Linnemann, Amelia K; Neuman, Joshua C; Battiola, Therese J; Wisinski, Jaclyn A; Kimple, Michelle E; Davis, Dawn Belt

2015-07-01

Cholecystokinin (CCK) is a classic gut hormone that is also expressed in the pancreatic islet, where it is highly up-regulated with obesity. Loss of CCK results in increased β-cell apoptosis in obese mice. Similarly, islet α-cells produce increased amounts of another gut peptide, glucagon-like peptide 1 (GLP-1), in response to cytokine and nutrient stimulation. GLP-1 also protects β-cells from apoptosis via cAMP-mediated mechanisms. Therefore, we hypothesized that the activation of islet-derived CCK and GLP-1 may be linked. We show here that both human and mouse islets secrete active GLP-1 as a function of body mass index/obesity. Furthermore, GLP-1 can rapidly stimulate β-cell CCK production and secretion through direct targeting by the cAMP-modulated transcription factor, cAMP response element binding protein (CREB). We find that cAMP-mediated signaling is required for Cck expression, but CCK regulation by cAMP does not require stimulatory levels of glucose or insulin secretion. We also show that CREB directly targets the Cck promoter in islets from obese (Leptin(ob/ob)) mice. Finally, we demonstrate that the ability of GLP-1 to protect β-cells from cytokine-induced apoptosis is partially dependent on CCK receptor signaling. Taken together, our work suggests that in obesity, active GLP-1 produced in the islet stimulates CCK production and secretion in a paracrine manner via cAMP and CREB. This intraislet incretin loop may be one mechanism whereby GLP-1 protects β-cells from apoptosis.

Glucagon-Like Peptide-1 Regulates Cholecystokinin Production in β-Cells to Protect From Apoptosis

PubMed Central

Linnemann, Amelia K.; Neuman, Joshua C.; Battiola, Therese J.; Wisinski, Jaclyn A.; Kimple, Michelle E.

2015-01-01

Cholecystokinin (CCK) is a classic gut hormone that is also expressed in the pancreatic islet, where it is highly up-regulated with obesity. Loss of CCK results in increased β-cell apoptosis in obese mice. Similarly, islet α-cells produce increased amounts of another gut peptide, glucagon-like peptide 1 (GLP-1), in response to cytokine and nutrient stimulation. GLP-1 also protects β-cells from apoptosis via cAMP-mediated mechanisms. Therefore, we hypothesized that the activation of islet-derived CCK and GLP-1 may be linked. We show here that both human and mouse islets secrete active GLP-1 as a function of body mass index/obesity. Furthermore, GLP-1 can rapidly stimulate β-cell CCK production and secretion through direct targeting by the cAMP-modulated transcription factor, cAMP response element binding protein (CREB). We find that cAMP-mediated signaling is required for Cck expression, but CCK regulation by cAMP does not require stimulatory levels of glucose or insulin secretion. We also show that CREB directly targets the Cck promoter in islets from obese (Leptinob/ob) mice. Finally, we demonstrate that the ability of GLP-1 to protect β-cells from cytokine-induced apoptosis is partially dependent on CCK receptor signaling. Taken together, our work suggests that in obesity, active GLP-1 produced in the islet stimulates CCK production and secretion in a paracrine manner via cAMP and CREB. This intraislet incretin loop may be one mechanism whereby GLP-1 protects β-cells from apoptosis. PMID:25984632

Recruitment of CREB1 and Histone Deacetylase 2 (HDAC2) to the Mouse Ltbp-1 Promoter Regulates its Constitutive Expression in a Dioxin Receptor-dependent Manner

PubMed Central

Gomez-Duran, Aurea; Ballestar, Esteban; Carvajal-Gonzalez, Jose M.; Marlowe, Jennifer L.; Puga, Alvaro; Esteller, Manel; Fernandez-Salguero, Pedro M.

2010-01-01

Latent TGFβ-binding protein 1 (LTBP-1) is a key regulator of TGFβ targeting and activation in the extracellular matrix. LTBP-1 is recognized as a major docking molecule to localize, and possibly to activate, TGFβ in the extracellular matrix. Despite this relevant function, the molecular mechanisms regulating Ltbp-1 transcription remain largely unknown. Previous results from our laboratory revealed that mouse embryonic fibroblasts (MEF) lacking dioxin receptor (AhR) had increased Ltbp-1 mRNA expression and elevated TGFβ activity, suggesting that AhR repressed Ltbp-1 transcription. Here, we have cloned the mouse Ltbp-1 gene promoter and analysed its mechanism of transcriptional repression by AhR. Reporter gene assays, AhR over-expression and site-directed mutagenesis showed that basal Ltbp-1 transcription is AhR-dependent. Chromatin immunoprecipitation (ChIP) and RNA interference (RNAi) revealed that AhR regulates Ltbp-1 transcription by a mechanism involving recruitment of co-activators such as CREB1 and co-repressors such as HDAC2 to the Ltbp-1 promoter. In AhR-expressing (AhR+/+) MEF cells, the recruitment of HDAC1, 2 and 4 correlated with decreased K8H4 acetylation and impaired binding of pCREBSer133 to the Ltbp-1 promoter, likely maintaining a constitutive repressed state. AhR−/− MEF cells had the opposite pattern of HDACs and pCREB1Ser133 binding to Ltbp-1 promoter, and therefore, over-expressed Ltbp-1 mRNA. In agreement, siRNA for HDAC2 increased Ltbp-1 expression and K8H4 acetylation in AhR+/+ but not in AhR−/− MEF cells. We suggest that HDAC2 binding keeps Ltbp-1 promoter repressed in AhR+/+ MEF cells, whereas in AhR-null MEF cells the absence of HDAC2 and the binding of pCREBSer133 allow Ltbp-1 transcription. Thus, epigenetics can contribute to constitutive Ltbp-1 repression by a mechanism requiring AhR activity. PMID:18508077

CREB Selectively Controls Learning-Induced Structural Remodeling of Neurons

ERIC Educational Resources Information Center

Middei, Silvia; Spalloni, Alida; Longone, Patrizia; Pittenger, Christopher; O'Mara, Shane M.; Marie, Helene; Ammassari-Teule, Martine

2012-01-01

The modulation of synaptic strength associated with learning is post-synaptically regulated by changes in density and shape of dendritic spines. The transcription factor CREB (cAMP response element binding protein) is required for memory formation and in vitro dendritic spine rearrangements, but its role in learning-induced remodeling of neurons…

Human T-lymphotropic virus type-1 p30 alters cell cycle G2 regulation of T lymphocytes to enhance cell survival

PubMed Central

Datta, Antara; Silverman, Lee; Phipps, Andrew J; Hiraragi, Hajime; Ratner, Lee; Lairmore, Michael D

2007-01-01

Background Human T-lymphotropic virus type-1 (HTLV-1) causes adult T-cell leukemia/lymphoma and is linked to a number of lymphocyte-mediated disorders. HTLV-1 contains both regulatory and accessory genes in four pX open reading frames. pX ORF-II encodes two proteins, p13 and p30, whose roles are still being defined in the virus life cycle and in HTLV-1 virus-host cell interactions. Proviral clones of HTLV-1 with pX ORF-II mutations diminish the ability of the virus to maintain viral loads in vivo. p30 expressed exogenously differentially modulates CREB and Tax-responsive element-mediated transcription through its interaction with CREB-binding protein/p300 and while acting as a repressor of many genes including Tax, in part by blocking tax/rex RNA nuclear export, selectively enhances key gene pathways involved in T-cell signaling/activation. Results Herein, we analyzed the role of p30 in cell cycle regulation. Jurkat T-cells transduced with a p30 expressing lentivirus vector accumulated in the G2-M phase of cell cycle. We then analyzed key proteins involved in G2-M checkpoint activation. p30 expression in Jurkat T-cells resulted in an increase in phosphorylation at serine 216 of nuclear cell division cycle 25C (Cdc25C), had enhanced checkpoint kinase 1 (Chk1) serine 345 phosphorylation, reduced expression of polo-like kinase 1 (PLK1), diminished phosphorylation of PLK1 at tyrosine 210 and reduced phosphorylation of Cdc25C at serine 198. Finally, primary human lymphocyte derived cell lines immortalized by a HTLV-1 proviral clone defective in p30 expression were more susceptible to camptothecin induced apoptosis. Collectively these data are consistent with a cell survival role of p30 against genotoxic insults to HTLV-1 infected lymphocytes. Conclusion Collectively, our data are the first to indicate that HTLV-1 p30 expression results in activation of the G2-M cell cycle checkpoint, events that would promote early viral spread and T-cell survival. PMID:17634129

Corticosteroids inhibit sphingosine 1-phosphate-induced interleukin-6 secretion from human airway smooth muscle via mitogen-activated protein kinase phosphatase 1-mediated repression of mitogen and stress-activated protein kinase 1.

PubMed

Che, Wenchi; Parmentier, Johannes; Seidel, Petra; Manetsch, Melanie; Ramsay, Emma E; Alkhouri, Hatem; Ge, Qi; Armour, Carol L; Ammit, Alaina J

2014-02-01

Sphingosine 1-phosphate (S1P) is a bioactive sphingolipid that plays an important proinflammatory role in asthmatic airways. Corticosteroids are first-line antiinflammatories in asthma; however, their repressive effects on S1P-induced cytokine secretion have not been investigated. To address this, our in vitro study reveals the molecular mechanisms by which corticosteroids inhibit S1P-induced IL-6 expression in the pivotal immunomodulatory cell type, airway smooth muscle (ASM). We first uncover the cellular signaling pathways responsible: S1P activates a cyclic adenosine monophosphate/cAMP response-element-binding protein (CREB)/CRE-dependent pathway to induce IL-6 transcription, concomitant with stimulation of the mitogen-activated protein kinase (MAPK) superfamily and downstream mitogen and stress-activated protein kinase 1 (MSK1) and histone H3 phosphorylation. In this way, S1P stimulates parallel signaling pathways to induce IL-6 secretion via CRE-driven transcription of the IL-6 gene promoter in a relaxed chromatin environment achieved through histone H3 phosphorylation. Second, we investigated how corticosteroids mediate their repressive effects. The corticosteroid dexamethasone inhibits S1P-induced IL-6 protein secretion and mRNA expression, but CREB/CRE transrepression, inhibition of IL-6 mRNA stability, or subcellular relocation of MSK1 were not responsible for the repressive effects of dexamethasone. Rather, we show that dexamethasone rapidly induces up-regulation of the MAPK deactivator MAPK phosphatase 1 (MKP-1) and that MKP-1 blocks the MAPK-driven activation of MSK1 and phosphorylation of histone H3. This was confirmed by treatment with triptolide, an inhibitor of MKP-1 up-regulation, where repressive effects of corticosteroids were reversed. Our study reveals the molecular mechanism underlying the antiinflammatory capacity of corticosteroids to repress proinflammatory functions induced by the potent bioactive sphingolipid S1P in the lung.

Human immunodeficiency virus type 1 enhancer-binding protein 3 is essential for the expression of asparagine-linked glycosylation 2 in the regulation of osteoblast and chondrocyte differentiation.

PubMed

Imamura, Katsuyuki; Maeda, Shingo; Kawamura, Ichiro; Matsuyama, Kanehiro; Shinohara, Naohiro; Yahiro, Yuhei; Nagano, Satoshi; Setoguchi, Takao; Yokouchi, Masahiro; Ishidou, Yasuhiro; Komiya, Setsuro

2014-04-04

Human immunodeficiency virus type 1 enhancer-binding protein 3 (Hivep3) suppresses osteoblast differentiation by inducing proteasomal degradation of the osteogenesis master regulator Runx2. In this study, we tested the possibility of cooperation of Hivep1, Hivep2, and Hivep3 in osteoblast and/or chondrocyte differentiation. Microarray analyses with ST-2 bone stroma cells demonstrated that expression of any known osteochondrogenesis-related genes was not commonly affected by the three Hivep siRNAs. Only Hivep3 siRNA promoted osteoblast differentiation in ST-2 cells, whereas all three siRNAs cooperatively suppressed differentiation in ATDC5 chondrocytes. We further used microarray analysis to identify genes commonly down-regulated in both MC3T3-E1 osteoblasts and ST-2 cells upon knockdown of Hivep3 and identified asparagine-linked glycosylation 2 (Alg2), which encodes a mannosyltransferase residing on the endoplasmic reticulum. The Hivep3 siRNA-mediated promotion of osteoblast differentiation was negated by forced Alg2 expression. Alg2 suppressed osteoblast differentiation and bone formation in cultured calvarial bone. Alg2 was immunoprecipitated with Runx2, whereas the combined transfection of Runx2 and Alg2 interfered with Runx2 nuclear localization, which resulted in suppression of Runx2 activity. Chondrocyte differentiation was promoted by Hivep3 overexpression, in concert with increased expression of Creb3l2, whose gene product is the endoplasmic reticulum stress transducer crucial for chondrogenesis. Alg2 silencing suppressed Creb3l2 expression and chondrogenesis of ATDC5 cells, whereas infection of Alg2-expressing virus promoted chondrocyte maturation in cultured cartilage rudiments. Thus, Alg2, as a downstream mediator of Hivep3, suppresses osteogenesis, whereas it promotes chondrogenesis. To our knowledge, this study is the first to link a mannosyltransferase gene to osteochondrogenesis.

Intra-Amygdala Injections of CREB Antisense Impair Inhibitory Avoidance Memory: Role of Norepinephrine and Acetylcholine

ERIC Educational Resources Information Center

Canal, Clinton E.; Chang, Qing; Gold, Paul E.

2008-01-01

Infusions of CREB antisense into the amygdala prior to training impair memory for aversive tasks, suggesting that the antisense may interfere with CRE-mediated gene transcription and protein synthesis important for the formation of new memories within the amygdala. However, the amygdala also appears to modulate memory formation in distributed…

Methotrexate in Atherogenesis and Cholesterol Metabolism

PubMed Central

Coomes, Eric; Chan, Edwin S. L.; Reiss, Allison B.

2011-01-01

Methotrexate is a disease-modifying antirheumatic drug commonly used to treat inflammatory conditions such as rheumatoid arthritis which itself is linked to increased cardiovascular risk. Treatments that target inflammation may also impact the cardiovascular system. While methotrexate improves cardiovascular risk, inhibition of the cyclooxygenase (COX)-2 enzyme promotes atherosclerosis. These opposing cardiovascular influences may arise from differing effects on the expression of proteins involved in cholesterol homeostasis. These proteins, ATP-binding cassette transporter (ABC) A1 and cholesterol 27-hydroxylase, facilitate cellular cholesterol efflux and defend against cholesterol overload. Methotrexate upregulates expression of cholesterol 27-hydroxylase and ABCA1 via adenosine release, while COX-2 inhibition downregulates these proteins. Adenosine, acting through the A2A and A3 receptors, may upregulate proteins involved in reverse cholesterol transport by cAMP-PKA-CREB activation and STAT inhibition, respectively. Elucidating underlying cardiovascular mechanisms of these drugs provides a framework for developing novel cardioprotective anti-inflammatory medications, such as selective A2A receptor agonists. PMID:21490773

Antisense protein tyrosine phosphatase 1B reverses activation of p38 mitogen-activated protein kinase in liver of ob/ob mice.

PubMed

Gum, Rebecca J; Gaede, Lori L; Heindel, Matthew A; Waring, Jeffrey F; Trevillyan, James M; Zinker, Bradley A; Stark, Margery E; Wilcox, Denise; Jirousek, Michael R; Rondinone, Cristina M; Ulrich, Roger G

2003-06-01

Phosphorylation of stress-activated kinase p38, a MAPK family member, was increased in liver of ob/ob diabetic mice relative to lean littermates. Treatment of ob/ob mice with protein tyrosine phosphatase 1B (PTP1B) antisense oligonucleotides (ASO) reduced phosphorylation of p38 in liver-to below lean littermate levels-and normalized plasma glucose while reducing plasma insulin. Phosphorylation of ERK, but not JNK, was also decreased in ASO-treated mice. PTP1B ASO decreased TNFalpha protein levels and phosphorylation of the transcription factor cAMP response element binding protein (CREB) in liver, both of which can occur through decreased phosphorylation of p38 and both of which have been implicated in insulin resistance or hyperglycemia. Decreased p38 phosphorylation was not directly due to decreased phosphorylation of the kinases that normally phosphorylate p38-MKK3 and MKK6. Additionally, p38 phosphorylation was not enhanced in liver upon insulin stimulation of ASO-treated ob/ob mice (despite increased activation of other signaling molecules) corroborating that p38 is not directly affected via the insulin receptor. Instead, decreased phosphorylation of p38 may be due to increased expression of MAPK phosphatases, particularly the p38/ERK phosphatase PAC1 (phosphatase of activated cells). This study demonstrates that reduction of PTP1B protein using ASO reduces activation of p38 and its substrates TNFalpha and CREB in liver of diabetic mice, which correlates with decreased hyperglycemia and hyperinsulinemia.

Polygalasaponin XXXII from Polygala tenuifolia root improves hippocampal-dependent learning and memory.

PubMed

Xue, Wei; Hu, Jin-feng; Yuan, Yu-he; Sun, Jian-dong; Li, Bo-yu; Zhang, Dong-ming; Li, Chuang-jun; Chen, Nai-hong

2009-09-01

The aim of this study was to investigate the cognition-enhancing activity and underlying mechanisms of a triterpenoid saponin (polygalasaponin XXXII, PGS32) isolated from the roots of Polygala tenuifolia Willd. The Morris water maze was used to evaluate the spatial learning and memory of mice. To detect the basic properties of synaptic transmission and long-term potentiation (LTP) in the dentate gyrus of rats, electrophysiological recordings were made of evoked potentials. Western blotting analysis and immunofluorescence assays were used to determine the phosphorylation of extracellular signal-regulated kinase (ERK), cAMP response element-binding protein (CREB), synapsin I and the expression of brain derived neurotrophic factor (BDNF). When administered at 0.125, 0.5, or 2 mg/kg, PGS32 could significantly prevent scopolamine-induced cognitive impairments in mice. Intracerebroventricular (icv) administration of PGS32 greatly enhanced basic synaptic transmission in the dentate gyrus of rats and induced LTP. In primary hippocampal neurons, as well as in the hippocampus of maze-trained mice, PGS32 activated the mitogen-activated protein (MAP) kinase cascade by promoting phosphorylation of ERK, CREB and synapsin I. The expression of BDNF was also greatly enhanced in the hippocampus. Our findings suggest that PGS32 can improve hippocampus-dependent learning and memory, possibly through improvement of synaptic transmission, activation of the MAP kinase cascade and enhancement of the level of BDNF. Therefore, PGS32 shows promise as a potential cognition-enhancing therapeutic drug.

Polygalasaponin XXXII from Polygala tenuifolia root improves hippocampal-dependent learning and memory

PubMed Central

Xue, Wei; Hu, Jin-feng; Yuan, Yu-he; Sun, Jian-dong; Li, Bo-yu; Zhang, Dong-ming; Li, Chuang-jun; Chen, Nai-hong

2009-01-01

Aim: The aim of this study was to investigate the cognition-enhancing activity and underlying mechanisms of a triterpenoid saponin (polygalasaponin XXXII, PGS32) isolated from the roots of Polygala tenuifolia Willd. Methods: The Morris water maze was used to evaluate the spatial learning and memory of mice. To detect the basic properties of synaptic transmission and long-term potentiation (LTP) in the dentate gyrus of rats, electrophysiological recordings were made of evoked potentials. Western blotting analysis and immunofluorescence assays were used to determine the phosphorylation of extracellular signal-regulated kinase (ERK), cAMP response element-binding protein (CREB), synapsin I and the expression of brain derived neurotrophic factor (BDNF). Results: When administered at 0.125, 0.5, or 2 mg/kg, PGS32 could significantly prevent scopolamine-induced cognitive impairments in mice. Intracerebroventricular (icv) administration of PGS32 greatly enhanced basic synaptic transmission in the dentate gyrus of rats and induced LTP. In primary hippocampal neurons, as well as in the hippocampus of maze-trained mice, PGS32 activated the mitogen-activated protein (MAP) kinase cascade by promoting phosphorylation of ERK, CREB and synapsin I. The expression of BDNF was also greatly enhanced in the hippocampus. Conclusion: Our findings suggest that PGS32 can improve hippocampus-dependent learning and memory, possibly through improvement of synaptic transmission, activation of the MAP kinase cascade and enhancement of the level of BDNF. Therefore, PGS32 shows promise as a potential cognition-enhancing therapeutic drug. PMID:19684611

Hypoglycemia induced behavioural deficit and decreased GABA receptor, CREB expression in the cerebellum of streptozoticin induced diabetic rats.

PubMed

Sherin, A; Peeyush, K T; Naijil, G; Chinthu, R; Paulose, C S

2010-11-20

Intensive glycemic control during diabetes is associated with an increased incidence of hypoglycemia, which is the major barrier in blood glucose homeostasis during diabetes therapy. The CNS neurotransmitters play an important role in the regulation of glucose homeostasis. In the present study, we showed the effects of hypoglycemia in diabetic and non- diabetic rats on motor functions and alterations of GABA receptor and CREB expression in the cerebellum. Cerebellar dysfunction is associated with seizure generation, motor deficits and memory impairment. Scatchard analysis of [(3)H]GABA binding in the cerebellum of diabetic hypoglycemic and control hypoglycemic rats showed significant (P<0.01) decrease in B(max) and K(d) compared to diabetic and control rats. Real-time PCR amplification of GABA receptor subunit GABA(Aα1) and GAD showed significant (P<0.001) down-regulation in the cerebellum of hypoglycemic rats compared to diabetic and control rats. Confocal imaging study confirmed the decreased GABA receptors in hypoglycemic rats. CREB mRNA expression was down-regulated during recurrent hypoglycemia. Both diabetic and non-diabetic hypoglycemic rats showed impaired performance in grid walk test compared to diabetic and control. Impaired GABA receptor and CREB expression along with motor function deficit were more prominent in hypoglycemic rats than hyperglycemic which showed that hypoglycemia is causing more neuronal damage at molecular level. These molecular changes observed during hypo/hyperglycemia contribute to motor and learning deficits which has clinical significance in diabetes treatment. 2010 Elsevier Inc. All rights reserved.

Neuronal Allocation to a Hippocampal Engram

PubMed Central

Park, Sungmo; Kramer, Emily E; Mercaldo, Valentina; Rashid, Asim J; Insel, Nathan; Frankland, Paul W; Josselyn, Sheena A

2016-01-01

The dentate gyrus (DG) is important for encoding contextual memories, but little is known about how a population of DG neurons comes to encode and support a particular memory. One possibility is that recruitment into an engram depends on a neuron's excitability. Here, we manipulated excitability by overexpressing CREB in a random population of DG neurons and examined whether this biased their recruitment to an engram supporting a contextual fear memory. To directly assess whether neurons overexpressing CREB at the time of training became critical components of the engram, we examined memory expression while the activity of these neurons was silenced. Chemogenetically (hM4Di, an inhibitory DREADD receptor) or optogenetically (iC++, a light-activated chloride channel) silencing the small number of CREB-overexpressing DG neurons attenuated memory expression, whereas silencing a similar number of random neurons not overexpressing CREB at the time of training did not. As post-encoding reactivation of the activity patterns present during initial experience is thought to be important in memory consolidation, we investigated whether post-training silencing of neurons allocated to an engram disrupted subsequent memory expression. We found that silencing neurons 5 min (but not 24 h) following training disrupted memory expression. Together these results indicate that the rules of neuronal allocation to an engram originally described in the lateral amygdala are followed in different brain regions including DG, and moreover, that disrupting the post-training activity pattern of these neurons prevents memory consolidation. PMID:27187069

Neuronal Allocation to a Hippocampal Engram.

PubMed

Park, Sungmo; Kramer, Emily E; Mercaldo, Valentina; Rashid, Asim J; Insel, Nathan; Frankland, Paul W; Josselyn, Sheena A

2016-12-01

The dentate gyrus (DG) is important for encoding contextual memories, but little is known about how a population of DG neurons comes to encode and support a particular memory. One possibility is that recruitment into an engram depends on a neuron's excitability. Here, we manipulated excitability by overexpressing CREB in a random population of DG neurons and examined whether this biased their recruitment to an engram supporting a contextual fear memory. To directly assess whether neurons overexpressing CREB at the time of training became critical components of the engram, we examined memory expression while the activity of these neurons was silenced. Chemogenetically (hM4Di, an inhibitory DREADD receptor) or optogenetically (iC++, a light-activated chloride channel) silencing the small number of CREB-overexpressing DG neurons attenuated memory expression, whereas silencing a similar number of random neurons not overexpressing CREB at the time of training did not. As post-encoding reactivation of the activity patterns present during initial experience is thought to be important in memory consolidation, we investigated whether post-training silencing of neurons allocated to an engram disrupted subsequent memory expression. We found that silencing neurons 5 min (but not 24 h) following training disrupted memory expression. Together these results indicate that the rules of neuronal allocation to an engram originally described in the lateral amygdala are followed in different brain regions including DG, and moreover, that disrupting the post-training activity pattern of these neurons prevents memory consolidation.

Sildenafil protects against 3-nitropropionic acid neurotoxicity through the modulation of calpain, CREB, and BDNF.

PubMed

Puerta, Elena; Hervias, Isabel; Barros-Miñones, Lucía; Jordan, Joaquin; Ricobaraza, Ana; Cuadrado-Tejedor, Mar; García-Osta, Ana; Aguirre, Norberto

2010-05-01

In this study we tested whether phosphodiesterase 5 (PDE5) inhibitors, sildenafil and vardenafil, would afford protection against 3-nitropropionic acid (3NP), which produces striatal lesions that closely mimic some of the neuropathological features of Huntington's Disease (HD). The neurotoxin was given over 5 days by constant systemic infusion using osmotic minipumps. Animals treated with PDE5 inhibitors (sildenafil or vardenafil) showed improved neurologic scores, reduced the loss of striatal DARPP-32 protein levels and lesion volumes, and decreased calpain activation produced by 3NP. This protective effect was independent of changes in 3NP-induced succinate dehydrogenase inhibition. Furthermore, striatal p-CREB levels along with the expression of BDNF were significantly increased in sildenafil-treated rats. In summary, PDE5 inhibitors protected against 3NP-induced striatal degeneration by reducing calpain activation and by promoting survival pathways. These data encourage further evaluation of PDE5 inhibitors in transgenic mouse models of HD. Copyright 2010 Elsevier Inc. All rights reserved.

Regulation of the kynurenine metabolism pathway by Xiaoyao San and the underlying effect in the hippocampus of the depressed rat.

PubMed

Wang, Jiajia; Li, Xiaofang; He, Shugui; Hu, Lijun; Guo, Jiewen; Huang, Xiangning; Hu, Jinqing; Qi, Yaoqun; Chen, Bin; Shang, Dewei; Wen, Yuguan

2018-03-25

Xiaoyao San (XYS) is a classic Chinese herbal formula for treatment of depression. The present study aimed to investigate the antidepressant effects of XYS in a rat model of chronic unpredictable mild stress (CUMS) and the underlying mechanisms. A CUMS rat model of depression was established via 4 weeks of unpredictable stimulation. Then the rats were orally administered paroxetine and XYS for 2 weeks with continued stress. Behavioral assessments, including an open field test (OFT), sucrose preference test (SPT) and forced swim test (FST), were conducted to evaluate the antidepressant effects of XYS. The concentrations in rat plasma of tryptophan (Trp) and its metabolic products, including kynurenine (Kyn) and quinolinic acid (QUIN), were determined using high performance liquid chromatography tandem mass spectrometry with electrochemical detection (HPLC-MS/MS). The mRNA and protein levels in rat hippocampus of depression-related brain derived neurotrophic factor (BDNF), cyclic AMP response element binding protein (CREB) and nerve cell adhesion molecule (NCAM) were determined by real-time qPCR and Western blot, respectively. Enzyme Linked Immunosorbent Assay (ELISA) was used to detect the activities of indoleamine 2,3-dioxygenase (IDO) and kynurenine-3-monooxygenase (KMO) in rat plasma. The results showed that a successful CUMS rat model was established through 4 weeks of continuous unpredictable stimulation, as indicated by the significant decrease in locomotor activity and increase in immobility time in the OFT, reduction in body weight and food intake etc. Compared with the normal group, the concentrations of Kyn and QUIN had significantly (p < 0.05) decreased at day 28 in the control group, but then improved after drug treatment with paroxetine and XYS. There were no obvious changes in the activities of IDO and KMO. Compared with the normal group, the mRNA of NCAM, CREB and BDNF were significantly down-regulated (p < 0.001) in the control group, BDNF gene was up-regulated by paroxetine or XYS treatment, NCAM and CREB gene did not change in XYS group, protein expressions of BDNF and CREB were significantly increased, and NCAM was significantly reduced (p < 0.05). XYS reversed the abnormalities of the tryptophan-kynurenine metabolic pathways in depressed rats and achieved an excellent antidepressant effect. Its direct impact may be observed as changes in biological indicators in rat hippocampus tissue. Copyright © 2017 Elsevier B.V. All rights reserved.

Tangeretin triggers melanogenesis through the activation of melanogenic signaling proteins and sustained extracellular signal- regulated kinase in B16/F10 murine melanoma cells.

PubMed

Yoon, Hoon Seok; Ko, Hee-Chul; Kim, Sang Suk; Park, Kyung Jin; An, Hyun Joo; Choi, Young Hun; Kim, Se-Jae; Lee, Nam-Ho; Hyun, Chang-Gu

2015-03-01

In order to test the effectiveness of tangeretin at ameliorating melanoma and melanoma-associated depigmentation, western blotting was used to assess the melanin content of treated melanoma cells. Tangeretin, a 4',5,6,7,8-pentamethoxyflavone, was found to trigger intracellular melanin production in a concentration-dependent manner in B16/F10 murine melanoma cells. Melanin content increased 1.74-fold in response to treatment with 25 μM of tangeretin, compared to that in non-treated cells. Examination of melanogenic protein expression showed that tyrosinase, tyrosinase-related protein (TRP)-1, and extracellular signal-regulated kinase (ERK) 1/2 levels increased in a dose-dependent manner. Furthermore, the expression of cyclic adenosine monophosphate response element binding protein (CREB) and microphthalmia transcription factor (MITF) was increased by tangeretin in 1 h and 4 h, respectively. Tangeretin- upregulated melanogenesis was suppressed by ERK 1/2 inhibitor and not by ERK1 inhibitor. These results suggest that tangeretin has therapeutic potential for melanoma and melanoma-associated depigmentation because it can induce hyperpigmentation through the activation of melanogenic signaling proteins and initiation of sustained ERK2 expression.

Novel CREB3L3 Nonsense Mutation in a Family With Dominant Hypertriglyceridemia.

PubMed

Cefalù, Angelo B; Spina, Rossella; Noto, Davide; Valenti, Vincenza; Ingrassia, Valeria; Giammanco, Antonina; Panno, Maria D; Ganci, Antonina; Barbagallo, Carlo M; Averna, Maurizio R

2015-12-01

Cyclic AMP responsive element-binding protein 3-like 3 (CREB3L3) is a novel candidate gene for dominant hypertriglyceridemia. To date, only 4 kindred with dominant hypertriglyceridemia have been found to be carriers of 2 nonsense mutations in CREB3L3 gene (245fs and W46X). We investigated a family in which hypertriglyceridemia displayed an autosomal dominant pattern of inheritance. The proband was a 49-year-old woman with high plasma triglycerides (≤1300 mg/dL; 14.68 mmol/L). Her father had a history of moderate hypertriglyceridemia, and her 51-year-old brother had triglycerides levels as high as 1600 mg/dL (18.06 mmol/L). To identify the causal mutation in this family, we analyzed the candidate genes of recessive and dominant forms of primary hypertriglyceridemia by direct sequencing. The sequencing of CREB3L3 gene led to the discovery of a novel minute frame shift mutation in exon 3 of CREB3L3 gene, predicted to result in the formation of a truncated protein devoid of function (c.359delG-p.K120fsX20). Heterozygosity for the c.359delG mutation resulted in a severe phenotype occurring later in life in the proband and her brother and a good response to diet and a hypotriglyceridemic treatment. The same mutation was detected in a 13-year-old daughter who to date is normotriglyceridemic. We have identified a novel pathogenic mutation in CREB3L3 gene in a family with dominant hypertriglyceridemia with a variable pattern of penetrance. © 2015 American Heart Association, Inc.

The Neuroprotective Effect of Curcumin Against Nicotine-Induced Neurotoxicity is Mediated by CREB-BDNF Signaling Pathway.

PubMed

Motaghinejad, Majid; Motevalian, Manijeh; Fatima, Sulail; Faraji, Fahimeh; Mozaffari, Shiva

2017-10-01

Nicotine abuse adversely affects brain and causes apoptotic neurodegeneration. Curcumin- a bright yellow chemical compound found in turmeric is associated with neuroprotective properties. The current study was designed to evaluate the role of CREB-BDNF signaling in mediating the neuroprotective effects of curcumin against nicotine-induced apoptosis, oxidative stress and inflammation in rats. Sixty adult male rats were divided randomly into six groups. Group 1 received 0.7 ml/rat normal saline, group 2 received 6 mg/kg nicotine. Groups 3, 4, 5 and 6 were treated concurrently with nicotine (6 mg/kg) and curcumin (10, 20, 40 and 60 mg/kg i.p. respectively) for 21 days. Open Field Test (OFT) was used to evaluate the motor activity. Hippocampal oxidative, anti-oxidant, inflammatory and apoptotic factors were evaluated. Furthermore, phosphorylated brain cyclic adenosine monophosphate (cAMP) response element binding protein (P-CREB) and brain derived neurotrophic factor (BDNF) levels were studied at gene and protein levels. We found that nicotine disturbed the motor activity in OFT and simultaneous treatment with curcumin (40 and 60 mg/kg) reduced the nicotine-induced motor activity disturbances. In addition, nicotine treatment increased lipid peroxidation and the levels of GSH, IL-1β, TNF-α and Bax, while reducing Bcl-2, P-CREB and BDNF levels in the hippocampus. Nicotine also reduced the activity of superoxide dismutase, glutathione peroxidase and glutathione reductase in hippocampus. In contrast, various doses of curcumin attenuated nicotine-induced apoptosis, oxidative stress and inflammation; while elevating P-CREB and BDNF levels. Thus, curcumin via activation of P-CREB/BDNF signaling pathway, confers neuroprotection against nicotine-induced inflammation, apoptosis and oxidative stress.

Protective effects of a green tea polyphenol, epigallocatechin-3-gallate, against sevoflurane-induced neuronal apoptosis involve regulation of CREB/BDNF/TrkB and PI3K/Akt/mTOR signalling pathways in neonatal mice.

PubMed

Ding, Mei-Li; Ma, Hui; Man, Yi-Gang; Lv, Hong-Yan

2017-12-01

Epigallocatechin-3-gallate (EGCG), a polyphenol in green tea, is an effective antioxidant and possesses neuroprotective effects. Brain-derived neurotrophic factor (BDNF) and cyclic AMP response element-binding protein (CREB) are crucial for neurogenesis and synaptic plasticity. In this study, we aimed to assess the protective effects of EGCG against sevoflurane-induced neurotoxicity in neonatal mice. Distinct groups of C57BL/6 mice were given EGCG (25, 50, or 75 mg/kg body weight) from postnatal day 3 (P3) to P21 and were subjected to sevoflurane (3%; 6 h) exposure on P7. EGCG significantly inhibited sevoflurane-induced neuroapoptosis as determined by Fluoro-Jade B staining and terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL). Increased levels of cleaved caspase-3, downregulated Bad and Bax, and significantly enhanced Bcl-2, Bcl-xL, xIAP, c-IAP-1, and survivin expression were observed. EGCG induced activation of the PI3K/Akt pathway as evidenced by increased Akt, phospho-Akt, GSK-3β, phospho-GSK-3β, and mTORc1 levels. Sevoflurane-mediated downregulation of cAMP/CREB and BDNF/TrkB signalling was inhibited by EGCG. Reverse transcription PCR analysis revealed enhanced BDNF and TrkB mRNA levels upon EGCG administration. Improved performance of mice in Morris water maze tests suggested enhanced learning and memory. The study indicates that EGCG was able to effectively inhibit sevoflurane-induced neurodegeneration and improve learning and memory retention of mice via activation of CREB/BDNF/TrkB-PI3K/Akt signalling.

The dense core vesicle protein IA-2, but not IA-2β, is required for active avoidance learning.

PubMed

Carmona, G N; Nishimura, T; Schindler, C W; Panlilio, L V; Notkins, A L

2014-06-06

The islet-antigens IA-2 and IA-2β are major autoantigens in type-1 diabetes and transmembrane proteins in dense core vesicles (DCV). Recently we showed that deletion of both IA-2 and IA-2β alters the secretion of hormones and neurotransmitters and impairs behavior and learning. The present study was designed to evaluate the contribution to learning of each of these genes by using single knockout (SKO) and double knockout (DKO) mice in an active avoidance test. After 5 days of training, wild-type (WT) mice showed 60-70% active avoidance responses, whereas the DKO mice showed only 10-15% active avoidance responses. The degree of active avoidance responses in the IA-2 SKO mice was similar to that of the DKO mice, but in contrast, the IA-2β SKO mice behaved like WT mice showing 60-70% active avoidance responses. Molecular studies revealed a marked decrease in the phosphorylation of the cAMP response element-binding protein (CREB) and Ca(2+)/calmodulin-dependent protein kinase II (CAMKII) in the striatum and hippocampus of the IA-2 SKO and DKO mice, but not in the IA-2β SKO mice. To evaluate the role of CREB and CAMKII in the SKO and DKO mice, GBR-12909, which selectively blocks the dopamine uptake transporter and increases CREB and CAMKII phosphorylation, was administered. GBR-12909 restored the phosphorylation of CREB and CAMKII and increased active avoidance learning in the DKO and IA-2 SKO to near the normal levels found in the WT and IA-2β SKO mice. We conclude that in the absence of the DCV protein IA-2, active avoidance learning is impaired. Published by Elsevier Ltd.

Cyclic phosphatidic acid and lysophosphatidic acid induce hyaluronic acid synthesis via CREB transcription factor regulation in human skin fibroblasts.

PubMed

Maeda-Sano, Katsura; Gotoh, Mari; Morohoshi, Toshiro; Someya, Takao; Murofushi, Hiromu; Murakami-Murofushi, Kimiko

2014-09-01

Cyclic phosphatidic acid (cPA) is a naturally occurring phospholipid mediator and an analog of the growth factor-like phospholipid lysophosphatidic acid (LPA). cPA has a unique cyclic phosphate ring at the sn-2 and sn-3 positions of its glycerol backbone. We showed before that a metabolically stabilized cPA derivative, 2-carba-cPA, relieved osteoarthritis pathogenesis in vivo and induced hyaluronic acid synthesis in human osteoarthritis synoviocytes in vitro. This study focused on hyaluronic acid synthesis in human fibroblasts, which retain moisture and maintain health in the dermis. We investigated the effects of cPA and LPA on hyaluronic acid synthesis in human fibroblasts (NB1RGB cells). Using particle exclusion and enzyme-linked immunosorbent assays, we found that both cPA and LPA dose-dependently induced hyaluronic acid synthesis. We revealed that the expression of hyaluronan synthase 2 messenger RNA and protein is up-regulated by cPA and LPA treatment time dependently. We then characterized the signaling pathways up-regulating hyaluronic acid synthesis mediated by cPA and LPA in NB1RGB cells. Pharmacological inhibition and reporter gene assays revealed that the activation of the LPA receptor LPAR1, Gi/o protein, phosphatidylinositol-3 kinase (PI3K), extracellular-signal-regulated kinase (ERK), and cyclic adenosine monophosphate response element-binding protein (CREB) but not nuclear factor κB induced hyaluronic acid synthesis by the treatment with cPA and LPA in NB1RGB cells. These results demonstrate for the first time that cPA and LPA induce hyaluronic acid synthesis in human skin fibroblasts mainly through the activation of LPAR1-Gi/o followed by the PI3K, ERK, and CREB signaling pathway. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

Co-regulation of nuclear respiratory factor-1 by NFκB and CREB links LPS-induced inflammation to mitochondrial biogenesis

PubMed Central

Suliman, Hagir B.; Sweeney, Timothy E.; Withers, Crystal M.; Piantadosi, Claude A.

2010-01-01

The nuclear respiratory factor-1 (NRF1) gene is activated by lipopolysaccharide (LPS), which might reflect TLR4-mediated mitigation of cellular inflammatory damage via initiation of mitochondrial biogenesis. To test this hypothesis, we examined NRF1 promoter regulation by NFκB, and identified interspecies-conserved κB-responsive promoter and intronic elements in the NRF1 locus. In mice, activation of Nrf1 and its downstream target, Tfam, by Escherichia coli was contingent on NFκB, and in LPS-treated hepatocytes, NFκB served as an NRF1 enhancer element in conjunction with NFκB promoter binding. Unexpectedly, optimal NRF1 promoter activity after LPS also required binding by the energy-state-dependent transcription factor CREB. EMSA and ChIP assays confirmed p65 and CREB binding to the NRF1 promoter and p65 binding to intron 1. Functionality for both transcription factors was validated by gene-knockdown studies. LPS regulation of NRF1 led to mtDNA-encoded gene expression and expansion of mtDNA copy number. In cells expressing plasmid constructs containing the NRF-1 promoter and GFP, LPS-dependent reporter activity was abolished by cis-acting κB-element mutations, and nuclear accumulation of NFκB and CREB demonstrated dependence on mitochondrial H2O2. These findings indicate that TLR4-dependent NFκB and CREB activation co-regulate the NRF1 promoter with NFκB intronic enhancement and redox-regulated nuclear translocation, leading to downstream target-gene expression, and identify NRF-1 as an early-phase component of the host antibacterial defenses. PMID:20587593

Rescuing prefrontal cAMP-CREB pathway reverses working memory deficits during withdrawal from prolonged alcohol exposure.

PubMed

Dominguez, G; Dagnas, M; Decorte, L; Vandesquille, M; Belzung, C; Béracochéa, D; Mons, N

2016-03-01

Both human and animal studies indicate that alcohol withdrawal following chronic alcohol consumption (CAC) impairs many of the cognitive functions which rely on the prefrontal cortex (PFC). A candidate signaling cascade contributing to memory deficits during alcohol withdrawal is the protein kinase A (PKA)/cAMP-responsive element binding (CREB) cascade, although the role of PKA/CREB cascade in behavioral and molecular changes during sustained withdrawal period remains largely unknown. We demonstrated that 1 week (1W) or 6 weeks (6W) withdrawal after 6-month CAC impairs working memory (WM) in a T-maze spontaneous alternation task and reduces phosphorylated CREB (pCREB) in the PFC but not the dorsal CA1 region (dCA1) of the hippocampus compared with CAC and water conditions. In contrast, both CAC-unimpaired and withdrawn-impaired mice exhibited decreased pCREB in dCA1 as well as reduced histone H4 acetylation in PFC and dCA1, compared with water controls. Next, we showed that enhancing CREB activity through rolipram administration prior to testing improved WM performance in withdrawn mice but impaired WM function in water mice. In addition, WM improvement correlates positively with increased pCREB level selectively in the PFC of withdrawn mice. Results further indicate that direct infusion of the PKA activator (Sp-cAMPS) into the PFC significantly improves or impairs, respectively, WM performance in withdrawn and water animals. In contrast, Sp-cAMPS had no effect on WM when infused into the dCA1. Collectively, these results provide strong support that dysregulation of PKA/CREB-dependent processes in prefrontal neurons is a critical molecular signature underlying cognitive decline during alcohol withdrawal.

Specific rescue by ortho-hydroxy atorvastatin of cortical GABAergic neurons from previous oxygen/glucose deprivation: role of pCREB.

PubMed

Guirao, Verónica; Martí-Sistac, Octavi; DeGregorio-Rocasolano, Núria; Ponce, Jovita; Dávalos, Antoni; Gasull, Teresa

2017-11-01

The statin atorvastatin (ATV) given as a post-treatment has been reported beneficial in stroke, although the mechanisms involved are not well understood so far. Here, we investigated in vitro the effect of post-treatment with ATV and its main bioactive metabolite ortho-hydroxy ATV (o-ATV) on neuroprotection after oxygen and glucose deprivation (OGD), and the role of the pro-survival cAMP response element-binding protein (CREB). Post-OGD treatment of primary cultures of rat cortical neurons with o-ATV, but not ATV, provided neuroprotection to a specific subset of cortical neurons that were large and positive for glutamic acid decarboxylase (large-GAD (+) neurons, GABAergic). Significantly, only these GABAergic neurons showed an increase in phosphorylated CREB (pCREB) early after neuronal cultures were treated post-OGD with o-ATV. We found that o-ATV, but not ATV, increased the neuronal uptake of glutamate from the medium; this provides a rationale for the specific effect of o-ATV on pCREB in large-GABAergic neurons, which have a higher ratio of synaptic (pCREB-promoting) vs extrasynaptic (pCREB-reducing) N-methyl-D-aspartate (NMDA) receptors (NMDAR) than that of small-non-GABAergic neurons. When we pharmacologically increased pCREB levels post-OGD in non-GABAergic neurons, through the selective activation of synaptic NMDAR, we observed as well long-lasting neuronal survival. We propose that the statin metabolite o-ATV given post-OGD boosts the intrinsic pro-survival factor pCREB in large-GABAergic cortical neurons in vitro, this contributing to protect them from OGD. © 2017 International Society for Neurochemistry.

Learning strategy selection in the water maze and hippocampal CREB phosphorylation differ in two inbred strains of mice.

PubMed

Sung, Jin-Young; Goo, June-Seo; Lee, Dong-Eun; Jin, Da-Qing; Bizon, Jennifer L; Gallagher, Michela; Han, Jung-Soo

2008-04-01

Learning strategy selection was assessed in two different inbred strains of mice, C57BL/6 and DBA/2, which are used for developing genetically modified mouse models. Male mice received a training protocol in a water maze using alternating blocks of visible and hidden platform trials, during which mice escaped to a single location. After training, mice were required to choose between the spatial location where the platform had been during training (a place strategy) and a visible platform presented in a new location (a cued/response strategy). Both strains of mice had similar escape performance on the visible and hidden platform trials during training. However, in the strategy preference test, C57BL/6 mice selected a place strategy significantly more often than DBA/2 mice. Because much evidence implicates the hippocampus and striatum as important neural substrates for spatial/place and cued/response learning, respectively, the engagement of the hippocampus was then assessed after either place or cue training by determining levels of cAMP response element-binding protein (CREB) and phosphorylated CREB (pCREB) in these two mouse strains. Results revealed that hippocampal CREB levels in both strains of mice were significantly increased after place in comparison to cued training. However, the relation of hippocampal pCREB levels to training was strain dependent; pCREB was significantly higher in C57BL/6 mice than in DBA/2 mice after place training, while hippocampal pCREB levels did not differ between strains after cued training. These findings indicate that pCREB, specifically associated with place/spatial training, is closely tied to differences in spatial/place strategy preference between C57BL/6 and DBA/2 mice.

Human T-cell leukemia virus type 1 Tax oncoprotein represses the expression of the BCL11B tumor suppressor in T-cells

PubMed Central

Takachi, Takayuki; Takahashi, Masahiko; Takahashi-Yoshita, Manami; Higuchi, Masaya; Obata, Miki; Mishima, Yukio; Okuda, Shujiro; Tanaka, Yuetsu; Matsuoka, Masao; Saitoh, Akihiko; Green, Patrick L; Fujii, Masahiro

2015-01-01

Human T-cell leukemia virus type 1 (HTLV-1) is the etiological agent of adult T cell leukemia (ATL), which is an aggressive form of T-cell malignancy. HTLV-1 oncoproteins, Tax and HBZ, play crucial roles in the immortalization of T-cells and/or leukemogenesis by dysregulating the cellular functions in the host. Recent studies show that HTLV-1-infected T-cells have reduced expression of the BCL11B tumor suppressor protein. In the present study, we explored whether Tax and/or HBZ play a role in downregulating BCL11B in HTLV-1-infected T-cells. Lentiviral transduction of Tax in a human T-cell line repressed the expression of BCL11B at both the protein and mRNA levels, whereas the transduction of HBZ had little effect on the expression. Tax mutants with a decreased activity for the NF-κB, CREB or PDZ protein pathways still showed a reduced expression of the BCL11B protein, thereby implicating a different function of Tax in BCL11B downregulation. In addition, the HTLV-2 Tax2 protein reduced the BCL11B protein expression in T-cells. Seven HTLV-1-infected T-cell lines, including three ATL-derived cell lines, showed reduced BCL11B mRNA and protein expression relative to an uninfected T-cell line, and the greatest reductions were in the cells expressing Tax. Collectively, these results indicate that Tax is responsible for suppressing BCL11B protein expression in HTLV-1-infected T-cells; Tax-mediated repression of BCL11B is another mechanism that Tax uses to promote oncogenesis of HTLV-1-infected T-cells. PMID:25613934

The Antidepressant Agomelatine Improves Memory Deterioration and Upregulates CREB and BDNF Gene Expression Levels in Unpredictable Chronic Mild Stress (UCMS)-Exposed Mice

PubMed Central

Gumuslu, Esen; Mutlu, Oguz; Sunnetci, Deniz; Ulak, Guner; Celikyurt, Ipek K.; Cine, Naci; Akar, Furuzan; Savlı, Hakan; Erden, Faruk

2014-01-01

Agomelatine, a novel antidepressant with established clinical efficacy, acts as an agonist of melatonergic MT1 and MT2 receptors and as an antagonist of 5-HT2C receptors. The present study was undertaken to investigate whether chronic treatment with agomelatine would block unpredictable chronic mild stress (UCMS)-induced cognitive deterioration in mice in passive avoidance (PA), modified elevated plus maze (mEPM), novel object recognition (NOR), and Morris water maze (MWM) tests. Moreover, the effects of stress and agomelatine on brain-derived neurotrophic factor (BDNF) and cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) messenger ribonucleic acid (mRNA) levels in the hippocampus was also determined using quantitative real-time polymerase chain reaction (RT-PCR). Male inbred BALB/c mice were treated with agomelatine (10 mg/kg, i.p.), melatonin (10 mg/kg), or vehicle daily for five weeks. The results of this study revealed that UCMS-exposed animals exhibited memory deterioration in the PA, mEPM, NOR, and MWM tests. The chronic administration of melatonin had a positive effect in the PA and +mEPM tests, whereas agomelatine had a partial effect. Both agomelatine and melatonin blocked stress-induced impairment in visual memory in the NOR test and reversed spatial learning and memory impairment in the stressed group in the MWM test. Quantitative RT-PCR revealed that CREB and BDNF gene expression levels were downregulated in UCMS-exposed mice, and these alterations were reversed by chronic agomelatine or melatonin treatment. Thus, agomelatine plays an important role in blocking stress-induced hippocampal memory deterioration and activates molecular mechanisms of memory storage in response to a learning experience. PMID:24634580

Differential role of the estrogen receptors ESR1 and ESR2 on the regulation of proteins involved with proliferation and differentiation of Sertoli cells from 15-day-old rats.

PubMed

Lucas, Thaís F G; Lazari, Maria Fatima M; Porto, Catarina S

2014-01-25

The aim of the present study was to investigate the role of each estrogen receptors on the regulation of proteins involved with proliferation and differentiation of Sertoli cells from 15-day-old rats. Activation of ESR1 by 17β-estradiol (E2) and ESR1-selective agonist PPT increased CCND1 expression, and this effect was dependent on NF-kB activation. E2 and the ESR2-selective agonist DPN, but not PPT, increased, in a PI3K and CREB-dependent manner, the expression of CDKN1B and the transcription factors GATA-1 and DMRT1. Analyzing the expression of ESR1 and ESR2 in different stages of development of Sertoli cells, we observed that the ESR1/ESR2 ratio decreased with age, and this ratio seems to be important to determine the end of cell proliferation and the start of cell differentiation. In Sertoli cells from 15-day-old rats, the ESR1/ESR2 ratio favors the effect of ESR1 and the activation of this receptor increased [Methyl-(3)H]thymidine incorporation. We propose that in Sertoli cells from 15-day-old rats E2 modulates Sertoli cell proliferation through ESR1/NF-kB-mediated increase of CCND1, and cell cycle exit and differentiation through ESR2/CREB-mediated increase of CDKN1B, GATA-1 and DMRT1. The present study reinforces the important role of estrogen for normal testis development. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Direct exposure to mild heat promotes proliferation and neuronal differentiation of neural stem/progenitor cells in vitro

PubMed Central

Hossain, Md Emon; Katakura, Masanori; Sugimoto, Naotoshi; Mamun, Abdullah Al; Islam, Rafiad; Hashimoto, Michio; Shido, Osamu

2017-01-01

Heat acclimation in rats is associated with enhanced neurogenesis in thermoregulatory centers of the hypothalamus. To elucidate the mechanisms for heat acclimation, we investigated the effects of direct mild heat exposure on the proliferation and differentiation of neural stem/progenitor cells (NSCs/NPCs). The NSCs/NPCs isolated from forebrain cortices of 14.5-day-old rat fetuses were propagated as neurospheres at either 37.0°C (control) or 38.5°C (mild heat exposure) for four days, and the effects on proliferation were investigated by MTS cell viability assay, measurement of neurosphere diameter, and counting the total number of cells. The mRNA expressions of heat shock proteins (HSPs) and brain-derived neurotrophic factor (BDNF), cAMP response element-binding (CREB) protein and Akt phosphorylation levels, and intracellular reactive oxygen species (ROS) levels were analyzed using real time PCR, Western blotting and CM-H2DCFDA assay respectively. Heat exposure under proliferation condition increased NSC/NPC viability, neurosphere diameter, and cell count. BDNF mRNA expression, CREB phosphorylation, and ROS level were also increased by heat exposure. Heat exposure increased HSP27 mRNA expression concomitant with enhanced p-Akt level. Moreover, treatment with LY294002 (a PI3K inhibitor) abolished the effects of heat exposure on NSC/NPC proliferation. Furthermore, heat exposure under differentiation conditions increased the proportion of cells positive for Tuj1 (a neuronal marker). These findings suggest that mild heat exposure increases NSC/NPC proliferation, possibly through activation of the Akt pathway, and also enhances neuronal differentiation. Direct effects of temperature on NSCs/NPCs may be one of the mechanisms involved in hypothalamic neurogenesis in heat-acclimated rats. Such heat-induced neurogenesis could also be an effective therapeutic strategy for neurodegenerative diseases. PMID:29287093

PPARγ activation attenuates cold-induced upregulation of thyroid status and brown adipose tissue PGC-1α and D2

PubMed Central

Festuccia, William T.; Blanchard, Pierre-Gilles; Oliveira, Thiago B.; Magdalon, Juliana; Paschoal, Vivian A.; Richard, Denis

2012-01-01

Here, we investigated whether pharmacological PPARγ activation modulates key early events in brown adipose tissue (BAT) recruitment induced by acute cold exposure with the aim of unraveling the interrelationships between sympathetic and PPARγ signaling. Sprague-Dawley rats treated or not with the PPARγ ligand rosiglitazone (15 mg·kg−1·day−1, 7 days) were kept at 23°C or exposed to cold (5°C) for 24 h and evaluated for BAT gene expression, sympathetic activity, thyroid status, and adrenergic signaling. Rosiglitazone did not affect the reduction in body weight gain and the increase in feed efficiency, V̇o2, and BAT sympathetic activity induced by 24-h cold exposure. Rosiglitazone strongly attenuated the increase in serum total and free T4 and T3 levels and BAT iodothyronine deiodinase type 2 (D2) and PGC-1α mRNA levels and potentiated the reduction in BAT thyroid hormone receptor (THR) β mRNA levels induced by cold. Administration of T3 to rosiglitazone-treated rats exacerbated the cold-induced increase in energy expenditure but did not restore a proper activation of D2 and PGC-1α, nor further increased uncoupling protein 1 expression. Regarding adrenergic signaling, rosiglitazone did not affect the changes in BAT cAMP content and PKA activity induced by cold. Rosiglitazone alone or in combination with cold increased CREB binding to DNA, but it markedly reduced the expression of one of its major coactivators, CREB binding protein. In conclusion, pharmacological PPARγ activation impairs short-term cold elicitation of BAT adrenergic and thyroid signaling, which may result in abnormal tissue recruitment and thermogenic activity. PMID:23100029

Estrogen Modulates Specific Life and Death Signals Induced by LH and hCG in Human Primary Granulosa Cells In Vitro.

PubMed

Casarini, Livio; Riccetti, Laura; De Pascali, Francesco; Gilioli, Lisa; Marino, Marco; Vecchi, Eugenia; Morini, Daria; Nicoli, Alessia; La Sala, Giovanni Battista; Simoni, Manuela

2017-04-28

Luteinizing hormone (LH) and human chorionic gonadotropin (hCG) are glycoprotein hormones used for assisted reproduction acting on the same receptor (LHCGR) and mediating different intracellular signaling. We evaluated the pro- and anti-apoptotic effect of 100 pM LH or hCG, in the presence or in the absence of 200 pg/mL 17β-estradiol, in long-term, serum-starved human primary granulosa cells (hGLC) and a transfected granulosa cell line overexpressing LHCGR (hGL5/LHCGR). To this purpose, phospho-extracellular-regulated kinase 1/2 (pERK1/2), protein kinase B (pAKT), cAMP-responsive element binding protein (pCREB) activation and procaspase 3 cleavage were evaluated over three days by Western blotting, along with the expression of target genes by real-time PCR and cell viability by colorimetric assay. We found that LH induced predominant pERK1/2 and pAKT activation STARD1 , CCND2 and anti-apoptotic XIAP gene expression, while hCG mediated more potent CREB phosphorylation, expression of CYP19A1 and procaspase 3 cleavage than LH. Cell treatment by LH is accompanied by increased (serum-starved) cell viability, while hCG decreased the number of viable cells. The hCG-specific, pro-apoptotic effect was blocked by a physiological dose of 17β-estradiol, resulting in pAKT activation, lack of procaspase 3 cleavage and increased cell viability. These results confirm that relatively high levels of steroidogenic pathway activation are linked to pro-apoptotic signals in vitro, which may be counteracted by other factors, i.e., estrogens.

Sigma 1 receptor activation regulates brain-derived neurotrophic factor through NR2A-CaMKIV-TORC1 pathway to rescue the impairment of learning and memory induced by brain ischaemia/reperfusion.

PubMed

Xu, Qian; Ji, Xue-Fei; Chi, Tian-Yan; Liu, Peng; Jin, Ge; Gu, Shao-Li; Zou, Li-Bo

2015-05-01

Sigma-1 receptor (Sig-1R) agonists showed anti-amnesic properties in Alzheimer's disease models and anti-inflammatory properties in cerebrum ischaemia models. The agonist of Sig-1R was reported to up-regulate brain-derived neurotrophic factor (BDNF) levels in the hippocampus of mice. Here, we investigate whether the activation of Sig-1R attenuates the learning and memory impairment induced by ischaemia/reperfusion and how it affects the expression of BDNF. Bilateral common carotid artery occlusion (BCCAO) was induced for 20 min in C57BL/6 mice. Sig-1R agonist, PRE084, sigma 1/2 non-selective agonist, DTG, Sig-1R antagonist and BD1047 were injected once daily throughout the experiment. Behavioural tests were performed from day 8. On day 22 after BCCAO, mice were sacrificed for biochemical analysis. PRE084 and DTG ameliorated learning and memory impairments in the Y maze, novel object recognition, and water maze tasks and prevented the decline of synaptic proteins and BDNF expression in the hippocampus of BCCAO mice. Furthermore, PRE084 and DTG up-regulated the level of NMDA receptor 2A (NR2A), calcium/calmodulin-dependent protein kinase type IV (CaMKIV) and CREB-specific co-activator transducer of regulated CREB activity 1 (TORC1). Additionally, the effects of PRE084 and DTG were antagonised by the co-administration of BD1047. Sig-1R activation showed an attenuation in the ischaemia/reperfusion model and the activation of Sig-1R increased the expression of BDNF, possibly through the NR2A-CaMKIV-TORC1 pathway, and Sig-1R agonists might function as neuroprotectant agents in vascular dementia.

Bi-functional, substrate mimicking RNA inhibits MSK1-mediated cAMP-response element-binding protein phosphorylation and reveals magnesium ion-dependent conformational changes of the kinase.

PubMed

Hamm, Jorg; Alessi, Dario R; Biondi, Ricardo M

2002-11-29

The design of specific inhibitors for protein kinases is an important step toward elucidation of intracellular signal transduction pathways and to guide drug discovery programs. We devised a model approach to generate specific, competitive kinase inhibitors by isolating substrate mimics containing two independent binding sites with an anti-idiotype strategy from combinatorial RNA libraries. As a general test for the ability to generate highly specific kinase inhibitors, we selected the transcription factor cAMP-response element-binding protein (CREB) that is phosphorylated on the same serine residue by the protein kinase MSK1 as well as by RSK1. The sequences and structures of these kinases are very similar, about 60% of their amino acids are identical. Nevertheless, we can demonstrate that the selected RNA inhibitors inhibit specifically CREB phosphorylation by MSK1 but do not affect CREB phosphorylation by RSK1. The inhibitors interact preferentially with the inactive form of MSK1. Furthermore, we demonstrate that RNA ligands can be conformation-specific probes, and this feature allowed us to describe magnesium ion-dependent conformational changes of MSK1 upon activation.

Neuroprotection of early and short-time applying berberine in the acute phase of cerebral ischemia: up-regulated pAkt, pGSK and pCREB, down-regulated NF-κB expression, ameliorated BBB permeability.

PubMed

Zhang, Xiaolin; Zhang, Xiangjian; Wang, Chaohui; Li, Yanhua; Dong, Lipeng; Cui, Lili; Wang, Lina; Liu, Zongjie; Qiao, Huimin; Zhu, Chunhua; Xing, Yinxue; Cao, Xiaoyun; Ji, Ye; Zhao, Kang

2012-06-12

Berberine (BBR) has gained attention for its vast beneficial biological effects through immunomodulation, anti-inflammatory and anti-apoptosis properties. Inflammatory and apoptosis damage play an important role in cerebral ischemic pathogenesis and may represent a target for treatment. The aim of this study was to explore BBR's effect in ischemic injury and the role of the Akt/GSK (glycogen synthase kinase) signaling cascade in mediating the anti-apoptosis and anti-inflammatory effects in the rat brain of permanent middle cerebral artery occlusion (pMCAO). Male Sprague-Dawley rats were subjected to pMCAO and randomly assigned into four groups: Sham (sham-operated) group, pMCAO (pMCAO+0.9% saline) group, BBR-L (pMCAO+BBR 10 mg/kg) and BBR-H (pMCAO+BBR 40 mg/kg) group. BBR was administered immediately after pMCAO and the neuroprotection was detected. Phospho-Akt (pAkt), phospho-glycogen synthase kinase 3-β (pGSK3β), phospho-cAMP response element binding protein (pCREB), nuclear factor-kappa B (NF-κB) and claudin-5 in ischemic cerebral cortex were detected by immunohistochemistry, reverse transcription-polymerase chain reaction and western blotting. Compared with pMCAO group, BBR dramatically lessened neurological deficits scores, brain water contents and infarct sizes, upregulated the expression of pAkt, pGSK3β, pCREB and claudin-5, and decreased the nuclear accumulation of NF-κB (P<0.05) in ischemic brain. The results showed that BBR reduced ischemic brain injury after pMACO, and this effect may be via the increasing the activation of Akt/GSK signaling and claudin-5, and decreasing NF-κB expression. Copyright © 2012 Elsevier B.V. All rights reserved.

CREB, NF-Y and MEIS1 conserved binding sites are essential to balance Myostatin promoter/enhancer activity during early myogenesis.

PubMed

Grade, Carla Vermeulen Carvalho; Mantovani, Carolina Stefano; Fontoura, Marina Alves; Yusuf, Faisal; Brand-Saberi, Beate; Alvares, Lúcia Elvira

2017-10-01

Myostatin (MSTN) is a strong inhibitor of skeletal muscle growth in human and other vertebrates. Its transcription is controlled by a proximal promoter/enhancer (Mstn P/E) containing a TATA box besides CREB, NF-Y, MEIS1 and FXR transcription factor binding sites (TFBSs), which are conserved throughout evolution. The aim of this work was to investigate the role of these TFBSs on Mstn P/E activity and evaluate the potential of their putative ligands as Mstn trans regulators. Mstn P/E mutant constructs were used to establish the role of conserved TFBSs using dual-luciferase assays. Expression analyses were performed by RT-PCR and in situ hybridization in C2C12 myoblasts and E10.5 mouse embryos, respectively. Our results revealed that CREB, NF-Y and MEIS1 sites are required to balance Mstn P/E activity, keeping Mstn transcription within basal levels during myoblast proliferation. Furthermore, our data showed that NF-Y site is essential, although not sufficient, to mediate Mstn P/E transcriptional activity. In turn, CREB and MEIS1 binding sites seem to depend on the presence of NF-Y site to induce Mstn P/E. FXR appears not to confer any effect on Mstn P/E activity, except in the absence of all other conserved TFBS. Accordingly, expression studies pointed to CREB, NF-Y and MEIS1 but not to FXR factors as possible regulators of Mstn transcription in the myogenic context. Altogether, our findings indicated that CREB, NF-Y and MEIS1 conserved sites are essential to control basal Mstn transcription during early myogenesis, possibly by interacting with these or other related factors.

Moringa oleifera Seed Extract Alleviates Scopolamine-Induced Learning and Memory Impairment in Mice

PubMed Central

Zhou, Juan; Yang, Wu-shuang; Suo, Da-qin; Li, Ying; Peng, Lu; Xu, Lan-xi; Zeng, Kai-yue; Ren, Tong; Wang, Ying; Zhou, Yu; Zhao, Yun; Yang, Li-chao; Jin, Xin

2018-01-01

The extract of Moringa oleifera seeds has been shown to possess various pharmacological properties. In the present study, we assessed the neuropharmacological effects of 70% ethanolic M. oleifera seed extract (MSE) on cognitive impairment caused by scopolamine injection in mice using the passive avoidance and Morris water maze (MWM) tests. MSE (250 or 500 mg/kg) was administered to mice by oral gavage for 7 or 14 days, and cognitive impairment was induced by intraperitoneal injection of scopolamine (4 mg/kg) for 1 or 6 days. Mice that received scopolamine alone showed impaired learning and memory retention and considerably decreased cholinergic system reactivity and neurogenesis in the hippocampus. MSE pretreatment significantly ameliorated scopolamine-induced cognitive impairment and enhanced cholinergic system reactivity and neurogenesis in the hippocampus. Additionally, the protein expressions of phosphorylated Akt, ERK1/2, and CREB in the hippocampus were significantly decreased by scopolamine, but these decreases were reversed by MSE treatment. These results suggest that MSE-induced ameliorative cognitive effects are mediated by enhancement of the cholinergic neurotransmission system and neurogenesis via activation of the Akt, ERK1/2, and CREB signaling pathways. These findings suggest that MSE could be a potent neuropharmacological drug against amnesia, and its mechanism might be modulation of cholinergic activity via the Akt, ERK1/2, and CREB signaling pathways. PMID:29740317

Identification of proteins interacting with Toxoplasma SRCAP by yeast two-hybrid screening.

PubMed

Nallani, Karuna C; Sullivan, William J

2005-03-01

Toxoplasma gondii is an opportunistic protozoan parasite that differentiates into latent cysts (bradyzoite) that can be reactivated during immunosuppression. TgSRCAP (Toxoplasma gondii Snf2-related CBP activator protein) is a SWI2/SNF2 family chromatin remodeler whose expression increases during cyst development. Identifying the proteins associating with TgSRCAP during the pre-cyst stage (tachyzoite) will increase our understanding of how parasite differentiation is initiated. We employed the yeast two-hybrid system to identify proteins that may interact directly with TgSRCAP. A stretch of 1,060 amino acids between ATPase subdomains IV and V of TgSRCAP was chosen as "bait" since the corresponding region in human SRCAP interacts with other proteins, including CREB binding protein. We have identified several novel parasite-specific transcription factors predicted to be in the T. gondii genome. Metabolic enzymes that may participate in cyst development were also identified as interacting with TgSRCAP.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Sun Young; Kim, Ji-Hee; Lee, Sang Joon

Surfactin, one of the most powerful biosurfactants, is a bacterial cyclic lipopeptide. Here, we investigated the anti-neuroinflammatory properties of surfactin in lipoteichoic acid (LTA)-stimulated BV-2 microglial cells. Surfactin significantly inhibited excessive production of the pro-inflammatory mediators TNF-α, IL-1β, IL-6, monocyte chemoattractant protein-1 (MCP-1), prostaglandin E{sub 2} (PGE{sub 2}), nitric oxide (NO) and reactive oxygen species (ROS), and suppressed the expression of matrix metalloproteinase-9 (MMP-9), inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2). Subsequent mechanistic studies revealed that surfactin inhibited LTA-induced nuclear factor-kappaB (NF-κB) and signal transducer and activator of transcription-1 (STAT-1) activation. However, surfactin increases the phosphorylation of the STAT-3, amore » component of the homeostatic mechanism causing anti-inflammatory events. We also demonstrated that surfactin induces heme oxygenase-1 (HO-1) expression and nuclear factor-regulated factor-2 (Nrf-2) activation, and that the anti-inflammatory effects of surfactin are abrogated by small interfering RNA-mediated knock-down of HO-1 or Nrf-2. Interestingly, we found that surfactin increased the level of cAMP and induced phosphorylation of cAMP responsive element binding protein (CREB) in microglial cells. Furthermore, treatment with the protein kinase A (PKA) inhibitor, H-89, blocked HO-1 induction by surfactin and abolished surfactin's suppressive effects on ROS and NO production. These results indicate that HO-1 and its upstream effector, PKA, play a pivotal role in the anti-neuroinflammatory response of surfactin in LTA-stimulated microglia. Therefore, surfactin might have therapeutic potential for neuroprotective agents to treat inflammatory and neurodegenerative diseases. - Highlights: ► Surfactin inhibits proinflammatory mediator synthesis in LTA-activated BV-2 cells. ► Surfactin suppresses NF-κB and STAT-1, but potentiates phosphorylation of STAT-3. ► Surfactin induces HO-1 expression and Nrf-2 activation. ► Surfactin induces cAMP and CREB phosphorylation. ► PKA inhibitor blocks HO-1 induction and surfactin’s antiinflammatory effects.« less

Membrane-Associated Effects of Glucocorticoid on BACE1 Upregulation and Aβ Generation: Involvement of Lipid Raft-Mediated CREB Activation.

PubMed

Choi, Gee Euhn; Lee, Sei-Jung; Lee, Hyun Jik; Ko, So Hee; Chae, Chang Woo; Han, Ho Jae

2017-08-30

Glucocorticoid has been widely accepted to induce Alzheimer's disease, but the nongenomic effect of glucocorticoid on amyloid β (Aβ) generation has yet to be studied. Here, we investigated the effect of the nongenomic pathway induced by glucocorticoid on amyloid precursor protein processing enzymes as well as Aβ production using male ICR mice and human neuroblastoma SK-N-MC cells. Mice groups exposed to restraint stress or intracerebroventricular injection of Aβ showed impaired cognition, decreased intracellular glucocorticoid receptor (GR) level, but elevated level of membrane GR (mGR). In this respect, we identified the mGR-dependent pathway evoked by glucocorticoid using impermeable cortisol conjugated to BSA (cortisol-BSA) on SK-N-MC cells. Cortisol-BSA augmented the expression of β-site amyloid precursor protein cleaving enzyme 1 (BACE1), the level of C-terminal fragment β of amyloid precursor protein (C99) and Aβ production, which were maintained even after blocking intracellular GR. We also found that cortisol-BSA enhanced the interaction between mGR and Gαs, which colocalized in the lipid raft. The subsequently activated CREB by cortisol-BSA bound to the CRE site of the BACE1 promoter increasing its expression, which was downregulated by inhibiting CBP. Consistently, blocking CBP attenuated cognitive impairment and Aβ production induced by corticosterone treatment or intracerebroventricular injection of Aβ more efficiently than inhibiting intracellular GR in mice. In conclusion, glucocorticoid couples mGR with Gαs and triggers cAMP-PKA-CREB axis dependent on the lipid raft to stimulate BACE1 upregulation and Aβ generation. SIGNIFICANCE STATEMENT Patients with Alzheimer's disease (AD) have been growing sharply and stress is considered as the major environment factor of AD. Glucocorticoid is the primarily responsive factor to stress and is widely known to induce AD. However, most AD patients usually have impaired genomic pathway of glucocorticoid due to intracellular glucocorticoid receptor deficiency. In this respect, the genomic mechanism of glucocorticoid faces difficulties in explaining the consistent amyloid β (Aβ) production. Therefore, it is necessary to investigate the novel pathway of glucocorticoid on Aβ generation to find a more selective therapeutic approach to AD patients. In this study, we revealed the importance of nongenomic pathway induced by glucocorticoid where membrane glucocorticoid receptor plays an important role in Aβ formation. Copyright © 2017 the authors 0270-6474/17/378459-18$15.00/0.

The Role of JMY in p53 Regulation.

PubMed

Adighibe, Omanma; Pezzella, Francesco

2018-05-31

Following the event of DNA damage, the level of tumour suppressor protein p53 increases inducing either cell cycle arrest or apoptosis. Junctional Mediating and Regulating Y protein (JMY) is a transcription co-factor involved in p53 regulation. In event of DNA damage, JMY levels also upregulate in the nucleus where JMY forms a co-activator complex with p300/CREB-binding protein (p300/CBP), Apoptosis-stimulating protein of p53 (ASPP) and Stress responsive activator of p53 (Strap). This co-activator complex then binds to and increases the ability of p53 to induce transcription of proteins triggering apoptosis but not cell cycle arrest. This then suggests that the increase of JMY levels due to DNA damage putatively "directs" p53 activity toward triggering apoptosis. JMY expression is also linked to increased cell motility as it: (1) downregulates the expression of adhesion molecules of the Cadherin family and (2) induces actin nucleation, making cells less adhesive and more mobile, favouring metastasis. All these characteristics taken together imply that JMY possesses both tumour suppressive and tumour metastasis promoting capabilities.

Ternary Complex Factors and Cofactors Are Essential for Human T-Cell Leukemia Virus Type 1 Tax Transactivation of the Serum Response Element

PubMed Central

Shuh, Maureen; Derse, David

2000-01-01

The human T-cell leukemia virus type 1 Tax protein activates the expression of cellular immediate early genes controlled by the serum response element (SRE), which contains both the serum response factor (SRF) binding element (CArG box) and the ternary complex factor (TCF) binding element (Ets box). We show that TCF binding is necessary for Tax activation of the SRE and that Tax directly interacts with TCFs in vitro. In addition, Tax interactions with CREB binding protein (CBP) and p300- and CBP-associated factor were found to be essential for Tax activation of SRF-mediated transcription. PMID:11070040

An Exposed KID-Like Domain in Human T-Cell Lymphotropic Virus Type 1 Tax Is Responsible for the Recruitment of Coactivators CBP/p300

PubMed Central

Harrod, Robert; Tang, Yong; Nicot, Christophe; Lu, Hsieng S.; Vassilev, Alex; Nakatani, Yoshihiro; Giam, Chou-Zen

1998-01-01

Human T-cell lymphotropic virus type 1 (HTLV-1) transcriptional activation is mediated by the viral transactivator, Tax, and three 21-bp repeats (Tax response element [TxRE]) located in the U3 region of the viral long terminal repeat (LTR). Each TxRE contains a core cyclic AMP response element (CRE) flanked by 5′ G-rich and 3′ C-rich sequences. The TxRE binds CREB (CRE-binding protein) and Tax to form a ternary complex and confers Tax-dependent transactivation. Recent data indicate that Tax functions as a specific link to connect CREB-binding protein (CBP)/p300 in a phosphorylation-independent manner to CREB/ATF-1 assembled on the viral 21-bp repeats. Glutathione S-transferase pull-down performed with Tax deletion mutants and peptide competition have localized the site in Tax critical for binding CBP/p300 to a highly protease-sensitive region around amino acid residues 81 to 95 (81QRTSKTLKVLTPPIT95) which lies between the domains previously proposed to be important for CREB binding and Tax subunit dimerization. Amino acid residues around the trypsin- and chymotrypsin-sensitive sites (88KVL90) of Tax bear resemblance to those in the kinase-inducible domain of CREB (129SRRPSYRKILNE140) surrounding Ser-133, which undergoes signal-induced phosphorylation to recruit CBP/p300. Site-directed mutagenesis of residues in this domain (R82A, K85A, K88A, and V89A) resulted in proteins which failed to transactivate from the HTLV-1 LTR in vivo. These mutants (K85A, K88A, and V89A) bind CREB with similar affinities as wild-type Tax, yet interaction with CBP/p300 is abrogated in various biochemical assays, indicating that the recruitment of CBP/p300 is crucial for Tax transactivation. A Tax mutant, M47, defective in the COOH-terminal transactivation domain, continued to interact with CBP/p300, suggesting that interactions with additional cellular factors are required for proper Tax function. PMID:9710589

LINC01121 Inhibits Cell Apoptosis While Facilitating Proliferation, Migration, and Invasion Though Negative Regulation of the Camp/PKA Signaling Pathway via GLP1R.

PubMed

Qian, Yi-Gang; Ye, Zhou; Chen, Hai-Yong; Lv, Zhen; Zhang, Ai-Bin; Fan, Le; Zhou, Jie; Zheng, Shu-Sen; Wang, Wei-Lin

2018-05-24

Pancreatic cancer is an aggressive malignancy as a result of highly metastatic potential. The current study was carried out to alter the expression of LINC01121 in pancreatic cancer, with the aim of elucidating its effects on the biological processes of cell proliferation, migration, invasion, and apoptosis. We hypothesized that both the GLP1R gene and cAMP/PKA signaling pathway participate in the aforementioned process. Microarray data (GSE14245, GSE27890 and GSE16515) and annotating probe files linked to pancreatic cancer were downloaded through the GEO database. The Multi Experiment Matrix (MEM) site was used to predict the target gene of lncRNA. Both pancreatic cancer tissues (n = 56) and paracancerous tissues (n = 45) were collected from patients diagnosed with pancreatic cancer. Immunohistochemistry was applied to identify the positive expression rate of GLP1R protein. Isolated pancreatic cancer cells and PANC-1 cells were independently classified into the blank, negative control (NC), LINC01121 vector, siRNA-LINC01121, siRNA-GLP1R and siRNA-LINC01121 + siRNA-GLP1R groups. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis were applied to detect the expressions of LINC01121, GLP1R, cAMP, PKA, CREB, Bcl-2, Bad and PCNA. Cell proliferation, migration, invasion, cycle progression, and apoptosis were examined by MTT assay, scratch test, Transwell assay and flow cytometry analyses of Annexin V-FITC/PI staining. Observations were made indicating that LINC01121 was highly expressed, while low expressions of GLP1R in pancreatic cancer were detected based on microarray data, which was largely in consistent with the data collected of LINC01121 and GLP1R within the tissues. The target prediction program and luciferase activity analysis was testament to the notion suggesting that GLP1R was indeed a target of LINC01121. In contrast to the blank and NC groups, the LINC01121 vector group exhibited increased expressions of LINC01121; decreased mRNA and protein levels of GLP1R, Bad, cAMP, and PKA; increased protein levels of CREB, Bcl-2, PCNA, p-PKA and p-CREB; increased cell proliferation, migration and invasion; and decreased cell apoptosis. There was no significant difference detected among the blank, NC, and siRNA-LINC01121 + siRNA-GLP1R groups, except that decreased LINC01121 expression was determined in the siRNA-LINC01121 + siRNA-GLP1R group. Parallel data were observed in the pancreatic cancer cells and PANC-1 cells. The current study presents evidence indicating that LINC01121 might inhibit apoptosis while acting to promote proliferation, migration, and invasion of pancreatic cancer cells, supplementing the stance held that LINC01121 functions as a tumor promoter by means of its involvement in the process of translational repression of the GLP1R and inhibition of the cAMP/PKA signaling pathway. © 2018 The Author(s). Published by S. Karger AG, Basel.

CREB and the discovery of cognitive enhancers.

PubMed

Scott, Roderick; Bourtchuladze, Rusiko; Gossweiler, Scott; Dubnau, Josh; Tully, Tim

2002-01-01

In the past few years, a series of molecular-genetic, biochemical, cellular and behavioral studies in fruit flies, sea slugs and mice have confirmed a long-standing notion that long-term memory formation depends on the synthesis of new proteins. Experiments focused on the cAMP-responsive transcription factor, CREB, have established that neural activity-induced regulation of gene transcription promotes a synaptic growth process that strengthens the connections among active neurons. This process constitutes a physical basis for the engram--and CREB is a "molecular switch" to produce the engram. Helicon Therapeutics has been formed to identify drug compounds that enhance memory formation via augmentation of CREB biochemistry. Candidate compounds have been identified from a high throughput cell-based screen and are being evaluated in animal models of memory formation. A gene discovery program also seeks to identify new genes, which function downstream of CREB during memory formation, as a source for new drug discoveries in the future. Together, these drug and gene discovery efforts promise new class of pharmaceutical therapies for the treatment of various forms of cognitive dysfunction.

Early-onset sleep defects in Drosophila models of Huntington's disease reflect alterations of PKA/CREB signaling

PubMed Central

Gonzales, Erin D.; Tanenhaus, Anne K.; Zhang, Jiabin; Chaffee, Ryan P.; Yin, Jerry C.P.

2016-01-01

Huntington's disease (HD) is a progressive neurological disorder whose non-motor symptoms include sleep disturbances. Whether sleep and activity abnormalities are primary molecular disruptions of mutant Huntingtin (mutHtt) expression or result from neurodegeneration is unclear. Here, we report Drosophila models of HD exhibit sleep and activity disruptions very early in adulthood, as soon as sleep patterns have developed. Pan-neuronal expression of full-length or N-terminally truncated mutHtt recapitulates sleep phenotypes of HD patients: impaired sleep initiation, fragmented and diminished sleep, and nighttime hyperactivity. Sleep deprivation of HD model flies results in exacerbated sleep deficits, indicating that homeostatic regulation of sleep is impaired. Elevated PKA/CREB activity in healthy flies produces patterns of sleep and activity similar to those in our HD models. We were curious whether aberrations in PKA/CREB signaling were responsible for our early-onset sleep/activity phenotypes. Decreasing signaling through the cAMP/PKA pathway suppresses mutHtt-induced developmental lethality. Genetically reducing PKA abolishes sleep/activity deficits in HD model flies, restores the homeostatic response and extends median lifespan. In vivo reporters, however, show dCREB2 activity is unchanged, or decreased when sleep/activity patterns are abnormal, suggesting dissociation of PKA and dCREB2 occurs early in pathogenesis. Collectively, our data suggest that sleep defects may reflect a primary pathological process in HD, and that measurements of sleep and cAMP/PKA could be prodromal indicators of disease, and serve as therapeutic targets for intervention. PMID:26604145

Context-specific modulation of cocaine-induced locomotor sensitization and ERK and CREB phosphorylation in rat nucleus accumbens

PubMed Central

Marin, Marcelo T.; Berkow, Alexander; Golden, Sam A.; Koya, Eisuke; Planeta, Cleopatra S.; Hope, Bruce T.

2009-01-01

Learned associations are hypothesized to develop between drug effects and contextual stimuli during repeated drug administration to produce context-specific sensitization that is expressed only in the drug-associated environment and not in a non-drug paired environment. Neuroadaptations that mediate such context-specific behavior are largely unknown. We investigated context-specific modulation of CREB phosphorylation and four upstream kinases in nucleus accumbens that phosphorylate CREB, including ERK, PKA, CaMKII and IV. Rats received seven once daily injections of cocaine or saline in one of two distinct environments outside their home cages. Seven days later, test injections of cocaine or saline were administered in either the Paired or the Non-paired environment. CREB and ERK phosphorylation were assessed with immunohistochemistry while phosphorylation of the remaining kinases, as well as CREB and ERK, were assessed by Western blotting. Repeated cocaine administration produced context-specific sensitized locomotor responses accompanied by context-specific enhancement of the number of cocaine-induced phosphoCREB and phosphoERK immunoreactive nuclei in a minority of neurons. In contrast, CREB and CaMKIV phosphorylation in nucleus accumbens homogenates were decreased by cocaine test injections. We have recently shown that a small number of cocaine-activated accumbens neurons mediate the learned association between cocaine effects and the drug administration environment to produce context-specific sensitization. The corresponding cocaine and context-specific phosphorylation of ERK and CREB in cocaine-activated accumbens neurons in the present study suggests that this signal transduction pathway is also selectively activated in the same set of accumbens neurons. PMID:19912338

Mechanical loading stimulates chondrogenesis via the PKA/CREB-Sox9 and PP2A pathways in chicken micromass cultures.

PubMed

Juhász, Tamás; Matta, Csaba; Somogyi, Csilla; Katona, Éva; Takács, Roland; Soha, Rudolf Ferenc; Szabó, István A; Cserháti, Csaba; Sződy, Róbert; Karácsonyi, Zoltán; Bakó, Eva; Gergely, Pál; Zákány, Róza

2014-03-01

Biomechanical stimuli play important roles in the formation of articular cartilage during early foetal life, and optimal mechanical load is a crucial regulatory factor of adult chondrocyte metabolism and function. In this study, we undertook to analyse mechanotransduction pathways during in vitro chondrogenesis. Chondroprogenitor cells isolated from limb buds of 4-day-old chicken embryos were cultivated as high density cell cultures for 6 days. Mechanical stimulation was carried out by a self-designed bioreactor that exerted uniaxial intermittent cyclic load transmitted by the culture medium as hydrostatic pressure and fluid shear to differentiating cells. The loading scheme (0.05 Hz, 600 Pa; for 30 min) was applied on culturing days 2 and 3, when final commitment and differentiation of chondroprogenitor cells occurred in this model. The applied mechanical load significantly augmented cartilage matrix production and elevated mRNA expression of several cartilage matrix constituents, including collagen type II and aggrecan core protein, as well as matrix-producing hyaluronan synthases through enhanced expression, phosphorylation and nuclear signals of the main chondrogenic transcription factor Sox9. Along with increased cAMP levels, a significantly enhanced protein kinase A (PKA) activity was also detected and CREB, the archetypal downstream transcription factor of PKA signalling, exhibited elevated phosphorylation levels and stronger nuclear signals in response to mechanical stimuli. All the above effects were diminished by the PKA-inhibitor H89. Inhibition of the PKA-independent cAMP-mediators Epac1 and Epac2 with HJC0197 resulted in enhanced cartilage formation, which was additive to that of the mechanical stimulation, implying that the chondrogenesis-promoting effect of mechanical load was independent of Epac. At the same time, PP2A activity was reduced following mechanical load and treatments with the PP2A-inhibitor okadaic acid were able to mimic the effects of the intervention. Our results indicate that proper mechanical stimuli augment in vitro cartilage formation via promoting both differentiation and matrix production of chondrogenic cells, and the opposing regulation of the PKA/CREB-Sox9 and the PP2A signalling pathways is crucial in this phenomenon. Copyright © 2014 Elsevier Inc. All rights reserved.

Molecular and Cellular Mechanisms for Trapping and Activating Emotional Memories

PubMed Central

Cai, Denise J.; Sano, Yoshitake; Lee, Yong-Seok; Zhou, Yu; Bekal, Pallavi; Deisseroth, Karl; Silva, Alcino J.

2016-01-01

Recent findings suggest that memory allocation to specific neurons (i.e., neuronal allocation) in the amygdala is not random, but rather the transcription factor cAMP-response element binding protein (CREB) modulates this process, perhaps by regulating the transcription of channels that control neuronal excitability. Here, optogenetic studies in the mouse lateral amygdala (LA) were used to demonstrate that CREB and neuronal excitability regulate which neurons encode an emotional memory. To test the role of CREB in memory allocation, we overexpressed CREB in the lateral amygdala to recruit the encoding of an auditory-fear conditioning (AFC) memory to a subset of neurons. Then, post-training activation of these neurons with Channelrhodopsin-2 was sufficient to trigger recall of the memory for AFC, suggesting that CREB regulates memory allocation. To test the role of neuronal excitability in memory allocation, we used a step function opsin (SFO) to transiently increase neuronal excitability in a subset of LA neurons during AFC. Post-training activation of these neurons with Volvox Channelrhodopsin-1 was able to trigger recall of that memory. Importantly, our studies show that activation of the SFO did not affect AFC by either increasing anxiety or by strengthening the unconditioned stimulus. Our findings strongly support the hypothesis that CREB regulates memory allocation by modulating neuronal excitability. PMID:27579481

Social isolation mediated anxiety like behavior is associated with enhanced expression and regulation of BDNF in the female mouse brain.

PubMed

Kumari, Anita; Singh, Padmanabh; Baghel, Meghraj Singh; Thakur, M K

2016-05-01

Adverse early life experience is prominent risk factors for numerous psychiatric illnesses, including mood and anxiety disorders. It imposes serious long-term costs on the individual as well as health and social systems. Hence, developing therapies that prevent the long-term consequences of early life stress is of utmost importance, and necessitates a better understanding of the mechanisms by which early life stress triggers long-lasting alterations in gene expression and behavior. Post-weaning isolation rearing of rodents models the behavioral consequences of adverse early life experiences in humans and it is reported to cause anxiety like behavior which is more common in case of females. Therefore, in the present study, we have studied the impact of social isolation of young female mice for 8weeks on the anxiety like behavior and the underlying molecular mechanism. Elevated plus maze and open field test revealed that social isolation caused anxiety like behavior. BDNF, a well-known molecule implicated in the anxiety like behavior, was up-regulated both at the message and protein level in cerebral cortex by social isolation. CREB-1 and CBP, which play a crucial role in BDNF transcription, were up-regulated at mRNA level in cerebral cortex by social isolation. HDAC-2, which negatively regulates BDNF expression, was down-regulated at mRNA and protein level in cerebral cortex by social isolation. Furthermore, BDNF acts in concert with Limk-1, miRNA-132 and miRNA-134 for the regulation of structural and morphological plasticity. Social isolation resulted in up-regulation of Limk-1 mRNA and miRNA-132 expression in the cerebral cortex. MiRNA-134, which inhibits the translation of Limk-1, was decreased in cerebral cortex by social isolation. Taken together, our study suggests that social isolation mediated anxiety like behavior is associated with up-regulation of BDNF expression and concomitant increase in the expression of CBP, CREB-1, Limk-1 and miRNA-132, and decrease in the expression of HDAC-2 and miRNA-134 in the cerebral cortex. Copyright © 2016. Published by Elsevier Inc.

Inositol polyphosphate multikinase is a coactivator for serum response factor-dependent induction of immediate early genes

PubMed Central

Kim, Eunha; Tyagi, Richa; Lee, Joo-Young; Park, Jina; Kim, Young-ran; Beon, Jiyoon; Chen, Po Yu; Cha, Jiyoung Y.; Snyder, Solomon H.; Kim, Seyun

2013-01-01

Inositol polyphosphate multikinase (IPMK) is a notably pleiotropic protein. It displays both inositol phosphate kinase and phosphatidylinositol kinase catalytic activities. Noncatalytically, IPMK stabilizes the mammalian target of rapamycin complex 1 and acts as a transcriptional coactivator for CREB-binding protein/E1A binding protein p300 and tumor suppressor protein p53. Serum response factor (SRF) is a major transcription factor for a wide range of immediate early genes. We report that IPMK, in a noncatalytic role, is a transcriptional coactivator for SRF mediating the transcription of immediate early genes. Stimulation by serum of many immediate early genes is greatly reduced by IPMK deletion. IPMK stimulates expression of these genes, an influence also displayed by catalytically inactive IPMK. IPMK acts by binding directly to SRF and thereby enhancing interactions of SRF with the serum response element of diverse genes. PMID:24248338

GPR30 Regulates Glutamate Transporter GLT-1 Expression in Rat Primary Astrocytes*

PubMed Central

Lee, Eunsook; Sidoryk-Wêgrzynowicz, Marta; Wang, Ning; Webb, Anton; Son, Deok-Soo; Lee, Kyuwon; Aschner, Michael

2012-01-01

The G protein-coupled estrogen receptor GPR30 contributes to the neuroprotective effects of 17β-estradiol (E2); however, the mechanisms associated with this protection have yet to be elucidated. Given that E2 increases astrocytic expression of glutamate transporter-1 (GLT-1), which would prevent excitotoxic-induced neuronal death, we proposed that GPR30 mediates E2 action on GLT-1 expression. To investigate this hypothesis, we examined the effects of G1, a selective agonist of GPR30, and GPR30 siRNA on astrocytic GLT-1 expression, as well as glutamate uptake in rat primary astrocytes, and explored potential signaling pathways linking GPR30 to GLT-1. G1 increased GLT-1 protein and mRNA levels, subject to regulation by both MAPK and PI3K signaling. Inhibition of TGF-α receptor suppressed the G1-induced increase in GLT-1 expression. Silencing GPR30 reduced the expression of both GLT-1 and TGF-α and abrogated the G1-induced increase in GLT-1 expression. Moreover, the G1-induced increase in GLT-1 protein expression was abolished by a protein kinase A inhibitor and an NF-κB inhibitor. G1 also enhanced cAMP response element-binding protein (CREB), as well as both NF-κB p50 and NF-κB p65 binding to the GLT-1 promoter. Finally, to model dysfunction of glutamate transporters, manganese was used, and G1 was found to attenuate manganese-induced impairment in GLT-1 protein expression and glutamate uptake. Taken together, the present data demonstrate that activation of GPR30 increases GLT-1 expression via multiple pathways, suggesting that GPR30 is worthwhile as a potential target to be explored for developing therapeutics of excitotoxic neuronal injury. PMID:22645130

Cellular mechanisms underlying an effect of "early handling" on pCREB and BDNF in the neonatal rat hippocampus.

PubMed

Garoflos, Efstathios; Stamatakis, Antonios; Mantelas, Athanasios; Philippidis, Helen; Stylianopoulou, Fotini

2005-08-09

Early experiences have long-term effects on brain function and behavior. However, the precise mechanisms involved still remain elusive. In an effort to address this issue, we employed the model of "early handling", which is known to affect the ability of the adult organism to respond to stressful stimuli, and determined its effects on hippocampal pCREB and BDNF 2, 4, and 8 h later. 8 h following "handling" on postnatal day 1, there was an increase in pCREB and BDNF positive cells in the hippocampus, a brain area which is a specific target of "handling". On the other hand, vehicle injection resulted in decreased pCREB and BDNF in both handled and non-handled animals 2 and 4 h later. The "handling"-induced increase of pCREB and BDNF was cancelled by inhibition of NMDA, AMPA/kainate, GABA-A, 5-HT1A or 5-HT2A/C receptors, as well as L-type voltage-gated Ca(2+) channels. It thus appears that "early handling" activates these neurotransmitter receptors, leading to increased intracellular Ca(2+), phosphorylation of the transcription factor CREB, and increased BDNF expression. BDNF can then exert its morphogenetic effects and thus "imprint" the effects of "handling" on the brain.

Cytoplasm-predominant Pten associates with increased region-specific brain tyrosine hydroxylase and dopamine D2 receptors in mouse model with autistic traits.

PubMed

He, Xin; Thacker, Stetson; Romigh, Todd; Yu, Qi; Frazier, Thomas W; Eng, Charis

2015-01-01

Autism spectrum disorder (ASD) is a group of neurodevelopmental disorders characterized by impairment in social communication/interaction and inflexible/repetitive behavior. Several lines of evidence support genetic factors as a predominant cause of ASD. Among those autism susceptibility genes that have been identified, the PTEN tumor suppressor gene, initially identified as predisposing to Cowden heritable cancer syndrome, was found to be mutated in a subset of ASD patients with extreme macrocephaly. However, the ASD-relevant molecular mechanism mediating the effect of PTEN mutations remains elusive. We developed a Pten knock-in murine model to study the effects of Pten germline mutations, specifically altering subcellular localization, in ASD. Proteins were isolated from the hemispheres of the male littermates, and Western blots were performed to determine protein expression levels of tyrosine hydroxylase (TH). Immunohistochemical stains were carried out to validate the localization of TH and dopamine D2 receptors (D2R). PC12 cells ectopically expressing either wild-type or missense mutant PTEN were then compared for the differences in TH expression. Mice carrying Pten mutations have high TH and D2R in the striatum and prefrontal cortex. They also have increased phosphorylation of cAMP response element-binding protein (CREB) and TH. Mechanistically, PTEN downregulates TH production in PC12 cells via inhibiting the phosphoinositide 3-kinase (PI3K)/CREB signaling pathway, while PTEN reduces TH phosphorylation via suppressing MAPK pathway. Unlike wild-type PTEN but similar to the mouse knock-in mutant Pten, three naturally occurring missense mutations of PTEN that we previously identified in ASD patients, H93R, F241S, and D252G, were not able to suppress TH when overexpressed in PC12 cells. In addition, two other PTEN missense mutations, C124S (pan phosphatase dead) and G129E (lipid phosphatase dead), failed to suppress TH when ectopically expressed in PC12 cells. Our data reveal a non-canonical PTEN-TH pathway in the brain that may work as a core regulator of dopamine signaling, which when dysfunctional is pathogenic in ASD.

Ssanghwa-tang, an oriental herbal cocktail, exerts anti-melanogenic activity by suppression of the p38 MAPK and PKA signaling pathways in B16F10 cells

PubMed Central

2013-01-01

Background Ssanghwa-tang (SHT) is a widely used medication for the treatment of fatigue, pain, inflammation, hypothermia, erectile dysfunction, cancer, and osteoporosis in Asia, however, role of SHT on the melanin synthesis has not been checked previously. Thus, the present study was designed to determine the effect of SHT on α-melanocyte stimulating hormone (α-MSH)-induced melanogensis and its mechanisms of action in murine B16F10 melanoma cells. Method Cellular melanin content and tyrosinase activity in murine B16F10 melanoma cells were determined after α-MSH stimulation with or without pre-treatment of SHT at the concentration of 250 and 500 μg/ml. Expression level of tyrosinase, tyrosinase-related protein 1 (TRP-1), TRP-2, microphthalmia-associated transcription factor (MITF), and activation of c-AMP-dependent protein kinase (PKA), c-AMP-related element binding protein (CREB), and mitogen-activated protein kinases (MAPKs) were examined by Western blot analysis. Results SHT significantly inhibited α-MSH-induced melanin synthesis and tyrosinase activity, and also decreased α-MSH-induced expression of MITF, tyrosinase, and TRP-1. In addition, SHT remarkably suppressed tyrosinase, CRE, and MITF luciferase reporter activity in a resting state as well as in α-MSH-stimulating condition. Phosphorylation of p38 MAPK by α-MSH stimulation was efficiently blocked by SHT pre-treatment. Moreover, SHT as an herbal cocktail showed synergistic anti-melanogenic effect compared with that of each single constituent herb. Conclusion SHT efficiently inhibited c-AMP-induced melanin synthesis in B16F10 cells via suppression of PKA and p38 MAPK signaling pathways and subsequently decreased the level of CREB phosphorylation, MITF, and melanogenic enzymes. These results indicate that SHT may be useful as herbal medicine for treating hyperpigmentation and cosmetics as a skin-whitening agent. PMID:23981281

v-src induction of the TIS10/PGS2 prostaglandin synthase gene is mediated by an ATF/CRE transcription response element.

PubMed

Xie, W; Fletcher, B S; Andersen, R D; Herschman, H R

1994-10-01

We recently reported the cloning of a mitogen-inducible prostaglandin synthase gene, TIS10/PGS2. In addition to growth factors and tumor promoters, the v-src oncogene induces TIS10/PGS2 expression in 3T3 cells. Deletion analysis, using luciferase reporters, identifies a region between -80 and -40 nucleotides 5' of the TIS10/PGS2 transcription start site that mediates pp60v-src induction in 3T3 cells. This region contains the sequence CGTCACGTG, which includes overlapping ATF/CRE (CGTCA) and E-box (CACGTG) sequences. Gel shift-oligonucleotide competition experiments with nuclear extracts from cells stably transfected with a temperature-sensitive v-src gene demonstrate that the CGTCACGTG sequence can bind proteins at both the ATF/CRE and E-box sequences. Dominant-negative CREB and Myc proteins that bind DNA, but do not transactivate, block v-src induction of a luciferase reporter driven by the first 80 nucleotides of the TIS10/PGS2 promoter. Mutational analysis distinguishes which TIS10/PGS2 cis-acting element mediates pp60v-src induction. E-box mutation has no effect on the fold induction in response to pp60v-src. In contrast, ATF/CRE mutation attenuates the pp60v-src response. Antibody supershift and methylation interference experiments demonstrate that CREB and at least one other ATF transcription factor in these extracts bind to the TIS10/PGS2 ATF/CRE element. Expression of a dominant-negative ras gene also blocks TIS10/PGS2 induction by v-src. Our data suggest that Ras mediates pp60v-src activation of an ATF transcription factor, leading to induced TIS10/PGS2 expression via the ATF/CRE element of the TIS10/PGS2 promoter. This is the first description of v-src activation of gene expression via an ATF/CRE element.

Human T cell lymphotropic virus type I genomic expression and impact on intracellular signaling pathways during neurodegenerative disease and leukemia.

PubMed

Yao, J; Wigdahl, B

2000-01-01

HTLV-I has been identified as the etiologic agent of neoplasia within the human peripheral blood T lymphocyte population, and a progressive neurologic disorder based primarily within the central nervous system. We have examined the role of HTLV-I in these two distinctly different clinical syndromes by examining the life cycle of the virus, with emphasis on the regulation of viral gene expression within relevant target cell populations. In particular, we have examined the impact of specific viral gene products, particularly Tax, on cellular metabolic function. Tax is a highly promiscuous and pleiotropic viral oncoprotein, and is the most important factor contributing to the initial stages of viral-mediated transformation of T cells after HTLV-I infection. Tax, which weakly binds to Tax response element 1 (TRE-1) in the viral long terminal repeat (LTR), can dramatically trans-activate viral gene expression by interacting with cellular transcription factors, such as activated transcription factors and cyclic AMP response element binding proteins (ATF/CREB), CREB binding protein (CBP/p300), and factors involved with the basic transcription apparatus. At the same time, Tax alters cellular gene expression by directly or indirectly interacting with a variety of cellular transcription factors, cell cycle control elements, and cellular signal transduction molecules ultimately resulting in dysregulated cell proliferation. The mechanisms associated with HTLV-I infection, leading to tropical spastic paraparesis (TSP) are not as clearly resolved. Possible explanations of viral-induced neurologic disease range from central nervous system (CNS) damage caused by direct viral invasion of the CNS to bystander CNS damage caused by the immune response to HTLV-I infection. It is interesting to note that it is very rare for an HTLV-I infected individual to develop both adult T cell leukemia (ATL) and TSP in his/her life time, suggesting that the mechanisms governing development of these two diseases are mutually exclusive.

Increasing CREB Function in the CA1 Region of Dorsal Hippocampus Rescues the Spatial Memory Deficits in a Mouse Model of Alzheimer's Disease

PubMed Central

Yiu, Adelaide P; Rashid, Asim J; Josselyn, Sheena A

2011-01-01

The principal defining feature of Alzheimer's disease (AD) is memory impairment. As the transcription factor CREB (cAMP/Ca2+ responsive element-binding protein) is critical for memory formation across species, we investigated the role of CREB in a mouse model of AD. We found that TgCRND8 mice exhibit a profound impairment in the ability to form a spatial memory, a process that critically relies on the dorsal hippocampus. Perhaps contributing to this memory deficit, we observed additional deficits in the dorsal hippocampus of TgCRND8 mice in terms of (1) biochemistry (decreased CREB activation in the CA1 region), (2) neuronal structure (decreased spine density and dendritic complexity of CA1 pyramidal neurons), and (3) neuronal network activity (decreased arc mRNA levels following behavioral training). Locally and acutely increasing CREB function in the CA1 region of dorsal hippocampus of TgCRND8 mice was sufficient to restore function in each of these key domains (biochemistry, neuronal structure, network activity, and most importantly, memory formation). The rescue produced by increasing CREB was specific both anatomically and behaviorally and independent of plaque load or Aβ levels. Interestingly, humans with AD show poor spatial memory/navigation and AD brains have disrupted (1) CREB activation, and (2) spine density and dendritic complexity in hippocampal CA1 pyramidal neurons. These parallel findings not only confirm that TgCRND8 mice accurately model key aspects of human AD, but furthermore, suggest the intriguing possibility that targeting CREB may be a useful therapeutic strategy in treating humans with AD. PMID:21734652

Transcriptional activation of peroxisome proliferator-activated receptor-{gamma} requires activation of both protein kinase A and Akt during adipocyte differentiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Sang-pil; Ha, Jung Min; Yun, Sung Ji

2010-08-13

Research highlights: {yields} Elevated cAMP activates both PKA and Epac. {yields} PKA activates CREB transcriptional factor and Epac activates PI3K/Akt pathway via Rap1. {yields} Akt modulates PPAR-{gamma} transcriptional activity in concert with CREB. -- Abstract: Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is required for the conversion of pre-adipocytes. However, the mechanism underlying activation of PPAR-{gamma} is unclear. Here we showed that cAMP-induced activation of protein kinase A (PKA) and Akt is essential for the transcriptional activation of PPAR-{gamma}. Hormonal induction of adipogenesis was blocked by a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002), by a protein kinase A (PKA) inhibitor (H89), and by amore » Rap1 inhibitor (GGTI-298). Transcriptional activity of PPAR-{gamma} was markedly enhanced by 3-isobutyl-1-methylxanthine (IBMX), but not insulin and dexamethasone. In addition, IBMX-induced PPAR-{gamma} transcriptional activity was blocked by PI3K/Akt, PKA, or Rap1 inhibitors. 8-(4-Chlorophenylthio)-2'-O-methyl-cAMP (8-pCPT-2'-O-Me-cAMP) which is a specific agonist for exchanger protein directly activated by cAMP (Epac) significantly induced the activation of Akt. Furthermore, knock-down of Akt1 markedly attenuated PPAR-{gamma} transcriptional activity. These results indicate that both PKA and Akt signaling pathways are required for transcriptional activation of PPAR-{gamma}, suggesting post-translational activation of PPAR-{gamma} might be critical step for adipogenic gene expression.« less

N-(4-methoxyphenyl) caffeamide-induced melanogenesis inhibition mechanisms.

PubMed

Kuo, Yueh-Hsiung; Chen, Chien-Chia; Wu, Po-Yuan; Wu, Chin-Sheng; Sung, Ping-Jyun; Lin, Chien-Yih; Chiang, Hsiu-Mei

2017-01-23

The derivative of caffeamide exhibits antioxidant and antityrosinase activity. The activity and mechanism of N-(4-methoxyphenyl) caffeamide (K36E) on melanogenesis was investigated. B16F0 cells were treated with various concentrations of K36E; the melanin contents and related signal transduction were studied. Western blotting assay was applied to determine the protein expression, and spectrophotometry was performed to identify the tyrosinase activity and melanin content. Our results indicated that K36E reduced α-melanocyte-stimulating hormone (α-MSH)-induced melanin content and tyrosinase activity in B16F0 cells. In addition, K36E inhibited the expression of phospho-cyclic adenosine monophosphate (cAMP)-response element-binding protein, microphthalmia-associated transcription factor (MITF), tyrosinase, and tyrosinase-related protein-1 (TRP-1). K36E activated the phosphorylation of protein kinase B (AKT) and glycogen synthase kinase 3 beta (GSK3β), leading to the inhibition of MITF transcription activity. K36E attenuated α-MSH induced cAMP pathways, contributing to hypopigmentation. K36E regulated melanin synthesis through reducing the expression of downstream proteins including p-CREB, p-AKT, p-GSK3β, tyrosinase, and TRP-1, and activated the transcription factor, MITF. K36E may have the potential to be developed as a skin whitening agent.

Cytoplasmic Location of α1A Voltage-Gated Calcium Channel C-Terminal Fragment (Cav2.1-CTF) Aggregate Is Sufficient to Cause Cell Death

PubMed Central

Takahashi, Makoto; Obayashi, Masato; Ishiguro, Taro; Sato, Nozomu; Niimi, Yusuke; Ozaki, Kokoro; Mogushi, Kaoru; Mahmut, Yasen; Tanaka, Hiroshi; Tsuruta, Fuminori; Dolmetsch, Ricardo; Yamada, Mitsunori; Takahashi, Hitoshi; Kato, Takeo; Mori, Osamu; Eishi, Yoshinobu; Mizusawa, Hidehiro; Ishikawa, Kinya

2013-01-01

The human α1A voltage-dependent calcium channel (Cav2.1) is a pore-forming essential subunit embedded in the plasma membrane. Its cytoplasmic carboxyl(C)-tail contains a small poly-glutamine (Q) tract, whose length is normally 4∼19 Q, but when expanded up to 20∼33Q, the tract causes an autosomal-dominant neurodegenerative disorder, spinocerebellar ataxia type 6 (SCA6). A recent study has shown that a 75-kDa C-terminal fragment (CTF) containing the polyQ tract remains soluble in normal brains, but becomes insoluble mainly in the cytoplasm with additional localization to the nuclei of human SCA6 Purkinje cells. However, the mechanism by which the CTF aggregation leads to neurodegeneration is completely elusive, particularly whether the CTF exerts more toxicity in the nucleus or in the cytoplasm. We tagged recombinant (r)CTF with either nuclear-localization or nuclear-export signal, created doxycyclin-inducible rat pheochromocytoma (PC12) cell lines, and found that the CTF is more toxic in the cytoplasm than in the nucleus, the observations being more obvious with Q28 (disease range) than with Q13 (normal-length). Surprisingly, the CTF aggregates co-localized both with cAMP response element-binding protein (CREB) and phosphorylated-CREB (p-CREB) in the cytoplasm, and Western blot analysis showed that the quantity of CREB and p-CREB were both decreased in the nucleus when the rCTF formed aggregates in the cytoplasm. In human brains, polyQ aggregates also co-localized with CREB in the cytoplasm of SCA6 Purkinje cells, but not in other conditions. Collectively, the cytoplasmic Cav2.1-CTF aggregates are sufficient to cause cell death, and one of the pathogenic mechanisms may be abnormal CREB trafficking in the cytoplasm and reduced CREB and p-CREB levels in the nuclei. PMID:23505410

Activity-dependent expression of miR-132 regulates immediate-early gene induction during olfactory learning in the greater short-nosed fruit bat, Cynopterus sphinx.

PubMed

Mukilan, Murugan; Ragu Varman, Durairaj; Sudhakar, Sivasubramaniam; Rajan, Koilmani Emmanuvel

2015-04-01

The activity-dependent expression of immediate-early genes (IEGs) and microRNA (miR)-132 has been implicated in synaptic plasticity and the formation of long-term memory (LTM). In the present study, we show that olfactory training induces the expression of IEGs (EGR-1, C-fos, C-jun) and miR-132 at similar time scale in olfactory bulb (OB) of Cynopterus sphinx. We examined the role of miR-132 in the OB using antisense oligodeoxynucleotide (AS-ODN) and demonstrated that a local infusion of AS-ODN in the OB 2h prior to training impaired olfactory memory formation in C. sphinx. However, the infusion of AS-ODN post-training did not cause a deficit in memory formation. Furthermore, the inhibition of miR-132 reduced the olfactory training-induced expression of IEGs and post synaptic density protein-95 (PSD-95) in the OB. Additionally, we show that miR-132 regulates the activation of calcium/calmodulin-dependent protein kinase-II (CaMKII) and cAMP response element binding protein (CREB), possibly through miR-148a. These data suggest that olfactory training induces the expression of miR-132 and IEGs, which in turn activates post-synaptic proteins that regulate olfactory memory formation. Copyright © 2015 Elsevier Inc. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yu, Shu; Cheng, Qiong; Li, Lu

Salidroside is proven to be a neuroprotective agent of natural origin, and its analog, 2-(4-Methoxyphenyl)ethyl-2-acetamido-2-deoxy-β-D-pyranoside (named SalA-4 g), has been synthesized in our lab. In this study, we showed that SalA-4 g promoted neuronal survival and inhibited neuronal apoptosis in primary hippocampal neurons exposed to oxygen and glucose deprivation (OGD) and in rats subjected to ischemia by transient middle cerebral artery occlusion (MCAO), respectively, and that SalA-4 g was more neuroprotective than salidroside. We further found that SalA-4 g elevated glucose uptake in OGD-injured primary hippocampal neurons and increased the expression and recruitment of glucose transporter 3 (GLUT3) in ischemicmore » brain. Signaling analysis revealed that SalA-4 g triggered the phosphorylation of CREB, and increased the expression of PKA RII in primary hippocampal neurons exposed to OGD injury, while inhibition of PKA/CREB by H-89 alleviated the elevation in glucose uptake and GLUT3 expression, and blocked the protective effects of SalA-4 g. Moreover, SalA-4 g was noted to inhibit intracellular Ca{sup 2+} influx and calpain1 activation in OGD-injured primary hippocampal neurons. Our results suggest that SalA-4 g neuroprotection might be mediated by increased glucose uptake and elevated GLUT3 expression through calpain1/PKA/CREB pathway. - Highlights: • A salidroside (Sal) analog (SalA-4 g) is prepared to be more neuroprotective than Sal. • SalA-4 g protected hippocampal neurons from oxygen and glucose deprivation insult. • SalA-4 g reduced ischemic injury after transient middle cerebral artery occlusion in rats. • Neuroprotection of SalA-4 g was mediated by GLUT3 level via calpain/PKA/CREB pathway.« less

Glucosensing capacity of rainbow trout telencephalon.

PubMed

Otero-Rodiño, C; Rocha, A; Álvarez-Otero, R; Ceinos, R M; López-Patiño, M A; Míguez, J M; Cerdá-Reverter, J M; Soengas, J L

2018-03-01

To assess the hypothesis of glucosensing systems present in fish telencephalon, we first demonstrated in rainbow trout, by in situ hybridisation, the presence of glucokinase (GK). Then, we assessed the response of glucosensing markers in rainbow trout telencephalon 6 hours after i.c.v. treatment with glucose or 2-deoxyglucose (inducing glucoprivation). We evaluated the response of parameters related to the mechanisms dependent on GK, liver X receptor (LXR), mitochondrial activity, sweet taste receptor and sodium-glucose linked transporter 1 (SGLT-1). We also assessed mRNA abundance of neuropeptides involved in the metabolic control of food intake (agouti-related protein, neuropeptide Y, pro-opiomelanocortin, and cocaine- and amphetamine-related transcript), as well as the abundance and phosphorylation status of proteins possibly involved in linking glucosensing with neuropeptide expression, such as protein kinase B (AkT), AMP-activated protein kinase (AMPK), mechanistic target of rapamycin and cAMP response element-binding protein (CREB). The responses obtained support the presence in the telencephalon of a glucosensing mechanism based on GK and maybe one based on LXR, although they do not support the presence of mechanisms dependent on mitochondrial activity and SGLT-1. The mechanism based on sweet taste receptor responded to glucose but in a converse way to that characterised previously in the hypothalamus. In general, systems responded only to glucose but not to glucoprivation. Neuropeptides did not respond to glucose or glucoprivation. By contrast, the presence of glucose activates Akt and inhibits AMPK, CREB and forkhead box01. This is the first study in any vertebrate species in which the response to glucose of putative glucosensing mechanisms is demonstrated in the telencephalon. Their role might relate to processes other than homeostatic control of food intake, such as the hedonic and reward system. © 2018 British Society for Neuroendocrinology.

LKB1 tumor suppressor and salt-inducible kinases negatively regulate human T-cell leukemia virus type 1 transcription

PubMed Central

2013-01-01

Background Human T-cell leukemia virus type 1 (HTLV-1) causes adult T-cell leukemia (ATL). Treatment options are limited and prophylactic agents are not available. We have previously demonstrated an essential role for CREB-regulating transcriptional coactivators (CRTCs) in HTLV-1 transcription. Results In this study we report on the negative regulatory role of LKB1 tumor suppressor and salt-inducible kinases (SIKs) in the activation of HTLV-1 long terminal repeats (LTR) by the oncoprotein Tax. Activation of LKB1 and SIKs effectively blunted Tax activity in a phosphorylation-dependent manner, whereas compromising these kinases, but not AMP-dependent protein kinases, augmented Tax function. Activated LKB1 and SIKs associated with Tax and suppressed Tax-induced LTR activation by counteracting CRTCs and CREB. Enforced expression of LKB1 or SIK1 in cells transfected with HTLV-1 molecular clone pX1MT repressed proviral transcription. On the contrary, depletion of LKB1 in pX1MT-transfected cells and in HTLV-1-transformed T cells boosted the expression of Tax. Treatment of HTLV-1 transformed cells with metformin led to LKB1/SIK1 activation, reduction in Tax expression, and inhibition of cell proliferation. Conclusions Our findings revealed a new function of LKB1 and SIKs as negative regulators of HTLV-1 transcription. Pharmaceutical activation of LKB1 and SIKs might be considered as a new strategy in anti-HTLV-1 and anti-ATL therapy. PMID:23577667

Impairment of the spatial learning and memory induced by learned helplessness and chronic mild stress.

PubMed

Song, Li; Che, Wang; Min-Wei, Wang; Murakami, Yukihisa; Matsumoto, Kinzo

2006-02-01

Increasing evidences indicate the concurrence and interrelationship of depression and cognitive impairments. The present study was undertaken to investigate the effects of two depressive animal models, learned helplessness (LH) and chronic mild stress (CMS), on the cognitive functions of mice in the Morris water maze task. Our results demonstrated that both LH and CMS significantly decreased the cognitive performance of stressed mice in the water maze task. The escaping latency to the platform was prolonged and the probe test percentage in the platform quadrant was reduced. These two models also increased the plasma corticosterone concentration and decreased the brain derived neurotrophic factor (BDNF) and cAMP-response element-biding protein (CREB) messenger ribonucleic acid (mRNA) levels in hippocampus, which might cause the spatial cognition deficits. Repeated treatment with antidepressant drugs, imipramine (Imi) and fluoxetine (Flu), significantly reduced the plasma corticosterone concentration and enhanced the BDNF and CREB levels. Furthermore, antidepressant treated animals showed an ameliorated cognitive performance compared with the vehicle treated stressed animals. These data suggest that both LH and CMS impair the spatial cognitive function and repeated treatment with antidepressant drugs decreases the prevalence of cognitive impairments induced by these two animal models. Those might in part be attributed to the reduced plasma corticosterone and enhanced hippocampal BDNF and CREB expressions. This study provided a better understanding of molecular mechanisms underlying interactions of depression and cognitive impairments, although animal models used in this study can mimic only some aspects of depression or cognition of human.

Comparative analysis of cis-regulation following stroke and seizures in subspaces of conserved eigensystems

PubMed Central

2010-01-01

Background It is often desirable to separate effects of different regulators on gene expression, or to identify effects of the same regulator across several systems. Here, we focus on the rat brain following stroke or seizures, and demonstrate how the two tasks can be approached simultaneously. Results We applied SVD to time-series gene expression datasets from the rat experimental models of stroke and seizures. We demonstrate conservation of two eigensystems, reflecting inflammation and/or apoptosis (eigensystem 2) and neuronal synaptic activity (eigensystem 3), between the stroke and seizures. We analyzed cis-regulation of gene expression in the subspaces of the conserved eigensystems. Bayesian networks analysis was performed separately for either experimental model, with cross-system validation of the highest-ranking features. In this way, we correctly re-discovered the role of AP1 in the regulation of apoptosis, and the involvement of Creb and Egr in the regulation of synaptic activity-related genes. We identified a novel antagonistic effect of the motif recognized by the nuclear matrix attachment region-binding protein Satb1 on AP1-driven transcriptional activation, suggesting a link between chromatin loop structure and gene activation by AP1. The effects of motifs binding Satb1 and Creb on gene expression in brain conform to the assumption of the linear response model of gene regulation. Our data also suggest that numerous enhancers of neuronal-specific genes are important for their responsiveness to the synaptic activity. Conclusion Eigensystems conserved between stroke and seizures separate effects of inflammation/apoptosis and neuronal synaptic activity, exerted by different transcription factors, on gene expression in rat brain. PMID:20565733

Enhanced wound healing of tissue-engineered human corneas through altered phosphorylation of the CREB and AKT signal transduction pathways.

PubMed

Couture, Camille; Desjardins, Pascale; Zaniolo, Karine; Germain, Lucie; Guérin, Sylvain L

2018-06-01

The cornea is a transparent organ, highly specialized and unique that is continually subjected to abrasive forces and occasional mechanical or chemical trauma because of its anatomical localization. Upon injury, the extracellular matrix (ECM) rapidly changes to promote wound healing through integrin-dependent activation of specific signal transduction mediators whose contribution is to favor faster closure of the wound by altering the adhesive and migratory properties of the cells surrounding the damaged area. In this study, we exploited the human tissue-engineered cornea (hTECs) as a model to study the signal transduction pathways that participate to corneal wound healing. By exploiting both gene profiling and activated kinases arrays, we could demonstrate the occurrence of important alterations in the level of expression and activation of a few mediators from the PI3K/Akt and CREB pathways in response to the ECM remodeling taking place during wound healing of damaged hTECs. Pharmacological inhibition of CREB with C646 considerably accelerated wound closure compared to controls. This process was considerably accelerated further when both C646 and SC79, an Akt agonist, were added together to wounded hTECs. Therefore, our study demonstrate that proper corneal wound healing requires the activation of Akt together with the inhibition of CREB and that wound healing in vitro can be altered by the use of pharmacological inhibitors (such as C646) or agonists (such as SC79) of these mediators. Corneal wounds account for a large proportion of all visual disabilities in North America. To our knowledge, this is the first time that a tissue-engineered human cornea (hTEC) entirely produced using normal untransformed human cells is used as a biomaterial to study the signal transduction pathways that are critical to corneal wound healing. Through the use of this biomaterial, we demonstrated that human corneal epithelial cells engaged in wound healing reduce phosphorylation of the signal transduction mediator CREB while, in the mean time, they increase that of AKT. By increasing the activation of AKT together with a decrease in CREB activation, we could considerably reduce wound closure time in our punch-damaged hTECs. Considering the increasing interest given to the reconstruction of different types of tissues, we believe these results will have a strong impact on the field of tissue-engineering and biomaterials. Altering the activation status of the Akt and CREB proteins might prove to be a therapeutically interesting avenue and may also find applications in wound healing of other tissues beside the cornea, such as the skin. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Increased Expression of Brain-Derived Neurotrophic Factor Transcripts I and VI, cAMP Response Element Binding, and Glucocorticoid Receptor in the Cortex of Patients with Temporal Lobe Epilepsy.

PubMed

Martínez-Levy, G A; Rocha, L; Rodríguez-Pineda, F; Alonso-Vanegas, M A; Nani, A; Buentello-García, R M; Briones-Velasco, M; San-Juan, D; Cienfuegos, J; Cruz-Fuentes, C S

2018-05-01

A body of evidence supports a relevant role of brain-derived neurotrophic factor (BDNF) in temporal lobe epilepsy (TLE). Magnetic resonance data reveal that the cerebral atrophy extends to regions that are functionally and anatomically connected with the hippocampus, especially the temporal cortex. We previously reported an increased expression of BDNF messenger for the exon VI in the hippocampus of temporal lobe epilepsy patients compared to an autopsy control group. Altered levels of this particular transcript were also associated with pre-surgical use of certain psychotropic. We extended here our analysis of transcripts I, II, IV, and VI to the temporal cortex since this cerebral region holds intrinsic communication with the hippocampus and is structurally affected in patients with TLE. We also assayed the cyclic adenosine monophosphate response element-binding (CREB) and glucocorticoid receptor (GR) genes as there is experimental evidence of changes in their expression associated with BDNF and epilepsy. TLE and pre-surgical pharmacological treatment were considered as the primary clinical independent variables. Transcripts BDNF I and BDNF VI increased in the temporal cortex of patients with pharmacoresistant TLE. The expression of CREB and GR expression follow the same direction. Pre-surgical use of selective serotonin reuptake inhibitors, carbamazepine (CBZ) and valproate (VPA), was associated with the differential expression of specific BDNF transcripts and CREB and GR genes. These changes could have functional implication in the plasticity mechanisms related to temporal lobe epilepsy.

Human T-cell leukemia virus type 1 Tax oncoprotein represses the expression of the BCL11B tumor suppressor in T-cells.

PubMed

Takachi, Takayuki; Takahashi, Masahiko; Takahashi-Yoshita, Manami; Higuchi, Masaya; Obata, Miki; Mishima, Yukio; Okuda, Shujiro; Tanaka, Yuetsu; Matsuoka, Masao; Saitoh, Akihiko; Green, Patrick L; Fujii, Masahiro

2015-04-01

Human T-cell leukemia virus type 1 (HTLV-1) is the etiological agent of adult T cell leukemia (ATL), which is an aggressive form of T-cell malignancy. HTLV-1 oncoproteins, Tax and HBZ, play crucial roles in the immortalization of T-cells and/or leukemogenesis by dysregulating the cellular functions in the host. Recent studies show that HTLV-1-infected T-cells have reduced expression of the BCL11B tumor suppressor protein. In the present study, we explored whether Tax and/or HBZ play a role in downregulating BCL11B in HTLV-1-infected T-cells. Lentiviral transduction of Tax in a human T-cell line repressed the expression of BCL11B at both the protein and mRNA levels, whereas the transduction of HBZ had little effect on the expression. Tax mutants with a decreased activity for the NF-κB, CREB or PDZ protein pathways still showed a reduced expression of the BCL11B protein, thereby implicating a different function of Tax in BCL11B downregulation. In addition, the HTLV-2 Tax2 protein reduced the BCL11B protein expression in T-cells. Seven HTLV-1-infected T-cell lines, including three ATL-derived cell lines, showed reduced BCL11B mRNA and protein expression relative to an uninfected T-cell line, and the greatest reductions were in the cells expressing Tax. Collectively, these results indicate that Tax is responsible for suppressing BCL11B protein expression in HTLV-1-infected T-cells; Tax-mediated repression of BCL11B is another mechanism that Tax uses to promote oncogenesis of HTLV-1-infected T-cells. © 2015 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

Evidence of Aβ- and transgene-dependent defects in ERK-CREB signaling in Alzheimer’s models

PubMed Central

Ma, Qiu-Lan; Harris-White, Marni E.; Ubeda, Oliver J.; Simmons, Mychica; Beech, Walter; Lim, Giselle P.; Teter, Bruce; Frautschy, Sally A.; Cole, Greg M.

2008-01-01

Extracellular-signal regulated kinase (ERK) signaling is critical for memory and tightly regulated by acute environmental stimuli. In Alzheimer disease transgenic models, active ERK is shown to first be increased, then later reduced, but whether these baseline changes reflect disruptions in ERK signaling is less clear. We investigated the influence of the familial Alzheimer’s disease transgene APPsw and β-amyloid peptide (Aβ) immunoneutralization on cannulation injury-associated (i.c.v. infusion) ERK activation. At both 12 and 22 months of age, the trauma-associated activation of ERK observed in Tg− mice was dramatically attenuated in Tg+. In cortices of 22-month-old non-infused mice, a reduction in ERK activation was observed in Tg+, relative to Tg− mice. Intracerebroventricular (i.c.v.) anti-Aβ infusion significantly increased phosphorylated ERK, its substrate cAMP-response element-binding protein (CREB) and a downstream target, the NMDA receptor subunit. We also demonstrated that Aβ oligomer decreased active ERK and subsequently active CREB in human neuroblastoma cells, which could be prevented by oligomer immunoneutralization. Aβ oligomers also inhibited active ERK and CREB in primary neurons, in addition to reducing the downstream post-synaptic protein NMDA receptor subunit. These effects were reversed by anti-oligomer. Our data strongly support the existence of an APPsw transgene-dependent and Aβ oligomer-mediated defect in regulation of ERK activation. PMID:17760871

A mouse model of Rubinstein-Taybi syndrome: Defective long-term memory is ameliorated by inhibitors of phosphodiesterase 4

PubMed Central

Bourtchouladze, Rusiko; Lidge, Regina; Catapano, Ray; Stanley, Jennifer; Gossweiler, Scott; Romashko, Darlene; Scott, Rod; Tully, Tim

2003-01-01

Mice carrying a truncated form of cAMP-responsive element binding protein (CREB)-binding protein (CBP) show several developmental abnormalities similar to patients with Rubinstein-Taybi syndrome (RTS). RTS patients suffer from mental retardation, whereas long-term memory formation is defective in mutant CBP mice. A critical role for cAMP signaling during CREB-dependent long-term memory formation appears to be evolutionarily conserved. From this observation, we reasoned that drugs that modulate CREB function by enhancing cAMP signaling might yield an effective treatment for the memory defect(s) of CBP+/− mice. To this end, we designed a cell-based drug screen and discovered inhibitors of phosphodiesterase 4 (PDE4) to be particularly effective enhancers of CREB function. We extend previous behavioral observations by showing that CBP+/− mutants have impaired long-term memory but normal learning and short-term memory in an object recognition task. We demonstrate that the prototypical PDE4 inhibitor, rolipram, and a novel one (HT0712) abolish the long-term memory defect of CBP+/− mice. Importantly, the genetic lesion in CBP acts specifically to shift the dose sensitivity for HT0712 to enhance memory formation, which conveys molecular specificity on the drug's mechanism of action. Our results suggest that PDE4 inhibitors may be used to treat the cognitive dysfunction of RTS patients. PMID:12930888

CRTC2 and Nedd4 ligase involvement in FSH and TGFβ1 upregulation of connexin43 gap junction.

PubMed

Fang, Wei-Ling; Lai, Si-Yi; Lai, Wei-An; Lee, Ming-Ting; Liao, Ching-Fong; Ke, Ferng-Chun; Hwang, Jiuan-Jiuan

2015-12-01

The major mission of the ovarian follicle is the timely production of the mature fertilizable oocyte, and this is achieved by gonadotropin-regulated, gap junction-mediated cell-cell communication between the oocyte and surrounding nurturing granulosa cells. We have demonstrated that FSH and transforming growth factor beta 1 (TGFβ1) stimulate Gja1 gene-encoded connexin43 (Cx43) gap junction formation/function in rat ovarian granulosa cells is important for their induction of steroidogenesis; additionally, cAMP-protein kinase A (PKA)- and calcium-calcineurin-sensitive cAMP response element-binding (CREB) coactivator CRTC2 plays a crucial role during steroidogenesis. This study was to explore the potential molecular mechanism whereby FSH and TGFβ1 regulate Cx43 synthesis and degradation, particularly the involvement of CRTC2 and ubiquitin ligase Nedd4. Primary culture of granulosa cells from ovarian antral follicles of gonadotropin-primed immature rats was used. At 48 h post-treatment, FSH plus TGFβ1 increased Cx43 level and gap junction function in a PKA- and calcineurin-dependent manner, and TGFβ1 acting through its type I receptor modulated FSH action. Chromatin-immunoprecipitation analysis reveals FSH induced an early-phase (45 min) and FSH+TGFβ1 further elicited a late-phase (24 h) increase in CRTC2, CREB and CBP binding to the Gja1 promoter. Additionally, FSH+TGFβ1 increased the half-life of hyper-phosphorylated Cx43 (Cx43-P2). Also, the proteasome inhibitor MG132 prevented the brefeldin A (blocker of protein transport through Golgi)-reduced Cx43-P2 level and membrane Cx43 gap junction plaque. This is associated with FSH+TGFβ1-attenuated Cx43 interaction with Nedd4 and Cx43 ubiquitination. In all, this study uncovers that FSH and TGFβ1 upregulation of Cx43 gap junctions in ovarian granulosa cells critically involves enhancing CRTC2/CREB/CBP-mediated Cx43 expression and attenuating ubiquitin ligase Nedd4-mediated proteosomal degradation of Cx43 protein. © 2015 Society for Endocrinology.

Calcineurin signaling and PGC-1alpha expression are suppressed during muscle atrophy due to diabetes.

PubMed

Roberts-Wilson, Tiffany K; Reddy, Ramesh N; Bailey, James L; Zheng, Bin; Ordas, Ronald; Gooch, Jennifer L; Price, S Russ

2010-08-01

PGC-1alpha is a transcriptional coactivator that controls energy homeostasis through regulation of glucose and oxidative metabolism. Both PGC-1alpha expression and oxidative capacity are decreased in skeletal muscle of patients and animals undergoing atrophy, suggesting that PGC-1alpha participates in the regulation of muscle mass. PGC-1alpha gene expression is controlled by calcium- and cAMP-sensitive pathways. However, the mechanism regulating PGC-1alpha in skeletal muscle during atrophy remains unclear. Therefore, we examined the mechanism responsible for decreased PGC-1alpha expression using a rodent streptozotocin (STZ) model of chronic diabetes and atrophy. After 21days, the levels of PGC-1alpha protein and mRNA were decreased. We examined the activation state of CREB, a potent activator of PGC-1alpha transcription, and found that phospho-CREB was paradoxically high in muscle of STZ-rats, suggesting that the cAMP pathway was not involved in PGC-1alpha regulation. In contrast, expression of calcineurin (Cn), a calcium-dependent phosphatase, was suppressed in the same muscles. PGC-1alpha expression is regulated by two Cn substrates, MEF2 and NFATc. Therefore, we examined MEF2 and NFATc activity in muscles from STZ-rats. Target genes MRF4 and MCIP1.4 mRNAs were both significantly reduced, consistent with reduced Cn signaling. Moreover, levels of MRF4, MCIP1.4, and PGC-1alpha were also decreased in muscles of CnAalpha-/- and CnAbeta-/- mice without diabetes indicating that decreased Cn signaling, rather than changes in other calcium- or cAMP-sensitive pathways, were responsible for decreased PGC-1alpha expression. These findings demonstrate that Cn activity is a major determinant of PGC-1alpha expression in skeletal muscle during diabetes and possibly other conditions associated with loss of muscle mass.

Calcineurin signaling and PGC-1α expression are suppressed during muscle atrophy due to diabetes

PubMed Central

Roberts-Wilson, Tiffany K.; Reddy, Ramesh N.; Bailey, James L.; Zheng, Bin; Ordas, Ronald; Gooch, Jennifer L.; Price, S. Russ

2010-01-01

PGC-1α is a transcriptional coactivator that controls energy homeostasis through regulation of glucose and oxidative metabolism. Both PGC-1α expression and oxidative capacity are decreased in skeletal muscle of patients and animals undergoing atrophy, suggesting that PGC-1α participates in the regulation of muscle mass. PGC-1α gene expression is controlled by calcium- and cAMP-sensitive pathways. However, the mechanism regulating PGC-1α in skeletal muscle during atrophy remains unclear. Therefore, we examined the mechanism responsible for decreased PGC-1α expression using a rodent streptozotocin (STZ) model of chronic diabetes and atrophy. After 21d, the levels of PGC-1α protein and mRNA were decreased. We examined the activation state of CREB, a potent activator of PGC-1α transcription, and found that phospho-CREB was paradoxically high in muscle of STZ-rats, suggesting that the cAMP pathway was not involved in PGC-1α regulation. In contrast, expression of calcineurin (Cn), a calcium-dependent phosphatase, was suppressed in the same muscles. PGC-1α expression is regulated by two Cn substrates, MEF2 and NFATc. Therefore, we examined MEF2 and NFATc activity in muscles from STZ-rats. Target genes MRF4 and MCIP1.4 were both significantly reduced, consistent with reduced Cn signaling. Moreover, levels of MRF4, MCIP1.4, and PGC-1α were also decreased in muscles of CnAα-/- and CnAβ-/- mice without diabetes indicating that decreased Cn signaling, rather than changes in other calcium- or cAMP-sensitive pathways, were responsible for decreased PGC-1α expression. These findings demonstrate that Cn activity is a major determinant of PGC-1α expression in skeletal muscle during diabetes and possibly other conditions associated with loss of muscle mass. PMID:20359506

CREB binding protein is required for both short-term and long-term memory formation.

PubMed

Chen, Guiquan; Zou, Xiaoyan; Watanabe, Hirotaka; van Deursen, Jan M; Shen, Jie

2010-09-29

CREB binding protein (CBP) is a transcriptional coactivator with histone acetyltransferase activity. Our prior study suggested that CBP might be a key target of presenilins in the regulation of memory formation and neuronal survival. To elucidate the role of CBP in the adult brain, we generated conditional knock-out (cKO) mice in which CBP is completely inactivated in excitatory neurons of the postnatal forebrain. Histological analysis revealed normal neuronal morphology and absence of age-dependent neuronal degeneration in the CBP cKO cerebral cortex. CBP cKO mice exhibited robust impairment in the formation of spatial, associative, and object-recognition memory. In addition to impaired long-term memory, CBP cKO mice also displayed deficits in short-term associative and object-recognition memory. Administration of a histone deacetylase inhibitor, trichostatin A, rescued the reduction of acetylated histones in the CBP cKO cortex but failed to rescue either short- or long-term memory deficits, suggesting that the memory impairment may not be caused by general reduction of histone acetyltransferase activity in CBP cKO mice. Further microarray and Western analysis showed decreased expression of calcium-calmodulin-dependent kinase isoforms and NMDA and AMPA receptor subunits in the cerebral cortex of CBP cKO mice. Collectively, these findings suggest a crucial role for CBP in the formation of both short- and long-term memory.

Chronic administration of aripiprazole activates GSK3β-dependent signalling pathways, and up-regulates GABAA receptor expression and CREB1 activity in rats.

PubMed

Pan, Bo; Huang, Xu-Feng; Deng, Chao

2016-07-20

Aripiprazole is a D2-like receptor (D2R) partial agonist with a favourable clinical profile. Previous investigations indicated that acute and short-term administration of aripiprazole had effects on PKA activity, GSK3β-dependent pathways, GABAA receptors, NMDA receptor and CREB1 in the brain. Since antipsychotics are used chronically in clinics, the present study investigated the long-term effects of chronic oral aripiprazole treatment on these cellular signalling pathways, in comparison with haloperidol (a D2R antagonist) and bifeprunox (a potent D2R partial agonist). We found that the Akt-GSK3β pathway was activated by aripiprazole and bifeprunox in the prefrontal cortex; NMDA NR2A levels were reduced by aripiprazole and haloperidol. In the nucleus accumbens, all three drugs increased Akt-GSK3β signalling; in addition, both aripiprazole and haloperidol, but not bifeprunox, increased the expression of Dvl-3, β-catenin and GABAA receptors, NMDA receptor subunits, as well as CREB1 phosphorylation levels. The results suggest that chronic oral administration of aripiprazole affects schizophrenia-related cellular signalling pathways and markers (including Akt-GSK3β signalling, Dvl-GSK3β-β-catenin signalling, GABAA receptor, NMDA receptor and CREB1) in a brain-region-dependent manner; the selective effects of aripiprazole on these signalling pathways might be associated with its unique clinical effects.

Involvement of AMPA/Kainate Glutamate Receptor in the Extinction and Reinstatement of Morphine-Induced Conditioned Place Preference: A Behavioral and Molecular Study.

PubMed

Siahposht-Khachaki, Ali; Fatahi, Zahra; Yans, Asal; Khodagholi, Fariba; Haghparast, Abbas

2017-03-01

Glutamate receptors in mesolimbic areas such as the nucleus accumbens, ventral tegmental area, prefrontal cortex (PFC), and hippocampus (HIP) are a component of the mechanisms of drug-induced reward and can modulate the firing pattern of dopaminergic neurons in the reward system. In addition, several lines of study have indicated that cAMP response element-binding protein (CREB) and c-fos have important role in morphine-induced conditioned place preference (CPP) induced by drugs of abuse, such as morphine, cocaine, nicotine, and alcohol. Therefore, in the present study, we investigated the changes in phosphorylated CREB (p-CREB) and c-fos induction within the nucleus accumbens (NAc), HIP, and PFC after intracerebroventricular (ICV) administration of different doses of CNQX or vehicle during extinction period or reinstatement of morphine-induced CPP. In all groups, the CPP procedure was done; afterward, the conditioning scores were recorded by Ethovision software. After behavioral test recording, we dissected out the NAc, HIP, and PFC regions and measured the p-CREB/CREB ratio and c-fos level by Western blot analysis. Our results showed that administration of CNQX significantly shortened the extinction of morphine CPP. Besides, ICV microinjection of CNQX following extinction period decreased the reinstatement of morphine CPP in extinguished rats. In molecular section, in treatment group, all mentioned factors were dose-dependently decreased in comparison with vehicle group (DMSO) after ICV microinjection of different doses of CNQX but not in pre-extinction microinjection. These findings suggested that antagonism of AMPA receptor decreased p-CREB/CREB ratio and c-fos level in the PFC, NAc, and HIP. Modulation of the drug memory reconsolidation may be useful for faster extinction of drug-induced reward and attenuation of drug-seeking behavior.

Theobromine up-regulates cerebral brain-derived neurotrophic factor and facilitates motor learning in mice.

PubMed

Yoneda, Mitsugu; Sugimoto, Naotoshi; Katakura, Masanori; Matsuzaki, Kentaro; Tanigami, Hayate; Yachie, Akihiro; Ohno-Shosaku, Takako; Shido, Osamu

2017-01-01

Theobromine, which is a caffeine derivative, is the primary methylxanthine produced by Theobroma cacao. Theobromine works as a phosphodiesterase (PDE) inhibitor to increase intracellular cyclic adenosine monophosphate (cAMP). cAMP activates the cAMP-response element-binding protein (CREB), which is involved in a large variety of brain processes, including the induction of the brain-derived neurotrophic factor (BDNF). BDNF supports cell survival and neuronal functions, including learning and memory. Thus, cAMP/CREB/BDNF pathways play an important role in learning and memory. Here, we investigated whether orally administered theobromine could act as a PDE inhibitor centrally and affect cAMP/CREB/BDNF pathways and learning behavior in mice. The mice were divided into two groups. The control group (CN) was fed a normal diet, whereas the theobromine group (TB) was fed a diet supplemented with 0.05% theobromine for 30 days. We measured the levels of theobromine, phosphorylated vasodilator-stimulated phosphoprotein (p-VASP), phosphorylated CREB (p-CREB), and BDNF in the brain. p-VASP was used as an index of cAMP increases. Moreover, we analyzed the performance of the mice on a three-lever motor learning task. Theobromine was detectable in the brains of TB mice. The brain levels of p-VASP, p-CREB, and BDNF were higher in the TB mice compared with those in the CN mice. In addition, the TB mice performed better on the three-lever task than the CN mice did. These results strongly suggested that orally administered theobromine acted as a PDE inhibitor in the brain, and it augmented the cAMP/CREB/BDNF pathways and motor learning in mice. Copyright © 2016 Elsevier Inc. All rights reserved.

Adenylyl cyclase 3/adenylyl cyclase-associated protein 1 (CAP1) complex mediates the anti-migratory effect of forskolin in pancreatic cancer cells.

PubMed

Quinn, Sierra N; Graves, Sarai H; Dains-McGahee, Clayton; Friedman, Emilee M; Hassan, Humma; Witkowski, Piotr; Sabbatini, Maria E

2017-04-01

Pancreatic cancer is one of the most lethal human malignancies. A better understanding of the intracellular mechanism of migration and invasion is urgently needed to develop treatment that will suppress metastases and improve overall survival. Cyclic adenosine monophosphate (cyclic AMP) is a second messenger that has shown to regulate migration and invasion of pancreatic cancer cells. The rise of cyclic AMP suppressed migration and invasion of pancreatic ductal adenocarcinoma cells. Cyclic AMP is formed from cytosolic ATP by the enzyme adenylyl cyclase (AC). There are ten isoforms of ACs; nine are anchored in the plasma membrane and one is soluble. What remains unknown is the extent to which the expression of transmembrane AC isoforms is both modified in pancreatic cancer and mediates the inhibitory effect of forskolin on cell motility. Using real-time PCR analysis, ADCY3 was found to be highly expressed in pancreatic tumor tissues, resulting in a constitutive increase in cyclic AMP levels. On the other hand, ADCY2 was down-regulated. Migration, invasion, and filopodia formation in two different pancreatic adenocarcinoma cell lines, HPAC and PANC-1 deficient in AC1 or AC3, were studied. We found that AC3, upon stimulation with forskolin, enhanced cyclic AMP levels and inhibited cell migration and invasion. Unlikely to be due to a cytotoxic effect, the inhibitory effects of forskolin involved the quick formation of AC3/adenylyl cyclase-associated protein 1 (CAP1)/G-actin complex, which inhibited filopodia formation and cell motility. Using Western blotting analysis, forskolin, through AC3 activation, caused phosphorylation of CREB, but not ERK. The effect of CREB phosphorylation is likely to be associated with long-term signaling changes. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Transcriptional and epigenetic substrates of methamphetamine addiction and withdrawal: evidence from a long-access self-administration model in the rat.

PubMed

Cadet, Jean Lud; Brannock, Christie; Jayanthi, Subramaniam; Krasnova, Irina N

2015-04-01

Methamphetamine use disorder is a chronic neuropsychiatric disorder characterized by recurrent binge episodes, intervals of abstinence, and relapses to drug use. Humans addicted to methamphetamine experience various degrees of cognitive deficits and other neurological abnormalities that complicate their activities of daily living and their participation in treatment programs. Importantly, models of methamphetamine addiction in rodents have shown that animals will readily learn to give themselves methamphetamine. Rats also accelerate their intake over time. Microarray studies have also shown that methamphetamine taking is associated with major transcriptional changes in the striatum measured within a short or longer time after cessation of drug taking. After a 2-h withdrawal time, there was increased expression of genes that participate in transcription regulation. These included cyclic AMP response element binding (CREB), ETS domain-containing protein (ELK1), and members of the FOS family of transcription factors. Other genes of interest include brain-derived neurotrophic factor (BDNF), tyrosine kinase receptor, type 2 (TrkB), and synaptophysin. Methamphetamine-induced transcription was found to be regulated via phosphorylated CREB-dependent events. After a 30-day withdrawal from methamphetamine self-administration, however, there was mostly decreased expression of transcription factors including junD. There was also downregulation of genes whose protein products are constituents of chromatin-remodeling complexes. Altogether, these genome-wide results show that methamphetamine abuse might be associated with altered regulation of a diversity of gene networks that impact cellular and synaptic functions. These transcriptional changes might serve as triggers for the neuropsychiatric presentations of humans who abuse this drug. Better understanding of the way that gene products interact to cause methamphetamine addiction will help to develop better pharmacological treatment of methamphetamine addicts.

Spinal IL-33/ST2 Signaling Contributes to Neuropathic Pain via Neuronal CaMKII-CREB and Astroglial JAK2-STAT3 Cascades in Mice.

PubMed

Liu, Shenbin; Mi, Wen-Li; Li, Qian; Zhang, Meng-Ting; Han, Ping; Hu, Shan; Mao-Ying, Qi-Liang; Wang, Yan-Qing

2015-11-01

Emerging evidence indicates that nerve damage-initiated neuroinflammation and immune responses, which are evidenced by the up-regulation of proinflammatory cytokines, contribute to the development of neuropathic pain. This study investigated the role of spinal interleukin (IL)-33 and its receptor ST2 in spared nerve injury (SNI)-induced neuropathic pain. The von Frey test and acetone test were performed to evaluate neuropathic pain behaviors (n = 8 to 12), and Western blot (n = 4 to 6), immunohistochemistry, real-time polymerase chain reaction (n = 5), and Bio-Plex (n = 5) assays were performed to understand the molecular mechanisms. Intrathecal administration of ST2-neutralizing antibody or ST2 gene knockout (ST2) significantly attenuated the SNI-induced mechanical and cold allodynia. On the 7th day after SNI, the expression of spinal IL-33 and ST2 was increased by 255.8 ± 27.3% and 266.4 ± 83.5% (mean ± SD), respectively. Mechanistic studies showed that the increased expression of the spinal N-methyl-D-aspartate (NMDA) receptor subunit 1 after SNI was reduced by ST2 antibody administration or ST2. The induction of nociceptive behaviors in naive mice due to recombinant IL-33 was reversed by the noncompetitive NMDA antagonist MK-801. ST2 antibody administration or ST2 markedly inhibited the increased activation of the astroglial janus kinase 2 (JAK2)-signal transducer and activator of transcription 3 (STAT3) cascade and the neuronal calcium-calmodulin-dependent kinase II (CaMKII)-cyclic adenosine monophosphate response element-binding protein (CREB) cascade after SNI. Moreover, intrathecal pretreatment with the CaMKII inhibitor KN-93 or the JAK2-STAT3 cascade inhibitor AG490 attenuated recombinant IL-33-induced nociceptive behaviors and NMDA subunit 1 up-regulation in naive mice. Spinal IL-33/ST2 signaling contributes to neuropathic pain by activating the astroglial JAK2-STAT3 cascade and the neuronal CaMKII-CREB cascade.

Transcriptional control, but not subcellular location, of PGC-1α is altered following exercise in a hot environment

PubMed Central

Shute, Robert J.; Kreiling, Jodi L.

2016-01-01

The purpose of this study was to determine mitochondrial biogenesis-related mRNA expression, binding of transcription factors to the peroxisome proliferator-activated receptor-γ coactivator 1-α (PGC-1α) promoter, and subcellular location of PGC-1α protein in human skeletal muscle following exercise in a hot environment compared with a room temperature environment. Recreationally trained males (n = 11) completed two trials in a temperature- and humidity-controlled environmental chamber. Each trial consisted of cycling in either a hot (H) or room temperature (C) environment (33 and 20°C, respectively) for 1 h at 60% of maximum wattage (Wmax) followed by 3 h of supine recovery at room temperature. Muscle biopsies were taken from the vastus lateralis pre-, post-, and 3 h postexercise. PGC-1α mRNA increased post (P = 0.039)- and 3 h postexercise in C (P = 0.002). PGC-1α, estrogen-related receptor-α (ERRα), and nuclear respiratory factor 1 (NRF-1) mRNA was all lower in H than C post (P = 0.038, P < 0.001, and P = 0.030, respectively)- and 3 h postexercise (P = 0.035, P = 0.007, and P < 0.001, respectively). Binding of cAMP response element-binding protein (CREB) (P = 0.005), myocyte enhancer factor 2 (MEF2) (P = 0.047), and FoxO forkhead box class-O1 (FoxO1) (P = 0.010) to the promoter region of the PGC-1α gene was lower in H than C. Nuclear PGC-1α protein increased postexercise in both H and C (P = 0.029) but was not different between trials (P = 0.602). These data indicate that acute exercise in a hot environment blunts expression of mitochondrial biogenesis-related mRNA, due to decreased binding of CREB, MEF2, and FoxO1 to the PGC-1α promoter. PMID:27445305

LIMK1 regulates long-term memory and synaptic plasticity via the transcriptional factor CREB.

PubMed

Todorovski, Zarko; Asrar, Suhail; Liu, Jackie; Saw, Ner Mu Nar; Joshi, Krutika; Cortez, Miguel A; Snead, O Carter; Xie, Wei; Jia, Zhengping

2015-04-01

Deletion of the LIMK1 gene is associated with Williams syndrome, a unique neurodevelopmental disorder characterized by severe defects in visuospatial cognition and long-term memory (LTM). However, whether LIMK1 contributes to these deficits remains elusive. Here, we show that LIMK1-knockout (LIMK1(-/-)) mice are drastically impaired in LTM but not short-term memory (STM). In addition, LIMK1(-/-) mice are selectively defective in late-phase long-term potentiation (L-LTP), a form of long-lasting synaptic plasticity specifically required for the formation of LTM. Furthermore, we show that LIMK1 interacts and regulates the activity of cyclic AMP response element-binding protein (CREB), an extensively studied transcriptional factor critical for LTM. Importantly, both L-LTP and LTM deficits in LIMK1(-/-) mice are rescued by increasing the activity of CREB. These results provide strong evidence that LIMK1 deletion is sufficient to lead to an LTM deficit and that this deficit is attributable to CREB hypofunction. Our study has identified a direct gene-phenotype link in mice and provides a potential strategy to restore LTM in patients with Williams syndrome through the enhancement of CREB activity in the adult brain. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Specific TATAA and bZIP requirements suggest that HTLV-I Tax has transcriptional activity subsequent to the assembly of an initiation complex

PubMed Central

Ching, Yick-Pang; Chun, Abel CS; Chin, King-Tung; Zhang, Zhi-Qing; Jeang, Kuan-Teh; Jin, Dong-Yan

2004-01-01

Background Human T-cell leukemia virus type I (HTLV-I) Tax protein is a transcriptional regulator of viral and cellular genes. In this study we have examined in detail the determinants for Tax-mediated transcriptional activation. Results Whereas previously the LTR enhancer elements were thought to be the sole Tax-targets, herein, we find that the core HTLV-I TATAA motif also provides specific responsiveness not seen with either the SV40 or the E1b TATAA boxes. When enhancer elements which can mediate Tax-responsiveness were compared, the authentic HTLV-I 21-bp repeats were found to be the most effective. Related bZIP factors such as CREB, ATF4, c-Jun and LZIP are often thought to recognize the 21-bp repeats equivalently. However, amongst bZIP factors, we found that CREB, by far, is preferred by Tax for activation. When LTR transcription was reconstituted by substituting either κB or serum response elements in place of the 21-bp repeats, Tax activated these surrogate motifs using surfaces which are different from that utilized for CREB interaction. Finally, we employed artificial recruitment of TATA-binding protein to the HTLV-I promoter in "bypass" experiments to show for the first time that Tax has transcriptional activity subsequent to the assembly of an initiation complex at the promoter. Conclusions Optimal activation of the HTLV-I LTR by Tax specifically requires the core HTLV-I TATAA promoter, CREB and the 21-bp repeats. In addition, we also provide the first evidence for transcriptional activity of Tax after the recruitment of TATA-binding protein to the promoter. PMID:15285791

Oleanolic Acid Ameliorates Aβ25-35 Injection-induced Spatial Learning and Memory Deficit in Alzheimer's Disease Model Rats.

PubMed

Wang, Kai; Sun, Weiming; Zhang, Linlin; Guo, Wei; Xu, Jiachun; Liu, Shuang; Zhou, Zhen; Zhang, Yulian

2018-05-24

Abnormal amyloid β (Aβ) accumulation and deposition in hippocampus is an essential process in Alzheimer's disease (AD). To investigate whether Oleanolic acid (OA) could improve learning and memory deficit and its possible mechanism. Forty-five SD rats were randomly divided into sham operation group, model group, and OA group. AD models by injection of Aβ25-35 were built. Morris water maze (MWM) was applied to investigate learning and memory, transmission electron microscope (TEM) to observe the ultrastructure of synapse, western blot to the key targets of synapse, electrophysiology for long-term potentiation (LTP), and Ca2+ concentration in synapse was also measured. The latency time in model group was significantly longer than that in sham operation group (P=0.0001<0.05); while it was significantly shorter in the OA group than that in model group (P=0.0001<0.05); compared with model group, the times of cross-platform in OA group significantly increased (P = 0.0001 <0.05). TEM results showed OA couldalleviate neuron damage and synapses changes induced by Aβ25-35. The expression of CaMKII, PKC, NMDAR2B, BDNF, TrkB, and CREB protein were significantly improved by OA; the concentration of Ca2+ were significantly lower and the slope and amplitude of f-EPSP increased in OA group. OA could ameliorate Aβ-induced spatial learning and memory loss of AD rats, and the mechanism might be involved with maintaining synaptic integrity to restore synaptic plasticity and increasing the NMDAR2B protein, CaMKII and PKC protein in postsynaptic density (PSD), reducing synaptic Ca2+ concentration, enhancing LTP by up-regulating BDNF, TrkB, CREB proteins. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Repeated pulses of serotonin required for long-term facilitation activate mitogen-activated protein kinase in sensory neurons of Aplysia

PubMed Central

Michael, Dan; Martin, Kelsey C.; Seger, Rony; Ning, Ming-Ming; Baston, Rene; Kandel, Eric R.

1998-01-01

Long-term facilitation of the connections between the sensory and motor neurons of the gill-withdrawal reflex in Aplysia requires five repeated pulses of serotonin (5-HT). The repeated pulses of 5-HT initiate a cascade of gene activation that leads ultimately to the growth of new synaptic connections. Several genes in this process have been identified, including the transcriptional regulators apCREB-1, apCREB-2, apC/EBP, and the cell adhesion molecule apCAM, which is thought to be involved in the formation of new synaptic connections. Here we report that the transcriptional regulators apCREB-2 and apC/EBP, as well as a peptide derived from the cytoplasmic domain of apCAM, are phosphorylated in vitro by Aplysia mitogen-activated protein kinase (apMAPK). We have cloned the cDNA encoding apMAPK and show that apMAPK activity is increased in sensory neurons treated with repeated pulses of 5-HT and by the cAMP pathway. These results suggest that apMAPK may participate with cAMP-dependent protein kinase during long-term facilitation in sensory cells by modifying some of the key elements involved in the consolidation of short- to long-lasting changes in synaptic strength. PMID:9465108

Modulation of mutant Huntingtin aggregates and toxicity by human myeloid leukemia factors.

PubMed

Banerjee, Manisha; Datta, Moumita; Bhattacharyya, Nitai P

2017-01-01

Increased poly glutamine (polyQ) stretch at N-terminal of Huntingtin (HTT) causes Huntington's disease. HTT interacts with large number of proteins, although the preference for such interactions with wild type or mutated HTT protein remains largely unknown. HYPK, an intrinsically unstructured protein chaperone and interactor of mutant HTT was found to interact with myeloid leukemia factor 1 (MLF1) and 2 (MLF2). To identify the role of these two proteins in mutant HTT mediated aggregate formation and toxicity in a cell model, both the proteins were found to preferentially interact with the mutated N-terminal HTT. They significantly reduced the number of cells containing mutant HTT aggregates and subsequent apoptosis in Neuro2A cells. Additionally, in FRAP assay, mobile fraction of mutant HTT aggregates was increased in the presence of MLF1 or MLF2. Further, MLF1 could release transcription factors like p53, CBP and CREB from mutant HTT aggregates. Moreover, in HeLa cell co-expressing mutant HTT exon1 and full length MLF1, p53 was released from the aggregates, leading to the recovery of the expression of the GADD45A transcript, a p53 regulated gene. Taking together, these results showed that MLF1 and MLF2 modulated the formation of aggregates and induction of apoptosis as well as the expressions of genes indirectly. Copyright © 2016 Elsevier Ltd. All rights reserved.

Neurogenic transdifferentiation of human adipose-derived stem cells? A critical protocol reevaluation with special emphasis on cell proliferation and cell cycle alterations.

PubMed

Kompisch, Kai Michael; Lange, Claudia; Steinemann, Doris; Skawran, Britta; Schlegelberger, Brigitte; Müller, Reinhard; Schumacher, Udo

2010-11-01

Adipose-derived stem cells (ASCs) are reported to display multilineage differentiation potential, including neuroectodermal pathways. The aim of the present study was to critically re-evaluate the potential neurogenic (trans-)differentiation capacity of ASCs using a neurogenic induction protocol based on the combination of isobutylmethylxanthine (IBMX), indomethacin and insulin. ASCs isolated from lipo-aspirate samples of five healthy female donors were characterized and potential neurogenic (trans-)differentiation was assessed by means of immunohistochemistry and gene expression analyses. Cell proliferation and cell cycle alterations were studied, and the expression of CREB/ATF transcription factors was analyzed. ASCs expressed CD59, CD90 and CD105, and were tested negative for CD34 and CD45. Under neurogenic induction, ASCs adopted a characteristic morphology comparable to neur(on)al progenitors and expressed musashi1, β-III-tubulin and nestin. Gene expression analyses revealed an increased expression of β-III-tubulin, GFAP, vimentin and BDNF, as well as SOX4 in induced ASCs. Cell proliferation was significantly reduced under neurogenic induction; cell cycle analyses showed a G2-cell cycle arrest accompanied by differential expression of key regulators of cell cycle progression. Differential expression of CREB/ATF transcription factors could be observed on neurogenic induction, pointing to a decisive role of the cAMP-CREB/ATF system. Our findings may point to a potential neurogenic (trans-)differentiation of ASCs into early neur(on)al progenitors, but do not present definite evidence for it. Especially, the adoption of a neural progenitor cell-like morphology must not automatically be misinterpreted as a specific characteristic of a respective (trans-)differentiation process, as this may as well be caused by alterations of cell cycle progression.

The parathyroid hormone-regulated transcriptome in osteocytes: parallel actions with 1,25-dihydroxyvitamin D3 to oppose gene expression changes during differentiation and to promote mature cell function.

PubMed

St John, Hillary C; Meyer, Mark B; Benkusky, Nancy A; Carlson, Alex H; Prideaux, Mathew; Bonewald, Lynda F; Pike, J Wesley

2015-03-01

Although localized to the mineralized matrix of bone, osteocytes are able to respond to systemic factors such as the calciotropic hormones 1,25(OH)2D3 and PTH. In the present studies, we examined the transcriptomic response to PTH in an osteocyte cell model and found that this hormone regulated an extensive panel of genes. Surprisingly, PTH uniquely modulated two cohorts of genes, one that was expressed and associated with the osteoblast to osteocyte transition and the other a cohort that was expressed only in the mature osteocyte. Interestingly, PTH's effects were largely to oppose the expression of differentiation-related genes in the former cohort, while potentiating the expression of osteocyte-specific genes in the latter cohort. A comparison of the transcriptional effects of PTH with those obtained previously with 1,25(OH)2D3 revealed a subset of genes that was strongly overlapping. While 1,25(OH)2D3 potentiated the expression of osteocyte-specific genes similar to that seen with PTH, the overlap between the two hormones was more limited. Additional experiments identified the PKA-activated phospho-CREB (pCREB) cistrome, revealing that while many of the differentiation-related PTH regulated genes were apparent targets of a PKA-mediated signaling pathway, a reduction in pCREB binding at sites associated with osteocyte-specific PTH targets appeared to involve alternative PTH activation pathways. That pCREB binding activities positioned near important hormone-regulated gene cohorts were localized to control regions of genes was reinforced by the presence of epigenetic enhancer signatures exemplified by unique modifications at histones H3 and H4. These studies suggest that both PTH and 1,25(OH)2D3 may play important and perhaps cooperative roles in limiting osteocyte differentiation from its precursors while simultaneously exerting distinct roles in regulating mature osteocyte function. Our results provide new insight into transcription factor-associated mechanisms through which PTH and 1,25(OH)2D3 regulate a plethora of genes important to the osteoblast/osteocyte lineage. Copyright © 2014 Elsevier Inc. All rights reserved.

Activation of Extracellular Signal-Regulated Kinase in the Anterior Cingulate Cortex Contributes to the Induction and Expression of Affective Pain

PubMed Central

Cao, Hong; Gao, Yong-Jing; Ren, Wen-Hua; Li, Ting-Ting; Duan, Kai-Zheng; Cui, Yi-Hui; Cao, Xiao-Hua; Zhao, Zhi-Qi; Ji, Ru-Rong; Zhang, Yu-Qiu

2009-01-01

The anterior cingulate cortex (ACC) is implicated in the affective response to noxious stimuli. However, little is known about the molecular mechanisms involved. The present study demonstrated that extracellular signal-regulated kinase (ERK) activation in the ACC plays a crucial role in pain-related negative emotion. Intraplantar formalin injection produced a transient ERK activation in laminae V–VI and a persistent ERK activation in laminae II–III of the rostral ACC (rACC) bilaterally. Using formalin-induced conditioned place avoidance (F-CPA) in rats, which is believed to reflect the pain-related negative emotion, we found that blockade of ERK activation in the rACC with MEK inhibitors prevented the induction of F-CPA. Interestingly, this blockade did not affect formalin-induced two-phase spontaneous nociceptive responses and CPA acquisition induced by electric foot-shock or U69,593, an innocuous aversive agent. Upstream, NMDA receptor, adenylyl cyclase (AC) and PKA activators activated ERK in rACC slices. Consistently, intra-rACC microinjection of AC or PKA inhibitors prevented F-CPA induction. Downstream, phosphorylation of cAMP response element binding protein (CREB) was induced in the rACC by formalin injection and by NMDA, AC and PKA activators in brain slices, which was suppressed by MEK inhibitors. Furthermore, ERK also contributed to the expression of pain-related negative emotion. Thus, when rats were re-exposed to the conditioning context for retrieval of pain experience, ERK and CREB were re-activated in the rACC, and inhibiting ERK activation blocked the expression of F-CPA. All together, our results demonstrate that ERK activation in the rACC is required for the induction and expression of pain-related negative affect. PMID:19279268

Nitric oxide increases Wnt-induced secreted protein-1 (WISP-1/CCN4) expression and function in colitis.

PubMed

Wang, Hongying; Zhang, Rui; Wen, Shoubin; McCafferty, Donna-Marie; Beck, Paul L; MacNaughton, Wallace K

2009-04-01

Nitric oxide (NO) derived from the inducible NO synthase (iNOS) is an important and complex mediator of inflammation in the intestine. Wnt-inducible secreted protein (WISP)-1 (CCN4), a member of the connective tissue growth factor family, is involved in tissue repair. We sought to determine the relationship between iNOS and WISP-1 in colitis. By analyzing human colonic biopsy samples, we showed that the expression of mRNA for both iNOS and WISP-1 was significantly higher in ulcerative colitis samples compared with control tissue. The upregulation of WISP-1 was positively correlated with iNOS expression in two models of colitis, induced by intrarectal trinitrobenzenesulfonic acid (TNBS) or occurring spontaneously in IL-10 deficient mice. Loss of iNOS, studied using iNOS(-/-) mice in both TNBS-induced and IL-10(-/-) colitis models, significantly attenuated the colitis-related WISP-1 increase. In human colonic epithelial cell lines, the NO donor, DETA-NONOate, elevated WISP-1 mRNA and protein expression through a beta-catenin and CREB-dependent, but Wnt-1-independent, pathway. In addition, NO-induced WISP-1 directly induced secretion of soluble collagen in colonic fibroblast cells. NO increases WISP-1 expression both in vitro and in vivo, suggesting a new role for iNOS and NO in colitis.

Lifespan extension induced by AMPK and calcineurin is mediated by CRTC-1 and CREB.

PubMed

Mair, William; Morantte, Ianessa; Rodrigues, Ana P C; Manning, Gerard; Montminy, Marc; Shaw, Reuben J; Dillin, Andrew

2011-02-17

Activating AMPK or inactivating calcineurin slows ageing in Caenorhabditis elegans and both have been implicated as therapeutic targets for age-related pathology in mammals. However, the direct targets that mediate their effects on longevity remain unclear. In mammals, CREB-regulated transcriptional coactivators (CRTCs) are a family of cofactors involved in diverse physiological processes including energy homeostasis, cancer and endoplasmic reticulum stress. Here we show that both AMPK and calcineurin modulate longevity exclusively through post-translational modification of CRTC-1, the sole C. elegans CRTC. We demonstrate that CRTC-1 is a direct AMPK target, and interacts with the CREB homologue-1 (CRH-1) transcription factor in vivo. The pro-longevity effects of activating AMPK or deactivating calcineurin decrease CRTC-1 and CRH-1 activity and induce transcriptional responses similar to those of CRH-1 null worms. Downregulation of crtc-1 increases lifespan in a crh-1-dependent manner and directly reducing crh-1 expression increases longevity, substantiating a role for CRTCs and CREB in ageing. Together, these findings indicate a novel role for CRTCs and CREB in determining lifespan downstream of AMPK and calcineurin, and illustrate the molecular mechanisms by which an evolutionarily conserved pathway responds to low energy to increase longevity.

Lifespan extension induced by AMPK and calcineurin is mediated by CRTC-1 and CREB

PubMed Central

Mair, William; Morantte, Ianessa; Rodrigues, Ana P. C.; Manning, Gerard; Montminy, Marc; Shaw, Reuben J.; Dillin, Andrew

2011-01-01

Activating AMPK or inactivating calcineurin slows ageing in Caenorhabditis elegans1,2 and both have been implicated as therapeutic targets for age-related pathology in mammals3–5. However, the direct targets that mediate their effects on longevity remain unclear. In mammals, CREB-regulated transcriptional coactivators (CRTCs)6 are a family of cofactors involved in diverse physiological processes including energy homeostasis7–9, cancer10 and endoplasmic reticulum stress11. Here we show that both AMPK and calcineurin modulate longevity exclusively through post-translational modification of CRTC-1, the sole C. elegans CRTC. We demonstrate that CRTC-1 is a direct AMPK target, and interacts with the CREB homologue-1 (CRH-1) transcription factor in vivo. The pro-longevity effects of activating AMPK or deactivating calcineurin decrease CRTC-1 and CRH-1 activity and induce transcriptional responses similar to those of CRH-1 null worms. Downregulation of crtc-1 increases lifespan in a crh-1-dependent manner and directly reducing crh-1 expression increases longevity, substantiating a role for CRTCs and CREB in ageing. Together, these findings indicate a novel role for CRTCs and CREB in determining lifespan downstream of AMPK and calcineurin, and illustrate the molecular mechanisms by which an evolutionarily conserved pathway responds to low energy to increase longevity. PMID:21331044

Voluntary resistance running with short distance enhances spatial memory related to hippocampal BDNF signaling.

PubMed

Lee, Min Chul; Okamoto, Masahiro; Liu, Yu Fan; Inoue, Koshiro; Matsui, Takashi; Nogami, Haruo; Soya, Hideaki

2012-10-15

Although voluntary running has beneficial effects on hippocampal cognitive functions if done abundantly, it is still uncertain whether resistance running would be the same. For this purpose, voluntary resistance wheel running (RWR) with a load is a suitable model, since it allows increased work levels and resultant muscular adaptation in fast-twitch muscle. Here, we examined whether RWR would have potential effects on hippocampal cognitive functions with enhanced hippocampal brain-derived neurotrophic factor (BDNF), as does wheel running without a load (WR). Ten-week-old male Wistar rats were assigned randomly to sedentary (Sed), WR, and RWR (to a maximum load of 30% of body weight) groups for 4 wk. We found that in RWR, work levels increased with load, but running distance decreased by about half, which elicited muscular adaptation for fast-twitch plantaris muscle without causing any negative stress effects. Both RWR and WR led to improved spatial learning and memory as well as gene expressions of hippocampal BDNF signaling-related molecules. RWR increased hippocampal BDNF, tyrosine-related kinase B (TrkB), and cAMP response element-binding (CREB) protein levels, whereas WR increased only BDNF. With both exercise groups, there were correlations between spatial memory and BDNF protein (r = 0.41), p-CREB protein (r = 0.44), and work levels (r = 0.77). These results suggest that RWR plays a beneficial role in hippocampus-related cognitive functions associated with hippocampal BDNF signaling, even with short distances, and that work levels rather than running distance are more determinant of exercise-induced beneficial effects in wheel running with and without a load.

Excitation-transcription coupling in parvalbumin-positive interneurons employs a novel CaM Kinase-dependent pathway distinct from excitatory neurons

PubMed Central

Cohen, Samuel M.; Ma, Huan; Kuchibhotla, Kishore V.; Watson, Brendon O.; Buzsáki, György; Froemke, Robert C.; Tsien, Richard W.

2016-01-01

Properly functional CNS circuits depend on inhibitory interneurons that in turn rely upon activity-dependent gene expression for morphological development, connectivity and excitatory-inhibitory coordination. Despite its importance, excitation-transcription coupling in inhibitory interneurons is poorly understood. Here, we report that PV+ interneurons employ a novel CaMK-dependent pathway to trigger CREB phosphorylation and gene expression. As in excitatory neurons, voltage-gated Ca2+ influx through CaV1 channels triggers CaM nuclear translocation via local Ca2+ signaling. However, PV+ interneurons are distinct in that nuclear signaling is mediated by γCaMKI, not γCaMKII. CREB phosphorylation also proceeds with slow, sigmoid kinetics, rate-limited by paucity of CaMKIV, protecting against saturation of phospho-CREB in the face of higher firing rates and bigger Ca2+ transients. Our findings support the generality of CaM shuttling to drive nuclear CaMK activity, and are relevant to disease pathophysiology, insofar as dysfunction of PV+ interneurons and molecules underpinning their excitation-transcription coupling both relate to neuropsychiatric disease. PMID:27041500

Molecular cloning and characterization of an SRCAP chromatin remodeling homologue in Toxoplasma gondii.

PubMed

Sullivan, William J; Monroy, M Alexandra; Bohne, Wolfgang; Nallani, Karuna C; Chrivia, John; Yaciuk, Peter; Smith, Charles K; Queener, Sherry F

2003-05-01

We have identified and mapped a gene in Toxoplasma gondii that encodes a homologue of SRCAP (Snf2-related CBP activator protein), a member of the SNF/SWI family of chromatin remodeling factors. The genomic locus (TgSRCAP) is present as a single copy and contains 16 introns. The predicted cDNA contains an open reading frame of 8,775 bp and encodes a protein of 2,924 amino acids. We have identified additional SRCAP-like sequences in Apicomplexa for comparison by screening genomic databases. An analysis of SRCAP homologues between species reveals signature features that may be indicative of SRCAP members. Expression of mRNA encoding TgSRCAP is upregulated when tachyzoite (invasive form) parasites are induced to differentiate into bradyzoites (encysted form) in vitro. Recombinant TgSRCAP protein is functionally equivalent to the human homologue, being capable of increasing transcription mediated by CREB.

Inhibitory effects of ginseng total saponin on up-regulation of cAMP pathway induced by repeated administration of morphine.

PubMed

Seo, Jeong-Ju; Lee, Jae-Woong; Lee, Wan-Kyu; Hong, Jin-Tae; Lee, Chong-Kil; Lee, Myung-Koo; Oh, Ki-Wan

2008-02-01

We have reported that ginseng total saponin (GTS) inhibited the development of physical and psychological dependence on morphine. However, the possible molecular mechanisms of GTS are unclear. Therefore, this study was undertaken to understand the possible molecular mechanism of GTS on the inhibitory effects of morphine-induced dependence. It has been reported that the up-regulated cAMP pathway in the LC of the mouse brain after repeated administration of morphine contributes to the feature of withdrawals. GTS inhibited up-regulation of cAMP pathway in the LC after repeated administration of morphine in this experiment. GTS inhibited cAMP levels and protein expression of protein kinase A (PKA). In addition, GTS inhibited the increase of cAMP response element binding protein (CREB) phosphorylation. Therefore, we conclude that the inhibitory effects of GTS on morphine-induced dependence might be mediated by the inhibition of cAMP pathway.

Molecular and genetic substrates linking stress and addiction.

PubMed

Briand, Lisa A; Blendy, Julie A

2010-02-16

Drug addiction is one of the top three health concerns in the United States in terms of economic and health care costs. Despite this, there are very few effective treatment options available. Therefore, understanding the causes and molecular mechanisms underlying the transition from casual drug use to compulsive drug addiction could aid in the development of treatment options. Studies in humans and animal models indicate that stress can lead to both vulnerability to develop addiction, and increased drug taking and relapse in addicted individuals. Exposure to stress or drugs of abuse results in long-term adaptations in the brain that are likely to involve persistent alterations in gene expression or activation of transcription factors, such as the cAMP Response Element Binding (CREB) protein. The signaling pathways controlled by CREB have been strongly implicated in drug addiction and stress. Many potential CREB target genes have been identified based on the presence of a CRE element in promoter DNA sequences. These include, but are not limited to CRF, BDNF, and dynorphin. These genes have been associated with initiation or reinstatement of drug reward and are altered in one direction or the other following stress. While many reviews have examined the interactions between stress and addiction, the goal of this review was to focus on specific molecules that play key roles in both stress and addiction and are therefore posed to mediate the interaction between the two. Focus on these molecules could provide us with new targets for pharmacological treatments for addiction. Copyright 2009 Elsevier B.V. All rights reserved.

Deficient Gene Expression in Protein Kinase Inhibitor α Null Mutant Mice

PubMed Central

Gangolli, Esha A.; Belyamani, Mouna; Muchinsky, Sara; Narula, Anita; Burton, Kimberly A.; McKnight, G. Stanley; Uhler, Michael D.; Idzerda, Rejean L.

2000-01-01

Protein kinase inhibitor (PKI) is a potent endogenous inhibitor of the cyclic AMP (cAMP)-dependent protein kinase (PKA). It functions by binding the free catalytic (C) subunit with a high affinity and is also known to export nuclear C subunit to the cytoplasm. The significance of these actions with respect to PKI's physiological role is not well understood. To address this, we have generated by homologous recombination mutant mice that are deficient in PKIα, one of the three isoforms of PKI. The mice completely lack PKI activity in skeletal muscle and, surprisingly, show decreased basal and isoproterenol-induced gene expression in muscle. Further examination revealed reduced levels of the phosphorylated (active) form of the transcription factor CREB (cAMP response element binding protein) in the knockouts. This phenomenon stems, at least in part, from lower basal PKA activity levels in the mutants, arising from a compensatory increase in the level of the RIα subunit of PKA. The deficit in gene induction, however, is not easily explained by current models of PKI function and suggests that PKI may play an as yet undescribed role in PKA signaling. PMID:10779334

Modulation of opiate-related signaling molecules in morphine-dependent conditioned behavior: conditioned place preference to morphine induces CREB phosphorylation.

PubMed

Morón, José A; Gullapalli, Srinivas; Taylor, Chirisse; Gupta, Achla; Gomes, Ivone; Devi, Lakshmi A

2010-03-01

Opiate addiction is a chronic, relapsing behavioral disorder where learned associations that develop between the abused opiate and the environment in which it is consumed are brought about through Pavlovian (classical) conditioning processes. However, the signaling mechanisms/pathways regulating the mechanisms that underlie the responses to opiate-associated cues or the development of sensitization as a consequence of repeated context-independent administration of opiates are unknown. In this study we examined the phosphorylation levels of various classic signaling molecules in brain regions implicated in addictive behaviors after acute and repeated morphine administration. An unbiased place conditioning protocol was used to examine changes in phosphorylation that are associated with (1) the expression of the rewarding effects of morphine and (2) the sensitization that develops to this effect. We also examined the effects of a delta-receptor antagonist on morphine-induced conditioned behavior and on the phosphorylation of classic signaling molecules in view of data showing that blockade of delta-opioid receptor (deltaOR) prevents the development of sensitization to the rewarding effects of morphine. We find that CREB phosphorylation is specifically induced upon the expression of a sensitized response to morphine-induced conditioned behavior in brain areas related to memory consolidation, such as the hippocampus and cortex. A similar effect is also observed, albeit to a lesser extent, in the case of the GluR1 subunit of AMPA glutamate receptor. These increases in the phosphorylation levels of CREB and pGluR1 are significantly blocked by pretreatment with a deltaOR antagonist. These results indicate a critical role for phospho-CREB, AMPA, and deltaOR activities in mediating the expression of a sensitized response to morphine-dependent conditioned behavior.

Continuing the search for the engram: examining the mechanism of fear memories.

PubMed

Josselyn, Sheena A

2010-07-01

The goal of my research is to gain insight using rodent models into the fundamental molecular, cellular and systems that make up the base of memory formation. My work focuses on fear memories. Aberrant fear and/or anxiety may be at the heart of many psychiatric disorders. In this article, I review the results of my research group; these results show that particular neurons in the lateral amygdala, a brain region important for fear, are specifically involved in particular fear memories. We started by showing that the transcription factor CREB (cAMP/Ca(2+) response element binding protein) plays a key role in the formation of fear memories. Next, we used viral vectors to overexpress CREB in a subset of lateral amygdala neurons. This not only facilitated fear memory formation but also "drove" the memory into the neurons with relatively increased CREB function. Finally, we showed that selective ablation of the neurons overexpressing CREB in the lateral amygdala selectively erased the fear memory. These findings are the first to show disruption of a specific memory by disrupting select neurons within a distributed network.

Molecular partners of hNOT/ALG3, the human counterpart of the Drosophila NOT and yeast ALG3 gene, suggest its involvement in distinct cellular processes relevant to congenital disorders of glycosylation, cancer, neurodegeneration and a variety of further pathologies.

PubMed

Hacker, Benedikt; Schultheiß, Christoph; Döring, Michael; Kurzik-Dumke, Ursula

2018-06-01

This study provides first insights into the involvement of hNOT/ALG3, the human counterpart of the Drosophila Neighbour of TID and yeast ALG3 gene, in various putative molecular networks. HNOT/ALG3 encodes two translated transcripts encoding precursor proteins differing in their N-terminus and showing 33% identity with the yeast asparagine-linked glycosylation 3 (ALG3) protein. Experimental evidence for the functional homology of the proteins of fly and man in the N-glycosylation has still to be provided. In this study, using the yeast two-hybrid technique we identify 17 molecular partners of hNOT-1/ALG3-1. We disclose the building of hNOT/ALG3 homodimers and provide experimental evidence for its in vivo interaction with the functionally linked proteins OSBP, OSBPL9 and LRP1, the SYPL1 protein and the transcription factor CREB3. Regarding the latter, we show that the 55 kDa N-glycosylated hNOT-1/ALG3-1 molecule binds the N-glycosylated CREB3 precursor but does not interact with CREB3's proteolytic products specific to the endoplasmic reticulum and to the nucleus. The interaction between the two partners is a prerequisite for the proteolytic activation of CREB3. In case of the further binding partners, our data suggest that hNOT-1/ALG3-1 interacts with both OSBPs and with their direct targets LRP1 and VAMP/VAP-A. Moreover, our results show that various partners of hNOT-1/ALG3-1 interact with its diverse post translationally processed products destined to distinct cellular compartments. Generally, our data suggest the involvement of hNOT-1/ALG3-1 in various molecular contexts determining essential processes associated with distinct cellular machineries and related to various pathologies, such as cancer, viral infections, neuronal and immunological disorders and CDG.

17β-estradiol-induced growth of triple-negative breast cancer cells is prevented by the reduction of GPER expression after treatment with gefitinib.

PubMed

Girgert, Rainer; Emons, Günter; Gründker, Carsten

2017-02-01

Triple-negative breast cancers (TNBCs) are neither susceptible to endocrine therapy due to a lack of estrogen receptor α expression nor trastuzumab. TNBCs frequently overexpress epidermal growth factor receptor (EGFR) and membrane bound estrogen receptor, GPER. To a certain extent the growth of TNBCs is stimulated by 17β-estradiol via GPER. We analyzed whether inhibition of EGFR by gefitinib reduces the expression of GPER and subsequent signal transduction in TNBC cells. Dependence of proliferation on 17β-estradiol was determined using Alamar Blue assay. Expression of GPR30 and activation of c-src, EGFR and cAMP-responsive element binding (CREB) protein by 17β-estradiol was analyzed by western blotting. Expression of c-fos, cyclin D1 and aromatase was determined using RT-PCR. Gefitinib reduced GPER expression concentration‑ and time‑dependently. In HCC70 cells, GPER expression was reduced to 15±11% (p<0.05) after treatment with 200 nM gefitinib for four days, and in HCC1806 cells GPER expression was reduced to 39±5% (p<0.01) of the control. 17β-estradiol significantly increased the percentage of HCC1806 cells within 7 days to 145±29% of the control (HCC70, 110±8%). This increase in cell growth was completely prevented in both TNBC cell lines after GPR30 expression was downregulated by treatment with 200 nM gefitinib. In HCC1806 cells, activation of c-src was increased by 17β-estradiol to 350±50% (p<0.01), and gefitinib reduced src activation to 110%. Similar results were obtained in the HCC70 cells. Phosphorylation of EGFR increased to 240±40% (p<0.05) in the HCC1806 cells treated with 17β-estradiol (HCC70, 147±25%). Gefitinib completely prevented this activation. Phosphorylation of CREB and induction of c-fos, cyclin D1 and aromatase expression by 17β-estradiol were all prevented by gefitinib. These experiments conclusively show that reduction of GPER expression is a promising therapeutic approach for TNBC.

Chronic administration of aripiprazole activates GSK3β-dependent signalling pathways, and up-regulates GABAA receptor expression and CREB1 activity in rats

PubMed Central

Pan, Bo; Huang, Xu-Feng; Deng, Chao

2016-01-01

Aripiprazole is a D2-like receptor (D2R) partial agonist with a favourable clinical profile. Previous investigations indicated that acute and short-term administration of aripiprazole had effects on PKA activity, GSK3β-dependent pathways, GABAA receptors, NMDA receptor and CREB1 in the brain. Since antipsychotics are used chronically in clinics, the present study investigated the long-term effects of chronic oral aripiprazole treatment on these cellular signalling pathways, in comparison with haloperidol (a D2R antagonist) and bifeprunox (a potent D2R partial agonist). We found that the Akt-GSK3β pathway was activated by aripiprazole and bifeprunox in the prefrontal cortex; NMDA NR2A levels were reduced by aripiprazole and haloperidol. In the nucleus accumbens, all three drugs increased Akt-GSK3β signalling; in addition, both aripiprazole and haloperidol, but not bifeprunox, increased the expression of Dvl-3, β-catenin and GABAA receptors, NMDA receptor subunits, as well as CREB1 phosphorylation levels. The results suggest that chronic oral administration of aripiprazole affects schizophrenia-related cellular signalling pathways and markers (including Akt-GSK3β signalling, Dvl-GSK3β-β-catenin signalling, GABAA receptor, NMDA receptor and CREB1) in a brain-region-dependent manner; the selective effects of aripiprazole on these signalling pathways might be associated with its unique clinical effects. PMID:27435909

Supercritical Fluid Extract of Spent Coffee Grounds Attenuates Melanogenesis through Downregulation of the PKA, PI3K/Akt, and MAPK Signaling Pathways.

PubMed

Huang, Huey-Chun; Wei, Chien-Mei; Siao, Jen-Hung; Tsai, Tsang-Chi; Ko, Wang-Ping; Chang, Kuei-Jen; Hii, Choon-Hoon; Chang, Tsong-Min

2016-01-01

The mode of action of spent coffee grounds supercritical fluid CO2 extract (SFE) in melanogenesis has never been reported. In the study, the spent coffee grounds were extracted by the supercritical fluid CO2 extraction method; the chemical constituents of the SFE were investigated by gas chromatography-mass spectrometry (GC-MS). The effects of the SFE and its major fatty acid components on melanogenesis were evaluated by mushroom tyrosinase activity assay and determination of intracellular tyrosinase activity and melanin content. The expression level of melanogenesis-related proteins was analyzed by western blotting assay. The results revealed that the SFE of spent coffee grounds (1-10 mg/mL) and its major fatty acids such as linoleic acid and oleic acid (6.25-50 μM) effectively suppressed melanogenesis in the B16F10 murine melanoma cells. Furthermore, the SFE decreased the expression of melanocortin 1 receptor (MC1R), microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein-1 (TRP-1), and tyrosinase-related protein-2 (TRP-2). The SFE also decreased the protein expression levels of p-JNK, p-p38, p-ERK, and p-CREB. Our results revealed that the SFE of spent coffee grounds attenuated melanogenesis in B16F10 cells by downregulation of protein kinase A (PKA), phosphatidylinositol-3-kinase (PI3K/Akt), and mitogen-activated protein kinases (MAPK) signaling pathways, which may be due to linoleic acid and oleic acid.

Noni (Morinda citrifolia L.) fruit juice reverses age-related decline in neural-immune interactions in the spleens of old F344 rats.

PubMed

Pratap, Uday P; Hima, Lalgi; Priyanka, Hannah P; ThyagaRajan, Srinivasan

2017-02-23

Various parts of the tropical plant, Morinda citrifolia L. (Noni), have been widely used in traditional medicine in South and Southeast Asia for several centuries. The therapeutic effects of the noni are believed to be mediated through several phytochemicals such as anthraquinones, iridoid, fatty acid glycosides, alcohols, etc. The aim of the study is to investigate the effects of Morinda citrifolia fruit juice (noni fruit juice; NFJ) on neural-immune interactions through the involvement of intracellular signaling pathways both in vitro and in vivo in the splenic lymphocytes of young and old male F344 rats. In the in vitro study, splenocytes from young and old F344 rats were isolated and treated with 0.0001-1% concentrations of NFJ for a period of 24h, while in the in vivo study, old F344 rats were orally administered (5ml/kg body weight) with NFJ (5%, 10% and 20%) twice daily for 60 days. After the treatment period, concanavalin A (Con A)-induced lymphocyte proliferation, cytokines (IL-2, IFN-γ, IL-6, and TNF-α) production, expression of tyrosine hydroxylase (p-TH), nerve growth factor (NGF), m-TOR, IκB-α, p-NF-κB (p50 and p65), p-ERK, p-Akt, p-CREB and lipid peroxidation, protein carbonyl formation, nitric oxide (NO) production were examined in the splenocytes. In vitro NFJ incubation of splenic lymphocytes increased Con A-induced lymphocyte proliferation, IL-2 and IFN-γ production, and expression of p-ERK, p-Akt, and p-CREB in young and old rats. In vivo treatment of old rats with NFJ increased lymphoproliferation, IL-2 and IFN-γ production, the expression of p-TH, NGF, and NO production, and suppressed IL-6 production, lipid peroxidation, protein carbonyl formation, and the expression of IκB-α and p-NF-κB (p50) in the splenocytes. Taken together, these results suggest that Morinda citrifolia fruit juice enhanced neural-immune interactions and cell survival pathways while inhibiting inflammatory processes that may be useful in the treatment of age-associated diseases. Copyright © 2017 Elsevier Ireland Ltd. All rights reserved.

Genome-wide analysis identifies colonic genes differentially associated with serum leptin and insulin concentrations in C57BL/6J mice fed a high-fat diet.

PubMed

Kim, Sung-Eun; Choo, Jinsil; Yoon, Joon; Chu, Jae Ryang; Bae, Yun Jung; Lee, Seungyeoun; Park, Taesung; Sung, Mi-Kyung

2017-01-01

Obesity-induced chronic inflammation is known to increase the risk of ulcerative colitis, Crohn's disease, and colorectal cancer. Accumulating evidence suggests that leptin and insulin are key molecules linking obesity with diseases of the lower intestine. Here, we identified serum phenotype-associated genes in the colon of diet-induced obese mice as early biomarkers of obesity-associated colonic diseases. C57BL/6J mice were fed with either normal diet (ND, 15% of fat calories) or high-fat diet (HFD, 45% of fat calories) for 8 weeks. Serum concentrations of insulin, insulin-like growth factor-1 (IGF-1), leptin, and adiponectin were measured as obesity-related phenotypic markers. Genome-wide gene expression profiles of colon tissue were determined, followed by statistical analyses to detect differentially expressed and serum phenotype-associated genes. HFD-fed mice showed higher serum concentrations of leptin (P < 0.001) and insulin (P < 0.01) than those in the ND group, whereas serum IGF-1 and adiponectin concentrations did not differ between the two dietary groups. Among differentially expressed genes affected by HFD, 135, 128, 110, and 341 genes were associated with serum levels of leptin, insulin, IGF-1, and adiponectin, respectively. We identified 17 leptin-associated genes and 4 insulin-associated genes that inversely responded to HFD and ND. Among these, leptin-associated Peli3 (Pellino E3 ubiquitin protein ligase family member 3), Creb1 (cAMP responsive element binding protein 1), and Enpp2 (ectonucleotide pyrophosphatase/phosphodiesterase 2, autotaxin) and insulin-associated Centg1 (AGAP2, ArfGAP with GTPase domain) are reported to play a role either in obesity or colonic diseases. mRNA expression of these genes was validated by RT-qPCR. Our data suggest Peli3, Creb1, Enpp2, and Centg1 as potential early biomarker candidates for obesity-induced pathophysiological changes in the colon. Future studies verifying the function of these candidates are needed for the prevention, early detection, and treatment of colon diseases.

Genome-wide analysis identifies colonic genes differentially associated with serum leptin and insulin concentrations in C57BL/6J mice fed a high-fat diet

PubMed Central

Yoon, Joon; Chu, Jae Ryang; Bae, Yun Jung; Lee, Seungyeoun; Park, Taesung; Sung, Mi-Kyung

2017-01-01

Obesity-induced chronic inflammation is known to increase the risk of ulcerative colitis, Crohn’s disease, and colorectal cancer. Accumulating evidence suggests that leptin and insulin are key molecules linking obesity with diseases of the lower intestine. Here, we identified serum phenotype-associated genes in the colon of diet-induced obese mice as early biomarkers of obesity-associated colonic diseases. C57BL/6J mice were fed with either normal diet (ND, 15% of fat calories) or high-fat diet (HFD, 45% of fat calories) for 8 weeks. Serum concentrations of insulin, insulin-like growth factor-1 (IGF-1), leptin, and adiponectin were measured as obesity-related phenotypic markers. Genome-wide gene expression profiles of colon tissue were determined, followed by statistical analyses to detect differentially expressed and serum phenotype-associated genes. HFD-fed mice showed higher serum concentrations of leptin (P < 0.001) and insulin (P < 0.01) than those in the ND group, whereas serum IGF-1 and adiponectin concentrations did not differ between the two dietary groups. Among differentially expressed genes affected by HFD, 135, 128, 110, and 341 genes were associated with serum levels of leptin, insulin, IGF-1, and adiponectin, respectively. We identified 17 leptin-associated genes and 4 insulin-associated genes that inversely responded to HFD and ND. Among these, leptin-associated Peli3 (Pellino E3 ubiquitin protein ligase family member 3), Creb1 (cAMP responsive element binding protein 1), and Enpp2 (ectonucleotide pyrophosphatase/phosphodiesterase 2, autotaxin) and insulin-associated Centg1 (AGAP2, ArfGAP with GTPase domain) are reported to play a role either in obesity or colonic diseases. mRNA expression of these genes was validated by RT-qPCR. Our data suggest Peli3, Creb1, Enpp2, and Centg1 as potential early biomarker candidates for obesity-induced pathophysiological changes in the colon. Future studies verifying the function of these candidates are needed for the prevention, early detection, and treatment of colon diseases. PMID:28170448

Changes in Global Transcriptional Profiling of Women Following Obesity Surgery Bypass.

PubMed

Pinhel, Marcela Augusta de Souza; Noronha, Natalia Yumi; Nicoletti, Carolina Ferreira; de Oliveira, Bruno Affonso Parente; Cortes-Oliveira, Cristiana; Pinhanelli, Vitor Caressato; Salgado Junior, Wilson; Machry, Ana Julia; da Silva Junior, Wilson Araújo; Souza, Dorotéia Rossi Silva; Marchini, Júlio Sérgio; Nonino, Carla Barbosa

2018-01-01

Differential gene expression in peripheral blood mononuclear cells (PBMCs) after Roux-en-Y gastric bypass (RYGB) is poorly characterized. Markers of these processes may provide a deeper understanding of the mechanisms that underlie these events. The main goal of this study was to identify changes in PBMC gene expression in women with obesity before and 6 months after RYGB-induced weight loss. The ribonucleic acid (RNA) of PBMCs from 13 obese women was analyzed before and 6 months after RYGB; the RNA of PBMCs from nine healthy women served as control. The gene expression levels were determined by microarray analysis. Significant differences in gene expression were validated by real-time quantitative polymerase chain reaction (RT-qPCR). Microarray analysis for comparison of the pre- and postoperative periods showed that 1366 genes were differentially expressed genes (DEGs). The main pathways were related to gene transcription; lipid, energy, and glycide metabolism; inflammatory and immunological response; cell differentiation; oxidative stress regulation; response to endogenous and exogenous stimuli; substrate oxidation; mTOR signaling pathway; interferon signaling; mitogen-activated protein kinases (MAPK), cAMP response element binding protein (CREB1), heat shock factor 1 (HSF1), and sterol regulatory element binding protein 1c (SREBP-1c) gene expression; adipocyte differentiation; and methylation. Six months after bariatric surgery and significant weight loss, many molecular pathways involved in obesity and metabolic diseases change. These findings are an important tool to identify potential targets for therapeutic intervention and clinical practice of nutritional genomics in obesity.

Prefrontal Consolidation Supports the Attainment of Fear Memory Accuracy

ERIC Educational Resources Information Center

Vieira, Philip A.; Lovelace, Jonathan W.; Corches, Alex; Rashid, Asim J.; Josselyn, Sheena A.; Korzus, Edward

2014-01-01

The neural mechanisms underlying the attainment of fear memory accuracy for appropriate discriminative responses to aversive and nonaversive stimuli are unclear. Considerable evidence indicates that coactivator of transcription and histone acetyltransferase cAMP response element binding protein (CREB) binding protein (CBP) is critically required…

Role of hippocampal and prefrontal cortical signaling pathways in dextromethorphan effect on morphine-induced memory impairment in rats.

PubMed

Ghasemzadeh, Zahra; Rezayof, Ameneh

2016-02-01

Evidence suggests that dextromethorphan (DM), an NMDA receptor antagonist, induces memory impairment. Considering that DM is widely used in cough-treating medications, and the co-abuse of DM with morphine has recently been reported, the aims of the present study was (1) to investigate whether there is a functional interaction between morphine and DM in passive avoidance learning and (2) to assess the possible role of the hippocampal and prefrontal cortical (PFC) signaling pathways in the effects of the drugs on memory formation. Our findings indicated that post-training or pre-test administration of morphine (2 and 6 mg/kg) or DM (10-30 mg/kg) impaired memory consolidation and retrieval which was associated with the attenuation of the levels of phosphorylated Ca(2+)/calmodulin-dependent protein kinase II (p-CAMKII) and cAMP responsive element-binding protein (p-CREB) in the targeted sites. Moreover, the memory impairment induced by post-training administration of morphine was reversed by pre-test administration of the same dose of morphine or DM (30 mg/kg), indicating state-dependent learning (SDL) and a cross-SDL between the drugs. It is important to note that the levels of p-CAMKII/CAMKII and p-CREB/CREB in the hippocampus and the PFC increased in drugs-induced SDL. In addition, DM administration potentiated morphine-induced SDL which was related to the enhanced levels of hippocampal and PFC CAMKII-CREB signaling pathways. It can be concluded that there is a relationship between the hippocampus and the PFC in the effect of DM and/or morphine on memory retrieval. Moreover, a cross SDL can be induced between the co-administration of DM and morphine. Interestingly, CAMKII-CREB signaling pathways also mediate the drugs-induced SDL. Copyright © 2015 Elsevier Inc. All rights reserved.

5-Lipoxygenase as an endogenous modulator of amyloid beta formation in vivo

PubMed Central

Chu, Jin; Praticò, Domenico

2010-01-01

Objective The 5-lipoxygenase (5-LO) enzymatic pathway is widely distributed within the central nervous system, and is up-regulated in Alzheimer's disease. However, the mechanism whereby it may influence the disease pathogenesis remains elusive. Methods We evaluated the molecular mechanism by which 5-LO regulates Amyloid β (Aβ) formation in vitro and in vivo by pharmacological and genetic approaches. Results Here we show that 5-LO regulates the formation of Aβ by activating the cAMP-response element binding protein (CREB), which in turn increases transcription of the γ-secretase complex. Preventing CREB activation by pharmacologic inhibition or dominant negative mutants blocks the 5-LO-dependent elevation of Aβ formation and the increase of γ-secretase mRNA and protein levels. Moreover, 5-LO targeted gene disruption or its in vivo selective pharmacological inhibition results in a significant reduction of Aβ, CREB and γ-secretase levels. Interpretation These data establish a novel functional role for 5-LO in regulating endogenous formation of Aβ levels in the central nervous system. Thus, 5-LO pharmacological inhibition may be beneficial in the treatment and prevention of Alzheimer's disease. PMID:21280074

NMDA receptor mediates proliferation and CREB phosphorylation in postnatal Müller glia-derived retinal progenitors

PubMed Central

Ramírez, Mónica

2009-01-01

Purpose Postnatal retinal Müller glia are considered to be retinal progenitors as they retain the ability to dedifferentiate, proliferate, and differentiate to new retinal glia and neurons after injury. The proliferation and differentiation processes are coordinated by several extrinsic factors and neurotransmitters, including glutamate. Thus, the appropriate numbers and proportions of the different cell types are generated to form a functional retina during development and during injury repair. Here we analyze the changes in the proliferation of postnatal Müller glia-derived progenitors after activation of the N-methyl-D-aspartate (NMDA) glutamate receptors. Methods Müller glia-derived progenitor cell cultures were characterized by immunocytochemistry with antibodies against the NR1 subunit of the NMDA receptor and the progenitor cell marker nestin. The effect of glutamate receptor agonists and antagonists on cell proliferation was analyzed by BrdU incorporation or Ki67 immunostaining, cell counting, and by immunolabeling of phosphorylated cAMP response element binding protein (P-CREB) transcription factor. The effect of NMDA receptor activation was analyzed in vivo by P-CREB immunohistochemistry in retinal sections of Long-Evans NMDA injected rats. Results We show that NMDA receptor activation significantly increases the proliferation rate of Müller-glia derived progenitor cells and that this increase can be blocked by NMDA receptor antagonists. Furthermore, we show that CREB phosphorylation is induced in NMDA-treated Müller-glia derived progenitor cells in culture and that specific pharmacological inhibition of CREB phosphorylation results in a decreased number of proliferating cells. We confirmed the relevance of these observations by the analysis of retinal sections after NMDA injection in vivo where immunoreactivity to phosphorylated CREB is also increased after treatment. Conclusions In the present study we show that NMDA receptor activation induces postnatal Müller glia-derived retinal cell progenitor proliferation and transcription factor CREB phosphorylation both in culture and in vivo. The identification of the molecular determinants of mature retinal progenitors such as transcription factor CREB and NMDA receptor-induced players should facilitate the control of growth and manipulation of progenitor cell cultures and the possible identification of the molecular mechanisms involved in progenitor self-renewal. PMID:19365572

A Low-Protein, High-Carbohydrate Diet Stimulates Thermogenesis in the Brown Adipose Tissue of Rats via ATF-2.

PubMed

de França, Suélem A; dos Santos, Maísa P; Przygodda, Franciele; Garófalo, Maria Antonieta R; Kettelhut, Isis C; Magalhães, Diego A; Bezerra, Kalinne S; Colodel, Edson M; Flouris, Andreas D; Andrade, Cláudia M B; Kawashita, Nair H

2016-03-01

The aim of this study was to evaluate thermogenesis in the interscapular brown adipose tissue (IBAT) of rats submitted to low-protein, high-carbohydrate (LPHC) diet and the involvement of adrenergic stimulation in this process. Male rats (~100 g) were submitted to LPHC (6%-protein; 74%-carbohydrate) or control (C; 17%-protein; 63%-carbohydrate) isocaloric diets for 15 days. The IBAT temperature was evaluated in the rats before and after the administration of noradrenaline (NA) (20 µg 100 g b w(-1) min(-1)). The expression levels of uncoupling protein 1 (UCP1) and other proteins involved in the regulation of UCP1 expression were determined by Western blot (Student's t test, P ≤ 0.05). The LPHC diet promoted a 1.1 °C increase in the basal temperature of IBAT when compared with the basal temperature in the IBAT of the C group. NA administration promoted a 0.3 °C increase in basal temperature in the IBAT of the C rats and a 0.5 °C increase in the IBAT of the LPHC group. The level of UCP1 increased 60% in the IBAT of LPHC-fed rats, and among the proteins involved in its expression, such as β3-AR and α1-AR, there was a 40% increase in the levels of p38-MAPK and a 30% decrease in CREB when compared to the C rats. The higher sympathetic flux to IBAT, which is a consequence of the administration of the LPHC diet to rats, activates thermogenesis and increases the expression of UCP1 in the tissue. Our results suggest that the increase in UCP1 content may occur via p38 MAPK and ATF2.

Tisp40 deficiency attenuates renal ischemia reperfusion injury induced apoptosis of tubular epithelial cells.

PubMed

Qin, Cong; Xiao, Chengcheng; Su, Yang; Zheng, Haizhou; Xu, Tao; Lu, Jingxiao; Luo, Pengcheng; Zhang, Jie

2017-10-01

Renal ischemia reperfusion (IR) is a major cause of acute kidney injury (AKI) and no effective treatments have been established. Tisp40 is a transcription factor of the CREB/ATF family and involves in cell apoptosis, proliferation and differentiation, but its role in renal IR remains unknown. Here, we investigated the role of Tisp40 in renal IR injury. In vivo, Tisp40 knockout (KO) and wild-type (WT) mice were subjected to thirty minutes of bilateral renal ischemia and 48h reperfusion, the blood and kidneys were harvested for analysis. In vitro, Tisp40 overexpression and vector cells were subjected to hypoxia/reoxygenation (HR), the apoptosis rate and the expressions of related proteins were measured. Following IR, the expressions of Tisp40 protein, serum creatinine (sCr), blood urea nitrogen (BUN) and apoptosis of tubular cells were significantly increased in WT mice. However, Tisp40 deficiency significantly attenuated the increase of sCr, BUN and apoptosis of tubular cells. Following HR, apoptosis of tubular cells was increased in Tisp40 overexpression cells compared with vector cells. Mechanistically, Tisp40 promoted the expressions of C/EBP homologous protein (CHOP), Bax and Cleaved caspase3 and suppressed the expression of Bcl-2 in renal IR injury. In conclusion, Tisp40 aggravates tubular cells apoptosis in renal IR injury. Copyright © 2017 Elsevier Inc. All rights reserved.

Bile Acid-regulated Peroxisome Proliferator-activated Receptor-α (PPARα) Activity Underlies Circadian Expression of Intestinal Peptide Absorption Transporter PepT1/Slc15a1*

PubMed Central

Okamura, Ayako; Koyanagi, Satoru; Dilxiat, Adila; Kusunose, Naoki; Chen, Jia Jun; Matsunaga, Naoya; Shibata, Shigenobu; Ohdo, Shigehiro

2014-01-01

Digested proteins are mainly absorbed as small peptides composed of two or three amino acids. The intestinal absorption of small peptides is mediated via only one transport system: the proton-coupled peptide transporter-1 (PepT1) encoded from the soluble carrier protein Slc15a1. In mammals, intestinal expression of PepT1/Slc15a1 oscillates during the daily feeding cycle. Although the oscillation in the intestinal expression of PepT1/Slc15a1 is suggested to be controlled by molecular components of circadian clock, we demonstrated here that bile acids regulated the oscillation of PepT1/Slc15a1 expression through modulating the activity of peroxisome proliferator-activated receptor α (PPARα). Nocturnally active mice mainly consumed their food during the dark phase. PPARα activated the intestinal expression of Slc15a1 mRNA during the light period, and protein levels of PepT1 peaked before the start of the dark phase. After food intake, bile acids accumulated in intestinal epithelial cells. Intestinal accumulated bile acids interfered with recruitment of co-transcriptional activator CREB-binding protein/p300 on the promoter region of Slc15a1 gene, thereby suppressing PPARα-mediated transactivation of Slc15a1. The time-dependent suppression of PPARα-mediated transactivation by bile acids caused an oscillation in the intestinal expression of PepT1/Slc15a1 during the daily feeding cycle that led to circadian changes in the intestinal absorption of small peptides. These findings suggest a molecular clock-independent mechanism by which bile acid-regulated PPARα activity governs the circadian expression of intestinal peptide transporter. PMID:25016014

Molecular mechanisms of cellular transformation by HTLV-1 Tax.

PubMed

Grassmann, Ralph; Aboud, Mordechai; Jeang, Kuan-Teh

2005-09-05

The HTLV Tax protein is crucial for viral replication and for initiating malignant transformation leading to the development of adult T-cell leukemia. Tax has been shown to be oncogenic, since it transforms and immortalizes rodent fibroblasts and human T-lymphocytes. Through CREB, NF-kappaB and SRF pathways Tax transactivates cellular promoters including those of cytokines (IL-13, IL-15), cytokine receptors (IL-2Ralpha) and costimulatory surface receptors (OX40/OX40L) leading to upregulated protein expression and activated signaling cascades (e.g. Jak/STAT, PI3Kinase, JNK). Tax also stimulates cell growth by direct binding to cyclin-dependent kinase holenzymes and/or inactivating tumor suppressors (e.g. p53, DLG). Moreover, Tax silences cellular checkpoints, which guard against DNA structural damage and chromosomal missegregation, thereby favoring the manifestation of a mutator phenotype in cells.

Daily rhythms of catalase and glutathione peroxidase expression and activity are endogenously driven in the hippocampus and are modified by a vitamin A-free diet.

PubMed

Navigatore-Fonzo, Lorena S; Delgado, Silvia M; Gimenez, Maria Sofia; Anzulovich, Ana C

2014-01-01

Alterations in enzymatic antioxidant defense systems lead to a deficit of cognitive functions and altered hippocampal synaptic plasticity. The objectives of this study were to investigate endogenous rhythms of catalase (CAT) and glutathione peroxidase (GPx) expression and activity, as well as CREB1 mRNA, in the rat hippocampus, and to evaluate to which extent the vitamin A deficiency could affect those temporal patterns. Rats from control and vitamin A-deficient (VAD) groups received a diet containing 4000 IU of vitamin A/kg diet, or the same diet devoid of vitamin A, respectively, during 3 months. Rats were maintained under 12-hour-dark conditions, during 10 days before the sacrifice. Circadian rhythms of CAT, GPx, RXRγ, and CREB1 mRNA levels were determined by reverse transcriptrase polymerase chain reaction in hippocampus samples isolated every 4 hours during a 24-hour period. CAT and GPx enzymatic activities were also determined by kinetic assays. Regulatory regions of clock and antioxidant enzymes genes were scanned for E-box, RXRE, and CRE sites. E-box, RXRE, and CRE sites were found on regulatory regions of GPx and CAT genes, which display a circadian expression in the rat hippocampus. VAD phase shifted CAT, GPx, and RXRγ endogenous rhythms without affecting circadian expression of CREB1. CAT and GPx expression and enzymatic activity are circadian in the rat hippocampus. The VAD affected the temporal patterns antioxidant genes expression, probably by altering circadian rhythms of its RXR receptors and clock factors; thus, it would impair the temporal orchestration of hippocampal daily cognitive performance.

Aqueous extracts from asparagus stems prevent memory impairments in scopolamine-treated mice.

PubMed

Sui, Zifang; Qi, Ce; Huang, Yunxiang; Ma, Shufeng; Wang, Xinguo; Le, Guowei; Sun, Jin

2017-04-19

Aqueous extracts from Asparagus officinalis L. stems (AEAS) are rich in polysaccharides, gamma-amino butyric acid (GABA), and steroidal saponin. This study was designed to investigate the effects of AEAS on learning, memory, and acetylcholinesterase-related activity in a scopolamine-induced model of amnesia. Sixty ICR mice were randomly divided into 6 groups (n = 10) including the control group (CT), scopolamine group (SC), donepezil group (DON), low, medium, and high dose groups of AEAS (LS, MS, HS; 1.6 mL kg -1 , 8 mL kg -1 , 16 mL kg -1 ). The results showed that 8 mL kg -1 of AEAS used in this study significantly reversed scopolamine-induced cognitive impairments in mice in the novel object recognition test (P < 0.05) and the Y-maze test (P < 0.05), and also improved the latency to escape in the Morris water maze test (P < 0.05). Moreover, it significantly increased acetylcholine and inhibited acetylcholinesterase activity in the hippocampus, which was directly related to the reduction in learning and memory impairments. It also reversed scopolamine-induced reduction in the hippocampal brain-derived neurotrophic factor (BDNF) and the cAMP response element-binding protein (CREB) mRNA expression. AEAS protected against scopolamine-induced memory deficits. In conclusion, AEAS protected learning and memory function in mice by enhancing the activity of the cholinergic nervous system, and increasing BDNF and CREB expression. This suggests that AEAS has the potential to prevent cognitive impairments in age-related diseases, such as Alzheimer's disease.

Differential involvement of 3', 5'-cyclic adenosine monophosphate-dependent protein kinase in regulation of Fos and tyrosine hydroxylase expression in the heart after naloxone induced morphine withdrawal.

PubMed

Almela, Pilar; Cerezo, Manuela; González-Cuello, A; Milanés, M Victoria; Laorden, M Luisa

2007-01-01

We previously demonstrated that morphine withdrawal induced hyperactivity of the heart by the activation of noradrenergic pathways innervating the left and right ventricle, as evaluated by noradrenaline (NA) turnover and Fos expression. We investigated whether cAMP-dependent protein kinase (PKA) plays a role in this process by estimating changes in PKA immunoreactivity and the influence of inhibitor of PKA on Fos protein expression, tyrosine hydroxylase (TH) immunoreactivity levels and NA turnover in the left and right ventricle. Dependence on morphine was induced by a 7-day s.c. implantation of morphine pellets. Morphine withdrawal was precipitated on day 8 by an injection of naloxone (5 mg/kg). When opioid withdrawal was precipitated, an increase in PKA immunoreactivity and phospho-CREB (cyclic AMP response element protein) levels were observed in the heart. Moreover, morphine withdrawal induces Fos expression, an enhancement of NA turnover and an increase in the total TH levels. When the selective PKA inhibitor HA-1004 was infused, concomitantly with morphine pellets, it diminished the increase in NA turnover and the total TH levels observed in morphine-withdrawn rats. However, this inhibitor neither modifies the morphine withdrawal induced Fos expression nor the increase of nonphosphorylated TH levels. The present findings indicate that an up-regulated PKA-dependent transduction pathway might contribute to the activation of the cardiac catecholaminergic neurons in response to morphine withdrawal and suggest that Fos is not a target of PKA at heart levels.

Low-level laser irradiation modulates brain-derived neurotrophic factor mRNA transcription through calcium-dependent activation of the ERK/CREB pathway.

PubMed

Yan, Xiaodong; Liu, Juanfang; Zhang, Zhengping; Li, Wenhao; Sun, Siguo; Zhao, Jian; Dong, Xin; Qian, Jixian; Sun, Honghui

2017-01-01

Low-level laser (LLL) irradiation has been reported to promote neuronal differentiation, but the mechanism remains unclear. Brain-derived neurotrophic factor (BDNF) has been confirmed to be one of the most important neurotrophic factors because it is critical for the differentiation and survival of neurons during development. Thus, this study aimed to investigate the effects of LLL irradiation on Bdnf messenger RNA (mRNA) transcription and the molecular pathway involved in LLL-induced Bdnf mRNA transcription in cultured dorsal root ganglion neurons (DRGNs) using Ca 2+ imaging, pharmacological detections, RNA interference, immunocytochemistry assay, Western blot, and qPCR analysis. We show here that LLL induced increases in the [Ca 2+ ] i level, Bdnf mRNA transcription, cAMP-response element-binding protein (CREB) phosphorylation, and extracellular signal-regulated kinase (ERK) phosphorylation, mediated by Ca 2+ release via inositol triphosphate receptor (IP3R)-sensitive calcium (Ca 2+ ) stores. Blockade of Ca 2+ increase suppressed Bdnf mRNA transcription, CREB phosphorylation, and ERK phosphorylation. Downregulation of phosphorylated (p)-CREB reduced Bdnf mRNA transcription triggered by LLL. Furthermore, blockade of ERK using PD98059 inhibitor reduced p-CREB and Bdnf mRNA transcription induced by LLL. Taken together, these findings establish the Ca 2+ -ERK-CREB cascade as a potential signaling pathway involved in LLL-induced Bdnf mRNA transcription. To our knowledge, this is the first report of the mechanisms of Ca 2+ -dependent Bdnf mRNA transcription triggered by LLL. These findings may help further explore the complex molecular signaling networks in LLL-triggered nerve regeneration in vivo and may also provide experimental evidence for the development of LLL for clinical applications.

Stress-Induced Transcriptional Regulation in the Developing Rat Brain Involves Increased Cyclic Adenosine 3′,5′-Monophosphate-Regulatory Element Binding Activity

PubMed Central

Hatalski, Carolyn G.; Baram, Tallie Z.

2012-01-01

The cAMP-regulatory element (CRE) binding protein (CREB) functions as a trans-acting regulator of genes containing the CRE sequence in their promoter. These include a number of critical genes, such as CRF, involved in the hypothalamic response to stressful stimuli in the adult. The ability of the developing rat (during the first 2 postnatal weeks) to mount the full complement of this stress response has been questioned. We have previously demonstrated the stress-induced up-regulation of the transcription of hypothalamic CRF during the second postnatal week in the rat. The focus of the current study was to explore the mechanism of transcriptional regulation in response to stress through the physiological induction of transcriptional trans-activators that bind to the CRE in the developing rat brain. CRE-binding activity was detected via gel shift analysis in extracts from both the hypothalamus and the cerebral cortex of the developing rat. CREB was identified in these extracts by Western blot analysis and was shown to be the major contributor to the CRE-binding activity by gel shift analysis with two specific antibodies directed against CREB. After acute hypothermic stress, the abundance of CRE-binding activity (but not of total immunoreactive CREB), increased in hypothalamic extracts. This enhanced CRE-binding activity was blocked by an antiserum directed against CREB and was accompanied by an apparent increase in CREB phosphorylation. These results indicate that posttranslational enhancement of CRE-binding activity is likely to constitute an important mechanism for up-regulation of genes possessing the CRE sequence in the developing rat hypothalamus by adverse external signals. PMID:9415405

Differences in the Flexibility of Switching Learning Strategies and CREB Phosphorylation Levels in Prefrontal Cortex, Dorsal Striatum and Hippocampus in Two Inbred Strains of Mice

PubMed Central

Cho, Woo-Hyun; Han, Jung-Soo

2016-01-01

Flexibility in using different learning strategies was assessed in two different inbred strains of mice, the C57BL/6 and DBA/2 strains. Mice were trained sequentially in two different Morris water maze protocols that tested their ability to switch their learning strategy to complete a new task after first being trained in a different task. Training consisted either of visible platform trials (cued training) followed by subsequent hidden platform trials (place training) or the reverse sequence (place training followed by cued training). Both strains of mice showed equivalent performance in the type of training (cued or place) that they received first. However, C57BL/6 mice showed significantly better performances than DBA/2 mice following the switch in training protocols, irrespective of the order of training. After completion of the switched training session, levels of cAMP response element-binding protein (CREB) and phosphorylated CREB (pCREB) were measured in the hippocampus, striatum and prefrontal cortex of the mice. Prefrontal cortical and hippocampal pCREB levels differed by strain, with higher levels found in C57BL/6 mice than in DBA/2 mice. No strain differences were observed in the medial or lateral region of the dorsal striatum. These findings indicate that the engagement (i.e., CREB signaling) of relevant neural structures may vary by the specific demands of the learning strategy, and this is closely tied to differences in the flexibility of C57BL/6 and DBA/2 mice to switch their learning strategies when given a new task. PMID:27695401

Cognitive-Enhancing Effect of Aronia melanocarpa Extract against Memory Impairment Induced by Scopolamine in Mice

PubMed Central

Lee, Hyeon Yong; Weon, Jin Bae; Jung, Youn Sik; Kim, Nam Young; Kim, Myong Ki; Ma, Choong Je

2016-01-01

Aronia melanocarpa (A. melanocarpa) berries are a fruit with a marked antioxidant effect. The objective of this study was to confirm the effect of A. melanocarpa berries extract against scopolamine-induced memory impairment in mice using the Morris water maze and passive avoidance test. Moreover, we determined a possible mechanism of the cognitive-enhancing effect involving AChE activity and BDNF and p-CREB expression in the hippocampus of mice. A. melanocarpa berries extract attenuated the learning and memory impairment induced by scopolamine in the Morris water maze (79.3 ± 0.8 s of 200 mg/kg and 64.4 ± 10.7 s of 400 mg/kg on day 4) and passive avoidance tests (46.0 ± 41.1 s of 200 mg/kg and 25.6 ± 18.7 s of 400 mg/kg). A. melanocarpa berries extract reduced the acetylcholinesterase level in the hippocampus of scopolamine-injected mice and increased BDNF and p-CREB expression in the hippocampus. The major compound, cyanidin-3-O-galactoside, also reversed memory impairment. These results showed that A. melanocarpa berries extract improved memory impairment by inhibiting AChE and increasing BDNF and p-CREB expression, and cyanidin-3-O-galactoside may be responsible for the effect of A. melanocarpa berries extract. PMID:27239211

Cognitive-Enhancing Effect of Aronia melanocarpa Extract against Memory Impairment Induced by Scopolamine in Mice.

PubMed

Lee, Hyeon Yong; Weon, Jin Bae; Jung, Youn Sik; Kim, Nam Young; Kim, Myong Ki; Ma, Choong Je

2016-01-01

Aronia melanocarpa (A. melanocarpa) berries are a fruit with a marked antioxidant effect. The objective of this study was to confirm the effect of A. melanocarpa berries extract against scopolamine-induced memory impairment in mice using the Morris water maze and passive avoidance test. Moreover, we determined a possible mechanism of the cognitive-enhancing effect involving AChE activity and BDNF and p-CREB expression in the hippocampus of mice. A. melanocarpa berries extract attenuated the learning and memory impairment induced by scopolamine in the Morris water maze (79.3 ± 0.8 s of 200 mg/kg and 64.4 ± 10.7 s of 400 mg/kg on day 4) and passive avoidance tests (46.0 ± 41.1 s of 200 mg/kg and 25.6 ± 18.7 s of 400 mg/kg). A. melanocarpa berries extract reduced the acetylcholinesterase level in the hippocampus of scopolamine-injected mice and increased BDNF and p-CREB expression in the hippocampus. The major compound, cyanidin-3-O-galactoside, also reversed memory impairment. These results showed that A. melanocarpa berries extract improved memory impairment by inhibiting AChE and increasing BDNF and p-CREB expression, and cyanidin-3-O-galactoside may be responsible for the effect of A. melanocarpa berries extract.

Dual Role of Protein Phosphorylation in DNA Activator/Coactivator Binding

PubMed Central

Dadarlat, Voichita M.; Skeel, Robert D.

2011-01-01

Binding free energies are calculated for the phosphorylated and unphosphorylated complexes between the kinase inducible domain (KID) of the DNA transcriptional activator cAMP response element binding (CREB) protein and the KIX domain of its coactivator, CREB-binding protein (CBP). To our knowledge, this is the first application of a method based on a potential of mean force (PMF) with restraining potentials to compute the binding free energy of protein-protein complexes. The KID:KIX complexes are chosen here because of their biological relevance to the DNA transcription process and their relatively small size (81 residues for the KIX domain of CBP, and 28 residues for KID). The results for pKID:KIX and KID:KIX are −9.55 and −4.96 kcal/mol, respectively, in good agreement with experimental estimates (−8.8 and −5.8 kcal/mol, respectively). A comparison between specific contributions to protein-protein binding for the phosphorylated and unphosphorylated complexes reveals a dual role for the phosphorylation of KID at Ser-133 in effecting a more favorable free energy of the bound system: 1), stabilization of the unbound conformation of phosphorylated KID due to favorable intramolecular interactions of the phosphate group of Ser-133 with the charged groups of an arginine-rich region spanning both α-helices, which lowers the configurational entropy; and 2), more favorable intermolecular electrostatic interactions between pSer-133 and Arg-131 of KID, and Lys-662, Tyr-658, and Glu-666 of KIX. Charge reduction through ligand phosphorylation emerges as a possible mechanism for controlling the unbound state conformation of KID and, ultimately, gene expression. This work also demonstrates that the PMF-based method with restraining potentials provides an added benefit in that important elements of the binding pathway are evidenced. Furthermore, the practicality of the PMF-based method for larger systems is validated by agreement with experiment. In addition, we provide a somewhat differently structured exposition of the PMF-based method with restraining potentials and outline its generalization to systems in which both protein and ligand may adopt unbound conformations that are different from those of the bound state. PMID:21244843

Continuous activation of pituitary adenylate cyclase-activating polypeptide receptors elicits antipodal effects on cyclic AMP and inositol phospholipid signaling pathways in CATH.a cells: role of protein synthesis and protein kinases.

PubMed

Muller, A; Lutz-Bucher, B; Kienlen-Campard, P; Koch, B; Loeffler, J P

1998-04-01

Continuous exposure of cells to agonists develops a process that determines the extent to which the cells eventually respond to further stimuli. Here we used CATH.a cells (a catecholaminergic neuron-like cell line), which express pituitary adenylate cyclase-activating polypeptide (PACAP) receptors linked to both adenylyl cyclase and phospholipase C-beta pathways, to investigate the influence of prolonged hormonal treatment on dual signaling and gene transcription. Prolonged incubation of cells with PACAP failed to down-regulate the density and affinity of membrane binding sites and caused opposite changes in messenger systems: PACAP-stimulated cyclic AMP accumulation was attenuated in a time- and dose-dependent fashion (t(1/2) = 6.7 h and IC50 = 0.1 nM), whereas phosphoinositide turnover was overstimulated. Both effects were insensitive to pertussis toxin, whereas the drop in cyclic AMP concentration was also unchanged in the presence of 3-isobutyl-1-methylxanthine, indicating that neither Gi-like proteins nor cyclic nucleotide phosphodiesterases play a critical role in these processes. Blockade of protein synthesis with cycloheximide, as well as inhibition by H89 of cyclic AMP-dependent protein kinase (but not by bisindolylmaleimide of protein kinase C) antagonized the influences exerted by PACAP on adenylyl cyclase activity and inositol phosphate formation. Transcription of the chimeric GAL4-CREB construct, transiently transfected into CATH.a cells, was stimulated by PACAP, and this effect was potentiated as a result of chronic PACAP treatment. The results of the present investigation provide new insight into the possible differential regulation and cross-talks of transduction signals of receptors linked to multiplex signaling. They demonstrate that prolonged exposure of CATH.a cells to PACAP results in the desensitization of the cyclic AMP pathway and superinduction of the inositol phosphate signal, through protein neosynthesis and cyclic AMP-dependent protein kinase activation. At the same time, they show that desensitization of cyclic AMP signaling not only fails to hamper, but actually amplifies PACAP-stimulated CREB-regulated transcription.

Essential Oils from the Medicinal Herbs Upregulate Dopamine Transporter in Rat Pheochromocytoma Cells.

PubMed

Choi, Min Sun; Choi, Bang-sub; Kim, Sang Heon; Pak, Sok Cheon; Jang, Chul Ho; Chin, Young-Won; Kim, Young-Mi; Kim, Dong-il; Jeon, Songhee; Koo, Byung-Soo

2015-10-01

The dopamine transporter (DAT) protein, a component of the dopamine system, undergoes adaptive neurobiological changes from drug abuse. Prevention of relapse and reduction of withdrawal symptoms are still the major limitations in the current pharmacological treatments of drug addiction. The present study aimed to investigate the effects of essential oils extracted from Elsholtzia ciliata, Shinchim, Angelicae gigantis Radix, and Eugenia caryophyllata, well-known traditional Korean medicines for addiction, on the modulation of dopamine system in amphetamine-treated cells and to explore the possible mechanism underlying its therapeutic effect. The potential cytotoxic effect of essential oils was evaluated in PC12 rat pheochromocytoma cells using cell viability assays. Quantification of DAT, p-CREB, p-MAPK, and p-Akt was done by immunoblotting. DAT was significantly reduced in cells treated with 50 μM of amphetamine in a time-dependent manner. No significant toxicity of essential oils from Elsholtzia ciliata and Shinchim was observed at doses of 10, 25, and 50 μg/mL. However, essential oils from A. gigantis Radix at a dose of 100 μg/mL and E. caryophyllata at doses of 50 and 100 μg/mL showed cytotoxicity. Treatment with GBR 12909, a highly selective DAT inhibitor, significantly increased DAT expression compared with that of amphetamine only by enhancing phosphorylation of mitogen-activated protein kinases (MAPK) and Akt. In addition, essential oils effectively induced hyperphosphorylation of cyclic-AMP response element-binding protein (CREB), MAPK, and Akt, which resulted in DAT upregulation. Our study implies that the essential oils may rehabilitate brain dopamine function through increased DAT availability in abstinent former drug users.