Transcription factor HBP1 is a direct anti-cancer target of transcription factor FOXO1 in invasive oral cancer.

PubMed

Chan, Chien-Yi; Huang, Shih-Yi; Sheu, Jim Jinn-Chyuan; Roth, Mendel M; Chou, I-Tai; Lien, Chia-Hsien; Lee, Ming-Fen; Huang, Chun-Yin

2017-02-28

Either FOXO1 or HBP1 transcription factor is a downstream effector of the PI3K/Akt pathway and associated with tumorigenesis. However, the relationship between FOXO1 and HBP1 in oral cancer remains unclear. Analysis of 30 oral tumor specimens revealed that mean mRNA levels of both FOXO1 and HBP1 in non-invasive and invasive oral tumors were found to be significantly lower than that of the control tissues, and the status of low FOXO1 and HBP1 (< 0.3 fold of the control) was associated with invasiveness of oral tumors. To investigate if HBP1 is a direct transcription target of FOXO1, we searched potential FOXO1 binding sites in the HBP1 promoter using the MAPPER Search Engine, and two putative FOXO1 binding sites located in the HBP1 promoter -132 to -125 bp and -343 to -336 bp were predicted. These binding sites were then confirmed by both reporter gene assays and the in cellulo ChIP assay. In addition, Akt activity manipulated by PI3K inhibitor LY294002 or Akt mutants was shown to negatively affect FOXO1-mediated HBP1 promoter activation and gene expression. Last, the biological significance of the FOXO1-HBP1 axis in oral cancer malignancy was evaluated in cell growth, colony formation, and invasiveness. The results indicated that HBP1 knockdown potently promoted malignant phenotypes of oral cancer and the suppressive effect of FOXO1 on cell growth, colony formation, and invasion was alleviated upon HBP1 knockdown in invasive oral cancer cells. Taken together, our data provide evidence for HBP1 as a direct downstream target of FOXO1 in oral cancer malignancy.

FOXO1 is a direct target of EWS-Fli1 oncogenic fusion protein in Ewing's sarcoma cells

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

Yang, Liu, E-mail: lyang@u.washington.edu; Medical Research Service, VA Puget Sound Health Care System, Seattle, WA 98108; Hu, Hsien-Ming

2010-11-05

Research highlights: {yields} Inducible and reversible siRNA knockdown of an oncogenic fusion protein such as EWS-Fli1 is feasible and more advantageous than other siRNA methods. {yields} The tumor suppressor gene FOXO1 is a new EWS-Fli1 target. {yields} While trans-activators are known for the FOXO1 gene, there has been no report on negative regulators of FOXO1 transcription. {yields} This study provides first evidence that the EWS-Fli1 oncogenic fusion protein can function as a transcriptional repressor of the FOXO1 gene. -- Abstract: Ewing's family tumors are characterized by a specific t(11;22) chromosomal translocation that results in the formation of EWS-Fli1 oncogenic fusionmore » protein. To investigate the effects of EWS-Fli1 on gene expression, we carried out DNA microarray analysis after specific knockdown of EWS-Fli1 through transfection of synthetic siRNAs. EWS-Fli1 knockdown increased expression of genes such as DKK1 and p57 that are known to be repressed by EWS-Fli1 fusion protein. Among other potential EWS-Fli1 targets identified by our microarray analysis, we have focused on the FOXO1 gene since it encodes a potential tumor suppressor and has not been previously reported in Ewing's cells. To better understand how EWS-Fli1 affects FOXO1 expression, we have established a doxycycline-inducible siRNA system to achieve stable and reversible knockdown of EWS-Fli1 in Ewing's sarcoma cells. Here we show that FOXO1 expression in Ewing's cells has an inverse relationship with EWS-Fli1 protein level, and FOXO1 promoter activity is increased after doxycycline-induced EWS-Fli1 knockdown. In addition, we have found that direct binding of EWS-Fli1 to FOXO1 promoter is attenuated after doxycycline-induced siRNA knockdown of the fusion protein. Together, these results suggest that suppression of FOXO1 function by EWS-Fli1 fusion protein may contribute to cellular transformation in Ewing's family tumors.« less

Isorhapontigenin (ISO) Inhibits Invasive Bladder Cancer Formation In Vivo and Human Bladder Cancer Invasion In Vitro by Targeting STAT1/FOXO1 Axis.

PubMed

Jiang, Guosong; Wu, Amy D; Huang, Chao; Gu, Jiayan; Zhang, Liping; Huang, Haishan; Liao, Xin; Li, Jingxia; Zhang, Dongyun; Zeng, Xingruo; Jin, Honglei; Huang, Haojie; Huang, Chuanshu

2016-07-01

Although our most recent studies have identified Isorhapontigenin (ISO), a novel derivative of stilbene that isolated from a Chinese herb Gnetum cleistostachyum, for its inhibition of human bladder cancer growth, nothing is known whether ISO possesses an inhibitory effect on bladder cancer invasion. Thus, we addressed this important question in current study and discovered that ISO treatment could inhibit mouse-invasive bladder cancer development following bladder carcinogen N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) exposure in vivo We also found that ISO suppressed human bladder cancer cell invasion accompanied by upregulation of the forkhead box class O 1 (FOXO1) mRNA transcription in vitro Accordingly, FOXO1 was profoundly downregulated in human bladder cancer tissues and was negatively correlated with bladder cancer invasion. Forced expression of FOXO1 specifically suppressed high-grade human bladder cancer cell invasion, whereas knockdown of FOXO1 promoted noninvasive bladder cancer cells becoming invasive bladder cancer cells. Moreover, knockout of FOXO1 significantly increased bladder cancer cell invasion and abolished the ISO inhibition of invasion in human bladder cancer cells. Further studies showed that the inhibition of Signal transducer and activator of transcription 1 (STAT1) phosphorylation at Tyr701 was crucial for ISO upregulation of FOXO1 transcription. Furthermore, this study revealed that metalloproteinase-2 (MMP-2) was a FOXO1 downstream effector, which was also supported by data obtained from mouse model of ISO inhibition BBN-induced mouse-invasive bladder cancer formation. These findings not only provide a novel insight into the understanding of mechanism of bladder cancer's propensity to invasion, but also identify a new role and mechanisms underlying the natural compound ISO that specifically suppresses such bladder cancer invasion through targeting the STAT1-FOXO1-MMP-2 axis. Cancer Prev Res; 9(7); 567-80. ©2016 AACR. ©2016 American

The inhibition of activated hepatic stellate cells proliferation by arctigenin through G0/G1 phase cell cycle arrest: persistent p27(Kip1) induction by interfering with PI3K/Akt/FOXO3a signaling pathway.

PubMed

Li, Ao; Wang, Jun; Wu, Mingjun; Zhang, Xiaoxun; Zhang, Hongzhi

2015-01-15

Proliferation of hepatic stellate cells (HSCs) is vital for the development of fibrosis during liver injury. In this study, we describe that arctigenin (ATG), a major bioactive component of Fructus Arctii, exhibited selective cytotoxic activity via inhibiting platelet-derived growth factor-BB (PDGF-BB)-activated HSCs proliferation and arrested cell cycle at G0/G1 phase, which could not be observed in normal human hepatocytes in vitro. The cyclin-dependent kinase (CDK) 4/6 activities could be strongly inhibited by ATG through down-regulation of cyclin D1 and CDK4/6 expression in early G1 phase arrest. In the ATG-treated HSCs, the expression level of p27(Kip1) and the formation of CDK2-p27(Kip1) complex were also increased. p27(Kip1) silencing significantly attenuated the effect of ATG, including cell cycle arrest and suppression of proliferation in activated HSCs. We also found that ATG suppressed PDGF-BB-induced phosphorylation of Akt and its downstream transcription factor Forkhead box O 3a (FOXO3a), decreased binding of FOXO3a to 14-3-3 protein, and stimulated nuclear translocation of FOXO3a in activated HSCs. Furthermore, knockdown of FOXO3a expression by FOXO3a siRNA attenuated ATG-induced up-regulation of p27(Kip1) in activated HSCs. All the above findings suggested that ATG could increase the levels of p27(Kip1) protein through inhibition of Akt and improvement of FOXO3a activity, in turn inhibited the CDK2 kinase activity, and eventually caused an overall inhibition of HSCs proliferation. Copyright © 2014 Elsevier B.V. All rights reserved.

SMAD3 augments FoxO3-induced MuRF-1 promoter activity in a DNA-binding-dependent manner

PubMed Central

Bollinger, Lance M.; Witczak, Carol A.; Houmard, Joseph A.

2014-01-01

Muscle-specific RING finger-1 (MuRF-1), a ubiquitin ligase and key regulator of proteasome-dependent protein degradation, is highly expressed during skeletal muscle atrophy. The transcription factor forkhead box O3 (FoxO3) induces MuRF-1 expression, but the direct role of other major atrophy-related transcription factors, such as SMAD3, is largely unknown. The goal of this study was to determine whether SMAD3 individually regulates, or with FoxO3 coordinately regulates, MuRF-1 expression. In cultured myotubes or human embryonic kidney cells, MuRF-1 mRNA content and promoter activity were increased by FoxO3 but not by SMAD3 overexpression. However, FoxO3 and SMAD3 coexpression synergistically increased MuRF-1 mRNA and promoter activity. Mutation of the SMAD-binding element (SBE) in the proximal MuRF-1 promoter or overexpression of a SMAD3 DNA-binding mutant attenuated FoxO3-dependent MuRF-1 promoter activation, showing that SMAD binding to DNA is required for optimal activation of FoxO3-induced transcription of MuRF-1. Using chromatin immunoprecipitation, SMAD3 DNA binding increased FoxO3 abundance and SBE mutation reduced FoxO3 abundance on the MuRF-1 promoter. Furthermore, SMAD3 overexpression dose-dependently increased FoxO3 protein content, and coexpression of FoxO3 and SMAD3 synergistically increased FoxO-dependent gene transcription [assessed with a FoxO response element (FRE)-driven reporter]. Collectively, these results show that SMAD3 regulates transcription of MuRF-1 by increasing FoxO3 binding at a conserved FRE-SBE motif within the proximal promoter region, and by increasing FoxO3 protein content and transcriptional activity. These data are the first to indicate that two major transcription factors regulating protein degradation, FoxO3 and SMAD3, converge to coordinately and directly regulate transcription of MuRF-1. PMID:24920680

PTEN regulates p300-dependent hypoxia-inducible factor 1 transcriptional activity through Forkhead transcription factor 3a (FOXO3a)

PubMed Central

Emerling, Brooke M.; Weinberg, Frank; Liu, Juinn-Lin; Mak, Tak W.; Chandel, Navdeep S.

2008-01-01

The tumor suppressor PTEN is mutated or deleted in many tumors, causing the activation of the PI3K pathway. Here, we show that the loss of PTEN increases the transcriptional activity of hypoxia-inducible factor 1 (HIF-1) through the inactivation of Forkhead transcription factors (FOXO) in PTEN-null cells. Reintroduction of PTEN into the nucleus, overexpression of a nonphosphorylatable FOXO3a, which accumulates in the nucleus, or inhibition of nuclear export of FOXO3a by leptomycin B represses HIF-1 transcriptional activity in PTEN-null cells. HIF-1 transcriptional activity increases in PTEN-positive cells depleted of FOXO3a with siRNA. PTEN and FOXO3a regulate the transactivation domain of HIF-1α. Chromatin immunoprecipitation indicates that FOXO3a complexes with HIF-1α and p300 on the Glut-1 promoter, a HIF-1 target gene. Overexpression of p300 reverses FOXO3a-mediated repression of HIF-1 transcriptional activity. Coimmunoprecipitation and GAL4-HIF-1α transactivation assays reveal that FOXO3a interferes with p300-dependent HIF-1 transcriptional activity. Thus, FOXO3a negatively regulates HIF-1 transcriptional activity. PMID:18268343

Parecoxib suppresses CHOP and Foxo1 nuclear translocation, but increases GRP78 levels in a rat model of focal ischemia.

PubMed

Ye, Zhi; Wang, Na; Xia, Pingping; Wang, E; Liao, Juan; Guo, Qulian

2013-04-01

Parecoxib, a novel COX-2 inhibitor, functions as a neuroprotective agent and rescues neurons from cerebral ischemic reperfusion injury-induced apoptosis. However, the molecular mechanisms underlying parecoxib neuroprotection remain to be elucidated. There is growing evidence that endoplasmic reticulum (ER) stress plays an important role in neuronal death caused by brain ischemia. However, very little is known about the role of parecoxib in mediating pathophysiological reactions to ER stress induced by ischemic reperfusion injury. Therefore, in the present study, we investigated whether delayed administration of parecoxib attenuates brain damage via suppressing ER stress-induced cell death. Adult male Sprague-Dawley rats were administered parecoxib (10 or 30 mg kg(-1), IP) or isotonic saline twice a day starting 24 h after middle cerebral artery occlusion (MCAO) for three consecutive days. The expressions of glucose-regulated protein 78 (GRP78) and oxygen-regulated protein 150 (ORP150) and C/EBP-homologous protein (CHOP) and forkhead box protein O 1 (Foxo1) in cytoplasmic and nuclear fraction were determined by Western blotting. The levels of caspase-12 expression were checked by immunohistochemistry analysis, served as a marker for ER stress-induced apoptosis. Parecoxib significantly suppressed cerebral ischemic injury-induced nuclear translocation of CHOP and Foxo1 and attenuated the immunoreactivity of caspase-12 in ischemic penumbra. Furthermore, the protective effect of delayed administration of parecoxib was accompanied by an increased GRP78 and ORP150 expression. Therefore, our study suggested that elevation of GRP78 and ORP150, and suppression of CHOP and Foxo1 nuclear translocation may contribute to parecoxib-mediated neuroprotection during ER stress responses.

GSK3 Protein Positively Regulates Type I Insulin-like Growth Factor Receptor through Forkhead Transcription Factors FOXO1/3/4

PubMed Central

Huo, Xiaodong; Liu, Shu; Shao, Ting; Hua, Hui; Kong, Qingbin; Wang, Jiao; Luo, Ting; Jiang, Yangfu

2014-01-01

Glycogen synthase kinase-3 (GSK3) has either tumor-suppressive roles or pro-tumor roles in different types of human tumors. A number of GSK3 targets in diverse signaling pathways have been uncovered, such as tuberous sclerosis complex subunit 2 and β-catenin. The O subfamily of forkhead/winged helix transcription factors (FOXO) is known as tumor suppressors that induce apoptosis. In this study, we find that FOXO binds to type I insulin-like growth factor receptor (IGF-IR) promoter and stimulates its transcription. GSK3 positively regulates the transactivation activity of FOXO and stimulates IGF-IR expression. Although kinase-dead GSK3β cannot up-regulate IGF-IR, the constitutively active GSK3β induces IGF-IR expression in a FOXO-dependent manner. Serum starvation or Akt inhibition leads to an increase in IGF-IR expression, which could be blunted by GSK3 inhibition. GSK3β knockdown or GSK3 inhibitor suppresses IGF-I-induced IGF-IR, Akt, and ERK1/2 phosphorylation. Moreover, knockdown of GSK3β or FOXO1/3/4 leads to a decrease in cellular proliferation and abrogates IGF-I-induced hepatoma cell proliferation. These results suggest that GSK3 and FOXO may positively regulate IGF-I signaling and hepatoma cell proliferation. PMID:25053419

Foxo3 activity promoted by non-coding effects of circular RNA and Foxo3 pseudogene in the inhibition of tumor growth and angiogenesis.

PubMed

Yang, W; Du, W W; Li, X; Yee, A J; Yang, B B

2016-07-28

It has recently been shown that the upregulation of a pseudogene specific to a protein-coding gene could function as a sponge to bind multiple potential targeting microRNAs (miRNAs), resulting in increased gene expression. Similarly, it was recently demonstrated that circular RNAs can function as sponges for miRNAs, and could upregulate expression of mRNAs containing an identical sequence. Furthermore, some mRNAs are now known to not only translate protein, but also function to sponge miRNA binding, facilitating gene expression. Collectively, these appear to be effective mechanisms to ensure gene expression and protein activity. Here we show that expression of a member of the forkhead family of transcription factors, Foxo3, is regulated by the Foxo3 pseudogene (Foxo3P), and Foxo3 circular RNA, both of which bind to eight miRNAs. We found that the ectopic expression of the Foxo3P, Foxo3 circular RNA and Foxo3 mRNA could all suppress tumor growth and cancer cell proliferation and survival. Our results showed that at least three mechanisms are used to ensure protein translation of Foxo3, which reflects an essential role of Foxo3 and its corresponding non-coding RNAs.

Active FOXO1 is a Key Determinant of Isoform-Specific Progesterone Receptor Transactivation and Senescence Programming

PubMed Central

Diep, Caroline H.; Knutson, Todd P.; Lange, Carol A.

2015-01-01

Progesterone promotes differentiation coupled to proliferation and pro-survival in the breast, but inhibits estrogen-driven growth in the reproductive tract and ovaries. Herein, it is demonstrated, using progesterone receptor (PR) isoform-specific ovarian cancer model systems, that PR-A and PR-B promote distinct gene expression profiles that differ from PR-driven genes in breast cancer cells. In ovarian cancer models, PR-A primarily regulates genes independently of progestin, while PR-B is the dominant ligand-dependent isoform. Notably, FOXO1 and the PR/FOXO1 target-gene p21 (CDKN1A) are repressed by PR-A, but induced by PR-B. In the presence of progestin, PR-B, but not PR-A, robustly induced cellular senescence via FOXO1-dependent induction of p21 and p15 (CDKN2B). Chromatin immunoprecipitation (ChIP) assays performed on PR-isoform specific cells demonstrated that while each isoform is recruited to the same PRE-containing region of the p21 promoter in response to progestin, only PR-B elicits active chromatin marks. Overexpression of constitutively active FOXO1 in PR-A-expressing cells conferred robust ligand-dependent upregulation of the PR-B target genes GZMA, IGFBP1, and p21, and induced cellular senescence. In the presence of endogenous active FOXO1, PR-A was phosphorylated on Ser294 and transactivated PR-B at PR-B target genes; these events were blocked by the FOXO1 inhibitor (AS1842856). PR isoform-specific regulation of the FOXO1/p21 axis recapitulated in human primary ovarian tumor explants treated with progestin; loss of progestin sensitivity correlated with high AKT activity. PMID:26577046

PDK-1/FoxO1 pathway in POMC neurons regulates Pomc expression and food intake.

PubMed

Iskandar, Kristy; Cao, Yongheng; Hayashi, Yoshitake; Nakata, Masanori; Takano, Eisuke; Yada, Toshihiko; Zhang, Changliang; Ogawa, Wataru; Oki, Miyo; Chua, Streamson; Itoh, Hiroshi; Noda, Tetsuo; Kasuga, Masato; Nakae, Jun

2010-04-01

Both insulin and leptin signaling converge on phosphatidylinositol 3-OH kinase [PI(3)K]/3-phosphoinositide-dependent protein kinase-1 (PDK-1)/protein kinase B (PKB, also known as Akt) in proopiomelanocortin (POMC) neurons. Forkhead box-containing protein-O1 (FoxO1) is inactivated in a PI(3)K-dependent manner. However, the interrelationship between PI(3)K/PDK-1/Akt and FoxO1, and the chronic effects of the overexpression of FoxO1 in POMC neurons on energy homeostasis has not been elucidated. To determine the extent to which PDK-1 and FoxO1 signaling in POMC neurons was responsible for energy homeostasis, we generated POMC neuron-specific Pdk1 knockout mice (POMCPdk1(-/-)) and mice selectively expressing a constitutively nuclear (CN)FoxO1 or transactivation-defective (Delta256)FoxO1 in POMC neurons (CNFoxO1(POMC) or Delta256FoxO1(POMC)). POMCPdk1(-/-) mice showed increased food intake and body weight accompanied by decreased expression of Pomc gene. The CNFoxO1(POMC) mice exhibited mild obesity and hyperphagia compared with POMCPdk1(-/-) mice. Although expression of the CNFoxO1 made POMCPdk1(-/-) mice more obese due to excessive suppression of Pomc gene, overexpression of Delta256FoxO1 in POMC neurons had no effects on metabolic phenotypes and Pomc expression levels of POMCPdk1(-/-) mice. These data suggest a requirement for PDK-1 and FoxO1 in transcriptional regulation of Pomc and food intake.

Upregulation of FoxO 1 Signaling Mediates the Proinflammatory Cytokine Upregulation in the Macrophage from Polycystic Ovary Syndrome Patients.

PubMed

Li, Ning; Wang, Xiaoyan; Wang, Xiaojie; Yu, Hongna; Lin, Li; Sun, Chengming; Liu, Peng; Chu, Yongli; Hou, Jianqing

2017-02-01

Chronic activation of macrophage-mediated inflammatory signals in insulin-sensitive metabolic tissues is thought to be one of the causes of insulin resistance-one of the hallmarks of the metabolic syndrome. Insulin resistance is a feature of polycystic ovary syndrome (PCOS) and is related to mitochondrial and endothelial function. In the present study, we investigated the phosphorylation level of FoxO 1, which is suppressed by the action of AKT, triggers the TLR4 inflammatory signaling pathway in the macrophages, from polycystic ovary syndrome patients or normal subjects. Then we investigated the influence of phosphorylation level of FoxO 1FoxO 1 on the induction of proinflammatory cytokines in the macrophages and the influence by FoxO FoxO 1 knockdown on the insulin-induced glucose uptake in PCOS macrophages. Our results demonstrated that the significantly high level of FoxO 1FoxO 1 phosphorylation correlated with the production of proinflammatory cytokines, such as IL-6, IL-1β, and TNF-α in the macrophages from PCOS patients. The high level of FoxO 1FoxO 1 phosphorylation enhanced the TLR-4 signaling in response to LPS, and the FoxO FoxO 1 knockdown inhibited the insulin-induced glucose uptake in PCOS macrophages. The findings of this paper suggest an intriguing regulatory transcriptional/signaling loop in macrophages that may contribute to maintain and exacerbate inflammation and insulin resistance in PCOS macrophages.

Inhibition of FOXO1/3 promotes vascular calcification.

PubMed

Deng, Liang; Huang, Lu; Sun, Yong; Heath, Jack M; Wu, Hui; Chen, Yabing

2015-01-01

Vascular calcification is a characteristic feature of atherosclerosis, diabetes mellitus, and end-stage renal disease. We have demonstrated that activation of protein kinase B (AKT) upregulates runt-related transcription factor 2 (Runx2), a key osteogenic transcription factor that is crucial for calcification of vascular smooth muscle cells (VSMC). Using mice with SMC-specific deletion of phosphatase and tensin homolog (PTEN), a major negative regulator of AKT, the present studies uncovered a novel molecular mechanism underlying PTEN/AKT/FOXO (forkhead box O)-mediated Runx2 upregulation and VSMC calcification. SMC-specific PTEN deletion mice were generated by crossing PTEN floxed mice with SM22α-Cre transgenic mice. The PTEN deletion resulted in sustained activation of AKT that upregulated Runx2 and promoted VSMC calcification in vitro and arterial calcification ex vivo. Runx2 knockdown did not affect proliferation but blocked calcification of the PTEN-deficient VSMC, suggesting that PTEN deletion promotes Runx2-depedent VSMC calcification that is independent of proliferation. At the molecular level, PTEN deficiency increased the amount of Runx2 post-transcriptionally by inhibiting Runx2 ubiquitination. AKT activation increased phosphorylation of FOXO1/3 that led to nuclear exclusion of FOXO1/3. FOXO1/3 knockdown in VSMC phenocopied the PTEN deficiency, demonstrating a novel function of FOXO1/3, as a downstream signaling of PTEN/AKT, in regulating Runx2 ubiquitination and VSMC calcification. Using heterozygous SMC-specific PTEN-deficient mice and atherogenic ApoE(-/-) mice, we further demonstrated AKT activation, FOXO phosphorylation, and Runx2 ubiquitination in vascular calcification in vivo. Our studies have determined a new causative effect of SMC-specific PTEN deficiency on vascular calcification and demonstrated that FOXO1/3 plays a crucial role in PTEN/AKT-modulated Runx2 ubiquitination and VSMC calcification. © 2014 American Heart Association, Inc.

Nodal enhances the activity of FoxO3a and its synergistic interaction with Smads to regulate cyclin G2 transcription in ovarian cancer cells.

PubMed

Fu, G; Peng, C

2011-09-15

Nodal, a member of the transforming growth factor-β superfamily, has been recently shown to suppress cell proliferation and to stimulate the expression of cyclin G2 (CCNG2) in human epithelial ovarian cancer cells. However, the precise mechanisms underlying these events are not fully understood. In this study, we investigated the transcriptional regulation of CCNG2 by the Nodal signaling pathway. In ovarian cancer cells, overexpression of Nodal or its receptors, activin receptor-like kinase 7 (ALK7) or ALK4, resulted in an increase in the CCNG2 promoter activity. Several putative Forkhead box class O (FoxO)3a-binding sites are present in the human CCNG2 promoter and overexpression of FoxO3a enhanced the CCNG2 promoter activity. The functional FoxO3a-binding element (FBE) was mapped to a proximal region located between -398 and -380 bp (FBE1) through deletion and mutation analyses, as well as chromatin immunoprecipitation (IP) assay. Interestingly, mutation of the FBE1 not only abolished the effect of FoxO3a, but also blocked Nodal-induced CCNG2 transcription. Nodal stimulated FoxO3a mRNA and protein expression through the canonical Smad pathway and suppressed FoxO3a inactivation by inhibiting AKT activity. Silencing of FoxO3a using small interfering RNA significantly reduced the effect of Nodal on the CCNG2 promoter activity. On the other hand, overexpression of Smad2 and Smad3 enhanced the FoxO3a-induced CCNG2 promoter activity whereas knockdown of Smad4 blocked the activity of FoxO3a. Furthermore, IP assays revealed that FoxO3a formed complexes with Smad proteins and that Nodal enhanced the binding of FoxO3a to the CCNG2 promoter. Finally, silencing of FoxO3a reversed the inhibitory effect of Nodal on cell proliferation. Taken together, these findings demonstrated that Nodal signaling promotes CCNG2 transcription by upregulating FoxO3a expression, inhibiting FoxO3a phosphorylation and enhancing its synergistic interaction with Smads. These results also suggest

Regulation of FOXO1-mediated transcription and cell proliferation by PARP-1

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

Sakamaki, Jun-ichi; Daitoku, Hiroaki; Yoshimochi, Kenji

2009-05-08

Forkhead box O (FOXO) transcription factors play an important role in a wide range of biological processes, including cell cycle control, apoptosis, detoxification of reactive oxygen species, and gluconeogenesis through regulation of gene expression. In this study, we demonstrated that PARP-1 functions as a negative regulator of FOXO1. We showed that PARP-1 directly binds to and poly(ADP-ribosyl)ates FOXO1 protein. PARP-1 represses FOXO1-mediated expression of cell cycle inhibitor p27{sup Kip1} gene. Notably, poly(ADP-ribosyl)ation activity was not required for the repressive effect of PARP-1 on FOXO1 function. Furthermore, knockdown of PARP-1 led to a decrease in cell proliferation in a manner dependentmore » on FOXO1 function. Chromatin immunoprecipitation experiments confirmed that PARP-1 is recruited to the p27{sup Kip1} gene promoter through a binding to FOXO1. These results suggest that PARP-1 acts as a corepressor for FOXO1, which could play an important role in proper cell proliferation by regulating p27{sup Kip1} gene expression.« less

FoxO6 Integrates Insulin Signaling With Gluconeogenesis in the Liver

PubMed Central

Kim, Dae Hyun; Perdomo, German; Zhang, Ting; Slusher, Sandra; Lee, Sojin; Phillips, Brett E.; Fan, Yong; Giannoukakis, Nick; Gramignoli, Roberto; Strom, Stephen; Ringquist, Steven; Dong, H. Henry

2011-01-01

OBJECTIVE Excessive endogenous glucose production contributes to fasting hyperglycemia in diabetes. This effect stems from inept insulin suppression of hepatic gluconeogenesis. To understand the underlying mechanisms, we studied the ability of forkhead box O6 (FoxO6) to mediate insulin action on hepatic gluconeogenesis and its contribution to glucose metabolism. RESEARCH DESIGN AND METHODS We characterized FoxO6 in glucose metabolism in cultured hepatocytes and in rodent models of dietary obesity, insulin resistance, or insulin-deficient diabetes. We determined the effect of FoxO6 on hepatic gluconeogenesis in genetically modified mice with FoxO6 gain- versus loss-of-function and in diabetic db/db mice with selective FoxO6 ablation in the liver. RESULTS FoxO6 integrates insulin signaling to hepatic gluconeogenesis. In mice, elevated FoxO6 activity in the liver augments gluconeogenesis, raising fasting blood glucose levels, and hepatic FoxO6 depletion suppresses gluconeogenesis, resulting in fasting hypoglycemia. FoxO6 stimulates gluconeogenesis, which is counteracted by insulin. Insulin inhibits FoxO6 activity via a distinct mechanism by inducing its phosphorylation and disabling its transcriptional activity, without altering its subcellular distribution in hepatocytes. FoxO6 becomes deregulated in the insulin-resistant liver, accounting for its unbridled activity in promoting gluconeogenesis and correlating with the pathogenesis of fasting hyperglycemia in diabetes. These metabolic abnormalities, along with fasting hyperglycemia, are reversible by selective inhibition of hepatic FoxO6 activity in diabetic mice. CONCLUSIONS Our data uncover a FoxO6-dependent pathway by which the liver orchestrates insulin regulation of gluconeogenesis, providing the proof-of-concept that selective FoxO6 inhibition is beneficial for curbing excessive hepatic glucose production and improving glycemic control in diabetes. PMID:21940782

SCP4 Promotes Gluconeogenesis Through FoxO1/3a Dephosphorylation.

PubMed

Cao, Jin; Yu, Yi; Zhang, Zhengmao; Chen, Xi; Hu, Zhaoyong; Tong, Qiang; Chang, Jiang; Feng, Xin-Hua; Lin, Xia

2018-01-01

FoxO1 and FoxO3a (collectively FoxO1/3a) proteins regulate a wide array of cellular processes, including hepatic gluconeogenesis. Phosphorylation of FoxO1/3a is a key event that determines its subcellular location and transcriptional activity. During glucose synthesis, the activity of FoxO1/3a is negatively regulated by Akt-mediated phosphorylation, which leads to the cytoplasmic retention of FoxO1/3a. However, the nuclear phosphatase that directly regulates FoxO1/3a remains to be identified. In this study, we discovered a nuclear phosphatase, SCP4/CTDSPL2 (SCP4), that dephosphorylated FoxO1/3a and promoted FoxO1/3a transcription activity. We found that SCP4 enhanced the transcription of FoxO1/3a target genes encoding PEPCK1 and G6PC, key enzymes in hepatic gluconeogenesis. Ectopic expression of SCP4 increased, while knockdown of SCP4 inhibited, glucose production. Moreover, we demonstrated that gene ablation of SCP4 led to hypoglycemia in neonatal mice. Consistent with the positive role of SCP4 in gluconeogenesis, expression of SCP4 was regulated under pathophysiological conditions. SCP4 expression was induced by glucose deprivation in vitro and in vivo and was elevated in obese mice caused by genetic (A vy ) and dietary (high-fat) changes. Thus, our findings provided experimental evidence that SCP4 regulates hepatic gluconeogenesis and could serve as a potential target for the prevention and treatment of diet-induced glucose intolerance and type 2 diabetes. © 2017 by the American Diabetes Association.

Green tea component EGCG, insulin and IGF-1 promote nuclear efflux of atrophy-associated transcription factor Foxo1 in skeletal muscle fibers.

PubMed

Wimmer, Robert J; Russell, Sarah J; Schneider, Martin F

2015-12-01

Prevention and slowing of skeletal muscle atrophy with nutritional approaches offers the potential to provide far-reaching improvements in the quality of life for our increasingly aging population. Here we show that polyphenol flavonoid epigallocatechin 3-gallate (EGCG), found in the popular beverage green tea (Camellia sinensis), demonstrates similar effects to the endogenous hormones insulin-like growth factor 1 (IGF-1) and insulin in the ability to suppress action of the atrophy-promoting transcription factor Foxo1 through a net translocation of Foxo1 out of the nucleus as monitored by nucleo-cytoplasmic movement of Foxo1-green fluorescent protein (GFP) in live skeletal muscle fibers. Foxo1-GFP nuclear efflux is rapid in IGF-1 or insulin, but delayed by an additional 30 min for EGCG. Once activated, kinetic analysis with a simple mathematical model shows EGCG, IGF-1 and insulin all produce similar apparent rate constants for Foxo1-GFP unidirectional nuclear influx and efflux. Interestingly, EGCG appears to have its effect at least partially via parallel signaling pathways that are independent of IGF-1's (and insulin's) downstream PI3K/Akt/Foxo1 signaling axis. Using the live fiber model system, we also determine the dose-response curve for both IGF-1 and insulin on Foxo1 nucleo-cytoplasmic distribution. The continued understanding of the activation mechanisms of EGCG could allow for nutritional promotion of green tea's antiatrophy skeletal muscle benefits and have implications in the development of a clinically significant parallel pathway for new drugs to target muscle wasting and the reduced insulin receptor sensitivity which causes type II diabetes mellitus. Copyright © 2015 Elsevier Inc. All rights reserved.

The metabolic activator FOXO1 binds hepatitis B virus DNA and activates its transcription

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

Shlomai, Amir, E-mail: amirsh@tasmc.health.gov.il; Institute for Gastroenterology and Liver disease, Tel-Aviv Sourasky Medical Center, 6 Weizmann street, Tel-Aviv; Shaul, Yosef

2009-04-17

Hepatitis B virus (HBV) is a small DNA virus that targets the liver and infects humans worldwide. Recently we have shown that the metabolic regulator PGC-1{alpha} coactivates HBV transcription thereby rendering the virus susceptible to fluctuations in the nutritional status of the liver. PGC-1{alpha} coactivation of HBV is mediated through the liver-enriched nuclear receptor HNF4{alpha} and through another yet unknown transcription factor(s). Here we show that the forkhead transcription factor FOXO1, a known target for PGC-1{alpha} coactivation and a central mediator of glucose metabolism in the liver, binds HBV core promoter and activates its transcription. This activation is further enhancedmore » in the presence of PGC-1{alpha}, implying that FOXO1 is a target for PGC-1{alpha} coactivation of HBV transcription. Thus, our results identify another key metabolic regulator as an activator of HBV transcription, thereby supporting the principle that HBV gene expression is regulated in a similar way to key hepatic metabolic genes.« less

FOXO/TXNIP pathway is involved in the suppression of hepatocellular carcinoma growth by glutamate antagonist MK-801

PubMed Central

2013-01-01

Background Accumulating evidence has suggested the importance of glutamate signaling in cancer growth, yet the signaling pathway has not been fully elucidated. N-methyl-D-aspartic acid (NMDA) receptor activates intracellular signaling pathways such as the extracellular-signal-regulated kinase (ERK) and forkhead box, class O (FOXO). Suppression of lung carcinoma growth by NMDA receptor antagonists via the ERK pathway has been reported. However, series of evidences suggested the importance of FOXO pathways for the regulation of normal and cancer cell growth. In the liver, FOXO1 play important roles for the cell proliferation such as hepatic stellate cells as well as liver metabolism. Our aim was to investigate the involvement of the FOXO pathway and the target genes in the growth inhibitory effects of NMDA receptor antagonist MK-801 in human hepatocellular carcinoma. Methods Expression of NMDAR1 in cancer cell lines from different tissues was examined by Western blot. NMDA receptor subunits in HepG2, HuH-7, and HLF were examined by reverse transcriptase polymerase chain reaction (RT-PCR), and growth inhibition by MK-801 and NBQX was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The effects of MK-801 on the cell cycle were examined by flow cytometry and Western blot analysis. Expression of thioredoxin-interacting protein (TXNIP) and p27 was determined by real-time PCR and Western blotting. Activation of the FOXO pathway and TXNIP induction were examined by Western blotting, fluorescence microscopy, Chromatin immunoprecipitation (ChIP) assay, and reporter gene assay. The effects of TXNIP on growth inhibition were examined using the gene silencing technique. Results NMDA receptor subunits were expressed in all cell lines examined, and MK-801, but not NBQX, inhibited cell growth of hepatocellular carcinomas. Cell cycle analysis showed that MK-801 induced G1 cell cycle arrest by down-regulating cyclin D1 and up-regulating p

Supplementation of Magnolol Attenuates Skeletal Muscle Atrophy in Bladder Cancer-Bearing Mice Undergoing Chemotherapy via Suppression of FoxO3 Activation and Induction of IGF-1.

PubMed

Chen, Meng-Chuan; Chen, Yen-Lin; Lee, Chi-Feng; Hung, Chih-Huang; Chou, Tz-Chong

2015-01-01

Skeletal muscle atrophy, the most prominent phenotypic feature of cancer cachexia, is often observed in cancer patients undergoing chemotherapy. Magnolol (M) extracted from Magnolia officinalis exhibits several pharmacological effects including anti-inflammatory and anticancer activities. In this study, we investigated whether magnolol supplementation protects against the development of cachexia symptoms in bladder cancer-bearing mice undergoing chemotherapy. Combined treatment of magnolol with chemotherapeutic drugs, such as gemcitabine and cisplatin (TGCM) or gemcitabine (TGM), markedly attenuates the body weight loss and skeletal muscle atrophy compared with conventional chemotherapy (TGC). The antiatrophic effect of magnolol may be associated with inhibition of myostatin and activin A formation, as well as FoxO3 transcriptional activity resulting from Akt activation, thereby suppressing ubiquitin ligases MuRF-1 and MAFbx/atrogin-1 expression, as well as proteasomal enzyme activity. Notably, magnolol-induced insulin-like growth factor 1 (IGF-1) production and related protein synthesis may also contribute to its protective effects. The decreased food intake, and intestinal injury and dysfunction observed in the mice of TGC group were significantly improved in the TGCM and TGM groups. Moreover, the increased inflammatory responses evidenced by elevation of proinflammatory cytokine formation and NF-κB activation occurred in the atrophying muscle of TGC group were markedly inhibited in mice of combined treatment with magnolol. In summary, these findings support that magnolol is a promising chemopreventive supplement for preventing chemotherapy-induced skeletal muscle atrophy associated with cancer cachexia by suppressing muscle protein degradation, and inflammatory responses, as well as increasing IGF-1-mediated protein synthesis.

Supplementation of Magnolol Attenuates Skeletal Muscle Atrophy in Bladder Cancer-Bearing Mice Undergoing Chemotherapy via Suppression of FoxO3 Activation and Induction of IGF-1

PubMed Central

Chen, Meng-Chuan; Chen, Yen-Lin; Lee, Chi-Feng; Hung, Chih-Huang; Chou, Tz-Chong

2015-01-01

Skeletal muscle atrophy, the most prominent phenotypic feature of cancer cachexia, is often observed in cancer patients undergoing chemotherapy. Magnolol (M) extracted from Magnolia officinalis exhibits several pharmacological effects including anti-inflammatory and anticancer activities. In this study, we investigated whether magnolol supplementation protects against the development of cachexia symptoms in bladder cancer-bearing mice undergoing chemotherapy. Combined treatment of magnolol with chemotherapeutic drugs, such as gemcitabine and cisplatin (TGCM) or gemcitabine (TGM), markedly attenuates the body weight loss and skeletal muscle atrophy compared with conventional chemotherapy (TGC). The antiatrophic effect of magnolol may be associated with inhibition of myostatin and activin A formation, as well as FoxO3 transcriptional activity resulting from Akt activation, thereby suppressing ubiquitin ligases MuRF-1 and MAFbx/atrogin-1 expression, as well as proteasomal enzyme activity. Notably, magnolol-induced insulin-like growth factor 1 (IGF-1) production and related protein synthesis may also contribute to its protective effects. The decreased food intake, and intestinal injury and dysfunction observed in the mice of TGC group were significantly improved in the TGCM and TGM groups. Moreover, the increased inflammatory responses evidenced by elevation of proinflammatory cytokine formation and NF-κB activation occurred in the atrophying muscle of TGC group were markedly inhibited in mice of combined treatment with magnolol. In summary, these findings support that magnolol is a promising chemopreventive supplement for preventing chemotherapy-induced skeletal muscle atrophy associated with cancer cachexia by suppressing muscle protein degradation, and inflammatory responses, as well as increasing IGF-1-mediated protein synthesis. PMID:26600425

Inhibition of FoxO1 acetylation by INHAT subunit SET/TAF-Iβ induces p21 transcription.

PubMed

Chae, Yun-Cheol; Kim, Kee-Beom; Kang, Joo-Young; Kim, Se-Ryeon; Jung, Hyeon-Soo; Seo, Sang-Beom

2014-08-25

Post-translational modification of forkhead family transcription factor, FoxO1, is an important regulatory mode for its diverse activities. FoxO1 is acetylated by HAT coactivators and its transcriptional activity is decreased via reduced DNA binding affinity. Here, we report that SET/TAF-Iβ inhibited p300-mediated FoxO1 acetylation in an INHAT domain-dependent manner. SET/TAF-Iβ interacted with FoxO1 and activated transcription of FoxO1 target gene, p21. Moreover, SET/TAF-Iβ inhibited acetylation of FoxO1 and increased p21 transcription induced by oxidative stress. Our results suggest that SET/TAF-Iβ inhibits FoxO1 acetylation and activates its transcriptional activity toward p21. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

FoxO1 integrates insulin signaling to VLDL production

PubMed Central

Kamagate, Adama; Dong, H. Henry

2009-01-01

Very low-density lipoproteins (VLDL) are triglyceride-rich particles. VLDL is synthesized in hepatocytes and secreted from the liver in a pathway that is tightly regulated by insulin. Hepatic VLDL production is stimulated in response to reduced insulin action, resulting in increased release of VLDL into the blood under fasting conditions. Circulating VLDL serves as a vehicle for transporting lipids to peripheral tissues for energy homeostasis. Conversely, hepatic VLDL production is suppressed in response to increased insulin release after meals. This effect is critical for preventing prolonged excursion of postprandial plasma lipid profiles in normal individuals. In subjects with obesity and type 2 diabetes, the ability of insulin to regulate VLDL production becomes impaired due to insulin resistance in the liver, resulting in excessive VLDL secretion and accumulation of triglyceride-rich particles in the blood. Such abnormality in lipid metabolism characterizes the pathogenesis of hypertriglyceridemia and accounts for increased risk of coronary artery disease in obesity and type 2 diabetes. Nevertheless, the molecular basis that links insulin resistance to VLDL overproduction remains poorly understood. Our recent studies illustrate that the forkhead transcription factor FoxO1 acts in the liver to integrate hepatic insulin action to VLDL production. Augmented FoxO1 activity in insulin resistant livers promotes hepatic VLDL overproduction and predisposes to the development of hypertriglyceridemia. These new findings raise an important question: Is FoxO1 a therapeutic target for ameliorating hypertriglyceridemia? Here we discuss this question in the context of recent advances toward our understanding of the pathophysiology of hypertriglyceridemia. PMID:18927507

Genetic association of FOXO1A and FOXO3A with longevity trait in Han Chinese populations

PubMed Central

Li, Yang; Wang, Wen-Jing; Cao, Huiqing; Lu, Jiehua; Wu, Chong; Hu, Fang-Yuan; Guo, Jian; Zhao, Ling; Yang, Fan; Zhang, Yi-Xin; Li, Wei; Zheng, Gu-Yan; Cui, Hanbin; Chen, Xiaomin; Zhu, Zhiming; He, Hongbo; Dong, Birong; Mo, Xianming; Zeng, Yi; Tian, Xiao-Li

2009-01-01

FOXO1A and FOXO3A are two members of the FoxO family. FOXO3A has recently been linked to human longevity in Japanese, German and Italian populations. Here we tested the genetic contribution of FOXO1A and FOXO3A to the longevity phenotype in Han Chinese population. Six tagging SNPs from FOXO1A and FOXO3A were selected and genotyped in 1817 centenarians and younger individuals. Two SNPs of FOXO1A were found to be associated with longevity in women (P = 0.01–0.005), whereas all three SNPs of FOXO3A were associated with longevity in both genders (P = 0.005–0.001). One SNP from FOXO1A was found not to be associated with longevity. In haplotype association tests, the OR (95% CI) for haplotypes TTG and CCG of FOXO1A in association with female longevity were 0.72 (0.58–0.90) and 1.38 (1.08–1.76), P = 0.0033 and 0.0063, respectively. The haplotypes of FOXO3A were associated with longevity in men [GTC: OR (95% CI) = 0.67 (0.51–0.86), P = 0.0014; CGT: OR (95% CI) = 1.48 (1.12–1.94), P = 0.0035] and in women [GTC: OR (95% CI) = 0.75 (0.60–0.94), P = 0.0094; CGT: OR (95% CI) = 1.47 (1.16–1.86), P = 0.0009]. The haplotype association tests were validated by permutation analysis. The association of FOXO1A with female longevity was replicated in 700 centenarians and younger individuals that were sampled geographically different from the original population. Thus, we demonstrate that, unlike FOXO3A, FOXO1A is more closely associated with human female longevity, suggesting that the genetic contribution to longevity trait may be affected by genders. PMID:19793722

C. elegans EAK-3 inhibits dauer arrest via nonautonomous regulation of nuclear DAF-16/FoxO activity

PubMed Central

Zhang, Yanmei; Xu, Jinling; Puscau, Cristina; Kim, Yongsoon; Wang, Xi; Alam, Hena; Hu, Patrick J.

2008-01-01

SUMMARY Insulin regulates development, metabolism, and lifespan via a conserved PI3K/Akt pathway that promotes cytoplasmic sequestration of FoxO transcription factors. The regulation of nuclear FoxO is poorly understood. In the nematode Caenorhabditis elegans, insulin-like signaling functions in larvae to inhibit dauer arrest and acts during adulthood to regulate lifespan. In a screen for genes that modulate C. elegans insulin-like signaling, we identified eak-3, which encodes a novel protein that is specifically expressed in the two endocrine XXX cells. The dauer arrest phenotype of eak-3 mutants is fully suppressed by mutations in daf-16/FoxO, which encodes the major target of C. elegans insulin-like signaling, and daf-12, which encodes a nuclear receptor regulated by steroid hormones known as dafachronic acids. eak-3 mutation does not affect DAF-16/FoxO subcellular localization but enhances expression of the direct DAF-16/FoxO target sod-3 in a daf-16/FoxO- and daf-12-dependent manner. eak-3 mutants have normal lifespans, suggesting that EAK-3 decouples insulin-like regulation of development and longevity. We propose that EAK-3 activity in the XXX cells promotes the synthesis and/or secretion of a hormone that acts in parallel to AKT-1 to inhibit the expression of DAF-16/FoxO target genes. Similar hormonal pathways may regulate FoxO target gene expression in mammals. PMID:18241854

Uncoupling of acetylation from phosphorylation regulates FoxO1 function independent of its subcellular localization.

PubMed

Qiang, Li; Banks, Alexander S; Accili, Domenico

2010-08-27

The activity of transcription factor FoxO1 is regulated by phosphorylation-dependent nuclear exclusion and deacetylation-dependent nuclear retention. It is unclear whether and how these two post-translational modifications affect each other. To answer this question, we expressed FoxO1 cDNAs with combined mutations of phosphorylation and acetylation sites in HEK-293 cells and analyzed their subcellular localization patterns. We show that mutations mimicking the acetylated state (KQ series) render FoxO1 more sensitive to Akt-mediated phosphorylation and nuclear exclusion and can reverse the constitutively nuclear localization of phosphorylation-defective FoxO1. Conversely, mutations mimicking the deacetylated state (KR series) promote FoxO1 nuclear retention. Oxidative stress and the Sirt1 activator resveratrol are thought to promote FoxO1 deacetylation and nuclear retention, thus increasing its activity. Accordingly, FoxO1 deacetylation was required for the effect of oxidative stress (induced by H(2)O(2)) to retain FoxO1 in the nucleus. H(2)O(2) also inhibited FoxO1 phosphorylation on Ser-253 and Thr-24, the key insulin-regulated sites, irrespective of its acetylation. In contrast, the effect of resveratrol was independent of FoxO1 acetylation and its phosphorylation on Ser-253 and Thr-24, suggesting that resveratrol acts on FoxO1 in a Sirt1- and Akt-independent manner. The dissociation of deacetylation from dephosphorylation in H(2)O(2)-treated cells indicates that the two modifications can occur independently of each other. It can be envisaged that FoxO1 exists in multiple nuclear forms with distinct activities depending on the balance of acetylation and phosphorylation.

Exendin-4 Upregulates Adiponectin Level in Adipocytes via Sirt1/Foxo-1 Signaling Pathway

PubMed Central

Wang, Anping; Li, Ting; An, Ping; Yan, Wenhua; Zheng, Hua; Wang, Baoan; Mu, Yiming

2017-01-01

Glucagon-like peptide-1 (GLP-1) receptor plays an essential role in regulating glucose metabolism. GLP-1 receptor agonists have been widely used for treating diabetes and other insulin resistance-related diseases. However, mechanisms underlying the anti-diabetic effects of GLP-1 receptor agonists remain largely unknown. In this study, we investigated the effects of GLP-1 agonist exendin-4 on the expression of adiponectin, an insulin sensitizing hormone. We found that exendin-4 increased the expression and secretion of adiponectin both in vitro and in vivo. Our data showed that exendin-4 upregulated adiponectin expression at both mRNA and protein levels in adipocytes and adipose tissues. The effects of exendin-4 on adiponectin expression were dependent on the GLP-1 receptor. We further demonstrated important roles of Sirt1 and transcriptional factor Foxo-1 in mediating the function of exendin-4 in regulating adiponectin expression. Suppression of Sirt1 or Foxo-1 expression significantly impaired exendin-4-induced adiponectin expression. Consistently, exendin-4 up-regulated Sirt1 and Foxo-1 expression in vivo. Our work is the first study demonstrating the role of Sirt1/Foxo-1 in regulating the regulatory function of a GLP-1 receptor agonist in adiponectin expression both in vitro and in vivo. The results provide important information for the mechanism underlying the function of GLP-1R on improving insulin resistance and related diseases. PMID:28122026

[Retracted] FOXO1 is crucial in glioblastoma cell tumorigenesis and regulates the expression of SIRT1 to suppress senescence in the brain.

PubMed

Yan, Han; Wu, Anhua

2018-03-01

The first author, Han Yan, wishes to retract the above article entitled 'FOXO1 is crucial in glioblastoma cell tumorigenesis and regulates the expression of SIRT1 to suppress senescence in the brain' to be published in Molecular Medicine Reports 17: 2535-2542, 2018. The article was submitted by the first author without the prior knowledge or authorization of the corresponding author, Anhua Wu, who did not participate in the design, preparation or the writing of the paper. The first author apologizes for the inconvenience caused in this regard. [The original article was published in Molecular Medicine Reports 17: 2535-2542, 2018; DOI: 10.3892/mmr.2017.8146].

Molecular Mechanism of Betaine on Hepatic Lipid Metabolism: Inhibition of Forkhead Box O1 (FoxO1) Binding to Peroxisome Proliferator-Activated Receptor Gamma (PPARγ).

PubMed

Kim, Dae Hyun; Lee, Bonggi; Kim, Min Jo; Park, Min Hi; An, Hye Jin; Lee, Eun Kyeong; Chung, Ki Wung; Park, June Whoun; Yu, Byung Pal; Choi, Jae Sue; Chung, Hae Young

2016-09-14

Betaine is a major water-soluble component of Lycium chinensis. Although there are reports about the protective effects of betaine on hepatic steatosis, the underlying mechanisms are unclear. We used db/db mice and HepG2 cells to examine the mechanism underlying betaine-mediated protection against hepatic steatosis. Here, we showed increased hepatic lipid accumulation in db/db mice, which is associated with increased activation of lipogenic transcription factors including forkhead box O1 (FoxO1) and peroxisome proliferator-activated receptor gamma (PPARγ), whereas betaine administration by oral gavage reversed these characteristics. We investigated whether betaine ameliorates hepatic steatosis by inhibiting FoxO1/PPARγ signaling in HepG2 cells. Although adenovirus-mediated FoxO1 overexpression notably increased mRNA expression levels of PPARγ and its target genes including FAS and ACC, betaine treatment reversed them. Furthermore, betaine inhibited FoxO1 binding to the PPARγ promoter and PPARγ transcriptional activity in HepG2 cells, which was previously shown to induce hepatic steatosis. We concluded that betaine ameliorates hepatic steatosis, at least in part, by inhibiting the FoxO1 binding to PPARγ and their downstream lipogenic signaling cascade.

Decoy Oligonucleotide Rescues IGF1R Expression from MicroRNA-223 Suppression

PubMed Central

Wang, Rong; He, Bao Mei; Qi, Bing; Xu, Chang Jun; Wu, Xing Zhong

2013-01-01

A mature miRNA generally suppresses hundreds of mRNA targets. To evaluate the selective effect of synthetic oligonucleotide decoys on hsa-miR-223 activity, reporters containing 3’ untranslated regions (UTR) of IGF1R, FOXO1, POLR3G, FOXO3, CDC27, FBXW7 and PAXIP1 mRNAs were constructed for the luciferase assay. The oligonucleotide decoys were designed and synthesized according to mature miR-223 sequence and its target mRNA sequence. Quantitative RT-PCR & western analysis were used to measure miR-223-targeted mRNA expression, Interestingly, apart from the antisense oligonucleotide, decoy nucleotides which were complementary to the 5’, central or 3’ region of mature miR-223 suppressed miR-223 targeting the 3’UTR of IGF1R, FOXO1, FOXO3, CDC27, POLR3G, and FBXW7 mRNAs and rescued the expression of these genes to varying degrees from miR-223 suppression at both mRNA and protein levels. All decoys had no effect on PAXIP1 which was not targeted by miR-223. The decoy 1 that was based on the sequence of IGF1R 3’UTR rescued the expression of IGF1R more significantly than other decoy nucleotides except the antisense decoy 4. Decoy 1 also rescued the expression of FOXO3 and POLR3G of which their 3’UTRs have similar binding sites for miR-223 with IGF1R 3’UTR. However decoy 1 failed to recover Sp1, CDC27 and FBXW7 expression. These data support that the sequence-specific decoy oligonucleotides might represent exogenous competing RNA which selectively inhibits microRNA targeting. PMID:24324762

Decoy oligonucleotide rescues IGF1R expression from MicroRNA-223 suppression.

PubMed

Wu, Li Hui; Cai, Qian Qian; Dong, Yi Wei; Wang, Rong; He, Bao Mei; Qi, Bing; Xu, Chang Jun; Wu, Xing Zhong

2013-01-01

A mature miRNA generally suppresses hundreds of mRNA targets. To evaluate the selective effect of synthetic oligonucleotide decoys on hsa-miR-223 activity, reporters containing 3' untranslated regions (UTR) of IGF1R, FOXO1, POLR3G, FOXO3, CDC27, FBXW7 and PAXIP1 mRNAs were constructed for the luciferase assay. The oligonucleotide decoys were designed and synthesized according to mature miR-223 sequence and its target mRNA sequence. Quantitative RT-PCR & western analysis were used to measure miR-223-targeted mRNA expression, Interestingly, apart from the antisense oligonucleotide, decoy nucleotides which were complementary to the 5', central or 3' region of mature miR-223 suppressed miR-223 targeting the 3'UTR of IGF1R, FOXO1, FOXO3, CDC27, POLR3G, and FBXW7 mRNAs and rescued the expression of these genes to varying degrees from miR-223 suppression at both mRNA and protein levels. All decoys had no effect on PAXIP1 which was not targeted by miR-223. The decoy 1 that was based on the sequence of IGF1R 3'UTR rescued the expression of IGF1R more significantly than other decoy nucleotides except the antisense decoy 4. Decoy 1 also rescued the expression of FOXO3 and POLR3G of which their 3'UTRs have similar binding sites for miR-223 with IGF1R 3'UTR. However decoy 1 failed to recover Sp1, CDC27 and FBXW7 expression. These data support that the sequence-specific decoy oligonucleotides might represent exogenous competing RNA which selectively inhibits microRNA targeting.

MicroRNA-21 regulates hepatic glucose metabolism by targeting FOXO1.

PubMed

Luo, Ailing; Yan, Haibo; Liang, Jichao; Du, Chunyuan; Zhao, Xuemei; Sun, Lijuan; Chen, Yong

2017-09-05

Abnormal activation of hepatic gluconeogenesis is a major contributor to fasting hyperglycemia in type 2 diabetes; however, the potential role of microRNAs in gluconeogenesis remains unclear. Here, we showed that hepatic expression levels of microRNA-21 (miR-21) were decreased in db/db and high-fat diet (HFD)-induced diabetic mice. Adenovirus-mediated overexpression of miR-21 decreased the expression of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) and inhibited glucose production in primary mouse hepatocytes. Silencing of miR-21 reversed this effect. Overexpression of miR-21 in the livers of db/db and HFD-induced mice was able to suppress hepatic gluconeogenesis, subsequently decreasing blood glucose levels and improving glucose and insulin intolerance. Furthermore, overexpression of miR-21 in primary mouse hepatocytes and mouse livers decreased the protein levels of FOXO1 and increased hepatic insulin sensitivity. By contrast, silencing of miR-21 increased the protein levels of FOXO1, subsequently leading to a decrease in insulin sensitivity and impaired glucose intolerance in C57BL/6 mice fed with high-fat diet for 4weeks. Finally, we confirmed that FOXO1 was a potential target of miR-21. These results suggest that miR-21 is a critical regulator in hepatic gluconeogenesis and may provide a novel therapeutic target for treating insulin resistance and type 2 diabetes. Copyright © 2017 Elsevier B.V. All rights reserved.

HTLV-1 Tax-mediated inhibition of FOXO3a activity is critical for the persistence of terminally differentiated CD4+ T cells.

PubMed

Olagnier, David; Sze, Alexandre; Bel Hadj, Samar; Chiang, Cindy; Steel, Courtney; Han, Xiaoying; Routy, Jean-Pierre; Lin, Rongtuan; Hiscott, John; van Grevenynghe, Julien

2014-12-01

The mechanisms involved in the persistence of activated CD4+ T lymphocytes following primary human T leukemia/lymphoma virus type 1 (HTLV-1) infection remain unclear. Here, we demonstrate that the HTLV-1 Tax oncoprotein modulates phosphorylation and transcriptional activity of the FOXO3a transcription factor, via upstream activation of the AKT pathway. De novo HTLV-1 infection of CD4+ T cells or direct lentiviral-mediated introduction of Tax led to AKT activation and AKT-dependent inactivation of FOXO3a, via phosphorylation of residues Ser253 and Thr32. Inhibition of FOXO3a signalling led to the long-term survival of a population of highly activated, terminally differentiated CD4+Tax+CD27negCCR7neg T cells that maintained the capacity to disseminate infectious HTLV-1. CD4+ T cell persistence was reversed by chemical inhibition of AKT activity, lentiviral-mediated expression of a dominant-negative form of FOXO3a or by specific small interfering RNA (siRNA)-mediated silencing of FOXO3a. Overall this study provides new mechanistic insight into the strategies used by HTLV-1 to increase long-term maintenance of Tax+CD4+ T lymphocytes during the early stages of HTLV-1 pathogenesis.

HTLV-1 Tax-Mediated Inhibition of FOXO3a Activity Is Critical for the Persistence of Terminally Differentiated CD4+ T Cells

PubMed Central

Bel Hadj, Samar; Chiang, Cindy; Steel, Courtney; Han, Xiaoying; Routy, Jean-Pierre; Lin, Rongtuan; Hiscott, John; van Grevenynghe, Julien

2014-01-01

The mechanisms involved in the persistence of activated CD4+ T lymphocytes following primary human T leukemia/lymphoma virus type 1 (HTLV-1) infection remain unclear. Here, we demonstrate that the HTLV-1 Tax oncoprotein modulates phosphorylation and transcriptional activity of the FOXO3a transcription factor, via upstream activation of the AKT pathway. De novo HTLV-1 infection of CD4+ T cells or direct lentiviral-mediated introduction of Tax led to AKT activation and AKT-dependent inactivation of FOXO3a, via phosphorylation of residues Ser253 and Thr32. Inhibition of FOXO3a signalling led to the long-term survival of a population of highly activated, terminally differentiated CD4+Tax+CD27negCCR7neg T cells that maintained the capacity to disseminate infectious HTLV-1. CD4+ T cell persistence was reversed by chemical inhibition of AKT activity, lentiviral-mediated expression of a dominant-negative form of FOXO3a or by specific small interfering RNA (siRNA)-mediated silencing of FOXO3a. Overall this study provides new mechanistic insight into the strategies used by HTLV-1 to increase long-term maintenance of Tax+CD4+ T lymphocytes during the early stages of HTLV-1 pathogenesis. PMID:25521510

Targeting forkhead box transcription factors FOXM1 and FOXO in leukemia (Review).

PubMed

Zhu, Hong

2014-10-01

Deregulation of forkhead box (FOX) proteins has been found in many genetic diseases and malignancies including leukemia. Leukemia is a common neoplastic disease of the blood or bone marrow characterized by the presence of immature leukocytes and is one of the leading causes of death due to cancer. Forkhead transcription factors, FOXM1 and FOXO family members (FOXOs), are important mediators in leukemia development. Aberrant expression of FOXM1 and FOXOs results in leukemogenesis. Usually the expression of FOXM1 is upregulated, whereas the expression of FOXOs is downregulated due to phosphorylation, nuclear exclusion and degradation in leukemia. On the one hand, FOXOs are bona fide tumor suppressors, on the other hand, active FOXOs maintain leukemia stem cells and stimulate drug resistance genes, contributing to leukemogenesis. FOXM1 and FOXOs have been proven to be potential targets for the development of leukemia therapeutics. They are also valuable diagnostic and prognostic markers in leukemia for clinical applications. This review summarizes the present knowledge concerning the molecular mechanisms by which FOXM1 and FOXOs modulate leukemogenesis and leukemia development, the clinical relevance of these FOX proteins in leukemia and related areas that warrant further investigation.

Reactive Oxygen Species Signaling Facilitates FOXO-3a/FBXO-Dependent Vascular BK Channel β1 Subunit Degradation in Diabetic Mice

PubMed Central

Lu, Tong; Chai, Qiang; Yu, Ling; d’Uscio, Livius V.; Katusic, Zvonimir S.; He, Tongrong; Lee, Hon-Chi

2012-01-01

Activity of the vascular large conductance Ca2+-activated K+ (BK) channel is tightly regulated by its accessory β1 subunit (BK-β1). Downregulation of BK-β1 expression in diabetic vessels is associated with upregulation of the forkhead box O subfamily transcription factor-3a (FOXO-3a)–dependent F-box–only protein (FBXO) expression. However, the upstream signaling regulating this process is unclear. Overproduction of reactive oxygen species (ROS) is a common finding in diabetic vasculopathy. We hypothesized that ROS signaling cascade facilitates the FOXO-3a/FBXO-mediated BK-β1 degradation and leads to diabetic BK channel dysfunction. Using cellular biology, patch clamp, and videomicroscopy techniques, we found that reduced BK-β1 expression in streptozotocin (STZ)-induced diabetic mouse arteries and in human coronary smooth muscle cells (SMCs) cultured with high glucose was attributable to an increase in protein kinase C (PKC)-β and NADPH oxidase expressions and accompanied by attenuation of Akt phosphorylation and augmentation of atrogin-1 expression. Treatment with ruboxistaurin (a PKCβ inhibitor) or with GW501516 (a peroxisome proliferator–activated receptor δ activator) reduced atrogin-1 expression and restored BK channel-mediated coronary vasodilation in diabetic mice. Our results suggested that oxidative stress inhibited Akt signaling and facilitated the FOXO-3a/FBXO-dependent BK-β1 degradation in diabetic vessels. Suppression of the FOXO-3a/FBXO pathway prevented vascular BK-β1 degradation and protected coronary function in diabetes. PMID:22586590

Cep55 overexpression causes male-specific sterility in mice by suppressing Foxo1 nuclear retention through sustained activation of PI3K/Akt signaling.

PubMed

Sinha, Debottam; Kalimutho, Murugan; Bowles, Josephine; Chan, Ai-Leen; Merriner, D Jo; Bain, Amanda L; Simmons, Jacinta L; Freire, Raimundo; Lopez, J Alejandro; Hobbs, Robin M; O'Bryan, Moira K; Khanna, Kum Kum

2018-04-17

( Sohlh1). Thus, Cep55 amplification leads to a shift toward the initial maintenance of undifferentiated spermatogonia and ultimately results in progressive germ cell loss. Collectively, our findings demonstrate that Cep55 overexpression causes change in germ cell proportions and manifests as a Sertoli cell only tubule phenotype, similar to that seen in many azoospermic men.-Sinha, D., Kalimutho, M., Bowles, J., Chan, A.-L., Merriner, D. J., Bain, A. L., Simmons, J. L., Freire, R., Lopez, J. A., Hobbs, R. M., O'Bryan, M. K., Khanna, K. K. Cep55 overexpression causes male-specific sterility in mice by suppressing Foxo1 nuclear retention through sustained activation of PI3K/Akt signaling.

FoxO1-negative cells are cancer stem-like cells in pancreatic ductal adenocarcinoma.

PubMed

Song, Weifeng; Li, Qi; Wang, Lei; Huang, Weiyi; Wang, Liwei

2015-06-11

Flow cytometry assays using aldehyde dehydrogenase (ALDH) activity or CD133 positivity to isolate cancer stem cells (CSCs) are widely applied but have limitations. Thus, characterization of CSC makers for a specific cancer is potentially important. We have previously shown that miR-21 regulates cancer cell growth via FoxO1 in pancreatic ductal adenocarcinoma (PDAC). Here, we areported evidence of FoxO1-negative PDAC cells as CSCs in PDAC. Both ALDH-high and CD133-high cell fractions isolated from PDAC of the patients expressed high levels of miR-21 and null FoxO1. Cultured PDAC cells were virally transduced with GFP under FoxO1 promoter. GFP (FoxO1)-null PDAC cells expressed high levels of miR-21, and grew more quickly than FoxO1-positive PDAC cells. Moreover, the fold increases in growth of FoxO1-negative vs FoxO1-positive cells were greater than CD133-high vs CD133-low cells, or ALDH-high vs ALDH-low cells. Further, FoxO1-negative cells formed tumor spheres in culture and developed tumors after serial adoptive transplantation into NOD/SCID mice, while the FoxO1-positive cells did not. Finally, selective elimination of FoxO1-negative cells completely inhibited the growth of PDAC cells. Together, these data suggest that FoxO1-negative cells as CSCs in PDAC, and targeting FoxO1-negative cells in PDAC may provide better therapeutic outcome.

Resveratrol Induces Growth Arrest and Apoptosis through Activation of FOXO Transcription Factors in Prostate Cancer Cells

PubMed Central

Chen, Qinghe; Ganapathy, Suthakar; Singh, Karan P.; Shankar, Sharmila; Srivastava, Rakesh K.

2010-01-01

Background Resveratrol, a naturally occurring phytopolyphenol compound, has attracted extensive interest in recent years because of its diverse pharmacological characteristics. Although resveratrol possesses chemopreventive properties against several cancers, the molecular mechanisms by which it inhibits cell growth and induces apoptosis have not been clearly understood. The present study was carried out to examine whether PI3K/AKT/FOXO pathway mediates the biological effects of resveratrol. Methodology/Principal Findings Resveratrol inhibited the phosphorylation of PI3K, AKT and mTOR. Resveratrol, PI3K inhibitors (LY294002 and Wortmannin) and AKT inhibitor alone slightly induced apoptosis in LNCaP cells. These inhibitors further enhanced the apoptosis-inducing potential of resveratrol. Overexpression of wild-type PTEN slightly induced apoptosis. Wild type PTEN and PTEN-G129E enhanced resveratrol-induced apoptosis, whereas PTEN-G129R had no effect on proapoptotic effects of resveratrol. Furthermore, apoptosis-inducing potential of resveratrol was enhanced by dominant negative AKT, and inhibited by wild-type AKT and constitutively active AKT. Resveratrol has no effect on the expression of FKHR, FKHRL1 and AFX genes. The inhibition of FOXO phosphorylation by resveratrol resulted in its nuclear translocation, DNA binding and transcriptional activity. The inhibition of PI3K/AKT pathway induced FOXO transcriptional activity resulting in induction of Bim, TRAIL, p27/KIP1, DR4 and DR5, and inhibition of cyclin D1. Similarly, resveratrol-induced FOXO transcriptional activity was further enhanced when activation of PI3K/AKT pathway was blocked. Over-expression of phosphorylation deficient mutants of FOXO proteins (FOXO1-TM, FOXO3A-TM and FOXO4-TM) induced FOXO transcriptional activity, which was further enhanced by resveratrol. Inhibition of FOXO transcription factors by shRNA blocked resveratrol-induced upregulation of Bim, TRAIL, DR4, DR5, p27/KIP1 and apoptosis, and

FoxO1 regulates myocardial glucose oxidation rates via transcriptional control of pyruvate dehydrogenase kinase 4 expression.

PubMed

Gopal, Keshav; Saleme, Bruno; Al Batran, Rami; Aburasayn, Hanin; Eshreif, Amina; Ho, Kim L; Ma, Wayne K; Almutairi, Malak; Eaton, Farah; Gandhi, Manoj; Park, Edwards A; Sutendra, Gopinath; Ussher, John R

2017-09-01

Pyruvate dehydrogenase (PDH) is the rate-limiting enzyme for glucose oxidation and a critical regulator of metabolic flexibility during the fasting to feeding transition. PDH is regulated via both PDH kinases (PDHK) and PDH phosphatases, which phosphorylate/inactivate and dephosphorylate/activate PDH, respectively. Our goal was to determine whether the transcription factor forkhead box O1 (FoxO1) regulates PDH activity and glucose oxidation in the heart via increasing the expression of Pdk4 , the gene encoding PDHK4. To address this question, we differentiated H9c2 myoblasts into cardiac myocytes and modulated FoxO1 activity, after which Pdk4 /PDHK4 expression and PDH phosphorylation/activity were assessed. We assessed binding of FoxO1 to the Pdk4 promoter in cardiac myocytes in conjunction with measuring the role of FoxO1 on glucose oxidation in the isolated working heart. Both pharmacological (1 µM AS1842856) and genetic (siRNA mediated) inhibition of FoxO1 decreased Pdk4 /PDHK4 expression and subsequent PDH phosphorylation in H9c2 cardiac myocytes, whereas 10 µM dexamethasone-induced Pdk4 /PDHK4 expression was abolished via pretreatment with 1 µM AS1842856. Furthermore, transfection of H9c2 cardiac myocytes with a vector expressing FoxO1 increased luciferase activity driven by a Pdk4 promoter construct containing the FoxO1 DNA-binding element region, but not in a Pdk4 promoter construct lacking this region. Finally, AS1842856 treatment in fasted mice enhanced glucose oxidation rates during aerobic isolated working heart perfusions. Taken together, FoxO1 directly regulates Pdk4 transcription in the heart, thereby controlling PDH activity and subsequent glucose oxidation rates. NEW & NOTEWORTHY Although studies have shown an association between FoxO1 activity and pyruvate dehydrogenase kinase 4 expression, our study demonstrated that pyruvate dehydrogenase kinase 4 is a direct transcriptional target of FoxO1 (but not FoxO3/FoxO4) in the heart. Furthermore, we

Cell proliferation is a key determinant of the outcome of FOXO3a activation

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

Poulsen, Raewyn C., E-mail: raewyn.poulsen@gmail.com; Carr, Andrew J.; Hulley, Philippa A.

2015-06-19

The FOXO family of forkhead transcription factors have a pivotal role in determining cell fate in response to oxidative stress. FOXO activity can either promote cell survival or induce cell death. Increased FOXO-mediated cell death has been implicated in the pathogenesis of degenerative diseases affecting musculoskeletal tissues. The aim of this study was to determine the conditions under which one member of the FOXO family, FOXO3a, promotes cell survival as opposed to cell death. Treatment of primary human tenocytes with 1 pM hydrogen peroxide for 18 h resulted in increased protein levels of FOXO3a. In peroxide-treated cells cultured in low serum media,more » FOXO3a inhibited cell proliferation and protected against apoptosis. However in peroxide treated cells cultured in high serum media, cell proliferation was unchanged but level of apoptosis significantly increased. Similarly, in tenocytes transduced to over-express FOXO3a, cell proliferation was inhibited and level of apoptosis unchanged in cells cultured in low serum. However there was a robust increase in cell death in FOXO3a-expressing cells cultured in high serum. Inhibition of cell proliferation in either peroxide-treated or FOXO3a-expressing cells cultured in high serum protected against apoptosis induction. Conversely, addition of a Chk2 inhibitor to peroxide-treated or FOXO3a-expressing cells overrode the inhibitory effect of FOXO3a on cell proliferation and led to increased apoptosis in cells cultured in low serum. This study demonstrates that proliferating cells may be particularly susceptible to the apoptosis-inducing actions of FOXO3a. Inhibition of cell proliferation by FOXO3a may be a critical event in allowing the pro-survival rather than the pro-apoptotic activity of FOXO3a to prevail. - Highlights: • FOXO3a activity can result in either promotion of cell survival or apoptosis. • The outcome of FOXO3a activation differs in proliferating compared to non-proliferating cells. �

NOTCH1 Inhibits Activation of ATM by Impairing the Formation of an ATM-FOXO3a-KAT5/Tip60 Complex.

PubMed

Adamowicz, Marek; Vermezovic, Jelena; d'Adda di Fagagna, Fabrizio

2016-08-23

The DNA damage response (DDR) signal transduction pathway is responsible for sensing DNA damage and further relaying this signal into the cell. ATM is an apical DDR kinase that orchestrates the activation and the recruitment of downstream DDR factors to induce cell-cycle arrest and repair. We have previously shown that NOTCH1 inhibits ATM activation upon DNA damage, but the underlying mechanism remains unclear. Here, we show that NOTCH1 does not impair ATM recruitment to DNA double-strand breaks (DSBs). Rather, NOTCH1 prevents binding of FOXO3a and KAT5/Tip60 to ATM through a mechanism in which NOTCH1 competes with FOXO3a for ATM binding. Lack of FOXO3a binding to ATM leads to the loss of KAT5/Tip60 association with ATM. Moreover, expression of NOTCH1 or depletion of ATM impairs the formation of the FOXO3a-KAT5/Tip60 protein complex. Finally, we show that pharmacological induction of FOXO3a nuclear localization sensitizes NOTCH1-driven cancers to DNA-damage-induced cell death. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

FOXOs modulate proteasome activity in human-induced pluripotent stem cells of Huntington's disease and their derived neural cells.

PubMed

Liu, Yanying; Qiao, Fangfang; Leiferman, Patricia C; Ross, Alan; Schlenker, Evelyn H; Wang, Hongmin

2017-11-15

Although it has been speculated that proteasome dysfunction may contribute to the pathogenesis of Huntington's disease (HD), a devastating neurodegenerative disorder, how proteasome activity is regulated in HD affected stem cells and somatic cells remains largely unclear. To better understand the pathogenesis of HD, we analyzed proteasome activity and the expression of FOXO transcription factors in three wild-type (WT) and three HD induced-pluripotent stem cell (iPSC) lines. HD iPSCs exhibited elevated proteasome activity and higher levels of FOXO1 and FOXO4 proteins. Knockdown of FOXO4 but not FOXO1 expression decreased proteasome activity. Following neural differentiation, the HD-iPSC-derived neural progenitor cells (NPCs) demonstrated lower levels of proteasome activity and FOXO expressions than their WT counterparts. More importantly, overexpression of FOXO4 but not FOXO1 in HD NPCs dramatically enhanced proteasome activity. When HD NPCs were further differentiated into DARPP32-positive neurons, these HD neurons were more susceptible to death than WT neurons and formed Htt aggregates under the condition of oxidative stress. Similar to HD NPCs, HD-iPSC-derived neurons showed reduced proteasome activity and diminished FOXO4 expression compared to WT-iPSC-derived neurons. Furthermore, HD iPSCs had lower AKT activities than WT iPSCs, whereas the neurons derived from HD iPSC had higher AKT activities than their WT counterparts. Inhibiting AKT activity increased both FOXO4 level and proteasome activity, indicating a potential role of AKT in regulating FOXO levels. These data suggest that FOXOs modulate proteasome activity, and thus represents a potentially valuable therapeutic target for HD. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

FOXO transcription factors directly activate bim gene expression and promote apoptosis in sympathetic neurons

PubMed Central

Gilley, Jonathan; Coffer, Paul J.; Ham, Jonathan

2003-01-01

Developing sympathetic neurons die by apoptosis when deprived of NGF. BIM, a BH3-only member of the BCL-2 family, is induced after NGF withdrawal in these cells and contributes to NGF withdrawal–induced death. Here, we have investigated the involvement of the Forkhead box, class O (FOXO) subfamily of Forkhead transcription factors in the regulation of BIM expression by NGF. We find that overexpression of FOXO transcription factors induces BIM expression and promotes death of sympathetic neurons in a BIM-dependent manner. In addition, we find that FKHRL1 (FOXO3a) directly activates the bim promoter via two conserved FOXO binding sites and that mutation of these sites abolishes bim promoter activation after NGF withdrawal. Finally, we show that FOXO activity contributes to the NGF deprivation–induced death of sympathetic neurons. PMID:12913110

Generation Of Functional Insulin-Producing Cells In The Gut By Foxo1 Ablation

PubMed Central

Talchai, Chutima; Xuan, Shouhong; Kitamura, Tadahiro; DePinho, Ronald A.; Accili, Domenico

2012-01-01

Restoration of regulated insulin secretion is the ultimate goal of type 1 diabetes therapy. Here we show that, surprisingly, somatic ablation of Foxo1 in Neurog3+ enteroendocrine progenitor cells gives rise to gut insulin-positive cells (Ins+) that express markers of mature β-cells, and secrete bioactive insulin as well as C-peptide in response to glucose and sulfonylureas. Lineage tracing experiments show that gut Ins+ cells arise cell-autonomously from Foxo1-deficient cells. Inducible Foxo1 ablation in adult mice also results in the generation of gut Ins+ cells. Following ablation by the β-cell toxin, streptozotocin, gut Ins+ cells regenerate and produce insulin, reversing hyperglycemia in mice. The data indicate that Neurog3+ enteroendocrine progenitors require active Foxo1 to prevent differentiation into Ins+ cells. Foxo1 ablation in gut epithelium may provide an approach to restore insulin production in type 1 diabetes. PMID:22406641

CHIP−/−-Mouse Liver: Adiponectin-AMPK-FOXO-Activation Overrides CYP2E1-Elicited JNK1-Activation, Delaying Onset of NASH: Therapeutic Implications

PubMed Central

Kim, Sung-Mi; Grenert, James P.; Patterson, Cam; Correia, Maria Almira

2016-01-01

Genetic ablation of C-terminus of Hsc70-interacting protein (CHIP) E3 ubiquitin-ligase impairs hepatic cytochrome P450 CYP2E1 degradation. Consequent CYP2E1 gain of function accelerates reactive O2 species (ROS) production, triggering oxidative/proteotoxic stress associated with sustained activation of c-Jun NH2-terminal kinase (JNK)-signaling cascades, pro-inflammatory effectors/cytokines, insulin resistance, progressive hepatocellular ballooning and microvesicular steatosis. Despite this, little evidence of nonalcoholic fatty liver disease (NAFLD)/nonalcoholic steatohepatitis (NASH) was found in CHIP−/−-mice over the first 8–9-months of life. We herein document that this lack of tissue injury is largely due to the concurrent up-regulation and/or activation of the adiponectin-5′-AMP-activated protein kinase (AMPK)-forkhead box O (FOXO)-signaling axis stemming from at the least three synergistic features: Up-regulated expression of adipose tissue adiponectin and its hepatic adipoR1/adipoR2 receptors, stabilization of hepatic AMPKα1-isoform, identified herein for the first time as a CHIP-ubiquitination substrate (unlike its AMPKα2-isoform), as well as nuclear stabilization of FOXOs, well-known CHIP-ubiquitination targets. Such beneficial predominance of the adiponectin-AMPK-FOXO-signaling axis over the sustained JNK-elevation and injurious insulin resistance in CHIP−/−-livers apparently counteracts/delays rapid progression of the hepatic microvesicular steatosis to the characteristic macrovesicular steatosis observed in clinical NASH and/or rodent NASH-models. PMID:27406999

Secreted frizzled related protein 3 (SFRP3) is required for tumorigenesis of PAX3-FOXO1-positive alveolar rhabdomyosarcoma

PubMed Central

Kephart, Julie J.G.; Tiller, Rosanne G.J.; Crose, Lisa E.S.; Slemmons, Katherine K.; Chen, Po-Han; Hinson, Ashley R.; Bentley, Rex C.; Chi, Jen-Tsan Ashley; Linardic, Corinne M.

2015-01-01

Purpose Rhabdomyosarcoma is a soft tissue sarcoma associated with the skeletal muscle lineage. Of the two predominant subtypes, known as embryonal (eRMS) and alveolar (aRMS), aRMS has the poorer prognosis, with a 5-year survival rate of <50%. The majority of aRMS tumors express the fusion protein PAX3-FOXO1. As PAX3-FOXO1 has proven chemically intractable, the current study aims to identify targetable proteins that are downstream from or cooperate with PAX3-FOXO1 to support tumorigenesis. Experimental Design Microarray analysis of the transcriptomes of human skeletal muscle myoblasts expressing PAX3-FOXO1 revealed alteration of several Wnt pathway gene members, including secreted frizzled related protein 3 (SFRP3), a secreted Wnt pathway inhibitor. Loss-of-function using shRNAs against SFRP3 were used to interrogate the role of SFRP3 in human aRMS cell lines in vitro and conditional murine xenograft systems in vivo. The combination of SFRP3 genetic suppression and the chemotherapeutic agent vincristine was also examined. Results In vitro, suppression of SFRP3 inhibited aRMS cell growth, reduced proliferation accompanied by a G1 arrest and induction of p21, and induced apoptosis. In vivo, doxycycline-inducible suppression of SFRP3 reduced aRMS tumor growth and weight by more than three-fold, in addition to increasing myogenic differentiation and β-catenin signaling. The combination of SFRP3 suppression and vincristine was more effective at reducing aRMS cell growth in vitro than either treatment alone, and ablated tumorigenesis in vivo. Conclusions SFRP3 is necessary for the growth of human aRMS cells both in vitro and in vivo and is a promising new target for investigation in aRMS. PMID:26071485

Secreted Frizzled-Related Protein 3 (SFRP3) Is Required for Tumorigenesis of PAX3-FOXO1-Positive Alveolar Rhabdomyosarcoma.

PubMed

Kephart, Julie J G; Tiller, Rosanne G J; Crose, Lisa E S; Slemmons, Katherine K; Chen, Po-Han; Hinson, Ashley R; Bentley, Rex C; Chi, Jen-Tsan Ashley; Linardic, Corinne M

2015-11-01

Rhabdomyosarcoma (RMS) is a soft tissue sarcoma associated with the skeletal muscle lineage. Of the two predominant subtypes, known as embryonal (eRMS) and alveolar (aRMS), aRMS has the poorer prognosis, with a five-year survival rate of <50%. The majority of aRMS tumors express the fusion protein PAX3-FOXO1. As PAX3-FOXO1 has proven chemically intractable, this study aims to identify targetable proteins that are downstream from or cooperate with PAX3-FOXO1 to support tumorigenesis. Microarray analysis of the transcriptomes of human skeletal muscle myoblasts expressing PAX3-FOXO1 revealed alteration of several Wnt pathway gene members, including secreted frizzled related protein 3 (SFRP3), a secreted Wnt pathway inhibitor. Loss-of-function using shRNAs against SFRP3 was used to interrogate the role of SFRP3 in human aRMS cell lines in vitro and conditional murine xenograft systems in vivo. The combination of SFRP3 genetic suppression and the chemotherapeutic agent vincristine was also examined. In vitro, suppression of SFRP3 inhibited aRMS cell growth, reduced proliferation accompanied by a G1 arrest and induction of p21, and induced apoptosis. In vivo, doxycycline-inducible suppression of SFRP3 reduced aRMS tumor growth and weight by more than three-fold, in addition to increasing myogenic differentiation and β-catenin signaling. The combination of SFRP3 suppression and vincristine was more effective at reducing aRMS cell growth in vitro than either treatment alone, and ablated tumorigenesis in vivo. SFRP3 is necessary for the growth of human aRMS cells both in vitro and in vivo and is a promising new target for investigation in aRMS. ©2015 American Association for Cancer Research.

Transcription factor FOXO1 promotes cell migration toward exogenous ATP via controlling P2Y1 receptor expression in lymphatic endothelial cells.

PubMed

Niimi, Kenta; Ueda, Mizuha; Fukumoto, Moe; Kohara, Misaki; Sawano, Toshinori; Tsuchihashi, Ryo; Shibata, Satoshi; Inagaki, Shinobu; Furuyama, Tatsuo

2017-08-05

Sprouting migration of lymphatic endothelial cell (LEC) is a pivotal step in lymphangiogenic process. However, its molecular mechanism remains unclear including effective migratory attractants. Meanwhile, forkhead transcription factor FOXO1 highly expresses in LEC nuclei, but its significance in LEC migratory activity has not been researched. In this study, we investigated function of FOXO1 transcription factor associated with LEC migration toward exogenous ATP which has recently gathered attentions as a cell migratory attractant. The transwell membrane assay indicated that LECs migrated toward exogenous ATP, which was impaired by FOXO1 knockdown. RT-PCR analysis showed that P2Y1, a purinergic receptor, expression was markedly reduced by FOXO1 knockdown in LECs. Moreover, P2Y1 blockage impaired LEC migration toward exogenous ATP. Western blot analysis revealed that Akt phosphorylation contributed to FOXO1-dependent LEC migration toward exogenous ATP and its blockage affected LEC migratory activity. Furthermore, luciferase reporter assay and ChIP assay suggested that FOXO1 directly bound to a conserved binding site in P2RY1 promoter and regulated its activity. These results indicated that FOXO1 serves a pivotal role in LEC migration toward exogenous ATP via direct transcriptional regulation of P2Y1 receptor. Copyright © 2017 Elsevier Inc. All rights reserved.

FOXO4-Knockdown Suppresses Oxidative Stress-Induced Apoptosis of Early Pro-Angiogenic Cells and Augments Their Neovascularization Capacities in Ischemic Limbs

PubMed Central

Nakayoshi, Takaharu; Sasaki, Ken-ichiro; Kajimoto, Hidemi; Koiwaya, Hiroshi; Ohtsuka, Masanori; Ueno, Takafumi; Chibana, Hidetoshi; Itaya, Naoki; Sasaki, Masahiro; Yokoyama, Shinji; Fukumoto, Yoshihiro; Imaizumi, Tsutomu

2014-01-01

The effects of therapeutic angiogenesis by intramuscular injection of early pro-angiogenic cells (EPCs) to ischemic limbs are unsatisfactory. Oxidative stress in the ischemic limbs may accelerate apoptosis of injected EPCs, leading to less neovascularization. Forkhead transcription factor 4 (FOXO4) was reported to play a pivotal role in apoptosis signaling of EPCs in response to oxidative stress. Accordingly, we assessed whether FOXO4-knockdown EPCs (FOXO4KD-EPCs) could suppress the oxidative stress-induced apoptosis and augment the neovascularization capacity in ischemic limbs. We transfected small interfering RNA targeted against FOXO4 of human EPCs to generate FOXO4KD-EPCs and confirmed a successful knockdown. FOXO4KD-EPCs gained resistance to apoptosis in response to hydrogen peroxide in vitro. Oxidative stress stained by dihydroethidium was stronger for the immunodeficient rat ischemic limb tissue than for the rat non-ischemic one. Although the number of apoptotic EPCs injected into the rat ischemic limb was greater than that of apoptotic EPCs injected into the rat non-ischemic limb, FOXO4KD-EPCs injected into the rat ischemic limb brought less apoptosis and more neovascularization than EPCs. Taken together, the use of FOXO4KD-EPCs with resistance to oxidative stress-induced apoptosis may be a new strategy to augment the effects of therapeutic angiogenesis by intramuscular injection of EPCs. PMID:24663349

Isotretinoin and FoxO1

PubMed Central

2011-01-01

Oral isotretinoin (13-cis retinoic acid) is the most effective drug in the treatment of acne and restores all major pathogenetic factors of acne vulgaris. isotretinoin is regarded as a prodrug which after isomerizisation to all-trans-retinoic acid (ATRA) induces apoptosis in cells cultured from human sebaceous glands, meibomian glands, neuroblastoma cells, hypothalamic cells, hippocampus cells, Dalton's lymphoma ascites cells, B16F-10 melanoma cells, and neuronal crest cells and others. By means of translational research this paper provides substantial indirect evidence for isotretinoin's mode of action by upregulation of forkhead box class O (FoxO) transcription factors. FoxOs play a pivotal role in the regulation of androgen receptor transactivation, insulin/insulin like growth factor-1 (IGF-1)-signaling, peroxisome proliferator-activated receptor-γ (PPArγ)- and liver X receptor-α (LXrα)-mediated lipogenesis, β-catenin signaling, cell proliferation, apoptosis, reactive oxygene homeostasis, innate and acquired immunity, stem cell homeostasis, as well as anti-cancer effects. An accumulating body of evidence suggests that the therapeutic, adverse, teratogenic and chemopreventive effecs of isotretinoin are all mediated by upregulation of FoxO-mediated gene transcription. These FoxO-driven transcriptional changes of the second response of retinoic acid receptor (RAR)-mediated signaling counterbalance gene expression of acne due to increased growth factor signaling with downregulated nuclear FoxO proteins. The proposed isotretinoin→ATRA→RAR→FoxO interaction offers intriguing new insights into the mode of isotretinoin action and explains most therapeutic, adverse and teratogenic effects of isotretinoin in the treatment of acne by a common mode of FoxO-mediated transcriptional regulation. PMID:22110774

Loss of TIMP3 underlies diabetic nephropathy via FoxO1/STAT1 interplay.

PubMed

Fiorentino, Loredana; Cavalera, Michele; Menini, Stefano; Marchetti, Valentina; Mavilio, Maria; Fabrizi, Marta; Conserva, Francesca; Casagrande, Viviana; Menghini, Rossella; Pontrelli, Paola; Arisi, Ivan; D'Onofrio, Mara; Lauro, Davide; Khokha, Rama; Accili, Domenico; Pugliese, Giuseppe; Gesualdo, Loreto; Lauro, Renato; Federici, Massimo

2013-03-01

ADAM17 and its inhibitor TIMP3 are involved in nephropathy, but their role in diabetic kidney disease (DKD) is unclear. Diabetic Timp3(-/-) mice showed increased albuminuria, increased membrane thickness and mesangial expansion. Microarray profiling uncovered a significant reduction of Foxo1 expression in diabetic Timp3(-/-) mice compared to WT, along with FoxO1 target genes involved in autophagy, while STAT1, a repressor of FoxO1 transcription, was increased. Re-expression of Timp3 in Timp3(-/-) mesangial cells rescued the expression of Foxo1 and its targets, and decreased STAT1 expression to control levels; abolishing STAT1 expression led to a rescue of FoxO1, evoking a role of STAT1 in linking Timp3 deficiency to FoxO1. Studies on kidney biopsies from patients with diabetic nephropathy confirmed a significant reduction in TIMP3, FoxO1 and FoxO1 target genes involved in autophagy compared to controls, while STAT1 expression was strongly increased. Our study suggests that loss of TIMP3 is a hallmark of DKD in human and mouse models and designates TIMP3 as a new possible therapeutic target for diabetic nephropathy. Copyright © 2013 The Authors. Published by John Wiley and Sons, Ltd on behalf of EMBO.

FoxO1 transcriptional activities in VEGF expression and beyond: a key regulator in functional angiogenesis?

PubMed

Ren, Bin

2018-04-24

FoxO1 has emerged as an important regulator of angiogenesis. Recent work published in this Journal shows that FoxO1 regulates VEGF expression in keratinocytes and is required for angiogenesis in wound healing. Since FoxO1 also regulates CD36 transcription, and endothelial cell differentiation and vascular maturation, this transcription factor may be essential for the formation of functional vascular networks via coupling the regulation of CD36 in vascular endothelial cells under physiological and pathological conditions. Although many outstanding questions remain to be answered, the mechanisms by which FoxO1 regulates VEGF in keratinocytes provide insight into the development of functional angiogenesis and further our understanding of vascular biology. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Regulation of glucose metabolism via hepatic forkhead transcription factor 1 (FoxO1) by Morinda citrifolia (noni) in high-fat diet-induced obese mice.

PubMed

Nerurkar, Pratibha V; Nishioka, Adrienne; Eck, Philip O; Johns, Lisa M; Volper, Esther; Nerurkar, Vivek R

2012-07-01

Renewed interest in alternative medicine among diabetic individuals prompted us to investigate anti-diabetic effects of Morinda citrifolia (noni) in high-fat diet (HFD)-fed mice. Type 2 diabetes is associated with increased glucose production due to the inability of insulin to suppress hepatic gluconeogenesis and promote glycolysis. Insulin inhibits gluconeogenesis by modulating transcription factors such as forkhead box O (FoxO1). Based on microarray analysis data, we tested the hypothesis that fermented noni fruit juice (fNJ) improves glucose metabolism via FoxO1 phosphorylation. C57BL/6 male mice were fed a HFD and fNJ for 12 weeks. Body weights and food intake were monitored daily. FoxO1 expression was analysed by real-time PCR and Western blotting. Specificity of fNJ-associated FoxO1 regulation of gluconeogenesis was confirmed by small interfering RNA (siRNA) studies using human hepatoma cells, HepG2. Supplementation with fNJ inhibited weight gain and improved glucose and insulin tolerance and fasting glucose in HFD-fed mice. Hypoglycaemic properties of fNJ were associated with the inhibition of hepatic FoxO1 mRNA expression, with a concomitant increase in FoxO1 phosphorylation and nuclear expulsion of the proteins. Gluconeogenic genes, phosphoenolpyruvate C kinase (PEPCK) and glucose-6-phosphatase (G6P), were significantly inhibited in mice fed a HFD+fNJ. HepG2 cells demonstrated more than 80 % inhibition of PEPCK and G6P mRNA expression in cells treated with FoxO1 siRNA and fNJ. These data suggest that fNJ improves glucose metabolism via FoxO1 regulation in HFD-fed mice.

Regulation of glucose metabolism via hepatic forkhead transcription factor 1 (FoxO1) by Morinda citrifolia (noni) in high-fat diet-induced obese mice

PubMed Central

Nerurkar, Pratibha V.; Nishioka, Adrienne; Eck, Philip O.; Nerurkar, Vivek R.

2016-01-01

Renewed interest in alternative medicine among diabetic individuals prompted us to investigate anti-diabetic effects of Morinda citrifolia (noni) in high-fat diet (HFD)-fed mice. Type 2 diabetes is associated with increased glucose production due to the inability of insulin to suppress hepatic gluconeogenesis and promote glycolysis. Insulin inhibits gluconeogenesis by modulating transcription factors such as forkhead box O (FoxO1). Based on microarray analysis data, we tested the hypothesis that fermented noni fruit juice (fNJ) improves glucose metabolism via FoxO1 phosphorylation. C57BL/6 male mice were fed a HFD and fNJ for 12 weeks. Body weights and food intake were monitored daily. FoxO1 expression was analysed by real-time PCR and Western blotting. Specificity of fNJ-associated FoxO1 regulation of gluconeogenesis was confirmed by small interfering RNA (siRNA) studies using human hepatoma cells, HepG2. Supplementation with fNJ inhibited weight gain and improved glucose and insulin tolerance and fasting glucose in HFD-fed mice. Hypoglycaemic properties of fNJ were associated with the inhibition of hepatic FoxO1 mRNA expression, with a concomitant increase in FoxO1 phosphorylation and nuclear expulsion of the proteins. Gluconeogenic genes, phosphoenolpyruvate C kinase (PEPCK) and glucose-6-phosphatase (G6P), were significantly inhibited in mice fed a HFD + fNJ. HepG2 cells demonstrated more than 80% inhibition of PEPCK and G6P mRNA expression in cells treated with FoxO1 siRNA and fNJ. These data suggest that fNJ improves glucose metabolism via FoxO1 regulation in HFD-fed mice. PMID:22011624

MicroRNA-9 promotes the proliferation, migration, and invasion of breast cancer cells via down-regulating FOXO1.

PubMed

Liu, D-Z; Chang, B; Li, X-D; Zhang, Q-H; Zou, Y-H

2017-09-01

The objective of the study was to investigate the role of microRNA-9 (miR-9) targeting forkhead box O1 (FOXO1) in the proliferation, migration, and invasion of breast cancer cells. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to determine the expressions of miR-9 and FOXO1 mRNA in breast cancer tissues, normal breast tissues, breast cancer cell lines, and normal breast epithelial cells. After the up-regulation of miR-9 expression, qRT-PCR and Western blotting were used to determine the expression of FOXO1. The luciferase reporter gene assay was used to validate the target gene. The CCK-8 assay, scratch-wound healing assay, and Transwell invasion assay were used to investigate the changes in the proliferation, migration, and invasion of breast cancer cells, respectively. MicroRNA-9 expression was significantly up-regulated in breast cancer tissues and breast cancer cell lines when compared with normal breast tissues and normal breast epithelial cells (both P < 0.05). FOXO1 mRNA and protein expressions were substantially down-regulated in breast cancer tissues and breast cancer cell lines when compared with normal breast tissues and normal breast epithelial cells (both P < 0.05). There can be a negative correlation between miR-9 and FOXO1 mRNA in breast cancer. Luciferase reporter gene assay indicated that miR-9 can down-regulate FOXO1 expression at a post-transcriptional level through binding specifically to FOXO1 3'UTR. The results of CCK-8 assay, scratch-wound healing assay, and Transwell invasion assay revealed that the inhibition of miR-9 can suppress MCF7 cell proliferation, migration, and invasion. Additionally, the expression of miR-9 increased significantly whilst that of FOXO1 decreased substantially as the disease progressed (P < 0.05). Our study provides evidence that miR-9 can promote the proliferation, migration, and invasion of breast cancer cells via down-regulating FOXO1.

Transcription Factor FoxO1 Is Essential for Enamel Biomineralization

PubMed Central

Poché, Ross A.; Sharma, Ramaswamy; Garcia, Monica D.; Wada, Aya M.; Nolte, Mark J.; Udan, Ryan S.; Paik, Ji-Hye; DePinho, Ronald A.; Bartlett, John D.; Dickinson, Mary E.

2012-01-01

The Transforming growth factor β (Tgf-β) pathway, by signaling via the activation of Smad transcription factors, induces the expression of many diverse downstream target genes thereby regulating a vast array of cellular events essential for proper development and homeostasis. In order for a specific cell type to properly interpret the Tgf-β signal and elicit a specific cellular response, cell-specific transcriptional co-factors often cooperate with the Smads to activate a discrete set of genes in the appropriate temporal and spatial manner. Here, via a conditional knockout approach, we show that mice mutant for Forkhead Box O transcription factor FoxO1 exhibit an enamel hypomaturation defect which phenocopies that of the Smad3 mutant mice. Furthermore, we determined that both the FoxO1 and Smad3 mutant teeth exhibit changes in the expression of similar cohort of genes encoding enamel matrix proteins required for proper enamel development. These data raise the possibility that FoxO1 and Smad3 act in concert to regulate a common repertoire of genes necessary for complete enamel maturation. This study is the first to define an essential role for the FoxO family of transcription factors in tooth development and provides a new molecular entry point which will allow researchers to delineate novel genetic pathways regulating the process of biomineralization which may also have significance for studies of human tooth diseases such as amelogenesis imperfecta. PMID:22291941

Elovl5 regulates the mTORC2-Akt-FOXO1 pathway by controlling hepatic cis-vaccenic acid synthesis in diet-induced obese mice[S

PubMed Central

Tripathy, Sasmita; Jump, Donald B.

2013-01-01

Elevated hepatic expression of fatty acid elongase-5 (Elovl5) induces FoxO1 phosphorylation, lowers FoxO1 nuclear content, and suppresses expression of genes involved in gluconeogenesis (GNG). In this report, we define the molecular and metabolic basis of Elovl5 control of FoxO1 phosphorylation. Adenoviral-mediated (Ad-Elovl5) induction of hepatic Elovl5 in diet-induced obese, glucose-intolerant mice and HepG2 cells increased the phosphorylation of Akt2-S473 [mammalian target of rapamycin complex-2 (mTORC2) site], but not Akt2-T308 (PDK1 site). The Akt2 inhibitor Akti1/2 blocked Elovl5 induction of FoxO1-S256 phosphorylation in HepG2 cells. Elevated Elovl5 activity in liver and HepG2 cells induced rictor mRNA, rictor protein, and rictor-mTOR interaction, whereas rictor knockdown (siRNA) attenuated Elovl5 induction of Akt2-S473 and FoxO1-S256 phosphorylation in HepG2 cells. FA analysis revealed that the abundance of cis-vaccenic acid (18:1,n-7) was increased in livers of obese mice and HepG2 cells following Ad-Elovl5 infection. Treating HepG2 cells with Elovl5 substrates established that palmitoleic acid (16:1,n-7), but not γ-linolenic acid (18:3,n-6), induced rictor protein, Akt-S473, and FoxO1-S256 phosphorylation. Inhibition of FA elongation blocked 16:1,n-7 but not 18:1,n-7 induction of rictor protein and Akt-S473 and FoxO1-S256 phosphorylation. These results establish a novel link between Elovl5-mediated synthesis of 18:1,n-7 and GNG through the control of the mTORC2-Akt-FoxO1 pathway. PMID:23099444

Protection against dexamethasone-induced muscle atrophy is related to modulation by testosterone of FOXO1 and PGC-1{alpha}

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

Qin, Weiping, E-mail: weiping.qin@mssm.edu; Department of Medicine, Mount Sinai School of Medicine, NY; Pan, Jiangping

Research highlights: {yields} In rat gastrocnemius muscle, dexamethasone reduced PGC-1{alpha} cellular and nuclear levels without altering mRNA levels for this factor. {yields} Dexamethasone reduced phosphorylating of p38 MAPK, which stabilizes PGC-1{alpha} and promotes its nuclear entry. {yields} Co-administration of testosterone with dexamethasone increased cellular and nuclear levels of PGC-1{alpha} protein without changing its mRNA levels. {yields} Co-administration of testosterone restored p38 MAPK levels to those of controls. -- Abstract: Glucocorticoid-induced muscle atrophy results from muscle protein catabolism and reduced protein synthesis, associated with increased expression of two muscle-specific ubiquitin ligases (MAFbx and MuRF1), and of two inhibitors of protein synthesis,more » REDD1 and 4EBP1. MAFbx, MuRF1, REDD1 and 4EBP1 are up-regulated by the transcription factors FOXO1 and FOXO3A. The transcriptional co-activator PGC-1{alpha} has been shown to attenuate many forms of muscle atrophy and to repress FOXO3A-mediated transcription of atrophy-specific genes. Dexamethasone-induced muscle atrophy can be prevented by testosterone, which blocks up-regulation by dexamethasone of FOXO1. Here, an animal model of dexamethasone-induced muscle atrophy was used to further characterize effects of testosterone to abrogate adverse actions of dexamethasone on FOXO1 levels and nuclear localization, and to determine how these agents affect PGC-1{alpha}, and its upstream activators, p38 MAPK and AMPK. In rat gastrocnemius muscle, testosterone blunted the dexamethasone-mediated increase in levels of FOXO1 mRNA, and FOXO1 total and nuclear protein. Dexamethasone reduced total and nuclear PGC-1{alpha} protein levels in the gastrocnemius; co-administration of testosterone with dexamethasone increased total and nuclear PGC-1{alpha} levels above those present in untreated controls. Testosterone blocked dexamethasone-induced decreases in activity of p38 MAPK in the

Mitochondria and FOXO3: breath or die.

PubMed

Hagenbuchner, Judith; Ausserlechner, Michael J

2013-01-01

Forkhead box O (FOXO) transcription factors are regulators of cell-type specific apoptosis and cell cycle arrest but also control longevity and reactive oxygen species (ROS). ROS-control by FOXO is mediated by transcriptional activation of detoxifying enzymes such as Superoxide dismutase 2 (SOD2), Catalase or Sestrins or by the repression of mitochondrial respiratory chain proteins resulting in reduced mitochondrial activity. FOXO3 also regulates the adaptation to hypoxia by reducing mitochondrial mass and oxygen consumption during HIF-1α activation. In neuronal tumor cells, FOXO3 triggers ROS-accumulation as a consequence of transient mitochondrial outer membrane permeabilization, which is essential for FOXO3-induced apoptosis in these cells. Cellular ROS levels are affected by the FOXO-targets Bim, BclxL, and Survivin. All three proteins localize to mitochondria and affect mitochondrial membrane potential, respiration and cellular ROS levels. Bim-activation by FOXO3 causes mitochondrial depolarization resulting in a transitory decrease of respiration and ROS production. Survivin, on the other hand, actively changes mitochondrial architecture, respiration-efficacy and energy metabolism. This ability distinguishes Survivin from other anti-apoptotic proteins such as BclxL, which inhibits ROS by inactivating Bim but does not alter mitochondrial function. Importantly, FOXO3 simultaneously also activates ROS-detoxification via induction of SESN3. In this paper we discuss the hypothesis that the delicate balance between ROS-accumulation by Bim-triggered mitochondrial damage, mitochondrial architecture and ROS-detoxifying proteins determines cell fate. We provide evidence for a FOXO self-reactivating loop and for novel functions of FOXO3 in controlling mitochondrial respiration of neuronal cells, which further supports the current view that FOXO transcription factors are information-integrating sentinels of cellular stress and critical modulators of cell homeostasis.

Inhibition of FoxO transcriptional activity prevents muscle fiber atrophy during cachexia and induces hypertrophy

PubMed Central

Reed, Sarah A.; Sandesara, Pooja B.; Senf, Sarah M.; Judge, Andrew R.

2012-01-01

Cachexia is characterized by inexorable muscle wasting that significantly affects patient prognosis and increases mortality. Therefore, understanding the molecular basis of this muscle wasting is of significant importance. Recent work showed that components of the forkhead box O (FoxO) pathway are increased in skeletal muscle during cachexia. In the current study, we tested the physiological significance of FoxO activation in the progression of muscle atrophy associated with cachexia. FoxO-DNA binding dependent transcription was blocked in the muscles of mice through injection of a dominant negative (DN) FoxO expression plasmid prior to inoculation with Lewis lung carcinoma cells or the induction of sepsis. Expression of DN FoxO inhibited the increased mRNA levels of atrogin-1, MuRF1, cathepsin L, and/or Bnip3 and inhibited muscle fiber atrophy during cancer cachexia and sepsis. Interestingly, during control conditions, expression of DN FoxO decreased myostatin expression, increased MyoD expression and satellite cell proliferation, and induced fiber hypertrophy, which required de novo protein synthesis. Collectively, these data show that FoxO-DNA binding-dependent transcription is necessary for normal muscle fiber atrophy during cancer cachexia and sepsis, and further suggest that basal levels of FoxO play an important role during normal conditions to depress satellite cell activation and limit muscle growth.—Reed, S. A., Sandesara, P. B., Senf, S. F., Judge, A. R. Inhibition of FoxO transcriptional activity prevents muscle fiber atrophy during cachexia and induces hypertrophy. PMID:22102632

CD11b activation suppresses TLR-dependent inflammation and autoimmunity in systemic lupus erythematosus

PubMed Central

Faridi, Mohd Hafeez; Khan, Samia Q.; Zhao, Wenpu; Lee, Ha Won; Altintas, Mehmet M.; Zhang, Kun; Kumar, Vinay; Armstrong, Andrew R.; Carmona-Rivera, Carmelo; Dorschner, Jessica M.; Schnaith, Abigail M.; Li, Xiaobo; Ghodke-Puranik, Yogita; Moore, Erica; Irizarry-Caro, Jorge; Zhang, Tingting; Day, Rachael; Stoub, Darren; Hoffmann, Victoria; Khaliqdina, Shehryar Jehangir; Bhargava, Prachal; Santander, Ana M.; Torroella-Kouri, Marta; Issac, Biju; Cimbaluk, David J.; Zloza, Andrew; Prabhakar, Rajeev; Deep, Shashank; Jolly, Meenakshi; Koh, Kwi Hye; Reichner, Jonathan S.; Bradshaw, Elizabeth M.; Chen, JianFeng; Moita, Luis F.; Yuen, Peter S.; Li Tsai, Wanxia; Singh, Bhupinder; Reiser, Jochen; Nath, Swapan K.; Niewold, Timothy B.; Vazquez-Padron, Roberto I.

2017-01-01

Genetic variations in the ITGAM gene (encoding CD11b) strongly associate with risk for systemic lupus erythematosus (SLE). Here we have shown that 3 nonsynonymous ITGAM variants that produce defective CD11b associate with elevated levels of type I interferon (IFN-I) in lupus, suggesting a direct link between reduced CD11b activity and the chronically increased inflammatory status in patients. Treatment with the small-molecule CD11b agonist LA1 led to partial integrin activation, reduced IFN-I responses in WT but not CD11b-deficient mice, and protected lupus-prone MRL/Lpr mice from end-organ injury. CD11b activation reduced TLR-dependent proinflammatory signaling in leukocytes and suppressed IFN-I signaling via an AKT/FOXO3/IFN regulatory factor 3/7 pathway. TLR-stimulated macrophages from CD11B SNP carriers showed increased basal expression of IFN regulatory factor 7 (IRF7) and IFN-β, as well as increased nuclear exclusion of FOXO3, which was suppressed by LA1-dependent activation of CD11b. This suggests that pharmacologic activation of CD11b could be a potential mechanism for developing SLE therapeutics. PMID:28263189

Posttranslational modulation of FoxO1 contributes to cardiac remodeling in post-ischemic heart failure.

PubMed

Kappel, Ben Arpad; Stöhr, Robert; De Angelis, Lorenzo; Mavilio, Maria; Menghini, Rossella; Federici, Massimo

2016-06-01

Forkhead box protein O1 (FoxO1) plays a key role in energy homeostasis, stress response and autophagy and is dysregulated in diabetes and ischemia. We investigated cardiac FoxO1 expression and posttranstranslational modifications after myocardial infarction (MI) and further tested if active posttranstranslational modulation of FoxO1 can alter cardiac remodeling in postischemic heart failure. Non-diabetic and diabetic C57BL/6 mice were subjected to MI by ligation of left anterior descending artery. In selected experiments we combined this model with intramyocardial injection of adenovirus expressing different isoforms of FoxO1. We used Millar catheter, histology, Western blot and metabolomics for further analyses. We show that after MI total cardiac FoxO1 is downregulated and partly recovers after 7 days. This downregulation is accompanied by fundamental posttranslational modifications of FoxO1, particularly acetylation. Adenovirus experiments revealed smaller infarction size and improved heart function in mice expressing a constitutively deacetylated variant of FoxO1 compared to a wild type variant of FoxO1 in both non-diabetic (MI size: -13.4 ± 3.5%; LVDP: +29.1 ± 9.4  mmHg; p < 0.05) and diabetic mice (MI size: -17.6 ± 3.7%; LVDP: +10.9 ± 3.6  mmHg; p < 0.05). Metabolomics analyses showed alterations in metabolites connected to muscle breakdown, collagen/elastin and energy metabolism between the two groups. First, our results demonstrate that myocardial ischemia is associated with downregulation and posttranslational modification of cardiac FoxO1. Second, we show in a mouse model of postischemic heart failure that posttranslational modulation of FoxO1 alters heart function involving collagen and protein metabolism. Therefore, posttranslational modifications of FoxO1 could be an option to target remodeling processes in postischemic heart failure. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

The oncoprotein HBXIP suppresses gluconeogenesis through modulating PCK1 to enhance the growth of hepatoma cells.

PubMed

Shi, Hui; Fang, Runping; Li, Yinghui; Li, Leilei; Zhang, Weiying; Wang, Huawei; Chen, Fuquan; Zhang, Shuqin; Zhang, Xiaodong; Ye, Lihong

2016-11-28

Hepatitis B X-interacting protein (HBXIP) as an oncoprotein plays crucial roles in the development of cancer, involving glucose metabolism reprogramming. In this study, we are interested in whether the oncoprotein HBXIP is involved in the modulation of gluconeogenesis in liver cancer. Here, we showed that the expression level of phosphoenolpyruvate carboxykinase (PCK1), a key enzyme of gluconeogenesis, was lower in clinical hepatocellular carcinoma (HCC) tissues than that in normal tissues. Mechanistically, HBXIP inhibited the expression of PCK1 through down-regulating transcription factor FOXO1 in hepatoma cells, and up-regulated miR-135a targeting the 3'UTR of FOXO1 mRNA in the cells. In addition, HBXIP increased the phosphorylation levels of FOXO1 protein by activating PI3K/Akt pathway, leading to the export of FOXO1 from nucleus to cytoplasm. Strikingly, over-expression of PCK1 could abolish the HBXIP-promoted growth of hepatoma cells in vitro and in vivo. Thus, we conclude that the oncoprotein HBXIP is able to depress the gluconeogenesis through suppressing PCK1 to promote hepatocarcinogenesis, involving miR-135a/FOXO1 axis and PI3K/Akt/p-FOXO1 pathway. Our finding provides new insights into the mechanism by which oncoprotein HBXIP modulates glucose metabolism reprogramming in HCC. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Astaxanthin attenuates contrast agent-induced acute kidney injury in vitro and in vivo via the regulation of SIRT1/FOXO3a expression.

PubMed

Liu, Nana; Chen, Jing; Gao, Dongmei; Li, Wenhua; Zheng, Di

2018-06-01

The study was processed to investigate the effect of astaxanthin (AST; 3,3-dihydroxybeta, beta-carotene-4,4-dione) on the acute kidney injury induced by iohexol and the relationship with SIRT1/FOXO3a signal pathway. Thirty male Sprague Dawley rats were randomly divided into five groups as follows: control group (CON; olive oil only), contrast media group, astaxanthin control group (100 mg/kg), low astaxanthin dose group (LAG, 50 mg/kg) and high astaxanthin dose group (HAG, 100 mg/kg). As followed, serum creatinine (SCr), blood urea nitrogen (BUN), the oxidative stress markers and apoptosis-related proteins were detected. Human proximal tubular epithelial cells (HK-2) were cultured in DMEM/F12 medium in vitro and then randomly divided into appropriate experimental groups: normal group (N), dimethyl sulfoxide (DMSO), iohexol group (I), iohexol + (1.0, 10.0 μmol/L) astaxanthin group (I + LAST; I + HAST), iohexol + SIRT1 inhibitors (nicotinamide) group (NA) and iohexol + si-RNA FOXO3a group (si-RNA FOXO3a); when cultured for 24 h, cell proliferation ability was tested by cell counting kit (CCK-8), reactive oxygen species (ROS) were detected by flow cytometry and the expression of SIRT1 and FOXO3a were observed using western blot. At the end of the experiment, the levels of SCr, BUN and malondialdehyde (MDA) were all increased in the CM group. The LAG and HAG reduce superoxide anion (SOD) activity, catalase (CAT) activity, glutathione peroxidase (GPx) activity and glutathione (GSH) content, as well as SCr and BUN level. Moreover, apoptosis-associated proteins, caspase 3 p17, bax and bcl-2 were upregulated. In HK-2 cells, after adding iohexol, proliferation and intracellular ROS levels were significantly increased. Using astaxanthin in advance after the intervention, the result is opposite. SIRTl inhibitors NA can reduce the expression of SIRTl and decrease the expression of FOXO3a protein. Si-RNA FOXO3a reduces the expression of FOXO3a but had no

Deacetylation of FOXO3 by SIRT1 or SIRT2 leads to Skp2-mediated FOXO3 ubiquitination and degradation

USDA-ARS?s Scientific Manuscript database

Sirtuin deacetylases and FOXO (Forkhead box, class O) transcription factors have important roles in many biological pathways, including cancer development. SIRT1 and SIRT2 deacetylate FOXO factors to regulate FOXO function. Because acetylation and ubiquitination both modify the '-amino group of lysi...

FOXO1, TGF-β Regulation and Wound Healing

PubMed Central

Hameedaldeen, Alhassan; Liu, Jian; Batres, Angelika; Graves, Gabrielle S.; Graves, Dana T.

2014-01-01

Re-epithelialization is a complex process that involves migration and proliferation of keratinocytes, in addition to the production of cytokines and growth factors that affect other cells. The induction of transcription factors during these processes is crucial for successful wound healing. The transcription factor forkhead boxO-1 (FOXO1) has recently been found to be an important regulator of wound healing. In particular, FOXO1 has significant effects through regulation of transforming growth factor-beta (TGF-β) expression and protecting keratinocytes from oxidative stress. In the absence of FOXO1, there is increased oxidative damage, reduced TGF-β1 expression, reduced migration and proliferation of keratinocytes and increased keratinocytes apoptosis leading to impaired re-epithelialization of wounds. PMID:25226535

Houttuynia cordata Thunb Promotes Activation of HIF-1A-FOXO3 and MEF2A Pathways to Induce Apoptosis in Human HepG2 Hepatocellular Carcinoma Cells.

PubMed

Kim, Jung Min; Hwang, In-Hu; Jang, Ik-Soon; Kim, Min; Bang, In Seok; Park, Soo Jung; Chung, Yun-Jo; Joo, Jong-Cheon; Lee, Min-Goo

2017-09-01

Houttuynia cordata Thunb ( H cordata), a medicinal plant, has anticancer activity, as it inhibits cell growth and induces cell apoptosis in cancer. However, the potential anti-cancer activity and mechanism of H cordata for human liver cancer cells is not well understood. Recently, we identified hypoxia-inducible factor (HIF)-1A, Forkhead box (FOX)O3, and MEF2A as proapoptotic factors induced by H cordata, suggesting that HIF-1A, FOXO3, and MEF2A contribute to the apoptosis of HepG2 hepatocellular carcinoma cells. FOXO3 transcription factors regulate target genes involved in apoptosis. H cordata significantly increased the mRNA and protein expression of HIF-1A and FOXO3 and stimulated MEF2A expression in addition to increased apoptosis in HepG2 cells within 24 hours. Therefore, we determined the potential role of FOXO3 on apoptosis and on H cordata-induced MEF2A in HepG2 cells. HIF-1A silencing by siRNA attenuated MEF2A and H cordata-mediated FOXO3 upregulation in HepG2 cells. Furthermore, H cordata-mediated MEF2A expression enhanced caspase-3 and caspase-7, which were abolished on silencing FOXO3 with siRNA. In addition, H cordata inhibited growth of human hepatocellular carcinoma xenografts in nude mice. Taken together, our results demonstrate that H cordata enhances HIF-1A/FOXO3 signaling, leading to MEF2A upregulation in HepG2 cells, and in parallel, it disturbs the expression of Bcl-2 family proteins (Bax, Bcl-2, and Bcl-xL), which results in apoptosis. Taken together, these findings demonstrate that H cordata promotes the activation of HIF-1A-FOXO3 and MEF2A pathways to induce apoptosis in human HepG2 hepatocellular carcinoma cells and is, therefore, a promising candidate for antitumor drug development.

Houttuynia cordata Thunb Promotes Activation of HIF-1A–FOXO3 and MEF2A Pathways to Induce Apoptosis in Human HepG2 Hepatocellular Carcinoma Cells

PubMed Central

Kim, Jung Min; Hwang, In-Hu; Jang, Ik-Soon; Kim, Min; Bang, In Seok; Park, Soo Jung; Chung, Yun-Jo; Joo, Jong-Cheon; Lee, Min-Goo

2016-01-01

Houttuynia cordata Thunb (H cordata), a medicinal plant, has anticancer activity, as it inhibits cell growth and induces cell apoptosis in cancer. However, the potential anti-cancer activity and mechanism of H cordata for human liver cancer cells is not well understood. Recently, we identified hypoxia-inducible factor (HIF)-1A, Forkhead box (FOX)O3, and MEF2A as proapoptotic factors induced by H cordata, suggesting that HIF-1A, FOXO3, and MEF2A contribute to the apoptosis of HepG2 hepatocellular carcinoma cells. FOXO3 transcription factors regulate target genes involved in apoptosis. H cordata significantly increased the mRNA and protein expression of HIF-1A and FOXO3 and stimulated MEF2A expression in addition to increased apoptosis in HepG2 cells within 24 hours. Therefore, we determined the potential role of FOXO3 on apoptosis and on H cordata–induced MEF2A in HepG2 cells. HIF-1A silencing by siRNA attenuated MEF2A and H cordata–mediated FOXO3 upregulation in HepG2 cells. Furthermore, H cordata–mediated MEF2A expression enhanced caspase-3 and caspase-7, which were abolished on silencing FOXO3 with siRNA. In addition, H cordata inhibited growth of human hepatocellular carcinoma xenografts in nude mice. Taken together, our results demonstrate that H cordata enhances HIF-1A/FOXO3 signaling, leading to MEF2A upregulation in HepG2 cells, and in parallel, it disturbs the expression of Bcl-2 family proteins (Bax, Bcl-2, and Bcl-xL), which results in apoptosis. Taken together, these findings demonstrate that H cordata promotes the activation of HIF-1A–FOXO3 and MEF2A pathways to induce apoptosis in human HepG2 hepatocellular carcinoma cells and is, therefore, a promising candidate for antitumor drug development. PMID:27698266

FOXO1 expression in keratinocytes promotes connective tissue healing

PubMed Central

Zhang, Chenying; Lim, Jason; Liu, Jian; Ponugoti, Bhaskar; Alsadun, Sarah; Tian, Chen; Vafa, Rameen; Graves, Dana T.

2017-01-01

Wound healing is complex and highly orchestrated. It is well appreciated that leukocytes, particularly macrophages, are essential for inducing the formation of new connective tissue, which requires the generation of signals that stimulate mesenchymal stem cells (MSC), myofibroblasts and fibroblasts. A key role for keratinocytes in this complex process has yet to be established. To this end, we investigated possible involvement of keratinocytes in connective tissue healing. By lineage-specific deletion of the forkhead box-O 1 (FOXO1) transcription factor, we demonstrate for the first time that keratinocytes regulate proliferation of fibroblasts and MSCs, formation of myofibroblasts and production of collagen matrix in wound healing. This stimulation is mediated by a FOXO1 induced TGFβ1/CTGF axis. The results provide direct evidence that epithelial cells play a key role in stimulating connective tissue healing through a FOXO1-dependent mechanism. Thus, FOXO1 and keratinocytes may be an important therapeutic target where healing is deficient or compromised by a fibrotic outcome. PMID:28220813

Cross-talk among HMGA1 and FoxO1 in control of nuclear insulin signaling.

PubMed

Chiefari, Eusebio; Arcidiacono, Biagio; Palmieri, Camillo; Corigliano, Domenica Maria; Morittu, Valeria Maria; Britti, Domenico; Armoni, Michal; Foti, Daniela Patrizia; Brunetti, Antonio

2018-06-04

As a mediator of insulin-regulated gene expression, the FoxO1 transcription factor represents a master regulator of liver glucose metabolism. We previously reported that the high-mobility group AT-hook 1 (HMGA1) protein, a molecular switch for the insulin receptor gene, functions also as a downstream target of the insulin receptor signaling pathway, representing a critical nuclear mediator of insulin function. Here, we investigated whether a functional relationship existed between FoxO1 and HMGA1, which might help explain insulin-mediated gene transcription in the liver. To this end, as a model study, we investigated the canonical FoxO1-HMGA1-responsive IGFBP1 gene, whose hepatic expression is regulated by insulin. By using a conventional GST-pull down assay combined with co-immunoprecipitation and Fluorescence Resonance Energy Transfer (FRET) analyses, we provide evidence of a physical interaction between FoxO1 and HMGA1. Further investigation with chromatin immunoprecipitation, confocal microscopy, and Fluorescence Recovery After Photobleaching (FRAP) technology indicated a functional significance of this interaction, in both basal and insulin-stimulated states, providing evidence that, by modulating FoxO1 transactivation, HMGA1 is essential for FoxO1-induced IGFBP1 gene expression, and thereby a critical modulator of insulin-mediated FoxO1 regulation in the liver. Collectively, our findings highlight a novel FoxO1/HMGA1-mediated mechanism by which insulin may regulate gene expression and metabolism.

Oxidative stress reduces trophoblast FOXO1 and integrin β3 expression that inhibits cell motility.

PubMed

Chen, Chie-Pein; Chen, Cheng-Yi; Wu, Yi-Hsin; Chen, Chia-Yu

2018-06-08

Preeclampsia is a serious pregnancy complication associated with placental oxidative stress and impaired trophoblast migration. The mechanism of defective trophoblast migration remains unknown. Forkhead box O1 (FOXO1) is a transcription factor. Integrin β3 is involved in cell motility. We hypothesized that FOXO1 mediates expression of trophoblast integrin β3, which could be impaired by oxidative stress and have implications in preeclampsia. The expressions of FOXO1 and integrin β3 were significantly reduced in preeclamptic placentas (n = 15) compared to that of controls (n = 15; p < 0.01). HTR-8/SVneo and JEG-3 trophoblasts were transfected to express wild-type FOXO1-WT or constitutively-expressed nuclear mutant form, FOXO1-AAA. The FOXO1 in HTR-8/SVneo and 3A-Sub-E trophoblasts was silenced by small interfering RNA. AKT-mediated phosphorylation inactivated FOXO1, but FOXO1-AAA was not phosphorylated. The expression of trophoblast integrin β3 was significantly elevated by FOXO1 overexpression and inhibited by FOXO1 knockdown. FOXO1 regulates integrin β3 at the transcriptional level via binding to the putative FOXO1 response element site between position -1154 to -1139 (TGAGATGTTTTGAAAG) in HTR-8/SVneo trophoblasts. The level of phosphorylated FOXO1 was decreased, and the FOXO1 level was increased in trophoblasts treated with AKT inhibitor MK2206, leading to upregulation of integrin β3. The capabilities of trophoblast adhesion and migration were enhanced by FOXO1-overexpression or MK2206, and inhibited by silencing FOXO1 or oxidative stress with H 2 O 2 . These results suggest that FOXO1 enhances trophoblast integrin β3 expression, and mediates cell adhesion and migration. By affecting the expression of FOXO1 and cell motility in trophoblasts, oxidative stress plays a role in the development of preeclampsia. Copyright © 2018 Elsevier Inc. All rights reserved.

Activation of PPAR alpha by fenofibrate inhibits apoptosis in vascular adventitial fibroblasts partly through SIRT1-mediated deacetylation of FoxO1.

PubMed

Wang, Wei-Rong; Liu, En-Qi; Zhang, Ji-Ye; Li, Yan-Xiang; Yang, Xiao-Feng; He, Yan-Hao; Zhang, Wei; Jing, Ting; Lin, Rong

2015-10-15

Recent studies demonstrated that the ligand-activated transcription factor peroxisome proliferator-activated receptorα (PPARα) acts in association with histone deacetylase sirtuin 1 (SIRT1) in the regulation of metabolism and inflammation involved in cardiovascular diseases. PPARα activation also participates in the modulation of cell apoptosis. Our previous study found that SIRT1 inhibits the apoptosis of vascular adventitial fibroblasts (VAFs). However, whether the role of PPARα in apoptosis of VAFs is mediated by SIRT1 remains unknown. In this study, we aimed to determine the effect of PPARα agonist fenofibrate on cell apoptosis and SIRT1 expression and related mechanisms in ApoE(-/-) mice and VAFs in vitro. We found that fenofibrate inhibited cell apoptosis in vascular adventitia and up-regulated SIRT1 expression in aorta of ApoE(-/-) mice. Moreover, SIRT1 activator resveratrol (RSV) further enhanced these effects of fenofibrate. In vitro study showed that activation of PPARα by fenofibrate inhibited TNF-α-induced cell apoptosis and cell cycle arrest in VAFs. Meanwhile, fenofibrate up-regulated SIRT1 expression and inhibited SIRT1 translocation from nucleus to cytoplasm in VAFs stimulated with TNF-α. Moreover, the effects of fenofibrate on cell apoptosis and SIRT1 expression in VAFs were reversed by PPARα antagonist GW6471. Importantly, treatment of VAFs with SIRT1 siRNA or pcDNA3.1(+)-SIRT1 showed that the inhibitory effect of fenofibrate on cell apoptosis in VAFs through SIRT1. On the other hand, knockdown of FoxO1 decreased cell apoptosis of VAFs compared with fenofibrate group. Overexpression of FoxO1 increased cell apoptosis of VAFs compared with fenofibrate group. Further study found that fenofibrate decreased the expression of acetylated-FoxO1 in TNF-α-stimulated VAFs, which was abolished by SIRT1 knockdown. Taken together, these findings indicate that activation of PPARα by fenofibrate inhibits cell apoptosis in VAFs partly through the SIRT1

Class IIa histone deacetylases are hormone-activated regulators of FOXO and mammalian glucose homeostasis.

PubMed

Mihaylova, Maria M; Vasquez, Debbie S; Ravnskjaer, Kim; Denechaud, Pierre-Damien; Yu, Ruth T; Alvarez, Jacqueline G; Downes, Michael; Evans, Ronald M; Montminy, Marc; Shaw, Reuben J

2011-05-13

Class IIa histone deacetylases (HDACs) are signal-dependent modulators of transcription with established roles in muscle differentiation and neuronal survival. We show here that in liver, class IIa HDACs (HDAC4, 5, and 7) are phosphorylated and excluded from the nucleus by AMPK family kinases. In response to the fasting hormone glucagon, class IIa HDACs are rapidly dephosphorylated and translocated to the nucleus where they associate with the promoters of gluconeogenic enzymes such as G6Pase. In turn, HDAC4/5 recruit HDAC3, which results in the acute transcriptional induction of these genes via deacetylation and activation of FOXO family transcription factors. Loss of class IIa HDACs in murine liver results in inhibition of FOXO target genes and lowers blood glucose, resulting in increased glycogen storage. Finally, suppression of class IIa HDACs in mouse models of type 2 diabetes ameliorates hyperglycemia, suggesting that inhibitors of class I/II HDACs may be potential therapeutics for metabolic syndrome. Copyright © 2011 Elsevier Inc. All rights reserved.

Metformin reduces lipid accumulation in macrophages by inhibiting FOXO1-mediated transcription of fatty acid-binding protein 4

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

Song, Jun; Division of Cardiothoracic Surgery, Michael E. DeBakey Department of Surgery, Baylor College of Medicine, Houston, TX; Texas Heart Institute at St. Luke's Episcopal Hospital, Houston, TX

2010-02-26

Objective: The accumulation of lipids in macrophages contributes to the development of atherosclerosis. Strategies to reduce lipid accumulation in macrophages may have therapeutic potential for preventing and treating atherosclerosis and cardiovascular complications. The antidiabetic drug metformin has been reported to reduce lipid accumulation in adipocytes. In this study, we examined the effects of metformin on lipid accumulation in macrophages and investigated the mechanisms involved. Methods and results: We observed that metformin significantly reduced palmitic acid (PA)-induced intracellular lipid accumulation in macrophages. Metformin promoted the expression of carnitine palmitoyltransferase I (CPT-1), while reduced the expression of fatty acid-binding protein 4 (FABP4)more » which was involved in PA-induced lipid accumulation. Quantitative real-time PCR showed that metformin regulates FABP4 expression at the transcriptional level. We identified forkhead transcription factor FOXO1 as a positive regulator of FABP4 expression. Inhibiting FOXO1 expression with FOXO1 siRNA significantly reduced basal and PA-induced FABP4 expression. Overexpression of wild-type FOXO1 and constitutively active FOXO1 significantly increased FABP4 expression, whereas dominant negative FOXO1 dramatically decreased FABP4 expression. Metformin reduced FABP4 expression by promoting FOXO1 nuclear exclusion and subsequently inhibiting its activity. Conclusions: Taken together, these results suggest that metformin reduces lipid accumulation in macrophages by repressing FOXO1-mediated FABP4 transcription. Thus, metformin may have a protective effect against lipid accumulation in macrophages and may serve as a therapeutic agent for preventing and treating atherosclerosis in metabolic syndrome.« less

mTOR-Independent Autophagy Inducer Trehalose Rescues against Insulin Resistance-Induced Myocardial Contractile Anomalies: Role of p38 MAPK and Foxo1

PubMed Central

Wang, Qiurong; Ren, Jun

2016-01-01

Insulin resistance is associated with cardiovascular diseases although the precise mechanisms remain elusive. Akt2, a critical member of the Akt family, plays an essential role in insulin signaling. This study was designed to examine the effect of trehalose, an mTOR-independent autophagy inducer, on myocardial function in an Akt2 knockout-induced insulin resistance model. Adult WT and Akt2 knockout (Akt2−/−) mice were administered trehalose (1 mg/g/day, i.p.) for two days and were then given 2% trehalose in drinking water for two more months. Echocardiographic and myocardial mechanics, intracellular Ca2+ properties, glucose tolerance, and autophagy were assessed. Apoptosis and ER stress were evaluated using TUNEL staining, Caspase 3 assay and Western blot. Autophagy and autophagy flux were examined with a focus on p38 mitogen activated protein kinase (MAPK), Forkhead box O (Foxo1) and Akt. Akt2 ablation impaired glucose tolerance, myocardial geometry and function accompanied with pronounced apoptosis, ER stress and dampened autophagy, the effects of which were ameliorated by trehalose treatment. Inhibition of lysosomal activity using bafilomycin A1 negated trehalose–induced induction of autophagy (LC3B–II and p62). Moreover, phosphorylation of p38 MAPK and Foxo1 were upregulated in Akt2−/− mice, the effect of which was attenuated by trehalose. Phosphorylation of Akt was suppressed in Akt2−/− mice and was unaffected by trehalose. In vitro findings revealed that the p38 MAPK activator anisomycin and the Foxo1 inhibitor (through phosphorylation) AS1842856 effectively masked trehalose-offered beneficial cardiomyocyte contractile response against Akt2 ablation. These data suggest that trehalose may rescue against insulin resistance-induced myocardial contractile defect and apoptosis, via autophagy associated with dephosphorylation of p38 MAPK and Foxo1 without affecting phosphorylation of Akt. PMID:27363949

mTOR-Independent autophagy inducer trehalose rescues against insulin resistance-induced myocardial contractile anomalies: Role of p38 MAPK and Foxo1.

PubMed

Wang, Qiurong; Ren, Jun

2016-09-01

Insulin resistance is associated with cardiovascular diseases although the precise mechanisms remain elusive. Akt2, a critical member of the Akt family, plays an essential role in insulin signaling. This study was designed to examine the effect of trehalose, an mTOR-independent autophagy inducer, on myocardial function in an Akt2 knockout-induced insulin resistance model. Adult WT and Akt2 knockout (Akt2(-/-)) mice were administered trehalose (1mg/g/day, i.p.) for two days and were then given 2% trehalose in drinking water for two more months. Echocardiographic and myocardial mechanics, intracellular Ca(2+) properties, glucose tolerance, and autophagy were assessed. Apoptosis and ER stress were evaluated using TUNEL staining, Caspase 3 assay and Western blot. Autophagy and autophagy flux were examined with a focus on p38 mitogen activated protein kinase (MAPK), Forkhead box O (Foxo1) and Akt. Akt2 ablation impaired glucose tolerance, myocardial geometry and function accompanied with pronounced apoptosis, ER stress and dampened autophagy, the effects of which were ameliorated by trehalose treatment. Inhibition of lysosomal activity using bafilomycin A1 negated trehalose-induced induction of autophagy (LC3B-II and p62). Moreover, phosphorylation of p38 MAPK and Foxo1 were upregulated in Akt2(-/-) mice, the effect of which was attenuated by trehalose. Phosphorylation of Akt was suppressed in Akt2(-/-) mice and was unaffected by trehalose. In vitro findings revealed that the p38 MAPK activator anisomycin and the Foxo1 inhibitor (through phosphorylation) AS1842856 effectively masked trehalose-offered beneficial cardiomyocyte contractile response against Akt2 ablation. These data suggest that trehalose may rescue against insulin resistance-induced myocardial contractile defect and apoptosis, via autophagy associated with dephosphorylation of p38 MAPK and Foxo1 without affecting phosphorylation of Akt. Copyright © 2016 Elsevier Ltd. All rights reserved.

dbl-1/TGF-β and daf-12/NHR Signaling Mediate Cell-Nonautonomous Effects of daf-16/FOXO on Starvation-Induced Developmental Arrest.

PubMed

Kaplan, Rebecca E W; Chen, Yutao; Moore, Brad T; Jordan, James M; Maxwell, Colin S; Schindler, Adam J; Baugh, L Ryan

2015-12-01

Nutrient availability has profound influence on development. In the nematode C. elegans, nutrient availability governs post-embryonic development. L1-stage larvae remain in a state of developmental arrest after hatching until they feed. This "L1 arrest" (or "L1 diapause") is associated with increased stress resistance, supporting starvation survival. Loss of the transcription factor daf-16/FOXO, an effector of insulin/IGF signaling, results in arrest-defective and starvation-sensitive phenotypes. We show that daf-16/FOXO regulates L1 arrest cell-nonautonomously, suggesting that insulin/IGF signaling regulates at least one additional signaling pathway. We used mRNA-seq to identify candidate signaling molecules affected by daf-16/FOXO during L1 arrest. dbl-1/TGF-β, a ligand for the Sma/Mab pathway, daf-12/NHR and daf-36/oxygenase, an upstream component of the daf-12 steroid hormone signaling pathway, were up-regulated during L1 arrest in a daf-16/FOXO mutant. Using genetic epistasis analysis, we show that dbl-1/TGF-β and daf-12/NHR steroid hormone signaling pathways are required for the daf-16/FOXO arrest-defective phenotype, suggesting that daf-16/FOXO represses dbl-1/TGF-β, daf-12/NHR and daf-36/oxygenase. The dbl-1/TGF-β and daf-12/NHR pathways have not previously been shown to affect L1 development, but we found that disruption of these pathways delayed L1 development in fed larvae, consistent with these pathways promoting development in starved daf-16/FOXO mutants. Though the dbl-1/TGF-β and daf-12/NHR pathways are epistatic to daf-16/FOXO for the arrest-defective phenotype, disruption of these pathways does not suppress starvation sensitivity of daf-16/FOXO mutants. This observation uncouples starvation survival from developmental arrest, indicating that DAF-16/FOXO targets distinct effectors for each phenotype and revealing that inappropriate development during starvation does not cause the early demise of daf-16/FOXO mutants. Overall, this study shows

dbl-1/TGF-β and daf-12/NHR Signaling Mediate Cell-Nonautonomous Effects of daf-16/FOXO on Starvation-Induced Developmental Arrest

PubMed Central

Moore, Brad T.; Jordan, James M.; Maxwell, Colin S.; Schindler, Adam J.; Baugh, L. Ryan

2015-01-01

Nutrient availability has profound influence on development. In the nematode C. elegans, nutrient availability governs post-embryonic development. L1-stage larvae remain in a state of developmental arrest after hatching until they feed. This “L1 arrest” (or "L1 diapause") is associated with increased stress resistance, supporting starvation survival. Loss of the transcription factor daf-16/FOXO, an effector of insulin/IGF signaling, results in arrest-defective and starvation-sensitive phenotypes. We show that daf-16/FOXO regulates L1 arrest cell-nonautonomously, suggesting that insulin/IGF signaling regulates at least one additional signaling pathway. We used mRNA-seq to identify candidate signaling molecules affected by daf-16/FOXO during L1 arrest. dbl-1/TGF-β, a ligand for the Sma/Mab pathway, daf-12/NHR and daf-36/oxygenase, an upstream component of the daf-12 steroid hormone signaling pathway, were up-regulated during L1 arrest in a daf-16/FOXO mutant. Using genetic epistasis analysis, we show that dbl-1/TGF-β and daf-12/NHR steroid hormone signaling pathways are required for the daf-16/FOXO arrest-defective phenotype, suggesting that daf-16/FOXO represses dbl-1/TGF-β, daf-12/NHR and daf-36/oxygenase. The dbl-1/TGF-β and daf-12/NHR pathways have not previously been shown to affect L1 development, but we found that disruption of these pathways delayed L1 development in fed larvae, consistent with these pathways promoting development in starved daf-16/FOXO mutants. Though the dbl-1/TGF-β and daf-12/NHR pathways are epistatic to daf-16/FOXO for the arrest-defective phenotype, disruption of these pathways does not suppress starvation sensitivity of daf-16/FOXO mutants. This observation uncouples starvation survival from developmental arrest, indicating that DAF-16/FOXO targets distinct effectors for each phenotype and revealing that inappropriate development during starvation does not cause the early demise of daf-16/FOXO mutants. Overall, this study

FoxO1 in dopaminergic neurons regulates energy homeostasis and targets tyrosine hydroxylase

PubMed Central

Doan, Khanh V.; Kinyua, Ann W.; Yang, Dong Joo; Ko, Chang Mann; Moh, Sang Hyun; Shong, Ko Eun; Kim, Hail; Park, Sang-Kyu; Kim, Dong-Hoon; Kim, Inki; Paik, Ji-Hye; DePinho, Ronald A.; Yoon, Seul Gi; Kim, Il Yong; Seong, Je Kyung; Choi, Yun-Hee; Kim, Ki Woo

2016-01-01

Dopaminergic (DA) neurons are involved in the integration of neuronal and hormonal signals to regulate food consumption and energy balance. Forkhead transcriptional factor O1 (FoxO1) in the hypothalamus plays a crucial role in mediation of leptin and insulin function. However, the homoeostatic role of FoxO1 in DA system has not been investigated. Here we report that FoxO1 is highly expressed in DA neurons and mice lacking FoxO1 specifically in the DA neurons (FoxO1 KODAT) show markedly increased energy expenditure and interscapular brown adipose tissue (iBAT) thermogenesis accompanied by reduced fat mass and improved glucose/insulin homoeostasis. Moreover, FoxO1 KODAT mice exhibit an increased sucrose preference in concomitance with higher dopamine and norepinephrine levels. Finally, we found that FoxO1 directly targets and negatively regulates tyrosine hydroxylase (TH) expression, the rate-limiting enzyme of the catecholamine synthesis, delineating a mechanism for the KO phenotypes. Collectively, these results suggest that FoxO1 in DA neurons is an important transcriptional factor that directs the coordinated control of energy balance, thermogenesis and glucose homoeostasis. PMID:27681312

FOXO1 regulates VEGFA expression and promotes angiogenesis in healing wounds.

PubMed

Jeon, Hyeran Helen; Yu, Quan; Lu, Yongjian; Spencer, Evelyn; Lu, Chanyi; Milovanova, Tatyana; Yang, Yang; Zhang, Chenying; Stepanchenko, Olga; Vafa, Rameen P; Coelho, Paulo G; Graves, Dana T

2018-03-25

Angiogenesis is a critical aspect of wound healing. We investigated the role of keratinocytes in promoting angiogenesis in mice with lineage-specific deletion of the transcription factor FOXO1. The results indicate that keratinocyte-specific deletion of Foxo1 reduces VEGFA expression in mucosal and skin wounds and leads to reduced endothelial cell proliferation, reduced angiogenesis, and impaired re-epithelialization and granulation tissue formation. In vitro FOXO1 was needed for VEGFA transcription and expression. In a porcine dermal wound-healing model that closely resembles healing in humans, local application of a FOXO1 inhibitor reduced angiogenesis. This is the first report that FOXO1 directly regulates VEGFA expression and that FOXO1 is needed for normal angiogenesis during wound healing. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

MiR-135 post-transcriptionally regulates FOXO1 expression and promotes cell proliferation in human malignant melanoma cells.

PubMed

Ren, Jian-Wen; Li, Zhang-Jun; Tu, Chen

2015-01-01

Malignant melanoma is the deadliest form of all skin cancers. Recently, microRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression by targeted repression of transcription and translation and play essential roles during cancer development. Our study showed that miR-135a is upregulated in malignant melanoma tissues and cell lines by using Real-time PCR assay. Enforced expression of miR-135a in malignant melanoma cells promotes cell proliferation, tumorigenicity, and cell cycle progression, whereas inhibition of miR-135a reverses the function. Additionally, we demonstrated FOXO1 is a direct target of miR-135a and transcriptionally down-regulated by miR-135a. Ectopic expression of miR-135a led to downregulation of the FOXO1 protein, resulting in upregulation of Cyclin D1, and downregulation of P21(Cip1) and P27(Kip1) through AKT pathway. Our findings suggested that miR-135a represents a potential onco-miRNA and plays an important role in malignant melanoma progression by suppressing FOXO1 expression.

Foxo Transcription Factors Blunt Cardiac Hypertrophy by Inhibiting Calcineurin Signaling

PubMed Central

Ni, Yan G.; Berenji, Kambeez; Wang, Na; Oh, Misook; Sachan, Nita; Dey, Asim; Cheng, Jun; Lu, Guangrong; Morris, David J.; Castrillon, Diego H.; Gerard, Robert D.; Rothermel, Beverly A.; Hill, Joseph A.

2014-01-01

Background Cellular hypertrophy requires coordinated regulation of progrowth and antigrowth mechanisms. In cultured neonatal cardiomyocytes, Foxo transcription factors trigger an atrophy-related gene program that counters hypertrophic growth. However, downstream molecular events are not yet well defined. Methods and Results Here, we report that expression of either Foxo1 or Foxo3 in cardiomyocytes attenuates calcineurin phosphatase activity and inhibits agonist-induced hypertrophic growth. Consistent with these results, Foxo proteins decrease calcineurin phosphatase activity and repress both basal and hypertrophic agonist-induced expression of MCIP1.4, a direct downstream target of the calcineurin/NFAT pathway. Furthermore, hearts from Foxo3-null mice exhibit increased MCIP1.4 abundance and a hypertrophic phenotype with normal systolic function at baseline. Together, these results suggest that Foxo proteins repress cardiac growth at least in part through inhibition of the calcineurin/NFAT pathway. Given that hypertrophic growth of the heart occurs in multiple contexts, our findings also suggest that certain hypertrophic signals are capable of overriding the antigrowth program induced by Foxo. Consistent with this, multiple hypertrophic agonists triggered inactivation of Foxo proteins in cardiomyocytes through a mechanism requiring the PI3K/Akt pathway. In addition, both Foxo1 and Foxo3 are phosphorylated and consequently inactivated in hearts undergoing hypertrophic growth induced by hemodynamic stress. Conclusions This study suggests that inhibition of the calcineurin/NFAT signaling cascade by Foxo and release of this repressive action by the PI3K/Akt pathway are important mechanisms whereby Foxo factors govern cell growth in the heart. PMID:16952979

Foxo transcription factors blunt cardiac hypertrophy by inhibiting calcineurin signaling.

PubMed

Ni, Yan G; Berenji, Kambeez; Wang, Na; Oh, Misook; Sachan, Nita; Dey, Asim; Cheng, Jun; Lu, Guangrong; Morris, David J; Castrillon, Diego H; Gerard, Robert D; Rothermel, Beverly A; Hill, Joseph A

2006-09-12

Cellular hypertrophy requires coordinated regulation of progrowth and antigrowth mechanisms. In cultured neonatal cardiomyocytes, Foxo transcription factors trigger an atrophy-related gene program that counters hypertrophic growth. However, downstream molecular events are not yet well defined. Here, we report that expression of either Foxo1 or Foxo3 in cardiomyocytes attenuates calcineurin phosphatase activity and inhibits agonist-induced hypertrophic growth. Consistent with these results, Foxo proteins decrease calcineurin phosphatase activity and repress both basal and hypertrophic agonist-induced expression of MCIP1.4, a direct downstream target of the calcineurin/NFAT pathway. Furthermore, hearts from Foxo3-null mice exhibit increased MCIP1.4 abundance and a hypertrophic phenotype with normal systolic function at baseline. Together, these results suggest that Foxo proteins repress cardiac growth at least in part through inhibition of the calcineurin/NFAT pathway. Given that hypertrophic growth of the heart occurs in multiple contexts, our findings also suggest that certain hypertrophic signals are capable of overriding the antigrowth program induced by Foxo. Consistent with this, multiple hypertrophic agonists triggered inactivation of Foxo proteins in cardiomyocytes through a mechanism requiring the PI3K/Akt pathway. In addition, both Foxo1 and Foxo3 are phosphorylated and consequently inactivated in hearts undergoing hypertrophic growth induced by hemodynamic stress. This study suggests that inhibition of the calcineurin/NFAT signaling cascade by Foxo and release of this repressive action by the PI3K/Akt pathway are important mechanisms whereby Foxo factors govern cell growth in the heart.

Akt/FOXO3a signaling modulates the endothelial stress response through regulation of heat shock protein 70 expression.

PubMed

Kim, Hyo-Soo; Skurk, Carsten; Maatz, Henrike; Shiojima, Ichiro; Ivashchenko, Yuri; Yoon, Suk-Won; Park, Young-Bae; Walsh, Kenneth

2005-06-01

To identify new antiapoptotic targets of the PI3K-Akt signaling pathway in endothelial cells, adenovirus-mediated Akt1 gene transfer and oligonucleotide microarrays were used to examine Akt-regulated transcripts. DNA microarray analysis revealed that HSP70 expression underwent the greatest fold activation of 12,532 transcripts examined in human umbilical vein endothelial cells (HUVEC) transduced with constitutively active Akt1. Akt1 gene transfer increased HSP70 transcript expression by 24.8-fold as determined by quantitative PCR and promoted a dose-dependent up-regulation of HSP70 protein as determined by Western immunoblot analysis. Gene transfer of FOXO3a, a downstream target of Akt in endothelial cells, significantly suppressed both basal and stress-induced HSP70 protein expression. FOXO3a induced caspase-9-dependent apoptosis in HUVEC, and cotransduction with Ad-HSP70 rescued endothelial cells from FOXO3a-induced apoptosis under basal and stress conditions. Our results identify HSP70 as a new antiapoptotic target of Akt-FOXO3a signaling in endothelial cells that controls viability through modulation of the stress-induced intrinsic cell death pathway.

FoxO1 integrates direct and indirect effects of insulin on hepatic glucose production and glucose utilization

PubMed Central

O-Sullivan, InSug; Zhang, Wenwei; Wasserman, David H.; Liew, Chong Wee; Liu, Jonathan; Paik, Jihye; DePinho, Ronald A.; Stolz, Donna Beer; Kahn, C. Ronald; Schwartz, Michael W.; Unterman, Terry G.

2016-01-01

FoxO proteins are major targets of insulin action. To better define the role of FoxO1 in mediating insulin effects in the liver, we generated liver-specific insulin receptor knockout (LIRKO) and IR/FoxO1 double knockout (LIRFKO) mice. Here we show that LIRKO mice are severely insulin resistant based on glucose, insulin and C-peptide levels, and glucose and insulin tolerance tests, and genetic deletion of hepatic FoxO1 reverses these effects. 13C-glucose and insulin clamp studies indicate that regulation of both hepatic glucose production (HGP) and glucose utilization is impaired in LIRKO mice, and these defects are also restored in LIRFKO mice corresponding to changes in gene expression. We conclude that (1) inhibition of FoxO1 is critical for both direct (hepatic) and indirect effects of insulin on HGP and utilization, and (2) extrahepatic effects of insulin are sufficient to maintain normal whole-body and hepatic glucose metabolism when liver FoxO1 activity is disrupted. PMID:25963540

GCN5L1 modulates cross-talk between mitochondria and cell signaling to regulate FoxO1 stability and gluconeogenesis.

PubMed

Wang, Lingdi; Scott, Iain; Zhu, Lu; Wu, Kaiyuan; Han, Kim; Chen, Yong; Gucek, Marjan; Sack, Michael N

2017-09-12

The mitochondrial enriched GCN5-like 1 (GCN5L1) protein has been shown to modulate mitochondrial protein acetylation, mitochondrial content and mitochondrial retrograde signaling. Here we show that hepatic GCN5L1 ablation reduces fasting glucose levels and blunts hepatic gluconeogenesis without affecting systemic glucose tolerance. PEPCK and G6Pase transcript levels are downregulated in hepatocytes from GCN5L1 liver specific knockout mice and their upstream regulator, FoxO1 protein levels are decreased via proteasome-dependent degradation and via reactive oxygen species mediated ERK-1/2 phosphorylation. ERK inhibition restores FoxO1, gluconeogenic enzyme expression and glucose production. Reconstitution of mitochondrial-targeted GCN5L1 blunts mitochondrial ROS, ERK activation and increases FoxO1, gluconeogenic enzyme expression and hepatocyte glucose production. We suggest that mitochondrial GCN5L1 modulates post-translational control of FoxO1, regulates gluconeogenesis and controls metabolic pathways via mitochondrial ROS mediated ERK activation. Exploring mechanisms underpinning GCN5L1 mediated ROS signaling may expand our understanding of the role of mitochondria in gluconeogenesis control.Hepatic gluconeogenesis is tightly regulated at transcriptional level and is essential for survival during prolonged fasting. Here Wang et al. show that the mitochondrial enriched GCN5-like 1 protein controls hepatic glucose production by regulating FoxO1 protein levels via proteasome-dependent degradation and, in turn, gluconeogenic gene expression.

WNK1 regulates skeletal muscle cell hypertrophy by modulating the nuclear localization and transcriptional activity of FOXO4.

PubMed

Mandai, Shintaro; Mori, Takayasu; Nomura, Naohiro; Furusho, Taisuke; Arai, Yohei; Kikuchi, Hiroaki; Sasaki, Emi; Sohara, Eisei; Rai, Tatemitsu; Uchida, Shinichi

2018-06-14

With-no-lysine (K) (WNK) kinases, which are mutated in the inherited form of hypertension pseudohypoaldosteronism type II, are essential regulators of membrane ion transporters. Here, we report that WNK1 positively regulates skeletal muscle cell hypertrophy via mediating the function of the pro-longevity transcription factor forkhead box protein O4 (FOXO4) independent of the conventional WNK signaling pathway linking SPS/STE20-related proline-alanine-rich kinase (SPAK)/oxidative stress response kinase 1 (OSR1) to downstream effector ion transporters. Small interfering RNA (siRNA)-mediated silencing of WNK1, but not SPAK/OSR1 kinases, induced myotube atrophy and remarkable increases in the mRNA expression of the muscle atrophy ubiquitin ligases MAFbx and MuRF1 in C2C12 mouse skeletal muscle cells. WNK1 silencing also increased FOXO4 nuclear localization, and co-transfection of Foxo4-targeted siRNA completely reversed the myotube atrophy and upregulation of atrogene transcription induced by WNK1 silencing. We further illustrated that WNK1 protein abundance in skeletal muscle was increased by chronic voluntary wheel running exercise (hypertrophic stimulus) and markedly decreased by adenine-induced chronic kidney disease (atrophic stimulus) in mice. These findings suggest that WNK1 is involved in the physiological regulation of mammalian skeletal muscle hypertrophy and atrophy via interactions with FOXO4. The WNK1-FOXO4 axis may be a potential therapeutic target in human diseases causing sarcopenia.

PAX3-FOXO1: Zooming in on an "undruggable" target.

PubMed

Wachtel, Marco; Schäfer, Beat W

2018-06-01

Driver oncogenes are prime targets for therapy in tumors many of which, including leukemias and sarcomas, express recurrent fusion transcription factors. One specific example for such a cancer type is alveolar rhabdomyosarcoma, which is associated in the majority of cases with the fusion protein PAX3-FOXO1. Since fusion transcription factors are challenging targets for development of small molecule inhibitors, indirect inhibitory strategies for this type of oncogenes represent a more promising approach. One can envision strategies at different molecular levels including upstream modifiers and activators, epigenetic and transcriptional co-regulators, and downstream effector targets. In this review, we will discuss the current knowledge regarding potential therapeutic targets that might contribute to indirect interference with PAX3-FOXO1 activity in alveolar rhabdomyosarcoma at the different molecular levels and extrapolate these findings to fusion transcription factors in general. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Excessive fluoride reduces Foxo1 expression in dental epithelial cells of the rat incisor.

PubMed

Gao, Jianghong; Ruan, Jianping; Gao, Liping

2014-10-01

Enamel fluorosis is characterized by hypomineralization, and forkhead box O1 (Foxo1) is essential for mouse enamel biomineralization. This study investigated the effect of fluoride on Foxo1 expression and its implications for enamel fluorosis. Mandibular incisors were extracted from Sprague Dawley rats treated for 3 months with water containing 0, 50, or 100 p.p.m. F⁻. Immunohistochemistry was used to localize and quantify FOXO1 expression in dental epithelial layer cells of the incisors. The effect of fluoride on expression of Foxo1, kallikrein-4 (Klk4), and amelotin (Amtn) mRNAs was analyzed by real-time RT-PCR, and western blotting was used to measure total and nuclear FOXO1 protein levels in mature dental epithelial cells. The results revealed that nuclear FOXO1 was mainly localized in the transition and the mature ameloblasts and exhibited weaker expression in the rats exposed to fluoride. In addition to the reduced levels of Foxo1, Klk4, and AmtnmRNAs, the protein levels of total and nuclearFOXO1 were decreased in the mature dental epithelial cells exposed to fluoride. Thus, excessive fluoride may have an effect on the expression levels of Foxo1 in dental epithelial cells and thereby affect hypomineralization of the enamel during fluorosis. © 2014 Eur J Oral Sci.

FOXO Regulates Organ-Specific Phenotypic Plasticity In Drosophila

PubMed Central

Tang, Hui Yuan; Smith-Caldas, Martha S. B.; Driscoll, Michael V.; Salhadar, Samy; Shingleton, Alexander W.

2011-01-01

Phenotypic plasticity, the ability for a single genotype to generate different phenotypes in response to environmental conditions, is biologically ubiquitous, and yet almost nothing is known of the developmental mechanisms that regulate the extent of a plastic response. In particular, it is unclear why some traits or individuals are highly sensitive to an environmental variable while other traits or individuals are less so. Here we elucidate the developmental mechanisms that regulate the expression of a particularly important form of phenotypic plasticity: the effect of developmental nutrition on organ size. In all animals, developmental nutrition is signaled to growing organs via the insulin-signaling pathway. Drosophila organs differ in their size response to developmental nutrition and this reflects differences in organ-specific insulin-sensitivity. We show that this variation in insulin-sensitivity is regulated at the level of the forkhead transcription factor FOXO, a negative growth regulator that is activated when nutrition and insulin signaling are low. Individual organs appear to attenuate growth suppression in response to low nutrition through an organ-specific reduction in FOXO expression, thereby reducing their nutritional plasticity. We show that FOXO expression is necessary to maintain organ-specific differences in nutritional-plasticity and insulin-sensitivity, while organ-autonomous changes in FOXO expression are sufficient to autonomously alter an organ's nutritional-plasticity and insulin-sensitivity. These data identify a gene (FOXO) that modulates a plastic response through variation in its expression. FOXO is recognized as a key player in the response of size, immunity, and longevity to changes in developmental nutrition, stress, and oxygen levels. FOXO may therefore act as a more general regulator of plasticity. These data indicate that the extent of phenotypic plasticity may be modified by changes in the expression of genes involved in

[Relationship between FoxO1 Expression and Wound Age during Skin Incised Wound Healing].

PubMed

Chen, Y; Ji, X Y; Fan, Y Y; Yu, L S

2018-02-01

To investigate FoxO1 expression and its time-dependent changes during the skin incised wound healing. After the establishment of the skin incised wound model in mice, the FoxO1 expression of skin in different time periods was detected by immunohistochemistry and Western blotting. Immunohistochemistry staining showed that FoxO1 was weakly expressed in a few fibroblasts of epidermis, hair follicles, sebaceous glands, vessel endothelium and dermis in the control group. The FoxO1 expression was enhanced in the epidermis and skin appendages around the wound during 6-12 h after injury, which could be detected in the infiltrating neutrophils and a small number of monocytes. FoxO1 was mainly expressed in monocytes during 1-3 d after injury, and in neovascular endothelial cells and fibroblasts during 5-10 d. On the 14th day after injury, the FoxO1 expression still could be detected in a few fibroblasts. The Western blotting results showed that the FoxO1 expression quantity of the tissue samples in injury group was higher than in control group. The FoxO1 expression peaked at 12 h and 7 d after injury. FoxO1 is time-dependently expressed in skin wound healing, which can be a useful marker for wound age determination. Copyright© by the Editorial Department of Journal of Forensic Medicine.

SIRT1 and FOXO1 mRNA expression in PBMC correlates to physical activity in COPD patients

PubMed Central

Taka, Chihiro; Hayashi, Ryuji; Shimokawa, Kazuki; Tokui, Kotaro; Okazawa, Seisuke; Kambara, Kenta; Inomata, Minehiko; Yamada, Toru; Matsui, Shoko; Tobe, Kazuyuki

2017-01-01

Background Physical activity (PA) is considered as one of the most important prognostic predictors in chronic obstructive pulmonary disease (COPD) patients. Longevity gene, SIRT1, is reported to be involved in the pathogenesis of COPD by regulating the signaling pathways of oxidative stress, inflammation, and aging. We hypothesize that SIRT1 and related genes are also associated with the benefits of PA in COPD patients. Methods Eighteen COPD outpatients were enrolled in this study, and their PA level was assessed with an accelerometer. We assessed the SIRT1 and related genes mRNA expression levels in the peripheral blood mononuclear cells (PBMCs) of the subjects. We carried out respiratory function testing, blood gas analysis, the 6-minute walk test, and measurement of the cross-sectional area of the erector spinae muscles (ESMCSA) by chest computed tomography. We analyzed the association of PA with the results of each of the examinations. Results The mean age was 72±9 years, and the mean forced expiratory volume in 1 second was 1.4±0.56 L (52%±19% predicted). Our findings revealed a correlation between the daily PA and ESMCSA. The SIRT1 and Forkhead box O (FOXO)1 mRNA expression levels in PBMCs were positively correlated with moderate-PA time (r=0.60, p=0.008 for SIRT1 and r=0.59, p=0.01 for FOXO1). PMID:29138552

GL-V9, a new synthetic flavonoid derivative, ameliorates DSS-induced colitis against oxidative stress by up-regulating Trx-1 expression via activation of AMPK/FOXO3a pathway.

PubMed

Zhao, Yue; Sun, Yang; Ding, Youxiang; Wang, Xiaoping; Zhou, Yuxin; Li, Wenjun; Huang, Shaoliang; Li, Zhiyu; Kong, Lingyi; Guo, Qinglong; Lu, Na

2015-09-22

GL-V9, a new synthesized flavonoid derivative, has been reported to possess anti-cancer properties in our previous studies. Uncontrolled overproduction of reactive oxygen species (ROS) has been implicated in oxidative damage of inflammatory bowel disease (IBD). In this study, we aimed to investigate the protective effect of GL-V9 against dextran sulfate sodium (DSS)-induced colitis. GL-V9 attenuated DSS-induced body weight loss, colon length shortening and colonic pathological damage. GL-V9 also inhibited inflammatory cells infiltration and decreased myeloperoxidase (MPO) and inducible nitric oxide synthase (iNOS) activities. Moreover, GL-V9 inhibited ROS and malondialdehyde (MDA) generation, but enhanced superoxide dismutase (SOD), glutathione (GSH) and total antioxidant capacity. GL-V9 reduced pro-inflammatory cytokines production in serum and colon as well. Mechanically, GL-V9 could increase Trx-1 via activation of AMPK/FOXO3a to suppress DSS-induced colonic oxidative stress. Furthermore, GL-V9 decreased pro-inflammatory cytokines and ROS production and increased the antioxidant defenses in the mouse macrophage cells RAW264.7 by promoting Trx-1 expression. In conclusion, our study demonstrated that GL-V9 attenuated DSS-induced colitis against oxidative stress by up-regulating Trx-1 via activation of AMPK/FOXO3a pathway, suggesting that GL-V9 might be a potential effective drug for colitis.

Long non-coding RNA MALAT1 interacts with transcription factor Foxo1 to regulate SIRT1 transcription in high glucose-induced HK-2 cells injury.

PubMed

Zhou, Ling; Xu, De-Yu; Sha, Wen-Gang; Shen, Lei; Lu, Guo-Yuan

2018-06-18

Tubular injury is considered as a crucial pathological feature of diabetic nephropathy. LncRNA MALAT1 is involved in diabetic complications. Hence the role of MALAT1 in high glucose-induced renal tubular epithelial cells (HK-2) injury deserves investigation. The diabetic mice model was established with streptozotocin (STZ) injection. The expression of NEAT1, SIRT1, and Foxo1 mRNA and protein was determined with qRT-PCR and western blot, respectively. The serum creatinine and urinary albumin were examined by enzyme linked immunosorbent assay (ELISA). Interaction between MALAT1 and Foxo1 was detected with RIP and RNA pull-down assay, respectively. Dual luciferase reporter assay was used to evaluate the binding between Foxo1 and SIRT1. LncRNA MALAT1 was up-regulated in kidney tissues of diabetic mice and in HK-2 cells treated with high glucose, while the expression of SIRT1 was decreased. Interaction between MALAT1 and Foxo1 was observed in HK-2 cells and the interaction was promoted by high glucose treatment. Foxo1 activated SIRT1 transcription by binding to its promoter, and MALAT1 repressed SIRT1 expression through targeting Foxo1. LncRNA MALAT1 interacts with transcription factor Foxo1 to represses SIRT1 transcription in high glucose incubated HK-2 cells, which promotes high glucose-induced HK-2 cells injury. Copyright © 2018. Published by Elsevier Inc.

Transcription Factor Foxo1 Is a Negative Regulator of NK Cell Maturation and Function

PubMed Central

Deng, Youcai; Kerdiles, Yann; Chu, Jianhong; Yuan, Shunzong; Wang, Youwei; Chen, Xilin; Mao, Hsiaoyin; Zhang, Lingling; Zhang, Jianying; Hughes, Tiffany; Deng, Yafei; Zhang, Qi; Wang, Fangjie; Zou, Xianghong; Liu, Chang-Gong; Freud, Aharon G.; Li, Xiaohui; Caligiuri, Michael A; Vivier, Eric; Yu, Jianhua

2015-01-01

SUMMARY Little is known about the role of negative regulators in controlling natural killer (NK) cell development and effector functions. Foxo1 is a multifunctional transcription factor of the forkhead family. Using a mouse model of conditional deletion in NK cells, we found that Foxo1 negatively controlled NK cell differentiation and function. Immature NK cells expressed abundant Foxo1 and little Tbx21 relative to mature NK cells, but these two transcription factors reversed their expression as NK cells proceeded through development. Foxo1 promoted NK cell homing to lymph nodes through upregulating CD62L expression, and impaired late-stage maturation and effector functions by repressing Tbx21 expression. Loss of Foxo1 rescued the defect in late-stage NK cell maturation in heterozygous Tbx21+/− mice. Collectively, our data reveal a regulatory pathway by which the negative regulator Foxo1 and the positive regulator Tbx21 play opposing roles in controlling NK cell development and effector functions. PMID:25769609

Effects of miR-155 on proliferation and apoptosis by regulating FoxO3a/BIM in liver cancer cell line HCCLM3.

PubMed

Liao, W-W; Zhang, C; Liu, F-R; Wang, W-J

2018-03-01

MiR-155 has been shown to be up-regulated in hepatocellular carcinoma (HCC) patients. B-cell lymphoma-2 (Bcl-2) interacting mediator of cell death (BIM) regulates cell proliferation and apoptosis, as its down-regulation is involved in HCC onset. Transcriptional factor FoxO3a mediates BIM expression and is related to HCC pathogenesis. Bioinformatics analysis showed targeted regulation of FoxO3a by miR-155. This study aims to investigate whether miR-155 plays a role in mediating FoxO3a/BIM signal pathway and HCC occurrence. HCC patients were collected for tumor and adjacent tissues, in which microRNA-155 (miR-155) and FoxO3a expressions were examined. In vitro cultured HCCLM3, HepG2 and L-02 cells were tested for basal apoptotic rate by flow cytometry and were compared for miR-155 and FoxO3a expression. Dual-luciferase reporter gene assay demonstrated the targeted relationship between miR-155 and FoxO3a. HCCLM3 cells were treated with miR-155 inhibitor and/or FoxO3a-pMD18-T. Cell apoptosis and proliferation were examined by using flow cytometry and MTT assays, respectively. Western blot and spectrometry assay were employed to quantify the FoxO3a, BIM expressions, and caspase activity. Compared to adjacent tissues, HCC tissues had significantly higher miR-155 and significantly lower FoxO3a expression (p<0.05). HCCLM3 and HepG2 cells had significantly lower FoxO3a expression and basal apoptotic rate compared to L02 cells, whilst miR-155 level was significantly higher (p<0.05). MiR-155 targeted and inhibited 3'-UTR of FoxO3a, increasing BIM expression, caspase-3, and caspase-9 activities, and enhancing cell apoptosis and weakening proliferation. HCC tissues elevated the miR-155 and suppressed the FoxO3a expressions. MiR-155 targeted and inhibited FoxO3a expression to suppress the BIM, depress caspase-3 and caspase-9 activities, therefore inhibiting the HCC cell apoptosis and facilitating proliferation.

Induction of tumor apoptosis through a circular RNA enhancing Foxo3 activity.

PubMed

Du, William W; Fang, Ling; Yang, Weining; Wu, Nan; Awan, Faryal Mehwish; Yang, Zhenguo; Yang, Burton B

2017-02-01

Circular RNAs are a class of non-coding RNAs that are receiving extensive attention. Despite reports showing circular RNAs acting as microRNA sponges, the biological functions of circular RNAs remain largely unknown. We show that in patient tumor samples and in a panel of cancer cells, circ-Foxo3 was minimally expressed. Interestingly, during cancer cell apoptosis, the expression of circ-Foxo3 was found to be significantly increased. We found that silencing endogenous circ-Foxo3 enhanced cell viability, whereas ectopic expression of circ-Foxo3 triggered stress-induced apoptosis and inhibited the growth of tumor xenografts. Also, expression of circ-Foxo3 increased Foxo3 protein levels but repressed p53 levels. By binding to both, circ-Foxo3 promoted MDM2-induced p53 ubiquitination and subsequent degradation, resulting in an overall decrease of p53. With low binding affinity to Foxo3 protein, circ-Foxo3 prevented MDM2 from inducing Foxo3 ubiquitination and degradation, resulting in increased levels of Foxo3 protein. As a result, cell apoptosis was induced by upregulation of the Foxo3 downstream target PUMA.

Association Between Genetic Variation in FOXO3 and Reductions in Inflammation and Disease Activity in Inflammatory Polyarthritis

PubMed Central

Viatte, Sebastien; Lee, James C.; Fu, Bo; Espéli, Marion; Lunt, Mark; De Wolf, Jack N. E.; Wheeler, Lily; Reynolds, John A.; Castelino, Madhura; Symmons, Deborah P. M.; Lyons, Paul A.

2016-01-01

Objective Genetic variation in FOXO3 (tagged by rs12212067) has been associated with a milder course of rheumatoid arthritis (RA) and shown to limit monocyte‐driven inflammation through a transforming growth factor β1–dependent pathway. This genetic association, however, has not been consistently observed in other RA cohorts. We sought to clarify the contribution of FOXO3 to prognosis in RA by combining detailed analysis of nonradiographic disease severity measures with an in vivo model of arthritis. Methods Collagen‐induced arthritis, the most commonly used mouse model of RA, was used to assess how Foxo3 contributes to arthritis severity. Using clinical, serologic, and biochemical methods, the arthritis that developed in mice carrying a loss‐of‐function mutation in Foxo3 was compared with that which occurred in littermate controls. The association of rs12212067 with nonradiographic measures of RA severity, including the C‐reactive protein level, the swollen joint count, the tender joint count, the Disease Activity Score in 28 joints, and the Health Assessment Questionnaire score, were modeled longitudinally in a large prospective cohort of patients with early RA. Results Loss of Foxo3 function resulted in more severe arthritis in vivo (both clinically and histologically) and was associated with higher titers of anticollagen antibodies and interleukin‐6 in the blood. Similarly, rs12212067 (a single‐nucleotide polymorphism that increases FOXO3 transcription) was associated with reduced inflammation, both biochemically and clinically, and with lower RA activity scores. Conclusion Consistent with its known role in restraining inflammatory responses, FOXO3 limits the severity of in vivo arthritis and, through genetic variation that increases its transcription, is associated with reduced inflammation and disease activity in RA patients, effects that result in less radiographic damage. PMID:27214848

A Genome-Wide RNAi Screen Identifies FOXO4 as a Metastasis-Suppressor through Counteracting PI3K/AKT Signal Pathway in Prostate Cancer

PubMed Central

Su, Bing; Gao, Lingqiu; Baranowski, Catherine; Gillard, Bryan; Wang, Jianmin; Ransom, Ryan; Ko, Hyun-Kyung; Gelman, Irwin H.

2014-01-01

Activation of the PI3K/AKT signal pathway is a known driving force for the progression to castration-recurrent prostate cancer (CR-CaP), which constitutes the major lethal phenotype of CaP. Here, we identify using a genomic shRNA screen the PI3K/AKT-inactivating downstream target, FOXO4, as a potential CaP metastasis suppressor. FOXO4 protein levels inversely correlate with the invasive potential of a panel of human CaP cell lines, with decreased mRNA levels correlating with increased incidence of clinical metastasis. Knockdown (KD) of FOXO4 in human LNCaP cells causes increased invasion in vitro and lymph node (LN) metastasis in vivo without affecting indices of proliferation or apoptosis. Increased Matrigel invasiveness was found by KD of FOXO1 but not FOXO3. Comparison of differentially expressed genes affected by FOXO4-KD in LNCaP cells in culture, in primary tumors and in LN metastases identified a panel of upregulated genes, including PIP, CAMK2N1, PLA2G16 and PGC, which, if knocked down by siRNA, could decrease the increased invasiveness associated with FOXO4 deficiency. Although only some of these genes encode FOXO promoter binding sites, they are all RUNX2-inducible, and RUNX2 binding to the PIP promoter is increased in FOXO4-KD cells. Indeed, the forced expression of FOXO4 reversed the increased invasiveness of LNCaP/shFOXO4 cells; the forced expression of FOXO4 did not alter RUNX2 protein levels, yet it decreased RUNX2 binding to the PIP promoter, resulting in PIP downregulation. Finally, there was a correlation between FOXO4, but not FOXO1 or FOXO3, downregulation and decreased metastasis-free survival in human CaP patients. Our data strongly suggest that increased PI3K/AKT-mediated metastatic invasiveness in CaP is associated with FOXO4 loss, and that mechanisms to induce FOXO4 re-expression might suppress CaP metastatic aggressiveness. PMID:24983969

Anoxia-responsive regulation of the FoxO transcription factors in freshwater turtles, Trachemys scripta elegans.

PubMed

Krivoruchko, Anastasia; Storey, Kenneth B

2013-11-01

The forkhead class O (FoxO) transcription factors are important regulators of multiple aspects of cellular metabolism. We hypothesized that activation of these transcription factors could play crucial roles in low oxygen survival in the anoxia-tolerant turtle, Trachemys scripta elegans. Two FoxOs, FoxO1 and FoxO3, were examined in turtle tissues in response to 5 and 20h of anoxic submergence using techniques of RT-PCR, western immunoblotting and DNA-binding assays to assess activation. Transcript levels of FoxO-responsive genes were also quantified using RT-PCR. FoxO1 was anoxia-responsive in the liver, with increases in transcript levels, protein levels, nuclear levels and DNA-binding of 1.7-4.8fold in response to anoxia. Levels of phosphorylated FoxO1 also decreased to 57% of control values in response to 5h of anoxia, indicating activation. FoxO3 was activated in the heart, kidney and liver in response to anoxia, with nuclear levels increasing by 1.5-3.7fold and DNA-binding activity increasing by 1.3-2.9fold. Transcript levels of two FoxO-target genes, p27kip1 and catalase, also rose by 2.4-2.5fold in the turtle liver under anoxia. The results suggest that the FoxO transcription factors are activated in response to anoxia in T. scripta elegans, potentially contributing to the regulation of stress resistance and metabolic depression. This study provides the first demonstration of activation of FoxOs in a natural model for vertebrate anoxia tolerance, further improving understanding of how tissues can survive without oxygen. © 2013.

Testosterone regulation of Akt/mTORC1/FoxO3a Signaling in Skeletal Muscle

PubMed Central

White, James P.; Gao, Song; Puppa, Melissa J.; Sato, Shuichi; Welle, Stephen L.; Carson, James A.

2012-01-01

Low endogenous testosterone production, known as hypogonadism is commonly associated with conditions inducing muscle wasting. Akt signaling can control skeletal muscle mass through mTOR regulation of protein synthesis and FoxO regulation of protein degradation, and this pathway has been previously identified as a target of androgen signaling. However, the testosterone sensitivity of Akt/mTOR signaling requires further understanding in order to grasp the significance of varied testosterone levels seen with wasting disease on muscle protein turnover regulation. Therefore, the purpose of this study is to determine the effect of androgen availability on muscle Akt/mTORC1/FoxO3a regulation in skeletal muscle and cultured C2C12 myotubes. C57BL/6 mice were either castrated for 42 days or castrated and treated with the nandrolone decanoate (ND) (6 mg/kg bw/wk). Testosterone loss (TL) significantly decreased volitional grip strength, body weight, and gastrocnemius (GAS) muscle mass, and ND reversed these changes. Related to muscle mass regulation, TL decreased muscle IGF-1 mRNA, the rate of myofibrillar protein synthesis, Akt phosphorylation, and the phosphorylation of Akt targets, GSK3β, PRAS40 and FoxO3a. TL induced expression of FoxO transcriptional targets, MuRF1, atrogin1 and REDD1. Muscle AMPK and raptor phosphorylation, mTOR inhibitors, were not altered by low testosterone. ND restored IGF-1 expression and Akt/mTORC1 signaling while repressing expression of FoxO transcriptional targets. Testosterone (T) sensitivity of Akt/mTORC1 signaling was examined in C2C12 myotubes, and mTOR phosphorylation was induced independent of Akt activation at low T concentrations, while a higher T concentration was required to activate Akt signaling. Interestingly, low concentration T was sufficient to amplify myotube mTOR and Akt signaling after 24h of T withdrawal, demonstrating the potential in cultured myotubes for a T initiated positive feedback mechanism to amplify Akt

Deletion of FoxO1, 3, and 4 in Osteoblast Progenitors Attenuates the Loss of Cancellous Bone Mass in a Mouse Model of Type 1 Diabetes

PubMed Central

Iyer, Srividhya; Han, Li; Ambrogini, Elena; Yavropoulou, Maria; Fowlkes, John; Manolagas, Stavros C; Almeida, Maria

2017-01-01

Type 1 diabetes is associated with osteopenia and increased fragility fractures, attributed to reduced bone formation. However, the molecular mechanisms mediating these effects remain unknown. Insulin promotes osteoblast formation and inhibits the activity of the FoxO transcription factors. FoxOs, on the other hand, inhibit osteoprogenitor proliferation and bone formation. Here, we investigated whether FoxOs play a role in the low bone mass associated with type 1 diabetes, using mice lacking FoxO1, 3, and 4 in osteoprogenitor cells (FoxO1,3,4ΔOsx1-Cre). Streptozotocin-induced diabetes caused a reduction in bone mass and strength in FoxO-intact mice. In contrast, cancellous bone was unaffected in diabetic FoxO1,3,4ΔOsx1-Cre mice. The low bone mass in the FoxO-intact diabetic mice was associated with decreased osteoblast number and bone formation, as well as decreased expression of the anti-osteoclastogenic cytokine osteoprotegerin (OPG) and increased osteoclast number. FoxO deficiency did not alter the effects of diabetes on bone formation; however, it did prevent the decrease in OPG and the increase in osteoclast number. Addition of high glucose to osteoblastic cell cultures decreased OPG mRNA, indicating that hyperglycemia in and of itself contributes to diabetic bone loss. Taken together, these results suggest that FoxOs exacerbate the loss of cancellous bone mass associated with type 1 diabetes and that inactivation of FoxOs might ameliorate the adverse effects of insulin deficiency. PMID:27491024

Hepatic IGF-1R overexpression combined with the activation of GSK-3β and FOXO3a in the development of liver cirrhosis.

PubMed

Liu, Wentao; Li, Jing; Cai, Yan; Wu, Qiong; Pan, Yue; Chen, Yang; Chen, Yujing; Zheng, Xiao; Li, Wei; Zhang, Xuewen; E, Changyong

2016-02-15

Liver cirrhosis is the common pathological histology manifest among a number of chronic liver diseases and liver cancer. Circulating levels of insulin growth factor-1 (IGF-1) have been recently linked to liver cirrhosis and the development of liver cancer. Herein, we hypothesized that IGF-1R overexpression combining the activation of GSK-3β and FOXO3a were involved in the development of human and murine chronic liver cirrhosis. Liver samples of patients were screened from the Tissue Bank of the China-Japan Union Hospital of Jilin University. Mice liver fibrosis model was performed using intraperitoneal injection of carbon tetrachloride (CCl4) for 12weeks. Serum IGF-1 levels were detected by enzyme-linked immunosorbent assays (ELISA). Microscopical examination of liver parenchyma was performed using conventional H&E and Masson's staining. Moreover, we investigated the IGF-1 receptor (IGF-1R) signaling pathway at different period after CCl4 administration. Serum IGF-1 levels were significantly decreased in patients with liver cirrhosis, which is concomitant with the declined circulating levels of IGF-1 in 8 to 12weeks CCl4-treated mice. Furthermore, the expression of IGF-1R was significantly higher at 12w compared with control group. In addition, activation of the GSK-3β and FOXO3a were activated during the process of murine chronic liver injury. The present study demonstrates that decreased circulating IGF-1 levels are involved in human and murine chronic liver disease. Interestingly, overexpression of the IGF-1R, and activation of GSK3β and FOXO3a might be the molecular mechanisms underlying the development of liver cirrhosis. Copyright © 2016 Elsevier Inc. All rights reserved.

De-novo NAD+ synthesis regulates SIRT1-FOXO1 apoptotic pathway in response to NQO1 substrates in lung cancer cells

PubMed Central

Cheng, Xuefang; Li, Qingran; Liu, Fang; Ye, Hui; Zhao, Min; Wang, Hong; Wang, Guangji; Hao, Haiping

2016-01-01

Tryptophan metabolism is essential in diverse kinds of tumors via regulating tumor immunology. However, the direct role of tryptophan metabolism and its signaling pathway in cancer cells remain largely elusive. Here, we establish a mechanistic link from L-type amino acid transporter 1 (LAT1) mediated transport of tryptophan and the subsequent de-novo NAD+ synthesis to SIRT1-FOXO1 regulated apoptotic signaling in A549 cells in response to NQO1 activation. In response to NQO1 activation, SIRT1 is repressed leading to the increased cellular accumulation of acetylated FOXO1 that transcriptionally activates apoptotic signaling. Decreased uptake of tryptophan due to the downregulation of LAT1 coordinates with PARP-1 hyperactivation to induce rapid depletion of NAD+ pool. Particularly, the LAT1-NAD+-SIRT1 signaling is activated in tumor tissues of patients with non-small cell lung cancer. Because NQO1 activation is characterized with oxidative challenge induced DNA damage, these results suggest that LAT1 and de-novo NAD+ synthesis in NSCLC cells may play essential roles in sensing excessive oxidative stress. PMID:27566573

De-novo NAD+ synthesis regulates SIRT1-FOXO1 apoptotic pathway in response to NQO1 substrates in lung cancer cells.

PubMed

Liu, Huiying; Xing, Rong; Cheng, Xuefang; Li, Qingran; Liu, Fang; Ye, Hui; Zhao, Min; Wang, Hong; Wang, Guangji; Hao, Haiping

2016-09-20

Tryptophan metabolism is essential in diverse kinds of tumors via regulating tumor immunology. However, the direct role of tryptophan metabolism and its signaling pathway in cancer cells remain largely elusive. Here, we establish a mechanistic link from L-type amino acid transporter 1 (LAT1) mediated transport of tryptophan and the subsequent de-novo NAD+ synthesis to SIRT1-FOXO1 regulated apoptotic signaling in A549 cells in response to NQO1 activation. In response to NQO1 activation, SIRT1 is repressed leading to the increased cellular accumulation of acetylated FOXO1 that transcriptionally activates apoptotic signaling. Decreased uptake of tryptophan due to the downregulation of LAT1 coordinates with PARP-1 hyperactivation to induce rapid depletion of NAD+ pool. Particularly, the LAT1-NAD+-SIRT1 signaling is activated in tumor tissues of patients with non-small cell lung cancer. Because NQO1 activation is characterized with oxidative challenge induced DNA damage, these results suggest that LAT1 and de-novo NAD+ synthesis in NSCLC cells may play essential roles in sensing excessive oxidative stress.

FoxO Transcription Factors and Regenerative Pathways in Diabetes Mellitus

PubMed Central

Maiese, Kenneth

2015-01-01

Mammalian forkhead transcription factors of the O class (FoxO) are exciting targets under consideration for the development of new clinical entities to treat metabolic disorders and diabetes mellitus (DM). DM, a disorder that currently affects greater than 350 million individuals globally, can become a devastating disease that leads to cellular injury through oxidative stress pathways and affects multiple systems of the body. FoxO proteins can regulate insulin signaling, gluconeogenesis, insulin resistance, immune cell migration, and cell senescence. FoxO proteins also control cell fate through oxidative stress and pathways of autophagy and apoptosis that either lead to tissue regeneration or cell demise. Furthermore, FoxO signaling can be dependent upon signal transduction pathways that include silent mating type information regulation 2 homolog 1 (S. cerevisiae) (SIRT1), Wnt, and Wnt1 inducible signaling pathway protein 1 (WISP1). Cellular metabolic pathways driven by FoxO proteins are complex, can lead to variable clinical outcomes, and require in-depth analysis of the epigenetic and post-translation protein modifications that drive FoxO protein activation and degradation. PMID:26256004

Dual effects of fructose on ChREBP and FoxO1/3α are responsible for AldoB up-regulation and vascular remodelling.

PubMed

Cao, Wei; Chang, Tuanjie; Li, Xiao-Qiang; Wang, Rui; Wu, Lingyun

2017-02-01

Increased production of methylglyoxal (MG) in vascular tissues is one of the causative factors for vascular remodelling in different subtypes of metabolic syndrome, including hypertension and insulin resistance. Fructose-induced up-regulation of aldolase B (AldoB) contributes to increased vascular MG production but the underlying mechanisms are unclear. Serum levels of MG and fructose were determined in diabetic patients with hypertension. MG level had significant positive correlations with blood pressure and fructose level respectively. C57BL/6 mice were fed with control or fructose-enriched diet for 3 months and ultrasonographic and histologic analyses were performed to evaluate arterial structural changes. Fructose-fed mice exhibited hypertension and high levels of serum MG with normal glucose level. Fructose intake increased blood vessel wall thickness and vascular smooth muscle cell (VSMC) proliferation. Western blotting and real-time PCR analysis revealed that AldoB level was significantly increased in both the aorta of fructose-fed mice and the fructose-treated VSMCs, whereas aldolase A (AldoA) expression was not changed. The knockdown of AldoB expression prevented fructose-induced MG overproduction and VSMC proliferation. Moreover, fructose significantly increased carbohydrate-responsive element-binding protein (ChREBP), phosphorylated FoxO1/3α and Akt1 levels. Fructose induced translocation of ChREBP from the cytosol to nucleus and activated AldoB gene expression, which was inhibited by the knockdown of ChREBP. Meanwhile, fructose caused FoxO1/3α shuttling from the nucleus to cytosol and inhibited its binding to AldoB promoter region. Fructose-induced AldoB up-regulation was suppressed by Akt1 inhibitor but enhanced by FoxO1/3α siRNA. Collectively, fructose activates ChREBP and inactivates FoxO1/3α pathways to up-regulate AldoB expression and MG production, leading to vascular remodelling. © 2017 The Author(s). published by Portland Press Limited on

Constitutive phosphorylation of the FOXO1A transcription factor as a prognostic variable in gastric cancer.

PubMed

Kim, Ji Hun; Kim, Min Kyu; Lee, Hee Eun; Cho, Sung Jin; Cho, Yu Jin; Lee, Byung Lan; Lee, Hye Seung; Nam, Seon Young; Lee, Jae-Seon; Kim, Woo Ho

2007-08-01

Increased phosphorylation of FOXO1A, a FOXO transcription factor, has been implicated in several human cancers; however, it has not been studied in the gastric cancer to date. To determine the status of pFOXO1A expression in human gastric cancers and to determine its relationship with other tumor-associated proteins, we performed immunohistochemical staining on tissue array slides containing 272 human gastric carcinoma specimens. In non-neoplastic gastric mucosa, the expression of pFOXO1A was observed primarily in cells in the proliferative zone and in areas of intestinal metaplasia. In gastric carcinomas, the expression of pFOXO1A was observed in 230 (84.6%) out of 272 cases examined, and was positively correlated with the Ki-67-labeling index (P=0.026). The expression of pFOXO1A was higher in the early stages of pTNM (P<0.001), and was inversely correlated with the intestinal type by Lauren's classification (P=0.001), lymphatic invasion (P=0.017) and lymph node metastasis (P<0.001). Moreover, the expression of pFOXO1A was correlated with a longer patient survival (P=0.004). In addition, the expression of pFOXO1A was correlated with that of pAKT1 (P<0.001), PTEN (P=0.009), CDKN2A (P=0.012), APC (P=0.048), SMAD4 (P<0.001), CD82 (P=0.011), and BCL2 (P=0.011). In conclusion, our results showed that the expression of pFOXO1A is a frequent and early event in gastric tumorigenesis and that there is a significant correlation between pFOXO1A and better prognosis. Thus, our data suggest that the expression of pFOXO1A may serve as a valuable prognostic variable in gastric carcinoma and have significant implications for the development and application of targeted therapy.

Cytokine-mediated FOXO3a phosphorylation suppresses FasL expression in hemopoietic cell lines: investigations of the role of Fas in apoptosis due to cytokine starvation.

PubMed

Behzad, Hayedeh; Jamil, Sarwat; Denny, Trisha A; Duronio, Vincent

2007-05-01

We have investigated phosphatidylinositol 3-kinase (PI3K)-dependent survival signalling pathways using several cytokines in three different hemopoietic cell lines, MC/9, FDC-P1, and TF-1. Cytokines caused PI3K- and PKB-dependent phosphorylation of FOXO3a (previously known as FKHRL1) at three distinct sites. Following cytokine withdrawal or PI3K inhibition, both of which are known to lead to apoptosis, there was a loss of FOXO3a phosphorylation, and a resulting increase in forkhead transcriptional activity, along with increased expression of Fas Ligand (FasL), which could be detected at the cell surface. Concurrently, an increase in cell surface expression of Fas was also detected. Despite the presence of both FasL and Fas, there was no detectable evidence that activation of Fas-mediated apoptotic events was contributing to apoptosis resulting from cytokine starvation or inhibition of PI3K activity. Thus, inhibition of FOXO3a activity is mediated by the PI3K-PKB pathway, but regulation of FasL is not the primary means by which cell survival is regulated in cytokine-dependent hemopoietic cells. We were also able to confirm increased expression of known FOXO3a targets, Bim and p27kip1. Together, these results support the conclusion that mitochondrial-mediated signals play the major role in apoptosis of hemopoietic cells due to loss of cytokine signalling.

Salsalate and Adiponectin Improve Palmitate-Induced Insulin Resistance via Inhibition of Selenoprotein P through the AMPK-FOXO1α Pathway.

PubMed

Jung, Tae Woo; Choi, Hae Yoon; Lee, So Young; Hong, Ho Cheol; Yang, Sae Jeong; Yoo, Hye Jin; Youn, Byung-Soo; Baik, Sei Hyun; Choi, Kyung Mook

2013-01-01

Selenoprotein P (SeP) was recently identified as a hepatokine that induces insulin resistance (IR) in rodents and humans. Recent clinical trials have shown that salsalate, a prodrug of salicylate, significantly lowers blood glucose levels and increases adiponectin concentrations. We examined the effects of salsalate and full length-adiponectin (fAd) on the expression of SeP under hyperlipidemic conditions and explored their regulatory mechanism on SeP. In palmitate-treated HepG2 cells as well as high fat diet (HFD)-fed male Spraque Dawley (SD) rats and male db/db mice, SeP expression and its regulatory pathway, including AMPK-FOXO1α, were evaluated after administration of salsalate and salicylate. Palmitate treatment significantly increased SeP expression and aggravated IR, while knock-down of SeP by siRNA restored these changes in HepG2 cells. Palmitate-induced SeP expression was inhibited by both salsalate and salicylate, which was mediated by AMPK activation, and was blocked by AMPK siRNA or an inhibitor of AMPK. Chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift (EMSA) assay showed that salsalate suppressed SeP expression by AMPK-mediated phosphorylation of FOXO1α. Moreover, fAd also reduced palmitate-induced SeP expression through the activation of AMPK, which results in improved IR. Both salsalate and salicylate treatment significantly improved glucose intolerance and insulin sensitivity, accompanied by reduced SeP mRNA and protein expression in HFD-fed rats and db/db mice, respectively. Taken together, we found that salsalate and adiponectin ameliorated palmitate-induced IR in hepatocytes via SeP inhibition through the AMPK-FOXO1α pathway. The regulation of SeP might be a novel mechanism mediating the anti-diabetic effects of salsalate and adiponectin.

Unexpected role for dosage compensation in the control of dauer arrest, insulin-like signaling, and FoxO transcription factor activity in Caenorhabditis elegans.

PubMed

Dumas, Kathleen J; Delaney, Colin E; Flibotte, Stephane; Moerman, Donald G; Csankovszki, Gyorgyi; Hu, Patrick J

2013-07-01

During embryogenesis, an essential process known as dosage compensation is initiated to equalize gene expression from sex chromosomes. Although much is known about how dosage compensation is established, the consequences of modulating the stability of dosage compensation postembryonically are not known. Here we define a role for the Caenorhabditis elegans dosage compensation complex (DCC) in the regulation of DAF-2 insulin-like signaling. In a screen for dauer regulatory genes that control the activity of the FoxO transcription factor DAF-16, we isolated three mutant alleles of dpy-21, which encodes a conserved DCC component. Knockdown of multiple DCC components in hermaphrodite and male animals indicates that the dauer suppression phenotype of dpy-21 mutants is due to a defect in dosage compensation per se. In dpy-21 mutants, expression of several X-linked genes that promote dauer bypass is elevated, including four genes encoding components of the DAF-2 insulin-like pathway that antagonize DAF-16/FoxO activity. Accordingly, dpy-21 mutation reduced the expression of DAF-16/FoxO target genes by promoting the exclusion of DAF-16/FoxO from nuclei. Thus, dosage compensation enhances dauer arrest by repressing X-linked genes that promote reproductive development through the inhibition of DAF-16/FoxO nuclear translocation. This work is the first to establish a specific postembryonic function for dosage compensation in any organism. The influence of dosage compensation on dauer arrest, a larval developmental fate governed by the integration of multiple environmental inputs and signaling outputs, suggests that the dosage compensation machinery may respond to external cues by modulating signaling pathways through chromosome-wide regulation of gene expression.

FOXO Transcriptional Factors and Long-Term Living

PubMed Central

Rashid, Rehana; Muneer, Saiqa; Hasan, Syed Muhammad Farid

2017-01-01

Several pathologies such as neurodegeneration and cancer are associated with aging, which is affected by many genetic and environmental factors. Healthy aging conceives human longevity, possibly due to carrying the defensive genes. For instance, FOXO (forkhead box O) genes determine human longevity. FOXO transcription factors are involved in the regulation of longevity phenomenon via insulin and insulin-like growth factor signaling. Only one FOXO gene (FOXO DAF-16) exists in invertebrates, while four FOXO genes, that is, FOXO1, FOXO3, FOXO4, and FOXO6 are found in mammals. These four transcription factors are involved in the multiple cellular pathways, which regulate growth, stress resistance, metabolism, cellular differentiation, and apoptosis in mammals. However, the accurate mode of longevity by FOXO factors is unclear until now. This article describes briefly the existing knowledge that is related to the role of FOXO factors in human longevity. PMID:28894507

The Inconvenience of Convenience Cohorts: Rhabdomyosarcoma and the PAX-FOXO1 Biomarker

PubMed Central

Rosenberg, Abby R.; Skapek, Stephen X.; Hawkins, Douglas S.

2012-01-01

“Convenience cohorts” comprise individuals thought to represent the general population, but chosen because they are readily available for evaluation, rather than at random. As such, these methods are subject to bias and may be misleading. Convenience cohorts have been used to investigate the prognostic significance of chromosomal translocations between the PAX3 or PAX7 and the FOXO1 genes in rhabdomyosarcoma, the most common pediatric sarcoma. However, retrospective studies assessing the role of PAX-FOXO1 translocations have yielded inconsistent results. This review highlights the findings from several clinical correlation studies of the PAX-FOXO1 biomarker and illustrates the challenges of using such methods to draw clinical conclusions. PMID:22564868

PAX3/7-FOXO1 fusion status in older rhabdomyosarcoma patient population by fluorescent in situ hybridization.

PubMed

Dumont, Sarah N; Lazar, Alexander J; Bridge, Julia A; Benjamin, Robert S; Trent, Jonathan C

2012-02-01

In pediatric alveolar rhabdomyosarcoma, the PAX3-FOXO1 and PAX7-FOXO1 gene fusions are prognostic indicators, while little is known concerning this disease in older patients. To determine whether PAX3/7-FOXO1 fusion gene status correlates with outcome in adolescent, young adult, and adult rhabdomyosarcoma patients, the histological, immunohistochemical, and clinical characteristics of 105 patients followed at The University of Texas MD Anderson Cancer Center from 1957 to 2001 were evaluated. The samples were assembled into a tissue microarray, and fusion gene status was determined by fluorescence in situ hybridization using PAX3, PAX7, and FOXO1 loci-specific probes. The disease characteristics and specific gene fusion were correlated with patient outcomes using the log-rank test. Fifty-two percent of the samples exhibited a PAX3-FOXO1 fusion, 15% the PAX7-FOXO1 fusion, and 33% were negative for a rearrangement of these loci. The presence of PAX3/7-FOXO1 translocation was significantly associated with a higher frequency of metastatic disease. Although a statistically significant correlation between the PAX3/7-FOXO1 fusion gene status and overall survival was not identified, there was a trend toward better outcomes for patients with fusion-negative RMS. Therefore, identification of a FOXO1 fusion appears to be an interesting tool for predicting outcomes in older rhabdomyosarcoma patients and is worth further investigations in this rare subgroup of RMS population.

FOXO1 content is reduced in cystic fibrosis and increases with IGF-I treatment.

PubMed

Smerieri, Arianna; Montanini, Luisa; Maiuri, Luigi; Bernasconi, Sergio; Street, Maria E

2014-10-08

Cystic fibrosis-related diabetes is to date the most frequent complication in cystic fibrosis (CF). The mechanisms underlying this condition are not well understood, and a possible role of insulin resistance is debated. We investigated insulin signal transduction in CF. Total insulin receptor, IRS1, p85 PI3K, and AKT contents were substantially normal in CF cells (CFBE41o-), whereas winged helix forkhead (FOX)O1 contents were reduced both in baseline conditions and after insulin stimulation. In addition, CF cells showed increased ERK1/2, and reduced β2 arrestin contents. No significant change in SOCS2 was observed. By using a CFTR inhibitor and siRNA, changes in FOXO1 were related to CFTR loss of function. In a CF-affected mouse model, FOXO1 content was reduced in the muscle while no significant difference was observed in liver and adipose tissue compared with wild-type. Insulin-like growth factor 1 (IGF-I) increased FOXO1 content in vitro and in vivo in muscle and adipose tissue. In conclusion; we present the first description of reduced FOXO1 content in CF, which is compatible with reduced gluconeogenesis and increased adipogenesis, both features of insulin insensitivity. IGF-I treatment was effective in increasing FOXO1, thereby suggesting that it could be considered as a potential treatment in CF patients possibly to prevent and treat cystic fibrosis-related diabetes.

FOXO family in regulating cancer and metabolism.

PubMed

Ma, Jian; Matkar, Smita; He, Xin; Hua, Xianxin

2018-06-01

FOXO proteins are a sub-group of a superfamily of forkhead box (FOX)-containing transcription factors (TFs). FOXOs play an important role in regulating a plethora of biological activities ranging from development, cell signaling, and tumorigenesis to cell metabolism. Here we mainly focus on reviewing the role of FOXOs in regulating tumor and metabolism. Moreover, how crosstalk among various pathways influences the function of FOXOs will be reviewed. Further, the paradoxical role for FOXOs in controlling the fate of cancer and especially resistance/sensitivity of cancer to the class of drugs that target PI3K/AKT will also be reviewed. Finally, how FOXOs regulate crosstalk between common cancer pathways and cell metabolism pathways, and how these crosstalks affect the fate of the cancer will be discussed. Copyright © 2018. Published by Elsevier Ltd.

Carbocysteine counteracts the effects of cigarette smoke on cell growth and on the SIRT1/FoxO3 axis in bronchial epithelial cells.

PubMed

Pace, E; Di Vincenzo, S; Ferraro, M; Bruno, A; Dino, P; Bonsignore, M R; Battaglia, S; Saibene, F; Lanata, L; Gjomarkaj, M

2016-08-01

Cigarette smoke may accelerate cellular senescence by increasing oxidative stress. Altered proliferation and altered expression of anti-aging factors, including SIRT1 and FoxO3, characterise cellular senescence. The effects of carbocysteine on the SIRT1/FoxO3 axis and on downstream molecular mechanisms in human bronchial epithelial cells exposed to cigarette smoke are largely unknown. Aim of this study was to explore whether carbocysteine modulated SIRT1/FoxO3 axis, and downstream molecular mechanisms associated to cellular senescence, in a bronchial epithelial cell line (16-HBE) exposed to cigarette smoke. 16HBE cells were stimulated with/without cigarette smoke extracts (CSE) and carbocysteine. Flow cytometry and clonogenic assay were used to assess cell proliferation; western blot analysis was used for assessing nuclear expression of SIRT1 and FoxO3. The nuclear co-localization of SIRT1 and FoxO3 was assessed by fluorescence microscopy. Beta galactosidase (a senescence marker) and SIRT1 activity were assessed by specific staining and colorimetric assays, respectively. ChiP Assay and flow cytometry were used for assessing survivin gene regulation and protein expression, respectively. CSE decreased cell proliferation, the nuclear expression of SIRT1 and FoxO3 and increased beta galactosidase staining. CSE, reduced SIRT1 activity and FoxO3 localization on survivin promoter thus increasing survivin expression. In CSE stimulated bronchial epithelial cells carbocysteine reverted these phenomena by increasing cell proliferation, and SIRT1 and FoxO3 nuclear expression, and by reducing beta galactosidase staining and survivin expression. The study shows for the first time that carbocysteine may revert some senescence processes induced by oxidative stress due to cigarette smoke exposure. Copyright © 2016 Elsevier Inc. All rights reserved.

Autophagy induction by leptin contributes to suppression of apoptosis in cancer cells and xenograft model: Involvement of p53/FoxO3A axis

PubMed Central

Nepal, Saroj; Kim, Mi Jin; Hong, Jin Tae; Kim, Sang Hyun; Sohn, Dong-Hwan; Lee, Sung Hee; Song, Kyung; Choi, Dong Young; Lee, Eung Seok; Park, Pil-Hoon

2015-01-01

Leptin, a hormone mainly produced from adipose tissue, has been shown to induce proliferation of cancer cells. However, the molecular mechanisms underlying leptin-induced tumor progression have not been clearly elucidated. In the present study, we investigated the role of autophagy in leptin-induced cancer cell proliferation using human hepatoma (HepG2) and breast cancer cells (MCF-7), and tumor growth in a xenograft model. Herein, we showed that leptin treatment caused autophagy induction as assessed by increase in expression of autophagy-related genes, including beclin-1, Atg5 and LC3 II, further induction of autophagosome formation and autophagic flux. Interestingly, inhibition of autophagic process by treatment with inhibitors and LC3B gene silencing blocked leptin-induced increase in cell number and suppression of apoptosis, indicating a crucial role of autophagy in leptin-induced tumor progression. Moreover, gene silencing of p53 or FoxO3A prevented leptin-induced LC3 II protein expression, suggesting an involvement of p53/FoxO3A axis in leptin-induced autophagy activation. Leptin administration also accelerated tumor growth in BALB/c nude mice, which was found to be autophagy dependent. Taken together, our results demonstrate that leptin-induced tumor growth is mediated by autophagy induction and autophagic process would be a promising target to regulate development of cancer caused by leptin production. PMID:25704884

Maternal diabetes and high glucose in vitro trigger Sca1+ cardiac progenitor cell apoptosis through FoxO3a.

PubMed

Yang, Penghua; Yang, Wendy W; Chen, Xi; Kaushal, Sunjay; Dong, Daoyin; Shen, Wei-Bin

2017-01-22

Recent controversies surrounding the authenticity of c-kit + cardiac progenitor cells significantly push back the advance in regenerative therapies for cardiovascular diseases. There is an urgent need for research in characterizing alternative types of cardiac progenitor cells. Towards this goal, in the present study, we determined the effect of maternal diabetes on Sca1 + cardiac progenitor cells. Maternal diabetes induced caspase 3-dependent apoptosis in Sca1 + cardiac progenitor cells derived from embryonic day 17.5 (E17.5). Similarly, high glucose in vitro but not the glucose osmotic control mannitol triggered Sca1 + cardiac progenitor cell apoptosis in a dose- and time-dependent manner. Both maternal diabetes and high glucose in vitro activated the pro-apoptotic transcription factor, Forkhead O 3a (FoxO3a) via dephosphorylation at threonine 32 (Thr-32) residue. foxo3a gene deletion abolished maternal diabetes-induced Sca1 + cardiac progenitor cell apoptosis. The dominant negative FoxO3a mutant without the transactivation domain from the C terminus blocked high glucose-induced Sca1 + cardiac progenitor cell apoptosis, whereas the constitutively active FoxO3a mutant with the three phosphorylation sites, Thr-32, Ser-253, and Ser-315, being replaced by alanine residues mimicked the pro-apoptotic effect of high glucose. Thus, maternal diabetes and high glucose in vitro may limit the regenerative potential of Sca1 + cardiac progenitor cells by inducing apoptosis through FoxO3a activation. These findings will serve as the guide in optimizing the autologous therapy using Sca1 + cardiac progenitor cells in cardiac defect babies born exposed to maternal diabetes. Copyright © 2016. Published by Elsevier Inc.

Associations between Forkhead Box O1 (FoxO1) Expression and Indicators of Hepatic Glucose Production in Transition Dairy Cows Supplemented with Dietary Nicotinic Acid.

PubMed

Kinoshita, Asako; Locher, Lena; Tienken, Reka; Meyer, Ulrich; Dänicke, Sven; Rehage, Jürgen; Huber, Korinna

2016-01-01

Forkhead box protein O1 (FoxO1) is a transcription factor which promotes hepatic glucose production (HGP) by up-regulating the transcription of gluconeogenic enzymes in monogastric species. The activity of FoxO1 is inhibited by insulin-induced phosphorylation. The aims of the present study were to find associations between FoxO1 expression and variables associated with HGP as affected by feeding regimen in dairy cows during the transition period. Twenty one healthy German Holstein cows were allocated to four groups (LC-CON, HC-CON, LC-NA with 5 cows/group and HC-NA with 6 cows/group, respectively). Cows received 0 (LC-CON and HC-CON) or 24 (LC-NA and HC-NA) g/d nicotinic acid with high (HC) or low (LC) concentrate proportion from -42 days (-41.8 + 4.8; mean + standard deviation) relative to expected calving date (d-42) to d24. Liver biopsy was taken at d-42, 1, 21, and 100. The total protein expression of FoxO1 (tFoxO1) and the extent of phosphorylation of FoxO1 at serine 256 (pFoxO1) were analysed semiquantitatively by Western Blotting. The expression of hepatic mRNA of FoxO1 and seven genes associated with HGP was measured by real-time RT-PCR. Mixed model and Pearson's correlation were used for statistical evaluation with the level of significance at P<0.05. No dietary effect was observed either on feed intake, energy balance, or on the concentration of blood metabolites. Neither time nor diet affected the expression of FoxO1 total protein and mRNA. A NA × concentrate interaction was found in pFoxO1. However, no corresponding dietary effect was found in the mRNA expression of investigated genes. Different patterns of correlations between FoxO1-related variables and investigated indicators for HGP were found at d21 and 100. The results indicated that the regulation of HGP did not take place on the levels of mRNA and protein expression and the phosphorylation of FoxO1 in dairy cows in early lactation.

Associations between Forkhead Box O1 (FoxO1) Expression and Indicators of Hepatic Glucose Production in Transition Dairy Cows Supplemented with Dietary Nicotinic Acid

PubMed Central

Kinoshita, Asako; Locher, Lena; Tienken, Reka; Meyer, Ulrich; Dänicke, Sven; Rehage, Jürgen; Huber, Korinna

2016-01-01

Forkhead box protein O1 (FoxO1) is a transcription factor which promotes hepatic glucose production (HGP) by up-regulating the transcription of gluconeogenic enzymes in monogastric species. The activity of FoxO1 is inhibited by insulin-induced phosphorylation. The aims of the present study were to find associations between FoxO1 expression and variables associated with HGP as affected by feeding regimen in dairy cows during the transition period. Twenty one healthy German Holstein cows were allocated to four groups (LC-CON, HC-CON, LC-NA with 5 cows/group and HC-NA with 6 cows/group, respectively). Cows received 0 (LC-CON and HC-CON) or 24 (LC-NA and HC-NA) g/d nicotinic acid with high (HC) or low (LC) concentrate proportion from -42 days (-41.8 + 4.8; mean + standard deviation) relative to expected calving date (d-42) to d24. Liver biopsy was taken at d-42, 1, 21, and 100. The total protein expression of FoxO1 (tFoxO1) and the extent of phosphorylation of FoxO1 at serine 256 (pFoxO1) were analysed semiquantitatively by Western Blotting. The expression of hepatic mRNA of FoxO1 and seven genes associated with HGP was measured by real-time RT-PCR. Mixed model and Pearson’s correlation were used for statistical evaluation with the level of significance at P<0.05. No dietary effect was observed either on feed intake, energy balance, or on the concentration of blood metabolites. Neither time nor diet affected the expression of FoxO1 total protein and mRNA. A NA × concentrate interaction was found in pFoxO1. However, no corresponding dietary effect was found in the mRNA expression of investigated genes. Different patterns of correlations between FoxO1-related variables and investigated indicators for HGP were found at d21 and 100. The results indicated that the regulation of HGP did not take place on the levels of mRNA and protein expression and the phosphorylation of FoxO1 in dairy cows in early lactation. PMID:26800252

Activation of MAPK and FoxO by Manganese (Mn) in Rat Neonatal Primary Astrocyte Cultures

PubMed Central

Exil, Vernat; Ping, Li; Yu, Yingchun; Chakraborty, Sudipta; Caito, Samuel W.; Wells, K. Sam; Karki, Pratap; Lee, Eunsook; Aschner, Michael

2014-01-01

Environmental exposure to manganese (Mn) leads to a neurodegenerative disease that has shared clinical characteristics with Parkinson's disease (PD). Mn-induced neurotoxicity is time- and dose-dependent, due in part to oxidative stress. We ascertained the molecular targets involved in Mn-induced neurodegeneration using astrocyte culture as: (1) Astrocytes are vital for information processing within the brain, (2) their redox potential is essential in mitigating reactive oxygen species (ROS) levels, and (3) they are targeted early in the course of Mn toxicity. We first tested protein levels of Mn superoxide dismutase -2 (SOD-2) and glutathione peroxidase (GPx-1) as surrogates of astrocytic oxidative stress response. We assessed levels of the forkhead winged-helix transcription factor O (FoxO) in response to Mn exposure. FoxO is highly regulated by the insulin-signaling pathway. FoxO mediates cellular responses to toxic stress and modulates adaptive responses. We hypothesized that FoxO is fundamental in mediating oxidative stress response upon Mn treatment, and may be a biomarker of Mn-induced neurodegeneration. Our results indicate that 100 or 500 µM of MnCl2 led to increased levels of FoxO (dephosphorylated and phosphorylated) compared with control cells (P<0.01). p-FoxO disappeared from the cytosol upon Mn exposure. Pre-treatment of cultured cells with (R)-(−)-2-oxothiazolidine-4-carboxylic acid (OTC), a cysteine analog rescued the cytosolic FoxO. At these concentrations, MAPK phosphorylation, in particular p38 and ERK, and PPAR gamma coactivator-1 (PGC-1) levels were increased, while AKT phosphorylation remained unchanged. FoxO phosphorylation level was markedly reduced with the use of SB203580 (a p38 MAPK inhibitor) and PD98059 (an ERK inhibitor). We conclude that FoxO phosphorylation after Mn exposure occurs in parallel with, and independent of the insulin-signaling pathway. FoxO levels and its translocation into the nucleus are part of early events

FoxO1 regulates apoptosis induced by asbestos in the MT-2 human T-cell line.

PubMed

Matsuzaki, Hidenori; Lee, Suni; Maeda, Megumi; Kumagai-Takei, Naoko; Nishimura, Yasumitsu; Otsuki, Takemi

2016-09-01

Asbestos is known to cause malignant mesothelioma and lung cancer. Recent studies implicate tumor immunity in the development of various tumors, including malignant mesothelioma. In order to establish an in vitro T-cell model to clarify the effects of long-term exposure of asbestos on tumor immunity, in this study, human T-cell line MT-2 cells were cultured with asbestos for longer than 8 months and the resultant cells (MT-2Rst) were assessed for the expression of forkhead transcription factor FoxO1. Gene expression analysis revealed that the amount of FoxO1 mRNA decreased after long-term exposure of the MT-2 cells to asbestos. In accordance with this reduction in FoxO1, pro-apoptotic Foxo1 target genes Puma, Fas ligand and Bim were also seen to be down-regulated in MT-2Rst cells. Furthermore, shRNA-mediated knock-down of FoxO1 reduced the number of apoptotic parental MT-2 cells after treatment with asbestos. On the other hand, over-expression of FoxO1 did not affect asbestos-induced apoptosis in MT-2Rst cells. These results suggested that FoxO1 played an important role in regulating asbestos-induced apoptosis and confirmed the presence of multiple pathways regulating resistance to asbestos in MT-2Rst cells.

PAX3-FOXO1 transgenic zebrafish models identify HES3 as a mediator of rhabdomyosarcoma tumorigenesis.

PubMed

Kendall, Genevieve C; Watson, Sarah; Xu, Lin; LaVigne, Collette A; Murchison, Whitney; Rakheja, Dinesh; Skapek, Stephen X; Tirode, Franck; Delattre, Olivier; Amatruda, James F

2018-06-05

Alveolar rhabdomyosarcoma is a pediatric soft-tissue sarcoma caused by PAX3/7-FOXO1 fusion oncogenes and is characterized by impaired skeletal muscle development. We developed human PAX3-FOXO1 -driven zebrafish models of tumorigenesis and found that PAX3-FOXO1 exhibits discrete cell lineage susceptibility and transformation. Tumors developed by 1.6-19 months and were primitive neuroectodermal tumors or rhabdomyosarcoma. We applied this PAX3-FOXO1 transgenic zebrafish model to study how PAX3-FOXO1 leverages early developmental pathways for oncogenesis and found that her3 is a unique target. Ectopic expression of the her3 human ortholog, HES3 , inhibits myogenesis in zebrafish and mammalian cells, recapitulating the arrested muscle development characteristic of rhabdomyosarcoma. In patients, HES3 is overexpressed in fusion-positive versus fusion-negative tumors. Finally, HES3 overexpression is associated with reduced survival in patients in the context of the fusion. Our novel zebrafish rhabdomyosarcoma model identifies a new PAX3-FOXO1 target, her3 / HES3 , that contributes to impaired myogenic differentiation and has prognostic significance in human disease. © 2018, Kendall et al.

MicroRNA-214 Suppresses Gluconeogenesis by Targeting Activating Transcriptional Factor 4*

PubMed Central

Li, Kai; Zhang, Jin; Yu, Junjie; Liu, Bin; Guo, Yajie; Deng, Jiali; Chen, Shanghai; Wang, Chunxia; Guo, Feifan

2015-01-01

Although the gluconeogenesis pathway is already a target for the treatment of type 2 diabetes, the potential role of microRNAs (miRNAs) in gluconeogenesis remains unclear. Here, we investigated the physiological functions of miR-214 in gluconeogenesis. The expression of miR-214 was suppressed by glucagon via protein kinase A signaling in primary hepatocytes, and miR-214 was down-regulated in the livers of fasted, high fat diet-induced diabetic and leptin receptor-mutated (db/db) mice. The overexpression of miR-214 in primary hepatocytes suppressed glucose production, and silencing miR-214 reversed this effect. Gluconeogenesis was suppressed in the livers of mice injected with an adenovirus expressing miR-214 (Ad-miR-214). Additionally, Ad-miR-214 alleviated high fat diet-induced elevation of gluconeogenesis and hyperglycemia. Furthermore, we found that activating transcription factor 4 (ATF4), a reported target of miR-214, can reverse the suppressive effect of miR-214 on gluconeogenesis in primary hepatocytes, and this suppressive effect was blocked in liver-specific ATF4 knock-out mice. ATF4 regulated gluconeogenesis via affecting forkhead box protein O1 (FOXO1) transcriptional activity. Finally, liver-specific miR-214 transgenic mice exhibited suppressed gluconeogenesis and reduced expression of ATF4, phosphoenolpyruvate carboxykinase, and glucose-6-phosphatase in liver. Taken together, our results suggest that the miR-214-ATF4 axis is a novel pathway for the regulation of hepatic gluconeogenesis. PMID:25657009

Resveratrol Ameliorates Contrast Induced Nephropathy Through the Activation of SIRT1-PGC-1α-Foxo1 Signaling in Mice.

PubMed

Hong, Yu Ah; Bae, So Yeon; Ahn, Shin Young; Kim, Jieun; Kwon, Young Joo; Jung, Woon Yong; Ko, Gang Jee

2017-01-01

SIRT1 activation promotes the resistance of renal tubular cells to oxidative stress, and resveratrol is known as a SIRT1 activator. Resveratrol was injected intraperitoneally with iohexol for 24 hours. NRK-52E cells were pretreated with resveratrol for 24 hours and then exposed to iohexol for 3 hours. Renal function was measured by serum creatinine and cell survival was assessed by MTT assay. We investigated whether resveratrol attenuates oxidative stress and apoptosis in contrast-induced nephropathy (CIN). Serum creatinine and tubular injury increased significantly after iohexol treatment, and resveratrol co-treatment attenuated the renal injury. Cell survival decreased after iohexol exposure and resveratrol reduced cell death induced by iohexol. Resveratrol was accompanied with the activation of SIRT1 and PGC-1α and dephosphorylation of FoxO1 in mice with CIN. SIRT1 and PGC-1α expression were decreased by iohexol, and increased significantly in resveratrol-pretreated cells. These processes resulted in reduction of oxidative stress and apoptosis both in vivo and in vitro experiments. Resveratrol decreased inflammatory cell infiltration induced by iohexol in mice with CIN. SIRT1 inhibition using siRNA in tubular cells accentuated the decrease of cell viability by iohexol. Resveratrol attenuated CIN by modulating renal oxidative stress and apoptosis through activation of SIRT1-PGC-1α-FoxO1 signaling. The Author(s). Published by S. Karger AG, Basel.

Saponarin activates AMPK in a calcium-dependent manner and suppresses gluconeogenesis and increases glucose uptake via phosphorylation of CRTC2 and HDAC5.

PubMed

Seo, Woo-Duck; Lee, Ji Hae; Jia, Yaoyao; Wu, Chunyan; Lee, Sung-Joon

2015-11-15

This study investigated the molecular mechanism of saponarin, a flavone glucoside, in the regulation of insulin sensitivity. Saponarin suppressed the rate of gluconeogenesis and increased cellular glucose uptake in HepG2 and TE671 cells by regulating AMPK. Using an in vitro kinase assay, we showed that saponarin did not directly interact with the AMPK protein. Instead, saponarin increased intracellular calcium levels and induced AMPK phosphorylation, which was diminished by co-stimulation with STO-609, an inhibitor of CAMKKβ. Transcription of hepatic gluconeogenesis genes was upregulated by nuclear translocation of CRTC2 and HDAC5, coactivators of CREB and FoxO1 transcription factors, respectively. This nuclear translocation was inhibited by increased phosphorylation of CRTC2 and HDAC5 by saponarin-induced AMPK in HepG2 cells and suppression of CREB and FoxO1 transactivation activities in cells stimulated by saponarin. The results from a chromatin immunoprecipitation assay confirmed the reduced binding of CRTC2 on the PEPCK and G6Pase promoters. In TE671 cells, AMPK phosphorylated HDAC5, which suppressed nuclear penetration and upregulated GLUT4 transcription, leading to enhanced glucose uptake. Collectively, these results suggest that saponarin activates AMPK in a calcium-dependent manner, thus regulating gluconeogenesis and glucose uptake. Copyright © 2015 Elsevier Ltd. All rights reserved.

Hepatic NPC1L1 overexpression ameliorates glucose metabolism in diabetic mice via suppression of gluconeogenesis.

PubMed

Kurano, Makoto; Hara, Masumi; Satoh, Hiroaki; Tsukamoto, Kazuhisa

2015-05-01

Inhibition of intestinal NPC1L1 by ezetimibe has been demonstrated to improve glucose metabolism in rodent models; however, the role of hepatic NPC1L1 in glucose metabolism has not been elucidated. In this study, we analyzed the effects of hepatic NPC1L1 on glucose metabolism. We overexpressed NPC1L1 in the livers of lean wild type mice, diet-induced obesity mice and db/db mice with adenoviral gene transfer. We found that in all three mouse models, hepatic NPC1L1 overexpression lowered fasting blood glucose levels as well as blood glucose levels on ad libitum; in db/db mice, hepatic NPC1L1 overexpression improved blood glucose levels to almost the same as those found in lean wild type mice. A pyruvate tolerance test revealed that gluconeogenesis was suppressed by hepatic NPC1L1 overexpression. Further analyses revealed that hepatic NPC1L1 overexpression decreased the expression of FoxO1, resulting in the reduced expression of G6Pase and PEPCK, key enzymes in gluconeogenesis. These results indicate that hepatic NPC1L1 might have distinct properties of suppressing gluconeogenesis via inhibition of FoxO1 pathways. Copyright © 2015 Elsevier Inc. All rights reserved.

PAX3/7–FOXO1 fusion status in older rhabdomyosarcoma patient population by fluorescent in situ hybridization

PubMed Central

Lazar, Alexander J.; Bridge, Julia A.; Benjamin, Robert S.; Trent, Jonathan C.

2014-01-01

Purpose In pediatric alveolar rhabdomyosarcoma, the PAX3–FOXO1 and PAX7–FOXO1 gene fusions are prognostic indicators, while little is known concerning this disease in older patients. To determine whether PAX3/7–FOXO1 fusion gene status correlates with outcome in adolescent, young adult, and adult rhabdomyosarcoma patients, the histological, immunohistochemical, and clinical characteristics of 105 patients followed at The University of Texas MD Anderson Cancer Center from 1957 to 2001 were evaluated. Methods The samples were assembled into a tissue microarray, and fusion gene status was determined by fluorescence in situ hybridization using PAX3, PAX7, and FOXO1 loci-specific probes. The disease characteristics and specific gene fusion were correlated with patient outcomes using the log-rank test. Results Fifty-two percent of the samples exhibited a PAX3–FOXO1 fusion, 15% the PAX7–FOXO1 fusion, and 33% were negative for a rearrangement of these loci. The presence of PAX3/7–FOXO1 translocation was significantly associated with a higher frequency of metastatic disease. Although a statistically significant correlation between the PAX3/7–FOXO1 fusion gene status and overall survival was not identified, there was a trend toward better outcomes for patients with fusion-negative RMS. Conclusions Therefore, identification of a FOXO1 fusion appears to be an interesting tool for predicting outcomes in older rhabdomyosarcoma patients and is worth further investigations in this rare subgroup of RMS population. PMID:22089931

Cutting Edge: Differential Regulation of PTEN by TCR, Akt, and FoxO1 Controls CD4+ T Cell Fate Decisions.

PubMed

Hawse, William F; Sheehan, Robert P; Miskov-Zivanov, Natasa; Menk, Ashley V; Kane, Lawrence P; Faeder, James R; Morel, Penelope A

2015-05-15

Signaling via the Akt/mammalian target of rapamycin pathway influences CD4(+) T cell differentiation; low levels favor regulatory T cell induction and high levels favor Th induction. Although the lipid phosphatase phosphatase and tensin homolog (PTEN) suppresses Akt activity, the control of PTEN activity is poorly studied in T cells. In this study, we identify multiple mechanisms that regulate PTEN expression. During Th induction, PTEN function is suppressed via lower mRNA levels, lower protein levels, and an increase in C-terminal phosphorylation. Conversely, during regulatory T cell induction, PTEN function is maintained through the stabilization of PTEN mRNA transcription and sustained protein levels. We demonstrate that differential Akt/mammalian target of rapamycin signaling regulates PTEN transcription via the FoxO1 transcription factor. A mathematical model that includes multiple modes of PTEN regulation recapitulates our experimental findings and demonstrates how several feedback loops determine differentiation outcomes. Collectively, this work provides novel mechanistic insights into how differential regulation of PTEN controls alternate CD4(+) T cell fate outcomes. Copyright © 2015 by The American Association of Immunologists, Inc.

FOXO1 promotes wound healing through the up-regulation of TGF-β1 and prevention of oxidative stress

PubMed Central

Ponugoti, Bhaskar; Xu, Fanxing; Zhang, Chenying; Tian, Chen; Pacios, Sandra

2013-01-01

Keratinocyte mobilization is a critical aspect of wound re-epithelialization, but the mechanisms that control its precise regulation remain poorly understood. We set out to test the hypothesis that forkhead box O1 (FOXO1) has a negative effect on healing because of its capacity to inhibit proliferation and promote apoptosis. Contrary to expectations, FOXO1 is required for keratinocyte transition to a wound-healing phenotype that involves increased migration and up-regulation of transforming growth factor β1 (TGF-β1) and its downstream targets, integrin-α3 and -β6 and MMP-3 and -9. Furthermore, we show that FOXO1 functions in keratinocytes to reduce oxidative stress, which is necessary to maintain cell migration and prevent cell death in a TGF-β1–independent manner. Thus, our studies identify a novel function for FOXO1 in coordinating the response of keratinocytes to wounding through up-regulation of TGF-β1 and other factors needed for keratinocyte migration and protection against oxidative stress, which together promote migration and decrease apoptosis. PMID:24145170

Role of FOXO1 in aldosterone-induced autophagy: A compensatory protective mechanism related to podocyte injury

PubMed Central

Wang, Bin; Ding, Wei; Zhang, Minmin; Li, Hongmei; Guo, Honglei; Lin, Lilu; Chen, Jing; Gu, Yong

2016-01-01

This study was undertaken to elucidate whether and how autophagy was regulated in aldosterone (Aldo)-induced podocyte injury and to examine its role in this model both in vitro and in vivo. In cultured podocytes, Aldo increased autophagy flux as indicated by the enhanced expression of LC3-II/LC3-I and the reduction of p62. Autophagy induction with rapamycin (RP) provided a cytoprotective effect, and inhibition of autophagy with Atg7-specific siRNA, chloroquine (CQ) or 3-methyladenine (3-MA) worsened Aldo-induced podocyte injury by attenuating endoplasmic reticulum (ER) stress. Aldo inhibited Akt phosphorylation but increased the mammalian target of rapamycin (mTOR) signaling pathway; however, Aldo up-regulated the expression of FOXO1 and its downstream effector Rab7. Either knockdown of FOXO1 or Rab7 inhibited Aldo-induced autophagy. Additionally, an elevated level of P300-regulated acetylation of FOXO1 and the interaction of acetylated FOXO1 and Atg7 were also confirmed to be involved in regulating autophagy in Aldo-induced podocytes. Similar results were further confirmed in vivo. We propose that autophagy enhancement through enhancing of the FOXO1/Rab7 axis and post-translational modification of FOXO1 may represent a potential therapeutic strategy against podocyte injury by promoting autophagy. PMID:27244896

Forkhead Transcription Factors: Formulating a FOXO Target for Cognitive Loss.

PubMed

Maiese, Kenneth

2017-01-01

With almost 47 million individuals worldwide suffering from some aspect of dementia, it is clear that cognitive loss impacts a significant proportion of the global population. Unfortunately, definitive treatments to resolve or prevent the onset of cognitive loss are limited. In most cases such care is currently non-existent prompting the need for novel treatment strategies. Mammalian forkhead transcription factors of the O class (FoxO) are one such avenue of investigation that offer an exciting potential to bring new treatments forward for disorders that involve cognitive loss. Here we examine the background, structure, expression, and function of FoxO transcription factors and their role in cognitive loss, programmed cell death in the nervous system with apoptosis and autophagy, and areas to target FoxOs for dementia and specific disorders such as Alzheimer's disease. FoxO proteins work in concert with a number of other cell survival pathways that involve growth factors, such as erythropoietin and neurotrophins, silent mating type information regulation 2 homolog 1 (Saccharomyces cerevisiae) (SIRT1), Wnt1 inducible signaling pathway protein 1 (WISP1), Wnt signaling, and cancer-related pathways. FoxO transcription factors oversee proinflammatory pathways, affect nervous system amyloid (Aβ) production and toxicity, lead to mitochondrial dysfunction, foster neuronal apoptotic cell death, and accelerate the progression of degenerative disease. However, under some scenarios such as those involving autophagy, FoxOs also can offer protection in the nervous system and reduce toxic intracellular protein accumulations and potentially limit Aβ toxicity. Given the ability of FoxOs to not only promote apoptotic cell death in the nervous system, but also through the induction of autophagy offer protection against degenerative disease that can lead to dementia, a fine balance in the activity of FoxOs may be required to target cognitive loss in individuals. Future work should

MicroRNA-214 suppresses gluconeogenesis by targeting activating transcriptional factor 4.

PubMed

Li, Kai; Zhang, Jin; Yu, Junjie; Liu, Bin; Guo, Yajie; Deng, Jiali; Chen, Shanghai; Wang, Chunxia; Guo, Feifan

2015-03-27

Although the gluconeogenesis pathway is already a target for the treatment of type 2 diabetes, the potential role of microRNAs (miRNAs) in gluconeogenesis remains unclear. Here, we investigated the physiological functions of miR-214 in gluconeogenesis. The expression of miR-214 was suppressed by glucagon via protein kinase A signaling in primary hepatocytes, and miR-214 was down-regulated in the livers of fasted, high fat diet-induced diabetic and leptin receptor-mutated (db/db) mice. The overexpression of miR-214 in primary hepatocytes suppressed glucose production, and silencing miR-214 reversed this effect. Gluconeogenesis was suppressed in the livers of mice injected with an adenovirus expressing miR-214 (Ad-miR-214). Additionally, Ad-miR-214 alleviated high fat diet-induced elevation of gluconeogenesis and hyperglycemia. Furthermore, we found that activating transcription factor 4 (ATF4), a reported target of miR-214, can reverse the suppressive effect of miR-214 on gluconeogenesis in primary hepatocytes, and this suppressive effect was blocked in liver-specific ATF4 knock-out mice. ATF4 regulated gluconeogenesis via affecting forkhead box protein O1 (FOXO1) transcriptional activity. Finally, liver-specific miR-214 transgenic mice exhibited suppressed gluconeogenesis and reduced expression of ATF4, phosphoenolpyruvate carboxykinase, and glucose-6-phosphatase in liver. Taken together, our results suggest that the miR-214-ATF4 axis is a novel pathway for the regulation of hepatic gluconeogenesis. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Inhibition of Gluconeogenesis in Primary Hepatocytes by Stromal Cell-derived Factor-1 (SDF-1) through a c-Src/Akt-dependent Signaling Pathway*

PubMed Central

Liu, Hui-Yu; Wen, Ge-Bo; Han, Jianmin; Hong, Tao; Zhuo, Degen; Liu, Zhenqi; Cao, Wenhong

2008-01-01

Hepatic gluconeogenesis is elevated in diabetes and a major contributor to hyperglycemia. Stromal cell-derived factor-1 (SDF-1) is a chemokine and an activator of Akt. In this study, we tested the hypothesis that SDF-1 suppresses hepatic gluconeogenesis through Akt. Our results from isolated primary hepatocytes show that SDF-1α and SDF-1β inhibited glucose production via gluconeogenesis and reduced transcript levels of key gluconeogenic genes glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase (PEPCK). Additionally, SDF-1α and SDF-1β both inhibited activation of the PEPCK promoter. In examining the mechanism by which SDF-1 inhibits gluconeogenesis, we found that SDF-1 promoted phosphorylation of Akt, FoxO1, and c-Src, but did not activate insulin receptor substrate-1-like insulin. Blockade of Akt activation by LY294002, FoxO1 translocation by constitutively nuclear FoxO1 mutant, or c-Src activation by the chemical inhibitor PP2, respectively, blunted SDF-1 suppression of gluconeogenesis. Finally, our results show that knocking down the level of SDF-1 receptor CXCR4 mRNA blocked SDF-1 suppression of gluconeogenesis. Together, our results demonstrate that SDF-1 is capable of inhibiting gluconeogenesis in primary hepatocytes through a signaling pathway distinct from the insulin signaling. PMID:18786922

FoxO6 regulates memory consolidation and synaptic function

PubMed Central

Salih, Dervis A.M.; Rashid, Asim J.; Colas, Damien; de la Torre-Ubieta, Luis; Zhu, Ruo P.; Morgan, Alexander A.; Santo, Evan E.; Ucar, Duygu; Devarajan, Keerthana; Cole, Christina J.; Madison, Daniel V.; Shamloo, Mehrdad; Butte, Atul J.; Bonni, Azad; Josselyn, Sheena A.; Brunet, Anne

2012-01-01

The FoxO family of transcription factors is known to slow aging downstream from the insulin/IGF (insulin-like growth factor) signaling pathway. The most recently discovered FoxO isoform in mammals, FoxO6, is highly enriched in the adult hippocampus. However, the importance of FoxO factors in cognition is largely unknown. Here we generated mice lacking FoxO6 and found that these mice display normal learning but impaired memory consolidation in contextual fear conditioning and novel object recognition. Using stereotactic injection of viruses into the hippocampus of adult wild-type mice, we found that FoxO6 activity in the adult hippocampus is required for memory consolidation. Genome-wide approaches revealed that FoxO6 regulates a program of genes involved in synaptic function upon learning in the hippocampus. Consistently, FoxO6 deficiency results in decreased dendritic spine density in hippocampal neurons in vitro and in vivo. Thus, FoxO6 may promote memory consolidation by regulating a program coordinating neuronal connectivity in the hippocampus, which could have important implications for physiological and pathological age-dependent decline in memory. PMID:23222102

Increased Hepatic Glucose Production in Fetal Sheep With Intrauterine Growth Restriction Is Not Suppressed by Insulin

PubMed Central

Thorn, Stephanie R.; Brown, Laura D.; Rozance, Paul J.; Hay, William W.; Friedman, Jacob E.

2013-01-01

Intrauterine growth restriction (IUGR) increases the risk for metabolic disease and diabetes, although the developmental origins of this remain unclear. We measured glucose metabolism during basal and insulin clamp periods in a fetal sheep model of placental insufficiency and IUGR. Compared with control fetuses (CON), fetuses with IUGR had increased basal glucose production rates and hepatic PEPCK and glucose-6-phosphatase expression, which were not suppressed by insulin. In contrast, insulin significantly increased peripheral glucose utilization rates in CON and IUGR fetuses. Insulin robustly activated AKT, GSK3β, and forkhead box class O (FOXO)1 in CON and IUGR fetal livers. IUGR livers, however, had increased basal FOXO1 phosphorylation, nuclear FOXO1 expression, and Jun NH2-terminal kinase activation during hyperinsulinemia. Expression of peroxisome proliferator–activated receptor γ coactivator 1α and hepatocyte nuclear factor-4α were increased in IUGR livers during basal and insulin periods. Cortisol and norepinephrine concentrations were positively correlated with glucose production rates. Isolated IUGR hepatocytes maintained increased glucose production in culture. In summary, fetal sheep with IUGR have increased hepatic glucose production, which is not suppressed by insulin despite insulin sensitivity for peripheral glucose utilization. These data are consistent with a novel mechanism involving persistent transcriptional activation in the liver that seems to be unique in the fetus with IUGR. PMID:22933111

Loss of immunological tolerance in Gimap5-deficient mice is associated with loss of Foxo in CD4+ T cells

PubMed Central

Aksoylar, H. Ibrahim; Lampe, Kristin; Barnes, Michael J.; Plas, David R.; Hoebe, Kasper

2011-01-01

Previously, we reported the abrogation of quiescence and reduced survival in lymphocytes from Gimap5sph/sph mice, an ENU germline mutant with a missense mutation in the GTPase of immunity-associated nucleotide binding protein 5 (Gimap5). These mice showed a progressive loss of peripheral lymphocyte populations and developed spontaneous colitis, resulting in early mortality. Here, we identify the molecular pathways that contribute to the onset of colitis in Gimap5sph/sph mice. We show that CD4+ T cells become Th1/Th17-polarized and are critically important for the development of colitis. Concomitantly, Treg cells become reduced in frequency in the peripheral tissues and their immune-suppressive capacity becomes impaired. Most importantly, these progressive changes in CD4+ T cells are associated with the loss of Foxo1, Foxo3 and Foxo4 expression. Our data establish a novel link between Gimap5 and Foxo expression and provide evidence for a regulatory mechanism that controls Foxo protein expression and may help maintain immunological tolerance. PMID:22106000

Targeting the UPR transcription factor XBP1 protects against Huntington's disease through the regulation of FoxO1 and autophagy

PubMed Central

Vidal, Rene L.; Figueroa, Alicia; Court, Felipe A.; Thielen, Peter; Molina, Claudia; Wirth, Craig; Caballero, Benjamin; Kiffin, Roberta; Segura-Aguilar, Juan; Cuervo, Ana Maria; Glimcher, Laurie H.; Hetz, Claudio

2012-01-01

Mutations leading to expansion of a poly-glutamine track in Huntingtin (Htt) cause Huntington's disease (HD). Signs of endoplasmic reticulum (ER) stress have been recently reported in animal models of HD, associated with the activation of the unfolded protein response (UPR). Here we have investigated the functional contribution of ER stress to HD by targeting the expression of two main UPR transcription factors, XBP1 and ATF4 (activating transcription factor 4), in full-length mutant Huntingtin (mHtt) transgenic mice. XBP1-deficient mice were more resistant to developing disease features, associated with improved neuronal survival and motor performance, and a drastic decrease in mHtt levels. The protective effects of XBP1 deficiency were associated with enhanced macroautophagy in both cellular and animal models of HD. In contrast, ATF4 deficiency did not alter mHtt levels. Although, XBP1 mRNA splicing was observed in the striatum of HD transgenic brains, no changes in the levels of classical ER stress markers were detected in symptomatic animals. At the mechanistic level, we observed that XBP1 deficiency led to augmented expression of Forkhead box O1 (FoxO1), a key transcription factor regulating autophagy in neurons. In agreement with this finding, ectopic expression of FoxO1 enhanced autophagy and mHtt clearance in vitro. Our results provide strong evidence supporting an involvement of XBP1 in HD pathogenesis probably due to an ER stress-independent mechanism involving the control of FoxO1 and autophagy levels. PMID:22337954

Modeling of the Human Alveolar Rhabdomyosarcoma Pax3-Foxo1 Chromosome Translocation in Mouse Myoblasts Using CRISPR-Cas9 Nuclease

PubMed Central

Lagutina, Irina V.; Valentine, Virginia; Picchione, Fabrizio; Harwood, Frank; Valentine, Marcus B.; Villarejo-Balcells, Barbara; Carvajal, Jaime J.; Grosveld, Gerard C.

2015-01-01

Many recurrent chromosome translocations in cancer result in the generation of fusion genes that are directly implicated in the tumorigenic process. Precise modeling of the effects of cancer fusion genes in mice has been inaccurate, as constructs of fusion genes often completely or partially lack the correct regulatory sequences. The reciprocal t(2;13)(q36.1;q14.1) in human alveolar rhabdomyosarcoma (A-RMS) creates a pathognomonic PAX3-FOXO1 fusion gene. In vivo mimicking of this translocation in mice is complicated by the fact that Pax3 and Foxo1 are in opposite orientation on their respective chromosomes, precluding formation of a functional Pax3-Foxo1 fusion via a simple translocation. To circumvent this problem, we irreversibly inverted the orientation of a 4.9 Mb syntenic fragment on chromosome 3, encompassing Foxo1, by using Cre-mediated recombination of two pairs of unrelated oppositely oriented LoxP sites situated at the borders of the syntenic region. We tested if spatial proximity of the Pax3 and Foxo1 loci in myoblasts of mice homozygous for the inversion facilitated Pax3-Foxo1 fusion gene formation upon induction of targeted CRISPR-Cas9 nuclease-induced DNA double strand breaks in Pax3 and Foxo1. Fluorescent in situ hybridization indicated that fore limb myoblasts show a higher frequency of Pax3/Foxo1 co-localization than hind limb myoblasts. Indeed, more fusion genes were generated in fore limb myoblasts via a reciprocal t(1;3), which expressed correctly spliced Pax3-Foxo1 mRNA encoding Pax3-Foxo1 fusion protein. We conclude that locus proximity facilitates chromosome translocation upon induction of DNA double strand breaks. Given that the Pax3-Foxo1 fusion gene will contain all the regulatory sequences necessary for precise regulation of its expression, we propose that CRISPR-Cas9 provides a novel means to faithfully model human diseases caused by chromosome translocation in mice. PMID:25659124

Ginsenoside Compound K suppresses the hepatic gluconeogenesis via activating adenosine-5'monophosphate kinase: A study in vitro and in vivo.

PubMed

Wei, Shengnan; Li, Wei; Yu, Yang; Yao, Fan; A, Lixiang; Lan, Xiaoxin; Guan, Fengying; Zhang, Ming; Chen, Li

2015-10-15

Compound K (CK) is a final intestinal metabolite of protopanaxadiol-type ginsenoside. We have reported that CK presented anti-diabetic effect via diminishing the expressions of hepatic gluconeogenesis key enzyme. Here, we further explore the possible mechanism of CK on suppression hepatic gluconeogenesis via activation of adenosine-5'monophosphate kinase (AMPK) on type 2 diabetes mice in vivo and in HepG2 cells. Type 2 diabetes mice model was developed by high fat diet combined with STZ injection. 30mg/kg/d CK was orally administrated for 4weeks, the fasting blood glucose level and 2h OGTT were conducted, and the protein expression of AMPK, peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), Phosphoenolpyruvate carboxykinase (PEPCK) and Glucose-6-phosphatase (G6Pase) were examined. The mechanism of Compound K on hepatic gluconeogenesis was further explored in HepG2 hepatocytes. Glucose production, the protein expression of AMPK, PEPCK, G6pase and PGC-1α, hepatic nuclear factor 4α (HNF-4α) and forkhead transcription factor O1 (FOXO1) were determined after Compound K treatment at the presence of AMPK inhibitor Compound C. We observed that CK inhibited the expression of PEPCK and G6Pase in the liver and in HepG2 hepatocytes. Meanwhile, CK treatment remarkably increased the activation of AMPK, while decreasing the expressions of PGC-1α, HNF-4α and FOXO1. However, AMPK inhibitor Compound C could reverse these effects of CK on gluconeogenesis in part. The results indicated that the effect of CK on suppression hepatic gluconeogenesis might be via the activation the AMPK activity. Copyright © 2015. Published by Elsevier Inc.

FOXO1 opposition of CD8+ T cell effector programming confers early memory properties and phenotypic diversity.

PubMed

Delpoux, Arnaud; Lai, Chen-Yen; Hedrick, Stephen M; Doedens, Andrew L

2017-10-17

The factors and steps controlling postinfection CD8 + T cell terminal effector versus memory differentiation are incompletely understood. Whereas we found that naive TCF7 (alias "Tcf-1") expression is FOXO1 independent, early postinfection we report bimodal, FOXO1-dependent expression of the memory-essential transcription factor TCF7 in pathogen-specific CD8 + T cells. We determined the early postinfection TCF7 high population is marked by low TIM3 expression and bears memory signature hallmarks before the appearance of established memory precursor marker CD127 (IL-7R). These cells exhibit diminished TBET, GZMB, mTOR signaling, and cell cycle progression. Day 5 postinfection, TCF7 high cells express higher memory-associated BCL2 and EOMES, as well as increased accumulation potential and capacity to differentiate into memory phenotype cells. TCF7 retroviral transduction opposes GZMB expression and the formation of KLRG1 pos phenotype cells, demonstrating an active role for TCF7 in extinguishing the effector program and forestalling terminal differentiation. Past the peak of the cellular immune response, we report a gradient of FOXO1 and TCF7 expression, which functions to oppose TBET and orchestrate a continuum of effector-to-memory phenotypes.

Investigation of FoxO3 dynamics during erythroblast development in β-thalassemia major

PubMed Central

Thanuthanakhun, Naruchit; Nuntakarn, Lalana; Sampattavanich, Somponnat; Anurathapan, Usanarat; Phuphanitcharoenkun, Suphanun; Pornpaiboonstid, Savichaya; Hongeng, Suradej

2017-01-01

The FoxO3 transcription factor is a key regulator of oxidative stress and erythroid maturation during erythropoiesis. In this study, we explored the involvement of FoxO3 in severe β-thalassemia. Using primary CD34+ hematopoietic progenitor cells from patients with β-thalassemia major, we successfully developed an in vitro model of ineffective erythropoiesis. Based on this model, FoxO3 activity was quantified in single cells using high throughput imaging flow cytometry. This study revealed a significant reduction of FoxO3 activity during the late stage of erythroblast differentiation in β-thalassemia, in contrast to erythropoiesis in normal cells that maintain persistent activation of FoxO3. In agreement with the decreased FoxO3 activity in β-thalassemia, the expression of FoxO3 target genes was also found to decrease, concurrent with elevated phosphorylation of AKT, most clearly at the late stage of erythroid differentiation. Our findings provide further evidence for the involvement of FoxO3 during terminal erythropoiesis and confirm the modulation of the PI3K/AKT pathway as a potential therapeutic strategy for β-thalassemia. PMID:29099866

Investigation of FoxO3 dynamics during erythroblast development in β-thalassemia major.

PubMed

Thanuthanakhun, Naruchit; Nuntakarn, Lalana; Sampattavanich, Somponnat; Anurathapan, Usanarat; Phuphanitcharoenkun, Suphanun; Pornpaiboonstid, Savichaya; Borwornpinyo, Suparerk; Hongeng, Suradej

2017-01-01

The FoxO3 transcription factor is a key regulator of oxidative stress and erythroid maturation during erythropoiesis. In this study, we explored the involvement of FoxO3 in severe β-thalassemia. Using primary CD34+ hematopoietic progenitor cells from patients with β-thalassemia major, we successfully developed an in vitro model of ineffective erythropoiesis. Based on this model, FoxO3 activity was quantified in single cells using high throughput imaging flow cytometry. This study revealed a significant reduction of FoxO3 activity during the late stage of erythroblast differentiation in β-thalassemia, in contrast to erythropoiesis in normal cells that maintain persistent activation of FoxO3. In agreement with the decreased FoxO3 activity in β-thalassemia, the expression of FoxO3 target genes was also found to decrease, concurrent with elevated phosphorylation of AKT, most clearly at the late stage of erythroid differentiation. Our findings provide further evidence for the involvement of FoxO3 during terminal erythropoiesis and confirm the modulation of the PI3K/AKT pathway as a potential therapeutic strategy for β-thalassemia.

Identification of the tumor‑suppressive function of circular RNA FOXO3 in non‑small cell lung cancer through sponging miR‑155.

PubMed

Zhang, Yanni; Zhao, Hui; Zhang, Lichuan

2018-06-01

Circular RNAs (circRNAs) are a class of endo-genous noncoding RNAs that have been demonstrated to be potential regulators in the development and progression of various types of human cancer. However, little is known about their roles in cancer initiation and progression, particular in non‑small cell lung cancer (NSCLC). In the present study, the expression level of circRNA‑forkhead box O3 class (FOXO3) in NSCLC specimens was determined and its functional role was investigated in NSCLC cells. By performing Taq‑man based RT‑qPCR, it was identified that circRNA‑FOXO3 was downregulated in NSCLC tissues and cell lines. Receiver operating curve analysis indicated that circRNA‑FOXO3 had a relatively higher diagnostic accuracy. The functional relevance was further examined by biological assays. circRNA‑FOXO3 significantly promoted the ability of cell proliferation, migration and invasion of NSCLC cells. The linear isomer of circRNA‑FOXO3, FOXO3 gene, was identified as a downstream target. RNA immunoprecipitation indicated that circRNA‑FOXO3 sequestering miR‑155, which further promoted linear FOXO3 expression. In addition, gain and loss functional assays indicated that circRNA‑FOXO3 served an anti‑oncogenic role through sequestering miR‑155 and enhancing FOXO3 expression. These results suggest that circRNA‑FOXO3 is a tumor‑suppressor in NSCLC and may serve as a promising therapeutic target. Therefore, restoration of circRNA‑FOXO3 expression could be a future approach to develop a novel treatment strategy.

Type I neuregulin1α is a novel local mediator to suppress hepatic gluconeogenesis in mice

PubMed Central

Arai, Takatomo; Ono, Yumika; Arimura, Yujiro; Sayama, Keimon; Suzuki, Tomohiro; Shinjo, Satoko; Kanai, Mai; Abe, Shin-ichi; Semba, Kentaro; Goda, Nobuhito

2017-01-01

Neuregulin1 is an epidermal growth factor (EGF)-like domain-containing protein that has multiple isoforms and functions as a local mediator in the control of various cellular functions. Here we show that type I isoform of neuregulin1 with an α-type EGF-like domain (Nrg1α) is the major isoform in mouse liver and regulates hepatic glucose production. Forced expression of Nrg1α in mouse liver enhanced systemic glucose disposal and decreased hepatic glucose production with reduced fasting blood glucose levels. Nuclear forkhead box protein O1 (FoxO1) and its downstream targets, PEPCK and G6Pase, were suppressed in liver and isolated hepatocytes by Nrg1α overexpression. In contrast, silencing of Nrg1α enhanced glucose production with increased PEPCK and G6Pase expressions in cAMP/dexamethasone-stimulated hepatocytes. Mechanistically, the recombinant α-type EGF-like domain of NRG1α (rNRG1α) stimulated the ERBB3 signalling pathway in hepatocytes, resulting in decreased nuclear FoxO1 accumulation via activation of both the AKT and ERK pathways. In addition, acute treatment with rNRG1α also suppressed elevation of blood glucose levels after both glucose and pyruvate challenge. Although a liver-specific deletion of Nrg1 gene in mice showed little effect on systemic glucose metabolism, these results suggest that NRG1α have a novel regulatory function in hepatic gluconeogenesis by regulating the ERBB3-AKT/ERK-FoxO1 cascade. PMID:28218289

Evolutionary acquisition of cysteines determines FOXO paralog-specific redox signaling.

PubMed

Putker, Marrit; Vos, Harmjan R; van Dorenmalen, Kim; de Ruiter, Hesther; Duran, Ana G; Snel, Berend; Burgering, Boudewijn M T; Vermeulen, Michiel; Dansen, Tobias B

2015-01-01

Reduction-oxidation (redox) signaling, the translation of an oxidative intracellular environment into a cellular response, is mediated by the reversible oxidation of specific cysteine thiols. The latter can result in disulfide formation between protein hetero- or homodimers that alter protein function until the local cellular redox environment has returned to the basal state. We have previously shown that this mechanism promotes the nuclear localization and activity of the Forkhead Box O4 (FOXO4) transcription factor. In this study, we sought to investigate whether redox signaling differentially controls the human FOXO3 and FOXO4 paralogs. We present evidence that FOXO3 and FOXO4 have acquired paralog-specific cysteines throughout vertebrate evolution. Using a proteome-wide screen, we identified previously unknown redox-dependent FOXO3 interaction partners. The nuclear import receptors Importin-7 (IPO7) and Importin-8 (IPO8) form a disulfide-dependent heterodimer with FOXO3, which is required for its reactive oxygen species-induced nuclear translocation. FOXO4 does not interact with IPO7 or IPO8. IPO7 and IPO8 control the nuclear import of FOXO3, but not FOXO4, in a redox-sensitive and disulfide-dependent manner. Our findings suggest that evolutionary acquisition of cysteines has contributed to regulatory divergence of FOXO paralogs, and that phylogenetic analysis can aid in the identification of cysteines involved in redox signaling.

FoxO and Stress Responses in the Cnidarian Hydra vulgaris

PubMed Central

Bridge, Diane; Theofiles, Alexander G.; Holler, Rebecca L.; Marcinkevicius, Emily; Steele, Robert E.; Martínez, Daniel E.

2010-01-01

Background In the face of changing environmental conditions, the mechanisms underlying stress responses in diverse organisms are of increasing interest. In vertebrates, Drosophila, and Caenorhabditis elegans, FoxO transcription factors mediate cellular responses to stress, including oxidative stress and dietary restriction. Although FoxO genes have been identified in early-arising animal lineages including sponges and cnidarians, little is known about their roles in these organisms. Methods/Principal Findings We have examined the regulation of FoxO activity in members of the well-studied cnidarian genus Hydra. We find that Hydra FoxO is expressed at high levels in cells of the interstitial lineage, a cell lineage that includes multipotent stem cells that give rise to neurons, stinging cells, secretory cells and gametes. Using transgenic Hydra that express a FoxO-GFP fusion protein in cells of the interstitial lineage, we have determined that heat shock causes localization of the fusion protein to the nucleus. Our results also provide evidence that, as in bilaterian animals, Hydra FoxO activity is regulated by both Akt and JNK kinases. Conclusions These findings imply that basic mechanisms of FoxO regulation arose before the evolution of bilaterians and raise the possibility that FoxO is involved in stress responses of other cnidarian species, including corals. PMID:20657733

FoxO and stress responses in the cnidarian Hydra vulgaris.

PubMed

Bridge, Diane; Theofiles, Alexander G; Holler, Rebecca L; Marcinkevicius, Emily; Steele, Robert E; Martínez, Daniel E

2010-07-21

In the face of changing environmental conditions, the mechanisms underlying stress responses in diverse organisms are of increasing interest. In vertebrates, Drosophila, and Caenorhabditis elegans, FoxO transcription factors mediate cellular responses to stress, including oxidative stress and dietary restriction. Although FoxO genes have been identified in early-arising animal lineages including sponges and cnidarians, little is known about their roles in these organisms. We have examined the regulation of FoxO activity in members of the well-studied cnidarian genus Hydra. We find that Hydra FoxO is expressed at high levels in cells of the interstitial lineage, a cell lineage that includes multipotent stem cells that give rise to neurons, stinging cells, secretory cells and gametes. Using transgenic Hydra that express a FoxO-GFP fusion protein in cells of the interstitial lineage, we have determined that heat shock causes localization of the fusion protein to the nucleus. Our results also provide evidence that, as in bilaterian animals, Hydra FoxO activity is regulated by both Akt and JNK kinases. These findings imply that basic mechanisms of FoxO regulation arose before the evolution of bilaterians and raise the possibility that FoxO is involved in stress responses of other cnidarian species, including corals.

Adenosine monophosphate-activated protein kinase attenuates cardiomyocyte hypertrophy through regulation of FOXO3a/MAFbx signaling pathway.

PubMed

Chen, Baolin; Wu, Qiang; Xiong, Zhaojun; Ma, Yuedong; Yu, Sha; Chen, Dandan; Huang, Shengwen; Dong, Yugang

2016-09-01

Control of cardiac muscle mass is thought to be determined by a dynamic balance of protein synthesis and degradation. Recent studies have demonstrated that atrophy-related forkhead box O 3a (FOXO3a)/muscle atrophy F-box (MAFbx) signaling pathway plays a central role in the modulation of proteolysis and exert inhibitory effect on cardiomyocyte hypertrophy. In this study, we tested the hypothesis that adenosine monophosphate-activated protein kinase (AMPK) activation attenuates cardiomyocyte hypertrophy by regulating FOXO3a/MAFbx signaling pathway and its downstream protein degradation. The results showed that activation of AMPK with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) attenuated cardiomyocyte hypertrophy induced by angiotensin II (Ang II). The antihypertrophic effects of AICAR were blunted by AMPK inhibitor Compound C. In addition, AMPK dramatically increased the activity of transcription factor FOXO3a, up-regulated the expression of its downstream ubiquitin ligase MAFbx, and enhanced cardiomyocyte proteolysis. Meanwhile, the effects of AMPK on protein degradation and cardiomyocyte hypertrophy were blocked after MAFbx was silenced by transfection of cardiomyocytes with MAFbx-siRNA. These results indicate that AMPK plays an important role in the inhibition of cardiomyocyte hypertrophy by activating protein degradation via FOXO3a/MAFbx signaling pathway. © The Author 2016. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Proliferation of murine c-kit(pos) cardiac stem cells stimulated with IGF-1 is associated with Akt-1 mediated phosphorylation and nuclear export of FoxO3a and its effect on downstream cell cycle regulators.

PubMed

Johnson, Ann Mary; Kartha, C C

2014-04-01

Insulin-like growth factor-1 (IGF-1) is known to promote proliferation in many cell types including c-kit(pos) cardiac stem cells (CSCs). Downstream signaling pathways of IGF-1 induced CSC proliferation have not been investigated. An important downstream target of IGF-1/Akt-1 signaling is FoxO3a, a key negative regulator of cell-cycle progression. We studied the effect of IGF-1 on proliferation of c-kit(pos) murine CSCs and found that IGF-1-mediated cell proliferation is associated with FoxO3a phosphorylation and inactivation of its transcriptional activity. PI3 inhibitors LY294002 and Wortmannin abolished the effect of IGF-1 on FoxO3a phosphorylation indicating that FoxO3a phosphorylation is mediated by PI3/Akt-1 pathway. In cells with FoxO3a translocation to the cytoplasm, there is decreased expression of cell-cycle inhibitors such as p27(kip1) and p57(kip2) and increased expression of CyclinD1. Our study provides evidence that IGF-1 induced CSC proliferation could be the result of FoxO3a inactivation and its downstream effect on cell-cycle regulators.

Autophagy protects chondrocytes from glucocorticoids-induced apoptosis via ROS/Akt/FOXO3 signaling.

PubMed

Shen, C; Cai, G-Q; Peng, J-P; Chen, X-D

2015-12-01

Glucocorticoids (GCs) have been widely used in the management of osteoarthritis (OA) and rheumatoid arthritis (RA). Nevertheless, there has been some concern about their ability of increasing reactive oxygen species (ROS) in the cartilage. Forkhead-box class O (FOXO) transcription factors have been proved to have a protective role in chondrocytes through regulation of autophagy and defending oxidative stress. The objective of this study was to investigate the role of FOXO3 in Dex-induce up-regulation of ROS. Healthy cartilages debris from six patients were used for chondrocytes culture. After the treatment of dexamethasone (Dex), the ROS levels, autophagic flux, the expression of FOXO3 in chondrocytes were measured. RNA interference technique was also used to determine the role of FOXO3 in Dex-induced autophagy. The metabolism of the extra-cellular matrix was also investigated. Dex increased intracellular ROS level, the expression of Akt, FOXO3 as well as autophagy flux in human chondrocytes. The expression of aggrecanases also increased after the treatment of Dex. Catalase, the ROS scavenger, suppressed Dex-induced up-regulation of autophagy flux and expression of aggrecanases and Akt. MK-2206 and LY294002, the PI3K/Akt inhibitors, repressed Dex-induced up-regulation of FOXO3. Silencing FOXO3 resulted in down-regulation of Dex-induced autophagy. Moreover, knockdown of FOXO3 increased Dex-induced apoptosis as well as ROS levels in chondrocytes. In addition, up-regulation of autophagy by Rapamycin resulted in decreasing ROS level in chondrocytes. Dex could advance the degenerative process in cartilage. Autophagy was induced in response to Dex-induced up-regulation of ROS via ROS/Akt/FOXO3 signal pathway. Copyright © 2015 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Attenuation of MUC4 potentiates the anticancer activity of auranofin via regulation of the Her2/Akt/FOXO3 pathway in ovarian cancer cells.

PubMed

Bae, Jun Sang; Lee, Jongsung; Park, Yoonkook; Park, Kyungmoon; Kim, Jung Ryul; Cho, Dong Hyu; Jang, Kyu Yun; Park, See-Hyoung

2017-10-01

Previously, we reported that auranofin induces apoptosis in SKOV3 cells via regulation of the IKKβ/FOXO3 pathway. In the present study, we reveal that the anticancer activity of auranofin in SKOV3 cells could be enhanced by the attenuation of MUC4 through the regulation of the Her2/Akt/FOXO3 pathway. Compared to the control-siRNA, siRNA transfection against MUC4 into SKOV3 cells accelerated the protein degradation of Her2. Under the same conditions, the expression level of phosphorylated Akt was also downregulated leading to an increase of FOXO3 in the nucleus. Notably, auranofin treatment in SKOV3 cells also resulted in the downregulation of the expression levels of both Her2 and phosphorylated Akt. Thus, Her2 was identified as the common molecular target protein by siRNA transfection against MUC4. Western blot analysis of total and nuclear fraction lysates from SKOV3 cells revealed that attenuation of MUC4 combined with auranofin treatment in SKOV3 cells synergistically activated FOXO3 translocation from the cytoplasm to the nucleus through the regulation of the Her2/Akt/FOXO3 pathway. Attenuation of MUC4 by siRNA transfection potentiated the antitumor effect of auranofin which was examined by performing in vitro assays such as WST-1, cell counting, colony formation, TUNEL and Annexin V staining. In addition, western blot analysis of the apoptosis‑related proteins such as PARP1, caspase-3, Bim extra large (EL), Bax and Bcl2 revealed that the attenuation of MUC4 by siRNA transfection potentiates the pro-apoptotic activity of auranofin in SKOV3 cells. Collectively, auranofin could regulate the Her2/Akt/FOXO3 signaling pathway in SKOV3 cells and be used as a potential antitumor agent considering the expression of MUC4 in ovarian cancer patients.

A role for FOXO1 in BCR–ABL1-independent tyrosine kinase inhibitor resistance in chronic myeloid leukemia

PubMed Central

Wagle, M; Eiring, A M; Wongchenko, M; Lu, S; Guan, Y; Wang, Y; Lackner, M; Amler, L; Hampton, G; Deininger, M W; O'Hare, T; Yan, Y

2016-01-01

Chronic myeloid leukemia (CML) patients who relapse on imatinib due to acquired ABL1 kinase domain mutations are successfully treated with second-generation ABL1-tyrosine kinase inhibitors (ABL-TKIs) such as dasatinib, nilotinib or ponatinib. However, ~40% of relapsed patients have uncharacterized BCR–ABL1 kinase-independent mechanisms of resistance. To identify these mechanisms of resistance and potential treatment options, we generated ABL-TKI-resistant K562 cells through prolonged sequential exposure to imatinib and dasatinib. Dual-resistant K562 cells lacked BCR–ABL1 kinase domain mutations, but acquired other genomic aberrations that were characterized by next-generation sequencing and copy number analyses. Proteomics showed that dual-resistant cells had elevated levels of FOXO1, phospho-ERK and BCL-2, and that dasatinib no longer inhibited substrates of the PI3K/AKT pathway. In contrast to parental cells, resistant cells were sensitive to growth inhibition and apoptosis induced by the class I PI3K inhibitor, GDC-0941 (pictilisib), which also induced FOXO1 nuclear translocation. FOXO1 was elevated in a subset of primary specimens from relapsed CML patients lacking BCR–ABL1 kinase domain mutations, and these samples were responsive to GDC-0941 treatment ex vivo. We conclude that elevated FOXO1 contributes to BCR–ABL1 kinase-independent resistance experienced by these CML patients and that PI3K inhibition coupled with BCR–ABL1 inhibition may represent a novel therapeutic approach. PMID:27044711

A role for FOXO1 in BCR-ABL1-independent tyrosine kinase inhibitor resistance in chronic myeloid leukemia.

PubMed

Wagle, M; Eiring, A M; Wongchenko, M; Lu, S; Guan, Y; Wang, Y; Lackner, M; Amler, L; Hampton, G; Deininger, M W; O'Hare, T; Yan, Y

2016-07-01

Chronic myeloid leukemia (CML) patients who relapse on imatinib due to acquired ABL1 kinase domain mutations are successfully treated with second-generation ABL1-tyrosine kinase inhibitors (ABL-TKIs) such as dasatinib, nilotinib or ponatinib. However, ~40% of relapsed patients have uncharacterized BCR-ABL1 kinase-independent mechanisms of resistance. To identify these mechanisms of resistance and potential treatment options, we generated ABL-TKI-resistant K562 cells through prolonged sequential exposure to imatinib and dasatinib. Dual-resistant K562 cells lacked BCR-ABL1 kinase domain mutations, but acquired other genomic aberrations that were characterized by next-generation sequencing and copy number analyses. Proteomics showed that dual-resistant cells had elevated levels of FOXO1, phospho-ERK and BCL-2, and that dasatinib no longer inhibited substrates of the PI3K/AKT pathway. In contrast to parental cells, resistant cells were sensitive to growth inhibition and apoptosis induced by the class I PI3K inhibitor, GDC-0941 (pictilisib), which also induced FOXO1 nuclear translocation. FOXO1 was elevated in a subset of primary specimens from relapsed CML patients lacking BCR-ABL1 kinase domain mutations, and these samples were responsive to GDC-0941 treatment ex vivo. We conclude that elevated FOXO1 contributes to BCR-ABL1 kinase-independent resistance experienced by these CML patients and that PI3K inhibition coupled with BCR-ABL1 inhibition may represent a novel therapeutic approach.

Elucidating the role of the FoxO3a transcription factor in the IGF-1-induced migration and invasion of uveal melanoma cancer cells.

PubMed

Yan, Fengxia; Liao, Rifang; Farhan, Mohd; Wang, Tinghuai; Chen, Jiashu; Wang, Zhong; Little, Peter J; Zheng, Wenhua

2016-12-01

Uveal melanoma (UM) is the most common primary intraocular malignant tumor of adults. It has high mortality rate due to liver metastasis. However, the epidemiology and pathogenesis of liver metastasis in UM are not elucidated and there is no effective therapy available for preventing the development of this disease. IGF-1 is a growth factor involved in cell proliferation, malignant transformation and inhibition of apoptosis. In previous report, IGF-1 receptor was found to be highly expressed in UM and this was related to tumor prognosis. FoxO3a is a Forkhead box O (FOXO) transcription factor and a downstream target of the IGF-1R/PI3K/Akt pathway involved in a number of physiological and pathological processes including cancer. However, the role of FoxO3a in UM is unknown. In the present study, we investigated fundamental mechanisms in the growth, migration and invasion of UM and the involvement of FoxO3a. IGF-1 increased the cell viability, invasion, migration and S-G2/M cell cycle phase accumulation of UM cells. Western blot analysis showed that IGF-1 led to activation of Akt and concomitant phosphorylation of FoxO3a. FoxO3a phosphorylation was associated with its translocation into the cytoplasm from the nucleus and its functional inhibition led to the inhibition of expression of Bim and p27, but an increase in the expression of Cyclin D1. The effects of IGF-1 on UM cells were reversed by LY294002 (a PI3K inhibitor) or Akt siRNA, and the overexpression of FoxO3a also attenuated basal invasion and migration of UM. Taken all together, these results suggest that inhibition of FoxO3a by IGF-1 via the PI3K/Akt pathway has an important role in IGF-1 induced proliferation and invasion of UM cells. These findings also support FoxO3a and IGF signaling may represent a valid target for investigating the development of new strategies for the treatment and prevention of the pathology of UM. Copyright © 2016. Published by Elsevier Masson SAS.

Alcohol alters hepatic FoxO1, p53, and mitochondrial SIRT5 deacetylation function

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

Lieber, Charles S.; Department of Medicine, Mount Sinai School of Medicine, New York, NY 10029; Leo, Maria Anna

2008-08-22

Chronic alcohol consumption affects the gene expression of a NAD-dependent deacetylase Sirtuis 1 (SIRT1) and the peroxisome proliferator-activated receptor-{gamma} coactivator1{alpha} (PGC-1{alpha}). Our aim was to verify that it also alters the forkhead (FoxO1) and p53 transcription factor proteins, critical in the hepatic response to oxidative stress and regulated by SIRT1 through its deacetylating capacity. Accordingly, rats were pair-fed the Lieber-DeCarli alcohol-containing liquid diets for 28 days. Alcohol increased hepatic mRNA expression of FoxO1 (p = 0.003) and p53 (p = 0.001) while corresponding protein levels remained unchanged. However phospho-FoxO1 and phospho-Akt (protein kinase) were both decreased by alcohol consumption (pmore » = 0.04 and p = 0.02, respectively) while hepatic p53 was found hyperacetylated (p = 0.017). Furthermore, mitochondrial SIRT5 was reduced (p = 0.0025), and PGC-1{alpha} hyperacetylated (p = 0.027), establishing their role in protein modification. Thus, alcohol consumption disrupts nuclear-mitochondrial interactions by post-translation protein modifications, which contribute to alteration of mitochondrial biogenesis through the newly discovered reduction of SIRT5.« less

Inactivation of the FoxO3a transcription factor is associated with the production of reactive oxygen species during protein kinase CK2 downregulation-mediated senescence in human colon cancer and breast cancer cells

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

Park, Seong-Yeol; Bae, Young-Seuk, E-mail: ysbae@knu.ac.kr

We previously showed that protein kinase CK2 downregulation mediates senescence through the reactive oxygen species (ROS)–p53–p21{sup Cip1/WAF1} pathway in various human cells. In the present study, we investigated whether the FoxO3a transcription factor is associated with ROS production during CK2 downregulation-induced senescence in human colon cancer HCT116 and breast cancer MCF-7 cells. FoxO3a overexpression suppressed ROS production and p53 stabilization induced by a CK2α knockdown. CK2α downregulation induced nuclear export of FoxO3a through stimulation of AKT-mediated phosphorylation of FoxO3a and decreased transcription of its target genes (Cu/ZnSOD, MnSOD, and catalase). In contrast, CK2α overexpression inhibited AKT-mediated FoxO3a phosphorylation. This resulted inmore » nuclear accumulation of FoxO3a, and elevated expression of its target genes. Therefore, these data indicate for the first time that CK2 downregulation stimulates ROS generation by inhibiting FoxO3a during premature senescence in human colon and breast cancer cells. - Highlights: • FoxO3a overexpression inhibited ROS production mediated by CK2α knockdown. • CK2α downregulation induced nuclear export of FoxO3a via AKT activation. • CK2α downregulation reduced transcription of FoxO3a target genes including SOD. • CK2α upregulation elevated nuclear import and target gene expression of FoxO3a. • This study indicates that CK2 can modulate the intracellular ROS level via FoxO3a.« less

The ins and outs of FoxO shuttling: mechanisms of FoxO translocation and transcriptional regulation.

PubMed Central

Van Der Heide, Lars P; Hoekman, Marco F M; Smidt, Marten P

2004-01-01

FoxO (forkhead box O; forkhead members of the O class) are transcription factors that function under the control of insulin/insulin-like signalling. FoxO factors have been associated with a multitude of biological processes, including cell-cycle, cell death, DNA repair, metabolism and protection from oxidative stress. Central to the regulation of FoxO factors is a shuttling system, which confines FoxO factors to either the nucleus or the cytosol. Shuttling of FoxO requires protein phosphorylation within several domains, and association with 14-3-3 proteins and the nuclear transport machinery. Description of the FoxO-shuttling mechanism contributes to the understanding of FoxO function in relation to signalling and gene regulation. PMID:15005655

Estrogen-mediated inactivation of FOXO3a by the G protein-coupled estrogen receptor GPER.

PubMed

Zekas, Erin; Prossnitz, Eric R

2015-10-15

Estrogen (17β-estradiol) promotes the survival and proliferation of breast cancer cells and its receptors represent important therapeutic targets. The cellular actions of estrogen are mediated by the nuclear estrogen receptors ERα and ERβ as well as the 7-transmembrane spanning G protein-coupled estrogen receptor (GPER). We previously reported that estrogen activates the phosphoinositide 3-kinase (PI3Kinase) pathway via GPER, resulting in phosphatidylinositol (3,4,5)-trisphosphate (PIP3) production within the nucleus of breast cancer cells; however, the mechanisms and consequences of this activity remained unclear. MCF7 breast cancer cells were transfected with GFP-fused Forkhead box O3 (FOXO3) as a reporter to assess localization in response to estrogen stimulation. Inhibitors of PI3Kinases and EGFR were employed to determine the mechanisms of estrogen-mediated FOXO3a inactivation. Receptor knockdown with siRNA and the selective GPER agonist G-1 elucidated the estrogen receptor(s) responsible for estrogen-mediated FOXO3a inactivation. The effects of selective estrogen receptor modulators and downregulators (SERMs and SERDs) on FOXO3a in MCF7 cells were also determined. Cell survival (inhibition of apoptosis) was assessed by caspase activation. In the estrogen-responsive breast cancer cell line MCF7, FOXO3a inactivation occurs on a rapid time scale as a result of GPER, but not ERα, stimulation by estrogen, established by the GPER-selective agonist G-1 and knockdown of GPER and ERα. GPER-mediated inactivation of FOXO3a is effected by the p110α catalytic subunit of PI3Kinase as a result of transactivation of the EGFR. The SERMs tamoxifen and raloxifene, as well as the SERD ICI182,780, were active in mediating FOXO3a inactivation in a GPER-dependent manner. Additionally, estrogen-and G-1-mediated stimulation of MCF7 cells results in a decrease in caspase activation under proapoptotic conditions. Our results suggest that non-genomic signaling by GPER contributes

Multiple elements regulate nuclear/cytoplasmic shuttling of FOXO1: characterization of phosphorylation- and 14-3-3-dependent and -independent mechanisms.

PubMed Central

Zhao, Xiangshan; Gan, Lixia; Pan, Haiyun; Kan, Donghui; Majeski, Michael; Adam, Stephen A; Unterman, Terry G

2004-01-01

FOXO1, a Forkhead transcription factor, is an important target of insulin and growth factor action. Phosphorylation of Thr-24, Ser-256 and Ser-319 promotes nuclear exclusion of FOXO1, yet the mechanisms regulating nuclear/cytoplasmic shuttling of FOXO1 are poorly understood. Previous studies have identified an NLS (nuclear localization signal) in the C-terminal basic region of the DBD (DNA-binding domain), and a leucine-rich, leptomycin-B sensitive NES (nuclear export signal) located further downstream. Here, we find that other elements in the DBD also contribute to nuclear localization, and that multiple mechanisms contribute to nuclear exclusion of FOXO1. Phosphorylation of Ser-319 and a cluster of nearby residues (Ser-322, Ser-325 and Ser-329) functions co-operatively with the nearby NES to promote nuclear exclusion. The N-terminal region of FOXO1 (amino acids 1-149) also is sufficient to promote nuclear exclusion, and does so through multiple mechanisms. Amino acids 1-50 are sufficient to promote nuclear exclusion of green fluorescent protein fusion proteins, and the phosphorylation of Thr-24 is required for this effect. A leucine-rich, leptomycin B-sensitive export signal is also present nearby. Phosphorylated FOXO1 binds 14-3-3 proteins, and co-precipitation studies with tagged proteins indicate that 14-3-3 binding involves co-operative interactions with both Thr-24 and Ser-256. Ser-256 is located in the C-terminal region of the DBD, where 14-3-3 proteins may interfere both with DNA-binding and with nuclear-localization functions. Together, these studies demonstrate that multiple elements contribute to nuclear/cytoplasmic shuttling of FOXO1, and that phosphorylation and 14-3-3 binding regulate the cellular distribution and function of FOXO1 through multiple mechanisms. The presence of these redundant mechanisms supports the concept that the regulation of FOXO1 function plays a critical role in insulin and growth factor action. PMID:14664696

Down-regulation of microRNA-135b inhibited growth of cervical cancer cells by targeting FOXO1.

PubMed

Xu, Yue; Zhao, Shuhua; Cui, Manhua; Wang, Qiang

2015-01-01

More and more evidence has confirmed that dysregulation of microRNAs (miRNAs) can conduce to the progression of human cancers. Previous studied have shown that dysregulation of miR-135b is in varieties of tumors. However, the roles of miR-135b in cervical cancer remain unknown. Therefore, our aim of this study was to explore the biological function and molecular mechanism of miR-135b in cervical cancer cell lines, discussing whether it could be a therapeutic biomarker of cervical cancer in the future. The MTT assay and ELISA-Brdu assay were used to assess cell proliferation. Cell cycle was detected by flow cytometry. Real-time quantitative polymerase chain reaction (PCR) and Western blot analyses were used to detect expressions of cyclin D1, p21, p27 and FOXO1. In our study, we found that miR-135b is up-regulated in cervical cancer cell lines. Down-regulation of miR-135b evidently inhibited proliferation and arrested cell cycle in cervical cancer cells. Bioinformatics analysis predicted that the FOXO1 was a potential target gene of miR-135b. Besides, miR-135b inhibition significantly increased expressions of the cyclin-dependent kinase inhibitors, p21(/CIP1) and p27(/KIP1), and decreased expression of cyclin D1. However, the high level of miR-135b was associated with increased expression of FOXO1 in cervical cancer cells. Further study by luciferase reporter assay demonstrated that miR-135b could directly target FOXO1. Down-regulation of FOXO1 in cervical cancer cells transfected with miR-135b inhibitor partially reversed its inhibitory effects. In conclusion, down-regulation of miR-135b inhibited cell growth in cervical cancer cells by up-regulation of FOXO1.

Tocotrienols inhibit AKT and ERK activation and suppress pancreatic cancer cell proliferation by suppressing the ErbB2 pathway.

PubMed

Shin-Kang, Sonyo; Ramsauer, Victoria P; Lightner, Janet; Chakraborty, Kanishka; Stone, William; Campbell, Sharon; Reddy, Shrikanth A G; Krishnan, Koyamangalath

2011-09-15

Tocotrienols are members of the vitamin E family but, unlike tocopherols, possess an unsaturated isoprenoid side chain that confers superior anti-cancer properties. The ability of tocotrienols to selectively inhibit the HMG-CoA reductase pathway through posttranslational degradation and to suppress the activity of transcription factor NF-κB could be the basis for some of these properties. Our studies indicate that γ- and δ-tocotrienols have potent antiproliferative activity in pancreatic cancer cells (Panc-28, MIA PaCa-2, Panc-1, and BxPC-3). Indeed both tocotrienols induced cell death (>50%) by the MTT cell viability assay in all four pancreatic cancer cell lines. We also examined the effects of the tocotrienols on the AKT and the Ras/Raf/MEK/ERK signaling pathways by Western blotting analysis. γ- and δ-tocotrienol treatment of cells reduced the activation of ERK MAP kinase and that of its downstream mediator RSK (ribosomal protein S6 kinase) in addition to suppressing the activation of protein kinase AKT. Suppression of activation of AKT by γ-tocotrienol led to downregulation of p-GSK-3β and upregulation accompanied by nuclear translocation of Foxo3. These effects were mediated by the downregulation of Her2/ErbB2 at the messenger level. Tocotrienols but not tocopherols were able to induce the observed effects. Our results suggest that the tocotrienol isoforms of vitamin E can induce apoptosis in pancreatic cancer cells through the suppression of vital cell survival and proliferative signaling pathways such as those mediated by the PI3-kinase/AKT and ERK/MAP kinases via downregulation of Her2/ErbB2 expression. The molecular components for this mechanism are not completely elucidated and need further investigation. Copyright © 2011 Elsevier Inc. All rights reserved.

KDM5 Interacts with Foxo to Modulate Cellular Levels of Oxidative Stress

PubMed Central

Liu, Xingyin; Greer, Christina; Secombe, Julie

2014-01-01

Increased cellular levels of oxidative stress are implicated in a large number of human diseases. Here we describe the transcription co-factor KDM5 (also known as Lid) as a new critical regulator of cellular redox state. Moreover, this occurs through a novel KDM5 activity whereby it alters the ability of the transcription factor Foxo to bind to DNA. Our microarray analyses of kdm5 mutants revealed a striking enrichment for genes required to regulate cellular levels of oxidative stress. Consistent with this, loss of kdm5 results in increased sensitivity to treatment with oxidizers, elevated levels of oxidized proteins, and increased mutation load. KDM5 activates oxidative stress resistance genes by interacting with Foxo to facilitate its recruitment to KDM5-Foxo co-regulated genes. Significantly, this occurs independently of KDM5's well-characterized demethylase activity. Instead, KDM5 interacts with the lysine deacetylase HDAC4 to promote Foxo deacetylation, which affects Foxo DNA binding. PMID:25329053

Concurrent acetylation of FoxO1/3a and p53 due to sirtuins inhibition elicit Bim/PUMA mediated mitochondrial dysfunction and apoptosis in berberine-treated HepG2 cells

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

Shukla, Shatrunajay; Department of Medical Elementology and Toxicology, Jamia Hamdard; Sharma, Ankita

Post-translational modifications i.e. phosphorylation and acetylation are pivotal requirements for proper functioning of eukaryotic proteins. The current study aimed to decode the impact of acetylation/deacetylation of non-histone targets i.e. FoxO1/3a and p53 of sirtuins (NAD{sup +} dependent enzymes with lysine deacetylase activity) in berberine treated human hepatoma cells. Berberine (100 μM) inhibited sirtuins significantly (P < 0.05) at transcriptional level as well as at translational level. Combination of nicotinamide (sirtuin inhibitor) with berberine potentiated sirtuins inhibition and increased the expression of FoxO1/3a and phosphorylation of p53 tumor suppressor protein. As sirtuins deacetylate non-histone targets including FoxO1/3a and p53, berberine increasedmore » the acetylation load of FoxO1/3a and p53 proteins. Acetylated FoxO and p53 proteins transcriptionally activate BH3-only proteins Bim and PUMA (3.89 and 3.87 fold respectively, P<0.001), which are known as direct activator of pro-apoptotic Bcl-2 family protein Bax that culminated into mitochondria mediated activation of apoptotic cascade. Bim/PUMA knock-down showed no changes in sirtuins' expression while cytotoxicity induced by berberine and nicotinamide was curtailed up to 28.3% (P < 0.001) and it restored pro/anti apoptotic protein ratio in HepG2 cells. Sirtuins inhibition was accompanied by decline in NAD{sup +}/NADH ratio, ATP generation, enhanced ROS production and decreased mitochondrial membrane potential. TEM analysis confirmed mitochondrial deterioration and cell damage. SRT-1720 (1–10 μM), a SIRT-1 activator, when pre-treated with berberine (25 μM), reversed sirtuins expression comparable to control and significantly restored the cell viability (P < 0.05). Thus, our findings suggest that berberine mediated sirtuins inhibition resulting into FoxO1/3a and p53 acetylation followed by BH3-only protein Bim/PUMA activation may in part be responsible for mitochondria

The Flavones Apigenin and Luteolin Induce FOXO1 Translocation but Inhibit Gluconeogenic and Lipogenic Gene Expression in Human Cells

PubMed Central

Bumke-Vogt, Christiane; Osterhoff, Martin A.; Borchert, Andrea; Guzman-Perez, Valentina; Sarem, Zeinab; Birkenfeld, Andreas L.; Bähr, Volker; Pfeiffer, Andreas F. H.

2014-01-01

The flavones apigenin (4′,5,7,-trihydroxyflavone) and luteolin (3′,4′,5,7,-tetrahydroxyflavone) are plant secondary metabolites with antioxidant, antiinflammatory, and anticancer activities. We evaluated their impact on cell signaling pathways related to insulin-resistance and type 2 diabetes. Apigenin and luteolin were identified in our U-2 OS (human osteosarcoma) cell screening assay for micronutrients triggering rapid intracellular translocation of the forkhead box transcription factor O1 (FOXO1), an important mediator of insulin signal transduction. Insulin reversed the translocation of FOXO1 as shown by live cell imaging. The impact on the expression of target genes was evaluated in HepG2 (human hepatoma) cells. The mRNA-expression of the gluconeogenic enzymes phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pc), the lipogenic enzymes fatty-acid synthase (FASN) and acetyl-CoA-carboxylase (ACC) were down-regulated by both flavones with smaller effective dosages of apigenin than for luteolin. PKB/AKT-, PRAS40-, p70S6K-, and S6-phosphorylation was reduced by apigenin and luteolin but not that of the insulin-like growth factor receptor IGF-1R by apigenin indicating a direct inhibition of the PKB/AKT-signaling pathway distal to the IGF-1 receptor. N-acetyl-L-cysteine did not prevent FOXO1 nuclear translocation induced by apigenin and luteolin, suggesting that these flavones do not act via oxidative stress. The roles of FOXO1, FOXO3a, AKT, sirtuin1 (SIRT1), and nuclear factor (erythroid-derived2)-like2 (NRF2), investigated by siRNA knockdown, showed differential patterns of signal pathways involved and a role of NRF2 in the inhibition of gluconeogenic enzyme expression. We conclude that these flavones show an antidiabetic potential due to reduction of gluconeogenic and lipogenic capacity despite inhibition of the PKB/AKT pathway which justifies detailed investigation in vivo. PMID:25136826

Gene expression profiles in Parkinson disease prefrontal cortex implicate FOXO1 and genes under its transcriptional regulation.

PubMed

Dumitriu, Alexandra; Latourelle, Jeanne C; Hadzi, Tiffany C; Pankratz, Nathan; Garza, Dan; Miller, John P; Vance, Jeffery M; Foroud, Tatiana; Beach, Thomas G; Myers, Richard H

2012-06-01

Parkinson disease (PD) is a complex neurodegenerative disorder with largely unknown genetic mechanisms. While the degeneration of dopaminergic neurons in PD mainly takes place in the substantia nigra pars compacta (SN) region, other brain areas, including the prefrontal cortex, develop Lewy bodies, the neuropathological hallmark of PD. We generated and analyzed expression data from the prefrontal cortex Brodmann Area 9 (BA9) of 27 PD and 26 control samples using the 44K One-Color Agilent 60-mer Whole Human Genome Microarray. All samples were male, without significant Alzheimer disease pathology and with extensive pathological annotation available. 507 of the 39,122 analyzed expression probes were different between PD and control samples at false discovery rate (FDR) of 5%. One of the genes with significantly increased expression in PD was the forkhead box O1 (FOXO1) transcription factor. Notably, genes carrying the FoxO1 binding site were significantly enriched in the FDR-significant group of genes (177 genes covered by 189 probes), suggesting a role for FoxO1 upstream of the observed expression changes. Single-nucleotide polymorphisms (SNPs) selected from a recent meta-analysis of PD genome-wide association studies (GWAS) were successfully genotyped in 50 out of the 53 microarray brains, allowing a targeted expression-SNP (eSNP) analysis for 52 SNPs associated with PD affection at genome-wide significance and the 189 probes from FoxO1 regulated genes. A significant association was observed between a SNP in the cyclin G associated kinase (GAK) gene and a probe in the spermine oxidase (SMOX) gene. Further examination of the FOXO1 region in a meta-analysis of six available GWAS showed two SNPs significantly associated with age at onset of PD. These results implicate FOXO1 as a PD-relevant gene and warrant further functional analyses of its transcriptional regulatory mechanisms.

Gene Expression Profiles in Parkinson Disease Prefrontal Cortex Implicate FOXO1 and Genes under Its Transcriptional Regulation

PubMed Central

Dumitriu, Alexandra; Latourelle, Jeanne C.; Hadzi, Tiffany C.; Pankratz, Nathan; Garza, Dan; Miller, John P.; Vance, Jeffery M.; Foroud, Tatiana; Beach, Thomas G.; Myers, Richard H.

2012-01-01

Parkinson disease (PD) is a complex neurodegenerative disorder with largely unknown genetic mechanisms. While the degeneration of dopaminergic neurons in PD mainly takes place in the substantia nigra pars compacta (SN) region, other brain areas, including the prefrontal cortex, develop Lewy bodies, the neuropathological hallmark of PD. We generated and analyzed expression data from the prefrontal cortex Brodmann Area 9 (BA9) of 27 PD and 26 control samples using the 44K One-Color Agilent 60-mer Whole Human Genome Microarray. All samples were male, without significant Alzheimer disease pathology and with extensive pathological annotation available. 507 of the 39,122 analyzed expression probes were different between PD and control samples at false discovery rate (FDR) of 5%. One of the genes with significantly increased expression in PD was the forkhead box O1 (FOXO1) transcription factor. Notably, genes carrying the FoxO1 binding site were significantly enriched in the FDR–significant group of genes (177 genes covered by 189 probes), suggesting a role for FoxO1 upstream of the observed expression changes. Single-nucleotide polymorphisms (SNPs) selected from a recent meta-analysis of PD genome-wide association studies (GWAS) were successfully genotyped in 50 out of the 53 microarray brains, allowing a targeted expression–SNP (eSNP) analysis for 52 SNPs associated with PD affection at genome-wide significance and the 189 probes from FoxO1 regulated genes. A significant association was observed between a SNP in the cyclin G associated kinase (GAK) gene and a probe in the spermine oxidase (SMOX) gene. Further examination of the FOXO1 region in a meta-analysis of six available GWAS showed two SNPs significantly associated with age at onset of PD. These results implicate FOXO1 as a PD–relevant gene and warrant further functional analyses of its transcriptional regulatory mechanisms. PMID:22761592

Histone methyltransferase SETDB1 maintains survival of mouse spermatogonial stem/progenitor cells via PTEN/AKT/FOXO1 pathway.

PubMed

Liu, Tiantian; Chen, Xiaoxu; Li, Tianjiao; Li, Xueliang; Lyu, Yinghua; Fan, Xiaoteng; Zhang, Pengfei; Zeng, Wenxian

2017-10-01

Spermatogonial stem cells (SSCs) possess the capacity of self-renewal and differentiation, which are the basis of spermatogenesis. In maintenance of SSC homeostasis, intrinsic/extrinsic factors and various signaling pathways tightly control the fate of SSCs. Methyltransferase SETDB1 (Set domain, bifurcated 1) catalyzes histone H3 lysine 9 (H3K9) trimethylation and represses gene expression. SETDB1 is required for maintaining the survival of spermatogonial stem cells in mice. However, the underlying molecular mechanism remains unclear. In the present study, we found that Setdb1 regulates PTEN/AKT/FOXO1 pathway to inhibit SSC apoptosis. Co-immunoprecipitation and reporter gene assay revealed that SETDB1 interacted and coordinated with AKT to regulate FOXO1 activity and expression of the downstream target genes Bim and Puma. Among the SETDB1-bound genes, the H3K9me3 levels on the promoter regions of Bim and Pten decreased in Setdb1-KD group; in contrast, H3K9me3 status on promoters of Bax and Puma remained unchanged. Therefore, SETDB1 was responsible for regulating the transcription activity of genes in the apoptotic pathway at least in part through modulating H3K9me3. This study replenishes the research on the epigenetic regulation of SSC survival, and provides a new insight for the future study of epigenetic regulation of spermatogenesis. Copyright © 2017 Elsevier B.V. All rights reserved.

FOXO1 is a TXN- and p300-dependent sensor and effector of oxidative stress in diffuse large B-cell lymphomas characterized by increased oxidative metabolism.

PubMed

Sewastianik, T; Szydlowski, M; Jablonska, E; Bialopiotrowicz, E; Kiliszek, P; Gorniak, P; Polak, A; Prochorec-Sobieszek, M; Szumera-Cieckiewicz, A; Kaminski, T S; Markowicz, S; Nowak, E; Grygorowicz, M A; Warzocha, K; Juszczynski, P

2016-11-17

Molecular profiling has led to identification of subtypes of diffuse large B-cell lymphomas (DLBCLs) differing in terms of oncogenic signaling and metabolic programs. The OxPhos-DLBCL subtype is characterized by enhanced mitochondrial oxidative phosphorylation. As increased oxidative metabolism leads to overproduction of potentially toxic reactive oxygen species (ROS), we sought to identify mechanisms responsible for adaptation of OxPhos cells to these conditions. Herein, we describe a mechanism involving the FOXO1-TXN-p300 redox-dependent circuit protecting OxPhos-DLBCL cells from ROS toxicity. We identify a BCL6-dependent transcriptional mechanism leading to relative TXN overexpression in OxPhos cells. We found that OxPhos cells lacking TXN were uniformly more sensitive to ROS and doxorubicin than control cells. Consistent with this, the overall survival of patients with high TXN mRNA expression, treated with doxorubicin-containing regimens, is significantly shorter than of those with low TXN mRNA expression. TXN overexpression curtails p300-mediated FOXO1 acetylation and its nuclear translocation in response to oxidative stress, thus attenuating FOXO1 transcriptional activity toward genes involved in apoptosis and cell cycle inhibition. We also demonstrate that FOXO1 knockdown in cells with silenced TXN expression markedly reduces ROS-induced apoptosis, indicating that FOXO1 is the major sensor and effector of oxidative stress in OxPhos-DLBCLs. These data highlight dynamic, context-dependent modulation of FOXO1 tumor-suppressor functions via acetylation and reveal potentially targetable vulnerabilities in these DLBCLs.

Annexin A11 knockdown inhibits in vitro proliferation and enhances survival of Hca-F cell via Akt2/FoxO1 pathway and MMP-9 expression.

PubMed

Liu, Shuqing; Wang, Jiasheng; Guo, Chunmei; Qi, Houbao; Sun, Ming-Zhong

2015-03-01

Annexin A11 (Anxa11), a Ca(2+)-regulated phospholipid-binding protein, is involved in cell apoptosis, differentiation, vesicle trafficking, cancer progression and autoimmune diseases. Previous study from our group indicated that Anxa11 was associated with lymphatic metastatic potential of murine hepatocarcinoma cells. Herein, we investigated the effects and action mechanism of Anxa11 knockdown on in vitro cell proliferation and apoptosis of Hca-F, a murine hepatocarcinoma cell with∼75% lymph node metastatic potential. Real-time PCR and western blotting assays indicated that Anxa11 was significantly downregulated in monoclonal Anxa11-shRNA-transfected Hca-F cells. Anxa11 knockdown in Hca-F suppressed its in vitro proliferation and cell apoptosis capacities. Following Anxa11 knockdown in Hca-F cells, Bax/Bcl-2 expression level ratio, Akt2 and FoxO1 (pSer319) expression levels as well as MMP-9 mRNA and active MMP-9 protein levels were significantly elevated in Hca-F cells. In conclusion, Annexin A11 knockdown inhibits the in vitro proliferation and cell apoptosis of Hca-F cell via Akt2/FoxO1 and/or MMP-9 expression pathway. Anxa11 might play an important role in hepatocarcinoma cell invasion and metastasis and hepatocarcinoma malignancy. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Concurrent acetylation of FoxO1/3a and p53 due to sirtuins inhibition elicit Bim/PUMA mediated mitochondrial dysfunction and apoptosis in berberine-treated HepG2 cells.

PubMed

Shukla, Shatrunajay; Sharma, Ankita; Pandey, Vivek Kumar; Raisuddin, Sheikh; Kakkar, Poonam

2016-01-15

Post-translational modifications i.e. phosphorylation and acetylation are pivotal requirements for proper functioning of eukaryotic proteins. The current study aimed to decode the impact of acetylation/deacetylation of non-histone targets i.e. FoxO1/3a and p53 of sirtuins (NAD(+) dependent enzymes with lysine deacetylase activity) in berberine treated human hepatoma cells. Berberine (100 μM) inhibited sirtuins significantly (P<0.05) at transcriptional level as well as at translational level. Combination of nicotinamide (sirtuin inhibitor) with berberine potentiated sirtuins inhibition and increased the expression of FoxO1/3a and phosphorylation of p53 tumor suppressor protein. As sirtuins deacetylate non-histone targets including FoxO1/3a and p53, berberine increased the acetylation load of FoxO1/3a and p53 proteins. Acetylated FoxO and p53 proteins transcriptionally activate BH3-only proteins Bim and PUMA (3.89 and 3.87 fold respectively, P<0.001), which are known as direct activator of pro-apoptotic Bcl-2 family protein Bax that culminated into mitochondria mediated activation of apoptotic cascade. Bim/PUMA knock-down showed no changes in sirtuins' expression while cytotoxicity induced by berberine and nicotinamide was curtailed up to 28.3% (P<0.001) and it restored pro/anti apoptotic protein ratio in HepG2 cells. Sirtuins inhibition was accompanied by decline in NAD(+)/NADH ratio, ATP generation, enhanced ROS production and decreased mitochondrial membrane potential. TEM analysis confirmed mitochondrial deterioration and cell damage. SRT-1720 (1-10 μM), a SIRT-1 activator, when pre-treated with berberine (25 μM), reversed sirtuins expression comparable to control and significantly restored the cell viability (P<0.05). Thus, our findings suggest that berberine mediated sirtuins inhibition resulting into FoxO1/3a and p53 acetylation followed by BH3-only protein Bim/PUMA activation may in part be responsible for mitochondria-mediated apoptosis. Copyright © 2015

FOXO3-mediated up-regulation of Bim contributes to rhein-induced cancer cell apoptosis.

PubMed

Wang, Jiao; Liu, Shu; Yin, Yancun; Li, Mingjin; Wang, Bo; Yang, Li; Jiang, Yangfu

2015-03-01

The anthraquinone compound rhein is a natural agent in the traditional Chinese medicine rhubarb. Preclinical studies demonstrate that rhein has anticancer activity. Treatment of a variety of cancer cells with rhein may induce apoptosis. Here, we report that rhein induces atypical unfolded protein response in breast cancer MCF-7 cells and hepatoma HepG2 cells. Rhein induces CHOP expression, eIF2α phosphorylation and caspase cleavage, while it does not induce glucose-regulated protein 78 (GRP78) expression in both MCF-7 and HepG2 cells. Meanwhile, rhein inhibits thapsigargin-induced GRP78 expression and X box-binding protein 1 splicing. In addition, rhein inhibits Akt phosphorylation and stimulates FOXO transactivation activity. Rhein induces Bim expression in MCF-7 and HepG2 cells, which can be abrogated by FOXO3a knockdown. Knockdown of FOXO3a or Bim abrogates rhein-induced caspase cleavage and apoptosis. The chemical chaperone 4-phenylbutyrate acid antagonizes the induction of FOXO activation, Bim expression and caspase cleavage by rhein, indicating that protein misfolding may be involved in triggering these deleterious effects. We conclude that FOXO3a-mediated up-regulation of Bim is a key mechanism underlying rhein-induced cancer cells apoptosis.

Elevated hepatic fatty acid elongase-5 activity corrects dietary fat-induced hyperglycemia in obese BL/6J mice[S

PubMed Central

Tripathy, Sasmita; Torres-Gonzalez, Moises; Jump, Donald B.

2010-01-01

Elevated hepatic fatty acid elongase-5 (Elovl5) activity lowers blood glucose in fasted chow-fed C57BL/6J mice. As high-fat diets induce hyperglycemia and suppress hepatic Elovl5 activity, we tested the hypothesis that elevated hepatic Elovl5 expression attenuates hyperglycemia in high-fat-diet-induced obese mice. Increasing hepatic Elovl5 activity by a recombinant adenoviral approach restored blood glucose and insulin, HOMA-IR, and glucose tolerance to normal values in obese mice. Elevated Elovl5 activity increased hepatic content of Elovl5 products (20:3,n-6, 22:4,n-6) and suppressed levels of enzymes (Pck1, G6Pc) and transcription factors (FoxO1 and PGC1α, but not CRTC2) involved in gluconeogenesis. Effects of Elovl5 on FoxO1 nuclear abundance correlated with increased phosphorylation of FoxO1, Akt, and the catalytic unit of PP2A, as well as a decline in cellular abundance of TRB3. Such changes are mechanistically linked to the regulation of FoxO1 nuclear abundance and gluconeogenesis. These results show that Elovl5 activity impacts the hepatic abundance and phosphorylation status of multiple proteins involved in gluconeogenesis. Our findings establish a link between fatty acid elongation and hepatic glucose metabolism and suggest a role for regulators of Elovl5 activity in the treatment of diet-induced hyperglycemia. PMID:20488798

AMP-activated protein kinase couples 3-bromopyruvate-induced energy depletion to apoptosis via activation of FoxO3a and upregulation of proapoptotic Bcl-2 proteins.

PubMed

Bodur, Cagri; Karakas, Bahriye; Timucin, Ahmet Can; Tezil, Tugsan; Basaga, Huveyda

2016-11-01

Most tumors primarily rely on glycolysis rather than mitochondrial respiration for ATP production. This phenomenon, also known as Warburg effect, renders tumors more sensitive to glycolytic disturbances compared to normal cells. 3-bromopyruvate is a potent inhibitor of glycolysis that shows promise as an anticancer drug candidate. Although investigations revealed that 3-BP triggers apoptosis through ATP depletion and subsequent AMPK activation, the underlying molecular mechanisms coupling AMPK to apoptosis are poorly understood. We showed that 3-BP leads to a rapid ATP depletion which was followed by growth inhibition and Bax-dependent apoptosis in HCT116 cells. Apoptosis was accompanied with activation of caspase-9 and -3 while pretreatment with a general caspase inhibitor attenuated cell death. AMPK, p38, JNK, and Akt were phosphorylated immediately upon treatment. Pharmacological inhibition and silencing of AMPK largely inhibited 3-BP-induced apoptosis and reversed phosphorylation of JNK. Transcriptional activity of FoxO3a was dramatically increased subsequent to AMPK-mediated phosphorylation of FoxO3a at Ser413. Cell death analysis of cells transiently transfected with wt or AMPK-phosphorylation-deficient FoxO3 expression plasmids verified the contributory role of AMPK-FoxO3a axis in 3-BP-induced apoptosis. In addition, expression of proapoptotic Bcl-2 proteins Bim and Bax were upregulated in an AMPK-dependent manner. Bim was transcriptionally activated in association with FoxO3a activity, while Bax upregulation was abolished in p53-null cells. Together, these data suggest that AMPK couples 3-BP-induced metabolic disruption to intrinsic apoptosis via modulation of FoxO3a-Bim axis and Bax expression. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

GxE Interactions between FOXO Genotypes and Tea Drinking Are Significantly Associated with Cognitive Disability at Advanced Ages in China

PubMed Central

Zeng, Yi; Chen, Huashuai; Ni, Ting; Ruan, Rongping; Feng, Lei; Nie, Chao; Cheng, Lingguo; Li, Yang; Tao, Wei; Gu, Jun; Land, Kenneth C.; Yashin, Anatoli; Tan, Qihua; Yang, Ze; Bolund, Lars; Yang, Huanming; Hauser, Elizabeth; Willcox, D. Craig; Willcox, Bradley J.; Tian, Xiao-Li; Vaupel, James W.

2015-01-01

Logistic regression analysis based on data from 822 Han Chinese oldest old aged 92+ demonstrated that interactions between carrying FOXO1A-266 or FOXO3-310 or FOXO3-292 and tea drinking at around age 60 or at present time were significantly associated with lower risk of cognitive disability at advanced ages. Associations between tea drinking and reduced cognitive disability were much stronger among carriers of the genotypes of FOXO1A-266 or FOXO3-310 or FOXO3-292 compared with noncarriers, and it was reconfirmed by analysis of three-way interactions across FOXO genotypes, tea drinking at around age 60, and at present time. Based on prior findings from animal and human cell models, we postulate that intake of tea compounds may activate FOXO gene expression, which in turn may positively affect cognitive function in the oldest old population. Our empirical findings imply that the health benefits of particular nutritional interventions, including tea drinking, may, in part, depend upon individual genetic profiles. PMID:24895270

The FOXO1 Gene-Obesity Interaction Increases the Risk of Type 2 Diabetes Mellitus in a Chinese Han Population.

PubMed

Gong, Lilin; Li, Rong; Ren, Wei; Wang, Zengchan; Wang, Zhihong; Yang, Maosheng; Zhang, Suhua

2017-02-01

Here, we aimed to study the effect of the forkhead box O1-insulin receptor substrate 2 (FOXO1-IRS2) gene interaction and the FOXO1 and IRS2 genes-environment interaction for the risk of type 2 diabetes mellitus (T2DM) in a Chinese Han population. We genotyped 7 polymorphism sites of FOXO1 gene and IRS2 gene in 780 unrelated Chinese Han people (474 cases of T2DM, 306 cases of healthy control). The risk of T2DM in individuals with AA genotype for rs7986407 and CC genotype for rs4581585 in FOXO1 gene was 2.092 and 2.57 times higher than that with GG genotype (odds ratio [OR] = 2.092; 95% confidence interval [CI] = 1.178-3.731; P = 0.011) and TT genotype (OR = 2.571; 95% CI = 1.404-4.695; P = 0.002), respectively. The risk of T2DM in individuals with GG genotype for Gly1057Asp in IRS2 gene was 1.42 times higher than that with AA genotype (OR = 1.422; 95% CI = 1.037-1.949; P = 0.029). The other 4 single nucleotide polymorphisms (SNPs) had no significant association with T2DM (P > 0.05). Multifactor dimensionality reduction (MDR) analysis showed that the interaction between SNPs rs7986407 and rs4325426 in FOXO1 gene and waist was the best model confirmed by interaction analysis, closely associating with T2DM. There was an increased risk for T2DM in the case of non-obesity with genotype combined AA/CC, AA/AC or AG/AA for rs7986407 and rs4325426, and obesity with genotype AA for rs7986407 or AA for rs4325426 (OR = 3.976; 95% CI = 1.156-13.675; P value from sign test [P(sign)] = 0.025; P value from permutation test [P(perm)] = 0.000-0.001). Together, this study indicates an association of FOXO1 and IRS2 gene polymorphisms with T2DM in Chinese Han population, supporting FOXO1-obesity interaction as a key factor for the risk of T2DM.

Intersection of FOXO- and RUNX1-mediated gene expression programs in single breast epithelial cells during morphogenesis and tumor progression.

PubMed

Wang, Lixin; Brugge, Joan S; Janes, Kevin A

2011-10-04

Gene expression networks are complicated by the assortment of regulatory factors that bind DNA and modulate transcription combinatorially. Single-cell measurements can reveal biological mechanisms hidden by population averages, but their value has not been fully explored in the context of mRNA regulation. Here, we adapted a single-cell expression profiling technique to examine the gene expression program downstream of Forkhead box O (FOXO) transcription factors during 3D breast epithelial acinar morphogenesis. By analyzing patterns of mRNA fluctuations among individual matrix-attached epithelial cells, we found that a subset of FOXO target genes was jointly regulated by the transcription factor Runt-related transcription factor 1 (RUNX1). Knockdown of RUNX1 causes hyperproliferation and abnormal morphogenesis, both of which require normal FOXO function. Down-regulating RUNX1 and FOXOs simultaneously causes widespread oxidative stress, which arrests proliferation and restores normal acinar morphology. In hormone-negative breast cancers lacking human epidermal growth factor receptor 2 (HER2) amplification, we find that RUNX1 down-regulation is strongly associated with up-regulation of FOXO1, which may be required to support growth of RUNX1-negative tumors. The coordinate function of these two tumor suppressors may provide a failsafe mechanism that inhibits cancer progression.

The transcription factor Prep1 controls hepatic insulin sensitivity and gluconeogenesis by targeting nuclear localization of FOXO1

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

Kulebyakin, Konstantin; Penkov, Dmitry; IFOM – the FIRC Institute of Molecular Oncology, Via Adamello 16, Milan, 20139

Liver plays a key role in controlling body carbohydrate homeostasis by switching between accumulation and production of glucose and this way maintaining constant level of glucose in blood. Increased blood glucose level triggers release of insulin from pancreatic β-cells. Insulin represses hepatic glucose production and increases glucose accumulation. Insulin resistance is the main cause of type 2 diabetes and hyperglycemia. Currently thiazolidinediones (TZDs) targeting transcriptional factor PPARγ are used as insulin sensitizers for treating patients with type 2 diabetes. However, TZDs are reported to be associated with cardiovascular and liver problems and stimulate obesity. Thus, it is necessary to searchmore » new approaches to improve insulin sensitivity. A promising candidate is transcriptional factor Prep1, as it was shown earlier it could affect insulin sensitivity in variety of insulin-sensitive tissues. The aim of the present study was to evaluate a possible involvement of transcriptional factor Prep1 in control of hepatic glucose accumulation and production. We created mice with liver-specific Prep1 knockout and discovered that hepatocytes derived from these mice are much more sensitive to insulin, comparing to their WT littermates. Incubation of these cells with 100 nM insulin results in almost complete inhibition of gluconeogenesis, while in WT cells this repression is only partial. However, Prep1 doesn't affect gluconeogenesis in the absence of insulin. Also, we observed that nuclear content of gluconeogenic transcription factor FOXO1 was greatly reduced in Prep1 knockout hepatocytes. These findings suggest that Prep1 may control hepatic insulin sensitivity by targeting FOXO1 nuclear stability. - Highlights: • A novel model of liver-specific Prep1 knockout is established. • Ablation of Prep1 in hepatocytes increases insulin sensitivity. • Prep1 controls hepatic insulin sensitivity by regulating localization of FOXO1. • Prep1 regulates

Molecular Insights into SIRT1 Protection Against UVB-Induced Skin Fibroblast Senescence by Suppression of Oxidative Stress and p53 Acetylation.

PubMed

Chung, Ki Wung; Choi, Yeon Ja; Park, Min Hi; Jang, Eun Ji; Kim, Dae Hyun; Park, Byung Hyun; Yu, Byung Pal; Chung, Hae Young

2015-08-01

Stresses, such as exposure to ultraviolet radiation and those associated with aging, are known to cause premature cellular senescence that is characterized by growth arrest and morphological and gene expression changes. This study was designed to investigate the protective effect of Sirtuin1 (SIRT1) on the UVB-induced premature senescence. Under in vitro experimental conditions, exposure to a subcytotoxic dose of UVB enhanced human skin fibroblasts senescence, as characterized by increased β-galactosidase activity and increased levels of senescence-associated proteins. However, adenovirus-mediated SIRT1 overexpression significantly protected fibroblasts from UVB-induced cellular deterioration. Exposure to UVB-induced cell senescence was associated with oxidative stress and p38 mitogen-activated protein kinase activation. Molecular analysis demonstrated that deacetylation of Forkhead box O3α (FOXO3α) by SIRT1 changed the transcriptional activity of FOXO3α and increased resistance to the oxidative stress. In addition, SIRT1 suppressed UVB-induced p53 acetylation and its transcriptional activity, which directly affected the cell cycle arrest induced by UVB. Further study demonstrated that SIRT1 activation inhibited cell senescence in the skin of the HR1 hairless mouse exposed to UVB. The study identifies a new role for SIRT1 in the UVB-induced senescence of skin fibroblats and provides a potential target for skin protection through molecuar insights into the mechanisms responsible for UVB-induced photoaging. © The Author 2014. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Advanced glycosylation end product promotes forkhead box O1 and inhibits Wnt pathway to suppress capacities of epidermal stem cells.

PubMed

Zhu, Jie; Wang, Peng; Yu, Zhimin; Lai, Wei; Cao, Yi; Huang, Pinbo; Xu, Qiaodong; Yu, Menglei; Xu, Junyao; Huang, Zitong; Zeng, Bing

2016-01-01

Diabetes mellitus is frequently accompanied by chronic complications like delayed wound healing, which is consider to be attributed to the accumulation of advanced glycosylation end product (AGE). However, the impacts of AGE on epidermal stem cells (ESCs) are largely unknown. This study aims to address the influence and mechanism of AGE on ESCs. ESCs isolated from rats were cultured in AGE-modified bovine serum albumin and transfected with small interfering RNA to knock down AGE-specific receptor (AGER). Expression of stem cell markers integrin β1 (ITGB1) and keratin 19 (KRT19), cell viability, apoptosis and reactive oxygen species (ROS) were examined. Wnt pathway-related factors Wnt family member 1 (WNT1), WNT3A, β-catenin, v-myc avian myelocytomatosis viral oncogene homolog (MYC), cyclin D1 (CCND1) and matrix metallopeptidase 7 (MMP7) were quantified. The interaction between forkhead box O1 (FOXO1) and β-catenin was assessed by co-immunoprecipitation. Results indicated that AGE down-regulated ITGB1 and KRT19 expression, suppressed ESC viability and promoted apoptosis, and ROS level ( P < 0.01), implying decreased capacities of ESCs. AGE also promoted AGER and FOXO1, while AGER knockdown had the opposite effects. Moreover, AGER knockdown elevated the level of WNT1, WNT3A, MYC, CCND1 and MMP7 that were suppressed by AGE ( P < 0.01). Immunoprecipitation analysis showed that FOXO1 could compete with lymphoid enhancer binding factor 1 to interact with β-catenin, which might help to elucidate the mechanism of AGE repressing ESCs. This study helps to understand the mechanism of accumulated AGE in affecting ESC capacities, and provides potential therapeutic targets to meliorate diabetic wound healing.

Assessing FOXO1A as a potential susceptibility locus for type 2 diabetes and obesity in American Indians.

PubMed

Muller, Yunhua L; Hanson, Robert L; Wiessner, Gregory; Nieboer, Lori; Kobes, Sayuko; Piaggi, Paolo; Abdussamad, Mahdi; Okani, Chidinma; Knowler, William C; Bogardus, Clifton; Baier, Leslie J

2015-10-01

A prior genome-wide association study (GWAS) in Pima Indians identified variation within FOXO1A that modestly associated with early-onset (onset age < 25 years) type 2 diabetes (T2D). FOXO1A encodes the forkhead transcription factor involved in pancreatic β-cell growth and hypothalamic energy balance; therefore, FOXO1A was analyzed as a candidate gene for T2D and obesity in a population-based sample of 7,710 American Indians. Tag SNPs in/near FOXO1A (minor allele frequency ≥ 0.05) were analyzed for association with T2D at early onset (n = 1,060) and all ages (n = 7,710) and with insulin secretion (n = 298). SNPs were also analyzed for association with maximum body mass index (BMI) in adulthood (n = 5,918), maximum BMI z-score in childhood (n = 5,350), and % body fat (n = 555). An intronic SNP rs2297627 associated with early-onset T2D [OR = 1.34 (1.13-1.58), P = 8.7 × 10(-4)] and T2D onset at any age [OR = 1.19 (1.09-1.30), P = 1 × 10(-4) ]. The T2D risk allele also associated with lower acute insulin secretion (β = 0.88, as a multiplier, P = 0.02). Another intronic SNP (rs1334241, D' = 0.99, r(2) = 0.49 with rs2297627) associated with maximum adulthood BMI (β = 1.02, as a multiplier, P = 3 × 10(-5)), maximum childhood BMI z-score (β = 0.08, P = 3 × 10(-4)), and % body fat (β = 0.83%, P = 0.04). Common variation in FOXO1A may modestly affect risk for T2D and obesity in American Indians. © 2015 The Obesity Society.

FOXO3a is broadly neuroprotective in vitro and in vivo against insults implicated in motor neuron diseases

PubMed Central

Mojsilovic-Petrovic, Jelena; Nedelsky, Natalia; Boccitto, Marco; Mano, Itzhak; Georgiades, Savvas N.; Zhou, Weiguo; Liu, Yuhong; Neve, Rachael L.; Taylor, J. Paul; Driscoll, Monica; Clardy, Jon; Merry, Diane; Kalb, Robert G.

2009-01-01

Aging is a risk factor for the development of adult-onset neuro-degenerative diseases. While some of the molecular pathways regulating longevity and stress resistance in lower organisms are defined (i.e., those activating the transcriptional regulators DAF-16 and HSF-1 in C. elegans), their relevance to mammals and disease susceptibility are unknown. We studied the signaling controlled by the mammalian homolog of DAF-16, FOXO3a, in model systems of motor neuron disease. Neuron death elicited in vitro by excitotoxic insult or the expression of mutant SOD1, mutant p150glued or polyQ expanded androgen receptor was abrogated by expression of nuclear-targeted FOXO3a. We identify a compound (Psammaplysene A, PA) that increases nuclear localization of FOXO3a in vitro and in vivo and show that PA also protects against these insults in vitro. Administration of PA to invertebrate model systems of neurodegeneration similarly blocked neuron death in a DAF-16/FOXO3a-dependent manner. These results indicate that activation of the DAF-16/FOXO3a pathway, genetically or pharmacologically, confers protection against the known causes of motor neuron diseases. PMID:19553463

The role of FOXO and PPAR transcription factors in diet-mediated inhibition of PDC activation and carbohydrate oxidation during exercise in humans and the role of pharmacological activation of PDC in overriding these changes.

PubMed

Constantin-Teodosiu, Dumitru; Constantin, Despina; Stephens, Francis; Laithwaite, David; Greenhaff, Paul L

2012-05-01

High-fat feeding inhibits pyruvate dehydrogenase complex (PDC)-controlled carbohydrate (CHO) oxidation, which contributes to muscle insulin resistance. We aimed to reveal molecular changes underpinning this process in resting and exercising humans. We also tested whether pharmacological activation of PDC overrides these diet-induced changes. Healthy males consumed a control diet (CD) and on two further occasions an isocaloric high-fat diet (HFD). After each diet, subjects cycled for 60 min after intravenous infusion with saline (CD and HFD) or dichloroacetate (HFD+DCA). Quadriceps muscle biopsies obtained before and after 10 and 60 min of exercise were used to estimate CHO use, PDC activation, and mRNAs associated with insulin, fat, and CHO signaling. Compared with CD, HFD increased resting pyruvate dehydrogenase kinase 2 (PDK2), PDK4, forkhead box class O transcription factor 1 (FOXO1), and peroxisome proliferator-activated receptor transcription factor α (PPARα) mRNA and reduced PDC activation. Exercise increased PDC activation and whole-body CHO use in HFD, but to a lower extent than in CD. Meanwhile PDK4 and FOXO1, but not PPARα or PDK2, mRNA remained elevated. HFD+DCA activated PDC throughout and restored whole-body CHO use during exercise. FOXO1 appears to play a role in HFD-mediated muscle PDK4 upregulation and inhibition of PDC and CHO oxidation in humans. Also, pharmacological activation of PDC restores HFD-mediated inhibition of CHO oxidation during exercise.

Esculetin ameliorates hepatic fibrosis in high fat diet induced non-alcoholic fatty liver disease by regulation of FoxO1 mediated pathway.

PubMed

Pandey, Anuradha; Raj, Priyank; Goru, Santosh Kumar; Kadakol, Almesh; Malek, Vajir; Sharma, Nisha; Gaikwad, Anil Bhanudas

2017-08-01

Non-alcoholic fatty liver disease (NAFLD), a chronic metabolic disorder is associated with oxidative stress, inflammation and fibrotic cascades. In this study, we aimed to examine the effects of Esculetin, a well-known anti-oxidant on TGF-β1 mediated liver fibrosis and FoxO1 activity. A non-genetic murine model for NAFLD was developed by chronic high fat diet (HFD) (58% calories from fats) feeding in Wistar rats. The plasma biochemical parameters, liver function tests, oxidative stress, and histopathological alterations were assessed. The alterations in extracellular matrix (ECM) deposition and FoxO1 activity were assessed by immunohistochemistry. The aberrations in plasma parameters, liver functioning, morphometric and microscopic changes in liver structure of HFD fed rats were significantly improved by treatment with Esculetin. Liver fibrosis, identified in the form of collagen deposition and expression of fibrotic proteins like TGF-β1 and fibronectin was also markedly controlled by Esculetin. The expression of phospho-FoxO1 was found to be reduced in HFD fed rats' liver, showing an increase in activation of FoxO1 under insulin resistant and hyperglycemic states. Esculetin treatment could improve phospho-FoxO1 expression, thus showing its ability to act on Akt/PI3K/FoxO1 pathway. As per the previous studies, a potential therapy for NAFLD may be the one with multi-faceted actions on insulin resistance, oxidative stress, inflammation and fibrosis. This study demonstrates the efficiency of Esculetin in improving liver fibrosis in HFD induced NAFLD. Copyright © 2017 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

Linking Alzheimer's disease to insulin resistance: the FoxO response to oxidative stress.

PubMed

Manolopoulos, K N; Klotz, L-O; Korsten, P; Bornstein, S R; Barthel, A

2010-11-01

Oxidative stress is an important determinant not only in the pathogenesis of Alzheimer's disease (AD), but also in insulin resistance (InsRes) and diabetic complications. Forkhead box class O (FoxO) transcription factors are involved in both insulin action and the cellular response to oxidative stress, thereby providing a potential integrative link between AD and InsRes. For example, the expression of intra- and extracellular antioxidant enzymes, such as manganese-superoxide dismutase and selenoprotein P, is regulated by FoxO proteins, as is the expression of important hepatic enzymes of gluconeogenesis. Here, we review the molecular mechanisms involved in the pathogenesis of AD and InsRes and discuss the function of FoxO proteins in these processes. Both InsRes and oxidative stress may promote the transcriptional activity of FoxO proteins, resulting in hyperglycaemia and a further increased production of reactive oxygen species (ROS). The consecutive activation of c-Jun N-terminal kinases and inhibition of Wingless (Wnt) signalling may result in the formation of β-amyloid plaques and τ protein phosphorylation. Wnt inhibition may also result in a sustained activation of FoxO proteins with induction of apoptosis and neuronal loss, thereby completing a vicious circle from oxidative stress, InsRes and hyperglycaemia back to the formation of ROS and consecutive neurodegeneration. In view of their central function in this model, FoxO proteins may provide a potential molecular target for the treatment of both InsRes and AD.

Regulation of neuroblastoma differentiation by forkhead transcription factors FOXO1/3/4 through the receptor tyrosine kinase PDGFRA

PubMed Central

Mei, Yang; Wang, Zhanxiang; Zhang, Lei; Zhang, Yiru; Li, Xiaoyu; Liu, Huihui; Ye, Jing; You, Han

2012-01-01

Neuroblastoma is a common childhood malignant tumor originated from the neural crest-derived sympathetic nervous system. A crucial early event in neuroblastoma pathogenesis is arrested differentiation of neuroblasts at various stages. Treatment of neuroblastoma with TPA and PDGF-BB leads to terminal differentiation of neuroblastoma cells. However, the signaling pathways that are involved in this process remain largely unknown. Here, we report that inhibition of endogenous FOXO proteins attenuated TPA/PDGF-BB mediated differentiation of neuroblastoma cells. Activated FOXO transcription factors acted on PDGFRA promoter to direct its basal mRNA expression as well as its induction upon serum deprivation. Depletion of endogenous PDGFRA in neuroblastoma cells significantly diminished neurite formation and extension under TPA/PDGF-BB treatment. Furthermore, ectopic expression of PDGFRA abolished the blockage of neuroblastoma differentiation by FOXOs inhibition. These findings define the FOXO–PDGFRA axis as crucial mechanistic components that govern TPA-induced neuroblastoma differentiation. PMID:22411791

Chronic resistance training activates autophagy and reduces apoptosis of muscle cells by modulating IGF-1 and its receptors, Akt/mTOR and Akt/FOXO3a signaling in aged rats.

PubMed

Luo, Li; Lu, A-Ming; Wang, Yan; Hong, An; Chen, Yulan; Hu, Juan; Li, Xiaoning; Qin, Zheng-Hong

2013-04-01

Resistance exercise training (RET) remains the most effective treatment for the loss of muscle mass and strength in elderly people. However, the underlying cellular and molecular mechanisms are not well understood. Recent evidence suggests that autophagic signaling is altered in aged skeletal muscles. This study aimed to investigate if RET affects IGF-1 and its receptors, the Akt/mTOR, and Akt/FOXO3a signaling pathways and regulates autophagy and apoptosis in the gastrocnemius muscles of 18-20 month old rats. The results showed that 9 weeks of RET prevented the loss of muscle mass and improved muscle strength, accompanied by reduced LC3-II/LC3-I ratio, reduced p62 protein levels, and increased levels of autophagy regulatory proteins, including Beclin 1, Atg5/12, Atg7, and the lysosomal enzyme cathepsin L. RET also reduced cytochrome c level in the cytosol but increased its level in mitochondrial fraction, and inhibited cleaved caspase 3 production and apoptosis. Furthermore, RET upregulated the expression of IGF-1 and its receptors but downregulated the phosphorylation of Akt and mTOR. In addition, RET upregulated the expression of total AMPK, phosphorylated AMPK, and FOXO3a. Taken together, these results suggest that the benefits of RET are associated with increased autophagy activity and reduced apoptosis of muscle cells by modulating IGF-1 and its receptors, the Akt/mTOR and Akt/FOXO3a signaling pathways in aged skeletal muscles. Copyright © 2013 Elsevier Inc. All rights reserved.

GxE interactions between FOXO genotypes and drinking tea are significantly associated with prevention of cognitive decline in advanced age in China.

PubMed

Zeng, Yi; Chen, Huashuai; Ni, Ting; Ruan, Rongping; Feng, Lei; Nie, Chao; Cheng, Lingguo; Li, Yang; Tao, Wei; Gu, Jun; Land, Kenneth C; Yashin, Anatoli; Tan, Qihua; Yang, Ze; Bolund, Lars; Yang, Huanming; Hauser, Elizabeth; Willcox, D Craig; Willcox, Bradley J; Tian, Xiao-Li; Vaupel, James W

2015-04-01

Logistic regression analysis based on data from 822 Han Chinese oldest old aged 92+ demonstrated that interactions between carrying FOXO1A-266 or FOXO3-310 or FOXO3-292 and tea drinking at around age 60 or at present time were significantly associated with lower risk of cognitive disability at advanced ages. Associations between tea drinking and reduced cognitive disability were much stronger among carriers of the genotypes of FOXO1A-266 or FOXO3-310 or FOXO3-292 compared with noncarriers, and it was reconfirmed by analysis of three-way interactions across FOXO genotypes, tea drinking at around age 60, and at present time. Based on prior findings from animal and human cell models, we postulate that intake of tea compounds may activate FOXO gene expression, which in turn may positively affect cognitive function in the oldest old population. Our empirical findings imply that the health benefits of particular nutritional interventions, including tea drinking, may, in part, depend upon individual genetic profiles. © The Author 2014. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Inactivation of the FoxO3a transcription factor is associated with the production of reactive oxygen species during protein kinase CK2 downregulation-mediated senescence in human colon cancer and breast cancer cells.

PubMed

Park, Seong-Yeol; Bae, Young-Seuk

2016-09-09

We previously showed that protein kinase CK2 downregulation mediates senescence through the reactive oxygen species (ROS)-p53-p21(Cip1/WAF1) pathway in various human cells. In the present study, we investigated whether the FoxO3a transcription factor is associated with ROS production during CK2 downregulation-induced senescence in human colon cancer HCT116 and breast cancer MCF-7 cells. FoxO3a overexpression suppressed ROS production and p53 stabilization induced by a CK2α knockdown. CK2α downregulation induced nuclear export of FoxO3a through stimulation of AKT-mediated phosphorylation of FoxO3a and decreased transcription of its target genes (Cu/ZnSOD, MnSOD, and catalase). In contrast, CK2α overexpression inhibited AKT-mediated FoxO3a phosphorylation. This resulted in nuclear accumulation of FoxO3a, and elevated expression of its target genes. Therefore, these data indicate for the first time that CK2 downregulation stimulates ROS generation by inhibiting FoxO3a during premature senescence in human colon and breast cancer cells. Copyright © 2016 Elsevier Inc. All rights reserved.

Lower FOXO3 mRNA expression in granulosa cells is involved in unexplained infertility.

PubMed

Yamamoto, Hikaru; Yamashita, Yoshiki; Saito, Natsuho; Hayashi, Atsushi; Hayashi, Masami; Terai, Yoshito; Ohmichi, Masahide

2017-06-01

The aim of this study was to investigate whether FOXO1 and FOXO3 mRNA expression in granulosa cells is the cause of unexplained infertility. Thirty-one patients aged <40 years (13 with unexplained infertility and 18 with male partner infertility as a control group) whose serum anti-Müllerian hormone level was >0.5 ng/μL were enrolled in the study. All patients underwent oocyte retrieval under a short protocol from June 2012 to October 2013. Real-time PCR was carried out using mRNA extracted from granulosa cells retrieved from mature follicles. We compared FOXO1 and FOXO3 mRNA expression ratios in granulosa cells between the unexplained infertility group and the male infertility group. The relation between FOXO1 and FOXO3 mRNA expression ratios in granulosa cells and assisted reproduction technology clinical outcome was also examined. FOXO3 mRNA expression ratio was significantly lower in the unexplained infertility group than in the male infertility group. Moreover, FOXO3 mRNA expression ratio showed a positive correlation with both the number of retrieved oocytes and serum anti-Müllerian hormone level. A positive correlation was also identified between FOXO1 mRNA expression and total dose of hMG. As well, the number of retrieved oocytes in the unexplained infertility group was statistically lower than that in the male infertility group. A lower FOXO3 mRNA expression in granulosa cells leads to poor oocyte development in patients with unexplained infertility undergoing controlled ovarian stimulation for in vitro fertilization-embryo transfer. © 2017 Japan Society of Obstetrics and Gynecology.

FOXO3 Modulates Endothelial Gene Expression and Function by Classical and Alternative Mechanisms*

PubMed Central

Czymai, Tobias; Viemann, Dorothee; Sticht, Carsten; Molema, Grietje; Goebeler, Matthias; Schmidt, Marc

2010-01-01

FOXO transcription factors represent targets of the phosphatidylinositol 3-kinase/protein kinase B survival pathway controlling important biological processes, such as cell cycle progression, apoptosis, vascular remodeling, stress responses, and metabolism. Recent studies suggested the existence of alternative mechanisms of FOXO-dependent gene expression beyond classical binding to a FOXO-responsive DNA-binding element (FRE). Here we analyzed the relative contribution of those mechanisms to vascular function by comparing the transcriptional and cellular responses to conditional activation of FOXO3 and a corresponding FRE-binding mutant in human primary endothelial cells. We demonstrate that FOXO3 controls expression of vascular remodeling genes in an FRE-dependent manner. In contrast, FOXO3-induced cell cycle arrest and apoptosis occurs independently of FRE binding, albeit FRE-dependent gene expression augments the proapoptotic response. These findings are supported by bioinformatical analysis, which revealed a statistical overrepresentation of cell cycle regulators and apoptosis-related genes in the group of co-regulated genes. Molecular analysis of FOXO3-induced endothelial apoptosis excluded modulators of the extrinsic death receptor pathway and demonstrated important roles for the BCL-2 family members BIM and NOXA in this process. Although NOXA essentially contributed to FRE-dependent apoptosis, BIM was effectively induced in the absence of FRE-binding, and small interfering RNA-mediated BIM depletion could rescue apoptosis induced by both FOXO3 mutants. These data suggest BIM as a critical cell type-specific mediator of FOXO3-induced endothelial apoptosis, whereas NOXA functions as an amplifying factor. Our study provides the first comprehensive analysis of alternatively regulated FOXO3 targets in relevant primary cells and underscores the importance of such genes for endothelial function and integrity. PMID:20123982

miR-182 attenuates atrophy-related gene expression by targeting FoxO3 in skeletal muscle

PubMed Central

Rahnert, Jill A.; Zheng, Bin; Woodworth-Hobbs, Myra E.; Franch, Harold A.; Russ Price, S.

2014-01-01

Skeletal muscle atrophy occurs in response to a variety of conditions including chronic kidney disease, diabetes, cancer, and elevated glucocorticoids. MicroRNAs (miR) may play a role in the wasting process. Activation of the forkhead box O3 (FoxO3) transcription factor causes skeletal muscle atrophy in patients, animals, and cultured cells by increasing the expression of components of the ubiquitin-proteasome and autophagy-lysosome proteolytic systems. To identify microRNAs that potentially modulate the atrophy process, an in silico target analysis was performed and miR-182 was predicted to target FoxO3 mRNA. Using a combination of immunoblot analysis, quantitative real-time RT-PCR, and FoxO3 3′-UTR luciferase reporter genes, miR-182 was confirmed to regulate FoxO3 expression in C2C12 myotubes. Transfection of miR-182 into muscle cells decreased FoxO3 mRNA 30% and FoxO3 protein 67% (P < 0.05) and also prevented a glucocorticoid-induced upregulation of multiple FoxO3 gene targets including MAFbx/atrogin-1, autophagy-related protein 12 (ATG12), cathepsin L, and microtubule-associated protein light chain 3 (LC3). Treatment of C2C12 myotubes with dexamethasone (Dex) (1 μM, 6 h) to induce muscle atrophy decreased miR-182 expression by 63% (P < 0.05). Similarly, miR-182 was decreased 44% (P < 0.05) in the gastrocnemius muscle of rats injected with streptozotocin to induce diabetes compared with controls. Finally, miR-182 was present in exosomes isolated from the media of C2C12 myotubes and Dex increased its abundance. These data identify miR-182 as an important regulator of FoxO3 expression that participates in the control of atrophy-inducing genes during catabolic diseases. PMID:24871856

IGF-1 prevents ANG II-induced skeletal muscle atrophy via Akt- and Foxo-dependent inhibition of the ubiquitin ligase atrogin-1 expression

PubMed Central

Yoshida, Tadashi; Semprun-Prieto, Laura; Sukhanov, Sergiy

2010-01-01

Congestive heart failure is associated with activation of the renin-angiotensin system and skeletal muscle wasting. Angiotensin II (ANG II) has been shown to increase muscle proteolysis and decrease circulating and skeletal muscle IGF-1. We have shown previously that skeletal muscle-specific overexpression of IGF-1 prevents proteolysis and apoptosis induced by ANG II. These findings indicated that downregulation of IGF-1 signaling in skeletal muscle played an important role in the wasting effect of ANG II. However, the signaling pathways and mechanisms whereby IGF-1 prevents ANG II-induced skeletal muscle atrophy are unknown. Here we show ANG II-induced transcriptional regulation of two ubiquitin ligases atrogin-1 and muscle ring finger-1 (MuRF-1) that precedes the reduction of skeletal muscle IGF-1 expression, suggesting that activation of atrogin-1 and MuRF-1 is an initial mechanism leading to skeletal muscle atrophy in response to ANG II. IGF-1 overexpression in skeletal muscle prevented ANG II-induced skeletal muscle wasting and the expression of atrogin-1, but not MuRF-1. Dominant-negative Akt and constitutively active Foxo-1 blocked the ability of IGF-1 to prevent ANG II-mediated upregulation of atrogin-1 and skeletal muscle wasting. Our findings demonstrate that the ability of IGF-1 to prevent ANG II-induced skeletal muscle wasting is mediated via an Akt- and Foxo-1-dependent signaling pathway that results in inhibition of atrogin-1 but not MuRF-1 expression. These data suggest strongly that atrogin-1 plays a critical role in mechanisms of ANG II-induced wasting in vivo. PMID:20228261

Oligonol, a Low-Molecular Weight Polyphenol Derived from Lychee, Alleviates Muscle Loss in Diabetes by Suppressing Atrogin-1 and MuRF1.

PubMed

Liu, Hung-Wen; Chen, Yen-Ju; Chang, Yun-Ching; Chang, Sue-Joan

2017-09-20

Stimulation of the ubiquitin-proteasome pathway-especially E3 ubiquitin ligases Atrogin-1 and MuRF1-is associated with muscle loss in diabetes. Elevated lipid metabolites impair myogenesis. Oligonol, a low molecular weight polyphenol derived from lychee, exhibited anti-diabetic and anti-obesity properties, suggesting it could be a proper supplement for attenuating muscle loss. Dietary (10 weeks) oligonol supplementation (20 or 200 mg/kg diet) on the skeletal muscle loss was investigated in diabetic db/db mice. Transcription factors NF-κB and FoxO3a involved in regulation of Atrogin-1 and MuRF1 were also investigated. Attenuation of muscle loss by oligonol (both doses) was associated with down-regulation of Atrogin-1 and MuRF1 gene expression. Oligonol supplementation decreased NF-κB expression in the nuclear fraction compared with db/db mice without oligonol supplement. Upregulation of sirtuin1 (SIRT1) expression prevented FoxO3a nuclear localization in db/db mice supplemented with oligonol. Marked increases in AMPKα activity and Ppara mRNA expression leading to lower lipid accumulation by oligonol provided additional benefits for attenuating muscle loss. Oligonol limited palmitate-induced senescent phenotype and cell cycle arrest and suppressed Atrogin-1 and MuRF1 mRNA expression in palmitate-treated C2C12 muscle cells, thus contributing to improving the impaired myotube formation. In conclusion, oligonol-mediated downregulation of Atrogin-1 and MuRF1 gene expression alleviates muscle loss and improves the impaired myotube formation, indicating that oligonol supplementation may be useful for the attenuation of myotube loss.

FoxO1 controls lysosomal acid lipase in adipocytes: implication of lipophagy during nutrient restriction and metformin treatment

PubMed Central

Lettieri Barbato, D; Tatulli, G; Aquilano, K; Ciriolo, M R

2013-01-01

Finding new molecular pathways and strategies modulating lipolysis in adipocytes is an attractive goal of the current research. Indeed, it is becoming clear that several human age-related pathologies are caused by adipose tissue expansion and altered lipid metabolism. In the present work, we show that transcription factor forkhead homeobox type protein O1 (FoxO1) is upregulated by nutrient restriction (NR) in adipocytes and exerts the transcriptional control of lipid catabolism via the induction of lysosomal acid lipase (Lipa). An increased autophagy and colocalization of lipid droplets (LDs) with lysosomes was observed implying lipophagy in Lipa-mediated LDs degradation. Interestingly, we found that metformin (Metf), a biguanide drug commonly used to treat type-2 diabetes, exerts effects comparable to that of NR. Actually, it was able to elicit FoxO1-dependent Lipa induction as well as LDs degradation through lipophagy. Moreover, we demonstrate that, during NR or Metf treatment, free fatty acids released by Lipa are directed toward AMP-activated protein kinase-mediated mitochondrial oxidation, thus maintaining energetic homeostasis in adipocytes. In conclusion, our data show that lysosomal-mediated lipid catabolism is activated by NR in adipocytes and give further support to the use of Metf as a NR mimetic to combat age-related diseases associated with altered lipid metabolism. PMID:24136225

dFOXO Activates Large and Small Heat Shock Protein Genes in Response to Oxidative Stress to Maintain Proteostasis in Drosophila.

PubMed

Donovan, Marissa R; Marr, Michael T

2016-09-02

Maintaining protein homeostasis is critical for survival at the cellular and organismal level (Morimoto, R. I. (2011) Cold Spring Harb. Symp. Quant. Biol. 76, 91-99). Cells express a family of molecular chaperones, the heat shock proteins, during times of oxidative stress to protect against proteotoxicity. We have identified a second stress responsive transcription factor, dFOXO, that works alongside the heat shock transcription factor to activate transcription of both the small heat shock protein and the large heat shock protein genes. This expression likely protects cells from protein misfolding associated with oxidative stress. Here we identify the regions of the Hsp70 promoter essential for FOXO-dependent transcription using in vitro methods and find a physiological role for FOXO-dependent expression of heat shock proteins in vivo. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

FOXO3a regulates BNIP3 and modulates mitochondrial calcium, dynamics, and function in cardiac stress

PubMed Central

Kohlbrenner, Erik; Gamb, Scott I.; Guenzel, Adam J.; Klaus, Katherine; Fayyaz, Ahmed U.; Nair, K. Sreekumaran; Hajjar, Roger J.

2016-01-01

The forkhead box O3a (FOXO3a) transcription factor has been shown to regulate glucose metabolism, muscle atrophy, and cell death in postmitotic cells. Its role in regulation of mitochondrial and myocardial function is not well studied. Based on previous work, we hypothesized that FOXO3a, through BCL2/adenovirus E1B 19-kDa protein-interacting protein 3 (BNIP3), modulates mitochondrial morphology and function in heart failure (HF). We modulated the FOXO3a-BNIP3 pathway in normal and phenylephrine (PE)-stressed adult cardiomyocytes (ACM) in vitro and developed a cardiotropic adeno-associated virus serotype 9 encoding dominant-negative FOXO3a (AAV9.dn-FX3a) for gene delivery in a rat model of HF with preserved ejection fraction (HFpEF). We found that FOXO3a upregulates BNIP3 expression in normal and PE-stressed ACM, with subsequent increases in mitochondrial Ca2+, leading to decreased mitochondrial membrane potential, mitochondrial fragmentation, and apoptosis. Whereas dn-FX3a attenuated the increase in BNIP3 expression and its consequences in PE-stressed ACM, AAV9.dn-FX3a delivery in an experimental model of HFpEF decreased BNIP3 expression, reversed adverse left ventricular remodeling, and improved left ventricular systolic and, particularly, diastolic function, with improvements in mitochondrial structure and function. Moreover, AAV9.dn-FX3a restored phospholamban phosphorylation at S16 and enhanced dynamin-related protein 1 phosphorylation at S637. Furthermore, FOXO3a upregulates maladaptive genes involved in mitochondrial apoptosis, autophagy, and cardiac atrophy. We conclude that FOXO3a activation in cardiac stress is maladaptive, in that it modulates Ca2+ cycling, Ca2+ homeostasis, and mitochondrial dynamics and function. Our results suggest an important role of FOXO3a in HF, making it an attractive potential therapeutic target. Listen to this article's corresponding podcast at http://ajpheart.podbean.com/e/role-of-foxo3a-in-heart-failure/. PMID:27694219

MicroRNA-142-5p Overexpression Inhibits Cell Growth and Induces Apoptosis by Regulating FOXO in Hepatocellular Carcinoma Cells.

PubMed

Lou, Kexin; Chen, Ning; Li, Zhihong; Zhang, Bei; Wang, Xiuli; Chen, Ye; Xu, Haining; Wang, Dongwei; Wang, Hao

2017-01-02

Abnormal expression of microRNA (miR)-142-5p has been reported in hepatocellular carcinoma (HCC). However, little information is available regarding the functional role of miR-142-5p in HCC. We aimed to explore the effects of miR-142-5p aberrant expression on HCC cell growth and cell apoptosis, as well as the underlying mechanism. Human HCC cell lines HepG2 and SMMC-7721 cells were transfected with miR-142-5p mimic, inhibitor, or a corresponding negative control. Cell viability, cell cycle distribution, and cell apoptosis were then analyzed. In addition, protein expression of Forkhead box, class O (FOXO) 1 and 3, a Bcl-2-interacting mediator of cell death (Bim), procaspase 3, and activated caspase 3 was measured. After transfection with miR-142-5p inhibitor, FOXO1 and FOXO3 were overexpressed, and then the cell viability and cell apoptosis were determined again. The relative cell viability in both HepG2 and SMMC-7721 cells was significantly reduced by miR-142-5p overexpression (p < 0.05). miR-142-5p overexpression displayed a significant blockage at the G1/S transition and significantly increased the percentages of G0/G1 phase. Moreover, the results showed that miR-142-5p overexpression significantly induced cell apoptosis and statistically elevated the protein expression levels of FOXO1, FOXO3, Bim, procaspase 3, and activated caspase 3. However, the cells transfected with miR-142-5p inhibitor showed contrary results. Additionally, the effects of miR-142-5p inhibitor on cell viability and apoptosis were reversed by overexpression of FOXO. In conclusion, our results suggest that miR-142-5p overexpression shows an important protective role in HCC by inhibiting cell growth and inducing apoptosis. These effects might be by regulating FOXO expression in HCC cells.

BAFF induces spleen CD4{sup +} T cell proliferation by down-regulating phosphorylation of FOXO3A and activates cyclin D2 and D3 expression

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

Ji, Fang; Chen, Rongjing; Liu, Baojun

2012-09-07

Highlights: Black-Right-Pointing-Pointer Firstly analyze the mechanism of BAFF and anti-CD3 co-stimulation on purified mouse splenic CD4{sup +} T cells. Black-Right-Pointing-Pointer Carrying out siRNA technology to study FOXO3A protein function. Black-Right-Pointing-Pointer Helpful to understand the T cell especially CD4{sup +} T cell's role in immunological reaction. -- Abstract: The TNF ligand family member 'B cell-activating factor belonging to the TNF family' (BAFF, also called BLyS, TALL-1, zTNF-4, and THANK) is an important survival factor for B and T cells. In this study, we show that BAFF is able to induce CD4{sup +} spleen T cell proliferation when co-stimulated with anti-CD3. Expressionmore » of phosphorylated FOXO3A was notably down-regulated and cyclins D2 and D3 were up-regulated and higher in the CD4{sup +} T cells when treated with BAFF and anti-CD3, as assessed by Western blotting. Furthermore, after FOXO3A was knocked down, expression of cyclin D1 was unchanged, compared with control group levels, but the expression of cyclins D2 and D3 increased, compared with the control group. In conclusion, our results suggest that BAFF induced CD4{sup +} spleen T cell proliferation by down-regulating the phosphorylation of FOXO3A and then activating cyclin D2 and D3 expression, leading to CD4{sup +} T cell proliferation.« less

Platelet activation suppresses HIV-1 infection of T cells

PubMed Central

2013-01-01

Background Platelets, anucleate cell fragments abundant in human blood, can capture HIV-1 and platelet counts have been associated with viral load and disease progression. However, the impact of platelets on HIV-1 infection of T cells is unclear. Results We found that platelets suppress HIV-1 spread in co-cultured T cells in a concentration-dependent manner. Platelets containing granules inhibited HIV-1 spread in T cells more efficiently than degranulated platelets, indicating that the granule content might exert antiviral activity. Indeed, supernatants from activated and thus degranulated platelets suppressed HIV-1 infection. Infection was inhibited at the stage of host cell entry and inhibition was independent of the viral strain or coreceptor tropism. In contrast, blockade of HIV-2 and SIV entry was less efficient. The chemokine CXCL4, a major component of platelet granules, blocked HIV-1 entry and neutralization of CXCL4 in platelet supernatants largely abrogated their anti-HIV-1 activity. Conclusions Release of CXCL4 by activated platelets inhibits HIV-1 infection of adjacent T cells at the stage of virus entry. The inhibitory activity of platelet-derived CXCL4 suggests a role of platelets in the defense against infection by HIV-1 and potentially other pathogens. PMID:23634812

Platelet activation suppresses HIV-1 infection of T cells.

PubMed

Solomon Tsegaye, Theodros; Gnirß, Kerstin; Rahe-Meyer, Niels; Kiene, Miriam; Krämer-Kühl, Annika; Behrens, Georg; Münch, Jan; Pöhlmann, Stefan

2013-05-01

Platelets, anucleate cell fragments abundant in human blood, can capture HIV-1 and platelet counts have been associated with viral load and disease progression. However, the impact of platelets on HIV-1 infection of T cells is unclear. We found that platelets suppress HIV-1 spread in co-cultured T cells in a concentration-dependent manner. Platelets containing granules inhibited HIV-1 spread in T cells more efficiently than degranulated platelets, indicating that the granule content might exert antiviral activity. Indeed, supernatants from activated and thus degranulated platelets suppressed HIV-1 infection. Infection was inhibited at the stage of host cell entry and inhibition was independent of the viral strain or coreceptor tropism. In contrast, blockade of HIV-2 and SIV entry was less efficient. The chemokine CXCL4, a major component of platelet granules, blocked HIV-1 entry and neutralization of CXCL4 in platelet supernatants largely abrogated their anti-HIV-1 activity. Release of CXCL4 by activated platelets inhibits HIV-1 infection of adjacent T cells at the stage of virus entry. The inhibitory activity of platelet-derived CXCL4 suggests a role of platelets in the defense against infection by HIV-1 and potentially other pathogens.

PAX-FOXO1 fusion status drives unfavorable outcome for children with rhabdomyosarcoma: a children's oncology group report.

PubMed

Skapek, Stephen X; Anderson, James; Barr, Frederic G; Bridge, Julia A; Gastier-Foster, Julie M; Parham, David M; Rudzinski, Erin R; Triche, Timothy; Hawkins, Douglas S

2013-09-01

Rhabdomyosarcoma (RMS) is divided into two major histological subtypes: alveolar (ARMS) and embryonal (ERMS), with most ARMS expressing one of two oncogenic genes fusing PAX3 or PAX7 with FOXO1 (P3F and P7F, respectively). The Children's Oncology Group (COG) carried out a multi-institutional clinical trial to evaluate the prognostic value of PAX-FOXO1 fusion status. Study participants were treated on COG protocol D9803 for intermediate risk ARMS or ERMS using multi-agent chemotherapy, radiotherapy, and surgery. Central diagnostic pathology review and molecular testing for fusion genes were carried out on prospectively collected specimens. Event-free (EFS) and overall survival (OS) at 5 years were correlated with histological subtype and PAX-FOXO1 status. Of 616 eligible D9803 enrollees, 434 cases had adequate clinical, molecular, and pathology data for definitive classification as ERMS, ARMS P3F+ or P7F+, or ARMSn (without detectable fusion). EFS was worse for those with ARMS P3F+ (54%) and P7F+ (65%) than those with ERMS (77%; P < 0.001). EFS for ARMSn and ERMS were not statistically different (90% vs. 77%, P = 0.15). ARMS P3F+ had poorer OS (64%) than ARMS P7F+ (87%), ARMSn (89%), and ERMS (82%; P = 0.006). ARMSn has an outcome similar to ERMS and superior EFS compared to ARMS with either P3F or P7F, when given therapy designed for children with intermediate risk RMS. This prospective analysis supports incorporation of PAX-FOXO1 fusion status into risk stratification and treatment allocation. Copyright © 2013 Wiley Periodicals, Inc.

Evidence for an association between increased oxidative stress and derangement of FOXO1 signaling in tumorigenesis of a cellular angiofibroma with monoallelic 13q14: a case report

PubMed Central

Arakaki, Kazunari; Chinen, Katsuya; Kamiya, Masuzo; Tanabe, Yasuka; Tawata, Natsumi; Ikehara, Fukino; Uehara, Karina; Shimabukuro, Hiroichi; Kinjo, Takao

2014-01-01

Cellular angiofibroma (CAF) is a rare soft tissue tumor characterized by random arrangement of spindle tumor cells in the stroma with short collagen bundles and thick- and hyalinized small vessels. CAFs share histological characteristics with spindle cell lipomas and mammary type myofibroblastomas. Because these tumors harbor monoallelic 13q14, common genetic and molecular mechanism for tumorigenesis is presumed. In this study, we reported a case of CAF in a 69-year-old man with monoallelic 13q14. Immunohistochemical analysis revealed that FOXO1, which is located in chromosome 13q14, was not expressed in the tumor. We also detected oxidative stress markers and found p38 MAPK activation, which is often induced by cellular stressors such as reactive oxygen species (ROS). Because FOXO1 induces the expression of genes encoding enzymes that generate antioxidants, oxidative stress induced by loss of FOXO1 expression may be common among CAFs, spindle cell lipomas, and mammary type myofibroblastomas. PMID:25674275

Evidence for an association between increased oxidative stress and derangement of FOXO1 signaling in tumorigenesis of a cellular angiofibroma with monoallelic 13q14: a case report.

PubMed

Arakaki, Kazunari; Chinen, Katsuya; Kamiya, Masuzo; Tanabe, Yasuka; Tawata, Natsumi; Ikehara, Fukino; Uehara, Karina; Shimabukuro, Hiroichi; Kinjo, Takao

2014-01-01

Cellular angiofibroma (CAF) is a rare soft tissue tumor characterized by random arrangement of spindle tumor cells in the stroma with short collagen bundles and thick- and hyalinized small vessels. CAFs share histological characteristics with spindle cell lipomas and mammary type myofibroblastomas. Because these tumors harbor monoallelic 13q14, common genetic and molecular mechanism for tumorigenesis is presumed. In this study, we reported a case of CAF in a 69-year-old man with monoallelic 13q14. Immunohistochemical analysis revealed that FOXO1, which is located in chromosome 13q14, was not expressed in the tumor. We also detected oxidative stress markers and found p38 MAPK activation, which is often induced by cellular stressors such as reactive oxygen species (ROS). Because FOXO1 induces the expression of genes encoding enzymes that generate antioxidants, oxidative stress induced by loss of FOXO1 expression may be common among CAFs, spindle cell lipomas, and mammary type myofibroblastomas.

Saponins from Aralia taibaiensis Attenuate D-Galactose-Induced Aging in Rats by Activating FOXO3a and Nrf2 Pathways

PubMed Central

Li, Ying-Na; Guo, Yu; Xi, Miao-Miao; Yang, Pei; Zhou, Xue-Ying; Yin, Shuang; Hai, Chun-Xu; Li, Jin-Gang; Qin, Xu-Jun

2014-01-01

Reactive oxygen species (ROS) are closely related to the aging process. In our previous studies, we found that the saponins from Aralia taibaiensis have potent antioxidant activity, suggesting the potential protective activity on the aging. However, the protective effect of the saponins and the possible underlying molecular mechanism remain unknown. In the present study, we employed a D-galactose-induced aging rat model to investigate the protective effect of the saponins. We found that D-galactose treatment induced obvious aging-related changes such as the decreased thymus and spleen coefficients, the increased advanced glycation end products (AGEs) level, senescence-associated β-galactosidase (SAβ-gal) activity, and malondialdehyde (MDA) level. Further results showed that Forkhead box O3a (FOXO3a), nuclear factor-erythroid 2-related factor 2 (Nrf2), and their targeted antioxidants such as superoxide dismutase 2 (SOD2), catalase (CAT), glutathione reductase (GR), glutathione (GSH), glutamate-cysteine ligase (GCL), and heme oxygenase 1 (HO-1) were all inhibited in the aging rats induced by D-galactose treatment. Saponins supplementation showed effective protection on these changes. These results demonstrate that saponins from Aralia taibaiensis attenuate the D-galactose-induced rat aging. By activating FOXO3a and Nrf2 pathways, saponins increase their downstream multiple antioxidants expression and function, at least in part contributing to the protection on the D-galactose-induced aging in rats. PMID:24669284

The Human T-Cell Leukemia Virus Type 1 Oncoprotein Tax Controls Forkhead Box O4 Activity through Degradation by the Proteasome▿

PubMed Central

Oteiza, Alexandra; Mechti, Nadir

2011-01-01

Activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway by the viral Tax oncoprotein plays a pivotal role in clonal expansion of human T-cell leukemia virus type 1 (HTLV-1)-infected cells. As the Forkhead box O (FoxO) tumor suppressors act as downstream effectors of PI3K/Akt, they represent good candidate targets whose dysregulation by Tax might be involved in HTLV-1-mediated activation and transformation of infected cells. In this report, we provide evidence showing that Tax induces a dose-dependent degradation of FoxO4 by the ubiquitin-proteasome pathway. Consistent with that, we demonstrate that Tax expression increases the interaction between FoxO4 and Mdm2 E3 ligase, leading to a strong FoxO4 polyubiquitination. These processes require the phosphorylation of FoxO4 by Akt, since a mutant of FoxO4 with mutations on its three Akt phosphorylation sites appears to be resistant to Tax-mediated degradation and ubiquitination. In addition, we show that Tax expression is associated with degradation and phosphorylation of endogenous FoxO4 in Jurkat T cells. Finally, we demonstrate that Tax represses FoxO4 transcriptional activity. Our study demonstrates that Tax can control FoxO4 protein stability and transcriptional activity and provides new insight into the subversion of cell signaling pathways during HTLV-1 infection. PMID:21525355

Foxo3 circular RNA promotes cardiac senescence by modulating multiple factors associated with stress and senescence responses.

PubMed

Du, William W; Yang, Weining; Chen, Yu; Wu, Zhong-Kai; Foster, Francis Stuart; Yang, Zhenguo; Li, Xiangmin; Yang, Burton B

2017-05-07

Circular RNAs are a subclass of non-coding RNAs detected within mammalian cells. This study was designed to test the roles of a circular RNA circ-Foxo3 in senescence using in vitro and in vivo approaches. Using the approaches of molecular and cellular biology, we show that a circular RNA generated from a member of the forkhead family of transcription factors, Foxo3, namely circ-Foxo3, was highly expressed in heart samples of aged patients and mice, which was correlated with markers of cellular senescence. Doxorubicin-induced cardiomyopathy was aggravated by ectopic expression of circ-Foxo3 but was relieved by silencing endogenous circ-Foxo3. We also found that silencing circ-Foxo3 inhibited senescence of mouse embryonic fibroblasts and that ectopic expression of circ-Foxo3 induced senescence. We found that circ-Foxo3 was mainly distributed in the cytoplasm, where it interacted with the anti-senescent protein ID-1 and the transcription factor E2F1, as well as the anti-stress proteins FAK and HIF1α. We conclude that ID-1, E2F1, FAK, and HIF1α interact with circ-Foxo3 and are retained in the cytoplasm and could no longer exert their anti-senescent and anti-stress roles, resulting in increased cellular senescence. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2016. For permissions please email: journals.permissions@oup.com.

Amelioration of streptozotocin‑induced pancreatic β cell damage by morin: Involvement of the AMPK‑FOXO3‑catalase signaling pathway.

PubMed

Wang, Ning; Zhang, Jiahui; Qin, Mengting; Yi, Wenjing; Yu, Shuang; Chen, Yi; Guan, Jing; Zhang, Rui

2018-03-01

Pancreatic β cells are sensitive to oxidative stress, which is one of the predominant causes of cell damage and the emergence of diabetes. The identification of effective therapeutic strategies to protect pancreatic cells from oxidative stress has increased interest in the screening of antioxidants from natural products. The present study aimed to investigate the protective effects of morin against streptozotocin (STZ)‑induced cell damage in a rat insulinoma cell line (RINm5F pancreatic β cells) and to identify the underlying mechanisms. The results indicated that morin inhibited the increase in intracellular reactive oxygen species, attenuated the activity of poly (ADP‑ribose) polymerase, restored intracellular nicotinamide adenine dinucleotide levels and reduced the apoptotic cell death of STZ‑treated pancreatic β cells. Treatment with morin significantly upregulated catalase in pancreatic β cells, and ameliorated the STZ‑induced loss of catalase at the genetic, protein and enzymatic level. In further experiments, morin induced the phosphorylation of 5' adenosine monophosphate‑activated protein kinase (AMPK), which subsequently promoted the translocation of forkhead box O3 (FOXO3) to the nucleus. Specific small interfering RNAs (siRNAs) against AMPK and FOXO3 suppressed morin‑induced catalase expression. Furthermore, catalase‑specific siRNA abolished the protective effects of morin against STZ‑stimulated cell death. Taken together, these results indicated that morin protected RINm5F cells from STZ‑induced cell damage by triggering the phosphorylation of AMPK, thus resulting in subsequent activation of FOXO3 and induction of catalase.

Effect of acute and chronic eccentric exercise on FOXO1 mRNA expression as fiber type transition factor in rat skeletal muscles.

PubMed

Azad, Milad; Khaledi, Neda; Hedayati, Mehdi

2016-06-15

Skeletal muscle is a highly elastic tissue which can respond to various functional demands by altering fiber-type composition. Exercise affects muscle fiber phenotype. One of the transcription factors that induce fiber-type transition is forkhead box O1 (FOXO1). Since eccentric contraction considered an essential part of exercise, so we are interested to see the effects of eccentric exercise (acute/chronic) on FOXO1 as an important factor of fiber-type transition in rat skeletal muscles. Twenty-four Sprague-Dawley rats (190-235g) were divided to 3 groups of 8 rats: 1) chronic eccentric exercise (CEE), 2) acute eccentric exercise (AEE), and 3) control (C). The exercise groups underwent downhill running protocol. CEE was running on treadmill in 3 days of week for 9 weeks, that slope and duration gradually managed from -4° to -16° and 15 to 90 min, respectively. AEE group was running with 16 m/min on -16° slope for 3 consecutive days that included 18 sets of 5 min with rest interval of 2 min in between. Soleus and super vastus lateralis (SVL) muscles mRNA were analyzed by real-time RT-PCR. SVL FOXO1 mRNA levels increased by 3.92-fold in the AEE and decreased 0.56-fold in the CEE group and were not significant in soleus muscle. In soleus muscle, myosin heavy chain (MHC) IIa, IIx, and IIb decreased in the AEE group and MHC IIa and IIx decreased in the CEE group. In SVL muscle, MHC I, IIa, and IIx increased in the AEE group and MHC IIa and IIX increased in the CEE group. In summary, both acute and chronic eccentric exercise could lead to change in FOXO1 mRNA only in fast SVL muscle of rat and so could induce fiber-type transition in both muscles regardless of changes in expression of FOXO1. So, oxidative stress can play important role in change of FOXO1. Copyright © 2016 Elsevier B.V. All rights reserved.

Constitutively expressed COX-2 in osteoblasts positively regulates Akt signal transduction via suppression of PTEN activity.

PubMed

Li, Ching-Ju; Chang, Je-Ken; Wang, Gwo-Jaw; Ho, Mei-Ling

2011-02-01

Cyclooxygenase-2 (COX-2) is thought to be an inducible enzyme, but increasing reports indicate that COX-2 is constitutively expressed in several organs. The status of COX-2 expression in bone and its physiological role remains undefined. Non-selective non-steroidal anti-inflammatory drugs (NSAIDs) and selective COX-2 inhibitors, which commonly suppress COX-2 activity, were reported to suppress osteoblast proliferation via Akt/FOXO3a/p27(Kip1) signaling, suggesting that COX-2 may be the key factor of the suppressive effects of NSAIDs on proliferation. Although Akt activation correlates with PTEN deficiency and cell viability, the role of COX-2 on PTEN/Akt regulation remains unclear. In this study, we hypothesized that COX-2 may be constitutively expressed in osteoblasts and regulate PTEN/Akt-related proliferation. We examined the localization and co-expression of COX-2 and p-Akt in normal mouse femurs and in cultured mouse (mOBs) and human osteoblasts (hOBs). Our results showed that osteoblasts adjacent to the trabeculae, periosteum and endosteum in mouse femurs constitutively expressed COX-2, while COX-2 co-expressed with p-Akt in osteoblasts sitting adjacent to trabeculae in vivo, and in mOBs and hOBs in vitro. We further used COX-2 siRNA to test the role of COX-2 in Akt signaling in hOBs; COX-2 silencing significantly inhibited PTEN phosphorylation, enhanced PTEN activity, and suppressed p-Akt level and proliferation. However, replenishment of the COX-2 enzymatic product, PGE2, failed to reverse COX-2-dependent Akt phosphorylation. Furthermore, transfection with recombinant human COX-2 (rhCOX-2) significantly reversed COX-2 siRNA-suppressed PTEN phosphorylation, but this effect was reduced when the enzymatic activity of rhCOX-2 was blocked. This finding indicated that the effect of COX-2 on PTEN/Akt signaling is not related to PGE2 but still dependent on COX-2 enzymatic activity. Conversely, COX-1 silencing did not affect PTEN/Akt signaling. Our findings provide

ACTIVE SUPPRESSION OF IMMUNOGLOBULIN ALLOTYPE SYNTHESIS

PubMed Central

Herzenberg, Leonore A.; Chan, Eva L.; Ravitch, Myrnice M.; Riblet, Roy J.; Herzenberg, Leonard A.

1973-01-01

Thymus-derived cells (T cells) that actively suppress production of IgG2a immunoglobulins carrying the Ig-1b allotype have been found in adult (SJL x BALB/c)F1 mice exposed to anti-Ig-1b early in life. The suppression is specific for Ig-1b. The allelic product, Ig-1a, is unaffected. Spleen, lymph node, bone marrow, or thymus cells from suppressed mice suppress production of Ig-1b by syngeneic spleen cells from normal F1 mice. When a mixture of suppressed and normal cells is transferred into lethally irradiated BALB/c mice, there is a short burst of Ig-1b production after which Ig-1b levels in the recipient fall rapidly below detectability. Pretreatment of the cells from the suppressed mice with antiserum specific for T cells (anti-Thy-1b) plus complement before mixture destroys the suppressing activity. Similar results with suppressor cells were obtained in vitro using Mishell-Dutton cultures. Mixture of spleen cells from suppressed animals with sheep erythrocyte (SRBC)-primed syngeneic normal spleen before culture suppresses Ig-1b plaque-forming cell (PFC) formation while leaving Ig-1a PFC unaffected. Treatment of the suppressed spleen with anti-Thy-1b before transfer removes the suppressing activity. PMID:4541122

Membrane Type 1–Matrix Metalloproteinase/Akt Signaling Axis Modulates TNF-α-Induced Procoagulant Activity and Apoptosis in Endothelial Cells

PubMed Central

Ohkawara, Hiroshi; Ishibashi, Toshiyuki; Sugimoto, Koichi; Ikeda, Kazuhiko; Ogawa, Kazuei; Takeishi, Yasuchika

2014-01-01

Membrane type 1–matrix metalloproteinase (MT1-MMP) functions as a signaling molecule in addition to a proteolytic enzyme. Our hypothesis was that MT1-MMP cooperates with protein kinase B (Akt) in tumor necrosis factor (TNF)-α-induced signaling pathways of vascular responses, including tissue factor (TF) procoagulant activity and endothelial apoptosis, in cultured human aortic endothelial cells (ECs). TNF-α (10 ng/mL) induced a decrease in Akt phosphorylation within 60 minutes in ECs. A chemical inhibitor of MMP, TIMP-2 and selective small interfering RNA (siRNA)-mediated suppression of MT1-MMP reversed TNF-α-triggered transient decrease of Akt phosphorylation within 60 minutes, suggesting that MT1-MMP may be a key regulator of Akt phosphorylation in TNF-α-stimulated ECs. In the downstream events, TNF-α increased TF antigen and activity, and suppressed the expression of thrombomodulin (TM) antigen. Inhibition of Akt markedly enhanced TNF-α-induced expression of TF antigen and activity, and further reduced the expression of TM antigen. Silencing of MT1-MMP by siRNA also reversed the changed expression of TF and TM induced by TNF-α. Moreover, TNF-α induced apoptosis of ECs through Akt- and forkhead box protein O1 (FoxO1)-dependent signaling pathway and nuclear factor-kB (NF-kB) activation. Knockdown of MT1-MMP by siRNA reversed apoptosis of ECs by inhibiting TNF-α-induced Akt-dependent regulation of FoxO1 in TNF-α-stimulated ECs. Immunoprecipitation demonstrated that TNF-α induced the changes in the associations between the cytoplasmic fraction of MT1-MMP and Akt in ECs. In conclusion, we show new evidence that MT1-MMP/Akt signaling axis is a key modifier for TNF-α-induced signaling pathways for modulation of procoagulant activity and apoptosis of ECs. PMID:25162582

PAX-FOXO1 Fusion Status Drives Unfavorable Outcome for Children With Rhabdomyosarcoma: A Children’s Oncology Group Report

PubMed Central

Skapek, Stephen X.; Anderson, James; Barr, Frederic G.; Bridge, Julia A.; Gastier-Foster, Julie M.; Parham, David M.; Rudzinski, Erin R.; Triche, Timothy; Hawkins, Douglas S.

2015-01-01

Background Rhabdomyosarcoma (RMS) is divided into two major histological subtypes: alveolar (ARMS) and embryonal (ERMS), with most ARMS expressing one of two oncogenic genes fusing PAX3 or PAX7 with FOXO1 (P3F and P7F, respectively). The Children’s Oncology Group (COG) carried out a multi-institutional clinical trial to evaluate the prognostic value of PAX-FOXO1 fusion status. Methods Study participants were treated on COG protocol D9803 for intermediate risk ARMS or ERMS using multi-agent chemotherapy, radiotherapy, and surgery. Central diagnostic pathology review and molecular testing for fusion genes were carried out on prospectively collected specimens. Event-free (EFS) and overall survival (OS) at 5 years were correlated with histological subtype and PAX-FOXO1 status. Results Of 616 eligible D9803 enrollees, 434 cases had adequate clinical, molecular, and pathology data for definitive classification as ERMS, ARMS P3F+ or P7F+, or ARMSn (without detectable fusion). EFS was worse for those with ARMS P3F+ (54%) and P7F+ (65%) than those with ERMS (77%; P < 0.001). EFS for ARMSn and ERMS were not statistically different (90% vs. 77%, P = 0.15). ARMS P3F+had poorer OS (64%) than ARMS P7F+ (87%), ARMSn (89%), and ERMS (82%; P = 0.006). Conclusions ARMSn has an outcome similar to ERMS and superior EFS compared to ARMS with either P3F or P7F, when given therapy designed for children with intermediate risk RMS. This prospective analysis supports incorporation of PAX-FOXO1 fusion status into risk stratification and treatment allocation. PMID:23526739

Transcriptional regulation of FoxO3 gene by glucocorticoids in murine myotubes

PubMed Central

Kuo, Taiyi; Liu, Patty H.; Chen, Tzu-Chieh; Lee, Rebecca A.; New, Jenny; Zhang, Danyun; Lei, Cassandra; Chau, Andy; Tang, Yicheng; Cheung, Edna

2016-01-01

Glucocorticoids and FoxO3 exert similar metabolic effects in skeletal muscle. FoxO3 gene expression was increased by dexamethasone (Dex), a synthetic glucocorticoid, both in vitro and in vivo. In C2C12 myotubes the increased expression is due to, at least in part, the elevated rate of FoxO3 gene transcription. In the mouse FoxO3 gene, we identified three glucocorticoid receptor (GR) binding regions (GBRs): one being upstream of the transcription start site, −17kbGBR; and two in introns, +45kbGBR and +71kbGBR. Together, these three GBRs contain four 15-bp glucocorticoid response elements (GREs). Micrococcal nuclease (MNase) assay revealed that Dex treatment increased the sensitivity to MNase in the GRE of +45kbGBR and +71kbGBR upon 30- and 60-min Dex treatment, respectively. Conversely, Dex treatment did not affect the chromatin structure near the −17kbGBR, in which the GRE is located in the linker region. Dex treatment also increased histone H3 and/or H4 acetylation in genomic regions near all three GBRs. Moreover, using chromatin conformation capture (3C) assay, we showed that Dex treatment increased the interaction between the −17kbGBR and two genomic regions: one located around +500 bp and the other around +73 kb. Finally, the transcriptional coregulator p300 was recruited to all three GBRs upon Dex treatment. The reduction of p300 expression decreased FoxO3 gene expression and Dex-stimulated interaction between distinct genomic regions of FoxO3 gene identified by 3C. Overall, our results demonstrate that glucocorticoids activated FoxO3 gene transcription through multiple GREs by chromatin structural change and DNA looping. PMID:26758684

Insulin-like Growth Factor 1 Regulates the Expression of ATP-Binding Cassette Transporter A1 in Pancreatic Beta Cells.

PubMed

Lyu, J; Imachi, H; Iwama, H; Zhang, H; Murao, K

2016-05-01

ATP-binding cassette transporter A1 (ABCA1) in pancreatic beta cells influences insulin secretion and cholesterol homeostasis. The present study investigates whether insulin-like growth factor 1 (IGF-1), which mediates stimulation of ABCA1 gene expression, could also interfere with the phosphatidylinositol 3-kinase (PI3-K) cascade.ABCA1 expression was examined by real-time polymerase chain reaction (PCR), Western blot analysis, and a reporter gene assay in rat insulin-secreting INS-1 cells incubated with IGF-1. The binding of forkhead box O1 (FoxO1) protein to the ABCA1 promoter was assessed by a chromatin immunoprecipitation (ChIP) assay. ABCA1 protein levels increased in response to rising concentrations of IGF-1. Real-time PCR analysis showed a significant increase in ABCA1 mRNA expression. However, both effects were suppressed after silencing the IGF-1 receptor. In parallel with its effect on endogenous ABCA1 mRNA levels, IGF-1 induced the activity of a reporter construct containing the ABCA1 promoter, while it was abrogated by LY294002, a specific inhibitor of PI3-K. Constitutively active Akt stimulated activity of the ABCA1 promoter, and a dominant-negative mutant of Akt or mutagenesis of the FoxO1 response element in the ABCA1 promoter abolished the ability of IGF-1 to stimulate promoter activity. A ChIP assay showed that FoxO1 mediated its transcriptional activity by directly binding to the ABCA1 promoter region. The knockdown of FoxO1 disrupted the effect of IGF-1 on ABCA1 expression. Furthermore, IGF-1 promoted cholesterol efflux and reduced the pancreatic lipotoxicity. These results demonstrate that the PI3-K/Akt/FoxO1 pathway contributes to the regulation of ABCA1 expression in response to IGF-1 stimulation. © Georg Thieme Verlag KG Stuttgart · New York.

FoxO is a critical regulator of stem cell maintenance in immortal Hydra.

PubMed

Boehm, Anna-Marei; Khalturin, Konstantin; Anton-Erxleben, Friederike; Hemmrich, Georg; Klostermeier, Ulrich C; Lopez-Quintero, Javier A; Oberg, Hans-Heinrich; Puchert, Malte; Rosenstiel, Philip; Wittlieb, Jörg; Bosch, Thomas C G

2012-11-27

Hydra's unlimited life span has long attracted attention from natural scientists. The reason for that phenomenon is the indefinite self-renewal capacity of its stem cells. The underlying molecular mechanisms have yet to be explored. Here, by comparing the transcriptomes of Hydra's stem cells followed by functional analysis using transgenic polyps, we identified the transcription factor forkhead box O (FoxO) as one of the critical drivers of this continuous self-renewal. foxO overexpression increased interstitial stem cell and progenitor cell proliferation and activated stem cell genes in terminally differentiated somatic cells. foxO down-regulation led to an increase in the number of terminally differentiated cells, resulting in a drastically reduced population growth rate. In addition, it caused down-regulation of stem cell genes and antimicrobial peptide (AMP) expression. These findings contribute to a molecular understanding of Hydra's immortality, indicate an evolutionarily conserved role of FoxO in controlling longevity from Hydra to humans, and have implications for understanding cellular aging.

Association of FOXO3A variation with human longevity confirmed in German centenarians

PubMed Central

Flachsbart, Friederike; Caliebe, Amke; Kleindorp, Rabea; Blanché, Hélène; von Eller-Eberstein, Huberta; Nikolaus, Susanna; Schreiber, Stefan; Nebel, Almut

2009-01-01

The human forkhead box O3A gene (FOXO3A) encodes an evolutionarily conserved key regulator of the insulin–IGF1 signaling pathway that is known to influence metabolism and lifespan in model organisms. A recent study described 3 SNPs in the FOXO3A gene that were statistically significantly associated with longevity in a discovery sample of long-lived men of Japanese ancestry [Willcox et al. (2008) Proc Natl Acad Sci USA 105:13987–13992]. However, this finding required replication in an independent population. Here, we have investigated 16 known FOXO3A SNPs in an extensive collection of 1,762 German centenarians/nonagenarians and younger controls and provide evidence that polymorphisms in this gene were indeed associated with the ability to attain exceptional old age. The FOXO3A association was considerably stronger in centenarians than in nonagenarians, highlighting the importance of centenarians for genetic longevity research. Our study extended the initial finding observed in Japanese men to women and indicates that both genders were likely to be equally affected by variation in FOXO3A. Replication in a French centenarian sample generated a trend that supported the previous results. Our findings confirmed the initial discovery in the Japanese sample and indicate FOXO3A as a susceptibility gene for prolonged survival in humans. PMID:19196970

Intracellular signals mediating the food intake suppressive effects of hindbrain glucagon-like-peptide-1 receptor activation

PubMed Central

Hayes, Matthew R.; Leichner, Theresa M.; Zhao, Shiru; Lee, Grace S.; Chowansky, Amy; Zimmer, Derek; De Jonghe, Bart C.; Kanoski, Scott E.; Grill, Harvey J.; Bence, Kendra K.

2011-01-01

Summary Glucagon-like-peptide-1 receptor (GLP-1R) activation within the nucleus tractus solitarius (NTS) suppresses food intake and body weight (BW), but the intracellular signals mediating these effects are unknown. Here, hindbrain (4th icv) GLP-1R activation by Exendin-4 increased PKA and MAPK activity and decreased phosphorylation of AMPK in NTS. PKA and MAPK signaling contribute to food intake and BW suppression by Exendin-4, as inhibitors RpcAMP and U0126 (4th icv), respectively, attenuated Exendin-4's effects. Hindbrain GLP-1R activation inhibited feeding by reducing meal number, not meal size. This effect was attenuated with stimulation of AMPK activity by AICAR (4th icv). The PKA, MAPK and AMPK signaling responses by Ex-4 were present in immortalized GLP-1R-expressing neurons (GT1-7). In conclusion, hindbrain GLP-1R activation suppresses food intake and BW through coordinated PKA-mediated suppression of AMPK and activation of MAPK. Pharmacotherapies targeting these signaling pathways, which mediate intake-suppressive effects of CNS GLP-1R activation, may prove efficacious in treating obesity. PMID:21356521

Ursolic acid improves domoic acid-induced cognitive deficits in mice

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

Wu, Dong-mei; Key Laboratory for Biotechnology on Medicinal Plants of Jiangsu Province, School of Life Science, Xuzhou Normal University, Xuzhou 221116, Jiangsu Province; Lu, Jun, E-mail: lu-jun75@163.com

Our previous findings suggest that mitochondrial dysfunction is the mechanism underlying cognitive deficits induced by domoic acid (DA). Ursolic acid (UA), a natural triterpenoid compound, possesses many important biological functions. Evidence shows that UA can activate PI3K/Akt signaling and suppress Forkhead box protein O1 (FoxO1) activity. FoxO1 is an important regulator of mitochondrial function. Here we investigate whether FoxO1 is involved in the oxidative stress-induced mitochondrial dysfunction in DA-treated mice and whether UA inhibits DA-induced mitochondrial dysfunction and cognitive deficits through regulating the PI3K/Akt and FoxO1 signaling pathways. Our results showed that FoxO1 knockdown reversed the mitochondrial abnormalities and cognitivemore » deficits induced by DA in mice through decreasing HO-1 expression. Mechanistically, FoxO1 activation was associated with oxidative stress-induced JNK activation and decrease of Akt phosphorylation. Moreover, UA attenuated the mitochondrial dysfunction and cognitive deficits through promoting Akt phosphorylation and FoxO1 nuclear exclusion in the hippocampus of DA-treated mice. LY294002, an inhibitor of PI3K/Akt signaling, significantly decreased Akt phosphorylation in the hippocampus of DA/UA mice, which weakened UA actions. These results suggest that UA could be recommended as a possible candidate for the prevention and therapy of cognitive deficits in excitotoxic brain disorders. - Highlights: • Ursolic acid (UA) is a naturally triterpenoid compound. • UA attenuated the mitochondrial dysfunction and cognitive deficits. • Mechanistically, UA activates PI3K/Akt signaling and suppresses FoxO1 activity. • UA could be recommended as a possible candidate for anti-excitotoxic brain disorders.« less

FoxO4 inhibits atherosclerosis through its function in bone marrow derived cells

PubMed Central

Zhu, Min; Zhang, Qing-Jun; Wang, Lin; Li, Hao; Liu, Zhi-Ping

2011-01-01

Objectives FoxO proteins are transcription factors involved in varieties of cellular processes, including immune cell homeostasis, cytokine production, anti-oxidative stress, and cell proliferation and differentiation. Although these processes are implicated in the development of atherosclerosis, very little is known about the role of FoxO proteins in the context of atherosclerosis. Our objectives were to determine whether and how inactivation of Foxo4, a member of the FoxO family, in vivo promotes atherosclerosis. Methods and Results Apolipoprotein E-deficient (apoE−/−) mice were crossbred with animals lacking Foxo4 (Foxo4−/−). After 10 weeks on a high fat diet (HFD), Foxo4−/−apoE−/− mice showed elevated atherosclerosis and increased amount of macrophages and T cells in the plaque compared to apoE−/− mice. Bone marrow transplantations of chimeric C57B/6 mice reconstituted with either wild-type or Foxo4−/− bone marrows indicate that Foxo4-deficiency in bone marrow derived cells sufficiently promoted atherosclerosis. Foxo4-null macrophages produced elevated inflammatory cytokine IL-6 and levels of reactive oxygen species (ROS) in response to lipopolysaccharides in vitro. Serum levels of IL-6 were upregulated in HFD-fed Foxo4−/−apoE−/− mice compared to those of apoE−/− mice. Conclusions FoxO4 inhibits atherosclerosis through bone marrow derived cells, possibly by inhibition of ROS and inflammatory cytokines that promote monocyte recruitment and/or retention. PMID:22005198

Soy Leaf Extract Containing Kaempferol Glycosides and Pheophorbides Improves Glucose Homeostasis by Enhancing Pancreatic β-Cell Function and Suppressing Hepatic Lipid Accumulation in db/db Mice.

PubMed

Li, Hua; Ji, Hyeon-Seon; Kang, Ji-Hyun; Shin, Dong-Ha; Park, Ho-Yong; Choi, Myung-Sook; Lee, Chul-Ho; Lee, In-Kyung; Yun, Bong-Sik; Jeong, Tae-Sook

2015-08-19

This study investigated the molecular mechanisms underlying the antidiabetic effect of an ethanol extract of soy leaves (ESL) in db/db mice. Control groups (db/+ and db/db) were fed a normal diet (ND), whereas the db/db-ESL group was fed ND with 1% ESL for 8 weeks. Dietary ESL improved glucose tolerance and lowered plasma glucose, glycated hemoglobin, HOMA-IR, and triglyceride levels. The pancreatic insulin content of the db/db-ESL group was significantly greater than that of the db/db group. ESL supplementation altered pancreatic IRS1, IRS2, Pdx1, Ngn3, Pax4, Ins1, Ins2, and FoxO1 expression. Furthermore, ESL suppressed lipid accumulation and increased glucokinase activity in the liver. ESL primarily contained kaempferol glycosides and pheophorbides. Kaempferol, an aglycone of kaempferol glycosides, improved β-cell proliferation through IRS2-related FoxO1 signaling, whereas pheophorbide a, a product of chlorophyll breakdown, improved insulin secretion and β-cell proliferation through IRS1-related signaling with protein kinase A in MIN6 cells. ESL effectively regulates glucose homeostasis by enhancing IRS-mediated β-cell insulin signaling and suppressing SREBP-1-mediated hepatic lipid accumulation in db/db mice.

Insulin Is Required to Maintain Albumin Expression by Inhibiting Forkhead Box O1 Protein*

PubMed Central

Chen, Qing; Lu, Mingjian; Monks, Bobby R.; Birnbaum, Morris J.

2016-01-01

Diabetes is accompanied by dysregulation of glucose, lipid, and protein metabolism. In recent years, much effort has been spent on understanding how insulin regulates glucose and lipid metabolism, whereas the effect of insulin on protein metabolism has received less attention. In diabetes, hepatic production of serum albumin decreases, and it has been long established that insulin positively controls albumin gene expression. In this study, we used a genetic approach in mice to identify the mechanism by which insulin regulates albumin gene transcription. Albumin expression was decreased significantly in livers with insulin signaling disrupted by ablation of the insulin receptor or Akt. Concomitant deletion of Forkhead Box O1 (Foxo1) in these livers rescued the decreased albumin secretion. Furthermore, activation of Foxo1 in the liver is sufficient to suppress albumin expression. These results suggest that Foxo1 acts as a repressor of albumin expression. PMID:26668316

Age-Based Differences in the Genetic Determinants of Glycemic Control: A Case of FOXO3 Variations.

PubMed

Sun, Liang; Hu, Caiyou; Qian, Yu; Zheng, Chenguang; Liang, Qinghua; Lv, Zeping; Huang, Zezhi; Qi, Keyan; Huang, Jin; Zhou, Qin; Yang, Ze

2015-01-01

Glucose homeostasis is a trait of healthy ageing and is crucial to the elderly, but less consideration has been given to the age composition in most studies involving genetics and hyperglycemia. Seven variants in FOXO3 were genotyped in three cohorts (n = 2037; LLI, MI_S and MI_N; mean age: 92.5 ± 3.6, 45.9 ± 8.2 and 46.8 ± 10.3, respectively) to compare the contribution of FOXO3 to fasting hyperglycemia (FH) between long-lived individuals (LLI, aged over 90 years) and middle-aged subjects (aged from 35-65 years). A different genetic predisposition of FOXO3 alleles to FH was observed between LLI and both of two middle-aged cohorts. In the LLI cohort, the longevity beneficial alleles of three variants with the haplotype "AGGC" in block 1 were significantly protective to FH, fasting glucose, hemoglobin A1C and HOMA-IR. Notably, combining multifactor dimensionality reduction and logistic regression, we identified a significant 3-factor interaction model (rs2802288, rs2802292 and moderate physical activity) associated with lower FH risk. However, not all of the findings were replicated in the two middle-aged cohorts. Our data provides a novel insight into the inconsistent genetic determinants between middle-aged and LLI subjects. FOXO3 might act as a shared genetic predisposition to hyperglycemia and lifespan.

Age-Based Differences in the Genetic Determinants of Glycemic Control: A Case of FOXO3 Variations

PubMed Central

Sun, Liang; Hu, Caiyou; Qian, Yu; Zheng, Chenguang; Liang, Qinghua; Lv, Zeping; Huang, Zezhi; Qi, Keyan; Huang, Jin; Zhou, Qin; Yang, Ze

2015-01-01

Background Glucose homeostasis is a trait of healthy ageing and is crucial to the elderly, but less consideration has been given to the age composition in most studies involving genetics and hyperglycemia. Methods Seven variants in FOXO3 were genotyped in three cohorts (n = 2037; LLI, MI_S and MI_N; mean age: 92.5±3.6, 45.9±8.2 and 46.8±10.3, respectively) to compare the contribution of FOXO3 to fasting hyperglycemia (FH) between long-lived individuals (LLI, aged over 90 years) and middle-aged subjects (aged from 35–65 years). Results A different genetic predisposition of FOXO3 alleles to FH was observed between LLI and both of two middle-aged cohorts. In the LLI cohort, the longevity beneficial alleles of three variants with the haplotype “AGGC” in block 1 were significantly protective to FH, fasting glucose, hemoglobin A1C and HOMA-IR. Notably, combining multifactor dimensionality reduction and logistic regression, we identified a significant 3-factor interaction model (rs2802288, rs2802292 and moderate physical activity) associated with lower FH risk. However, not all of the findings were replicated in the two middle-aged cohorts. Conclusion Our data provides a novel insight into the inconsistent genetic determinants between middle-aged and LLI subjects. FOXO3 might act as a shared genetic predisposition to hyperglycemia and lifespan. PMID:25993007

Flavonoids as Putative Inducers of the Transcription Factors Nrf2, FoxO, and PPARγ

PubMed Central

Duckstein, Nils; Hasler, Mario; Rimbach, Gerald

2017-01-01

Dietary flavonoids have been shown to extend the lifespan of some model organisms and may delay the onset of chronic ageing-related diseases. Mechanistically, the effects could be explained by the compounds scavenging free radicals or modulating signalling pathways. Transcription factors Nrf2, FoxO, and PPARγ possibly affect ageing by regulating stress response, adipogenesis, and insulin sensitivity. Using Hek-293 cells transfected with luciferase reporter constructs, we tested the potency of flavonoids from different subclasses (flavonols, flavones, flavanols, and isoflavones) to activate these transcription factors. Under cell-free conditions (ABTS and FRAP assays), we tested their free radical scavenging activities and used α-tocopherol and ascorbic acid as positive controls. Most of the tested flavonoids, but not the antioxidant vitamins, stimulated Nrf2-, FoxO-, and PPARγ-dependent promoter activities. Flavonoids activating Nrf2 also tended to induce a FoxO and PPARγ response. Interestingly, activation patterns of cellular stress response by flavonoids were not mirrored by their activities in ABTS and FRAP assays, which depended mostly on hydroxylation in the flavonoid B ring and, in some cases, extended that of the vitamins. In conclusion, the free radical scavenging properties of flavonoids do not predict whether these molecules can stimulate a cellular response linked to activation of longevity-associated transcription factors. PMID:28761622

[Preparation and characterization of mouse polyclonal antibody against conserved region of human FOXO3].

PubMed

Li, Lei; Lyu, Dan

2017-06-01

Objective To purify the recombinant protein specific to conserved region of forkhead box O3 (FOXO3) and prepare mouse anti-human FOXO3 polyclonal antibody. Methods The DNA fragment (aa290-472) encoding conserved domain of FOXO3 was amplified by PCR, and subsequently cloned into pET28a vector. Following transformation into E.coli BL21, the soluble fusion protein His-FOXO3 was induced by IPTG and purified by Ni-NTA affinity chromatography. The purified protein was used to immunize BALB/c mice to generate polyclonal antibody. The characteristics of the polyclonal antibody were assessed by ELISA, Western blotting and immunoprecipitation assays. Results We successfully prepared the expression vector pET28a-FOXO3 (aa290-472) and expressed the purified fusion protein in a soluble form. By immunizing mice with the fusion protein, we obtained anti-human FOXO3 polyclonal antibody. ELISA and Western blotting showed that the mouse antibody could recognize specifically the endogenous FOXO3 protein. Conclusion The polyclonal antibody against conserved domain of FOXO3 can identify the endogenous FOXO3 protein. It can be used to analyze the endogenous FOXO3 expression level.

Dissociation of lipotoxicity and glucotoxicity in a mouse model of obesity associated diabetes: role of forkhead box O1 (FOXO1) in glucose-induced beta cell failure.

PubMed

Kluth, O; Mirhashemi, F; Scherneck, S; Kaiser, D; Kluge, R; Neschen, S; Joost, H-G; Schürmann, A

2011-03-01

Carbohydrate-free diet prevents hyperglycaemia and beta cell destruction in the New Zealand Obese (NZO) mouse model. Here we have used a sequential dietary regimen to dissociate the effects of obesity and hyperglycaemia on beta cell function and integrity, and to study glucose-induced alterations of key transcription factors over 16 days. Mice were rendered obese by feeding a carbohydrate-free diet for 18 weeks. Thereafter, a carbohydrate-containing diet was given. Plasma glucose, plasma insulin and total pancreatic insulin were determined, and forkhead box O1 protein (FOXO1) phosphorylation and the transcription factors pancreatic and duodenal homeobox 1 (PDX1), NK6 homeobox 1 protein (NKX6.1) and v-maf musculoaponeurotic fibrosarcoma oncogene family, protein A (avian) (MAFA) were monitored by immunohistochemistry for 16 days. Dietary carbohydrates produced a rapid and continuous increase in plasma glucose in NZO mice between day 2 and 16 after the dietary challenge. Hyperglycaemia caused a dramatic dephosphorylation of FOXO1 at day 2, followed by a progressive depletion of insulin stores. The loss of beta cells was triggered by apoptosis (detectable at day 8), associated with reduction of crucial transcription factors (PDX1, NKX6.1 and MAFA). Incubation of isolated islets from carbohydrate-restricted NZO mice or MIN6 cells with palmitate and glucose for 48 h resulted in a dephosphorylation of FOXO1 and thymoma viral proto-oncogene 1 (AKT) without changing the protein levels of both proteins. The dietary regimen dissociates the effects of obesity (lipotoxicity) from those of hyperglycaemia (glucotoxicity) in NZO mice. Obese NZO mice are unable to compensate for the carbohydrate challenge by increasing insulin secretion or synthesising adequate amounts of insulin. In response to the hyperglycaemia, FOXO1 is dephosphorylated, leading to reduced levels of beta cell-specific transcription factors and to apoptosis of the cells.

A splice junction-targeted CRISPR approach (spJCRISPR) reveals human FOXO3B to be a protein-coding gene.

PubMed

Santo, Evan E; Paik, Jihye

2018-06-17

The rapid development of CRISPR technology is revolutionizing molecular approaches to the dissection of complex biological phenomena. Here we describe an alternative generally applicable implementation of the CRISPR-Cas9 system that allows for selective knockdown of extremely homologous genes. This strategy employs the lentiviral delivery of paired sgRNAs and nickase Cas9 (Cas9D10A) to achieve targeted deletion of splice junctions. This general strategy offers several advantages over standard single-guide exon-targeting CRISPR-Cas9 such as greatly reduced off-target effects, more restricted genomic editing, routine disruption of target gene mRNA expression and the ability to differentiate between closely related genes. Here we demonstrate the utility of this strategy by achieving selective knockdown of the highly homologous human genes FOXO3A and suspected pseudogene FOXO3B. We find the spJCRISPR strategy to efficiently and selectively disrupt FOXO3A and FOXO3B mRNA and protein expression; thus revealing that the human FOXO3B locus encodes a bona fide human gene. Unlike FOXO3A, we find the FOXO3B protein to be cytosolically localized in both the presence and absence of active Akt. The ability to selectively target and efficiently disrupt the expression of the closely-related FOXO3A and FOXO3B genes demonstrates the efficacy of the spJCRISPR approach. Copyright © 2018. Published by Elsevier B.V.

FoxO proteins restrain osteoclastogenesis and bone resorption by attenuating H2O2 accumulation

PubMed Central

Bartell, Shoshana M.; Kim, Ha-Neui; Ambrogini, Elena; Han, Li; Iyer, Srividhya; Serra Ucer, S.; Rabinovitch, Peter; Jilka, Robert L.; Weinstein, Robert S.; Zhao, Haibo; O’Brien, Charles A.; Manolagas, Stavros C.; Almeida, Maria

2014-01-01

Besides their cell-damaging effects in the setting of oxidative stress, reactive oxygen species (ROS) play an important role in physiological intracellular signalling by triggering proliferation and survival. FoxO transcription factors counteract ROS generation by upregulating antioxidant enzymes. Here we show that intracellular H2O2 accumulation is a critical and purposeful adaptation for the differentiation and survival of osteoclasts, the bone cells responsible for the resorption of mineralized bone matrix. Using mice with conditional loss or gain of FoxO transcription factor function, or mitochondria-targeted catalase in osteoclasts, we demonstrate this is achieved, at least in part, by downregulating the H2O2-inactivating enzyme catalase. Catalase downregulation results from the repression of the transcriptional activity of FoxO1, 3 and 4 by RANKL, the indispensable signal for the generation of osteoclasts, via an Akt-mediated mechanism. Notably, mitochondria-targeted catalase prevented the loss of bone caused by loss of oestrogens, suggesting that decreasing H2O2 production in mitochondria may represent a rational pharmacotherapeutic approach to diseases with increased bone resorption. PMID:24781012

Mevastatin ameliorates sphingosine 1‐phosphate‐induced COX‐2/PGE2‐dependent cell migration via FoxO1 and CREB phosphorylation and translocation

PubMed Central

Hsu, Chih‐Kai; Lin, Chih‐Chung; Hsiao, Li‐Der

2015-01-01

Background and Purpose Sphingosine 1‐phosphate (S1P), an important inflammatory mediator, has been shown to regulate COX‐2 production and promote various cellular responses such as cell migration. Mevastatin, an inhibitor of 3‐hydroxy‐3‐methylglutaryl‐CoA reductase (HMG‐CoA), effectively inhibits inflammatory responses. However, the mechanisms underlying S1P‐evoked COX‐2‐dependent cell migration, which is modulated by mevastatin in human tracheal smooth muscle cells (HTSMCs) remain unclear. Experimental Approach The expression of COX‐2 was determined by Western blotting, real time‐PCR and promoter analyses. The signalling molecules were investigated by pretreatment with respective pharmacological inhibitors or transfection with siRNAs. The interaction between COX‐2 promoter and transcription factors was determined by chromatin immunoprecipitation assay. Finally, the effect of mevastatin on HTSMC migration and leukocyte counts in BAL fluid and COX‐2 expression induced by S1P was determined by a cell migration assay, cell counting and Western blot. Key Results S1P stimulated mTOR activation through the Nox2/ROS and PI3K/Akt pathways, which can further stimulate FoxO1 phosphorylation and translocation to the cytosol. We also found that S1P induced CREB activation and translocation via an mTOR‐independent signalling pathway. Finally, we showed that pretreatment with mevastatin markedly reduced S1P‐induced cell migration and COX‐2/PGE2 production via a PPARγ‐dependent signalling pathway. Conclusions and Implications Mevastatin attenuates the S1P‐induced increased expression of COX‐2 and cell migration via the regulation of FoxO1 and CREB phosphorylation and translocation by PPARγ in HTSMCs. Mevastatin could be beneficial for prevention of airway inflammation in the future. PMID:26359950

Association between FOXO3A gene polymorphisms and human longevity: a meta-analysis

PubMed Central

Bao, Ji-Ming; Song, Xian-Lu; Hong, Ying-Qia; Zhu, Hai-Li; Li, Cui; Zhang, Tao; Chen, Wei; Zhao, Shan-Chao; Chen, Qing

2014-01-01

Numerous studies have shown associations between the FOXO3A gene, encoding the forkhead box O3 transcription factor, and human or specifically male longevity. However, the associations of specific FOXO3A polymorphisms with longevity remain inconclusive. We performed a meta-analysis of existing studies to clarify these potential associations. A comprehensive search was conducted to identify studies of FOXO3A gene polymorphisms and longevity. Pooled odds ratios (ORs) and 95% confidence intervals (CIs) were calculated by comparing the minor and major alleles. A total of seven articles reporting associations of FOXO3A polymorphisms with longevity were identified and included in this meta-analysis. These comprised 11 independent studies with 5241 cases and 5724 controls from different ethnic groups. rs2802292, rs2764264, rs13217795, rs1935949 and rs2802288 polymorphisms were associated with human longevity (OR = 1.36, 95% CI = 1.10–1.69, P = 0.005; OR = 1.20, 95% CI = 1.04–1.37, P = 0.01; OR = 1.27, 95% CI = 1.10–1.46, P = 0.001; OR = 1.14, 95% CI = 1.01–1.27 and OR = 1.24, 95% CI = 1.07–1.43, P = 0.003, respectively). Analysis stratified by gender indicated significant associations between rs2802292, rs2764264 and rs13217795 and male longevity (OR = 1.54, 95% CI = 1.33–1.79, P < 0.001; OR = 1.38, 95% CI = 1.15–1.66, P = 0.001; and OR = 1.39, 95% CI = 1.15–1.67, P = 0.001), but rs2802292, rs2764264 and rs1935949 were not linked to female longevity. Moreover, our study showed no association between rs2153960, rs7762395 or rs13220810 polymorphisms and longevity. In conclusion, this meta-analysis indicates a significant association of five FOXO3A gene polymorphisms with longevity, with the effects of rs2802292 and rs2764264 being male-specific. Further investigations are required to confirm these findings. PMID:24589462

Association between FOXO3A gene polymorphisms and human longevity: a meta-analysis.

PubMed

Bao, Ji-Ming; Song, Xian-Lu; Hong, Ying-Qia; Zhu, Hai-Li; Li, Cui; Zhang, Tao; Chen, Wei; Zhao, Shan-Chao; Chen, Qing

2014-01-01

Numerous studies have shown associations between the FOXO3A gene, encoding the forkhead box O3 transcription factor, and human or specifically male longevity. However, the associations of specific FOXO3A polymorphisms with longevity remain inconclusive. We performed a meta-analysis of existing studies to clarify these potential associations. A comprehensive search was conducted to identify studies of FOXO3A gene polymorphisms and longevity. Pooled odds ratios (ORs) and 95% confidence intervals (CIs) were calculated by comparing the minor and major alleles. A total of seven articles reporting associations of FOXO3A polymorphisms with longevity were identified and included in this meta-analysis. These comprised 11 independent studies with 5241 cases and 5724 controls from different ethnic groups. rs2802292, rs2764264, rs13217795, rs1935949 and rs2802288 polymorphisms were associated with human longevity (OR = 1.36, 95% CI = 1.10-1.69, P= 0.005; OR = 1.20, 95% CI = 1.04-1.37, P= 0.01; OR = 1.27, 95% CI = 1.10-1.46, P= 0.001; OR = 1.14, 95% CI = 1.01-1.27 and OR = 1.24, 95% CI = 1.07-1.43, P= 0.003, respectively). Analysis stratified by gender indicated significant associations between rs2802292, rs2764264 and rs13217795 and male longevity (OR = 1.54, 95% CI = 1.33-1.79, P < 0.001; OR = 1.38, 95% CI = 1.15-1.66, P= 0.001; and OR = 1.39, 95% CI = 1.15-1.67, P= 0.001), but rs2802292, rs2764264 and rs1935949 were not linked to female longevity. Moreover, our study showed no association between rs2153960, rs7762395 or rs13220810 polymorphisms and longevity. In conclusion, this meta-analysis indicates a significant association of five FOXO3A gene polymorphisms with longevity, with the effects of rs2802292 and rs2764264 being male-specific. Further investigations are required to confirm these findings.

Evodia alkaloids suppress gluconeogenesis and lipogenesis by activating the constitutive androstane receptor.

PubMed

Yu, Lushan; Wang, Zhangting; Huang, Minmin; Li, Yingying; Zeng, Kui; Lei, Jinxiu; Hu, Haihong; Chen, Baian; Lu, Jing; Xie, Wen; Zeng, Su

2016-09-01

The constitutive androstane receptor (CAR) is a key sensor in xenobiotic detoxification and endobiotic metabolism. Increasing evidence suggests that CAR also plays a role in energy metabolism by suppressing the hepatic gluconeogenesis and lipogenesis. In this study, we investigated the effects of two evodia alkaloids, rutaecarpine (Rut) and evodiamine (Evo), on gluconeogenesis and lipogenesis through their activation of the human CAR (hCAR). We found that both Rut and Evo exhibited anti-lipogenic and anti-gluconeogenic effects in the hyperlipidemic HepG2 cells. Both compounds can potently activate hCAR, and treatment of cells with hCAR antagonists reversed the anti-lipogenic and anti-gluconeogenic effects of Rut and Evo. The anti-gluconeogenic effect of Rut and Evo was due to the CAR-mediated inhibition of the recruitment of forkhead box O1 (FoxO1) and hepatocyte nuclear factor 4α (HNF4α) onto the phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) gene promoters. In vivo, we showed that treatment of mice with Rut improved glucose tolerance in a CAR-dependent manner. Our results suggest that the evodia alkaloids Rut and Evo may have a therapeutic potential for the treatment of hyperglycemia and type 2 diabetes. This article is part of a Special Issue entitled: Xenobiotic nuclear receptors: New Tricks for An Old Dog, edited by Dr. Wen Xie. Copyright © 2015 Elsevier B.V. All rights reserved.

Simvastatin prevents triple-negative breast cancer metastasis in pre-clinical models through regulation of FOXO3a

PubMed Central

Wolfe, Adam R.; Debeb, Bisrat G.; Lacerda, Lara; Larson, Richard; Bambhroliya, Arvind; Huang, Xuelin; Bertucci, Francois; Finetti, Pascal; Birnbaum, Daniel; Van Laere, Steven; Diagaradjan, Parmeswaran; Ruffell, Brian; Trenton, Nicholaus J.; Chu, Khoi; Hittelman, Walter; Diehl, Michael; Levental, Ilya; Ueno, Naoto T.; Woodward, Wendy A.

2016-01-01

Purpose We previously reported using statins was correlated with improved metastasis free survival in aggressive breast cancer. The purpose of this study was to examine the effect of statins on metastatic colonization by triple negative breast cancer (TNBC) cells. Experimental Design TNBC cell lines were treated with simvastatin and then studied for cell cycle progression and proliferation in vitro, and metastasis formation in vivo, following injection of statin-treated cells. Reverse-phase protein assay (RPPA) analysis was performed on statin-treated and control breast cancer cells. RNA interference targeting FOXO3a was used to measure the impact of simvastatin on FOXO3a-expressing cells. The prognostic value of FOXO3a mRNA expression was examined in 8 public breast cancer gene expression data sets including 1,479 patients. Results Simvastatin increased G1/S phase arrest of the cell cycle and inhibited both proliferation and migration of TNBC cells in vitro. In vitro pretreatment and in vivo treatment with simvastatin reduced metastases. Phosphorylated FOXO3a was downregulated after simvastatin treatment in (RPPA) analysis. Ectopic expression of FOXO3a enhanced mammosphere formation and migratory capacity in vitro. Knockdown of FOXO3a attenuated the effect of simvastatin on mammosphere formation and migration. Analysis of public gene expression data demonstrates FOXO3a mRNA downregulation was independently associated with shorter metastasis-free survival in all breast cancers, as well as in TNBC breast cancers. Conclusions Simvastatin inhibits in vitro endpoints associated with metastasis through a FOXO3a mechanism and reduced metastasis formation in vivo. FOXO3a expression is prognostic for metastasis formation in patient data. Further investigation of simvastatin as a cancer therapy is warranted. PMID:26590814

Time course expression of Foxo transcription factors in skeletal muscle following corticosteroid administration

PubMed Central

Cho, John E.; Fournier, Mario; Da, Xiaoyu

2010-01-01

Increased expression of forkhead box O (Foxo) transcription factors were reported in cultured myotubes and mouse limb muscle with corticosteroid (CS) treatment. We previously reported that administration of CS to rats resulted in muscle fiber atrophy only by day 7. The aim of this study, therefore, was to evaluate the time-course changes in the expression of Foxo transcription factors and muscle-specific ubiquitin E3 ligases in rat limb muscle following CS administration. Triamcinolone (TRI; 1 mg · kg−1 · day−1 im) was administered for 1, 3, or 7 days. Control (CTL) rats were given saline. Muscle mRNA was analyzed by real-time RT-PCR. Compared with CTL, body weights of TRI-treated animals decreased by 3, 12, and 21% at days 1, 3, and 7, respectively. Muscle IGF-1 mRNA levels decreased by 33, 65, and 58% at days 1, 3, and 7 in TRI-treated rats compared with CTL. Levels of phosphorylated Akt were 28, 50, and 36% lower in TRI animals at these time points. Foxo1 mRNA increased progressively by 1.2-, 1.4-, and 2.5-fold at days 1, 3, and 7 in TRI animals. Similar changes were noted in the expression of Foxo3a mRNA (1.3-, 1.4-, and 2.6-fold increments). By contrast, Foxo4 mRNA was not significantly changed in TRI animals. With TRI, muscle atrophy F box/Atrogin-1 increased by 1.8-, 4.1-, and 7.5-fold at days 1, 3, and 7 compared with CTL rats. By contrast, muscle RING finger 1 increased only from day 7 (2.7-fold). Gradual reduction in IGF-I expression with TRI over the time series paralleled that of Akt. These findings are consistent with a progressive stimulus to muscle protein degradation and the need to process/remove disassembled muscle proteins via the ubiquitin-proteasome system. Elucidating the dynamic catabolic responses to CS challenge is important in understanding the mechanisms underlying muscle atrophy and therapeutic measures to offset this. PMID:19850732

FOXO3a-mediated suppression of the self-renewal capacity of sphere-forming cells derived from the ovarian cancer SKOV3 cell line by 7-difluoromethoxyl-5,4'-di-n-octyl genistein.

PubMed

Ning, Yingxia; Luo, Chaoyuan; Ren, Kaiqun; Quan, Meifang; Cao, Jianguo

2014-05-01

Carcinogenesis is predominantly dependent on the cancer stem cells (CSCs) residing or populating within the cancer. We previously demonstrated that the novel synthetic genistein analogue, 7-difluoromethoxyl-5,4'-di-n-octylgenistein (DFOG), induced apoptotic cell death of ovarian and gastric cancer cells. The present study demonstrated that sphere‑forming cells (SFCs) derived from the ovarian cancer cell-line SKOV3 possessed ovarian cancer stem-like cell (OCSLC) properties, including self-renewal and high tumorigenicity. DFOG may be effective in inhibiting the self‑renewal capacity of SFCs derived from the SKOV3 cell line. DFOG decreased the level of phosphorylated FOXO3a protein in SKOV3 cell‑derived SFCs. The inhibition of FOXO3a expression by siRNA significantly attenuated the ability of DFOG to inhibit the self-renewal capacity of SKOV3-derived SFCs. Our results suggested that DFOG has been demonstrated to significantly inhibit the self-renewal capacity of ovarian cancer stem cells (OCSCs) through a mechanism partly dependent on the activation of FOXO3a.

Luteolin and fisetin suppress oxidative stress by modulating sirtuins and forkhead box O3a expression under in vitro diabetic conditions.

PubMed

Kim, Arang; Lee, Wooje; Yun, Jung-Mi

2017-10-01

Chronic hyperglycemia induces oxidative stress via accumulation of reactive oxygen species (ROS) and contributes to diabetic complications. Hyperglycemia induces mitochondrial superoxide anion production through the increased activity of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. This study aimed to determine whether fisetin and luteolin treatments suppress the oxidative stress by modulating the expression of sirtuins (SIRTs) and forkhead box O3a (FOXO3a) under hyperglycemic conditions in human monocytes. Human monocytic cells (THP-1) were cultured under osmotic control (14.5 mmol/L mannitol), normoglycemic (NG, 5.5 mmol/L glucose), or hyperglycemic (HG, 20 mmol/L glucose) conditions, in the absence or presence of fisetin and luteolin for 48 h. To determine the effect of fisetin and luteolin treatments on high glucose-induced oxidative stress, western blotting and intracellular staining were performed. Hyperglycemic conditions increased the ROS production, as compared to normoglycemic condition. However, fisetin and luteolin treatments inhibited ROS production under hyperglycemia. To obtain further insight into ROS production in hyperglycemic conditions, evaluation of p47phox expression revealed that fisetin and luteolin treatments inhibited p47phox expression under hyperglycemic conditions. Conversely, the expression levels of SIRT1, SIRT3, SIRT6, and FOXO3a were decreased under high glucose conditions compared to normal glucose conditions, but exposure to fisetin and luteolin induced the expression of SIRT1, SIRT3, SIRT6, and FOXO3a. The above findings suggest that fisetin and luteolin inhibited high glucose-induced ROS production in monocytes through the activation of SIRTs and FOXO3a. The results of our study supports current researches that state fisetin and luteolin as potential agents for the development of novel strategies for diabetes.

Luteolin and fisetin suppress oxidative stress by modulating sirtuins and forkhead box O3a expression under in vitro diabetic conditions

PubMed Central

Kim, Arang; Lee, Wooje

2017-01-01

BACKGROUND/OBJECTIVE Chronic hyperglycemia induces oxidative stress via accumulation of reactive oxygen species (ROS) and contributes to diabetic complications. Hyperglycemia induces mitochondrial superoxide anion production through the increased activity of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. This study aimed to determine whether fisetin and luteolin treatments suppress the oxidative stress by modulating the expression of sirtuins (SIRTs) and forkhead box O3a (FOXO3a) under hyperglycemic conditions in human monocytes. MATERIALS/METHODS Human monocytic cells (THP-1) were cultured under osmotic control (14.5 mmol/L mannitol), normoglycemic (NG, 5.5 mmol/L glucose), or hyperglycemic (HG, 20 mmol/L glucose) conditions, in the absence or presence of fisetin and luteolin for 48 h. To determine the effect of fisetin and luteolin treatments on high glucose-induced oxidative stress, western blotting and intracellular staining were performed. RESULTS Hyperglycemic conditions increased the ROS production, as compared to normoglycemic condition. However, fisetin and luteolin treatments inhibited ROS production under hyperglycemia. To obtain further insight into ROS production in hyperglycemic conditions, evaluation of p47phox expression revealed that fisetin and luteolin treatments inhibited p47phox expression under hyperglycemic conditions. Conversely, the expression levels of SIRT1, SIRT3, SIRT6, and FOXO3a were decreased under high glucose conditions compared to normal glucose conditions, but exposure to fisetin and luteolin induced the expression of SIRT1, SIRT3, SIRT6, and FOXO3a. The above findings suggest that fisetin and luteolin inhibited high glucose-induced ROS production in monocytes through the activation of SIRTs and FOXO3a. CONCLUSIONS The results of our study supports current researches that state fisetin and luteolin as potential agents for the development of novel strategies for diabetes. PMID:28989580

Cryptic insertion of 3'FOXO1 into inverted chromosome arm 2q in the presence of two normal chromosome 13s and 13 small interstitial duplications in a patient with alveolar rhabdomyosarcoma.

PubMed

Hackman, Sarah; Calvey, Laura; Bernreuter, Kristen; Mark, Mengya Wang; Starnes, Sarah; Batanian, Jacqueline R

2015-09-01

Alveolar rhabdomyosarcoma (ARMS) is a pediatric soft tissue neoplasm with a characteristic translocation, t(2;13)(q35;q14), which is detected in 70-80% of cases. This well-described translocation produces the gene fusion product PAX3-FOXO1. Cryptic rearrangements of this fusion have never before been reported in ARMS. Here we describe a patient with ARMS that showed, by fluorescence in situ hybridization and G-banded chromosomes, a cryptic insertion of 3'FOXO1 into inverted chromosome 2q. The inversion breakpoints were depicted by array comparative genomic hybridization as two small interstitial duplications, one of which involved the PAX3 gene. In addition, the array comparative genomic hybridization results revealed 1q gain, 16q loss, and 11 more small duplications, with one of them involving the FOXO1 gene. Although the pathogenesis in classic ARMS cases is thought to be driven by the 5'PAX3-3'FOXO1 fusion on derivative chromosome 13, here we report a novel cryptic insertion of 3'FOXO1 resulting in a pathogenic fusion with 5'PAX3 on inverted chromosome 2q. Copyright © 2015 Elsevier Inc. All rights reserved.

Overexpression of FOXO4 induces apoptosis of clear-cell renal carcinoma cells through downregulation of Bim.

PubMed

Wang, Wei; Zhou, Pang-Hu; Hu, Wei

2016-03-01

Forkhead box O4 (FOXO4) has been reported to be a novel tumor suppressor gene in gastrointestinal cancers; however, its role in clear‑cell renal carcinoma cells (ccRCC) has remained largely elusive. The present study assessed the expression levels of FOXO4 in RCC tissues and cells. Furthermore, the effects of vector‑mediated overexpression of FOXO4 on the apoptotic rate of the 786‑0 and Caki‑1 cell lines and the role of Bim in this process were investigated. The results demonstrated that the protein and mRNA expression levels of FOXO4 were decreased in renal cancer tissues and cell lines compared with those in normal tissues and cell lines. FOXO4 overexpression significantly increased the apoptotic rate of ccRCC cells in vitro, along with increased protein expression levels of Bim, cleaved‑caspase 3, B‑cell lymphoma 2 (Bcl‑2)‑associated X protein (Bax) and cytochrome c, as well as a decrease in Bcl‑2 expression. Of note, the apoptotic effects of FOXO4 overexpression in 786‑0 cells were inhibited by small interfering RNA‑mediated knockdown of Bim. The results of the present study indicated that FOXO4 has an inhibitory effect in ccRCC, at least in part through inducing apoptosis via upregulation of Bim in the mitochondria-dependent pathway.

PUMA promotes Bax translocation in FOXO3a-dependent pathway during STS-induced apoptosis

NASA Astrophysics Data System (ADS)

Zhang, Yingjie; Chen, Qun

2009-08-01

PUMA (p53 up-regulated modulator of apoptosis, also called Bbc3) was first identified as a BH3-only Bcl-2 family protein that is transcriptionally up-regulated by p53 and activated upon p53-dependent apoptotic stimuli, such as treatment with DNA-damaging drugs or UV irradiation. Recently studies have been shown that Puma is also up-regulated in response to certain p53-independent apoptotic stimuli, such as growth factor deprivation or treatment with glucocorticoids or STS (staurosporine). However, the molecular mechanisms of PUMA up-regulation and how PUMA functions in response to p53-independent apoptotic stimuli remain poorly understood. In this study, based on real-time single cell analysis, flow cytometry and western blotting technique, we investigated the function of PUMA in living human lung adenocarcinoma cells (ASTC-a-1) after STS treatment. Our results show that FOXO3a was activated by STS stimulation and then translocated from cytosol to nucleus. The expression of PUMA was up-regulated via a FOXO3a-dependent manner after STS treatment, while p53 had little function in this process. Moreover, cell apoptosis and Bax translocation induced by STS were not blocked by Pifithrin-α (p53 inhibitor), which suggested that p53 was not involved in this signaling pathway. Taken together, these results indicate that PUMA promoted Bax translocation in a FOXO3a-dependment pathway during STS-induced apoptosis, while p53 was dispensable in this process.

Inhibition of SH2-domain-containing inositol 5-phosphatase (SHIP2) ameliorates palmitate induced-apoptosis through regulating Akt/FOXO1 pathway and ROS production in HepG2 cells

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

Gorgani-Firuzjaee, Sattar; Adeli, Khosrow; Meshkani, Reza, E-mail: rmeshkani@tums.ac.ir

The serine–threonine kinase Akt regulates proliferation and survival by phosphorylating a network of protein substrates; however, the role of a negative regulator of the Akt pathway, the SH2-domain-containing inositol 5-phosphatase (SHIP2) in apoptosis of the hepatocytes, remains unknown. In the present study, we studied the molecular mechanisms linking SHIP2 expression to apoptosis using overexpression or suppression of SHIP2 gene in HepG2 cells exposed to palmitate (0.5 mM). Overexpression of the dominant negative mutant SHIP2 (SHIP2-DN) significantly reduced palmitate-induced apoptosis in HepG2 cells, as these cells had increased cell viability, decreased apoptotic cell death and reduced the activity of caspase-3, cytochrome cmore » and poly (ADP-ribose) polymerase. Overexpression of the wild-type SHIP2 gene led to a massive apoptosis in HepG2 cells. The protection from palmitate-induced apoptosis by SHIP2 inhibition was accompanied by a decrease in the generation of reactive oxygen species (ROS). In addition, SHIP2 inhibition was accompanied by an increased Akt and FOXO-1 phosphorylation, whereas overexpression of the wild-type SHIP2 gene had the opposite effects. Taken together, these findings suggest that SHIP2 expression level is an important determinant of hepatic lipoapotosis and its inhibition can potentially be a target in treatment of hepatic lipoapoptosis in diabetic patients. - Highlights: • Lipoapoptosis is the major contributor to the development of NAFLD. • The PI3-K/Akt pathway regulates apoptosis in different cells. • The role of negative regulator of this pathway, SHIP2 in lipoapoptosis is unknown. • SHIP2 inhibition significantly reduces palmitate-induced apoptosis in HepG2 cells. • SHIP2 inhibition prevents palmitate induced-apoptosis by regulating Akt/FOXO1 pathway.« less

FoxO regulates microtubule dynamics and polarity to promote dendrite branching in Drosophila sensory neurons

PubMed Central

Sears, James C.; Broihier, Heather T.

2016-01-01

The size and shape of dendrite arbors are defining features of neurons and critical determinants of neuronal function. The molecular mechanisms establishing arborization patterns during development are not well understood, though properly regulated microtubule (MT) dynamics and polarity are essential. We previously found that FoxO regulates axonal MTs, raising the question of whether it also regulates dendritic MTs and morphology. Here we demonstrate that FoxO promotes dendrite branching in all classes of Drosophila dendritic arborization (da) neurons. FoxO is required both for initiating growth of new branches and for maintaining existing branches. To elucidate FoxO function, we characterized MT organization in both foxO null and overexpressing neurons. We find that FoxO directs MT organization and dynamics in dendrites. Moreover, it is both necessary and sufficient for anterograde MT polymerization, which is known to promote dendrite branching. Lastly, FoxO promotes proper larval nociception, indicating a functional consequence of impaired da neuron morphology in foxO mutants. Together, our results indicate that FoxO regulates dendrite structure and function and suggest that FoxO-mediated pathways control MT dynamics and polarity. PMID:27546375

Insulin-like growth factor 1 rescues R28 retinal neurons from apoptotic death through ERK-mediated BimEL phosphorylation independent of Akt.

PubMed

Kong, Dejuan; Gong, Lijie; Arnold, Edith; Shanmugam, Sumathi; Fort, Patrice E; Gardner, Thomas W; Abcouwer, Steven F

2016-10-01

Insulin-like growth factor 1 (IGF-1) can provide long-term neurotrophic support by activation of Akt, inhibition of FoxO nuclear localization and suppression of Bim gene transcription in multiple neuronal systems. However, MEK/ERK activation can also promote neuron survival through phosphorylation of BimEL. We explored the contribution of the PI3K/Akt/FoxO and MEK/ERK/BimEL pathways in IGF-1 stimulated survival after serum deprivation (SD) of R28 cells differentiated to model retinal neurons. IGF-1 caused rapid activation of Akt leading to FoxO1/3-T32/T24 phosphorylation, and prevented FoxO1/3 nuclear translocation and Bim mRNA upregulation in response to SD. IGF-1 also caused MAPK/MEK pathway activation as indicated by ERK1/2-T202/Y204 and Bim-S65 phosphorylation. Overexpression of FoxO1 increased Bim mRNA expression and amplified the apoptotic response to SD without shifting the serum response curve. Inhibition of Akt activation with LY294002 or by Rictor knockdown did not block the protective effect of IGF-1, while inhibition of MEK activity with PD98059 prevented Bim phosphorylation and blocked IGF-1 protection. In addition, knockdown of Bim expression was protective during SD, while co-silencing of FoxO1 and Fox03 expression had little effect. Thus, the PI3K/Akt/FoxO pathway was not essential for protection from SD-induced apoptosis by IGF-1 in R28 cells. Instead, IGF-1 protection was dependent on activation of the MEK/ERK pathway leading to BimEL phosphorylation, which is known to prevent Bax/Bak oligomerization and activation of the intrinsic mitochondrial apoptosis pathway. These studies demonstrate the requirement of the MEK/ERK pathway in a model of retinal neuron cell survival and highlight the cell specificity for IGF-1 signaling in this response. Copyright © 2016 Elsevier Ltd. All rights reserved.

Insulin-like Growth Factor 1 Rescues R28 Retinal Neurons from Apoptotic Death through ERK-mediated BimEL Phosphorylation Independent of Akt

PubMed Central

Kong, Dejuan; Gong, Lijie; Arnold, Edith; Shanmugam, Sumathi; Fort, Patrice E.; Gardner, Thomas W.; Abcouwer, Steven F.

2016-01-01

Insulin-like growth factor 1 (IGF-1) can provide long-term neurotrophic support by activation of Akt, inhibition of FoxO nuclear localization and suppression of Bim gene transcription in multiple neuronal systems. However, MEK/ERK activation can also promote neuron survival through phosphorylation of BimEL. We explored the contribution of the PI3K/Akt/FoxO and MEK/ERK/BimEL pathways in IGF-1 stimulated survival after serum deprivation (SD) of R28 cells differentiated to model retinal neurons. IGF-1 caused rapid activation of Akt leading to FoxO1/3-T32/T24 phosphorylation, and prevented FoxO1/3 nuclear translocation and Bim mRNA upregulation in response to SD. IGF-1 also caused MAPK/MEK pathway activation as indicated by ERK1/2-T202/Y204 and Bim-S65 phosphorylation. Overexpression of FoxO1 increased Bim mRNA expression and amplified the apoptotic response to SD without shifting the serum response curve. Inhibition of Akt activation with LY294002 or by Rictor knockdown did not block the protective effect of IGF-1, while inhibition of MEK activity with PD98059 prevented Bim phosphorylation and blocked IGF-1 protection. In addition, knockdown of Bim expression was protective during SD, while co-silencing of FoxO1 and Fox03 expression had little effect. Thus, the PI3K/Akt/FoxO pathway was not essential for protection from SD-induced apoptosis by IGF-1 in R28 cells. Instead, IGF-1 protection was dependent on activation of the MEK/ERK pathway leading to BimEL phosphorylation, which is known to prevent Bax/Bak oligomerization and activation of the intrinsic mitochondrial apoptosis pathway. These studies demonstrate the requirement of the MEK/ERK pathway in a model of retinal neuron cell survival and highlight the cell specificity for IGF-1 signaling in this response. PMID:27511131

Simvastatin prevents triple-negative breast cancer metastasis in pre-clinical models through regulation of FOXO3a.

PubMed

Wolfe, Adam R; Debeb, Bisrat G; Lacerda, Lara; Larson, Richard; Bambhroliya, Arvind; Huang, Xuelin; Bertucci, Francois; Finetti, Pascal; Birnbaum, Daniel; Van Laere, Steven; Diagaradjan, Parmeswaran; Ruffell, Brian; Trenton, Nicholaus J; Chu, Khoi; Hittelman, Walter; Diehl, Michael; Levental, Ilya; Ueno, Naoto T; Woodward, Wendy A

2015-12-01

We previously reported using statins was correlated with improved metastasis-free survival in aggressive breast cancer. The purpose of this study was to examine the effect of statins on metastatic colonization by triple-negative breast cancer (TNBC) cells. TNBC cell lines were treated with simvastatin and then studied for cell cycle progression and proliferation in vitro, and metastasis formation in vivo, following injection of statin-treated cells. Reverse-phase protein assay (RPPA) analysis was performed on statin-treated and control breast cancer cells. RNA interference targeting FOXO3a was used to measure the impact of simvastatin on FOXO3a-expressing cells. The prognostic value of FOXO3a mRNA expression was examined in eight public breast cancer gene expression datasets including 1479 patients. Simvastatin increased G1/S-phase arrest of the cell cycle and inhibited both proliferation and migration of TNBC cells in vitro. In vitro pre-treatment and in vivo treatment with simvastatin reduced metastases. Phosphorylated FOXO3a was downregulated after simvastatin treatment in (RPPA) analysis. Ectopic expression of FOXO3a enhanced mammosphere formation and migratory capacity in vitro. Knockdown of FOXO3a attenuated the effect of simvastatin on mammosphere formation and migration. Analysis of public gene expression data demonstrates FOXO3a mRNA downregulation was independently associated with shorter metastasis-free survival in all breast cancers, as well as in TNBC breast cancers. Simvastatin inhibits in vitro endpoints associated with metastasis through a FOXO3a mechanism and reduced metastasis formation in vivo. FOXO3a expression is prognostic for metastasis formation in patient data. Further investigation of simvastatin as a cancer therapy is warranted.

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

Park, Joo-Man; Kim, Tae-Hyun; Brain Korea 21 Project for Medical Sciences, Yonsei University College of Medicine, Seoul 120-752

Research highlights: {yields} Insulin-suppression of PEPCK and G6Pase gene expression is counteracted by resveratrol. {yields} Resveratrol upregulates hepatic gluconeogenic genes by attenuating insulin signaling and deacetylating FOXO1, which are SIRT1-independent in the cytosol and SIRT1-dependent in the nucleus, respectively. {yields} Resveratrol increases the binding activity of Foxo1 to the IRE of PEPCK and G6Pase. -- Abstract: During a state of fasting, the blood glucose level is maintained by hepatic gluconeogenesis. SIRT1 is an important metabolic regulator during nutrient deprivation and the liver-specific knockdown of SIRT1 resulted in decreased glucose production. We hypothesize that SIRT1 is responsible for the upregulation ofmore » insulin-suppressed gluconeogenic genes through the deacetylation of FOXO1. Treatment of primary cultured hepatocytes with resveratrol increased insulin-repressed PEPCK and G6Pase mRNA levels, which depend on SIRT1 activity. We found that the resveratrol treatment resulted in a decrease in the phosphorylation of Akt and FOXO1, which are independent of SIRT1 action. Fluorescence microscopy revealed that resveratrol caused the nuclear localization of FOXO1. In the nucleus, FOXO1 is deacetylated by SIRT1, which might make it more accessible to the IRE of the PEPCK and G6Pase promoter, causing an increase in their gene expression. Our results indicate that resveratrol upregulates the expression of gluconeogenic genes by attenuating insulin signaling and by deacetylating FOXO1, which are SIRT1-independent in the cytosol and SIRT1-dependent in the nucleus, respectively.« less

Activation of the ζ receptor 1 suppresses NMDA responses in rat retinal ganglion cells.

PubMed

Zhang, X-J; Liu, L-L; Jiang, S-X; Zhong, Y-M; Yang, X-L

2011-03-17

The sigma receptor 1 (σR1) has been shown to modulate the activity of several voltage- and ligand-gated channels. Using patch-clamp techniques in rat retinal slice preparations, we demonstrated that activation of σR1 by SKF10047 (SKF) or PRE-084 suppressed N-methyl-D-aspartate (NMDA) receptor-mediated current responses from both ON and OFF type ganglion cells (GCs), dose-dependently, and the effect could be blocked by the σR1 antagonist BD1047 or the σR antagonist haloperidol. The suppression by SKF of NMDA currents was abolished with pre-incubation of the G protein inhibitor GDP-β-S or the Gi/o activator mastoparan. We further explored the intracellular signaling pathway responsible for the SKF-induced suppression of NMDA responses. Application of either cAMP/the PKA inhibitor Rp-cAMP or cGMP/the PKG inhibitor KT5823 did not change the SKF-induced effect, suggesting the involvement of neither cAMP/PKA nor cGMP/PKG pathway. In contrast, suppression of NMDA responses by SKF was abolished by internal infusion of the phosphatidylinostiol-specific phospholipase C (PLC) inhibitor U73122, but not by the phosphatidylcholine-PLC inhibitor D609. SKF-induced suppression of NMDA responses was dependent on intracellular Ca2+ concentration ([Ca2+]i), as evidenced by the fact that the effect was abolished when [Ca2+]i was buffered with 10 mM BAPTA. The SKF effect was blocked by xestospongin-C/heparin, IP3 receptor antagonists, but unchanged by ryanodine/caffeine, ryanodine receptor modulators. Furthermore, application of protein kinase C inhibitors Bis IV and Gö6976 eliminated the SKF effect. These results suggest that the suppression of NMDA responses of rat retinal GCs caused by the activation of σR1 may be mediated by a distinct [Ca2+]i-dependent PLC-PKC pathway. This effect of SKF could help ameliorate malfunction of GCs caused by excessive stimulation of NMDA receptors under pathological conditions. Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights

CREB and FoxO1: two transcription factors for the regulation of hepatic gluconeogenesis

PubMed Central

Oh, Kyoung-Jin; Han, Hye-Sook; Kim, Min-Jung; Koo, Seung-Hoi

2013-01-01

Liver plays a major role in maintaining glucose homeostasis in mammals. Under fasting conditions, hepatic glucose production is critical as a source of fuel to maintain the basic functions in other tissues, including skeletal muscle, red blood cells, and the brain. Fasting hormones glucagon and cortisol play major roles during the process, in part by activating the transcription of key enzyme genes in the gluconeogenesis such as phosphoenol pyruvate carboxykinase (PEPCK) and glucose 6 phosphatase catalytic subunit (G6Pase). Conversely, gluconeogenic transcription is repressed by pancreatic insulin under feeding conditions, which effectively inhibits transcriptional activator complexes by either promoting post-translational modifications or activating transcriptional inhibitors in the liver, resulting in the reduction of hepatic glucose output. The transcriptional regulatory machineries have been highlighted as targets for type 2 diabetes drugs to control glycemia, so understanding of the complex regulatory mechanisms for transcription circuits for hepatic gluconeogenesis is critical in the potential development of therapeutic tools for the treatment of this disease. In this review, the current understanding regarding the roles of two key transcriptional activators, CREB and FoxO1, in the regulation of hepatic gluconeogenic program is discussed. [BMB Reports 2013; 46(12): 567-574] PMID:24238363

Ion channel TRPV1-dependent activation of PTP1B suppresses EGFR-associated intestinal tumorigenesis

PubMed Central

de Jong, Petrus R.; Takahashi, Naoki; Harris, Alexandra R.; Lee, Jihyung; Bertin, Samuel; Jeffries, James; Jung, Michael; Duong, Jen; Triano, Amy I.; Lee, Jongdae; Niv, Yaron; Herdman, David S.; Taniguchi, Koji; Kim, Chang-Whan; Dong, Hui; Eckmann, Lars; Stanford, Stephanie M.; Bottini, Nunzio; Corr, Maripat; Raz, Eyal

2014-01-01

The intestinal epithelium has a high rate of turnover, and dysregulation of pathways that regulate regeneration can lead to tumor development; however, the negative regulators of oncogenic events in the intestinal epithelium are not fully understood. Here we identified a feedback loop between the epidermal growth factor receptor (EGFR), a known mediator of proliferation, and the transient receptor potential cation channel, subfamily V, member 1 (TRPV1), in intestinal epithelial cells (IECs). We found that TRPV1 was expressed by IECs and was intrinsically activated upon EGFR stimulation. Subsequently, TRPV1 activation inhibited EGFR-induced epithelial cell proliferation via activation of Ca2+/calpain and resulting activation of protein tyrosine phosphatase 1B (PTP1B). In a murine model of multiple intestinal neoplasia (ApcMin/+ mice), TRPV1 deficiency increased adenoma formation, and treatment of these animals with an EGFR kinase inhibitor reversed protumorigenic phenotypes, supporting a functional association between TRPV1 and EGFR signaling in IECs. Administration of a TRPV1 agonist suppressed intestinal tumorigenesis in ApcMin/+ mice, similar to — as well as in conjunction with — a cyclooxygenase-2 (COX-2) inhibitor, which suggests that targeting both TRPV1 and COX-2 has potential as a therapeutic approach for tumor prevention. Our findings implicate TRPV1 as a regulator of growth factor signaling in the intestinal epithelium through activation of PTP1B and subsequent suppression of intestinal tumorigenesis. PMID:25083990

The transcription factor Prep1 controls hepatic insulin sensitivity and gluconeogenesis by targeting nuclear localization of FOXO1.

PubMed

Kulebyakin, Konstantin; Penkov, Dmitry; Blasi, Francesco; Akopyan, Zhanna; Tkachuk, Vsevolod

2016-12-02

Liver plays a key role in controlling body carbohydrate homeostasis by switching between accumulation and production of glucose and this way maintaining constant level of glucose in blood. Increased blood glucose level triggers release of insulin from pancreatic β-cells. Insulin represses hepatic glucose production and increases glucose accumulation. Insulin resistance is the main cause of type 2 diabetes and hyperglycemia. Currently thiazolidinediones (TZDs) targeting transcriptional factor PPARγ are used as insulin sensitizers for treating patients with type 2 diabetes. However, TZDs are reported to be associated with cardiovascular and liver problems and stimulate obesity. Thus, it is necessary to search new approaches to improve insulin sensitivity. A promising candidate is transcriptional factor Prep1, as it was shown earlier it could affect insulin sensitivity in variety of insulin-sensitive tissues. The aim of the present study was to evaluate a possible involvement of transcriptional factor Prep1 in control of hepatic glucose accumulation and production. We created mice with liver-specific Prep1 knockout and discovered that hepatocytes derived from these mice are much more sensitive to insulin, comparing to their WT littermates. Incubation of these cells with 100 nM insulin results in almost complete inhibition of gluconeogenesis, while in WT cells this repression is only partial. However, Prep1 doesn't affect gluconeogenesis in the absence of insulin. Also, we observed that nuclear content of gluconeogenic transcription factor FOXO1 was greatly reduced in Prep1 knockout hepatocytes. These findings suggest that Prep1 may control hepatic insulin sensitivity by targeting FOXO1 nuclear stability. Copyright © 2016 Elsevier Inc. All rights reserved.

Prolactin Signaling through the Short Form of Its Receptor Represses Forkhead Transcription Factor FOXO3 and Its Target Gene Galt Causing a Severe Ovarian Defect

PubMed Central

Halperin, Julia; Devi, Sangeeta Y.; Elizur, Shai; Stocco, Carlos; Shehu, Aurora; Rebourcet, Diane; Unterman, Terry G.; Leslie, Nancy D.; Le, Jamie; Binart, Nadine; Gibori, Geula

2008-01-01

Prolactin (PRL) is a hormone with over 300 biological activities. Although the signaling pathway downstream of the long form of its receptor (RL) has been well characterized, little is known about PRL actions upon activation of the short form (RS). Here, we show that mice expressing only RS exhibit an ovarian phenotype of accelerated follicular recruitment followed by massive follicular death leading to premature ovarian failure. Consequently, RS-expressing ovaries of young adults are depleted of functional follicles and formed mostly by interstitium. We also show that activation of RS represses the expression of the transcription factor Forkhead box O3 (FOXO3) and that of the enzyme galactose-1-phosphate uridyltransferase (Galt), two proteins known to be essential for normal follicular development. Our finding that FOXO3 regulates the expression of Galt and enhances its transcriptional activity indicates that it is the repression of FOXO3 by PRL acting through RS that prevents Galt expression in the ovary and causes follicular death. Coexpression of RL with RS prevents PRL inhibition of Galt, and the ovarian defect is no longer seen in RS transgenic mice that coexpress RL, suggesting that RL prevents RS-induced ovarian impairment. In summary, we show that PRL signals through RS and causes, in the absence of RL, a severe ovarian pathology by repressing the expression of FOXO3 and that of its target gene Galt. We also provide evidence of a link between the premature ovarian failure seen in mice expressing RS and in mice with FOXO3 gene deletion as well as in human with Galt mutation. PMID:17975019

Sphingosine 1-phosphate (S1P) suppresses the collagen-induced activation of human platelets via S1P4 receptor.

PubMed

Onuma, Takashi; Tanabe, Kumiko; Kito, Yuko; Tsujimoto, Masanori; Uematsu, Kodai; Enomoto, Yukiko; Matsushima-Nishiwaki, Rie; Doi, Tomoaki; Nagase, Kiyoshi; Akamatsu, Shigeru; Tokuda, Haruhiko; Ogura, Shinji; Iwama, Toru; Kozawa, Osamu; Iida, Hiroki

2017-08-01

Sphingosine 1-phosphate (S1P) is as an extracellular factor that acts as a potent lipid mediator by binding to specific receptors, S1P receptors (S1PRs). However, the precise role of S1P in human platelets that express S1PRs has not yet been fully clarified. We previously reported that heat shock protein 27 (HSP27) is released from human platelets accompanied by its phosphorylation stimulated by collagen. In the present study, we investigated the effect of S1P on the collagen-induced platelet activation. S1P pretreatment markedly attenuated the collagen-induced aggregation. Co-stimulation with S1P and collagen suppressed collagen-induced platelet activation, but the effect was weaker than that of S1P-pretreatment. The collagen-stimulated secretion of platelet-derived growth factor (PDGF)-AB and the soluble CD40 ligand (sCD40L) release were significantly reduced by S1P. In addition, S1P suppressed the collagen-induced release of HSP27 as well as the phosphorylation of HSP27. S1P significantly suppressed the collagen-induced phosphorylation of p38 mitogen-activated protein kinase. S1P increased the levels of GTP-bound Gαi and GTP-bound Gα13 coupled to S1PPR1 and/or S1PR4. CYM50260, a selective S1PR4 agonist, but not SEW2871, a selective S1PR1 agonist, suppressed the collagen-stimulated platelet aggregation, PDGF-AB secretion and sCD40L release. In addition, CYM50260 reduced the release of phosphorylated-HSP27 by collagen as well as the phosphorylation of HSP27. The selective S1PR4 antagonist CYM50358, which failed to affect collagen-induced HSP27 phosphorylation, reversed the S1P-induced attenuation of HSP27 phosphorylation by collagen. These results strongly suggest that S1P inhibits the collagen-induced human platelet activation through S1PR4 but not S1PR1. Copyright © 2017 Elsevier Ltd. All rights reserved.

Chronic innate immune activation of TBK1 suppresses mTORC1 activity and dysregulates cellular metabolism.

PubMed

Hasan, Maroof; Gonugunta, Vijay K; Dobbs, Nicole; Ali, Aktar; Palchik, Guillermo; Calvaruso, Maria A; DeBerardinis, Ralph J; Yan, Nan

2017-01-24

Three-prime repair exonuclease 1 knockout (Trex1 -/- ) mice suffer from systemic inflammation caused largely by chronic activation of the cyclic GMP-AMP synthase-stimulator of interferon genes-TANK-binding kinase-interferon regulatory factor 3 (cGAS-STING-TBK1-IRF3) signaling pathway. We showed previously that Trex1-deficient cells have reduced mammalian target of rapamycin complex 1 (mTORC1) activity, although the underlying mechanism is unclear. Here, we performed detailed metabolic analysis in Trex1 -/- mice and cells that revealed both cellular and systemic metabolic defects, including reduced mitochondrial respiration and increased glycolysis, energy expenditure, and fat metabolism. We also genetically separated the inflammatory and metabolic phenotypes by showing that Sting deficiency rescued both inflammatory and metabolic phenotypes, whereas Irf3 deficiency only rescued inflammation on the Trex1 -/- background, and many metabolic defects persist in Trex1 -/- Irf3 -/- cells and mice. We also showed that Leptin deficiency (ob/ob) increased lipogenesis and prolonged survival of Trex1 -/- mice without dampening inflammation. Mechanistically, we identified TBK1 as a key regulator of mTORC1 activity in Trex1 -/- cells. Together, our data demonstrate that chronic innate immune activation of TBK1 suppresses mTORC1 activity, leading to dysregulated cellular metabolism.

Synergic chemoprevention with dietary carbohydrate restriction and supplementation of AMPK-activating phytochemicals: the role of SIRT1.

PubMed