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Sample records for regulators mediating powdery

  1. Temporal global expression data reveal known and novel salicylate-impacted processes and regulators mediating powdery mildew growth and reproduction on Arabidopsis.

    PubMed

    Chandran, Divya; Tai, Yu Chuan; Hather, Gregory; Dewdney, Julia; Denoux, Carine; Burgess, Diane G; Ausubel, Frederick M; Speed, Terence P; Wildermuth, Mary C

    2009-03-01

    Salicylic acid (SA) is a critical mediator of plant innate immunity. It plays an important role in limiting the growth and reproduction of the virulent powdery mildew (PM) Golovinomyces orontii on Arabidopsis (Arabidopsis thaliana). To investigate this later phase of the PM interaction and the role played by SA, we performed replicated global expression profiling for wild-type and SA biosynthetic mutant isochorismate synthase1 (ics1) Arabidopsis from 0 to 7 d after infection. We found that ICS1-impacted genes constitute 3.8% of profiled genes, with known molecular markers of Arabidopsis defense ranked very highly by the multivariate empirical Bayes statistic (T(2) statistic). Functional analyses of T(2)-selected genes identified statistically significant PM-impacted processes, including photosynthesis, cell wall modification, and alkaloid metabolism, that are ICS1 independent. ICS1-impacted processes include redox, vacuolar transport/secretion, and signaling. Our data also support a role for ICS1 (SA) in iron and calcium homeostasis and identify components of SA cross talk with other phytohormones. Through our analysis, 39 novel PM-impacted transcriptional regulators were identified. Insertion mutants in one of these regulators, PUX2 (for plant ubiquitin regulatory X domain-containing protein 2), results in significantly reduced reproduction of the PM in a cell death-independent manner. Although little is known about PUX2, PUX1 acts as a negative regulator of Arabidopsis CDC48, an essential AAA-ATPase chaperone that mediates diverse cellular activities, including homotypic fusion of endoplasmic reticulum and Golgi membranes, endoplasmic reticulum-associated protein degradation, cell cycle progression, and apoptosis. Future work will elucidate the functional role of the novel regulator PUX2 in PM resistance. PMID:19176722

  2. Histo-chemical and biochemical analysis reveals association of er1 mediated powdery mildew resistance and redox balance in pea.

    PubMed

    Mohapatra, Chinmayee; Chand, Ramesh; Navathe, Sudhir; Sharma, Sandeep

    2016-09-01

    Powdery mildew caused by Erysiphe pisi is one of the important diseases responsible for heavy yield losses in pea crop worldwide. The most effective method of controlling the disease is the use of resistant varieties. The resistance to powdery mildew in pea is recessive and governed by a single gene er1. The objective of present study is to investigate if er1 mediated powdery mildew resistance is associated with changes in the redox status of the pea plant. 16 pea genotypes were screened for powdery mildew resistance in field condition for two years and, also, analyzed for the presence/absence of er1 gene. Histochemical analysis with DAB and NBT staining indicates accumulation of reactive oxygen species (ROS) in surrounding area of powdery mildew infection which was higher in susceptible genotypes as compared to resistant genotypes. A biochemical study revealed that the activity of superoxide dismutase (SOD) and catalase, enzymes involved in scavenging ROS, was increased in, both, resistant and susceptible genotypes after powdery mildew infection. However, both enzymes level was always higher in resistant than susceptible genotypes throughout time course of infection. Moreover, irrespective of any treatment, the total phenol (TP) and malondialdehyde (MDA) content was significantly high and low in resistant genotypes, respectively. The powdery mildew infection elevated the MDA content but decreased the total phenol in pea genotypes. Statistical analysis showed a strong positive correlation between AUDPC and MDA; however, a negative correlation was observed between AUDPC and SOD, CAT and TP. Heritability of antioxidant was also high. The study identified few novel genotypes resistant to powdery mildew infection that carried the er1 gene and provided further clue that er1 mediated defense response utilizes antioxidant machinery to confer powdery mildew resistance in pea. PMID:27135819

  3. Mla- and Rom1-mediated control of microRNA398 and chloroplast copper/zinc superoxide dismutase regulates cell death in response to the barley powdery mildew fungus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Barley Mla (Mildew resistance locus a) confers allele-specific interactions with natural variants of the ascomycete fungus, Blumeria graminis f. sp. hordei (Bgh), causal agent of powdery mildew disease. Significant reprogramming of host gene expression occurs upon infection by this obligate biotrop...

  4. Gene-For-Gene-Mediated Transcriptome Reprogramming in Barley-Powdery Mildew Interactions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Barley has a complex interaction with powdery mildew [Blumeria graminis f. sp. hordei (Bgh)] that begins with early recognition of microbe-associated molecular patterns (MAMPs) from the pathogen. During Bgh invasion of the epidermis, the fate of cells is decided by the presence of resistance (R) gen...

  5. Temperature regulates the initiation of cleistothecia in powdery mildew of strawberry

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The formation of cleistothecia by the strawberry powdery mildew pathogen (Podosphaera aphanis) is widespread, but often sporadic throughout the range of strawberry cultivation. In some production regions, notably in warmer climates, they are reportedly rare. We confirmed that the pathogen is heterot...

  6. Coexpression network analysis of the genes regulated by two types of resistance responses to powdery mildew in wheat.

    PubMed

    Zhang, Juncheng; Zheng, Hongyuan; Li, Yiwen; Li, Hongjie; Liu, Xin; Qin, Huanju; Dong, Lingli; Wang, Daowen

    2016-01-01

    Powdery mildew disease caused by Blumeria graminis f. sp. tritici (Bgt) inflicts severe economic losses in wheat crops. A systematic understanding of the molecular mechanisms involved in wheat resistance to Bgt is essential for effectively controlling the disease. Here, using the diploid wheat Triticum urartu as a host, the genes regulated by immune (IM) and hypersensitive reaction (HR) resistance responses to Bgt were investigated through transcriptome sequencing. Four gene coexpression networks (GCNs) were developed using transcriptomic data generated for 20 T. urartu accessions showing IM, HR or susceptible responses. The powdery mildew resistance regulated (PMRR) genes whose expression was significantly correlated with Bgt resistance were identified, and they tended to be hubs and enriched in six major modules. A wide occurrence of negative regulation of PMRR genes was observed. Three new candidate immune receptor genes (TRIUR3_13045, TRIUR3_01037 and TRIUR3_06195) positively associated with Bgt resistance were discovered. Finally, the involvement of TRIUR3_01037 in Bgt resistance was tentatively verified through cosegregation analysis in a F2 population and functional expression assay in Bgt susceptible leaf cells. This research provides insights into the global network properties of PMRR genes. Potential molecular differences between IM and HR resistance responses to Bgt are discussed. PMID:27033636

  7. Coexpression network analysis of the genes regulated by two types of resistance responses to powdery mildew in wheat

    PubMed Central

    Zhang, Juncheng; Zheng, Hongyuan; Li, Yiwen; Li, Hongjie; Liu, Xin; Qin, Huanju; Dong, Lingli; Wang, Daowen

    2016-01-01

    Powdery mildew disease caused by Blumeria graminis f. sp. tritici (Bgt) inflicts severe economic losses in wheat crops. A systematic understanding of the molecular mechanisms involved in wheat resistance to Bgt is essential for effectively controlling the disease. Here, using the diploid wheat Triticum urartu as a host, the genes regulated by immune (IM) and hypersensitive reaction (HR) resistance responses to Bgt were investigated through transcriptome sequencing. Four gene coexpression networks (GCNs) were developed using transcriptomic data generated for 20 T. urartu accessions showing IM, HR or susceptible responses. The powdery mildew resistance regulated (PMRR) genes whose expression was significantly correlated with Bgt resistance were identified, and they tended to be hubs and enriched in six major modules. A wide occurrence of negative regulation of PMRR genes was observed. Three new candidate immune receptor genes (TRIUR3_13045, TRIUR3_01037 and TRIUR3_06195) positively associated with Bgt resistance were discovered. Finally, the involvement of TRIUR3_01037 in Bgt resistance was tentatively verified through cosegregation analysis in a F2 population and functional expression assay in Bgt susceptible leaf cells. This research provides insights into the global network properties of PMRR genes. Potential molecular differences between IM and HR resistance responses to Bgt are discussed. PMID:27033636

  8. Powdery Scab

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery scab (PS) is incited by a member of the Plamodiophorales, or parasitic slime molds, Spongospora subterranea. It invades the roots forming multinucleate plasmodia, release zoospores, and forms galls on the roots and lesions on the tuber skin. It is an emerging disease in the Pacific Northwe...

  9. Powdery Mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew of beet, a disease caused by Erysiphe polygoni, is a problem in most beet growing areas. Symptoms include whitish mats of hyphae and conidia that can cover leaves and flower stalks and cause leaves to turn yellow, then purplish brown. Characteristics of the pathogen, epidemiology and...

  10. Powdery mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew is found sporadically on alfalfa and annual medic species. It has been reported on commercial hay and seed fields in Europe, the People's Republic of China, Egypt, and the U.S. It is a common foliar disease in greenhouse-grown plants. Severe infections can cause defoliation and redu...

  11. Powdery Mildew Resistance Conferred by Loss of the ENHANCED DISEASE RESISTANCE1 Protein Kinase Is Suppressed by a Missense Mutation in KEEP ON GOING, a Regulator of Abscisic Acid Signaling1[W][OA

    PubMed Central

    Wawrzynska, Anna; Christiansen, Katy M.; Lan, Yinan; Rodibaugh, Natalie L.; Innes, Roger W.

    2008-01-01

    Loss-of-function mutations in the Arabidopsis (Arabidopsis thaliana) ENHANCED DISEASE RESISTANCE1 (EDR1) gene confer enhanced resistance to infection by powdery mildew (Golovinomyces cichoracearum). EDR1 encodes a protein kinase, but its substrates and the pathways regulated by EDR1 are unknown. To identify components of the EDR1 signal transduction pathway(s), we conducted a forward genetic screen for mutations that suppressed edr1-mediated disease resistance. Genetic mapping and cloning of one of these suppressor mutations revealed a recessive missense mutation in the KEEP ON GOING gene (KEG; At5g13530), which we designated keg-4. KEG encodes a multidomain protein that includes a RING E3 ligase domain, a kinase domain, ankyrin repeats, and HERC2-like repeats. The KEG protein has previously been shown to have ubiquitin ligase activity and to negatively regulate protein levels of the transcription factor ABCISIC ACID INSENSITIVE5. KEG mRNA levels were found to be 3-fold higher in edr1 mutant plants compared to wild type. Loss-of-function mutations in KEG are seedling lethal and are hypersensitive to glucose and abscisic acid (ABA). The keg-4 mutation, in contrast, conferred resistance to 6% glucose and suppressed edr1-mediated hypersensitivity to ABA, suggesting that the keg-4 mutation suppresses ABA signaling by altering KEG function. Several ABA-responsive genes were found to be further up-regulated in the edr1 mutant following ABA treatment, and this up-regulation was suppressed by the keg-4 mutation. We conclude that edr1-mediated resistance to powdery mildew is mediated, in part, by enhanced ABA signaling. PMID:18815384

  12. The Novel Gene VpPR4-1 from Vitis pseudoreticulata Increases Powdery Mildew Resistance in Transgenic Vitis vinifera L.

    PubMed Central

    Dai, Lingmin; Wang, Dan; Xie, Xiaoqing; Zhang, Chaohong; Wang, Xiping; Xu, Yan; Wang, Yuejin; Zhang, Jianxia

    2016-01-01

    Pathogenesis-related proteins (PRs) can lead to increased resistance of the whole plant to pathogen attack. Here, we isolate and characterize a PR-4 protein (VpPR4-1) from a wild Chinese grape Vitis pseudoreticulata which shows greatly elevated transcription following powdery mildew infection. Its expression profiles under a number of abiotic stresses were also investigated. Powdery mildew, salicylic acid, and jasmonic acid methyl ester significantly increased the VpPR4-1 induction while NaCl and heat treatments just slightly induced VpPR4-1 expression. Abscisic acid and cold treatment slightly affected the expression level of VpPR4-1. The VpPR4-1 gene was overexpressed in 30 regenerated V. vinifera cv. Red Globe via Agrobacterium tumefaciens-mediated transformation and verified by the Western blot. The 26 transgenic grapevines exhibited higher expression levels of PR-4 protein content than wild-type vines and six of them were inoculated with powdery mildew which showed that the growth of powdery mildew was repressed. The powdery mildew-resistance of Red Globe transformed with VpPR4-1 was enhanced inoculated with powdery mildew. Moreover, other powdery mildew resistant genes were associated with feedback regulation since VpPR4-1 is in abundance. This study demonstrates that PR-4 protein in grapes plays a vital role in defense against powdery mildew invasion. PMID:27303413

  13. Flor revisited (again): eQTL and mutational analysis of NB-LRR mediated immunity to powdery mildew in barley

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genes encoding early signaling events in pathogen defense often are identified only by their phenotype. Such genes involved in barley-powdery mildew interactions include Mla, specifying race-specific resistance; Rar1 (Required for Mla12-specified resistance1), and Rom1 (Restoration of Mla-specified...

  14. The knottin-like Blufensin family regulates genes involved in nuclear import and the secretory pathway in barley-powdery mildew interactions

    PubMed Central

    Xu, Weihui; Meng, Yan; Surana, Priyanka; Fuerst, Greg; Nettleton, Dan; Wise, Roger P.

    2015-01-01

    Plants have evolved complex regulatory mechanisms to control a multi-layered defense response to microbial attack. Both temporal and spatial gene expression are tightly regulated in response to pathogen ingress, modulating both positive and negative control of defense. BLUFENSINs, small knottin-like peptides in barley, wheat, and rice, are highly induced by attack from fungal pathogens, in particular, the obligate biotrophic fungus, Blumeria graminis f. sp. hordei (Bgh), causal agent of barley powdery mildew. Previous research indicated that Blufensin1 (Bln1) functions as a negative regulator of basal defense mechanisms. In the current report, we show that BLN1 and BLN2 can both be secreted to the apoplast and Barley stripe mosaic virus (BSMV)-mediated overexpression of Bln2 increases susceptibility of barley to Bgh. Bimolecular fluorescence complementation (BiFC) assays signify that BLN1 and BLN2 can interact with each other, and with calmodulin. We then used BSMV-induced gene silencing to knock down Bln1, followed by Barley1 GeneChip transcriptome analysis, to identify additional host genes influenced by Bln1. Analysis of differential expression revealed a gene set enriched for those encoding proteins annotated to nuclear import and the secretory pathway, particularly Importin α1-b and Sec61 γ subunits. Further functional analysis of these two affected genes showed that when silenced, they also reduced susceptibility to Bgh. Taken together, we postulate that Bln1 is co-opted by Bgh to facilitate transport of disease-related host proteins or effectors, influencing the establishment of Bgh compatibility on its barley host. PMID:26089830

  15. Powdery mildew of chickpea

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew of chickpea is caused by several fungi including Leveillula taurica, Erysiphe pisi and possible other Erysiphe spp. The most conspicuous sign of infection is diffuse, powdery sporulation on leaves and pods. The disease is favored by cool and dry weather. The relative roles of the sex...

  16. Silicon-mediated resistance of Arabidopsis against powdery mildew involves mechanisms other than the salicylic acid (SA)-dependent defence pathway.

    PubMed

    Vivancos, Julien; Labbé, Caroline; Menzies, James G; Bélanger, Richard R

    2015-08-01

    On absorption by plants, silicon (Si) offers protection against many fungal pathogens, including powdery mildews. The mechanisms by which Si exerts its prophylactic role remain enigmatic, although a prevailing hypothesis suggests that Si positively influences priming. Attempts to decipher Si properties have been limited to plants able to absorb Si, which excludes the model plant Arabidopsis because it lacks Si influx transporters. In this work, we were able to engineer Arabidopsis plants with an Si transporter from wheat (TaLsi1) and to exploit mutants (pad4 and sid2) deficient in salicylic acid (SA)-dependent defence responses to study their phenotypic response and changes in defence expression against Golovinomyces cichoracearum (Gc) following Si treatment. Our results showed that TaLsi1 plants contained significantly more Si and were significantly more resistant to Gc infection than control plants when treated with Si, the first such demonstration in a plant transformed with a heterologous Si transporter. The resistant plants accumulated higher levels of SA and expressed higher levels of transcripts encoding defence genes, thus suggesting a role for Si in the process. However, TaLsi1 pad4 and TaLsi1 sid2 plants were also more resistant to Gc than were pad4 and sid2 plants following Si treatment. Analysis of the resistant phenotypes revealed a significantly reduced production of SA and expression of defence genes comparable with susceptible controls. These results indicate that Si contributes to Arabidopsis defence priming following pathogen infection, but highlight that Si will confer protection even when priming is altered. We conclude that Si-mediated protection involves mechanisms other than SA-dependent defence responses. PMID:25346281

  17. Deubiquitinating enzymes regulate PARK2-mediated mitophagy

    PubMed Central

    Wang, Yuqing; Serricchio, Mauro; Jauregui, Miluska; Shanbhag, Riya; Stoltz, Tasha; Di Paolo, Caitlin T; Kim, Peter K; McQuibban, G Angus

    2015-01-01

    The selective degradation of mitochondria by the process of autophagy, termed mitophagy, is one of the major mechanisms of mitochondrial quality control. The best-studied mitophagy pathway is the one mediated by PINK1 and PARK2/Parkin. From recent studies it has become clear that ubiquitin-ligation plays a pivotal role and most of the focus has been on the role of ubiquitination of mitochondrial proteins in mitophagy. Even though ubiquitination is a reversible process, very little is known about the role of deubiquitinating enzymes (DUBs) in mitophagy. Here, we report that 2 mitochondrial DUBs, USP30 and USP35, regulate PARK2-mediated mitophagy. We show that USP30 and USP35 can delay PARK2-mediated mitophagy using a quantitative mitophagy assay. Furthermore, we show that USP30 delays mitophagy by delaying PARK2 recruitment to the mitochondria during mitophagy. USP35 does not delay PARK2 recruitment, suggesting that it regulates mitophagy through an alternative mechanism. Interestingly, USP35 only associates with polarized mitochondria, and rapidly translocates to the cytosol during CCCP-induced mitophagy. It is clear that PARK2-mediated mitophagy is regulated at many steps in this important quality control pathway. Taken together, these findings demonstrate an important role of mitochondrial-associated DUBs in mitophagy. Because defects in mitochondria quality control are implicated in many neurodegenerative disorders, our study provides clear rationales for the design and development of drugs for the therapeutic treatment of neurodegenerative diseases such as Parkinson and Alzheimer diseases. PMID:25915564

  18. Powdery Mildew Disease Resistance

    SciTech Connect

    Somerville, Shauna C.

    2010-08-31

    The overall goal of this project was to characterize the PMR5 protein, a member of the DUF231/TBR family, and to determine its role in plant cell wall biogenesis. Since the pmr5 mutants are also resistant to the fungal powdery mildew pathogen, we wished to determine what specific cell wall changes are associated with disease resistance and why. The graduate student working on this project made mutations in the putative active site of PMR5, assuming it is a member of the SGNH/GDSL esterase superfamily (Anantharaman and Aravind, 2010, Biology Direct 5, 1). These mutants were inactive in planta suggesting that PMR5 is a functional enzyme and not a binding protein or chaperone. In addition, she determined that cell wall preparations from the pmr5 mutant exhibited a modest reduction (13%) in total acetyl groups. To pursue characterization further, the graduate student expressed the PMR5 protein in a heterologous E. coli system. She could purify PMR5 using a two step protocol based on tags added to the N and C terminus of the protein. She was able to show the PMR5 protein bound to pectins, including homogalacturonan, but not to other cell wall components (e.g., xyloglucans, arabinans). Based on these observations, a postdoctoral fellow is currently developing an enzyme assay for PMR5 based on the idea that it may be acetylating the homogalacturonic acid pectin fraction. Our initial experiments to localize PMR5 subcellularly suggested that it occurred in the endoplasmic reticulum. However, since the various pectins are believed to be synthesized in the Golgi apparatus, we felt it necessary to repeat our results using a native promoter expression system. Within the past year, we have demonstrated conclusively that PMR5 is localized to the endoplasmic reticulum, a location that sets it apart from most cell wall biogenesis and modification enzymes. The graduate student contributed to the characterization of two suppressor mutants, which were selected as restoring powdery

  19. Down-regulation of Arabidopsis DND1 orthologs in potato and tomato leads to broad-spectrum resistance to late blight and powdery mildew.

    PubMed

    Sun, Kaile; Wolters, Anne-Marie A; Loonen, Annelies E H M; Huibers, Robin P; van der Vlugt, René; Goverse, Aska; Jacobsen, Evert; Visser, Richard G F; Bai, Yuling

    2016-04-01

    Multiple susceptibility genes (S), identified in Arabidopsis, have been shown to be functionally conserved in crop plants. Mutations in these S genes result in resistance to different pathogens, opening a new way to achieve plant disease resistance. The aim of this study was to investigate the role of Defense No Death 1 (DND1) in susceptibility of tomato and potato to late blight (Phytophthora infestans). In Arabidopsis, the dnd1 mutant has broad-spectrum resistance against several fungal, bacterial, and viral pathogens. However this mutation is also associated with a dwarfed phenotype. Using an RNAi approach, we silenced AtDND1 orthologs in potato and tomato. Our results showed that silencing of the DND1 ortholog in both crops resulted in resistance to the pathogenic oomycete P. infestans and to two powdery mildew species, Oidium neolycopersici and Golovinomyces orontii. The resistance to P. infestans in potato was effective to four different isolates although the level of resistance (complete or partial) was dependent on the aggressiveness of the isolate. In tomato, DND1-silenced plants showed a severe dwarf phenotype and autonecrosis, whereas DND1-silenced potato plants were not dwarfed and showed a less pronounced autonecrosis. Our results indicate that S gene function of DND1 is conserved in tomato and potato. We discuss the possibilities of using RNAi silencing or loss-of-function mutations of DND1 orthologs, as well as additional S gene orthologs from Arabidopsis, to breed for resistance to pathogens in crop plants. PMID:26577903

  20. Detection of powdery scab on hairy nightshades

    Technology Transfer Automated Retrieval System (TEKTRAN)

    During the 2002 and 2003 growing seasons, powdery scab like root galls were detected on roots of hairynightshades (Solanum physalifolim, formerly S. sarrachoides) grown in potato fields where populations of the powdery scab pathogen were high. At the end of the 2007 growing season, hairy nightshades...

  1. Resistance to Powdery Mildew in Hydrangeas

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew is an important foliar disease of hydrangeas. For developing efficient integrated disease management strategies involving the utilization of resistant cultivars, it is essential to understand how the powdery mildew fungus infects hydrangea plants and how the host resists the parasitis...

  2. Differential gene expression during conidiation in the grape powdery mildew fungus, Erysiphe necator

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Asexual sporulation (conidiation) is coordinately regulated in the grape powdery mildew fungus Erysiphe necator, but nothing is known about its genetic regulation. We hypothesized that genes required for conidiation in other fungi would be up-regulated at conidiophore initiation and/or full conidia...

  3. Cell mediated immune regulation in autoimmunity.

    PubMed

    Gillissen, G; Pusztai-Markos, Z

    1979-01-01

    Autoimmunity is the term for the immune conditions characterized by a specific humoral or cell mediated response to the body's own tissues. The termination of the natural state of self tolerance may lead to immunopathological manifestations with clinical consequences, i.e. autoimmune diseases. In a very general sense, one may classify autoimmune diseases into two groups with respect to the underlying mechanism: 1. There are autoimmune diseases which develop in the presence of a normal intact regulation mechanism. 2. Another group whose development must be understood on the basis of a cellular dysfunction. In the first case, dequestered or semi-sequestered autoantigens are liberated as a consequence of exogenic influences inducing the sensitization of immunocompetent cells. The immune system then reacts with these autoantigens in the same way as with foreign substances. This kind of autoimmune disease will, however, not be dealt with here. In the second case, autoantigens are normally, i.e. in healthy individuals, accessible to the immunocompetent cells. To understand the reason for the development of an autoimmune reaction one must first clarify the mechanism of self tolerance. Then one must examine the way in which a break of this physiological state takes place. One of the major unanswered questions is the relative importance of antibody-mediated and cell-mediated immune mechanisms in the onset and further development of autoimmune diseases. Recently it has been suggested that a dysfunction at the cellular level might represent the basic cause which induces the termination of selftolerance. Most of the conceptions about the mechanism by which autoimmune diseases are triggered were gained through experiments with animals. It is, however, difficult to use these experimental results to explain human diseases; in humans many questions are still open. Undoubtedly, the mechanisms of induction and maintenance of self tolerance and also the ways in which autoimmune

  4. LIFEGUARD proteins support plant colonization by biotrophic powdery mildew fungi.

    PubMed

    Weis, Corina; Hückelhoven, Ralph; Eichmann, Ruth

    2013-09-01

    Pathogenic microbes manipulate eukaryotic cells during invasion and target plant proteins to achieve host susceptibility. BAX INHIBITOR-1 (BI-1) is an endoplasmic reticulum-resident cell death suppressor in plants and animals and is required for full susceptibility of barley to the barley powdery mildew fungus Blumeria graminis f.sp. hordei. LIFEGUARD (LFG) proteins resemble BI-1 proteins in terms of predicted membrane topology and cell-death-inhibiting function in metazoans, but display clear sequence-specific distinctions. This work shows that barley (Hordeum vulgare L.) and Arabidopsis thaliana genomes harbour five LFG genes, HvLFGa-HvLFGe and AtLFG1-AtLFG5, whose functions are largely uncharacterized. As observed for HvBI-1, single-cell overexpression of HvLFGa supports penetration success of B. graminis f.sp. hordei into barley epidermal cells, while transient-induced gene silencing restricts it. In penetrated barley epidermal cells, a green fluorescent protein-tagged HvLFGa protein accumulates at the site of fungal entry, around fungal haustoria and in endosomal or vacuolar membranes. The data further suggest a role of LFG proteins in plant-powdery mildew interactions in both monocot and dicot plants, because stable overexpression or knockdown of AtLFG1 or AtLFG2 also support or delay development of the powdery mildew fungus Erysiphe cruciferarum on the respective Arabidopsis mutants. Together, this work has identified new modulators of plant-powdery mildew interactions, and the data further support functional similarities between BI-1 and LFG proteins beyond cell death regulation. PMID:23888068

  5. LIFEGUARD proteins support plant colonization by biotrophic powdery mildew fungi

    PubMed Central

    Weis, Corina; Hückelhoven, Ralph; Eichmann, Ruth

    2013-01-01

    Pathogenic microbes manipulate eukaryotic cells during invasion and target plant proteins to achieve host susceptibility. BAX INHIBITOR-1 (BI-1) is an endoplasmic reticulum-resident cell death suppressor in plants and animals and is required for full susceptibility of barley to the barley powdery mildew fungus Blumeria graminis f.sp. hordei. LIFEGUARD (LFG) proteins resemble BI-1 proteins in terms of predicted membrane topology and cell-death-inhibiting function in metazoans, but display clear sequence-specific distinctions. This work shows that barley (Hordeum vulgare L.) and Arabidopsis thaliana genomes harbour five LFG genes, HvLFGa–HvLFGe and AtLFG1–AtLFG5, whose functions are largely uncharacterized. As observed for HvBI-1, single-cell overexpression of HvLFGa supports penetration success of B. graminis f.sp. hordei into barley epidermal cells, while transient-induced gene silencing restricts it. In penetrated barley epidermal cells, a green fluorescent protein-tagged HvLFGa protein accumulates at the site of fungal entry, around fungal haustoria and in endosomal or vacuolar membranes. The data further suggest a role of LFG proteins in plant–powdery mildew interactions in both monocot and dicot plants, because stable overexpression or knockdown of AtLFG1 or AtLFG2 also support or delay development of the powdery mildew fungus Erysiphe cruciferarum on the respective Arabidopsis mutants. Together, this work has identified new modulators of plant–powdery mildew interactions, and the data further support functional similarities between BI-1 and LFG proteins beyond cell death regulation. PMID:23888068

  6. Quantitative and temporal definition of the Mla transcriptional regulon during barley-powdery mildew interactions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Barley Mildew resistance locus a (Mla) is a major determinant of immunity to the powdery mildew pathogen, Blumeria graminis f. sp. hordei. Alleles of Mla encode cytoplasmic- and membrane-localized coiled-coil, nucleotide binding site, leucine-rich repeat proteins that mediate resistance when complem...

  7. A small GTP-binding host protein is required for entry of powdery mildew fungus into epidermal cells of barley.

    PubMed

    Schultheiss, Holger; Dechert, Cornelia; Kogel, Karl-Heinz; Hückelhoven, Ralph

    2002-04-01

    Small GTP-binding proteins such as those from the RAC family are cytosolic signal transduction proteins that often are involved in processing of extracellular stimuli. Plant RAC proteins are implicated in regulation of plant cell architecture, secondary wall formation, meristem signaling, and defense against pathogens. We isolated a RacB homolog from barley (Hordeum vulgare) to study its role in resistance to the barley powdery mildew fungus (Blumeria graminis f.sp. hordei). RacB was constitutively expressed in the barley epidermis and its expression level was not strongly influenced by inoculation with B. graminis. However, after biolistic bombardment of barley leaf segments with RacB-double-stranded RNA, sequence-specific RNA interference with RacB function inhibited fungal haustorium establishment in a cell-autonomous and genotype-specific manner. Mutants compromised in function of the Mlo wild-type gene and the Ror1 gene (genotype mlo5 ror1) that are moderately susceptible to B. graminis showed no alteration in powdery mildew resistance upon RacB-specific RNA interference. Thus, the phenotype, induced by RacB-specific RNA interference, was apparently dependent on the same processes as mlo5-mediated broad resistance, which is suppressed by ror1. We conclude that an RAC small GTP-binding protein is required for successful fungal haustorium establishment and that this function may be linked to MLO-associated functions. PMID:11950993

  8. Transcript-based Cloning of RRP46, a Regulator of rRNA Processing and R-Gene-Independent Cell Death in Barley–Powdery Mildew Interactions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Programmed cell death (PCD) plays a pivotal role in plant development and defense. To investigate the degree of interaction between PCD and R-gene mediated defense, we used the 22K Barley1 GeneChip to compare and contrast time-course expression profiles of Blumeria graminis f. sp. hordei (Bgh) chal...

  9. The knottin-like Blufensin family regulates genes involved in nuclear import and the secretory pathway in barley-powdery mildew interactions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plants have evolved complex regulatory mechanisms to control a multi-layered defense response to microbial attack. Both temporal and spatial gene expression are tightly regulated in response to pathogen ingress, modulating both positive and negative control of defense. BLUFENSINs, small knottin-like...

  10. Potential alternative hosts for a powdery mildew on pea

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew of pea (Pisum sativum) is an important disease in the field and in the greenhouse. The most widely documented powdery mildew on pea is Erysiphe pisi, but E. trifolii and E. baeumleri have also been reported. From greenhouse-grown peas, we obtained powdery mildew samples with rDNA ITS ...

  11. Receptor Complex Mediated Regulation of Symplastic Traffic.

    PubMed

    Stahl, Yvonne; Faulkner, Christine

    2016-05-01

    Plant receptor kinases (RKs) and receptor proteins (RPs) are involved in a plethora of cellular processes, including developmental decisions and immune responses. There is increasing evidence that plasmodesmata (PD)-localized RKs and RPs act as nexuses that perceive extracellular signals and convey them into intra- and intercellular responses by regulating the exchange of molecules through PD. How RK/RP complexes regulate the specific and nonspecific traffic of molecules through PD, and how these receptors are specifically targeted to PD, have been elusive but underpin comprehensive understanding of the function and regulation of the symplast. In this review we gather the current knowledge of RK/RP complex function at PD and how they might regulate intercellular traffic. PMID:26655263

  12. Inferring RBP-Mediated Regulation in Lung Squamous Cell Carcinoma

    PubMed Central

    Lafzi, Atefeh; Kazan, Hilal

    2016-01-01

    RNA-binding proteins (RBPs) play key roles in post-transcriptional regulation of mRNAs. Dysregulations in RBP-mediated mechanisms have been found to be associated with many steps of cancer initiation and progression. Despite this, previous studies of gene expression in cancer have ignored the effect of RBPs. To this end, we developed a lasso regression model that predicts gene expression in cancer by incorporating RBP-mediated regulation as well as the effects of other well-studied factors such as copy-number variation, DNA methylation, TFs and miRNAs. As a case study, we applied our model to Lung squamous cell carcinoma (LUSC) data as we found that there are several RBPs differentially expressed in LUSC. Including RBP-mediated regulatory effects in addition to the other features significantly increased the Spearman rank correlation between predicted and measured expression of held-out genes. Using a feature selection procedure that accounts for the adaptive search employed by lasso regularization, we identified the candidate regulators in LUSC. Remarkably, several of these candidate regulators are RBPs. Furthermore, majority of the candidate regulators have been previously found to be associated with lung cancer. To investigate the mechanisms that are controlled by these regulators, we predicted their target gene sets based on our model. We validated the target gene sets by comparing against experimentally verified targets. Our results suggest that the future studies of gene expression in cancer must consider the effect of RBP-mediated regulation. PMID:27186987

  13. Soluble Mediators Regulating Immunity in Early Life

    PubMed Central

    Pettengill, Matthew Aaron; van Haren, Simon Daniël; Levy, Ofer

    2014-01-01

    Soluble factors in blood plasma have a substantial impact on both the innate and adaptive immune responses. The complement system, antibodies, and anti-microbial proteins and peptides can directly interact with potential pathogens, protecting against systemic infection. Levels of these innate effector proteins are generally lower in neonatal circulation at term delivery than in adults, and lower still at preterm delivery. The extracellular environment also has a critical influence on immune cell maturation, activation, and effector functions, and many of the factors in plasma, including hormones, vitamins, and purines, have been shown to influence these processes for leukocytes of both the innate and adaptive immune systems. The ontogeny of plasma factors can be viewed in the context of a lower effectiveness of immune responses to infection and immunization in early life, which may be influenced by the striking neonatal deficiency of complement system proteins or enhanced neonatal production of the anti-inflammatory cytokine IL-10, among other ontogenic differences. Accordingly, we survey here a number of soluble mediators in plasma for which age-dependent differences in abundance may influence the ontogeny of immune function, particularly direct innate interaction and skewing of adaptive lymphocyte activity in response to infectious microorganisms and adjuvanted vaccines. PMID:25309541

  14. Regulated lysosomal exocytosis mediates cancer progression

    PubMed Central

    Machado, Eda; White-Gilbertson, Shai; van de Vlekkert, Diantha; Janke, Laura; Moshiach, Simon; Campos, Yvan; Finkelstein, David; Gomero, Elida; Mosca, Rosario; Qiu, Xiaohui; Morton, Christopher L.; Annunziata, Ida; d’Azzo, Alessandra

    2015-01-01

    Understanding how tumor cells transition to an invasive and drug-resistant phenotype is central to cancer biology, but the mechanisms underlying this transition remain unclear. We show that sarcomas gain these malignant traits by inducing lysosomal exocytosis, a ubiquitous physiological process. During lysosomal exocytosis, the movement of exocytic lysosomes along the cytoskeleton and their docking at the plasma membrane involve LAMP1, a sialylated membrane glycoprotein and target of the sialidase NEU1. Cleavage of LAMP1 sialic acids by NEU1 limits the extent of lysosomal exocytosis. We found that by down-regulation of NEU1 and accumulation of oversialylated LAMP1, tumor cells exacerbate lysosomal exocytosis of soluble hydrolases and exosomes. This facilitates matrix invasion and propagation of invasive signals, and purging of lysosomotropic chemotherapeutics. In Arf−⁄− mice, Neu1 haploinsufficiency fostered the development of invasive, pleomorphic sarcomas, expressing epithelial and mesenchymal markers, and lysosomal exocytosis effectors, LAMP1 and Myosin-11. These features are analogous to those of metastatic, pleomorphic human sarcomas, where low NEU1 levels correlate with high expression of lysosomal exocytosis markers. In a therapeutic proof of principle, we demonstrate that inhibiting lysosomal exocytosis reversed invasiveness and chemoresistance in aggressive sarcoma cells. Thus, we reveal that this unconventional, lysosome-regulated pathway plays a primary role in tumor progression and chemoresistance. PMID:26824057

  15. Tropomyosin-Mediated Regulation of Cytoplasmic Myosins.

    PubMed

    Manstein, Dietmar J; Mulvihill, Daniel P

    2016-08-01

    The ability of the actin-based cytoskeleton to rapidly reorganize is critical for maintaining cell organization and viability. The plethora of activities in which actin polymers participate require different biophysical properties, which can vary significantly between the different events that often occur simultaneously at separate cellular locations. In order to modify the biophysical properties of an actin polymer for a particular function, the cell contains diverse actin-binding proteins that modulate the growth, regulation and molecular interactions of actin-based structures according to functional requirements. In metazoan and yeast cells, tropomyosin is a key regulator of actin-based structures. Cells have the capacity to produce multiple tropomyosin isoforms, each capable of specifically associating as copolymers with actin at distinct cellular locations to fine-tune the functional properties of discrete actin structures. Here, we present a unifying theory in which tropomyosin isoforms critically define the surface landscape of copolymers with cytoplasmic β- or γ-actin. Decoration of filamentous actin with different tropomyosin isoforms determines the identity and modulates the activity of the interacting myosin motor proteins. Conversely, changes in the nucleotide state of actin and posttranslational modifications affect the composition, morphology, subcellular localization and allosteric coupling of the associated actin-based superstructures. PMID:27060364

  16. Inoculum detection for managing grape powdery mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Grape powdery mildew (caused by Erysiphe necator) often develops explosively and can result in significant yield and quality losses, despite your best efforts. The threat of quality losses often leads to a low risk prophylactic regimen rather than a higher risk IPM approach. Gary Grove and I have co...

  17. An Update on Resistance to Powdery Scab

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery scab infections in the Columbia Basin occur early in the growing season when soil temperatures are relatively cool and irrigation water supplies the soil moisture needed for infection. Root galls usually appear 3 weeks after infection, which roughly correlates to 1.5 to 2 months past plant e...

  18. Resistance to Powdery Mildew in Hydrangea

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew, caused by Erysiphe polygoni, is an important foliar disease of hydrangeas. Infection process and development of E. polygoni on hydrangeas were determined using a leaf disk assay. On susceptible hydrangea cultivars, conidia of E. polygoni germinated and formed primary appressoria with...

  19. Resistance to Powdery Scab in Potato

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery scab is a serious disease of potato that is among the most damaging emerging potato pathogens throughout the world. Twenty years ago it was rarity in the Columbia Basin but today it is widespread, damaging and a threat to the profitability of the industry. Flagellated zoospores which swim ...

  20. Controlling Powdery Scab: The Breeding Approach

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery scab is a serious disease of potato caused by the fungus Spongospora subterranea. S. subterranea is a member of the Plasmodiophorales, also known as the parasitic slime molds. The primary diagnostic sign of the disease are cystosori, spongelike aggregates of cysts or resting spores. The ...

  1. Mechanisms of Resistance to Grapevine Powdery Mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Although all high quality Vitis vinifera cultivars are highly susceptible to powdery mildew, current efforts show promise for the identification or modulation of resistance within the species and for continued introgression of resistance from resistant accessions of wild Vitis spp. Examination of t...

  2. Identification of powdery mildew fungi anno 2006

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This is an update of the previous laboratory exercise on powdery mildews at the Plant Health Instructor website. In 2002, there was a drastic change in the taxonomy of the Erysiphales, and most teaching documents had become outdated and even unusable. This exercise was essentially a new creation, ...

  3. Small RNA mediated regulation of seed germination

    PubMed Central

    Das, Shabari Sarkar; Karmakar, Prakash; Nandi, Asis Kumar; Sanan-Mishra, Neeti

    2015-01-01

    Mature seeds of most of the higher plants harbor dormant embryos and go through the complex process of germination under favorable environmental conditions. The germination process involves dynamic physiological, cellular and metabolic events that are controlled by the interplay of several gene products and different phytohormones. The small non-coding RNAs comprise key regulatory modules in the process of seed dormancy and germination. Recent studies have implicated the small RNAs in plant growth in correlation with various plant physiological processes including hormone signaling and stress response. In this review we provide a brief overview of the regulation of seed germination or dormancy while emphasizing on the current understanding of the role of small RNAs in this regard. We have also highlighted specific examples of stress responsive small RNAs in seed germination and discussed their future potential. PMID:26528301

  4. Regulation of cell proliferation and apoptosis by bioactive lipid mediators.

    PubMed

    Clària, Joan

    2006-11-01

    Bioactive lipid mediators are increasingly being recognized as important endogenous regulators of cell activation, signaling, apoptosis and proliferation. Most of these lipid mediators are originated from cleavage of constituents of cellular membranes under the activity of phospholipases and sphingomyelinases. One of the major cascades of bioactive lipid mediator production involves the release of arachidonic acid from membrane phospholipids followed by the formation of eicosanoids (i.e. prostaglandins, leukotrienes and lipoxins). These biologically active metabolites of arachidonic acid are emerging as key regulators of cell proliferation and neo-angiogenesis and agents that specifically target these lipid mediators are being investigated as potential anticancer drugs. On the other hand, the lysophospholipid family, which includes members of the sphingomyelin-ceramide-sphingosine-1-phosphate and lysophosphatidic acid subfamilies, has evolved as an important group of lipid signaling molecules implicated in cellular differentiation, cell growth and apoptosis. This article reviews the most recent patents in this field of research, covering the following strategies based on the modulation of bioactive lipid mediators: (1) prostaglandin H synthase-2 inhibitors, (2) lipoxin analogs and aspirin-triggered lipid mediators, and (3) lysophosphatidic acid and other lysophospholipids. PMID:18221047

  5. Hydrogen Sulfide Regulates Homocysteine-Mediated Glomerulosclerosis

    PubMed Central

    Sen, Utpal; Munjal, Charu; Qipshidze, Natia; Abe, Oluwasegun; Gargoum, Riyad; Tyagi, Suresh C.

    2010-01-01

    Background/Aims In this study we tested the hypothesis that H2S regulates collagen deposition, matrix metalloproteinases (MMP) and inflammatory molecules during hyperhomocysteinemia (HHcy) resulting in attenuation of glomerulosclerosis and improved renal function. Materials and Methods A genetic model of HHcy, cystathionine β-synthase heterozygous (CBS+/−) and wild-type (WT) 2-kidney (2K) mice were used in this study and supplemented with or without NaHS (30 μmol/l, H2S donor) in drinking water for 8 weeks. To expedite the renal damage associated with HHcy, uninephrectomized (1K) mice of similar groups were also used. Results Results demonstrated that NAD(P)H oxidase (p47phox subunit) and blood pressure were upregulated in WT 1K, CBS+/− 2K and CBS+/− 1K mice with downregulation of H2S production and reduced glomerular filtration rate. These changes were normalized with H2S supplementation. Both pro- and active MMP-2 and -9 and collagen protein expressions and glomerular depositions were also upregulated in WT 1K, CBS+/− 2K and CBS+/− 1K mice. Increased expressions of inflammatory molecules, intercellular cell adhesion molecule-1 and vascular cell adhesion molecule-1, as well as increased macrophage infiltration, were detected in WT 1K, CBS+/− 2K and CBS+/− 1K mice. These changes were ameliorated with H2S supplementation. Conclusion Together, these results suggest that increased oxidative stress and decreased H2S in HHcy causes matrix remodeling and inflammation resulting in glomerulosclerosis and reduced renal function. PMID:20395677

  6. Mediator Complex Dependent Regulation of Cardiac Development and Disease

    PubMed Central

    Grueter, Chad E.

    2013-01-01

    Cardiovascular disease (CVD) is a leading cause of morbidity and mortality. The risk factors for CVD include environmental and genetic components. Human mutations in genes involved in most aspects of cardiovascular function have been identified, many of which are involved in transcriptional regulation. The Mediator complex serves as a pivotal transcriptional regulator that functions to integrate diverse cellular signals by multiple mechanisms including recruiting RNA polymerase II, chromatin modifying proteins and non-coding RNAs to promoters in a context dependent manner. This review discusses components of the Mediator complex and the contribution of the Mediator complex to normal and pathological cardiac development and function. Enhanced understanding of the role of this core transcriptional regulatory complex in the heart will help us gain further insights into CVD. PMID:23727265

  7. Theoretical studies on sRNA-mediated regulation in bacteria

    NASA Astrophysics Data System (ADS)

    Chang, Xiao-Xue; Xu, Liu-Fang; Shi, Hua-Lin

    2015-12-01

    Small RNA(sRNA)-mediated post-transcriptional regulation differs from protein-mediated regulation. Through base-pairing, sRNA can regulate the target mRNA in a catalytic or stoichiometric manner. Some theoretical models were built for comparison of the protein-mediated and sRNA-mediated modes in the steady-state behaviors and noise properties. Many experiments demonstrated that a single sRNA can regulate several mRNAs, which causes crosstalk between the targets. Here, we focus on some models in which two target mRNAs are silenced by the same sRNA to discuss their crosstalk features. Additionally, the sequence-function relationship of sRNA and its role in the kinetic process of base-pairing have been highlighted in model building. Project supported by the National Basic Research Program of China (Grant No. 2013CB834100), the National Natural Science Foundation of China (Grant Nos. 11121403 and 11274320), the Open Project Program of State Key Laboratory of Theoretical Physics, Institute of Theoretical Physics, Chinese Academy of Sciences, China (Grant No. Y4KF171CJ1), the National Natural Science Foundation for Young Scholar of China (Grant No. 11304115), and the China Postdoctoral Science Foundation (Grant No. 2013M541282).

  8. Widespread context dependency of microRNA-mediated regulation

    PubMed Central

    Erhard, Florian; Haas, Jürgen; Lieber, Diana; Malterer, Georg; Jaskiewicz, Lukasz; Zavolan, Mihaela; Dölken, Lars; Zimmer, Ralf

    2014-01-01

    Gene expression is regulated in a context-dependent, cell-type-specific manner. Condition-specific transcription is dependent on the presence of transcription factors (TFs) that can activate or inhibit its target genes (global context). Additional factors, such as chromatin structure, histone, or DNA modifications, also influence the activity of individual target genes (individual context). The role of the global and individual context for post-transcriptional regulation has not systematically been investigated on a large scale and is poorly understood. Here we show that global and individual context dependency is a pervasive feature of microRNA-mediated regulation. Our comprehensive and highly consistent data set from several high-throughput technologies (PAR-CLIP, RIP-chip, 4sU-tagging, and SILAC) provides strong evidence that context-dependent microRNA target sites (CDTS) are as frequent and functionally relevant as constitutive target sites (CTS). Furthermore, we found the global context to be insufficient to explain the CDTS, and that flanking sequence motifs provide individual context that is an equally important factor. Our results demonstrate that, similar to TF-mediated regulation, global and individual context dependency are prevalent in microRNA-mediated gene regulation, implying a much more complex post-transcriptional regulatory network than is currently known. The necessary tools to unravel post-transcriptional regulations and mechanisms need to be much more involved, and much more data will be needed for particular cell types and cellular conditions in order to understand microRNA-mediated regulation and the context-dependent post-transcriptional regulatory network. PMID:24668909

  9. Widespread context dependency of microRNA-mediated regulation.

    PubMed

    Erhard, Florian; Haas, Jürgen; Lieber, Diana; Malterer, Georg; Jaskiewicz, Lukasz; Zavolan, Mihaela; Dölken, Lars; Zimmer, Ralf

    2014-06-01

    Gene expression is regulated in a context-dependent, cell-type-specific manner. Condition-specific transcription is dependent on the presence of transcription factors (TFs) that can activate or inhibit its target genes (global context). Additional factors, such as chromatin structure, histone, or DNA modifications, also influence the activity of individual target genes (individual context). The role of the global and individual context for post-transcriptional regulation has not systematically been investigated on a large scale and is poorly understood. Here we show that global and individual context dependency is a pervasive feature of microRNA-mediated regulation. Our comprehensive and highly consistent data set from several high-throughput technologies (PAR-CLIP, RIP-chip, 4sU-tagging, and SILAC) provides strong evidence that context-dependent microRNA target sites (CDTS) are as frequent and functionally relevant as constitutive target sites (CTS). Furthermore, we found the global context to be insufficient to explain the CDTS, and that flanking sequence motifs provide individual context that is an equally important factor. Our results demonstrate that, similar to TF-mediated regulation, global and individual context dependency are prevalent in microRNA-mediated gene regulation, implying a much more complex post-transcriptional regulatory network than is currently known. The necessary tools to unravel post-transcriptional regulations and mechanisms need to be much more involved, and much more data will be needed for particular cell types and cellular conditions in order to understand microRNA-mediated regulation and the context-dependent post-transcriptional regulatory network. PMID:24668909

  10. SUMOylation-mediated regulation of cell cycle progression and cancer

    PubMed Central

    Eifler, Karolin; Vertegaal, Alfred C.O.

    2016-01-01

    SUMOylation plays critical roles during cell cycle progression. Many important cell cycle regulators, including many oncogenes and tumor suppressors, are functionally regulated via SUMOylation. The dynamic SUMOylation pattern observed throughout the cell cycle is ensured via distinct spatial and temporal regulation of the SUMO machinery. Additionally, SUMOylation cooperates with other post-translational modifications to mediate cell cycle progression. Deregulation of these SUMOylation and deSUMOylation enzymes causes severe defects in cell proliferation and genome stability. Different types of cancers were recently shown to be dependent on a functioning SUMOylation system, a finding that could potentially be exploited in anti-cancer therapies. PMID:26601932

  11. Lasiodiplodia theobromae is a Mycoparasite of a Powdery Mildew Pathogen.

    PubMed

    Sreerama Kumar, P; Singh, Leena

    2009-12-01

    Powdery mildews on over 40 plants in Bangalore were screened during July-December of 2003~2008. Isolates from mycoparasitised Oidium caesalpiniacearum of Bauhinia purpurea comprised Lasiodiplodia theobromae, in addition to Ampelomyces quisqualis. Koch's postulates were satisfied to establish the mycoparasitism of L. theobromae. This is the first report that L. theobromae acts as a mycoparasite of a powdery mildew. PMID:23983554

  12. Lasiodiplodia theobromae is a Mycoparasite of a Powdery Mildew Pathogen

    PubMed Central

    Singh, Leena

    2009-01-01

    Powdery mildews on over 40 plants in Bangalore were screened during July-December of 2003~2008. Isolates from mycoparasitised Oidium caesalpiniacearum of Bauhinia purpurea comprised Lasiodiplodia theobromae, in addition to Ampelomyces quisqualis. Koch's postulates were satisfied to establish the mycoparasitism of L. theobromae. This is the first report that L. theobromae acts as a mycoparasite of a powdery mildew. PMID:23983554

  13. NFAT regulates calcium-sensing receptor-mediated TNF production

    SciTech Connect

    abdullah, huda ismail; Pedraza, Paulina L.; Hao, Shoujin; Rodland, Karin D.; McGiff, John C.; Ferreri, Nicholas R.

    2006-05-01

    Because nuclear factor of activated T cells (NFAT) has been implicated in TNF production as well as osmoregulation and salt and water homeostasis, we addressed whether calcium-sensing receptor (CaR)-mediated TNF production in medullary thick ascending limb (mTAL) cells was NFAT dependent. TNF production in response to addition of extracellular Ca2+ (1.2 mM) was abolished in mTAL cells transiently transfected with a dominant-negative CaR construct (R796W) or pretreated with the phosphatidylinositol phospholipase C (PI-PLC) inhibitor U-73122. Cyclosporine A (CsA), an inhibitor of the serine/threonine phosphatase calcineurin, and a peptide ligand, VIVIT, that selectively inhibits calcineurin-NFAT signaling, also prevented CaR-mediated TNF production. Increases in calcineurin activity in cells challenged with Ca2+ were inhibited after pretreatment with U-73122 and CsA, suggesting that CaR activation increases calcineurin activity in a PI-PLC-dependent manner. Moreover, U-73122, CsA, and VIVIT inhibited CaR-dependent activity of an NFAT construct that drives expression of firefly luciferase in transiently transfected mTAL cells. Collectively, these data verify the role of calcineurin and NFAT in CaR-mediated TNF production by mTAL cells. Activation of the CaR also increased the binding of NFAT to a consensus oligonucleotide, an effect that was blocked by U-73122 and CsA, suggesting that a calcineurin- and NFAT-dependent pathway increases TNF production in mTAL cells. This mechanism likely regulates TNF gene transcription as U-73122, CsA, and VIVIT blocked CaR-dependent activity of a TNF promoter construct. Elucidating CaR-mediated signaling pathways that regulate TNF production in the mTAL will be crucial to understanding mechanisms that regulate extracellular fluid volume and salt balance.

  14. Biocontrol agent Bacillus amyloliquefaciens LJ02 induces systemic resistance against cucurbits powdery mildew

    PubMed Central

    Li, Yunlong; Gu, Yilin; Li, Juan; Xu, Mingzhu; Wei, Qing; Wang, Yuanhong

    2015-01-01

    Powdery mildew is a fungal disease found in a wide range of plants and can significantly reduce crop yields. Bacterial strain LJ02 is a biocontrol agent (BCA) isolated from a greenhouse in Tianjin, China. In combination of morphological, physiological, biochemical and phylogenetic analyses, strain LJ02 was classified as a new member of Bacillus amyloliquefaciens. Greenhouse trials showed that LJ02 fermentation broth (LJ02FB) can effectively diminish the occurrence of cucurbits powdery mildew. When treated with LJ02FB, cucumber seedlings produced significantly elevated production of superoxide dismutase, peroxidase, polyphenol oxidase and phenylalanine ammonia lyase as compared to that of the control. We further confirmed that the production of free salicylic acid (SA) and expression of one pathogenesis-related (PR) gene PR-1 in cucumber leaves were markedly elevated after treating with LJ02FB, suggesting that SA-mediated defense response was stimulated. Moreover, LJ02FB-treated cucumber leaves could secrete resistance-related substances into rhizosphere that inhibit the germination of fungi spores and the growth of pathogens. Finally, we separated bacterium and its fermented substances to test their respective effects and found that both components have SA-inducing activity and bacterium plays major roles. Altogether, we identified a BCA against powdery mildew and its mode of action by inducing systemic resistance such as SA signaling pathway. PMID:26379654

  15. Dscam-Mediated Cell Recognition Regulates Neural Circuit Formation

    PubMed Central

    Hattori, Daisuke; Millard, S. Sean; Wojtowicz, Woj M.; Zipursky, S. Lawrence

    2009-01-01

    The Dscam family of immunoglobulin cell surface proteins mediates recognition events between neurons that play an essential role in the establishment of neural circuits. The Drosophila Dscam1 locus encodes tens of thousands of cell surface proteins via alternative splicing. These isoforms exhibit exquisite isoform-specific binding in vitro that mediates homophilic repulsion in vivo. These properties provide the molecular basis for self-avoidance, an essential developmental mechanism that allows axonal and dendritic processes to uniformly cover their synaptic fields. In a mechanistically similar fashion, homophilic repulsion mediated by Drosophila Dscam2 prevents processes from the same class of cells from occupying overlapping synaptic fields through a process called tiling. Genetic studies in the mouse visual system support the view that vertebrate DSCAM also promotes both self-avoidance and tiling. By contrast, DSCAM and DSCAM-L promote layer-specific targeting in the chick visual system, presumably through promoting homophilic adhesion. The fly and mouse studies underscore the importance of homophilic repulsion in regulating neural circuit assembly, whereas the chick studies suggest that DSCA Mproteins may mediate a variety of different recognition events during wiring in a context-dependent fashion. PMID:18837673

  16. Problems of Subject Mediator Development for Gene Expression Regulation Domain

    NASA Astrophysics Data System (ADS)

    Kalinichenko, L. A.; Briukhov, D. O.; Zakharov, V. N.; Podkolodnaja, O. A.; Podkolodny, N. L.

    For efficient organization of research in the domain of bioinformatics it is required to organize properly the relevant information in specific research areas. One of the important outcomes of such organization would be provision of access to and querying of a large number of distributed information sources including various data on the primary and spatial structure of DNA and RNA macromolecules, proteins and their complexes as well as data on peculiarities of their interactions with each other. To provide for semantic integration of nonsystematic population of autonomous information sources kept by different information providers into a well-structured information collection it is required to create the global unified representation of the existing information sources and services. To reach that it is proposed to form a special middleware consisting of the subject mediators. For each subject mediator, the application domain model is to be defined by the experts in the field. This model may include specifications of data structures, terminologies (thesauri), concepts (ontologies), methods applicable to data, processes (workflows), characteristic for the domain. The mediators provide a uniform query interface to the multiple data and procedure service sources, thereby freeing the users from having to locate the relevant sources, query each one in isolation, and combine manually the information from them. In the paper we discuss an approach for development of the mediator for integration of heterogeneous molecular-genetic data in the gene expression regulation domain.

  17. Family Emotion Expressiveness Mediates the Relations Between Maternal Emotion Regulation and Child Emotion Regulation.

    PubMed

    Are, Funlola; Shaffer, Anne

    2016-10-01

    While there is a growing body of literature examining the influence of emotion socialization on children's emotional and social development, there is less research on what predicts emotion socialization behaviors among parents. The current study explores maternal emotion regulation difficulties as a predictor of emotion socialization practices, specifically, family emotion expressiveness. Further, the current study examines the role of family emotion expressiveness as a possible mediator of the relations between maternal and child emotion regulation in a community sample of 110 mother-child dyads with preschool-aged children. Analyses revealed that positive family expressiveness mediated the relations between maternal emotion dysregulation and child emotion regulation and thus presents important clinical implications for existing emotion socialization interventions. PMID:26573929

  18. Mechanism and regulation of the nonsense-mediated decay pathway

    PubMed Central

    Hug, Nele; Longman, Dasa; Cáceres, Javier F.

    2016-01-01

    The Nonsense-mediated mRNA decay (NMD) pathway selectively degrades mRNAs harboring premature termination codons (PTCs) but also regulates the abundance of a large number of cellular RNAs. The central role of NMD in the control of gene expression requires the existence of buffering mechanisms that tightly regulate the magnitude of this pathway. Here, we will focus on the mechanism of NMD with an emphasis on the role of RNA helicases in the transition from NMD complexes that recognize a PTC to those that promote mRNA decay. We will also review recent strategies aimed at uncovering novel trans-acting factors and their functional role in the NMD pathway. Finally, we will describe recent progress in the study of the physiological role of the NMD response. PMID:26773057

  19. Hypoxia-mediated regulation of gene expression in mammalian cells

    PubMed Central

    Shih, Shu-Ching; Claffey, Kevin P.

    1998-01-01

    The molecular mechanism underlying oxygen sensing in mammalian cells has been extensively investigated in the areas of glucose transport, glycolysis, erythropoiesis, angiogenesis and catecholamine metabolism. Expression of functionally operative representative proteins in these specific areas, such as the glucose transporter 1, glycolytic enzymes, erythropoietin, vascular endothelial growth factor and tyrosine hydroxylase are all induced by hypoxia. Recent studies demonstrated that both transcriptional activation and post-transcriptional mechanisms are important to the hypoxia-mediated regulation of gene expression. In this article, the cis-acting elements and trans-acting factors involved in the transcriptional activation of gene expression will be reviewed. In addition, the mechanisms of post-transcriptional mRNA stabilization will also be addressed. We will discuss whether these two processes of regulation of hypoxia-responsive genes are mechanistically linked and co-operative in nature. PMID:10319016

  20. Expression of a Grapevine NAC Transcription Factor Gene Is Induced in Response to Powdery Mildew Colonization in Salicylic Acid-Independent Manner

    PubMed Central

    Toth, Zsofia; Winterhagen, Patrick; Kalapos, Balazs; Su, Yingcai; Kovacs, Laszlo; Kiss, Erzsebet

    2016-01-01

    Tissue colonization by grape powdery mildew (PM) pathogen Erysiphe necator (Schw.) Burr triggers a major remodeling of the transcriptome in the susceptible grapevine Vitis vinifera L. While changes in the expression of many genes bear the signature of salicylic acid (SA) mediated regulation, the breadth of PM-induced changes suggests the involvement of additional regulatory networks. To explore PM-associated gene regulation mediated by other SA-independent systems, we designed a microarray experiment to distinguish between transcriptome changes induced by E. necator colonization and those triggered by elevated SA levels. We found that the majority of genes responded to both SA and PM, but certain genes were responsive to PM infection alone. Among them, we identified genes of stilbene synthases, PR-10 proteins, and several transcription factors. The microarray results demonstrated that the regulation of these genes is either independent of SA, or dependent, but SA alone is insufficient to bring about their regulation. We inserted the promoter-reporter fusion of a PM-responsive transcription factor gene into a wild-type and two SA-signaling deficient Arabidopsis lines and challenged the resulting transgenic plants with an Arabidopsis-adapted PM pathogen. Our results provide experimental evidence that this grape gene promoter is activated by the pathogen in a SA-independent manner. PMID:27488171

  1. Expression of a Grapevine NAC Transcription Factor Gene Is Induced in Response to Powdery Mildew Colonization in Salicylic Acid-Independent Manner.

    PubMed

    Toth, Zsofia; Winterhagen, Patrick; Kalapos, Balazs; Su, Yingcai; Kovacs, Laszlo; Kiss, Erzsebet

    2016-01-01

    Tissue colonization by grape powdery mildew (PM) pathogen Erysiphe necator (Schw.) Burr triggers a major remodeling of the transcriptome in the susceptible grapevine Vitis vinifera L. While changes in the expression of many genes bear the signature of salicylic acid (SA) mediated regulation, the breadth of PM-induced changes suggests the involvement of additional regulatory networks. To explore PM-associated gene regulation mediated by other SA-independent systems, we designed a microarray experiment to distinguish between transcriptome changes induced by E. necator colonization and those triggered by elevated SA levels. We found that the majority of genes responded to both SA and PM, but certain genes were responsive to PM infection alone. Among them, we identified genes of stilbene synthases, PR-10 proteins, and several transcription factors. The microarray results demonstrated that the regulation of these genes is either independent of SA, or dependent, but SA alone is insufficient to bring about their regulation. We inserted the promoter-reporter fusion of a PM-responsive transcription factor gene into a wild-type and two SA-signaling deficient Arabidopsis lines and challenged the resulting transgenic plants with an Arabidopsis-adapted PM pathogen. Our results provide experimental evidence that this grape gene promoter is activated by the pathogen in a SA-independent manner. PMID:27488171

  2. Carbonic anhydrase enzymes regulate mast cell-mediated inflammation.

    PubMed

    Henry, Everett K; Sy, Chandler B; Inclan-Rico, Juan M; Espinosa, Vanessa; Ghanny, Saleena S; Dwyer, Daniel F; Soteropoulos, Patricia; Rivera, Amariliz; Siracusa, Mark C

    2016-08-22

    Type 2 cytokine responses are necessary for the development of protective immunity to helminth parasites but also cause the inflammation associated with allergies and asthma. Recent studies have found that peripheral hematopoietic progenitor cells contribute to type 2 cytokine-mediated inflammation through their enhanced ability to develop into mast cells. In this study, we show that carbonic anhydrase (Car) enzymes are up-regulated in type 2-associated progenitor cells and demonstrate that Car enzyme inhibition is sufficient to prevent mouse mast cell responses and inflammation after Trichinella spiralis infection or the induction of food allergy-like disease. Further, we used CRISPR/Cas9 technology and illustrate that genetically editing Car1 is sufficient to selectively reduce mast cell development. Finally, we demonstrate that Car enzymes can be targeted to prevent human mast cell development. Collectively, these experiments identify a previously unrecognized role for Car enzymes in regulating mast cell lineage commitment and suggest that Car enzyme inhibitors may possess therapeutic potential that can be used to treat mast cell-mediated inflammation. PMID:27526715

  3. Innate Immune Regulation by STAT-mediated Transcriptional Mechanisms

    PubMed Central

    Li, Haiyan S.; Watowich, Stephanie S.

    2014-01-01

    Summary The term innate immunity typically refers to a quick but nonspecific host defense response against invading pathogens. The innate immune system comprises particular immune cell populations, epithelial barriers, and numerous secretory mediators including cytokines, chemokines, and defense peptides. Innate immune cells are also now recognized to play important contributing roles in cancer and pathological inflammatory conditions. Innate immunity relies on rapid signal transduction elicited upon pathogen recognition via pattern recognition receptors (PRRs) and cell:cell communication conducted by soluble mediators, including cytokines. A majority of cytokines involved in innate immune signaling use a molecular cascade encompassing receptor-associated Jak protein tyrosine kinases and STAT (signal transducer and activator of transcription) transcriptional regulators. Here, we focus on roles for STAT proteins in three major innate immune subsets: neutrophils, macrophages, and dendritic cells (DCs). While knowledge in this area is only now emerging, understanding the molecular regulation of these cell types is necessary for developing new approaches to treat human disorders such as inflammatory conditions, autoimmunity, and cancer. PMID:25123278

  4. Identification of Powdery Mildew Responsive Genes in Hevea brasiliensis through mRNA Differential Display

    PubMed Central

    Li, Xiang; Bi, Zhenghong; Di, Rong; Liang, Peng; He, Qiguang; Liu, Wenbo; Miao, Weiguo; Zheng, Fucong

    2016-01-01

    Powdery mildew is an important disease of rubber trees caused by Oidium heveae B. A. Steinmann. As far as we know, none of the resistance genes related to powdery mildew have been isolated from the rubber tree. There is little information available at the molecular level regarding how a rubber tree develops defense mechanisms against this pathogen. We have studied rubber tree mRNA transcripts from the resistant RRIC52 cultivar by differential display analysis. Leaves inoculated with the spores of O. heveae were collected from 0 to 120 hpi in order to identify pathogen-regulated genes at different infection stages. We identified 78 rubber tree genes that were differentially expressed during the plant–pathogen interaction. BLAST analysis for these 78 ESTs classified them into seven functional groups: cell wall and membrane pathways, transcription factor and regulatory proteins, transporters, signal transduction, phytoalexin biosynthesis, other metabolism functions, and unknown functions. The gene expression for eight of these genes was validated by qRT-PCR in both RRIC52 and the partially susceptible Reyan 7-33-97 cultivars, revealing the similar or differential changes of gene expressions between these two cultivars. This study has improved our overall understanding of the molecular mechanisms of rubber tree resistance to powdery mildew. PMID:26840302

  5. Identification of Powdery Mildew Responsive Genes in Hevea brasiliensis through mRNA Differential Display.

    PubMed

    Li, Xiang; Bi, Zhenghong; Di, Rong; Liang, Peng; He, Qiguang; Liu, Wenbo; Miao, Weiguo; Zheng, Fucong

    2016-01-01

    Powdery mildew is an important disease of rubber trees caused by Oidium heveae B. A. Steinmann. As far as we know, none of the resistance genes related to powdery mildew have been isolated from the rubber tree. There is little information available at the molecular level regarding how a rubber tree develops defense mechanisms against this pathogen. We have studied rubber tree mRNA transcripts from the resistant RRIC52 cultivar by differential display analysis. Leaves inoculated with the spores of O. heveae were collected from 0 to 120 hpi in order to identify pathogen-regulated genes at different infection stages. We identified 78 rubber tree genes that were differentially expressed during the plant-pathogen interaction. BLAST analysis for these 78 ESTs classified them into seven functional groups: cell wall and membrane pathways, transcription factor and regulatory proteins, transporters, signal transduction, phytoalexin biosynthesis, other metabolism functions, and unknown functions. The gene expression for eight of these genes was validated by qRT-PCR in both RRIC52 and the partially susceptible Reyan 7-33-97 cultivars, revealing the similar or differential changes of gene expressions between these two cultivars. This study has improved our overall understanding of the molecular mechanisms of rubber tree resistance to powdery mildew. PMID:26840302

  6. Nonsense-mediated decay regulates key components of homologous recombination

    PubMed Central

    Janke, Ryan; Kong, Jeremy; Braberg, Hannes; Cantin, Greg; Yates, John R.; Krogan, Nevan J.; Heyer, Wolf-Dietrich

    2016-01-01

    Cells frequently experience DNA damage that requires repair by homologous recombination (HR). Proteins involved in HR are carefully coordinated to ensure proper and efficient repair without interfering with normal cellular processes. In Saccharomyces cerevisiae, Rad55 functions in the early steps of HR and is regulated in response to DNA damage through phosphorylation by the Mec1 and Rad53 kinases of the DNA damage response. To further identify regulatory processes that target HR, we performed a high-throughput genetic interaction screen with RAD55 phosphorylation site mutants. Genes involved in the mRNA quality control process, nonsense-mediated decay (NMD), were found to genetically interact with rad55 phospho-site mutants. Further characterization revealed that RAD55 transcript and protein levels are regulated by NMD. Regulation of HR by NMD extends to multiple targets beyond RAD55, including RAD51, RAD54 and RAD57. Finally, we demonstrate that loss of NMD results in an increase in recombination rates and resistance to the DNA damaging agent methyl methanesulfonate, suggesting this pathway negatively regulates HR under normal growth conditions. PMID:27001511

  7. Estrogen Receptor α Regulates Dlx3-Mediated Osteoblast Differentiation

    PubMed Central

    Lee, Sung Ho; Oh, Kyo-Nyeo; Han, Younho; Choi, You Hee; Lee, Kwang-Youl

    2016-01-01

    Estrogen receptor α (ER-α), which is involved in bone metabolism and breast cancer, has been shown to have transcriptional targets. Dlx3 is essential for the skeletal development and plays an important role in osteoblast differentiation. Various osteogenic stimulators and transcription factors can induce the protein expression of Dlx3. However, the regulatory function of ER-α in the Dlx3 mediated osteogenic process remains unknown. Therefore, we investigated the regulation of Dlx3 and found that ER-α is a positive regulator of Dlx3 transcription in BMP2-induced osteoblast differentiation. We also found that ER-α interacts with Dlx3 and increases its transcriptional activity and DNA binding affinity. Furthermore, we demonstrated that the regulation of Dlx3 activity by ER-α is independent of the ligand (estradiol) binding domain. These results indicate that Dlx3 is a novel target of ER-α, and that ER-α regulates the osteoblast differentiation through modulation of Dlx3 expression and/or interaction with Dlx3. PMID:26674964

  8. Nonsense-mediated decay regulates key components of homologous recombination.

    PubMed

    Janke, Ryan; Kong, Jeremy; Braberg, Hannes; Cantin, Greg; Yates, John R; Krogan, Nevan J; Heyer, Wolf-Dietrich

    2016-06-20

    Cells frequently experience DNA damage that requires repair by homologous recombination (HR). Proteins involved in HR are carefully coordinated to ensure proper and efficient repair without interfering with normal cellular processes. In Saccharomyces cerevisiae, Rad55 functions in the early steps of HR and is regulated in response to DNA damage through phosphorylation by the Mec1 and Rad53 kinases of the DNA damage response. To further identify regulatory processes that target HR, we performed a high-throughput genetic interaction screen with RAD55 phosphorylation site mutants. Genes involved in the mRNA quality control process, nonsense-mediated decay (NMD), were found to genetically interact with rad55 phospho-site mutants. Further characterization revealed that RAD55 transcript and protein levels are regulated by NMD. Regulation of HR by NMD extends to multiple targets beyond RAD55, including RAD51, RAD54 and RAD57 Finally, we demonstrate that loss of NMD results in an increase in recombination rates and resistance to the DNA damaging agent methyl methanesulfonate, suggesting this pathway negatively regulates HR under normal growth conditions. PMID:27001511

  9. Thioredoxin-dependent regulation of AIF-mediated DNA damage.

    PubMed

    Shelar, Sandeep B; Kaminska, Kamila K; Reddy, Shridhivya A; Kumar, Dilip; Tan, Chong-Teik; Yu, Victor C; Lu, Jun; Holmgren, Arne; Hagen, Thilo; Chew, Eng-Hui

    2015-10-01

    The thioredoxin (Trx) system is one major redox system in mammalian cells. One of its component, Trx, is involved in redox homeostasis and many cellular biological processes through participating in disulfide reduction, S-nitrosylation/S-denitrosylation reactions and protein-protein interactions. In this study, we report the identification of a novel interaction between cytosolic/nuclear Trx1 and apoptosis inducing factor (AIF), and the redox sensitivity and biological significance of the Trx-AIF interaction was characterized. Cytosolic Trx1 but not mitochondrial Trx2 was observed to interact with AIF under physiological conditions and Trx1's active site cysteines were crucial for the interaction. Under oxidative stress conditions, Trx-AIF interaction was disrupted. When the treated cells were allowed to recover from oxidative stress by means of removal of the oxidants, interaction between Trx1 and AIF was re-established time-dependently, which underpins the biological relevance of a Trx-dependent redox regulation of AIF-mediated cell death. Indeed, in times of oxidative stress, nuclear translocation of AIF was found to occur concurrently with perturbations to the Trx-AIF interaction. Once localized in the nucleus, reduced Trx1 hindered the interaction between AIF and DNA, thereby bringing about an attenuation of AIF-mediated DNA damage. In conclusion, characterization of the Trx-AIF interaction has led to an understanding of the effect of reduced Trx1 on possibly regulating AIF-dependent cell death through impeding AIF-mediated DNA damage. Importantly, identification of the novel interaction between Trx1 and AIF has provided opportunities to design and develop therapeutically relevant strategies that either promote or prevent this protein-protein interaction for the treatment of different disease states. PMID:26119781

  10. A disulphide isomerase gene (PDI-V) from Haynaldia villosa contributes to powdery mildew resistance in common wheat

    PubMed Central

    Faheem, Muhammad; Li, Yingbo; Arshad, Muhammad; Jiangyue, Cheng; Jia, Zhao; Wang, Zongkuan; Xiao, Jin; Wang, Haiyan; Cao, Aizhong; Xing, Liping; Yu, Feifei; Zhang, Ruiqi; Xie, Qi; Wang, Xiue

    2016-01-01

    In this study, we report the contribution of a PDI-like gene from wheat wild relative Haynaldia villosa in combating powdery mildew. PDI-V protein contains two conserved thioredoxin (TRX) active domains (a and a′) and an inactive domain (b). PDI-V interacted with E3 ligase CMPG1-V protein, which is a positive regulator of powdery mildew response. PDI-V was mono-ubiquitinated by CMPG1-V without degradation being detected. PDI-V was located on H. villosa chromosome 5V and encoded for a protein located in the endoplasmic reticulum. Bgt infection in leaves of H. villosa induced PDI-V expression. Virus induced gene silencing of PDIs in a T. durum-H. villosa amphiploid compromised the resistance. Single cell transient over-expression of PDI-V or a truncated version containing the active TXR domain a decreased the haustorial index in moderately susceptible wheat cultivar Yangmai 158. Stable transgenic lines over-expressing PDI-V in Yangmai 158 displayed improved powdery mildew resistance at both the seedling and adult stages. By contrast over-expression of point-mutated PDI-VC57A did not increase the level of resistance in Yangmai 158. The above results indicate a pivotal role of PDI-V in powdery mildew resistance and showed that conserved TRX domain a is critical for its function. PMID:27071705

  11. A disulphide isomerase gene (PDI-V) from Haynaldia villosa contributes to powdery mildew resistance in common wheat.

    PubMed

    Faheem, Muhammad; Li, Yingbo; Arshad, Muhammad; Jiangyue, Cheng; Jia, Zhao; Wang, Zongkuan; Xiao, Jin; Wang, Haiyan; Cao, Aizhong; Xing, Liping; Yu, Feifei; Zhang, Ruiqi; Xie, Qi; Wang, Xiue

    2016-01-01

    In this study, we report the contribution of a PDI-like gene from wheat wild relative Haynaldia villosa in combating powdery mildew. PDI-V protein contains two conserved thioredoxin (TRX) active domains (a and a') and an inactive domain (b). PDI-V interacted with E3 ligase CMPG1-V protein, which is a positive regulator of powdery mildew response. PDI-V was mono-ubiquitinated by CMPG1-V without degradation being detected. PDI-V was located on H. villosa chromosome 5V and encoded for a protein located in the endoplasmic reticulum. Bgt infection in leaves of H. villosa induced PDI-V expression. Virus induced gene silencing of PDIs in a T. durum-H. villosa amphiploid compromised the resistance. Single cell transient over-expression of PDI-V or a truncated version containing the active TXR domain a decreased the haustorial index in moderately susceptible wheat cultivar Yangmai 158. Stable transgenic lines over-expressing PDI-V in Yangmai 158 displayed improved powdery mildew resistance at both the seedling and adult stages. By contrast over-expression of point-mutated PDI-V(C57A) did not increase the level of resistance in Yangmai 158. The above results indicate a pivotal role of PDI-V in powdery mildew resistance and showed that conserved TRX domain a is critical for its function. PMID:27071705

  12. Structural basis of antizyme-mediated regulation of polyamine homeostasis

    PubMed Central

    Wu, Hsiang-Yi; Chen, Shin-Fu; Hsieh, Ju-Yi; Chou, Fang; Wang, Yu-Hsuan; Lin, Wan-Ting; Lee, Pei-Ying; Yu, Yu-Jen; Lin, Li-Ying; Lin, Te-Sheng; Lin, Chieh-Liang; Liu, Guang-Yaw; Tzeng, Shiou-Ru; Hung, Hui-Chih; Chan, Nei-Li

    2015-01-01

    Polyamines are organic polycations essential for cell growth and differentiation; their aberrant accumulation is often associated with diseases, including many types of cancer. To maintain polyamine homeostasis, the catalytic activity and protein abundance of ornithine decarboxylase (ODC), the committed enzyme for polyamine biosynthesis, are reciprocally controlled by the regulatory proteins antizyme isoform 1 (Az1) and antizyme inhibitor (AzIN). Az1 suppresses polyamine production by inhibiting the assembly of the functional ODC homodimer and, most uniquely, by targeting ODC for ubiquitin-independent proteolytic destruction by the 26S proteasome. In contrast, AzIN positively regulates polyamine levels by competing with ODC for Az1 binding. The structural basis of the Az1-mediated regulation of polyamine homeostasis has remained elusive. Here we report crystal structures of human Az1 complexed with either ODC or AzIN. Structural analysis revealed that Az1 sterically blocks ODC homodimerization. Moreover, Az1 binding triggers ODC degradation by inducing the exposure of a cryptic proteasome-interacting surface of ODC, which illustrates how a substrate protein may be primed upon association with Az1 for ubiquitin-independent proteasome recognition. Dynamic and functional analyses further indicated that the Az1-induced binding and degradation of ODC by proteasome can be decoupled, with the intrinsically disordered C-terminal tail fragment of ODC being required only for degradation but not binding. Finally, the AzIN–Az1 structure suggests how AzIN may effectively compete with ODC for Az1 to restore polyamine production. Taken together, our findings offer structural insights into the Az-mediated regulation of polyamine homeostasis and proteasomal degradation. PMID:26305948

  13. Sirtuin 4 Regulates Lipopolysaccharide Mediated Leydig Cell Dysfunction.

    PubMed

    Ramatchandirin, Balamurugan; Sadasivam, Mohanraj; Kannan, Arun; Prahalathan, Chidambaram

    2016-04-01

    Bacterial lipopolysaccharide (LPS) is the most important contributing factor in pathogenesis of bacterial infection in male accessory glands; and it has shown to inhibit testicular steroidogenesis and induce apoptosis. The present study demonstrates that LPS causes mitochondrial dysfunction via suppression of sirtuin 4 (SIRT4); which in turn affects Leydig cell function by modulating steroidogenesis and apoptosis. LC-540 Leydig cells treated with LPS (10 µg/ml) showed impaired steroidogenesis and increased cellular apoptosis. The mRNA and protein expression of SIRT4 were decreased in LPS treated cells when compared to controls. The obtained data suggest that the c-Jun N-terminal kinase (JNK) activation suppresses SIRT4 expression in LPS treated Leydig cells. Furthermore, the overexpression of SIRT4 prevented LPS induced impaired steroidogenesis and cellular apoptosis by improving mitochondrial function. These findings provide valuable information that SIRT4 regulates LPS mediated Leydig cell dysfunction. PMID:26365714

  14. Hypoxia-mediated regulation of stem cell fate.

    PubMed

    Singh, Rashim Pal; Franke, Kristin; Wielockx, Ben

    2012-09-01

    Hypoxia-mediated regulation of stem cell fate, or reduced oxygen availability, is a prominent feature during mammalian development and under physiological and pathological conditions in adults. Oxygen-sensing is therefore indispensable as it enables the cells to adapt instantaneously to an inappropriate pO(2). This machinery relies primarily on hypoxia inducible factor (HIF). Moreover, a growing body of evidence proposes that different types of stem cells exist in a very hypoxic microenvironment, which may be beneficial for the maintenance of these cells and ensures continuous replenishment of dead or damaged cells in virtually all tissues of the body. Recent reports have shown that HIF is a critical player in these responses. However, a better understanding of the different HIF-related mechanisms is of utmost importance for the improvement of therapeutic strategies for tissue regeneration as well as hematological malignancies. PMID:22994515

  15. Dopamine receptor-mediated regulation of neuronal "clock" gene expression.

    PubMed

    Imbesi, M; Yildiz, S; Dirim Arslan, A; Sharma, R; Manev, H; Uz, T

    2009-01-23

    Using a transgenic mice model (i.e. "clock" knockouts), clock transcription factors have been suggested as critical regulators of dopaminergic behaviors induced by drugs of abuse. Moreover, it has been shown that systemic administration of psychostimulants, such as cocaine and methamphetamine regulates the striatal expression of clock genes. However, it is not known whether dopamine receptors mediate these regulatory effects of psychostimulants at the cellular level. Primary striatal neurons in culture express dopamine receptors as well as clock genes and have been successfully used in studying dopamine receptor functioning. Therefore, we investigated the role of dopamine receptors on neuronal clock gene expression in this model using specific receptor agonists. We found an inhibitory effect on the expression of mClock and mPer1 genes with the D2-class (i.e. D2/D3) receptor agonist quinpirole. We also found a generalized stimulatory effect on the expression of clock genes mPer1, mClock, mNPAS2 (neuronal PAS domain protein 2), and mBmal1 with the D1-class (i.e. D1) receptor agonist SKF38393. Further, we tested whether systemic administration of dopamine receptor agonists causes similar changes in striatal clock gene expression in vivo. We found quinpirole-induced alterations in mPER1 protein levels in the mouse striatum (i.e. rhythm shift). Collectively, our results indicate that the dopamine receptor system may mediate psychostimulant-induced changes in clock gene expression. Using striatal neurons in culture as a model, further research is needed to better understand how dopamine signaling modulates the expression dynamics of clock genes (i.e. intracellular signaling pathways) and thereby influences neuronal gene expression, neuronal transmission, and brain functioning. PMID:19017537

  16. Loss of Function in Mlo Orthologs Reduces Susceptibility of Pepper and Tomato to Powdery Mildew Disease Caused by Leveillula taurica

    PubMed Central

    Zheng, Zheng; Pavan, Stefano; Matsuda, Yoshinori; Toyoda, Hideyoshi; Wolters, Anne-Marie A.; Visser, Richard G. F.; Bai, Yuling

    2013-01-01

    Powdery mildew disease caused by Leveillula taurica is a serious fungal threat to greenhouse tomato and pepper production. In contrast to most powdery mildew species which are epiphytic, L. taurica is an endophytic fungus colonizing the mesophyll tissues of the leaf. In barley, Arabidopsis, tomato and pea, the correct functioning of specific homologues of the plant Mlo gene family has been found to be required for pathogenesis of epiphytic powdery mildew fungi. The aim of this study was to investigate the involvement of the Mlo genes in susceptibility to the endophytic fungus L. taurica. In tomato (Solanum lycopersicum), a loss-of-function mutation in the SlMlo1 gene results in resistance to powdery mildew disease caused by Oidium neolycopersici. When the tomato Slmlo1 mutant was inoculated with L. taurica in this study, it proved to be less susceptible compared to the control, S. lycopersicum cv. Moneymaker. Further, overexpression of SlMlo1 in the tomato Slmlo1 mutant enhanced susceptibility to L. taurica. In pepper, the CaMlo2 gene was isolated by applying a homology-based cloning approach. Compared to the previously identified CaMlo1 gene, the CaMlo2 gene is more similar to SlMlo1 as shown by phylogenetic analysis, and the expression of CaMlo2 is up-regulated at an earlier time point upon L. taurica infection. However, results of virus-induced gene silencing suggest that both CaMlo1 and CaMlo2 may be involved in the susceptibility of pepper to L. taurica. The fact that overexpression of CaMlo2 restored the susceptibility of the tomato Slmlo1 mutant to O. neolycopersici and increased its susceptibility to L. taurica confirmed the role of CaMlo2 acting as a susceptibility factor to different powdery mildews, though the role of CaMlo1 as a co-factor for susceptibility cannot be excluded. PMID:23923019

  17. FLC-mediated flowering repression is positively regulated by sumoylation

    PubMed Central

    Seo, Hak Soo

    2014-01-01

    Flowering locus C (FLC), a floral repressor, is a critical factor for the transition from the vegetative to the reproductive phase. Here, the mechanisms regulating the activity and stability of the FLC protein were investigated. Bimolecular fluorescence complementation and in vitro pull-down analyses showed that FLC interacts with the E3 small ubiquitin-like modifier (SUMO) ligase AtSIZ1, suggesting that AtSIZ1 is an E3 SUMO ligase for FLC. In vitro sumoylation assays showed that FLC is modified by SUMO in the presence of SUMO-activating enzyme E1 and conjugating enzyme E2, but its sumoylation is inhibited by AtSIZ1. In transgenic plants, inducible AtSIZ1 overexpression led to an increase in the concentration of FLC and delayed the post-translational decay of FLC, indicating that AtSIZ1 stabilizes FLC through direct binding. Also, the flowering time in mutant FLC (K154R, a mutation of the sumoylation site)-overexpressing plants was comparable with that in the wild type, whereas flowering was considerably delayed in FLC-overexpressing plants, supporting the notion that sumoylation is an important mechanism for FLC function. The data indicate that the sumoylation of FLC is critical for its role in the control of flowering time and that AtSIZ1 positively regulates FLC-mediated floral suppression. PMID:24218331

  18. Site Specific Cleavage Mediated by MMPs Regulates Function of Agrin

    PubMed Central

    McFarlane, Ainsley; Xie, Irene; Overall, Christopher M.; Stetefeld, Jörg

    2012-01-01

    Background Agrin is the key inducer of postsynaptic differentiations at the neuromuscular junction. The multidomain heparan sulfate proteoglycan is mediating via its N-terminal segment the interaction with laminin, whereas the C-terminal portion is responsible for Dystroglycan binding and clustering of the Acetylcholine receptor. Matrix metalloproteinases (MMP) are known to play essential roles in matrix remodeling, degradation and regulation of extracellular signaling networks. Principal Findings Site-specific processing of Agrin provides key insight into regulatory effects of Matrix metalloproteinases (MMPs). Here, we present a detailed study of agrin processing by different MMPs together with a molecular understanding of binding and cleavage at both terminal fragments. The data suggest for a regulatory effect of MMP cleavage at particularly important functional sites of agrin. Cleave of agrin abolishes the agrin-laminin complex formation and the Acetylcholine receptor clustering at the neuromuscular junction. Conclusion/Significance Agrin is a target of specific MMP processing resulting in agrin subfragments with different regulatory activities. MMP processing is a powerful tool to regulate extracellular signaling networks. PMID:22984437

  19. Lysine mediation of neuroendocrine food regulation in guinea fowl.

    PubMed

    Payne, A; Wang, X; Ivy, M T; Stewart, A; Nelson, K; Darris, C; Nahashon, S N

    2016-02-01

    In poultry, obesity is partly influenced by food intake, and is increasingly becoming a nationwide problem. Hypothalamic food intake mechanisms are involved metabolically and neurologically via two peptide hormones, leptin and ghrelin, and the amino acid glutamate, which is enzymatically derived from lysine metabolism. We hypothesize that lysine homeostasis mediates regulation of feed intake and performance characteristics via the brain-liver axis through glutamate sensing. The objective was to examine the effects of lysine homeostasis in avian food regulation and performance through neuroendocrine signaling. One-day-old male French Guinea fowl (GF) keets (n = 270) were weighed and randomly assigned to 5 dietary treatments (0.80%, 0.86%, 0.92%, 1.10% control, and 1.22% lysine) in 3 replicates. At 4 and 8 wk of age 20% of experimental birds were randomly selected, weighed and euthanatized. The liver, pancreas, and hypothalamus were excised, snap frozen in liquid nitrogen and stored at -80°C until use. Tissue mRNA was extracted and cDNA synthesized for qPCR assays. Lysine at 0.80 and 0.86% hindered growth, development of digestive organs, expression of brain and liver glutamate and leptin receptors, and caused high mortality in GF. The fold change for metabotropic glutamate receptor I was lower (P < 0.05) in liver and higher in brain at 0.86 and 0.92% than the control (1.10%) and 1.22% lysine. The 1.22% lysine exhibited highest expression of ionotropic glutamate receptor, while brain ghrelin receptor expression was highest at 0.86 and 0.92% lysine. Therefore, dietary lysine concentration may influence signaling pathways regulating food intake in brain-liver axis via glutamate synthesis. PMID:26614682

  20. Apoptosis signal-regulating kinase 1 mediates striatal degeneration via the regulation of C1q

    PubMed Central

    Cho, Kyoung Joo; Cheon, So Young; Kim, Gyung Whan

    2016-01-01

    Apoptosis signal-regulating kinase-1 (ASK1), an early signaling element in the cell death pathway, has been hypothesized to participate in the pathology of neurodegenerative diseases. The systemic administration of 3-nitropropionic acid (3-NP) facilitates the development of selective striatal lesions. However, it remains unclear whether specific neurons are selectively targeted in 3-NP-infused striatal degeneration. Recently, it has been proposed that complement-mediated synapse elimination may be reactivated aberrantly in the pathology of neurodegenerative diseases. We hypothesized that ASK1 is involved in striatal astrocyte reactivation; reactive astrocyte secretes molecules detrimental to neuron; and striatal neurons are more susceptible to these factors. Our results indicate that striatal astrocyte is reactivated and ASK1 level increases after 3-NP general and chronic infusion. Reactive striatal astrocyte increases TGF-beta differentially to cortex and striatum. ASK1 may be involved in regulation of astrocyte TGF-beta and it is linked to the C1q level in spatial and temporal, and moreover in the earlier stage of progressing striatal neuronal loss. Conclusively the present study suggests that ASK1 mediates 3-NP toxicity and regulates C1q level through the astrocyte TGF-beta. And also it may suggest that C1q level may be a surrogate of prediction marker representing neurodegenerative disease progress before developing behavioral impairment. PMID:26728245

  1. Variation within and between Vitis species for foliar resistance to the powdery mildew pathogen Erysiphe necator

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To complement existing control strategies, grape growers desire cultivars with resistance to powdery mildew caused by Erysiphe necator. Numerous disease resistance screens of diverse Vitis germplasm have been conducted previously to identify powdery mildew resistance, but ratings of named cultivars...

  2. p53 isoforms regulate astrocyte-mediated neuroprotection and neurodegeneration.

    PubMed

    Turnquist, C; Horikawa, I; Foran, E; Major, E O; Vojtesek, B; Lane, D P; Lu, X; Harris, B T; Harris, C C

    2016-09-01

    Bidirectional interactions between astrocytes and neurons have physiological roles in the central nervous system and an altered state or dysfunction of such interactions may be associated with neurodegenerative diseases, such as Alzheimer's disease (AD) and amyotrophic lateral sclerosis (ALS). Astrocytes exert structural, metabolic and functional effects on neurons, which can be either neurotoxic or neuroprotective. Their neurotoxic effect is mediated via the senescence-associated secretory phenotype (SASP) involving pro-inflammatory cytokines (e.g., IL-6), while their neuroprotective effect is attributed to neurotrophic growth factors (e.g., NGF). We here demonstrate that the p53 isoforms Δ133p53 and p53β are expressed in astrocytes and regulate their toxic and protective effects on neurons. Primary human astrocytes undergoing cellular senescence upon serial passaging in vitro showed diminished expression of Δ133p53 and increased p53β, which were attributed to the autophagic degradation and the SRSF3-mediated alternative RNA splicing, respectively. Early-passage astrocytes with Δ133p53 knockdown or p53β overexpression were induced to show SASP and to exert neurotoxicity in co-culture with neurons. Restored expression of Δ133p53 in near-senescent, otherwise neurotoxic astrocytes conferred them with neuroprotective activity through repression of SASP and induction of neurotrophic growth factors. Brain tissues from AD and ALS patients possessed increased numbers of senescent astrocytes and, like senescent astrocytes in vitro, showed decreased Δ133p53 and increased p53β expression, supporting that our in vitro findings recapitulate in vivo pathology of these neurodegenerative diseases. Our finding that Δ133p53 enhances the neuroprotective function of aged and senescent astrocytes suggests that the p53 isoforms and their regulatory mechanisms are potential targets for therapeutic intervention in neurodegenerative diseases. PMID:27104929

  3. Regulation of T cell receptor complex-mediated signaling by ubiquitin and ubiquitin-like modifications

    PubMed Central

    Friend, Samantha F; Deason-Towne, Francina; Peterson, Lisa K; Berger, Allison J; Dragone, Leonard L

    2014-01-01

    Post-translational protein modifications are a dynamic method of regulating protein function in response to environmental signals. As with any cellular process, T cell receptor (TCR) complex-mediated signaling is highly regulated, since the strength and duration of TCR-generated signals governs T cell development and activation. While regulation of TCR complex-mediated signaling by phosphorylation has been well studied, regulation by ubiquitin and ubiquitin-like modifiers is still an emerging area of investigation. This review will examine how ubiquitin, E3 ubiquitin ligases, and other ubiquitin-like modifications such as SUMO and NEDD8 regulate TCR complex-mediated signaling. PMID:25628960

  4. Regulation of T cell receptor complex-mediated signaling by ubiquitin and ubiquitin-like modifications.

    PubMed

    Friend, Samantha F; Deason-Towne, Francina; Peterson, Lisa K; Berger, Allison J; Dragone, Leonard L

    2014-01-01

    Post-translational protein modifications are a dynamic method of regulating protein function in response to environmental signals. As with any cellular process, T cell receptor (TCR) complex-mediated signaling is highly regulated, since the strength and duration of TCR-generated signals governs T cell development and activation. While regulation of TCR complex-mediated signaling by phosphorylation has been well studied, regulation by ubiquitin and ubiquitin-like modifiers is still an emerging area of investigation. This review will examine how ubiquitin, E3 ubiquitin ligases, and other ubiquitin-like modifications such as SUMO and NEDD8 regulate TCR complex-mediated signaling. PMID:25628960

  5. Identification and structure of the mating-type locus and development of PCR-based markers for mating type in powdery mildew fungi

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In fungi, mating compatibility is regulated by mating-type loci. The objectives of this study were to identify and sequence mating-type genes at the MAT1 locus in the grape powdery mildew fungus, Erysiphe necator, to develop a PCR-based marker for determining mating type in E. necator, and to devel...

  6. Vesicular Nucleotide Transporter-Mediated ATP Release Regulates Insulin Secretion

    PubMed Central

    Geisler, Jessica C.; Corbin, Kathryn L.; Li, Qin; Feranchak, Andrew P.; Nunemaker, Craig S.

    2013-01-01

    Extracellular ATP plays a critical role in regulating insulin secretion in pancreatic β cells. The ATP released from insulin secretory vesicles has been proposed to be a major source of extracellular ATP. Currently, the mechanism by which ATP accumulates into insulin secretory granules remains elusive. In this study, the authors identified the expression of a vesicular nucleotide transporter (VNUT) in mouse pancreas, isolated mouse islets, and MIN6 cells, a mouse β cell line. Immunohistochemistry and immunofluorescence revealed that VNUT colocalized extensively with insulin secretory granules. Functional studies showed that suppressing endogenous VNUT expression in β cells by small hairpin RNA knockdown greatly reduced basal- and glucose-induced ATP release. Importantly, knocking down VNUT expression by VNUT small hairpin RNA in MIN6 cells and isolated mouse islets dramatically suppressed basal insulin release and glucose-stimulated insulin secretion (GSIS). Moreover, acute pharmacologic blockade of VNUT with Evans blue, a VNUT antagonist, greatly attenuated GSIS in a dose-dependent manner. Exogenous ATP treatment effectively reversed the insulin secretion defect induced by both VNUT knockdown and functional inhibition, indicating that VNUT-mediated ATP release is essential for maintaining normal insulin secretion. In contrast to VNUT knockdown, overexpression of VNUT in β cells resulted in excessive ATP release and enhanced basal insulin secretion and GSIS. Elevated insulin secretion induced by VNUT overexpression was reversed by pharmacologic inhibition of P2X but not P2Y purinergic receptors. This study reveals VNUT is expressed in pancreatic β cells and plays an essential and novel role in regulating insulin secretion through vesicular ATP release and extracellular purinergic signaling. PMID:23254199

  7. Calcineurin mediates homeostatic synaptic plasticity by regulating retinoic acid synthesis

    PubMed Central

    Arendt, Kristin L.; Zhang, Zhenjie; Ganesan, Subhashree; Hintze, Maik; Shin, Maggie M.; Tang, Yitai; Cho, Ahryon; Graef, Isabella A.; Chen, Lu

    2015-01-01

    Homeostatic synaptic plasticity is a form of non-Hebbian plasticity that maintains stability of the network and fidelity for information processing in response to prolonged perturbation of network and synaptic activity. Prolonged blockade of synaptic activity decreases resting Ca2+ levels in neurons, thereby inducing retinoic acid (RA) synthesis and RA-dependent homeostatic synaptic plasticity; however, the signal transduction pathway that links reduced Ca2+-levels to RA synthesis remains unknown. Here we identify the Ca2+-dependent protein phosphatase calcineurin (CaN) as a key regulator for RA synthesis and homeostatic synaptic plasticity. Prolonged inhibition of CaN activity promotes RA synthesis in neurons, and leads to increased excitatory and decreased inhibitory synaptic transmission. These effects of CaN inhibitors on synaptic transmission are blocked by pharmacological inhibitors of RA synthesis or acute genetic deletion of the RA receptor RARα. Thus, CaN, acting upstream of RA, plays a critical role in gating RA signaling pathway in response to synaptic activity. Moreover, activity blockade-induced homeostatic synaptic plasticity is absent in CaN knockout neurons, demonstrating the essential role of CaN in RA-dependent homeostatic synaptic plasticity. Interestingly, in GluA1 S831A and S845A knockin mice, CaN inhibitor- and RA-induced regulation of synaptic transmission is intact, suggesting that phosphorylation of GluA1 C-terminal serine residues S831 and S845 is not required for CaN inhibitor- or RA-induced homeostatic synaptic plasticity. Thus, our study uncovers an unforeseen role of CaN in postsynaptic signaling, and defines CaN as the Ca2+-sensing signaling molecule that mediates RA-dependent homeostatic synaptic plasticity. PMID:26443861

  8. Calcineurin mediates homeostatic synaptic plasticity by regulating retinoic acid synthesis.

    PubMed

    Arendt, Kristin L; Zhang, Zhenjie; Ganesan, Subhashree; Hintze, Maik; Shin, Maggie M; Tang, Yitai; Cho, Ahryon; Graef, Isabella A; Chen, Lu

    2015-10-20

    Homeostatic synaptic plasticity is a form of non-Hebbian plasticity that maintains stability of the network and fidelity for information processing in response to prolonged perturbation of network and synaptic activity. Prolonged blockade of synaptic activity decreases resting Ca(2+) levels in neurons, thereby inducing retinoic acid (RA) synthesis and RA-dependent homeostatic synaptic plasticity; however, the signal transduction pathway that links reduced Ca(2+)-levels to RA synthesis remains unknown. Here we identify the Ca(2+)-dependent protein phosphatase calcineurin (CaN) as a key regulator for RA synthesis and homeostatic synaptic plasticity. Prolonged inhibition of CaN activity promotes RA synthesis in neurons, and leads to increased excitatory and decreased inhibitory synaptic transmission. These effects of CaN inhibitors on synaptic transmission are blocked by pharmacological inhibitors of RA synthesis or acute genetic deletion of the RA receptor RARα. Thus, CaN, acting upstream of RA, plays a critical role in gating RA signaling pathway in response to synaptic activity. Moreover, activity blockade-induced homeostatic synaptic plasticity is absent in CaN knockout neurons, demonstrating the essential role of CaN in RA-dependent homeostatic synaptic plasticity. Interestingly, in GluA1 S831A and S845A knockin mice, CaN inhibitor- and RA-induced regulation of synaptic transmission is intact, suggesting that phosphorylation of GluA1 C-terminal serine residues S831 and S845 is not required for CaN inhibitor- or RA-induced homeostatic synaptic plasticity. Thus, our study uncovers an unforeseen role of CaN in postsynaptic signaling, and defines CaN as the Ca(2+)-sensing signaling molecule that mediates RA-dependent homeostatic synaptic plasticity. PMID:26443861

  9. Examination of marker-assisted selection for powdery and downy mildew resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Field grown seedlings potentially carrying both the Run1 and Rpv1 loci for powdery and downy mildew resistance, respectively, from muscadine introgressions were assessed for resistance to both powdery and downy mildew. Powdery mildew was assessed in the field under no-spray conditions, while down mi...

  10. Affect Regulation as a Mediator of Attachment and Deliberate Self-Harm

    ERIC Educational Resources Information Center

    Kimball, Joan S.; Diddams, Margaret

    2007-01-01

    The authors used structural equation modeling to test the mediational role of affect regulation on attachment and deliberate self-harm in 216 undergraduates. Results suggest that affect regulation mediates the relationship between attachment and deliberate self-harm, providing support for the theoretical importance of attachment and affect…

  11. H-Ras regulation of TRAIL death receptor mediated apoptosis

    PubMed Central

    Chen, Jun-Jie; Bozza, William P.; Di, Xu; Zhang, Yaqin; Hallett, William; Zhang, Baolin

    2014-01-01

    TNF-related apoptosis-inducing ligand (TRAIL) induces apoptosis through the death receptors (DRs) 4 and/or 5 expressed on the cell surface. Multiple clinical trials are underway to evaluate the antitumor activity of recombinant human TRAIL and agonistic antibodies to DR4 or DR5. However, their therapeutic potential is limited by the high frequency of cancer resistance. Here we provide evidence demonstrating the role of H-Ras in TRAIL receptor mediated apoptosis. By analyzing the genome wide mRNA expression data of the NCI60 cancer cell lines, we found that H-Ras expression was consistently upregulated in TRAIL-resistant cell lines. By contrast, no correlation was found between TRAIL sensitivity and K-Ras expression levels or their mutational profiles. Notably, H-Ras upregulation associated with a surface deficiency of TRAIL death receptors. Selective inhibition of H-Ras activity in TRAIL-resistant cells restored the surface expression of both DR4 and DR5 without changing their total protein levels. The resulting cells became highly susceptible to both TRAIL and agonistic DR5 antibody, whereas K-Ras inhibition had little or no effect on TRAIL-induced apoptosis, indicating H-Ras plays a distinct role in the regulation of TRAIL death receptors. Further studies are warranted to determine the therapeutic potential of H-Ras-specific inhibitors in combination with TRAIL receptor agonists. PMID:25026275

  12. Sympathetic dysfunction mediating cardiovascular regulation in alcoholic neuropathy.

    PubMed

    Chida, K; Takasu, T; Mori, N; Tokunaga, K; Komatsu, K; Kawamura, H

    1994-01-01

    We evaluated autonomic function in alcoholic neuropathy by non-invasive hemodynamic studies using servo-plethysmo-manometry and laser Doppler flowmetry. In 16 alcoholics compared with 17 age-matched healthy controls, the extent of AP responses to phase IV of the Valsalva maneuver, cold pressor test and isometric exercise decreased significantly, indicating sympathetic hypofunction. Five patients had paradoxical depressor response to cold pressor test, which was reproducible by submersion of the hand in warm water. The data suggest involvement of the higher autonomic center. Reflex bradycardia in phase IV of the Valsalva maneuver and the variation coefficient of R-R intervals in the electrocardiogram were lower, indicating parasympathetic hypofunction. Cutaneous blood flow response in the foot upon local warming and submersion of the hand in cold water was significantly impaired, but that in the contralateral hand was not. Abnormalities in the autonomic function tests may result from a toxic effect of ethanol on the peripheral and central autonomic nervous system and the cardiovascular system. The results indicate not only parasympathetic (vagal) but sympathetic dysfunction mediating cardiovascular regulation in patients with alcoholic neuropathy. PMID:7926889

  13. Phosphatidic Acid-Mediated Signaling Regulates Microneme Secretion in Toxoplasma.

    PubMed

    Bullen, Hayley E; Jia, Yonggen; Yamaryo-Botté, Yoshiki; Bisio, Hugo; Zhang, Ou; Jemelin, Natacha Klages; Marq, Jean-Baptiste; Carruthers, Vern; Botté, Cyrille Y; Soldati-Favre, Dominique

    2016-03-01

    The obligate intracellular lifestyle of apicomplexan parasites necessitates an invasive phase underpinned by timely and spatially controlled secretion of apical organelles termed micronemes. In Toxoplasma gondii, extracellular potassium levels and other stimuli trigger a signaling cascade culminating in phosphoinositide-phospholipase C (PLC) activation, which generates the second messengers diacylglycerol (DAG) and IP3 and ultimately results in microneme secretion. Here we show that a delicate balance between DAG and its downstream product, phosphatidic acid (PA), is essential for controlling microneme release. Governing this balance is the apicomplexan-specific DAG-kinase-1, which interconverts PA and DAG, and whose depletion impairs egress and causes parasite death. Additionally, we identify an acylated pleckstrin-homology (PH) domain-containing protein (APH) on the microneme surface that senses PA during microneme secretion and is necessary for microneme exocytosis. As APH is conserved in Apicomplexa, these findings highlight a potentially widely used mechanism in which key lipid mediators regulate microneme exocytosis. PMID:26962945

  14. Osmotic regulation of Rab-mediated organelle docking

    PubMed Central

    Brett, Christopher L.; Merz, Alexey J.

    2009-01-01

    SUMMARY Osmotic gradients across organelle and plasma membranes modulate the rates of membrane fission and fusion; sufficiently large gradients can cause membrane rupture [1–6]. Hypotonic gradients applied to living yeast cells trigger prompt (within seconds) swelling and fusion of Saccharomyces cerevisiae vacuoles, while hypertonic gradients cause vacuoles to fragment on a slower time scale [7–11]. Here, we analyze the influence of osmotic strength on homotypic fusion of isolated yeast vacuoles. Consistent with previously reported in vivo results, we find that decreases in osmolyte concentration increase the rate and extent of vacuole fusion in vitro, while increases in osmolyte concentration prevent fusion. Unexpectedly, our results reveal that osmolytes regulate fusion by inhibiting early, Rab-dependent docking or predocking events, not late events. Our experiments reveal an organelle-autonomous pathway that may control organelle surface to volume ratio, size and copy number: decreasing the osmolyte concentration in the cytoplasmic compartment accelerates Rab-mediated docking and fusion. Fusion, by altering the organelle surface-to-enclosed volume relationship, in turn reduces the risk of membrane rupture. PMID:18619842

  15. Hoarding and eating pathology: the mediating role of emotion regulation.

    PubMed

    Raines, Amanda M; Boffa, Joseph W; Allan, Nicholas P; Short, Nicole A; Schmidt, Norman B

    2015-02-01

    Hoarding disorder is characterized by persistent difficulty discarding possessions resulting in clutter that precludes one from using living areas for their intended purposes. The limited empirical work available has suggested a strong link between hoarding and various non-psychiatric conditions, including obesity. Despite these associations, no research has examined the link between hoarding and other forms of eating pathology including symptoms associated with binge eating. Moreover, little is known about mechanisms that may account for this relationship. The current study examined the associations between hoarding severity, obesity, and symptoms associated with binge eating in a sample (N=97) of individuals with elevated hoarding symptoms. Results revealed that hoarding severity was associated with increased body mass index (BMI) and symptoms of binge eating. In addition, difficulties regulating emotions mediated the association between hoarding and eating concerns. Considering the lack of information on hoarding behaviors, as well as its classification as a new diagnosis within DSM-5, these findings add considerably to a growing body of literature on hoarding disorder. PMID:25440599

  16. Osmotic regulation of Rab-mediated organelle docking.

    PubMed

    Brett, Christopher L; Merz, Alexey J

    2008-07-22

    Osmotic gradients across organelle and plasma membranes modulate the rates of membrane fission and fusion; sufficiently large gradients can cause membrane rupture [1-6]. Hypotonic gradients applied to living yeast cells trigger prompt (within seconds) swelling and fusion of Saccharomyces cerevisiae vacuoles, whereas hypertonic gradients cause vacuoles to fragment on a slower time scale [7-11]. Here, we analyze the influence of osmotic strength on homotypic fusion of isolated yeast vacuoles. Consistent with previously reported in vivo results, we find that decreases in osmolyte concentration increase the rate and extent of vacuole fusion in vitro, whereas increases in osmolyte concentration prevent fusion. Unexpectedly, our results reveal that osmolytes regulate fusion by inhibiting early Rab-dependent docking or predocking events, not late events. Our experiments reveal an organelle-autonomous pathway that may control organelle surface-to-volume ratio, size, and copy number: Decreasing the osmolyte concentration in the cytoplasmic compartment accelerates Rab-mediated docking and fusion. By altering the relationship between the organelle surface and its enclosed volume, fusion in turn reduces the risk of membrane rupture. PMID:18619842

  17. Notch-mediated lateral inhibition regulates proneural wave propagation when combined with EGF-mediated reaction diffusion.

    PubMed

    Sato, Makoto; Yasugi, Tetsuo; Minami, Yoshiaki; Miura, Takashi; Nagayama, Masaharu

    2016-08-30

    Notch-mediated lateral inhibition regulates binary cell fate choice, resulting in salt and pepper patterns during various developmental processes. However, how Notch signaling behaves in combination with other signaling systems remains elusive. The wave of differentiation in the Drosophila visual center or "proneural wave" accompanies Notch activity that is propagated without the formation of a salt and pepper pattern, implying that Notch does not form a feedback loop of lateral inhibition during this process. However, mathematical modeling and genetic analysis clearly showed that Notch-mediated lateral inhibition is implemented within the proneural wave. Because partial reduction in EGF signaling causes the formation of the salt and pepper pattern, it is most likely that EGF diffusion cancels salt and pepper pattern formation in silico and in vivo. Moreover, the combination of Notch-mediated lateral inhibition and EGF-mediated reaction diffusion enables a function of Notch signaling that regulates propagation of the wave of differentiation. PMID:27535937

  18. The Arabidopsis mediator complex subunits MED16, MED14, and MED2 regulate mediator and RNA polymerase II recruitment to CBF-responsive cold-regulated genes.

    PubMed

    Hemsley, Piers A; Hurst, Charlotte H; Kaliyadasa, Ewon; Lamb, Rebecca; Knight, Marc R; De Cothi, Elizabeth A; Steele, John F; Knight, Heather

    2014-01-01

    The Mediator16 (MED16; formerly termed SENSITIVE TO FREEZING6 [SFR6]) subunit of the plant Mediator transcriptional coactivator complex regulates cold-responsive gene expression in Arabidopsis thaliana, acting downstream of the C-repeat binding factor (CBF) transcription factors to recruit the core Mediator complex to cold-regulated genes. Here, we use loss-of-function mutants to show that RNA polymerase II recruitment to CBF-responsive cold-regulated genes requires MED16, MED2, and MED14 subunits. Transcription of genes known to be regulated via CBFs binding to the C-repeat motif/drought-responsive element promoter motif requires all three Mediator subunits, as does cold acclimation-induced freezing tolerance. In addition, these three subunits are required for low temperature-induced expression of some other, but not all, cold-responsive genes, including genes that are not known targets of CBFs. Genes inducible by darkness also required MED16 but required a different combination of Mediator subunits for their expression than the genes induced by cold. Together, our data illustrate that plants control transcription of specific genes through the action of subsets of Mediator subunits; the specific combination defined by the nature of the stimulus but also by the identity of the gene induced. PMID:24415770

  19. The LIF-mediated molecular signature regulating murine embryo implantation.

    PubMed

    Rosario, Gracy X; Hondo, Eiichi; Jeong, Jae-Wook; Mutalif, Rafidah; Ye, Xiaoqian; Yee, Li Xuan; Stewart, Colin L

    2014-09-01

    The establishment of a receptive uterus is the prime requirement for embryo implantation. In mice, the E2-induced cytokine leukemia inhibitory factor (LIF) is essential in switching the uterine luminal epithelium (LE) from a nonreceptive to a receptive state. Here we define the LIF-mediated switch using array analysis and informatics to identify LIF-induced changes in gene expression and annotated signaling pathways specific to the LE. We compare gene expression profiles at 0, 1, 3, and 6 h, following LIF treatment. During the first hour, the JAK-STAT signaling pathway is activated and the expression of 54 genes declines, primarily affecting LE cytoskeletal and chromatin organization as well as a transient reduction in the progesterone, TGFbetaR1, and ACVR1 receptors. Simultaneously 256 genes increase expression, of which 42 are transcription factors, including Sox, Kfl, Hes, Hey, and Hox families. Within 3 h, the expression of 3987 genes belonging to more than 25 biological process pathways was altered. We confirmed the mRNA and protein distribution of key genes from 10 pathways, including the Igf-1, Vegf, Toll-like receptors, actin cytoskeleton, ephrin, integrins, TGFbeta, Wnt, and Notch pathways. These data identify novel LIF-activated pathways in the LE and define the molecular basis between the refractory and receptive uterine phases. More broadly, these findings highlight the staggering capacity of a single cytokine to induce a dynamic and complex network of changes in a simple epithelium essential to mammalian reproduction and provide a basis for identifying new routes to regulating female reproduction. PMID:25031358

  20. Dominant negative RPW8.2 fusion proteins reveal the importance of haustorium-oriented protein trafficking for resistance against powdery mildew in Arabidopsis.

    PubMed

    Zhang, Qiong; Berkey, Robert; Pan, Zhiyong; Wang, Wenming; Zhang, Yi; Ma, Xianfeng; King, Harlan; Xiao, Shunyuan

    2015-01-01

    Powdery mildew fungi form feeding structures called haustoria inside epidermal cells of host plants to extract photosynthates for their epiphytic growth and reproduction. The haustorium is encased by an interfacial membrane termed the extrahaustorial membrane (EHM). The atypical resistance protein RPW8.2 from Arabidopsis is specifically targeted to the EHM where RPW8.2 activates haustorium-targeted (thus broad-spectrum) resistance against powdery mildew fungi. EHM-specific localization of RPW8.2 suggests the existence of an EHM-oriented protein/membrane trafficking pathway during EHM biogenesis. However, the importance of this specific trafficking pathway for host defense has not been evaluated via a genetic approach without affecting other trafficking pathways. Here, we report that expression of EHM-oriented, nonfunctional RPW8.2 chimeric proteins exerts dominant negative effect over functional RPW8.2 and potentially over other EHM-localized defense proteins, thereby compromising both RPW8.2-mediated and basal resistance to powdery mildew. Thus, our results highlight the importance of the EHM-oriented protein/membrane trafficking pathway for host resistance against haustorium-forming pathogens such as powdery mildew fungi. PMID:25830634

  1. Mutations in PMR5 result in powdery mildew resistance and altered cell wall composition.

    PubMed

    Vogel, John P; Raab, Theodore K; Somerville, Chris R; Somerville, Shauna C

    2004-12-01

    Powdery mildews and other obligate biotrophic pathogens are highly adapted to their hosts and often show limited host ranges. One facet of such host specialization is likely to be penetration of the host cell wall, a major barrier to infection. A mutation in the pmr5 gene rendered Arabidopsis resistant to the powdery mildew species Erysiphe cichoracearum and Erysiphe orontii, but not to the unrelated pathogens Pseudomonas syringae or Peronospora parasitica. PMR5 belongs to a large family of plant-specific genes of unknown function. pmr5-mediated resistance did not require signaling through either the salicylic acid or jasmonic acid/ethylene defense pathways, suggesting resistance in this mutant may be due either to the loss of a susceptibility factor or to the activation of a novel form of defense. Based on Fourier transform infrared analysis, the pmr5 cell walls were enriched in pectin and exhibited a reduced degree of pectin modification relative to wild-type cell walls. In addition, the mutant had smaller cells, suggesting a defect in cell expansion. A double mutant with pmr6 (defective in a glycosylphosphatidylinositol-anchored pectate lyase-like gene) exhibited a strong increase in total uronic acid content and a more severe reduction in size, relative to the single mutants, suggesting that the two genes affect pectin composition, either directly or indirectly, via different mechanisms. These two mutants highlight the importance of the host cell wall in plant-microbe interactions. PMID:15584961

  2. Powdery Emulsion Explosive: A New Excellent Industrial Explosive

    NASA Astrophysics Data System (ADS)

    Ni, Ouqi; Zhang, Kaiming; Yu, Zhengquan; Tang, Shujuan

    2012-07-01

    Powdery emulsion explosive (PEE), a new powdery industrial explosive with perfect properties, has been made using an emulsification-spray drying technique. PEE is composed of 91-92.5 wt% ammonium nitrate (AN), 4.5-6 wt% organic fuels, and 1.5-1.8 wt% water. Due to its microstructure as a water-in-oil (W/O) emulsion and low water content, it has excellent detonation performance, outstanding water resistance, reliable safety, and good application compared with other industrial explosives, such as ammonite, emulsion explosives, and ANFO.

  3. Dysfunctional miRNA-Mediated Regulation in Chromophobe Renal Cell Carcinoma

    PubMed Central

    Sun, Xiaohan; Zhang, Junying

    2016-01-01

    Past research on pathogenesis of a complex disease suggests that differentially expressed message RNAs (mRNAs) can be noted as biomarkers of a disease. However, significant miRNA-mediated regulation change might also be more deep underlying cause of a disease. In this study, a miRNA-mediated regulation module is defined based on GO terms (Gene Ontology terms) from which dysfunctional modules are identified as the suspected cause of a disease. A miRNA-mediated regulation module contains mRNAs annotated to a GO term and MicroRNAs (miRNAs) which regulate the mRNAs. Based on the miRNA-mediated regulation coefficients estimated from the expression profiles of the mRNA and the miRNAs, a SW (single regulation-weight) value is then designed to evaluate the miRNA-mediated regulation change of an mRNA, and the modules with significantly differential SW values are thus identified as dysfunctional modules. The approach is applied to Chromophobe renal cell carcinoma and it identifies 70 dysfunctional miRNA-mediated regulation modules from initial 4381 modules. The identified dysfunctional modules are detected to be comprehensive reflection of chromophobe renal cell carcinoma. The proposed approach suggests that accumulated alteration in miRNA-mediated regulation might cause functional alterations, which further cause a disease. Moreover, this approach can also be used to identify diffentially miRNA-mediated regulated mRNAs showing more comprehensive underlying association with a disease than differentially expressed mRNAs. PMID:27258182

  4. Lyn, PKC-δ, SHIP-1 interactions regulate GPVI-mediated platelet-dense granule secretion

    PubMed Central

    Chari, Ramya; Kim, Soochong; Murugappan, Swaminathan; Sanjay, Archana; Daniel, James L.

    2009-01-01

    Protein kinase C-δ (PKC-δ) is expressed in platelets and activated downstream of protease-activated receptors (PARs) and glycoprotein VI (GPVI) receptors. We have previously shown that PKC-δ positively regulates PAR-mediated dense granule secretion, whereas it negatively regulates GPVI-mediated dense granule secretion. We further investigated the mechanism of such differential regulation of dense granule release by PKC-δ in platelets. SH2 domain–containing inositol phosphatase-1 (SHIP-1) is phosphorylated on Y1020, a marker for its activation, upon stimulation of human platelets with PAR agonists SFLLRN and AYPGKF or GPVI agonist convulxin. GPVI-mediated SHIP-1 phosphorylation occurred rapidly at 15 seconds, whereas PAR-mediated phosphorylation was delayed, occurring at 1 minute. Lyn and SHIP-1, but not SHIP-2 or Shc, preferentially associated with PKC-δ on stimulation of platelets with a GPVI agonist, but not with a PAR agonist. In PKC-δ–null murine platelets, convulxin-induced SHIP-1 phosphorylation was inhibited. Furthermore, in Lyn null murine platelets, GPVI-mediated phosphorylations on Y-1020 of SHIP-1 and Y311 of PKC-δ were inhibited. In murine platelets lacking Lyn or SHIP-1, GPVI-mediated dense granule secretions are potentiated, whereas PAR-mediated dense granule secretions are inhibited. Therefore, we conclude that Lyn-mediated phosphorylations of PKC-δ and SHIP-1 and their associations negatively regulate GPVI-mediated dense granule secretion in platelets. PMID:19587372

  5. Mutation of the Glucosinolate Biosynthesis Enzyme Cytochrome P450 83A1 Monooxygenase Increases Camalexin Accumulation and Powdery Mildew Resistance

    PubMed Central

    Liu, Simu; Bartnikas, Lisa M.; Volko, Sigrid M.; Ausubel, Frederick M.; Tang, Dingzhong

    2016-01-01

    Small secondary metabolites, including glucosinolates and the major phytoalexin camalexin, play important roles in immunity in Arabidopsis thaliana. We isolated an Arabidopsis mutant with increased resistance to the powdery mildew fungus Golovinomyces cichoracearum and identified a mutation in the gene encoding cytochrome P450 83A1 monooxygenase (CYP83A1), which functions in glucosinolate biosynthesis. The cyp83a1-3 mutant exhibited enhanced defense responses to G. cichoracearum and double mutant analysis showed that this enhanced resistance requires NPR1, EDS1, and PAD4, but not SID2 or EDS5. In cyp83a1-3 mutants, the expression of genes related to camalexin synthesis increased upon G. cichoracearum infection. Significantly, the cyp83a1-3 mutant also accumulated higher levels of camalexin. Decreasing camalexin levels by mutation of the camalexin synthetase gene PAD3 or the camalexin synthesis regulator AtWRKY33 compromised the powdery mildew resistance in these mutants. Consistent with these observations, overexpression of PAD3 increased camalexin levels and enhanced resistance to G. cichoracearum. Taken together, our data indicate that accumulation of higher levels of camalexin contributes to increased resistance to powdery mildew. PMID:26973671

  6. Emotion Regulation Factors as Mediators between Body Dissatisfaction and Bulimic Symptoms in Early Adolescent Girls

    ERIC Educational Resources Information Center

    Sim, Leslie; Zeman, Janice

    2005-01-01

    Research suggests that negative affect is an important mediator in the relationship between body dissatisfaction and bulimic symptoms. This study examines the mediational role of specific emotion regulation processes (i.e., negative emotionality, poor awareness of emotion, nonconstructive coping with negative emotion) in bulimic symptoms. In…

  7. Marker-assisted selection for powdery mildew in pea

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew, caused by Erysiphe pisi Syd. is an important pathogen of pea (Pisum sativum L.) worldwide. It causes significant yield loss and reduced crop quality when present in epidemic proportion. Genetic resistance is controlled by two recessive alleles, er-1 and er-2. Availability of co-domin...

  8. Phoma glomerata as a mycoparasite of powdery mildew.

    PubMed

    Sullivan, R F; White, J F

    2000-01-01

    Ampelomyces and Phoma species are frequently confused with each other. Isolates previously attributed to the genus Ampelomyces were shown to be Phoma isolates through studies of their morphology and life cycle and ribosomal DNA internal transcribed spacer region 1 sequence analysis. Phoma glomerata can colonize and suppress development of powdery mildew on oak and may have utility as a mycoparasitic agent. PMID:10618259

  9. Phoma glomerata as a Mycoparasite of Powdery Mildew

    PubMed Central

    Sullivan, Raymond F.; White, James F.

    2000-01-01

    Ampelomyces and Phoma species are frequently confused with each other. Isolates previously attributed to the genus Ampelomyces were shown to be Phoma isolates through studies of their morphology and life cycle and ribosomal DNA internal transcribed spacer region 1 sequence analysis. Phoma glomerata can colonize and suppress development of powdery mildew on oak and may have utility as a mycoparasitic agent. PMID:10618259

  10. Breeding Table Grapes with Resistance to Powdery Mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Different mechanisms for resistance to powdery mildew exist in grape germplasm. M. rotundifolia and V. aesitvalis progeny show a gradation in response from resistant to susceptible, suggesting multiple resistance genes. V. romanetii progeny segregate into the distinct classes of highly resistant o...

  11. Developmnent of molecular markers for powdery mildew resistance in grapevines

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Molecular markers have been used as a tool for plant breeding for two decades. Markers linked to disease resistance can be used to pyramid resistance genes for improved durability and to improve the efficiency of evaluation. To develop molecular markers linked to resistance to grape powdery mildew ...

  12. New Sources of Resistance to Cucurbit Powdery Mildew in Melon

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Many physiological races of the cucurbit powdery mildew pathogen (CPM) Podosphaera xanthii (Castagne) Braun & Shishkoff have been reported on melon (Cucumis melo L.). Melon accession PI 313970 is the only reported source of host plant resistance to race S, which first appeared in Imperial Valley, CA...

  13. Hop powdery mildew control through alteration of spring pruning practices

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Since 1997, Podosphaera macularis, the causal agent of hop powdery mildew, has become a recurrent threat to hops in the Pacific Northwest because of the potential to reduce cone yield and quality. Disease management practices often involve preventative fungicide applications, but alternative approac...

  14. QTL mapping of resistance to Powdery Mildew in lettuce.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Erysiphe cichoracearam causes powdery mildew on most compositae including lettuce and chicory. Variation in susceptibility has been documented both in cultivated lettuce and wild relatives. Little is known about the genetic architecture of resistance to the pathogen, but monogenic resistance has bee...

  15. Erysiphe trifolii causing powdery mildew of lentil (Lens culinaris)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The taxonomy of the powdery mildew fungus infecting lentil in the Pacific Northwest (PNW) of the USA was investigated on the basis of morphology and rDNA ITS sequences. Anamorphic characters were in close agreement with descriptions of E. trifolii. However, teleomorphs formed chasmothecial appenda...

  16. Evaluation of Powdery Mildew Resistance in Hydrangea macrophylla

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew, Erysiphe polygoni DC, can be a significant problem on Hydrangea macrophylla (Thunb.) Ser. in the landscape during late summer to fall and during greenhouse propagation or production of potted plants. Because very little information related to sources of resistance is available, 90 H...

  17. Powdery mildew suppresses herbivore-induced plant volatiles and interferes with parasitoid attraction in Brassica rapa.

    PubMed

    Desurmont, Gaylord A; Xu, Hao; Turlings, Ted C J

    2016-09-01

    The co-occurrence of different antagonists on a plant can greatly affect infochemicals with ecological consequences for higher trophic levels. Here we investigated how the presence of a plant pathogen, the powdery mildew Erysiphe cruciferarum, on Brassica rapa affects (1) plant volatiles emitted in response to damage by a specialist herbivore, Pieris brassicae; (2) the attraction of the parasitic wasp Cotesia glomerata and (3) the performance of P. brassicae and C. glomerata. Plant volatiles were significantly induced by herbivory in both healthy and mildew-infected plants, but were quantitatively 41% lower for mildew-infected plants compared to healthy plants. Parasitoids strongly preferred Pieris-infested plants to dually-infested (Pieris + mildew) plants, and preferred dually infested plants over only mildew-infected plants. The performance of P. brassicae was unaffected by powdery mildew, but C. glomerata cocoon mass was reduced when parasitized caterpillars developed on mildew-infected plants. Thus, avoidance of mildew-infested plants may be adaptive for C. glomerata parasitoids, whereas P. brassicae caterpillars may suffer less parasitism on mildew-infected plants in nature. From a pest management standpoint, the concurrent presence of multiple plant antagonists can affect the efficiency of specific natural enemies, which may in turn have a negative impact on the regulation of pest populations. PMID:27043839

  18. Negative regulation of RIG-I-mediated antiviral signaling by TRK-fused gene (TFG) protein

    SciTech Connect

    Lee, Na-Rae; Shin, Han-Bo; Kim, Hye-In; Choi, Myung-Soo; Inn, Kyung-Soo

    2013-07-19

    Highlights: •TRK-fused gene product (TFG) interacts with TRIM25 upon viral infection. •TFG negatively regulates RIG-I mediated antiviral signaling. •TFG depletion leads to enhanced viral replication. •TFG act downstream of MAVS. -- Abstract: RIG-I (retinoic acid inducible gene I)-mediated antiviral signaling serves as the first line of defense against viral infection. Upon detection of viral RNA, RIG-I undergoes TRIM25 (tripartite motif protein 25)-mediated K63-linked ubiquitination, leading to type I interferon (IFN) production. In this study, we demonstrate that TRK-fused gene (TFG) protein, previously identified as a TRIM25-interacting protein, binds TRIM25 upon virus infection and negatively regulates RIG-I-mediated type-I IFN signaling. RIG-I-mediated IFN production and nuclear factor (NF)-κB signaling pathways were upregulated by the suppression of TFG expression. Furthermore, vesicular stomatitis virus (VSV) replication was significantly inhibited by small inhibitory hairpin RNA (shRNA)-mediated knockdown of TFG, supporting the suppressive role of TFG in RIG-I-mediated antiviral signaling. Interestingly, suppression of TFG expression increased not only RIG-I-mediated signaling but also MAVS (mitochondrial antiviral signaling protein)-induced signaling, suggesting that TFG plays a pivotal role in negative regulation of RNA-sensing, RIG-I-like receptor (RLR) family signaling pathways.

  19. PMR6, a pectate lyase-like gene required for powdery mildew susceptibility in Arabidopsis.

    PubMed

    Vogel, John P; Raab, Theodore K; Schiff, Celine; Somerville, Shauna C

    2002-09-01

    The plant genes required for the growth and reproduction of plant pathogens are largely unknown. In an effort to identify these genes, we isolated Arabidopsis mutants that do not support the normal growth of the powdery mildew pathogen Erysiphe cichoracearum. Here, we report on the cloning and characterization of one of these genes, PMR6. PMR6 encodes a pectate lyase-like protein with a novel C-terminal domain. Consistent with its predicted gene function, mutations in PMR6 alter the composition of the plant cell wall, as shown by Fourier transform infrared spectroscopy. pmr6-mediated resistance requires neither salicylic acid nor the ability to perceive jasmonic acid or ethylene, indicating that the resistance mechanism does not require the activation of well-described defense pathways. Thus, pmr6 resistance represents a novel form of disease resistance based on the loss of a gene required during a compatible interaction rather than the activation of known host defense pathways. PMID:12215508

  20. How Do Motivational Regulation Strategies Affect Achievement: Mediated by Effort Management and Moderated by Intelligence

    ERIC Educational Resources Information Center

    Schwinger, Malte; Steinmayr, Ricarda; Spinath, Birgit

    2009-01-01

    It was assumed that the effect of motivational regulation strategies on achievement is mediated by effort management and moderated by intelligence. A sample of 231 11th and 12th grade German high-school students provided self-reports on their use of motivational regulation strategies and effort management and completed an intelligence test.…

  1. Emotion Regulation and Aggressive Behavior in Preschoolers: The Mediating Role of Social Information Processing

    ERIC Educational Resources Information Center

    Helmsen, Johanna; Koglin, Ute; Petermann, Franz

    2012-01-01

    This study examined whether the relation between maladaptive emotion regulation and aggression was mediated by deviant social information processing (SIP). Participants were 193 preschool children. Emotion regulation and aggression were rated by teachers. Deviant SIP (i.e., attribution of hostile intent, aggressive response generation, aggressive…

  2. CERAMIDE SYNTHASE 1 IS REGULATED BY PROTEASOMAL MEDIATED TURNOVER

    PubMed Central

    Sridevi, Priya; Alexander, Hannah; Laviad, Elad L.; Pewzner-Jung, Yael; Hannink, Mark; Futerman, Anthony H.; Alexander, Stephen

    2009-01-01

    Ceramide is an important bioactive lipid, intimately involved in many cellular functions, including the regulation of cell death, and in cancer and chemotherapy. Ceramide is synthesized de novo from sphinganine and acyl CoA via a family of 6 ceramide synthase enzymes, each having a unique preference for different fatty acyl CoA substrates and a unique tissue distribution. However, little is known regarding the regulation of these important enzymes. In this study we focus on ceramide synthase 1 (CerS1) which is the most structurally and functionally distinct of the enzymes, and describe a regulatory mechanism that specifically controls the level of CerS1 via ubiquitination and proteasome dependent protein turnover. We show that both endogenous and ectopically expressed CerS1 have rapid basal turnover and that diverse stresses including chemotherapeutic drugs, UV light and DTT can induce CerS1 turnover. The turnover requires CerS1 activity and is regulated by the opposing actions of p38 MAP kinase and protein kinase C (PKC). p38 MAP kinase is a positive regulator of turnover, while PKC is a negative regulator of turnover. CerS1 is phosphorylated in vivo and activation of PKC increases the phosphorylation of the protein. This study reveals a novel and highly specific mechanism by which CerS1 protein levels are regulated and which directly impacts ceramide homeostasis. PMID:19393694

  3. Difficulties with emotion regulation mediate the relationship between borderline personality disorder symptom severity and interpersonal problems.

    PubMed

    Herr, Nathaniel R; Rosenthal, M Zachary; Geiger, Paul J; Erikson, Karen

    2013-08-01

    Problems with interpersonal functioning and difficulties with emotion regulation are core characteristics of borderline personality disorder (BPD). Little is known, however, about the interrelationship between these areas of dysfunction in accounting for BPD symptom severity. The present study examines a model of the relationship between difficulties with emotion regulation and interpersonal dysfunction in a community sample of adults (n = 124) with the full range of BPD symptoms. Results showed that difficulties with emotion regulation fully mediated the relationship between BPD symptom severity and interpersonal dysfunction. An alternative model indicated that interpersonal problems partially mediated the relationship between difficulties with emotion regulation and BPD symptom severity. These findings support existing theories of BPD, which propose that difficulties with emotion regulation may account for the types of interpersonal problems experienced by individuals with BPD and suggest further examination of the possibility that interpersonal dysfunction may worsen these individuals' difficulties with emotion regulation. PMID:24343962

  4. Ferroportin-mediated iron transport: expression and regulation

    PubMed Central

    Ward, Diane; Kaplan, Jerry

    2013-01-01

    The distinguishing feature between iron homeostasis in single versus multicellular organisms is the need for multicellular organisms to transfer iron from sites of absorption to sites of utilization and storage. Ferroportin is the only known iron exporter and ferroportin plays an essential role in the export of iron from cells to blood. Ferroportin can be regulated at many different levels including transcriptionally, post-transcriptionally, through mRNA stability and post-translationally, through protein turnover. Additionally, ferroportin may be regulated in both cell-dependent and cell-autonomous fashions. Regulation of ferroportin is critical for iron homeostasis as alterations in ferroportin may result in either iron deficiency or iron overload. PMID:22440327

  5. KAP1 regulates type I interferon/STAT1-mediated IRF-1 gene expression

    SciTech Connect

    Kamitani, Shinya; Ohbayashi, Norihiko; Ikeda, Osamu; Togi, Sumihito; Muromoto, Ryuta; Sekine, Yuichi; Ohta, Kazuhide; Ishiyama, Hironobu; Matsuda, Tadashi

    2008-05-30

    Signal transducers and activators of transcription (STATs) mediate cell proliferation, differentiation, and survival in immune responses, hematopoiesis, neurogenesis, and other biological processes. Recently, we showed that KAP1 is a novel STAT-binding partner that regulates STAT3-mediated transactivation. KAP1 is a universal co-repressor protein for the KRAB zinc finger protein superfamily of transcriptional repressors. In this study, we found KAP1-dependent repression of interferon (IFN)/STAT1-mediated signaling. We also demonstrated that endogenous KAP1 associates with endogenous STAT1 in vivo. Importantly, a small-interfering RNA-mediated reduction in KAP1 expression enhanced IFN-induced STAT1-dependent IRF-1 gene expression. These results indicate that KAP1 may act as an endogenous regulator of the IFN/STAT1 signaling pathway.

  6. Emotion regulation as mediator of treatment outcome in therapy for deliberate self-harm.

    PubMed

    Slee, Nadja; Spinhoven, Philip; Garnefski, Nadia; Arensman, Ella

    2008-01-01

    This study presents the outcomes of mediator analyses as part of a randomized controlled trial of Cognitive-Behavioural Therapy (CBT) for young people who engage in deliberate self-harm (DSH). The study involved 90 people, aged 15-35 years, who were randomly assigned to CBT in addition to treatment as usual or to treatment as usual only. The findings showed that changes in DSH were partially mediated by changes in emotion-regulation difficulties, particularly difficulties with impulse control and goal-directed behaviours. In addition, the potential mediating role of symptoms of depression, anxiety and suicidal cognitions was examined. Although the CBT intervention significantly reduced depression, anxiety and suicidal cognitions, these measures of symptom severity did not play a mediating role. These findings suggest that interventions for DSH should not primarily focus on mental disorders associated with DSH, but should be DSH-specific and should target specific emotion-regulation difficulties. PMID:19115441

  7. SUMO-mediated regulation of DNA damage repair and responses

    PubMed Central

    Sarangi, Prabha; Zhao, Xiaolan

    2015-01-01

    Sumoylation plays important roles during DNA damage repair and responses. Recent broad-scope and substrate-based studies have shed light on the regulation and significance of sumoylation during these processes. An emerging paradigm is that sumoylation of many DNA metabolism proteins is controlled by DNA engagement. Such “on-site modification” can explain low substrate modification levels and has important implications in sumoylation mechanisms and effects. New studies also suggest that sumoylation can regulate a process through an ensemble effect or via major substrates. Additionally, we describe new trends in the functional effects of sumoylation, such as bi-directional changes in biomolecule binding and multi-level coordination with other modifications. These emerging themes and models will stimulate our thinking and research in sumoylation and genome maintenance. PMID:25778614

  8. Light-Mediated Hormonal Regulation of Plant Growth and Development.

    PubMed

    de Wit, Mieke; Galvão, Vinicius Costa; Fankhauser, Christian

    2016-04-29

    Light is crucial for plant life, and perception of the light environment dictates plant growth, morphology, and developmental changes. Such adjustments in growth and development in response to light conditions are often established through changes in hormone levels and signaling. This review discusses examples of light-regulated processes throughout a plant's life cycle for which it is known how light signals lead to hormonal regulation. Light acts as an important developmental switch in germination, photomorphogenesis, and transition to flowering, and light cues are essential to ensure light capture through architectural changes during phototropism and the shade avoidance response. In describing well-established links between light perception and hormonal changes, we aim to give insight into the mechanisms that enable plants to thrive in variable light environments. PMID:26905653

  9. A novel phenylcyclohex-1-enecarbothioamide derivative inhibits CXCL8-mediated chemotaxis through selective regulation of CXCR2-mediated signalling

    PubMed Central

    Ha, Helen; Bensman, Tim; Ho, Henry; Beringer, Paul M; Neamati, Nouri

    2014-01-01

    Background and Purpose Since the CXC chemokine receptor CXCR2 and its cognate ligand CXCL8 (IL-8) critically regulate neutrophil trafficking during inflammation, they have been implicated in a number of inflammatory lung diseases. Several CXCR2 antagonists have been described and the blockade of CXCR2 has shown promise in pre-clinical disease models and early clinical trials. However, given its potential, there are fewer distinct classes of antagonists of CXCR2 than of other clinically relevant molecular targets. Thus, we sought to identify additional classes of compounds that alter CXCR2 function. Experimental Approach We used the CXCR2 Tango™ assay to screen an in-house library of highly diverse chemical compounds. CX4338 [2-(benzylamino)-4,4-dimethyl-6-oxo-N-phenylcyclohex-1-enecarbothioamide] was identified from our screen and additional studies to characterize the compound were performed. Receptor internalization and second-messenger assays were used to assess the effects of CX4338 on CXCR2-mediated signalling. Wound healing, transwell cell migration and LPS-induced lung inflammation in mice were used to determine the in vitro and in vivo effects of CX4338. Key Results CX4338 selectively inhibited CXCR2-mediated recruitment of β-arrestin-2 and receptor internalization, while enhancing CXCR2-mediated MAPK activation. Additionally, CX4338 inhibited CXCL8-induced chemotaxis in CXCR2-overexpressing cells and human neutrophils. In vivo, CX4338 significantly reduced neutrophils in bronchoalveolar lavage induced by LPS in mice. Conclusions and Implications A novel compound CX4338 inhibited CXCR2-mediated cell migration with a mechanism of action not previously reported. Also, selective inhibition of CXCR2-mediated β-arrestin-2 activation is sufficient to inhibit CXCL8-mediated chemotaxis. PMID:24354854

  10. Simian virus 40 T antigen can regulate p53-mediated transcription independent of binding p53.

    PubMed Central

    Rushton, J J; Jiang, D; Srinivasan, A; Pipas, J M; Robbins, P D

    1997-01-01

    A simian virus 40 (SV40) T-antigen mutant containing only the N-terminal 136 amino acids, able to bind to Rb and p300 but not p53, partially inhibited p53-mediated transcription without affecting the ability of p53 to bind DNA. These results suggest that SV40 T antigen can regulate p53-mediated transcription either directly through protein-protein association or indirectly through interaction with factors which may function to confer p53-mediated transcription. PMID:9188637

  11. Rationalization of pesticide treatments against powdery mildew of grape.

    PubMed

    Spera, G; La Torre, A; Gianferro, M; Bugliosi, R

    2007-01-01

    The powdery mildew represents one of the diseases which affect the grape, it is diffused in all agricultural regions with variable intensity and epidemic course in operation of many microclimatic factors. The powdery mildew of grape is caused from Uncinala necator (Schw.) Burr. (nowadays named Erysiphe necator Schwein.); it is controlled with systemic therapy and contact chemicaL products. In some vineyards located in Latium (central Italy), different field trials have been carried out purposely to rationalize the treatments against E. necator. We have studied the powdery mildew infections through monitoring a set of environmental parameters, the evaluation of cultivar sensibility, the agricultural production method and the area characteristics. We have analysed the following environmental parameters monitoring every 15 minutes: precipitation, soil temperature, solar radiation, wind direction, wind speed, atmospheric relative humidity, atmospheric temperature, leaf wetness, soil humidity to cm 20 and soil humidity to cm 40. Besides, we have used Artificial Intelligence analysis techniques to try to forecast U. necator infections. Guideline EPPO/OEPP PP 1/4 (4) has been used. The trials were conducted in conventional and organic farms. In 2 conventional farms and in organic farm we have considered 1 untreated control thesis, in order to follow the course of infection, 1 standard farm reference thesis (standard), where the treatments were carried out according to the usual farm procedures and 1 thesis where the treatments were carried out according to examining the environmental data. In another conventional vineyard, we have considered only 1 untreated control thesis and 1 standard farm reference thesis (standard) to study disease trend. The achieved results have underlined the possibility (through the knowledge of data pedoclimatic and cultural) to position the treatments against the powdery mildew so that to reduce their number. The lower number of treatments that

  12. Adolescent Depression and Negative Life Events, the Mediating Role of Cognitive Emotion Regulation

    PubMed Central

    Stikkelbroek, Yvonne; Bodden, Denise H. M.; Kleinjan, Marloes; Reijnders, Mirjam; van Baar, Anneloes L.

    2016-01-01

    Background Depression during adolescence is a serious mental health problem. Difficulties in regulating evoked emotions after stressful life events are considered to lead to depression. This study examined if depressive symptoms were mediated by various cognitive emotion regulation strategies after stressful life events, more specifically, the loss of a loved one, health threats or relational challenges. Methods We used a sample of 398 adolescents (Mage = 16.94, SD = 2.90), including 52 depressed outpatients, who all reported stressful life event(s). Path analyses in Mplus were used to test mediation, for the whole sample as well as separately for participants scoring high versus low on depression, using multigroup analyses. Results Health threats and relational challenging stressful life events were associated with depressive symptoms, while loss was not. More frequent use of maladaptive strategies was related to more depressive symptoms. More frequent use of adaptive strategies was related to less depressive symptoms. Specific life events were associated with specific emotion regulation strategies. The relationship between challenging, stressful life events and depressive symptoms in the whole group was mediated by maladaptive strategies (self-blame, catastrophizing and rumination). No mediation effect was found for adaptive strategies. Conclusion The association between relational challenging, stressful life events and depressive symptoms was mediated by maladaptive, cognitive emotion regulation strategies. PMID:27571274

  13. ERK2 Mediates Metabolic Stress Response to Regulate Cell Fate.

    PubMed

    Shin, Sejeong; Buel, Gwen R; Wolgamott, Laura; Plas, David R; Asara, John M; Blenis, John; Yoon, Sang-Oh

    2015-08-01

    Insufficient nutrients disrupt physiological homeostasis, resulting in diseases and even death. Considering the physiological and pathological consequences of this metabolic stress, the adaptive responses that cells utilize under this condition are of great interest. We show that under low-glucose conditions, cells initiate adaptation followed by apoptosis responses using PERK/Akt and MEK1/ERK2 signaling, respectively. For adaptation, cells engage the ER stress-induced unfolded protein response, which results in PERK/Akt activation and cell survival. Sustained and extreme energetic stress promotes a switch to isoform-specific MEK1/ERK2 signaling, induction of GCN2/eIF2α phosphorylation, and ATF4 expression, which overrides PERK/Akt-mediated adaptation and induces apoptosis through ATF4-dependent expression of pro-apoptotic factors including Bid and Trb3. ERK2 activation during metabolic stress contributes to changes in TCA cycle and amino acid metabolism, and cell death, which is suppressed by glutamate and α-ketoglutarate supplementation. Taken together, our results reveal promising targets to protect cells or tissues from metabolic stress. PMID:26190261

  14. Chirality-mediated polypeptide micelles for regulated drug delivery.

    PubMed

    Ding, Jianxun; Li, Chen; Zhang, Ying; Xu, Weiguo; Wang, Jincheng; Chen, Xuesi

    2015-01-01

    Two kinds of triblock poly(ethylene glycol)-polyleucine (PEG-PLeu) copolymers were synthesized through the ring-opening polymerization of L-Leu N-carboxyanhydride (NCA), or equivalent D-Leu NCA and L-Leu NCA with amino-terminated PEG as a macroinitiator. The amphiphilic copolymers spontaneously self-assembled into spherical micellar aggregations in an aqueous environment. The micelle with a racemic polypeptide core exhibited smaller critical micelle concentration and diameter compared to those with a levorotatory polypeptide core. A model anthracycline antineoplastic agent, i.e., doxorubicin (DOX), was loaded into micelles through nanoprecipitation, and the PEG-P(D,L-Leu) micelle exhibited higher drug-loading efficacy than that with a P(L-Leu) core-this difference was attributed to the flexible and compact P(L-Leu) core. Sustained in vitro DOX release from micelles with both levorotatory and racemic polypeptide cores was observed, and the DOX-loaded PEG-P(D,L-Leu) micelle exhibited a slower release rate. More interestingly, DOX-loaded micelles exhibited chirality-mediated antitumor efficacy in vitro and in vivo, which are all better than that of free DOX. Furthermore, both enhanced tumor inhibition and excellent security in vivo were confirmed by histopathological or in situ cell apoptosis analyses. Therefore, DOX-loaded PEG-PLeu micelles appear to be an interesting nanoscale polymeric formulation for promising malignancy chemotherapy. PMID:25278445

  15. GSK3 is a regulator of RAR-mediated differentiation

    PubMed Central

    Gupta, K; Gulen, F; Sun, L; Aguilera, R; Chakrabarti, A; Kiselar, J; Agarwal, MK; Wald, DN

    2015-01-01

    Acute myeloid leukemia (AML) is the most common form of leukemia in adults. Unfortunately, the standard therapeutic agents used for this disease have high toxicities and poor efficacy. The one exception to these poor outcomes is the use of the retinoid, all-trans retinoic acid (ATRA), for a rare subtype of AML (APL). The use of the differentiation agent, ATRA, in combination with low-dose chemotherapy leads to the long-term survival and presumed cure of 75–85% of patients. Unfortunately ATRA has not been clinically useful for other subtypes of AML. Though many non-APL leukemic cells respond to ATRA, they require significantly higher concentrations of ATRA for effective differentiation. Here we show that the combination of ATRA with glycogen synthase kinase 3 (GSK3) inhibition significantly enhances ATRA-mediated AML differentiation and growth inhibition. These studies have revealed that ATRA's receptor, the retinoic acid receptor (RAR), is a novel target of GSK3 phosphorylation and that GSK3 can impact the expression and transcriptional activity of the RAR. Overall, our studies suggest the clinical potential of ATRA and GSK3 inhibition for AML and provide a mechanistic framework to explain the promising activity of this combination regimen. PMID:22222598

  16. GSK3 is a regulator of RAR-mediated differentiation.

    PubMed

    Gupta, K; Gulen, F; Sun, L; Aguilera, R; Chakrabarti, A; Kiselar, J; Agarwal, M K; Wald, D N

    2012-06-01

    Acute myeloid leukemia (AML) is the most common form of leukemia in adults. Unfortunately, the standard therapeutic agents used for this disease have high toxicities and poor efficacy. The one exception to these poor outcomes is the use of the retinoid, all-trans retinoic acid (ATRA), for a rare subtype of AML (APL). The use of the differentiation agent, ATRA, in combination with low-dose chemotherapy leads to the long-term survival and presumed cure of 75-85% of patients. Unfortunately ATRA has not been clinically useful for other subtypes of AML. Though many non-APL leukemic cells respond to ATRA, they require significantly higher concentrations of ATRA for effective differentiation. Here we show that the combination of ATRA with glycogen synthase kinase 3 (GSK3) inhibition significantly enhances ATRA-mediated AML differentiation and growth inhibition. These studies have revealed that ATRA's receptor, the retinoic acid receptor (RAR), is a novel target of GSK3 phosphorylation and that GSK3 can impact the expression and transcriptional activity of the RAR. Overall, our studies suggest the clinical potential of ATRA and GSK3 inhibition for AML and provide a mechanistic framework to explain the promising activity of this combination regimen. PMID:22222598

  17. TLR signals posttranscriptionally regulate the cytokine trafficking mediator sortilin

    PubMed Central

    Yabe-Wada, Toshiki; Matsuba, Shintaro; Takeda, Kazuya; Sato, Tetsuya; Suyama, Mikita; Ohkawa, Yasuyuki; Takai, Toshiyuki; Shi, Haifeng; Philpott, Caroline C.; Nakamura, Akira

    2016-01-01

    Regulating the transcription, translation and secretion of cytokines is crucial for controlling the appropriate balance of inflammation. Here we report that the sorting receptor sortilin plays a key role in cytokine production. We observed interactions of sortilin with multiple cytokines including IFN-α, and sortilin depletion in plasmacytoid dendritic cells (pDCs) led to a reduction of IFN-α secretion, suggesting a pivotal role of sortilin in the exocytic trafficking of IFN-α in pDCs. Moreover, sortilin mRNA was degraded posttranscriptionally upon stimulation with various TLR ligands. Poly-rC-binding protein 1 (PCBP1) recognized the C-rich element (CRE) in the 3′ UTR of sortilin mRNA, and depletion of PCBP1 enhanced the degradation of sortilin transcripts, suggesting that PCBP1 can act as a trans-acting factor to stabilize sortilin transcripts. The nucleotide-binding ability of PCBP1 was impaired by zinc ions and alterations of intracellular zinc affect sortilin expression. PCBP1 may therefore control the stability of sortilin transcripts by sensing intracellular zinc levels. Collectively, our findings provide insights into the posttranslational regulation of cytokine production through the posttranscriptional control of sortilin expression by TLR signals. PMID:27220277

  18. Negative regulation of Vps34 by Cdk mediated phosphorylation

    PubMed Central

    Furuya, Tsuyoshi; Kim, Minsu; Lipinski, Marta; Li, Juying; Kim, Dohoon; Lu, Tao; Shen, Yong; Rameh, Lucia; Yankner, Bruce; Tsai, Li-Huei; Yuan, Junying

    2010-01-01

    Summary Vps34 (vacuolar protein sorting 34) complexes, the class III PtdIns3 kinase, specifically phosphorylate the D3-position of PtdIns to produce PtdIns3P. Vps34 is involved in the control of multiple key intracellular membrane trafficking pathways including endocytic sorting and autophagy. In mammalian cells, Vps34 interacts with Beclin 1, an orthologue of Atg6 in yeast, to regulate the production of PtdIns3P and autophagy. We show that Vps34 is phosphorylated on Thr159 by Cdk1, which negatively regulates its interaction with Beclin1 during mitosis. Cdk5/p25, a neuronal cdk shown to play a role in Alzheimer’s disease, can also phosphorylate Thr159 of Vps34. Phosphorylation of Vps34 on Thr159 inhibits its interaction with Beclin 1. We propose that phosphorylation of Thr159 in Vps34 is a key regulatory mechanism that controls the class III PtdIns3 kinase activity in cell cycle progression, development and human diseases including neurodegeneration and cancers. PMID:20513426

  19. TLR signals posttranscriptionally regulate the cytokine trafficking mediator sortilin.

    PubMed

    Yabe-Wada, Toshiki; Matsuba, Shintaro; Takeda, Kazuya; Sato, Tetsuya; Suyama, Mikita; Ohkawa, Yasuyuki; Takai, Toshiyuki; Shi, Haifeng; Philpott, Caroline C; Nakamura, Akira

    2016-01-01

    Regulating the transcription, translation and secretion of cytokines is crucial for controlling the appropriate balance of inflammation. Here we report that the sorting receptor sortilin plays a key role in cytokine production. We observed interactions of sortilin with multiple cytokines including IFN-α, and sortilin depletion in plasmacytoid dendritic cells (pDCs) led to a reduction of IFN-α secretion, suggesting a pivotal role of sortilin in the exocytic trafficking of IFN-α in pDCs. Moreover, sortilin mRNA was degraded posttranscriptionally upon stimulation with various TLR ligands. Poly-rC-binding protein 1 (PCBP1) recognized the C-rich element (CRE) in the 3' UTR of sortilin mRNA, and depletion of PCBP1 enhanced the degradation of sortilin transcripts, suggesting that PCBP1 can act as a trans-acting factor to stabilize sortilin transcripts. The nucleotide-binding ability of PCBP1 was impaired by zinc ions and alterations of intracellular zinc affect sortilin expression. PCBP1 may therefore control the stability of sortilin transcripts by sensing intracellular zinc levels. Collectively, our findings provide insights into the posttranslational regulation of cytokine production through the posttranscriptional control of sortilin expression by TLR signals. PMID:27220277

  20. Distinct Brain Systems Mediate the Effects of Nociceptive Input and Self-Regulation on Pain

    PubMed Central

    Woo, Choong-Wan; Roy, Mathieu; Buhle, Jason T.; Wager, Tor D.

    2015-01-01

    Cognitive self-regulation can strongly modulate pain and emotion. However, it is unclear whether self-regulation primarily influences primary nociceptive and affective processes or evaluative ones. In this study, participants engaged in self-regulation to increase or decrease pain while experiencing multiple levels of painful heat during functional magnetic resonance imaging (fMRI) imaging. Both heat intensity and self-regulation strongly influenced reported pain, but they did so via two distinct brain pathways. The effects of stimulus intensity were mediated by the neurologic pain signature (NPS), an a priori distributed brain network shown to predict physical pain with over 90% sensitivity and specificity across four studies. Self-regulation did not influence NPS responses; instead, its effects were mediated through functional connections between the nucleus accumbens and ventromedial prefrontal cortex. This pathway was unresponsive to noxious input, and has been broadly implicated in valuation, emotional appraisal, and functional outcomes in pain and other types of affective processes. These findings provide evidence that pain reports are associated with two dissociable functional systems: nociceptive/affective aspects mediated by the NPS, and evaluative/functional aspects mediated by a fronto-striatal system. PMID:25562688

  1. Dexamethasone-mediated transcriptional regulation of rat carboxylesterase 2 gene.

    PubMed

    Hori, Takeshi; Jin, Liangjing; Fujii, Ayako; Furihata, Tomomi; Nagahara, Yuko; Chiba, Kan; Hosokawa, Masakiyo

    2012-07-01

    Rat carboxylesterase 2 (rCES2), which was previously identified as a methylprednisolone 21-hemisuccinate hydrolase, is highly inducible by dexamethasone in the liver. In the present study, we investigated the molecular mechanisms by which this induction occurs. Injection of dexamethasone (1 mg/kg weight) into rats resulted in increases in the expression of rCES2 mRNA in a time-dependent manner with a peak at 12 h after injection. In primary rat hepatocytes, the expression level of rCES2 mRNA was increased by treatment with 100 nM dexamethasone, and the increase was completely blocked in the presence of 10 µM mifepristone (RU-486), a potent inhibitor of glucocorticoid receptor (GR), or 10 µg/mL cycloheximide, a translation inhibitor. Luciferase assays revealed that 100 nM dexamethasone increased rCES2 promoter activities, although the effect of dexamethasone on the promoter activity was smaller than that on rCES2 mRNA expression. The increased activities were completely inhibited by treatment of the hepatocytes with 10 µM RU-486. Based on these results, it is concluded that dexamethasone enhances transcription of the rCES2 gene via GR in the rat liver and that the dexamethasone-mediated induction of rCES2 mRNA may be dependent on de novo protein synthesis. Our results provide clues to understanding what compounds induce rCES2. PMID:22235919

  2. Tespa1 negatively regulates FcεRI-mediated signaling and the mast cell–mediated allergic response

    PubMed Central

    Zheng, Mingzhu; Qiu, Yuanjun; Guo, Chuansheng; Ji, Jian; Lei, Lei; Zhang, Xue; Liang, Jingjing; Lou, Jun; Huang, Wei; Dong, Bowen; Wu, Songquan; Wang, Jianli; Ke, Yuehai; Cao, Xuetao; Zhou, Yi Ting

    2014-01-01

    Antigen-mediated cross-linking of IgE on mast cells triggers a signaling cascade that results in their degranulation and proinflammatory cytokine production, which are key effectors in allergic reactions. We show that the activation of mast cells is negatively regulated by the newly identified adaptor protein Tespa1. Loss of Tespa1 in mouse mast cells led to hyper-responsiveness to stimulation via FcεRI. Mice lacking Tespa1 also displayed increased sensitivity to IgE-mediated allergic responses. The dysregulated signaling in KO mast cells was associated with increased activation of Grb2-PLC-γ1-SLP-76 signaling within the LAT1 (linker for activation of T cells family, member 1) signalosome versus the LAT2 signalosome. Collectively, these findings show that Tespa1 orchestrates mast cell activation by tuning the balance of LAT1 and LAT2 signalosome assembly. PMID:25422497

  3. Clinical Neuroanatomy and Neurotransmitter-Mediated Regulation of Penile Erection

    PubMed Central

    Jo, Hyun Woo; Kwon, Hyunseob

    2014-01-01

    Erectile dysfunction (ED) has an adverse impact on men's quality of life. Penile erection, which is regulated by nerves that are innervated into the erectile tissue, can be affected by functional or anatomical trauma of the perineal region, including specific structures of the penis, causing ED. Penile erection is neurologically controlled by the autonomic nervous system. Therefore, it is of utmost importance to understand the neurogenic structure of the erectile tissue and the types of neurotransmitters involved in the penile erection process. Here, we highlight the basic clinical anatomy and erectile function of the penis. Understanding the clinical connotation of the relationship between penile erectile structure and function may provide fresh insights for identifying the main mechanisms involved in ED and help develop surgical techniques for the treatment of ED. PMID:24987557

  4. slo K+ channel gene regulation mediates rapid drug tolerance

    NASA Astrophysics Data System (ADS)

    Ghezzi, Alfredo; Al-Hasan, Yazan M.; Larios, Leo E.; Bohm, Rudolf A.; Atkinson, Nigel S.

    2004-12-01

    Changes in neural activity caused by exposure to drugs may trigger homeostatic mechanisms that attempt to restore normal neural excitability. In Drosophila, a single sedation with the anesthetic benzyl alcohol changes the expression of the slo K+ channel gene and induces rapid drug tolerance. We demonstrate linkage between these two phenomena by using a mutation and a transgene. A mutation that eliminates slo expression prevents tolerance, whereas expression from an inducible slo transgene mimics tolerance in naïve animals. The behavioral response to benzyl alcohol can be separated into an initial phase of hyperkinesis and a subsequent phase of sedation. The hyperkinetic phase causes a drop in slo gene expression and makes animals more sensitive to benzyl alcohol. It is the sedative phase that stimulates slo gene expression and induces tolerance. We demonstrate that the expression level of slo is a predictor of drug sensitivity. drug abuse | potassium channel | transcription regulation

  5. Complement-Mediated Regulation of Metabolism and Basic Cellular Processes.

    PubMed

    Hess, Christoph; Kemper, Claudia

    2016-08-16

    Complement is well appreciated as a critical arm of innate immunity. It is required for the removal of invading pathogens and works by directly destroying them through the activation of innate and adaptive immune cells. However, complement activation and function is not confined to the extracellular space but also occurs within cells. Recent work indicates that complement activation regulates key metabolic pathways and thus can impact fundamental cellular processes, such as survival, proliferation, and autophagy. Newly identified functions of complement include a key role in shaping metabolic reprogramming, which underlies T cell effector differentiation, and a role as a nexus for interactions with other effector systems, in particular the inflammasome and Notch transcription-factor networks. This review focuses on the contributions of complement to basic processes of the cell, in particular the integration of complement with cellular metabolism and the potential implications in infection and other disease settings. PMID:27533012

  6. Mediators and molecular pathways involved in the regulation of neutrophil extracellular trap formation mediated by activated platelets.

    PubMed

    Carestia, Agostina; Kaufman, Tomás; Rivadeneyra, Leonardo; Landoni, Verónica Inés; Pozner, Roberto Gabriel; Negrotto, Soledad; D'Atri, Lina Paola; Gómez, Ricardo Martín; Schattner, Mirta

    2016-01-01

    In addition to being key elements in hemostasis and thrombosis, platelets amplify neutrophil function. We aimed to gain further insight into the stimuli, mediators, molecular pathways, and regulation of neutrophil extracellular trap formation mediated by human platelets. Platelets stimulated by lipopolysaccharide, a wall component of gram-negative bacteria, Pam3-cysteine-serine-lysine 4, a mimetic of lipopeptide from gram-positive bacteria, Escherichia coli, Staphylococcus aureus, or physiologic platelet agonists promoting neutrophil extracellular trap formation and myeloperoxidase-associated DNA activity under static and flow conditions. Although P-selectin or glycoprotein IIb/IIIa were not involved, platelet glycoprotein Ib, neutrophil cluster of differentiation 18, and the release of von Willebrand factor and platelet factor 4 seemed to be critical for the formation of neutrophil extracellular traps. The secretion of these molecules depended on thromboxane A(2) production triggered by lipopolysaccharide or Pam3-cysteine-serine-lysine 4 but not on high concentrations of thrombin. Accordingly, aspirin selectively inhibited platelet-mediated neutrophil extracellular trap generation. Signaling through extracellular signal-regulated kinase, phosphatidylinositol 3-kinase, and Src kinases, but not p38 or reduced nicotinamide adenine dinucleotide phosphate oxidase, was involved in platelet-triggered neutrophil extracellular trap release. Platelet-mediated neutrophil extracellular trap formation was inhibited by prostacyclin. Our results support a role for stimulated platelets in promoting neutrophil extracellular trap formation, reveal that an endothelium-derived molecule contributes to limiting neutrophil extracellular trap formation, and highlight platelet inhibition as a potential target for controlling neutrophil extracellular trap cell death. PMID:26320263

  7. Silencing Mediator of Retinoid and Thyroid Hormone Receptors (SMRT) regulates glucocorticoid action in adipocytes

    PubMed Central

    Emont, Margo P.; Mantis, Stelios; Kahn, Jonathan H.; Landeche, Michael; Han, Xuan; Sargis, Robert M

    2015-01-01

    Local modulation of glucocorticoid action in adipocytes regulates adiposity and systemic insulin sensitivity. However, the specific cofactors that mediate glucocorticoid receptor (GR) action in adipocytes remain unclear. Here we show that the silencing mediator of retinoid and thyroid hormone receptors (SMRT) is recruited to GR in adipocytes and regulates ligand-dependent GR function. Decreased SMRT expression in adipocytes in vivo increases expression of glucocorticoid-responsive genes. Moreover, adipocytes with decreased SMRT expression exhibit altered glucocorticoid regulation of lipolysis. We conclude that SMRT regulates the metabolic functions of GR in adipocytes in vivo. Modulation of GR-SMRT interactions in adipocytes represents a novel approach to control the local degree of glucocorticoid action and thus influence adipocyte metabolic function. PMID:25766503

  8. First report of powdery mildew of chickpea (Cicer arietinum) caused by Leveillula taurica in Washington State

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Chickpea is an important grain legume worldwide, and an important rotational and specialty crop in the US. In October 2007, powdery mildew was found on chickpea plants in a field near Pullman, Washington. Typical powdery mildew symptoms and signs were observed on petioles and adaxial leaf surfaces....

  9. Genetic characterization and targeted mapping of a Triticum timopheevii-dervied powdery mildew resistance gene

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There are five alleles conferring race-specific resistance to powdery mildew (caused by Blumeria graminis f. sp. tritici) at the Pm1 locus on the long arm of chromosome 7A of wheat (Triticum aestivum. L). A dominant powdery mildew resistance gene transferred to the hexaploid germplasm line NC99BGTAG...

  10. DELAY OF EXPRESSION OF POWDERY MILDEW ON ZINNIA GROWN HYDROPONICALLY IN HOAGLAND'S SOLUTION FORTIFIED WITH SILICON

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew, caused by the fungus Erysiphe cichoracearum, is one of the most common foliar diseases that occur in greenhouse bedding plant production. Although powdery mildews are somewhat host specific, E. cichoracearum is reported to have a wide host range which includes the commonly grown be...

  11. Pm37, a new broadly effective powdery mildew resistance gene from Triticum timopheevii

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew, caused by Blumeria graminis f. sp. tritici, is an important foliar disease in wheat (Triticum aestivum L.), especially in areas with a cool or maritime climate. A dominant powdery mildew resistance gene transferred to the hexaploid germplasm line NC99BGTAG11 (NCAG11) from T. timophee...

  12. QTL mapping of powdery mildew resistance in WI 2757 cucumber (Cucumis sativus L)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew is a serious fungal disease of cucumber and other cucurbot crops in the US and many other parts of the world. Resistant cultivars have been deployed in production for a long time, but the genetic mechanisms of powdery mildew resistance in cucumber are not well understood. In a three-y...

  13. Crop damage caused by Powdery Mildew on Hop and its relationship to late season management

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew of hop (Podosphaera macularis) may cause economic loss due to reductions in cone yield and quality. Quantitative estimates of crop damage from powdery mildew remain poorly characterised, especially the effect of late season disease management on crop yield and quality. Field studies i...

  14. Virulence structure of the eastern U.S. wheat powdery mildew population

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Little is known about the population structure of wheat powdery mildew in the eastern 2 U.S., and the most recent report on virulence in this pathogen population involved isolates 3 collected in 1993-94. In the present study, wheat leaves naturally infected with powdery mildew 4 were collected from ...

  15. Powdery mildew caused by Podosphaera macularis on hop (Humulus lupulus) in North Carolina

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In June 2015, a grower in western North Carolina detected powdery mildew in a small hop yard. Characteristic colonies of the pathogen where observed on cultivars Cashmere, Cascade, and Chinook. Leaves with powdery mildew were collected from cultivar Cashmere for confirmation of the pathogen identi...

  16. Genetic Mapping of a Triticum monococcum-derived Powdery Mildew Resistance Gene in Common Wheat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew of wheat (Triticum aestivum L.) is a major fungal disease caused by Blumeria graminis DC f sp. tritici. A microsatellite linkage map was developed for the T. monococcum-derived powdery mildew resistant gene present in the North Carolina germplasm line NCBGT96A6 (NCA6). Genetic analys...

  17. Epidemiology of Powdery Mildew on Resistant and Susceptible Flowering Dogwood Cultivars

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew, caused by Erysiphe pulchra (syn. Microsphaera pulchra) is an important disease on flowering dogwood (Cornus florida) in the Eastern United States. Temporal progress of powdery mildew on flowering dogwood cultivars with different levels of resistance was investigated in the field in 2...

  18. Sequential sampling for estimation and classification of the incidence of hop powdery mildew I: Leaf sampling

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Hop powdery mildew (caused by Podosphaera macularis) is an important disease of hops (Humulus lupulus) in the Pacific Northwest. Sequential sampling models for estimation and classification of the incidence of powdery mildew on leaves of hop were developed based on the beta-binomial distribution, u...

  19. Characterization of resistance to powdery mildew in the Hop cultivars Newport and Comet

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Hop powdery mildew, caused by Podosphaera macularis, is an important disease in the Northwestern U.S. Outbreaks of powdery mildew on cultivars previously resistant to the disease have been reported increasingly with the emergence of virulent pathogen strains capable of overcoming a commonly deployed...

  20. Effect of grafting on resistant rootstocks on development of powdery mildew on susceptible watermelon scion, 2012

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An experiment was conducted in a greenhouse at the U.S. Vegetable Laboratory in Charleston, SC, to determine if grafting a powdery mildew susceptible variety (scion) onto watermelon rootstocks with resistance to powdery mildew would provide resistance to foliage of the susceptible scion. Plants of ...

  1. Evaluation and Quantitative trait loci mapping of resistance to powdery mildew in lettuce

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Lettuce (Lactuca sativa L.) is the major leafy vegetable that is susceptible to powdery mildew disease under greenhouse and field conditions. We mapped quantitative trait loci (QTLs) for resistance to powdery mildew under greenhouse conditions in an interspecific population derived from a cross betw...

  2. Oxidative Stress-Mediated Regulation of Proteasome Complexes*

    PubMed Central

    Aiken, Charity T.; Kaake, Robyn M.; Wang, Xiaorong; Huang, Lan

    2011-01-01

    Oxidative stress has been implicated in aging and many human diseases, notably neurodegenerative disorders and various cancers. The reactive oxygen species that are generated by aerobic metabolism and environmental stressors can chemically modify proteins and alter their biological functions. Cells possess protein repair pathways to rescue oxidized proteins and restore their functions. If these repair processes fail, oxidized proteins may become cytotoxic. Cell homeostasis and viability are therefore dependent on the removal of oxidatively damaged proteins. Numerous studies have demonstrated that the proteasome plays a pivotal role in the selective recognition and degradation of oxidized proteins. Despite extensive research, oxidative stress-triggered regulation of proteasome complexes remains poorly defined. Better understanding of molecular mechanisms underlying proteasome function in response to oxidative stress will provide a basis for developing new strategies aimed at improving cell viability and recovery as well as attenuating oxidation-induced cytotoxicity associated with aging and disease. Here we highlight recent advances in the understanding of proteasome structure and function during oxidative stress and describe how cells cope with oxidative stress through proteasome-dependent degradation pathways. PMID:21543789

  3. Bacteriophage-mediated toxin gene regulation in Clostridium difficile.

    PubMed

    Govind, Revathi; Vediyappan, Govindsamy; Rolfe, Rial D; Dupuy, Bruno; Fralick, Joe A

    2009-12-01

    Clostridium difficile has been identified as the most important single identifiable cause of nosocomial antibiotic-associated diarrhea and colitis. Virulent strains of C. difficile produce two large protein toxins, toxin A and toxin B, which are involved in pathogenesis. In this study, we examined the effect of lysogeny by PhiCD119 on C. difficile toxin production. Transcriptional analysis demonstrated a decrease in the expression of pathogenicity locus (PaLoc) genes tcdA, tcdB, tcdR, tcdE, and tcdC in PhiCD119 lysogens. During this study we found that repR, a putative repressor gene of PhiCD119, was expressed in C. difficile lysogens and that its product, RepR, could downregulate tcdA::gusA and tcdR::gusA reporter fusions in Escherichia coli. We cloned and purified a recombinant RepR containing a C-terminal six-His tag and documented its binding to the upstream regions of tcdR in C. difficile PaLoc and in repR upstream region in PhiCD119 by gel shift assays. DNA footprinting experiments revealed similarities between the RepR binding sites in tcdR and repR upstream regions. These findings suggest that presence of a CD119-like temperate phage can influence toxin gene regulation in this nosocomially important pathogen. PMID:19776116

  4. Regulation of PKC Mediated Signaling by Calcium during Visceral Leishmaniasis

    PubMed Central

    Roy, Nivedita; Chakraborty, Supriya; Paul Chowdhury, Bidisha; Banerjee, Sayantan; Halder, Kuntal; Majumder, Saikat; Majumdar, Subrata; Sen, Parimal C.

    2014-01-01

    Calcium is an ubiquitous cellular signaling molecule that controls a variety of cellular processes and is strictly maintained in the cellular compartments by the coordination of various Ca2+ pumps and channels. Two such fundamental calcium pumps are plasma membrane calcium ATPase (PMCA) and Sarco/endoplasmic reticulum calcium ATPase (SERCA) which play a pivotal role in maintaining intracellular calcium homeostasis. This intracellular Ca2+ homeostasis is often disturbed by the protozoan parasite Leishmania donovani, the causative organism of visceral leishmaniasis. In the present study we have dileneated the involvement of PMCA4 and SERCA3 during leishmaniasis. We have observed that during leishmaniasis, intracellular Ca2+ concentration was up-regulated and was further controlled by both PMCA4 and SERCA3. Inhibition of these two Ca2+-ATPases resulted in decreased parasite burden within the host macrophages due to enhanced intracellular Ca2+. Contrastingly, on the other hand, activation of PMCA4 was found to enhance the parasite burden. Our findings also highlighted the importance of Ca2+ in the modulation of cytokine balance during leishmaniasis. These results thus cumulatively suggests that these two Ca2+-ATPases play prominent roles during visceral leishmaniasis. PMID:25329062

  5. Bacteriophage-Mediated Toxin Gene Regulation in Clostridium difficile▿

    PubMed Central

    Govind, Revathi; Vediyappan, Govindsamy; Rolfe, Rial D.; Dupuy, Bruno; Fralick, Joe A.

    2009-01-01

    Clostridium difficile has been identified as the most important single identifiable cause of nosocomial antibiotic-associated diarrhea and colitis. Virulent strains of C. difficile produce two large protein toxins, toxin A and toxin B, which are involved in pathogenesis. In this study, we examined the effect of lysogeny by ΦCD119 on C. difficile toxin production. Transcriptional analysis demonstrated a decrease in the expression of pathogenicity locus (PaLoc) genes tcdA, tcdB, tcdR, tcdE, and tcdC in ΦCD119 lysogens. During this study we found that repR, a putative repressor gene of ΦCD119, was expressed in C. difficile lysogens and that its product, RepR, could downregulate tcdA::gusA and tcdR::gusA reporter fusions in Escherichia coli. We cloned and purified a recombinant RepR containing a C-terminal six-His tag and documented its binding to the upstream regions of tcdR in C. difficile PaLoc and in repR upstream region in ΦCD119 by gel shift assays. DNA footprinting experiments revealed similarities between the RepR binding sites in tcdR and repR upstream regions. These findings suggest that presence of a CD119-like temperate phage can influence toxin gene regulation in this nosocomially important pathogen. PMID:19776116

  6. Regulation of PKC mediated signaling by calcium during visceral leishmaniasis.

    PubMed

    Roy, Nivedita; Chakraborty, Supriya; Paul Chowdhury, Bidisha; Banerjee, Sayantan; Halder, Kuntal; Majumder, Saikat; Majumdar, Subrata; Sen, Parimal C

    2014-01-01

    Calcium is an ubiquitous cellular signaling molecule that controls a variety of cellular processes and is strictly maintained in the cellular compartments by the coordination of various Ca2+ pumps and channels. Two such fundamental calcium pumps are plasma membrane calcium ATPase (PMCA) and Sarco/endoplasmic reticulum calcium ATPase (SERCA) which play a pivotal role in maintaining intracellular calcium homeostasis. This intracellular Ca2+ homeostasis is often disturbed by the protozoan parasite Leishmania donovani, the causative organism of visceral leishmaniasis. In the present study we have dileneated the involvement of PMCA4 and SERCA3 during leishmaniasis. We have observed that during leishmaniasis, intracellular Ca2+ concentration was up-regulated and was further controlled by both PMCA4 and SERCA3. Inhibition of these two Ca2+-ATPases resulted in decreased parasite burden within the host macrophages due to enhanced intracellular Ca2+. Contrastingly, on the other hand, activation of PMCA4 was found to enhance the parasite burden. Our findings also highlighted the importance of Ca2+ in the modulation of cytokine balance during leishmaniasis. These results thus cumulatively suggests that these two Ca2+-ATPases play prominent roles during visceral leishmaniasis. PMID:25329062

  7. Nrf2-Mediated Regulation of Skeletal Muscle Glycogen Metabolism.

    PubMed

    Uruno, Akira; Yagishita, Yoko; Katsuoka, Fumiki; Kitajima, Yasuo; Nunomiya, Aki; Nagatomi, Ryoichi; Pi, Jingbo; Biswal, Shyam S; Yamamoto, Masayuki

    2016-06-01

    Nrf2 (NF-E2-related factor 2) contributes to the maintenance of glucose homeostasis in vivo Nrf2 suppresses blood glucose levels by protecting pancreatic β cells from oxidative stress and improving peripheral tissue glucose utilization. To elucidate the molecular mechanisms by which Nrf2 contributes to the maintenance of glucose homeostasis, we generated skeletal muscle (SkM)-specific Keap1 knockout (Keap1MuKO) mice that express abundant Nrf2 in their SkM and then examined Nrf2 target gene expression in that tissue. In Keap1MuKO mice, blood glucose levels were significantly downregulated and the levels of the glycogen branching enzyme (Gbe1) and muscle-type PhKα subunit (Phka1) mRNAs, along with those of the glycogen branching enzyme (GBE) and the phosphorylase b kinase α subunit (PhKα) protein, were significantly upregulated in mouse SkM. Consistent with this result, chemical Nrf2 inducers promoted Gbe1 and Phka1 mRNA expression in both mouse SkM and C2C12 myotubes. Chromatin immunoprecipitation analysis demonstrated that Nrf2 binds the Gbe1 and Phka1 upstream promoter regions. In Keap1MuKO mice, muscle glycogen content was strongly reduced and forced GBE expression in C2C12 myotubes promoted glucose uptake. Therefore, our results demonstrate that Nrf2 induction in SkM increases GBE and PhKα expression and reduces muscle glycogen content, resulting in improved glucose tolerance. Our results also indicate that Nrf2 differentially regulates glycogen metabolism in SkM and the liver. PMID:27044864

  8. Phasevarion mediated epigenetic gene regulation in Helicobacter pylori.

    PubMed

    Srikhanta, Yogitha N; Gorrell, Rebecca J; Steen, Jason A; Gawthorne, Jayde A; Kwok, Terry; Grimmond, Sean M; Robins-Browne, Roy M; Jennings, Michael P

    2011-01-01

    Many host-adapted bacterial pathogens contain DNA methyltransferases (mod genes) that are subject to phase-variable expression (high-frequency reversible ON/OFF switching of gene expression). In Haemophilus influenzae and pathogenic Neisseria, the random switching of the modA gene, associated with a phase-variable type III restriction modification (R-M) system, controls expression of a phase-variable regulon of genes (a "phasevarion"), via differential methylation of the genome in the modA ON and OFF states. Phase-variable type III R-M systems are also found in Helicobacter pylori, suggesting that phasevarions may also exist in this key human pathogen. Phylogenetic studies on the phase-variable type III modH gene revealed that there are 17 distinct alleles in H. pylori, which differ only in their DNA recognition domain. One of the most commonly found alleles was modH5 (16% of isolates). Microarray analysis comparing the wild-type P12modH5 ON strain to a P12ΔmodH5 mutant revealed that six genes were either up- or down-regulated, and some were virulence-associated. These included flaA, which encodes a flagella protein important in motility and hopG, an outer membrane protein essential for colonization and associated with gastric cancer. This study provides the first evidence of this epigenetic mechanism of gene expression in H. pylori. Characterisation of H. pylori modH phasevarions to define stable immunological targets will be essential for vaccine development and may also contribute to understanding H. pylori pathogenesis. PMID:22162751

  9. Parental conflict resolution styles and children's adjustment: children's appraisals and emotion regulation as mediators.

    PubMed

    Siffert, Andrea; Schwarz, Beate

    2011-01-01

    Guided by the emotional security hypothesis and the cognitive-contextual framework, the authors investigated whether the associations between negative parental conflict resolution styles and children's internalizing and externalizing problems were mediated by children's appraisals of threat and self-blame and their emotion regulation. Participants were 192 Swiss 2-parent families with children aged 9-12 years (M age = 10.62 years, SD = 0.41 years). Structural equation modeling was used to test the empirical validity of the theoretical model. Results indicated that children's maladaptive emotion regulation mediated the association between negative parental conflict resolution styles and children's internalizing as well as externalizing problems. Whereas perceived threat was related only to children's internalizing problems, self-blame did not mediate the links between negative parental conflict resolution styles and children's adjustment. Implications for understanding the mechanisms by which exposure to interparental conflict could lead to children's maladjustment and limitations of the study are discussed. PMID:21452750

  10. Sex bias in CNS autoimmune disease mediated by androgen control of autoimmune regulator.

    PubMed

    Zhu, Meng-Lei; Bakhru, Pearl; Conley, Bridget; Nelson, Jennifer S; Free, Meghan; Martin, Aaron; Starmer, Joshua; Wilson, Elizabeth M; Su, Maureen A

    2016-01-01

    Male gender is protective against multiple sclerosis and other T-cell-mediated autoimmune diseases. This protection may be due, in part, to higher androgen levels in males. Androgen binds to the androgen receptor (AR) to regulate gene expression, but how androgen protects against autoimmunity is not well understood. Autoimmune regulator (Aire) prevents autoimmunity by promoting self-antigen expression in medullary thymic epithelial cells, such that developing T cells that recognize these self-antigens within the thymus undergo clonal deletion. Here we show that androgen upregulates Aire-mediated thymic tolerance to protect against autoimmunity. Androgen recruits AR to Aire promoter regions, with consequent enhancement of Aire transcription. In mice and humans, thymic Aire expression is higher in males compared with females. Androgen administration and male gender protect against autoimmunity in a multiple sclerosis mouse model in an Aire-dependent manner. Thus, androgen control of an intrathymic Aire-mediated tolerance mechanism contributes to gender differences in autoimmunity. PMID:27072778

  11. Emotion regulation difficulties mediate associations between betrayal trauma and symptoms of posttraumatic stress, depression, and anxiety.

    PubMed

    Goldsmith, Rachel E; Chesney, Samantha A; Heath, Nicole M; Barlow, M Rose

    2013-06-01

    Emotion regulation difficulties following trauma exposure have received increasing attention among researchers and clinicians. Previous work highlights the role of emotion regulation difficulties in multiple forms of psychological distress and identifies emotion regulation capacities as especially compromised among survivors of betrayal trauma: physical, sexual, or emotional maltreatment perpetrated by someone to whom the victim is close, such as a parent or partner. It is unknown, however, whether links between emotion regulation difficulties and psychological symptoms differ following exposure to betrayal trauma as compared with other trauma types. In the present study, 593 male and female university undergraduates completed the Difficulties with Emotion Regulation Scale (Gratz & Roemer, 2004), the Brief Betrayal Trauma Scale (Goldberg & Freyd, 2006), the Impact of Event Scale (Horowitz, Wilner, & Alvarez, 1979), and the Trauma Symptom Checklist (Elliott & Briere, 1992). A path analytic model demonstrated that betrayal trauma indirectly impacted symptoms of intrusion (β = .11), avoidance (β = .13), depression (β = .17), and anxiety (β = .14) via emotion regulation difficulties, an effect consistent with mediation. Emotion regulation difficulties did not mediate the relationship between other trauma exposure and psychological symptoms. Results may inform treatment-matching efforts, and suggest that emotion regulation difficulties may constitute a key therapeutic target following betrayal trauma. PMID:23737296

  12. Naturally occurring broad-spectrum powdery mildew resistance in a Central American tomato accession is caused by loss of mlo function.

    PubMed

    Bai, Yuling; Pavan, Stefano; Zheng, Zheng; Zappel, Nana F; Reinstädler, Anja; Lotti, Concetta; De Giovanni, Claudio; Ricciardi, Luigi; Lindhout, Pim; Visser, Richard; Theres, Klaus; Panstruga, Ralph

    2008-01-01

    The resistant cherry tomato (Solanum lycopersicum var. cerasiforme) line LC-95, derived from an accession collected in Ecuador, harbors a natural allele (ol-2) that confers broad-spectrum and recessively inherited resistance to powdery mildew (Oidium neolycopersici). As both the genetic and phytopathological characteristics of ol-2-mediated resistance are reminiscent of powdery mildew immunity conferred by loss-of-function mlo alleles in barley and Arabidopsis, we initiated a candidate-gene approach to clone Ol-2. A tomato Mlo gene (SlMlo1) with high sequence-relatedness to barley Mlo and Arabidopsis AtMLO2 mapped to the chromosomal region harboring the Ol-2 locus. Complementation experiments using transgenic tomato lines as well as virus-induced gene silencing assays suggested that loss of SlMlo1 function is responsible for powdery mildew resistance conferred by ol-2. In progeny of a cross between a resistant line bearing ol-2 and the susceptible tomato cultivar Moneymaker, a 19-bp deletion disrupting the SlMlo1 coding region cosegregated with resistance. This polymorphism results in a frameshift and, thus, a truncated nonfunctional SlMlo1 protein. Our findings reveal the second example of a natural mlo mutant that possibly arose post-domestication, suggesting that natural mlo alleles might be evolutionarily short-lived due to fitness costs related to loss of mlo function. PMID:18052880

  13. Neuronal Wiskott-Aldrich syndrome protein regulates TGF-β1-mediated lung vascular permeability.

    PubMed

    Wagener, Brant M; Hu, Meng; Zheng, Anni; Zhao, Xueke; Che, Pulin; Brandon, Angela; Anjum, Naseem; Snapper, Scott; Creighton, Judy; Guan, Jun-Lin; Han, Qimei; Cai, Guo-Qiang; Han, Xiaosi; Pittet, Jean-Francois; Ding, Qiang

    2016-07-01

    TGF-β1 induces an increase in paracellular permeability and actin stress fiber formation in lung microvascular endothelial and alveolar epithelial cells via small Rho GTPase. The molecular mechanism involved is not fully understood. Neuronal Wiskott-Aldrich syndrome protein (N-WASP) has an essential role in actin structure dynamics. We hypothesized that N-WASP plays a critical role in these TGF-β1-induced responses. In these cell monolayers, we demonstrated that N-WASP down-regulation by short hairpin RNA prevented TGF-β1-mediated disruption of the cortical actin structure, actin stress filament formation, and increased permeability. Furthermore, N-WASP down-regulation blocked TGF-β1 activation mediated by IL-1β in alveolar epithelial cells, which requires actin stress fiber formation. Control short hairpin RNA had no effect on these TGF-β1-induced responses. TGF-β1-induced phosphorylation of Y256 of N-WASP via activation of small Rho GTPase and focal adhesion kinase mediates TGF-β1-induced paracellular permeability and actin cytoskeleton dynamics. In vivo, compared with controls, N-WASP down-regulation increases survival and prevents lung edema in mice induced by bleomycin exposure-a lung injury model in which TGF-β1 plays a critical role. Our data indicate that N-WASP plays a crucial role in the development of TGF-β1-mediated acute lung injury by promoting pulmonary edema via regulation of actin cytoskeleton dynamics.-Wagener, B. M., Hu, M., Zheng, A., Zhao, X., Che, P., Brandon, A., Anjum, N., Snapper, S., Creighton, J., Guan, J.-L., Han, Q., Cai, G.-Q., Han, X., Pittet, J.-F., Ding, Q. Neuronal Wiskott-Aldrich syndrome protein regulates TGF-β1-mediated lung vascular permeability. PMID:27025963

  14. Mother's, Father's Regulation and Child's Self-Regulation in a Computer-Mediated Learning Situation

    ERIC Educational Resources Information Center

    Nader-Grosbois, Nathalie; Normandeau, Sylvie; Ricard-Cossette, Marcelle; Quintal, Germain

    2008-01-01

    This study has explored the parents' regulation strategies that were more likely to support children's self-regulation in learning situations with computers. These strategies have been analysed by means of new grids involving seven categories of behaviour: cognitive strategies relating to identification of objective, exploration of means,…

  15. Conditional Cooperativity of Toxin - Antitoxin Regulation Can Mediate Bistability between Growth and Dormancy

    PubMed Central

    Cataudella, Ilaria; Sneppen, Kim; Gerdes, Kenn; Mitarai, Namiko

    2013-01-01

    Many toxin-antitoxin operons are regulated by the toxin/antitoxin ratio by mechanisms collectively coined “conditional cooperativity”. Toxin and antitoxin form heteromers with different stoichiometric ratios, and the complex with the intermediate ratio works best as a transcription repressor. This allows transcription at low toxin level, strong repression at intermediate toxin level, and then again transcription at high toxin level. Such regulation has two interesting features; firstly, it provides a non-monotonous response to the concentration of one of the proteins, and secondly, it opens for ultra-sensitivity mediated by the sequestration of the functioning heteromers. We explore possible functions of conditional regulation in simple feedback motifs, and show that it can provide bistability for a wide range of parameters. We then demonstrate that the conditional cooperativity in toxin-antitoxin systems combined with the growth-inhibition activity of free toxin can mediate bistability between a growing state and a dormant state. PMID:24009488

  16. Conditional cooperativity of toxin - antitoxin regulation can mediate bistability between growth and dormancy.

    PubMed

    Cataudella, Ilaria; Sneppen, Kim; Gerdes, Kenn; Mitarai, Namiko

    2013-01-01

    Many toxin-antitoxin operons are regulated by the toxin/antitoxin ratio by mechanisms collectively coined "conditional cooperativity". Toxin and antitoxin form heteromers with different stoichiometric ratios, and the complex with the intermediate ratio works best as a transcription repressor. This allows transcription at low toxin level, strong repression at intermediate toxin level, and then again transcription at high toxin level. Such regulation has two interesting features; firstly, it provides a non-monotonous response to the concentration of one of the proteins, and secondly, it opens for ultra-sensitivity mediated by the sequestration of the functioning heteromers. We explore possible functions of conditional regulation in simple feedback motifs, and show that it can provide bistability for a wide range of parameters. We then demonstrate that the conditional cooperativity in toxin-antitoxin systems combined with the growth-inhibition activity of free toxin can mediate bistability between a growing state and a dormant state. PMID:24009488

  17. Arabidopsis Phospholipase Dδ Is Involved in Basal Defense and Nonhost Resistance to Powdery Mildew Fungi1[W

    PubMed Central

    Pinosa, Francesco; Buhot, Nathalie; Kwaaitaal, Mark; Fahlberg, Per; Thordal-Christensen, Hans; Ellerström, Mats; Andersson, Mats X.

    2013-01-01

    Plants have evolved a complex array of defensive responses against pathogenic microorganisms. Recognition of microbes initiates signaling cascades that activate plant defenses. The membrane lipid phosphatidic acid, produced by phospholipase D (PLD), has been shown to take part in both abiotic and biotic stress signaling. In this study, the involvement of PLD in the interaction between Arabidopsis (Arabidopsis thaliana) and the barley powdery mildew fungus Blumeria graminis f. sp. hordei (Bgh) was investigated. This nonadapted pathogen is normally resisted by a cell wall-based defense, which stops the fungal hyphae from penetrating the epidermal cell wall. Chemical inhibition of phosphatidic acid production by PLD increased the penetration rate of Bgh spores on wild-type leaves. The analysis of transfer DNA knockout lines for all Arabidopsis PLD genes revealed that PLDδ is involved in penetration resistance against Bgh, and chemical inhibition of PLDs in plants mutated in PLDδ indicated that this isoform alone is involved in Bgh resistance. In addition, we confirmed the involvement of PLDδ in penetration resistance against another nonadapted pea powdery mildew fungus, Erysiphe pisi. A green fluorescent protein fusion of PLDδ localized to the plasma membrane at the Bgh attack site, where it surrounded the cell wall reinforcement. Furthermore, in the pldδ mutant, transcriptional up-regulation of early microbe-associated molecular pattern response genes was delayed after chitin stimulation. In conclusion, we propose that PLD is involved in defense signaling in nonhost resistance against powdery mildew fungi and put PLDδ forward as the main isoform participating in this process. PMID:23979971

  18. TPL-2 Regulates Macrophage Lipid Metabolism and M2 Differentiation to Control TH2-Mediated Immunopathology

    PubMed Central

    Entwistle, Lewis J.; Khoury, Hania; Papoutsopoulou, Stamatia; Mahmood, Radma; Mansour, Nuha R.; Ching-Cheng Huang, Stanley; Pearce, Edward J.; Pedro S. de Carvalho, Luiz; Ley, Steven C.

    2016-01-01

    Persistent TH2 cytokine responses following chronic helminth infections can often lead to the development of tissue pathology and fibrotic scarring. Despite a good understanding of the cellular mechanisms involved in fibrogenesis, there are very few therapeutic options available, highlighting a significant medical need and gap in our understanding of the molecular mechanisms of TH2-mediated immunopathology. In this study, we found that the Map3 kinase, TPL-2 (Map3k8; Cot) regulated TH2-mediated intestinal, hepatic and pulmonary immunopathology following Schistosoma mansoni infection or S. mansoni egg injection. Elevated inflammation, TH2 cell responses and exacerbated fibrosis in Map3k8–/–mice was observed in mice with myeloid cell-specific (LysM) deletion of Map3k8, but not CD4 cell-specific deletion of Map3k8, indicating that TPL-2 regulated myeloid cell function to limit TH2-mediated immunopathology. Transcriptional and metabolic assays of Map3k8–/–M2 macrophages identified that TPL-2 was required for lipolysis, M2 macrophage activation and the expression of a variety of genes involved in immuno-regulatory and pro-fibrotic pathways. Taken together this study identified that TPL-2 regulated TH2-mediated inflammation by supporting lipolysis and M2 macrophage activation, preventing TH2 cell expansion and downstream immunopathology and fibrosis. PMID:27487182

  19. TPL-2 Regulates Macrophage Lipid Metabolism and M2 Differentiation to Control TH2-Mediated Immunopathology.

    PubMed

    Kannan, Yashaswini; Perez-Lloret, Jimena; Li, Yanda; Entwistle, Lewis J; Khoury, Hania; Papoutsopoulou, Stamatia; Mahmood, Radma; Mansour, Nuha R; Ching-Cheng Huang, Stanley; Pearce, Edward J; Pedro S de Carvalho, Luiz; Ley, Steven C; Wilson, Mark S

    2016-08-01

    Persistent TH2 cytokine responses following chronic helminth infections can often lead to the development of tissue pathology and fibrotic scarring. Despite a good understanding of the cellular mechanisms involved in fibrogenesis, there are very few therapeutic options available, highlighting a significant medical need and gap in our understanding of the molecular mechanisms of TH2-mediated immunopathology. In this study, we found that the Map3 kinase, TPL-2 (Map3k8; Cot) regulated TH2-mediated intestinal, hepatic and pulmonary immunopathology following Schistosoma mansoni infection or S. mansoni egg injection. Elevated inflammation, TH2 cell responses and exacerbated fibrosis in Map3k8-/-mice was observed in mice with myeloid cell-specific (LysM) deletion of Map3k8, but not CD4 cell-specific deletion of Map3k8, indicating that TPL-2 regulated myeloid cell function to limit TH2-mediated immunopathology. Transcriptional and metabolic assays of Map3k8-/-M2 macrophages identified that TPL-2 was required for lipolysis, M2 macrophage activation and the expression of a variety of genes involved in immuno-regulatory and pro-fibrotic pathways. Taken together this study identified that TPL-2 regulated TH2-mediated inflammation by supporting lipolysis and M2 macrophage activation, preventing TH2 cell expansion and downstream immunopathology and fibrosis. PMID:27487182

  20. A GRHL3-regulated repair pathway suppresses immune-mediated epidermal hyperplasia

    PubMed Central

    Gordon, William M.; Zeller, Michael D.; Klein, Rachel H.; Swindell, William R.; Ho, Hsiang; Espetia, Francisco; Gudjonsson, Johann E.; Baldi, Pierre F.; Andersen, Bogi

    2014-01-01

    Dermal infiltration of T cells is an important step in the onset and progression of immune-mediated skin diseases such as psoriasis; however, it is not known whether epidermal factors play a primary role in the development of these diseases. Here, we determined that the prodifferentiation transcription factor grainyhead-like 3 (GRHL3), which is essential during epidermal development, is dispensable for adult skin homeostasis, but required for barrier repair after adult epidermal injury. Consistent with activation of a GRHL3-regulated repair pathway in psoriasis, we found that GRHL3 is upregulated in lesional skin and binds known epidermal differentiation gene targets. Using an imiquimod-induced model of immune-mediated epidermal hyperplasia, we found that mice lacking GRHL3 have an exacerbated epidermal damage response, greater sensitivity to disease induction, delayed resolution of epidermal lesions, and resistance to anti–IL-22 therapy compared with WT animals. ChIP-Seq and gene expression profiling of murine skin revealed that while GRHL3 regulates differentiation pathways both during development and during repair from immune-mediated damage, it targets distinct sets of genes in the 2 processes. In particular, GRHL3 suppressed a number of alarmin and other proinflammatory genes after immune injury. This study identifies a GRHL3-regulated epidermal barrier repair pathway that suppresses disease initiation and helps resolve existing lesions in immune-mediated epidermal hyperplasia. PMID:25347468

  1. PKCα-Mediated Signals Regulate the Motile Responses of Cochlear Outer Hair Cells

    PubMed Central

    Park, Channy; Kalinec, Federico

    2015-01-01

    There is strong evidence that changes in the actin/spectrin-based cortical cytoskeleton of outer hair cells (OHCs) regulate their motile responses as well as cochlear amplification, the process that optimizes the sensitivity and frequency selectivity of the mammalian inner ear. Since a RhoA/protein kinase C (PKC)-mediated pathway is known to inhibit the actin-spectrin interaction in other cell models, we decided to investigate whether this signaling cascade could also participate in the regulation of OHC motility. We used high-speed video microscopy and confocal microscopy to explore the effects of pharmacological activation of PKCα, PKCβI, PKCβII, PKCδ, PKCε, and PKCζ with lysophosphatidic acid (LPA) and their inhibition with bisindolylmaleimide I, as well as inhibition of RhoA and Rho-associated protein kinase (ROCK) with C3 and Y-27632, respectively. Motile responses were induced in isolated guinea pig OHCs by stimulation with an 8 V/cm external alternating electrical field as 50 Hz bursts of square wave pulses (100 ms on/off). We found that LPA increased expression of PKCα and PKCζ only, with PKCα, but not PKCζ, phosphorylating the cytoskeletal protein adducin of both Ser-726 and Thr-445. Interestingly, however, inhibition of PKCα reduced adducin phosphorylation only at Ser-726. We also determined that LPA activation of a PKCα-mediated signaling pathway simultaneously enhanced OHC electromotile amplitude and cell shortening, and facilitated RhoA/ROCK/LIMK1-mediated cofilin phosphorylation. Altogether, our results suggest that PKCα-mediated signals, probably via adducin-mediated inhibition of actin-spectrin binding and cofilin-mediated depolymerization of actin filaments, play an essential role in the homeostatic regulation of OHC motility and cochlear amplification. PMID:25954875

  2. Protein kinase C regulates tonic GABAA receptor-mediated inhibition in the hippocampus and thalamus

    PubMed Central

    Bright, Damian P; Smart, Trevor G

    2013-01-01

    Tonic inhibition mediated by extrasynaptic GABAA receptors (GABAARs) is an important regulator of neuronal excitability. Phosphorylation by protein kinase C (PKC) provides a key mode of regulation for synaptic GABAARs underlying phasic inhibition; however, less attention has been focused on the plasticity of tonic inhibition and whether this can also be modulated by receptor phosphorylation. To address this issue, we used whole-cell patch clamp recording in acute murine brain slices at both room and physiological temperatures to examine the effects of PKC-mediated phosphorylation on tonic inhibition. Recordings from dentate gyrus granule cells in the hippocampus and dorsal lateral geniculate relay neurons in the thalamus demonstrated that PKC activation caused downregulation of tonic GABAAR-mediated inhibition. Conversely, inhibition of PKC resulted in an increase in tonic GABAAR activity. These findings were corroborated by experiments on human embryonic kidney 293 cells expressing recombinant α4β2δ GABAARs, which represent a key extrasynaptic GABAAR isoform in the hippocampus and thalamus. Using bath application of low GABA concentrations to mimic activation by ambient neurotransmitter, we demonstrated a similar inhibition of receptor function following PKC activation at physiological temperature. Live cell imaging revealed that this was correlated with a loss of cell surface GABAARs. The inhibitory effects of PKC activation on α4β2δ GABAAR activity appeared to be mediated by direct phosphorylation at a previously identified site on the β2 subunit, serine 410. These results indicate that PKC-mediated phosphorylation can be an important physiological regulator of tonic GABAAR-mediated inhibition. PMID:24102973

  3. Importance of Mediator complex in the regulation and integration of diverse signaling pathways in plants

    PubMed Central

    Samanta, Subhasis; Thakur, Jitendra K.

    2015-01-01

    Basic transcriptional machinery in eukaryotes is assisted by a number of cofactors, which either increase or decrease the rate of transcription. Mediator complex is one such cofactor, and recently has drawn a lot of interest because of its integrative power to converge different signaling pathways before channeling the transcription instructions to the RNA polymerase II machinery. Like yeast and metazoans, plants do possess the Mediator complex across the kingdom, and its isolation and subunit analyses have been reported from the model plant, Arabidopsis. Genetic, and molecular analyses have unraveled important regulatory roles of Mediator subunits at every stage of plant life cycle starting from flowering to embryo and organ development, to even size determination. It also contributes immensely to the survival of plants against different environmental vagaries by the timely activation of its resistance mechanisms. Here, we have provided an overview of plant Mediator complex starting from its discovery to regulation of stoichiometry of its subunits. We have also reviewed involvement of different Mediator subunits in different processes and pathways including defense response pathways evoked by diverse biotic cues. Wherever possible, attempts have been made to provide mechanistic insight of Mediator's involvement in these processes. PMID:26442070

  4. Phytochrome-Mediated Light Regulation of PHYA- and PHYB-GUS Transgenes in Arabidopsis thaliana Seedlings.

    PubMed Central

    Somers, D. E.; Quail, P. H.

    1995-01-01

    Phytochrome wild-type gene-[beta]-glucuronidase (PHY-GUS) gene fusions were used in transgenic Arabidopsis to compare the activity levels and light regulation of the PHYA and PHYB promoters and to identify the photoreceptors mediating this regulation. In dark-grown seedlings, both promoters are 4-fold more active in shoots than in roots,but the PHYA promoter is nearly 20-fold more active than that of PHYB in both organs. In shoots, white light represses the activities of the PHYA and PHYB promoters 10- and 2-fold, respectively, whereas in roots light has no effect on the PHYA promoter but increases PHYB promoter activity 2-fold. Consequently, PHYA promoter activity remains higher than that of PHYB in light in both shoots (5-fold) and roots (11-fold). Experiments with narrow-waveband light and photomorphogenic mutants suggest that no single photoreceptor is necessary for full white-light-directed PHYA repression in shoots, but that multiple, independent photoreceptor pathways are sufficient alone or in combination. In contrast, phytochrome B appears both necessary and sufficient for a light-mediated decrease in PHYB activity in shoots, and phytochrome A mediates a far-red-light-stimulated increase in PHYB promoter activity. Together, the data indicate that the PHYA and PHYB genes are regulated in divergent fashion at the transcriptional level, both developmentally and by the spectral distribution of the prevailing light, and that this regulation may be important to the photosensory function of the two photoreceptors. PMID:12228380

  5. Bacillus sp. BS061 Suppresses Powdery Mildew and Gray Mold

    PubMed Central

    Kim, Young-Sook; Song, Ja-Gyeong; Lee, In-Kyoung; Yeo, Woon-Hyung

    2013-01-01

    The use of a microorganism, or its secretions, to prevent plant disease offers an attractive alternative or supplement to synthetic fungicides for the management of plant disease without the negative effects of chemical control mechanisms. During a screening for microorganisms with the potential to be used as microbial fungicides, Bacillus sp. BS061 was isolated from a plant leaf. The strain BS061 potently inhibited the mycelial growth of Botrytis cinerea, and significantly reduced disease incidence of powdery mildew in cucumber and strawberry. We also found that the culture filtrate of BS061 inhibited the mycelial growth of various plant pathogens. PMID:23874134

  6. Arabidopsis PROTEASOME REGULATOR1 is required for auxin-mediated suppression of proteasome activity and regulates auxin signalling.

    PubMed

    Yang, Bao-Jun; Han, Xin-Xin; Yin, Lin-Lin; Xing, Mei-Qing; Xu, Zhi-Hong; Xue, Hong-Wei

    2016-01-01

    The plant hormone auxin is perceived by the nuclear F-box protein TIR1 receptor family and regulates gene expression through degradation of Aux/IAA transcriptional repressors. Several studies have revealed the importance of the proteasome in auxin signalling, but details on how the proteolytic machinery is regulated and how this relates to degradation of Aux/IAA proteins remains unclear. Here we show that an Arabidopsis homologue of the proteasome inhibitor PI31, which we name PROTEASOME REGULATOR1 (PTRE1), is a positive regulator of the 26S proteasome. Loss-of-function ptre1 mutants are insensitive to auxin-mediated suppression of proteasome activity, show diminished auxin-induced degradation of Aux/IAA proteins and display auxin-related phenotypes. We found that auxin alters the subcellular localization of PTRE1, suggesting this may be part of the mechanism by which it reduces proteasome activity. Based on these results, we propose that auxin regulates proteasome activity via PTRE1 to fine-tune the homoeostasis of Aux/IAA repressor proteins thus modifying auxin activity. PMID:27109828

  7. Arabidopsis PROTEASOME REGULATOR1 is required for auxin-mediated suppression of proteasome activity and regulates auxin signalling

    PubMed Central

    Yang, Bao-Jun; Han, Xin-Xin; Yin, Lin-Lin; Xing, Mei-Qing; Xu, Zhi-Hong; Xue, Hong-Wei

    2016-01-01

    The plant hormone auxin is perceived by the nuclear F-box protein TIR1 receptor family and regulates gene expression through degradation of Aux/IAA transcriptional repressors. Several studies have revealed the importance of the proteasome in auxin signalling, but details on how the proteolytic machinery is regulated and how this relates to degradation of Aux/IAA proteins remains unclear. Here we show that an Arabidopsis homologue of the proteasome inhibitor PI31, which we name PROTEASOME REGULATOR1 (PTRE1), is a positive regulator of the 26S proteasome. Loss-of-function ptre1 mutants are insensitive to auxin-mediated suppression of proteasome activity, show diminished auxin-induced degradation of Aux/IAA proteins and display auxin-related phenotypes. We found that auxin alters the subcellular localization of PTRE1, suggesting this may be part of the mechanism by which it reduces proteasome activity. Based on these results, we propose that auxin regulates proteasome activity via PTRE1 to fine-tune the homoeostasis of Aux/IAA repressor proteins thus modifying auxin activity. PMID:27109828

  8. Regulation of Cell Cycle Regulators by SIRT1 Contributes to Resveratrol-Mediated Prevention of Pulmonary Arterial Hypertension

    PubMed Central

    Zhou, Shuang; Li, Meng-Tao; Jia, Yu-Yan; Liu, Jin-Jing; Wang, Qian; Tian, Zhuang; Liu, Yong-Tai; Chen, Hou-Zao; Liu, De-Pei; Zeng, Xiao-Feng

    2015-01-01

    Pulmonary arterial hypertension (PAH) is a major cause of morbidity and mortality in rheumatic diseases. Vascular remodeling due to the proliferation of pulmonary arterial smooth muscle cells (PASMCs) is central to the development of PAH. To date, it is still unclear if Silence Information Regulator 1 (SIRT1) regulates cell cycle regulators in the proliferation of PASMCs and contributes to prevention of PAH by resveratrol. In this study, we found that a significant decrease of SIRT1 expression levels in platelet-derived growth factor BB (PDGF-BB) treated human PASMCs (HPASMCs) and in monocrotaline (MCT) induced PAH rat. Overexpression of SIRT1 induced G1 phase arrest and increased p21 expression but decreased cyclin D1 expression in PDGF-BB treated HPASMCs. Moreover, resveratrol attenuated pulmonary arterial remodeling, decreased pulmonary arterial pressure, and upregulated SIRT1 and p21 expression but downregulated cyclin D1 expression in MCT induced PAH rat. Notably, knockdown of SIRT1 eliminated the regulation of resveratrol on p21 and cyclin D1 expression in PDGF-BB treated HPASMCs. These results demonstrated that SIRT1 mediated the regulation of resveratrol on the expression of cell cycle regulatory molecules. It suggests that SIRT1 exerts a protective role in PAH associated with rheumatic diseases and can be a potential treatment target. PMID:26273643

  9. Posttraumatic stress and growth among Tibetan refugees: the mediating role of cognitive-emotional regulation strategies.

    PubMed

    Hussain, Dilwar; Bhushan, Braj

    2011-07-01

    This study examined posttraumatic stress (PTS) and posttraumatic growth (PTG) among 226 Tibetan refugees across two generations. Additional objectives were to (i) examine the sex and generation differences on the scores of trauma, PTS, and PTG, (ii) explore the relationship between traumatic experiences, PTS and PTG, and (iii) investigate the mediating effect of cognitive-emotional regulation strategies between the traumatic experiences and PTS as well as PTG. Females scored higher on trauma, PTS, and PTG. The trauma, PTS, and PTG scores of the two generations were significantly different. Acceptance and putting into perspective partially mediated the relationship between traumatic experience and PTS. Positive refocusing, refocus on planning, putting into perspective, and catastrophisizing partially mediated the relationship between traumatic experiences and PTG. PMID:21455959

  10. sRNA-Mediated Regulation of P-Fimbriae Phase Variation in Uropathogenic Escherichia coli.

    PubMed

    Khandige, Surabhi; Kronborg, Tina; Uhlin, Bernt Eric; Møller-Jensen, Jakob

    2015-08-01

    Uropathogenic Escherichia coli (UPEC) are capable of occupying physiologically distinct intracellular and extracellular niches within the urinary tract. This feat requires the timely regulation of gene expression and small RNAs (sRNAs) are known to mediate such rapid adjustments in response to changing environmental cues. This study aimed to uncover sRNA-mediated gene regulation in the UPEC strain UTI89, during infection of bladder epithelial cells. Hfq is an RNA chaperone known to facilitate and stabilize sRNA and target mRNA interactions with bacterial cells. The co-immunoprecipitation and high throughput RNA sequencing of Hfq bound sRNAs performed in this study, revealed distinct sRNA profiles in UPEC in the extracellular and intracellular environments. Our findings emphasize the importance of studying regulatory sRNAs in a biologically relevant niche. This strategy also led to the discovery of a novel virulence-associated trans-acting sRNA-PapR. Deletion of papR was found to enhance adhesion of UTI89 to both bladder and kidney cell lines in a manner independent of type-1 fimbriae. We demonstrate PapR mediated posttranscriptional repression of the P-fimbriae phase regulator gene papI and postulate a role for such regulation in fimbrial cross-talk at the population level in UPEC. Our results further implicate the Leucine responsive protein (LRP) as a transcriptional activator regulating PapR expression. Our study reports, for the first time, a role for sRNAs in regulation of P-fimbriae phase variation and emphasizes the importance of studying pathogenesis-specific sRNAs within a relevant biological niche. PMID:26291711

  11. Regulation of secretory transport by protein kinase D-mediated phosphorylation of the ceramide transfer protein.

    PubMed

    Fugmann, Tim; Hausser, Angelika; Schöffler, Patrik; Schmid, Simone; Pfizenmaier, Klaus; Olayioye, Monilola A

    2007-07-01

    Protein kinase D (PKD) has been identified as a crucial regulator of secretory transport at the trans-Golgi network (TGN). Recruitment and activation of PKD at the TGN is mediated by the lipid diacylglycerol, a pool of which is generated by sphingomyelin synthase from ceramide and phosphatidylcholine. The nonvesicular transfer of ceramide from the endoplasmic reticulum to the Golgi complex is mediated by the lipid transfer protein CERT (ceramide transport). In this study, we identify CERT as a novel in vivo PKD substrate. Phosphorylation on serine 132 by PKD decreases the affinity of CERT toward its lipid target phosphatidylinositol 4-phosphate at Golgi membranes and reduces ceramide transfer activity, identifying PKD as a regulator of lipid homeostasis. We also show that CERT, in turn, is critical for PKD activation and PKD-dependent protein cargo transport to the plasma membrane. Thus, the interdependence of PKD and CERT is key to the maintenance of Golgi membrane integrity and secretory transport. PMID:17591919

  12. Regulation of secretory transport by protein kinase D–mediated phosphorylation of the ceramide transfer protein

    PubMed Central

    Fugmann, Tim; Hausser, Angelika; Schöffler, Patrik; Schmid, Simone; Pfizenmaier, Klaus; Olayioye, Monilola A.

    2007-01-01

    Protein kinase D (PKD) has been identified as a crucial regulator of secretory transport at the trans-Golgi network (TGN). Recruitment and activation of PKD at the TGN is mediated by the lipid diacylglycerol, a pool of which is generated by sphingomyelin synthase from ceramide and phosphatidylcholine. The nonvesicular transfer of ceramide from the endoplasmic reticulum to the Golgi complex is mediated by the lipid transfer protein CERT (ceramide transport). In this study, we identify CERT as a novel in vivo PKD substrate. Phosphorylation on serine 132 by PKD decreases the affinity of CERT toward its lipid target phosphatidylinositol 4-phosphate at Golgi membranes and reduces ceramide transfer activity, identifying PKD as a regulator of lipid homeostasis. We also show that CERT, in turn, is critical for PKD activation and PKD-dependent protein cargo transport to the plasma membrane. Thus, the interdependence of PKD and CERT is key to the maintenance of Golgi membrane integrity and secretory transport. PMID:17591919

  13. Self-Regulation Mediates the Relationship between Learner Typology and Achievement in At - Risk Children

    PubMed Central

    Weed, Keri; Keogh, Deborah; Borkowski, John G.; Whitman, Thomas; Noria, Christine W.

    2010-01-01

    A person-centered approach was used to explore the mediating role of self-regulation between learner typology at age 8 and academic achievement at age 14while controlling for domain-specific achievement in a longitudinal sample of 113 children born to adolescent mothers. Children were classified into one of 5 learner typologies at age 8based on interactive patterns of intellectual, achievement, and adaptive abilities. Typology classification explained significant variance in both reading and mathematics achievement at age 14. A bootstrapping approach confirmed that self-regulation mediated the relationship between typology and reading and mathematical achievement for children from all typologies except those classified as Cognitively and Adaptively Challenged. Implications of person-centered approaches for understanding processes involved with achievement are discussed. PMID:21278904

  14. TRIM29 regulates the p63-mediated pathway in cervical cancer cells.

    PubMed

    Masuda, Yasushi; Takahashi, Hidehisa; Hatakeyama, Shigetsugu

    2015-10-01

    Cell invasion and adhesion play an important role in cancer metastasis and are orchestrated by a complicated network of transcription factors including p63. Here, we show that a member of the tripartite motif protein family, TRIM29, is required for regulation of the p63-mediated pathway in cervical cancer cells. TRIM29 knockdown alters the adhesion and invasion activities of cervical cancer cells. TRIM29 knockdown and overexpression cause a significant decrease and increase of TAp63α expression, respectively. TRIM29 knockdown alters the expression pattern of integrins and increases ZEB1 expression. TRIM29 is required for suppression of an increase in the adhesion activity of cells by TAp63α. These findings suggest that TRIM29 regulates the p63-mediated pathway and the behavior of cervical cancer cells. PMID:26071105

  15. Nitric oxide mediates bleomycin-induced angiogenesis and pulmonary fibrosis via regulation of VEGF.

    PubMed

    Iyer, Anand Krishnan V; Ramesh, Vani; Castro, Carlos A; Kaushik, Vivek; Kulkarni, Yogesh M; Wright, Clayton A; Venkatadri, Rajkumar; Rojanasakul, Yon; Azad, Neelam

    2015-11-01

    Pulmonary fibrosis is a progressive lung disease hallmarked by increased fibroblast proliferation, amplified levels of extracellular matrix deposition and increased angiogenesis. Although dysregulation of angiogenic mediators has been implicated in pulmonary fibrosis, the specific rate-limiting angiogenic markers involved and their role in the progression of pulmonary fibrosis remains unclear. We demonstrate that bleomycin treatment induces angiogenesis, and inhibition of the central angiogenic mediator VEGF using anti-VEGF antibody CBO-P11 significantly attenuates bleomycin-induced pulmonary fibrosis in vivo. Bleomycin-induced nitric oxide (NO) was observed to be the key upstream regulator of VEGF via the PI3k/Akt pathway. VEGF regulated other important angiogenic proteins including PAI-1 and IL-8 in response to bleomycin exposure. Inhibition of NO and VEGF activity significantly mitigated bleomycin-induced angiogenic and fibrogenic responses. NO and VEGF are key mediators of bleomycin-induced pulmonary fibrosis, and could serve as important targets against this debilitating disease. Overall, our data suggests an important role for angiogenic mediators in the pathogenesis of bleomycin-induced pulmonary fibrosis. PMID:25919965

  16. Mechanisms of cohesin-mediated gene regulation and lessons learned from cohesinopathies

    PubMed Central

    Ball, Alexander R.; Chen, Yen-Yun; Yokomori, Kyoko

    2014-01-01

    Cohesins are conserved and essential Structural Maintenance of Chromosomes (SMC) protein-containing complexes that physically interact with chromatin and modulate higher-order chromatin organization. Cohesins mediate sister chromatid cohesion and cellular long-distance chromatin interactions affecting genome maintenance and gene expression. Discoveries of mutations in cohesin’s subunits and its regulator proteins in human developmental disorders, so-called “cohesinopathies,” reveal crucial roles for cohesins in development and cellular growth and differentiation. In this review, we discuss the latest findings concerning cohesin’s functions in higher-order chromatin architecture organization and gene regulation and new insight gained from studies of cohesinopathies. PMID:24269489

  17. TRIM45 negatively regulates NF-{kappa}B-mediated transcription and suppresses cell proliferation

    SciTech Connect

    Shibata, Mio; Sato, Tomonobu; Nukiwa, Ryota; Ariga, Tadashi; Hatakeyama, Shigetsugu

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer NF-{kappa}B plays an important role in cell survival and carcinogenesis. Black-Right-Pointing-Pointer TRIM45 negatively regulates TNF{alpha}-induced NF-{kappa}B-mediated transcription. Black-Right-Pointing-Pointer TRIM45 overexpression suppresses cell growth. Black-Right-Pointing-Pointer TRIM45 acts as a repressor for the NF-{kappa}B signal and regulates cell growth. -- Abstract: The NF-{kappa}B signaling pathway plays an important role in cell survival, immunity, inflammation, carcinogenesis, and organogenesis. Activation of NF-{kappa}B is regulated by several posttranslational modifications including phosphorylation, neddylation and ubiquitination. The NF-{kappa}B signaling pathway is activated by two distinct signaling mechanisms and is strictly modulated by the ubiquitin-proteasome system. It has been reported that overexpression of TRIM45, one of the TRIM family ubiquitin ligases, suppresses transcriptional activities of Elk-1 and AP-1, which are targets of the MAPK signaling pathway. In this study, we showed that TRIM45 also negatively regulates TNF{alpha}-induced NF-{kappa}B-mediated transcription by a luciferase reporter assay and that TRIM45 lacking a RING domain also has an activity to inhibit the NF-{kappa}B signal. Moreover, we found that TRIM45 overexpression suppresses cell growth. These findings suggest that TRIM45 acts as a repressor for the NF-{kappa}B signal and regulates cell growth.

  18. 20-hydroxyecdysone mediates non-canonical regulation of mosquito vitellogenins through alternative splicing.

    PubMed

    Provost-Javier, K N; Rasgon, J L

    2014-08-01

    Vitellogenesis is one of the most well-studied physiological processes in mosquitoes. Expression of mosquito vitellogenin genes is classically described as being restricted to female adult reproduction. We report premature vitellogenin transcript expression in three vector mosquitoes: Culex tarsalis, Aedes aegypti and Anopheles gambiae. Vitellogenins expressed during non-reproductive stages are alternatively spliced to retain their first intron and encode premature termination codons. We show that intron retention results in transcript degradation by translation-dependent nonsense-mediated mRNA decay. This is probably an example of regulated unproductive splicing and translation (RUST), a mechanism known to regulate gene expression in numerous organisms but which has never been described in mosquitoes. We demonstrate that the hormone 20-hydroxyecdysone (20E) is responsible for regulating post-transcriptional splicing of vitellogenin. After exposure of previtellogenic fat bodies to 20E, vitellogenin expression switches from a non-productive intron-retaining transcript to a spliced protein-coding transcript. This effect is independent of factors classically known to influence transcription, such as juvenile hormone-mediated competence and amino acid signalling through the target of rapamycin pathway. Non-canonical regulation of vitellogenesis through RUST is a novel role for the multifunctional hormone 20E, and may have important implications for general patterns of gene regulation in mosquitoes. PMID:24720618

  19. 20-hydroxyecdysone mediates non-canonical regulation of mosquito vitellogenins through alternative splicing

    PubMed Central

    Provost-Javier, K. N.; Rasgon, J. L.

    2015-01-01

    Vitellogenesis is one of the most well-studied physiological processes in mosquitoes. Expression of mosquito vitellogenin genes is classically described as being restricted to female adult reproduction. We report premature vitellogenin transcript expression in three vector mosquitoes: Culex tarsalis, Aedes aegypti and Anopheles gambiae. Vitellogenins expressed during non-reproductive stages are alternatively spliced to retain their first intron and encode premature termination codons. We show that intron retention results in transcript degradation by translation-dependent nonsense-mediated mRNA decay. This is probably an example of regulated unproductive splicing and translation (RUST), a mechanism known to regulate gene expression in numerous organisms but which has never been described in mosquitoes. We demonstrate that the hormone 20-hydroxyecdysone (20E) is responsible for regulating post-transcriptional splicing of vitellogenin. After exposure of previtellogenic fat bodies to 20E, vitellogenin expression switches from a non-productive intron-retaining transcript to a spliced protein-coding transcript. This effect is independent of factors classically known to influence transcription, such as juvenile hormone-mediated competence and amino acid signalling through the target of rapamycin pathway. Non-canonical regulation of vitellogenesis through RUST is a novel role for the multifunctional hormone 20E, and may have important implications for general patterns of gene regulation in mosquitoes. PMID:24720618

  20. Thermo-Regulation of Genes Mediating Motility and Plant Interactions in Pseudomonas syringae

    PubMed Central

    Hockett, Kevin L.; Burch, Adrien Y.; Lindow, Steven E.

    2013-01-01

    Pseudomonas syringae is an important phyllosphere colonist that utilizes flagellum-mediated motility both as a means to explore leaf surfaces, as well as to invade into leaf interiors, where it survives as a pathogen. We found that multiple forms of flagellum-mediated motility are thermo-suppressed, including swarming and swimming motility. Suppression of swarming motility occurs between 28° and 30°C, which coincides with the optimal growth temperature of P. syringae. Both fliC (encoding flagellin) and syfA (encoding a non-ribosomal peptide synthetase involved in syringafactin biosynthesis) were suppressed with increasing temperature. RNA-seq revealed 1440 genes of the P. syringae genome are temperature sensitive in expression. Genes involved in polysaccharide synthesis and regulation, phage and IS elements, type VI secretion, chemosensing and chemotaxis, translation, flagellar synthesis and motility, and phytotoxin synthesis and transport were generally repressed at 30°C, while genes involved in transcriptional regulation, quaternary ammonium compound metabolism and transport, chaperone/heat shock proteins, and hypothetical genes were generally induced at 30°C. Deletion of flgM, a key regulator in the transition from class III to class IV gene expression, led to elevated and constitutive expression of fliC regardless of temperature, but did not affect thermo-regulation of syfA. This work highlights the importance of temperature in the biology of P. syringae, as many genes encoding traits important for plant-microbe interactions were thermo-regulated. PMID:23527276

  1. Deciphering Cis-Regulatory Element Mediated Combinatorial Regulation in Rice under Blast Infected Condition.

    PubMed

    Deb, Arindam; Kundu, Sudip

    2015-01-01

    Combinations of cis-regulatory elements (CREs) present at the promoters facilitate the binding of several transcription factors (TFs), thereby altering the consequent gene expressions. Due to the eminent complexity of the regulatory mechanism, the combinatorics of CRE-mediated transcriptional regulation has been elusive. In this work, we have developed a new methodology that quantifies the co-occurrence tendencies of CREs present in a set of promoter sequences; these co-occurrence scores are filtered in three consecutive steps to test their statistical significance; and the significantly co-occurring CRE pairs are presented as networks. These networks of co-occurring CREs are further transformed to derive higher order of regulatory combinatorics. We have further applied this methodology on the differentially up-regulated gene-sets of rice tissues under fungal (Magnaporthe) infected conditions to demonstrate how it helps to understand the CRE-mediated combinatorial gene regulation. Our analysis includes a wide spectrum of biologically important results. The CRE pairs having a strong tendency to co-occur often exhibit very similar joint distribution patterns at the promoters of rice. We couple the network approach with experimental results of plant gene regulation and defense mechanisms and find evidences of auto and cross regulation among TF families, cross-talk among multiple hormone signaling pathways, similarities and dissimilarities in regulatory combinatorics between different tissues, etc. Our analyses have pointed a highly distributed nature of the combinatorial gene regulation facilitating an efficient alteration in response to fungal attack. All together, our proposed methodology could be an important approach in understanding the combinatorial gene regulation. It can be further applied to unravel the tissue and/or condition specific combinatorial gene regulation in other eukaryotic systems with the availability of annotated genomic sequences and suitable

  2. Megakaryocytes regulate expression of Pyk2 isoforms and caspase-mediated cleavage of actin in osteoblasts.

    PubMed

    Kacena, Melissa A; Eleniste, Pierre P; Cheng, Ying-Hua; Huang, Su; Shivanna, Mahesh; Meijome, Tomas E; Mayo, Lindsey D; Bruzzaniti, Angela

    2012-05-18

    The proliferation and differentiation of osteoblast (OB) precursors are essential for elaborating the bone-forming activity of mature OBs. However, the mechanisms regulating OB proliferation and function are largely unknown. We reported that OB proliferation is enhanced by megakaryocytes (MKs) via a process that is regulated in part by integrin signaling. The tyrosine kinase Pyk2 has been shown to regulate cell proliferation and survival in a variety of cells. Pyk2 is also activated by integrin signaling and regulates actin remodeling in bone-resorbing osteoclasts. In this study, we examined the role of Pyk2 and actin in the MK-mediated increase in OB proliferation. Calvarial OBs were cultured in the presence of MKs for various times, and Pyk2 signaling cascades in OBs were examined by Western blotting, subcellular fractionation, and microscopy. We found that MKs regulate the temporal expression of Pyk2 and its subcellular localization. We also found that MKs regulate the expression of two alternatively spliced isoforms of Pyk2 in OBs, which may regulate OB differentiation and proliferation. MKs also induced cytoskeletal reorganization in OBs, which was associated with the caspase-mediated cleavage of actin, an increase in focal adhesions, and the formation of apical membrane ruffles. Moreover, BrdU incorporation in MK-stimulated OBs was blocked by the actin-polymerizing agent, jasplakinolide. Collectively, our studies reveal that Pyk2 and actin play an important role in MK-regulated signaling cascades that control OB proliferation and may be important for therapeutic interventions aimed at increasing bone formation in metabolic diseases of the skeleton. PMID:22447931

  3. Megakaryocytes Regulate Expression of Pyk2 Isoforms and Caspase-mediated Cleavage of Actin in Osteoblasts*

    PubMed Central

    Kacena, Melissa A.; Eleniste, Pierre P.; Cheng, Ying-Hua; Huang, Su; Shivanna, Mahesh; Meijome, Tomas E.; Mayo, Lindsey D.; Bruzzaniti, Angela

    2012-01-01

    The proliferation and differentiation of osteoblast (OB) precursors are essential for elaborating the bone-forming activity of mature OBs. However, the mechanisms regulating OB proliferation and function are largely unknown. We reported that OB proliferation is enhanced by megakaryocytes (MKs) via a process that is regulated in part by integrin signaling. The tyrosine kinase Pyk2 has been shown to regulate cell proliferation and survival in a variety of cells. Pyk2 is also activated by integrin signaling and regulates actin remodeling in bone-resorbing osteoclasts. In this study, we examined the role of Pyk2 and actin in the MK-mediated increase in OB proliferation. Calvarial OBs were cultured in the presence of MKs for various times, and Pyk2 signaling cascades in OBs were examined by Western blotting, subcellular fractionation, and microscopy. We found that MKs regulate the temporal expression of Pyk2 and its subcellular localization. We also found that MKs regulate the expression of two alternatively spliced isoforms of Pyk2 in OBs, which may regulate OB differentiation and proliferation. MKs also induced cytoskeletal reorganization in OBs, which was associated with the caspase-mediated cleavage of actin, an increase in focal adhesions, and the formation of apical membrane ruffles. Moreover, BrdU incorporation in MK-stimulated OBs was blocked by the actin-polymerizing agent, jasplakinolide. Collectively, our studies reveal that Pyk2 and actin play an important role in MK-regulated signaling cascades that control OB proliferation and may be important for therapeutic interventions aimed at increasing bone formation in metabolic diseases of the skeleton. PMID:22447931

  4. Deciphering Cis-Regulatory Element Mediated Combinatorial Regulation in Rice under Blast Infected Condition

    PubMed Central

    Deb, Arindam; Kundu, Sudip

    2015-01-01

    Combinations of cis-regulatory elements (CREs) present at the promoters facilitate the binding of several transcription factors (TFs), thereby altering the consequent gene expressions. Due to the eminent complexity of the regulatory mechanism, the combinatorics of CRE-mediated transcriptional regulation has been elusive. In this work, we have developed a new methodology that quantifies the co-occurrence tendencies of CREs present in a set of promoter sequences; these co-occurrence scores are filtered in three consecutive steps to test their statistical significance; and the significantly co-occurring CRE pairs are presented as networks. These networks of co-occurring CREs are further transformed to derive higher order of regulatory combinatorics. We have further applied this methodology on the differentially up-regulated gene-sets of rice tissues under fungal (Magnaporthe) infected conditions to demonstrate how it helps to understand the CRE-mediated combinatorial gene regulation. Our analysis includes a wide spectrum of biologically important results. The CRE pairs having a strong tendency to co-occur often exhibit very similar joint distribution patterns at the promoters of rice. We couple the network approach with experimental results of plant gene regulation and defense mechanisms and find evidences of auto and cross regulation among TF families, cross-talk among multiple hormone signaling pathways, similarities and dissimilarities in regulatory combinatorics between different tissues, etc. Our analyses have pointed a highly distributed nature of the combinatorial gene regulation facilitating an efficient alteration in response to fungal attack. All together, our proposed methodology could be an important approach in understanding the combinatorial gene regulation. It can be further applied to unravel the tissue and/or condition specific combinatorial gene regulation in other eukaryotic systems with the availability of annotated genomic sequences and suitable

  5. An Arabidopsis mutant with enhanced resistance to powdery mildew.

    PubMed Central

    Frye, C A; Innes, R W

    1998-01-01

    We have identified an Arabidopsis mutant that displays enhanced disease resistance to the fungus Erysiphe cichoracearum, causal agent of powdery mildew. The edr1 mutant does not constitutively express the pathogenesis-related genes PR-1, BGL2, or PR-5 and thus differs from previously described disease-resistant mutants of Arabidopsis. E. cichoracearum conidia (asexual spores) germinated normally and formed extensive hyphae on edr1 plants, indicating that the initial stages of infection were not inhibited. Production of conidiophores on edr1 plants, however, was <16% of that observed on wild-type Arabidopsis. Reduction in sporulation correlated with a more rapid induction of defense responses. Autofluorescent compounds and callose accumulated in edr1 leaves 3 days after inoculation with E. cichoracearum, and dead mesophyll cells accumulated in edr1 leaves starting 5 days after inoculation. Macroscopic patches of dead cells appeared 6 days after inoculation. This resistance phenotype is similar to that conferred by "late-acting" powdery mildew resistance genes of wheat and barley. The edr1 mutation is recessive and maps to chromosome 1 between molecular markers ATEAT1 and NCC1. We speculate that the edr1 mutation derepresses multiple defense responses, making them more easily induced by virulent pathogens. PMID:9634583

  6. Method to characterize dielectric properties of powdery substances

    NASA Astrophysics Data System (ADS)

    Tuhkala, M.; Juuti, J.; Jantunen, H.

    2013-07-01

    An open ended coaxial cavity method for dielectric characterization of powdery substance operating at 4.5 GHz in TEM mode is presented. Classical mixing rules and electromagnetic modeling were utilized with measured effective permittivities and Q factors to determine the relative permittivity and dielectric loss tangent of different powders with ɛr up to 30. The modeling enabled determination of the correction factor for the simplified equation for the relative permittivity of an open ended coaxial resonator and mixing rules having the best correlation with experiments. SiO2, Al2O3, LTCC CT 2000, ZrO2, and La2O3 powders were used in the experiments. Based on the measured properties and Bruggeman symmetric and Looyenga mixing rules, the determined dielectric characteristics of the powders exhibited good correlation with values in the literature. The presented characterization method enabled the determination of dielectric properties of powdery substances within the presented range, and therefore could be applied to various research fields and applications where dielectric properties of powders need to be known and controlled.

  7. The Arabidopsis NRG2 Protein Mediates Nitrate Signaling and Interacts with and Regulates Key Nitrate Regulators[OPEN

    PubMed Central

    Zhao, Lufei; Zhang, Chengfei; Li, Zehui; Lei, Zhao; Liu, Fei; Guan, Peizhu; Crawford, Nigel M.

    2016-01-01

    We show that NITRATE REGULATORY GENE2 (NRG2), which we identified using forward genetics, mediates nitrate signaling in Arabidopsis thaliana. A mutation in NRG2 disrupted the induction of nitrate-responsive genes after nitrate treatment by an ammonium-independent mechanism. The nitrate content in roots was lower in the mutants than in the wild type, which may have resulted from reduced expression of NRT1.1 (also called NPF6.3, encoding a nitrate transporter/receptor) and upregulation of NRT1.8 (also called NPF7.2, encoding a xylem nitrate transporter). Genetic and molecular data suggest that NRG2 functions upstream of NRT1.1 in nitrate signaling. Furthermore, NRG2 directly interacts with the nitrate regulator NLP7 in the nucleus, but nuclear retention of NLP7 in response to nitrate is not dependent on NRG2. Transcriptomic analysis revealed that genes involved in four nitrogen-related clusters including nitrate transport and response to nitrate were differentially expressed in the nrg2 mutants. A nitrogen compound transport cluster containing some members of the NRT/PTR family was regulated by both NRG2 and NRT1.1, while no nitrogen-related clusters showed regulation by both NRG2 and NLP7. Thus, NRG2 plays a key role in nitrate regulation in part through modulating NRT1.1 expression and may function with NLP7 via their physical interaction. PMID:26744214

  8. NEK6-mediated phosphorylation of human TPP1 regulates telomere length through telomerase recruitment.

    PubMed

    Hirai, Yugo; Tamura, Miki; Otani, Junji; Ishikawa, Fuyuki

    2016-08-01

    Shelterin component TPP1 plays critical roles in chromosome end protection and telomere length regulation. Specifically, TPP1 contains an OB-fold domain that provides an interface to recruit telomerase. However, it remains largely unknown how telomerase recruitment is regulated by cell cycle regulators. We show that TPP1 interacts with the cell cycle regulator kinase NEK6 in human cells. We found that NEK6-mediated phosphorylation of TPP1 Ser255 in G2/M phase regulates the association between telomerase activity and TPP1. Furthermore, we found evidence that POT1 negatively regulates TPP1 phosphorylation because the level of Ser255 phosphorylation was elevated when telomeres were elongated by a POT1 mutant lacking its OB-fold domains. Ser255 is located in the intervening region between the telomerase-recruiting OB-fold and the POT1 recruitment domains. Ser255 and the surrounding amino acids are conserved among vertebrates. These observations suggest that a region adjacent to the OB-fold domain of TPP1 is involved in telomere length regulation via telomerase recruitment. PMID:27396482

  9. Transcription Factor-Mediated Regulation of Carboxylesterase Enzymes in Livers of Mice

    PubMed Central

    Zhang, Youcai; Cheng, Xingguo; Aleksunes, Lauren

    2012-01-01

    The induction of drug-metabolizing enzymes by chemicals is one of the major reasons for drug-drug interactions. In the present study, the regulation of mRNA expression of one arylacetamide deacetylase (Aadac) and 11 carboxylesterases (Cess) by 15 microsomal enzyme inducers (MEIs) was examined in livers of male C57BL/6 mice. The data demonstrated that Aadac mRNA expression was suppressed by three aryl hydrocarbon receptor (AhR) ligands, two constitutive androstane receptor (CAR) activators, two pregnane X receptor (PXR) ligands, and one nuclear factor erythroid 2-related factor 2 (Nrf2) activator. Ces1 subfamily mRNA expression was not altered by most of the MEIs, whereas Ces2 subfamily mRNA was readily induced by the activators of CAR, PXR, and Nrf2 but not by peroxisome proliferator-activated receptor α activators. Studies using null mice demonstrated that 1) AhR was required for the 2,3,7,8-tetrachlorodibenzo-p-dioxin–mediated suppression of Aadac and Ces3a; 2) CAR was involved in the 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene–mediated induction of Aadac, Ces2c, Ces2a, and Ces3a; 3) PXR was required for the pregnenolone-16α-carbonitrile–mediated induction of Aadac, Ces2c, and Ces2a; 4) Nrf2 was required for the oltipraz-mediated induction of Ces1g and Ces2c; and 5) PXR was not required for the DEX-mediated suppression of Cess in livers of mice. In conclusion, the present study systematically investigated the regulation of Cess by MEIs in livers of mice and demonstrated that MEIs modulated mRNA expression of mouse hepatic Cess through the activation of AhR, CAR, PXR, and/or Nrf2 transcriptional pathways. PMID:22429928

  10. Regulation of synaptic activity by snapin-mediated endolysosomal transport and sorting

    PubMed Central

    Di Giovanni, Jerome; Sheng, Zu-Hang

    2015-01-01

    Recycling synaptic vesicles (SVs) transit through early endosomal sorting stations, which raises a fundamental question: are SVs sorted toward endolysosomal pathways? Here, we used snapin mutants as tools to assess how endolysosomal sorting and trafficking impact presynaptic activity in wild-type and snapin−/− neurons. Snapin acts as a dynein adaptor that mediates the retrograde transport of late endosomes (LEs) and interacts with dysbindin, a subunit of the endosomal sorting complex BLOC-1. Expressing dynein-binding defective snapin mutants induced SV accumulation at presynaptic terminals, mimicking the snapin−/− phenotype. Conversely, over-expressing snapin reduced SV pool size by enhancing SV trafficking to the endolysosomal pathway. Using a SV-targeted Ca2+ sensor, we demonstrate that snapin–dysbindin interaction regulates SV positional priming through BLOC-1/AP-3-dependent sorting. Our study reveals a bipartite regulation of presynaptic activity by endolysosomal trafficking and sorting: LE transport regulates SV pool size, and BLOC-1/AP-3-dependent sorting fine-tunes the Ca2+ sensitivity of SV release. Therefore, our study provides new mechanistic insights into the maintenance and regulation of SV pool size and synchronized SV fusion through snapin-mediated LE trafficking and endosomal sorting. PMID:26108535

  11. New Regulators of Clathrin-Mediated Endocytosis Identified in Saccharomyces cerevisiae by Systematic Quantitative Fluorescence Microscopy.

    PubMed

    Farrell, Kristen B; Grossman, Caitlin; Di Pietro, Santiago M

    2015-11-01

    Despite the importance of clathrin-mediated endocytosis (CME) for cell biology, it is unclear if all components of the machinery have been discovered and many regulatory aspects remain poorly understood. Here, using Saccharomyces cerevisiae and a fluorescence microscopy screening approach we identify previously unknown regulatory factors of the endocytic machinery. We further studied the top scoring protein identified in the screen, Ubx3, a member of the conserved ubiquitin regulatory X (UBX) protein family. In vivo and in vitro approaches demonstrate that Ubx3 is a new coat component. Ubx3-GFP has typical endocytic coat protein dynamics with a patch lifetime of 45 ± 3 sec. Ubx3 contains a W-box that mediates physical interaction with clathrin and Ubx3-GFP patch lifetime depends on clathrin. Deletion of the UBX3 gene caused defects in the uptake of Lucifer Yellow and the methionine transporter Mup1 demonstrating that Ubx3 is needed for efficient endocytosis. Further, the UBX domain is required both for localization and function of Ubx3 at endocytic sites. Mechanistically, Ubx3 regulates dynamics and patch lifetime of the early arriving protein Ede1 but not later arriving coat proteins or actin assembly. Conversely, Ede1 regulates the patch lifetime of Ubx3. Ubx3 likely regulates CME via the AAA-ATPase Cdc48, a ubiquitin-editing complex. Our results uncovered new components of the CME machinery that regulate this fundamental process. PMID:26362318

  12. Moderate differences in circulating corticosterone alter receptor-mediated regulation of 5-hydroxytryptamine neuronal activity.

    PubMed

    Judge, Sarah J; Ingram, Colin D; Gartside, Sarah E

    2004-12-01

    Circulating glucocorticoid levels vary with stress and psychiatric illness and play a potentially important role in regulating transmitter systems that regulate mood. To determine whether chronic variation in corticosterone levels within the normal diurnal range altered the control of 5-hydroxytryptamine (5-HT) neuronal activity, male rats were adrenalectomized and implanted with either a 2% or 70% corticosterone/cholesterol pellet (100 mg). Two weeks later, the regulation of 5-HT neuronal activity in the dorsal raphe nucleus was studied by in vitro electrophysiology. At this time, serum corticosterone levels approximated the low-point (2%) and mid-point (70%) of the diurnal range. The excitatory response of 5-HT neurones to the alpha1-adrenoceptor agonist phenylephrine (1-11 microM) was significantly greater in the 2% group compared to the 70% group. By contrast, the inhibitory response to 5-HT (10-50 microM) was significantly lower in the 2% group compared to the 70% group. Thus, chronic variation in circulating corticosterone over a narrow part of the normal diurnal range causes a shift in the balance of positive and negative regulation of 5-HT neurones, with increased alpha 1-adrenoceptor-mediated excitation and reduced 5-HT-mediated autoinhibition at lower corticosterone levels. This shift would have a major impact on control of 5-HT neuronal activity. PMID:15582914

  13. CHIP mediates down-regulation of nucleobindin-1 in preosteoblast cell line models.

    PubMed

    Xue, Fuying; Wu, Yanping; Zhao, Xinghui; Zhao, Taoran; Meng, Ying; Zhao, Zhanzhong; Guo, Junwei; Chen, Wei

    2016-08-01

    Nucleobindin-1 (NUCB1), also known as Calnuc, is a highly conserved, multifunctional protein widely expressed in tissues and cells. It contains two EF-hand motifs which have been shown to play a crucial role in binding Ca(2+) ions. In this study, we applied comparative two-dimensional gel electrophoresis to characterize differentially expressed proteins in HA-CHIP over-expressed and endogenous CHIP depleted MC3T3-E1 stable cell lines, identifying NUCB1 as a novel CHIP/Stub1 targeted protein. NUCB1 interacts with and is down-regulated by CHIP by both proteasomal dependent and independent pathways, suggesting that CHIP-mediated down-regulation of nucleobindin-1 might play a role in osteoblast differentiation. The chaperone protein Hsp70 was found to be important for CHIP and NUCB1 interaction as well as CHIP-mediated NUCB1 down-regulation. Our findings provide new insights into understanding the stability regulation of NUCB1. PMID:27178152

  14. New concept for determination and denomination of pathotypes and races of cucurbit powdery mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cucurbit powdery mildew (CPM), which is caused most frequently by two obligate erysiphaceous ectoparasites (Golovinomyces cichoracearum, Podosphaera xanthii), occurs on field and glasshouse cucurbit crops worldwide. We review the current state of understanding of variation for pathogenicity of these...

  15. Differential Regulation of White-Opaque Switching by Individual Subunits of Candida albicans Mediator

    PubMed Central

    Zhang, Anda; Liu, Zhongle

    2013-01-01

    The multisubunit eukaryotic Mediator complex integrates diverse positive and negative gene regulatory signals and transmits them to the core transcription machinery. Mutations in individual subunits within the complex can lead to decreased or increased transcription of certain subsets of genes, which are highly specific to the mutated subunit. Recent studies suggest a role for Mediator in epigenetic silencing. Using white-opaque morphological switching in Candida albicans as a model, we have shown that Mediator is required for the stability of both the epigenetic silenced (white) and active (opaque) states of the bistable transcription circuit driven by the master regulator Wor1. Individual deletions of eight C. albicans Mediator subunits have shown that different Mediator subunits have dramatically diverse effects on the directionality, frequency, and environmental induction of epigenetic switching. Among the Mediator deletion mutants analyzed, only Med12 has a steady-state transcriptional effect on the components of the Wor1 circuit that clearly corresponds to its effect on switching. The MED16 and MED9 genes have been found to be among a small subset of genes that are required for the stability of both the white and opaque states. Deletion of the Med3 subunit completely destabilizes the opaque state, even though the Wor1 transcription circuit is intact and can be driven by ectopic expression of Wor1. The highly impaired ability of the med3 deletion mutant to mate, even when Wor1 expression is ectopically induced, reveals that the activation of the Wor1 circuit can be decoupled from the opaque state and one of its primary biological consequences. PMID:23873866

  16. Transcriptional regulation of lycopene metabolism mediated by rootstock during the ripening of grafted watermelons.

    PubMed

    Kong, Qiusheng; Yuan, Jingxian; Gao, Lingyun; Liu, Peng; Cao, Lei; Huang, Yuan; Zhao, Liqiang; Lv, Huifang; Bie, Zhilong

    2017-01-01

    Rootstocks have comprehensive effects on lycopene accumulation in grafted watermelon fruits. However, little is known about lycopene metabolic regulation in grafted watermelon. To address this problem, parallel changes in lycopene contents and the expression of its metabolic genes were analyzed during the fruit ripening of nongrafted watermelon and watermelon grafted onto bottle gourd, pumpkin, and wild watermelon. Results showed that rootstocks mediated the transcriptional regulations of lycopene accumulation in different ways. Bottle gourd and wild watermelon promoted lycopene accumulation in grafted watermelon fruits by upregulating the biosynthetic genes phytoene synthase (PSY) and ζ-carotene desaturase (ZDS), and downregulating the catabolic genes β-carotene hydroxylase (CHYB), zeaxanthin epoxidase (ZEP), 9-cis-epoxycarotenoid dioxygenase (NCED), and carotenoid cleavage dioxygenase (CCD). However, pumpkin did not affect lycopene accumulation by upregulating both biosynthetic and catabolic genes. The rootstock-dependent characteristic of lycopene accumulation in grafted watermelon fruits provided an alternative model for investigating lycopene metabolic regulation. PMID:27507492

  17. FBXL5 targets cortactin for ubiquitination-mediated destruction to regulate gastric cancer cell migration.

    PubMed

    Cen, Gang; Ding, Hong-Hua; Liu, Bin; Wu, Wei-Dong

    2014-09-01

    Cortactin, an actin-interacting protein, is implicated in cytoskeletal architecture and often amplified in several types of cancer including gastric adenocarcinomas. Downregulation of cortactin decreases cell migration and invasion. However, how to regulate cortactin in gastric cancer remains largely unknown. Here, we report that FBXL5 interacts with and targets cortactin for ubiquitylation and subsequent proteasomal degradation. Furthermore, we showed that FBXL5-induced cortactin degradation is mediated by extracellular regulated signal kinase (ERK). Serine phosphorylation sites mutant, cortactinS405A/S418A, prevent FBXL5-induced cortactin degradation. Moreover, CortactinS405A/S418A exhibited stronger effects in promoting gastric cancer cell migration when compared to wild-type cortactin. Taken together, our data suggested a novel molecular mechanism for the negative regulation of cortactin by FBXL5 in gastric cancer cells migration. PMID:24867096

  18. Decoding the noncoding: Prospective of lncRNA-mediated innate immune regulation

    PubMed Central

    Li, Zhonghan; Rana, Tariq M

    2014-01-01

    The innate immune system is the first line of defense against microbial pathogens, but tight regulation of gene expression is necessary to prevent the detrimental effects of unrestrained activation. Although the functions of most long noncoding RNAs (lncRNAs; >200 nucleotides) are unknown, many have been shown to regulate diverse cellular activities. Recent reports by us and others have suggested that lncRNAs may also play critical roles in transcriptional regulation of gene expression during innate immune responses. Following engagement of Toll-like receptors, lncRNAs form functional RNA–protein complexes that recruit activators or remove repressors of transcription, leading to rapid expression of inflammatory mediators. These discoveries suggest that lncRNAs may contribute to the gene regulatory networks that govern host–pathogen interactions. PMID:25482890

  19. Parent Emotion Socialization Practices and Child Self-regulation as Predictors of Child Anxiety: The Mediating Role of Cardiac Variability.

    PubMed

    Williams, Sarah R; Woodruff-Borden, Janet

    2015-08-01

    The importance of the parent-child relationship in emotional development is well supported. The parental role of facilitating a child's self-regulation may provide a more focused approach for examining the role of parenting in child anxiety. The current study hypothesized that parent emotion socialization practices would predict a child's abilities in self-regulation. Given that physiological arousal has been implicated in emotional development, this was hypothesized to mediate the relationship between parental emotion socialization and child emotion regulation to predict child anxiety. Eighty-five parent and child dyads participated in the study. Parents reporting higher degrees of unsupportive emotion socialization were more likely to have children with fewer abilities in emotion regulation. Cardiac responsiveness mediated the relationship between unsupportive emotion socialization and child emotion regulation. The model of cardiac responsiveness mediating the relationship between unsupportive emotion socialization and child emotion regulation failed to reach statistical significance in predicting child anxiety symptoms. PMID:25204571

  20. MicroRNA-30b-Mediated Regulation of Catalase Expression in Human ARPE-19 Cells

    PubMed Central

    Haque, Rashidul; Chun, Eugene; Howell, Jennifer C.; Sengupta, Trisha; Chen, Dan; Kim, Hana

    2012-01-01

    Background Oxidative injury to retinal pigment epithelium (RPE) and retinal photoreceptors has been linked to a number of retinal diseases, including age-related macular degeneration (AMD). Reactive oxygen species (ROS)-mediated gene expression has been extensively studied at transcriptional levels. Also, the post-transcriptional control of gene expression at the level of translational regulation has been recently reported. However, the microRNA (miRNA/miR)-mediated post-transcriptional regulation in human RPE cells has not been thoroughly looked at. Increasing evidence points to a potential role of miRNAs in diverse physiological processes. Methodology/Principal Findings We demonstrated for the first time in a human retinal pigment epithelial cell line (ARPE-19) that the post-transcriptional control of gene expression via miRNA modulation regulates human catalase, an important and potent component of cell's antioxidant defensive network, which detoxifies hydrogen peroxide (H2O2) radicals. Exposure to several stress-inducing agents including H2O2 has been reported to alter miRNA expression profile. Here, we demonstrated that a sublethal dose of H2O2 (200 µM) up-regulated the expression of miR-30b, a member of the miR-30 family, which inhibited the expression of endogenous catalase both at the transcript and protein levels. However, antisense (antagomirs) of miR-30b was not only found to suppress the miR-30b mimics-mediated inhibitions, but also to dramatically increase the expression of catalase even under an oxidant environment. Conclusions/Significance We propose that a microRNA antisense approach could enhance cytoprotective mechanisms against oxidative stress by increasing the antioxidant defense system. PMID:22880027

  1. Regulation of Noxa-mediated apoptosis in Helicobacter pylori–infected gastric epithelial cells

    PubMed Central

    Rath, Suvasmita; Das, Lopamudra; Kokate, Shrikant Babanrao; Pratheek, B. M.; Chattopadhyay, Subhasis; Goswami, Chandan; Chattopadhyay, Ranajoy; Crowe, Sheila Eileen; Bhattacharyya, Asima

    2015-01-01

    Helicobacter pylori induces the antiapoptotic protein myeloid cell leukemia 1 (Mcl1) in human gastric epithelial cells (GECs). Apoptosis of oncogenic protein Mcl1-expressing cells is mainly regulated by Noxa-mediated degradation of Mcl1. We wanted to elucidate the status of Noxa in H. pylori–infected GECs. For this, various GECs such as AGS, MKN45, and KATO III were either infected with H. pylori or left uninfected. The effect of infection was examined by immunoblotting, immunoprecipitation, chromatin immunoprecipitation assay, in vitro binding assay, flow cytometry, and confocal microscopy. Infected GECs, surgical samples collected from patients with gastric adenocarcinoma as well as biopsy samples from patients infected with H. pylori showed significant up-regulation of both Mcl1 and Noxa compared with noninfected samples. Coexistence of Mcl1 and Noxa was indicative of an impaired Mcl-Noxa interaction. We proved that Noxa was phosphorylated at Ser13 residue by JNK in infected GECs, which caused cytoplasmic retention of Noxa. JNK inhibition enhanced Mcl1-Noxa interaction in the mitochondrial fraction of infected cells, whereas overexpression of nonphosphorylatable Noxa resulted in enhanced mitochondria-mediated apoptosis in the infected epithelium. Because phosphorylation-dephosphorylation can regulate the apoptotic function of Noxa, this could be a potential target molecule for future treatment approaches for H. pylori–induced gastric cancer.—Rath, S., Das, L., Kokate, S. B., Pratheek, B. M., Chattopadhyay, S., Goswami, C., Chattopadhyay, R., Crowe, S. E., Bhattacharyya, A. Regulation of Noxa-mediated apoptosis in Helicobacter pylori–infected gastric epithelial cells. PMID:25404713

  2. Epigenetic and Glucocorticoid Receptor-Mediated Regulation of Glutathione Peroxidase 3 in Lung Cancer Cells

    PubMed Central

    An, Byung Chull; Jung, Nak-Kyun; Park, Chun Young; Oh, In-Jae; Choi, Yoo-Duk; Park, Jae-Il; Lee, Seung-won

    2016-01-01

    Glutathione peroxidase 3 (GPx3), an antioxidant enzyme, acts as a modulator of redox signaling, has immunomodulatory function, and catalyzes the detoxification of reactive oxygen species (ROS). GPx3 has been identified as a tumor suppressor in many cancers. Although hyper-methylation of the GPx3 promoter has been shown to down-regulate its expression, other mechanisms by which GPx3 expression is regulated have not been reported. The aim of this study was to further elucidate the mechanisms of GPx3 regulation. GPx3 gene analysis predicted the presence of ten glucocorticoid response elements (GREs) on the GPx3 gene. This result prompted us to investigate whether GPx3 expression is regulated by the glucocorticoid receptor (GR), which is implicated in tumor response to chemotherapy. The corticosteroid dexamethasone (Dex) was used to examine the possible relationship between GR and GPx3 expression. Dex significantly induced GPx3 expression in H1299, H1650, and H1975 cell lines, which exhibit low levels of GPx3 expression under normal conditions. The results of EMSA and ChIP-PCR suggest that GR binds directly to GRE 6 and 7, both of which are located near the GPx3 promoter. Assessment of GPx3 transcription efficiency using a luciferase reporter system showed that blocking formation of the GR-GRE complexes reduced luciferase activity by 7–8-fold. Suppression of GR expression by siRNA transfection also induced down-regulation of GPx3. These data indicate that GPx3 expression can be regulated independently via epigenetic or GR-mediated mechanisms in lung cancer cells, and suggest that GPx3 could potentiate glucocorticoid (GC)-mediated anti-inflammatory signaling in lung cancer cells. PMID:27484907

  3. Epigenetic and Glucocorticoid Receptor-Mediated Regulation of Glutathione Peroxidase 3 in Lung Cancer Cells.

    PubMed

    An, Byung Chull; Jung, Nak-Kyun; Park, Chun Young; Oh, In-Jae; Choi, Yoo-Duk; Park, Jae-Il; Lee, Seung-Won

    2016-08-31

    Glutathione peroxidase 3 (GPx3), an antioxidant enzyme, acts as a modulator of redox signaling, has immunomodulatory function, and catalyzes the detoxification of reactive oxygen species (ROS). GPx3 has been identified as a tumor suppressor in many cancers. Although hyper-methylation of the GPx3 promoter has been shown to down-regulate its expression, other mechanisms by which GPx3 expression is regulated have not been reported. The aim of this study was to further elucidate the mechanisms of GPx3 regulation. GPx3 gene analysis predicted the presence of ten glucocorticoid response elements (GREs) on the GPx3 gene. This result prompted us to investigate whether GPx3 expression is regulated by the glucocorticoid receptor (GR), which is implicated in tumor response to chemotherapy. The corticosteroid dexamethasone (Dex) was used to examine the possible relationship between GR and GPx3 expression. Dex significantly induced GPx3 expression in H1299, H1650, and H1975 cell lines, which exhibit low levels of GPx3 expression under normal conditions. The results of EMSA and ChIP-PCR suggest that GR binds directly to GRE 6 and 7, both of which are located near the GPx3 promoter. Assessment of GPx3 transcription efficiency using a luciferase reporter system showed that blocking formation of the GR-GRE complexes reduced luciferase activity by 7-8-fold. Suppression of GR expression by siRNA transfection also induced down-regulation of GPx3. These data indicate that GPx3 expression can be regulated independently via epigenetic or GR-mediated mechanisms in lung cancer cells, and suggest that GPx3 could potentiate glucocorticoid (GC)-mediated anti-inflammatory signaling in lung cancer cells. PMID:27484907

  4. Tetraspanin CD151 Is a Negative Regulator of FcεRI-Mediated Mast Cell Activation

    PubMed Central

    Abdala-Valencia, Hiam; Bryce, Paul J.; Schleimer, Robert P.; Wechsler, Joshua B.; Loffredo, Lucas F.; Cook-Mills, Joan M.; Hsu, Chia-Lin; Berdnikovs, Sergejs

    2016-01-01

    Mast cells are critical in the pathogenesis of allergic disease due to the release of preformed and newly synthesized mediators, yet the mechanisms controlling mast cell activation are not well understood. Members of the tetraspanin family are recently emerging as modulators of FcεRI-mediated mast cell activation; however, mechanistic understanding of their function is currently lacking. The tetraspanin CD151 is a poorly understood member of this family and is specifically induced on mouse and human mast cells upon FcεRI aggregation but its functional effects are unknown. In this study, we show that CD151 deficiency significantly exacerbates the IgE-mediated late phase inflammation in a murine model of passive cutaneous anaphylaxis. Ex vivo, FcεRI stimulation of bone marrow–derived mast cells from CD151−/− mice resulted in significantly enhanced expression of proinflammatory cytokines IL-4, IL-13, and TNF-α compared with wild-type controls. However, FcεRI -induced mast cell degranulation was unaffected. At the molecular signaling level, CD151 selectively regulated IgE-induced activation of ERK1/2 and PI3K, associated with cytokine production, but had no effect on the phospholipase Cγ1 signaling, associated with degranulation. Collectively, our data indicate that CD151 exerts negative regulation over IgE-induced late phase responses and cytokine production in mast cells. PMID:26136426

  5. Tetraspanin CD151 Is a Negative Regulator of FcεRI-Mediated Mast Cell Activation.

    PubMed

    Abdala-Valencia, Hiam; Bryce, Paul J; Schleimer, Robert P; Wechsler, Joshua B; Loffredo, Lucas F; Cook-Mills, Joan M; Hsu, Chia-Lin; Berdnikovs, Sergejs

    2015-08-15

    Mast cells are critical in the pathogenesis of allergic disease due to the release of preformed and newly synthesized mediators, yet the mechanisms controlling mast cell activation are not well understood. Members of the tetraspanin family are recently emerging as modulators of FcεRI-mediated mast cell activation; however, mechanistic understanding of their function is currently lacking. The tetraspanin CD151 is a poorly understood member of this family and is specifically induced on mouse and human mast cells upon FcεRI aggregation but its functional effects are unknown. In this study, we show that CD151 deficiency significantly exacerbates the IgE-mediated late phase inflammation in a murine model of passive cutaneous anaphylaxis. Ex vivo, FcεRI stimulation of bone marrow-derived mast cells from CD151(-/-) mice resulted in significantly enhanced expression of proinflammatory cytokines IL-4, IL-13, and TNF-α compared with wild-type controls. However, FcεRI-induced mast cell degranulation was unaffected. At the molecular signaling level, CD151 selectively regulated IgE-induced activation of ERK1/2 and PI3K, associated with cytokine production, but had no effect on the phospholipase Cγ1 signaling, associated with degranulation. Collectively, our data indicate that CD151 exerts negative regulation over IgE-induced late phase responses and cytokine production in mast cells. PMID:26136426

  6. Down-regulation of lipoprotein lipase increases ABCA1-mediated cholesterol efflux in THP-1 macrophages.

    PubMed

    Kawashima, Ryoko L; Medh, Jheem D

    2014-08-01

    The ATP-binding cassette transporter A1 (ABCA1) mediates the efflux of excess cholesterol from foam cells to lipid-poor apolipoprotein A-I, in a process called reverse cholesterol transport. Lipoprotein lipase (LPL) is a lipolytic enzyme expressed by macrophages within atherosclerotic lesions. Lentivirus-mediated RNA interference was used to genetically knock-down (KD) the expression of LPL in THP-1 macrophages. Silencing of the LPL gene was confirmed by end-point PCR, real time PCR, and protein analysis. Suppression of LPL expression correlated with a 1.6-fold up-regulation of ABCA1 mRNA levels, and resulted in a 4.5-fold increase in ABCA1-dependent cholesterol efflux. Replenishing LPL by addition of purified bovine LPL to the cell culture media resulted in down-regulation of ABCA1-mediated cholesterol efflux in both wild-type and LPL knockdown cells. These findings suggest an inverse correlation between macrophage LPL levels and ABCA1 cholesterol transport activity. PMID:25017912

  7. Substrate Stiffness Regulates Proinflammatory Mediator Production through TLR4 Activity in Macrophages.

    PubMed

    Previtera, Michelle L; Sengupta, Amitabha

    2015-01-01

    Clinical data show that disease adversely affects tissue elasticity or stiffness. While macrophage activity plays a critical role in driving disease pathology, there are limited data available on the effects of tissue stiffness on macrophage activity. In this study, the effects of substrate stiffness on inflammatory mediator production by macrophages were investigated. Bone marrow-derived macrophages were grown on polyacrylamide gels that mimicked the stiffness of a variety of soft biological tissues. Overall, macrophages grown on soft substrates produced less proinflammatory mediators than macrophages grown on stiff substrates when the endotoxin LPS was added to media. In addition, the pathways involved in stiffness-regulated proinflammation were investigated. The TLR4 signaling pathway was examined by evaluating TLR4, p-NF-κB p65, MyD88, and p-IκBα expression as well as p-NF-κB p65 translocation. Expression and translocation of the various signaling molecules were higher in macrophages grown on stiff substrates than on soft substrates. Furthermore, TLR4 knockout experiments showed that TLR4 activity enhanced proinflammation on stiff substrates. In conclusion, these results suggest that proinflammatory mediator production initiated by TLR4 is mechanically regulated in macrophages. PMID:26710072

  8. Substrate Stiffness Regulates Proinflammatory Mediator Production through TLR4 Activity in Macrophages

    PubMed Central

    Previtera, Michelle L.; Sengupta, Amitabha

    2015-01-01

    Clinical data show that disease adversely affects tissue elasticity or stiffness. While macrophage activity plays a critical role in driving disease pathology, there are limited data available on the effects of tissue stiffness on macrophage activity. In this study, the effects of substrate stiffness on inflammatory mediator production by macrophages were investigated. Bone marrow–derived macrophages were grown on polyacrylamide gels that mimicked the stiffness of a variety of soft biological tissues. Overall, macrophages grown on soft substrates produced less proinflammatory mediators than macrophages grown on stiff substrates when the endotoxin LPS was added to media. In addition, the pathways involved in stiffness–regulated proinflammation were investigated. The TLR4 signaling pathway was examined by evaluating TLR4, p–NF–κB p65, MyD88, and p–IκBα expression as well as p–NF–κB p65 translocation. Expression and translocation of the various signaling molecules were higher in macrophages grown on stiff substrates than on soft substrates. Furthermore, TLR4 knockout experiments showed that TLR4 activity enhanced proinflammation on stiff substrates. In conclusion, these results suggest that proinflammatory mediator production initiated by TLR4 is mechanically regulated in macrophages. PMID:26710072

  9. Regulation of retinoid mediated cholesterol efflux involves liver X receptor activation in mouse macrophages.

    PubMed

    Manna, Pulak R; Sennoune, Souad R; Martinez-Zaguilan, Raul; Slominski, Andrzej T; Pruitt, Kevin

    2015-08-14

    Removal of cholesterol from macrophage-derived foam cells is a critical step to the prevention of atherosclerotic lesions. We have recently demonstrated the functional importance of retinoids in the regulation of the steroidogenic acute regulatory (StAR) protein that predominantly mediates the intramitochondrial transport of cholesterol in target tissues. In the present study, treatment of mouse macrophages with retinoids, particularly all-trans retinoic acid (atRA) and 9-cis RA, resulted in increases in cholesterol efflux to apolipoprotein AI (Apo-A1). Activation of the PKA pathway by a cAMP analog, (Bu)2cAMP, markedly augmented retinoid mediated cholesterol efflux. Macrophages overexpressing hormone-sensitive lipase increased the hydrolysis of cholesteryl esters and concomitantly enhanced the efficacy of retinoic acid receptor and liver X receptor (LXR) ligands on StAR and ATP-binding cassette transporter A1 (ABCA1) protein levels. RAs elevated StAR promoter activity in macrophages, and an increase in StAR levels augmented cholesterol efflux to Apo-A1, suggesting retinoid-mediated efflux of cholesterol involves enhanced oxysterol production. Further studies revealed that retinoids activate the LXR regulated genes, sterol receptor-element binding protein-1c and ABCA1. These findings provide insights into the regulatory events in which retinoid signaling effectively enhances macrophage cholesterol efflux and indicate that retinoid therapy may have important implications in limiting and/or regressing atherosclerotic cardiovascular disease. PMID:26119689

  10. Agrin regulates CLASP2-mediated capture of microtubules at the neuromuscular junction synaptic membrane

    PubMed Central

    Schmidt, Nadine; Basu, Sreya; Sladecek, Stefan; Gatti, Sabrina; van Haren, Jeffrey; Treves, Susan; Pielage, Jan

    2012-01-01

    Agrin is the major factor mediating the neuronal regulation of postsynaptic structures at the vertebrate neuromuscular junction, but the details of how it orchestrates this unique three-dimensional structure remain unknown. Here, we show that agrin induces the formation of the dense network of microtubules in the subsynaptic cytoplasm and that this, in turn, regulates acetylcholine receptor insertion into the postsynaptic membrane. Agrin acted in part by locally activating phosphatidylinositol 3-kinase and inactivating GSK3β, which led to the local capturing of dynamic microtubules at agrin-induced acetylcholine receptor (AChR) clusters, mediated to a large extent by the microtubule plus-end tracking proteins CLASP2 and CLIP-170. Indeed, in the absence of CLASP2, microtubule plus ends at the subsynaptic muscle membrane, the density of synaptic AChRs, the size of AChR clusters, and the numbers of subsynaptic muscle nuclei with their selective gene expression programs were all reduced. Thus, the cascade linking agrin to CLASP2-mediated microtubule capturing at the synaptic membrane is essential for the maintenance of a normal neuromuscular phenotype. PMID:22851317

  11. Regnase-1 in microglia negatively regulates high mobility group box 1-mediated inflammation and neuronal injury

    PubMed Central

    Liu, Xiao-Xi; Wang, Chen; Huang, Shao-Fei; Chen, Qiong; Hu, Ya-Fang; Zhou, Liang; Gu, Yong

    2016-01-01

    Extracellular high mobility group box 1 (HMGB1) has been demonstrated to function as a proinflammatory cytokine and induces neuronal injury in response to various pathological stimuli in central nervous system (CNS). However, the regulatory factor involved in HMGB1-mediated inflammatory signaling is largely unclear. Regulatory RNase 1 (Regnase-1) is a potent anti-inflammation enzyme that can degrade a set of mRNAs encoding proinflammatory cytokines. The present study aims to determine the role of Regnase-1 in the regulation of HMGB1-mediated inflammatory injury in CNS. Cultured microglia and rat brain were treated with recombinant HMGB1 to examine the induction of Regnase-1 expression. Moreover, the role of Regnase-1 in modulating the expression of inflammatory cytokines and neuronal injury was then investigated in microglia by specific siRNA knockdown upon HMGB1 treatment. Results showed that HMGB1 could significantly induce the de novo synthesis of Regnase-1 in cultured microglia. Consistently, Regnase-1 was elevated and found to be co-localized with microglia marker in the brain of rat treated with HMGB1. Silencing Regnase-1 in microglia enhanced HMGB1-induced expression of proinflammatory cytokines and exacerbated neuronal toxicity. Collectively, these results suggest that Regnase-1 can be induced by HMGB1 in microglia and negatively regulates HMGB1-mediated neuroinflammation and neuronal toxicity. PMID:27044405

  12. β-catenin mediates behavioral resilience through Dicer1/microRNA regulation

    PubMed Central

    Dias, Caroline; Feng, Jian; Sun, Haosheng; Shao, Ning-yi; Mazei-Robison, Michelle S.; Damez-Werno, Diane; Scobie, Kimberly; Bagot, Rosemary; LaBonte, Benoit; Ribeiro, Efrain; Liu, XiaoChuan; Kennedy, Pamela; Vialou, Vincent; Ferguson, Deveroux; Pena, Catherine; Calipari, Erin; Koo, Jawook; Mouzon, Ezekiell; Ghose, Subruto; Tamminga, Carol; Neve, Rachael; Shen, Li

    2014-01-01

    β-catenin is a multi-functional protein that plays an important role in the mature central nervous system; its dysfunction has been implicated in several neuropsychiatric disorders, including depression. Here we show that β-catenin mediates pro-resilient and anxiolytic effects in mice in the nucleus accumbens, a key brain reward region, an effect mediated by D2-type medium spiny neurons. Using genome-wide β-catenin enrichment mapping, we identify Dicer1—important in small RNA (e.g., microRNA) biogenesis—as a β-catenin target gene that mediates resilience. Small RNA profiling after excising β-catenin from nucleus accumbens in the context of chronic stress reveals β-catenin-dependent microRNA regulation associated with resilience. Together, these findings establish β-catenin as a critical regulator in the development of behavioral resilience, activating a network that includes Dicer1 and downstream microRNAs. We thus present a foundation for the development of novel therapeutic targets to promote stress resilience. PMID:25383518

  13. Cathepsin-B-mediated cleavage of Disabled-2 regulates TGF-β-induced autophagy.

    PubMed

    Jiang, Yong; Woosley, Alec N; Sivalingam, Nageswaran; Natarajan, Sneha; Howe, Philip H

    2016-08-01

    Transforming growth factor-β (TGF-β) induces the expression of Disabled-2 (Dab2), an endocytic adaptor and tumour suppressor, concomitant with the induction of an epithelial-mesenchymal transition (EMT) in mammary epithelial cells. Here we show that following TGF-β-mediated EMT, sustained TGF-β treatment leads to proteolytic degradation of Dab2 by cathepsin B (CTSB), loss of the mesenchymal phenotype and induction of autophagy. CTSB inhibition or expression of a CTSB-resistant Dab2 mutant maintains Dab2 expression and shifts long-term TGF-β-treated cells from autophagy to apoptosis. We further show that Dab2 interacts with Beclin-1 to promote casein-kinase-2-mediated phosphorylation of Beclin-1, preventing Beclin-1-Vps34 interaction and subsequent autophagosome assembly. Thus, CTSB-mediated degradation of Dab2 allows Beclin-1-Vps34 induction of autophagy, whereas sustained Dab2 expression prevents autophagy and promotes apoptosis by stabilizing the pro-apoptotic Bim protein. In vivo studies suggest that Dab2-mediated regulation of autophagy modulates chemotherapeutic resistance and tumour metastasis. PMID:27398911

  14. Emotional Regulation and Depression: A Potential Mediator between Heart and Mind.

    PubMed

    Compare, Angelo; Zarbo, Cristina; Shonin, Edo; Van Gordon, William; Marconi, Chiara

    2014-01-01

    A narrative review of the major evidence concerning the relationship between emotional regulation and depression was conducted. The literature demonstrates a mediating role of emotional regulation in the development of depression and physical illness. Literature suggests in fact that the employment of adaptive emotional regulation strategies (e.g., reappraisal) causes a reduction of stress-elicited emotions leading to physical disorders. Conversely, dysfunctional emotional regulation strategies and, in particular, rumination and emotion suppression appear to be influential in the pathogenesis of depression and physiological disease. More specifically, the evidence suggests that depression and rumination affect both cognitive (e.g., impaired ability to process negative information) and neurobiological mechanisms (e.g., hypothalamic pituitary adrenal axis overactivation and higher rates of cortisol production). Understanding the factors that govern the variety of health outcomes that different people experience following exposure to stress has important implications for the development of effective emotion-regulation interventional approaches (e.g., mindfulness-based therapy, emotion-focused therapy, and emotion regulation therapy). PMID:25050177

  15. Emotional Regulation and Depression: A Potential Mediator between Heart and Mind

    PubMed Central

    Van Gordon, William

    2014-01-01

    A narrative review of the major evidence concerning the relationship between emotional regulation and depression was conducted. The literature demonstrates a mediating role of emotional regulation in the development of depression and physical illness. Literature suggests in fact that the employment of adaptive emotional regulation strategies (e.g., reappraisal) causes a reduction of stress-elicited emotions leading to physical disorders. Conversely, dysfunctional emotional regulation strategies and, in particular, rumination and emotion suppression appear to be influential in the pathogenesis of depression and physiological disease. More specifically, the evidence suggests that depression and rumination affect both cognitive (e.g., impaired ability to process negative information) and neurobiological mechanisms (e.g., hypothalamic pituitary adrenal axis overactivation and higher rates of cortisol production). Understanding the factors that govern the variety of health outcomes that different people experience following exposure to stress has important implications for the development of effective emotion-regulation interventional approaches (e.g., mindfulness-based therapy, emotion-focused therapy, and emotion regulation therapy). PMID:25050177

  16. TRIM13 Is a Negative Regulator of MDA5-Mediated Type I Interferon Production

    PubMed Central

    Narayan, Kavitha; Waggoner, Lisa; Pham, Serena T.; Hendricks, Gabriel L.; Waggoner, Stephen N.; Conlon, Joseph; Wang, Jennifer P.

    2014-01-01

    ABSTRACT Retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA5) are essential intracellular detectors of viral RNA. They contribute to the type I interferon (IFN) response that is crucial for host defense against viral infections. Given the potent antiviral and proinflammatory activities elicited by the type I IFNs, induction of the type I IFN response is tightly regulated. Members of the tripartite motif (TRIM) family of proteins have recently emerged as key regulators of antiviral immunity. We show that TRIM13, an E3 ubiquitin ligase, is expressed in immune cells and is upregulated in bone marrow-derived macrophages upon stimulation with inducers of type I IFN. TRIM13 interacts with MDA5 and negatively regulates MDA5-mediated type I IFN production in vitro, acting upstream of IFN regulatory factor 3. We generated Trim13−/− mice and show that upon lethal challenge with encephalomyocarditis virus (EMCV), which is sensed by MDA5, Trim13−/− mice produce increased amounts of type I IFNs and survive longer than wild-type mice. Trim13−/− murine embryonic fibroblasts (MEFs) challenged with EMCV or poly(I·C) also show a significant increase in beta IFN (IFN-β) levels, but, in contrast, IFN-β responses to the RIG-I-detected Sendai virus were diminished, suggesting that TRIM13 may play a role in positively regulating RIG-I function. Together, these results demonstrate that TRIM13 regulates the type I IFN response through inhibition of MDA5 activity and that it functions nonredundantly to modulate MDA5 during EMCV infection. IMPORTANCE The type I interferon (IFN) response is crucial for host defense against viral infections, and proper regulation of this pathway contributes to maintaining immune homeostasis. Retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA5) are intracellular detectors of viral RNA that induce the type I IFN response. In this study, we show that expression of the

  17. STEM CELL AGING. A mitochondrial UPR-mediated metabolic checkpoint regulates hematopoietic stem cell aging

    PubMed Central

    Mohrin, Mary; Shin, Jiyung; Liu, Yufei; Brown, Katharine; Luo, Hanzhi; Xi, Yannan; Haynes, Cole M.; Chen, Danica

    2015-01-01

    Deterioration of adult stem cells accounts for much of aging-associated compromised tissue maintenance. How stem cells maintain metabolic homeostasis remains elusive. Here, we identified a regulatory branch of the mitochondrial unfolded protein response (UPRmt), which is mediated by the interplay of SIRT7 and NRF1 and is coupled to cellular energy metabolism and proliferation. SIRT7 inactivation caused reduced quiescence, increased mitochondrial protein folding stress (PFSmt), and compromised regenerative capacity of hematopoietic stem cells (HSCs). SIRT7 expression was reduced in aged HSCs, and SIRT7 up-regulation improved the regenerative capacity of aged HSCs. These findings define the deregulation of a UPRmt-mediated metabolic checkpoint as a reversible contributing factor for HSC aging. PMID:25792330

  18. ARMS/Kidins220 and synembryn-B levels regulate NGF-mediated secretion.

    PubMed

    López-Benito, Saray; Lillo, Concepción; Hernández-Hernández, Ángel; Chao, Moses V; Arévalo, Juan C

    2016-05-01

    Proper development of the nervous system requires a temporally and spatially orchestrated set of events including differentiation, synapse formation and neurotransmission. Nerve growth factor (NGF) acting through the TrkA neurotrophin receptor (also known as NTRK1) regulates many of these events. However, the molecular mechanisms responsible for NGF-regulated secretion are not completely understood. Here, we describe a new signaling pathway involving TrkA, ARMS (also known as Kidins220), synembryn-B and Rac1 in NGF-mediated secretion in PC12 cells. Whereas overexpression of ARMS blocked NGF-mediated secretion, without affecting basal secretion, a decrease in ARMS resulted in potentiation. Similar effects were observed with synembryn-B, a protein that interacts directly with ARMS. Downstream of ARMS and synembryn-B are Gαq and Trio proteins, which modulate the activity of Rac1 in response to NGF. Expression of dominant-negative Rac1 rescued the secretion defects of cells overexpressing ARMS or synembryn-B. Thus, this neurotrophin pathway represents a new mechanism responsible for NGF-regulated secretion. PMID:26966186

  19. Regulation of Transcription Factor Yin Yang 1 by SET7/9-mediated Lysine Methylation

    PubMed Central

    Zhang, Wen-juan; Wu, Xiao-nan; Shi, Tao-tao; Xu, Huan-teng; Yi, Jia; Shen, Hai-feng; Huang, Ming-feng; Shu, Xing-yi; Wang, Fei-fei; Peng, Bing-ling; Xiao, Rong-quan; Gao, Wei-wei; Ding, Jian-cheng; Liu, Wen

    2016-01-01

    Yin Yang 1 (YY1) is a multifunctional transcription factor shown to be critical in a variety of biological processes. Although it is regulated by multiple types of post-translational modifications (PTMs), whether YY1 is methylated, which enzyme methylates YY1, and hence the functional significance of YY1 methylation remains completely unknown. Here we reported the first methyltransferase, SET7/9 (KMT7), capable of methylating YY1 at two highly conserved lysine (K) residues, K173 and K411, located in two distinct domains, one in the central glycine-rich region and the other in the very carboxyl-terminus. Functional studies revealed that SET7/9-mediated YY1 methylation regulated YY1 DNA-binding activity both in vitro and at specific genomic loci in cultured cells. Consistently, SET7/9-mediated YY1 methylation was shown to involve in YY1-regulated gene transcription and cell proliferation. Our findings revealed a novel regulatory strategy, methylation by lysine methyltransferase, imposed on YY1 protein, and linked YY1 methylation with its biological functions. PMID:26902152

  20. Neurotrophin-mediated degradation of histone methyltransferase by S-nitrosylation cascade regulates neuronal differentiation

    PubMed Central

    Sen, Nilkantha; Snyder, Solomon H.

    2011-01-01

    Epigenetic regulation of histones mediates neurotrophin actions with histone acetylation enhancing cAMP response element-binding (CREB)-associated transcription elicited by brain-derived neurotrophic factor (BDNF) and nerve-growth factor (NGF). Roles for histone methylation in CREB's transcriptional activity have not been well characterized. We show that depletion of the histone methyltransferase suppressor of variegation 3–9 homolog 1 (SUV39H1) selectively augments BDNF- and NGF-mediated neurite outgrowth. SUV39H1 is the principal enzyme responsible for trimethylation of histone H3 at lysine 9, a molecular mark associated with transcriptional silencing. BDNF and NGF act via a signaling cascade wherein degradation of SUV39H1 down-regulates trimethylation of H3K9 in a nitric oxide-dependent pathway. BDNF activates neuronal NOS with the nitrosylated GAPDH/seven in absentia (Siah) homolog complex translocating to the nucleus. Degradation of SUV39H1 by Siah facilitates histone H3 on lysine 9 acetylation, CREB binding to DNA, enhanced expression of CREB-regulated genes and neurite outgrowth. PMID:22123949

  1. Ca2+-Dependent Endoplasmic Reticulum Stress Regulates Mechanical Stress-Mediated Cartilage Thinning.

    PubMed

    Zhu, M; Zhou, S; Huang, Z; Wen, J; Li, H

    2016-07-01

    Our previous study identified that endoplasmic reticulum stress (ERS) plays a critical role in chondrocyte apoptosis and mandibular cartilage thinning in response to compressive mechanical force, although the underlying mechanisms remain elusive. Because the endoplasmic reticulum (ER) is a primary site of intracellular Ca(2+) storage, we hypothesized that Ca(2+)-dependent ERS might be involved in mechanical stress-mediated mandibular cartilage thinning. In this study, we used in vitro and in vivo models to determine Ca(2+) concentrations, histological changes, subcellular changes, apoptosis, and the expression of ERS markers in mandibular cartilage and chondrocytes. The results showed that in chondrocytes, cytosolic Ca(2+) ([Ca(2+)]i) was dramatically increased by compressive mechanical force. Interestingly, the inhibition of Ca(2+) channels by ryanodine and 2-aminoethoxydiphenyl borate, inhibitors of ryanodine receptors and inositol trisphosphate receptors, respectively, partially rescued mechanical force-mediated mandibular cartilage thinning. Furthermore, chondrocyte apoptosis was also compromised by inhibiting the increase in [Ca(2+)]i that occurred in response to compressive mechanical force. Mechanistically, the ERS induced by compressive mechanical force was also repressed by [Ca(2+)]i inhibition, as demonstrated by a decrease in the expression of the ER stress markers 78 kDa glucose-regulated protein (GRP78) and 94 kDa glucose-regulated protein (GRP94) at both the mRNA and protein levels. Collectively, these data identified [Ca(2+)]i as a critical mediator of the pathological changes that occur in mandibular cartilage under compressive mechanical force and shed light on the treatment of mechanical stress-mediated cartilage degradation. PMID:27053115

  2. Clathrin Light Chains Regulate Clathrin-Mediated Trafficking, Auxin Signaling, and Development in Arabidopsis[C][W][OA

    PubMed Central

    Wang, Chao; Yan, Xu; Chen, Qian; Jiang, Nan; Fu, Wei; Ma, Bojun; Liu, Jianzhong; Li, Chuanyou; Bednarek, Sebastian Y.; Pan, Jianwei

    2013-01-01

    Plant clathrin-mediated membrane trafficking is involved in many developmental processes as well as in responses to environmental cues. Previous studies have shown that clathrin-mediated endocytosis of the plasma membrane (PM) auxin transporter PIN-FORMED1 is regulated by the extracellular auxin receptor AUXIN BINDING PROTEIN1 (ABP1). However, the mechanisms by which ABP1 and other factors regulate clathrin-mediated trafficking are poorly understood. Here, we applied a genetic strategy and time-resolved imaging to dissect the role of clathrin light chains (CLCs) and ABP1 in auxin regulation of clathrin-mediated trafficking in Arabidopsis thaliana. Auxin was found to differentially regulate the PM and trans-Golgi network/early endosome (TGN/EE) association of CLCs and heavy chains (CHCs) in an ABP1-dependent but TRANSPORT INHIBITOR RESPONSE1/AUXIN-BINDING F-BOX PROTEIN (TIR1/AFB)-independent manner. Loss of CLC2 and CLC3 affected CHC membrane association, decreased both internalization and intracellular trafficking of PM proteins, and impaired auxin-regulated endocytosis. Consistent with these results, basipetal auxin transport, auxin sensitivity and distribution, and root gravitropism were also found to be dramatically altered in clc2 clc3 double mutants, resulting in pleiotropic defects in plant development. These results suggest that CLCs are key regulators in clathrin-mediated trafficking downstream of ABP1-mediated signaling and thus play a critical role in membrane trafficking from the TGN/EE and PM during plant development. PMID:23424247

  3. Activity of natural products against courgette powdery mildew.

    PubMed

    La Torre, A; Spera, G; Lolleti, D

    2004-01-01

    The effectiveness of several natural products in field trials were tested against powdery mildew on courgette. Sulphur, sodium bicarbonate (alone or in mixture with pinolene, mineral oil or sulphur), sodium silicate in mixture with mineral oil, potassium permanganate, lecithin, Equisetum arvense in mixture with Timus vulgaris, Ampelomyces quisqualis and Reynoutria sachalinensis were evaluated. Generally, in our working conditions (very strong pathogen infection), only some products were effective. Good results were obtained using sulphur and potassium permanganate in the first year and very interesting results about disease control were obtained, in the second year of the trial, using sulphur alone or in mixture with sodium bicarbonate. The products tested did not show any phytotoxic symptoms. PMID:15756857

  4. Bacillus sp. BS061 Suppresses Gray Mold and Powdery Mildew through the Secretion of Different Bioactive Substances.

    PubMed

    Kim, Young-Sook; Song, Ja-Gyeong; Lee, In-Kyoung; Yeo, Woon-Hyung; Yun, Bong-Sik

    2013-09-01

    A Bacillus sp. BS061 significantly reduced disease incidence of gray mold and powdery mildew. To identify the active principle, the culture filtrate was partitioned between butanol and water. The antifungal activity against B. cinerea was evident in the butanol-soluble portion, and active substances were identified as cyclic lipopeptides, iturin A series, by nuclear magnetic resonance spectrometry (NMR) and mass analysis. Interestingly, antifungal activity against powdery mildew was observed in the water-soluble portion, suggesting that cyclic lipopeptides have no responsibility to suppress powdery mildew. This finding reveals that biocontrol agents of Bacillus origin suppress gray mold and powdery mildew through the secretion of different bioactive substances. PMID:24198673

  5. Cell-Mediated Immune Function and Cytokine Regulation During Space Flight

    NASA Technical Reports Server (NTRS)

    Sams, Clarence F.; Pierson, Duane L.; Paloski, W. H. (Technical Monitor)

    2000-01-01

    The changes in immune function which occur during space flight potentially expose the crews to an increased risk for development of illness. Decreased cellular immune function has been repeatedly documented after space flight and confirmed during flight by in vivo delayed-type hypersensitivity testing. However, correlation of immune changes with a clinically significant risk factor has not yet been performed. Our hypothesis is that space flight induces a decrease in cell-mediated immune function accompanied by a shift from a type 1 cytokine pattern (favoring cell-mediated immunity) to a type 2 cytokine pattern (favoring humoral immunity). We further hypothesize that reactivation of latent viruses will occur during space flight in association with the decreased cellular immunity. To test these hypotheses, we will determine the effects of space flight on cell-mediated immunity and viral reactivation. We will utilize delayed-type hypersensitivity testing as an in vivo measure of integrated cell-mediated immune function. The production of cytokines and immunoregulatory factors by lymphocytes and monocytes will be measured to determine whether changes in cytokine patterns are associated with the space flight-induced immune dysregulation. Correlation of antigen-specific immune changes with reactivation of latent herpes viruses will be determined by measuring peripheral levels of viral (CMV, VZV, EBV) antigen-specific T cells and comparing to the levels of EBV-infected B-cells by fluorescence in situ hybridization and flow cytometry. A comparison of cell-mediated immune function, cytokine regulation and viral reactivation will provide new insights into crew member health risks during flight.

  6. RNF4-mediated polyubiquitination regulates the Fanconi anemia/BRCA pathway.

    PubMed

    Xie, Jenny; Kim, Hyungjin; Moreau, Lisa A; Puhalla, Shannon; Garber, Judy; Al Abo, Muthana; Takeda, Shunichi; D'Andrea, Alan D

    2015-04-01

    The Fanconi anemia/BRCA (FA/BRCA) pathway is a DNA repair pathway that is required for excision of DNA interstrand cross-links. The 17 known FA proteins, along with several FA-associated proteins (FAAPs), cooperate in this pathway to detect, unhook, and excise DNA cross-links and to subsequently repair the double-strand breaks generated in the process. In the current study, we identified a patient with FA with a point mutation in FANCA, which encodes a mutant FANCA protein (FANCAI939S). FANCAI939S failed to bind to the FAAP20 subunit of the FA core complex, leading to decreased stability. Loss of FAAP20 binding exposed a SUMOylation site on FANCA at amino acid residue K921, resulting in E2 SUMO-conjugating enzyme UBC9-mediated SUMOylation, RING finger protein 4-mediated (RNF4-mediated) polyubiquitination, and proteasome-mediated degradation of FANCA. Mutation of the SUMOylation site of FANCA rescued the expression of the mutant protein. Wild-type FANCA was also subject to SUMOylation, RNF4-mediated polyubiquitination, and degradation, suggesting that regulated release of FAAP20 from FANCA is a critical step in the normal FA pathway. Consistent with this model, cells lacking RNF4 exhibited interstrand cross-linker hypersensitivity, and the gene encoding RNF4 was epistatic with the other genes encoding members of the FA/BRCA pathway. Together, the results from our study underscore the importance of analyzing unique patient-derived mutations for dissecting complex DNA repair processes. PMID:25751062

  7. Differential dopaminergic regulation of inwardly rectifying potassium channel mediated subthreshold dynamics in striatal medium spiny neurons.

    PubMed

    Zhao, Bo; Zhu, Junling; Dai, Dongqing; Xing, Junling; He, Jiahou; Fu, Zhanyan; Zhang, Lei; Li, Zhuyi; Wang, Wenting

    2016-08-01

    The dorsal striatum plays a key role in motor control and cognitive processes. Proper functioning of the striatum relies on the fine dynamic balance between the direct pathway projection medium spiny neurons (MSNs) that express D1 dopamine receptor (D1 MSNs) and indirect pathway projection MSNs that express D2 dopamine receptor (D2 MSNs). The inwardly rectifying K(+) channels (Kir), which express on both D1 and D2 MSNs, participate in the subthreshold dynamics including the membrane resonance and dendritic integration. However, it remains unclear whether dopamine differentially regulates Kir mediated subthreshold dynamics in two subtypes MSNs. Using transgenic mice that express either tdTomato in D1 MSNs or eGFP in D2 MSNs, we explored the Kir mediated subthreshold dynamics in D1 or D2 MSNs with whole cell patch clamp recording in acute brain slices. We found that D1 receptor agonist increased the Kir current while D2 receptor activation decreased the Kir conductance. The dopamine regulation of the Kir enhanced the resonant frequency and reduced the resonant impedance of D1 MSNs. The converse is ture for D2 MSNs. It also caused an opposing effect on dendritic integration between D1 and D2 MSNs, which can promote stability of the two pathways. The D1 receptor activation modulated Kir through cAMP-PKA signaling, whereas the D2 receptor modulated Kir through PLC-PKC signaling. Our findings demonstrated the differential dopaminergic regulation role of Kir, which mediates distinct subthreshold dynamics, and thus, contributes to the role of dopamine in fine tuning the balance of the striatal direct and indirect pathway activities. PMID:27018450

  8. c-Cbl regulates αPix-mediated cell migration and invasion

    SciTech Connect

    Seong, Min Woo; Park, Ji Ho; Yoo, Hee Min; Yang, Seung Wook; Oh, Kyu Hee; Ka, Seung Hyeun; Park, Dong Eun; Lee, Soon-Tae; Chung, Chin Ha

    2014-12-12

    Highlights: • c-Cbl ubiquitinates αPix for proteasome-mediated degradation. • C6 and A172 glioma cells lack c-Cbl, which leads to stabilization of αPix. • The accumulated αPix promotes migration and invasion of the cancer cells. • The lack of c-Cbl in the cells appears responsible for their malignant behavior. - Abstract: c-Cbl, a RING-type ubiquitin E3 ligase, down-regulates receptor tyrosine kinases, including EGF receptor, and inhibits cell proliferation. Moreover, c-Cbl mutations are frequently found in patients with myeloid neoplasm. Therefore, c-Cbl is known as a tumor suppressor. αPix is expressed only in highly proliferative and mobile cells, including immune cells, and up-regulated in certain invasive tumors, such as glioblastoma multiforme. Here, we showed that c-Cbl serves as an ubiquitin E3 ligase for proteasome-mediated degradation of αPix, but not βPix. Remarkably, the rat C6 and human A172 glioma cells were unable to express c-Cbl, which leads to a dramatic accumulation of αPix. Depletion of αPix by shRNA markedly reduced the ability of the glioma cells to migrate and invade, whereas complementation of shRNA-insensitive αPix promoted it. These results indicate that c-Cbl negatively regulates αPix-mediated cell migration and invasion and the lack of c-Cbl in the C6 and A172 glioma cells is responsible for their malignant behavior.

  9. Sucrose-mediated transcriptional regulation of sucrose symporter activity in the phloem.

    SciTech Connect

    Matt Vaughn Greg Harrington Daniel R Bush

    2002-08-06

    This project was based on our discovery that sucrose acts as a signaling molecule that regulates the activity of a proton-sucrose symporter in sugar beet leaf tissue. A major objective here was determining how sucrose transporter activity is being regulated. When sucrose accumulates in the phloem sucrose transport activity drops dramatically. Western blots of plasma membrane proteins isolated from sucrose treated leaves showed that the loss of sucrose transport activity was proportional to a decline in symporter abundance, demonstrating that sucrose transport is regulated by changes in the amount of BvSUT1 protein. BvSUT1 transcript levels decreased in parallel with the loss of sucrose transport activity. Nuclear run-on experiments demonstrated that BvSUT1 gene transcription was repressed significantly in nuclei from leaves fed 100 mM exogenous sucrose, showing that sucrose-dependent modulation of BvSUT1 mRNA levels is mediated by changes in transcription. To identify which secondary messenger systems might be involved in regulating symporter activity, we used a variety of pharmacological agents to probe for a role of calcium or protein phosphorylation in sucrose signaling. In a detailed analysis, only okadaic acid altered sucrose transport activity. These results suggest a protein phosphatase is involved. We hypothesized that protein kinase inhibitors would have a neutral affect or increase symporter transcription. Transpirational feeding of the protein kinase inhibitor staurosporine had no impact on sucrose transport while calphostin C, an inhibitor of protein kinase C, caused a 60% increase. These data provided good evidence that protein phosphorylation plays a central role in regulating sucrose symporter expression and sucrose transport activity. To determine whether protein phosphorylation is involved in sucrose regulation of proton-sucrose symporter activity, we pre-fed leaves with staurosporine for 4 h and then fed the treated leaves water or 100 mM sucrose

  10. Does rumination mediate the relationship between emotion regulation ability and posttraumatic stress disorder?

    PubMed Central

    Ehring, Thomas; Ehlers, Anke

    2014-01-01

    Background and objectives Trauma-related rumination has been suggested to be involved in the maintenance of posttraumatic stress disorder (PTSD). This view has empirically been supported by extensive evidence using cross-sectional, prospective, and experimental designs. However, it is unclear why trauma survivors engage in rumination despite its negative consequences. The current study aimed to explore the hypothesis that low emotion regulation ability underlies trauma-related rumination. Methods Emotion regulation ability and trauma-related rumination were assessed in 93 road traffic accident survivors 2 weeks post-trauma. In addition, symptom levels of PTSD were assessed at 2 weeks as well as 1, 3, and 6 months follow-up. Results Emotion regulation ability was significantly related to trauma-related rumination as well as levels of PTSD symptoms. In addition, the association between low emotion regulation ability and PTSD was mediated by rumination. Conclusions The findings support the view that rumination is used as a dysfunctional emotion regulation strategy by trauma survivors. PMID:25206955

  11. Genome-wide analysis of FOXO3 mediated transcription regulation through RNA polymerase II profiling.

    PubMed

    Eijkelenboom, Astrid; Mokry, Michal; de Wit, Elzo; Smits, Lydia M; Polderman, Paulien E; van Triest, Miranda H; van Boxtel, Ruben; Schulze, Almut; de Laat, Wouter; Cuppen, Edwin; Burgering, Boudewijn M T

    2013-01-01

    Forkhead box O (FOXO) transcription factors are key players in diverse cellular processes affecting tumorigenesis, stem cell maintenance and lifespan. To gain insight into the mechanisms of FOXO-regulated target gene expression, we studied genome-wide effects of FOXO3 activation. Profiling RNA polymerase II changes shows that FOXO3 regulates gene expression through transcription initiation. Correlative analysis of FOXO3 and RNA polymerase II ChIP-seq profiles demonstrates FOXO3 to act as a transcriptional activator. Furthermore, this analysis reveals a significant part of FOXO3 gene regulation proceeds through enhancer regions. FOXO3 binds to pre-existing enhancers and further activates these enhancers as shown by changes in histone acetylation and RNA polymerase II recruitment. In addition, FOXO3-mediated enhancer activation correlates with regulation of adjacent genes and pre-existence of chromatin loops between FOXO3 bound enhancers and target genes. Combined, our data elucidate how FOXOs regulate gene transcription and provide insight into mechanisms by which FOXOs can induce different gene expression programs depending on chromatin architecture. PMID:23340844

  12. TRIM39 negatively regulates the NFκB-mediated signaling pathway through stabilization of Cactin.

    PubMed

    Suzuki, Masanobu; Watanabe, Masashi; Nakamaru, Yuji; Takagi, Dai; Takahashi, Hidehisa; Fukuda, Satoshi; Hatakeyama, Shigetsugu

    2016-03-01

    NFκB is one of the central regulators of cell survival, immunity, inflammation, carcinogenesis and organogenesis. The activation of NFκB is strictly regulated by several posttranslational modifications including phosphorylation, neddylation and ubiquitination. Several types of ubiquitination play important roles in multi-step regulations of the NFκB pathway. Some of the tripartite motif-containing (TRIM) proteins functioning as E3 ubiquitin ligases are known to regulate various biological processes such as inflammatory signaling pathways. One of the TRIM family proteins, TRIM39, for which the gene has single nucleotide polymorphisms, has been identified as one of the genetic factors in Behcet's disease. However, the role of TRIM39 in inflammatory signaling had not been fully elucidated. In this study, to elucidate the function of TRIM39 in inflammatory signaling, we performed yeast two-hybrid screening using TRIM39 as a bait and identified Cactin, which has been reported to inhibit NFκB- and TLR-mediated transcriptions. We show that TRIM39 stabilizes Cactin protein and that Cactin is upregulated after TNFα stimulation. TRIM39 knockdown also causes activation of the NFκB signal. These findings suggest that TRIM39 negatively regulates the NFκB signal in collaboration with Cactin induced by inflammatory stimulants such as TNFα. PMID:26363554

  13. Dual Function of NAC072 in ABF3-Mediated ABA-Responsive Gene Regulation in Arabidopsis

    PubMed Central

    Li, Xiaoyun; Li, Xiaoling; Li, Meijuan; Yan, Youcheng; Liu, Xu; Li, Ling

    2016-01-01

    The NAM, ATAF1/2, and CUC2 (NAC) domain proteins play various roles in plant growth and stress responses. Arabidopsis NAC transcription factor NAC072 has been reported as a transcriptional activator in Abscisic acid (ABA)-responsive gene expression. However, the exact function of NAC072 in ABA signaling is still elusive. In this study, we present evidence for the interrelation between NAC072 and ABA-responsive element binding factor 3 (ABF3) that act as a positive regulator of ABA-responsive gene expression in Arabidopsis. The transcript of NAC072 is up-regulated by ABF3 in ABA response, and NAC072 protein interacts with ABF3. Enhanced ABA sensitivity occurs in nac072 mutant plants that overexpressed ABF3. However, overexpression of NAC072 weakened the ABA sensitivity in the abf3 mutant plants, but instead of recovering the ABA sensitivity of abf3. NAC072 and ABF3 cooperate to regulate RD29A expression, but are antagonistic when regulating RD29B expression. Therefore, NAC072 displays a dual function in ABF3-mediated ABA-responsive gene regulation. PMID:27486475

  14. Peroxide-Dependent MGL Sulfenylation Regulates 2-AG-Mediated Endocannabinoid Signaling in Brain Neurons.

    PubMed

    Dotsey, Emmanuel Y; Jung, Kwang-Mook; Basit, Abdul; Wei, Don; Daglian, Jennifer; Vacondio, Federica; Armirotti, Andrea; Mor, Marco; Piomelli, Daniele

    2015-05-21

    The second messenger hydrogen peroxide transduces changes in the cellular redox state by reversibly oxidizing protein cysteine residues to sulfenic acid. This signaling event regulates many cellular processes but has never been shown to occur in the brain. Here, we report that hydrogen peroxide heightens endocannabinoid signaling in brain neurons through sulfenylation of cysteines C201 and C208 in monoacylglycerol lipase (MGL), a serine hydrolase that deactivates the endocannabinoid 2-arachidonoyl-sn-glycerol (2-AG) in nerve terminals. The results suggest that MGL sulfenylation may provide a presynaptic control point for 2-AG-mediated endocannabinoid signaling. PMID:26000748

  15. P-cadherin-mediated Rho GTPase regulation during collective cell migration

    PubMed Central

    Plutoni, Cédric; Bazellières, Elsa; Gauthier-Rouvière, Cécile

    2016-01-01

    ABSTRACT This commentary addresses the role of P-cadherin in collective cell migration (CCM), a cooperative and coordinated migration mode, used by cells during normal and pathological migration processes. We discuss how cadherin-mediated cell-cell junctions (CCJs) play a critical role in CCM through their ability to regulate Rho GTPase-dependent pathways and how this leads to the generation and orientation of mechanical forces. We will also highlight the key function of P-cadherin (a poor prognostic marker in several tumors) in promoting collective cell movement in epithelial and mesenchymal cells. PMID:27152729

  16. Neutrophil-Mediated Regulation of Innate and Adaptive Immunity: The Role of Myeloperoxidase

    PubMed Central

    Odobasic, Dragana; Kitching, A. Richard; Holdsworth, Stephen R.

    2016-01-01

    Neutrophils are no longer seen as leukocytes with a sole function of being the essential first responders in the removal of pathogens at sites of infection. Being armed with numerous pro- and anti-inflammatory mediators, these phagocytes can also contribute to the development of various autoimmune diseases and can positively or negatively regulate the generation of adaptive immune responses. In this review, we will discuss how myeloperoxidase, the most abundant neutrophil granule protein, plays a key role in the various functions of neutrophils in innate and adaptive immunity. PMID:26904693

  17. Regulation of competence-mediated horizontal gene transfer in the natural habitat of Vibrio cholerae.

    PubMed

    Metzger, Lisa C; Blokesch, Melanie

    2016-04-01

    The human pathogen Vibrio cholerae is an autochthonous inhabitant of aquatic environments where it often interacts with zooplankton and their chitinous molts. Chitin induces natural competence for transformation in V. cholerae, a key mode of horizontal gene transfer (HGT). Recent comparative genomic analyses were indicative of extensive HGT in this species. However, we can still expand our understanding of the complex regulatory network that drives competence in V. cholerae. Here, we present recent advances, including the elucidation of bipartite competence regulation mediated by QstR, the inclusion of the type VI secretion system in the competence regulon of pandemic O1 El Tor strains, and the identification of TfoS as a transcriptional regulator that links chitin to competence induction in V. cholerae. PMID:26615332

  18. Antennally mediated negative feedback regulation of pheromone production in the pine engraver beetle, Ips pini

    NASA Astrophysics Data System (ADS)

    Ginzel, Matthew D.; Bearfield, Jeremy C.; Keeling, Christopher I.; McCormack, Colin C.; Blomquist, Gary J.; Tittiger, Claus

    2007-01-01

    Bark beetles use monoterpenoid aggregation pheromones to coordinate host colonization and mating. These chemical signals are produced de novo in midgut cells via the mevalonate pathway, and pheromone production may be regulated by a negative feedback system mediated through the antennae. In this study, we explored the effect of antennectomy on pheromone production and transcript levels of key mevalonate pathway genes in juvenile hormone III-treated male pine engraver beetles, Ips pini (Say). Antennectomized males produced significantly greater amounts of pheromone than podectomized males and those with intact antennae. Likewise, mRNA levels of three mevalonate pathway genes important in pheromone biosynthesis were measured by quantitative real-time PCR and found to be induced to a greater extent with antennectomy, suggesting a transcriptional regulation of pheromone production.

  19. Neuroligin 1 regulates spines and synaptic plasticity via LIMK1/cofilin-mediated actin reorganization.

    PubMed

    Liu, An; Zhou, Zikai; Dang, Rui; Zhu, Yuehua; Qi, Junxia; He, Guiqin; Leung, Celeste; Pak, Daniel; Jia, Zhengping; Xie, Wei

    2016-02-15

    Neuroligin (NLG) 1 is important for synapse development and function, but the underlying mechanisms remain unclear. It is known that at least some aspects of NLG1 function are independent of the presynaptic neurexin, suggesting that the C-terminal domain (CTD) of NLG1 may be sufficient for synaptic regulation. In addition, NLG1 is subjected to activity-dependent proteolytic cleavage, generating a cytosolic CTD fragment, but the significance of this process remains unknown. In this study, we show that the CTD of NLG1 is sufficient to (a) enhance spine and synapse number, (b) modulate synaptic plasticity, and (c) exert these effects via its interaction with spine-associated Rap guanosine triphosphatase-activating protein and subsequent activation of LIM-domain protein kinase 1/cofilin-mediated actin reorganization. Our results provide a novel postsynaptic mechanism by which NLG1 regulates synapse development and function. PMID:26880202

  20. Feedback regulation via AMPK and HIF-1 mediates ROS-dependent longevity in Caenorhabditis elegans

    PubMed Central

    Hwang, Ara B.; Ryu, Eun-A; Artan, Murat; Chang, Hsin-Wen; Kabir, Mohammad Humayun; Nam, Hyun-Jun; Lee, Dongyeop; Yang, Jae-Seong; Kim, Sanguk; Mair, William B.; Lee, Cheolju; Lee, Siu Sylvia; Lee, Seung-Jae

    2014-01-01

    Mild inhibition of mitochondrial respiration extends the lifespan of many species. In Caenorhabditis elegans, reactive oxygen species (ROS) promote longevity by activating hypoxia-inducible factor 1 (HIF-1) in response to reduced mitochondrial respiration. However, the physiological role and mechanism of ROS-induced longevity are poorly understood. Here, we show that a modest increase in ROS increases the immunity and lifespan of C. elegans through feedback regulation by HIF-1 and AMP-activated protein kinase (AMPK). We found that activation of AMPK as well as HIF-1 mediates the longevity response to ROS. We further showed that AMPK reduces internal levels of ROS, whereas HIF-1 amplifies the levels of internal ROS under conditions that increase ROS. Moreover, mitochondrial ROS increase resistance to various pathogenic bacteria, suggesting a possible association between immunity and long lifespan. Thus, AMPK and HIF-1 may control immunity and longevity tightly by acting as feedback regulators of ROS. PMID:25288734

  1. EPAS-1 Mediates SP-1-Dependent FBI-1 Expression and Regulates Tumor Cell Survival and Proliferation

    PubMed Central

    Wang, Xiaogang; Cao, Peng; Li, Zhiqing; Wu, Dongyang; Wang, Xi; Liang, Guobiao

    2014-01-01

    Factor binding IST-1 (FBI-1) plays an important role in oncogenic transformation and tumorigenesis. As FBI-1 is over-expressed in multiple human cancers, the regulation of itself would provide new effective options for cancer intervention. In this work, we aimed to study the role that EPAS-1 plays in regulating FBI-1. We use the fact that specificity protein-1 (SP-1) is one of the crucial transcription factors of FBI-1, and that SP-1 can interact with the endothelial pas domain protein-1 (EPAS-1) for the induction of hypoxia related genes. The study showed that EPAS-1 plays an indispensible role in SP-1 transcription factor-mediated FBI-1 induction, and participated in tumor cell survival and proliferation. Thus, EPAS-1 could be a novel target for cancer therapeutics. PMID:25192290

  2. EPAS-1 mediates SP-1-dependent FBI-1 expression and regulates tumor cell survival and proliferation.

    PubMed

    Wang, Xiaogang; Cao, Peng; Li, Zhiqing; Wu, Dongyang; Wang, Xi; Liang, Guobiao

    2014-01-01

    Factor binding IST-1 (FBI-1) plays an important role in oncogenic transformation and tumorigenesis. As FBI-1 is over-expressed in multiple human cancers, the regulation of itself would provide new effective options for cancer intervention. In this work, we aimed to study the role that EPAS-1 plays in regulating FBI-1. We use the fact that specificity protein-1 (SP-1) is one of the crucial transcription factors of FBI-1, and that SP-1 can interact with the endothelial pas domain protein-1 (EPAS-1) for the induction of hypoxia related genes. The study showed that EPAS-1 plays an indispensible role in SP-1 transcription factor-mediated FBI-1 induction, and participated in tumor cell survival and proliferation. Thus, EPAS-1 could be a novel target for cancer therapeutics. PMID:25192290

  3. Coping, emotion regulation, and self-blame as mediators of sexual abuse and psychological symptoms in adult sexual assault.

    PubMed

    Ullman, Sarah E; Peter-Hagene, Liana C; Relyea, Mark

    2014-01-01

    This study examined whether coping, emotion regulation, and self-blame mediate relationships of trauma histories with post-traumatic stress disorder and depression in adult sexual assault victims (N = 1863). A path analysis showed that theorized mediators partially mediated associations between trauma history variables and psychological symptoms. Specifically, child sexual abuse severity was related to greater post-traumatic stress disorder and depression indirectly through maladaptive coping and decreased emotion regulation but not self-blame. Other traumas had direct relationships with symptoms and partially mediated effects through maladaptive coping and emotion regulation. Child sexual abuse was unrelated to self-blame, but other traumas were related to greater self-blame. Results differed according to whether women had counseling post-assault. Implications are drawn for future research and clinical treatment of adult sexual assault victims. PMID:24393091

  4. COPI-mediated retrograde trafficking from the Golgi to the ER regulates EGFR nuclear transport

    SciTech Connect

    Wang, Ying-Nai; Wang, Hongmei; Yamaguchi, Hirohito; Lee, Hong-Jen; Lee, Heng-Huan; Hung, Mien-Chie

    2010-09-03

    Research highlights: {yields} ARF1 activation is involved in the EGFR transport to the ER and the nucleus. {yields} Assembly of {gamma}-COP coatomer mediates EGFR transport to the ER and the nucleus. {yields} Golgi-to-ER retrograde trafficking regulates nuclear transport of EGFR. -- Abstract: Emerging evidence indicates that cell surface receptors, such as the entire epidermal growth factor receptor (EGFR) family, have been shown to localize in the nucleus. A retrograde route from the Golgi to the endoplasmic reticulum (ER) is postulated to be involved in the EGFR trafficking to the nucleus; however, the molecular mechanism in this proposed model remains unexplored. Here, we demonstrate that membrane-embedded vesicular trafficking is involved in the nuclear transport of EGFR. Confocal immunofluorescence reveals that in response to EGF, a portion of EGFR redistributes to the Golgi and the ER, where its NH{sub 2}-terminus resides within the lumen of Golgi/ER and COOH-terminus is exposed to the cytoplasm. Blockage of the Golgi-to-ER retrograde trafficking by brefeldin A or dominant mutants of the small GTPase ADP-ribosylation factor, which both resulted in the disassembly of the coat protein complex I (COPI) coat to the Golgi, inhibit EGFR transport to the ER and the nucleus. We further find that EGF-dependent nuclear transport of EGFR is regulated by retrograde trafficking from the Golgi to the ER involving an association of EGFR with {gamma}-COP, one of the subunits of the COPI coatomer. Our findings experimentally provide a comprehensive pathway that nuclear transport of EGFR is regulated by COPI-mediated vesicular trafficking from the Golgi to the ER, and may serve as a general mechanism in regulating the nuclear transport of other cell surface receptors.

  5. Serotonin Receptor 2B Mediates Mechanical Hyperalgesia by Regulating Transient Receptor Potential Vanilloid 1.

    PubMed

    Su, Yeu-Shiuan; Chiu, Yuan-Yi; Lin, Shih-Yuan; Chen, Chih-Cheng; Sun, Wei-Hsin

    2016-05-01

    Serotonin [5-hydroxytryptamine (5-HT)], an inflammatory mediator, contributes to inflammatory pain. The presence of multiple 5-HT subtype receptors on peripheral and central nociceptors complicates the role of 5-HT in pain. Previously, we found that 5-HT2B/2C antagonist could block 5-HT-induced mechanical hyperalgesia. However, the types of neurons or circuits underlying this effect remained unsolved. Here, we demonstrate that the Gq/11-phospholipase Cβ-protein kinase Cε (PKCε) pathway mediated by 5-HT2B is involved in 5-HT-induced mechanical hyperalgesia in mice. Administration of a transient receptor potential vanilloid 1 (TRPV1) antagonist inhibited the 5-HT-induced mechanical hyperalgesia. 5-HT injection enhanced 5-HT- and capsaicin-evoked calcium signals specifically in isolectin B4 (IB4)-negative neurons; signals were inhibited by a 5-HT2B/2C antagonist and PKCε blocker. Thus, 5-HT2B mediates 5-HT-induced mechanical hyperalgesia by regulating TRPV1 function. PMID:26635025

  6. Palmitoylation of cytoskeleton associated protein 4 by DHHC2 regulates antiproliferative factor-mediated signaling.

    PubMed

    Planey, Sonia L; Keay, Susan K; Zhang, Chen-Ou; Zacharias, David A

    2009-03-01

    Previously, we identified cytoskeleton-associated protein 4 (CKAP4) as a major substrate of the palmitoyl acyltransferase, DHHC2, using a novel proteomic method called palmitoyl-cysteine identification, capture and analysis (PICA). CKAP4 is a reversibly palmitoylated and phosphorylated protein that links the ER to the cytoskeleton. It is also a high-affinity receptor for antiproliferative factor (APF), a small sialoglycopeptide secreted from bladder epithelial cells of patients with interstitial cystitis (IC). The role of DHHC2-mediated palmitoylation of CKAP4 in the antiproliferative response of HeLa and normal bladder epithelial cells to APF was investigated. Our data show that siRNA-mediated knockdown of DHHC2 and consequent suppression of CKAP4 palmitoylation inhibited the ability of APF to regulate cellular proliferation and blocked APF-induced changes in the expression of E-cadherin, vimentin, and ZO-1 (genes known to play a role in cellular proliferation and tumorigenesis). Immunocytochemistry revealed that CKAP4 palmitoylation by DHHC2 is required for its trafficking from the ER to the plasma membrane and for its nuclear localization. These data suggest an important role for DHHC2-mediated palmitoylation of CKAP4 in IC and in opposing cancer-related cellular behaviors and support the idea that DHHC2 is a tumor suppressor. PMID:19144824

  7. RNF43 interacts with NEDL1 and regulates p53-mediated transcription

    SciTech Connect

    Shinada, Keisuke; Tsukiyama, Tadasuke; Sho, Takuya; Okumura, Fumihiko; Asaka, Masahiro; Hatakeyama, Shigetsugu

    2011-01-07

    Research highlights: {yields} RNF43 binds to NEDD-4-like ubiquitin-protein ligase-1 (NEDL1). {yields} RNF43 interacts with p53 and suppresses transcriptional activity of p53. {yields} RNF43 attenuates apoptosis induced by ultraviolet irradiation. {yields} RNF43 is likely associated with p53-mediated apoptosis in collaboration with NEDL1 in colorectal carcinogenesis. -- Abstract: The ubiquitin-proteasomal system plays a crucial role in oncogenesis in colorectal tissues. Recent studies have shown that stability of {beta}-catenin, which functions as an oncogene for colorectal cancer, is regulated by ubiquitin-mediated degradation. It has been reported that a putative E3 ubiquitin ligase, RNF43, is highly expressed in human colorectal carcinoma and that RNF43 promotes cell growth. However, the involvement of RNF43 in carcinogenesis has not been fully elucidated. In this study, we found by using yeast two-hybrid screening that RNF43 binds to NEDD-4-like ubiquitin-protein ligase-1 (NEDL1), which enhances pro-apoptotic activity by p53. In addition, we found that RNF43 also interacts with p53 and that RNF43 suppresses transcriptional activity of p53 in H1299 cells and attenuates apoptosis induced by ultraviolet irradiation. These findings suggest that RNF43 is associated with p53-mediated apoptosis in collaboration with NEDL1 in colorectal carcinogenesis.

  8. Influence of Student Learning Experience on Academic Performance: The Mediator and Moderator Effects of Self-Regulation and Motivation

    ERIC Educational Resources Information Center

    Ning, Hoi Kwan; Downing, Kevin

    2012-01-01

    This study examined the mediator and moderator roles of self-regulation and motivation constructs in the relationship between learning experience and academic success. Self-reported measures of learning experience, self-regulation and motivation were obtained from 384 undergraduate students from a university in Hong Kong. Structural equation…

  9. Regulation of clathrin coat assembly by Eps15 homology domain–mediated interactions during endocytosis

    PubMed Central

    Suzuki, Ryohei; Toshima, Junko Y.; Toshima, Jiro

    2012-01-01

    Clathrin-mediated endocytosis involves a coordinated series of molecular events regulated by interactions among a variety of proteins and lipids through specific domains. One such domain is the Eps15 homology (EH) domain, a highly conserved protein–protein interaction domain present in a number of proteins distributed from yeast to mammals. Several lines of evidence suggest that the yeast EH domain–containing proteins Pan1p, End3p, and Ede1p play important roles during endocytosis. Although genetic and cell-biological studies of these proteins suggested a role for the EH domains in clathrin-mediated endocytosis, it was unclear how they regulate clathrin coat assembly. To explore the role of the EH domain in yeast endocytosis, we mutated those of Pan1p, End3p, or Ede1p, respectively, and examined the effects of single, double, or triple mutation on clathrin coat assembly. We found that mutations of the EH domain caused a defect of cargo internalization and a delay of clathrin coat assembly but had no effect on assembly of the actin patch. We also demonstrated functional redundancy among the EH domains of Pan1p, End3p, and Ede1p for endocytosis. Of interest, the dynamics of several endocytic proteins were differentially affected by various EH domain mutations, suggesting functional diversity of each EH domain. PMID:22190739

  10. Differences in kinase-mediated regulation of cell cycle progression in normal and transformed cells

    SciTech Connect

    Crissman, H.A.; Gadbois, D.M.; Tobey, R.A.; Stevenson, A.P.; Kraemer, P.M.; Bustos, L.D.; Dickson, J.A.; Bradbury, E.M. )

    1993-01-01

    Staurosporine (Stsp), a general protein kinase inhibitor, was used to investigate the role of kinase-mediated mechanisms in regulating mammalian cell proliferation. Low levels of Stsp (1-2nM) prevented nontransformed cells from entering S phase, indicating that protein phosphorylation processes are essential for commitment of DNA replication in normal cells. Cells resumed cycling when Stsp was removed. The period of sensitivity of nontransformed human diploid fibroblasts to low levels of the drug commenced 3 h later than the G0/G1 boundary and extended through the G1/S boundary. The initial block point at 3 h corresponds neither to the serum nor the amino acid restriction point. In contrast, neither low nor high concentrations (100nm) of Stsp affected G1 progression of transformed cells. High drug concentrations blocked normal cells in G1 and G2 but affected only G2-progression in transformed cells. These results indicate that kinase-mediated regulation of DNA replication is lost as a result of neoplastic transformation, but the G2-arrest mechanism remains intact.

  11. FIAT represses ATF4-mediated transcription to regulate bone mass in transgenic mice.

    PubMed

    Yu, Vionnie W C; Ambartsoumian, Gourgen; Verlinden, Lieve; Moir, Janet M; Prud'homme, Josée; Gauthier, Claude; Roughley, Peter J; St-Arnaud, René

    2005-05-23

    We report the characterization of factor inhibiting activating transcription factor 4 (ATF4)-mediated transcription (FIAT), a leucine zipper nuclear protein. FIAT interacted with ATF4 to inhibit binding of ATF4 to DNA and block ATF4-mediated transcription of the osteocalcin gene in vitro. Transgenic mice overexpressing FIAT in osteoblasts also had reduced osteocalcin gene expression and decreased bone mineral density, bone volume, mineralized volume, trabecular thickness, trabecular number, and decreased rigidity of long bones. Mineral homeostasis, osteoclast number and activity, and osteoblast proliferation and apoptosis were unchanged in transgenics. Expression of osteoblastic differentiation markers was largely unaffected and type I collagen synthesis was unchanged. Mineral apposition rate was reduced in transgenic mice, suggesting that the lowered bone mass was due to a decline in osteoblast activity. This cell-autonomous decrease in osteoblast activity was confirmed by measuring reduced alkaline phosphatase activity and mineralization in primary osteoblast cultures. These results show that FIAT regulates bone mass accrual and establish FIAT as a novel transcriptional regulator of osteoblastic function. PMID:15911876

  12. Ubiquitin Ligase TRIM62 Regulates CARD9-Mediated Anti-fungal Immunity and Intestinal Inflammation.

    PubMed

    Cao, Zhifang; Conway, Kara L; Heath, Robert J; Rush, Jason S; Leshchiner, Elizaveta S; Ramirez-Ortiz, Zaida G; Nedelsky, Natalia B; Huang, Hailiang; Ng, Aylwin; Gardet, Agnès; Cheng, Shih-Chin; Shamji, Alykhan F; Rioux, John D; Wijmenga, Cisca; Netea, Mihai G; Means, Terry K; Daly, Mark J; Xavier, Ramnik J

    2015-10-20

    CARD9 is a central component of anti-fungal innate immune signaling via C-type lectin receptors, and several immune-related disorders are associated with CARD9 alterations. Here, we used a rare CARD9 variant that confers protection against inflammatory bowel disease as an entry point to investigating CARD9 regulation. We showed that the protective variant of CARD9, which is C-terminally truncated, acted in a dominant-negative manner for CARD9-mediated cytokine production, indicating an important role for the C terminus in CARD9 signaling. We identified TRIM62 as a CARD9 binding partner and showed that TRIM62 facilitated K27-linked poly-ubiquitination of CARD9. We identified K125 as the ubiquitinated residue on CARD9 and demonstrated that this ubiquitination was essential for CARD9 activity. Furthermore, we showed that similar to Card9-deficient mice, Trim62-deficient mice had increased susceptibility to fungal infection. In this study, we utilized a rare protective allele to uncover a TRIM62-mediated mechanism for regulation of CARD9 activation. PMID:26488816

  13. TNL-mediated immunity in Arabidopsis requires complex regulation of the redundant ADR1 gene family.

    PubMed

    Dong, Oliver Xiaoou; Tong, Meixuezi; Bonardi, Vera; El Kasmi, Farid; Woloshen, Virginia; Wünsch, Lisa K; Dangl, Jeffery L; Li, Xin

    2016-05-01

    Nucleotide-binding leucine-rich repeat proteins (NLRs) serve as intracellular immune receptors in animals and plants. Sensor NLRs perceive pathogen-derived effector molecules and trigger robust host defense. Recent studies revealed the role of three coiled-coil-type NLRs (CNLs) of the ADR1 family - ADR1, ADR1-L1 and ADR1-L2 - as redundant helper NLRs, whose function is required for defense mediated by multiple sensor NLRs. From a mutant snc1-enhancing (MUSE) forward genetic screen in Arabidopsis targeted to identify negative regulators of snc1 that encodes a TIR-type NLR (TNL), we isolated two alleles of muse15, both carrying mutations in ADR1-L1. Interestingly, loss of ADR1-L1 also enhances immunity-related phenotypes in other autoimmune mutants including cpr1, bal and lsd1. This immunity-enhancing effect is not mediated by increased SNC1 protein stability, nor is it fully dependent on the accumulation of the defense hormone salicylic acid (SA). Transcriptional analysis revealed an upregulation of ADR1 and ADR1-L2 in the adr1-L1 background, which may overcompensate the loss of ADR1-L1, resulting in enhanced immunity. Interestingly, autoimmunity of snc1 and chs2, which encode typical TNLs, is fully suppressed by the adr1 triple mutant, suggesting that the ADRs are required for TNL downstream signaling. This study extends our knowledge on the interplay among ADRs and reveals their complexity in defense regulation. PMID:27074399

  14. MBNL1-mediated regulation of differentiation RNAs promotes myofibroblast transformation and the fibrotic response

    PubMed Central

    Davis, Jennifer; Salomonis, Nathan; Ghearing, Natasha; Lin, Suh-Chin J.; Kwong, Jennifer Q.; Mohan, Apoorva; Swanson, Maurice S.; Molkentin, Jeffery D.

    2015-01-01

    The differentiation of fibroblasts into myofibroblasts mediates tissue wound healing and fibrotic remodelling, although the molecular programme underlying this process remains poorly understood. Here we perform a genome-wide screen for genes that control myofibroblast transformation, and identify the RNA-binding protein muscleblind-like1 (MBNL1). MBNL1 overexpression promotes transformation of fibroblasts into myofibroblasts, whereas loss of Mbnl1 abrogates transformation and impairs the fibrotic phase of wound healing in mouse models of myocardial infarction and dermal injury. Mechanistically, MBNL1 directly binds to and regulates a network of differentiation-specific and cytoskeletal/matrix-assembly transcripts to promote myofibroblast differentiation. One of these transcripts is the nodal transcriptional regulator serum response factor (SRF), whereas another is calcineurin Aβ. CRISPR-Cas9-mediated gene-editing of the MBNL1-binding site within the Srf 3′UTR impairs myofibroblast differentiation, whereas in vivo deletion of Srf in fibroblasts impairs wound healing and fibrosis. These data establish a new RNA-dependent paradigm for myofibroblast formation through MBNL1. PMID:26670661

  15. Regulation of DNA methylation patterns by CK2-mediated phosphorylation of Dnmt3a.

    PubMed

    Deplus, Rachel; Blanchon, Loïc; Rajavelu, Arumugam; Boukaba, Abdelhalim; Defrance, Matthieu; Luciani, Judith; Rothé, Françoise; Dedeurwaerder, Sarah; Denis, Hélène; Brinkman, Arie B; Simmer, Femke; Müller, Fabian; Bertin, Benjamin; Berdasco, Maria; Putmans, Pascale; Calonne, Emilie; Litchfield, David W; de Launoit, Yvan; Jurkowski, Tomasz P; Stunnenberg, Hendrik G; Bock, Christoph; Sotiriou, Christos; Fraga, Mario F; Esteller, Manel; Jeltsch, Albert; Fuks, François

    2014-08-01

    DNA methylation is a central epigenetic modification that is established by de novo DNA methyltransferases. The mechanisms underlying the generation of genomic methylation patterns are still poorly understood. Using mass spectrometry and a phosphospecific Dnmt3a antibody, we demonstrate that CK2 phosphorylates endogenous Dnmt3a at two key residues located near its PWWP domain, thereby downregulating the ability of Dnmt3a to methylate DNA. Genome-wide DNA methylation analysis shows that CK2 primarily modulates CpG methylation of several repeats, most notably of Alu SINEs. This modulation can be directly attributed to CK2-mediated phosphorylation of Dnmt3a. We also find that CK2-mediated phosphorylation is required for localization of Dnmt3a to heterochromatin. By revealing phosphorylation as a mode of regulation of de novo DNA methyltransferase function and by uncovering a mechanism for the regulation of methylation at repetitive elements, our results shed light on the origin of DNA methylation patterns. PMID:25066127

  16. Versatile in vivo regulation of tumor phenotypes by dCas9-mediated transcriptional perturbation.

    PubMed

    Braun, Christian J; Bruno, Peter M; Horlbeck, Max A; Gilbert, Luke A; Weissman, Jonathan S; Hemann, Michael T

    2016-07-01

    Targeted transcriptional regulation is a powerful tool to study genetic mediators of cellular behavior. Here, we show that catalytically dead Cas9 (dCas9) targeted to genomic regions upstream or downstream of the transcription start site allows for specific and sustainable gene-expression level alterations in tumor cells in vitro and in syngeneic immune-competent mouse models. We used this approach for a high-coverage pooled gene-activation screen in vivo and discovered previously unidentified modulators of tumor growth and therapeutic response. Moreover, by using dCas9 linked to an activation domain, we can either enhance or suppress target gene expression simply by changing the genetic location of dCas9 binding relative to the transcription start site. We demonstrate that these directed changes in gene-transcription levels occur with minimal off-target effects. Our findings highlight the use of dCas9-mediated transcriptional regulation as a versatile tool to reproducibly interrogate tumor phenotypes in vivo. PMID:27325776

  17. Role of scleraxis in mechanical stretch-mediated regulation of cardiac myofibroblast phenotype.

    PubMed

    Roche, Patricia L; Nagalingam, Raghu S; Bagchi, Rushita A; Aroutiounova, Nina; Belisle, Breanna M J; Wigle, Jeffrey T; Czubryt, Michael P

    2016-08-01

    The phenotype conversion of fibroblasts to myofibroblasts plays a key role in the pathogenesis of cardiac fibrosis. Numerous triggers of this conversion process have been identified, including plating of cells on solid substrates, cytokines such as transforming growth factor-β, and mechanical stretch; however, the underlying mechanisms remain incompletely defined. Recent studies from our laboratory revealed that the transcription factor scleraxis is a key regulator of cardiac fibroblast phenotype and extracellular matrix expression. Here we report that mechanical stretch induces type I collagen expression and morphological changes indicative of cardiac myofibroblast conversion, as well as scleraxis expression via activation of the scleraxis promoter. Scleraxis causes phenotypic changes similar to stretch, and the effect of stretch is attenuated in scleraxis null cells. Scleraxis was also sufficient to upregulate expression of vinculin and F-actin, to induce stress fiber and focal adhesion formation, and to attenuate both cell migration and proliferation, further evidence of scleraxis-mediated regulation of fibroblast to myofibroblast conversion. Together, these data confirm that scleraxis is sufficient to promote the myofibroblast phenotype and is a required effector of stretch-mediated conversion. Scleraxis may thus represent a potential target for the development of novel antifibrotic therapies aimed at inhibiting myofibroblast formation. PMID:27357547

  18. Identification of signaling pathways regulating primary cilium length and flow-mediated adaptation

    PubMed Central

    Besschetnova, Tatiana Y.; Kolpakova-Hart, Elona; Guan, Yinghua; Zhou, Jing; Olsen, Bjorn R.; Shah, Jagesh V.

    2009-01-01

    Summary The primary cilium acts as a transducer of extracellular stimuli into intracellular signaling [1, 2]. Its regulation, particularly with respect to length, has been defined primarily by genetic experiments and human disease states in which molecular components that are necessary for its proper construction have been mutated or deleted [1]. However, dynamic modulation of cilium length, a phenomenon observed in ciliated protists [3, 4], has not been well-characterized in vertebrates. Here we demonstrate that decreased intracellular calcium (Ca2+) or increased cyclic AMP (cAMP), and subsequent PKA activation, increases primary cilium length in mammalian epithelial and mesenchymal cells. Anterograde intraflagellar transport is sped up in lengthened cilia, potentially increasing delivery flux of cilium components. The cilium length response creates a negative feedback loop whereby fluid shear-mediated deflection of the primary cilium, which decreases intracellular cAMP, leads to cilium shortening and thus decreases mechanotransductive signaling. This adaptive response is blocked when the autosomal dominant polycystic kidney disease (ADPKD) gene products, polycystin-1 or -2, are reduced. Dynamic regulation of cilium length is thus intertwined with cilium-mediated signaling and provides a natural braking mechanism in response to external stimuli that may be compromised in PKD. PMID:20096584

  19. FGFR2c-mediated ERK-MAPK activity regulates coronal suture development.

    PubMed

    Pfaff, Miles J; Xue, Ke; Li, Li; Horowitz, Mark C; Steinbacher, Derek M; Eswarakumar, Jacob V P

    2016-07-15

    Fibroblast growth factor receptor 2 (FGFR2) signaling is critical for proper craniofacial development. A gain-of-function mutation in the 2c splice variant of the receptor's gene is associated with Crouzon syndrome, which is characterized by craniosynostosis, the premature fusion of one or more of the cranial vault sutures, leading to craniofacial maldevelopment. Insight into the molecular mechanism of craniosynostosis has identified the ERK-MAPK signaling cascade as a critical regulator of suture patency. The aim of this study is to investigate the role of FGFR2c-induced ERK-MAPK activation in the regulation of coronal suture development. Loss-of-function and gain-of-function Fgfr2c mutant mice have overlapping phenotypes, including coronal synostosis and craniofacial dysmorphia. In vivo analysis of coronal sutures in loss-of-function and gain-of-function models demonstrated fundamentally different pathogenesis underlying coronal suture synostosis. Calvarial osteoblasts from gain-of-function mice demonstrated enhanced osteoblastic function and maturation with concomitant increase in ERK-MAPK activation. In vitro inhibition with the ERK protein inhibitor U0126 mitigated ERK protein activation levels with a concomitant reduction in alkaline phosphatase activity. This study identifies FGFR2c-mediated ERK-MAPK signaling as a key mediator of craniofacial growth and coronal suture development. Furthermore, our results solve the apparent paradox between loss-of-function and gain-of-function FGFR2c mutants with respect to coronal suture synostosis. PMID:27034231

  20. PI3K regulates BMAL1/CLOCK-mediated circadian transcription from the Dbp promoter.

    PubMed

    Morishita, Yoshikazu; Miura, Daiki; Kida, Satoshi

    2016-06-01

    The circadian rhythm generated by circadian clock underlies a molecular mechanism of rhythmic transcriptional regulation by transcription factor BMAL1/CLOCK. Importantly, the circadian clock is coordinated by exogenous cues to accommodate to changes in the external environment. However, the molecular mechanisms by which intracellular-signaling pathways mediate the adjustments of the circadian transcriptional rhythms remain unclear. In this study, we found that pharmacological inhibition or shRNA-mediated knockdown of phosphatidylinositol 3-kinase (PI3K) blocked upregulation of Dbp mRNA induced by serum shock in NIH 3T3 cells. Moreover, the inhibition of PI3K significantly reduced the promoter activity of the Dbp gene, as well as decreased the recruitment of BMAL1/CLOCK to the E-box in the Dbp promoter. Interestingly, the inhibition of PI3K blocked heterodimerization of BMAL1 and CLOCK. Our findings suggest that PI3K signaling plays a modulatory role in the regulation of the transcriptional rhythm of the Dbp gene by targeting BMAL1 and CLOCK. PMID:27022680

  1. Mammalian mediator of transcriptional regulation and its possible role as an end-point of signal transduction pathways.

    PubMed

    Jiang, Y W; Veschambre, P; Erdjument-Bromage, H; Tempst, P; Conaway, J W; Conaway, R C; Kornberg, R D

    1998-07-21

    A multiprotein complex isolated from murine cells is identified as a counterpart of the yeast Mediator of transcriptional regulation on the basis of the following: homologs of two subunits of yeast Mediator, Srb7 and Med7, copurify with the complex; peptide sequencing reveals, in addition, homologs of the yeast Mediator subunits Rgr1 and Med6; as with yeast Mediator, the mouse complex binds to the RNA polymerase II C-terminal domain (CTD) and stimulates phosphorylation of the CTD by TFIIH. Peptide sequencing also identifies a component of mouse Mediator as a relative of Ring-3 protein, a mitogen-activated nuclear protein kinase, raising the possibility of Mediator as an end point of signal transduction pathways. PMID:9671713

  2. Arginine methylation of HSP70 regulates retinoid acid-mediated RARβ2 gene activation

    PubMed Central

    Gao, Wei-wei; Xiao, Rong-quan; Peng, Bing-ling; Xu, Huan-teng; Shen, Hai-feng; Huang, Ming-feng; Shi, Tao-tao; Yi, Jia; Zhang, Wen-juan; Wu, Xiao-nan; Gao, Xiang; Lin, Xiang-zhi; Dorrestein, Pieter C.; Rosenfeld, Michael G.; Liu, Wen

    2015-01-01

    Although “histone” methyltransferases and demethylases are well established to regulate transcriptional programs and to use nonhistone proteins as substrates, their possible roles in regulation of heat-shock proteins in the nucleus have not been investigated. Here, we report that a highly conserved arginine residue, R469, in HSP70 (heat-shock protein of 70 kDa) proteins, an evolutionarily conserved protein family of ATP-dependent molecular chaperone, was monomethylated (me1), at least partially, by coactivator-associated arginine methyltransferase 1/protein arginine methyltransferase 4 (CARM1/PRMT4) and demethylated by jumonji-domain–containing 6 (JMJD6), both in vitro and in cultured cells. Functional studies revealed that HSP70 could directly regulate retinoid acid (RA)-induced retinoid acid receptor β2 (RARβ2) gene transcription through its binding to chromatin, with R469me1 being essential in this process. HSP70’s function in gene transcriptional regulation appears to be distinct from its protein chaperon activity. R469me1 was shown to mediate the interaction between HSP70 and TFIIH, which involves in RNA polymerase II phosphorylation and thus transcriptional initiation. Our findings expand the repertoire of nonhistone substrates targeted by PRMT4 and JMJD6, and reveal a new function of HSP70 proteins in gene transcription at the chromatin level aside from its classic role in protein folding and quality control. PMID:26080448

  3. Cytokinin as a mediator for regulating root system architecture in response to environmental cues.

    PubMed

    Ramireddy, Eswarayya; Chang, Ling; Schmülling, Thomas

    2014-01-01

    Root system architecture (RSA) is developmentally controlled by genetic pathways and their interaction with various environmental cues, in particular soil conditions. One important player in shaping RSA is the hormone cytokinin, which acts as a negative regulator of root elongation and branching. The redundant roles of cytokinin metabolism and signaling genes of Arabidopsis thaliana in regulating early stages of lateral root formation has recently been shown and it has been proposed that this redundancy reflects a role in mediating different environmental cues. Here we report that the transcript levels of cytokinin genes in the root responds to changes in nutrient availability in distinct ways. IPT3, IPT5, CYP735A2, LOG5, and CKX4 are particularly responsive cytokinin metabolism genes, genes encoding different type-A response regulators and the transcriptions factor genes ARR10 and CRF6 are among the most responsive signaling genes. This finding supports the hypothesis that environmental cues operate through fine-tuned transcriptional regulation of cytokinin genes to modulate root development. PMID:24509549

  4. Cellular context–mediated Akt dynamics regulates MAP kinase signaling thresholds during angiogenesis

    PubMed Central

    Hellesøy, Monica; Lorens, James B.

    2015-01-01

    The formation of new blood vessels by sprouting angiogenesis is tightly regulated by contextual cues that affect angiogeneic growth factor signaling. Both constitutive activation and loss of Akt kinase activity in endothelial cells impair angiogenesis, suggesting that Akt dynamics mediates contextual microenvironmental regulation. We explored the temporal regulation of Akt in endothelial cells during formation of capillary-like networks induced by cell–cell contact with vascular smooth muscle cells (vSMCs) and vSMC-associated VEGF. Expression of constitutively active Akt1 strongly inhibited network formation, whereas hemiphosphorylated Akt1 epi-alleles with reduced kinase activity had an intermediate inhibitory effect. Conversely, inhibition of Akt signaling did not affect endothelial cell migration or morphogenesis in vSMC cocultures that generate capillary-like structures. We found that endothelial Akt activity is transiently blocked by proteasomal degradation in the presence of SMCs during the initial phase of capillary-like structure formation. Suppressed Akt activity corresponded to the increased endothelial MAP kinase signaling that was required for angiogenic endothelial morphogenesis. These results reveal a regulatory principle by which cellular context regulates Akt protein dynamics, which determines MAP kinase signaling thresholds necessary drive a morphogenetic program during angiogenesis. PMID:26023089

  5. Loss of MBNL Leads to Disruption of Developmentally Regulated Alternative Polyadenylation in RNA-Mediated Disease

    PubMed Central

    Batra, Ranjan; Charizanis, Konstantinos; Manchanda, Mini; Mohan, Apoorva; Li, Moyi; Finn, Dustin J.; Goodwin, Marianne; Zhang, Chaolin; Sobczak, Krzysztof; Thornton, Charles A.; Swanson, Maurice S.

    2014-01-01

    SUMMARY Inhibition of muscleblind-like (MBNL) activity due to sequestration by microsatellite expansion RNAs is a major pathogenic event in the RNA-mediated disease myotonic dystrophy (DM). Although MBNL1 and MBNL2 bind to nascent transcripts to regulate alternative splicing during muscle and brain development, another major binding site for the MBNL protein family is the 3′ untranslated region of target RNAs. Here, we report that depletion of Mbnl proteins in mouse embryo fibroblasts leads to mis-regulation of thousands of alternative polyadenylation events. HITS-CLIP and minigene reporter analyses indicate that these polyadenylation switches are a direct consequence of MBNL binding to target RNAs. Mis-regulated alternative polyadenylation also occurs in skeletal muscle in a mouse polyCUG model and human DM resulting in the persistence of neonatal polyadenylation patterns. These findings reveal a novel developmental function for MBNL proteins and demonstrate that DM is characterized by mis-regulation of pre-mRNA processing at multiple levels. PMID:25263597

  6. Spliceosome-mediated decay (SMD) regulates expression of nonintronic genes in budding yeast

    PubMed Central

    Volanakis, Adam; Passoni, Monica; Hector, Ralph D.; Shah, Sneha; Kilchert, Cornelia; Granneman, Sander; Vasiljeva, Lidia

    2013-01-01

    We uncovered a novel role for the spliceosome in regulating mRNA expression levels that involves splicing coupled to RNA decay, which we refer to as spliceosome-mediated decay (SMD). Our transcriptome-wide studies identified numerous transcripts that are not known to have introns but are spliced by the spliceosome at canonical splice sites in Saccharomyces cerevisiae. Products of SMD are primarily degraded by the nuclear RNA surveillance machinery. We demonstrate that SMD can significantly down-regulate mRNA levels; splicing at canonical splice sites in the bromodomain factor 2 (BDF2) transcript reduced transcript levels roughly threefold by generating unstable products that are rapidly degraded by the nuclear surveillance machinery. Regulation of BDF2 mRNA levels by SMD requires Bdf1, a functionally redundant Bdf2 paralog that plays a role in recruiting the spliceosome to the BDF2 mRNA. Interestingly, mutating BDF2 5′ splice site and branch point consensus sequences partially suppresses the bdf1Δ temperature-sensitive phenotype, suggesting that maintaining proper levels of Bdf2 via SMD is biologically important. We propose that the spliceosome can also repress protein-coding gene expression by promoting nuclear turnover of spliced RNA products and provide an insight for coordinated regulation of Bdf1 and Bdf2 levels in the cell. PMID:24065768

  7. Op18/Stathmin Mediates Multiple Region-Specific Tubulin and Microtubule-Regulating Activities

    PubMed Central

    Larsson, Niklas; Segerman, Bo; Howell, Bonnie; Fridell, Kajsa; Cassimeris, Lynne; Gullberg, Martin

    1999-01-01

    Oncoprotein18/stathmin (Op18) is a regulator of microtubule (MT) dynamics that binds tubulin heterodimers and destabilizes MTs by promoting catastrophes (i.e., transitions from growing to shrinking MTs). Here, we have performed a deletion analysis to mechanistically dissect Op18 with respect to (a) modulation of tubulin GTP hydrolysis and exchange, (b) tubulin binding in vitro, and (c) tubulin association and MT-regulating activities in intact cells. The data reveal distinct types of region-specific Op18 modulation of tubulin GTP metabolism, namely inhibition of nucleotide exchange and stimulation or inhibition of GTP hydrolysis. These regulatory activities are mediated via two-site cooperative binding to tubulin by multiple nonessential physically separated regions of Op18. In vitro analysis revealed that NH2- and COOH-terminal truncations of Op18 have opposite effects on the rates of tubulin GTP hydrolysis. Transfection of human leukemia cells with these two types of mutants result in similar decrease of MT content, which in both cases appeared independent of a simple tubulin sequestering mechanism. However, the NH2- and COOH-terminal–truncated Op18 mutants regulate MTs by distinct mechanisms as evidenced by morphological analysis of microinjected newt lung cells. Hence, mutant analysis shows that Op18 has the potential to regulate tubulin/MTs by more than one specific mechanism. PMID:10491392

  8. Role of TARP interaction in S-SCAM-mediated regulation of AMPA receptors.

    PubMed

    Danielson, Eric; Metallo, Jacob; Lee, Sang H

    2012-01-01

    Scaffolding proteins are involved in the incorporation, anchoring, maintenance, and removal of AMPA receptors (AMPARs) at synapses, either through a direct interaction with AMPARs or via indirect association through auxiliary subunits of transmembrane AMPAR regulatory proteins (TARPs). Synaptic scaffolding molecule (S-SCAM) is a newly characterized member of the scaffolding proteins critical for the regulation and maintenance of AMPAR levels at synapses, and directly binds to TARPs through a PDZ interaction. However, the functional significance of S-SCAM-TARP interaction in the regulation of AMPARs has not been tested. Here we show that overexpression of the C-terminal peptide of TARP-γ2 fused to EGFP abolished the S-SCAM-mediated enhancement of surface GluA2 expression. Conversely, the deletion of the PDZ-5 domain of S-SCAM that binds TARPs greatly attenuated the S-SCAM-induced increase of surface GluA2 expression. In contrast, the deletion of the guanylate kinase domain of S-SCAM did not show a significant effect on the regulation of AMPARs. Together, these results suggest that S-SCAM is regulating AMPARs through TARPs. PMID:22878254

  9. Dynamic regulation of partner abundance mediates response of reef coral symbioses to environmental change.

    PubMed

    Cunning, R; Vaughan, N; Gillette, P; Capo, T R; Matté, J L; Baker, A C

    2015-05-01

    Regulating partner abunclance may allow symmotic organisms to mediate interaction outcomes, facilitating adaptive responses to environmental change. To explore the capacity for-adaptive regulation in an ecologically important endosymbiosis, we studied the population dynamics of symbiotic algae in reef-building corals under different abiotic contexts. We found high natural variability in symbiont abundance in corals across reefs, but this variability converged to different symbiont-specific abundances when colonies were maintained under constant conditions. When conditions changed seasonally, symbiont abundance readjusted to new equilibria. We explain these patterns using an a priori model of symbiotic costs and benefits to the coral host, which shows that the observed changes in symbiont abundance are consistent with the maximization of interaction benefit under different environmental conditions. These results indicate that, while regulating symbiont abundance helps hosts sustain maximum benefit in a dynamic environment, spatiotemporal variation in abiotic factors creates a broad range of symbiont abundances (and interaction outcomes) among corals that may account for observed natural variability in performance (e.g., growth rate) and stress tolerance (e.g., bleaching susceptibility). This cost or benefit framework provides a new perspective on the dynamic regulation of reef coral symbioses and illustrates that the dependence of interaction outcomes on biotic and abiotic contexts may be important in understanding how diverse mutualisms respond to environmental change. PMID:26236853

  10. Blimp-1-Dependent IL-10 Production by Tr1 Cells Regulates TNF-Mediated Tissue Pathology

    PubMed Central

    Montes de Oca, Marcela; Kumar, Rajiv; de Labastida Rivera, Fabian; Amante, Fiona H; Sheel, Meru; Faleiro, Rebecca J.; Bunn, Patrick T.; Best, Shannon E.; Beattie, Lynette; Ng, Susanna S.; Edwards, Chelsea L.; Muller, Werner; Cretney, Erika; Nutt, Stephen L.; Smyth, Mark J.; Haque, Ashraful; Hill, Geoffrey R.; Sundar, Shyam; Kallies, Axel; Engwerda, Christian R.

    2016-01-01

    Tumor necrosis factor (TNF) is critical for controlling many intracellular infections, but can also contribute to inflammation. It can promote the destruction of important cell populations and trigger dramatic tissue remodeling following establishment of chronic disease. Therefore, a better understanding of TNF regulation is needed to allow pathogen control without causing or exacerbating disease. IL-10 is an important regulatory cytokine with broad activities, including the suppression of inflammation. IL-10 is produced by different immune cells; however, its regulation and function appears to be cell-specific and context-dependent. Recently, IL-10 produced by Th1 (Tr1) cells was shown to protect host tissues from inflammation induced following infection. Here, we identify a novel pathway of TNF regulation by IL-10 from Tr1 cells during parasitic infection. We report elevated Blimp-1 mRNA levels in CD4+ T cells from visceral leishmaniasis (VL) patients, and demonstrate IL-12 was essential for Blimp-1 expression and Tr1 cell development in experimental VL. Critically, we show Blimp-1-dependent IL-10 production by Tr1 cells prevents tissue damage caused by IFNγ-dependent TNF production. Therefore, we identify Blimp-1-dependent IL-10 produced by Tr1 cells as a key regulator of TNF-mediated pathology and identify Tr1 cells as potential therapeutic tools to control inflammation. PMID:26765224

  11. Comparative transcriptomics of Central Asian Vitis vinifera accessions reveals distinct defense strategies against powdery mildew

    PubMed Central

    Amrine, Katherine C H; Blanco-Ulate, Barbara; Riaz, Summaira; Pap, Dániel; Jones, Laura; Figueroa-Balderas, Rosa; Walker, M Andrew; Cantu, Dario

    2015-01-01

    Grape powdery mildew (PM), caused by the biotrophic ascomycete Erysiphe necator, is a devastating fungal disease that affects most Vitis vinifera cultivars. We have previously identified a panel of V. vinifera accessions from Central Asia with partial resistance to PM that possess a Ren1-like local haplotype. In this study, we show that in addition to the typical Ren1-associated late post-penetration resistance, these accessions display a range of different levels of disease development suggesting that alternative alleles or additional genes contribute to determining the outcome of the interaction with the pathogen. To identify potential Ren1-dependent transcriptional responses and functions associated with the different levels of resistance, we sequenced and analyzed the transcriptomes of these Central Asian accessions at two time points of PM infection. Transcriptomes were compared to identify constitutive differences and PM-inducible responses that may underlie their disease resistant phenotype. Responses to E. necator in all resistant accessions were characterized by an early up-regulation of 13 genes, most encoding putative defense functions, and a late down-regulation of 32 genes, enriched in transcriptional regulators and protein kinases. Potential Ren1-dependent responses included a hotspot of co-regulated genes on chromosome 18. We also identified 81 genes whose expression levels and dynamics correlated with the phenotypic differences between the most resistant accessions ‘Karadzhandahal’, DVIT3351.27, and O34-16 and the other genotypes. This study provides a first exploration of the functions associated with varying levels of partial resistance to PM in V. vinifera accessions that can be exploited as sources of genetic resistance in grape breeding programs. PMID:26504579

  12. Regulation of Arabidopsis defense responses against Spodoptera littoralis by CPK-mediated calcium signaling

    PubMed Central

    2010-01-01

    Background Plant Ca2+ signals are involved in a wide array of intracellular signaling pathways after pest invasion. Ca2+-binding sensory proteins such as Ca2+-dependent protein kinases (CPKs) have been predicted to mediate the signaling following Ca2+ influx after insect herbivory. However, until now this prediction was not testable. Results To investigate the roles CPKs play in a herbivore response-signaling pathway, we screened the characteristics of Arabidopsis CPK mutants damaged by a feeding generalist herbivore, Spodoptera littoralis. Following insect attack, the cpk3 and cpk13 mutants showed lower transcript levels of plant defensin gene PDF1.2 compared to wild-type plants. The CPK cascade was not directly linked to the herbivory-induced signaling pathways that were mediated by defense-related phytohormones such as jasmonic acid and ethylene. CPK3 was also suggested to be involved in a negative feedback regulation of the cytosolic Ca2+ levels after herbivory and wounding damage. In vitro kinase assays of CPK3 protein with a suite of substrates demonstrated that the protein phosphorylates transcription factors (including ERF1, HsfB2a and CZF1/ZFAR1) in the presence of Ca2+. CPK13 strongly phosphorylated only HsfB2a, irrespective of the presence of Ca2+. Furthermore, in vivo agroinfiltration assays showed that CPK3-or CPK13-derived phosphorylation of a heat shock factor (HsfB2a) promotes PDF1.2 transcriptional activation in the defense response. Conclusions These results reveal the involvement of two Arabidopsis CPKs (CPK3 and CPK13) in the herbivory-induced signaling network via HsfB2a-mediated regulation of the defense-related transcriptional machinery. This cascade is not involved in the phytohormone-related signaling pathways, but rather directly impacts transcription factors for defense responses. PMID:20504319

  13. N-Cadherin-Mediated Signaling Regulates Cell Phenotype for Nucleus Pulposus Cells of the Intervertebral Disc

    PubMed Central

    Hwang, Priscilla Y.; Jing, Liufang; Michael, Keith W.; Richardson, William J.; Chen, Jun; Setton, Lori A.

    2015-01-01

    Juvenile nucleus pulposus (NP) cells of the intervertebral disc (IVD) are large, vacuolated cells that form cell clusters with strong cell–cell interactions. With maturation and aging, NP cells lose their ability to form these cell clusters, with aging-associated changes in NP cell phenotype, morphology, and proteoglycan synthesis that may contribute to IVD degeneration. Therefore, it is important to understand the mechanisms governing juvenile NP cell cluster behavior towards the goal of revealing factors that can promote juvenile, healthy NP cell phenotypes. N-cadherin has been identified as a cell–cell adhesion marker that is present in juvenile NP cells, but disappears with age. The goal of this study was to reveal the importance of N-cadherin in regulating cell–cell interactions in juvenile NP cell cluster formation and test for a regulatory role in maintaining a juvenile NP phenotype in vitro. Juvenile porcine IVD cells, of notochordal origin, were promoted to form cell clusters in vitro, and analyzed for preservation of the juvenile NP phenotype. Additionally, cadherin-blocking experiments were performed to prevent cluster formation in order to study the importance of cluster formation in NP cell signaling. Findings reveal N-cadherin-mediated cell–cell contacts promote cell clustering behavior and regulate NP cell matrix production and preservation of NP-specific markers. Inhibition of N-cadherin-mediated contacts resulted in loss of all features of the juvenile NP cell. These results establish a regulatory role for N-cadherin in juvenile NP cells, and suggest that preservation of the N-cadherin mediated cell–cell contact is important for preserving juvenile NP cell phenotype and morphology. PMID:25848407

  14. Epigenetic Regulation in Particulate Matter-Mediated Cardiopulmonary Toxicities: A Systems Biology Perspective.

    PubMed

    Wang, Ting; Garcia, Joe Gn; Zhang, Wei

    2012-12-01

    Particulate matter (PM) air pollution exerts significant adverse health effects in global populations, particularly in developing countries with extensive air pollution. Understanding of the mechanisms of PM-induced health effects including the risk for cardiovascular diseases remains limited. In addition to the direct cellular physiological responses such as mitochondrial dysfunction and oxidative stress, PM mediates remarkable dysregulation of gene expression, especially in cardiovascular tissues. The PM-mediated gene dysregulation is likely to be a complex mechanism affected by various genetic and non-genetic factors. Notably, PM is known to alter epigenetic markers (e.g., DNA methylation and histone modifications), which may contribute to air pollution-mediated health consequences including the risk for cardiovascular diseases. Notably, epigenetic changes induced by ambient PM exposure have emerged to play a critical role in gene regulation. Though the underlying mechanism(s) are not completely clear, the available evidence suggests that the modulated activities of DNA methyltransferase (DNMT), histone acetylase (HAT) and histone deacetylase (HDAC) may contribute to the epigenetic changes induced by PM or PM-related chemicals. By employing genome-wide epigenomic and systems biology approaches, PM toxicogenomics could conceivably progress greatly with the potential identification of individual epigenetic loci associated with dysregulated gene expression after PM exposure, as well the interactions between epigenetic pathways and PM. Furthermore, novel therapeutic targets based on epigenetic markers could be identified through future epigenomic studies on PM-mediated cardiopulmonary toxicities. These considerations collectively inform the future population health applications of genomics in developing countries while benefiting global personalized medicine at the same time. PMID:23185213

  15. Mechanisms underlying the protein-kinase mediated regulation of the HERG potassium channel synthesis

    PubMed Central

    Krishnan, Yamini; Li, Yan; Zheng, Renjian; Kanda, Vikram; McDonald, Thomas V.

    2012-01-01

    The HERG (human ether-a-go-go related gene) potassium channel aids in repolarization of the cardiomyocyte membrane at the end of each action potential. We have previously shown that sustained protein kinase A or C (PKA and PKC) activity specifically enhances channel synthesis over the course of hours to days in heterologous expression and cardiac myocytes. The kinase-mediated augmentation of the channel is post-transcriptional and occurs near or at the endoplasmic reticulum. Here we report our further investigations into the mechanisms of kinase-mediated augmentation of HERG channel protein. We show that HERG channel phosphorylation alone is not sufficient for the PKA-dependent increase to occur. In vitro translation studies indicate that an additional factor is required for the process. Pharmacologic inhibitors suggest that the channel augmentation is not due to kinase-mediated alteration in proteasome or lysosome activity. PKA activation had no effect on stability of HERG mRNA and polyribosomal profiling showed that kinase activity did not elevate translation from low to high rates. Transcriptional inhibition results suggest that the additional cellular factor is a PKA-regulated protein. Together, these findings suggest that PKA-mediated augmentation of HERG abundance is more complex than previously appreciated involving enhancement of already active translation rates, phosphorylation of the channel protein and at least one other cAMP/PKA-responsive protein. Further exploration of molecular components of this regulatory pathway will be necessary to determine exact mechanism and the biomedical impact of this process in vivo. PMID:22613764

  16. KSHV-Mediated Regulation of Par3 and SNAIL Contributes to B-Cell Proliferation.

    PubMed

    Jha, Hem C; Sun, Zhiguo; Upadhyay, Santosh K; El-Naccache, Darine W; Singh, Rajnish K; Sahu, Sushil K; Robertson, Erle S

    2016-07-01

    Studies have suggested that Epithelial-Mesenchymal Transition (EMT) and transformation is an important step in progression to cancer. Par3 (partitioning-defective protein) is a crucial factor in regulating epithelial cell polarity. However, the mechanism by which the latency associated nuclear antigen (LANA) encoded by Kaposi's Sarcoma associated herpesvirus (KSHV) regulates Par3 and EMTs markers (Epithelial-Mesenchymal Transition) during viral-mediated B-cell oncogenesis has not been fully explored. Moreover, several studies have demonstrated a crucial role for EMT markers during B-cell malignancies. In this study, we demonstrate that Par3 is significantly up-regulated in KSHV-infected primary B-cells. Further, Par3 interacted with LANA in KSHV positive and LANA expressing cells which led to translocation of Par3 from the cell periphery to a predominantly nuclear signal. Par3 knockdown led to reduced cell proliferation and increased apoptotic induction. Levels of SNAIL was elevated, and E-cadherin was reduced in the presence of LANA or Par3. Interestingly, KSHV infection in primary B-cells led to enhancement of SNAIL and down-regulation of E-cadherin in a temporal manner. Importantly, knockdown of SNAIL, a major EMT regulator, in KSHV cells resulted in reduced expression of LANA, Par3, and enhanced E-cadherin. Also, SNAIL bound to the promoter region of p21 and can regulate its activity. Further a SNAIL inhibitor diminished NF-kB signaling through upregulation of Caspase3 in KSHV positive cells in vitro. This was also supported by upregulation of SNAIL and Par3 in BC-3 transplanted NOD-SCID mice which has potential as a therapeutic target for KSHV-associated B-cell lymphomas. PMID:27463802

  17. KSHV-Mediated Regulation of Par3 and SNAIL Contributes to B-Cell Proliferation

    PubMed Central

    Jha, Hem C.; Sun, Zhiguo; Upadhyay, Santosh K.; El-Naccache, Darine W.; Singh, Rajnish K.; Sahu, Sushil K.; Robertson, Erle S.

    2016-01-01

    Studies have suggested that Epithelial–Mesenchymal Transition (EMT) and transformation is an important step in progression to cancer. Par3 (partitioning-defective protein) is a crucial factor in regulating epithelial cell polarity. However, the mechanism by which the latency associated nuclear antigen (LANA) encoded by Kaposi's Sarcoma associated herpesvirus (KSHV) regulates Par3 and EMTs markers (Epithelial-Mesenchymal Transition) during viral-mediated B-cell oncogenesis has not been fully explored. Moreover, several studies have demonstrated a crucial role for EMT markers during B-cell malignancies. In this study, we demonstrate that Par3 is significantly up-regulated in KSHV-infected primary B-cells. Further, Par3 interacted with LANA in KSHV positive and LANA expressing cells which led to translocation of Par3 from the cell periphery to a predominantly nuclear signal. Par3 knockdown led to reduced cell proliferation and increased apoptotic induction. Levels of SNAIL was elevated, and E-cadherin was reduced in the presence of LANA or Par3. Interestingly, KSHV infection in primary B-cells led to enhancement of SNAIL and down-regulation of E-cadherin in a temporal manner. Importantly, knockdown of SNAIL, a major EMT regulator, in KSHV cells resulted in reduced expression of LANA, Par3, and enhanced E-cadherin. Also, SNAIL bound to the promoter region of p21 and can regulate its activity. Further a SNAIL inhibitor diminished NF-kB signaling through upregulation of Caspase3 in KSHV positive cells in vitro. This was also supported by upregulation of SNAIL and Par3 in BC-3 transplanted NOD-SCID mice which has potential as a therapeutic target for KSHV-associated B-cell lymphomas. PMID:27463802

  18. Are ineffective defence reactions potential target for induced resistance during the compatible wheat-powdery mildew interaction?

    PubMed

    Tayeh, Ch; Randoux, B; Tisserant, B; Khong, G; Jacques, Ph; Reignault, Ph

    2015-11-01

    Powdery mildew caused by Blumeria graminis f.sp. tritici, an obligate aerial biotrophic fungus, would be one of the most damaging wheat (Triticum aestivum) diseases without the extensive use of conventional fungicides. In our study, the expression levels of some basal defence-related genes were investigated during a compatible interaction in order to evaluate wheat reactions to infection, along with the different stages of the infectious process in planta. As fungal conidia initiated their germination and developed appressorial germ tube (AGT), early defence reactions involved the expression of a lipoxygenase (LOX)- and an oxalate oxidase (OXO)-encoding genes, followed by activations of corresponding LOX (EC 1.13.11.12) and OXO (EC 1.2.3.4) activities, respectively. When penetration of AGT took place, up-regulation of chitinases (CHI) and PR1-encoding genes expression occurred along with an increase of CHI (EC 3.2.1.14) activity. Meanwhile, expression of a phenylalanine ammonia-lyase-encoding gene also took place. Up-regulation of a phospholipase C- and lipid transfer proteins-encoding genes expression occurred during the latest stages of infection. Neither the phi glutathione S-transferase (GST)-encoding gene expression nor the GST (EC 2.5.1.13) activity was modified upon wheat infection by powdery mildew. Whether these defence reactions during such a compatible interaction are markers of immunity or susceptibility, and whether they have the ability to contribute to protection upon modulation of their timing and their intensity by resistance inducers are discussed. PMID:26218548

  19. Antisense expression of peach mildew resistance locus O (PpMlo1) gene confers cross-species resistance to powdery mildew in Fragaria x ananassa.

    PubMed

    Jiwan, Derick; Roalson, Eric H; Main, Dorrie; Dhingra, Amit

    2013-12-01

    Powdery mildew (PM) is one of the major plant pathogens. The conventional method of PM control includes frequent use of sulfur-based fungicides adding to production costs and potential harm to the environment. PM remains a major scourge for Rosaceae crops where breeding approaches mainly resort to gene-for-gene resistance. We have tested an alternate source of PM resistance in Rosaceae. Mildew resistance locus O (MLO) has been well studied in barley due to its role in imparting broad spectrum resistance to PM. We identified PpMlo1 (Prunus persica Mlo) in peach and characterized it further to test if a similar mechanism of resistance is conserved in Rosaceae. Due to its recalcitrance in tissue culture, reverse genetic studies involving PpMloI were not feasible in peach. Therefore, Fragaria x ananassa LF9 line, a taxonomic surrogate, was used for functional analysis of PpMlo1. Agrobacterium-mediated transformation yielded transgenic strawberry plants expressing PpMlo1 in sense and antisense orientation. Antisense expression of PpMlo1 in transgenic strawberry plants conferred resistance to Fragaria-specific powdery mildew, Podosphaera macularis. Phylogenetic analysis of 208 putative Mlo gene copies from 35 plant species suggests a large number of duplications of this gene family prior to the divergence of monocots and eudicots, early in eudicot diversification. Our results indicate that the Mlo-based resistance mechanism is functional in Rosaceae, and that Fragaria can be used as a host to test mechanistic function of genes derived from related tree species. To the best of our knowledge, this work is one of the first attempts at testing the potential of using a Mlo-based resistance strategy to combat powdery mildew in Rosaceae. PMID:23728780

  20. Peroxisome proliferators and fatty acids negatively regulate liver X receptor-mediated activity and sterol biosynthesis.

    PubMed

    Johnson, T E; Ledwith, B J

    2001-04-01

    Peroxisome proliferators (PPs) are potent tumor promoters in rodents. The mechanism of hepatocarcinogenesis requires the nuclear receptor peroxisome proliferator activated receptor-alpha (PPARalpha), but might also involve the PPARalpha independent alteration of signaling pathways that regulate cell growth. Here, we studied the effects of PPs on the mevalonate pathway, a critical pathway that controls cell proliferation. Liver X receptors (LXRs) are nuclear receptors that act as sterol sensors in the mevalonate pathway. In gene reporter assays in COS-7 cells, the basal activity of the LXR responsive reporter gene (LXRE-luc) was suppressed by 10 microM lovastatin and zaragozic acid A, suggesting that this activity was attributed to the activation of native LXRs, by endogenously produced mevalonate products. The potent PP and rodent tumor promoter, pirinixic acid (WY-14643) also inhibited LXR-mediated transcription in a dose related manner (approximate IC(50) of 100 microM). As did several other PPs including ciprofibric acid and mono-ethylhexylphthalate. Polyunsaturated and medium to long chain fatty acids at 100 microM were also potent inhibitors; the arachidonic acid analogue eicosatetraynoic acid being the most active (approximate IC(50) of 10 microM). Of the PPs and fatty acids tested, there was a strong correlation between the ability of these agents to suppress de novo sterol synthesis in a rat hepatoma cell line, H4IIEC3, and inhibit LXR-mediated transcription in COS-7 cells, but a discordance between these endpoints and PPARalpha activation and fatty acid acyl-CoA oxidase induction. Taken together, these results suggest that PPs and fatty acids negatively regulate the mevalonate pathway through a mechanism that is not entirely dependent on PPARalpha activation. Because of the importance of the mevalonate pathway in regulating cell proliferation, the modulation of this pathway by PPs and fatty acids might contribute to their actions on cell growth

  1. Using Temperature-Sensitive Smart Polymers to Regulate DNA-mediated Nanoassembly

    NASA Astrophysics Data System (ADS)

    Hamner, Kristen L.

    Nanoparticle (NP) self-assembly has been proven as an effective route to organize nanoscale building blocks into ordered structures for potential technological applications. In order to successfully exploit the self-assembly processes a high level of direction and control is required. In my dissertation research, I synthesized a temperature responsive copolymer (p) to modify gold nanoparticles (AuNP) for controlling self-assembly. The copolymers' ability to regulate DNA-mediated NP self-assembly is a particular focus. In Chapter 2, the results show that by the addition of the p to create thermally responsive NP interfaces allows for controlled aggregation behavior and interparticle distances defined by the transition temperature (TC) of the p, to aid in NP assembly and help to regulate DNA-mediated interactions between NP. The work in Chapter 3 revealed that the reconfigurable conformation of the p sterically regulates the assembly: at T < TC, the chains extended beyond the hydrodynamic reach of the single stranded DNA and prohibited recognition, while at T > TC, assembly was observed, due the hydrophobic collapse of the p and the subsequent exposure of the complementary DNA bases. In Chapter 4, to gain insight into the mechanism, the rate of assembly was monitored, with DNA lengths that had hydrodynamic diameters more comparable to that of the p, and found the p was capable of slowing the kinetics. I further investigated to find that the addition of p extended the interparticle distances while disrupting the long range ordering. Finally, how the temperature responsive behavior of the p acted on the interparticle distances was probed, and it was found that without p, the interparticle distances expanded, while the addition of p compressed the interparticle distances.

  2. STIM1/ORAI1-mediated Ca2+ Influx Regulates Enolase-1 Exteriorization*

    PubMed Central

    Didiasova, Miroslava; Zakrzewicz, Dariusz; Magdolen, Viktor; Nagaraj, Chandran; Bálint, Zoltán; Rohde, Manfred; Preissner, Klaus T.; Wygrecka, Malgorzata

    2015-01-01

    Tumor cells use broad spectrum proteolytic activity of plasmin to invade tissue and form metastatic foci. Cell surface-associated enolase-1 (ENO-1) enhances plasmin formation and thus participates in the regulation of pericellular proteolysis. Although increased levels of cell surface bound ENO-1 have been described in different types of cancer, the molecular mechanism responsible for ENO-1 exteriorization remains elusive. In the present study, increased ENO-1 protein levels were found in ductal breast carcinoma and on the cell surface of highly metastatic breast cancer cell line MDA-MB-231. Elevated cell surface-associated ENO-1 expression correlated with augmented MDA-MB-231 cell migratory and invasive properties. Exposure of MDA-MB-231 cells to LPS potentiated translocation of ENO-1 to the cell surface and its release into the extracellular space in the form of exosomes. These effects were independent of de novo protein synthesis and did not require the classical endoplasmic reticulum/Golgi pathway. LPS-triggered ENO-1 exteriorization was suppressed by pretreatment of MDA-MB-231 cells with the Ca2+ chelator BAPTA or an inhibitor of endoplasmic reticulum Ca2+-ATPase pump, cyclopiazonic acid. In line with these observations, the stromal interaction molecule (STIM) 1 and the calcium release-activated calcium modulator (ORAI) 1-mediated store-operated Ca2+ entry were found to regulate LPS-induced ENO-1 exteriorization. Pharmacological blockage or knockdown of STIM1 or ORAI1 reduced ENO-1-dependent migration of MDA-MB-231 cells. Collectively, our results demonstrate the pivotal role of store-operated Ca2+ channel-mediated Ca2+ influx in the regulation of ENO-1 exteriorization and thus in the modulation of cancer cell migratory and invasive properties. PMID:25805497

  3. Glucocorticoid mediated regulation of inflammation in human monocytes is associated with depressive mood and obesity.

    PubMed

    Cheng, Tiefu; Dimitrov, Stoyan; Pruitt, Christopher; Hong, Suzi

    2016-04-01

    Dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis is observed in various conditions, including depression and obesity, which are also often related. Glucocorticoid (GC) resistance and desensitization of peripheral GC receptors (GRs) are often the case in HPA dysregulation seen in depression, and GC plays a critical role in regulation of inflammation. Given the growing evidence that inflammation is a central feature of some depression cases and obesity, we aimed to investigate the immune-regulatory role of GC-GR in relation to depressive mood and obesity in 35 healthy men and women. Depressive mood and level of obesity were assessed, using Beck Depression Inventory (BDI-Ia) and body mass index (BMI), respectively. We measured plasma cortisol levels via enzyme-linked immunosorbent assay and lipopolysaccharide-stimulated intracellular tumor necrosis factor (TNF) production by monocytes, using flow cytometry. Cortisol sensitivity was determined by the difference in monocytic TNF production between the conditions of 1 and 0μM cortisol incubation ("cortisol-mediated inflammation regulation, CoMIR"). GR vs. mineralocorticoid receptor (MR) antagonism for CoMIR was examined by using mifepristone and spironolactone. A series of multiple regression analyses were performed to investigate independent contribution of depressive mood vs. obesity after controlling for age, gender, systolic blood pressure (SBP), and plasma cortisol in predicting CoMIR. CoMIR was explained by somatic subcomponents of depressive mood (BDI-S: β=-0.499, p=0.001), or BMI (β=-0.466, p<0.01) in separate models. The effects of BMI disappeared when BDI-S was controlled for in the model, while BDI-S remained a significant independent predictor for CoMIR (β=-0.369, p<0.05). However, BMI remained the only independent predictor when BDI-T or BDI-C were controlled for in the model. Mediation analyses also revealed that the relationship between BMI and CoMIR was mediated by BDI-S. The exploratory

  4. MlAB10: a Triticum turgidum subsp. dicoccoides derived powdery mildew resistance gene identified in common wheat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew is an economically important disease in wheat growing areas with a cool maritime environment. Host genetic resistance is the most economical, consistent, and environmentally sound method of control. NC97BGTAB10 is a germplasm line containing powdery mildew resistance introgressed fr...

  5. MIAG12: A Triticum timopheevii-derived powdery mildew resistance gene in common wheat on chromosome 7AL

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Wheat powdery mildew is an economically important disease in cool and humid 2 environments. Powdery mildew causes yield losses as high as 48 percent through a reduction in 3 tiller survival, kernels per head and kernel size. Race-specific host resistance is the most 4 consistent, environmentally fri...

  6. MlNCD1: A novel Aegilops tauschii derived powdery mildew resistance gene identified in common wheat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew is a major fungal disease in wheat, especially in cool maritime climates. A novel Aegilops tauschii derived wheat powdery mildew resistance gene present in the germplasm line NC96BGTD1 was genetically characterized as a monogenic trait in field trials using F2 and F4-derived lines fr...

  7. Confirmation of Three Quantitative Trait Loci Conferring Adult Plant Resistance to Powdery Mildew in Two Winter Wheat Populations

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Hypersensitive, race specific genes primarily have been deployed to control powdery mildew (caused by Blumeria graminis) in wheat (Triticum aestivum); however, recent efforts have shifted to breeding for more durable resistance. Previously, three QTL for adult plant resistance (APR) to powdery milde...

  8. MicroRNA-146a: A Key Regulator of Astrocyte-Mediated Inflammatory Response

    PubMed Central

    Prabowo, Avanita; Fluiter, Kees; Spliet, Wim G. M.; van Rijen, Peter C.; Gorter, Jan A.; Aronica, Eleonora

    2012-01-01

    Increasing evidence supports the involvement of microRNAs (miRNA) in the regulation of inflammation in human neurological disorders. In the present study we investigated the role of miR-146a, a key regulator of the innate immune response, in the modulation of astrocyte-mediated inflammation. Using Taqman PCR and in situ hybridization, we studied the expression of miR-146a in epilepsy-associated glioneuronal lesions which are characterized by prominent activation of the innate immune response. In addition, cultured human astrocytes were used to study the regulation of miR-146a expression in response to proinflammatory cytokines. qPCR and western blot were used to evaluate the effects of overexpression or knockdown of miR-146a on IL-1β signaling. Downstream signaling in the IL-1β pathway, as well as the expression of IL-6 and COX-2 were evaluated by western blot and ELISA. Release several cytokines was evaluated using a human magnetic multiplex cytokine assay on a Luminex® 100™/200™ platform. Increased expression of miR-146a was observed in glioneuronal lesions by Taqman PCR. MiR-146a expression in human glial cell cultures was strongly induced by IL-1β and blocked by IL-1β receptor antagonist. Modulation of miR-146a expression by transfection of astrocytes with anti-miR146a or mimic, regulated the mRNA expression levels of downstream targets of miR-146a (IRAK-1, IRAK-2 and TRAF-6) and the expression of IRAK-1 protein. In addition, the expression of IL-6 and COX-2 upon IL-1β stimulation was suppressed by increased levels of miR-146a and increased by the reduction of miR-146a. Modulation of miR-146a expression affected also the release of several cytokines such as IL-6 and TNF-α. Our observations indicate that in response to inflammatory cues, miR-146a was induced as a negative-feedback regulator of the astrocyte-mediated inflammatory response. This supports an important role of miR-146a in human neurological disorders associated with chronic inflammation

  9. MicroRNA-146a: a key regulator of astrocyte-mediated inflammatory response.

    PubMed

    Iyer, Anand; Zurolo, Emanuele; Prabowo, Avanita; Fluiter, Kees; Spliet, Wim G M; van Rijen, Peter C; Gorter, Jan A; Aronica, Eleonora

    2012-01-01

    Increasing evidence supports the involvement of microRNAs (miRNA) in the regulation of inflammation in human neurological disorders. In the present study we investigated the role of miR-146a, a key regulator of the innate immune response, in the modulation of astrocyte-mediated inflammation. Using Taqman PCR and in situ hybridization, we studied the expression of miR-146a in epilepsy-associated glioneuronal lesions which are characterized by prominent activation of the innate immune response. In addition, cultured human astrocytes were used to study the regulation of miR-146a expression in response to proinflammatory cytokines. qPCR and western blot were used to evaluate the effects of overexpression or knockdown of miR-146a on IL-1β signaling. Downstream signaling in the IL-1β pathway, as well as the expression of IL-6 and COX-2 were evaluated by western blot and ELISA. Release several cytokines was evaluated using a human magnetic multiplex cytokine assay on a Luminex® 100™/200™ platform. Increased expression of miR-146a was observed in glioneuronal lesions by Taqman PCR. MiR-146a expression in human glial cell cultures was strongly induced by IL-1β and blocked by IL-1β receptor antagonist. Modulation of miR-146a expression by transfection of astrocytes with anti-miR146a or mimic, regulated the mRNA expression levels of downstream targets of miR-146a (IRAK-1, IRAK-2 and TRAF-6) and the expression of IRAK-1 protein. In addition, the expression of IL-6 and COX-2 upon IL-1β stimulation was suppressed by increased levels of miR-146a and increased by the reduction of miR-146a. Modulation of miR-146a expression affected also the release of several cytokines such as IL-6 and TNF-α. Our observations indicate that in response to inflammatory cues, miR-146a was induced as a negative-feedback regulator of the astrocyte-mediated inflammatory response. This supports an important role of miR-146a in human neurological disorders associated with chronic inflammation

  10. Biotrophy at Its Best: Novel Findings and Unsolved Mysteries of the Arabidopsis-Powdery Mildew Pathosystem.

    PubMed

    Kuhn, Hannah; Kwaaitaal, Mark; Kusch, Stefan; Acevedo-Garcia, Johanna; Wu, Hongpo; Panstruga, Ralph

    2016-01-01

    It is generally accepted in plant-microbe interactions research that disease is the exception rather than a common outcome of pathogen attack. However, in nature, plants with symptoms that signify colonization by obligate biotrophic powdery mildew fungi are omnipresent. The pervasiveness of the disease and the fact that many economically important plants are prone to infection by powdery mildew fungi drives research on this interaction. The competence of powdery mildew fungi to establish and maintain true biotrophic relationships renders the interaction a paramount example of a pathogenic plant-microbe biotrophy. However, molecular details underlying the interaction are in many respects still a mystery. Since its introduction in 1990, the Arabidopsis-powdery mildew pathosystem has become a popular model to study molecular processes governing powdery mildew infection. Due to the many advantages that the host Arabidopsis offers in terms of molecular and genetic tools this pathosystem has great capacity to answer some of the questions of how biotrophic pathogens overcome plant defense and establish a persistent interaction that nourishes the invader while in parallel maintaining viability of the plant host. PMID:27489521

  11. Biotrophy at Its Best: Novel Findings and Unsolved Mysteries of the Arabidopsis-Powdery Mildew Pathosystem

    PubMed Central

    Kuhn, Hannah; Kwaaitaal, Mark; Kusch, Stefan; Acevedo-Garcia, Johanna; Wu, Hongpo; Panstruga, Ralph

    2016-01-01

    It is generally accepted in plant-microbe interactions research that disease is the exception rather than a common outcome of pathogen attack. However, in nature, plants with symptoms that signify colonization by obligate biotrophic powdery mildew fungi are omnipresent. The pervasiveness of the disease and the fact that many economically important plants are prone to infection by powdery mildew fungi drives research on this interaction. The competence of powdery mildew fungi to establish and maintain true biotrophic relationships renders the interaction a paramount example of a pathogenic plant-microbe biotrophy. However, molecular details underlying the interaction are in many respects still a mystery. Since its introduction in 1990, the Arabidopsis-powdery mildew pathosystem has become a popular model to study molecular processes governing powdery mildew infection. Due to the many advantages that the host Arabidopsis offers in terms of molecular and genetic tools this pathosystem has great capacity to answer some of the questions of how biotrophic pathogens overcome plant defense and establish a persistent interaction that nourishes the invader while in parallel maintaining viability of the plant host. PMID:27489521

  12. Histone deacetylase 6–mediated selective autophagy regulates COPD-associated cilia dysfunction

    PubMed Central

    Lam, Hilaire C.; Cloonan, Suzanne M.; Bhashyam, Abhiram R.; Haspel, Jeffery A.; Singh, Anju; Sathirapongsasuti, J. Fah; Cervo, Morgan; Yao, Hongwei; Chung, Anna L.; Mizumura, Kenji; An, Chang Hyeok; Shan, Bin; Franks, Jonathan M.; Haley, Kathleen J.; Owen, Caroline A.; Tesfaigzi, Yohannes; Washko, George R.; Quackenbush, John; Silverman, Edwin K.; Rahman, Irfan; Kim, Hong Pyo; Mahmood, Ashfaq; Biswal, Shyam S.; Ryter, Stefan W.; Choi, Augustine M.K.

    2013-01-01

    Chronic obstructive pulmonary disease (COPD) involves aberrant airway inflammatory responses to cigarette smoke (CS) that are associated with epithelial cell dysfunction, cilia shortening, and mucociliary clearance disruption. Exposure to CS reduced cilia length and induced autophagy in vivo and in differentiated mouse tracheal epithelial cells (MTECs). Autophagy-impaired (Becn1+/– or Map1lc3B–/–) mice and MTECs resisted CS-induced cilia shortening. Furthermore, CS increased the autophagic turnover of ciliary proteins, indicating that autophagy may regulate cilia homeostasis. We identified cytosolic deacetylase HDAC6 as a critical regulator of autophagy-mediated cilia shortening during CS exposure. Mice bearing an X chromosome deletion of Hdac6 (Hdac6–/Y) and MTECs from these mice had reduced autophagy and were protected from CS-induced cilia shortening. Autophagy-impaired Becn1–/–, Map1lc3B–/–, and Hdac6–/Y mice or mice injected with an HDAC6 inhibitor were protected from CS-induced mucociliary clearance (MCC) disruption. MCC was preserved in mice given the chemical chaperone 4-phenylbutyric acid, but was disrupted in mice lacking the transcription factor NRF2, suggesting that oxidative stress and altered proteostasis contribute to the disruption of MCC. Analysis of human COPD specimens revealed epigenetic deregulation of HDAC6 by hypomethylation and increased protein expression in the airways. We conclude that an autophagy-dependent pathway regulates cilia length during CS exposure and has potential as a therapeutic target for COPD. PMID:24200693

  13. Insulin and mTOR Pathway Regulate HDAC3-Mediated Deacetylation and Activation of PGK1

    PubMed Central

    Wang, Shiwen; Jiang, Bowen; Zhang, Tengfei; Liu, Lixia; Wang, Yi; Wang, Yiping; Chen, Xiufei; Lin, Huaipeng; Zhou, Lisha; Xia, Yukun; Chen, Leilei; Yang, Chen; Xiong, Yue; Ye, Dan; Guan, Kun-Liang

    2015-01-01

    Phosphoglycerate kinase 1 (PGK1) catalyzes the reversible transfer of a phosphoryl group from 1, 3-bisphosphoglycerate (1, 3-BPG) to ADP, producing 3-phosphoglycerate (3-PG) and ATP. PGK1 plays a key role in coordinating glycolytic energy production with one-carbon metabolism, serine biosynthesis, and cellular redox regulation. Here, we report that PGK1 is acetylated at lysine 220 (K220), which inhibits PGK1 activity by disrupting the binding with its substrate, ADP. We have identified KAT9 and HDAC3 as the potential acetyltransferase and deacetylase, respectively, for PGK1. Insulin promotes K220 deacetylation to stimulate PGK1 activity. We show that the PI3K/AKT/mTOR pathway regulates HDAC3 S424 phosphorylation, which promotes HDAC3-PGK1 interaction and PGK1 K220 deacetylation. Our study uncovers a previously unknown mechanism for the insulin and mTOR pathway in regulation of glycolytic ATP production and cellular redox potential via HDAC3-mediated PGK1 deacetylation. PMID:26356530

  14. Interplay between arginine methylation and ubiquitylation regulates KLF4-mediated genome stability and carcinogenesis

    PubMed Central

    Hu, Dong; Gur, Mert; Zhou, Zhuan; Gamper, Armin; Hung, Mien-Chie; Fujita, Naoya; Lan, Li; Bahar, Ivet; Wan, Yong

    2015-01-01

    KLF4 is an important regulator of cell-fate decision, including DNA damage response and apoptosis. We identify a novel interplay between protein modifications in regulating KLF4 function. Here we show that arginine methylation of KLF4 by PRMT5 inhibits KLF4 ubiquitylation by VHL and thereby reduces KLF4 turnover, resulting in the elevation of KLF4 protein levels concomitant with increased transcription of KLF4-dependent p21 and reduced expression of KLF4-repressed Bax. Structure-based modelling and simulations provide insight into the molecular mechanisms of KLF4 recognition and catalysis by PRMT5. Following genotoxic stress, disruption of PRMT5-mediated KLF4 methylation leads to abrogation of KLF4 accumulation, which, in turn, attenuates cell cycle arrest. Mutating KLF4 methylation sites suppresses breast tumour initiation and progression, and immunohistochemical stain shows increased levels of both KLF4 and PRMT5 in breast cancer tissues. Taken together, our results point to a critical role for aberrant KLF4 regulation by PRMT5 in genome stability and breast carcinogenesis. PMID:26420673

  15. Transcriptional regulation mechanism mediated by miRNA-DNA•DNA triplex structure stabilized by Argonaute.

    PubMed

    Toscano-Garibay, Julia D; Aquino-Jarquin, Guillermo

    2014-11-01

    Transcription regulation depends on interactions between repressor or activator proteins with promoter sequences, while post-transcriptional regulation typically relies on microRNA (miRNA) interaction with sequences in 5' and 3'-Untranslated regions (UTRs) of messenger RNA (mRNA). However, several pieces of evidence suggest that miRNA:Argonaute (AGO) complexes may also suppress transcription through RNA interference (RNAi) components and epigenetic mechanisms. However, recent observations suggest that miRNA-induced transcriptional silencing could be exerted by an unknown mechanism independent of chromatin modifiers. The RNA-DNA•DNA triplex structure has emerged as an important RNA tertiary motif in which successive non-canonical base pairs form between a DNA-DNA duplex and a third strand. Frequently, promoters have Purine (PU)-rich tracts, and some Triplex-forming oligonucleotides (TFOs) targeting these regulatory regions have been shown to inhibit transcription selectively. Here, we summarize observations suggesting that miRNAs exert regulation over promoter regions through miRNA-DNA•DNA triplex structure formation stabilized by AGO proteins which represents a plausible model of RNA-mediated Transcriptional gene silencing (TGS). PMID:25086339

  16. MicroRNA-mediated networks underlie immune response regulation in papillary thyroid carcinoma

    NASA Astrophysics Data System (ADS)

    Huang, Chen-Tsung; Oyang, Yen-Jen; Huang, Hsuan-Cheng; Juan, Hsueh-Fen

    2014-09-01

    Papillary thyroid carcinoma (PTC) is a common endocrine malignancy with low death rate but increased incidence and recurrence in recent years. MicroRNAs (miRNAs) are small non-coding RNAs with diverse regulatory capacities in eukaryotes and have been frequently implied in human cancer. Despite current progress, however, a panoramic overview concerning miRNA regulatory networks in PTC is still lacking. Here, we analyzed the expression datasets of PTC from The Cancer Genome Atlas (TCGA) Data Portal and demonstrate for the first time that immune responses are significantly enriched and under specific regulation in the direct miRNA-target network among distinctive PTC variants to different extents. Additionally, considering the unconventional properties of miRNAs, we explore the protein-coding competing endogenous RNA (ceRNA) and the modulatory networks in PTC and unexpectedly disclose concerted regulation of immune responses from these networks. Interestingly, miRNAs from these conventional and unconventional networks share general similarities and differences but tend to be disparate as regulatory activities increase, coordinately tuning the immune responses that in part account for PTC tumor biology. Together, our systematic results uncover the intensive regulation of immune responses underlain by miRNA-mediated networks in PTC, opening up new avenues in the management of thyroid cancer.

  17. MicroRNA29a regulates IL-33-mediated tissue remodelling in tendon disease

    PubMed Central

    Millar, Neal L.; Gilchrist, Derek S.; Akbar, Moeed; Reilly, James H.; Kerr, Shauna C.; Campbell, Abigail L.; Murrell, George A. C.; Liew, Foo Y.; Kurowska-Stolarska, Mariola; McInnes, Iain B.

    2015-01-01

    MicroRNA (miRNA) has the potential for cross-regulation and functional integration of discrete biological processes during complex physiological events. Utilizing the common human condition tendinopathy as a model system to explore the cross-regulation of immediate inflammation and matrix synthesis by miRNA we observed that elevated IL-33 expression is a characteristic of early tendinopathy. Using in vitro tenocyte cultures and in vivo models of tendon damage, we demonstrate that such IL-33 expression plays a pivotal role in the transition from type 1 to type 3 collagen (Col3) synthesis and thus early tendon remodelling. Both IL-33 effector function, via its decoy receptor sST2, and Col3 synthesis are regulated by miRNA29a. Downregulation of miRNA29a in human tenocytes is sufficient to induce an increase in Col3 expression. These data provide a molecular mechanism of miRNA-mediated integration of the early pathophysiologic events that facilitate tissue remodelling in human tendon after injury. PMID:25857925

  18. Interplay between arginine methylation and ubiquitylation regulates KLF4-mediated genome stability and carcinogenesis.

    PubMed

    Hu, Dong; Gur, Mert; Zhou, Zhuan; Gamper, Armin; Hung, Mien-Chie; Fujita, Naoya; Lan, Li; Bahar, Ivet; Wan, Yong

    2015-01-01

    KLF4 is an important regulator of cell-fate decision, including DNA damage response and apoptosis. We identify a novel interplay between protein modifications in regulating KLF4 function. Here we show that arginine methylation of KLF4 by PRMT5 inhibits KLF4 ubiquitylation by VHL and thereby reduces KLF4 turnover, resulting in the elevation of KLF4 protein levels concomitant with increased transcription of KLF4-dependent p21 and reduced expression of KLF4-repressed Bax. Structure-based modelling and simulations provide insight into the molecular mechanisms of KLF4 recognition and catalysis by PRMT5. Following genotoxic stress, disruption of PRMT5-mediated KLF4 methylation leads to abrogation of KLF4 accumulation, which, in turn, attenuates cell cycle arrest. Mutating KLF4 methylation sites suppresses breast tumour initiation and progression, and immunohistochemical stain shows increased levels of both KLF4 and PRMT5 in breast cancer tissues. Taken together, our results point to a critical role for aberrant KLF4 regulation by PRMT5 in genome stability and breast carcinogenesis. PMID:26420673

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

    SciTech Connect

    Sakamaki, Jun-ichi; Daitoku, Hiroaki; Yoshimochi, Kenji; Miwa, Masanao; Fukamizu, Akiyoshi

    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 dependent 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.

  20. Complement regulates TLR4-mediated inflammatory responses during intestinal ischemia reperfusion

    PubMed Central

    Pope, Michael R.; Hoffman, Sara M.; Tomlinson, Stephen; Fleming, Sherry D.

    2010-01-01

    Innate immune responses including TLR4 and complement activation are required for mesenteric ischemia/reperfusion (IR)-induced tissue damage. We examined the regulation of TLR4 and complement activation in a mouse model of intestinal IR. Intestinal IR induced C3 deposition in a TLR4 dependent manner. In addition, in wild-type but not TLR4 deficient mice, IR significantly increased C3 and Factor B (FB) mRNA expression within the intestine. To further examine the role of TLR4 and complement, we administered the complement inhibitor, CR2-Crry, to target local complement activation in wild-type C57Bl/10, and TLR4 deficient B10/ScN mice. TLR4 deficient mice sustained less damage and inflammation after IR than wild-type mice, but administration of CR2-Crry did not further reduce tissue damage. In contrast, CR2-Crry treatment of wild-type mice was accompanied by a reduction in complement activation and in C3 and FB transcription in response to IR. CR2-Crry also significantly decreased intestinal IL-6 and IL-12p40 production in both the wild-type and TLR4 deficient mice. These data indicate that TLR4 regulates extrahepatic complement production while complement regulates TLR4-mediated cytokine production during intestinal IR. PMID:20800895

  1. WDR82 Negatively Regulates Cellular Antiviral Response by Mediating TRAF3 Polyubiquitination in Multiple Cell Lines.

    PubMed

    Zhu, Kun; Wang, Xiang; Ju, Lin-Gao; Zhu, Yuan; Yao, Jie; Wang, Yanyi; Wu, Min; Li, Lian-Yun

    2015-12-01

    Upon virus infection, retinoic acid-inducible gene I-like receptors in host cells recognize viral RNA and activate type I IFN expression. Previously, we identified WD repeat domain (WDR) 5 as one positive regulator for pathway activation. In this study, we report that WDR82, a homolog protein of WDR5, acts opposite to WDR5 and inhibits the activation of the retinoic acid-inducible gene I signaling pathway. WDR82 overexpression inhibits virus-triggered pathway activation, whereas its knockdown enhances induced IFN-β expression. WDR82 is localized on the mitochondria, and its first N-terminal WD40 domain is critical for localization. WDR82 interacts with TNFR-associated factor (TRAF) 3, and its overexpression promotes K48-linked, but not K63-linked, polyubiquitination on TRAF3. Furthermore, WDR82 knockdown inhibits viral replication in the cell, whereas its overexpression has the opposite effect. Interestingly, WDR82 regulates Sendai virus-induced IFNB1 expression in a cell type-specific manner. Taken together, our findings demonstrate that WDR82 is a negative regulator of virus-triggered type I IFNs pathway through mediating TRAF3 polyubiquitination status and stability on mitochondria. PMID:26519536

  2. [Regulation of Positive and Negative Emotions as Mediator between Maternal Emotion Socialization and Child Problem Behavior].

    PubMed

    Fäsche, Anika; Gunzenhauser, Catherine; Friedlmeier, Wolfgang; von Suchodoletz, Antje

    2015-01-01

    The present study investigated five to six year old children's ability to regulate negative and positive emotions in relation to psychosocial problem behavior (N=53). It was explored, whether mothers' supportive and nonsupportive strategies of emotion socialization influence children's problem behavior by shaping their emotion regulation ability. Mothers reported on children's emotion regulation and internalizing and externalizing problem behavior via questionnaire, and were interviewed about their preferences for socialization strategies in response to children's expression of negative affect. Results showed that children with more adaptive expression of adequate positive emotions had less internalizing behavior problems. When children showed more control of inadequate negative emotions, children were less internalizing as well as externalizing in their behavior. Furthermore, results indicated indirect relations of mothers' socialization strategies with children's problem behavior. Control of inadequate negative emotions mediated the link between non-supportive strategies on externalizing problem behavior. Results suggest that emotion regulatory processes should be part of interventions to reduce the development of problematic behavior in young children. Parents should be trained in dealing with children's emotions in a constructive way. PMID:26032031

  3. MAR-mediated integration of plasmid vectors for in vivo gene transfer and regulation

    PubMed Central

    2013-01-01

    Background The in vivo transfer of naked plasmid DNA into organs such as muscles is commonly used to assess the expression of prophylactic or therapeutic genes in animal disease models. Results In this study, we devised vectors allowing a tight regulation of transgene expression in mice from such non-viral vectors using a doxycycline-controlled network of activator and repressor proteins. Using these vectors, we demonstrate proper physiological response as consequence of the induced expression of two therapeutically relevant proteins, namely erythropoietin and utrophin. Kinetic studies showed that the induction of transgene expression was only transient, unless epigenetic regulatory elements termed Matrix Attachment Regions, or MAR, were inserted upstream of the regulated promoters. Using episomal plasmid rescue and quantitative PCR assays, we observed that similar amounts of plasmids remained in muscles after electrotransfer with or without MAR elements, but that a significant portion had integrated into the muscle fiber chromosomes. Interestingly, the MAR elements were found to promote plasmid genomic integration but to oppose silencing effects in vivo, thereby mediating long-term expression. Conclusions This study thus elucidates some of the determinants of transient or sustained expression from the use of non-viral regulated vectors in vivo. PMID:24295286

  4. MicroRNA29a regulates IL-33-mediated tissue remodelling in tendon disease.

    PubMed

    Millar, Neal L; Gilchrist, Derek S; Akbar, Moeed; Reilly, James H; Kerr, Shauna C; Campbell, Abigail L; Murrell, George A C; Liew, Foo Y; Kurowska-Stolarska, Mariola; McInnes, Iain B

    2015-01-01

    MicroRNA (miRNA) has the potential for cross-regulation and functional integration of discrete biological processes during complex physiological events. Utilizing the common human condition tendinopathy as a model system to explore the cross-regulation of immediate inflammation and matrix synthesis by miRNA we observed that elevated IL-33 expression is a characteristic of early tendinopathy. Using in vitro tenocyte cultures and in vivo models of tendon damage, we demonstrate that such IL-33 expression plays a pivotal role in the transition from type 1 to type 3 collagen (Col3) synthesis and thus early tendon remodelling. Both IL-33 effector function, via its decoy receptor sST2, and Col3 synthesis are regulated by miRNA29a. Downregulation of miRNA29a in human tenocytes is sufficient to induce an increase in Col3 expression. These data provide a molecular mechanism of miRNA-mediated integration of the early pathophysiologic events that facilitate tissue remodelling in human tendon after injury. PMID:25857925

  5. Yeast Actin-Related Protein ARP6 Negatively Regulates Agrobacterium-Mediated Transformation of Yeast Cell.

    PubMed

    Luo, Yumei; Chen, Zikai; Zhu, Detu; Tu, Haitao; Pan, Shen Quan

    2015-01-01

    The yeasts, including Saccharomyces cerevisiae and Pichia pastoris, are single-cell eukaryotic organisms that can serve as models for human genetic diseases and hosts for large scale production of recombinant proteins in current biopharmaceutical industry. Thus, efficient genetic engineering tools for yeasts are of great research and economic values. Agrobacterium tumefaciens-mediated transformation (AMT) can transfer T-DNA into yeast cells as a method for genetic engineering. However, how the T-DNA is transferred into the yeast cells is not well established yet. Here our genetic screening of yeast knockout mutants identified a yeast actin-related protein ARP6 as a negative regulator of AMT. ARP6 is a critical member of the SWR1 chromatin remodeling complex (SWR-C); knocking out some other components of the complex also increased the transformation efficiency, suggesting that ARP6 might regulate AMT via SWR-C. Moreover, knockout of ARP6 led to disruption of microtubule integrity, higher uptake and degradation of virulence proteins, and increased DNA stability inside the cells, all of which resulted in enhanced transformation efficiency. Our findings have identified molecular and cellular mechanisms regulating AMT and a potential target for enhancing the transformation efficiency in yeast cells. PMID:26425545

  6. DNA methylation-mediated transcription factors regulate Piwil1 expression during chicken spermatogenesis

    PubMed Central

    QIU, Lingling; XU, Lu; CHANG, Guobin; GUO, Qixin; LIU, Xiangping; BI, Yulin; ZHANG, Yu; WANG, Hongzhi; WANG, Kehua; LU, Wei; REN, Lichen; ZHU, Pengfei; WU, Yun; ZHANG, Yang; XU, Qi; CHEN, Guohong

    2016-01-01

    The P-element induced wimpy testis (Piwi) protein family is responsible for initiating spermatogenesis and maintaining the integrity of germ cells and stem cells, but little is known regarding its transcriptional regulation in poultry. Here, we characterized the methylation status of the Piwil1 promoter in five different spermatogenic cell lines using direct bisulfite pyrosequencing and determined that methylation correlates negatively with germ cell type-specific expression patterns of piwil1. We demonstrated that methylation of the −148 CpG site, which is the predicted binding site for the transcription factors TCF3 and NRF1, was differentially methylated in different spermatogenic cells. This site was completely methylated in PGCs (primordial germ cells), but was unmethylated in round spermatids. A similar result was obtained in the region from +121 to +139 CpG sites of the Piwil1 promoter CpG island, which was predicted to contain SOX2 binding sites. In addition, demethylation assays further demonstrated that DNA methylation indeed regulates Piwil1 expression during chicken spermatogenesis. Combined with transcription factor binding site prediction, we speculate that methylation influences the recruitment of corresponding transcription factors. Collectively, we show the negative correlation between promoter methylation and piwil1 expression and that the spatiotemporal expression of chicken Piwil1 from the PGC stage to the round spermatid stage is influenced by methylation-mediated transcription factor regulation. PMID:27108736

  7. Maternal Attachment Style and Responses to Adolescents’ Negative Emotions: The Mediating Role of Maternal Emotion Regulation

    PubMed Central

    Jones, Jason D.; Brett, Bonnie E.; Ehrlich, Katherine B.; Lejuez, Carl W.; Cassidy, Jude

    2014-01-01

    SYNOPSIS Objective Previous research has examined the developmental consequences, particularly in early childhood, of parents’ supportive and unsupportive responses to children’s negative emotions. Much less is known about factors that explain why parents respond in ways that may support or undermine their children’s emotions, and even less is known about how these parenting processes unfold with adolescents. We examined the associations between mothers’ attachment styles and their distress, harsh, and supportive responses to their adolescents’ negative emotions two years later and whether these links were mediated by maternal emotion regulation difficulties. Design Mothers in a longitudinal study (n = 230) reported on their attachment style, difficulties regulating their emotions, and their hypothetical responses to their adolescents’ negative emotions, respectively, at consecutive laboratory visits one year apart. Results Mothers who reported greater attachment-related avoidance and anxiety reported having greater difficulties with emotion regulation one year later. Emotion dysregulation, in turn, predicted more distressed, harsher, and less supportive maternal responses to adolescents’ negative emotions the following year. In addition, greater avoidance directly predicted harsher maternal responses two years later. Conclusions These findings extend previous research by identifying maternal attachment style as a predictor of responses to adolescent distress and by documenting the underlying role of emotion dysregulation in the link between adult attachment style and parenting. PMID:25568638

  8. Yeast Actin-Related Protein ARP6 Negatively Regulates Agrobacterium-Mediated Transformation of Yeast Cell

    PubMed Central

    Luo, Yumei; Chen, Zikai; Zhu, Detu; Tu, Haitao; Pan, Shen Quan

    2015-01-01

    The yeasts, including Saccharomyces cerevisiae and Pichia pastoris, are single-cell eukaryotic organisms that can serve as models for human genetic diseases and hosts for large scale production of recombinant proteins in current biopharmaceutical industry. Thus, efficient genetic engineering tools for yeasts are of great research and economic values. Agrobacterium tumefaciens-mediated transformation (AMT) can transfer T-DNA into yeast cells as a method for genetic engineering. However, how the T-DNA is transferred into the yeast cells is not well established yet. Here our genetic screening of yeast knockout mutants identified a yeast actin-related protein ARP6 as a negative regulator of AMT. ARP6 is a critical member of the SWR1 chromatin remodeling complex (SWR-C); knocking out some other components of the complex also increased the transformation efficiency, suggesting that ARP6 might regulate AMT via SWR-C. Moreover, knockout of ARP6 led to disruption of microtubule integrity, higher uptake and degradation of virulence proteins, and increased DNA stability inside the cells, all of which resulted in enhanced transformation efficiency. Our findings have identified molecular and cellular mechanisms regulating AMT and a potential target for enhancing the transformation efficiency in yeast cells. PMID:26425545

  9. Src-mediated Post-translational Regulation of Endoglin Stability and Function Is Critical for Angiogenesis*

    PubMed Central

    Pan, Christopher C.; Kumar, Sanjay; Shah, Nirav; Hoyt, Dale G.; Hawinkels, Lukas J. A. C.; Mythreye, Karthikeyan; Lee, Nam Y.

    2014-01-01

    Endoglin is a transforming growth factor β (TGF-β) co-receptor essential for angiogenesis and tumor vascularization. Endoglin modulates the crucial balance between pro- and anti-angiogenic signaling by activin receptor-like kinase (ALK) 1, 5, and TGF-β type II (TβRII) receptors. Despite its established role in physiology and disease, the mechanism of endoglin down-regulation remains unknown. Here we report that the conserved juxtamembrane cytoplasmic tyrosine motif (612YIY614) is a critical determinant of angiogenesis. Src directly phosphorylates this motif to induce endoglin internalization and degradation via the lysosome. We identified epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF) as Src-activators that induce endoglin turnover following 612YIY614 phosphorylation. Interestingly, Src phosphorylation of endoglin-612YIY614 was also an important process for receptor down-regulation by TRACON105 (TRC105), an endoglin-targeting antibody currently in clinical trials. The regulation of 612YIY614 phosphorylation was critical for angiogenesis, as both the phosphomimetic and unphosphorylatable mutants impaired endothelial functions including proliferation, migration, and capillary tube formation. Collectively, these findings establish Src and pro-angiogenic mitogens as critical mediators of endoglin stability and function. PMID:25070888

  10. Effects of Myoga on Memory and Synaptic Plasticity by Regulating Nerve Growth Factor-Mediated Signaling.

    PubMed

    Kim, Hyo Geun; Lim, Soonmin; Hong, Jongki; Kim, Ae-Jung; Oh, Myung Sook

    2016-02-01

    The flower bud of Zingiber mioga Roscoe, known as 'myoga' or Japanese ginger, has a pungent aroma and is commonly consumed as a spice, with pickles, or as a health supplement in Eastern Asia. Here, we evaluated the activity of myoga in the brain, focusing especially on nerve growth factor (NGF), which is believed to mediate synaptic plasticity, supporting learning and memory. In a rat primary hippocampal astrocyte culture system, treatment with myoga extract for 24 h significantly stimulated the production of NGF. In mice administered myoga extract for 14 days, 200 and 400 mg/kg/day treatment resulted in increased NGF levels in the hippocampus. Myoga extract treatment also regulated the phosphorylation of extracellular signal-regulated kinases and cAMP response element-binding protein in the mouse hippocampus, leading to increased synaptic plasticity. In addition, it significantly increased novel object recognition time and spontaneous alternation, indicating improvement in learning and memory. These results suggest that myoga helps regulate NGF and synaptic plasticity, increasing memory ability. PMID:26563629

  11. Membrane-Mediated Regulation of the Intrinsically Disordered CD3ϵ Cytoplasmic Tail of the TCR

    PubMed Central

    López, Cesar A.; Sethi, Anurag; Goldstein, Byron; Wilson, Bridget S.; Gnanakaran, S.

    2015-01-01

    The regulation of T-cell-mediated immune responses depends on the phosphorylation of immunoreceptor tyrosine-based activation motifs (ITAMs) on T-cell receptors. Although many details of the signaling cascades are well understood, the initial mechanism and regulation of ITAM phosphorylation remains unknown. We used molecular dynamics simulations to study the influence of different compositions of lipid bilayers on the membrane association of the CD3ϵ cytoplasmic tails of the T-cell receptors. Our results show that binding of CD3ϵ to membranes is modulated by both the presence of negatively charged lipids and the lipid order of the membrane. Free-energy calculations reveal that the protein-membrane interaction is favored by the presence of nearby basic residues and the ITAM tyrosines. Phosphorylation minimizes membrane association, rendering the ITAM motif more accessible to binding partners. In systems mimicking biological membranes, the CD3ϵ chain localization is modulated by different facilitator lipids (e.g., gangliosides or phosphoinositols), revealing a plausible regulatory effect on activation through the regulation of lipid composition in cell membranes. PMID:25992726

  12. Impulsivity, self-regulation,and pathological video gaming among youth: testing a mediation model.

    PubMed

    Liau, Albert K; Neo, Eng Chuan; Gentile, Douglas A; Choo, Hyekyung; Sim, Timothy; Li, Dongdong; Khoo, Angeline

    2015-03-01

    Given the potential negative mental health consequences of pathological video gaming, understanding its etiology may lead to useful treatment developments. The purpose of the study was to examine the influence of impulsive and regulatory processes on pathological video gaming. Study 1 involved 2154 students from 6 primary and 4 secondary schools in Singapore. Study 2 involved 191 students from 2 secondary schools. The results of study 1 and study 2 supported the hypothesis that self-regulation is a mediator between impulsivity and pathological video gaming. Specifically, higher levels of impulsivity was related to lower levels of self-regulation, which in turn was related to higher levels of pathological video gaming. The use of impulsivity and self-regulation in predicting pathological video gaming supports the dual-system model of incorporating both impulsive and reflective systems in the prediction of self-control outcomes. The study highlights the development of self-regulatory resources as a possible avenue for future prevention and treatment research. PMID:22199156

  13. Lipid droplet-mediated ER homeostasis regulates autophagy and cell survival during starvation.

    PubMed

    Velázquez, Ariadna P; Tatsuta, Takashi; Ghillebert, Ruben; Drescher, Ingmar; Graef, Martin

    2016-03-14

    Lipid droplets (LDs) are conserved organelles for intracellular neutral lipid storage. Recent studies suggest that LDs function as direct lipid sources for autophagy, a central catabolic process in homeostasis and stress response. Here, we demonstrate that LDs are dispensable as a membrane source for autophagy, but fulfill critical functions for endoplasmic reticulum (ER) homeostasis linked to autophagy regulation. In the absence of LDs, yeast cells display alterations in their phospholipid composition and fail to buffer de novo fatty acid (FA) synthesis causing chronic stress and morphologic changes in the ER. These defects compromise regulation of autophagy, including formation of multiple aberrant Atg8 puncta and drastically impaired autophagosome biogenesis, leading to severe defects in nutrient stress survival. Importantly, metabolically corrected phospholipid composition and improved FA resistance of LD-deficient cells cure autophagy and cell survival. Together, our findings provide novel insight into the complex interrelation between LD-mediated lipid homeostasis and the regulation of autophagy potentially relevant for neurodegenerative and metabolic diseases. PMID:26953354

  14. DNA methylation-mediated transcription factors regulate Piwil1 expression during chicken spermatogenesis.

    PubMed

    Qiu, Lingling; Xu, Lu; Chang, Guobin; Guo, Qixin; Liu, Xiangping; Bi, Yulin; Zhang, Yu; Wang, Hongzhi; Wang, Kehua; Lu, Wei; Ren, Lichen; Zhu, Pengfei; Wu, Yun; Zhang, Yang; Xu, Qi; Chen, Guohong

    2016-08-25

    The P-element induced wimpy testis (Piwi) protein family is responsible for initiating spermatogenesis and maintaining the integrity of germ cells and stem cells, but little is known regarding its transcriptional regulation in poultry. Here, we characterized the methylation status of the Piwil1 promoter in five different spermatogenic cell lines using direct bisulfite pyrosequencing and determined that methylation correlates negatively with germ cell type-specific expression patterns of piwil1. We demonstrated that methylation of the -148 CpG site, which is the predicted binding site for the transcription factors TCF3 and NRF1, was differentially methylated in different spermatogenic cells. This site was completely methylated in PGCs (primordial germ cells), but was unmethylated in round spermatids. A similar result was obtained in the region from +121 to +139 CpG sites of the Piwil1 promoter CpG island, which was predicted to contain SOX2 binding sites. In addition, demethylation assays further demonstrated that DNA methylation indeed regulates Piwil1 expression during chicken spermatogenesis. Combined with transcription factor binding site prediction, we speculate that methylation influences the recruitment of corresponding transcription factors. Collectively, we show the negative correlation between promoter methylation and piwil1 expression and that the spatiotemporal expression of chicken Piwil1 from the PGC stage to the round spermatid stage is influenced by methylation-mediated transcription factor regulation. PMID:27108736

  15. Chk2 regulates transcription-independent p53-mediated apoptosis in response to DNA damage

    SciTech Connect

    Chen Chen; Shimizu, Shigeomi; Tsujimoto, Yoshihide; Motoyama, Noboru . E-mail: motoyama@nils.go.jp

    2005-07-29

    The tumor suppressor protein p53 plays a central role in the induction of apoptosis in response to genotoxic stress. The protein kinase Chk2 is an important regulator of p53 function in mammalian cells exposed to ionizing radiation (IR). Cells derived from Chk2-deficient mice are resistant to the induction of apoptosis by IR, and this resistance has been thought to be a result of the defective transcriptional activation of p53 target genes. It was recently shown, however, that p53 itself and histone H1.2 translocate to mitochondria and thereby induces apoptosis in a transcription-independent manner in response to IR. We have now examined whether Chk2 also regulates the transcription-independent induction of apoptosis by p53 and histone H1.2. The reduced ability of IR to induce p53 stabilization in Chk2-deficient thymocytes was associated with a marked impairment of p53 and histone H1 translocation to mitochondria. These results suggest that Chk2 regulates the transcription-independent mechanism of p53-mediated apoptosis by inducing stabilization of p53 in response to IR.

  16. A New Oxidative Sensing and Regulation Pathway Mediated by the MgrA Homologue SarZ in Staphylococcus aureus

    PubMed Central

    Chen, Peng, R.; Nishida, Satoshi; Poor, Catherine B.; Cheng, Alice; Bae, Taeok; Kuechenmeister, Lisa; Dunman, Paul M.; Missiakas, Dominique; He, Chuan

    2009-01-01

    Summary Oxidative stress serves as an important host/environmental signal that triggers a wide range of responses from the human pathogen Staphylococcus aureus. Among these, a thiol-based oxidation sensing pathway through a global regulator MgrA controls the virulence and antibiotic resistance of the bacterium. Herein, we report a new thiol-based oxidation sensing and regulation system that is mediated through a parallel global regulator SarZ. SarZ is a functional homologue of MgrA and is shown to affect the expression of ~87 genes in S. aureus. It uses a key Cys residue, Cys13, to sense oxidative stress and to coordinate the expression of genes involved in metabolic switching, antibiotic resistance, peroxide stress defense, virulence, and cell wall properties. The discovery of this SarZ-mediated regulation, mostly independent from the MgrA-based regulation, fills a missing gap of oxidation sensing and response in S. aureus. PMID:19007410

  17. The submergence tolerance regulator Sub1A mediates stress-responsive expression of AP2/ERF transcription factors.

    PubMed

    Jung, Ki-Hong; Seo, Young-Su; Walia, Harkamal; Cao, Peijian; Fukao, Takeshi; Canlas, Patrick E; Amonpant, Fawn; Bailey-Serres, Julia; Ronald, Pamela C

    2010-03-01

    We previously characterized the rice (Oryza sativa) Submergence1 (Sub1) locus encoding three ethylene-responsive factor (ERF) transcriptional regulators. Genotypes carrying the Sub1A-1 allele are tolerant of prolonged submergence. To elucidate the mechanism of Sub1A-1-mediated tolerance, we performed transcriptome analyses comparing the temporal submergence response of Sub1A-1-containing tolerant M202(Sub1) with the intolerant isoline M202 lacking this gene. We identified 898 genes displaying Sub1A-1-dependent regulation. Integration of the expression data with publicly available metabolic pathway data identified submergence tolerance-associated pathways governing anaerobic respiration, hormone responses, and antioxidant systems. Of particular interest were a set of APETALA2 (AP2)/ERF family transcriptional regulators that are associated with the Sub1A-1-mediated response upon submergence. Visualization of expression patterns of the AP2/ERF superfamily members in a phylogenetic context resolved 12 submergence-regulated AP2/ERFs into three putative functional groups: (1) anaerobic respiration and cytokinin-mediated delay in senescence via ethylene accumulation during submergence (three ERFs); (2) negative regulation of ethylene-dependent gene expression (five ERFs); and (3) negative regulation of gibberellin-mediated shoot elongation (four ERFs). These results confirm that the presence of Sub1A-1 impacts multiple pathways of response to submergence. PMID:20107022

  18. PPAR-γ activation by Tityus serrulatus venom regulates lipid body formation and lipid mediator production.

    PubMed

    Zoccal, Karina Furlani; Paula-Silva, Francisco Wanderley Garcia; Bitencourt, Claudia da Silva; Sorgi, Carlos Artério; Bordon, Karla de Castro Figueiredo; Arantes, Eliane Candiani; Faccioli, Lúcia Helena

    2015-01-01

    Tityus serrulatus venom (TsV) consists of numerous peptides with different physiological and pharmacological activities. Studies have shown that scorpion venom increases pro-inflammatory cytokine production, contributing to immunological imbalance, multiple organ dysfunction, and patient death. We have previously demonstrated that TsV is a venom-associated molecular pattern (VAMP) recognized by TLRs inducing intense inflammatory reaction through the production of pro-inflammatory cytokines and arachidonic acid-derived lipid mediators prostaglandin (PG)E2 and leukotriene (LT)B4. Lipid bodies (LBs) are potential sites for eicosanoid production by inflammatory cells. Moreover, recent studies have shown that the peroxisome proliferator-activated receptor gamma (PPAR-γ) is implicated in LB formation and acts as an important modulator of lipid metabolism during inflammation. In this study, we used murine macrophages to evaluate whether the LB formation induced by TsV after TLR recognition correlates with lipid mediator generation by macrophages and if it occurs through PPAR-γ activation. We demonstrate that TsV acts through TLR2 and TLR4 stimulation and PPAR-γ activation to induce LB formation and generation of PGE2 and LTB4. Our data also show that PPAR-γ negatively regulates the pro-inflammatory NF-κB transcription factor. Based on these results, we suggest that during envenomation, LBs constitute functional organelles for lipid mediator production through signaling pathways that depend on cell surface and nuclear receptors. These findings point to the inflammatory mechanisms that might also be triggered during human envenomation by TsV. PMID:25450800

  19. Ceramidases: regulators of cellular responses mediated by ceramide, sphingosine, and sphingosine-1-phosphate

    PubMed Central

    Mao, Cungui; Obeid, Lina M.

    2008-01-01

    Summary Ceramidases catalyze hydrolysis of ceramides to generate sphingosine (SPH), which is phosphorylated to form sphingosine-1-phosphate (S1P). Ceramide, SPH, and S1P are bioactive lipids that mediate cell proliferation, differentiation, apoptosis, adhesion, and migration, likely by controlling hydrolysis of ceramides and generation of SPH and S1P. Presently, 5 human ceramidases encoded by 5 distinct genes have been cloned: acid ceramidase (AC), neutral ceramidase (NC), alkaline ceramidase 1 (ACER1), alkaline ceramidase 2 (ACER2), and alkaline ceramidase 3 (ACER3). Each human ceramidase has a mouse counterpart. AC, NC, and ACER1–3 have maximal activities in acidic, neutral, and alkaline environments, respectively. ACER1–3 have similar protein sequences but no homology to AC and NC. AC and NC also have distinct protein sequences. The human AC (hAC) was implicated in Farber disease, and hAC may be important for cell survival. The mouse AC (mAC) is needed for early embryo survival. NC is protective against inflammatory cytokines, and the mouse NC (mNC) is required for the catabolism of ceramides in the digestive tract. ACER1 is critical in mediating cell differentiation by controlling the generation of SPH and S1P and that ACER2’s role in cell proliferation and survival depends on its expression or the cell type in which it is found. Here, we discuss the role of each ceramidase in regulating cellular responses mediated by ceramides, SPH, and S1P. PMID:18619555

  20. IL-4 regulates specific Arg-1(+) macrophage sFlt-1-mediated inhibition of angiogenesis.

    PubMed

    Wu, Wei-Kang; Georgiadis, Anastasios; Copland, David A; Liyanage, Sidath; Luhmann, Ulrich F O; Robbie, Scott J; Liu, Jian; Wu, Jiahui; Bainbridge, James W; Bates, David O; Ali, Robin R; Nicholson, Lindsay B; Dick, Andrew D

    2015-08-01

    One of the main drivers for neovascularization in age-related macular degeneration is activation of innate immunity in the presence of macrophages. Here, we demonstrate that T helper cell type 2 cytokines and, in particular, IL-4 condition human and murine monocyte phenotype toward Arg-1(+), and their subsequent behavior limits angiogenesis by increasing soluble fms-like tyrosine kinase 1 (sFlt-1) gene expression. We document that T helper cell type 2 cytokine-conditioned murine macrophages neutralize vascular endothelial growth factor-mediated endothelial cell proliferation (human umbilical vein endothelial cell and choroidal vasculature) in a sFlt-1-dependent manner. We demonstrate that in vivo intravitreal administration of IL-4 attenuates laser-induced choroidal neovascularization (L-CNV) due to specific IL-4 conditioning of macrophages. IL-4 induces the expression of sFlt-1 by resident CD11b(+) retinal microglia and infiltrating myeloid cells but not from retinal pigment epithelium. IL-4-induced suppression of L-CNV is not prevented when sFlt-1 expression is attenuated in retinal pigment epithelium. IL-4-mediated suppression of L-CNV was abrogated in IL-4R-deficient mice and in bone marrow chimeras reconstituted with myeloid cells that had undergone lentiviral-mediated shRNA silencing of sFlt-1, demonstrating the critical role of this cell population. Together, these data establish how lL-4 directly drives macrophage sFlt-1 production expressing an Arg-1(+) phenotype and support the therapeutic potential of targeted IL-4 conditioning within the tissue to regulate disease conditions such as neovascular age-related macular degeneration. PMID:26079814

  1. Nipbl and Mediator Cooperatively Regulate Gene Expression to Control Limb Development

    PubMed Central

    Muto, Akihiko; Ikeda, Shingo; Lopez-Burks, Martha E.

    2014-01-01

    Haploinsufficiency for Nipbl, a cohesin loading protein, causes Cornelia de Lange Syndrome (CdLS), the most common “cohesinopathy”. It has been proposed that the effects of Nipbl-haploinsufficiency result from disruption of long-range communication between DNA elements. Here we use zebrafish and mouse models of CdLS to examine how transcriptional changes caused by Nipbl deficiency give rise to limb defects, a common condition in individuals with CdLS. In the zebrafish pectoral fin (forelimb), knockdown of Nipbl expression led to size reductions and patterning defects that were preceded by dysregulated expression of key early limb development genes, including fgfs, shha, hand2 and multiple hox genes. In limb buds of Nipbl-haploinsufficient mice, transcriptome analysis revealed many similar gene expression changes, as well as altered expression of additional classes of genes that play roles in limb development. In both species, the pattern of dysregulation of hox-gene expression depended on genomic location within the Hox clusters. In view of studies suggesting that Nipbl colocalizes with the mediator complex, which facilitates enhancer-promoter communication, we also examined zebrafish deficient for the Med12 Mediator subunit, and found they resembled Nipbl-deficient fish in both morphology and gene expression. Moreover, combined partial reduction of both Nipbl and Med12 had a strongly synergistic effect, consistent with both molecules acting in a common pathway. In addition, three-dimensional fluorescent in situ hybridization revealed that Nipbl and Med12 are required to bring regions containing long-range enhancers into close proximity with the zebrafish hoxda cluster. These data demonstrate a crucial role for Nipbl in limb development, and support the view that its actions on multiple gene pathways result from its influence, together with Mediator, on regulation of long-range chromosomal interactions. PMID:25255084

  2. Regulation of the LPA2 Receptor Signaling through the Carboxyl-Terminal Tail-Mediated Protein-Protein Interactions

    PubMed Central

    Lin, Fang-Tsyr; Lai, Yun-Ju

    2008-01-01

    While it is well known that lysophosphatidic acid (LPA) mediates diverse physiological and pathophysiological responses through the activation of G protein-coupled LPA receptors, the specificity and molecular mechanisms by which different LPA receptors mediate these biological responses remain largely unknown. Recent identification of several PDZ proteins and zinc finger proteins that interact with the carboxyl-terminal tail of the LPA2 receptor provides a considerable progress towards the understanding of the mechanisms how the LPA2 receptor specifically mediates LPA signaling pathways. These findings have led to the proposal that there are at least two distinct protein interaction motifs present in the carboxyl terminus of the LPA2 receptor. Together, these data provide a new concept that the efficiency and specificity of the LPA2 receptor-mediated signal transduction can be achieved through the cross-regulation between the classical G protein-activated signaling cascades and the interacting partner-mediated signaling pathways. PMID:18501721

  3. Inhibition of protein prenylation down-regulates signalling by inflammatory mediators in human keratinocytes.

    PubMed

    Alaei, P; MacNulty, E E; Ryder, N S

    1996-05-01

    Several inflammatory mediators have been shown to activate phospholipase C in human keratinocytes via GTP-binding protein-coupled receptors. Since GTP-binding proteins are prenylated proteins, we have examined the role of prenylation in signal transduction in HaCaT keratinocytes. Indirect inhibition of prenylation with the HMG CoA reductase inhibitors fluvastatin or compactin decreased bradykinin-stimulated inositol 1,4,5-triphosphate generation. This effect was abolished by mevalonic acid but not by serum, indicating a requirement for a non-sterol metabolite for signal generation. The BK response was also inhibited by zaragozic acids B and C, known inhibitors of prenyl protein transferases. These results suggest that protein prenylation may be a novel therapeutic target in dermatological conditions where an up-regulation of the inositol lipid pathway has been demonstrated. PMID:8630058

  4. PML tumor suppressor is regulated by HIPK2-mediated phosphorylation in response to DNA damage.

    PubMed

    Gresko, E; Ritterhoff, S; Sevilla-Perez, J; Roscic, A; Fröbius, K; Kotevic, I; Vichalkovski, A; Hess, D; Hemmings, B A; Schmitz, M L

    2009-02-01

    The promyelocytic leukemia (PML) tumor suppressor protein, a central regulator of cell proliferation and apoptosis, is frequently fused to the retinoic acid receptor-alpha (RARalpha) in acute PML. Here we show the interaction of PML with another tumor suppressor protein, the serine/threonine kinase homeodomain-interacting protein kinase (HIPK2). In response to DNA damage, HIPK2 phosphorylates PML at serines 8 and 38. Although HIPK2-mediated phosphorylation of PML occurs early during the DNA damage response, the oncogenic PML-RARalpha fusion protein is phosphorylated with significantly delayed kinetics. DNA damage or HIPK2 expression leads to the stabilization of PML and PML-RARalpha proteins. The N-terminal phosphorylation sites contribute to the DNA damage-induced PML SUMOylation and are required for the ability of PML to cooperate with HIPK2 for the induction of cell death. PMID:19015637

  5. An electrostatic mechanism for Ca(2+)-mediated regulation of gap junction channels.

    PubMed

    Bennett, Brad C; Purdy, Michael D; Baker, Kent A; Acharya, Chayan; McIntire, William E; Stevens, Raymond C; Zhang, Qinghai; Harris, Andrew L; Abagyan, Ruben; Yeager, Mark

    2016-01-01

    Gap junction channels mediate intercellular signalling that is crucial in tissue development, homeostasis and pathologic states such as cardiac arrhythmias, cancer and trauma. To explore the mechanism by which Ca(2+) blocks intercellular communication during tissue injury, we determined the X-ray crystal structures of the human Cx26 gap junction channel with and without bound Ca(2+). The two structures were nearly identical, ruling out both a large-scale structural change and a local steric constriction of the pore. Ca(2+) coordination sites reside at the interfaces between adjacent subunits, near the entrance to the extracellular gap, where local, side chain conformational rearrangements enable Ca(2+)chelation. Computational analysis revealed that Ca(2+)-binding generates a positive electrostatic barrier that substantially inhibits permeation of cations such as K(+) into the pore. Our results provide structural evidence for a unique mechanism of channel regulation: ionic conduction block via an electrostatic barrier rather than steric occlusion of the channel pore. PMID:26753910

  6. Long non-coding RNA-mediated regulation of glucose homeostasis and diabetes

    PubMed Central

    Sun, Xinghui; Wong, Danny

    2016-01-01

    Long non-coding RNAs (lncRNAs) represent an important class of non-coding RNAs that plays key roles in regulating the expression of genes in health and disease. Accumulating genetic, experimental, and epidemiological studies highlight a growing list of lncRNAs that control glucose homeostasis and diabetic pathologies and complications. Through interactions with chromatin, RNA, and protein, lncRNAs modulate chromatin modification, mRNA stability, microRNA activity, and the function of proteins such as transcription factors. This review highlights emerging concepts in lncRNA-mediated control of glucose homeostasis as well as some of the challenges and therapeutic opportunities in the pathogenesis of diabetes and its complications. PMID:27335687

  7. IRTKS negatively regulates antiviral immunity through PCBP2 sumoylation-mediated MAVS degradation

    PubMed Central

    Xia, Pengyan; Wang, Shuo; Xiong, Zhen; Ye, Buqing; Huang, Li-Yu; Han, Ze-Guang; Fan, Zusen

    2015-01-01

    RNA virus infection is recognized by the RIG-I family of receptors that activate the mitochondrial adaptor MAVS, leading to the clearance of viruses. Antiviral signalling activation requires strict modulation to avoid damage to the host from exacerbated inflammation. Insulin receptor tyrosine kinase substrate (IRTKS) participates in actin bundling and insulin signalling and its deficiency causes insulin resistance. However, whether IRTKS is involved in the regulation of innate immunity remains elusive. Here we show that IRTKS deficiency causes enhanced innate immune responses against RNA viruses. IRTKS-mediated suppression of antiviral responses depends on the RIG-I-MAVS signalling pathway. IRTKS recruits the E2 ligase Ubc9 to sumoylate PCBP2 in the nucleus, which causes its cytoplasmic translocation during viral infection. The sumoylated PCBP2 associates with MAVS to initiate its degradation, leading to downregulation of antiviral responses. Thus, IRTKS functions as a negative modulator of excessive inflammation. PMID:26348439

  8. Bim mediates mitochondria-regulated particulate matter-induced apoptosis in alveolar epithelial cells

    PubMed Central

    Zhang, J.; Ghio, A.J.; Chang, W.; Kamdar, O.; Rosen, G.D.; Upadhyay, D.

    2007-01-01

    We studied the role of Bim, a pro-apoptotic BCL-2 family member in Airborne particulate matter (PM 2.5 μm)-induced apoptosis in alveolar epithelial cells (AEC). PM induced AEC apoptosis by causing significant reduction of mitochondrial membrane potential and increase in caspase-9, caspase-3 and PARP-1 activation. PM upregulated pro-apoptotic protein Bim and enhanced translocation of Bim to the mitochondria. ShRNABim blocked PM-induced apoptosis by preventing activation of the mitochondrial death pathway suggesting a role of Bim in the regulation of mitochondrial pathway in AEC. Accordingly, we provide the evidence that Bim mediates PM-induced apoptosis via mitochondrial pathway. PMID:17716672

  9. Negative Regulation of Interferon-induced Transmembrane Protein 3 by SET7-mediated Lysine Monomethylation*

    PubMed Central

    Shan, Zhao; Han, Qinglin; Nie, Jia; Cao, Xuezhi; Chen, Zuojia; Yin, Shuying; Gao, Yayi; Lin, Fang; Zhou, Xiaohui; Xu, Ke; Fan, Huimin; Qian, Zhikang; Sun, Bing; Zhong, Jin; Li, Bin; Tsun, Andy

    2013-01-01

    Although lysine methylation is classically known to regulate histone function, its role in modulating antiviral restriction factor activity remains uncharacterized. Interferon-induced transmembrane protein 3 (IFITM3) was found monomethylated on its lysine 88 residue (IFITM3-K88me1) to reduce its antiviral activity, mediated by the lysine methyltransferase SET7. Vesicular stomatitis virus and influenza A virus infection increased IFITM3-K88me1 levels by promoting the interaction between IFITM3 and SET7, suggesting that this pathway could be hijacked to support infection; conversely, IFN-α reduced IFITM3-K88me1 levels. These findings may have important implications in the design of therapeutics targeting protein methylation against infectious diseases. PMID:24129573

  10. An electrostatic mechanism for Ca2+-mediated regulation of gap junction channels

    PubMed Central

    Bennett, Brad C.; Purdy, Michael D.; Baker, Kent A.; Acharya, Chayan; McIntire, William E.; Stevens, Raymond C.; Zhang, Qinghai; Harris, Andrew L.; Abagyan, Ruben; Yeager, Mark

    2016-01-01

    Gap junction channels mediate intercellular signalling that is crucial in tissue development, homeostasis and pathologic states such as cardiac arrhythmias, cancer and trauma. To explore the mechanism by which Ca2+ blocks intercellular communication during tissue injury, we determined the X-ray crystal structures of the human Cx26 gap junction channel with and without bound Ca2+. The two structures were nearly identical, ruling out both a large-scale structural change and a local steric constriction of the pore. Ca2+ coordination sites reside at the interfaces between adjacent subunits, near the entrance to the extracellular gap, where local, side chain conformational rearrangements enable Ca2+chelation. Computational analysis revealed that Ca2+-binding generates a positive electrostatic barrier that substantially inhibits permeation of cations such as K+ into the pore. Our results provide structural evidence for a unique mechanism of channel regulation: ionic conduction block via an electrostatic barrier rather than steric occlusion of the channel pore. PMID:26753910

  11. CHIP-mediated degradation of transglutaminase 2 negatively regulates tumor growth and angiogenesis in renal cancer.

    PubMed

    Min, B; Park, H; Lee, S; Li, Y; Choi, J-M; Lee, J Y; Kim, J; Choi, Y D; Kwon, Y-G; Lee, H-W; Bae, S-C; Yun, C-O; Chung, K C

    2016-07-14

    The multifunctional enzyme transglutaminase 2 (TG2) primarily catalyzes cross-linking reactions of proteins via (γ-glutamyl) lysine bonds. Several recent findings indicate that altered regulation of intracellular TG2 levels affects renal cancer. Elevated TG2 expression is observed in renal cancer. However, the molecular mechanism underlying TG2 degradation is not completely understood. Carboxyl-terminus of Hsp70-interacting protein (CHIP) functions as an ubiquitin E3 ligase. Previous studies reveal that CHIP deficiency mice displayed a reduced life span with accelerated aging in kidney tissues. Here we show that CHIP promotes polyubiquitination of TG2 and its subsequent proteasomal degradation. In addition, TG2 upregulation contributes to enhanced kidney tumorigenesis. Furthermore, CHIP-mediated TG2 downregulation is critical for the suppression of kidney tumor growth and angiogenesis. Notably, our findings are further supported by decreased CHIP expression in human renal cancer tissues and renal cancer cells. The present work reveals that CHIP-mediated TG2 ubiquitination and proteasomal degradation represent a novel regulatory mechanism that controls intracellular TG2 levels. Alterations in this pathway result in TG2 hyperexpression and consequently contribute to renal cancer. PMID:26568304

  12. Multiple regulation of Rad51-mediated homologous recombination by fission yeast Fbh1.

    PubMed

    Tsutsui, Yasuhiro; Kurokawa, Yumiko; Ito, Kentaro; Siddique, Md Shahjahan P; Kawano, Yumiko; Yamao, Fumiaki; Iwasaki, Hiroshi

    2014-08-01

    Fbh1, an F-box helicase related to bacterial UvrD, has been proposed to modulate homologous recombination in fission yeast. We provide several lines of evidence for such modulation. Fbh1, but not the related helicases Srs2 and Rqh1, suppressed the formation of crossover recombinants from single HO-induced DNA double-strand breaks. Purified Fbh1 in complex with Skp1 (Fbh1-Skp1 complex) inhibited Rad51-driven DNA strand exchange by disrupting Rad51 nucleoprotein filaments in an ATP-dependent manner; this disruption was alleviated by the Swi5-Sfr1 complex, an auxiliary activator of Rad51. In addition, the reconstituted SCFFbh1 complex, composed of purified Fbh1-Skp1 and Pcu1-Rbx1, displayed ubiquitin-ligase E3 activity toward Rad51. Furthermore, Fbh1 reduced the protein level of Rad51 in stationary phase in an F-box-dependent, but not in a helicase domain-independent manner. These results suggest that Fbh1 negatively regulates Rad51-mediated homologous recombination via its two putative, unrelated activities, namely DNA unwinding/translocation and ubiquitin ligation. In addition to its anti-recombinase activity, we tentatively suggest that Fbh1 might also have a pro-recombination role in vivo, because the Fbh1-Skp1 complex stimulated Rad51-mediated strand exchange in vitro after strand exchange had been initiated. PMID:25165823

  13. A conserved polybasic domain mediates plasma membrane targeting of Lgl and its regulation by hypoxia

    PubMed Central

    Dong, Wei; Zhang, Xuejing; Liu, Weijie; Chen, Yi-jiun; Huang, Juan; Austin, Erin; Celotto, Alicia M.; Jiang, Wendy Z.; Palladino, Michael J.; Jiang, Yu; Hammond, Gerald R.V.

    2015-01-01

    Lethal giant larvae (Lgl) plays essential and conserved functions in regulating both cell polarity and tumorigenesis in Drosophila melanogaster and vertebrates. It is well recognized that plasma membrane (PM) or cell cortex localization is crucial for Lgl function in vivo, but its membrane-targeting mechanisms remain poorly understood. Here, we discovered that hypoxia acutely and reversibly inhibits Lgl PM targeting through a posttranslational mechanism that is independent of the well-characterized atypical protein kinase C (aPKC) or Aurora kinase–mediated phosphorylations. Instead, we identified an evolutionarily conserved polybasic (PB) domain that targets Lgl to the PM via electrostatic binding to membrane phosphatidylinositol phosphates. Such PB domain–mediated PM targeting is inhibited by hypoxia, which reduces inositol phospholipid levels on the PM through adenosine triphosphate depletion. Moreover, Lgl PB domain contains all the identified phosphorylation sites of aPKC and Aurora kinases, providing a molecular mechanism by which phosphorylations neutralize the positive charges on the PB domain to inhibit Lgl PM targeting. PMID:26483556

  14. NADPH oxidases regulate septin-mediated cytoskeletal remodeling during plant infection by the rice blast fungus.

    PubMed

    Ryder, Lauren S; Dagdas, Yasin F; Mentlak, Thomas A; Kershaw, Michael J; Thornton, Christopher R; Schuster, Martin; Chen, Jisheng; Wang, Zonghua; Talbot, Nicholas J

    2013-02-19

    The rice blast fungus Magnaporthe oryzae infects plants with a specialized cell called an appressorium, which uses turgor to drive a rigid penetration peg through the rice leaf cuticle. Here, we show that NADPH oxidases (Nox) are necessary for septin-mediated reorientation of the F-actin cytoskeleton to facilitate cuticle rupture and plant cell invasion. We report that the Nox2-NoxR complex spatially organizes a heteroligomeric septin ring at the appressorium pore, required for assembly of a toroidal F-actin network at the point of penetration peg emergence. Maintenance of the cortical F-actin network during plant infection independently requires Nox1, a second NADPH oxidase, which is necessary for penetration hypha elongation. Organization of F-actin in appressoria is disrupted by application of antioxidants, whereas latrunculin-mediated depolymerization of appressorial F-actin is competitively inhibited by reactive oxygen species, providing evidence that regulated synthesis of reactive oxygen species by fungal NADPH oxidases directly controls septin and F-actin dynamics. PMID:23382235

  15. Polycomb PRC2 complex mediates epigenetic silencing of a critical osteogenic master regulator in the hippocampus.

    PubMed

    Aguilar, Rodrigo; Bustos, Fernando J; Saez, Mauricio; Rojas, Adriana; Allende, Miguel L; van Wijnen, Andre J; van Zundert, Brigitte; Montecino, Martin

    2016-08-01

    During hippocampal neuron differentiation, the expression of critical inducers of non-neuronal cell lineages must be efficiently silenced. Runx2 transcription factor is the master regulator of mesenchymal cells responsible for intramembranous osteoblast differentiation and formation of the craniofacial bone tissue that surrounds and protects the central nervous system (CNS) in mammalian embryos. The molecular mechanisms that mediate silencing of the Runx2 gene and its downstream target osteogenic-related genes in neuronal cells have not been explored. Here, we assess the epigenetic mechanisms that mediate silencing of osteoblast-specific genes in CNS neurons. In particular, we address the contribution of histone epigenetic marks and histone modifiers on the silencing of the Runx2/p57 bone-related isoform in rat hippocampal tissues at embryonic to adult stages. Our results indicate enrichment of repressive chromatin histone marks and of the Polycomb PRC2 complex at the Runx2/p57 promoter region. Knockdown of PRC2 H3K27-methyltransferases Ezh2 and Ezh1, or forced expression of the Trithorax/COMPASS subunit Wdr5 activates Runx2/p57 mRNA expression in both immature and mature hippocampal cells. Together these results indicate that complementary epigenetic mechanisms progressively and efficiently silence critical osteoblastic genes during hippocampal neuron differentiation. PMID:27216774

  16. Complement Component 3 Is Regulated by TWIST1 and Mediates Epithelial-Mesenchymal Transition.

    PubMed

    Cho, Min Soon; Rupaimoole, Rajesha; Choi, Hyun-Jin; Noh, Kyunghee; Chen, Jichao; Hu, Qianghua; Sood, Anil K; Afshar-Kharghan, Vahid

    2016-02-01

    We have previously shown that complement component 3 (C3) is secreted by malignant epithelial cells. To understand the mechanism of upregulation of C3 expression in tumor cells, we studied the C3 promoter and identified that twist basic helix-loop-helix transcription factor 1 (TWIST1) binds to the C3 promoter and enhances its expression. Because TWIST1 mediates epithelial-mesenchymal transition (EMT), we studied the effect of C3 on EMT and found that C3 decreased E-cadherin expression on cancer cells and promoted EMT. We showed that C3-induced reduction in E-cadherin expression in ovarian cancer cells was mediated by C3a and is Krüppel-like factor 5 dependent. We investigated the association between TWIST1 and C3 in malignant tumors and in murine embryos. TWIST1 and C3 colocalized at the invasive tumor edges, and in the neural crest and limb buds of mouse embryos. Our results identified TWIST1 as a transcription factor that regulates C3 expression during pathologic and physiologic EMT. PMID:26718342

  17. SET9-Mediated Regulation of TGF-β Signaling Links Protein Methylation to Pulmonary Fibrosis.

    PubMed

    Elkouris, Maximilianos; Kontaki, Haroula; Stavropoulos, Athanasios; Antonoglou, Anastasia; Nikolaou, Kostas C; Samiotaki, Martina; Szantai, Eszter; Saviolaki, Dimitra; Brown, Peter J; Sideras, Paschalis; Panayotou, George; Talianidis, Iannis

    2016-06-21

    TGF-β signaling regulates a variety of cellular processes, including proliferation, apoptosis, differentiation, immune responses, and fibrogenesis. Here, we describe a lysine methylation-mediated mechanism that controls the pro-fibrogenic activity of TGF-β. We find that the methyltransferase Set9 potentiates TGF-β signaling by targeting Smad7, an inhibitory downstream effector. Smad7 methylation promotes interaction with the E3 ligase Arkadia and, thus, ubiquitination-dependent degradation. Depletion or pharmacological inhibition of Set9 results in elevated Smad7 protein levels and inhibits TGF-β-dependent expression of genes encoding extracellular matrix components. The inhibitory effect of Set9 on TGF-β-mediated extracellular matrix production is further demonstrated in mouse models of pulmonary fibrosis. Lung fibrosis induced by bleomycin or Ad-TGF-β treatment was highly compromised in Set9-deficient mice. These results uncover a complex regulatory interplay among multiple Smad7 modifications and highlight the possibility that protein methyltransferases may represent promising therapeutic targets for treating lung fibrosis. PMID:27292644

  18. Syndecan-4 negatively regulates antiviral signalling by mediating RIG-I deubiquitination via CYLD

    PubMed Central

    Lin, Wei; Zhang, Jing; Lin, Haiyan; Li, Zexing; Sun, Xiaofeng; Xin, Di; Yang, Meng; Sun, Liwei; Li, Lin; Wang, Hongmei; Chen, Dahua; Sun, Qinmiao

    2016-01-01

    Retinoic acid-inducible gene I (RIG-I) plays important roles in pathogen recognition and antiviral signalling transduction. Here we show that syndecan-4 (SDC4) is a RIG-I-interacting partner identified in a yeast two-hybrid screen. We find that SDC4 negatively regulates the RIG-I-mediated antiviral signalling in a feedback-loop control manner. The genetic evidence obtained by using knockout mice further emphasizes this biological role of SDC4 in antiviral signalling. Mechanistically, we show that SDC4 interacts with both RIG-I and deubiquitinase CYLD via its carboxyl-terminal intracellular region. SDC4 likely promotes redistribution of RIG-I and CYLD in a perinuclear pattern post viral infection, and thus enhances the RIG-I–CYLD interaction and potentiates the K63-linked deubiquitination of RIG-I. Collectively, our findings uncover a mechanism by which SDC4 antagonizes the activation of RIG-I in a CYLD-mediated deubiquitination-dependent process, thereby balancing antiviral signalling to avoid deleterious effects on host cells. PMID:27279133

  19. Mitochondria-Mediated Protein Regulation Mechanism of Polymorphs-Dependent Inhibition of Nanoselenium on Cancer Cells.

    PubMed

    Wang, Ge; Guo, Yuming; Yang, Gai; Yang, Lin; Ma, Xiaoming; Wang, Kui; Zhu, Lin; Sun, Jiaojiao; Wang, Xiaobing; Zhang, Hua

    2016-01-01

    The present study was (i) to prepare two types of selenium nanoparticles, namely an amorphous form of selenium quantum dots (A-SeQDs) and a crystalline form of selenium quantum dots (C-SeQDs); and (ii) to investigate the nano-bio interactions of A-SeQDs and C-SeQDs in MCF-7, HepG2, HeLa, NIH/3T3, L929 cells and BRL-3A cells. It was found that A-SeQDs could induce the mitochondria-mediated apoptosis, necrosis and death of cells, while C-SeQDs had much weaker effects. This polymorphs-dependent anti-proliferative activity of nano-selenium was scarcely reported. Further investigation demonstrated that A-SeQDs could differentially regulate 61 proteins and several pathways related to stress response, protein synthesis, cell migration and cell cycle, including "p38 MAPK Signaling", "p53 Signaling", "14-3-3-mediated Signaling", "p70S6K Signaling" and "Protein Ubiquitination Pathway". This was the first report to demonstrate the involvement of protein synthesis and post-translational modification pathways in the anti-proliferative activity associated with NMs. Compared with previously fragmentary studies, this study use a nanomics approach combining bioinformatics and proteomics to systematically investigate the nano-bio interactions of selenium nanoparticles in cancer cells. PMID:27514819

  20. Atg1-mediated myosin II activation regulates autophagosome formation during starvation-induced autophagy.

    PubMed

    Tang, Hong-Wen; Wang, Yu-Bao; Wang, Shiu-Lan; Wu, Mei-Hsuan; Lin, Shu-Yu; Chen, Guang-Chao

    2011-02-16

    Autophagy is a membrane-mediated degradation process of macromolecule recycling. Although the formation of double-membrane degradation vesicles (autophagosomes) is known to have a central role in autophagy, the mechanism underlying this process remains elusive. The serine/threonine kinase Atg1 has a key role in the induction of autophagy. In this study, we show that overexpression of Drosophila Atg1 promotes the phosphorylation-dependent activation of the actin-associated motor protein myosin II. A novel myosin light chain kinase (MLCK)-like protein, Spaghetti-squash activator (Sqa), was identified as a link between Atg1 and actomyosin activation. Sqa interacts with Atg1 through its kinase domain and is a substrate of Atg1. Significantly, myosin II inhibition or depletion of Sqa compromised the formation of autophagosomes under starvation conditions. In mammalian cells, we found that the Sqa mammalian homologue zipper-interacting protein kinase (ZIPK) and myosin II had a critical role in the regulation of starvation-induced autophagy and mammalian Atg9 (mAtg9) trafficking when cells were deprived of nutrients. Our findings provide evidence of a link between Atg1 and the control of Atg9-mediated autophagosome formation through the myosin II motor protein. PMID:21169990

  1. NK cell regulation of CD4 T cell-mediated graft-versus-host disease.

    PubMed

    Noval Rivas, Magali; Hazzan, Marc; Weatherly, Kathleen; Gaudray, Florence; Salmon, Isabelle; Braun, Michel Y

    2010-06-15

    CD3-negative NK cells are granular lymphocytes capable of producing inflammatory cytokines and killing malignant, infected, or stressed cells. We have recently observed a new role for NK cells in the control of the proliferation of CD4 T cells under persistent antigenic stimulation. Monoclonal anti-male CD4 T cells transferred into Rag2-/- male recipients did not expand or were rapidly eliminated. Remarkably, T cells transferred into NK cell-deficient Rag2-/- Il-2Rgammac-/- male hosts expanded extensively and mediated tissue lesions usually observed in chronic graft-versus-host disease (GVHD). T cell failure to proliferate and to induce chronic GVHD was the result of NK cell activity, because depletion of the recipient's NK1.1+ cells by Ab treatment induced T cell expansion and chronic GVHD. T cells under chronic Ag stimulation upregulated ligands of the activating receptor NKG2D, and regulatory activity of NK cells was inhibited by the injection of Abs directed to NKG2D. On the contrary, blocking NKG2A inhibitory receptors did not increase NK cell regulatory activity. Finally, we show that NK regulation of T cell expansion did not involve perforin-mediated lytic activity of NK cells, but depended on T cell surface expression of a functional Fas molecule. These results highlight the potential role played by NK cells in controlling the Ag-specific CD4+ T cells responsible for chronic GVHD. PMID:20488796

  2. The H3K9 methyltransferase Setdb1 regulates TLR4-mediated inflammatory responses in macrophages

    PubMed Central

    Hachiya, Rumi; Shiihashi, Takuya; Shirakawa, Ibuki; Iwasaki, Yorihiro; Matsumura, Yoshihiro; Oishi, Yumiko; Nakayama, Yukiteru; Miyamoto, Yoshihiro; Manabe, Ichiro; Ochi, Kozue; Tanaka, Miyako; Goda, Nobuhito; Sakai, Juro; Suganami, Takayoshi; Ogawa, Yoshihiro

    2016-01-01

    Proinflammatory cytokine production in macrophages involves multiple regulatory mechanisms, which are affected by environmental and intrinsic stress. In particular, accumulating evidence has suggested epigenetic control of macrophage differentiation and function mainly in vitro. SET domain, bifurcated 1 (Setdb1, also known as Eset) is a histone 3 lysine 9 (H3K9)-specific methyltransferase and is essential for early development of embryos. Here we demonstrate that Setdb1 in macrophages potently suppresses Toll-like receptor 4 (TLR4)-mediated expression of proinflammatory cytokines including interleukin-6 through its methyltransferase activity. As a molecular mechanism, Setdb1-deficiency decreases the basal H3K9 methylation levels and augments TLR4-mediated NF-κB recruitment on the proximal promoter region of interleukin-6, thereby accelerating interleukin-6 promoter activity. Moreover, macrophage-specific Setdb1-knockout mice exhibit higher serum interleukin-6 concentrations in response to lipopolysaccharide challenge and are more susceptible to endotoxin shock than wildtype mice. This study provides evidence that the H3K9 methyltransferase Setdb1 is a novel epigenetic regulator of proinflammatory cytokine expression in macrophages in vitro and in vivo. Our data will shed insight into the better understanding of how the immune system reacts to a variety of conditions. PMID:27349785

  3. Mitochondria-Mediated Protein Regulation Mechanism of Polymorphs-Dependent Inhibition of Nanoselenium on Cancer Cells

    PubMed Central

    Wang, Ge; Guo, Yuming; Yang, Gai; Yang, Lin; Ma, Xiaoming; Wang, Kui; Zhu, Lin; Sun, Jiaojiao; Wang, Xiaobing; Zhang, Hua

    2016-01-01

    The present study was (i) to prepare two types of selenium nanoparticles, namely an amorphous form of selenium quantum dots (A-SeQDs) and a crystalline form of selenium quantum dots (C-SeQDs); and (ii) to investigate the nano-bio interactions of A-SeQDs and C-SeQDs in MCF-7, HepG2, HeLa, NIH/3T3, L929 cells and BRL-3A cells. It was found that A-SeQDs could induce the mitochondria-mediated apoptosis, necrosis and death of cells, while C-SeQDs had much weaker effects. This polymorphs-dependent anti-proliferative activity of nano-selenium was scarcely reported. Further investigation demonstrated that A-SeQDs could differentially regulate 61 proteins and several pathways related to stress response, protein synthesis, cell migration and cell cycle, including “p38 MAPK Signaling”, “p53 Signaling”, “14-3-3-mediated Signaling”, “p70S6K Signaling” and “Protein Ubiquitination Pathway”. This was the first report to demonstrate the involvement of protein synthesis and post-translational modification pathways in the anti-proliferative activity associated with NMs. Compared with previously fragmentary studies, this study use a nanomics approach combining bioinformatics and proteomics to systematically investigate the nano-bio interactions of selenium nanoparticles in cancer cells. PMID:27514819

  4. Ciliopathy proteins regulate paracrine signaling by modulating proteasomal degradation of mediators

    PubMed Central

    Liu, Yangfan P.; Tsai, I-Chun; Morleo, Manuela; Oh, Edwin C.; Leitch, Carmen C.; Massa, Filomena; Lee, Byung-Hoon; Parker, David S.; Finley, Daniel; Zaghloul, Norann A.; Franco, Brunella; Katsanis, Nicholas

    2014-01-01

    Cilia are critical mediators of paracrine signaling; however, it is unknown whether proteins that contribute to ciliopathies converge on multiple paracrine pathways through a common mechanism. Here, we show that loss of cilopathy-associated proteins Bardet-Biedl syndrome 4 (BBS4) or oral-facial-digital syndrome 1 (OFD1) results in the accumulation of signaling mediators normally targeted for proteasomal degradation. In WT cells, several BBS proteins and OFD1 interacted with proteasomal subunits, and loss of either BBS4 or OFD1 led to depletion of multiple subunits from the centrosomal proteasome. Furthermore, overexpression of proteasomal regulatory components or treatment with proteasomal activators sulforaphane (SFN) and mevalonolactone (MVA) ameliorated signaling defects in cells lacking BBS1, BBS4, and OFD1, in morphant zebrafish embryos, and in induced neurons from Ofd1-deficient mice. Finally, we tested the hypothesis that other proteasome-dependent pathways not known to be associated with ciliopathies are defective in the absence of ciliopathy proteins. We found that loss of BBS1, BBS4, or OFD1 led to decreased NF-κB activity and concomitant IκBβ accumulation and that these defects were ameliorated with SFN treatment. Taken together, our data indicate that basal body proteasomal regulation governs paracrine signaling pathways and suggest that augmenting proteasomal function might benefit ciliopathy patients. PMID:24691443

  5. Nuclear DISC1 regulates CRE-mediated gene transcription and sleep homeostasis in the fruit fly.

    PubMed

    Sawamura, N; Ando, T; Maruyama, Y; Fujimuro, M; Mochizuki, H; Honjo, K; Shimoda, M; Toda, H; Sawamura-Yamamoto, T; Makuch, L A; Hayashi, A; Ishizuka, K; Cascella, N G; Kamiya, A; Ishida, N; Tomoda, T; Hai, T; Furukubo-Tokunaga, K; Sawa, A

    2008-12-01

    Disrupted-in-schizophrenia-1 (DISC1) is one of major susceptibility factors for a wide range of mental illnesses, including schizophrenia, bipolar disorder, major depression and autism spectrum conditions. DISC1 is located in several subcellular domains, such as the centrosome and the nucleus, and interacts with various proteins, including NudE-like (NUDEL/NDEL1) and activating transcription factor 4 (ATF4)/CREB2. Nevertheless, a role for DISC1 in vivo remains to be elucidated. Therefore, we have generated a Drosophila model for examining normal functions of DISC1 in living organisms. DISC1 transgenic flies with preferential accumulation of exogenous human DISC1 in the nucleus display disturbance in sleep homeostasis, which has been reportedly associated with CREB signaling/CRE-mediated gene transcription. Thus, in mammalian cells, we characterized nuclear DISC1, and identified a subset of nuclear DISC1 that colocalizes with the promyelocytic leukemia (PML) bodies, a nuclear compartment for gene transcription. Furthermore, we identified three functional cis-elements that regulate the nuclear localization of DISC1. We also report that DISC1 interacts with ATF4/CREB2 and a corepressor N-CoR, modulating CRE-mediated gene transcription. PMID:18762802

  6. Deficits in emotion regulation mediate the relationship between childhood abuse and later eating disorder symptoms.

    PubMed

    Burns, Erin E; Fischer, Sarah; Jackson, Joan L; Harding, Hilary G

    2012-01-01

    The present study examined the relationship of child maltreatment to both emotion dysregulation and subsequent eating pathology. In an effort to extend previous research, the authors examined the unique impact of childhood emotional abuse (CEA) on emotion dysregulation and eating disorder (ED) symptoms while controlling for the effects of sexual and physical abuse. Structural equation modeling was utilized to simultaneously examine the effects of all three abuse types on multiple dependent variables as well as examine whether deficits in emotion regulation mediated the relationship between abuse and eating pathology. Results from a survey of 1,254 female college students revealed significant paths from abuse subtypes to specific eating disorder symptoms, with CEA evidencing the strongest association with ED symptoms. Additionally, emotion dysregulation was positively associated with ED symptoms, and mediated the effects of emotional abuse on symptoms. Findings support previous research on the enduring effects of emotional abuse as well as highlight the importance of the assessment of CEA in the treatment of ED symptoms. PMID:22265934

  7. The H3K9 methyltransferase Setdb1 regulates TLR4-mediated inflammatory responses in macrophages.

    PubMed

    Hachiya, Rumi; Shiihashi, Takuya; Shirakawa, Ibuki; Iwasaki, Yorihiro; Matsumura, Yoshihiro; Oishi, Yumiko; Nakayama, Yukiteru; Miyamoto, Yoshihiro; Manabe, Ichiro; Ochi, Kozue; Tanaka, Miyako; Goda, Nobuhito; Sakai, Juro; Suganami, Takayoshi; Ogawa, Yoshihiro

    2016-01-01

    Proinflammatory cytokine production in macrophages involves multiple regulatory mechanisms, which are affected by environmental and intrinsic stress. In particular, accumulating evidence has suggested epigenetic control of macrophage differentiation and function mainly in vitro. SET domain, bifurcated 1 (Setdb1, also known as Eset) is a histone 3 lysine 9 (H3K9)-specific methyltransferase and is essential for early development of embryos. Here we demonstrate that Setdb1 in macrophages potently suppresses Toll-like receptor 4 (TLR4)-mediated expression of proinflammatory cytokines including interleukin-6 through its methyltransferase activity. As a molecular mechanism, Setdb1-deficiency decreases the basal H3K9 methylation levels and augments TLR4-mediated NF-κB recruitment on the proximal promoter region of interleukin-6, thereby accelerating interleukin-6 promoter activity. Moreover, macrophage-specific Setdb1-knockout mice exhibit higher serum interleukin-6 concentrations in response to lipopolysaccharide challenge and are more susceptible to endotoxin shock than wildtype mice. This study provides evidence that the H3K9 methyltransferase Setdb1 is a novel epigenetic regulator of proinflammatory cytokine expression in macrophages in vitro and in vivo. Our data will shed insight into the better understanding of how the immune system reacts to a variety of conditions. PMID:27349785

  8. EMX1 regulates NRP1-mediated wiring of the mouse anterior cingulate cortex.

    PubMed

    Lim, Jonathan W C; Donahoo, Amber-Lee S; Bunt, Jens; Edwards, Timothy J; Fenlon, Laura R; Liu, Ying; Zhou, Jing; Moldrich, Randal X; Piper, Michael; Gobius, Ilan; Bailey, Timothy L; Wray, Naomi R; Kessaris, Nicoletta; Poo, Mu-Ming; Rubenstein, John L R; Richards, Linda J

    2015-11-01

    Transcription factors act during cortical development as master regulatory genes that specify cortical arealization and cellular identities. Although numerous transcription factors have been identified as being crucial for cortical development, little is known about their downstream targets and how they mediate the emergence of specific neuronal connections via selective axon guidance. The EMX transcription factors are essential for early patterning of the cerebral cortex, but whether EMX1 mediates interhemispheric connectivity by controlling corpus callosum formation remains unclear. Here, we demonstrate that in mice on the C57Bl/6 background EMX1 plays an essential role in the midline crossing of an axonal subpopulation of the corpus callosum derived from the anterior cingulate cortex. In the absence of EMX1, cingulate axons display reduced expression of the axon guidance receptor NRP1 and form aberrant axonal bundles within the rostral corpus callosum. EMX1 also functions as a transcriptional activator of Nrp1 expression in vitro, and overexpression of this protein in Emx1 knockout mice rescues the midline-crossing phenotype. These findings reveal a novel role for the EMX1 transcription factor in establishing cortical connectivity by regulating the interhemispheric wiring of a subpopulation of neurons within the mouse anterior cingulate cortex. PMID:26534986

  9. Regulation of adenovirus-mediated elafin transgene expression by bacterial lipopolysaccharide.

    PubMed

    Simpson, A J; Cunningham, G A; Porteous, D J; Haslett, C; Sallenave, J M

    2001-07-20

    Lipopolysaccharide (LPS) is a mediator of inflammatory lung injury. Selective augmentation of host defense molecules such as elafin (an elastase inhibitor with antimicrobial activity) at the onset of pulmonary inflammation is an attractive potential therapeutic strategy. The aim of this study was to determine whether elafin expression could be induced by LPS administered after transfection with adenovirus (Ad) encoding human elafin downstream of the murine cytomegalovirus (CMV) promoter (known to be potentially responsive to LPS). In addition, we aimed to determine the effect of local elafin augmentation on neutrophil migration to the lung. LPS significantly up-regulated elafin expression from pulmonary epithelial cells transfected with Ad-elafin in vitro. In murine airways expression of human elafin was achieved using doses low enough (3 x 10(7) plaque forming units) to circumvent overt vector-induced inflammation. LPS significantly up-regulated human elafin secretion in murine airways treated with Ad-elafin [117 ng/ml in bronchoalveolar lavage fluid (BALF) after LPS administration, 5.9 ng/ml after PBS, p < 0.01)]. Over-expression of elafin significantly augmented LPS-mediated neutrophil migration into the airways in vivo (1.30 x 10(6) neutrophils in BALF after Ad-elafin/LPS treatment, 0.54 x 10(6) after Ad-lacZ/LPS (p < 0.05), 0.63 x 10(6) after PBS/LPS (p < 0.05)) and significantly enhanced human neutrophil migration in vitro. These data suggest novel functions for elafin in neutrophil migration, and that judicious selection of promoters may allow single, low-dose adenoviral administration to effect inflammation-specific expression of potentially therapeutic transgenes. PMID:11485631

  10. Oncogenic mutations in intestinal adenomas regulate Bim-mediated apoptosis induced by TGF-β

    PubMed Central

    Wiener, Zoltán; Band, Arja M.; Kallio, Pauliina; Högström, Jenny; Hyvönen, Ville; Kaijalainen, Seppo; Ritvos, Olli; Haglund, Caj; Kruuna, Olli; Robine, Sylvie; Louvard, Daniel; Ben-Neriah, Yinon; Alitalo, Kari

    2014-01-01

    In the majority of microsatellite-stable colorectal cancers (CRCs), an initiating mutation occurs in the adenomatous polyposis coli (APC) or β-catenin gene, activating the β-catenin/TCF pathway. The progression of resulting adenomas is associated with oncogenic activation of KRas and inactivation of the p53 and TGF-β/Smad functions. Most established CRC cell lines contain mutations in the TGF-β/Smad pathway, but little is known about the function of TGF-β in the early phases of intestinal tumorigenesis. We used mouse and human ex vivo 3D intestinal organoid cultures and in vivo mouse models to study the effect of TGF-β on the Lgr5+ intestinal stem cells and their progeny in intestinal adenomas. We found that the TGF-β–induced apoptosis in Apc-mutant organoids, including the Lgr5+ stem cells, was mediated by up-regulation of the BH3-only proapoptotic protein Bcl-2–like protein 11 (Bim). BH3-mimetic compounds recapitulated the effect of Bim not only in the adenomas but also in human CRC organoids that had lost responsiveness to TGF-β–induced apoptosis. However, wild-type intestinal crypts were markedly less sensitive to TGF-β than Apc-mutant adenomas, whereas the KRas oncogene increased resistance to TGF-β via the activation of the Erk1/2 kinase pathway, leading to Bim down-regulation. Our studies identify Bim as a critical mediator of TGF-β–induced apoptosis in intestinal adenomas and show that the common progression mutations modify Bim levels and sensitivity to TGF-β during intestinal adenoma development. PMID:24825889

  11. Nucleus accumbens cocaine-amphetamine regulated transcript mediates food intake during novelty conflict.

    PubMed

    Burghardt, P R; Krolewski, D M; Dykhuis, K E; Ching, J; Pinawin, A M; Britton, S L; Koch, L G; Watson, S J; Akil, H

    2016-05-01

    Obesity is a persistent and pervasive problem, particularly in industrialized nations. It has come to be appreciated that the metabolic health of an individual can influence brain function and subsequent behavioral patterns. To examine the relationship between metabolic phenotype and central systems that regulate behavior, we tested rats with divergent metabolic phenotypes (Low Capacity Runner: LCR vs. High Capacity Runner: HCR) for behavioral responses to the conflict between hunger and environmental novelty using the novelty suppressed feeding (NSF) paradigm. Additionally, we measured expression of mRNA, for peptides involved in energy management, in response to fasting. Following a 24-h fast, LCR rats showed lower latencies to begin eating in a novel environment compared to HCR rats. A 48-h fast equilibrated the latency to begin eating in the novel environment. A 24-h fast differentially affected expression of cocaine-amphetamine regulated transcript (CART) mRNA in the nucleus accumbens (NAc), where 24-h of fasting reduced CART mRNA in LCR rats. Bilateral microinjections of CART 55-102 peptide into the NAc increased the latency to begin eating in the NSF paradigm following a 24-h fast in LCR rats. These results indicate that metabolic phenotype influences how animals cope with the conflict between hunger and novelty, and that these differences are at least partially mediated by CART signaling in the NAc. For individuals with poor metabolic health who have to navigate food-rich and stressful environments, changes in central systems that mediate conflicting drives may feed into the rates of obesity and exacerbate the difficulty individuals have in maintaining weight loss. PMID:26926827

  12. Regulation of alkaline ceramidase activity by the c-Src-mediated pathway.

    PubMed

    Sasaki, Hirotsune; Toyomura, Kaori; Matsuzaki, Wataru; Okamoto, Aya; Yamaguchi, Naoto; Nakamura, Hiroyuki; Murayama, Toshihiko

    2014-05-15

    Ceramidase hydrolyzes ceramide to fatty acids and sphingosine, and sphingosine is then converted to sphingosine-1-phosphate. Ceramide and sphingosine-1-phosphate act as signaling molecules. Although stimuli coupling to protein kinases-dependent systems have been shown to regulate ceramidase activity, the exact role of c-Src-mediated signal has not been elucidated. We examined the effects of the downregulation of c-Src activity and c-Src overexpression on ceramidase activity in cells. In A549, CHO, and HeLa cells labeled with a fluorescent ceramide, 4-nitrobenzo-2-oxa-1,3-diazole-labeled C6-ceramide (NBD-ceramide), the downregulation of c-Src by c-Src-shRNA and pharmacological inhibitors including SU6656 decreased levels of NBD-caproic acid. The overexpression of c-Src increased NBD-caproic acid levels in CHO and HeLa cells. Similar results were obtained in Na3VO4-treated cells having higher NBD-caproic acid levels. The downregulation and overexpression of c-Src decreased and increased ceramidase activity, respectively, in the lysates of A549 cells at pH 8.8. The ceramidase sensitivity to substrates, pH, and Ca(2+) suggest that the c-Src- and SU6656-sensitive ceramidase is alkaline ceramidase (ACER), possibly Ca(2+)-activated ACER2. Serum starvation increased both ceramidase activity at pH 8.8 and expression of ACER2. Our data suggest that c-Src-mediated signal positively regulates ACER activity in a Ca(2+)-independent manner. PMID:24708996

  13. Differential regulation of a CLC anion channel by SPAK kinase ortholog-mediated multisite phosphorylation

    PubMed Central

    Miyazaki, Hiroaki

    2012-01-01

    Shrinkage-induced inhibition of the Caenorhabditis elegans cell volume and cell cycle-dependent CLC anion channel CLH-3b occurs by concomitant phosphorylation of S742 and S747, which are located on a 175 amino acid linker domain between cystathionine-β-synthase 1 (CBS1) and CBS2. Phosphorylation is mediated by the SPAK kinase homolog GCK-3 and is mimicked by substituting serine residues with glutamate. Type 1 serine/threonine protein phosphatases mediate swelling-induced channel dephosphorylation. S742E/S747E double mutant channels are constitutively inactive and cannot be activated by cell swelling. S742E and S747E mutant channels were fully active in the absence of GCK-3 and were inactive when coexpressed with the kinase. Both channels responded to cell volume changes. However, the S747E mutant channel activated and inactivated in response to cell swelling and shrinkage, respectively, much more slowly than either wild-type or S742E mutant channels. Slower activation and inactivation of S747E was not due to altered rates of dephosphorylation or dephosphorylation-dependent conformational changes. GCK-3 binds to the 175 amino acid inter-CBS linker domain. Coexpression of wild-type CLH-3b and GCK-3 with either wild-type or S742E linkers gave rise to similar channel activity and regulation. In contrast, coexpression with the S747E linker greatly enhanced basal channel activity and increased the rate of shrinkage-induced channel inactivation. Our findings suggest the intriguing possibility that the phosphorylation state of S742 in S747E mutant channels modulates GCK-3/channel interaction and hence channel phosphorylation. These results provide a foundation for further detailed studies of the role of multisite phosphorylation in regulating CLH-3b and GCK-3 activity. PMID:22357738

  14. EGF-mediated regulation of IGFBP-3 determines esophageal epithelial cellular response to IGF-I

    PubMed Central

    Takaoka, Munenori; Smith, Caitlin E.; Mashiba, Michael K.; Okawa, Takaomi; Andl, Claudia D.; El-Deiry, Wafik S.; Nakagawa, Hiroshi

    2010-01-01

    IGF and EGF regulate various physiological and pathological processes. IGF binding protein (IGFBP)-3 regulates cell proliferation in IGF-dependent and -independent fashions. Recently, we identified IGFBP-3 as a novel EGF receptor (EGFR) downstream target molecule in primary and immortalized human esophageal epithelial cells, suggesting an interplay between the EGF and IGF signaling pathways. However, the regulatory mechanisms for IGFBP-3 expression and its functional role in esophageal cell proliferation remain to be elucidated. Herein, we report that IGFBP-3 mRNA and protein were induced upon growth factor deprivation in primary and immortalized human esophageal cells through mechanisms requiring p53-independent de novo mRNA transcription and protein synthesis. This occurred in the face of the activated phosphatidylinositol 3-OH-kinase (PI3K)/mammalian target of rapamycin (mTOR) pathway. Secreted IGFBP-3 neutralized IGFs and prevented IGF-I receptor (IGF-IR) activation. In contrast, EGF suppressed IGFBP-3 mRNA and protein expression through activation of MAPK in an EGFR-tyrosine kinase-dependent manner to restore the cellular response to IGF-I. When stably overexpressed, wild-type IGFBP-3 but not I56G/L80G/L81G (GGG) mutant IGFBP-3, which has a reduced affinity to IGFs, prevented IGF-I from activating IGF-IR and Akt as well as stimulating cell proliferation. However, unlike other cell types where IGFBP-3 exerts antiproliferative effects, neither wild-type nor GGG mutant IGFBP-3 alone affected cell proliferation or EGFR activity. These results indicate that IGF signaling is subject to negative regulation through IGFBP-3 and positive regulation by EGF, the latter of which suppresses IGFBP-3. This provides a platform for understanding the novel cross talk between EGF- and IGF-mediated pathways. PMID:16210470

  15. Calmodulin activity regulates group I metabotropic glutamate receptor-mediated signal transduction and synaptic depression.

    PubMed

    Sethna, Ferzin; Zhang, Ming; Kaphzan, Hanoch; Klann, Eric; Autio, Dawn; Cox, Charles L; Wang, Hongbing

    2016-05-01

    Group I metabotropic glutamate receptors (mGluR), including mGluR1 and mGluR 5 (mGluR1/5), are coupled to Gq and modulate activity-dependent synaptic plasticity. Direct activation of mGluR1/5 causes protein translation-dependent long-term depression (LTD). Although it has been established that intracellular Ca(2+) and the Gq-regulated signaling molecules are required for mGluR1/5 LTD, whether and how Ca(2+) regulates Gq signaling and upregulation of protein expression remain unknown. Through pharmacological inhibition, we tested the function of the Ca(2+) sensor calmodulin (CaM) in intracellular signaling triggered by the activation of mGluR1/5. CaM inhibitor N-[4-aminobutyl]-5-chloro-2-naphthalenesulfonamide hydrochloride (W13) suppressed the mGluR1/5-stimulated activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and p70-S6 kinase 1 (S6K1) in hippocampal neurons. W13 also blocked the mGluR1/5 agonist-induced synaptic depression in hippocampal slices and in anesthetized mice. Consistent with the function of CaM, inhibiting the downstream targets Ca(2+) /CaM-dependent protein kinases (CaMK) blocked ERK1/2 and S6K1 activation. Furthermore, disruption of the CaM-CaMK-ERK1/2 signaling cascade suppressed the mGluR1/5-stimulated upregulation of Arc expression. Altogether, our data suggest CaM as a new Gq signaling component for coupling Ca(2+) and protein upregulation and regulating mGluR1/5-mediated synaptic modification. PMID:26864654

  16. Regulation of Natural mRNAs by the Nonsense-Mediated mRNA Decay Pathway

    PubMed Central

    Peccarelli, Megan

    2014-01-01

    The nonsense-mediated mRNA decay (NMD) pathway is a specialized mRNA degradation pathway that degrades select mRNAs. This pathway is conserved in all eukaryotes examined so far, and it triggers the degradation of mRNAs that prematurely terminate translation. Originally identified as a pathway that degrades mRNAs with premature termination codons as a result of errors during transcription, splicing, or damage to the mRNA, NMD is now also recognized as a pathway that degrades some natural mRNAs. The degradation of natural mRNAs by NMD has been identified in multiple eukaryotes, including Saccharomyces cerevisiae, Drosophila melanogaster, Arabidopsis thaliana, and humans. S. cerevisiae is used extensively as a model to study natural mRNA regulation by NMD. Inactivation of the NMD pathway in S. cerevisiae affects approximately 10% of the transcriptome. Similar percentages of natural mRNAs in the D. melanogaster and human transcriptomes are also sensitive to the pathway, indicating that NMD is important for the regulation of gene expression in multiple organisms. NMD can either directly or indirectly regulate the decay rate of natural mRNAs. Direct NMD targets possess NMD-inducing features. This minireview focuses on the regulation of natural mRNAs by the NMD pathway, as well as the features demonstrated to target these mRNAs for decay by the pathway in S. cerevisiae. We also compare NMD-targeting features identified in S. cerevisiae with known NMD-targeting features in other eukaryotic organisms. PMID:25038084

  17. Green mechanochemical oxidative decomposition of powdery decabromodiphenyl ether with persulfate.

    PubMed

    Huang, Aizhen; Zhang, Zhimin; Wang, Nan; Zhu, Lihua; Zou, Jing

    2016-01-25

    A method was developed for efficiently degrading powdery decabromodiphenyl ether (BDE209) by using mechanochemical (MC) activation of persulfate (PS). Characteristic Raman spectra of BDE209 corresponding to CBr and CO bonds were decreased in intensity and finally disappeared as the MC reaction proceeded. The BDE209 removal was influenced by the molar ratio of PS to BDE209, the mass ratio of milling ball to reaction mixtures, the ball size, and the ball rotation speed. Under optimal conditions, the new method could achieve a complete degradation, debromination and mineralization of BDE209 within 3h of milling. However, the degradation removal (or debromination efficiency) was decreased to only 51.7% (15.6%) and 67.8% (31.5%) for the use of CaO and peroxymonosulfate, respectively. The analyses of products demonstrated that once the degradation was initiated, BDE209 molecules were deeply debrominated and fully mineralized in the MC-PS system. The strong oxidizing ability of this system was due to the reactive sulfate radicals generated from the MC-enhanced activation of PS, which was confirmed with electron spin resonance spectroscopy. Because no toxic low brominated polybrominated diphenyl ethers were accumulated as byproducts, the proposed MC oxidative degradation method will have promising applications in the treatment of solid BDE209 at high concentrations. PMID:26474378

  18. The wheat powdery mildew genome shows the unique evolution of an obligate biotroph.

    PubMed

    Wicker, Thomas; Oberhaensli, Simone; Parlange, Francis; Buchmann, Jan P; Shatalina, Margarita; Roffler, Stefan; Ben-David, Roi; Doležel, Jaroslav; Šimková, Hana; Schulze-Lefert, Paul; Spanu, Pietro D; Bruggmann, Rémy; Amselem, Joelle; Quesneville, Hadi; Ver Loren van Themaat, Emiel; Paape, Timothy; Shimizu, Kentaro K; Keller, Beat

    2013-09-01

    Wheat powdery mildew, Blumeria graminis forma specialis tritici, is a devastating fungal pathogen with a poorly understood evolutionary history. Here we report the draft genome sequence of wheat powdery mildew, the resequencing of three additional isolates from different geographic regions and comparative analyses with the barley powdery mildew genome. Our comparative genomic analyses identified 602 candidate effector genes, with many showing evidence of positive selection. We characterize patterns of genetic diversity and suggest that mildew genomes are mosaics of ancient haplogroups that existed before wheat domestication. The patterns of diversity in modern isolates suggest that there was no pronounced loss of genetic diversity upon formation of the new host bread wheat 10,000 years ago. We conclude that the ready adaptation of B. graminis f.sp. tritici to the new host species was based on a diverse haplotype pool that provided great genetic potential for pathogen variation. PMID:23852167

  19. NMR Spectroscopy Identifies Metabolites Translocated from Powdery Mildew Resistant Rootstocks to Susceptible Watermelon Scions.

    PubMed

    Mahmud, Iqbal; Kousik, Chandrasekar; Hassell, Richard; Chowdhury, Kamal; Boroujerdi, Arezue F

    2015-09-16

    Powdery mildew (PM) disease causes significant loss in watermelon. Due to the unavailability of a commercial watermelon variety that is resistant to PM, grafting susceptible cultivars on wild resistant rootstocks is being explored as a short-term management strategy to combat this disease. Nuclear magnetic resonance-based metabolic profiles of susceptible and resistant rootstocks of watermelon and their corresponding susceptible scions (Mickey Lee) were compared to screen for potential metabolites related to PM resistance using multivariate principal component analysis. Significant score plot differences between the susceptible and resistant groups were revealed through Mahalanobis distance analysis. Significantly different spectral buckets and their corresponding metabolites (including choline, fumarate, 5-hydroxyindole-3-acetate, and melatonin) have been identified quantitatively using multivariate loading plots and verified by volcano plot analyses. The data suggest that these metabolites were translocated from the powdery mildew resistant rootstocks to their corresponding powdery mildew susceptible scions and can be related to PM disease resistance. PMID:26302171

  20. Nuclear Function of Subclass I Actin-Depolymerizing Factor Contributes to Susceptibility in Arabidopsis to an Adapted Powdery Mildew Fungus1[OPEN

    PubMed Central

    Inada, Noriko; Higaki, Takumi; Hasezawa, Seiichiro

    2016-01-01

    Actin-depolymerizing factors (ADFs) are conserved proteins that function in regulating the structure and dynamics of actin microfilaments in eukaryotes. In this study, we present evidence that Arabidopsis (Arabidopsis thaliana) subclass I ADFs, particularly ADF4, functions as a susceptibility factor for an adapted powdery mildew fungus. The null mutant of ADF4 significantly increased resistance against the adapted powdery mildew fungus Golovinomyces orontii. The degree of resistance was further enhanced in transgenic plants in which the expression of all subclass I ADFs (i.e. ADF1–ADF4) was suppressed. Microscopic observations revealed that the enhanced resistance of adf4 and ADF1-4 knockdown plants (ADF1-4Ri) was associated with the accumulation of hydrogen peroxide and cell death specific to G. orontii-infected cells. The increased resistance and accumulation of hydrogen peroxide in ADF1-4Ri were suppressed by the introduction of mutations in the salicylic acid- and jasmonic acid-signaling pathways but not by a mutation in the ethylene-signaling pathway. Quantification by microscopic images detected an increase in the level of actin microfilament bundling in ADF1-4Ri but not in adf4 at early G. orontii infection time points. Interestingly, complementation analysis revealed that nuclear localization of ADF4 was crucial for susceptibility to G. orontii. Based on its G. orontii-infected-cell-specific phenotype, we suggest that subclass I ADFs are susceptibility factors that function in a direct interaction between the host plant and the powdery mildew fungus. PMID:26747284

  1. Genome-Wide Study of the Tomato SlMLO Gene Family and Its Functional Characterization in Response to the Powdery Mildew Fungus Oidium neolycopersici.

    PubMed

    Zheng, Zheng; Appiano, Michela; Pavan, Stefano; Bracuto, Valentina; Ricciardi, Luigi; Visser, Richard G F; Wolters, Anne-Marie A; Bai, Yuling

    2016-01-01

    The MLO (Mildew Locus O) gene family encodes plant-specific proteins containing seven transmembrane domains and likely acting in signal transduction in a calcium and calmodulin dependent manner. Some members of the MLO family are susceptibility factors toward fungi causing the powdery mildew disease. In tomato, for example, the loss-of-function of the MLO gene SlMLO1 leads to a particular form of powdery mildew resistance, called ol-2, which arrests almost completely fungal penetration. This type of penetration resistance is characterized by the apposition of papillae at the sites of plant-pathogen interaction. Other MLO homologs in Arabidopsis regulate root response to mechanical stimuli (AtMLO4 and AtMLO11) and pollen tube reception by the female gametophyte (AtMLO7). However, the role of most MLO genes remains unknown. In this work, we provide a genome-wide study of the tomato SlMLO gene family. Besides SlMLO1, other 15 SlMLO homologs were identified and characterized with respect to their structure, genomic organization, phylogenetic relationship, and expression profile. In addition, by analysis of transgenic plants, we demonstrated that simultaneous silencing of SlMLO1 and two of its closely related homologs, SlMLO5 and SlMLO8, confer higher level of resistance than the one associated with the ol-2 mutation. The outcome of this study provides evidence for functional redundancy among tomato homolog genes involved in powdery mildew susceptibility. Moreover, we developed a series of transgenic lines silenced for individual SlMLO homologs, which lay the foundation for further investigations aimed at assigning new biological functions to the MLO gene family. PMID:27579028

  2. Genome-Wide Study of the Tomato SlMLO Gene Family and Its Functional Characterization in Response to the Powdery Mildew Fungus Oidium neolycopersici

    PubMed Central

    Zheng, Zheng; Appiano, Michela; Pavan, Stefano; Bracuto, Valentina; Ricciardi, Luigi; Visser, Richard G. F.; Wolters, Anne-Marie A.; Bai, Yuling

    2016-01-01

    The MLO (Mildew Locus O) gene family encodes plant-specific proteins containing seven transmembrane domains and likely acting in signal transduction in a calcium and calmodulin dependent manner. Some members of the MLO family are susceptibility factors toward fungi causing the powdery mildew disease. In tomato, for example, the loss-of-function of the MLO gene SlMLO1 leads to a particular form of powdery mildew resistance, called ol-2, which arrests almost completely fungal penetration. This type of penetration resistance is characterized by the apposition of papillae at the sites of plant-pathogen interaction. Other MLO homologs in Arabidopsis regulate root response to mechanical stimuli (AtMLO4 and AtMLO11) and pollen tube reception by the female gametophyte (AtMLO7). However, the role of most MLO genes remains unknown. In this work, we provide a genome-wide study of the tomato SlMLO gene family. Besides SlMLO1, other 15 SlMLO homologs were identified and characterized with respect to their structure, genomic organization, phylogenetic relationship, and expression profile. In addition, by analysis of transgenic plants, we demonstrated that simultaneous silencing of SlMLO1 and two of its closely related homologs, SlMLO5 and SlMLO8, confer higher level of resistance than the one associated with the ol-2 mutation. The outcome of this study provides evidence for functional redundancy among tomato homolog genes involved in powdery mildew susceptibility. Moreover, we developed a series of transgenic lines silenced for individual SlMLO homologs, which lay the foundation for further investigations aimed at assigning new biological functions to the MLO gene family. PMID:27579028

  3. A variable homopolymeric G-repeat defines small RNA-mediated posttranscriptional regulation of a chemotaxis receptor in Helicobacter pylori

    PubMed Central

    Pernitzsch, Sandy R.; Tirier, Stephan M.; Beier, Dagmar; Sharma, Cynthia M.

    2014-01-01

    Phase variation of hypermutable simple sequence repeats (SSRs) is a widespread and stochastic mechanism to generate phenotypic variation within a population and thereby contributes to host adaptation of bacterial pathogens. Although several examples of SSRs that affect transcription or coding potential have been reported, we now show that a SSR also impacts small RNA-mediated posttranscriptional regulation. Based on in vitro and in vivo analyses, we demonstrate that a variable homopolymeric G-repeat in the leader of the TlpB chemotaxis receptor mRNA of the human pathogen Helicobacter pylori is directly targeted by a small RNA (sRNA), RepG (Regulator of polymeric G-repeats). Whereas RepG sRNA is highly conserved, the tlpB G-repeat length varies among diverse H. pylori strains, resulting in strain-specific RepG-mediated tlpB regulation. Based on modification of the G-repeat length within one strain, we demonstrate that the G-repeat length determines posttranscriptional regulation and can mediate both repression and activation of tlpB through RepG. In vitro translation assays show that this regulation occurs at the translational level and that RepG influences tlpB translation dependent on the G-repeat length. In contrast to the digital ON–OFF switches through frame-shift mutations within coding sequences, such modulation of posttranscriptional regulation allows for a gradual control of gene expression. This connection to sRNA-mediated posttranscriptional regulation might also apply to other genes with SSRs, which could be targeting sites of cis- or trans-encoded sRNAs, and thereby could facilitate host adaptation through sRNA-mediated fine-tuning of virulence gene expression. PMID:24474799

  4. The Powdery Mildew Disease of Arabidopsis: A Paradigm for the Interaction between Plants and Biotrophic Fungi

    PubMed Central

    Micali, Cristina; Göllner, Katharina; Humphry, Matt; Consonni, Chiara; Panstruga, Ralph

    2008-01-01

    The powdery mildew diseases, caused by fungal species of the Erysiphales, have an important economic impact on a variety of plant species and have driven basic and applied research efforts in the field of phytopathology for many years. Although the first taxonomic reports on the Erysiphales date back to the 1850's, advances into the molecular biology of these fungal species have been hampered by their obligate biotrophic nature and difficulties associated with their cultivation and genetic manipulation in the laboratory. The discovery in the 1990's of a few species of powdery mildew fungi that cause disease on Arabidopsis has opened a new chapter in this research field. The great advantages of working with a model plant species have translated into remarkable progress in our understanding of these complex pathogens and their interaction with the plant host. Herein we summarize advances in the study of Arabidopsis-powdery mildew interactions and discuss their implications for the general field of plant pathology. We provide an overview of the life cycle of the pathogens on Arabidopsis and describe the structural and functional changes that occur during infection in the host and fungus in compatible and incompatible interactions, with special emphasis on defense signaling, resistance pathways, and compatibility factors. Finally, we discuss the future of powdery mildew research in anticipation of the sequencing of multiple powdery mildew genomes. The cumulative body of knowledge on powdery mildews of Arabidopsis provides a valuable tool for the study and understanding of disease associated with many other obligate biotrophic pathogen species. PMID:22303240

  5. Two Pathways Mediate Inter-Domain Allosteric Regulation in Pin1

    PubMed Central

    Guo, Jingjing; Pang, Xiaodong; Zhou, Huan-Xiang

    2014-01-01

    Summary Allostery is an essential means for regulating biomolecular functions and provides unique opportunities for drug design, yet our ability to elucidate allosteric mechanisms remains limited. Here, based on extensive molecular dynamics simulations, we present an atomistic picture of the pathways mediating the allosteric regulation of the PPIase domain of Pin1 by its WW domain. Two pathways jointly propagate the action of substrate-WW binding to produce closure and rigidification of three PPIase catalytic-site loops. One pathway preexists in the apo protein but remains dormant until substrate-WW binding completes the second. The reduction in conformational entropy and preorganization of the catalytic-site loops observed here may explain why substrate-WW binding enhances ligand affinity and catalytic activity of the PPIase domain, and suggest a combination drug therapy for Pin1-related diseases. Whereas the traditional view of allostery has emphasized conformational transition, our study uniquely identifies a distinct role of conformational dynamics in eliciting allostery. PMID:25543254

  6. Diacylglycerol mediates regulation of TASK potassium channels by Gq-coupled receptors.

    PubMed

    Wilke, Bettina U; Lindner, Moritz; Greifenberg, Lea; Albus, Alexandra; Kronimus, Yannick; Bünemann, Moritz; Leitner, Michael G; Oliver, Dominik

    2014-01-01

    The two-pore domain potassium (K2P) channels TASK-1 (KCNK3) and TASK-3 (KCNK9) are important determinants of background K(+) conductance and membrane potential. TASK-1/3 activity is regulated by hormones and transmitters that act through G protein-coupled receptors (GPCR) signalling via G proteins of the Gαq/11 subclass. How the receptors inhibit channel activity has remained unclear. Here, we show that TASK-1 and -3 channels are gated by diacylglycerol (DAG). Receptor-initiated inhibition of TASK required the activity of phospholipase C, but neither depletion of the PLC substrate PI(4,5)P2 nor release of the downstream messengers IP3 and Ca(2+). Attenuation of cellular DAG transients by DAG kinase or lipase suppressed receptor-dependent inhibition, showing that the increase in cellular DAG-but not in downstream lipid metabolites-mediates channel inhibition. The findings identify DAG as the signal regulating TASK channels downstream of GPCRs and define a novel role for DAG that directly links cellular DAG dynamics to excitability. PMID:25420509

  7. DELLA-mediated gibberellin signalling regulates Nod factor signalling and rhizobial infection.

    PubMed

    Fonouni-Farde, Camille; Tan, Sovanna; Baudin, Maël; Brault, Mathias; Wen, Jiangqi; Mysore, Kirankumar S; Niebel, Andreas; Frugier, Florian; Diet, Anouck

    2016-01-01

    Legumes develop symbiotic interactions with rhizobial bacteria to form nitrogen-fixing nodules. Bacterial Nod factors (NFs) and plant regulatory pathways modulating NF signalling control rhizobial infections and nodulation efficiency. Here we show that gibberellin (GA) signalling mediated by DELLA proteins inhibits rhizobial infections and controls the NF induction of the infection marker ENOD11 in Medicago truncatula. Ectopic expression of a constitutively active DELLA protein in the epidermis is sufficient to promote ENOD11 expression in the absence of symbiotic signals. We show using heterologous systems that DELLA proteins can interact with the nodulation signalling pathway 2 (NSP2) and nuclear factor-YA1 (NF-YA1) transcription factors that are essential for the activation of NF responses. Furthermore, MtDELLA1 can bind the ERN1 (ERF required for nodulation 1) promoter and positively transactivate its expression. Overall, we propose that GA-dependent action of DELLA proteins may directly regulate the NSP1/NSP2 and NF-YA1 activation of ERN1 transcription to regulate rhizobial infections. PMID:27586842

  8. USP15 regulates SMURF2 kinetics through C-lobe mediated deubiquitination

    PubMed Central

    Iyengar, Prasanna Vasudevan; Jaynes, Patrick; Rodon, Laura; Lama, Dilraj; Law, Kai Pong; Lim, Yoon Pin; Verma, Chandra; Seoane, Joan; Eichhorn, Pieter Johan Adam

    2015-01-01

    Ubiquitin modification of the TGF-β pathway components is emerging as a key mechanism of TGF-β pathway regulation. To limit TGF-β responses, TGF-β signaling is regulated through a negative feedback loop whereby the E3 ligase SMURF2 targets the TGF-β receptor (TβR) complex for ubiquitin-mediated degradation. Counteracting this process, a number of deubiquitinating (DUBs) enzymes have recently been identified that deubiquitinate and stabilize the TβR. However the precise mechanism by which these DUBs act on TβR function remains poorly defined. Here, we demonstrate that apart from targeting the TβR complex directly, USP15 also deubiquitinates SMURF2 resulting in enhanced TβR stability and downstream pathway activation. Through proteomic analysis, we show that USP15 modulates the ubiquitination of Lys734, a residue required for SMURF2 catalytic activity. Our results show that SMURF2 is a critical target of USP15 in the TGF-β pathway and may also explain how USP15 and SMURF2 target multiple complementary protein complexes in other pathways. PMID:26435193

  9. Phospholipase D-mediated autophagic regulation is a potential target for cancer therapy

    PubMed Central

    Jang, Y H; Choi, K Y; Min, D S

    2014-01-01

    Autophagy is a catabolic process in which cell components are degraded to maintain cellular homeostasis by nutrient limitations. Defects of autophagy are involved in numerous diseases, including cancer. Here, we demonstrate a new role of phospholipase D (PLD) as a regulator of autophagy. PLD inhibition enhances autophagic flux via ATG1 (ULK1), ATG5 and ATG7, which are essential autophagy gene products critical for autophagosome formation. Moreover, PLD suppresses autophagy by differentially modulating phosphorylation of ULK1 mediated by mTOR and adenosine monophosphate-activated protein kinase (AMPK), and by suppressing the interaction of Beclin 1 with vacuolar-sorting protein 34 (Vps34), indicating that PLD coordinates major players of the autophagic pathway, AMPK-mTOR-ULK1 and Vps34/Beclin 1. Ultimately, PLD inhibition significantly sensitized in vitro and in vivo cancer regression via genetic and pharmacological inhibition of autophagy, providing rationale for a new therapeutic approach to enhancing the anticancer efficacy of PLD inhibition. Collectively, we show a novel role for PLD in the molecular machinery regulating autophagy. PMID:24317201

  10. Role of a COP1 interactive protein in mediating light-regulated gene expression in arabidopsis.

    PubMed Central

    Yamamoto, Y Y; Matsui, M; Ang, L H; Deng, X W

    1998-01-01

    Arabidopsis seedlings display distinct patterns of gene expression and morphogenesis according to the ambient light condition. An Arabidopsis nuclear protein, CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1), acts to repress photomorphogenesis in the absence of light. The Arabidopsis CIP7 protein was identified by its capability to interact with COP1. CIP7 is a novel nuclear protein that contains transcriptional activation activity without a recognizable DNA binding motif. CIP7 requires light for its high level of expression, and COP1 seems to play a role in repressing its expression in darkness. Decreasing CIP7 expression by introducing antisense CIP7 RNA resulted in defects in light-dependent anthocyanin and chlorophyll accumulation. Antisense plants also displayed reduced expression of light-inducible genes for anthocyanin biosynthesis and photosynthesis. However, no defect was observed in light-dependent inhibition of hypocotyl elongation. Taken together, our data indicate that CIP7 acts as a positive regulator of light-regulated genes and is a potential direct downstream target of COP1 for mediating light control of gene expression. PMID:9668129

  11. Role of cocaine- and amphetamine-regulated transcript in estradiol-mediated neuroprotection

    NASA Astrophysics Data System (ADS)

    Xu, Yun; Zhang, Wenri; Klaus, Judith; Young, Jennifer; Koerner, Ines; Sheldahl, Laird C.; Hurn, Patricia D.; Martínez-Murillo, Francisco; Alkayed, Nabil J.

    2006-09-01

    Estrogen reduces brain injury after experimental cerebral ischemia in part through a genomic mechanism of action. Using DNA microarrays, we analyzed the genomic response of the brain to estradiol, and we identified a transcript, cocaine- and amphetamine-regulated transcript (CART), that is highly induced in the cerebral cortex by estradiol under ischemic conditions. Using in vitro and in vivo models of neural injury, we confirmed and characterized CART mRNA and protein up-regulation by estradiol in surviving neurons, and we demonstrated that i.v. administration of a rat CART peptide is protective against ischemic brain injury in vivo. We further demonstrated binding of cAMP response element (CRE)-binding protein to a CART promoter CRE site in ischemic brain and rapid activation by CART of ERK in primary cultured cortical neurons. The findings suggest that CART is an important player in estrogen-mediated neuroprotection and a potential therapeutic agent for stroke and other neurodegenerative diseases. ischemia | stroke | estrogen

  12. Anks1a regulates COPII-mediated anterograde transport of receptor tyrosine kinases critical for tumorigenesis.

    PubMed

    Lee, Haeryung; Noh, Hyuna; Mun, Jiyoung; Gu, Changkyu; Sever, Sanja; Park, Soochul

    2016-01-01

    ErbB2 signalling, which is amplified by EphA2 binding, is an important therapeutic target for breast cancer. Despite the importance of the EphA2/ErbB2 complex in promoting breast tumorigenesis, the mechanism by which these receptor tyrosine kinases (RTKs) are exported from the endoplasmic reticulum (ER) remains poorly understood. Here we report that the PTB adaptor Anks1a is specifically localized to the ER on its own serine phosphorylation. Once there, Anks1a acts as an important regulator of COPII-mediated EphA2 ER export. The Anks1a ankyrin repeat domain binds EphA2 and causes it to accumulate at sites of ER exit. Simultaneously, the Anks1a PTB domain binds Sec23. This induces internalization of EphA2 via COPII vesicles, while Anks1a remains behind on the ER membrane. EphA2 also binds ErbB2 in the ER and seems to load ErbB2 into growing COPII carriers. Together, our study reveals a novel mechanism that regulates the loading of RTKs into COPII vesicles. PMID:27619642

  13. ERβ regulation of NF-kB activation in prostate cancer is mediated by HIF-1.

    PubMed

    Mak, Paul; Li, Jiarong; Samanta, Sanjoy; Mercurio, Arthur M

    2015-11-24

    We examined the regulation of NF-κB in prostate cancer by estrogen receptor β (ERβ) based on the inverse correlation between p65 and ERβ expression that exists in prostate carcinomas and reports that ERβ can inhibit NF-κB activation, although the mechanism is not known. We demonstrate that ERβ functions as a gate-keeper for NF-κB p65 signaling by repressing its expression and nuclear translocation. ERβ regulation of NF-κB signaling is mediated by HIF-1. Loss of ERβ or hypoxia stabilizes HIF-1α, which we found to be a direct driver of IKKβ transcription through a hypoxia response element present in the promoter of the IKKβ gene. The increase of IKKβ expression in ERβ-ablated cells correlates with an increase in phospho-IκBα and concomitant p65 nuclear translocation. An inverse correlation between the expression of ERβ and IKKβ/p65 was also observed in the prostates of ERβ knockout (BERKO) mice, Gleason grade 5 prostate tumors and analysis of prostate cancer databases. These findings provide a novel mechanism for how ERβ prevents NF-κB activation and raise the exciting possibility that loss of ERβ expression is linked to chronic inflammation in the prostate, which contributes to the development of high-grade prostate cancer. PMID:26450901

  14. Phosphorylation and SCF-mediated degradation regulate CREB-H transcription of metabolic targets

    PubMed Central

    Barbosa, Sónia; Carreira, Suzanne; Bailey, Daniel; Abaitua, Fernando; O'Hare, Peter

    2015-01-01

    CREB‑H, an endoplasmic reticulum–anchored transcription factor, plays a key role in regulating secretion and in metabolic and inflammatory pathways, but how its activity is modulated remains unclear. We examined processing of the nuclear active form and identified a motif around S87–S90 with homology to DSG-type phosphodegrons. We show that this region is subject to multiple phosphorylations, which regulate CREB-H stability by targeting it to the SCFFbw1a E3 ubiquitin ligase. Data from phosphatase treatment, use of phosophospecific antibody, and substitution of serine residues demonstrate phosphorylation of candidate serines in the region, with the core S87/S90 motif representing a critical determinant promoting proteasome-mediated degradation. Candidate kinases CKII and GSK-3b phosphorylate CREB-H in vitro with specificities for different serines. Prior phosphorylation with GSK-3 at one or more of the adjacent serines substantially increases S87/S90-dependent phosphorylation by CKII. In vivo expression of a dominant-negative Cul1 enhances steady-state levels of CREB‑H, an effect augmented by Fbw1a. CREB-H directly interacts with Fbw1a in a phosphorylation-dependent manner. Finally, mutations within the phosphodegron, when incorporated into the full-length protein, result in increased levels of constitutively cleaved nuclear protein and increased transcription and secretion of a key endogenous target gene, apolipoprotein A IV. PMID:26108621

  15. Cell mediated contraction in 3D cell-matrix constructs leads to spatially regulated osteogenic differentiation

    PubMed Central

    Klumpers, Darinka D.; Zhao, Xuanhe; Mooney, David J.; Smit, Theo H.

    2013-01-01

    During embryonic development, morphogenetic processes give rise to a variety of shapes and patterns that lead to functional tissues and organs. While the impact of chemical signals in these processes is widely studied, the role of physical cues is less understood. The aim of this study was to test the hypothesis that the interplay of cell mediated contraction and mechanical boundary conditions alone can result in spatially regulated differentiation in simple 3D constructs. An experimental model consisting of a 3D cell-gel construct and a finite element (FE) model were used to study the effect of cellular traction exerted by mesenchymal stem cells (MSCs) on an initially homogeneous matrix under inhomogeneous boundary conditions. A robust shape change is observed due to contraction under time-varying mechanical boundary conditions, which is explained by the finite element model. Furthermore, distinct local differences of osteogenic differentiation are observed, with a spatial pattern independent of osteogenic factors in the culture medium. Regions that are predicted to have experienced relatively high shear stress at any time during contraction, correlate with the regions of distinct osteogenesis. Taken together, these results support the underlying hypothesis that cellular contractility and mechanical boundary conditions alone can result in spatially regulated differentiation. These results will have important implications for tissue engineering and regeneration. PMID:23925497

  16. GABA-mediated regulation of the activity-dependent olfactory bulb dopaminergic phenotype

    PubMed Central

    Akiba, Yosuke; Sasaki, Hayato; Huerta, Patricio T.; Estevez, Alvaro G.; Baker, Harriet; Cave, John W.

    2009-01-01

    Gamma-amino-butyric acid (GABA) regulates the proliferation and migration of olfactory bulb (OB) interneuron progenitors derived from the subventricular zone (SVZ), but the role of GABA in the differentiation of these progenitors has been largely unexplored. This study examined the role of GABA in the differentiation of OB dopaminergic interneurons using neonatal forebrain organotypic slice cultures prepared from transgenic mice expressing GFP under the control of the tyrosine hydroxylase (Th) gene promoter (ThGFP). KCl-mediated depolarization of the slices induced ThGFP expression. The addition of GABA to the depolarized slices further increased GFP fluorescence by inducing ThGFP expression in an additional set of periglomerular cells. These findings showed that GABA promoted differentiation of SVZ-derived OB dopaminergic interneurons and suggested that GABA indirectly regulated Th expression and OB dopaminergic neuron differentiation through an acceleration of the maturation rate for the dopaminergic progenitors. Additional studies revealed that the effect of GABA on ThGFP expression required activation of L- and P/Q-type Ca+2 channels as well as GABAA and GABAB receptors. These voltage-gated Ca+2 channels and GABA receptors have previously been shown to be required for the co-expressed GABAergic phenotype in the OB interneurons. Together, these findings suggest that Th expression and the differentiation of OB dopaminergic interneurons are coupled to the co-expressed GABAergic phenotype, and demonstrate a novel role for GABA in neurogenesis. PMID:19301430

  17. SP PROTEINS AND RUNX2 MEDIATE REGULATION OF MATRIX GLA PROTEIN (MGP) EXPRESSION BY PARATHYROID HORMONE

    PubMed Central

    Suttamanatwong, Supaporn; Jensen, Eric D; Shilling, Jody; Franceschi, Renny T.; Carlson, Ann E.; Mansky, Kim C.; Gopalakrishnan, Rajaram

    2009-01-01

    As part of its catabolic action in bone, parathyroid hormone (PTH) inhibits extracellular matrix mineralization. We previously showed that PTH dose-dependently induces matrix gla protein (MGP) expression in osteoblasts and this induction is at least partially responsible for PTH-mediated inhibition of mineralization. Recently, we identified PKA and ERK/MAPK as the key signaling pathways involved in PTH regulation of MGP expression. The goal of this study was to further characterize the mechanism by which PTH stimulates expression of MGP. Deletion analysis of the murine Mgp gene promoter identified a PTH-responsive region between -173 bp and -49 bp. Using gel-mobility shift assays we found that Sp1, Sp3 and Runx2 bind to distinct sites within this region. Mutation of either the Sp or the Runx2 site reduced MGP induction by PTH, while mutation of both sites completely abolished PTH responsiveness. Overexpression of Runx2 or Sp1 activated the Mgp reporter, while Sp3 was a dose-dependent repressor of MGP expression. Collectively, these data show that PTH regulates MGP gene transcription in osteoblasts through altered activities of Sp and Runx2 transcription factors. PMID:19306294

  18. CO-CBS-H2 S Axis: From Vascular Mediator to Cancer Regulator.

    PubMed

    Suematsu, Makoto; Nakamura, Takashi; Tokumoto, Yasuhito; Yamamoto, Takehiro; Kajimura, Mayumi; Kabe, Yasuaki

    2016-04-01

    CO is a gaseous mediator generated by HO. Our previous studies revealed that CO generated from inducible HO-1 or from constitutive HO-2 modulates function of different heme proteins or enzymes through binding to their prosthetic ferrous heme to alter their structures, regulating biological function of cells and organs. Such CO-directed target macromolecules include sGC and CBS. In the liver, CO serves as a sinusoidal dilator through its action on sGC in hepatic stellate cells, while the same gas accounts for vasoconstrictor that inhibits H2 S generated by CO-sensitive CBS in astrocytes. Since molecular O2 is a substrate for HO, the latter mechanism contributes to hypoxic vasodilation in neurovascular units. We have recently uncovered that stress-inducible CO in and around cancer cells suppresses CBS to result in decreased methylation of PFKFB3, the enzyme regulating PFK-1, leading to a shift of glucose biotransformation from glycolysis toward pentose phosphate pathway; such a metabolic remodeling causes chemoresistance through increasing NADPH and reduced glutathione under stress conditions for cancer cells. This article reviews the intriguing networks of CO-sensitive metabolic regulatory mechanisms in microcirculation and cancer. PMID:26537442

  19. Neutrophils Self-Regulate Immune Complex-Mediated Cutaneous Inflammation through CXCL2.

    PubMed

    Li, Jackson LiangYao; Lim, Chun Hwee; Tay, Fen Wei; Goh, Chi Ching; Devi, Sapna; Malleret, Benoit; Lee, Bernett; Bakocevic, Nadja; Chong, Shu Zhen; Evrard, Maximilien; Tanizaki, Hideaki; Lim, Hwee Ying; Russell, Bruce; Renia, Laurent; Zolezzi, Francesca; Poidinger, Michael; Angeli, Veronique; St John, Ashley L; Harris, John E; Tey, Hong Liang; Tan, Suet Mien; Kabashima, Kenji; Weninger, Wolfgang; Larbi, Anis; Ng, Lai Guan

    2016-02-01

    Deposition of immune complexes (ICs) in tissues triggers acute inflammatory pathology characterized by massive neutrophil influx leading to edema and hemorrhage, and is especially associated with vasculitis of the skin, but the mechanisms that regulate this type III hypersensitivity process remain poorly understood. Here, using a combination of multiphoton intravital microscopy and genomic approaches, we re-examined the cutaneous reverse passive Arthus reaction and observed that IC-activated neutrophils underwent transmigration, triggered further IC formation, and transported these ICs into the interstitium, whereas neutrophil depletion drastically reduced IC formation and ameliorated vascular leakage in vivo. Thereafter, we show that these neutrophils expressed high levels of CXCL2, which further amplified neutrophil recruitment and activation in an autocrine and/or paracrine manner. Notably, CXCL1 expression was restricted to tissue-resident cell types, but IC-activated neutrophils may also indirectly, via soluble factors, modulate macrophage CXCL1 expression. Consistent with their distinct cellular origins and localization, only neutralization of CXCL2 but not CXCL1 in the interstitium effectively reduced neutrophil recruitment. In summary, our study establishes that neutrophils are able to self-regulate their own recruitment and responses during IC-mediated inflammation through a CXCL2-driven feed forward loop. PMID:26802238

  20. GAD67-mediated GABA Synthesis and Signaling Regulate Inhibitory Synaptic Innervation in the Visual Cortex

    PubMed Central

    Chattopadhyaya, Bidisha; Di Cristo, Graziella; Wu, Cai Zhi; Knott, Graham; Kuhlman, Sandra; Fu, Yu; Palmiter, Richard D.; Huang, Z. Josh

    2007-01-01

    The development of GABAergic inhibitory circuits is shaped by neural activity, but the underlying mechanisms are unclear. we demonstrate a novel function of GABA in regulating GABAergic innervation in the adolescent brain, when GABA is mainly known as an inhibitory transmitter. Conditional knockdown of the rate-limiting synthetic enzyme GAD67 in basket interneurons in adolescent visual cortex resulted in cell autonomous deficits in axon branching, perisomatic synapse formation around pyramidal neurons, and complexity of the innervation fields; the same manipulation had little influence on the subsequent maintenance of perisomatic synapses. These effects of GABA deficiency were rescued by suppressing GABA re-uptake and by GABA receptor agonists. Germ-line knockdown of GAD67 but not GAD65 showed similar deficits, suggesting a specific role of GAD67 in the maturation of perisomatic innervation. Since intracellular GABA levels are modulated by neuronal activity, our results implicate GAD67-mediated GABA synthesis in activity-dependent regulation of inhibitory innervation patterns. PMID:17582330

  1. Endoglin Regulation of Smad2 Function Mediates Beclin1 Expression and Endothelial Autophagy*

    PubMed Central

    Pan, Christopher C.; Kumar, Sanjay; Shah, Nirav; Bloodworth, Jeffrey C.; Hawinkels, Lukas J. A. C.; Mythreye, Karthikeyan; Hoyt, Dale G.; Lee, Nam Y.

    2015-01-01

    Autophagy is the targeted degradation of proteins and organelles critical for homeostasis and cell survival. Transforming growth factor β (TGF-β) differentially regulates autophagy in a context-specific manner, although the precise intracellular mechanisms remain less clear. Importantly, how TGF-β controls autophagic responses in endothelial cells (EC) during angiogenesis is unknown. Here we identified endoglin, an EC-specific TGF-β co-receptor essential for angiogenesis, as a key determinant of autophagy. Among the two opposing TGF-β Smad pathways in the EC system (Smad1/5/8 and Smad2/3), we found Smad2 as the major transcriptional regulator of autophagy that targets beclin1 (BECN1) gene expression. Smad2, but not Smad3, acts as a repressor upstream of the BECN1 promoter region. Overall, endoglin promotes autophagy by impeding Smad2 transcriptional repressor activity. Notably, increased beclin1 levels upon Smad2 knockdown directly correlated with enhanced autophagy during angiogenesis. Taken together, these results establish endoglin as a critical mediator of autophagy and demonstrate a new transcriptional mechanism by which Smad2 inhibits angiogenesis. PMID:25931117

  2. MCPIP1 Endoribonuclease Activity Negatively Regulates Interleukin-17-Mediated Signaling and Inflammation.

    PubMed

    Garg, Abhishek V; Amatya, Nilesh; Chen, Kong; Cruz, J Agustin; Grover, Prerna; Whibley, Natasha; Conti, Heather R; Hernandez Mir, Gerard; Sirakova, Tatiana; Childs, Erin C; Smithgall, Thomas E; Biswas, Partha S; Kolls, Jay K; McGeachy, Mandy J; Kolattukudy, Pappachan E; Gaffen, Sarah L

    2015-09-15

    Interleukin-17 (IL-17) induces pathology in autoimmunity and infections; therefore, constraint of this pathway is an essential component of its regulation. We demonstrate that the signaling intermediate MCPIP1 (also termed Regnase-1, encoded by Zc3h12a) is a feedback inhibitor of IL-17 receptor signal transduction. MCPIP1 knockdown enhanced IL-17-mediated signaling, requiring MCPIP1's endoribonuclease but not deubiquitinase domain. MCPIP1 haploinsufficient mice showed enhanced resistance to disseminated Candida albicans infection, which was reversed in an Il17ra(-/-) background. Conversely, IL-17-dependent pathology in Zc3h12a(+/-) mice was exacerbated in both EAE and pulmonary inflammation. MCPIP1 degraded Il6 mRNA directly but only modestly downregulated the IL-6 promoter. However, MCPIP1 strongly inhibited the Lcn2 promoter by regulating the mRNA stability of Nfkbiz, encoding the IκBζ transcription factor. Unexpectedly, MCPIP1 degraded Il17ra and Il17rc mRNA, independently of the 3' UTR. The cumulative impact of MCPIP1 on IL-6, IκBζ, and possibly IL-17R subunits results in a biologically relevant inhibition of IL-17 signaling. PMID:26320658

  3. PPARγ isoforms differentially regulate metabolic networks to mediate mouse prostatic epithelial differentiation

    PubMed Central

    Strand, D W; Jiang, M; Murphy, T A; Yi, Y; Konvinse, K C; Franco, O E; Wang, Y; Young, J D; Hayward, S W

    2012-01-01

    Recent observations indicate prostatic diseases are comorbidities of systemic metabolic dysfunction. These discoveries revealed fundamental questions regarding the nature of prostate metabolism. We previously showed that prostate-specific ablation of PPARγ in mice resulted in tumorigenesis and active autophagy. Here, we demonstrate control of overlapping and distinct aspects of prostate epithelial metabolism by ectopic expression of individual PPARγ isoforms in PPARγ knockout prostate epithelial cells. Expression and activation of either PPARγ 1 or 2 reduced de novo lipogenesis and oxidative stress and mediated a switch from glucose to fatty acid oxidation through regulation of genes including Pdk4, Fabp4, Lpl, Acot1 and Cd36. Differential effects of PPARγ isoforms included decreased basal cell differentiation, Scd1 expression and triglyceride fatty acid desaturation and increased tumorigenicity by PPARγ1. In contrast, PPARγ2 expression significantly increased basal cell differentiation, Scd1 expression and AR expression and responsiveness. Finally, in confirmation of in vitro data, a PPARγ agonist versus high-fat diet (HFD) regimen in vivo confirmed that PPARγ agonization increased prostatic differentiation markers, whereas HFD downregulated PPARγ-regulated genes and decreased prostate differentiation. These data provide a rationale for pursuing a fundamental metabolic understanding of changes to glucose and fatty acid metabolism in benign and malignant prostatic diseases associated with systemic metabolic stress. PMID:22874998

  4. Hypoxia-Inducible Factor 1: Regulator of Mitochondrial Metabolism and Mediator of Ischemic Preconditioning

    PubMed Central

    Semenza, Gregg L.

    2010-01-01

    Hypoxia-inducible factor 1 (HIF-1) mediates adaptive responses to reduced oxygen availability by regulating gene expression. A critical cell-autonomous adaptive response to chronic hypoxia controlled by HIF-1 is reduced mitochondrial mass and/or metabolism. Exposure of HIF-1-deficient fibroblasts to chronic hypoxia results in cell death due to excessive levels of reactive oxygen species (ROS). HIF-1 reduces ROS production under hypoxic conditions by multiple mechanisms including: a subunit switch in cytochrome c oxidase from the COX4-1 to COX4-2 regulatory subunit that increases the efficiency of complex IV; induction of pyruvate dehydrogenase kinase 1, which shunts pyruvate away from the mitochondria; induction of BNIP3, which triggers mitochondrial selective autophagy; and induction of microRNA-210, which blocks assembly of Fe/S clusters that are required for oxidative phosphorylation. HIF-1 is also required for ischemic preconditioning and this effect may be due in part to its induction of CD73, the enzyme that produces adenosine. HIF-1-dependent regulation of mitochondrial metabolism may also contribute to the protective effects of ischemic preconditioning. PMID:20732359

  5. Multiple membrane-cytoplasmic domain contacts in the cystic fibrosis transmembrane conductance regulator (CFTR) mediate regulation of channel gating.

    PubMed

    He, Lihua; Aleksandrov, Andrei A; Serohijos, Adrian W R; Hegedus, Tamás; Aleksandrov, Luba A; Cui, Liying; Dokholyan, Nikolay V; Riordan, John R

    2008-09-26

    The cystic fibrosis transmembrane conductance regulator (CFTR) is a unique ATP-binding cassette (ABC) ion channel mutated in patients with cystic fibrosis. The most common mutation, deletion of phenylalanine 508 (DeltaF508) and many other disease-associated mutations occur in the nucleotide binding domains (NBD) and the cytoplasmic loops (CL) of the membrane-spanning domains (MSD). A recently constructed computational model of the CFTR three-dimensional structure, supported by experimental data (Serohijos, A. W., Hegedus, T., Aleksandrov, A. A., He, L., Cui, L., Dokholyan, N. V., and Riordan, J. R. (2008) Proc. Natl. Acad. Sci. U. S. A. 105, 3256-3261) revealed that several of these mutations including DeltaF508 disrupted interfaces between these domains. Here we have used cysteine cross-linking experiments to verify all NBD/CL interfaces predicted by the structural model and observed that their cross-linking has a variety of different effects on channel gating. The interdomain contacts comprise aromatic clusters important for stabilization of the interfaces and also involve the Q-loops and X-loops that are in close proximity to the ATP binding sites. Cross-linking of all domain-swapping contacts between NBDs and MSD cytoplasmic loops in opposite halves of the protein rapidly and reversibly arrest single channel gating while those in the same halves have lesser impact. These results reinforce the idea that mediation of regulatory signals between cytoplasmic- and membrane-integrated domains of the CFTR channel apparently relies on an array of precise but highly dynamic interdomain structural joints. PMID:18658148

  6. FOG-2 mediated recruitment of the NuRD complex regulates cardiomyocyte proliferation during heart development.

    PubMed

    Garnatz, Audrey S; Gao, Zhiguang; Broman, Michael; Martens, Spencer; Earley, Judy U; Svensson, Eric C

    2014-11-01

    FOG-2 is a multi-zinc finger protein that binds the transcriptional activator GATA4 and modulates GATA4-mediated regulation of target genes during heart development. Our previous work has demonstrated that the Nucleosome Remodeling and Deacetylase (NuRD) complex physically interacts with FOG-2 and is necessary for FOG-2 mediated repression of GATA4 activity in vitro. However, the relevance of this interaction for FOG-2 function in vivo has remained unclear. In this report, we demonstrate the importance of FOG-2/NuRD interaction through the generation and characterization of mice homozygous for a mutation in FOG-2 that disrupts NuRD binding (FOG-2(R3K5A)). These mice exhibit a perinatal lethality and have multiple cardiac malformations, including ventricular and atrial septal defects and a thin ventricular myocardium. To investigate the etiology of the thin myocardium, we measured the rate of cardiomyocyte proliferation in wild-type and FOG-2(R3K5A) developing hearts. We found cardiomyocyte proliferation was reduced by 31±8% in FOG-2(R3K5A) mice. Gene expression analysis indicated that the cell cycle inhibitor Cdkn1a (p21(cip1)) is up-regulated 2.0±0.2-fold in FOG-2(R3K5A) hearts. In addition, we demonstrate that FOG-2 can directly repress the activity of the Cdkn1a gene promoter, suggesting a model by which FOG-2/NuRD promotes ventricular wall thickening by repression of this cell cycle inhibitor. Consistent with this notion, the genetic ablation of Cdkn1a in FOG-2(R3K5A) mice leads to an improvement in left ventricular function and a partial rescue of left ventricular wall thickness. Taken together, our results define a novel mechanism in which FOG-2/NuRD interaction is required for cardiomyocyte proliferation by directly down-regulating the cell cycle inhibitor Cdkn1a during heart development. PMID:25196150

  7. CXCR4 mediated chemotaxis is regulated by 5T4 oncofetal glycoprotein in mouse embryonic cells.

    PubMed

    Southgate, Thomas D; McGinn, Owen J; Castro, Fernanda V; Rutkowski, Andrzej J; Al-Muftah, Mariam; Marinov, Georgi; Smethurst, Graeme J; Shaw, David; Ward, Christopher M; Miller, Crispin J; Stern, Peter L

    2010-01-01

    5T4 oncofetal molecules are highly expressed during development and upregulated in cancer while showing only low levels in some adult tissues. Upregulation of 5T4 expression is a marker of loss of pluripotency in the early differentiation of embryonic stem (ES) cells and forms an integrated component of an epithelial-mesenchymal transition, a process important during embryonic development and metastatic spread of epithelial tumors. Investigation of the transcriptional changes in early ES differentiation showed upregulation of CXCL12 and down-regulation of a cell surface protease, CD26, which cleaves this chemokine. CXCL12 binds to the widely expressed CXCR4 and regulates key aspects of development, stem cell motility and tumour metastasis to tissues with high levels of CXCL12. We show that the 5T4 glycoprotein is required for optimal functional cell surface expression of the chemokine receptor CXCR4 and CXCL12 mediated chemotaxis in differentiating murine embryonic stem cells and embryo fibroblasts (MEF). Cell surface expression of 5T4 and CXCR4 molecules is co-localized in differentiating ES cells and MEF. By contrast, differentiating ES and MEF derived from 5T4 knockout (KO) mice show only intracellular CXCR4 expression but infection with adenovirus encoding mouse 5T4 restores CXCL12 chemotaxis and surface co-localization with 5T4 molecules. A series of chimeric constructs with interchanged domains of 5T4 and the glycoprotein CD44 were used to map the 5T4 sequences relevant for CXCR4 membrane expression and function in 5T4KO MEF. These data identified the 5T4 transmembrane domain as sufficient and necessary to enable CXCR4 cell surface expression and chemotaxis. Furthermore, some monoclonal antibodies against m5T4 can inhibit CXCL12 chemotaxis of differentiating ES cells and MEF which is not mediated by simple antigenic modulation. Collectively, these data support a molecular interaction of 5T4 and CXCR4 occurring at the cell surface which directly facilitates

  8. FOG-2 Mediated Recruitment of the NuRD Complex Regulates Cardiomyocyte Proliferation during Heart Development

    PubMed Central

    Garnatz, Audrey S.; Gao, Zhiguang; Broman, Michael; Martens, Spencer; Earley, Judy U.; Svensson, Eric C.

    2014-01-01

    SUMMARY FOG-2 is a multi-zinc finger protein that binds the transcriptional activator GATA4 and modulates GATA4-mediated regulation of target genes during heart development. Our previous work has demonstrated that the Nucleosome Remodeling and Deacetylase (NuRD) complex physically interacts with FOG-2 and is necessary for FOG-2 mediated repression of GATA4 activity in vitro. However, the relevance of this interaction for FOG-2 function in vivo has remained unclear. In this report, we demonstrate the importance of FOG-2/NuRD interaction through the generation and characterization of mice homozygous for a mutation in FOG-2 that disrupts NuRD binding (FOG-2R3K5A). These mice exhibit a perinatal lethality and have multiple cardiac malformations, including ventricular and atrial septal defects and a thin ventricular myocardium. To investigate the etiology of the thin myocardium, we measured the rate of cardiomyocyte proliferation in wild-type and FOG-2R3K5A developing hearts. We found cardiomyocyte proliferation was reduced by 31 ± 8% in FOG-2R3K5A mice. Gene expression analysis indicated that the cell cycle inhibitor Cdkn1a (p21cip1) is up-regulated 2.0 ± 0.2-fold in FOG-2R3K5A hearts. In addition, we demonstrate that FOG-2 can directly repress the activity of the Cdkn1a gene promoter, suggesting a model by which FOG-2/NuRD promotes ventricular wall thickening by repression of this cell cycle inhibitor. Consistent with this notion, the genetic ablation of Cdkn1a in FOG-2R3K5A mice leads to an improvement in left ventricular function and a partial rescue of left ventricular wall thickness. Taken together, our results define a novel mechanism in which FOG-2/NuRD interaction is required for cardiomyocyte proliferation by directly down-regulating the cell cycle inhibitor Cdkn1a during heart development. PMID:25196150

  9. Mildew-Omics: How Global Analyses Aid the Understanding of Life and Evolution of Powdery Mildews

    PubMed Central

    Bindschedler, Laurence V.; Panstruga, Ralph; Spanu, Pietro D.

    2016-01-01

    The common powdery mildew plant diseases are caused by ascomycete fungi of the order Erysiphales. Their characteristic life style as obligate biotrophs renders functional analyses in these species challenging, mainly because of experimental constraints to genetic manipulation. Global large-scale (“-omics”) approaches are thus particularly valuable and insightful for the characterisation of the life and evolution of powdery mildews. Here we review the knowledge obtained so far from genomic, transcriptomic and proteomic studies in these fungi. We consider current limitations and challenges regarding these surveys and provide an outlook on desired future investigations on the basis of the various –omics technologies. PMID:26913042

  10. Protein kinase C beta II suppresses colorectal cancer by regulating IGF-1 mediated cell survival.

    PubMed

    Dowling, Catríona M; Phelan, James; Callender, Julia A; Cathcart, Mary Clare; Mehigan, Brian; McCormick, Paul; Dalton, Tara; Coffey, John C; Newton, Alexandra C; O'Sullivan, Jacintha; Kiely, Patrick A

    2016-04-12

    Despite extensive efforts, cancer therapies directed at the Protein Kinase C (PKC) family of serine/threonine kinases have failed in clinical trials. These therapies have been directed at inhibiting PKC and have, in some cases, worsened disease outcome. Here we examine colon cancer patients and show not only that PKC Beta II is a tumour suppressor, but patients with low levels of this isozyme have significantly decreased disease free survival. Specifically, analysis of gene expression levels of all PKC genes in matched normal and cancer tissue samples from colon cancer patients revealed a striking down-regulation of the gene coding PKC Beta in the cancer tissue (n = 21). Tissue microarray analysis revealed a dramatic down-regulation of PKC Beta II protein levels in both the epithelial and stromal diseased tissue (n = 166). Of clinical significance, low levels of the protein in the normal tissue of patients is associated with a low (10%) 10 year survival compared with a much higher (60%) survival in patients with relatively high levels of the protein. Consistent with PKC Beta II levels protecting against colon cancer, overexpression of PKC Beta II in colon cancer cell lines reveals that PKC Beta II reverses transformation in cell based assays. Further to this, activation of PKC Beta II results in a dramatic downregulation of IGF-I-induced AKT, indicating a role for PKCs in regulating IGF-1 mediated cell survival. Thus, PKC Beta II is a tumour suppressor in colon cancer and low levels serve as a predictor for poor survival outcome. PMID:26989024

  11. Opposite effects of genistein on the regulation of insulin-mediated glucose homeostasis in adipose tissue

    PubMed Central

    Wang, M; Gao, X J; Zhao, W W; Zhao, W J; Jiang, C H; Huang, F; Kou, J P; Liu, B L; Liu, K

    2013-01-01

    BACKGROUND AND PURPOSE Genistein is an isoflavone phytoestrogen found in a number of plants such as soybeans and there is accumulating evidence that it has beneficial effects on the regulation of glucose homeostasis. In this study we evaluated the effect of genistein on glucose homeostasis and its underlying mechanisms in normal and insulin-resistant conditions. EXPERIMENTAL APPROACH To induce insulin resistance, mice or differentiated 3T3-L1 adipocytes were treated with macrophage-derived conditioned medium. A glucose tolerance test was used to investigate the effect of genistein. Insulin signalling activation, glucose transporter-4 (GLUT4) translocation and AMP-activated PK (AMPK) activation were detected by Western blot analysis or elisa. KEY RESULTS Genistein impaired glucose tolerance and attenuated insulin sensitivity in normal mice by inhibiting the insulin-induced phosphorylation of insulin receptor substrate-1 (IRS1) at tyrosine residues, leading to inhibition of insulin-mediated GLUT4 translocation in adipocytes. Mac-CM, an inflammatory stimulus induced glucose intolerance accompanied by impaired insulin sensitivity; genistein reversed these changes by restoring the disturbed IRS1 function, leading to an improvement in GLUT4 translocation. In addition, genistein increased AMPK activity under both normal and inflammatory conditions; this was shown to contribute to the anti-inflammatory effect of genistein, which leads to an improvement in insulin signalling and the amelioration of insulin resistance. CONCLUSION AND IMPLICATIONS Genistein showed opposite effects on insulin sensitivity under normal and inflammatory conditions in adipose tissue and this action was derived from its negative or positive regulation of IRS1 function. Its up-regulation of AMPK activity contributes to the inhibition of inflammation implicated in insulin resistance. PMID:23763311

  12. Complex Regulation Pathways of AmpC-Mediated β-Lactam Resistance in Enterobacter cloacae Complex

    PubMed Central

    Guérin, François; Isnard, Christophe; Giard, Jean Christophe

    2015-01-01

    Enterobacter cloacae complex (ECC), an opportunistic pathogen causing numerous infections in hospitalized patients worldwide, is able to resist β-lactams mainly by producing the AmpC β-lactamase enzyme. AmpC expression is highly inducible in the presence of some β-lactams, but the underlying genetic regulation, which is intricately linked to peptidoglycan recycling, is still poorly understood. In this study, we constructed different mutant strains that were affected in genes encoding enzymes suspected to be involved in this pathway. As expected, the inactivation of ampC, ampR (which encodes the regulator protein of ampC), and ampG (encoding a permease) abolished β-lactam resistance. Reverse transcription-quantitative PCR (qRT-PCR) experiments combined with phenotypic studies showed that cefotaxime (at high concentrations) and cefoxitin induced the expression of ampC in different ways: one involving NagZ (a N-acetyl-β-d-glucosaminidase) and another independent of NagZ. Unlike the model established for Pseudomonas aeruginosa, inactivation of DacB (also known as PBP4) was not responsible for a constitutive ampC overexpression in ECC, whereas it caused AmpC-mediated high-level β-lactam resistance, suggesting a post-transcriptional regulation mechanism. Global transcriptomic analysis by transcriptome sequencing (RNA-seq) of a dacB deletion mutant confirmed these results. Lastly, analysis of 37 ECC clinical isolates showed that amino acid changes in the AmpD sequence were likely the most crucial event involved in the development of high-level β-lactam resistance in vivo as opposed to P. aeruginosa where dacB mutations have been commonly found. These findings bring new elements for a better understanding of β-lactam resistance in ECC, which is essential for the identification of novel potential drug targets. PMID:26438498

  13. Lipocalin 2, a Regulator of Retinoid Homeostasis and Retinoid-mediated Thermogenic Activation in Adipose Tissue.

    PubMed

    Guo, Hong; Foncea, Rocio; O'Byrne, Sheila M; Jiang, Hongfeng; Zhang, Yuanyuan; Deis, Jessica A; Blaner, William S; Bernlohr, David A; Chen, Xiaoli

    2016-05-20

    We have recently characterized the role of lipocalin 2 (Lcn2) as a new adipose-derived cytokine in the regulation of adaptive thermogenesis via a non-adrenergic pathway. Herein, we explored a potential non-adrenergic mechanism by which Lcn2 regulates thermogenesis and lipid metabolism. We found that Lcn2 is a retinoic acid target gene, and retinoic acid concurrently stimulated UCP1 and Lcn2 expression in adipocytes. Lcn2 KO mice exhibited a blunted effect of all-trans-retinoic acid (ATRA) on body weight and fat mass, lipid metabolism, and retinoic acid signaling pathway activation in adipose tissue under the high fat diet-induced obese condition. We further demonstrated that Lcn2 is required for the full action of ATRA on the induction of UCP1 and PGC-1α expression in brown adipocytes and the restoration of cold intolerance in Lcn2 KO mice. Interestingly, we discovered that Lcn2 KO mice have decreased levels of retinoic acid and retinol in adipose tissue. The protein levels of STRA6 responsible for retinol uptake were significantly decreased in adipose tissue. The retinol transporter RBP4 was increased in adipose tissue but decreased in the circulation, suggesting the impairment of RBP4 secretion in Lcn2 KO adipose tissue. Moreover, Lcn2 deficiency abolished the ATRA effect on RBP4 expression in adipocytes. All the data suggest that the decreased retinoid level and action are associated with impaired retinol transport and storage in adipose tissue in Lcn2 KO mice. We conclude that Lcn2 plays a critical role in regulating metabolic homeostasis of retinoids and retinoid-mediated thermogenesis in adipose tissue. PMID:27008859

  14. Protein kinase C beta II suppresses colorectal cancer by regulating IGF-1 mediated cell survival

    PubMed Central

    Dowling, Catríona M.; Phelan, James; Callender, Julia A.; Cathcart, Mary Clare; Mehigan, Brian; McCormick, Paul; Dalton, Tara; Coffey, John C.; Newton, Alexandra C.; O'sullivan, Jacintha; Kiely, Patrick A.

    2016-01-01

    Despite extensive efforts, cancer therapies directed at the Protein Kinase C (PKC) family of serine/threonine kinases have failed in clinical trials. These therapies have been directed at inhibiting PKC and have, in some cases, worsened disease outcome. Here we examine colon cancer patients and show not only that PKC Beta II is a tumour suppressor, but patients with low levels of this isozyme have significantly decreased disease free survival. Specifically, analysis of gene expression levels of all PKC genes in matched normal and cancer tissue samples from colon cancer patients revealed a striking down-regulation of the gene coding PKC Beta in the cancer tissue (n = 21). Tissue microarray analysis revealed a dramatic down-regulation of PKC Beta II protein levels in both the epithelial and stromal diseased tissue (n = 166). Of clinical significance, low levels of the protein in the normal tissue of patients is associated with a low (10%) 10 year survival compared with a much higher (60%) survival in patients with relatively high levels of the protein. Consistent with PKC Beta II levels protecting against colon cancer, overexpression of PKC Beta II in colon cancer cell lines reveals that PKC Beta II reverses transformation in cell based assays. Further to this, activation of PKC Beta II results in a dramatic downregulation of IGF-I-induced AKT, indicating a role for PKCs in regulating IGF-1 mediated cell survival. Thus, PKC Beta II is a tumour suppressor in colon cancer and low levels serve as a predictor for poor survival outcome. PMID:26989024

  15. Luteinizing Hormone Receptor mRNA Down-Regulation Is Mediated through ERK-Dependent Induction of RNA Binding Protein

    PubMed Central

    Menon, Bindu; Franzo-Romain, Megan; Damanpour, Shadi

    2011-01-01

    The ligand-induced down-regulation of LH receptor (LHR) expression in the ovaries, at least in part, is regulated by a posttranscriptional process mediated by a specific LH receptor mRNA binding protein (LRBP). The LH-mediated signaling pathways involved in this process were examined in primary cultures of human granulosa cells. Treatment with 10 IU human chorionic gonadotropin (hCG) for 12 h resulted in the down-regulation of LHR mRNA expression while producing an increase in LHR mRNA binding to LRBP as well as a 2-fold increase in LRBP levels. The activation of ERK½ pathway in LH-mediated LHR mRNA down-regulation was also established by demonstrating the translocation of ERK½ from the cytosol to the nucleus using confocal microcopy. Inhibition of protein kinase A using H-89 or ERK½ by U0126 abolished the LH-induced LHR mRNA down-regulation. These treatments also abrogated both the increases in LRBP levels as well as the LHR mRNA binding activity. The abolishment of the hCG-induced increase in LRBP levels and LHR mRNA binding activity was further confirmed by transfecting granulosa cells with ERK½ specific small interfering RNA. This treatment also reversed the hCG-induced down-regulation of LHR mRNA. These data show that LH-regulated ERK½ signaling is required for the LRBP-mediated down-regulation of LHR mRNA. PMID:21147848

  16. A novel crosstalk between TLR4- and NOD2-mediated signaling in the regulation of intestinal inflammation.

    PubMed

    Kim, Hajeong; Zhao, Quanju; Zheng, Hua; Li, Xin; Zhang, Tuo; Ma, Xiaojing

    2015-01-01

    Although Toll-like receptor 4 (TLR4)- and nucleotide-binding oligomerization domain 2 (NOD2)-mediated signaling mechanisms have been extensively studied individually, the crosstalk between them in the regulation of intestinal mucosal defense and tissue homeostasis has been underappreciated. Here, we uncover some novel activities of NOD2 by gene expression profiling revealing the global nature of the cross-regulation between TLR4- and NOD2-mediated signaling. Specifically, NOD2 is able to sense the intensity of TLR4-mediated signaling, resulting in either synergistic stimulation of Interluekin-12 (IL-12) production when the TLR signaling intensity is low; or in the inhibition of IL-12 synthesis and maintenance of intestinal mucosal homeostasis when the TLR signaling intensifies. This balancing act is mediated through receptor-interacting serine/threonine kinase 2, and the transcriptional regulator CCAAT/enhancer-binding protein α (C/EBPα) via its serine 248 phosphorylation by Protein Kinase C. Mice deficient in C/EBPα in the hematopoietic compartment are highly susceptible to chemically induced experimental colitis in an IL-12-dependent manner. Additionally, in contrast to the dogma, we find that the major Crohn's disease-associated NOD2 mutations could cause a primarily immunodeficient phenotype by selectively impairing TLR4-mediated IL-12 production and host defense. To restore the impaired homeostasis would be a way forward to developing novel therapeutic strategies for inflammatory bowel diseases. PMID:26153766

  17. Accumbal α-adrenoceptors, but not β-adrenoceptors, regulate behaviour that is mediated by reserpine-sensitive storage vesicles.

    PubMed

    Verheij, Michel M M; Saigusa, Tadashi; Koshikawa, Noriaki; Cools, Alexander R

    2015-02-01

    It has previously been demonstrated that mesolimbic α-adrenoceptors, but not β-adrenoceptors, control the release of dopamine that is derived from reserpine-sensitive storage vesicles. The aim of the present study was to investigate whether these storage vesicles also regulate α-adrenoceptor-mediated or β-adrenoceptor-mediated changes in behaviour. Accordingly, rats were pretreated with reserpine before the α-adrenoceptor antagonist phentolamine or the β-adrenoceptor agonist isoproterenol was locally applied to the nucleus accumbens. Both phentolamine and isoproterenol increased the duration of walking, rearing and grooming and decreased the duration of sitting. Reserpine counteracted the behavioural response elicited by phentolamine but not by isoproterenol. The results of the present study demonstrate that mesolimbic α-adrenoceptors, but not β-adrenoceptors, regulate behaviour that is mediated by reserpine-sensitive storage pools. It is hypothesized that the observed α-adrenoceptor-mediated increase in locomotor activity is due to the α-adrenoceptor-mediated increase in the release of accumbal intravesicular dopamine. Our finding that α-adrenoceptors inhibit, whereas β-adrenoceptors stimulate, locomotor activity may help explain why noradrenaline or environmental stressors have previously been found to have opposing effects on the regulation of behaviour. PMID:25325287

  18. A novel crosstalk between TLR4- and NOD2-mediated signaling in the regulation of intestinal inflammation

    PubMed Central

    Kim, Hajeong; Zhao, Quanju; Zheng, Hua; Li, Xin; Zhang, Tuo; Ma, Xiaojing

    2015-01-01

    Although Toll-like receptor 4 (TLR4)- and nucleotide-binding oligomerization domain 2 (NOD2)-mediated signaling mechanisms have been extensively studied individually, the crosstalk between them in the regulation of intestinal mucosal defense and tissue homeostasis has been underappreciated. Here, we uncover some novel activities of NOD2 by gene expression profiling revealing the global nature of the cross-regulation between TLR4- and NOD2-mediated signaling. Specifically, NOD2 is able to sense the intensity of TLR4-mediated signaling, resulting in either synergistic stimulation of Interluekin-12 (IL-12) production when the TLR signaling intensity is low; or in the inhibition of IL-12 synthesis and maintenance of intestinal mucosal homeostasis when the TLR signaling intensifies. This balancing act is mediated through receptor-interacting serine/threonine kinase 2, and the transcriptional regulator CCAAT/enhancer-binding protein α (C/EBPα) via its serine 248 phosphorylation by Protein Kinase C. Mice deficient in C/EBPα in the hematopoietic compartment are highly susceptible to chemically induced experimental colitis in an IL-12-dependent manner. Additionally, in contrast to the dogma, we find that the major Crohn’s disease-associated NOD2 mutations could cause a primarily immunodeficient phenotype by selectively impairing TLR4-mediated IL-12 production and host defense. To restore the impaired homeostasis would be a way forward to developing novel therapeutic strategies for inflammatory bowel diseases. PMID:26153766

  19. RNA-mediated regulation in Gram-positive pathogens: an overview punctuated with examples from the group A Streptococcus

    PubMed Central

    Miller, Eric W.; Cao, Tram N.; Pflughoeft, Kathryn J.; Sumby, Paul

    2014-01-01

    RNA-based mechanisms of regulation represent a ubiquitous class of regulators that are associated with diverse processes including nutrient sensing, stress response, modulation of horizontal gene transfer, and virulence factor expression. While better studied in Gram-negative bacteria, the literature is replete with examples of the importance of RNA-mediated regulatory mechanisms to the virulence and fitness of Gram-positives. Regulatory RNAs are classified as cis-acting, e.g. riboswitches, which modulate the transcription, translation, or stability of co-transcribed RNA, or trans-acting, e.g. small regulatory RNAs, which target separate mRNAs or proteins. The group A Streptococcus (GAS, Streptococcus pyogenes) is a Gram-positive bacterial pathogen from which several regulatory RNA mechanisms have been characterized. The study of RNA-mediated regulation in GAS has uncovered novel concepts with respect to how small regulatory RNAs may positively regulate target mRNA stability, and to how CRISPR RNAs are processed from longer precursors. This review provides an overview of RNA-mediated regulation in Gram-positive bacteria, and is highlighted with specific examples from GAS research. The key roles that these systems play in regulating bacterial virulence are discussed and future perspectives outlined. PMID:25091277

  20. Androgen receptor-mediated non-genomic regulation of prostate cancer cell proliferation

    PubMed Central

    Liao, Ross S.; Ma, Shihong; Miao, Lu; Li, Rui; Yin, Yi

    2013-01-01

    Androgen receptor (AR)-mediated signaling is necessary for prostate cancer cell proliferation and an important target for therapeutic drug development. Canonically, AR signals through a genomic or transcriptional pathway, involving the translocation of androgen-bound AR to the nucleus, its binding to cognate androgen response elements on promoter, with ensuing modulation of target gene expression, leading to cell proliferation. However, prostate cancer cells can show dose-dependent proliferation responses to androgen within minutes, without the need for genomic AR signaling. This proliferation response known as the non-genomic AR signaling is mediated by cytoplasmic AR, which facilitates the activation of kinase-signaling cascades, including the Ras-Raf-1, phosphatidyl-inositol 3-kinase (PI3K)/Akt and protein kinase C (PKC), which in turn converge on mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) activation, leading to cell proliferation. Further, since activated ERK may also phosphorylate AR and its coactivators, the non-genomic AR signaling may enhance AR genomic activity. Non-genomic AR signaling may occur in an ERK-independent manner, via activation of mammalian target of rapamycin (mTOR) pathway, or modulation of intracellular Ca2+ concentration through plasma membrane G protein-coupled receptors (GPCRs). These data suggest that therapeutic strategies aimed at preventing AR nuclear translocation and genomic AR signaling alone may not completely abrogate AR signaling. Thus, elucidation of mechanisms that underlie non-genomic AR signaling may identify potential mechanisms of resistance to current anti-androgens and help developing novel therapies that abolish all AR signaling in prostate cancer. PMID:26816736

  1. Chenodeoxycholic acid-mediated activation of the farnesoid X receptor negatively regulates hydroxysteroid sulfotransferase.

    PubMed

    Miyata, Masaaki; Matsuda, Yoshiki; Tsuchiya, Hiroyuki; Kitada, Hirotaka; Akase, Takanori; Shimada, Miki; Nagata, Kiyoshi; Gonzalez, Frank J; Yamazoe, Yasushi

    2006-08-01

    Hydroxysteroid sulfotransferase catalyzing bile acid sulfation plays an essential role in protection against lithocholic acid (LCA)-induced liver toxicity. Hepatic levels of Sult2a is up to 8-fold higher in farnesoid X receptor-null mice than in the wild-type mice. Thus, the influence of FXR ligand (chenodeoxycholic acid (CDCA) and LCA) feeding on hepatic Sult2a expression was examined in FXR-null and wild-type mice. Hepatic Sult2a protein content was elevated in FXR-null and wild-type mice fed a LCA (1% and 0.5%) diet. Treatment with 0.5% CDCA diet decreased hepatic Sult2a to 20% of the control in wild-type mice, but increased the content in FXR-null mice. Liver Sult2a1 (St2a4) mRNA levels were reduced to 26% in wild-type mice after feeding of a CDCA diet, while no decrease was observed on Sult2a1 mRNA levels in FXR-null mice after CDCA feeding. A significant inverse relationship (r(2)=0.523) was found between hepatic Sult2a protein content and small heterodimer partner (SHP) mRNA level. PCN-mediated increase in Sult2a protein levels were attenuated by CDCA feeding in wild-type mice, but not in FXR-null mice. Human SULT2A1 protein and mRNA levels were decreased in HepG2 cells treated with the FXR agonists, CDCA or GW4064 in dose-dependent manners, although SHP mRNA levels were increased. These results suggest that SULT2A is negatively regulated through CDCA-mediated FXR activation in mice and humans. PMID:16946559

  2. FOXM1 mediates resistance to docetaxel in gastric cancer via up-regulating Stathmin.

    PubMed

    Li, Xiaoxiao; Yao, Ruyong; Yue, Lu; Qiu, Wensheng; Qi, Weiwei; Liu, Shihai; Yao, Yasai; Liang, Jun

    2014-05-01

    Docetaxel is commonly used as an effective chemotherapeutic drug for gastric cancer patients recently. With the increasing emergence of docetaxel resistance nowadays, identification of suitable biomarkers for predicting chemosensitivity to docetaxel may be a key role for improving therapeutic effects for gastric cancer patients. In this study, we investigated the correlation between the expression of transcription factor forkhead box protein M1 (FOXM1) and chemotherapy response to docetaxel in gastric cancer, the possible mechanism for which was further explored. As a result, FOXM1 overexpression was shown to mediate resistance to docetaxel in gastric cancers. It altered microtubule dynamics to protect tumour cells from docetaxel-induced apoptosis. Mechanistic investigations revealed that tubulin-destabilizing protein Stathmin, which mediated docetaxel resistance in FOXM1-silenced gastric cancer cells, is a direct down-stream target of FOXM1, whereas another microtubule dynamics protein mitotic centromere-associated kinesin (MCAK), shown to be related to docetaxel resistance in gastric cancer cells, is not associated with FOXM1 expression significantly. These results were further provided by immunohistochemical analysis, indicating that FOXM1 and Stathmin expression levels were correlated in 103 post-operational gastric cancer specimens. Moreover, when we attenuated FOXM1 expression with FOXM1 inhibitor thiostrepton, docetaxel resistance in gastric cancers was found to be reversed, simultaneously with the down-regulation of FOXM1 and Stathmin. Therefore, FOXM1 can be a useful marker for predicting and monitoring docetaxel response. Through the inhibition of FOXM1, docetaxel resistance can be reversed, and thus FOXM1 could be a new therapeutic target in docetaxel-resistant gastric cancer. PMID:24628949

  3. An Integrated Genomic Strategy Delineates Candidate Mediator Genes Regulating Grain Size and Weight in Rice

    PubMed Central

    Malik, Naveen; Dwivedi, Nidhi; Singh, Ashok K.; Parida, Swarup K.; Agarwal, Pinky; Thakur, Jitendra K.; Tyagi, Akhilesh K.

    2016-01-01

    The present study deployed a Mediator (MED) genes-mediated integrated genomic strategy for understanding the complex genetic architecture of grain size/weight quantitative trait in rice. The targeted multiplex amplicon resequencing of 55 MED genes annotated from whole rice genome in 384 accessions discovered 3971 SNPs, which were structurally and functionally annotated in diverse coding and non-coding sequence-components of genes. Association analysis, using the genotyping information of 3971 SNPs in a structured population of 384 accessions (with 50–100 kb linkage disequilibrium decay), detected 10 MED gene-derived SNPs significantly associated (46% combined phenotypic variation explained) with grain length, width and weight in rice. Of these, one strong grain weight-associated non-synonymous SNP (G/A)-carrying OsMED4_2 gene was validated successfully in low- and high-grain weight parental accessions and homozygous individuals of a rice mapping population. The seed-specific expression, including differential up/down-regulation of three grain size/weight-associated MED genes (including OsMED4_2) in six low and high-grain weight rice accessions was evident. Altogether, combinatorial genomic approach involving haplotype-based association analysis delineated diverse functionally relevant natural SNP-allelic variants in 10 MED genes, including three potential novel SNP haplotypes in an OsMED4_2 gene governing grain size/weight differentiation in rice. These molecular tags have potential to accelerate genomics-assisted crop improvement in rice. PMID:27000976

  4. TUSC5 regulates insulin-mediated adipose tissue glucose uptake by modulation of GLUT4 recycling

    PubMed Central

    Beaton, Nigel; Rudigier, Carla; Moest, Hansjörg; Müller, Sebastian; Mrosek, Nadja; Röder, Eva; Rudofsky, Gottfried; Rülicke, Thomas; Ukropec, Jozef; Ukropcova, Barbara; Augustin, Robert; Neubauer, Heike; Wolfrum, Christian

    2015-01-01

    Objective Failure to properly dispose of glucose in response to insulin is a serious health problem, occurring during obesity and is associated with type 2 diabetes development. Insulin-stimulated glucose uptake is facilitated by the translocation and plasma membrane fusion of vesicles containing glucose transporter 4 (GLUT4), the rate-limiting step of post-prandial glucose disposal. Methods We analyzed the role of Tusc5 in the regulation of insulin-stimulated Glut4-mediated glucose uptake in vitro and in vivo. Furthermore, we measured Tusc5 expression in two patient cohorts. Results Herein, we report that TUSC5 controls insulin-stimulated glucose uptake in adipocytes, in vitro and in vivo. TUSC5 facilitates the proper recycling of GLUT4 and other key trafficking proteins during prolonged insulin stimulation, thereby enabling proper protein localization and complete vesicle formation, processes that ultimately enable insulin-stimulated glucose uptake. Tusc5 knockout mice exhibit impaired glucose disposal and TUSC5 expression is predictive of glucose tolerance in obese individuals, independent of body weight. Furthermore, we show that TUSC5 is a PPARγ target and in its absence the anti-diabetic effects of TZDs are significantly blunted. Conclusions Collectively, these findings establish TUSC5 as an adipose tissue-specific protein that enables proper protein recycling, linking the ubiquitous vesicle traffic machinery with tissue-specific insulin-mediated glucose uptake into adipose tissue and the maintenance of a healthy metabolic phenotype in mice and humans. PMID:26629404

  5. The Mediation Effects of Dysfunctional Beliefs and Emotional Regulation on Children's Perceived Parental Conflict and Internalizing and Externalizing Problems

    ERIC Educational Resources Information Center

    Lee, Ji-yeon; Wesbecher, Kristen; Lee, Mihwa; Lee, Jeeyon

    2015-01-01

    The purpose of this study was to examine the mediational effects of dysfunctional beliefs and difficulties in emotional regulation on children's perception of interparental conflict and subsequent internalizing and externalizing problems. The participants in this study were 335 fifth grade elementary school students in Korea. We hypothesized that…

  6. Empathy and Self-Regulation as Mediators between Parenting and Adolescents' Prosocial Behavior toward Strangers, Friends, and Family

    ERIC Educational Resources Information Center

    Padilla-Walker, Laura M.; Christensen, Katherine J.

    2011-01-01

    The current study examined the role of empathy and self-regulation as mediators between positive parenting (mothering and fathering) and early adolescents' prosocial behavior toward 3 targets (strangers, friends, and family). Data were taken from Time 1 and Time 2 of the "Flourishing Families Project," and included reports from 500 families with…

  7. Links between Maternal and Child Psychopathology Symptoms: Mediation through Child Emotion Regulation and Moderation through Maternal Behavior

    ERIC Educational Resources Information Center

    Suveg, Cynthia; Shaffer, Anne; Morelen, Diana; Thomassin, Kristel

    2011-01-01

    This study examined the intergenerational transmission of psychopathology symptoms with 7-12 year-old children (N = 97; 44 boys, 53 girls, M age = 9.14, SD = 1.38) and their mothers (M age = 38.46, SD = 6.86). Child emotion regulation mediated the links between maternal psychopathology and child internalizing and externalizing symptoms. In turn,…

  8. The Mediation Role of Intrinsic and Extrinsic Motivation in the Relationship between Creative Educational Environment and Metacognitive Self-Regulation

    ERIC Educational Resources Information Center

    Maralani, Farnaz Mehdipour

    2016-01-01

    This study investigated the mediation role of intrinsic and extrinsic motivation in the relationship between creative educational environment and metacognitive self-regulation. Participants were 300 girls, selected randomly from the girl hostel in university of Tehran. Participants completed Akoal's creative educational environment questionnaire,…

  9. Cumulative Risk and Adolescent's Internalizing and Externalizing Problems: The Mediating Roles of Maternal Responsiveness and Self-Regulation

    ERIC Educational Resources Information Center

    Doan, Stacey N.; Fuller-Rowell, Thomas E.; Evans, Gary W.

    2012-01-01

    The purpose of the present study was to examine longitudinal associations among maternal responsiveness, self-regulation, and behavioral adjustment in adolescents. The authors used structural equation modeling to test a model that demonstrates that the effects of early cumulative risk on behavioral problems is mediated by maternal responsiveness…

  10. CDK8-Cyclin C Mediates Nutritional Regulation of Developmental Transitions through the Ecdysone Receptor in Drosophila

    PubMed Central

    Xie, Xiao-Jun; Hsu, Fu-Ning; Gao, Xinsheng; Xu, Wu; Ni, Jian-Quan; Xing, Yue; Huang, Liying; Hsiao, Hao-Ching; Zheng, Haiyan; Wang, Chenguang; Zheng, Yani; Xiaoli, Alus M.; Yang, Fajun; Bondos, Sarah E.; Ji, Jun-Yuan

    2015-01-01

    The steroid hormone ecdysone and its receptor (EcR) play critical roles in orchestrating developmental transitions in arthropods. However, the mechanism by which EcR integrates nutritional and developmental cues to correctly activate transcription remains poorly understood. Here, we show that EcR-dependent transcription, and thus, developmental timing in Drosophila, is regulated by CDK8 and its regulatory partner Cyclin C (CycC), and the level of CDK8 is affected by nutrient availability. We observed that cdk8 and cycC mutants resemble EcR mutants and EcR-target genes are systematically down-regulated in both mutants. Indeed, the ability of the EcR-Ultraspiracle (USP) heterodimer to bind to polytene chromosomes and the promoters of EcR target genes is also diminished. Mass spectrometry analysis of proteins that co-immunoprecipitate with EcR and USP identified multiple Mediator subunits, including CDK8 and CycC. Consistently, CDK8-CycC interacts with EcR-USP in vivo; in particular, CDK8 and Med14 can directly interact with the AF1 domain of EcR. These results suggest that CDK8-CycC may serve as transcriptional cofactors for EcR-dependent transcription. During the larval–pupal transition, the levels of CDK8 protein positively correlate with EcR and USP levels, but inversely correlate with the activity of sterol regulatory element binding protein (SREBP), the master regulator of intracellular lipid homeostasis. Likewise, starvation of early third instar larvae precociously increases the levels of CDK8, EcR and USP, yet down-regulates SREBP activity. Conversely, refeeding the starved larvae strongly reduces CDK8 levels but increases SREBP activity. Importantly, these changes correlate with the timing for the larval–pupal transition. Taken together, these results suggest that CDK8-CycC links nutrient intake to developmental transitions (EcR activity) and fat metabolism (SREBP activity) during the larval–pupal transition. PMID:26222308

  11. CDK8-Cyclin C Mediates Nutritional Regulation of Developmental Transitions through the Ecdysone Receptor in Drosophila.

    PubMed

    Xie, Xiao-Jun; Hsu, Fu-Ning; Gao, Xinsheng; Xu, Wu; Ni, Jian-Quan; Xing, Yue; Huang, Liying; Hsiao, Hao-Ching; Zheng, Haiyan; Wang, Chenguang; Zheng, Yani; Xiaoli, Alus M; Yang, Fajun; Bondos, Sarah E; Ji, Jun-Yuan

    2015-07-01

    The steroid hormone ecdysone and its receptor (EcR) play critical roles in orchestrating developmental transitions in arthropods. However, the mechanism by which EcR integrates nutritional and developmental cues to correctly activate transcription remains poorly understood. Here, we show that EcR-dependent transcription, and thus, developmental timing in Drosophila, is regulated by CDK8 and its regulatory partner Cyclin C (CycC), and the level of CDK8 is affected by nutrient availability. We observed that cdk8 and cycC mutants resemble EcR mutants and EcR-target genes are systematically down-regulated in both mutants. Indeed, the ability of the EcR-Ultraspiracle (USP) heterodimer to bind to polytene chromosomes and the promoters of EcR target genes is also diminished. Mass spectrometry analysis of proteins that co-immunoprecipitate with EcR and USP identified multiple Mediator subunits, including CDK8 and CycC. Consistently, CDK8-CycC interacts with EcR-USP in vivo; in particular, CDK8 and Med14 can directly interact with the AF1 domain of EcR. These results suggest that CDK8-CycC may serve as transcriptional cofactors for EcR-dependent transcription. During the larval-pupal transition, the levels of CDK8 protein positively correlate with EcR and USP levels, but inversely correlate with the activity of sterol regulatory element binding protein (SREBP), the master regulator of intracellular lipid homeostasis. Likewise, starvation of early third instar larvae precociously increases the levels of CDK8, EcR and USP, yet down-regulates SREBP activity. Conversely, refeeding the starved larvae strongly reduces CDK8 levels but increases SREBP activity. Importantly, these changes correlate with the timing for the larval-pupal transition. Taken together, these results suggest that CDK8-CycC links nutrient intake to developmental transitions (EcR activity) and fat metabolism (SREBP activity) during the larval-pupal transition. PMID:26222308

  12. Cumulative childhood trauma and psychological maladjustment of sexually abused children in Korea: mediating effects of emotion regulation.

    PubMed

    Choi, Ji Young; Oh, Kyung Ja

    2014-02-01

    The purpose of the present study was to identify the mediating effects of emotion regulation on the association between cumulative childhood trauma and behavior problems in sexually abused children in Korea, using structural equation modeling (SEM). Data were collected on 171 children (ages 6-13 years) referred to a public counseling center for sexual abuse in Seoul, Korea. Cumulative childhood traumas were defined on the basis of number of traumas (physical abuse, witnessing domestic violence, neglect, traumatic separation from parent, and sexual abuse) and the severity and duration of traumas. Children were evaluated by their parents on emotion regulation using the Emotion Regulation Checklist and internalizing and externalizing behavior problems using the Korean-Child Behavior Checklist. SEM analyses confirmed the complete mediation model, in which emotion dysregulation fully mediates the relationship between cumulative childhood traumas and internalizing/externalizing behavior problems. These findings indicate that emotion regulation is an important mechanism that can explain the negative effects of cumulative childhood traumas and that there is a need to focus on emotion regulation in sexually abused children exposed to cumulative trauma. PMID:24210271

  13. Emotion Regulation Mediates the Association Between ADHD and Depressive Symptoms in a Community Sample of Youth

    PubMed Central

    Seymour, Karen E.; Chronis-Tuscano, Andrea; Iwamoto, Derek K.; Kurdziel, Gretchen; MacPherson, Laura

    2014-01-01

    The purpose of this study was to examine the longitudinal relationship between attention-deficit/ hyperactivity disorder (ADHD) symptoms, emotion regulation (ER) ability, and depressive symptoms within a diverse community sample of 277 youth, ages 9–12 (56 % male). Participants were drawn from a larger study examining adolescent risk behaviors, and completed annual assessments over 3 years. Youth ADHD symptoms were assessed at Time 1 (T1) using the parent-reported Disruptive Behavior Disorders Rating Scale, ER was assessed with the parent-reported Emotion Regulation Checklist at Time 2 (T2), and youth depressive symptoms were assessed using the self-reported Revised Child Anxiety and Depression Scales at Time 3 (T3). Analyses examined T2 ER as a mediator between T1 ADHD symptoms (including the unique contributions of inattentive [IA] versus hyperactive/impulsive [HI] symptoms) and T3 depressive symptoms. Structural equation modeling (SEM) indicated the path model specified provided an excellent fit to the data. Tests of indirect effects suggested that T2 ER appears to be a significant mechanism that underlies the relationship between T1 ADHD and T3 depression, even when accounting for T1 oppositional defiant and depressive symptoms. Furthermore, while both T1 IA and HI symptoms had significant indirect effects on T3 depression through the mechanism T2 ER, HI proved a more robust predictor of T2 ER than IA. Results of this prospective study support cross-sectional findings pointing to ER as a potential mechanism linking ADHD and depressive symptoms in youth. Clinical implications and future directions are discussed. PMID:24221724

  14. Phosphorylation Regulates Id2 Degradation and Mediates the Proliferation of Neural Precursor Cells

    PubMed Central

    Sullivan, Jaclyn M.; Havrda, Matthew C.; Kettenbach, Arminja N.; Paolella, Brenton R.; Zhang, Zhonghua; Gerber, Scott A.; Israel, Mark A.

    2016-01-01

    Inhibitor of DNA binding proteins (Id1-Id4) function to inhibit differentiation and promote proliferation of many different cell types. Among the Id family members, Id2 has been most extensively studied in the central nervous system (CNS). Id2 contributes to cultured neural precursor cell (NPC) proliferation as well as to the proliferation of CNS tumors such as glioblastoma that are likely to arise from NPC-like cells. We identified three phosphorylation sites near the N-terminus of Id2 in NPCs. To interrogate the importance of Id2 phosphorylation, Id2−/− NPCs were modified to express wild type (WT) Id2 or an Id2 mutant protein that could not be phosphorylated at the identified sites. We observed that NPCs expressing this mutant lacking phosphorylation near the N-terminus had higher steady-state levels of Id2 when compared to NPCs expressing WT Id2. This elevated level was the result of a longer half-life and reduced proteasome-mediated degradation. Moreover, NPCs expressing constitutively de-phosphorylated Id2 proliferated more rapidly than NPCs expressing WT Id2, a finding consistent with the well-characterized function of Id2 in driving proliferation. Observing that phosphorylation of Id2 modulates the degradation of this important cell-cycle regulator, we sought to identify a phosphatase that would stabilize Id2 enhancing its activity in NPCs and extended our analysis to include human glioblastoma-derived stem cells (GSCs). We found that expression of the phosphatase PP2A altered Id2 levels. Our findings suggest that inhibition of PP2A may be a novel strategy to regulate the proliferation of normal NPCs and malignant GSCs by decreasing Id2 levels. PMID:26756672

  15. Emotion regulation mediates the association between ADHD and depressive symptoms in a community sample of youth.

    PubMed

    Seymour, Karen E; Chronis-Tuscano, Andrea; Iwamoto, Derek K; Kurdziel, Gretchen; Macpherson, Laura

    2014-05-01

    The purpose of this study was to examine the longitudinal relationship between attention-deficit/hyperactivity disorder (ADHD) symptoms, emotion regulation (ER) ability, and depressive symptoms within a diverse community sample of 277 youth, ages 9-12 (56 % male). Participants were drawn from a larger study examining adolescent risk behaviors, and completed annual assessments over 3 years. Youth ADHD symptoms were assessed at Time 1 (T1) using the parent-reported Disruptive Behavior Disorders Rating Scale, ER was assessed with the parent-reported Emotion Regulation Checklist at Time 2 (T2), and youth depressive symptoms were assessed using the self-reported Revised Child Anxiety and Depression Scales at Time 3 (T3). Analyses examined T2 ER as a mediator between T1 ADHD symptoms (including the unique contributions of inattentive [IA] versus hyperactive/impulsive [HI] symptoms) and T3 depressive symptoms. Structural equation modeling (SEM) indicated the path model specified provided an excellent fit to the data. Tests of indirect effects suggested that T2 ER appears to be a significant mechanism that underlies the relationship between T1 ADHD and T3 depression, even when accounting for T1 oppositional defiant and depressive symptoms. Furthermore, while both T1 IA and HI symptoms had significant indirect effects on T3 depression through the mechanism T2 ER, HI proved a more robust predictor of T2 ER than IA. Results of this prospective study support cross-sectional findings pointing to ER as a potential mechanism linking ADHD and depressive symptoms in youth. Clinical implications and future directions are discussed. PMID:24221724

  16. Extracellular calmodulin regulates growth and cAMP-mediated chemotaxis in Dictyostelium discoideum

    SciTech Connect

    O'Day, Danton H.; Huber, Robert J.; Suarez, Andres

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer Extracellular calmodulin is present throughout growth and development in Dictyostelium. Black-Right-Pointing-Pointer Extracellular calmodulin localizes within the ECM during development. Black-Right-Pointing-Pointer Extracellular calmodulin inhibits cell proliferation and increases chemotaxis. Black-Right-Pointing-Pointer Extracellular calmodulin exists in eukaryotic microbes. Black-Right-Pointing-Pointer Extracellular calmodulin may be functionally as important as intracellular calmodulin. -- Abstract: The existence of extracellular calmodulin (CaM) has had a long and controversial history. CaM is a ubiquitous calcium-binding protein that has been found in every eukaryotic cell system. Calcium-free apo-CaM and Ca{sup 2+}/CaM exert their effects by binding to and regulating the activity of CaM-binding proteins (CaMBPs). Most of the research done to date on CaM and its CaMBPs has focused on their intracellular functions. The presence of extracellular CaM is well established in a number of plants where it functions in proliferation, cell wall regeneration, gene regulation and germination. While CaM has been detected extracellularly in several animal species, including frog, rat, rabbit and human, its extracellular localization and functions are less well established. In contrast the study of extracellular CaM in eukaryotic microbes remains to be done. Here we show that CaM is constitutively expressed and secreted throughout asexual development in Dictyostelium where the presence of extracellular CaM dose-dependently inhibits cell proliferation but increases cAMP mediated chemotaxis. During development, extracellular CaM localizes within the slime sheath where it coexists with at least one CaMBP, the matricellular CaM-binding protein CyrA. Coupled with previous research, this work provides direct evidence for the existence of extracellular CaM in the Dictyostelium and provides insight into its functions in this model amoebozoan.

  17. Molecular Mechanisms Mediating the Adaptive Regulation of Intestinal Riboflavin Uptake Process

    PubMed Central

    Subramanian, Veedamali S.; Ghosal, Abhisek; Kapadia, Rubina; Nabokina, Svetlana M.; Said, Hamid M.

    2015-01-01

    The intestinal absorption process of vitamin B2 (riboflavin, RF) is carrier-mediated, and all three known human RF transporters, i.e., hRFVT-1, -2, and -3 (products of the SLC52A1, 2 & 3 genes, respectively) are expressed in the gut. We have previously shown that the intestinal RF uptake process is adaptively regulated by substrate level, but little is known about the molecular mechanism(s) involved. Using human intestinal epithelial NCM460 cells maintained under RF deficient and over-supplemented (OS) conditions, we now show that the induction in RF uptake in RF deficiency is associated with an increase in expression of the hRFVT-2 & -3 (but not hRFVT-1) at the protein and mRNA levels. Focusing on hRFVT-3, the predominant transporter in the intestine, we also observed an increase in the level of expression of its hnRNA and activity of its promoter in the RF deficiency state. An increase in the level of expression of the nuclear factor Sp1 (which is important for activity of the SLC52A3 promoter) was observed in RF deficiency, while mutating the Sp1/GC site in the SLC52A3 promoter drastically decreased the level of induction in SLC52A3 promoter activity in RF deficiency. We also observed specific epigenetic changes in the SLC52A3 promoter in RF deficiency. Finally, an increase in hRFVT-3 protein expression at the cell surface was observed in RF deficiency. Results of these investigations show, for the first time, that transcriptional and post-transcriptional mechanisms are involved in the adaptive regulation of intestinal RF uptake by the prevailing substrate level. PMID:26121134

  18. Ouabain Regulates CFTR-Mediated Anion Secretion and Na,K-ATPase Transport in ADPKD Cells.

    PubMed

    Jansson, Kyle; Venugopal, Jessica; Sánchez, Gladis; Magenheimer, Brenda S; Reif, Gail A; Wallace, Darren P; Calvet, James P; Blanco, Gustavo

    2015-12-01

    Cyst enlargement in autosomal dominant polycystic kidney disease (ADPKD) requires the transepithelial secretion of fluid into the cyst lumen. We previously showed that physiological amounts of ouabain enhance cAMP-dependent fluid secretion and cyst growth of human ADPKD cyst epithelial cells in culture and formation of cyst-like dilations in metanephric kidneys from Pkd1 mutant mice. Here, we investigated the mechanisms by which ouabain promotes cAMP-dependent fluid secretion and cystogenesis. Ouabain (3 nM) enhanced cAMP-induced cyst-like dilations in embryonic kidneys from Pkd1 (m1Bei) mice, but had no effect on metanephroi from Pkd1 (m1Bei) mice that lack expression of the cystic fibrosis transmembrane conductance regulator (CFTR). Similarly, ouabain stimulation of cAMP-induced fluid secretion and in vitro cyst growth of ADPKD cells were abrogated by CFTR inhibition, showing that CFTR is required for ouabain effects on ADPKD fluid secretion. Moreover, ouabain directly enhanced the cAMP-dependent Cl(-) efflux mediated by CFTR in ADPKD monolayers. Ouabain increased the trafficking of CFTR to the plasma membrane and up-regulated the expression of the CFTR activator PDZK1. Finally, ouabain decreased plasma membrane expression and activity of the Na,K-ATPase in ADPKD cells. Altogether, these results show that ouabain enhances net fluid secretion and cyst formation by activating apical anion secretion via CFTR and decreasing basolateral Na(+) transport via Na,K-ATPase. These results provide new information on the mechanisms by which ouabain affects ADPKD cells and further highlight the importance of ouabain as a non-genomic stimulator of cystogenesis in ADPKD. PMID:26289599

  19. Differential localization of A-Raf regulates MST2-mediated apoptosis during epithelial differentiation.

    PubMed

    Rauch, J; Vandamme, D; Mack, B; McCann, B; Volinsky, N; Blanco, A; Gires, O; Kolch, W

    2016-08-01

    A-Raf belongs to the family of oncogenic Raf kinases that are involved in mitogenic signaling by activating the MEK-ERK pathway. Low kinase activity of A-Raf toward MEK suggested that A-Raf might have alternative functions. We recently identified A-Raf as a potent inhibitor of the proapoptotic mammalian sterile 20-like kinase (MST2) tumor suppressor pathway in several cancer entities including head and neck, colon, and breast. Independent of kinase activity, A-Raf binds to MST2 thereby efficiently inhibiting apoptosis. Here, we show that the interaction of A-Raf with the MST2 pathway is regulated by subcellular compartmentalization. Although in proliferating normal cells and tumor cells A-Raf localizes to the mitochondria, differentiated non-carcinogenic cells of head and neck epithelia, which express A-Raf at the plasma membrane. The constitutive or induced re-localization of A-Raf to the plasma membrane compromises its ability to efficiently sequester and inactivate MST2, thus rendering cells susceptible to apoptosis. Physiologically, A-Raf re-localizes to the plasma membrane upon epithelial differentiation in vivo. This re-distribution is regulated by the scaffold protein kinase suppressor of Ras 2 (KSR2). Downregulation of KSR2 during mammary epithelial cell differentiation or siRNA-mediated knockdown re-localizes A-Raf to the plasma membrane causing the release of MST2. By using the MCF7 cell differentiation system, we could demonstrate that overexpression of A-Raf in MCF7 cells, which induces differentiation. Our findings offer a new paradigm to understand how differential localization of Raf complexes affects diverse signaling functions in normal cells and carcinomas. PMID:26891695

  20. Phosphorylation Regulates Id2 Degradation and Mediates the Proliferation of Neural Precursor Cells.

    PubMed

    Sullivan, Jaclyn M; Havrda, Matthew C; Kettenbach, Arminja N; Paolella, Brenton R; Zhang, Zhonghua; Gerber, Scott A; Israel, Mark A

    2016-05-01

    Inhibitor of DNA binding proteins (Id1-Id4) function to inhibit differentiation and promote proliferation of many different cell types. Among the Id family members, Id2 has been most extensively studied in the central nervous system (CNS). Id2 contributes to cultured neural precursor cell (NPC) proliferation as well as to the proliferation of CNS tumors such as glioblastoma that are likely to arise from NPC-like cells. We identified three phosphorylation sites near the N-terminus of Id2 in NPCs. To interrogate the importance of Id2 phosphorylation, Id2(-/-) NPCs were modified to express wild type (WT) Id2 or an Id2 mutant protein that could not be phosphorylated at the identified sites. We observed that NPCs expressing this mutant lacking phosphorylation near the N-terminus had higher steady-state levels of Id2 when compared to NPCs expressing WT Id2. This elevated level was the result of a longer half-life and reduced proteasome-mediated degradation. Moreover, NPCs expressing constitutively de-phosphorylated Id2 proliferated more rapidly than NPCs expressing WT Id2, a finding consistent with the well-characterized function of Id2 in driving proliferation. Observing that phosphorylation of Id2 modulates the degradation of this important cell-cycle regulator, we sought to identify a phosphatase that would stabilize Id2 enhancing its activity in NPCs and extended our analysis to include human glioblastoma-derived stem cells (GSCs). We found that expression of the phosphatase PP2A altered Id2 levels. Our findings suggest that inhibition of PP2A may be a novel strategy to regulate the proliferation of normal NPCs and malignant GSCs by decreasing Id2 levels. Stem Cells 2016;34:1321-1331. PMID:26756672

  1. Wnt5a-mediated non-canonical Wnt signalling regulates human endothelial cell proliferation and migration

    SciTech Connect

    Cheng Chingwen Yeh Juching; Fan Taiping; Smith, Stephen K.; Charnock-Jones, D. Stephen

    2008-01-11

    Cell to cell interaction is one of the key processes effecting angiogenesis and endothelial cell function. Wnt signalling is mediated through cell-cell interaction and is involved in many developmental processes and cellular functions. In this study, we investigated the possible function of Wnt5a and the non-canonical Wnt pathway in human endothelial cells. We found that Wnt5a-mediated non-canonical Wnt signalling regulated endothelial cell proliferation. Blocking this pathway using antibody, siRNA or a down-stream inhibitor led to suppression of endothelial cell proliferation, migration, and monolayer wound closure. We also found that the mRNA level of Wnt5a is up-regulated when endothelial cells are treated with a cocktail of inflammatory cytokines. Our findings suggest non-canonical Wnt signalling plays a role in regulating endothelial cell growth and possibly in angiogenesis.

  2. The effects of childhood abuse on symptom complexity in a clinical sample: mediating effects of emotion regulation difficulties.

    PubMed

    Choi, Ji Young; Choi, Young Min; Gim, Min Sook; Park, Jun Hyun; Park, Soo Hyun

    2014-08-01

    The purpose of the present study was to first examine whether childhood abuse predicts symptom complexity, as indicated by the number of clinically elevated scales on the MMPI-2 in an adult clinical sample. Secondly, we investigated whether emotion regulation difficulties mediated the relationship between childhood abuse and symptom complexity. A total of 162 adult outpatients not presenting with psychotic symptoms completed the Korean Childhood Trauma Questionnaire (K-CTQ), Life Events Checklist (LEC), Difficulties in Emotion Regulation Scale (DERS), and Korean Minnesota Multiphasic Personality Inventory-2 (MMPI-2). Partial correlation analysis results indicated that after controlling for the presence of adulthood trauma, childhood abuse was associated with more symptom complexity, or more clinically elevated scales on the MMPI-2. Furthermore, structural equation modeling results showed that emotion regulation difficulties partially mediated the relationship between childhood abuse and symptom complexity. These findings indicate that individuals who had experienced childhood abuse evidence simultaneous presentation of diverse clinical symptoms. PMID:24841063

  3. Confirming resistance in bottle gourd germplasm by quantifying powdery mildew conidia using a cellometer

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew (PM) caused by Podosphaera xanthii, an important foliar disease affecting cucurbit crops grown in the United States, commonly occurs on foliage, petioles, and stems. We have developed two highly resistant bottle gourd (Lagenaria siceraria) germplasm (USVL351 and USVL482) for use in o...

  4. Transcriptomes that confer to plant defense against powdery mildew disease in Lagerstroemia indica

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genus Lagerstroemia (common name crape myrtle) is a deciduous shrub or small tree, originaly native to southeast Asia and has became one of the most distinctive and popular summer flowering woody ornamentals anywhere south of USDA from east to west coast. Powdery mildew was one of the most serio...

  5. Mechanisms of resistance to an azole fungicide in the grapevine powdery mildew fungus, Erysiphe necator

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We studied the mechanisms of azole resistance in the grapevine powdery mildew fungus, Erysiphe necator, by quantifying the sensitivity to myclobutanil (EC50) in 65 isolates from the eastern U.S. and 12 from Chile. From each isolate, we sequenced the gene for sterol 14a-demethylase (CYP51), and measu...

  6. The perfect stage of powdery mildew of Beta vulgaris found in Michigan

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew (Erysiphe polygoni [synonym E. betae]) affects several different crops in the genus Beta, including sugar beet, Swiss chard, and table beet. The disease has been prevalent in many sugar beet growing areas of the United States since the first major epidemic on beet in the United States...

  7. Tolerance to Cucurbit Powdery Mildew in USDA Bottle Gourd (Lagenaria siceraria) Plant Introductions (PI)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew (Podosphaera xanthii) can cause severe damage to cucurbit crops grown in open fields and greenhouses. Bottle gourd plants (Lagenaria siceraria) are being used throughout the world as rootstocks for grafting watermelon. In recent years, there has been a growing interest in the USA in...

  8. Golovinomyces spadaceus causing powdery mildew on Coreopsis hybrid 'Full Moon' (Heliantheae, Asteraceae) in Washington State

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Symptoms of powdery mildew were observed on a Coreopsis cultivar in the Horticulture and Landscape Architecture Garden on the Washington State University campus, Pullman, Whitman County, Washington. White to off-white sporulating mycelial areas were ~5mm in diam to confluent and confined to adaxial...

  9. Evaluation of watermelon and related species for resistance to race 1W powdery mildew resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Race 1 and race 2 powdery mildew (Podosphaera xanthii) has been reported on watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) in the United States for six years. In this study, the entire available U.S. Plant Introduction collection of Citrullus sp. was evaluated for resistance to P. xanthii r...

  10. Strategies for durable resistance to the grapevine powdery mildew fungus, Erysiphe necator

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nearly all cultivars of Vitis vinifera are highly susceptible to the grapevine powdery mildew fungus, Erysiphe necator. Grape breeders around the world are working to introgress resistance from wild Vitis. Of the widely-used introgressions, most involve dominant, race-specific resistance phenotype...

  11. VitisGenPM: a precision phenotyping center for powdery mildew resistance breeding in grapevine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Analyzing resistance to powdery mildew (Erysiphe necator) can be complicated by race specificity, environment, phenology, and other factors. Of necessity, breeding programs often rely upon natural infection in greenhouses, nurseries, and/or vineyards and use categorical ratings to assess resistance ...

  12. Hairy nightshade is an alternative host of spongospora subterranea, the potato powdery scab pathogen

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Structures, similar to galls developing on potato roots infected by the powdery scab pathogen S. subterranea, were observed on roots of hairy nightshades (Solanum physalifolium, formerly S. sarrachoides) collected from commercial potato fields in Washington State where populations of S. subterranea ...

  13. PROPOSED MAJOR POWDERY MILDEW GENES IN EASTERN AND SOUTHERN WHEAT GERMPLASM; IN THE PAST TEN YEARS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Resistance to powdery mildew of wheat is controlled by major genes in adult and seedling stages or quantitative resistance in the adult stage. Entries in the PM nursery from Southern and Eastern US were screened on detached seedling leaves. The observed virulence patterns were then compared to virul...

  14. Suppression of cucumber powdery mildew by UV-B is affected by background light quality

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Brief (5-10 min) exposure to UV-B radiation (280-300 nm) suppressed powdery mildew (Podosphaera xanthii) on Cucumis sativus. The effect was enhanced by red light (600-660 nm), but offset by blue light (420-500 nm) and UV-A (300-420 nm). Compared to untreated controls, 2 h red light from specific lig...

  15. Identifying resistance to powdery mildew race 2W in the USDA-ARS watermelon germplasm collection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew caused by Podosphaera xanthii has become a common disease of watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai] in the United States. The disease can be controlled with fungicides. However, it is more economical and environmentally safe to use genetic resistance against this di...

  16. Molecular characterization of a new powdery mildew resistance gene Pm54 in soft red winter wheat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew has caused increasing damage to wheat production in the southeastern USA. To combat the disease, there is a continuing need to discover new genes or quantitative trait loci for mildew resistance and promptly adopt those loci in breeding programs. Pioneer® variety 26R61 (shortened as 2...

  17. FIRST REPORT OF POWDERY MILDEW ON DIPSACUS SYLVESTRIS CAUSED BY SPHAEROTHECA DIPSACEARUM IN NORTH AMERICA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Dipsacus sylvestris (syn. D. fullonum, common teasel) (Dipsacaceae) is a European species introduced into North America, and now widely established and regarded as a noxious weed. Powdery mildew fungi (Erysiphales) reported previously from this host included Phyllactinia species in Washington State...

  18. Suppression of Zinnia Powdery Mildew in the Greenhouse with Silicon-containing Media Amendments

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We have previously reported that supplemental silicon supplied in hydroponic solution or soilless media can reduce the incidence and severity of powdery mildew (PM) on zinnia. This research reports the use of 1) silicon-containing organic amendments, 2) mineral compositions high in silicon content,...

  19. Improving management of grape powdery mildew with new tools and knowledge

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The assumption that inoculum of the grape powdery mildew pathogen is always available once conditions are suitable for inoculum release has been shown to be incorrect. Using various molecular techniques, we have shown that viticulturist can reduce their fungicide applications, on average, by 2.4 ap...

  20. First report of Oidiopsis taurica causing powdery mildew outbreak on pepper in Maryland

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pepper plants grown in large experimental plots at Beltsville Maryland showed widespread powdery mildew infection in the late summer of 2008. Extensive coverage of the abaxial surface by white patches of conidia was noted, along with chlorotic regions on the adaxial surface. Samples were taken for ...

  1. QTL Mapping of Powdery Mildew Susceptibility in Hop (Humulus lupulus L)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Hop powdery mildew is best controlled via the production of resistant varieties. Recent evidence supports selection for absence of susceptibility to fungal pathogens as a more durable resistance mechanism. The objective of this study was to identify molecular markers associated with susceptibility...

  2. Taxonomic complexity of powdery mildew pathogens found on lentil and pea in the US Pacific Northwest

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Classification of powdery mildews found on lentil and pea in greenhouse and field production conditions in the US Pacific Northwest was investigated using morphological and molecular characters. Isolates collected from lentil plants grown in the greenhouse or field displayed morphologies in substant...

  3. Tolerance to powdery mildew conferred in susceptible watermelon scion by grafting on resistant rootstocks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cucurbit powdery mildew (PM) caused by Podosphaera xanthii, can impact seedling growth and cause serious losses in greenhouse and open field production. We have developed several watermelon and bottle gourd germplasm lines with high levels of resistance to PM. A PM susceptible cultivar Mickey Lee ...

  4. Cucurbit powdery mildews: methodology for objective determination and denomination of races

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cucurbit powdery mildew (CPM), a disease on field and greenhouse cucurbit crops worldwide, is caused most frequently by two obligate erysiphaceous ectoparasites (Golovinomyces orontii s.l., Podosphaera xanthii) that are highly variable in their pathogenicity and virulence. Various, independent syste...

  5. Resistance to Root Galling Caused by the Powdery Scab Pathogen Spongospora subterranea in Potato

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Potato selections (clones and commercial cultivars) were examined for resistance to root galling, caused by the powdery scab pathogen Spongospora subterranea in 7 field trials conducted between 2003 and 2007 in the states of Washington (WA) and Idaho (ID). In 2003, Shepody demonstrated the highest l...

  6. Specific isolation of RNA from the grape powdery mildew pathogen Erysiphe necator, an epiphytic, obligate parasite

    Technology Transfer Automated Retrieval System (TEKTRAN)

    RNA expression profiling of obligately parasitic plant microbes is hampered by the requisite interaction of host and parasite. For superficial pathogens like grape powdery mildew as well as for epiphytic saprophytes, growth along the outside surface of the plant allows separation from the host and ...

  7. Outbreak of powdery mildew on watermelon fruit caused by podosphera xanthii in southwest Florida

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew caused by the obligate parasite Podosphera xanthii occurs commonly on foliage, petioles and stems of most cucurbit vegetable crops grown in the U.S. (1). However, in the field, fruit infection on cucurbits including watermelon (Citrullus lanatus), is rarely, if ever, observed. Consequ...

  8. Genetic mapping of the powdery mildew resistance gene in soybean PI 567301B

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Powdery mildew (PMD) of soybean [Glycine max (L.) Merr.] is caused by the fungus Microsphaera diffusa. Severe infection of PMD on susceptible varieties often cause premature defoliation and chlorosis of the leaves, which can result in considerable yield losses under favorable environmental condition...

  9. Registration of ‘Wyandot-14’ soybean with resistance to soybean aphid and powdery mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    ‘Wyandot-14’ soybean [Glycine max (L.) Merr.] with resistance to soybean aphid biotypes 1 and 2 and resistance to powdery mildew was jointly released by the USDA-Agricultural Research Service and The Ohio Agricultural Research and Development Center (OARDC) as a late maturity group (MG) II (2.9) foo...

  10. Development of a grower performed inoculum detection assay for management of grape powdery mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Management of grape powdery mildew (Erysiphe necator) and other polycyclic diseases relies on calendar-based pesticide application schedules. However, these schedules may be adjusted to optimize and often reduce the number of applications by utilizing information generated from infection risk foreca...

  11. First report of powdery mildew caused by Podosphaera leucotricha on Callery pear in North America

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Podosphaera leucotricha (Ellis & Everh.) E.S. Salmon (Ascomycetes, Erysiphales) is the etiological agent of a powdery mildew disease that occurs on rosaceous plants, primarily Malus and Pyrus. This fungus is nearly circumglobal. In May 2009, leaves of Bradford pear (Pyrus calleryana Decne.), some di...

  12. Sequential sampling for estimation and classification of the incidence of hop powdery mildew II: Cone sampling

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sequential sampling models for estimation and classification of the incidence of powdery mildew (caused by Podosphaera macularis) on hop (Humulus lupulus) cones were developed using parameter estimates of the binary power law derived from the analysis of 221 transect data sets (model construction da...

  13. Resistance to a New Race of the Cucurbit Powdery Mildew Present in Arizona and California

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A new strain of the powdery mildew fungus Podosphaera xanthii, designated race S, was first detected on melon (Cucumis melo L.) in the lower desert areas of Yuma, Ariz. and Imperial Valley, Calif. in 2003 when it caused economic losses in some fields. Race S overcomes resistance in 19 reported melon...

  14. Grapevine powdery mildew resistance and susceptibility loci identified on a high-resolution SNP map

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Improved efficacy and durability of powdery mildew resistance requires knowledge of the genetics of resistance and susceptibility and high resolution maps to facilitate combining of multiple resistance genes with desirable traits. In the current study, we studied the inheritance of resistance and s...

  15. Genetic dissection of resistance to anthracnose and powdery mildew in Medicago truncatula

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Medicago truncatula was used to characterize resistance to anthracnose and powdery mildew, respectively caused by Colletotrichum trifolii and Erysiphe pisi. Two isolates of E. pisi (Ep-p from pea and Ep-a from alfalfa) and two races of C. trifolii (races 1 and 2) were used in this study. The A17 gen...

  16. Cyclic AMP Represents a Crucial Component of Treg Cell-Mediated Immune Regulation.

    PubMed

    Klein, Matthias; Bopp, Tobias

    2016-01-01

    T regulatory (Treg) cells are one of the key players in the immune tolerance network, and a plethora of manuscripts have described their development and function in the course of the last two decades. Nevertheless, it is still a matter of debate as to which mechanisms and agents are employed by Treg cells, providing the basis of their suppressive potency. One of the important candidates is cyclic AMP (cAMP), which is long known as a potent suppressor at least of T cell activation and function. While this suppressive function by itself is widely accepted, the source and the mechanism of action of cAMP are less clear, and a multitude of seemingly contradictory data allow for, in principle, two different scenarios of cAMP-mediated suppression. In one scenario, Treg cells contain high amounts of cAMP and convey this small molecule via gap junction intercellular communication directly to the effector T cells (Teff) leading to their suppression. Alternatively, it was shown that Treg cells represent the origin of considerable amounts of adenosine, which trigger the adenylate cyclases in Teff cells via A2A and A2B receptors, thus strongly increasing intracellular cAMP. This review will present and discuss initial findings and recent developments concerning the function of cAMP for Treg cells and its impact on immune regulation. PMID:27621729

  17. Rab GTPase regulation of retromer-mediated cargo export during endosome maturation

    PubMed Central

    Liu, Ting-Ting; Gomez, Timothy S.; Sackey, Bridget K.; Billadeau, Daniel D.; Burd, Christopher G.

    2012-01-01

    The retromer complex, composed of sorting nexin subunits and a Vps26/Vps29/Vps35 trimer, mediates sorting of retrograde cargo from the endosome to the trans-Golgi network. The retromer trimer subcomplex is an effector of Rab7 (Ypt7 in yeast). Whereas endosome targeting of human retromer has been shown to require Rab7-GTP, targeting of yeast retromer to the endosome is independent of Ypt7-GTP and requires the Vps5 and Vps17 retromer sorting nexin subunits. An evolutionarily conserved amino acid segment within Vps35 is required for Ypt7/Rab7 recognition in vivo by both yeast and human retromer, establishing that Rab recognition is a conserved feature of this subunit. Recognition of Ypt7 by retromer is required for its function in retrograde sorting, and in yeast cells lacking the guanine nucleotide exchange factor for Ypt7, retrograde cargo accumulates in endosomes that are decorated with retromer, revealing an additional role for Rab recognition at the cargo export stage of the retromer functional cycle. In addition, yeast retromer trimer antagonizes Ypt7-regulated organelle tethering and fusion of endosomes/vacuoles via recognition of Ypt7. Thus retromer has dual roles in retrograde cargo export and in controlling the fusion dynamics of the late endovacuolar system. PMID:22593205

  18. Mechanical regulation of transcription controls Polycomb-mediated gene silencing during lineage commitment.

    PubMed

    Le, Huy Quang; Ghatak, Sushmita; Yeung, Ching-Yan Chloé; Tellkamp, Frederik; Günschmann, Christian; Dieterich, Christoph; Yeroslaviz, Assa; Habermann, Bianca; Pombo, Ana; Niessen, Carien M; Wickström, Sara A

    2016-08-01

    Tissue mechanics drive morphogenesis, but how forces are sensed and transmitted to control stem cell fate and self-organization remains unclear. We show that a mechanosensory complex of emerin (Emd), non-muscle myosin IIA (NMIIA) and actin controls gene silencing and chromatin compaction, thereby regulating lineage commitment. Force-driven enrichment of Emd at the outer nuclear membrane of epidermal stem cells leads to defective heterochromatin anchoring to the nuclear lamina and a switch from H3K9me2,3 to H3K27me3 occupancy at constitutive heterochromatin. Emd enrichment is accompanied by the recruitment of NMIIA to promote local actin polymerization that reduces nuclear actin levels, resulting in attenuation of transcription and subsequent accumulation of H3K27me3 at facultative heterochromatin. Perturbing this mechanosensory pathway by deleting NMIIA in mouse epidermis leads to attenuated H3K27me3-mediated silencing and precocious lineage commitment, abrogating morphogenesis. Our results reveal how mechanics integrate nuclear architecture and chromatin organization to control lineage commitment and tissue morphogenesis. PMID:27398909

  19. Mechanisms of leiomodin 2-mediated regulation of actin filament in muscle cells

    PubMed Central

    Chen, Xiaorui; Ni, Fengyun; Kondrashkina, Elena; Ma, Jianpeng; Wang, Qinghua

    2015-01-01

    Leiomodin (Lmod) is a class of potent tandem-G-actin–binding nucleators in muscle cells. Lmod mutations, deletion, or instability are linked to lethal nemaline myopathy. However, the lack of high-resolution structures of Lmod nucleators in action severely hampered our understanding of their essential cellular functions. Here we report the crystal structure of the actin–Lmod2162–495 nucleus. The structure contains two actin subunits connected by one Lmod2162–495 molecule in a non–filament-like conformation. Complementary functional studies suggest that the binding of Lmod2 stimulates ATP hydrolysis and accelerates actin nucleation and polymerization. The high level of conservation among Lmod proteins in sequence and functions suggests that the mechanistic insights of human Lmod2 uncovered here may aid in a molecular understanding of other Lmod proteins. Furthermore, our structural and mechanistic studies unraveled a previously unrecognized level of regulation in mammalian signal transduction mediated by certain tandem-G-actin–binding nucleators. PMID:26417072

  20. Photoperiodic regulation of testis function in rats: mediation by a circadian mechanism

    SciTech Connect

    Nelson, R.J.; Bamat, M.K.; Zucker, I.

    1982-05-01

    Laboratory rats traditionally are classified as nonphotoperiodic because variations in daylength have little or no effect on their gonadal function. After olfactory bulbectomy, however, rats show clear evidence of photoperiodic regulation of the gonads. The present study demonstrates, by means of resonance experiments, that the testicular response to daylength in rats is mediated by a circadian photoperiodic time measurement system similar to that of photoperiodic rodents. Olfactory-bulbectomized rats were maintained in fixed photoperiods in which a 6 h light period was coupled with dark periods of 18, 30, 42, or 54 h; a fifth group was maintained in a 45L:10D photoperiod. Rats from the 6L:30D, 6L:54D and 14L:10D photoperiods had testes and seminal vesicle weights, plasma testosterone titers and spermatogenesis indices indicative of functional reproductive status. Rats exposed to the 6L:18D and 6L:42D photoperiods had reduced testicular and seminal vesicle weights, lower testosterone levels and reduced spermatogenesis. Researchers hypothesize that photo- and nonphotoperiodic rodent species use similar mechanisms for distinguishing long from short photoperiods, but differ in the extent to which discrimination of short daylengths is transduced into altered gonadal activity.

  1. Hypothalamus-Pituitary-Adrenal cell-mediated immunity regulation in the Immune Restoration Inflammatory Syndrome

    PubMed Central

    Khakshooy, Allen; Chiappelli, Francesco

    2016-01-01

    Over one third of the patients sero-positive for the human immunodeficiency virus (HIV) with signs of the acquired immune deficiency syndrome (AIDS), and under treatment with anti-retroviral therapy (ART), develop the immune reconstitution inflammatory syndrome (IRIS). It is not clear what variables are that determine whether a patient with HIV/AIDS will develop ART-related IRIS, but the best evidence base thus far indicates that HIV/AIDS patients with low CD4 cell count, and HIV/AIDS patients whose CD4 count recovery shows a sharp slope, suggesting a particularly fast "immune reconstitution", are at greater risk of developing IRIS. Here, we propose the hypothesis that one important variable that can contribute to low CD4 cell count number and function in ART-treated HIV/AIDS patients is altered hypothalamic-pituitary-adrenal (HPA) cell-mediated immune (CMI) regulation. We discuss HPA-CMI deregulation in IRIS as the new frontier in comparative effectiveness research (CRE) for obtaining and utilizing the best evidence base for treatment of patients with HIV/AIDS in specific clinical settings. We propose that our hypothesis about altered HPA-CMI may extend to the pathologies observed in related viral infection, including Zika PMID:27212842

  2. Role of SWI/SNF in acute leukemia maintenance and enhancer-mediated Myc regulation

    PubMed Central

    Shi, Junwei; Whyte, Warren A.; Zepeda-Mendoza, Cinthya J.; Milazzo, Joseph P.; Shen, Chen; Roe, Jae-Seok; Minder, Jessica L.; Mercan, Fatih; Wang, Eric; Eckersley-Maslin, Melanie A.; Campbell, Amy E.; Kawaoka, Shinpei; Shareef, Sarah; Zhu, Zhu; Kendall, Jude; Muhar, Matthias; Haslinger, Christian; Yu, Ming; Roeder, Robert G.; Wigler, Michael H.; Blobel, Gerd A.; Zuber, Johannes; Spector, David L.; Young, Richard A.; Vakoc, Christopher R.

    2013-01-01

    Cancer cells frequently depend on chromatin regulatory activities to maintain a malignant phenotype. Here, we show that leukemia cells require the mammalian SWI/SNF chromatin remodeling complex for their survival and aberrant self-renewal potential. While Brg1, an ATPase subunit of SWI/SNF, is known to suppress tumor formation in several cell types, we found that leukemia cells instead rely on Brg1 to support their oncogenic transcriptional program, which includes Myc as one of its key targets. To account for this context-specific function, we identify a cluster of lineage-specific enhancers located 1.7 Mb downstream from Myc that are occupied by SWI/SNF as well as the BET protein Brd4. Brg1 is required at these distal elements to maintain transcription factor occupancy and for long-range chromatin looping interactions with the Myc promoter. Notably, these distal Myc enhancers coincide with a region that is focally amplified in ∼3% of acute myeloid leukemias. Together, these findings define a leukemia maintenance function for SWI/SNF that is linked to enhancer-mediated gene regulation, providing general insights into how cancer cells exploit transcriptional coactivators to maintain oncogenic gene expression programs. PMID:24285714

  3. Signal transduction pathways mediating parathyroid hormone regulation of osteoblastic gene expression

    NASA Technical Reports Server (NTRS)

    Partridge, N. C.; Bloch, S. R.; Pearman, A. T.

    1994-01-01

    Parathyroid hormone (PTH) plays a central role in regulation of calcium metabolism. For example, excessive or inappropriate production of PTH or the related hormone, parathyroid hormone related protein (PTHrP), accounts for the majority of the causes of hypercalcemia. Both hormones act through the same receptor on the osteoblast to elicit enhanced bone resorption by the osteoclast. Thus, the osteoblast mediates the effect of PTH in the resorption process. In this process, PTH causes a change in the function and phenotype of the osteoblast from a cell involved in bone formation to one directing the process of bone resorption. In response to PTH, the osteoblast decreases collagen, alkaline phosphatase, and osteopontin expression and increases production of osteocalcin, cytokines, and neutral proteases. Many of these changes have been shown to be due to effects on mRNA abundance through either transcriptional or post-transcriptional mechanisms. However, the signal transduction pathway for the hormone to cause these changes is not completely elucidated in any case. Binding of PTH and PTHrP to their common receptor has been shown to result in activation of protein kinases A and C and increases in intracellular calcium. The latter has not been implicated in any changes in mRNA of osteoblastic genes. On the other hand activation of PKA can mimic all the effects of PTH; protein kinase C may be involved in some responses. We will discuss possible mechanisms linking PKA and PKC activation to changes in gene expression, particularly at the nuclear level.

  4. A novel TRIM family member, Trim69, regulates zebrafish development through p53-mediated apoptosis.

    PubMed

    Han, Ruiqin; Zhao, Qing; Zong, Shudong; Miao, Shiying; Song, Wei; Wang, Linfang

    2016-05-01

    Trim69 contains the hallmark domains of a tripartite motif (TRIM) protein, including a Ring-finger domain, B-box domain, and coiled-coil domain. Trim69 is structurally and evolutionarily conserved in zebrafish, mouse, rat, human, and chimpanzee. The role of this protein is unclear, however, so we investigated its function in zebrafish development. Trim69 is extensively expressed in zebrafish adults and developing embryos-particularly in the testis, brain, ovary, and heart-and its expression decreases in a time- and stage-dependent manner. Loss of trim69 in zebrafish induces apoptosis and activates apoptosis-related processes; indeed, the tp53 pathway was up-regulated in response to the knockdown. Expression of human trim69 rescued the apoptotic phenotype, while overexpression of trim69 does not increase cellular apoptosis. Taken together, our results suggest that trim69 participates in tp53-mediated apoptosis during zebrafish development. Mol. Reprod. Dev. 83: 442-454, 2016. © 2016 Wiley Periodicals, Inc. PMID:27031046

  5. Peripheral Brain Derived Neurotrophic Factor Precursor Regulates Pain as an Inflammatory Mediator.

    PubMed

    Luo, Cong; Zhong, Xiao-Lin; Zhou, Fiona H; Li, Jia-Yi; Zhou, Pei; Xu, Jun-Mei; Song, Bo; Li, Chang-Qi; Zhou, Xin-Fu; Dai, Ru-Ping

    2016-01-01

    The precursor of brain derived neurotrophic factor (proBDNF), the unprocessed BDNF gene product, binds to its receptors and exerts the opposing biologic functions of mature BDNF. proBDNF is expressed in the peripheral tissues but the functions of peripheral proBDNF remain elusive. Here we showed that proBDNF and its predominant receptor, p75 pan-neurotrophin receptor were upregulated in the nerve fibers and inflammatory cells in the local tissue in inflammatory pain. Neutralization of proBDNF by polyclonal antibody attenuated pain in different models of inflammatory pain. Unilateral intra-plantar supplementation of proBDNF by injecting exogenous proBDNF or ectopic overexpression resulted in pain hypersensitivity and induced spinal phosphorylated extracellular signal-regulated kinase activation. Exogenous proBDNF injection induced the infiltration of inflammatory cells and the activation of proinflammatory cytokines, suggesting that inflammatory reaction contributed to the pro-algesic effect of proBDNF. Finally, we generated monoclonal anti-proBDNF antibody that could biologically block proBDNF. Administration of monoclonal Ab-proBDNF attenuated various types of inflammatory pain and surgical pain. Thus, peripheral proBDNF is a potential pain mediator and anti-proBDNF pretreatment may alleviate the development of inflammatory pain. PMID:27251195

  6. Ku proteins function as corepressors to regulate farnesoid X receptor-mediated gene expression

    SciTech Connect

    Ohno, Masae; Kunimoto, Masaaki; Nishizuka, Makoto; Osada, Shigehiro; Imagawa, Masayoshi

    2009-12-18

    The farnesoid X receptor (FXR; NR1H4) is a member of the nuclear receptor superfamily and regulates the expression of genes involved in enterohepatic circulation and the metabolism of bile acids. Based on functional analyses, nuclear receptors are divided into regions A-F. To explore the cofactors interacting with FXR, we performed a pull-down assay using GST-fused to the N-terminal A/B region and the C region, which are required for the ligand-independent transactivation and DNA-binding, respectively, of FXR, and nuclear extracts from HeLa cells. We identified DNA-dependent protein kinase catalytic subunit (DNA-PKcs), Ku80, and Ku70 as FXR associated factors. These proteins are known to have an important role in DNA repair, recombination, and transcription. DNA-PKcs mainly interacted with the A/B region of FXR, whereas the Ku proteins interacted with the C region and with the D region (hinge region). Chromatin immunoprecipitation assays revealed that the Ku proteins associated with FXR on the bile salt export pump (BSEP) promoter. Furthermore, we demonstrated that ectopic expression of the Ku proteins decreased the promoter activity and expression of BSEP gene mediated by FXR. These results suggest that the Ku proteins function as corepressors for FXR.

  7. MicroRNA-mediated gene regulation: potential applications for plant genetic engineering.

    PubMed

    Zhou, Man; Luo, Hong

    2013-09-01

    Food security is one of the most important issues challenging the world today. Any strategies to solve this problem must include increasing crop yields and quality. MicroRNA-based genetic modification technology (miRNA-based GM tech) can be one of the most promising solutions that contribute to agricultural productivity directly by developing superior crop cultivars with enhanced biotic and abiotic stress tolerance and increased biomass yields. Indirectly, the technology may increase usage of marginal soils and decrease pesticide use, among other benefits. This review highlights the most recent progress of transgenic studies utilizing various miRNAs and their targets for plant trait modifications, and analyzes the potential of miRNA-mediated gene regulation for use in crop improvement. Strategies for manipulating miRNAs and their targets in transgenic plants including constitutive, stress-induced, or tissue-specific expression of miRNAs or their targets, RNA interference, expressing miRNA-resistant target genes, artificial target mimic and artificial miRNAs were discussed. We also discussed potential risks of utilizing miRNA-based GM tech. In general, miRNAs and their targets not only provide an invaluable source of novel transgenes, but also inspire the development of several new GM strategies, allowing advances in breeding novel crop cultivars with agronomically useful characteristics. PMID:23771582

  8. Arginase 2 deficiency reduces hyperoxia-mediated retinal neurodegeneration through the regulation of polyamine metabolism.

    PubMed

    Narayanan, S P; Xu, Z; Putluri, N; Sreekumar, A; Lemtalsi, T; Caldwell, R W; Caldwell, R B

    2014-01-01

    Hyperoxia treatment has been known to induce neuronal and glial death in the developing central nervous system. Retinopathy of prematurity (ROP) is a devastating disease in premature infants and a major cause of childhood vision impairment. Studies indicate that, in addition to vascular injury, retinal neurons are also affected in ROP. Using an oxygen-induced retinopathy (OIR) mouse model for ROP, we have previously shown that deletion of the arginase 2 (A2) significantly reduced neuro-glial injury and improved retinal function. In the current study, we investigated the mechanism of A2 deficiency-mediated neuroprotection in the OIR retina. Hyperoxia treatment has been known to induce neuronal death in neonates. During the hyperoxia phase of OIR, a significant increase in the number of apoptotic cells was observed in the wild-type (WT) OIR retina compared with A2-deficient OIR. Mass spectrometric analysis showed alterations in polyamine metabolism in WT OIR retina. Further, increased expression level of spermine oxidase was observed in WT OIR retina, suggesting increased oxidation of polyamines in OIR retina. These changes were minimal in A2-deficient OIR retina. Treatment using the polyamine oxidase inhibitor, N, N'-bis (2, 3-butadienyl)-1, 4-butanediamine dihydrochloride, significantly improved neuronal survival during OIR treatment. Our data suggest that retinal arginase is involved in the hyperoxia-induced neuronal degeneration in the OIR model, through the regulation of polyamine metabolism. PMID:24556690

  9. Pan1 regulates transitions between stages of clathrin-mediated endocytosis

    PubMed Central

    Bradford, Mary Katherine; Whitworth, Karen; Wendland, Beverly

    2015-01-01

    Endocytosis is a well-conserved process by which cells invaginate small portions of the plasma membrane to create vesicles containing extracellular and transmembrane cargo proteins. Dozens of proteins and hundreds of specific binding interactions are needed to coordinate and regulate these events. Saccharomyces cerevisiae is a powerful model system with which to study clathrin-mediated endocytosis (CME). Pan1 is believed to be a scaffolding protein due to its interactions with numerous proteins that act throughout the endocytic process. Previous research characterized many Pan1 binding interactions, but due to Pan1's essential nature, the exact mechanisms of Pan1's function in endocytosis have been difficult to define. We created a novel Pan1-degron allele, Pan1-AID, in which Pan1 can be specifically and efficiently degraded in <1 h upon addition of the plant hormone auxin. The loss of Pan1 caused a delay in endocytic progression and weakened connections between the coat/actin machinery and the membrane, leading to arrest in CME. In addition, we determined a critical role for the central region of Pan1 in endocytosis and viability. The regions important for endocytosis and viability can be separated, suggesting that Pan1 may have a distinct role in the cell that is essential for viability. PMID:25631817

  10. Phosphorylation of Src by phosphoinositide 3-kinase regulates beta-adrenergic receptor-mediated EGFR transactivation.

    PubMed

    Watson, Lewis J; Alexander, Kevin M; Mohan, Maradumane L; Bowman, Amber L; Mangmool, Supachoke; Xiao, Kunhong; Naga Prasad, Sathyamangla V; Rockman, Howard A

    2016-10-01

    β2-Adrenergic receptors (β2AR) transactivate epidermal growth factor receptors (EGFR) through formation of a β2AR-EGFR complex that requires activation of Src to mediate signaling. Here, we show that both lipid and protein kinase activities of the bifunctional phosphoinositide 3-kinase (PI3K) enzyme are required for β2AR-stimulated EGFR transactivation. Mechanistically, the generation of phosphatidylinositol (3,4,5)-tris-phosphate (PIP3) by the lipid kinase function stabilizes β2AR-EGFR complexes while the protein kinase activity of PI3K regulates Src activation by direct phosphorylation. The protein kinase activity of PI3K phosphorylates serine residue 70 on Src to enhance its activity and induce EGFR transactivation following βAR stimulation. This newly identified function for PI3K, whereby Src is a substrate for the protein kinase activity of PI3K, is of importance since Src plays a key role in pathological and physiological signaling. PMID:27169346

  11. Synaptonuclear messenger PRR7 inhibits c-Jun ubiquitination and regulates NMDA-mediated excitotoxicity.

    PubMed

    Kravchick, Dana O; Karpova, Anna; Hrdinka, Matous; Lopez-Rojas, Jeffrey; Iacobas, Sanda; Carbonell, Abigail U; Iacobas, Dumitru A; Kreutz, Michael R; Jordan, Bryen A

    2016-09-01

    Elevated c-Jun levels result in apoptosis and are evident in neurodegenerative disorders such as Alzheimer's disease and dementia and after global cerebral insults including stroke and epilepsy. NMDA receptor (NMDAR) antagonists block c-Jun upregulation and prevent neuronal cell death following excitotoxic insults. However, the molecular mechanisms regulating c-Jun abundance in neurons are poorly understood. Here, we show that the synaptic component Proline rich 7 (PRR7) accumulates in the nucleus of hippocampal neurons following NMDAR activity. We find that PRR7 inhibits the ubiquitination of c-Jun by E3 ligase SCF(FBW) (7) (FBW7), increases c-Jun-dependent transcriptional activity, and promotes neuronal death. Microarray assays show that PRR7 abundance is directly correlated with transcripts associated with cellular viability. Moreover, PRR7 knockdown attenuates NMDAR-mediated excitotoxicity in neuronal cultures in a c-Jun-dependent manner. Our results show that PRR7 links NMDAR activity to c-Jun function and provide new insights into the molecular processes that underlie NMDAR-dependent excitotoxicity. PMID:27458189

  12. Rab5-mediated endosome-endosome fusion regulates hemoglobin endocytosis in Leishmania donovani.

    PubMed

    Singh, Sudha B; Tandon, Ruchi; Krishnamurthy, Ganga; Vikram, Rajagopal; Sharma, Nimisha; Basu, Sandip K; Mukhopadhyay, Amitabha

    2003-11-01

    To understand the trafficking of endocytosed hemoglobin (Hb) in Leishmania, we investigated the characteristics of in vitro fusion between endosomes containing biotinylated Hb (BHb) and avidin-horseradish peroxidase (AHRP). We showed that early endosome fusion in Leishmania is temperature and cytosol dependent and is inhibited by ATP depletion, ATPgammaS, GTPgammaS and N-ethylmaleimide treatment. The Rab5 homolog from Leishmania donovani, LdRab5, was cloned and expressed. Our results showed that homotypic fusion between the early endosomes in Leishmania is Rab5 dependent. Early endosomes containing BHb fused efficiently with late endosomes in a process regulated by Rab7, whereas no fusion between early and late endosomes was detected using fluid phase markers. Pre-treatment of early endosomes containing BHb with monoclonal antibody specific for the C-terminus of the Hb receptor (HbR) or the addition of the C-terminal cytoplasmic fragment of the HbR specifically inhibited the fusion with late endosomes, suggesting that signal(s) mediated through the HbR cytoplasmic tail promotes the fusion of early endosomes containing Hb with late endosomes. PMID:14592970

  13. Cyclic AMP Represents a Crucial Component of Treg Cell-Mediated Immune Regulation

    PubMed Central

    Klein, Matthias; Bopp, Tobias

    2016-01-01

    T regulatory (Treg) cells are one of the key players in the immune tolerance network, and a plethora of manuscripts have described their development and function in the course of the last two decades. Nevertheless, it is still a matter of debate as to which mechanisms and agents are employed by Treg cells, providing the basis of their suppressive potency. One of the important candidates is cyclic AMP (cAMP), which is long known as a potent suppressor at least of T cell activation and function. While this suppressive function by itself is widely accepted, the source and the mechanism of action of cAMP are less clear, and a multitude of seemingly contradictory data allow for, in principle, two different scenarios of cAMP-mediated suppression. In one scenario, Treg cells contain high amounts of cAMP and convey this small molecule via gap junction intercellular communication directly to the effector T cells (Teff) leading to their suppression. Alternatively, it was shown that Treg cells represent the origin of considerable amounts of adenosine, which trigger the adenylate cyclases in Teff cells via A2A and A2B receptors, thus strongly increasing intracellular cAMP. This review will present and discuss initial findings and recent developments concerning the function of cAMP for Treg cells and its impact on immune regulation.

  14. Cyclic nucleotide gated channel 10 negatively regulates salt tolerance by mediating Na+ transport in Arabidopsis.

    PubMed

    Jin, Yakang; Jing, Wen; Zhang, Qun; Zhang, Wenhua

    2015-01-01

    A number of cyclic nucleotide gated channel (CNGC) genes have been identified in plant genomes, but their functions are mainly undefined. In this study, we identified the role of CNGC10 in the response of Arabidopsis thaliana to salt stress. The cngc10 T-DNA insertion mutant showed greater tolerance to salt than wild-type A. thaliana during seed germination and seedling growth. The cngc10 mutant accumulated less Na(+) and K(+), but not less Ca(2+), in shoots in response to salt stress. By contrast, overexpression of CNGC10 resulted in greater sensitivity to salt stress, and complementation of this gene recovered salt sensitivity. In response to salt stress, heterologous expression of CNGC10 in the Na(+) sensitive yeast mutant strain B31 inhibited growth due to accumulation of Na(+) at a rate greater than that of yeast transformed with an empty vector. Quantitative RT-PCR analysis demonstrated that CNGC10 was expressed mainly in roots and flowers. GUS analysis of a root cross section indicated that CNGC10 was expressed mainly in the endodermis and epidermis. Furthermore, the expression of CNGC10 in roots was dramatically inhibited by exposure to 200 mM NaCl for 6 h. These data suggest that CNGC10 negatively regulates salt tolerance in A. thaliana and may be involved in mediating Na(+) transport. PMID:25416933

  15. Conserved cellular function and stress-mediated regulation among members of the proteolipid protein family.

    PubMed

    Fernández, María E; Alfonso, Julieta; Brocco, Marcela A; Frasch, Alberto C

    2010-05-01

    Chronic stress causes morphological alterations in the hippocampus of rodents and tree shrews, including atrophy of CA3 dendrites and loss of synapses. The molecular mechanisms underlying these structural changes remain largely unknown. We have previously identified M6a as a stress responsive gene and shown that M6a is involved in filopodium/spine outgrowth and, likely, synapse formation. M6a belongs to the proteolipid protein (PLP) family, all of their members having four transmembrane domains that allow their localization at the plasma membrane. In the present work, we analyzed other members of this family, the closely related M6b as well as PLP and its splice variant DM20. We found that chronic restraint stress in mice reduces M6b and DM20, but not PLP, mRNA levels in the hippocampus. In addition, M6b and DM20, but again not PLP, induce filopodium formation in primary cultures of hippocampal neurons. Several M6b protein isoforms were studied, all of them having similar effects except for the one lacking the transmembrane domains. Our results reveal a conserved cellular function and a stress-mediated regulation among members of the proteolipid protein family, suggesting an involvement of proteolipid proteins in the stress response. PMID:19937804

  16. Wnt-mediated reciprocal regulation between cartilage and bone development during endochondral ossification.

    PubMed

    Lu, Cheng; Wan, Yong; Cao, Jingjing; Zhu, Xuming; Yu, Jian; Zhou, Rujiang; Yao, Yiyun; Zhang, Lingling; Zhao, Haixia; Li, Hanjun; Zhao, Jianzhi; He, Lin; Ma, Gang; Yang, Xiao; Yao, Zhengju; Guo, Xizhi

    2013-04-01

    The role of Wnt signaling is extensively studied in skeletal development and postnatal bone remodeling, mostly based on the genetic approaches of β-catenin manipulation. However, given their independent function, a requirement for β-catenin is not the same as that for Wnt. Here, we investigated the effect of Wnt proteins in both tissues through generating cartilage- or bone-specific Wls null mice, respectively. Depletion of Wls by Col2-Cre, which would block Wnt secretion in the chondrocytes and perichondrium, delayed chondrocyte hypertrophy in the growth plate and impaired perichondrial osteogenesis. Loss of Wls in chondrocytes also disturbed the proliferating chondrocyte morphology and division orientation, which was similar to the defect observed in Wnt5a null mice. On the other hand, inactivation of Wls in osteoblasts by Col1-Cre resulted in a shorter hypertrophic zone and an increase of TRAP positive cell number in the chondro-osseous junction of growth plate, coupled with a decrease in bone mass. Taken together, our studies reveal that Wnt proteins not only modulate differentiation and cellular communication within populations of chondrocytes, but also mediate the cross regulation between the chondrocytes and osteoblasts in growth plate. PMID:23274346

  17. Trbp regulates heart function through miRNA-mediated Sox6 repression

    PubMed Central

    Ding, Jian; Chen, Jinghai; Wang, Yanqun; Kataoka, Masaharu; Ma, Lixin; Zhou, Pingzhu; Hu, Xiaoyun; Lin, Zhiqiang; Nie, Mao; Deng, Zhong-Liang; Pu, William T; Wang, Da-Zhi

    2015-01-01

    Cardiomyopathy is associated with altered expression of genes encoding contractile proteins. Here we show that Trbp (Tarbp2), an RNA binding protein, is required for normal heart function. Cardiac-specific inactivation of Trbp (TrbpcKO) caused progressive cardiomyopathy and lethal heart failure. Trbp loss of function resulted in upregulation of Sox6, repression of genes encoding normal cardiac slow-twitch myofiber proteins, and pathologically increased expression of skeletal fast-twitch myofiber genes. Remarkably, knockdown of Sox6 fully rescued the Trbp mutant phenotype, whereas Sox6 overexpression phenocopied the TrbpcKO phenotype. Trbp inactivation was mechanistically linked to Sox6 upregulation through altered processing of miR-208a, which is a direct inhibitor of Sox6. Transgenic overexpression of miR-208a sufficiently repressed Sox6, restored the balance of fast- and slow- twitch myofiber gene expression, and rescued cardiac function in TrbpcKO mice. Together, our studies reveal a novel Trbp-mediated microRNA processing mechanism in regulating a linear genetic cascade essential for normal heart function. PMID:26029872

  18. Intersectin-2L Regulates Caveola Endocytosis Secondary to Cdc42-mediated Actin Polymerization*

    PubMed Central

    Klein, Irene K.; Predescu, Dan N.; Sharma, Tiffany; Knezevic, Ivana; Malik, Asrar B.; Predescu, Sanda

    2009-01-01

    Here we addressed the role of intersectin-2L (ITSN-2L), a guanine nucleotide exchange factor for the Rho GTPase Cdc42, in the mechanism of caveola endocytosis in endothelial cells (ECs). Immunoprecipitation and co-localization studies showed that ITSN-2L associates with members of the Cdc42-WASp-Arp2/3 actin polymerization pathway. Expression of Dbl homology-pleckstrin homology (DH-PH) region of ITSN-2L (DH-PHITSN-2L) induced specific activation of Cdc42, resulting in formation of extensive filopodia, enhanced cortical actin, as well as a shift from G-actin to F-actin. The “catalytically dead” DH-PH domain reversed these effects and induced significant stress fiber formation, without a detectable shift in actin pools. A biotin assay for caveola internalization indicated a significant decrease in the uptake of biotinylated proteins in DH-PHITSN-2L-transfected cells compared with control and 1 μm jasplakinolide-treated cells. ECs depleted of ITSN-2L by small interfering RNA, however, showed decreased Cdc42 activation and actin remodeling similar to the defective DH-PH, resulting in 62% increase in caveola-mediated uptake compared with controls. Thus, ITSN-2L, a guanine nucleotide exchange factor for Cdc42, regulates different steps of caveola endocytosis in ECs by controlling the temporal and spatial actin polymerization and remodeling sub-adjacent to the plasma membrane. PMID:19622753

  19. Intersectin-2L regulates caveola endocytosis secondary to Cdc42-mediated actin polymerization.

    PubMed

    Klein, Irene K; Predescu, Dan N; Sharma, Tiffany; Knezevic, Ivana; Malik, Asrar B; Predescu, Sanda

    2009-09-18

    Here we addressed the role of intersectin-2L (ITSN-2L), a guanine nucleotide exchange factor for the Rho GTPase Cdc42, in the mechanism of caveola endocytosis in endothelial cells (ECs). Immunoprecipitation and co-localization studies showed that ITSN-2L associates with members of the Cdc42-WASp-Arp2/3 actin polymerization pathway. Expression of Dbl homology-pleckstrin homology (DH-PH) region of ITSN-2L (DH-PH(ITSN-2L)) induced specific activation of Cdc42, resulting in formation of extensive filopodia, enhanced cortical actin, as well as a shift from G-actin to F-actin. The "catalytically dead" DH-PH domain reversed these effects and induced significant stress fiber formation, without a detectable shift in actin pools. A biotin assay for caveola internalization indicated a significant decrease in the uptake of biotinylated proteins in DH-PH(ITSN-2L)-transfected cells compared with control and 1 microM jasplakinolide-treated cells. ECs depleted of ITSN-2L by small interfering RNA, however, showed decreased Cdc42 activation and actin remodeling similar to the defective DH-PH, resulting in 62% increase in caveola-mediated uptake compared with controls. Thus, ITSN-2L, a guanine nucleotide exchange factor for Cdc42, regulates different steps of caveola endocytosis in ECs by controlling the temporal and spatial actin polymerization and remodeling sub-adjacent to the plasma membrane. PMID:19622753

  20. Peripheral Brain Derived Neurotrophic Factor Precursor Regulates Pain as an Inflammatory Mediator

    PubMed Central

    Luo, Cong; Zhong, Xiao-Lin; Zhou, Fiona H.; Li, Jia-yi; Zhou, Pei; Xu, Jun-Mei; Song, Bo; Li, Chang-Qi; Zhou, Xin-Fu; Dai, Ru-Ping

    2016-01-01

    The precursor of brain derived neurotrophic factor (proBDNF), the unprocessed BDNF gene product, binds to its receptors and exerts the opposing biologic functions of mature BDNF. proBDNF is expressed in the peripheral tissues but the functions of peripheral proBDNF remain elusive. Here we showed that proBDNF and its predominant receptor, p75 pan-neurotrophin receptor were upregulated in the nerve fibers and inflammatory cells in the local tissue in inflammatory pain. Neutralization of proBDNF by polyclonal antibody attenuated pain in different models of inflammatory pain. Unilateral intra-plantar supplementation of proBDNF by injecting exogenous proBDNF or ectopic overexpression resulted in pain hypersensitivity and induced spinal phosphorylated extracellular signal-regulated kinase activation. Exogenous proBDNF injection induced the infiltration of inflammatory cells and the activation of proinflammatory cytokines, suggesting that inflammatory reaction contributed to the pro-algesic effect of proBDNF. Finally, we generated monoclonal anti-proBDNF antibody that could biologically block proBDNF. Administration of monoclonal Ab-proBDNF attenuated various types of inflammatory pain and surgical pain. Thus, peripheral proBDNF is a potential pain mediator and anti-proBDNF pretreatment may alleviate the development of inflammatory pain. PMID:27251195