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Sample records for regulates transcriptional activity

  1. Post-translational regulation of Oct4 transcriptional activity.

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    Jonathan P Saxe

    Full Text Available Oct4 is a key component of the molecular circuitry which regulates embryonic stem cell proliferation and differentiation. It is essential for maintenance of undifferentiated, pluripotent cell populations, and accomplishes these tasks by binding DNA in multiple heterodimer and homodimer configurations. Very little is known about how formation of these complexes is regulated, or the mechanisms through which Oct4 proteins respond to complex extracellular stimuli which regulate pluripotency. Here, we provide evidence for a phosphorylation-based mechanism which regulates specific Oct4 homodimer conformations. Point mutations of a putative phosphorylation site can specifically abrogate transcriptional activity of a specific homodimer assembly, with little effect on other configurations. Moreover, we performed bioinformatic predictions to identify a subset of Oct4 target genes which may be regulated by this specific assembly, and show that altering Oct4 protein levels affects transcription of Oct4 target genes which are regulated by this assembly but not others. Finally, we identified several signaling pathways which may mediate this phosphorylation and act in combination to regulate Oct4 transcriptional activity and protein stability. These results provide a mechanism for rapid and reversible alteration of Oct4 transactivation potential in response to extracellular signals.

  2. Activating transcription factor 4 regulates osteoclast differentiation in mice

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    Cao, Huiling; Yu, Shibing; Yao, Zhi; Galson, Deborah L.; Jiang, Yu; Zhang, Xiaoyan; Fan, Jie; Lu, Binfeng; Guan, Youfei; Luo, Min; Lai, Yumei; Zhu, Yibei; Kurihara, Noriyoshi; Patrene, Kenneth; Roodman, G. David; Xiao, Guozhi

    2010-01-01

    Activating transcription factor 4 (ATF4) is a critical transcription factor for osteoblast (OBL) function and bone formation; however, a direct role in osteoclasts (OCLs) has not been established. Here, we targeted expression of ATF4 to the OCL lineage using the Trap promoter or through deletion of Atf4 in mice. OCL differentiation was drastically decreased in Atf4–/– bone marrow monocyte (BMM) cultures and bones. Coculture of Atf4–/– BMMs with WT OBLs or a high concentration of RANKL failed to restore the OCL differentiation defect. Conversely, Trap-Atf4-tg mice displayed severe osteopenia with dramatically increased osteoclastogenesis and bone resorption. We further showed that ATF4 was an upstream activator of the critical transcription factor Nfatc1 and was critical for RANKL activation of multiple MAPK pathways in OCL progenitors. Furthermore, ATF4 was crucial for M-CSF induction of RANK expression on BMMs, and lack of ATF4 caused a shift in OCL precursors to macrophages. Finally, ATF4 was largely modulated by M-CSF signaling and the PI3K/AKT pathways in BMMs. These results demonstrate that ATF4 plays a direct role in regulating OCL differentiation and suggest that it may be a therapeutic target for treating bone diseases associated with increased OCL activity. PMID:20628199

  3. Activating transcription factor 4 regulates osteoclast differentiation in mice.

    Science.gov (United States)

    Cao, Huiling; Yu, Shibing; Yao, Zhi; Galson, Deborah L; Jiang, Yu; Zhang, Xiaoyan; Fan, Jie; Lu, Binfeng; Guan, Youfei; Luo, Min; Lai, Yumei; Zhu, Yibei; Kurihara, Noriyoshi; Patrene, Kenneth; Roodman, G David; Xiao, Guozhi

    2010-08-01

    Activating transcription factor 4 (ATF4) is a critical transcription factor for osteoblast (OBL) function and bone formation; however, a direct role in osteoclasts (OCLs) has not been established. Here, we targeted expression of ATF4 to the OCL lineage using the Trap promoter or through deletion of Atf4 in mice. OCL differentiation was drastically decreased in Atf4-/- bone marrow monocyte (BMM) cultures and bones. Coculture of Atf4-/- BMMs with WT OBLs or a high concentration of RANKL failed to restore the OCL differentiation defect. Conversely, Trap-Atf4-tg mice displayed severe osteopenia with dramatically increased osteoclastogenesis and bone resorption. We further showed that ATF4 was an upstream activator of the critical transcription factor Nfatc1 and was critical for RANKL activation of multiple MAPK pathways in OCL progenitors. Furthermore, ATF4 was crucial for M-CSF induction of RANK expression on BMMs, and lack of ATF4 caused a shift in OCL precursors to macrophages. Finally, ATF4 was largely modulated by M-CSF signaling and the PI3K/AKT pathways in BMMs. These results demonstrate that ATF4 plays a direct role in regulating OCL differentiation and suggest that it may be a therapeutic target for treating bone diseases associated with increased OCL activity.

  4. Resveratrol regulates gene transcription via activation of stimulus-responsive transcription factors.

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    Thiel, Gerald; Rössler, Oliver G

    2017-03-01

    Resveratrol (trans-3,4',5-trihydroxystilbene), a polyphenolic phytoalexin of grapes and other fruits and plants, is a common constituent of our diet and of dietary supplements. Many health-promoting benefits have been connected with resveratrol in the treatment of cardiovascular diseases, cancer, diabetes, inflammation, neurodegeneration, and diseases connected with aging. To explain the pleiotropic effects of resveratrol, the molecular targets of this compound have to be identified on the cellular level. Resveratrol induces intracellular signal transduction pathways which ultimately lead to changes in the gene expression pattern of the cells. Here, we review the effect of resveratrol on the activation of the stimulus-responsive transcription factors CREB, AP-1, Egr-1, Elk-1, and Nrf2. Following activation, these transcription factors induce transcription of delayed response genes. The gene products of these delayed response genes are ultimately responsible for the changes in the biochemistry and physiology of resveratrol-treated cells. The activation of stimulus-responsive transcription factors may explain many of the intracellular activities of resveratrol. However, results obtained in vitro may not easily be transferred to in vivo systems.

  5. Analyzing phosphorylation-dependent regulation of subcellular localization and transcriptional activity of transcriptional coactivator NT-PGC-1α.

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    Chang, Ji Suk; Gettys, Thomas W

    2013-01-01

    Peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α) is a nuclear transcriptional coactivator that regulates the genes involved in energy metabolism. Recent evidence has been provided that alternative splicing of PPARGC1A gene produces a functional but predominantly cytosolic isoform of PGC-1α (NT-PGC-1α). We have demonstrated that transcriptional coactivation capacity of NT-PGC-1α is directly correlated with its nuclear localization in a PKA phosphorylation-dependent manner. In this chapter, we describe quantitative imaging analysis methods that are developed to measure the relative fluorescence intensity of the protein of interest in the nucleus and cytoplasm in a single cell and the frequency distribution of nuclear/cytoplasmic intensity ratios in the population of cells, respectively. This chapter also describes transient cotransfection and dual-luciferase reporter gene assay that examine the ability of coactivators to activate the transcriptional activity of transcription factors.

  6. Inferring yeast cell cycle regulators and interactions using transcription factor activities

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    Galbraith Simon J

    2005-06-01

    Full Text Available Abstract Background Since transcription factors are often regulated at the post-transcriptional level, their activities, rather than expression levels may provide valuable information for investigating functions and their interactions. The recently developed Network Component Analysis (NCA and its generalized form (gNCA provide a robust framework for deducing the transcription factor activities (TFAs from various types of DNA microarray data and transcription factor-gene connectivity. The goal of this work is to demonstrate the utility of TFAs in inferring transcription factor functions and interactions in Saccharomyces cerevisiae cell cycle regulation. Results Using gNCA, we determined 74 TFAs from both wild type and fkh1 fkh2 deletion mutant microarray data encompassing 1529 ORFs. We hypothesized that transcription factors participating in the cell cycle regulation exhibit cyclic activity profiles. This hypothesis was supported by the TFA profiles of known cell cycle factors and was used as a basis to uncover other potential cell cycle factors. By combining the results from both cluster analysis and periodicity analysis, we recovered nearly 90% of the known cell cycle regulators, and identified 5 putative cell cycle-related transcription factors (Dal81, Hap2, Hir2, Mss11, and Rlm1. In addition, by analyzing expression data from transcription factor knockout strains, we determined 3 verified (Ace2, Ndd1, and Swi5 and 4 putative interaction partners (Cha4, Hap2, Fhl1, and Rts2 of the forkhead transcription factors. Sensitivity of TFAs to connectivity errors was determined to provide confidence level of these predictions. Conclusion By subjecting TFA profiles to analyses based upon physiological signatures we were able to identify cell cycle related transcription factors consistent with current literature, transcription factors with potential cell cycle dependent roles, and interactions between transcription factors.

  7. Osterix represses adipogenesis by negatively regulating PPARγ transcriptional activity

    OpenAIRE

    Younho Han; Chae Yul Kim; Heesun Cheong; Kwang Youl Lee

    2016-01-01

    Osterix is a novel bone-related transcription factor involved in osteoblast differentiation, and bone maturation. Because a reciprocal relationship exists between adipocyte and osteoblast differentiation of bone marrow derived mesenchymal stem cells, we hypothesized that Osterix might have a role in adipogenesis. Ablation of Osterix enhanced adipogenesis in 3T3-L1 cells, whereas overexpression suppressed this process and inhibited the expression of adipogenic markers including CCAAT/enhancer-...

  8. Regulation and function of signal transducer and activator of transcription 3

    Institute of Scientific and Technical Information of China (English)

    Qian-Rong; Qi; Zeng-Ming; Yang

    2014-01-01

    Signal transducer and activator of transcription 3(STAT3), a member of the STAT family, is a key regulator of many physiological and pathological processes. Significant progress has been made in understanding the transcriptional control, posttranslational modification, cellular localization and functional regulation of STAT3. STAT3 can translocate into the nucleus and bind to specific promoter sequences, thereby exerting transcriptional regulation. Recent studies have shown that STAT3 can also translocate into mitochondria, participating in aerobic respiration and apoptosis. In addition, STAT3 plays an important role in inflammation and tumorigenesis by regulating cell proliferation, differentiation and metabolism. Conditional knockout mouse models make it possible to study the physiological function of STAT3 in specific tissues and organs. This review summarizes the latest advances in the understanding of the expression, regulation and function of STAT3 in physiological and tumorigenic processes.

  9. Nutritional conditions regulate transcriptional activity of SF-1 by controlling sumoylation and ubiquitination.

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    Lee, Jiwon; Yang, Dong Joo; Lee, Syann; Hammer, Gary D; Kim, Ki Woo; Elmquist, Joel K

    2016-01-11

    Steroidogenic factor 1 (SF-1) is a transcription factor expressed in the ventral medial nucleus of the hypothalamus that regulates energy homeostasis. However, the molecular mechanisms of SF-1 in the control of energy balance are largely unknown. Here, we show that nutritional conditions, such as the presence or absence of serum, affect SF-1 action. Serum starvation significantly decreased hypothalamic SF-1 levels by promoting ubiquitin-dependent degradation, and sumoylation was required for this process. SF-1 transcriptional activity was also differentially regulated by nutritional status. Under normal conditions, the transcriptional activity of hypothalamic SF-1 was activated by SUMO, but this was attenuated during starvation. Taken together, these results indicate that sumoylation and ubiquitination play crucial roles in the regulation of SF-1 function and that these effects are dependent on nutritional conditions, further supporting the importance of SF-1 in the control of energy homeostasis.

  10. Proximal genomic localization of STAT1 binding and regulated transcriptional activity

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    Smyth Gordon K

    2006-10-01

    Full Text Available Abstract Background Signal transducer and activator of transcription (STAT proteins are key regulators of gene expression in response to the interferon (IFN family of anti-viral and anti-microbial cytokines. We have examined the genomic relationship between STAT1 binding and regulated transcription using multiple tiling microarray and chromatin immunoprecipitation microarray (ChIP-chip experiments from public repositories. Results In response to IFN-γ, STAT1 bound proximally to regions of the genome that exhibit regulated transcriptional activity. This finding was consistent between different tiling microarray platforms, and between different measures of transcriptional activity, including differential binding of RNA polymerase II, and differential mRNA transcription. Re-analysis of tiling microarray data from a recent study of IFN-γ-induced STAT1 ChIP-chip and mRNA expression revealed that STAT1 binding is tightly associated with localized mRNA transcription in response to IFN-γ. Close relationships were also apparent between STAT1 binding, STAT2 binding, and mRNA transcription in response to IFN-α. Furthermore, we found that sites of STAT1 binding within the Encyclopedia of DNA Elements (ENCODE region are precisely correlated with sites of either enhanced or diminished binding by the RNA polymerase II complex. Conclusion Together, our results indicate that STAT1 binds proximally to regions of the genome that exhibit regulated transcriptional activity. This finding establishes a generalized basis for the positioning of STAT1 binding sites within the genome, and supports a role for STAT1 in the direct recruitment of the RNA polymerase II complex to the promoters of IFN-γ-responsive genes.

  11. O-GlcNAc modification of Sp3 and Sp4 transcription factors negatively regulates their transcriptional activities.

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    Ha, Changhoon; Lim, Kihong

    2015-11-13

    The addition of O-linked N-acetylglucosamine (O-GlcNAc) on serine or threonine modifies a myriad of proteins and regulates their function, stability and localization. O-GlcNAc modification is common among chromosome-associated proteins, such as transcription factors, suggesting its extensive involvement in gene expression regulation. In this study, we demonstrate the O-GlcNAc status of the Sp family members of transcription factors and the functional impact on their transcriptional activities. We highlight the presence of O-GlcNAc residues in Sp3 and Sp4, but not Sp2, as demonstrated by their enrichment in GlcNAc positive protein fractions and by detection of O-GlcNAc residues on Sp3 and Sp4 co-expressed in Escherichia coli together with O-GlcNAc transferase (OGT) using an O-GlcNAc-specific antibody. Deletion mutants of Sp3 and Sp4 indicate that the majority of O-GlcNAc sites reside in their N-terminal transactivation domain. Overall, using reporter gene assays and co-immunoprecipitations, we demonstrate a functional inhibitory role of O-GlcNAc modifications in Sp3 and Sp4 transcription factors. Thereby, our study strengthens the current notion that O-GlcNAc modification is an important regulator of protein interactome.

  12. An activator of transcription regulates phage TP901-1 late gene expression

    DEFF Research Database (Denmark)

    Brøndsted, Lone; Pedersen, Margit; Hammer, Karin

    2001-01-01

    to activate transcription of the promoter. Several lactococcal bacteriophages encode ORF29 homologous proteins, indicating that late transcription may be controlled by a similar mechanism in these phages. With the identification of this novel regulator, our results suggest that within the P335 group...... bp contains both the promoter and the region necessary for activation by ORF29. The transcriptional start site of the promoter was identified by primer extension to position 13073 on the TP901-1 genome, thus located 87 bp downstream of orf29 in a 580-bp intergenic region between orf29 and orf30....... Furthermore, the region located -85 to -61 bp upstream of the start site was shown to be necessary for promoter activity. During infection, the transcript arising from the late promoter is fully induced at 40 min postinfection, and our results suggest that a certain level of ORF29 must he reached in order...

  13. Nitrogen-regulated transcription and enzyme activities in continuous cultures of Saccharomyces cerevisiae.

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    ter Schure, E G; Silljé, H H; Raeven, L J; Boonstra, J; Verkleij, A J; Verrips, C T

    1995-05-01

    Variations in the transcription of nitrogen-regulated genes and in the activities of nitrogen-regulated enzymes of the yeast Saccharomyces cerevisiae were studied by changing the carbon and nitrogen fluxes. S. cerevisiae was grown in continuous culture at various dilution rates (D) under nitrogen limitation with NH4Cl as sole nitrogen source. With an increase in D from 0.05 to 0.29 h-1, both the glucose and the ammonia flux increased sixfold. The activities of the two ammonia-incorporating enzymes, NADPH-dependent glutamate dehydrogenase (NADPH-GDH) and glutamine synthetase (GS), encoded by GDH1 and GLN1, respectively, increased with increasing D, while the activity of the glutamate-degrading enzyme, NAD-dependent glutamate dehydrogenase (NAD-GDH), decreased. Surprisingly, no changes were observed in the transcription of GDH1 and GLN1; however increased D was accompanied by an increase in GAP1 transcription. At the metabolite level, the increase in the glucose and nitrogen flux did not result in changes in the intracellular 2-oxoglutarate, glutamate or glutamine concentrations. It is shown that growth on ammonia alone is not sufficient to cause repression of GAP1 and GLN1 transcription and that the regulation of GAP1 transcription and both NADPH-GDH and GS activity is not an on/off switch, but is gradually modulated in correlation with the ammonia concentration.

  14. O-GlcNAc transferase regulates transcriptional activity of human Oct4.

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    Constable, Sandii; Lim, Jae-Min; Vaidyanathan, Krithika; Wells, Lance

    2017-10-01

    O-linked β-N-acetylglucosamine (O-GlcNAc) is a single sugar modification found on many different classes of nuclear and cytoplasmic proteins. Addition of this modification, by the enzyme O-linked N-acetylglucosamine transferase (OGT), is dynamic and inducible. One major class of proteins modified by O-GlcNAc is transcription factors. O-GlcNAc regulates transcription factor properties through a variety of different mechanisms including localization, stability and transcriptional activation. Maintenance of embryonic stem (ES) cell pluripotency requires tight regulation of several key transcription factors, many of which are modified by O-GlcNAc. Octamer-binding protein 4 (Oct4) is one of the key transcription factors required for pluripotency of ES cells and more recently, the generation of induced pluripotent stem (iPS) cells. The action of Oct4 is modulated by the addition of several post-translational modifications, including O-GlcNAc. Previous studies in mice found a single site of O-GlcNAc addition responsible for transcriptional regulation. This study was designed to determine if this mechanism is conserved in humans. We mapped 10 novel sites of O-GlcNAc attachment on human Oct4, and confirmed a role for OGT in transcriptional activation of Oct4 at a site distinct from that found in mouse that allows distinction between different Oct4 target promoters. Additionally, we uncovered a potential new role for OGT that does not include its catalytic function. These results confirm that human Oct4 activity is being regulated by OGT by a mechanism that is distinct from mouse Oct4. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  15. CDPK1, a calcium-dependent protein kinase, regulates transcriptional activator RSG in response to gibberellins.

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    Nakata, Masaru; Yuasa, Takashi; Takahashi, Yohsuke; Ishida, Sarahmi

    2009-05-01

    The homeostasis of gibberellins (GAs) is maintained by negative-feedback regulation in plant cells. REPRESSION OF SHOOT GROWTH (RSG) is a transcriptional activator with a basic Leu zipper domain suggested to contribute GA feedback regulation by the transcriptional regulation of genes encoding GA biosynthetic enzymes. The 14-3-3 signaling proteins negatively regulate RSG by sequestering it in the cytoplasm in response to GAs. The phosphorylation on Ser-114 of RSG is essential for 14-3-3 binding of RSG; however, the kinase that catalyzes the reaction is unknown. Recently a Ca(2+)-dependent protein kinase (CDPK) was identified as an RSG kinase that promotes 14-3-3 binding of RSG by phosphorylation of the Ser-114 of RSG. Our results suggest that CDPK decodes the Ca(2+) signal produced by GAs and regulates the intracellular localization of RSG in plant cells.

  16. Hypoxia-Inducible Factor 3 Is an Oxygen-Dependent Transcription Activator and Regulates a Distinct Transcriptional Response to Hypoxia

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    Peng Zhang

    2014-03-01

    Full Text Available Hypoxia-inducible factors (HIFs play key roles in the cellular response to hypoxia. It is widely accepted that whereas HIF-1 and HIF-2 function as transcriptional activators, HIF-3 inhibits HIF-1/2α action. Contrary to this idea, we show that zebrafish Hif-3α has strong transactivation activity. Hif-3α is degraded under normoxia. Mutation of P393, P493, and L503 inhibits this oxygen-dependent degradation. Transcriptomics and chromatin immunoprecipitation analyses identify genes that are regulated by Hif-3α, Hif-1α, or both. Under hypoxia or when overexpressed, Hif-3α binds to its target gene promoters and upregulates their expression. Dominant-negative inhibition and knockdown of Hif-3α abolish hypoxia-induced Hif-3α-promoter binding and gene expression. Hif-3α not only mediates hypoxia-induced growth and developmental retardation but also possesses hypoxia-independent activities. Importantly, transactivation activity is conserved and human HIF-3α upregulates similar genes in human cells. These findings suggest that Hif-3 is an oxygen-dependent transcription factor and activates a distinct transcriptional response to hypoxia.

  17. FBXO3 Protein Promotes Ubiquitylation and Transcriptional Activity of AIRE (Autoimmune Regulator)*

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    Shao, Wei; Zumer, Kristina; Fujinaga, Koh; Peterlin, B. Matija

    2016-01-01

    The autoimmune regulator (AIRE) is a transcription factor which is expressed in medullary thymic epithelial cells. It directs the expression of otherwise tissue-specific antigens, which leads to the elimination of autoreactive T cells during development. AIRE is modified post-translationally by phosphorylation and ubiquitylation. In this report we connected these modifications. AIRE, which is phosphorylated on two specific residues near its N terminus, then binds to the F-box protein 3 (FBXO3) E3 ubiquitin ligase. In turn, this SCFFBXO3 (SKP1-CUL1-F box) complex ubiquitylates AIRE, increases its binding to the positive transcription elongation factor b (P-TEFb), and potentiates its transcriptional activity. Because P-TEFb is required for the transition from initiation to elongation of transcription, this interaction ensures proper expression of AIRE-responsive tissue-specific antigens in the thymus. PMID:27365398

  18. FBXO3 Protein Promotes Ubiquitylation and Transcriptional Activity of AIRE (Autoimmune Regulator).

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    Shao, Wei; Zumer, Kristina; Fujinaga, Koh; Peterlin, B Matija

    2016-08-19

    The autoimmune regulator (AIRE) is a transcription factor which is expressed in medullary thymic epithelial cells. It directs the expression of otherwise tissue-specific antigens, which leads to the elimination of autoreactive T cells during development. AIRE is modified post-translationally by phosphorylation and ubiquitylation. In this report we connected these modifications. AIRE, which is phosphorylated on two specific residues near its N terminus, then binds to the F-box protein 3 (FBXO3) E3 ubiquitin ligase. In turn, this SCF(FBXO3) (SKP1-CUL1-F box) complex ubiquitylates AIRE, increases its binding to the positive transcription elongation factor b (P-TEFb), and potentiates its transcriptional activity. Because P-TEFb is required for the transition from initiation to elongation of transcription, this interaction ensures proper expression of AIRE-responsive tissue-specific antigens in the thymus. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. IscR regulates RNase LS activity by repressing rnlA transcription.

    Science.gov (United States)

    Otsuka, Yuichi; Miki, Kumiko; Koga, Mitsunori; Katayama, Natsu; Morimoto, Wakako; Takahashi, Yasuhiro; Yonesaki, Tetsuro

    2010-07-01

    The Escherichia coli endoribonuclease LS was originally identified as a potential antagonist of bacteriophage T4. When the T4 dmd gene is defective, RNase LS cleaves T4 mRNAs and antagonizes T4 reproduction. This RNase also plays an important role in RNA metabolisms in E. coli. rnlA is an essential gene for RNase LS activity, but the transcriptional regulation of this gene remains to be elucidated. An Fe-S cluster protein, IscR, acts as a transcription factor and controls the expression of genes that are necessary for Fe-S cluster biogenesis. Here, we report that overexpression of IscR suppressed RNase LS activity, causing the loss of antagonist activity against phage T4. This suppressive effect did not require the ligation of Fe-S cluster into IscR. beta-Galactosidase reporter assays showed that transcription from an rnlA promoter increased in iscR-deleted cells compared to wild-type cells, and gel-mobility shift assays revealed specific binding of IscR to the rnlA promoter region. RT-PCR analysis demonstrated that endogenous rnlA mRNA was reduced by overexpression of IscR and increased by deletion of iscR. From these results, we conclude that IscR negatively regulates transcription of rnlA and represses RNase LS activity.

  20. Alternative Polyadenylation Regulates CELF1/CUGBP1 Target Transcripts Following T Cell Activation

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    Beisang, Daniel; Reilly, Cavan; Bohjanen, Paul R.

    2014-01-01

    Alternative polyadenylation (APA) is an evolutionarily conserved mechanism for regulating gene expression. Transcript 3′ end shortening through changes in polyadenylation site usage occurs following T cell activation, but the consequences of APA on gene expression are poorly understood. We previously showed that GU-rich elements (GREs) found in the 3′ untranslated regions of select transcripts mediate rapid mRNA decay by recruiting the protein CELF1/CUGBP1. Using a global RNA sequencing approach, we found that a network of CELF1 target transcripts involved in cell division underwent preferential 3′ end shortening via APA following T cell activation, resulting in decreased inclusion of CELF1 binding sites and increased transcript expression. We present a model whereby CELF1 regulates APA site selection following T cell activation through reversible binding to nearby GRE sequences. These findings provide insight into the role of APA in controlling cellular proliferation during biological processes such as development, oncogenesis and T cell activation PMID:25123787

  1. Adaptation of the Agrobacterium tumefaciens VirG response regulator to activate transcription in plants.

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    Czarnecka-Verner, Eva; Salem, Tarek A; Gurley, William B

    2016-02-01

    The Agrobacterium tumefaciens VirG response regulator of the VirA/VirG two-component system was adapted to function in tobacco protoplasts. The subcellular localization of VirG and VirA proteins transiently expressed in onion cells was determined using GFP fusions. Preliminary studies using Gal4DBD-VP16 fusions with VirG and Escherichia coli UhpA, and NarL response regulators indicated compatibility of these bacterial proteins with the eukaryotic transcriptional apparatus. A strong transcriptional activator based on tandem activation domains from the Drosophila fushi tarazu and Herpes simplex VP16 was created. Selected configurations of the two-site Gal4-vir box GUS reporters were activated by chimeric effectors dependent on either the yeast Gal4 DNA-binding domain or that of VirG. Transcriptional induction of the GUS reporter was highest for the VirE19-element promoter with both constitutive and wild-type VirG-tandem activation domain effectors. Multiple VirE19 elements increased the reporter activity proportionately, indicating that the VirG DNA binding domain was functional in plants. The VirG constitutive-Q-VP16 effector was more active than the VirG wild-type. In both the constitutive and wild-type forms of VirG, Q-VP16 activated transcription of the GUS reporter best when located at the C-terminus, i.e. juxtaposed to the VirG DNA binding domain. These results demonstrate the possibility of using DNA binding domains from bacterial response regulators and their cognate binding elements in the engineering of plant gene expression.

  2. RIT1 GTPase Regulates Sox2 Transcriptional Activity and Hippocampal Neurogenesis.

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    Mir, Sajad; Cai, Weikang; Andres, Douglas A

    2017-02-10

    Adult neurogenesis, the process of generating mature neurons from neuronal progenitor cells, makes critical contributions to neural circuitry and brain function in both healthy and disease states. Neurogenesis is a highly regulated process in which diverse environmental and physiological stimuli are relayed to resident neural stem cell populations to control the transcription of genes involved in self-renewal and differentiation. Understanding the molecular mechanisms governing neurogenesis is necessary for the development of translational strategies to harness this process for neuronal repair. Here we report that the Ras-related GTPase RIT1 serves to control the sequential proliferation and differentiation of adult hippocampal neural progenitor cells, with in vivo expression of active RIT1 driving robust adult neurogenesis. Gene expression profiling analysis demonstrates increased expression of a specific set of transcription factors known to govern adult neurogenesis in response to active RIT1 expression in the hippocampus, including sex-determining region Y-related HMG box 2 (Sox2), a well established regulator of stem cell self-renewal and neurogenesis. In adult hippocampal neuronal precursor cells, RIT1 controls an Akt-dependent signaling cascade, resulting in the stabilization and transcriptional activation of phosphorylated Sox2. This study supports a role for RIT1 in relaying niche-derived signals to neural/stem progenitor cells to control transcription of genes involved in self-renewal and differentiation.

  3. Regulating retrotransposon activity through the use of alternative transcription start sites

    DEFF Research Database (Denmark)

    Persson, Jenna; Steglich, Babett; Smialowska, Agata

    2016-01-01

    a new mechanism of retrotransposon regulation through transcription start site (TSS) selection by altered nucleosome occupancy. We show that Fun30 chromatin remodelers cooperate to maintain a high level of nucleosome occupancy at retrotransposon-flanking long terminal repeat (LTR) elements......Retrotransposons, the ancestors of retroviruses, have the potential for gene disruption and genomic takeover if not kept in check. Paradoxically, although host cells repress these elements by multiple mechanisms, they are transcribed and are even activated under stress conditions. Here, we describe....... This enforces the use of a downstream TSS and the production of a truncated RNA incapable of reverse transcription and retrotransposition. However, in stressed cells, nucleosome occupancy at LTR elements is reduced, and the TSS shifts to allow for productive transcription. We propose that controlled...

  4. The transcriptional regulator Aire binds to and activates super-enhancers.

    Science.gov (United States)

    Bansal, Kushagra; Yoshida, Hideyuki; Benoist, Christophe; Mathis, Diane

    2017-03-01

    Aire is a transcription factor that controls T cell tolerance by inducing the expression of a large repertoire of genes specifically in thymic stromal cells. It interacts with scores of protein partners of diverse functional classes. We found that Aire and some of its partners, notably those implicated in the DNA-damage response, preferentially localized to and activated long chromatin stretches that were overloaded with transcriptional regulators, known as super-enhancers. We also identified topoisomerase 1 as a cardinal Aire partner that colocalized on super-enhancers and was required for the interaction of Aire with all of its other associates. We propose a model that entails looping of super-enhancers to efficiently deliver Aire-containing complexes to local and distal transcriptional start sites.

  5. Identification of Sp1 as a Transcription Activator to Regulate Fibroblast Growth Factor 21 Gene Expression

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    Shuqin Chen

    2017-01-01

    Full Text Available Fibroblast growth factor 21 (FGF21 is a metabolic hormone with multiple beneficial effects on lipid and glucose homeostasis. Previous study demonstrated that FGF21 might be one of the Sp1 target genes. However, the transcriptional role of Sp1 on FGF21 in adipose tissue and liver has not been reported. In this study, we found that the proximal promoter of mouse FGF21 is located between −63 and −20 containing two putative Sp1-binding sites. Sp1 is a mammalian transcription factor involved in the regulation of many genes during physiological and pathological processes. Our study showed that overexpression of Sp1 or suppressing Sp1 expression resulted in increased or reduced FGF21 promoter activity, respectively. Mutation analysis demonstrated that the Sp1-binding site located between −46 and −38 plays a primary role in transcription of FGF21. Electrophoretic mobility shift assay and chromatin immunoprecipitation analysis indicated that Sp1 specifically bound to this region. Furthermore, the binding activity of Sp1 was significantly increased in adipose tissues of HFD-induced obese mouse and liver of DEN-treated mouse. Thus, our results demonstrate that Sp1 positively regulates the basal transcription of FGF21 in the liver and adipose tissue and contributes to the obesity-induced FGF21 upregulation in mouse adipose tissue and hepatic FGF21 upregulation in hepatocarcinogenesis.

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

    Energy Technology Data Exchange (ETDEWEB)

    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

  7. Signal transducer and activator of transcription 3 regulation by novel binding partners

    Institute of Scientific and Technical Information of China (English)

    Tadashi; Matsuda; Ryuta; Muromoto; Yuichi; Sekine; Sumihito; Togi; Yuichi; Kitai; Shigeyuki; Kon; Kenji; Oritani

    2015-01-01

    Signal transducers and activators of transcription(STATs) mediate essential signals for various biological processes,including immune responses,hematopoiesis,and neurogenesis. STAT3,for example,is involved in the pathogenesis of various human diseases,including cancers,autoimmune and inflammatory disorders. STAT3 activation is therefore tightly regulated at multiple levels to prevent these pathological conditions. A number of proteins have been reported to associate with STAT3 and regulate its activity. These STAT3-interacting proteins function to modulate STAT3-mediated signaling at various steps and mediate the crosstalk of STAT3 with other cellular signaling pathways. This article reviews the roles of novel STAT3 binding partners such as DAXX,zipperinteracting protein kinase,Krüppel-associated box-associated protein 1,Y14,PDZ and LIM domain 2 and signal transducing adaptor protein-2,in the regulation of STAT3-mediated signaling.

  8. RNA Polymerase II Regulates Topoisomerase 1 Activity to Favor Efficient Transcription.

    Science.gov (United States)

    Baranello, Laura; Wojtowicz, Damian; Cui, Kairong; Devaiah, Ballachanda N; Chung, Hye-Jung; Chan-Salis, Ka Yim; Guha, Rajarshi; Wilson, Kelli; Zhang, Xiaohu; Zhang, Hongliang; Piotrowski, Jason; Thomas, Craig J; Singer, Dinah S; Pugh, B Franklin; Pommier, Yves; Przytycka, Teresa M; Kouzine, Fedor; Lewis, Brian A; Zhao, Keji; Levens, David

    2016-04-01

    We report a mechanism through which the transcription machinery directly controls topoisomerase 1 (TOP1) activity to adjust DNA topology throughout the transcription cycle. By comparing TOP1 occupancy using chromatin immunoprecipitation sequencing (ChIP-seq) versus TOP1 activity using topoisomerase 1 sequencing (TOP1-seq), a method reported here to map catalytically engaged TOP1, TOP1 bound at promoters was discovered to become fully active only after pause-release. This transition coupled the phosphorylation of the carboxyl-terminal-domain (CTD) of RNA polymerase II (RNAPII) with stimulation of TOP1 above its basal rate, enhancing its processivity. TOP1 stimulation is strongly dependent on the kinase activity of BRD4, a protein that phosphorylates Ser2-CTD and regulates RNAPII pause-release. Thus the coordinated action of BRD4 and TOP1 overcame the torsional stress opposing transcription as RNAPII commenced elongation but preserved negative supercoiling that assists promoter melting at start sites. This nexus between transcription and DNA topology promises to elicit new strategies to intercept pathological gene expression.

  9. Occludin controls HIV transcription in brain pericytes via regulation of SIRT-1 activation.

    Science.gov (United States)

    Castro, Victor; Bertrand, Luc; Luethen, Mareen; Dabrowski, Sebastian; Lombardi, Jorge; Morgan, Laura; Sharova, Natalia; Stevenson, Mario; Blasig, Ingolf E; Toborek, Michal

    2016-03-01

    HIV invades the brain early after infection; however, its interactions with the cells of the blood-brain barrier (BBB) remain poorly understood. Our goal was to evaluate the role of occludin, one of the tight junction proteins that regulate BBB functions in HIV infection of BBB pericytes. We provide evidence that occludin levels largely control the metabolic responses of human pericytes to HIV. Occludin in BBB pericytes decreased by 10% during the first 48 h after HIV infection, correlating with increased nuclear translocation of the gene repressor C-terminal-binding protein (CtBP)-1 and NFκB-p65 activation. These changes were associated with decreased expression and activation of the class III histone deacetylase sirtuin (SIRT)-1. Occludin levels recovered 96 h after infection, restoring SIRT-1 and reducing HIV transcription to 20% of its highest values. We characterized occludin biochemically as a novel NADH oxidase that controls the expression and activation of SIRT-1. The inverse correlation between occludin and HIV transcription was then replicated in human primary macrophages and differentiated monocytic U937 cells, in which occludin silencing resulted in 75 and 250% increased viral transcription, respectively. Our work shows that occludin has previously unsuspected metabolic properties and is a target of HIV infection, opening the possibility of designing novel pharmacological approaches to control HIV transcription.

  10. The Proteasome Activator PA28γ, a Negative Regulator of p53, Is Transcriptionally Up-Regulated by p53

    Directory of Open Access Journals (Sweden)

    Zhen-Xing Wan

    2014-02-01

    Full Text Available PA28γ (also called REGγ, 11Sγ or PSME3 negatively regulates p53 activity by promoting its nuclear export and/or degradation. Here, using the RNA ligase-mediated rapid amplification of cDNA ends (RLM-RACE method, we identified the transcription start site of the PA28γ gene. Assessment with the luciferase assay demonstrated that the sequence −193 to +16 is the basal promoter. Three p53 binding sites were found within the PA28γ promoter utilizing a bioinformatics approach and were confirmed by chromatin immunoprecipitation and biotinylated DNA affinity precipitation experiments. The p53 protein promotes PA28γ transcription, and p53-stimulated transcription of PA28γ can be inhibited by PA28γ itself. Our results suggest that PA28γ and p53 form a negative feedback loop, which maintains the balance of p53 and PA28γ in cells.

  11. Yap5 is an iron-responsive transcriptional activator that regulates vacuolar iron storage in yeast.

    Science.gov (United States)

    Li, Liangtao; Bagley, Dustin; Ward, Diane M; Kaplan, Jerry

    2008-02-01

    The transporter Ccc1 imports iron into the vacuole, which is the major site of iron storage in fungi and plants. CCC1 mRNA is destabilized under low-iron conditions by the binding of Cth1 and Cth2 to the 3' untranslated region (S. Puig, E. Askeland, and D. J. Thiele, Cell 120:99-110, 2005). Here, we show that the transcription of CCC1 is stimulated by iron through a Yap consensus site in the CCC1 promoter. We identified YAP5 as being the iron-sensitive transcription factor and show that a yap5Delta strain is sensitive to high iron. Green fluorescent protein-tagged Yap5 is localized to the nucleus and occupies the CCC1 promoter independent of the iron concentration. Yap5 contains two cysteine-rich domains, and the mutation of the cysteines to alanines in each of the domains affects the transcription of CCC1 but not DNA binding. The fusion of the Yap5 cysteine-containing domains to a GAL4 DNA binding domain results in iron-sensitive GAL1-lacZ expression. Iron affects the sulfhydryl status of Yap5, which is indicative of the generation of intramolecular disulfide bonds. These results show that Yap5 is an iron-sensing transcription factor and that iron regulates transcriptional activation.

  12. Yap5 Is an Iron-Responsive Transcriptional Activator That Regulates Vacuolar Iron Storage in Yeast▿

    Science.gov (United States)

    Li, Liangtao; Bagley, Dustin; Ward, Diane M.; Kaplan, Jerry

    2008-01-01

    The transporter Ccc1 imports iron into the vacuole, which is the major site of iron storage in fungi and plants. CCC1 mRNA is destabilized under low-iron conditions by the binding of Cth1 and Cth2 to the 3′ untranslated region (S. Puig, E. Askeland, and D. J. Thiele, Cell 120:99-110, 2005). Here, we show that the transcription of CCC1 is stimulated by iron through a Yap consensus site in the CCC1 promoter. We identified YAP5 as being the iron-sensitive transcription factor and show that a yap5Δ strain is sensitive to high iron. Green fluorescent protein-tagged Yap5 is localized to the nucleus and occupies the CCC1 promoter independent of the iron concentration. Yap5 contains two cysteine-rich domains, and the mutation of the cysteines to alanines in each of the domains affects the transcription of CCC1 but not DNA binding. The fusion of the Yap5 cysteine-containing domains to a GAL4 DNA binding domain results in iron-sensitive GAL1-lacZ expression. Iron affects the sulfhydryl status of Yap5, which is indicative of the generation of intramolecular disulfide bonds. These results show that Yap5 is an iron-sensing transcription factor and that iron regulates transcriptional activation. PMID:18070921

  13. A central region of Gli2 regulates its localization to the primary cilium and transcriptional activity.

    Science.gov (United States)

    Santos, Nicole; Reiter, Jeremy F

    2014-04-01

    Signaling through vertebrate Hedgehog (Hh) proteins depends on the primary cilium. In response to Hh signals, the transcriptional activator of the pathway, Gli2, accumulates at the ciliary tip, raising the possibility that ciliary localization is important for Gli2 activation. To test this hypothesis, we used the Floxin system to create knock-in Gli2 alleles in embryonic stem cells (ESCs) to allow methodical testing of which domains and residues are essential for the ciliary localization of Gli2. The Gli2 zinc fingers, transcriptional activation domain, repressor domain, phosphorylation cluster and a Sufu binding motif were each dispensable for ciliary localization. Mutating residues that are required for Gli2 sumoylation and nuclear trafficking also did not abrogate ciliary localization. By contrast, several other domains restricted Gli2 nuclear localization, and a central region, distinct from previously characterized domains, was required for ciliary localization. In addition to an inability to localize to cilia, Gli2 lacking this central domain was unable to activate target genes. Thus, our systematic analysis in ESCs reveals that distinct regions of Gli2 regulate its nuclear and ciliary localization. The identification of a domain essential for both ciliary localization and transcriptional activity suggests that ciliary localization of Gli2 is required for its activation.

  14. Locked and proteolysis-based transcription activator-like effector (TALE) regulation.

    Science.gov (United States)

    Lonzarić, Jan; Lebar, Tina; Majerle, Andreja; Manček-Keber, Mateja; Jerala, Roman

    2016-02-18

    Development of orthogonal, designable and adjustable transcriptional regulators is an important goal of synthetic biology. Their activity has been typically modulated through stimulus-induced oligomerization or interaction between the DNA-binding and activation/repression domain. We exploited a feature of the designable Transcription activator-like effector (TALE) DNA-binding domain that it winds around the DNA which allows to topologically prevent it from binding by intramolecular cyclization. This new approach was investigated through noncovalent ligand-induced cyclization or through a covalent split intein cyclization strategy, where the topological inhibition of DNA binding by cyclization and its restoration by a proteolytic release of the topologic constraint was expected. We show that locked TALEs indeed have diminished DNA binding and regain full transcriptional activity by stimulation with the rapamycin ligand or site-specific proteolysis of the peptide linker, with much higher level of activation than rapamycin-induced heterodimerization. Additionally, we demonstrated reversibility, activation of genomic targets and implemented logic gates based on combinations of protein cyclization, proteolytic cleavage and ligand-induced dimerization, where the strongest fold induction was achieved by the proteolytic cleavage of a repression domain from a linear TALE.

  15. Mechanosensitive mechanisms in transcriptional regulation.

    Science.gov (United States)

    Mammoto, Akiko; Mammoto, Tadanori; Ingber, Donald E

    2012-07-01

    Transcriptional regulation contributes to the maintenance of pluripotency, self-renewal and differentiation in embryonic cells and in stem cells. Therefore, control of gene expression at the level of transcription is crucial for embryonic development, as well as for organogenesis, functional adaptation, and regeneration in adult tissues and organs. In the past, most work has focused on how transcriptional regulation results from the complex interplay between chemical cues, adhesion signals, transcription factors and their co-regulators during development. However, chemical signaling alone is not sufficient to explain how three-dimensional (3D) tissues and organs are constructed and maintained through the spatiotemporal control of transcriptional activities. Accumulated evidence indicates that mechanical cues, which include physical forces (e.g. tension, compression or shear stress), alterations in extracellular matrix (ECM) mechanics and changes in cell shape, are transmitted to the nucleus directly or indirectly to orchestrate transcriptional activities that are crucial for embryogenesis and organogenesis. In this Commentary, we review how the mechanical control of gene transcription contributes to the maintenance of pluripotency, determination of cell fate, pattern formation and organogenesis, as well as how it is involved in the control of cell and tissue function throughout embryogenesis and adult life. A deeper understanding of these mechanosensitive transcriptional control mechanisms should lead to new approaches to tissue engineering and regenerative medicine.

  16. Cyclin C regulates adipogenesis by stimulating transcriptional activity of CCAAT/enhancer binding protein alpha.

    Science.gov (United States)

    Song, Ziyi; Xiaoli, Alus M; Zhang, Quanwei; Zhang, Yi; Yang, Ellen S T; Wang, Sven; Chang, Rui; Zhang, Zhengdong D; Yang, Gongshe; Strich, Randy; Pessin, Jeffrey E; Yang, Fajun

    2017-03-28

    Brown adipose tissue (BAT) is important for maintaining energy homeostasis and adaptive thermogenesis in rodents and humans. As disorders arising from dysregulated energy metabolism, such as obesity and metabolic diseases, have increased, so has interest in the molecular mechanisms in adipocyte biology. Using a functional screen, we identified cyclin C (CycC), a conserved subunit of the Mediator complex, as a novel regulator for brown adipocyte formation. siRNA-mediated CycC knockdown (KD) in brown preadipocytes impaired the early transcriptional program of differentiation, and genetic knockout (KO) of CycC completely blocked the differentiation process. RNA-seq analyses of CycC-KD revealed a critical role of CycC in activating genes co-regulated by peroxisome proliferator activated receptor gamma (PPARγ) and CCAAT/enhancer binding protein alpha (C/EBPα). Overexpression of PPARγ2 or addition of the PPARγ ligand rosiglitazone rescued the defects in CycC-KO brown preadipocytes, and efficiently activated the PPARγ-responsive promoters in both wild-type (WT) and CycC-KO cells, suggesting that CycC is not essential for PPARγ transcriptional activity. In contrast, CycC-KO significantly reduced C/EBPα-dependent gene expression. Unlike for PPARγ, overexpression of C/EBPα could not induce C/EBPα target gene expression in CycC-KO cells or rescue the CycC-KO defects in brown adipogenesis, suggesting that CycC is essential for C/EBPα-mediated gene activation. CycC physically interacted with C/EBPα and this interaction was required for C/EBPα transactivation domain activity. Consistent with the role of C/EBPα in white adipogenesis, CycC-KD also inhibited differentiation of 3T3-L1 cells into white adipocytes. Together, these data indicate that CycC activates adipogenesis by stimulating the transcriptional activity of C/EBPα.

  17. A role for RNA post-transcriptional regulation in satellite cell activation

    Directory of Open Access Journals (Sweden)

    Farina Nicholas H

    2012-10-01

    Full Text Available Abstract Background Satellite cells are resident skeletal muscle stem cells responsible for muscle maintenance and repair. In resting muscle, satellite cells are maintained in a quiescent state. Satellite cell activation induces the myogenic commitment factor, MyoD, and cell cycle entry to facilitate transition to a population of proliferating myoblasts that eventually exit the cycle and regenerate muscle tissue. The molecular mechanism involved in the transition of a quiescent satellite cell to a transit-amplifying myoblast is poorly understood. Methods Satellite cells isolated by FACS from uninjured skeletal muscle and 12 h post-muscle injury from wild type and Syndecan-4 null mice were probed using Affymetrix 430v2 gene chips and analyzed by Spotfiretm and Ingenuity Pathway analysis to identify gene expression changes and networks associated with satellite cell activation, respectively. Additional analyses of target genes identify miRNAs exhibiting dynamic changes in expression during satellite cell activation. The function of the miRNAs was assessed using miRIDIAN hairpin inhibitors. Results An unbiased gene expression screen identified over 4,000 genes differentially expressed in satellite cells in vivo within 12 h following muscle damage and more than 50% of these decrease dramatically. RNA binding proteins and genes involved in post-transcriptional regulation were significantly over-represented whereas splicing factors were preferentially downregulated and mRNA stability genes preferentially upregulated. Furthermore, six computationally identified miRNAs demonstrated novel expression through muscle regeneration and in satellite cells. Three of the six miRNAs were found to regulate satellite cell fate. Conclusions The quiescent satellite cell is actively maintained in a state poised to activate in response to external signals. Satellite cell activation appears to be regulated by post-transcriptional gene regulation.

  18. SUMOylation regulates the transcriptional repression activity of FOG-2 and its association with GATA-4.

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    José Perdomo

    Full Text Available Friend of GATA 2 (FOG-2, a co-factor of several GATA transcription factors (GATA-4, -5 and 6, is a critical regulator of coronary vessel formation and heart morphogenesis. Here we demonstrate that FOG-2 is SUMOylated and that this modification modulates its transcriptional activity. FOG-2 SUMOylation occurs at four lysine residues (K324, 471, 915, 955 [corrected]. Three of these residues are part of the characteristic SUMO consensus site (ψKXE, while K955 is found in the less frequent TKXE motif. Absence of SUMOylation did not affect FOG-2's nuclear localization. However, mutation of the FOG-2 SUMOylation sites, or de-SUMOylation, with SENP-1 or SENP-8 resulted in stronger transcriptional repression activity in both heterologous cells and cardiomyocytes. Conversely, increased FOG-2 SUMOylation by overexpression of SUMO-1 or expression of a SUMO-1-FOG-2 fusion protein rendered FOG-2 incapable of repressing GATA-4-mediated activation of the B-type natriuretic peptide (BNP promoter. Moreover, we demonstrate both increased interaction between a FOG-2 SUMO mutant and GATA-4 and enhanced SUMOylation of wild-type FOG-2 by co-expression of GATA-4. These data suggest a new dynamics in which GATA-4 may alter the activity of FOG-2 by influencing its SUMOylation status.

  19. A co-activator of nitrogen-regulated transcription in Saccharomyces cerevisiae.

    Science.gov (United States)

    Soussi-Boudekou, S; André, B

    1999-02-01

    In Saccharomyces cerevisiae, the transcription factors Gln3p and Nil1p of the GATA family play a determinant role in expression of genes that are subject to nitrogen catabolite repression. Here we report the isolation of a new yeast mutant, gan1-1, exhibiting dramatically decreased NAD-linked glutamate dehydrogenase (NAD-GDH) and glutamine synthetase (GS) activities. The GAN1 gene was cloned and found to encode a 488-amino-acid polypeptide bearing no typical DNA binding domain. Gan1p is required for full expression of GLN1, GDH2 and also other nitrogen utilization genes, including GAP1, PUT4, MEP2 and GDH1. The extent to which Gan1p is required, however, varies according to the gene and to the nitrogen source available. We show that Gan1p is in fact involved in Gln3p- and Nil1p-dependent transcription. In the case of Gln3p-dependent transcription, the degree to which Gan1p is required appears to be gene specific. The contribution of Gan1p to gene expression is also influenced by the nitrogen status of the cell. We found that GAN1 is identical to ADA1, which encodes a component of the ADA/GCN5 co-activator complex. Ada1/Gan1p thus represents the first reported case of an accessory protein (a co-activator) linking the GATA-binding proteins Gln3p and Nil1p, mediating nitrogen-regulated transcription, to the basal transcription machinery.

  20. Activated α2-Macroglobulin Regulates Transcriptional Activation of c-MYC Target Genes through Cell Surface GRP78 Protein.

    Science.gov (United States)

    Gopal, Udhayakumar; Gonzalez-Gronow, Mario; Pizzo, Salvatore Vincent

    2016-05-13

    Activated α2-macroglobulin (α2M*) signals predominantly through cell surface GRP78 (CS-GRP78) to promote proliferation and survival of cancer cells; however, the molecular mechanism remains obscure. c-MYC is an essential transcriptional regulator that controls cell proliferation. We hypothesize that α2M*/CS-GRP78-evoked key signaling events are required for transcriptional activation of c-MYC target genes. Activation of CS-GRP78 by α2M* requires ligation of the GRP78 primary amino acid sequence (Leu(98)-Leu(115)). After stimulation with α2M*, CS-GRP78 signaling activates 3-phosphoinositide-dependent protein kinase-1 (PDK1) to induce phosphorylation of PLK1, which in turn induces c-MYC transcription. We demonstrate that PLK1 binds directly to c-MYC and promotes its transcriptional activity by phosphorylating Ser(62) Moreover, activated c-MYC is recruited to the E-boxes of target genes FOSL1 and ID2 by phosphorylating histone H3 at Ser(10) In addition, targeting the carboxyl-terminal domain of CS-GRP78 with a mAb suppresses transcriptional activation of c-MYC target genes and impairs cell proliferation. This work demonstrates that α2M*/CS-GRP78 acts as an upstream regulator of the PDK1/PLK1 signaling axis to modulate c-MYC transcription and its target genes, suggesting a therapeutic strategy for targeting c-MYC-associated malignant progression. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Activated α2-Macroglobulin Regulates Transcriptional Activation of c-MYC Target Genes through Cell Surface GRP78 Protein*

    Science.gov (United States)

    Gopal, Udhayakumar; Gonzalez-Gronow, Mario; Pizzo, Salvatore Vincent

    2016-01-01

    Activated α2-macroglobulin (α2M*) signals predominantly through cell surface GRP78 (CS-GRP78) to promote proliferation and survival of cancer cells; however, the molecular mechanism remains obscure. c-MYC is an essential transcriptional regulator that controls cell proliferation. We hypothesize that α2M*/CS-GRP78-evoked key signaling events are required for transcriptional activation of c-MYC target genes. Activation of CS-GRP78 by α2M* requires ligation of the GRP78 primary amino acid sequence (Leu98–Leu115). After stimulation with α2M*, CS-GRP78 signaling activates 3-phosphoinositide-dependent protein kinase-1 (PDK1) to induce phosphorylation of PLK1, which in turn induces c-MYC transcription. We demonstrate that PLK1 binds directly to c-MYC and promotes its transcriptional activity by phosphorylating Ser62. Moreover, activated c-MYC is recruited to the E-boxes of target genes FOSL1 and ID2 by phosphorylating histone H3 at Ser10. In addition, targeting the carboxyl-terminal domain of CS-GRP78 with a mAb suppresses transcriptional activation of c-MYC target genes and impairs cell proliferation. This work demonstrates that α2M*/CS-GRP78 acts as an upstream regulator of the PDK1/PLK1 signaling axis to modulate c-MYC transcription and its target genes, suggesting a therapeutic strategy for targeting c-MYC-associated malignant progression. PMID:27002159

  2. Sumoylation regulates the transcriptional activity of MafA in pancreatic beta cells.

    Science.gov (United States)

    Shao, Chunli; Cobb, Melanie H

    2009-01-30

    MafA is a transcriptional regulator expressed primarily in pancreatic beta cells. It binds to the RIPE3b/C1-binding site within the ins gene promoter, which plays a critical role in regulating ins gene expression in response to glucose. Here, we show that MafA is post-translationally modified by the small ubiquitin-related modifiers SUMO-1 and -2. Mutation of a single site in MafA, Lys(32), blocks its sumoylation in beta cells. Incubation of beta cells in low glucose (2 mm) or exposure to hydrogen peroxide increases sumoylation of endogenous MafA. Forced sumoylation of MafA results in reduced transcriptional activity toward the ins gene promoter and increased suppression of the CHOP-10 gene promoter. Sumoylation of MafA has no apparent effect on either its nuclear localization in beta cells or its ubiquitin-dependent degradation. This study suggests that modification of MafA by SUMO modulates gene transcription and thereby beta cell function.

  3. Computational inference of replication and transcription activator regulator activity in herpesvirus from gene expression data

    OpenAIRE

    Recchia, A; Wit, E; Vinciotti, V; Kellam, P

    2008-01-01

    One of the main aims of system biology is to understand the structure and dynamics of genomic systems. A computational approach, facilitated by new technologies for high-throughput quantitative experimental data, is put forward to investigate the regulatory system of dynamic interaction among genes in Kaposi's sarcoma-associated herpesvirus network after induction of lytic replication. A reconstruction of transcription factor activity and gene-regulatory kinetics using data from a time-course...

  4. Statins Increase Plasminogen Activator Inhibitor Type 1 Gene Transcription through a Pregnane X Receptor Regulated Element.

    Directory of Open Access Journals (Sweden)

    Frederick M Stanley

    Full Text Available Plasminogen activator inhibitor type 1 (PAI-1 is a multifunctional protein that has important roles in inflammation and wound healing. Its aberrant regulation may contribute to many disease processes such as heart disease. The PAI-1 promoter is responsive to multiple inputs including cytokines, growth factors, steroids and oxidative stress. The statin drugs, atorvastatin, mevastatin and rosuvastatin, increased basal and stimulated expression of the PAI-1 promoter 3-fold. A statin-responsive, nuclear hormone response element was previously identified in the PAI-1 promoter, but it was incompletely characterized. We characterized this direct repeat (DR of AGGTCA with a 3-nucleotide spacer at -269/-255 using deletion and directed mutagenesis. Deletion or mutation of this element increased basal transcription from the promoter suggesting that it repressed PAI-1 transcription in the unliganded state. The half-site spacing and the ligand specificity suggested that this might be a pregnane X receptor (PXR responsive element. Computational molecular docking showed that atorvastatin, mevastatin and rosuvastatin were structurally compatible with the PXR ligand-binding pocket in its agonist conformation. Experiments with Gal4 DNA binding domain fusion proteins showed that Gal4-PXR was activated by statins while other DR + 3 binding nuclear receptor fusions were not. Overexpression of PXR further enhanced PAI-1 transcription in response to statins. Finally, ChIP experiments using Halo-tagged PXR and RXR demonstrated that both components of the PXR-RXR heterodimer bound to this region of the PAI-1 promoter.

  5. AMP-Activated Protein Kinase Regulates Oxidative Metabolism in Caenorhabditis elegans through the NHR-49 and MDT-15 Transcriptional Regulators.

    Science.gov (United States)

    Moreno-Arriola, Elizabeth; El Hafidi, Mohammed; Ortega-Cuéllar, Daniel; Carvajal, Karla

    2016-01-01

    Cellular energy regulation relies on complex signaling pathways that respond to fuel availability and metabolic demands. Dysregulation of these networks is implicated in the development of human metabolic diseases such as obesity and metabolic syndrome. In Caenorhabditis elegans the AMP-activated protein kinase, AAK, has been associated with longevity and stress resistance; nevertheless its precise role in energy metabolism remains elusive. In the present study, we find an evolutionary conserved role of AAK in oxidative metabolism. Similar to mammals, AAK is activated by AICAR and metformin and leads to increased glycolytic and oxidative metabolic fluxes evidenced by an increase in lactate levels and mitochondrial oxygen consumption and a decrease in total fatty acids and lipid storage, whereas augmented glucose availability has the opposite effects. We found that these changes were largely dependent on the catalytic subunit AAK-2, since the aak-2 null strain lost the observed metabolic actions. Further results demonstrate that the effects due to AAK activation are associated to SBP-1 and NHR-49 transcriptional factors and MDT-15 transcriptional co-activator, suggesting a regulatory pathway that controls oxidative metabolism. Our findings establish C. elegans as a tractable model system to dissect the relationship between distinct molecules that play a critical role in the regulation of energy metabolism in human metabolic diseases.

  6. Liver x receptors regulate the transcriptional activity of the glucocorticoid receptor: implications for the carbohydrate metabolism.

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    Nancy Nader

    Full Text Available GLUCOCORTICOIDS are steroid hormones that strongly influence intermediary carbohydrate metabolism by increasing the transcription rate of glucose-6-phosphatase (G6Pase, a key enzyme of gluconeogenesis, and suppress the immune system through the glucocorticoid receptor (GR. The liver X receptors (LXRs, on the other hand, bind to cholesterol metabolites, heterodimerize with the retinoid X receptor (RXR, and regulate the cholesterol turnover, the hepatic glucose metabolism by decreasing the expression of G6Pase, and repress a set of inflammatory genes in immune cells. Since the actions of these receptors overlap with each other, we evaluated the crosstalk between the GR- and LXR-mediated signaling systems. Transient transfection-based reporter assays and gene silencing methods using siRNAs for LXRs showed that overexpression/ligand (GW3965 activation of LXRs/RXRs repressed GR-stimulated transactivation of certain glucocorticoid response element (GRE-driven promoters in a gene-specific fashion. Activation of LXRs by GW3965 attenuated dexamethasone-stimulated elevation of circulating glucose in rats. It also suppressed dexamethasone-induced mRNA expression of hepatic glucose-6-phosphatase (G6Pase in rats, mice and human hepatoma HepG2 cells, whereas endogenous, unliganded LXRs were required for dexamethasone-induced mRNA expression of phosphoenolpyruvate carboxylase. In microarray transcriptomic analysis of rat liver, GW3965 differentially regulated glucocorticoid-induced transcriptional activity of about 15% of endogenous glucocorticoid-responsive genes. To examine the mechanism through which activated LXRs attenuated GR transcriptional activity, we examined LXRα/RXRα binding to GREs. Endogenous LXRα/RXRα bound GREs and inhibited GR binding to these DNA sequences both in in vitro and in vivo chromatin immunoprecipitation assays, while their recombinant proteins did so on classic or G6Pase GREs in gel mobility shift assays. We propose that

  7. Liver X Receptors Regulate the Transcriptional Activity of the Glucocorticoid Receptor: Implications for the Carbohydrate Metabolism

    Science.gov (United States)

    Nader, Nancy; Ng, Sinnie Sin Man; Wang, Yonghong; Abel, Brent S.; Chrousos, George P.; Kino, Tomoshige

    2012-01-01

    GLUCOCORTICOIDS are steroid hormones that strongly influence intermediary carbohydrate metabolism by increasing the transcription rate of glucose-6-phosphatase (G6Pase), a key enzyme of gluconeogenesis, and suppress the immune system through the glucocorticoid receptor (GR). The liver X receptors (LXRs), on the other hand, bind to cholesterol metabolites, heterodimerize with the retinoid X receptor (RXR), and regulate the cholesterol turnover, the hepatic glucose metabolism by decreasing the expression of G6Pase, and repress a set of inflammatory genes in immune cells. Since the actions of these receptors overlap with each other, we evaluated the crosstalk between the GR- and LXR-mediated signaling systems. Transient transfection-based reporter assays and gene silencing methods using siRNAs for LXRs showed that overexpression/ligand (GW3965) activation of LXRs/RXRs repressed GR-stimulated transactivation of certain glucocorticoid response element (GRE)-driven promoters in a gene-specific fashion. Activation of LXRs by GW3965 attenuated dexamethasone-stimulated elevation of circulating glucose in rats. It also suppressed dexamethasone-induced mRNA expression of hepatic glucose-6-phosphatase (G6Pase) in rats, mice and human hepatoma HepG2 cells, whereas endogenous, unliganded LXRs were required for dexamethasone-induced mRNA expression of phosphoenolpyruvate carboxylase. In microarray transcriptomic analysis of rat liver, GW3965 differentially regulated glucocorticoid-induced transcriptional activity of about 15% of endogenous glucocorticoid-responsive genes. To examine the mechanism through which activated LXRs attenuated GR transcriptional activity, we examined LXRα/RXRα binding to GREs. Endogenous LXRα/RXRα bound GREs and inhibited GR binding to these DNA sequences both in in vitro and in vivo chromatin immunoprecipitation assays, while their recombinant proteins did so on classic or G6Pase GREs in gel mobility shift assays. We propose that administration of

  8. HBXIP up-regulates ACSL1 through activating transcriptional factor Sp1 in breast cancer.

    Science.gov (United States)

    Wang, Yue; Cai, Xiaoli; Zhang, Shuqin; Cui, Ming; Liu, Fabao; Sun, Baodi; Zhang, Weiying; Zhang, Xiaodong; Ye, Lihong

    2017-03-11

    The oncoprotein hepatitis B X-interacting protein (HBXIP) results in the dysregulation of lipid metabolism to enhance the development of breast cancer. Acyl-CoA synthetase long-chain family member 1 (ACSL1) is required for thioesterification of long-chain fatty acids into their acyl-CoA derivatives. In this study, we present a hypothesis that HBXIP might be involved in the regulation of ACSL1 in breast cancer. Interestingly, we found that the overexpression of HBXIP was able to up-regulate ACSL1 at the levels of mRNA and protein in a dose-dependent manner in breast cancer cells. Conversely, silencing of HBXIP led to the opposite results. Mechanistically, HBXIP as a coactivator interacted with transcriptional factor Sp1 through binding to the promoter of ACSL1 by ChIP assays analysis, leading to the transcription of ACSL1 in breast cancer cells. Immunohistochemistry staining revealed that the positive rate of ACSL1 was 71.4% (35/49) in clinical breast cancer tissues, HBXIP 79.6% (39/49), in which the positive rate of ACSL1 was 76.9% (30/39) in the HBXIP-positive specimens. But, few positive rate of ACSL1 10% (1/10) was observed in normal breast tissues. The mRNA levels of ACSL1 were significantly higher in clinical breast cancer tissues than those in their corresponding peritumor tissues. The mRNA levels of ACSL1 were positively associated with those of HBXIP in clinical breast cancer tissues. Thus, we conclude that the oncoprotein HBXIP is able to up-regulate ACSL1 through activating the transcriptional factor Sp1 in breast cancer.

  9. Regulation of WRKY46 transcription factor function by mitogen-activated protein kinases in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Arsheed Hussain Sheikh

    2016-02-01

    Full Text Available AbstractMitogen-activated protein kinase (MAPK cascades are central signalling pathways activated in plants after sensing internal developmental and external stress cues. Knowledge about the downstream substrate proteins of MAPKs is still limited in plants. We screened Arabidopsis WRKY transcription factors as potential targets downstream of MAPKs, and concentrated on characterizing WRKY46 as a substrate of the MAPK, MPK3. Mass spectrometry revealed in vitro phosphorylation of WRKY46 at amino acid position S168 by MPK3. However, mutagenesis studies showed that a second phosphosite, S250, can also be phosphorylated. Elicitation with pathogen-associated molecular patterns (PAMPs, such as the bacterial flagellin-derived flg22 peptide led to in vivo destabilization of WRKY46 in Arabidopsis protoplasts. Mutation of either phosphorylation site reduced the PAMP-induced degradation of WRKY46. Furthermore, the protein for the double phosphosite mutant is expressed at higher levels compared to wild-type proteins or single phosphosite mutants. In line with its nuclear localization and predicted function as a transcriptional activator, overexpression of WRKY46 in protoplasts raised basal plant defence as reflected by the increase in promoter activity of the PAMP-responsive gene, NHL10, in a MAPK-dependent manner. Thus, MAPK-mediated regulation of WRKY46 is a mechanism to control plant defence.

  10. Stathmin regulates mutant p53 stability and transcriptional activity in ovarian cancer.

    Science.gov (United States)

    Sonego, Maura; Schiappacassi, Monica; Lovisa, Sara; Dall'Acqua, Alessandra; Bagnoli, Marina; Lovat, Francesca; Libra, Massimo; D'Andrea, Sara; Canzonieri, Vincenzo; Militello, Loredana; Napoli, Marco; Giorda, Giorgio; Pivetta, Barbara; Mezzanzanica, Delia; Barbareschi, Mattia; Valeri, Barbara; Canevari, Silvana; Colombatti, Alfonso; Belletti, Barbara; Del Sal, Giannino; Baldassarre, Gustavo

    2013-05-01

    Stathmin is a p53-target gene, frequently overexpressed in late stages of human cancer progression. Type II High Grade Epithelial Ovarian Carcinomas (HG-EOC) represents the only clear exception to this observation. Here, we show that stathmin expression is necessary for the survival of HG-EOC cells carrying a p53 mutant (p53(MUT) ) gene. At molecular level, stathmin favours the binding and the phosphorylation of p53(MUT) by DNA-PKCS , eventually modulating p53(MUT) stability and transcriptional activity. Inhibition of stathmin or DNA-PKCS impaired p53(MUT) -dependent transcription of several M phase regulators, resulting in M phase failure and EOC cell death, both in vitro and in vivo. In primary human EOC a strong correlation exists between stathmin, DNA-PKCS , p53(MUT) overexpression and its transcriptional targets, further strengthening the relevance of the new pathway here described. Overall our data support the hypothesis that the expression of stathmin and p53 could be useful for the identification of high risk patients that will benefit from a therapy specifically acting on mitotic cancer cells.

  11. Neuronal activity rapidly induces transcription of the CREB-regulated microRNA-132, in vivo.

    Science.gov (United States)

    Nudelman, Aaron S; DiRocco, Derek P; Lambert, Talley J; Garelick, Michael G; Le, Josh; Nathanson, Neil M; Storm, Daniel R

    2010-04-01

    Activity-dependent changes in gene-expression are believed to underlie the molecular representation of memory. In this study, we report that in vivo activation of neurons rapidly induces the CREB-regulated microRNA miR-132. To determine if production of miR-132 is regulated by neuronal activity its expression in mouse brain was monitored by quantitative RT-PCR (RT-qPCR). Pilocarpine-induced seizures led to a robust, rapid, and transient increase in the primary transcript of miR-132 (pri-miR-132) followed by a subsequent rise in mature microRNA (miR-132). Activation of neurons in the hippocampus, olfactory bulb, and striatum by contextual fear conditioning, odor-exposure, and cocaine-injection, respectively, also increased pri-miR-132. Induction kinetics of pri-miR-132 were monitored and found to parallel those of immediate early genes, peaking at 45 min and returning to basal levels within 2 h of stimulation. Expression levels of primary and mature-miR-132 increased significantly between postnatal Days 10 and 24. We conclude that miR-132 is an activity-dependent microRNA in vivo, and may contribute to the long-lasting proteomic changes required for experience-dependent neuronal plasticity.

  12. Neuronal activity rapidly induces transcription of the CREB-regulated microRNA-132, in vivo

    DEFF Research Database (Denmark)

    Nudelman, Aaron Samuel; DiRocco, Derek P; Lambert, Talley J

    2010-01-01

    of stimulation. Expression levels of primary and mature-miR-132 increased significantly between postnatal Days 10 and 24. We conclude that miR-132 is an activity-dependent microRNA in vivo, and may contribute to the long-lasting proteomic changes required for experience-dependent neuronal plasticity.......Activity-dependent changes in gene-expression are believed to underlie the molecular representation of memory. In this study, we report that in vivo activation of neurons rapidly induces the CREB-regulated microRNA miR-132. To determine if production of miR-132 is regulated by neuronal activity its...... expression in mouse brain was monitored by quantitative RT-PCR (RT-qPCR). Pilocarpine-induced seizures led to a robust, rapid, and transient increase in the primary transcript of miR-132 (pri-miR-132) followed by a subsequent rise in mature microRNA (miR-132). Activation of neurons in the hippocampus...

  13. RNA exosome-regulated long non-coding RNA transcription controls super-enhancer activity.

    Science.gov (United States)

    Pefanis, Evangelos; Wang, Jiguang; Rothschild, Gerson; Lim, Junghyun; Kazadi, David; Sun, Jianbo; Federation, Alexander; Chao, Jaime; Elliott, Oliver; Liu, Zhi-Ping; Economides, Aris N; Bradner, James E; Rabadan, Raul; Basu, Uttiya

    2015-05-01

    We have ablated the cellular RNA degradation machinery in differentiated B cells and pluripotent embryonic stem cells (ESCs) by conditional mutagenesis of core (Exosc3) and nuclear RNase (Exosc10) components of RNA exosome and identified a vast number of long non-coding RNAs (lncRNAs) and enhancer RNAs (eRNAs) with emergent functionality. Unexpectedly, eRNA-expressing regions accumulate R-loop structures upon RNA exosome ablation, thus demonstrating the role of RNA exosome in resolving deleterious DNA/RNA hybrids arising from active enhancers. We have uncovered a distal divergent eRNA-expressing element (lncRNA-CSR) engaged in long-range DNA interactions and regulating IgH 3' regulatory region super-enhancer function. CRISPR-Cas9-mediated ablation of lncRNA-CSR transcription decreases its chromosomal looping-mediated association with the IgH 3' regulatory region super-enhancer and leads to decreased class switch recombination efficiency. We propose that the RNA exosome protects divergently transcribed lncRNA expressing enhancers by resolving deleterious transcription-coupled secondary DNA structures, while also regulating long-range super-enhancer chromosomal interactions important for cellular function.

  14. Negative regulation of TLR-signaling pathways by activating transcription factor-3.

    Science.gov (United States)

    Whitmore, Mark M; Iparraguirre, Amaya; Kubelka, Lindsey; Weninger, Wolfgang; Hai, Tsonwin; Williams, Bryan R G

    2007-09-15

    Activating transcription factor-3 (ATF3) is rapidly induced by LPS in mouse macrophages and regulates TLR4 responses. We show that ATF3 is rapidly induced by various TLRs in mouse macrophages and plasmacytoid dendritic cells (DCs), as well as plasmacytoid and myeloid subsets of human DCs. In primary macrophages from mice with a targeted deletion of the atf3 gene (ATF3-knockout (KO)), TLR-stimulated levels of IL-12 and IL-6 were elevated relative to responses in wild-type macrophages. Similarly, targeted deletion of atf3 correlated with enhanced responsiveness of myeloid DCs to TLR activation as measured by IL-12 secretion. Ectopic expression of ATF3 antagonized TLR-stimulated IL-12p40 activation in a reporter assay. In vivo, CpG-oligodeoxynucleotide, a TLR9 agonist, given i.p. to ATF3-KO mice resulted in enhanced cytokine production from splenocytes. Furthermore, while ATF3-KO mice challenged with a sublethal dose of PR8 influenza virus were delayed in body weight recovery in comparison to wild type, the ATF3-KO mice showed higher titers of serum neutralizing Ab against PR8 5 mo postinfection. Thus, ATF3 behaves as a negative regulatory transcription factor in TLR pathways and, accordingly, deficiency in atf3 alters responses to immunological challenges in vivo. ATF3 dysregulation merits further exploration in diseases such as type I diabetes and cancer, where altered innate immunity has been implicated in their pathogenesis.

  15. Ribbon regulates morphogenesis of the Drosophila embryonic salivary gland through transcriptional activation and repression.

    Science.gov (United States)

    Loganathan, Rajprasad; Lee, Joslynn S; Wells, Michael B; Grevengoed, Elizabeth; Slattery, Matthew; Andrew, Deborah J

    2016-01-01

    Transcription factors affect spatiotemporal patterns of gene expression often regulating multiple aspects of tissue morphogenesis, including cell-type specification, cell proliferation, cell death, cell polarity, cell shape, cell arrangement and cell migration. In this work, we describe a distinct role for Ribbon (Rib) in controlling cell shape/volume increases during elongation of the Drosophila salivary gland (SG). Notably, the morphogenetic changes in rib mutants occurred without effects on general SG cell attributes such as specification, proliferation and apoptosis. Moreover, the changes in cell shape/volume in rib mutants occurred without compromising epithelial-specific morphological attributes such as apicobasal polarity and junctional integrity. To identify the genes regulated by Rib, we performed ChIP-seq analysis in embryos driving expression of GFP-tagged Rib specifically in the SGs. To learn if the Rib binding sites identified in the ChIP-seq analysis were linked to changes in gene expression, we performed microarray analysis comparing RNA samples from age-matched wild-type and rib null embryos. From the superposed ChIP-seq and microarray gene expression data, we identified 60 genomic sites bound by Rib likely to regulate SG-specific gene expression. We confirmed several of the identified Rib targets by qRT-pCR and/or in situ hybridization. Our results indicate that Rib regulates cell growth and tissue shape in the Drosophila salivary gland via a diverse array of targets through both transcriptional activation and repression. Furthermore, our results suggest that autoregulation of rib expression may be a key component of the SG morphogenetic gene network.

  16. Three promoters regulate the transcriptional activity of the human holocarboxylase synthetase gene.

    Science.gov (United States)

    Xia, Mengna; Malkaram, Sridhar A; Zempleni, Janos

    2013-11-01

    Holocarboxylase synthetase (HLCS) is the only protein biotin ligase in the human proteome. HLCS-dependent biotinylation of carboxylases plays crucial roles in macronutrient metabolism. HLCS appears to be an essential part of multiprotein complexes in the chromatin that cause gene repression and contribute toward genome stability. Consistent with these essential functions, HLCS knockdown causes strong phenotypes including shortened life span and low stress resistance in Drosophila melanogaster, and de-repression of long-terminal repeats in humans, other mammalian cell lines and Drosophila. Despite previous observations that the expression of HLCS depends on biotin status in rats and in human cell lines, little is known about the regulation of HLCS expression. The goal of this study was to identify promoters that regulate the expression of the human HLCS gene. Initially, the human HLCS locus was interrogated in silico using predictors of promoters including sequences of HLCS mRNA and expressed sequence tags, CpG islands, histone marks denoting transcriptionally poised chromatin, transcription factor binding sites and DNaseI hypersensitive regions. Our predictions revealed three putative HLCS promoters, denoted P1, P2 and P3. Promoters lacked a TATA box, which is typical for housekeeping genes. When the three promoters were cloned into a luciferase reporter plasmid, reporter gene activity was at least three times background noise in human breast, colon and kidney cell lines; activities consistently followed the pattern P1>P3>P2. Promoter activity depended on the concentration of biotin in culture media, but the effect was moderate. We conclude that we have identified promoters in the human HLCS gene.

  17. Transcriptional regulator-mediated activation of adaptation genes triggers CRISPR de novo spacer acquisition

    DEFF Research Database (Denmark)

    Liu, Tao; Li, Yingjun; Wang, Xiaodi;

    2015-01-01

    , it was demonstrated that the transcription level of csa1, cas1, cas2 and cas4 was significantly enhanced in a csa3a-overexpression strain and, moreover, the Csa1 and Cas1 protein levels were increased in this strain. Furthermore, we demonstrated the hyperactive uptake of unique spacers within both CRISPR loci......Acquisition of de novo spacer sequences confers CRISPR-Cas with a memory to defend against invading genetic elements. However, the mechanism of regulation of CRISPR spacer acquisition remains unknown. Here we examine the transcriptional regulation of the conserved spacer acquisition genes in Type I...... in the presence of the csa3a overexpression vector. The spacer acquisition process is dependent on the CCN PAM sequence and protospacer selection is random and non-directional. These results suggested a regulation mechanism of CRISPR spacer acquisition where a single transcriptional regulator senses the presence...

  18. Transcriptional regulator-mediated activation of adaptation genes triggers CRISPR de novo spacer acquisition

    DEFF Research Database (Denmark)

    Liu, Tao; Li, Yingjun; Wang, Xiaodi

    2015-01-01

    Acquisition of de novo spacer sequences confers CRISPR-Cas with a memory to defend against invading genetic elements. However, the mechanism of regulation of CRISPR spacer acquisition remains unknown. Here we examine the transcriptional regulation of the conserved spacer acquisition genes in Type I......, it was demonstrated that the transcription level of csa1, cas1, cas2 and cas4 was significantly enhanced in a csa3a-overexpression strain and, moreover, the Csa1 and Cas1 protein levels were increased in this strain. Furthermore, we demonstrated the hyperactive uptake of unique spacers within both CRISPR loci...... in the presence of the csa3a overexpression vector. The spacer acquisition process is dependent on the CCN PAM sequence and protospacer selection is random and non-directional. These results suggested a regulation mechanism of CRISPR spacer acquisition where a single transcriptional regulator senses the presence...

  19. Regulating retrotransposon activity through the use of alternative transcription start sites

    DEFF Research Database (Denmark)

    Persson, Jenna; Steglich, Babett; Smialowska, Agata;

    2016-01-01

    a new mechanism of retrotransposon regulation through transcription start site (TSS) selection by altered nucleosome occupancy. We show that Fun30 chromatin remodelers cooperate to maintain a high level of nucleosome occupancy at retrotransposon-flanking long terminal repeat (LTR) elements....... This enforces the use of a downstream TSS and the production of a truncated RNA incapable of reverse transcription and retrotransposition. However, in stressed cells, nucleosome occupancy at LTR elements is reduced, and the TSS shifts to allow for productive transcription. We propose that controlled...

  20. Deciphering Transcriptional Regulation

    DEFF Research Database (Denmark)

    Valen, Eivind

    RNA); and ii) translation, in which the mRNA is translated into a protein. This thesis focus on the ¿rst of these steps, transcription, and speci¿cally the initiation of this. Simpli¿ed, initiation is preceded by the binding of several proteins, known as transcription factors (TFs), to DNA. This takes place......The myriad of cells in the human body are all made from the same blueprint: the human genome. At the heart of this diversity lies the concept of gene regulation, the process in which it is decided which genes are used where and when. Genes do not function as on/off buttons, but more like a volume...... control spanning the range from completely muted to cranked up to maximum. The volume, in this case, is the production rate of proteins. This production is the result of a two step procedure: i) transcription, in which a small part of DNA from the genome (a gene) is transcribed into an RNA molecule (an m...

  1. Epidermal growth factor receptor regulates β-catenin location, stability, and transcriptional activity in oral cancer

    Directory of Open Access Journals (Sweden)

    Hung Hsing-Wen

    2010-03-01

    Full Text Available Abstract Background Many cancerous cells accumulate β-catenin in the nucleus. We examined the role of epidermal growth factor receptor (EGFR signaling in the accumulation of β-catenin in the nuclei of oral cancer cells. Results We used two strains of cultured oral cancer cells, one with reduced EGFR expression (OECM1 cells and one with elevated EGFR expression (SAS cells, and measured downstream effects, such as phosphorylation of β-catenin and GSK-3β, association of β-catenin with E-cadherin, and target gene regulation. We also studied the expression of EGFR, β-catenin, and cyclin D1 in 112 samples of oral cancer by immunostaining. Activation of EGFR signaling increased the amount of β-catenin in the nucleus and decreased the amount in the membranes. EGF treatment increased phosphorylation of β-catenin (tyrosine and GSK-3β(Ser-(9, resulting in a loss of β-catenin association with E-cadherin. TOP-FLASH and FOP-FLASH reporter assays demonstrated that the EGFR signal regulates β-catenin transcriptional activity and mediates cyclin D1 expression. Chromatin immunoprecipitation experiments indicated that the EGFR signal affects chromatin architecture at the regulatory element of cyclin D1, and that the CBP, HDAC1, and Suv39h1 histone/chromatin remodeling complex is involved in this process. Immunostaining showed a significant association between EGFR expression and aberrant accumulation of β-catenin in oral cancer. Conclusions EGFR signaling regulates β-catenin localization and stability, target gene expression, and tumor progression in oral cancer. Moreover, our data suggest that aberrant accumulation of β-catenin under EGFR activation is a malignancy marker of oral cancer.

  2. Analysis of Polygenic Mutants Suggests a Role for Mediator in Regulating Transcriptional Activation Distance in Saccharomyces cerevisiae.

    Science.gov (United States)

    Reavey, Caitlin T; Hickman, Mark J; Dobi, Krista C; Botstein, David; Winston, Fred

    2015-10-01

    Studies of natural populations of many organisms have shown that traits are often complex, caused by contributions of mutations in multiple genes. In contrast, genetic studies in the laboratory primarily focus on studying the phenotypes caused by mutations in a single gene. However, the single mutation approach may be limited with respect to the breadth and degree of new phenotypes that can be found. We have taken the approach of isolating complex, or polygenic mutants in the lab to study the regulation of transcriptional activation distance in yeast. While most aspects of eukaryotic transcription are conserved from yeast to human, transcriptional activation distance is not. In Saccharomyces cerevisiae, the upstream activating sequence (UAS) is generally found within 450 base pairs of the transcription start site (TSS) and when the UAS is moved too far away, activation no longer occurs. In contrast, metazoan enhancers can activate from as far as several hundred kilobases from the TSS. Previously, we identified single mutations that allow transcription activation to occur at a greater-than-normal distance from the GAL1 UAS. As the single mutant phenotypes were weak, we have now isolated polygenic mutants that possess strong long-distance phenotypes. By identification of the causative mutations we have accounted for most of the heritability of the phenotype in each strain and have provided evidence that the Mediator coactivator complex plays both positive and negative roles in the regulation of transcription activation distance.

  3. SUMOylation of the KRAB zinc-finger transcription factor PARIS/ZNF746 regulates its transcriptional activity

    Energy Technology Data Exchange (ETDEWEB)

    Nishida, Tamotsu, E-mail: nishida@gene.mie-u.ac.jp; Yamada, Yoshiji

    2016-05-13

    Parkin-interacting substrate (PARIS), a member of the family of Krüppel-associated box (KRAB)-containing zinc-finger transcription factors, is a substrate of the ubiquitin E3 ligase parkin. PARIS represses the expression of peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), although the underlying mechanisms remain largely unknown. In the present study, we demonstrate that PARIS can be SUMOylated, and its SUMOylation plays a role in the repression of PGC-1a promoter activity. Protein inhibitor of activated STAT y (PIASy) was identified as an interacting protein of PARIS and shown to enhance its SUMOylation. PIASy repressed PGC-1a promoter activity, and this effect was attenuated by PARIS in a manner dependent on its SUMOylation status. Co-expression of SUMO-1 with PIASy completely repressed PGC-1a promoter activity independently of PARIS expression. PARIS-mediated PGC-1a promoter repression depended on the activity of histone deacetylases (HDAC), whereas PIASy repressed the PGC-1a promoter in an HDAC-independent manner. Taken together, these results suggest that PARIS and PIASy modulate PGC-1a gene transcription through distinct molecular mechanisms. -- Highlights: •PARIS can be SUMOylated in vivo and in vitro. •SUMOylation of PARIS functions in the repression of PGC-1a promoter activity. •PIASy interacts with PARIS and enhances its SUMOylation. •PIASy influences PARIS-mediated repression of PGC-1a promoter activity.

  4. Transcriptional activation of human CDCA8 gene regulated by transcription factor NF-Y in embryonic stem cells and cancer cells.

    Science.gov (United States)

    Dai, Can; Miao, Cong-Xiu; Xu, Xiao-Ming; Liu, Lv-Jun; Gu, Yi-Fan; Zhou, Di; Chen, Lian-Sheng; Lin, Ge; Lu, Guang-Xiu

    2015-09-11

    The cell division cycle associated 8 (CDCA8) gene plays an important role in mitosis. Overexpression of CDCA8 was reported in some human cancers and is required for cancer growth and progression. We found CDCA8 expression was also high in human ES cells (hESCs) but dropped significantly upon hESC differentiation. However, the regulation of CDCA8 expression has not yet been studied. Here, we characterized the CDCA8 promoter and identified its cis-elements and transcription factors. Three transcription start sites were identified. Reporter gene assays revealed that the CDCA8 promoter was activated in hESCs and cancer cell lines. The promoter drove the reporter expression specifically to pluripotent cells during early mouse embryo development and to tumor tissues in tumor-bearing mice. These results indicate that CDCA8 is transcriptionally activated in hESCs and cancer cells. Mechanistically, two key activation elements, bound by transcription factor NF-Y and CREB1, respectively, were identified in the CDCA8 basic promoter by mutation analyses and electrophoretic motility shift assays. NF-Y binding is positively correlated with promoter activities in different cell types. Interestingly, the NF-YA subunit, binding to the promoter, is primarily a short isoform in hESCs and a long isoform in cancer cells, indicating a different activation mechanism of the CDCA8 transcription between hESCs and cancer cells. Finally, enhanced CDCA8 promoter activities by NF-Y overexpression and reduced CDCA8 transcription by NF-Y knockdown further verified that NF-Y is a positive regulator of CDCA8 transcription. Our study unearths the molecular mechanisms underlying the activation of CDCA8 expression in hESCs and cancer cells, which provides a better understanding of its biological functions.

  5. TAZ: a novel transcriptional co-activator regulated by interactions with 14-3-3 and PDZ domain proteins.

    Science.gov (United States)

    Kanai, F; Marignani, P A; Sarbassova, D; Yagi, R; Hall, R A; Donowitz, M; Hisaminato, A; Fujiwara, T; Ito, Y; Cantley, L C; Yaffe, M B

    2000-12-15

    The highly conserved and ubiquitously expressed 14-3-3 proteins regulate differentiation, cell cycle progression and apoptosis by binding intracellular phosphoproteins involved in signal transduction. By screening in vitro translated cDNA pools for the ability to bind 14-3-3, we identified a novel transcriptional co-activator, TAZ (transcriptional co-activator with PDZ-binding motif) as a 14-3-3-binding molecule. TAZ shares homology with Yes-associated protein (YAP), contains a WW domain and functions as a transcriptional co-activator by binding to the PPXY motif present on transcription factors. 14-3-3 binding requires TAZ phosphorylation on a single serine residue, resulting in the inhibition of TAZ transcriptional co-activation through 14-3-3-mediated nuclear export. The C-terminus of TAZ contains a highly conserved PDZ-binding motif that localizes TAZ into discrete nuclear foci and is essential for TAZ-stimulated gene transcription. TAZ uses this same motif to bind the PDZ domain-containing protein NHERF-2, a molecule that tethers plasma membrane ion channels and receptors to cytoskeletal actin. TAZ may link events at the plasma membrane and cytoskeleton to nuclear transcription in a manner that can be regulated by 14-3-3.

  6. Regulation of selected genome loci using de novo-engineered transcription activator-like effector (TALE)-type transcription factors.

    Science.gov (United States)

    Morbitzer, Robert; Römer, Patrick; Boch, Jens; Lahaye, Thomas

    2010-12-14

    Proteins that can be tailored to bind desired DNA sequences are key tools for molecular biology. Previous studies suggested that DNA-binding specificity of transcription activator-like effectors (TALEs) from the bacterial genus Xanthomonas is defined by repeat-variable diresidues (RVDs) of tandem-arranged 34/35-amino acid repeat units. We have studied chimeras of two TALEs differing in RVDs and non-RVDs and found that, in contrast to the critical contributions by RVDs, non-RVDs had no major effect on the DNA-binding specificity of the chimeras. This finding suggests that one needs only to modify the RVDs to generate designer TALEs (dTALEs) to activate transcription of user-defined target genes. We used the scaffold of the TALE AvrBs3 and changed its RVDs to match either the tomato Bs4, the Arabidopsis EGL3, or the Arabidopsis KNAT1 promoter. All three dTALEs transcriptionally activated the desired promoters in a sequence-specific manner as mutations within the targeted DNA sequences abolished promoter activation. This study is unique in showing that chromosomal loci can be targeted specifically by dTALEs. We also engineered two AvrBs3 derivatives with four additional repeat units activating specifically either the pepper Bs3 or UPA20 promoter. Because AvrBs3 activates both promoters, our data show that addition of repeat units facilitates TALE-specificity fine-tuning. Finally, we demonstrate that the RVD NK mediates specific interaction with G nucleotides that thus far could not be targeted specifically by any known RVD type. In summary, our data demonstrate that the TALE scaffold can be tailored to target user-defined DNA sequences in whole genomes.

  7. The ETS-5 transcription factor regulates activity states in Caenorhabditis elegans by controlling satiety

    Science.gov (United States)

    Juozaityte, Vaida; Pladevall-Morera, David; Podolska, Agnieszka; Nørgaard, Steffen; Pocock, Roger

    2017-01-01

    Animal behavior is shaped through interplay among genes, the environment, and previous experience. As in mammals, satiety signals induce quiescence in Caenorhabditis elegans. Here we report that the C. elegans transcription factor ETS-5, an ortholog of mammalian FEV/Pet1, controls satiety-induced quiescence. Nutritional status has a major influence on C. elegans behavior. When foraging, food availability controls behavioral state switching between active (roaming) and sedentary (dwelling) states; however, when provided with high-quality food, C. elegans become sated and enter quiescence. We show that ETS-5 acts to promote roaming and inhibit quiescence by setting the internal “satiety quotient” through fat regulation. Acting from the ASG and BAG sensory neurons, we show that ETS-5 functions in a complex network with serotonergic and neuropeptide signaling pathways to control food-regulated behavioral state switching. Taken together, our results identify a neuronal mechanism for controlling intestinal fat stores and organismal behavioral states in C. elegans, and establish a paradigm for the elucidation of obesity-relevant mechanisms. PMID:28193866

  8. Crystal structure of the caseinolytic protease gene regulator, a transcriptional activator in actinomycetes.

    Science.gov (United States)

    Russo, Santina; Schweitzer, Jens-Eric; Polen, Tino; Bott, Michael; Pohl, Ehmke

    2009-02-20

    Human pathogens of the genera Corynebacterium and Mycobacterium possess the transcriptional activator ClgR (clp gene regulator) which in Corynebacterium glutamicum has been shown to regulate the expression of the ClpCP protease genes. ClgR specifically binds to pseudo-palindromic operator regions upstream of clpC and clpP1P2. Here, we present the first crystal structure of a ClgR protein from C. glutamicum. The structure was determined from two different crystal forms to resolutions of 1.75 and 2.05 A, respectively. ClgR folds into a five-helix bundle with a helix-turn-helix motif typical for DNA-binding proteins. Upon dimerization the two DNA-recognition helices are arranged opposite to each other at the protein surface in a distance of approximately 30 A, which suggests that they bind into two adjacent major grooves of B-DNA in an anti-parallel manner. A binding pocket is situated at a strategic position in the dimer interface and could possess a regulatory role altering the positions of the DNA-binding helices.

  9. hSSB1 regulates both the stability and the transcriptional activity of p53

    Institute of Scientific and Technical Information of China (English)

    Shuangbing Xu; Yuanzhong Wu; Qiong Chen; Jingying Cao; Kaishun Hu; Jianjun Tang; Yi Sang

    2013-01-01

    The tumor suppressor p53 is essential for several cellular processes that are involved in the response to diverse genotoxic stress,including cell cycle arrest,DNA repair,apoptosis and senescence.Studies of the regulation of p53 have mostly focused on its stability and transactivation; however,new regulatory molecules for p53 have also been frequently identified.Here,we report that human ssDNA binding protein SSB1 (hSSB1),a novel DNA damageassociated protein,can interact with p53 and protect p53 from ubiquitin-mediated degradation.Furthermore,hSSB1 also associates with the acetyltransferase p300 and is required for efficient transcriptional activation of the p53 target gene p21 by affecting the acetylation of p53 at lysine382.Functionally,the hSSB1 knockdown-induced abrogation of the G2/M checkpoint is partially dependent on p53 or p300.Collectively,our results indicate that hSSB1 may regulate DNA damage checkpoints by positively modulating p53 and its downstream target p21.

  10. Ligand-regulated association of ErbB-4 to the transcriptional co-activator YAP65 controls transcription at the nuclear level.

    Science.gov (United States)

    Omerovic, Jasminka; Puggioni, Eleonora M R; Napoletano, Silvia; Visco, Vincenzo; Fraioli, Rocco; Frati, Luigi; Gulino, Alberto; Alimandi, Maurizio

    2004-04-01

    It has been proposed that ligand-dependent Regulated Intramembrane Proteolysis (RIP) of ErbB-4 receptors generates 80 kDa Intra-Cellular Domains (E4.ICDs) that relocate to the nuclear compartments where they implement the signaling abilities of the ErbB-4 receptors. The E4.ICD may directly regulate gene transcription or, in an alternative scenario, the tyrosine kinase activity of E4.ICDs may target proteins involved in transcriptional regulation upon its relocation into the nucleus. We have identified the transcriptional coactivator YAP65, here referred as YAP (Yes Associated Protein), as binding partner of ErbB-4 in a two hybrid screening in yeast. Interaction between YAP and ErbB-4 occurs via the WW domain of YAP and the PPPPY at positions 1297-1301 and the PPPAY at positions 1052-1056 of the amino acid sequence of the Cyt-1 isoform of ErbB-4. Stechiometry of binding is regulated by the ligand-dependent phosphorylation of Tyr 1056 in the PPPAYTPM module that function as "biochemical switch" to decrease the association of YAP to ErbB-4. In principle, this novel interaction highlights new mechanisms of signaling propagation from the ErbB-4 receptors, offering supporting evidences that the E4.ICDs forms released following ligand-receptor engagement may recruit YAP and relocate to the nucleus to implement or regulate transcription.

  11. The FERM-domain protein Expanded regulates Hippo pathway activity via direct interactions with the transcriptional activator Yorkie.

    Science.gov (United States)

    Badouel, Caroline; Gardano, Laura; Amin, Nancy; Garg, Ankush; Rosenfeld, Robyn; Le Bihan, Thierry; McNeill, Helen

    2009-03-01

    The Hippo kinase pathway plays a central role in growth regulation and tumor suppression from flies to man. The Hippo/Mst kinase phosphorylates and activates the NDR family kinase Warts/Lats, which phosphorylates and inhibits the transcriptional activator Yorkie/YAP. Current models place the FERM-domain protein Expanded upstream of Hippo kinase in growth control. To understand how Expanded regulates Hippo pathway activity, we used affinity chromatography and mass spectrometry to identify Expanded-binding proteins. Surprisingly we find that Yorkie is the major Expanded-binding protein in Drosophila S2 cells. Expanded binds Yorkie at endogenous levels via WW-domain-PPxY interactions, independently of Yorkie phosphorylation at S168, which is critical for 14-3-3 binding. Expanded relocalizes Yorkie from the nucleus, abrogating its nuclear activity, and it can regulate growth downstream of warts in vivo. These data lead to a new model whereby Expanded functions downstream of Warts, in concert with 14-3-3 proteins to sequester Yorkie in the cytoplasm, inhibiting growth activity of the Hippo pathway.

  12. Mechanisms of transcriptional regulation and prognostic significance of activated leukocyte cell adhesion molecule in cancer

    Directory of Open Access Journals (Sweden)

    Chen Hairu

    2010-10-01

    Full Text Available Abstract Background Activated leukocyte cell adhesion molecule (ALCAM is implicated in the prognosis of multiple cancers with low level expression associated with metastasis and early death in breast cancer. Despite this significance, mechanisms that regulate ALCAM gene expression and ALCAM's role in adhesion of pre-metastatic circulating tumor cells have not been defined. We studied ALCAM expression in 20 tumor cell lines by real-time PCR, western blot and immunochemistry. Epigenetic alterations of the ALCAM promoter were assessed using methylation-specific PCR and bisulfite sequencing. ALCAM's role in adhesion of tumor cells to the vascular wall was studied in isolated perfused lungs. Results A common site for transcription initiation of the ALCAM gene was identified and the ALCAM promoter sequenced. The promoter contains multiple cis-active elements including a functional p65 NF-κB motif, and it harbors an extensive array of CpG residues highly methylated exclusively in ALCAM-negative tumor cells. These CpG residues were modestly demethylated after 5-aza-2-deoxycytidine treatment. Restoration of high-level ALCAM expression using an ALCAM cDNA increased clustering of MDA-MB-435 tumor cells perfused through the pulmonary vasculature of ventilated rat lungs. Anti-ALCAM antibodies reduced the number of intravascular tumor cell clusters. Conclusion Our data suggests that loss of ALCAM expression, due in part to DNA methylation of extensive segments of the promoter, significantly impairs the ability of circulating tumor cells to adhere to each other, and may therefore promote metastasis. These findings offer insight into the mechanisms for down-regulation of ALCAM gene expression in tumor cells, and for the positive prognostic value of high-level ALCAM in breast cancer.

  13. Potential of Natural Products in the Inhibition of Adipogenesis through Regulation of PPARγ Expression and/or Its Transcriptional Activity.

    Science.gov (United States)

    Feng, Shi; Reuss, Laura; Wang, Yu

    2016-09-23

    Obesity is a global health problem characterized as an increase in the mass of adipose tissue. Adipogenesis is one of the key pathways that increases the mass of adipose tissue, by which preadipocytes mature into adipocytes through cell differentiation. Peroxisome proliferator-activated receptor γ (PPARγ), the chief regulator of adipogenesis, has been acutely investigated as a molecular target for natural products in the development of anti-obesity treatments. In this review, the regulation of PPARγ expression by natural products through inhibition of CCAAT/enhancer-binding protein β (C/EBPβ) and the farnesoid X receptor (FXR), increased expression of GATA-2 and GATA-3 and activation of the Wnt/β-catenin pathway were analyzed. Furthermore, the regulation of PPARγ transcriptional activity associated with natural products through the antagonism of PPARγ and activation of Sirtuin 1 (Sirt1) and AMP-activated protein kinase (AMPK) were discussed. Lastly, regulation of mitogen-activated protein kinase (MAPK) by natural products, which might regulate both PPARγ expression and PPARγ transcriptional activity, was summarized. Understanding the role natural products play, as well as the mechanisms behind their regulation of PPARγ activity is critical for future research into their therapeutic potential for fighting obesity.

  14. Potential of Natural Products in the Inhibition of Adipogenesis through Regulation of PPARγ Expression and/or Its Transcriptional Activity

    Directory of Open Access Journals (Sweden)

    Shi Feng

    2016-09-01

    Full Text Available Obesity is a global health problem characterized as an increase in the mass of adipose tissue. Adipogenesis is one of the key pathways that increases the mass of adipose tissue, by which preadipocytes mature into adipocytes through cell differentiation. Peroxisome proliferator-activated receptor γ (PPARγ, the chief regulator of adipogenesis, has been acutely investigated as a molecular target for natural products in the development of anti-obesity treatments. In this review, the regulation of PPARγ expression by natural products through inhibition of CCAAT/enhancer-binding protein β (C/EBPβ and the farnesoid X receptor (FXR, increased expression of GATA-2 and GATA-3 and activation of the Wnt/β-catenin pathway were analyzed. Furthermore, the regulation of PPARγ transcriptional activity associated with natural products through the antagonism of PPARγ and activation of Sirtuin 1 (Sirt1 and AMP-activated protein kinase (AMPK were discussed. Lastly, regulation of mitogen-activated protein kinase (MAPK by natural products, which might regulate both PPARγ expression and PPARγ transcriptional activity, was summarized. Understanding the role natural products play, as well as the mechanisms behind their regulation of PPARγ activity is critical for future research into their therapeutic potential for fighting obesity.

  15. Sumoylation of transcription factor Tec1 regulates signaling of mitogen-activated protein kinase pathways in yeast.

    Science.gov (United States)

    Wang, Yuqi; Abu Irqeba, Ameair; Ayalew, Mihretu; Suntay, Kristina

    2009-10-14

    Tec1 is a transcription factor in the yeast mitogen-activated protein kinase (MAPK) pathway that controls invasive growth. Previously we reported that a fraction of Tec1 protein is sumoylated on residue lysine 54 in normally growing cells. Here we describe regulation and functional consequences of Tec1 sumoylation. We found that activation of Kss1, the MAPK that directly activates Tec1, results in a decrease in Tec1 sumoylation and a concurrent increase of Tec1 transcriptional activity. Consistent with a role of sumoylation in inhibiting Tec1 activity, specifically increasing sumoylation of Tec1 by fusing it to the sumoylating enzyme Ubc9 leads to a dramatic decrease of Tec1 transcriptional activity. Invasive growth is also compromised in Tec1-Ubc9. In contrast, fusing sumoylation-site mutant Tec1, i.e., Tec1(K54R), to Ubc9 did not significantly alter transcriptional activation and had a less effect on invasive growth. Taken together, these findings provide evidence for regulated sumoylation as a mechanism to modulate the activity of Tec1 and validate Ubc9 fusion-directed sumoylation as a useful approach for studying protein sumoylation.

  16. Sumoylation of transcription factor Tec1 regulates signaling of mitogen-activated protein kinase pathways in yeast.

    Directory of Open Access Journals (Sweden)

    Yuqi Wang

    Full Text Available Tec1 is a transcription factor in the yeast mitogen-activated protein kinase (MAPK pathway that controls invasive growth. Previously we reported that a fraction of Tec1 protein is sumoylated on residue lysine 54 in normally growing cells. Here we describe regulation and functional consequences of Tec1 sumoylation. We found that activation of Kss1, the MAPK that directly activates Tec1, results in a decrease in Tec1 sumoylation and a concurrent increase of Tec1 transcriptional activity. Consistent with a role of sumoylation in inhibiting Tec1 activity, specifically increasing sumoylation of Tec1 by fusing it to the sumoylating enzyme Ubc9 leads to a dramatic decrease of Tec1 transcriptional activity. Invasive growth is also compromised in Tec1-Ubc9. In contrast, fusing sumoylation-site mutant Tec1, i.e., Tec1(K54R, to Ubc9 did not significantly alter transcriptional activation and had a less effect on invasive growth. Taken together, these findings provide evidence for regulated sumoylation as a mechanism to modulate the activity of Tec1 and validate Ubc9 fusion-directed sumoylation as a useful approach for studying protein sumoylation.

  17. Regulation of U6 Promoter Activity by Transcriptional Interference in Viral Vector-Based RNAi

    Institute of Scientific and Technical Information of China (English)

    Linghu Nie; Meghna Das Thakur; Yumei Wang; Qin Su; Yongliang Zhao; Yunfeng Feng

    2010-01-01

    The direct negative impact of the transcriptional activity of one component on the second one in c/s is referred to as transcriptional interference (TI).U6 is a type Ⅲ RNA polymerase Ⅲ promoter commonly used for driving small hairpin RNA (shRNA) expression in vector-based RNAi.In the design and construction of viral vectors,multiple transcription units may be arranged in close proximity in a space-limited vector.Determining if U6 promoter activity can be affected by TI is critical for the expression of target shRNA in gene therapy or loss-of-function studies.In this research,we designed and implemented a modified retroviral system where shRNA and exogenous gene expressions were driven by two independent transcriptional units.We arranged U6 promoter driving.shRNA expression and UbiC promoter in two promoter arrangements.In primary macrophages,we found U6 promoter activity was inhibited by UbiC promoter when in the divergent arrangement but not in tandem.In contrast,PKG promoter had no such negative impact.Instead of enhancing U6 promoter activity,CMV enhancer had significant negative impact on U6 promoter activity in the presence of UbiC promoter.Our results indicate that U6 promoter activity can be affected by TI in a proximal promoter-specific and arrangement-dependent manner.

  18. Resveratrol post-transcriptionally regulates pro-inflammatory gene expression via regulation of KSRP RNA binding activity

    Science.gov (United States)

    Bollmann, Franziska; Art, Julia; Henke, Jenny; Schrick, Katharina; Besche, Verena; Bros, Matthias; Li, Huige; Siuda, Daniel; Handler, Norbert; Bauer, Florian; Erker, Thomas; Behnke, Felix; Mönch, Bettina; Härdle, Lorena; Hoffmann, Markus; Chen, Ching-Yi; Förstermann, Ulrich; Dirsch, Verena M.; Werz, Oliver; Kleinert, Hartmut; Pautz, Andrea

    2014-01-01

    Resveratrol shows beneficial effects in inflammation-based diseases like cancer, cardiovascular and chronic inflammatory diseases. Therefore, the molecular mechanisms of the anti-inflammatory resveratrol effects deserve more attention. In human epithelial DLD-1 and monocytic Mono Mac 6 cells resveratrol decreased the expression of iNOS, IL-8 and TNF-α by reducing mRNA stability without inhibition of the promoter activity. Shown by pharmacological and siRNA-mediated inhibition, the observed effects are SIRT1-independent. Target-fishing and drug responsive target stability experiments showed selective binding of resveratrol to the RNA-binding protein KSRP, a central post-transcriptional regulator of pro-inflammatory gene expression. Knockdown of KSRP expression prevented resveratrol-induced mRNA destabilization in human and murine cells. Resveratrol did not change KSRP expression, but immunoprecipitation experiments indicated that resveratrol reduces the p38 MAPK-related inhibitory KSRP threonine phosphorylation, without blocking p38 MAPK activation or activity. Mutation of the p38 MAPK target site in KSRP blocked the resveratrol effect on pro-inflammatory gene expression. In addition, resveratrol incubation enhanced KSRP-exosome interaction, which is important for mRNA degradation. Finally, resveratrol incubation enhanced its intra-cellular binding to the IL-8, iNOS and TNF-α mRNA. Therefore, modulation of KSRP mRNA binding activity and, thereby, enhancement of mRNA degradation seems to be the common denominator of many anti-inflammatory effects of resveratrol. PMID:25352548

  19. A cytoplasmic negative regulator isoform of ATF7 impairs ATF7 and ATF2 phosphorylation and transcriptional activity.

    Science.gov (United States)

    Diring, Jessica; Camuzeaux, Barbara; Donzeau, Mariel; Vigneron, Marc; Rosa-Calatrava, Manuel; Kedinger, Claude; Chatton, Bruno

    2011-01-01

    Alternative splicing and post-translational modifications are processes that give rise to the complexity of the proteome. The nuclear ATF7 and ATF2 (activating transcription factor) are structurally homologous leucine zipper transcription factors encoded by distinct genes. Stress and growth factors activate ATF2 and ATF7 mainly via sequential phosphorylation of two conserved threonine residues in their activation domain. Distinct protein kinases, among which mitogen-activated protein kinases (MAPK), phosphorylate ATF2 and ATF7 first on Thr71/Thr53 and next on Thr69/Thr51 residues respectively, resulting in transcriptional activation. Here, we identify and characterize a cytoplasmic alternatively spliced isoform of ATF7. This variant, named ATF7-4, inhibits both ATF2 and ATF7 transcriptional activities by impairing the first phosphorylation event on Thr71/Thr53 residues. ATF7-4 indeed sequesters the Thr53-phosphorylating kinase in the cytoplasm. Upon stimulus-induced phosphorylation, ATF7-4 is poly-ubiquitinated and degraded, enabling the release of the kinase and ATF7/ATF2 activation. Our data therefore conclusively establish that ATF7-4 is an important cytoplasmic negative regulator of ATF7 and ATF2 transcription factors.

  20. A cytoplasmic negative regulator isoform of ATF7 impairs ATF7 and ATF2 phosphorylation and transcriptional activity.

    Directory of Open Access Journals (Sweden)

    Jessica Diring

    Full Text Available Alternative splicing and post-translational modifications are processes that give rise to the complexity of the proteome. The nuclear ATF7 and ATF2 (activating transcription factor are structurally homologous leucine zipper transcription factors encoded by distinct genes. Stress and growth factors activate ATF2 and ATF7 mainly via sequential phosphorylation of two conserved threonine residues in their activation domain. Distinct protein kinases, among which mitogen-activated protein kinases (MAPK, phosphorylate ATF2 and ATF7 first on Thr71/Thr53 and next on Thr69/Thr51 residues respectively, resulting in transcriptional activation. Here, we identify and characterize a cytoplasmic alternatively spliced isoform of ATF7. This variant, named ATF7-4, inhibits both ATF2 and ATF7 transcriptional activities by impairing the first phosphorylation event on Thr71/Thr53 residues. ATF7-4 indeed sequesters the Thr53-phosphorylating kinase in the cytoplasm. Upon stimulus-induced phosphorylation, ATF7-4 is poly-ubiquitinated and degraded, enabling the release of the kinase and ATF7/ATF2 activation. Our data therefore conclusively establish that ATF7-4 is an important cytoplasmic negative regulator of ATF7 and ATF2 transcription factors.

  1. Control of gene expression during T cell activation: alternate regulation of mRNA transcription and mRNA stability

    Directory of Open Access Journals (Sweden)

    Gorospe Myriam

    2005-05-01

    Full Text Available Abstract Background Microarray technology has become highly valuable for identifying complex global changes in gene expression patterns. The effective correlation of observed changes in gene expression with shared transcription regulatory elements remains difficult to demonstrate convincingly. One reason for this difficulty may result from the intricate convergence of both transcriptional and mRNA turnover events which, together, directly influence steady-state mRNA levels. Results In order to investigate the relative contribution of gene transcription and changes in mRNA stability regulation to standard analyses of gene expression, we used two distinct microarray methods which individually measure nuclear gene transcription and changes in polyA mRNA gene expression. Gene expression profiles were obtained from both polyA mRNA (whole-cell and nuclear run-on (newly transcribed RNA across a time course of one hour following the activation of human Jurkat T cells with PMA plus ionomycin. Comparative analysis revealed that regulation of mRNA stability may account for as much as 50% of all measurements of changes in polyA mRNA in this system, as inferred by the absence of any corresponding regulation of nuclear gene transcription activity for these groups of genes. Genes which displayed dramatic elevations in both mRNA and nuclear run-on RNA were shown to be inhibited by Actinomycin D (ActD pre-treatment of cells while large numbers of genes regulated only through altered mRNA turnover (both up and down were ActD-resistant. Consistent patterns across the time course were observed for both transcribed and stability-regulated genes. Conclusion We propose that regulation of mRNA stability contributes significantly to the observed changes in gene expression in response to external stimuli, as measured by high throughput systems.

  2. A central region of Gli2 regulates its localization to the primary cilium and transcriptional activity

    OpenAIRE

    Santos, Nicole; Reiter, Jeremy F.

    2014-01-01

    Signaling through vertebrate Hedgehog (Hh) proteins depends on the primary cilium. In response to Hh signals, the transcriptional activator of the pathway, Gli2, accumulates at the ciliary tip, raising the possibility that ciliary localization is important for Gli2 activation. To test this hypothesis, we used the Floxin system to create knock-in Gli2 alleles in embryonic stem cells (ESCs) to allow methodical testing of which domains and residues are essential for the ciliary localization of G...

  3. Peripheral CLOCK Regulates Target-Tissue Glucocorticoid Receptor Transcriptional Activity in a Circadian Fashion in Man

    Science.gov (United States)

    Charmandari, Evangelia; Chrousos, George P.; Lambrou, George I.; Pavlaki, Aikaterini; Koide, Hisashi; Ng, Sinnie Sin Man; Kino, Tomoshige

    2011-01-01

    Context and Objective Circulating cortisol fluctuates diurnally under the control of the “master” circadian CLOCK, while the peripheral “slave” counterpart of the latter regulates the transcriptional activity of the glucocorticoid receptor (GR) at local glucocorticoid target tissues through acetylation. In this manuscript, we studied the effect of CLOCK-mediated GR acetylation on the sensitivity of peripheral tissues to glucocorticoids in humans. Design and Participants We examined GR acetylation and mRNA expression of GR, CLOCK-related and glucocorticoid-responsive genes in peripheral blood mononuclear cells (PBMCs) obtained at 8 am and 8 pm from 10 healthy subjects, as well as in PBMCs obtained in the morning and cultured for 24 hours with exposure to 3-hour hydrocortisone pulses every 6 hours. We used EBV-transformed lymphocytes (EBVLs) as non-synchronized controls. Results GR acetylation was higher in the morning than in the evening in PBMCs, mirroring the fluctuations of circulating cortisol in reverse phase. All known glucocorticoid-responsive genes tested responded as expected to hydrocortisone in non-synchronized EBVLs, however, some of these genes did not show the expected diurnal mRNA fluctuations in PBMCs in vivo. Instead, their mRNA oscillated in a Clock- and a GR acetylation-dependent fashion in naturally synchronized PBMCs cultured ex vivo in the absence of the endogenous glucocorticoid, suggesting that circulating cortisol might prevent circadian GR acetylation-dependent effects in some glucocorticoid-responsive genes in vivo. Conclusions Peripheral CLOCK-mediated circadian acetylation of the human GR may function as a target-tissue, gene-specific counter regulatory mechanism to the actions of diurnally fluctuating cortisol, effectively decreasing tissue sensitivity to glucocorticoids in the morning and increasing it at night. PMID:21980503

  4. Peripheral CLOCK regulates target-tissue glucocorticoid receptor transcriptional activity in a circadian fashion in man.

    Directory of Open Access Journals (Sweden)

    Evangelia Charmandari

    Full Text Available CONTEXT AND OBJECTIVE: Circulating cortisol fluctuates diurnally under the control of the "master" circadian CLOCK, while the peripheral "slave" counterpart of the latter regulates the transcriptional activity of the glucocorticoid receptor (GR at local glucocorticoid target tissues through acetylation. In this manuscript, we studied the effect of CLOCK-mediated GR acetylation on the sensitivity of peripheral tissues to glucocorticoids in humans. DESIGN AND PARTICIPANTS: We examined GR acetylation and mRNA expression of GR, CLOCK-related and glucocorticoid-responsive genes in peripheral blood mononuclear cells (PBMCs obtained at 8 am and 8 pm from 10 healthy subjects, as well as in PBMCs obtained in the morning and cultured for 24 hours with exposure to 3-hour hydrocortisone pulses every 6 hours. We used EBV-transformed lymphocytes (EBVLs as non-synchronized controls. RESULTS: GR acetylation was higher in the morning than in the evening in PBMCs, mirroring the fluctuations of circulating cortisol in reverse phase. All known glucocorticoid-responsive genes tested responded as expected to hydrocortisone in non-synchronized EBVLs, however, some of these genes did not show the expected diurnal mRNA fluctuations in PBMCs in vivo. Instead, their mRNA oscillated in a Clock- and a GR acetylation-dependent fashion in naturally synchronized PBMCs cultured ex vivo in the absence of the endogenous glucocorticoid, suggesting that circulating cortisol might prevent circadian GR acetylation-dependent effects in some glucocorticoid-responsive genes in vivo. CONCLUSIONS: Peripheral CLOCK-mediated circadian acetylation of the human GR may function as a target-tissue, gene-specific counter regulatory mechanism to the actions of diurnally fluctuating cortisol, effectively decreasing tissue sensitivity to glucocorticoids in the morning and increasing it at night.

  5. Nuclear cereblon modulates transcriptional activity of Ikaros and regulates its downstream target, enkephalin, in human neuroblastoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Wada, Takeyoshi [Faculty of Science and Engineering, Waseda University, TWIns, 2-2 Wakamatsu, Shinjuku, Tokyo, 162-8480 (Japan); Asahi, Toru [Faculty of Science and Engineering, Waseda University, TWIns, 2-2 Wakamatsu, Shinjuku, Tokyo, 162-8480 (Japan); Research Organization for Nano & Life Innovation, Waseda University #03C309, TWIns, 2-2 Wakamatsu, Shinjuku, Tokyo, 162-8480 (Japan); Sawamura, Naoya, E-mail: naoya.sawamura@gmail.com [Faculty of Science and Engineering, Waseda University, TWIns, 2-2 Wakamatsu, Shinjuku, Tokyo, 162-8480 (Japan); Research Organization for Nano & Life Innovation, Waseda University #03C309, TWIns, 2-2 Wakamatsu, Shinjuku, Tokyo, 162-8480 (Japan)

    2016-08-26

    The gene coding cereblon (CRBN) was originally identified in genetic linkage analysis of mild autosomal recessive nonsyndromic intellectual disability. CRBN has broad localization in both the cytoplasm and nucleus. However, the significance of nuclear CRBN remains unknown. In the present study, we aimed to elucidate the role of CRBN in the nucleus. First, we generated a series of CRBN deletion mutants and determined the regions responsible for the nuclear localization. Only CRBN protein lacking the N-terminal region was localized outside of the nucleus, suggesting that the N-terminal region is important for its nuclear localization. CRBN was also identified as a thalidomide-binding protein and component of the cullin-4-containing E3 ubiquitin ligase complex. Thalidomide has been reported to be involved in the regulation of the transcription factor Ikaros by CRBN-mediated degradation. To investigate the nuclear functions of CRBN, we performed co-immunoprecipitation experiments and evaluated the binding of CRBN to Ikaros. As a result, we found that CRBN was associated with Ikaros protein, and the N-terminal region of CRBN was required for Ikaros binding. In luciferase reporter gene experiments, CRBN modulated transcriptional activity of Ikaros. Furthermore, we found that CRBN modulated Ikaros-mediated transcriptional repression of the proenkephalin gene by binding to its promoter region. These results suggest that CRBN binds to Ikaros via its N-terminal region and regulates transcriptional activities of Ikaros and its downstream target, enkephalin. - Highlights: • We found that CRBN is a nucleocytoplasmic shutting protein and identified the key domain for nucleocytoplasmic shuttling. • CRBN associates with the transcription factor Ikaros via the N-terminal domain. • CRBN modulates Ikaros-mediated transcriptional regulation and its downstream target, enkephalin.

  6. RNA-guided transcriptional regulation

    Energy Technology Data Exchange (ETDEWEB)

    Church, George M.; Mali, Prashant G.; Esvelt, Kevin M.

    2016-02-23

    Methods of modulating expression of a target nucleic acid in a cell are provided including introducing into the cell a first foreign nucleic acid encoding one or more RNAs complementary to DNA, wherein the DNA includes the target nucleic acid, introducing into the cell a second foreign nucleic acid encoding a nuclease-null Cas9 protein that binds to the DNA and is guided by the one or more RNAs, introducing into the cell a third foreign nucleic acid encoding a transcriptional regulator protein or domain, wherein the one or more RNAs, the nuclease-null Cas9 protein, and the transcriptional regulator protein or domain are expressed, wherein the one or more RNAs, the nuclease-null Cas9 protein and the transcriptional regulator protein or domain co-localize to the DNA and wherein the transcriptional regulator protein or domain regulates expression of the target nucleic acid.

  7. Genomewide Analyses Define Different Modes of Transcriptional Regulation by Peroxisome Proliferator-Activated Receptor-β/δ (PPARβ/δ)

    Science.gov (United States)

    Schönbauer, Anne; Meissner, Wolfgang; Scharfe, Maren; Jarek, Michael; Blöcker, Helmut; Müller-Brüsselbach, Sabine; Müller, Rolf

    2011-01-01

    Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors with essential functions in lipid, glucose and energy homeostasis, cell differentiation, inflammation and metabolic disorders, and represent important drug targets. PPARs heterodimerize with retinoid X receptors (RXRs) and can form transcriptional activator or repressor complexes at specific DNA elements (PPREs). It is believed that the decision between repression and activation is generally governed by a ligand-mediated switch. We have performed genomewide analyses of agonist-treated and PPARβ/δ-depleted human myofibroblasts to test this hypothesis and to identify global principles of PPARβ/δ-mediated gene regulation. Chromatin immunoprecipitation sequencing (ChIP-Seq) of PPARβ/δ, H3K4me3 and RNA polymerase II enrichment sites combined with transcriptional profiling enabled the definition of 112 bona fide PPARβ/δ target genes showing either of three distinct types of transcriptional response: (I) ligand-independent repression by PPARβ/δ; (II) ligand-induced activation and/or derepression by PPARβ/δ; and (III) ligand-independent activation by PPARβ/δ. These data identify PPRE-mediated repression as a major mechanism of transcriptional regulation by PPARβ/δ, but, unexpectedly, also show that only a subset of repressed genes are activated by a ligand-mediated switch. Our results also suggest that the type of transcriptional response by a given target gene is connected to the structure of its associated PPRE(s) and the biological function of its encoded protein. These observations have important implications for understanding the regulatory PPAR network and PPARβ/δ ligand-based drugs. PMID:21283829

  8. Tet1 oxidase regulates neuronal gene transcription, active DNA hydroxymethylation, object location memory, and threat recognition memory

    Directory of Open Access Journals (Sweden)

    Dinesh Kumar

    2015-10-01

    Full Text Available A dynamic equilibrium between DNA methylation and demethylation of neuronal activity-regulated genes is crucial for memory processes. However, the mechanisms underlying this equilibrium remain elusive. Tet1 oxidase has been shown to play a key role in the active DNA demethylation in the central nervous system. In this study, we used Tet1 gene knockout (Tet1KO mice to examine the involvement of Tet1 in memory consolidation and storage in the adult brain. We found that Tet1 ablation leads to altered expression of numerous neuronal activity-regulated genes, compensatory upregulation of active demethylation pathway genes, and upregulation of various epigenetic modifiers. Moreover, Tet1KO mice showed an enhancement in the consolidation and storage of threat recognition (cued and contextual fear conditioning and object location memories. We conclude that Tet1 plays a critical role in regulating neuronal transcription and in maintaining the epigenetic state of the brain associated with memory consolidation and storage.

  9. Signal transducer and activator of transcription 3 and 5 regulate system Xc- and redox balance in human breast cancer cells.

    Science.gov (United States)

    Linher-Melville, Katja; Haftchenary, Sina; Gunning, Patrick; Singh, Gurmit

    2015-07-01

    System Xc- is a cystine/glutamate antiporter that contributes to the maintenance of cellular redox balance. The human xCT (SLC7A11) gene encodes the functional subunit of system Xc-. Transcription factors regulating antioxidant defense mechanisms including system Xc- are of therapeutic interest, especially given that aggressive breast cancer cells exhibit increased system Xc- function. This investigation provides evidence that xCT expression is regulated by STAT3 and/or STAT5A, functionally affecting the antiporter in human breast cancer cells. Computationally analyzing two kilobase pairs of the xCT promoter/5' flanking region identified a distal gamma-activated site (GAS) motif, with truncations significantly increasing luciferase reporter activity. Similar transcriptional increases were obtained after treating cells transiently transfected with the full-length xCT promoter construct with STAT3/5 pharmacological inhibitors. Knock-down of STAT3 or STAT5A with siRNAs produced similar results. However, GAS site mutation significantly reduced xCT transcriptional activity, suggesting that STATs may interact with other transcription factors at more proximal promoter sites. STAT3 and STAT5A were bound to the xCT promoter in MDA-MB-231 cells, and binding was disrupted by pre-treatment with STAT inhibitors. Pharmacologically suppressing STAT3/5 activation significantly increased xCT mRNA and protein levels, as well as cystine uptake, glutamate release, and total levels of intracellular glutathione. Our data suggest that STAT proteins negatively regulate basal xCT expression. Blocking STAT3/5-mediated signaling induces an adaptive, compensatory mechanism to protect breast cancer cells from stress, including reactive oxygen species, by up-regulating xCT expression and the function of system Xc-. We propose that targeting system Xc- together with STAT3/5 inhibitors may heighten therapeutic anti-cancer effects.

  10. Master regulator for chondrogenesis, Sox9, regulates transcriptional activation of the endoplasmic reticulum stress transducer BBF2H7/CREB3L2 in chondrocytes.

    Science.gov (United States)

    Hino, Kenta; Saito, Atsushi; Kido, Miori; Kanemoto, Soshi; Asada, Rie; Takai, Tomoko; Cui, Min; Cui, Xiang; Imaizumi, Kazunori

    2014-05-16

    The endoplasmic reticulum (ER) stress transducer, box B-binding factor 2 human homolog on chromosome 7 (BBF2H7), is a basic leucine zipper (bZIP) transmembrane transcription factor. This molecule is activated in response to ER stress during chondrogenesis. The activated BBF2H7 accelerates cartilage matrix protein secretion through the up-regulation of Sec23a, which is responsible for protein transport from the ER to the Golgi apparatus and is a target of BBF2H7. In the present study, we elucidated the mechanisms of the transcriptional activation of Bbf2h7 in chondrocytes. The transcription of Bbf2h7 is regulated by Sex determining region Y-related high-mobility group box 9 (Sox9), a critical factor for chondrocyte differentiation that facilitates the expression of one of the major cartilage matrix proteins Type II collagen (Col2), through binding to the Sox DNA-binding motif in the Bbf2h7 promoter. BBF2H7 is activated as a transcription factor in response to physiological ER stress caused by abundant synthesis of cartilage matrix proteins, and consequently regulates the secretion of cartilage matrix proteins. Taken together, our findings demonstrate novel regulatory mechanisms of Sox9 for controlling the secretion of cartilage matrix proteins through the activation of BBF2H7-Sec23a signaling during chondrogenesis.

  11. Altered LKB1/CREB-regulated transcription co-activator (CRTC) signaling axis promotes esophageal cancer cell migration and invasion.

    Science.gov (United States)

    Gu, Y; Lin, S; Li, J-L; Nakagawa, H; Chen, Z; Jin, B; Tian, L; Ucar, D A; Shen, H; Lu, J; Hochwald, S N; Kaye, F J; Wu, L

    2012-01-26

    LKB1 is a tumor susceptibility gene for the Peutz-Jeghers cancer syndrome and is a target for mutational inactivation in sporadic human malignancies. LKB1 encodes a serine/threonine kinase that has critical roles in cell growth, polarity and metabolism. A novel and important function of LKB1 is its ability to regulate the phosphorylation of CREB-regulated transcription co-activators (CRTCs) whose aberrant activation is linked with oncogenic activities. However, the roles and mechanisms of LKB1 and CRTC in the pathogenesis of esophageal cancer have not been previously investigated. In this study, we observed altered LKB1-CRTC signaling in a subset of human esophageal cancer cell lines and patient samples. LKB1 negatively regulates esophageal cancer cell migration and invasion in vitro. Mechanistically, we determined that CRTC signaling becomes activated because of LKB1 loss, which results in the transcriptional activation of specific downstream targets including LYPD3, a critical mediator for LKB1 loss-of-function. Our data indicate that de-regulated LKB1-CRTC signaling might represent a crucial mechanism for esophageal cancer progression.

  12. Nuclear cereblon modulates transcriptional activity of Ikaros and regulates its downstream target, enkephalin, in human neuroblastoma cells.

    Science.gov (United States)

    Wada, Takeyoshi; Asahi, Toru; Sawamura, Naoya

    2016-08-26

    The gene coding cereblon (CRBN) was originally identified in genetic linkage analysis of mild autosomal recessive nonsyndromic intellectual disability. CRBN has broad localization in both the cytoplasm and nucleus. However, the significance of nuclear CRBN remains unknown. In the present study, we aimed to elucidate the role of CRBN in the nucleus. First, we generated a series of CRBN deletion mutants and determined the regions responsible for the nuclear localization. Only CRBN protein lacking the N-terminal region was localized outside of the nucleus, suggesting that the N-terminal region is important for its nuclear localization. CRBN was also identified as a thalidomide-binding protein and component of the cullin-4-containing E3 ubiquitin ligase complex. Thalidomide has been reported to be involved in the regulation of the transcription factor Ikaros by CRBN-mediated degradation. To investigate the nuclear functions of CRBN, we performed co-immunoprecipitation experiments and evaluated the binding of CRBN to Ikaros. As a result, we found that CRBN was associated with Ikaros protein, and the N-terminal region of CRBN was required for Ikaros binding. In luciferase reporter gene experiments, CRBN modulated transcriptional activity of Ikaros. Furthermore, we found that CRBN modulated Ikaros-mediated transcriptional repression of the proenkephalin gene by binding to its promoter region. These results suggest that CRBN binds to Ikaros via its N-terminal region and regulates transcriptional activities of Ikaros and its downstream target, enkephalin.

  13. Localizing potentially active post-transcriptional regulations in the Ewing's sarcoma gene regulatory network

    Directory of Open Access Journals (Sweden)

    Delyon Bernard

    2010-11-01

    Full Text Available Abstract Background A wide range of techniques is now available for analyzing regulatory networks. Nonetheless, most of these techniques fail to interpret large-scale transcriptional data at the post-translational level. Results We address the question of using large-scale transcriptomic observation of a system perturbation to analyze a regulatory network which contained several types of interactions - transcriptional and post-translational. Our method consisted of post-processing the outputs of an open-source tool named BioQuali - an automatic constraint-based analysis mimicking biologist's local reasoning on a large scale. The post-processing relied on differences in the behavior of the transcriptional and post-translational levels in the network. As a case study, we analyzed a network representation of the genes and proteins controlled by an oncogene in the context of Ewing's sarcoma. The analysis allowed us to pinpoint active interactions specific to this cancer. We also identified the parts of the network which were incomplete and should be submitted for further investigation. Conclusions The proposed approach is effective for the qualitative analysis of cancer networks. It allows the integrative use of experimental data of various types in order to identify the specific information that should be considered a priority in the initial - and possibly very large - experimental dataset. Iteratively, new dataset can be introduced into the analysis to improve the network representation and make it more specific.

  14. Transcription regulation mechanisms of bacteriophages

    Science.gov (United States)

    Yang, Haiquan; Ma, Yingfang; Wang, Yitian; Yang, Haixia; Shen, Wei; Chen, Xianzhong

    2014-01-01

    Phage diversity significantly contributes to ecology and evolution of new bacterial species through horizontal gene transfer. Therefore, it is essential to understand the mechanisms underlying phage-host interactions. After initial infection, the phage utilizes the transcriptional machinery of the host to direct the expression of its own genes. This review presents a view on the transcriptional regulation mechanisms of bacteriophages, and its contribution to phage diversity and classification. Through this review, we aim to broaden the understanding of phage-host interactions while providing a reference source for researchers studying the regulation of phage transcription. PMID:25482231

  15. Nuclear IL-33 is a transcriptional regulator of NF-{kappa}B p65 and induces endothelial cell activation

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Yeon-Sook; Park, Jeong Ae; Kim, Jihye; Rho, Seung-Sik; Park, Hyojin [Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of); Kim, Young-Myeong [Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon (Korea, Republic of); Kwon, Young-Guen, E-mail: ygkwon@yonsei.ac.kr [Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of)

    2012-05-04

    Highlights: Black-Right-Pointing-Pointer IL-33 as nuclear factor regulated expression of ICAM-1 and VCAM-1. Black-Right-Pointing-Pointer Nuclear IL-33 increased the transcription of NF-{kappa}B p65 by binding to the p65 promoter. Black-Right-Pointing-Pointer Nuclear IL-33 controls NF-{kappa}B-dependent inflammatory responses. -- Abstract: Interleukin (IL)-33, an IL-1 family member, acts as an extracellular cytokine by binding its cognate receptor, ST2. IL-33 is also a chromatin-binding transcriptional regulator highly expressed in the nuclei of endothelial cells. However, the function of IL-33 as a nuclear factor is poorly defined. Here, we show that IL-33 is a novel transcriptional regulator of the p65 subunit of the NF-{kappa}B complex and is involved in endothelial cell activation. Quantitative reverse transcriptase PCR and Western blot analyses indicated that IL-33 mediates the expression of intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 in endothelial cells basally and in response to tumor necrosis factor-{alpha}-treatment. IL-33-induced ICAM-1/VCAM-1 expression was dependent on the regulatory effect of IL-33 on the nuclear factor (NF)-{kappa}B pathway; NF-{kappa}B p65 expression was enhanced by IL-33 overexpression and, conversely, reduced by IL-33 knockdown. Moreover, NF-{kappa}B p65 promoter activity and chromatin immunoprecipitation analysis revealed that IL-33 binds to the p65 promoter region in the nucleus. Our data provide the first evidence that IL-33 in the nucleus of endothelial cells participates in inflammatory reactions as a transcriptional regulator of NF-{kappa}B p65.

  16. Catching transcriptional regulation by thermostatistical modeling

    Science.gov (United States)

    Frank, Till D.; Cheong, Alex; Okada-Hatakeyama, Mariko; Kholodenko, Boris N.

    2012-08-01

    Gene expression is frequently regulated by multiple transcription factors (TFs). Thermostatistical methods allow for a quantitative description of interactions between TFs, RNA polymerase and DNA, and their impact on the transcription rates. We illustrate three different scales of the thermostatistical approach: the microscale of TF molecules, the mesoscale of promoter energy levels and the macroscale of transcriptionally active and inactive cells in a cell population. We demonstrate versatility of combinatorial transcriptional activation by exemplifying logic functions, such as AND and OR gates. We discuss a metric for cell-to-cell transcriptional activation variability known as Fermi entropy. Suitability of thermostatistical modeling is illustrated by describing the experimental data on transcriptional induction of NFκB and the c-Fos protein.

  17. Transcriptional Regulation of Plant Secondary Metabolism

    Institute of Scientific and Technical Information of China (English)

    Chang-Qing Yang; Xin Fang; Xiu-Ming Wu; Ying-Bo Mao; Ling-Jian Wang; Xiao-Ya Chen

    2012-01-01

    Plant secondary metabolites play critical roles in plant-environment interactions.They are synthesized in different organs or tissues at particular developmental stages,and in response to various environmental stimuli,both biotic and abiotic.Accordingly,corresponding genes are regulated at the transcriptional level by multiple transcription factors.Several families of transcription factors have been identified to participate in controlling the biosynthesis and accumulation of secondary metabolites.These regulators integrate internal (often developmental) and external signals,bind to corresponding cis-elements — which are often in the promoter regions — to activate or repress the expression of enzyme-coding genes,and some of them interact with other transcription factors to form a complex.In this review,we summarize recent research in these areas,with an emphasis on newly-identified transcription factors and their functions in metabolism regulation.

  18. A super-family of transcriptional activators regulates bacteriophage packaging and lysis in Gram-positive bacteria.

    Science.gov (United States)

    Quiles-Puchalt, Nuria; Tormo-Más, María Ángeles; Campoy, Susana; Toledo-Arana, Alejandro; Monedero, Vicente; Lasa, Iñigo; Novick, Richard P; Christie, Gail E; Penadés, José R

    2013-08-01

    The propagation of bacteriophages and other mobile genetic elements requires exploitation of the phage mechanisms involved in virion assembly and DNA packaging. Here, we identified and characterized four different families of phage-encoded proteins that function as activators required for transcription of the late operons (morphogenetic and lysis genes) in a large group of phages infecting Gram-positive bacteria. These regulators constitute a super-family of proteins, here named late transcriptional regulators (Ltr), which share common structural, biochemical and functional characteristics and are unique to this group of phages. They are all small basic proteins, encoded by genes present at the end of the early gene cluster in their respective phage genomes and expressed under cI repressor control. To control expression of the late operon, the Ltr proteins bind to a DNA repeat region situated upstream of the terS gene, activating its transcription. This involves the C-terminal part of the Ltr proteins, which control specificity for the DNA repeat region. Finally, we show that the Ltr proteins are the only phage-encoded proteins required for the activation of the packaging and lysis modules. In summary, we provide evidence that phage packaging and lysis is a conserved mechanism in Siphoviridae infecting a wide variety of Gram-positive bacteria.

  19. A super-family of transcriptional activators regulates bacteriophage packaging and lysis in Gram-positive bacteria

    Science.gov (United States)

    Quiles-Puchalt, Nuria; Tormo-Más, María Ángeles; Campoy, Susana; Toledo-Arana, Alejandro; Monedero, Vicente; Lasa, Íñigo; Novick, Richard P.; Christie, Gail E.; Penadés, José R.

    2013-01-01

    The propagation of bacteriophages and other mobile genetic elements requires exploitation of the phage mechanisms involved in virion assembly and DNA packaging. Here, we identified and characterized four different families of phage-encoded proteins that function as activators required for transcription of the late operons (morphogenetic and lysis genes) in a large group of phages infecting Gram-positive bacteria. These regulators constitute a super-family of proteins, here named late transcriptional regulators (Ltr), which share common structural, biochemical and functional characteristics and are unique to this group of phages. They are all small basic proteins, encoded by genes present at the end of the early gene cluster in their respective phage genomes and expressed under cI repressor control. To control expression of the late operon, the Ltr proteins bind to a DNA repeat region situated upstream of the terS gene, activating its transcription. This involves the C-terminal part of the Ltr proteins, which control specificity for the DNA repeat region. Finally, we show that the Ltr proteins are the only phage-encoded proteins required for the activation of the packaging and lysis modules. In summary, we provide evidence that phage packaging and lysis is a conserved mechanism in Siphoviridae infecting a wide variety of Gram-positive bacteria. PMID:23771138

  20. Histone deacetylases regulate gonadotropin-releasing hormone I gene expression via modulating Otx2-driven transcriptional activity.

    Directory of Open Access Journals (Sweden)

    Lu Gan

    Full Text Available BACKGROUND: Precise coordination of the hypothalamic-pituitary-gonadal axis orchestrates the normal reproductive function. As a central regulator, the appropriate synthesis and secretion of gonadotropin-releasing hormone I (GnRH-I from the hypothalamus is essential for the coordination. Recently, emerging evidence indicates that histone deacetylases (HDACs play an important role in maintaining normal reproductive function. In this study, we identify the potential effects of HDACs on Gnrh1 gene transcription. METHODOLOGY/PRINCIPAL FINDINGS: Inhibition of HDACs activities by trichostatin A (TSA and valproic acid (VPA promptly and dramatically repressed transcription of Gnrh1 gene in the mouse immortalized mature GnRH neuronal cells GT1-7. The suppression was connected with a specific region of Gnrh1 gene promoter, which contains two consensus Otx2 binding sites. Otx2 has been known to activate the basal and also enhancer-driven transcription of Gnrh1 gene. The transcriptional activity of Otx2 is negatively modulated by Grg4, a member of the Groucho-related-gene (Grg family. In the present study, the expression of Otx2 was downregulated by TSA and VPA in GT1-7 cells, accompanied with the opposite changes of Grg4 expression. Chromatin immunoprecipitation and electrophoretic mobility shift assays demonstrated that the DNA-binding activity of Otx2 to Gnrh1 gene was suppressed by TSA and VPA. Overexpression of Otx2 partly abolished the TSA- and VPA-induced downregulation of Gnrh1 gene expression. CONCLUSIONS/SIGNIFICANCE: Our data indicate that HDAC inhibitors downregulate Gnrh1 gene expression via repressing Otx2-driven transcriptional activity. This study should provide an insight for our understanding on the effects of HDACs in the reproductive system and suggests that HDACs could be potential novel targets for the therapy of GnRH-related diseases.

  1. VEGF-A isoform-specific regulation of calcium ion flux, transcriptional activation and endothelial cell migration

    Directory of Open Access Journals (Sweden)

    Gareth W. Fearnley

    2015-07-01

    Full Text Available Vascular endothelial growth factor A (VEGF-A regulates many aspects of vascular physiology such as cell migration, proliferation, tubulogenesis and cell-cell interactions. Numerous isoforms of VEGF-A exist but their physiological significance is unclear. Here we evaluated two different VEGF-A isoforms and discovered differential regulation of cytosolic calcium ion flux, transcription factor localisation and endothelial cell response. Analysis of VEGF-A isoform-specific stimulation of VEGFR2-dependent signal transduction revealed differential capabilities for isoform activation of multiple signal transduction pathways. VEGF-A165 treatment promoted increased phospholipase Cγ1 phosphorylation, which was proportional to the subsequent rise in cytosolic calcium ions, in comparison to cells treated with VEGF-A121. A major consequence of this VEGF-A isoform-specific calcium ion flux in endothelial cells is differential dephosphorylation and subsequent nuclear translocation of the transcription factor NFATc2. Using reverse genetics, we discovered that NFATc2 is functionally required for VEGF-A-stimulated endothelial cell migration but not tubulogenesis. This work presents a new mechanism for understanding how VEGF-A isoforms program complex cellular outputs by converting signal transduction pathways into transcription factor redistribution to the nucleus, as well as defining a novel role for NFATc2 in regulating the endothelial cell response.

  2. Protein inhibitor of activated STAT 4 (PIAS4) regulates pro-inflammatory transcription in hepatocytes by repressing SIRT1.

    Science.gov (United States)

    Sun, Lina; Fan, Zhiwen; Chen, Junliang; Tian, Wenfang; Li, Min; Xu, Huihui; Wu, Xiaoyan; Fang, Mingming; Xia, Jun; Xu, Yong

    2016-07-12

    Excessive nutrition promotes the pathogenesis of non-alcoholic steatohepatitis (NASH), characterized by the accumulation of pro-inflammation mediators in the liver. In the present study we investigated the regulation of pro-inflammatory transcription in hepatocytes by protein inhibitor of activated STAT 4 (PIAS4) in this process and the underlying mechanisms. We report that expression of the class III deacetylase SIRT1 was down-regulated in the livers of NASH mice accompanied by a simultaneous increase in the expression and binding activity of PIAS4. Exposure to high glucose stimulated the expression PIAS4 in cultured hepatocytes paralleling SIRT1 repression. Estrogen, a known NASH-protective hormone, ameliorated SIRT1 trans-repression by targeting PIAS4. Over-expression of PIAS4 enhanced, while PIAS4 knockdown alleviated, repression of SIRT1 transcription by high glucose. Lentiviral delivery of short hairpin RNA (shRNA) targeting PIAS4 attenuated hepatic inflammation in NASH mice by restoring SIRT1 expression. Mechanistically, PIAS4 promoted NF-κB-mediated pro-inflammatory transcription in a SIRT1 dependent manner. In conclusion, our study indicates that PIAS4 mediated SIRT1 repression in response to nutrient surplus contributes to the pathogenesis of NASH. Therefore, targeting PIAS4 might provide novel therapeutic strategies in the intervention of NASH.

  3. Distinct regulation of activity-dependent transcription of immediate early genes in cultured rat cortical neurons.

    Science.gov (United States)

    Fukuchi, Mamoru; Sanabe, Tomofumi; Watanabe, Toshifumi; Kubota, Takane; Tabuchi, Akiko; Tsuda, Masaaki

    2017-08-26

    The activity-regulated expression of immediate early genes (IEGs) contributes to long-lasting neuronal functions underlying long-term memory. However, their response properties following neuronal activity are unique and remain poorly understood. To address this knowledge gap, here we further investigated the response properties of two representative IEGs, c-fos and brain-derived neurotrophic factor (Bdnf). Treatment of cultured cortical cells with KCl produces a depolarization process that results in the increase of intracellular calcium concentration in a KCl concentration-dependent manner. Consistent with this increase, c-fos expression was induced in a KCl concentration-dependent manner. In contrast, however, Bdnf expression was optimally activated by both 25 and 50 mM concentration of KCl. Similar results were observed when the cells were treated with okadaic acid, which inhibits protein phosphatases and elicits the hyper-phosphorylation of signaling molecules. Thus, Bdnf expression is strictly regulated by a neuronal activity threshold in an all or nothing manner, whereas c-fos expression is activated in a neuronal activity-dependent manner. Our findings also suggest that these differential responses might be due to the presence or absence of a TATA box. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Bordetella pertussis fim3 gene regulation by BvgA: phosphorylation controls the formation of inactive vs. active transcription complexes.

    Science.gov (United States)

    Boulanger, Alice; Moon, Kyung; Decker, Kimberly B; Chen, Qing; Knipling, Leslie; Stibitz, Scott; Hinton, Deborah M

    2015-02-10

    Two-component systems [sensor kinase/response regulator (RR)] are major tools used by microorganisms to adapt to environmental conditions. RR phosphorylation is typically required for gene activation, but few studies have addressed how and if phosphorylation affects specific steps during transcription initiation. We characterized transcription complexes made with RNA polymerase and the Bordetella pertussis RR, BvgA, in its nonphosphorylated or phosphorylated (BvgA∼P) state at P(fim3), the promoter for the virulence gene fim3 (fimbrial subunit), using gel retardation, potassium permanganate and DNase I footprinting, cleavage reactions with protein conjugated with iron bromoacetamidobenzyl-EDTA, and in vitro transcription. Previous work has shown that the level of nonphosphorylated BvgA remains high in vivo under conditions in which BvgA is phosphorylated. Our results here indicate that surprisingly both BvgA and BvgA∼P form open and initiating complexes with RNA polymerase at P(fim3). However, phosphorylation of BvgA is needed to generate the correct conformation that can transition to competent elongation. Footprints obtained with the complexes made with nonphosphorylated BvgA are atypical; while the initiating complex with BvgA synthesizes short RNA, it does not generate full-length transcripts. Extended incubation of the BvgA/RNA polymerase initiated complex in the presence of heparin generates a stable, but defective species that depends on the initial transcribed sequence of fim3. We suggest that the presence of nonphosphorylated BvgA down-regulates P(fim3) activity when phosphorylated BvgA is present and may allow the bacterium to quickly adapt to the loss of inducing conditions by rapidly eliminating P(fim3) activation once the signal for BvgA phosphorylation is removed.

  5. Combinatorial Regulation in Yeast Transcription Networks

    Science.gov (United States)

    Li, Hao

    2006-03-01

    Yeast has evolved a complex network to regulate its transcriptional program in response to changes in environment. It is quite common that in response to an external stimulus, several transcription factors will be activated and they work in combinations to control different subsets of genes in the genome. We are interested in how the promoters of genes are designed to integrate signals from multiple transcription factors and what are the functional and evolutionary constraints. To answer how, we have developed a number of computational algorithms to systematically map the binding sites and target genes of transcription factors using sequence and gene expression data. To analyze the functional constraints, we have employed mechanistic models to study the dynamic behavior of genes regulated by multiple factors. We have also developed methods to trace the evolution of transcriptional networks via comparative analysis of multiple species.

  6. FTH1P3, a Novel H-Ferritin Pseudogene Transcriptionally Active, Is Ubiquitously Expressed and Regulated during Cell Differentiation.

    Directory of Open Access Journals (Sweden)

    Maddalena Di Sanzo

    Full Text Available Ferritin, the major iron storage protein, performs its essential functions in the cytoplasm, nucleus and mitochondria. The variable assembly of 24 subunits of the Heavy (H and Light (L type composes the cytoplasmic molecule. In humans, two distinct genes code these subunits, both belonging to complex multigene families. Until now, one H gene has been identified with the coding sequence interrupted by three introns and more than 20 intronless copies widely dispersed on different chromosomes. Two of the intronless genes are actively transcribed in a tissue-specific manner. Herein, we report that FTH1P3, another intronless pseudogene, is transcribed. FTH1P3 transcript was detected in several cell lines and tissues, suggesting that its transcription is ubiquitary, as it happens for the parental ferritin H gene. Moreover, FTH1P3 expression is positively regulated during the cell differentiation process.

  7. FTH1P3, a Novel H-Ferritin Pseudogene Transcriptionally Active, Is Ubiquitously Expressed and Regulated during Cell Differentiation.

    Science.gov (United States)

    Di Sanzo, Maddalena; Aversa, Ilenia; Santamaria, Gianluca; Gagliardi, Monica; Panebianco, Mariafranca; Biamonte, Flavia; Zolea, Fabiana; Faniello, Maria Concetta; Cuda, Giovanni; Costanzo, Francesco

    2016-01-01

    Ferritin, the major iron storage protein, performs its essential functions in the cytoplasm, nucleus and mitochondria. The variable assembly of 24 subunits of the Heavy (H) and Light (L) type composes the cytoplasmic molecule. In humans, two distinct genes code these subunits, both belonging to complex multigene families. Until now, one H gene has been identified with the coding sequence interrupted by three introns and more than 20 intronless copies widely dispersed on different chromosomes. Two of the intronless genes are actively transcribed in a tissue-specific manner. Herein, we report that FTH1P3, another intronless pseudogene, is transcribed. FTH1P3 transcript was detected in several cell lines and tissues, suggesting that its transcription is ubiquitary, as it happens for the parental ferritin H gene. Moreover, FTH1P3 expression is positively regulated during the cell differentiation process.

  8. Transcriptional Activation of the Rhodobacter sphaeroides Cytochrome c2 Gene P2 Promoter by the Response Regulator PrrA

    OpenAIRE

    Comolli, James C; Carl, Audrey J.; Hall, Christine; Donohue, Timothy

    2002-01-01

    Anoxygenic photosynthetic growth of Rhodobacter sphaeroides, a member of the α subclass of the class Proteobacteria, requires the response regulator PrrA. PrrA and the sensor kinase PrrB are part of a two-component signaling pathway that influences a wide range of processes under oxygen-limited conditions. In this work we characterized the pathway of transcription activation by PrrB and PrrA by purifying these proteins, analyzing them in vitro, and characterizing a mutant PrrA protein in vivo...

  9. Regulation of RUNX2 transcription factor-DNA interactions and cell proliferation by vitamin D3 (cholecalciferol) prohormone activity.

    Science.gov (United States)

    Underwood, Karen F; D'Souza, David R; Mochin-Peters, Maria; Pierce, Adam D; Kommineni, Sravya; Choe, Moran; Bennett, Jessica; Gnatt, Averell; Habtemariam, Bahru; MacKerell, Alexander D; Passaniti, Antonino

    2012-04-01

    The fat-soluble prohormone cholecalciferol (Vitamin D3) is a precursor of the circulating 25-OH Vitamin D3, which is converted by 1α-hydroxylase to the biologically active 1,25-OH Vitamin D3. Active Vitamin D3 interacts with the Vitamin D receptor (VDR), a transcription factor that plays an important role in calcium mobilization and bone formation. RUNX2 is a DNA-binding transcription factor that regulates target genes important in bone formation, angiogenesis, and cancer metastasis. Using computer-assisted drug design (CADD) and a microtiter plate-based DNA-binding enzyme-linked immunosorbent assay (D-ELISA) to measure nuclear RUNX2 DNA binding, we have found that Vitamin D3 prohormones can modulate RUNX2 DNA binding, which was dose-dependent and sensitive to trypsin, salt, and phosphatase treatment. Unlabeled oligonucleotide or truncated, dominant negative RUNX2 proteins were competitive inhibitors of RUNX2 DNA binding. The RUNX2 heterodimeric partner, Cbfβ, was detected in the binding complexes with specific antibodies. Evaluation of several RUNX2:DNA targeted small molecules predicted by CADD screening revealed a previously unknown biological activity of the inactive Vitamin D3 precursor, cholecalciferol. Cholecalciferol modulated RUNX2:DNA binding at nanomolar concentrations even in cells with low VDR. Cholecalciferol and 25-OH Vitamin D3 prohormones were selective inhibitors of RUNX2-positive endothelial, bone, and breast cancer cell proliferation, but not of cells lacking RUNX2 expression. These compounds may have application in modulating RUNX2 activity in an angiogenic setting, in metastatic cells, and to promote bone formation in disease-mediated osteoporosis. The combination CADD discovery and D-ELISA screening approaches allows the testing of other novel derivatives of Vitamin D and/or transcriptional inhibitors with the potential to regulate DNA binding and biological function.

  10. Vibrio parahaemolyticus CalR down regulates the thermostable direct hemolysin (TDH) gene transcription and thereby inhibits hemolytic activity.

    Science.gov (United States)

    Zhang, Yiquan; Zhang, Ying; Gao, He; Zhang, Lingyu; Yin, Zhe; Huang, Xinxiang; Zhou, Dongsheng; Yang, Huiying; Yang, Wenhui; Wang, Li

    2017-03-04

    TDH, encoded by tdh gene, is a major virulent determinant of V. parahaemolyticus that controls various biological activities, such as hemolytic activity, cytotoxicity, and enterotoxicity. The hemolytic activity on Wagatsuma agar ascribed to TDH is called Kanagawa phenomenon (KP). All KP positive strains contain tdh1 and tdh2 genes, but tdh2 is predominantly responsible for KP. CalR is a regulatory protein that was originally identified as a repressor of swarming motility and T3SS1 gene expression in V. parahaemolyticus. In the present study, the regulation of tdh2 by CalR was investigated using a set of experiments including qRT-PCR, primer extension, LacZ fusion, hemolytic phenotype, EMSA, and DNase I footprinting assays. The results showed that His-CalR protected a single region from 224bp to 318bp upstream of tdh2 against DNase I digestion, and a transcriptional start site located at 42bp upstream of tdh2 was detected and its transcribed activity was inhibited by CalR. Moreover, the KP test results showed that the hemolytic activity of V. parahaemolyticus is also under negative control of CalR. The data demonstrated that CalR is a repressor of the tdh2 transcription and thereby inhibits the hemolytic activity of V. parahaemolyticus.

  11. The activation of peroxisome proliferator-activated receptor γ is regulated by Krüppel-like transcription factors 6 & 9 under steatotic conditions.

    Science.gov (United States)

    Escalona-Nandez, Ivonne; Guerrero-Escalera, Dafne; Estanes-Hernández, Alma; Ortíz-Ortega, Victor; Tovar, Armando R; Pérez-Monter, Carlos

    2015-03-20

    Liver steatosis is characterised by lipid droplet deposition in hepatocytes that can leads to an inflammatory and fibrotic phenotype. Peroxisome proliferator-activated receptors (PPARs) play key roles in energetic homeostasis by regulating lipid metabolism in hepatic tissue. In adipose tissue PPARγ regulates the adipocyte differentiation by promoting the expression of lipid-associated genes. Within the liver PPARγ is up-regulated under steatotic conditions; however, which transcription factors participate in its expression is not completely understood. Krüppel-like transcription factors (KLFs) regulate various cellular mechanisms, such as cell proliferation and differentiation. KLFs are key components of adipogenesis by regulating the expression of PPARγ and other proteins such as the C-terminal enhancer binding protein (C/EBP). Here, we demonstrate that the transcript levels of Klf6, Klf9 and Pparγ are increased in response to a steatotic insult in vitro. Chromatin immunoprecipitation (ChIp) experiments showed that klf6 and klf9 are actively recruited to the Pparγ promoter region under these conditions. Accordingly, the loss-of-function experiments reduced cytoplasmic triglyceride accumulation. Here, we demonstrated that KLF6 and KLF9 proteins directly regulate PPARγ expression under steatotic conditions. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Grape seed extract regulates androgen receptor-mediated transcription in prostate cancer cells through potent anti-histone acetyltransferase activity.

    Science.gov (United States)

    Park, Si Yong; Lee, Yoo-Hyun; Choi, Kyung-Chul; Seong, Ah-Reum; Choi, Hyo-Kyoung; Lee, Ok-Hee; Hwang, Han-Joon; Yoon, Ho-Geun

    2011-01-01

    Histone acetylation, which is regulated by histone acetyltransferases (HATs) and deacetylases, is an epigenetic mechanism that influences eukaryotic transcription. Significant changes in histone acetylation are associated with cancer; therefore, manipulating the acetylation status of key gene targets is likely crucial for effective cancer therapy. Grape seed extract (GSE) has a known protective effect against prostate cancer. Here, we showed that GSE significantly inhibited HAT activity by 30-80% in vitro (P cancer cells by measuring luciferase activity using a pGL3-PSA construct bearing the AR element in the human prostate cancer cell line LNCaP (P cancer cell growth, and implicate GSE as a novel candidate for therapeutic activity against prostate cancer.

  13. Synergy and Antagonism of Active Constituents of ADAPT-232 on Transcriptional Level of Metabolic Regulation of Isolated Neuroglial Cells.

    Science.gov (United States)

    Panossian, Alexander; Hamm, Rebecca; Kadioglu, Onat; Wikman, Georg; Efferth, Thomas

    2013-01-01

    interactions result in suppression some genes activated by individual substances. These interactions can have an influence on transcriptional control of metabolic regulation both on the cellular level and the level of the whole organism. Merging of deregulated genes array profiles and intracellular networks is specific to the new substance with unique pharmacological characteristics. Presumably, this phenomenon could be used to eliminate undesirable effects (e.g., toxic effects) and increase the selectivity of pharmacological intervention.

  14. Transcriptional regulation of the redD transcriptional activator gene accounts for growth-phase-dependent production of the antibiotic undecylprodigiosin in Streptomyces coelicolor A3(2)

    NARCIS (Netherlands)

    Takano, E.; Gramajo, H.C.; Strauch, E.; White, J.; Bibb, M.J.

    1992-01-01

    Transcription of redD, the activator gene required for production of the red-pigmented antibiotic undecylprodigiosin by Streptomyces coelicolor A3(2), showed a dramatic increase during the transition from exponential to stationary phase. The increase in redD expression was followed by transcription

  15. Peroxisome proliferator-activated receptor alpha, PPARα, directly regulates transcription of cytochrome P450 CYP2C8.

    Science.gov (United States)

    Thomas, Maria; Winter, Stefan; Klumpp, Britta; Turpeinen, Miia; Klein, Kathrin; Schwab, Matthias; Zanger, Ulrich M

    2015-01-01

    The cytochrome P450, CYP2C8, metabolizes more than 60 clinically used drugs as well as endogenous substances including retinoic acid and arachidonic acid. However, predictive factors for interindividual variability in the efficacy and toxicity of CYP2C8 drug substrates are essentially lacking. Recently we demonstrated that peroxisome proliferator-activated receptor alpha (PPARα), a nuclear receptor primarily involved in control of lipid and energy homeostasis directly regulates the transcription of CYP3A4. Here we investigated the potential regulation of CYP2C8 by PPARα. Two linked intronic SNPs in PPARα (rs4253728, rs4823613) previously associated with hepatic CYP3A4 status showed significant association with CYP2C8 protein level in human liver samples (N = 150). Furthermore, siRNA-mediated knock-down of PPARα in HepaRG human hepatocyte cells resulted in up to ∼60 and ∼50% downregulation of CYP2C8 mRNA and activity, while treatment with the PPARα agonist WY14,643 lead to an induction by >150 and >100%, respectively. Using chromatin immunoprecipitation scanning assay we identified a specific upstream gene region that is occupied in vivo by PPARα. Electromobility shift assay demonstrated direct binding of PPARα to a DR-1 motif located at positions -2762/-2775 bp upstream of the CYP2C8 transcription start site. We further validated the functional activity of this element using luciferase reporter gene assays in HuH7 cells. Moreover, based on our previous studies we demonstrated that WNT/β-catenin acts as a functional inhibitor of PPARα-mediated inducibility of CYP2C8 expression. In conclusion, our data suggest direct involvement of PPARα in both constitutive and inducible regulation of CYP2C8 expression in human liver, which is further modulated by WNT/β-catenin pathway. PPARA gene polymorphism could have a modest influence on CYP2C8 phenotype.

  16. Peroxisome proliferator-activated receptor alpha, PPARα, directly regulates transcription of cytochrome P450 CYP2C8

    Directory of Open Access Journals (Sweden)

    Maria eThomas

    2015-11-01

    Full Text Available The cytochrome P450, CYP2C8, metabolises more than 60 clinically used drugs as well as endogenous substances including retinoic acid and arachidonic acid. However predictive factors for interindividual variability in the efficacy and toxicity of CYP2C8 drug substrates are essentially lacking. Recently we demonstrated that peroxisome proliferator-activated receptor alpha (PPARα, a nuclear receptor primarily involved in control of lipid and energy homeostasis directly regulates the transcription of CYP3A4. Here we investigated the potential regulation of CYP2C8 by PPARα. Two linked intronic SNPs in PPARα (rs4253728, rs4823613 previously associated with hepatic CYP3A4 status showed significant association with CYP2C8 protein level in human liver samples (N=150. Furthermore, siRNA-mediated knock-down of PPARα in HepaRG human hepatocyte cells resulted in up to ~60% and ~50% downregulation of CYP2C8 mRNA and activity, while treatment with the PPARα agonist WY14,643 lead to an induction by >150% and >100%, respectively. Using chromatin immunoprecipitation scanning assay we identified a specific upstream gene region that is occupied in vivo by PPARα. Electromobility shift assay demonstrated direct binding of PPARα to a DR-1 motif located at positions -2762/-2775bp upstream of the CYP2C8 transcription start site. We further validated the functional activity of this element using luciferase reporter gene assays in HuH7 cells. Moreover, based on our previous studies we demonstrated that WNT/β-catenin acts as a functional inhibitor of PPARα-mediated inducibility of CYP2C8 expression. In conclusion, our data suggest direct involvement of PPARα in both constitutive and inducible regulation of CYP2C8 expression in human liver, which is further modulated by WNT/ β-catenin pathway. PPARA gene polymorphism could have a modest influence on CYP2C8 phenotype.

  17. ID4 regulates transcriptional activity of wild type and mutant p53 via K373 acetylation.

    Science.gov (United States)

    Morton, Derrick J; Patel, Divya; Joshi, Jugal; Hunt, Aisha; Knowell, Ashley E; Chaudhary, Jaideep

    2017-01-10

    Given that mutated p53 (50% of all human cancers) is over-expressed in many cancers, restoration of mutant p53 to its wild type biological function has been sought after as cancer therapy. The conformational flexibility has allowed to restore the normal biological function of mutant p53 by short peptides and small molecule compounds. Recently, studies have focused on physiological mechanisms such as acetylation of lysine residues to rescue the wild type activity of mutant p53. Using p53 null prostate cancer cell line we show that ID4 dependent acetylation promotes mutant p53 DNA-binding capabilities to its wild type consensus sequence, thus regulating p53-dependent target genes leading to subsequent cell cycle arrest and apoptosis. Specifically, by using wild type, mutant (P223L, V274F, R175H, R273H), acetylation mimics (K320Q and K373Q) and non-acetylation mimics (K320R and K373R) of p53, we identify that ID4 promotes acetylation of K373 and to a lesser extent K320, in turn restoring p53-dependent biological activities. Together, our data provides a molecular understanding of ID4 dependent acetylation that suggests a strategy of enhancing p53 acetylation at sites K373 and K320 that may serve as a viable mechanism of physiological restoration of mutant p53 to its wild type biological function.

  18. The ETS-5 transcription factor regulates activity states in Caenorhabditis elegans by controlling satiety

    DEFF Research Database (Denmark)

    Juozaityte, Vaida; Pladevall-Morera, David; Podolska, Agnieszka

    2017-01-01

    Animal behavior is shaped through interplay among genes, the environment, and previous experience. As in mammals, satiety signals induce quiescence in Caenorhabditis elegans Here we report that the C. elegans transcription factor ETS-5, an ortholog of mammalian FEV/Pet1, controls satiety-induced ...

  19. Menin and RNF20 recruitment is associated with dynamic histone modifications that regulate signal transducer and activator of transcription 1 (STAT1-activated transcription of the interferon regulatory factor 1 gene (IRF1

    Directory of Open Access Journals (Sweden)

    Buro Lauren J

    2010-09-01

    Full Text Available Abstract Background Signal transducer and activator of transcription (STAT activation of gene expression is both rapid and transient, and when properly executed it affects growth, differentiation, homeostasis and the immune response, but when dysregulated it contributes to human disease. Transcriptional activation is regulated by alterations to the chromatin template. However, the role of histone modification at gene loci that are activated for transcription in response to STAT signaling is poorly defined. Results Using chromatin immunoprecipitation, we profiled several histone modifications during STAT1 activation of the interferon regulatory factor 1 gene (IRF1. Methylated lysine histone proteins H3K4me2, H3K4me3, H3K79me3, H3K36me3 and monoubiquitinated histone ubH2B are dynamic and correlate with interferon (IFNγ induction of STAT1 activity. Chemical inhibition of H3K4 methylation downregulates IRF1 transcription and decreases RNA polymerase II (Pol II occupancy at the IRF1 promoter. MEN1, a component of a complex proteins associated with Set1 (COMPASS-like complex and the hBRE1 component, RNF20, are localized to IRF1 in the uninduced state and are further recruited when IRF1 is activated. RNAi-mediated depletion of RNF20 lowers both ubH2B and H3K4me3, but surprisingly, upregulates IFNγ induced IRF1 transcription. The dynamics of phosphorylation in the C-terminal domain (CTD of Pol II are disrupted during gene activation as well. Conclusions H2B monoubiquitination promotes H3K4 methylation, but the E3 ubiquitin ligase, RNF20, is repressive of inducible transcription at the IRF1 gene locus, suggesting that ubH2B can, directly or indirectly, affect Pol II CTD phosphorylation cycling to exert control on ongoing transcription.

  20. Unique role of SRSF2 in transcription activation and diverse functions of the SR and hnRNP proteins in gene expression regulation.

    Science.gov (United States)

    Mo, Sudong; Ji, Xiong; Fu, Xiang-Dong

    2013-01-01

    Transcription pause release from gene promoters has been recognized to be a critical point for transcriptional regulation in higher eukaryotes. Recent studies suggest that regulatory RNAs are extensively involved in transcriptional control, which may enlist various RNA binding proteins. We recently showed a key role of SRSF2, a member of the SR family of splicing regulators, in binding to promoter-associated small RNA to mediate transcription pause release, a regulatory strategy akin to the function of the HIV Tat protein via binding to the TAR element in nascent RNA to activate transcription. In this report, we further dissect the structural requirement for SRSF2 to function as a transcription activator and extend the analysis to multiple SR and hnRNP proteins by using the MS2 tethering strategy. Our results reveal that SRSF2 is a unique SR protein that activates transcription in a position-dependent manner while three other SR proteins enhance translation in a position-independent fashion. In contrast, multiple hnRNP proteins appear to negatively influence mRNA levels, especially when tethered in the gene body. These findings suggest broad participation of RNA binding proteins in diverse aspects of regulated gene expression at both the transcriptional and posttranscriptional levels in mammalian cells.

  1. Regulated assembly of transcription factors and control of transcription initiation.

    Science.gov (United States)

    Beckett, D

    2001-11-30

    Proteins that function in regulation of transcription initiation are typically homo or hetero-oligomeric. Results of recent biophysical studies of transcription regulators indicate that the assembly of these proteins is often subject to regulation. This regulation of assembly dictates the frequency of transcription initiation via its influence on the affinity of a transcription regulator for DNA and its affect on target site selection. Factors that modulate transcription factor assembly include binding of small molecules, post-translational modification, DNA binding and interactions with other proteins. Here, the results of recent structural and/or thermodynamic studies of a number of transcription regulators that are subject to regulated assembly are reviewed. The accumulated data indicate that this phenomenon is ubiquitous and that mechanisms utilized in eukaryotes and prokaryotes share common features. Copyright 2001 Academic Press.

  2. Regulation of the transcriptional coactivator FHL2 licenses activation of the androgen receptor in castrate-resistant prostate cancer.

    Science.gov (United States)

    McGrath, Meagan J; Binge, Lauren C; Sriratana, Absorn; Wang, Hong; Robinson, Paul A; Pook, David; Fedele, Clare G; Brown, Susan; Dyson, Jennifer M; Cottle, Denny L; Cowling, Belinda S; Niranjan, Birunthi; Risbridger, Gail P; Mitchell, Christina A

    2013-08-15

    It is now clear that progression from localized prostate cancer to incurable castrate-resistant prostate cancer (CRPC) is driven by continued androgen receptor (AR), signaling independently of androgen. Thus, there remains a strong rationale to suppress AR activity as the single most important therapeutic goal in CRPC treatment. Although the expression of ligand-independent AR splice variants confers resistance to AR-targeted therapy and progression to lethal castrate-resistant cancer, the molecular regulators of AR activity in CRPC remain unclear, in particular those pathways that potentiate the function of mutant AR in CRPC. Here, we identify FHL2 as a novel coactivator of ligand-independent AR variants that are important in CRPC. We show that the nuclear localization of FHL2 and coactivation of the AR is driven by calpain cleavage of the cytoskeletal protein filamin, a pathway that shows differential activation in prostate epithelial versus prostate cancer cell lines. We further identify a novel FHL2-AR-filamin transcription complex, revealing how deregulation of this axis promotes the constitutive, ligand-independent activation of AR variants, which are present in CRPC. Critically, the calpain-cleaved filamin fragment and FHL2 are present in the nucleus only in CRPC and not benign prostate tissue or localized prostate cancer. Thus, our work provides mechanistic insight into the enhanced AR activation, most notably of the recently identified AR variants, including AR-V7 that drives CRPC progression. Furthermore, our results identify the first disease-specific mechanism for deregulation of FHL2 nuclear localization during cancer progression. These results offer general import beyond prostate cancer, given that nuclear FHL2 is characteristic of other human cancers where oncogenic transcription factors that drive disease are activated like the AR in prostate cancer.

  3. The nickel-responsive regulator NikR controls activation and repression of gene transcription in Helicobacter pylori.

    NARCIS (Netherlands)

    F.D.J. Ernst (Florian); E.J. Kuipers (Ernst); A. Heijens (Angela); R. Sarwari (Roya); J. Stoof (Jeroen); C.W. Penn (Charles); J.G. Kusters (Johannes); A.H.M. van Vliet (Arnoud)

    2005-01-01

    textabstractThe NikR protein is a nickel-dependent regulatory protein which is a member of the ribbon-helix-helix family of transcriptional regulators. The gastric pathogen Helicobacter pylori expresses a NikR ortholog, which was previously shown to mediate regulation of metal metabolism and urease

  4. Analysis of DNA binding and transcriptional activation by the LysR-type transcriptional regulator CbbR of Xanthobacter flavus

    NARCIS (Netherlands)

    van Keulen, G; Ridder, ANJA; Dijkhuizen, L; Meijer, WG; Meijer, Wim G.

    2003-01-01

    The LysR-type transcriptional regulator CbbR controls the expression of the cbb and gap-pgk operons in Xanthobacter flavus, which encode the majority of the enzymes of the Calvin cycle required for autotrophic CO2 fixation. The cbb operon promoter of this chemoautotrophic bacterium contains three po

  5. p38 mitogen-activated protein kinase up-regulates NF-{kappa}B transcriptional activation through RelA phosphorylation during stretch-induced myogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Ji, Guoping [Department of Orthodontics, College of Stomatology, Ninth People' s Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai Research Institute of Stomatology, Shanghai 200011 (China); Liu, Dongxu [Department of Orthodontics, College of Stomatology, Shandong University, Jinan, Shandong Province 250012 (China); Liu, Jing [Department of Orthodontics, The Affiliated Qingdao Municipal Hospital, Qingdao University, Qingdao, Shandong Province 266075 (China); Gao, Hui [Department of Orthodontics, Tianjin Stomatological Hospital, Tianjin 300041 (China); Yuan, Xiao, E-mail: yuanxiaoqd@163.com [Department of Orthodontics, The Affiliated Qingdao Municipal Hospital, Qingdao University, Qingdao, Shandong Province 266075 (China); Shen, Gang, E-mail: ganshen2007@163.com [Department of Orthodontics, College of Stomatology, Ninth People' s Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai Research Institute of Stomatology, Shanghai 200011 (China)

    2010-01-01

    p38 MAPK and nuclear factor-B (NF-B) signaling pathways play an indispensable role in the control of skeletal myogenesis. The specific contribution of these signaling pathways to the response of myoblast to the mechanical stimulation and the molecular mechanisms underlying this response remain unresolved. Using an established in vitro model, we now show that p38 MAP kinase activity regulates the transcriptional activation of NF-{kappa}B in response to mechanical stimulation of myoblasts. Furthermore, SB203580 blocked stretch-induced NF-{kappa}B activation during myogenesis, not through down-regulation of degradation of I{kappa}B-{alpha}, and consequent translocation of the p65 subunit of NF-{kappa}B to the nucleus. It is likely that stretch-induced NF-{kappa}B activation by phosphorylation of p65 NF-{kappa}B. Moreover, depletion of p38{alpha} using siRNA significantly reduces stretch-induced phosphorylation of RelA and NF-{kappa}B activity. These results provides the first evidence of a cross-talk between p38 MAPK and NF-{kappa}B signaling pathways during stretch-induced myogenesis, with phosphorylation of RelA being one of the effectors of this promyogenic mechanism. The {alpha} isoform of p38MAP kinase regulates the transcriptional activation of NF-{kappa}B following stimulation with cyclic stretch.

  6. Transcriptional activator Cat8 is involved in regulation of xylose alcoholic fermentation in the thermotolerant yeast Ogataea (Hansenula) polymorpha.

    Science.gov (United States)

    Ruchala, Justyna; Kurylenko, Olena O; Soontorngun, Nitnipa; Dmytruk, Kostyantyn V; Sibirny, Andriy A

    2017-02-28

    Efficient xylose alcoholic fermentation is one of the key to a successful lignocellulosic ethanol production. However, regulation of this process in the native xylose-fermenting yeasts is poorly understood. In this work, we paid attention to the transcriptional factor Cat8 and its possible role in xylose alcoholic fermentation in Ogataea (Hansenula) polymorpha. In Saccharomyces cerevisiae, organism, which does not metabolize xylose, gene CAT8 encodes a Zn-cluster transcriptional activator necessary for expression of genes involved in gluconeogenesis, respiration, glyoxylic cycle and ethanol utilization. Xylose is a carbon source that could be fermented to ethanol and simultaneously could be used in gluconeogenesis for hexose synthesis. This potentially suggests involvement of CAT8 in xylose metabolism. Here, the role of CAT8 homolog in the natural xylose-fermenting thermotolerant yeast O. polymorpha was characterized. The CAT8 ortholog was identified in O. polymorpha genome and deleted both in the wild-type strain and in advanced ethanol producer from xylose. Constructed cat8Δ strain isolated from wild strain showed diminished growth on glycerol, ethanol and xylose as well as diminished respiration on the last substrate. At the same time, cat8Δ mutant isolated from the best available O. polymorpha ethanol producer showed only visible defect in growth on ethanol. CAT8 deletant was characterized by activated transcription of genes XYL3, DAS1 and RPE1 and slight increase in the activity of several enzymes involved in xylose metabolism and alcoholic fermentation. Ethanol production from xylose in cat8Δ mutants in the background of wild-type strain and the best available ethanol producer from xylose increased for 50 and 30%, respectively. The maximal titer of ethanol during xylose fermentation was 12.5 g ethanol/L at 45 °C. Deletion of CAT8 did not change ethanol production from glucose. Gene CAT8 was also overexpressed under control of the strong constitutive

  7. The Hinge-Helix 1 Region of Peroxisome Proliferator-Activated Receptor γ1 (PPARγ1) Mediates Interaction with Extracellular Signal-Regulated Kinase 5 and PPARγ1 Transcriptional Activation: Involvement in Flow-Induced PPARγ Activation in Endothelial Cells

    OpenAIRE

    Akaike, Masashi; Che, Wenyi; Marmarosh, Nicole-Lerner; Ohta, Shinsuke; Osawa, Masaki; Ding,Bo; Berk, Bradford C.; Yan, Chen; Abe, Jun-ichi

    2004-01-01

    Peroxisome proliferator-activated receptors (PPAR) are ligand-activated transcription factors that form a subfamily of the nuclear receptor gene family. Since both flow and PPARγ have atheroprotective effects and extracellular signal-regulated kinase 5 (ERK5) kinase activity is significantly increased by flow, we investigated whether ERK5 kinase regulates PPARγ activity. We found that activation of ERK5 induced PPARγ1 activation in endothelial cells (ECs). However, we could not detect PPARγ p...

  8. CREB-regulated transcription co-activator family stimulates promoter II-driven aromatase expression in preadipocytes.

    Science.gov (United States)

    Samarajeewa, Nirukshi U; Docanto, Maria M; Simpson, Evan R; Brown, Kristy A

    2013-08-01

    The dramatically increased prevalence of breast cancer after menopause is of great concern and is correlated with elevated local levels of estrogens. This is mainly due to an increase in aromatase expression driven by its proximal promoter II (PII). We have previously demonstrated that the CREB co-activator CRTC2 binds directly to PII and stimulates its activity via mechanisms involving LKB1-AMPK in response to prostaglandin E(2) (PGE(2)). There are three members of the CRTC family (CRTC1-3) and this study aimed to characterize the role of other CRTCs in the activation of aromatase PII. The expression and subcellular localization of CRTCs were examined in preadipocytes using qPCR and immunofluorescence. Under basal conditions, CRTC1 expression was the lowest, whereas CRTC3 transcripts were present at higher levels. Basally, CRTC2 and CRTC3 were mainly cytoplasmic and PGE(2) caused their nuclear translocation. Reporter assays and chromatin immunoprecipitation (ChIP) were performed to assess the effect of CRTCs on PII activity and binding. Basal PII activity was significantly increased with all CRTCs. Forskolin (FSK)/phorbol 12-myristate 13-acetate (PMA), to mimic PGE(2), resulted in a further significant increase in PII activity with all CRTCs, with CRTC2 and CRTC3 having greater effects. This was consistent with ChIP data showing an increased binding of CRTCs to PII with FSK/PMA. Moreover, gene silencing of CRTC2 and CRTC3 significantly reduced the FSK/PMA-mediated stimulation of aromatase activity. Interestingly, CRTCs acted cooperatively with CREB1 to increase PII activity, and both CREs were found to be essential for the maximal induction of PII activity by CRTCs. Phosphorylation of CRTC2 at its AMPK target site, Ser 171, dictated its subcellular localization, and the activation of aromatase PII in preadipocytes. In conclusion, this study demonstrates that aromatase regulation in primary human breast preadipocytes involves more than one CRTC.

  9. An activating transcription factor of Litopenaeus vannamei involved in WSSV genes Wsv059 and Wsv166 regulation.

    Science.gov (United States)

    Li, Xiao-Yun; Yue, Hai-Tao; Zhang, Ze-Zhi; Bi, Hai-Tao; Chen, Yong-Gui; Weng, Shao-Ping; Chan, Siuming; He, Jian-Guo; Chen, Yi-Hong

    2014-12-01

    Members of activating transcription factor/cyclic adenosine 3', 5'-monophosphate response element binding protein (ATF/CREB) family are induced by various stress signals and function as effector molecules. Consequently, cellular changes occur in response to discrete sets of instructions. In this work, we found an ATF transcription factor in Litopenaeus vannamei designated as LvATFβ. The full-length cDNA of LvATFβ was 1388 bp long with an open reading frame of 939 bp that encoded a putative 313 amino acid protein. The protein contained a basic region-leucine zipper (bZip) domain that was a common feature among ATF/CREB transcription factors. LvATFβ was highly expressed in intestines, gills, and heart. LvATFβ expression was dramatically upregulated by white spot syndrome virus (WSSV) infection. Pull-down assay revealed that LvATFβ had strong affinity to promoters of WSSV genes, namely, wsv059 and wsv166. Dual-luciferase reporter assay showed that LvATFβ could upregulate the expression of wsv059 and wsv166. Knocked down LvATFβ resulted in decreased expression of wsv059 and wsv166 in WSSV-challenged L. vannamei. Knocked down expression of wsv059 and wsv166 by RNA interference inhibited the replication and reduce the mortality of L. vannamei during WSSV challenge inoculation. The copy numbers of WSSV in wsv059 and wsv166 knocked down group were significant lower than in the control. These results suggested that LvATFβ may be involved in WSSV replication by regulating the expression of wsv059 and wsv166.

  10. Transcription activator-like effector-mediated regulation of gene expression based on the inducible packaging and delivery via designed extracellular vesicles.

    Science.gov (United States)

    Lainšček, Duško; Lebar, Tina; Jerala, Roman

    2017-02-26

    Transcription activator-like effector (TALE) proteins present a powerful tool for genome editing and engineering, enabling introduction of site-specific mutations, gene knockouts or regulation of the transcription levels of selected genes. TALE nucleases or TALE-based transcription regulators are introduced into mammalian cells mainly via delivery of the coding genes. Here we report an extracellular vesicle-mediated delivery of TALE transcription regulators and their ability to upregulate the reporter gene in target cells. Designed transcriptional activator TALE-VP16 fused to the appropriate dimerization domain was enriched as a cargo protein within extracellular vesicles produced by mammalian HEK293 cells stimulated by Ca-ionophore and using blue light- or rapamycin-inducible dimerization systems. Blue light illumination or rapamycin increased the amount of the TALE-VP16 activator in extracellular vesicles and their addition to the target cells resulted in an increased expression of the reporter gene upon addition of extracellular vesicles to the target cells. This technology therefore represents an efficient delivery for the TALE-based transcriptional regulators.

  11. The activation of peroxisome proliferator-activated receptor γ is regulated by Krüppel-like transcription factors 6 & 9 under steatotic conditions

    Energy Technology Data Exchange (ETDEWEB)

    Escalona-Nandez, Ivonne; Guerrero-Escalera, Dafne; Estanes-Hernández, Alma [Departamento de Gastroenterología, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Vasco de Quiroga 15 Sección XVI, Tlalpan, 14000, México, D.F. (Mexico); Ortíz-Ortega, Victor; Tovar, Armando R. [Departamento de Fisiología de la Nutrición, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Vasco de Quiroga 15 Sección XVI, Tlalpan, 14000, México, D.F. (Mexico); Pérez-Monter, Carlos, E-mail: carlos.perezm@incmnsz.mx [Departamento de Gastroenterología, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Vasco de Quiroga 15 Sección XVI, Tlalpan, 14000, México, D.F. (Mexico)

    2015-03-20

    Liver steatosis is characterised by lipid droplet deposition in hepatocytes that can leads to an inflammatory and fibrotic phenotype. Peroxisome proliferator-activated receptors (PPARs) play key roles in energetic homeostasis by regulating lipid metabolism in hepatic tissue. In adipose tissue PPARγ regulates the adipocyte differentiation by promoting the expression of lipid-associated genes. Within the liver PPARγ is up-regulated under steatotic conditions; however, which transcription factors participate in its expression is not completely understood. Krüppel-like transcription factors (KLFs) regulate various cellular mechanisms, such as cell proliferation and differentiation. KLFs are key components of adipogenesis by regulating the expression of PPARγ and other proteins such as the C-terminal enhancer binding protein (C/EBP). Here, we demonstrate that the transcript levels of Klf6, Klf9 and Pparγ are increased in response to a steatotic insult in vitro. Chromatin immunoprecipitation (ChIp) experiments showed that klf6 and klf9 are actively recruited to the Pparγ promoter region under these conditions. Accordingly, the loss-of-function experiments reduced cytoplasmic triglyceride accumulation. Here, we demonstrated that KLF6 and KLF9 proteins directly regulate PPARγ expression under steatotic conditions. - Highlights: • Palmitic acid promotes expression of KlF6 & KLF9 in HepG2 cells. • KLF6 and KLF9 promote the expression of PPARγ in response to palmitic acid. • Binding of KLF6 and KLF9 to the PPARγ promoter promotes steatosis in HepG2 cells. • KLF6 and KLF9 loss-of function diminishes the steatosis in HepG2 cells.

  12. Arabidopsis ABI5 plays a role in regulating ROS homeostasis by activating CATALASE 1 transcription in seed germination.

    Science.gov (United States)

    Bi, Chao; Ma, Yu; Wu, Zhen; Yu, Yong-Tao; Liang, Shan; Lu, Kai; Wang, Xiao-Fang

    2017-05-01

    It has been known that ABA INSENSITIVE 5 (ABI5) plays a vital role in regulating seed germination. In the present study, we showed that inhibition of the catalase activity with 3-amino-1,2,4-triazole (3-AT) inhibits seed germination of Col-0, abi5 mutants and ABI5-overexpression transgenic lines. Compared with Col-0, the seeds of abi5 mutants showed more sensitive to 3-AT during seed germination, while the seeds of ABI5-overexpression transgenic lines showed more insensitive. H2O2 showed the same effect on seed germination of Col-0, abi5 mutants and ABI5-overexpression transgenic lines as 3-AT. These results suggest that ROS is involved in the seed germination mediated by ABI5. Further, we observed that T-DNA insertion mutants of the three catalase members in Arabidopsis displayed 3-AT-insensitive or -hypersensitive phenotypes during seed germination, suggesting that these catalase members regulate ROS homeostasis in a highly complex way. ABI5 affects reactive oxygen species (ROS) homeostasis by affecting CATALASE expression and catalase activity. Furthermore, we showed that ABI5 directly binds to the CAT1 promoter and activates CAT1 expression. Genetic evidence supports the idea that CAT1 functions downstream of ABI5 in ROS signaling during seed germination. RNA-sequencing analysis indicates that the transcription of the genes involved in ROS metabolic process or genes responsive to ROS stress is impaired in abi5-1 seeds. Additionally, expression changes in some genes correlative to seed germination were showed due to the change in ABI5 expression under 3-AT treatment. Together, all the findings suggest that ABI5 regulates seed germination at least partly by affecting ROS homeostasis.

  13. Heat shock transcription factor 1 is activated as a consequence of lymphocyte activation and regulates a major proteostasis network in T cells critical for cell division during stress.

    Science.gov (United States)

    Gandhapudi, Siva K; Murapa, Patience; Threlkeld, Zachary D; Ward, Martin; Sarge, Kevin D; Snow, Charles; Woodward, Jerold G

    2013-10-15

    Heat shock transcription factor 1 (HSF1) is a major transcriptional regulator of the heat shock response in eukaryotic cells. HSF1 is evoked in response to a variety of cellular stressors, including elevated temperatures, oxidative stress, and other proteotoxic stressors. Previously, we demonstrated that HSF1 is activated in naive T cells at fever range temperatures (39.5°C) and is critical for in vitro T cell proliferation at fever temperatures. In this study, we demonstrated that murine HSF1 became activated to the DNA-binding form and transactivated a large number of genes in lymphoid cells strictly as a consequence of receptor activation in the absence of apparent cellular stress. Microarray analysis comparing HSF1(+/+) and HSF1(-/-) gene expression in T cells activated at 37°C revealed a diverse set of 323 genes significantly regulated by HSF1 in nonstressed T cells. In vivo proliferation studies revealed a significant impairment of HSF1(-/-) T cell expansion under conditions mimicking a robust immune response (staphylococcal enterotoxin B-induced T cell activation). This proliferation defect due to loss of HSF1 is observed even under nonfebrile temperatures. HSF1(-/-) T cells activated at fever temperatures show a dramatic reduction in cyclin E and cyclin A proteins during the cell cycle, although the transcription of these genes was modestly affected. Finally, B cell and hematopoietic stem cell proliferation from HSF1(-/-) mice, but not HSF1(+/+) mice, were also attenuated under stressful conditions, indicating that HSF1 is critical for the cell cycle progression of lymphoid cells activated under stressful conditions.

  14. An overview on transcriptional regulators in Streptomyces.

    Science.gov (United States)

    Romero-Rodríguez, Alba; Robledo-Casados, Ivonne; Sánchez, Sergio

    2015-08-01

    Streptomyces are Gram-positive microorganisms able to adapt and respond to different environmental conditions. It is the largest genus of Actinobacteria comprising over 900 species. During their lifetime, these microorganisms are able to differentiate, produce aerial mycelia and secondary metabolites. All of these processes are controlled by subtle and precise regulatory systems. Regulation at the transcriptional initiation level is probably the most common for metabolic adaptation in bacteria. In this mechanism, the major players are proteins named transcription factors (TFs), capable of binding DNA in order to repress or activate the transcription of specific genes. Some of the TFs exert their action just like activators or repressors, whereas others can function in both manners, depending on the target promoter. Generally, TFs achieve their effects by using one- or two-component systems, linking a specific type of environmental stimulus to a transcriptional response. After DNA sequencing, many streptomycetes have been found to have chromosomes ranging between 6 and 12Mb in size, with high GC content (around 70%). They encode for approximately 7000 to 10,000 genes, 50 to 100 pseudogenes and a large set (around 12% of the total chromosome) of regulatory genes, organized in networks, controlling gene expression in these bacteria. Among the sequenced streptomycetes reported up to now, the number of transcription factors ranges from 471 to 1101. Among these, 315 to 691 correspond to transcriptional regulators and 31 to 76 are sigma factors. The aim of this work is to give a state of the art overview on transcription factors in the genus Streptomyces.

  15. Transcription factor ZBED6 mediates IGF2 gene expression by regulating promoter activity and DNA methylation in myoblasts

    Science.gov (United States)

    Zinc finger, BED-type containing 6 (ZBED6) is an important transcription factor in placental mammals, affecting development, cell proliferation and growth. In this study, we found that the expression of the ZBED6 and IGF2 were up regulated during C2C12 differentiation. The IGF2 expression levels wer...

  16. LRP-1 ectodomain shedding as a post-transcriptional mechanism to regulate its endocytic activity

    OpenAIRE

    Selvais, Charlotte

    2010-01-01

    The low-density lipoprotein (LDL) receptor-related protein-1 (LRP-1) is a type I integral membrane receptor that mediates endocytosis of a wide array of ligands. LRP-1-dependent endocytosis emerges as a main mechanism controlling the extracellular amounts of various matrix metalloproteinase (MMP) family members. The LRP-1-mediated internalization of MMP-2 was identified as a pivotal mechanism for controlling the extracellular activity of this proteinase. In particular, we have documented that...

  17. Transcriptional activity of neural retina leucine zipper (Nrl) is regulated by c-Jun N-terminal kinase and Tip60 during retina development.

    Science.gov (United States)

    Kim, Jung-Woong; Jang, Sang-Min; Kim, Chul-Hong; An, Joo-Hee; Choi, Kyung-Hee

    2012-05-01

    Neural retina leucine zipper (Nrl), a key basic motif leucine zipper (bZIP) transcription factor, modulates rod photoreceptor differentiation by activating rod-specific target genes. In searching for factors that might couple with Nrl to modulate its transcriptional activity through posttranslational modification, we observed the novel interactions of Nrl with c-Jun N-terminal kinase 1 (JNK1) and HIV Tat-interacting protein 60 (Tip60). JNK1 directly interacted with and phosphorylated Nrl at serine 50, which enhanced Nrl transcriptional activity on the rhodopsin and Ppp2r5c promoters. Use of an inactive JNK1 mutant or treatment with a JNK inhibitor (SP600125) significantly reduced JNK1-mediated phosphorylation and transcriptional activity of Nrl in cultured retinal explants. We also found that Nrl activated rhodopsin and Ppp2r5c transcription by recruiting Tip60 to promote histone H3/H4 acetylation. The binding affinity of phospho-Nrl for Tip60 was significantly greater than that of the unphosphorylated Nrl. Thus, the histone acetyltransferase-containing Tip60 behaved as a coactivator in the Nrl-dependent transcriptional regulation of the rhodopsin and Ppp2r5c genes in the developing mouse retina. A transcriptional network of interactive proteins, including Nrl, JNK1, and Tip60, may be required to precisely control spatiotemporal photoreceptor-specific gene expression during retinal development.

  18. Targeted genome regulation via synthetic programmable transcriptional regulators

    KAUST Repository

    Piatek, Agnieszka Anna

    2016-04-19

    Regulation of gene transcription controls cellular functions and coordinates responses to developmental, physiological and environmental cues. Precise and efficient molecular tools are needed to characterize the functions of single and multiple genes in linear and interacting pathways in a native context. Modular DNA-binding domains from zinc fingers (ZFs) and transcriptional activator-like proteins (TALE) are amenable to bioengineering to bind DNA target sequences of interest. As a result, ZF and TALE proteins were used to develop synthetic programmable transcription factors. However, these systems are limited by the requirement to re-engineer proteins for each new target sequence. The clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR associated 9 (Cas9) genome editing tool was recently repurposed for targeted transcriptional regulation by inactivation of the nuclease activity of Cas9. Due to the facile engineering, simplicity, precision and amenability to library construction, the CRISPR/Cas9 system is poised to revolutionize the functional genomics field across diverse eukaryotic species. In this review, we discuss the development of synthetic customizable transcriptional regulators and provide insights into their current and potential applications, with special emphasis on plant systems, in characterization of gene functions, elucidation of molecular mechanisms and their biotechnological applications. © 2016 Informa UK Limited, trading as Taylor & Francis Group

  19. The quorum sensing transcriptional regulator TraR has separate binding sites for DNA and the anti-activator

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Zhida; Fuqua, Clay [Department of Molecular and Cellular Biochemistry, 212 S. Hawthorne Dr. Simon Hall 400A, Indiana University, Bloomington, IN 47405 (United States); Chen, Lingling, E-mail: linchen@indiana.edu [Department of Molecular and Cellular Biochemistry, 212 S. Hawthorne Dr. Simon Hall 400A, Indiana University, Bloomington, IN 47405 (United States)

    2012-02-10

    Highlights: Black-Right-Pointing-Pointer Quorum sensing transcription factor TraR is inhibited by forming TraR-TraM complex. Black-Right-Pointing-Pointer K213 is a key DNA binding residue, but not involved in interaction with TraM. Black-Right-Pointing-Pointer Mutations of TraM-interacting TraR residues did not affect DNA-binding of TraR. Black-Right-Pointing-Pointer Mutations of TraR residues reduced the TraR-TraM interaction more than those of TraM. Black-Right-Pointing-Pointer TraM inhibition on DNA-binding of TraR is driven by thermodynamics. -- Abstract: Quorum sensing represents a mechanism by which bacteria control their genetic behaviors via diffusible signals that reflect their population density. TraR, a quorum sensing transcriptional activator in the Rhizobiaceae family, is regulated negatively by the anti-activator TraM via formation of a TraR-TraM heterocomplex. Prior structural analysis suggests that TraM and DNA bind to TraR in distinct sites. Here we combined isothermal titration calorimetry (ITC) and electrophoretic mobility shift assays (EMSA) to investigate roles of TraR residues from Rhizobium sp. NGR234 in binding of both TraM and DNA. We found that K213A mutation of TraR{sub NGR} abolished DNA binding, however, did not alter TraM binding. Mutations of TraM-interfacing TraR{sub NGR} residues decreased the TraR-TraM interaction, but did not affect the DNA-binding activity of TraR{sub NGR}. Thus, our biochemical studies support the independent binding sites on TraR for TraM and DNA. We also found that point mutations in TraR{sub NGR} appeared to decrease the TraR-TraM interaction more effectively than those in TraM{sub NGR}, consistent with structural observations that individual TraR{sub NGR} residues contact with more TraM{sub NGR} residues than each TraM{sub NGR} residues with TraR{sub NGR} residues. Finally, we showed that TraM inhibition on DNA-binding of TraR was driven thermodynamically. We discussed subtle mechanistic differences in Tra

  20. The transcriptional regulation of pluripotency

    Institute of Scientific and Technical Information of China (English)

    Jia-Chi Yeo; Huck-Hui Ng

    2013-01-01

    The defining features of embryonic stem cells (ESCs) are their self-renewing and pluripotent capacities.Indeed,the ability to give rise into all cell types within the organism not only allows ESCs to function as an ideal in vitro tool to study embryonic development,but also offers great therapeutic potential within the field of regenerative medicine.However,it is also this same remarkable developmental plasticity that makes the efficient control of ESC differentiation into the desired cell type very difficult.Therefore,in order to harness ESCs for clinical applications,a detailed understanding of the molecular and cellular mechanisms controlling ESC pluripotency and lineage commitment is necessary.In this respect,through a variety of transcriptomic approaches,ESC pluripotency has been found to be regulated by a system of ESC-associated transcription factors; and the external signalling environment also acts as a key factor in modulating the ESC transcriptome.Here in this review,we summarize our current understanding of the transcriptional regulatory network in ESCs,discuss how the control of various signalling pathways could influence pluripotency,and provide a future outlook of ESC research.

  1. Switching on cilia: transcriptional networks regulating ciliogenesis.

    Science.gov (United States)

    Choksi, Semil P; Lauter, Gilbert; Swoboda, Peter; Roy, Sudipto

    2014-04-01

    Cilia play many essential roles in fluid transport and cellular locomotion, and as sensory hubs for a variety of signal transduction pathways. Despite having a conserved basic morphology, cilia vary extensively in their shapes and sizes, ultrastructural details, numbers per cell, motility patterns and sensory capabilities. Emerging evidence indicates that this diversity, which is intimately linked to the different functions that cilia perform, is in large part programmed at the transcriptional level. Here, we review our understanding of the transcriptional control of ciliary biogenesis, highlighting the activities of FOXJ1 and the RFX family of transcriptional regulators. In addition, we examine how a number of signaling pathways, and lineage and cell fate determinants can induce and modulate ciliogenic programs to bring about the differentiation of distinct cilia types.

  2. Dedicator of cytokinesis 8 regulates signal transducer and activator of transcription 3 activation and promotes TH17 cell differentiation.

    Science.gov (United States)

    Keles, Sevgi; Charbonnier, Louis Marie; Kabaleeswaran, Venkataraman; Reisli, Ismail; Genel, Ferah; Gulez, Nesrin; Al-Herz, Waleed; Ramesh, Narayanaswamy; Perez-Atayde, Antonio; Karaca, Neslihan E; Kutukculer, Necil; Wu, Hao; Geha, Raif S; Chatila, Talal A

    2016-11-01

    The autosomal recessive hyper-IgE syndrome (HIES) caused by dedicator of cytokinesis 8 (DOCK8) deficiency shares clinical features with autosomal dominant HIES because of signal transducer and activator of transcription 3 (STAT3) mutations, including recurrent infections and mucocutaneous candidiasis, which are suggestive of TH17 cell dysfunction. The mechanisms underlying this phenotypic overlap are unclear. We sought to elucidate common mechanisms operating in the different forms of HIES. We analyzed the differentiation of CD4(+) TH cell subsets in control and DOCK8-deficient subjects. We also examined the role of DOCK8 in regulating STAT3 activation in T cells. TH cell differentiation was analyzed by ELISA, flow cytometry, and real-time PCR measurements of cytokines and TH cell transcription factors. The interaction of DOCK8 and STAT3 signaling pathways was examined by using flow cytometry, immunofluorescence, coimmunoprecipitation, and gene expression analysis. There was a profound block in the differentiation of DOCK8-deficient naive CD4(+) T cells into TH17 cells. A missense mutation that disrupts DOCK8 guanine nucleotide exchange factor (GEF) activity while sparing protein expression also impaired TH17 cell differentiation. DOCK8 constitutively associated with STAT3 independent of GEF activity, whereas it regulated STAT3 phosphorylation in a GEF activity-dependent manner. DOCK8 also promoted STAT3 translocation to the nucleus and induction of STAT3-dependent gene expression. DOCK8 interacts with STAT3 and regulates its activation and the outcome of STAT3-dependent TH17 differentiation. These findings might explain the phenotypic overlap between DOCK8 deficiency and autosomal dominant HIES. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  3. The solution structure of DNA-free Pax-8 paired box domain accounts for redox regulation of transcriptional activity in the Pax protein family

    NARCIS (Netherlands)

    Codutti, L.; van Ingen, H.; Vascotto, C.; Fogolari, F.; Corazza, A.; Tell, G.; Quadrifoglio, F.; Viglino, P.; Boelens, R.; Esposito, G.

    2008-01-01

    Pax-8 is a transcription factor belonging to the PAX genes superfamily and its crucial role has been proven both in embryo and in the adult organism. Pax-8 activity is regulated via a redoxbased mechanism centered on the glutathionylation of specific cysteines in the N-terminal region (Cys45 and

  4. The transcriptional activators AraR and XlnR from Aspergillus niger regulate expression of pentose catabolic and pentose phosphate pathway genes

    NARCIS (Netherlands)

    Battaglia, Evy; Zhou, M.; de Vries, Ronald P; van den Brink, J.

    2014-01-01

    The pentose catabolic pathway (PCP) and the pentose phosphate pathway (PPP) are required for the conversion of pentose sugars in fungi and are linked via d-xylulose-5-phosphate. Previously, it was shown that the PCP is regulated by the transcriptional activators XlnR and AraR in Aspergillus niger. H

  5. GALNT3, a gene associated with hyperphosphatemic familial tumoral calcinosis, is transcriptionally regulated by extracellular phosphate and modulates matrix metalloproteinase activity

    OpenAIRE

    2008-01-01

    GALNT3, a gene associated with Hyperphosphatemic Familial Tumoral Calcinosis, is transcriptionally regulated by extracellular phosphate and modulates matrix metalloproteinase activity correspondence: Corresponding author. Laboratory of Molecular Dermatology Department of Dermatology Rambam Medical Center POB 9602, Haifa 31096, Israel. Tel.: +972 4 8541919; fax: +972 4 8542951. (Sprecher, Eli) (Sprecher, Eli) Labo...

  6. The Mediator complex and transcription regulation

    Science.gov (United States)

    Poss, Zachary C.; Ebmeier, Christopher C.

    2013-01-01

    The Mediator complex is a multi-subunit assembly that appears to be required for regulating expression of most RNA polymerase II (pol II) transcripts, which include protein-coding and most non-coding RNA genes. Mediator and pol II function within the pre-initiation complex (PIC), which consists of Mediator, pol II, TFIIA, TFIIB, TFIID, TFIIE, TFIIF and TFIIH and is approximately 4.0 MDa in size. Mediator serves as a central scaffold within the PIC and helps regulate pol II activity in ways that remain poorly understood. Mediator is also generally targeted by sequence-specific, DNA-binding transcription factors (TFs) that work to control gene expression programs in response to developmental or environmental cues. At a basic level, Mediator functions by relaying signals from TFs directly to the pol II enzyme, thereby facilitating TF-dependent regulation of gene expression. Thus, Mediator is essential for converting biological inputs (communicated by TFs) to physiological responses (via changes in gene expression). In this review, we summarize an expansive body of research on the Mediator complex, with an emphasis on yeast and mammalian complexes. We focus on the basics that underlie Mediator function, such as its structure and subunit composition, and describe its broad regulatory influence on gene expression, ranging from chromatin architecture to transcription initiation and elongation, to mRNA processing. We also describe factors that influence Mediator structure and activity, including TFs, non-coding RNAs and the CDK8 module. PMID:24088064

  7. Piwil2 modulates the proliferation and metastasis of colon cancer via regulation of matrix metallopeptidase 9 transcriptional activity.

    Science.gov (United States)

    Li, Dawei; Sun, Xing; Yan, Dongwang; Huang, Jianfeng; Luo, Qiongzhen; Tang, Huamei; Peng, Zhihai

    2012-10-01

    Piwi-like protein 2 (Piwil2) has recently emerged as a putative oncogene which is amplified in several human malignancies. However, the role of Piwil2 in colon cancer remains poorly understood. The aim of this study was to investigate the clinical and pathological significance of Piwil2, and the possible role in the proliferation and metastasis of colon cancer. Primary colon cancer paired with adjacent normal colon tissue and lymph node metastasis (LNM) lesions in 66 patients' tissue microarrays (TMA) were used to determine the expression of Piwil2. Knocked down Piwil2 expression in SW620 and SW480 colon cancer cell lines was performed to evaluate the role of Piwil2 in cell proliferation, invasion, metastasis in vitro and tumorigenicity in vivo. The possible roles of Piwil2 in the regulation of a 2 kb matrix metallopeptidase 9 (MMP9) promoter fragment and on the regulation of apoptotic pathways were evaluated by using a luciferase reporter construct and Western blots, respectively. Significantly higher expression levels of Piwil2 were observed in primary colon cancer tissue and in LNM in comparison with normal colon mucosa. Piwil2 expression significantly correlated with more aggressive clinical and pathological parameters with poorer five-year metastasis-free survival and overall survival. Piwil2 silencing significantly reduced cancer cell proliferation, colony formation ability and increased apoptosis in vitro and inhibited tumor growth in vivo. Piwil2 knockdown also attenuated migration and invasion of colon cancer cells via modulation of MMP9 transcriptional activities. Our results indicate that Piwil2 moderates the proliferation and metastasis potential of colon cancer.

  8. Transcription of Tnfaip3 is regulated by NF-κB and p38 via C/EBPβ in activated macrophages.

    Directory of Open Access Journals (Sweden)

    Ting-Yu Lai

    Full Text Available Macrophages play a pivotal role in the immune system through recognition and elimination of microbial pathogens. Toll-like receptors (TLRs on macrophages interact with microbial substances and initiate signal transduction through intracellular adapters. TLR4, which recognizes the lipopolysaccharides (LPS on Gram-positive and Gram-negative bacteria, triggers downstream signaling mediators and eventually activates IκB kinase (IKK complex and mitogen-activated protein kinases (MAPKs such as p38. Previous reports revealed that, in addition to NF-κB, a core transcription factor of the innate immune response, the induction of some LPS-induced genes in macrophages required another transcription factor whose activity depends on p38. However, these additional transcription factors remain to be identified. In order to identify p38-activated transcription factors that cooperate with NF-κB in response to LPS stimulation, microarrays were used to identify genes regulated by both NF-κB and p38 using wild-type, IKK-depleted, and p38 inhibitor-treated mouse bone marrow-derived macrophages (BMDMs. In silico analysis of transcription factor binding sites was used to predict the potential synergistic transcription factors from the co-expressed genes. Among these genes, NF-κB and C/EBPβ, a p38 downstream transcription factor, were predicted to co-regulate genes in LPS-stimulated BMDMs. Based on the subsequent results of a chromatin immunoprecipitation assay and TNFAIP3 expression in C/EBPβ-ablated macrophages, we demonstrated that Tnfaip3 is regulated by both NF-κB and p38-dependent C/EBPβ. These results identify a novel regulatory mechanism in TLR4-mediated innate immunity.

  9. Nickel-responsive transcriptional regulators.

    Science.gov (United States)

    Musiani, Francesco; Zambelli, Barbara; Bazzani, Micaela; Mazzei, Luca; Ciurli, Stefano

    2015-09-01

    Nickel is an essential micronutrient for a large number of living organisms, but it is also a toxic metal ion when it accumulates beyond the sustainable level as it may result if and when its cellular trafficking is not properly governed. Therefore, the homeostasis and metabolism of nickel is tightly regulated through metal-specific protein networks that respond to the available Ni(II) concentration. These are directed by specific nickel sensors, able to couple Ni(II) binding to a change in their DNA binding affinity and/or specificity, thus translating the cellular level of Ni(II) into a modification of the expression of the proteins devoted to modulating nickel uptake, efflux and cellular utilization. This review describes the Ni(II)-dependent transcriptional regulators discovered so far, focusing on their structural features, metal coordination modes and metal binding thermodynamics. Understanding these properties is essential to comprehend how these sensors correlate nickel availability to metal coordination and functional responses. A broad and comparative study, described here, reveals some general traits that characterize the binding stoichiometry and Ni(II) affinity of these metallo-sensors.

  10. Consensus PP1 binding motifs regulate transcriptional corepression and alternative RNA splicing activities of the steroid receptor coregulators, p54nrb and PSF.

    Science.gov (United States)

    Liu, Liangliang; Xie, Ning; Rennie, Paul; Challis, John R G; Gleave, Martin; Lye, Stephen J; Dong, Xuesen

    2011-07-01

    Originally identified as essential pre-mRNA splicing factors, non-POU-domain-containing, octamer binding protein (p54nrb) and PTB-associated RNA splicing factor (PSF) are also steroid receptor corepressors. The mechanisms by which p54nrb and PSF regulate gene transcription remain unclear. Both p54nrb and PSF contain protein phosphatase 1 (PP1) consensus binding RVxF motifs, suggesting that PP1 may regulate phosphorylation status of p54nrb and PSF and thus their function in gene transcription. In this report, we demonstrated that PP1 forms a protein complex with both p54nrb and PSF. PP1 interacts directly with the RVxF motif only in p54nrb, but not in PSF. Association with PP1 results in dephosphorylation of both p54nrb and PSF in vivo and the loss of their transcriptional corepressor activities. Using the CD44 minigene as a reporter, we showed that PP1 regulates p54nrb and PSF alternative splicing activities that determine exon skipping vs. inclusion in the final mature RNA for translation. In addition, changes in transcriptional corepression and RNA splicing activities of p54nrb and PSF are correlated with alterations in protein interactions of p54nrb and PSF with transcriptional corepressors such as Sin3A and histone deacetylase 1, and RNA splicing factors such as U1A and U2AF. Furthermore, we demonstrated a novel function of the RVxF motif within PSF that enhances its corepression and RNA splicing activities independent of PP1. We conclude that the RVxF motifs play an important role in controlling the multifunctional properties of p54nrb and PSF in the regulation of gene transcription.

  11. Transcriptional Regulation of Telomerase Reverse Transcriptase (TERT) by MYC

    Science.gov (United States)

    Khattar, Ekta; Tergaonkar, Vinay

    2017-01-01

    Telomerase elongates telomeres and is crucial for maintaining genomic stability. While stem cells and cancer cells display high telomerase activity, normal somatic cells lack telomerase activity primarily due to transcriptional repression of telomerase reverse transcriptase (TERT), the catalytic component of telomerase. Transcription factor binding, chromatin status as well as epigenetic modifications at the TERT promoter regulates TERT transcription. Myc is an important transcriptional regulator of TERT that directly controls its expression by promoter binding and associating with other transcription factors. In this review, we discuss the current understanding of the molecular mechanisms behind regulation of TERT transcription by Myc. We also discuss future perspectives in investigating the regulation of Myc at TERT promoter during cancer development.

  12. The roles of the catalytic and noncatalytic activities of Rpd3L and Rpd3S in the regulation of gene transcription in yeast.

    Directory of Open Access Journals (Sweden)

    Daniella Yeheskely-Hayon

    Full Text Available In budding yeasts, the histone deacetylase Rpd3 resides in two different complexes called Rpd3L (large and Rpd3S (small that exert opposing effects on the transcription of meiosis-specific genes. By introducing mutations that disrupt the integrity and function of either Rpd3L or Rpd3S, we show here that Rpd3 function is determined by its association with either of these complexes. Specifically, the catalytic activity of Rpd3S activates the transcription of the two major positive regulators of meiosis, IME1 and IME2, under all growth conditions and activates the transcription of NDT80 only during vegetative growth. In contrast, the effects of Rpd3L depends on nutrients; it represses or activates transcription in the presence or absence of a nitrogen source, respectively. Further, we show that transcriptional activation does not correlate with histone H4 deacetylation, suggesting an effect on a nonhistone protein. Comparison of rpd3-null and catalytic-site point mutants revealed an inhibitory activity that is independent of either the catalytic activity of Rpd3 or the integrity of Rpd3L and Rpd3S.

  13. Signal transducer and activator of transcription 5a inhibited by pimozide may regulate survival of goat mammary gland epithelial cells by regulating parathyroid hormone-related protein.

    Science.gov (United States)

    Li, Hui; Zheng, Huiling; Sun, Yongsen; Yu, Qian; Li, Lihui

    2014-11-10

    The signal transducer and activator of transcription 5a (Stat5a) modulates genes involved in proliferation and survival and plays pivotal roles in regulating the function of the mammary gland during pregnancy, lactation, and involution. However, there is little information about the effects of Stat5a on apoptosis of goat mammary gland epithelial cells (GMECs). In addition, parathyroid hormone-related protein (PTHrP) is a key regulator in cellular calcium transport, mammary gland development and breast tumor biology. This study aimed to explore the interaction of Stat5a and PTHrP in GMEC apoptosis. Quantitative real time PCR (qRT-PCR) suggested that Stat5a was predominantly expressed in the mammary gland, lung, liver and spleen of goats. Treating the GMECs with pimozide, an inhibitor of Stat5a that decreases Stat5a tyrosine phosphorylation, increased PTHrP levels in GMECs in a dose-dependent manner and simultaneously promoted apoptosis of the GMECs. We also demonstrated that PTHrP inhibition induced GMEC apoptosis and restrained cell proliferation. In contrast, PTHrP overexpression protected GMECs from pimozide- and calcium-induced apoptosis, and promoted cell proliferation. Furthermore, pimozide and CaCl2 downregulated the antiapoptotic protein Bcl-2 mRNA expression, respectively, and these effects were protected by PTHrP overexpression. Interestingly, we also found that Stat5a suppressed the expression of matrix metalloproteinase 9 (MMP-9) which can induce goat mammary epithelial cell migration, but PTHrP increased MMP-9 mRNA level. Thus, Stat5a may regulate GMEC survival by regulating the expression of PTHrP. Copyright © 2014. Published by Elsevier B.V.

  14. Transcriptional regulation of topology modulators and transcription regulators of Mycobacterium tuberculosis.

    Science.gov (United States)

    Ghosh, Soumitra; Padmanabhan, Bhavna; Godbole, Adwait Anand; Tare, Priyanka; Ahmed, Wareed; Vasu, Kommireddy; China, Arnab; Kumar, Rupesh; Mitra, Anirban; Nagaraja, Valakunja

    2016-07-01

    Mycobacterium tuberculosis (Mtb) is a formidable pathogen which has the ability to survive the hostile environment of the host by evading the host defense system. The re-configuration of its transcriptional and metabolic process allows the pathogen to confront the adverse environment within the host macrophages. The factors that assist the transcription and modulate the DNA topology would have to play a key role in the regulation of global gene expression of the organism. How transcription of these essential housekeeping genes alters in response to growth conditions and environmental stress has not been addressed together in a set of experimental conditions in Mtb. Now, we have mapped the transcription start sites (TSS) and promoters of several genes that play a central role in the regulation of DNA topology and transcription in Mtb. Using in vivo reporter assays, we validated the activity of the identified promoter elements in different growth conditions. The variation in transcript abundance of these essential genes was also analyzed in growth phase-dependent manner. These data provide the first glimpse into the specific adaptive changes in the expression of genes involved in transcription and DNA topology modulation in Mtb.

  15. Downregulated long non-coding RNA MEG3 in breast cancer regulates proliferation, migration and invasion by depending on p53’s transcriptional activity

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Lin [West Biostatistics and Cost-effectiveness Research Center, Medical Insurance Office, West China Hospital of Sichuan University, 610041, Sichuan (China); Li, Yu [Department of Anesthesiology, West China Hospital, Sichuan University, 610041, Sichuan (China); Yang, Bangxiang, E-mail: b19933009@qq.coom [Department of Pain Management, West China Hospital of Sichuan University, 610041, Sichuan (China)

    2016-09-09

    Long non-coding RNAs (lncRNAs) was found to play critical roles in tumorigenesis, hence, screen of tumor-related lncRNAs, identification of their biological roles is important for understanding the processes of tumorigenesis. In this study, we identified the expressing difference of several tumor-related lncRNAs in breast cancer samples and found that, MEG3, which is downregulated in non-small cell lung cancer (NSCLC) tumor tissues, is also downregulated in breast cancer samples compared with adjacent tissues. For figuring out the effect of MEG3 in breast cancer cells MCF7 and MB231, we overexpressed MEG3 in these cells, and found that it resulted the inhibition of proliferation, colony formation, migration and invasion capacities by enhancing p53’s transcriptional activity on its target genes, including p21, Maspin and KAI1. MEG3 presented similar effects in MB157, which is a p53-null breast cancer cell line, when functional p53 but not p53R273H mutant, which lacks transcriptional activity, was introduced. Surprisingly, overexpression of MEG3 activates p53’s transcriptional activity by decreasing MDM2’s transcription level, and thus stabilizes and accumulates P53. Taken together, our findings indicate that MEG3 is downregulated in breast cancer tissues and affects breast cancer cells’ malignant behaviors, which indicate MEG3 a potential therapeutic target for breast cancer. - Highlights: • MEG3 RNA is widely downregulated in breast tumor tissue. • MEG3 regulates P53 indirectly through transcriptional regulation of MDM2. • Under unstressed condition, MEG3-related P53 accumulation transcriptionally activates p53’s target genes. • MEG3 expression level tightly regulates proliferation, colony formation, migration and invasion in breast tumor cells.

  16. The transcription factor Gfi1 regulates G-CSF signaling and neutrophil development through the Ras activator RasGRP1

    Science.gov (United States)

    de la Luz Sierra, Maria; Sakakibara, Shuhei; Gasperini, Paola; Salvucci, Ombretta; Jiang, Kan; McCormick, Peter J.; Segarra, Marta; Stone, Jim; Maric, Dragan; Zhu, Jinfang; Qian, Xiaolan; Lowy, Douglas R.

    2010-01-01

    The transcription factor growth factor independence 1 (Gfi1) and the growth factor granulocyte colony-stimulating factor (G-CSF) are individually essential for neutrophil differentiation from myeloid progenitors. Here, we provide evidence that the functions of Gfi1 and G-CSF are linked in the regulation of granulopoiesis. We report that Gfi1 promotes the expression of Ras guanine nucleotide releasing protein 1 (RasGRP1), an exchange factor that activates Ras, and that RasGRP1 is required for G-CSF signaling through the Ras/mitogen–activated protein/extracellular signal-regulated kinase (MEK/Erk) pathway. Gfi1-null mice have reduced levels of RasGRP1 mRNA and protein in thymus, spleen, and bone marrow, and Gfi1 transduction in myeloid cells promotes RasGRP1 expression. When stimulated with G-CSF, Gfi1-null myeloid cells are selectively defective at activating Erk1/2, but not signal transducer and activator of transcription 1 (STAT1) or STAT3, and fail to differentiate into neutrophils. Expression of RasGRP1 in Gfi1-deficient cells rescues Erk1/2 activation by G-CSF and allows neutrophil maturation by G-CSF. These results uncover a previously unknown function of Gfi1 as a regulator of RasGRP1 and link Gfi1 transcriptional control to G-CSF signaling and regulation of granulopoiesis. PMID:20203268

  17. Transcriptional Regulation of Heart Development in Zebrafish

    Science.gov (United States)

    Lu, Fei; Langenbacher, Adam D.; Chen, Jau-Nian

    2016-01-01

    Cardiac transcription factors orchestrate the complex cellular and molecular events required to produce a functioning heart. Misregulation of the cardiac transcription program leads to embryonic developmental defects and is associated with human congenital heart diseases. Recent studies have expanded our understanding of the regulation of cardiac gene expression at an additional layer, involving the coordination of epigenetic and transcriptional regulators. In this review, we highlight and discuss discoveries made possible by the genetic and embryological tools available in the zebrafish model organism, with a focus on the novel functions of cardiac transcription factors and epigenetic and transcriptional regulatory proteins during cardiogenesis. PMID:27148546

  18. The Serratia LuxR family regulator CarR 39006 activates transcription independently of cognate quorum sensing signals.

    Science.gov (United States)

    Poulter, Simon; Carlton, Timothy M; Spring, David R; Salmond, George P C

    2011-05-01

    In Gram-negative bacteria, quorum sensing control of gene expression is mediated by transcription factors of the LuxR family, whose DNA-binding affinity is modulated by diffusible N-acyl homoserine lactone (AHL) signalling molecules. In Serratia sp. ATCC 39006 and the plant pathogen Erwinia carotovora ssp. carotovora (Ecc), the biosynthesis of the β-lactam antibiotic 1-carbapen-2-em-3-carboxylic acid (Car) is under quorum sensing control. This study has revealed that, uniquely, the LuxR family transcriptional activator CarR(39006) from Serratia 39006 has no detectable affinity for cognate AHL molecules. Furthermore, CarR(39006) was shown to be naturally competent to bind to its target promoter with high affinity, activate transcription and resist cellular proteolysis, and was unaffected by AHL signals. Experiments with chimeric proteins suggest that the C-terminal DNA-binding domain of CarR(39006) may be responsible for conferring AHL independence. In contrast, we show that the homologous CarR(Ecc) protein binds to its 3O-C6-HSL ligand with high affinity, and that the highly conserved Trp-44 residue is critical for this interaction. Unlike TraR from Agrobacterium tumefaciens, CarR(Ecc) is not directly protected from cellular proteolysis by AHL binding, but via AHL-induced DNA binding. At physiological protein concentrations, AHL binding induces CarR(Ecc) to bind to its target promoter with higher affinity and activate transcription. © 2011 Blackwell Publishing Ltd.

  19. HECT E3 Ubiquitin Ligase Itch Functions as a Novel Negative Regulator of Gli-Similar 3 (Glis3 Transcriptional Activity.

    Directory of Open Access Journals (Sweden)

    Gary T ZeRuth

    Full Text Available The transcription factor Gli-similar 3 (Glis3 plays a critical role in the generation of pancreatic ß cells and the regulation insulin gene transcription and has been implicated in the development of several pathologies, including type 1 and 2 diabetes and polycystic kidney disease. However, little is known about the proteins and posttranslational modifications that regulate or mediate Glis3 transcriptional activity. In this study, we identify by mass-spectrometry and yeast 2-hybrid analyses several proteins that interact with the N-terminal region of Glis3. These include the WW-domain-containing HECT E3 ubiquitin ligases, Itch, Smurf2, and Nedd4. The interaction between Glis3 and the HECT E3 ubiquitin ligases was verified by co-immunoprecipitation assays and mutation analysis. All three proteins interact through their WW-domains with a PPxY motif located in the Glis3 N-terminus. However, only Itch significantly contributed to Glis3 polyubiquitination and reduced Glis3 stability by enhancing its proteasomal degradation. Itch-mediated degradation of Glis3 required the PPxY motif-dependent interaction between Glis3 and the WW-domains of Itch as well as the presence of the Glis3 zinc finger domains. Transcription analyses demonstrated that Itch dramatically inhibited Glis3-mediated transactivation and endogenous Ins2 expression by increasing Glis3 protein turnover. Taken together, our study identifies Itch as a critical negative regulator of Glis3-mediated transcriptional activity. This regulation provides a novel mechanism to modulate Glis3-driven gene expression and suggests that it may play a role in a number of physiological processes controlled by Glis3, such as insulin transcription, as well as in Glis3-associated diseases.

  20. Nobiletin suppresses adipogenesis by regulating the expression of adipogenic transcription factors and the activation of AMP-activated protein kinase (AMPK).

    Science.gov (United States)

    Choi, Youngmin; Kim, Younghwa; Ham, Hyeonmi; Park, Yooheon; Jeong, Heon-Sang; Lee, Junsoo

    2011-12-28

    The objective of this study was to elucidate the effect of nobiletin (5,6,7,8,3',4'-hexamethoxyflavone) on adipogenesis in 3T3-L1 cells. To determine the effect of nobiletin on adipogenesis, preadipocyte differentiation was induced in the presence or absence of nobiletin (10-100 μM) for 4 days. The results revealed that nobiletin markedly inhibited lipid accumulation and glycerol-3-phosphate dehydrogenase (GPDH) activity and blocked the expression of adipogenic transcription factors, including peroxisome proliferator-activated receptors (PPARγ) and CCAAT/enhancer binding proteins (C/EBPα). Moreover, nobiletin significantly increased AMP-activated protein kinase (AMPK), a major regulator of cellular energy balance, phosphorylation, and intracellular reactive oxygen species (ROS) generation. This study also investigated the involvement of AMPK in the expression of a major transcription factor, PPARγ. It was found that pretreatment with compound C, a cell permeable inhibitor of AMPK, abolished the inhibitory effects of nobiletin on PPARγ expression. The results suggest that nobiletin exerts antiadipogenic effects through modulation of the PPARγ and AMPK signaling pathway and, therefore, may be a promising antiobesity agent.

  1. Identification of a Small Molecule Activator for AphB, a LysR-Type Virulence Transcriptional Regulator in Vibrio cholerae.

    Science.gov (United States)

    Privett, Britney R; Pellegrini, Maria; Kovacikova, Gabriela; Taylor, Ronald K; Skorupski, Karen; Mierke, Dale; Kull, F Jon

    2017-07-25

    AphB is a LysR-type transcriptional regulator (LTTR) that cooperates with a second transcriptional activator, AphA, at the tcpPH promoter to initiate expression of the virulence cascade in Vibrio cholerae. Because it is not yet known whether AphB responds to a natural ligand in V. cholerae that influences its ability to activate transcription, we used a computational approach to identify small molecules that influence its activity. In silico docking was used to identify potential ligands for AphB, and saturation transfer difference nuclear magnetic resonance was subsequently employed to access the validity of promising targets. We identified a small molecule, BP-15, that specifically binds the C-terminal regulatory domain of AphB and increases its activity. Interestingly, molecular docking predicts that BP-15 does not bind in the putative primary effector-binding pocket located at the interface of RD-I and RD-II as in other LTTRs, but rather at the dimerization interface. The information gained in this study helps us to further understand the mechanism by which transcriptional activation by AphB is regulated by suggesting that AphB has a secondary ligand binding site, as observed in other LTTRs. This study also lays the groundwork for the future design of inhibitory molecules to block the V. cholerae virulence cascade, thereby preventing the devastating symptoms of cholera infection.

  2. Involvement of V-Ets erythroblastosis virus E26 oncogene homolog 2 in regulation of transcription activity of MDR1 gene

    Institute of Scientific and Technical Information of China (English)

    Jian Wang; Xiaoqing Zeng; Tiancheng Luo; Wei Jin; Shiyao Chen

    2012-01-01

    Over-expression of MDR1 confers multidrug resistance (MDR) in cancers and remains a major cause for the failure of chemotherapy.In the present study,we found that V-Ets erythroblastosis virus E26 oncogene homolog 2(ETS2) could activate MDR1 transcription and P-glyco-protein (P-gp) expression in SGC7901 cells.Knockdown of ETS2 attenuated MDR1 transcription and P-gp expression,and increased the sensitivity of MDR cancer cells to cytotoxic drugs that were transported by P-gp in SGC7901/VCR cells.ETS2 could bind to the ETS2 sites on the MDR1 promoter and activate its transcription.The regulation of MDR1 expression by ETS2 may provide potential ways to overcome MDR in cancer treatment.

  3. Core promoter-specific gene regulation: TATA box selectivity and Initiator-dependent bi-directionality of serum response factor-activated transcription.

    Science.gov (United States)

    Xu, Muyu; Gonzalez-Hurtado, Elsie; Martinez, Ernest

    2016-04-01

    Gene-specific activation by enhancers involves their communication with the basal RNA polymerase II transcription machinery at the core promoter. Core promoters are diverse and may contain a variety of sequence elements such as the TATA box, the Initiator (INR), and the downstream promoter element (DPE) recognized, respectively, by the TATA-binding protein (TBP) and TBP-associated factors of the TFIID complex. Core promoter elements contribute to the gene selectivity of enhancers, and INR/DPE-specific enhancers and activators have been identified. Here, we identify a TATA box-selective activating sequence upstream of the human β-actin (ACTB) gene that mediates serum response factor (SRF)-induced transcription from TATA-dependent but not INR-dependent promoters and requires the TATA-binding/bending activity of TBP, which is otherwise dispensable for transcription from a TATA-less promoter. The SRF-dependent ACTB sequence is stereospecific on TATA promoters but activates in an orientation-independent manner a composite TATA/INR-containing promoter. More generally, we show that SRF-regulated genes of the actin/cytoskeleton/contractile family tend to have a TATA box. These results suggest distinct TATA-dependent and INR-dependent mechanisms of TFIID-mediated transcription in mammalian cells that are compatible with only certain stereospecific combinations of activators, and that a TBP-TATA binding mechanism is important for SRF activation of the actin/cytoskeleton-related gene family.

  4. The transcriptional activator GAL4-VP16 regulates the intra-molecular interactions of the TATA-binding protein

    Indian Academy of Sciences (India)

    Anurag Kumar Mishra; Perumal Vanathi; Purnima Bhargava

    2003-06-01

    Binding characteristics of yeast TATA-binding protein (yTBP) over five oligomers having different TATA variants and lacking a UASGAL, showed that TATA-binding protein (TBP)-TATA complex gets stabilized in the presence of the acidic activator GAL4-VP16. Activator also greatly suppressed the non-specific TBP-DNA complex formation. The effects were more pronounced over weaker TATA boxes. Activator also reduced the TBP dimer levels both in vitro and in vivo, suggesting the dimer may be a direct target of transcriptional activators. The transcriptional activator facilitated the dimer to monomer transition and activated monomers further to help TBP bind even the weaker TATA boxes stably. The overall stimulatory effect of the GAL4-VP16 on the TBP-TATA complex formation resembles the known effects of removal of the N-terminus of TBP on its activity, suggesting that the activator directly targets the N-terminus of TBP and facilitates its binding to the TATA box.

  5. Methylation of an intragenic alternative promoter regulates transcription of GARP.

    Science.gov (United States)

    Haupt, Sonja; Söntgerath, Viktoria Sophie Apollonia; Leipe, Jan; Schulze-Koops, Hendrik; Skapenko, Alla

    2016-02-01

    Alternative promoter usage has been proposed as a mechanism regulating transcriptional and translational diversity in highly elaborated systems like the immune system in humans. Here, we report that transcription of human glycoprotein A repetitions predominant (GARP) in regulatory CD4 T cells (Tregs) is tightly regulated by two alternative promoters. An intragenic promoter contains several CpGs and acts as a weak promoter that is demethylated and initiates transcription Treg-specifically. The strong up-stream promoter containing a CpG-island is, in contrast, fully demethylated throughout tissues. Transcriptional activity of the strong promoter was surprisingly down-regulated upon demethylation of the weak promoter. This demethylation-induced transcriptional attenuation regulated the magnitude of GARP expression and correlated with disease activity in rheumatoid arthritis. Treg-specific GARP transcription was initiated by synergistic interaction of forkhead box protein 3 (Foxp3) with nuclear factor of activated T cells (NFAT) and was underpinned by permissive chromatin remodeling caused by release of the H3K4 demethylase, PLU-1. Our findings describe a novel function of alternative promoters in regulating the extent of transcription. Moreover, since GARP functions as a transporter of transforming growth factor β (TGFβ), a cytokine with broad pleiotropic traits, GARP transcriptional attenuation by alternative promoters might provide a mechanism regulating peripheral TGFβ to avoid unwanted harmful effects.

  6. DNA-binding specificity, transcriptional activation potential, and the rin mutation effect for the tomato fruit-ripening regulator RIN.

    Science.gov (United States)

    Ito, Yasuhiro; Kitagawa, Mamiko; Ihashi, Nao; Yabe, Kimiko; Kimbara, Junji; Yasuda, Junichi; Ito, Hirotaka; Inakuma, Takahiro; Hiroi, Seiji; Kasumi, Takafumi

    2008-07-01

    The RIN gene encodes a putative MADS box transcription factor that controls tomato fruit ripening, and its ripening inhibitor (rin) mutation yields non-ripening fruit. In this study, the molecular properties of RIN and the rin mutant protein were clarified. The results revealed that the RIN protein accumulates in ripening fruit specifically and is localized in the nucleus of the cell. In vitro studies revealed that RIN forms a stable homodimer that binds to MADS domain-specific DNA sites. Analysis of binding site selection experiments revealed that the consensus binding sites of RIN highly resemble those of the SEPALLATA (SEP) proteins, which are Arabidopsis MADS box proteins that control the identity of floral organs. RIN exhibited a transcription-activating function similar to that exhibited by the SEP proteins. These results indicate that RIN exhibits similar molecular functions to SEP proteins although they play distinctly different biological roles. In vivo assays revealed that RIN binds to the cis-element of LeACS2. Our results also revealed that the rin mutant protein accumulates in the mutant fruit and exhibits a DNA-binding activity similar to that exhibited by the wild-type protein, but has lost its transcription-activating function, which in turn would inhibit ripening in mutant fruit.

  7. Histone variants in plant transcriptional regulation.

    Science.gov (United States)

    Jiang, Danhua; Berger, Frédéric

    2017-01-01

    Chromatin based organization of eukaryotic genome plays a profound role in regulating gene transcription. Nucleosomes form the basic subunits of chromatin by packaging DNA with histone proteins, impeding the access of DNA to transcription factors and RNA polymerases. Exchange of histone variants in nucleosomes alters the properties of nucleosomes and thus modulates DNA exposure during transcriptional regulation. Growing evidence indicates the important function of histone variants in programming transcription during developmental transitions and stress response. Here we review how histone variants and their deposition machineries regulate the nucleosome stability and dynamics, and discuss the link between histone variants and transcriptional regulation in plants. This article is part of a Special Issue entitled: Plant Gene Regulatory Mechanisms and Networks, edited by Dr. Erich Grotewold and Dr. Nathan Springer.

  8. Distal Interleukin-1β (IL-1β) Response Element of Human Matrix Metalloproteinase-13 (MMP-13) Binds Activator Protein 1 (AP-1) Transcription Factors and Regulates Gene Expression*

    Science.gov (United States)

    Schmucker, Adam C.; Wright, Jason B.; Cole, Michael D.; Brinckerhoff, Constance E.

    2012-01-01

    The collagenase matrix metalloproteinase-13 (MMP-13) plays an important role in the destruction of cartilage in arthritic joints. MMP-13 expression is strongly up-regulated in arthritis, largely because of stimulation by inflammatory cytokines such as IL-1β. Treatment of chondrocytes with IL-1β induces transcription of MMP-13 in vitro. IL-1β signaling converges upon the activator protein-1 transcription factors, which have been shown to be required for IL-1β-induced MMP-13 gene expression. Using chromatin immunoprecipitation (ChIP), we detected activator protein-1 binding within an evolutionarily conserved DNA sequence ∼20 kb 5′ relative to the MMP-13 transcription start site (TSS). Also using ChIP, we detected histone modifications and binding of RNA polymerase II within this conserved region, all of which are consistent with transcriptional activation. Chromosome conformation capture indicates that chromosome looping brings this region in close proximity with the MMP-13 TSS. Finally, a luciferase reporter construct driven by a component of the conserved region demonstrated an expression pattern similar to that of endogenous MMP-13. These data suggest that a conserved region at 20 kb upstream from the MMP-13 TSS includes a distal transcriptional response element of MMP-13, which contributes to MMP-13 gene expression. PMID:22102411

  9. Phosphorylation of the transcription activator CLOCK regulates progression through a ∼ 24-h feedback loop to influence the circadian period in Drosophila.

    Science.gov (United States)

    Mahesh, Guruswamy; Jeong, EunHee; Ng, Fanny S; Liu, Yixiao; Gunawardhana, Kushan; Houl, Jerry H; Yildirim, Evrim; Amunugama, Ravi; Jones, Richard; Allen, David L; Edery, Isaac; Kim, Eun Young; Hardin, Paul E

    2014-07-11

    Circadian (≅ 24 h) clocks control daily rhythms in metabolism, physiology, and behavior in animals, plants, and microbes. In Drosophila, these clocks keep circadian time via transcriptional feedback loops in which clock-cycle (CLK-CYC) initiates transcription of period (per) and timeless (tim), accumulating levels of PER and TIM proteins feed back to inhibit CLK-CYC, and degradation of PER and TIM allows CLK-CYC to initiate the next cycle of transcription. The timing of key events in this feedback loop are controlled by, or coincide with, rhythms in PER and CLK phosphorylation, where PER and CLK phosphorylation is high during transcriptional repression. PER phosphorylation at specific sites controls its subcellular localization, activity, and stability, but comparatively little is known about the identity and function of CLK phosphorylation sites. Here we identify eight CLK phosphorylation sites via mass spectrometry and determine how phosphorylation at these sites impacts behavioral and molecular rhythms by transgenic rescue of a new Clk null mutant. Eliminating phosphorylation at four of these sites accelerates the feedback loop to shorten the circadian period, whereas loss of CLK phosphorylation at serine 859 increases CLK activity, thereby increasing PER levels and accelerating transcriptional repression. These results demonstrate that CLK phosphorylation influences the circadian period by regulating CLK activity and progression through the feedback loop. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. Brain-specific transcriptional regulator T-brain-1 controls brain wiring and neuronal activity in autism spectrum disorders

    Directory of Open Access Journals (Sweden)

    Tzyy-Nan eHuang

    2015-11-01

    Full Text Available T-brain-1 (TBR1 is a brain-specific T-box transcription factor. In 1995, Tbr1 was first identified from a subtractive hybridization that compared mouse embryonic and adult telencephalons. Previous studies of Tbr1–/– mice have indicated critical roles for TBR1 in the development of the cerebral cortex, amygdala and olfactory bulb. Neuronal migration and axonal projection are two important developmental features controlled by TBR1. Recently, recurrent de novo disruptive mutations in the TBR1 gene have been found in patients with autism spectrum disorders (ASDs. Human genetic studies have identified TBR1 as a high-confidence risk factor for ASDs. Because only one allele of the TBR1 gene is mutated in these patients, Tbr1+/– mice serve as a good genetic mouse model to explore the mechanism by which de novo TBR1 mutation leads to ASDs. Although neuronal migration and axonal projection defects of cerebral cortex are the most prominent phenotypes in Tbr1–/– mice, these features are not found in Tbr1+/– mice. Instead, inter- and intra-amygdalar axonal projections and NMDAR expression and activity in amygdala are particularly susceptible to Tbr1 haploinsufficiency. The studies indicated that both abnormal brain wiring (abnormal amygdalar connections and excitation/inhibition imbalance (NMDAR hypoactivity, two prominent models for ASD etiology, are present in Tbr1+/– mice. Moreover, calcium/calmodulin-dependent serine protein kinase (CASK was found to interact with TBR1. The CASK-TBR1 complex had been shown to directly bind the promoter of the Grin2b gene, which is also known as Nmdar2b, and upregulate Grin2b expression. This molecular function of TBR1 provides an explanation for NMDAR hypoactivity in Tbr1+/– mice. In addition to Grin2b, cell adhesion molecules-including Ntng1, Cdh8 and Cntn2-are also regulated by TBR1 to control axonal projections of amygdala. Taken together, the studies of Tbr1 provide an integrated picture of ASD

  11. Catabolite control protein E (CcpE) is a LysR-type transcriptional regulator of tricarboxylic acid cycle activity in Staphylococcus aureus.

    Science.gov (United States)

    Hartmann, Torsten; Zhang, Bo; Baronian, Grégory; Schulthess, Bettina; Homerova, Dagmar; Grubmüller, Stephanie; Kutzner, Erika; Gaupp, Rosmarie; Bertram, Ralph; Powers, Robert; Eisenreich, Wolfgang; Kormanec, Jan; Herrmann, Mathias; Molle, Virginie; Somerville, Greg A; Bischoff, Markus

    2013-12-13

    The tricarboxylic acid cycle (TCA cycle) is a central metabolic pathway that provides energy, reducing potential, and biosynthetic intermediates. In Staphylococcus aureus, TCA cycle activity is controlled by several regulators (e.g. CcpA, CodY, and RpiRc) in response to the availability of sugars, amino acids, and environmental stress. Developing a bioinformatic search for additional carbon catabolite-responsive regulators in S. aureus, we identified a LysR-type regulator, catabolite control protein E (CcpE), with homology to the Bacillus subtilis CcpC regulator. Inactivation of ccpE in S. aureus strain Newman revealed that CcpE is a positive transcriptional effector of the first two enzymes of the TCA cycle, aconitase (citB) and to a lesser extent citrate synthase (citZ). Consistent with the transcriptional data, aconitase activity dramatically decreased in the ccpE mutant relative to the wild-type strain. The effect of ccpE inactivation on citB transcription and the lesser effect on citZ transcription were also reflected in electrophoretic mobility shift assays where CcpE bound to the citB promoter but not the citZ promoter. Metabolomic studies showed that inactivation of ccpE resulted in increased intracellular concentrations of acetate, citrate, lactate, and alanine, consistent with a redirection of carbon away from the TCA cycle. Taken together, our data suggest that CcpE is a major direct positive regulator of the TCA cycle gene citB.

  12. The schizophrenia- and autism-associated gene, transcription factor 4 regulates the columnar distribution of layer 2/3 prefrontal pyramidal neurons in an activity-dependent manner.

    Science.gov (United States)

    Page, S C; Hamersky, G R; Gallo, R A; Rannals, M D; Calcaterra, N E; Campbell, M N; Mayfield, B; Briley, A; Phan, B N; Jaffe, A E; Maher, B J

    2017-03-14

    Disruption of the laminar and columnar organization of the brain is implicated in several psychiatric disorders. Here, we show in utero gain-of-function of the psychiatric risk gene transcription factor 4 (TCF4) severely disrupts the columnar organization of medial prefrontal cortex (mPFC) in a transcription- and activity-dependent manner. This morphological phenotype was rescued by co-expression of TCF4 plus calmodulin in a calcium-dependent manner and by dampening neuronal excitability through co-expression of an inwardly rectifying potassium channel (Kir2.1). For we believe the first time, we show that N-methyl-d-aspartate (NMDA) receptor-dependent Ca(2+) transients are instructive to minicolumn organization because Crispr/Cas9-mediated mutation of NMDA receptors rescued TCF4-dependent morphological phenotypes. Furthermore, we demonstrate that the transcriptional regulation by the psychiatric risk gene TCF4 enhances NMDA receptor-dependent early network oscillations. Our novel findings indicate that TCF4-dependent transcription directs the proper formation of prefrontal cortical minicolumns by regulating the expression of genes involved in early spontaneous neuronal activity, and thus our results provides insights into potential pathophysiological mechanisms of TCF4-associated psychiatric disorders.Molecular Psychiatry advance online publication, 14 March 2017; doi:10.1038/mp.2017.37.

  13. Transcriptional Regulation of Pyruvate Dehydrogenase Kinase

    Directory of Open Access Journals (Sweden)

    Ji Yun Jeong

    2012-10-01

    Full Text Available The pyruvate dehydrogenase complex (PDC activity is crucial to maintains blood glucose and ATP levels, which largely depends on the phosphorylation status by pyruvate dehydrogenase kinase (PDK isoenzymes. Although it has been reported that PDC is phosphorylated and inactivated by PDK2 and PDK4 in metabolically active tissues including liver, skeletal muscle, heart, and kidney during starvation and diabetes, the precise mechanisms by which expression of PDK2 and PDK4 are transcriptionally regulated still remains unclear. Insulin represses the expression of PDK2 and PDK4 via phosphorylation of FOXO through PI3K/Akt signaling pathway. Several nuclear hormone receptors activated due to fasting or increased fat supply, including peroxisome proliferator-activated receptors, glucocorticoid receptors, estrogen-related receptors, and thyroid hormone receptors, also participate in the up-regulation of PDK2 and PDK4; however, the endogenous ligands that bind those nuclear receptors have not been identified. It has been recently suggested that growth hormone, adiponectin, epinephrine, and rosiglitazone also control the expression of PDK4 in tissue-specific manners. In this review, we discuss several factors involved in the expressional regulation of PDK2 and PDK4, and introduce current studies aimed at providing a better understanding of the molecular mechanisms that underlie the development of metabolic diseases such as diabetes.

  14. APE1/Ref-1 regulates STAT3 transcriptional activity and APE1/Ref-1-STAT3 dual-targeting effectively inhibits pancreatic cancer cell survival.

    Science.gov (United States)

    Cardoso, Angelo A; Jiang, Yanlin; Luo, Meihua; Reed, April M; Shahda, Safi; He, Ying; Maitra, Anirban; Kelley, Mark R; Fishel, Melissa L

    2012-01-01

    Pancreatic cancer is a largely incurable disease, and increasing evidence supports strategies targeting multiple molecular mediators of critical functions of pancreatic ductal adenocarcinoma cells. Intracellular redox state modulates the activity of various signal transduction pathways and biological processes, including cell survival, drug resistance and responsiveness to microenvironmental factors. Recently, it has been shown that the transcription factor STAT3 is under redox control, but the mechanisms involved in its regulation are unknown. Here, we demonstrate for the first time that STAT3 DNA binding and transcriptional activity is directly regulated by the redox function of the APE1/Ref-1 endonuclease, using overexpression and redox-specific mutational strategies, and gene knockdown. Also, pharmacological blockade of APE1/Ref-1 by the redox-selective inhibitor E3330 abrogates STAT3 DNA binding. Since APE1/Ref-1 also exerts redox control on other cancer-associated transcription factors, we assessed the impact of dual-targeting of STAT3 signaling and APE1/Ref-1 redox on pancreatic cancer cell functions. We observed that disruption of APE1/Ref-1 redox activity synergizes with STAT3 blockade to potently inhibit the proliferation and viability of human PDAC cells. Mechanistically, we show that STAT3-APE1/Ref-1 dual targeting promotes marked tumor cell apoptosis, with engagement of caspase-3 signaling, which are significantly increased in comparison to the effects triggered by single target blockade. Also, we show that STAT3-APE1/Ref-1 dual blockade results in significant inhibition of tumor cell migration. Overall, this work demonstrates that the transcriptional activity of STAT3 is directly regulated by the redox function of APE1/Ref-1, and that concurrent blockade of STAT3 and APE1/Ref-1 redox synergize effectively inhibit critical PDAC cell functions.

  15. Transcriptional regulation of mononuclear phagocyte development

    Directory of Open Access Journals (Sweden)

    Roxane eTussiwand

    2015-10-01

    Full Text Available IntroductionThe mononuclear-phagocyte system (MPS, which comprises dendritic cells (DCs, macrophages and monocytes, is a heterogeneous group of myeloid cells. The complexity of the MPS is equally reflected by the plasticity in function and phenotype that characterizes each subset depending on their location and activation state. Specialized subsets of Mononuclear Phagocytes (MP reside in defined anatomical locations, are critical for the homeostatic maintenance of tissues, and provide the link between innate and adaptive immune responses during infections. The ability of MP to maintain or to induce the correct tolerogenic or inflammatory milieu also resides in their complex subset specialization. Such subset heterogeneity is obtained through lineage diversification and specification, which is controlled by defined transcriptional networks and programs. Understanding the MP biology means to define their transcriptional signature, which is required during lineage commitment, and which characterizes each subset’s features. This review will focus on the transcriptional regulation of the MPS; in particular what determines lineage commitment and functional identity; we will emphasizes recent advances in the field of single cell analysis and highlight unresolved questions in the field.

  16. Regulating transcription traffic around DSBs.

    Science.gov (United States)

    Plosky, Brian S

    2015-05-07

    If a double-strand break (DSB) occurs and either a DNA polymerase or RNA polymerase is coming along, how do we save the train? In this issue of Molecular Cell, Ui et al. (2015) describe a connection between an elongation factor and a repressive complex to prevent transcription in proximity to a DSB.

  17. Transcriptional regulation of IL-2 in health and autoimmunity

    Science.gov (United States)

    Crispín, José C.; Tsokos, George C.

    2009-01-01

    The regulation of IL-2 production is central to our understanding of the immune system. Key during T cell activation, it also plays an essential role in the regulation of the immune response. This review discusses the function of recently described factors that modulate transcription and chromatin remodeling at the IL2 promoter. Also, it addresses the role of FoxP3 as a transcriptional regulator in conventional T cells and regulatory T cells, and the mechanisms whereby CD28 stabilizes IL2 transcription and translation. Finally, the alterations that prevent T cells from SLE patients from producing normal amounts of IL-2 upon stimulation are described. PMID:18723131

  18. Structural basis of transcription activation.

    Science.gov (United States)

    Feng, Yu; Zhang, Yu; Ebright, Richard H

    2016-06-10

    Class II transcription activators function by binding to a DNA site overlapping a core promoter and stimulating isomerization of an initial RNA polymerase (RNAP)-promoter closed complex into a catalytically competent RNAP-promoter open complex. Here, we report a 4.4 angstrom crystal structure of an intact bacterial class II transcription activation complex. The structure comprises Thermus thermophilus transcription activator protein TTHB099 (TAP) [homolog of Escherichia coli catabolite activator protein (CAP)], T. thermophilus RNAP σ(A) holoenzyme, a class II TAP-dependent promoter, and a ribotetranucleotide primer. The structure reveals the interactions between RNAP holoenzyme and DNA responsible for transcription initiation and reveals the interactions between TAP and RNAP holoenzyme responsible for transcription activation. The structure indicates that TAP stimulates isomerization through simple, adhesive, stabilizing protein-protein interactions with RNAP holoenzyme. Copyright © 2016, American Association for the Advancement of Science.

  19. Dual-level regulation of ACC synthase activity by MPK3/MPK6 cascade and its downstream WRKY transcription factor during ethylene induction in Arabidopsis.

    Science.gov (United States)

    Li, Guojing; Meng, Xiangzong; Wang, Ruigang; Mao, Guohong; Han, Ling; Liu, Yidong; Zhang, Shuqun

    2012-06-01

    Plants under pathogen attack produce high levels of ethylene, which plays important roles in plant immunity. Previously, we reported the involvement of ACS2 and ACS6, two Type I ACS isoforms, in Botrytis cinerea-induced ethylene biosynthesis and their regulation at the protein stability level by MPK3 and MPK6, two Arabidopsis pathogen-responsive mitogen-activated protein kinases (MAPKs). The residual ethylene induction in the acs2/acs6 double mutant suggests the involvement of additional ACS isoforms. It is also known that a subset of ACS genes, including ACS6, is transcriptionally induced in plants under stress or pathogen attack. However, the importance of ACS gene activation and the regulatory mechanism(s) are not clear. In this report, we demonstrate using genetic analysis that ACS7 and ACS11, two Type III ACS isoforms, and ACS8, a Type II ACS isoform, also contribute to the B. cinerea-induced ethylene production. In addition to post-translational regulation, transcriptional activation of the ACS genes also plays a critical role in sustaining high levels of ethylene induction. Interestingly, MPK3 and MPK6 not only control the stability of ACS2 and ACS6 proteins via direct protein phosphorylation but also regulate the expression of ACS2 and ACS6 genes. WRKY33, another MPK3/MPK6 substrate, is involved in the MPK3/MPK6-induced ACS2/ACS6 gene expression based on genetic analyses. Furthermore, chromatin-immunoprecipitation assay reveals the direct binding of WRKY33 to the W-boxes in the promoters of ACS2 and ACS6 genes in vivo, suggesting that WRKY33 is directly involved in the activation of ACS2 and ACS6 expression downstream of MPK3/MPK6 cascade in response to pathogen invasion. Regulation of ACS activity by MPK3/MPK6 at both transcriptional and protein stability levels plays a key role in determining the kinetics and magnitude of ethylene induction.

  20. A novel dual kinase function of the RET proto-oncogene negatively regulates activating transcription factor 4-mediated apoptosis.

    Science.gov (United States)

    Bagheri-Yarmand, Rozita; Sinha, Krishna M; Gururaj, Anupama E; Ahmed, Zamal; Rizvi, Yasmeen Q; Huang, Su-Chen; Ladbury, John E; Bogler, Oliver; Williams, Michelle D; Cote, Gilbert J; Gagel, Robert F

    2015-05-01

    The RET proto-oncogene, a tyrosine kinase receptor, is widely known for its essential role in cell survival. Germ line missense mutations, which give rise to constitutively active oncogenic RET, were found to cause multiple endocrine neoplasia type 2, a dominant inherited cancer syndrome that affects neuroendocrine organs. However, the mechanisms by which RET promotes cell survival and prevents cell death remain elusive. We demonstrate that in addition to cytoplasmic localization, RET is localized in the nucleus and functions as a tyrosine-threonine dual specificity kinase. Knockdown of RET by shRNA in medullary thyroid cancer-derived cells stimulated expression of activating transcription factor 4 (ATF4), a master transcription factor for stress-induced apoptosis, through activation of its target proapoptotic genes NOXA and PUMA. RET knockdown also increased sensitivity to cisplatin-induced apoptosis. We observed that RET physically interacted with and phosphorylated ATF4 at tyrosine and threonine residues. Indeed, RET kinase activity was required to inhibit the ATF4-dependent activation of the NOXA gene because the site-specific substitution mutations that block threonine phosphorylation increased ATF4 stability and activated its targets NOXA and PUMA. Moreover, chromatin immunoprecipitation assays revealed that ATF4 occupancy increased at the NOXA promoter in TT cells treated with tyrosine kinase inhibitors or the ATF4 inducer eeyarestatin as well as in RET-depleted TT cells. Together these findings reveal RET as a novel dual kinase with nuclear localization and provide mechanisms by which RET represses the proapoptotic genes through direct interaction with and phosphorylation-dependent inactivation of ATF4 during the pathogenesis of medullary thyroid cancer.

  1. PTS regulation domain-containing transcriptional activator CelR and sigma factor σ(54) control cellobiose utilization in Clostridium acetobutylicum.

    Science.gov (United States)

    Nie, Xiaoqun; Yang, Bin; Zhang, Lei; Gu, Yang; Yang, Sheng; Jiang, Weihong; Yang, Chen

    2016-04-01

    The phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) regulation domain (PRD)-containing enhancer binding proteins (EBPs) are an important class of σ(54) -interacting transcriptional activators. Although PRD-containing EBPs are present in many Firmicutes, most of their regulatory functions remain unclear. In this study, the transcriptional regulons of about 50 PRD-containing EBPs in diverse Firmicutes species are reconstructed by using a comparative genomic approach, which contain the genes associated with utilization of β-glucosides, fructose/levan, mannose/glucose, pentitols, and glucosamine/fructosamine. We then present experimental evidence that the cel operon involved in cellobiose utilization is directly regulated by CelR and σ(54) (SigL) in Clostridium acetobutylicum. The predicted three CelR-binding sites and σ(54) promoter elements upstream of the cel operon are verified by in vitro binding assays. We show that CelR has an ATPase activity, which is strongly stimulated by the presence of DNA containing the CelR-binding sites. Moreover, mutations in any one of the three CelR-binding sites significantly decreased the cel promoter activity probably due to the need for all three DNA sites for maximal ATPase activity of CelR. It is suggested that CelR is regulated by PTS-mediated phosphorylation at His-551 and His-829, which exerts a positive effect and an inhibitory effect, respectively, on the CelR activity. © 2015 John Wiley & Sons Ltd.

  2. Crystal structure and DNA binding activity of a PadR family transcription regulator from hypervirulent Clostridium difficile R20291.

    Science.gov (United States)

    Isom, Catherine E; Menon, Smita K; Thomas, Leonard M; West, Ann H; Richter-Addo, George B; Karr, Elizabeth A

    2016-10-04

    Clostridium difficile is a spore-forming obligate anaerobe that can remain viable for extended periods, even in the presence of antibiotics, which contributes to the persistence of this bacterium as a human pathogen during host-to-host transmission and in hospital environments. We examined the structure and function of a gene product with the locus tag CDR20291_0991 (cdPadR1) as part of our broader goal aimed at elucidating transcription regulatory mechanisms involved in virulence and antibiotic resistance of the recently emergent hypervirulent C. difficile strain R20291. cdPadR1 is genomically positioned near genes that are involved in stress response and virulence. In addition, it was previously reported that cdPadR1 and a homologue from the historical C. difficile strain 630 (CD630_1154) were differentially expressed when exposed to stressors, including antibiotics. The crystal structure of cdPadR1 was determined to 1.9 Å resolution, which revealed that it belongs to the PadR-s2 subfamily of PadR transcriptional regulators. cdPadR1 binds its own promoter and other promoter regions from within the C. difficile R20291 genome. DNA binding experiments demonstrated that cdPadR1 binds a region comprised of inverted repeats and an AT-rich core with the predicted specific binding motif, GTACTAT(N2)ATTATA(N)AGTA, within its own promoter that is also present in 200 other regions in the C. difficile R20291 genome. Mutation of the highly conserved W in α4 of the effector binding/oligomerization domain, which is predicted to be involved in multi-drug recognition and dimerization in other PadR-s2 proteins, resulted in alterations of cdPadR1 binding to the predicted binding motif, potentially due to loss of higher order oligomerization. Our results indicate that cdPadR1 binds a region within its own promoter consisting of the binding motif GTACTAT(N2)ATTATA(N)AGTA and seems to associate non-specifically with longer DNA fragments in vitro, which may facilitate promoter and

  3. The switch from fermentation to respiration in Saccharomyces cerevisiae is regulated by the Ert1 transcriptional activator/repressor.

    Science.gov (United States)

    Gasmi, Najla; Jacques, Pierre-Etienne; Klimova, Natalia; Guo, Xiao; Ricciardi, Alessandra; Robert, François; Turcotte, Bernard

    2014-10-01

    In the yeast Saccharomyces cerevisiae, fermentation is the major pathway for energy production, even under aerobic conditions. However, when glucose becomes scarce, ethanol produced during fermentation is used as a carbon source, requiring a shift to respiration. This adaptation results in massive reprogramming of gene expression. Increased expression of genes for gluconeogenesis and the glyoxylate cycle is observed upon a shift to ethanol and, conversely, expression of some fermentation genes is reduced. The zinc cluster proteins Cat8, Sip4, and Rds2, as well as Adr1, have been shown to mediate this reprogramming of gene expression. In this study, we have characterized the gene YBR239C encoding a putative zinc cluster protein and it was named ERT1 (ethanol regulated transcription factor 1). ChIP-chip analysis showed that Ert1 binds to a limited number of targets in the presence of glucose. The strongest enrichment was observed at the promoter of PCK1 encoding an important gluconeogenic enzyme. With ethanol as the carbon source, enrichment was observed with many additional genes involved in gluconeogenesis and mitochondrial function. Use of lacZ reporters and quantitative RT-PCR analyses demonstrated that Ert1 regulates expression of its target genes in a manner that is highly redundant with other regulators of gluconeogenesis. Interestingly, in the presence of ethanol, Ert1 is a repressor of PDC1 encoding an important enzyme for fermentation. We also show that Ert1 binds directly to the PCK1 and PDC1 promoters. In summary, Ert1 is a novel factor involved in the regulation of gluconeogenesis as well as a key fermentation gene.

  4. Transcription dynamics of inducible genes modulated by negative regulations.

    Science.gov (United States)

    Li, Yanyan; Tang, Moxun; Yu, Jianshe

    2015-06-01

    Gene transcription is a stochastic process in single cells, in which genes transit randomly between active and inactive states. Transcription of many inducible genes is also tightly regulated: It is often stimulated by extracellular signals, activated through signal transduction pathways and later repressed by negative regulations. In this work, we study the nonlinear dynamics of the mean transcription level of inducible genes modulated by the interplay of the intrinsic transcriptional randomness and the repression by negative regulations. In our model, we integrate negative regulations into gene activation process, and make the conventional assumption on the production and degradation of transcripts. We show that, whether or not the basal transcription is temporarily terminated when cells are stimulated, the mean transcription level grows in the typical up and down pattern commonly observed in immune response genes. With the help of numerical simulations, we clarify the delicate impact of the system parameters on the transcription dynamics, and demonstrate how our model generates the distinct temporal gene-induction patterns in mouse fibroblasts discerned in recent experiments.

  5. Thermodynamics-based models of transcriptional regulation by enhancers: the roles of synergistic activation, cooperative binding and short-range repression.

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    Xin He

    Full Text Available Quantitative models of cis-regulatory activity have the potential to improve our mechanistic understanding of transcriptional regulation. However, the few models available today have been based on simplistic assumptions about the sequences being modeled, or heuristic approximations of the underlying regulatory mechanisms. We have developed a thermodynamics-based model to predict gene expression driven by any DNA sequence, as a function of transcription factor concentrations and their DNA-binding specificities. It uses statistical thermodynamics theory to model not only protein-DNA interaction, but also the effect of DNA-bound activators and repressors on gene expression. In addition, the model incorporates mechanistic features such as synergistic effect of multiple activators, short range repression, and cooperativity in transcription factor-DNA binding, allowing us to systematically evaluate the significance of these features in the context of available expression data. Using this model on segmentation-related enhancers in Drosophila, we find that transcriptional synergy due to simultaneous action of multiple activators helps explain the data beyond what can be explained by cooperative DNA-binding alone. We find clear support for the phenomenon of short-range repression, where repressors do not directly interact with the basal transcriptional machinery. We also find that the binding sites contributing to an enhancer's function may not be conserved during evolution, and a noticeable fraction of these undergo lineage-specific changes. Our implementation of the model, called GEMSTAT, is the first publicly available program for simultaneously modeling the regulatory activities of a given set of sequences.

  6. Synergy and antagonism of active constituents of ADAPT-232 on transcriptional level of metabolic regulation of isolated neuroglial cells.

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    Alexander George Panossian

    2013-02-01

    Full Text Available Gene expression profiling was performed on the human neuroglial cell line T98G after treatment with adaptogen ADAPT-232 and its constituents – extracts of Eleutherococcus senticosus root, Schisandra chinensis berry, and Rhodiola rosea root as well as several constituents individually, namely, eleutheroside E, schizandrin B, salidroside, triandrin, and tyrosol. A common feature for all tested adaptogens was their effect on G-protein-coupled receptor (GPCR signaling pathways, i.e. cAMP, phospholipase C and phosphatidylinositol signal transduction pathways. Adaptogens may reduce the cAMP level in brain cells by downregulation of adenylate cyclase gene ADC2Y and upregulation of phosphodiestherase gene PDE4D that is essential for energy homeostasis as well as for switching from catabolic to anabolic states and vice versa. All tested adaptogens up-regulated the PLCB1 gene, which encodes phosphoinositide-specific phospholipase C (PLC and phosphatidylinositol 3-kinases (PI3Ks, key players for the regulation of NF-B-mediated defense responses. Other common targets of adaptogens included genes encoding ERα estrogen receptor(2.9-22.6 fold down-regulation, cholesterol ester transfer protein (5.1-10.6 fold down-regulation, heat shock protein Hsp70 (3.0-45.0 fold up-regulation, serpin peptidase inhibitor (neuroserpin, and 5-HT3 receptor of serotonin (2.2-6.6 fold down-regulation. These findings can be reconciled with the observed beneficial effects of adaptogens in behavioral, mental and aging-associated disorders. Combining two or more active substances in one mixture significantly changes deregulated genes profiles: synergetic interactions result in activation of genes that none of the individual substances affected, while antagonistic interactions result in suppression some genes activated by individual substances. Merging of deregulated genes array profiles and intracellular networks is specific to the new substance with unique pharmacological

  7. Krüppel-like factor 4, a novel transcription factor regulates microglial activation and subsequent neuroinflammation

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    Das Sulagna

    2010-10-01

    Full Text Available Abstract Background Activation of microglia, the resident macrophages of the central nervous system (CNS, is the hallmark of neuroinflammation in neurodegenerative diseases and other pathological conditions associated with CNS infection. The activation of microglia is often associated with bystander neuronal death. Nuclear factor-κB (NF-κB is one of the important transcription factors known to be associated with microglial activation which upregulates the expression of inducible nitric oxide synthase (iNOS, cyclooxygenase-2 (Cox-2 and other pro-inflammatory cytokines. Recent studies have focused on the role of Krüppel-like factor 4 (Klf4, one of the zinc-finger transcription factors, in mediating inflammation. However, these studies were limited to peripheral system and its role in CNS is not understood. Our studies focused on the possible role of Klf4 in mediating CNS inflammation. Methods For in vitro studies, mouse microglial BV-2 cell lines were treated with 500 ng/ml Salmonella enterica lipopolysacchride (LPS. Brain tissues were isolated from BALB/c mice administered with 5 mg/kg body weight of LPS. Expressions of Klf4, Cox-2, iNOS and pNF-κB were evaluated using western blotting, quantitative real time PCR, and reverse transcriptase polymerase chain reactions (RT-PCRs. Klf4 knockdown was carried out using SiRNA specific for Klf4 mRNA and luciferase assays and electromobility shift assay (EMSA were performed to study the interaction of Klf4 to iNOS promoter elements in vitro. Co-immunoprecipitation of Klf4 and pNF-κB was done in order to study a possible interaction between the two transcription factors. Results LPS stimulation increased Klf4 expression in microglial cells in a time- and dose-dependent manner. Knockdown of Klf4 resulted in decreased levels of the pro-inflammatory cytokines TNF-α, MCP-1 and IL-6, along with a significant decrease in iNOS and Cox-2 expression. NO production also decreased as a result of Klf4 knockdown

  8. Transcriptional Regulation by CHIP/LDB Complexes

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    Bronstein, Revital; Levkovitz, Liron; Yosef, Nir; Yanku, Michaela; Ruppin, Eytan; Sharan, Roded; Westphal, Heiner; Oliver, Brian; Segal, Daniel

    2010-01-01

    It is increasingly clear that transcription factors play versatile roles in turning genes “on” or “off” depending on cellular context via the various transcription complexes they form. This poses a major challenge in unraveling combinatorial transcription complex codes. Here we use the powerful genetics of Drosophila combined with microarray and bioinformatics analyses to tackle this challenge. The nuclear adaptor CHIP/LDB is a major developmental regulator capable of forming tissue-specific transcription complexes with various types of transcription factors and cofactors, making it a valuable model to study the intricacies of gene regulation. To date only few CHIP/LDB complexes target genes have been identified, and possible tissue-dependent crosstalk between these complexes has not been rigorously explored. SSDP proteins protect CHIP/LDB complexes from proteasome dependent degradation and are rate-limiting cofactors for these complexes. By using mutations in SSDP, we identified 189 down-stream targets of CHIP/LDB and show that these genes are enriched for the binding sites of APTEROUS (AP) and PANNIER (PNR), two well studied transcription factors associated with CHIP/LDB complexes. We performed extensive genetic screens and identified target genes that genetically interact with components of CHIP/LDB complexes in directing the development of the wings (28 genes) and thoracic bristles (23 genes). Moreover, by in vivo RNAi silencing we uncovered novel roles for two of the target genes, xbp1 and Gs-alpha, in early development of these structures. Taken together, our results suggest that loss of SSDP disrupts the normal balance between the CHIP-AP and the CHIP-PNR transcription complexes, resulting in down-regulation of CHIP-AP target genes and the concomitant up-regulation of CHIP-PNR target genes. Understanding the combinatorial nature of transcription complexes as presented here is crucial to the study of transcription regulation of gene batteries required

  9. The Lrp family of transcriptional regulators

    NARCIS (Netherlands)

    Brinkman, A.B.; Ettema, T.J.G.; Vos, de W.M.; Oost, van der J.

    2003-01-01

    Genome analysis has revealed that members of the Lrp family of transcriptional regulators are widely distributed among prokaryotes, both bacteria and archaea. The archetype Leucine-responsive Regulatory Protein from Escherichia coli is a global regulator involved in modulating a variety of metabolic

  10. Transcriptional Regulation and Macrophage Differentiation.

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    Hume, David A; Summers, Kim M; Rehli, Michael

    2016-06-01

    Monocytes and macrophages are professional phagocytes that occupy specific niches in every tissue of the body. Their survival, proliferation, and differentiation are controlled by signals from the macrophage colony-stimulating factor receptor (CSF-1R) and its two ligands, CSF-1 and interleukin-34. In this review, we address the developmental and transcriptional relationships between hematopoietic progenitor cells, blood monocytes, and tissue macrophages as well as the distinctions from dendritic cells. A huge repertoire of receptors allows monocytes, tissue-resident macrophages, or pathology-associated macrophages to adapt to specific microenvironments. These processes create a broad spectrum of macrophages with different functions and individual effector capacities. The production of large transcriptomic data sets in mouse, human, and other species provides new insights into the mechanisms that underlie macrophage functional plasticity.

  11. Regulation of Lactobacillus casei Sorbitol Utilization Genes Requires DNA-Binding Transcriptional Activator GutR and the Conserved Protein GutM▿

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    Alcántara, Cristina; Sarmiento-Rubiano, Luz Adriana; Monedero, Vicente; Deutscher, Josef; Pérez-Martínez, Gaspar; Yebra, María J.

    2008-01-01

    Sequence analysis of the five genes (gutRMCBA) downstream from the previously described sorbitol-6-phosphate dehydrogenase-encoding Lactobacillus casei gutF gene revealed that they constitute a sorbitol (glucitol) utilization operon. The gutRM genes encode putative regulators, while the gutCBA genes encode the EIIC, EIIBC, and EIIA proteins of a phosphoenolpyruvate-dependent sorbitol phosphotransferase system (PTSGut). The gut operon is transcribed as a polycistronic gutFRMCBA messenger, the expression of which is induced by sorbitol and repressed by glucose. gutR encodes a transcriptional regulator with two PTS-regulated domains, a galactitol-specific EIIB-like domain (EIIBGat domain) and a mannitol/fructose-specific EIIA-like domain (EIIAMtl domain). Its inactivation abolished gut operon transcription and sorbitol uptake, indicating that it acts as a transcriptional activator. In contrast, cells carrying a gutB mutation expressed the gut operon constitutively, but they failed to transport sorbitol, indicating that EIIBCGut negatively regulates GutR. A footprint analysis showed that GutR binds to a 35-bp sequence upstream from the gut promoter. A sequence comparison with the presumed promoter region of gut operons from various firmicutes revealed a GutR consensus motif that includes an inverted repeat. The regulation mechanism of the L. casei gut operon is therefore likely to be operative in other firmicutes. Finally, gutM codes for a conserved protein of unknown function present in all sequenced gut operons. A gutM mutant, the first constructed in a firmicute, showed drastically reduced gut operon expression and sorbitol uptake, indicating a regulatory role also for GutM. PMID:18676710

  12. Regulation of human protein S gene (PROS1) transcription

    NARCIS (Netherlands)

    Wolf, Cornelia de

    2006-01-01

    This thesis describes the investigation of the transcriptional regulation of the gene for anticoagulant plasma Protein S, PROS1. Protein S is a cofactor for Protein C in the Protein C anticoagulant pathway. The coagulation cascade is negatively regulated by this pathway through inactivation of activ

  13. The Ikaros transcription factor regulates responsiveness to IL-12 and expression of IL-2 receptor alpha in mature, activated CD8 T cells.

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    Eric T Clambey

    Full Text Available The Ikaros family of transcription factors is critical for normal T cell development while limiting malignant transformation. Mature CD8 T cells express multiple Ikaros family members, yet little is known about their function in this context. To test the functions of this gene family, we used retroviral transduction to express a naturally occurring, dominant negative (DN isoform of Ikaros in activated CD8 T cells. Notably, expression of DN Ikaros profoundly enhanced the competitive advantage of activated CD8 T cells cultured in IL-12, such that by 6 days of culture, DN Ikaros-transduced cells were 100-fold more abundant than control cells. Expression of a DN isoform of Helios, a related Ikaros-family transcription factor, conferred a similar advantage to transduced cells in IL-12. While DN Ikaros-transduced cells had higher expression of the IL-2 receptor alpha chain, DN Ikaros-transduced cells achieved their competitive advantage through an IL-2 independent mechanism. Finally, the competitive advantage of DN Ikaros-transduced cells was manifested in vivo, following adoptive transfer of transduced cells. These data identify the Ikaros family of transcription factors as regulators of cytokine responsiveness in activated CD8 T cells, and suggest a role for this family in influencing effector and memory CD8 T cell differentiation.

  14. Sesamol and sesame (Sesamum indicum) oil enhance macrophage cholesterol efflux via up-regulation of PPARγ1 and LXRα transcriptional activity in a MAPK-dependent manner.

    Science.gov (United States)

    Majdalawieh, Amin F; Ro, Hyo-Sung

    2015-08-01

    Cholesterol clearance by macrophages is a vital process to eliminate excess cholesterol from the body. Internalization of modified cholesterol by macrophages triggers overexpression of peroxisome proliferator-activated receptor γ1 (PPARγ1) and liver X receptor α (LXRα), two transcription factors that are critically involved in macrophage cholesterol efflux. Recent studies demonstrate that oral administration of sesamol derivative (INV-403) and sesame oil leads to a significant attenuation of atherosclerosis in Watanabe heritable hyperlipidemic rabbits and LDLR(-/-) mice, respectively. However, the exact molecular mechanisms underlying such anti-atherogenic effects remain largely unrevealed. Luciferase reporter assays were performed to assess the effects of sesamol and sesame oil on PPARγ1 and LXRα gene expression. The potential of sesamol and sesame oil to modulate cholesterol efflux was evaluated using (3)H-cholesterol efflux assays. Sesamol and sesame oil treatments lead to a significant up-regulation of PPARγ1 and LXRα expression and transcriptional activity in a MAPK-dependent manner. Importantly, primary macrophages display a significantly enhanced cholesterol efflux potential upon treatment with sesamol and sesame oil, and this stimulatory effect is mediated by MAPK signaling. Our findings suggest that the previously reported anti-atherogenic effects of sesamol and sesame oil could be attributed, at least in part, to enhanced PPARγ1 and LXRα expression and transcriptional activity leading to improved macrophage cholesterol efflux. Our study is novel in elucidating the molecular and cellular mechanisms underlying the protective effects of sesamol and sesame oil against atherosclerosis.

  15. Berberine regulates peroxisome proliferator-activated receptors and positive transcription elongation factor b expression in diabetic adipocytes.

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    Zhou, Jiyin; Zhou, Shiwen

    2010-12-15

    Berberine has hypoglycemic and hypolipidemic effects on diabetic rats. This study investigated the relationship between hypoglycemic and hypolipidemic effects of berberine and peroxisome proliferator-activated receptors (PPARs) and positive transcription elongation factor b (P-TEFb) (including cyclin-dependent kinase 9 (CDK9) and cyclin T1) in white adipose tissue of diabetic rats and RNA interference-treated 3T3-L1 cells. Berberine promoted differentiation and inhibited lipid accumulation of 3T3-L1 cells, further decreased PPARα/δ/γ, CDK9 and cyclin T1 mRNA and protein expression and decreased tumor necrosis factor α content in supernatants of both control and RNA interference-treated 3T3-L1 cells. After a 16-week induction with 35 mg/kg streptozotocin (i.p.) and high-carbohydrate/high-fat diet, diabetic rats were treated with 75, 150 and 300 mg/kg berberine and 100 mg/kg fenofibrate or 4 mg/kg rosiglitazone for another 16 weeks. Berberine decreased white adipose tissue to body weight ratio and adipocyte size and increased adipocyte number. Berberine upregulated PPARα/δ/γ, CDK9 and cyclin T1 mRNA and protein expression in adipose tissue, decreased tumor necrosis factor α and free fatty acid content and increased lipoprotein lipase activity in serum and adipose tissue. Berberine modulated metabolic related PPARs expression and differentiation related P-TEFb expression in adipocytes, which are associated with its hypoglycemic and hypolipidemic effects.

  16. Regulation of the Hippo Pathway Transcription Factor TEAD.

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    Lin, Kimberly C; Park, Hyun Woo; Guan, Kun-Liang

    2017-09-27

    The TEAD transcription factor family is best known for transcriptional output of the Hippo signaling pathway and has been implicated in processes such as development, cell growth and proliferation, tissue homeostasis, and regeneration. Our understanding of the functional importance of TEADs has increased dramatically since its initial discovery three decades ago. The majority of our knowledge of TEADs is in the context of Hippo signaling as nuclear DNA-binding proteins passively activated by Yes-associated protein (YAP) and transcriptional activator with PDZ-binding domain (TAZ), transcription coactivators downstream of the Hippo pathway. However, recent studies suggest that TEAD itself is actively regulated. Here, we highlight evidence demonstrating Hippo-independent regulation of TEADs and the potential impacts these studies may have on new cancer therapeutics. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. The NAD-Dependent Deacetylase Sirtuin-1 Regulates the Expression of Osteogenic Transcriptional Activator Runt-Related Transcription Factor 2 (Runx2 and Production of Matrix Metalloproteinase (MMP-13 in Chondrocytes in Osteoarthritis

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    Koh Terauchi

    2016-06-01

    Full Text Available Aging is one of the major pathologic factors associated with osteoarthritis (OA. Recently, numerous reports have demonstrated the impact of sirtuin-1 (Sirt1, which is the NAD-dependent deacetylase, on human aging. It has been demonstrated that Sirt1 induces osteogenic and chondrogenic differentiation of mesenchymal stem cells. However, the role of Sirt1 in the OA chondrocytes still remains unknown. We postulated that Sirt1 regulates a hypertrophic chondrocyte lineage and degeneration of articular cartilage through the activation of osteogenic transcriptional activator Runx2 and matrix metalloproteinase (MMP-13 in OA chondrocytes. To verify whether sirtuin-1 (Sirt1 regulates chondrocyte activity in OA, we studied expressions of Sirt1, Runx2 and production of MMP-13, and their associations in human OA chondrocytes. The expression of Sirt1 was ubiquitously observed in osteoarthritic chondrocytes; in contrast, Runx2 expressed in the osteophyte region in patients with OA and OA model mice. OA relating catabolic factor IL-1βincreased the expression of Runx2 in OA chondrocytes. OA chondrocytes, which were pretreated with Sirt1 inhibitor, inhibited the IL-1β-induced expression of Runx2 compared to the control. Since the Runx2 is a promotor of MMP-13 expression, Sirt1 inactivation may inhibit the Runx2 expression and the resultant down-regulation of MMP-13 production in chondrocytes. Our findings suggest thatSirt1 may regulate the expression of Runx2, which is the osteogenic transcription factor, and the production of MMP-13 from chondrocytes in OA. Since Sirt1 activity is known to be affected by several stresses, including inflammation and oxidative stress, as well as aging, SIRT may be involved in the development of OA.

  18. A bacteriophage transcription regulator inhibits bacterial transcription initiation by σ-factor displacement.

    Science.gov (United States)

    Liu, Bing; Shadrin, Andrey; Sheppard, Carol; Mekler, Vladimir; Xu, Yingqi; Severinov, Konstantin; Matthews, Steve; Wigneshweraraj, Sivaramesh

    2014-04-01

    Bacteriophages (phages) appropriate essential processes of bacterial hosts to benefit their own development. The multisubunit bacterial RNA polymerase (RNAp) enzyme, which catalyses DNA transcription, is targeted by phage-encoded transcription regulators that selectively modulate its activity. Here, we describe the structural and mechanistic basis for the inhibition of bacterial RNAp by the transcription regulator P7 encoded by Xanthomonas oryzae phage Xp10. We reveal that P7 uses a two-step mechanism to simultaneously interact with the catalytic β and β' subunits of the bacterial RNAp and inhibits transcription initiation by inducing the displacement of the σ(70)-factor on initial engagement of RNAp with promoter DNA. The new mode of interaction with and inhibition mechanism of bacterial RNAp by P7 underscore the remarkable variety of mechanisms evolved by phages to interfere with host transcription.

  19. Transcriptional master regulator analysis in breast cancer genetic networks.

    Science.gov (United States)

    Tovar, Hugo; García-Herrera, Rodrigo; Espinal-Enríquez, Jesús; Hernández-Lemus, Enrique

    2015-12-01

    Gene regulatory networks account for the delicate mechanisms that control gene expression. Under certain circumstances, gene regulatory programs may give rise to amplification cascades. Such transcriptional cascades are events in which activation of key-responsive transcription factors called master regulators trigger a series of gene expression events. The action of transcriptional master regulators is then important for the establishment of certain programs like cell development and differentiation. However, such cascades have also been related with the onset and maintenance of cancer phenotypes. Here we present a systematic implementation of a series of algorithms aimed at the inference of a gene regulatory network and analysis of transcriptional master regulators in the context of primary breast cancer cells. Such studies were performed in a highly curated database of 880 microarray gene expression experiments on biopsy-captured tissue corresponding to primary breast cancer and healthy controls. Biological function and biochemical pathway enrichment analyses were also performed to study the role that the processes controlled - at the transcriptional level - by such master regulators may have in relation to primary breast cancer. We found that transcription factors such as AGTR2, ZNF132, TFDP3 and others are master regulators in this gene regulatory network. Sets of genes controlled by these regulators are involved in processes that are well-known hallmarks of cancer. This kind of analyses may help to understand the most upstream events in the development of phenotypes, in particular, those regarding cancer biology.

  20. Anticancer activity of eugenol is not related to regulation of the oncogenic transcription factor Forkhead Box M1

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    Luiz Alexandre Marques Wiirzler

    2016-09-01

    Full Text Available Genome-wide gene expression profiling of cancers has consistently identified the FOXM1 as one of the most commonly upregulated genes in cancer cells that plays an essential role in the regulation of a wide spectrum of biological processes, including inhibition of apoptosis. Since the anticancer activity of EUG reported in the literature is related to induction of apoptosis in cancer cells, we hypothesized that there is a correlation between the EUG-induced apoptosis effect and downregulation of FOXM1. A series of experiments were conducted to evaluate the effect of EUG on cellular viability of cancer cells (MTT and its potential regulatory effect on FOXM1 protein levels (western blots. Our findings confirm the anticancer effect of EUG on different human cancer cell lines as previously reported in the literature (SKBR3 LC50: 318.6; HT29 LC50: 525.5; and HepG2 LC50: 2090.0 µM. However, we demonstrated that EUG does not regulate the FOXM1. The results evidenced the anticancer effect of EUG on three cancer cell lines and showed that the EUG- apoptosis induced effect is not related to regulation of FOXM1 at the protein level. Further studies must be done to provide information on the mechanism of action of this agent.

  1. Redox-regulated transcription in plants: Emerging concepts

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    Jehad Shaikhali

    2017-09-01

    Full Text Available In plants, different stimuli, both internal and external, activate production of reactive oxygen species (ROS. Photosynthesis is considered as high rate redox-metabolic process with rapid transients including light/photon capture, electron fluxes, and redox potentials that can generate ROS; thus, regulatory systems are required to minimize ROS production. Despite their potential for causing harmful oxidations, it is now accepted that redox homeostasis mechanisms that maintain the intracellular reducing environment make it possible to use ROS as powerful signaling molecules within and between cells. Redox and ROS information from the chloroplasts is a fine-tuning mechanism both inside the chloroplast and as retrograde signal to the cytosol and nucleus to control processes such as gene expression/transcription and translation. Wide repertoires of downstream target genes expression (activation/repression is regulated by transcription factors. In many cases, transcription factors function through various mechanisms that affect their subcellular localization and or activity. Some post-translational modifications (PTMs known to regulate the functional state of transcription factors are phosphorylation, acetylation, and SUMOylation, ubiquitylation and disulfide formation. Recently, oxPTMs, targeted in redox proteomics, can provide the bases to study redox regulation of low abundant nuclear proteins. This review summarizes the recent advances on how cellular redox status can regulate transcription factor activity, the implications of this regulation for plant growth and development, and by which plants respond to environmental/abiotic stresses.

  2. Soybean miR172c Targets the Repressive AP2 Transcription Factor NNC1 to Activate ENOD40 Expression and Regulate Nodule Initiation[C][W

    Science.gov (United States)

    Wang, Youning; Wang, Lixiang; Zou, Yanmin; Chen, Liang; Cai, Zhaoming; Zhang, Senlei; Zhao, Fang; Tian, Yinping; Jiang, Qiong; Ferguson, Brett J.; Gresshoff, Peter M.; Li, Xia

    2014-01-01

    MicroRNAs are noncoding RNAs that act as master regulators to modulate various biological processes by posttranscriptionally repressing their target genes. Repression of their target mRNA(s) can modulate signaling cascades and subsequent cellular events. Recently, a role for miR172 in soybean (Glycine max) nodulation has been described; however, the molecular mechanism through which miR172 acts to regulate nodulation has yet to be explored. Here, we demonstrate that soybean miR172c modulates both rhizobium infection and nodule organogenesis. miR172c was induced in soybean roots inoculated with either compatible Bradyrhizobium japonicum or lipooligosaccharide Nod factor and was highly upregulated during nodule development. Reduced activity and overexpression of miR172c caused dramatic changes in nodule initiation and nodule number. We show that soybean miR172c regulates nodule formation by repressing its target gene, Nodule Number Control1, which encodes a protein that directly targets the promoter of the early nodulin gene, ENOD40. Interestingly, transcriptional levels of miR172c were regulated by both Nod Factor Receptor1α/5α-mediated activation and by autoregulation of nodulation-mediated inhibition. Thus, we established a direct link between miR172c and the Nod factor signaling pathway in addition to adding a new layer to the precise nodulation regulation mechanism of soybean. PMID:25549672

  3. The NDR/LATS kinase Cbk1 controls the activity of the transcriptional regulator Bcr1 during biofilm formation in Candida albicans.

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    Pilar Gutiérrez-Escribano

    Full Text Available In nature, many microorganisms form specialized complex, multicellular, surface-attached communities called biofilms. These communities play critical roles in microbial pathogenesis. The fungal pathogen Candida albicans is associated with catheter-based infections due to its ability to establish biofilms. The transcription factor Bcr1 is a master regulator of C. albicans biofilm development, although the full extent of its regulation remains unknown. Here, we report that Bcr1 is a phosphoprotein that physically interacts with the NDR kinase Cbk1 and undergoes Cbk1-dependent phosphorylation. Mutating the two putative Cbk1 phosphoacceptor residues in Bcr1 to alanine markedly impaired Bcr1 function during biofilm formation and virulence in a mouse model of disseminated candidiasis. Cells lacking Cbk1, or any of its upstream activators, also had reduced biofilm development. Notably, mutating the two putative Cbk1 phosphoacceptor residues in Bcr1 to glutamate in cbk1Δ cells upregulated the transcription of Bcr1-dependent genes and partially rescued the biofilm defects of a cbk1Δ strain. Therefore, our data uncovered a novel role of the NDR/LATS kinase Cbk1 in the regulation of biofilm development through the control of Bcr1.

  4. Global transcription regulation by DNA topoisomerase I in exponentially growing Saccharomyces cerevisiae cells: activation of telomere-proximal genes by TOP1 deletion.

    Science.gov (United States)

    Lotito, Luca; Russo, Alessandra; Chillemi, Giovanni; Bueno, Susana; Cavalieri, Duccio; Capranico, Giovanni

    2008-03-21

    To establish the cellular functions of DNA topoisomerase I-B (Top1p) at a global level, we have determined the expression profiles and histone modification patterns affected by TOP1 gene deletion (DeltaTOP1) in Saccharomyces cerevisiae. In exponentially growing cells, DeltaTOP1 specifically increases transcription of telomere-proximal genes and decreases glucose utilization and energy production pathways. Immunoprecipitation data demonstrate that Top1p can bind to and is catalytically active at telomeric DNA repeats, and that both DeltaTOP1 and an inactive Y727F Top1p mutant increase H4 histone acetylation at telomere-proximal regions. Interestingly, while the Y727F mutation has no influence on enzyme recruitment to chromatin sites, it has a marked effect on H4 K16 acetylation at subtelomeric regions. The Top1p mutation also increases H3 histone K4 dimethylation, which has been associated with gene transcription, at 3' termini of subtelomeric genes. No major effect of DeltaTOP1 or mutation was detected on Sir3p recruitment; however, DeltaTOP1 has an effect on transcript levels of genes known to regulate telomeric silencing. Thus, the findings indicate that Top1p activity can favor both a repressed chromatin organization and a reduced gene expression level at telomere-proximal regions in yeast. As telomere-proximal regions are known to be enriched for stress-activated genes, our findings show that Top1p can optimize transcript levels for cell growth in exponentially growing cells under a synthetic medium with glucose.

  5. The post-transcriptional regulator rsmA/csrA activates T3SS by stabilizing the 5' UTR of hrpG, the master regulator of hrp/hrc genes, in Xanthomonas.

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    Maxuel O Andrade

    2014-02-01

    Full Text Available The RsmA/CsrA family of the post-transcriptional regulators of bacteria is involved in the regulation of many cellular processes, including pathogenesis. In this study, we demonstrated that rsmA not only is required for the full virulence of the phytopathogenic bacterium Xanthomonas citri subsp. citri (XCC but also contributes to triggering the hypersensitive response (HR in non-host plants. Deletion of rsmA resulted in significantly reduced virulence in the host plant sweet orange and a delayed and weakened HR in the non-host plant Nicotiana benthamiana. Microarray, quantitative reverse-transcription PCR, western-blotting, and GUS assays indicated that RsmA regulates the expression of the type 3 secretion system (T3SS at both transcriptional and post-transcriptional levels. The regulation of T3SS by RsmA is a universal phenomenon in T3SS-containing bacteria, but the specific mechanism seems to depend on the interaction between a particular bacterium and its hosts. For Xanthomonads, the mechanism by which RsmA activates T3SS remains unknown. Here, we show that RsmA activates the expression of T3SS-encoding hrp/hrc genes by directly binding to the 5' untranslated region (UTR of hrpG, the master regulator of the hrp/hrc genes in XCC. RsmA stabilizes hrpG mRNA, leading to increased accumulation of HrpG proteins and subsequently, the activation of hrp/hrc genes. The activation of the hrp/hrc genes by RsmA via HrpG was further supported by the observation that ectopic overexpression of hrpG in an rsmA mutant restored its ability to cause disease in host plants and trigger HR in non-host plants. RsmA also stabilizes the transcripts of another T3SS-associated hrpD operon by directly binding to the 5' UTR region. Taken together, these data revealed that RsmA primarily activates T3SS by acting as a positive regulator of hrpG and that this regulation is critical to the pathogenicity of XCC.

  6. The regulation of transcription in memory consolidation.

    Science.gov (United States)

    Alberini, Cristina M; Kandel, Eric R

    2014-12-04

    De novo transcription of DNA is a fundamental requirement for the formation of long-term memory. It is required during both consolidation and reconsolidation, the posttraining and postreactivation phases that change the state of the memory from a fragile into a stable and long-lasting form. Transcription generates both mRNAs that are translated into proteins, which are necessary for the growth of new synaptic connections, as well as noncoding RNA transcripts that have regulatory or effector roles in gene expression. The result is a cascade of events that ultimately leads to structural changes in the neurons that mediate long-term memory storage. The de novo transcription, critical for synaptic plasticity and memory formation, is orchestrated by chromatin and epigenetic modifications. The complexity of transcription regulation, its temporal progression, and the effectors produced all contribute to the flexibility and persistence of long-term memory formation. In this article, we provide an overview of the mechanisms contributing to this transcriptional regulation underlying long-term memory formation.

  7. Dual-level regulation of ACC synthase activity by MPK3/MPK6 cascade and its downstream WRKY transcription factor during ethylene induction in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Guojing Li

    2012-06-01

    Full Text Available Plants under pathogen attack produce high levels of ethylene, which plays important roles in plant immunity. Previously, we reported the involvement of ACS2 and ACS6, two Type I ACS isoforms, in Botrytis cinerea-induced ethylene biosynthesis and their regulation at the protein stability level by MPK3 and MPK6, two Arabidopsis pathogen-responsive mitogen-activated protein kinases (MAPKs. The residual ethylene induction in the acs2/acs6 double mutant suggests the involvement of additional ACS isoforms. It is also known that a subset of ACS genes, including ACS6, is transcriptionally induced in plants under stress or pathogen attack. However, the importance of ACS gene activation and the regulatory mechanism(s are not clear. In this report, we demonstrate using genetic analysis that ACS7 and ACS11, two Type III ACS isoforms, and ACS8, a Type II ACS isoform, also contribute to the B. cinerea-induced ethylene production. In addition to post-translational regulation, transcriptional activation of the ACS genes also plays a critical role in sustaining high levels of ethylene induction. Interestingly, MPK3 and MPK6 not only control the stability of ACS2 and ACS6 proteins via direct protein phosphorylation but also regulate the expression of ACS2 and ACS6 genes. WRKY33, another MPK3/MPK6 substrate, is involved in the MPK3/MPK6-induced ACS2/ACS6 gene expression based on genetic analyses. Furthermore, chromatin-immunoprecipitation assay reveals the direct binding of WRKY33 to the W-boxes in the promoters of ACS2 and ACS6 genes in vivo, suggesting that WRKY33 is directly involved in the activation of ACS2 and ACS6 expression downstream of MPK3/MPK6 cascade in response to pathogen invasion. Regulation of ACS activity by MPK3/MPK6 at both transcriptional and protein stability levels plays a key role in determining the kinetics and magnitude of ethylene induction.

  8. The liver-enriched transcription factor CREBH is nutritionally regulated and activated by fatty acids and PPAR{alpha}

    Energy Technology Data Exchange (ETDEWEB)

    Danno, Hirosuke; Ishii, Kiyo-aki; Nakagawa, Yoshimi; Mikami, Motoki; Yamamoto, Takashi; Yabe, Sachiko; Furusawa, Mika; Kumadaki, Shin; Watanabe, Kazuhisa; Shimizu, Hidehisa; Matsuzaka, Takashi; Kobayashi, Kazuto; Takahashi, Akimitsu; Yatoh, Shigeru; Suzuki, Hiroaki; Yamada, Nobuhiro [Department of Internal Medicine (Endocrinology and Metabolism), Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba Ibaraki 305-8575 (Japan); Shimano, Hitoshi, E-mail: hshimano@md.tsukuba.ac.jp [Department of Internal Medicine (Endocrinology and Metabolism), Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba Ibaraki 305-8575 (Japan)

    2010-01-08

    To elucidate the physiological role of CREBH, the hepatic mRNA and protein levels of CREBH were estimated in various feeding states of wild and obesity mice. In the fast state, the expression of CREBH mRNA and nuclear protein were high and profoundly suppressed by refeeding in the wild-type mice. In ob/ob mice, the refeeding suppression was impaired. The diet studies suggested that CREBH expression was activated by fatty acids. CREBH mRNA levels in the mouse primary hepatocytes were elevated by addition of the palmitate, oleate and eicosapenonate. It was also induced by PPAR{alpha} agonist and repressed by PPAR{alpha} antagonist. Luciferase reporter gene assays indicated that the CREBH promoter activity was induced by fatty acids and co-expression of PPAR{alpha}. Deletion studies identified the PPRE for PPAR{alpha} activation. Electrophoretic mobility shift assay and chromatin immunoprecipitation (ChIP) assay confirmed that PPAR{alpha} directly binds to the PPRE. Activation of CREBH at fasting through fatty acids and PPAR{alpha} suggest that CREBH is involved in nutritional regulation.

  9. Regulation of the Ets transcription factor Tel

    NARCIS (Netherlands)

    Roukens, Mark Guido

    2010-01-01

    In this thesis we report novel studies on the molecular regulation of the transcriptional repressor Tel (Translocation Ets Leukemia). The work in this thesis is presented as follows: Chapter 1 is an introduction which summarizes the literature about Tel and its Drosophila orthologue Yan as it was k

  10. MrkH, a novel c-di-GMP-dependent transcriptional activator, controls Klebsiella pneumoniae biofilm formation by regulating type 3 fimbriae expression.

    Directory of Open Access Journals (Sweden)

    Jonathan J Wilksch

    2011-08-01

    Full Text Available Klebsiella pneumoniae causes significant morbidity and mortality worldwide, particularly amongst hospitalized individuals. The principle mechanism for pathogenesis in hospital environments involves the formation of biofilms, primarily on implanted medical devices. In this study, we constructed a transposon mutant library in a clinical isolate, K. pneumoniae AJ218, to identify the genes and pathways implicated in biofilm formation. Three mutants severely defective in biofilm formation contained insertions within the mrkABCDF genes encoding the main structural subunit and assembly machinery for type 3 fimbriae. Two other mutants carried insertions within the yfiN and mrkJ genes, which encode GGDEF domain- and EAL domain-containing c-di-GMP turnover enzymes, respectively. The remaining two isolates contained insertions that inactivated the mrkH and mrkI genes, which encode for novel proteins with a c-di-GMP-binding PilZ domain and a LuxR-type transcriptional regulator, respectively. Biochemical and functional assays indicated that the effects of these factors on biofilm formation accompany concomitant changes in type 3 fimbriae expression. We mapped the transcriptional start site of mrkA, demonstrated that MrkH directly activates transcription of the mrkA promoter and showed that MrkH binds strongly to the mrkA regulatory region only in the presence of c-di-GMP. Furthermore, a point mutation in the putative c-di-GMP-binding domain of MrkH completely abolished its function as a transcriptional activator. In vivo analysis of the yfiN and mrkJ genes strongly indicated their c-di-GMP-specific function as diguanylate cyclase and phosphodiesterase, respectively. In addition, in vitro assays showed that purified MrkJ protein has strong c-di-GMP phosphodiesterase activity. These results demonstrate for the first time that c-di-GMP can function as an effector to stimulate the activity of a transcriptional activator, and explain how type 3 fimbriae

  11. Arabidopsis FHY3 and FAR1 Regulate Light-Induced myo-Inositol Biosynthesis and Oxidative Stress Responses by Transcriptional Activation of MIPS1.

    Science.gov (United States)

    Ma, Lin; Tian, Tian; Lin, Rongcheng; Deng, Xing-Wang; Wang, Haiyang; Li, Gang

    2016-04-04

    myo-Inositol-1-phosphate synthase (MIPS) catalyzes the limiting step of inositol biosynthesis and has crucial roles in plant growth and development. In response to stress, the transcription of MIPS1 is induced and the biosynthesis of inositol or inositol derivatives is promoted by unknown mechanisms. Here, we found that the light signaling protein FAR-RED ELONGATED HYPOCOTYL3 (FHY3) and its homolog FAR-RED IMPAIRED RESPONSE1 (FAR1) regulate light-induced inositol biosynthesis and oxidative stress responses by activating the transcription of MIPS1. Disruption of FHY3 and FAR1 caused light-induced cell death after dark-light transition, precocious leaf senescence, and increased sensitivity to oxidative stress. Reduction of salicylic acid (SA) accumulation by overexpression of SALICYLIC ACID 3-HYDROXYLASE largely suppressed the cell death phenotype of fhy3 far1 mutant plants, suggesting that FHY3- and FAR1-mediated cell death is dependent on SA. Furthermore, comparative analysis of chromatin immunoprecipitation sequencing and microarray results revealed that FHY3 and FAR1 directly target both MIPS1 and MIPS2. The fhy3 far1 mutant plants showed severely decreased MIPS1/2 transcript levels and reduced inositol levels. Conversely, constitutive expression of MIPS1 partially rescued the inositol contents, caused reduced transcript levels of SA-biosynthesis genes, and prevented oxidative stress in fhy3 far1. Taken together, our results indicate that the light signaling proteins FHY3 and FAR1 directly bind the promoter of MIPS1 to activate its expression and thereby promote inositol biosynthesis to prevent light-induced oxidative stress and SA-dependent cell death.

  12. Transcriptional activation of pyoluteorin operon mediated by the LysR-type regulator PltR bound at a 22 bp lys box in Pseudomonas aeruginosa M18.

    Directory of Open Access Journals (Sweden)

    Sainan Li

    Full Text Available Pseudomonas aeruginosa M18, a rhizosphere-isolated bacterial strain showing strong antifungal activity, can produce secondary metabolites such as phenazine-1-carboxylic acid and pyoluteorin (Plt. The LysR-type transcriptional regulator PltR activates the Plt biosynthesis operon pltLABCDEFG, the expression of which is induced by Plt. Here, we identified and characterized the non-conserved pltL promoter (pltLp specifically activated by PltR and its upstream neighboring lys box from the complicated pltR-pltL intergenic sequence. The 22 bp palindromic lys box, which consists of two 9 bp complementary inverted repeats interrupted by 4 bp, was found to contain the conserved, GC-rich LysR-binding motif (T-N(11-A. Evidence obtained in vivo from mutational and lacZ report analyses and in vitro from electrophoretic mobility shift assays reveals that the PltR protein directly bound to the pltLp region as the indispensable binding motif "lys box", thereby transcriptionally activating the pltLp-driven plt operon expression. Plt, as a potential non-essential coinducer of PltR, specifically induced the pltLp expression and thus strengthened its biosynthetic plt operon expression.

  13. Cyclin-dependent kinase 5 modulates the transcriptional activity of the mineralocorticoid receptor and regulates expression of brain-derived neurotrophic factor.

    Science.gov (United States)

    Kino, Tomoshige; Jaffe, Howard; Amin, Niranjana D; Chakrabarti, Mayukh; Zheng, Ya-Li; Chrousos, George P; Pant, Harish C

    2010-05-01

    Glucocorticoids, major end effectors of the stress response, play an essential role in the homeostasis of the central nervous system (CNS) and contribute to memory consolidation and emotional control through their intracellular receptors, the glucocorticoid and mineralocorticoid receptors. Cyclin-dependent kinase 5 (CDK5), on the other hand, plays important roles in the morphogenesis and functions of the central nervous system, and its aberrant activation has been associated with development of neurodegenerative disorders. We previously reported that CDK5 phosphorylated the glucocorticoid receptor and modulated its transcriptional activity. Here we found that CDK5 also regulated mineralocorticoid receptor-induced transcriptional activity by phosphorylating multiple serine and threonine residues located in its N-terminal domain through physical interaction. Aldosterone and dexamethasone, respectively, increased and suppressed mRNA/protein expression of brain-derived neurotrophic factor (BDNF) in rat cortical neuronal cells, whereas the endogenous glucocorticoid corticosterone showed a biphasic effect. CDK5 enhanced the effect of aldosterone and dexamethasone on BDNF expression. Because this neurotrophic factor plays critical roles in neuronal viability, synaptic plasticity, consolidation of memory, and emotional changes, we suggest that aberrant activation of CDK5 might influence these functions through corticosteroid receptors/BDNF.

  14. Transcriptional regulation of bone and joint remodeling by NFAT

    OpenAIRE

    2010-01-01

    Osteoporosis and arthritis are highly prevalent diseases and a significant cause of morbidity and mortality worldwide. These diseases result from aberrant tissue remodeling leading to weak, fracture-prone bones or painful, dysfunctional joints. The nuclear factor of activated T cells (NFAT) transcription factor family controls diverse biologic processes in vertebrates. Here, we review the scientific evidence that links NFAT-regulated gene transcription to bone and joint pathology. A particula...

  15. Id transcriptional regulators in adipogenesis and adipose tissue metabolism.

    Science.gov (United States)

    Patil, Mallikarjun; Sharma, Bal Krishan; Satyanarayana, Ande

    2014-06-01

    Id proteins (Id1-Id4) are helix-loop-helix (HLH) transcriptional regulators that lack a basic DNA binding domain. They act as negative regulators of basic helix-loop-helix (bHLH) transcription factors by forming heterodimers and inhibit their DNA binding and transcriptional activity. Id proteins are implicated in the regulation of various cellular mechanisms such as cell proliferation, cellular differentiation, cell fate determination, angiogenesis and tumorigenesis. A handful of recent studies also disclosed that Id proteins have critical functions in adipocyte differentiation and adipose tissue metabolism. Here, we reviewed the progress made thus far in understanding the specific functions of Id proteins in adipose tissue differentiation and metabolism. In addition to reviewing the known mechanisms of action, we also discuss possible additional mechanisms in which Id proteins might participate in regulating adipogenic and metabolic pathways.

  16. Arabidopsis STO/BBX24 negatively regulates UV-B signaling by interacting with COP1 and repressing HY5 transcriptional activity.

    Science.gov (United States)

    Jiang, Lei; Wang, Yan; Li, Qian-Feng; Björn, Lars Olof; He, Jun-Xian; Li, Shao-Shan

    2012-06-01

    UV-B (280-315 nm) is an integral part of solar radiation and can act either as a stress inducer or as a developmental signal. In recent years, increasing attention has been paid to the low-fluence UV-B-induced photomorphogenic response and several key players in this response have been identified, which include UVR8 (a UV-B-specific photoreceptor), COP1 (a WD40-repeat-containing RING finger protein), HY5 (a basic zipper transcription factor), and RUP1/2 (two UVR8-interacting proteins). Here we report that Arabidopsis SALT TOLERANCE (STO/BBX24), a known regulator for light signaling in plants, defines a new signaling component in UV-B-mediated photomorphogenesis. The bbx24 mutant is hypersensitive to UV-B radiation and becomes extremely dwarfed under UV-B treatment. By contrast, BBX24 overexpression transgenic lines respond much more weakly to UV-B than the bbx24 and wild-type plants. BBX24 expression is UV-B-inducible and its accumulation under UV-B requires COP1. Co-immunoprecipitation experiments indicate that BBX24 interacts with COP1 in planta upon UV-B illumination. Moreover, BBX24 interacts with HY5 and acts antagonistically with HY5 in UV-B-induced inhibition of hypocotyl elongation. Furthermore, BBX24 attenuates UV-B-induced HY5 accumulation and suppresses its transcription-activation activity. Taken together, our results reveal a previously uncharacterized function of the light-regulated BBX24 in UV-B responses and demonstrate that BBX24 functions as a negative regulator of photomorphogenic UV-B responses by interacting with both COP1 and HY5. The UV-B-inducible expression pattern and its suppression of HY5 activity suggest that BBX24 could be a new component of the feedback regulatory module of UV-B signaling in plants.

  17. CRTR-1, a developmentally regulated transcriptional repressor related to the CP2 family of transcription factors.

    Science.gov (United States)

    Rodda, S; Sharma, S; Scherer, M; Chapman, G; Rathjen, P

    2001-02-02

    CP2-related proteins comprise a family of DNA-binding transcription factors that are generally activators of transcription and expressed ubiquitously. We reported a differential display polymerase chain reaction fragment, Psc2, which was expressed in a regulated fashion in mouse pluripotent cells in vitro and in vivo. Here, we report further characterization of the Psc2 cDNA and function. The Psc2 cDNA contained an open reading frame homologous to CP2 family proteins. Regions implicated in DNA binding and oligomeric complex formation, but not transcription activation, were conserved. Psc2 expression in vivo during embryogenesis and in the adult mouse demonstrated tight spatial and temporal regulation, with the highest levels of expression in the epithelial lining of distal convoluted tubules in embryonic and adult kidneys. Functional analysis demonstrated that PSC2 repressed transcription 2.5-15-fold when bound to a heterologous promoter in ES, 293T, and COS-1 cells. The N-terminal 52 amino acids of PSC2 were shown to be necessary and sufficient for this activity and did not share obvious homology with reported repressor motifs. These results represent the first report of a CP2 family member that is expressed in a developmentally regulated fashion in vivo and that acts as a direct repressor of transcription. Accordingly, the protein has been named CP2-Related Transcriptional Repressor-1 (CRTR-1).

  18. Transcriptional activation and cell cycle block are the keys for 5-fluorouracil induced up-regulation of human thymidylate synthase expression.

    Directory of Open Access Journals (Sweden)

    Alessio Ligabue

    Full Text Available BACKGROUND: 5-fluorouracil, a commonly used chemotherapeutic agent, up-regulates expression of human thymidylate synthase (hTS. Several different regulatory mechanisms have been proposed to mediate this up-regulation in distinct cell lines, but their specific contributions in a single cell line have not been investigated to date. We have established the relative contributions of these previously proposed regulatory mechanisms in the ovarian cancer cell line 2008 and the corresponding cisplatin-resistant and 5-FU cross-resistant-subline C13*. METHODOLOGY/PRINCIPAL FINDINGS: Using RNA polymerase II inhibitor DRB treated cell cultures, we showed that 70-80% of up-regulation of hTS results from transcriptional activation of TYMS mRNA. Moreover, we report that 5-FU compromises the cell cycle by blocking the 2008 and C13* cell lines in the S phase. As previous work has established that TYMS mRNA is synthesized in the S and G(1 phase and hTS is localized in the nuclei during S and G(2-M phase, the observed cell cycle changes are also expected to affect the intracellular regulation of hTS. Our data also suggest that the inhibition of the catalytic activity of hTS and the up-regulation of the hTS protein level are not causally linked, as the inactivated ternary complex, formed by hTS, deoxyuridine monophosphate and methylenetetrahydrofolate, was detected already 3 hours after 5-FU exposure, whereas substantial increase in global TS levels was detected only after 24 hours. CONCLUSIONS/SIGNIFICANCE: Altogether, our data indicate that constitutive TYMS mRNA transcription, cell cycle-induced hTS regulation and hTS enzyme stability are the three key mechanisms responsible for 5-fluorouracil induced up-regulation of human thymidylate synthase expression in the two ovarian cancer cell lines studied. As these three independent regulatory phenomena occur in a precise order, our work provides a feasible rationale for earlier observed synergistic combinations of 5

  19. Regulation of PES1 on estrogen receptor transcriptional activity%PES1对雌激素受体转录活性的调节作用

    Institute of Scientific and Technical Information of China (English)

    李杰萍; 庄庆仁; 兰小鹏; 曾国彬; 罗小锋

    2012-01-01

    目的 探讨PES1与雌激素受体(ER)的相互作用及其对ER转录活性的影响.方法 将体外翻译的PES1与纯化的GST-ERα和GST-ERβ蛋白分别混合,用GST pull-down验证在体外PES1与ERα和ERβ是否存在相互作用.将HA-PES1与FLAG-ERα或FLAGC-ERβ共转染293T细胞后进行免疫共沉淀,以验证PES1与ER是否在体内有相互作用.用含雌激素受体作用元件的荧光素酶报告基因检测PES1对ERα和ERβ转录活性的影响.结果 PES1与ERα、ERβ在体内外均存在相互作用,而且PES1与ERα的结合比与ERβ的强.在体内,在雌激素(E2)存在下,E2可以增强PES1与ERα的结合,而对PES1与ERβ的结合没有明显影响.PES1对ER转录活性的影响是E2依赖性的,PES1能升高ERα的转录活性而降低ERβ的转录活性(P<0.01).结论 PES1是一种新的ER共调节因子,能反向调节ERα和ERβ的转录活性,需要进一步研究的是其在ER信号通路以及在E2诱发的肿瘤发生发展中的作用.%Objective To study the intERαction of PES1 with estrogen receptor(ER) and its effect on the transcriptional activity of ER. Methods GST pull-down was used to verify the intERαction of PES1 with Erα and Erβ in vitro when PES1, which was previously translated, was mixed with the purified GST-Erα or GST-Erβ. The intERαction of PES1 with ER in vivo was tested by co-immunoprecipitation (co-IP) after HA-PES1 been co-transfected with FLAG-Erα or FLAGC-Erβ in 293T cells. The effects of PES1 on Ers' transcriptional activities were analyzed with estrogen receptor element lucifERαse ( ERE-LUC). Results PES1 intERαcted with both Erα and Erβ. Compared to ER α, PES1 binded Erβ strongerly E2 enhanced the binding of PES1 and Erα but had no effect on that of PES1 and Erβ. PES1 regulated the transcriptional activity of both Erα and Erβ in an E2-denpent manner PES1 raised the transcriptional activity of Erα but inhibited the transcriptional activity of Erβ. Conclusions PES1 may be a new

  20. Genetic manipulation of a transcription-regulating sequence of porcine reproductive and respiratory syndrome virus reveals key nucleotides determining its activity.

    Science.gov (United States)

    Zheng, Haihong; Zhang, Keyu; Zhu, Xing-Quan; Liu, Changlong; Lu, Jiaqi; Gao, Fei; Zhou, Yan; Zheng, Hao; Lin, Tao; Li, Liwei; Tong, Guangzhi; Wei, Zuzhang; Yuan, Shishan

    2014-08-01

    The factors that determine the transcription-regulating sequence (TRS) activity of porcine reproductive and respiratory syndrome virus (PRRSV) remain largely unclear. In this study, the effect of mutagenesis of conserved C nucleotides at positions 5 and 6 in the leader TRS (TRS-L) and/or canonical body TRS7 (TRS-B7) on the synthesis of subgenomic (sg) mRNA and virus infectivity was investigated in the context of a type 2 PRRSV infectious cDNA clone. The results showed that a double C mutation in the leader TRS completely abolished sg mRNAs synthesis and virus infectivity, but a single C mutation did not. A single C or double C mutation in TRS-B7.1 or/and TRS-B7.2 impaired or abolished the corresponding sg mRNA synthesis. Introduction of identical mutations in the leader and body TRSs partially restored sg mRNA7.1 and/or sg mRNA7.2 transcription, indicating that the base-pairing interaction between sense TRS-L and cTRS-B is a crucial factor influencing sg mRNA synthesis. Analysis of the mRNA leader-body junctions of mutants provided evidence for a mechanism of discontinuous minus-strand transcription. This study also showed that mutational inactivation of TRS-B7.1 or TRS-B7.2 did not affect the production of infectious progeny virus, and the sg mRNA formed from each of them could express N protein. However, TRS-B7.1 plays more important roles than TRS-B7.2 in maintaining the growth characteristic of type 2 PRRSV. These results provide more insight into the molecular mechanism of genome expression and subgenomic mRNA transcription of PRRSV.

  1. Peroxisome proliferator-activated receptor gamma (PPARγ) in yellow catfish Pelteobagrus fulvidraco: molecular characterization, mRNA expression and transcriptional regulation by insulin in vivo and in vitro.

    Science.gov (United States)

    Zheng, Jia-Lang; Zhuo, Mei-Qin; Luo, Zhi; Pan, Ya-Xiong; Song, Yu-Feng; Huang, Chao; Zhu, Qing-Ling; Hu, Wei; Chen, Qi-Liang

    2015-02-01

    Peroxisome proliferator-activated receptor gamma (PPARγ) is ligand-inducible transcription factor and has important roles in lipid metabolism, cell proliferation and inflammation. In the present study, yellow catfish Pelteobagrus fulvidraco PPARγ cDNA was isolated from liver by RT-PCR and RACE, and its molecular characterization and transcriptional regulation by insulin in vivo and in vitro were determined. The generation of PPARγ1 and PPARγ2 was due to alternative promoter of PPARγ gene. PPARγ1 and PPARγ2 mRNA covered 2426 bp and 2537 bp, respectively, with an open reading frame (ORF) of 1584 bp encoding 527 amino acid residues. Yellow catfish PPARγ gene was organized in a manner similar to that of their mammalian homologs, implying a modular organization of the protein's domains. A comparison between the yellow catfish PPARγ amino acid sequence and the correspondent sequences of several other species revealed the identity of 55-76.2%. Two PPARγ transcripts (PPARγ1 and PPARγ2) mRNAs were expressed in a wide range of tissues, but the abundance of each PPARγ mRNA showed the tissue- and developmental stage-dependent expression patterns. Intraperitoneal injection of insulin in vivo significantly stimulated the mRNA expression of total PPARγ and PPARγ1, but not PPARγ2 in the liver of yellow catfish. In contrast, incubation of hepatocytes with insulin in vitro increased the mRNA levels of PPARγ1, PPARγ2 and total PPARγ. To our knowledge, for the first time, the present study provides evidence that PPARγ1 and PPARγ2 are differentially expressed with and among tissues during different developmental stages and also regulated by insulin both in vivo and in vitro, which serves to increase our understanding on PPARγ physiological function in fish.

  2. Genome-Wide Binding Analysis of the Transcription Activator IDEAL PLANT ARCHITECTURE1 Reveals a Complex Network Regulating Rice Plant Architecture[W

    Science.gov (United States)

    Lu, Zefu; Yu, Hong; Xiong, Guosheng; Wang, Jing; Jiao, Yongqing; Liu, Guifu; Jing, Yanhui; Meng, Xiangbing; Hu, Xingming; Qian, Qian; Fu, Xiangdong; Wang, Yonghong; Li, Jiayang

    2013-01-01

    IDEAL PLANT ARCHITECTURE1 (IPA1) is critical in regulating rice (Oryza sativa) plant architecture and substantially enhances grain yield. To elucidate its molecular basis, we first confirmed IPA1 as a functional transcription activator and then identified 1067 and 2185 genes associated with IPA1 binding sites in shoot apices and young panicles, respectively, through chromatin immunoprecipitation sequencing assays. The SQUAMOSA PROMOTER BINDING PROTEIN-box direct binding core motif GTAC was highly enriched in IPA1 binding peaks; interestingly, a previously uncharacterized indirect binding motif TGGGCC/T was found to be significantly enriched through the interaction of IPA1 with proliferating cell nuclear antigen PROMOTER BINDING FACTOR1 or PROMOTER BINDING FACTOR2. Genome-wide expression profiling by RNA sequencing revealed IPA1 roles in diverse pathways. Moreover, our results demonstrated that IPA1 could directly bind to the promoter of rice TEOSINTE BRANCHED1, a negative regulator of tiller bud outgrowth, to suppress rice tillering, and directly and positively regulate DENSE AND ERECT PANICLE1, an important gene regulating panicle architecture, to influence plant height and panicle length. The elucidation of target genes of IPA1 genome-wide will contribute to understanding the molecular mechanisms underlying plant architecture and to facilitating the breeding of elite varieties with ideal plant architecture. PMID:24170127

  3. Genome-wide binding analysis of the transcription activator ideal plant architecture1 reveals a complex network regulating rice plant architecture.

    Science.gov (United States)

    Lu, Zefu; Yu, Hong; Xiong, Guosheng; Wang, Jing; Jiao, Yongqing; Liu, Guifu; Jing, Yanhui; Meng, Xiangbing; Hu, Xingming; Qian, Qian; Fu, Xiangdong; Wang, Yonghong; Li, Jiayang

    2013-10-01

    Ideal plant architecture1 (IPA1) is critical in regulating rice (Oryza sativa) plant architecture and substantially enhances grain yield. To elucidate its molecular basis, we first confirmed IPA1 as a functional transcription activator and then identified 1067 and 2185 genes associated with IPA1 binding sites in shoot apices and young panicles, respectively, through chromatin immunoprecipitation sequencing assays. The Squamosa promoter binding protein-box direct binding core motif GTAC was highly enriched in IPA1 binding peaks; interestingly, a previously uncharacterized indirect binding motif TGGGCC/T was found to be significantly enriched through the interaction of IPA1 with proliferating cell nuclear antigen promoter binding factor1 or promoter binding factor2. Genome-wide expression profiling by RNA sequencing revealed IPA1 roles in diverse pathways. Moreover, our results demonstrated that IPA1 could directly bind to the promoter of rice teosinte branched1, a negative regulator of tiller bud outgrowth, to suppress rice tillering, and directly and positively regulate dense and erect panicle1, an important gene regulating panicle architecture, to influence plant height and panicle length. The elucidation of target genes of IPA1 genome-wide will contribute to understanding the molecular mechanisms underlying plant architecture and to facilitating the breeding of elite varieties with ideal plant architecture.

  4. Bacillus subtilis response regulator DegU is a direct activator of pgsB transcription involved in gamma-poly-glutamic acid synthesis.

    Science.gov (United States)

    Ohsawa, Taku; Tsukahara, Kensuke; Ogura, Mitsuo

    2009-09-01

    pgsB encodes gamma-poly glutamic acid (gamma-PGA) synthetase and constitutes an operon with pgsC, pgsAA, and pgsE. Genetic analysis revealed that degQ and swrA, the known regulators of pgsB, are not required for pgsB expression when high cellular concentrations of phosphorylated form of the response regulator DegU (DegU-P) are present. However, swrA appeared still to be required for gamma-PGA synthesis under the conditions we tested. Since genetic analysis suggested that DegU-P activates pgsB directly, we performed gel retardation and footprint analyses using purified His-tagged DegU and the pgsB promoter. The in vitro experiments revealed that His-tagged DegU bound to the immediate upstream region of the -35 region of the pgsB promoter. A six-base deletion within the sequence (the -44 to -39 region) abolished DegU-binding to the pgsB promoter and pgsB transcription, confirming the importance of the sequence for DegU-dependent regulation of pgsB. Hence we conclude that DegU is a direct activator of the pgsB operon.

  5. pRB binds to and modulates the transrepressing activity of the E1A-regulated transcription factor p120E4F

    NARCIS (Netherlands)

    Fajas, L.; Paul, C.; Zugasti, O.; Cam, L. Le; Polanowska, J.; Fabbrizio, E.; Medema, R.H.; Vignais, M.-L.; Sardet, C.

    2000-01-01

    The retinoblastoma protein pRB is involved in the transcriptional control of genes essential for cell cycle progression and differentiation. pRB interacts with different transcription factors and thereby modulates their activity by sequestration, corepression, or activation. We report that pRB, but

  6. Sry is a transcriptional activator.

    Science.gov (United States)

    Dubin, R A; Ostrer, H

    1994-09-01

    The SRY gene functions as a genetic switch in gonadal ridge initiating testis determination. The mouse Sry and human SRY open reading frames (ORFs) share a conserved DNA-binding domain (the HMG-box) yet exhibit no additional homology outside this region. As judged by the accumulation of lacZ-SRY hybrid proteins in the nucleus, both the human and mouse SRY ORFs contain a nuclear localization signal. The mouse Sry HMG-box domain selectively binds the sequence NACAAT in vitro when challenged with a random pool of oligonucleotides and binds AACAAT with the highest affinity. When put under the control of a heterologous promotor, the mouse Sry gene activated transcription of a reporter gene containing multiple copies of the AACAAT binding site. Activation was likewise observed for a GAL4-responsive reporter gene, when the mouse Sry gene was linked to the DNA-binding domain of GAL4. Using this system, the activation function was mapped to a glutamine/histidine-rich domain. In addition, LexA-mouse Sry fusion genes activated a LexA-responsive reporter gene in yeast. In contrast, a GAL4-human SRY fusion gene did not cause transcriptional activation. These studies suggest that both the human and the mouse SRY ORFs encode nuclear, DNA-binding proteins and that the mouse Sry ORF can function as a transcriptional activator with separable DNA-binding and activator domains.

  7. TRANSFAC: transcriptional regulation, from patterns to profiles.

    Science.gov (United States)

    Matys, V; Fricke, E; Geffers, R; Gössling, E; Haubrock, M; Hehl, R; Hornischer, K; Karas, D; Kel, A E; Kel-Margoulis, O V; Kloos, D-U; Land, S; Lewicki-Potapov, B; Michael, H; Münch, R; Reuter, I; Rotert, S; Saxel, H; Scheer, M; Thiele, S; Wingender, E

    2003-01-01

    The TRANSFAC database on eukaryotic transcriptional regulation, comprising data on transcription factors, their target genes and regulatory binding sites, has been extended and further developed, both in number of entries and in the scope and structure of the collected data. Structured fields for expression patterns have been introduced for transcription factors from human and mouse, using the CYTOMER database on anatomical structures and developmental stages. The functionality of Match, a tool for matrix-based search of transcription factor binding sites, has been enhanced. For instance, the program now comes along with a number of tissue-(or state-)specific profiles and new profiles can be created and modified with Match Profiler. The GENE table was extended and gained in importance, containing amongst others links to LocusLink, RefSeq and OMIM now. Further, (direct) links between factor and target gene on one hand and between gene and encoded factor on the other hand were introduced. The TRANSFAC public release is available at http://www.gene-regulation.com. For yeast an additional release including the latest data was made available separately as TRANSFAC Saccharomyces Module (TSM) at http://transfac.gbf.de. For CYTOMER free download versions are available at http://www.biobase.de:8080/index.html.

  8. Activating Transcription Factor 3 regulates in part the enhanced tumour cell cytotoxicity of the histone deacetylase inhibitor M344 and cisplatin in combination

    Directory of Open Access Journals (Sweden)

    St Germain Carly

    2010-09-01

    Full Text Available Abstract Background Activating Transcription Factor (ATF 3 is a key regulator of the cellular integrated stress response whose expression has also been correlated with pro-apoptotic activities in tumour cell models. Combination treatments with chemotherapeutic drugs, such as cisplatin, and histone deacetylase (HDAC inhibitors have been demonstrated to enhance tumour cell cytotoxicity. We recently demonstrated a role for ATF3 in regulating cisplatin-induced apoptosis and others have shown that HDAC inhibition can also induce cellular stress. In this study, we evaluated the role of ATF3 in regulating the co-operative cytotoxicity of cisplatin in combination with an HDAC inhibitor. Results The HDAC inhibitor M344 induced ATF3 expression at the protein and mRNA level in a panel of human derived cancer cell lines as determined by Western blot and quantitative RT-PCR analyses. Combination treatment with M344 and cisplatin lead to increased induction of ATF3 compared with cisplatin alone. Utilizing the MTT cell viability assay, M344 treatments also enhanced the cytotoxic effects of cisplatin in these cancer cell lines. The mechanism of ATF3 induction by M344 was found to be independent of MAPKinase pathways and dependent on ATF4, a known regulator of ATF3 expression. ATF4 heterozygote (+/- and knock out (-/- mouse embryonic fibroblast (MEF as well as chromatin immunoprecipitation (ChIP assays were utilized in determining the mechanistic induction of ATF3 by M344. We also demonstrated that ATF3 regulates the enhanced cytotoxicity of M344 in combination with cisplatin as evidenced by attenuation of cytotoxicity in shRNAs targeting ATF3 expressing cells. Conclusion This study identifies the pro-apoptotic factor, ATF3 as a novel target of M344, as well as a mediator of the co-operative effects of cisplatin and M344 induced tumour cell cytotoxicity.

  9. RNA-guided transcriptional regulation in planta via synthetic dCas9-based transcription factors

    KAUST Repository

    Piatek, Agnieszka Anna

    2014-11-14

    Targeted genomic regulation is a powerful approach to accelerate trait discovery and development in agricultural biotechnology. Bacteria and archaea use clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) regulatory systems for adaptive molecular immunity against foreign nucleic acids introduced by invading phages and conjugative plasmids. The type II CRISPR/Cas system has been adapted for genome editing in many cell types and organisms. A recent study used the catalytically inactive Cas9 (dCas9) protein combined with guide-RNAs (gRNAs) as a DNA-targeting platform to modulate gene expression in bacterial, yeast, and human cells. Here, we modified this DNA-targeting platform for targeted transcriptional regulation in planta by developing chimeric dCas9-based transcriptional activators and repressors. To generate transcriptional activators, we fused the dCas9 C-terminus with the activation domains of EDLL and TAL effectors. To generate a transcriptional repressor, we fused the dCas9 C-terminus with the SRDX repression domain. Our data demonstrate that dCas9 fusion with the EDLL activation domain (dCas9:EDLL) and the TAL activation domain (dCas9:TAD), guided by gRNAs complementary to selected promoter elements, induce strong transcriptional activation on Bs3

  10. Transcriptional and chromatin regulation during fasting – The genomic era

    Science.gov (United States)

    Goldstein, Ido; Hager, Gordon L.

    2015-01-01

    An elaborate metabolic response to fasting is orchestrated by the liver and is heavily reliant upon transcriptional regulation. In response to hormones (glucagon, glucocorticoids) many transcription factors (TFs) are activated and regulate various genes involved in metabolic pathways aimed at restoring homeostasis: gluconeogenesis, fatty acid oxidation, ketogenesis and amino acid shuttling. We summarize the recent discoveries regarding fasting-related TFs with an emphasis on genome-wide binding patterns. Collectively, the summarized findings reveal a large degree of co-operation between TFs during fasting which occurs at motif-rich DNA sites bound by a combination of TFs. These new findings implicate transcriptional and chromatin regulation as major determinants of the response to fasting and unravels the complex, multi-TF nature of this response. PMID:26520657

  11. Transcriptionally regulated, prostate-targeted gene therapy for prostate cancer.

    Science.gov (United States)

    Lu, Yi

    2009-07-02

    Prostate cancer is the most frequently diagnosed cancer and the second leading cause of cancer deaths in American males today. Novel and effective treatment such as gene therapy is greatly desired. The early viral based gene therapy uses tissue-nonspecific promoters, which causes unintended toxicity to other normal tissues. In this chapter, we will review the transcriptionally regulated gene therapy strategy for prostate cancer treatment. We will describe the development of transcriptionally regulated prostate cancer gene therapy in the following areas: (1) Comparison of different routes for best viral delivery to the prostate; (2) Study of transcriptionally regulated, prostate-targeted viral vectors: specificity and activity of the transgene under several different prostate-specific promoters were compared in vitro and in vivo; (3) Selection of therapeutic transgenes and strategies for prostate cancer gene therapy (4) Oncolytic virotherapy for prostate cancer. In addition, the current challenges and future directions in this field are also discussed.

  12. The response regulator SsrB activates transcription and binds to a region overlapping OmpR binding sites at Salmonella pathogenicity island 2.

    Science.gov (United States)

    Feng, Xiuhong; Walthers, Don; Oropeza, Ricardo; Kenney, Linda J

    2004-11-01

    OmpR activates expression of the two-component regulatory system located on Salmonella pathogenicity island 2 (SPI-2) that controls the expression of a type III secretion system, as well as many other genes required for systemic infection in mice. Measurements of SsrA and SsrB protein levels under different growth conditions indicate that expression of these two components is uncoupled, i.e. SsrB is produced in the absence of ssrA and vice versa. This result was suggested from our previous studies, in which two promoters at ssrA/B were identified. The isolated C-terminus of SsrB binds to DNA and protects regions upstream of ssrA, ssrB and srfH from DNase I digestion. Furthermore, the C-terminus of SsrB alone is capable of activating transcription in the absence of the N-terminus. Results from beta-galactosidase assays indicate that the N-terminal phosphorylation domain inhibits the C-terminal effector domain. A previous study from our laboratory reported that ssrA-lacZ and ssrB-lacZ transcriptional fusions were substantially reduced in an ssrB null strain. Results from DNase I protection assays provide direct evidence that SsrB binds at ssrA and ssrB, although the binding sites lie within the transcribed regions. Additional regulators clearly affect gene expression at this important locus, and here we provide evidence that SlyA, a transcription factor that contributes to Salmonella virulence, also affects ssrA/B gene expression.

  13. Gene expression and metabolite profiling of developing highbush blueberry fruit indicates transcriptional regulation of flavonoid metabolism and activation of abscisic acid metabolism.

    Science.gov (United States)

    Zifkin, Michael; Jin, Alena; Ozga, Jocelyn A; Zaharia, L Irina; Schernthaner, Johann P; Gesell, Andreas; Abrams, Suzanne R; Kennedy, James A; Constabel, C Peter

    2012-01-01

    Highbush blueberry (Vaccinium corymbosum) fruits contain substantial quantities of flavonoids, which are implicated in a wide range of health benefits. Although the flavonoid constituents of ripe blueberries are known, the molecular genetics underlying their biosynthesis, localization, and changes that occur during development have not been investigated. Two expressed sequence tag libraries from ripening blueberry fruit were constructed as a resource for gene identification and quantitative real-time reverse transcription-polymerase chain reaction primer design. Gene expression profiling by quantitative real-time reverse transcription-polymerase chain reaction showed that flavonoid biosynthetic transcript abundance followed a tightly regulated biphasic pattern, and transcript profiles were consistent with the abundance of the three major classes of flavonoids. Proanthocyanidins (PAs) and corresponding biosynthetic transcripts encoding anthocyanidin reductase and leucoanthocyanidin reductase were most concentrated in young fruit and localized predominantly to the inner fruit tissue containing the seeds and placentae. Mean PA polymer length was seven to 8.5 subunits, linked predominantly via B-type linkages, and was relatively constant throughout development. Flavonol accumulation and localization patterns were similar to those of the PAs, and the B-ring hydroxylation pattern of both was correlated with flavonoid-3'-hydroxylase transcript abundance. By contrast, anthocyanins accumulated late in maturation, which coincided with a peak in flavonoid-3-O-glycosyltransferase and flavonoid-3'5'-hydroxylase transcripts. Transcripts of VcMYBPA1, which likely encodes an R2R3-MYB transcriptional regulator of PA synthesis, were prominent in both phases of development. Furthermore, the initiation of ripening was accompanied by a substantial rise in abscisic acid, a growth regulator that may be an important component of the ripening process and contribute to the regulation of

  14. The transcriptional activators AraR and XlnR from Aspergillus niger regulate expression of pentose catabolic and pentose phosphate pathway genes.

    Science.gov (United States)

    Battaglia, Evy; Zhou, Miaomiao; de Vries, Ronald P

    2014-09-01

    The pentose catabolic pathway (PCP) and the pentose phosphate pathway (PPP) are required for the conversion of pentose sugars in fungi and are linked via d-xylulose-5-phosphate. Previously, it was shown that the PCP is regulated by the transcriptional activators XlnR and AraR in Aspergillus niger. Here we assessed whether XlnR and AraR also regulate the PPP. Expression of two genes, rpiA and talB, was reduced in the ΔaraR/ΔxlnR strain and increased in the xylulokinase negative strain (xkiA1) on d-xylose and/or l-arabinose. Bioinformatic analysis of the 1 kb promoter regions of rpiA and talB showed the presence of putative XlnR binding sites. Combining all results in this study, it strongly suggests that these two PPP genes are under regulation of XlnR in A. niger.

  15. Transcriptional Auto-Regulation of RUNX1 P1 Promoter.

    Directory of Open Access Journals (Sweden)

    Milka Martinez

    Full Text Available RUNX1 a member of the family of runt related transcription factors (RUNX, is essential for hematopoiesis. The expression of RUNX1 gene is controlled by two promoters; the distal P1 promoter and the proximal P2 promoter. Several isoforms of RUNX1 mRNA are generated through the use of both promoters and alternative splicing. These isoforms not only differs in their temporal expression pattern but also exhibit differences in tissue specificity. The RUNX1 isoforms derived from P2 are expressed in a variety of tissues, but expression of P1-derived isoform is restricted to cells of hematopoietic lineage. However, the control of hematopoietic-cell specific expression is poorly understood. Here we report regulation of P1-derived RUNX1 mRNA by RUNX1 protein. In silico analysis of P1 promoter revealed presence of two evolutionary conserved RUNX motifs, 0.6kb upstream of the transcription start site, and three RUNX motifs within 170bp of the 5'UTR. Transcriptional contribution of these RUNX motifs was studied in myeloid and T-cells. RUNX1 genomic fragment containing all sites show very low basal activity in both cell types. Mutation or deletion of RUNX motifs in the UTR enhances basal activity of the RUNX1 promoter. Chromatin immunoprecipitation revealed that RUNX1 protein is recruited to these sites. Overexpression of RUNX1 in non-hematopoietic cells results in a dose dependent activation of the RUNX1 P1 promoter. We also demonstrate that RUNX1 protein regulates transcription of endogenous RUNX1 mRNA in T-cell. Finally we show that SCL transcription factor is recruited to regions containing RUNX motifs in the promoter and the UTR and regulates activity of the RUNX1 P1 promoter in vitro. Thus, multiple lines of evidence show that RUNX1 protein regulates its own gene transcription.

  16. Transcriptional Auto-Regulation of RUNX1 P1 Promoter.

    Science.gov (United States)

    Martinez, Milka; Hinojosa, Marcela; Trombly, Daniel; Morin, Violeta; Stein, Janet; Stein, Gary; Javed, Amjad; Gutierrez, Soraya E

    2016-01-01

    RUNX1 a member of the family of runt related transcription factors (RUNX), is essential for hematopoiesis. The expression of RUNX1 gene is controlled by two promoters; the distal P1 promoter and the proximal P2 promoter. Several isoforms of RUNX1 mRNA are generated through the use of both promoters and alternative splicing. These isoforms not only differs in their temporal expression pattern but also exhibit differences in tissue specificity. The RUNX1 isoforms derived from P2 are expressed in a variety of tissues, but expression of P1-derived isoform is restricted to cells of hematopoietic lineage. However, the control of hematopoietic-cell specific expression is poorly understood. Here we report regulation of P1-derived RUNX1 mRNA by RUNX1 protein. In silico analysis of P1 promoter revealed presence of two evolutionary conserved RUNX motifs, 0.6kb upstream of the transcription start site, and three RUNX motifs within 170bp of the 5'UTR. Transcriptional contribution of these RUNX motifs was studied in myeloid and T-cells. RUNX1 genomic fragment containing all sites show very low basal activity in both cell types. Mutation or deletion of RUNX motifs in the UTR enhances basal activity of the RUNX1 promoter. Chromatin immunoprecipitation revealed that RUNX1 protein is recruited to these sites. Overexpression of RUNX1 in non-hematopoietic cells results in a dose dependent activation of the RUNX1 P1 promoter. We also demonstrate that RUNX1 protein regulates transcription of endogenous RUNX1 mRNA in T-cell. Finally we show that SCL transcription factor is recruited to regions containing RUNX motifs in the promoter and the UTR and regulates activity of the RUNX1 P1 promoter in vitro. Thus, multiple lines of evidence show that RUNX1 protein regulates its own gene transcription.

  17. Role of Activator Protein-1 in the Transcription of Interleukin-5 Gene Regulated by Protein Kinase C Signal in Asthmatic Human T Lymphocytes

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    In order to explore the role of activator protein-1 (AP-1) in the transcription of interleukin-5 (IL-5) gene regulated by protein kinase C (PKC) signal in peripheral blood T lymphocytes from asthmatic patient, T lymphocytes were isolated and purified from peripheral blood of each asthmatic patient. The T lymphocytes were randomly divided int9 4 groups: group A (blank control), group B (treated with PKC agonist phorbol 12-myristate 13-acetate (PMA)), Group C (treated with PMA and AP-1 cis-element decoy oligodeoxynucleotides (decoy ODNs)), and group D (treated with PMA and AP-1 mutant decoy ODNs). The ODNs were transfected into the T cells of group C and D by cation liposome respectively. Reverse transcription-polymerase chain reaction (RT-PCR) was employed to assess IL-5 mRNA expression, and electrophoretic mobility shift assays (EMSA) for the activation of AP-1. The results showed that the activation of AP-1 (88 003.58±1 626.57) and the expression of IL5 mRNA (0. 8300±0. 0294) in T lymphocytes stimulated with PMA were significantly higher than these in blank control (20 888.47±1103.56 and 0. 3050±0. 0208, respectively, P< 0.01), while the indexes (23 219.83±1 024.86 and 0. 3425±0. 0171 respectively) of T lymphocytes stimulated with PMA and AP-1 decoy ODNs were significantly inhibited, as compared with group B (P<0.01). The indexes (87 107. 41±1 342.92 and 0. 8225±0. 0222, respectively) in T lymphocytes stimulated with PMA and AP-1 mutant decoy ODNs did not exhibit significant changes, as compared with group B (P>0.05). The significant positive correlation was found between the activation of AP-1 and the expression of IL-5 mRNA (P< 0.01). It was concluded that AP-1 might participate in the signal transduction of PKC-triggered transcription of IL-5 gene in asthmatic T lymphocytes. This suggests the activation of PKC/AP-1 signal transduction cascade of T lymphocytes may play an important role in the pathogenesis of asthma.

  18. Computational Analysis of the Transcriptional Regulation of the Actin Family

    Institute of Scientific and Technical Information of China (English)

    郑家顺; 吴加金; 孙之荣

    2002-01-01

    Transcriptional regulation is a very important regulatory step in the regulation of gene expression. Transcription factors (TFs) play an important role in controlling the temporal special specificity of gene expression. The regulation area of actin genes was analyzed statistically to predict the transcription factor binding sites in the regulatory area. A group of transcription factors located in most of the sequences is believed to play an important role in co-regulating the expression of actin genes.

  19. Repetitive Elements in Mycoplasma hyopneumoniae Transcriptional Regulation

    Science.gov (United States)

    Cattani, Amanda Malvessi; Siqueira, Franciele Maboni; Guedes, Rafael Lucas Muniz; Schrank, Irene Silveira

    2016-01-01

    Transcriptional regulation, a multiple-step process, is still poorly understood in the important pig pathogen Mycoplasma hyopneumoniae. Basic motifs like promoters and terminators have already been described, but no other cis-regulatory elements have been found. DNA repeat sequences have been shown to be an interesting potential source of cis-regulatory elements. In this work, a genome-wide search for tandem and palindromic repetitive elements was performed in the intergenic regions of all coding sequences from M. hyopneumoniae strain 7448. Computational analysis demonstrated the presence of 144 tandem repeats and 1,171 palindromic elements. The DNA repeat sequences were distributed within the 5’ upstream regions of 86% of transcriptional units of M. hyopneumoniae strain 7448. Comparative analysis between distinct repetitive sequences found in related mycoplasma genomes demonstrated different percentages of conservation among pathogenic and nonpathogenic strains. qPCR assays revealed differential expression among genes showing variable numbers of repetitive elements. In addition, repeats found in 206 genes already described to be differentially regulated under different culture conditions of M. hyopneumoniae strain 232 showed almost 80% conservation in relation to M. hyopneumoniae strain 7448 repeats. Altogether, these findings suggest a potential regulatory role of tandem and palindromic DNA repeats in the M. hyopneumoniae transcriptional profile. PMID:28005945

  20. Arabidopsis STO/BBX24 negatively regulates UV-B signaling by interacting with COP1 and repressing HY5 transcriptional activity

    Institute of Scientific and Technical Information of China (English)

    Lei Jiang; Yan Wang; Qian-Feng Li; Lars Olof Bj(o)rn; Jun-Xian He; Shao-Shan Li

    2012-01-01

    UV-B (280-315 nm) is an integral part of solar radiation and can act either as a stress inducer or as a developmental signal.In recent years,increasing attention has been paid to the Iow-fluence UV-B-induced photomorphogenic response and several key players in this response have been identified,which include UVR8 (a UV-B-specific photoreceptor),COPI (a WD40-repeat-containing RING finger protein),HY5 (a basic zipper transcription factor),and RUP1/2 (two UVR8-interacting proteins).Here we report that Arabidopsis SALT TOLERANCE (STO/BBX24),a known regulator for light signaling in plants,defines a new signaling component in UV-B-mediated photomorphogenesis.The bbx24 mutant is hypersensitive to UV-B radiation and becomes extremely dwarfed under UV-B treatment.By contrast,BBX24 overexpression transgenic lines respond much more weakly to UV-B than the bbx24 and wild-type plants.BBX24 expression is UV-B-inducible and its accumulation under UV-B requires COP1.Co-immunoprecipitation experiments indicate that BBX24 interacts with COP1 in planta upon UV-B illumination.Moreover,BBX24 interacts with HY5 and acts antagonistically with HY5 in UV-B-induced inhibition of hypocotyl elongation.Furthermore,BBX24 attenuates UV-B-induced HY5 accumulation and suppresses its transcription-activation activity.Taken together,our results reveal a previously uncharacterized function of the light-regulated BBX24 in UV-B responses and demonstrate that BBX24 functions as a negative regulator of photomorphogenic UV-B responses by interacting with both COP1 and HY5.The UV-B-inducible expression pattern and its suppression of HY5 activity suggest that BBX24 could be a new component of the feedback regulatory module of UV-B signaling in plants.

  1. Rad51 activates polyomavirus JC early transcription.

    Directory of Open Access Journals (Sweden)

    Martyn K White

    Full Text Available The human neurotropic polyomavirus JC (JCV causes the fatal CNS demyelinating disease progressive multifocal leukoencephalopathy (PML. JCV infection is very common and after primary infection, the virus is able to persist in an asymptomatic state. Rarely, and usually only under conditions of immune impairment, JCV re-emerges to actively replicate in the astrocytes and oligodendrocytes of the brain causing PML. The regulatory events involved in the reactivation of active viral replication in PML are not well understood but previous studies have implicated the transcription factor NF-κB acting at a well-characterized site in the JCV noncoding control region (NCCR. NF-κB in turn is regulated in a number of ways including activation by cytokines such as TNF-α, interactions with other transcription factors and epigenetic events involving protein acetylation--all of which can regulate the transcriptional activity of JCV. Active JCV infection is marked by the occurrence of rapid and extensive DNA damage in the host cell and the induction of the expression of cellular proteins involved in DNA repair including Rad51, a major component of the homologous recombination-directed double-strand break DNA repair machinery. Here we show that increased Rad51 expression activates the JCV early promoter. This activation is co-operative with the stimulation caused by NF-κB p65, abrogated by mutation of the NF-κB binding site or siRNA to NFκB p65 and enhanced by the histone deacetylase inhibitor sodium butyrate. These data indicate that the induction of Rad51 resulting from infection with JCV acts through NF-κB via its binding site to stimulate JCV early transcription. We suggest that this provides a novel positive feedback mechanism to enhance viral gene expression during the early stage of JCV infection.

  2. Computational Investigations of Post-Transcriptional Regulation

    DEFF Research Database (Denmark)

    Rasmussen, Simon Horskjær

    are the “switches” of combinatorial regulation. RBP hotspots are highly accessible AU-rich regions that are more frequently bound by RBPs and they are frequently in the vicinity of miRNA target sites. To further investigate this, an experimental design and analysis method, to further unravel combinatorial...... investigated using high-throughput data. Analysis of IMP RIP-seq, iCLIP and RNA-seq datasets identified transcripts associated with cytoplasmic IMP ribonucleoproteins. Many of these transcripts were functionally involved in actin cytoskeletal remodeling. Further analyses of this data permitted estimation...... of a bipartite motif, composed of an AU-rich and a CA-rich domain. In addition, a regulatory motif discovery method was developed and applied to identify motifs using differential expression data and CLIP-data in the above investigations. This thesis increased the understanding of the role of RBPs in miRNA...

  3. Computational Investigations of Post-Transcriptional Regulation

    DEFF Research Database (Denmark)

    Rasmussen, Simon Horskjær

    are the “switches” of combinatorial regulation. RBP hotspots are highly accessible AU-rich regions that are more frequently bound by RBPs and they are frequently in the vicinity of miRNA target sites. To further investigate this, an experimental design and analysis method, to further unravel combinatorial...... investigated using high-throughput data. Analysis of IMP RIP-seq, iCLIP and RNA-seq datasets identified transcripts associated with cytoplasmic IMP ribonucleoproteins. Many of these transcripts were functionally involved in actin cytoskeletal remodeling. Further analyses of this data permitted estimation...... of a bipartite motif, composed of an AU-rich and a CA-rich domain. In addition, a regulatory motif discovery method was developed and applied to identify motifs using differential expression data and CLIP-data in the above investigations. This thesis increased the understanding of the role of RBPs in mi...

  4. Suppressor of cytokine signaling 2 (SOCS2) negatively regulates the expression of antimicrobial peptides by affecting the Stat transcriptional activity in shrimp Marsupenaeus japonicus.

    Science.gov (United States)

    Sun, Jie-Jie; Lan, Jiang-Feng; Xu, Ji-Dong; Niu, Guo-Juan; Wang, Jin-Xing

    2016-09-01

    The suppressor of cytokine signaling (SOCS) family is a kind of negative regulators in the Janus kinase/signal transducer and activator of transcription (Jak/Stat) pathway in mammals and Drosophila. In kuruma shrimp, Marsupenaeus japonicus, SOCS2 is identified and its expression can be stimulated by peptidoglycan and polycytidylic acid. However, if SOCS2 participates in regulating Jak/Stat pathway in shrimp still needs further study. In this study, SOCS2 with Src homology 2 domain and SOCS box was identified in kuruma shrimp, M. japonicus. SOCS2 existed in hemocytes, heart, hepatopancreas, gills, stomach, and intestine, the expression of SOCS2 was upregulated significantly in the hemocytes and intestine of shrimp challenged with Vibrio anguillarum at 6 h. To analyze SOCS2 function in shrimp immunity, bacterial clearance and survival rate were analyzed after knockdown of SOCS2 in shrimp challenged with V. anguillarum. Results showed that bacterial clearance increased, and the survival rate improved significantly comparing with controls. The SOCS2 was expressed in Escherichia coli and the recombinant SOCS2 was injected into shrimp, and Stat phosphorylation and translocation were analyzed. The result showed that "overexpression" of SOCS2 declined Stat phosphorylation level and inhibited Stat translocation into the nucleus. After knockdown of SOCS2 in shrimp prior to V. anguillarum infection, the expression level of antimicrobial peptides, including anti-lipopolysaccharide factors C1, C2 and D1, and Crustin I was upregulated significantly, and the expression of the AMPs was declined after recombinant SOCS2 injection. The SOCS2 expression was also decreased in Stat-knockdown shrimp challenged by V. anguillarum at 6 and 12 h. Therefore, SOCS2 negatively regulates the AMP expression by inhibiting Stat phosphorylation and translocation into nucleus in shrimp, meanwhile, SOCS2 expression was also regulated by Jak/Stat pathway.

  5. How the ubiquitin proteasome system regulates the regulators of transcription.

    Science.gov (United States)

    Ee, Gary; Lehming, Norbert

    2012-01-01

    The ubiquitin proteasome system plays an important role in transcription. Monoubiquitination of activators is believed to aid their function, while the 26S proteasomal degradation of repressors is believed to restrict their function. What remains controversial is the question of whether the degradation of activators aids or restricts their function.

  6. Regulation of signal transducer and activator of transcription 3 and apoptotic pathways by betaine attenuates isoproterenol-induced acute myocardial injury in rats.

    Science.gov (United States)

    Zheng, P; Liu, J; Mai, S; Yuan, Y; Wang, Y; Dai, G

    2015-05-01

    The present study was designed to investigate the cardioprotective effects of betaine on acute myocardial ischemia induced experimentally in rats focusing on regulation of signal transducer and activator of transcription 3 (STAT3) and apoptotic pathways as the potential mechanism underlying the drug effect. Male Sprague Dawley rats were treated with betaine (100, 200, and 400 mg/kg) orally for 40 days. Acute myocardial ischemic injury was induced in rats by subcutaneous injection of isoproterenol (85 mg/kg), for two consecutive days. Serum cardiac marker enzyme, histopathological variables and expression of protein levels were analyzed. Oral administration of betaine (200 and 400 mg/kg) significantly reduced the level of cardiac marker enzyme in the serum and prevented left ventricular remodeling. Western blot analysis showed that isoproterenol-induced phosphorylation of STAT3 was maintained or further enhanced by betaine treatment in myocardium. Furthermore, betaine (200 and 400 mg/kg) treatment increased the ventricular expression of Bcl-2 and reduced the level of Bax, therefore causing a significant increase in the ratio of Bcl-2/Bax. The protective role of betaine on myocardial damage was further confirmed by histopathological examination. In summary, our results showed that betaine pretreatment attenuated isoproterenol-induced acute myocardial ischemia via the regulation of STAT3 and apoptotic pathways.

  7. Exploring the active site of acyl homoserine lactones-dependent transcriptional regulators with bacterial quorum sensing modulators using molecular mechanics and docking studies.

    Science.gov (United States)

    Soulère, Laurent; Frezza, Marine; Queneau, Yves; Doutheau, Alain

    2007-09-01

    A comparative molecular modelling study of acyl homoserine lactones-dependent transcriptional regulators (TraR, SdiA, LuxR and LasR) involved in bacterial quorum sensing (QS) revealed a high structural homology of their active site. Docking studies within the active site of TraR of fixed conformations obtained using molecular mechanics calculations showed that TraR, for which the crystalline structure is known, is a relevant model for the study of other protein-ligand interactions in the same protein family. Structure-activity relationships of AHLs derived QS modulators including carboxamides, sulfonamides and ureas were thus investigated. The results show that Tyr61, a residue conserved in the LuxR-proteins family, is involved in attractive interactions with aromatic carboxamide antagonists. Tyr53, Tyr61 and Asp70, conserved residues, are implicated in both the development of additional hydrogen bonds and attractive interactions with the N-sulfonyl homoserine lactones and AHLs derived ureas antagonists.

  8. Transcriptional regulation by Polycomb group proteins

    DEFF Research Database (Denmark)

    Di Croce, Luciano; Helin, Kristian

    2013-01-01

    Polycomb group (PcG) proteins are epigenetic regulators of transcription that have key roles in stem-cell identity, differentiation and disease. Mechanistically, they function within multiprotein complexes, called Polycomb repressive complexes (PRCs), which modify histones (and other proteins......) and silence target genes. The dynamics of PRC1 and PRC2 components has been the focus of recent research. Here we discuss our current knowledge of the PRC complexes, how they are targeted to chromatin and how the high diversity of the PcG proteins allows these complexes to influence cell identity....

  9. Activation of AP-1 Transcription Factors Differentiates FGF2 and Vascular Endothelial Growth Factor Regulation of Endothelial Nitric-oxide Synthase Expression in Placental Artery Endothelial Cells*

    Science.gov (United States)

    Mata-Greenwood, Eugenia; Liao, Wu-xiang; Wang, Wen; Zheng, Jing; Chen, Dong-bao

    2010-01-01

    FGF2 (fibroblast growth factor 2), but not vascular endothelial growth factor (VEGF), stimulates sustained activation of ERK2/1 for endothelial NOS3 (nitric-oxide synthase 3) protein expression in ovine fetoplacental artery endothelial cells (oFPAEC). We deciphered herein the downstream signaling of ERK2/1 responsible for NOS3 expression by FGF2 in oFPAEC. FGF2, but not VEGF, increased NOS3 mRNA levels without altering its degradation. FGF2, but not VEGF, trans-activated sheep NOS3 promoter, and this was dependent on ERK2/1 activation. FGF2 did not trans-activate NOS3 promoters with deletions upstream of the consensus AP-1 site (TGAGTC A, −678 to −685). Trans-activation of wild-type NOS3 promoter by FGF2 was significantly inhibited when either the AP-1 or the cAMP-response element (CRE)-like sequence (TGCGTCA, −752 to −758) was mutated and was completely blocked when both were mutated. EMSA analyses showed that FGF2, but not VEGF, stimulated AP-1 and CRE DNA-protein complexes primarily composed of JunB and Fra1. Chromatin immunoprecipitation assays confirmed JunB/Fra1 binding to NOS3 promoter AP-1 and CRE elements in intact cells. FGF2, but not VEGF, stimulated JunB and Fra1 expressions; all preceded NOS3 up-regulation and were inhibited by PD98059. Down-regulation of JunB or Fra-1, but not c-Jun, blocked FGF2 stimulation of NOS3 expression and NO production. AP-1 inhibition suppressed FGF2 stimulation of NOS3 expression in human umbilical vein EC and uterine artery endothelial cells. Thus, FGF2 induction of NOS3 expression is mainly mediated by AP-1-dependent transcription involving JunB and Fra1 up-regulation via sustained ERK2/1 activation in endothelial cells. PMID:20371606

  10. Transcription regulation by CHD proteins to control plant development

    Directory of Open Access Journals (Sweden)

    Yongfeng eHu

    2014-05-01

    Full Text Available CHD (Chromodomain-Helicase-DNA binding proteins have been characterized in various species as important transcription regulators by their chromatin remodeling activity. However, in plant the function of these proteins has hardly been analyzed before except that Arabidopsis PICKLE and rice CHR729 are identified to play critical roles in the regulation of series of genes involved in developmental or stress responding process. In this review we focus on how plant CHD proteins regulate gene expression and the role of these proteins in controlling plant development and stress response.

  11. Signal transducer and activator of transcription 3 (Stat3) regulates host defense and protects mice against herpes simplex virus-1 (HSV-1) infection.

    Science.gov (United States)

    Hsia, Hung-Ching; Stopford, Charles M; Zhang, Zhigang; Damania, Blossom; Baldwin, Albert S

    2016-12-13

    Signal transducer and activator of transcription 3 (STAT3) mediates cellular responses to multiple cytokines, governs gene expression, and regulates the development and activation of immune cells. STAT3 also modulates reactivation of latent herpes simplex virus-1 (HSV-1) in ganglia. However, it is unclear how STAT3 regulates the innate immune response during the early phase of HSV-1 lytic infection. Many cell types critical for the innate immunity are derived from the myeloid lineage. Therefore, in this study, we used myeloid-specific Stat3 knockout mice to investigate the role of STAT3 in the innate immune response against HSV-1. Our results demonstrate that Stat3 knockout bone marrow-derived macrophages (BMMs) expressed decreased levels of interferon-α (IFN-α) and interferon-stimulated genes (ISGs) upon HSV-1 infection. In vivo, knockout mice were more susceptible to HSV-1, as marked by higher viral loads and more significant weight loss. Splenic expression of IFN-α and ISGs was reduced in the absence of STAT3, indicating that STAT3 is required for optimal type I interferon response to HSV-1. Expression of TNF-α and IL-12, cytokines that have been shown to limit HSV-1 replication and pathogenesis, was also significantly lower in knockout mice. Interestingly, Stat3 knockout mice failed to expand the CD8(+) conventional DC (cDC) population upon HSV-1 infection, and this was accompanied by impaired NK and CD8 T cell activation. Collectively, our data demonstrate that myeloid-specific Stat3 deletion causes defects in multiple aspects of the immune system and that STAT3 has a protective role at the early stage of systemic HSV-1 infection.

  12. The ubiquitin ligase HectH9 regulates transcriptional activation by Myc and is essential for tumor cell proliferation

    DEFF Research Database (Denmark)

    Adhikary, Sovana; Marinoni, Federica; Hock, Andreas

    2005-01-01

    The Myc oncoprotein forms a binary activating complex with its partner protein, Max, and a ternary repressive complex that, in addition to Max, contains the zinc finger protein Miz1. Here we show that the E3 ubiquitin ligase HectH9 ubiquitinates Myc in vivo and in vitro, forming a lysine 63-linked...

  13. The cell cycle rallies the transcription cycle: Cdc28/Cdk1 is a cell cycle-regulated transcriptional CDK.

    Science.gov (United States)

    Chymkowitch, Pierre; Enserink, Jorrit M

    2013-01-01

    In the budding yeast Saccharomyces cerevisiae, the cyclin-dependent kinases (CDKs) Kin28, Bur1 and Ctk1 regulate basal transcription by phosphorylating the carboxyl-terminal domain (CTD) of RNA polymerase II. However, very little is known about the involvement of the cell cycle CDK Cdc28 in the transcription process. We have recently shown that, upon cell cycle entry, Cdc28 kinase activity boosts transcription of a subset of genes by directly stimulating the basal transcription machinery. Here, we discuss the biological significance of this finding and give our view of the kinase-dependent role of Cdc28 in regulation of RNA polymerase II.

  14. SIRT1 regulates HIV transcription via Tat deacetylation.

    Directory of Open Access Journals (Sweden)

    Sara Pagans

    2005-02-01

    Full Text Available The human immunodeficiency virus (HIV Tat protein is acetylated by the transcriptional coactivator p300, a necessary step in Tat-mediated transactivation. We report here that Tat is deacetylated by human sirtuin 1 (SIRT1, a nicotinamide adenine dinucleotide-dependent class III protein deacetylase in vitro and in vivo. Tat and SIRT1 coimmunoprecipitate and synergistically activate the HIV promoter. Conversely, knockdown of SIRT1 via small interfering RNAs or treatment with a novel small molecule inhibitor of the SIRT1 deacetylase activity inhibit Tat-mediated transactivation of the HIV long terminal repeat. Tat transactivation is defective in SIRT1-null mouse embryonic fibroblasts and can be rescued by expression of SIRT1. These results support a model in which cycles of Tat acetylation and deacetylation regulate HIV transcription. SIRT1 recycles Tat to its unacetylated form and acts as a transcriptional coactivator during Tat transactivation.

  15. Thermodynamics-based models of transcriptional regulation with gene sequence.

    Science.gov (United States)

    Wang, Shuqiang; Shen, Yanyan; Hu, Jinxing

    2015-12-01

    Quantitative models of gene regulatory activity have the potential to improve our mechanistic understanding of transcriptional regulation. However, the few models available today have been based on simplistic assumptions about the sequences being modeled or heuristic approximations of the underlying regulatory mechanisms. In this work, we have developed a thermodynamics-based model to predict gene expression driven by any DNA sequence. The proposed model relies on a continuous time, differential equation description of transcriptional dynamics. The sequence features of the promoter are exploited to derive the binding affinity which is derived based on statistical molecular thermodynamics. Experimental results show that the proposed model can effectively identify the activity levels of transcription factors and the regulatory parameters. Comparing with the previous models, the proposed model can reveal more biological sense.

  16. α-catenin is a tumor suppressor that controls cell accumulation by regulating the localization and activity of the transcriptional coactivator Yap1.

    Science.gov (United States)

    Silvis, Mark R; Kreger, Bridget T; Lien, Wen-Hui; Klezovitch, Olga; Rudakova, G Marianna; Camargo, Fernando D; Lantz, Dan M; Seykora, John T; Vasioukhin, Valeri

    2011-05-24

    The Hippo pathway regulates contact inhibition of cell proliferation and, ultimately, organ size in diverse multicellular organisms. Inactivation of the Hippo pathway promotes nuclear localization of the transcriptional coactivator Yap1, a Hippo pathway effector, and can cause cancer. Here, we show that deletion of αE (α epithelial) catenin in the hair follicle stem cell compartment resulted in the development of skin squamous cell carcinoma in mice. Tumor formation was accelerated by simultaneous deletion of αE-catenin and the tumor suppressor-encoding gene p53. A small interfering RNA screen revealed a functional connection between αE-catenin and Yap1. By interacting with Yap1, αE-catenin promoted its cytoplasmic localization, and Yap1 showed constitutive nuclear localization in αE-catenin-null cells. We also found an inverse correlation between αE-catenin abundance and Yap1 activation in human squamous cell carcinoma tumors. These findings identify αE-catenin as a tumor suppressor that inhibits Yap1 activity and sequesters it in the cytoplasm.

  17. IKAROS: a multifunctional regulator of the polymerase II transcription cycle.

    Science.gov (United States)

    Bottardi, Stefania; Mavoungou, Lionel; Milot, Eric

    2015-09-01

    Transcription factors are important determinants of lineage specification during hematopoiesis. They favor recruitment of cofactors involved in epigenetic regulation, thereby defining patterns of gene expression in a development- and lineage-specific manner. Additionally, transcription factors can facilitate transcription preinitiation complex (PIC) formation and assembly on chromatin. Interestingly, a few lineage-specific transcription factors, including IKAROS, also regulate transcription elongation. IKAROS is a tumor suppressor frequently inactivated in leukemia and associated with a poor prognosis. It forms a complex with the nucleosome remodeling and deacetylase (NuRD) complex and the positive transcription elongation factor b (P-TEFb), which is required for productive transcription elongation. It has also been reported that IKAROS interacts with factors involved in transcription termination. Here we review these and other recent findings that establish IKAROS as the first transcription factor found to act as a multifunctional regulator of the transcription cycle in hematopoietic cells.

  18. Activation of BmGSTd1 promoter and regulation by transcription factor Krüppel (Kr) in silkworm, Bombyx mori.

    Science.gov (United States)

    Zhao, Guodong; Wang, Binbin; Liu, Yunlei; Du, Jie; Li, Bing; Chen, Yuhua; Xu, Yaxiang; Shen, Weide; Xia, Qingyou; Wei, Zhengguo

    2014-11-10

    The Glutathione S-transferases (GSTs) are a large family of multifunctional enzymes, many of which play an important role in the detoxification of endogenous and exogenous toxic substances. In this research, firstly, we measured the rutin-induced transcriptional level of BmGSTd1 gene by using real-time quantitative RT-PCR method and dual spike-in strategy. The activities of the BmGSTd1 promoter in various tissues of silkworm were measured by firefly luciferase activity and normalized by the Renilla luciferase activity. Results showed that the activity of the BmGSTd1 promoter were highest in Malpighian tubule, followed by fat body, silk gland, hemocyte, epidermis, and midgut. The essential region for basal and rutin-induced transcriptional activity was -1573 to -931bp in Malpighian tubule and fat body of silkworm. Promoter truncation analysis using a dual-luciferase reporter assay in BmN cells showed that the region -1288 to -1202bp for BmGSTd1 gene was essential for basal and rutin-induced transcriptional activity. Sequence analysis of this region revealed several potential transcriptional regulatory elements such as Bcd and Kr. The mutation of core base of Kr site demonstrated that Kr functioned positively in rutin-mediated BmGSTd1 transcription.

  19. Repurposing CRISPR System for Transcriptional Activation.

    Science.gov (United States)

    Chen, Meng; Qi, Lei Stanley

    2017-01-01

    In recent years, Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system has become the most popular one for genome editing. When the nuclease domains of Cas9 protein are mutated into deactivated form (dCas9), CRISPR/dCas9 still retains the ability to bind the targeted DNA sequence, but loses the endonuclease cleavage activity. Taking advantage of the characteristics of this engineered nuclease inactive Cas9, the CRISPR/dCas system has been repurposed into versatile RNA-guided, DNA-targeting platforms, such as genome imaging, gene regulation, and epigenetic modification. Specifically, fusion of dCas9 with activation domains allows specific and efficient transcriptional activation on a genome-wide scale among diverse organisms. The purpose of this chapter is to review most important the recently published literature on CRISPR/dCas9-based transcriptional activation systems. Compared with the conventional approaches for enhancement of the expression of specific genes of interest, CRISPR/Cas9-based system has emerged as a promising technology for genome regulation, allowing specificity, convenience, robustness, and scalability for endogenous gene activation.

  20. Autopalmitoylation of TEAD Proteins Regulates Transcriptional Output of Hippo Pathway

    Science.gov (United States)

    Chan, PuiYee; Han, Xiao; Zheng, Baohui; DeRan, Michael; Yu, Jianzhong; Jarugumilli, Gopala K.; Deng, Hua; Pan, Duojia; Luo, Xuelian; Wu, Xu

    2016-01-01

    TEA domain (TEAD) transcription factors bind to the co-activator YAP/TAZ, and regulate the transcriptional output of Hippo pathway, playing critical roles in organ size control and tumorigenesis. Protein S-palmitoylation attaches fatty acid (palmitate) to cysteine residues, and regulates protein trafficking, membrane localization and signaling activities. Using activity-based chemical probes, we discovered that human TEADs possess intrinsic palmitoylating enzyme-like activities, and undergo autopalmitoylation at evolutionarily conserved cysteine residues under physiological conditions. We determined the crystal structures of lipid-bound TEADs, and found that the lipid chain of palmitate inserts into a conserved deep hydrophobic pocket. Strikingly, palmitoylation is required for TEAD’s binding to YAP/TAZ, but dispensable for the binding to Vgll4 tumor suppressor. In addition, palmitoylation does not alter TEAD’s localization. Moreover, TEAD palmitoylation-deficient mutants impaired TAZ-mediated muscle differentiation in vitro, and Yorkie-mediated tissue overgrowth in Drosophila in vivo. Our study directly linked autopalmitoylation to the transcriptional regulation of Hippo pathway. PMID:26900866

  1. Transcriptional regulation of the human mitochondrial peptide deformylase (PDF).

    Science.gov (United States)

    Pereira-Castro, Isabel; Costa, Luís Teixeira da; Amorim, António; Azevedo, Luisa

    2012-05-18

    The last years of research have been particularly dynamic in establishing the importance of peptide deformylase (PDF), a protein of the N-terminal methionine excision (NME) pathway that removes formyl-methionine from mitochondrial-encoded proteins. The genomic sequence of the human PDF gene is shared with the COG8 gene, which encodes a component of the oligomeric golgi complex, a very unusual case in Eukaryotic genomes. Since PDF is crucial in maintaining mitochondrial function and given the atypical short distance between the end of COG8 coding sequence and the PDF initiation codon, we investigated whether the regulation of the human PDF is affected by the COG8 overlapping partner. Our data reveals that PDF has several transcription start sites, the most important of which only 18 bp from the initiation codon. Furthermore, luciferase-activation assays using differently-sized fragments defined a 97 bp minimal promoter region for human PDF, which is capable of very strong transcriptional activity. This fragment contains a potential Sp1 binding site highly conserved in mammalian species. We show that this binding site, whose mutation significantly reduces transcription activation, is a target for the Sp1 transcription factor, and possibly of other members of the Sp family. Importantly, the entire minimal promoter region is located after the end of COG8's coding region, strongly suggesting that the human PDF preserves an independent regulation from its overlapping partner. Copyright © 2012 Elsevier Inc. All rights reserved.

  2. Plastid sigma factors: Their individual functions and regulation in transcription.

    Science.gov (United States)

    Chi, Wei; He, Baoye; Mao, Juan; Jiang, Jingjing; Zhang, Lixin

    2015-09-01

    Sigma factors are the predominant factors involved in transcription regulation in bacteria. These factors can recruit the core RNA polymerase to promoters with specific DNA sequences and initiate gene transcription. The plastids of higher plants originating from an ancestral cyanobacterial endosymbiont also contain sigma factors that are encoded by a small family of nuclear genes. Although all plastid sigma factors contain sequences conserved in bacterial sigma factors, a considerable number of distinct traits have been acquired during evolution. The present review summarises recent advances concerning the regulation of the structure, function and activity of plastid sigma factors since their discovery nearly 40 years ago. We highlight the specialised roles and overlapping redundant functions of plastid sigma factors according to their promoter selectivity. We also focus on the mechanisms that modulate the activity of sigma factors to optimise plastid function in response to developmental cues and environmental signals. This article is part of a Special Issue entitled: Chloroplast Biogenesis.

  3. Regulation of neural gene transcription by optogenetic inhibition of the RE1-silencing transcription factor.

    Science.gov (United States)

    Paonessa, Francesco; Criscuolo, Stefania; Sacchetti, Silvio; Amoroso, Davide; Scarongella, Helena; Pecoraro Bisogni, Federico; Carminati, Emanuele; Pruzzo, Giacomo; Maragliano, Luca; Cesca, Fabrizia; Benfenati, Fabio

    2016-01-01

    Optogenetics provides new ways to activate gene transcription; however, no attempts have been made as yet to modulate mammalian transcription factors. We report the light-mediated regulation of the repressor element 1 (RE1)-silencing transcription factor (REST), a master regulator of neural genes. To tune REST activity, we selected two protein domains that impair REST-DNA binding or recruitment of the cofactor mSin3a. Computational modeling guided the fusion of the inhibitory domains to the light-sensitive Avena sativa light-oxygen-voltage-sensing (LOV) 2-phototrophin 1 (AsLOV2). By expressing AsLOV2 chimeras in Neuro2a cells, we achieved light-dependent modulation of REST target genes that was associated with an improved neural differentiation. In primary neurons, light-mediated REST inhibition increased Na(+)-channel 1.2 and brain-derived neurotrophic factor transcription and boosted Na(+) currents and neuronal firing. This optogenetic approach allows the coordinated expression of a cluster of genes impinging on neuronal activity, providing a tool for studying neuronal physiology and correcting gene expression changes taking place in brain diseases.

  4. Loss of Interdependent Binding by the FoxO1 and FoxA1/A2 Forkhead Transcription Factors Culminates in Perturbation of Active Chromatin Marks and Binding of Transcriptional Regulators at Insulin-sensitive Genes.

    Science.gov (United States)

    Yalley, Akua; Schill, Daniel; Hatta, Mitsutoki; Johnson, Nicole; Cirillo, Lisa Ann

    2016-04-15

    FoxO1 binds to insulin response elements located in the promoters of insulin-like growth factor-binding protein 1 (IGFBP1) and glucose-6-phosphatase (G6Pase), activating their expression. Insulin-mediated phosphorylation of FoxO1 promotes cytoplasmic translocation, inhibiting FoxO1-mediated transactivation. We have previously demonstrated that FoxO1 opens and remodels chromatin assembled from the IGFBP1 promoter via a highly conserved winged helix motif. This finding, which established FoxO1 as a "pioneer" factor, suggested a model whereby FoxO1 chromatin remodeling at regulatory targets facilitates binding and recruitment of additional regulatory factors. However, the impact of FoxO1 phosphorylation on its ability to bind chromatin and the effect of FoxO1 loss on recruitment of neighboring transcription factors at its regulatory targets in liver chromatin is unknown. In this study, we demonstrate that an amino acid substitution that mimics insulin-mediated phosphorylation of a serine in the winged helix DNA binding motif curtails FoxO1 nucleosome binding. We also demonstrate that shRNA-mediated loss of FoxO1 binding to the IGFBP1 and G6Pase promoters in HepG2 cells significantly reduces binding of RNA polymerase II and the pioneer factors FoxA1/A2. Knockdown of FoxA1 similarly reduced binding of RNA polymerase II and FoxO1. Reduction in acetylation of histone H3 Lys-27 accompanies loss of FoxO1 and FoxA1/A2 binding. Interdependent binding of FoxO1 and FoxA1/A2 possibly entails cooperative binding because FoxO1 and FoxA1/A2 facilitate one another's binding to IGFPB1 promoter DNA. These results illustrate how transcription factors can nucleate transcriptional events in chromatin in response to signaling events and suggest a model for regulation of hepatic glucose metabolism through interdependent FoxO/FoxA binding.

  5. Transcriptional Regulation in Mammalian Cells by Sequence-Specific DNA Binding Proteins

    Science.gov (United States)

    Mitchell, Pamela J.; Tjian, Robert

    1989-07-01

    The cloning of genes encoding mammalian DNA binding transcription factors for RNA polymerase II has provided the opportunity to analyze the structure and function of these proteins. This review summarizes recent studies that define structural domains for DNA binding and transcriptional activation functions in sequence-specific transcription factors. The mechanisms by which these factors may activate transcriptional initiation and by which they may be regulated to achieve differential gene expression are also discussed.

  6. Harnessing the master transcriptional repressor REST to reciprocally regulate neurogenesis

    Science.gov (United States)

    Nesti, Edmund

    2015-01-01

    Neurogenesis begins in embryonic development and continues at a reduced rate into adulthood in vertebrate species, yet the signaling cascades regulating this process remain poorly understood. Plasma membrane-initiated signaling cascades regulate neurogenesis via downstream pathways including components of the transcriptional machinery. A nuclear factor that temporally regulates neurogenesis by repressing neuronal differentiation is the repressor element 1 (RE1) silencing transcription (REST) factor. We have recently discovered a regulatory site on REST that serves as a molecular switch for neuronal differentiation. Specifically, C-terminal domain small phosphatase 1, CTDSP1, present in non-neuronal cells, maintains REST activity by dephosphorylating this site. Reciprocally, extracellular signal-regulated kinase, ERK, activated by growth factor signaling in neural progenitors, and peptidylprolyl cis/trans isomerase Pin1, decrease REST activity through phosphorylation-dependent degradation. Our findings further resolve the mechanism for temporal regulation of REST and terminal neuronal differentiation. They also provide new potential therapeutic targets to enhance neuronal regeneration after injury. PMID:27535341

  7. Incorporating nucleosomes into thermodynamic models of transcription regulation.

    Science.gov (United States)

    Raveh-Sadka, Tali; Levo, Michal; Segal, Eran

    2009-08-01

    Transcriptional control is central to many cellular processes, and, consequently, much effort has been devoted to understanding its underlying mechanisms. The organization of nucleosomes along promoter regions is important for this process, since most transcription factors cannot bind nucleosomal sequences and thus compete with nucleosomes for DNA access. This competition is governed by the relative concentrations of nucleosomes and transcription factors and by their respective sequence binding preferences. However, despite its importance, a mechanistic understanding of the quantitative effects that the competition between nucleosomes and factors has on transcription is still missing. Here we use a thermodynamic framework based on fundamental principles of statistical mechanics to explore theoretically the effect that different nucleosome organizations along promoters have on the activation dynamics of promoters in response to varying concentrations of the regulating factors. We show that even simple landscapes of nucleosome organization reproduce experimental results regarding the effect of nucleosomes as general repressors and as generators of obligate binding cooperativity between factors. Our modeling framework also allows us to characterize the effects that various sequence elements of promoters have on the induction threshold and on the shape of the promoter activation curves. Finally, we show that using only sequence preferences for nucleosomes and transcription factors, our model can also predict expression behavior of real promoter sequences, thereby underscoring the importance of the interplay between nucleosomes and factors in determining expression kinetics.

  8. Transcriptional regulation by nonclassical action of thyroid hormone

    Directory of Open Access Journals (Sweden)

    Moeller Lars C

    2011-08-01

    Full Text Available Abstract Thyroid hormone (TH is essential for normal development, growth and metabolism. Its effects were thought to be principally mediated through triiodothyronine (T3, acting as a ligand for the nuclear TH receptors (TRs α and β residing on thyroid hormone response elements (TREs in the promoter of TH target genes. In this classical model of TH action, T3 binding to TRs leads to recruitment of basal transcription factors and increased transcription of TH responsive genes. Recently, the concept of TH action on gene expression has become more diverse and now includes nonclassical actions of T3 and T4: T3 has been shown to activate PI3K via the TRs, which ultimately increases transcription of certain genes, e.g. HIF-1α. Additionally, both T3 and thyroxine (T4 can bind to a membrane integrin, αvβ3, which leads to activation of the PI3K and MAPK signal transduction pathways and finally also increases gene transcription, e.g. of the FGF2 gene. Therefore, these initially nongenomic, nonclassical actions seem to serve as additional interfaces for transcriptional regulation by TH. Aim of this perspective is to summarize the genes that are currently known to be induced by nonclassical TH action and the mechanisms involved.

  9. Constitutive activation of signal transducer and activator of transcription 3 (STAT3) and nuclear factor κB signaling in glioblastoma cancer stem cells regulates the Notch pathway.

    Science.gov (United States)

    Garner, Jo Meagan; Fan, Meiyun; Yang, Chuan He; Du, Ziyun; Sims, Michelle; Davidoff, Andrew M; Pfeffer, Lawrence M

    2013-09-06

    Malignant gliomas are locally aggressive, highly vascular tumors that have a dismal prognosis, and present therapies provide little improvement in the disease course and outcome. Many types of malignancies, including glioblastoma, originate from a population of cancer stem cells (CSCs) that are able to initiate and maintain tumors. Although CSCs only represent a small fraction of cells within a tumor, their high tumor-initiating capacity and therapeutic resistance drives tumorigenesis. Therefore, it is imperative to identify pathways associated with CSCs to devise strategies to selectively target them. In this study, we describe a novel relationship between glioblastoma CSCs and the Notch pathway, which involves the constitutive activation of STAT3 and NF-κB signaling. Glioma CSCs were isolated and maintained in vitro using an adherent culture system, and the biological properties were compared with the traditional cultures of CSCs grown as multicellular spheres under nonadherent culture conditions. Interestingly, both adherent and spheroid glioma CSCs show constitutive activation of the STAT3/NF-κB signaling pathway and up-regulation of STAT3- and NF-κB-dependent genes. Gene expression profiling also identified components of the Notch pathway as being deregulated in glioma CSCs, and the deregulated expression of these genes was sensitive to treatment with STAT3 and NF-κB inhibitors. This finding is particularly important because Notch signaling appears to play a key role in CSCs in a variety of cancers and controls cell fate determination, survival, proliferation, and the maintenance of stem cells. The constitutive activation of STAT3 and NF-κB signaling pathways that leads to the regulation of Notch pathway genes in glioma CSCs identifies novel therapeutic targets for the treatment of glioma.

  10. Hydrogen peroxide sensing, signaling and regulation of transcription factors

    Directory of Open Access Journals (Sweden)

    H. Susana Marinho

    2014-01-01

    Full Text Available The regulatory mechanisms by which hydrogen peroxide (H2O2 modulates the activity of transcription factors in bacteria (OxyR and PerR, lower eukaryotes (Yap1, Maf1, Hsf1 and Msn2/4 and mammalian cells (AP-1, NRF2, CREB, HSF1, HIF-1, TP53, NF-κB, NOTCH, SP1 and SCREB-1 are reviewed. The complexity of regulatory networks increases throughout the phylogenetic tree, reaching a high level of complexity in mammalians. Multiple H2O2 sensors and pathways are triggered converging in the regulation of transcription factors at several levels: (1 synthesis of the transcription factor by upregulating transcription or increasing both mRNA stability and translation; (ii stability of the transcription factor by decreasing its association with the ubiquitin E3 ligase complex or by inhibiting this complex; (iii cytoplasm–nuclear traffic by exposing/masking nuclear localization signals, or by releasing the transcription factor from partners or from membrane anchors; and (iv DNA binding and nuclear transactivation by modulating transcription factor affinity towards DNA, co-activators or repressors, and by targeting specific regions of chromatin to activate individual genes. We also discuss how H2O2 biological specificity results from diverse thiol protein sensors, with different reactivity of their sulfhydryl groups towards H2O2, being activated by different concentrations and times of exposure to H2O2. The specific regulation of local H2O2 concentrations is also crucial and results from H2O2 localized production and removal controlled by signals. Finally, we formulate equations to extract from typical experiments quantitative data concerning H2O2 reactivity with sensor molecules. Rate constants of 140 M−1 s−1 and ≥1.3 × 103 M−1 s−1 were estimated, respectively, for the reaction of H2O2 with KEAP1 and with an unknown target that mediates NRF2 protein synthesis. In conclusion, the multitude of H2O2 targets and mechanisms provides an opportunity for

  11. Global transcriptional regulator TrmB family members in prokaryotes.

    Science.gov (United States)

    Kim, Minwook; Park, Soyoung; Lee, Sung-Jae

    2016-10-01

    Members of the TrmB family act as global transcriptional regulators for the activation or repression of sugar ABC transporters and central sugar metabolic pathways, including glycolytic, gluconeogenic, and other metabolic pathways, and also as chromosomal stabilizers in archaea. As a relatively newly classified transcriptional regulator family, there is limited experimental evidence for their role in Thermococcales, halophilic archaeon Halobacterium salinarum NRC1, and crenarchaea Sulfolobus strains, despite being one of the extending protein families in archaea. Recently, the protein structures of Pyrococcus furiosus TrmB and TrmBL2 were solved, and the transcriptomic data uncovered by microarray and ChIP-Seq were published. In the present review, recent evidence of the functional roles of TrmB family members in archaea is explained and extended to bacteria.

  12. VLDL hydrolysis by hepatic lipase regulates PPARδ transcriptional responses.

    Directory of Open Access Journals (Sweden)

    Jonathan D Brown

    Full Text Available BACKGROUND: PPARs (α,γ,δ are a family of ligand-activated transcription factors that regulate energy balance, including lipid metabolism. Despite these critical functions, the integration between specific pathways of lipid metabolism and distinct PPAR responses remains obscure. Previous work has revealed that lipolytic pathways can activate PPARs. Whether hepatic lipase (HL, an enzyme that regulates VLDL and HDL catabolism, participates in PPAR responses is unknown. METHODS/PRINCIPAL FINDINGS: Using PPAR ligand binding domain transactivation assays, we found that HL interacted with triglyceride-rich VLDL (>HDL≫LDL, IDL to activate PPARδ preferentially over PPARα or PPARγ, an effect dependent on HL catalytic activity. In cell free ligand displacement assays, VLDL hydrolysis by HL activated PPARδ in a VLDL-concentration dependent manner. Extended further, VLDL stimulation of HL-expressing HUVECs and FAO hepatoma cells increased mRNA expression of canonical PPARδ target genes, including adipocyte differentiation related protein (ADRP, angiopoietin like protein 4 and pyruvate dehydrogenase kinase-4. HL/VLDL regulated ADRP through a PPRE in the promoter region of this gene. In vivo, adenoviral-mediated hepatic HL expression in C57BL/6 mice increased hepatic ADRP mRNA levels by 30%. In ob/ob mice, a model with higher triglycerides than C57BL/6 mice, HL overexpression increased ADRP expression by 70%, demonstrating the importance of triglyceride substrate for HL-mediated PPARδ activation. Global metabolite profiling identified HL/VLDL released fatty acids including oleic acid and palmitoleic acid that were capable of recapitulating PPARδ activation and ADRP gene regulation in vitro. CONCLUSIONS: These data define a novel pathway involving HL hydrolysis of VLDL that activates PPARδ through generation of specific monounsaturated fatty acids. These data also demonstrate how integrating cell biology with metabolomic approaches provides insight

  13. Transcriptional Regulation of the Streptococcus salivarius 57.I Urease Operon

    Science.gov (United States)

    Chen, Yi-Ywan M.; Weaver, Cheryl A.; Mendelsohn, David R.; Burne, Robert A.

    1998-01-01

    The Streptococcus salivarius 57.I ure cluster was organized as an operon, beginning with ureI, followed by ureABC (structural genes) and ureEFGD (accessory genes). Northern analyses revealed transcripts encompassing structural genes and transcripts containing the entire operon. A ς70-like promoter could be mapped 5′ to ureI (PureI) by primer extension analysis. The intensity of the signal increased when cells were grown at an acidic pH and was further enhanced by excess carbohydrate. To determine the function(s) of two inverted repeats located 5′ to PureI, transcriptional fusions of the full-length promoter region (PureI), or a deletion derivative (PureIΔ100), and a promoterless chloramphenicol acetyltransferase (CAT) gene were constructed and integrated into the chromosome to generate strains PureICAT and PureIΔ100CAT, respectively. CAT specific activities of PureICAT were repressed at pH 7.0 and induced at pH 5.5 and by excess carbohydrate. In PureIΔ100CAT, CAT activity was 60-fold higher than in PureICAT at pH 7.0 and pH induction was nearly eliminated, indicating that expression was negatively regulated. Thus, it was concluded that PureI was the predominant, regulated promoter and that regulation was governed by a mechanism differing markedly from other known mechanisms for bacterial urease expression. PMID:9791132

  14. Insulin-like growth factor 1 receptor and p38 mitogen-activated protein kinase signals inversely regulate signal transducer and activator of transcription 3 activity to control human dental pulp stem cell quiescence, propagation, and differentiation.

    Science.gov (United States)

    Vandomme, Jerome; Touil, Yasmine; Ostyn, Pauline; Olejnik, Cecile; Flamenco, Pilar; El Machhour, Raja; Segard, Pascaline; Masselot, Bernadette; Bailliez, Yves; Formstecher, Pierre; Polakowska, Renata

    2014-04-15

    Dental pulp stem cells (DPSCs) remain quiescent until activated in response to severe dental pulp damage. Once activated, they exit quiescence and enter regenerative odontogenesis, producing reparative dentin. The factors and signaling molecules that control the quiescence/activation and commitment to differentiation of human DPSCs are not known. In this study, we determined that the inhibition of insulin-like growth factor 1 receptor (IGF-1R) and p38 mitogen-activated protein kinase (p38 MAPK) signaling commonly activates DPSCs and promotes their exit from the G0 phase of the cell cycle as well as from the pyronin Y(low) stem cell compartment. The inhibition of these two pathways, however, inversely determines DPSC fate. In contrast to p38 MAPK inhibitors, IGF-1R inhibitors enhance dental pulp cell sphere-forming capacity and reduce the cells' colony-forming capacity without inducing cell death. The inverse cellular changes initiated by IGF-1R and p38 MAPK inhibitors were accompanied by inverse changes in the levels of active signal transducer and activator of transcription 3 (STAT3) factor, inactive glycogen synthase kinase 3, and matrix extracellular phosphoglycoprotein, a marker of early odontoblast differentiation. Our data suggest that there is cross talk between the IGF-1R and p38 MAPK signaling pathways in DPSCs and that the signals provided by these pathways converge at STAT3 and inversely regulate its activity to maintain quiescence or to promote self-renewal and differentiation of the cells. We propose a working model that explains the possible interactions between IGF-1R and p38 MAPK at the molecular level and describes the cellular consequences of these interactions. This model may inspire further fundamental study and stimulate research on the clinical applications of DPSC in cellular therapy and tissue regeneration.

  15. Transcriptional and post-transcriptional regulation of a NAC1 transcription factor in Medicago truncatula roots.

    Science.gov (United States)

    D'haeseleer, Katrien; Den Herder, Griet; Laffont, Carole; Plet, Julie; Mortier, Virginie; Lelandais-Brière, Christine; De Bodt, Stefanie; De Keyser, Annick; Crespi, Martin; Holsters, Marcelle; Frugier, Florian; Goormachtig, Sofie

    2011-08-01

    • Legume roots develop two types of lateral organs, lateral roots and nodules. Nodules develop as a result of a symbiotic interaction with rhizobia and provide a niche for the bacteria to fix atmospheric nitrogen for the plant. • The Arabidopsis NAC1 transcription factor is involved in lateral root formation, and is regulated post-transcriptionally by miRNA164 and by SINAT5-dependent ubiquitination. We analyzed in Medicago truncatula the role of the closest NAC1 homolog in lateral root formation and in nodulation. • MtNAC1 shows a different expression pattern in response to auxin than its Arabidopsis homolog and no changes in lateral root number or nodulation were observed in plants affected in MtNAC1 expression. In addition, no interaction was found with SINA E3 ligases, suggesting that post-translational regulation of MtNAC1 does not occur in M. truncatula. Similar to what was found in Arabidopsis, a conserved miR164 target site was retrieved in MtNAC1, which reduced protein accumulation of a GFP-miR164 sensor. Furthermore, miR164 and MtNAC1 show an overlapping expression pattern in symbiotic nodules, and overexpression of this miRNA led to a reduction in nodule number. • This work suggests that regulatory pathways controlling a conserved transcription factor are complex and divergent between M. truncatula and Arabidopsis.

  16. Negative transcriptional regulation of mitochondrial transcription factor A (TFAM) by nuclear TFAM

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Eun Jin; Kang, Young Cheol; Park, Wook-Ha; Jeong, Jae Hoon; Pak, Youngmi Kim, E-mail: ykpak@khu.ac.kr

    2014-07-18

    Highlights: • TFAM localizes in nuclei and mitochondria of neuronal cells. • Nuclear TFAM does not bind the Tfam promoter. • Nuclear TFAM reduced the Tfam promoter activity via suppressing NRF-1 activity. • A novel self-negative feedback regulation of Tfam gene expression is explored. • FAM may play different roles depending on its subcellular localizations. - Abstract: The nuclear DNA-encoded mitochondrial transcription factor A (TFAM) is synthesized in cytoplasm and transported into mitochondria. TFAM enhances both transcription and replication of mitochondrial DNA. It is unclear, however, whether TFAM plays a role in regulating nuclear gene expression. Here, we demonstrated that TFAM was localized to the nucleus and mitochondria by immunostaining, subcellular fractionation, and TFAM-green fluorescent protein hybrid protein studies. In HT22 hippocampal neuronal cells, human TFAM (hTFAM) overexpression suppressed human Tfam promoter-mediated luciferase activity in a dose-dependent manner. The mitochondria targeting sequence-deficient hTFAM also repressed Tfam promoter activity to the same degree as hTFAM. It indicated that nuclear hTFAM suppressed Tfam expression without modulating mitochondrial activity. The repression required for nuclear respiratory factor-1 (NRF-1), but hTFAM did not bind to the NRF-1 binding site of its promoter. TFAM was co-immunoprecipitated with NRF-1. Taken together, we suggest that nuclear TFAM down-regulate its own gene expression as a NRF-1 repressor, showing that TFAM may play different roles depending on its subcellular localizations.

  17. Plant NAC-type transcription factor proteins contain a NARD domain for repression of transcriptional activation.

    Science.gov (United States)

    Hao, Yu-Jun; Song, Qing-Xin; Chen, Hao-Wei; Zou, Hong-Feng; Wei, Wei; Kang, Xu-Sheng; Ma, Biao; Zhang, Wan-Ke; Zhang, Jin-Song; Chen, Shou-Yi

    2010-10-01

    Plant-specific transcription factor NAC proteins play essential roles in many biological processes such as development, senescence, morphogenesis, and stress signal transduction pathways. In the NAC family, some members function as transcription activators while others act as repressors. In the present study we found that though the full-length GmNAC20 from soybean did not have transcriptional activation activity, the carboxy-terminal activation domain of GmNAC20 had high transcriptional activation activity in the yeast assay system. Deletion experiments revealed an active repression domain with 35 amino acids, named NARD (NAC Repression Domain), in the d subdomain of NAC DNA-binding domain. NARD can reduce the transcriptional activation ability of diverse transcription factors when fused to either the amino-terminal or the carboxy-terminal of the transcription factors. NARD-like sequences are also present in other NAC family members and they are functional repression domain when fused to VP16 in plant protoplast assay system. Mutation analysis of conserved amino acid residues in NARD showed that the hydrophobic LVFY motif may partially contribute to the repression function. It is hypothesized that the interactions between the repression domain NARD and the carboxy-terminal activation domain may finally determine the ability of NAC family proteins to regulate downstream gene expressions.

  18. ATF2, a paradigm of the multifaceted regulation of transcription factors in biology and disease.

    Science.gov (United States)

    Watson, Gregory; Ronai, Ze'ev; Lau, Eric

    2017-02-15

    Stringent transcriptional regulation is crucial for normal cellular biology and organismal development. Perturbations in the proper regulation of transcription factors can result in numerous pathologies, including cancer. Thus, understanding how transcription factors are regulated and how they are dysregulated in disease states is key to the therapeutic targeting of these factors and/or the pathways that they regulate. Activating transcription factor 2 (ATF2) has been studied in a number of developmental and pathological conditions. Recent findings have shed light on the transcriptional, post-transcriptional, and post-translational regulatory mechanisms that influence ATF2 function, and thus, the transcriptional programs coordinated by ATF2. Given our current knowledge of its multiple levels of regulation and function, ATF2 represents a paradigm for the mechanistic complexity that can regulate transcription factor function. Thus, increasing our understanding of the regulation and function of ATF2 will provide insights into fundamental regulatory mechanisms that influence how cells integrate extracellular and intracellular signals into a genomic response through transcription factors. Characterization of ATF2 dysfunction in the context of pathological conditions, particularly in cancer biology and response to therapy, will be important in understanding how pathways controlled by ATF2 or other transcription factors might be therapeutically exploited. In this review, we provide an overview of the currently known upstream regulators and downstream targets of ATF2.

  19. Characterization of Evolutionarily ConservedMotifs Involved in Activity and Regulation of theABA-INSENSITIVE (ABI) 4 Transcription Factor

    Institute of Scientific and Technical Information of China (English)

    2014-01-01

    In recent years, the transcription factor ABI4 has emerged as an important node of integration for externaland internal signals such as nutrient status and hormone signaling that modulates critical transitions during the growthand development of plants. For this reason, understanding the mechanism of action and regulation of this protein rep-resents an important step towards the elucidation of crosstalk mechanisms in plants. However, this understanding hasbeen hindered due to the negligible levels of this protein as a result of multiple posttranscriptional regulations. To betterunderstand the function and regulation of the ABI4 protein in this work, we performed a functional analysis of severalevolutionarily conserved motifs. Based on these conserved motifs, we identified ortholog genes of ABI4 in differentplant species. The functionality of the putative ortholog from Theobroma cacao was demonstrated in transient expres-sion assays and in complementation studies in plants. The function of the highly conserved motifs was analyzed aftertheir deletion or mutagenesis in the Arabidopsis ABI4 sequence using mesophyll protoplasts. This approach permitted usto immunologically detect the ABI4 protein and identify some of the mechanisms involved in its regulation. We identi-fied sequences required for the nuclear localization (AP2-associated motif) as well as those for transcriptional activationfunction (LRP motif). Moreover, this approach showed that the protein stability of this transcription factor is controlledthrough protein degradation and subcellular localization and involves the AP2-associated and the PEST motifs. We dem-onstrated that the degradation of ABI4 protein through the PEST motif is mediated by the 26S proteasome in responseto changes in the sugar levels.

  20. Transcriptional networks that regulate muscle stem cell function.

    Science.gov (United States)

    Punch, Vincent G; Jones, Andrew E; Rudnicki, Michael A

    2009-01-01

    Muscle stem cells comprise different populations of stem and progenitor cells found in embryonic and adult tissues. A number of signaling and transcriptional networks are responsible for specification and survival of these cell populations and regulation of their behavior during growth and regeneration. Muscle progenitor cells are mostly derived from the somites of developing embryos, while satellite cells are the progenitor cells responsible for the majority of postnatal growth and adult muscle regeneration. In resting muscle, these stem cells are quiescent, but reenter the cell cycle during their activation, whereby they undergo decisions to self-renew, proliferate, or differentiate and fuse into multinucleated myofibers to repair damaged muscle. Regulation of muscle stem cell activity is under the precise control of a number of extrinsic signaling pathways and active transcriptional networks that dictate their behavior, fate, and regenerative potential. Here, we review the networks responsible for these different aspects of muscle stem cell biology and discuss prevalent parallels between mechanisms regulating the activity of embryonic muscle progenitor cells and adult satellite cells.

  1. A matter of hierarchy: activation of orfamide production by the post-transcriptional Gac-Rsm cascade of Pseudomonas protegens CHA0 through expression upregulation of the two dedicated transcriptional regulators.

    Science.gov (United States)

    Sobrero, Patricio Martín; Muzlera, Andrés; Frescura, Julieta; Jofré, Edgardo; Valverde, Claudio

    2017-07-13

    In this work, we surveyed the genome of P. protegens CHA0 in order to identify novel mRNAs possibly under the control of the Gac-Rsm cascade that might, for their part, serve to elucidate as-yet-unknown functions involved in the biocontrol of plant pathogens and/or in cellular processes required for fitness in natural environments. In view of the experimental evidence from former studies on the Gac-Rsm cascade, we developed a computational screen supported by a combination of sequence, structural and evolutionary constraints that led to a dataset of 43 potential novel mRNA targets. We then confirmed several mRNA targets experimentally and next focused on two of the respective genes that are physically linked to the orfamide biosynthetic gene cluster and whose predicted open-reading frames resembled cognate LuxR-type transcriptional regulators of cyclic lipopeptide clusters in related pseudomonads. In this report, we demonstrate that in strain CHA0, orfamide production is stringently dependent on a functional Gac-Rsm cascade and that both mRNAs encoding transcriptional regulatory proteins are under direct translational control of the RsmA/E proteins. Our results have thus revealed a hierarchical control over the expression of orfamide biosynthetic genes with the final transcriptional control subordinated to the global Gac-Rsm post-transcriptional regulatory system. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  2. Transcription factor regulation by mechanical stress.

    Science.gov (United States)

    Mendez, Melissa G; Janmey, Paul A

    2012-05-01

    New technologies and interest in cell mechanics are generating exciting new discoveries about how material properties and forces affect biological structure and function. Mechanical forces are transduced via a variety of mechanisms, recently beginning to be revealed, into signals capable of altering cell function and structure. Responses to physical stimuli occur at multiple levels, from changes in the structures of single proteins to global cascades capable of altering cell proliferation and differentiation. This review describes recent findings in which physical stimuli were shown to modulate transcription factor activity, including that of armadillo/β-catenin, serum response factor (SRF), yes-associated protein (YAP) and nuclear factor κB (NF-κB). Copyright © 2012 Elsevier Ltd. All rights reserved.

  3. Transcriptional Regulation of Urokinase-type Plasminogen Activator Receptor by Hypoxia-Inducible Factor 1 Is Crucial for Invasion of Pancreatic and Liver Cancer

    Directory of Open Access Journals (Sweden)

    Peter Büchler

    2009-02-01

    Full Text Available Angioinvasion is critical for metastasis with urokinase-type plasminogen activator receptor (uPAR and tumor hypoxia-activated hypoxia-inducible factor 1 (HIF-1 as key players. Transcriptional control of uPAR expression by HIF has never been reported. The aim of the present study, therefore, was to test whether tumor hypoxia-induced HIF expression may be linked to transcriptional activation of uPAR and dependent angioinvasion. We used human pancreatic cancer cells and a model of parental and derived HIF-1β-deficient mouse liver cancer cell lines and performed Northern blot analysis, nuclear runoff assays, electrophoretic mobility shift assay, polymerase chain reaction-generated deletion mutants, luciferase assays, Matrigel invasion assays, and in vivo angioinvasion assays in the chorioallantoic membrane of fertilized chicken eggs. Urokinase-type plasminogen activator receptor promoter analysis resulted in four putative HIF binding sites. Hypoxia strongly induced de novo transcription of uPAR mRNA. With sequential deletion mutants of the uPAR promoter, it was possible to identify one HIF binding site causing a nearly 200-fold increase in luciferase activity. Hypoxia enhanced the number of invading tumor cells in vitro and in vivo. In contrast, HIF-1β-deficient cells failed to upregulate uPAR expression, to activate luciferase activity, and to invade on hypoxia. Taken together, we show for the first time that uPAR is under transcriptional control of HIF and that this is important for hypoxia-induced metastasis.

  4. Differential gene expression regulated by oscillatory transcription factors.

    Directory of Open Access Journals (Sweden)

    Luca Cerone

    Full Text Available Cells respond to changes in the internal and external environment by a complex regulatory system whose end-point is the activation of transcription factors controlling the expression of a pool of ad-hoc genes. Recent experiments have shown that certain stimuli may trigger oscillations in the concentration of transcription factors such as NF-κB and p53 influencing the final outcome of the genetic response. In this study we investigate the role of oscillations in the case of three different well known gene regulatory mechanisms using mathematical models based on ordinary differential equations and numerical simulations. We considered the cases of direct regulation, two-step regulation and feed-forward loops, and characterized their response to oscillatory input signals both analytically and numerically. We show that in the case of indirect two-step regulation the expression of genes can be turned on or off in a frequency dependent manner, and that feed-forward loops are also able to selectively respond to the temporal profile of oscillating transcription factors.

  5. In silico and wet lab approaches to study transcriptional regulation

    NARCIS (Netherlands)

    Hestand, Matthew Scott

    2010-01-01

    Gene expression is a complicated process with multiple types of regulation, including binding of proteins termed transcription factors. This thesis looks at transcription factors and transcription factor binding site discovery through computational predictions and wet lab work to better elucidate th

  6. The TrmB family: a versatile group of transcriptional regulators in Archaea.

    Science.gov (United States)

    Gindner, Antonia; Hausner, Winfried; Thomm, Michael

    2014-09-01

    Microbes are organisms which are well adapted to their habitat. Their survival depends on the regulation of gene expression levels in response to environmental signals. The most important step in regulation of gene expression takes place at the transcriptional level. This regulation is intriguing in Archaea because the eu-karyotic-like transcription apparatus is modulated by bacterial-like transcription regulators. The transcriptional regulator of mal operon (TrmB) family is well known as a very large group of regulators in Archaea with more than 250 members to date. One special feature of these regulators is that some of them can act as repressor, some as activator and others as both repressor and activator. This review gives a short updated overview of the TrmB family and their regulatory patterns in different Archaea as a lot of new data have been published on this topic since the last review from 2008.

  7. Transcription factors regulating B cell fate in the germinal centre.

    Science.gov (United States)

    Recaldin, T; Fear, D J

    2016-01-01

    Diversification of the antibody repertoire is essential for the normal operation of the vertebrate adaptive immune system. Following antigen encounter, B cells are activated, proliferate rapidly and undergo two diversification events; somatic hypermutation (followed by selection), which enhances the affinity of the antibody for its cognate antigen, and class-switch recombination, which alters the effector functions of the antibody to adapt the response to the challenge faced. B cells must then differentiate into antibody-secreting plasma cells or long-lived memory B cells. These activities take place in specialized immunological environments called germinal centres, usually located in the secondary lymphoid organs. To complete the germinal centre activities successfully, a B cell adopts a transcriptional programme that allows it to migrate to specific sites within the germinal centre, proliferate, modify its DNA recombination and repair pathways, alter its apoptotic potential and finally undergo terminal differentiation. To co-ordinate these processes, B cells employ a number of 'master regulator' transcription factors which mediate wholesale transcriptomic changes. These master transcription factors are mutually antagonistic and form a complex regulatory network to maintain distinct gene expression programs. Within this network, multiple points of positive and negative feedback ensure the expression of the 'master regulators', augmented by a number of 'secondary' factors that reinforce these networks and sense the progress of the immune response. In this review we will discuss the different activities B cells must undertake to mount a successful T cell-dependent immune response and describe how a regulatory network of transcription factors controls these processes.

  8. Redox regulation of cardiomyocyte cell cycling via an ERK1/2 and c-Myc-dependent activation of cyclin D2 transcription

    Science.gov (United States)

    Murray, Thomas V.A.; Smyrnias, Ioannis; Schnelle, Moritz; Mistry, Rajesh K.; Zhang, Min; Beretta, Matteo; Martin, Daniel; Anilkumar, Narayana; de Silva, Shana M.; Shah, Ajay M.; Brewer, Alison C.

    2015-01-01

    Adult mammalian cardiomyocytes have a very limited capacity to proliferate, and consequently the loss of cells after cardiac stress promotes heart failure. Recent evidence suggests that administration of hydrogen peroxide (H2O2), can regulate redox-dependent signalling pathway(s) to promote cardiomyocyte proliferation in vitro, but the potential relevance of such a pathway in vivo has not been tested. We have generated a transgenic (Tg) mouse model in which the H2O2-generating enzyme, NADPH oxidase 4 (Nox4), is overexpressed within the postnatal cardiomyocytes, and observed that the hearts of 1–3 week old Tg mice pups are larger in comparison to wild type (Wt) littermate controls. We demonstrate that the cardiomyocytes of Tg mouse pups have increased cell cycling capacity in vivo as determined by incorporation of 5-bromo-2′-deoxyuridine. Further, microarray analyses of the transcriptome of these Tg mouse hearts suggested that the expression of cyclin D2 is significantly increased. We investigated the molecular mechanisms which underlie this more proliferative phenotype in isolated neonatal rat cardiomyocytes (NRCs) in vitro, and demonstrate that Nox4 overexpression mediates an H2O2-dependent activation of the ERK1/2 signalling pathway, which in turn phosphorylates and activates the transcription factor c-myc. This results in a significant increase in cyclin D2 expression, which we show to be mediated, at least in part, by cis-acting c-myc binding sites within the proximal cyclin D2 promoter. Overexpression of Nox4 in NRCs results in an increase in their proliferative capacity that is ablated by the silencing of cyclin D2. We further demonstrate activation of the ERK1/2 signalling pathway, increased phosphorylation of c-myc and significantly increased expression of cyclin D2 protein in the Nox4 Tg hearts. We suggest that this pathway acts to maintain the proliferative capacity of cardiomyocytes in Nox4 Tg pups in vivo and so delays their exit from the cell

  9. FOXO Transcription Factors: Their Clinical Significance and Regulation

    Directory of Open Access Journals (Sweden)

    Yu Wang

    2014-01-01

    Full Text Available Members of the class O of forkhead box transcription factors (FOXO have important roles in metabolism, cellular proliferation, stress resistance, and apoptosis. The activity of FOXOs is tightly regulated by posttranslational modification, including phosphorylation, acetylation, and ubiquitylation. Activation of cell survival pathways such as phosphoinositide-3-kinase/AKT/IKK or RAS/mitogen-activated protein kinase phosphorylates FOXOs at different sites which regulate FOXOs nuclear localization or degradation. FOXO transcription factors are upregulated in a number of cell types including hepatocytes, fibroblasts, osteoblasts, keratinocytes, endothelial cells, pericytes, and cardiac myocytes. They are involved in a number of pathologic and physiologic processes that include proliferation, apoptosis, autophagy, metabolism, inflammation, cytokine expression, immunity, differentiation, and resistance to oxidative stress. These processes impact a number of clinical conditions such as carcinogenesis, diabetes, diabetic complications, cardiovascular disease, host response, and wound healing. In this paper, we focus on the potential role of FOXOs in different disease models and the regulation of FOXOs by various stimuli.

  10. Bipartite functions of the CREB co-activators selectively direct alternative splicing or transcriptional activation.

    Science.gov (United States)

    Amelio, Antonio L; Caputi, Massimo; Conkright, Michael D

    2009-09-16

    The CREB regulated transcription co-activators (CRTCs) regulate many biological processes by integrating and converting environmental inputs into transcriptional responses. Although the mechanisms by which CRTCs sense cellular signals are characterized, little is known regarding how CRTCs contribute to the regulation of cAMP inducible genes. Here we show that these dynamic regulators, unlike other co-activators, independently direct either pre-mRNA splice-site selection or transcriptional activation depending on the cell type or promoter context. Moreover, in other scenarios, the CRTC co-activators coordinately regulate transcription and splicing. Mutational analyses showed that CRTCs possess distinct functional domains responsible for regulating either pre-mRNA splicing or transcriptional activation. Interestingly, the CRTC1-MAML2 oncoprotein lacks the splicing domain and is incapable of altering splice-site selection despite robustly activating transcription. The differential usage of these distinct domains allows CRTCs to selectively mediate multiple facets of gene regulation, indicating that co-activators are not solely restricted to coordinating alternative splicing with increase in transcriptional activity.

  11. Characterization of E2F8, a novel E2F-like cell-cycle regulated repressor of E2F-activated transcription

    OpenAIRE

    Christensen, Jesper; Cloos, Paul; Toftegaard, Ulla; Klinkenberg, David; Bracken, Adrian P.; Trinh, Emmanuelle; Heeran, Mel; Di Stefano, Luisa; Helin, Kristian

    2005-01-01

    The E2F family of transcription factors are downstream effectors of the retinoblastoma protein, pRB, pathway and are essential for the timely regulation of genes necessary for cell-cycle progression. Here we describe the characterization of human and murine E2F8, a new member of the E2F family. Sequence analysis of E2F8 predicts the presence of two distinct E2F-related DNA binding domains suggesting that E2F8 and, the recently, identified E2F7 form a subgroup within the E2F family. We show th...

  12. Genomic dissection of conserved transcriptional regulation in intestinal epithelial cells

    Science.gov (United States)

    Camp, J. Gray; Weiser, Matthew; Cocchiaro, Jordan L.; Kingsley, David M.; Furey, Terrence S.; Sheikh, Shehzad Z.; Rawls, John F.

    2017-01-01

    The intestinal epithelium serves critical physiologic functions that are shared among all vertebrates. However, it is unknown how the transcriptional regulatory mechanisms underlying these functions have changed over the course of vertebrate evolution. We generated genome-wide mRNA and accessible chromatin data from adult intestinal epithelial cells (IECs) in zebrafish, stickleback, mouse, and human species to determine if conserved IEC functions are achieved through common transcriptional regulation. We found evidence for substantial common regulation and conservation of gene expression regionally along the length of the intestine from fish to mammals and identified a core set of genes comprising a vertebrate IEC signature. We also identified transcriptional start sites and other putative regulatory regions that are differentially accessible in IECs in all 4 species. Although these sites rarely showed sequence conservation from fish to mammals, surprisingly, they drove highly conserved IEC expression in a zebrafish reporter assay. Common putative transcription factor binding sites (TFBS) found at these sites in multiple species indicate that sequence conservation alone is insufficient to identify much of the functionally conserved IEC regulatory information. Among the rare, highly sequence-conserved, IEC-specific regulatory regions, we discovered an ancient enhancer upstream from her6/HES1 that is active in a distinct population of Notch-positive cells in the intestinal epithelium. Together, these results show how combining accessible chromatin and mRNA datasets with TFBS prediction and in vivo reporter assays can reveal tissue-specific regulatory information conserved across 420 million years of vertebrate evolution. We define an IEC transcriptional regulatory network that is shared between fish and mammals and establish an experimental platform for studying how evolutionarily distilled regulatory information commonly controls IEC development and physiology. PMID

  13. Regulation of Transcriptional Networks by PKC Isozymes: Identification of c-Rel as a Key Transcription Factor for PKC-Regulated Genes.

    Directory of Open Access Journals (Sweden)

    Rachana Garg

    Full Text Available Activation of protein kinase C (PKC, a family of serine-threonine kinases widely implicated in cancer progression, has major impact on gene expression. In a recent genome-wide analysis of prostate cancer cells we identified distinctive gene expression profiles controlled by individual PKC isozymes and highlighted a prominent role for PKCδ in transcriptional activation.Here we carried out a thorough bioinformatics analysis to dissect transcriptional networks controlled by PKCα, PKCδ, and PKCε, the main diacylglycerol/phorbol ester PKCs expressed in prostate cancer cells. Despite the remarkable differences in the patterns of transcriptional responsive elements (REs regulated by each PKC, we found that c-Rel represents the most frequent RE in promoters regulated by all three PKCs. In addition, promoters of PKCδ-regulated genes were particularly enriched with REs for CREB, NF-E2, RREB, SRF, Oct-1, Evi-1, and NF-κB. Most notably, by using transcription factor-specific RNAi we were able to identify subsets of PKCδ-regulated genes modulated by c-Rel and CREB. Furthermore, PKCδ-regulated genes condensed under the c-Rel transcriptional regulation display significant functional interconnections with biological processes such as angiogenesis, inflammatory response, and cell motility.Our study identified candidate transcription factors in the promoters of PKC regulated genes, in particular c-Rel was found as a key transcription factor in the control of PKCδ-regulated genes. The deconvolution of PKC-regulated transcriptional networks and their nodes may greatly help in the identification of PKC effectors and have significant therapeutics implications.

  14. Detection, characterization and regulation of antisense transcripts in HIV-1

    Directory of Open Access Journals (Sweden)

    Mesnard Jean-Michel

    2007-10-01

    Full Text Available Abstract Background We and others have recently demonstrated that the human retrovirus HTLV-I was producing a spliced antisense transcript, which led to the synthesis of the HBZ protein. The objective of the present study was to demonstrate the existence of antisense transcription in HIV-1 and to provide a better characterization of the transcript and its regulation. Results Initial experiments conducted by standard RT-PCR analysis in latently infected J1.1 cell line and pNL4.3-transfected 293T cells confirmed the existence of antisense transcription in HIV-1. A more adapted RT-PCR protocol with limited RT-PCR artefacts also led to a successful detection of antisense transcripts in several infected cell lines. RACE analyses demonstrated the existence of several transcription initiation sites mapping near the 5' border of the 3'LTR (in the antisense strand. Interestingly, a new polyA signal was identified on the antisense strand and harboured the polyA signal consensus sequence. Transfection experiments in 293T and Jurkat cells with an antisense luciferase-expressing NL4.3 proviral DNA showed luciferase reporter gene expression, which was further induced by various T-cell activators. In addition, the viral Tat protein was found to be a positive modulator of antisense transcription by transient and stable transfections of this proviral DNA construct. RT-PCR analyses in 293T cells stably transfected with a pNL4.3-derived construct further confirmed these results. Infection of 293T, Jurkat, SupT1, U937 and CEMT4 cells with pseudotyped virions produced from the antisense luciferase-expressing NL4.3 DNA clone led to the production of an AZT-sensitive luciferase signal, which was however less pronounced than the signal from NL4.3Luc-infected cells. Conclusion These results demonstrate for the first time that antisense transcription exists in HIV-1 in the context of infection. Possible translation of the predicted antisense ORF in this transcript should

  15. Transcriptional regulation by the numbers: models.

    Science.gov (United States)

    Bintu, Lacramioara; Buchler, Nicolas E; Garcia, Hernan G; Gerland, Ulrich; Hwa, Terence; Kondev, Jané; Phillips, Rob

    2005-04-01

    The expression of genes is regularly characterized with respect to how much, how fast, when and where. Such quantitative data demands quantitative models. Thermodynamic models are based on the assumption that the level of gene expression is proportional to the equilibrium probability that RNA polymerase (RNAP) is bound to the promoter of interest. Statistical mechanics provides a framework for computing these probabilities. Within this framework, interactions of activators, repressors, helper molecules and RNAP are described by a single function, the "regulation factor". This analysis culminates in an expression for the probability of RNA polymerase binding at the promoter of interest as a function of the number of regulatory proteins in the cell.

  16. ATF-2 regulates lipopolysaccharide-induced transcription in macrophage cells.

    Science.gov (United States)

    Hirose, Noriyuki; Maekawa, Toshio; Shinagawa, Toshie; Ishii, Shunsuke

    2009-07-17

    The transcription factor ATF-2, a member of the ATF/CREB family, is a target of p38 that are involved in stress-induced apoptosis and in Toll-like receptor (TLR)-mediated signaling. Phosphorylation of ATF-2 at Thr-71 was enhanced by treating of RAW264.7 macrophage cells with either LPS, MALP-2, or CpG-ODN. LPS treatment enhanced the trans-activation capacity of ATF-2. Among multiple LPS-induced genes, the LPS-induced expression of Socs-3 was significantly reduced by the treatment of RAW264.7 cells with an Atf-2 siRNA. Transcription from the Socs-3 promoter was synergistically stimulated by ATF-2 and LPS, whereas it was suppressed by Atf-2 siRNA. Histone deacetylase 1 (HDAC1) interacted with ATF-2 after LPS treatment, but not before treatment. Treatment of RAW264.7 cells with trichostatin A, an inhibitor of HDAC, suppressed the LPS-induced Socs-3 expression, suggesting that HDAC1 positively regulates the LPS-induced transcription of Socs-3. Thus, ATF-2 plays an important role in TLR-mediated transcriptional control in macrophage cells.

  17. A transcriptional cofactor YAP regulates IFNT expression via transcription factor TEAD in bovine conceptuses.

    Science.gov (United States)

    Kusama, K; Bai, R; Sakurai, T; Bai, H; Ideta, A; Aoyagi, Y; Imakawa, K

    2016-10-01

    Interferon tau (IFNT) is the pregnancy recognition protein in all ruminants, and its expression is restricted to trophoblast cells. Interferon tau production increases as the conceptus elongates; however, its expression is downregulated soon after the initiation of conceptus attachment to the uterine epithelium. Our previous study identified that among 8 bovine IFNT genes, only 2 forms of IFNTs, IFNT2 and IFN-tau-c1, were expressed by the conceptuses during the periattachment period. To characterize whether Hippo signaling including a transcription cofactor yes-associated protein (YAP) was involved in the IFNT regulation, we examined the expression and effects of YAP and/or TEAD in human choriocarcinoma JEG3 and bovine trophoblast CT-1 cells, and in bovine conceptuses obtained from day 17, 20 or 22 pregnant animals (pregnant day 19.5 = day of conceptus attachment to the endometrium). YAP was expressed in bovine conceptuses and transfection of YAP or TEAD4, a transcription factor partner of YAP, expression plasmid increased the luciferase activity of IFNT2 and IFN-tau-c1 reporter plasmids in JEG3 cells. In the presence of YAP expression plasmid, TEAD2 or TEAD4 expression plasmid further upregulated transcriptional activity of IFNT2 or IFN-tau-c1 constructs, which were substantially reduced in the absence of the TEAD-binding site on IFNT2 or IFN-tau-c1 promoter region in JEG3 cells. In CT-1 cells, treatment with TEAD2, TEAD4, or YAP small-interfering RNA downregulated endogenous IFNT expression. It should be noted that TEAD2 and TEAD4 were predominantly localized in the nuclei of trophectoderm of Day 17 conceptuses, but nuclear localization appeared to be lower in those cells of conceptuses on days 20 and 22 of pregnancy. Moreover, the binding of TEAD4 to the TEAD-binding site of the IFN-tau-c1 promoter region in day 17 conceptuses was less in day 20 and 22 conceptuses. Furthermore, the level of YAP phosphorylation increased in day 20 and 22 conceptuses. These

  18. Engineering transcriptional regulation to control Pdu microcompartment formation.

    Science.gov (United States)

    Kim, Edward Y; Jakobson, Christopher M; Tullman-Ercek, Danielle

    2014-01-01

    Bacterial microcompartments (MCPs) show great promise for the organization of engineered metabolic pathways within the bacterial cytoplasm. This subcellular organelle is composed of a protein shell of 100-200 nm diameter that natively encapsulates multi-enzyme pathways. The high energy cost of synthesizing the thousands of protein subunits required for each MCP demands precise regulation of MCP formation for both native and engineered systems. Here, we study the regulation of the propanediol utilization (Pdu) MCP, for which growth on 1,2-propanediol induces expression of the Pdu operon for the catabolism of 1,2-propanediol. We construct a fluorescence-based transcriptional reporter to investigate the activation of the Ppdu promoter, which drives the transcription of 21 pdu genes. Guided by this reporter, we find that MCPs can be expressed in strains grown in rich media, provided that glucose is not present. We also characterize the response of the Ppdu promoter to a transcriptional activator of the pdu operon, PocR, and find PocR to be a necessary component of Pdu MCP formation. Furthermore, we find that MCPs form normally upon the heterologous expression of PocR even in the absence of the natural inducer 1,2-propanediol and in the presence of glucose, and that Pdu MCPs formed in response to heterologous PocR expression can metabolize 1,2-propanediol in vivo. We anticipate that this technique of overexpressing a key transcription factor may be used to study and engineer the formation, size, and/or number of MCPs for the Pdu and related MCP systems.

  19. Engineering transcriptional regulation to control Pdu microcompartment formation.

    Directory of Open Access Journals (Sweden)

    Edward Y Kim

    Full Text Available Bacterial microcompartments (MCPs show great promise for the organization of engineered metabolic pathways within the bacterial cytoplasm. This subcellular organelle is composed of a protein shell of 100-200 nm diameter that natively encapsulates multi-enzyme pathways. The high energy cost of synthesizing the thousands of protein subunits required for each MCP demands precise regulation of MCP formation for both native and engineered systems. Here, we study the regulation of the propanediol utilization (Pdu MCP, for which growth on 1,2-propanediol induces expression of the Pdu operon for the catabolism of 1,2-propanediol. We construct a fluorescence-based transcriptional reporter to investigate the activation of the Ppdu promoter, which drives the transcription of 21 pdu genes. Guided by this reporter, we find that MCPs can be expressed in strains grown in rich media, provided that glucose is not present. We also characterize the response of the Ppdu promoter to a transcriptional activator of the pdu operon, PocR, and find PocR to be a necessary component of Pdu MCP formation. Furthermore, we find that MCPs form normally upon the heterologous expression of PocR even in the absence of the natural inducer 1,2-propanediol and in the presence of glucose, and that Pdu MCPs formed in response to heterologous PocR expression can metabolize 1,2-propanediol in vivo. We anticipate that this technique of overexpressing a key transcription factor may be used to study and engineer the formation, size, and/or number of MCPs for the Pdu and related MCP systems.

  20. The molecular clock regulates circadian transcription of tissue factor gene.

    Science.gov (United States)

    Oishi, Katsutaka; Koyanagi, Satoru; Ohkura, Naoki

    2013-02-01

    Tissue factor (TF) is involved in endotoxin-induced inflammation and mortality. We found that the circadian expression of TF mRNA, which peaked at the day to night transition (activity onset), was damped in the liver of Clock mutant mice. Luciferase reporter and chromatin immunoprecipitation analyses using embryonic fibroblasts derived from wild-type or Clock mutant mice showed that CLOCK is involved in transcription of the TF gene. Furthermore, the results of real-time luciferase reporter experiments revealed that the circadian expression of TF mRNA is regulated by clock molecules through a cell-autonomous mechanism via an E-box element located in the promoter region.

  1. Role of CTCF protein in regulating FMR1 locus transcription.

    Directory of Open Access Journals (Sweden)

    Stella Lanni

    Full Text Available Fragile X syndrome (FXS, the leading cause of inherited intellectual disability, is caused by epigenetic silencing of the FMR1 gene, through expansion and methylation of a CGG triplet repeat (methylated full mutation. An antisense transcript (FMR1-AS1, starting from both promoter and intron 2 of the FMR1 gene, was demonstrated in transcriptionally active alleles, but not in silent FXS alleles. Moreover, a DNA methylation boundary, which is lost in FXS, was recently identified upstream of the FMR1 gene. Several nuclear proteins bind to this region, like the insulator protein CTCF. Here we demonstrate for the first time that rare unmethylated full mutation (UFM alleles present the same boundary described in wild type (WT alleles and that CTCF binds to this region, as well as to the FMR1 gene promoter, exon 1 and intron 2 binding sites. Contrariwise, DNA methylation prevents CTCF binding to FXS alleles. Drug-induced CpGs demethylation does not restore this binding. CTCF knock-down experiments clearly established that CTCF does not act as insulator at the active FMR1 locus, despite the presence of a CGG expansion. CTCF depletion induces heterochromatinic histone configuration of the FMR1 locus and results in reduction of FMR1 transcription, which however is not accompanied by spreading of DNA methylation towards the FMR1 promoter. CTCF depletion is also associated with FMR1-AS1 mRNA reduction. Antisense RNA, like sense transcript, is upregulated in UFM and absent in FXS cells and its splicing is correlated to that of the FMR1-mRNA. We conclude that CTCF has a complex role in regulating FMR1 expression, probably through the organization of chromatin loops between sense/antisense transcriptional regulatory regions, as suggested by bioinformatics analysis.

  2. The EDLL motif: a potent plant transcriptional activation domain from AP2/ERF transcription factors.

    Science.gov (United States)

    Tiwari, Shiv B; Belachew, Alemu; Ma, Siu Fong; Young, Melinda; Ade, Jules; Shen, Yu; Marion, Colleen M; Holtan, Hans E; Bailey, Adina; Stone, Jeffrey K; Edwards, Leslie; Wallace, Andreah D; Canales, Roger D; Adam, Luc; Ratcliffe, Oliver J; Repetti, Peter P

    2012-06-01

    In plants, the ERF/EREBP family of transcriptional regulators plays a key role in adaptation to various biotic and abiotic stresses. These proteins contain a conserved AP2 DNA-binding domain and several uncharacterized motifs. Here, we describe a short motif, termed 'EDLL', that is present in AtERF98/TDR1 and other clade members from the same AP2 sub-family. We show that the EDLL motif, which has a unique arrangement of acidic amino acids and hydrophobic leucines, functions as a strong activation domain. The motif is transferable to other proteins, and is active at both proximal and distal positions of target promoters. As such, the EDLL motif is able to partly overcome the repression conferred by the AtHB2 transcription factor, which contains an ERF-associated amphiphilic repression (EAR) motif. We further examined the activation potential of EDLL by analysis of the regulation of flowering time by NF-Y (nuclear factor Y) proteins. Genetic evidence indicates that NF-Y protein complexes potentiate the action of CONSTANS in regulation of flowering in Arabidopsis; we show that the transcriptional activation function of CONSTANS can be substituted by direct fusion of the EDLL activation motif to NF-YB subunits. The EDLL motif represents a potent plant activation domain that can be used as a tool to confer transcriptional activation potential to heterologous DNA-binding proteins.

  3. Transcriptional regulation of human thromboxane synthase gene expression

    Energy Technology Data Exchange (ETDEWEB)

    Lee, K.D.; Baek, S.J.; Fleischer, T [Univ. of Maryland Medical School, Baltimore, MD (United States)] [and others

    1994-09-01

    The human thromboxane synthase (TS) gene encodes a microsomal enzyme catalyzing the conversion of prostaglandin endoperoxide into thromboxane A{sub 2}(TxA{sub 2}), a potent inducer of vasoconstriction and platelet aggregation. A deficiency in platelet TS activity results in bleeding disorders, but the underlying molecular mechanism remains to be elucidated. Increased TxA{sub 2} has been associated with many pathophysiological conditions such as cardiovascular disease, pulmonary hypertension, pre-eclampsia, and thrombosis in sickle cell patients. Since the formation of TxA{sub 2} is dependent upon TS, the regulation of TS gene expression may presumably play a crucial role in vivo. Abrogation of the regulatory mechanism in TS gene expression might contribute, in part, to the above clinical manifestations. To gain insight into TS gene regulation, a 1.7 kb promoter of the human TS gene was cloned and sequenced. RNase protection assay and 5{prime} RACE protocols were used to map the transcription initiation site to nucleotide A, 30 bp downstream from a canonical TATA box. Several transcription factor binding sites, including AP-1, PU.1, and PEA3, were identified within this sequence. Transient expression studies in HL-60 cells transfected with constructs containing various lengths (0.2 to 5.5 kb) of the TS promoter/luciferase fusion gene indicated the presence of multiple repressor elements within the 5.5 kb TS promoter. However, a lineage-specific up-regulation of TS gene expression was observed in HL-60 cells induced by TPA to differentiate along the macrophage lineage. The increase in TS transcription was not detectable until 36 hr after addition of the inducer. These results suggest that expression of the human TS gene may be regulated by a mechanism involving repression and derepression of the TS promoter.

  4. Regulation of Insulin Gene Transcription by Multiple Histone Acetyltransferases

    OpenAIRE

    2012-01-01

    Glucose-stimulated insulin gene transcription is mainly regulated by a 340-bp promoter region upstream of the transcription start site by beta-cell-enriched transcription factors Pdx-1, MafA, and NeuroD1. Previous studies have shown that histone H4 hyperacetylation is important for acute up-regulation of insulin gene transcription. Until now, only the histone acetyltransferase (HAT) protein p300 has been shown to be involved in this histone H4 acetylation event. In this report we investigated...

  5. Ascorbic acid-dependent gene expression in Streptococcus pneumoniae and the activator function of the transcriptional regulator UlaR2.

    Science.gov (United States)

    Afzal, Muhammad; Shafeeq, Sulman; Kuipers, Oscar P

    2015-01-01

    In this study, we have explored the impact of ascorbic acid on the transcriptome of Streptococcus pneumoniae D39. The expression of several genes and operons, including the ula operon (which has been previously shown to be involved in ascorbic acid utilization), the AdcR regulon (which has been previously shown to be involved in zinc transport and virulence) and a PTS operon (which we denote here as ula2 operon) were altered in the presence of ascorbic acid. The ula2 operon consists of five genes, including the transcriptional activator ulaR2. Our β-galactosidase assay data and transcriptome comparison of the ulaR2 mutant with the wild-type demonstrated that the transcriptional activator UlaR2 in the presence of ascorbic acid activates the expression of the ula2 operon. We further predict a 16-bp regulatory site (5'-ATATTGTGCTCAAATA-3') for UlaR2 in the Pula2. Furthermore, we have explored the effect of ascorbic acid on the expression of the AdcR regulon. Our ICP-MS analysis showed that addition of ascorbic acid to the medium causes zinc starvation in the cell which leads to the activation of the AdcR regulon.

  6. Contraction-induced Interleukin-6 Gene Transcription in Skeletal Muscle Is Regulated by c-Jun Terminal Kinase/Activator Protein-1*

    Science.gov (United States)

    Whitham, Martin; Chan, M. H. Stanley; Pal, Martin; Matthews, Vance B.; Prelovsek, Oja; Lunke, Sebastian; El-Osta, Assam; Broenneke, Hella; Alber, Jens; Brüning, Jens C.; Wunderlich, F. Thomas; Lancaster, Graeme I.; Febbraio, Mark A.

    2012-01-01

    Exercise increases the expression of the prototypical myokine IL-6, but the precise mechanism by which this occurs has yet to be identified. To mimic exercise conditions, C2C12 myotubes were mechanically stimulated via electrical pulse stimulation (EPS). We compared the responses of EPS with the pharmacological Ca2+ carrier calcimycin (A23187) because contraction induces marked increases in cytosolic Ca2+ levels or the classical IκB kinase/NFκB inflammatory response elicited by H2O2. We demonstrate that, unlike H2O2-stimulated increases in IL-6 mRNA, neither calcimycin- nor EPS-induced IL-6 mRNA expression is under the transcriptional control of NFκB. Rather, we show that EPS increased the phosphorylation of JNK and the reporter activity of the downstream transcription factor AP-1. Furthermore, JNK inhibition abolished the EPS-induced increase in IL-6 mRNA and protein expression. Finally, we observed an exercise-induced increase in both JNK phosphorylation and IL-6 mRNA expression in the skeletal muscles of mice after 30 min of treadmill running. Importantly, exercise did not increase IL-6 mRNA expression in skeletal muscle-specific JNK-deficient mice. These data identify a novel contraction-mediated transcriptional regulatory pathway for IL-6 in skeletal muscle. PMID:22351769

  7. Transcriptional regulation of cathelicidin genes in chicken bone marrow cells.

    Science.gov (United States)

    Lee, Sang In; Jang, Hyun June; Jeon, Mi-hyang; Lee, Mi Ock; Kim, Jeom Sun; Jeon, Ik-Soo; Byun, Sung June

    2016-04-01

    Cathelicidins form a family of vertebrate-specific immune molecules with an evolutionarily conserved gene structure. We analyzed the expression patterns of cathelicidin genes (CAMP, CATH3, and CATHB1) in chicken bone marrow cells (BMCs) and chicken embryonic fibroblasts (CEFs). We found that CAMP and CATHB1 were significantly up-regulated in BMCs, whereas the expression of CATH3 did not differ significantly between BMCs and CEFs. To study the mechanism underlying the up-regulation of cathelicidin genes in BMCs, we predicted the transcription factors (TFs) that bind to the 5'-flanking regions of cathelicidin genes. CEBPA, EBF1, HES1, MSX1, and ZIC3 were up-regulated in BMCs compared to CEFs. Subsequently, when a siRNA-mediated knockdown assay was performed for MSX1, the expression of CAMP and CATHB1 was decreased in BMCs. We also showed that the transcriptional activity of the CAMP promoter was decreased by mutation of the MSX1-binding sites present within the 5'-flanking region of CAMP. These results increase our understanding of the regulatory mechanisms controlling cathelicidin genes in BMCs.

  8. FOXL2 Is an Essential Activator of SF-1-Induced Transcriptional Regulation of Anti-Müllerian Hormone in Human Granulosa Cells.

    Science.gov (United States)

    Jin, Hanyong; Won, Miae; Park, Si Eun; Lee, Seunghwa; Park, Mira; Bae, Jeehyeon

    2016-01-01

    Anti-Müllerian hormone (AMH) is required for proper sexual differentiation by regulating the regression of the Müllerian ducts in males. Recent studies indicate that AMH could be an important factor for maintaining the ovarian reserve. However, the mechanisms of AMH regulation in the ovary are largely unknown. Here, we provide evidence that AMH is an ovarian target gene of steroidogenic factor-1 (SF-1), an orphan nuclear receptor required for proper follicle development. FOXL2 is an evolutionally conserved transcription factor, and its mutations cause blepharophimosis, ptosis, and epicanthus inversus syndrome (BPES), wherein affected females display eyelid defects and premature ovarian failure (POF). Notably, we found that functional FOXL2 is essential for SF-1-induced AMH regulation, via protein-protein interactions between FOXL2 and SF-1. A BPES-inducing mutant of FOXL2 (290-291delCA) was unable to interact with SF-1 and failed to mediate the association between SF-1 and the AMH promoter. Therefore, this study identified a novel regulatory circuit for ovarian AMH production; specifically, through the coordinated interplay between FOXL2 and SF-1 that could control ovarian follicle development.

  9. FOXL2 Is an Essential Activator of SF-1-Induced Transcriptional Regulation of Anti-Mullerian Hormone in Human Granulosa Cells.

    Directory of Open Access Journals (Sweden)

    Hanyong Jin

    Full Text Available Anti-Müllerian hormone (AMH is required for proper sexual differentiation by regulating the regression of the Müllerian ducts in males. Recent studies indicate that AMH could be an important factor for maintaining the ovarian reserve. However, the mechanisms of AMH regulation in the ovary are largely unknown. Here, we provide evidence that AMH is an ovarian target gene of steroidogenic factor-1 (SF-1, an orphan nuclear receptor required for proper follicle development. FOXL2 is an evolutionally conserved transcription factor, and its mutations cause blepharophimosis, ptosis, and epicanthus inversus syndrome (BPES, wherein affected females display eyelid defects and premature ovarian failure (POF. Notably, we found that functional FOXL2 is essential for SF-1-induced AMH regulation, via protein-protein interactions between FOXL2 and SF-1. A BPES-inducing mutant of FOXL2 (290-291delCA was unable to interact with SF-1 and failed to mediate the association between SF-1 and the AMH promoter. Therefore, this study identified a novel regulatory circuit for ovarian AMH production; specifically, through the coordinated interplay between FOXL2 and SF-1 that could control ovarian follicle development.

  10. Oct-2 transcription factor binding activity and expression up-regulation in rat cerebral ischaemia is associated with a diminution of neuronal damage in vitro.

    Science.gov (United States)

    Camós, Susanna; Gubern, Carme; Sobrado, Mónica; Rodríguez, Rocío; Romera, Víctor G; Moro, María Ángeles; Lizasoain, Ignacio; Serena, Joaquín; Mallolas, Judith; Castellanos, Mar

    2014-06-01

    Brain plasticity provides a mechanism to compensate for lesions produced as a result of stroke. The present study aims to identify new transcription factors (TFs) following focal cerebral ischaemia in rat as potential therapeutic targets. A transient focal cerebral ischaemia model was used for TF-binding activity and TF-TF interaction profile analysis. A permanent focal cerebral ischaemia model was used for the transcript gene analysis and for the protein study. The identification of TF variants, mRNA analysis, and protein study was performed using conventional polymerase chain reaction (PCR), qPCR, and Western blot and immunofluorescence, respectively. Rat cortical neurons were transfected with small interfering RNA against the TF in order to study its role. The TF-binding analysis revealed a differential binding activity of the octamer family in ischaemic brain in comparison with the control brain samples both in acute and late phases. In this study, we focused on Oct-2 TF. Five of the six putative Oct-2 transcript variants are expressed in both control and ischaemic rat brain, showing a significant increase in the late phase of ischaemia. Oct-2 protein showed neuronal localisation both in control and ischaemic rat brain cortical slices. Functional studies revealed that Oct-2 interacts with TFs involved in important brain processes (neuronal and vascular development) and basic cellular functions and that Oct-2 knockdown promotes neuronal injury. The present study shows that Oct-2 expression and binding activity increase in the late phase of cerebral ischaemia and finds Oct-2 to be involved in reducing ischaemic-mediated neuronal injury.

  11. Transcriptional regulation is a major controller of cell cycle transition dynamics

    DEFF Research Database (Denmark)

    Romanel, Alessandro; Jensen, Lars Juhl; Cardelli, Luca

    2012-01-01

    in various organisms showed the importance of positive feedbacks in other transitions as well. Here we investigate if a universal control system with transcriptional regulation(s) and post-translational positive feedback(s) can be proposed for the regulation of all cell cycle transitions. Through...... computational modeling, we analyze the transition dynamics in all possible combinations of transcriptional and post-translational regulations. We find that some combinations lead to 'sloppy' transitions, while others give very precise control. The periodic transcriptional regulation through the activator...... or the inhibitor leads to radically different dynamics. Experimental evidence shows that in cell cycle transitions of organisms investigated for cell cycle dependent periodic transcription, only the inhibitor OR the activator is under cyclic control and never both of them. Based on these observations, we propose...

  12. Transcription factor FOXA2-centered transcriptional regulation network in non-small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Sang-Min; An, Joo-Hee; Kim, Chul-Hong; Kim, Jung-Woong, E-mail: jungkim@cau.ac.kr; Choi, Kyung-Hee, E-mail: khchoi@cau.ac.kr

    2015-08-07

    Lung cancer is the leading cause of cancer-mediated death. Although various therapeutic approaches are used for lung cancer treatment, these mainly target the tumor suppressor p53 transcription factor, which is involved in apoptosis and cell cycle arrest. However, p53-targeted therapies have limited application in lung cancer, since p53 is found to be mutated in more than half of lung cancers. In this study, we propose tumor suppressor FOXA2 as an alternative target protein for therapies against lung cancer and reveal a possible FOXA2-centered transcriptional regulation network by identifying new target genes and binding partners of FOXA2 by using various screening techniques. The genes encoding Glu/Asp-rich carboxy-terminal domain 2 (CITED2), nuclear receptor subfamily 0, group B, member 2 (NR0B2), cell adhesion molecule 1 (CADM1) and BCL2-associated X protein (BAX) were identified as putative target genes of FOXA2. Additionally, the proteins including highly similar to heat shock protein HSP 90-beta (HSP90A), heat shock 70 kDa protein 1A variant (HSPA1A), histone deacetylase 1 (HDAC1) and HDAC3 were identified as novel interacting partners of FOXA2. Moreover, we showed that FOXA2-dependent promoter activation of BAX and p21 genes is significantly reduced via physical interactions between the identified binding partners and FOXA2. These results provide opportunities to understand the FOXA2-centered transcriptional regulation network and novel therapeutic targets to modulate this network in p53-deficient lung cancer. - Highlights: • Identification of new target genes of FOXA2. • Identifications of novel interaction proteins of FOXA2. • Construction of FOXA2-centered transcriptional regulatory network in non-small cell lung cancer.

  13. Transcriptional regulation of mitochondrial HMG-CoA synthase in the control of ketogenesis.

    Science.gov (United States)

    Hegardt, F G

    1998-10-01

    Mitochondrial and cytosolic HMG-CoA synthases are encoded by two different genes. Control of ketogenesis is exerted by transcriptional regulation of mitochondrial HMG-CoA synthase. Fasting, cAMP, and fatty acids increase its transcriptional rate, while refeeding and insulin repress it. Fatty acids increase transcription through peroxisomal proliferator regulatory element (PPRE), to which peroxisome proliferator activated receptor (PPAR) can bind. Other transcription factors such as chicken ovalbumin upstream promoter transcription factor (COUP-TF) and hepatocyte nuclear factor 4 (HNF-4) compete for the PPRE site, modulating the response of PPAR.

  14. Antioxidative Activities of Both Oleic Acid and Camellia tenuifolia Seed Oil Are Regulated by the Transcription Factor DAF-16/FOXO in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Chia-Cheng Wei

    Full Text Available Tea seed oil is a high quality edible oil, yet lacking sufficient scientific evidences to support the nutritional and medical purposes. We identified major and minor components in Camellia tenuifolia seed oil and investigated the antioxidative activity and its underlying mechanisms in Caenorhabditis elegans.The results showed that the major constitutes in C. tenuifolia seed oil were unsaturated fatty acids (~78.4%. Moreover, two minor compounds, β-amyrin and β-sitosterol, were identified and their antioxidative activity was examined. We found that oleic acid was the major constitute in C. tenuifolia seed oil and plays a key role in the antioxidative activity of C. tenuifolia seed oil in C. elegans.This study found evidences that the transcription factor DAF-16/FOXO was involved in both oleic acid- and C. tenuifolia seed oil-mediated oxidative stress resistance in C. elegans. This study suggests the potential of C. tenuifolia seed oil as nutrient or functional foods.

  15. Protein kinase A regulates molecular chaperone transcription and protein aggregation.

    Directory of Open Access Journals (Sweden)

    Yue Zhang

    Full Text Available Heat shock factor 1 (HSF1 regulates one of the major pathways of protein quality control and is essential for deterrence of protein-folding disorders, particularly in neuronal cells. However, HSF1 activity declines with age, a change that may open the door to progression of neurodegenerative disorders such as Huntington's disease. We have investigated mechanisms of HSF1 regulation that may become compromised with age. HSF1 binds stably to the catalytic domain of protein kinase A (PKAcα and becomes phosphorylated on at least one regulatory serine residue (S320. We show here that PKA is essential for effective transcription of HSP genes by HSF1. PKA triggers a cascade involving HSF1 binding to the histone acetylase p300 and positive translation elongation factor 1 (p-TEFb and phosphorylation of the c-terminal domain of RNA polymerase II, a key mechanism in the downstream steps of HSF1-mediated transcription. This cascade appears to play a key role in protein quality control in neuronal cells expressing aggregation-prone proteins with long poly-glutamine (poly-Q tracts. Such proteins formed inclusion bodies that could be resolved by HSF1 activation during heat shock. Resolution of the inclusions was inhibited by knockdown of HSF1, PKAcα, or the pTEFb component CDK9, indicating a key role for the HSF1-PKA cascade in protein quality control.

  16. Mitotic Transcriptional Activation: Clearance of Actively Engaged Pol II via Transcriptional Elongation Control in Mitosis.

    Science.gov (United States)

    Liang, Kaiwei; Woodfin, Ashley R; Slaughter, Brian D; Unruh, Jay R; Box, Andrew C; Rickels, Ryan A; Gao, Xin; Haug, Jeffrey S; Jaspersen, Sue L; Shilatifard, Ali

    2015-11-05

    Although it is established that some general transcription factors are inactivated at mitosis, many details of mitotic transcription inhibition (MTI) and its underlying mechanisms are largely unknown. We have identified mitotic transcriptional activation (MTA) as a key regulatory step to control transcription in mitosis for genes with transcriptionally engaged RNA polymerase II (Pol II) to activate and transcribe until the end of the gene to clear Pol II from mitotic chromatin, followed by global impairment of transcription reinitiation through MTI. Global nascent RNA sequencing and RNA fluorescence in situ hybridization demonstrate the existence of transcriptionally engaged Pol II in early mitosis. Both genetic and chemical inhibition of P-TEFb in mitosis lead to delays in the progression of cell division. Together, our study reveals a mechanism for MTA and MTI whereby transcriptionally engaged Pol II can progress into productive elongation and finish transcription to allow proper cellular division.

  17. Complexes containing activating transcription factor (ATF)/cAMP-responsive-element-binding protein (CREB) interact with the CCAAT/enhancer-binding protein (C/EBP)-ATF composite site to regulate Gadd153 expression during the stress response.

    Science.gov (United States)

    Fawcett, T W; Martindale, J L; Guyton, K Z; Hai, T; Holbrook, N J

    1999-01-01

    Gadd153, also known as chop, encodes a member of the CCAAT/enhancer-binding protein (C/EBP) transcription factor family and is transcriptionally activated by cellular stress signals. We recently demonstrated that arsenite treatment of rat pheochromocytoma PC12 cells results in the biphasic induction of Gadd153 mRNA expression, controlled in part through binding of C/EBPbeta and two uncharacterized protein complexes to the C/EBP-ATF (activating transcription factor) composite site in the Gadd153 promoter. In this report, we identified components of these additional complexes as two ATF/CREB (cAMP-responsive-element-binding protein) transcription factors having differential binding activities dependent upon the time of arsenite exposure. During arsenite treatment of PC12 cells, we observed enhanced binding of ATF4 to the C/EBP-ATF site at 2 h as Gadd153 mRNA levels increased, and enhanced binding of ATF3 complexes at 6 h as Gadd153 expression declined. We further demonstrated that ATF4 activates, while ATF3 represses, Gadd153 promoter activity through the C/EBP-ATF site. ATF3 also repressed ATF4-mediated transactivation and arsenite-induced activation of the Gadd153 promoter. Our results suggest that numerous members of the ATF/CREB family are involved in the cellular stress response, and that regulation of stress-induced biphasic Gadd153 expression in PC12 cells involves the ordered, sequential binding of multiple transcription factor complexes to the C/EBP-ATF composite site. PMID:10085237

  18. Preferential associations between co-regulated genes reveal a transcriptional interactome in erythroid cells.

    Science.gov (United States)

    Schoenfelder, Stefan; Sexton, Tom; Chakalova, Lyubomira; Cope, Nathan F; Horton, Alice; Andrews, Simon; Kurukuti, Sreenivasulu; Mitchell, Jennifer A; Umlauf, David; Dimitrova, Daniela S; Eskiw, Christopher H; Luo, Yanquan; Wei, Chia-Lin; Ruan, Yijun; Bieker, James J; Fraser, Peter

    2010-01-01

    The discovery of interchromosomal interactions in higher eukaryotes points to a functional interplay between genome architecture and gene expression, challenging the view of transcription as a one-dimensional process. However, the extent of interchromosomal interactions and the underlying mechanisms are unknown. Here we present the first genome-wide analysis of transcriptional interactions using the mouse globin genes in erythroid tissues. Our results show that the active globin genes associate with hundreds of other transcribed genes, revealing extensive and preferential intra- and interchromosomal transcription interactomes. We show that the transcription factor Klf1 mediates preferential co-associations of Klf1-regulated genes at a limited number of specialized transcription factories. Our results establish a new gene expression paradigm, implying that active co-regulated genes and their regulatory factors cooperate to create specialized nuclear hot spots optimized for efficient and coordinated transcriptional control.

  19. Calcium regulates caveolin-1 expression at the transcriptional level

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Xiao-Yan; Huang, Cheng-Cheng; Kan, Qi-Ming [Laboratory of Tumor Biology and Glycobiology, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China); Li, Yan [Experimental Animal Center, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China); Liu, Dan; Zhang, Xue-Cheng [Laboratory of Tumor Biology and Glycobiology, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China); Sato, Toshinori [Department of Biosciences and Informatics, Keio University, Hiyoshi, Yokohama 223-8522 (Japan); Yamagata, Sadako [Laboratory of Tumor Biology and Glycobiology, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China); Yamagata, Tatsuya, E-mail: tcyamagata@gmail.com [Laboratory of Tumor Biology and Glycobiology, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China)

    2012-09-28

    Highlights: Black-Right-Pointing-Pointer Caveolin-1 expression is regulated by calcium signaling at the transcriptional level. Black-Right-Pointing-Pointer An inhibitor of or siRNA to L-type calcium channel suppressed caveolin-1 expression. Black-Right-Pointing-Pointer Cyclosporine A or an NFAT inhibitor markedly reduced caveolin-1 expression. Black-Right-Pointing-Pointer Caveolin-1 regulation by calcium signaling is observed in several mouse cell lines. -- Abstract: Caveolin-1, an indispensable component of caveolae serving as a transformation suppressor protein, is highly expressed in poorly metastatic mouse osteosarcoma FBJ-S1 cells while highly metastatic FBJ-LL cells express low levels of caveolin-1. Calcium concentration is higher in FBJ-S1 cells than in FBJ-LL cells; therefore, we investigated the possibility that calcium signaling positively regulates caveolin-1 in mouse FBJ-S1 cells. When cells were treated with the calcium channel blocker nifedipine, cyclosporin A (a calcineurin inhibitor), or INCA-6 (a nuclear factor of activated T-cells [NFAT] inhibitor), caveolin-1 expression at the mRNA and protein levels decreased. RNA silencing of voltage-dependent L-type calcium channel subunit alpha-1C resulted in suppression of caveolin-1 expression. This novel caveolin-1 regulation pathway was also identified in mouse NIH 3T3 cells and Lewis lung carcinoma cells. These results indicate that caveolin-1 is positively regulated at the transcriptional level through a novel calcium signaling pathway mediated by L-type calcium channel/Ca{sup 2+}/calcineurin/NFAT.

  20. Identification of sumoylation sites in CCDC6, the first identified RET partner gene in papillary thyroid carcinoma, uncovers a mode of regulating CCDC6 function on CREB1 transcriptional activity.

    Directory of Open Access Journals (Sweden)

    Chiara Luise

    Full Text Available CCDC6 was originally identified in chimeric genes as caused by chromosomal translocation involving the RET protooncogene in some thyroid tumors. Recognised as a 65 kDa pro-apoptotic phosphoprotein, CCDC6 has been enrolled as an ATM substrate that contribute to protect genome integrity by modulating PP4c activity in response to genotoxic stress. Recently, CCDC6 has been identified as a repressor of CREB1-dependent transcription. Sumoylation has emerged as an important mechanism in transcriptional control. Here, we report the identification and characterization of three sites of sumoylation in CCDC6 (K74, K266 and K424 which are highly conserved in vertebrates. We demonstrate that the post-translational modifications by SUMO2 constrain most of the CCDC6 protein in the cytosol and affect its functional interaction with CREB1 with a decrease of CCDC6 repressive function on CREB1 transcriptional activity. Indeed, the impairment of functional outcome of sumoylated CCDC6 is obtained knocking down all three the sumoylation sites. Interestingly, in thyroid cells the SUMO2-mediated CCDC6 post-translational modifications are induced by Forskolin, a cAMP analog. Signal transduction via the cAMP pathway is known to be ubiquitous and represents a major line of communication between many organisms and their environment. We believe that CCDC6 could be an important player in the dynamics of cAMP signaling by fine regulating CREB1 transcriptional activity in normal and transformed thyroid cells.

  1. Human mediator subunit MED15 promotes transcriptional activation.

    Science.gov (United States)

    Nakatsubo, Takuya; Nishitani, Saori; Kikuchi, Yuko; Iida, Satoshi; Yamada, Kana; Tanaka, Aki; Ohkuma, Yoshiaki

    2014-10-01

    In eukaryotes, the Mediator complex is an essential transcriptional cofactor of RNA polymerase II (Pol II). In humans, it contains up to 30 subunits and consists of four modules: head, middle, tail, and CDK/Cyclin. One of the subunits, MED15, is located in the tail module, and was initially identified as Gal11 in budding yeast, where it plays an essential role in the transcriptional regulation of galactose metabolism with the potent transcriptional activator Gal4. For this reason, we investigated the function of the human MED15 subunit (hMED15) in transcriptional activation. First, we measured the effect of hMED15 knockdown on cell growth in HeLa cells. The growth rate was greatly reduced. By immunostaining, we observed the colocalization of hMED15 with the general transcription factors TFIIE and TFIIH in the nucleus. We measured the effects of siRNA-mediated knockdown of hMED15 on transcriptional activation using two different transcriptional activators, VP16 and SREBP1a. Treatment with siRNAs reduced transcriptional activation, and this reduction could be rescued by overexpression of HA/Flag-tagged, wild-type hMED15. To investigate hMED15 localization, we treated human MCF-7 cells with the MDM2 inhibitor Nutlin-3, thus inducing p21 transcription. We found that hMED15 localized to both the p53 binding site and the p21 promoter region, along with TFIIE and TFIIH. These results indicate that hMED15 promotes transcriptional activation.

  2. Modeling Writing Development: Contribution of Transcription and Self-Regulation to Portuguese Students' Text Generation Quality

    Science.gov (United States)

    Limpo, Teresa; Alves, Rui A.

    2013-01-01

    Writing is a complex activity that requires transcription and self-regulation. We used multiple-group structural equation modeling to test the contribution of transcription (handwriting and spelling), planning, revision, and self-efficacy to writing quality at 2 developmental points (Grades 4-6 vs. 7-9). In Grades 4-6, the model explained 76% of…

  3. Sumoylation delays the ATF7 transcription factor subcellular localization and inhibits its transcriptional activity.

    Science.gov (United States)

    Hamard, Pierre-Jacques; Boyer-Guittaut, Michaël; Camuzeaux, Barbara; Dujardin, Denis; Hauss, Charlotte; Oelgeschläger, Thomas; Vigneron, Marc; Kedinger, Claude; Chatton, Bruno

    2007-01-01

    Over the past few years, small ubiquitin-like modifier (SUMO) modification has emerged as an important regulator of diverse pathways and activities including protein localization and transcriptional regulation. We identified a consensus sumoylation motif (IKEE), located within the N-terminal activation domain of the ATF7 transcription factor and thus investigated the role of this modification. ATF7 is a ubiquitously expressed transcription factor, homologous to ATF2, that binds to CRE elements within specific promoters. This protein is able to heterodimerize with Jun or Fos proteins and its transcriptional activity is mediated by interaction with TAF12, a subunit of the general transcription factor TFIID. In the present article, we demonstrate that ATF7 is sumoylated in vitro (using RanBP2 as a E3-specific ligase) and in vivo. Moreover, we show that ATF7 sumoylation affects its intranuclear localization by delaying its entry into the nucleus. Furthermore, SUMO conjugation inhibits ATF7 transactivation activity by (i) impairing its association with TAF12 and (ii) blocking its binding-to-specific sequences within target promoters.

  4. MaJAZ1 Attenuates the MaLBD5-Mediated Transcriptional Activation of Jasmonate Biosynthesis Gene MaAOC2 in Regulating Cold Tolerance of Banana Fruit.

    Science.gov (United States)

    Ba, Liang-jie; Kuang, Jian-fei; Chen, Jian-ye; Lu, Wang-jin

    2016-02-01

    Previous studies indicated that methyl jasmonate (MeJA) treatment could effectively reduce the chilling injury of many fruits, including banana, but the underlying mechanism is poorly understood. In this study, one lateral organ boundaries (LOB) domain (LBD) gene, designated as MaLBD5, was isolated and characterized from banana fruit. Expression analysis revealed that accumulation of MaLBD5 was induced by cold temperature and MeJA treatment. Subcellular localization and transactivation assays showed that MaLBD5 was localized to the nucleus and possessed transcriptional activation activity. Protein-protein interaction analysis demonstrated that MaLBD5 physically interacted with MaJAZ1, a potential repressor of jasmonate signaling. Furthermore, transient expression assays indicated that MaLBD5 transactivated a jasmonate biosynthesis gene, termed MaAOC2, which was also induced by cold and MeJA. More interestingly, MaJAZ1 attenuated the MaLBD5-mediated transactivation of MaAOC2. These results suggest that MaLBD5 and MaJAZ1 might act antagonistically in relation to MeJA-induced cold tolerance of banana fruit, at least partially via affecting jasmonate biosynthesis. Collectively, our findings expand the knowledge of the transcriptional regulatory network of MeJA-mediated cold tolerance of banana fruit.

  5. FRUITING GENES OF SCHIZOPHYLLUM-COMMUNE ARE TRANSCRIPTIONALLY REGULATED

    NARCIS (Netherlands)

    SCHUREN, FHJ; VANDERLENDE, TR; WESSELS, JGH

    Fruiting genes in Schizophyllum commune are controlled by the mating-type genes and other regulatory genes. To examine whether differential accumulation of mRNAs for these fruiting genes is caused by transcriptional regulation, run-on transcription assaYs were performed with nuclei isolated from

  6. FRUITING GENES OF SCHIZOPHYLLUM-COMMUNE ARE TRANSCRIPTIONALLY REGULATED

    NARCIS (Netherlands)

    SCHUREN, FHJ; VANDERLENDE, TR; WESSELS, JGH

    1993-01-01

    Fruiting genes in Schizophyllum commune are controlled by the mating-type genes and other regulatory genes. To examine whether differential accumulation of mRNAs for these fruiting genes is caused by transcriptional regulation, run-on transcription assaYs were performed with nuclei isolated from cul

  7. FRUITING GENES OF SCHIZOPHYLLUM-COMMUNE ARE TRANSCRIPTIONALLY REGULATED

    NARCIS (Netherlands)

    SCHUREN, FHJ; VANDERLENDE, TR; WESSELS, JGH

    1993-01-01

    Fruiting genes in Schizophyllum commune are controlled by the mating-type genes and other regulatory genes. To examine whether differential accumulation of mRNAs for these fruiting genes is caused by transcriptional regulation, run-on transcription assaYs were performed with nuclei isolated from cul

  8. Heterogeneity of Calcium Channel/cAMP-Dependent Transcriptional Activation.

    Science.gov (United States)

    Kobrinsky, Evgeny

    2015-01-01

    The major function of the voltage-gated calcium channels is to provide the Ca(2+) flux into the cell. L-type voltage-gated calcium channels (Cav1) serve as voltage sensors that couple membrane depolarization to many intracellular processes. Electrical activity in excitable cells affects gene expression through signaling pathways involved in the excitation-transcription (E-T) coupling. E-T coupling starts with activation of the Cav1 channel and results in initiation of the cAMP-response element binding protein (CREB)-dependent transcription. In this review we discuss the new quantitative approaches to measuring E-T signaling events. We describe the use of wavelet transform to detect heterogeneity of transcriptional activation in nuclei. Furthermore, we discuss the properties of discovered microdomains of nuclear signaling associated with the E-T coupling and the basis of the frequency-dependent transcriptional regulation.

  9. Regulation of transcription functions of the p53 tumor suppressor by the mdm-2 oncogene.

    OpenAIRE

    1995-01-01

    BACKGROUND: Mdm-2, a zinc finger protein, negatively regulates the p53 tumor suppressor gene product by binding to it and preventing transcriptional activation (16). MATERIALS AND METHODS: Assays for p53 mediated transcription, repression and activation by mutant and wild-type p53 proteins were used to measure the ability of mdm-2 to block each activity. RESULTS: Mdm-2 was able to inhibit all three functions of the wild-type and mutant p53 activities; transcriptional activation by the wild-ty...

  10. RNA-binding proteins involved in post-transcriptional regulation in bacteria

    Directory of Open Access Journals (Sweden)

    Elke eVan Assche

    2015-03-01

    Full Text Available Post-transcriptional regulation is a very important mechanism to control gene expression in changing environments. In the past decade, a lot of interest has been directed towards the role of small RNAs in bacterial post-transcriptional regulation. However, small RNAs are not the only molecules controlling gene expression at this level, RNA-binding proteins play an important role as well. CsrA and Hfq are the two best studied bacterial proteins of this type, but recently, additional proteins involved in post-transcriptional control have been identified. This review focuses on the general working mechanisms of post-transcriptionally active RNA-binding proteins, which include (i adaptation of the susceptibility of mRNAs and sRNAs to RNases, (ii modulating the accessibility of the ribosome binding site of mRNAs, (iii recruiting and assisting in the interaction of mRNAs with other molecules and (iv regulating transcription terminator / antiterminator formation, and gives an overview of both the well-studied and the newly identified proteins that are involved in post-transcriptional regulatory processes. Additionally, the post-transcriptional mechanisms by which the expression or the activity of these proteins is regulated, are described. For many of the newly identified proteins, however, mechanistic questions remain. Most likely, more post-transcriptionally active proteins will be identified in the future.

  11. Tissue-specific 5' heterogeneity of PPARα transcripts and their differential regulation by leptin.

    Science.gov (United States)

    Garratt, Emma S; Vickers, Mark H; Gluckman, Peter D; Hanson, Mark A; Burdge, Graham C; Lillycrop, Karen A

    2013-01-01

    The genes encoding nuclear receptors comprise multiple 5'untranslated exons, which give rise to several transcripts encoding the same protein, allowing tissue-specific regulation of expression. Both human and mouse peroxisome proliferator activated receptor (PPAR) α genes have multiple promoters, although their function is unknown. Here we have characterised the rat PPARα promoter region and have identified three alternative PPARα transcripts, which have different transcription start sites owing to the utilisation of distinct first exons. Moreover these alternative PPARα transcripts were differentially expressed between adipose tissue and liver. We show that while the major adipose (P1) and liver (P2) transcripts were both induced by dexamethasone, they were differentially regulated by the PPARα agonist, clofibric acid, and leptin. Leptin had no effect on the adipose-specific P1 transcript, but induced liver-specific P2 promoter activity via a STAT3/Sp1 mechanism. Moreover in Wistar rats, leptin treatment between postnatal day 3-13 led to an increase in P2 but not P1 transcription in adipose tissue which was sustained into adulthood. This suggests that the expression of the alternative PPARα transcripts are in part programmed by early life exposure to leptin leading to persistent change in adipose tissue fatty acid metabolism through specific activation of a quiescent PPARα promoter. Such complexity in the regulation of PPARα may allow the expression of PPARα to be finely regulated in response to environmental factors.

  12. Codependent activators direct myoblast-specific MyoD transcription.

    Science.gov (United States)

    Hu, Ping; Geles, Kenneth G; Paik, Ji-Hye; DePinho, Ronald A; Tjian, Robert

    2008-10-01

    Although FoxO and Pax proteins represent two important families of transcription factors in determining cell fate, they had not been functionally or physically linked together in mediating regulation of a common target gene during normal cellular transcription programs. Here, we identify MyoD, a key regulator of myogenesis, as a direct target of FoxO3 and Pax3/7 in myoblasts. Our cell-based assays and in vitro studies reveal a tight codependent partnership between FoxO3 and Pax3/7 to coordinately recruit RNA polymerase II and form a preinitiation complex (PIC) to activate MyoD transcription in myoblasts. The role of FoxO3 in regulating muscle differentiation is confirmed in vivo by observed defects in muscle regeneration caused by MyoD downregulation in FoxO3 null mice. These data establish a mutual interdependence and functional link between two families of transcription activators serving as potential signaling sensors and regulators of cell fate commitment in directing tissue specific MyoD transcription.

  13. SOX6 attenuates glucose-stimulated insulin secretion by repressing PDX1 transcriptional activity and is down-regulated in hyperinsulinemic obese mice.

    Science.gov (United States)

    Iguchi, Haruhisa; Ikeda, Yukio; Okamura, Masashi; Tanaka, Toshiya; Urashima, Yasuyo; Ohguchi, Hiroto; Takayasu, Shinobu; Kojima, Noriaki; Iwasaki, Satoshi; Ohashi, Riuko; Jiang, Shuying; Hasegawa, Go; Ioka, Ryoichi X; Magoori, Kenta; Sumi, Koichi; Maejima, Takashi; Uchida, Aoi; Naito, Makoto; Osborne, Timothy F; Yanagisawa, Masashi; Yamamoto, Tokuo T; Kodama, Tatsuhiko; Sakai, Juro

    2005-11-11

    In obesity-related insulin resistance, pancreatic islets compensate for insulin resistance by increasing secretory capacity. Here, we report the identification of sex-determining region Y-box 6 (SOX6), a member of the high mobility group box superfamily of transcription factors, as a co-repressor for pancreatic-duodenal homeobox factor-1 (PDX1). SOX6 mRNA levels were profoundly reduced by both a long term high fat feeding protocol in normal mice and in genetically obese ob/ob mice on a normal chow diet. Interestingly, we show that SOX6 is expressed in adult pancreatic insulin-producing beta-cells and that overexpression of SOX6 decreased glucose-stimulated insulin secretion, which was accompanied by decreased ATP/ADP ratio, Ca(2+) mobilization, proinsulin content, and insulin gene expression. In a complementary fashion, depletion of SOX6 by small interfering RNAs augmented glucose-stimulated insulin secretion in insulinoma mouse MIN6 and rat INS-1E cells. These effects can be explained by our mechanistic studies that show SOX6 acts to suppress PDX1 stimulation of the insulin II promoter through a direct protein/protein interaction. Furthermore, SOX6 retroviral expression decreased acetylation of histones H3 and H4 in chromatin from the promoter for the insulin II gene, suggesting that SOX6 may decrease PDX1 stimulation through changes in chromatin structure at specific promoters. These results suggest that perturbations in transcriptional regulation that are coordinated through SOX6 and PDX1 in beta-cells may contribute to the beta-cell adaptation in obesity-related insulin resistance.

  14. Post-Transcriptional Regulation of Cytokine Signaling by AU-Rich and GU-Rich Elements

    Science.gov (United States)

    Bohjanen, Paul R.

    2014-01-01

    Cytokines are necessary for cell communication to enable responses to external stimuli that are imperative for the survival and maintenance of homeostasis. Dysfunction of the cytokine network has detrimental effects on intra- and extracellular environments. Thus, it is critical that the expression of cytokines and the signals transmitted by cytokines to target cells are tightly regulated at numerous levels, including transcriptional and post-transcriptional levels. Here, we briefly summarize the role of AU-rich elements (AREs) in the regulation of cytokine gene expression at the post-transcriptional level and describe a role for GU-rich elements (GREs) in coordinating the regulation of cytokine signaling. GREs function as post-transcriptional regulators of proteins that control cellular activation, growth, and apoptosis. GREs and AREs work in concert to coordinate cytokine signal transduction pathways. The precise regulation of cytokine signaling is particularly important, because its dysregulation can lead to human diseases. PMID:24697201

  15. Bmp indicator mice reveal dynamic regulation of transcriptional response.

    Directory of Open Access Journals (Sweden)

    Anna L Javier

    Full Text Available Cellular responses to Bmp ligands are regulated at multiple levels, both extracellularly and intracellularly. Therefore, the presence of these growth factors is not an accurate indicator of Bmp signaling activity. While a common approach to detect Bmp signaling activity is to determine the presence of phosphorylated forms of Smad1, 5 and 8 by immunostaining, this approach is time consuming and not quantitative. In order to provide a simpler readout system to examine the presence of Bmp signaling in developing animals, we developed BRE-gal mouse embryonic stem cells and a transgenic mouse line that specifically respond to Bmp ligand stimulation. Our reporter identifies specific transcriptional responses that are mediated by Smad1 and Smad4 with the Schnurri transcription factor complex binding to a conserved Bmp-Responsive Element (BRE, originally identified among Drosophila, Xenopus and human Bmp targets. Our BRE-gal mES cells specifically respond to Bmp ligands at concentrations as low as 5 ng/ml; and BRE-gal reporter mice, derived from the BRE-gal mES cells, show dynamic activity in many cellular sites, including extraembryonic structures and mammary glands, thereby making this a useful scientific tool.

  16. Activation of the MAPK11/12/13/14 (p38 MAPK) pathway regulates the transcription of autophagy genes in response to oxidative stress induced by a novel copper complex in HeLa cells.

    Science.gov (United States)

    Zhong, Wu; Zhu, Haichuan; Sheng, Fugeng; Tian, Yonglu; Zhou, Jun; Chen, Yingyu; Li, Song; Lin, Jian

    2014-07-01

    Transition metal copper (Cu) can exist in oxidized or reduced states in cells, leading to cytotoxicity in cancer cells through oxidative stress. Recently, copper complexes are emerging as a new class of anticancer compounds. Here, we report that a novel anticancer copper complex (HYF127c/Cu) induces oxidative stress-dependent cell death in cancer cells. Further, transcriptional analysis revealed that oxidative stress elicits broad transcriptional changes of genes, in which autophagy-related genes are significantly changed in HYF127c/Cu-treated cells. Consistently, autophagy was induced in HYF127c/Cu-treated cells and inhibitors of autophagy promoted cell death induced by HYF127c/Cu. Further analysis identified that the MAPK11/12/13/14 (formerly known as p38 MAPK) pathway was also activated in HYF127c/Cu-treated cells. Meanwhile, the MAPK11/12/13/14 inhibitor SB203580 downregulated autophagy by inhibiting the transcription of the autophagy genes MAP1LC3B, BAG3, and HSPA1A, and promoted HYF127c/Cu-induced cell death. These data suggest that copper-induced oxidative stress will induce protective autophagy through transcriptional regulation of autophagy genes by activation of the MAPK11/12/13/14 pathway in HeLa cells.

  17. Axon Regeneration Is Regulated by Ets–C/EBP Transcription Complexes Generated by Activation of the cAMP/Ca2+ Signaling Pathways

    Science.gov (United States)

    Matsumoto, Kunihiro

    2015-01-01

    The ability of specific neurons to regenerate their axons after injury is governed by cell-intrinsic regeneration pathways. In Caenorhabditis elegans, the JNK and p38 MAPK pathways are important for axon regeneration. Axonal injury induces expression of the svh-2 gene encoding a receptor tyrosine kinase, stimulation of which by the SVH-1 growth factor leads to activation of the JNK pathway. Here, we identify ETS-4 and CEBP-1, related to mammalian Ets and C/EBP, respectively, as transcriptional activators of svh-2 expression following axon injury. ETS-4 and CEBP-1 function downstream of the cAMP and Ca2+–p38 MAPK pathways, respectively. We show that PKA-dependent phosphorylation of ETS-4 promotes its complex formation with CEBP-1. Furthermore, activation of both cAMP and Ca2+ signaling is required for activation of svh-2 expression. Thus, the cAMP/Ca2+ signaling pathways cooperatively activate the JNK pathway, which then promotes axon regeneration. PMID:26484536

  18. Axon Regeneration Is Regulated by Ets-C/EBP Transcription Complexes Generated by Activation of the cAMP/Ca2+ Signaling Pathways.

    Directory of Open Access Journals (Sweden)

    Chun Li

    2015-10-01

    Full Text Available The ability of specific neurons to regenerate their axons after injury is governed by cell-intrinsic regeneration pathways. In Caenorhabditis elegans, the JNK and p38 MAPK pathways are important for axon regeneration. Axonal injury induces expression of the svh-2 gene encoding a receptor tyrosine kinase, stimulation of which by the SVH-1 growth factor leads to activation of the JNK pathway. Here, we identify ETS-4 and CEBP-1, related to mammalian Ets and C/EBP, respectively, as transcriptional activators of svh-2 expression following axon injury. ETS-4 and CEBP-1 function downstream of the cAMP and Ca2+-p38 MAPK pathways, respectively. We show that PKA-dependent phosphorylation of ETS-4 promotes its complex formation with CEBP-1. Furthermore, activation of both cAMP and Ca2+ signaling is required for activation of svh-2 expression. Thus, the cAMP/Ca2+ signaling pathways cooperatively activate the JNK pathway, which then promotes axon regeneration.

  19. Axon Regeneration Is Regulated by Ets-C/EBP Transcription Complexes Generated by Activation of the cAMP/Ca2+ Signaling Pathways.

    Science.gov (United States)

    Li, Chun; Hisamoto, Naoki; Matsumoto, Kunihiro

    2015-10-01

    The ability of specific neurons to regenerate their axons after injury is governed by cell-intrinsic regeneration pathways. In Caenorhabditis elegans, the JNK and p38 MAPK pathways are important for axon regeneration. Axonal injury induces expression of the svh-2 gene encoding a receptor tyrosine kinase, stimulation of which by the SVH-1 growth factor leads to activation of the JNK pathway. Here, we identify ETS-4 and CEBP-1, related to mammalian Ets and C/EBP, respectively, as transcriptional activators of svh-2 expression following axon injury. ETS-4 and CEBP-1 function downstream of the cAMP and Ca2+-p38 MAPK pathways, respectively. We show that PKA-dependent phosphorylation of ETS-4 promotes its complex formation with CEBP-1. Furthermore, activation of both cAMP and Ca2+ signaling is required for activation of svh-2 expression. Thus, the cAMP/Ca2+ signaling pathways cooperatively activate the JNK pathway, which then promotes axon regeneration.

  20. Chromatin insulation by a transcriptional activator.

    Science.gov (United States)

    Sutter, Nathan B; Scalzo, David; Fiering, Steven; Groudine, Mark; Martin, David I K

    2003-02-04

    In eukaryotic genomes, transcriptionally active regions are interspersed with silent chromatin that may repress genes in its vicinity. Chromatin insulators are elements that can shield a locus from repressive effects of flanking chromatin. Few such elements have been characterized in higher eukaryotes, but transcriptional activating elements are an invariant feature of active loci and have been shown to suppress transgene silencing. Hence, we have assessed the ability of a transcriptional activator to cause chromatin insulation, i.e., to relieve position effects at transgene integration sites in cultured cells. The transgene contained a series of binding sites for the metal-inducible transcriptional activator MTF, linked to a GFP reporter. Clones carrying single integrated transgenes were derived without selection for expression, and in most clones the transgene was silent. Induction of MTF resulted in transition of the transgene from the silent to the active state, prolongation of the active state, and a marked narrowing of the range of expression levels at different genomic sites. At one genomic site, prolonged induction of MTF resulted in suppression of transgene silencing that persisted after withdrawal of the induction stimulus. These results are consistent with MTF acting as a chromatin insulator and imply that transcriptional activating elements can insulate active loci against chromatin repression.

  1. Human herpesvirus 6B U19 protein is a PML-regulated transcriptional activator that localizes to nuclear foci in a PML-independent manner

    DEFF Research Database (Denmark)

    Kofod-Olsen, Emil; Ross-Hansen, Katrine; Mikkelsen, Jacob Giehm

    2008-01-01

    Human herpesvirus 6B (HHV-6B) contains an IE-B domain spanning open reading frames U16/17-U19, based on homology with human cytomegalovirus. Here, the protein product, U19, of the HHV-6B U19 gene is identified as a 47 kDa transcriptional activator. HHV-6B infection or overexpression of U19...... transactivated the RANTES promoter. Mutational analysis of the promoter indicated that transactivation was not critically dependent on the promoter sites CRE, NF-kappaB, ISRE or NF-IL6. ND10 are nuclear substructures that are involved in several cellular regulatory pathways, including those controlling gene...... structure, U19 also localized to the centre of ND10. Knockdown of PML by small interfering RNA did not prevent U19 localization to ND10-like foci, but instead led to a fourfold increase in U19-induced transcription from the RANTES promoter. Generation of four truncated U19 proteins indicated that the N...

  2. Evolution of transcriptional regulation in "Escherichia coli"

    OpenAIRE

    Wolf, Luise

    2014-01-01

    During gene expression, transcription initiation marks the first step towards synthesis of functional proteins. Expression levels of specific types of RNA molecules in the cell depend on the underlying genotype of the promoter sequence. Prediction of expression levels from the promoter sequence alone can have important implications for the design of artificial promoters. In this work, we explored promoter determinants that cause differences in expression levels and tracked how ...

  3. Insights into the regulation of transcription by scanning force microscopy.

    Science.gov (United States)

    Dame, R T; Wyman, C; Goosen, N

    2003-12-01

    The scanning force microscope (SFM) is a valuable tool for the structural analysis of complexes between protein(s) and DNA. In recent years the application of scanning force microscopy to the field of transcription regulation has been reported in numerous studies. Using this technique, novel insights could be obtained into the architecture and dynamics of complexes, which are relevant to the transcription process and the mechanisms by which this process is regulated. In this article an overview is given of SFM studies addressing, in particular, topics in the field of transcription in prokaryotic organisms.

  4. Regulation of Na(+)/K(+)-ATPase by neuron-specific transcription factor Sp4: implication in the tight coupling of energy production, neuronal activity and energy consumption in neurons.

    Science.gov (United States)

    Johar, Kaid; Priya, Anusha; Wong-Riley, Margaret T T

    2014-02-01

    A major source of energy demand in neurons is the Na(+)/K(+)-ATPase pump that restores the ionic gradient across the plasma membrane subsequent to depolarizing neuronal activity. The energy comes primarily from mitochondrial oxidative metabolism, of which cytochrome c oxidase (COX) is a key enzyme. Recently, we found that all 13 subunits of COX are regulated by specificity (Sp) factors, and that the neuron-specific Sp4, but not Sp1 or Sp3, regulates the expression of key glutamatergic receptor subunits as well. The present study sought to test our hypothesis that Sp4 also regulates Na(+)/K(+)-ATPase subunit genes in neurons. By means of multiple approaches, including in silico analysis, electrophoretic mobility shift and supershift assays, chromatin immunoprecipitation, promoter mutational analysis, over-expression, and RNA interference studies, we found that Sp4, with minor contributions from Sp1 and Sp3, functionally regulate the Atp1a1, Atp1a3, and Atp1b1 subunit genes of Na(+)/K(+)-ATPase in neurons. Transcripts of all three genes were up-regulated by depolarizing KCl stimulation and down-regulated by the impulse blocker tetrodotoxin (TTX), indicating that their expression was activity-dependent. Silencing of Sp4 blocked the up-regulation of these genes induced by KCl, whereas over-expression of Sp4 rescued them from TTX-induced suppression. The effect of silencing or over-expressing Sp4 on primary neurons was much greater than those of Sp1 or Sp3. The binding sites of Sp factors on these genes are conserved among mice, rats and humans. Thus, Sp4 plays an important role in the transcriptional coupling of energy generation and energy consumption in neurons.

  5. Transcriptional regulation of bone sialoprotein gene by interleukin-11.

    Science.gov (United States)

    Wang, Shuang; Sasaki, Yoko; Zhou, Liming; Matsumura, Hiroyoshi; Araki, Shouta; Mezawa, Masaru; Takai, Hideki; Chen, Zhen; Ogata, Yorimasa

    2011-05-01

    Interleukin-11 (IL-11) is a stromal cell-derived cytokine that belongs to the interleukin-6 family of cytokines. IL-11 has many biological activities and has roles in hematopoiesis, immune responses, the nervous system and bone metabolism. Bone sialoprotein (BSP) is a mineralized tissue-specific protein expressed in differentiated osteoblasts that appears to function in the initial mineralization of bone. IL-11 (20 ng/ml) increased BSP mRNA and protein levels at 12h in osteoblast-like ROS 17/2.8 cells. In a transient transfection assay, IL-11 (20 ng/ml) increased luciferase activity of the construct (-116 to +60) in ROS 17/2.8 cells and rat bone marrow stromal cells. Introduction of 2 bp mutations to the luciferase constructs showed that the effects of IL-11 were mediated by a cAMP response element (CRE), a fibroblast growth factor 2 response element (FRE) and a homeodomain protein-binding site (HOX). Luciferase activities induced by IL-11 were blocked by protein kinase A inhibitor, tyrosine kinase inhibitor and ERK1/2 inhibitor. Gel shift analyses showed that IL-11 (20 ng/ml) increased nuclear protein binding to CRE, FRE and HOX. CREB1, phospho-CREB1, c-Fos, c-Jun, JunD and Fra2 antibodies disrupted the formation of CRE-protein complexes. Dlx5, Msx2, Runx2 and Smad1 antibodies disrupted FRE- and HOX-protein complex formations. These studies demonstrate that IL-11 stimulates BSP transcription by targeting CRE, FRE and HOX sites in the proximal promoter of the rat BSP gene. Moreover, phospho-CREB1, c-Fos, c-Jun, JunD, Fra2, Dlx5, Msx2, Runx2 and Smadl transcription factors appear to be key regulators of IL-11 effects on BSP transcription.

  6. Transcription factor FOXA2-centered transcriptional regulation network in non-small cell lung cancer.

    Science.gov (United States)

    Jang, Sang-Min; An, Joo-Hee; Kim, Chul-Hong; Kim, Jung-Woong; Choi, Kyung-Hee

    2015-08-01

    Lung cancer is the leading cause of cancer-mediated death. Although various therapeutic approaches are used for lung cancer treatment, these mainly target the tumor suppressor p53 transcription factor, which is involved in apoptosis and cell cycle arrest. However, p53-targeted therapies have limited application in lung cancer, since p53 is found to be mutated in more than half of lung cancers. In this study, we propose tumor suppressor FOXA2 as an alternative target protein for therapies against lung cancer and reveal a possible FOXA2-centered transcriptional regulation network by identifying new target genes and binding partners of FOXA2 by using various screening techniques. The genes encoding Glu/Asp-rich carboxy-terminal domain 2 (CITED2), nuclear receptor subfamily 0, group B, member 2 (NR0B2), cell adhesion molecule 1 (CADM1) and BCL2-associated X protein (BAX) were identified as putative target genes of FOXA2. Additionally, the proteins including highly similar to heat shock protein HSP 90-beta (HSP90A), heat shock 70 kDa protein 1A variant (HSPA1A), histone deacetylase 1 (HDAC1) and HDAC3 were identified as novel interacting partners of FOXA2. Moreover, we showed that FOXA2-dependent promoter activation of BAX and p21 genes is significantly reduced via physical interactions between the identified binding partners and FOXA2. These results provide opportunities to understand the FOXA2-centered transcriptional regulation network and novel therapeutic targets to modulate this network in p53-deficient lung cancer.

  7. Transcriptional responses and regulations to deficient phosphorus in plants

    Institute of Scientific and Technical Information of China (English)

    Jinxiang BAO; Shuhua ZHANG; Wenjing LU; Chengjin GUO; Juntao GU; Kai XIAO

    2009-01-01

    Significant progress has been made over the past several years in the understanding of phosphorus (Pi)-starvation responses in plants and their regulation. The transcriptional changes that occur in response to Pi starvation are beginning to be revealed, although much is left to understand about their significance. In this paper, the recent progresses on the gene expression changes under deficient-Pi, cis-regulatory elements involved in response to deficient-Pi, the transcriptional control of Pi-starvation responses in eukaryotes, transcription factors involved in response to Pi-starvation, the role of MicroRNA on regulation of phosphate homeostasis, and phosphate sensing and signal transduction in plants have been summarized. The purpose of this review is to provide some basis for further elucidation of the transcriptional responses and regulations, and the networks of Pi sensing and signal transduction under deficient-Pi in plants in the future.

  8. The N-terminus region of the putative C2H2 transcription factor Ada1 harbors a species-specific activation motif that regulates asexual reproduction in Fusarium verticillioides.

    Science.gov (United States)

    Malapi-Wight, Martha; Kim, Jung-Eun; Shim, Won-Bo

    2014-01-01

    Fusarium verticillioides is an important plant pathogenic fungus causing maize ear and stalk rots. In addition, the fungus is directly associated with fumonisin contamination of food and feeds. Here, we report the functional characterization of Ada1, a putative Cys2-His2 zinc finger transcription factor with a high level of similarity to Aspergillus nidulans FlbC, which is required for the activation of the key regulator of conidiation brlA. ADA1 is predicted to encode a protein with two DNA binding motifs at the C terminus and a putative activator domain at the N terminus region. Deletion of the flbC gene in A. nidulans results in "fluffy" cotton-like colonies, with a defect in transition from vegetative growth to asexual development. In this study we show that Ada1 plays a key role in asexual development in F. verticillioides. Conidia production was significantly reduced in the knockout mutant (Δada1), in which aberrant conidia and conidiophores were also observed. We identified genes that are predicted to be downstream of ADA1, based on A. nidulans conidiation signaling pathway. Among them, the deletion of stuA homologue, FvSTUA, resulted in near absence of conidia production. To further investigate the functional conservation of this transcription factor, we complemented the Δada1 strain with A. nidulans flbC, F. verticillioides ADA1, and chimeric constructs. A. nidulans flbC failed to restore conidia production similar to the wild-type level. However, the Ada1N-terminal domain, which contains a putative activator, fused to A. nidulans FlbC C-terminal motif successfully complemented the Δada1 mutant. Taken together, Ada1 is an important transcriptional regulator of asexual development in F. verticillioides and that the N-terminus domain is critical for proper function of this transcription factor.

  9. Conserved regulation of the Hansenula polymorpha MOX promoter in Saccharomyces cerevisiae reveals insights in the transcriptional activation by Adr1p.

    Science.gov (United States)

    Pereira, G G; Hollenberg, C P

    1996-05-15

    The Hansenula polymorpha MOX gene encodes a peroxisomal enzyme that catalyzes the first step of the highly specialized methanol metabolism. MOX is strongly transcribed in cells growing in methanol and completely repressed in glucose. We show here that the MOX promoter confers a glucose-repressible expression upon a lacZ reporter gene in Saccharomyces cerevisiae, an unrelated yeast species that lacks the methanol metabolism. Repression was mediated by a 200-bp region of the MOX promoter, termed MOX-B, and was counteracted by Adr1p, a transcription factor involved in the derepression of S. cerevisiae genes encoding peroxisomal proteins, the class to which MOX belongs. Binding of Adr1p to MOX-B was demonstrated by gel retardation and DNaseI-footprinting, and Adr1p was shown to interact with a DNA region containing only a half of the putative Adr1p consensus binding site. Our findings suggest that Adr1p is a conserved regulator for genes encoding peroxisomal proteins at least in other yeast species, and that its interaction with the DNA is dependent on the promoter context.

  10. Potential Role of Activating Transcription Factor 5 during Osteogenesis

    Directory of Open Access Journals (Sweden)

    Luisa Vicari

    2016-01-01

    Full Text Available Human adipose-derived stem cells are an abundant population of stem cells readily isolated from human adipose tissue that can differentiate into connective tissue lineages including bone, cartilage, fat, and muscle. Activating transcription factor 5 is a transcription factor of the ATF/cAMP response element-binding protein (CREB family. It is transcribed in two types of mRNAs (activating transcription factor 5 isoform 1 and activating transcription factor 5 isoform 2, encoding the same single 30-kDa protein. Although it is well demonstrated that it regulates the proliferation, differentiation, and apoptosis, little is known about its potential role in osteogenic differentiation. The aim of this study was to evaluate the expression levels of the two isoforms and protein during osteogenic differentiation of human adipose-derived stem cells. Our data indicate that activating transcription factor 5 is differentially expressed reaching a peak of expression at the stage of bone mineralization. These findings suggest that activating transcription factor 5 could play an interesting regulatory role during osteogenesis, which would provide a powerful tool to study bone physiology.

  11. Structural basis for transcription regulation by alarmone ppGpp.

    Science.gov (United States)

    Artsimovitch, Irina; Patlan, Vsevolod; Sekine, Shun-ichi; Vassylyeva, Marina N; Hosaka, Takeshi; Ochi, Kozo; Yokoyama, Shigeyuki; Vassylyev, Dmitry G

    2004-04-30

    Guanosine-tetraphosphate (ppGpp) is a major regulator of stringent control, an adaptive response of bacteria to amino acid starvation. The 2.7 A resolution structure of the Thermus thermophilus RNA polymerase (RNAP) holoenzyme in complex with ppGpp reveals that ppGpp binds to the same site near the active center in both independent RNAP molecules in the crystal but in strikingly distinct orientations. Binding is symmetrical with respect to the two diphosphates of ppGpp and is relaxed with respect to the orientation of the nucleotide base. Different modes of ppGpp binding are coupled with asymmetry of the active site configurations. The results suggest that base pairing of ppGpp with cytosines in the nontemplate DNA strand might be an essential component of transcription control by ppGpp. We present experimental evidence highlighting the importance of base-specific contacts between ppGpp and specific cytosine residues during both transcription initiation and elongation.

  12. Antioxidative Activities of Both Oleic Acid and Camellia tenuifolia Seed Oil Are Regulated by the Transcription Factor DAF-16/FOXO in Caenorhabditis elegans

    Science.gov (United States)

    Wei, Chia-Cheng; Yen, Pei-Ling; Chang, Shang-Tzen; Cheng, Pei-Ling; Lo, Yi-Chen; Liao, Vivian Hsiu-Chuan

    2016-01-01

    Background Tea seed oil is a high quality edible oil, yet lacking sufficient scientific evidences to support the nutritional and medical purposes. We identified major and minor components in Camellia tenuifolia seed oil and investigated the antioxidative activity and its underlying mechanisms in Caenorhabditis elegans. Principal Findings The results showed that the major constitutes in C. tenuifolia seed oil were unsaturated fatty acids (~78.4%). Moreover, two minor compounds, β-amyrin and β-sitosterol, were identified and their antioxidative activity was examined. We found that oleic acid was the major constitute in C. tenuifolia seed oil and plays a key role in the antioxidative activity of C. tenuifolia seed oil in C. elegans. Conclusions This study found evidences that the transcription factor DAF-16/FOXO was involved in both oleic acid- and C. tenuifolia seed oil-mediated oxidative stress resistance in C. elegans. This study suggests the potential of C. tenuifolia seed oil as nutrient or functional foods. PMID:27275864

  13. Role of an expansin-like molecule in Dictyostelium morphogenesis and regulation of its gene expression by the signal transducer and activator of transcription protein Dd-STATa.

    Science.gov (United States)

    Ogasawara, Shun; Shimada, Nao; Kawata, Takefumi

    2009-02-01

    Expansins are proteins involved in plant morphogenesis, exerting their effects on cellulose to extend cell walls. Dictyostelium is an organism that possesses expansin-like molecules, but their functions are not known. In this study, we analyzed the expL7 (expansin-like 7) gene, which has been identified as a putative target of Dd-STATa, a Dictyostelium homolog of the metazoan signal transducer and activator of transcription (STAT) proteins. Promoter fragments of the expL7 were fused to a lacZ reporter and the expression patterns determined. As expected from the behavior of the endogenous expL7 gene, the expL7/lacZ fusion gene was downregulated in Dd-STATa null slugs. In the parental strain, the expL7 promoter was activated in the anterior tip region. Mutational analysis of the promoter identified a sequence that was necessary for expression in tip cells. In addition, an activator sequence for pstAB cells was identified. These sequences act in combination with the repressor region to prevent ectopic expL7 expression in the prespore and prestalk regions of the slug and culminant. Although the expL7 null mutant showed no phenotypic change, the expL7 overexpressor showed aberrant stalk formation. These results indicate that the expansin-like molecule is important for morphogenesis in Dictyostelium.

  14. Regulation of human protein S gene (PROS1) transcription

    NARCIS (Netherlands)

    Wolf, Cornelia de

    2006-01-01

    This thesis describes the investigation of the transcriptional regulation of the gene for anticoagulant plasma Protein S, PROS1. Protein S is a cofactor for Protein C in the Protein C anticoagulant pathway. The coagulation cascade is negatively regulated by this pathway through inactivation of

  15. Dissecting specific and global transcriptional regulation of bacterial gene expression

    NARCIS (Netherlands)

    Gerosa, Luca; Kochanowski, Karl; Heinemann, Matthias; Sauer, Uwe

    2013-01-01

    Gene expression is regulated by specific transcriptional circuits but also by the global expression machinery as a function of growth. Simultaneous specific and global regulation thus constitutes an additional-but often neglected-layer of complexity in gene expression. Here, we develop an experiment

  16. Exposure to static magnetic fields increases insulin secretion in rat INS-1 cells by activating the transcription of the insulin gene and up-regulating the expression of vesicle-secreted proteins.

    Science.gov (United States)

    Mao, Libin; Wang, Huiqin; Ma, Fenghui; Guo, Zhixia; He, Hongpeng; Zhou, Hao; Wang, Nan

    2017-08-01

    To evaluate the effect of static magnetic fields (SMFs) on insulin secretion and explore the mechanisms underlying exposure to SMF-induced insulin secretion in rat insulinoma INS-1 cells. INS-1 cells were exposed to a 400 mT SMF for 72 h, and the proliferation of INS-1 cells was detected by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The secretion of insulin was measured with an enzyme linked immunosorbent assays (ELISA), the expression of genes was detected by real-time PCR, and the expression of proteins was measured by Western blotting. Exposure to an SMF increased the expression and secretion of insulin by INS-1 cells but did not affect cell proliferation. Moreover, SMF exposure up-regulated the expression of several pancreas-specific transcriptional factors. Specifically, the activity of the rat insulin promoter was enhanced in INS-1 cells exposed to an SMF, and the expression levels of synaptosomal-associated protein 25 (SNAP-25) and syntaxin-1A were up-regulated after exposure to an SMF. SMF exposure can promote insulin secretion in rat INS-1 cells by activating the transcription of the insulin gene and up-regulating the expression of vesicle-secreted proteins.

  17. Transcriptional regulation of the stress response by mTOR.

    Science.gov (United States)

    Aramburu, Jose; Ortells, M Carmen; Tejedor, Sonia; Buxadé, Maria; López-Rodríguez, Cristina

    2014-07-01

    The kinase mammalian target of rapamycin (mTOR) is a central regulator of cell growth and proliferation that integrates inputs from growth factor receptors, nutrient availability, intracellular ATP (adenosine 5'-triphosphate), and a variety of stressors. Since early works in the mid-1990s uncovered the role of mTOR in stimulating protein translation, this kinase has emerged as a rather multifaceted regulator of numerous processes. Whereas mTOR is generally activated by growth- and proliferation-stimulating signals, its activity can be reduced and even suppressed when cells are exposed to a variety of stress conditions. However, cells can also adapt to stress while maintaining their growth capacity and mTOR function. Despite knowledge accumulated on how stress represses mTOR, less is known about mTOR influencing stress responses. In this review, we discuss the capability of mTOR, in particular mTOR complex 1 (mTORC1), to activate stress-responsive transcription factors, and we outline open questions for future investigation.

  18. STREAM: Static Thermodynamic REgulAtory Model of transcription.

    Science.gov (United States)

    Bauer, Denis C; Bailey, Timothy L

    2008-11-01

    Understanding the transcriptional regulation of a gene in detail is a crucial step towards uncovering and ultimately utilizing the regulatory grammar of the genome. Modeling transcriptional regulation using thermodynamic equations has become an increasingly important approach towards this goal. Here, we present stream, the first publicly available framework for modeling, visualizing and predicting the regulation of the transcription rate of a target gene. Given the concentrations of a set of transcription factors (TFs), the TF binding sites (TFBSs) in a regulatory DNA region, and the transcription rate of the target gene, stream will optimize its parameters to generate a model that best fits the input data. This trained model can then be used to (a) validate that the given set of TFs is able to regulate the target gene and (b) to predict the transcription rate under different conditions (e.g. different tissues, knockout/additional TFs or mutated/missing TFBSs). The platform independent executable of stream, as well as a tutorial and the full documentation, are available at http://bioinformatics.org.au/stream/. stream requires Java version 5 or higher.

  19. NFAT5 regulates transcription of the mouse telomerase reverse transcriptase gene

    Energy Technology Data Exchange (ETDEWEB)

    Fujiki, Tsukasa; Udono, Miyako; Kotake, Yojiro; Yamashita, Makiko; Shirahata, Sanetaka; Katakura, Yoshinori, E-mail: katakura.yoshinori.528@m.kyushu-u.ac.jp

    2010-12-10

    We aimed to clarify the transcription-regulation mechanisms of the mouse telomerase reverse transcriptase gene (mTERT). First, we searched for the promoter region required for transcriptional activation of mTERT and identified an enhancer cis-element (named mTERT-EE) located between - 200 and - 179 bp of the mouse TERT gene (mTERT). EMSA results suggested that nuclear factor of activated T cells (NFAT) member proteins bind to mTERT-EE. We then identified NFAT5 as the factor binding to mTERT-EE and found that it activates the transcription of the mTERT core promoter. The results that siRNA directed against NFAT5 significantly reduced mTERT expression and mTERT core promoter activity and that the expressions of NFAT5 and mTERT were well correlated in various mouse tissues except liver suggest that NFAT5 dominantly and directly regulates mTERT expression. To clarify their functionality further, we investigated the effect of hypertonic stress, a known stimulus affecting the expression and transcriptional activity of NFAT5, on mTERT expression. The result indicated that hypertonic stress activates mTERT transcription via the activation and recruitment of NFAT5 to the mTERT promoter. These results provide useful information about the transcription-regulation mechanisms of mTERT.

  20. Regulation of Estrogen-Dependent Transcription by the LIM Cofactors CLIM and RLIM in Breast Cancer

    Science.gov (United States)

    Johnsen, Steven A.; Güngör, Cenap; Prenzel, Tanja; Riethdorf, Sabine; Riethdorf, Lutz; Taniguchi-Ishigaki, Naoko; Rau, Thomas; Tursun, Baris; Furlow, J. David; Sauter, Guido; Scheffner, Martin; Pantel, Klaus; Gannon, Frank; Bach, Ingolf

    2009-01-01

    Mammary oncogenesis is profoundly influenced by signaling pathways controlled by Estrogen Receptor-alpha (ERα). Although it is known that ERα exerts its oncogenic effect by stimulating the proliferation of many human breast cancers through the activation of target genes, our knowledge of the underlying transcriptional mechanisms remains limited. Our published work has shown that the in vivo activity of LIM homeodomain transcription factors (LIM-HDs) is critically regulated by Cofactors of LIM-HD proteins (CLIM) and the ubiquitin ligase RING finger LIM domain interacting protein (RLIM). Here, we identify CLIM and RLIM as novel ERα cofactors that co-localize and interact with ERα in primary human breast tumors. We show that both cofactors associate with estrogen responsive promoters and regulate the expression of endogenous ERα target genes in breast cancer cells. Surprisingly, our results indicate opposing functions of LIM cofactors for ERα and LIM-HDs: whereas CLIM enhances transcriptional activity of LIM-HDs, it inhibits transcriptional activation mediated by ERα on most target genes in vivo. In turn, the ubiquitin ligase RLIM inhibits transcriptional activity of LIM-HDs, but enhances transcriptional activation of endogenous ERα target genes. Results from a human breast cancer tissue microarray (TMA) of 1,335 patients revealed a highly significant correlation of elevated CLIM levels to ER/PR positivity and poor differentiation of tumors. Combined, these results indicate that LIM cofactors CLIM and RLIM regulate the biological activity of ERα during the development of human breast cancer. PMID:19117995

  1. Promoter proximal polyadenylation sites reduce transcription activity

    DEFF Research Database (Denmark)

    Andersen, Pia Kjølhede; Lykke-Andersen, Søren; Jensen, Torben Heick

    2012-01-01

    Gene expression relies on the functional communication between mRNA processing and transcription. We previously described the negative impact of a point-mutated splice donor (SD) site on transcription. Here we demonstrate that this mutation activates an upstream cryptic polyadenylation (CpA) site...... RNA polymerase II-transcribed genes use specialized termination mechanisms to maintain high transcription levels.......Gene expression relies on the functional communication between mRNA processing and transcription. We previously described the negative impact of a point-mutated splice donor (SD) site on transcription. Here we demonstrate that this mutation activates an upstream cryptic polyadenylation (CpA) site......, which in turn causes reduced transcription. Functional depletion of U1 snRNP in the context of the wild-type SD triggers the same CpA event accompanied by decreased RNA levels. Thus, in accordance with recent findings, U1 snRNP can shield premature pA sites. The negative impact of unshielded pA sites...

  2. Global analysis of gene transcription regulation in prokaryotes.

    Science.gov (United States)

    Zhou, D; Yang, R

    2006-10-01

    Prokaryotes have complex mechanisms to regulate their gene transcription, through the action of transcription factors (TFs). This review deals with current strategies, approaches and challenges in the understanding of i) how to map the repertoires of TF and operon on a genome, ii) how to identify the specific cis-acting DNA elements and their DNA-binding TFs that are required for expression of a given gene, iii) how to define the regulon members of a given TF, iv) how a given TF interacts with its target promoters, v) how these TF-promoter DNA interactions constitute regulatory networks, and vi) how transcriptional regulatory networks can be reconstructed by the reverse-engineering methods. Our goal is to depict the power of newly developed genomic techniques and computational tools, alone or in combination, to dissect the genetic circuitry of transcription regulation, and how this has the tremendous potential to model the regulatory networks in the prokaryotic cells.

  3. Initiation and regulation of paramyxovirus transcription and replication.

    Science.gov (United States)

    Noton, Sarah L; Fearns, Rachel

    2015-05-01

    The paramyxovirus family has a genome consisting of a single strand of negative sense RNA. This genome acts as a template for two distinct processes: transcription to generate subgenomic, capped and polyadenylated mRNAs, and genome replication. These viruses only encode one polymerase. Thus, an intriguing question is, how does the viral polymerase initiate and become committed to either transcription or replication? By answering this we can begin to understand how these two processes are regulated. In this review article, we present recent findings from studies on the paramyxovirus, respiratory syncytial virus, which show how its polymerase is able to initiate transcription and replication from a single promoter. We discuss how these findings apply to other paramyxoviruses. Then, we examine how trans-acting proteins and promoter secondary structure might serve to regulate transcription and replication during different phases of the paramyxovirus replication cycle.

  4. Germline transcription: a key regulator of accessibility and recombination.

    Science.gov (United States)

    Abarrategui, Iratxe; Krangel, Michael S

    2009-01-01

    The developmental control of V(D)J recombination is imposed at the level of chromatin accessibility of recombination signal sequences (RSSs) to the recombinase machinery. Cis-acting transcriptional regulatory elements such as promoters and enhancers play a central role in the control of accessibility in vivo. However, the molecular mechanisms by which these elements influence accessibility are still under investigation. Although accessibility for V(D)J recombination is usually accompanied by germline transcription at antigen receptor loci, the functional significance of this transcription in directing RSS accessibility has been elusive. In this chapter, we review past studies outlining the complex relationship between V(D)J recombination and transcription as well as our current understanding on how chromatin structure is regulated during gene expression. We then summarize recent work that directly addresses the functional role of transcription in V(D)J recombination.

  5. Retinoic acid regulates the expression of photoreceptor transcription factor NRL.

    Science.gov (United States)

    Khanna, Hemant; Akimoto, Masayuki; Siffroi-Fernandez, Sandrine; Friedman, James S; Hicks, David; Swaroop, Anand

    2006-09-15

    NRL (neural retina leucine zipper) is a key basic motif-leucine zipper (bZIP) transcription factor, which orchestrates rod photoreceptor differentiation by activating the expression of rod-specific genes. The deletion of Nrl in mice results in functional cones that are derived from rod precursors. However, signaling pathways modulating the expression or activity of NRL have not been elucidated. Here, we show that retinoic acid (RA), a diffusible factor implicated in rod development, activates the expression of NRL in serum-deprived Y79 human retinoblastoma cells and in primary cultures of rat and porcine photoreceptors. The effect of RA is mimicked by TTNPB, a RA receptor agonist, and requires new protein synthesis. DNaseI footprinting and electrophoretic mobility shift assays (EMSA) using bovine retinal nuclear extract demonstrate that RA response elements (RAREs) identified within the Nrl promoter bind to RA receptors. Furthermore, in transiently transfected Y79 and HEK293 cells the activity of Nrl-promoter driving a luciferase reporter gene is induced by RA, and this activation is mediated by RAREs. Our data suggest that signaling by RA via RA receptors regulates the expression of NRL, providing a framework for delineating early steps in photoreceptor cell fate determination.

  6. Large-scale transcriptome data reveals transcriptional activity of fission yeast LTR retrotransposons

    DEFF Research Database (Denmark)

    Mourier, Tobias; Willerslev, Eske

    2010-01-01

    transcriptional activity from Long Terminal Repeat (LTR) retrotransposons. LTR retrotransposons are normally flanked by two LTR sequences. However, the majority of LTR sequences in S. pombe exist as solitary LTRs, i.e. as single terminal repeat sequences not flanking a retrotransposon. Transcriptional activity...... of transcriptional activity are observed from both strands of solitary LTR sequences. Transcriptome data collected during meiosis suggests that transcription of solitary LTRs is correlated with the transcription of nearby protein-coding genes. CONCLUSIONS: Presumably, the host organism negatively regulates...... proliferation of LTR retrotransposons. The finding of considerable transcriptional activity of retrotransposons suggests that part of this regulation is likely to take place at a posttranscriptional level. Alternatively, the transcriptional activity may signify a hitherto unrecognized activity level...

  7. Coordinated transcriptional regulation patterns associated with infertility phenotypes in men.

    Science.gov (United States)

    Ellis, Peter J I; Furlong, Robert A; Conner, Sarah J; Kirkman-Brown, Jackson; Afnan, Masoud; Barratt, Christopher; Griffin, Darren K; Affara, Nabeel A

    2007-08-01

    Microarray gene-expression profiling is a powerful tool for global analysis of the transcriptional consequences of disease phenotypes. Understanding the genetic correlates of particular pathological states is important for more accurate diagnosis and screening of patients, and thus for suggesting appropriate avenues of treatment. As yet, there has been little research describing gene-expression profiling of infertile and subfertile men, and thus the underlying transcriptional events involved in loss of spermatogenesis remain unclear. Here we present the results of an initial screen of 33 patients with differing spermatogenic phenotypes. Oligonucleotide array expression profiling was performed on testis biopsies for 33 patients presenting for testicular sperm extraction. Significantly regulated genes were selected using a mixed model analysis of variance. Principle components analysis and hierarchical clustering were used to interpret the resulting dataset with reference to the patient history, clinical findings and histological composition of the biopsies. Striking patterns of coordinated gene expression were found. The most significant contains multiple germ cell-specific genes and corresponds to the degree of successful spermatogenesis in each patient, whereas a second pattern corresponds to inflammatory activity within the testis. Smaller-scale patterns were also observed, relating to unique features of the individual biopsies.

  8. Transcription regulation mechanisms of bacteriophages: Recent advances and future prospects

    OpenAIRE

    Yang, Haiquan; Ma, Yingfang; Wang, Yitian; Yang, Haixia; Shen, Wei; Chen, Xianzhong

    2014-01-01

    Phage diversity significantly contributes to ecology and evolution of new bacterial species through horizontal gene transfer. Therefore, it is essential to understand the mechanisms underlying phage-host interactions. After initial infection, the phage utilizes the transcriptional machinery of the host to direct the expression of its own genes. This review presents a view on the transcriptional regulation mechanisms of bacteriophages, and its contribution to phage diversity and classification...

  9. Cellular adaptation to hypoxia and p53 transcription regulation

    Institute of Scientific and Technical Information of China (English)

    Yang ZHAO; Xue-qun CHEN; Ji-zeng DU

    2009-01-01

    Tumor suppressor p53 is the most frequently mutated gene in human tumors. Meanwhile, under stress conditions, p53 also acts as a transcription factor, regulating the expression of a series of target genes to maintain the integrity of genome. The target genes of p53 can be classified into genes regulating cell cycle arrest, genes involved in apoptosis, and genes inhibiting angiogenesis. p53 protein contains a transactivation domain, a sequence-specific DNA binding domain, a tetramerization domain, a non-specific DNA binding domain that recognizes damaged DNA, and a later identified proline-rich domain. Under stress, p53 proteins accumulate and are activated through two mechanisms. One, involving ataxia telangiectasia-mutated protein (ATM), is that the interaction between p53 and its down-regulation factor murine double minute 2 (MDM2) decreases, leading to p53 phosphorylation on Ser15, as determined by the post-translational mechanism; the other holds that p53 increases and is activated through the binding of ribosomal protein L26 (RPL26) or nucleolin to p53 mRNA 5' untranslated region (UTR), regulating p53 translation. Under hypoxia, p53 decreases transactivation and increases transrepression. The mutations outside the DNA binding domain of p53 also contribute to tumor progress, so further studies on p53 should also be focused on this direction. The subterranean blind mole rat Spalax in Israel is a good model for hypoxia-adaptation. The p53 of Spalax mutated in residue 172 and residue 207 from arginine to lysine, conferring it the ability to survive hypoxic conditions. This model indicates that p53 acts as a master gene of diversity formation during evolution.

  10. Understanding Transcription Factor Regulation by Integrating Gene Expression and DNase I Hypersensitive Sites

    Directory of Open Access Journals (Sweden)

    Guohua Wang

    2015-01-01

    Full Text Available Transcription factors are proteins that bind to DNA sequences to regulate gene transcription. The transcription factor binding sites are short DNA sequences (5–20 bp long specifically bound by one or more transcription factors. The identification of transcription factor binding sites and prediction of their function continue to be challenging problems in computational biology. In this study, by integrating the DNase I hypersensitive sites with known position weight matrices in the TRANSFAC database, the transcription factor binding sites in gene regulatory region are identified. Based on the global gene expression patterns in cervical cancer HeLaS3 cell and HelaS3-ifnα4h cell (interferon treatment on HeLaS3 cell for 4 hours, we present a model-based computational approach to predict a set of transcription factors that potentially cause such differential gene expression. Significantly, 6 out 10 predicted functional factors, including IRF, IRF-2, IRF-9, IRF-1 and IRF-3, ICSBP, belong to interferon regulatory factor family and upregulate the gene expression levels responding to the interferon treatment. Another factor, ISGF-3, is also a transcriptional activator induced by interferon alpha. Using the different transcription factor binding sites selected criteria, the prediction result of our model is consistent. Our model demonstrated the potential to computationally identify the functional transcription factors in gene regulation.

  11. Engineering prokaryotic transcriptional activators as metabolite biosensors in yeast

    DEFF Research Database (Denmark)

    Skjødt, Mette Louise; Snoek, Tim; Kildegaard, Kanchana Rueksomtawin

    2016-01-01

    real-time monitoring of production has attracted attention. Here we applied systematic engineering of multiple parameters to search for a general biosensor design in the budding yeast Saccharomyces cerevisiae based on small-molecule binding transcriptional activators from the prokaryote superfamily......,cis-muconic acid at different levels, and found that reporter gene output correlated with production. The transplantation of prokaryotic transcriptional activators into the eukaryotic chassis illustrates the potential of a hitherto untapped biosensor resource useful for biotechnological applications....... of LysR-type transcriptional regulators (LTTRs). We identified a design supporting LTTR-dependent activation of reporter gene expression in the presence of cognate small-molecule inducers. As proof of principle, we applied the biosensors for in vivo screening of cells producing naringenin or cis...

  12. Regulating expression of cell and tissue-specific genes by modifying transcription

    Energy Technology Data Exchange (ETDEWEB)

    Beachy, Roger N; Dai, Shunhong

    2010-06-14

    Transcriptional regulation is the primary step to control gene expression, therefore function. Such regulation is achieved primarily via a combination of the activities of the promoter cis regulatory DNA elements and trans regulatory proteins that function through binding to these DNA elements. Rice bZIP transcription factors RF2a, RF2b and RLP1 play key roles in regulating the activity of a vascular tissue specific promoter isolated from Rice Tungro Bacilliform Virus (RTBV), through their interactions with the Box II essential cis element located in the promoter (Dai et al., 2006., Dai et al., 2004., Yin et al., 1997). RF2a, RF2b and RLP1 possess multiple regulatory domains. Functional characterization reveals that those domains can activate or repress the activity of the RTBV promoter. It is equally as important to recognize that these proteins control plant development by regulating differentiation and/or function of the vascular tissues. Studies of transcriptional regulation of the RTBV promoter by this group of bZIP proteins will not only provide insights about gene expression in the vascular tissue, but also insights about general mechanisms of transcription activation and repression. The knowledge gained from this research will also enable us to develop a well-described set of tools that can be used to control expression of multiple genes in transgenic plants. We have proposed characterize the function domains of RF2a, RF2b and RLP1 and explore the biological function of the transcription repressor RLP1.

  13. Regulating expression of cell and tissue-specific genes by modifying transcription

    Energy Technology Data Exchange (ETDEWEB)

    Beachy, Roger N; Dai, Shunhong

    2010-06-14

    Transcriptional regulation is the primary step to control gene expression, therefore function. Such regulation is achieved primarily via a combination of the activities of the promoter cis regulatory DNA elements and trans regulatory proteins that function through binding to these DNA elements. Rice bZIP transcription factors RF2a, RF2b and RLP1 play key roles in regulating the activity of a vascular tissue specific promoter isolated from Rice Tungro Bacilliform Virus (RTBV), through their interactions with the Box II essential cis element located in the promoter (Dai et al., 2006., Dai et al., 2004., Yin et al., 1997). RF2a, RF2b and RLP1 possess multiple regulatory domains. Functional characterization reveals that those domains can activate or repress the activity of the RTBV promoter. It is equally as important to recognize that these proteins control plant development by regulating differentiation and/or function of the vascular tissues. Studies of transcriptional regulation of the RTBV promoter by this group of bZIP proteins will not only provide insights about gene expression in the vascular tissue, but also insights about general mechanisms of transcription activation and repression. The knowledge gained from this research will also enable us to develop a well-described set of tools that can be used to control expression of multiple genes in transgenic plants. We have proposed characterize the function domains of RF2a, RF2b and RLP1 and explore the biological function of the transcription repressor RLP1.

  14. Transcriptional regulation of dendritic cell diversity.

    Science.gov (United States)

    Chopin, Michaël; Allan, Rhys S; Belz, Gabrielle T

    2012-01-01

    Dendritic cells (DCs) are specialized antigen presenting cells that are exquisitely adapted to sense pathogens and induce the development of adaptive immune responses. They form a complex network of phenotypically and functionally distinct subsets. Within this network, individual DC subsets display highly specific roles in local immunosurveillance, migration, and antigen presentation. This division of labor amongst DCs offers great potential to tune the immune response by harnessing subset-specific attributes of DCs in the clinical setting. Until recently, our understanding of DC subsets has been limited and paralleled by poor clinical translation and efficacy. We have now begun to unravel how different DC subsets develop within a complex multilayered system. These findings open up exciting possibilities for targeted manipulation of DC subsets. Furthermore, ground-breaking developments overcoming a major translational obstacle - identification of similar DC populations in mouse and man - now sets the stage for significant advances in the field. Here we explore the determinants that underpin cellular and transcriptional heterogeneity within the DC network, how these influence DC distribution and localization at steady-state, and the capacity of DCs to present antigens via direct or cross-presentation during pathogen infection.

  15. Navigating the transcriptional roadmap regulating plant secondary cell wall deposition

    Directory of Open Access Journals (Sweden)

    Steven Grant Hussey

    2013-08-01

    Full Text Available The current status of lignocellulosic biomass as an invaluable resource in industry, agriculture and health has spurred increased interest in understanding the transcriptional regulation of secondary cell wall (SCW biosynthesis. The last decade of research has revealed an extensive network of NAC, MYB and other families of transcription factors regulating Arabidopsis SCW biosynthesis, and numerous studies have explored SCW-related transcription factors in other dicots and monocots. Whilst the general structure of the Arabidopsis network has been a topic of several reviews, they have not comprehensively represented the detailed protein-DNA and protein-protein interactions described in the literature, and an understanding of network dynamics and functionality has not yet been achieved for SCW formation. Furthermore the methodologies employed in studies of SCW transcriptional regulation have not received much attention, especially in the case of non-model organisms. In this review, we have reconstructed the most exhaustive literature-based network representations to date of SCW transcriptional regulation in Arabidopsis. We include a manipulable Cytoscape representation of the Arabidopsis SCW transcriptional network to aid in future studies, along with a list of supporting literature for each documented interaction. Amongst other topics, we discuss the various components of the network, its evolutionary conservation in plants, putative modules and dynamic mechanisms that may influence network function, and the approaches that have been employed in network inference. Future research should aim to better understand network function and its response to dynamic perturbations, whilst the development and application of genome-wide approaches such as ChIP-seq and systems genetics are in progress for the study of SCW transcriptional regulation in non-model organisms.

  16. Ranges of control in the transcriptional regulation of Escherichia coli

    Directory of Open Access Journals (Sweden)

    Stoyan Helga

    2009-12-01

    Full Text Available Abstract Background The positioning of genes in the genome is an important evolutionary degree of freedom for organizing gene regulation. Statistical properties of these distributions have been studied particularly in relation to the transcriptional regulatory network. The systematics of gene-gene distances then become important sources of information on the control, which different biological mechanisms exert on gene expression. Results Here we study a set of categories, which has to our knowledge not been analyzed before. We distinguish between genes that do not participate in the transcriptional regulatory network (i.e. that are according to current knowledge not producing transcription factors and do not possess binding sites for transcription factors in their regulatory region, and genes that via transcription factors either are regulated by or regulate other genes. We find that the two types of genes ("isolated" and "regulatory" genes show a clear statistical repulsion and have different ranges of correlations. In particular we find that isolated genes have a preference for shorter intergenic distances. Conclusions These findings support previous evidence from gene expression patterns for two distinct logical types of control, namely digital control (i.e. network-based control mediated by dedicated transcription factors and analog control (i.e. control based on genome structure and mediated by neighborhood on the genome.

  17. Both mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (ERK) 1/2 and phosphatidylinositide-3-OH kinase (PI3K)/Akt pathways regulate activation of E-twenty-six (ETS)-like transcription factor 1 (Elk-1) in U138 glioblastoma cells.

    Science.gov (United States)

    Mut, Melike; Lule, Sevda; Demir, Ozlem; Kurnaz, Isil Aksan; Vural, Imran

    2012-02-01

    Epidermal growth factor (EGF) and its receptor (EGFR) have been shown to play a significant role in the pathogenesis of glioblastoma. In our study, the EGFR was stimulated with EGF in human U138 glioblastoma cells. We show that the activated mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (ERK) 1/2 pathway phosphorylated the E twenty-six (ETS)-like transcription factor 1 (Elk-1) mainly at serine 383 residue. Mitogen-activated protein kinase kinase (MEK) 1/2 inhibitor, UO126 and ERK inhibitor II, FR180204 blocked the Elk-1 phosphorylation and activation. The phosphatidylinositide-3-OH kinase (PI3K)/Akt pathway was also involved in the Elk-1 activation. Activation of the Elk-1 led to an increased survival and a proliferative response with the EGF stimulation in the U138 glioblastoma cells. Knocking-down the Elk-1 using an RNA interference technique caused a decrease in survival of the unstimulated U138 glioblastoma cells and also decreased the proliferative response to the EGF stimulation. The Elk-1 transcription factor was important for the survival and proliferation of U138 glioblastoma cells upon the stimulation of EGFR with EGF. The MAPK/ERK1/2 and PI3K/Akt pathways regulated this response via activation of the Elk-1 transcription factor. The Elk-1 may be one of the convergence points for pathways located downstream of EGFR in glioblastoma cells. Utilization of the Elk-1 as a therapeutic target may lead to a novel strategy in treatment of glioblastoma.

  18. Expression and activity of the Calvin-Benson-Bassham cycle transcriptional regulator CbbR from Acidithiobacillus ferrooxidans in Ralstonia eutropha.

    Science.gov (United States)

    Esparza, Mario; Jedlicki, Eugenia; Dopson, Mark; Holmes, David S

    2015-08-01

    Autotrophic fixation of carbon dioxide into cellular carbon occurs via several pathways but quantitatively, the Calvin-Benson-Bassham cycle is the most important. CbbR regulates the expression of the cbb genes involved in CO2 fixation via the Calvin-Benson-Bassham cycle in a number of autotrophic bacteria. A gene potentially encoding CbbR (cbbR(AF)) has been predicted in the genome of the chemolithoautotrophic, extreme acidophile Acidithiobacillus ferrooxidans. However, this microorganism is recalcitrant to genetic manipulation impeding the experimental validation of bioinformatic predictions. Two novel functional assays were devised to advance our understanding of cbbR(AF) function using the mutated facultative autotroph Ralstonia eutropha H14 ΔcbbR as a surrogate host to test gene function: (i) cbbR(AF) was expressed in R. eutropha and was able to complement ΔcbbR; and (ii) CbbR(AF) was able to regulate the in vivo activity of four A. ferrooxidans cbb operon promoters in R. eutropha. These results open up the use of R. eutropha as a surrogate host to explore cbbR(AF) activity.

  19. Post-transcriptional regulation of gene expression in Yersinia species

    Directory of Open Access Journals (Sweden)

    Chelsea A Schiano

    2012-11-01

    Full Text Available Proper regulation of gene expression is required by bacterial pathogens to respond to continually changing environmental conditions and the host response during the infectious process. While transcriptional regulation is perhaps the most well understood form of controlling gene expression, recent studies have demonstrated the importance of post-transcriptional mechanisms of gene regulation that allow for more refined management of the bacterial response to host conditions. Yersinia species of bacteria are known to use various forms of post-transcriptional regulation for control of many virulence-associated genes. These include regulation by cis- and trans-acting small non-coding RNAs, RNA-binding proteins, RNases, and thermoswitches. The effects of these and other regulatory mechanisms on Yersinia physiology can be profound and have been shown to influence type III secretion, motility, biofilm formation, host cell invasion, intracellular survival and replication, and more. In this review, we will discuss these and other post-transcriptional mechanisms and their influence on virulence gene regulation, with a particular emphasis on how these processes influence the virulence of Yersinia in the host.

  20. The sequence-specific transcription factor c-Jun targets Cockayne syndrome protein B to regulate transcription and chromatin structure.

    Directory of Open Access Journals (Sweden)

    Robert J Lake

    2014-04-01

    Full Text Available Cockayne syndrome is an inherited premature aging disease associated with numerous developmental and neurological defects, and mutations in the gene encoding the CSB protein account for the majority of Cockayne syndrome cases. Accumulating evidence suggests that CSB functions in transcription regulation, in addition to its roles in DNA repair, and those defects in this transcriptional activity might contribute to the clinical features of Cockayne syndrome. Transcription profiling studies have so far uncovered CSB-dependent effects on gene expression; however, the direct targets of CSB's transcriptional activity remain largely unknown. In this paper, we report the first comprehensive analysis of CSB genomic occupancy during replicative cell growth. We found that CSB occupancy sites display a high correlation to regions with epigenetic features of promoters and enhancers. Furthermore, we found that CSB occupancy is enriched at sites containing the TPA-response element. Consistent with this binding site preference, we show that CSB and the transcription factor c-Jun can be found in the same protein-DNA complex, suggesting that c-Jun can target CSB to specific genomic regions. In support of this notion, we observed decreased CSB occupancy of TPA-response elements when c-Jun levels were diminished. By modulating CSB abundance, we found that CSB can influence the expression of nearby genes and impact nucleosome positioning in the vicinity of its binding site. These results indicate that CSB can be targeted to specific genomic loci by sequence-specific transcription factors to regulate transcription and local chromatin structure. Additionally, comparison of CSB occupancy sites with the MSigDB Pathways database suggests that CSB might function in peroxisome proliferation, EGF receptor transactivation, G protein signaling and NF-κB activation, shedding new light on the possible causes and mechanisms of Cockayne syndrome.

  1. Transcription factor p53 can regulate proliferation, apoptosis and secretory activity of luteinizing porcine ovarian granulosa cell cultured with and without ghrelin and FSH.

    Science.gov (United States)

    Sirotkin, A V; Benco, A; Tandlmajerova, A; Vasícek, D; Kotwica, J; Darlak, K; Valenzuela, F

    2008-11-01

    The aim of our in vitro experiments was to examine the role of transcription factor p53 in controlling the basic functions of ovarian cells and their response to hormonal treatments. Porcine ovarian granulosa cells, transfected and non-transfected with a gene construct encoding p53, were cultured with ghrelin and FSH (all at concentrations of 0, 1, 10, or 100 ng/ml). Accumulation of p53, of apoptosis-related (MAP3K5) and proliferation-related (cyclin B1) substances was evaluated by immunocytochemistry. The secretion of progesterone (P(4)), oxytocin (OT), prostaglandin F (PGF), and E (PGE) was measured by RIA. Transfection with the p53 gene construct promoted accumulation of this transcription factor within cells. It also stimulated the expression of a marker of apoptosis (MAP3K5). Over-expression of p53 resulted in reduced accumulation of a marker of proliferation (cyclin B1), P(4), and PGF secretion and increased OT and PGE secretion. Ghrelin, when added alone, did not affect p53 or P(4), but reduced MAP3K5 and increased PGF and PGE secretion. Over-expression of p53 reversed the effect of ghrelin on OT, caused it to be inhibitory to P(4) secretion, but did not modify its action on MAP3K5, PGF, or PGE. FSH promoted the accumulation of p53, MAP3K5, and cyclin B1; these effects were unaffected by p53 transfection. These multiple effects of the p53 gene construct on luteinizing granulosa cells, cultured with and without hormones 1) demonstrate the effects of ghrelin and FSH on porcine ovarian cell apoptosis and secretory activity, 2) confirm the involvement of p53 in promoting apoptosis and inhibiting P(4) secretion in these cells, 3) provide the first evidence that p53 suppress proliferation of ovarian cells, 4) provide the first evidence that p53 is involved in the control of ovarian peptide hormone (OT) and prostaglandin (PGF and PGE) secretion, and 5) suggest that p53 can modulate, but probably not mediate, the effects of ghrelin and FSH on the ovary.

  2. Activation of rapid signaling pathways and the subsequent transcriptional regulation for the proliferation of breast cancer MCF-7 cells by the treatment with an extract of Glycyrrhiza glabra root.

    Science.gov (United States)

    Dong, Sijun; Inoue, Akio; Zhu, Yun; Tanji, Masao; Kiyama, Ryoiti

    2007-12-01

    Glycyrrhiza glabra root is one of the common traditional Chinese medicines and used as flavoring and sweetening agents for tobaccos, chewing gums, candies, toothpaste and beverages. While glycyrrhizin is one of the main components in the extract of G. glabra root and has been characterized, the other components have not been well characterized. The mechanism of growth activation of breast cancer MCF-7 cells, including the activation of Erk1/2 and Akt, and the transcriptional regulation of estrogen-responsive genes, was examined by means of sulforhodamine B, luciferase reporter gene, real-time RT-PCR and Western blotting assays after the induction of the cells with the extract of G. glabra root. The extract has similar activity to that induced by 17beta-estradiol (E(2)), although glycyrrhizin did not show such an activity. Moreover, the estrogen receptor alpha-dependent neurite outgrowth induced by the extract was similar to that by E(2), whereas glycyrrhizin had no effect. Furthermore, the expression profile examined by cDNA microarray assay using a set of 120 estrogen-responsive genes, which were related to proliferation, transcription, transport, enzymes and signaling, showed a statistically significant correlation (R=0.47, Pglabra root. Furthermore, the extract had estrogenic activity and a distinguishable profile of gene expression, suggesting the presence of potentially useful components other than glycyrrhizin in G. glabra root for hormone and anti-cancer therapies.

  3. Transcriptional and epigenetic regulation of autophagy in aging.

    Science.gov (United States)

    Lapierre, Louis R; Kumsta, Caroline; Sandri, Marco; Ballabio, Andrea; Hansen, Malene

    2015-01-01

    Macroautophagy is a major intracellular degradation process recognized as playing a central role in cell survival and longevity. This multistep process is extensively regulated at several levels, including post-translationally through the action of conserved longevity factors such as the nutrient sensor TOR. More recently, transcriptional regulation of autophagy genes has emerged as an important mechanism for ensuring the somatic maintenance and homeostasis necessary for a long life span. Autophagy is increased in many long-lived model organisms and contributes significantly to their longevity. In turn, conserved transcription factors, particularly the helix-loop-helix transcription factor TFEB and the forkhead transcription factor FOXO, control the expression of many autophagy-related genes and are important for life-span extension. In this review, we discuss recent progress in understanding the contribution of these transcription factors to macroautophagy regulation in the context of aging. We also review current research on epigenetic changes, such as histone modification by the deacetylase SIRT1, that influence autophagy-related gene expression and additionally affect aging. Understanding the molecular regulation of macroautophagy in relation to aging may offer new avenues for the treatment of age-related diseases.

  4. DNA Topoisomerases Maintain Promoters in a State Competent for Transcriptional Activation in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Pedersen, Jakob Madsen; Fredsøe, Jacob Christian; Rødgaard, Morten Terpager;

    2012-01-01

    To investigate the role of DNA topoisomerases in transcription, we have studied global gene expression in Saccharomyces cerevisiae cells deficient for topoisomerases I and II and performed single-gene analyses to support our findings. The genome-wide studies show a general transcriptional down-re...... transcriptional activation of genes with a repressible/inducible mode of regulation....

  5. HER4 Cyt1 and Cyt2 Isoforms Regulate Transcription Through Differential Interactions with a Transcriptional Regulator, Yap

    Science.gov (United States)

    2011-10-01

    Smad7[29]. However, the primary target of Yap is thought to be the family of TEF/ TEAD transcription factors, shown by Zhao et al. to be required for...phosphorylation of Yap by HER4 isoforms modulate the ability of Yap to regulate TEF/ TEAD -, RunX2-, and p73-dependent transcription. We will also examine...whether HER4 s80–Cyt1 and –Cyt2 interact with the Yap:transcription factor complex, 11    specifically Yap:TEF/ TEAD , and will evaluate the ability

  6. The tumor suppressor gene hypermethylated in cancer 1 is transcriptionally regulated by E2F1

    DEFF Research Database (Denmark)

    Jenal, Mathias; Trinh, Emmanuelle; Britschgi, Christian;

    2009-01-01

    The Hypermethylated in Cancer 1 (HIC1) gene encodes a zinc finger transcriptional repressor that cooperates with p53 to suppress cancer development. We and others recently showed that HIC1 is a transcriptional target of p53. To identify additional transcriptional regulators of HIC1, we screened...... to the HIC1 promoter was shown by chromatin immunoprecipitation assays in human TIG3 fibroblasts expressing tamoxifen-activated E2F1. In agreement, activation of E2F1 in TIG3-E2F1 cells markedly increased HIC1 expression. Interestingly, expression of E2F1 in the p53(-/-) hepatocellular carcinoma cell line...

  7. Dual role of G-runs and hnRNP F in the regulation of a mutation-activated pseudoexon in the fibrinogen gamma-chain transcript.

    Directory of Open Access Journals (Sweden)

    Valeria Rimoldi

    Full Text Available Most pathological pseudoexon inclusion events originate from single activating mutations, suggesting that many intronic sequences are on the verge of becoming exons. However, the precise mechanisms controlling pseudoexon definition are still largely unexplored. Here, we investigated the cis-acting elements and trans-acting regulatory factors contributing to the regulation of a previously described fibrinogen gamma-chain (FGG pseudoexon, which is activated by a deep-intronic mutation (IVS6-320A>T. This pseudoexon contains several G-run elements, which may be bound by heterogeneous nuclear ribonucleoproteins (hnRNPs F and H. To explore the effect of these proteins on FGG pseudoexon inclusion, both silencing and overexpression experiments were performed in eukaryotic cells. While hnRNP H did not significantly affect pseudoexon splicing, hnRNP F promoted pseudoexon inclusion, indicating that these two proteins have only partially redundant functions. To verify the binding of hnRNP F and the possible involvement of other trans-acting splicing modulators, pulldown experiments were performed on the region of the pseudoexon characterized by both a G-run and enrichment for exonic splicing enhancers. This 25-bp-long region strongly binds hnRNP F/H and weakly interacts with Serine/Arginine-rich protein 40, which however was demonstrated to be dispensable for FGG pseudoexon inclusion in overexpression experiments. Deletion analysis, besides confirming the splicing-promoting role of the G-run within this 25-bp region, demonstrated that two additional hnRNP F binding sites might instead function as silencer elements. Taken together, our results indicate a major role of hnRNP F in regulating FGG pseudoexon inclusion, and strengthen the notion that G-runs may function either as splicing enhancers or silencers of the same exon.

  8. Dual role of G-runs and hnRNP F in the regulation of a mutation-activated pseudoexon in the fibrinogen gamma-chain transcript.

    Science.gov (United States)

    Rimoldi, Valeria; Soldà, Giulia; Asselta, Rosanna; Spena, Silvia; Stuani, Cristiana; Buratti, Emanuele; Duga, Stefano

    2013-01-01

    Most pathological pseudoexon inclusion events originate from single activating mutations, suggesting that many intronic sequences are on the verge of becoming exons. However, the precise mechanisms controlling pseudoexon definition are still largely unexplored. Here, we investigated the cis-acting elements and trans-acting regulatory factors contributing to the regulation of a previously described fibrinogen gamma-chain (FGG) pseudoexon, which is activated by a deep-intronic mutation (IVS6-320A>T). This pseudoexon contains several G-run elements, which may be bound by heterogeneous nuclear ribonucleoproteins (hnRNPs) F and H. To explore the effect of these proteins on FGG pseudoexon inclusion, both silencing and overexpression experiments were performed in eukaryotic cells. While hnRNP H did not significantly affect pseudoexon splicing, hnRNP F promoted pseudoexon inclusion, indicating that these two proteins have only partially redundant functions. To verify the binding of hnRNP F and the possible involvement of other trans-acting splicing modulators, pulldown experiments were performed on the region of the pseudoexon characterized by both a G-run and enrichment for exonic splicing enhancers. This 25-bp-long region strongly binds hnRNP F/H and weakly interacts with Serine/Arginine-rich protein 40, which however was demonstrated to be dispensable for FGG pseudoexon inclusion in overexpression experiments. Deletion analysis, besides confirming the splicing-promoting role of the G-run within this 25-bp region, demonstrated that two additional hnRNP F binding sites might instead function as silencer elements. Taken together, our results indicate a major role of hnRNP F in regulating FGG pseudoexon inclusion, and strengthen the notion that G-runs may function either as splicing enhancers or silencers of the same exon.

  9. Regulation of heterochromatin transcription by Snail1/LOXL2 during epithelial-to-mesenchymal transition.

    Science.gov (United States)

    Millanes-Romero, Alba; Herranz, Nicolás; Perrera, Valentina; Iturbide, Ane; Loubat-Casanovas, Jordina; Gil, Jesús; Jenuwein, Thomas; García de Herreros, Antonio; Peiró, Sandra

    2013-12-12

    Although heterochromatin is enriched with repressive traits, it is also actively transcribed, giving rise to large amounts of noncoding RNAs. Although these RNAs are responsible for the formation and maintenance of heterochromatin, little is known about how their transcription is regulated. Here, we show that the Snail1 transcription factor represses mouse pericentromeric transcription, acting through the H3K4 deaminase LOXL2. Since Snail1 plays a key role in the epithelial-to-mesenchymal transition (EMT), we analyzed the regulation of heterochromatin transcription in this process. At the onset of EMT, one of the major structural heterochromatin proteins, HP1α, is transiently released from heterochromatin foci in a Snail1/LOXL2-dependent manner, concomitantly with a downregulation of major satellite transcription. Moreover, preventing the downregulation of major satellite transcripts compromised the migratory and invasive behavior of mesenchymal cells. We propose that Snail1 regulates heterochromatin transcription through LOXL2, thus creating the favorable transcriptional state necessary for completing EMT.

  10. Transcriptional Regulation of the p16 Tumor Suppressor Gene.

    Science.gov (United States)

    Kotake, Yojiro; Naemura, Madoka; Murasaki, Chihiro; Inoue, Yasutoshi; Okamoto, Haruna

    2015-08-01

    The p16 tumor suppressor gene encodes a specific inhibitor of cyclin-dependent kinase (CDK) 4 and 6 and is found altered in a wide range of human cancers. p16 plays a pivotal role in tumor suppressor networks through inducing cellular senescence that acts as a barrier to cellular transformation by oncogenic signals. p16 protein is relatively stable and its expression is primary regulated by transcriptional control. Polycomb group (PcG) proteins associate with the p16 locus in a long non-coding RNA, ANRIL-dependent manner, leading to repression of p16 transcription. YB1, a transcription factor, also represses the p16 transcription through direct association with its promoter region. Conversely, the transcription factors Ets1/2 and histone H3K4 methyltransferase MLL1 directly bind to the p16 locus and mediate p16 induction during replicative and premature senescence. In the present review, we discuss the molecular mechanisms by which these factors regulate p16 transcription.

  11. Integration of the transcriptional networks regulating limb morphogenesis.

    Science.gov (United States)

    Rabinowitz, Adam H; Vokes, Steven A

    2012-08-15

    The developing limb is one of the best described vertebrate systems for understanding how coordinated gene expression during embryogenesis leads to the structures present in the mature organism. This knowledge, derived from decades of research, is largely based upon gain- and loss-of-function experiments. These studies have provided limited information about how the key signaling pathways interact with each other and the downstream effectors of these pathways. We summarize our current understanding of known genetic interactions in the context of three temporally defined gene regulatory networks. These networks crystallize our current knowledge, depicting a dynamic process involving multiple feedback loops between the ectoderm and mesoderm. At the same time, they highlight the fact that many essential processes are still largely undescribed. Much of the dynamic transcriptional activity occurring during development is regulated by distal cis-regulatory elements. Modern genomic tools have provided new approaches for studying the function of cis-regulatory elements and we discuss the results of these studies in regard to understanding limb development. Ultimately, these genomic techniques will allow scientists to understand how multiple signaling pathways are integrated in space and time to drive gene expression and regulate the formation of the limb.

  12. Post-transcriptional regulation of ribosomal protein genes during serum starvation in Entamoeba histolytica.

    Science.gov (United States)

    Ahamad, Jamaluddin; Ojha, Sandeep; Srivastava, Ankita; Bhattacharya, Alok; Bhattacharya, Sudha

    2015-06-01

    Ribosome synthesis involves all three RNA polymerases which are co-ordinately regulated to produce equimolar amounts of rRNAs and ribosomal proteins (RPs). Unlike model organisms where transcription of rRNA and RP genes slows down during stress, in E. histolytica rDNA transcription continues but pre-rRNA processing slows down and unprocessed pre-rRNA accumulates during serum starvation. To investigate the regulation of RP genes under stress we measured transcription of six selected RP genes from the small- and large-ribosomal subunits (RPS6, RPS3, RPS19, RPL5, RPL26, RPL30) representing the early-, mid-, and late-stages of ribosomal assembly. Transcripts of these genes persisted in growth-stressed cells. Expression of luciferase reporter under the control of two RP genes (RPS19 and RPL30) was studied during serum starvation and upon serum replenishment. Although luciferase transcript levels remained unchanged during starvation, luciferase activity steadily declined to 7.8% and 15% of control cells, respectively. After serum replenishment the activity increased to normal levels, suggesting post-transcriptional regulation of these genes. Mutations in the sequence -2 to -9 upstream of AUG in the RPL30 gene resulted in the phenotype expected of post-transcriptional regulation. Transcription of luciferase reporter was unaffected in this mutant, and luciferase activity did not decline during serum starvation, showing that this sequence is required to repress translation of RPL30 mRNA, and mutations in this region relieve repression. Our data show that during serum starvation E. histolytica blocks ribosome biogenesis post-transcriptionally by inhibiting pre-rRNA processing on the one hand, and the translation of RP mRNAs on the other.

  13. DDX3, a DEAD box RNA helicase with tumor growth-suppressive property and transcriptional regulation activity of the p21waf1/cip1 promoter, is a candidate tumor suppressor.

    Science.gov (United States)

    Chao, Chi-Hong; Chen, Chun-Ming; Cheng, Pei-Lin; Shih, Jing-Wen; Tsou, Ann-Ping; Lee, Yan-Hwa Wu

    2006-07-01

    DDX3 is a DEAD box RNA helicase with diverse biological functions. Using colony formation assay, our results revealed that DDX3 inhibited the colony formation ability of various tumor cells, and this inhibition might be due to a reduced growth rate caused by DDX3. Additionally, we identified p21(waf1/cip1), a cyclin-dependent kinase inhibitor, as a target gene of DDX3, and the up-regulation of p21(waf1/cip1) expression accounted for the colony-suppressing activity of DDX3. Moreover, DDX3 exerted its transactivation function on p21(waf1/cip1) promoter through an ATPase-dependent but helicase-independent mechanism, and the four Sp1 sites located within the -123 to -63 region, relative to the transcription start site of p21(waf1/cip1) promoter, were essential for the response to DDX3. Furthermore, DDX3 interacted and cooperated with Sp1 to up-regulate the promoter activity of p21(waf1/cip1). To determine the relevance of DDX3 in clinical cancers, the expression profile of DDX3 in various tumors was also examined. A declined expression of DDX3 mRNA and protein was found in approximately 58% to 73% of hepatoma specimens, which led to the reduction of p21(waf1/cip1) expression in a manner independent of p53 status. Additionally, an alteration of subcellular localization from nuclei to cytoplasm was also observed in >70% of cutaneous squamous cell carcinoma samples. Because DDX3 exhibits tumor suppressor functions, such as a growth-suppressive property and transcriptional activation of the p21(waf1/cip1) promoter, and is inactivated through down-regulation of gene expression or alteration of subcellular localization in tumor cells, all these features together suggest that DDX3 might be a candidate tumor suppressor.

  14. Transcriptional regulators of Na, K-ATPase subunits

    Directory of Open Access Journals (Sweden)

    Zhiqin eLi

    2015-10-01

    Full Text Available The Na,K-ATPase classically serves as an ion pump creating an electrochemical gradient across the plasma membrane that is essential for transepithelial transport, nutrient uptake and membrane potential. In addition, Na,K-ATPase also functions as a receptor, a signal transducer and a cell adhesion molecule. With such diverse roles, it is understandable that the Na,K-ATPase subunits, the catalytic alpha-subunit, the beta-subunit and the FXYD proteins, are controlled extensively during development and to accommodate physiological needs. The spatial and temporal expression of Na,K-ATPase is partially regulated at the transcriptional level. Numerous transcription factors, hormones, growth factors, lipids and extracellular stimuli modulate the transcription of the Na,K-ATPase subunits. Moreover, epigenetic mechanisms also contribute to the regulation of Na,K-ATPase expression. With the ever growing knowledge about diseases associated with the malfunction of Na,K-ATPase, this review aims at summarizing the best-characterized transcription regulators that modulate Na,K-ATPase subunit levels. As abnormal expression of Na,K-ATPase subunits have been observed in many carcinoma, we will also discuss transcription factors that are associated with epithelial-to-mesenchymal transition, a crucial step in the progression of many tumors to malignant disease.

  15. Transcriptional Regulation of Fucosyltransferase 1 Gene Expression in Colon Cancer Cells

    Directory of Open Access Journals (Sweden)

    Fumiko Taniuchi

    2013-01-01

    Full Text Available The α1,2-fucosyltransferase I (FUT1 enzyme is important for the biosynthesis of H antigens, Lewis B, and Lewis Y. In this study, we clarified the transcriptional regulation of FUT1 in the DLD-1 colon cancer cell line, which has high expression of Lewis B and Lewis Y antigens, expresses the FUT1 gene, and shows α1,2-fucosyltransferase (FUT activity. 5′-rapid amplification of cDNA ends revealed a FUT1 transcriptional start site −10 nucleotides upstream of the site registered at NM_000148 in the DataBase of Human Transcription Start Sites (DBTSS. Using the dual luciferase assay, FUT1 gene expression was shown to be regulated at the region −91 to −81 nt to the transcriptional start site, which contains the Elk-1 binding site. Site-directed mutagenesis of this region revealed the Elk-1 binding site to be essential for FUT1 transcription. Furthermore, transfection of the dominant negative Elk-1 gene, and the chromatin immunoprecipitation (CHIp assay, supported Elk-1-dependent transcriptional regulation of FUT1 gene expression in DLD-1 cells. These results suggest that a defined region in the 5′-flanking region of FUT1 is critical for FUT1 transcription and that constitutive gene expression of FUT1 is regulated by Elk-1 in DLD-1 cells.

  16. Combinatorial Gene Regulation through Kinetic Control of the Transcription Cycle.

    Science.gov (United States)

    Scholes, Clarissa; DePace, Angela H; Sánchez, Álvaro

    2017-01-25

    Cells decide when, where, and to what level to express their genes by "computing" information from transcription factors (TFs) binding to regulatory DNA. How is the information contained in multiple TF-binding sites integrated to dictate the rate of transcription? The dominant conceptual and quantitative model is that TFs combinatorially recruit one another and RNA polymerase to the promoter by direct physical interactions. Here, we develop a quantitative framework to explore kinetic control, an alternative model in which combinatorial gene regulation can result from TFs working on different kinetic steps of the transcription cycle. Kinetic control can generate a wide range of analog and Boolean computations without requiring the input TFs to be simultaneously bound to regulatory DNA. We propose experiments that will illuminate the role of kinetic control in transcription and discuss implications for deciphering the cis-regulatory "code."

  17. Transcriptional and epigenetic regulation of PPARγ expression during adipogenesis

    OpenAIRE

    Lee, Ji-Eun; Ge, Kai

    2014-01-01

    The nuclear receptor PPARγ is a master regulator of adipogenesis. PPARγ is highly expressed in adipose tissues and its expression is markedly induced during adipogenesis. In this review, we describe the current knowledge, as well as future directions, on transcriptional and epigenetic regulation of PPARγ expression during adipogenesis. Investigating the molecular mechanisms that control PPARγ expression during adipogenesis is critical for understanding the development of white and brown adipo...

  18. Regulation of cell proliferation by the E2F transcription factors

    DEFF Research Database (Denmark)

    Helin, K

    1998-01-01

    demonstrated that individual members of the E2F transcription factor family are likely to have distinct roles in mammalian development and homeostasis. Additional mechanisms regulating the activity of the E2F transcription factors have been reported, including subcellular localization and proteolysis of the E2......Experimental data generated in the past year have further emphasized the essential role for the E2F transcription factors in the regulation of cell proliferation. Genetic studies have shown that E2F activity is required for normal development in fruitflies, and the generation of E2F-1(-/-) mice has......Fs in the proteasomes. Novel target genes for the E2F transcription factors have been identified that link the E2Fs directly to the initiation of DNA replication....

  19. Dynamic Post-Transcriptional Regulation of HIV-1 Gene Expression

    Science.gov (United States)

    Kula, Anna; Marcello, Alessandro

    2012-01-01

    Gene expression of the human immunodeficiency virus type 1 (HIV-1) is a highly regulated process. Basal transcription of the integrated provirus generates early transcripts that encode for the viral products Tat and Rev. Tat promotes the elongation of RNA polymerase while Rev mediates the nuclear export of viral RNAs that contain the Rev-responsive RNA element (RRE). These RNAs are exported from the nucleus to allow expression of Gag-Pol and Env proteins and for the production of full-length genomic RNAs. A balance exists between completely processed mRNAs and RRE-containing RNAs. Rev functions as an adaptor that recruits cellular factors to re-direct singly spliced and unspliced viral RNAs to nuclear export. The aim of this review is to address the dynamic regulation of this post-transcriptional pathway in light of recent findings that implicate several novel cellular cofactors of Rev function. PMID:24832221

  20. Dynamic Post-Transcriptional Regulation of HIV-1 Gene Expression

    Directory of Open Access Journals (Sweden)

    Alessandro Marcello

    2012-07-01

    Full Text Available Gene expression of the human immunodeficiency virus type 1 (HIV-1 is a highly regulated process. Basal transcription of the integrated provirus generates early transcripts that encode for the viral products Tat and Rev. Tat promotes the elongation of RNA polymerase while Rev mediates the nuclear export of viral RNAs that contain the Rev-responsive RNA element (RRE. These RNAs are exported from the nucleus to allow expression of Gag-Pol and Env proteins and for the production of full-length genomic RNAs. A balance exists between completely processed mRNAs and RRE-containing RNAs. Rev functions as an adaptor that recruits cellular factors to re-direct singly spliced and unspliced viral RNAs to nuclear export. The aim of this review is to address the dynamic regulation of this post-transcriptional pathway in light of recent findings that implicate several novel cellular cofactors of Rev function.

  1. Uncovering transcriptional regulation of metabolism by using metabolic network topology

    DEFF Research Database (Denmark)

    Patil, Kiran Raosaheb; Nielsen, Jens

    2005-01-01

    therefore developed an algorithm that is based on hypothesis-driven data analysis to uncover the transcriptional regulatory architecture of metabolic networks. By using information on the metabolic network topology from genome-scale metabolic reconstruction, we show that it is possible to reveal patterns...... in the metabolic network that follow a common transcriptional response. Thus, the algorithm enables identification of so-called reporter metabolites (metabolites around which the most significant transcriptional changes occur) and a set of connected genes with significant and coordinated response to genetic...... changes induced by complex regulatory mechanisms coordinating the activity of different metabolic pathways. It is difficult to map such global transcriptional responses by using traditional methods, because many genes in the metabolic network have relatively small changes at their transcription level. We...

  2. The Csr system regulates genome-wide mRNA stability and transcription and thus gene expression in Escherichia coli.

    Science.gov (United States)

    Esquerré, Thomas; Bouvier, Marie; Turlan, Catherine; Carpousis, Agamemnon J; Girbal, Laurence; Cocaign-Bousquet, Muriel

    2016-04-26

    Bacterial adaptation requires large-scale regulation of gene expression. We have performed a genome-wide analysis of the Csr system, which regulates many important cellular functions. The Csr system is involved in post-transcriptional regulation, but a role in transcriptional regulation has also been suggested. Two proteins, an RNA-binding protein CsrA and an atypical signaling protein CsrD, participate in the Csr system. Genome-wide transcript stabilities and levels were compared in wildtype E. coli (MG1655) and isogenic mutant strains deficient in CsrA or CsrD activity demonstrating for the first time that CsrA and CsrD are global negative and positive regulators of transcription, respectively. The role of CsrA in transcription regulation may be indirect due to the 4.6-fold increase in csrD mRNA concentration in the CsrA deficient strain. Transcriptional action of CsrA and CsrD on a few genes was validated by transcriptional fusions. In addition to an effect on transcription, CsrA stabilizes thousands of mRNAs. This is the first demonstration that CsrA is a global positive regulator of mRNA stability. For one hundred genes, we predict that direct control of mRNA stability by CsrA might contribute to metabolic adaptation by regulating expression of genes involved in carbon metabolism and transport independently of transcriptional regulation.

  3. Transcriptional diversity and regulation across time and states

    DEFF Research Database (Denmark)

    Vitting-Seerup, Kristoffer

    and disease and can for example by themselves drive tumor formation. Unfortunately many researches are generally not considering isoforms when analyzing gene expression data leaving a large gap in our understanding of transcriptional output. Another recent finding is that an essential part of gene regulation...

  4. Transcriptional regulation of central amino acid metabolism in Lactococcus lactis

    NARCIS (Netherlands)

    Larsen, Rasmus

    2005-01-01

    This thesis describes the functional characterisation of the transcriptional regulators GlnR, ArgR and AhrC of Lactococcus lactis, which are responsible for the control of genes involved in the metabolism of the amino acids glutamine, glutamate and arginine. A chromosomal glnR deletion mutant was ma

  5. Serotonin transporter evolution and impact of polymorphic transcriptional regulation

    DEFF Research Database (Denmark)

    Søeby, Karen; Larsen, Svend Ask; Olsen, Line

    2005-01-01

    in the VNTRs of all mammalian SERT genes. The number of these putative binding sites varies proportionally to the length of the VNTR. We propose that the intronic VNTR have been selectively targeted through mammalian evolution to finetune transcriptional regulation of the serotonin expression....

  6. MRTF potentiates TEAD-YAP transcriptional activity causing metastasis.

    Science.gov (United States)

    Kim, Tackhoon; Hwang, Daehee; Lee, Dahye; Kim, Jeong-Hwan; Kim, Seon-Young; Lim, Dae-Sik

    2017-02-15

    Yes-associated protein (YAP) and myocardin-related transcription factor (MRTF) play similar roles and exhibit significant crosstalk in directing transcriptional responses to chemical and physical extracellular cues. The mechanism underlying this crosstalk, however, remains unclear. Here, we show MRTF family proteins bind YAP via a conserved PPXY motif that interacts with the YAP WW domain. This interaction allows MRTF to recruit NcoA3 to the TEAD-YAP transcriptional complex and potentiate its transcriptional activity. We show this interaction of MRTF and YAP is critical for LPA-induced cancer cell invasion in vitro and breast cancer metastasis to the lung in vivo We also demonstrate the significance of MRTF-YAP binding in regulation of YAP activity upon acute actin cytoskeletal damage. Acute actin disruption induces nucleo-cytoplasmic shuttling of MRTF, and this process underlies the LATS-independent regulation of YAP activity. Our results provide clear evidence of crosstalk between MRTF and YAP independent of the LATS kinases that normally act upstream of YAP signaling. Our results also suggest a mechanism by which extracellular stimuli can coordinate physiological events downstream of YAP. © 2016 The Authors.

  7. AT(1) receptor Gαq protein-independent signalling transcriptionally activates only a few genes directly, but robustly potentiates gene regulation from the β2-adrenergic receptor

    DEFF Research Database (Denmark)

    Christensen, Gitte Lund; Knudsen, Steen; Schneider, Mikael;

    2011-01-01

    of Gαq protein-dependent and -independent regulation of AT(1)R mediated gene expression. We found angiotensin II to regulate 212 genes, whereas Gαq-independent signalling obtained with the biased agonist, SII angiotensin II only regulated few genes. Interestingly, SII angiotensin II, like Ang II vastly...

  8. Transcription co-activator Arabidopsis ANGUSTIFOLIA3 (AN3) regulates water-use efficiency and drought tolerance by modulating stomatal density and improving root architecture by the transrepression of YODA (YDA).

    Science.gov (United States)

    Meng, Lai-Sheng; Yao, Shun-Qiao

    2015-09-01

    One goal of modern agriculture is the improvement of plant drought tolerance and water-use efficiency (WUE). Although stomatal density has been linked to WUE, the causal molecular mechanisms and engineered alternations of this relationship are not yet fully understood. Moreover, YODA (YDA), which is a MAPKK kinase gene, negatively regulates stomatal development. BR-INSENSITIVE 2 interacts with phosphorylates and inhibits YDA. However, whether YDA is modulated in the transcriptional level is still unclear. Plants lacking ANGUSTIFOLIA3 (AN3) activity have high drought stress tolerance because of low stomatal densities and improved root architecture. Such plants also exhibit enhanced WUE through declining transpiration without a demonstrable reduction in biomass accumulation. AN3 negatively regulated YDA expression at the transcriptional level by target-gene analysis. Chromatin immunoprecipitation analysis indicated that AN3 was associated with a region of the YDA promoter in vivo. YDA mutation significantly decreased the stomatal density and root length of an3 mutant, thus proving the participation of YDA in an3 drought tolerance and WUE enhancement. These components form an AN3-YDA complex, which allows the integration of water deficit stress signalling into the production or spacing of stomata and cell proliferation, thus leading to drought tolerance and enhanced WUE.

  9. Transcript Regulation of Human Telomerase Reverse Transcriptase by c-myc and mad1

    Institute of Scientific and Technical Information of China (English)

    Lin ZOU; Peng-Hui ZHANG; Chun-Li LUO; Zhi-Guang TU

    2005-01-01

    Telomerase activity is highly positive correlated to most malignant neoplasms. Human telomerase reverse transcriptase (hTERT) is the rate-limiting factor of telomerase activity. Recent studies have shown that the expression of hTERT is mainly determined by its transcript regulation. Among the transcript regulation factors of hTERT, c-myc and mad1 are well known. Here, we constructed c-myc and mad1 eukaryotic expression vectors, then co-transfected them with the wild-type (Tw) or mutant hTERT promoter (Td)luciferase reporter plasmid, which were double-mutated in the E-box sequences from CACGTG to CACCTG of Tw. The change of luciferase activity in different cells was detected. The results showed that Tw was obviously activated in T24 and EJ bladder cancer cells, but not in normal fibrocytes. c-myc and mad1 had positive and negative effects respectively on the Tw transcript in a dose-dependent manner, while the roles of c-myc and mad1 in regulating the Td transcript were reversed. c-myc combined with mad1 can downregulate Tw but not Td. These observations indicate that c-myc and mad1 can regulate the hTERT transcript in a different manner in hTERT positive cells, but not in normal cells. This may provide an insight into some telomerase-related carcinogenesis mechanisms.

  10. Sp1 and KLF15 regulate basal transcription of the human LRP5 gene

    Directory of Open Access Journals (Sweden)

    Zou Yongxin

    2010-02-01

    Full Text Available Abstract Background LRP5, a member of the low density lipoprotein receptor superfamily, regulates diverse developmental processes in embryogenesis and maintains physiological homeostasis in adult organisms. However, how the expression of human LRP5 gene is regulated remains unclear. Results In order to characterize the transcriptional regulation of human LRP5 gene, we cloned the 5' flanking region and evaluated its transcriptional activity in a luciferase reporter system. We demonstrated that both KLF15 and Sp1 binding sites between -72 bp and -53 bp contribute to the transcriptional activation of human LRP5 promoter. Chromatin immunoprecipitation assay demonstrated that the ubiquitous transcription factors KLF15 and Sp1 bind to this region. Using Drosophila SL2 cells, we showed that KLF15 and Sp1 trans-activated the LRP5 promoter in a manner dependent on the presence of Sp1-binding and KLF15-binding motifs. Conclusions Both KLF15 and Sp1 binding sites contribute to the basal activity of human LRP5 promoter. This study provides the first insight into the mechanisms by which transcription of human LRP5 gene is regulated.

  11. HIPK2: a versatile switchboard regulating the transcription machinery and cell death.

    Science.gov (United States)

    Calzado, Marco A; Renner, Florian; Roscic, Ana; Schmitz, M Lienhard

    2007-01-15

    Homeodomain interacting protein kinase 2 (HIPK2) is an evolutionary conserved serine/threonine kinase that regulates gene expression by phosphorylation of transcription factors and accessory components of the transcription machinery. HIPK2 is activated in response to DNA-damaging agents or morphogenic signals and accordingly HIPK2-guided gene expression programs trigger differentiation and development or alternatively apoptosis. The kinase contributes to the regulation of remarkably diverse pathways such as p53 activation or Wnt signaling. Here we discuss recent findings from biochemical and functional experiments that allow a deeper understanding of the pleiotropic effects mediated by HIPK2.

  12. Bidirectional Transcription Arises from Two Distinct Hubs of Transcription Factor Binding and Active Chromatin.

    Science.gov (United States)

    Scruggs, Benjamin S; Gilchrist, Daniel A; Nechaev, Sergei; Muse, Ginger W; Burkholder, Adam; Fargo, David C; Adelman, Karen

    2015-06-18

    Anti-sense transcription originating upstream of mammalian protein-coding genes is a well-documented phenomenon, but remarkably little is known about the regulation or function of anti-sense promoters and the non-coding RNAs they generate. Here we define at nucleotide resolution the divergent transcription start sites (TSSs) near mouse mRNA genes. We find that coupled sense and anti-sense TSSs precisely define the boundaries of a nucleosome-depleted region (NDR) that is highly enriched in transcription factor (TF) motifs. Notably, as the distance between sense and anti-sense TSSs increases, so does the size of the NDR, the level of signal-dependent TF binding, and gene activation. We further discover a group of anti-sense TSSs in macrophages with an enhancer-like chromatin signature. Interestingly, this signature identifies divergent promoters that are activated during immune challenge. We propose that anti-sense promoters serve as platforms for TF binding and establishment of active chromatin to further regulate or enhance sense-strand mRNA expression.

  13. Thioredoxin interacting protein inhibits hypoxia-inducible factor transcriptional activity

    Science.gov (United States)

    Farrell, Michael R; Rogers, Lynette K; Liu, Yusen; Welty, Stephen E; Tipple, Trent E

    2010-01-01

    Vascular endothelial growth factor (VEGF) is required for proper lung development and is transcriptionally regulated in alveolar epithelial cells by hypoxia inducible factor (HIF). Previous findings in a newborn mouse model of bronchopulmonary dysplasia (BPD) suggest that thioredoxin interacting protein (Txnip) is a novel regulator of VEGF expression. The present studies were designed to test the hypothesis that Txnip negatively regulates VEGF through effects on HIF-mediated gene expression. To test this hypothesis, we first examined the levels of VEGF and Txnip protein in the lungs of 1 day-old newborn and E19 embryos and detected a significant inverse correlation. To elucidate the mechanisms underlying this relationship, we studied the effects of Txnip overexpression on HIF-mediated transcription using murine lung epithelial (MLE-12) cells. Overexpression of Txnip inhibited HIF-mediated reporter activity in both hypoxia and room air. Suppression of HIF activity by Txnip appeared to be independent of the ability of Txnip to bind to thioredoxin. Thus, our studies support a model in which Txnip is a potentially critical regulator of HIF-mediated gene transcription in the murine lung. Alterations in Txnip expression could alter lung VEGF expression in prematurely born human infants and contribute to the development of BPD. PMID:20692333

  14. Mlc is a transcriptional activator with a key role in integrating cyclic AMP receptor protein and integration host factor regulation of leukotoxin RNA synthesis in Aggregatibacter actinomycetemcomitans

    Science.gov (United States)

    Aggregatibacter actinomycetemcomitans, a periodontal pathogen, synthesizes leukotoxin (LtxA), a protein that helps the bacterium evade the host immune response. Transcription of the ltxA operon is induced during anaerobic growth. The cAMP receptor protein (CRP) indirectly increases ltxA expression...

  15. Large-scale transcriptome data reveals transcriptional activity of fission yeast LTR retrotransposons

    DEFF Research Database (Denmark)

    Mourier, Tobias; Willerslev, Eske

    2010-01-01

    BACKGROUND: Retrotransposons are transposable elements that proliferate within eukaryotic genomes through a process involving reverse transcription. The numbers of retrotransposons within genomes and differences between closely related species may yield insight into the evolutionary history...... of transcriptional activity are observed from both strands of solitary LTR sequences. Transcriptome data collected during meiosis suggests that transcription of solitary LTRs is correlated with the transcription of nearby protein-coding genes. CONCLUSIONS: Presumably, the host organism negatively regulates...

  16. Observation of Transcription Regulation in the Mouse Heart Nuclear DNA Fragments and the Specific-protein Interaction by AFM

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    Using atom force microscopy (AFM), in vitro transcription, PAGE and other experimental technologies, it is observed that, in active genes of mice (Balb/c) nuclear DNA fragments of non-transcriptional state, only regulation sequences at both ends are associated with scaffold proteins (indissociable proteins) and some transcriptional factors such as complexes (dissociable proteins) made of gene-coding proteins and specific auxiliary small molecules, while there are no combining proteins in intermediate coding sequences. However, in active genes of transcriptional state, both regulation sequences and intermediate coding sequences are associated with active transcriptional factors by non-covalent bonds.This paper shows the prospective application of AFM observation and in vitro transcription in the research on gene expression and regulation. It also offers some theoretical basis for localization of specific genes in human genomes.

  17. A CONSTANS-like transcriptional activator, OsCOL13, functions as a negative regulator of flowering downstream of OsphyB and upstream of Ehd1 in rice.

    Science.gov (United States)

    Sheng, Peike; Wu, Fuqing; Tan, Junjie; Zhang, Huan; Ma, Weiwei; Chen, Liping; Wang, Jiachang; Wang, Jie; Zhu, Shanshan; Guo, Xiuping; Wang, Jiulin; Zhang, Xin; Cheng, Zhijun; Bao, Yiqun; Wu, Chuanyin; Liu, Xuanming; Wan, Jianmin

    2016-09-01

    Flowering time determines the adaptability of crop plants to different local environments, thus being one of the most important agronomic traits targeted in breeding programs. Photoperiod is one of the key factors that control flowering in plant. A number of genes that participate in the photoperiod pathway have been characterized in long-day plants such as Arabidopsis, as well as in short-day plants such as Oryza sativa. Of those, CONSTANS (CO) as a floral integrator promotes flowering in Arabidopsis under long day conditions. In rice, Heading date1 (Hd1), a homologue of CO, functions in an opposite way, which inhibits flowering under long day conditions and induces flowering under short day conditions. Here, we show that another CONSTANS-like (COL) gene, OsCOL13, negatively regulates flowering in rice under both long and short day conditions. Overexpression of OsCOL13 delays flowering regardless of day length. We also demonstrated that OsCOL13 has a constitutive and rhythmic expression pattern, and that OsCOL13 is localized to the nucleus. OsCOL13 displays transcriptional activation activity in the yeast assays and likely forms homodimers in vivo. OsCOL13 suppresses the florigen genes Hd3a and RFT1 by repressing Ehd1, but has no relationship with other known Ehd1 regulators as determined by using mutants or near isogenic lines. In addition, the transcriptional level of OsCOL13 significantly decreased in the osphyb mutant, but remained unchanged in the osphya and osphyc mutants. Thus, we conclude that OsCOL13 functions as a negative regulator downstream of OsphyB and upstream of Ehd1 in the photoperiodic flowering in rice.

  18. Genome-wide signatures of transcription factor activity: connecting transcription factors, disease, and small molecules.

    Directory of Open Access Journals (Sweden)

    Jing Chen

    Full Text Available Identifying transcription factors (TF involved in producing a genome-wide transcriptional profile is an essential step in building mechanistic model that can explain observed gene expression data. We developed a statistical framework for constructing genome-wide signatures of TF activity, and for using such signatures in the analysis of gene expression data produced by complex transcriptional regulatory programs. Our framework integrates ChIP-seq data and appropriately matched gene expression profiles to identify True REGulatory (TREG TF-gene interactions. It provides genome-wide quantification of the likelihood of regulatory TF-gene interaction that can be used to either identify regulated genes, or as genome-wide signature of TF activity. To effectively use ChIP-seq data, we introduce a novel statistical model that integrates information from all binding "peaks" within 2 Mb window around a gene's transcription start site (TSS, and provides gene-level binding scores and probabilities of regulatory interaction. In the second step we integrate these binding scores and regulatory probabilities with gene expression data to assess the likelihood of True REGulatory (TREG TF-gene interactions. We demonstrate the advantages of TREG framework in identifying genes regulated by two TFs with widely different distribution of functional binding events (ERα and E2f1. We also show that TREG signatures of TF activity vastly improve our ability to detect involvement of ERα in producing complex diseases-related transcriptional profiles. Through a large study of disease-related transcriptional signatures and transcriptional signatures of drug activity, we demonstrate that increase in statistical power associated with the use of TREG signatures makes the crucial difference in identifying key targets for treatment, and drugs to use for treatment. All methods are implemented in an open-source R package treg. The package also contains all data used in the analysis

  19. Immunosuppressive effect of isopropanol: down-regulation of cytokine production results from the alteration of discrete transcriptional pathways in activated lymphocytes.

    Science.gov (United States)

    Désy, Olivier; Carignan, Damien; Caruso, Manuel; de Campos-Lima, Pedro O

    2008-08-15

    Isopropanol (IPA) is widely used in household applications and constitutes a leading cause of acute alcohol intoxication second only to ethanol. Although the effects of ethanol on the immune system have been extensively studied, far fewer data are available on IPA. Given the structural similarity between the two molecules, we hypothesized that IPA could as well have immunomodulatory properties. We report here that acute IPA exposure is detrimental to human T lymphocyte and NK cell activity in vitro in concentrations as low as 0.08-0.16% (13-26 mM). IPA treatment did not affect receptor-mediated early signaling but had a reproducible and dose-dependent effect on the nuclear translocation of NFAT and AP-1. Furthermore, we show in a model of acute IPA intoxication that animals became immunosuppressed as judged by their reduced ability to release IL-2 and IFN-gamma in the serum in response to staphylococcal enterotoxin B. This effect was also associated to the down-regulation of TNF-alpha production and was sufficiently strong to rescue susceptible animals from enterotoxin-induced toxic shock. Our results suggest that IPA is potentially immunosuppressive to the adaptive and innate immune system and have broad significance given the exposure of the general population to this ubiquitous chemical.

  20. MOF Acetyl Transferase Regulates Transcription and Respiration in Mitochondria.

    Science.gov (United States)

    Chatterjee, Aindrila; Seyfferth, Janine; Lucci, Jacopo; Gilsbach, Ralf; Preissl, Sebastian; Böttinger, Lena; Mårtensson, Christoph U; Panhale, Amol; Stehle, Thomas; Kretz, Oliver; Sahyoun, Abdullah H; Avilov, Sergiy; Eimer, Stefan; Hein, Lutz; Pfanner, Nikolaus; Becker, Thomas; Akhtar, Asifa

    2016-10-20

    A functional crosstalk between epigenetic regulators and metabolic control could provide a mechanism to adapt cellular responses to environmental cues. We report that the well-known nuclear MYST family acetyl transferase MOF and a subset of its non-specific lethal complex partners reside in mitochondria. MOF regulates oxidative phosphorylation by controlling expression of respiratory genes from both nuclear and mtDNA in aerobically respiring cells. MOF binds mtDNA, and this binding is dependent on KANSL3. The mitochondrial pool of MOF, but not a catalytically deficient mutant, rescues respiratory and mtDNA transcriptional defects triggered by the absence of MOF. Mof conditional knockout has catastrophic consequences for tissues with high-energy consumption, triggering hypertrophic cardiomyopathy and cardiac failure in murine hearts; cardiomyocytes show severe mitochondrial degeneration and deregulation of mitochondrial nutrient metabolism and oxidative phosphorylation pathways. Thus, MOF is a dual-transcriptional regulator of nuclear and mitochondrial genomes connecting epigenetics and metabolism.

  1. In silico comparative genomic analysis of GABAA receptor transcriptional regulation

    Directory of Open Access Journals (Sweden)

    Joyce Christopher J

    2007-06-01

    Full Text Available Abstract Background Subtypes of the GABAA receptor subunit exhibit diverse temporal and spatial expression patterns. In silico comparative analysis was used to predict transcriptional regulatory features in individual mammalian GABAA receptor subunit genes, and to identify potential transcriptional regulatory components involved in the coordinate regulation of the GABAA receptor gene clusters. Results Previously unreported putative promoters were identified for the β2, γ1, γ3, ε, θ and π subunit genes. Putative core elements and proximal transcriptional factors were identified within these predicted promoters, and within the experimentally determined promoters of other subunit genes. Conserved intergenic regions of sequence in the mammalian GABAA receptor gene cluster comprising the α1, β2, γ2 and α6 subunits were identified as potential long range transcriptional regulatory components involved in the coordinate regulation of these genes. A region of predicted DNase I hypersensitive sites within the cluster may contain transcriptional regulatory features coordinating gene expression. A novel model is proposed for the coordinate control of the gene cluster and parallel expression of the α1 and β2 subunits, based upon the selective action of putative Scaffold/Matrix Attachment Regions (S/MARs. Conclusion The putative regulatory features identified by genomic analysis of GABAA receptor genes were substantiated by cross-species comparative analysis and now require experimental verification. The proposed model for the coordinate regulation of genes in the cluster accounts for the head-to-head orientation and parallel expression of the α1 and β2 subunit genes, and for the disruption of transcription caused by insertion of a neomycin gene in the close vicinity of the α6 gene, which is proximal to a putative critical S/MAR.

  2. Estrogen Receptor α (ERα) and Estrogen Related Receptor α (ERRα) are both transcriptional regulators of the Runx2-I isoform.

    Science.gov (United States)

    Kammerer, Martial; Gutzwiller, Sabine; Stauffer, Daniela; Delhon, Isabelle; Seltenmeyer, Yves; Fournier, Brigitte

    2013-04-30

    Runx2 is a master regulator of bone development and has also been described as an oncogene. Estrogen Receptor α (ERα) and Estrogen Related Receptor α (ERRα), both implicated in bone metabolism and breast cancer, have been shown to share common transcriptional targets. Here, we show that ERα is a positive regulator of Runx2-I transcription. Moreover, ERRα can act as a transcriptional activator of Runx2-I in presence of peroxisome proliferator activated receptor gamma coactivator-1 alpha (PGC-1α). In contrast, ERRα behaves as a negative regulator of Runx2-I transcription in presence of PGC-1β. ERα and ERRα cross-talk via a common estrogen receptor response element on the Runx2-I promoter. In addition, estrogen regulates PGC-1β that in turn is able to modulate both ERα and ERRα transcriptional activity.

  3. Insulin transcriptionally regulates argininosuccinate synthase to maintain vascular endothelial function.

    Science.gov (United States)

    Haines, Ricci J; Corbin, Karen D; Pendleton, Laura C; Meininger, Cynthia J; Eichler, Duane C

    2012-04-27

    Diminished vascular endothelial cell nitric oxide (NO) production is a major factor in the complex pathogenesis of diabetes mellitus. In this report, we demonstrate that insulin not only maintains endothelial NO production through regulation of endothelial nitric oxide synthase (eNOS), but also via the regulation of argininosuccinate synthase (AS), which is the rate-limiting step of the citrulline-NO cycle. Using serum starved, cultured vascular endothelial cells, we show that insulin up-regulates AS and eNOS transcription to support NO production. Moreover, we show that insulin enhances NO production in response to physiological cues such as bradykinin. To translate these results to an in vivo model, we show that AS transcription is diminished in coronary endothelial cells isolated from rats with streptozotocin (STZ)-induced diabetes. Importantly, we demonstrate restoration of AS and eNOS transcription by insulin treatment in STZ-diabetic rats, and show that this restoration was accompanied by improved endothelial function as measured by endothelium-dependent vasorelaxation. Overall, this report demonstrates, both in cell culture and whole animal studies, that insulin maintains vascular function, in part, through the maintenance of AS transcription, thus ensuring an adequate supply of arginine to maintain vascular endothelial response to physiological cues. Copyright © 2012 Elsevier Inc. All rights reserved.

  4. Mechanisms of post-transcriptional gene regulation in bacterial biofilms

    Directory of Open Access Journals (Sweden)

    Viveka eVadyvaloo

    2014-03-01

    Full Text Available Abstract Biofilms are characterized by a dense multicellular community of microorganisms that can be formed by the attachment of bacteria to an inert surface and to each other. The development of biofilm involves the initial attachment of planktonic bacteria to a surface, followed by replication, cell-to-cell adhesion to form microcolonies, maturation and detachment. Mature biofilms are embedded in a self-produced extracellular polymeric matrix composed primarily of bacterial-derived exopolysaccharides, specialized proteins, adhesins and occasionally DNA. Because the synthesis and assembly of biofilm matrix components is an exceptionally complex process, the transition between its different phases requires the coordinate expression and simultaneous regulation of many genes by complex genetic networks involving all levels of gene regulation. The finely controlled intracellular level of the chemical second messenger molecule, cyclic-di-GMP is central to the post-transcriptional mechanisms governing the switch between the motile planktonic lifestyle and the sessile biofilm forming state in many bacteria. Several other post-transcriptional regulatory mechanisms are known to dictate biofilm development and assembly and these include RNA-binding proteins, small non-coding RNAs, toxin-antitoxin systems, riboswitches and RNases. Post-transcriptional regulation is therefore a powerful molecular mechanism employed by bacteria to rapidly adjust to the changing environment and to fine tune gene expression to the developmental needs of the cell. In this review, we discuss post-transcriptional mechanisms that influence the biofilm developmental cycle in a variety of pathogenic bacteria.

  5. ChrR positively regulates transcription of the Rhodobacter sphaeroides cytochrome c2 gene.

    Science.gov (United States)

    Schilke, B A; Donohue, T J

    1995-04-01

    Transcription of the Rhodobacter sphaeroides cytochrome c2 gene (cycA) is negatively regulated by both the presence of oxygen and intermediates in tetrapyrrole biosynthesis. A mutation responsible for uncoupling cycA transcription from tetrapyrrole availability was localized to a gene (chrR) that encodes a 357-amino-acid protein. Analysis of a defined chrR null mutation indicated that this protein positively regulated cycA transcription. From this and other results, it appeared that the positive action of ChrR on cycA transcription is blocked by altering the availability of either heme or some intermediate in tetrapyrrole biosynthesis. A single missense mutation which substitutes an Arg for a Cys at residue 182 of ChrR (C182R) was shown to be necessary and sufficient for the increased cycA transcription seen in the mutant strain Chr4. Thus, it appears that this C182R substitution generated an altered-function form of ChrR. In addition, by analyzing cycA transcription in delta ChrR strains, we showed that ChrR was not required for increased cycA transcription under anaerobic conditions. Instead, our results indicated that ChrR and the response regulator PrrA (J. M. Eraso and S. Kaplan, J. Bacteriol. 176:32-43, 1994) functioned independently at the upstream cycA promoter that is activated under anaerobic conditions.

  6. Transcriptional and post-transcriptional regulation of nucleotide excision repair genes in human cells

    Energy Technology Data Exchange (ETDEWEB)

    Lefkofsky, Hailey B. [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Veloso, Artur [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI (United States); Bioinformatics Program, Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, MI (United States); Ljungman, Mats, E-mail: ljungman@umich.edu [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Environmental Health Sciences, School of Public Health, University of Michigan, Ann Arbor, MI (United States)

    2015-06-15

    Nucleotide excision repair (NER) removes DNA helix-distorting lesions induced by UV light and various chemotherapeutic agents such as cisplatin. These lesions efficiently block the elongation of transcription and need to be rapidly removed by transcription-coupled NER (TC-NER) to avoid the induction of apoptosis. Twenty-nine genes have been classified to code for proteins participating in nucleotide excision repair (NER) in human cells. Here we explored the transcriptional and post-transcriptional regulation of these NER genes across 13 human cell lines using Bru-seq and BruChase-seq, respectively. Many NER genes are relatively large in size and therefore will be easily inactivated by UV-induced transcription-blocking lesions. Furthermore, many of these genes produce transcripts that are rather unstable. Thus, these genes are expected to rapidly lose expression leading to a diminished function of NER. One such gene is ERCC6 that codes for the CSB protein critical for TC-NER. Due to its large gene size and high RNA turnover rate, the ERCC6 gene may act as dosimeter of DNA damage so that at high levels of damage, ERCC6 RNA levels would be diminished leading to the loss of CSB expression, inhibition of TC-NER and the promotion of cell death.

  7. Characterization of E2F8, a novel E2F-like cell-cycle regulated repressor of E2F-activated transcription

    DEFF Research Database (Denmark)

    Christensen, Jesper; Cloos, Paul; Toftegaard, Ulla

    2005-01-01

    . Sequence analysis of E2F8 predicts the presence of two distinct E2F-related DNA binding domains suggesting that E2F8 and, the recently, identified E2F7 form a subgroup within the E2F family. We show that E2F transcription factors bind the E2F8 promoter in vivo and that expression of E2F8 is being induced...... at the G1/S transition. Purified recombinant E2F8 binds specifically to a consensus E2F-DNA-binding site indicating that E2F8, like E2F7, binds DNA without the requirement of co-factors such as DP1. E2F8 inhibits E2F-driven promoters suggesting that E2F8 is transcriptional repressor like E2F7. Ectopic...

  8. Serotonin transporter evolution and impact of polymorphic transcriptional regulation

    DEFF Research Database (Denmark)

    Søeby, Karen; Larsen, Svend Ask; Olsen, Line

    2005-01-01

    extensively across the great apes and monkeys as well as in rodents while it is absent in non-mammals. As in humans, the VNTR sequence may be polymorphic within species and thus it may underlie both inter- and intraspecies differences. Also, we find new putative binding sites for several transcription factors...... in the VNTRs of all mammalian SERT genes. The number of these putative binding sites varies proportionally to the length of the VNTR. We propose that the intronic VNTR have been selectively targeted through mammalian evolution to finetune transcriptional regulation of the serotonin expression....

  9. DNA supercoiling and aerobic regulation of transcription from the Klebsiella pneumoniae nifLA promoter.

    Science.gov (United States)

    Dixon, R A; Henderson, N C; Austin, S

    1988-11-11

    Expression from the K. pneumoniae nifLA promoter is oxygen sensitive and is also inhibited by the DNA gyrase inhibitor coumermycin A1 under anaerobic growth conditions. The activity of this promoter was found to be highly sensitive to changes in DNA topology in vitro. Transcription was completely dependent on negative supercoiling at physiological salt concentrations although transcription from linear or fully relaxed closed circular templates was detectable at KCl concentrations lower than 50 mM. These observations suggest that aerobic regulation of nif transcription may be mediated through the level of DNA supercoiling.

  10. Transcriptional regulation of early T-cell development in the thymus.

    Science.gov (United States)

    Seo, Wooseok; Taniuchi, Ichiro

    2016-03-01

    T-cell development occurs in multipotent progenitors arriving in the thymus, which provides a highly specialized microenvironment. Specification and sequential commitment processes to T cells begin in early thymic progenitors upon receiving thymus-specific environmental cues, resulting in the activation of the genetically programmed transcriptional cascade that includes turning on and off numerous transcription factors in a precise manner. Thus, early thymocyte differentiation has been an excellent model system to study cell differentiation processes. This review summarizes recent advances in our knowledge on thymic T-cell development from newly arrived multipotent T-cell progenitors to fully committed T-cell precursors, from the transcriptional regulation perspective.

  11. Mechanisms of transcriptional activation of the stimulator of interferon genes by transcription factors CREB and c-Myc.

    Science.gov (United States)

    Wang, Yan-Yan; Jin, Rui; Zhou, Guo-Ping; Xu, Hua-Guo

    2016-12-20

    Stimulator of interferon genes (STING) plays an important role in host defense, autoimmune disease, osteoclast differentiation and anti-tumor response. Although many downstream targets have been studied in depth, the regulation of STING gene expression remains largely unknown. Here we demonstrate that transcription factors CREB and c-Myc maintain the transcriptional activity of STING. By 5'-rapid amplification of cDNA ends analysis, we identified the transcriptional start site (TSS) of STING. We illustrated that the region -124/+1 relative to TSS was sufficient for full promoter activity by a series of 5' deletion promoter constructs. Transcriptional activity of the STING minimal promoter was dependent on CREB and c-Myc binding motifs and was abolished after mutation of these two DNA elements. Chromatin immunoprecipitation assays demonstrated that transcription factors CREB and c-Myc bind to STING promoter in vivo. Overexpression of CREB and c-Myc increased the STING promoter activity. Meanwhile, knocking-down of CREB and c-Myc by a small interfering RNA (siRNA) strategy markedly reduced endogenous STING expression. In summary, these results demonstrated that transcription factors CREB and c-Myc are involved in the regulation of STING transcription.

  12. Human DJ-1-specific Transcriptional Activation of Tyrosine Hydroxylase Gene*

    Science.gov (United States)

    Ishikawa, Shizuma; Taira, Takahiro; Takahashi-Niki, Kazuko; Niki, Takeshi; Ariga, Hiroyoshi; Iguchi-Ariga, Sanae M. M.

    2010-01-01

    Loss-of-function mutation in the DJ-1 gene causes a subset of familial Parkinson disease. The mechanism underlying DJ-1-related selective vulnerability in the dopaminergic pathway is, however, not known. DJ-1 has multiple functions, including transcriptional regulation, and one of transcriptional target genes for DJ-1 is the tyrosine hydroxylase (TH) gene, the product of which is a key enzyme for dopamine biosynthesis. It has been reported that DJ-1 is a neuroprotective transcriptional co-activator that sequesters a transcriptional co-repressor polypyrimidine tract-binding protein-associated splicing factor (PSF) from the TH gene promoter. In this study, we found that knockdown of human DJ-1 by small interference RNA in human dopaminergic cell lines attenuated TH gene expression and 4-dihydroxy-l-phenylalanine production but that knockdown or knock-out of mouse DJ-1 in mouse cell lines or in mice did not affect such expression and TH activity. In reporter assays using the human TH gene promoter linked to the luciferase gene, stimulation of TH promoter activity was observed in human cells, but not mouse cells, that had been transfected with DJ-1. Although human DJ-1 and mouse DJ-1 were associated either with human or with mouse PSF, TH promoter activity inhibited by PSF was restored by human DJ-1 but not by mouse DJ-1. Chromatin immunoprecipitation assays revealed that the complex of PSF with DJ-1 bound to the human but not the mouse TH gene promoter. These results suggest a novel species-specific transcriptional regulation of the TH promoter by DJ-1 and one of the mechanisms for no reduction of TH in DJ-1-knock-out mice. PMID:20938049

  13. Regulation of the BMP Signaling-Responsive Transcriptional Network in the Drosophila Embryo.

    Science.gov (United States)

    Deignan, Lisa; Pinheiro, Marco T; Sutcliffe, Catherine; Saunders, Abbie; Wilcockson, Scott G; Zeef, Leo A H; Donaldson, Ian J; Ashe, Hilary L

    2016-07-01

    The BMP signaling pathway has a conserved role in dorsal-ventral axis patterning during embryonic development. In Drosophila, graded BMP signaling is transduced by the Mad transcription factor and opposed by the Brinker repressor. In this study, using the Drosophila embryo as a model, we combine RNA-seq with Mad and Brinker ChIP-seq to decipher the BMP-responsive transcriptional network underpinning differentiation of the dorsal ectoderm during dorsal-ventral axis patterning. We identify multiple new BMP target genes, including positive and negative regulators of EGF signaling. Manipulation of EGF signaling levels by loss- and gain-of-function studies reveals that EGF signaling negatively regulates embryonic BMP-responsive transcription. Therefore, the BMP gene network has a self-regulating property in that it establishes a balance between its activity and that of the antagonistic EGF signaling pathway to facilitate correct patterning. In terms of BMP-dependent transcription, we identify key roles for the Zelda and Zerknüllt transcription factors in establishing the resulting expression domain, and find widespread binding of insulator proteins to the Mad and Brinker-bound genomic regions. Analysis of embryos lacking the BEAF-32 insulator protein shows reduced transcription of a peak BMP target gene and a reduction in the number of amnioserosa cells, the fate specified by peak BMP signaling. We incorporate our findings into a model for Mad-dependent activation, and discuss its relevance to BMP signal interpretation in vertebrates.

  14. Transcriptional Regulation of Neural Retina Leucine Zipper (Nrl), a Photoreceptor Cell Fate Determinant*

    Science.gov (United States)

    Montana, Cynthia L.; Lawrence, Karen A.; Williams, Natecia L.; Tran, Nicholas M.; Peng, Guang-Hua; Chen, Shiming; Corbo, Joseph C.

    2011-01-01

    The transcription factor neural retina leucine zipper (Nrl) is a critical determinant of rod photoreceptor cell fate and a key regulator of rod differentiation. Nrl−/− rod precursors fail to turn on rod genes and instead differentiate as cones. Furthermore, NRL mutations in humans cause retinitis pigmentosa. Despite the developmental and clinical significance of this gene, little is known about the transcriptional regulation of Nrl itself. In this study, we sought to define the cis- and trans-acting factors responsible for initiation and maintenance of Nrl transcription in the mouse retina. Utilizing a quantitative mouse retinal explant electroporation assay, we discovered a phylogenetically conserved, 30-base pair region immediately upstream of the transcription start site that is required for Nrl promoter activity. This region contains binding sites for the retinal transcription factors CRX, OTX2, and RORβ, and point mutations in these sites completely abolish promoter activity in living retinas. Gel-shift experiments show that CRX, OTX2, and RORβ can bind to the critical region in vitro, whereas ChIP experiments demonstrate binding of CRX and OTX2 to the critical region in vivo. Thus, our results indicate that CRX, OTX2, and RORβ directly regulate Nrl transcription by binding to critical sites within the Nrl promoter. We propose a model in which Nrl expression is primarily initiated by OTX2 and RORβ and later maintained at high levels by CRX and RORβ. PMID:21865162

  15. Transcriptional regulation of neural retina leucine zipper (Nrl), a photoreceptor cell fate determinant.

    Science.gov (United States)

    Montana, Cynthia L; Lawrence, Karen A; Williams, Natecia L; Tran, Nicholas M; Peng, Guang-Hua; Chen, Shiming; Corbo, Joseph C

    2011-10-21

    The transcription factor neural retina leucine zipper (Nrl) is a critical determinant of rod photoreceptor cell fate and a key regulator of rod differentiation. Nrl(-/-) rod precursors fail to turn on rod genes and instead differentiate as cones. Furthermore, NRL mutations in humans cause retinitis pigmentosa. Despite the developmental and clinical significance of this gene, little is known about the transcriptional regulation of Nrl itself. In this study, we sought to define the cis- and trans-acting factors responsible for initiation and maintenance of Nrl transcription in the mouse retina. Utilizing a quantitative mouse retinal explant electroporation assay, we discovered a phylogenetically conserved, 30-base pair region immediately upstream of the transcription start site that is required for Nrl promoter activity. This region contains binding sites for the retinal transcription factors CRX, OTX2, and RORβ, and point mutations in these sites completely abolish promoter activity in living retinas. Gel-shift experiments show that CRX, OTX2, and RORβ can bind to the critical region in vitro, whereas ChIP experiments demonstrate binding of CRX and OTX2 to the critical region in vivo. Thus, our results indicate that CRX, OTX2, and RORβ directly regulate Nrl transcription by binding to critical sites within the Nrl promoter. We propose a model in which Nrl expression is primarily initiated by OTX2 and RORβ and later maintained at high levels by CRX and RORβ.

  16. GSK3beta is a negative regulator of the transcriptional coactivator MAML1.

    Science.gov (United States)

    Saint Just Ribeiro, Mariana; Hansson, Magnus L; Lindberg, Mikael J; Popko-Scibor, Anita E; Wallberg, Annika E

    2009-11-01

    Glycogen synthase kinase 3beta (GSK3beta) is involved in several cellular signaling systems through regulation of the activity of diverse transcription factors such as Notch, p53 and beta-catenin. Mastermind-like 1 (MAML1) was originally identified as a Notch coactivator, but has also been reported to function as a transcriptional coregulator of p53, beta-catenin and MEF2C. In this report, we show that active GSK3beta directly interacts with the MAML1 N-terminus and decreases MAML1 transcriptional activity, suggesting that GSK3beta might target a coactivator in its regulation of gene expression. We have previously shown that MAML1 increases global acetylation of histones, and here we show that the GSK3 inhibitor SB41, further enhances MAML1-dependent histone acetylation in cells. Finally, MAML1 translocates GSK3beta to nuclear bodies; this function requires full-length MAML1 protein.

  17. Regulating expressin of cell and tissue-specific genes by modifying transcription

    Energy Technology Data Exchange (ETDEWEB)

    Beachy, R N; Dai, Shunhong

    2009-12-15

    Transcriptional regulation is the primary step to control gene expression, therefore function. Such regulation is achieved primarily via a combination of the activities of the promoter cis regulatory DNA elements and trans regulatory proteins that function through binding to these DNA elements. Our research supported by this program has led to the identification of rice bZIP transcription factors RF2a, RF2b and RLP1 that play key roles in regulating the activity of a vascular tissue specific promoter isolated from Rice Tungro Bacilliform Virus (RTBV) through their interactions with the Box II essential cis element located in the promoter. RF2a, RF2b and RLP1 possess multiple regulatory domains. Functional characterization reveals that those domains can activate or repress the activity of the RTBV promoter. Studies of transcriptional regulation of the RTBV promoter by this group of bZIP proteins not only provide insights about gene expression in the vascular tissue, but also insights about general mechanisms of transcription activation and repression. The knowledge gained from this research will also enable us to develop a well-described set of tools that can be used to control expression of multiple genes in transgenic plants and to improve biofuel feedstock.

  18. NLP is a novel transcription regulator involved in VSG expression site control in Trypanosoma brucei.

    Science.gov (United States)

    Narayanan, Mani Shankar; Kushwaha, Manish; Ersfeld, Klaus; Fullbrook, Alexander; Stanne, Tara M; Rudenko, Gloria

    2011-03-01

    Trypanosoma brucei mono-allelically expresses one of approximately 1500 variant surface glycoprotein (VSG) genes while multiplying in the mammalian bloodstream. The active VSG is transcribed by RNA polymerase I in one of approximately 15 telomeric VSG expression sites (ESs). T. brucei is unusual in controlling gene expression predominantly post-transcriptionally, and how ESs are mono-allelically controlled remains a mystery. Here we identify a novel transcription regulator, which resembles a nucleoplasmin-like protein (NLP) with an AT-hook motif. NLP is key for ES control in bloodstream form T. brucei, as NLP knockdown results in 45- to 65-fold derepression of the silent VSG221 ES. NLP is also involved in repression of transcription in the inactive VSG Basic Copy arrays, minichromosomes and procyclin loci. NLP is shown to be enriched on the 177- and 50-bp simple sequence repeats, the non-transcribed regions around rDNA and procyclin, and both active and silent ESs. Blocking NLP synthesis leads to downregulation of the active ES, indicating that NLP plays a role in regulating appropriate levels of transcription of ESs in both their active and silent state. Discovery of the unusual transcription regulator NLP provides new insight into the factors that are critical for ES control.

  19. Activator protein 1 promotes the transcriptional activation of IRAK-M.

    Science.gov (United States)

    Jin, Peipei; Bo, Lulong; Liu, Yongjian; Lu, Wenbin; Lin, Shengwei; Bian, Jinjun; Deng, Xiaoming

    2016-10-01

    Interleukin-1 receptor-associated kinase M (IRAK-M) is a well-known negative regulator for Toll-like receptor signaling, which can regulate immune homeostasis and tolerance in a number of pathological settings. However, the mechanism for IRAK-M regulation at transcriptional level remains largely unknown. In this study, a 1.4kb upstream sequence starting from the major IRAK-M transcriptional start site was cloned into luciferase reporter vector pGL3-basic to construct the full-length IRAK-M promoter. Luciferase reporter plasmids harboring the full-length and the deletion mutants of IRAK-M were transfected into 293T and A549 cells, and their relative luciferase activity was measured. The results demonstrated that activator protein 1(AP-1) cis-element plays a crucial role in IRAK-M constitutive gene transcription. Silencing of c-Fos and/or c-Jun expression suppressed the IRAK-M promoter activity as well as its mRNA and protein expressions. As a specific inhibitor for AP-1 activation, SP600125 also significantly suppressed the basal transcriptional activity of IRAK-M, the binding activity of c-Fos/c-Jun with IRAK-M promoter, and IRAK-M protein expression. Taken together, the result of this study highlights the importance of AP-1 in IRAK-M transcription, which offers more information on the role of IRAK-M in infectious and non-infectious diseases.

  20. Integration of the Transcription Factor-Regulated and Epigenetic Mechanisms in the Control of Keratinocyte Differentiation

    Science.gov (United States)

    Botchkarev, Vladimir A.

    2016-01-01

    The epidermal differentiation program is regulated at several levels including signaling pathways, lineage-specific transcription factors, and epigenetic regulators that establish well-coordinated process of terminal differentiation resulting in formation of the epidermal barrier. The epigenetic regulatory machinery operates at several levels including modulation of covalent DNA/histone modifications, as well as through higher-order chromatin remodeling to establish long-range topological interactions between the genes and their enhancer elements. Epigenetic regulators exhibit both activating and repressive effects on chromatin in keratinocytes (KCs): whereas some of them promote terminal differentiation, the others stimulate proliferation of progenitor cells, as well as inhibit premature activation of terminal differentiation-associated genes. Transcription factor-regulated and epigenetic mechanisms are highly connected, and the p63 transcription factor has an important role in the higher-order chromatin remodeling of the KC-specific gene loci via direct control of the genome organizer Satb1 and ATP-dependent chromatin remodeler Brg1. However, additional efforts are required to fully understand the complexity of interactions between distinct transcription factors and epigenetic regulators in the control of KC differentiation. Further understanding of these interactions and their alterations in different pathological skin conditions will help to progress toward the development of novel approaches for the treatment of skin disorders by targeting epigenetic regulators and modulating chromatin organization in KCs. PMID:26551942

  1. Enhancer Sharing Promotes Neighborhoods of Transcriptional Regulation Across Eukaryotes

    Science.gov (United States)

    Quintero-Cadena, Porfirio; Sternberg, Paul W.

    2016-01-01

    Enhancers physically interact with transcriptional promoters, looping over distances that can span multiple regulatory elements. Given that enhancer–promoter (EP) interactions generally occur via common protein complexes, it is unclear whether EP pairing is predominantly deterministic or proximity guided. Here, we present cross-organismic evidence suggesting that most EP pairs are compatible, largely determined by physical proximity rather than specific interactions. By reanalyzing transcriptome datasets, we find that the transcription of gene neighbors is correlated over distances that scale with genome size. We experimentally show that nonspecific EP interactions can explain such correlation, and that EP distance acts as a scaling factor for the transcriptional influence of an enhancer. We propose that enhancer sharing is commonplace among eukaryotes, and that EP distance is an important layer of information in gene regulation. PMID:27799341

  2. Evolution of proline biosynthesis: enzymology, bioinformatics, genetics, and transcriptional regulation.

    Science.gov (United States)

    Fichman, Yosef; Gerdes, Svetlana Y; Kovács, Hajnalka; Szabados, László; Zilberstein, Aviah; Csonka, Laszlo N

    2015-11-01

    Proline is not only an essential component of proteins but it also has important roles in adaptation to osmotic and dehydration stresses, redox control, and apoptosis. Here, we review pathways of proline biosynthesis in the three domains of life. Pathway reconstruction from genome data for hundreds of eubacterial and dozens of archaeal and eukaryotic organisms revealed evolutionary conservation and variations of this pathway across different taxa. In the most prevalent pathway of proline synthesis, glutamate is phosphorylated to γ-glutamyl phosphate by γ-glutamyl kinase, reduced to γ-glutamyl semialdehyde by γ-glutamyl phosphate reductase, cyclized spontaneously to Δ(1)-pyrroline-5-carboxylate and reduced to proline by Δ(1)-pyrroline-5-carboxylate reductase. In higher plants and animals the first two steps are catalysed by a bi-functional Δ(1) -pyrroline-5-carboxylate synthase. Alternative pathways of proline formation use the initial steps of the arginine biosynthetic pathway to ornithine, which can be converted to Δ(1)-pyrroline-5-carboxylate by ornithine aminotransferase and then reduced to proline or converted directly to proline by ornithine cyclodeaminase. In some organisms, the latter pathways contribute to or could be fully responsible for the synthesis of proline. The conservation of proline biosynthetic enzymes and significance of specific residues for catalytic activity and allosteric regulation are analysed on the basis of protein structural data, multiple sequence alignments, and mutant studies, providing novel insights into proline biosynthesis in organisms. We also discuss the transcriptional control of the proline biosynthetic genes in bacteria and plants.

  3. Transcriptional regulation of cadherins during development and carcinogenesis.

    Science.gov (United States)

    Peinado, Héctor; Portillo, Francisco; Cano, Amparo

    2004-01-01

    The cadherin superfamily of Ca(2+)-dependent homophilic adhesion molecules plays a critical role in regulating cell-to-cell interactions. During development, the expression of different cadherins is highly dynamic, since they are associated with the morphogenesis, establishment and/or maintenance of different tissues. Alterations in cadherin expression or function occur frequently during carcinogenesis, such as the loss of the epithelial cadherin (E-cadherin) and/or the aberrant expression of other cadherins. Indeed, the aberrant expression of cadherins has been detected during carcinoma invasion, a process which is reminiscent of the epithelial-mesenchymal transition (EMT) so important in many critical developmental processes. The functional regulation of cadherins can occur at many different levels, from transcriptional regulation to the control of the strength of the cadherin-mediated cell-cell interaction. In this review, we will focus on the transcriptional control of cadherin expression, both in development and carcinogenesis, paying particular attention to the regulation of E-cadherin given its proposed role as a suppressor of invasion. We will discuss the main genetic and epigenetic mechanisms involved in down-regulating E-cadherin expression, and we will analyse the mechanisms involved in regulating EMT, in an attempt to elucidate which elements are common to this process in both physiological and pathological situations.

  4. Evolution of transcriptional regulation in closely related bacteria

    Directory of Open Access Journals (Sweden)

    Tsoy Olga V

    2012-10-01

    Full Text Available Abstract Background The exponential growth of the number of fully sequenced genomes at varying taxonomic closeness allows one to characterize transcriptional regulation using comparative-genomics analysis instead of time-consuming experimental methods. A transcriptional regulatory unit consists of a transcription factor, its binding site and a regulated gene. These units constitute a graph which contains so-called “network motifs”, subgraphs of a given structure. Here we consider genomes of closely related Enterobacteriales and estimate the fraction of conserved network motifs and sites as well as positions under selection in various types of non-coding regions. Results Using a newly developed technique, we found that the highest fraction of positions under selection, approximately 50%, was observed in synvergon spacers (between consecutive genes from the same strand, followed by ~45% in divergon spacers (common 5’-regions, and ~10% in convergon spacers (common 3’-regions. The fraction of selected positions in functional regions was higher, 60% in transcription factor-binding sites and ~45% in terminators and promoters. Small, but significant differences were observed between Escherichia coli and Salmonella enterica. This fraction is similar to the one observed in eukaryotes. The conservation of binding sites demonstrated some differences between types of regulatory units. In E. coli, strains the interactions of the type “local transcriptional factor gene” turned out to be more conserved in feed-forward loops (FFLs compared to non-motif interactions. The coherent FFLs tend to be less conserved than the incoherent FFLs. A natural explanation is that the former imply functional redundancy. Conclusions A naïve hypothesis that FFL would be highly conserved turned out to be not entirely true: its conservation depends on its status in the transcriptional network and also from its usage. The fraction of positions under selection in

  5. Functional analysis of transcriptional regulation of herpes simplex virus type 1 tegument protein VP22

    Institute of Scientific and Technical Information of China (English)

    YU Xian; LI WeiZhong; LIU LongDing; CHE YanChun; CUN Wei; WU Wen Juan; HE ChunYan; SHAO CongWen; LI QiHan

    2008-01-01

    The herpes simplex virus type 1 (HSV-1) tegument proteins have important functions in the viral repli-cation process. In order to investigate the role of the HSV-1 tegument protein VP22 in viral replication, its transcriptional regulation of viral promoters was investigated using the chloramphenicol acetyl-transferase (CAT) assay. The results indicate that VP22 exerts a dose-dependent transcriptional in-hibitory effect on the HSV-1 α4, TK, and gC gene promoters. VP22 had the capacity to repress tran-scriptional activation of promoters via different viral transcription regulatory factors such as VP16 and ICP0, as evidenced by the specific repression of the TK and gC gene promoters by ICP0. In addition, VP22 was capable of inhibiting the promotion of ICP0 transcriptional activation in the presence of HAT PCAF, which is even more remarkable than the VP22 repression of ICP0 transcriptional activation. Fi-nally, the transcriptional inhibitory effect of VP22 on other viral promoters was demonstrated by the analysis of β-galactosidase activities in internal controls.

  6. Transcriptional profiling of UlaR-regulated genes in Streptococcus pneumoniae

    NARCIS (Netherlands)

    Shafeeq, Sulman; Afzal, Muhammad; Henriques-Normark, Birgitta; Kuipers, Oscar P

    2015-01-01

    The transcriptional regulator UlaR belongs to the family of PRD-containing transcriptional regulators, which are mostly involved in the regulation of carbohydrate metabolism. The role of the transcriptional regulator UlaR in Streptococcus pneumoniae has recently been described [1]. Here, we report d

  7. TRANSFAC and its module TRANSCompel: transcriptional gene regulation in eukaryotes.

    Science.gov (United States)

    Matys, V; Kel-Margoulis, O V; Fricke, E; Liebich, I; Land, S; Barre-Dirrie, A; Reuter, I; Chekmenev, D; Krull, M; Hornischer, K; Voss, N; Stegmaier, P; Lewicki-Potapov, B; Saxel, H; Kel, A E; Wingender, E

    2006-01-01

    The TRANSFAC database on transcription factors, their binding sites, nucleotide distribution matrices and regulated genes as well as the complementing database TRANSCompel on composite elements have been further enhanced on various levels. A new web interface with different search options and integrated versions of Match and Patch provides increased functionality for TRANSFAC. The list of databases which are linked to the common GENE table of TRANSFAC and TRANSCompel has been extended by: Ensembl, UniGene, EntrezGene, HumanPSD and TRANSPRO. Standard gene names from HGNC, MGI and RGD, are included for human, mouse and rat genes, respectively. With the help of InterProScan, Pfam, SMART and PROSITE domains are assigned automatically to the protein sequences of the transcription factors. TRANSCompel contains now, in addition to the COMPEL table, a separate table for detailed information on the experimental EVIDENCE on which the composite elements are based. Finally, for TRANSFAC, in respect of data growth, in particular the gain of Drosophila transcription factor binding sites (by courtesy of the Drosophila DNase I footprint database) and of Arabidopsis factors (by courtesy of DATF, Database of Arabidopsis Transcription Factors) has to be stressed. The here described public releases, TRANSFAC 7.0 and TRANSCompel 7.0, are accessible under http://www.gene-regulation.com/pub/databases.html.

  8. Temperature regulates transcription in the zebrafish circadian clock.

    Directory of Open Access Journals (Sweden)

    Kajori Lahiri

    2005-11-01

    Full Text Available It has been well-documented that temperature influences key aspects of the circadian clock. Temperature cycles entrain the clock, while the period length of the circadian cycle is adjusted so that it remains relatively constant over a wide range of temperatures (temperature compensation. In vertebrates, the molecular basis of these properties is poorly understood. Here, using the zebrafish as an ectothermic model, we demonstrate first that in the absence of light, exposure of embryos and primary cell lines to temperature cycles entrains circadian rhythms of clock gene expression. Temperature steps drive changes in the basal expression of certain clock genes in a gene-specific manner, a mechanism potentially contributing to entrainment. In the case of the per4 gene, while E-box promoter elements mediate circadian clock regulation, they do not direct the temperature-driven changes in transcription. Second, by studying E-box-regulated transcription as a reporter of the core clock mechanism, we reveal that the zebrafish clock is temperature-compensated. In addition, temperature strongly influences the amplitude of circadian transcriptional rhythms during and following entrainment by light-dark cycles, a property that could confer t