WorldWideScience

Sample records for activating transcription factor

  1. Human cytomegalovirus IE2 protein interacts with transcription activating factors

    Institute of Scientific and Technical Information of China (English)

    XU; Jinping(徐进平); YE; Linbai(叶林柏)

    2002-01-01

    The human cytomegalovirus (HCMV) IE86 Cdna was cloned into Pgex-2T and fusion protein GST-IE86 was expressed in E. Coli. SDS-PAGE and Western blot assay indicated that fusion protein GST-IE86 with molecular weight of 92 ku is soluble in the supernatant of cell lysate. Protein GST and fusion protein GST-IE86 were purified by affinity chromatography. The technology of co-separation and specific affinity chromatography was used to study the interactions of HCMV IE86 protein with some transcriptional regulatory proteins and transcriptional factors. The results indicated that IE86 interacts separately with transcriptional factor TFIIB and promoter DNA binding transcription trans-activating factors SP1, AP1 and AP2 to form a heterogenous protein complex. These transcriptional trans-activating factors, transcriptional factor and IE86 protein were adsorbed and retained in the affinity chromatography simultaneously. But IE86 protein could not interact with NF-Кb, suggesting that the function of IE86 protein that can interact with transcriptional factor and transcriptional trans-activating factors has no relevance to protein glycosylation. IE86 protein probably has two domains responsible for binding transcriptional trans-activating regulatory proteins and transcriptional factors respectively, thus activating the transcription of many genes. The interactions accelerated the assembly of the transcriptional initiation complexes.

  2. 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.

  3. 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

  4. 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.

  5. 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

    2010-01-01

    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

  6. 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.

  7. 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.

  8. 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.

  9. 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

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

    Science.gov (United States)

    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.

  11. Activating transcription factor 6 derepression mediates neuroprotection in Huntington disease.

    Science.gov (United States)

    Naranjo, José R; Zhang, Hongyu; Villar, Diego; González, Paz; Dopazo, Xose M; Morón-Oset, Javier; Higueras, Elena; Oliveros, Juan C; Arrabal, María D; Prieto, Angela; Cercós, Pilar; González, Teresa; De la Cruz, Alicia; Casado-Vela, Juan; Rábano, Alberto; Valenzuela, Carmen; Gutierrez-Rodriguez, Marta; Li, Jia-Yi; Mellström, Britt

    2016-02-01

    Deregulated protein and Ca2+ homeostasis underlie synaptic dysfunction and neurodegeneration in Huntington disease (HD); however, the factors that disrupt homeostasis are not fully understood. Here, we determined that expression of downstream regulatory element antagonist modulator (DREAM), a multifunctional Ca2+-binding protein, is reduced in murine in vivo and in vitro HD models and in HD patients. DREAM downregulation was observed early after birth and was associated with endogenous neuroprotection. In the R6/2 mouse HD model, induced DREAM haplodeficiency or blockade of DREAM activity by chronic administration of the drug repaglinide delayed onset of motor dysfunction, reduced striatal atrophy, and prolonged life span. DREAM-related neuroprotection was linked to an interaction between DREAM and the unfolded protein response (UPR) sensor activating transcription factor 6 (ATF6). Repaglinide blocked this interaction and enhanced ATF6 processing and nuclear accumulation of transcriptionally active ATF6, improving prosurvival UPR function in striatal neurons. Together, our results identify a role for DREAM silencing in the activation of ATF6 signaling, which promotes early neuroprotection in HD.

  12. Comprehensive Behavioral Analysis of Activating Transcription Factor 5-Deficient Mice.

    Science.gov (United States)

    Umemura, Mariko; Ogura, Tae; Matsuzaki, Ayako; Nakano, Haruo; Takao, Keizo; Miyakawa, Tsuyoshi; Takahashi, Yuji

    2017-01-01

    Activating transcription factor 5 (ATF5) is a member of the CREB/ATF family of basic leucine zipper transcription factors. We previously reported that ATF5-deficient (ATF5(-/-)) mice demonstrated abnormal olfactory bulb development due to impaired interneuron supply. Furthermore, ATF5(-/-) mice were less aggressive than ATF5(+/+) mice. Although ATF5 is widely expressed in the brain, and involved in the regulation of proliferation and development of neurons, the physiological role of ATF5 in the higher brain remains unknown. Our objective was to investigate the physiological role of ATF5 in the higher brain. We performed a comprehensive behavioral analysis using ATF5(-/-) mice and wild type littermates. ATF5(-/-) mice exhibited abnormal locomotor activity in the open field test. They also exhibited abnormal anxiety-like behavior in the light/dark transition test and open field test. Furthermore, ATF5(-/-) mice displayed reduced social interaction in the Crawley's social interaction test and increased pain sensitivity in the hot plate test compared with wild type. Finally, behavioral flexibility was reduced in the T-maze test in ATF5(-/-) mice compared with wild type. In addition, we demonstrated that ATF5(-/-) mice display disturbances of monoamine neurotransmitter levels in several brain regions. These results indicate that ATF5 deficiency elicits abnormal behaviors and the disturbance of monoamine neurotransmitter levels in the brain. The behavioral abnormalities of ATF5(-/-) mice may be due to the disturbance of monoamine levels. Taken together, these findings suggest that ATF5(-/-) mice may be a unique animal model of some psychiatric disorders.

  13. 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.

  14. Transcription factor PIF4 controls the thermosensory activation of flowering

    KAUST Repository

    Kumar, S. Vinod

    2012-03-21

    Plant growth and development are strongly affected by small differences in temperature. Current climate change has already altered global plant phenology and distribution, and projected increases in temperature pose a significant challenge to agriculture. Despite the important role of temperature on plant development, the underlying pathways are unknown. It has previously been shown that thermal acceleration of flowering is dependent on the florigen, FLOWERING LOCUS T (FT). How this occurs is, however, not understood, because the major pathway known to upregulate FT, the photoperiod pathway, is not required for thermal acceleration of flowering. Here we demonstrate a direct mechanism by which increasing temperature causes the bHLH transcription factor PHYTOCHROME INTERACTING FACTOR4 (PIF4) to activate FT. Our findings provide a new understanding of how plants control their timing of reproduction in response to temperature. Flowering time is an important trait in crops as well as affecting the life cycles of pollinator species. A molecular understanding of how temperature affects flowering will be important for mitigating the effects of climate change. © 2012 Macmillan Publishers Limited. All rights reserved.

  15. Peroxisome proliferator-activated receptor gamma recruits the positive transcription elongation factor b complex to activate transcription and promote adipogenesis

    DEFF Research Database (Denmark)

    Iankova, Irena; Petersen, Rasmus K; Annicotte, Jean-Sébastien

    2006-01-01

    Positive transcription elongation factor b (P-TEFb) phosphorylates the C-terminal domain of RNA polymerase II, facilitating transcriptional elongation. In addition to its participation in general transcription, P-TEFb is recruited to specific promoters by some transcription factors such as c-Myc...

  16. Identification of Post-Transcriptional Modulators of Breast Cancer Transcription Factor Activity Using MINDy

    Science.gov (United States)

    Campbell, Thomas M.; Castro, Mauro A. A.; Ponder, Bruce A. J.

    2016-01-01

    We have recently identified transcription factors (TFs) that are key drivers of breast cancer risk. To better understand the pathways or sub-networks in which these TFs mediate their function we sought to identify upstream modulators of their activity. We applied the MINDy (Modulator Inference by Network Dynamics) algorithm to four TFs (ESR1, FOXA1, GATA3 and SPDEF) that are key drivers of estrogen receptor-positive (ER+) breast cancer risk, as well as cancer progression. Our computational analysis identified over 500 potential modulators. We assayed 189 of these and identified 55 genes with functional characteristics that were consistent with a role as TF modulators. In the future, the identified modulators may be tested as potential therapeutic targets, able to alter the activity of TFs that are critical in the development of breast cancer. PMID:27997592

  17. Building gene expression signatures indicative of transcription factor activation to predict AOP modulation

    Science.gov (United States)

    Building gene expression signatures indicative of transcription factor activation to predict AOP modulation Adverse outcome pathways (AOPs) are a framework for predicting quantitative relationships between molecular initiatin...

  18. Glucagon and Insulin Cooperatively Stimulate Fibroblast Growth Factor 21 Gene Transcription by Increasing the Expression of Activating Transcription Factor 4.

    Science.gov (United States)

    Alonge, Kimberly M; Meares, Gordon P; Hillgartner, F Bradley

    2017-03-31

    Previous studies have shown that glucagon cooperatively interacts with insulin to stimulate hepatic FGF21 gene expression. Here we investigated the mechanism by which glucagon and insulin increased FGF21 gene transcription in primary hepatocyte cultures. Transfection analyses demonstrated that glucagon plus insulin induction of FGF21 transcription was conferred by two activating transcription factor 4 (ATF4) binding sites in the FGF21 gene. Glucagon plus insulin stimulated a 5-fold increase in ATF4 protein abundance, and knockdown of ATF4 expression suppressed the ability of glucagon plus insulin to increase FGF21 expression. In hepatocytes incubated in the presence of insulin, treatment with a PKA-selective agonist mimicked the ability of glucagon to stimulate ATF4 and FGF21 expression. Inhibition of PKA, PI3K, Akt, and mammalian target of rapamycin complex 1 (mTORC1) suppressed the ability of glucagon plus insulin to stimulate ATF4 and FGF21 expression. Additional analyses demonstrated that chenodeoxycholic acid (CDCA) induced a 6-fold increase in ATF4 expression and that knockdown of ATF4 expression suppressed the ability of CDCA to increase FGF21 gene expression. CDCA increased the phosphorylation of eIF2α, and inhibition of eIF2α signaling activity suppressed CDCA regulation of ATF4 and FGF21 expression. These results demonstrate that glucagon plus insulin increases FGF21 transcription by stimulating ATF4 expression and that activation of cAMP/PKA and PI3K/Akt/mTORC1 mediates the effect of glucagon plus insulin on ATF4 expression. These results also demonstrate that CDCA regulation of FGF21 transcription is mediated at least partially by an eIF2α-dependent increase in ATF4 expression. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. Activating Transcription Factor 3 (ATF3 and the Nervous System

    Directory of Open Access Journals (Sweden)

    Patrick Norval Anderson

    2012-02-01

    Full Text Available It has been recognised for over a century that the ability of axons to regenerate in peripheral nerves is fundamentally greater than that of axons in the brain, spinal cord or optic nerves [early literature was reviewed in (Ramon y Cajal, 1928]. One factor that contributes to the successful regeneration of the axons in peripheral nerves is the complex cell body response the neurons show to axotomy. That transcription factors must play an important role in enabling neurons to regrow their axons is implicit to the observation that several hundred genes are regulated in neurons during axonal regeneration (Costigan et al., 2002; Boeshore et al., 2004. In addition, similarly large numbers of genes are regulated in the non-neuronal cells present in injured peripheral nerves [especially Schwann cells (Barrette et al., 2010] and CNS tissue. Of the transcription factors that regulate these changes in gene expression, the function of c-jun is best understood but ATF-3 (also known as LRF-1, LRG-21, CRG-5 and TI-241 is also upregulated in most of the neurons (Fig. 1 and Schwann cells that express c-jun. Indeed, ATF-3 has become a standard marker for neurons axotomised by peripheral nerve injury (Tsuzuki et al., 2001; Yamanaka et al., 2005; Yano et al., 2008; Linda et al., 2011 and its expression by injured neurons is closely correlated with a regenerative response. None the less, surprisingly little is known about the functions of ATF3 in neurons or glia within the injured nervous system, especially when compared with those of its potential binding partner, c-Jun.

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

    Directory of Open Access Journals (Sweden)

    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.

  1. 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.

  2. Characterization of the transcriptional activation domains of human TEF3-1 (transcription enhancer factor 3 isoform 1).

    Science.gov (United States)

    Qiao, Cheng; Jiang, Yajie; Deng, Cuilan; Huang, Zebo; Teng, Kaixuan; Chen, Lan; Liu, Xin

    2015-03-01

    TEF3-1 (transcription enhancer factor 3 isoform 1) is a human transcriptional factor, which has a N-terminal TEA/ATTS domain supposedly for DNA binding and C-terminal PRD and STY domains for transcriptional activation. Taking advantage of the efficient reporter design of yeast two-hybrid system, we characterized the TEF3-1 domains in activating gene expression. Previously study usually mentioned that the C-terminal domain of TEF3-1 has the transcriptional activity, however, our data shows that the peptides TEF3-11-66 and TEF3-1197-434 functioned as two independent activation domains, suggesting that N-terminal domain of TEF3-1 also has transcriptional activation capacity. Additionally, more deletions of amino acids 197-434 showed that only the peptides TEF3-1197-265 contained the minimum sequences for the C-terminal transcriptional activation domain. The protein structure is predicted to contain a helix-turn-helix structure in TEF3-11-66 and four β sheets in TEF3-1197-265. Finally, after the truncated fragments of TEF3-1 were expressed in HUVEC cells, the whole TEF3-1 and the two activation domains could increase F-actin stress fiber, cell proliferation, migration and targeted gene expression. Further analysis and characterization of the activation domains in TEF3-1 may broaden our understanding of the gene involved in angiogenesis and other pathological processes.

  3. WRKY Transcription Factors Involved in Activation of SA Biosynthesis Genes

    Directory of Open Access Journals (Sweden)

    Bol John F

    2011-05-01

    Full Text Available Abstract Background Increased defense against a variety of pathogens in plants is achieved through activation of a mechanism known as systemic acquired resistance (SAR. The broad-spectrum resistance brought about by SAR is mediated through salicylic acid (SA. An important step in SA biosynthesis in Arabidopsis is the conversion of chorismate to isochorismate through the action of isochorismate synthase, encoded by the ICS1 gene. Also AVRPPHB SUSCEPTIBLE 3 (PBS3 plays an important role in SA metabolism, as pbs3 mutants accumulate drastically reduced levels of SA-glucoside, a putative storage form of SA. Bioinformatics analysis previously performed by us identified WRKY28 and WRKY46 as possible regulators of ICS1 and PBS3. Results Expression studies with ICS1 promoter::β-glucuronidase (GUS genes in Arabidopsis thaliana protoplasts cotransfected with 35S::WRKY28 showed that over expression of WRKY28 resulted in a strong increase in GUS expression. Moreover, qRT-PCR analyses indicated that the endogenous ICS1 and PBS3 genes were highly expressed in protoplasts overexpressing WRKY28 or WRKY46, respectively. Electrophoretic mobility shift assays indentified potential WRKY28 binding sites in the ICS1 promoter, positioned -445 and -460 base pairs upstream of the transcription start site. Mutation of these sites in protoplast transactivation assays showed that these binding sites are functionally important for activation of the ICS1 promoter. Chromatin immunoprecipitation assays with haemagglutinin-epitope-tagged WRKY28 showed that the region of the ICS1 promoter containing the binding sites at -445 and -460 was highly enriched in the immunoprecipitated DNA. Conclusions The results obtained here confirm results from our multiple microarray co-expression analyses indicating that WRKY28 and WRKY46 are transcriptional activators of ICS1 and PBS3, respectively, and support this in silico screening as a powerful tool for identifying new components of stress

  4. Altered activities of transcription factors and their related gene expression in cardiac tissues of diabetic rats.

    Science.gov (United States)

    Nishio, Y; Kashiwagi, A; Taki, H; Shinozaki, K; Maeno, Y; Kojima, H; Maegawa, H; Haneda, M; Hidaka, H; Yasuda, H; Horiike, K; Kikkawa, R

    1998-08-01

    Gene regulation in the cardiovascular tissues of diabetic subjects has been reported to be altered. To examine abnormal activities in transcription factors as a possible cause of this altered gene regulation, we studied the activity of two redox-sensitive transcription factors--nuclear factor-kappaB (NF-kappaB) and activating protein-1 (AP-1)--and the change in the mRNA content of heme oxygenase-1, which is regulated by these transcription factors in the cardiac tissues of rats with streptozotocin-induced diabetes. Increased activity of NF-kappaB and AP-1 but not nuclear transcription-activating factor, as determined by an electrophoretic mobility shift assay, was found in the hearts of 4-week diabetic rats. Glycemic control by a subcutaneous injection of insulin prevented these diabetes-induced changes in transcription factor activity. In accordance with these changes, the mRNA content of heme oxygenase-1 was increased fourfold in 4-week diabetic rats and threefold in 24-week diabetic rats as compared with control rats (P oxidative stress is involved in the activation of the transcription factors NF-kappaB and AP-1 in the cardiac tissues of diabetic rats, and that these abnormal activities of transcription factors could be associated with the altered gene regulation observed in the cardiovascular tissues of diabetic rats.

  5. Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression.

    Science.gov (United States)

    Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression Exposure to many drugs and environmentally-relevant chemicals can cause adverse outcomes. These adverse outcomes, such as cancer, have been linked to mol...

  6. A New Microsphere-Based Immunoassay for Measuring the Activity of Transcription Factors

    Directory of Open Access Journals (Sweden)

    Tsai Chueh-Jen

    2010-01-01

    Full Text Available Abstract There are several traditional and well-developed methods for analyzing the activity of transcription factors, such as EMSA, enzyme-linked immunosorbent assay, and reporter gene activity assays. All of these methods have their own distinct disadvantages, but none can analyze the changes in transcription factors in the few cells that are cultured in the wells of 96-well titer plates. Thus, a new microsphere-based immunoassay to measure the activity of transcription factors (MIA-TF was developed. In MIA-TF, NeutrAvidin-labeled microspheres were used as the solid phase to capture biotin-labeled double-strand DNA fragments which contain certain transcription factor binding elements. The activity of transcription factors was detected by immunoassay using a transcription factor-specific antibody to monitor the binding with the DNA probe. Next, analysis was performed by flow cytometry. The targets hypoxia-inducible factor-1α (HIF-1α and nuclear factor-kappa B (NF-κB were applied and detected in this MIA-TF method; the results that we obtained demonstrated that this method could be used to monitor the changes of NF-κB or HIF within 50 or 100 ng of nuclear extract. Furthermore, MIA-TF could detect the changes in NF-κB or HIF in cells that were cultured in wells of a 96-well plate without purification of the nuclear protein, an important consideration for applying this method to high-throughput assays in the future. The development of MIA-TF would support further progress in clinical analysis and drug screening systems. Overall, MIA-TF is a method with high potential to detect the activity of transcription factors.

  7. Inhibition of the TEF/TEAD transcription factor activity by nuclear calcium and distinct kinase pathways.

    Science.gov (United States)

    Thompson, M; Andrade, V A; Andrade, S J; Pusl, T; Ortega, J M; Goes, A M; Leite, M F

    2003-02-07

    Transcription enhancer factor (TEF/TEAD) is a family of four transcription factors that share a common TEA-DNA binding domain and are involved in similar cellular functions, such as cell differentiation and proliferation. All adult tissues express at least one of the four TEAD genes, so this family of transcription factors may be of widespread importance, yet little is known about their regulation. Here we examine the factors that regulate TEAD activity in CHO cells. RT-PCR indicated the presence of TEAD-1, TEAD-3, and both isoforms of TEAD-4, but not TEAD-2. Quantitative measurements showed that TEAD-4 is most abundant, followed by TEAD-3, then TEAD-1. We examined the relative effects of nuclear and cytosolic Ca(2+) on TEAD activity, since TEAD proteins are localized to the nucleus and since free Ca(2+) within the nucleus selectively regulates transcription in some systems. Chelation of nuclear but not cytosolic Ca(2+) increased TEAD activity two times above control. Inhibition of mitogen-activated protein kinase (MAPK) also increased TEAD activity, while cAMP decreased TEAD activity, and protein kinase C had no effect. Together, these results show that nuclear Ca(2+), MAPK, and cAMP each negatively regulate the activity of the TEAD transcription factor.

  8. Arabidopsis sigma factor binding proteins are activators of the WRKY33 transcription factor in plant defense.

    Science.gov (United States)

    Lai, Zhibing; Li, Ying; Wang, Fei; Cheng, Yuan; Fan, Baofang; Yu, Jing-Quan; Chen, Zhixiang

    2011-10-01

    Necrotrophic pathogens are important plant pathogens that cause many devastating plant diseases. Despite their impact, our understanding of the plant defense response to necrotrophic pathogens is limited. The WRKY33 transcription factor is important for plant resistance to necrotrophic pathogens; therefore, elucidation of its functions will enhance our understanding of plant immunity to necrotrophic pathogens. Here, we report the identification of two WRKY33-interacting proteins, nuclear-encoded SIGMA FACTOR BINDING PROTEIN1 (SIB1) and SIB2, which also interact with plastid-encoded plastid RNA polymerase SIGMA FACTOR1. Both SIB1 and SIB2 contain an N-terminal chloroplast targeting signal and a putative nuclear localization signal, suggesting that they are dual targeted. Bimolecular fluorescence complementation indicates that WRKY33 interacts with SIBs in the nucleus of plant cells. Both SIB1 and SIB2 contain a short VQ motif that is important for interaction with WRKY33. The two VQ motif-containing proteins recognize the C-terminal WRKY domain and stimulate the DNA binding activity of WRKY33. Like WRKY33, both SIB1 and SIB2 are rapidly and strongly induced by the necrotrophic pathogen Botrytis cinerea. Resistance to B. cinerea is compromised in the sib1 and sib2 mutants but enhanced in SIB1-overexpressing transgenic plants. These results suggest that dual-targeted SIB1 and SIB2 function as activators of WRKY33 in plant defense against necrotrophic pathogens.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Science.gov (United States)

    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.

  11. Expression and Activation of STAT Transcription Factors in Breast Cancer

    Science.gov (United States)

    1998-05-08

    clinicians. J~, 273: 577-585, 1995. 183 Hundertmark 5, Buhler H, Rudolf M, Weitzel HK, Ragosch V: Inhibition of 11 beta-hydroxysteroid dehydrogenase...activated protein kinase through a Jakl-dependent pathway. Mol. Cell. Bioi., 17:3833-40, 1997. Stewart JF, Rubens RO, King RJ, Minton MJ, Steiner R

  12. Smad transcription factors.

    Science.gov (United States)

    Massagué, Joan; Seoane, Joan; Wotton, David

    2005-12-01

    Smad transcription factors lie at the core of one of the most versatile cytokine signaling pathways in metazoan biology-the transforming growth factor-beta (TGFbeta) pathway. Recent progress has shed light into the processes of Smad activation and deactivation, nucleocytoplasmic dynamics, and assembly of transcriptional complexes. A rich repertoire of regulatory devices exerts control over each step of the Smad pathway. This knowledge is enabling work on more complex questions about the organization, integration, and modulation of Smad-dependent transcriptional programs. We are beginning to uncover self-enabled gene response cascades, graded Smad response mechanisms, and Smad-dependent synexpression groups. Our growing understanding of TGFbeta signaling through the Smad pathway provides general principles for how animal cells translate complex inputs into concrete behavior.

  13. Identification of HTF (HER2 transcription factor) as an AP-2 (activator protein-2) transcription factor and contribution of the HTF binding site to ERBB2 gene overexpression.

    Science.gov (United States)

    Vernimmen, Douglas; Begon, Dominique; Salvador, Christophe; Gofflot, Stéphanie; Grooteclaes, Madeleine; Winkler, Rosita

    2003-02-15

    The ERBB2 gene is overexpressed in 30% of human breast cancers and this is correlated with poor prognosis. Overexpression of the ERBB2 gene is due to increased transcription and gene amplification. Our previous studies have identified a new cis element in the ERBB2 promoter which is involved in the gene's overexpression. This cis element, located 501 bp upstream from the main ERBB2 transcription initiation site, binds a transcription factor called HTF (HER2 transcription factor). We report here the identification of HTF as an AP-2 (activator protein-2) transcription factor. The new cis element is bound by AP-2 with high affinity, compared with a previously described AP-2 binding site located 284 bp downstream. Co-transfection of an AP-2alpha expression vector with a reporter vector containing the newly identified AP-2 binding site in front of a minimal ERBB2 promoter induced a dose-dependent increase in transcriptional activity. We examined the contribution of the new AP-2 binding site to ERBB2 overexpression. For this purpose we abolished the new and/or the previously described AP-2 binding sequence by site-directed mutagenesis. The results show that the two functional AP-2 sites in the first 700 bp of the ERBB2 promoter co-operate to achieve maximal transcriptional activity.

  14. PEA3/ETV4-related transcription factors coupled with active ERK signalling are associated with poor prognosis in gastric adenocarcinoma

    LENUS (Irish Health Repository)

    Keld, R

    2011-06-28

    Background: Transcription factors often play important roles in tumourigenesis. Members of the PEA3 subfamily of ETS-domain transcription factors fulfil such a role and have been associated with tumour metastasis in several different cancers. Moreover, the activity of the PEA3 subfamily transcription factors is potentiated by Ras-ERK pathway signalling, which is itself often deregulated in tumour cells.\\r\

  15. 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.

  16. Role of SIRT1 and FOXO factors in eNOS transcriptional activation by resveratrol.

    Science.gov (United States)

    Xia, Ning; Strand, Susanne; Schlufter, Frank; Siuda, Daniel; Reifenberg, Gisela; Kleinert, Hartmut; Förstermann, Ulrich; Li, Huige

    2013-08-01

    Many of the cardiovascular protective effects of resveratrol are attributable to an enhanced production of nitric oxide (NO) by the endothelial NO synthase (eNOS). Resveratrol has been shown to enhance eNOS gene expression as well as eNOS enzymatic activity. The aim of the present study was to analyze the molecular mechanisms of eNOS transcriptional activation by resveratrol. Treatment of human EA.hy 926 endothelial cells with resveratrol led to a concentration-dependent upregulation of eNOS expression. In luciferase reporter gene assay, resveratrol enhanced the activity of human eNOS promoter fragments (3500, 1600, 633 and 263bp in length, respectively), indicating that the proximal promoter region is required for resveratrol-induced eNOS transcriptional activation. Knockdown of the NAD(+)-dependent protein deacetylase sirtuin 1 (SIRT1) by siRNA prevented the upregulation of eNOS mRNA and protein by resveratrol. Forkhead box O (FOXO) transcription factors are established downstream targets of SIRT1. siRNA-mediated knockdown of FOXO1 and FOXO3a abolished the effect of resveratrol on eNOS expression, indicating the involvement of these factors. Resveratrol treatment enhanced the expression of FOXO1 and FOXO3a in EA.hy 926 cells. Reporter gene assay using promoter containing forkhead response elements showed increased FOXO factor activity by resveratrol. In electrophoretic mobility shift assay, the enhanced binding of nuclear proteins to the eNOS promoter regions by resveratrol could be blocked by antibodies against FOXO1 and FOXO3a. In conclusion, resveratrol enhances the expression and activity of FOXO transcription factors. The SIRT1/FOXO factor axis is involved in resveratrol-induced eNOS transcriptional activation.

  17. Elk3 from hamster--a ternary complex factor with strong transcriptional repressor activity.

    Science.gov (United States)

    Hjortoe, Gertrud Malene; Weilguny, Dietmar; Willumsen, Berthe Marie

    2005-01-01

    Elk3 belongs to the Ets family of transcription factors, which are regulated by the Ras/mitogen-activated protein kinase-signaling pathway. In the absence of Ras, this protein is a strong inhibitor of transcription and may be directly involved in regulation of growth by downregulating the transcription of genes that are activated during entry into G1. We have isolated the Cricetulus griseus Elk3 gene from the Chinese hamster ovary (CHO) cell line and investigated the transcriptional potential of this factor. Transient transfections revealed that, in addition to its regulation of the c-fos promoter, Elk3 from CHO cells seems to inhibit other promoters controlling expression of proteins involved in G1/S phase progression; Cyclin D1 and DHFR. As has been described for the Elk3 homologs Net (Mouse) and Sap-2 (Human), the results of the present study further indicate that hamster Elk3 is a target of the Ras-Raf-MAPK pathway, and cotransfections with constitutively active H-ras relieves its negative transcriptional activity. No cells stably expressing exogenous Elk3 could be obtained, possibly due to an unspecified toxic or growth retarding effect. These findings support a possible role for Elk3 in growth regulation and reveal a high degree of homology for this protein across species.

  18. MiT/TFE transcription factors are activated during mitophagy downstream of Parkin and Atg5.

    Science.gov (United States)

    Nezich, Catherine L; Wang, Chunxin; Fogel, Adam I; Youle, Richard J

    2015-08-03

    The kinase PINK1 and ubiquitin ligase Parkin can regulate the selective elimination of damaged mitochondria through autophagy (mitophagy). Because of the demand on lysosomal function by mitophagy, we investigated a role for the transcription factor EB (TFEB), a master regulator of lysosomal biogenesis, in this process. We show that during mitophagy TFEB translocates to the nucleus and displays transcriptional activity in a PINK1- and Parkin-dependent manner. MITF and TFE3, homologues of TFEB belonging to the same microphthalmia/transcription factor E (MiT/TFE) family, are similarly regulated during mitophagy. Unlike TFEB translocation after starvation-induced mammalian target of rapamycin complex 1 inhibition, Parkin-mediated TFEB relocalization required Atg9A and Atg5 activity. However, constitutively active Rag guanosine triphosphatases prevented TFEB translocation during mitophagy, suggesting cross talk between these two MiT/TFE activation pathways. Analysis of clustered regularly interspaced short palindromic repeats-generated TFEB/MITF/TFE3/TFEC single, double, and triple knockout cell lines revealed that these proteins partly facilitate Parkin-mediated mitochondrial clearance. These results illuminate a pathway leading to MiT/TFE transcription factor activation, distinct from starvation-induced autophagy, which occurs during mitophagy.

  19. Inflammatory transcription factors as activation markers and functional readouts in immune-to-brain communication.

    Science.gov (United States)

    Rummel, Christoph

    2016-05-01

    Immune-to-brain communication pathways involve humoral mediators, including cytokines, central modulation by neuronal afferents and immune cell trafficking to the brain. During systemic inflammation these pathways contribute to mediating brain-controlled sickness symptoms including fever. Experimentally, activation of these signaling pathways can be mimicked and studied when injecting animals with pathogen associated molecular patterns (PAMPS). One central component of the brain inflammatory response, which leads, for example, to fever induction, is transcriptional activation of brain cells via cytokines and PAMPS. We and others have studied the spatiotemporal activation and the physiological significance of transcription factors for the induction of inflammation within the brain and the manifestation of fever. Evidence has revealed a role of nuclear factor (NF)κB in the initiation, signal transducer and activator of transcription (STAT)3 in the maintenance and NF-interleukin (IL)6 in the maintenance or even termination of brain-inflammation and fever. Moreover, psychological stressors, such as exposure to a novel environment, leads to increased body core temperature and genomic NF-IL6-activation, suggesting a potential use of NF-IL6-immunohistochemistry as a multimodal brain cell activation marker and a role for NF-IL6 for differential brain activity. In addition, the nutritional status, as reflected by circulating levels of the cytokine-like hormone leptin, influence immune-to-brain communication and age-dependent changes in LPS-induced fever. Overall, transcription factors remain therapeutically important targets for the treatment of brain-inflammation and fever induction during infectious/non-infectious inflammatory and psychological stress. However, the exact physiological role and significance of these transcription factors requires to be further investigated.

  20. The Drosophila Transcription Factors Tinman and Pannier Activate and Collaborate with Myocyte Enhancer Factor-2 to Promote Heart Cell Fate.

    Directory of Open Access Journals (Sweden)

    TyAnna L Lovato

    Full Text Available Expression of the MADS domain transcription factor Myocyte Enhancer Factor 2 (MEF2 is regulated by numerous and overlapping enhancers which tightly control its transcription in the mesoderm. To understand how Mef2 expression is controlled in the heart, we identified a late stage Mef2 cardiac enhancer that is active in all heart cells beginning at stage 14 of embryonic development. This enhancer is regulated by the NK-homeodomain transcription factor Tinman, and the GATA transcription factor Pannier through both direct and indirect interactions with the enhancer. Since Tinman, Pannier and MEF2 are evolutionarily conserved from Drosophila to vertebrates, and since their vertebrate homologs can convert mouse fibroblast cells to cardiomyocytes in different activator cocktails, we tested whether over-expression of these three factors in vivo could ectopically activate known cardiac marker genes. We found that mesodermal over-expression of Tinman and Pannier resulted in approximately 20% of embryos with ectopic Hand and Sulphonylurea receptor (Sur expression. By adding MEF2 alongside Tinman and Pannier, a dramatic expansion in the expression of Hand and Sur was observed in almost all embryos analyzed. Two additional cardiac markers were also expanded in their expression. Our results demonstrate the ability to initiate ectopic cardiac fate in vivo by the combination of only three members of the conserved Drosophila cardiac transcription network, and provide an opportunity for this genetic model system to be used to dissect the mechanisms of cardiac specification.

  1. Eukaryotic transcription factors

    DEFF Research Database (Denmark)

    Staby, Lasse; O'Shea, Charlotte; Willemoës, Martin

    2017-01-01

    Gene-specific transcription factors (TFs) are key regulatory components of signaling pathways, controlling, for example, cell growth, development, and stress responses. Their biological functions are determined by their molecular structures, as exemplified by their structured DNA-binding domains...... targeting specific cis-acting elements in genes, and by the significant lack of fixed tertiary structure in their extensive intrinsically disordered regions. Recent research in protein intrinsic disorder (ID) has changed our understanding of transcriptional activation domains from 'negative noodles' to ID...... them to participate in large interactomes, how they use only a few hydrophobic residues, short sequence motifs, prestructured motifs, and coupled folding and binding for their interactions with co-activators, and how their accessibility to post-translational modification affects their interactions...

  2. Nucleocytoplasmic shuttling of transcription factors

    DEFF Research Database (Denmark)

    Cartwright, P; Helin, K

    2000-01-01

    To elicit the transcriptional response following intra- or extracellular stimuli, the signals need to be transmitted to their site of action within the nucleus. The nucleocytoplasmic shuttling of transcription factors is a mechanism mediating this process. The activation and inactivation...... of the transcriptional response is essential for cells to progress through the cell cycle in a normal manner. The involvement of cytoplasmic and nuclear accessory molecules, and the general nuclear membrane transport components, are essential for this process. Although nuclear import and export for different...... transcription factor families are regulated by similar mechanisms, there are several differences that allow for the specific activation of each transcription factor. This review discusses the general import and export pathways found to be common amongst many different transcription factors, and highlights...

  3. Nucleocytoplasmic shuttling of transcription factors

    DEFF Research Database (Denmark)

    Cartwright, P; Helin, K

    2000-01-01

    To elicit the transcriptional response following intra- or extracellular stimuli, the signals need to be transmitted to their site of action within the nucleus. The nucleocytoplasmic shuttling of transcription factors is a mechanism mediating this process. The activation and inactivation...... transcription factor families are regulated by similar mechanisms, there are several differences that allow for the specific activation of each transcription factor. This review discusses the general import and export pathways found to be common amongst many different transcription factors, and highlights...... of the transcriptional response is essential for cells to progress through the cell cycle in a normal manner. The involvement of cytoplasmic and nuclear accessory molecules, and the general nuclear membrane transport components, are essential for this process. Although nuclear import and export for different...

  4. Multiple transcription factor codes activate epidermal wound-response genes in Drosophila.

    Science.gov (United States)

    Pearson, Joseph C; Juarez, Michelle T; Kim, Myungjin; Drivenes, Øyvind; McGinnis, William

    2009-02-17

    Wounds in Drosophila and mouse embryos induce similar genetic pathways to repair epidermal barriers. However, the transcription factors that transduce wound signals to repair epidermal barriers are largely unknown. We characterize the transcriptional regulatory enhancers of 4 genes-Ddc, ple, msn, and kkv-that are rapidly activated in epidermal cells surrounding wounds in late Drosophila embryos and early larvae. These epidermal wound enhancers all contain evolutionarily conserved sequences matching binding sites for JUN/FOS and GRH transcription factors, but vary widely in trans- and cis-requirements for these inputs and their binding sites. We propose that the combination of GRH and FOS is part of an ancient wound-response pathway still used in vertebrates and invertebrates, but that other mechanisms have evolved that result in similar transcriptional output. A common, but largely untested assumption of bioinformatic analyses of gene regulatory networks is that transcription units activated in the same spatial and temporal patterns will require the same cis-regulatory codes. Our results indicate that this is an overly simplistic view.

  5. Resveratrol inhibits growth of orthotopic pancreatic tumors through activation of FOXO transcription factors.

    Directory of Open Access Journals (Sweden)

    Sanjit K Roy

    Full Text Available BACKGROUND: The forkhead transcription factors of the O class (FOXO play a direct role in cellular proliferation, oxidative stress response, and tumorigenesis. The objectives of this study were to examine whether FOXOs regulate antitumor activities of resveratrol in pancreatic cancer cells in vitro and in vivo. METHODOLOGY/PRINCIPAL FINDINGS: Pancreatic cancer cell lines were treated with resveratrol. Cell viability, colony formation, apoptosis and cell cycle were measured by XTT, soft agar, TUNEL and flow cytometry assays, respectively. FOXO nuclear translocation, DNA binding and transcriptional activities were measured by fluorescence technique, gelshift and luciferase assay, respectively. Mice were orthotopically implanted with PANC1 cells and orally gavaged with resveratrol. The components of PI3K and ERK pathways, FOXOs and their target gene expressions were measured by the Western blot analysis. Resveratrol inhibited cell viability and colony formations, and induced apoptosis through caspase-3 activation in four pancreatic cancer cell lines (PANC-1, MIA PaCa-2, Hs766T, and AsPC-1. Resveratrol induced cell cycle arrest by up-regulating the expression of p21/CIP1, p27/KIP1 and inhibiting the expression of cyclin D1. Resveratrol induced apoptosis by up-regulating Bim and activating caspase-3. Resveratrol inhibited phosphorylation of FOXOs, and enhanced their nuclear translocation, FOXO-DNA binding and transcriptional activities. The inhibition of PI3K/AKT and MEK/ERK pathways induced FOXO transcriptional activity and apoptosis. Furthermore, deletion of FOXO genes abrogated resveratrol-induced cell cycle arrest and apoptosis. Finally, resveratrol-treated mice showed significant inhibition in tumor growth which was associated with reduced phosphorylation of ERK, PI3K, AKT, FOXO1 and FOXO3a, and induction of apoptosis and FOXO target genes. CONCLUSIONS: These data suggest that inhibition of ERK and AKT pathways act together to activate FOXO

  6. Elk3 from hamster-a ternary complex factor with strong transcriptional repressor activity

    DEFF Research Database (Denmark)

    Hjortoe, G.M.; Weilguny, D.; Willumsen, Berthe Marie

    2005-01-01

    the transcription of genes that are activated during entry into G1. We have isolated the Cricetulus griseus Elk3 gene from the Chinese hamster ovary (CHO) cell line and investigated the transcriptional potential of this factor. Transient transfections revealed that, in addition to its regulation of the c......-fos promoter, Elk3 from CHO cells seems to inhibit other promoters controlling expression of proteins involved in G1/S phase progression; Cyclin D1 and DHFR. As has been described for the Elk3 homologs Net (Mouse) and Sap-2 (Human), the results of the present study further indicate that hamster Elk3...

  7. Genetic factors affecting gene transcription and catalytic activity of UDP-glucuronosyltransferases in human liver.

    Science.gov (United States)

    Liu, Wanqing; Ramírez, Jacqueline; Gamazon, Eric R; Mirkov, Snezana; Chen, Peixian; Wu, Kehua; Sun, Chang; Cox, Nancy J; Cook, Edwin; Das, Soma; Ratain, Mark J

    2014-10-15

    The aim of this study was to discover cis- and trans-acting factors significantly affecting mRNA expression and catalytic activity of human hepatic UDP-glucuronosyltransferases (UGTs). Transcription levels of five major hepatic UGT1A (UGT1A1, UGT1A3, UGT1A4, UGT1A6 and UGT1A9) and five UGT2B (UGT2B4, UGT2B7, UGT2B10, UGT2B15 and UGT2B17) genes were quantified in human liver tissue samples (n = 125) using real-time PCR. Glucuronidation activities of 14 substrates were measured in 47 livers. We genotyped 167 tagSNPs (single-nucleotide polymorphisms) in UGT1A (n = 43) and UGT2B (n = 124), as well as the known functional UGT1A1*28 and UGT2B17 CNV (copy number variation) polymorphisms. Transcription levels of 15 transcription factors (TFs) known to regulate these UGTs were quantified. We found that UGT expression and activity were highly variable among the livers (median and range of coefficient of variations: 135%, 74-217% and 52%, 39-105%, respectively). CAR, PXR and ESR1 were found to be the most important trans-regulators of UGT transcription (median and range of correlation coefficients: 46%, 6-58%; 47%, 9-58%; and 52%, 24-75%, respectively). Hepatic UGT activities were mainly determined by UGT gene transcription levels. Twenty-one polymorphisms were significantly (FDR-adjusted P transcription and testosterone glucuronidation rate, in addition to that attributable to the UGT2B17 CNV. Our study discovered novel pharmacogenetic markers and provided detailed insight into the genetic network regulating hepatic UGTs.

  8. Cisplatin induces cytotoxicity through the mitogen-activated protein kinase pathways and activating transcription factor 3.

    Science.gov (United States)

    St Germain, Carly; Niknejad, Nima; Ma, Laurie; Garbuio, Kyla; Hai, Tsonwin; Dimitroulakos, Jim

    2010-07-01

    The mechanisms underlying the proapoptotic effect of the chemotherapeutic agent, cisplatin, are largely undefined. Understanding the mechanisms regulating cisplatin cytotoxicity may uncover strategies to enhance the efficacy of this important therapeutic agent. This study evaluates the role of activating transcription factor 3 (ATF3) as a mediator of cisplatin-induced cytotoxicity. Cytotoxic doses of cisplatin and carboplatin treatments consistently induced ATF3 expression in five tumor-derived cell lines. Characterization of this induction revealed a p53, BRCA1, and integrated stress response-independent mechanism, all previously implicated in stress-mediated ATF3 induction. Analysis of mitogen-activated protein kinase (MAPK) pathway involvement in ATF3 induction by cisplatin revealed a MAPK-dependent mechanism. Cisplatin treatment combined with specific inhibitors to each MAPK pathway (c-Jun N-terminal kinase, extracellular signal-regulated kinase, and p38) resulted in decreased ATF3 induction at the protein level. MAPK pathway inhibition led to decreased ATF3 messenger RNA expression and reduced cytotoxic effects of cisplatin as measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide cell viability assay. In A549 lung carcinoma cells, targeting ATF3 with specific small hairpin RNA also attenuated the cytotoxic effects of cisplatin. Similarly, ATF3-/- murine embryonic fibroblasts (MEFs) were shown to be less sensitive to cisplatin-induced cytotoxicity compared with ATF3+/+ MEFs. This study identifies cisplatin as a MAPK pathway-dependent inducer of ATF3, whose expression influences cisplatin's cytotoxic effects.

  9. Arsenic Directly Binds to and Activates the Yeast AP-1-Like Transcription Factor Yap8

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, Nallani Vijay; Yang, Jianbo; Pillai, Jitesh K.; Rawat, Swati; Solano, Carlos; Kumar, Abhay; Grøtli, Morten; Stemmler, Timothy L.; Rosen, Barry P.; Tamás, Markus J.

    2015-12-28

    The AP-1-like transcription factor Yap8 is critical for arsenic tolerance in the yeastSaccharomyces cerevisiae. However, the mechanism by which Yap8 senses the presence of arsenic and activates transcription of detoxification genes is unknown. Here we demonstrate that Yap8 directly binds to trivalent arsenite [As(III)]in vitroandin vivoand that approximately one As(III) molecule is bound per molecule of Yap8. As(III) is coordinated by three sulfur atoms in purified Yap8, and our genetic and biochemical data identify the cysteine residues that form the binding site as Cys132, Cys137, and Cys274. As(III) binding by Yap8 does not require an additional yeast protein, and Yap8 is regulated neither at the level of localization nor at the level of DNA binding. Instead, our data are consistent with a model in which a DNA-bound form of Yap8 acts directly as an As(III) sensor. Binding of As(III) to Yap8 triggers a conformational change that in turn brings about a transcriptional response. Thus, As(III) binding to Yap8 acts as a molecular switch that converts inactive Yap8 into an active transcriptional regulator. This is the first report to demonstrate how a eukaryotic protein couples arsenic sensing to transcriptional activation.

  10. An Efficient Method to Identify Conditionally Activated Transcription Factors and their Corresponding Signal Transduction Pathway Segments

    Directory of Open Access Journals (Sweden)

    Haiyan Hu

    2009-11-01

    Full Text Available A signal transduction pathway (STP is a cascade composed of a series of signal transferring steps, which often activate one or more transcription factors (TFs to control the transcription of target genes. Understanding signaling pathways is important to our understanding of the molecular mechanisms of disease. Many condition-annotated pathways have been deposited in public databases. However, condition-annotated pathways are far from complete, considering the large number of possible conditions. Computational methods to assist in the identification of conditionally activated pathways are greatly needed. In this paper, we propose an efficient method to identify conditionally activated pathway segments starting from the identification of conditionally activated TFs, by incorporating protein-DNA binding data, gene expression data and protein interaction data. Applying our methods on several microarray datasets, we have discovered many significantly activated TFs and their corresponding pathway segments, which are supported by evidence in the literature.

  11. The Transcription Factor Encyclopedia

    DEFF Research Database (Denmark)

    Yusuf, Dimas; Butland, Stefanie L; Swanson, Magdalena I

    2012-01-01

    ABSTRACT: Here we present the Transcription Factor Encyclopedia (TFe), a new web-based compendium of mini review articles on transcription factors (TFs) that is founded on the principles of open access and collaboration. Our consortium of over 100 researchers has collectively contributed over 130...

  12. Plants contain a novel multi-member class of heat shock factors without transcriptional activator potential.

    Science.gov (United States)

    Czarnecka-Verner, E; Yuan, C X; Scharf, K D; Englich, G; Gurley, W B

    2000-07-01

    Based on phylogeny of DNA-binding domains and the organization of hydrophobic repeats, two families of heat shock transcription factors (HSFs) exist in plants. Class A HSFs are involved in the activation of the heat shock response, but the role of class B HSFs is not clear. When transcriptional activities of full-length HSFs were monitored in tobacco protoplasts, no class B HSFs from soybean or Arabidopsis showed activity under control or heat stress conditions. Additional assays confirmed the finding that the class B HSFs lacked the capacity to activate transcription. Fusion of a heterologous activation domain from human HSF1 (AD2) to the C-terminus of GmHSFB1-34 gave no evidence of synergistic enhancement of AD2 activity, which would be expected if weak activation domains were present. Furthermore, activity of AtHSFB1-4 (class B) was not rescued by coexpression with AtHSFA4-21 (class A) indicating that the class A HSF was not able to provide a missing function required for class B activity. The transcriptional activation potential of Arabidopsis AtHSFA4-21 was mapped primarily to a 39 amino acid fragment in the C-terminus enriched in bulky hydrophobic and acidic residues. Deletion mutagenesis of the C-terminal activator regions of tomato and Arabidopsis HSFs indicated that these plant HSFs lack heat-inducible regulatory regions analogous to those of mammalian HSF1. These findings suggest that heat shock regulation in plants may differ from metazoans by partitioning negative and positive functional domains onto separate HSF proteins. Class A HSFs are primarily responsible for stress-inducible activation of heat shock genes whereas some of the inert class B HSFs may be specialized for repression, or down-regulation, of the heat shock response.

  13. Transcription factor TnrA inhibits the biosynthetic activity of glutamine synthetase in Bacillus subtilis.

    Science.gov (United States)

    Fedorova, Ksenia; Kayumov, Airat; Woyda, Kathrin; Ilinskaja, Olga; Forchhammer, Karl

    2013-05-02

    The Bacillus subtilis glutamine synthetase (GS) plays a dual role in cell metabolism by functioning as catalyst and regulator. GS catalyses the ATP-dependent synthesis of glutamine from glutamate and ammonium. Under nitrogen-rich conditions, GS becomes feedback-inhibited by high intracellular glutamine levels and then binds transcription factors GlnR and TnrA, which control the genes of nitrogen assimilation. While GS-bound TnrA is no longer able to interact with DNA, GlnR-DNA binding is shown to be stimulated by GS complex formation. In this paper we show a new physiological feature of the interaction between glutamine synthetase and TnrA. The transcription factor TnrA inhibits the biosynthetic activity of glutamine synthetase in vivo and in vitro, while the GlnR protein does not affect the activity of the enzyme.

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

    Directory of Open Access Journals (Sweden)

    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.

  15. Copper is required for cobalt-induced transcriptional activity of hypoxia-inducible factor-1.

    Science.gov (United States)

    Qiu, Liying; Ding, Xueqin; Zhang, Zhen; Kang, Y James

    2012-08-01

    Cobalt inhibits prolyl hydroxylases, leading to the accumulation of hypoxia-inducible factor-1α (HIF-1α) and a concomitant increase in the transcriptional activity of HIF-1. Therefore, cobalt has been under development as a drug for activating HIF-1 under some disease conditions. However, it has been shown that ischemic conditions resulted in the loss of copper, and the activation of HIF-1 would not occur unless copper was supplemented. The present study was undertaken to test the hypothesis that copper is also required for the cobalt activation of HIF-1 transcriptional activity. Human umbilical vein endothelial cells subjected to treatment with cobalt chloride (CoCl(2)) at concentrations above 25 μM for 2 h resulted in an accumulation of HIF-1α, which was determined by Western blot analysis, and an increase in the expression of vascular endothelial growth factor (VEGF), which was determined by real-time reverse transcription-polymerase chain reaction analysis for mRNA levels and enzyme-linked immunosorbent assay analysis for protein levels. The copper chelator tetraethylenepentamine at 25 μM did not significantly affect the accumulation of HIF-1α but blocked increases in VEGF mRNA and protein levels, an effect that could be reversed by the addition of 25 μM copper sulfate (CuSO(4)). In addition, gene silencing of the copper chaperone for Cu,Zn-superoxide dismutase blocked VEGF expression with little effect on cobalt-induced HIF-1α accumulation. The present study thus demonstrates that copper was required for cobalt-activated transcriptional activity of HIF-1, although copper did not affect cobalt-induced accumulation of HIF-1α in the cells.

  16. Combinatorial activation and repression by seven transcription factors specify Drosophila odorant receptor expression.

    Directory of Open Access Journals (Sweden)

    Shadi Jafari

    Full Text Available The mechanism that specifies olfactory sensory neurons to express only one odorant receptor (OR from a large repertoire is critical for odor discrimination but poorly understood. Here, we describe the first comprehensive analysis of OR expression regulation in Drosophila. A systematic, RNAi-mediated knock down of most of the predicted transcription factors identified an essential function of acj6, E93, Fer1, onecut, sim, xbp1, and zf30c in the regulation of more than 30 ORs. These regulatory factors are differentially expressed in antennal sensory neuron classes and specifically required for the adult expression of ORs. A systematic analysis reveals not only that combinations of these seven factors are necessary for receptor gene expression but also a prominent role for transcriptional repression in preventing ectopic receptor expression. Such regulation is supported by bioinformatics and OR promoter analyses, which uncovered a common promoter structure with distal repressive and proximal activating regions. Thus, our data provide insight into how combinatorial activation and repression can allow a small number of transcription factors to specify a large repertoire of neuron classes in the olfactory system.

  17. 24. The transcription factors and the relevant signaling pathways activated by low concentration MNNG

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Aims: To explore the transcription factors and related signal transduction pathways activated in the alkylating agents N-methyl-N'-nitro-N-nitrosoguanindine (MNNG) exposed cells which may involved in the mechanism of MNNG induced changes of gene expression, especially the elevation of DNA polymerase β expression and also the consequence of JNK kinase activation which were reported previously in this lab. Methods: Clontech Mercury pathway profiling system containing 8 different vectors in which a specific response element is located upstream from the SEAP-reporter gene were employed to detect the transcription factor activation in Vero cells treated with 0.2 μmol/L MNNG for 2 hours. Thoroughly, CREB phosphorylation, protein kinase A (PKA) and the cellular cAMP content were also assayed with PhosphoPlus CREB (ser-133) antibody kit, protein kinase assay kit and cAMP RIA kit respectively. Results: Among 8 different response elements, the expression of the reporter gene governed by the transcription factors CREB (cAMP response element binding protein), AP1 (activator protein 1), NF-κB (nuclear factor κ B) were elevated by 1.3, 1.4 and 1.3 times higber than control respectively. The level of activated CREB by Ser-133 phosphorylation was 2.08 times higher than control in cells treated with MNNG for 60 min, as measured by immunoblotting. The activity of CREB upstream kinase protein kinase A (PKA), which can phosphorylate CREB on ser-133 was also activated, and the activation peaked at 60 min (11.03±2.80 arbitrary units vs 0.86±0.43 of control). Also, cAMP levels were significantly raised after 60-minute-treatment, 1.52 times higher vs those in solvent control. Conclusion: In addition of previously reported JNK activation, we show here that low concentration alkylating agent MNNG can also activate the cAMP-PKA and NF-κB pathway. These in consequence induce the activation of transcription factors APl, CREB and NF-κB, which may related to the MNNG induced changes in

  18. A light- and calcium-gated transcription factor for imaging and manipulating activated neurons.

    Science.gov (United States)

    Wang, Wenjing; Wildes, Craig P; Pattarabanjird, Tanyaporn; Sanchez, Mateo I; Glober, Gordon F; Matthews, Gillian A; Tye, Kay M; Ting, Alice Y

    2017-09-01

    Activity remodels neurons, altering their molecular, structural, and electrical characteristics. To enable the selective characterization and manipulation of these neurons, we present FLARE, an engineered transcription factor that drives expression of fluorescent proteins, opsins, and other genetically encoded tools only in the subset of neurons that experienced activity during a user-defined time window. FLARE senses the coincidence of elevated cytosolic calcium and externally applied blue light, which together produce translocation of a membrane-anchored transcription factor to the nucleus to drive expression of any transgene. In cultured rat neurons, FLARE gives a light-to-dark signal ratio of 120 and a high- to low-calcium signal ratio of 10 after 10 min of stimulation. Opsin expression permitted functional manipulation of FLARE-marked neurons. In adult mice, FLARE also gave light- and motor-activity-dependent transcription in the cortex. Due to its modular design, minute-scale temporal resolution, and minimal dark-state leak, FLARE should be useful for the study of activity-dependent processes in neurons and other cells that signal with calcium.

  19. MYRF is a membrane-associated transcription factor that autoproteolytically cleaves to directly activate myelin genes.

    Directory of Open Access Journals (Sweden)

    Helena Bujalka

    Full Text Available The myelination of axons is a crucial step during vertebrate central nervous system (CNS development, allowing for rapid and energy efficient saltatory conduction of nerve impulses. Accordingly, the differentiation of oligodendrocytes, the myelinating cells of the CNS, and their expression of myelin genes are under tight transcriptional control. We previously identified a putative transcription factor, Myelin Regulatory Factor (Myrf, as being vital for CNS myelination. Myrf is required for the generation of CNS myelination during development and also for its maintenance in the adult. It has been controversial, however, whether Myrf directly regulates transcription, with reports of a transmembrane domain and lack of nuclear localization. Here we show that Myrf is a membrane-associated transcription factor that undergoes an activating proteolytic cleavage to separate its transmembrane domain-containing C-terminal region from a nuclear-targeted N-terminal region. Unexpectedly, this cleavage event occurs via a protein domain related to the autoproteolytic intramolecular chaperone domain of the bacteriophage tail spike proteins, the first time this domain has been found to play a role in eukaryotic proteins. Using ChIP-Seq we show that the N-terminal cleavage product directly binds the enhancer regions of oligodendrocyte-specific and myelin genes. This binding occurs via a defined DNA-binding consensus sequence and strongly promotes the expression of target genes. These findings identify Myrf as a novel example of a membrane-associated transcription factor and provide a direct molecular mechanism for its regulation of oligodendrocyte differentiation and CNS myelination.

  20. SENSITIVE TO PROTON RHIZOTOXICITY1, CALMODULIN BINDING TRANSCRIPTION ACTIVATOR2, and other transcription factors are involved in ALUMINUM-ACTIVATED MALATE TRANSPORTER1 expression.

    Science.gov (United States)

    Tokizawa, Mutsutomo; Kobayashi, Yuriko; Saito, Tatsunori; Kobayashi, Masatomo; Iuchi, Satoshi; Nomoto, Mika; Tada, Yasuomi; Yamamoto, Yoshiharu Y; Koyama, Hiroyuki

    2015-03-01

    In Arabidopsis (Arabidopsis thaliana) the root apex is protected from aluminum (Al) rhizotoxicity by excretion of malate, an Al chelator, by ALUMINUM-ACTIVATED MALATE TRANSPORTER1 (AtALMT1). AtALMT1 expression is fundamentally regulated by the SENSITIVE TO PROTON RHIZOTOXICITY1 (STOP1) zinc finger protein, but other transcription factors have roles that enable Al-inducible expression with a broad dynamic range. In this study, we characterized multiple cis-elements in the AtALMT1 promoter that interact with transcription factors. In planta complementation assays of AtALMT1 driven by 5' truncated promoters of different lengths showed that the promoter region between -540 and 0 (the first ATG) restored the Al-sensitive phenotype of atalm1 and thus contains cis-elements essential for AtALMT1 expression for Al tolerance. Computation of overrepresented octamers showed that eight regions in this promoter region contained potential cis-elements involved in Al induction and STOP1 regulation. Mutation in a position around -297 from the first ATG completely inactivated AtALMT1 expression and Al response. In vitro binding assays showed that this region contained the STOP1 binding site, which accounted for the recognition by four zinc finger domains of the protein. Other positions were characterized as cis-elements that regulated expression by repressors and activators and a transcription factor that determines root tip expression of AtALMT1. From the consensus of known cis-elements, we identified CALMODULIN-BINDING TRANSCRIPTION ACTIVATOR2 to be an activator of AtALMT1 expression. Al-inducible expression of AtALMT1 changed transcription starting sites, which increased the abundance of transcripts with a shortened 5' untranslated region. The present analyses identified multiple mechanisms that regulate AtALMT1 expression.

  1. Brain-derived neurotrophic factor activation of NFAT (nuclear factor of activated T-cells)-dependent transcription: a role for the transcription factor NFATc4 in neurotrophin-mediated gene expression.

    Science.gov (United States)

    Groth, Rachel D; Mermelstein, Paul G

    2003-09-03

    A member of the neurotrophin family, brain-derived neurotrophic factor (BDNF) regulates neuronal survival and differentiation during development. Within the adult brain, BDNF is also important in neuronal adaptive processes, such as the activity-dependent plasticity that underlies learning and memory. These long-term changes in synaptic strength are mediated through alterations in gene expression. However, many of the mechanisms by which BDNF is linked to transcriptional and translational regulation remain unknown. Recently, the transcription factor NFATc4 (nuclear factor of activated T-cells isoform 4) was discovered in neurons, where it is believed to play an important role in long-term changes in neuronal function. Interestingly, NFATc4 is particularly sensitive to the second messenger systems activated by BDNF. Thus, we hypothesized that NFAT-dependent transcription may be an important mediator of BDNF-induced plasticity. In cultured rat CA3-CA1 hippocampal neurons, BDNF activated NFAT-dependent transcription via TrkB receptors. Inhibition of calcineurin blocked BDNF-induced nuclear translocation of NFATc4, thus preventing transcription. Further, phospholipase C was a critical signaling intermediate between BDNF activation of TrkB and the initiation of NFAT-dependent transcription. Both inositol 1,4,5-triphosphate (IP3)-mediated release of calcium from intracellular stores and activation of protein kinase C were required for BDNF-induced NFAT-dependent transcription. Finally, increased expression of IP3 receptor 1 and BDNF after neuronal exposure to BDNF was linked to NFAT-dependent transcription. These results suggest that NFATc4 plays a crucial role in neurotrophin-mediated synaptic plasticity.

  2. Resveratrol induces growth arrest and apoptosis through activation of FOXO transcription factors in prostate cancer cells.

    Directory of Open Access Journals (Sweden)

    Qinghe Chen

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

  3. Molecular genetic analysis of activation-tagged transcription factors thought to be involved in photomorphogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Neff, Michael M.

    2011-06-23

    This is a final report for Department of Energy Grant No. DE-FG02-08ER15927 entitled “Molecular Genetic Analysis of Activation-Tagged Transcription Factors Thought to be Involved in Photomorphogenesis”. Based on our preliminary photobiological and genetic analysis of the sob1-D mutant, we hypothesized that OBP3 is a transcription factor involved in both phytochrome and cryptochrome-mediated signal transduction. In addition, we hypothesized that OBP3 is involved in auxin signaling and root development. Based on our preliminary photobiological and genetic analysis of the sob2-D mutant, we also hypothesized that a related gene, LEP, is involved in hormone signaling and seedling development.

  4. Differential expression of two activating transcription factor 5 isoforms in papillary thyroid carcinoma

    Science.gov (United States)

    Vicari, Luisa; La Rosa, Cristina; Forte, Stefano; Calabrese, Giovanna; Colarossi, Cristina; Aiello, Eleonora; Salluzzo, Salvatore; Memeo, Lorenzo

    2016-01-01

    Background Activating transcription factor 5 (ATF5) is a member of the activating transcription/cAMP response element-binding protein family of basic leucine zipper proteins that plays an important role in cell survival, differentiation, proliferation, and apoptosis. The ATF5 gene generates two transcripts: ATF5 isoform 1 and ATF5 isoform 2. A number of studies indicate that ATF5 could be an attractive target for therapeutic intervention in several tumor types; however, so far, the role of ATF5 has not been investigated in papillary thyroid carcinoma (PTC). Methods Quantitative real-time reverse transcription polymerase chain reaction and immuno-histochemical staining were used to study ATF5 mRNA and protein expression in PTC. Results We report here that ATF5 is expressed more in PTC tissue than in normal thyroid tissue. Furthermore, this is the first study that describes the presence of both ATF5 isoforms in PTC. Conclusion These findings could provide potential applications in PTC cancer treatment.

  5. Cisplatin Induces Cytotoxicity through the Mitogen-Activated Protein Kinase Pathways ana Activating Transcription Factor 3

    Directory of Open Access Journals (Sweden)

    Carly St. Germain

    2010-07-01

    Full Text Available The mechanisms underlying the proapoptotic effect of the chemotherapeutic agent, cisplatin, are largely undefined. Understanding the mechanisms regulating cisplatin cytotoxicity may uncover strategies to enhance the efficacy of this important therapeutic agent. This study evaluates the role of activating transcription factor 3 (ATF3 as a mediator of cisplatin-induced cytotoxicity. Cytotoxic doses of cisplatin and carboplatin treatments consistently induced ATF3 expression in five tumor-derived cell lines. Characterization of this induction revealed a p53, BRCA1, and integrated stress response-independent mechanism, all previously implicated in stress-mediated ATF3 induction. Analysis of mitogenactivated protein kinase (MAPK pathway involvement in ATF3 induction by cisplatin revealed a MAPK-dependent mechanism. Cisplatin treatment combined with specific inhibitors to each MAPK pathway (c-Jun N-terminal kinase, extracellularsignal-regulated kinase, and p38 resulted in decreasedATF3 induction at the protein level. MAPK pathway inhibition led to decreased ATF3 messenger RNA expression and reduced cytotoxic effects of cisplatin as measured by the 3-(4,5-dimethylthiazol-2-ylF2,5-diphenyltetrazolium bromide cell viability assay. In A549 lung carcinoma cells, targeting ATF3 with specific small hairpin RNA also attenuated the cytotoxic effects of cisplatin. Similarly, ATF3-/murine embryonic fibroblasts (MEFs were shown to be less sensitive to cisplatin-induced cytotoxicity compared with ATF3+/+ MEFs. This study identifies cisplatin as a MAPK pathway-dependent inducer of ATF3, whose expression influences cisplatin’s cytotoxic effects.

  6. A Modified Reverse One-Hybrid Screen Identifies Transcriptional Activation Domains in PHYTOCHROME-INTERACTING FACTOR 3.

    Science.gov (United States)

    Dalton, Jutta C; Bätz, Ulrike; Liu, Jason; Curie, Gemma L; Quail, Peter H

    2016-01-01

    Transcriptional activation domains (TADs) are difficult to predict and identify, since they are not conserved and have little consensus. Here, we describe a yeast-based screening method that is able to identify individual amino acid residues involved in transcriptional activation in a high throughput manner. A plant transcriptional activator, PIF3 (phytochrome interacting factor 3), was fused to the yeast GAL4-DNA-binding Domain (BD), driving expression of the URA3 (Orotidine 5'-phosphate decarboxylase) reporter, and used for negative selection on 5-fluroorotic acid (5FOA). Randomly mutagenized variants of PIF3 were then selected for a loss or reduction in transcriptional activation activity by survival on FOA. In the process, we developed a strategy to eliminate false positives from negative selection that can be used for both reverse-1- and 2-hybrid screens. With this method we were able to identify two distinct regions in PIF3 with transcriptional activation activity, both of which are functionally conserved in PIF1, PIF4, and PIF5. Both are collectively necessary for full PIF3 transcriptional activity, but neither is sufficient to induce transcription autonomously. We also found that the TAD appear to overlap physically with other PIF3 functions, such as phyB binding activity and consequent phosphorylation. Our protocol should provide a valuable tool for identifying, analyzing and characterizing novel TADs in eukaryotic transcription factors, and thus potentially contribute to the unraveling of the mechanism underlying transcriptional activation.

  7. A Modified Reverse One-Hybrid Screen Identifies Transcriptional Activation Domains in PHYTOCHROME-INTERACTING FACTOR 3

    Science.gov (United States)

    Dalton, Jutta C.; Bätz, Ulrike; Liu, Jason; Curie, Gemma L.; Quail, Peter H.

    2016-01-01

    Transcriptional activation domains (TADs) are difficult to predict and identify, since they are not conserved and have little consensus. Here, we describe a yeast-based screening method that is able to identify individual amino acid residues involved in transcriptional activation in a high throughput manner. A plant transcriptional activator, PIF3 (phytochrome interacting factor 3), was fused to the yeast GAL4-DNA-binding Domain (BD), driving expression of the URA3 (Orotidine 5′-phosphate decarboxylase) reporter, and used for negative selection on 5-fluroorotic acid (5FOA). Randomly mutagenized variants of PIF3 were then selected for a loss or reduction in transcriptional activation activity by survival on FOA. In the process, we developed a strategy to eliminate false positives from negative selection that can be used for both reverse-1- and 2-hybrid screens. With this method we were able to identify two distinct regions in PIF3 with transcriptional activation activity, both of which are functionally conserved in PIF1, PIF4, and PIF5. Both are collectively necessary for full PIF3 transcriptional activity, but neither is sufficient to induce transcription autonomously. We also found that the TAD appear to overlap physically with other PIF3 functions, such as phyB binding activity and consequent phosphorylation. Our protocol should provide a valuable tool for identifying, analyzing and characterizing novel TADs in eukaryotic transcription factors, and thus potentially contribute to the unraveling of the mechanism underlying transcriptional activation. PMID:27379152

  8. Differential requirement of the epidermal growth factor receptor for G protein-mediated activation of transcription factors by lysophosphatidic acid

    Directory of Open Access Journals (Sweden)

    Dent Paul

    2010-01-01

    Full Text Available Abstract Background The role of the epidermal growth factor receptor (EGFR and other receptor tyrosine kinases (RTKs in provoking biological actions of G protein-coupled receptors (GPCRs has been one of the most disputed subjects in the field of GPCR signal transduction. The purpose of the current study is to identify EGFR-mediated mechanisms involved in activation of G protein cascades and the downstream transcription factors by lysophosphatidic acid (LPA. Results In ovarian cancer cells highly responsive to LPA, activation of AP-1 by LPA was suppressed by inhibition of EGFR, an effect that could be reversed by co-stimulation of another receptor tyrosine kinase c-Met with hepatocyte growth factor, indicating that LPA-mediated activation of AP-1 requires activity of a RTK, not necessarily EGFR. Induction of AP-1 components by LPA lied downstream of Gi, G12/13, and Gq. Activation of the effectors of Gi, but not Gq or G12/13 was sensitive to inhibition of EGFR. In contrast, LPA stimulated another prominent transcription factor NF-κB via the Gq-PKC pathway in an EGFR-independent manner. Consistent with the importance of Gi-elicited signals in a plethora of biological processes, LPA-induced cytokine production, cell proliferation, migration and invasion require intact EGFR. Conclusions An RTK activity is required for activation of the AP-1 transcription factor and other Gi-dependent cellular responses to LPA. In contrast, activation of G12/13, Gq and Gq-elicited NF-κB by LPA is independent of such an input. These results provide a novel insight into the role of RTK in GPCR signal transduction and biological functions.

  9. Molecular genetics of blood-fleshed peach reveals activation of anthocyanin biosynthesis by NAC transcription factors.

    Science.gov (United States)

    Zhou, Hui; Lin-Wang, Kui; Wang, Huiliang; Gu, Chao; Dare, Andrew P; Espley, Richard V; He, Huaping; Allan, Andrew C; Han, Yuepeng

    2015-04-01

    Anthocyanin pigmentation is an important consumer trait in peach (Prunus persica). In this study, the genetic basis of the blood-flesh trait was investigated using the cultivar Dahongpao, which shows high levels of cyanidin-3-glucoside in the mesocarp. Elevation of anthocyanin levels in the flesh was correlated with the expression of an R2R3 MYB transcription factor, PpMYB10.1. However, PpMYB10.1 did not co-segregate with the blood-flesh trait. The blood-flesh trait was mapped to a 200-kb interval on peach linkage group (LG) 5. Within this interval, a gene encoding a NAC domain transcription factor (TF) was found to be highly up-regulated in blood-fleshed peaches when compared with non-red-fleshed peaches. This NAC TF, designated blood (BL), acts as a heterodimer with PpNAC1 which shows high levels of expression in fruit at late developmental stages. We show that the heterodimer of BL and PpNAC1 can activate the transcription of PpMYB10.1, resulting in anthocyanin pigmentation in tobacco. Furthermore, silencing the BL gene reduces anthocyanin pigmentation in blood-fleshed peaches. The transactivation activity of the BL-PpNAC1 heterodimer is repressed by a SQUAMOSA promoter-binding protein-like TF, PpSPL1. Low levels of PpMYB10.1 expression in fruit at early developmental stages is probably attributable to lower levels of expression of PpNAC1 plus the presence of high levels of repressors such as PpSPL1. We present a mechanism whereby BL is the key gene for the blood-flesh trait in peach via its activation of PpMYB10.1 in maturing fruit. Partner TFs such as basic helix-loop-helix proteins and NAC1 are required, as is the removal of transcriptional repressors.

  10. (-)-Epicatechin gallate (ECG) stimulates osteoblast differentiation via Runt-related transcription factor 2 (RUNX2) and transcriptional coactivator with PDZ-binding motif (TAZ)-mediated transcriptional activation.

    Science.gov (United States)

    Byun, Mi Ran; Sung, Mi Kyung; Kim, A Rum; Lee, Cham Han; Jang, Eun Jung; Jeong, Mi Gyeong; Noh, Minsoo; Hwang, Eun Sook; Hong, Jeong-Ho

    2014-04-01

    Osteoporosis is a degenerative bone disease characterized by low bone mass and is caused by an imbalance between osteoblastic bone formation and osteoclastic bone resorption. It is known that the bioactive compounds present in green tea increase osteogenic activity and decrease the risk of fracture by improving bone mineral density. However, the detailed mechanism underlying these beneficial effects has yet to be elucidated. In this study, we investigated the osteogenic effect of (-)-epicatechin gallate (ECG), a major bioactive compound found in green tea. We found that ECG effectively stimulates osteoblast differentiation, indicated by the increased expression of osteoblastic marker genes. Up-regulation of osteoblast marker genes is mediated by increased expression and interaction of the transcriptional coactivator with PDZ-binding motif (TAZ) and Runt-related transcription factor 2 (RUNX2). ECG facilitates nuclear localization of TAZ through PP1A. PP1A is essential for osteoblast differentiation because inhibition of PP1A activity was shown to suppress ECG-mediated osteogenic differentiation. Taken together, the results showed that ECG stimulates osteoblast differentiation through the activation of TAZ and RUNX2, revealing a novel mechanism for green tea-stimulated osteoblast differentiation.

  11. Dynamic transcription factor activity and networks during ErbB2 breast oncogenesis and targeted therapy.

    Science.gov (United States)

    Weiss, M S; Peñalver Bernabé, B; Shin, S; Asztalos, S; Dubbury, S J; Mui, M D; Bellis, A D; Bluver, D; Tonetti, D A; Saez-Rodriguez, J; Broadbelt, L J; Jeruss, J S; Shea, L D

    2014-12-01

    Tissue development and disease progression are multi-stage processes controlled by an evolving set of key regulatory factors, and identifying these factors necessitates a dynamic analysis spanning relevant time scales. Current omics approaches depend on incomplete biological databases to identify critical cellular processes. Herein, we present TRACER (TRanscriptional Activity CEll aRrays), which was employed to quantify the dynamic activity of numerous transcription factor (TFs) simultaneously in 3D and networks for TRACER (NTRACER), a computational algorithm that allows for cellular rewiring to establish dynamic regulatory networks based on activity of TF reporter constructs. We identified major hubs at various stages of culture associated with normal and abnormal tissue growth (i.e., ELK-1 and E2F1, respectively) and the mechanism of action for a targeted therapeutic, lapatinib, through GATA-1, which were confirmed in human ErbB2 positive breast cancer patients and human ErbB2 positive breast cancer cell lines that were either sensitive or resistant to lapatinib.

  12. Transcriptional Activity of Nuclear Factor κB Family Genes in Patients with Systemic Sclerosis.

    Science.gov (United States)

    Lis-Święty, Anna; Gola, Joanna; Mazurek, Urszula; Brzezińska-Wcisło, Ligia

    2017-05-01

    Systemic sclerosis (SSc) is a connective tissue disease of unknown etiology and unclear pathogenesis. Evaluation of the activation of nuclear factor κB (NF-κB) family genes IκBα, p50, p52, p65, and c-Rel, potentially involved in the regulation of immunity, inflammation, angiogenesis, and tissue remodeling in SSc, was carried out. The study included 19 patients with limited SSc, 11 patients with early SSc, and 10 healthy persons constituting the control group. Real-time QRT-PCR was used to evaluate the mRNAs in peripheral blood samples. The patients with early SSc showed a decrease in transcriptional activity of IκBα inhibitor and c-Rel subunit. Transcriptional activity decrease in the other patients with limited SSc included genes encoding c-Rel and p50, subunits of NF-κB factor. Deregulation of intracellular signal transduction by NF-κB takes place at the beginning of SSc and in its fibrosis stage. Associations between clinical variables and NF-κB related gene expression as well as the activation of NF-κB family members in SSc patients should be addressed in future studies. © 2017 by the Association of Clinical Scientists, Inc.

  13. An inducible transcription factor activates expression of human immunodeficiency virus in T cells

    Science.gov (United States)

    Nabel, Gary; Baltimore, David

    1987-04-01

    Human immunodeficiency virus (HIV) production from latently infected T lymphocytes can be induced with compounds that activate the cells to secrete lymphokines1,2. The elements in the HIV genome which control activation are not known but expression might be regulated through a variety of DNA elements. The cis-acting control elements of the viral genome are enhancer and promoter regions. The virus also encodes trans-acting factors specified by the tat-III (refs 3-6) and art genes7. We have examined whether products specific to activated T cells might stimulate viral transcription by binding to regions on viral DNA. Activation of T cells, which increases HIV expression up to 50-fold, correlated with induction of a DNA binding protein indistinguishable from a recognized transcription factor, called NF-κB (ref. 8), with binding sites in the viral enhancer. Mutation of these binding sites abolished inducibility. That NF-κB acts in synergy with the viral tat-III gene product to enhance HIV expression in T cells may have implications for the pathogenesis of AIDS (acquired immune deficiency syndrome).

  14. On involvement of transcription factors nuclear factor kappa-light-chain-enhancer of activated B cells, activator protein-1 and signal transducer and activator of transcription-3 in photodynamic therapy-induced death of crayfish neurons and satellite glial cells

    Science.gov (United States)

    Berezhnaya, Elena; Neginskaya, Marya; Kovaleva, Vera; Sharifulina, Svetlana; Ischenko, Irina; Komandirov, Maxim; Rudkovskii, Mikhail; Uzdensky, Anatoly B.

    2015-07-01

    Photodynamic therapy (PDT) is currently used in the treatment of brain tumors. However, not only malignant cells but also neighboring normal neurons and glial cells are damaged during PDT. In order to study the potential role of transcription factors-nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), activator protein (AP-1), and signal transducer and activator of transcription-3 (STAT-3)-in photodynamic injury of normal neurons and glia, we photosensitized the isolated crayfish mechanoreceptor consisting of a single sensory neuron enveloped by glial cells. Application of different inhibitors and activators showed that transcription factors NF-κB (inhibitors caffeic acid phenethyl ester and parthenolide, activator betulinic acid), AP-1 (inhibitor SR11302), and STAT-3 (inhibitors stattic and cucurbitacine) influenced PDT-induced death and survival of neurons and glial cells in different ways. These experiments indicated involvement of NF-κB in PDT-induced necrosis of neurons and apoptosis of glial cells. However, in glial cells, it played the antinecrotic role. AP-1 was not involved in PDT-induced necrosis of neurons and glia, but mediated glial apoptosis. STAT-3 was involved in PDT-induced apoptosis of glial cells and necrosis of neurons and glia. Therefore, signaling pathways that regulate cell death and survival in neurons and glial cells are different. Using various inhibitors or activators of transcription factors, one can differently influence the sensitivity and resistance of neurons and glial cells to PDT.

  15. Effects of calmodulin on DNA-binding activity of heat shock transcription factor in vitro

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The DNA-binding activity of heat shock transcription factor (HSF) was induced by heat shock (HS) of a whole cell extract. Addition of antiserum, specific to CaM, to a whole cell extract reduced bind of the HSF to the heat shock element (HSE) with maize, and the re-addition of CaM to the sample restored the activity of the HSF for binding to HSE. In addition, DNA-binding activity of the HSF was also induced by directly adding CaM to a whole cell extract at non-HS temperature with maize. Similar results were obtained with wheat and tomato. Our observations provide the first example of the involvement of CaM in regulation of the DNA-binding activity of the HSF.

  16. 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.

  17. Modulation of NF-кB transcription factor activation by Molluscum contagiosum virus proteins

    Directory of Open Access Journals (Sweden)

    Justyna Struzik

    2014-02-01

    Full Text Available Molluscum contagiosum virus is a human and animal dermatotropic pathogen, which causes a severe disease in immunocompromised individuals. MCV belongs to the Poxviridae family whose members exert immunomodulatory effects on the host antiviral response. Poxviruses interfere with cell signaling pathways that lead to the activation of nuclear factor кB, a pleiotropic transcription factor which is crucial for regulation of the immune response, the cell cycle and apoptosis. In resting cells, NF-κB is present in the cytoplasm, where it is associated with inhibitor κB. Upon stimulation by activators, such as proinflammatory cytokines and bacterial or viral products, the inhibitory protein undergoes phosphorylation and proteasomal degradation. NF-κB, in turn, translocates to the nucleus, where it regulates the transcription of various genes that are essential for processes mentioned above. Since poxviruses replicate exclusively in the cell cytoplasm, NF-кB became a good target for poxviral immunomodulation. MCV encodes various proteins which interfere with the signaling pathways that lead to the activation of NF-κB. Ligand inhibitor encoded by MCV, MC54, binds interleukin-18 and inhibits interferon-γ production. Other MCV proteins, MC159 and MC160, belong to intracellular inhibitors of NF-κB and are members of viral FLICE-inhibitory proteins (vFLIPs. MC159 protein encoded by MCV was shown to inhibit apoptosis of virus-infected cells. Such interactions serve immune evasion and are responsible for the persistence of MCV.

  18. Germ cell nuclear factor directly represses the transcription of peroxisome proliferator-activated receptor delta gene

    Institute of Scientific and Technical Information of China (English)

    Chengqiang He; Naizheng Ding; Jie Kang

    2008-01-01

    Germ cell nuclear factor (GCNF) is a transcription factor that can repress gene transcription and plays an important role during spermatogenesis. Peroxisome proliferator-activated receptor delta (PPARδ) is a nuclear hormone receptor belonging to the steroid receptor superfamily.It can activate the expression of many genes,including those involved in lipid metabolism.In this report,we showed that GCNF specifically interacts with PPARδ promoter.Overexpression of GCNF in African green monkey SV40 transformed kidney fibroblast COS7 cells and mouse embryo fibroblast NIH 3T3 cells represses the activity of PPARδ promoter.The mutation of GCNF response element in PPARδ promoter relieves the repression in NIH 3T3 cells and mouse testis.Moreover,we showed that GCNF in nuclear extracts of mouse testis is able to bind to PPARδ promoter directly.We also found that GCNF and PPARδ mRNA were expressed with different patterns in mouse testis by in situ hybridization.These results suggested that GCNF might be a negative regulator of PPARδ gene expression through its direct interaction with PPARδ promoter in mouse testis.

  19. Transcription activation of a UV-inducible Clostridium perfringens bacteriocin gene by a novel sigma factor.

    Science.gov (United States)

    Dupuy, Bruno; Mani, Nagraj; Katayama, Seiichi; Sonenshein, Abraham L

    2005-02-01

    Expression of the plasmid-encoded Clostridium perfringens gene for bacteriocin BCN5 was shown to depend in vivo and in vitro on the activity of UviA protein. UviA, also plasmid-encoded, proved to be an RNA polymerase sigma factor and was also partly autoregulatory. The uviA gene has two promoters; one provided a UviA-independent, basal level of gene expression while the stronger, UviA-dependent promoter was only utilized after the cell experienced DNA damage. As a result, BCN5 synthesis is induced by treatment with UV light or mitomycin C. UviA is related to a special class of sigma factors found to date only in Clostridium species and responsible for activating transcription of toxin genes in Clostridium difficile, Clostridium tetani, and Clostridium botulinum.

  20. Inducing goat pluripotent stem cells with four transcription factor mRNAs that activate endogenous promoters.

    Science.gov (United States)

    Chen, Hao; Zuo, Qisheng; Wang, Yingjie; Song, Jiuzhou; Yang, Huilin; Zhang, Yani; Li, Bichun

    2017-02-13

    Traditional approaches for generating goat pluripotent stem cells (iPSCs) suffer from complexity and low preparation efficiency. Therefore, we tried to derive goat iPSCs with a new method by transfecting exogenous Oct4, Sox2, Klf4 and c-Myc mRNAs into goat embryonic fibroblasts (GEFs), and explore the mechanisms regarding the transcription regulation of the reprogramming factors in goat iPSCs induction. mRNAs of the four reprogramming factors were transfected into GEFs, and were localized in nucleus with approximately 90% transfection efficiency. After five consecutive transfections, GEFs tended to aggregate by day 10. Clones appeared on day 15-18, and typical embryonic stem cell -like clones formed on day 20. One thousand AKP staining positive clones were achieved in 10(4) GEFs, with approximately 1.0% induction efficiency. Immunofluorescence staining and qRT-PCR detection of the ESCs markers confirmed the properties of the goat iPSCs. The achieved goat iPSCs could be cultured to 22nd passage, which showed normal karyotype. The goat iPSCs were able to differentiate into embryoid bodies with three germ layers. qRT-PCR and western blot showed activated endogenous pluripotent factors expression in the later phase of mRNA-induced goat iPSCs induction. Epigenetic analysis of the endogenous pluripotent gene Nanog revealed its demethylation status in derived goat iPSCs. Core promoter regions of the four reprogramming factors were determined. Transcription factor binding sites, including Elf-1, AP-2, SP1, C/EBP and MZF1, were identified to be functional in the core promoter regions of these reprogramming genes. Demethylation and deacetylation of the promoters enhanced their transcription activities. We successfully generated goat iPSCs by transfection of Oct4, Sox2, Klf4 and c-Myc mRNAs into GEFs, which initiated the endogenous reprogramming network and altered the methylation status of pluripotent genes. Core promoter regions and functional transcription binding sites of

  1. Characterization of the regulatory mechanisms of activating transcription factor 3 by hypertrophic stimuli in rat cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Elina Koivisto

    Full Text Available AIMS: Activating transcription factor 3 (ATF3 is a stress-activated immediate early gene suggested to have both detrimental and cardioprotective role in the heart. Here we studied the mechanisms of ATF3 activation by hypertrophic stimuli and ATF3 downstream targets in rat cardiomyocytes. METHODS AND RESULTS: When neonatal rat cardiomyocytes were exposed to endothelin-1 (ET-1, 100 nM and mechanical stretching in vitro, maximal increase in ATF3 expression occurred at 1 hour. Inhibition of extracellular signal-regulated kinase (ERK by PD98059 decreased ET-1- and stretch-induced increase of ATF3 protein but not ATF3 mRNA levels, whereas protein kinase A (PKA inhibitor H89 attenuated both ATF3 mRNA transcription and protein expression in response to ET-1 and stretch. To characterize further the regulatory mechanisms upstream of ATF3, p38 mitogen-activated protein kinase (MAPK signaling was investigated using a gain-of-function approach. Adenoviral overexpression of p38α, but not p38β, increased ATF3 mRNA and protein levels as well as DNA binding activity. To investigate the role of ATF3 in hypertrophic process, we overexpressed ATF3 by adenovirus-mediated gene transfer. In vitro, ATF3 gene delivery attenuated the mRNA transcription of interleukin-6 (IL-6 and plasminogen activator inhibitor-1 (PAI-1, and enhanced nuclear factor-κB (NF-κB and Nkx-2.5 DNA binding activities. Reduced PAI-1 expression was also detected in vivo in adult rat heart by direct intramyocardial adenovirus-mediated ATF3 gene delivery. CONCLUSIONS: These data demonstrate that ATF3 activation by ET-1 and mechanical stretch is partly mediated through ERK and cAMP-PKA pathways, whereas p38 MAPK pathway is involved in ATF3 activation exclusively through p38α isoform. ATF3 activation caused induction of modulators of the inflammatory response NF-κB and Nkx-2.5, as well as attenuation of pro-fibrotic and pro-inflammatory proteins IL-6 and PAI-1, suggesting cardioprotective role

  2. The transcription factor encyclopedia.

    Science.gov (United States)

    Yusuf, Dimas; Butland, Stefanie L; Swanson, Magdalena I; Bolotin, Eugene; Ticoll, Amy; Cheung, Warren A; Zhang, Xiao Yu Cindy; Dickman, Christopher T D; Fulton, Debra L; Lim, Jonathan S; Schnabl, Jake M; Ramos, Oscar H P; Vasseur-Cognet, Mireille; de Leeuw, Charles N; Simpson, Elizabeth M; Ryffel, Gerhart U; Lam, Eric W-F; Kist, Ralf; Wilson, Miranda S C; Marco-Ferreres, Raquel; Brosens, Jan J; Beccari, Leonardo L; Bovolenta, Paola; Benayoun, Bérénice A; Monteiro, Lara J; Schwenen, Helma D C; Grontved, Lars; Wederell, Elizabeth; Mandrup, Susanne; Veitia, Reiner A; Chakravarthy, Harini; Hoodless, Pamela A; Mancarelli, M Michela; Torbett, Bruce E; Banham, Alison H; Reddy, Sekhar P; Cullum, Rebecca L; Liedtke, Michaela; Tschan, Mario P; Vaz, Michelle; Rizzino, Angie; Zannini, Mariastella; Frietze, Seth; Farnham, Peggy J; Eijkelenboom, Astrid; Brown, Philip J; Laperrière, David; Leprince, Dominique; de Cristofaro, Tiziana; Prince, Kelly L; Putker, Marrit; del Peso, Luis; Camenisch, Gieri; Wenger, Roland H; Mikula, Michal; Rozendaal, Marieke; Mader, Sylvie; Ostrowski, Jerzy; Rhodes, Simon J; Van Rechem, Capucine; Boulay, Gaylor; Olechnowicz, Sam W Z; Breslin, Mary B; Lan, Michael S; Nanan, Kyster K; Wegner, Michael; Hou, Juan; Mullen, Rachel D; Colvin, Stephanie C; Noy, Peter John; Webb, Carol F; Witek, Matthew E; Ferrell, Scott; Daniel, Juliet M; Park, Jason; Waldman, Scott A; Peet, Daniel J; Taggart, Michael; Jayaraman, Padma-Sheela; Karrich, Julien J; Blom, Bianca; Vesuna, Farhad; O'Geen, Henriette; Sun, Yunfu; Gronostajski, Richard M; Woodcroft, Mark W; Hough, Margaret R; Chen, Edwin; Europe-Finner, G Nicholas; Karolczak-Bayatti, Magdalena; Bailey, Jarrod; Hankinson, Oliver; Raman, Venu; LeBrun, David P; Biswal, Shyam; Harvey, Christopher J; DeBruyne, Jason P; Hogenesch, John B; Hevner, Robert F; Héligon, Christophe; Luo, Xin M; Blank, Marissa Cathleen; Millen, Kathleen Joyce; Sharlin, David S; Forrest, Douglas; Dahlman-Wright, Karin; Zhao, Chunyan; Mishima, Yuriko; Sinha, Satrajit; Chakrabarti, Rumela; Portales-Casamar, Elodie; Sladek, Frances M; Bradley, Philip H; Wasserman, Wyeth W

    2012-01-01

    Here we present the Transcription Factor Encyclopedia (TFe), a new web-based compendium of mini review articles on transcription factors (TFs) that is founded on the principles of open access and collaboration. Our consortium of over 100 researchers has collectively contributed over 130 mini review articles on pertinent human, mouse and rat TFs. Notable features of the TFe website include a high-quality PDF generator and web API for programmatic data retrieval. TFe aims to rapidly educate scientists about the TFs they encounter through the delivery of succinct summaries written and vetted by experts in the field. TFe is available at http://www.cisreg.ca/tfe.

  3. The Transcription Factor Encyclopedia

    DEFF Research Database (Denmark)

    Yusuf, Dimas; Butland, Stefanie L; Swanson, Magdalena I

    2012-01-01

    mini review articles on pertinent human, mouse and rat TFs. Notable features of the TFe website include a high-quality PDF generator and web API for programmatic data retrieval. TFe aims to rapidly educate scientists about the TFs they encounter through the delivery of succinct summaries written......ABSTRACT: Here we present the Transcription Factor Encyclopedia (TFe), a new web-based compendium of mini review articles on transcription factors (TFs) that is founded on the principles of open access and collaboration. Our consortium of over 100 researchers has collectively contributed over 130...

  4. A modified reverse one-hybrid screen identifies transcriptional activation in Phyochrome-Interacting Factor 3

    Science.gov (United States)

    Transcriptional activation domains (TAD) are difficult to predict and identify, since they are not conserved and have little consensus. Here, we describe a yeast-based screening method that is able to identify individual amino acid residues involved in transcriptional activation in a high throughput...

  5. 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.

  6. The oncoprotein HBXIP upregulates PDGFB via activating transcription factor Sp1 to promote the proliferation of breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Yingyi; Zhao, Yu; Li, Leilei; Shen, Yu; Cai, Xiaoli [Department of Biochemistry, College of Life Sciences, Nankai University, Tianjin 300071 (China); Zhang, Xiaodong, E-mail: zhangxd@nankai.edu.cn [Department of Cancer Research, Institute for Molecular Biology, College of Life Sciences, Nankai University, Tianjin 300071 (China); Ye, Lihong, E-mail: yelihong@nankai.edu.cn [Department of Biochemistry, College of Life Sciences, Nankai University, Tianjin 300071 (China)

    2013-05-03

    Highlights: •HBXIP is able to upregulate the expression of PDGFB in breast cancer cells. •HBXIP serves as a coactivator of activating transcription factor Sp1. •HBXIP stimulates the PDGFB promoter via activating transcription factor Sp1. •HBXIP promotes the proliferation of breast cancer cell via upregulating PDGFB. -- Abstract: We have reported that the oncoprotein hepatitis B virus X-interacting protein (HBXIP) acts as a novel transcriptional coactivator to promote proliferation and migration of breast cancer cells. Previously, we showed that HBXIP was able to activate nuclear factor-κB (NF-κB) in breast cancer cells. As an oncogene, the platelet-derived growth factor beta polypeptide (PDGFB) plays crucial roles in carcinogenesis. In the present study, we found that both HBXIP and PDGFB were highly expressed in breast cancer cell lines. Interestingly, HBXIP was able to increase transcriptional activity of NF-κB through PDGFB, suggesting that HBXIP is associated with PDGFB in the cells. Moreover, HBXIP was able to upregulate PDGFB at the levels of mRNA, protein and promoter in the cells. Then, we identified that HBXIP stimulated the promoter of PDGFB through activating transcription factor Sp1. In function, HBXIP enhanced the proliferation of breast cancer cells through PDGFB in vitro. Thus, we conclude that HBXIP upregulates PDGFB via activating transcription factor Sp1 to promote proliferation of breast cancer cells.

  7. Statistical reconstruction of transcription factor activity using Michaelis-Menten kinetics.

    Science.gov (United States)

    Khanin, R; Vinciotti, V; Mersinias, V; Smith, C P; Wit, E

    2007-09-01

    The basic building block of a gene regulatory network consists of a gene encoding a transcription factor (TF) and the gene(s) it regulates. Considerable efforts have been directed recently at devising experiments and algorithms to determine TFs and their corresponding target genes using gene expression and other types of data. The underlying problem is that the expression of a gene coding for the TF provides only limited information about the activity of the TF, which can also be controlled posttranscriptionally. In the absence of a reliable technology to routinely measure the activity of regulators, it is of great importance to understand whether this activity can be inferred from gene expression data. We here develop a statistical framework to reconstruct the activity of a TF from gene expression data of the target genes in its regulatory module. The novelty of our approach is that we embed the deterministic Michaelis-Menten model of gene regulation in this statistical framework. The kinetic parameters of the gene regulation model are inferred together with the profile of the TF regulator. We also obtain a goodness-of-fit test to verify the fit of the model. The model is applied to a time series involving the Streptomyces coelicolor bacterium. We focus on the transcriptional activator cdaR, which is partly responsible for the production of a particular type of antibiotic. The aim is to reconstruct the activity profile of this regulator. Our approach can be extended to include more complex regulatory relationships, such as multiple regulatory factors, competition, and cooperativity.

  8. Reinstate the Damaged VEGF Signaling Pathway with VEGF-activating Transcription Factor

    Institute of Scientific and Technical Information of China (English)

    Yao-guo Yang; Heng Guan; Chang-wei Liu; Yong-jun Li

    2009-01-01

    Objective To investigate the role of vascular endothelial growth factor-activating transcriptional factor(VEGF-ATF)on the VEGF signaling pathway in diabetes mellitus.Methods Totally,20 C57BL/6 mice fed with high fat diet was induced into diabetes mellitus.Ten diabetes mellitus mice received a lower limb muscle injection with VEGF-ATF plasmid,and another ten were as control.VEGF-ATF is an engineered transcription factor designed to increase VEGF expression.Three days later,mice were sacrificed and the injected gastrocnemius was used for analysis.VEGF mRNA and protein expressions were examined by real-time PCR and ELISA respectively.VEGF receptor 2 mRNA expression was tested with RT-PCR.Phosphorylated Akt,Akt,endothelial nitric oxide synthase(eNOS),and phosphorylated eNOS were assessed by western blot.Results At 3 days post-injection,in mice with diabetes mellitus,VEGF gene transfer increased VEGF mRNA copies and VEGF protein expression in injected muscles compared with control;and reinstated the impaired VEGF signaling pathway with increasing the ratios of phosphorylated Akt/Akt and phosphorylated eNOS/eNOS.However,it did not affect the expression of VEGF receptor 2 mRNA.Conclusion Gene transfer with VEGF-ATF is able to reinstate the impaired VEGF downstream pathway,and potentially promote therapeutic angiogenesis in mice with diabetes mcllitus.

  9. A two-subunit bacterial sigma-factor activates transcription in Bacillus subtilis.

    Science.gov (United States)

    MacLellan, Shawn R; Guariglia-Oropeza, Veronica; Gaballa, Ahmed; Helmann, John D

    2009-12-15

    The sigma-like factor YvrI and coregulator YvrHa activate transcription from a small set of conserved promoters in Bacillus subtilis. We report here that these two proteins independently contribute sigma-region 2 and sigma-region 4 functions to a holoenzyme-promoter DNA complex. YvrI binds RNA polymerase (RNAP) through a region 4 interaction with the beta-subunit flap domain and mediates specific promoter recognition but cannot initiate DNA melting at the -10 promoter element. Conversely, YvrHa possesses sequence similarity to a conserved core-binding motif in sigma-region 2 and binds to the N-terminal coiled-coil element in the RNAP beta'-subunit previously implicated in interaction with region 2 of sigma-factors. YvrHa plays an essential role in stabilizing the open complex and interacts specifically with the N-terminus of YvrI. Based on these results, we propose that YvrHa is situated in the transcription complex proximal to the -10 element of the promoter, whereas YvrI is responsible for -35 region recognition. This system presents an unusual example of a two-subunit bacterial sigma-factor.

  10. Expression and activity of SNAIL transcription factor during Epithelial to Mesenchymal Transition (EMT) in cancer progression.

    Science.gov (United States)

    Papiewska-Pająk, Izabela; Kowalska, Maria A; Boncela, Joanna

    2016-09-19

    Inhibition of E-cadherin gene expression by transcription factor SNAIL is known to be a crucial element of Epithelial to Mesenchymal Transition; EMT. Epigenetic regulation of E-cadherin expression is regulated by SNAIL binding to E-box sequences in the CDH1 gene promoter and recruiting enzymes belonging to repressor complexes that are directly engaged in histone modifications and DNA methylation leading to the modification of chromatin structure. SNAIL involvement in cell acquisition of invasive phenotype is based on direct suppression of tight-junction and gap junction proteins. The nuclear localization of SNAIL is required for SNAIL activity and protects this factor from proteasomal degradation in the cytoplasm. The main factor engaged in that process is GSK- 3β kinase. Expression and stability of SNAIL is regulated on the transctriptional and posttranscriptional levels by a number of signaling molecules and biological factors, for example: TGF-β, TNF-α, ILK and NFκB. The expression of SNAIL in cancer cells is also regulated by micro-RNA, mainly by miR-34. Increased expression of SNAIL, observed in many human cancers, has been correlated with increased resistance to chemio-, radio - or immunotherapy, gain of cancer stem cells features and migrative and invasive characteristics, which leads to tumor metastases. Understanding of the SNAIL's mechanism of action may lead to new treatment strategies in cancer directed to interfere with signaling pathways that either activate SNAIL or are activated by SNAIL.

  11. hypoxia-inducible factors activate CD133 promoter through ETS family transcription factors.

    Directory of Open Access Journals (Sweden)

    Shunsuke Ohnishi

    Full Text Available CD133 is a cellular surface protein that has been reported to be a cancer stem cell marker, and thus it is considered to be a potential target for cancer treatment. However, the mechanism regulating CD133 expression is not yet understood. In this study, we analyzed the activity of five putative promoters (P1-P5 of CD133 in human embryonic kidney (HEK 293 cells and colon cancer cell line WiDr, and found that the activity of promoters, particularly of P5, is elevated by overexpression of hypoxia-inducible factors (HIF-1α and HIF-2α. Deletion and mutation analysis identified one of the two E-twenty six (ETS binding sites (EBSs in the P5 region as being essential for its promoter activity induced by HIF-1α and HIF-2α. In addition, a chromatin imunoprecipitation assay demonstrated that HIF-1α and HIF-2α bind to the proximal P5 promoter at the EBSs. The immunoprecipitation assay showed that HIF-1α physically interacts with Elk1; however, HIF-2α did not bind to Elk1 or ETS1. Furthermore, knockdown of both HIF-1α and HIF-2α resulted in a reduction of CD133 expression in WiDr. Taken together, our results revealed that HIF-1α and HIF-2α activate CD133 promoter through ETS proteins.

  12. Hypoxia-Inducible Factor 1 Is an Inductor of Transcription Factor Activating Protein 2 Epsilon Expression during Chondrogenic Differentiation

    Directory of Open Access Journals (Sweden)

    Stephan Niebler

    2015-01-01

    Full Text Available The transcription factor AP-2ε (activating enhancer-binding protein epsilon is expressed in cartilage of humans and mice. However, knowledge about regulatory mechanisms influencing AP-2ε expression is limited. Using quantitative real time PCR, we detected a significant increase in AP-2ε mRNA expression comparing initial and late stages of chondrogenic differentiation processes in vitro and in vivo. Interestingly, in these samples the expression pattern of the prominent hypoxia marker gene angiopoietin-like 4 (Angptl4 strongly correlated with that of AP-2ε suggesting that hypoxia might represent an external regulator of AP-2ε expression in mammals. In order to show this, experiments directly targeting the activity of hypoxia-inducible factor-1 (HIF1, the complex mediating responses to oxygen deprivation, were performed. While the HIF1-activating compounds 2,2′-dipyridyl and desferrioxamine resulted in significantly enhanced mRNA concentration of AP-2ε, siRNA against HIF1α led to a significantly reduced expression rate of AP-2ε. Additionally, we detected a significant upregulation of the AP-2ε mRNA level after oxygen deprivation. In sum, these different experimental approaches revealed a novel role for the HIF1 complex in the regulation of the AP-2ε gene in cartilaginous cells and underlined the important role of hypoxia as an important external regulatory stimulus during chondrogenic differentiation modulating the expression of downstream transcription factors.

  13. Nε-lysine acetylation of a bacterial transcription factor inhibits Its DNA-binding activity.

    Directory of Open Access Journals (Sweden)

    Sandy Thao

    Full Text Available Evidence suggesting that eukaryotes and archaea use reversible N(ε-lysine (N(ε-Lys acetylation to modulate gene expression has been reported, but evidence for bacterial use of N(ε-Lys acetylation for this purpose is lacking. Here, we report data in support of the notion that bacteria can control gene expression by modulating the acetylation state of transcription factors (TFs. We screened the E. coli proteome for substrates of the bacterial Gcn5-like protein acetyltransferase (Pat. Pat acetylated four TFs, including the RcsB global regulatory protein, which controls cell division, and capsule and flagellum biosynthesis in many bacteria. Pat acetylated residue Lys180 of RcsB, and the NAD(+-dependent Sir2 (sirtuin-like protein deacetylase (CobB deacetylated acetylated RcsB (RcsB(Ac, demonstrating that N(ε-Lys acetylation of RcsB is reversible. Analysis of RcsB(Ac and variant RcsB proteins carrying substitutions at Lys180 provided biochemical and physiological evidence implicating Lys180 as a critical residue for RcsB DNA-binding activity. These findings further the likelihood that reversible N(ε-Lys acetylation of transcription factors is a mode of regulation of gene expression used by all cells.

  14. A Global Genomic and Genetic Strategy to Predict Pathway Activation of Xenobiotic Responsive Transcription Factors in the Mouse Liver

    Science.gov (United States)

    Many drugs and environmentally-relevant chemicals activate xenobiotic-responsive transcription factors(TF). Identification of target genes of these factors would be useful in predicting pathway activation in in vitro chemical screening. Starting with a large compendium of Affymet...

  15. 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.

  16. Activating transcription factor 1 directs Mhem atheroprotective macrophages through coordinated iron handling and foam cell protection.

    Science.gov (United States)

    Boyle, Joseph J; Johns, Michael; Kampfer, Theresa; Nguyen, Aivi T; Game, Laurence; Schaer, Dominik J; Mason, Justin C; Haskard, Dorian O

    2012-01-06

    Intraplaque hemorrhage (IPH) drives atherosclerosis through the dual metabolic stresses of cholesterol-enriched erythrocyte membranes and pro-oxidant heme/iron. When clearing tissue hemorrhage, macrophages are typically seen storing either iron or lipid. We have recently defined hemorrhage-associated macrophages (HA-mac) as a plaque macrophage population that responds adaptively to IPH. This study aimed to define the key transcription factor(s) involved in HO-1 induction by heme. To address this question, we used microarray analysis and transfection with siRNA and plasmids. To maintain physiological relevance, we focused on human blood-derived monocytes. We found that heme stimulates monocytes through induction of activating transcription factor 1 (ATF-1). ATF-1 coinduces heme oxygenase-1 (HO-1) and Liver X receptor beta (LXR-β). Heme-induced HO-1 and LXR-β were suppressed by knockdown of ATF-1, and HO-1 and LXR-β were induced by ATF-1 transfection. ATF-1 required phosphorylation for full functional activity. Expression of LXR-β in turn led to induction of other genes central to cholesterol efflux, such as LXR-α and ABCA1. This heme-directed state was distinct from known macrophage states (M1, M2, Mox) and, following the same format, we have designated them Mhem. These results show that ATF-1 mediates HO-1 induction by heme and drives macrophage adaptation to intraplaque hemorrhage. Our definition of an ATF-1-mediated pathway for linked protection from foam cell formation and oxidant stress may have therapeutic potential.

  17. 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.

  18. Dynamic, large-scale profiling of transcription factor activity from live cells in 3D culture.

    Directory of Open Access Journals (Sweden)

    Michael S Weiss

    Full Text Available BACKGROUND: Extracellular activation of signal transduction pathways and their downstream target transcription factors (TFs are critical regulators of cellular processes and tissue development. The intracellular signaling network is complex, and techniques that quantify the activities of numerous pathways and connect their activities to the resulting phenotype would identify the signals and mechanisms regulating tissue development. The ability to investigate tissue development should capture the dynamic pathway activity and requires an environment that supports cellular organization into structures that mimic in vivo phenotypes. Taken together, our objective was to develop cellular arrays for dynamic, large-scale quantification of TF activity as cells organized into spherical structures within 3D culture. METHODOLOGY/PRINCIPAL FINDINGS: TF-specific and normalization reporter constructs were delivered in parallel to a cellular array containing a well-established breast cancer cell line cultured in Matrigel. Bioluminescence imaging provided a rapid, non-invasive, and sensitive method to quantify luciferase levels, and was applied repeatedly on each sample to monitor dynamic activity. Arrays measuring 28 TFs identified up to 19 active, with 13 factors changing significantly over time. Stimulation of cells with β-estradiol or activin A resulted in differential TF activity profiles evolving from initial stimulation of the ligand. Many TFs changed as expected based on previous reports, yet arrays were able to replicate these results in a single experiment. Additionally, arrays identified TFs that had not previously been linked with activin A. CONCLUSIONS/SIGNIFICANCE: This system provides a method for large-scale, non-invasive, and dynamic quantification of signaling pathway activity as cells organize into structures. The arrays may find utility for investigating mechanisms regulating normal and abnormal tissue growth, biomaterial design, or as a

  19. 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.

  20. A Nonradioactive Method for Detecting DNA-binding Activity of Nuclear Transcription Factors

    Institute of Scientific and Technical Information of China (English)

    张宁; 徐永健; 张珍祥; 熊维宁

    2003-01-01

    To determine the feasibility of a nonradioactive electrophoresis mobility shift assay fordetecting nuclear transcription factor, double-stranded oligonucleotides encoding the consensus tar-get sequence of NF-κB were labled with DIG by terminal transferase. After nuclear protein stimula-ted with phorbol 12-myristate 13-acetate (PMA) or PMA and pyrrolidine dithiocarbamate (PDT C)electrophoresed on 8 % nondenaturing poliacrylamide gel together with oligeonucleotide probe, theywere electro-blotted nylon membrane positively charged. Anti-DIG-AP antibody catalyzed chemilu-minescent substrate CSPD to image on X-film. The results showed that nuclear proteins binded spe-cifically to the NF-κB consensus sequence in the EMSA by chemiluminescent technique method andthe activity of NF-κB in PMA group was more than that in PMA+PDTC group. It is suggestedthat detection of NF-κB by EMSA with chemiluminescent technique is feasible and simple, whichcan be performed in ordinary laboratories.

  1. The adaptor protein MITA links virus-sensing receptors to IRF3 transcription factor activation.

    Science.gov (United States)

    Zhong, Bo; Yang, Yan; Li, Shu; Wang, Yan-Yi; Li, Ying; Diao, Feici; Lei, Caoqi; He, Xiao; Zhang, Lu; Tien, Po; Shu, Hong-Bing

    2008-10-17

    Viral infection triggers activation of transcription factors such as NF-kappaB and IRF3, which collaborate to induce type I interferons (IFNs) and elicit innate antiviral response. Here, we identified MITA as a critical mediator of virus-triggered type I IFN signaling by expression cloning. Overexpression of MITA activated IRF3, whereas knockdown of MITA inhibited virus-triggered activation of IRF3, expression of type I IFNs, and cellular antiviral response. MITA was found to localize to the outer membrane of mitochondria and to be associated with VISA, a mitochondrial protein that acts as an adaptor in virus-triggered signaling. MITA also interacted with IRF3 and recruited the kinase TBK1 to the VISA-associated complex. MITA was phosphorylated by TBK1, which is required for MITA-mediated activation of IRF3. Our results suggest that MITA is a critical mediator of virus-triggered IRF3 activation and IFN expression and further demonstrate the importance of certain mitochondrial proteins in innate antiviral immunity.

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

    Directory of Open Access Journals (Sweden)

    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

  3. Interaction with general transcription factor IIF (TFIIF) is required for the suppression of activated transcription by RPB5-mediating protein(RMP)

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    RMP was reported to regulate transcription via competing with HBx to bind the general transcription factor IIB (TFIIB) and interacting with RPB5 subunit of RNA polymerase Ⅱ as a corepressor of transcription regulator. However, our present research uncovered that RMP also regulates the transcription through interaction with the general transcription factors IIF (TFIIF), which assemble in the preinitiation complex and function in both transcription initiation and elongation. With in vitro pull-down assay and Far-Western analysis, we demonstrated that RMP could bind with bacterially expressed recombinant RAP30 and RAP74of TFIIF subunits. In the immunoprecipitation assay in COS 1 cells cotransfected with FLAG-tagged RMP or its mutants, GST-fused RAP30 and RAP74 were co-immunoprecipitated with RMP in approximately equal molar ratio, which suggests that RAP30 and RAP74 interact with RMP as a TFIIF complex. Interestingly both RAP30 and RAP74 interact with the same domain (D5) of the C-terminal RMP of 118-amino-acid residuals which overlaps with its TFIIB-binding domain. Internal deletion of D5 region of RMP abolished its binding ability with both subunits of TFIIF, while D5 domain alone was sufficient to interact with TFIIF subunits. The result of luciferase assay showed that overexpression of RMP, but not the mutant RMP lacking D5 region, suppressed the transcription activated by Gal-VP16, suggesting that interaction with TFIIF is required for RMP to suppress the activated transcription. The interaction between RMP and TFIIF may be an additional passway for RMP to regulate the transcription, or alternatively TFIIF may cooperate with RPB5 and TFIIB for the corepressor function of RMP.

  4. Transcription factor-based biosensor

    Science.gov (United States)

    Dietrich, Jeffrey A; Keasling, Jay D

    2013-10-08

    The present invention provides for a system comprising a BmoR transcription factor, a .sigma..sup.54-RNA polymerase, and a pBMO promoter operatively linked to a reporter gene, wherein the pBMO promoter is capable of expression of the reporter gene with an activated form of the BmoR and the .sigma..sup.54-RNA polymerase.

  5. Bromodomain and extra-terminal (BET) bromodomain inhibition activate transcription via transient release of positive transcription elongation factor b (P-TEFb) from 7SK small nuclear ribonucleoprotein.

    Science.gov (United States)

    Bartholomeeusen, Koen; Xiang, Yanhui; Fujinaga, Koh; Peterlin, B Matija

    2012-10-19

    By phosphorylating elongation factors and the C-terminal domain of RNA polymerase II, the positive transcription elongation factor b (P-TEFb) is the critical kinase for transcription elongation and co-transcriptional processing of eukaryotic genes. It exists in inactive small nuclear ribonucleoprotein (7SK snRNP) and active (free P-TEFb) complexes in cells. The P-TEFb equilibrium determines the state of cellular activation, proliferation, and differentiation. Free P-TEFb, which is required for growth, can be recruited to RNA polymerase II via transcription factors, BRD4, or the super elongation complex (SEC). UV light, various signaling cascades, transcriptional blockade, or compounds such as hexamethylene bisacetamide (HMBA), suberoylanilide hydroxamic acid (SAHA), and other histone deacetylase inhibitors lead to a rapid release of free P-TEFb, followed by its reassembly into the 7SK snRNP. As a consequence, transcription of HEXIM1, a critical 7SK snRNP subunit, and HIV is induced. In this study, we found that a bromodomain and extra-terminal (BET) bromodomain inhibitor, JQ1, which inhibits BRD4 by blocking its association with chromatin, also leads to the rapid release of free P-TEFb from the 7SK snRNP. Indeed, JQ1 transiently increased levels of free P-TEFb and BRD4·P-TEFb and SEC·P-TEFb complexes in cells. As a consequence, the levels of HEXIM1 and HIV proteins rose. Importantly, the knockdown of ELL2, a subunit of the SEC, blocked the ability of JQ1 to increase HIV transcription. Finally, the effects of JQ1 and HMBA or SAHA on the P-TEFb equilibrium were cooperative. We conclude that HMBA, SAHA, and JQ1 affect transcription elongation by a similar and convergent mechanism.

  6. RAI1 transcription factor activity is impaired in mutants associated with Smith-Magenis Syndrome.

    Directory of Open Access Journals (Sweden)

    Paulina Carmona-Mora

    Full Text Available Smith-Magenis Syndrome (SMS is a complex genomic disorder mostly caused by the haploinsufficiency of the Retinoic Acid Induced 1 gene (RAI1, located in the chromosomal region 17p11.2. In a subset of SMS patients, heterozygous mutations in RAI1 are found. Here we investigate the molecular properties of these mutated forms and their relationship with the resulting phenotype. We compared the clinical phenotype of SMS patients carrying a mutation in RAI1 coding region either in the N-terminal or the C-terminal half of the protein and no significant differences were found. In order to study the molecular mechanism related to these two groups of RAI1 mutations first we analyzed those mutations that result in the truncated protein corresponding to the N-terminal half of RAI1 finding that they have cytoplasmic localization (in contrast to full length RAI1 and no ability to activate the transcription through an endogenous target: the BDNF enhancer. Similar results were found in lymphoblastoid cells derived from a SMS patient carrying RAI1 c.3103insC, where both mutant and wild type products of RAI1 were detected. The wild type form of RAI1 was found in the chromatin bound and nuclear matrix subcellular fractions while the mutant product was mainly cytoplasmic. In addition, missense mutations at the C-terminal half of RAI1 presented a correct nuclear localization but no activation of the endogenous target. Our results showed for the first time a correlation between RAI1 mutations and abnormal protein function plus they suggest that a reduction of total RAI1 transcription factor activity is at the heart of the SMS clinical presentation.

  7. RAI1 transcription factor activity is impaired in mutants associated with Smith-Magenis Syndrome.

    Science.gov (United States)

    Carmona-Mora, Paulina; Canales, Cesar P; Cao, Lei; Perez, Irene C; Srivastava, Anand K; Young, Juan I; Walz, Katherina

    2012-01-01

    Smith-Magenis Syndrome (SMS) is a complex genomic disorder mostly caused by the haploinsufficiency of the Retinoic Acid Induced 1 gene (RAI1), located in the chromosomal region 17p11.2. In a subset of SMS patients, heterozygous mutations in RAI1 are found. Here we investigate the molecular properties of these mutated forms and their relationship with the resulting phenotype. We compared the clinical phenotype of SMS patients carrying a mutation in RAI1 coding region either in the N-terminal or the C-terminal half of the protein and no significant differences were found. In order to study the molecular mechanism related to these two groups of RAI1 mutations first we analyzed those mutations that result in the truncated protein corresponding to the N-terminal half of RAI1 finding that they have cytoplasmic localization (in contrast to full length RAI1) and no ability to activate the transcription through an endogenous target: the BDNF enhancer. Similar results were found in lymphoblastoid cells derived from a SMS patient carrying RAI1 c.3103insC, where both mutant and wild type products of RAI1 were detected. The wild type form of RAI1 was found in the chromatin bound and nuclear matrix subcellular fractions while the mutant product was mainly cytoplasmic. In addition, missense mutations at the C-terminal half of RAI1 presented a correct nuclear localization but no activation of the endogenous target. Our results showed for the first time a correlation between RAI1 mutations and abnormal protein function plus they suggest that a reduction of total RAI1 transcription factor activity is at the heart of the SMS clinical presentation.

  8. In Vitro Anticancer Activity of Phlorofucofuroeckol A via Upregulation of Activating Transcription Factor 3 against Human Colorectal Cancer Cells

    Directory of Open Access Journals (Sweden)

    Hyun Ji Eo

    2016-03-01

    Full Text Available Phlorofucofuroeckol A (PFF-A, one of the phlorotannins found in brown algae, has been reported to exert anti-cancer property. However, the molecular mechanism for the anti-cancer effect of PFF-A has not been known. Activating transcription factor 3 (ATF3 has been reported to be associated with apoptosis in colorectal cancer. The present study was performed to investigate the molecular mechanism by which PFF-A stimulates ATF3 expression and apoptosis in human colorectal cancer cells. PFF-A decreased cell viability through apoptosis of human colorectal cancer cells. PFF-A increased ATF3 expression through regulating transcriptional activity. The responsible cis-element for ATF3 transcriptional activation by PFF-A was cAMP response element binding protein (CREB, located between positions −147 and −85 of the ATF3 promoter. Inhibition of p38, c-Jun N-terminal kinases (JNK, glycogen synthase kinase (GSK 3β, and IκB kinase (IKK-α blocked PFF-A-mediated ATF3 expression. ATF3 knockdown by ATF3 siRNA attenuated the cleavage of poly (ADP-ribose polymerase (PARP by PFF-A, while ATF3 overexpression increased PFF-A-mediated cleaved PARP. These results suggest that PFF-A may exert anti-cancer property through inducing apoptosis via the ATF3-mediated pathway in human colorectal cancer cells.

  9. Nuclear factor-κB p65 (RelA) transcription factor is constitutively activated in human colorectal carcinoma tissue

    Institute of Scientific and Technical Information of China (English)

    Liang-Liang Yu; Hong-Gang Yu; Jie-Ping Yu; He-Sheng Luo; Xi-Ming Xu; Jun-Hua Li

    2004-01-01

    AIM: Activation of transcription factor nuclear factor-κB (NF-κB) has been shown to play a role in cell proliferation,apoptosis, cytokine production, and oncogenesis. The purpose of this study was to determine whether NF-κB was constitutively activated in human colorectal tumor tissues and, if so, to determine the role of NF-κB in colorectal tumorigenesis, and furthermore, to determine the association of RelA expression with tumor cell apoptosis and the expression of Bcl-2 and Bcl-xL.METHODS: Paraffin sections of normal epithelial, adenomatous and adenocarcinoma tissues were analysed immunohistochemically for expression of RelA, Bcl-2 and Bcl-xL proteins.Electrophoretic mobility shift assay (EMSA) was used to confirm the increased nuclear translocation of RelA in colorectal tumor tissues. The mRNA expressions of Bcl-2 and Bcl-xL were determined by reverse transcription polymerase chain reaction (RT-PCR) analysis. Apoptotic cells were detected by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate fluorescence nick end labeling (TUNEL) method.RESULTS: The activity of NF-κB was significantly higher in adenocarcinoma tissue in comparison with that in adenomatous and normal epithelial tissues. The apoptotic index (AI)significantly decreased in the transition from adenoma to adenocarcinoma. Meanwhile, the expressions of Bcl-2 and Bcl-xL protein and their mRNAs were significantly higher in adenocarcinoma tissues than that in adenomatous and normal epithelial tissues.CONCLUSION: NF-κB may inhibit apoptosis via enhancing the expression of the apoptosis genes Bcl-2 and BCl-xL. And the increased expression of RelA/nuclear factor-κB plays an important rote in the pathogenesis of colorectal carcinoma.

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

    Science.gov (United States)

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

    2015-01-01

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

  11. Promoter-proximal transcription factor binding is transcriptionally active when coupled with nucleosome repositioning in immediate vicinity

    Science.gov (United States)

    Yadav, Vinod Kumar; Thakur, Ram Krishna; Eckloff, Bruce; Baral, Aradhita; Singh, Ankita; Halder, Rashi; Kumar, Akinchan; Alam, Mohammad Parwez; Kundu, Tapas K.; Pandita, Raj; Pandita, Tej K.; Wieben, Eric D.; Chowdhury, Shantanu

    2014-01-01

    Previous studies have analyzed patterns of transcription, transcription factor (TF) binding or mapped nucleosome occupancy across the genome. These suggest that the three aspects are genetically connected but the cause and effect relationships are still unknown. For example, physiologic TF binding studies involve many TFs, consequently, it is difficult to assign nucleosome reorganization to the binding site occupancy of any particular TF. Therefore, several aspects remain unclear: does TF binding influence nucleosome (re)organizations locally or impact the chromatin landscape at a more global level; are all or only a fraction of TF binding a result of reorganization in nucleosome occupancy and do all TF binding and associated changes in nucleosome occupancy result in altered gene expression? With these in mind, following characterization of two states (before and after induction of a single TF of choice) we determined: (i) genomic binding sites of the TF, (ii) promoter nucleosome occupancy and (iii) transcriptome profiles. Results demonstrated that promoter-proximal TF binding influenced expression of the target gene when it was coupled to nucleosome repositioning at or close to its binding site in most cases. In contrast, only in few cases change in target gene expression was found when TF binding occurred without local nucleosome reorganization. PMID:25081206

  12. Number of active transcription factor binding sites is essential for the Hes7 oscillator

    Directory of Open Access Journals (Sweden)

    de Angelis Martin

    2006-02-01

    Full Text Available Abstract Background It is commonly accepted that embryonic segmentation of vertebrates is regulated by a segmentation clock, which is induced by the cycling genes Hes1 and Hes7. Their products form dimers that bind to the regulatory regions and thereby repress the transcription of their own encoding genes. An increase of the half-life of Hes7 protein causes irregular somite formation. This was shown in recent experiments by Hirata et al. In the same work, numerical simulations from a delay differential equations model, originally invented by Lewis, gave additional support. For a longer half-life of the Hes7 protein, these simulations exhibited strongly damped oscillations with, after few periods, severely attenuated the amplitudes. In these simulations, the Hill coefficient, a crucial model parameter, was set to 2 indicating that Hes7 has only one binding site in its promoter. On the other hand, Bessho et al. established three regulatory elements in the promoter region. Results We show that – with the same half life – the delay system is highly sensitive to changes in the Hill coefficient. A small increase changes the qualitative behaviour of the solutions drastically. There is sustained oscillation and hence the model can no longer explain the disruption of the segmentation clock. On the other hand, the Hill coefficient is correlated with the number of active binding sites, and with the way in which dimers bind to them. In this paper, we adopt response functions in order to estimate Hill coefficients for a variable number of active binding sites. It turns out that three active transcription factor binding sites increase the Hill coefficient by at least 20% as compared to one single active site. Conclusion Our findings lead to the following crucial dichotomy: either Hirata's model is correct for the Hes7 oscillator, in which case at most two binding sites are active in its promoter region; or at least three binding sites are active, in which

  13. The Journey of a Transcription Factor

    DEFF Research Database (Denmark)

    Pireyre, Marie

    MYBs to activate transcription of GLS biosynthetic genes. A lot is known about transcriptional regulation of these nine GLS regulators. This thesis aimed at identifying regulatory mechanisms at the protein level, allowing rapid and specific regulation of transcription factors using GLS as a model....... The general introduction and the first chapter provide background on protein level regulation and underline the importance of these mechanisms in regulating transcription factors. The remaining chapters report the identification of multiple new regulators of MYB transcription factors, potentially involved...... in their regulation at multiple steps of their activation. Plant signaling in connection with transcription factor regulation is an exciting field, allowing research on multiple regulatory mechanisms. This thesis shed light on the importance of integrating all steps of transcription factor activation in a regulatory...

  14. Role of hypoxia-inducible factor-1 in transcriptional activation of ceruloplasmin by iron deficiency

    Science.gov (United States)

    Mukhopadhyay, C. K.; Mazumder, B.; Fox, P. L.

    2000-01-01

    A role of the copper protein ceruloplasmin (Cp) in iron metabolism is suggested by its ferroxidase activity and by the tissue iron overload in hereditary Cp deficiency patients. In addition, plasma Cp increases markedly in several conditions of anemia, e.g. iron deficiency, hemorrhage, renal failure, sickle cell disease, pregnancy, and inflammation. However, little is known about the cellular and molecular mechanism(s) involved. We have reported that iron chelators increase Cp mRNA expression and protein synthesis in human hepatocarcinoma HepG2 cells. Furthermore, we have shown that the increase in Cp mRNA is due to increased rate of transcription. We here report the results of new studies designed to elucidate the molecular mechanism underlying transcriptional activation of Cp by iron deficiency. The 5'-flanking region of the Cp gene was cloned from a human genomic library. A 4774-base pair segment of the Cp promoter/enhancer driving a luciferase reporter was transfected into HepG2 or Hep3B cells. Iron deficiency or hypoxia increased luciferase activity by 5-10-fold compared with untreated cells. Examination of the sequence showed three pairs of consensus hypoxia-responsive elements (HREs). Deletion and mutation analysis showed that a single HRE was necessary and sufficient for gene activation. The involvement of hypoxia-inducible factor-1 (HIF-1) was shown by gel-shift and supershift experiments that showed HIF-1alpha and HIF-1beta binding to a radiolabeled oligonucleotide containing the Cp promoter HRE. Furthermore, iron deficiency (and hypoxia) did not activate Cp gene expression in Hepa c4 hepatoma cells deficient in HIF-1beta, as shown functionally by the inactivity of a transfected Cp promoter-luciferase construct and by the failure of HIF-1 to bind the Cp HRE in nuclear extracts from these cells. These results are consistent with in vivo findings that iron deficiency increases plasma Cp and provides a molecular mechanism that may help to understand these

  15. The osmoprotective function of the NFAT5 transcription factor in T cell development and activation.

    Science.gov (United States)

    Trama, Jason; Go, William Y; Ho, Steffan N

    2002-11-15

    The NFAT5/TonEBP transcription factor, a recently identified rel/NF-kappaB family member, activates transcription of osmocompensatory genes in response to extracellular hyperosmotic stress. However, the function of NFAT5 under isosmotic conditions present in vivo remains unknown. Here we demonstrate that NFAT5 is necessary for optimal T cell development in vivo and allows for optimal cell growth ex vivo under conditions associated with osmotic stress. Transgenic mice expressing an inhibitory form of NFAT5 in developing and mature T cells exhibited a 30% reduction in thymic cellularity evenly distributed among thymic subsets, consistent with the uniform expression and nuclear localization of NFAT5 in each subset. This was associated with a 25% reduction in peripheral CD4(+) T cells and a 50% reduction in CD8(+) T cells. While transgenic T cells exhibited no impairment in cell growth or cytokine production under normal culture conditions, impaired cell growth was observed under both hyperosmotic conditions and isosmotic conditions associated with osmotic stress. Transgenic thymocytes also demonstrated increased sensitivity to osmotic stress. Consistent with this, the system A amino acid transporter gene ATA2 exhibited NFAT5 dependence under hypertonic conditions but not in response to amino acid deprivation. Expression of the TNF-alpha gene, a putative NFAT5 target, was not altered in transgenic T cells. These results not only demonstrate an osmoprotective function for NFAT5 in primary cells but also show that NFAT5 is necessary for optimal thymic development in vivo, suggesting that developing thymocytes within the thymic microenvironment are subject to an osmotic stress that is effectively countered by NFAT5-dependent responses.

  16. Statistical Reconstruction of Transcription Factor Activity Using Michaelis–Menten Kinetics

    NARCIS (Netherlands)

    Khanin, R.; Vinciotti, V.; Mersinias, V.; Smith, C.P.; Wit, Ernst

    The basic building block of a gene regulatory network consists of a gene encoding a transcription factor (TF) and the gene(s) it regulates. Considerable efforts have been directed recently at devising experiments and algorithms to determine TFs and their corresponding target genes using gene

  17. 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 ...

  18. Statistical Reconstruction of Transcription Factor Activity Using Michaelis–Menten Kinetics

    NARCIS (Netherlands)

    Khanin, R.; Vinciotti, V.; Mersinias, V.; Smith, C.P.; Wit, Ernst

    2007-01-01

    The basic building block of a gene regulatory network consists of a gene encoding a transcription factor (TF) and the gene(s) it regulates. Considerable efforts have been directed recently at devising experiments and algorithms to determine TFs and their corresponding target genes using gene express

  19. Evaluating Transcription Factor Activity Changes by Scoring Unexplained Target Genes in Expression Data

    Science.gov (United States)

    Berchtold, Evi; Csaba, Gergely; Zimmer, Ralf

    2016-01-01

    Several methods predict activity changes of transcription factors (TFs) from a given regulatory network and measured expression data. But available gene regulatory networks are incomplete and contain many condition-dependent regulations that are not relevant for the specific expression measurement. It is not known which combination of active TFs is needed to cause a change in the expression of a target gene. A method to systematically evaluate the inferred activity changes is missing. We present such an evaluation strategy that indicates for how many target genes the observed expression changes can be explained by a given set of active TFs. To overcome the problem that the exact combination of active TFs needed to activate a gene is typically not known, we assume a gene to be explained if there exists any combination for which the predicted active TFs can possibly explain the observed change of the gene. We introduce the i-score (inconsistency score), which quantifies how many genes could not be explained by the set of activity changes of TFs. We observe that, even for these minimal requirements, published methods yield many unexplained target genes, i.e. large i-scores. This holds for all methods and all expression datasets we evaluated. We provide new optimization methods to calculate the best possible (minimal) i-score given the network and measured expression data. The evaluation of this optimized i-score on a large data compendium yields many unexplained target genes for almost every case. This indicates that currently available regulatory networks are still far from being complete. Both the presented Act-SAT and Act-A* methods produce optimal sets of TF activity changes, which can be used to investigate the difficult interplay of expression and network data. A web server and a command line tool to calculate our i-score and to find the active TFs associated with the minimal i-score is available from https://services.bio.ifi.lmu.de/i-score. PMID:27723775

  20. Lycopene activates antioxidant enzymes and nuclear transcription factor systems in heat-stressed broilers.

    Science.gov (United States)

    Sahin, K; Orhan, C; Tuzcu, M; Sahin, N; Hayirli, A; Bilgili, S; Kucuk, O

    2016-05-01

    This study was conducted to evaluate the effects of dietary lycopene supplementation on growth performance, antioxidant status, and muscle nuclear transcription factor [Kelch like-ECH-associated protein 1 (Keap1) and (erythroid-derived 2)-like 2 (Nrf2)] expressions in broiler chickens exposed to heat stress (HS). A total of 180 one-day-old male broiler chicks (Ross 308) were assigned randomly to one of 2×3 factorially arranged treatments: two housing temperatures (22°C for 24 h/d; thermoneutral, TN or 34°C for 8 h/d HS) and three dietary lycopene levels (0, 200, or 400 mg/kg). Each treatment consisted of three replicates of 10 birds. Birds were reared to 42 d of age. Heat stress caused reductions in feed intake and weight gain by 12.2 and 20.7% and increased feed efficiency by 10.8% (Plycopene level improved performance in both environments. Birds reared under the HS environment had lower serum and muscle lycopene concentration (0.34 vs. 0.50 μg/mL and 2.80 vs. 2.13 μg/g), activities of superoxide dismutase (151 vs. 126 U/mL and 131 vs. 155 U/mg protein), glutathione peroxidase (184 vs. 154 U/mL and 1.39 vs. 1.74 U/mg protein), and higher malondialdehyde (MDA) concentration (0.53 vs. 0.83 μg/mL and 0.78 vs. 0.45 μg/ mg protein) than birds reared under the TN environment. Changes in levels of lycopene and MDA and activities of enzymes in serum and muscle varied by the environmental temperature as dietary lycopene level increased. Moreover, increasing dietary lycopene level suppressed muscle Keap1 expression and enhanced muscle Nrf2 expression, which had increased by 150% and decreased by 40%, respectively in response to HS. In conclusion, lycopene supplementation alleviates adverse effects of HS on performance through modulating expressions of stress-related nuclear transcription factors.

  1. Heterogeneous nuclear ribonucleoprotein K inhibits heat shock-induced transcriptional activity of heat shock factor 1.

    Science.gov (United States)

    Kim, Hee-Jung; Lee, Jae-Jin; Cho, Jin-Hwan; Jeong, Jaeho; Park, A Young; Kang, Wonmo; Lee, Kong-Joo

    2017-08-04

    When cells are exposed to heat shock and various other stresses, heat shock factor 1 (HSF1) is activated, and the heat shock response (HSR) is elicited. To better understand the molecular regulation of the HSR, we used 2D-PAGE-based proteome analysis to screen for heat shock-induced post-translationally modified cellular proteins. Our analysis revealed that two protein spots typically present on 2D-PAGE gels and containing heterogeneous nuclear ribonucleoprotein K (hnRNP K) with trioxidized Cys(132) disappeared after the heat shock treatment and reappeared during recovery, but the total amount of hnRNP K protein remained unchanged. We next tested whether hnRNP K plays a role in HSR by regulating HSF1 and found that hnRNP K inhibits HSF1 activity, resulting in reduced expression of hsp70 and hsp27 mRNAs. hnRNP K also reduced binding affinity of HSF1 to the heat shock element by directly interacting with HSF1 but did not affect HSF1 phosphorylation-dependent activation or nuclear localization. hnRNP K lost its ability to induce these effects when its Cys(132) was substituted with Ser, Asp, or Glu. These findings suggest that hnRNP K inhibits transcriptional activity of HSF1 by inhibiting its binding to heat shock element and that the oxidation status of Cys(132) in hnRNP K is critical for this inhibition. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Transcription factors in alkaloid biosynthesis.

    Science.gov (United States)

    Yamada, Yasuyuki; Sato, Fumihiko

    2013-01-01

    Higher plants produce a large variety of low-molecular weight secondary compounds. Among them, nitrogen-containing alkaloids are the most biologically active and are often used pharmaceutically. Whereas alkaloid chemistry has been intensively investigated, alkaloid biosynthesis, including the relevant biosynthetic enzymes, genes and their regulation, and especially transcription factors, is largely unknown, as only a limited number of plant species produce certain types of alkaloids and they are difficult to study. Recently, however, several groups have succeeded in isolating the transcription factors that are involved in the biosynthesis of several types of alkaloids, including bHLH, ERF, and WRKY. Most of them show Jasmonate (JA) responsiveness, which suggests that the JA signaling cascade plays an important role in alkaloid biosynthesis. Here, we summarize the types and functions of transcription factors that have been isolated in alkaloid biosynthesis, and characterize their similarities and differences compared to those in other secondary metabolite pathways, such as phenylpropanoid and terpenoid biosyntheses. The evolution of this biosynthetic pathway and regulatory network, as well as the application of these transcription factors to metabolic engineering, is discussed.

  3. Activation of the Ig Iα1 promoter by the transcription factor Ets-1 triggers Ig Iα1-Cα1 germline transcription in epithelial cancer cells.

    Science.gov (United States)

    Duan, Zhi; Zheng, Hui; Xu, San; Jiang, Yiqun; Liu, Haidan; Li, Ming; Hu, Duosha; Li, Wei; Bode, Ann M; Dong, Zigang; Cao, Ya

    2014-03-01

    Immunoglobulins (Igs) are known to be synthesized and secreted only by B lymphocytes. Class switch recombination (CSR) is a key event that enables B cells to express Igs, and one of the crucial steps for CSR initiation is the germline transcription of Ig genes. Surprisingly, recent studies have demonstrated that the Ig genes are also expressed in some epithelial cancer cells; however, the mechanisms underlying how cancer cells initiate CSR and express Igs are still unknown. In this study, we confirmed that the Ig Iα1 promoter in cancer cell lines was activated by the Ets-1 transcription factor, and the activity of the Ig Iα1 promoter and Ig Iα1-Cα1 germline transcription were attenuated after knockdown of Ets-1 by specific small interfering RNAs (siRNA). Furthermore, the expression of Ets-1 and Igα heavy chain in cancer cells was dose dependently upregulated by TGF-β1. These results indicate that activation of the Ig Iα1 promoter by the transcription factor Ets-1 is a critical pathway and provides a novel mechanism for Ig expression in non-B cell cancers.

  4. Critical Role of Activating Transcription Factor 4 in the Anabolic Actions of Parathyroid Hormone in Bone

    Science.gov (United States)

    Yu, Shibing; Franceschi, Renny T.; Luo, Min; Fan, Jie; Jiang, Di; Cao, Huiling; Kwon, Tae-Geon; Lai, Yumei; Zhang, Jian; Patrene, Kenneth; Hankenson, Kurt; Roodman, G. David; Xiao, Guozhi

    2009-01-01

    Parathyroid hormone (PTH) is a potent anabolic agent for the treatment of osteoporosis. However, its mechanism of action in osteoblast and bone is not well understood. In this study, we show that the anabolic actions of PTH in bone are severely impaired in both growing and adult ovariectomized mice lacking bone-related activating transcription factor 4 (ATF4). Our study demonstrates that ATF4 deficiency suppresses PTH-stimulated osteoblast proliferation and survival and abolishes PTH-induced osteoblast differentiation, which, together, compromise the anabolic response. We further demonstrate that the PTH-dependent increase in osteoblast differentiation is correlated with ATF4-dependent up-regulation of Osterix. This regulation involves interactions of ATF4 with a specific enhancer sequence in the Osterix promoter. Furthermore, actions of PTH on Osterix require this same element and are associated with increased binding of ATF4 to chromatin. Taken together these experiments establish a fundamental role for ATF4 in the anabolic actions of PTH on the skeleton. PMID:19851510

  5. Critical role of activating transcription factor 4 in the anabolic actions of parathyroid hormone in bone.

    Directory of Open Access Journals (Sweden)

    Shibing Yu

    Full Text Available Parathyroid hormone (PTH is a potent anabolic agent for the treatment of osteoporosis. However, its mechanism of action in osteoblast and bone is not well understood. In this study, we show that the anabolic actions of PTH in bone are severely impaired in both growing and adult ovariectomized mice lacking bone-related activating transcription factor 4 (ATF4. Our study demonstrates that ATF4 deficiency suppresses PTH-stimulated osteoblast proliferation and survival and abolishes PTH-induced osteoblast differentiation, which, together, compromise the anabolic response. We further demonstrate that the PTH-dependent increase in osteoblast differentiation is correlated with ATF4-dependent up-regulation of Osterix. This regulation involves interactions of ATF4 with a specific enhancer sequence in the Osterix promoter. Furthermore, actions of PTH on Osterix require this same element and are associated with increased binding of ATF4 to chromatin. Taken together these experiments establish a fundamental role for ATF4 in the anabolic actions of PTH on the skeleton.

  6. 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

  7. The phytoestrogen genistein modulates lysosomal metabolism and transcription factor EB (TFEB) activation.

    Science.gov (United States)

    Moskot, Marta; Montefusco, Sandro; Jakóbkiewicz-Banecka, Joanna; Mozolewski, Paweł; Węgrzyn, Alicja; Di Bernardo, Diego; Węgrzyn, Grzegorz; Medina, Diego L; Ballabio, Andrea; Gabig-Cimińska, Magdalena

    2014-06-13

    Genistein (5,7-dihydroxy-3-(4-hydroxyphenyl)-4H-1-benzopyran-4-one) has been previously proposed as a potential drug for use in substrate reduction therapy for mucopolysaccharidoses, a group of inherited metabolic diseases caused by mutations leading to inefficient degradation of glycosaminoglycans (GAGs) in lysosomes. It was demonstrated that this isoflavone can cross the blood-brain barrier, making it an especially desirable potential drug for the treatment of neurological symptoms present in most lysosomal storage diseases. So far, no comprehensive genomic analyses have been performed to elucidate the molecular mechanisms underlying the effect elicited by genistein. Therefore, the aim of this work was to identify the genistein-modulated gene network regulating GAG biosynthesis and degradation, taking into consideration the entire lysosomal metabolism. Our analyses identified over 60 genes with known roles in lysosomal biogenesis and/or function whose expression was enhanced by genistein. Moreover, 19 genes whose products are involved in both GAG synthesis and degradation pathways were found to be remarkably differentially regulated by genistein treatment. We found a regulatory network linking genistein-mediated control of transcription factor EB (TFEB) gene expression, TFEB nuclear translocation, and activation of TFEB-dependent lysosome biogenesis to lysosomal metabolism. Our data indicate that the molecular mechanism of genistein action involves not only impairment of GAG synthesis but more importantly lysosomal enhancement via TFEB. These findings contribute to explaining the beneficial effects of genistein in lysosomal storage diseases as well as envisage new therapeutic approaches to treat these devastating diseases.

  8. Activation of the transcription factor EB rescues lysosomal abnormalities in cystinotic kidney cells.

    Science.gov (United States)

    Rega, Laura R; Polishchuk, Elena; Montefusco, Sandro; Napolitano, Gennaro; Tozzi, Giulia; Zhang, Jinzhong; Bellomo, Francesco; Taranta, Anna; Pastore, Anna; Polishchuk, Roman; Piemonte, Fiorella; Medina, Diego L; Catz, Sergio D; Ballabio, Andrea; Emma, Francesco

    2016-04-01

    Nephropathic cystinosis is a rare autosomal recessive lysosomal storage disease characterized by accumulation of cystine into lysosomes secondary to mutations in the cystine lysosomal transporter, cystinosin. The defect initially causes proximal tubular dysfunction (Fanconi syndrome) which in time progresses to end-stage renal disease. Cystinotic patients treated with the cystine-depleting agent, cysteamine, have improved life expectancy, delayed progression to chronic renal failure, but persistence of Fanconi syndrome. Here, we have investigated the role of the transcription factor EB (TFEB), a master regulator of the autophagy-lysosomal pathway, in conditionally immortalized proximal tubular epithelial cells derived from the urine of a healthy volunteer or a cystinotic patient. Lack of cystinosin reduced TFEB expression and induced TFEB nuclear translocation. Stimulation of endogenous TFEB activity by genistein, or overexpression of exogenous TFEB lowered cystine levels within 24 hours in cystinotic cells. Overexpression of TFEB also stimulated delayed endocytic cargo processing within 24 hours. Rescue of other abnormalities of the lysosomal compartment was observed but required prolonged expression of TFEB. These abnormalities could not be corrected with cysteamine. Thus, these data show that the consequences of cystinosin deficiency are not restricted to cystine accumulation and support the role of TFEB as a therapeutic target for the treatment of lysosomal storage diseases, in particular of cystinosis. Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

  9. 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.

  10. Transcription Factors Bind Thousands of Active and InactiveRegions in the Drosophila Blastoderm

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xiao-Yong; MacArthur, Stewart; Bourgon, Richard; Nix, David; Pollard, Daniel A.; Iyer, Venky N.; Hechmer, Aaron; Simirenko, Lisa; Stapleton, Mark; Luengo Hendriks, Cris L.; Chu, Hou Cheng; Ogawa, Nobuo; Inwood, William; Sementchenko, Victor; Beaton, Amy; Weiszmann, Richard; Celniker, Susan E.; Knowles, David W.; Gingeras, Tom; Speed, Terence P.; Eisen, Michael B.; Biggin, Mark D.

    2008-01-10

    Identifying the genomic regions bound by sequence-specific regulatory factors is central both to deciphering the complex DNA cis-regulatory code that controls transcription in metazoans and to determining the range of genes that shape animal morphogenesis. Here, we use whole-genome tiling arrays to map sequences bound in Drosophila melanogaster embryos by the six maternal and gap transcription factors that initiate anterior-posterior patterning. We find that these sequence-specific DNA binding proteins bind with quantitatively different specificities to highly overlapping sets of several thousand genomic regions in blastoderm embryos. Specific high- and moderate-affinity in vitro recognition sequences for each factor are enriched in bound regions. This enrichment, however, is not sufficient to explain the pattern of binding in vivo and varies in a context-dependent manner, demonstrating that higher-order rules must govern targeting of transcription factors. The more highly bound regions include all of the over forty well-characterized enhancers known to respond to these factors as well as several hundred putative new cis-regulatory modules clustered near developmental regulators and other genes with patterned expression at this stage of embryogenesis. The new targets include most of the microRNAs (miRNAs) transcribed in the blastoderm, as well as all major zygotically transcribed dorsal-ventral patterning genes, whose expression we show to be quantitatively modulated by anterior-posterior factors. In addition to these highly bound regions, there are several thousand regions that are reproducibly bound at lower levels. However, these poorly bound regions are, collectively, far more distant from genes transcribed in the blastoderm than highly bound regions; are preferentially found in protein-coding sequences; and are less conserved than highly bound regions. Together these observations suggest that many of these poorly-bound regions are not involved in early

  11. Transcription factors bind thousands of active and inactive regions in the Drosophila blastoderm.

    Directory of Open Access Journals (Sweden)

    Xiao-yong Li

    2008-02-01

    Full Text Available Identifying the genomic regions bound by sequence-specific regulatory factors is central both to deciphering the complex DNA cis-regulatory code that controls transcription in metazoans and to determining the range of genes that shape animal morphogenesis. We used whole-genome tiling arrays to map sequences bound in Drosophila melanogaster embryos by the six maternal and gap transcription factors that initiate anterior-posterior patterning. We find that these sequence-specific DNA binding proteins bind with quantitatively different specificities to highly overlapping sets of several thousand genomic regions in blastoderm embryos. Specific high- and moderate-affinity in vitro recognition sequences for each factor are enriched in bound regions. This enrichment, however, is not sufficient to explain the pattern of binding in vivo and varies in a context-dependent manner, demonstrating that higher-order rules must govern targeting of transcription factors. The more highly bound regions include all of the over 40 well-characterized enhancers known to respond to these factors as well as several hundred putative new cis-regulatory modules clustered near developmental regulators and other genes with patterned expression at this stage of embryogenesis. The new targets include most of the microRNAs (miRNAs transcribed in the blastoderm, as well as all major zygotically transcribed dorsal-ventral patterning genes, whose expression we show to be quantitatively modulated by anterior-posterior factors. In addition to these highly bound regions, there are several thousand regions that are reproducibly bound at lower levels. However, these poorly bound regions are, collectively, far more distant from genes transcribed in the blastoderm than highly bound regions; are preferentially found in protein-coding sequences; and are less conserved than highly bound regions. Together these observations suggest that many of these poorly bound regions are not involved in

  12. Profiling of histone H3 lysine 9 trimethylation levels predicts transcription factor activity and survival in acute myeloid leukemia

    DEFF Research Database (Denmark)

    Müller-Tidow, Carsten; Klein, Hans-Ulrich; Hascher, Antje

    2010-01-01

    . The altered genomic regions showed an overrepresentation of cis-binding sites for ets and c-AMP response elements (CREs) for transcription factors of the CREB/CREM/ATF1 family. The decrease in H3K9me3 levels at CREs was associated with increased CRE-driven promoter activity in AML blasts in vivo. AML...

  13. Krebs cycle function is required for activation of the Spo0A transcription factor in Bacillus subtilis.

    OpenAIRE

    Ireton, K; Jin, S.; Grossman, A D; Sonenshein, A L

    1995-01-01

    Expression of genes early during sporulation in Bacillus subtilis requires the activity of the transcription factor encoded by spo0A. The active, phosphorylated form of Spo0A is produced through the action of a multicomponent pathway, the phosphorelay. A mutant defective in the first three enzymes of the Krebs citric acid cycle was unable to express early sporulation genes, apparently because of a failure to activate the phosphorelay. Cells that produce an altered Spo0A protein that can be ph...

  14. Krebs cycle function is required for activation of the Spo0A transcription factor in Bacillus subtilis.

    OpenAIRE

    Ireton, K.; Jin, S.; Grossman, A D; Sonenshein, A L

    1995-01-01

    Expression of genes early during sporulation in Bacillus subtilis requires the activity of the transcription factor encoded by spo0A. The active, phosphorylated form of Spo0A is produced through the action of a multicomponent pathway, the phosphorelay. A mutant defective in the first three enzymes of the Krebs citric acid cycle was unable to express early sporulation genes, apparently because of a failure to activate the phosphorelay. Cells that produce an altered Spo0A protein that can be ph...

  15. Characterization of human constitutive photomorphogenesis protein 1, a RING finger ubiquitin ligase that interacts with Jun transcription factors and modulates their transcriptional activity.

    Science.gov (United States)

    Bianchi, Elisabetta; Denti, Simona; Catena, Raffaella; Rossetti, Grazisa; Polo, Simona; Gasparian, Sona; Putignano, Stella; Rogge, Lars; Pardi, Ruggero

    2003-05-30

    RING finger proteins have been implicated in many fundamental cellular processes, including the control of gene expression. A key regulator of light-dependent development in Arabidopsis thaliana is the constitutive photomorphogenesis protein 1 (atCOP1), a RING finger protein that plays an essential role in translating light/dark signals into specific changes in gene transcription. atCOP1 binds the basic leucine zipper factor HY5 and suppresses its transcriptional activity through a yet undefined mechanism that results in HY5 degradation in response to darkness. Furthermore, the pleiotropic phenotype of atCOP1 mutants indicates that atCOP1 may be a central regulator of several transcriptional pathways. Here we report the cloning and characterization of the human orthologue of atCOP1. Human COP1 (huCOP1) distributes both to the cytoplasm and the nucleus of cells and shows a striking degree of sequence conservation with atCOP1, suggesting the possibility of a functional conservation as well. In co-immunoprecipitation assays huCOP1 specifically binds basic leucine zipper factors of the Jun family. As a functional consequence of this interaction, expression of huCOP1 in mammalian cells down-regulates c-Jun-dependent transcription and the expression of the AP-1 target genes, urokinase and matrix metalloproteinase 1. The RING domain of huCOP1 displays ubiquitin ligase activity in an autoubiquitination assay in vitro; however, suppression of AP-1-dependent transcription by huCOP1 occurs in the absence of changes in c-Jun protein levels, suggesting that this inhibitory effect is independent of c-Jun degradation. Our findings indicate that huCOP1 is a novel regulator of AP-1-dependent transcription sharing the important properties of Arabidopsis COP1 in the control of gene expression.

  16. Cartography of hematopoietic stem cell commitment dependent upon a reporter for transcription factor activation.

    Science.gov (United States)

    Akashi, Koichi

    2007-06-01

    A hierarchical hematopoietic developmental tree has been proposed based on the result of prospective purification of lineage-restricted progenitors. For more detailed mapping for hematopoietic stem cell (HSC) commitment, we tracked the expression of PU.1, a major granulocyte/monocyte (GM)- and lymphoid-related transcription factor, from the HSC to the myelolymphoid progenitor stages by using a mouse line harboring a knockin reporter for PU.1. This approach enabled us to find a new progenitor population committed to GM and lymphoid lineages within the HSC fraction. This result suggests that there should be another developmental pathway independent of the conventional one with myeloid versus lymphoid bifurcation, represented by common myeloid progenitors and common lymphoid progenitors, respectively. The utilization of the transcription factor expression as a functional marker might be useful to obtain cartography of the hematopoietic development at a higher resolution.

  17. Functional diversification of FD transcription factors in rice, components of florigen activation complexes.

    Science.gov (United States)

    Tsuji, Hiroyuki; Nakamura, Hiroyuki; Taoka, Ken-ichiro; Shimamoto, Ko

    2013-03-01

    Florigen, a protein encoded by the FLOWERING LOCUS T (FT) in Arabidopsis and Heading date 3a (Hd3a) in rice, is the universal flowering hormone in plants. Florigen is transported from leaves to the shoot apical meristem and initiates floral evocation. In shoot apical cells, conserved cytoplasmic 14-3-3 proteins act as florigen receptors. A hexameric florigen activation complex (FAC) composed of Hd3a, 14-3-3 proteins, and OsFD1, a transcription factor, activates OsMADS15, a rice homolog of Arabidopsis APETALA1, leading to flowering. Because FD is a key component of the FAC, we characterized the FD gene family and their functions. Phylogenetic analysis of FD genes indicated that this family is divided into two groups: (i) canonical FD genes that are conserved among eudicots and non-Poaceae monocots; and (ii) Poaceae-specific FD genes that are organized into three subgroups: Poaceae FD1, FD2 and FD3. The Poaceae FD1 group shares a small sequence motif, T(A/V)LSLNS, with FDs of eudicots and non-Poaceae monocots. Overexpression of OsFD2, a member of the Poaceae FD2 group, produced smaller leaves with shorter plastochrons, suggesting that OsFD2 controls leaf development. In vivo subcellular localization of Hd3a, 14-3-3 and OsFD2 suggested that in contrast to OsFD1, OsFD2 is restricted to the cytoplasm through its interaction with the cytoplasmic 14-3-3 proteins, and interaction of Hd3a with 14-3-3 facilitates nuclear translocation of the FAC containing OsFD2. These results suggest that FD function has diverged between OsFD1 and OsFD2, but formation of a FAC is essential for their function.

  18. Transforming growth factor β signaling upregulates the expression of human GDP-fucose transporter by activating transcription factor Sp1.

    Science.gov (United States)

    Xu, Yu-Xin; Ma, Anna; Liu, Li

    2013-01-01

    GDP-fucose transporter plays a crucial role in fucosylation of glycoproteins by providing activated fucose donor, GDP-fucose, for fucosyltransferases in the lumen of the Golgi apparatus. Fucose-containing glycans are involved in many biological processes, which are essential for growth and development. Mutations in the GDP-fucose transporter gene cause leukocyte adhesion deficiency syndrome II, a disease characterized by slow growth, mental retardation and immunodeficiency. However, no information is available regarding its transcriptional regulation. Here, by using human cells, we show that TGF-β1 specifically induces the GDP-fucose transporter expression, but not other transporters tested such as CMP-sialic acid transporter, suggesting a diversity of regulatory pathways for the expression of these transporters. The regulatory elements that are responsive to the TGF-β1 stimulation are present in the region between bp -330 and -268 in the GDP-fucose transporter promoter. We found that this region contains two identical octamer GC-rich motifs (GGGGCGTG) that were demonstrated to be essential for the transporter expression. We also show that the transcription factor Sp1 specifically binds to the GC-rich motifs in vitro and Sp1 coupled with phospho-Smad2 is associated with the promoter region covering the Sp1-binding motifs in vivo using chromatin immunoprecipitation (ChIP) assays. In addition, we further confirmed that Sp1 is essential for the GDP-fucose transporter expression stimulated by TGF-β1 using a luciferase reporter system. These results highlight the role of TGF-β signaling in regulation of the GDP-fucose transporter expression via activating Sp1. This is the first transcriptional study for any nucleotide sugar transporters that have been identified so far. Notably, TGF-β1 receptor itself is known to be modified by fucosylation. Given the essential role of GDP-fucose transporter in fucosylation, the finding that TGF-β1 stimulates the expression of

  19. Expression of a splice variant of the platelet-activating factor receptor transcript 2 in various human cancer cell lines

    Directory of Open Access Journals (Sweden)

    Ibtissam Youlyouz

    2002-01-01

    Full Text Available Platelet-activating factor receptor (PAF-R transcripts were analysed by reverse transcriptase-polymerase chain reaction in five human cancer cell lines derived from the breast (BT20, SKBR3 and T47D cells, the pancreas (Miapaca cells and the bladder (5637 cells in order to confirm the existence of a splice variant of the PAF-R transcript 2. After cloning and sequencing, we confirmed its existence in all cell lines. It consisted of the PAF-R transcript 2 lengthening with 82 nucleotides from the 3' end of exon 1 of the PAF-R gene. The role of this elongated form of the tissue-type PAF-R transcript in cell physiology remains to be elucidated.

  20. Occupancy by key transcription factors is a more accurate predictor of enhancer activity than histone modifications or chromatin accessibility.

    Science.gov (United States)

    Dogan, Nergiz; Wu, Weisheng; Morrissey, Christapher S; Chen, Kuan-Bei; Stonestrom, Aaron; Long, Maria; Keller, Cheryl A; Cheng, Yong; Jain, Deepti; Visel, Axel; Pennacchio, Len A; Weiss, Mitchell J; Blobel, Gerd A; Hardison, Ross C

    2015-01-01

    Regulated gene expression controls organismal development, and variation in regulatory patterns has been implicated in complex traits. Thus accurate prediction of enhancers is important for further understanding of these processes. Genome-wide measurement of epigenetic features, such as histone modifications and occupancy by transcription factors, is improving enhancer predictions, but the contribution of these features to prediction accuracy is not known. Given the importance of the hematopoietic transcription factor TAL1 for erythroid gene activation, we predicted candidate enhancers based on genomic occupancy by TAL1 and measured their activity. Contributions of multiple features to enhancer prediction were evaluated based on the results of these and other studies. TAL1-bound DNA segments were active enhancers at a high rate both in transient transfections of cultured cells (39 of 79, or 56%) and transgenic mice (43 of 66, or 65%). The level of binding signal for TAL1 or GATA1 did not help distinguish TAL1-bound DNA segments as active versus inactive enhancers, nor did the density of regulation-related histone modifications. A meta-analysis of results from this and other studies (273 tested predicted enhancers) showed that the presence of TAL1, GATA1, EP300, SMAD1, H3K4 methylation, H3K27ac, and CAGE tags at DNase hypersensitive sites gave the most accurate predictors of enhancer activity, with a success rate over 80% and a median threefold increase in activity. Chromatin accessibility assays and the histone modifications H3K4me1 and H3K27ac were sensitive for finding enhancers, but they have high false positive rates unless transcription factor occupancy is also included. Occupancy by key transcription factors such as TAL1, GATA1, SMAD1, and EP300, along with evidence of transcription, improves the accuracy of enhancer predictions based on epigenetic features.

  1. 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.

  2. 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.

  3. A Global Genomic and Genetic Strategy to Identify, Validate and Use Gene Signatures of Xenobiotic-Responsive Transcription Factors in Prediction of Pathway Activation in the Mouse Liver

    Science.gov (United States)

    Many drugs and environmentally-relevant chemicals activate xenobiotic-responsive transcription factors. Identification of target genes of these factors would be useful in predicting pathway activation in in vitro chemical screening as well as their involvement in disease states. ...

  4. Fungal-specific transcription factor AbPf2 activates pathogenicity in Alternaria brassicicola

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Yangrae; Ohm, Robin A.; Grigoriev, Igor V.; Srivastava, Akhil

    2012-12-03

    Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. To identify molecular determinants of pathogenicity, we created non-pathogenic mutants of a transcription factor-encoding gene, AbPf2. The frequency and timing of germination and appressorium formation on host plants were similar between the non-pathogenic abpf2 mutants and wild-type A. brassicicola. The mutants were also similar in vitro to wild-type A. brassicicola in terms of vegetative growth, conidium production, and responses to a phytoalexin, reactive oxygen species and osmolites. The hyphae of the mutants grew slowly but did not cause disease symptoms on the surface of host plants. Transcripts of the AbPf2 gene increased exponentially soon after wild-type conidia contacted their host plants . A small amount of AbPf2 protein, as monitored using GFP fusions, was present in young, mature conidia. The protein level decreased during saprophytic growth, but increased and was located primarily in fungal nuclei during pathogenesis. Levels of the proteins and transcripts sharply decreased following colonization of host tissues beyond the initial infection site. When expression of the transcription factor was induced in the wild-type during early pathogenesis, 106 fungal genes were also induced in the wild-type but not in the abpf2 mutants. Notably, 33 of the 106 genes encoded secreted proteins, including eight putative effector proteins. Plants inoculated with abpf2 mutants expressed higher levels of genes associated with photosynthesis, the pentose phosphate pathway and primary metabolism, but lower levels of defense-related genes. Our results suggest that AbPf2 is an important regulator of pathogenesis, but does not affect other cellular processes in A. brassicicola.

  5. Release of positive transcription elongation factor b (P-TEFb) from 7SK small nuclear ribonucleoprotein (snRNP) activates hexamethylene bisacetamide-inducible protein (HEXIM1) transcription.

    Science.gov (United States)

    Liu, Pingyang; Xiang, Yanhui; Fujinaga, Koh; Bartholomeeusen, Koen; Nilson, Kyle A; Price, David H; Peterlin, B Matija

    2014-04-04

    By phosphorylating negative elongation factors and the C-terminal domain of RNA polymerase II (RNAPII), positive transcription elongation factor b (P-TEFb), which is composed of CycT1 or CycT2 and CDK9, activates eukaryotic transcription elongation. In growing cells, it is found in active and inactive forms. In the former, free P-TEFb is a potent transcriptional coactivator. In the latter, it is inhibited by HEXIM1 or HEXIM2 in the 7SK small nuclear ribonucleoprotein (snRNP), which contains, additionally, 7SK snRNA, methyl phosphate-capping enzyme (MePCE), and La-related protein 7 (LARP7). This P-TEFb equilibrium determines the state of growth and proliferation of the cell. In this study, the release of P-TEFb from the 7SK snRNP led to increased synthesis of HEXIM1 but not HEXIM2 in HeLa cells, and this occurred only from an unannotated, proximal promoter. ChIP with sequencing revealed P-TEFb-sensitive poised RNA polymerase II at this proximal but not the previously annotated distal HEXIM1 promoter. Its immediate upstream sequences were fused to luciferase reporters and were found to be responsive to many P-TEFb-releasing compounds. The superelongation complex subunits AF4/FMR2 family member 4 (AFF4) and elongation factor RNA polymerase II 2 (ELL2) were recruited to this proximal promoter after P-TEFb release and were required for its transcriptional effects. Thus, P-TEFb regulates its own equilibrium in cells, most likely to maintain optimal cellular homeostasis.

  6. Signal Transducer and Activator of Transcription (STAT)-3 Activates Nuclear Factor (NF)-κB in Chronic Lymphocytic Leukemia Cells

    Science.gov (United States)

    Liu, Zhiming; Hazan-Halevy, Inbal; Harris, David M.; Li, Ping; Ferrajoli, Alessandra; Faderl, Stefan; Keating, Michael J.; Estrov, Zeev

    2014-01-01

    Nuclear factor (NF)-κB plays a major role in the pathogenesis of B-cell neoplasms. A broad array of mostly extracellular stimuli has been reported to activate NF-κB, to various degrees, in chronic lymphocytic leukemia (CLL) cells. Because CLL cells harbor high levels of unphosphorylated (U) signal transducer and activator of transcription (STAT)-3 protein and U-STAT3 was reported to activate NF-κB, we sought to determine whether U-STAT3 activates NF-κB in CLL. Using the electrophoretic mobility shift assay (EMSA) we studied peripheral blood low-density cells from 15 patients with CLL and found that CLL cell nuclear extracts from all the samples bound to an NF-κB DNA probe, suggesting that NF-κB is constitutively activated in CLL. Immunoprecipitation studies showed that STAT3 bound NF-κB p65, and confocal microscopy studies detected U-STAT3/NF-κB complexes in the nuclei of CLL cells, thereby confirming these findings. Furthermore, infection of CLL cells with retroviral STAT3-shRNA attenuated the binding of NF-κB to DNA, as assessed by EMSA, and downregulated mRNA levels of NF-κB-regulated genes, as assessed by quantitative polymerase chain reaction. Taken together, our data suggest that U-STAT3 binds to the NF-κB p50/p65 dimers and that the U-STAT3/NF-κB complexes bind to DNA and activate NF-κB-regulated genes in CLL cells. PMID:21364020

  7. Protein intrinsic disorder in Arabidopsis NAC transcription factors: transcriptional activation by ANAC013 and ANAC046 and their interactions with RCD1.

    Science.gov (United States)

    O'Shea, Charlotte; Kryger, Mikael; Stender, Emil G P; Kragelund, Birthe B; Willemoës, Martin; Skriver, Karen

    2015-01-15

    Protein ID (intrinsic disorder) plays a significant, yet relatively unexplored role in transcription factors (TFs). In the present paper, analysis of the transcription regulatory domains (TRDs) of six phylogenetically representative, plant-specific NAC [no apical meristem, ATAF (Arabidopsis transcription activation factor), cup-shaped cotyledon] TFs shows that the domains are present in similar average pre-molten or molten globule-like states, but have different patterns of order/disorder and MoRFs (molecular recognition features). ANAC046 (Arabidopsis NAC 046) was selected for further studies because of its simple MoRF pattern and its ability to interact with RCD1 (radical-induced cell death 1). Experiments in yeast and thermodynamic characterization suggest that its single MoRF region is sufficient for both transcriptional activation and interaction with RCD1. The remainder of the large regulatory domain is unlikely to contribute to the interaction, since the domain and truncations thereof have similar affinities for RCD1, which are also similar for ANAC013-RCD1 interactions. However, different enthalpic and entropic contributions to binding were revealed for ANAC046 and ANAC013, suggestive of differences in binding mechanisms. Although substitution of both hydrophobic and acidic residues of the ANAC046 MoRF region abolished binding, substitution of other residues, even with α-helix-breaking proline, was less disruptive. Together, the biophysical analyses suggest that RCD1-ANAC046 complex formation does not involve folding-upon-binding, but rather fuzziness or an unknown structure in ANAC046. We suggest that the ANAC046 regulatory domain functions as an entropic chain with a terminal hot spot interacting with RCD1. RCD1, a cellular hub, may be able to interact with many different TFs by exploiting their ID-based flexibility, as demonstrated for its interactions with ANAC046 and ANAC013.

  8. Activating transcription factor-4 promotes mineralization in vascular smooth muscle cells

    Science.gov (United States)

    Masuda, Masashi; Miyazaki-Anzai, Shinobu; Keenan, Audrey L.; Shiozaki, Yuji; Okamura, Kayo; Chick, Wallace S.; Williams, Kristina; Zhao, Xiaoyun; Rahman, Shaikh Mizanoor; Tintut, Yin; Adams, Christopher M.

    2016-01-01

    Emerging evidence indicates that upregulation of the ER stress–induced pro-osteogenic transcription factor ATF4 plays an important role in vascular calcification, a common complication in patients with aging, diabetes, and chronic kidney disease (CKD). In this study, we demonstrated the pathophysiological role of ATF4 in vascular calcification using global Atf4 KO, smooth muscle cell–specific (SMC-specific) Atf4 KO, and transgenic (TG) mouse models. Reduced expression of ATF4 in global ATF4-haplodeficient and SMC-specific Atf4 KO mice reduced medial and atherosclerotic calcification under normal kidney and CKD conditions. In contrast, increased expression of ATF4 in SMC-specific Atf4 TG mice caused severe medial and atherosclerotic calcification. We further demonstrated that ATF4 transcriptionally upregulates the expression of type III sodium-dependent phosphate cotransporters (PiT1 and PiT2) by interacting with C/EBPβ. These results demonstrate that the ER stress effector ATF4 plays a critical role in the pathogenesis of vascular calcification through increased phosphate uptake in vascular SMCs. PMID:27812542

  9. Cocaine induces cell death and activates the transcription nuclear factor kappa-b in pc12 cells

    Science.gov (United States)

    Lepsch, Lucilia B; Munhoz, Carolina D; Kawamoto, Elisa M; Yshii, Lidia M; Lima, Larissa S; Curi-Boaventura, Maria F; Salgado, Thais ML; Curi, Rui; Planeta, Cleopatra S; Scavone, Cristoforo

    2009-01-01

    Cocaine is a worldwide used drug and its abuse is associated with physical, psychiatric and social problems. The mechanism by which cocaine causes neurological damage is very complex and involves several neurotransmitter systems. For example, cocaine increases extracellular levels of dopamine and free radicals, and modulates several transcription factors. NF-κB is a transcription factor that regulates gene expression involved in cellular death. Our aim was to investigate the toxicity and modulation of NF-κB activity by cocaine in PC 12 cells. Treatment with cocaine (1 mM) for 24 hours induced DNA fragmentation, cellular membrane rupture and reduction of mitochondrial activity. A decrease in Bcl-2 protein and mRNA levels, and an increase in caspase 3 activity and cleavage were also observed. In addition, cocaine (after 6 hours treatment) activated the p50/p65 subunit of NF-κB complex and the pretreatment of the cells with SCH 23390, a D1 receptor antagonist, attenuated the NF-κB activation. Inhibition of NF-κB activity by using PDTC and Sodium Salicilate increased cell death caused by cocaine. These results suggest that cocaine induces cell death (apoptosis and necrosis) and activates NF-κB in PC12 cells. This activation occurs, at least partially, due to activation of D1 receptors and seems to have an anti-apoptotic effect on these cells. PMID:19183502

  10. Cocaine induces cell death and activates the transcription nuclear factor kappa-b in pc12 cells

    Directory of Open Access Journals (Sweden)

    Lepsch Lucilia B

    2009-02-01

    Full Text Available Abstract Cocaine is a worldwide used drug and its abuse is associated with physical, psychiatric and social problems. The mechanism by which cocaine causes neurological damage is very complex and involves several neurotransmitter systems. For example, cocaine increases extracellular levels of dopamine and free radicals, and modulates several transcription factors. NF-κB is a transcription factor that regulates gene expression involved in cellular death. Our aim was to investigate the toxicity and modulation of NF-κB activity by cocaine in PC 12 cells. Treatment with cocaine (1 mM for 24 hours induced DNA fragmentation, cellular membrane rupture and reduction of mitochondrial activity. A decrease in Bcl-2 protein and mRNA levels, and an increase in caspase 3 activity and cleavage were also observed. In addition, cocaine (after 6 hours treatment activated the p50/p65 subunit of NF-κB complex and the pretreatment of the cells with SCH 23390, a D1 receptor antagonist, attenuated the NF-κB activation. Inhibition of NF-κB activity by using PDTC and Sodium Salicilate increased cell death caused by cocaine. These results suggest that cocaine induces cell death (apoptosis and necrosis and activates NF-κB in PC12 cells. This activation occurs, at least partially, due to activation of D1 receptors and seems to have an anti-apoptotic effect on these cells.

  11. Effects of chronic renal failure rat serum on histone acetyltransferase p300 and activation of activating transcription factor 4 of arterial smooth muscle cells cultured in vitro

    Institute of Scientific and Technical Information of China (English)

    张耀全

    2014-01-01

    Objective To investigate the effects of the rat serum with chronic renal failure(CRF)on ubiquitin-proteasome pathway,histone acetyltransferase p300 and activation of activating transcription factor 4(ATF4)of rat arterial vascular smooth muscle cells(VSMCs)cultured in vitro,and explore the possible mechanism.Methods Objective To establish the rat model of

  12. The leucine zipper domains of the transcription factors GCN4 and c-Jun have ribonuclease activity.

    Directory of Open Access Journals (Sweden)

    Yaroslav Nikolaev

    Full Text Available Basic-region leucine zipper (bZIP proteins are one of the largest transcription factor families that regulate a wide range of cellular functions. Owing to the stability of their coiled coil structure leucine zipper (LZ domains of bZIP factors are widely employed as dimerization motifs in protein engineering studies. In the course of one such study, the X-ray structure of the retro-version of the LZ moiety of yeast transcriptional activator GCN4 suggested that this retro-LZ may have ribonuclease activity. Here we show that not only the retro-LZ but also the authentic LZ of GCN4 has weak but distinct ribonuclease activity. The observed cleavage of RNA is unspecific, it is not suppressed by the ribonuclease A inhibitor RNasin and involves the breakage of 3',5'-phosphodiester bonds with formation of 2',3'-cyclic phosphates as the final products as demonstrated by HPLC/electrospray ionization mass spectrometry. Several mutants of the GCN4 leucine zipper are catalytically inactive, providing important negative controls and unequivocally associating the enzymatic activity with the peptide under study. The leucine zipper moiety of the human factor c-Jun as well as the entire c-Jun protein are also shown to catalyze degradation of RNA. The presented data, which was obtained in the test-tube experiments, adds GCN4 and c-Jun to the pool of proteins with multiple functions (also known as moonlighting proteins. If expressed in vivo, the endoribonuclease activity of these bZIP-containing factors may represent a direct coupling between transcription activation and controlled RNA turnover. As an additional result of this work, the retro-leucine zipper of GCN4 can be added to the list of functional retro-peptides.

  13. LIM homeobox transcription factor Lhx2 inhibits skeletal muscle differentiation in part via transcriptional activation of Msx1 and Msx2.

    Science.gov (United States)

    Kodaka, Yusaku; Tanaka, Kiyoko; Kitajima, Kenji; Tanegashima, Kosuke; Matsuda, Ryoichi; Hara, Takahiko

    2015-02-15

    LIM homeobox transcription factor Lhx2 is known to be an important regulator of neuronal development, homeostasis of hair follicle stem cells, and self-renewal of hematopoietic stem cells; however, its function in skeletal muscle development is poorly understood. In this study, we found that overexpression of Lhx2 completely inhibits the myotube-forming capacity of C2C12 cells and primary myoblasts. The muscle dedifferentiation factors Msx1 and Msx2 were strongly induced by the Lhx2 overexpression. Short interfering RNA-mediated knockdown of Lhx2 in the developing limb buds of mouse embryos resulted in a reduction in Msx1 and Msx2 mRNA levels, suggesting that they are downstream target genes of Lhx2. We found two Lhx2 consensus-binding sites in the -2097 to -1189 genomic region of Msx1 and two additional sites in the -536 to +73 genomic region of Msx2. These sequences were shown by luciferase reporter assay to be essential for Lhx2-mediated transcriptional activation. Moreover, electrophoretic mobility shift assays and chromatin immunoprecipitation assays showed that Lhx2 is present in chromatin DNA complexes bound to the enhancer regions of the Msx1 and Msx2 genes. These data demonstrate that Msx1 and Msx2 are direct transcriptional targets of Lhx2. In addition, overexpression of Lhx2 significantly enhanced the mRNA levels of bone morphogenetic protein 4 and transforming growth factor beta family genes. We propose that Lhx2 is involved in the early stage of skeletal muscle development by inducing multiple differentiation inhibitory factors.

  14. Functional interplay of SP family members and nuclear factor Y is essential for transcriptional activation of the human Calreticulin gene.

    Science.gov (United States)

    Schardt, Julian A; Keller, Manuela; Seipel, Katja; Pabst, Thomas

    2015-09-01

    Calreticulin (CALR) is a highly conserved, multifunctional protein involved in a variety of cellular processes including the maintenance of intracellular calcium homeostasis, proper protein folding, differentiation and immunogenic cell death. More recently, a crucial role for CALR in the pathogenesis of certain hematologic malignancies was discovered: in clinical subgroups of acute myeloid leukemia, CALR overexpression mediates a block in differentiation, while somatic mutations have been found in the majority of patients with myeloproliferative neoplasms with nonmutated Janus kinase 2 gene (JAK2) or thrombopoietin receptor gene (MPL). However, the mechanisms underlying CALR promoter activation have insufficiently been investigated so far. By dissecting the core promoter region, we could identify a functional TATA-box relevant for transcriptional activation. In addition, we characterized two evolutionary highly conserved cis-regulatory modules (CRMs) within the proximal promoter each composed of one binding site for the transcription factors SP1 and SP3 as well as for the nuclear transcription factor Y (NFY) and we verified binding of these factors to their cognate sites in vitro and in vivo.

  15. Sigma Factors for Cyanobacterial Transcription

    Directory of Open Access Journals (Sweden)

    Sousuke Imamura

    2009-04-01

    Full Text Available Cyanobacteria are photosynthesizing microorganisms that can be used as a model for analyzing gene expression. The expression of genes involves transcription and translation. Transcription is performed by the RNA polymerase (RNAP holoenzyme, comprising a core enzyme and a sigma (σ factor which confers promoter selectivity. The unique structure, expression, and function of cyanobacterial σ factors (and RNAP core subunits are summarized here based on studies, reported previously. The types of promoter recognized by the σ factors are also discussed with regard to transcriptional regulation.

  16. Conserved region 2.1 of Escherichia coli heat shock transcription factor sigma32 is required for modulating both metabolic stability and transcriptional activity.

    Science.gov (United States)

    Horikoshi, Mina; Yura, Takashi; Tsuchimoto, Sachie; Fukumori, Yoshihiro; Kanemori, Masaaki

    2004-11-01

    Escherichia coli heat shock transcription factor sigma32 is rapidly degraded in vivo, with a half-life of about 1 min. A set of proteins that includes the DnaK chaperone team (DnaK, DnaJ, GrpE) and ATP-dependent proteases (FtsH, HslUV, etc.) are involved in degradation of sigma32. To gain further insight into the regulation of sigma32 stability, we isolated sigma32 mutants that were markedly stabilized. Many of the mutants had amino acid substitutions in the N-terminal half (residues 47 to 55) of region 2.1, a region highly conserved among bacterial sigma factors. The half-lives ranged from about 2-fold to more than 10-fold longer than that of the wild-type protein. Besides greater stability, the levels of heat shock proteins, such as DnaK and GroEL, increased in cells producing stable sigma32. Detailed analysis showed that some stable sigma32 mutants have higher transcriptional activity than the wild type. These results indicate that the N-terminal half of region 2.1 is required for modulating both metabolic stability and the activity of sigma32. The evidence suggests that sigma32 stabilization does not result from an elevated affinity for core RNA polymerase. Region 2.1 may, therefore, be involved in interactions with the proteolytic machinery, including molecular chaperones.

  17. Transcription factor CP2 is involved in activating mBMP4 in mouse mesenchymal stem cells.

    Science.gov (United States)

    Kang, Ho Chul; Chae, Ji Hyung; Kim, Beom Sue; Han, Su Youne; Kim, Sung-Hyun; Auh, Chung-Kyoon; Yang, Sung-Il; Kim, Chul Geun

    2004-06-30

    CP2 is a member of a family of transcription factors that regulate genes involved in events from early development to terminal differentiation. In an effort to understand how it selects its target genes we carried out a database search, and located several CP2 binding motifs in the promoter region of bone morphogenetic protein-4 (BMP4). BMP4 is a key regulator of cell fate and body patterning throughout development. For the CP2 binding motifs in BMP4 promoter region to be relevant in vivo, CP2 and BMP4 should be expressed together. We found that CP2b and CP2c, two potent transcriptional activators, are expressed in a manner similar to BMP4 during osteoblast differentiation of C3H10T1/2 cells. In in vitro assays, the CP2 proteins bound to two CP2 binding motifs (-715 to -676 and -147 to -118) in the BMP4 promoter, and luciferase reporter assays indicated that this binding was essential for transcription of BMP4 during osteoblast differentiation. Taken together, our data indicate that CP2b and CP2c play important roles during bone development by activating BMP4 transcription.

  18. The forkhead transcription factor Foxl2 is sumoylated in both human and mouse: sumoylation affects its stability, localization, and activity.

    Directory of Open Access Journals (Sweden)

    Mara Marongiu

    Full Text Available The FOXL2 forkhead transcription factor is expressed in ovarian granulosa cells, and mutated FOXL2 causes the blepharophimosis, ptosis and epicanthus inversus syndrome (BPES and predisposes to premature ovarian failure. Inactivation of Foxl2 in mice demonstrated its indispensability for female gonadal sex determination and ovary development and revealed its antagonism of Sox9, the effector of male testis development. To help to define the regulatory activities of FOXL2, we looked for interacting proteins. Based on yeast two-hybrid screening, we found that FOXL2 interacts with PIAS1 and UBC9, both parts of the sumoylation machinery. We showed that human FOXL2 is sumoylated in transfected cell lines, and that endogenous mouse Foxl2 is comparably sumoylated. This modification changes its cellular localization, stability and transcriptional activity. It is intriguing that similar sumoylation and regulatory consequences have also been reported for SOX9, the male counterpart of FOXL2 in somatic gonadal tissues.

  19. Structure and regulatory function of plant transcription factors

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    The expression of inducible genes in plants is regulated byspecific transcription factors at the transcriptional level. A typical transcription factor usually contains a DNA-binding domain, a transcription regulation domain, a dimerization site and a nuclear localization domain. These functional domains define the characteristic, localization and regulatory role of a transcription factor. Transcription factors recognize and bind to specific cis-acting elements or interact with other proteins, and then activate or repress the transcription of target genes by their functional domains. In recent years, elucidation on the structure and function of transcription factors has become an important subject in plant molecular biology.

  20. Fisetin stimulates autophagic degradation of phosphorylated tau via the activation of TFEB and Nrf2 transcription factors.

    Science.gov (United States)

    Kim, Sunhyo; Choi, Ki Ju; Cho, Sun-Jung; Yun, Sang-Moon; Jeon, Jae-Pil; Koh, Young Ho; Song, Jihyun; Johnson, Gail V W; Jo, Chulman

    2016-04-26

    The neuronal accumulation of phosphorylated tau plays a critical role in the pathogenesis of Alzheimer's disease (AD). Here, we examined the effect of fisetin, a flavonol, on tau levels. Treatment of cortical cells or primary neurons with fisetin resulted in significant decreases in the levels of phosphorylated tau. In addition, fisetin decreased the levels of sarkosyl-insoluble tau in an active GSK-3β-induced tau aggregation model. However, there was no difference in activities of tau kinases and phosphatases such as protein phosphatase 2A, irrespective of fisetin treatment. Fisetin activated autophagy together with the activation of transcription factor EB (TFEB) and Nrf2 transcriptional factors. The activation of autophagy including TFEB is likely due to fisetin-mediated mammalian target of rapamycin complex 1 (mTORC1) inhibition, since the phosphorylation levels of p70S6 kinase and 4E-BP1 were decreased in the presence of fisetin. Indeed, fisetin-induced phosphorylated tau degradation was attenuated by chemical inhibitors of the autophagy-lysosome pathway. Together the results indicate that fisetin reduces levels of phosphorylated tau through the autophagy pathway activated by TFEB and Nrf2. Our result suggests fisetin should be evaluated further as a potential preventive and therapeutic drug candidate for AD.

  1. Intracellular ascorbate enhances hypoxia-inducible factor (HIF)-hydroxylase activity and preferentially suppresses the HIF-1 transcriptional response.

    Science.gov (United States)

    Kuiper, Caroline; Dachs, Gabi U; Currie, Margaret J; Vissers, Margreet C M

    2014-04-01

    Hypoxia-inducible factor (HIF)-1 drives the transcription of hundreds of genes to support cell survival under conditions of microenvironmental and metabolic stress. HIF-1 is downregulated by iron-containing 2-oxoglutarate-dependent enzymes that require ascorbate as a cofactor. The HIF hydroxylases control both protein stability and the formation of an active transcription complex and, consequently, ascorbate could affect HIF-1α stabilization and/or gene expression, but the relative effect of ascorbate on these separate processes has not been well characterized. In this study we examined the effects of known intracellular ascorbate concentrations on both processes in response to various means of hydroxylase inhibition, including CoCl2, NiCl2, desferrioxamine, dimethyloxalylglycine, and hypoxia. Ascorbate inhibited HIF-1 activity most dramatically with all mechanisms of iron competition. In addition, HIF-1-dependent gene expression was effectively prevented by ascorbate and was inhibited even under conditions that allowed HIF-1α protein stabilization. This suggests that (1) ascorbate acts primarily to stabilize and reduce the iron atom in the hydroxylase active site and (2) the asparagine hydroxylase controlling HIF-1 transcriptional activity is particularly susceptible to fluctuations in intracellular ascorbate. These findings suggest that ascorbate plays a significant role in supporting HIF-hydroxylase function and that it could thereby modulate the cell survival response.

  2. Activation of transcription factor AP-1 in response to thermal injury in rat small intestine and IEC-6 cells.

    Science.gov (United States)

    Zhang, Yonghong; Zhao, Hong; Liu, Tao; Wan, Changrong; Liu, Xiaoxi; Gao, Zhimin; Hou, Xiaolin; Jiang, Linshu; Liu, Fenghua

    2015-07-11

    Our previous studies indicated that heat stress can cause significant damage to the intestinal epithelium and induce differential expression of many genes in rat small intestine. The transcription factors AP-1 and NF-κB, which act as important mediators by binding to specific DNA sequences within gene promoters, regulate the transcription of genes associated with immune regulation, stress response and cell fate. To determine whether AP-1 and NF-κB are involved in hyperthermia-induced injury in rat small intestine and IEC-6 cells, we investigated their activity, and the expression of related proteins, by electrophoretic mobility shift assays and western blotting, respectively. Heat stress resulted in severe damage to the epithelium of the small intestine. The cell morphology and viability were obviously altered when IEC-6 cell was exposed to hyperthermia. AP-1 was activated in the small intestine of heat-stressed rats, as was phosphorylation of the JNK signaling pathway. In IEC-6 cell line, AP-1 activation in groups exposed to 42 °C for 1 h, 2 h and 4 h was significantly increased. In contrast, NF-κB was not activated in both in vivo and in vitro models. These results reveal that AP-1 is likely to play an important role in regulating gene transcription in rat small intestine and IEC-6 cells during exposure to heat stress.

  3. 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

  4. Myocardin-related transcription factor-A controls myofibroblast activation and fibrosis in response to myocardial infarction

    Science.gov (United States)

    Small, Eric M.; Thatcher, Jeffrey E.; Sutherland, Lillian B.; Kinoshita, Hideyuki; Gerard, Robert D.; Richardson, James A.; DiMaio, J. Michael; Sadek, Hesham; Kuwahara, Koichiro; Olson, Eric N.

    2010-01-01

    Rationale Myocardial infarction (MI) results in loss of cardiac myocytes in the ischemic zone of the heart followed by fibrosis and scar formation, which diminish cardiac contractility and impede angiogenesis and repair. Myofibroblasts, a specialized cell type that switches from a fibroblast-like state to a contractile, smooth muscle-like state, are believed to be primarily responsible for fibrosis of the injured heart and other tissues, although the transcriptional mediators of fibrosis and myofibroblast activation remain poorly defined. Myocardin-related transcription factors (MRTFs) are SRF co-factors that promote a smooth muscle phenotype and are emerging as components of stress-responsive signaling. Objective We aimed to examine the effect of MRTF-A on cardiac remodeling and fibrosis. Methods and Results Here we show that MRTF-A controls the expression of a fibrotic gene program that includes genes involved in extracellular matrix production and smooth muscle cell differentiation in the heart. In MRTF-A null mice, fibrosis and scar formation following MI or Angiotensin (Ang) II treatment are dramatically diminished compared with wild-type littermates. This protective effect of MRTF-A deletion is associated with a reduction in expression of fibrosis-associated genes, including collagen 1a2, a direct transcriptional target of SRF/MRTF-A. Conclusions We conclude that MRTF-A regulates myofibroblast activation and fibrosis in response to the renin-angiotensin system and post-MI remodeling. PMID:20558820

  5. The role of factor inhibiting HIF (FIH-1 in inhibiting HIF-1 transcriptional activity in glioblastoma multiforme.

    Directory of Open Access Journals (Sweden)

    Enfeng Wang

    Full Text Available Glioblastoma multiforme (GBM accounts for about 38% of primary brain tumors in the United States. GBM is characterized by extensive angiogenesis induced by vascular growth factors and cytokines. The transcription of these growth factors and cytokines is regulated by the Hypoxia-Inducible-Factor-1(HIF-1, which is a key regulator mediating the cellular response to hypoxia. It is known that Factor Inhibiting HIF-1, or FIH-1, is also involved in the cellular response to hypoxia and has the capability to physically interact with HIF-1 and block its transcriptional activity under normoxic conditions. Delineation of the regulatory role of FIH-1 will help us to better understand the molecular mechanism responsible for tumor growth and progression and may lead to the design of new therapies targeting cellular pathways in response to hypoxia. Previous studies have shown that the chromosomal region of 10q24 containing the FIH-1 gene is often deleted in GBM, suggesting a role for the FIH-1 in GBM tumorigenesis and progression. In the current study, we found that FIH-1 is able to inhibit HIF-mediated transcription of GLUT1 and VEGF-A, even under hypoxic conditions in human glioblastoma cells. FIH-1 has been found to be more potent in inhibiting HIF function than PTEN. This observation points to the possibility that deletion of 10q23-24 and loss or decreased expression of FIH-1 gene may lead to a constitutive activation of HIF-1 activity, an alteration of HIF-1 targets such as GLUT-1 and VEGF-A, and may contribute to the survival of cancer cells in hypoxia and the development of hypervascularization observed in GBM. Therefore FIH-1 can be potential therapeutic target for the treatment of GBM patients with poor prognosis.

  6. c-Abl mediates high NaCl-induced phosphorylation and activation of the transcription factor TonEBP/OREBP

    Science.gov (United States)

    Gallazzini, Morgan; Yu, Ming-Jiun; Gunaratne, Ruwan; Burg, Maurice B.; Ferraris, Joan D.

    2010-01-01

    The transcription factor TonEBP/OREBP promotes cell survival during osmotic stress. High NaCl-induced phosphorylation of TonEBP/OREBP at tyrosine-143 was known to be an important factor in increasing its activity in cell culture. We now find that TonEBP/OREBP also is phosphorylated at tyrosine-143 in rat renal inner medulla, dependent on the interstitial osmolality. c-Abl seemed likely to be the kinase that phosphorylates TonEBP/OREBP because Y143 is in a consensus c-Abl phosphorylation site. We now confirm that, as follows. High NaCl increases c-Abl activity. Specific inhibition of c-Abl by imatinib, siRNA, or c-Abl kinase dead drastically reduces high NaCl-induced TonEBP/OREBP activity by reducing its nuclear location and transactivating activity. c-Abl associates with TonEBP/OREBP (coimmunoprecipitation) and phosphorylates TonEBP/OREBP-Y143 both in cell and in vitro. High NaCl-induced activation of ataxia telangiectasia mutated, previously known to contribute to activation of TonEBP/OREBP, depends on c-Abl activity. Thus, c-Abl is the kinase responsible for high NaCl-induced phosphorylation of TonEBP/OREBP-Y143, which contributes to its increased activity.—Gallazzini, M., Yu, M.-J., Gunaratne, R., Burg, M. B., Ferraris, J. D. c-Abl mediates high NaCl-induced phosphorylation and activation of the transcription factor TonEBP/OREBP. PMID:20585028

  7. Analysis of the DNA-binding and dimerization activities of Neurospora crassa transcription factor NUC-1.

    Science.gov (United States)

    Peleg, Y; Metzenberg, R L

    1994-12-01

    NUC-1, a positive regulatory protein of Neurospora crassa, controls the expression of several unlinked target genes involved in phosphorus acquisition. The carboxy-terminal end of the NUC-1 protein has sequence similarity to the helix-loop-helix family of transcription factors. Bacterially expressed and in vitro-synthesized proteins, which consist of the carboxy-terminal portion of NUC-1, bind specifically to upstream sequences of two of its target genes, pho2+ and pho-4+. These upstream sequences contain the core sequence, CACGTG, a target for many helix-loop-helix proteins. A large loop region (47 amino acids) separates the helix I and helix II domains. Mutations and deletion within the loop region did not interfere with the in vitro or in vivo functions of the protein. Immediately carboxy-proximal to the helix II domain, the NUC-1 protein contains an atypical zipper domain which is essential for function. This domain consists of a heptad repeat of alanine and methionine rather than leucine residues. Analysis of mutant NUC-1 proteins suggests that the helix II and the zipper domains are essential for the protein dimerization, whereas the basic and the helix I domains are involved in DNA binding. The helix I domain, even though likely to participate in dimer formation while NUC-1 is bound to DNA, is not essential for in vitro dimerization.

  8. A Sequentially Priming Phosphorylation Cascade Activates the Gliomagenic Transcription Factor Olig2

    Directory of Open Access Journals (Sweden)

    Jing Zhou

    2017-03-01

    Full Text Available During development of the vertebrate CNS, the basic helix-loop-helix (bHLH transcription factor Olig2 sustains replication competence of progenitor cells that give rise to neurons and oligodendrocytes. A pathological counterpart of this developmental function is seen in human glioma, wherein Olig2 is required for maintenance of stem-like cells that drive tumor growth. The mitogenic/gliomagenic functions of Olig2 are regulated by phosphorylation of a triple serine motif (S10, S13, and S14 in the amino terminus. Here, we identify a set of three serine/threonine protein kinases (glycogen synthase kinase 3α/β [GSK3α/β], casein kinase 2 [CK2], and cyclin-dependent kinases 1/2 [CDK1/2] that are, collectively, both necessary and sufficient to phosphorylate the triple serine motif. We show that phosphorylation of the motif itself serves as a template to prime phosphorylation of additional serines and creates a highly charged “acid blob” in the amino terminus of Olig2. Finally, we show that small molecule inhibitors of this forward-feeding phosphorylation cascade have potential as glioma therapeutics.

  9. Genotype-dependent activation or repression of HBV enhancer Ⅱ by transcription factor COUP-TF1

    Institute of Scientific and Technical Information of China (English)

    Silke F Fischer; Katja Schmidt; Nicola Fiedler; Dieter Glebe; Christian Schüttler; Jianguang Sun; Wolfram H Gerlich; Reinald Repp; Stephan Schaefer

    2006-01-01

    AIM: To study the expression of HBV enhancer Ⅱ by transcription factor COUP-TF1.METHODS: In order to study the regulation of HBV variants in the vicinity of the NRRE we cloned luciferase constructs containing the HBV enhancer Ⅱ from variants and from HBV genotypes A and D and cotransfected them together with expression vectors for COUP-TF1 into HepG2 cells.RESULTS: Our findings show that enhancer Ⅱ of HBV genotype A is also repressed by COUP-TF1. In contrast,two different enhancer Ⅱ constructs of HBV genotype D were activated by COUP-TF1. The activation was independent of the NRRE because a natural variant with a deletion of nt 1763-1770 was still activated by COUP-TF1.CONCLUSION: Regulation of transcription of the HBV genome seems to differ among HBV genomes derived from different genotypes. These differences in transcriptional control among HBV genotypes may be the molecular basis for differences in the clinical course among HBV genotypes.

  10. The Pioneer Transcription Factor FoxA Maintains an Accessible Nucleosome Configuration at Enhancers for Tissue-Specific Gene Activation.

    Science.gov (United States)

    Iwafuchi-Doi, Makiko; Donahue, Greg; Kakumanu, Akshay; Watts, Jason A; Mahony, Shaun; Pugh, B Franklin; Lee, Dolim; Kaestner, Klaus H; Zaret, Kenneth S

    2016-04-01

    Nuclear DNA wraps around core histones to form nucleosomes, which restricts the binding of transcription factors to gene regulatory sequences. Pioneer transcription factors can bind DNA sites on nucleosomes and initiate gene regulatory events, often leading to the local opening of chromatin. However, the nucleosomal configuration of open chromatin and the basis for its regulation is unclear. We combined low and high levels of micrococcal nuclease (MNase) digestion along with core histone mapping to assess the nucleosomal configuration at enhancers and promoters in mouse liver. We find that MNase-accessible nucleosomes, bound by transcription factors, are retained more at liver-specific enhancers than at promoters and ubiquitous enhancers. The pioneer factor FoxA displaces linker histone H1, thereby keeping enhancer nucleosomes accessible in chromatin and allowing other liver-specific transcription factors to bind and stimulate transcription. Thus, nucleosomes are not exclusively repressive to gene regulation when they are retained with, and exposed by, pioneer factors.

  11. The Journey of a Transcription Factor

    DEFF Research Database (Denmark)

    Pireyre, Marie

    Plants have developed astonishing networks regulating their metabolism to adapt to their environment. The complexity of these networks is illustrated by the expansion of families of regulators such as transcription factors in the plant kingdom. Transcription factors specifically impact...... transcriptional networks by integrating exogenous and endogenous stimuli and regulating gene expression accordingly. Regulation of transcription factors and their activation is thus highly important to modulate the transcriptional programs and increase fitness of the plant in a given environment. Plant metabolism...... is regulated to allocate resources to growth and/or defense at different time points. Among plant chemical defenses are the amino acid-derived glucosinolates (GLS). Their absolute and relative accumulation is tightly regulated at basal level, but also in response to e.g. pathogen attack and hormone stimuli...

  12. 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.

  13. A Taiwanese Propolis Derivative Induces Apoptosis through Inducing Endoplasmic Reticular Stress and Activating Transcription Factor-3 in Human Hepatoma Cells

    Directory of Open Access Journals (Sweden)

    Fat-Moon Suk

    2013-01-01

    Full Text Available Activating transcription factor-(ATF- 3, a stress-inducible transcription factor, is rapidly upregulated under various stress conditions and plays an important role in inducing cancer cell apoptosis. NBM-TP-007-GS-002 (GS-002 is a Taiwanese propolin G (PPG derivative. In this study, we examined the antitumor effects of GS-002 in human hepatoma Hep3B and HepG2 cells in vitro. First, we found that GS-002 significantly inhibited cell proliferation and induced cell apoptosis in dose-dependent manners. Several main apoptotic indicators were found in GS-002-treated cells, such as the cleaved forms of caspase-3, caspase-9, and poly(ADP-ribose polymerase (PARP. GS-002 also induced endoplasmic reticular (ER stress as evidenced by increases in ER stress-responsive proteins including glucose-regulated protein 78 (GRP78, growth arrest- and DNA damage-inducible gene 153 (GADD153, phosphorylated eukaryotic initiation factor 2α (eIF2α, phosphorylated protein endoplasmic-reticular-resident kinase (PERK, and ATF-3. The induction of ATF-3 expression was mediated by mitogen-activated protein kinase (MAPK signaling pathways in GS-002-treated cells. Furthermore, we found that GS-002 induced more cell apoptosis in ATF-3-overexpressing cells. These results suggest that the induction of apoptosis by the propolis derivative, GS-002, is partially mediated through ER stress and ATF-3-dependent pathways, and GS-002 has the potential for development as an antitumor drug.

  14. The transcriptional activity of hepatocyte nuclear factor 4 alpha is inhibited via phosphorylation by ERK1/2

    Science.gov (United States)

    Bacquet, Caroline; Kiss, Judit; Sipeki, Szabolcs; Martin, Ludovic; Buday, László; Bálint, Bálint L.; Arányi, Tamás

    2017-01-01

    Hepatocyte nuclear factor 4 alpha (HNF4α) nuclear receptor is a master regulator of hepatocyte development, nutrient transport and metabolism. HNF4α is regulated both at the transcriptional and post-transcriptional levels by different mechanisms. Several kinases (PKA, PKC, AMPK) were shown to phosphorylate and decrease the activity of HNF4α. Activation of the ERK1/2 signalling pathway, inducing proliferation and survival, inhibits the expression of HNF4α. However, based on our previous results we hypothesized that HNF4α is also regulated at the post-transcriptional level by ERK1/2. Here we show that ERK1/2 is capable of directly phosphorylating HNF4α in vitro at several phosphorylation sites including residues previously shown to be targeted by other kinases, as well. Furthermore, we also demonstrate that phosphorylation of HNF4α leads to a reduced trans-activational capacity of the nuclear receptor in luciferase reporter gene assay. We confirm the functional relevance of these findings by demonstrating with ChIP-qPCR experiments that 30-minute activation of ERK1/2 leads to reduced chromatin binding of HNF4α. Accordingly, we have observed decreasing but not disappearing binding of HNF4α to the target genes. In addition, 24-hour activation of the pathway further decreased HNF4α chromatin binding to specific loci in ChIP-qPCR experiments, which confirms the previous reports on the decreased expression of the HNF4a gene due to ERK1/2 activation. Our data suggest that the ERK1/2 pathway plays an important role in the regulation of HNF4α-dependent hepatic gene expression. PMID:28196117

  15. Transcription factors - Methods and protocols

    Directory of Open Access Journals (Sweden)

    CarloAlberto Redi

    2011-03-01

    Full Text Available A hearty wellcome to prof. Higgins editorial toil: a necessary tool for those colleagues (young and older fighting each day with the transcription factor they are involved with. In fact, the book is a full coverage compendium of state of the art papers dealing with practical thecniques and theoretical concepts about transcription factors. Each of the chapters (twenty-four is written by colleagues already working with one of the many trascription factors we become acquainted with. For the sake of the reader the volume is divided in four parts: Part I is a brief (when compared to the others three ! introductory presentation of the shuttling (i.e., transcription factor nuclear-cytoplasmic trafficking achieved by three reviews presentation of this biologically critical phenomenon. Part II (nine chapters is devoted to the necessary techniques to study nuclear translocation ...............

  16. Activating transcription factor 4 is involved in endoplasmic reticulum stress-mediated apoptosis contributing to vascular calcification.

    Science.gov (United States)

    Duan, Xiao-Hui; Chang, Jin-Rui; Zhang, Jing; Zhang, Bao-Hong; Li, Yu-Lin; Teng, Xu; Zhu, Yi; Du, Jie; Tang, Chao-Shu; Qi, Yong-Fen

    2013-09-01

    Our previous work reported that endoplasmic reticulum stress (ERS)-mediated apoptosis was activated during vascular calcification (VC). Activating transcription factor 4 (ATF4) is a critical transcription factor in osteoblastogenesis and ERS-induced apoptosis. However, whether ATF4 is involved in ERS-mediated apoptosis contributing to VC remains unclear. In the present study, in vivo VC was induced in rats by administering vitamin D3 plus nicotine. Vascular smooth muscle cell (VSMC) calcification in vitro was induced by incubation in calcifying media containing β-glycerophosphate and CaCl2. ERS inhibitors taurine or 4-phenylbutyric acid attenuated ERS and VSMC apoptosis in calcified rat arteries, reduced calcification and retarded the VSMC contractile phenotype transforming into an osteoblast-like phenotype in vivo. Inhibition of ERS retarded the VSMC phenotypic transition into an osteoblast-like cell phenotype and reduced VSMC calcification and apoptosis in vitro. Interestingly, ATF4 was activated in calcified aortas and calcified VSMCs in vitro. ATF4 knockdown attenuated ERS-induced apoptosis in calcified VSMCs. ATF4 deficiency blocked VSMC calcification and negatively regulated the osteoblast phenotypic transition of VSMCs in vitro. Our results demonstrate that ATF4 was involved at least in part in the process of ERS-mediated apoptosis contributing to VC.

  17. 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.

  18. Metformin Suppresses Lipopolysaccharide (LPS)-induced Inflammatory Response in Murine Macrophages via Activating Transcription Factor-3 (ATF-3) Induction*

    Science.gov (United States)

    Kim, Juyoung; Kwak, Hyun Jeong; Cha, Ji-Young; Jeong, Yun-Seung; Rhee, Sang Dahl; Kim, Kwang Rok; Cheon, Hyae Gyeong

    2014-01-01

    Metformin, a well known antidiabetic agent that improves peripheral insulin sensitivity, also elicits anti-inflammatory actions, but its mechanism is unclear. Here, we investigated the mechanism responsible for the anti-inflammatory effect of metformin action in lipopolysaccharide (LPS)-stimulated murine macrophages. Metformin inhibited LPS-induced production of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in a concentration-dependent manner and in parallel induction of activating transcription factor-3 (ATF-3), a transcription factor and member of the cAMP-responsive element-binding protein family. ATF-3 knockdown abolished the inhibitory effects of metformin on LPS-induced proinflammatory cytokine production accompanied with reversal of metformin-induced suppression of mitogen-activated protein kinase (MAPK) phosphorylation. Conversely, AMP-activated protein kinase (AMPK) phosphorylation and NF-κB suppression by metformin were unaffected by ATF-3 knockdown. ChIP-PCR analysis revealed that LPS-induced NF-κB enrichments on the promoters of IL-6 and TNF-α were replaced by ATF-3 upon metformin treatment. AMPK knockdown blunted all the effects of metformin (ATF-3 induction, proinflammatory cytokine inhibition, and MAPK inactivation), suggesting that AMPK activation by metformin is required for and precedes ATF-3 induction. Oral administration of metformin to either mice with LPS-induced endotoxemia or ob/ob mice lowered the plasma and tissue levels of TNF-α and IL-6 and increased ATF-3 expression in spleen and lungs. These results suggest that metformin exhibits anti-inflammatory action in macrophages at least in part via pathways involving AMPK activation and ATF-3 induction. PMID:24973221

  19. Metformin suppresses lipopolysaccharide (LPS)-induced inflammatory response in murine macrophages via activating transcription factor-3 (ATF-3) induction.

    Science.gov (United States)

    Kim, Juyoung; Kwak, Hyun Jeong; Cha, Ji-Young; Jeong, Yun-Seung; Rhee, Sang Dahl; Kim, Kwang Rok; Cheon, Hyae Gyeong

    2014-08-15

    Metformin, a well known antidiabetic agent that improves peripheral insulin sensitivity, also elicits anti-inflammatory actions, but its mechanism is unclear. Here, we investigated the mechanism responsible for the anti-inflammatory effect of metformin action in lipopolysaccharide (LPS)-stimulated murine macrophages. Metformin inhibited LPS-induced production of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in a concentration-dependent manner and in parallel induction of activating transcription factor-3 (ATF-3), a transcription factor and member of the cAMP-responsive element-binding protein family. ATF-3 knockdown abolished the inhibitory effects of metformin on LPS-induced proinflammatory cytokine production accompanied with reversal of metformin-induced suppression of mitogen-activated protein kinase (MAPK) phosphorylation. Conversely, AMP-activated protein kinase (AMPK) phosphorylation and NF-κB suppression by metformin were unaffected by ATF-3 knockdown. ChIP-PCR analysis revealed that LPS-induced NF-κB enrichments on the promoters of IL-6 and TNF-α were replaced by ATF-3 upon metformin treatment. AMPK knockdown blunted all the effects of metformin (ATF-3 induction, proinflammatory cytokine inhibition, and MAPK inactivation), suggesting that AMPK activation by metformin is required for and precedes ATF-3 induction. Oral administration of metformin to either mice with LPS-induced endotoxemia or ob/ob mice lowered the plasma and tissue levels of TNF-α and IL-6 and increased ATF-3 expression in spleen and lungs. These results suggest that metformin exhibits anti-inflammatory action in macrophages at least in part via pathways involving AMPK activation and ATF-3 induction. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Overexpression of the transcription factor Sp1 activates the OAS-RNAse L-RIG-I pathway.

    Directory of Open Access Journals (Sweden)

    Valéryane Dupuis-Maurin

    Full Text Available Deregulated expression of oncogenes or transcription factors such as specificity protein 1 (Sp1 is observed in many human cancers and plays a role in tumor maintenance. Paradoxically in untransformed cells, Sp1 overexpression induces late apoptosis but the early intrinsic response is poorly characterized. In the present work, we studied increased Sp1 level consequences in untransformed cells and showed that it turns on an early innate immune transcriptome. Sp1 overexpression does not activate known cellular stress pathways such as DNA damage response or endoplasmic reticulum stress, but induces the activation of the OAS-RNase L pathway and the generation of small self-RNAs, leading to the upregulation of genes of the antiviral RIG-I pathway at the transcriptional and translational levels. Finally, Sp1-induced intrinsic innate immune response leads to the production of the chemokine CXCL4 and to the recruitment of inflammatory cells in vitro and in vivo. Altogether our results showed that increased Sp1 level in untransformed cells constitutes a novel danger signal sensed by the OAS-RNase L axis leading to the activation of the RIG-I pathway. These results suggested that the OAS-RNase L-RIG-I pathway may be activated in sterile condition in absence of pathogen.

  1. Integration of G-Protein Coupled Receptor Signaling Pathways for Activation of a Transcription Factor (EGR-3)

    Institute of Scientific and Technical Information of China (English)

    Xuehai Tan; Pam Sanders; Jack Bolado Jr.; Mike Whitney

    2003-01-01

    We recently reported the use of a gene-trapping approach to isolate cell clones in which a reporter gene had integrated into genes modulated by T-cell activation. We have now tested a panel of clones from that report and identified the one that responds to a variety of G-protein coupled receptors (GPCR). The βlactamase tagged EGR-3 Jurkat cell was used to dissect specific GPCR signaling in vivo. Three GPCRs were studied, including the chemokine receptor CXCR4 (Gicoupled) that was endogenously expressed, the platelet activation factor (PAF) receptor (Gq-coupled), andβ2 adrenergic receptor (Gs-coupled) that was both stably transfected. Agonists for each receptor activated transcription of theβ-lactamase tagged EGR-3 gene. Induction of EGR-3 through CXCR4 was blocked by pertussis toxin and PD58059, a specific inhibitor of MEK (MAPK/ERK kinase). Neither of these inhibitors blocked isoproterenol or PAF-mediated activation of EGR-3. Conversely, β2- and PAF-mediated EGR-3 activation was blocked by the p38, specific inhibitor SB580. In addition, bothβ2- and PAF-mediated EGR-3 activation could be synergistically activated by CXCR4 activation. This combined result indicates that EGR-3 can be activated through distinct signal transduction pathways by different GPCRs and that signals can be integrated and amplified to efficiently tune the level of activation.

  2. 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.

  3. SUMOylation of the Forkhead transcription factor FOXL2 promotes its stabilization/activation through transient recruitment to PML bodies.

    Directory of Open Access Journals (Sweden)

    Adrien Georges

    Full Text Available BACKGROUND: FOXL2 is a transcription factor essential for ovarian development and maintenance. It is mutated in the genetic condition called Blepharophimosis Ptosis Epicantus inversus Syndrome (BPES and in cases of isolated premature ovarian failure. We and others have previously shown that FOXL2 undergoes several post-translational modifications. METHODS AND PRINCIPAL FINDINGS: Here, using cells in culture, we show that interference with FOXL2 SUMOylation leads to a robust inhibition of its transactivation ability, which correlates with a decreased stability. Interestingly, FOXL2 SUMOylation promotes its transient recruitment to subnuclear structures that we demonstrate to be PML (Promyelocytic Leukemia Nuclear Bodies. Since PML bodies are known to be sites where post-translational modifications of nuclear factors take place, we used tandem mass spectrometry to identify new post-translational modifications of FOXL2. Specifically, we detected four phosphorylated, one sulfated and three acetylated sites. CONCLUSIONS: By analogy with other transcription factors, we propose that PML Nuclear Bodies might transiently recruit FOXL2 to the vicinity of locally concentrated enzymes that could be involved in the post-translational maturation of FOXL2. FOXL2 acetylation, sulfation, phosphorylation as well as other modifications yet to be discovered might alter the transactivation capacity of FOXL2 and/or its stability, thus modulating its global intracellular activity.

  4. 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.

  5. Interactions of transcription factors with chromatin.

    Science.gov (United States)

    van Bakel, Harm

    2011-01-01

    Sequence-specific transcription factors (TFs) play a central role in regulating transcription initiation by directing the recruitment and activity of the general transcription machinery and accessory factors. It is now well established that many of the effects exerted by TFs in eukaryotes are mediated through interactions with a host of coregulators that modify the chromatin state, resulting in a more open (in case of activation) or closed conformation (in case of repression). The relationship between TFs and chromatin is a two-way street, however, as chromatin can in turn influence the recognition and binding of target sequences by TFs. The aim of this chapter is to highlight how this dynamic interplay between TF-directed remodelling of chromatin and chromatin-adjusted targeting of TF binding determines where and how transcription is initiated, and to what degree it is productive.

  6. Transcription Factors in Xylem Development. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Sederoff, Ronald; Whetten, Ross; O' Malley, David; Campbell, Malcolm

    1999-07-01

    Answers to the following questions are answered in this report. do the two pine Byb proteins previously identified as candidate transcription factors bind to DNA and activate transcription? In what cell types are tehse Myb proteins expressed? Are these proteins localized to the nucleus? Do other proteins in pine xylem interact with these Myb proteins? Does altered expression of these genes have an impact on xylogenesis, specifically the expression of monolignol biosynthetic genes?

  7. Transcription Factors in Xylem Development. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Sederoff, Ronald; Whetten, Ross; O' Malley, David; Campbell, Malcolm

    1999-07-01

    Answers to the following questions are answered in this report. do the two pine Byb proteins previously identified as candidate transcription factors bind to DNA and activate transcription? In what cell types are tehse Myb proteins expressed? Are these proteins localized to the nucleus? Do other proteins in pine xylem interact with these Myb proteins? Does altered expression of these genes have an impact on xylogenesis, specifically the expression of monolignol biosynthetic genes?

  8. Thymocytes are activated by toluene inhalation through the transcription factors NF-κB, STAT5 and NF-AT.

    Science.gov (United States)

    Liu, Jiqin; Yoshida, Yasuhiro; Kunugita, Naoki; Noguchi, Junko; Sugiura, Tsutomu; Ding, Ning; Arashidani, Keiichi; Fujimaki, Hidekazu; Yamashita, Uki

    2010-10-01

    Toluene has been extensively examined for effects on the central nervous system. To investigate the influence of low-level inhalation of toluene on the naive immune cells, male C3H/HeN mice were exposed to filtered air (control) and 50 ppm of toluene for 3 weeks. Low-level exposure resulted in (1) increased proliferation of thymocytes, (2) IL-2 production induced in thymocytes and (3) activation of the transcription factors NF-κB, STAT5 and NF-AT in thymocytes. These results suggest that thymocytes are sensitive cells and T cell activators are candidates for biomarkers for low-level exposure to toluene on naive immune cells. Copyright © 2010 John Wiley & Sons, Ltd.

  9. Common and distinct DNA-binding and regulatory activities of the BEN-solo transcription factor family.

    Science.gov (United States)

    Dai, Qi; Ren, Aiming; Westholm, Jakub O; Duan, Hong; Patel, Dinshaw J; Lai, Eric C

    2015-01-01

    Recently, the BEN (BANP, E5R, and NAC1) domain was recognized as a new class of conserved DNA-binding domain. The fly genome encodes three proteins that bear only a single BEN domain ("BEN-solo" factors); namely, Insensitive (Insv), Bsg25A (Elba1), and CG9883 (Elba2). Insv homodimers preferentially bind CCAATTGG palindromes throughout the genome to mediate transcriptional repression, whereas Bsg25A and Elba2 heterotrimerize with their obligate adaptor, Elba3 (i.e., the ELBA complex), to recognize a CCAATAAG motif in the Fab-7 insulator. While these data suggest distinct DNA-binding properties of BEN-solo proteins, we performed reporter assays that indicate that both Bsg25A and Elba2 can individually recognize Insv consensus sites efficiently. We confirmed this by solving the structure of Bsg25A complexed to the Insv site, which showed that key aspects of the BEN:DNA recognition strategy are similar between these proteins. We next show that both Insv and ELBA proteins are competent to mediate transcriptional repression via Insv consensus sequences but that the ELBA complex appears to be selective for the ELBA site. Reciprocally, genome-wide analysis reveals that Insv exhibits significant cobinding to class I insulator elements, indicating that it may also contribute to insulator function. Indeed, we observed abundant Insv binding within the Hox complexes with substantial overlaps with class I insulators, many of which bear Insv consensus sites. Moreover, Insv coimmunoprecipitates with the class I insulator factor CP190. Finally, we observed that Insv harbors exclusive activity among fly BEN-solo factors with respect to regulation of Notch-mediated cell fate choices in the peripheral nervous system. This in vivo activity is recapitulated by BEND6, a mammalian BEN-solo factor that conserves the Notch corepressor function of Insv but not its capacity to bind Insv consensus sites. Altogether, our data define an array of common and distinct biochemical and functional

  10. TEAD/TEF transcription factors utilize the activation domain of YAP65, a Src/Yes-associated protein localized in the cytoplasm.

    Science.gov (United States)

    Vassilev, A; Kaneko, K J; Shu, H; Zhao, Y; DePamphilis, M L

    2001-05-15

    Mammals express four highly conserved TEAD/TEF transcription factors that bind the same DNA sequence, but serve different functions during development. TEAD-2/TEF-4 protein purified from mouse cells was associated predominantly with a novel TEAD-binding domain at the amino terminus of YAP65, a powerful transcriptional coactivator. YAP65 interacted specifically with the carboxyl terminus of all four TEAD proteins. Both this interaction and sequence-specific DNA binding by TEAD were required for transcriptional activation in mouse cells. Expression of YAP in lymphocytic cells that normally do not support TEAD-dependent transcription (e.g., MPC11) resulted in up to 300-fold induction of TEAD activity. Conversely, TEAD overexpression squelched YAP activity. Therefore, the carboxy-terminal acidic activation domain in YAP is the transcriptional activation domain for TEAD transcription factors. However, whereas TEAD was concentrated in the nucleus, excess YAP65 accumulated in the cytoplasm as a complex with the cytoplasmic localization protein, 14-3-3. Because TEAD-dependent transcription was limited by YAP65, and YAP65 also binds Src/Yes protein tyrosine kinases, we propose that YAP65 regulates TEAD-dependent transcription in response to mitogenic signals.

  11. RNA helicase HEL-1 promotes longevity by specifically activating DAF-16/FOXO transcription factor signaling in Caenorhabditis elegans.

    Science.gov (United States)

    Seo, Mihwa; Seo, Keunhee; Hwang, Wooseon; Koo, Hee Jung; Hahm, Jeong-Hoon; Yang, Jae-Seong; Han, Seong Kyu; Hwang, Daehee; Kim, Sanguk; Jang, Sung Key; Lee, Yoontae; Nam, Hong Gil; Lee, Seung-Jae V

    2015-08-01

    The homeostatic maintenance of the genomic DNA is crucial for regulating aging processes. However, the role of RNA homeostasis in aging processes remains unknown. RNA helicases are a large family of enzymes that regulate the biogenesis and homeostasis of RNA. However, the functional significance of RNA helicases in aging has not been explored. Here, we report that a large fraction of RNA helicases regulate the lifespan of Caenorhabditis elegans. In particular, we show that a DEAD-box RNA helicase, helicase 1 (HEL-1), promotes longevity by specifically activating the DAF-16/forkhead box O (FOXO) transcription factor signaling pathway. We find that HEL-1 is required for the longevity conferred by reduced insulin/insulin-like growth factor 1 (IGF-1) signaling (IIS) and is sufficient for extending lifespan. We further show that the expression of HEL-1 in the intestine and neurons contributes to longevity. HEL-1 enhances the induction of a large fraction of DAF-16 target genes. Thus, the RNA helicase HEL-1 appears to promote longevity in response to decreased IIS as a transcription coregulator of DAF-16. Because HEL-1 and IIS are evolutionarily well conserved, a similar mechanism for longevity regulation via an RNA helicase-dependent regulation of FOXO signaling may operate in mammals, including humans.

  12. HDG1 transcription factor targets

    NARCIS (Netherlands)

    Horstman, A.; Boutilier, K.A.; Sanchez Perez, Gabino

    2015-01-01

    The AIL transcription factor BABY BOOM (BBM) is required together with the related PLETHORA proteins for embryo and root meristem development and its expression is sufficient to confer pluripotency and totipotency to somatic tissues. We show that BBM and other AIL proteins interact with multiple

  13. Structure and associated DNA-helicase activity of a general transcription initiation factor that binds to RNA polymerase II.

    Science.gov (United States)

    Sopta, M; Burton, Z F; Greenblatt, J

    1989-10-05

    RAP30/74 is a heteromeric general transcription initiation factor which binds to RNA polymerase II. Here we report that preparations of RAP30/74 contain an ATP-dependent DNA helicase whose probable function is to melt the DNA at transcriptional start sites. The sequence of the RAP30 subunit of RAP30/74 indicates that RAP30 may be distantly related to bacterial sigma factors.

  14. Increased accumulation of hypoxia-inducible factor-1α with reduced transcriptional activity mediates the antitumor effect of triptolide

    Directory of Open Access Journals (Sweden)

    Li Zheng

    2010-10-01

    Full Text Available Abstract Background Hypoxia-inducible factor-1α (HIF-1α, a critical transcription factor to reduced O2 availability, has been demonstrated to be extensively involved in tumor survival, aggressive progression, drug resistance and angiogenesis. Thus it has been considered as a potential anticancer target. Triptolide is the main principle responsible for the biological activities of the Traditional Chinese Medicine tripterygium wilfordii Hook F. Triptolide possesses great chemotherapy potential for cancer with its broad-spectrum anticancer, antiangiogenesis, and drug-resistance circumvention activities. Numerous biological molecules inhibited by triptolide have been viewed as its possible targets. However, the anticancer action mechanisms of triptolide remains to be further investigated. Here we used human ovarian SKOV-3 cancer cells as a model to probe the effect of triptolide on HIF-1α. Results Triptolide was observed to inhibit the proliferation of SKOV-3 cells, and meanwhile, to enhance the accumulation of HIF-1α protein in SKOV-3, A549 and DU145 cells under different conditions. Triptolide did not change the kinetics or nuclear localization of HIF-1α protein or the 26 S proteasome activity in SKOV-3 cells. However, triptolide was found to increase the levels of HIF-1α mRNA. Unexpectedly, the HIF-1α protein induced by triptolide appeared to lose its transcriptional activity, as evidenced by the decreased mRNA levels of its target genes including VEGF, BNIP3 and CAIX. The results were further strengthened by the lowered secretion of VEGF protein, the reduced sprout outgrowth from the rat aorta rings and the inhibitory expression of the hypoxia responsive element-driven luciferase reporter gene. Moreover, the silencing of HIF-1α partially prevented the cytotoxicity and apoptosis triggered by triptolide. Conclusions The potent induction of HIF-1α protein involved in its cytotoxicity, together with the suppression of HIF-1 transcriptional

  15. Activation of the Long Terminal Repeat of Human Endogenous Retrovirus K by Melanoma-Specific Transcription Factor MITF-M

    Directory of Open Access Journals (Sweden)

    Iyoko Katoh

    2011-11-01

    Full Text Available The human and Old World primate genomes possess conserved endogenous retrovirus sequences that have been implicated in evolution, reproduction, and carcinogenesis. Human endogenous retrovirus (HERV-K with 5′LTR-gag-pro-pol-env-rec/np9-3′LTR sequences represents the newest retrovirus family that integrated into the human genome 1 to 5 million years ago. Although a high-level expression of HERV-K in melanomas, breast cancers, and terato-carcinomas has been demonstrated, the mechanism of the lineage-specific activation of the long terminal repeat (LTR remains obscure. We studied chromosomal HERV-K expression in MeWo melanoma cells in comparison with the basal expression in human embryonic kidney 293 (HEK293 cells. Cloned LTR of HERV-K (HML-2.HOM was also characterized by mutation and transactivation experiments. We detected multiple transcriptional initiator (Inr sites in the LTR by rapid amplification of complementary DNA ends (5′ RACE. HEK293 and MeWo showed different Inr usage. The most potent Inr was associated with a TATA box and three binding motifs of microphthalmia-associated transcription factor (MITF. Both chromosomal HERV-K expression and the cloned LTR function were strongly activated in HEK293 by transfection with MITF-M, a melanocyte/melanoma–specific isoform of MITF. Coexpression of MITF and the HERV-K core antigen was detected in retinal pigmented epithelium by an immunofluorescence analysis. Although malignant melanoma lines MeWo, G361, and SK-MEL-28 showed enhanced HERV-K transcription compared with normal melanocytes, the level of MITF-M messenger RNA persisted from normal to transformed melanocytes. Thus, MITF-M may be a prerequisite for the pigmented cell lineage–specific function of HERV-K LTR, leading to the high-level expression in malignant melanomas.

  16. NAC transcription factors in senescence

    DEFF Research Database (Denmark)

    Podzimska-Sroka, Dagmara; O'Shea, Charlotte; Gregersen, Per L.;

    2015-01-01

    Within the last decade, NAC transcription factors have been shown to play essential roles in senescence, which is the focus of this review. Transcriptome analyses associate approximately one third of Arabidopsis NAC genes and many crop NAC genes with senescence, thereby implicating NAC genes...... as important regulators of the senescence process. The consensus DNA binding site of the NAC domain is used to predict NAC target genes, and protein interaction sites can be predicted for the intrinsically disordered transcription regulatory domains of NAC proteins. The molecular characteristics...

  17. Expression of peroxisome proliferator-activated receptor and CCAAT/enhancer binding protein transcription factors in cultured human sebocytes.

    Science.gov (United States)

    Chen, WenChieh; Yang, Chao-Chun; Sheu, Hamm-Ming; Seltmann, Holger; Zouboulis, Christos C

    2003-09-01

    Lipid synthesis and accumulation represent a major step in sebocyte differentiation and it may be of importance for sebocytes to express two families of transcription factors, CCAAT/enhancer binding proteins (c/EBPs) and peroxisome proliferator-activated receptors (PPARs), which were found to play a crucial role in the differentiation of adipocytes. Using the immortalized human sebaceous gland cell line SZ95 we examined the expression of the molecules before and after treatment with testosterone, 5alpha-dihydrotestosterone, dexamethasone, 17beta-estradiol and genistein, at 6, 12, 24, and 48 h, respectively. Reverse transcription-PCR analysis showed expression of peroxisome proliferator-activated receptors -alpha, -delta, -gamma1, -gamma2 and CCAAT/enhancer binding proteins-alpha, -beta, -gamma-delta in native SZ95 sebocytes. In western blot studies, high levels of CCAAT/enhancer binding proteins-alpha and -beta, and peroxisome proliferator-activated receptors-gamma were expressed at 6, 24, and 12 h, respectively. Immunostaining of the cultured sebocytes showed the CCAAT/enhancer binding proteins-alpha and -beta mainly localized within nuclei, whereas peroxisome proliferator-activated receptors-gamma in the cytoplasm. Strong staining of sebocytes was immunohistochemically revealed in the basal layer of sebaceous glands in human scalp and sebaceous nevus. Genistein down-regulated the expression of CCAAT/enhancer binding proteins-alpha and -beta, and peroxisome proliferator-activated receptors-gamma on the protein level. Treatment with linoleic acid for 48 h induced further differentiation of sebocytes leading to abundant lipid synthesis.

  18. Transcription factor CTCF and mammalian genome organization

    Directory of Open Access Journals (Sweden)

    Kotova E. S.

    2014-07-01

    Full Text Available The CTCF transcription factor is thought to be one of the main participants in various gene regulatory networks including transcription activation and repression, formation of independently functioning chromatin domains, regulation of imprinting etc. Sequencing of human and other genomes opened up a possibility to ascertain the genomic distribution of CTCF binding sites and to identify CTCF-dependent cis-regulatory elements, including insulators. In the review, we summarized recent data on CTCF functioning within a framework of the chromatin loop domain hypothesis of large-scale regulation of the genome activity. Its fundamental properties allow CTCF to serve as a transcription factor, an insulator protein and a dispersed genome-wide demarcation tool able to recruit various factors that emerge in response to diverse external and internal signals, and thus to exert its signal-specific function(s.

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

    Science.gov (United States)

    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.

  20. Unbiased identification of signal-activated transcription factors by barcoded synthetic tandem repeat promoter screening (BC-STAR-PROM).

    Science.gov (United States)

    Gosselin, Pauline; Rando, Gianpaolo; Fleury-Olela, Fabienne; Schibler, Ueli

    2016-08-15

    The discovery of transcription factors (TFs) controlling pathways in health and disease is of paramount interest. We designed a widely applicable method, dubbed barcorded synthetic tandem repeat promoter screening (BC-STAR-PROM), to identify signal-activated TFs without any a priori knowledge about their properties. The BC-STAR-PROM library consists of ∼3000 luciferase expression vectors, each harboring a promoter (composed of six tandem repeats of synthetic random DNA) and an associated barcode of 20 base pairs (bp) within the 3' untranslated mRNA region. Together, the promoter sequences encompass >400,000 bp of random DNA, a sequence complexity sufficient to capture most TFs. Cells transfected with the library are exposed to a signal, and the mRNAs that it encodes are counted by next-generation sequencing of the barcodes. This allows the simultaneous activity tracking of each of the ∼3000 synthetic promoters in a single experiment. Here we establish proof of concept for BC-STAR-PROM by applying it to the identification of TFs induced by drugs affecting actin and tubulin cytoskeleton dynamics. BC-STAR-PROM revealed that serum response factor (SRF) is the only immediate early TF induced by both actin polymerization and microtubule depolymerization. Such changes in cytoskeleton dynamics are known to occur during the cell division cycle, and real-time bioluminescence microscopy indeed revealed cell-autonomous SRF-myocardin-related TF (MRTF) activity bouts in proliferating cells.

  1. The LIM Homeodomain Transcription Factor LHX6

    Science.gov (United States)

    Zhang, Zichao; Gutierrez, Diana; Li, Xiao; Bidlack, Felicitas; Cao, Huojun; Wang, Jianbo; Andrade, Kelsey; Margolis, Henry C.; Amendt, Brad A.

    2013-01-01

    LHX6 is a LIM-homeobox transcription factor expressed during embryogenesis; however, the molecular mechanisms regulating LHX6 transcriptional activities are unknown. LHX6 and the PITX2 homeodomain transcription factor have overlapping expression patterns during tooth and craniofacial development, and in this report, we demonstrate new transcriptional mechanisms for these factors. PITX2 and LHX6 are co-expressed in the oral and dental epithelium and epithelial cell lines. Lhx6 expression is increased in Pitx2c transgenic mice and decreased in Pitx2 null mice. PITX2 activates endogenous Lhx6 expression and the Lhx6 promoter, whereas LHX6 represses its promoter activity. Chromatin immunoprecipitation experiments reveal endogenous PITX2 binding to the Lhx6 promoter. LHX6 directly interacts with PITX2 to inhibit PITX2 transcriptional activities and activation of multiple promoters. Bimolecular fluorescence complementation assays reveal an LHX6·PITX2 nuclear interaction in living cells. LHX6 has a dominant repressive effect on the PITX2 synergistic activation with LEF-1 and β-catenin co-factors. Thus, LHX6 acts as a transcriptional repressor and represses the expression of several genes involved in odontogenesis. We have identified specific defects in incisor, molar, mandible, bone, and root development and late stage enamel formation in Lhx6 null mice. Amelogenin and ameloblastin expression is reduced and/or delayed in the Lhx6 null mice, potentially resulting from defects in dentin deposition and ameloblast differentiation. Our results demonstrate that LHX6 regulates cell proliferation in the cervical loop and promotes cell differentiation in the anterior region of the incisor. We demonstrate new molecular mechanisms for LHX6 and an interaction with PITX2 for normal craniofacial and tooth development. PMID:23229549

  2. Activating transcription factor 4 underlies the pathogenesis of arsenic trioxide-mediated impairment of macrophage innate immune functions.

    Science.gov (United States)

    Srivastava, Ritesh K; Li, Changzhao; Wang, Yong; Weng, Zhiping; Elmets, Craig A; Harrod, Kevin S; Deshane, Jessy S; Athar, Mohammad

    2016-10-01

    Chronic arsenic exposure to humans is considered immunosuppressive with augmented susceptibility to several infectious diseases. The exact molecular mechanisms, however, remain unknown. Earlier, we showed the involvement of unfolded protein response (UPR) signaling in arsenic-mediated impairment of macrophage functions. Here, we show that activating transcription factor 4 (ATF4), a UPR transcription factor, regulates arsenic trioxide (ATO)-mediated dysregulation of macrophage functions. In ATO-treated ATF4(+/+) wild-type mice, a significant down-regulation of CD11b expression was associated with the reduced phagocytic functions of peritoneal and lung macrophages. This severe immuno-toxicity phenotype was not observed in ATO-treated ATF4(+/-) heterozygous mice. To confirm these observations, we demonstrated in Raw 264.7 cells that ATF4 knock-down rescues ATO-mediated impairment of macrophage functions including cytokine production, bacterial engulfment and clearance of engulfed bacteria. Sustained activation of ATF4 by ATO in macrophages induces apoptosis, while diminution of ATF4 expression protects against ATO-induced apoptotic cell death. Raw 264.7 cells treated with ATO also manifest dysregulated Ca(++) homeostasis. ATO induces Ca(++)-dependent calpain-1 and caspase-12 expression which together regulated macrophage apoptosis. Additionally, apoptosis was also induced by mitochondria-regulated pathway. Restoring ATO-impaired Ca(++) homeostasis in ER/mitochondria by treatments with the inhibitors of inositol 1,4,5-trisphosphate receptor (IP3R) and voltage-dependent anion channel (VDAC) attenuate innate immune functions of macrophages. These studies identify a novel role for ATF4 in underlying pathogenesis of macrophage dysregulation and immuno-toxicity of arsenic.

  3. Parathyroid hormone-related protein protects renal tubuloepithelial cells from apoptosis by activating transcription factor Runx2.

    Science.gov (United States)

    Ardura, Juan A; Sanz, Ana B; Ortiz, Alberto; Esbrit, Pedro

    2013-05-01

    Runx2 is a key transcription factor in bone development regulating several processes, including osteoblast apoptosis. The antiapoptotic effects of parathyroid hormone (PTH) in osteoblasts depend on Runx2-mediated transcription of prosurvival genes. In the kidney, PTH-related protein (PTHrP) promotes tubulointerstitial cell survival by activating the PTH/PTHrP type 1 receptor. We found that Runx2 is expressed in renal tubuloepithelial MCT and HK2 cell lines in vitro and in the mouse kidney tubuloepithelium in vivo. The 1-36 amino-acid fragment of PTHrP was found to increase the expression and nuclear translocation of Runx2 in both cell lines in a dose- and time-dependent manner. PTHrP(1-36) protected renal tubuloepithelial cells from folic acid toxicity and serum deprivation, an effect inhibited by a dominant-negative Runx2 construct or a Runx2 siRNA. Furthermore, PTHrP(1-36) upregulated the antiapoptotic proteins Bcl-2 and osteopontin, and these effects were abolished by Runx2 siRNA. Runx2, osteopontin, and Bcl-2 were increased in tubuloepithelial cells from transgenic mice with PTHrP overexpression and in wild-type mice with acute or chronic renal failure. Thus, PTHrP regulates renal tubuloepithelial cell survival via Runx2 in the mammalian kidney.

  4. Tailor-made zinc-finger transcription factors activate FLO11 gene expression with phenotypic consequences in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Shieh, Jia-Ching; Cheng, Yu-Che; Su, Mao-Chang; Moore, Michael; Choo, Yen; Klug, Aaron

    2007-08-15

    Cys2His2 zinc fingers are eukaryotic DNA-binding motifs, capable of distinguishing different DNA sequences, and are suitable for engineering artificial transcription factors. In this work, we used the budding yeast Saccharomyces cerevisiae to study the ability of tailor-made zinc finger proteins to activate the expression of the FLO11 gene, with phenotypic consequences. Two three-finger peptides were identified, recognizing sites from the 5' UTR of the FLO11 gene with nanomolar DNA-binding affinity. The three-finger domains and their combined six-finger motif, recognizing an 18-bp site, were fused to the activation domain of VP16 or VP64. These transcription factor constructs retained their DNA-binding ability, with the six-finger ones being the highest in affinity. However, when expressed in haploid yeast cells, only one three-finger recombinant transcription factor was able to activate the expression of FLO11 efficiently. Unlike in the wild-type, cells with such transcriptional activation displayed invasive growth and biofilm formation, without any requirement for glucose depletion. The VP16 and VP64 domains appeared to act equally well in the activation of FLO11 expression, with comparable effects in phenotypic alteration. We conclude that the functional activity of tailor-made transcription factors in cells is not easily predicted by the in vitro DNA-binding activity.

  5. The active spatial organization of the β-globin locus requires the transcription factor EKLF

    NARCIS (Netherlands)

    R.P.M. Drissen (Roy); R.-J.T.S. Palstra (Robert-Jan); N. Gillemans (Nynke); D. Splinter (Daniël); F.G. Grosveld (Frank); J.N.J. Philipsen (Sjaak); W.L. de Laat (Wouter)

    2004-01-01

    textabstractThree-dimensional organization of a gene locus is important for its regulation, as recently demonstrated for the β-globin locus. When actively expressed, the cis-regulatory elements of the β-globin locus are in proximity in the nuclear space, forming a compartment termed the Active

  6. The forkhead transcription factor FOXO3a controls microglial inflammatory activation and eventual apoptotic injury through caspase 3.

    Science.gov (United States)

    Shang, Yan Chen; Chong, Zhao Zhong; Hou, Jinling; Maiese, Kenneth

    2009-02-01

    Memory loss and cognitive failure are increasingly being identified as potential risks with the recognized increase in life expectancy of the general population. As a result, the development of novel therapeutic strategies for disorders such as Alzheimer's disease have garnered increased attention. The etiologies that can lead to Alzheimer's disease are extremely varied, but a number of therapeutic options are directed against amyloid-beta peptide and inflammatory cell regulation to prevent or halt progressive cognitive loss. In particular, inflammatory microglial cells may have disparate functions that in some scenarios lead to disability through the removal of functional neurovascular cells and in other circumstances foster tissue repair. Given the significance microglial cells hold for neurodegenerative disorders, we therefore examined the function that amyloid (Abeta(1-42)) has upon the microglial cell line EOC 2 and identified a novel role for the forkhead transcription factor FoxO3a and caspase 3. Here we show that Abeta(1-42) leads to progressive injury and apoptotic cell loss in microglial cells that involves both early phosphatidylserine (PS) externalization and late genomic DNA fragmentation over a 24 hour course. Prior to these injury programs, Abeta(1-42) results in the activation and proliferation of microglia as demonstrated by increased proliferating cell nuclear antigen (PCNA) expression and bromodeoxyuridine (BrdU) uptake. Both apoptotic injury as well as the prior activation and proliferation of microglial cells relies upon the presence of FoxO3a, since specific gene silencing of FoxO3a promotes microglial cell protection and prevents the early activation and proliferation of these cells. Furthermore, Abeta(1-42) exposure maintained FoxO3a in an unphosphorylated "active" state and facilitated the cellular trafficking of FoxO3a from the cytoplasm to the cell nucleus to potentially lead to "pro-apoptotic" programs by this transcription factor. One

  7. Fibronectin upregulates cGMP-dependent protein kinase type Iβ through C/EBP transcription factor activation in contractile cells.

    Science.gov (United States)

    Chamorro-Jorganes, Aranzazu; Calleros, Laura; Griera, Mercedes; Saura, Marta; Luengo, Alicia; Rodriguez-Puyol, D; Rodriguez-Puyol, M

    2011-03-01

    The nitric oxide (NO)-soluble guanylate cyclase (sGC) pathway exerts most of its cellular actions through the activation of the cGMP-dependent protein kinase (PKG). Accumulation of extracellular matrix is one of the main structural changes in pathological conditions characterized by a decreased activity of this pathway, such as hypertension, diabetes, or aging, and it is a well-known fact that extracellular matrix proteins modulate cell phenotype through the interaction with membrane receptors such as integrins. The objectives of this study were 1) to evaluate whether extracellular matrix proteins, particularly fibronectin (FN), modulate PKG expression in contractile cells, 2) to analyze the mechanisms involved, and 3) to evaluate the functional consequences. FN increased type I PKG (PKG-I) protein content in human mesangial cells, an effect dependent on the interaction with β(1)-integrin. The FN upregulation of PKG-I protein content was due to increased mRNA expression, determined by augmented transcriptional activity of the PKG-I promoter region. Akt and the transcription factor CCAAT enhancer-binding protein (C/EBP) mediated the genesis of these changes. FN also increased PKG-I in another type of contractile cell, rat vascular smooth muscle cells (RVSMC). Tirofiban, a pharmacological analog of FN, increased PKG-I protein content in RVSMC and rat aortic walls and magnified the hypotensive effect of dibutyryl cGMP in conscious Wistar rats. The present results provide evidence of a mechanism able to increase PKG-I protein content in contractile cells. Elucidation of this novel mechanism provides a rationale for future pharmacotherapy in certain vascular diseases.

  8. A WRKY Transcription Factor Recruits the SYG1-Like Protein SHB1 to Activate Gene Expression and Seed Cavity Enlargement

    Science.gov (United States)

    Kang, Xiaojun; Li, Wei; Zhou, Yun; Ni, Min

    2013-01-01

    Seed development in Arabidopsis and in many dicots involves an early proliferation of the endosperm to form a large embryo sac or seed cavity close to the size of the mature seed, followed by a second phase during which the embryo grows and replaces the endosperm. SHORT HYPOCOTYL UNDER BLUE1 (SHB1) is a member of the SYG1 protein family in fungi, Caenorhabditis elegans, flies, and mammals. SHB1 gain-of-function enhances endosperm proliferation, increases seed size, and up-regulates the expression of the WRKY transcription factor gene MINISEED3 (MINI3) and the LRR receptor kinase gene HAIKU2 (IKU2). Mutations in either IKU2 or MINI3 retard endosperm proliferation and reduce seed size. However, the molecular mechanisms underlying the establishment of the seed cavity and hence the seed size remain largely unknown. Here, we show that the expression of MINI3 and IKU2 is repressed before fertilization and after 4 days after pollination (DAP), but is activated by SHB1 from 2 to 4 DAP prior to the formation of the seed cavity. SHB1 associates with their promoters but without a recognizable DNA binding motif, and this association is abolished in mini3 mutant. MINI3 binds to W-boxes in, and recruits SHB1 to, its own and IKU2 promoters. Interestingly, SHB1, but not MINI3, activates transcription of pMINI3::GUS or pIKU2::GUS. We reveal a critical developmental switch through the activation of MINI3 expression by SHB1. The recruitment of SHB1 by MINI3 to its own and IKU2 promoters represents a novel two-step amplification to counter the low expression level of IKU2, which is a trigger for endosperm proliferation and seed cavity enlargement. PMID:23505389

  9. Cross-Family Transcription Factor Interactions

    NARCIS (Netherlands)

    Bemer, Marian; Dijk, van Aalt-Jan; Immink, Richard G.H.; Angenent, Gerco C.

    2017-01-01

    Specific and dynamic gene expression strongly depends on transcription factor (TF) activity and most plant TFs function in a combinatorial fashion. They can bind to DNA and control the expression of the corresponding gene in an additive fashion or cooperate by physical interactions, forming larger p

  10. Involvement of transcription factor activator protein-2α in doxazosin-induced HeLa cell apoptosis

    Institute of Scientific and Technical Information of China (English)

    Lu GAN; Dong-xing ZHU; Li-ping YANG; Ru-shi LIU; Feng YAN; Jian ZHANG

    2008-01-01

    Aim:To investigate the pro-apoptotic effects of α- 1-adrenergic inhibitor doxazosin in HeLa cells and the potential involvement of transcription factor activator pro-tein-2α (AP-2α) in doxazosin-indueed apoptosis. Methods:The HeLa cells were exposed to various concentrations of doxazosin for 16 h. Apoptosis was detected using a DNA fragmentation assay, Hoechst 33258 staining, and flow cytometric analysis. The expression of AP-2α and caspase-3 was detected by relative quan-titative RT-PCR and Western blot assays, respectively. After the respective trans-fections of the HeLa cells with AP-2α overexpressing constructs and an antisense oligonucleotide against AP-2α, apoptosis was assessed by flow cytometric analysis, and the expression of AP-2α and easpase-3 was detected by relative quantitative RT-PCR and Western blot assays. The colorimetric assay was per-formed to detect the caspase-3 activity. Results:Treatment with various concen-trations of doxazosin for 16 h increased the apoptotic rate and total cell death rate of the HeLa cells in a dose-dependent manner and upregulated the expression of AP-2α and caspase-3 in a dose-dependent manner. A dose-dependent increase was observed in the caspase-3 activity. Overexpressing AP-2α led to the in-creased rate of doxazosin-induced apoptosis and the total cell death, whereas doxazosin-induced apoptosis and the total cell death in HeLa cells decreased by antisense AP-2α. Furthermore, overexpressing AP-2α increased the expression and activity of caspase-3, whereas antisense AP-2α in part abolished the increased effects of doxazosin on caspase-3 expression and activity. Conclusion:Doxazosin induces apoptosis in HeLa cells in a dose-dependent manner, and transcription factor AP-2α is functionally involved in doxazosin-induced HeLa cell apoptosis.

  11. Quercetin induces growth arrest through activation of FOXO1 transcription factor in EGFR-overexpressing oral cancer cells.

    Science.gov (United States)

    Huang, Chun-Yin; Chan, Chien-Yi; Chou, I-Tai; Lien, Chia-Hsien; Hung, Hsiao-Chi; Lee, Ming-Fen

    2013-09-01

    The squamous cell carcinomas of the head and neck (SCCHNs) with aberrant epidermal growth factor receptor (EGFR) signaling are often associated with poor prognosis and low survival. Therefore, efficient inhibition of the EGFR signaling could intervene with the development of malignancy. Quercetin appears to be antitumorigenesis, but the underlying mechanism remains unclear in oral cancer. Fork-head box O (FOXO) transcription factors, Akt downstream effectors, are important regulators of cell growth. Here, we hypothesized that FOXO1 might be crucial in quercetin-induced growth inhibition in EGFR-overexpressing oral cancer. Quercetin treatment suppressed cell growth by inducing G2 arrest and apoptosis in EGFR-overexpressing HSC-3 and TW206 oral cancer cells. Quercetin inhibited EGFR/Akt activation with a concomitant induction of FOXO1 activation. FOXO1 knockdown attenuated quercetin-induced p21 and FasL expression and subsequent G2 arrest and apoptosis, respectively. Likewise, quercetin suppressed tumor growth in HSC-3 xenograft mice. Taken together, our data indicate that quercetin is an effective anticancer agent and that FOXO1 is crucial in quercetin-induced growth suppression in EGFR-overexpressing oral cancer. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. Hepatitis C virus nonstructural protein-5A activates sterol regulatory element-binding protein-1c through transcription factor Sp1

    Energy Technology Data Exchange (ETDEWEB)

    Xiang, Zhonghua; Qiao, Ling; Zhou, Yan [Vaccine and Infectious Disease Organization, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E3 (Canada); Babiuk, Lorne A. [University of Alberta, Edmonton, Alberta (Canada); Liu, Qiang, E-mail: qiang.liu@usask.ca [Vaccine and Infectious Disease Organization, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E3 (Canada)

    2010-11-19

    Research highlights: {yields} A chimeric subgenomic HCV replicon expresses HCV-3a NS5A in an HCV-1b backbone. {yields} HCV-3a NS5A increases mature SREBP-1c protein level. {yields} HCV-3a NS5A activates SREBP-1c transcription. {yields} Domain II of HCV-3a NS5A is more effective in SREBP-1c promoter activation. {yields} Transcription factor Sp1 is required for SREBP-1c activation by HCV-3a NS5A. -- Abstract: Steatosis is an important clinical manifestation of hepatitis C virus (HCV) infection. The molecular mechanisms of HCV-associated steatosis are not well understood. Sterol regulatory element-binding protein-1c (SREBP-1c) is a key transcription factor which activates the transcription of lipogenic genes. Here we showed that the nuclear, mature SREBP-1c level increases in the nucleus of replicon cells expressing HCV-3a nonstructural protein-5A (NS5A). We further showed that HCV-3a NS5A up-regulates SREBP-1c transcription. Additional analysis showed that transcriptional factor Sp1 is involved in SREBP-1c activation by HCV-3a NS5A because inhibition of Sp1 activity by mithramycin A or a dominant-negative Sp1 construct abrogated SREBP-1c promoter activation by HCV-3a NS5A. In addition, chromatin immunoprecipitation (ChIP) assay demonstrated enhanced binding of Sp1 on the SREBP-1c promoter in HCV-3a NS5A replicon cells. These results showed that HCV-3a NS5A activates SREBP-1c transcription through Sp1. Taken together, our results suggest that HCV-3a NS5A is a contributing factor for steatosis caused by HCV-3a infection.

  13. Transcription factor network downstream of protease activated receptors (PARs modulating mouse bladder inflammation

    Directory of Open Access Journals (Sweden)

    Hurst Robert E

    2007-08-01

    Full Text Available Abstract Background All four PARs are present in the urinary bladder, and their expression is altered during inflammation. In order to search for therapeutic targets other than the receptors themselves, we set forth to determine TFs downstream of PAR activation in the C57BL/6 urinary bladders. Methods For this purpose, we used a protein/DNA combo array containing 345 different TF consensus sequences. Next, the TF selected was validated by EMSA and IHC. As mast cells seem to play a fundamental role in bladder inflammation, we determined whether c-kit receptor deficient (Kitw/Kitw-v mice have an abrogated response to PAR stimulation. Finally, TFEB antibody was used for CHIP/Q-PCR assay and revealed up-regulation of genes known to be downstream of TFEB. Results TFEB, a member of the MiTF family of basic helix-loop-helix leucine zipper, was the only TF commonly up-regulated by all PAR-APs. IHC results confirm a correlation between inflammation and TFEB expression in C57BL/6 mice. In contrast, Kitw/Kitw-v mice did not exhibit inflammation in response to PAR activation. EMSA results confirmed the increased TFEB binding activity in C57BL/6 but not in Kitw/Kitw-v mice. Conclusion This is the first report describing the increased expression of TFEB in bladder inflammation in response to PAR activation. As TFEB belongs to a family of TFs essential for mast cell survival, our findings suggest that this molecule may influence the participation of mast cells in PAR-mediated inflammation and that targeting TFEB/MiTF activity may be a novel approach for the treatment of bladder inflammatory disorders.

  14. 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.

  15. Coffee modulates transcription factor Nrf2 and highly increases the activity of antioxidant enzymes in rats.

    Science.gov (United States)

    Vicente, Silvio J V; Ishimoto, Emília Y; Torres, Elizabeth A F S

    2014-01-08

    This study investigated the effect of a 28 day administration of coffee brew on the activity of antioxidant enzymes in rats. After this period of 2.0 mL/day dosages of this beverage, the activities of hepatic superoxide dismutase, catalase, and glutathione peroxidase increased 74.8, 59.4, and 135.2%, respectively, whereas the cytosolic level of Nrf2 increased 131.3%. At the same time, the total antioxidant capacity of the hepatic tissue increased 25.1%, improving the defensive status against oxidative stress. At the end of the experiment, the levels of biomarkers alanine transaminase and aspartate transaminase remained equal to the control group, and no changes were observed in the hepatic histoarchiteture of the animals, suggesting that the liver tissue was not impaired by the exposure to coffee. The changes in enzyme activities and antioxidant capacity were statistically significant (p < 0.05), indicating that coffee could be considered an important alternative against oxidative stress and its correlated degenerative diseases.

  16. Polyphenol Compound as a Transcription Factor Inhibitor

    Directory of Open Access Journals (Sweden)

    Seyeon Park

    2015-10-01

    Full Text Available A target-based approach has been used to develop novel drugs in many therapeutic fields. In the final stage of intracellular signaling, transcription factor–DNA interactions are central to most biological processes and therefore represent a large and important class of targets for human therapeutics. Thus, we focused on the idea that the disruption of protein dimers and cognate DNA complexes could impair the transcriptional activation and cell transformation regulated by these proteins. Historically, natural products have been regarded as providing the primary leading compounds capable of modulating protein–protein or protein-DNA interactions. Although their mechanism of action is not fully defined, polyphenols including flavonoids were found to act mostly as site-directed small molecule inhibitors on signaling. There are many reports in the literature of screening initiatives suggesting improved drugs that can modulate the transcription factor interactions responsible for disease. In this review, we focus on polyphenol compound inhibitors against dimeric forms of transcription factor components of intracellular signaling pathways (for instance, c-jun/c-fos (Activator Protein-1; AP-1, c-myc/max, Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB and β-catenin/T cell factor (Tcf.

  17. 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.

  18. Activation of Transcription Factor MEF2D by Bis(3)-cognitin Protects Dopaminergic Neurons and Ameliorates Parkinsonian Motor Defects*

    Science.gov (United States)

    Yao, Lu; Li, Wenming; She, Hua; Dou, Juan; Jia, Leili; He, Yingli; Yang, Qian; Zhu, Jinqiu; Cápiro, Natalie L.; Walker, Douglas I.; Pennell, Kurt D.; Pang, Yuanping; Liu, Yong; Han, Yifan; Mao, Zixu

    2012-01-01

    Parkinson disease (PD) is characterized by the selective demise of dopaminergic (DA) neurons in the substantial nigra pars compacta. Dysregulation of transcriptional factor myocyte enhancer factor 2D (MEF2D) has been implicated in the pathogenic process in in vivo and in vitro models of PD. Here, we identified a small molecule bis(3)-cognitin (B3C) as a potent activator of MEF2D. We showed that B3C attenuated the toxic effects of neurotoxin 1-methyl-4-phenylpyridinium (MPP+) by activating MEF2D via multiple mechanisms. B3C significantly reduced MPP+-induced oxidative stress and potentiated Akt to down-regulate the activity of MEF2 inhibitor glycogen synthase kinase 3β (GSK3β) in a DA neuronal cell line SN4741. Furthermore, B3C effectively rescued MEF2D from MPP+-induced decline in both nucleic and mitochondrial compartments. B3C offered SN4741 cells potent protection against MPP+-induced apoptosis via MEF2D. Interestingly, B3C also protected SN4741 cells from wild type or mutant A53T α-synuclein-induced cytotoxicity. Using the in vivo PD model of C57BL/6 mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP), we showed that B3C maintained redox homeostasis, promoted Akt function activity, and restored MEF2D level in midbrain neurons. Moreover, B3C greatly prevented the loss of tyrosine hydroxylase signal in substantial nigra pars compacta DA neurons and ameliorated behavioral impairments in mice treated with MPTP. Collectedly, our studies identified B3C as a potent neuroprotective agent whose effectiveness relies on its ability to effectively up-regulate MEF2D in DA neurons against toxic stress in models of PD in vitro and in vivo. PMID:22891246

  19. Activation of transcription factor MEF2D by bis(3)-cognitin protects dopaminergic neurons and ameliorates Parkinsonian motor defects.

    Science.gov (United States)

    Yao, Lu; Li, Wenming; She, Hua; Dou, Juan; Jia, Leili; He, Yingli; Yang, Qian; Zhu, Jinqiu; Cápiro, Natalie L; Walker, Douglas I; Pennell, Kurt D; Pang, Yuanping; Liu, Yong; Han, Yifan; Mao, Zixu

    2012-10-05

    Parkinson disease (PD) is characterized by the selective demise of dopaminergic (DA) neurons in the substantial nigra pars compacta. Dysregulation of transcriptional factor myocyte enhancer factor 2D (MEF2D) has been implicated in the pathogenic process in in vivo and in vitro models of PD. Here, we identified a small molecule bis(3)-cognitin (B3C) as a potent activator of MEF2D. We showed that B3C attenuated the toxic effects of neurotoxin 1-methyl-4-phenylpyridinium (MPP(+)) by activating MEF2D via multiple mechanisms. B3C significantly reduced MPP(+)-induced oxidative stress and potentiated Akt to down-regulate the activity of MEF2 inhibitor glycogen synthase kinase 3β (GSK3β) in a DA neuronal cell line SN4741. Furthermore, B3C effectively rescued MEF2D from MPP(+)-induced decline in both nucleic and mitochondrial compartments. B3C offered SN4741 cells potent protection against MPP(+)-induced apoptosis via MEF2D. Interestingly, B3C also protected SN4741 cells from wild type or mutant A53T α-synuclein-induced cytotoxicity. Using the in vivo PD model of C57BL/6 mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP), we showed that B3C maintained redox homeostasis, promoted Akt function activity, and restored MEF2D level in midbrain neurons. Moreover, B3C greatly prevented the loss of tyrosine hydroxylase signal in substantial nigra pars compacta DA neurons and ameliorated behavioral impairments in mice treated with MPTP. Collectedly, our studies identified B3C as a potent neuroprotective agent whose effectiveness relies on its ability to effectively up-regulate MEF2D in DA neurons against toxic stress in models of PD in vitro and in vivo.

  20. The p38 MAP kinase inhibitor SB203580 enhances nuclear factor-kappa B transcriptional activity by a non-specific effect upon the ERK pathway

    NARCIS (Netherlands)

    Birkenkamp, KU; Tuyt, LML; Lummen, C; Wierenga, LTJ; Kruijer, W; Vellenga, E

    2000-01-01

    1 In the present study we investigated a possible role for the p38 mitogen-activated protein (MAP) kinase pathway in mediating nuclear factor-kappa B (NF-kappa B) transcriptional activity in the erythroleukaemic cell line TF-1. 2 TF-1 cells stimulated with the phosphatase inhibitor okadaic acid (OA)

  1. Activating transcription factor 6 poly morphisms and haplotypes ars associated with impaired glucose homeostasis ans type 2 diabetis in dutch Caucasians

    NARCIS (Netherlands)

    Meex, S.J.; Greevenbroek, van M.M.J.; Ayoubi, T.A.; Vlietinck, R.; Vliet-Ostaptchouk, J.V.; Hofker, M.H.; Vermeulen, V.; Schalkwijk, C.G.; Feskens, E.J.M.; Boer, J.M.A.; Stehouwer, C.D.A.; Kallen, van der C.J.H.; Bruin, de T.W.A.

    2007-01-01

    Context: Activating transcription factor 6 (ATF6) is critical for initiation and full activation of the unfolded protein response. An association between genetic variation in ATF6 and type 2 diabetes (DM2) was recently reported in Pima Indians. Objectives: To investigate the broader significance of

  2. 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.

  3. Protein Kinases and Transcription Factors Activation in Response to UV-Radiation of Skin: Implications for Carcinogenesis

    Directory of Open Access Journals (Sweden)

    Laurence A. Marchat

    2011-12-01

    Full Text Available Solar ultraviolet (UV radiation is an important environmental factor that leads to immune suppression, inflammation, photoaging, and skin carcinogenesis. Here, we reviewed the specific signal transduction pathways and transcription factors involved in the cellular response to UV-irradiation. Increasing experimental data supporting a role for p38, MAPK, JNK, ERK1/2, and ATM kinases in the response network to UV exposure is discussed. We also reviewed the participation of NF-κB, AP-1, and NRF2 transcription factors in the control of gene expression after UV-irradiation. In addition, we discussed the promising chemotherapeutic intervention of transcription factors signaling by natural compounds. Finally, we focused on the review of data emerging from the use of DNA microarray technology to determine changes in global gene expression in keratinocytes and melanocytes in response to UV treatment. Efforts to obtain a comprehensive portrait of the transcriptional events regulating photodamage of intact human epidermis after UV exposure reveals the existence of novel factors participating in UV-induced cell death. Progress in understanding the multitude of mechanisms induced by UV-irradiation could lead to the potential use of protein kinases and novel proteins as specific targets for the prevention and control of skin cancer.

  4. Signal transduction mechanisms involved in S100A4-induced activation of the transcription factor NF-κB

    Directory of Open Access Journals (Sweden)

    Mælandsmo Gunhild M

    2010-05-01

    Full Text Available Abstract Background The metastasis-promoting protein S100A4 activates the transcription factor NF-κB through the classical NF-κB activation pathway. The upstream signal transduction mechanisms leading to increased NF-κB activity are, however, incompletely characterized. Methods The human osteosarcoma cell line II-11b was stimulated with recombinant S100A4 in the presence or absence of inhibitors of common signal transduction pathways, and NF-κB activity was examined using a luciferase-based reporter assay and phosphorylation of IκBα. mRNA expression was analyzed by real-time RT-PCR, protein expression was examined by Western blotting and IKK activity was measured using an in vitro kinase assay. The role of upstream kinases and the cell surface receptor RAGE was investigated by overexpression of dominant negative proteins and by siRNA transfection. Results The Ser/Thr kinase inhibitors H-7 and staurosporine inhibited S100A4-induced IκBα phosphorylation and subsequent NF-κB activation. The protein tyrosine kinase inhibitor genistein and the phospholipase C inhibitor compound 48/80 had a partial inhibitory effect on IκBα phosphorylation, whereas inhibitors of protein kinase C, G-protein coupled receptors and PI 3-kinases had no effect on the level of phosphorylation. Interestingly, S100A4 treatment induced activating phosphorylations of IKKα/β, but neither H-7 nor staurosporine was able to significantly inhibit IKK activation. Dominant negative MEKK1 or NIK did not inhibit S100A4-induced NF-κB activity, and S100A4 stimulation did not influence AKT phosphorylation. Furthermore, diminished expression of the putative S100 protein receptor RAGE did not affect the observed phosphorylation of IκBα. Conclusions S100A4 activates NF-κB by inducing phosphorylation of IKKα/β, leading to increased IκBα phosphorylation. The Ser/Thr kinase inhibitors H-7 and staurosporine attenuated S100A4-induced NF-κB activation and inhibited IKK

  5. FGF19 (fibroblast growth factor 19) as a novel target gene for activating transcription factor 4 in response to endoplasmic reticulum stress.

    Science.gov (United States)

    Shimizu, Makoto; Li, Juan; Maruyama, Ryuto; Inoue, Jun; Sato, Ryuichiro

    2013-02-15

    FGF19 (fibroblast growth factor 19), expressed in the small intestine, acts as an enterohepatic hormone by mediating inhibitory effects on the bile acid synthetic pathway and regulating carbohydrate and lipid metabolism. In an attempt to identify novel agents other than bile acids that induce increased FGF19 expression, we found that some ER (endoplasmic reticulum) stress inducers were effective. When intestinal epithelial Caco-2 cells were incubated with thapsigargin, marked increases were observed in the mRNA and secreted protein levels of FGF19. This was not associated with the farnesoid X receptor. Reporter gene analyses using the 5'-promoter region of FGF19 revealed that a functional AARE (amino-acid-response element) was localized in this region, and this site was responsible for inducing its transcription through ATF4 (activating transcription factor 4), which is activated in response to ER stress. EMSAs (electrophoretic mobility-shift assays) and ChIP (chromatin immunoprecipitation) assays showed that ATF4 bound to this site and enhanced FGF19 expression. Overexpression of ATF4 in Caco-2 cells induced increased FGF19 mRNA expression, whereas shRNA (short hairpin RNA)-mediated depletion of ATF4 significantly attenuated a thapsigargin-induced increase in FGF19 mRNA.

  6. Arabidopsis IRE1 catalyses unconventional splicing of bZIP60 mRNA to produce the active transcription factor

    KAUST Repository

    Nagashima, Yukihiro

    2011-07-01

    IRE1 plays an essential role in the endoplasmic reticulum (ER) stress response in yeast and mammals. We found that a double mutant of Arabidopsis IRE1A and IRE1B (ire1a/ire1b) is more sensitive to the ER stress inducer tunicamycin than the wild-type. Transcriptome analysis revealed that genes whose induction was reduced in ire1a/ire1b largely overlapped those in the bzip60 mutant. We observed that the active form of bZIP60 protein detected in the wild-type was missing in ire1a/ire1b. We further demonstrated that bZIP60 mRNA is spliced by ER stress, removing 23 ribonucleotides and therefore causing a frameshift that replaces the C-terminal region of bZIP60 including the transmembrane domain (TMD) with a shorter region without a TMD. This splicing was detected in ire1a and ire1b single mutants, but not in the ire1a/ire1b double mutant. We conclude that IRE1A and IRE1B catalyse unconventional splicing of bZIP60 mRNA to produce the active transcription factor.

  7. 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...

  8. DNA residence time is a regulatory factor of transcription repression.

    Science.gov (United States)

    Clauß, Karen; Popp, Achim P; Schulze, Lena; Hettich, Johannes; Reisser, Matthias; Escoter Torres, Laura; Uhlenhaut, N Henriette; Gebhardt, J Christof M

    2017-08-21

    Transcription comprises a highly regulated sequence of intrinsically stochastic processes, resulting in bursts of transcription intermitted by quiescence. In transcription activation or repression, a transcription factor binds dynamically to DNA, with a residence time unique to each factor. Whether the DNA residence time is important in the transcription process is unclear. Here, we designed a series of transcription repressors differing in their DNA residence time by utilizing the modular DNA binding domain of transcription activator-like effectors (TALEs) and varying the number of nucleotide-recognizing repeat domains. We characterized the DNA residence times of our repressors in living cells using single molecule tracking. The residence times depended non-linearly on the number of repeat domains and differed by more than a factor of six. The factors provoked a residence time-dependent decrease in transcript level of the glucocorticoid receptor-activated gene SGK1. Down regulation of transcription was due to a lower burst frequency in the presence of long binding repressors and is in accordance with a model of competitive inhibition of endogenous activator binding. Our single molecule experiments reveal transcription factor DNA residence time as a regulatory factor controlling transcription repression and establish TALE-DNA binding domains as tools for the temporal dissection of transcription regulation. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  9. 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.

  10. Interaction of Restin with transcription factors

    Institute of Scientific and Technical Information of China (English)

    WU; Yousheng; LU; Fan; QI; Yinxin; WANG; Ruihua; ZHANG; Jia

    2005-01-01

    Restin, a member of melanoma-associated antigen superfamily gene, was first cloned from differentiated leukemia cell induced by all trans-retinoic acid, and was able to inhibit cell proliferation, but the molecular mechanism was not clear. Since Restin was localized in cell nucleus, and its homolog member, Necdin (neuronal growth suppressor factor), could interact with transcription factors p53 and E2F1, we proposed that Restin might also function as Necdin through interacting with some transcription factors. In this study, transcription factors p53, AP1,ATFs and E2Fs were cloned and used in the mammalian two-hybrid system to identify their interaction with Restin. The results showed that only ATF3 had a strong interaction with Restin. It is interesting to know that ATF3 was an important transcription factor for G1 cell cycle initiation in physiological stress response. It was possible that the inhibition of cell proliferation by Restin might be related with the inhibition of ATF3 activity.

  11. Possible Involvement of Nitric Oxide and Reactive Oxygen Species in Glucose Deprivation-Induced Activation of Transcription Factor Rst2

    OpenAIRE

    Toshiaki Kato; Xin Zhou; Yan Ma

    2013-01-01

    Glucose is one of the most important sources of cellular nutrition and glucose deprivation induces various cellular responses. In Schizosaccharomyces pombe, zinc finger protein Rst2 is activated upon glucose deprivation, and regulates gene expression via the STREP (stress response element of Schizosaccharomyces pombe) motif. However, the activation mechanism of Rst2 is not fully understood. We monitored Rst2 transcriptional activity in living cells using a Renilla luciferase reporter system. ...

  12. Differential expression of brain-derived neurotrophic factor transcripts after pilocarpine-induced seizure-like activity is related to mode of Ca2+ entry.

    Science.gov (United States)

    Poulsen, F R; Lauterborn, J; Zimmer, J; Gall, C M

    2004-01-01

    Activity-dependent brain-derived neurotrophic factor (BDNF) expression is Ca2+-dependent, yet little is known about the Ca2+ channel contributions that might direct selective expression of the multiple BDNF transcripts. Here, effects of pilocarpine-induced seizure activity on total BDNF expression and on the individual sensitivity of BDNF transcripts to glutamate receptor and Ca2+ channel blockers were evaluated using hippocampal slice cultures and in situ hybridization of transcript-specific cRNA probes directed against mRNAs for the four 5' exons (I-IV) of the BDNF gene. mRNAs for nerve growth factor (NGF) and tyrosine kinase B (trkB) also were studied. Pilocarpine (5 mM) induced a dose- and time-dependent increase in total BDNF (exon V) mRNA expression in the dentate granule cells and CA3-CA1 pyramidal cells with maximal effects at 6 and 24 h, respectively. Increases were blocked by co-treatment with the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid/kainate 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX: 25 microM) and the N-methyl-d-aspartic acid receptor antagonist 2-amino-5-phosphonovaleric acid (APV; 25 microM), whereas the L-type voltage sensitive Ca2+ channel blocker nifedipine (20 microM) was without detectable effect. Maximal NGF and trkB mRNA expression was induced by pilocarpine at 4 and 12 h, respectively. For the individual BDNF transcripts, APV blocked pilocarpine-induced increases in transcript II, whereas nifedipine blocked increases in transcripts I and III. Transcript IV levels were not altered by treatment. These results indicate that transcript II makes the greatest contribution to pilocarpine effects on total BDNF mRNA content in this model and provides evidence for regional and Ca2+ channel-specific differences in activity-dependent regulation of the different BDNF transcripts in hippocampus.

  13. Polymorphisms in SREBF1 and SREBF2, two antipsychotic-activated transcription factors controlling cellular lipogenesis, are associated with schizophrenia in German and Scandinavian samples

    DEFF Research Database (Denmark)

    Le Hellard, S; Mühleisen, T W; Djurovic, S

    2010-01-01

    Several studies have reported structural brain abnormalities, decreased myelination and oligodendrocyte dysfunction in schizophrenia. In the central nervous system, glia-derived de novo synthesized cholesterol is essential for both myelination and synaptogenesis. Previously, we demonstrated...... in glial cell lines that antipsychotic drugs induce the expression of genes involved in cholesterol and fatty acids biosynthesis through activation of the sterol regulatory element binding protein (SREBP) transcription factors, encoded by the sterol regulatory element binding transcription factor 1 (SREBF1......) and sterol regulatory element binding transcription factor 2 (SREBF2) genes. Considering the importance of these factors in the lipid biosynthesis and their possible involvement in antipsychotic drug effects, we hypothesized that genetic variants of SREBF1 and/or SREBF2 could affect schizophrenia...

  14. Rac1/p21-activated kinase pathway controls retinoblastoma protein phosphorylation and E2F transcription factor activation in B lymphocytes.

    Science.gov (United States)

    Zaldua, Natalia; Llavero, Francisco; Artaso, Alain; Gálvez, Patricia; Lacerda, Hadriano M; Parada, Luis A; Zugaza, José L

    2016-02-01

    Small GTPases of the Ras superfamily are capable of activating E2F-dependent transcription leading to cell proliferation, but the molecular mechanisms are poorly understood. In this study, using immortalized chicken DT40 B cell lines to investigate the role of the Vav/Rac signalling cascade on B cell proliferation, it is shown that the proliferative response triggered by B cell receptor activation is dramatically reduced in the absence of Vav3 expression. Analysis of this proliferative defect shows that in the absence of Vav3 expression, retinoblastoma protein (RB) phosphorylation and the subsequent E2F activation do not take place. By combining pharmacological and genetic approaches, phosphatidylinositol-3-kinase and phospholipase Cγ2 (PLCγ2) were identified as the key regulatory signalling molecules upstream of the Vav3/Rac pathway leading to RB phosphorylation and E2F transcription factor activation. Additionally, vav3(-/-) and plcγ2(-/-) DT40 B cells were not able to activate the RB-E2F complex wild-type phenotype when these genetically modified cells were transfected with constitutively active forms of RhoA or Cdc42. However, when these knockout cells were transfected with different constitutively active versions of PLCγ, Vav or Rac1, not only activation of the RB-E2F complex wild-type phenotype was recovered but also the cellular proliferation. Furthermore, by evaluating the effect of two known effector mutants of Rac1 (Rac1(Q61L/F37A) and Rac1(Q61L/Y40C) ), the RB-E2F complex activation dependency on p21-activated kinase (PAK) and protein kinase Cε (PKCε) activities was established, being independent of both actin cytoskeleton reorganization and Ras activity. These results suggest that PAK1 and PKCε may be potential therapeutic targets to stop uncontrolled B cell proliferation mediated by the Vav/Rac pathway.

  15. Mechanism for transcriptional synergy between interferon regulatory factor (IRF)-3 and IRF-7 in activation of the interferon-beta gene promoter.

    Science.gov (United States)

    Yang, Hongmei; Ma, Gang; Lin, Charles H; Orr, Melissa; Wathelet, Marc G

    2004-09-01

    The interferon-beta promoter has been studied extensively as a model system for combinatorial transcriptional regulation. In virus-infected cells the transcription factors ATF-2, c-Jun, interferon regulatory factor (IRF)-3, IRF-7 and NF-kappaB, and the coactivators p300/CBP play critical roles in the activation of this and other promoters. It remains unclear, however, why most other combinations of AP-1, IRF and Rel proteins fail to activate the interferon-beta gene. Here we have explored how different IRFs may cooperate with other factors to activate transcription. First we showed in undifferentiated embryonic carcinoma cells that ectopic expression of either IRF-3 or IRF-7, but not IRF-1, was sufficient to allow virus-dependent activation of the interferon-beta promoter. Moreover, the activity of IRF-3 and IRF-7 was strongly affected by promoter context, with IRF-7 preferentially being recruited to the natural interferon-beta promoter. We fully reconstituted activation of this promoter in insect cells. Maximal synergy required IRF-3 and IRF-7 but not IRF-1, and was strongly dependent on the presence of p300/CBP, even when these coactivators only modestly affected the activity of each factor by itself. These results suggest that specificity in activation of the interferon-beta gene depends on a unique promoter context and on the role played by coactivators as architectural factors. Copyright 2004 FEBS

  16. Pregnenolone sulfate activates basic region leucine zipper transcription factors in insulinoma cells: role of voltage-gated Ca2+ channels and transient receptor potential melastatin 3 channels.

    Science.gov (United States)

    Müller, Isabelle; Rössler, Oliver G; Thiel, Gerald

    2011-12-01

    The neurosteroid pregnenolone sulfate activates a signaling cascade in insulinoma cells involving activation of extracellular signal-regulated protein kinase and enhanced expression of the transcription factor Egr-1. Here, we show that pregnenolone sulfate stimulation leads to a significant elevation of activator protein-1 (AP-1) activity in insulinoma cells. Expression of the basic region leucine zipper (bZIP) transcription factors c-Jun and c-Fos is up-regulated in insulinoma cells and pancreatic β-cells in primary culture after pregnenolone sulfate stimulation. Up-regulation of a chromatin-embedded c-Jun promoter/luciferase reporter gene transcription in pregnenolone sulfate-stimulated insulinoma cells was impaired when the AP-1 binding sites were mutated, indicating that these motifs function as pregnenolone sulfate response elements. In addition, phosphorylation of cAMP response element (CRE)-binding protein is induced and transcription of a CRE-controlled reporter gene is stimulated after pregnenolone sulfate treatment, indicating that the CRE functions as a pregnenolone sulfate response element as well. Pharmacological and genetic experiments revealed that both L-type Ca(2+) channels and transient receptor potential melastatin 3 (TRPM3) channels are essential for connecting pregnenolone sulfate stimulation with enhanced AP-1 activity and bZIP-mediated transcription in insulinoma cells. In contrast, pregnenolone sulfate stimulation did not enhance AP-1 activity or c-Jun and c-Fos expression in pituitary corticotrophs that express functional L-type Ca(2+) channels but only trace amounts of TRPM3. We conclude that expression of L-type Ca(2+) channels is not sufficient to activate bZIP-mediated gene transcription by pregnenolone sulfate. Rather, additional expression of TRPM3 or depolarization of the cells is required to connect pregnenolone sulfate stimulation with enhanced gene transcription.

  17. 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.

  18. Influence of the adipose derived hormone resistin onsignal transducer and activator of transcription factors, steroidogenesis and proliferation of Leydig cells

    Institute of Scientific and Technical Information of China (English)

    Stephanie Jean; David Landry; Mikella Daigle; Luc J. Martin

    2012-01-01

    Objective:To explore whether resistin, an adipose derived hormone linked to insulin resistance, influence steroidogenic genes expressions andLeydig cells function or not.Methods:Various Leydig cell lines were exposed to increasing doses of resistin with or without cAMP.Changes were monitored at the protein level for signal transducer and activator of transcription(STAT) factors and steroidogenic components, steroidogenic acute regulatory protein(STAR) and cholesterol side-chain cleavage enzyme(CYP11A1), for progesterone production and cell viability.Results:Resistin had no effect on progesterone production, despite an increase in nuclear translocation ofSTAT1,STAT3 andSTAT5 and unexpected synergy with cAMP in the synthesis ofSTAR and CYP11A1.In addition, exposure to normal levels of resistin(10 ng/mL) seemed to have beneficial effects onLeydig cell function, as it increased cells viability and proliferation.Conclusions:Our results suggest that resistin may function as an endocrine mediator linking metabolism and male reproduction.

  19. β-Catenin-independent activation of TCF1/LEF1 in human hematopoietic tumor cells through interaction with ATF2 transcription factors.

    Directory of Open Access Journals (Sweden)

    Luca Grumolato

    Full Text Available The role of Wnt signaling in embryonic development and stem cell maintenance is well established and aberrations leading to the constitutive up-regulation of this pathway are frequent in several types of human cancers. Upon ligand-mediated activation, Wnt receptors promote the stabilization of β-catenin, which translocates to the nucleus and binds to the T-cell factor/lymphoid enhancer factor (TCF/LEF family of transcription factors to regulate the expression of Wnt target genes. When not bound to β-catenin, the TCF/LEF proteins are believed to act as transcriptional repressors. Using a specific lentiviral reporter, we identified hematopoietic tumor cells displaying constitutive TCF/LEF transcriptional activation in the absence of β-catenin stabilization. Suppression of TCF/LEF activity in these cells mediated by an inducible dominant-negative TCF4 (DN-TCF4 inhibited both cell growth and the expression of Wnt target genes. Further, expression of TCF1 and LEF1, but not TCF4, stimulated TCF/LEF reporter activity in certain human cell lines independently of β-catenin. By a complementary approach in vivo, TCF1 mutants, which lacked the ability to bind to β-catenin, induced Xenopus embryo axis duplication, a hallmark of Wnt activation, and the expression of the Wnt target gene Xnr3. Through generation of different TCF1-TCF4 fusion proteins, we identified three distinct TCF1 domains that participate in the β-catenin-independent activity of this transcription factor. TCF1 and LEF1 physically interacted and functionally synergized with members of the activating transcription factor 2 (ATF2 family of transcription factors. Moreover, knockdown of ATF2 expression in lymphoma cells phenocopied the inhibitory effects of DN-TCF4 on the expression of target genes associated with the Wnt pathway and on cell growth. Together, our findings indicate that, through interaction with ATF2 factors, TCF1/LEF1 promote the growth of hematopoietic malignancies in the

  20. 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.

  1. Dual functions of the nucleus-encoded factor TDA1 in trapping and translation activation of atpA transcripts in Chlamydomonas reinhardtii chloroplasts.

    Science.gov (United States)

    Eberhard, Stephan; Loiselay, Christelle; Drapier, Dominique; Bujaldon, Sandrine; Girard-Bascou, Jacqueline; Kuras, Richard; Choquet, Yves; Wollman, Francis-André

    2011-09-01

    After endosymbiosis, organelles lost most of their initial genome. Moreover, expression of the few remaining genes became tightly controlled by the nucleus through trans-acting protein factors that are required for post-transcriptional expression (maturation/stability or translation) of a single (or a few) specific organelle target mRNA(s). Here, we characterize the nucleus-encoded TDA1 factor, which is specifically required for translation of the chloroplast atpA transcript that encodes subunit α of ATP synthase in Chlamydomonas reinhardtii. The sequence of TDA1 contains eight copies of a degenerate 38-residue motif, that we named octotrico peptide repeat (OPR), which has been previously described in a few other trans-acting factors targeted to the C. reinhardtii chloroplast. Interestingly, a proportion of the untranslated atpA transcripts are sequestered into high-density, non-polysomic, ribonucleoprotein complexes. Our results suggest that TDA1 has a dual function: (i) trapping a subset of untranslated atpA transcripts into non-polysomic complexes, and (ii) translational activation of these transcripts. We discuss these results in light of our previous observation that only a proportion of atpA transcripts are translated at any given time in the chloroplast of C. reinhardtii.

  2. Lipid Synthetic Transcription Factor SREBP-1a Activates p21WAF1/CIP1, a Universal Cyclin-Dependent Kinase Inhibitor

    OpenAIRE

    INOUE, Noriyuki; Shimano, Hitoshi; Nakakuki, Masanori; Matsuzaka, Takashi; Nakagawa, Yoshimi; Yamamoto, Takashi; Sato, Ryuichiro; Takahashi, Akimitsu; Sone, Hirohito; Yahagi, Naoya; Suzuki, Hiroaki; Toyoshima, Hideo; Yamada, Nobuhiro

    2005-01-01

    Sterol regulatory element-binding proteins (SREBPs) are membrane-bound transcription factors that regulate lipid synthetic genes. In contrast to SREBP-2, which regulates cellular cholesterol level in normal cells, SREBP-1a is highly expressed in actively growing cells and activates entire programs of genes involved in lipid synthesis such as cholesterol, fatty acids, triglycerides, and phospholipids. Previously, the physiological relevance of this potent activity of SREBP-1a has been thought ...

  3. Transcriptional activation is a conserved feature of the early embryonic factor Zelda that requires a cluster of four zinc fingers for DNA binding and a low-complexity activation domain.

    Science.gov (United States)

    Hamm, Danielle C; Bondra, Eliana R; Harrison, Melissa M

    2015-02-01

    Delayed transcriptional activation of the zygotic genome is a nearly universal phenomenon in metazoans. Immediately following fertilization, development is controlled by maternally deposited products, and it is not until later stages that widespread activation of the zygotic genome occurs. Although the mechanisms driving this genome activation are currently unknown, the transcriptional activator Zelda (ZLD) has been shown to be instrumental in driving this process in Drosophila melanogaster. Here we define functional domains of ZLD required for both DNA binding and transcriptional activation. We show that the C-terminal cluster of four zinc fingers mediates binding to TAGteam DNA elements in the promoters of early expressed genes. All four zinc fingers are required for this activity, and splice isoforms lacking three of the four zinc fingers fail to activate transcription. These truncated splice isoforms dominantly suppress activation by the full-length, embryonically expressed isoform. We map the transcriptional activation domain of ZLD to a central region characterized by low complexity. Despite relatively little sequence conservation within this domain, ZLD orthologs from Drosophila virilis, Anopheles gambiae, and Nasonia vitripennis activate transcription in D. melanogaster cells. Transcriptional activation by these ZLD orthologs suggests that ZLD functions through conserved interactions with a protein cofactor(s). We have identified distinct DNA-binding and activation domains within the critical transcription factor ZLD that controls the initial activation of the zygotic genome.

  4. Rapid, sequential activation of mitogen-activated protein kinases and transcription factors precedes proinflammatory cytokine mRNA expression in spleens of mice exposed to the trichothecene vomitoxin.

    Science.gov (United States)

    Zhou, Hui-Ren; Islam, Zahidul; Pestka, James J

    2003-03-01

    Since proinflammatory cytokine mRNA expression is induced within lymphoid tissue in vivo by the trichothecene vomitoxin (VT) in a rapid (1-2 h) and transient (4-8 h) fashion, it was hypothesized that mitogen-activated protein kinases (MAPKs) and transcription factors associated upstream with gene transcription of these cytokines are activated prior to or within these time windows. To test this hypothesis, mice were first treated with a single oral dose of VT and then analyzed for MAPK phosphorylation in the spleen. As little as 1 mg/kg of VT induced JNK 1/2, ERK 1/2, and p38 phosphorylation with maximal effects being observed at 5 to 100 mg/kg of VT. VT transiently induced JNK and p38 phosphorylation over a 60-min time period with peak effects being observed at 15 and 30 min, respectively. In contrast, ERK remained phosphorylated from 15 to 120 min. Next, the binding of activating protein 1 (AP-1), CCAAT enhancer-binding protein (C/EBP), CRE-binding protein (CREB), and nuclear factor-kappaB (NF-kappaB) was measured by electrophoretic mobility shift assay (EMSA) using four different consensus transcriptional control motifs at 0, 0.5, 1.5, 4, and 8 h after oral exposure to 25 mg/kg of VT. AP-1 binding activity was differentially elevated from 0.5 h to 8 h, whereas C/EBP binding was elevated only at 0.5 h. CREB binding decreased slightly at 0.5 h but gradually increased, reaching a maximum at 4 h. NF-kappaB binding was increased only slightly at 4 and 8 h. The specificities of AP-1, C/EBP, CREB, and NF-kappaB for relevant DNA motifs were verified by competition assays, using an excess of unlabeled consensus and mutant oligonucleotides. Supershift EMSAs and Western blot analysis identified specific VT-inducible DNA binding proteins for AP-1 (cJun, phospho c-jun, JunB, and JunD), C/EBP (C/EBPbeta), CREB (CREB-1 and ATF-2), and NF-kappaB (p50 and cRel). Finally, when the effects of oral VT exposure on proinflammatory gene expression were assessed at 3, 6, and 9 h

  5. 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.

  6. Insights into transcription enhancer factor 1 (TEF-1) activity from the solution structure of the TEA domain.

    Science.gov (United States)

    Anbanandam, Asokan; Albarado, Diana C; Nguyen, Catherine T; Halder, Georg; Gao, Xiaolian; Veeraraghavan, Sudha

    2006-11-14

    Transcription enhancer factor 1 is essential for cardiac, skeletal, and smooth muscle development and uses its N-terminal TEA domain (TEAD) to bind M-CAT elements. Here, we present the first structure of TEAD and show that it is a three-helix bundle with a homeodomain fold. Structural data reveal how TEAD binds DNA. Using structure-function correlations, we find that the L1 loop is essential for cooperative loading of TEAD molecules on to tandemly duplicated M-CAT sites. Furthermore, using a microarray chip-based assay, we establish that known binding sites of the full-length protein are only a subset of DNA elements recognized by TEAD. Our results provide a model for understanding the regulation of genome-wide gene expression during development by TEA/ATTS family of transcription factors.

  7. The LIM homeobox transcription factor Lhx2 is required to specify the retina field and synergistically cooperates with Pax6 for Six6 trans-activation.

    Science.gov (United States)

    Tétreault, Nicolas; Champagne, Marie-Pier; Bernier, Gilbert

    2009-03-15

    In mammals, a limited set of homeobox-containing transcription factors are expressed in the presumptive eye field and required to initiate eye development. How these factors interact together at the genetic and molecular level to coordinate this developmental process is poorly understood. We found that the Lhx2 and Pax6 transcription factors operate in a concerted manner during retinal development to promote transcriptional activation of the Six6 homeobox-gene in primitive and mature retinal progenitors. Lhx2 demarcates the presumptive retina field at the neural plate stage and Lhx2 inactivation delays initiation of Rx, Six3 and Pax6 expression in this domain. The later expressed Six6 is properly activated in the pituitary/hypothalamic axis of Lhx2(-/-) embryos, but expression fails to be initiated in the optic vesicle. Lhx2 and Pax6 associate with the chromatin at several regions of Six6 in vivo and cooperate for trans-activation of Six6 regulatory elements in vitro. In retinal progenitor/stem cells, both Lhx2 and Pax6 are genetically required for proper Six6 expression and forced co-expression of Lhx2 and Pax6 can synergistically trans-activate the Six6 locus. Our work reveals how two master regulators of eye development coordinate their action to sequentially promote tissue-specific transcriptional initiation and full activation of a retinal determinant gene.

  8. Activation of Hypodermal Differentiation in the Caenorhabditis elegans Embryo by GATA Transcription Factors ELT-1 and ELT-3

    OpenAIRE

    Gilleard, J.S.; McGhee, J D

    2001-01-01

    The Caenorhabditis elegans GATA transcription factor genes elt-1 and elt-3 are expressed in the embryonic hypodermis (also called the epidermis). elt-1 is expressed in precursor cells and is essential for the production of most hypodermal cells (22). elt-3 is expressed in all of the major hypodermal cells except the lateral seam cells, and expression is initiated immediately after the terminal division of precursor lineages (13). Although this expression pattern suggests a role for ELT-3 in h...

  9. 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.

  10. Trm1p, a Zn(II)₂Cys₆-type transcription factor, is essential for the transcriptional activation of genes of methanol utilization pathway, in Pichia pastoris.

    Science.gov (United States)

    Sahu, Umakant; Krishna Rao, Kamisetty; Rangarajan, Pundi N

    2014-08-15

    The zinc finger transcription factors Mxr1p and Rop are key regulators of methanol metabolism in the methylotrophic yeast, Pichia pastoris, while Trm1p and Trm2p regulate methanol metabolism in Candida boidinii. Here, we demonstrate that Trm1p is essential for the expression of genes of methanol utilization (mut) pathway in P. pastoris as well. Expression of AOXI and other genes of mut pathway is severely compromised in P. pastoris ΔTrm1 strain resulting in impaired growth on media containing methanol as the sole source of carbon. Trm1p localizes to the nucleus of cells cultured on glucose or methanol. The zinc finger domain of Mxr1p but not Trm1p binds to AOXI promoter sequences in vitro, indicating that these two positive regulators act by different mechanisms. We conclude that both Trm1p and Mxr1p are essential for the expression of genes of mut pathway in P. pastoris and the mechanism of transcriptional regulation of mut pathway may be similar in P. pastoris and C. boidinii.

  11. Extent of liver resection modulates the activation of transcription factors and the production of cytokines involved in liver regeneration

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    AIM: To investigate the molecular events involved in liver regeneration following subtotal hepatectomy (SH) as previous studies have largely focused on partial hepatectomy (PH).METHODS: Male Wistar rats were subjected to 70% PH or 90% SH, respectively, and sacrificed at different times after surgery. Untreated and sham-operated animals served as controls. Serum and liver samples were obtained to investigate liver function, apoptosis (TUNEL assay) and transcription factors (NF-κB, Stat3; ELISA) or cytokines (HGF, TNF-α, IL-6, TGF-α, TGF-β; quantitative RT-PCR) involved in liver regeneration. RESULTS: Serum levels of ALT and AST in animals with 70% PH differed significantly from sham-operated and control animals. We found that the peak concentration 12 h after surgery returned to control levels 7 d after surgery. LDH was increased only at 12 h after 70% PH compared to sham. Bilirubin showed no differences between the sham and 70% resection. After PH, early NF-κB activation was detected 12 h after surgery (313.21 ± 17.22 ng/mL), while there was no activation after SH (125.22 ± 44.36 ng/mL) compared to controls (111.43 ± 32.68 ng/mL) at this time point. In SH, however, NF-κB activation was delayed until 24 h (475.56 ± 144.29 ng/mL). Star3 activation was similar in both groups. These findings correlated with suppressed and delayed induction of regenerative genes after SH (i.e. TNF-α 24 h postoperatively: 2375 ± 1220 in 70% and 88±31 in 90%; IL-6 12 h postoperatively: 2547 ± 441 in 70% and 173 ± 82 in 90%). TUNEL staining revealed elevated apoptosis rates in SH (0.44% at 24 h; 0.63% at 7 d) compared to PH (0.27% at 24 h; 0.15% at 7 d). CONCLUSION: The molecular events involved in liver regeneration are significantly influenced by the extent of resection as SH leads to suppression and delay of liver regeneration compared to PH, which is associated with delayed activation of NF-κB and suppression of proregenerative cytokines.

  12. DBD: a transcription factor prediction database.

    Science.gov (United States)

    Kummerfeld, Sarah K; Teichmann, Sarah A

    2006-01-01

    Regulation of gene expression influences almost all biological processes in an organism; sequence-specific DNA-binding transcription factors are critical to this control. For most genomes, the repertoire of transcription factors is only partially known. Hitherto transcription factor identification has been largely based on genome annotation pipelines that use pairwise sequence comparisons, which detect only those factors similar to known genes, or on functional classification schemes that amalgamate many types of proteins into the category of 'transcription factor'. Using a novel transcription factor identification method, the DBD transcription factor database fills this void, providing genome-wide transcription factor predictions for organisms from across the tree of life. The prediction method behind DBD identifies sequence-specific DNA-binding transcription factors through homology using profile hidden Markov models (HMMs) of domains. Thus, it is limited to factors that are homologus to those HMMs. The collection of HMMs is taken from two existing databases (Pfam and SUPERFAMILY), and is limited to models that exclusively detect transcription factors that specifically recognize DNA sequences. It does not include basal transcription factors or chromatin-associated proteins, for instance. Based on comparison with experimentally verified annotation, the prediction procedure is between 95% and 99% accurate. Between one quarter and one-half of our genome-wide predicted transcription factors represent previously uncharacterized proteins. The DBD (www.transcriptionfactor.org) consists of predicted transcription factor repertoires for 150 completely sequenced genomes, their domain assignments and the hand curated list of DNA-binding domain HMMs. Users can browse, search or download the predictions by genome, domain family or sequence identifier, view families of transcription factors based on domain architecture and receive predictions for a protein sequence.

  13. 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

  14. Teratogenic factors affect transcription factor expression.

    Science.gov (United States)

    Kojima, Takuya; Asano, Shinya; Takahashi, Naoki

    2013-01-01

    Chemical compounds are produced every day, many with adverse effects on human health, and hence it is vital to predict the risks to humans simply, rapidly, and accurately. Teratogens have a serious impact on fetal development. This has been studied mainly by phenotypic analysis of experimental animals. However, since phenotypes can vary within different species, we established a new evaluation system based on our recent finding that teratogens influence Hox gene expression in mice. Similarly to the Hox gene expression changes, the expression patterns of several transcription factors involved in development, including the Dlx, Irx, Sall, and T-box families, were altered after 6 h of exposure to retinoic acid (RA) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The expression changes in Dlx4, Dlx6, Irx5, Sall2, Sall3, Sall4, Tbx10, and Tbx22 were linked to teratogen-induced phenotypes, and our results indicate that expression changes in developmental transcription factors can help to predict teratogenic risk.

  15. The Etched1 gene of Zea mays (L.) encodes a zinc ribbon protein that belongs to the transcriptionally active chromosome (TAC) of plastids and is similar to the transcription factor TFIIS.

    Science.gov (United States)

    da Costa e Silva, Oswaldo; Lorbiecke, René; Garg, Preeti; Müller, Lenard; Wassmann, Martina; Lauert, Patricia; Scanlon, Mike; Hsia, An-Ping; Schnable, Patrick S; Krupinska, Karin; Wienand, Udo

    2004-06-01

    Etched1 (et1) is a pleiotropic, recessive mutation of maize that causes fissured and cracked mature kernels and virescent seedlings. Microscopic examinations of the et1 phenotype revealed an aberrant plastid development in mutant kernels and mutant leaves. Here, we report on the cloning of the et1 gene by transposon tagging, the localization of the gene product in chloroplasts, and its putative function in the plastid transcriptional apparatus. Several alleles of Mutator (Mu)-induced et1 mutants, the et1-reference (et1-R) mutant, and Et1 wild-type were cloned and analyzed at the molecular level. Northern analyses with wild-type plants revealed that Et1 transcripts are present in kernels, leaves, and other types of tissue, and no Et1 expression could be detected in the et1 mutants analyzed. The ET1 protein is imported by chloroplasts and has been immunologically detected in transcriptionally active chromosome (TAC) fractions derived from chloroplasts. Accordingly, the relative transcriptional activity of TAC fractions was significantly reduced in chloroplasts of et1-R plants. ET1 is the first zinc ribbon (ZR) protein shown to be targeted to plastids. With regard to its localization and its striking structural similarity to the eukaryotic transcription elongation factor TFIIS, it is feasible that ET1 functions in plastid transcription elongation by reactivation of arrested RNA polymerases.

  16. Synthesis and biological evaluation of diaryl-substituted carboranes as inhibitors of hypoxia inducible factor (HIF)-1 transcriptional activity.

    Science.gov (United States)

    Minegishi, Hidemitsu; Matsukawa, Takuya; Nakamura, Hiroyuki

    2013-02-01

    Diaromatic-substituted ortho- and meta-carboranes were synthesized as mimics of manassantin A. Among the carboranes synthesized, compounds 1 and 2 showed significant inhibition of hypoxia-induced HIF-1 transcriptional activity, with IC(50) values of 3.2 and 2.2 μM, respectively. Compounds 1 and 2 similarly suppressed hypoxia-induced HIF-1α accumulation in a concentration-dependent manner without affecting the expression level of HIF-1α mRNA. The hypoxia-induced accumulation and translocation of HIF-1α into nuclei were not observed in HeLa cells treated with compounds 1 and 2 by immunofluorescence analysis, revealing that the inhibition of hypoxia-induced HIF-1 transcriptional activity is induced by compounds 1 and 2 through a degradation pathway of the HIF-1α protein under hypoxic conditions.

  17. The BABY BOOM Transcription Factor Activates the LEC1-ABI3-FUS3-LEC2 Network to Induce Somatic Embryogenesis.

    Science.gov (United States)

    Horstman, Anneke; Li, Mengfan; Heidmann, Iris; Weemen, Mieke; Chen, Baojian; Muino, Jose M; Angenent, Gerco C; Boutilier, Kim

    2017-10-01

    Somatic embryogenesis is an example of induced cellular totipotency, where embryos develop from vegetative cells rather than from gamete fusion. Somatic embryogenesis can be induced in vitro by exposing explants to growth regulators and/or stress treatments. The BABY BOOM (BBM) and LEAFY COTYLEDON1 (LEC1) and LEC2 transcription factors are key regulators of plant cell totipotency, as ectopic overexpression of either transcription factor induces somatic embryo formation from Arabidopsis (Arabidopsis thaliana) seedlings without exogenous growth regulators or stress treatments. Although LEC and BBM proteins regulate the same developmental process, it is not known whether they function in the same molecular pathway. We show that BBM transcriptionally regulates LEC1 and LEC2, as well as the two other LAFL genes, FUSCA3 (FUS3) and ABSCISIC ACIDINSENSITIVE3 (ABI3). LEC2 and ABI3 quantitatively regulate BBM-mediated somatic embryogenesis, while FUS3 and LEC1 are essential for this process. BBM-mediated somatic embryogenesis is dose and context dependent, and the context-dependent phenotypes are associated with differential LAFL expression. We also uncover functional redundancy for somatic embryogenesis among other Arabidopsis BBM-like proteins and show that one of these proteins, PLETHORA2, also regulates LAFL gene expression. Our data place BBM upstream of other major regulators of plant embryo identity and totipotency. © 2017 American Society of Plant Biologists. All Rights Reserved.

  18. EFFECTS OF IL-4 UPON THE ACTIVITY OF STAT6 TRANSCRIPTION FACTOR IN PERIPHERAL BLOOD LYMPHOCYTES IN BRONCHIAL ASTHMA

    Directory of Open Access Journals (Sweden)

    V. N. Mineev

    2009-01-01

    Full Text Available Abstract. The aim of the study is to specify the levels of STAT6, and phospho-STAT6 under the influence of IL-4 in patients with bronchial asthma (BA. The samples from ten healthy controls and thirty-three BA patients with allergic and non-allergic clinical forms of different severity were under investigation. Peripheral blood lymphocytes were treated with 10 ng/ml of IL-4 (Sigma Aldrich, USA for 1 h. Then the proteins (STAT6 and phospho-STAT6 expressed in peripheral lymphocytes were analyzed by Western blot of cell lysates. Preparation of the cell lysates and Western blotting were carried out using standard procedures. Antibodies against phospho-STAT6 and STAT6 (10 ng/ml were used (Cell Signaling, USA. Levels of thespecific proteins were standardized according to β-actin (Cell Signaling, USA. Treatment with IL-4 caused an increase of phospho-STAT6 levels in lymphocytes of all BA patients, as compared with control group. In allergic BA, the phospho-STAT6 levels were significantly higher than in non-allergic clinical forms. Expression of STAT6 in lymphocytes of patients with severe BA was significantly higher, as compared to BA of moderate severity. An IL-4-induced activation of the STAT6 transcription factor was revealed in an in vitro system, being mostly expressed in allergic BA. The level of STAT6 may serve as a BA severity index. This study was supported by a «Scholarship of the Year» grant from the St. Petersburg State Medical I. Pavlov University (2007.

  19. 14-3-3-β and -{varepsilon} contribute to activation of the osmoprotective transcription factor NFAT5 by increasing its protein abundance and its transactivating activity.

    Science.gov (United States)

    Izumi, Yuichiro; Burg, Maurice B; Ferraris, Joan D

    2014-01-01

    Abstract Having previously found that high NaCl causes rapid exit of 14-3-3 isoforms from the nucleus, we used siRNA-mediated knockdown to test whether 14-3-3s contribute to the high NaCl-induced increase in the activity of the osmoprotective transcription factor NFAT5. We find that, when NaCl is elevated, knockdown of 14-3-3-β and/or 14-3-3-ε decreases NFAT5 transcriptional activity, as assayed both by luciferase reporter and by the mRNA abundance of the NFAT5 target genes aldose reductase and the sodium- and chloride-dependent betaine transporter, BGT1. Knockdown of other 14-3-3 isoforms does not significantly affect NFAT5 activity. 14-3-3-β and/or 14-3-3-ε do not act by affecting the nuclear localization of NFAT5, but by at least two other mechanisms: (1) 14-3-3-β and 14-3-3-ε increase protein abundance of NFAT5 and (2) they increase NFAT5 transactivating activity. When NaCl is elevated, knockdown of 14-3-3-β and/or 14-3-3-ε reduces the protein abundance of NFAT5, as measured by Western blot, without affecting the level of NFAT5 mRNA, and the knockdown also decreases NFAT5 transactivating activity, as measured by luciferase reporter. The 14-3-3s increase NFAT5 protein, not by increasing its translation, but by decreasing the rate at which it is degraded, as measured by cycloheximide chase. It is not clear at this point whether the 14-3-3s affect NFAT5 directly or indirectly through their effects on other proteins that signal activation of NFAT5.

  20. Modulation of p53 activity by IκBα: Evidence suggesting a common phylogeny between NF-κB and p53 transcription factors

    Directory of Open Access Journals (Sweden)

    Gelfand Erwin W

    2005-06-01

    Full Text Available Abstract Background In this work we present evidence that the p53 tumor suppressor protein and NF-κB transcription factors could be related through common descent from a family of ancestral transcription factors regulating cellular proliferation and apoptosis. P53 is a homotetrameric transcription factor known to interact with the ankyrin protein 53BP2 (a fragment of the ASPP2 protein. NF-κB is also regulated by ankyrin proteins, the prototype of which is the IκB family. The DNA binding sequences of the two transcription factors are similar, sharing 8 out of 10 nucleotides. Interactions between the two proteins, both direct and indirect, have been noted previously and the two proteins play central roles in the control of proliferation and apoptosis. Results Using previously published structure data, we noted a significant degree of structural alignment between p53 and NF-κB p65. We also determined that IκBα and p53 bind in vitro through a specific interaction in part involving the DNA binding region of p53, or a region proximal to it, and the amino terminus of IκBα independently or cooperatively with the ankyrin 3 domain of IκBα In cotransfection experiments, κBα could significantly inhibit the transcriptional activity of p53. Inhibition of p53-mediated transcription was increased by deletion of the ankyrin 2, 4, or 5 domains of IκBα Co-precipitation experiments using the stably transfected ankyrin 5 deletion mutant of κBα and endogenous wild-type p53 further support the hypothesis that p53 and IκBα can physically interact in vivo. Conclusion The aggregate results obtained using bacterially produced IκBα and p53 as well as reticulocyte lysate produced proteins suggest a correlation between in vitro co-precipitation in at least one of the systems and in vivo p53 inhibitory activity. These observations argue for a mechanism involving direct binding of IκBα to p53 in the inhibition of p53 transcriptional activity, analogous to

  1. Fungal-specific transcription factor AbPf2 activates pathogenicity in Alternaria brassicicola

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Yangrae; Ohm, Robin A. [US Department of Energy Joint Genome Institute, 2800 Mitchell Drive, Walnut Creek, CA, 94598, USA; Grigoriev, Igor V. [US Department of Energy Joint Genome Institute, 2800 Mitchell Drive, Walnut Creek, CA, 94598, USA; Srivastava, Akhil [Plant and Environmental Protection Sciences, University of Hawaii at Manoa, 3190 Maile Way, St John 317, Honolulu, HI, 96822, USA

    2013-05-24

    Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. To identify molecular determinants of pathogenicity, we created non-pathogenic mutants of a transcription factor-encoding gene, AbPf2. The frequency and timing of germination and appressorium formation on host plants were similar between the non-pathogenic abpf2 mutants and wild-type A. brassicicola. The mutants were also similar in vitro to wild-type A. brassicicola in terms of vegetative growth, conidium production, and responses to a phytoalexin, reactive oxygen species and osmolites. The hyphae of the mutants grew slowly but did not cause disease symptoms on the surface of host plants. Transcripts of the AbPf2 gene increased exponentially soon after wild-type conidia contacted their host plants . A small amount of AbPf2 protein, as monitored using GFP fusions, was present in young, mature conidia. The protein level decreased during saprophytic growth, but increased and was located primarily in fungal nuclei during pathogenesis. Levels of the proteins and transcripts sharply decreased following colonization of host tissues beyond the initial infection site. When expression of the transcription factor was induced in the wild-type during early pathogenesis, 106 fungal genes were also induced in the wild-type but not in the abpf2 mutants. Notably, 33 of the 106 genes encoded secreted proteins, including eight putative effector proteins. Plants inoculated with abpf2 mutants expressed higher levels of genes associated with photosynthesis, the pentose phosphate pathway and primary metabolism, but lower levels of defense-related genes. Our results suggest that AbPf2 is an important regulator of pathogenesis, but does not affect other cellular processes in A. brassicicola.

  2. Phosphorylation of activating transcription factor-2 (ATF-2) within the activation domain is a key determinant of sensitivity to tamoxifen in breast cancer.

    Science.gov (United States)

    Rudraraju, Bharath; Droog, Marjolein; Abdel-Fatah, Tarek M A; Zwart, Wilbert; Giannoudis, Athina; Malki, Mohammed I; Moore, David; Patel, Hetal; Shaw, Jacqui; Ellis, Ian O; Chan, Steve; Brooke, Greg N; Nevedomskaya, Ekaterina; Lo Nigro, Christiana; Carroll, Jason; Coombes, R Charles; Bevan, Charlotte; Ali, Simak; Palmieri, Carlo

    2014-09-01

    Activating transcription factor-2 (ATF-2) has been implicated as a tumour suppressor in breast cancer (BC). c-JUN N-terminal kinase (JNK) and p38 MAPK phosphorylate ATF-2 within the activation domain (AD), which is required for its transcriptional activity. To date, the role of ATF-2 in determining response to endocrine therapy has not been explored. Effects of ATF-2 loss in the oestrogen receptor (ER)-positive luminal BC cell line MCF7 were explored, as well as its role in response to tamoxifen treatment. Genome-wide chromatin binding patterns of ATF-2 when phosphorylated within the AD in MCF-7 cells were determined using ChIP-seq. The expression of ATF-2 and phosphorylated ATF-2 (pATF-2-Thr71) was determined in a series of 1,650 BC patients and correlated with clinico-pathological features and clinical outcome. Loss of ATF-2 diminished the growth-inhibitory effects of tamoxifen, while tamoxifen treatment induced ATF-2 phosphorylation within the AD, to regulate the expression of a set of 227 genes for proximal phospho-ATF-2 binding, involved in cell development, assembly and survival. Low expression of both ATF-2 and pATF-2-Thr71 was significantly associated with aggressive pathological features. Furthermore, pATF-2 was associated with both p-p38 and pJNK1/2 (ATF-2 is not associated with outcome, pATF-2 is associated with longer disease-free (p = 0.002) and BC-specific survival in patients exposed to tamoxifen (p = 0.01). Furthermore, multivariate analysis confirmed pATF-2-Thr71 as an independent prognostic factor. ATF-2 is important for modulating the effect of tamoxifen and phosphorylation of ATF-2 within the AD at Thr71 predicts for improved outcome for ER-positive BC receiving tamoxifen.

  3. 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

  4. The yeast transcription factor Crz1 is activated by light in a Ca2+/calcineurin-dependent and PKA-independent manner.

    Directory of Open Access Journals (Sweden)

    Kristofer Bodvard

    Full Text Available Light in the visible range can be stressful to non-photosynthetic organisms. The yeast Saccharomyces cerevisiae has earlier been reported to respond to blue light via activation of the stress-regulated transcription factor Msn2p. Environmental changes also induce activation of calcineurin, a Ca(2+/calmodulin dependent phosphatase, which in turn controls gene transcription by dephosphorylating the transcription factor Crz1p. We investigated the connection between cellular stress caused by blue light and Ca(2+ signalling in yeast by monitoring the nuclear localization dynamics of Crz1p, Msn2p and Msn4p. The three proteins exhibit distinctly different stress responses in relation to light exposure. Msn2p, and to a lesser degree Msn4p, oscillate rapidly between the nucleus and the cytoplasm in an apparently stochastic fashion. Crz1p, in contrast, displays a rapid and permanent nuclear localization induced by illumination, which triggers Crz1p-dependent transcription of its target gene CMK2. Moreover, increased extracellular Ca(2+ levels stimulates the light-induced responses of all three transcription factors, e.g. Crz1p localizes much quicker to the nucleus and a larger fraction of cells exhibits permanent Msn2p nuclear localization at higher Ca(2+ concentration. Studies in mutants lacking Ca(2+ transporters indicate that influx of extracellular Ca(2+ is crucial for the initial stages of light-induced Crz1p nuclear localization, while mobilization of intracellular Ca(2+ stores appears necessary for a sustained response. Importantly, we found that Crz1p nuclear localization is dependent on calcineurin and the carrier protein Nmd5p, while not being affected by increased protein kinase A activity (PKA, which strongly inhibits light-induced nuclear localization of Msn2/4p. We conclude that the two central signalling pathways, cAMP-PKA-Msn2/4 and Ca(2+-calcineurin-Crz1, are both activated by blue light illumination.

  5. Selective activation of tumor growth-promoting Ca2+ channel MS4A12 in colon cancer by caudal type homeobox transcription factor CDX2

    Directory of Open Access Journals (Sweden)

    Huber Christoph

    2009-09-01

    Full Text Available Abstract Colon cancer-associated MS4A12 is a novel colon-specific component of store-operated Ca2+ (SOC entry sensitizing cells for epidermal growth factor (EGF-mediated effects on proliferation and chemotaxis. In the present study, we investigated regulation of the MS4A12 promoter to understand the mechanisms responsible for strict transcriptional restriction of this gene to the colonic epithelial cell lineage. DNA-binding assays and luciferase reporter assays showed that MS4A12 promoter activity is governed by a single CDX homeobox transcription factor binding element. RNA interference (RNAi-mediated silencing of intestine-specific transcription factors CDX1 and CDX2 and chromatin immunoprecipitation (ChIP in LoVo and SW48 colon cancer cells revealed that MS4A12 transcript and protein expression is essentially dependent on the presence of endogenous CDX2. In summary, our findings provide a rationale for colon-specific expression of MS4A12. Moreover, this is the first report establishing CDX2 as transactivator of tumor growth-promoting gene expression in colon cancer, adding to untangle the complex and conflicting biological functions of CDX2 in colon cancer and supporting MS4A12 as important factor for normal colonic development as well as for the biology and treatment of colon cancer.

  6. TALE-induced bHLH transcription factors that activate a pectate lyase contribute to water soaking in bacterial spot of tomato

    Science.gov (United States)

    Schwartz, Allison R.; Morbitzer, Robert; Lahaye, Thomas; Staskawicz, Brian J.

    2017-01-01

    AvrHah1 [avirulence (avr) gene homologous to avrBs3 and hax2, no. 1] is a transcription activator-like (TAL) effector (TALE) in Xanthomonas gardneri that induces water-soaked disease lesions on fruits and leaves during bacterial spot of tomato. We observe that water from outside the leaf is drawn into the apoplast in X. gardneri-infected, but not X. gardneriΔavrHah1 (XgΔavrHah1)-infected, plants, conferring a dark, water-soaked appearance. The pull of water can facilitate entry of additional bacterial cells into the apoplast. Comparing the transcriptomes of tomato infected with X. gardneri vs. XgΔavrHah1 revealed the differential up-regulation of two basic helix–loop–helix (bHLH) transcription factors with predicted effector binding elements (EBEs) for AvrHah1. We mined our RNA-sequencing data for differentially up-regulated genes that could be direct targets of the bHLH transcription factors and therefore indirect targets of AvrHah1. We show that two pectin modification genes, a pectate lyase and pectinesterase, are targets of both bHLH transcription factors. Designer TALEs (dTALEs) for the bHLH transcription factors and the pectate lyase, but not for the pectinesterase, complement water soaking when delivered by XgΔavrHah1. By perturbing transcriptional networks and/or modifying the plant cell wall, AvrHah1 may promote water uptake to enhance tissue damage and eventual bacterial egression from the apoplast to the leaf surface. Understanding how disease symptoms develop may be a useful tool for improving the tolerance of crops from damaging disease lesions. PMID:28100489

  7. Urinary exosomal activating transcriptional factor 3 as the early diagnostic biomarker for sepsis-induced acute kidney injury.

    Science.gov (United States)

    Panich, Tanaporn; Chancharoenthana, Wiwat; Somparn, Poorichaya; Issara-Amphorn, Jiraphorn; Hirankarn, Nattiya; Leelahavanichkul, Asada

    2017-01-07

    An early sepsis-induced acute kidney injury (sepsis-AKI) biomarker is currently in needed. Urinary neutrophil gelatinase-associated lipocalin (uNGAL) is a candidate of sepsis-AKI biomarker but with different cut-point values. Urinary exosomal activating transcriptional factor 3 (uATF3) has been mentioned as an interesting biomarker. We conducted experiments in mice and a prospective, multicenter study in patients as a proof of concept that urine exosome is an interesting biomarker. An early expression of ATF3 in kidney of CD-1 mice at 6 h after cecal ligation and puncture implied the possibility of uATF3 as an early sepsis-AKI biomarker. Increase serum creatinine (Scr) ≥0.3 mg/dL from the baseline was used as an AKI diagnosis and urine was analyzed for uATF3 and uNGAL. Patients with baseline Scr at admission ≥1.5 mg/dL were excluded. The analysis showed higher Scr, uNGAL and uATF3 in patients with sepsis-AKI in comparison with patients with sepsis-non-AKI and healthy volunteers. A fair correlation, r(2) = 0.47, between uATF3 and uNGAL was showed in sepsis-AKI group with Scr ≥2 mg/dL. To see if uATF3 could be an early sepsis-AKI biomarker, urine sample was collected daily during the first week of the admission. In sepsis-AKI and sepsis-non-AKI groups, uNGAL were 367 ± 43 ng/mL and 183 ± 23 ng/mL, respectively; and uATF3 were 19 ± 4 ng/mL and 1.4 ± 0.8 ng/mL, respectively. With the mean value of uNGAL and uATF3 in sepsis AKI as a cut-off level, AUROC of uNGAL and uATF3 were 64% (95% CI 0.54 to 0.74) and 84% (95% CI 0.77 to 0.91), respectively. Urine exosome is an interesting source of urine biomarker and uATF3 is an interesting sepsis-AKI biomarker.

  8. Transcription-dependent degradation controls the stability of the SREBP family of transcription factors.

    Science.gov (United States)

    Sundqvist, Anders; Ericsson, Johan

    2003-11-25

    Cholesterol metabolism is tightly controlled by members of the sterol regulatory element-binding protein (SREBP) family of transcription factors. Here we demonstrate that the ubiquitination and degradation of SREBPs depend on their transcriptional activity. Mutations in the transactivation or DNA-binding domains of SREBPs inhibit their transcriptional activity and stabilize the proteins. The transcriptional activity and degradation of these mutants are restored when fused to heterologous transactivation or DNA-binding domains. When SREBP1a was fused to the DBD of Gal4, the ubiquitination and degradation of the fusion protein depended on coexpression of a promoter-reporter gene containing Gal4-binding sites. In addition, disruption of the interaction between WT SREBP and endogenous p300/CBP resulted in inhibition of SREBP-dependent transcription and stabilization of SREBP. Chemical inhibitors of transcription reduced the degradation of transcriptionally active SREBP1a, whereas they had no effect on the stability of transcriptionally inactive mutants, demonstrating that transcriptional activation plays an important role in the degradation of SREBPs. Thus, transcription-dependent degradation of SREBP constitutes a feedback mechanism to regulate the expression of genes involved in cholesterol metabolism and may represent a general mechanism to regulate the duration of transcriptional responses.

  9. Prunus transcription factors: Breeding perspectives

    Directory of Open Access Journals (Sweden)

    Valmor João Bianchi

    2015-06-01

    Full Text Available Many plant processes depend on differential gene expression, which is generally controlled by complex proteins called transcription factors (TFs. In peach, 1,533 TFs have been identified, accounting for about 5.5% of the 27,852 protein-coding genes. These TFs are the reference for the rest of the Prunus species. TF studies in Prunus have been performed on the gene expression analysis of different agronomic traits, including control of the flowering process, fruit quality, and biotic and abiotic stress resistance. These studies, using quantitative RT-PCR, have mainly been performed in peach, and to a lesser extent in other species, including almond, apricot, black cherry, Fuji cherry, Japanese apricot, plum, and sour and sweet cherry. Other tools have also been used in TF studies, including cDNA-AFLP, LC-ESI-MS, RNA and DNA blotting or mapping. More recently, new tools assayed include microarray and high-throughput DNA sequencing (DNA-Seq and RNA sequencing (RNA-Seq. New functional genomics opportunities include genome resequencing and the well-known synteny among Prunus genomes and transcriptomes. These new functional studies should be applied in breeding programs in the development of molecular markers. With the genome sequences available, some strategies that have been used in model systems (such as SNP genotyping assays and genotyping-by-sequencing may be applicable in the functional analysis of Prunus TFs as well. In addition, the knowledge of the gene functions and position in the peach reference genome of the TFs represents an additional advantage. These facts could greatly facilitate the isolation of genes via QTL (quantitative trait loci map-based cloning in the different Prunus species, following the association of these TFs with the identified QTLs using the peach reference genome.

  10. 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

  11. Mediator of DNA damage checkpoint 1 (MDC1 contributes to high NaCl-induced activation of the osmoprotective transcription factor TonEBP/OREBP.

    Directory of Open Access Journals (Sweden)

    Margarita Kunin

    Full Text Available BACKGROUND: Hypertonicity, such as induced by high NaCl, increases the activity of the transcription factor TonEBP/OREBP whose target genes increase osmoprotective organic osmolytes and heat shock proteins. METHODOLOGY: We used mass spectrometry to analyze proteins that coimmunoprecipitate with TonEBP/OREBP in order to identify ones that might contribute to its high NaCl-induced activation. PRINCIPAL FINDINGS: We identified 20 unique peptides from Mediator of DNA Damage Checkpoint 1 (MDC1 with high probability. The identification was confirmed by Western analysis. We used small interfering RNA knockdown of MDC1 to characterize its osmotic function. Knocking down MDC1 reduces high NaCl-induced increases in TonEBP/OREBP transcriptional and transactivating activity, but has no significant effect on its nuclear localization. We confirm six previously known phosphorylation sites in MDC1, but do not find evidence that high NaCl increases phosphorylation of MDC1. It is suggestive that MDC1 acts as a DNA damage response protein since hypertonicity reversibly increases DNA breaks, and other DNA damage response proteins, like ATM, also associate with TonEBP/OREBP and contribute to its activation by hypertonicity. CONCLUSIONS/SIGNIFICANCE: MDC1 associates with TonEBP/OREBP and contributes to high NaCl-induced increase of that factor's transcriptional activity.

  12. The activity of Mblk-1, a mushroom body-selective transcription factor from the honeybee, is modulated by the ras/MAPK pathway.

    Science.gov (United States)

    Park, Jung-Min; Kunieda, Takekazu; Kubo, Takeo

    2003-05-16

    We previously identified a gene, termed Mblk-1, that encodes a putative transcription factor with two DNA-binding motifs expressed preferentially in the mushroom body of the honeybee brain, and its preferred binding sequence, termed Mblk-1-binding element (MBE) (Takeuchi, H., Kage, E., Sawata, M., Kamikouchi, A., Ohashi, K., Ohara, M., Fujiyuki, T., Kunieda, T., Sekimizu, K., Natori, S., and Kubo, T. (2001) Insect Mol Biol 10, 487-494; Park, J.-M., Kunieda. T., Takeuchi, H., and Kubo, T. (2002) Biochem. Biophys. Res. Commun. 291, 23-28). In the present study, the effect of Mblk-1 on transcription of genes containing MBE in Drosophila Schneider's Line 2 cells was examined using a luciferase assay. Mblk-1 expression transactivated promoters containing MBEs approximately 2-7-fold. Deletion experiments revealed that RHF2, the second DNA-binding domain of Mblk-1, was necessary for the transcriptional activity. Furthermore, mitogen-activated protein kinase (MAPK) phosphorylated Mblk-1 at Ser-444 in vitro, and the Mblk-1-induced transactivation was stimulated by phosphorylation of Ser-444 by the Ras/MAPK pathway in the luciferase assay. These results suggest that Mblk-1 is a transcription factor that might function in the mushroom body neuronal circuits downstream of the Ras/MAPK pathway in the honeybee brain.

  13. 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.

  14. Transcription Factors Ets2 and Sp1 Act Synergistically with Histone Acetyltransferase p300 in Activating Human Interleukin-12 p40 Promoter

    Institute of Scientific and Technical Information of China (English)

    Hai-Jing SUN; Xin XU; Xiu-Li WANG; Liang WEI; Fen LI; Jun LU; Bai-Qu HUANG

    2006-01-01

    There has been considerable interest in researching the regulatory mechanisms that control the synthesis of interleukin (IL)-12, which plays a central role in the differentiation of T-helper-1 cells. In this study, we performed a series of transient transfection experiments designed to elucidate the functional relationship between the IL-12 promoter-specific transcription factors (Ets2 and Spl) and histone acetylation modification in IL-12 regulation mediated by p300 and various histone deacetylases (HDACs). Results presented in this report demonstrated that the transcription factors Ets2 and Spl acted synergistically with p300to activate the human IL-12 promoter. The histone acetyltransferase (HAT) activity of p300 was required for this synergic effect, because the adenovirus E1A protein inhibited the synergy. Conversely, HDACs repressed the synergic effect of transcription factors and histone acetylation on the activation of IL-12, while p300 was able to rectify it. These data indicated that Ets2 and Sp1 worked concertedly and synergistically with p300 in the regulation of human IL-12 expression.

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

    Directory of Open Access Journals (Sweden)

    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.

  16. YB-1 gene expression is kept constant during myocyte differentiation through replacement of different transcription factors and then falls gradually under the control of neural activity.

    Science.gov (United States)

    Kobayashi, Shunsuke; Tanaka, Toru; Moue, Masamitsu; Ohashi, Sachiyo; Nishikawa, Taishi

    2015-11-01

    We have previously reported that translation of acetylcholine receptor α-subunit (AChR α) mRNA in skeletal muscle cells is regulated by Y-box binding protein 1 (YB-1) in response to neural activity, and that in the postnatal mouse developmental changes in the amount of YB-1 mRNA are similar to those of AChR α mRNA, which is known to be regulated by myogenic transcription factors. Here, we examined transcriptional regulation of the YB-1 gene in mouse skeletal muscle and differentiating C2C12 myocytes. Although neither YB-1 nor AChR α was detected at either the mRNA or protein level in adult hind limb muscle, YB-1 expression was transiently activated in response to denervation of the sciatic nerve and completely paralleled that of AChR α, suggesting that these genes are regulated by the same transcription factors. However, during differentiation of C2C12 cells to myotubes, the level of YB-1 remained constant even though the level of AChR α increased markedly. Reporter gene, gel mobility shift and ChIP assays revealed that in the initial stage of myocyte differentiation, transcription of the YB-1 gene was regulated by E2F1 and Sp1, and was then gradually replaced under the control of both MyoD and myogenin through an E-box sequence in the proximal region of the YB-1 gene promoter. These results suggest that transcription factors for the YB-1 gene are exchanged during skeletal muscle cell differentiation, perhaps playing a role in translational control of mRNAs by YB-1 in both myotube formation and the response of skeletal muscle tissues to neural stimulation.

  17. First molluscan transcription factor activator protein-1 (Ap-1) member from disk abalone and its expression profiling against immune challenge and tissue injury.

    Science.gov (United States)

    De Zoysa, Mahanama; Nikapitiya, Chamilani; Lee, Youngdeuk; Lee, Sukkyoung; Oh, Chulhong; Whang, Ilson; Yeo, Sang-Yeop; Choi, Cheol Young; Lee, Jehee

    2010-12-01

    The regulation of transcriptional activation is an essential and critical point in gene expression. In this study, we describe a novel transcription factor activator protein-1 (Ap-1) gene from disk abalone Haliotis discus discus (AbAp-1) for the first time in mollusk. It was identified by homology screening of an abalone normalized cDNA library. The cloned AbAp-1 consists of a 945 bp coding region that encodes a putative protein containing 315 amino acids. The AbAp-1 gene is composed of a characteristic Jun transcription factor domain and a highly conserved basic leucine zipper (bZIP) signature similar to known Ap-1 genes. The AbAp-1 shares 46, 43 and, 40% amino acid identities with fish (Takifugu rubripes), human and insect (Ixodes scapularis) Ap-1, respectively. Quantitative real time RT-PCR analysis confirmed that AbAp-1 gene expression is constitutive in all selected tissues. AbAp-1 was upregulated in gills after bacteria (Vibrio alginolyticus, Vibrio parahemolyticus and Lysteria monocytogenes) challenge; and, upregulated in hemocytes and gills by viral hemorrhagic septicemia virus (VHSV) challenge. Shell damage and tissue injury also increased the transcriptional level of Ap-1 in mantle together with other transcription factors (NF-kB, LITAF) and pro-inflammatory TNF-α. All results considered, identification and gene expression data demonstrate that abalone Ap-1 is an important regulator in innate immune response against bacteria and virus, as well as in the inflammatory response during tissue injury. In addition, stimulation of Ap-1 under different external stimuli could be useful to understand the Ap-1 biology and its downstream target genes, especially in abalone-like mollusks.

  18. Tumor Suppressor Pten Inhibits Nuclear Accumulation of β-Catenin and T Cell/Lymphoid Enhancer Factor 1–Mediated Transcriptional Activation

    Science.gov (United States)

    Persad, Sujata; A.Troussard, Armelle; McPhee, Timothy R.; Mulholland, David J.; Dedhar, Shoukat

    2001-01-01

    β-Catenin is a protein that plays a role in intercellular adhesion as well as in the regulation of gene expression. The latter role of β-catenin is associated with its oncogenic properties due to the loss of expression or inactivation of the tumor suppressor adenomatous polyposis coli (APC) or mutations in β-catenin itself. We now demonstrate that another tumor suppressor, PTEN, is also involved in the regulation of nuclear β-catenin accumulation and T cell factor (TCF) transcriptional activation in an APC-independent manner. We show that nuclear β-catenin expression is constitutively elevated in PTEN null cells and this elevated expression is reduced upon reexpression of PTEN. TCF promoter/luciferase reporter assays and gel mobility shift analysis demonstrate that PTEN also suppresses TCF transcriptional activity. Furthermore, the constitutively elevated expression of cyclin D1, a β-catenin/TCF–regulated gene, is also suppressed upon reexpression of PTEN. Mechanistically, PTEN increases the phosphorylation of β-catenin and enhances its rate of degradation. We define a pathway that involves mainly integrin-linked kinase and glycogen synthase kinase 3 in the PTEN-dependent regulation of β-catenin stability, nuclear β-catenin expression, and transcriptional activity. Our data indicate that β-catenin/TCF–mediated gene transcription is regulated by PTEN, and this may represent a key mechanism by which PTEN suppresses tumor progression. PMID:11402061

  19. Resveratrol increases anti-aging Klotho gene expression via the activating transcription factor 3/c-Jun complex-mediated signaling pathway.

    Science.gov (United States)

    Hsu, Shih-Che; Huang, Shih-Ming; Chen, Ann; Sun, Chiao-Yin; Lin, Shih-Hua; Chen, Jin-Shuen; Liu, Shu-Ting; Hsu, Yu-Juei

    2014-08-01

    The Klotho gene functions as an aging suppressor gene. Evidence from animal models suggests that induction of Klotho expression may be a potential treatment for age-associated diseases. However, the molecular mechanism involved in regulating renal Klotho gene expression remains unclear. In this study, we determined that resveratrol, a natural polyphenol, induced renal Klotho expression both in vivo and in vitro. In the mouse kidney, resveratrol administration markedly increased both Klotho mRNA and protein expression. In resveratrol-treated NRK-52E cells, increased Klotho expression was accompanied by the upregulation and nuclear translocation of activating transcription factor 3 (ATF3) and c-Jun. ATF3 or c-Jun overexpression enhanced the transcriptional activation of Klotho. Conversely, resveratrol-induced Klotho expression was attenuated in the presence of dominant-negative ATF3 or c-Jun. Coimmunoprecipitation and a chromatin immunoprecipitation assay revealed that ATF3 physically interacted with c-Jun and that the ATF3/c-Jun complex directly bound to the Klotho promoter through ATF3- and AP-1-binding elements. c-Jun cotransfection augmented the effects of ATF3 on Klotho transcription in vitro. Although Sirtuin 1 mRNA expression was induced by resveratrol and involved in regulating Klotho mRNA expression, it was not the primary cause for the aforementioned ATF3/c-Jun pathway. In summary, resveratrol enhances the renal expression of the anti-aging Klotho gene, and the transcriptional factors ATF3 and c-Jun functionally interact and coordinately regulate the resveratrol-mediated transcriptional activation of Klotho.

  20. RBP-J kappa repression activity is mediated by a co-repressor and antagonized by the Epstein-Barr virus transcription factor EBNA2.

    Science.gov (United States)

    Waltzer, L; Bourillot, P Y; Sergeant, A; Manet, E

    1995-01-01

    The Epstein-Barr virus (EBV) protein EBNA2 is a transcriptional activator that can be targeted to its DNA responsive elements by direct interaction with the cellular protein RBP-J kappa. RBP-J kappa is a ubiquitous factor, highly conserved between man, mouse and Drosophila, whose function in mammalian cells is largely unknown. Here we provide evidence that RBP-J kappa is a transcriptional repressor and, more importantly, that RBP-J kappa repression is mediated by a co-repressor. The function of the co-repressor could be counterbalanced by making a fusion protein (RBP-VP16) between RBP-J kappa and the VP16 activation domain. This RBP-VP16-mediated activation could be strongly increased by an EBNA2 protein deprived of its activation domain, but not by an EBNA2 protein incapable of making physical contact with RBP-J kappa. Our results suggest that EBNA2 activates transcription by both interfering with the function of a co-repressor recruited by RBP-J kappa and providing an activation domain. Images PMID:8559649

  1. 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.

  2. HIF transcription factors, inflammation, and immunity.

    Science.gov (United States)

    Palazon, Asis; Goldrath, Ananda W; Nizet, Victor; Johnson, Randall S

    2014-10-16

    The hypoxic response in cells and tissues is mediated by the family of hypoxia-inducible factor (HIF) transcription factors; these play an integral role in the metabolic changes that drive cellular adaptation to low oxygen availability. HIF expression and stabilization in immune cells can be triggered by hypoxia, but also by other factors associated with pathological stress: e.g., inflammation, infectious microorganisms, and cancer. HIF induces a number of aspects of host immune function, from boosting phagocyte microbicidal capacity to driving T cell differentiation and cytotoxic activity. Cellular metabolism is emerging as a key regulator of immunity, and it constitutes another layer of fine-tuned immune control by HIF that can dictate myeloid cell and lymphocyte development, fate, and function. Here we discuss how oxygen sensing in the immune microenvironment shapes immunological response and examine how HIF and the hypoxia pathway control innate and adaptive immunity.

  3. A vascular endothelial growth factor activating transcription factor increases the endothelial progenitor cells population and induces therapeutic angiogenesis in a type 1 diabetic mouse with hindlimb ischemia

    Institute of Scientific and Technical Information of China (English)

    Diao Yongpeng; Lian Lishan; Guo Lilong; Chen Houzao; Chen Yuexin; Song Xiaojun; Li Yongjun

    2014-01-01

    Background Therapeutic angiogenesis has been shown to promote blood vessel growth and improve tissue perfusion.Vascular endothelial growth factor (VEGF) plays an important role in angiogenesis.However,it has side effects that limit its therapeutic utility in vivo,especially at high concentrations.This study aimed to investigate whether an intramuscular injection of a genetically engineered zinc finger VEGF-activating transcription factor modulates the endothelial progenitor cells (EPC) and promotes therapeutic angiogenesis in a hindlimb ischemia model with type 1 diabetes.Methods Alloxan (intravenous injection) was used to induce type Ⅰ diabetes in C57BL/6 mice (n=58).The ischemic limb received ZFP-VEGF (125 μg ZFP-VEGF plasmid in 1% poloxamer) or placebo (1% poloxamer) intramuscularly.Mice were sacrificed 3,5,10,or 20 days post-injection.Limb blood flow was monitored using laser Doppler perfusion imaging.VEGF mRNA and protein expression were examined using real-time PCR and ELISA,respectively.Capillary density,proliferation,and apoptosis were examined using immunohistochemistry techniques.Flow cytometry was used to detect the EPC population in bone marrow.Two-tailed Student's paired t test and repeated-measures analysis of variance were used for statistical analysis.Results ZFP-VEGF increased VEGF mRNA and protein expression at 3 and 10 days post-injection,and increased EPC in bone marrow at day 5 and 20 post-injection compared with controls (P<0.05).ZFP-VEGF treatment resulted in better perfusion recovery,a higher capillary density and proliferation,and less apoptosis compared with controls (P<0.05).Conclusions Intramuscular ZFP-VEGF injection promotes therapeutic angiogenesis in an ischemic hindlimb model with type 1 diabetes.This might be due to the effects of VEGF on cell survival and EPC recruitment.

  4. Transcriptional factors, Mafs and their biological roles

    Institute of Scientific and Technical Information of China (English)

    Mariko Tsuchiya; Ryoichi Misaka; Kosaku Nitta; Ken Tsuchiya

    2015-01-01

    The Maf family of transcription factors is characterizedby a typical bZip structure; these transcription factorsact as important regulators of the development anddifferentiation of many organs and tissues, includingthe kidney. The Maf family consists of two subgroupsthat are characterized according to their structure largeMaf transcription factors and small Maf transcriptionfactors. The large Maf subgroup consists of fourproteins, designated as MAFA, MAFB, c-MAF and neuralretina-specific leucine zipper. In particular, MAFA is adistinct molecule that has been attracting the attentionof researchers because it acts as a strong transactivatorof insulin, suggesting that Maf transcription factors arelikely to be involved in systemic energy homeostasis. Inthis review, we focused on the regulation of glucose/energy balance by Maf transcription factors in variousorgans.

  5. Transcription Factors Synergistically Activated at the Crossing of the Restriction Point between G1 and S Cell Cycle Phases. Pathologic Gate Opening during Multi-Hit Malignant Transformation

    Directory of Open Access Journals (Sweden)

    Nicoletta Castagnino

    2016-12-01

    Full Text Available Transcription factors (TFs represent key regulators of gene-expression patterns controlling cell behavior. TFs are active at nuclear – chromatin levels. TFs do not act in isolation; small sets of TFs cooperate toward the transcription of sets of mRNAs and consequently the translation of new proteins (the molecular phenotypes of a cell. Most TFs are activated through a cascade of biochemical reactions mediated by receptors expressed on the target cell surface. Nuclear Receptors (NRs are transcription factors activated instead by small hydrophobic molecules capable of crossing the plasma membrane. The convergence of different pathways on TFs and their posttranslational modifications ensure that the external stimuli generate appropriate and integrated responses. The reconstruction of the molecular anatomy of these pathways through Molecular Interactions Maps (MIMs can depict these intricate interactions. A mathematical modeling approach simulates/mimics their mechanism of action in normal and pathological conditions. We can simulate the effect of virtual hits in neoplastic transformation as mutations/alterations in these pathways. We can also simulate the effect of targeted inhibitors on these deregulated pathways. This strategy can help to guide an appropriate combination of targeted drugs in the treatment of a cancer patient, a major innovative perspective of incoming years.

  6. 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.

  7. A New Vaccinia Virus Intermediate Transcription Factor

    OpenAIRE

    Sanz, Patrick; Moss, Bernard

    1998-01-01

    Transcription of the vaccinia virus genome is mediated by a virus-encoded multisubunit DNA-dependent RNA polymerase in conjunction with early-, intermediate-, and late-stage-specific factors. Previous studies indicated that two virus-encoded proteins (capping enzyme and VITF-1) and one unidentified cellular protein (VITF-2) are required for specific transcription of an intermediate promoter template in vitro. We have now extensively purified an additional virus-induced intermediate transcript...

  8. 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.

  9. Analysis of the spacing between the two palindromes of activation sequence-1 with respect to binding to different TGA factors and transcriptional activation potential.

    OpenAIRE

    Krawczyk, Stefanie; Thurow, Corinna; Niggeweg, Ricarda; Gatz, Christiane

    2002-01-01

    In higher plants, activation sequence-1 (as-1) of the cauliflower mosaic virus 35S promoter mediates both salicylic acid- and auxin-inducible transcriptional activation. Originally found in viral and T-DNA promoters, as-1-like elements are also functional elements of plant promoters activated in the course of a defence response upon pathogen attack. as-1-like elements are characterised by two imperfect palindromes with the palindromic centres being spaced by 12 bp. They are recognised by plan...

  10. Yin Yang 1: a multifaceted protein beyond a transcription factor.

    Science.gov (United States)

    Deng, Zhiyong; Cao, Paul; Wan, Mei Mei; Sui, Guangchao

    2010-01-01

    As a transcription factor, Yin Yang 1 (YY1) regulates the transcription of a dazzling list of genes and the number of its targets still mounts. Recent studies revealed that YY1 possesses functions independent of its DNA binding activity and its regulatory role in tumorigenesis has started to emerge.

  11. Biochemical characterization of a molecular switch involving the heat shock protein ClpC, which controls the activity of ComK, the competence transcription factor of Bacillus subtilis

    NARCIS (Netherlands)

    Turgay, K; Hamoen, LW; Venema, G; Dubnau, D

    1997-01-01

    Development of genetic competence in Bacillus subtilis is controlled by the competence-specific transcription factor ComK. ComK activates transcription of itself and several other genes required for competence. The activity of ComK is controlled by other genes including mecA, clpC, and comS. We have

  12. Differential expression of brain-derived neurotrophic factor transcripts after pilocarpine-induced seizure-like activity is related to mode of Ca2+ entry

    DEFF Research Database (Denmark)

    Poulsen, F R; Lauterborn, J; Zimmer, J;

    2004-01-01

    ) and tyrosine kinase B (trkB) also were studied. Pilocarpine (5 mM) induced a dose- and time-dependent increase in total BDNF (exon V) mRNA expression in the dentate granule cells and CA3-CA1 pyramidal cells with maximal effects at 6 and 24 h, respectively. Increases were blocked by co-treatment with the alpha......Activity-dependent brain-derived neurotrophic factor (BDNF) expression is Ca2+-dependent, yet little is known about the Ca2+ channel contributions that might direct selective expression of the multiple BDNF transcripts. Here, effects of pilocarpine-induced seizure activity on total BDNF expression...... and on the individual sensitivity of BDNF transcripts to glutamate receptor and Ca2+ channel blockers were evaluated using hippocampal slice cultures and in situ hybridization of transcript-specific cRNA probes directed against mRNAs for the four 5' exons (I-IV) of the BDNF gene. mRNAs for nerve growth factor (NGF...

  13. Transcriptional elongation factor ENL phosphorylated by ATM recruits polycomb and switches off transcription for DSB repair.

    Science.gov (United States)

    Ui, Ayako; Nagaura, Yuko; Yasui, Akira

    2015-05-07

    Transcription is repressed if a DNA double-strand break (DSB) is introduced in close proximity to a transcriptional activation site at least in part by H2A-ubiquitination. While ATM signaling is involved, how it controls H2A-ubiquitination remains unclear. Here, we identify that, in response to DSBs, a transcriptional elongation factor, ENL (MLLT1), is phosphorylated by ATM at conserved SQ sites. This phosphorylation increases the interaction between ENL and the E3-ubiquitin-ligase complex of Polycomb Repressive Complex 1 (PRC1) via BMI1. This interaction promotes enrichment of PRC1 at transcription elongation sites near DSBs to ubiquitinate H2A leading to transcriptional repression. ENL SQ sites and BMI1 are necessary for KU70 accumulation at DSBs near active transcription sites and cellular resistance to DSBs. Our data suggest that ATM-dependent phosphorylation of ENL functions as switch from elongation to Polycomb-mediated repression to preserve genome integrity.

  14. In vivo identification of promoter elements and transcription factors mediating activation of hepatic HMG-CoA reductase by T{sub 3}

    Energy Technology Data Exchange (ETDEWEB)

    Boone, Lindsey R.; Niesen, Melissa I. [Department of Molecular Medicine, College of Medicine, University of South Florida, Tampa, FL (United States); Jaroszeski, Mark [Department of Chemical and Biomedical Engineering, College of Engineering, University of South Florida, Tampa, FL (United States); Ness, Gene C., E-mail: gness@hsc.usf.edu [Department of Molecular Medicine, College of Medicine, University of South Florida, Tampa, FL (United States)

    2009-07-31

    The promoter elements and transcription factors necessary for triiodothyronine (T{sub 3}) induction of hepatic HMG-CoA reductase (HMGR) were investigated by transfecting rat livers with wild type and mutant HMGR promoter-luciferase constructs using in vivo electroporation. Mutations in the sterol response element (SRE), nuclear factor-y (NF-Y) site, and the newly identified upstream transcription factor-2 (USF-2) site essentially abolished the T{sub 3} response. Chromatin immunoprecipitation (ChIP) analysis demonstrated that T{sub 3} treatment caused a 4-fold increase in in vivo binding of USF-2 to the HMGR promoter. Co-transfection of the wild type HMGR promoter with siRNAs to USF-2, SREBP-2, or NF-Y nearly abolished the T{sub 3} induction, as measured by promoter activity. These data provide in vivo evidence for functional roles for USF-2, SREBP-2, and NF-Y in mediating the T{sub 3}-induction of hepatic HMGR transcription.

  15. Ultraviolet irradiation induces CYR61/CCN1, a mediator of collagen homeostasis, through activation of transcription factor AP-1 in human skin fibroblasts.

    Science.gov (United States)

    Quan, Taihao; Qin, Zhaoping; Xu, Yiru; He, Tianyuan; Kang, Sewon; Voorhees, John J; Fisher, Gary J

    2010-06-01

    UV irradiation from the sun elevates the production of collagen-degrading matrix metalloproteinases (MMPs) and reduces the production of new collagen. This imbalance of collagen homeostasis impairs the structure and function of the dermal collagenous extracellular matrix (ECM), thereby promoting premature skin aging (photoaging). We report here that aberrant dermal collagen homeostasis in UV-irradiated human skin is mediated in part by a CCN-family member, cysteine-rich protein-61 (CYR61/CCN1). CYR61 is significantly elevated in acutely UV-irradiated human skin in vivo, and UV-irradiated human skin fibroblasts. Knockdown of CYR61 significantly attenuates UV irradiation-induced inhibition of type-I procollagen and upregulation of MMP-1. Determination of CYR61 mRNA and protein indicates that the primary mechanism of CYR61 induction by UV irradiation is transcriptional. Analysis of CYR61 proximal promoter showed that a sequence conforming to the consensus binding site for transcription factor activator protein-1 (AP-1) is required for promoter activity. UV irradiation increased the binding of AP-1-family members c-Jun and c-Fos to this AP-1 site. Furthermore, functional blockade of c-Jun or knockdown of c-Jun significantly reduced the UV irradiation-induced activation of CYR61 promoter and CYR61 gene expression. These data show that CYR61 is transcriptionally regulated by UV irradiation through transcription factor AP-1, and mediates altered collagen homeostasis that occurs in response to UV irradiation in human skin fibroblasts.

  16. Changes in the Expression of Transcription Factors Involved in Modulating the Expression of EPO-R in Activated Human CD4-Positive Lymphocytes.

    Science.gov (United States)

    Lisowska, Katarzyna A; Frackowiak, Joanna E; Mikosik, Anna; Witkowski, Jacek M

    2013-01-01

    We have recently described the presence of the erythropoietin receptor (EPO-R) on CD4(+) lymphocytes and demonstrated that its expression increases during their activation, reaching a level reported to be typical for erythroid progenitors. This observation suggests that EPO-R expression is modulated during lymphocyte activation, which may be important for the cells' function. Here we investigated whether the expression of GATA1, GATA3 and Sp1 transcription factors is correlated with the expression of EPO-R in human CD4(+) lymphocytes stimulated with monoclonal anti-CD3 antibody. The expression of GATA1, GATA3 and Sp1 transcription factors in CD4(+) cells was estimated before and after stimulation with anti-CD3 antibody by Western Blot and flow cytometry. The expression of EPO-R was measured using real-time PCR and flow cytometry. There was no change in the expression of GATA1 and GATA3 in CD4(+) lymphocytes after stimulation with anti-CD3 antibody. However, stimulation resulted in the significantly increased expression of the Sp1 factor. CD4(+) lymphocytes stimulated with anti-CD3 antibody exhibited an increase in both the expression level of EPOR gene and the number of EPO-R molecules on the cells' surface, the latter being significantly correlated with the increased expression of Sp1. Sp1 is noted to be the single transcription factor among the ones studied whose level changes as a result of CD4(+) lymphocyte stimulation. It seems that Sp1 may significantly affect the number of EPO-R molecules present on the surface of activated CD4(+) lymphocytes.

  17. Enhancing lysosomal biogenesis and autophagic flux by activating the transcription factor EB protects against cadmium-induced neurotoxicity

    Science.gov (United States)

    Pi, Huifeng; Li, Min; Tian, Li; Yang, Zhiqi; Yu, Zhengping; Zhou, Zhou

    2017-01-01

    Cadmium (Cd), a highly ubiquitous heavy metal, is a well-known inducer of neurotoxicity. However, the mechanism underlying cadmium-induced neurotoxicity remains unclear. In this study, we found that Cd inhibits autophagosome-lysosome fusion and impairs lysosomal function by reducing the levels of lysosomal-associated membrane proteins, inhibiting lysosomal proteolysis and altering lysosomal pH, contributing to defects in autophagic clearance and subsequently leading to nerve cell death. In addition, Cd decreases transcription factor EB (TFEB) expression at both the mRNA and protein levels. Furthermore, Cd induces the nuclear translocation of TFEB and TFEB target-gene expression, associated with compromised lysosomal function or a compensatory effect after the impairment of the autophagic flux. Notably, restoration of the levels of lysosomal-associated membrane protein, lysosomal proteolysis, lysosomal pH and autophagic flux through Tfeb overexpression protects against Cd-induced neurotoxicity, and this protective effect is incompletely dependent on TFEB nuclear translocation. Moreover, gene transfer of the master autophagy regulator TFEB results in the clearance of toxic proteins and the correction of Cd-induced neurotoxicity in vivo. Our study is the first to demonstrate that Cd disrupts lysosomal function and autophagic flux and manipulation of TFEB signalling may be a therapeutic approach for antagonizing Cd-induced neurotoxicity. PMID:28240313

  18. Transcription Factor Networks in Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    David Y. Rhee

    2014-09-01

    Full Text Available Specific cellular fates and functions depend on differential gene expression, which occurs primarily at the transcriptional level and is controlled by complex regulatory networks of transcription factors (TFs. TFs act through combinatorial interactions with other TFs, cofactors, and chromatin-remodeling proteins. Here, we define protein-protein interactions using a coaffinity purification/mass spectrometry method and study 459 Drosophila melanogaster transcription-related factors, representing approximately half of the established catalog of TFs. We probe this network in vivo, demonstrating functional interactions for many interacting proteins, and test the predictive value of our data set. Building on these analyses, we combine regulatory network inference models with physical interactions to define an integrated network that connects combinatorial TF protein interactions to the transcriptional regulatory network of the cell. We use this integrated network as a tool to connect the functional network of genetic modifiers related to mastermind, a transcriptional cofactor of the Notch pathway.

  19. Serine Arginine-Rich Splicing Factor 1 (SRSF1) Contributes to the Transcriptional Activation of CD3ζ in Human T Cells.

    Science.gov (United States)

    Moulton, Vaishali R; Gillooly, Andrew R; Perl, Marcel A; Markopoulou, Anastasia; Tsokos, George C

    2015-01-01

    T lymphocytes from many patients with systemic lupus erythematosus (SLE) express decreased levels of the T cell receptor (TCR)-associated CD3 zeta (ζ) signaling chain, a feature directly linked to their abnormal phenotype and function. Reduced mRNA expression partly due to defective alternative splicing, contributes to the reduced expression of CD3ζ chain. We previously identified by oligonucleotide pulldown and mass spectrometry approaches, the serine arginine-rich splicing factor 1 (SRSF1) binding to the 3' untranslated region (UTR) of CD3ζ mRNA. We showed that SRSF1 regulates alternative splicing of the 3'UTR of CD3ζ to promote expression of the normal full length 3`UTR over an unstable splice variant in human T cells. In this study we show that SRSF1 regulates transcriptional activation of CD3ζ. Specifically, overexpression and silencing of SRSF1 respectively increases and decreases CD3ζ total mRNA and protein expression in Jurkat and primary T cells. Using promoter-luciferase assays, we show that SRSF1 enhances transcriptional activity of the CD3ζ promoter in a dose dependent manner. Chromatin immunoprecipitation assays show that SRSF1 is recruited to the CD3ζ promoter. These results indicate that SRSF1 contributes to transcriptional activation of CD3ζ. Thus our study identifies a novel mechanism whereby SRSF1 regulates CD3ζ expression in human T cells and may contribute to the T cell defect in SLE.

  20. From tissue mechanics to transcription factors.

    Science.gov (United States)

    Janmey, Paul A; Wells, Rebecca G; Assoian, Richard K; McCulloch, Christopher A

    2013-10-01

    Changes in tissue stiffness are frequently associated with diseases such as cancer, fibrosis, and atherosclerosis. Several recent studies suggest that, in addition to resulting from pathology, mechanical changes may play a role akin to soluble factors in causing the progression of disease, and similar mechanical control might be essential for normal tissue development and homeostasis. Many cell types alter their structure and function in response to exogenous forces or as a function of the mechanical properties of the materials to which they adhere. This review summarizes recent progress in identifying intracellular signaling pathways, and especially transcriptional programs, that are differentially activated when cells adhere to materials with different mechanical properties or when they are subject to tension arising from external forces. Several cytoplasmic or cytoskeletal signaling pathways involving small GTPases, focal adhesion kinase and transforming growth factor beta as well as the transcriptional regulators MRTF-A, NFκB, and Yap/Taz have emerged as important mediators of mechanical signaling. © 2013 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  1. Male-dominant activation of rat renal organic anion transporter 1 (Oat1 and 3 (Oat3 expression by transcription factor BCL6.

    Directory of Open Access Journals (Sweden)

    Waja Wegner

    Full Text Available BACKGROUND: Organic anion transporters 1 (Oat1 and 3 (Oat3 mediate the transport of organic anions, including frequently prescribed drugs, across cell membranes in kidney proximal tubule cells. In rats, these transporters are known to be male-dominant and testosterone-dependently expressed. The molecular mechanisms that are involved in the sex-dependent expression are unknown. Our aim was to identify genes that show a sex-dependent expression and could be involved in male-dominant regulation of Oat1 and Oat3. METHODOLOGY/PRINCIPAL FINDINGS: Promoter activities of Oat1 and Oat3 were analyzed using luciferase assays. Expression profiling was done using a SurePrint G3 rat GE 8 × 60K microarray. RNA was isolated from renal cortical slices of four adult rats per sex. To filter the achieved microarray data for genes expressed in proximal tubule cells, transcription database alignment was carried out. We demonstrate that predicted androgen response elements in the promoters of Oat1 and Oat3 are not functional when the promoters were expressed in OK cells. Using microarray analyses we analyzed 17,406 different genes. Out of these genes, 56 exhibit a sex-dependent expression in rat proximal tubule cells. As genes potentially involved in the regulation of Oat1 and Oat3 expression, we identified, amongst others, the male-dominant hydroxysteroid (17-beta dehydrogenase 1 (Hsd17b1, B-cell CLL/lymphoma 6 (BCL6, and polymerase (RNA III (DNA directed polypeptide G (Polr3g. Moreover, our results revealed that the transcription factor BCL6 activates promoter constructs of Oat1 and Oat3. CONCLUSION: The results indicate that the male-dominant expression of both transporters, Oat1 and Oat3, is possibly not directly regulated by the classical androgen receptor mediated transcriptional pathway but appears to be regulated by the transcription factor BCL6.

  2. 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.

  3. Zinc finger artificial transcription factor-based nearest inactive analogue/nearest active analogue strategy used for the identification of plant genes controlling homologous recombination

    NARCIS (Netherlands)

    Jia, Qi; van Verk, Marcel C.; Pinas, Johan E.; Lindhout, Beatrice I.; Hooykaas, Paul J.J.; Van der Zaal, Bert J.

    2013-01-01

    In previous work, we selected a particular transcription factor, designated VP16-HRU, from a pool of zinc finger artificial transcription factors (ZF-ATFs) used for genome interrogation. When expressed in Arabidopsis thaliana under control of the ribosomal protein S5A promoter, the RPS5A::VP16-HRU c

  4. Research progress of activator of transcription factor Krox-20%转录激活因子Krox-20

    Institute of Scientific and Technical Information of China (English)

    马凯丽; 李聪; 刘增力; 王志钢

    2013-01-01

    早期生长应答基因2(early growth response 2,EGR2/Krox-20)是血清诱导的立早基因(immediate early)家族成员之一,其编码的蛋白质有3个锌指结构,可以与特异DNA序列结合,起转录激活作用.Krox-20的表达受多个信号通路的调控,进而对动物脂肪细胞的分化及神经系统的发育产生影响.对转基因鼠的研究表明,Krox-20可以促进动物脂肪形成及毛发生长等生理过程相关基因的表达,从而改良性状.本文概述了Krox-20的研究现状.%Early growth response 2 (EGR2/Krox-20) gene is one member of the immediate early gene family which is transiently activated after serum stimulation,and the Krox-20 protein,with three zinc fingers,can combine to specific DNA sequence and play the role of transcriptional activation.Its expression is regulated by multiple signaling pathways,and it is involved in animal fat cell differentiation as well as the development of nervous system.Studies on transgenic mice have indicated that Krox-20 specifically promotes the expression of certain genes which are related to the formation of animal fat,the development of hair follicle and other physiological processes.This review provides an overview of advance in study of Krox-20.

  5. Comparison of Transcription Factor Binding Site Models

    KAUST Repository

    Bhuyan, Sharifulislam

    2012-05-01

    Modeling of transcription factor binding sites (TFBSs) and TFBS prediction on genomic sequences are important steps to elucidate transcription regulatory mechanism. Dependency of transcription regulation on a great number of factors such as chemical specificity, molecular structure, genomic and epigenetic characteristics, long distance interaction, makes this a challenging problem. Different experimental procedures generate evidence that DNA-binding domains of transcription factors show considerable DNA sequence specificity. Probabilistic modeling of TFBSs has been moderately successful in identifying patterns from a family of sequences. In this study, we compare performances of different probabilistic models and try to estimate their efficacy over experimental TFBSs data. We build a pipeline to calculate sensitivity and specificity from aligned TFBS sequences for several probabilistic models, such as Markov chains, hidden Markov models, Bayesian networks. Our work, containing relevant statistics and evaluation for the models, can help researchers to choose the most appropriate model for the problem at hand.

  6. 3,3′-diindolylmethane induces activating transcription factor 3 (ATF3) via ATF4 in human colorectal cancer cells

    OpenAIRE

    Lee, Seong-Ho; Min, Kyung-Won; Zhang, Xiaobo; Baek, Seung Joon

    2012-01-01

    A 3,3′-diindolylmethane (DIM) is a major in vivo condensation product of indole-3-carbinol (I3C), which are present in cruciferous vegetables. Although these compounds have been widely implicated in anti-tumorigenic and pro-apoptotic properties in animal as well as in vitro models of cancer, the underlying cellular mechanisms regulated by DIM are only partially understood. Activating transcription factor 3 (ATF3) is a member of the ATF/CREB subfamily of the basic-region leucine zipper (bZIP) ...

  7. 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.

  8. [The Effect of Transcription on Enhancer Activity in Drosophila melanogaster].

    Science.gov (United States)

    Erokhin, M M; Davydova, A I; Lomaev, D V; Georgiev, P G; Chetverina, D A

    2016-01-01

    In higher eukaryotes, the level of gene transcription is under the control of DNA regulatory elements, such as promoter, from which transcription is initiated with the participation of RNA polymerase II and general transcription factors, as well as the enhancer, which increase the rate of transcription with the involvement of activator proteins and cofactors. It was demonstrated that enhancers are often located in the transcribed regions of the genome. We showed earlier that transcription negatively affected the activity of enhancers in Drosophila in model transgenic systems. In this study, we tested the effect of the distance between the leading promoter, enhancer, and target promoter on the inhibitory effect of transcriptions of different strengths. It was demonstrated that the negative effect of transcription remained, but weakened with increased distance between the leading promoter and enhancer and with decreased distance between the enhancer and target promoter. Thus, transcription can modulate the activity of enhancers by controlling its maximum level.

  9. Dehydroepiandrosterone sulfate mediates activation of transcription factors CREB and ATF-1 via a Gα11-coupled receptor in the spermatogenic cell line GC-2.

    Science.gov (United States)

    Shihan, Mazen; Kirch, Ulrike; Scheiner-Bobis, Georgios

    2013-12-01

    Dehydroepiandrosterone sulfate (DHEAS) is a circulating steroid produced in the adrenal cortex, brain, and gonads. Whereas a series of investigations attest to neuroprotective effects of the steroid in the brain, surprisingly little is known about the physiological effects of DHEAS on cells of the reproductive system. Here we demonstrate that DHEAS acting on the spermatogenic cell line GC-2 induces a time- and concentration-dependent phosphorylation of c-Src and Erk1/2 and activates the transcription factors activating transforming factor-1 (ATF-1) and cyclic AMP-responsive element binding protein (CREB). These actions are consistent with the non-classical signaling pathway of testosterone and suggest that DHEAS is a pro-androgen that is converted into testosterone in order to exert its biological activity. The fact, however, that steroid sulfatase mRNA was not detected in the GC-2 cells and the clear demonstration of DHEAS-induced activation of Erk1/2, ATF-1 and CREB after silencing the androgen receptor by small interfering RNA (siRNA) clearly contradict this assumption and make it appear unlikely that DHEAS has to be converted in the cytosol into a different steroid in order to activate the kinases and transcription factors mentioned. Instead, it is likely that the DHEAS-induced signaling is mediated through the interaction of the steroid with a membrane-bound G-protein-coupled receptor, since silencing of Guanine nucleotide-binding protein subunit alpha-11 (Gnα11) leads to the abolition of the DHEAS-induced stimulation of Erk1/2, ATF-1, and CREB. The investigation presented here shows a hormone-like activity of DHEAS on a spermatogenic cell line. Since DHEAS is produced in male and female reproductive organs, these findings could help to define new roles for DHEAS in the physiology of reproduction.

  10. PlnTFDB: an integrative plant transcription factor database

    Directory of Open Access Journals (Sweden)

    Ruzicic Slobodan

    2007-02-01

    Full Text Available Abstract Background Transcription factors (TFs are key regulatory proteins that enhance or repress the transcriptional rate of their target genes by binding to specific promoter regions (i.e. cis-acting elements upon activation or de-activation of upstream signaling cascades. TFs thus constitute master control elements of dynamic transcriptional networks. TFs have fundamental roles in almost all biological processes (development, growth and response to environmental factors and it is assumed that they play immensely important functions in the evolution of species. In plants, TFs have been employed to manipulate various types of metabolic, developmental and stress response pathways. Cross-species comparison and identification of regulatory modules and hence TFs is thought to become increasingly important for the rational design of new plant biomass. Up to now, however, no computational repository is available that provides access to the largely complete sets of transcription factors of sequenced plant genomes. Description PlnTFDB is an integrative plant transcription factor database that provides a web interface to access large (close to complete sets of transcription factors of several plant species, currently encompassing Arabidopsis thaliana (thale cress, Populus trichocarpa (poplar, Oryza sativa (rice, Chlamydomonas reinhardtii and Ostreococcus tauri. It also provides an access point to its daughter databases of a species-centered representation of transcription factors (OstreoTFDB, ChlamyTFDB, ArabTFDB, PoplarTFDB and RiceTFDB. Information including protein sequences, coding regions, genomic sequences, expressed sequence tags (ESTs, domain architecture and scientific literature is provided for each family. Conclusion We have created lists of putatively complete sets of transcription factors and other transcriptional regulators for five plant genomes. They are publicly available through http://plntfdb.bio.uni-potsdam.de. Further data will be

  11. Porins from Salmonella enterica Serovar Typhimurium Activate the Transcription Factors Activating Protein 1 and NF-κB through the Raf-1-Mitogen-Activated Protein Kinase Cascade

    Science.gov (United States)

    Galdiero, Massimiliano; Vitiello, Mariateresa; Sanzari, Emma; D’Isanto, Marina; Tortora, Annalisa; Longanella, Anna; Galdiero, Stefania

    2002-01-01

    In this study we examined the ability of Salmonella enterica serovar Typhimurium porins to activate activating protein 1 (AP-1) and nuclear factor κB (NF-κB) through the mitogen-activated protein kinase (MAPK) cascade, and we identified the AP-1-induced protein subunits. Our results demonstrate that these enzymes may participate in cell signaling pathways leading to AP-1 and NF-κB activation following porin stimulation of cells. Raf-1 was phosphorylated in response to the treatment of U937 cells with porins; moreover, the porin-mediated increase in Raf-1 phosphorylation is accompanied by the phosphorylation of MAPK kinase 1/2 (MEK1/2), p38, extracellular-signal-regulated kinase 1/2, and c-Jun N-terminal kinase. We used three different inhibitors of phosphorylation pathways: 2′-amino-3′-methoxyflavone (PD-098059), a selective inhibitor of MEK1 activator and the MAPK cascade; 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole (SB203580), a specific inhibitor of the p38 pathway; and 7β-acetoxy-1α,6β,9α-trihydroxy-8,13-epoxy-labd-14-en-11-one (forskolin), an inhibitor at the level of Raf-1 kinase. PD-098059 pretreatment of cells decreases AP-1 and NF-κB activation by lipopolysaccharide (LPS) but not by porins, and SB203580 pretreatment of cells decreases mainly AP-1 and NF-κB activation by porins; in contrast, forskolin pretreatment of cells does not affect AP-1 and NF-κB activation following either porin or LPS stimulation. Our data suggest that the p38 signaling pathway mainly regulates AP-1 and NF-κB activation in cells treated with S. enterica serovar Typhimurium porins. Antibody electrophoretic mobility shift assays showed that JunD and c-Fos binding is found in cells treated with porins, in cells treated with LPS, and in unstimulated cells. However, by 30 to 60 min of stimulation, a different complex including c-Jun appears in cells treated with porins or LPS, while the Fra-2 subunit is present only after porin stimulation

  12. 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

    Directory of Open Access Journals (Sweden)

    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.

  13. ETS transcription factors in embryonic vascular development.

    Science.gov (United States)

    Craig, Michael P; Sumanas, Saulius

    2016-07-01

    At least thirteen ETS-domain transcription factors are expressed during embryonic hematopoietic or vascular development and potentially function in the formation and maintenance of the embryonic vasculature or blood lineages. This review summarizes our current understanding of the specific roles played by ETS factors in vasculogenesis and angiogenesis and the implications of functional redundancies between them.

  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. Expression of activating transcription factor 3 (ATF 3) and caspase 3 in Schwann cells and axonal outgrowth after sciatic nerve repair in diabetic BB rats.

    Science.gov (United States)

    Stenberg, Lena; Kanje, Martin; Dolezal, Katarina; Dahlin, Lars B

    2012-04-25

    The aim of this study was to evaluate nerve regeneration in relation to the transcription factor, Activating Transcription Factor 3 (ATF 3), and an apoptotic marker, caspase 3, in the Schwann cells of diabetic BB rats (i.e. display type 1 diabetes phenotype). Sciatic nerves in healthy Wistar rats and in diabetic BB rats were transected and immediately repaired. Axonal outgrowth (neurofilament staining) and expression of ATF 3 and caspase 3 were quantified by immunohistochemistry after six days. There was no difference in axonal outgrowth between healthy and diabetic rats. However, the sciatic nerve in the diabetic rats exhibited a larger number of ATF 3 expressing Schwann cells at the site of the lesion and also a higher number of caspase 3 expressing Schwann cells. Similar differences were observed in the distal nerve segment between the healthy and diabetic rats. There were no correlations between the number of Schwann cells expressing ATF 3 and caspase 3. Thus, diabetic BB rats display an increased activation of ATF 3 and also a rise in apoptotic caspase 3 expressing Schwann cells, but with no discrepancy in length of axonal outgrowth after nerve injury and repair at six days. Knowledge about signal transduction mechanisms in diabetes after stress may provide new insights into the development of diabetic neuropathy and neuropathic pain. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  16. Differential effects of Sp cellular transcription factors on viral promoter activation by varicella-zoster virus (VZV) IE62 protein.

    Science.gov (United States)

    Khalil, Mohamed I; Ruyechan, William T; Hay, John; Arvin, Ann

    2015-11-01

    The immediate early (IE) 62 protein is the major varicella-zoster virus (VZV) regulatory factor. Analysis of the VZV genome revealed 40 predicted GC-rich boxes within 36 promoters. We examined effects of ectopic expression of Sp1-Sp4 on IE62- mediated transactivation of three viral promoters. Ectopic expression of Sp3 and Sp4 enhanced IE62 activation of ORF3 and gI promoters while Sp3 reduced IE62 activation of ORF28/29 promoter and VZV DNA replication. Sp2 reduced IE62 transactivation of gI while Sp1 had no significant influence on IE62 activation with any of these viral promoters. Electrophoretic mobility shift assays (EMSA) confirmed binding of Sp1 and Sp3 but not Sp2 and Sp4 to the gI promoter. Sp1-4 bound to IE62 and amino acids 238-258 of IE62 were important for the interaction with Sp3 and Sp4 as well as Sp1. This work shows that Sp family members have differential effects on IE62-mediated transactivation in a promoter-dependent manner.

  17. Angiotensin II-induced protein kinase D activates the ATF/CREB family of transcription factors and promotes StAR mRNA expression.

    Science.gov (United States)

    Olala, Lawrence O; Choudhary, Vivek; Johnson, Maribeth H; Bollag, Wendy B

    2014-07-01

    Aldosterone synthesis is initiated upon the transport of cholesterol from the outer to the inner mitochondrial membrane, where the cholesterol is hydrolyzed to pregnenolone. This process is the rate-limiting step in acute aldosterone production and is mediated by the steroidogenic acute regulatory (StAR) protein. We have previously shown that angiotensin II (AngII) activation of the serine/threonine protein kinase D (PKD) promotes acute aldosterone production in bovine adrenal glomerulosa cells, but the mechanism remains unclear. Thus, the purpose of this study was to determine the downstream signaling effectors of AngII-stimulated PKD activity. Our results demonstrate that overexpression of the constitutively active serine-to-glutamate PKD mutant enhances, whereas the dominant-negative serine-to-alanine PKD mutant inhibits, AngII-induced StAR mRNA expression relative to the vector control. PKD has been shown to phosphorylate members of the activating transcription factor (ATF)/cAMP response element binding protein (CREB) family of leucine zipper transcription factors, which have been shown previously to bind the StAR proximal promoter and induce StAR mRNA expression. In primary glomerulosa cells, AngII induces ATF-2 and CREB phosphorylation in a time-dependent manner. Furthermore, overexpression of the constitutively active PKD mutant enhances the AngII-elicited phosphorylation of ATF-2 and CREB, and the dominant-negative mutant inhibits this response. Furthermore, the constitutively active PKD mutant increases the binding of phosphorylated CREB to the StAR promoter. Thus, these data provide insight into the previously reported role of PKD in AngII-induced acute aldosterone production, providing a mechanism by which PKD may be mediating steroidogenesis in primary bovine adrenal glomerulosa cells.

  18. Advanced glycation end-products induce heparanase expression in endothelial cells by the receptor for advanced glycation end products and through activation of the FOXO4 transcription factor.

    Science.gov (United States)

    An, Xiao-Fei; Zhou, Lei; Jiang, Peng-Jun; Yan, Ming; Huang, Yu-Jun; Zhang, Su-Na; Niu, Yun-Fei; Ten, Shi-Chao; Yu, Jiang-Yi

    2011-08-01

    As an endo-β (1-4)-D: -glucuronidase, heparanase can specifically cleave carbohydrate chains of heparan sulfate (HS) and has been implicated in development of endothelial cells dsyfunction. The advanced glycation end products (AGEs) play a pivotal role in the pathology of diabetic complications. In the present study, we investigated the effect of AGE-bovine serum albumin (AGE-BSA) on heparanase expression in human microvascular endothelial cells (HMVECs) and the underlying molecular mechanisms. The results indicated that in vitro direct exposure of HMVECs to AGE-BSA (300, 1000, and 3000 μg/ml) could increase heparanase mRNA and protein expression in a dose and time-dependent manner. The effect of 1000 μg/ml AGE-BSA could be abolished by neutralization with antibody of the receptor for advanced glycation end products (RAGE). Moreover, pretreatment with inhibitors of nuclear factor-κB (NF-κB) or PI3-kinase did not affect heparanase expression induced by AGE-BSA. Nevertheless, small interference RNA (siRNA) for transcriptional factor FOXO4 could reduce the increase of heparanase expression in HMVECs induced by 1000 μg/ml AGE-BSA. These results suggest that AGEs could induce heparanase expression in HMVECs by RAGE and predominantly through activation of the FOXO4 transcription factor.

  19. 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.

  20. Schisandrin B exhibits anti-inflammatory activity through modulation of the redox-sensitive transcription factors Nrf2 and NF-κB.

    Science.gov (United States)

    Checker, Rahul; Patwardhan, Raghavendra S; Sharma, Deepak; Menon, Jisha; Thoh, Maikho; Bhilwade, Hari N; Konishi, Tetsuya; Sandur, Santosh K

    2012-10-01

    Schisandrin B (SB), a dibenzocyclooctadiene derivative isolated from Schisandra chinensis and used commonly in traditional Chinese medicine for the treatment of hepatitis and myocardial disorders, has been recently shown to modulate cellular redox balance. Since we have shown that cellular redox plays an important role in the modulation of immune responses, the present studies were undertaken to study the effects of SB on activation and effector functions of lymphocytes. SB altered the redox status of lymphocytes by enhancing the basal reactive oxygen species levels and altering the GSH/GSSG ratio in lymphocytes. It also induced nuclear translocation of redox sensitive transcription factor Nrf2 and increased the transcription of its dependent genes. SB inhibited mitogen-induced proliferation and cytokine secretion by lymphocytes. SB also significantly inhibited mitogen-induced upregulation of T cell costimulatory molecules and activation markers. It was observed that SB inhibited mitogen-induced phosphorylation of c-Raf, MEK, ERK, JNK, and p38. It suppressed IκBα degradation and nuclear translocation of NF-κB in activated lymphocytes. Anti-inflammatory effects of SB were significantly abrogated by the inhibitors of Nrf2 and HO-1, suggesting the involvement of this pathway. Similar anti-inflammatory effects of SB on lymphocyte proliferation and cytokine secretion were also observed in vivo. To our knowledge, this is the first report showing that the anti-inflammatory effects of SB are mediated via modulation of Nrf2 and NF-κB in lymphocytes.

  1. 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.

  2. SIRT1 Protein, by Blocking the Activities of Transcription Factors FoxO1 and FoxO3, Inhibits Muscle Atrophy and Promotes Muscle Growth*

    Science.gov (United States)

    Lee, Donghoon; Goldberg, Alfred L.

    2013-01-01

    In several cell types, the protein deacetylase SIRT1 regulates the activities of FoxO transcription factors whose activation is critical in muscle atrophy. However, the possible effects of SIRT1 on the activity of FoxOs in skeletal muscle and on the regulation of muscle size have not been investigated. Here, we show that after food deprivation, SIRT1 levels fall dramatically in type II skeletal muscles (tibialis anterior), which show marked atrophy, unlike in the liver (where SIRT1 rises) or heart or the soleus, a type I muscle (where SIRT1 is unchanged). Maintenance of high SIRT1 levels by electroporation in mouse muscle inhibits markedly the muscle wasting induced by fasting as well as by denervation, and these protective effects require its deacetylase activity. SIRT1 overexpression reduces muscle wasting by blocking the activation of FoxO1 and 3. It thus prevents the induction of key atrogenes, including the muscle-specific ubiquitin ligases, atrogin1 and MuRF1, and multiple autophagy (Atg) genes and the increase in overall proteolysis. In normal muscle, SIRT1 overexpression by electroporation causes rapid fiber hypertrophy without, surprisingly, activation of the PI3K-AKT signaling pathway. Thus, SIRT1 activation favors postnatal muscle growth, and its fall appears to be critical for atrophy during fasting. Consequently, SIRT1 activation represents an attractive possible pharmacological approach to prevent muscle wasting and cachexia. PMID:24003218

  3. 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.

  4. Keap1 silencing boosts lipopolysaccharide-induced transcription of interleukin 6 via activation of nuclear factor κB in macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Lv, Peng [Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Xue, Peng; Dong, Jian [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Peng, Hui [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Evaluation and Research Center for Toxicology, Institute of Disease Control and Prevention, Academy of Military Medical Sciences (China); Clewell, Rebecca [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Wang, Aiping [Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Wang, Yue [Institute for Medical Device Standardization Administration, National Institutes for Food and Drug Control, Beijing (China); Peng, Shuangqing [Evaluation and Research Center for Toxicology, Institute of Disease Control and Prevention, Academy of Military Medical Sciences (China); Qu, Weidong [Key Laboratory of the Public Health Safety, Ministry of Education, School of Public Health, Fudan University, Shanghai (China); Zhang, Qiang; Andersen, Melvin E. [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Pi, Jingbo, E-mail: jpi@thehamner.org [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States)

    2013-11-01

    Interleukin-6 (IL6) is a multifunctional cytokine that regulates immune and inflammatory responses. Multiple transcription factors, including nuclear factor κB (NF-κB) and nuclear factor E2-related factor 2 (Nrf2), regulate IL6 transcription. Kelch-like ECH-associated protein 1 (Keap1) is a substrate adaptor protein for the Cullin 3-dependent E3 ubiquitin ligase complex, which regulates the degradation of many proteins, including Nrf2 and IκB kinase β (IKKβ). Here, we found that stable knockdown of Keap1 (Keap1-KD) in RAW 264.7 (RAW) mouse macrophages and human monocyte THP-1 cells significantly increased expression of Il6, and Nrf2-target genes, under basal and lipopolysaccharide (LPS, 0.001–0.1 μg/ml)-challenged conditions. However, Nrf2 activation alone, by tert-butylhydroquinone treatment of RAW cells, did not increase expression of Il6. Compared to cells transduced with scrambled non-target negative control shRNA, Keap1-KD RAW cells showed enhanced protein levels of IKKβ and increased expression and phosphorylation of NF-κB p65 under non-stressed and LPS-treated conditions. Because the expression of Il6 in Keap1-KD RAW cells was significantly attenuated by silencing of Ikkβ, but not Nrf2, it appears that stabilized IKKβ is responsible for the enhanced transactivation of Il6 in Keap1-KD cells. This study demonstrated that silencing of Keap1 in macrophages boosts LPS-induced transcription of Il6 via NF-κB activation. Given the importance of IL6 in the inflammatory response, the Keap1–IKKβ–NF-κB pathway may be a novel target for treatment and prevention of inflammation and associated disorders. - Highlights: • Knockdown of Keap1 increases expression of Il6 in macrophages. • Silencing of Keap1 results in protein accumulation of IKKβ and NF-κB p65. • Induction of Il6 resulting from Keap1 silencing is attributed to NF-κB activation.

  5. CCAAT/enhancer-binding protein delta activates insulin-like growth factor-I gene transcription in osteoblasts. Identification of a novel cyclic AMP signaling pathway in bone

    Science.gov (United States)

    Umayahara, Y.; Ji, C.; Centrella, M.; Rotwein, P.; McCarthy, T. L.

    1997-01-01

    Insulin-like growth factor-I (IGF-I) plays a key role in skeletal growth by stimulating bone cell replication and differentiation. We previously showed that prostaglandin E2 (PGE2) and other cAMP-activating agents enhanced IGF-I gene transcription in cultured primary rat osteoblasts through promoter 1, the major IGF-I promoter, and identified a short segment of the promoter, termed HS3D, that was essential for hormonal regulation of IGF-I gene expression. We now demonstrate that CCAAT/enhancer-binding protein (C/EBP) delta is a major component of a PGE2-stimulated DNA-protein complex involving HS3D and find that C/EBPdelta transactivates IGF-I promoter 1 through this site. Competition gel shift studies first indicated that a core C/EBP half-site (GCAAT) was required for binding of a labeled HS3D oligomer to osteoblast nuclear proteins. Southwestern blotting and UV-cross-linking studies showed that the HS3D probe recognized a approximately 35-kDa nuclear protein, and antibody supershift assays indicated that C/EBPdelta comprised most of the PGE2-activated gel-shifted complex. C/EBPdelta was detected by Western immunoblotting in osteoblast nuclear extracts after treatment of cells with PGE2. An HS3D oligonucleotide competed effectively with a high affinity C/EBP site from the rat albumin gene for binding to osteoblast nuclear proteins. Co-transfection of osteoblast cell cultures with a C/EBPdelta expression plasmid enhanced basal and PGE2-activated IGF-I promoter 1-luciferase activity but did not stimulate a reporter gene lacking an HS3D site. By contrast, an expression plasmid for the related protein, C/EBPbeta, did not alter basal IGF-I gene activity but did increase the response to PGE2. In osteoblasts and in COS-7 cells, C/EBPdelta, but not C/EBPbeta, transactivated a reporter gene containing four tandem copies of HS3D fused to a minimal promoter; neither transcription factor stimulated a gene with four copies of an HS3D mutant that was unable to bind osteoblast

  6. CCAAT/enhancer-binding protein delta activates insulin-like growth factor-I gene transcription in osteoblasts. Identification of a novel cyclic AMP signaling pathway in bone

    Science.gov (United States)

    Umayahara, Y.; Ji, C.; Centrella, M.; Rotwein, P.; McCarthy, T. L.

    1997-01-01

    Insulin-like growth factor-I (IGF-I) plays a key role in skeletal growth by stimulating bone cell replication and differentiation. We previously showed that prostaglandin E2 (PGE2) and other cAMP-activating agents enhanced IGF-I gene transcription in cultured primary rat osteoblasts through promoter 1, the major IGF-I promoter, and identified a short segment of the promoter, termed HS3D, that was essential for hormonal regulation of IGF-I gene expression. We now demonstrate that CCAAT/enhancer-binding protein (C/EBP) delta is a major component of a PGE2-stimulated DNA-protein complex involving HS3D and find that C/EBPdelta transactivates IGF-I promoter 1 through this site. Competition gel shift studies first indicated that a core C/EBP half-site (GCAAT) was required for binding of a labeled HS3D oligomer to osteoblast nuclear proteins. Southwestern blotting and UV-cross-linking studies showed that the HS3D probe recognized a approximately 35-kDa nuclear protein, and antibody supershift assays indicated that C/EBPdelta comprised most of the PGE2-activated gel-shifted complex. C/EBPdelta was detected by Western immunoblotting in osteoblast nuclear extracts after treatment of cells with PGE2. An HS3D oligonucleotide competed effectively with a high affinity C/EBP site from the rat albumin gene for binding to osteoblast nuclear proteins. Co-transfection of osteoblast cell cultures with a C/EBPdelta expression plasmid enhanced basal and PGE2-activated IGF-I promoter 1-luciferase activity but did not stimulate a reporter gene lacking an HS3D site. By contrast, an expression plasmid for the related protein, C/EBPbeta, did not alter basal IGF-I gene activity but did increase the response to PGE2. In osteoblasts and in COS-7 cells, C/EBPdelta, but not C/EBPbeta, transactivated a reporter gene containing four tandem copies of HS3D fused to a minimal promoter; neither transcription factor stimulated a gene with four copies of an HS3D mutant that was unable to bind osteoblast

  7. 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.

  8. Eomesodermin controls interleukin-5 production in memory T helper 2 cells through inhibition of activity of the transcription factor GATA3.

    Science.gov (United States)

    Endo, Yusuke; Iwamura, Chiaki; Kuwahara, Makoto; Suzuki, Akane; Sugaya, Kaoru; Tumes, Damon J; Tokoyoda, Koji; Hosokawa, Hiroyuki; Yamashita, Masakatsu; Nakayama, Toshinori

    2011-11-23

    The regulation of memory CD4(+) helper T (Th) cell function, such as polarized cytokine production, remains unclear. Here we show that memory T helper 2 (Th2) cells are divided into four subpopulations by CD62L and CXCR3 expression. All four subpopulations produced interleukin-4 (IL-4) and IL-13, whereas only the CD62L(lo)CXCR3(lo) population produced IL-5 accompanied by increased H3-K4 methylation at the Il5 gene locus. The transcription factor Eomesodermin (encoded by Eomes) was highly expressed in memory Th2 cells, whereas its expression was selectively downregulated in the IL-5-producing cells. Il5 expression was enhanced in Eomes-deficient cells, and Eomesodermin was shown to interact with the transcription factor GATA3, preventing GATA3 binding to the Il5 promoter. Memory Th2 cell-dependent airway inflammation was attenuated in the absence of the CD62L(lo)CXCR3(lo) population but was enhanced by Eomes-deficient memory Th2 cells. Thus, IL-5 production in memory Th2 cells is regulated by Eomesodermin via the inhibition of GATA3 activity.

  9. Modulation of DNA binding by gene-specific transcription factors.

    Science.gov (United States)

    Schleif, Robert F

    2013-10-01

    The transcription of many genes, particularly in prokaryotes, is controlled by transcription factors whose activity can be modulated by controlling their DNA binding affinity. Understanding the molecular mechanisms by which DNA binding affinity is regulated is important, but because forming definitive conclusions usually requires detailed structural information in combination with data from extensive biophysical, biochemical, and sometimes genetic experiments, little is truly understood about this topic. This review describes the biological requirements placed upon DNA binding transcription factors and their consequent properties, particularly the ways that DNA binding affinity can be modulated and methods for its study. What is known and not known about the mechanisms modulating the DNA binding affinity of a number of prokaryotic transcription factors, including CAP and lac repressor, is provided.

  10. NAC Transcription Factors in Stress Responses and Senescence

    DEFF Research Database (Denmark)

    O'Shea, Charlotte

    Plant-specific NAM/ATAF/CUC (NAC) transcription factors have recently received considerable attention due to their significant roles in plant development and stress signalling. This interest has resulted in a number of physiological, genetic and cell biological studies of their functions. Some......RF is a functional hotspot for both transcriptional activity and interaction with the cellular hub protein Radical Induced Cell Death1 (RCD1). Specific amino acid residues essential for the interaction were identified. These studies and structural analysis suggested that RCD1-ANAC046 complex formation does......-associated RCD1-transcription factor network represents a much-needed model for translation of in vitro obtained ID analysis, as presented here, to the in vivo organismal level. A systematic analysis was performed of the DNA-binding-site landscape and regulatory network of NAC transcription factors...

  11. Activity of transcription factor JACKDAW is essential for SHR/SCR-dependent activation of SCARECROW and MAGPIE and is modulated by reciprocal interactions with MAGPIE, SCARECROW and SHORT ROOT.

    Science.gov (United States)

    Ogasawara, Hiromi; Kaimi, Ryuji; Colasanti, Joseph; Kozaki, Akiko

    2011-11-01

    Two GRAS family transcription factors, SHORT-ROOT (SHR) and SCARECROW (SCR), are required for ground tissue and quiescent center formation in Arabidopsis roots. The action of SHR and SCR is regulated by two INDETERMINATE DOMAIN (IDD) family proteins, JACKDAW (JKD) and MAGPIE (MGP). Although the reciprocal interaction of these transcription factors is considered to be involved in the modulation of SHR and SCR action by JKD and MGP, the underlying mechanism remains unclear. In this study, we use a transient assay with Arabidopsis culture cells to show that the physical interaction of these transcription factors modulate their transcriptional activity. Transient expression of LUC reporter genes with the proximal sequences upstream from the ATG codon of SCR and MGP in protoplasts were activated by JKD. Moreover, promoter activities were enhanced further by the addition of SHR and SCR to JKD, but not by the combination of SHR and SCR in the absence of JKD. Yeast one-hybrid analysis showed that JKD binds to the SCR and MGP promoter sequences, indicating the existence of another binding sequences of JKD different from the previously determined IDD binding sequence. Our findings suggest that JKD directly regulates SCR and MGP expression in cooperation with SHR, SCR and MGP.

  12. Runx transcription factors in neuronal development

    Directory of Open Access Journals (Sweden)

    Shiga Takashi

    2008-08-01

    Full Text Available Abstract Runt-related (Runx transcription factors control diverse aspects of embryonic development and are responsible for the pathogenesis of many human diseases. In recent years, the functions of this transcription factor family in the nervous system have just begun to be understood. In dorsal root ganglion neurons, Runx1 and Runx3 play pivotal roles in the development of nociceptive and proprioceptive sensory neurons, respectively. Runx appears to control the transcriptional regulation of neurotrophin receptors, numerous ion channels and neuropeptides. As a consequence, Runx contributes to diverse aspects of the sensory system in higher vertebrates. In this review, we summarize recent progress in determining the role of Runx in neuronal development.

  13. Transcriptional activation of hypoxia-inducible factor-1 (HIF-1) in myeloid cells promotes angiogenesis through VEGF and S100A8.

    Science.gov (United States)

    Ahn, G-One; Seita, Jun; Hong, Beom-Ju; Kim, Young-Eun; Bok, Seoyeon; Lee, Chan-Ju; Kim, Kwang Soon; Lee, Jerry C; Leeper, Nicholas J; Cooke, John P; Kim, Hak Jae; Kim, Il Han; Weissman, Irving L; Brown, J Martin

    2014-02-18

    Emerging evidence indicates that myeloid cells are essential for promoting new blood vessel formation by secreting various angiogenic factors. Given that hypoxia-inducible factor (HIF) is a critical regulator for angiogenesis, we questioned whether HIF in myeloid cells also plays a role in promoting angiogenesis. To address this question, we generated a unique strain of myeloid-specific knockout mice targeting HIF pathways using human S100A8 as a myeloid-specific promoter. We observed that mutant mice where HIF-1 is transcriptionally activated in myeloid cells (by deletion of the von Hippel-Lindau gene) resulted in erythema, enhanced neovascularization in matrigel plugs, and increased production of vascular endothelial growth factor (VEGF) in the bone marrow, all of which were completely abrogated by either genetic or pharmacological inactivation of HIF-1. We further found that monocytes were the major effector producing VEGF and S100A8 proteins driving neovascularization in matrigel. Moreover, by using a mouse model of hindlimb ischemia we observed significantly improved blood flow in mice intramuscularly injected with HIF-1-activated monocytes. This study therefore demonstrates that HIF-1 activation in myeloid cells promotes angiogenesis through VEGF and S100A8 and that this may become an attractive therapeutic strategy to treat diseases with vascular defects.

  14. ZCT1 and ZCT2 transcription factors repress the activity of a gene promoter from the methyl erythritol phosphate pathway in Madagascar periwinkle cells.

    Science.gov (United States)

    Chebbi, Mouadh; Ginis, Olivia; Courdavault, Vincent; Glévarec, Gaëlle; Lanoue, Arnaud; Clastre, Marc; Papon, Nicolas; Gaillard, Cécile; Atanassova, Rossitza; St-Pierre, Benoit; Giglioli-Guivarc'h, Nathalie; Courtois, Martine; Oudin, Audrey

    2014-10-15

    In Catharanthus roseus, accumulating data highlighted the existence of a coordinated transcriptional regulation of structural genes that takes place within the secoiridoid biosynthetic branch, including the methyl erythritol phosphate (MEP) pathway and the following steps leading to secologanin. To identify transcription factors acting in these pathways, we performed a yeast one-hybrid screening using as bait a promoter region of the hydroxymethylbutenyl 4-diphosphate synthase (HDS) gene involved in the responsiveness of C. roseus cells to hormonal signals inducing monoterpene indole alkaloid (MIA) production. We identified that ZCT2, one of the three members of the zinc finger Catharanthus protein (ZCT) family, can bind to a HDS promoter region involved in hormonal responsiveness. By trans-activation assays, we demonstrated that ZCT1 and ZCT2 but not ZCT3 repress the HDS promoter activity. Gene expression analyses in C. roseus cells exposed to methyljasmonate revealed a persistence of induction of ZCT2 gene expression suggesting the existence of feed-back regulatory events acting on HDS gene expression in correlation with the MIA production.

  15. Activating transcription factor 4 mediates a multidrug resistance phenotype of esophageal squamous cell carcinoma cells through transactivation of STAT3 expression.

    Science.gov (United States)

    Zhu, Hongwu; Chen, Xiong; Chen, Bin; Chen, Bei; Fan, Jianyong; Song, Weibing; Xie, Ziying; Jiang, Dan; Li, Qiuqiong; Zhou, Meihua; Sun, Dayong; Zhao, Yagang

    2014-11-01

    Multidrug resistance (MDR) is a major challenge to the clinical treatment of esophageal cancer. The stress response gene activating transcription factor 4 (ATF4) is involved in homeostasis and cellular protection. However, relatively little is known about the expression and function of ATF4 in esophageal squamous cell carcinoma (ESCC) MDR. In this study, we investigate the potential role and mechanisms of ATF4 in ESCC MDR. We demonstrated that overexpression of ATF4 promotes the MDR phenotype in ESCC cells, while depletion of ATF4 in the MDR ESCC cell line induces drug re-sensitization. We also demonstrated that ATF4 transactivates STAT3 expression by directly binding to the signal transducers and activators of transcription 3 (STAT3) promoter, resulting in MDR in ESCC cells. Significantly, inhibition of STAT3 by small interfering RNA (siRNA) or a selective inhibitor (JSI-124) reintroduces therapeutic sensitivity. In addition, increased Bcl-2, survivin, and MRP1 expression levels were observed in ATF4-overexpressing cells. In conclusion, ATF4 may promote MDR in ESCC cells through the up-regulation of STAT3 expression, and thus is an attractive therapeutic target to combat therapeutic resistance in ESCC.

  16. NAC transcription factors: structurally distinct, functionally diverse

    DEFF Research Database (Denmark)

    Olsen, Addie Nina; Ernst, Heidi A; Leggio, Leila Lo;

    2005-01-01

    NAC proteins constitute one of the largest families of plant-specific transcription factors, and the family is present in a wide range of land plants. Here, we summarize the biological and molecular functions of the NAC family, paying particular attention to the intricate regulation of NAC protei...

  17. TCP transcription factors: architectures of plant form.

    Science.gov (United States)

    Manassero, Nora G Uberti; Viola, Ivana L; Welchen, Elina; Gonzalez, Daniel H

    2013-04-01

    After its initial definition in 1999, the TCP family of transcription factors has become the focus of a multiplicity of studies related with plant development at the cellular, organ, and tissue levels. Evidence has accumulated indicating that TCP transcription factors are the main regulators of plant form and architecture and constitute a tool through which evolution shapes plant diversity. The TCP transcription factors act in a multiplicity of pathways related with cell proliferation and hormone responses. In recent years, the molecular pathways of TCP protein action and biochemical studies on their mode of interaction with DNA have begun to shed light on their mechanism of action. However, the available information is fragmented and a unifying view of TCP protein action is lacking, as well as detailed structural studies of the TCP-DNA complex. Also important, the possible role of TCP proteins as integrators of plant developmental responses to the environment has deserved little attention. In this review, we summarize the current knowledge about the structure and functions of TCP transcription factors and analyze future perspectives for the study of the role of these proteins and their use to modify plant development.

  18. The activating protein 1 transcription factor basic leucine zipper transcription factor, ATF-like (BATF), regulates lymphocyte- and mast cell-driven immune responses in the setting of allergic asthma.

    Science.gov (United States)

    Übel, Caroline; Sopel, Nina; Graser, Anna; Hildner, Kai; Reinhardt, Cornelia; Zimmermann, Theodor; Rieker, Ralf Joachim; Maier, Anja; Neurath, Markus F; Murphy, Kenneth M; Finotto, Susetta

    2014-01-01

    Mice without the basic leucine zipper transcription factor, ATF-like (BATF) gene (Batf(-/-)) lack TH17 and follicular helper T cells, which demonstrates that Batf is a transcription factor important for T- and B-cell differentiation. In this study we examined whether BATF expression would influence allergic asthma. In a cohort of preschool control children and children with asthma, we analyzed BATF mRNA expression using real-time PCR in PBMCs. In a murine model of allergic asthma, we analyzed differences in this allergic disease between wild-type, Batf transgenic, and Batf(-/-) mice. In the absence of corticosteroid treatment, children with recurrent asthma have a significant increase in BATF mRNA expression in their PBMCs. Batf(-/-) mice display a significant reduction in the pathophysiologic responses seen in asthmatic wild-type littermates. Moreover, we discovered a decrease in IL-3 production and IL-3-dependent mast cell development in Batf(-/-) mice. By contrast, IFN-γ was induced in lung CD4(+) and CD8(+) T cells. Intranasal delivery of anti-IFN-γ antibodies induced airway hyperresponsiveness and inflammation in wild-type but not in Batf(-/-) mice. Transgenic overexpression of Batf under the control of the CD2 promoter/enhancer augmented lung inflammation and IgE levels in the setting of experimental asthma. BATF is increased in non-steroid-treated asthmatic children. Targeting BATF expression resulted in amelioration of the pathophysiologic responses seen in children with allergic asthma, and BATF has emerged as a novel target for antiasthma interventions. Copyright © 2013 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

  19. Regulation of the Hippo Pathway Transcription Factor TEAD.

    Science.gov (United States)

    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.

  20. Role of Flightless-I (Drosophila) homolog in the transcription activation of type I collagen gene mediated by transforming growth factor beta

    Energy Technology Data Exchange (ETDEWEB)

    Lim, Mi-Sun; Jeong, Kwang Won, E-mail: kwjeong@gachon.ac.kr

    2014-11-21

    Highlights: • FLII activates TGFβ-mediated expression of COL1A2 gene. • TGFβ induces the association of FLII with SMAD3 and BRG1 in A549 cells. • FLII is required for the recruitment of SWI/SNF complex and chromatin accessibility to COL1A2 promoter. - Abstract: Flightless-I (Drosophila) homolog (FLII) is a nuclear receptor coactivator that is known to interact with other transcriptional regulators such as the SWI/SNF complex, an ATP-dependent chromatin-remodeling complex, at the promoter or enhancer region of estrogen receptor (ER)-α target genes. However, little is known about the role of FLII during transcription initiation in the transforming growth factor beta (TGFβ)/SMAD-dependent signaling pathway. Here, we demonstrate that FLII functions as a coactivator in the expression of type I collagen gene induced by TGFβ in A549 cells. FLII activates the reporter gene driven by COL1A2 promoter in a dose-dependent manner. Co-expression of GRIP1, CARM1, or p300 did not show any synergistic activation of transcription. Furthermore, the level of COL1A2 expression correlated with the endogenous level of FLII mRNA level. Depletion of FLII resulted in a reduction of TGFβ-induced expression of COL1A2 gene. In contrast, over-expression of FLII caused an increase in the endogenous expression of COL1A2. We also showed that FLII is associated with Brahma-related gene 1 (BRG1) as well as SMAD in A549 cells. Notably, the recruitment of BRG1 to the COL1A2 promoter region was decreased in FLII-depleted A549 cells, suggesting that FLII is required for TGFβ-induced chromatin remodeling, which is carried out by the SWI/SNF complex. Furthermore, formaldehyde-assisted isolation of regulatory elements (FAIRE)-quantitative polymerase chain reaction (qPCR) experiments revealed that depletion of FLII caused a reduction in chromatin accessibility at the COL1A2 promoter. These results suggest that FLII plays a critical role in TGFβ/SMAD-mediated transcription of the COL1A2 gene

  1. TOBFAC: the database of tobacco transcription factors

    Directory of Open Access Journals (Sweden)

    Brannock Jennifer F

    2008-01-01

    Full Text Available Abstract Background Regulation of gene expression at the level of transcription is a major control point in many biological processes. Transcription factors (TFs can activate and/or repress the transcriptional rate of target genes and vascular plant genomes devote approximately 7% of their coding capacity to TFs. Global analysis of TFs has only been performed for three complete higher plant genomes – Arabidopsis (Arabidopsis thaliana, poplar (Populus trichocarpa and rice (Oryza sativa. Presently, no large-scale analysis of TFs has been made from a member of the Solanaceae, one of the most important families of vascular plants. To fill this void, we have analysed tobacco (Nicotiana tabacum TFs using a dataset of 1,159,022 gene-space sequence reads (GSRs obtained by methylation filtering of the tobacco genome. An analytical pipeline was developed to isolate TF sequences from the GSR data set. This involved multiple (typically 10–15 independent searches with different versions of the TF family-defining domain(s (normally the DNA-binding domain followed by assembly into contigs and verification. Our analysis revealed that tobacco contains a minimum of 2,513 TFs representing all of the 64 well-characterised plant TF families. The number of TFs in tobacco is higher than previously reported for Arabidopsis and rice. Results TOBFAC: the database of tobacco transcription factors, is an integrative database that provides a portal to sequence and phylogeny data for the identified TFs, together with a large quantity of other data concerning TFs in tobacco. The database contains an individual page dedicated to each of the 64 TF families. These contain background information, domain architecture via Pfam links, a list of all sequences and an assessment of the minimum number of TFs in this family in tobacco. Downloadable phylogenetic trees of the major families are provided along with detailed information on the bioinformatic pipeline that was used to find

  2. A new type of NtrC transcriptional activator.

    OpenAIRE

    Foster-Hartnett, D; Cullen, P. J.; Monika, E M; Kranz, R G

    1994-01-01

    The enteric NtrC (NRI) protein has been the paradigm for a class of bacterial enhancer-binding proteins (EBPs) that activate transcription of RNA polymerase containing the sigma 54 factor. Activators in the NtrC class are characterized by essentially three properties: (i) they bind to sites distant from the promoters that they activate (> 100 bp upstream of the transcriptional start site), (ii) they contain a conserved nucleotide-binding fold and exhibit ATPase activity that is required for a...

  3. 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

  4. XRP44X, an Inhibitor of Ras/Erk Activation of the Transcription Factor Elk3, Inhibits Tumour Growth and Metastasis in Mice

    Science.gov (United States)

    Cheung, Henry; Tourrette, Yves; Maas, Peter; Schalken, Jack A; van der Pluijm, Gabri

    2016-01-01

    Transcription factors have an important role in cancer but are difficult targets for the development of tumour therapies. These factors include the Ets family, and in this study Elk3 that is activated by Ras oncogene /Erk signalling, and is involved in angiogenesis, malignant progression and epithelial-mesenchymal type processes. We previously described the identification and in-vitro characterisation of an inhibitor of Ras / Erk activation of Elk3 that also affects microtubules, XRP44X. We now report an initial characterisation of the effects of XRP44X in-vivo on tumour growth and metastasis in three preclinical models mouse models, subcutaneous xenografts, intra-cardiac injection-bone metastasis and the TRAMP transgenic mouse model of prostate cancer progression. XRP44X inhibits tumour growth and metastasis, with limited toxicity. Tumours from XRP44X-treated animals have decreased expression of genes containing Elk3-like binding motifs in their promoters, Elk3 protein and phosphorylated Elk3, suggesting that perhaps XRP44X acts in part by inhibiting the activity of Elk3. Further studies are now warranted to develop XRP44X for tumour therapy. PMID:27427904

  5. Expression and characterization of SUMO-conjugated metal-responsive transcription factor 1: SIM-dependent cross-interaction and distinct DNA binding activity.

    Science.gov (United States)

    Lin, Chang-Yi; Liu, Ya-Chuan; Lin, Meng-Chieh; Thi Nguyen, Thao; Tam, Ming F; Chein, Chih-Yuan; Lin, Meng-Ting; Lin, Lih-Yuan

    2013-04-01

    Metal-responsive transcription factor 1 (MTF-1) regulates a variety of genes involving in metal homeostasis and oxidative stress. We have shown that MTF-1 can be conjugated by small ubiquitin-like modifier (SUMO) and forms complexes with cellular factor(s) in a SUMO-interacting motif (SIM)-dependent manner. To investigate whether the interaction of MTF-1 and its SUMO conjugate occurs, we expressed and isolated MTF-1 and sumoylated MTF-1 (S-MTF-1) for functional studies. Various conditions were examined to optimize the expressions of MTF-1 and S-MTF-1. Results from affinity column chromatography demonstrated that the unmodified MTF-1 consistently co-eluted with the S-MTF-1. Mutations at the SIM did not reduce the level of MTF-1 sumoylation but the sumoylated product can then be purified to homogeneity. The presence of MTF-1 cross-interaction was further supported by in vitro pull-down assays. The ability of the purified proteins in binding metal-responsive element (MRE) was assessed with electrophoretic mobility shift assay. Noticeably, MTF-1 required the presence of cell extracts to render the binding activity. However, S-MTF-1 binds MRE in void of other cellular factors. The same characteristic was found for MTF-1 with SUMO fusion at the carboxyl terminus. These results indicate that the presence of SUMO moiety allows the protein to interact directly with MRE.

  6. Death-domain associated protein-6 (DAXX) mediated apoptosis in hantavirus infection is counter-balanced by activation of interferon-stimulated nuclear transcription factors

    Energy Technology Data Exchange (ETDEWEB)

    Khaiboullina, Svetlana F., E-mail: sv.khaiboullina@gmail.com [Whittemore Peterson Institute, University of Nevada-Reno, Reno (United States); Morzunov, Sergey P. [Department of Pathology and Nevada State Health Laboratory, University of Nevada-Reno, Reno (United States); Boichuk, Sergei V. [Kazan State Medical University, Kazan (Russian Federation); Palotás, András [Asklepios-Med (private medical practice and research center), Szeged (Hungary); Jeor, Stephen St. [Department of Microbiology and Immunology, University of Nevada-Reno, Reno (United States); Lombardi, Vincent C. [Whittemore Peterson Institute, University of Nevada-Reno, Reno (United States); Rizvanov, Albert A. [Department of Genetics, Kazan (Volga Region) Federal University, Kazan (Russian Federation)

    2013-09-01

    Hantaviruses are negative strand RNA species that replicate predominantly in the cytoplasm. They also activate numerous cellular responses, but their involvement in nuclear processes is yet to be established. Using human umbilical vein endothelial cells (HUVECs), this study investigates the molecular finger-print of nuclear transcription factors during hantavirus infection. The viral-replication-dependent activation of pro-myelocytic leukemia protein (PML) was followed by subsequent localization in nuclear bodies (NBs). PML was also found in close proximity to activated Sp100 nuclear antigen and interferon-stimulated gene 20 kDa protein (ISG-20), but co-localization with death-domain associated protein-6 (DAXX) was not observed. These data demonstrate that hantavirus triggers PML activation and localization in NBs in the absence of DAXX-PLM-NB co-localization. The results suggest that viral infection interferes with DAXX-mediated apoptosis, and expression of interferon-activated Sp100 and ISG-20 proteins may indicate intracellular intrinsic antiviral attempts.

  7. Conjugated bilirubin affects cytokine profiles in hepatitis A virus infection by modulating function of signal transducer and activator of transcription factors.

    Science.gov (United States)

    Castro-García, Flor P; Corral-Jara, Karla F; Escobedo-Melendez, Griselda; Sandoval-Hernandez, Monserrat A; Rosenstein, Yvonne; Roman, Sonia; Panduro, Arturo; Fierro, Nora A

    2014-12-01

    Hepatitis A virus (HAV) infection is the major cause of acute liver failure in paediatric patients. The clinical spectrum of infection is variable, and liver injury is determined by altered hepatic enzyme function and bilirubin concentration. We recently reported differences in cytokine profiles between distinct HAV-induced clinical courses, and bilirubin has been recognized as a potential immune-modulator. However, how bilirubin may affect cytokine profiles underlying the variability in the course of infection has not been determined. Herein, we used a transcription factor (TF) binding site identification approach to retrospectively analyse cytokine expression in HAV-infected children and to predict the entire set of TFs associated with the expression of specific cytokine profiles. The results suggested that modulation of the activity of signal transducers and activators of transcription proteins (STATs) may play a central role during HAV infection. This led us to compare the degree of STAT phosphorylation in peripheral blood lymphoid cells (PBLCs) from paediatric patients with distinct levels of conjugated bilirubin (CB). Low CB levels in sera were associated with increased STAT-1 and STAT-5 phosphorylation. A positive correlation was observed between the serum interleukin-6 (IL-6) content and CB values, whereas higher levels of CB correlated with reduced serum IL-8 values and with a reduction in the proportion of PBLCs positive for STAT-5 phosphorylation. When CB was used to stimulate patients' PBLCs in vitro, the levels of IL-6 and tumour necrosis factor-α were increased. The data showed that bilirubin plays a role in STAT function and affects cytokine profile expression during HAV infection.

  8. Transcriptional profiling of intrinsic PNS factors in the postnatal mouse.

    Science.gov (United States)

    Smith, Robin P; Lerch-Haner, Jessica K; Pardinas, Jose R; Buchser, William J; Bixby, John L; Lemmon, Vance P

    2011-01-01

    Neurons in the peripheral nervous system (PNS) display a higher capacity to regenerate after injury than those in the central nervous system, suggesting cell specific transcriptional modules underlying axon growth and inhibition. We report a systems biology based search for PNS specific transcription factors (TFs). Messenger RNAs enriched in dorsal root ganglion (DRG) neurons compared to cerebellar granule neurons (CGNs) were identified using subtractive hybridization and DNA microarray approaches. Network and transcription factor binding site enrichment analyses were used to further identify TFs that may be differentially active. Combining these techniques, we identified 32 TFs likely to be enriched and/or active in the PNS. Twenty-five of these TFs were then tested for an ability to promote CNS neurite outgrowth in an overexpression screen. Real-time PCR and immunohistochemical studies confirmed that one representative TF, STAT3, is intrinsic to PNS neurons, and that constitutively active STAT3 is sufficient to promote CGN neurite outgrowth.

  9. TEAD/TEF transcription factors utilize the activation domain of YAP65, a Src/Yes-associated protein localized in the cytoplasm

    OpenAIRE

    Vassilev, Alex; Kaneko, Kotaro J.; Shu, Hongjun; Zhao, Yingming; DePamphilis, Melvin L.

    2001-01-01

    Mammals express four highly conserved TEAD/TEF transcription factors that bind the same DNA sequence, but serve different functions during development. TEAD-2/TEF-4 protein purified from mouse cells was associated predominantly with a novel TEAD-binding domain at the amino terminus of YAP65, a powerful transcriptional coactivator. YAP65 interacted specifically with the carboxyl terminus of all four TEAD proteins. Both this interaction and sequence-specific DNA binding by TEAD were required fo...

  10. Role of cysteines in the stability and DNA-binding activity of the hypochlorite-specific transcription factor HypT.

    Directory of Open Access Journals (Sweden)

    Adrian Drazic

    Full Text Available Reactive oxygen species are important components of the immune response. Hypochlorite (HOCl is produced by neutrophils to kill invading microorganisms. The bactericidal activity of HOCl is due to proteome-wide unfolding and oxidation of proteins at cysteine and methionine residues. Escherichia coli cells are protected from HOCl-killing by the previously identified dodecameric transcription factor HypT (YjiE. Here, we aimed to unravel whether HOCl activates HypT directly or via a reaction product of HOCl with a cellular component. Bacterial viability assays and analysis of target gene regulation indicate that HypT is highly specific to activation by HOCl and that no reaction products of HOCl such as monochloramine, hydroxyl radicals, or methionine sulfoxide activate HypT in vivo. Surprisingly, purified HypT lost its DNA-binding activity upon incubation with HOCl or reaction products that oxidize HypT to form a disulfide-linked dimer, and regained DNA-binding activity upon reduction. Thus, we postulate that the cysteines in HypT contribute to control the DNA-binding activity of HypT in vitro. HypT contains five cysteine residues; a HypT mutant with all cysteines substituted by serine is aggregation-prone and forms tetramers in addition to the typical dodecamers. Using single and multiple cysteine-to-serine mutants, we identified Cys150 to be required for stability and Cys4 being important for oligomerization of HypT to dodecamers. Further, oxidation of Cys4 is responsible for the loss of DNA-binding of HypT upon oxidation in vitro. It appears that Cys4 oxidation upon conditions that are insufficient to stimulate the DNA-binding activity of HypT prevents unproductive interactions of HypT with DNA. Thus, Cys4 oxidation may be a check point in the activation process of HypT.

  11. Brain-derived neurotrophic factor promotes vesicular glutamate transporter 3 expression and neurite outgrowth of dorsal root ganglion neurons through the activation of the transcription factors Etv4 and Etv5.

    Science.gov (United States)

    Liu, Dong; Liu, Zhen; Liu, Huaxiang; Li, Hao; Pan, Xinliang; Li, Zhenzhong

    2016-03-01

    Brain-derived neurotrophic factor (BDNF) is critical for sensory neuron survival and is necessary for vesicular glutamate transporter 3 (VGLUT3) expression. Whether the transcription factors Etv4 and Etv5 are involved in these BDNF-induced effects remains unclear. In the present study, primary cultured dorsal root ganglion (DRG) neurons were used to test the link between BDNF and transcription factors Etv4 and Etv5 on VGLUT3 expression and neurite outgrowth. BDNF promoted the mRNA and protein expression of Etv4 and Etv5 in DRG neurons. These effects were blocked by extracellular signal-regulated protein kinase 1/2 (ERK1/2) inhibitor PD98059 but not phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 or phospholipase C-γ (PLC-γ) inhibitor U73122. Etv4 siRNA and Etv5 siRNA effectively blocked the VGLUT3 expression and neurite elongation induced by BNDF. The overexpression of Etv4 or Etv5 potentiated the effects of BNDF-induced neurite elongation and growth-associated protein 43 (GAP-43), medium neurofilament (NF-M), and light neurofilament (NF-L) expression while these effects could be inhibited by Etv4 and Etv5 siRNA. These data imply that Etv4 and Etv5 are essential transcription factors in modulating BDNF/TrkB signaling-mediated VGLUT3 expression and neurite outgrowth. BDNF, through the ERK1/2 signaling pathway, activates Etv4 and Etv5 to initiate GAP-43 expression, promote neurofilament (NF) protein expression, induce neurite outgrowth, and mediate VGLUT3 expression for neuronal function improvement. The biological effects initiated by BDNF/TrkB signaling linked to E26 transformation-specific (ETS) transcription factors are important to elucidate neuronal differentiation, axonal regeneration, and repair in various pathological states.

  12. Adaptive evolution of transcription factor binding sites

    Directory of Open Access Journals (Sweden)

    Berg Johannes

    2004-10-01

    Full Text Available Abstract Background The regulation of a gene depends on the binding of transcription factors to specific sites located in the regulatory region of the gene. The generation of these binding sites and of cooperativity between them are essential building blocks in the evolution of complex regulatory networks. We study a theoretical model for the sequence evolution of binding sites by point mutations. The approach is based on biophysical models for the binding of transcription factors to DNA. Hence we derive empirically grounded fitness landscapes, which enter a population genetics model including mutations, genetic drift, and selection. Results We show that the selection for factor binding generically leads to specific correlations between nucleotide frequencies at different positions of a binding site. We demonstrate the possibility of rapid adaptive evolution generating a new binding site for a given transcription factor by point mutations. The evolutionary time required is estimated in terms of the neutral (background mutation rate, the selection coefficient, and the effective population size. Conclusions The efficiency of binding site formation is seen to depend on two joint conditions: the binding site motif must be short enough and the promoter region must be long enough. These constraints on promoter architecture are indeed seen in eukaryotic systems. Furthermore, we analyse the adaptive evolution of genetic switches and of signal integration through binding cooperativity between different sites. Experimental tests of this picture involving the statistics of polymorphisms and phylogenies of sites are discussed.

  13. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells

    Science.gov (United States)

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C.; Côté, Maxime C.; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-10-01

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors.

  14. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells.

    Science.gov (United States)

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C; Côté, Maxime C; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-10-14

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors.

  15. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells

    Science.gov (United States)

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C.; Côté, Maxime C.; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-01-01

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors. PMID:27739523

  16. Fisetin stimulates autophagic degradation of phosphorylated tau via the activation of TFEB and Nrf2 transcription factors

    OpenAIRE

    Sunhyo Kim; Ki Ju Choi; Sun-Jung Cho; Sang-Moon Yun; Jae-Pil Jeon; Young Ho Koh; Jihyun Song; Johnson, Gail V.W.; Chulman Jo

    2016-01-01

    The neuronal accumulation of phosphorylated tau plays a critical role in the pathogenesis of Alzheimer?s disease (AD). Here, we examined the effect of fisetin, a flavonol, on tau levels. Treatment of cortical cells or primary neurons with fisetin resulted in significant decreases in the levels of phosphorylated tau. In addition, fisetin decreased the levels of sarkosyl-insoluble tau in an active GSK-3?-induced tau aggregation model. However, there was no difference in activities of tau kinase...

  17. Fisetin stimulates autophagic degradation of phosphorylated tau via the activation of TFEB and Nrf2 transcription factors

    OpenAIRE

    Sunhyo Kim; Ki Ju Choi; Sun-Jung Cho; Sang-Moon Yun; Jae-Pil Jeon; Young Ho Koh; Jihyun Song; Johnson, Gail V.W.; Chulman Jo

    2016-01-01

    The neuronal accumulation of phosphorylated tau plays a critical role in the pathogenesis of Alzheimer’s disease (AD). Here, we examined the effect of fisetin, a flavonol, on tau levels. Treatment of cortical cells or primary neurons with fisetin resulted in significant decreases in the levels of phosphorylated tau. In addition, fisetin decreased the levels of sarkosyl-insoluble tau in an active GSK-3β-induced tau aggregation model. However, there was no difference in activities of tau kinase...

  18. Activation of Mechanosensitive Transcription Factors in Murine C2C12 Mesenchymal Precursors by Focused Low-Intensity Pulsed Ultrasound (FLIPUS).

    Science.gov (United States)

    Puts, Regina; Rikeit, Paul; Ruschke, Karen; Kadow-Romacker, Anke; Hwang, Soyoung; Jenderka, Klaus-Vitold; Knaus, Petra; Raum, Kay

    2016-10-01

    In this paper, we investigated the mechanoresponse of C2C12 mesenchymal precursor cells to focused low-intensity pulsed ultrasound (FLIPUS). The setup has been developed for in vitro stimulation of adherent cells in the defocused far field of the ultrasound propagating through the bottom of the well plate. Twenty-four-well tissue culture plates, carrying the cell monolayers, were incubated in a temperature-controlled water tank. The ultrasound was applied at 3.6-MHz frequency, pulsed at 100-Hz repetition frequency with a 27.8% duty cycle, and calibrated at an output intensity of ISATA = 44.5 ±7.1 mW/cm(2). Numerical sound propagation simulations showed no generation of standing waves in the well plate. The response of murine C2C12 cells to FLIPUS was evaluated by measuring activation of mechanosensitive transcription factors, i.e., activator protein-1 (AP-1), specificity protein 1 (Sp1), and transcriptional enhancer factor (TEAD), and expression of mechanosensitive genes, i.e., c-fos, c-jun, heparin binding growth associated molecule (HB-GAM), and Cyr-61. FLIPUS induced 50% ( p ≤ 0.05 ) and 70% ( p ≤ 0.05 ) increases in AP-1 and TEAD promoter activities, respectively, when stimulated for 5 min. The Sp1 activity was enhanced by about 20% ( p ≤ 0.05 ) after 5-min FLIPUS exposure and the trend persisted for 30-min ( p ≤ 0.05 ) and 1-h ( p ≤ 0.05 ) stimulation times. Expressions of mechanosensitive genes c-fos ( p ≤ 0.05 ), c-jun ( p ≤ 0.05 ), HB-GAM ( p ≤ 0.05 ), and cystein-rich protein 61 ( p ≤ 0.05 ) were enhanced in response to 5-min FLIPUS stimulation. The increase in proliferation of C2C12s occurred after the FLIPUS stimulation ( p ≤ 0.05 ), with AP-1, Sp1, and TEAD possibly regulating the observed cellular activities.

  19. 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).

  20. The world according to GARP transcription factors.

    Science.gov (United States)

    Safi, Alaeddine; Medici, Anna; Szponarski, Wojciech; Ruffel, Sandrine; Lacombe, Benoît; Krouk, Gabriel

    2017-10-01

    Plant specific GARP transcription factor family (made of ARR-B and G2-like) contains genes with very diverse in planta functions: nutrient sensing, root and shoot development, floral transition, chloroplast development, circadian clock oscillation maintenance, hormonal transport and signaling. In this work we review: first, their structural but distant relationships with MYB transcription factors, second, their role in planta, third, the diversity of their Cis-regulatory elements, fourth, their potential protein partners. We conclude that the GARP family may hold keys to understand the interactions between nutritional signaling pathways (nitrogen and phosphate at least) and development. Understanding how plant nutrition and development are coordinated is central to understand how to adapt plants to an ever-changing environment. Consequently GARPs are likely to attract increasing research attentions, as they are likely at the crossroads of these fundamental processes. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Co-operation of the transcription factor hepatocyte nuclear factor-4 with Sp1 or Sp3 leads to transcriptional activation of the human haem oxygenase-1 gene promoter in a hepatoma cell line.

    Science.gov (United States)

    Takahashi, Shigeru; Matsuura, Naomi; Kurokawa, Takako; Takahashi, Yuji; Miura, Takashi

    2002-11-01

    We reported previously that the 5'-flanking region (nucleotides -1976 to -1655) of the human haem oxygenase-1 ( hHO-1 ) gene enhances hHO-1 promoter activity in human hepatoma HepG2 cells, but not in HeLa cells [Takahashi, Takahashi, Ito, Nagano, Shibahara and Miura (1999) Biochim. Biophys. Acta 1447, 231-235]. To define more precisely the regulatory elements involved, in the present study we have functionally dissected this region and localized the enhancer to a 50 bp fragment (-1793 to -1744). Site-direct mutagenesis analysis revealed that two regions were responsible for this enhancer activity, i.e. a hepatocyte nuclear factor-4 (HNF-4) homologous region and a GC box motif homologous region. Mutation in either region alone moderately decreased enhancer activity. However, mutations in both regions reduced promoter activity to the basal level. Electrophoretic mobility-shift assays demonstrated that the P5-2 fragment (-1793 to -1744) interacted with at least two nuclear factors, i.e. HNF-4 and Sp1/Sp3. Co-transfection experiments using Drosophila SL2 cells revealed that HNF-4 and Sp1/Sp3 synergistically stimulated the enhancer activity of the P5-2 fragment. These results indicate that co-operation of HNF-4 with Sp1 or Sp3 leads to the activation of hHO-1 gene expression in hepatoma cells.

  2. HTF: A b-ZIP transcription factor that is closely related to the human XBP/TREB5 and is activated by hepatocellular carcinoma in rats.

    Science.gov (United States)

    Kishimoto, T; Kokura, K; Kumagai, Y; Wakamatsu, T; Makino, Y; Tamura, T

    1996-06-25

    We screened for rat hepatocellular carcinoma (HCC)-related genes by a novel cDNA subtraction method and obtained one gene. This gene was transcribed as 2.0- and 2.5-kb mRNAs, and its transcription was specifically enhanced in HCC. These cDNAs had the same open reading frame, but the 2.5 kb transcript had an extra 495 bases of 5'-UTR at the 5'-terminus. The deduced aa sequence revealed a basic-leucine zipper (b-ZIP) and proline/glutamine-rich structures, both of which are characteristic motifs for transcription factors. We designated the translation product of this gene HTF (Hepatocarcinogenesis-related Transcription Factor). Electrophoretic mobility shift assay demonstrated the DNA-binding ability of the recombinant HTF. It is most interesting that HTF had a considerable homology with human XBP/TREB5, which has been reported to be a binding factor for the X-box of the MHC class II gene and for the 21-bp enhancer of the HTLV-1 LTR. Genomic Southern analysis suggested that the 2.0- and 2.5-kb mRNAs are transcribed by a dual promoter of a single gene. Our results may suggest that HTF is a b-ZIP-type transcription factor involved in rat hepatocellular carcinoma.

  3. 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.

  4. 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.

  5. c-MYC Generates Repair Errors via Increased Transcription of Alternative-NHEJ Factors, LIG3 and PARP1, in Tyrosine Kinase-Activated Leukemias.

    Science.gov (United States)

    Muvarak, Nidal; Kelley, Shannon; Robert, Carine; Baer, Maria R; Perrotti, Danilo; Gambacorti-Passerini, Carlo; Civin, Curt; Scheibner, Kara; Rassool, Feyruz V

    2015-04-01

    Leukemias expressing the constitutively activated tyrosine kinases (TK) BCR-ABL1 and FLT3/ITD activate signaling pathways that increase genomic instability through generation of reactive oxygen species (ROS), DNA double-strand breaks (DSB), and error-prone repair. The nonhomologous end-joining (NHEJ) pathway is a major pathway for DSB repair and is highly aberrant in TK-activated leukemias; an alternative form of NHEJ (ALT-NHEJ) predominates, evidenced by increased expression of DNA ligase IIIα (LIG3) and PARP1, increased frequency of large genomic deletions, and repair using DNA sequence microhomologies. This study, for the first time, demonstrates that the TK target c-MYC plays a role in transcriptional activation and subsequent expression of LIG3 and PARP1 and contributes to the increased error-prone repair observed in TK-activated leukemias. c-MYC negatively regulates microRNAs miR-150 and miR-22, which demonstrate an inverse correlation with LIG3 and PARP1 expression in primary and cultured leukemia cells and chronic myelogenous leukemia human patient samples. Notably, inhibition of c-MYC and overexpression of miR-150 and -22 decreases ALT-NHEJ activity. Thus, BCR-ABL1 or FLT3/ITD induces c-MYC expression, leading to genomic instability via augmented expression of ALT-NHEJ repair factors that generate repair errors. In the context of TK-activated leukemias, c-MYC contributes to aberrant DNA repair through downstream targets LIG3 and PARP1, which represent viable and attractive therapeutic targets. ©2015 American Association for Cancer Research.

  6. Cooperative antiproliferative signaling by aspirin and indole-3-carbinol targets microphthalmia-associated transcription factor gene expression and promoter activity in human melanoma cells.

    Science.gov (United States)

    Poindexter, Kevin M; Matthew, Susanne; Aronchik, Ida; Firestone, Gary L

    2016-04-01

    Antiproliferative signaling of combinations of the nonsteroidal anti-inflammatory drug acetylsalicylic acid (aspirin) and indole-3-carbinol (I3C), a natural indolecarbinol compound derived from cruciferous vegetables, was investigated in human melanoma cells. Melanoma cell lines with distinct mutational profiles were sensitive to different extents to the antiproliferative response of aspirin, with oncogenic BRAF-expressing G361 cells and wild-type BRAF-expressing SK-MEL-30 cells being the most responsive. I3C triggered a strong proliferative arrest of G361 melanoma cells and caused only a modest decrease in the proliferation of SK-MEL-30 cells. In both cell lines, combinations of aspirin and I3C cooperatively arrested cell proliferation and induced a G1 cell cycle arrest, and nearly ablated protein and transcript levels of the melanocyte master regulator microphthalmia-associated transcription factor isoform M (MITF-M). In melanoma cells transfected with a -333/+120-bp MITF-M promoter-luciferase reporter plasmid, treatment with aspirin and I3C cooperatively disrupted MITF-M promoter activity, which accounted for the loss of MITF-M gene products. Mutational analysis revealed that the aspirin required the LEF1 binding site, whereas I3C required the BRN2 binding site to mediate their combined and individual effects on MITF-M promoter activity. Consistent with LEF1 being a downstream effector of Wnt signaling, aspirin, but not I3C, downregulated protein levels of the Wnt co-receptor LDL receptor-related protein-6 and β-catenin and upregulated the β-catenin destruction complex component Axin. Taken together, our results demonstrate that aspirin-regulated Wnt signaling and I3C-targeted signaling pathways converge at distinct DNA elements in the MITF-M promoter to cooperatively disrupt MITF-M expression and melanoma cell proliferation.

  7. [All signs of metabolic syndrome in the hypertensive ISIAH rats are associated with increased activity of transcription factors PPAR, LXR, PXR, and CAR in the liver].

    Science.gov (United States)

    Pivovarova, E N; Dushkin, M I; Perepechaeva, M L; Kobzev, V F; Trufakin, V A; Markel', A L

    2011-01-01

    It is known that the metabolic syndrome (MS), which includes hypertension, dislipidemia, glucose intolerance, and obesity leads to cardiovascular diseases. The MS risk is growing catastrophically. Molecular mechanisms allowing to understand the reason of integrated dysfunctions, taking place at MS cases, have remained almost unstudied. The chronical stress plays a crucial role in MS development; therefore in the present work a hypertensive rat strain with Inherited Stress-Induced Arterial Hypertension (ISIAH) was used as a model. It was shown that ISIAH rat strain as compared with the control WAG rat strain is characterized by increased content of triglyceride, VLDL and LDL cholesterols, a decreased content of HDL cholesterol, a high level of apolipoprotein B-100, and decreased level of apolipoprotein A-I. The ISIAH rats body weight was higher as compared with WAG rats; ISIAH rats blood glucose content was higher too. Thus, strain hypertension for ISIAH rat is accompanied by dislipidemia, increased glucose content, and increased body weight, representing a whole set of MS signs. Since at MS cases the systemic abnormalities in lipid and carbohydrate metabolism take place, the functional activity of transcription factors (TFs) participating in integral regulation of lipid and carbohydrate metabolism genes in liver was measured. PPAR, LXR, PXR, CAR DNA-binding activity was increased in ISIAH rats, suggesting involvement of these TFs in MS development. Integrated investigation of PPAR, LXR, PXR, CAR regulatory mechanisms, signal transduction and transcriptional targets will provide insights into the pathogenesis of MS and offer valuable information for designing of drugs for MS treatment.

  8. 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.

  9. Heterodimerization of the transcription factors E2F-1 and DP-1 leads to cooperative trans-activation

    DEFF Research Database (Denmark)

    Helin, K; Wu, C L; Fattaey, A R;

    1993-01-01

    the hypophosphorylated form of the retinoblastoma protein (pRB). The other protein, murine DP-1, was purified from an E2F DNA-affinity column, and it was subsequently shown to bind the consensus E2F DNA-binding site. To study a possible interaction between E2F-1 and DP-1, we have now isolated a cDNA for the human...... is required for stable interaction with pRB in vivo and that trans-activation by E2F-1/DP-1 heterodimers is inhibited by pRB. We suggest that "E2F" is the activity that is formed when an E2F-1-related protein and a DP-1-related protein dimerize....

  10. Heterodimerization of the transcription factors E2F-1 and DP-1 leads to cooperative trans-activation

    DEFF Research Database (Denmark)

    Helin, K; Wu, C L; Fattaey, A R

    1993-01-01

    homolog of DP-1. Human DP-1 and E2F-1 associate both in vivo and in vitro, and this interaction leads to enhanced binding to E2F DNA-binding sites. The association of E2F-1 and DP-1 leads to cooperative activation of an E2F-responsive promoter. Finally, we demonstrate that E2F-1 and DP-1 association...

  11. Systematic genetic analysis of transcription factors to map the fission yeast transcription-regulatory network.

    Science.gov (United States)

    Chua, Gordon

    2013-12-01

    Mapping transcriptional-regulatory networks requires the identification of target genes, binding specificities and signalling pathways of transcription factors. However, the characterization of each transcription factor sufficiently for deciphering such networks remains laborious. The recent availability of overexpression and deletion strains for almost all of the transcription factor genes in the fission yeast Schizosaccharomyces pombe provides a valuable resource to better investigate transcription factors using systematic genetics. In the present paper, I review and discuss the utility of these strain collections combined with transcriptome profiling and genome-wide chromatin immunoprecipitation to identify the target genes of transcription factors.

  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. Comparison of the molecular topologies of stress-activated transcription factors HSF1, AP-1, NRF2, and NF-κB in their induction kinetics of HMOX1.

    Science.gov (United States)

    Pronk, Tessa E; van der Veen, Jochem W; Vandebriel, Rob J; van Loveren, Henk; de Vink, Erik P; Pennings, Jeroen L A

    2014-10-01

    For cells, reacting aptly to changes in their environment is of critical importance. The protein Heme oxygenase-1 (HMOX1) plays a critical role as a guard of cellular homeostasis and is considered as a reliable indicator of cellular oxidative stress. A better insight in the regulation of HMOX1 would assist in understanding the physiological role of HMOX1 as well as improving functional interpretation of the gene as a biomarker in toxicogenomics. Remarkably, as many as four transcription factors are known to regulate the HMOX1 gene: HSF1, AP-1, NRF2, and NF-κB. To investigate induction kinetics of these transcription factors, we constructed mathematical simulation models for each of them. We included the topology of the known interactions of molecules involved in the activation of the transcription factors, and the feedback loops resulting in their down-regulation. We evaluate how the molecular circuitries associated with the different transcription factors differ in their kinetics regarding HMOX1 induction, under different scenarios of acute and less acute stress. We also evaluate the combined effect of the four transcription factors on HMOX1 expression and the resulting alleviation of stress. Overall, the results support the assumption of different biological roles for the four transcription factors, with AP-1 being a fast acting general stress response protein at the expense of efficiency, and NRF2 being important for cellular homeostasis in maintaining low levels of oxidative stress.

  14. Transcriptional activators enhance polyadenylation of mRNA precursors

    OpenAIRE

    Nagaike, Takashi; Manley, James L.

    2011-01-01

    3′ processing of mRNA precursors is frequently coupled to transcription by RNA polymerase II (RNAP II). This coupling is well known to involve the C-terminal domain of the RNAP II largest subunit, but a variety of other transcription-associated factors have also been suggested to mediate coupling. Our recent studies have provided direct evidence that transcriptional activators can enhance the efficiency of transcription-coupled 3′ processing. In this point-of-view, we discuss the mechanisms t...

  15. Cooperative binding of transcription factors promotes bimodal gene expression response.

    Directory of Open Access Journals (Sweden)

    Pablo S Gutierrez

    Full Text Available In the present work we extend and analyze the scope of our recently proposed stochastic model for transcriptional regulation, which considers an arbitrarily complex cis-regulatory system using only elementary reactions. Previously, we determined the role of cooperativity on the intrinsic fluctuations of gene expression for activating transcriptional switches, by means of master equation formalism and computer simulation. This model allowed us to distinguish between two cooperative binding mechanisms and, even though the mean expression levels were not affected differently by the acting mechanism, we showed that the associated fluctuations were different. In the present generalized model we include other regulatory functions in addition to those associated to an activator switch. Namely, we introduce repressive regulatory functions and two theoretical mechanisms that account for the biphasic response that some cis-regulatory systems show to the transcription factor concentration. We have also extended our previous master equation formalism in order to include protein production by stochastic translation of mRNA. Furthermore, we examine the graded/binary scenarios in the context of the interaction energy between transcription factors. In this sense, this is the first report to show that the cooperative binding of transcription factors to DNA promotes the "all-or-none" phenomenon observed in eukaryotic systems. In addition, we confirm that gene expression fluctuation levels associated with one of two cooperative binding mechanism never exceed the fluctuation levels of the other.

  16. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  18. The transcription factors MS188 and AMS form a complex to activate the expression of CYP703A2 for sporopollenin biosynthesis in Arabidopsis thaliana.

    Science.gov (United States)

    Xiong, Shuang-Xi; Lu, Jie-Yang; Lou, Yue; Teng, Xiao-Dong; Gu, Jing-Nan; Zhang, Cheng; Shi, Qiang-Sheng; Yang, Zhong-Nan; Zhu, Jun

    2016-12-01

    The sexine layer of pollen grain is mainly composed of sporopollenins. The sporophytic secretory tapetum is required for the biosynthesis of sporopollenin. Although several enzymes involved in sporopollenin biosynthesis have been reported, the regulatory mechanism of these enzymes in tapetal layer remains elusive. ABORTED MICROSPORES (AMS) and MALE STERILE 188/MYB103/MYB80 (MS188/MYB103/MYB80) are two tapetal cell-specific transcription factors required for pollen wall formation. AMS functions upstream of MS188. Here we report that AMS and MS188 target the CYP703A2 gene, which is involved in sporopollenin biosynthesis. We found that AMS and MS188 were localized in tapetum while CYP703A2 was localized in both tapetum and locule. Chromatin immunoprecipitation (ChIP) showed that MS188 directly bound to the promoter of CYP703A2 and luciferase-inducible assay showed that MS188 activated the expression of CYP703A2. Yeast two-hybrid and electrophoretic mobility shift assays (EMSAs) further demonstrated that MS188 complexed with AMS. The expression of CYP703A2 could be partially restored by the elevated levels of MS188 in the ams mutant. Therefore, our data reveal that MS188 coordinates with AMS to activate CYP703A2 in sporopollenin biosynthesis of plant tapetum.

  19. Gomisin A enhances tumor necrosis factor-α-induced G1 cell cycle arrest via signal transducer and activator of transcription 1-mediated phosphorylation of retinoblastoma protein.

    Science.gov (United States)

    Waiwut, Pornthip; Shin, Myoung-Sook; Yokoyama, Satoru; Saiki, Ikuo; Sakurai, Hiroaki

    2012-01-01

    Gomisin A, a dibenzocyclooctadiene lignan isolated from the fruit of Schisandra chinensis, has been reported as an anti-cancer substance. In this study, we investigated the effects of gomisin A on cancer cell proliferation and cell cycle arrest in HeLa cells. Gomisin A significantly inhibited cell proliferation in a dose-dependent manner after 72 h treatment, especially in the presence of tumor necrosis factor-α (TNF-α), due to cell cycle arrest in the G1 phase with the downregulation of cyclin D1 expression and Retinoblastoma (RB) phosphorylation. In addition, gomisin A in combination with TNF-α strongly suppressed the expression of signal transducer and activator of transcription 1 (STAT1). Inhibition of STAT1 pathways by a small-interfering RNA against STAT1 and AG490 Janus kinase (JAK) kinase inhibitor AG490 reduced the cyclin D1 expression and RB phosphorylation, indicating that JAK-mediated STAT1 activation is involved in gomisin A-induced G1 cell cycle arrest.

  20. The promoter of the white spot syndrome virus immediate-early gene WSSV108 is activated by the cellular KLF transcription factor.

    Science.gov (United States)

    Liu, Wang-Jing; Lo, Chu-Fang; Kou, Guang-Hsiung; Leu, Jiann-Horng; Lai, Ying-Jang; Chang, Li-Kwan; Chang, Yun-Shiang

    2015-03-01

    A series of deletion and mutation assays of the white spot syndrome virus (WSSV) immediate-early gene WSSV108 promoter showed that a Krüppel-like factor (KLF) binding site located from -504 to -495 (relative to the transcription start site) is important for the overall level of WSSV108 promoter activity. Electrophoretic mobility shift assays further showed that overexpressed recombinant Penaeus monodon KLF (rPmKLF) formed a specific protein-DNA complex with the (32)P-labeled KLF binding site of the WSSV108 promoter, and that higher levels of Litopenaeus vannamei KLF (LvKLF) were expressed in WSSV-infected shrimp. A transactivation assay indicated that the WSSV108 promoter was strongly activated by rPmKLF in a dose-dependent manner. Lastly, we found that specific silencing of LvKLF expression in vivo by dsRNA injection dramatically reduced both WSSV108 expression and WSSV replication. We conclude that shrimp KLF is important for WSSV genome replication and gene expression, and that it binds to the WSSV108 promoter to enhance the expression of this immediate-early gene.

  1. Cdk phosphorylation of the Ste11 transcription factor constrains differentiation-specific transcription to G1

    DEFF Research Database (Denmark)

    Kjaerulff, Søren; Andersen, Nicoline Resen; Borup, Mia Trolle;

    2007-01-01

    Eukaryotic cells normally differentiate from G(1); here we investigate the mechanism preventing expression of differentiation-specific genes outside G(1). In fission yeast, induction of the transcription factor Ste11 triggers sexual differentiation. We find that Ste11 is only active in G(1) when...... S phase. When we mutated T82 to aspartic acid, mimicking constant phosphorylation, cells no longer underwent differentiation. Conversely, changing T82 to alanine rendered Ste11-controlled transcription constitutive through the cell cycle, and allowed mating from S phase with increased frequency...

  2. Systematic identification of transcription factors associated with patient survival in cancers

    Directory of Open Access Journals (Sweden)

    Alves Pedro

    2009-05-01

    Full Text Available Abstract Background Aberrant activation or expression of transcription factors has been implicated in the tumorigenesis of various types of cancer. In spite of the prevalent application of microarray experiments for profiling gene expression in cancer samples, they provide limited information regarding the activities of transcription factors. However, the association between transcription factors and cancers is largely dependent on the transcription regulatory activities rather than mRNA expression levels. Results In this paper, we propose a computational approach that integrates microarray expression data with the transcription factor binding site information to systematically identify transcription factors associated with patient survival given a specific cancer type. This approach was applied to two gene expression data sets for breast cancer and acute myeloid leukemia. We found that two transcription factor families, the steroid nuclear receptor family and the ATF/CREB family, are significantly correlated with the survival of patients with breast cancer; and that a transcription factor named T-cell acute lymphocytic leukemia 1 is significantly correlated with acute myeloid leukemia patient survival. Conclusion Our analysis identifies transcription factors associating with patient survival and provides insight into the regulatory mechanism underlying the breast cancer and leukemia. The transcription factors identified by our method are biologically meaningful and consistent with prior knowledge. As an insightful tool, this approach can also be applied to other microarray cancer data sets to help researchers better understand the intricate relationship between transcription factors and diseases.

  3. Isl1 is a direct transcriptional target of Forkhead transcription factors in second heart field-derived mesoderm

    Science.gov (United States)

    Kang, Jione; Nathan, Elisha; Xu, Shan-Mei; Tzahor, Eldad; Black, Brian L.

    2009-01-01

    The cells of the second heart field (SHF) contribute to the outflow tract and right ventricle, as well as to parts of the left ventricle and atria. Isl1, a member of the LIM-homeodomain transcription factor family, is expressed early in this cardiac progenitor population and functions near the top of a transcriptional pathway essential for heart development. Isl1 is required for the survival and migration of SHF-derived cells into the early developing heart at the inflow and outflow poles. Despite this important role for Isl1 in early heart formation, the transcriptional regulation of Isl1 has remained largely undefined. Therefore, to identify transcription factors that regulate Isl1 expression in vivo, we screened the conserved noncoding sequences from the mouse Isl1 locus for enhancer activity in transgenic mouse embryos. Here, we report the identification of an enhancer from the mouse Isl1 gene that is sufficient to direct expression to the SHF and its derivatives. The Isl1 SHF enhancer contains three consensus Forkhead transcription factor binding sites that are efficiently and specifically bound by Forkhead transcription factors. Importantly, the activity of the enhancer is dependent on these three Forkhead binding sites in transgenic mouse embryos. Thus, these studies demonstrate that Isl1 is a direct transcriptional target of Forkhead transcription factors in the SHF and establish a transcriptional pathway upstream of Isl1 in the SHF. PMID:19580802

  4. Uncovering Transcriptional Regulatory Networks by Sparse Bayesian Factor Model

    Science.gov (United States)

    Meng, Jia; Zhang, Jianqiu(Michelle); Qi, Yuan(Alan); Chen, Yidong; Huang, Yufei

    2010-12-01

    The problem of uncovering transcriptional regulation by transcription factors (TFs) based on microarray data is considered. A novel Bayesian sparse correlated rectified factor model (BSCRFM) is proposed that models the unknown TF protein level activity, the correlated regulations between TFs, and the sparse nature of TF-regulated genes. The model admits prior knowledge from existing database regarding TF-regulated target genes based on a sparse prior and through a developed Gibbs sampling algorithm, a context-specific transcriptional regulatory network specific to the experimental condition of the microarray data can be obtained. The proposed model and the Gibbs sampling algorithm were evaluated on the simulated systems, and results demonstrated the validity and effectiveness of the proposed approach. The proposed model was then applied to the breast cancer microarray data of patients with Estrogen Receptor positive ([InlineEquation not available: see fulltext.]) status and Estrogen Receptor negative ([InlineEquation not available: see fulltext.]) status, respectively.

  5. Uncovering Transcriptional Regulatory Networks by Sparse Bayesian Factor Model

    Directory of Open Access Journals (Sweden)

    Qi Yuan(Alan

    2010-01-01

    Full Text Available Abstract The problem of uncovering transcriptional regulation by transcription factors (TFs based on microarray data is considered. A novel Bayesian sparse correlated rectified factor model (BSCRFM is proposed that models the unknown TF protein level activity, the correlated regulations between TFs, and the sparse nature of TF-regulated genes. The model admits prior knowledge from existing database regarding TF-regulated target genes based on a sparse prior and through a developed Gibbs sampling algorithm, a context-specific transcriptional regulatory network specific to the experimental condition of the microarray data can be obtained. The proposed model and the Gibbs sampling algorithm were evaluated on the simulated systems, and results demonstrated the validity and effectiveness of the proposed approach. The proposed model was then applied to the breast cancer microarray data of patients with Estrogen Receptor positive ( status and Estrogen Receptor negative ( status, respectively.

  6. Graded Proteasome Dysfunction in Caenorhabditis elegans Activates an Adaptive Response Involving the Conserved SKN-1 and ELT-2 Transcription Factors and the Autophagy-Lysosome Pathway.

    Science.gov (United States)

    Keith, Scott A; Maddux, Sarah K; Zhong, Yayu; Chinchankar, Meghna N; Ferguson, Annabel A; Ghazi, Arjumand; Fisher, Alfred L

    2016-02-01

    The maintenance of cellular proteins in a biologically active and structurally stable state is a vital endeavor involving multiple cellular pathways. One such pathway is the ubiquitin-proteasome system that represents a major route for protein degradation, and reductions in this pathway usually have adverse effects on the health of cells and tissues. Here, we demonstrate that loss-of-function mutants of the Caenorhabditis elegans proteasome subunit, RPN-10, exhibit moderate proteasome dysfunction and unexpectedly develop both increased longevity and enhanced resistance to multiple threats to the proteome, including heat, oxidative stress, and the presence of aggregation prone proteins. The rpn-10 mutant animals survive through the activation of compensatory mechanisms regulated by the conserved SKN-1/Nrf2 and ELT-2/GATA transcription factors that mediate the increased expression of genes encoding proteasome subunits as well as those mediating oxidative- and heat-stress responses. Additionally, we find that the rpn-10 mutant also shows enhanced activity of the autophagy-lysosome pathway as evidenced by increased expression of the multiple autophagy genes including atg-16.2, lgg-1, and bec-1, and also by an increase in GFP::LGG-1 puncta. Consistent with a critical role for this pathway, the enhanced resistance of the rpn-10 mutant to aggregation prone proteins depends on autophagy genes atg-13, atg-16.2, and prmt-1. Furthermore, the rpn-10 mutant is particularly sensitive to the inhibition of lysosome activity via either RNAi or chemical means. We also find that the rpn-10 mutant shows a reduction in the numbers of intestinal lysosomes, and that the elt-2 gene also plays a novel and vital role in controlling the production of functional lysosomes by the intestine. Overall, these experiments suggest that moderate proteasome dysfunction could be leveraged to improve protein homeostasis and organismal health and longevity, and that the rpn-10 mutant provides a unique

  7. Graded Proteasome Dysfunction in Caenorhabditis elegans Activates an Adaptive Response Involving the Conserved SKN-1 and ELT-2 Transcription Factors and the Autophagy-Lysosome Pathway.

    Directory of Open Access Journals (Sweden)

    Scott A Keith

    2016-02-01

    Full Text Available The maintenance of cellular proteins in a biologically active and structurally stable state is a vital endeavor involving multiple cellular pathways. One such pathway is the ubiquitin-proteasome system that represents a major route for protein degradation, and reductions in this pathway usually have adverse effects on the health of cells and tissues. Here, we demonstrate that loss-of-function mutants of the Caenorhabditis elegans proteasome subunit, RPN-10, exhibit moderate proteasome dysfunction and unexpectedly develop both increased longevity and enhanced resistance to multiple threats to the proteome, including heat, oxidative stress, and the presence of aggregation prone proteins. The rpn-10 mutant animals survive through the activation of compensatory mechanisms regulated by the conserved SKN-1/Nrf2 and ELT-2/GATA transcription factors that mediate the increased expression of genes encoding proteasome subunits as well as those mediating oxidative- and heat-stress responses. Additionally, we find that the rpn-10 mutant also shows enhanced activity of the autophagy-lysosome pathway as evidenced by increased expression of the multiple autophagy genes including atg-16.2, lgg-1, and bec-1, and also by an increase in GFP::LGG-1 puncta. Consistent with a critical role for this pathway, the enhanced resistance of the rpn-10 mutant to aggregation prone proteins depends on autophagy genes atg-13, atg-16.2, and prmt-1. Furthermore, the rpn-10 mutant is particularly sensitive to the inhibition of lysosome activity via either RNAi or chemical means. We also find that the rpn-10 mutant shows a reduction in the numbers of intestinal lysosomes, and that the elt-2 gene also plays a novel and vital role in controlling the production of functional lysosomes by the intestine. Overall, these experiments suggest that moderate proteasome dysfunction could be leveraged to improve protein homeostasis and organismal health and longevity, and that the rpn-10 mutant

  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. Statistics for Transcription Factor Binding Sites

    OpenAIRE

    2008-01-01

    Transcription factors (TFs) play a key role in gene regulation. They interact with specific binding sites or motifs on the DNA sequence and regulate expression of genes downstream of these binding sites. In silico prediction of potential binding of a TF to a binding site is an important task in computational biology. From a statistical point of view, the DNA sequence is a long text consisting of four different letters ('A','C','G', and 'T'). The binding of a TF to the sequence corresponds to ...

  10. Zinc finger artificial transcription factor-based nearest inactive analogue/nearest active analogue strategy used for the identification of plant genes controlling homologous recombination.

    Science.gov (United States)

    Jia, Qi; van Verk, Marcel C; Pinas, Johan E; Lindhout, Beatrice I; Hooykaas, Paul J J; van der Zaal, Bert J

    2013-12-01

    In previous work, we selected a particular transcription factor, designated VP16-HRU, from a pool of zinc finger artificial transcription factors (ZF-ATFs) used for genome interrogation. When expressed in Arabidopsis thaliana under control of the ribosomal protein S5A promoter, the RPS5A::VP16-HRU construct led to a 200- to 300-fold increase in the frequency of somatic intrachromosomal homologous recombination (iHR). Because the expression of each ZF-ATF leads to a large number of transcriptional changes, we designed a strategy employing a collection of structurally similar ZF-ATFs to filter out the transcriptional changes relevant to the phenotype by deep sequencing. In that manner, 30 transcripts were found to be consistently induced in plants with enhanced homologous recombination (HR). For 25 of the cognate genes, their effect on the HR process was assessed using cDNA/gDNA expression constructs. For three genes, ectopic expression indeed led to enhanced iHR frequencies, albeit much lower than the frequency observed when a HR-inducing ZF-ATF was present. Altogether, our data demonstrate that despite the large number of transcriptional changes brought about by individual ZF-ATFs, causal changes can be identified. In our case, the picture emerged that a natural regulatory switch for iHR does not exist but that ZF-ATFs-like VP16-HRU act as an ectopic master switch, orchestrating the timely expression of a set of plant genes that each by themselves only have modest effects, but when acting together support an extremely high iHR frequency.

  11. 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

  12. 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.

  13. Interference with Activator Protein-2 transcription factors leads to induction of apoptosis and an increase in chemo- and radiation- sensitivity in breast cancer cells

    LENUS (Irish Health Repository)

    Thewes, Verena

    2010-05-11

    Abstract Background Activator Protein-2 (AP-2) transcription factors are critically involved in a variety of fundamental cellular processes such as proliferation, differentiation and apoptosis and have also been implicated in carcinogenesis. Expression of the family members AP-2α and AP-2γ is particularly well documented in malignancies of the female breast. Despite increasing evaluation of single AP-2 isoforms in mammary tumors the functional role of concerted expression of multiple AP-2 isoforms in breast cancer remains to be elucidated. AP-2 proteins can form homo- or heterodimers, and there is growing evidence that the net effect whether a cell will proliferate, undergo apoptosis or differentiate is partly dependent on the balance between different AP-2 isoforms. Methods We simultaneously interfered with all AP-2 isoforms expressed in ErbB-2-positive murine N202.1A breast cancer cells by conditionally over-expressing a dominant-negative AP-2 mutant. Results We show that interference with AP-2 protein function lead to reduced cell number, induced apoptosis and increased chemo- and radiation-sensitivity. Analysis of global gene expression changes upon interference with AP-2 proteins identified 139 modulated genes (90 up-regulated, 49 down-regulated) compared with control cells. Gene Ontology (GO) investigations for these genes revealed Cell Death and Cell Adhesion and Migration as the main functional categories including 25 and 12 genes, respectively. By using information obtained from Ingenuity Pathway Analysis Systems we were able to present proven or potential connections between AP-2 regulated genes involved in cell death and response to chemo- and radiation therapy, (i.e. Ctgf, Nrp1, Tnfaip3, Gsta3) and AP-2 and other main apoptosis players and to create a unique network. Conclusions Expression of AP-2 transcription factors in breast cancer cells supports proliferation and contributes to chemo- and radiation-resistance of tumor cells by impairing the

  14. Influence of the valine zipper region on the structure and aggregation of the basic leucine zipper (bZIP) domain of activating transcription factor 5 (ATF5).

    Science.gov (United States)

    Ciaccio, Natalie A; Reynolds, T Steele; Middaugh, C Russell; Laurence, Jennifer S

    2012-11-01

    Protein aggregation is a major problem for biopharmaceuticals. While the control of aggregation is critically important for the future of protein pharmaceuticals, mechanisms of aggregate assembly, particularly the role that structure plays, are still poorly understood. Increasing evidence indicates that partially folded intermediates critically influence the aggregation pathway. We have previously reported the use of the basic leucine zipper (bZIP) domain of activating transcription factor 5 (ATF5) as a partially folded model system to investigate protein aggregation. This domain contains three regions with differing structural propensity: a N-terminal polybasic region, a central helical leucine zipper region, and a C-terminal extended valine zipper region. Additionally, a centrally positioned cysteine residue readily forms an intermolecular disulfide bond that reduces aggregation. Computational analysis of ATF5 predicts that the valine zipper region facilitates self-association. Here we test this hypothesis using a truncated mutant lacking the C-terminal valine zipper region. We compare the structure and aggregation of this mutant to the wild-type (WT) form under both reducing and nonreducing conditions. Our data indicate that removal of this region results in a loss of α-helical structure in the leucine zipper and a change in the mechanism of self-association. The mutant form displays increased association at low temperature but improved resistance to thermally induced aggregation.

  15. Hair dyes resorcinol and lawsone reduce production of melanin in melanoma cells by tyrosinase activity inhibition and decreasing tyrosinase and microphthalmia-associated transcription factor (MITF) expression.

    Science.gov (United States)

    Lee, Shu-Mei; Chen, Yi-Shyan; Lin, Chih-Chien; Chen, Kuan-Hung

    2015-01-09

    Hair coloring products are one of the most important cosmetics for modern people; there are three major types of hair dyes, including the temporary, semi-permanent and permanent hair dyes. The selected hair dyes (such as ammonium persulfate, sodium persulfate, resorcinol and lawsone) are the important components for hair coloring products. Therefore, we analyzed the effects of these compounds on melanogenesis in B16-F10 melanoma cells. The results proved that hair dyes resorcinol and lawsone can reduce the production of melanin. The results also confirmed that resorcinol and lawsone inhibit mushroom and cellular tyrosinase activities in vitro. Resorcinol and lawsone can also downregulate the protein levels of tyrosinase and microphthalmia-associated transcription factor (MITF) in B16-F10 cells. Thus, we suggest that frequent use of hair dyes may have the risk of reducing natural melanin production in hair follicles. Moreover, resorcinol and lawsone may also be used as hypopigmenting agents to food, agricultural and cosmetic industry in the future.

  16. Transcription Factor Activating Protein-2β (TFAP-2β) genotype and symptoms of attention deficit hyperactivity disorder in relation to symptoms of depression in two independent samples.

    Science.gov (United States)

    Nilsson, Kent W; Sonnby, Karin; Nordquist, Niklas; Comasco, Erika; Leppert, Jerzy; Oreland, Lars; Sjöberg, Rickard L

    2014-04-01

    The Transcription Factor Activating Protein-2β (TFAP-2β) gene has been shown to influence monoaminergic neurotransmission, and several genes important for monoaminergic function have binding sites for TFAP-2β. Familial studies of attention deficit hyperactivity disorder (ADHD) suggest a hereditary-determined subtype of ADHD with comorbid depression. We examined a functional variation of the TFAP-2β gene in the context of co-occurring symptoms of ADHD and depression in two independent population-based samples of adolescents (Group A, n = 175 and Group B, n = 1,506) from Sweden. Results indicated 6.1 to 7.8% of adolescents screened positively for ADHD and depression symptoms. Symptoms of depression were more common among girls who screened positively for ADHD and did not carry the nine-repeat allele of the TFAP-2β intron 1 Variable Number Tandem Repeat (VNTR) polymorphism. The presence of the nine-repeat variant of the TFAP-2β intron 1 VNTR appears to protect girls with ADHD symptoms from the co-expression of symptoms of depression.

  17. Hair Dyes Resorcinol and Lawsone Reduce Production of Melanin in Melanoma Cells by Tyrosinase Activity Inhibition and Decreasing Tyrosinase and Microphthalmia-Associated Transcription Factor (MITF Expression

    Directory of Open Access Journals (Sweden)

    Shu-Mei Lee

    2015-01-01

    Full Text Available Hair coloring products are one of the most important cosmetics for modern people; there are three major types of hair dyes, including the temporary, semi-permanent and permanent hair dyes. The selected hair dyes (such as ammonium persulfate, sodium persulfate, resorcinol and lawsone are the important components for hair coloring products. Therefore, we analyzed the effects of these compounds on melanogenesis in B16-F10 melanoma cells. The results proved that hair dyes resorcinol and lawsone can reduce the production of melanin. The results also confirmed that resorcinol and lawsone inhibit mushroom and cellular tyrosinase activities in vitro. Resorcinol and lawsone can also downregulate the protein levels of tyrosinase and microphthalmia-associated transcription factor (MITF in B16-F10 cells. Thus, we suggest that frequent use of hair dyes may have the risk of reducing natural melanin production in hair follicles. Moreover, resorcinol and lawsone may also be used as hypopigmenting agents to food, agricultural and cosmetic industry in the future.

  18. The nuclear-encoded sigma factor SIG4 directly activates transcription of chloroplast psbA and ycf17 genes in the unicellular red alga Cyanidioschyzon merolae.

    Science.gov (United States)

    Fujii, Gaku; Imamura, Sousuke; Era, Atsuko; Miyagishima, Shin-ya; Hanaoka, Mitsumasa; Tanaka, Kan

    2015-05-01

    The plant organelle chloroplast originated from the endosymbiosis of a cyanobacterial-like photosynthetic bacterium, and still retains its own genome derived from this ancestor. We have been focusing on a unicellular red alga, Cyanidioschyzon merolae, as a model photosynthetic eukaryote. In this study, we analyzed the transcriptional specificity of SIG4, which is one of four nuclear-encoded chloroplast RNA polymerase sigma factors in this alga. Accumulation of the SIG4 protein was observed in response to nitrogen depletion or high light conditions. By comparing the chloroplast transcriptomes under nitrogen depletion and SIG4-overexpressing conditions, we identified several candidate genes as SIG4 targets. Together with the results of chromatin immunoprecipitation analysis, the promoters of the psbA (encoding the D1 protein of the photosystem II reaction center) and ycf17 (encoding a protein of the early light-inducible protein family) genes were shown to be direct activation targets. The phycobilisome (PBS) CpcB protein was decreased by SIG4 overexpression, which suggests the negative involvement of SIG4 in PBS accumulation. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  19. Myxoma virus attenuates expression of activating transcription factor 4 (ATF4 which has implications for the treatment of proteasome inhibitor–resistant multiple myeloma

    Directory of Open Access Journals (Sweden)

    Dunlap KM

    2015-01-01

    Full Text Available Katherine M Dunlap, Mee Y Bartee, Eric Bartee Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, SC, USA Abstract: The recent development of chemotherapeutic proteasome inhibitors, such as bortezomib, has improved the outcomes of patients suffering from the plasma cell malignancy multiple myeloma. Unfortunately, many patients treated with these drugs still suffer relapsing disease due to treatment-induced upregulation of the antiapoptotic protein Mcl1. We have recently demonstrated that an oncolytic poxvirus, known as myxoma, can rapidly eliminate primary myeloma cells by inducing cellular apoptosis. The efficacy of myxoma treatment on proteasome inhibitor–relapsed or –refractory myeloma, however, remains unknown. We now demonstrate that myxoma-based elimination of myeloma is not affected by cellular resistance to proteasome inhibitors. Additionally, myxoma virus infection specifically prevents expression of Mcl1 following induction of the unfolded protein response, by blocking translation of the unfolded protein response activating transcription factor (ATF4. These results suggest that myxoma-based oncolytic therapy represents an attractive option for myeloma patients whose disease is refractory to chemotherapeutic proteasome inhibitors due to upregulation of Mcl1. Keywords: drug resistance, oncolytic

  20. Mitochondrial nucleoid and transcription factor A.

    Science.gov (United States)

    Kanki, Tomotake; Nakayama, Hiroshi; Sasaki, Narie; Takio, Koji; Alam, Tanfis Istiaq; Hamasaki, Naotaka; Kang, Dongchon

    2004-04-01

    Nuclear DNA is tightly packed into nucleosomal structure. In contrast, human mitochondrial DNA (mtDNA) had long been believed to be rather naked because mitochondria lack histone. Mitochondrial transcription factor A (TFAM), a member of a high mobility group (HMG) protein family and a first-identified mitochondrial transcription factor, is essential for maintenance of mitochondrial DNA. Abf2, a yeast counterpart of human TFAM, is abundant enough to cover the whole region of mtDNA and to play a histone-like role in mitochondria. Human TFAM is indeed as abundant as Abf2, suggesting that TFAM also has a histone-like architectural role for maintenance of mtDNA. When human mitochondria are solubilized with non-ionic detergent Nonidet-P40 and then separated into soluble and particulate fractions, most TFAM is recovered from the particulate fraction together with mtDNA, suggesting that human mtDNA forms a nucleoid structure. TFAM is tightly associated with mtDNA as a main component of the nucleoid.

  1. 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.

  2. Transcription factors mediate long-range enhancer-promoter interactions.

    Science.gov (United States)

    Nolis, Ilias K; McKay, Daniel J; Mantouvalou, Eva; Lomvardas, Stavros; Merika, Menie; Thanos, Dimitris

    2009-12-01

    We examined how remote enhancers establish physical communication with target promoters to activate gene transcription in response to environmental signals. Although the natural IFN-beta enhancer is located immediately upstream of the core promoter, it also can function as a classical enhancer element conferring virus infection-dependent activation of heterologous promoters, even when it is placed several kilobases away from these promoters. We demonstrated that the remote IFN-beta enhancer "loops out" the intervening DNA to reach the target promoter. These chromatin loops depend on sequence-specific transcription factors bound to the enhancer and the promoter and thus can explain the specificity observed in enhancer-promoter interactions, especially in complex genetic loci. Transcription factor binding sites scattered between an enhancer and a promoter can work as decoys trapping the enhancer in nonproductive loops, thus resembling insulator elements. Finally, replacement of the transcription factor binding sites involved in DNA looping with those of a heterologous prokaryotic protein, the lambda repressor, which is capable of loop formation, rescues enhancer function from a distance by re-establishing enhancer-promoter loop formation.

  3. 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.

  4. 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.

  5. Identification and Small Molecule Inhibition of an Activating Transcription Factor 4 (ATF4)-dependent Pathway to Age-related Skeletal Muscle Weakness and Atrophy*

    Science.gov (United States)

    Ebert, Scott M.; Dyle, Michael C.; Bullard, Steven A.; Dierdorff, Jason M.; Murry, Daryl J.; Fox, Daniel K.; Bongers, Kale S.; Lira, Vitor A.; Meyerholz, David K.; Talley, John J.; Adams, Christopher M.

    2015-01-01

    Aging reduces skeletal muscle mass and strength, but the underlying molecular mechanisms remain elusive. Here, we used mouse models to investigate molecular mechanisms of age-related skeletal muscle weakness and atrophy as well as new potential interventions for these conditions. We identified two small molecules that significantly reduce age-related deficits in skeletal muscle strength, quality, and mass: ursolic acid (a pentacyclic triterpenoid found in apples) and tomatidine (a steroidal alkaloid derived from green tomatoes). Because small molecule inhibitors can sometimes provide mechanistic insight into disease processes, we used ursolic acid and tomatidine to investigate the pathogenesis of age-related muscle weakness and atrophy. We found that ursolic acid and tomatidine generate hundreds of small positive and negative changes in mRNA levels in aged skeletal muscle, and the mRNA expression signatures of the two compounds are remarkably similar. Interestingly, a subset of the mRNAs repressed by ursolic acid and tomatidine in aged muscle are positively regulated by activating transcription factor 4 (ATF4). Based on this finding, we investigated ATF4 as a potential mediator of age-related muscle weakness and atrophy. We found that a targeted reduction in skeletal muscle ATF4 expression reduces age-related deficits in skeletal muscle strength, quality, and mass, similar to ursolic acid and tomatidine. These results elucidate ATF4 as a critical mediator of age-related muscle weakness and atrophy. In addition, these results identify ursolic acid and tomatidine as potential agents and/or lead compounds for reducing ATF4 activity, weakness, and atrophy in aged skeletal muscle. PMID:26338703

  6. The chloroplast transcription apparatus from mustard (Sinapis alba L.). Evidence for three different transcription factors which resemble bacterial sigma factors.

    Science.gov (United States)

    Tiller, K; Eisermann, A; Link, G

    1991-05-23

    A chloroplast protein fraction with sigma-like activity [Bülow, S. & Link, G. (1988) Plant Mol. Biol. 10, 349-357], was further purified and characterized. Chromatography on heparin-Sepharose, DEAE-Sepharose and Sephacryl S-300 led to the separation of three sigma-like factors (SLF) polypeptides with Mr 67,000 (SLF67), 52,000 (SLF52) and 29,000 (SLF29). None of these polypeptides bind to DNA itself, but each one confers enhanced binding and transcriptional activity when added to Escherichia coli RNA-polymerase core enzyme and DNA fragments carrying a chloroplast promoter. SLF67, SLF52, and SLF29 differ in their ionic-strength requirements for activity. They each mediate the binding to promoters of the chloroplast genes psbA, trnQ, and rps16, with different efficiencies. It is suggested that chloroplast transcription in vivo might be controlled at least in part by these functionally distinct factors.

  7. The ubiquitous transcription factor CTCF promotes lineage-specific epigenomic remodeling and establishment of transcriptional networks driving cell differentiation.

    Science.gov (United States)

    Dubois-Chevalier, Julie; Staels, Bart; Lefebvre, Philippe; Eeckhoute, Jérôme

    2015-01-01

    Cell differentiation relies on tissue-specific transcription factors (TFs) that cooperate to establish unique transcriptomes and phenotypes. However, the role of ubiquitous TFs in these processes remains poorly defined. Recently, we have shown that the CCCTC-binding factor (CTCF) is required for adipocyte differentiation through epigenomic remodelling of adipose tissue-specific enhancers and transcriptional activation of Peroxisome proliferator-activated receptor gamma (PPARG), the main driver of the adipogenic program (PPARG), and its target genes. Here, we discuss how these findings, together with the recent literature, illuminate a functional role for ubiquitous TFs in lineage-determining transcriptional networks.

  8. Determination and inference of eukaryotic transcription factor sequence specificity.

    Science.gov (United States)

    Weirauch, Matthew T; Yang, Ally; Albu, Mihai; Cote, Atina G; Montenegro-Montero, Alejandro; Drewe, Philipp; Najafabadi, Hamed S; Lambert, Samuel A; Mann, Ishminder; Cook, Kate; Zheng, Hong; Goity, Alejandra; van Bakel, Harm; Lozano, Jean-Claude; Galli, Mary; Lewsey, Mathew G; Huang, Eryong; Mukherjee, Tuhin; Chen, Xiaoting; Reece-Hoyes, John S; Govindarajan, Sridhar; Shaulsky, Gad; Walhout, Albertha J M; Bouget, François-Yves; Ratsch, Gunnar; Larrondo, Luis F; Ecker, Joseph R; Hughes, Timothy R

    2014-09-11

    Transcription factor (TF) DNA sequence preferences direct their regulatory activity, but are currently known for only ∼1% of eukaryotic TFs. Broadly sampling DNA-binding domain (DBD) types from multiple eukaryotic clades, we determined DNA sequence preferences for >1,000 TFs encompassing 54 different DBD classes from 131 diverse eukaryotes. We find that closely related DBDs almost always have very similar DNA sequence preferences, enabling inference of motifs for ∼34% of the ∼170,000 known or predicted eukaryotic TFs. Sequences matching both measured and inferred motifs are enriched in chromatin immunoprecipitation sequencing (ChIP-seq) peaks and upstream of transcription start sites in diverse eukaryotic lineages. SNPs defining expression quantitative trait loci in Arabidopsis promoters are also enriched for predicted TF binding sites. Importantly, our motif "library" can be used to identify specific TFs whose binding may be altered by human disease risk alleles. These data present a powerful resource for mapping transcriptional networks across eukaryotes.

  9. Human RNA polymerase II associated factor 1 complex promotes tumorigenesis by activating c-MYC transcription in non-small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Zhi, Xiuyi [Department of Thoracic Surgery, Xuanwu Hospital, Capital Medical University, Beijing, 100053 (China); Giroux-Leprieur, Etienne [ER2 GRC UPMC04 Theranoscan, Pierre et Marie Curie University, Tenon Hospital, 4 Rue de La Chine, 75020, Paris (France); Respiratory Diseases and Thoracic Oncology Department, Ambroise Pare Hospital – APHP, Versailles Saint Quentin en Yvelines University, 9 Avenue Charles de Gaulle, 92100, Boulogne-Billancourt (France); Wislez, Marie [ER2 GRC UPMC04 Theranoscan, Pierre et Marie Curie University, Tenon Hospital, 4 Rue de La Chine, 75020, Paris (France); Hu, Mu; Zhang, Yi [Department of Thoracic Surgery, Xuanwu Hospital, Capital Medical University, Beijing, 100053 (China); Shi, Huaiyin [Department of Pathology, Chinese PLA General Hospital, Fu-xing Road #28, Beijing, 100853 (China); Du, Kaiqi, E-mail: kaiqidu_zhejiang@163.com [Department of Cardiothoracic Surgery, Chinese People' s Armed Police Force, Zhejiang Corps Hospital, Jiaxing, Zhejiang Province (China); Wang, Lei, E-mail: leiwang_hebei@163.com [Department of Human Anatomy, Hebei Medical University, Hebei Province (China)

    2015-10-02

    Human RNA polymerase II (RNAPII)-associated factor 1 complex (hPAF1C) plays a crucial role in protein-coding gene transcription. Overexpression of hPAF1C has been implicated in the initiation and progression of various human cancers. However, the molecular pathways involved in tumorigenesis through hPAF1C remain to be elucidated. The current study suggested hPAF1C expression as a prognostic biomarker for early stage non-small cell lung cancer (NSCLC) and patients with low hPAF1C expression levels had significantly better overall survival. Furthermore, the expression of hPAF1C was found to be positively correlated with c-MYC expression in patient tumor samples and in cancer cell lines. Mechanistic studies indicated that hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription. These results demonstrated the prognostic value of hPAF1C in early-stage NSCLC and the role of hPAF1C in the transcriptional regulation of c-MYC oncogene during NSCLC tumorigenesis. - Highlights: • hPAF1C expression is a prognostic biomarker for early stage non-small cell lung cancer. • The expression of hPAF1C was positively correlated with c-MYC in tumor samples of patients and in several NSCLC cell lines. • hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription.

  10. Mapping functional regions of transcription factor TFIIIA.

    Science.gov (United States)

    Vrana, K E; Churchill, M E; Tullius, T D; Brown, D D

    1988-04-01

    Functional deletion mutants of the trans-acting factor TFIIIA, truncated at both ends of the molecule, have been expressed by in vitro transcription of a cDNA clone and subsequent cell-free translation of the synthetic mRNAs. A region of TFIIIA 19 amino acids or less, near the carboxyl terminus, is critical for maximal transcription and lies outside the DNA-binding domain. The elongated protein can be aligned over the internal control region (ICR) of the Xenopus 5S RNA gene with its carboxyl terminus oriented toward the 5' end of the gene and its amino terminus oriented toward the 3' end of the gene. The nine "zinc fingers" and the linkers that separate them comprise 80% of the protein mass and correspond to the DNA-binding domain of TFIIIA. The zinc fingers near the amino terminus of the protein contribute more to the overall binding energy of the protein to the ICR than do the zinc fingers near the carboxyl end. The most striking feature of TFIIIA is its modular structure. This is demonstrated by the fact that each zinc finger binds to just one of three short nucleotide sequences within the ICR.

  11. Transcription Factor Families Regulate the Anthocyanin Biosynthetic Pathway in Capsicum

    Science.gov (United States)

    Anthocyanin structural gene transcription requires the expression of at least one member of each of three transcription factor families - MYC, MYB and WD40. These transcription factors form a complex that binds to structural gene promoters, thereby modulating gene expression. Capsicum annuum display...

  12. Characterization of the MADS domain transcription factor family in Arabidopsis thaliana

    NARCIS (Netherlands)

    Folter, Stefan de

    2006-01-01

    Gene regulation at the level of transcription is crucial for almost all biological processes in a cell or organism. Transcription factors are sequence-specific DNA-binding proteins that are capable of activating and/or repressing transcription. The genome of Arabidopsis thaliana, for instance, consi

  13. Bayesian non-negative factor analysis for reconstructing transcription factor mediated regulatory networks

    Directory of Open Access Journals (Sweden)

    Chen Yidong

    2011-10-01

    Full Text Available Abstract Background Transcriptional regulation by transcription factor (TF controls the time and abundance of mRNA transcription. Due to the limitation of current proteomics technologies, large scale measurements of protein level activities of TFs is usually infeasible, making computational reconstruction of transcriptional regulatory network a difficult task. Results We proposed here a novel Bayesian non-negative factor model for TF mediated regulatory networks. Particularly, the non-negative TF activities and sample clustering effect are modeled as the factors from a Dirichlet process mixture of rectified Gaussian distributions, and the sparse regulatory coefficients are modeled as the loadings from a sparse distribution that constrains its sparsity using knowledge from database; meantime, a Gibbs sampling solution was developed to infer the underlying network structure and the unknown TF activities simultaneously. The developed approach has been applied to simulated system and breast cancer gene expression data. Result shows that, the proposed method was able to systematically uncover TF mediated transcriptional regulatory network structure, the regulatory coefficients, the TF protein level activities and the sample clustering effect. The regulation target prediction result is highly coordinated with the prior knowledge, and sample clustering result shows superior performance over previous molecular based clustering method. Conclusions The results demonstrated the validity and effectiveness of the proposed approach in reconstructing transcriptional networks mediated by TFs through simulated systems and real data.

  14. 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

  15. 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.

  16. Oncovirus Kaposi sarcoma herpesvirus (KSHV) represses tumor suppressor PDLIM2 to persistently activate nuclear factor κB (NF-κB) and STAT3 transcription factors for tumorigenesis and tumor maintenance.

    Science.gov (United States)

    Sun, Fan; Xiao, Yadong; Qu, Zhaoxia

    2015-03-20

    Kaposi sarcoma herpesvirus (KSHV) is the most common cause of malignancies among AIDS patients. However, how KSHV induces tumorigenesis remains largely unknown. Here, we demonstrate that one important mechanism underlying the tumorigenesis of KSHV is through transcriptional repression of the tumor suppressor gene PDZ-LIM domain-containing protein 2 (PDLIM2). PDLIM2 expression is repressed in KSHV-transformed human umbilical vascular endothelial cells as well as in KSHV-associated cancer cell lines and primary tumors. Importantly, PDLIM2 repression is essential for KSHV-induced persistent activation of nuclear factor κB (NF-κB) and signal transducer and activator of transcription 3 (STAT3) and subsequent tumorigenesis and tumor maintenance. Our mechanistic studies indicate that PDLIM2 repression by KSHV involves DNA methylation. Notably, the epigenetic repression of PDLIM2 can be reversed by 5-aza-2-deoxycytidine and vitamin D to suppress KSHV-associated cancer cell growth. These studies not only improve our understanding of KSHV pathogenesis but also provide immediate therapeutic strategies for KSHV-mediated cancers, particularly those associated with AIDS.

  17. Insect neuropeptide bursicon homodimers induce innate immune and stress genes during molting by activating the NF-κB transcription factor Relish.

    Directory of Open Access Journals (Sweden)

    Shiheng An

    Full Text Available BACKGROUND: Bursicon is a heterodimer neuropeptide composed of two cystine knot proteins, bursicon α (burs α and bursicon β (burs β, that elicits cuticle tanning (melanization and sclerotization through the Drosophila leucine-rich repeats-containing G protein-coupled receptor 2 (DLGR2. Recent studies show that both bursicon subunits also form homodimers. However, biological functions of the homodimers have remained unknown until now. METHODOLOGY/PRINCIPAL FINDINGS: In this report, we show in Drosophila melanogaster that both bursicon homodimers induced expression of genes encoding antimicrobial peptides (AMPs in neck-ligated adults following recombinant homodimer injection and in larvae fat body after incubation with recombinant homodimers. These AMP genes were also up-regulated in 24 h old unligated flies (when the endogenous bursicon level is low after injection of recombinant homodimers. Up-regulation of AMP genes by the homodimers was accompanied by reduced bacterial populations in fly assay preparations. The induction of AMP expression is via activation of the NF-κB transcription factor Relish in the immune deficiency (Imd pathway. The influence of bursicon homodimers on immune function does not appear to act through the heterodimer receptor DLGR2, i.e. novel receptors exist for the homodimers. CONCLUSIONS/SIGNIFICANCE: Our results reveal a mechanism of CNS-regulated prophylactic innate immunity during molting via induced expression of genes encoding AMPs and genes of the Turandot family. Turandot genes are also up-regulated by a broader range of extreme insults. From these data we infer that CNS-generated bursicon homodimers mediate innate prophylactic immunity to both stress and infection during the vulnerable molting cycle.

  18. Phosphoproteome and transcription factor activity profiling identify actions of the anti-inflammatory agent UTL-5g in LPS stimulated RAW 264.7 cells including disrupting actin remodeling and STAT-3 activation.

    Science.gov (United States)

    Carruthers, Nicholas J; Stemmer, Paul M; Chen, Ben; Valeriote, Frederick; Gao, Xiaohua; Guatam, Subhash C; Shaw, Jiajiu

    2017-09-15

    UTL-5g is a novel small-molecule TNF-alpha modulator. It reduces cisplatin-induced side effects by protecting kidney, liver, and platelets, thereby increasing tolerance for cisplatin. UTL-5g also reduces radiation-induced acute liver toxicity. The mechanism of action for UTL-5g is not clear at the present time. A phosphoproteomic analysis to a depth of 4943 phosphopeptides and a luminescence-based transcription factor activity assay were used to provide complementary analyses of signaling events that were disrupted by UTL-5g in RAW 264.7 cells. Transcriptional activity downstream of the interferon gamma, IL-6, type 1 Interferon, TGF-β, PKC/Ca(2+) and the glucocorticoid receptor pathways were disrupted by UTL-5g. Phosphoproteomic analysis indicated that hyperphosphorylation of proteins involved in actin remodeling was suppressed by UTL-5g (gene set analysis, FDR 5g. This global characterization of UTL-5g activity in a macrophage cell line discovered that it disrupts selected aspects of LPS signaling including Stat3 activation and actin remodeling providing new insight on how UTL-5g acts to reduce cisplatin-induced side effects. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Transcription profile of Escherichia coli: genomic SELEX search for regulatory targets of transcription factors.

    Science.gov (United States)

    Ishihama, Akira; Shimada, Tomohiro; Yamazaki, Yukiko

    2016-03-18

    Bacterial genomes are transcribed by DNA-dependent RNA polymerase (RNAP), which achieves gene selectivity through interaction with sigma factors that recognize promoters, and transcription factors (TFs) that control the activity and specificity of RNAP holoenzyme. To understand the molecular mechanisms of transcriptional regulation, the identification of regulatory targets is needed for all these factors. We then performed genomic SELEX screenings of targets under the control of each sigma factor and each TF. Here we describe the assembly of 156 SELEX patterns of a total of 116 TFs performed in the presence and absence of effector ligands. The results reveal several novel concepts: (i) each TF regulates more targets than hitherto recognized; (ii) each promoter is regulated by more TFs than hitherto recognized; and (iii) the binding sites of some TFs are located within operons and even inside open reading frames. The binding sites of a set of global regulators, including cAMP receptor protein, LeuO and Lrp, overlap with those of the silencer H-NS, suggesting that certain global regulators play an anti-silencing role. To facilitate sharing of these accumulated SELEX datasets with the research community, we compiled a database, 'Transcription Profile of Escherichia coli' (www.shigen.nig.ac.jp/ecoli/tec/). © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  20. 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.

  1. Aldosterone stimulates nuclear factor-kappa B activity and transcription of intercellular adhesion molecule-1 and connective tissue growth factor in rat mesangial cells via serum- and glucocorticoid-inducible protein kinase-1.

    Science.gov (United States)

    Terada, Yoshio; Ueda, Satoko; Hamada, Kazu; Shimamura, Yoshiko; Ogata, Koji; Inoue, Kosuke; Taniguchi, Yoshinori; Kagawa, Toru; Horino, Taro; Takao, Toshihiro

    2012-02-01

    Several clinical and experimental data support the hypothesis that aldosterone contributes to the progression of renal injury. To determine the signaling pathway of aldosterone in relation to fibrosis and inflammation in mesangial cells, we investigated the effects of aldosterone on expression and activation of serum- and glucocorticoid-inducible protein kinase-1 (SGK1), the activation of nuclear factor-kappa B (NF-κB activation, and the expressions of intercellular adhesion molecule-1 (ICAM-1) and connective tissue growth factor (CTGF). Aldosterone stimulated SGK1 expression, phosphorylation (Ser-256), and kinase activity. The increments of phosphorylation and expression of SGK1 induced by aldosterone were inhibited by mineralocorticoid receptor (MR) inhibitor (eplerenone). Aldosterone stimulated NF-κB activity measured by NF-κB responsive elements, luciferase assay, and the levels of inhibitor of kappa B (IκB) phosphorylation. This aldosterone-induced activation of NF-κB was inhibited by the transfection of dominant-negative SGK1. Furthermore, aldosterone augmented the promoter activities and protein expressions of ICAM-1 and CTGF. The effects of aldosterone on ICAM-1 and CTGF promoter activities and protein expressions were inhibited by the transfection of dominant-negative SGK1 and dominant-negative IκBα. We also found that the MR antagonist significantly ameliorated the glomerular injury and enhancements in SGK1, ICAM-1, and CTGF expressions induced by 1% sodium chloride and aldosterone in vivo. In conclusion, our findings suggest that aldosterone stimulates ICAM-1 and CTGF transcription via activation of SGK1 and NF-κB, which may be involved in the progression of aldosterone-induced mesangial fibrosis and inflammation. MR antagonists may serve as useful therapeutic targets for the treatment of glomerular inflammatory disease.

  2. Inhibition of Nuclear Transcription Factor-κB and Activation of Peroxisome Proliferator-Activated Receptors in HepG2 Cells by Cucurbitane-Type Triterpene Glycosides from Momordica charantia

    Science.gov (United States)

    Nhiem, Nguyen Xuan; Yen, Pham Hai; Ngan, Nguyen Thi Thanh; Quang, Tran Hong; Kiem, Phan Van; Minh, Chau Van; Tai, Bui Huu; Cuong, Nguyen Xuan; Song, Seok Bean

    2012-01-01

    Abstract Momordica charantia: is used to treat various diseases, including inflammatory conditions. Previous reports indicated that the extract of this plant inhibits activation of nuclear transcription factor-κB (NF-κB) but activates peroxisome proliferator-activated receptor (PPAR). Additionally, cucurbitane-type triterpene glycosides are the main bioactive components of the fruit of M. charantia. Therefore, we investigated the anti-inflammatory activity of 17 cucurbitane-type triterpene glycosides (1–17) isolated from this plant. Their inhibition of NF-κB and activation of PPAR activities in HepG2 cells were measured using luciferase reporter and PPAR subtype transactivation assays. Compounds 6 and 8 were found to inhibit NF-κB activation stimulated by tumor necrosis factor-α (TNFα) in a dose-dependent manner. With 50% inhibition concentration (IC50) values of 0.4 μM, compounds 6 and 8 were more potent inhibitors than the positive control, sulfasalazine (IC50=0.9 μM). Compounds 4, 6, and 8 also inhibited TNFα-induced expressions of inducible nitric oxide synthase and cyclooxygenase-2 mRNA. However, only compound 13 significantly increased PPARγ transactivation. PMID:22248180

  3. Inhibition of nuclear transcription factor-κB and activation of peroxisome proliferator-activated receptors in HepG2 cells by cucurbitane-type triterpene glycosides from Momordica charantia.

    Science.gov (United States)

    Nhiem, Nguyen Xuan; Yen, Pham Hai; Ngan, Nguyen Thi Thanh; Quang, Tran Hong; Kiem, Phan Van; Minh, Chau Van; Tai, Bui Huu; Cuong, Nguyen Xuan; Song, Seok Bean; Kim, Young Ho

    2012-04-01

    Momordica charantia is used to treat various diseases, including inflammatory conditions. Previous reports indicated that the extract of this plant inhibits activation of nuclear transcription factor-κB (NF-κB) but activates peroxisome proliferator-activated receptor (PPAR). Additionally, cucurbitane-type triterpene glycosides are the main bioactive components of the fruit of M. charantia. Therefore, we investigated the anti-inflammatory activity of 17 cucurbitane-type triterpene glycosides (1-17) isolated from this plant. Their inhibition of NF-κB and activation of PPAR activities in HepG2 cells were measured using luciferase reporter and PPAR subtype transactivation assays. Compounds 6 and 8 were found to inhibit NF-κB activation stimulated by tumor necrosis factor-α (TNFα) in a dose-dependent manner. With 50% inhibition concentration (IC(50)) values of 0.4 μM, compounds 6 and 8 were more potent inhibitors than the positive control, sulfasalazine (IC(50)=0.9 μM). Compounds 4, 6, and 8 also inhibited TNFα-induced expressions of inducible nitric oxide synthase and cyclooxygenase-2 mRNA. However, only compound 13 significantly increased PPARγ transactivation.

  4. 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...

  5. Absence of the Birt-Hogg-Dubé gene product is associated with increased hypoxia-inducible factor transcriptional activity and a loss of metabolic flexibility.

    Science.gov (United States)

    Preston, R S; Philp, A; Claessens, T; Gijezen, L; Dydensborg, A B; Dunlop, E A; Harper, K T; Brinkhuizen, T; Menko, F H; Davies, D M; Land, S C; Pause, A; Baar, K; van Steensel, M A M; Tee, A R

    2011-03-10

    Under conditions of reduced tissue oxygenation, hypoxia-inducible factor (HIF) controls many processes, including angiogenesis and cellular metabolism, and also influences cell proliferation and survival decisions. HIF is centrally involved in tumour growth in inherited diseases that give rise to renal cell carcinoma (RCC), such as Von Hippel-Lindau syndrome and tuberous sclerosis complex. In this study, we examined whether HIF is involved in tumour formation of RCC in Birt-Hogg-Dubé syndrome. For this, we analysed a Birt-Hogg-Dubé patient-derived renal tumour cell line (UOK257) that is devoid of the Birt-Hogg-Dubé protein (BHD) and observed high levels of HIF activity. Knockdown of BHD expression also caused a threefold activation of HIF, which was not as a consequence of more HIF1α or HIF2α protein. Transcription of HIF target genes VEGF, BNIP3 and CCND1 was also increased. We found nuclear localization of HIF1α and increased expression of VEGF, BNIP3 and GLUT1 in a chromophobe carcinoma from a Birt-Hogg-Dubé patient. Our data also reveal that UOK257 cells have high lactate dehydrogenase, pyruvate kinase and 3-hydroxyacyl-CoA dehydrogenase activity. We observed increased expression of pyruvate dehydrogenase kinase 1 (a HIF gene target), which in turn leads to increased phosphorylation and inhibition of pyruvate dehydrogenase. Together with increased protein levels of GLUT1, our data reveal that UOK257 cells favour glycolytic rather than lipid metabolism (a cancer phenomenon termed the 'Warburg effect'). UOK257 cells also possessed a higher expression level of the L-lactate influx monocarboxylate transporter 1 and consequently utilized L-lactate as a metabolic fuel. As a result of their higher dependency on glycolysis, we were able to selectively inhibit the growth of these UOK257 cells by treatment with 2-deoxyglucose. This work suggests that targeting glycolytic metabolism may be used therapeutically to treat Birt-Hogg-Dubé-associated renal lesions.

  6. Effects of cytosine methylation on transcription factor binding sites

    KAUST Repository

    Medvedeva, Yulia A

    2014-03-26

    Background: DNA methylation in promoters is closely linked to downstream gene repression. However, whether DNA methylation is a cause or a consequence of gene repression remains an open question. If it is a cause, then DNA methylation may affect the affinity of transcription factors (TFs) for their binding sites (TFBSs). If it is a consequence, then gene repression caused by chromatin modification may be stabilized by DNA methylation. Until now, these two possibilities have been supported only by non-systematic evidence and they have not been tested on a wide range of TFs. An average promoter methylation is usually used in studies, whereas recent results suggested that methylation of individual cytosines can also be important.Results: We found that the methylation profiles of 16.6% of cytosines and the expression profiles of neighboring transcriptional start sites (TSSs) were significantly negatively correlated. We called the CpGs corresponding to such cytosines " traffic lights" We observed a strong selection against CpG " traffic lights" within TFBSs. The negative selection was stronger for transcriptional repressors as compared with transcriptional activators or multifunctional TFs as well as for core TFBS positions as compared with flanking TFBS positions.Conclusions: Our results indicate that direct and selective methylation of certain TFBS that prevents TF binding is restricted to special cases and cannot be considered as a general regulatory mechanism of transcription. 2013 Medvedeva et al.; licensee BioMed Central Ltd.

  7. Functional characterization of tomato membrane-bound NAC transcription factors.

    Science.gov (United States)

    Bhattacharjee, Payel; Das, Rohit; Mandal, Arunava; Kundu, Pallob

    2017-03-01

    Genome-wide analysis was carried out to identify and analyze differential expression pattern of tomato membrane bound NAC transcription factors (SlNACMTFs) during stresses. Two biotic-stress-related SlNACMTFs have been characterized to elucidate their regulatory function. NAC transcription factors are known regulators of stress-related gene expression. As Stresses are perceived and transmitted by membrane-bound proteins, functional characterization of membrane-associated NAC transcription factors in tomato can reveal valuable insight about membrane-mediated stress-signalling. Tomato genome encodes 13 NAC genes which have predicted transmembrane domain(s) (SlNACMTFs). mRNA of 12 SlNACMTFs were readily detected in multiple tissues, and also in polysome isolated from leaf, confirming active transcription and translation from these genes occur under normal physiological condition. Additionally, most of the SlNACMTFs were differentially regulated during stresses and stress-related transcription factor binding sites are prevalent in their promoters. SlNACMTF3 and 8 were majorly regulated in biotic and abiotic stresses. Like other MTFs, SlNACMTF3 was translocated to the plasma membrane, whereas the C-terminus truncated (ΔC) form localized in the cytoplasm and the nucleus. Accordingly, the ΔC forms significantly influenced the activity of promoters harbouring NAC binding sites (NACbs). Furthermore, the NAC domain of these transcription factors could directly interact with an NACbs, and the proteins failed to regulate a promoter lacking a crucial NACbs. Interestingly, the type of influence to an NACbs containing promoter was dependent on the context of the NACbs, as the same SlNACMTF showed an alternative mode of regulation on different promoters, as well as the same promoter activity was oppositely regulated by two different SlNACMTF. Finally, both SlNACMTFs demonstrated the differential regulatory effect on the expression of several stress-related genes by interacting

  8. Transcriptional interference by RNA polymerase pausing and dislodgement of transcription factors.

    Science.gov (United States)

    Palmer, Adam C; Egan, J Barry; Shearwin, Keith E

    2011-01-01

    Transcriptional interference is the in cis suppression of one transcriptional process by another. Mathematical modeling shows that promoter occlusion by elongating RNA polymerases cannot produce strong interference. Interference may instead be generated by (1) dislodgement of slow-to-assemble pre-initiation complexes and transcription factors and (2) prolonged occlusion by paused RNA polymerases.

  9. Ketamine inhibits transcription factors activator protein 1 and nuclear factor-kappaB, interleukin-8 production, as well as CD11b and CD16 expression: studies in human leukocytes and leukocytic cell lines.

    NARCIS (Netherlands)

    Welters, I.D.; Hafer, G.; Menzebach, A.; Muhling, J.; Neuhauser, C.; Browning, P.; Goumon, Y.

    2010-01-01

    BACKGROUND: Recent data indicate that ketamine exerts antiinflammatory actions. However, little is known about the signaling mechanisms involved in ketamine-induced immune modulation. In this study, we investigated the effects of ketamine on lipopolysaccharide-induced activation of transcription

  10. 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.

  11. 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.

  12. The solution configurations of inactive and activated DntR have implications for the sliding dimer mechanism of LysR transcription factors.

    Science.gov (United States)

    Lerche, Michael; Dian, Cyril; Round, Adam; Lönneborg, Rosa; Brzezinski, Peter; Leonard, Gordon A

    2016-01-28

    LysR Type Transcriptional Regulators (LTTRs) regulate basic metabolic pathways or virulence gene expression in prokaryotes. Evidence suggests that the activation of LTTRs involves a conformational change from an inactive compact apo- configuration that represses transcription to an active, expanded holo- form that promotes it. However, no LTTR has yet been observed to adopt both configurations. Here, we report the results of structural studies of various forms of the LTTR DntR. Crystal structures of apo-DntR and of a partially autoinducing mutant H169T-DntR suggest that active and inactive DntR maintain a compact homotetrameric configuration. However, Small Angle X-ray Scattering (SAXS) studies on solutions of apo-, H169T- and inducer-bound holo-DntR indicate a different behaviour, suggesting that while apo-DntR maintains a compact configuration in solution both H169T- and holo-DntR adopt an expanded conformation. Models of the SAXS-obtained solution conformations of apo- and holo-DntR homotetramers in complex with promoter-operator region DNA are consistent with previous observations of a shifting of LTTR DNA binding sites upon activation and a consequent relaxation in the bend of the promoter-operator region DNA. Our results thus provide clear evidence at the molecular level which strongly supports the 'sliding dimer' hypothesis concerning LTTR activation mechanisms.

  13. Pathologically Relevant Prelamin A Interactions with Transcription Factors.

    Science.gov (United States)

    Infante, Arantza; Rodríguez, Clara I

    2016-01-01

    LMNA-linked laminopathies are a group of rare human diseases caused by mutations in LMNA or by disrupted posttranslational processing of its largest encoded isoform, prelamin A. The accumulation of mutated or immature forms of farnesylated prelamin A, named progerin or prelamin A, respectively, dominantly disrupts nuclear lamina structure with toxic effects in cells. One hypothe