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Sample records for additional regulatory factors

  1. Additive functions in boolean models of gene regulatory network modules.

    PubMed

    Darabos, Christian; Di Cunto, Ferdinando; Tomassini, Marco; Moore, Jason H; Provero, Paolo; Giacobini, Mario

    2011-01-01

    Gene-on-gene regulations are key components of every living organism. Dynamical abstract models of genetic regulatory networks help explain the genome's evolvability and robustness. These properties can be attributed to the structural topology of the graph formed by genes, as vertices, and regulatory interactions, as edges. Moreover, the actual gene interaction of each gene is believed to play a key role in the stability of the structure. With advances in biology, some effort was deployed to develop update functions in boolean models that include recent knowledge. We combine real-life gene interaction networks with novel update functions in a boolean model. We use two sub-networks of biological organisms, the yeast cell-cycle and the mouse embryonic stem cell, as topological support for our system. On these structures, we substitute the original random update functions by a novel threshold-based dynamic function in which the promoting and repressing effect of each interaction is considered. We use a third real-life regulatory network, along with its inferred boolean update functions to validate the proposed update function. Results of this validation hint to increased biological plausibility of the threshold-based function. To investigate the dynamical behavior of this new model, we visualized the phase transition between order and chaos into the critical regime using Derrida plots. We complement the qualitative nature of Derrida plots with an alternative measure, the criticality distance, that also allows to discriminate between regimes in a quantitative way. Simulation on both real-life genetic regulatory networks show that there exists a set of parameters that allows the systems to operate in the critical region. This new model includes experimentally derived biological information and recent discoveries, which makes it potentially useful to guide experimental research. The update function confers additional realism to the model, while reducing the complexity

  2. Additive Functions in Boolean Models of Gene Regulatory Network Modules

    PubMed Central

    Darabos, Christian; Di Cunto, Ferdinando; Tomassini, Marco; Moore, Jason H.; Provero, Paolo; Giacobini, Mario

    2011-01-01

    Gene-on-gene regulations are key components of every living organism. Dynamical abstract models of genetic regulatory networks help explain the genome's evolvability and robustness. These properties can be attributed to the structural topology of the graph formed by genes, as vertices, and regulatory interactions, as edges. Moreover, the actual gene interaction of each gene is believed to play a key role in the stability of the structure. With advances in biology, some effort was deployed to develop update functions in Boolean models that include recent knowledge. We combine real-life gene interaction networks with novel update functions in a Boolean model. We use two sub-networks of biological organisms, the yeast cell-cycle and the mouse embryonic stem cell, as topological support for our system. On these structures, we substitute the original random update functions by a novel threshold-based dynamic function in which the promoting and repressing effect of each interaction is considered. We use a third real-life regulatory network, along with its inferred Boolean update functions to validate the proposed update function. Results of this validation hint to increased biological plausibility of the threshold-based function. To investigate the dynamical behavior of this new model, we visualized the phase transition between order and chaos into the critical regime using Derrida plots. We complement the qualitative nature of Derrida plots with an alternative measure, the criticality distance, that also allows to discriminate between regimes in a quantitative way. Simulation on both real-life genetic regulatory networks show that there exists a set of parameters that allows the systems to operate in the critical region. This new model includes experimentally derived biological information and recent discoveries, which makes it potentially useful to guide experimental research. The update function confers additional realism to the model, while reducing the complexity

  3. Transcription factor trapping by RNA in gene regulatory elements.

    PubMed

    Sigova, Alla A; Abraham, Brian J; Ji, Xiong; Molinie, Benoit; Hannett, Nancy M; Guo, Yang Eric; Jangi, Mohini; Giallourakis, Cosmas C; Sharp, Phillip A; Young, Richard A

    2015-11-20

    Transcription factors (TFs) bind specific sequences in promoter-proximal and -distal DNA elements to regulate gene transcription. RNA is transcribed from both of these DNA elements, and some DNA binding TFs bind RNA. Hence, RNA transcribed from regulatory elements may contribute to stable TF occupancy at these sites. We show that the ubiquitously expressed TF Yin-Yang 1 (YY1) binds to both gene regulatory elements and their associated RNA species across the entire genome. Reduced transcription of regulatory elements diminishes YY1 occupancy, whereas artificial tethering of RNA enhances YY1 occupancy at these elements. We propose that RNA makes a modest but important contribution to the maintenance of certain TFs at gene regulatory elements and suggest that transcription of regulatory elements produces a positive-feedback loop that contributes to the stability of gene expression programs.

  4. Circuitry and dynamics of human transcription factor regulatory networks

    PubMed Central

    Neph, Shane; Stergachis, Andrew B.; Reynolds, Alex; Sandstrom, Richard; Borenstein, Elhanan; Stamatoyannopoulos, John A.

    2012-01-01

    SUMMARY The combinatorial cross-regulation of hundreds of sequence-specific transcription factors defines a regulatory network that underlies cellular identity and function. Here we use genome-wide maps of in vivo DNaseI footprints to assemble an extensive core human regulatory network comprising connections among 475 sequence-specific transcription factors, and to analyze the dynamics of these connections across 41 diverse cell and tissue types. We find that human transcription factor networks are highly cell-selective and are driven by cohorts of factors that include regulators with previously unrecognized roles in control of cellular identity. Moreover, we identify many widely expressed factors that impact transcriptional regulatory networks in a cell-selective manner. Strikingly, in spite of their inherent diversity, all cell type regulatory networks independently converge on a common architecture that closely resembles the topology of living neuronal networks. Together, our results provide the first description of the circuitry, dynamics, and organizing principles of the human transcription factor regulatory network. PMID:22959076

  5. Foxo transcription factors control regulatory T cell development and function

    PubMed Central

    Kerdiles, Yann M.; Stone, Erica L.; Beisner, Daniel L.; McGargill, Maureen A.; Ch'en, Irene L.; Stockmann, Christian; Katayama, Carol D.; Hedrick, Stephen M.

    2010-01-01

    SUMMARY Foxo transcription factors integrate extrinsic signals to regulate cell division, differentiation and survival, and specific functions of lymphoid and myeloid cells. Here we showed the absence of Foxo1 severely curtailed the development of Foxp3+ regulatory T (Treg) cells, and those that developed were nonfunctional in vivo. The loss of function included diminished CTLA-4 receptor expression as the Ctla4 gene was a direct target of Foxo1. T cell specific loss of Foxo1 resulted in exocrine pancreatitis, hind limb paralysis, multi-organ lymphocyte infiltration, anti-nuclear antibodies and expanded germinal centers. Foxo-mediated control over Treg cell specification was further revealed by the inability of TGF-β cytokine to suppress T-bet transcription factor in the absence of Foxo1, resulting in IFN-γ-secretion. In addition the absence of Foxo3 exacerbated the effects of the loss of Foxo1. Thus, Foxo transcription factors guide the contingencies of T cell differentiation and specific functions of effector cell populations. PMID:21167754

  6. Anti-Sigma Factors in E. coli: Common Regulatory Mechanisms Controlling Sigma Factors Availability

    PubMed Central

    Treviño-Quintanilla, Luis Gerardo; Freyre-González, Julio Augusto; Martínez-Flores, Irma

    2013-01-01

    In bacteria, transcriptional regulation is a key step in cellular gene expression. All bacteria contain a core RNA polymerase that is catalytically competent but requires an additional σ factor for specific promoter recognition and correct transcriptional initiation. The RNAP core is not able to selectively bind to a given σ factor. In contrast, different σ factors have different affinities for the RNAP core. As a consequence, the concentration of alternate σ factors requires strict regulation in order to properly control the delicate interplay among them, which favors the competence for the RNAP core. This control is archived by different σ/anti-σ controlling mechanisms that shape complex regulatory networks and cascades, and enable the response to sudden environmental cues, whose global understanding is a current challenge for systems biology. Although there have been a number of excellent studies on each of these σ/anti-σ post-transcriptional regulatory systems, no comprehensive comparison of these mechanisms in a single model organism has been conducted. Here, we survey all these systems in E. coli dissecting and analyzing their inner workings and highlightin their differences. Then, following an integral approach, we identify their commonalities and outline some of the principles exploited by the cell to effectively and globally reprogram the transcriptional machinery. These principles provide guidelines for developing biological synthetic circuits enabling an efficient and robust response to sudden stimuli. PMID:24396271

  7. Regulatory coding of lymphoid lineage choice by hematopoietic transcription factors

    NASA Technical Reports Server (NTRS)

    Warren, Luigi A.; Rothenberg, Ellen V.

    2003-01-01

    During lymphopoiesis, precursor cells negotiate a complex regulatory space, defined by the levels of several competing and cross-regulating transcription factors, before arriving at stable states of commitment to the B-, T- and NK-specific developmental programs. Recent perturbation experiments provide evidence that this space has three major axes, corresponding to the PU.1 versus GATA-1 balance, the intensity of Notch signaling through the CSL pathway, and the ratio of E-box transcription factors to their Id protein antagonists.

  8. 14 CFR 1203.406 - Additional classification factors.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 14 Aeronautics and Space 5 2010-01-01 2010-01-01 false Additional classification factors. 1203.406... PROGRAM Guides for Original Classification § 1203.406 Additional classification factors. In determining the appropriate classification category, the following additional factors should be considered:...

  9. Roles of Interferon Regulatory Factors in Chronic Myeloid Leukemia.

    PubMed

    Manzella, Livia; Tirrò, Elena; Pennisi, Maria Stella; Massimino, Michele; Stella, Stefania; Romano, Chiara; Vitale, Silvia Rita; Vigneri, Paolo

    2016-01-01

    The Interferon Regulatory Factor (IRF) family consists of multiple transcription factors involved in the regulation of a variety of biological processes. Originally identified as transcriptional regulators of the type I interferon system, IRFs play a pivotal role in adaptive immunity, cell growth, differentiation and tumorigenesis. Hence, understanding IRF biology has important implications in the host response to cancer development and progression. Many lines of evidence suggest that different IRFs are involved in the pathogenesis of Chronic Myeloid Leukemia (CML), a myeloproliferative disorder caused by the BCR-ABL oncoprotein. BCR-ABL displays constitutive tyrosine kinase activity that favors cell proliferation, inhibits apoptosis and allows cell survival even in the absence of proper adhesion to the extracellular matrix. Different BCR-ABL tyrosine kinase inhibitors are currently available for CML treatment. These drugs are able to generate eight year CML-specific overall survival rates >90%, only a minority of patients will achieve molecular responses compatible with drug discontinuation. Thus, there is an unmet need for additional therapeutic targets that may lead to the cure of most patients diagnosed with CML. A growing body of evidence has suggested a role for both IRF4 and IRF8 in the pathogenesis of CML. Furthermore, IRF1 is consistently deleted at one or both alleles in patients with leukemia and myelodysplasia. Finally, we have recently demonstrated that IRF5 is a target of BCR-ABL kinase activity and reduces CML cell proliferation. In this article, we provide an update on the current knowledge of the role of the IRFs in CML. PMID:26728039

  10. Putative Regulatory Factors Associated with Intramuscular Fat Content

    PubMed Central

    Cesar, Aline S. M.; Regitano, Luciana C. A.; Koltes, James E.; Fritz-Waters, Eric R.; Lanna, Dante P. D.; Gasparin, Gustavo; Mourão, Gerson B.; Oliveira, Priscila S. N.; Reecy, James M.; Coutinho, Luiz L.

    2015-01-01

    Intramuscular fat (IMF) content is related to insulin resistance, which is an important prediction factor for disorders, such as cardiovascular disease, obesity and type 2 diabetes in human. At the same time, it is an economically important trait, which influences the sensorial and nutritional value of meat. The deposition of IMF is influenced by many factors such as sex, age, nutrition, and genetics. In this study Nellore steers (Bos taurus indicus subspecies) were used to better understand the molecular mechanisms involved in IMF content. This was accomplished by identifying differentially expressed genes (DEG), biological pathways and putative regulatory factors. Animals included in this study had extreme genomic estimated breeding value (GEBV) for IMF. RNA-seq analysis, gene set enrichment analysis (GSEA) and co-expression network methods, such as partial correlation coefficient with information theory (PCIT), regulatory impact factor (RIF) and phenotypic impact factor (PIF) were utilized to better understand intramuscular adipogenesis. A total of 16,101 genes were analyzed in both groups (high (H) and low (L) GEBV) and 77 DEG (FDR 10%) were identified between the two groups. Pathway Studio software identified 13 significantly over-represented pathways, functional classes and small molecule signaling pathways within the DEG list. PCIT analyses identified genes with a difference in the number of gene-gene correlations between H and L group and detected putative regulatory factors involved in IMF content. Candidate genes identified by PCIT include: ANKRD26, HOXC5 and PPAPDC2. RIF and PIF analyses identified several candidate genes: GLI2 and IGF2 (RIF1), MPC1 and UBL5 (RIF2) and a host of small RNAs, including miR-1281 (PIF). These findings contribute to a better understanding of the molecular mechanisms that underlie fat content and energy balance in muscle and provide important information for the production of healthier beef for human consumption. PMID:26042666

  11. Putative regulatory factors associated with intramuscular fat content.

    PubMed

    Cesar, Aline S M; Regitano, Luciana C A; Koltes, James E; Fritz-Waters, Eric R; Lanna, Dante P D; Gasparin, Gustavo; Mourão, Gerson B; Oliveira, Priscila S N; Reecy, James M; Coutinho, Luiz L

    2015-01-01

    Intramuscular fat (IMF) content is related to insulin resistance, which is an important prediction factor for disorders, such as cardiovascular disease, obesity and type 2 diabetes in human. At the same time, it is an economically important trait, which influences the sensorial and nutritional value of meat. The deposition of IMF is influenced by many factors such as sex, age, nutrition, and genetics. In this study Nellore steers (Bos taurus indicus subspecies) were used to better understand the molecular mechanisms involved in IMF content. This was accomplished by identifying differentially expressed genes (DEG), biological pathways and putative regulatory factors. Animals included in this study had extreme genomic estimated breeding value (GEBV) for IMF. RNA-seq analysis, gene set enrichment analysis (GSEA) and co-expression network methods, such as partial correlation coefficient with information theory (PCIT), regulatory impact factor (RIF) and phenotypic impact factor (PIF) were utilized to better understand intramuscular adipogenesis. A total of 16,101 genes were analyzed in both groups (high (H) and low (L) GEBV) and 77 DEG (FDR 10%) were identified between the two groups. Pathway Studio software identified 13 significantly over-represented pathways, functional classes and small molecule signaling pathways within the DEG list. PCIT analyses identified genes with a difference in the number of gene-gene correlations between H and L group and detected putative regulatory factors involved in IMF content. Candidate genes identified by PCIT include: ANKRD26, HOXC5 and PPAPDC2. RIF and PIF analyses identified several candidate genes: GLI2 and IGF2 (RIF1), MPC1 and UBL5 (RIF2) and a host of small RNAs, including miR-1281 (PIF). These findings contribute to a better understanding of the molecular mechanisms that underlie fat content and energy balance in muscle and provide important information for the production of healthier beef for human consumption.

  12. The effect of hyperammonemia on myostatin and myogenic regulatory factor gene expression in broiler embryos

    PubMed Central

    Stern, R.A.; Ashwell, C.M.; Dasarathy, S.; Mozdziak, P.E.

    2015-01-01

    Myogenesis is facilitated by four myogenic regulatory factors and is significantly inhibited by myostatin. The objective of the current study was to examine embryonic gene regulation of myostatin/myogenic regulatory factors, and subsequent manipulations of protein synthesis, in broiler embryos under induced hyperammonemia. Broiler eggs were injected with ammonium acetate solution four times over 48 hours beginning on either embryonic day (ED) 15 or 17. Serum ammonia concentration was significantly higher (P < 0.05) in ammonium acetate injected embryos for both ED17 and ED19 collected samples when compared to sham-injected controls. Expression of mRNA, extracted from pectoralis major of experimental and control embryos, was measured using real-time quantitative PCR for myostatin, myogenic regulatory factors myogenic factor 5, myogenic determination factor 1, myogenin, myogenic regulatory factor 4, and paired box 7. A significantly lower (P < 0.01) myostatin expression was accompanied by a higher serum ammonia concentration in both ED17 and ED19 collected samples. Myogenic factor 5 expression was higher (P < 0.05) in ED17 collected samples administered ammonium acetate. In both ED17 and ED19 collected samples, myogenic regulatory factor 4 was lower (P ≤ 0.05) in ammonium acetate injected embryos. No significant difference was seen in myogenic determination factor 1, myogenin, or paired box 7 expression between treatment groups for either age of sample collection. Additionally, there was no significant difference in BrdU staining of histological samples taken from treated and control embryos. Myostatin protein levels were evaluated by Western blot analysis, and also showed lower myostatin expression (P < 0.05). Overall, it appears possible to inhibit myostatin expression through hyperammonemia, which is expected to have a positive effect on embryonic myogenesis and postnatal muscle growth. PMID:25689990

  13. Regulatory role of the respiratory supercomplex factors in Saccharomyces cerevisiae

    PubMed Central

    Rydström Lundin, Camilla; Ott, Martin; Ädelroth, Pia; Brzezinski, Peter

    2016-01-01

    The respiratory supercomplex factors (Rcf) 1 and 2 mediate supramolecular interactions between mitochondrial complexes III (ubiquinol-cytochrome c reductase; cyt. bc1) and IV (cytochrome c oxidase; CytcO). In addition, removal of these polypeptides results in decreased activity of CytcO, but not of cyt. bc1. In the present study, we have investigated the kinetics of ligand binding, the single-turnover reaction of CytcO with O2, and the linked cyt. bc1-CytcO quinol oxidation-oxygen-reduction activities in mitochondria in which Rcf1 or Rcf2 were removed genetically (strains rcf1Δ and rcf2Δ, respectively). The data show that in the rcf1Δ and rcf2Δ strains, in a significant fraction of the population, ligand binding occurs over a time scale that is ∼100-fold faster (τ ≅ 100 μs) than observed with the wild-type mitochondria (τ ≅ 10 ms), indicating structural changes. This effect is specific to removal of Rcf and not dissociation of the cyt. bc1–CytcO supercomplex. Furthermore, in the rcf1Δ and rcf2Δ strains, the single-turnover reaction of CytcO with O2 was incomplete. This observation indicates that the lower activity of CytcO is caused by a fraction of inactive CytcO rather than decreased CytcO activity of the entire population. Furthermore, the data suggest that the Rcf1 polypeptide mediates formation of an electron-transfer bridge from cyt. bc1 to CytcO via a tightly bound cyt. c. We discuss the significance of the proposed regulatory mechanism of Rcf1 and Rcf2 in the context of supramolecular interactions between cyt. bc1 and CytcO. PMID:27432958

  14. Regulation of APC Activity by Phosphorylation and Regulatory Factors

    PubMed Central

    Kotani, Shuji; Tanaka, Hirofumi; Yasuda, Hideyo; Todokoro, Kazuo

    1999-01-01

    Ubiquitin-dependent proteolysis of Cut2/Pds1 and Cyclin B is required for sister chromatid separation and exit from mitosis, respectively. Anaphase-promoting complex/cyclosome (APC) specifically ubiquitinates Cut2/Pds1 at metaphase–anaphase transition, and ubiquitinates Cyclin B in late mitosis and G1 phase. However, the exact regulatory mechanism of substrate-specific activation of mammalian APC with the right timing remains to be elucidated. We found that not only the binding of the activators Cdc20 and Cdh1 and the inhibitor Mad2 to APC, but also the phosphorylation of Cdc20 and Cdh1 by Cdc2-Cyclin B and that of APC by Polo-like kinase and cAMP-dependent protein kinase, regulate APC activity. The cooperation of the phosphorylation/dephosphorylation and the regulatory factors in regulation of APC activity may thus control the precise progression of mitosis. PMID:10459014

  15. 14 CFR 1203.406 - Additional classification factors.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 14 Aeronautics and Space 5 2013-01-01 2013-01-01 false Additional classification factors. 1203.406 Section 1203.406 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.406 Additional classification factors. In...

  16. 14 CFR 1203.406 - Additional classification factors.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 14 Aeronautics and Space 5 2011-01-01 2010-01-01 true Additional classification factors. 1203.406 Section 1203.406 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.406 Additional classification factors. In...

  17. 14 CFR 1203.406 - Additional classification factors.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 14 Aeronautics and Space 5 2012-01-01 2012-01-01 false Additional classification factors. 1203.406 Section 1203.406 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.406 Additional classification factors. In...

  18. Interaction of Trypanosoma cruzi adenylate cyclase with liver regulatory factors.

    PubMed Central

    Eisenschlos, C; Flawiá, M M; Torruella, M; Torres, H N

    1986-01-01

    Trypanosoma cruzi adenylate cyclase catalytic subunits may interact with regulatory factors from rat liver membranes, reconstituting heterologous systems which are catalytically active in assay mixtures containing MgATP. The systems show stimulatory responses to glucagon and guanosine 5'-[beta gamma-imido]triphosphate (p[NH]ppG) or fluoride. Reconstitution was obtained by three different methods: fusion of rat liver membranes (pretreated with N-ethylmaleimide) to T. cruzi membranes; interaction of detergent extracts of rat liver membranes with T. cruzi membranes; or interaction of purified preparations of T. cruzi adenylate cyclase and of liver membrane factors in phospholipid vesicles. The liver factors responsible for the guanine nucleotide effect were characterized as the NS protein. Data also indicate that reconstitution requires the presence of a membrane substrate. PMID:2947568

  19. Interaction between Major Nitrogen Regulatory Protein NIT2 and Pathway-Specific Regulatory Factor NIT4 Is Required for Their Synergistic Activation of Gene Expression in Neurospora crassa

    PubMed Central

    Feng, Bo; Marzluf, George A.

    1998-01-01

    In Neurospora crassa, the major nitrogen regulatory protein, NIT2, a member of the GATA family of transcription factors, controls positively the expression of numerous genes which specify nitrogen catabolic enzymes. Expression of the highly regulated structural gene nit-3, which encodes nitrate reductase, is dependent upon a synergistic interaction of NIT2 with a pathway-specific control protein, NIT4, a member of the GAL4 family of fungal regulatory factors. The NIT2 and NIT4 proteins both bind at specific recognition elements in the nit-3 promoter, but, in addition, we show that a direct protein-protein interaction between NIT2 and NIT4 is essential for optimal expression of the nit-3 structural gene. Neurospora possesses at least five different GATA factors which control different areas of cellular function, but which have a similar DNA binding specificity. Significantly, only NIT2, of the several Neurospora GATA factors examined, interacts with NIT4. We propose that protein-protein interactions of the individual GATA factors with additional pathway-specific regulatory factors determine each of their specific regulatory functions. PMID:9632783

  20. Uncovering Transcriptional Regulatory Networks by Sparse Bayesian Factor Model

    NASA Astrophysics Data System (ADS)

    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.

  1. Bayesian non-negative factor analysis for reconstructing transcription factor mediated regulatory networks

    PubMed Central

    2011-01-01

    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. PMID:22166063

  2. 14 CFR § 1203.406 - Additional classification factors.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 14 Aeronautics and Space 5 2014-01-01 2014-01-01 false Additional classification factors. § 1203.406 Section § 1203.406 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.406 Additional classification...

  3. Clinical trials in "emerging markets": regulatory considerations and other factors.

    PubMed

    Singh, Romi; Wang, Ouhong

    2013-11-01

    Clinical studies are being placed in emerging markets as part of global drug development programs to access large pool of eligible patients and to benefit from a cost effective structure. However, over the last few years, the definition of "emerging markets" is being revisited, especially from a regulatory perspective. For purposes of this article, countries outside US, EU and the traditional "western countries" are discussed. Multiple factors are considered for placement of clinical studies such as adherence to Good Clinical Practice (GCP), medical infrastructure & standard of care, number of eligible patients, etc. This article also discusses other quantitative factors such as country's GDP, patent applications, healthcare expenditure, healthcare infrastructure, corruption, innovation, etc. These different factors and indexes are correlated to the number of clinical studies ongoing in the "emerging markets". R&D, healthcare expenditure, technology infrastructure, transparency, and level of innovation, show a significant correlation with the number of clinical trials being conducted in these countries. This is the first analysis of its kind to evaluate and correlate the various other factors to the number of clinical studies in a country. PMID:24070788

  4. Clinical trials in "emerging markets": regulatory considerations and other factors.

    PubMed

    Singh, Romi; Wang, Ouhong

    2013-11-01

    Clinical studies are being placed in emerging markets as part of global drug development programs to access large pool of eligible patients and to benefit from a cost effective structure. However, over the last few years, the definition of "emerging markets" is being revisited, especially from a regulatory perspective. For purposes of this article, countries outside US, EU and the traditional "western countries" are discussed. Multiple factors are considered for placement of clinical studies such as adherence to Good Clinical Practice (GCP), medical infrastructure & standard of care, number of eligible patients, etc. This article also discusses other quantitative factors such as country's GDP, patent applications, healthcare expenditure, healthcare infrastructure, corruption, innovation, etc. These different factors and indexes are correlated to the number of clinical studies ongoing in the "emerging markets". R&D, healthcare expenditure, technology infrastructure, transparency, and level of innovation, show a significant correlation with the number of clinical trials being conducted in these countries. This is the first analysis of its kind to evaluate and correlate the various other factors to the number of clinical studies in a country.

  5. Plants with genetically modified events combined by conventional breeding: an assessment of the need for additional regulatory data.

    PubMed

    Pilacinski, W; Crawford, A; Downey, R; Harvey, B; Huber, S; Hunst, P; Lahman, L K; MacIntosh, S; Pohl, M; Rickard, C; Tagliani, L; Weber, N

    2011-01-01

    Crop varieties with multiple GM events combined by conventional breeding have become important in global agriculture. The regulatory requirements in different countries for such products vary considerably, placing an additional burden on regulatory agencies in countries where the submission of additional data is required and delaying the introduction of innovative products to meet agricultural needs. The process of conventional plant breeding has predictably provided safe food and feed products both historically and in the modern era of plant breeding. Thus, previously approved GM events that have been combined by conventional plant breeding and contain GM traits that are not likely to interact in a manner affecting safety should be considered to be as safe as their conventional counterparts. Such combined GM event crop varieties should require little, if any, additional regulatory data to meet regulatory requirements.

  6. Pervasive variation of transcription factor orthologs contributes to regulatory network evolution.

    PubMed

    Nadimpalli, Shilpa; Persikov, Anton V; Singh, Mona

    2015-03-01

    Differences in transcriptional regulatory networks underlie much of the phenotypic variation observed across organisms. Changes to cis-regulatory elements are widely believed to be the predominant means by which regulatory networks evolve, yet examples of regulatory network divergence due to transcription factor (TF) variation have also been observed. To systematically ascertain the extent to which TFs contribute to regulatory divergence, we analyzed the evolution of the largest class of metazoan TFs, Cys2-His2 zinc finger (C2H2-ZF) TFs, across 12 Drosophila species spanning ~45 million years of evolution. Remarkably, we uncovered that a significant fraction of all C2H2-ZF 1-to-1 orthologs in flies exhibit variations that can affect their DNA-binding specificities. In addition to loss and recruitment of C2H2-ZF domains, we found diverging DNA-contacting residues in ~44% of domains shared between D. melanogaster and the other fly species. These diverging DNA-contacting residues, found in ~70% of the D. melanogaster C2H2-ZF genes in our analysis and corresponding to ~26% of all annotated D. melanogaster TFs, show evidence of functional constraint: they tend to be conserved across phylogenetic clades and evolve slower than other diverging residues. These same variations were rarely found as polymorphisms within a population of D. melanogaster flies, indicating their rapid fixation. The predicted specificities of these dynamic domains gradually change across phylogenetic distances, suggesting stepwise evolutionary trajectories for TF divergence. Further, whereas proteins with conserved C2H2-ZF domains are enriched in developmental functions, those with varying domains exhibit no functional enrichments. Our work suggests that a subset of highly dynamic and largely unstudied TFs are a likely source of regulatory variation in Drosophila and other metazoans.

  7. Identification of tissue-specific cis-regulatory modules based on interactions between transcription factors

    PubMed Central

    Yu, Xueping; Lin, Jimmy; Zack, Donald J; Qian, Jiang

    2007-01-01

    Background Evolutionary conservation has been used successfully to help identify cis-acting DNA regions that are important in regulating tissue-specific gene expression. Motivated by increasing evidence that some DNA regulatory regions are not evolutionary conserved, we have developed an approach for cis-regulatory region identification that does not rely upon evolutionary sequence conservation. Results The conservation-independent approach is based on an empirical potential energy between interacting transcription factors (TFs). In this analysis, the potential energy is defined as a function of the number of TF interactions in a genomic region and the strength of the interactions. By identifying sets of interacting TFs, the analysis locates regions enriched with the binding sites of these interacting TFs. We applied this approach to 30 human tissues and identified 6232 putative cis-regulatory modules (CRMs) regulating 2130 tissue-specific genes. Interestingly, some genes appear to be regulated by different CRMs in different tissues. Known regulatory regions are highly enriched in our predicted CRMs. In addition, DNase I hypersensitive sites, which tend to be associated with active regulatory regions, significantly overlap with the predicted CRMs, but not with more conserved regions. We also find that conserved and non-conserved CRMs regulate distinct gene groups. Conserved CRMs control more essential genes and genes involved in fundamental cellular activities such as transcription. In contrast, non-conserved CRMs, in general, regulate more non-essential genes, such as genes related to neural activity. Conclusion These results demonstrate that identifying relevant sets of binding motifs can help in the mapping of DNA regulatory regions, and suggest that non-conserved CRMs play an important role in gene regulation. PMID:17996093

  8. CaMKII in addition to MLCK contributes to phosphorylation of regulatory light chain in cardiomyocytes.

    PubMed

    Eikemo, Hilde; Moltzau, Lise Román; Hussain, Rizwan I; Nguyen, Cam H T; Qvigstad, Eirik; Levy, Finn Olav; Skomedal, Tor; Osnes, Jan-Bjørn

    2016-02-26

    The aim was to identify kinase activities involved in the phosphorylation of regulatory light chain (RLC) in situ in cardiomyocytes. In electrically stimulated rat cardiomyocytes, phosphatase inhibition by calyculin A unmasked kinase activities evoking an increase of phosphorylated RLC (P-RLC) from about 16% to about 80% after 80 min. The phosphorylation rate in cardiomyocytes was reduced by about 40% by the myosin light chain kinase (MLCK) inhibitor, ML-7. In rat ventricular muscle strips, calyculin A induced a positive inotropic effect that correlated with P-RLC levels. The inotropic effect and P-RLC elevation were abolished by ML-7 treatment. The kinase activities phosphorylating RLC in cardiomyocytes were reduced by about 60% by the non-selective kinase inhibitor staurosporine and by about 50% by the calmodulin antagonist W7. W7 eliminated the inhibitory effect of ML-7, suggesting that the cardiac MLCK is Ca(2+)/calmodulin (CaM)-dependent. The CaM-dependent kinase II (CaMKII) inhibitor KN-93 attenuated the calyculin A-induced RLC phosphorylation by about 40%, indicating a contribution from CaMKII. The residual phosphorylation in the presence of W7 indicated that also CaM-independent kinase activities might contribute. RLC phosphorylation was insensitive to protein kinase C inhibition. In conclusion, in addition to MLCK, CaMKII phosphorylates RLC in cardiomyocytes. Involvement of other kinases cannot be excluded.

  9. [Requirements for drug approval and additional benefits assessment: Regulatory aspects and experiences].

    PubMed

    Broich, K; Löbker, W; Schulte, A; Beinlich, P; Müller, T

    2016-04-01

    The early assessment of benefits of newly approved drugs with novel active substances or new applications, which came into force on 1 January 2011 still represents a challenge to all parties involved. This article highlights the definitions, regulatory requirements and interaction between drug marketing approval and early assessment of benefits in Germany. The constellation of an extensively harmonized European and even international drug authorization process with a predominantly national regulation of drug reimbursement situation inevitably causes friction, which could be markedly reduced through early joint advisory discussions during the planning phase for pivotal clinical trials. During the year 2015 the Federal Institute for Drugs and Medical Devices (BfArM) carried out 300 scientific advice procedures of which 34 were concerned with applications in the field of indications for the central nervous system (CNS). In comparison 98 advisory meetings were held by the Federal Joint Committee (G-BA) of which the BfArM provided advice in 12 instances and in 2 cases on CNS indications. Study design, endpoints and appropriate comparative therapies are the key issues in exchanges and discussions between the BfArM, the G‑BA and applicants. Under these aspects the BfArM and G‑BA promote an early and consistent involvement in early advice procedures regarding the prerequisites for drug approval and assessment of additional benefits.

  10. [Requirements for drug approval and additional benefits assessment: Regulatory aspects and experiences].

    PubMed

    Broich, K; Löbker, W; Schulte, A; Beinlich, P; Müller, T

    2016-04-01

    The early assessment of benefits of newly approved drugs with novel active substances or new applications, which came into force on 1 January 2011 still represents a challenge to all parties involved. This article highlights the definitions, regulatory requirements and interaction between drug marketing approval and early assessment of benefits in Germany. The constellation of an extensively harmonized European and even international drug authorization process with a predominantly national regulation of drug reimbursement situation inevitably causes friction, which could be markedly reduced through early joint advisory discussions during the planning phase for pivotal clinical trials. During the year 2015 the Federal Institute for Drugs and Medical Devices (BfArM) carried out 300 scientific advice procedures of which 34 were concerned with applications in the field of indications for the central nervous system (CNS). In comparison 98 advisory meetings were held by the Federal Joint Committee (G-BA) of which the BfArM provided advice in 12 instances and in 2 cases on CNS indications. Study design, endpoints and appropriate comparative therapies are the key issues in exchanges and discussions between the BfArM, the G‑BA and applicants. Under these aspects the BfArM and G‑BA promote an early and consistent involvement in early advice procedures regarding the prerequisites for drug approval and assessment of additional benefits. PMID:27003322

  11. Arabidopsis Ensemble Reverse-Engineered Gene Regulatory Network Discloses Interconnected Transcription Factors in Oxidative Stress[W

    PubMed Central

    Vermeirssen, Vanessa; De Clercq, Inge; Van Parys, Thomas; Van Breusegem, Frank; Van de Peer, Yves

    2014-01-01

    The abiotic stress response in plants is complex and tightly controlled by gene regulation. We present an abiotic stress gene regulatory network of 200,014 interactions for 11,938 target genes by integrating four complementary reverse-engineering solutions through average rank aggregation on an Arabidopsis thaliana microarray expression compendium. This ensemble performed the most robustly in benchmarking and greatly expands upon the availability of interactions currently reported. Besides recovering 1182 known regulatory interactions, cis-regulatory motifs and coherent functionalities of target genes corresponded with the predicted transcription factors. We provide a valuable resource of 572 abiotic stress modules of coregulated genes with functional and regulatory information, from which we deduced functional relationships for 1966 uncharacterized genes and many regulators. Using gain- and loss-of-function mutants of seven transcription factors grown under control and salt stress conditions, we experimentally validated 141 out of 271 predictions (52% precision) for 102 selected genes and mapped 148 additional transcription factor-gene regulatory interactions (49% recall). We identified an intricate core oxidative stress regulatory network where NAC13, NAC053, ERF6, WRKY6, and NAC032 transcription factors interconnect and function in detoxification. Our work shows that ensemble reverse-engineering can generate robust biological hypotheses of gene regulation in a multicellular eukaryote that can be tested by medium-throughput experimental validation. PMID:25549671

  12. Molecular characterization of interferon regulatory factor 1 in Bubalus bubalis.

    PubMed

    Stafuzza, N B; Borges, M M; Amaral-Trusty, M E J

    2015-01-01

    Interferon regulatory factor 1 (IRF1) is functionally diverse in the regulation of immune response and is considered to be an important candidate gene for studying disease susceptibility in mammals. In this paper, we characterized the whole sequence of the IRF1 gene in river buffalo (Bubalus bubalis) and compared genomic and the amino acid sequences between different species. The buffalo IRF1 gene was 7099 bp long and organized into 10 exons and nine introns. Its molecular structure showed exactly the same number of exons (10) and introns (nine) in bovids, mice, horses, humans, and chickens. However, rats did not have exon 5, but had the largest exon 4, which suggests that exon 5 was incorporated into exon 4. The coding and the amino acid sequences of the gene showed that identity varied from 73 to 99% at the coding sequence level and from 61 to 100% at the amino acid level when compared with other mammals and chickens. Comparative analysis of the gene sequence between two different buffalo breeds, Murrah and Mediterranean, revealed six potential SNPs that are primarily located in the 5' and 3'UTRs. PMID:26400319

  13. Altered oncomodules underlie chromatin regulatory factors driver mutations.

    PubMed

    Frigola, Joan; Iturbide, Ane; Lopez-Bigas, Nuria; Peiro, Sandra; Gonzalez-Perez, Abel

    2016-05-24

    Chromatin regulatory factors (CRFs), are known to be involved in tumorigenesis in several cancer types. Nevertheless, the molecular mechanisms through which driver alterations of CRFs cause tumorigenesis remain unknown. Here, we developed a CRFs Oncomodules Discovery approach, which mines several sources of cancer genomics and perturbaomics data. The approach prioritizes sets of genes significantly miss-regulated in primary tumors (oncomodules) bearing mutations of driver CRFs. We applied the approach to eleven TCGA tumor cohorts and uncovered oncomodules potentially associated to mutations of five driver CRFs in three cancer types. Our results revealed, for example, the potential involvement of the mTOR pathway in the development of tumors with loss-of-function mutations of MLL2 in head and neck squamous cell carcinomas. The experimental validation that MLL2 loss-of-function increases the sensitivity of cancer cell lines to mTOR inhibition lends further support to the validity of our approach. The potential oncogenic modules detected by our approach may guide experiments proposing ways to indirectly target driver mutations of CRFs. PMID:27095575

  14. Biosynthesis of nitric oxide activates iron regulatory factor in macrophages.

    PubMed

    Drapier, J C; Hirling, H; Wietzerbin, J; Kaldy, P; Kühn, L C

    1993-09-01

    Biosynthesis of nitric oxide (NO) from L-arginine modulates activity of iron-dependent enzymes, including mitochondrial acontiase, an [Fe-S] protein. We examined the effect of NO on the activity of iron regulatory factor (IRF), a cytoplasmic protein which modulates both ferritin mRNA translation and transferrin receptor mRNA stability by binding to specific mRNA sequences called iron responsive elements (IREs). Murine macrophages were activated with interferon-gamma and lipopolysaccharide to induce NO synthase activity and cultured in the presence or absence of NG-substituted analogues of L-arginine which served as selective inhibitors of NO synthesis. Measurement of the nitrite concentration in the culture medium was taken as an index of NO production. Mitochondria-free cytosols were then prepared and aconitase activity as well as IRE binding activity and induction of IRE binding activity were correlated and depended on NO synthesis after IFN-gamma and/or LPS stimulation. Authentic NO gas as well as the NO-generating compound 3-morpholinosydnonimine (SIN-1) also conversely modulated aconitase and IRE binding activities of purified recombinant IRF. These results provide evidence that endogenously produced NO may modulate the post-transcriptional regulation of genes involved in iron homeostasis and support the hypothesis that the [Fe-S] cluster of IRF mediates iron-dependent regulation. PMID:7504626

  15. A role for interferon regulatory factor 4 in receptor editing.

    PubMed

    Pathak, Simanta; Ma, Shibin; Trinh, Long; Lu, Runqing

    2008-04-01

    Receptor editing is the primary means through which B cells revise antigen receptors and maintain central tolerance. Previous studies have demonstrated that interferon regulatory factor 4 (IRF-4) and IRF-8 promote immunoglobulin light-chain rearrangement and transcription at the pre-B stage. Here, the roles of IRF-4 and -8 in receptor editing were analyzed. Our results show that secondary rearrangement was impaired in IRF-4 but not IRF-8 mutant mice, suggesting that receptor editing is defective in the absence of IRF-4. The role of IRF-4 in receptor editing was further examined in B-cell-receptor (BCR) transgenic mice. Our results show that secondary rearrangement triggered by membrane-bound antigen was defective in the IRF-4-deficient mice. Our results further reveal that the defect in secondary rearrangement is more severe at the immunoglobulin lambda locus than at the kappa locus, indicating that IRF-4 is more critical for the lambda rearrangement. We provide evidence demonstrating that the expression of IRF-4 in immature B cells is rapidly induced by self-antigen and that the reconstitution of IRF-4 expression in the IRF-4 mutant immature B cells promotes secondary rearrangement. Thus, our studies identify IRF-4 as a nuclear effector of a BCR signaling pathway that promotes secondary rearrangement at the immature B-cell stage.

  16. Interferon regulatory factors: at the crossroads of immunity, metabolism, and disease.

    PubMed

    Zhao, Guang-Nian; Jiang, Ding-Sheng; Li, Hongliang

    2015-02-01

    The interferon-regulatory factor (IRF) family comprises nine members in mammals. Although this transcription factor family was originally thought to function primarily in the immune system, contributing to both the innate immune response and the development of immune cells, recent advances have revealed that IRFs plays critical roles in other biological processes, such as metabolism. Accordingly, abnormalities in the expression and/or function of IRFs have increasingly been linked to disease. Herein, we provide an update on the recent progress regarding the regulation of immune responses and immune cell development associated with IRFs. Additionally, we discuss the relationships between IRFs and immunity, metabolism, and disease, with a particular focus on the role of IRFs as stress sensors. This article is part of a Special Issue entitled: Autophagy and protein quality control in cardiometabolic diseases.

  17. Increments and duplication events of enzymes and transcription factors influence metabolic and regulatory diversity in prokaryotes.

    PubMed

    Martínez-Núñez, Mario Alberto; Poot-Hernandez, Augusto Cesar; Rodríguez-Vázquez, Katya; Perez-Rueda, Ernesto

    2013-01-01

    In this work, the content of enzymes and DNA-binding transcription factors (TFs) in 794 non-redundant prokaryotic genomes was evaluated. The identification of enzymes was based on annotations deposited in the KEGG database as well as in databases of functional domains (COG and PFAM) and structural domains (Superfamily). For identifications of the TFs, hidden Markov profiles were constructed based on well-known transcriptional regulatory families. From these analyses, we obtained diverse and interesting results, such as the negative rate of incremental changes in the number of detected enzymes with respect to the genome size. On the contrary, for TFs the rate incremented as the complexity of genome increased. This inverse related performance shapes the diversity of metabolic and regulatory networks and impacts the availability of enzymes and TFs. Furthermore, the intersection of the derivatives between enzymes and TFs was identified at 9,659 genes, after this point, the regulatory complexity grows faster than metabolic complexity. In addition, TFs have a low number of duplications, in contrast to the apparent high number of duplications associated with enzymes. Despite the greater number of duplicated enzymes versus TFs, the increment by which duplicates appear is higher in TFs. A lower proportion of enzymes among archaeal genomes (22%) than in the bacterial ones (27%) was also found. This low proportion might be compensated by the interconnection between the metabolic pathways in Archaea. A similar proportion was also found for the archaeal TFs, for which the formation of regulatory complexes has been proposed. Finally, an enrichment of multifunctional enzymes in Bacteria, as a mechanism of ecological adaptation, was detected. PMID:23922780

  18. Increments and Duplication Events of Enzymes and Transcription Factors Influence Metabolic and Regulatory Diversity in Prokaryotes

    PubMed Central

    Martínez-Núñez, Mario Alberto; Poot-Hernandez, Augusto Cesar; Rodríguez-Vázquez, Katya; Perez-Rueda, Ernesto

    2013-01-01

    In this work, the content of enzymes and DNA-binding transcription factors (TFs) in 794 non-redundant prokaryotic genomes was evaluated. The identification of enzymes was based on annotations deposited in the KEGG database as well as in databases of functional domains (COG and PFAM) and structural domains (Superfamily). For identifications of the TFs, hidden Markov profiles were constructed based on well-known transcriptional regulatory families. From these analyses, we obtained diverse and interesting results, such as the negative rate of incremental changes in the number of detected enzymes with respect to the genome size. On the contrary, for TFs the rate incremented as the complexity of genome increased. This inverse related performance shapes the diversity of metabolic and regulatory networks and impacts the availability of enzymes and TFs. Furthermore, the intersection of the derivatives between enzymes and TFs was identified at 9,659 genes, after this point, the regulatory complexity grows faster than metabolic complexity. In addition, TFs have a low number of duplications, in contrast to the apparent high number of duplications associated with enzymes. Despite the greater number of duplicated enzymes versus TFs, the increment by which duplicates appear is higher in TFs. A lower proportion of enzymes among archaeal genomes (22%) than in the bacterial ones (27%) was also found. This low proportion might be compensated by the interconnection between the metabolic pathways in Archaea. A similar proportion was also found for the archaeal TFs, for which the formation of regulatory complexes has been proposed. Finally, an enrichment of multifunctional enzymes in Bacteria, as a mechanism of ecological adaptation, was detected. PMID:23922780

  19. Increments and duplication events of enzymes and transcription factors influence metabolic and regulatory diversity in prokaryotes.

    PubMed

    Martínez-Núñez, Mario Alberto; Poot-Hernandez, Augusto Cesar; Rodríguez-Vázquez, Katya; Perez-Rueda, Ernesto

    2013-01-01

    In this work, the content of enzymes and DNA-binding transcription factors (TFs) in 794 non-redundant prokaryotic genomes was evaluated. The identification of enzymes was based on annotations deposited in the KEGG database as well as in databases of functional domains (COG and PFAM) and structural domains (Superfamily). For identifications of the TFs, hidden Markov profiles were constructed based on well-known transcriptional regulatory families. From these analyses, we obtained diverse and interesting results, such as the negative rate of incremental changes in the number of detected enzymes with respect to the genome size. On the contrary, for TFs the rate incremented as the complexity of genome increased. This inverse related performance shapes the diversity of metabolic and regulatory networks and impacts the availability of enzymes and TFs. Furthermore, the intersection of the derivatives between enzymes and TFs was identified at 9,659 genes, after this point, the regulatory complexity grows faster than metabolic complexity. In addition, TFs have a low number of duplications, in contrast to the apparent high number of duplications associated with enzymes. Despite the greater number of duplicated enzymes versus TFs, the increment by which duplicates appear is higher in TFs. A lower proportion of enzymes among archaeal genomes (22%) than in the bacterial ones (27%) was also found. This low proportion might be compensated by the interconnection between the metabolic pathways in Archaea. A similar proportion was also found for the archaeal TFs, for which the formation of regulatory complexes has been proposed. Finally, an enrichment of multifunctional enzymes in Bacteria, as a mechanism of ecological adaptation, was detected.

  20. Expression of myogenic regulatory factors and myo-endothelial remodeling in sporadic inclusion body myositis

    PubMed Central

    Wanschitz, Julia V.; Dubourg, Odile; Lacene, Emmanuelle; Fischer, Michael B.; Höftberger, Romana; Budka, Herbert; Romero, Norma B.; Eymard, Bruno; Herson, Serge; Butler-Browne, Gillian S.; Voit, Thomas; Benveniste, Olivier

    2013-01-01

    Muscle repair relies on coordinated activation and differentiation of satellite cells, a process that is unable to counterbalance progressive degeneration in sporadic inclusion body myositis (s-IBM). To explore features of myo regeneration, the expression of myogenic regulatory factors Pax7, MyoD and Myogenin and markers of regenerating fibers was analyzed by immunohistochemistry in s-IBM muscle compared with polymyositis, dermatomyositis, muscular dystrophy and age-matched controls. In addition, the capillary density and number of interstitial CD34+ hematopoietic progenitor cells was determined by double-immunoflourescence staining. Satellite cells and regenerating fibers were significantly increased in s-IBM similar to other inflammatory myopathies and correlated with the intensity of inflammation (R > 0.428). Expression of MyoD, visualizing activated satellite cells and proliferating myoblasts, was lower in s-IBM compared to polymyosits. In contrast, Myogenin a marker of myogenic cell differentiation was strongly up-regulated in s-IBM muscle. The microvascular architecture in s-IBM was distorted, although the capillary density was normal. Notably, CD34+ hematopoietic cells were significantly increased in the interstitial compartment. Our findings indicate profound myo-endothelial remodeling of s-IBM muscle concomitant to inflammation. An altered expression of myogenic regulatory factors involved in satellite cell activation and differentiation, however, might reflect perturbations of muscle repair in s-IBM. PMID:23058947

  1. Identification of Neurodegenerative Factors Using Translatome-Regulatory Network Analysis

    PubMed Central

    Brichta, Lars; Shin, William; Jackson-Lewis, Vernice; Blesa, Javier; Yap, Ee-Lynn; Walker, Zachary; Zhang, Jack; Roussarie, Jean-Pierre; Alvarez, Mariano J.; Califano, Andrea; Przedborski, Serge; Greengard, Paul

    2016-01-01

    For degenerative disorders of the central nervous system, the major obstacle to therapeutic advancement has been the challenge of identifying the key molecular mechanisms underlying neuronal loss. We developed a combinatorial approach including translational profiling and brain regulatory network analysis to search for key determinants of neuronal survival or death. Following the generation of transgenic mice for cell type-specific profiling of midbrain dopaminergic neurons, we established and compared translatome libraries reflecting the molecular signature of these cells at baseline or under degenerative stress. Analysis of these libraries by interrogating a context-specific brain regulatory network led to the identification of a repertoire of intrinsic upstream regulators that drive the dopaminergic stress response. The altered activity of these regulators was not associated with changes in their expression levels. This strategy can be generalized for the elucidation of novel molecular determinants involved in the degeneration of other classes of neurons. PMID:26214373

  2. Chromatin properties of regulatory DNA probed by manipulation of transcription factors.

    PubMed

    Sharov, Alexei A; Nishiyama, Akira; Qian, Yong; Dudekula, Dawood B; Longo, Dan L; Schlessinger, David; Ko, Minoru S H

    2014-08-01

    Transcription factors (TFs) bind to DNA and regulate the transcription of nearby genes. However, only a small fraction of TF binding sites have such regulatory effects. Here we search for the predictors of functional binding sites by carrying out a systematic computational screening of a variety of contextual factors (histone modifications, nuclear lamin-bindings, and cofactor bindings). We used regression analysis to test if contextual factors are associated with upregulation or downregulation of neighboring genes following the induction or knockdown of the 9 TFs in mouse embryonic stem (ES) cells. Functional TF binding sites appeared to be either active (i.e., bound by P300, CHD7, mediator, cohesin, and SWI/SNF) or repressed (i.e., with H3K27me3 histone marks and bound by Polycomb factors). Active binding sites mediated the downregulation of nearby genes upon knocking down the activating TFs or inducing repressors. Repressed TF binding sites mediated the upregulation of nearby genes (e.g., poised developmental regulators) upon inducing TFs. In addition, repressed binding sites mediated repressive effects of TFs, identified by the downregulation of target genes after the induction of TFs or by the upregulation of target genes after the knockdown of TFs. The contextual factors associated with functions of DNA-bound TFs were used to improve the identification of candidate target genes regulated by TFs.

  3. Using graphical adaptive lasso approach to construct transcription factor and microRNA's combinatorial regulatory network in breast cancer.

    PubMed

    Su, Naifang; Dai, Ding; Deng, Chao; Qian, Minping; Deng, Minghua

    2014-06-01

    Discovering the regulation of cancer-related gene is of great importance in cancer biology. Transcription factors and microRNAs are two kinds of crucial regulators in gene expression, and they compose a combinatorial regulatory network with their target genes. Revealing the structure of this network could improve the authors' understanding of gene regulation, and further explore the molecular pathway in cancer. In this article, the authors propose a novel approach graphical adaptive lasso (GALASSO) to construct the regulatory network in breast cancer. GALASSO use a Gaussian graphical model with adaptive lasso penalties to integrate the sequence information as well as gene expression profiles. The simulation study and the experimental profiles verify the accuracy of the authors' approach. The authors further reveal the structure of the regulatory network, and explore the role of feedforward loops in gene regulation. In addition, the authors discuss the combinatorial regulatory effect between transcription factors and microRNAs, and select miR-155 for detailed analysis of microRNA's role in cancer. The proposed GALASSO approach is an efficient method to construct the combinatorial regulatory network. It also provides a new way to integrate different data sources and could find more applications in meta-analysis problem.

  4. The safety and regulatory status of food, drug and cosmetics colour additives exempt from certification.

    PubMed

    Hallagan, J B; Allen, D C; Borzelleca, J F

    1995-06-01

    Colour additives exempt from certification (also known as 'natural colour additives') are commonly used in the United States to colour foods, drugs and cosmetics. The US Food and Drug Administration established regulations governing the use of these colour additives, and the labelling of the products that contain them. The safety of these colour additives has been demonstrated by safety testing programs that have yielded a significant amount of toxicology data, and also by a long and well documented history of safe use in the United States and elsewhere. In this report we review the data supporting the safety of the colour additives that are exempt from certification and approved for use in the USA, and conclude that the safety of these colour additives is well supported by the available data. Relevant aspects of the regulation of these colour additives are also described. PMID:7797179

  5. Additives

    NASA Technical Reports Server (NTRS)

    Smalheer, C. V.

    1973-01-01

    The chemistry of lubricant additives is discussed to show what the additives are chemically and what functions they perform in the lubrication of various kinds of equipment. Current theories regarding the mode of action of lubricant additives are presented. The additive groups discussed include the following: (1) detergents and dispersants, (2) corrosion inhibitors, (3) antioxidants, (4) viscosity index improvers, (5) pour point depressants, and (6) antifouling agents.

  6. MicroRNA and transcription factor mediated regulatory network for ovarian cancer: regulatory network of ovarian cancer.

    PubMed

    Ying, Huanchun; Lv, Jing; Ying, Tianshu; Li, Jun; Yang, Qing; Ma, Yuan

    2013-10-01

    A better understanding on the regulatory interactions of microRNA (miRNA) target genes and transcription factor (TF) target genes in ovarian cancer may be conducive for developing early diagnosis strategy. Thus, gene expression data and miRNA expression data were downloaded from The Cancer Genome Atlas in this study. Differentially expressed genes and miRNAs were selected out with t test, and Gene Ontology enrichment analysis was performed with DAVID tools. Regulatory interactions were retrieved from miRTarBase, TRED, and TRANSFAC, and then networks for miRNA target genes and TF target genes were constructed to globally present the mechanisms. As a result, a total of 1,939 differentially expressed genes were identified, and they were enriched in 28 functions, among which cell cycle was affected to the most degree. Besides, 213 differentially expressed miRNAs were identified. Two regulatory networks for miRNA target genes and TF target genes were established and then both were combined, in which E2F transcription factor 1, cyclin-dependent kinase inhibitor 1A, cyclin E1, and miR-16 were the hub genes. These genes may be potential biomarkers for ovarian cancer.

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

    PubMed Central

    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

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

    NASA Astrophysics Data System (ADS)

    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.

  9. Construction and analysis of dynamic transcription factor regulatory networks in the progression of glioma.

    PubMed

    Li, Yongsheng; Shao, Tingting; Jiang, Chunjie; Bai, Jing; Wang, Zishan; Zhang, Jinwen; Zhang, Lili; Zhao, Zheng; Xu, Juan; Li, Xia

    2015-11-03

    The combinatorial cross-regulation of transcription factors (TFs) plays an important role in cellular identity and function; however, the dynamic regulation of TFs during glioma progression remains largely unknown. Here, we used the genome-wide expression of TFs to construct an extensive human TF network comprising interactions among 513 TFs and to analyse the dynamics of the TF-TF network during glioma progression. We found that the TF regulatory networks share a common architecture and that the topological structures are conserved. Strikingly, despite the conservation of the network architecture, TF regulatory networks are highly grade specific, and TF circuitry motifs are dynamically rewired during glioma progression. In addition, the most frequently observed structure in the grade-specific TF networks was the feedforward loop (FFL). We described applications that show how investigating the behaviour of FFLs in glioblastoma can reveal FFLs (such as RARG-NR1I2-CDX2) that are associated with prognosis. We constructed comprehensive TF-TF networks and systematically analysed the circuitry, dynamics, and topological principles of the networks during glioma progression, which will further enhance our understanding of the functions of TFs in glioma.

  10. Tumor suppressor properties of the splicing regulatory factor RBM10

    PubMed Central

    Hernández, Jordi; Bechara, Elias; Schlesinger, Doerte; Delgado, Javier; Serrano, Luis; Valcárcel, Juan

    2016-01-01

    ABSTRACT RBM10 is an RNA binding protein and alternative splicing regulator frequently mutated in lung adenocarcinomas. Recent results indicate that RBM10 inhibits proliferation of lung cancer cells by promoting skipping of exon 9 of the gene NUMB, a frequent alternative splicing change in lung cancer generating a negative regulator of Notch signaling. Complementing these observations, we show that knock down of RBM10 in human cancer cells enhances growth of mouse tumor xenografts, confirming that RBM10 acts as a tumor suppressor, while knock down of an oncogenic mutant version of RBM10 reduces xenograft tumor growth. A RBM10 mutation found in lung cancer cells, V354E, disrupts RBM10-mediated regulation of NUMB alternative splicing, inducing the cell proliferation-promoting isoform. We now show that 2 natural RBM10 isoforms that differ by the presence or absence of V354 in the second RNA Recognition Motif (RRM2), display similar regulatory effects on NUMB alternative splicing, suggesting that V354E actively disrupts RBM10 activity. Structural modeling localizes V354 in the outside surface of one α-helix opposite to the RNA binding surface of RBM10, and we show that the mutation does not compromise binding of the RRM2 domain to NUMB RNA regulatory sequences. We further show that other RBM10 mutations found in lung adenocarcinomas also compromise regulation of NUMB exon 9. Collectively, our previous and current results reveal that RBM10 is a tumor suppressor that represses Notch signaling and cell proliferation through the regulation of NUMB alternative splicing. PMID:26853560

  11. Effect of multiplicative and additive noise on genetic transcriptional regulatory mechanism

    NASA Astrophysics Data System (ADS)

    Liu, Xue-Mei; Xie, Hui-Zhang; Liu, Liang-Gang; Li, Zhi-Bing

    2009-02-01

    A multiplicative noise and an additive noise are introduced in the kinetic model of Smolen-Baxter-Byrne [P. Smolen, D.A. Baxter, J.H. Byrne, Amer. J. Physiol. Cell. Physiol. 274 (1998) 531], in which the expression of gene is controlled by protein concentration of transcriptional activator. The Fokker-Planck equation is solved and the steady-state probability distribution is obtained numerically. It is found that the multiplicative noise converts the bistability to monostability that can be regarded as a noise-induced transition. The additive noise reduces the transcription efficiency. The correlation between the multiplicative noise and the additive noise works as a genetic switch and regulates the gene transcription effectively.

  12. AP-42 ADDITIONS AND REVISIONS - TRANSPORTABILITY FACTORS FOR FUGITIVE DUST

    EPA Science Inventory

    The product is a table of factors, one for each county in the US, reflecting the portion of fugitive dust removed very close to the source via impaction on vegetation and similar mechanisms. Factors were based on land cover in area (county or grid cell) A praft final product was...

  13. Transcription factor binding at enhancers: shaping a genomic regulatory landscape in flux

    PubMed Central

    Palstra, Robert-Jan; Grosveld, Frank

    2012-01-01

    The mammalian genome is packed tightly in the nucleus of the cell. This packing is primarily facilitated by histone proteins and results in an ordered organization of the genome in chromosome territories that can be roughly divided in heterochromatic and euchromatic domains. On top of this organization several distinct gene regulatory elements on the same chromosome or other chromosomes are thought to dynamically communicate via chromatin looping. Advances in genome-wide technologies have revealed the existence of a plethora of these regulatory elements in various eukaryotic genomes. These regulatory elements are defined by particular in vitro assays as promoters, enhancers, insulators, and boundary elements. However, recent studies indicate that the in vivo distinction between these elements is often less strict. Regulatory elements are bound by a mixture of common and lineage-specific transcription factors which mediate the long-range interactions between these elements. Inappropriate modulation of the binding of these transcription factors can alter the interactions between regulatory elements, which in turn leads to aberrant gene expression with disease as an ultimate consequence. Here we discuss the bi-modal behavior of regulatory elements that act in cis (with a focus on enhancers), how their activity is modulated by transcription factor binding and the effect this has on gene regulation. PMID:23060900

  14. Enhancement of alkaloid production in opium and California poppy by transactivation using heterologous regulatory factors.

    PubMed

    Apuya, Nestor R; Park, Joon-Hyun; Zhang, Liping; Ahyow, Maurice; Davidow, Patricia; Van Fleet, Jennifer; Rarang, Joel C; Hippley, Matthew; Johnson, Thomas W; Yoo, Hye-Dong; Trieu, Anthony; Krueger, Shannon; Wu, Chuan-yin; Lu, Yu-ping; Flavell, Richard B; Bobzin, Steven C

    2008-02-01

    Genes encoding regulatory factors isolated from Arabidopsis, soybean and corn have been screened to identify those that modulate the expression of genes encoding for enzymes involved in the biosynthesis of morphinan alkaloids in opium poppy (Papaver somniferum) and benzophenanthridine alkaloids in California poppy (Eschscholzia californica). In opium poppy, the over-expression of selected regulatory factors increased the levels of PsCOR (codeinone reductase), Ps4'OMT (S-adenosyl-l-methionine:3'-hydroxy-N-methylcoclaurine 4'-O-methyltransferase) and Ps6OMT [(R,S)-norcoclaurine 6-O-methyltransferase] transcripts by 10- to more than 100-fold. These transcriptional activations translated into an enhancement of alkaloid production in opium poppy of up to at least 10-fold. In California poppy, the transactivation effect of regulatory factor WRKY1 resulted in an increase of up to 60-fold in the level of EcCYP80B1 [(S)-N-methylcoclaurine 3'-hydroxylase] and EcBBE (berberine bridge enzyme) transcripts. As a result, the accumulations of selected alkaloid intermediates were enhanced up to 30-fold. The transactivation effects of other regulatory factors led to the accumulation of the same intermediates. These regulatory factors also led to the production of new alkaloids in California poppy callus culture.

  15. Enhancement of alkaloid production in opium and California poppy by transactivation using heterologous regulatory factors.

    PubMed

    Apuya, Nestor R; Park, Joon-Hyun; Zhang, Liping; Ahyow, Maurice; Davidow, Patricia; Van Fleet, Jennifer; Rarang, Joel C; Hippley, Matthew; Johnson, Thomas W; Yoo, Hye-Dong; Trieu, Anthony; Krueger, Shannon; Wu, Chuan-yin; Lu, Yu-ping; Flavell, Richard B; Bobzin, Steven C

    2008-02-01

    Genes encoding regulatory factors isolated from Arabidopsis, soybean and corn have been screened to identify those that modulate the expression of genes encoding for enzymes involved in the biosynthesis of morphinan alkaloids in opium poppy (Papaver somniferum) and benzophenanthridine alkaloids in California poppy (Eschscholzia californica). In opium poppy, the over-expression of selected regulatory factors increased the levels of PsCOR (codeinone reductase), Ps4'OMT (S-adenosyl-l-methionine:3'-hydroxy-N-methylcoclaurine 4'-O-methyltransferase) and Ps6OMT [(R,S)-norcoclaurine 6-O-methyltransferase] transcripts by 10- to more than 100-fold. These transcriptional activations translated into an enhancement of alkaloid production in opium poppy of up to at least 10-fold. In California poppy, the transactivation effect of regulatory factor WRKY1 resulted in an increase of up to 60-fold in the level of EcCYP80B1 [(S)-N-methylcoclaurine 3'-hydroxylase] and EcBBE (berberine bridge enzyme) transcripts. As a result, the accumulations of selected alkaloid intermediates were enhanced up to 30-fold. The transactivation effects of other regulatory factors led to the accumulation of the same intermediates. These regulatory factors also led to the production of new alkaloids in California poppy callus culture. PMID:17961129

  16. Mangiferin inhibits macrophage classical activation via downregulating interferon regulatory factor 5 expression.

    PubMed

    Wei, Zhiquan; Yan, Li; Chen, Yixin; Bao, Chuanhong; Deng, Jing; Deng, Jiagang

    2016-08-01

    Mangiferin is a natural polyphenol and the predominant effective component of Mangifera indica Linn. leaves. For hundreds of years, Mangifera indica Linn. leaf has been used as an ingredient in numerous traditional Chinese medicine preparations for the treatment of bronchitis. However, the pharmacological mechanism of mangiferin in the treatment of bronchitis remains to be elucidated. Macrophage classical activation is important role in the process of bronchial airway inflammation, and interferon regulatory factor 5 (IRF5) has been identified as a key regulatory factor for macrophage classical activation. The present study used the THP‑1 human monocyte cell line to investigate whether mangiferin inhibits macrophage classical activation via suppressing IRF5 expression in vitro. THP‑1 cells were differentiated to macrophages by phorbol 12‑myristate 13‑acetate. Macrophages were polarized to M1 macrophages following stimulation with lipopolysaccharide (LPS)/interferon‑γ (IFN‑γ). Flow cytometric analysis was conducted to detect the M1 macrophages. Reverse transcription‑quantitative polymerase chain reaction was used to investigate cellular IRF5 gene expression. Levels of proinflammatory cytokines and IRF5 were assessed following cell culture and cellular homogenization using enzyme‑linked immunosorbent assay. IRF5 protein and nuclei co‑localization was performed in macrophages with laser scanning confocal microscope immunofluorescence analysis. The results of the present study demonstrated that mangiferin significantly inhibits LPS/IFN‑γ stimulation‑induced classical activation of macrophages in vitro and markedly decreases proinflammatory cytokine release. In addition, cellular IRF5 expression was markedly downregulated. These results suggest that the inhibitory effect of mangiferin on classical activation of macrophages may be exerted via downregulation of cellular IRF5 expression levels. PMID:27277156

  17. Mangiferin inhibits macrophage classical activation via downregulating interferon regulatory factor 5 expression

    PubMed Central

    Wei, Zhiquan; Yan, Li; Chen, Yixin; Bao, Chuanhong; Deng, Jing; Deng, Jiagang

    2016-01-01

    Mangiferin is a natural polyphenol and the predominant effective component of Mangifera indica Linn. leaves. For hundreds of years, Mangifera indica Linn. leaf has been used as an ingredient in numerous traditional Chinese medicine preparations for the treatment of bronchitis. However, the pharmacological mechanism of mangiferin in the treatment of bronchitis remains to be elucidated. Macrophage classical activation is important role in the process of bronchial airway inflammation, and interferon regulatory factor 5 (IRF5) has been identified as a key regulatory factor for macrophage classical activation. The present study used the THP-1 human monocyte cell line to investigate whether mangiferin inhibits macrophage classical activation via suppressing IRF5 expression in vitro. THP-1 cells were differentiated to macrophages by phorbol 12-myristate 13-acetate. Macrophages were polarized to M1 macrophages following stimulation with lipopolysaccharide (LPS)/interferon-γ (IFN-γ). Flow cytometric analysis was conducted to detect the M1 macrophages. Reverse transcription-quantitative polymerase chain reaction was used to investigate cellular IRF5 gene expression. Levels of proinflammatory cytokines and IRF5 were assessed following cell culture and cellular homogenization using enzyme-linked immunosorbent assay. IRF5 protein and nuclei co-localization was performed in macrophages with laser scanning confocal microscope immunofluorescence analysis. The results of the present study demonstrated that mangiferin significantly inhibits LPS/IFN-γ stimulation-induced classical activation of macrophages in vitro and markedly decreases proinflammatory cytokine release. In addition, cellular IRF5 expression was markedly downregulated. These results suggest that the inhibitory effect of mangiferin on classical activation of macrophages may be exerted via downregulation of cellular IRF5 expression levels. PMID:27277156

  18. Reconstruction and topological characterization of the sigma factor regulatory network of Mycobacterium tuberculosis.

    PubMed

    Chauhan, Rinki; Ravi, Janani; Datta, Pratik; Chen, Tianlong; Schnappinger, Dirk; Bassler, Kevin E; Balázsi, Gábor; Gennaro, Maria Laura

    2016-03-31

    Accessory sigma factors, which reprogram RNA polymerase to transcribe specific gene sets, activate bacterial adaptive responses to noxious environments. Here we reconstruct the complete sigma factor regulatory network of the human pathogen Mycobacterium tuberculosis by an integrated approach. The approach combines identification of direct regulatory interactions between M. tuberculosis sigma factors in an E. coli model system, validation of selected links in M. tuberculosis, and extensive literature review. The resulting network comprises 41 direct interactions among all 13 sigma factors. Analysis of network topology reveals (i) a three-tiered hierarchy initiating at master regulators, (ii) high connectivity and (iii) distinct communities containing multiple sigma factors. These topological features are likely associated with multi-layer signal processing and specialized stress responses involving multiple sigma factors. Moreover, the identification of overrepresented network motifs, such as autoregulation and coregulation of sigma and anti-sigma factor pairs, provides structural information that is relevant for studies of network dynamics.

  19. Reconstruction and topological characterization of the sigma factor regulatory network of Mycobacterium tuberculosis

    PubMed Central

    Chauhan, Rinki; Ravi, Janani; Datta, Pratik; Chen, Tianlong; Schnappinger, Dirk; Bassler, Kevin E.; Balázsi, Gábor; Gennaro, Maria Laura

    2016-01-01

    Accessory sigma factors, which reprogram RNA polymerase to transcribe specific gene sets, activate bacterial adaptive responses to noxious environments. Here we reconstruct the complete sigma factor regulatory network of the human pathogen Mycobacterium tuberculosis by an integrated approach. The approach combines identification of direct regulatory interactions between M. tuberculosis sigma factors in an E. coli model system, validation of selected links in M. tuberculosis, and extensive literature review. The resulting network comprises 41 direct interactions among all 13 sigma factors. Analysis of network topology reveals (i) a three-tiered hierarchy initiating at master regulators, (ii) high connectivity and (iii) distinct communities containing multiple sigma factors. These topological features are likely associated with multi-layer signal processing and specialized stress responses involving multiple sigma factors. Moreover, the identification of overrepresented network motifs, such as autoregulation and coregulation of sigma and anti-sigma factor pairs, provides structural information that is relevant for studies of network dynamics. PMID:27029515

  20. Duplication of a promiscuous transcription factor drives the emergence of a new regulatory network.

    PubMed

    Pougach, Ksenia; Voet, Arnout; Kondrashov, Fyodor A; Voordeckers, Karin; Christiaens, Joaquin F; Baying, Bianka; Benes, Vladimir; Sakai, Ryo; Aerts, Jan; Zhu, Bo; Van Dijck, Patrick; Verstrepen, Kevin J

    2014-01-01

    The emergence of new genes throughout evolution requires rewiring and extension of regulatory networks. However, the molecular details of how the transcriptional regulation of new gene copies evolves remain largely unexplored. Here we show how duplication of a transcription factor gene allowed the emergence of two independent regulatory circuits. Interestingly, the ancestral transcription factor was promiscuous and could bind different motifs in its target promoters. After duplication, one paralogue evolved increased binding specificity so that it only binds one type of motif, whereas the other copy evolved a decreased activity so that it only activates promoters that contain multiple binding sites. Interestingly, only a few mutations in both the DNA-binding domains and in the promoter binding sites were required to gradually disentangle the two networks. These results reveal how duplication of a promiscuous transcription factor followed by concerted cis and trans mutations allows expansion of a regulatory network.

  1. Duplication of a promiscuous transcription factor drives the emergence of a new regulatory network

    PubMed Central

    Pougach, Ksenia; Voet, Arnout; Kondrashov, Fyodor A.; Voordeckers, Karin; Christiaens, Joaquin F.; Baying, Bianka; Benes, Vladimir; Sakai, Ryo; Aerts, Jan; Zhu, Bo; Van Dijck, Patrick; Verstrepen, Kevin J.

    2014-01-01

    The emergence of new genes throughout evolution requires rewiring and extension of regulatory networks. However, the molecular details of how the transcriptional regulation of new gene copies evolves remain largely unexplored. Here we show how duplication of a transcription factor gene allowed the emergence of two independent regulatory circuits. Interestingly, the ancestral transcription factor was promiscuous and could bind different motifs in its target promoters. After duplication, one paralogue evolved increased binding specificity so that it only binds one type of motif, whereas the other copy evolved a decreased activity so that it only activates promoters that contain multiple binding sites. Interestingly, only a few mutations in both the DNA-binding domains and in the promoter binding sites were required to gradually disentangle the two networks. These results reveal how duplication of a promiscuous transcription factor followed by concerted cis and trans mutations allows expansion of a regulatory network. PMID:25204769

  2. Sparse Additive Ordinary Differential Equations for Dynamic Gene Regulatory Network Modeling

    PubMed Central

    Wu, Hulin; Lu, Tao; Xue, Hongqi; Liang, Hua

    2014-01-01

    Summary The gene regulation network (GRN) is a high-dimensional complex system, which can be represented by various mathematical or statistical models. The ordinary differential equation (ODE) model is one of the popular dynamic GRN models. High-dimensional linear ODE models have been proposed to identify GRNs, but with a limitation of the linear regulation effect assumption. In this article, we propose a sparse additive ODE (SA-ODE) model, coupled with ODE estimation methods and adaptive group LASSO techniques, to model dynamic GRNs that could flexibly deal with nonlinear regulation effects. The asymptotic properties of the proposed method are established and simulation studies are performed to validate the proposed approach. An application example for identifying the nonlinear dynamic GRN of T-cell activation is used to illustrate the usefulness of the proposed method. PMID:25061254

  3. Sparse Additive Ordinary Differential Equations for Dynamic Gene Regulatory Network Modeling.

    PubMed

    Wu, Hulin; Lu, Tao; Xue, Hongqi; Liang, Hua

    2014-04-01

    The gene regulation network (GRN) is a high-dimensional complex system, which can be represented by various mathematical or statistical models. The ordinary differential equation (ODE) model is one of the popular dynamic GRN models. High-dimensional linear ODE models have been proposed to identify GRNs, but with a limitation of the linear regulation effect assumption. In this article, we propose a sparse additive ODE (SA-ODE) model, coupled with ODE estimation methods and adaptive group LASSO techniques, to model dynamic GRNs that could flexibly deal with nonlinear regulation effects. The asymptotic properties of the proposed method are established and simulation studies are performed to validate the proposed approach. An application example for identifying the nonlinear dynamic GRN of T-cell activation is used to illustrate the usefulness of the proposed method. PMID:25061254

  4. Decreased Transcription Factor Binding Levels Nearby Primate Pseudogenes Suggest Regulatory Degeneration

    PubMed Central

    Douglas, Gavin M.; Wilson, Michael D.; Moses, Alan M.

    2016-01-01

    Characteristics of pseudogene degeneration at the coding level are well-known, such as a shift toward neutral rates of nonsynonymous substitutions and gain of frameshift mutations. In contrast, degeneration of pseudogene transcriptional regulation is not well understood. Here, we test two predictions of regulatory degeneration along a pseudogenized lineage: 1) Decreased transcription factor (TF) binding and 2) accelerated evolution in putative cis-regulatory regions. We find evidence for decreased TF binding levels nearby two primate pseudogenes compared with functional liver genes. However, the majority of TF-bound sequences nearby pseudogenes do not show evidence for lineage-specific accelerated rates of evolution. We conclude that decreases in TF binding level could be a marker for regulatory degeneration, while sequence degeneration in primate cis-regulatory modules may be obscured by background rates of TF binding site turnover. PMID:26882985

  5. Gene Regulatory Network Inference of Immunoresponsive Gene 1 (IRG1) Identifies Interferon Regulatory Factor 1 (IRF1) as Its Transcriptional Regulator in Mammalian Macrophages

    PubMed Central

    Tallam, Aravind; Perumal, Thaneer M.; Antony, Paul M.; Jäger, Christian; Fritz, Joëlle V.; Vallar, Laurent; Balling, Rudi; del Sol, Antonio; Michelucci, Alessandro

    2016-01-01

    Immunoresponsive gene 1 (IRG1) is one of the highest induced genes in macrophages under pro-inflammatory conditions. Its function has been recently described: it codes for immune-responsive gene 1 protein/cis-aconitic acid decarboxylase (IRG1/CAD), an enzyme catalysing the production of itaconic acid from cis-aconitic acid, a tricarboxylic acid (TCA) cycle intermediate. Itaconic acid possesses specific antimicrobial properties inhibiting isocitrate lyase, the first enzyme of the glyoxylate shunt, an anaplerotic pathway that bypasses the TCA cycle and enables bacteria to survive on limited carbon conditions. To elucidate the mechanisms underlying itaconic acid production through IRG1 induction in macrophages, we examined the transcriptional regulation of IRG1. To this end, we studied IRG1 expression in human immune cells under different inflammatory stimuli, such as TNFα and IFNγ, in addition to lipopolysaccharides. Under these conditions, as previously shown in mouse macrophages, IRG1/CAD accumulates in mitochondria. Furthermore, using literature information and transcription factor prediction models, we re-constructed raw gene regulatory networks (GRNs) for IRG1 in mouse and human macrophages. We further implemented a contextualization algorithm that relies on genome-wide gene expression data to infer putative cell type-specific gene regulatory interactions in mouse and human macrophages, which allowed us to predict potential transcriptional regulators of IRG1. Among the computationally identified regulators, siRNA-mediated gene silencing of interferon regulatory factor 1 (IRF1) in macrophages significantly decreased the expression of IRG1/CAD at the gene and protein level, which correlated with a reduced production of itaconic acid. Using a synergistic approach of both computational and experimental methods, we here shed more light on the transcriptional machinery of IRG1 expression and could pave the way to therapeutic approaches targeting itaconic acid levels

  6. Specific detection of interferon regulatory factor 5 (IRF5): A case of antibody inequality

    PubMed Central

    Li, Dan; De, Saurav; Li, Dan; Song, Su; Matta, Bharati; Barnes, Betsy J.

    2016-01-01

    Interferon regulatory factor 5 (IRF5) is a member of the IRF family of transcription factors. IRF5 was first identified and characterized as a transcriptional regulator of type I interferon expression after virus infection. In addition to its critical role(s) in the regulation and development of host immunity, subsequent studies revealed important roles for IRF5 in autoimmunity, cancer, obesity, pain, cardiovascular disease, and metabolism. Based on these important disease-related findings, a large number of commercial antibodies have become available to study the expression and function of IRF5. Here we validate a number of these antibodies for the detection of IRF5 by immunoblot, flow cytometry, and immunofluorescence or immunohistochemistry using well-established positive and negative controls. Somewhat surprising, the majority of commercial antibodies tested were unable to specifically recognize human or mouse IRF5. We present data on antibodies that do specifically recognize human or mouse IRF5 in a particular application. These findings reiterate the importance of proper controls and molecular weight standards for the analysis of protein expression. Given that dysregulated IRF5 expression has been implicated in the pathogenesis of numerous diseases, including autoimmune and cancer, results indicate that caution should be used in the evaluation and interpretation of IRF5 expression analysis. PMID:27481535

  7. Aberrant expression of interferon regulatory factor 3 in human lung cancer

    SciTech Connect

    Tokunaga, Takayuki; Naruke, Yuki; Shigematsu, Sayuri; Kohno, Tomoko; Yasui, Kiyoshi; Ma, Yuhua; Chua, Koon Jiew; Katayama, Ikuo; Nakamura, Takashi; Hishikawa, Yoshitaka; Koji, Takehiko; Yatabe, Yasushi; Nagayasu, Takeshi; Fujita, Takashi; Matsuyama, Toshifumi; and others

    2010-06-25

    We analyzed the subcellular distributions and gene structures of interferon regulatory factor 3 (IRF3) transcription factor in 50 cases of human primary lung cancer. The immunohistochemical analyses revealed substantially aberrant IRF3 expression specific to the cancer lesions (2 and 6 tumors with nuclear staining, and 4 and 5 tumors with negative staining, in adenocarcinoma and squamous cell carcinoma, respectively), while the morphologically normal region around the tumors exhibited only cytoplasmic staining. In addition, we determined the sequence of the entire IRF3 coding region, and found two novel variants with the amino acid changes (S{sup 175}(AGC) {yields} R{sup 175}(CGC) and A{sup 208}(GCC) {yields} D{sup 208}(GAC)). The R{sup 175} variant was also detected in a morphologically normal region around the nuclear staining squamous cell carcinoma, and exhibited almost the same functions as the wild type IRF3. On the other hand, the D{sup 208} variant, found in the negative staining squamous cell carcinoma cases, reduced the nuclear translocation in response to I{kappa}B kinase {epsilon} stimulation, as compared to the wild type IRF3, but the same variant was detected in the surrounding morphologically normal region. The aberrant expression of IRF3 and the novel D{sup 208} variant may provide clues to elucidate the etiology of primary lung cancer.

  8. Multiple signaling pathways leading to the activation of interferon regulatory factor 3.

    PubMed

    Servant, Marc J; Grandvaux, Nathalie; Hiscott, John

    2002-09-01

    Virus infection of susceptible cells activates multiple signaling pathways that orchestrate the activation of genes, such as cytokines, involved in the antiviral and innate immune response. Among the kinases induced are the mitogen-activated protein (MAP) kinases, Jun-amino terminal kinases (JNK) and p38, the IkappaB kinase (IKK) and DNA-PK. In addition, virus infection also activates an uncharacterized VAK responsible for the C-terminal phosphorylation and subsequent activation of interferon regulatory factor 3 (IRF-3). Virus-mediated activation of IRF-3 through VAK is dependent on viral entry and transcription, since replication deficient virus failed to induce IRF-3 activity. The pathways leading to VAK activation are not well characterized, but IRF-3 appears to represent a novel cellular detection pathway that recognizes viral nucleocapsid (N) structure. Recently, the range of inducers responsible for IRF-3 activation has increased. In addition to virus infection, recognition of bacterial infection mediated through lipopolysaccharide by Toll-like receptor 4 has also been reported. Furthermore, MAP kinase kinase kinase (MAP KKK)-related pathways and DNA-PK induce N-terminal phosphorylation of IRF-3. This review summarizes recent observations in the identification of novel signaling pathways leading to IRF-3 activation.

  9. Discovering transcription factor regulatory targets using gene expression and binding data

    PubMed Central

    Maienschein-Cline, Mark; Zhou, Jie; White, Kevin P.; Sciammas, Roger; Dinner, Aaron R.

    2012-01-01

    Motivation: Identifying the target genes regulated by transcription factors (TFs) is the most basic step in understanding gene regulation. Recent advances in high-throughput sequencing technology, together with chromatin immunoprecipitation (ChIP), enable mapping TF binding sites genome wide, but it is not possible to infer function from binding alone. This is especially true in mammalian systems, where regulation often occurs through long-range enhancers in gene-rich neighborhoods, rather than proximal promoters, preventing straightforward assignment of a binding site to a target gene. Results: We present EMBER (Expectation Maximization of Binding and Expression pRofiles), a method that integrates high-throughput binding data (e.g. ChIP-chip or ChIP-seq) with gene expression data (e.g. DNA microarray) via an unsupervised machine learning algorithm for inferring the gene targets of sets of TF binding sites. Genes selected are those that match overrepresented expression patterns, which can be used to provide information about multiple TF regulatory modes. We apply the method to genome-wide human breast cancer data and demonstrate that EMBER confirms a role for the TFs estrogen receptor alpha, retinoic acid receptors alpha and gamma in breast cancer development, whereas the conventional approach of assigning regulatory targets based on proximity does not. Additionally, we compare several predicted target genes from EMBER to interactions inferred previously, examine combinatorial effects of TFs on gene regulation and illustrate the ability of EMBER to discover multiple modes of regulation. Availability: All code used for this work is available at http://dinner-group.uchicago.edu/downloads.html Contact: dinner@uchicago.edu Supplementary Information: Supplementary data are available at Bioinformatics online. PMID:22084256

  10. Tissue factor activation: is disulfide bond switching a regulatory mechanism?

    PubMed Central

    Ghosh, Samit; Mandal, Samir K.

    2007-01-01

    A majority of tissue factor (TF) on cell surfaces exists in a cryptic form (ie, coagulation function inactive) but retains its functionality in cell signaling. Recent studies have suggested that cryptic TF contains unpaired cysteine thiols and that activation involves the formation of the disulfide bond Cys186-Cys 209 and that protein disulfide isomerase (PDI) regulates TF coagulant and signaling activities by targeting this disulfide bond. This study was carried out to investigate the validity of this novel concept. Although treatment of MDA 231 tumor cells, fibroblasts, and stimulated endothelial cells with the oxidizing agent HgCl2 markedly increased the cell-surface TF coagulant activity, the increase is associated with increased anionic phospholipids at the cell surface. Annexin V, which binds to anionic phospholipids, attenuated the increased TF coagulant activity. It is noteworthy that treatment of cells with reducing agents also increased the cell surface TF activity. No evidence was found for either detectable expression of PDI at the cell surface or association of TF with PDI. Furthermore, reduction of PDI with the gene silencing had no effect on either TF coagulant or cell signaling functions. Overall, the present data undermine the recently proposed hypothesis that PDI-mediated disulfide exchange plays a role in regulating TF procoagulant and cell signaling functions. PMID:17726162

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

    PubMed

    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. Insights into the Interferon Regulatory Factor Activation from the Crystal Structure of Dimeric IRF5

    SciTech Connect

    Chen, W.; Lam, S; Srinath, H; Jiang, Z; Correia, J; Schiffer, C; Fitzgerald, K; Lin, K; Royer, Jr., W

    2008-01-01

    The interferon regulatory factors (IRFs) are involved in the innate immune response and are activated by phosphorylation. The structure of a pseudophosphorylated IRF5 activation domain now reveals structural changes in the activated form that would turn an autoinhibitory region into a dimerization interface. In vivo analysis supports the relevance of such a dimer to transcriptional activation.

  13. Affinity Density: a novel genomic approach to the identification of transcription factor regulatory targets

    PubMed Central

    Hazelett, Dennis J.; Lakeland, Daniel L.; Weiss, Joseph B.

    2009-01-01

    Methods: A new method was developed for identifying novel transcription factor regulatory targets based on calculating Local Affinity Density. Techniques from the signal-processing field were used, in particular the Hann digital filter, to calculate the relative binding affinity of different regions based on previously published in vitro binding data. To illustrate this approach, the complete genomes of Drosophila melanogaster and D.pseudoobscura were analyzed for binding sites of the homeodomain proteinc Tinman, an essential heart development gene in both Drosophila and Mouse. The significant binding regions were identified relative to genomic background and assigned to putative target genes. Valid candidates common to both species of Drosophila were selected as a test of conservation. Results: The new method was more sensitive than cluster searches for conserved binding motifs with respect to positive identification of known Tinman targets. Our Local Affinity Density method also identified a significantly greater proportion of Tinman-coexpressed genes than equivalent, optimized cluster searching. In addition, this new method predicted a significantly greater than expected number of genes with previously published RNAi phenotypes in the heart. Availability: Algorithms were implemented in Python, LISP, R and maxima, using MySQL to access locally mirrored sequence data from Ensembl (D.melanogaster release 4.3) and flybase (D.pseudoobscura). All code is licensed under GPL and freely available at http://www.ohsu.edu/cellbio/dev_biol_prog/affinitydensity/. Contact: hazelett@ohsu.edu PMID:19401399

  14. Interferon Regulatory Factor 6 Has a Protective Role in the Host Response to Endotoxic Shock

    PubMed Central

    Volk, Paige; Moreland, Jessica G.; Dunnwald, Martine

    2016-01-01

    Interferon Regulatory Factor (IRF) 6, a member of the IRF family, is essential for epidermal and orofacial embryonic development. Irf6 is strongly expressed in keratinocytes, in which it regulates epidermal proliferation, differentiation, and migration. A recent role for Irf6 in Toll-like receptor 2-dependent chemokine gene expression was also reported in an epithelial cell line. However, a function for Irf6 in innate immune cells was not previously reported. In the present study, we investigated the expression and function of Irf6 in bone marrow-derived neutrophils and macrophages. We show here, using a conditional knockout of Irf6 in lysosymeM expressing cells, that Irf6 is required for resistance to LPS-induced endotoxic shock. In addition, Irf6-deficient bone marrow-derived neutrophils exhibited increased chemotactic index and velocity compared with wild-type cells in vitro. TLR4-specific KC and IL6 secretions were upregulated in Irf6-deficient bone marrow-derived macrophages in vitro. These cells also exhibited an increased level of phosphorylated IkBa. Collectively, our findings suggest a role for Irf6 in the resistance to endotoxic shock due to NFk-B-mediated alteration of cytokine production. PMID:27035130

  15. From System-Wide Differential Gene Expression to Perturbed Regulatory Factors: A Combinatorial Approach.

    PubMed

    Mahajan, Gaurang; Mande, Shekhar C

    2015-01-01

    High-throughput experiments such as microarrays and deep sequencing provide large scale information on the pattern of gene expression, which undergoes extensive remodeling as the cell dynamically responds to varying environmental cues or has its function disrupted under pathological conditions. An important initial step in the systematic analysis and interpretation of genome-scale expression alteration involves identification of a set of perturbed transcriptional regulators whose differential activity can provide a proximate hypothesis to account for these transcriptomic changes. In the present work, we propose an unbiased and logically natural approach to transcription factor enrichment. It involves overlaying a list of experimentally determined differentially expressed genes on a background regulatory network coming from e.g. literature curation or computational motif scanning, and identifying that subset of regulators whose aggregated target set best discriminates between the altered and the unaffected genes. In other words, our methodology entails testing of all possible regulatory subnetworks, rather than just the target sets of individual regulators as is followed in most standard approaches. We have proposed an iterative search method to efficiently find such a combination, and benchmarked it on E. coli microarray and regulatory network data available in the public domain. Comparative analysis carried out on artificially generated differential expression profiles, as well as empirical factor overexpression data for M. tuberculosis, shows that our methodology provides marked improvement in accuracy of regulatory inference relative to the standard method that involves evaluating factor enrichment in an individual manner. PMID:26562430

  16. From System-Wide Differential Gene Expression to Perturbed Regulatory Factors: A Combinatorial Approach.

    PubMed

    Mahajan, Gaurang; Mande, Shekhar C

    2015-01-01

    High-throughput experiments such as microarrays and deep sequencing provide large scale information on the pattern of gene expression, which undergoes extensive remodeling as the cell dynamically responds to varying environmental cues or has its function disrupted under pathological conditions. An important initial step in the systematic analysis and interpretation of genome-scale expression alteration involves identification of a set of perturbed transcriptional regulators whose differential activity can provide a proximate hypothesis to account for these transcriptomic changes. In the present work, we propose an unbiased and logically natural approach to transcription factor enrichment. It involves overlaying a list of experimentally determined differentially expressed genes on a background regulatory network coming from e.g. literature curation or computational motif scanning, and identifying that subset of regulators whose aggregated target set best discriminates between the altered and the unaffected genes. In other words, our methodology entails testing of all possible regulatory subnetworks, rather than just the target sets of individual regulators as is followed in most standard approaches. We have proposed an iterative search method to efficiently find such a combination, and benchmarked it on E. coli microarray and regulatory network data available in the public domain. Comparative analysis carried out on artificially generated differential expression profiles, as well as empirical factor overexpression data for M. tuberculosis, shows that our methodology provides marked improvement in accuracy of regulatory inference relative to the standard method that involves evaluating factor enrichment in an individual manner.

  17. metagene Profiles Analyses Reveal Regulatory Element’s Factor-Specific Recruitment Patterns

    PubMed Central

    Samb, Rawane; Lemaçon, Audrey; Bilodeau, Steve; Droit, Arnaud

    2016-01-01

    ChIP-Sequencing (ChIP-Seq) provides a vast amount of information regarding the localization of proteins across the genome. The aggregation of ChIP-Seq enrichment signal in a metagene plot is an approach commonly used to summarize data complexity and to obtain a high level visual representation of the general occupancy pattern of a protein. Here we present the R package metagene, the graphical interface Imetagene and the companion package similaRpeak. Together, they provide a framework to integrate, summarize and compare the ChIP-Seq enrichment signal from complex experimental designs. Those packages identify and quantify similarities or dissimilarities in patterns between large numbers of ChIP-Seq profiles. We used metagene to investigate the differential occupancy of regulatory factors at noncoding regulatory regions (promoters and enhancers) in relation to transcriptional activity in GM12878 B-lymphocytes. The relationships between occupancy patterns and transcriptional activity suggest two different mechanisms of action for transcriptional control: i) a “gradient effect” where the regulatory factor occupancy levels follow transcription and ii) a “threshold effect” where the regulatory factor occupancy levels max out prior to reaching maximal transcription. metagene, Imetagene and similaRpeak are implemented in R under the Artistic license 2.0 and are available on Bioconductor. PMID:27538250

  18. metagene Profiles Analyses Reveal Regulatory Element's Factor-Specific Recruitment Patterns.

    PubMed

    Joly Beauparlant, Charles; Lamaze, Fabien C; Deschênes, Astrid; Samb, Rawane; Lemaçon, Audrey; Belleau, Pascal; Bilodeau, Steve; Droit, Arnaud

    2016-08-01

    ChIP-Sequencing (ChIP-Seq) provides a vast amount of information regarding the localization of proteins across the genome. The aggregation of ChIP-Seq enrichment signal in a metagene plot is an approach commonly used to summarize data complexity and to obtain a high level visual representation of the general occupancy pattern of a protein. Here we present the R package metagene, the graphical interface Imetagene and the companion package similaRpeak. Together, they provide a framework to integrate, summarize and compare the ChIP-Seq enrichment signal from complex experimental designs. Those packages identify and quantify similarities or dissimilarities in patterns between large numbers of ChIP-Seq profiles. We used metagene to investigate the differential occupancy of regulatory factors at noncoding regulatory regions (promoters and enhancers) in relation to transcriptional activity in GM12878 B-lymphocytes. The relationships between occupancy patterns and transcriptional activity suggest two different mechanisms of action for transcriptional control: i) a "gradient effect" where the regulatory factor occupancy levels follow transcription and ii) a "threshold effect" where the regulatory factor occupancy levels max out prior to reaching maximal transcription. metagene, Imetagene and similaRpeak are implemented in R under the Artistic license 2.0 and are available on Bioconductor. PMID:27538250

  19. Polymyxin Resistance of Pseudomonas aeruginosa phoQ Mutants Is Dependent on Additional Two-Component Regulatory Systems

    PubMed Central

    Gutu, Alina D.; Sgambati, Nicole; Strasbourger, Pnina; Brannon, Mark K.; Jacobs, Michael A.; Haugen, Eric; Kaul, Rajinder K.; Johansen, Helle Krogh; Høiby, Niels

    2013-01-01

    Pseudomonas aeruginosa can develop resistance to polymyxin as a consequence of mutations in the PhoPQ regulatory system, mediated by covalent lipid A modification. Transposon mutagenesis of a polymyxin-resistant phoQ mutant defined 41 novel loci required for resistance, including two regulatory systems, ColRS and CprRS. Deletion of the colRS genes, individually or in tandem, abrogated the polymyxin resistance of a ΔphoQ mutant, as did individual or tandem deletion of cprRS. Individual deletion of colR or colS in a ΔphoQ mutant also suppressed 4-amino-l-arabinose addition to lipid A, consistent with the known role of this modification in polymyxin resistance. Surprisingly, tandem deletion of colRS or cprRS in the ΔphoQ mutant or individual deletion of cprR or cprS failed to suppress 4-amino-l-arabinose addition to lipid A, indicating that this modification alone is not sufficient for PhoPQ-mediated polymyxin resistance in P. aeruginosa. Episomal expression of colRS or cprRS in tandem or of cprR individually complemented the Pm resistance phenotype in the ΔphoQ mutant, while episomal expression of colR, colS, or cprS individually did not. Highly polymyxin-resistant phoQ mutants of P. aeruginosa isolated from polymyxin-treated cystic fibrosis patients harbored mutant alleles of colRS and cprS; when expressed in a ΔphoQ background, these mutant alleles enhanced polymyxin resistance. These results define ColRS and CprRS as two-component systems regulating polymyxin resistance in P. aeruginosa, indicate that addition of 4-amino-l-arabinose to lipid A is not the only PhoPQ-regulated biochemical mechanism required for resistance, and demonstrate that colRS and cprS mutations can contribute to high-level clinical resistance. PMID:23459479

  20. [ASSESSMENT OF EXTREME FACTORS OF SHIFT WORK IN ARCTIC CONDITIONS BY WORKERS WITH DIFFERENT REGULATORY PROCESSES].

    PubMed

    Korneeva, Ya A; Simonova, N N

    2016-01-01

    A man working on a shift basis in the Arctic, every day is under the influence of various extreme factors which are inevitable for oil and gas indudtry. To adapt to shift work employees use various resources of the individual. The purpose of research is the determination of personal resources of shift workers to overcome the adverse factors of the environment in the Arctic. The study involved 191 builder of main gas pipelines, working in shifts in the Tyumen region (the length of the shift 52 days of arrival) at the age of 23 to 59 (mean age 34.9 ± 8.1) years. Methods: psychological testing, questioning, observation, descriptive statistics, discriminant step by step analysis. There was revealed the correlation between the subjective assessment of the majority of adverse climatic factors in the regulatory process "assessment of results"; production factors--regulatory processes such as flexibility, autonomy, simulation, and the general level of self-regulation; social factors are more associated with the severity of such regulatory processes, flexibility and evaluation of results. PMID:27430072

  1. Scientific and regulatory standards for assessing product performance using the similarity factor, f2.

    PubMed

    Stevens, Ruth E; Gray, Vivian; Dorantes, Angelica; Gold, Lynn; Pham, Loan

    2015-03-01

    The similarity factor, f2, measures the sameness of dissolution profiles. The following commentary is an overview of discussions and presentations from a group of industry and US regulatory experts that have integrated the science and regulatory research and practice for assessing product performance, particularly for modified-release (MR) dosage forms, using f2. For a drug development sponsor or applicant with an orally complex dosage formulation, it is critical to understand dissolution methods and the similarity factor and how and/or when to apply it in their NDA, ANDA, or PMA submission. As part of any regulatory submission, it is critical to justify that the product performance has not been impacted by any change in the manufacturing process and/or the delayed and/or prolonged drug release characteristics compared to a similar conventional or another orally complex dosage form. The purposes of this document are (1) to provide a description of appropriate dissolution methods, how is the f2 calculated and how it can be used to justify product performance similarity, or not; (2) to provide an overview of alternative methods available for dissolution profile comparisons, and (3) to illustrate how applying these concepts in a focused way supports approval of submissions and regulatory dossiers and aligns them with on-going science and regulatory initiatives. A case study will be used as an example to demonstrate how dissolution testing and the f2 calculation results can impact regulatory outcomes from an NDA (505(b)(1)), NDA (505(b)(2)), ANDA (505(j)), supplemental NDAs/ANDAs, or PMA perspective.

  2. Characterizing the interplay betwen mulitple levels of organization within bacterial sigma factor regulatory networks

    SciTech Connect

    Yu, Qiu; Nagarajan, Harish; Embree, Mallory; Shieu, Wendy; Abate, Elisa; Juarez, Katy; Cho, Byung-Kwan; Elkins, James G; Nevin, Kelly P.; Barrett, Christian; Lovley, Derek; Palsson, Bernhard O.; Zengler, Karsten

    2013-01-01

    Bacteria contain multiple sigma factors, each targeting diverse, but often overlapping sets of promoters, thereby forming a complex network. The layout and deployment of such a sigma factor network directly impacts global transcriptional regulation and ultimately dictates the phenotype. Here we integrate multi-omic data sets to determine the topology, the operational, and functional states of the sigma factor network in Geobacter sulfurreducens, revealing a unique network topology of interacting sigma factors. Analysis of the operational state of the sigma factor network shows a highly modular structure with sN being the major regulator of energy metabolism. Surprisingly, the functional state of the network during the two most divergent growth conditions is nearly static, with sigma factor binding profiles almost invariant to environmental stimuli. This first comprehensive elucidation of the interplay between different levels of the sigma factor network organization is fundamental to characterize transcriptional regulatory mechanisms in bacteria.

  3. Muscle regulatory factors regulate T1R3 taste receptor expression.

    PubMed

    Kokabu, Shoichiro; Lowery, Jonathan W; Toyono, Takashi; Seta, Yuji; Hitomi, Suzuro; Sato, Tsuyoshi; Enoki, Yuichiro; Okubo, Masahiko; Fukushima, Yosuke; Yoda, Tetsuya

    2015-12-25

    T1R3 is a T1R class of G protein-coupled receptors, composing subunit of the umami taste receptor when complexed with T1R1. T1R3 was originally discovered in gustatory tissue but is now known to be expressed in a wide variety of tissues and cell types such the intestine, pancreatic β-cells, skeletal muscle, and heart. In addition to taste recognition, the T1R1/T1R3 complex functions as an amino acid sensor and has been proposed to be a control mechanism for the secretion of hormones, such as cholecystokinin, insulin, and duodenal HCO3(-) and activates the mammalian rapamycin complex 1 (MTORC1) to inhibit autophagy. T1R3 knockout mice have increased rate of autophagy in the heart, skeletal muscle and liver. Thus, T1R3 has multiple physiological functions and is widely expressed in vivo. However, the exact mechanisms regulating T1R3 expression are largely unknown. Here, we used comparative genomics and functional analyses to characterize the genomic region upstream of the annotated transcriptional start of human T1R3. This revealed that the T1R3 promoter in human and mouse resides in an evolutionary conserved region (ECR). We also identified a repressive element located upstream of the human T1R3 promoter that has relatively high degree of conservation with rhesus macaque. Additionally, the muscle regulatory factors MyoD and Myogenin regulate T1R3 expression and T1R3 expression increases with skeletal muscle differentiation of murine myoblast C2C12 cells. Taken together, our study raises the possibility that MyoD and Myogenin might control skeletal muscle metabolism and homeostasis through the regulation of T1R3 promoter activity. PMID:26545778

  4. Ethylene Response Factors: A Key Regulatory Hub in Hormone and Stress Signaling.

    PubMed

    Müller, Maren; Munné-Bosch, Sergi

    2015-09-01

    Ethylene is essential for many developmental processes and a key mediator of biotic and abiotic stress responses in plants. The ethylene signaling and response pathway includes Ethylene Response Factors (ERFs), which belong to the transcription factor family APETALA2/ERF. It is well known that ERFs regulate molecular response to pathogen attack by binding to sequences containing AGCCGCC motifs (the GCC box), a cis-acting element. However, recent studies suggest that several ERFs also bind to dehydration-responsive elements and act as a key regulatory hub in plant responses to abiotic stresses. Here, we review some of the recent advances in our understanding of the ethylene signaling and response pathway, with emphasis on ERFs and their role in hormone cross talk and redox signaling under abiotic stresses. We conclude that ERFs act as a key regulatory hub, integrating ethylene, abscisic acid, jasmonate, and redox signaling in the plant response to a number of abiotic stresses.

  5. Retinoic acid exerts dual regulatory actions on the expression and nuclear localization of interferon regulatory factor-1.

    PubMed

    Luo, Xin M; Ross, A Catharine

    2006-05-01

    Interferon regulatory factor-1 (IRF-1), a transcription factor and tumor suppressor involved in cell growth regulation and immune responses, has been shown to be induced by all-trans retinoic acid (ATRA). However, the factors controlling the cellular location and activity of IRF-1 are not well understood. In this study, we examined the expression of IRF-1 and its nuclear localization, DNA-binding activity, and target gene expression in human mammary epithelial MCF10A cells, a model of breast epithelial cell differentiation and carcinogenesis. Following initial treatment with ATRA, IRF-1 mRNA and protein were induced within 2 hrs, reached a peak (>30-fold induction) at 8 hrs, and declined afterwards. IRF-1 protein was predominantly cytoplasmic during this treatment. Although a second dose of ATRA or Am580 (a related retinoid selective for retinoic acid receptor-alpha [RARalpha]), given 16 hrs after the first dose, restimulated IRF-1 mRNA and protein levels to a similar level to that obtained by the first dose, IRF-1 was predominantly concentrated in the nucleus after restimulation. ATRA and Am580 also increased nuclear RARalpha, whereas retinoid X receptor-alpha (RXRalpha)--a dimerization partner for RARalpha, was localized to the nucleus upon second exposure to ATRA. However, ATRA and Am580 did not regulate the expression or activation of signal transducer and activator of transcription-1 (STAT-1), a transcription factor capable of inducing the expression of IRF-1, indicating an STAT-1-independent mechanism of regulation by ATRA and Am580. The increase in nuclear IRF-1 after retinoid restimulation was accompanied by enhanced binding to an IRF-E DNA response element, and elevated expression of an IRF-1 target gene, 2',5'-oligoadenylate synthetase-2. The dual effect of retinoids in increasing IRF-1 mRNA and protein and in augmenting the nuclear localization of IRF-1 protein may be essential for maximizing the tumor suppressor activity and the immunosurveillance

  6. Mining 3D genome structure populations identifies major factors governing the stability of regulatory communities

    PubMed Central

    Dai, Chao; Li, Wenyuan; Tjong, Harianto; Hao, Shengli; Zhou, Yonggang; Li, Qingjiao; Chen, Lin; Zhu, Bing; Alber, Frank; Jasmine Zhou, Xianghong

    2016-01-01

    Three-dimensional (3D) genome structures vary from cell to cell even in an isogenic sample. Unlike protein structures, genome structures are highly plastic, posing a significant challenge for structure-function mapping. Here we report an approach to comprehensively identify 3D chromatin clusters that each occurs frequently across a population of genome structures, either deconvoluted from ensemble-averaged Hi-C data or from a collection of single-cell Hi-C data. Applying our method to a population of genome structures (at the macrodomain resolution) of lymphoblastoid cells, we identify an atlas of stable inter-chromosomal chromatin clusters. A large number of these clusters are enriched in binding of specific regulatory factors and are therefore defined as ‘Regulatory Communities.' We reveal two major factors, centromere clustering and transcription factor binding, which significantly stabilize such communities. Finally, we show that the regulatory communities differ substantially from cell to cell, indicating that expression variability could be impacted by genome structures. PMID:27240697

  7. Uncovering MicroRNA and Transcription Factor Mediated Regulatory Networks in Glioblastoma.

    PubMed

    Sun, Jingchun; Gong, Xue; Purow, Benjamin; Zhao, Zhongming

    2012-01-01

    Glioblastoma multiforme (GBM) is the most common and lethal brain tumor in humans. Recent studies revealed that patterns of microRNA (miRNA) expression in GBM tissue samples are different from those in normal brain tissues, suggesting that a number of miRNAs play critical roles in the pathogenesis of GBM. However, little is yet known about which miRNAs play central roles in the pathology of GBM and their regulatory mechanisms of action. To address this issue, in this study, we systematically explored the main regulation format (feed-forward loops, FFLs) consisting of miRNAs, transcription factors (TFs) and their impacting GBM-related genes, and developed a computational approach to construct a miRNA-TF regulatory network. First, we compiled GBM-related miRNAs, GBM-related genes, and known human TFs. We then identified 1,128 3-node FFLs and 805 4-node FFLs with statistical significance. By merging these FFLs together, we constructed a comprehensive GBM-specific miRNA-TF mediated regulatory network. Then, from the network, we extracted a composite GBM-specific regulatory network. To illustrate the GBM-specific regulatory network is promising for identification of critical miRNA components, we specifically examined a Notch signaling pathway subnetwork. Our follow up topological and functional analyses of the subnetwork revealed that six miRNAs (miR-124, miR-137, miR-219-5p, miR-34a, miR-9, and miR-92b) might play important roles in GBM, including some results that are supported by previous studies. In this study, we have developed a computational framework to construct a miRNA-TF regulatory network and generated the first miRNA-TF regulatory network for GBM, providing a valuable resource for further understanding the complex regulatory mechanisms in GBM. The observation of critical miRNAs in the Notch signaling pathway, with partial verification from previous studies, demonstrates that our network-based approach is promising for the identification of new and important

  8. Transcriptional Regulatory Network Analysis of MYB Transcription Factor Family Genes in Rice

    PubMed Central

    Smita, Shuchi; Katiyar, Amit; Chinnusamy, Viswanathan; Pandey, Dev M.; Bansal, Kailash C.

    2015-01-01

    MYB transcription factor (TF) is one of the largest TF families and regulates defense responses to various stresses, hormone signaling as well as many metabolic and developmental processes in plants. Understanding these regulatory hierarchies of gene expression networks in response to developmental and environmental cues is a major challenge due to the complex interactions between the genetic elements. Correlation analyses are useful to unravel co-regulated gene pairs governing biological process as well as identification of new candidate hub genes in response to these complex processes. High throughput expression profiling data are highly useful for construction of co-expression networks. In the present study, we utilized transcriptome data for comprehensive regulatory network studies of MYB TFs by “top-down” and “guide-gene” approaches. More than 50% of OsMYBs were strongly correlated under 50 experimental conditions with 51 hub genes via “top-down” approach. Further, clusters were identified using Markov Clustering (MCL). To maximize the clustering performance, parameter evaluation of the MCL inflation score (I) was performed in terms of enriched GO categories by measuring F-score. Comparison of co-expressed cluster and clads analyzed from phylogenetic analysis signifies their evolutionarily conserved co-regulatory role. We utilized compendium of known interaction and biological role with Gene Ontology enrichment analysis to hypothesize function of coexpressed OsMYBs. In the other part, the transcriptional regulatory network analysis by “guide-gene” approach revealed 40 putative targets of 26 OsMYB TF hubs with high correlation value utilizing 815 microarray data. The putative targets with MYB-binding cis-elements enrichment in their promoter region, functional co-occurrence as well as nuclear localization supports our finding. Specially, enrichment of MYB binding regions involved in drought-inducibility implying their regulatory role in drought

  9. Population genetic and phylogenetic evidence for positive selection on regulatory mutations at the factor VII locus in humans.

    PubMed Central

    Hahn, Matthew W; Rockman, Matthew V; Soranzo, Nicole; Goldstein, David B; Wray, Gregory A

    2004-01-01

    The abundance of cis-regulatory polymorphisms in humans suggests that many may have been important in human evolution, but evidence for their role is relatively rare. Four common polymorphisms in the 5' promoter region of factor VII (F7), a coagulation factor, have been shown to affect its transcription and protein abundance both in vitro and in vivo. Three of these polymorphisms have low-frequency alleles that decrease expression of F7 and may provide protection against myocardial infarction (heart attacks). The fourth polymorphism has a minor allele that increases the level of transcription. To look for evidence of natural selection on the cis-regulatory variants flanking F7, we genotyped three of the polymorphisms in six Old World populations for which we also have data from a group of putatively neutral SNPs. Our population genetic analysis shows evidence for selection within humans; surprisingly, the strongest evidence is due to a large increase in frequency of the high-expression variant in Singaporean Chinese. Further characterization of a Japanese population shows that at least part of the increase in frequency of the high-expression allele is found in other East Asian populations. In addition, to examine interspecific patterns of selection we sequenced the homologous 5' noncoding region in chimpanzees, bonobos, a gorilla, an orangutan, and a baboon. Analysis of these data reveals an excess of fixed differences within transcription factor binding sites along the human lineage. Our results thus further support the hypothesis that regulatory mutations have been important in human evolution. PMID:15238535

  10. Histone deacetylase inhibitor trichostatin A enhances myogenesis by coordinating muscle regulatory factors and myogenic repressors

    SciTech Connect

    Hagiwara, Hiroki; Saito, Fumiaki; Masaki, Toshihiro; Ikeda, Miki; Nakamura-Ohkuma, Ayami; Shimizu, Teruo; Matsumura, Kiichiro

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer We investigated the effect of TSA, one of most potent HDACIs, on myogenesis using the C2C12 skeletal muscle cell line. Black-Right-Pointing-Pointer TSA enhances the expression of myosin heavy chain without affecting DAPC expression. Black-Right-Pointing-Pointer TSA enhances the expression of the early MRFs, Myf5 and MEF2, and suppresses the late MRF, myogenin, after 24 h treatment. Black-Right-Pointing-Pointer TSA enhances the expression of the myogenic repressors, Ids, which inhibit myogenic differentiation. Black-Right-Pointing-Pointer TSA promotes myogenesis by coordinating the expression of MRFs and myogenic repressors. -- Abstract: Histone deacetylase inhibitors (HDACIs) are known to promote skeletal muscle formation. However, their mechanisms that include effects on the expression of major muscle components such as the dystrophin-associated proteins complex (DAPC) or myogenic regulatory factors (MRFs) remain unknown. In this study, we investigated the effects of HDACIs on skeletal muscle formation using the C2C12 cell culture system. C2C12 myoblasts were exposed to trichostatin A (TSA), one of the most potent HDACIs, and differentiation was subsequently induced. We found that TSA enhances the expression of myosin heavy chain without affecting DAPC expression. In addition, TSA increases the expression of the early MRFs, Myf5 and MEF2, whereas it suppresses the expression of the late MRF, myogenin. Interestingly, TSA also enhances the expression of Id1, Id2, and Id3 (Ids). Ids are myogenic repressors that inhibit myogenic differentiation. These findings suggest that TSA promotes gene expression in proliferation and suppresses it in the differentiation stage of muscle formation. Taken together, our data demonstrate that TSA enhances myogenesis by coordinating the expression of MRFs and myogenic repressors.

  11. Haploinsufficiency of interferon regulatory factor 6 alters brain morphology in the mouse.

    PubMed

    Aerts, Andrea; DeVolder, Ian; Weinberg, Seth M; Thedens, Dan; Dunnwald, Martine; Schutte, Brian C; Nopoulos, Peg

    2014-03-01

    Orofacial clefts are among the commonest birth defects. Among many genetic contributors to orofacial clefting, Interferon Regulatory Factor 6 (IRF6) is unique since mutations in this gene cause Van der Woude (VWS), the most common clefting syndrome. Furthermore, variants in IRF6 contribute to increased risk for non-syndromic cleft lip and/or palate (NSCL/P). Our previous work shows that individuals with either VWS or NSCL/P may have cerebral anomalies (larger anterior, smaller posterior regions), and a smaller cerebellum. The objective of this study was to test the hypothesis that disrupting Irf6 in the mouse will result in quantitative brain changes similar to those reported for humans with VWS and NSCL/P. Male mice heterozygous for Irf6 (Irf6(gt1/+); n = 9) and wild-type (Irf6(+/+) ; n = 6) mice at comparable age underwent a 4.7-T MRI scan to obtain quantitative measures of cortical and subcortical brain structures. There was no difference in total brain volume between groups. However, the frontal cortex was enlarged in the Irf6(gt1/+) mice compared to that of wild types (P = 0.028) while the posterior cortex did not differ. In addition, the volume of the cerebellum of Irf6(gt1/+) mice was decreased (P = 0.004). Mice that were heterozygous for Irf6 showed a similar pattern of brain anomalies previously reported in humans with VWS and NSCL/P. These structural differences were present in the absence of overt oral clefts. These results support a role for IRF6 in brain morphometry and provide evidence for a potential genetic link to abnormal brain development in orofacial clefting. PMID:24357509

  12. Regulatory mechanisms underlying sepsis progression in patients with tumor necrosis factor-α genetic variations

    PubMed Central

    LIU, YANGZHOU; HAN, NING; LI, QINCHUAN; LI, ZENGCHUN

    2016-01-01

    The present study aimed to investigate the regulatory mechanisms underlying sepsis progression in patients with tumor necrosis factor (TNF)-α genetic variations. The GSE5760 expression profile data, which was downloaded from the Gene Expression Omnibus database, contained 30 wild-type (WT) and 28 mutation (MUT) samples. Differentially expressed genes (DEGs) between the two types of samples were identified using the Student's t-test, and the corresponding microRNAs (miRNAs) were screened using WebGestalt software. An integrated miRNA-DEG network was constructed using the Cytoscape software, based on the interactions between the DEGs, as identified using the Search Tool for the Retrieval of Interacting Genes/Proteins database, and the correlation between miRNAs and their target genes. Furthermore, Gene Ontology and pathway enrichment analyses were conducted for the DEGs using the Database for Annotation, Visualization and Integrated Discovery and the KEGG Orthology Based Annotation System, respectively. A total of 390 DEGS between the WT and MUT samples, along with 11 -associated miRNAs, were identified. The integrated miRNA-DEG network consisted of 38 DEGs and 11 miRNAs. Within this network, COPS2 was found to be associated with transcriptional functions, while FUS was found to be involved in mRNA metabolic processes. Other DEGs, including FBXW7 and CUL3, were enriched in the ubiquitin-mediated proteolysis pathway. In addition, miR-15 was predicted to target COPS2 and CUL3. The results of the present study suggested that COPS2, FUS, FBXW7 and CUL3 may be associated with sepsis in patients with TNF-α genetic variations. In the progression of sepsis, FBXW7 and CUL3 may participate in the ubiquitin-mediated proteolysis pathway, whereas COPS2 may regulate the phosphorylation and ubiquitination of the FUS protein. Furthermore, COPS2 and CUL3 may be novel targets of miR-15. PMID:27347057

  13. Comparative analysis of the transcription-factor gene regulatory networks of E. coli and S. cerevisiae

    PubMed Central

    Guzmán-Vargas, Lev; Santillán, Moisés

    2008-01-01

    Background The regulatory interactions between transcription factors (TF) and regulated genes (RG) in a species genome can be lumped together in a single directed graph. The TF's and RG's conform the nodes of this graph, while links are drawn whenever a transcription factor regulates a gene's expression. Projections onto TF nodes can be constructed by linking every two nodes regulating a common gene. Similarly, projections onto RG nodes can be made by linking every two regulated genes sharing at least one common regulator. Recent studies of the connectivity pattern in the transcription-factor regulatory network of many organisms have revealed some interesting properties. However, the differences between TF and RG nodes have not been widely explored. Results After analysing the RG and TF projections of the transcription-factor gene regulatory networks of Escherichia coli and Saccharomyces cerevisiae, we found several common characteristic as well as some noticeable differences. To better understand these differences, we compared the properties of the E. coli and S. cerevisiae RG- and TF-projected networks with those of the corresponding projections built from randomized versions of the original bipartite networks. These last results indicate that the observed differences are mostly due to the very different ratios of TF to RG counts of the E. coli and S. cerevisiae bipartite networks, rather than to their having different connectivity patterns. Conclusion Since E. coli is a prokaryotic organism while S. cerevisiae is eukaryotic, there are important differences between them concerning processing of mRNA before translation, DNA packing, amount of junk DNA, and gene regulation. From the results in this paper we conclude that the most important effect such differences have had on the development of the corresponding transcription-factor gene regulatory networks is their very different ratios of TF to RG numbers. This ratio is more than three times larger in S

  14. Role of cellular caspases, nuclear factor-kappa B and interferon regulatory factors in Bluetongue virus infection and cell fate

    PubMed Central

    2010-01-01

    Background Bluetongue virus (BTV) infection causes haemorrhagic disease in ruminants and induces cell death. The pathogenesis in animals and in cell culture has been linked to BTV-induced apoptosis. Results In this report, we investigated BTV-induced apoptosis in cell culture in depth and show that both extrinsic (caspase-8 activation) and intrinsic (caspase-9 activation) pathways play roles in BTV apoptosis. Further, by using chemical inhibitors and knock-out cell lines, we show that these pathways act independently of each other in BTV infected cells. In addition to activation of caspase-8, -9 and executioner caspase-3, we also identified that BTV infection causes the activation of caspase-7, which results in the cleavage of poly (ADP-ribose) polymerase (PARP). BTV-induced cell death appears to be due to apoptosis rather than necrosis, as the HMBG-1 was not translocated from the nucleus. We also examined if NF-κB response is related to BTV-induced apoptosis as in reovirus. Our data suggests that NF-κB response is not linked to the induction of apoptosis. It is controlled by the degradation of only IκBα but not IκBβ, resulting in a rapid transient response during BTV infection. This was supported using an NF-κB dependent luciferase reporter gene assay, which demonstrated early response, that appeared to be suppressed by the late stage of BTV replication. Furthermore, virus titres were higher in the presence of NF-κB inhibitor (SN50), indicating that NF-κB has a role in initiating an antiviral environment. In addition, we show that BTV infection induces the translocation of interferon regulatory factors (IRF-3 and IRF-7) into the nucleus. The induction of IRF responses, when measured by IRF dependent luciferase reporter gene assay, revealed that the IRF responses, like NF-κB response, were also at early stage of infection and mirrored the timing of NF-κB induction. Conclusion BTV triggers a wide range of caspase activities resulting in cell apoptosis

  15. Evolution of DNA specificity in a transcription factor family produced a new gene regulatory module.

    PubMed

    McKeown, Alesia N; Bridgham, Jamie T; Anderson, Dave W; Murphy, Michael N; Ortlund, Eric A; Thornton, Joseph W

    2014-09-25

    Complex gene regulatory networks require transcription factors (TFs) to bind distinct DNA sequences. To understand how novel TF specificity evolves, we combined phylogenetic, biochemical, and biophysical approaches to interrogate how DNA recognition diversified in the steroid hormone receptor (SR) family. After duplication of the ancestral SR, three mutations in one copy radically weakened binding to the ancestral estrogen response element (ERE) and improved binding to a new set of DNA sequences (steroid response elements, SREs). They did so by establishing unfavorable interactions with ERE and abolishing unfavorable interactions with SRE; also required were numerous permissive substitutions, which nonspecifically improved cooperativity and affinity of DNA binding. Our findings indicate that negative determinants of binding play key roles in TFs' DNA selectivity and-with our prior work on the evolution of SR ligand specificity during the same interval-show how a specific new gene regulatory module evolved without interfering with the integrity of the ancestral module. PMID:25259920

  16. Using RSAT to scan genome sequences for transcription factor binding sites and cis-regulatory modules.

    PubMed

    Turatsinze, Jean-Valery; Thomas-Chollier, Morgane; Defrance, Matthieu; van Helden, Jacques

    2008-01-01

    This protocol shows how to detect putative cis-regulatory elements and regions enriched in such elements with the regulatory sequence analysis tools (RSAT) web server (http://rsat.ulb.ac.be/rsat/). The approach applies to known transcription factors, whose binding specificity is represented by position-specific scoring matrices, using the program matrix-scan. The detection of individual binding sites is known to return many false predictions. However, results can be strongly improved by estimating P value, and by searching for combinations of sites (homotypic and heterotypic models). We illustrate the detection of sites and enriched regions with a study case, the upstream sequence of the Drosophila melanogaster gene even-skipped. This protocol is also tested on random control sequences to evaluate the reliability of the predictions. Each task requires a few minutes of computation time on the server. The complete protocol can be executed in about one hour.

  17. PcFKH1, a novel regulatory factor from the forkhead family, controls the biosynthesis of penicillin in Penicillium chrysogenum.

    PubMed

    Domínguez-Santos, Rebeca; García-Estrada, Carlos; Kosalková, Katarina; Prieto, Carlos; Santamarta, Irene; Martín, Juan-Francisco

    2015-08-01

    Penicillin biosynthesis in Penicillium chrysogenum (re-identified as Penicillium rubens) is a good example of a biological process subjected to complex global regulatory networks and serves as a model to study fungal secondary metabolism. The winged-helix family of transcription factors recently described, which includes the forkhead type of proteins, is a key type of regulatory proteins involved in this process. In yeasts and humans, forkhead transcription factors are involved in different processes (cell cycle regulation, cell death control, pre-mRNA processing and morphogenesis); one member of this family of proteins has been identified in the P. chrysogenum genome (Pc18g00430). In this work, we have characterized this novel transcription factor (named PcFKH1) by generating knock-down mutants and overexpression strains. Results clearly indicate that PcFKH1 positively controls antibiotic biosynthesis through the specific interaction with the promoter region of the penDE gene, thus regulating penDE mRNA levels. PcFKH1 also binds to the pcbC promoter, but with low affinity. In addition, it also controls other ancillary genes of the penicillin biosynthetic process, such as phlA (encoding phenylacetyl CoA ligase) and ppt (encoding phosphopantetheinyl transferase). PcFKH1 also plays a role in conidiation and spore pigmentation, but it does not seem to be involved in hyphal morphology or cell division in the improved laboratory reference strain Wisconsin 54-1255. A genome-wide analysis of processes putatively coregulated by PcFKH1 and PcRFX1 (another winged-helix transcription factor) in P. chrysogenum provided evidence of the global effect of these transcription factors in P. chrysogenum metabolism.

  18. Tumor Suppressor Interferon-Regulatory Factor 1 Counteracts the Germinal Center Reaction Driven by a Cancer-Associated Gammaherpesvirus

    PubMed Central

    Mboko, Wadzanai P.; Olteanu, Horatiu; Ray, Avijit; Xin, Gang; Darrah, Eric J.; Kumar, Suresh N.; Kulinski, Joseph M.; Cui, Weiguo; Dittel, Bonnie N.; Gauld, Stephen B.

    2015-01-01

    ABSTRACT Gammaherpesviruses are ubiquitous pathogens that are associated with the development of B cell lymphomas. Gammaherpesviruses employ multiple mechanisms to transiently stimulate a broad, polyclonal germinal center reaction, an inherently mutagenic stage of B cell differentiation that is thought to be the primary target of malignant transformation in virus-driven lymphomagenesis. We found that this gammaherpesvirus-driven germinal center expansion was exaggerated and lost its transient nature in the absence of interferon-regulatory factor 1 (IRF-1), a transcription factor with antiviral and tumor suppressor functions. Uncontrolled and persistent expansion of germinal center B cells led to pathological changes in the spleens of chronically infected IRF-1-deficient animals. Additionally, we found decreased IRF-1 expression in cases of human posttransplant lymphoproliferative disorder, a malignant condition associated with gammaherpesvirus infection. The results of our study define an unappreciated role for IRF-1 in B cell biology and provide insight into the potential mechanism of gammaherpesvirus-driven lymphomagenesis. IMPORTANCE Gammaherpesviruses establish lifelong infection in most adults and are associated with B cell lymphomas. While the infection is asymptomatic in many hosts, it is critical to identify individuals who may be at an increased risk of virus-induced cancer. Such identification is currently impossible, as the host risk factors that predispose individuals toward viral lymphomagenesis are poorly understood. The current study identifies interferon-regulatory factor 1 (IRF-1) to be one of such candidate host factors. Specifically, we found that IRF-1 enforces long-term suppression of an inherently mutagenic stage of B cell differentiation that gammaherpesviruses are thought to target for transformation. Correspondingly, in the absence of IRF-1, chronic gammaherpesvirus infection induced pathological changes in the spleens of infected animals

  19. Dysregulated T helper cell differentiation in the absence of interferon regulatory factor 4.

    PubMed

    Lohoff, Michael; Mittrücker, Hans-Willi; Prechtl, Stefan; Bischof, Susi; Sommer, Frank; Kock, Sonja; Ferrick, David A; Duncan, Gordon S; Gessner, Andre; Mak, Tak W

    2002-09-01

    Certain IFN regulatory factor (IRF) transcription factors indirectly influence T helper (Th) cell differentiation by regulating the production of IL-12. Here, we show that IRF4 directly regulates Th cell differentiation in vitro and in vivo during murine leishmaniasis. In the absence of IRF4, IL-12-induced Th1 cell differentiation was compromised, while IL-4 failed to induce Th2 cell differentiation. Instead, IL-4 tended to induce Th1 cells, defined by production of IFN-gamma and TNF. Although early IL-4 signaling was normal in IRF4(-/-) Th cells, the protein GATA-3, a transcription factor critical for Th2 development, was not up-regulated following IL-4 treatment. Retroviral overexpression of GATA-3 rescued Th2 differentiation. Therefore, IRF4 deficiency manifests itself as severely dysregulated Th cell differentiation.

  20. Transcription factor abundance controlled by an auto-regulatory mechanism involving a transcription start site switch

    PubMed Central

    Ngondo, Richard Patryk; Carbon, Philippe

    2014-01-01

    A transcriptional feedback loop is the simplest and most direct means for a transcription factor to provide an increased stability of gene expression. In this work performed in human cells, we reveal a new negative auto-regulatory mechanism involving an alternative transcription start site (TSS) usage. Using the activating transcription factor ZNF143 as a model, we show that the ZNF143 low-affinity binding sites, located downstream of its canonical TSS, play the role of protein sensors to induce the up- or down-regulation of ZNF143 gene expression. We uncovered that the TSS switch that mediates this regulation implies the differential expression of two transcripts with an opposite protein production ability due to their different 5′ untranslated regions. Moreover, our analysis of the ENCODE data suggests that this mechanism could be used by other transcription factors to rapidly respond to their own aberrant expression level. PMID:24234445

  1. Selection of terrestrial transfer factors for radioecological assessment models and regulatory guides

    SciTech Connect

    Ng, Y.C.; Hoffman, F.O.

    1983-01-01

    A parameter value for a radioecological assessment model is not a single value but a distribution of values about a central value. The sources that contribute to the variability of transfer factors to predict foodchain transport of radionuclides are enumerated. Knowledge of these sources, judgement in interpreting the available data, consideration of collateral information, and established criteria that specify the desired level of conservatism in the resulting predictions are essential elements when selecting appropriate parameter values for radioecological assessment models and regulatory guides. 39 references, 4 figures, 5 tables.

  2. Mining regulatory network connections by ranking transcription factor target genes using time series expression data.

    PubMed

    Honkela, Antti; Rattray, Magnus; Lawrence, Neil D

    2013-01-01

    Reverse engineering the gene regulatory network is challenging because the amount of available data is very limited compared to the complexity of the underlying network. We present a technique addressing this problem through focussing on a more limited problem: inferring direct targets of a transcription factor from short expression time series. The method is based on combining Gaussian process priors and ordinary differential equation models allowing inference on limited potentially unevenly sampled data. The method is implemented as an R/Bioconductor package, and it is demonstrated by ranking candidate targets of the p53 tumour suppressor.

  3. Direct lineage reprogramming via pioneer factors; a detour through developmental gene regulatory networks.

    PubMed

    Morris, Samantha A

    2016-08-01

    Although many approaches have been employed to generate defined fate in vitro, the resultant cells often appear developmentally immature or incompletely specified, limiting their utility. Growing evidence suggests that current methods of direct lineage conversion may rely on the transition through a developmental intermediate. Here, I hypothesize that complete conversion between cell fates is more probable and feasible via reversion to a developmentally immature state. I posit that this is due to the role of pioneer transcription factors in engaging silent, unmarked chromatin and activating hierarchical gene regulatory networks responsible for embryonic patterning. Understanding these developmental contexts will be essential for the precise engineering of cell identity. PMID:27486230

  4. How microRNA and transcription factor co-regulatory networks affect osteosarcoma cell proliferation.

    PubMed

    Poos, Kathrin; Smida, Jan; Nathrath, Michaela; Maugg, Doris; Baumhoer, Daniel; Korsching, Eberhard

    2013-01-01

    Osteosarcomas (OS) are complex bone tumors with various genomic alterations. These alterations affect the expression and function of several genes due to drastic changes in the underlying gene regulatory network. However, we know little about critical gene regulators and their functional consequences on the pathogenesis of OS. Therefore, we aimed to determine microRNA and transcription factor (TF) co-regulatory networks in OS cell proliferation. Cell proliferation is an essential part in the pathogenesis of OS and deeper understanding of its regulation might help to identify potential therapeutic targets. Based on expression data of OS cell lines divided according to their proliferative activity, we obtained 12 proliferation-related microRNAs and corresponding target genes. Therewith, microRNA and TF co-regulatory networks were generated and analyzed regarding their structure and functional influence. We identified key co-regulators comprising the microRNAs miR-9-5p, miR-138, and miR-214 and the TFs SP1 and MYC in the derived networks. These regulators are implicated in NFKB- and RB1-signaling and focal adhesion processes based on their common or interacting target genes (e.g., CDK6, CTNNB1, E2F4, HES1, ITGA6, NFKB1, NOTCH1, and SIN3A). Thus, we proposed a model of OS cell proliferation which is primarily co-regulated through the interactions of the mentioned microRNA and TF combinations. This study illustrates the benefit of systems biological approaches in the analysis of complex diseases. We integrated experimental data with publicly available information to unravel the coordinated (post)-transcriptional control of microRNAs and TFs to identify potential therapeutic targets in OS. The resulting microRNA and TF co-regulatory networks are publicly available for further exploration to generate or evaluate own hypotheses of the pathogenesis of OS (http://www.complex-systems.uni-muenster.de/co_networks.html).

  5. Identification of High-Impact cis-Regulatory Mutations Using Transcription Factor Specific Random Forest Models

    PubMed Central

    Svetlichnyy, Dmitry; Imrichova, Hana; Fiers, Mark; Kalender Atak, Zeynep; Aerts, Stein

    2015-01-01

    Cancer genomes contain vast amounts of somatic mutations, many of which are passenger mutations not involved in oncogenesis. Whereas driver mutations in protein-coding genes can be distinguished from passenger mutations based on their recurrence, non-coding mutations are usually not recurrent at the same position. Therefore, it is still unclear how to identify cis-regulatory driver mutations, particularly when chromatin data from the same patient is not available, thus relying only on sequence and expression information. Here we use machine-learning methods to predict functional regulatory regions using sequence information alone, and compare the predicted activity of the mutated region with the reference sequence. This way we define the Predicted Regulatory Impact of a Mutation in an Enhancer (PRIME). We find that the recently identified driver mutation in the TAL1 enhancer has a high PRIME score, representing a “gain-of-target” for MYB, whereas the highly recurrent TERT promoter mutation has a surprisingly low PRIME score. We trained Random Forest models for 45 cancer-related transcription factors, and used these to score variations in the HeLa genome and somatic mutations across more than five hundred cancer genomes. Each model predicts only a small fraction of non-coding mutations with a potential impact on the function of the encompassing regulatory region. Nevertheless, as these few candidate driver mutations are often linked to gains in chromatin activity and gene expression, they may contribute to the oncogenic program by altering the expression levels of specific oncogenes and tumor suppressor genes. PMID:26562774

  6. A Minimal Regulatory Network of Extrinsic and Intrinsic Factors Recovers Observed Patterns of CD4+ T Cell Differentiation and Plasticity

    PubMed Central

    Martinez-Sanchez, Mariana Esther; Mendoza, Luis; Villarreal, Carlos; Alvarez-Buylla, Elena R.

    2015-01-01

    CD4+ T cells orchestrate the adaptive immune response in vertebrates. While both experimental and modeling work has been conducted to understand the molecular genetic mechanisms involved in CD4+ T cell responses and fate attainment, the dynamic role of intrinsic (produced by CD4+ T lymphocytes) versus extrinsic (produced by other cells) components remains unclear, and the mechanistic and dynamic understanding of the plastic responses of these cells remains incomplete. In this work, we studied a regulatory network for the core transcription factors involved in CD4+ T cell-fate attainment. We first show that this core is not sufficient to recover common CD4+ T phenotypes. We thus postulate a minimal Boolean regulatory network model derived from a larger and more comprehensive network that is based on experimental data. The minimal network integrates transcriptional regulation, signaling pathways and the micro-environment. This network model recovers reported configurations of most of the characterized cell types (Th0, Th1, Th2, Th17, Tfh, Th9, iTreg, and Foxp3-independent T regulatory cells). This transcriptional-signaling regulatory network is robust and recovers mutant configurations that have been reported experimentally. Additionally, this model recovers many of the plasticity patterns documented for different T CD4+ cell types, as summarized in a cell-fate map. We tested the effects of various micro-environments and transient perturbations on such transitions among CD4+ T cell types. Interestingly, most cell-fate transitions were induced by transient activations, with the opposite behavior associated with transient inhibitions. Finally, we used a novel methodology was used to establish that T-bet, TGF-β and suppressors of cytokine signaling proteins are keys to recovering observed CD4+ T cell plastic responses. In conclusion, the observed CD4+ T cell-types and transition patterns emerge from the feedback between the intrinsic or intracellular regulatory core

  7. Discovery of transcription factors and regulatory regions driving in vivo tumor development by ATAC-seq and FAIRE-seq open chromatin profiling.

    PubMed

    Davie, Kristofer; Jacobs, Jelle; Atkins, Mardelle; Potier, Delphine; Christiaens, Valerie; Halder, Georg; Aerts, Stein

    2015-02-01

    Genomic enhancers regulate spatio-temporal gene expression by recruiting specific combinations of transcription factors (TFs). When TFs are bound to active regulatory regions, they displace canonical nucleosomes, making these regions biochemically detectable as nucleosome-depleted regions or accessible/open chromatin. Here we ask whether open chromatin profiling can be used to identify the entire repertoire of active promoters and enhancers underlying tissue-specific gene expression during normal development and oncogenesis in vivo. To this end, we first compare two different approaches to detect open chromatin in vivo using the Drosophila eye primordium as a model system: FAIRE-seq, based on physical separation of open versus closed chromatin; and ATAC-seq, based on preferential integration of a transposon into open chromatin. We find that both methods reproducibly capture the tissue-specific chromatin activity of regulatory regions, including promoters, enhancers, and insulators. Using both techniques, we screened for regulatory regions that become ectopically active during Ras-dependent oncogenesis, and identified 3778 regions that become (over-)activated during tumor development. Next, we applied motif discovery to search for candidate transcription factors that could bind these regions and identified AP-1 and Stat92E as key regulators. We validated the importance of Stat92E in the development of the tumors by introducing a loss of function Stat92E mutant, which was sufficient to rescue the tumor phenotype. Additionally we tested if the predicted Stat92E responsive regulatory regions are genuine, using ectopic induction of JAK/STAT signaling in developing eye discs, and observed that similar chromatin changes indeed occurred. Finally, we determine that these are functionally significant regulatory changes, as nearby target genes are up- or down-regulated. In conclusion, we show that FAIRE-seq and ATAC-seq based open chromatin profiling, combined with motif

  8. Modulation of Mitochondrial Antiviral Signaling by Human Herpesvirus 8 Interferon Regulatory Factor 1

    PubMed Central

    Hwang, Keun Young

    2015-01-01

    ABSTRACT Mitochondrial lipid raft-like microdomains, experimentally also termed mitochondrial detergent-resistant membrane fractions (mDRM), play a role as platforms for recruiting signaling molecules involved in antiviral responses such as apoptosis and innate immunity. Viruses can modulate mitochondrial functions for their own survival and replication. However, viral regulation of the antiviral responses via mDRM remains incompletely understood. Here, we report that human herpesvirus 8 (HHV-8) gene product viral interferon regulatory factor 1 (vIRF-1) is targeted to mDRM during virus replication and negatively regulates the mitochondrial antiviral signaling protein (MAVS)-mediated antiviral responses. The N-terminal region of vIRF-1 interacts directly with membrane lipids, including cardiolipin. In addition, a GxRP motif within the N terminus of vIRF-1, conserved in the mDRM-targeting region of mitochondrial proteins, including PTEN-induced putative kinase 1 (PINK1) and MAVS, was found to be important for vIRF-1 association with mitochondria. Furthermore, MAVS, which has the potential to promote vIRF-1 targeting to mDRM possibly by inducing cardiolipin exposure on the outer membrane of mitochondria, interacts with vIRF-1, which, in turn, inhibits MAVS-mediated antiviral signaling. Consistent with these results, vIRF-1 targeting to mDRM contributes to promotion of HHV-8 productive replication and inhibition of associated apoptosis. Combined, our results suggest novel molecular mechanisms for negative-feedback regulation of MAVS by vIRF-1 during virus replication. IMPORTANCE Successful virus replication is in large part achieved by the ability of viruses to counteract apoptosis and innate immune responses elicited by infection of host cells. Recently, mitochondria have emerged to play a central role in antiviral signaling. In particular, mitochondrial lipid raft-like microdomains appear to function as platforms in cell apoptosis signaling. However, viral regulation

  9. Ciliary dyslexia candidate genes DYX1C1 and DCDC2 are regulated by Regulatory Factor X (RFX) transcription factors through X-box promoter motifs

    PubMed Central

    Tammimies, Kristiina; Bieder, Andrea; Lauter, Gilbert; Sugiaman-Trapman, Debora; Torchet, Rachel; Hokkanen, Marie-Estelle; Burghoorn, Jan; Castrén, Eero; Kere, Juha; Tapia-Páez, Isabel; Swoboda, Peter

    2016-01-01

    DYX1C1, DCDC2, and KIAA0319 are three of the most replicated dyslexia candidate genes (DCGs). Recently, these DCGs were implicated in functions at the cilium. Here, we investigate the regulation of these DCGs by Regulatory Factor X transcription factors (RFX TFs), a gene family known for transcriptionally regulating ciliary genes. We identify conserved X-box motifs in the promoter regions of DYX1C1, DCDC2, and KIAA0319 and demonstrate their functionality, as well as the ability to recruit RFX TFs using reporter gene and electrophoretic mobility shift assays. Furthermore, we uncover a complex regulation pattern between RFX1, RFX2, and RFX3 and their significant effect on modifying the endogenous expression of DYX1C1 and DCDC2 in a human retinal pigmented epithelial cell line immortalized with hTERT (hTERT-RPE1). In addition, induction of ciliogenesis increases the expression of RFX TFs and DCGs. At the protein level, we show that endogenous DYX1C1 localizes to the base of the cilium, whereas DCDC2 localizes along the entire axoneme of the cilium, thereby validating earlier localization studies using overexpression models. Our results corroborate the emerging role of DCGs in ciliary function and characterize functional noncoding elements, X-box promoter motifs, in DCG promoter regions, which thus can be targeted for mutation screening in dyslexia and ciliopathies associated with these genes.—Tammimies, K., Bieder, A., Lauter, G., Sugiaman-Trapman, D., Torchet, R., Hokkanen, M.-E., Burghoorn, J., Castrén, E., Kere, J., Tapia-Páez, I., Swoboda, P. Ciliary dyslexia candidate genes DYX1C1 and DCDC2 are regulated by Regulatory Factor (RF) X transcription factors through X-box promoter motifs. PMID:27451412

  10. Transcription factor MITF and remodeller BRG1 define chromatin organisation at regulatory elements in melanoma cells.

    PubMed

    Laurette, Patrick; Strub, Thomas; Koludrovic, Dana; Keime, Céline; Le Gras, Stéphanie; Seberg, Hannah; Van Otterloo, Eric; Imrichova, Hana; Siddaway, Robert; Aerts, Stein; Cornell, Robert A; Mengus, Gabrielle; Davidson, Irwin

    2015-03-24

    Microphthalmia-associated transcription factor (MITF) is the master regulator of the melanocyte lineage. To understand how MITF regulates transcription, we used tandem affinity purification and mass spectrometry to define a comprehensive MITF interactome identifying novel cofactors involved in transcription, DNA replication and repair, and chromatin organisation. We show that MITF interacts with a PBAF chromatin remodelling complex comprising BRG1 and CHD7. BRG1 is essential for melanoma cell proliferation in vitro and for normal melanocyte development in vivo. MITF and SOX10 actively recruit BRG1 to a set of MITF-associated regulatory elements (MAREs) at active enhancers. Combinations of MITF, SOX10, TFAP2A, and YY1 bind between two BRG1-occupied nucleosomes thus defining both a signature of transcription factors essential for the melanocyte lineage and a specific chromatin organisation of the regulatory elements they occupy. BRG1 also regulates the dynamics of MITF genomic occupancy. MITF-BRG1 interplay thus plays an essential role in transcription regulation in melanoma.

  11. Identification and functional analysis of interferon regulatory factor 3 in Lateolabrax maculatus.

    PubMed

    Chen, Xiao-Wu; Wei, Qun; Wang, Zhi-Peng; Wang, Chun-Lei; Bi, Yan-Hui; Gu, Yi-Feng

    2016-10-01

    The interferon (IFN) regulatory factor 3 (IRF3) is a member of the IFN regulatory transcription factor family, which binds to the IFN-stimulated response element (ISRE) within the promoter of IFN genes and IFN-stimulated genes. In this study, the IRF3 cDNA of sea perch Lateolabrax maculatus (SpIRF3) was identified, which contained 1781 bp with an open reading frame of 1398 bp that coded a 465 amino acid protein. The SpIRF3 protein shared conserved characterizations with its homologues and displayed the conserved DNA-binding domain, IRF association domain, serine-rich C-terminal domain, and tryptophan residue cluster. Phylogenetic analysis illustrated that SpIRF3 belonged to the IRF3 subfamily. Subcellular localization analysis showed that SpIRF3 mainly resided in the cytoplasm without stimuli but translocated into nuclei in the presence of poly I:C. Real-time PCR data indicated that SpIRF3 was transcriptionally up-regulated by poly I:C stimulation in various organs. Moreover, reporter assay revealed that SpIRF3 functioned as a modulator in triggering the IFN response by inducing the activity of IFN and ISRE-containing promoter. These data revealed that SpIRF3 was a potential molecule in the IFN immune defense system against viral infection. PMID:27181713

  12. Transcription factor MITF and remodeller BRG1 define chromatin organisation at regulatory elements in melanoma cells

    PubMed Central

    Laurette, Patrick; Strub, Thomas; Koludrovic, Dana; Keime, Céline; Le Gras, Stéphanie; Seberg, Hannah; Van Otterloo, Eric; Imrichova, Hana; Siddaway, Robert; Aerts, Stein; Cornell, Robert A; Mengus, Gabrielle; Davidson, Irwin

    2015-01-01

    Microphthalmia-associated transcription factor (MITF) is the master regulator of the melanocyte lineage. To understand how MITF regulates transcription, we used tandem affinity purification and mass spectrometry to define a comprehensive MITF interactome identifying novel cofactors involved in transcription, DNA replication and repair, and chromatin organisation. We show that MITF interacts with a PBAF chromatin remodelling complex comprising BRG1 and CHD7. BRG1 is essential for melanoma cell proliferation in vitro and for normal melanocyte development in vivo. MITF and SOX10 actively recruit BRG1 to a set of MITF-associated regulatory elements (MAREs) at active enhancers. Combinations of MITF, SOX10, TFAP2A, and YY1 bind between two BRG1-occupied nucleosomes thus defining both a signature of transcription factors essential for the melanocyte lineage and a specific chromatin organisation of the regulatory elements they occupy. BRG1 also regulates the dynamics of MITF genomic occupancy. MITF-BRG1 interplay thus plays an essential role in transcription regulation in melanoma. DOI: http://dx.doi.org/10.7554/eLife.06857.001 PMID:25803486

  13. [Butanol as a regulatory factor of ompC gene expression in E. coli cells].

    PubMed

    Seregina, T A; Shakulov, R S; Mironov, A S

    2012-11-01

    The influence of butanol on the expression of ompC gene encoding synthesis of OmpC porin in the MG 1655 strain of E. coli and butanol-tolerant mutant ButR was studied. It was shown that in the case of wild bacteria, the addition of butanol to the growth medium results in an increased level of ompC transcription. However, OmpC porin is not detected in the membrane fraction of cells. ButR mutant exhibits a higher level of ompC gene expression. A direct correlation is observed between the level of OmpC porin expression and its content in the membrane fraction of ButR mutant cells. In the regulatory region of the ompC gene of the ButR mutant, three nucleotide substitutions located in the binding sites of OmpR and CpxR activator proteins were identified. It was shown that mutations in the regulatory region of the ompC gene in the ButR mutant are responsible for the decreased level of OmpC porin expression under normal growth conditions. However, these mutations lead to an increased level of OmpC porin synthesis in the presence of butanol. These data suggest an additional mechanism of ompC gene regulation with the participation of butanol as a positive transcription effector.

  14. Molecular cloning and expression of an additional epidermal growth factor receptor-related gene.

    PubMed Central

    Plowman, G D; Whitney, G S; Neubauer, M G; Green, J M; McDonald, V L; Todaro, G J; Shoyab, M

    1990-01-01

    Epidermal growth factor (EGF), transforming growth factor alpha (TGF-alpha), and amphiregulin are structurally and functionally related growth regulatory proteins. These secreted polypeptides all bind to the 170-kDa cell-surface EGF receptor, activating its intrinsic kinase activity. However, amphiregulin exhibits different activities than EGF and TGF-alpha in a number of biological assays. Amphiregulin only partially competes with EGF for binding EGF receptor, and amphiregulin does not induce anchorage-independent growth of normal rat kidney cells (NRK) in the presence of TGF-beta. Amphiregulin also appears to abrogate the stimulatory effect of TGF-alpha on the growth of several aggressive epithelial carcinomas that overexpress EGF receptor. These findings suggest that amphiregulin may interact with a separate receptor in certain cell types. Here we report the cloning of another member of the human EGF receptor (HER) family of receptor tyrosine kinases, which we have named "HER3/ERRB3." The cDNA was isolated from a human carcinoma cell line, and its 6-kilobase transcript was identified in various human tissues. We have generated peptide-specific antisera that recognizes the 160-kDa HER3 protein when transiently expressed in COS cells. These reagents will allow us to determine whether HER3 binds amphiregulin or other growth regulatory proteins and what role HER3 protein plays in the regulation of cell growth. Images PMID:2164210

  15. Functional analysis of microRNA and transcription factor synergistic regulatory network based on identifying regulatory motifs in non-small cell lung cancer

    PubMed Central

    2013-01-01

    Background Lung cancer, especially non-small cell lung cancer, is a leading cause of malignant tumor death worldwide. Understanding the mechanisms employed by the main regulators, such as microRNAs (miRNAs) and transcription factors (TFs), still remains elusive. The patterns of their cooperation and biological functions in the synergistic regulatory network have rarely been studied. Results Here, we describe the first miRNA-TF synergistic regulation network in human lung cancer. We identified important regulators (MYC, NFKB1, miR-590, and miR-570) and significant miRNA-TF synergistic regulatory motifs by random simulations. The two most significant motifs were the co-regulation of miRNAs and TFs, and TF-mediated cascade regulation. We also developed an algorithm to uncover the biological functions of the human lung cancer miRNA-TF synergistic regulatory network (regulation of apoptosis, cellular protein metabolic process, and cell cycle), and the specific functions of each miRNA-TF synergistic subnetwork. We found that the miR-17 family exerted important effects in the regulation of non-small cell lung cancer, such as in proliferation and cell cycle regulation by targeting the retinoblastoma protein (RB1) and forming a feed forward loop with the E2F1 TF. We proposed a model for the miR-17 family, E2F1, and RB1 to demonstrate their potential roles in the occurrence and development of non-small cell lung cancer. Conclusions This work will provide a framework for constructing miRNA-TF synergistic regulatory networks, function analysis in diseases, and identification of the main regulators and regulatory motifs, which will be useful for understanding the putative regulatory motifs involving miRNAs and TFs, and for predicting new targets for cancer studies. PMID:24200043

  16. 34 CFR 377.22 - What additional factors does the Secretary consider in making grants?

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... PROJECTS TO INCREASE CLIENT CHOICE PROGRAM How Does the Secretary Make an Award? § 377.22 What additional factors does the Secretary consider in making grants? In addition to the criteria in § 377.21, the... strategies to increase client choice, in order to ensure that a variety of approaches are demonstrated...

  17. A Consensus Network of Gene Regulatory Factors in the Human Frontal Lobe.

    PubMed

    Berto, Stefano; Perdomo-Sabogal, Alvaro; Gerighausen, Daniel; Qin, Jing; Nowick, Katja

    2016-01-01

    Cognitive abilities, such as memory, learning, language, problem solving, and planning, involve the frontal lobe and other brain areas. Not much is known yet about the molecular basis of cognitive abilities, but it seems clear that cognitive abilities are determined by the interplay of many genes. One approach for analyzing the genetic networks involved in cognitive functions is to study the coexpression networks of genes with known importance for proper cognitive functions, such as genes that have been associated with cognitive disorders like intellectual disability (ID) or autism spectrum disorders (ASD). Because many of these genes are gene regulatory factors (GRFs) we aimed to provide insights into the gene regulatory networks active in the human frontal lobe. Using genome wide human frontal lobe expression data from 10 independent data sets, we first derived 10 individual coexpression networks for all GRFs including their potential target genes. We observed a high level of variability among these 10 independently derived networks, pointing out that relying on results from a single study can only provide limited biological insights. To instead focus on the most confident information from these 10 networks we developed a method for integrating such independently derived networks into a consensus network. This consensus network revealed robust GRF interactions that are conserved across the frontal lobes of different healthy human individuals. Within this network, we detected a strong central module that is enriched for 166 GRFs known to be involved in brain development and/or cognitive disorders. Interestingly, several hubs of the consensus network encode for GRFs that have not yet been associated with brain functions. Their central role in the network suggests them as excellent new candidates for playing an essential role in the regulatory network of the human frontal lobe, which should be investigated in future studies. PMID:27014338

  18. A Consensus Network of Gene Regulatory Factors in the Human Frontal Lobe

    PubMed Central

    Berto, Stefano; Perdomo-Sabogal, Alvaro; Gerighausen, Daniel; Qin, Jing; Nowick, Katja

    2016-01-01

    Cognitive abilities, such as memory, learning, language, problem solving, and planning, involve the frontal lobe and other brain areas. Not much is known yet about the molecular basis of cognitive abilities, but it seems clear that cognitive abilities are determined by the interplay of many genes. One approach for analyzing the genetic networks involved in cognitive functions is to study the coexpression networks of genes with known importance for proper cognitive functions, such as genes that have been associated with cognitive disorders like intellectual disability (ID) or autism spectrum disorders (ASD). Because many of these genes are gene regulatory factors (GRFs) we aimed to provide insights into the gene regulatory networks active in the human frontal lobe. Using genome wide human frontal lobe expression data from 10 independent data sets, we first derived 10 individual coexpression networks for all GRFs including their potential target genes. We observed a high level of variability among these 10 independently derived networks, pointing out that relying on results from a single study can only provide limited biological insights. To instead focus on the most confident information from these 10 networks we developed a method for integrating such independently derived networks into a consensus network. This consensus network revealed robust GRF interactions that are conserved across the frontal lobes of different healthy human individuals. Within this network, we detected a strong central module that is enriched for 166 GRFs known to be involved in brain development and/or cognitive disorders. Interestingly, several hubs of the consensus network encode for GRFs that have not yet been associated with brain functions. Their central role in the network suggests them as excellent new candidates for playing an essential role in the regulatory network of the human frontal lobe, which should be investigated in future studies. PMID:27014338

  19. Different papillomaviruses have different repertoires of transcription factor binding sites: convergence and divergence in the upstream regulatory region

    PubMed Central

    García-Vallvé, Santiago; Iglesias-Rozas, José R; Alonso, Ángel; Bravo, Ignacio G

    2006-01-01

    Background Papillomaviruses (PVs) infect stratified squamous epithelia in warm-blooded vertebrates and have undergone a complex evolutionary process. The control of the expression of the early ORFs in PVs depends on the binding of cellular and viral transcription factors to the upstream regulatory region (URR) of the virus. It is believed that there is a core of transcription factor binding sites (TFBS) common to all PVs, with additional individual differences, although most of the available information focuses only on a handful of viruses. Results We have studied the URR of sixty-one PVs, covering twenty different hosts. We have predicted the TFBS present in the URR and analysed these results by principal component analysis and genetic algorithms. The number and nature of TFBS in the URR might be much broader than thus far described, and different PVs have different repertoires of TFBS. Conclusion There are common fingerprints in the URR in PVs that infect primates, although the ancestors of these viruses diverged a long time ago. Additionally, there are obvious differences between the URR of alpha and beta PVs, despite these PVs infect similar histological cell types in the same host, i.e. human. A thorough analysis of the TFBS in the URR might provide crucial information about the differential biology of cancer-associated PVs. PMID:16526953

  20. Modulation of interferon regulatory factor 5 activities by the Kaposi sarcoma-associated herpesvirus-encoded viral interferon regulatory factor 3 contributes to immune evasion and lytic induction.

    PubMed

    Bi, Xiaohui; Yang, Lisong; Mancl, Margo E; Barnes, Betsy J

    2011-04-01

    Multiple Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded proteins with potential roles in KSHV-associated neoplasms have been identified. KSHV encodes 4 genes with homology to transcription factors of the interferon (IFN) regulatory factor (IRF) family. Viral IRF3 (vIRF3) is expressed in latently KSHV-infected primary effusion lymphoma (PEL) cells and was recently shown to be essential for the survival of PEL cells. The focus of this study was to determine the mechanism(s) of vIRF3 oncogenic activity contributing to KSHV-associated lymphoma. We report that vIRF3 interacts with the amino-terminal DNA binding domain of human IRF5, leading to a complex manipulation of IRF5 function. vIRF3 associated with both exogenous and endogenous IRF5, thereby inhibiting IRF5-mediated IFN promoter activation and the synthesis of biologically active type I IFNs by blocking its binding to endogenous IFNA promoters. The function of this interaction was not limited to the IFN system as IRF5-mediated cell growth regulation was significantly altered by overexpression of vIRF3 in B cells. vIRF3 prevented IRF5-mediated growth inhibition and G2/M cell cycle arrest. Important, IRF5 was upregulated by the protein kinase C agonist 12-O-tetradecanoyl-phorbol-13-acetate in BCBL1 PEL cells and interaction with vIRF3 was observed at the endogenous p21 promoter in response to 12-O-tetradecanoyl-phorbol-13-acetate, suggesting that these 2 proteins cooperate in the regulation of lytic cycle-induced G1 arrest, which is an important early step for the reactivation of KSHV. In conclusion, cellular IRF5 and vIRF3 interact, leading to the functional modulation of IRF5-mediated type I IFN expression and cell cycle regulation. These findings support an important role for vIRF3 in immune evasion and cell proliferation that likely contribute to the survival of PEL cells.

  1. Transcription profile of Escherichia coli: genomic SELEX search for regulatory targets of transcription factors

    PubMed Central

    Ishihama, Akira; Shimada, Tomohiro; Yamazaki, Yukiko

    2016-01-01

    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/). PMID:26843427

  2. Transcription profile of Escherichia coli: genomic SELEX search for regulatory targets of transcription factors.

    PubMed

    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/). PMID:26843427

  3. Regulatory elements responsible for inducible expression of the granulocyte colony-stimulating factor gene in macrophages.

    PubMed Central

    Nishizawa, M; Nagata, S

    1990-01-01

    Granulocyte colony-stimulating factor (G-CSF) plays an essential role in granulopoiesis during bacterial infection. Macrophages produce G-CSF in response to bacterial endotoxins such as lipopolysaccharide (LPS). To elucidate the mechanism of the induction of G-CSF gene in macrophages or macrophage-monocytes, we have examined regulatory cis elements in the promoter of mouse G-CSF gene. Analyses of linker-scanning and internal deletion mutants of the G-CSF promoter by the chloramphenicol acetyltransferase assay have indicated that at least three regulatory elements are indispensable for the LPS-induced expression of the G-CSF gene in macrophages. When one of the three elements was reiterated and placed upstream of the TATA box of the G-CSF promoter, it mediated inducibility as a tissue-specific and orientation-independent enhancer. Although this element contains a conserved NF-kappa B-like binding site, the gel retardation assay and DNA footprint analysis with nuclear extracts from macrophage cell lines demonstrated that nuclear proteins bind to the DNA sequence downstream of the NF-kappa B-like element, but not to the conserved element itself. The DNA sequence of the binding site was found to have some similarities to the LPS-responsive element which was recently identified in the promoter of the mouse class II major histocompatibility gene. Images PMID:1691438

  4. Medusa structure of the gene regulatory network: dominance of transcription factors in cancer subtype classification.

    PubMed

    Guo, Yuchun; Feng, Ying; Trivedi, Niraj S; Huang, Sui

    2011-05-01

    Gene expression profiles consisting of ten thousands of transcripts are used for clustering of tissue, such as tumors, into subtypes, often without considering the underlying reason that the distinct patterns of expression arise because of constraints in the realization of gene expression profiles imposed by the gene regulatory network. The topology of this network has been suggested to consist of a regulatory core of genes represented most prominently by transcription factors (TFs) and microRNAs, that influence the expression of other genes, and of a periphery of 'enslaved' effector genes that are regulated but not regulating. This 'medusa' architecture implies that the core genes are much stronger determinants of the realized gene expression profiles. To test this hypothesis, we examined the clustering of gene expression profiles into known tumor types to quantitatively demonstrate that TFs, and even more pronounced, microRNAs, are much stronger discriminators of tumor type specific gene expression patterns than a same number of randomly selected or metabolic genes. These findings lend support to the hypothesis of a medusa architecture and of the canalizing nature of regulation by microRNAs. They also reveal the degree of freedom for the expression of peripheral genes that are less stringently associated with a tissue type specific global gene expression profile.

  5. TMREC: A Database of Transcription Factor and MiRNA Regulatory Cascades in Human Diseases

    PubMed Central

    Wang, Shuyuan; Li, Wei; Lian, Baofeng; Liu, Xinyi; Zhang, Yan; Dai, Enyu; Yu, Xuexin; Meng, Fanlin; Jiang, Wei; Li, Xia

    2015-01-01

    Over the past decades, studies have reported that the combinatorial regulation of transcription factors (TFs) and microRNAs (miRNAs) is essential for the appropriate execution of biological events and developmental processes. Dysregulations of these regulators often cause diseases. However, there are no available resources on the regulatory cascades of TFs and miRNAs in the context of human diseases. To fulfill this vacancy, we established the TMREC database in this study. First, we integrated curated transcriptional and post-transcriptional regulations to construct the TF and miRNA regulatory network. Next, we identified all linear paths using the Breadth First Search traversal method. Finally, we used known disease-related genes and miRNAs to measure the strength of association between cascades and diseases. Currently, TMREC consists of 74,248 cascades and 25,194 cascade clusters, involving in 412 TFs, 266 miRNAs and 545 diseases. With the expanding of experimental support regulation data, we will regularly update the database. TMREC aims to help experimental biologists to comprehensively analyse gene expression regulation, to understand the aetiology and to predict novel therapeutic targets.TMREC is freely available at http://bioinfo.hrbmu.edu.cn/TMREC/. PMID:25932650

  6. Molecular characterization of interferon regulatory factor 2 (IRF-2) homolog in pearl oyster Pinctada fucata.

    PubMed

    Huang, Xian-De; Liu, Wen-Guang; Wang, Qi; Zhao, Mi; Wu, Shan-Zeng; Guan, Yun-Yan; Shi, Yu; He, Mao-Xian

    2013-05-01

    Interferon regulatory factors (IRFs) control many facets of the innate and adaptive immune responses, regulate the development of the immune system itself and involve in reproduction and morphogenesis. In the present study, the IRF-2 homology gene, PfIRF-2 from pearl oyster Pinctada fucata was cloned and its genomic structure and promoter were analyzed. PfIRF-2 encodes a putative protein of 350 amino acids, and contains a highly conserved N-terminal DNA-binding domain and a variable C-terminal regulatory domain. Comparison and phylogenetic analysis revealed that PfIRF-2 shared a relatively higher identity with other mollusk but relatively lower identity with vertebrate IRF-2, and was clustered with IRF-1 subfamily composed of IRF-2 and IRF-1. Furthermore, gene expression analysis revealed that PfIRF-2 involved in the immune response to LPS and poly(I:C) stimulation. Immunofluorescence assay showed that the expressed PfIRF-2 was translocated into the nucleus and dual-luciferase reporter assays indicated that PfIRF-2 could involved and activate interferon signaling or NF-κB signal pathway in HEK293 cells. The study of PfIRF-2 may help better understand the innate immune in mollusk.

  7. Transcription factors GAF and HSF act at distinct regulatory steps to modulate stress-induced gene activation

    PubMed Central

    Fuda, Nicholas J.; Mahat, Dig B.; Core, Leighton J.; Guertin, Michael J.

    2016-01-01

    The coordinated regulation of gene expression at the transcriptional level is fundamental to development and homeostasis. Inducible systems are invaluable when studying transcription because the regulatory process can be triggered instantaneously, allowing the tracking of ordered mechanistic events. Here, we use precision run-on sequencing (PRO-seq) to examine the genome-wide heat shock (HS) response in Drosophila and the function of two key transcription factors on the immediate transcription activation or repression of all genes regulated by HS. We identify the primary HS response genes and the rate-limiting steps in the transcription cycle that GAGA-associated factor (GAF) and HS factor (HSF) regulate. We demonstrate that GAF acts upstream of promoter-proximally paused RNA polymerase II (Pol II) formation (likely at the step of chromatin opening) and that GAF-facilitated Pol II pausing is critical for HS activation. In contrast, HSF is dispensable for establishing or maintaining Pol II pausing but is critical for the release of paused Pol II into the gene body at a subset of highly activated genes. Additionally, HSF has no detectable role in the rapid HS repression of thousands of genes. PMID:27492368

  8. The Transcription Factor Interferon Regulatory Factor 1 Is Expressed after Cerebral Ischemia and Contributes to Ischemic Brain Injury

    PubMed Central

    Iadecola, Costantino; Salkowski, Cindy A.; Zhang, Fangyi; Aber, Tracy; Nagayama, Masao; Vogel, Stefanie N.; Elizabeth Ross, M.

    1999-01-01

    The transcription factor interferon regulatory factor 1 (IRF-1) is involved in the molecular mechanisms of inflammation and apoptosis, processes that contribute to ischemic brain injury. In this study, the induction of IRF-1 in response to cerebral ischemia and its role in ischemic brain injury were investigated. IRF-1 gene expression was markedly upregulated within 12 h of occlusion of the middle cerebral artery in C57BL/6 mice. The expression reached a peak 4 d after ischemia (6.0 ± 1.8-fold; P < 0.001) and was restricted to the ischemic regions of the brain. The volume of ischemic injury was reduced by 23 ± 3% in IRF-1+/− and by 46 ± 9% in IRF-1−/− mice (P < 0.05). The reduction in infarct volume was paralleled by a substantial attenuation in neurological deficits. Thus, IRF-1 is the first nuclear transacting factor demonstrated to contribute directly to cerebral ischemic damage and may be a novel therapeutic target in ischemic stroke. PMID:9989987

  9. Regulatory factor X1-induced down-regulation of transforming growth factor β2 transcription in human neuroblastoma cells.

    PubMed

    Feng, Chenzhuo; Zuo, Zhiyi

    2012-06-29

    Regulatory factor X (RFX) proteins are transcription factors. Seven mammalian RFX proteins have been identified. RFX1 is the prototype RFX. However, its biological functions are not known. Here, RFX1 overexpression reduced fetal bovine serum-stimulated proliferation of SH-SY5Y cells, a human neuroblastoma cell line. This inhibition is associated with decreased transforming growth factor β2 (TGFβ2) and phospho-extracellular signal-regulated kinase (ERK). Exogenous TGFβ2 increased cell proliferation and phospho-ERK in cells overexpressing RFX1. An anti-TGFβ2 antibody and PD98059, an ERK activation inhibitor, inhibited SH-SY5Y cell proliferation. TGFβ2 promoter activity was decreased in cells overexpressing RFX1. Chromosome immunoprecipitation assay showed that RFX1 bound the TGFβ2 promoter. RFX1 down-regulation increased TGFβ2 in SH-SY5Y and HCN-1A cells, a normal human neuronal cell line. More importantly, TGFβ2 concentrations were negatively correlated with RFX1 levels in human medulloblastoma tissues with a R(2) of 0.464. These results suggest that RFX1 reduces cell proliferation through inhibiting the TGFβ2-ERK signaling pathway. RFX1 blocks TGFβ2 expression through its direct action on TGFβ2 transcription. This effect also appears in human brain tumor tissues. Because TGFβ is known to be involved in cancer development, our results provide initial evidence to suggest that RFX1 may play an important role in human tumor biology.

  10. Somite subdomains, muscle cell origins, and the four muscle regulatory factor proteins

    PubMed Central

    1994-01-01

    We show by immunohistology that distinct expression patterns of the four muscle regulatory factor (MRF) proteins identify subdomains of mouse somites. Myf-5 and MyoD are, at specific stages, each expressed in both myotome and dermatome cells. Myf-5 expression is initially restricted to dorsal cells in all somites, as is MyoD expression in neck somites. In trunk somites, however, MyoD is initially expressed in ventral cells. Myogenin and MRF4 are restricted to myotome cells, though the MRF4-expressing cells are initially less widely distributed than the myogenin-expressing cells, which are at all stages found throughout the myotome. All somitic myocytes express one or more MRFs. The transiently distinct expression patterns of the four MRF proteins identify dorsal and ventral subdomains of somites, and suggest that skeletal muscle cells in somites originate at multiple sites and via multiple molecular pathways. PMID:7929574

  11. Machine learning for regulatory analysis and transcription factor target prediction in yeast

    PubMed Central

    Holloway, Dustin T.; Kon, Mark

    2006-01-01

    High throughput technologies, including array-based chromatin immunoprecipitation, have rapidly increased our knowledge of transcriptional maps—the identity and location of regulatory binding sites within genomes. Still, the full identification of sites, even in lower eukaryotes, remains largely incomplete. In this paper we develop a supervised learning approach to site identification using support vector machines (SVMs) to combine 26 different data types. A comparison with the standard approach to site identification using position specific scoring matrices (PSSMs) for a set of 104 Saccharomyces cerevisiae regulators indicates that our SVM-based target classification is more sensitive (73 vs. 20%) when specificity and positive predictive value are the same. We have applied our SVM classifier for each transcriptional regulator to all promoters in the yeast genome to obtain thousands of new targets, which are currently being analyzed and refined to limit the risk of classifier over-fitting. For the purpose of illustration we discuss several results, including biochemical pathway predictions for Gcn4 and Rap1. For both transcription factors SVM predictions match well with the known biology of control mechanisms, and possible new roles for these factors are suggested, such as a function for Rap1 in regulating fermentative growth. We also examine the promoter melting temperature curves for the targets of YJR060W, and show that targets of this TF have potentially unique physical properties which distinguish them from other genes. The SVM output automatically provides the means to rank dataset features to identify important biological elements. We use this property to rank classifying k-mers, thereby reconstructing known binding sites for several TFs, and to rank expression experiments, determining the conditions under which Fhl1, the factor responsible for expression of ribosomal protein genes, is active. We can see that targets of Fhl1 are differentially expressed in

  12. Are major behavioral and sociodemographic risk factors for mortality additive or multiplicative in their effects?

    PubMed

    Mehta, Neil; Preston, Samuel

    2016-04-01

    All individuals are subject to multiple risk factors for mortality. In this paper, we consider the nature of interactions between certain major sociodemographic and behavioral risk factors associated with all-cause mortality in the United States. We develop the formal logic pertaining to two forms of interaction between risk factors, additive and multiplicative relations. We then consider the general circumstances in which additive or multiplicative relations might be expected. We argue that expectations about interactions among socio-demographic variables, and their relation to behavioral variables, have been stated in terms of additivity. However, the statistical models typically used to estimate the relation between risk factors and mortality assume that risk factors act multiplicatively. We examine empirically the nature of interactions among five major risk factors associated with all-cause mortality: smoking, obesity, race, sex, and educational attainment. Data were drawn from the cross-sectional NHANES III (1988-1994) and NHANES 1999-2010 surveys, linked to death records through December 31, 2011. Our analytic sample comprised 35,604 respondents and 5369 deaths. We find that obesity is additive with each of the remaining four variables. We speculate that its additivity is a reflection of the fact that obese status is generally achieved later in life. For all pairings of socio-demographic variables, risks are multiplicative. For survival chances, it is much more dangerous to be poorly educated if you are black or if you are male. And it is much riskier to be a male if you are black. These traits, established at birth or during childhood, literally result in deadly combinations. We conclude that the identification of interactions among risk factors can cast valuable light on the nature of the process being studied. It also has public health implications by identifying especially vulnerable groups and by properly identifying the proportion of deaths

  13. Intracellular Concentrations of 65 Species of Transcription Factors with Known Regulatory Functions in Escherichia coli

    PubMed Central

    Kori, Ayako; Koshio, Etsuko; Yamada, Kayoko; Maeda, Hiroto; Shimada, Tomohiro; Makinoshima, Hideki; Iwata, Akira; Fujita, Nobuyuki

    2014-01-01

    The expression pattern of the Escherichia coli genome is controlled in part by regulating the utilization of a limited number of RNA polymerases among a total of its approximately 4,600 genes. The distribution pattern of RNA polymerase changes from modulation of two types of protein-protein interactions: the interaction of core RNA polymerase with seven species of the sigma subunit for differential promoter recognition and the interaction of RNA polymerase holoenzyme with about 300 different species of transcription factors (TFs) with regulatory functions. We have been involved in the systematic search for the target promoters recognized by each sigma factor and each TF using the newly developed Genomic SELEX system. In parallel, we developed the promoter-specific (PS)-TF screening system for identification of the whole set of TFs involved in regulation of each promoter. Understanding the regulation of genome transcription also requires knowing the intracellular concentrations of the sigma subunits and TFs under various growth conditions. This report describes the intracellular levels of 65 species of TF with known function in E. coli K-12 W3110 at various phases of cell growth and at various temperatures. The list of intracellular concentrations of the sigma factors and TFs provides a community resource for understanding the transcription regulation of E. coli under various stressful conditions in nature. PMID:24837290

  14. Autoimmune manifestations in human myelodysplasia: a positive correlation with interferon regulatory factor-1 (IRF-1) expression

    PubMed Central

    Giannouli, S; Tzoanopoulos, D; Ritis, K; Kartalis, G; Moutsopoulos, H; Voulgarelis, M

    2004-01-01

    Background: Patients with myelodysplasia may have autoimmune manifestations (AIM). Interferon regulatory factor-1 (IRF-1) is a transcription factor involved in interferon signalling, leukaemogenesis, and the development of the immune system. Objectives: To determine whether IRF-1 is implicated in the pathophysiology of AIM in myelodysplasia. Methods: 14 patients with myelodysplasia were studied, seven with AIM and seven without. Five patients with vasculitis and seven normal subjects served as controls. The expression of IRF-1 was studied in bone marrow mononuclear cells taken from patients and controls, using a relative quantitative reverse transcriptase polymerase chain reaction. Results: A 10-fold reduction in full length IRF-1 mRNA was detected in the myelodysplasia patients without AIM compared with the normal controls. In contrast, the group with AIM had increased IRF-1 transcripts, to a level almost equal to that observed in patients with vasculitis and normal controls. Conclusions: Myelodysplasia patients without IRF-1 expression had a decreased incidence of AIM. Thus the absence of IRF-1 transcription factor appears to protect against the development of autoimmunity in myelodysplasia. PMID:15082491

  15. Inferring regulatory element landscapes and transcription factor networks from cancer methylomes.

    PubMed

    Yao, Lijing; Shen, Hui; Laird, Peter W; Farnham, Peggy J; Berman, Benjamin P

    2015-05-21

    Recent studies indicate that DNA methylation can be used to identify transcriptional enhancers, but no systematic approach has been developed for genome-wide identification and analysis of enhancers based on DNA methylation. We describe ELMER (Enhancer Linking by Methylation/Expression Relationships), an R-based tool that uses DNA methylation to identify enhancers and correlates enhancer state with expression of nearby genes to identify transcriptional targets. Transcription factor motif analysis of enhancers is coupled with expression analysis of transcription factors to infer upstream regulators. Using ELMER, we investigated more than 2,000 tumor samples from The Cancer Genome Atlas. We identified networks regulated by known cancer drivers such as GATA3 and FOXA1 (breast cancer), SOX17 and FOXA2 (endometrial cancer), and NFE2L2, SOX2, and TP63 (squamous cell lung cancer). We also identified novel networks with prognostic associations, including RUNX1 in kidney cancer. We propose ELMER as a powerful new paradigm for understanding the cis-regulatory interface between cancer-associated transcription factors and their functional target genes.

  16. Behavioral and regulatory abnormalities in mice deficient in the NPAS1 and NPAS3 transcription factors.

    PubMed

    Erbel-Sieler, Claudia; Dudley, Carol; Zhou, Yudong; Wu, Xinle; Estill, Sandi Jo; Han, Tina; Diaz-Arrastia, Ramon; Brunskill, Eric W; Potter, S Steven; McKnight, Steven L

    2004-09-14

    Laboratory mice bearing inactivating mutations in the genes encoding the NPAS1 and NPAS3 transcription factors have been shown to exhibit a spectrum of behavioral and neurochemical abnormalities. Behavioral abnormalities included diminished startle response, as measured by prepulse inhibition, and impaired social recognition. NPAS1/NPAS3-deficient mice also exhibited stereotypic darting behavior at weaning and increased locomotor activity. Immunohistochemical staining assays showed that the NPAS1 and NPAS3 proteins are expressed in inhibitory interneurons and that the viability and anatomical distribution of these neurons are unaffected by the absence of either transcription factor. Adult brain tissues from NPAS3- and NPAS1/NPAS3-deficient mice exhibited a distinct reduction in reelin, a large, secreted protein whose expression has been reported to be attenuated in the postmortem brain tissue of patients with schizophrenia. These observations raise the possibility that a regulatory program controlled in inhibitory interneurons by the NPAS1 and NPAS3 transcription factors may be either substantively or tangentially relevant to psychosis.

  17. Behavioral and regulatory abnormalities in mice deficient in the NPAS1 and NPAS3 transcription factors.

    PubMed

    Erbel-Sieler, Claudia; Dudley, Carol; Zhou, Yudong; Wu, Xinle; Estill, Sandi Jo; Han, Tina; Diaz-Arrastia, Ramon; Brunskill, Eric W; Potter, S Steven; McKnight, Steven L

    2004-09-14

    Laboratory mice bearing inactivating mutations in the genes encoding the NPAS1 and NPAS3 transcription factors have been shown to exhibit a spectrum of behavioral and neurochemical abnormalities. Behavioral abnormalities included diminished startle response, as measured by prepulse inhibition, and impaired social recognition. NPAS1/NPAS3-deficient mice also exhibited stereotypic darting behavior at weaning and increased locomotor activity. Immunohistochemical staining assays showed that the NPAS1 and NPAS3 proteins are expressed in inhibitory interneurons and that the viability and anatomical distribution of these neurons are unaffected by the absence of either transcription factor. Adult brain tissues from NPAS3- and NPAS1/NPAS3-deficient mice exhibited a distinct reduction in reelin, a large, secreted protein whose expression has been reported to be attenuated in the postmortem brain tissue of patients with schizophrenia. These observations raise the possibility that a regulatory program controlled in inhibitory interneurons by the NPAS1 and NPAS3 transcription factors may be either substantively or tangentially relevant to psychosis. PMID:15347806

  18. Individual interferon regulatory factor-3 thiol residues are not critical for its activation following virus infection.

    PubMed

    Zucchini, Nicolas; Williams, Virginie; Grandvaux, Nathalie

    2012-09-01

    The interferon regulatory factor (IRF)-3 transcription factor plays a central role in the capacity of the host to mount an efficient innate antiviral immune defense, mainly through the regulation of type I Interferon genes. A tight regulation of IRF-3 is crucial for an adapted intensity and duration of the response. Redox-dependent processes are now well known to regulate signaling cascades. Recent reports have revealed that signaling molecules upstream of IRF-3, including the mitochondrial antiviral-signalling protein (MAVS) and the TNF receptor associated factors (TRAFs) adaptors, are sensitive to redox regulation. In the present study, we assessed whether redox regulation of thiol residues contained in IRF-3, which are priviledged redox sensors, play a role in its regulation following Sendai virus infection, using a combination of mutation of Cysteine (Cys) residues into Alanine and thiols alkylation using N-ethyl maleimide. Alkylation of IRF-3 on Cys289 appears to destabilize IRF-3 dimer in vitro. However, a detailed analysis of IRF-3 phosphorylation, dimerization, nuclear accumulation, and induction of target gene promoter in vivo led us to conclude that IRF-3 specific, individual Cys residues redox status does not play an essential role in its activation in vivo.

  19. Abnormal skin, limb and craniofacial morphogenesis in mice deficient for interferon regulatory factor 6 (Irf6)

    PubMed Central

    Ingraham, Christopher R; Kinoshita, Akira; Kondo, Shinji; Yang, Baoli; Sajan, Samin; Trout, Kurt J; Malik, Margaret I; Dunnwald, Martine; Goudy, Stephen L; Lovett, Michael; Murray, Jeffrey C; Schutte, Brian C

    2007-01-01

    Transcription factor paralogs may share a common role in staged or overlapping expression in specific tissues, as in the Hox family. In other cases, family members have distinct roles in a range of embryologic, differentiation or response pathways (as in the Tbx and Pax families). For the interferon regulatory factor (IRF) family of transcription factors, mice deficient in Irf1, Irf2, Irf3, Irf4, Irf5, Irf7, Irf8 or Irf9 have defects in the immune response but show no embryologic abnormalities1–7. Mice deficient for Irf6 have not been reported, but in humans, mutations in IRF6 cause two mendelian orofacial clefting syndromes8–10, and genetic variation in IRF6 confers risk for isolated cleft lip and palate11–15. Here we report that mice deficient for Irf6 have abnormal skin, limb and craniofacial development. Histological and gene expression analyses indicate that the primary defect is in keratinocyte differentiation and proliferation. This study describes a new role for an IRF family member in epidermal development. PMID:17041601

  20. A novel tumor necrosis factor-responsive transcription factor which recognizes a regulatory element in hemopoietic growth factor genes

    SciTech Connect

    Shannon, M.F.; Pell, L.M.; Kuczek, E.S.; Occhiodoro, F.S.; Dunn, S.M.; Vadas, M.A. ); Lenardo, M.J. )

    1990-06-01

    A conserved DNA sequence element, termed cytokine 1 (CK-1), is found in the promoter regions of many hemopoietic growth factor (HGF) genes. Mutational analyses and modification interference experiments show that this sequence specifically binds a nuclear transcription factor, NF-GMa, which is a protein with a molecular mass of 43 kilodaltons. It interacts with different affinities with the CK-1-like sequence from a number of HGF genes, including granulocyte macrophage colony-stimulating factor (GM-CSF), granulocyte (G)-CSF, interleukin 3 (IL-3), and IL-5. The authors show that the level of NF-GMa binding is induced in embryonic fibroblasts by tumor necrosis factor {alpha} (TNF-{alpha}) treatment and that the CK-1 sequence from the G-CSF gene is a TNF-{alpha}-responsive enhancer in these cells.

  1. Interferon-Regulatory Factor 5-Dependent Signaling Restricts Orthobunyavirus Dissemination to the Central Nervous System

    PubMed Central

    Proenca-Modena, Jose Luiz; Hyde, Jennifer L.; Sesti-Costa, Renata; Lucas, Tiffany; Pinto, Amelia K.; Richner, Justin M.; Gorman, Matthew J.; Lazear, Helen M.

    2015-01-01

    ABSTRACT Interferon (IFN)-regulatory factor 5 (IRF-5) is a transcription factor that induces inflammatory responses after engagement and signaling by pattern recognition receptors. To define the role of IRF-5 during bunyavirus infection, we evaluated Oropouche virus (OROV) and La Crosse virus (LACV) pathogenesis and immune responses in primary cells and in mice with gene deletions in Irf3, Irf5, and Irf7 or in Irf5 alone. Deletion of Irf3, Irf5, and Irf7 together resulted in uncontrolled viral replication in the liver and spleen, hypercytokinemia, extensive liver injury, and an early-death phenotype. Remarkably, deletion of Irf5 alone resulted in meningoencephalitis and death on a more protracted timeline, 1 to 2 weeks after initial OROV or LACV infection. The clinical signs in OROV-infected Irf5−/− mice were associated with abundant viral antigen and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL)-positive cells in several regions of the brain. Circulating dendritic cell (DC) subsets in Irf5−/− mice had higher levels of OROV RNA in vivo yet produced lower levels of type I IFN than wild-type (WT) cells. This result was supported by data obtained in vitro, since a deficiency of IRF-5 resulted in enhanced OROV infection and diminished type I IFN production in bone marrow-derived DCs. Collectively, these results indicate a key role for IRF-5 in modulating the host antiviral response in peripheral organs that controls bunyavirus neuroinvasion in mice. IMPORTANCE Oropouche virus (OROV) and La Crosse virus (LACV) are orthobunyaviruses that are transmitted by insects and cause meningitis and encephalitis in subsets of individuals in the Americas. Recently, we demonstrated that components of the type I interferon (IFN) induction pathway, particularly the regulatory transcription factors IRF-3 and IRF-7, have key protective roles during OROV infection. However, the lethality in Irf3−/− Irf7−/− (DKO) mice infected with OROV

  2. Overexpression of the Dominant-Negative Form of Interferon Regulatory Factor 1 in Oligodendrocytes Protects against Experimental Autoimmune Encephalomyelitis

    PubMed Central

    Ren, Zhihua; Wang, Yan; Tao, Duan; Liebenson, David; Liggett, Thomas; Goswami, Rajendra; Clarke, Robert; Stefoski, Dusan

    2011-01-01

    Interferon regulatory factor 1 (IRF-1) is a transcription factor that has been implicated in the pathogenesis of the human autoimmune demyelinating disease multiple sclerosis (MS) and in its animal model, experimental autoimmune encephalomyelitis (EAE). The goal of the present study was to directly examine the role of IRF-1 in oligodendrocyte injury and inflammatory demyelination. For the purpose of this study, we generated a transgenic mouse line (CNP/dnIRF-1) that overexpresses the dominant-negative form of IRF-1 (dnIRF1) specifically in oligodendrocytes. CNP/dnIRF-1 mice exhibited no phenotypic abnormalities but displayed suppressed IRF-1 signaling in oligodendrocytes. The major finding of our study was that the CNP/dnIRF-1 mice, compared with the wild-type mice, were protected against EAE, a phenomenon associated with significant reduction of inflammatory demyelination and with oligodendrocyte and axonal preservation. The observed protection was related to suppressed IRF-1 signaling and impaired expression of immune and proapoptotic genes in oligodendrocytes. No significant differences in the peripheral immune responses between the wild-type and the CNP/dnIRF-1 mice were identified throughout the experiments. This study indicates that IRF-1 plays a critical role in the pathogenesis of EAE by mediating oligodendrocyte response to inflammation and injury. It also suggests that oligodendrocytes are actively involved in the neuroimmune network, and that exploring oligodendrocyte-related pathogenic mechanisms, in addition to the conventional immune-based ones, may have important therapeutic implications in MS. PMID:21653838

  3. Regulation of the germinal center gene program by interferon (IFN) regulatory factor 8/IFN consensus sequence-binding protein

    PubMed Central

    Lee, Chang Hoon; Melchers, Mark; Wang, Hongsheng; Torrey, Ted A.; Slota, Rebecca; Qi, Chen-Feng; Kim, Ji Young; Lugar, Patricia; Kong, Hee Jeong; Farrington, Lila; van der Zouwen, Boris; Zhou, Jeff X.; Lougaris, Vassilios; Lipsky, Peter E.; Grammer, Amrie C.; Morse, Herbert C.

    2006-01-01

    Interferon (IFN) consensus sequence-binding protein/IFN regulatory factor 8 (IRF8) is a transcription factor that regulates the differentiation and function of macrophages, granulocytes, and dendritic cells through activation or repression of target genes. Although IRF8 is also expressed in lymphocytes, its roles in B cell and T cell maturation or function are ill defined, and few transcriptional targets are known. Gene expression profiling of human tonsillar B cells and mouse B cell lymphomas showed that IRF8 transcripts were expressed at highest levels in centroblasts, either from secondary lymphoid tissue or transformed cells. In addition, staining for IRF8 was most intense in tonsillar germinal center (GC) dark-zone centroblasts. To discover B cell genes regulated by IRF8, we transfected purified primary tonsillar B cells with enhanced green fluorescent protein–tagged IRF8, generated small interfering RNA knockdowns of IRF8 expression in a mouse B cell lymphoma cell line, and examined the effects of a null mutation of IRF8 on B cells. Each approach identified activation-induced cytidine deaminase (AICDA) and BCL6 as targets of transcriptional activation. Chromatin immunoprecipitation studies demonstrated in vivo occupancy of 5′ sequences of both genes by IRF8 protein. These results suggest previously unappreciated roles for IRF8 in the transcriptional regulation of B cell GC reactions that include direct regulation of AICDA and BCL6. PMID:16380510

  4. Regulation of the germinal center gene program by interferon (IFN) regulatory factor 8/IFN consensus sequence-binding protein.

    PubMed

    Lee, Chang Hoon; Melchers, Mark; Wang, Hongsheng; Torrey, Ted A; Slota, Rebecca; Qi, Chen-Feng; Kim, Ji Young; Lugar, Patricia; Kong, Hee Jeong; Farrington, Lila; van der Zouwen, Boris; Zhou, Jeff X; Lougaris, Vassilios; Lipsky, Peter E; Grammer, Amrie C; Morse, Herbert C

    2006-01-23

    Interferon (IFN) consensus sequence-binding protein/IFN regulatory factor 8 (IRF8) is a transcription factor that regulates the differentiation and function of macrophages, granulocytes, and dendritic cells through activation or repression of target genes. Although IRF8 is also expressed in lymphocytes, its roles in B cell and T cell maturation or function are ill defined, and few transcriptional targets are known. Gene expression profiling of human tonsillar B cells and mouse B cell lymphomas showed that IRF8 transcripts were expressed at highest levels in centroblasts, either from secondary lymphoid tissue or transformed cells. In addition, staining for IRF8 was most intense in tonsillar germinal center (GC) dark-zone centroblasts. To discover B cell genes regulated by IRF8, we transfected purified primary tonsillar B cells with enhanced green fluorescent protein-tagged IRF8, generated small interfering RNA knockdowns of IRF8 expression in a mouse B cell lymphoma cell line, and examined the effects of a null mutation of IRF8 on B cells. Each approach identified activation-induced cytidine deaminase (AICDA) and BCL6 as targets of transcriptional activation. Chromatin immunoprecipitation studies demonstrated in vivo occupancy of 5' sequences of both genes by IRF8 protein. These results suggest previously unappreciated roles for IRF8 in the transcriptional regulation of B cell GC reactions that include direct regulation of AICDA and BCL6.

  5. Impact of treadmill locomotor training on skeletal muscle IGF1 and myogenic regulatory factors in spinal cord injured rats.

    PubMed

    Liu, Min; Stevens-Lapsley, Jennifer E; Jayaraman, Arun; Ye, Fan; Conover, Christine; Walter, Glenn A; Bose, Prodip; Thompson, Floyd J; Borst, Stephen E; Vandenborne, Krista

    2010-07-01

    The objective of this study was to determine the impact of treadmill locomotor training on the expression of insulin-like growth factor I (IGF1) and changes in myogenic regulatory factors (MRFs) in rat soleus muscle following spinal cord injury (SCI). Moderate, midthoracic (T(8)) contusion SCIs were produced using a NYU (New York University) impactor. Animals were randomly assigned to treadmill training or untrained groups. Rats in the training group were trained starting at 1 week after SCI, for either 3 bouts of 20 min over 1.5 days or 10 bouts over 5 days. Five days of treadmill training completely prevented the decrease in soleus fiber size resulting from SCI. In addition, treadmill training triggered increases in IGF1, MGF and IGFBP4 mRNA expression, and a concurrent reduction of IGFBP5 mRNA in skeletal muscle. Locomotor training also caused an increase in markers of muscle regeneration, including small muscle fibers expressing embryonic myosin and Pax7 positive nuclei and increased expression of the MRFs, myogenin and MyoD. We concluded that treadmill locomotor training ameliorated muscle atrophy in moderate contusion SCI rats. Training-induced muscle regeneration and fiber hypertrophy following SCI was associated with an increase in IGF1, an increase in Pax7 positive nuclei, and upregulation of MRFs.

  6. Ubiquitination of Tumor Necrosis Factor Receptor-associated Factor 4 (TRAF4) by Smad Ubiquitination Regulatory Factor 1 (Smurf1) Regulates Motility of Breast Epithelial and Cancer Cells*

    PubMed Central

    Wang, Xiangchun; Jin, Chaoyang; Tang, Yi; Tang, Liu-Ya; Zhang, Ying E.

    2013-01-01

    Smad ubiquitin regulatory factors (Smurfs) are HECT-domain ubiquitin E3 ligases that regulate diverse cellular processes, including normal and tumor cell migration. However, the underlying mechanism of the Smurfs' role in cell migration is not fully understood. Here we show that Smurf1 induces ubiquitination of tumor necrosis factor receptor-associated factor 4 (TRAF4) at K190. Using the K190R mutant of TRAF4, we demonstrate that Smurf1-induced ubiquitination is required for proper localization of TRAF4 to tight junctions in confluent epithelial cells. We further show that TRAF4 is essential for the migration of both normal mammary epithelial and breast cancer cells. The ability of TRAF4 to promote cell migration is also dependent on Smurf1-mediated ubiquitination, which is associated with Rac1 activation by TRAF4. These results reveal a new regulatory circuit for cell migration, consisting of Smurf1-mediated ubiquitination of TRAF4 and Rac1 activation. PMID:23760265

  7. Imbalanced signal transduction in regulatory T cells expressing the transcription factor FoxP3

    PubMed Central

    Yan, Dapeng; Farache, Julia; Mingueneau, Michael; Mathis, Diane; Benoist, Christophe

    2015-01-01

    FoxP3+ T regulatory (Treg) cells have a fundamental role in immunological tolerance, with transcriptional and functional phenotypes that demarcate them from conventional CD4+ T cells (Tconv). Differences between these two lineages in the signaling downstream of T-cell receptor-triggered activation have been reported, and there are different requirements for some signaling factors. Seeking a comprehensive view, we found that Treg cells have a broadly dampened activation of several pathways and signaling nodes upon TCR-mediated activation, with low phosphorylation of CD3ζ, SLP76, Erk1/2, AKT, or S6 and lower calcium flux. In contrast, STAT phosphorylation triggered by interferons, IL2 or IL6, showed variations between Treg and Tconv in magnitude or choice of preferential STAT activation but no general Treg signaling defect. Much, but not all, of the Treg/Tconv difference in TCR-triggered responses could be attributed to lower responsiveness of antigen-experienced cells with CD44hi or CD62Llo phenotypes, which form a greater proportion of the Treg pool. Candidate regulators were tested, but the Treg/Tconv differential could not be explained by overexpression in Treg cells of the signaling modulator CD5, the coinhibitors PD-1 and CTLA4, or the regulatory phosphatase DUSP4. However, transcriptome profiling in Dusp4-deficient mice showed that DUSP4 enhances the expression of a segment of the canonical Treg transcriptional signature, which partially overlaps with the TCR-dependent Treg gene set. Thus, Treg cells, likely because of their intrinsically higher reactivity to self, tune down TCR signals but seem comparatively more attuned to cytokines or other intercellular signals. PMID:26627244

  8. Restricted maternal nutrition alters myogenic regulatory factor expression in satellite cells of ovine offspring.

    PubMed

    Raja, J S; Hoffman, M L; Govoni, K E; Zinn, S A; Reed, S A

    2016-07-01

    Poor maternal nutrition inhibits muscle development and postnatal muscle growth. Satellite cells are myogenic precursor cells that contribute to postnatal muscle growth, and their activity can be evaluated by the expression of several transcription factors. Paired-box (Pax)7 is expressed in quiescent and active satellite cells. MyoD is expressed in activated and proliferating satellite cells and myogenin is expressed in terminally differentiating cells. Disruption in the expression pattern or timing of expression of myogenic regulatory factors negatively affects muscle development and growth. We hypothesized that poor maternal nutrition during gestation would alter the in vitro temporal expression of MyoD and myogenin in satellite cells from offspring at birth and 3 months of age. Ewes were fed 100% or 60% of NRC requirements from day 31±1.3 of gestation. Lambs from control-fed (CON) or restricted-fed (RES) ewes were euthanized within 24 h of birth (birth; n=5) or were fed a control diet until 3 months of age (n=5). Satellite cells isolated from the semitendinosus muscle were used for gene expression analysis or cultured for 24, 48 or 72 h and immunostained for Pax7, MyoD or myogenin. Fusion index was calculated from a subset of cells allowed to differentiate. Compared with CON, temporal expression of MyoD and myogenin was altered in cultured satellite cells isolated from RES lambs at birth. The percent of cells expressing MyoD was greater in RES than CON (P=0.03) after 24 h in culture. After 48 h of culture, there was a greater percent of cells expressing myogenin in RES compared with CON (P0.05). In satellite cells from RES lambs at 3 months of age, the percent of cells expressing MyoD and myogenin were greater than CON after 72 h in culture (P<0.05). Fusion index was reduced in RES lambs at 3 months of age compared with CON (P<0.001). Restricted nutrition during gestation alters the temporal expression of myogenic regulatory factors in satellite cells of the

  9. Multiple Ets factors and interferon regulatory factor-4 modulate CD68 expression in a cell type-specific manner.

    PubMed

    O'Reilly, Dawn; Quinn, Carmel M; El-Shanawany, Tariq; Gordon, Siamon; Greaves, David R

    2003-06-13

    CD68 is a transmembrane glycoprotein expressed in all cells of the mononuclear phagocyte lineage including monocytes and tissue resident macrophages. Deletion analysis of the 5'-flanking sequences of the gene demonstrated that the proximal -150-bp sequence of the CD68 promoter exhibits high level promoter activity in macrophages. Mutations that abolish Ets factor binding at positions -106 and -89 reduce promoter activity in macrophages to 12 and 30%, respectively. Band shift experiments show that PU.1 associates with the -89 site whereas, Elf-1 preferentially binds the -106 Ets binding site and enhances CD68 activity in vitro. Furthermore, chromatin immunoprecipitation experiments confirm that Elf-1 and PU.1 associate with the CD68 proximal promoter in vivo in THP-1 cells. PU.1 does not bind to the CD68 promoter alone but instead forms heterocomplexes with members of the interferon regulatory factor family (IRF) including IRF-4 and IRF-8. IRF-4 and IRF-8 typically mediate transcriptional activation when associated with PU.1 on composite elements. However, our data show that PU.1/IRF-4 and IRF-8 heterocomplexes down-regulate CD68 promoter activity in macrophages and repression is dependent on the integrity of both the IRF and PU.1 half-sites of this composite element. Chromatin immunoprecipitation data reveal that neither IRF-4 nor IRF-8 associate with the CD68 proximal promoter in macrophages in vivo but IRF-4 is associated with the promoter in B lymphocytes. We propose that expression of CD68 in myeloid cells requires the Ets transcription factors Elf-1 and PU.1 and CD68 expression is down-regulated in lymphoid cells by combinatorial interactions between PU.1 and IRF-4.

  10. Antiviral Activity of Porcine Interferon Regulatory Factor 1 against Swine Viruses in Cell Culture.

    PubMed

    Li, Yongtao; Chang, Hongtao; Yang, Xia; Zhao, Yongxiang; Chen, Lu; Wang, Xinwei; Liu, Hongying; Wang, Chuanqing; Zhao, Jun

    2015-11-17

    Interferon regulatory factor 1 (IRF1), as an important transcription factor, is abundantly induced upon virus infections and participates in host antiviral immune responses. However, the roles of porcine IRF1 (poIRF1) in host antiviral defense remain poorly understood. In this study, we determined that poIRF1 was upregulated upon infection with viruses and distributed in nucleus in porcine PK-15 cells. Subsequently, we tested the antiviral activities of poIRF1 against several swine viruses in cells. Overexpression of poIRF1 can efficiently suppress the replication of viruses, and knockdown of poIRF1 promotes moderately viral replication. Interestingly, overexpression of poIRF1 enhances dsRNA-induced IFN-β and IFN-stimulated response element (ISRE) promoter activation, whereas knockdown of poIRF1 cannot significantly affect the activation of IFN-β promoter induced by RNA viruses. This study suggests that poIRF1 plays a significant role in cellular antiviral response against swine viruses, but might be dispensable for IFN-β induction triggered by RNA viruses in PK-15 cells. Given these results, poIRF1 plays potential roles in cellular antiviral responses against swine viruses.

  11. Interferon Regulatory Factor 4 controls TH1 cell effector function and metabolism

    PubMed Central

    Mahnke, Justus; Schumacher, Valéa; Ahrens, Stefanie; Käding, Nadja; Feldhoff, Lea Marie; Huber, Magdalena; Rupp, Jan; Raczkowski, Friederike; Mittrücker, Hans-Willi

    2016-01-01

    The transcription factor Interferon Regulatory Factor 4 (IRF4) is essential for TH2 and TH17 cell formation and controls peripheral CD8+ T cell differentiation. We used Listeria monocytogenes infection to characterize the function of IRF4 in TH1 responses. IRF4−/− mice generated only marginal numbers of listeria-specific TH1 cells. After transfer into infected mice, IRF4−/− CD4+ T cells failed to differentiate into TH1 cells as indicated by reduced T-bet and IFN-γ expression, and showed limited proliferation. Activated IRF4−/− CD4+ T cells exhibited diminished uptake of the glucose analog 2-NBDG, limited oxidative phosphorylation and strongly reduced aerobic glycolysis. Insufficient metabolic adaptation contributed to the limited proliferation and TH1 differentiation of IRF4−/− CD4+ T cells. Our study identifies IRF4 as central regulator of TH1 responses and cellular metabolism. We propose that this function of IRF4 is fundamental for the initiation and maintenance of all TH cell responses. PMID:27762344

  12. Ikkepsilon regulates viral-induced interferon regulatory factor-3 activation via a redox-sensitive pathway

    SciTech Connect

    Indukuri, Hemalatha; Castro, Shawn M.; Liao, S.-M.; Feeney, Lee Ann; Dorsch, Marion; Coyle, Anthony J.; Garofalo, Roberto P.; Brasier, Allan R.; Casola, Antonella . E-mail: ancasola@utmb.edu

    2006-09-15

    Respiratory syncytial virus (RSV)-induced chemokine gene expression occurs through the activation of a subset of transcription factors, including Interferon Regulatory Factor (IRF)-3. In this study, we have investigated the signaling pathway leading to RSV-induced IRF-3 activation and whether it is mediated by intracellular reactive oxygen species (ROS) generation. Our results show that RSV infection induces expression and catalytic activity of IKK{epsilon}, a noncanonical IKK-like kinase. Expression of a kinase-inactive IKK{epsilon} blocks RSV-induced IRF-3 serine phosphorylation, nuclear translocation and DNA-binding, leading to inhibition of RANTES gene transcription, mRNA expression and protein synthesis. Treatment of alveolar epithelial cells with antioxidants or with NAD(P)H oxidase inhibitors abrogates RSV-induced chemokine secretion, IRF-3 phosphorylation and IKK{epsilon} induction, indicating that ROS generation plays a fundamental role in the signaling pathway leading to IRF-3 activation, therefore, identifying a novel molecular target for the development of strategies aimed to modify the inflammatory response associated with RSV infection of the lung.

  13. Origin of a novel regulatory module by duplication and degeneration of an ancient plant transcription factor.

    PubMed

    Floyd, Sandra K; Ryan, Joseph G; Conway, Stephanie J; Brenner, Eric; Burris, Kellie P; Burris, Jason N; Chen, Tao; Edger, Patrick P; Graham, Sean W; Leebens-Mack, James H; Pires, J Chris; Rothfels, Carl J; Sigel, Erin M; Stevenson, Dennis W; Neal Stewart, C; Wong, Gane Ka-Shu; Bowman, John L

    2014-12-01

    It is commonly believed that gene duplications provide the raw material for morphological evolution. Both the number of genes and size of gene families have increased during the diversification of land plants. Several small proteins that regulate transcription factors have recently been identified in plants, including the LITTLE ZIPPER (ZPR) proteins. ZPRs are post-translational negative regulators, via heterodimerization, of class III Homeodomain Leucine Zipper (C3HDZ) proteins that play a key role in directing plant form and growth. We show that ZPR genes originated as a duplication of a C3HDZ transcription factor paralog in the common ancestor of euphyllophytes (ferns and seed plants). The ZPRs evolved by degenerative mutations resulting in loss all of the C3HDZ functional domains, except the leucine zipper that modulates dimerization. ZPRs represent a novel regulatory module of the C3HDZ network unique to the euphyllophyte lineage, and their origin correlates to a period of rapid morphological changes and increased complexity in land plants. The origin of the ZPRs illustrates the significance of gene duplications in creating developmental complexity during land plant evolution that likely led to morphological evolution. PMID:25263420

  14. 34 CFR 648.32 - What additional factors does the Secretary consider?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 34 Education 3 2010-07-01 2010-07-01 false What additional factors does the Secretary consider? 648.32 Section 648.32 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF POSTSECONDARY EDUCATION, DEPARTMENT OF EDUCATION GRADUATE ASSISTANCE IN AREAS OF NATIONAL...

  15. 34 CFR 491.22 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 34 Education 3 2013-07-01 2013-07-01 false What additional factor does the Secretary consider? 491.22 Section 491.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION ADULT EDUCATION FOR THE...

  16. 34 CFR 491.22 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 34 Education 3 2012-07-01 2012-07-01 false What additional factor does the Secretary consider? 491.22 Section 491.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION ADULT EDUCATION FOR THE...

  17. 34 CFR 491.22 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 34 Education 3 2014-07-01 2014-07-01 false What additional factor does the Secretary consider? 491.22 Section 491.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION ADULT EDUCATION FOR THE...

  18. 34 CFR 491.22 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 34 Education 3 2011-07-01 2011-07-01 false What additional factor does the Secretary consider? 491.22 Section 491.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION ADULT EDUCATION FOR THE...

  19. 34 CFR 491.22 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 34 Education 3 2010-07-01 2010-07-01 false What additional factor does the Secretary consider? 491.22 Section 491.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION ADULT EDUCATION FOR THE...

  20. 34 CFR 636.22 - What additional factors does the Secretary consider?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 34 Education 3 2010-07-01 2010-07-01 false What additional factors does the Secretary consider? 636.22 Section 636.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF POSTSECONDARY EDUCATION, DEPARTMENT OF EDUCATION URBAN COMMUNITY SERVICE PROGRAM How Does...

  1. 34 CFR 636.22 - What additional factors does the Secretary consider?

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 34 Education 3 2014-07-01 2014-07-01 false What additional factors does the Secretary consider? 636.22 Section 636.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF POSTSECONDARY EDUCATION, DEPARTMENT OF EDUCATION URBAN COMMUNITY SERVICE PROGRAM How Does...

  2. 34 CFR 636.22 - What additional factors does the Secretary consider?

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 34 Education 3 2012-07-01 2012-07-01 false What additional factors does the Secretary consider? 636.22 Section 636.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF POSTSECONDARY EDUCATION, DEPARTMENT OF EDUCATION URBAN COMMUNITY SERVICE PROGRAM How Does...

  3. 34 CFR 636.22 - What additional factors does the Secretary consider?

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 34 Education 3 2013-07-01 2013-07-01 false What additional factors does the Secretary consider? 636.22 Section 636.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF POSTSECONDARY EDUCATION, DEPARTMENT OF EDUCATION URBAN COMMUNITY SERVICE PROGRAM How Does...

  4. 34 CFR 636.22 - What additional factors does the Secretary consider?

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 34 Education 3 2011-07-01 2011-07-01 false What additional factors does the Secretary consider? 636.22 Section 636.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF POSTSECONDARY EDUCATION, DEPARTMENT OF EDUCATION URBAN COMMUNITY SERVICE PROGRAM How Does...

  5. 21 CFR 1311.115 - Additional requirements for two-factor authentication.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 9 2013-04-01 2013-04-01 false Additional requirements for two-factor authentication. 1311.115 Section 1311.115 Food and Drugs DRUG ENFORCEMENT ADMINISTRATION, DEPARTMENT OF JUSTICE... criteria of FIPS 140-2 Security Level 1, as incorporated by reference in § 1311.08, for...

  6. 21 CFR 1311.115 - Additional requirements for two-factor authentication.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 9 2012-04-01 2012-04-01 false Additional requirements for two-factor authentication. 1311.115 Section 1311.115 Food and Drugs DRUG ENFORCEMENT ADMINISTRATION, DEPARTMENT OF JUSTICE... criteria of FIPS 140-2 Security Level 1, as incorporated by reference in § 1311.08, for...

  7. 34 CFR 425.22 - What additional factors does the Secretary consider?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 34 Education 3 2010-07-01 2010-07-01 false What additional factors does the Secretary consider? 425.22 Section 425.22 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION DEMONSTRATION PROJECTS FOR...

  8. 34 CFR 648.32 - What additional factors does the Secretary consider?

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... private institutions of higher education. (Authority: 20 U.S.C. 1135-1135c) ... 34 Education 3 2011-07-01 2011-07-01 false What additional factors does the Secretary consider? 648.32 Section 648.32 Education Regulations of the Offices of the Department of Education...

  9. 34 CFR 472.23 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 34 Education 3 2010-07-01 2010-07-01 false What additional factor does the Secretary consider? 472.23 Section 472.23 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION NATIONAL WORKPLACE LITERACY PROGRAM...

  10. 34 CFR 472.23 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 34 Education 3 2012-07-01 2012-07-01 false What additional factor does the Secretary consider? 472.23 Section 472.23 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION NATIONAL WORKPLACE LITERACY PROGRAM...

  11. 34 CFR 472.23 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 34 Education 3 2014-07-01 2014-07-01 false What additional factor does the Secretary consider? 472.23 Section 472.23 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION NATIONAL WORKPLACE LITERACY PROGRAM...

  12. 34 CFR 472.23 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 34 Education 3 2011-07-01 2011-07-01 false What additional factor does the Secretary consider? 472.23 Section 472.23 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION NATIONAL WORKPLACE LITERACY PROGRAM...

  13. 34 CFR 472.23 - What additional factor does the Secretary consider?

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 34 Education 3 2013-07-01 2013-07-01 false What additional factor does the Secretary consider? 472.23 Section 472.23 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF VOCATIONAL AND ADULT EDUCATION, DEPARTMENT OF EDUCATION NATIONAL WORKPLACE LITERACY PROGRAM...

  14. Tissue-specific targeting of cell fate regulatory genes by E2f factors.

    PubMed

    Julian, L M; Liu, Y; Pakenham, C A; Dugal-Tessier, D; Ruzhynsky, V; Bae, S; Tsai, S-Y; Leone, G; Slack, R S; Blais, A

    2016-04-01

    Cell cycle proteins are important regulators of diverse cell fate decisions, and in this capacity have pivotal roles in neurogenesis and brain development. The mechanisms by which cell cycle regulation is integrated with cell fate control in the brain and other tissues are poorly understood, and an outstanding question is whether the cell cycle machinery regulates fate decisions directly or instead as a secondary consequence of proliferative control. Identification of the genes targeted by E2 promoter binding factor (E2f) transcription factors, effectors of the pRb/E2f cell cycle pathway, will provide essential insights into these mechanisms. We identified the promoter regions bound by three neurogenic E2f factors in neural precursor cells in a genome-wide manner. Through bioinformatic analyses and integration of published genomic data sets we uncovered hundreds of transcriptionally active E2f-bound promoters corresponding to genes that control cell fate processes, including key transcriptional regulators and members of the Notch, fibroblast growth factor, Wnt and Tgf-β signaling pathways. We also demonstrate a striking enrichment of the CCCTC binding factor transcription factor (Ctcf) at E2f3-bound nervous system-related genes, suggesting a potential regulatory co-factor for E2f3 in controlling differentiation. Finally, we provide the first demonstration of extensive tissue specificity among E2f target genes in mammalian cells, whereby E2f3 promoter binding is well conserved between neural and muscle precursors at genes associated with cell cycle processes, but is tissue-specific at differentiation-associated genes. Our findings implicate the cell cycle pathway as a widespread regulator of cell fate genes, and suggest that E2f3 proteins control cell type-specific differentiation programs by regulating unique sets of target genes. This work significantly enhances our understanding of how the cell cycle machinery impacts cell fate and differentiation, and will

  15. Expression regulation of zebrafish interferon regulatory factor 9 by promoter analysis.

    PubMed

    Shi, Jun; Zhang, Yi-Bing; Zhang, Jian-She; Gui, Jian-Fang

    2013-12-01

    We previously showed that a fish interferon (IFN) regulatory factor 9 (IRF9) homologue, crucian carp Carassius auratus IRF9, displays constitutively nuclear localization and involvement in fish IFN-dependent JAK-STAT signaling; however, little is known about the expression regulation of fish IRF9. Here, we characterized the expression of zebrafish IRF9 by promoter analysis. Zebrafish IRF9 gene promoter contained several putative transcription factor binding sites, including one ISRE (IFN-stimulated response element), one GAS (IFN gamma activation sequence) and three GATEs (IFNγ activated transcriptional element, GATE1/2/3). Further sequence analyses revealed that GAS and GATE motifs existed in all promoters of IRF9 from mammals and fishes. Luciferase assays confirmed that zebrafish IRF9 promoter could be activated by zebrafish IFNφs and zebrafish IFNγ2, as well as transcription factors IRF3, IRF7, and combination of IRF9 and STAT2. Treatment of recombinant crucian carp IFN protein or overexpression of zebrafish IFNγ2 both led to significant increase in crucian carp IRF9 mRNA and protein in cultured fish cells. Comparison of IFN-stimulated promoter activity revealed much more significant induction of zebrafish IRF9 by zebrafish IFNγ2 than by zebrafish IFNφs. Mutation analyses showed that the putative GAS and GATE3 contributed to zebrafish IFNγ2-triggered IRF9 expression, whereas the putative ISRE and the other two GATEs were not functional for induction of zebrafish IRF9. These results together indicated that the expression property of IRF9 might be conserved from fish to mammals and that some not yet identified mechanisms could exist in IRF9 gene transcription regulation in response to IFNs.

  16. Inhibition of protein kinase C catalytic activity by additional regions within the human protein kinase Calpha-regulatory domain lying outside of the pseudosubstrate sequence.

    PubMed Central

    Kirwan, Angie F; Bibby, Ashley C; Mvilongo, Thierry; Riedel, Heimo; Burke, Thomas; Millis, Sherri Z; Parissenti, Amadeo M

    2003-01-01

    The N-terminal pseudosubstrate site within the protein kinase Calpha (PKCalpha)-regulatory domain has long been regarded as the major determinant for autoinhibition of catalytic domain activity. Previously, we observed that the PKC-inhibitory capacity of the human PKCalpha-regulatory domain was only reduced partially on removal of the pseudosubstrate sequence [Parissenti, Kirwan, Kim, Colantonio and Schimmer (1998) J. Biol. Chem. 273, 8940-8945]. This finding suggested that one or more additional region(s) contributes to the inhibition of catalytic domain activity. To assess this hypothesis, we first examined the PKC-inhibitory capacity of a smaller fragment of the PKCalpha-regulatory domain consisting of the C1a, C1b and V2 regions [GST-Ralpha(39-177): this protein contained the full regulatory domain of human PKCalpha fused to glutathione S-transferase (GST), but lacked amino acids 1-38 (including the pseudosubstrate sequence) and amino acids 178-270 (including the C2 region)]. GST-Ralpha(39-177) significantly inhibited PKC in a phorbol-independent manner and could not bind the peptide substrate used in our assays. These results suggested that a region within C1/V2 directly inhibits catalytic domain activity. Providing further in vivo support for this hypothesis, we found that expression of N-terminally truncated pseudosubstrate-less bovine PKCalpha holoenzymes in yeast was capable of inhibiting cell growth in a phorbol-dependent manner. This suggested that additional autoinhibitory force(s) remained within the truncated holoenzymes that could be relieved by phorbol ester. Using tandem PCR-mediated mutagenesis, we observed that mutation of amino acids 33-86 within GST-Ralpha(39-177) dramatically reduced its PKC-inhibitory capacity when protamine was used as substrate. Mutagenesis of a broad range of sequences within C2 (amino acids 159-242) also significantly reduced PKC-inhibitory capacity. Taken together, these observations support strongly the existence of

  17. Isolation of an additional member of the fibroblast growth factor receptor family, FGFR-3.

    PubMed Central

    Keegan, K; Johnson, D E; Williams, L T; Hayman, M J

    1991-01-01

    The fibroblast growth factors are a family of polypeptide growth factors involved in a variety of activities including mitogenesis, angiogenesis, and wound healing. Fibroblast growth factor receptors (FGFRs) have previously been identified in chicken, mouse, and human and have been shown to contain an extracellular domain with either two or three immunoglobulin-like domains, a transmembrane domain, and a cytoplasmic tyrosine kinase domain. We have isolated a human cDNA for another tyrosine kinase receptor that is highly homologous to the previously described FGFR. Expression of this receptor cDNA in COS cells directs the expression of a 125-kDa glycoprotein. We demonstrate that this cDNA encodes a biologically active receptor by showing that human acidic and basic fibroblast growth factors activate this receptor as measured by 45Ca2+ efflux assays. These data establish the existence of an additional member of the FGFR family that we have named FGFR-3. Images PMID:1847508

  18. Are good ideas enough? The impact of socio-economic and regulatory factors on GMO commercialisation.

    PubMed

    Vàzquez-Salat, Núria

    2013-01-01

    In recent years scientific literature has seen an increase in publications describing new transgenic applications. Although technically-sound, these promising developments might not necessarily translate into products available to the consumer. This article highlights the impact of external factors on the commercial viability of Genetically Modified (GM) animals in the pharmaceutical and food sectors. Through the division of the production chain into three Policy Domains -Science, Market and Public- I present an overview of the broad range of regulatory and socio-economic components that impacts on the path towards commercialisation of GM animals. To further illustrate the unique combination of forces that influence each application, I provide an in-depth analysis of two real cases: GM rabbits producing human polyclonal antibodies (pharmaceutical case study) and GM cows producing recombinant human lactoferrin (food case study). The inability to generalise over the commercial success of a given transgenic application should encourage researchers to perform these type of exercises early in the R & D process. Furthermore, through the analysis of these case studies we can observe a change in the biopolitics of Genetically Modified Organisms (GMOs). Contrary to the GM plant biopolitical landscape, developing states such as China and Argentina are placing themselves as global leaders in GM animals. The pro-GM attitude of these states is likely to cause a shift in the political evolution of global GMO governance. PMID:24510133

  19. Na+/H+ Exchanger Regulatory Factor 1 (NHERF1) Directly Regulates Osteogenesis*

    PubMed Central

    Liu, Li; Alonso, Veronica; Guo, Lida; Tourkova, Irina; Henderson, Sarah E.; Almarza, Alejandro J.; Friedman, Peter A.; Blair, Harry C.

    2012-01-01

    Bone formation requires synthesis, secretion, and mineralization of matrix. Deficiencies in these processes produce bone defects. The absence of the PDZ domain protein Na+/H+ exchange regulatory factor 1 (NHERF1) in mice, or its mutation in humans, causes osteomalacia believed to reflect renal phosphate wasting. We show that NHERF1 is expressed by mineralizing osteoblasts and organizes Na+/H+ exchangers (NHEs) and the PTH receptor. NHERF1-null mice display reduced bone formation and wide mineralizing fronts despite elimination of phosphate wasting by dietary supplementation. Bone mass was normal, reflecting coordinated reduction of bone resorption and formation. NHERF1-null bone had decreased strength, consistent with compromised matrix quality. Mesenchymal stem cells from NHERF1-null mice showed limited osteoblast differentiation but enhanced adipocyte differentiation. PTH signaling and Na+/H+ exchange were dysregulated in these cells. Osteoclast differentiation from monocytes was unaffected. Thus, NHERF1 is required for normal osteoblast differentiation and matrix synthesis. In its absence, compensatory mechanisms maintain bone mass, but bone strength is reduced. PMID:23109343

  20. FootprintDB: Analysis of Plant Cis-Regulatory Elements, Transcription Factors, and Binding Interfaces.

    PubMed

    Contreras-Moreira, Bruno; Sebastian, Alvaro

    2016-01-01

    FootprintDB is a database and search engine that compiles regulatory sequences from open access libraries of curated DNA cis-elements and motifs, and their associated transcription factors (TFs). It systematically annotates the binding interfaces of the TFs by exploiting protein-DNA complexes deposited in the Protein Data Bank. Each entry in footprintDB is thus a DNA motif linked to the protein sequence of the TF(s) known to recognize it, and in most cases, the set of predicted interface residues involved in specific recognition. This chapter explains step-by-step how to search for DNA motifs and protein sequences in footprintDB and how to focus the search to a particular organism. Two real-world examples are shown where this software was used to analyze transcriptional regulation in plants. Results are described with the aim of guiding users on their interpretation, and special attention is given to the choices users might face when performing similar analyses. PMID:27557773

  1. Are good ideas enough? The impact of socio-economic and regulatory factors on GMO commercialisation.

    PubMed

    Vàzquez-Salat, Núria

    2013-01-01

    In recent years scientific literature has seen an increase in publications describing new transgenic applications. Although technically-sound, these promising developments might not necessarily translate into products available to the consumer. This article highlights the impact of external factors on the commercial viability of Genetically Modified (GM) animals in the pharmaceutical and food sectors. Through the division of the production chain into three Policy Domains -Science, Market and Public- I present an overview of the broad range of regulatory and socio-economic components that impacts on the path towards commercialisation of GM animals. To further illustrate the unique combination of forces that influence each application, I provide an in-depth analysis of two real cases: GM rabbits producing human polyclonal antibodies (pharmaceutical case study) and GM cows producing recombinant human lactoferrin (food case study). The inability to generalise over the commercial success of a given transgenic application should encourage researchers to perform these type of exercises early in the R & D process. Furthermore, through the analysis of these case studies we can observe a change in the biopolitics of Genetically Modified Organisms (GMOs). Contrary to the GM plant biopolitical landscape, developing states such as China and Argentina are placing themselves as global leaders in GM animals. The pro-GM attitude of these states is likely to cause a shift in the political evolution of global GMO governance.

  2. Zinc regulates a key transcriptional pathway for epileptogenesis via metal-regulatory transcription factor 1

    PubMed Central

    van Loo, Karen M. J.; Schaub, Christina; Pitsch, Julika; Kulbida, Rebecca; Opitz, Thoralf; Ekstein, Dana; Dalal, Adam; Urbach, Horst; Beck, Heinz; Yaari, Yoel; Schoch, Susanne; Becker, Albert J.

    2015-01-01

    Temporal lobe epilepsy (TLE) is the most common focal seizure disorder in adults. In many patients, transient brain insults, including status epilepticus (SE), are followed by a latent period of epileptogenesis, preceding the emergence of clinical seizures. In experimental animals, transcriptional upregulation of CaV3.2 T-type Ca2+-channels, resulting in an increased propensity for burst discharges of hippocampal neurons, is an important trigger for epileptogenesis. Here we provide evidence that the metal-regulatory transcription factor 1 (MTF1) mediates the increase of CaV3.2 mRNA and intrinsic excitability consequent to a rise in intracellular Zn2+ that is associated with SE. Adeno-associated viral (rAAV) transfer of MTF1 into murine hippocampi leads to increased CaV3.2 mRNA. Conversely, rAAV-mediated expression of a dominant-negative MTF1 abolishes SE-induced CaV3.2 mRNA upregulation and attenuates epileptogenesis. Finally, data from resected human hippocampi surgically treated for pharmacoresistant TLE support the Zn2+-MTF1-CaV3.2 cascade, thus providing new vistas for preventing and treating TLE. PMID:26498180

  3. Activation of interferon regulatory factor 3 is inhibited by the influenza A virus NS1 protein.

    PubMed

    Talon, J; Horvath, C M; Polley, R; Basler, C F; Muster, T; Palese, P; García-Sastre, A

    2000-09-01

    We present a novel mechanism by which viruses may inhibit the alpha/beta interferon (IFN-alpha/beta) cascade. The double-stranded RNA (dsRNA) binding protein NS1 of influenza virus is shown to prevent the potent antiviral interferon response by inhibiting the activation of interferon regulatory factor 3 (IRF-3), a key regulator of IFN-alpha/beta gene expression. IRF-3 activation and, as a consequence, IFN-beta mRNA induction are inhibited in wild-type (PR8) influenza virus-infected cells but not in cells infected with an isogenic virus lacking the NS1 gene (delNS1 virus). Furthermore, NS1 is shown to be a general inhibitor of the interferon signaling pathway. Inhibition of IRF-3 activation can be achieved by the expression of wild-type NS1 in trans, not only in delNS1 virus-infected cells but also in cells infected with a heterologous RNA virus (Newcastle disease virus). We propose that inhibition of IRF-3 activation by a dsRNA binding protein significantly contributes to the virulence of influenza A viruses and possibly to that of other viruses.

  4. Interferon regulatory factor-1 polymorphisms are associated with the control of Plasmodium falciparum infection

    PubMed Central

    Mangano, Valentina D; Luoni, Gaia; Rockett, Kirk A; Sirima, Bienvenu S; Konaté, Amadou; Forton, Julian; Clark, Taane; Bancone, Germana; Akha, Elham Sadighi; Kwiatkowski, Dominic P; Modiano, David

    2010-01-01

    We describe the haplotypic structure of the Interferon Regulatory Factor-1 (IRF-1) locus in two West African ethnic groups, Fulani and Mossi, that differ in their susceptibility and immune response to Plasmodium falciparum malaria. Both populations showed significant associations between IRF-1 polymorphisms and carriage of P. falciparum infection, with different patterns of association that may reflect their different haplotypic architecture. Genetic variation at this locus does not therefore account for the Fulani-specific resistance to malaria while it could contribute to parasite clearance's ability in populations living in endemic areas. We then conducted a case-control study of three haplotype-tagging Single Nucleotide Polymorphisms (htSNPs) in 370 hospitalized malaria patients (160 severe and 210 uncomplicated) and 410 healthy population controls, all from the Mossi ethnic group. All 3 htSNPs showed correlation with blood infection levels in malaria patients, and the rs10065633 polymorphism was associated with severe disease (p=0.02). These findings provide the first evidence of the involvement in malaria susceptibility of a specific locus within the 5q31 region, previously shown to be linked with P. falciparum infection levels. PMID:18200030

  5. Immune regulatory and neuroprotective properties of preimplantation factor: From newborn to adult.

    PubMed

    Barnea, E R; Almogi-Hazan, O; Or, R; Mueller, M; Ria, F; Weiss, L; Paidas, M J

    2015-12-01

    Embryonic-maternal interaction from the earliest stages of gestation has a key, sustained role in neurologic development, persisting into adulthood. Early adverse events may be detrimental in adulthood. Protective factors present during gestation could significantly impact post-natal therapy. The role of PreImplantation Factor (PIF) within this context is herein examined. Secreted by viable early embryos, PIF establishes effective embryonic-maternal communication and exerts essential trophic and protective roles by reducing oxidative stress and protein misfolding and by blunting the nocive let-7 microRNA related pathway. PIF's effects on systemic immunity lead to comprehensive immune modulation, not immune suppression. We examine PIF's role in protecting embryos from adverse maternal environment, which can lead to neurological disorders that may only manifest post-nataly: Synthetic PIF successfully translates endogenous PIF features in both pregnant and non-pregnant clinically relevant models. Specifically PIF has neuroprotective effects in neonatal prematurity. In adult relapsing-remitting neuroinflammation, PIF reverses advanced paralysis while promoting neurogenesis. PIF reversed Mycobacterium smegmatis induced brain infection. In graft-vs.-host disease, PIF reduced skin ulceration, liver inflammation and colon ulceration while maintaining beneficial anti-cancer, graft-vs.-leukemia effect. Clinical-grade PIF has high-safety profile even at supraphysiological doses. The FDA awarded Fast-Track designation, and university-sponsored clinical trials for autoimmune disorder are ongoing. Altogether, PIF properties point to its determining regulatory role in immunity, inflammation and transplant acceptance. Specific plans for using PIF for the treatment of complex neurological disorders (ie. traumatic brain injury, progressive paralysis), including neuroprotection from newborn to adult, are presented.

  6. Characterization of Amphioxus IFN Regulatory Factor Family Reveals an Archaic Signaling Framework for Innate Immune Response.

    PubMed

    Yuan, Shaochun; Zheng, Tingting; Li, Peiyi; Yang, Rirong; Ruan, Jie; Huang, Shengfeng; Wu, Zhenxin; Xu, Anlong

    2015-12-15

    The IFN regulatory factor (IRF) family encodes transcription factors that play important roles in immune defense, stress response, reproduction, development, and carcinogenesis. Although the origin of the IRF family has been dated back to multicellular organisms, invertebrate IRFs differ from vertebrate IRFs in genomic structure and gene synteny, and little is known about their functions. Through comparison of multiple amphioxus genomes, in this study we suggested that amphioxus contains nine IRF members, whose orthologs are supposed to be shared among three amphioxus species. As the orthologs to the vertebrate IRF1 and IRF4 subgroups, Branchiostoma belcheri tsingtauense (bbt)IRF1 and bbtIRF8 bind the IFN-stimulated response element (ISRE) and were upregulated when amphioxus intestinal cells were stimulated with poly(I:C). As amphioxus-specific IRFs, both bbtIRF3 and bbtIRF7 bind ISRE. When activated, they can be phosphorylated by bbtTBK1 and then translocate into nucleus for target gene transcription. As transcriptional repressors, bbtIRF2 and bbtIRF4 can inhibit the transcriptional activities of bbtIRF1, 3, 7, and 8 by competing for the binding of ISRE. Interestingly, amphioxus IRF2, IRF8, and Rel were identified as target genes of bbtIRF1, bbtIRF7, and bbtIRF3, respectively, suggesting a dynamic feedback regulation among amphioxus IRF and NF-κB. Collectively, to our knowledge we present for the first time an archaic IRF signaling framework in a basal chordate, shedding new insights into the origin and evolution of vertebrate IFN-based antiviral networks.

  7. Contribution of Ser386 and Ser396 to activation of interferon regulatory factor 3.

    PubMed

    Chen, Weijun; Srinath, Hema; Lam, Suvana S; Schiffer, Celia A; Royer, William E; Lin, Kai

    2008-05-30

    IRF-3, a member of the interferon regulatory factor (IRF) family of transcription factors, functions in innate immune defense against viral infection. Upon infection, host cell IRF-3 is activated by phosphorylation at its seven C-terminal Ser/Thr residues: (385)SSLENTVDLHISNSHPLSLTS(405). This phosphoactivation triggers IRF-3 to react with the coactivators, CREB-binding protein (CBP)/p300, to form a complex that activates target genes in the nucleus. However, the role of each phosphorylation site for IRF-3 phosphoactivation remains unresolved. To address this issue, all seven Ser/Thr potential phosphorylation sites were screened by mutational studies, size-exclusion chromatography, and isothermal titration calorimetry. Using purified proteins, we show that CBP (amino acid residues 2067-2112) interacts directly with IRF-3 (173-427) and six of its single-site mutants to form heterodimers, but when CBP interacts with IRF-3 S396D, oligomerization is evident. CBP also interacts in vitro with IRF-3 double-site mutants to form different levels of oligomerization. Among all the single-site mutants, IRF-3 S396D showed the strongest binding to CBP. Although IRF-3 S386D alone did not interact as strongly with CBP as did other mutants, it strengthened the interaction and oligomerization of IRF-3 S396D with CBP. In contrast, IRF-3 S385D weakened the interaction and oligomerization of IRF-3 S396D and S386/396D with CBP. Thus, it appears that Ser385 and Ser386 serve antagonistic functions in regulating IRF-3 phosphoactivation. These results indicate that Ser386 and Ser396 are critical for IRF-3 activation, and support a phosphorylation-oligomerization model for IRF-3 activation.

  8. Immune regulatory and neuroprotective properties of preimplantation factor: From newborn to adult.

    PubMed

    Barnea, E R; Almogi-Hazan, O; Or, R; Mueller, M; Ria, F; Weiss, L; Paidas, M J

    2015-12-01

    Embryonic-maternal interaction from the earliest stages of gestation has a key, sustained role in neurologic development, persisting into adulthood. Early adverse events may be detrimental in adulthood. Protective factors present during gestation could significantly impact post-natal therapy. The role of PreImplantation Factor (PIF) within this context is herein examined. Secreted by viable early embryos, PIF establishes effective embryonic-maternal communication and exerts essential trophic and protective roles by reducing oxidative stress and protein misfolding and by blunting the nocive let-7 microRNA related pathway. PIF's effects on systemic immunity lead to comprehensive immune modulation, not immune suppression. We examine PIF's role in protecting embryos from adverse maternal environment, which can lead to neurological disorders that may only manifest post-nataly: Synthetic PIF successfully translates endogenous PIF features in both pregnant and non-pregnant clinically relevant models. Specifically PIF has neuroprotective effects in neonatal prematurity. In adult relapsing-remitting neuroinflammation, PIF reverses advanced paralysis while promoting neurogenesis. PIF reversed Mycobacterium smegmatis induced brain infection. In graft-vs.-host disease, PIF reduced skin ulceration, liver inflammation and colon ulceration while maintaining beneficial anti-cancer, graft-vs.-leukemia effect. Clinical-grade PIF has high-safety profile even at supraphysiological doses. The FDA awarded Fast-Track designation, and university-sponsored clinical trials for autoimmune disorder are ongoing. Altogether, PIF properties point to its determining regulatory role in immunity, inflammation and transplant acceptance. Specific plans for using PIF for the treatment of complex neurological disorders (ie. traumatic brain injury, progressive paralysis), including neuroprotection from newborn to adult, are presented. PMID:26546485

  9. Identification of the Staphylococcus aureus vfrAB Operon, a Novel Virulence Factor Regulatory Locus

    PubMed Central

    Daly, Seth M.; Hall, Pamela R.; Bayles, Kenneth W.

    2014-01-01

    During a screen of the Nebraska Transposon Mutant Library, we identified 71 mutations in the Staphylococcus aureus genome that altered hemolysis on blood agar medium. Although many of these mutations disrupted genes known to affect the production of alpha-hemolysin, two of them were associated with an apparent operon, designated vfrAB, that had not been characterized previously. Interestingly, a ΔvfrB mutant exhibited only minor effects on the transcription of the hla gene, encoding alpha-hemolysin, when grown in broth, as well as on RNAIII, a posttranscriptional regulatory RNA important for alpha-hemolysin translation, suggesting that VfrB may function at the posttranscriptional level. Indeed, a ΔvfrB mutant had increased aur and sspAB protease expression under these conditions. However, disruption of the known secreted proteases in the ΔvfrB mutant did not restore hemolytic activity in the ΔvfrB mutant on blood agar. Further analysis revealed that, in contrast to the minor effects of VfrB on hla transcription when strains were cultured in liquid media, the level of hla transcription was decreased 50-fold in the absence of VfrB on solid media. These results demonstrate that while VfrB represses protease expression when strains are grown in broth, hla regulation is highly responsive to factors associated with growth on solid media. Intriguingly, the ΔvfrB mutant displayed increased pathogenesis in a model of S. aureus dermonecrosis, further highlighting the complexity of VfrB-dependent virulence regulation. The results of this study describe a phenotype associated with a class of highly conserved yet uncharacterized proteins found in Gram-positive bacteria, and they shed new light on the regulation of virulence factors necessary for S. aureus pathogenesis. PMID:24549328

  10. Identification of the Staphylococcus aureus vfrAB operon, a novel virulence factor regulatory locus.

    PubMed

    Bose, Jeffrey L; Daly, Seth M; Hall, Pamela R; Bayles, Kenneth W

    2014-05-01

    During a screen of the Nebraska Transposon Mutant Library, we identified 71 mutations in the Staphylococcus aureus genome that altered hemolysis on blood agar medium. Although many of these mutations disrupted genes known to affect the production of alpha-hemolysin, two of them were associated with an apparent operon, designated vfrAB, that had not been characterized previously. Interestingly, a ΔvfrB mutant exhibited only minor effects on the transcription of the hla gene, encoding alpha-hemolysin, when grown in broth, as well as on RNAIII, a posttranscriptional regulatory RNA important for alpha-hemolysin translation, suggesting that VfrB may function at the posttranscriptional level. Indeed, a ΔvfrB mutant had increased aur and sspAB protease expression under these conditions. However, disruption of the known secreted proteases in the ΔvfrB mutant did not restore hemolytic activity in the ΔvfrB mutant on blood agar. Further analysis revealed that, in contrast to the minor effects of VfrB on hla transcription when strains were cultured in liquid media, the level of hla transcription was decreased 50-fold in the absence of VfrB on solid media. These results demonstrate that while VfrB represses protease expression when strains are grown in broth, hla regulation is highly responsive to factors associated with growth on solid media. Intriguingly, the ΔvfrB mutant displayed increased pathogenesis in a model of S. aureus dermonecrosis, further highlighting the complexity of VfrB-dependent virulence regulation. The results of this study describe a phenotype associated with a class of highly conserved yet uncharacterized proteins found in Gram-positive bacteria, and they shed new light on the regulation of virulence factors necessary for S. aureus pathogenesis. PMID:24549328

  11. Regulatory SNPs Alter the Gene Expression of Diabetic Retinopathy Associated Secretary Factors

    PubMed Central

    Chen, Chian-Feng; Liou, Shiow-Wen; Wu, Hsin-Han; Lin, Chin-Hui; Huang, Li-Shan; Woung, Lin-Chung; Tsai, Ching-Yao

    2016-01-01

    Objectives: Diabetic retinopathy (DR) is a common microvascular complication in both type I and type II diabetes. Several previous reports indicated the serum centration of some secretary factors were highly associated with DR. Therefore, we hypothesis regulatory SNPs (rSNPs) genotype in secretary factors may alter these gene expression and lead to DR. Methods: At first, pyrosequencing were applying to screen the SNPs which present allele frequency different in DR and DNR. Then individual genotyping was processed by Taqman assays in Taiwanese DR and DNR patients. To evaluate the effect of SNP allele on transcriptional activity, we measured promoter activity using luciferase reporter constructs. Results: We found the frequencies of the CC, CG, and GG genotype of the rs2010963 polymorphism were 15.09%, 47.14%, and 37.74% in DR and 12.90%, 19.35%, and 67.74% in DNR, respectively (p = 0.0205). The prevalence of DR was higher (p = 0.00793) in patients with the CC or CG genotype (62.26% and 32.26% for DR and DNR, respectively) compared with the patients with the GG genotype. To evaluate the effect of rs2010963-C allele on transcriptional activity, we measured promoter activity using luciferase reporter constructs. The rs2010963-C reporter showed 1.6 to 2-fold higher luciferase activity than rs2010963-G in 3 cell lines. Conclusion: Our data proposed rs2010963-C altered the expression level of VEGFA in different tissues. We suggested small increase but long term exposure to VEGFA may lead to DR finally.

  12. Characterization of Amphioxus IFN Regulatory Factor Family Reveals an Archaic Signaling Framework for Innate Immune Response.

    PubMed

    Yuan, Shaochun; Zheng, Tingting; Li, Peiyi; Yang, Rirong; Ruan, Jie; Huang, Shengfeng; Wu, Zhenxin; Xu, Anlong

    2015-12-15

    The IFN regulatory factor (IRF) family encodes transcription factors that play important roles in immune defense, stress response, reproduction, development, and carcinogenesis. Although the origin of the IRF family has been dated back to multicellular organisms, invertebrate IRFs differ from vertebrate IRFs in genomic structure and gene synteny, and little is known about their functions. Through comparison of multiple amphioxus genomes, in this study we suggested that amphioxus contains nine IRF members, whose orthologs are supposed to be shared among three amphioxus species. As the orthologs to the vertebrate IRF1 and IRF4 subgroups, Branchiostoma belcheri tsingtauense (bbt)IRF1 and bbtIRF8 bind the IFN-stimulated response element (ISRE) and were upregulated when amphioxus intestinal cells were stimulated with poly(I:C). As amphioxus-specific IRFs, both bbtIRF3 and bbtIRF7 bind ISRE. When activated, they can be phosphorylated by bbtTBK1 and then translocate into nucleus for target gene transcription. As transcriptional repressors, bbtIRF2 and bbtIRF4 can inhibit the transcriptional activities of bbtIRF1, 3, 7, and 8 by competing for the binding of ISRE. Interestingly, amphioxus IRF2, IRF8, and Rel were identified as target genes of bbtIRF1, bbtIRF7, and bbtIRF3, respectively, suggesting a dynamic feedback regulation among amphioxus IRF and NF-κB. Collectively, to our knowledge we present for the first time an archaic IRF signaling framework in a basal chordate, shedding new insights into the origin and evolution of vertebrate IFN-based antiviral networks. PMID:26573836

  13. Regulatory SNPs Alter the Gene Expression of Diabetic Retinopathy Associated Secretary Factors

    PubMed Central

    Chen, Chian-Feng; Liou, Shiow-Wen; Wu, Hsin-Han; Lin, Chin-Hui; Huang, Li-Shan; Woung, Lin-Chung; Tsai, Ching-Yao

    2016-01-01

    Objectives: Diabetic retinopathy (DR) is a common microvascular complication in both type I and type II diabetes. Several previous reports indicated the serum centration of some secretary factors were highly associated with DR. Therefore, we hypothesis regulatory SNPs (rSNPs) genotype in secretary factors may alter these gene expression and lead to DR. Methods: At first, pyrosequencing were applying to screen the SNPs which present allele frequency different in DR and DNR. Then individual genotyping was processed by Taqman assays in Taiwanese DR and DNR patients. To evaluate the effect of SNP allele on transcriptional activity, we measured promoter activity using luciferase reporter constructs. Results: We found the frequencies of the CC, CG, and GG genotype of the rs2010963 polymorphism were 15.09%, 47.14%, and 37.74% in DR and 12.90%, 19.35%, and 67.74% in DNR, respectively (p = 0.0205). The prevalence of DR was higher (p = 0.00793) in patients with the CC or CG genotype (62.26% and 32.26% for DR and DNR, respectively) compared with the patients with the GG genotype. To evaluate the effect of rs2010963-C allele on transcriptional activity, we measured promoter activity using luciferase reporter constructs. The rs2010963-C reporter showed 1.6 to 2-fold higher luciferase activity than rs2010963-G in 3 cell lines. Conclusion: Our data proposed rs2010963-C altered the expression level of VEGFA in different tissues. We suggested small increase but long term exposure to VEGFA may lead to DR finally. PMID:27648002

  14. Myogenic regulatory factor (MRF) expression is affected by exercise in postnatal chicken skeletal muscles.

    PubMed

    Yin, Huadong; Li, Diyan; Wang, Yan; Zhao, Xiaoling; Liu, Yiping; Yang, Zhiqin; Zhu, Qing

    2015-05-01

    The MyoD1, MyoG, Myf5, and Mrf4 proteins belong to the family of muscle regulatory factors (MRFs) and play important roles in skeletal muscle hyperplasia and hypertrophy. We hypothesized that exercise would affect MRF mRNA and protein abundance in postnatal chicken skeletal muscle driving molecular changes that could ultimately lead to increased muscle fiber diameter. At day (d) 43, twelve hundred chickens with similar body weight were randomly assigned to cage, pen, and free-range groups. The MRF mRNA abundance was measured in the pectoralis major and thigh muscle at d56, d70, and d84, and the protein levels of MRFs were determined from the thigh muscle at d84. The results showed no significant difference in mRNA of the MRFs among the three groups at d56 (P>0.05). At d84, chicken in the pen and free-range group showed higher MyoD1, MyoG, Myf5, and Mrf4 mRNA abundance compared to the caged chickens (P<0.05). Free-range chickens had higher Mrf4 and MyoG expression than those in penned ones (P<0.05). Protein abundances of all four factors were lowest in the caged group, and Mrf4 and MyoG protein quantities were greatest in free-range chickens (P<0.05), but Myf5 and MyoD1 protein abundance did not differ between penned and caged groups. The results suggested that exercise up-regulated MRF expression in the postnatal skeletal muscles, which led to an increase in muscle fiber diameter, and eventually affected the meat quality of the skeletal muscles in adult chickens.

  15. A functional SNP in the regulatory region of the decay-accelerating factor gene associates with extraocular muscle pareses in myasthenia gravis

    PubMed Central

    Heckmann, J M; Uwimpuhwe, H; Ballo, R; Kaur, M; Bajic, V B; Prince, S

    2009-01-01

    Complement activation in myasthenia gravis (MG) may damage muscle endplate and complement regulatory proteins such as decay-accelerating factor (DAF) or CD55 may be protective. We hypothesize that the increased prevalence of severe extraocular muscle (EOM) dysfunction among African MG subjects reported earlier may result from altered DAF expression. To test this hypothesis, we screened the DAF gene sequences relevant to the classical complement pathway and found an association between myasthenics with EOM paresis and the DAF regulatory region c.-198C>G SNP (odds ratio=8.6; P=0.0003). This single nucleotide polymorphism (SNP) results in a twofold activation of a DAF 5′-flanking region luciferase reporter transfected into three different cell lines. Direct matching of the surrounding SNP sequence within the DAF regulatory region with the known transcription factor-binding sites suggests a loss of an Sp1-binding site. This was supported by the observation that the c.-198C>G SNP did not show the normal lipopolysaccharide-induced DAF transcriptional upregulation in lymphoblasts from four patients. Our findings suggest that at critical periods during autoimmune MG, this SNP may result in inadequate DAF upregulation with consequent complement-mediated EOM damage. Susceptible individuals may benefit from anti-complement therapy in addition to immunosuppression. PMID:19675582

  16. Goblet-cell-specific transcription of mouse intestinal trefoil factor gene results from collaboration of complex series of positive and negative regulatory elements.

    PubMed Central

    Itoh, H; Inoue, N; Podolsky, D K

    1999-01-01

    Intestinal trefoil factor (ITF) is expressed selectively in intestinal goblet cells. Previous studies of the rat ITF gene identified one cis-regulatory element, designated the goblet-cell-response element (GCRE), present in the proximal region of the promoter. To identify additional cis-regulatory elements responsible for goblet-cell-specific expression, a DNA fragment containing 6353 bp of the 5'-flanking region of the mouse ITF gene was cloned and its promoter activity was examined extensively. In human and murine intestinal-derived cell lines (LS174T and CMT-93), the luciferase activities of a 6.3-kb construct were 5- and 2-fold greater than the smaller 1.8-kb construct, respectively. In contrast, the activity in non-intestinal cell lines (HepG2 and HeLa) was 2-4-fold lower than the smaller construct. In the region downstream from the 1.8-kb position, strong luciferase activities in LS174T and HepG2 cells were observed using a 201-bp construct. Interestingly, increased activity was almost completely suppressed in cells transfected with a 391-bp construct. Detailed analyses of this region revealed the existence of a 11-bp positive regulatory element (-181 to -170; ACCTCTTCCTG) and a 9-bp negative regulatory element (-208 to -200; ATTGACAGA) in addition to the GCRE. All three elements were well conserved among human, rat and mouse ITF gene promoters. In addition, a mutant 1.8-kb construct in which the negative regulatory region was deleted yielded the same approximate luciferase activity as a 6.3-kb construct, suggesting binding of a goblet-cell-specific silencer inhibitor (SI) between -6.3 and -1.8 kb. The SI present in goblet cells may block the silencers' binding to the pre-initiation complex and allow increased transcriptional activity driven by specific and non-specific enhancers. High-level expression of the mouse ITF gene specifically in intestinal goblet cells may be achieved through the combined effects of these regulatory elements. PMID:10393106

  17. Antiviral factors and type I/III interferon expression associated with regulatory factors in the oral epithelial cells from HIV-1-serodiscordant couples

    PubMed Central

    Cervantes, Cesar A. C.; Oliveira, Luanda M. S.; Manfrere, Kelly C. G.; Lima, Josenilson F.; Pereira, Natalli Z.; Duarte, Alberto J. S.; Sato, Maria N.

    2016-01-01

    Individuals who remain HIV-seronegative despite repeated unprotected exposure to the virus are defined as exposed seronegative (ESN) individuals. Innate and adaptive immunity, as well as genetic factors, provide ESNs with important advantages that allow for low infection susceptibility. The majority of HIV-1-infected individuals undergo antiretroviral therapy, which can decrease the level of HIV-1 exposure in ESNs. We analyzed type I interferon (IFN)-related antiviral and regulatory factors in peripheral blood mononuclear cells (PBMCs) and oral epithelial cells from serodiscordant couples. Our findings revealed that ESNs did not induce the expression of antiviral factors (APOBEC-3G, TRIM5-α, SAMDH1, STING, TBk1) or regulatory factors (Trex, Foxo3, Socs3, IL-10) in PBMCs, unlike their HIV-1-infected partners. In contrast, ESNs upregulated APOBEC-3G and type I/III IFNs (IFNs-α,-β/-λ) in oral mucosal epithelial cells similar to their HIV-infected partners. The serodiscordant groups exhibited an increased expression of type I IFN-induced regulators, such as Trex and Foxo3, in oral epithelial cells. TLR7, TLR8 and TLR9 were expressed in oral epithelial cells of both ESNs and HIV-1-infected subjects. These findings revealed evidence of antiviral factors, type I/III interferon and regulatory factor expression only in the oral mucosal compartment of ESNs, while HIV-1-infected partners systemically and oral mucosal expressed the antiviral profile. PMID:27168019

  18. Dynamic changes in binding of immunoglobulin heavy chain 3' regulatory region to protein factors during class switching.

    PubMed

    Chatterjee, Sanjukta; Ju, Zhongliang; Hassan, Rabih; Volpi, Sabrina A; Emelyanov, Alexander V; Birshtein, Barbara K

    2011-08-19

    The 3' regulatory region (3' RR) of the Igh locus works at long distances on variable region (V(H)) and switch region (I) region promoters to initiate germ line (non-coding) transcription (GT) and promote class switch recombination (CSR). The 3' RR contains multiple elements, including enhancers (hs3a, hs1.2, hs3b, and hs4) and a proposed insulator region containing CTCF (CCCTC-binding factor) binding sites, i.e. hs5/6/7 and the downstream region ("38"). Notably, deletion of each individual enhancer (hs3a-hs4) has no significant phenotypic consequence, suggesting that the 3' RR has considerable structural flexibility in its function. To better understand how the 3' RR functions, we identified transcription factor binding sites and used chromatin immunoprecipitation (ChIP) assays to monitor their occupancy in splenic B cells that initiate GT and undergo CSR (LPS±IL4), are deficient in GT and CSR (p50(-/-)), or do not undergo CSR despite efficient GT (anti-IgM+IL4). Like 3' RR enhancers, hs5-7 and the 38 region were observed to contain multiple Pax5 binding sites (in addition to multiple CTCF sites). We found that the Pax5 binding profile to the 3' RR dynamically changed during CSR independent of the specific isotype to which switching was induced, and binding focused on hs1.2, hs4, and hs7. CTCF-associated and CTCF-independent cohesin interactions were also identified. Our observations are consistent with a scaffold model in which a platform of active protein complexes capable of facilitating GT and CSR can be formed by varying constellations of 3' RR elements.

  19. Activation of Sterol Regulatory Element Binding Factors by Fenofibrate and Gemfibrozil Stimulates Myelination in Zebrafish.

    PubMed

    Ashikawa, Yoshifumi; Nishimura, Yuhei; Okabe, Shiko; Sasagawa, Shota; Murakami, Soichiro; Yuge, Mizuki; Kawaguchi, Koki; Kawase, Reiko; Tanaka, Toshio

    2016-01-01

    Oligodendrocytes are major myelin-producing cells and play essential roles in the function of a healthy nervous system. However, they are also one of the most vulnerable neural cell types in the central nervous system (CNS), and myelin abnormalities in the CNS are found in a wide variety of neurological disorders, including multiple sclerosis, adrenoleukodystrophy, and schizophrenia. There is an urgent need to identify small molecular weight compounds that can stimulate myelination. In this study, we performed comparative transcriptome analysis to identify pharmacodynamic effects common to miconazole and clobetasol, which have been shown to stimulate myelination by mouse oligodendrocyte progenitor cells (OPCs). Of the genes differentially expressed in both miconazole- and clobetasol-treated mouse OPCs compared with untreated cells, we identified differentially expressed genes (DEGs) common to both drug treatments. Gene ontology analysis revealed that these DEGs are significantly associated with the sterol biosynthetic pathway, and further bioinformatics analysis suggested that sterol regulatory element binding factors (SREBFs) might be key upstream regulators of the DEGs. In silico screening of a public database for chemicals associated with SREBF activation identified fenofibrate, a peroxisome proliferator-activated receptor α (PPARα) agonist, as a drug that increases the expression of known SREBF targets, raising the possibility that fenofibrate may also stimulate myelination. To test this, we performed in vivo imaging of zebrafish expressing a fluorescent reporter protein under the control of the myelin basic protein (mbp) promoter. Treatment of zebrafish with fenofibrate significantly increased expression of the fluorescent reporter compared with untreated zebrafish. This increase was attenuated by co-treatment with fatostatin, a specific inhibitor of SREBFs, confirming that the fenofibrate effect was mediated via SREBFs. Furthermore, incubation of zebrafish

  20. Activation of Sterol Regulatory Element Binding Factors by Fenofibrate and Gemfibrozil Stimulates Myelination in Zebrafish

    PubMed Central

    Ashikawa, Yoshifumi; Nishimura, Yuhei; Okabe, Shiko; Sasagawa, Shota; Murakami, Soichiro; Yuge, Mizuki; Kawaguchi, Koki; Kawase, Reiko; Tanaka, Toshio

    2016-01-01

    Oligodendrocytes are major myelin-producing cells and play essential roles in the function of a healthy nervous system. However, they are also one of the most vulnerable neural cell types in the central nervous system (CNS), and myelin abnormalities in the CNS are found in a wide variety of neurological disorders, including multiple sclerosis, adrenoleukodystrophy, and schizophrenia. There is an urgent need to identify small molecular weight compounds that can stimulate myelination. In this study, we performed comparative transcriptome analysis to identify pharmacodynamic effects common to miconazole and clobetasol, which have been shown to stimulate myelination by mouse oligodendrocyte progenitor cells (OPCs). Of the genes differentially expressed in both miconazole- and clobetasol-treated mouse OPCs compared with untreated cells, we identified differentially expressed genes (DEGs) common to both drug treatments. Gene ontology analysis revealed that these DEGs are significantly associated with the sterol biosynthetic pathway, and further bioinformatics analysis suggested that sterol regulatory element binding factors (SREBFs) might be key upstream regulators of the DEGs. In silico screening of a public database for chemicals associated with SREBF activation identified fenofibrate, a peroxisome proliferator-activated receptor α (PPARα) agonist, as a drug that increases the expression of known SREBF targets, raising the possibility that fenofibrate may also stimulate myelination. To test this, we performed in vivo imaging of zebrafish expressing a fluorescent reporter protein under the control of the myelin basic protein (mbp) promoter. Treatment of zebrafish with fenofibrate significantly increased expression of the fluorescent reporter compared with untreated zebrafish. This increase was attenuated by co-treatment with fatostatin, a specific inhibitor of SREBFs, confirming that the fenofibrate effect was mediated via SREBFs. Furthermore, incubation of zebrafish

  1. Activation of Sterol Regulatory Element Binding Factors by Fenofibrate and Gemfibrozil Stimulates Myelination in Zebrafish.

    PubMed

    Ashikawa, Yoshifumi; Nishimura, Yuhei; Okabe, Shiko; Sasagawa, Shota; Murakami, Soichiro; Yuge, Mizuki; Kawaguchi, Koki; Kawase, Reiko; Tanaka, Toshio

    2016-01-01

    Oligodendrocytes are major myelin-producing cells and play essential roles in the function of a healthy nervous system. However, they are also one of the most vulnerable neural cell types in the central nervous system (CNS), and myelin abnormalities in the CNS are found in a wide variety of neurological disorders, including multiple sclerosis, adrenoleukodystrophy, and schizophrenia. There is an urgent need to identify small molecular weight compounds that can stimulate myelination. In this study, we performed comparative transcriptome analysis to identify pharmacodynamic effects common to miconazole and clobetasol, which have been shown to stimulate myelination by mouse oligodendrocyte progenitor cells (OPCs). Of the genes differentially expressed in both miconazole- and clobetasol-treated mouse OPCs compared with untreated cells, we identified differentially expressed genes (DEGs) common to both drug treatments. Gene ontology analysis revealed that these DEGs are significantly associated with the sterol biosynthetic pathway, and further bioinformatics analysis suggested that sterol regulatory element binding factors (SREBFs) might be key upstream regulators of the DEGs. In silico screening of a public database for chemicals associated with SREBF activation identified fenofibrate, a peroxisome proliferator-activated receptor α (PPARα) agonist, as a drug that increases the expression of known SREBF targets, raising the possibility that fenofibrate may also stimulate myelination. To test this, we performed in vivo imaging of zebrafish expressing a fluorescent reporter protein under the control of the myelin basic protein (mbp) promoter. Treatment of zebrafish with fenofibrate significantly increased expression of the fluorescent reporter compared with untreated zebrafish. This increase was attenuated by co-treatment with fatostatin, a specific inhibitor of SREBFs, confirming that the fenofibrate effect was mediated via SREBFs. Furthermore, incubation of zebrafish

  2. Cloning and expression analysis of interferon regulatory factor 7 in the Pacific cod, Gadus macrocephalus.

    PubMed

    Sun, Hang; Jiang, Zhiqiang; Mao, Mingguang; Huo, Yuan; Han, Yuzhe; Zhang, Saisai

    2016-02-01

    Interferon regulatory factor 7 (IRF7) plays an important role in regulating the response of type I interferon (IFN) to viral infection. To understand the mechanisms underlying immune reactions in the Pacific cod, Gadus macrocephalus, the gene encoding G. macrocephalus IRF7 was cloned and characterized. The cDNA of G. macrocephalus IRF7 was also cloned and sequenced. A cDNA sequence of 2032 bp was assembled using polymerase chain reaction (PCR) products. It contains an open reading frame of 1323 bp in length, which encoded a 440-amino acid polypeptide that comprised a DNA-binding domain (DBD), an IRF association domain (IAD), and a serine-rich domain (SRD). In the DBD, the tryptophan cluster consisted of only four tryptophans, which is a unique characteristic in fish IRF7. The mRNA of IRF7 was detected in various tissues, including in the spleen, thymus, kidney, intestine, and gills, using relative quantification PCR (R-qPCR). Dynamic expression of IRF7 was observed in larvae throughout post-hatching (ph) development, with the highest level detected at day of ph (dph) 25. Response to immune stimulation was examined by challenging larvae with polyriboinosinic polyribocytidylic acid (pIC) to mimic viral infection and elicit an immune reaction. R-qPCR revealed that the expression of IRF7 significantly increased in pIC-treated groups relative to that in the control groups, in a time-dependent manner, with peak responses at 48 and 72 h after pIC-treatment. These results show that IRF7 is expressed in various tissues of adult fish and larvae and is sensitive to viral infection, suggesting that it plays a role in antiviral immune defense in G. macrocephalus. PMID:26702560

  3. S. macrurus myogenic regulatory factors induce mammalian skeletal muscle differentiation: Evidence for functional conservation of MRFs

    PubMed Central

    Kim, Hyun-Jung; Güth, Robert; Jonsson, Colleen B.; Unguez, Graciela A.

    2009-01-01

    The current-producing cells of the electric organ (EO), i.e., electrocytes, in Sternopygus macrurus derive from skeletal muscle fibers. Mature electrocytes are not contractile but they do retain some muscle proteins, are multinucleated, and receive cholinergic innervation. Electrocytes express the myogenic regulatory factors (MRFs) MyoD, myogenin, Myf5 and MRF4 despite their incomplete muscle phenotype. Although S. macrurus MRFs share functional domains that are highly conserved and their expression is confined to the myogenic lineage, their capability to induce the muscle phenotype has not been determined. To test the functional conservation of S. macrurus MRFs to transcriptionally activate skeletal muscle gene expression and induce the myogenic program, we transiently over-expressed S. macrurus MyoD (SmMyoD) and myogenin (SmMyoG) in mouse C3H/10T1/2 and NIH3T3 embryonic cells. RT-PCR and immunolabeling studies showed that SmMyoD and SmMyoG efficiently can convert these two cell lines into multinucleated myotubes that expressed differentiated muscle markers. The levels of myogenic induction by SmMyoD and SmMyoG were comparable to those obtained with mouse MRF homologs. Furthermore, SmMyoD and SmMyoG proteins were able to induce mouse MyoD and myogenin in C3H/10T1/2 cells. We conclude that S. macrurus MRFs are functionally conserved as they can transcriptionally activate skeletal muscle gene expression and induce the myogenic program in mammalian non-muscle cells. Hence, these data suggest that the partial muscle phenotype of electrocytes is not likely due to differences in the MRF-dependent transcriptional program between skeletal muscle and electric organ. PMID:19598116

  4. Molecular cloning and characterization of interferon regulatory factor 9 (IRF9) in Japanese flounder, Paralichthys olivaceus.

    PubMed

    Hu, Guo-Bin; Zhao, Ming-Yu; Lin, Jing-Yun; Liu, Qiu-Ming; Zhang, Shi-Cui

    2014-08-01

    Interferon regulatory factor 9 (IRF9) in mammals is known to be involved in antiviral response. In this study, we studied the structure, mRNA tissue distribution and regulation of IRF9 from Japanese flounder, Paralichthys olivaceus. The cDNA sequence of IRF9 is 3305 bp long, containing an open reading frame (ORF) of 1308 bp that encodes a peptide of 435 amino acids. The predicted protein sequence shares 33.7-72.0% identity to other fish IRF9s. Japanese flounder IRF9 possesses a DNA-binding domain (DBD), an IRF association domain (IAD), two nuclear localization signals (NLSs) and a proline-rich domain (PRD). The IRF9 transcripts were detectable in all examined tissues of healthy Japanese flounders, with higher levels in the head kidney, kidney, liver and spleen. The IRF9 mRNA levels were up-regulated in the gills, head kidney, spleen and muscle when challenged with polyinosinic:polycytidylic acid (poly I:C) or lymphocystis disease virus (LCDV). The up-regulations were stronger and arose earlier in the case of poly I:C treatment in most tested organs in a 7-day time course, with maximum increases ranging from 1.37- to 8.59-fold and peak time points from 3 h to 3 d post injection depending on different organs, relative to those in the case of LCDV treatment which ranged from 1.32- to 3.21-fold and from 18 h to 3 d post injection, respectively. The highest and earliest inductions were detected in the spleen in both challenge cases, while the inductions by LCDV in the muscle were quite faint. These results demonstrate a role of Japanese flounder IRF9 in the host's antiviral responses. PMID:24837327

  5. Construction of pancreatic cancer double-factor regulatory network based on chip data on the transcriptional level.

    PubMed

    Zhao, Li-Li; Zhang, Tong; Liu, Bing-Rong; Liu, Tie-Fu; Tao, Na; Zhuang, Li-Wei

    2014-05-01

    Transcription factor (TF) and microRNA (miRNA) have been discovered playing crucial roles in cancer development. However, the effect of TFs and miRNAs in pancreatic cancer pathogenesis remains vague. We attempted to reveal the possible mechanism of pancreatic cancer based on transcription level. Using GSE16515 datasets downloaded from gene expression omnibus database, we first identified the differentially expressed genes (DEGs) in pancreatic cancer by the limma package in R. Then the DEGs were mapped into DAVID to conduct the kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis. TFs and miRNAs that DEGs significantly enriched were identified by Fisher's test, and then the pancreatic cancer double-factor regulatory network was constructed. In our study, total 1117 DEGs were identified and they significantly enriched in 4 KEGG pathways. A double-factor regulatory network was established, including 29 DEGs, 24 TFs, 25 miRNAs. In the network, LAMC2, BRIP1 and miR155 were identified which may be involved in pancreatic cancer development. In conclusion, the double-factor regulatory network was found to play an important role in pancreatic cancer progression and our results shed new light on the molecular mechanism of pancreatic cancer.

  6. Three WRKY transcription factors additively repress abscisic acid and gibberellin signaling in aleurone cells.

    PubMed

    Zhang, Liyuan; Gu, Lingkun; Ringler, Patricia; Smith, Stanley; Rushton, Paul J; Shen, Qingxi J

    2015-07-01

    Members of the WRKY transcription factor superfamily are essential for the regulation of many plant pathways. Functional redundancy due to duplications of WRKY transcription factors, however, complicates genetic analysis by allowing single-mutant plants to maintain wild-type phenotypes. Our analyses indicate that three group I WRKY genes, OsWRKY24, -53, and -70, act in a partially redundant manner. All three showed characteristics of typical WRKY transcription factors: each localized to nuclei and yeast one-hybrid assays indicated that they all bind to W-boxes, including those present in their own promoters. Quantitative real time-PCR (qRT-PCR) analyses indicated that the expression levels of the three WRKY genes varied in the different tissues tested. Particle bombardment-mediated transient expression analyses indicated that all three genes repress the GA and ABA signaling in a dosage-dependent manner. Combination of all three WRKY genes showed additive antagonism of ABA and GA signaling. These results suggest that these WRKY proteins function as negative transcriptional regulators of GA and ABA signaling. However, different combinations of these WRKY genes can lead to varied strengths in suppression of their targets.

  7. SNPs in putative regulatory regions identified by human mouse comparative sequencing and transcription factor binding site data

    SciTech Connect

    Banerjee, Poulabi; Bahlo, Melanie; Schwartz, Jody R.; Loots, Gabriela G.; Houston, Kathryn A.; Dubchak, Inna; Speed, Terence P.; Rubin, Edward M.

    2002-01-01

    Genome wide disease association analysis using SNPs is being explored as a method for dissecting complex genetic traits and a vast number of SNPs have been generated for this purpose. As there are cost and throughput limitations of genotyping large numbers of SNPs and statistical issues regarding the large number of dependent tests on the same data set, to make association analysis practical it has been proposed that SNPs should be prioritized based on likely functional importance. The most easily identifiable functional SNPs are coding SNPs (cSNPs) and accordingly cSNPs have been screened in a number of studies. SNPs in gene regulatory sequences embedded in noncoding DNA are another class of SNPs suggested for prioritization due to their predicted quantitative impact on gene expression. The main challenge in evaluating these SNPs, in contrast to cSNPs is a lack of robust algorithms and databases for recognizing regulatory sequences in noncoding DNA. Approaches that have been previously used to delineate noncoding sequences with gene regulatory activity include cross-species sequence comparisons and the search for sequences recognized by transcription factors. We combined these two methods to sift through mouse human genomic sequences to identify putative gene regulatory elements and subsequently localized SNPs within these sequences in a 1 Megabase (Mb) region of human chromosome 5q31, orthologous to mouse chromosome 11 containing the Interleukin cluster.

  8. Genome-wide survey of tissue-specific microRNA and transcription factor regulatory networks in 12 tissues.

    PubMed

    Guo, Zhiyun; Maki, Miranda; Ding, Ruofan; Yang, Yalan; Zhang, Bao; Xiong, Lili

    2014-06-03

    Tissue-specific miRNAs (TS miRNA) specifically expressed in particular tissues play an important role in tissue identity, differentiation and function. However, transcription factor (TF) and TS miRNA regulatory networks across multiple tissues have not been systematically studied. Here, we manually extracted 116 TS miRNAs and systematically investigated the regulatory network of TF-TS miRNA in 12 human tissues. We identified 2,347 TF-TS miRNA regulatory relations and revealed that most TF binding sites tend to enrich close to the transcription start site of TS miRNAs. Furthermore, we found TS miRNAs were regulated widely by non-tissue specific TFs and the tissue-specific expression level of TF have a close relationship with TF-genes regulation. Finally, we describe TSmiR (http://bioeng.swjtu.edu.cn/TSmiR), a novel and web-searchable database that houses interaction maps of TF-TS miRNA in 12 tissues. Taken together, these observations provide a new suggestion to better understand the regulatory network and mechanisms of TF-TS miRNAs underlying different tissues.

  9. Genetic Variation of Goat Interferon Regulatory Factor 3 Gene and Its Implication in Goat Evolution

    PubMed Central

    Shu, Liping; Zhang, Yesheng; Wang, Yangzi; Sanni, Timothy M.; Imumorin, Ikhide G.; Peters, Sunday O.; Zhang, Jiajin; Dong, Yang; Wang, Wen

    2016-01-01

    The immune systems are fundamentally vital for evolution and survival of species; as such, selection patterns in innate immune loci are of special interest in molecular evolutionary research. The interferon regulatory factor (IRF) gene family control many different aspects of the innate and adaptive immune responses in vertebrates. Among these, IRF3 is known to take active part in very many biological processes. We assembled and evaluated 1356 base pairs of the IRF3 gene coding region in domesticated goats from Africa (Nigeria, Ethiopia and South Africa) and Asia (Iran and China) and the wild goat (Capra aegagrus). Five segregating sites with θ value of 0.0009 for this gene demonstrated a low diversity across the goats’ populations. Fu and Li tests were significantly positive but Tajima’s D test was significantly negative, suggesting its deviation from neutrality. Neighbor joining tree of IRF3 gene in domesticated goats, wild goat and sheep showed that all domesticated goats have a closer relationship than with the wild goat and sheep. Maximum likelihood tree of the gene showed that different domesticated goats share a common ancestor and suggest single origin. Four unique haplotypes were observed across all the sequences, of which, one was particularly common to African goats (MOCH-K14-0425, Poitou and WAD). In assessing the evolution mode of the gene, we found that the codon model dN/dS ratio for all goats was greater than one. Phylogenetic Analysis by Maximum Likelihood (PAML) gave a ω0 (dN/dS) value of 0.067 with LnL value of -6900.3 for the first Model (M1) while ω2 = 1.667 in model M2 with LnL value of -6900.3 with positive selection inferred in 3 codon sites. Mechanistic empirical combination (MEC) model for evaluating adaptive selection pressure on particular codons also confirmed adaptive selection pressure in three codons (207, 358 and 408) in IRF3 gene. Positive diversifying selection inferred with recent evolutionary changes in domesticated goat

  10. Interferon regulatory factor 1 and histone H4 acetylation in systemic lupus erythematosus

    PubMed Central

    Leung, Yiu Tak; Shi, Lihua; Maurer, Kelly; Song, Li; Zhang, Zhe; Petri, Michelle; Sullivan, Kathleen E

    2015-01-01

    Histone acetylation modulates gene expression and has been described as increased in systemic lupus erythematosus (SLE). We investigated interferon regulatory factor 1 (IRF1) interactions that influence H4 acetylation (H4ac) in SLE. Intracellular flow cytometry for H4 acetylated lysine (K) 5, K8, K12, and K16 was performed. Histone acetylation was defined in monocytes and T cells from controls and SLE patients. RNA-Seq studies were performed on monocytes to look for an imbalance in histone acetyltransferases and histone deacetylase enzyme expression. Expression levels were validated using real-time quantitative RT-PCR. IRF1 induction of H4ac was evaluated using D54MG cells overexpressing IRF1. IRF1 protein interactions were studied using co-immunoprecipitation assays. IRF1-dependent recruitment of histone acetyltransferases to target genes was examined by ChIP assays using p300 antibody. Flow cytometry data showed significantly increased H4K5, H4K8, H4K12, and H4K16 acetylation in SLE monocytes. HDAC3 and HDAC11 gene expression were decreased in SLE monocytes. PCAF showed significantly higher gene expression in SLE than controls. IRF1-overexpressing D54MG cells were associated with significantly increased H4K5, H4K8, and H4K12 acetylation compared to vector-control D54MG cells both globally and at specific target genes. Co-immunoprecipitation studies using D54MG cells revealed IRF1 protein-protein interactions with PCAF, P300, CBP, GCN5, ATF2, and HDAC3. ChIP experiments demonstrated increased p300 recruitment to known IRF1 targets in D54MG cells overexpressing IRF1. In contrast, p300 binding to IRF1 targets decreased in D54MG cells with IRF1 knockdown. SLE appears to be associated with an imbalance in histone acetyltransferases and histone deacetylase enzymes favoring pathologic H4 acetylation. Furthermore, IRF1 directly interacts with chromatin modifying enzymes, supporting a model where recruitment to specific target genes is mediated in part by IRF1. PMID

  11. Genetic Variation of Goat Interferon Regulatory Factor 3 Gene and Its Implication in Goat Evolution.

    PubMed

    Okpeku, Moses; Esmailizadeh, Ali; Adeola, Adeniyi C; Shu, Liping; Zhang, Yesheng; Wang, Yangzi; Sanni, Timothy M; Imumorin, Ikhide G; Peters, Sunday O; Zhang, Jiajin; Dong, Yang; Wang, Wen

    2016-01-01

    The immune systems are fundamentally vital for evolution and survival of species; as such, selection patterns in innate immune loci are of special interest in molecular evolutionary research. The interferon regulatory factor (IRF) gene family control many different aspects of the innate and adaptive immune responses in vertebrates. Among these, IRF3 is known to take active part in very many biological processes. We assembled and evaluated 1356 base pairs of the IRF3 gene coding region in domesticated goats from Africa (Nigeria, Ethiopia and South Africa) and Asia (Iran and China) and the wild goat (Capra aegagrus). Five segregating sites with θ value of 0.0009 for this gene demonstrated a low diversity across the goats' populations. Fu and Li tests were significantly positive but Tajima's D test was significantly negative, suggesting its deviation from neutrality. Neighbor joining tree of IRF3 gene in domesticated goats, wild goat and sheep showed that all domesticated goats have a closer relationship than with the wild goat and sheep. Maximum likelihood tree of the gene showed that different domesticated goats share a common ancestor and suggest single origin. Four unique haplotypes were observed across all the sequences, of which, one was particularly common to African goats (MOCH-K14-0425, Poitou and WAD). In assessing the evolution mode of the gene, we found that the codon model dN/dS ratio for all goats was greater than one. Phylogenetic Analysis by Maximum Likelihood (PAML) gave a ω0 (dN/dS) value of 0.067 with LnL value of -6900.3 for the first Model (M1) while ω2 = 1.667 in model M2 with LnL value of -6900.3 with positive selection inferred in 3 codon sites. Mechanistic empirical combination (MEC) model for evaluating adaptive selection pressure on particular codons also confirmed adaptive selection pressure in three codons (207, 358 and 408) in IRF3 gene. Positive diversifying selection inferred with recent evolutionary changes in domesticated goat IRF3

  12. Mechanisms of immune suppression by interleukin-10 and transforming growth factor-β: the role of T regulatory cells

    PubMed Central

    Taylor, Alison; Verhagen, Johan; Blaser, Kurt; Akdis, Mübeccel; Akdis, Cezmi A

    2006-01-01

    Specific immune suppression and induction of tolerance are essential processes in the regulation and circumvention of immune defence. The balance between allergen-specific type 1 regulatory (Tr1) cells and T helper (Th) 2 cells appears to be decisive in the development of allergy. Tr1 cells consistently represent the dominant subset specific for common environmental allergens in healthy individuals. In contrast, there is a high frequency of allergen-specific interleukin-4 (IL-4)-secreting T cells in allergic individuals. Allergen-specific immunotherapy can induce specific Tr1 cells that abolish allergen-induced proliferation of Th1 and Th2 cells, as well as their cytokine production. Tr1 cells utilize multiple suppressor mechanisms, such as IL-10 and transforming growth factor-β (TGF-β) as secreted cytokines and various surface molecules, such as cytotoxic T-lymphocyte antigen 4 and programmed death-1. IL-10 only inhibits T cells stimulated by low numbers of triggered T-cell receptors, which depend on CD28 costimulation. IL-10 inhibits CD28 tyrosine phosphorylation, preventing the binding of phosphatidylinositol 3-kinase p85 and consequently inhibiting the CD28 signalling pathway. In addition, IL-10 and TGF-β secreted by Tr1 cells skew the antibody production from immunoglobulin E (IgE) towards the non-inflammatory isotypes IgG4 and IgA, respectively. Induction of antigen-specific Tr1 cells can thus re-direct an inappropriate immune response against allergens or auto-antigens using a broad range of suppressor mechanisms. PMID:16556256

  13. A Regulatory Circuit Composed of a Transcription Factor, IscR, and a Regulatory RNA, RyhB, Controls Fe-S Cluster Delivery

    PubMed Central

    Chareyre, Sylvia; Barras, Frédéric

    2016-01-01

    ABSTRACT Fe-S clusters are cofactors conserved through all domains of life. Once assembled by dedicated ISC and/or SUF scaffolds, Fe-S clusters are conveyed to their apo-targets via A-type carrier proteins (ATCs). Escherichia coli possesses four such ATCs. ErpA is the only ATC essential under aerobiosis. Recent studies reported a possible regulation of the erpA mRNA by the small RNA (sRNA) RyhB, which controls the expression of many genes under iron starvation. Surprisingly, erpA has not been identified in recent transcriptomic analysis of the iron starvation response, thus bringing into question the actual physiological significance of the putative regulation of erpA by RyhB. Using an sRNA library, we show that among 26 sRNAs, only RyhB represses the expression of an erpA-lacZ translational fusion. We further demonstrate that this repression occurs during iron starvation. Using mutational analysis, we show that RyhB base pairs to the erpA mRNA, inducing its disappearance. In addition, IscR, the master regulator of Fe-S homeostasis, represses expression of erpA at the transcriptional level when iron is abundant, but depleting iron from the medium alleviates this repression. The conjunction of transcriptional derepression by IscR and posttranscriptional repression by RyhB under Fe-limiting conditions is best described as an incoherent regulatory circuit. This double regulation allows full expression of erpA at iron concentrations for which Fe-S biogenesis switches from the ISC to the SUF system. We further provide evidence that this regulatory circuit coordinates ATC usage to iron availability. PMID:27651365

  14. Regulatory Lymphocytes Are Key Factors in MHC-Independent Resistance to EAE

    PubMed Central

    Marín, Nieves; Mecha, Miriam; Espejo, Carmen; Mestre, Leyre; Eixarch, Herena; Montalban, Xavier; Álvarez-Cermeño, José C.; Guaza, Carmen; Villar, Luisa M.

    2014-01-01

    Background and Objectives. Resistant and susceptible mouse strains to experimental autoimmune encephalomyelitis (EAE), an inducible demyelinating experimental disease serving as animal model for multiple sclerosis, have been described. We aimed to explore MHC-independent mechanisms inducing resistance to EAE. Methods. For EAE induction, female C57BL/6 (susceptible strain) and CD1 (resistant outbred strain showing heterogeneous MHC antigens) mice were immunized with the 35–55 peptide of myelin oligodendrocyte glycoprotein (MOG35−55). We studied T cell proliferation, regulatory and effector cell subpopulations, intracellular and serum cytokine patterns, and titers of anti-MOG serum antibodies. Results. Upon immunization with MOG35−55, T lymphocytes from susceptible mice but not that of resistant strain were capable of proliferating when stimulated with MOG35−55. Accordingly, resistant mice experienced a rise in regulatory B cells (P = 0.001) and, to a lower extent, in regulatory T cells (P = 0.02) compared with C57BL/6 susceptible mice. As a consequence, MOG35−55-immunized C57BL/6 mice showed higher percentages of CD4+ T cells producing both IFN-gamma (P = 0.02) and IL-17 (P = 0.009) and higher serum levels of IL-17 (P = 0.04) than resistant mice. Conclusions. Expansion of regulatory B and T cells contributes to the induction of resistance to EAE by an MHC-independent mechanism. PMID:24868560

  15. Activation of Vago by interferon regulatory factor (IRF) suggests an interferon system-like antiviral mechanism in shrimp.

    PubMed

    Li, Chaozheng; Li, Haoyang; Chen, Yixiao; Chen, Yonggui; Wang, Sheng; Weng, Shao-Ping; Xu, Xiaopeng; He, Jianguo

    2015-01-01

    There is a debate on whether invertebrates possess an antiviral immunity similar to the interferon (IFN) system of vertebrates. The Vago gene from arthropods encodes a viral-activated secreted peptide that restricts virus infection through activating the JAK-STAT pathway and is considered to be a cytokine functionally similar to IFN. In this study, the first crustacean IFN regulatory factor (IRF)-like gene was identified in Pacific white shrimp, Litopenaeus vannamei. The L. vannamei IRF showed similar protein nature to mammalian IRFs and could be activated during virus infection. As a transcriptional regulatory factor, L. vannamei IRF could activate the IFN-stimulated response element (ISRE)-containing promoter to regulate the expression of mammalian type I IFNs and initiate an antiviral state in mammalian cells. More importantly, IRF could bind the 5'-untranslated region of L. vannamei Vago4 gene and activate its transcription, suggesting that shrimp Vago may be induced in a similar manner to that of IFNs and supporting the opinion that Vago might function as an IFN-like molecule in invertebrates. These suggested that shrimp might possess an IRF-Vago-JAK/STAT regulatory axis, which is similar to the IRF-IFN-JAK/STAT axis of vertebrates, indicating that invertebrates might possess an IFN system-like antiviral mechanism.

  16. Activation of Vago by interferon regulatory factor (IRF) suggests an interferon system-like antiviral mechanism in shrimp

    PubMed Central

    Li, Chaozheng; Li, Haoyang; Chen, Yixiao; Chen, Yonggui; Wang, Sheng; Weng, Shao-Ping; Xu, Xiaopeng; He, Jianguo

    2015-01-01

    There is a debate on whether invertebrates possess an antiviral immunity similar to the interferon (IFN) system of vertebrates. The Vago gene from arthropods encodes a viral-activated secreted peptide that restricts virus infection through activating the JAK-STAT pathway and is considered to be a cytokine functionally similar to IFN. In this study, the first crustacean IFN regulatory factor (IRF)-like gene was identified in Pacific white shrimp, Litopenaeus vannamei. The L. vannamei IRF showed similar protein nature to mammalian IRFs and could be activated during virus infection. As a transcriptional regulatory factor, L. vannamei IRF could activate the IFN-stimulated response element (ISRE)-containing promoter to regulate the expression of mammalian type I IFNs and initiate an antiviral state in mammalian cells. More importantly, IRF could bind the 5′-untranslated region of L. vannamei Vago4 gene and activate its transcription, suggesting that shrimp Vago may be induced in a similar manner to that of IFNs and supporting the opinion that Vago might function as an IFN-like molecule in invertebrates. These suggested that shrimp might possess an IRF-Vago-JAK/STAT regulatory axis, which is similar to the IRF-IFN-JAK/STAT axis of vertebrates, indicating that invertebrates might possess an IFN system-like antiviral mechanism. PMID:26459861

  17. Factors which Limit the Value of Additional Redundancy in Human Rated Launch Vehicle Systems

    NASA Technical Reports Server (NTRS)

    Anderson, Joel M.; Stott, James E.; Ring, Robert W.; Hatfield, Spencer; Kaltz, Gregory M.

    2008-01-01

    The National Aeronautics and Space Administration (NASA) has embarked on an ambitious program to return humans to the moon and beyond. As NASA moves forward in the development and design of new launch vehicles for future space exploration, it must fully consider the implications that rule-based requirements of redundancy or fault tolerance have on system reliability/risk. These considerations include common cause failure, increased system complexity, combined serial and parallel configurations, and the impact of design features implemented to control premature activation. These factors and others must be considered in trade studies to support design decisions that balance safety, reliability, performance and system complexity to achieve a relatively simple, operable system that provides the safest and most reliable system within the specified performance requirements. This paper describes conditions under which additional functional redundancy can impede improved system reliability. Examples from current NASA programs including the Ares I Upper Stage will be shown.

  18. Regulation of Peroxisome Proliferator-Activated Receptor γ Expression by Adipocyte Differentiation and Determination Factor 1/Sterol Regulatory Element Binding Protein 1: Implications for Adipocyte Differentiation and Metabolism

    PubMed Central

    Fajas, Lluis; Schoonjans, Kristina; Gelman, Laurent; Kim, Jae B.; Najib, Jamila; Martin, Genevieve; Fruchart, Jean-Charles; Briggs, Michael; Spiegelman, Bruce M.; Auwerx, Johan

    1999-01-01

    Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor implicated in adipocyte differentiation and insulin sensitivity. We investigated whether PPARγ expression is dependent on the activity of adipocyte differentiation and determination factor 1/sterol regulatory element binding protein 1 (ADD-1/SREBP-1), another transcription factor associated with both adipocyte differentiation and cholesterol homeostasis. Ectopic expression of ADD-1/SREBP-1 in 3T3-L1 and HepG2 cells induced endogenous PPARγ mRNA levels. The related transcription factor SREBP-2 likewise induced PPARγ expression. In addition, cholesterol depletion, a condition known to result in proteolytic activation of transcription factors of the SREBP family, induced PPARγ expression and improved PPRE-driven transcription. The effect of the SREBPs on PPARγ expression was mediated through the PPARγ1 and -3 promoters. Both promoters contain a consensus E-box motif that mediates the regulation of the PPARγ gene by ADD-1/SREBP-1 and SREBP-2. These results suggest that PPARγ expression can be controlled by the SREBP family of transcription factors and demonstrate new interactions between transcription factors that can regulate different pathways of lipid metabolism. PMID:10409739

  19. The Transcription Factor Interferon Regulatory Factor-1 (IRF1) Plays a Key Role in the Terminal Effector Pathways of Human Preterm Labor.

    PubMed

    Lim, Ratana; Tran, Ha Thi; Liong, Stella; Barker, Gillian; Lappas, Martha

    2016-02-01

    Preterm birth is the largest single cause of neonatal death and morbidity. By activating cytokine- and Toll-like receptor (TLR)-signaling pathways, infection and/or inflammation are strongly associated with preterm delivery. Interferon regulatory factor-1 (IRF1) is an important regulator of the inflammatory response. The aims of this study were to establish the effect of 1) labor on IRF1 expression in human fetal membranes and myometrium, 2) prolabor mediators on IRF1 expression and activity, and 3) IRF1 small interfering RNA on the expression of prolabor mediators. IRF1 expression was higher in fetal membranes and myometrium after spontaneous term labor and in preterm fetal membranes with infection. The proinflammatory cytokine IL1B, the bacterial product fsl-1, and viral analog polyinosinic:polycytidylic acid (poly [I:C]) significantly increased IRF1 mRNA expression and transcriptional activity in human primary myometrial cells. In addition, IL1B increased IRF1 activity in primary amnion cells. IRF1 silencing in myometrial cells decreased IL1B-, fsl-1-, and poly (I:C)-induced cytokine (IL6, TNF, IL1B) and chemokine (CXCL8, CCL2) mRNA expression and IL6, CXCL8, and CCL2 release. IL1B-, fsl-1-, and poly (I:C)-induced PTGS2 mRNA expression and IL1B-induced prostaglandin release was also decreased by IRF1 silencing. In conclusion, IRF1 upregulation in fetal membranes and myometrium after term labor indicates a proinflammatory role for IRF1 in human parturition. IRF1 is involved in TLR- and cytokine-mediated signaling in human myometrium. These data provide new insights into the mechanisms associated with inflammation- and infection-associated preterm birth. IRF1 inhibitors as therapeutics for the management of spontaneous preterm birth warrants further investigation. PMID:26674566

  20. Interferon Regulatory Factor 6 (IRF6) and Fibroblast Growth Factor Receptor 1 (FGFR1) Contribute to Human Tooth Agenesis

    PubMed Central

    Vieira, Alexandre R.; Modesto, Adriana; Meira, Raquel; Barbosa, Anna Renata Schneider; Lidral, Andrew C.; Murray, Jeffrey C.

    2008-01-01

    Phenotypic characteristics expressed in syndromes give clues to the factors involved in the cause of isolated forms of the same defects. We investigated two genes responsible for craniofacial syndromes, FGFR1 and IRF6, in a collection of families with isolated tooth agenesis. Cheek swab samples were obtained for DNA analysis from 116 case/parent trios. Probands had at least one developmentally missing tooth, excluding third molars. In addition, we studied 89 cases and 50 controls from Ohio to replicate any positive findings. Genotyping was performed by kinetic polymerase chain-reaction or TaqMan assays. Linkage disequilibrium analysis and transmission distortion of the marker alleles were performed. The same variants in the IRF6 gene that are associated with isolated orofacial clefts are also associated with human tooth agenesis (rs861019, P = 0.058; rs17015215—V274I, P = 0.0006; rs7802, P = 0.004). Mutations in IRF6 cause Van der Woude and popliteal pterygium syndromes. The craniofacial phenotypic characteristics of these syndromes include oral clefts and preferential tooth agenesis of incisors and premolars, besides pits on the lower lips. Also it appears that preferential premolar agenesis is associated with FGFR1 (P = 0.014) and IRF6 (P = 0.002) markers. There were statistically significant data suggesting that IRF6 interacts not only with MSX1 (P = 0.001), but also with TGFA (P = 0.03). PMID:17318851

  1. A Hox Transcription Factor Collective Binds a Highly Conserved Distal-less cis-Regulatory Module to Generate Robust Transcriptional Outcomes

    PubMed Central

    Uhl, Juli D.; Zandvakili, Arya; Gebelein, Brian

    2016-01-01

    cis-regulatory modules (CRMs) generate precise expression patterns by integrating numerous transcription factors (TFs). Surprisingly, CRMs that control essential gene patterns can differ greatly in conservation, suggesting distinct constraints on TF binding sites. Here, we show that a highly conserved Distal-less regulatory element (DCRE) that controls gene expression in leg precursor cells recruits multiple Hox, Extradenticle (Exd) and Homothorax (Hth) complexes to mediate dual outputs: thoracic activation and abdominal repression. Using reporter assays, we found that abdominal repression is particularly robust, as neither individual binding site mutations nor a DNA binding deficient Hth protein abolished cooperative DNA binding and in vivo repression. Moreover, a re-engineered DCRE containing a distinct configuration of Hox, Exd, and Hth sites also mediated abdominal Hox repression. However, the re-engineered DCRE failed to perform additional segment-specific functions such as thoracic activation. These findings are consistent with two emerging concepts in gene regulation: First, the abdominal Hox/Exd/Hth factors utilize protein-protein and protein-DNA interactions to form repression complexes on flexible combinations of sites, consistent with the TF collective model of CRM organization. Second, the conserved DCRE mediates multiple cell-type specific outputs, consistent with recent findings that pleiotropic CRMs are associated with conserved TF binding and added evolutionary constraints. PMID:27058369

  2. Regulatory review time for approval of oncology drugs in Japan between 2001 and 2014. Considerations of changes, factors that affect review time, and difference with the United States.

    PubMed

    Maeda, Hideki; Kurokawa, Tatsuo

    2015-05-01

    In this study, we comprehensively and historically studied the review time of oncology drugs approved by the regulatory authorities in Japan with publicly available information. A total of 120 applications of oncology drugs were approved in Japan between April 2001 and July 2014. The review time peaked with 732.0 days (24.4 months) in 2005, and showed a tendency to decline gradually each year thereafter. After 2012, a significant reduction of the review time was seen in comparison to the median of 13-year median time. In addition, we compared the review time with that in the United States. The median review time lag with the United States was significantly peaked in 2005. After 2005, the review time lag with the FDA has decreased, but lag did not significantly reduce by 2014. We also examined factors influencing the review time in Japan with multiple regression analysis. It was found that the factors related to a use of overseas data and expedited program for accelerating the reviews influenced the direction of shortening the review time. We consider that regulatory authorities in Japan need to keep making efforts to reduce the review time further and eliminate the review time lag with the United States.

  3. A Hox Transcription Factor Collective Binds a Highly Conserved Distal-less cis-Regulatory Module to Generate Robust Transcriptional Outcomes.

    PubMed

    Uhl, Juli D; Zandvakili, Arya; Gebelein, Brian

    2016-04-01

    cis-regulatory modules (CRMs) generate precise expression patterns by integrating numerous transcription factors (TFs). Surprisingly, CRMs that control essential gene patterns can differ greatly in conservation, suggesting distinct constraints on TF binding sites. Here, we show that a highly conserved Distal-less regulatory element (DCRE) that controls gene expression in leg precursor cells recruits multiple Hox, Extradenticle (Exd) and Homothorax (Hth) complexes to mediate dual outputs: thoracic activation and abdominal repression. Using reporter assays, we found that abdominal repression is particularly robust, as neither individual binding site mutations nor a DNA binding deficient Hth protein abolished cooperative DNA binding and in vivo repression. Moreover, a re-engineered DCRE containing a distinct configuration of Hox, Exd, and Hth sites also mediated abdominal Hox repression. However, the re-engineered DCRE failed to perform additional segment-specific functions such as thoracic activation. These findings are consistent with two emerging concepts in gene regulation: First, the abdominal Hox/Exd/Hth factors utilize protein-protein and protein-DNA interactions to form repression complexes on flexible combinations of sites, consistent with the TF collective model of CRM organization. Second, the conserved DCRE mediates multiple cell-type specific outputs, consistent with recent findings that pleiotropic CRMs are associated with conserved TF binding and added evolutionary constraints. PMID:27058369

  4. A Hox Transcription Factor Collective Binds a Highly Conserved Distal-less cis-Regulatory Module to Generate Robust Transcriptional Outcomes.

    PubMed

    Uhl, Juli D; Zandvakili, Arya; Gebelein, Brian

    2016-04-01

    cis-regulatory modules (CRMs) generate precise expression patterns by integrating numerous transcription factors (TFs). Surprisingly, CRMs that control essential gene patterns can differ greatly in conservation, suggesting distinct constraints on TF binding sites. Here, we show that a highly conserved Distal-less regulatory element (DCRE) that controls gene expression in leg precursor cells recruits multiple Hox, Extradenticle (Exd) and Homothorax (Hth) complexes to mediate dual outputs: thoracic activation and abdominal repression. Using reporter assays, we found that abdominal repression is particularly robust, as neither individual binding site mutations nor a DNA binding deficient Hth protein abolished cooperative DNA binding and in vivo repression. Moreover, a re-engineered DCRE containing a distinct configuration of Hox, Exd, and Hth sites also mediated abdominal Hox repression. However, the re-engineered DCRE failed to perform additional segment-specific functions such as thoracic activation. These findings are consistent with two emerging concepts in gene regulation: First, the abdominal Hox/Exd/Hth factors utilize protein-protein and protein-DNA interactions to form repression complexes on flexible combinations of sites, consistent with the TF collective model of CRM organization. Second, the conserved DCRE mediates multiple cell-type specific outputs, consistent with recent findings that pleiotropic CRMs are associated with conserved TF binding and added evolutionary constraints.

  5. Iron- and Quorum-sensing Signals Converge on Small Quorum-regulatory RNAs for Coordinated Regulation of Virulence Factors in Vibrio vulnificus.

    PubMed

    Wen, Yancheng; Kim, In Hwang; Kim, Kun-Soo

    2016-07-01

    Vibrio vulnificus is a marine bacterium that causes human infections resulting in high mortality. This pathogen harbors five quorum-regulatory RNAs (Qrr1-5) that affect the expression of pathogenicity genes by modulating the expression of the master regulator SmcR. The qrr genes are activated by phosphorylated LuxO to different degrees; qrr2 is strongly activated; qrr3 and qrr5 are moderately activated, and qrr1 and qrr4 are marginally activated and are the only two that do not respond to cell density-dependent regulation. Qrrs function redundantly to inhibit SmcR at low cell density and fully repress when all five are activated. In this study, we found that iron inhibits qrr expression in three distinct ways. First, the iron-ferric uptake regulator (Fur) complex directly binds to qrr promoter regions, inhibiting LuxO activation by competing with LuxO for cis-acting DNA elements. Second, qrr transcription is repressed by iron independently of Fur. Third, LuxO expression is repressed by iron independently of Fur. We also found that, under iron-limiting conditions, the five Qrrs functioned additively, not redundantly, to repress SmcR, suggesting that cells lacking iron enter a high cell density mode earlier and could thereby modulate expression of virulence factors sooner. This study suggests that iron and quorum sensing, along with their cognate regulatory circuits, are linked together in the coordinated expression of virulence factors.

  6. Regulatory T-cell suppressor program co-opts transcription factor IRF4 to control TH2 responses

    PubMed Central

    Zheng, Ye; Chaudhry, Ashutosh; Kas, Arnold; deRoos, Paul; Kim, Jeong M.; Chu, Tin-Tin; Corcoran, Lynn; Treuting, Piper; Klein, Ulf; Rudensky, Alexander Y.

    2010-01-01

    In the course of infection or autoimmunity, particular transcription factors orchestrate the differentiation of TH1, TH2 or TH17 effector cells, the responses of which are limited by a distinct lineage of suppressive regulatory T cells (Treg). Treg cell differentiation and function are guided by the transcription factor Foxp3, and their deficiency due to mutations in Foxp3 results in aggressive fatal autoimmune disease associated with sharply augmented TH1 and TH2 cytokine production1–3. Recent studies suggested that Foxp3 regulates the bulk of the Foxp3-dependent transcriptional program indirectly through a set of transcriptional regulators serving as direct Foxp3 targets4,5. Here we show that in mouse Treg cells, high amounts of interferon regulatory factor-4 (IRF4), a transcription factor essential for TH2 effector cell differentiation, is dependent on Foxp3 expression. We proposed that IRF4 expression endows Treg cells with the ability to suppress TH2 responses. Indeed, ablation of a conditional Irf4 allele in Treg cells resulted in selective dysregulation of TH2 responses, IL4-dependent immunoglobulin isotype production, and tissue lesions with pronounced plasma cell infiltration, in contrast to the mononuclear-cell-dominated pathology typical of mice lacking Treg cells. Our results indicate that Treg cells use components of the transcriptional machinery, promoting a particular type of effector CD4+ T cell differentiation, to efficiently restrain the corresponding type of the immune response. PMID:19182775

  7. Characterization of the microRNA pool and the factors affecting its regulatory potential.

    PubMed

    Cui, Kai; Lyu, Qing; Xu, Naihan; Liu, Qing; Zhang, Jiarong; Xing, Wei; Bai, Linfu; Liao, Meijian; He, Jie; Yuan, Bo; Chen, Deheng; Xie, Weidong; Zhang, Yaou

    2014-12-01

    The regulation of gene expression by microRNAs (miRNAs) is complex due to a number of variables involved. The potential for one miRNA to target many genes, the presence of multiple miRNA response elements (MREs) in one mRNA molecule and the interplay between RNAs that share common MREs each add a layer of complexity to the process; making it difficult to determine how regulation of gene expression by miRNAs works within the context of the system as a whole. In this study, we used luciferase report vectors inserted with different 3'UTR fragments as probes to detect the repressive effect of the miRNA pool on gene expression and uncovered some essential characteristics of gene regulation mediated by the miRNA pool, such as the nonlinear correlative relationship between the regulatory potential of a miRNA pool and the number of potential MREs, the buffering effect and the saturating effect of the miRNA pool, and the restrictive effect caused by the density of MREs. Through expressing gradient concentration of 3'UTR fragments, we indirectly detected the regulatory potential of the competing endogenous RNA (ceRNA) pool and analysed its effect on the regulatory potential of the miRNA pool. Our results provide some new insights into miRNA pool mediated gene regulation.

  8. Additive Factors Do Not Imply Discrete Processing Stages: A Worked Example Using Models of the Stroop Task

    PubMed Central

    Stafford, Tom; Gurney, Kevin N.

    2011-01-01

    Previously, it has been shown experimentally that the psychophysical law known as Piéron’s Law holds for color intensity and that the size of the effect is additive with that of Stroop condition (Stafford et al., 2011). According to the additive factors method (Donders, 1868–1869/1969; Sternberg, 1998), additivity is assumed to indicate independent and discrete processing stages. We present computational modeling work, using an existing Parallel Distributed Processing model of the Stroop task (Cohen et al., 1990) and a standard model of decision making (Ratcliff, 1978). This demonstrates that additive factors can be successfully accounted for by existing single stage models of the Stroop effect. Consequently, it is not valid to infer either discrete stages or separate loci of effects from additive factors. Further, our modeling work suggests that information binding may be a more important architectural property for producing additive factors than discrete stages. PMID:22102842

  9. Additive cytotoxicity of different monoclonal antibody-cobra venom factor conjugates for human neuroblastoma cells.

    PubMed

    Juhl, H; Petrella, E C; Cheung, N K; Bredehorst, R; Vogel, C W

    1997-11-01

    Insufficient numbers of antigen molecules and heterogeneity of antigen expression on tumor cells are major factors limiting the immunotherapeutic potential of the few clinically useful monoclonal antibodies capable of mediating complement cytotoxicity and antibody-dependent cellular cytotoxicity. To overcome this limitation, we converted two non-cytotoxic monoclonal anti-neuroblastoma antibodies, designated 3E7 (IgG2b) and 8H9 (IgG1), and the non-cytotoxic F(ab')2 fragment of the cytotoxic monoclonal anti-GD2 antibody 3F8 (IgG3) into cytotoxic antibody conjugates by covalent attachment of cobra venom factor (CVF), a structural and functional homologue of the activated third component of complement. Competitive binding experiments confirmed the different specificities of the three antibodies. In the presence of human complement, all three antibody-CVF conjugates mediated selective complement-dependent lysis of human neuroblastoma cells. Consistent with the kinetics of the alternative pathway of complement, approximately seven hours incubation were required to reach maximum cytotoxicity of up to 25% for the 3E7-CVF conjugate, up to 60% for the 8H9-CVF conjugate, and up to 95% for the 3F8 F(ab')2-CVF conjugate. The different extent of maximal cytotoxic activity of the three conjugates was reflected by corresponding differences in the extent of binding of both unconjugated antibodies and the respective conjugates. Any combination of the three antibody-CVF conjugates caused an additive effect in complement-mediated lysis. Using a cocktail of all three conjugates, the extent of complement-mediated killing could be increased up to 100%. These data demonstrate that by coupling of CVF the relative large number of non-cytotoxic monoclonal anti-tumor antibodies of interesting specificity can be used to design cocktails of cytotoxic conjugates and, thereby, to overcome the problem of insufficient and heterogeneous antigen expression on tumor cells for immunotherapy.

  10. Integrating products of Bessel functions with an additional exponential or rational factor

    NASA Astrophysics Data System (ADS)

    Van Deun, Joris; Cools, Ronald

    2008-04-01

    We provide two MATLAB programs to compute integrals of the form ex∏i=1kJν_i(ax)dxand 0∞xr+x∏i=1kJν_i(ax)dx with Jν_i(x) the Bessel function of the first kind and (real) order ν. The parameter m is a real number such that ∑ν+m>-1 (to assure integrability near zero), r is real and the numbers c and a are all strictly positive. The program can deliver accurate error estimates. Program summaryProgram title: BESSELINTR, BESSELINTC Catalogue identifier: AEAH_v1_0 Program summary URL:http://cpc.cs.qub.ac.uk/summaries/AEAH_v1_0.html Program obtainable from: CPC Program Library, Queen's University, Belfast, N. Ireland Licensing provisions: Standard CPC licence, http://cpc.cs.qub.ac.uk/licence/licence.html No. of lines in distributed program, including test data, etc.: 1601 No. of bytes in distributed program, including test data, etc.: 13 161 Distribution format: tar.gz Programming language: Matlab (version ⩾6.5), Octave (version ⩾2.1.69) Computer: All supporting Matlab or Octave Operating system: All supporting Matlab or Octave RAM: For k Bessel functions our program needs approximately ( 500+140k) double precision variables Classification: 4.11 Nature of problem: The problem consists in integrating an arbitrary product of Bessel functions with an additional rational or exponential factor over a semi-infinite interval. Difficulties arise from the irregular oscillatory behaviour and the possible slow decay of the integrand, which prevents truncation at a finite point. Solution method: The interval of integration is split into a finite and infinite part. The integral over the finite part is computed using Gauss-Legendre quadrature. The integrand on the infinite part is approximated using asymptotic expansions and this approximation is integrated exactly with the aid of the upper incomplete gamma function. In the case where a rational factor is present, this factor is first expanded in a Taylor series around infinity. Restrictions: Some (and eventually all

  11. KLF2 is a rate-limiting transcription factor that can be targeted to enhance regulatory T-cell production

    PubMed Central

    Pabbisetty, Sudheer K.; Rabacal, Whitney; Maseda, Damian; Cendron, Delphine; Collins, Patrick L.; Hoek, Kristen L.; Parekh, Vrajesh V.; Aune, Thomas M.; Sebzda, Eric

    2014-01-01

    Regulatory T cells (Tregs) are a specialized subset of CD4+ T cells that maintain self-tolerance by functionally suppressing autoreactive lymphocytes. The Treg compartment is composed of thymus-derived Tregs (tTregs) and peripheral Tregs (pTregs) that are generated in secondary lymphoid organs after exposure to antigen and specific cytokines, such as TGF-β. With regard to this latter lineage, pTregs [and their ex vivo generated counterparts, induced Tregs (iTregs)] offer particular therapeutic potential because these cells can be raised against specific antigens to limit autoimmunity. We now report that transcription factor Krüppel-like factor 2 (KLF2) is necessary for the generation of iTregs but not tTregs. Moreover, drugs that limit KLF2 proteolysis during T-cell activation enhance iTreg development. To the authors’ knowledge, this study identifies the first transcription factor to distinguish between i/pTreg and tTreg ontogeny and demonstrates that KLF2 is a therapeutic target for the production of regulatory T cells. PMID:24979767

  12. An integrated approach to characterize transcription factor and microRNA regulatory networks involved in Schwann cell response to peripheral nerve injury

    PubMed Central

    2013-01-01

    Background The regenerative response of Schwann cells after peripheral nerve injury is a critical process directly related to the pathophysiology of a number of neurodegenerative diseases. This SC injury response is dependent on an intricate gene regulatory program coordinated by a number of transcription factors and microRNAs, but the interactions among them remain largely unknown. Uncovering the transcriptional and post-transcriptional regulatory networks governing the Schwann cell injury response is a key step towards a better understanding of Schwann cell biology and may help develop novel therapies for related diseases. Performing such comprehensive network analysis requires systematic bioinformatics methods to integrate multiple genomic datasets. Results In this study we present a computational pipeline to infer transcription factor and microRNA regulatory networks. Our approach combined mRNA and microRNA expression profiling data, ChIP-Seq data of transcription factors, and computational transcription factor and microRNA target prediction. Using mRNA and microRNA expression data collected in a Schwann cell injury model, we constructed a regulatory network and studied regulatory pathways involved in Schwann cell response to injury. Furthermore, we analyzed network motifs and obtained insights on cooperative regulation of transcription factors and microRNAs in Schwann cell injury recovery. Conclusions This work demonstrates a systematic method for gene regulatory network inference that may be used to gain new information on gene regulation by transcription factors and microRNAs. PMID:23387820

  13. Translation regulatory factor RBM3 is a proto-oncogene that prevents mitotic catastrophe

    PubMed Central

    Sureban, SM; Ramalingam, S; Natarajan, G; May, R; Subramaniam, D; Bishnupuri, KS; Morrison, AR; Dieckgraefe, BK; Brackett, DJ; Postier, RG; Houchen, CW; Anant, S

    2009-01-01

    RNA-binding proteins play a key role in post-transcriptional regulation of mRNA stability and translation. We have identified that RBM3, a translation regulatory protein, is significantly upregulated in human tumors, including a stage-dependent increase in colorectal tumors. Forced RBM3 overexpression in NIH3T3 mouse fibro-blasts and SW480 human colon epithelial cells increases cell proliferation and development of compact multicellular spheroids in soft agar suggesting the ability to induce anchorage-independent growth. In contrast, down-regulating RBM3 in HCT116 colon cancer cells with specific siRNA decreases cell growth in culture, which was partially overcome when treated with prostaglandin E2, a product of cyclooxygenase (COX)-2 enzyme activity. Knockdown also resulted in the growth arrest of tumor xenografts. We have also identified that RBM3 knockdown increases caspase-mediated apoptosis coupled with nuclear cyclin B1, and phosphorylated Cdc25c, Chk1 and Chk2 kinases, implying that under conditions of RBM3 downregulation, cells undergo mitotic catastrophe. RBM3 enhances COX-2, IL-8 and VEGF mRNA stability and translation. Conversely, RBM3 knockdown results in loss in the translation of these transcripts. These data demonstrate that the RNA stabilizing and translation regulatory protein RBM3 is a novel proto-oncogene that induces transformation when overexpressed and is essential for cells to progress through mitosis. PMID:18427544

  14. Altered expression of hypoxia-inducible factor-1α (HIF-1α) and its regulatory genes in gastric cancer tissues.

    PubMed

    Wang, Jihan; Ni, Zhaohui; Duan, Zipeng; Wang, Guoqing; Li, Fan

    2014-01-01

    Tissue hypoxia induces reprogramming of cell metabolism and may result in normal cell transformation and cancer progression. Hypoxia-inducible factor 1-alpha (HIF-1α), the key transcription factor, plays an important role in gastric cancer development and progression. This study aimed to investigate the underlying regulatory signaling pathway in gastric cancer using gastric cancer tissue specimens. The integration of gene expression profile and transcriptional regulatory element database (TRED) was pursued to identify HIF-1α ↔ NFκB1 → BRCA1 → STAT3 ← STAT1 gene pathways and their regulated genes. The data showed that there were 82 differentially expressed genes that could be regulated by these five transcription factors in gastric cancer tissues and these genes formed 95 regulation modes, among which seven genes (MMP1, TIMP1, TLR2, FCGR3A, IRF1, FAS, and TFF3) were hub molecules that are regulated at least by two of these five transcription factors simultaneously and were associated with hypoxia, inflammation, and immune disorder. Real-Time PCR and western blot showed increasing of HIF-1α in mRNA and protein levels as well as TIMP1, TFF3 in mRNA levels in gastric cancer tissues. The data are the first study to demonstrate HIF-1α-regulated transcription factors and their corresponding network genes in gastric cancer. Further study with a larger sample size and more functional experiments is needed to confirm these data and then translate into clinical biomarker discovery and treatment strategy for gastric cancer.

  15. Altered Expression of Hypoxia-Inducible Factor-1α (HIF-1α) and Its Regulatory Genes in Gastric Cancer Tissues

    PubMed Central

    Wang, Jihan; Ni, Zhaohui; Duan, Zipeng; Wang, Guoqing; Li, Fan

    2014-01-01

    Tissue hypoxia induces reprogramming of cell metabolism and may result in normal cell transformation and cancer progression. Hypoxia-inducible factor 1-alpha (HIF-1α), the key transcription factor, plays an important role in gastric cancer development and progression. This study aimed to investigate the underlying regulatory signaling pathway in gastric cancer using gastric cancer tissue specimens. The integration of gene expression profile and transcriptional regulatory element database (TRED) was pursued to identify HIF-1α ↔ NFκB1 → BRCA1 → STAT3 ← STAT1 gene pathways and their regulated genes. The data showed that there were 82 differentially expressed genes that could be regulated by these five transcription factors in gastric cancer tissues and these genes formed 95 regulation modes, among which seven genes (MMP1, TIMP1, TLR2, FCGR3A, IRF1, FAS, and TFF3) were hub molecules that are regulated at least by two of these five transcription factors simultaneously and were associated with hypoxia, inflammation, and immune disorder. Real-Time PCR and western blot showed increasing of HIF-1α in mRNA and protein levels as well as TIMP1, TFF3 in mRNA levels in gastric cancer tissues. The data are the first study to demonstrate HIF-1α-regulated transcription factors and their corresponding network genes in gastric cancer. Further study with a larger sample size and more functional experiments is needed to confirm these data and then translate into clinical biomarker discovery and treatment strategy for gastric cancer. PMID:24927122

  16. Enhancer sequence variants and transcription-factor deregulation synergize to construct pathogenic regulatory circuits in B-cell lymphoma.

    PubMed

    Koues, Olivia I; Kowalewski, Rodney A; Chang, Li-Wei; Pyfrom, Sarah C; Schmidt, Jennifer A; Luo, Hong; Sandoval, Luis E; Hughes, Tyler B; Bednarski, Jeffrey J; Cashen, Amanda F; Payton, Jacqueline E; Oltz, Eugene M

    2015-01-20

    Most B-cell lymphomas arise in the germinal center (GC), where humoral immune responses evolve from potentially oncogenic cycles of mutation, proliferation, and clonal selection. Although lymphoma gene expression diverges significantly from GC B cells, underlying mechanisms that alter the activities of corresponding regulatory elements (REs) remain elusive. Here we define the complete pathogenic circuitry of human follicular lymphoma (FL), which activates or decommissions REs from normal GC B cells and commandeers enhancers from other lineages. Moreover, independent sets of transcription factors, whose expression was deregulated in FL, targeted commandeered versus decommissioned REs. Our approach revealed two distinct subtypes of low-grade FL, whose pathogenic circuitries resembled GC B or activated B cells. FL-altered enhancers also were enriched for sequence variants, including somatic mutations, which disrupt transcription-factor binding and expression of circuit-linked genes. Thus, the pathogenic regulatory circuitry of FL reveals distinct genetic and epigenetic etiologies for GC B-cell transformation.

  17. 34 CFR 359.32 - What additional factors does the Secretary consider in making a grant under this program?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... EDUCATION DISABILITY AND REHABILITATION RESEARCH: SPECIAL PROJECTS AND DEMONSTRATIONS FOR SPINAL CORD INJURIES How Does the Secretary Make a Grant? § 359.32 What additional factors does the Secretary...

  18. 34 CFR 359.32 - What additional factors does the Secretary consider in making a grant under this program?

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... EDUCATION DISABILITY AND REHABILITATION RESEARCH: SPECIAL PROJECTS AND DEMONSTRATIONS FOR SPINAL CORD INJURIES How Does the Secretary Make a Grant? § 359.32 What additional factors does the Secretary...

  19. 34 CFR 359.32 - What additional factors does the Secretary consider in making a grant under this program?

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... EDUCATION DISABILITY AND REHABILITATION RESEARCH: SPECIAL PROJECTS AND DEMONSTRATIONS FOR SPINAL CORD INJURIES How Does the Secretary Make a Grant? § 359.32 What additional factors does the Secretary...

  20. 34 CFR 359.32 - What additional factors does the Secretary consider in making a grant under this program?

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... EDUCATION DISABILITY AND REHABILITATION RESEARCH: SPECIAL PROJECTS AND DEMONSTRATIONS FOR SPINAL CORD INJURIES How Does the Secretary Make a Grant? § 359.32 What additional factors does the Secretary...

  1. 34 CFR 359.32 - What additional factors does the Secretary consider in making a grant under this program?

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... EDUCATION DISABILITY AND REHABILITATION RESEARCH: SPECIAL PROJECTS AND DEMONSTRATIONS FOR SPINAL CORD INJURIES How Does the Secretary Make a Grant? § 359.32 What additional factors does the Secretary...

  2. Two new monoclonal antibodies for biochemical and flow cytometric analyses of human interferon regulatory factor-3 activation, turnover, and depletion.

    PubMed

    Rustagi, Arjun; Doehle, Brian P; McElrath, M Juliana; Gale, Michael

    2013-02-01

    Interferon regulatory factor-3 (IRF-3) is a master transcription factor that drives the host intracellular innate immune response to virus infection. The importance of IRF-3 in innate immune responses is highlighted by the fact that pathogenic viruses have developed strategies for antagonism of IRF-3. Several tools exist for evaluation of viral regulation of IRF-3 activation and function, but high-quality monoclonal antibodies that mark the differential activation states of human IRF-3 are lacking. To study IRF-3 activation, turnover, and depletion in a high-throughput manner in the context of virus infection, we have developed two new monoclonal antibodies to human IRF-3. These antibodies detect IRF-3 in virus-infected cells in a wide variety of assays and provide a new tool to study virus-host interactions and innate immune signaling.

  3. Two new monoclonal antibodies for biochemical and flow cytometric analyses of human interferon regulatory factor-3 activation, turnover, and depletion

    PubMed Central

    Rustagi, Arjun; Doehle, Brian P.; McElrath, M. Juliana; Gale, Michael

    2012-01-01

    Interferon regulatory factor-3 (IRF-3) is a master transcription factor that drives the host intracellular innate immune response to virus infection. The importance of IRF-3 in innate immune responses is highlighted by the fact that pathogenic viruses have developed strategies for antagonism of IRF-3. Several tools exist for evaluation of viral regulation of IRF-3 activation and function, but high-quality monoclonal antibodies that mark the differential activation states of human IRF-3 are lacking. To study IRF-3 activation, turnover, and depletion in a high-throughput manner in the context of virus infection, we have developed two new monoclonal antibodies to human IRF-3. These antibodies detect IRF-3 in virus-infected cells in a wide variety of assays and provide a new tool to study virus-host interactions and innate immune signaling. PMID:22705311

  4. Inhibitor of apoptosis proteins (IAPs) as regulatory factors of hepatic apoptosis.

    PubMed

    Wang, Kewei; Lin, Bingliang

    2013-10-01

    IAPs are a group of regulatory proteins that are structurally related. Their conserved homologues have been identified in various organisms. In human, eight IAP members have been recognized based on baculoviral IAP repeat (BIR) domains. IAPs are key regulators of apoptosis, cytokinesis and signal transduction. The antiapoptotic property of IAPs depends on their professional role for caspases. IAPs are functionally non-equivalent and regulate effector caspases through distinct mechanisms. IAPs impede apoptotic process via membrane receptor-dependent (extrinsic) cascade and mitochondrial dependent (intrinsic) pathway. IAP-mediated apoptosis affects the progression of liver diseases. Therapeutic options of liver diseases may depend on the understanding toward mechanisms of the IAP-mediated apoptosis.

  5. Relative Importance and Additive Effects of Maternal and Infant Risk Factors on Childhood Asthma

    PubMed Central

    Rosas-Salazar, Christian; James, Kristina; Escobar, Gabriel; Gebretsadik, Tebeb; Li, Sherian Xu; Carroll, Kecia N.; Walsh, Eileen; Mitchel, Edward; Das, Suman; Kumar, Rajesh; Yu, Chang; Dupont, William D.; Hartert, Tina V.

    2016-01-01

    Background Environmental exposures that occur in utero and during early life may contribute to the development of childhood asthma through alteration of the human microbiome. The objectives of this study were to estimate the cumulative effect and relative importance of environmental exposures on the risk of childhood asthma. Methods We conducted a population-based birth cohort study of mother-child dyads who were born between 1995 and 2003 and were continuously enrolled in the PRIMA (Prevention of RSV: Impact on Morbidity and Asthma) cohort. The individual and cumulative impact of maternal urinary tract infections (UTI) during pregnancy, maternal colonization with group B streptococcus (GBS), mode of delivery, infant antibiotic use, and older siblings at home, on the risk of childhood asthma were estimated using logistic regression. Dose-response effect on childhood asthma risk was assessed for continuous risk factors: number of maternal UTIs during pregnancy, courses of infant antibiotics, and number of older siblings at home. We further assessed and compared the relative importance of these exposures on the asthma risk. In a subgroup of children for whom maternal antibiotic use during pregnancy information was available, the effect of maternal antibiotic use on the risk of childhood asthma was estimated. Results Among 136,098 singleton birth infants, 13.29% developed asthma. In both univariate and adjusted analyses, maternal UTI during pregnancy (odds ratio [OR] 1.2, 95% confidence interval [CI] 1.18, 1.25; adjusted OR [AOR] 1.04, 95%CI 1.02, 1.07 for every additional UTI) and infant antibiotic use (OR 1.21, 95%CI 1.20, 1.22; AOR 1.16, 95%CI 1.15, 1.17 for every additional course) were associated with an increased risk of childhood asthma, while having older siblings at home (OR 0.92, 95%CI 0.91, 0.93; AOR 0.85, 95%CI 0.84, 0.87 for each additional sibling) was associated with a decreased risk of childhood asthma, in a dose-dependent manner. Compared with vaginal

  6. Genomic analysis of host - Peste des petits ruminants vaccine viral transcriptome uncovers transcription factors modulating immune regulatory pathways.

    PubMed

    Manjunath, Siddappa; Kumar, Gandham Ravi; Mishra, Bishnu Prasad; Mishra, Bina; Sahoo, Aditya Prasad; Joshi, Chaitanya G; Tiwari, Ashok K; Rajak, Kaushal Kishore; Janga, Sarath Chandra

    2015-01-01

    Peste des petits ruminants (PPR), is an acute transboundary viral disease of economic importance, affecting goats and sheep. Mass vaccination programs around the world resulted in the decline of PPR outbreaks. Sungri 96 is a live attenuated vaccine, widely used in Northern India against PPR. This vaccine virus, isolated from goat works efficiently both in sheep and goat. Global gene expression changes under PPR vaccine virus infection are not yet well defined. Therefore, in this study we investigated the host-vaccine virus interactions by infecting the peripheral blood mononuclear cells isolated from goat with PPRV (Sungri 96 vaccine virus), to quantify the global changes in the transcriptomic signature by RNA-sequencing. Viral genome of Sungri 96 vaccine virus was assembled from the PPRV infected transcriptome confirming the infection and demonstrating the feasibility of building a complete non-host genome from the blood transcriptome. Comparison of infected transcriptome with control transcriptome revealed 985 differentially expressed genes. Functional analysis showed enrichment of immune regulatory pathways under PPRV infection. Key genes involved in immune system regulation, spliceosomal and apoptotic pathways were identified to be dysregulated. Network analysis revealed that the protein - protein interaction network among differentially expressed genes is significantly disrupted in infected state. Several genes encoding TFs that govern immune regulatory pathways were identified to co-regulate the differentially expressed genes. These data provide insights into the host - PPRV vaccine virus interactome for the first time. Our findings suggested dysregulation of immune regulatory pathways and genes encoding Transcription Factors (TFs) that govern these pathways in response to viral infection. PMID:25827022

  7. Dynamic control of gene regulatory logic by seemingly redundant transcription factors

    PubMed Central

    AkhavanAghdam, Zohreh; Sinha, Joydeb; Tabbaa, Omar P; Hao, Nan

    2016-01-01

    Many transcription factors co-express with their homologs to regulate identical target genes, however the advantages of such redundancies remain elusive. Using single-cell imaging and microfluidics, we study the yeast general stress response transcription factor Msn2 and its seemingly redundant homolog Msn4. We find that gene regulation by these two factors is analogous to logic gate systems. Target genes with fast activation kinetics can be fully induced by either factor, behaving as an 'OR' gate. In contrast, target genes with slow activation kinetics behave as an 'AND' gate, requiring distinct contributions from both factors, upon transient stimulation. Furthermore, such genes become an 'OR' gate when the input duration is prolonged, suggesting that the logic gate scheme is not static but rather dependent on the input dynamics. Therefore, Msn2 and Msn4 enable a time-based mode of combinatorial gene regulation that might be applicable to homologous transcription factors in other organisms. DOI: http://dx.doi.org/10.7554/eLife.18458.001 PMID:27690227

  8. Comparative analysis of transcription factor gene families from Papaver somniferum: identification of regulatory factors involved in benzylisoquinoline alkaloid biosynthesis.

    PubMed

    Agarwal, Parul; Pathak, Sumya; Lakhwani, Deepika; Gupta, Parul; Asif, Mehar Hasan; Trivedi, Prabodh Kumar

    2016-05-01

    Opium poppy (Papaver somniferum L.), known for biosynthesis of several therapeutically important benzylisoquinoline alkaloids (BIAs), has emerged as the premier organism to study plant alkaloid metabolism. The most prominent molecules produced in opium poppy include narcotic analgesic morphine, the cough suppressant codeine, the muscle relaxant papaverine and the anti-microbial agent sanguinarine and berberine. Despite several health benefits, biosynthesis of some of these molecules is very low due to tight temporal and spatial regulation of the genes committed to their biosynthesis. Transcription factors, one of the prime regulators of secondary plant product biosynthesis, might be involved in controlled biosynthesis of BIAs in P. somniferum. In this study, identification of members of different transcription factor gene families using transcriptome datasets of 10 cultivars of P. somniferum with distinct chemoprofile has been carried out. Analysis suggests that most represented transcription factor gene family in all the poppy cultivars is WRKY. Comparative transcriptome analysis revealed differential expression pattern of the members of a set of transcription factor gene families among 10 cultivars. Through analysis, two members of WRKY and one member of C3H gene family were identified as potential candidates which might regulate thebaine and papaverine biosynthesis, respectively, in poppy.

  9. Comparative analysis of transcription factor gene families from Papaver somniferum: identification of regulatory factors involved in benzylisoquinoline alkaloid biosynthesis.

    PubMed

    Agarwal, Parul; Pathak, Sumya; Lakhwani, Deepika; Gupta, Parul; Asif, Mehar Hasan; Trivedi, Prabodh Kumar

    2016-05-01

    Opium poppy (Papaver somniferum L.), known for biosynthesis of several therapeutically important benzylisoquinoline alkaloids (BIAs), has emerged as the premier organism to study plant alkaloid metabolism. The most prominent molecules produced in opium poppy include narcotic analgesic morphine, the cough suppressant codeine, the muscle relaxant papaverine and the anti-microbial agent sanguinarine and berberine. Despite several health benefits, biosynthesis of some of these molecules is very low due to tight temporal and spatial regulation of the genes committed to their biosynthesis. Transcription factors, one of the prime regulators of secondary plant product biosynthesis, might be involved in controlled biosynthesis of BIAs in P. somniferum. In this study, identification of members of different transcription factor gene families using transcriptome datasets of 10 cultivars of P. somniferum with distinct chemoprofile has been carried out. Analysis suggests that most represented transcription factor gene family in all the poppy cultivars is WRKY. Comparative transcriptome analysis revealed differential expression pattern of the members of a set of transcription factor gene families among 10 cultivars. Through analysis, two members of WRKY and one member of C3H gene family were identified as potential candidates which might regulate thebaine and papaverine biosynthesis, respectively, in poppy. PMID:26108744

  10. Cleavage of interferon regulatory factor 7 by enterovirus 71 3C suppresses cellular responses.

    PubMed

    Lei, Xiaobo; Xiao, Xia; Xue, Qinghua; Jin, Qi; He, Bin; Wang, Jianwei

    2013-02-01

    Enterovirus 71 (EV71) is a positive-stranded RNA virus which is capable of inhibiting innate immunity. Among virus-encoded proteins, the 3C protein compromises the type I interferon (IFN-I) response mediated by retinoid acid-inducible gene-I (RIG-I) or Toll-like receptor 3 that activates interferon regulatory 3 (IRF3) and IRF7. In the present study, we report that enterovirus 71 downregulates IRF7 through the 3C protein, which inhibits the function of IRF7. When expressed in mammalian cells, the 3C protein mediates cleavage of IRF7 rather than that of IRF3. This process is insensitive to inhibitors of caspase, proteasome, lysosome, and autophagy. H40D substitution in the 3C active site abolishes its activity, whereas R84Q or V154S substitution in the RNA binding motif has no effect. Furthermore, 3C-mediated cleavage occurs at the Q189-S190 junction within the constitutive activation domain of IRF7, resulting in two cleaved IRF7 fragments that are incapable of activating IFN expression. Ectopic expression of wild-type IRF7 limits EV71 replication. On the other hand, expression of the amino-terminal domain of IRF7 enhances EV71 infection, which correlates with its ability to interact with and inhibit IRF3. These results suggest that control of IRF7 by the 3C protein may represent a viral mechanism to escape cellular responses. PMID:23175366

  11. Genetic and environmental factors affecting cryptic variations in gene regulatory networks

    PubMed Central

    2013-01-01

    Background Cryptic genetic variation (CGV) is considered to facilitate phenotypic evolution by producing visible variations in response to changes in the internal and/or external environment. Several mechanisms enabling the accumulation and release of CGVs have been proposed. In this study, we focused on gene regulatory networks (GRNs) as an important mechanism for producing CGVs, and examined how interactions between GRNs and the environment influence the number of CGVs by using individual-based simulations. Results Populations of GRNs were allowed to evolve under various stabilizing selections, and we then measured the number of genetic and phenotypic variations that had arisen. Our results showed that CGVs were not depleted irrespective of the strength of the stabilizing selection for each phenotype, whereas the visible fraction of genetic variation in a population decreased with increasing strength of selection. On the other hand, increasing the number of different environments that individuals encountered within their lifetime (i.e., entailing plastic responses to multiple environments) suppressed the accumulation of CGVs, whereas the GRNs with more genes and interactions were favored in such heterogeneous environments. Conclusions Given the findings that the number of CGVs in a population was largely determined by the size (order) of GRNs, we propose that expansion of GRNs and adaptation to novel environments are mutually facilitating and sustainable sources of evolvability and hence the origins of biological diversity and complexity. PMID:23622056

  12. Insulin counter-regulatory factors, fibrinogen and C-reactive protein during olanzapine administration: effects of the antidiabetic metformin.

    PubMed

    Baptista, Trino; Sandia, Ignacio; Lacruz, Anny; Rangel, Nairy; de Mendoza, Soaira; Beaulieu, Serge; Contreras, Quilianio; Galeazzi, Tatiana; Vargas, Doritza

    2007-03-01

    In this study, the Authors assessed some insulin counter-regulatory factors, fibrinogen and C-reactive protein after olanzapine administration, and the effect of metformin on these variables, 37 patients with chronic schizophrenia were given olanzapine (10 mg/day for 14 weeks). Nineteen patients received metformin (850-2550 mg/day) and 18 received placebo in a randomized, double-blind protocol. The following variables were quantified before and after olanzapine: cortisol, leptin, tumor necrosis factor-alpha, glucagon, growth hormone, fibrinogen and C-reactive protein. Results were correlated with the changes in body weight and the insulin resistance index. We have reported elsewhere that metformin did not prevent olanzapine-induced weight gain, and the insulin resistance index significantly decreased after metformin and placebo; Baptista T, et al. Can J Psychiatry 2006; 51: 192-196. Cortisol, tumor necrosis factor-alpha and fibrinogen levels significantly decreased in both groups. Glucagon significantly increased after metformin (P=0.03). Leptin tended to increase after placebo (P=0.1) and displayed a small nonsignificant reduction after metformin. The C-reactive protein did not change significantly in any group. Contrarily to most published studies, olanzapine was associated with decreased insulin resistance. Decrements in cortisol, fibrinogen and tumor necrosis factor-alpha levels point to an improvement in the metabolic profile. The trend for leptin to increase after placebo, but not after metformin in spite of similar weight gain suggests a beneficial effect of this antidiabetic agent. PMID:17293706

  13. A direct regulatory interaction between chaperonin TRiC and stress-responsive transcription factor HSF1.

    PubMed

    Neef, Daniel W; Jaeger, Alex M; Gomez-Pastor, Rocio; Willmund, Felix; Frydman, Judith; Thiele, Dennis J

    2014-11-01

    Heat shock transcription factor 1 (HSF1) is an evolutionarily conserved transcription factor that protects cells from protein-misfolding-induced stress and apoptosis. The mechanisms by which cytosolic protein misfolding leads to HSF1 activation have not been elucidated. Here, we demonstrate that HSF1 is directly regulated by TRiC/CCT, a central ATP-dependent chaperonin complex that folds cytosolic proteins. A small-molecule activator of HSF1, HSF1A, protects cells from stress-induced apoptosis, binds TRiC subunits in vivo and in vitro, and inhibits TRiC activity without perturbation of ATP hydrolysis. Genetic inactivation or depletion of the TRiC complex results in human HSF1 activation, and HSF1A inhibits the direct interaction between purified TRiC and HSF1 in vitro. These results demonstrate a direct regulatory interaction between the cytosolic chaperone machine and a critical transcription factor that protects cells from proteotoxicity, providing a mechanistic basis for signaling perturbations in protein folding to a stress-protective transcription factor. PMID:25437552

  14. A trans-acting Variant within the Transcription Factor RIM101 Interacts with Genetic Background to Determine its Regulatory Capacity.

    PubMed

    Read, Timothy; Richmond, Phillip A; Dowell, Robin D

    2016-01-01

    Most genetic variants associated with disease occur within regulatory regions of the genome, underscoring the importance of defining the mechanisms underlying differences in regulation of gene expression between individuals. We discovered a pair of co-regulated, divergently oriented transcripts, AQY2 and ncFRE6, that are expressed in one strain of Saccharomyces cerevisiae, ∑1278b, but not in another, S288c. By combining classical genetics techniques with high-throughput sequencing, we identified a trans-acting single nucleotide polymorphism within the transcription factor RIM101 that causes the background-dependent expression of both transcripts. Subsequent RNA-seq experiments revealed that RIM101 regulates many more targets in S288c than in ∑1278b and that deletion of RIM101 in both backgrounds abrogates the majority of differential expression between the strains. Strikingly, only three transcripts undergo a significant change in expression after swapping RIM101 alleles between backgrounds, implying that the differences in the RIM101 allele lead to a remarkably focused transcriptional response. However, hundreds of RIM101-dependent targets undergo a subtle but consistent shift in expression in the S288c RIM101-swapped strain, but not its ∑1278b counterpart. We conclude that ∑1278b may harbor a variant(s) that buffers against widespread transcriptional dysregulation upon introduction of a non-native RIM101 allele, emphasizing the importance of accounting for genetic background when assessing the impact of a regulatory variant.

  15. 21 CFR 1311.115 - Additional requirements for two-factor authentication.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ..., such as a password or response to a challenge question. (2) Something the practitioner is, biometric... modules or one-time-password devices. (c) If one factor is a biometric, the biometric subsystem...

  16. 21 CFR 1311.115 - Additional requirements for two-factor authentication.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ..., such as a password or response to a challenge question. (2) Something the practitioner is, biometric... modules or one-time-password devices. (c) If one factor is a biometric, the biometric subsystem...

  17. Regulatory and functional connection of microphthalmia-associated transcription factor and anti-metastatic pigment epithelium derived factor in melanoma.

    PubMed

    Fernández-Barral, Asunción; Orgaz, Jose Luis; Baquero, Pablo; Ali, Zaheer; Moreno, Alberto; Tiana, María; Gómez, Valentí; Riveiro-Falkenbach, Erica; Cañadas, Carmen; Zazo, Sandra; Bertolotto, Corine; Davidson, Irwin; Rodríguez-Peralto, Jose Luis; Palmero, Ignacio; Rojo, Federico; Jensen, Lasse Dahl; del Peso, Luis; Jiménez, Benilde

    2014-06-01

    Pigment epithelium-derived factor (PEDF), a member of the serine protease inhibitor superfamily, has potent anti-metastatic effects in cutaneous melanoma through its direct actions on endothelial and melanoma cells. Here we show that PEDF expression positively correlates with microphthalmia-associated transcription factor (MITF) in melanoma cell lines and human samples. High PEDF and MITF expression is characteristic of low aggressive melanomas classified according to molecular and pathological criteria, whereas both factors are decreased in senescent melanocytes and naevi. Importantly, MITF silencing down-regulates PEDF expression in melanoma cell lines and primary melanocytes, suggesting that the correlation in the expression reflects a causal relationship. In agreement, analysis of Chromatin immunoprecipitation coupled to high throughput sequencing (ChIP-seq) data sets revealed three MITF binding regions within the first intron of SERPINF1, and reporter assays demonstrated that the binding of MITF to these regions is sufficient to drive transcription. Finally, we demonstrate that exogenous PEDF expression efficiently halts in vitro migration and invasion, as well as in vivo dissemination of melanoma cells induced by MITF silencing. In summary, these results identify PEDF as a novel transcriptional target of MITF and support a relevant functional role for the MITF-PEDF axis in the biology of melanoma.

  18. Asynchronous combinatorial action of four regulatory factors activates Bcl11b for T cell commitment.

    PubMed

    Kueh, Hao Yuan; Yui, Mary A; Ng, Kenneth K H; Pease, Shirley S; Zhang, Jingli A; Damle, Sagar S; Freedman, George; Siu, Sharmayne; Bernstein, Irwin D; Elowitz, Michael B; Rothenberg, Ellen V

    2016-08-01

    During T cell development, multipotent progenitors relinquish competence for other fates and commit to the T cell lineage by turning on Bcl11b, which encodes a transcription factor. To clarify lineage commitment mechanisms, we followed developing T cells at the single-cell level using Bcl11b knock-in fluorescent reporter mice. Notch signaling and Notch-activated transcription factors collaborate to activate Bcl11b expression irrespectively of Notch-dependent proliferation. These inputs work via three distinct, asynchronous mechanisms: an early locus 'poising' function dependent on TCF-1 and GATA-3, a stochastic-permissivity function dependent on Notch signaling, and a separate amplitude-control function dependent on Runx1, a factor already present in multipotent progenitors. Despite their necessity for Bcl11b expression, these inputs act in a stage-specific manner, providing a multitiered mechanism for developmental gene regulation. PMID:27376470

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

    SciTech Connect

    Xiang, Zhonghua; Qiao, Ling; Zhou, Yan; Babiuk, Lorne A.; Liu, Qiang

    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.

  20. Interferon regulatory factor-8 modulates the development of tumour-induced CD11b+Gr-1+ myeloid cells.

    PubMed

    Stewart, Trina J; Greeneltch, Kristy M; Reid, Julia E; Liewehr, David J; Steinberg, Seth M; Liu, Kebin; Abrams, Scott I

    2009-09-01

    Tumour-induced myeloid-derived suppressor cells (MDSC) promote immune suppression and mediate tumour progression. However, the molecular basis for the generation of MDSC, which in mice co-express the CD11b(+) and Gr-1(+) cell surface markers remains unclear. Because CD11b(+)Gr-1(+) cells expand during progressive tumour growth, this suggests that tumour-induced events alter signalling pathways that affect normal myeloid cell development. Interferon regulatory factor-8 (IRF-8), a member of the IFN-gamma regulatory factor family, is essential for normal myelopoiesis. We therefore examined whether IRF-8 modulated tumour-induced CD11b(+)Gr-1(+) cell development or accumulation using both implantable (4T1) and transgenic (MMTV-PyMT) mouse models of mammary tumour growth. In the 4T1 model, both splenic and bone marrow-derived CD11b(+)Gr-1(+) cells of tumour-bearing mice displayed a marked reduction in IRF-8 expression compared to control populations. A causal link between IRF-8 expression and the emergence of tumour-induced CD11b(+)Gr-1(+) cells was explored in vivo using a double transgenic (dTg) mouse model designed to express transgenes for both IRF-8 and mammary carcinoma development. Despite the fact that tumour growth was unaffected, splenomegaly, as well as the frequencies and absolute numbers of CD11b(+)Gr-1(+) cells were significantly lower in dTg mice when compared with single transgenic tumour-bearing mice. Overall, these data reveal that IRF-8 plays an important role in tumour-induced development and/or accumulation of CD11b(+)Gr-1(+) cells, and establishes a molecular basis for the potential manipulation of these myeloid populations for cancer therapy. PMID:20196788

  1. Calcifying nanoparticles (nanobacteria): an additional potential factor for urolithiasis in space flight crews.

    PubMed

    Jones, Jeffrey A; Ciftcioglu, Neva; Schmid, Josef F; Barr, Yael R; Griffith, Donald

    2009-01-01

    Spaceflight-induced microgravity appears to be a risk factor for the development of urinary calculi, resulting in urolithiasis during and after spaceflight. Calcifying nanoparticles, or nanobacteria, multiply more rapidly in simulated microgravity and create external shells of calcium phosphate. The question arises whether calcifying nanoparticles are nidi for calculi and contribute to the development of clinically significant urolithiasis in those who are predisposed to the development of urinary calculi because of intrinsic or extrinsic factors. This case report describes a calculus recovered after flight from an astronaut that, on morphologic and immunochemical analysis (including specific monoclonal antibody staining), demonstrated characteristics of calcifying nanoparticles. PMID:18718644

  2. Interferon regulatory factor 3 is a key regulation factor for inducing the expression of SAMHD1 in antiviral innate immunity

    PubMed Central

    Yang, Shen; Zhan, Yuan; Zhou, Yanjun; Jiang, Yifeng; Zheng, Xuchen; Yu, Lingxue; Tong, Wu; Gao, Fei; Li, Liwei; Huang, Qinfeng; Ma, Zhiyong; Tong, Guangzhi

    2016-01-01

    SAMHD1 is a type I interferon (IFN) inducible host innate immunity restriction factor that inhibits an early step of the viral life cycle. The underlying mechanisms of SAMHD1 transcriptional regulation remains elusive. Here, we report that inducing SAMHD1 upregulation is part of an early intrinsic immune response via TLR3 and RIG-I/MDA5 agonists that ultimately induce the nuclear translocation of the interferon regulation factor 3 (IRF3) protein. Further studies show that IRF3 plays a major role in upregulating endogenous SAMHD1 expression in a mechanism that is independent of the classical IFN-induced JAK-STAT pathway. Both overexpression and activation of IRF3 enhanced the SAMHD1 promoter luciferase activity, and activated IRF3 was necessary for upregulating SAMHD1 expression in a type I IFN cascade. We also show that the SAMHD1 promoter is a direct target of IRF3 and an IRF3 binding site is sufficient to render this promoter responsive to stimulation. Collectively, these findings indicate that upregulation of endogenous SAMHD1 expression is attributed to the phosphorylation and nuclear translocation of IRF3 and we suggest that type I IFN induction and induced SAMHD1 expression are coordinated. PMID:27411355

  3. Regulatory role of the 90-kDa-heat-shock protein (Hsp90) and associated factors on gene expression.

    PubMed

    Erlejman, Alejandra G; Lagadari, Mariana; Toneatto, Judith; Piwien-Pilipuk, Graciela; Galigniana, Mario D

    2014-02-01

    The term molecular chaperone was first used to describe the ability of nucleoplasmin to prevent the aggregation of histones with DNA during the assembly of nucleosomes. Subsequently, the name was extended to proteins that mediate the post-translational assembly of oligomeric complexes protecting them from denaturation and/or aggregation. Hsp90 is a 90-kDa molecular chaperone that represents the major soluble protein of the cell. In contrast to most conventional chaperones, Hsp90 functions as a refined sensor of protein function and its principal role in the cell is to facilitate biological activity to properly folded client proteins that already have a preserved tertiary structure. Consequently, Hsp90 is related to basic cell functions such as cytoplasmic transport of soluble proteins, translocation of client proteins to organelles, and regulation of the biological activity of key signaling factors such as protein kinases, ubiquitin ligases, steroid receptors, cell cycle regulators, and transcription factors. A growing amount of evidence links the protective action of this molecular chaperone to mechanisms related to posttranslational modifications of soluble nuclear factors as well as histones. In this article, we discuss some aspects of the regulatory action of Hsp90 on transcriptional regulation and how this effect could have impacted genetic assimilation mechanism in some organisms.

  4. Regulatory role of the 90-kDa-heat-shock protein (Hsp90) and associated factors on gene expression.

    PubMed

    Erlejman, Alejandra G; Lagadari, Mariana; Toneatto, Judith; Piwien-Pilipuk, Graciela; Galigniana, Mario D

    2014-02-01

    The term molecular chaperone was first used to describe the ability of nucleoplasmin to prevent the aggregation of histones with DNA during the assembly of nucleosomes. Subsequently, the name was extended to proteins that mediate the post-translational assembly of oligomeric complexes protecting them from denaturation and/or aggregation. Hsp90 is a 90-kDa molecular chaperone that represents the major soluble protein of the cell. In contrast to most conventional chaperones, Hsp90 functions as a refined sensor of protein function and its principal role in the cell is to facilitate biological activity to properly folded client proteins that already have a preserved tertiary structure. Consequently, Hsp90 is related to basic cell functions such as cytoplasmic transport of soluble proteins, translocation of client proteins to organelles, and regulation of the biological activity of key signaling factors such as protein kinases, ubiquitin ligases, steroid receptors, cell cycle regulators, and transcription factors. A growing amount of evidence links the protective action of this molecular chaperone to mechanisms related to posttranslational modifications of soluble nuclear factors as well as histones. In this article, we discuss some aspects of the regulatory action of Hsp90 on transcriptional regulation and how this effect could have impacted genetic assimilation mechanism in some organisms. PMID:24389346

  5. 21 CFR 1311.115 - Additional requirements for two-factor authentication.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ...) separate from the computer to which the practitioner is gaining access. (b) If one factor is a hard token, it must be separate from the computer to which it is gaining access and must meet at least the criteria of FIPS 140-2 Security Level 1, as incorporated by reference in § 1311.08, for...

  6. MicroRNA-23a downregulates the expression of interferon regulatory factor-1 in hepatocellular carcinoma cells.

    PubMed

    Yan, Yihe; Liang, Zhihai; Du, Qiang; Yang, Muqing; Geller, David A

    2016-08-01

    Interferon regulatory factor-1 (IRF-1) is a tumor-suppressor gene induced by interferon-γ (IFNγ) and plays an important role in the cell death of hepatocellular carcinoma (HCC). HCC tumors evade death in part by downregulating IRF-1 expression, yet the molecular mechanisms accounting for IRF-1 suppression in HCC have not yet been characterized. Previous studies have shown that microRNA-23a (miR-23a) can suppress apoptosis by targeting IRF-1. Therefore, we hypothesized that miR-23a promotes HCC growth by downregulating IRF-1. For the in vivo studies, 7 cases of resected HCC and adjacent liver samples were analyzed. For the in vitro studies, IRF-1 mRNA and protein were examined in HepG2 and Huh-7 HCC cells after IFNγ stimulation by real-time PCR and western blotting, respectively. To determine the role of miR-23a in regulating IRF-1, HepG2 cells were transfected with an miR-23a mimic or inhibitor, and IRF-1 expression was examined. Binding of miR-23a was assessed by cloning the 528-bp human IRF-1 3'-untranslated region (3'UTR) into luciferase reporter plasmid pMIR-IRF-1-3'UTR. The results showed that IRF-1 mRNA expression was downregulated in the human HCC tumor tissues compared to that in the adjacent background liver tissues. IFNγ-induced IRF-1 protein was less in the HepG2 tumor cells compared to that in the primary human hepatocytes. miR-23a expression was inversely correlated with IRF-1, and addition of the miR-23a inhibitor increased basal IRF-1 mRNA and protein. Likewise, the miR-23a mimic downregulated IFNγ-induced IRF-1 protein expression, while the miR-23a inhibitor increased IRF-1. Furthermore, the miR-23a mimic repressed IRF-1-3'UTR reporter activity, while the miR-23a inhibitor increased the reporter activity. These results demonstrated that IRF-1 expression is downregulated in human HCC tumors compared to that noted in the background liver. miR-23a downregulates the expression of IRF-1 in HCC cells, and the IRF-1 3'UTR has an miR‑23a binding

  7. MicroRNA-23a downregulates the expression of interferon regulatory factor-1 in hepatocellular carcinoma cells

    PubMed Central

    Yan, Yihe; Liang, Zhihai; Du, Qiang; Yang, Muqing; Geller, David A.

    2016-01-01

    Interferon regulatory factor-1 (IRF-1) is a tumor-suppressor gene induced by interferon-γ (IFNγ) and plays an important role in the cell death of hepatocellular carcinoma (HCC). HCC tumors evade death in part by downregulating IRF-1 expression, yet the molecular mechanisms accounting for IRF-1 suppression in HCC have not yet been characterized. Previous studies have shown that microRNA-23a (miR-23a) can suppress apoptosis by targeting IRF-1. Therefore, we hypothesized that miR-23a promotes HCC growth by down-regulating IRF-1. For the in vivo studies, 7 cases of resected HCC and adjacent liver samples were analyzed. For the in vitro studies, IRF-1 mRNA and protein were examined in HepG2 and Huh-7 HCC cells after IFNγ stimulation by real-time PCR and western blotting, respectively. To determine the role of miR-23a in regulating IRF-1, HepG2 cells were transfected with an miR-23a mimic or inhibitor, and IRF-1 expression was examined. Binding of miR-23a was assessed by cloning the 528-bp human IRF-1 3′-untranslated region (3′UTR) into luciferase reporter plasmid pMIR-IRF-1-3′UTR. The results showed that IRF-1 mRNA expression was down-regulated in the human HCC tumor tissues compared to that in the adjacent background liver tissues. IFNγ-induced IRF-1 protein was less in the HepG2 tumor cells compared to that in the primary human hepatocytes. miR-23a expression was inversely correlated with IRF-1, and addition of the miR-23a inhibitor increased basal IRF-1 mRNA and protein. Likewise, the miR-23a mimic downregulated IFNγ-induced IRF-1 protein expression, while the miR-23a inhibitor increased IRF-1. Furthermore, the miR-23a mimic repressed IRF-1-3′UTR reporter activity, while the miR-23a inhibitor increased the reporter activity. These results demonstrated that IRF-1 expression is downregulated in human HCC tumors compared to that noted in the background liver. miR-23a downregulates the expression of IRF-1 in HCC cells, and the IRF-1 3′UTR has an miR-23a

  8. The effect of nutritional additives on anti-infective factors in human milk.

    PubMed

    Quan, R; Yang, C; Rubinstein, S; Lewiston, N J; Stevenson, D K; Kerner, J A

    1994-06-01

    It has become a common practice to supplement human milk with a variety of additives to improve the nutritive content of the feeding for the premature infant. Twenty-two freshly frozen human milk samples were measured for lysozyme activity, total IgA, and specific IgA to Escherichia coli serotypes 01, 04, and 06. One mL aliquots were mixed with the following: 1 mL of Similac, Similac Special Care, Enfamil, Enfamil Premature Formula, and sterile water; 33 mL of Poly-Vi-Sol, 33 mg of Moducal, and 38 mg of breast-milk fortifier, and then reanalyzed. Significant decreases (41% to 74%) in lysozyme activity were seen with the addition of all formulas; breast-milk fortifier reduced activity by 19%, while no differences were seen with Moducal, sterile water, or Poly-Vi-Sol. No differences were seen in total IgA content, but some decreases were seen in specific IgA to E. coli serotypes 04 and 06. E. coli growth was determined after 3 1/2 hours of incubation at 37 degrees C after mixing. All cow-milk formulas enhanced E. coli growth; soy formulas and other additives preserved inhibition of bacterial growth. Nutritional additives can impair anti-infective properties of human milk, and such interplay should be considered in the decision on the feeding regimen of premature infants.

  9. 34 CFR 377.22 - What additional factors does the Secretary consider in making grants?

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... (Continued) OFFICE OF SPECIAL EDUCATION AND REHABILITATIVE SERVICES, DEPARTMENT OF EDUCATION DEMONSTRATION PROJECTS TO INCREASE CLIENT CHOICE PROGRAM How Does the Secretary Make an Award? § 377.22 What additional... strategies to increase client choice, in order to ensure that a variety of approaches are demonstrated...

  10. 34 CFR 377.22 - What additional factors does the Secretary consider in making grants?

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... (Continued) OFFICE OF SPECIAL EDUCATION AND REHABILITATIVE SERVICES, DEPARTMENT OF EDUCATION DEMONSTRATION PROJECTS TO INCREASE CLIENT CHOICE PROGRAM How Does the Secretary Make an Award? § 377.22 What additional... strategies to increase client choice, in order to ensure that a variety of approaches are demonstrated...

  11. Identification of hub genes and regulatory factors of glioblastoma multiforme subgroups by RNA-seq data analysis

    PubMed Central

    Li, Yanan; Min, Weijie; Li, Mengmeng; Han, Guosheng; Dai, Dongwei; Zhang, Lei; Chen, Xin; Wang, Xinglai; Zhang, Yuhui; Yue, Zhijian; Liu, Jianmin

    2016-01-01

    Glioblastoma multiforme (GBM) is the most common malignant brain tumor. This study aimed to identify the hub genes and regulatory factors of GBM subgroups by RNA sequencing (RNA-seq) data analysis, in order to explore the possible mechanisms responsbile for the progression of GBM. The dataset RNASeqV2 was downloaded by TCGA-Assembler, containing 169 GBM and 5 normal samples. Gene expression was calculated by the reads per kilobase per million reads measurement, and nor malized with tag count comparison. Following subgroup classification by the non-negative matrix factorization, the differentially expressed genes (DEGs) were screened in 4 GBM subgroups using the method of significance analysis of microarrays. Functional enrichment analysis was performed by DAVID, and the protein-protein interaction (PPI) network was constructed based on the HPRD database. The subgroup-related microRNAs (miRNAs or miRs), transcription factors (TFs) and small molecule drugs were predicted with predefined criteria. A cohort of 19,515 DEGs between the GBM and control samples was screened, which were predominantly enriched in cell cycle- and immunoreaction-related pathways. In the PPI network, lymphocyte cytosolic protein 2 (LCP2), breast cancer 1 (BRCA1), specificity protein 1 (Sp1) and chromodomain-helicase-DNA-binding protein 3 (CHD3) were the hub nodes in subgroups 1–4, respectively. Paired box 5 (PAX5), adipocyte protein 2 (aP2), E2F transcription factor 1 (E2F1) and cAMP-response element-binding protein-1 (CREB1) were the specific TFs in subgroups 1–4, respectively. miR-147b, miR-770-5p, miR-220a and miR-1247 were the particular miRNAs in subgroups 1–4, respectively. Natalizumab was the predicted small molecule drug in subgroup 2. In conclusion, the molecular regulatory mechanisms of GBM pathogenesis were distinct in the different subgroups. Several crucial genes, TFs, miRNAs and small molecules in the different GBM subgroups were identified, which may be used as potential

  12. Identification of hub genes and regulatory factors of glioblastoma multiforme subgroups by RNA-seq data analysis.

    PubMed

    Li, Yanan; Min, Weijie; Li, Mengmeng; Han, Guosheng; Dai, Dongwei; Zhang, Lei; Chen, Xin; Wang, Xinglai; Zhang, Yuhui; Yue, Zhijian; Liu, Jianmin

    2016-10-01

    Glioblastoma multiforme (GBM) is the most common malignant brain tumor. This study aimed to identify the hub genes and regulatory factors of GBM subgroups by RNA sequencing (RNA-seq) data analysis, in order to explore the possible mechanisms responsbile for the progression of GBM. The dataset RNASeqV2 was downloaded by TCGA-Assembler, containing 169 GBM and 5 normal samples. Gene expression was calculated by the reads per kilobase per million reads measurement, and nor malized with tag count comparison. Following subgroup classification by the non-negative matrix factorization, the differentially expressed genes (DEGs) were screened in 4 GBM subgroups using the method of significance analysis of microarrays. Functional enrichment analysis was performed by DAVID, and the protein-protein interaction (PPI) network was constructed based on the HPRD database. The subgroup-related microRNAs (miRNAs or miRs), transcription factors (TFs) and small molecule drugs were predicted with pre-defined criteria. A cohort of 19,515 DEGs between the GBM and control samples was screened, which were predominantly enriched in cell cycle- and immunoreaction-related pathways. In the PPI network, lymphocyte cytosolic protein 2 (LCP2), breast cancer 1 (BRCA1), specificity protein 1 (Sp1) and chromodomain-helicase-DNA-binding protein 3 (CHD3) were the hub nodes in subgroups 1-4, respectively. Paired box 5 (PAX5), adipocyte protein 2 (aP2), E2F transcription factor 1 (E2F1) and cAMP-response element-binding protein-1 (CREB1) were the specific TFs in subgroups 1-4, respectively. miR‑147b, miR‑770-5p, miR‑220a and miR‑1247 were the particular miRNAs in subgroups 1-4, respectively. Natalizumab was the predicted small molecule drug in subgroup 2. In conclusion, the molecular regulatory mechanisms of GBM pathogenesis were distinct in the different subgroups. Several crucial genes, TFs, miRNAs and small molecules in the different GBM subgroups were identified

  13. Regulatory role of neuron-restrictive silencing factor in expression of TRPC1

    SciTech Connect

    Ohba, Takayoshi; Watanabe, Hiroyuki; Takahashi, Yoichiro; Suzuki, Takashi; Miyoshi, Ichiro; Nakayama, Shinnsuke; Satoh, Eisaku; Iino, Kenji; Sasano, Hironobu; Mori, Yasuo; Kuromitsu, Sadao; Imagawa, Keiichi; Saito, Yoshihiko; Iijima, Toshihiko; Ito, Hiroshi; Murakami, Manabu . E-mail: mmura0123@hotmail.co.jp

    2006-12-22

    Neuron-restrictive silencer factor (NRSF) binds its consensus element to repress the transcription of various genes. The dominant-negative form (dnNRSF) has a hypertrophic effect on cardiogenesis through an unidentified mechanism. We examined the involvement of transient receptor potential (TRP) channel proteins, using transgenic mice overexpressing dnNRSF (dnNRSF mice). Electrophoretic mobility-shift assays revealed an interaction between NRSF and a neuron-restrictive silencer element-like sequence in intron 4 of TRPC1 genomic DNA. According to RT-PCR and Western analyses, TRPC1 was up-regulated in dnNRSF mouse heart. Transient overexpression of TRPC1 in HEK 293T cells increased the activity of the nuclear factor in activated T cells (NFAT) promoter and stimulated store-operated Ca{sup 2+} channel (SOCC)-mediated Ca{sup 2+} entry. Transfection of TRPC1 into primary cardiomyocytes increased NFAT activity, indicating a major role for TRPC1 in NFAT activation. Our findings strongly suggest that NRSF regulates TRP1 gene expression and causes changes in the levels of calcium entry through SOCCs.

  14. A model for genetic and epigenetic regulatory networks identifies rare pathways for transcription factor induced pluripotency

    NASA Astrophysics Data System (ADS)

    Artyomov, Maxim; Meissner, Alex; Chakraborty, Arup

    2010-03-01

    Most cells in an organism have the same DNA. Yet, different cell types express different proteins and carry out different functions. This is because of epigenetic differences; i.e., DNA in different cell types is packaged distinctly, making it hard to express certain genes while facilitating the expression of others. During development, upon receipt of appropriate cues, pluripotent embryonic stem cells differentiate into diverse cell types that make up the organism (e.g., a human). There has long been an effort to make this process go backward -- i.e., reprogram a differentiated cell (e.g., a skin cell) to pluripotent status. Recently, this has been achieved by transfecting certain transcription factors into differentiated cells. This method does not use embryonic material and promises the development of patient-specific regenerative medicine, but it is inefficient. The mechanisms that make reprogramming rare, or even possible, are poorly understood. We have developed the first computational model of transcription factor-induced reprogramming. Results obtained from the model are consistent with diverse observations, and identify the rare pathways that allow reprogramming to occur. If validated, our model could be further developed to design optimal strategies for reprogramming and shed light on basic questions in biology.

  15. Regulatory role of the OsWOX3A transcription factor in rice root development.

    PubMed

    Cho, Sung-Hwan; Paek, Nam-Chon

    2016-06-01

    WUSCHEL-related homeobox (WOX) transcription factors play important roles in cell fate determination during plant development and function in the signaling pathways of phytohormones such as gibberellic acid (GA). In a recent study, we demonstrated that OsWOX3A directly binds to the promoter of the KAO gene, which encodes ent-kaurenoic acid oxidase, an enzyme involved in GA biosynthesis, and represses KAO expression. Interestingly, we observed that OsWOX3A overexpression causes not only severe dwarfism, but also an increase in the number of lateral roots. Moreover, the expression of PIN-FORMED 1 (PIN1), involved in polar auxin transport, is significantly upregulated in OsWOX3A-OX plants. These findings indicate that OsWOX3A may be involved in modulation of GA-auxin crosstalk in rice root development. PMID:27171233

  16. The regulatory role of serum response factor pathway in neutrophil inflammatory response

    PubMed Central

    Taylor, Ashley; Halene, Stephanie

    2015-01-01

    Purpose of review Neutrophils rapidly migrate to sites of injury and infection. Egress of neutrophils from the circulation into tissues is a highly regulated process involving several distinct steps. Cell–cell interactions mediated by selectins and integrins and reorganization of the actin cytoskeleton are key mechanisms facilitating appropriate neutrophil recruitment. Neutrophil function is impaired in inherited and acquired disorders, such as leukocyte adhesion deficiency and myelodysplasia. Since the discovery that deletion of all or part of chromosome 5 is the most common genetic aberration in myelodysplasia, the roles of several of the deleted genes have been investigated in hematopoiesis. Several genes encoding proteins of the serum response factor (SRF) pathway are located on 5q. This review focuses, in particular, on the role of SRF in myeloid maturation and neutrophil function. Recent findings SRF and its pathway fulfill multiple complex roles in the regulation of the innate and adaptive immune system. Loss of SRF leads to defects in B-cell and T-cell development. SRF-deficient macrophages fail to spread, transmigrate, and phagocytose bacteria, and SRF-deficient neutrophils show defective chemotaxis in vitro and in vivo with failure of inside-out activation and trafficking of the Mac1 integrin complex. Loss of the formin mammalian Diaphanous 1, a regulator of linear actin polymerization and mediator of Ras homolog family member A signaling to SRF, results in aberrant myeloid differentiation and hyperactivity of the immune system. Summary SRF is an essential transcription factor in hematopoiesis and mature myeloid cell function. SRF regulates neutrophil migration, integrin activation, and trafficking. Disruption of the SRF pathway results in myelodysplasia and immune dysfunction. PMID:25402621

  17. Transcription factors RUNX1 and RUNX3 in the induction and suppressive function of Foxp3+ inducible regulatory T cells

    PubMed Central

    Klunker, Sven; Chong, Mark M.W.; Mantel, Pierre-Yves; Palomares, Oscar; Bassin, Claudio; Ziegler, Mario; Rückert, Beate; Meiler, Flurina; Akdis, Mübeccel; Littman, Dan R.

    2009-01-01

    Forkhead box P3 (FOXP3)+CD4+CD25+ inducible regulatory T (iT reg) cells play an important role in immune tolerance and homeostasis. In this study, we show that the transforming growth factor-β (TGF-β) induces the expression of the Runt-related transcription factors RUNX1 and RUNX3 in CD4+ T cells. This induction seems to be a prerequisite for the binding of RUNX1 and RUNX3 to three putative RUNX binding sites in the FOXP3 promoter. Inactivation of the gene encoding RUNX cofactor core-binding factor-β (CBFβ) in mice and small interfering RNA (siRNA)-mediated suppression of RUNX1 and RUNX3 in human T cells resulted in reduced expression of Foxp3. The in vivo conversion of naive CD4+ T cells into Foxp3+ iT reg cells was significantly decreased in adoptively transferred CbfbF/F CD4-cre naive T cells into Rag2−/− mice. Both RUNX1 and RUNX3 siRNA silenced human T reg cells and CbfbF/F CD4-cre mouse T reg cells showed diminished suppressive function in vitro. Circulating human CD4+ CD25high CD127− T reg cells significantly expressed higher levels of RUNX3, FOXP3, and TGF-β mRNA compared with CD4+CD25− cells. Furthermore, FOXP3 and RUNX3 were colocalized in human tonsil T reg cells. These data demonstrate Runx transcription factors as a molecular link in TGF-β–induced Foxp3 expression in iT reg cell differentiation and function. PMID:19917773

  18. Suppression of preoptic sleep-regulatory neuronal activity during corticotropin-releasing factor-induced sleep disturbance.

    PubMed

    Gvilia, Irma; Suntsova, Natalia; Kumar, Sunil; McGinty, Dennis; Szymusiak, Ronald

    2015-11-01

    Corticotropin releasing factor (CRF) is implicated in sleep and arousal regulation. Exogenous CRF causes sleep suppression that is associated with activation of at least two important arousal systems: pontine noradrenergic and hypothalamic orexin/hypocretin neurons. It is not known whether CRF also impacts sleep-promoting neuronal systems. We hypothesized that CRF-mediated changes in wake and sleep involve decreased activity of hypothalamic sleep-regulatory neurons localized in the preoptic area. To test this hypothesis, we examined the effects of intracerebroventricular administration of CRF on sleep-wake measures and c-Fos expression in GABAergic neurons in the median preoptic nucleus (MnPN) and ventrolateral preoptic area (VLPO) in different experimental conditions. Administration of CRF (0.1 nmol) during baseline rest phase led to delayed sleep onset and decreases in total amount and mean duration of non-rapid eye movement (NREM) sleep. Administration of CRF during acute sleep deprivation (SD) resulted in suppression of recovery sleep and decreased c-Fos expression in MnPN/VLPO GABAergic neurons. Compared with vehicle controls, intracerebroventricular CRF potentiated disturbances of both NREM and REM sleep in rats exposed to a species-specific psychological stressor, the dirty cage of a male conspecific. The number of MnPN/VLPO GABAergic neurons expressing c-Fos was reduced in the CRF-treated group of dirty cage-exposed rats. These findings confirm the involvement of CRF in wake-sleep cycle regulation and suggest that increased CRF signaling in the brain 1) negatively affects homeostatic responses to sleep loss, 2) exacerbates stress-induced disturbances of sleep, and 3) suppresses the activity of sleep-regulatory neurons of the MnPN and VLPO. PMID:26333784

  19. Evaluation of transcription factor that regulates T helper 17 and regulatory T cells function in periodontal health and disease

    PubMed Central

    Karthikeyan, B.; Talwar; Arun, K. V.; Kalaivani, S.

    2015-01-01

    Background: The differentiation of naοve T helper (Th) cells towards Th17 and regulatory T cells (Treg) is regulated by the transcription factors retinoic acid related orphan receptor gamma transcription (RORYt) and Forkhead box p3 (Foxp3), respectively. An imbalance in the activity of these transcription factors could result in the dysregulation of Th17/Treg response. Materials and Methods: Total RNA was isolated from gingival tissue obtained from 10 patients, each from periodontally healthy and diseased groups. The gene expression of RORYt and Foxp3 was measured by real-time reverse transcription polymerization chain reaction using total RNA isolates from gingival tissues group when compared to the healthy group, while Foxp3 demonstrated a 6.68 ± 0.03 fold decrease of expression in diseased group when compared to healthy group. Conclusion: Our results indicate a functional imbalance in the Th17/Treg response in periodontal disease group when compared to the periodontally healthy group. PMID:26538941

  20. HIV-1, interferon and the interferon regulatory factor system: an interplay between induction, antiviral responses and viral evasion.

    PubMed

    Marsili, Giulia; Remoli, Anna Lisa; Sgarbanti, Marco; Perrotti, Edvige; Fragale, Alessandra; Battistini, Angela

    2012-01-01

    Thirty years after the first isolation of the etiological agent of AIDS, the virus HIV-1 is still a major threat worldwide with millions of individuals currently infected. Although current combination therapies allow viral replication to be controlled, HIV-1 is not eradicated and persists in drug- and immune system-insensitive reservoirs and a cure is still lacking. Pathogens such as HIV-1 that cause chronic infections are able to adapt to the host in a manner that ensures long term residence and survival, via the evolution of numerous mechanisms that evade various aspects of the innate and adaptive immune response. One such mechanism is targeted to members of the interferon (IFN) regulatory factor (IRF) family of proteins. These transcription factors regulate a variety of biological processes including interferon induction, immune cell activation and downstream pattern recognition receptors (PRRs). HIV-1 renders IRFs harmless and hijacks them to its own advantage in order to facilitate its replication and evasion of immune responses. Type I interferon (IFN), the canonical antiviral innate response, can be induced in both acute and chronic HIV-1 infection in vivo, but in the majority of individuals this initial response is not protective and can contribute to disease progression. Type I IFN expression is largely inhibited in T cells and macrophages in order to successfully establish productive infection, whereas sustained IFN production by plasmacytoid dendritic cells is considered an important source of chronic immune activation, a hallmark to AIDS progression.

  1. Foot-and-mouth disease virus leader proteinase inhibits dsRNA-induced type I interferon transcription by decreasing interferon regulatory factor 3/7 in protein levels

    SciTech Connect

    Wang, Dang; Fang, Liurong; Luo, Rui; Ye, Rui; Fang, Ying; Xie, Lilan; Chen, Huanchun; Xiao, Shaobo

    2010-08-13

    Research highlights: {yields} FMDV L{sup pro} inhibits poly(I:C)-induced IFN-{alpha}1/{beta} mRNA expression. {yields} L{sup pro} inhibits MDA5-mediated activation of the IFN-{alpha}1/{beta} promoter. {yields} L{sup pro} significantly reduced the transcription of multiple IRF-responsive genes. {yields} L{sup pro} inhibits IFN-{alpha}1/{beta} promoter activation by decreasing IRF-3/7 in protein levels. {yields} The ability to process eIF-4G of L{sup pro} is not necessary to inhibit IFN-{alpha}1/{beta} activation. -- Abstract: The leader proteinase (L{sup pro}) of foot-and-mouth disease virus (FMDV) has been identified as an interferon-{beta} (IFN-{beta}) antagonist that disrupts the integrity of transcription factor nuclear factor {kappa}B (NF-{kappa}B). In this study, we showed that the reduction of double stranded RNA (dsRNA)-induced IFN-{alpha}1/{beta} expression caused by L{sup pro} was also associated with a decrease of interferon regulatory factor 3/7 (IRF-3/7) in protein levels, two critical transcription factors for activation of IFN-{alpha}/{beta}. Furthermore, overexpression of L{sup pro} significantly reduced the transcription of multiple IRF-responsive genes including 2',5'-OAS, ISG54, IP-10, and RANTES. Screening L{sup pro} mutants indicated that the ability to process eIF-4G of L{sup pro} is not required for suppressing dsRNA-induced activation of the IFN-{alpha}1/{beta} promoter and decreasing IRF-3/7 expression. Taken together, our results demonstrate that, in addition to disrupting NF-{kappa}B, L{sup pro} also decreases IRF-3/7 expression to suppress dsRNA-induced type I IFN production, suggesting multiple strategies used by FMDV to counteract the immune response to viral infection.

  2. Nerve growth factor-induced derepression of peripherin gene expression is associated with alterations in proteins binding to a negative regulatory element.

    PubMed Central

    Thompson, M A; Lee, E; Lawe, D; Gizang-Ginsberg, E; Ziff, E B

    1992-01-01

    The peripherin gene, which encodes a neuronal-specific intermediate filament protein, is transcriptionally induced with a late time course when nerve growth factor (NGF) stimulates PC12 cells to differentiate into neurons. We have studied its transcriptional regulation in order to better understand the neuronal-specific end steps of the signal transduction pathway of NGF. By 5' deletion mapping of the peripherin promoter, we have localized two positive regulatory elements necessary for full induction by NGF: a distal positive element and a proximal constitutive element within 111 bp of the transcriptional start site. In addition, there is a negative regulatory element (NRE; -179 to -111), the deletion of which results in elevated basal expression of the gene. Methylation interference footprinting of the NRE defined a unique sequence, GGCAGGGCGCC, as the binding site for proteins present in nuclear extracts from both undifferentiated and differentiated PC12 cells. However, DNA mobility shift assays using an oligonucleotide probe containing the footprinted sequence demonstrate a prominent retarded complex in extracts from undifferentiated PC12 cells which migrates with slower mobility than do the complexes produced by using differentiated PC12 cell extract. Transfection experiments using peripherin-chloramphenicol acetyltransferase constructs in which the footprinted sequence has been mutated confirm that the NRE has a functional, though not exclusive, role in repressing peripherin expression in undifferentiated and nonneuronal cells. We propose a two-step model of activation of peripherin by NGF in which dissociation of a repressor from the protein complex at the NRE, coupled with a positive signal from the distal positive element, results in depression of the gene. Images PMID:1588954

  3. Soil Pore Characteristics, an Underappreciated Regulatory Factor in GHGs Emission and C Stabilization

    NASA Astrophysics Data System (ADS)

    Toosi, E. R.; Yu, J.; Doane, T. A.; Guber, A.; Rivers, M. L.; Marsh, T. L.; Ali, K.; Kravchenko, A. N.

    2015-12-01

    Enduring challenges in understanding soil organic matter (SOM) stability and emission of greenhouse gases (GHGs) from soil stem from complexities of soil processes, many of which occur at micro-scales. The goal of this study is to evaluate the interactive effects soil pore characteristics, soil moisture levels, inherent SOM levels and properties, and substrate quality, on GHGs emission, and accelerated decomposition of native SOM following addition of fresh substrate i.e. priming. Our core hypothesis is that soil pore characteristics play a major role as a mediator in (i) the decomposition of organic matter regardless of its source (i.e. litter vs. native SOM) or substrate quality, as well as in (ii) GHGs emissions. Samples with prevalence of small (<10 μm) vs. large (>30 μm) pores were prepared from soils with similar properties but under long-term contrasting management. The samples were incubated (110 d) at low and optimum soil moisture conditions after addition of high quality (13C-soybean) and low quality (13C-corn) substrate. Headspace gas was analyzed for 13C-CO2 and GHGs on a regularly basis (day 1, 3, 7, 14, 24, 36, 48, 60, 72, 90, and 110). Selected samples were scanned at the early stage of decomposition (7, 14, 24 d) at 2-6 μm resolutions using X-ray computed μ tomography in order to: (1) quantify soil pore characteristics; (2) visualize and quantify distribution of soil moisture within samples of different pore characteristics; and (3) to visualize and measure losses of decomposing plant residue. Initial findings indicate that, consistent with our hypotheses, pore characteristics influenced GHGs emission, and intensity and pattern of plant residue decomposition. The importance of pores was highly pronounced in presence of added plant residue where greater N2O emission occurred in samples with dominant large pores, in contrast to CO2. Further findings will be discussed upon completion of the study and analysis of the results.

  4. The Regulatory Role of Nuclear Factor Kappa B in the Heart of Hereditary Hypertriglyceridemic Rat

    PubMed Central

    Vranková, Stanislava; Barta, Andrej; Klimentová, Jana; Dovinová, Ima; Líšková, Silvia; Dobešová, Zdenka; Pecháňová, Oľga; Kuneš, Jaroslav; Zicha, Josef

    2016-01-01

    Activation of nuclear factor-κB (NF-κB) by increased production of reactive oxygen species (ROS) might induce transcription and expression of different antioxidant enzymes and also of nitric oxide synthase (NOS) isoforms. Thus, we aimed at studying the effect of NF-κB inhibition, caused by JSH-23 (4-methyl-N1-(3-phenyl-propyl)-benzene-1,2-diamine) injection, on ROS and NO generation in hereditary hypertriglyceridemic (HTG) rats. 12-week-old, male Wistar and HTG rats were treated with JSH-23 (bolus, 10 μmol, i.v.). After one week, blood pressure (BP), superoxide dismutase (SOD) activity, SOD1, endothelial NOS (eNOS), and NF-κB (p65) protein expressions were higher in the heart of HTG rats compared to control rats. On the other hand, NOS activity was decreased. In HTG rats, JSH-23 treatment increased BP and heart conjugated dienes (CD) concentration (measured as the marker of tissue oxidative damage). Concomitantly, SOD activity together with SOD1 expression was decreased, while NOS activity and eNOS protein expression were increased significantly. In conclusion, NF-κB inhibition in HTG rats led to decreased ROS degradation by SOD followed by increased oxidative damage in the heart and BP elevation. In these conditions, increased NO generation may represent rather a counterregulatory mechanism activated by ROS. Nevertheless, this mechanism was not sufficient enough to compensate BP increase in HTG rats. PMID:27148433

  5. Transcription factor evolution in eukaryotes and the assembly of the regulatory toolkit in multicellular lineages.

    PubMed

    de Mendoza, Alex; Sebé-Pedrós, Arnau; Šestak, Martin Sebastijan; Matejcic, Marija; Torruella, Guifré; Domazet-Loso, Tomislav; Ruiz-Trillo, Iñaki

    2013-12-10

    Transcription factors (TFs) are the main players in transcriptional regulation in eukaryotes. However, it remains unclear what role TFs played in the origin of all of the different eukaryotic multicellular lineages. In this paper, we explore how the origin of TF repertoires shaped eukaryotic evolution and, in particular, their role into the emergence of multicellular lineages. We traced the origin and expansion of all known TFs through the eukaryotic tree of life, using the broadest possible taxon sampling and an updated phylogenetic background. Our results show that the most complex multicellular lineages (i.e., those with embryonic development, Metazoa and Embryophyta) have the most complex TF repertoires, and that these repertoires were assembled in a stepwise manner. We also show that a significant part of the metazoan and embryophyte TF toolkits evolved earlier, in their respective unicellular ancestors. To gain insights into the role of TFs in the development of both embryophytes and metazoans, we analyzed TF expression patterns throughout their ontogeny. The expression patterns observed in both groups recapitulate those of the whole transcriptome, but reveal some important differences. Our comparative genomics and expression data reshape our view on how TFs contributed to eukaryotic evolution and reveal the importance of TFs to the origins of multicellularity and embryonic development. PMID:24277850

  6. Sources and biology of regulatory factors active on mouse myeloid leukemic cells

    SciTech Connect

    Metcalf, D.

    1982-01-01

    The action of serum or cells in enforcing differentiation in mouse myelomonocytic leukemic cells was monitored in agar cultures of WEHI-3B leukemic cells. The repeated intravenous injection of 5 ..mu..g endotoxin initially increased serum differentiating activity but after the third injection responses to further injections decreased markedly. Congenitally athymic (nude) mice exhibited normal rises in serum differentiating activity when injected with endotoxin but C3H HeJ mice failed to respond to challenge with purified lipid A. Whole body irradiation up to 1,200 rads did not increase serum differentiating activity but did not suppress responses to challenge injection of endotoxin. Coculture of WEHI-3B cells with peritoneal cells from normal or irradiated BALB/c mice caused marked granulocytic differentiation in WEHI-3B colonies. This effect was not seen if leukemic cells were cultured with thymus, spleen, or bone marrow cells. The serum halflife of the factor in postendotoxin serum enforcing differentiation of WEHI-3B cells was shown to be 1.5-2.3 hr.

  7. Myogenic regulatory factors during regeneration of skeletal muscle in young, adult, and old rats

    NASA Technical Reports Server (NTRS)

    Marsh, D. R.; Criswell, D. S.; Carson, J. A.; Booth, F. W.

    1997-01-01

    Myogenic factor mRNA expression was examined during muscle regeneration after bupivacaine injection in Fischer 344/Brown Norway F1 rats aged 3, 18, and 31 mo of age (young, adult, and old, respectively). Mass of the tibialis anterior muscle in the young rats had recovered to control values by 21 days postbupivacaine injection but in adult and old rats remained 40% less than that of contralateral controls at 21 and 28 days of recovery. During muscle regeneration, myogenin mRNA was significantly increased in muscles of young, adult, and old rats 5 days after bupivacaine injection. Subsequently, myogenin mRNA levels in young rat muscle decreased to postinjection control values by day 21 but did not return to control values in 28-day regenerating muscles of adult and old rats. The expression of MyoD mRNA was also increased in muscles at day 5 of regeneration in young, adult, and old rats, decreased to control levels by day 14 in young and adult rats, and remained elevated in the old rats for 28 days. In summary, either a diminished ability to downregulate myogenin and MyoD mRNAs in regenerating muscle occurs in old rat muscles, or the continuing myogenic effort includes elevated expression of these mRNAs.

  8. Porcine complement regulatory protein CD46 and heparan sulfates are the major factors for classical swine fever virus attachment in vitro.

    PubMed

    Dräger, Carolin; Beer, Martin; Blome, Sandra

    2015-03-01

    Classical swine fever virus (CSFV) is the causative agent of a severe multi-systemic disease of pigs. While several aspects of virus-host-interaction are known, the early steps of infection remain unclear. For the closely related bovine viral diarrhea virus (BVDV), a cellular receptor is known: bovine complement regulatory protein CD46. Given that these two pestiviruses are closely related, porcine CD46 is also a candidate receptor for CSFV. In addition to CD46, cell-culture-adapted CSFV strains have been shown to use heparan sulfates as an additional cellular factor. In the present study, the interaction of field-type and cell-culture-adapted CSFV with a permanent porcine cell line or primary macrophages was assessed using anti-porcine CD46 monoclonal antibodies and a heparan-sulfate-blocking compound, DSTP-27. The influence of receptor blocking was assessed using virus titration and quantitative PCR. Treatment of cells with monoclonal antibodies against porcine CD46 led to a reduction of viral growth in both cell types. The effect was most pronounced with field-type CSFV. The blocking could be enhanced by addition of DSTP-27, especially for cell-culture-adapted CSFV. The combined use of both blocking agents led to a significant reduction of viral growth but was also not able to abolish infection completely. The results obtained in this study showed that both porcine CD46 and heparan sulfates play a major role in the initial steps of CSFV infection. Additional receptors might also play a role for attachment and entry; however, their impact is obviously limited in vitro in comparison to CD46 and heparan sulfates.

  9. Studying Dynamic Features in Myocardial Infarction Progression by Integrating miRNA-Transcription Factor Co-Regulatory Networks and Time-Series RNA Expression Data from Peripheral Blood Mononuclear Cells.

    PubMed

    Shi, Hongbo; Zhang, Guangde; Wang, Jing; Wang, Zhenzhen; Liu, Xiaoxia; Cheng, Liang; Li, Weimin

    2016-01-01

    Myocardial infarction (MI) is a serious heart disease and a leading cause of mortality and morbidity worldwide. Although some molecules (genes, miRNAs and transcription factors (TFs)) associated with MI have been studied in a specific pathological context, their dynamic characteristics in gene expressions, biological functions and regulatory interactions in MI progression have not been fully elucidated to date. In the current study, we analyzed time-series RNA expression data from peripheral blood mononuclear cells. We observed that significantly differentially expressed genes were sharply up- or down-regulated in the acute phase of MI, and then changed slowly until the chronic phase. Biological functions involved at each stage of MI were identified. Additionally, dynamic miRNA-TF co-regulatory networks were constructed based on the significantly differentially expressed genes and miRNA-TF co-regulatory motifs, and the dynamic interplay of miRNAs, TFs and target genes were investigated. Finally, a new panel of candidate diagnostic biomarkers (STAT3 and ICAM1) was identified to have discriminatory capability for patients with or without MI, especially the patients with or without recurrent events. The results of the present study not only shed new light on the understanding underlying regulatory mechanisms involved in MI progression, but also contribute to the discovery of true diagnostic biomarkers for MI. PMID:27367417

  10. Studying Dynamic Features in Myocardial Infarction Progression by Integrating miRNA-Transcription Factor Co-Regulatory Networks and Time-Series RNA Expression Data from Peripheral Blood Mononuclear Cells

    PubMed Central

    Wang, Jing; Wang, Zhenzhen; Liu, Xiaoxia; Cheng, Liang; Li, Weimin

    2016-01-01

    Myocardial infarction (MI) is a serious heart disease and a leading cause of mortality and morbidity worldwide. Although some molecules (genes, miRNAs and transcription factors (TFs)) associated with MI have been studied in a specific pathological context, their dynamic characteristics in gene expressions, biological functions and regulatory interactions in MI progression have not been fully elucidated to date. In the current study, we analyzed time-series RNA expression data from peripheral blood mononuclear cells. We observed that significantly differentially expressed genes were sharply up- or down-regulated in the acute phase of MI, and then changed slowly until the chronic phase. Biological functions involved at each stage of MI were identified. Additionally, dynamic miRNA–TF co-regulatory networks were constructed based on the significantly differentially expressed genes and miRNA–TF co-regulatory motifs, and the dynamic interplay of miRNAs, TFs and target genes were investigated. Finally, a new panel of candidate diagnostic biomarkers (STAT3 and ICAM1) was identified to have discriminatory capability for patients with or without MI, especially the patients with or without recurrent events. The results of the present study not only shed new light on the understanding underlying regulatory mechanisms involved in MI progression, but also contribute to the discovery of true diagnostic biomarkers for MI. PMID:27367417

  11. Effects of estrogen on osteoprogenitor cells and cytokines/bone-regulatory factors in postmenopausal women.

    PubMed

    Mödder, Ulrike I; Roforth, Matthew M; Hoey, Kelley; McCready, Louise K; Peterson, James M; Monroe, David G; Oursler, Merry Jo; Khosla, Sundeep

    2011-08-01

    Decreases in estrogen levels contribute not only to early postmenopausal bone loss but also to bone loss with aging. While estrogen is critical for the maintenance of bone formation, the mechanism(s) of this effect remain unclear. Thus, we assessed the effects of 4months of transdermal estradiol treatment (0.05mg/day) of postmenopausal women as compared to no treatment (n=16 per group) on the expression of genes in pre-specified pathways in freshly isolated bone marrow osteoprogenitor cells (hematopoietic lineage [lin]-/Stro1+). We also evaluated whether estrogen treatment modulated peripheral blood or bone marrow plasma levels of the Wnt antagonists, sclerostin and DKK1, as well as serotonin, OPG, RANKL, adiponectin, oxytocin, and inflammatory cytokines (TNFα, IL-1β, and IL-6), as each of these molecules have recently been shown to play an important role in regulating osteoblast function and/or being responsive to estrogen. We observed a significant decrease in the expression of several proliferation markers (cyclin B1, cyclin E1, E2F1) and increase in adhesion molecules (N-cadherin) in bone marrow lin-/Stro1+ cells from estrogen-treated compared to control women. None of the peripheral blood or bone marrow plasma marker levels differed between the two groups, with the exception of sclerostin levels, which were significantly lower in the estrogen-treated as compared to the control women in peripheral serum (by 32%, P=0.009) and in bone marrow plasma (by 34%, P=0.017). There were significant differences in bone marrow versus peripheral plasma levels of several factors: sclerostin and OPG levels were higher in bone marrow as compared to peripheral plasma, whereas serotonin and adiponectin levels were higher in peripheral as compared to bone marrow plasma. In summary, our data directly assessing possible regulation by estrogen of osteoprogenitor cells in humans indicate that, consistent with previous studies in mice, estrogen suppresses the proliferation of human

  12. Basolateral Na+/HCO3– cotransport activity is regulated by the dissociable Na+/H+ exchanger regulatory factor

    PubMed Central

    Bernardo, Angelito A.; Kear, Felicidad T.; Santos, Anna V.P.; Ma, Jianfei; Steplock, Debra; Robey, R. Brooks; Weinman, Edward J.

    1999-01-01

    In the renal proximal tubule, the activities of the basolateral Na+/HCO3– cotransporter (NBC) and the apical Na+/H+ exchanger (NHE3) uniformly vary in parallel, suggesting that they are coordinately regulated. PKA-mediated inhibition of NHE3 is mediated by a PDZ motif–containing protein, the Na+/H+ exchanger regulatory factor (NHE-RF). Given the common inhibition of these transporters after protein kinase A (PKA) activation, we sought to determine whether NHE-RF also plays a role in PKA-regulated NBC activity. Renal cortex immunoblot analysis using anti-peptide antibodies directed against rabbit NHE-RF demonstrated the presence of this regulatory factor in both brush-border membranes (BBMs) and basolateral membranes (BLMs). Using a reconstitution assay, we found that limited trypsin digestion of detergent solubilized rabbit renal BLM preparations resulted in NBC activity that was unaffected by PKA activation. Co-reconstitution of these trypsinized preparations with a recombinant protein corresponding to wild-type rabbit NHE-RF restored the inhibitory effect of PKA on NBC activity in a concentration-dependent manner. NBC activity was inhibited 60% by 10–8M NHE-RF; this effect was not observed in the absence of PKA. Reconstitution with heat-denatured NHE-RF also failed to attenuate NBC activity. To establish further a physiologic role for NHE-RF in NBC regulation, the renal epithelial cell line B-SC-1, which lacks detectable endogenous NHE-RF expression, was engineered to express stably an NHE-RF transgene. NHE-RF–expressing B-SC-1 cells (B-SC-RF) exhibited markedly lower basal levels of NBC activity than did wild-type controls. Inhibition of NBC activity in B-SC-RF cells was enhanced after 10 μM of forskolin treatment, consistent with a postulated role for NHE-RF in mediating the inhibition of NBC activity by PKA. These findings not only suggest NHE-RF involvement in PKA-regulated NBC activity, but also provide a unique molecular mechanism whereby

  13. Aitchbone hanging and ageing period are additive factors influencing pork eating quality.

    PubMed

    Channon, H A; Taverner, M R; D'Souza, D N; Warner, R D

    2014-01-01

    The effects of abattoir, carcase weight (60 or 80 kg HCW), hanging method (Achilles or aitchbone) and ageing period (2 or 7 day post-slaughter) on eating quality attributes of pork were investigated in this 3×2×2×2 factorial study. A total of 144 Large White×Landrace female pigs were slaughtered at one of three abattoirs and sides hung from either the Achilles tendon or the aitchbone. After 24 h chilling, loin (M. longissimus thoracis et lumborum) and topside (M. semimembranosus) muscles were individually vacuum packaged and aged for 2 or 7 days post-slaughter. Consumers (n=852) evaluated eating quality. Neither abattoir nor carcase weight influenced tenderness, flavour or overall liking of pork. Improvements in tenderness, flavour and overall liking were found due to aitchbone hanging (P<0.001) and ageing (P<0.001) for 7 days compared with Achilles-hung carcases and pork aged for 2 days, respectively. This study demonstrated that aitchbone hanging and 7 day ageing can improve eating quality, but these effects were additive as the interaction term was not significant. PMID:24013699

  14. Electrical inhibition of lens epithelial cell proliferation: an additional factor in secondary cataract?

    PubMed Central

    Wang, Entong; Reid, Brian; Lois, Noemi; Forrester, John V.; McCaig, Colin D.; Zhao, Min

    2005-01-01

    Cataract is the most common cause of blindness but is at least curable by surgery. Unfortunately, many patients gradually develop the complication of posterior capsule opacification (PCO) or secondary cataract. This arises from stimulated cell growth within the lens capsule and can greatly impair vision. It is not fully understood why residual lens epithelial cell growth occurs after surgery. We propose and show that cataract surgery might remove an important inhibitory factor for lens cell growth, namely electric fields. The lens generates a unique pattern of electric currents constantly flowing out from the equator and entering the anterior and posterior poles. We show here that cutting and removing part of the anterior capsule as in cataract surgery significantly decreases the equatorial outward electric currents. Application of electric fields in culture inhibits proliferation of human lens epithelial cells. This inhibitory effect is likely to be mediated through a cell cycle control mechanism that decreases entry of cells into S phase from G1 phase by decreasing the G1-specific cell cycle protein cyclin E and increasing the cyclin-Cdk complex inhibitor p27kip1. Capsulorrhexis in vivo, which reduced endogenous lens electric fields, significantly increased LEC growth. This, together with our previous findings that electric fields have significant effects on the direction of lens cell migration, points to a controlling mechanism for the aberrant cell growth in posterior capsule opacification. A novel approach to control growth of lens epithelial cells using electric fields combined with other controlling mechanisms may be more effective in the prevention and treatment of this common complication of cataract surgery. PMID:15764648

  15. Insulin resistance: an additional risk factor in the pathogenesis of cardiovascular disease in type 2 diabetes.

    PubMed

    Patel, Tushar P; Rawal, Komal; Bagchi, Ashim K; Akolkar, Gauri; Bernardes, Nathalia; Dias, Danielle da Silva; Gupta, Sarita; Singal, Pawan K

    2016-01-01

    Sedentary life style and high calorie dietary habits are prominent leading cause of metabolic syndrome in modern world. Obesity plays a central role in occurrence of various diseases like hyperinsulinemia, hyperglycemia and hyperlipidemia, which lead to insulin resistance and metabolic derangements like cardiovascular diseases (CVDs) mediated by oxidative stress. The mortality rate due to CVDs is on the rise in developing countries. Insulin resistance (IR) leads to micro or macro angiopathy, peripheral arterial dysfunction, hampered blood flow, hypertension, as well as the cardiomyocyte and the endothelial cell dysfunctions, thus increasing risk factors for coronary artery blockage, stroke and heart failure suggesting that there is a strong association between IR and CVDs. The plausible linkages between these two pathophysiological conditions are altered levels of insulin signaling proteins such as IR-β, IRS-1, PI3K, Akt, Glut4 and PGC-1α that hamper insulin-mediated glucose uptake as well as other functions of insulin in the cardiomyocytes and the endothelial cells of the heart. Reduced AMPK, PFK-2 and elevated levels of NADP(H)-dependent oxidases produced by activated M1 macrophages of the adipose tissue and elevated levels of circulating angiotensin are also cause of CVD in diabetes mellitus condition. Insulin sensitizers, angiotensin blockers, superoxide scavengers are used as therapeutics in the amelioration of CVD. It evidently becomes important to unravel the mechanisms of the association between IR and CVDs in order to formulate novel efficient drugs to treat patients suffering from insulin resistance-mediated cardiovascular diseases. The possible associations between insulin resistance and cardiovascular diseases are reviewed here. PMID:26542377

  16. The Transcription Factors Grainyhead-like 2 and NK2-Homeobox 1 Form a Regulatory Loop That Coordinates Lung Epithelial Cell Morphogenesis and Differentiation*

    PubMed Central

    Varma, Saaket; Cao, Yuxia; Tagne, Jean-Bosco; Lakshminarayanan, Meenakshi; Li, Jun; Friedman, Thomas B.; Morell, Robert J.; Warburton, David; Kotton, Darrell N.; Ramirez, Maria I.

    2012-01-01

    The Grainyhead family of transcription factors controls morphogenesis and differentiation of epithelial cell layers in multicellular organisms by regulating cell junction- and proliferation-related genes. Grainyhead-like 2 (Grhl2) is expressed in developing mouse lung epithelium and is required for normal lung organogenesis. The specific epithelial cells expressing Grhl2 and the genes regulated by Grhl2 in normal lungs are mostly unknown. In these studies we identified the NK2-homeobox 1 transcription factor (Nkx2-1) as a direct transcriptional target of Grhl2. By binding and transcriptional assays and by confocal microscopy we showed that these two transcription factors form a positive feedback loop in vivo and in cell lines and are co-expressed in lung bronchiolar and alveolar type II cells. The morphological changes observed in flattening lung alveolar type II cells in culture are associated with down-regulation of Grhl2 and Nkx2-1. Reduction of Grhl2 in lung epithelial cell lines results in lower expression levels of Nkx2-1 and of known Grhl2 target genes. By microarray analysis we identified that in addition to Cadherin1 and Claudin4, Grhl2 regulates other cell interaction genes such as semaphorins and their receptors, which also play a functional role in developing lung epithelium. Impaired collective cell migration observed in Grhl2 knockdown cell monolayers is associated with reduced expression of these genes and may contribute to the altered epithelial phenotype reported in Grhl2 mutant mice. Thus, Grhl2 functions at the nexus of a novel regulatory network, connecting lung epithelial cell identity, migration, and cell-cell interactions. PMID:22955271

  17. Physiological and receptor-selective retinoids modulate interferon gamma signaling by increasing the expression, nuclear localization, and functional activity of interferon regulatory factor-1.

    PubMed

    Luo, Xin M; Ross, A Catharine

    2005-10-28

    Synergistic actions between all-trans-retinoic acid (atRA) and interferon gamma (IFNgamma) on modulation of cellular functions have been reported both in vitro and in vivo. However, the mechanism of atRA-mediated regulation of IFNgamma signaling is poorly understood. In this study, we have used the human lung epithelial cell line A549 to examine the effect of atRA on IFNgamma-induced expression of IFN regulatory factor-1 (IRF-1), an important transcription factor involved in cell growth and apoptosis, differentiation, and antiviral and antibacterial immune responses. At least 4 h of pretreatment with atRA followed by suboptimal concentrations of IFNgamma induced a faster, higher, and more stable expression of IRF-1 than IFNgamma alone. Actinomycin D completely blocked the induction of IRF-1 by the combination, suggesting regulation at the transcriptional level. Further, we found that activation of signal transducer and activator of transcription-1 was induced more dramatically by atRA and IFNgamma than by IFNgamma alone. Expression of IFNgamma receptor-1 on the cell surface was also increased upon atRA pretreatment. Experiments using receptor-selective retinoids revealed that ligands for retinoic acid receptor-alpha (RARalpha), including atRA, 9-cis-retinoic acid, and Am580, sequentially increased the levels of IFNgamma receptor-1, activated signal transducer and activator of transcription-1, and IRF-1 and that an RARalpha antagonist was able to inhibit the effects of atRA and Am580. In addition, atRA pretreatment affected the transcriptional functions of IFNgamma-induced IRF-1, increasing its nuclear localization and DNA binding activity as well as the transcript levels of IRF-1 target genes. These results suggest that atRA, an RARalpha ligand, regulates IFNgamma-induced IRF-1 by affecting multiple components of the IFNgamma signaling pathway, from the plasma membrane to the nuclear transcription factors.

  18. Regulatory SNPs and transcriptional factor binding sites in ADRBK1, AKT3, ATF3, DIO2, TBXA2R and VEGFA

    PubMed Central

    Buroker, Norman E

    2014-01-01

    Abstract Regulatory single nucleotide polymorphisms (rSNPs) which change the transcriptional factor binding sites (TFBS) for transcriptional factors (TFs) to bind DNA were reviewed for the ADRBK1 (GRK2), AKT3, ATF3, DIO2, TBXA2R and VEGFA genes. Changes in the TFBS where TFs attach to regulate these genes may result in human sickness and disease. The highlights of this previous work were reviewed for these genes. PMID:25483406

  19. Auxin Response Factor SlARF2 Is an Essential Component of the Regulatory Mechanism Controlling Fruit Ripening in Tomato.

    PubMed

    Hao, Yanwei; Hu, Guojian; Breitel, Dario; Liu, Mingchun; Mila, Isabelle; Frasse, Pierre; Fu, Yongyao; Aharoni, Asaph; Bouzayen, Mondher; Zouine, Mohamed

    2015-12-01

    Ethylene is the main regulator of climacteric fruit ripening, by contrast the putative role of other phytohormones in this process remains poorly understood. The present study brings auxin signaling components into the mechanism regulating tomato fruit ripening through the functional characterization of Auxin Response Factor2 (SlARF2) which encodes a downstream component of auxin signaling. Two paralogs, SlARF2A and SlARF2B, are found in the tomato genome, both displaying a marked ripening-associated expression but distinct responsiveness to ethylene and auxin. Down-regulation of either SlARF2A or SlARF2B resulted in ripening defects while simultaneous silencing of both genes led to severe ripening inhibition suggesting a functional redundancy among the two ARFs. Tomato fruits under-expressing SlARF2 produced less climacteric ethylene and exhibited a dramatic down-regulation of the key ripening regulators RIN, CNR, NOR and TAGL1. Ethylene treatment failed to reverse the non-ripening phenotype and the expression of ethylene signaling and biosynthesis genes was strongly altered in SlARF2 down-regulated fruits. Although both SlARF proteins are transcriptional repressors the data indicate they work as positive regulators of tomato fruit ripening. Altogether, the study defines SlARF2 as a new component of the regulatory network controlling the ripening process in tomato. PMID:26716451

  20. Three novel acetylation sites in the Foxp3 transcription factor regulate the suppressive activity of regulatory T cells

    PubMed Central

    Kwon, Hye-Sook; Lim, Hyung W.; Wu, Jessica; Schnoelzer, Martina; Verdin, Eric; Ott, Melanie

    2012-01-01

    The Foxp3 transcription factor is the master regulator of regulatory T cell (Treg) differentiation and function. Its activity is regulated by reversible acetylation. Using mass spectrometry of immunoprecipitated proteins, we identify three novel acetylation sites in murine Foxp3 (K31, K262, and K267) and the corresponding sites in human FoxP3 proteins. Newly raised modification-specific antibodies against acetylated K31 and K267 confirm acetylation of these residues in murine Tregs. Mutant Foxp3 proteins carrying arginine substitutions at the three acetylation sites (3KR) accumulate in T cells to higher levels than wildtype Foxp3 and exert better suppressive activity in co-culture experiments. Acetylation and stability of wildtype, but not mutant, Foxp3 is enhanced when cells are treated with Ex-527, an inhibitor of the NAD+-dependent deacetylase SIRT1. Treatment with Ex-527 promotes Foxp3 expression during induced Treg differentiation, enhances Foxp3 levels in natural Tregs, and prevents loss of Foxp3 expression in adoptively transferred Tregs in mice. Our data identify SIRT1 as a negative regulator of Treg function via deacetylation of three novel target sites in Foxp3. SIRT1 inhibitors strengthen the suppressive activity of Tregs and may be useful in enhancing Treg-based therapeutic approaches to autoimmune diseases or graft rejections. PMID:22312127

  1. The PDZ Protein Na+/H+ Exchanger Regulatory Factor-1 (NHERF1) Regulates Planar Cell Polarity and Motile Cilia Organization

    PubMed Central

    Stolz, Donna B.; Tsang, Michael; Friedman, Peter A.; Romero, Guillermo

    2016-01-01

    Directional flow of the cerebrospinal fluid requires coordinated movement of the motile cilia of the ependymal epithelium that lines the cerebral ventricles. Here we report that mice lacking the Na+/H+ Exchanger Regulatory Factor 1 (NHERF1/Slc9a3r1, also known as EBP50) develop profound communicating hydrocephalus associated with fewer and disorganized ependymal cilia. Knockdown of NHERF1/slc9a3r1 in zebrafish embryos also causes severe hydrocephalus of the hindbrain and impaired ciliogenesis in the otic vesicle. Ultrastructural analysis did not reveal defects in the shape or organization of individual cilia. Similar phenotypes have been described in animals with deficiencies in Wnt signaling and the Planar Cell Polarity (PCP) pathway. We show that NHERF1 binds the PCP core genes Frizzled (Fzd) and Vangl. We further show that NHERF1 assembles a ternary complex with Fzd4 and Vangl2 and promotes translocation of Vangl2 to the plasma membrane, in particular to the apical surface of ependymal cells. Taken together, these results strongly support an important role for NHERF1 in the regulation of PCP signaling and the development of functional motile cilia. PMID:27055101

  2. Auxin Response Factor SlARF2 Is an Essential Component of the Regulatory Mechanism Controlling Fruit Ripening in Tomato

    PubMed Central

    Hao, Yanwei; Hu, Guojian; Breitel, Dario; Liu, Mingchun; Mila, Isabelle; Frasse, Pierre; Fu, Yongyao; Aharoni, Asaph; Bouzayen, Mondher; Zouine, Mohamed

    2015-01-01

    Ethylene is the main regulator of climacteric fruit ripening, by contrast the putative role of other phytohormones in this process remains poorly understood. The present study brings auxin signaling components into the mechanism regulating tomato fruit ripening through the functional characterization of Auxin Response Factor2 (SlARF2) which encodes a downstream component of auxin signaling. Two paralogs, SlARF2A and SlARF2B, are found in the tomato genome, both displaying a marked ripening-associated expression but distinct responsiveness to ethylene and auxin. Down-regulation of either SlARF2A or SlARF2B resulted in ripening defects while simultaneous silencing of both genes led to severe ripening inhibition suggesting a functional redundancy among the two ARFs. Tomato fruits under-expressing SlARF2 produced less climacteric ethylene and exhibited a dramatic down-regulation of the key ripening regulators RIN, CNR, NOR and TAGL1. Ethylene treatment failed to reverse the non-ripening phenotype and the expression of ethylene signaling and biosynthesis genes was strongly altered in SlARF2 down-regulated fruits. Although both SlARF proteins are transcriptional repressors the data indicate they work as positive regulators of tomato fruit ripening. Altogether, the study defines SlARF2 as a new component of the regulatory network controlling the ripening process in tomato. PMID:26716451

  3. Virulence factor regulation and regulatory networks in Streptococcus pyogenes and their impact on pathogen-host interactions.

    PubMed

    Kreikemeyer, Bernd; McIver, Kevin S; Podbielski, Andreas

    2003-05-01

    Streptococcus pyogenes (group A streptococcus, GAS) is a very important human pathogen with remarkable adaptation capabilities. Survival within the harsh host surroundings requires sensing potential on the bacterial side, which leads in particular to coordinately regulated virulence factor expression. GAS 'stand-alone' response regulators (RRs) and two-component signal transduction systems (TCSs) link the signals from the host environment with adaptive responses of the bacterial cell. Numerous putative regulatory systems emerged from GAS genome sequences. Only three RRs [Mga, RofA-like protein (RALP) and Rgg/RopB] and three TCSs (CsrRS/CovRS, FasBCAX and Ihk/Irr) have been studied in some detail with respect to their growth-phase-dependent activity and their influence on GAS-host cell interaction. In particular, the Mga-, RALP- and Rgg/RopB-regulated pathways display interconnected activities that appear to influence GAS colonization, persistence and spreading mechanisms, in a growth-phase-related fashion. Here, we have summarized our current knowledge about these RRs and TCSs to highlight the questions that should be addressed in future research on GAS pathogenicity.

  4. Auxin Response Factor SlARF2 Is an Essential Component of the Regulatory Mechanism Controlling Fruit Ripening in Tomato.

    PubMed

    Hao, Yanwei; Hu, Guojian; Breitel, Dario; Liu, Mingchun; Mila, Isabelle; Frasse, Pierre; Fu, Yongyao; Aharoni, Asaph; Bouzayen, Mondher; Zouine, Mohamed

    2015-12-01

    Ethylene is the main regulator of climacteric fruit ripening, by contrast the putative role of other phytohormones in this process remains poorly understood. The present study brings auxin signaling components into the mechanism regulating tomato fruit ripening through the functional characterization of Auxin Response Factor2 (SlARF2) which encodes a downstream component of auxin signaling. Two paralogs, SlARF2A and SlARF2B, are found in the tomato genome, both displaying a marked ripening-associated expression but distinct responsiveness to ethylene and auxin. Down-regulation of either SlARF2A or SlARF2B resulted in ripening defects while simultaneous silencing of both genes led to severe ripening inhibition suggesting a functional redundancy among the two ARFs. Tomato fruits under-expressing SlARF2 produced less climacteric ethylene and exhibited a dramatic down-regulation of the key ripening regulators RIN, CNR, NOR and TAGL1. Ethylene treatment failed to reverse the non-ripening phenotype and the expression of ethylene signaling and biosynthesis genes was strongly altered in SlARF2 down-regulated fruits. Although both SlARF proteins are transcriptional repressors the data indicate they work as positive regulators of tomato fruit ripening. Altogether, the study defines SlARF2 as a new component of the regulatory network controlling the ripening process in tomato.

  5. Interferon Regulatory Factor-1 Mediates Alveolar Macrophage Pyroptosis During LPS-Induced Acute Lung Injury in Mice

    PubMed Central

    Wu, Dongdong; Pan, Pinhua; Su, Xiaoli; Zhang, Lemeng; Qin, Qingwu; Tan, Hongyi; Huang, Li; Li, Yuanyuan

    2016-01-01

    ABSTRACT Previously, we demonstrated that pyroptosis in alveolar macrophages (AMs) plays an essential role in lipopolysaccharide (LPS)-induced acute lung injury. However, the underlying mechanism remains largely unclear. Here, we show that the absence of interferon regulatory factor 1 (IRF-1) in genetic knock-out mice strongly abrogates pyroptosis in AMs and alleviates the LPS-induced lung injury and systemic inflammation. Our study demonstrates that IRF-1 contributes to caspase-1 activation and apoptosis-associated speck-like protein containing a caspase activation and recruitment domain pyroptosome formation in AMs and leads to downstream inflammatory cytokine release, including that of IL-1β, IL-18, and HMGB1. The nuclear translocation of IRF-1 is linked to the presence of toll-like receptor 4 (TLR4). Our findings suggest that pyroptosis and the downstream inflammatory response in AMs induced by LPS is a process that is dependent on TLR4-mediated up-regulation of IRF-1. In summary, IRF-1 plays a key role in controlling caspase-1-dependent pyroptosis and inflammation. PMID:26939040

  6. Vpu Mediates Depletion of Interferon Regulatory Factor 3 during HIV Infection by a Lysosome-Dependent Mechanism

    PubMed Central

    Doehle, Brian P.; Chang, Kristina; Rustagi, Arjun; McNevin, John; McElrath, M. Juliana

    2012-01-01

    HIV has evolved sophisticated mechanisms to avoid restriction by intracellular innate immune defenses that otherwise serve to control acute viral infection and virus dissemination. Innate defenses are triggered when pattern recognition receptor (PRR) proteins of the host cell engage pathogen-associated molecule patterns (PAMPs) present in viral products. Interferon regulatory factor 3 (IRF3) plays a central role in PRR signaling of innate immunity to drive the expression of type I interferon (IFN) and interferon-stimulated genes (ISGs), including a variety of HIV restriction factors, that serve to limit viral replication directly and/or program adaptive immunity. Productive infection of T cells by HIV is dependent upon the targeted proteolysis of IRF3 that occurs through a virus-directed mechanism that results in suppression of innate immune defenses. However, the mechanisms by which HIV controls innate immune signaling and IRF3 function are not defined. Here, we examined the innate immune response induced by HIV strains identified through their differential control of PRR signaling. We identified viruses that, unlike typical circulating HIV strains, lack the ability to degrade IRF3. Our studies show that IRF3 regulation maps specifically to the HIV accessory protein Vpu. We define a molecular interaction between Vpu and IRF3 that redirects IRF3 to the endolysosome for proteolytic degradation, thus allowing HIV to avoid the innate antiviral immune response. Our studies reveal that Vpu is an important IRF3 regulator that supports acute HIV infection through innate immune suppression. These observations define the Vpu-IRF3 interface as a novel target for therapeutic strategies aimed at enhancing the immune response to HIV. PMID:22593165

  7. Vpu mediates depletion of interferon regulatory factor 3 during HIV infection by a lysosome-dependent mechanism.

    PubMed

    Doehle, Brian P; Chang, Kristina; Rustagi, Arjun; McNevin, John; McElrath, M Juliana; Gale, Michael

    2012-08-01

    HIV has evolved sophisticated mechanisms to avoid restriction by intracellular innate immune defenses that otherwise serve to control acute viral infection and virus dissemination. Innate defenses are triggered when pattern recognition receptor (PRR) proteins of the host cell engage pathogen-associated molecule patterns (PAMPs) present in viral products. Interferon regulatory factor 3 (IRF3) plays a central role in PRR signaling of innate immunity to drive the expression of type I interferon (IFN) and interferon-stimulated genes (ISGs), including a variety of HIV restriction factors, that serve to limit viral replication directly and/or program adaptive immunity. Productive infection of T cells by HIV is dependent upon the targeted proteolysis of IRF3 that occurs through a virus-directed mechanism that results in suppression of innate immune defenses. However, the mechanisms by which HIV controls innate immune signaling and IRF3 function are not defined. Here, we examined the innate immune response induced by HIV strains identified through their differential control of PRR signaling. We identified viruses that, unlike typical circulating HIV strains, lack the ability to degrade IRF3. Our studies show that IRF3 regulation maps specifically to the HIV accessory protein Vpu. We define a molecular interaction between Vpu and IRF3 that redirects IRF3 to the endolysosome for proteolytic degradation, thus allowing HIV to avoid the innate antiviral immune response. Our studies reveal that Vpu is an important IRF3 regulator that supports acute HIV infection through innate immune suppression. These observations define the Vpu-IRF3 interface as a novel target for therapeutic strategies aimed at enhancing the immune response to HIV.

  8. Interferon Regulatory Factor 8-Deficiency Determines Massive Neutrophil Recruitment but T Cell Defect in Fast Growing Granulomas during Tuberculosis

    PubMed Central

    Sali, Michela; Anfossi, Antonio Giovanni; Abalsamo, Laura; Palucci, Ivana; Mattei, Fabrizio; Sanchez, Massimo; Giagu, Anna; Antuofermo, Elisabetta; Fadda, Giovanni; Belardelli, Filippo; Delogu, Giovanni; Gabriele, Lucia

    2013-01-01

    Following Mycobacterium tuberculosis (Mtb) infection, immune cell recruitment in lungs is pivotal in establishing protective immunity through granuloma formation and neogenesis of lymphoid structures (LS). Interferon regulatory factor-8 (IRF-8) plays an important role in host defense against Mtb, although the mechanisms driving anti-mycobacterial immunity remain unclear. In this study, IRF-8 deficient mice (IRF-8−/−) were aerogenously infected with a low-dose Mtb Erdman virulent strain and the course of infection was compared with that induced in wild-type (WT-B6) counterparts. Tuberculosis (TB) progression was examined in both groups using pathological, microbiological and immunological parameters. Following Mtb exposure, the bacterial load in lungs and spleens progressed comparably in the two groups for two weeks, after which IRF-8−/− mice developed a fatal acute TB whereas in WT-B6 the disease reached a chronic stage. In lungs of IRF-8−/−, uncontrolled growth of pulmonary granulomas and impaired development of LS were observed, associated with unbalanced homeostatic chemokines, progressive loss of infiltrating T lymphocytes and massive prevalence of neutrophils at late infection stages. Our data define IRF-8 as an essential factor for the maintenance of proper immune cell recruitment in granulomas and LS required to restrain Mtb infection. Moreover, IRF-8−/− mice, relying on a common human and mouse genetic mutation linked to susceptibility/severity of mycobacterial diseases, represent a valuable model of acute TB for comparative studies with chronically-infected congenic WT-B6 for dissecting protective and pathological immune reactions. PMID:23717393

  9. Interferon regulatory factor 8-deficiency determines massive neutrophil recruitment but T cell defect in fast growing granulomas during tuberculosis.

    PubMed

    Rocca, Stefano; Schiavoni, Giovanna; Sali, Michela; Anfossi, Antonio Giovanni; Abalsamo, Laura; Palucci, Ivana; Mattei, Fabrizio; Sanchez, Massimo; Giagu, Anna; Antuofermo, Elisabetta; Fadda, Giovanni; Belardelli, Filippo; Delogu, Giovanni; Gabriele, Lucia

    2013-01-01

    Following Mycobacterium tuberculosis (Mtb) infection, immune cell recruitment in lungs is pivotal in establishing protective immunity through granuloma formation and neogenesis of lymphoid structures (LS). Interferon regulatory factor-8 (IRF-8) plays an important role in host defense against Mtb, although the mechanisms driving anti-mycobacterial immunity remain unclear. In this study, IRF-8 deficient mice (IRF-8⁻/⁻) were aerogenously infected with a low-dose Mtb Erdman virulent strain and the course of infection was compared with that induced in wild-type (WT-B6) counterparts. Tuberculosis (TB) progression was examined in both groups using pathological, microbiological and immunological parameters. Following Mtb exposure, the bacterial load in lungs and spleens progressed comparably in the two groups for two weeks, after which IRF-8⁻/⁻ mice developed a fatal acute TB whereas in WT-B6 the disease reached a chronic stage. In lungs of IRF-8⁻/⁻, uncontrolled growth of pulmonary granulomas and impaired development of LS were observed, associated with unbalanced homeostatic chemokines, progressive loss of infiltrating T lymphocytes and massive prevalence of neutrophils at late infection stages. Our data define IRF-8 as an essential factor for the maintenance of proper immune cell recruitment in granulomas and LS required to restrain Mtb infection. Moreover, IRF-8⁻/⁻ mice, relying on a common human and mouse genetic mutation linked to susceptibility/severity of mycobacterial diseases, represent a valuable model of acute TB for comparative studies with chronically-infected congenic WT-B6 for dissecting protective and pathological immune reactions. PMID:23717393

  10. Signals and myogenic regulatory factors restrict pax3 and pax7 expression to dermomyotome-like tissue in zebrafish

    PubMed Central

    Hammond, Christina L.; Hinits, Yaniv; Osborn, Daniel P.S.; Minchin, James E.N.; Tettamanti, Gianluca; Hughes, Simon M.

    2014-01-01

    Pax3/7 paired homeodomain transcription factors are important markers of muscle stem cells. Pax3 is required upstream of myod for lateral dermomyotomal cells in the amniote somite to form particular muscle cells. Later Pax3/7-dependent cells generate satellite cells and most body muscle. Here we analyse early myogenesis from, and regulation of, a population of Pax3-expressing dermomyotome-like cells in the zebrafish. Zebrafish pax3 is widely expressed in the lateral somite and, along with pax7, becomes restricted anteriorly and then to the external cells on the lateral somite surface. Midline-derived Hedgehog signals appear to act directly on lateral somite cells to repress Pax3/7. Both Hedgehog and Fgf8, signals that induce muscle formation within the somite, suppress Pax3/7 and promote expression of myogenic regulatory factors (MRFs) myf5 and myod in specific muscle precursor cell populations. Loss of MRF function leads to loss of myogenesis by specific populations of muscle fibres, with parallel up-regulation of Pax3/7. Myod is required for lateral fast muscle differentiation from pax3-expressing cells. In contrast, either Myf5 or Myod is sufficient to promote slow muscle formation from adaxial cells. Thus, myogenic signals act to drive somite cells to a myogenic fate through up-regulation of distinct combinations of MRFs. Our data show that the relationship between Pax3/7 genes and myogenesis is evolutionarily ancient, but that changes in the MRF targets for particular signals contribute to myogenic differences between species. PMID:17094960

  11. 75 FR 18213 - Determination of Regulatory Review Period for Purposes of Patent Extension; MOZOBIL

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-04-09

    ... product, animal drug product, medical device, food additive, or color additive) was subject to regulatory...- colony stimulating factor to mobilize hematopoietic stem cells to the peripheral blood for collection...

  12. SND1 transcription factor-directed quantitative functional hierarchical genetic regulatory network in wood formation in Populus trichocarpa.

    PubMed

    Lin, Ying-Chung; Li, Wei; Sun, Ying-Hsuan; Kumari, Sapna; Wei, Hairong; Li, Quanzi; Tunlaya-Anukit, Sermsawat; Sederoff, Ronald R; Chiang, Vincent L

    2013-11-01

    Wood is an essential renewable raw material for industrial products and energy. However, knowledge of the genetic regulation of wood formation is limited. We developed a genome-wide high-throughput system for the discovery and validation of specific transcription factor (TF)-directed hierarchical gene regulatory networks (hGRNs) in wood formation. This system depends on a new robust procedure for isolation and transfection of Populus trichocarpa stem differentiating xylem protoplasts. We overexpressed Secondary Wall-Associated NAC Domain 1s (Ptr-SND1-B1), a TF gene affecting wood formation, in these protoplasts and identified differentially expressed genes by RNA sequencing. Direct Ptr-SND1-B1-DNA interactions were then inferred by integration of time-course RNA sequencing data and top-down Graphical Gaussian Modeling-based algorithms. These Ptr-SND1-B1-DNA interactions were verified to function in differentiating xylem by anti-PtrSND1-B1 antibody-based chromatin immunoprecipitation (97% accuracy) and in stable transgenic P. trichocarpa (90% accuracy). In this way, we established a Ptr-SND1-B1-directed quantitative hGRN involving 76 direct targets, including eight TF and 61 enzyme-coding genes previously unidentified as targets. The network can be extended to the third layer from the second-layer TFs by computation or by overexpression of a second-layer TF to identify a new group of direct targets (third layer). This approach would allow the sequential establishment, one two-layered hGRN at a time, of all layers involved in a more comprehensive hGRN. Our approach may be particularly useful to study hGRNs in complex processes in plant species resistant to stable genetic transformation and where mutants are unavailable.

  13. Regulatory domains of the A-Myb transcription factor and its interaction with the CBP/p300 adaptor molecules.

    PubMed Central

    Facchinetti, V; Loffarelli, L; Schreek, S; Oelgeschläger, M; Lüscher, B; Introna, M; Golay, J

    1997-01-01

    The A-Myb transcription factor belongs to the Myb family of oncoproteins and is likely to be involved in the regulation of proliferation and/or differentiation of normal B cells and Burkitt's lymphoma cells. To characterize in detail the domains of A-Myb that regulate its function, we have generated a series of deletion mutants and have investigated their trans-activation potential as well as their DNA-binding activity. Our results have allowed us to delineate the trans-activation domain as well as two separate regulatory regions. The boundaries of the trans-activation domain (amino acid residues 218-319) are centred on a sequence rich in charged amino acids (residues 259-281). A region (residues 320-482) localized immediately downstream of the trans-activation domain and containing a newly identified conserved stretch of 48 residues markedly inhibits specific DNA binding. Finally the last 110 residues of A-Myb (residues 643-752), which include a sequence conserved in all mammalian myb genes (region III), negatively regulate the maximal trans-activation potential of A-Myb. We have also investigated the functional interaction between A-Myb and the nuclear adaptor molecule CBP [cAMP response element-binding protein (CREB)-binding protein]. We demonstrate that CBP synergizes with A-Myb in a dose-dependent fashion, and that this co-operative effect can be inhibited by E1A and can also be observed with the CBP homologue p300. We show that this functional synergism requires the presence of the A-Myb charged sequence and that it involves physical interaction between A-Myb and the CREB-binding domain of CBP. PMID:9210395

  14. Conserved Gene Regulatory Function of the Carboxy-Terminal Domain of Dictyostelid C-Module-Binding Factor

    PubMed Central

    Schmith, Anika; Groth, Marco; Ratka, Josephine; Gatz, Sara; Spaller, Thomas; Siol, Oliver; Glöckner, Gernot

    2013-01-01

    C-module-binding factor A (CbfA) is a jumonji-type transcription regulator that is important for maintaining the expression and mobility of the retrotransposable element TRE5-A in the social amoeba Dictyostelium discoideum. CbfA-deficient cells have lost TRE5-A retrotransposition, are impaired in the ability to feed on bacteria, and do not enter multicellular development because of a block in cell aggregation. In this study, we performed Illumina RNA-seq of growing CbfA mutant cells to obtain a list of CbfA-regulated genes. We demonstrate that the carboxy-terminal domain of CbfA alone is sufficient to mediate most CbfA-dependent gene expression. The carboxy-terminal domain of CbfA from the distantly related social amoeba Polysphondylium pallidum restored the expression of CbfA-dependent genes in the D. discoideum CbfA mutant, indicating a deep conservation in the gene regulatory function of this domain in the dictyostelid clade. The CbfA-like protein CbfB displays ∼25% sequence identity with CbfA in the amino-terminal region, which contains a JmjC domain and two zinc finger regions and is thought to mediate chromatin-remodeling activity. In contrast to CbfA proteins, where the carboxy-terminal domains are strictly conserved in all dictyostelids, CbfB proteins have completely unrelated carboxy-terminal domains. Outside the dictyostelid clade, CbfA-like proteins with the CbfA-archetypical JmjC/zinc finger arrangement and individual carboxy-terminal domains are prominent in filamentous fungi but are not found in yeasts, plants, and metazoans. Our data suggest that two functional regions of the CbfA-like proteins evolved at different rates to allow the occurrence of species-specific adaptation processes during genome evolution. PMID:23355006

  15. Drug Transporters and Na+/H+ Exchange Regulatory Factor PSD-95/Drosophila Discs Large/ZO-1 Proteins

    PubMed Central

    Walsh, Dustin R.; Nolin, Thomas D.

    2015-01-01

    Drug transporters govern the absorption, distribution, and elimination of pharmacologically active compounds. Members of the solute carrier and ATP binding-cassette drug transporter family mediate cellular drug uptake and efflux processes, thereby coordinating the vectorial movement of drugs across epithelial barriers. To exert their physiologic and pharmacological function in polarized epithelia, drug transporters must be targeted and stabilized to appropriate regions of the cell membrane (i.e., apical versus basolateral). Despite the critical importance of drug transporter membrane targeting, the mechanisms that underlie these processes are largely unknown. Several clinically significant drug transporters possess a recognition sequence that binds to PSD-95/Drosophila discs large/ZO-1 (PDZ) proteins. PDZ proteins, such as the Na+/H+ exchanger regulatory factor (NHERF) family, act to stabilize and organize membrane targeting of multiple transmembrane proteins, including many clinically relevant drug transporters. These PDZ proteins are normally abundant at apical membranes, where they tether membrane-delimited transporters. NHERF expression is particularly high at the apical membrane in polarized tissue such as intestinal, hepatic, and renal epithelia, tissues important to drug disposition. Several recent studies have highlighted NHERF proteins as determinants of drug transporter function secondary to their role in controlling membrane abundance and localization. Mounting evidence strongly suggests that NHERF proteins may have clinically significant roles in pharmacokinetics and pharmacodynamics of several pharmacologically active compounds and may affect drug action in cancer and chronic kidney disease. For these reasons, NHERF proteins represent a novel class of post-translational mediators of drug transport and novel targets for new drug development. PMID:26092975

  16. Postnatal developmental expression of the PDZ scaffolds Na+-H+ exchanger regulatory factors 1 and 2 in the rat cochlea

    PubMed Central

    Kanjhan, Refik; Hryciw, Deanne H.; Bellingham, Mark C.; Poronnik, Philip; Yun, C. Chris

    2006-01-01

    Sensory transduction in the mammalian cochlea requires the maintenance of specialized fluid compartments with distinct ionic compositions. This is achieved by the concerted action of diverse ion channels and transporters, some of which can interact with the PDZ scaffolds, Na+-H+ exchanger regulatory factors 1 and 2 (NHERF-1, NHERF-2). Here, we report that NHERF-1 and NHERF-2 are widely expressed in the rat cochlea, and that their expression is developmentally regulated. Reverse transcription/polymerase chain reaction (RT-PCR) and Western blotting initially confirmed the RNA and protein expression of NHERFs. We then performed immunohistochemistry on cochlea during various stages of postnatal development. Prior to the onset of hearing (P8), NHERF-1 immunolabeling was prominently polarized to the apical membrane of cells lining the endolymphatic compartment, including the stereocilia and cuticular plates of the inner and outer hair cells, marginal cells of the stria vascularis, Reissner’s epithelia, and tectorial membrane. With maturation (P21, P70), NHERF-1 immunolabeling was reduced in the above structures, whereas labeling increased in the apical membrane of the interdental cells of the spiral limbus and the inner and outer sulcus cells, Hensen’s cells, the inner and outer pillar cells, Deiters cells, the inner border cells, spiral ligament fibrocytes, and spiral ganglion neurons (particularly type II). NHERF-1 expression in strial basal and intermediate cells was persistent. NHERF-2 immunolabeling was similar to that for NHERF-1 during postnatal development, with the exception of expression in the synaptic regions beneath the outer hair cells. NHERF-1 and NHERF-2 co-localized with glial fibrillary acidic protein and vimentin in glia. The cochlear localization of NHERF scaffolds suggests that they play important roles in the developmental regulation of ion transport, homeostasis, and auditory neurotransmission. PMID:16160858

  17. Infliximab therapy balances regulatory T cells, tumour necrosis factor receptor 2 (TNFR2) expression and soluble TNFR2 in sarcoidosis.

    PubMed

    Verwoerd, A; Hijdra, D; Vorselaars, A D M; Crommelin, H A; van Moorsel, C H M; Grutters, J C; Claessen, A M E

    2016-08-01

    Sarcoidosis is a systemic granulomatous disease of unknown aetiology that most commonly affects the lungs. Although elevated levels of regulatory T cells (Tregs ) have been reported, the extent to which they play a role in sarcoidosis pathogenesis remains unclear. Tumour necrosis factor (TNF) is thought to be one of the driving forces behind granuloma formation, illustrated by the efficacy of infliximab in severe sarcoidosis. Tregs express TNF receptor 2 (TNFR2) highly. Here, we examined the influence of infliximab therapy on Tregs and (soluble) TNFR2 levels in sarcoidosis, and correlated these with response to therapy. We observed that relative frequencies of Tregs were significantly higher in patients (n = 54) compared to healthy controls (n = 26; median 6·73 versus 4·36%; P < 0·001) and decreased following therapy (4·95; P < 0·001). Baseline TNFR2 expression on Tregs was increased significantly in patients versus controls (99·4 versus 96·2%; P = 0·031), and also in responders to therapy versus non-responders (99·6 versus 97·3%; P = 0·012). Furthermore, baseline soluble TNFR2 (sTNFR2) was higher in responders than in non-responders (mean 174 versus 107 pg/ml; P = 0·015). After treatment, responders showed a significant reduction in sTNFR2 levels in peripheral blood (-44·7 pg/ml; P < 0·001), in contrast to non-responders (+3·59 pg/ml). Our results demonstrated that Treg frequencies and TNFR2 expression on Tregs are increased in sarcoidosis, followed by a decline during infliximab therapy, suggesting a pathophysiological role of this T cell subset. Interestingly, sTNFR2 levels at baseline differed significantly between responders and non-responders, making it a potential marker in predicting which patients might benefit from infliximab.

  18. Molecular identification and functional characterisation of the interferon regulatory factor 1 in the blunt snout bream (Megalobrama amblycephala).

    PubMed

    Zhan, Fan-Bin; Liu, Han; Lai, Rui-Fang; Jakovlić, Ivan; Wang, Wen-Bin; Wang, Wei-Min

    2016-07-01

    Interferon regulatory factors (IRFs) play a key role in mediating the host response against pathogen infection and other important biological processes. This is the first report of an IRF family member in blunt snout bream Megalobrama amblycephala. The complete cDNA of M. amblycephala (Ma) IRF1 gene has 1422 nucleotides (nt.), with an open reading frame of 858 nt, encoding a polypeptide of 285 amino acids. The putative MaIRF1 polypeptide shared significant structural homology with known IRF1 homologs: a conserved IRF domain was found at the N-terminal and an IRF association domain 2 at the C-terminal. Phylogenetic analysis showed that MaIRF1 amino acid sequence clustered with other teleost IRF1s, with a grass carp ortholog exhibiting the highest similarity. MaIRF1 mRNA expression patterns were studied using quantitative real-time PCR in healthy fish tissues and after a challenge with Aeromonas hydrophila bacterium. It was constitutively expressed in all examined tissues: the highest in blood, the lowest in muscle. The expression after A. hydrophila challenge was up-regulated in liver, spleen and kidney, but down-regulated in intestine and gills. At the protein level, similar expression patterns were observed in liver and gills. Patterns differed in intestine (up-regulation), spleen (down-regulation) and kidney (expression mostly unchanged). This study indicates that MaIRF1 gene plays an important role in the blunt snout bream immune system, hence providing an important base for further studies. PMID:27150048

  19. LGP2 Downregulates Interferon Production during Infection with Seasonal Human Influenza A Viruses That Activate Interferon Regulatory Factor 3

    PubMed Central

    Malur, Meghana; Gale, Michael

    2012-01-01

    LGP2, a member of the RIG-I-like receptor family, lacks the amino-terminal caspase activation recruitment domains (CARDs) required for initiating the activation of interferon regulatory factor 3 (IRF3) and interferon (IFN) transcription. The role of LGP2 in virus infection is controversial, and the only LGP2 experiments previously carried out with mammalian influenza A viruses employed an attenuated, mouse-adapted H1N1 A/PR/8/34 (PR8) virus that does not encode the NS1 protein. Here we determine whether LGP2 has a role during infection with wild-type, nonattenuated influenza A viruses that have circulated in the human population, specifically two types of seasonal influenza A viruses: (i) H3N2 and H1N1 viruses that activate IRF3 and IFN transcription and (ii) recent H1N1 viruses that block these two activations. In human cells infected with an H3N2 virus that activates IRF3, overexpression of LGP2 or its repressor domain decreased STAT1 activation and IFN-β transcription approximately 10-fold. Overexpression of LGP2 also caused a 10-fold decrease of STAT1 activation during infection with other seasonal influenza A viruses that activate IRF3. Using LGP2+/+ and LGP2−/− mouse cells, we show that endogenous LGP2 decreased IFN production during H3N2 virus infection 3- to 4-fold. In contrast, in both mouse and human cells infected with H1N1 viruses that do not activate IRF3, LGP2 had no detectable role. These results demonstrate that LGP2 downregulates IFN production during infection by seasonal influenza A viruses that activate IRF3 and IFN transcription. It is intriguing that LGP2, a host protein induced during influenza A virus infection, downregulates the host antiviral IFN response. PMID:22837208

  20. Inhibition of mRNA export and dimerization of interferon regulatory factor 3 by Theiler's virus leader protein.

    PubMed

    Ricour, Céline; Delhaye, Sophie; Hato, Stanleyson V; Olenyik, Tamara D; Michel, Bénédicte; van Kuppeveld, Frank J M; Gustin, Kurt E; Michiels, Thomas

    2009-01-01

    Theiler's murine encephalomyelitis virus (TMEV or Theiler's virus) is a neurotropic picornavirus that can persist lifelong in the central nervous system of infected mice, causing a chronic inflammatory demyelinating disease. The leader (L) protein of the virus is an important determinant of viral persistence and has been shown to inhibit transcription of type I interferon (IFN) genes and to cause nucleocytoplasmic redistribution of host proteins. In this study, it was shown that expression of the L protein shuts off synthesis of the reporter proteins green fluorescent protein and firefly luciferase, suggesting that it induces a global shut-off of host protein expression. The L protein did not inhibit transcription or translation of the reporter genes, but blocked cellular mRNA export from the nucleus. This activity correlated with the phosphorylation of nucleoporin 98 (Nup98), an essential component of the nuclear pore complex. In contrast, the data confirmed that the L protein inhibited IFN expression at the transcriptional level, and showed that transcription of other chemokine or cytokine genes was affected by the L protein. This transcriptional inhibition correlated with inhibition of interferon regulatory factor 3 (IRF-3) dimerization. Whether inhibition of IRF-3 dimerization and dysfunction of the nuclear pore complex are related phenomena remains an open question. In vivo, IFN antagonism appears to be an important role of the L protein early in infection, as a virus bearing a mutation in the zinc finger of the L protein replicated as efficiently as the wild-type virus in type I IFN receptor-deficient mice, but had impaired fitness in IFN-competent mice.

  1. MEK inhibition prevents tumour-shed transforming growth factor-β-induced T-regulatory cell augmentation in tumour milieu

    PubMed Central

    Hossain, Dewan M S; Panda, Abir K; Chakrabarty, Sreeparna; Bhattacharjee, Pushpak; Kajal, Kirti; Mohanty, Suchismita; Sarkar, Irene; Sarkar, Diptendra K; Kar, Santosh K; Sa, Gaurisankar

    2015-01-01

    Tumour progression is associated with immune-suppressive conditions that facilitate the escape of tumour cells from the regimen of immune cells, subsequently paralysing the host defence mechanisms. Induction of CD4+ CD25+ FoxP3+ T regulatory (Treg) cells has been implicated in the tumour immune escape mechanism, although the novel anti-cancer treatment strategies targeting Treg cells remain unknown. The focus of this study is to define the interaction between tumour and immune system, i.e. how immune tolerance starts and gradually leads to the induction of adaptive Treg cells in the tumour microenvironment. Our study identified hyperactivated mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) -signalling as a potential target for reversing Treg cell augmentation in breast cancer patients. In more mechanistic detail, pharmacological inhibitors of MEK/ERK signalling inhibited transforming growth factor-β (TGF-β) production in tumour cells that essentially blocked TGF-β-SMAD3/SMAD4-mediated induction of CD25/interleukin-2 receptor α on CD4+ T-cell surface. As a result high-affinity binding of interleukin-2 on those cells was prohibited, causing lack of Janus kinase 1 (JAK1)/JAK3-mediated signal transducer and activator of transcription 3 (STAT3)/STAT5 activation required for FoxP3 expression. Finally, for a more radical approach towards a safe MEK inhibitor, we validate the potential of multi-kinase inhibitor curcumin, especially the nano-curcumin made out of pure curcumin with greater bioavailability; in repealing tumour-shed TGF-β-induced Treg cell augmentation. PMID:25284464

  2. LGP2 downregulates interferon production during infection with seasonal human influenza A viruses that activate interferon regulatory factor 3.

    PubMed

    Malur, Meghana; Gale, Michael; Krug, Robert M

    2012-10-01

    LGP2, a member of the RIG-I-like receptor family, lacks the amino-terminal caspase activation recruitment domains (CARDs) required for initiating the activation of interferon regulatory factor 3 (IRF3) and interferon (IFN) transcription. The role of LGP2 in virus infection is controversial, and the only LGP2 experiments previously carried out with mammalian influenza A viruses employed an attenuated, mouse-adapted H1N1 A/PR/8/34 (PR8) virus that does not encode the NS1 protein. Here we determine whether LGP2 has a role during infection with wild-type, nonattenuated influenza A viruses that have circulated in the human population, specifically two types of seasonal influenza A viruses: (i) H3N2 and H1N1 viruses that activate IRF3 and IFN transcription and (ii) recent H1N1 viruses that block these two activations. In human cells infected with an H3N2 virus that activates IRF3, overexpression of LGP2 or its repressor domain decreased STAT1 activation and IFN-β transcription approximately 10-fold. Overexpression of LGP2 also caused a 10-fold decrease of STAT1 activation during infection with other seasonal influenza A viruses that activate IRF3. Using LGP2(+/+) and LGP2(-/-) mouse cells, we show that endogenous LGP2 decreased IFN production during H3N2 virus infection 3- to 4-fold. In contrast, in both mouse and human cells infected with H1N1 viruses that do not activate IRF3, LGP2 had no detectable role. These results demonstrate that LGP2 downregulates IFN production during infection by seasonal influenza A viruses that activate IRF3 and IFN transcription. It is intriguing that LGP2, a host protein induced during influenza A virus infection, downregulates the host antiviral IFN response.

  3. Regulatory roles for Tiam1, a guanine nucleotide exchange factor for Rac1, in glucose-stimulated insulin secretion in pancreatic beta-cells.

    PubMed

    Veluthakal, Rajakrishnan; Madathilparambil, Suresh Vasu; McDonald, Phillip; Olson, Lawrence Karl; Kowluru, Anjaneyulu

    2009-01-01

    Using various biochemical, pharmacological and molecular biological approaches, we have recently reported regulatory roles for Rac1, a small G-protein, in glucose-stimulated insulin secretion (GSIS). However, little is understood with respect to localization of, and regulation by, specific regulatory factors of Rac1 in GSIS. Herein, we investigated regulatory roles for Tiam1, a specific nucleotide exchange factor (GEF) for Rac1, in GSIS in pancreatic beta-cells. Western blot analysis indicated that Tiam1 is predominantly cytosolic in distribution. NSC23766, a specific inhibitor of Tiam1-mediated activation of Rac1, markedly attenuated glucose-induced, but not KCl-induced insulin secretion in INS 832/13 cells and normal rat islets. Further, NSC23766 significantly reduced glucose-induced activation (i.e. GTP-bound form) and membrane association of Rac1 in INS 832/13 cells and rat islets. Moreover, siRNA-mediated knock-down of Tiam1 markedly inhibited glucose-induced membrane trafficking and activation of Rac1 in INS 832/13 cells. Interestingly, however, in contrast to the inhibitory effects of NSC23766, Tiam1 gene depletion potentiated GSIS in these cells; such a potentiation of GSIS was sensitive to extracellular calcium. Together, our studies present the first evidence for a regulatory role for Tiam1/Rac1-sensitive signaling step in GSIS. They also provide evidence for the existence of a potential Rac1/Tiam1-independent, but calcium-sensitive component for GSIS in these cells.

  4. Homeobox genes and melatonin synthesis: regulatory roles of the cone-rod homeobox transcription factor in the rodent pineal gland.

    PubMed

    Rohde, Kristian; Møller, Morten; Rath, Martin Fredensborg

    2014-01-01

    Nocturnal synthesis of melatonin in the pineal gland is controlled by a circadian rhythm in arylalkylamine N-acetyltransferase (AANAT) enzyme activity. In the rodent, Aanat gene expression displays a marked circadian rhythm; release of norepinephrine in the gland at night causes a cAMP-based induction of Aanat transcription. However, additional transcriptional control mechanisms exist. Homeobox genes, which are generally known to encode transcription factors controlling developmental processes, are also expressed in the mature rodent pineal gland. Among these, the cone-rod homeobox (CRX) transcription factor is believed to control pineal-specific Aanat expression. Based on recent advances in our understanding of Crx in the rodent pineal gland, we here suggest that homeobox genes play a role in adult pineal physiology both by ensuring pineal-specific Aanat expression and by facilitating cAMP response element-based circadian melatonin production.

  5. Homeobox Genes and Melatonin Synthesis: Regulatory Roles of the Cone-Rod Homeobox Transcription Factor in the Rodent Pineal Gland

    PubMed Central

    Rath, Martin Fredensborg

    2014-01-01

    Nocturnal synthesis of melatonin in the pineal gland is controlled by a circadian rhythm in arylalkylamine N-acetyltransferase (AANAT) enzyme activity. In the rodent, Aanat gene expression displays a marked circadian rhythm; release of norepinephrine in the gland at night causes a cAMP-based induction of Aanat transcription. However, additional transcriptional control mechanisms exist. Homeobox genes, which are generally known to encode transcription factors controlling developmental processes, are also expressed in the mature rodent pineal gland. Among these, the cone-rod homeobox (CRX) transcription factor is believed to control pineal-specific Aanat expression. Based on recent advances in our understanding of Crx in the rodent pineal gland, we here suggest that homeobox genes play a role in adult pineal physiology both by ensuring pineal-specific Aanat expression and by facilitating cAMP response element-based circadian melatonin production. PMID:24877149

  6. Molecular architecture of the regulatory Locus sae of Staphylococcus aureus and its impact on expression of virulence factors.

    PubMed

    Steinhuber, Andrea; Goerke, Christiane; Bayer, Manfred G; Döring, Gerd; Wolz, Christiane

    2003-11-01

    We characterized the sae operon, a global regulator for virulence gene expression in Staphylococcus aureus. A Tn917 sae mutant was obtained by screening a Tn917 library of the agr mutant ISP479Mu for clones with altered hemolytic activity. Sequence analysis of the sae operon revealed two additional open reading frames (ORFs) (ORF3 and ORF4) upstream of the two-component regulatory genes saeR and saeS. Four overlapping sae-specific transcripts (T1 to T4) were detected by Northern blot analysis, and the transcriptional initiation points were mapped by primer extension analysis. The T1, T2, and T3 mRNAs are probably terminated at the same stem-loop sequence downstream of saeS. The T1 message (3.1 kb) initiates upstream of ORF4, T2 (2.4 kb) initiates upstream of ORF3, and T3 (2.0 kb) initiates in front of saeR. T4 (0.7 kb) represents a monocistronic mRNA encompassing ORF4 only. sae-specific transcripts were detectable in all of the 40 different clinical S. aureus isolates investigated. Transcript levels were at maximum during the post-exponential growth phase. The sae mutant showed a significantly reduced rate of invasion of human endothelial cells, consistent with diminished transcription and expression of fnbA. The expression of type 5 capsular polysaccharide is activated in the sae mutant of strain Newman, as shown by immunofluorescence and promoter-reporter fusion experiments. In summary, the sae operon constitutes a four-component regulator system which acts on virulence gene expression in S. aureus.

  7. Enhancing Interferon Regulatory Factor 7 Mediated Antiviral Responses and Decreasing Nuclear Factor Kappa B Expression Limit HIV-1 Replication in Cervical Tissues

    PubMed Central

    Rollenhage, Christiane; Macura, Sherrill L.; Lathrop, Melissa J.; Mackenzie, Todd A.; Doncel, Gustavo F.; Asin, Susana N.

    2015-01-01

    Establishment of a productive HIV-1 infection in the female reproductive tract likely depends on the balance between anti-viral and pro-inflammatory responses leading to activation and proliferation of HIV target cells. Immune modulators that boost anti-viral and depress pro-inflammatory immune responses may decrease HIV-1 infection or replication. Polyinosinic:polycytidylic [Poly (I:C)] has been reported to down-regulate HIV-1 replication in immune cell subsets and lymphoid tissues, yet the scope and mechanisms of poly (I:C) regulation of HIV-1 replication in the cervicovaginal mucosa, the main portal of viral entry in women remain unknown. Using a relevant, underexplored ex vivo cervical tissue model, we demonstrated that poly (I:C) enhanced Interferon Regulatory Factor (IRF)7 mediated antiviral responses and decreased tissue Nuclear Factor Kappa B (NFκB) RNA expression. This pattern of cellular transcription factor expression correlated with decreased HIV-1 transcription and viral release. Reducing IRF7 expression up-regulated HIV-1 and NFκB transcription, providing proof of concept for the critical involvement of IRF7 in cervical tissues. By combining poly (I:C) with a suboptimal concentration of tenofovir, the leading anti-HIV prophylactic microbicide candidate, we demonstrated an earlier and greater decrease in HIV replication in poly (I:C)/tenofovir treated tissues compared with tissues treated with tenofovir alone, indicating overall improved efficacy. Poly (I:C) decreases HIV-1 replication by stimulating IRF7 mediated antiviral responses while reducing NFκB expression. Early during the infection, poly (I:C) improved the anti-HIV-1 activity of suboptimal concentrations of tenofovir likely to be present during periods of poor adherence i.e. inconsistent or inadequate drug use. Understanding interactions between anti-viral and pro-inflammatory immune responses in the genital mucosa will provide crucial insights for the identification of targets that can be

  8. Regulatory roles of tumor necrosis factor-alpha and interleukin-1 beta in monocyte chemoattractant protein-1-mediated pulmonary granuloma formation in the rat.

    PubMed Central

    Flory, C. M.; Jones, M. L.; Miller, B. F.; Warren, J. S.

    1995-01-01

    Intravenous infusion of particulate yeast cell wall glucan into rats results in the synchronous development of angiocentric pulmonary granulomas that are composed almost entirely of monocytes and macrophages. Previous studies indicate that locally produced monocyte chemoattractant protein-1 (MCP-1) is required for full granuloma development. Because tumor necrosis factor-alpha (TNF-alpha) and interleukin 1 (IL-1) can induce MCP-1 production in a variety of cell types, we sought to determine their potential regulatory roles in this model. A single infusion of anti-TNF-alpha antibody at the time of glucan infusion (time 0) markedly reduced MCP-1 mRNA levels at 1 and 6 hours but not at later time points; there was no effect on granuloma size or number measured at 48 hours. When multiple infusions of anti-TNF-alpha antibody were administered over a 23-hour period (0 to 23 hours), MCP-1 mRNA was reduced through 24 hours, there was a significant reduction in peak bronchoalveolar lavage fluid MCP-1 activity at 48 hours, and there were marked reductions in granuloma size and number at 48 hours. Similar results were observed in animals that received infusions of anti-IL-1 beta. Infusion of anti-IL-1 beta at time 0 resulted in moderate reductions in MCP-1 mRNA at 1 and 6 hours and had no effect on granuloma size or number measured at 48 hours. When multiple infusions of anti-IL-1 beta were administered over a 23-hour period (0 to 23 hours), MCP-1 mRNA was reduced through 24 hours, there was a moderate reduction in peak bronchoalveolar lavage fluid MCP-1 activity at 48 hours, and there were marked reductions in granuloma size and number at 48 hours. A single infusion of anti-TNF-alpha and anti-IL-1 beta together at time 0 resulted in marked reductions in whole lung MCP-1 and mRNA at 1 and 6 hours, but not at 24 hours. Multiple combined infusions of anti-TNF-alpha and anti-IL-1 beta over a 23-hour period resulted in additive reductions in MCP-1 mRNA through 24 hours

  9. Effects of chondrogenic and osteogenic regulatory factors on composite constructs grown using human mesenchymal stem cells, silk scaffolds and bioreactors.

    PubMed

    Augst, Alexander; Marolt, Darja; Freed, Lisa E; Vepari, Charu; Meinel, Lorenz; Farley, Michelle; Fajardo, Robert; Patel, Nipun; Gray, Martha; Kaplan, David L; Vunjak-Novakovic, Gordana

    2008-08-01

    Human mesenchymal stem cells (hMSCs) isolated from bone marrow aspirates were cultured on silk scaffolds in rotating bioreactors for three weeks with either chondrogenic or osteogenic medium supplements to engineer cartilage- or bone-like tissue constructs. Osteochondral composites formed from these cartilage and bone constructs were cultured for an additional three weeks in culture medium that was supplemented with chondrogenic factors, supplemented with osteogenic factors or unsupplemented. Progression of cartilage and bone formation and the integration between the two regions were assessed by medical imaging (magnetic resonance imaging and micro-computerized tomography imaging), and by biochemical, histological and mechanical assays. During composite culture (three to six weeks), bone-like tissue formation progressed in all three media to a markedly larger extent than cartilage-like tissue formation. The integration of the constructs was most enhanced in composites cultured in chondrogenic medium. The results suggest that tissue composites with well-mineralized regions and substantially less developed cartilage regions can be generated in vitro by culturing hMSCs on silk scaffolds in bioreactors, that hMSCs have markedly higher capacity for producing engineered bone than engineered cartilage, and that chondrogenic factors play major roles at early stages of bone formation by hMSCs and in the integration of the two tissue constructs into a tissue composite. PMID:18230586

  10. Nuclear factor kappaB/p49 is a negative regulatory factor in nerve growth factor-induced choline acetyltransferase promoter activity in PC12 cells.

    PubMed

    Toliver-Kinsky, T; Wood, T; Perez-Polo, J R

    2000-12-01

    Anovel nuclear factor kappaB (NF-kappaB) binding site has been identified within the promoter region of the mouse gene encoding choline acetyltransferase (ChAT), the enzyme that synthesizes acetylcholine and has been implicated in the cognitive deficits associated with aging and Alzheimer's disease. This binding site, which is located within the nerve growth factor (NGF)-responsive enhancer element, was recognized by the NF-kappaB protein p49 but not p65 or p50. p49 from both basal forebrain and PC12 nuclear extracts interacted with this specific sequence in electrophoretic mobility shift assays. Mutation of the NF-kappaB site caused an increase in NGF-induced promoter activation, whereas overexpression of p49 in NGF-differentiated PC12 cells caused a decrease in endogenous ChAT enzyme activity and a decrease in promoter activity that was specifically mediated through this NF-kappaB binding site. Treatment of PC12 cells with NGF resulted in a drastic reduction in nuclear p49 binding to the ChAT NF-kappaB site after 24 h, but nuclear p49 levels were not altered, suggesting that late NGF-mediated events prevent binding of p49 to the ChAT promoter by an unknown mechanism other than nuclear translocation. Decreased ChAT expression and increased NF-kappaB activity in the brain are associated with aging and Alzheimer's disease. These data indicate that p49 is a negative regulator of ChAT expression and suggest a possible mechanism for aging-associated declines in cholinergic function.

  11. A combination of structural and cis-regulatory factors drives biochemical differences in Drosophila melanogaster malic enzyme.

    PubMed

    Rzezniczak, Teresa Z; Lum, Thomas E; Harniman, Robert; Merritt, Thomas J S

    2012-12-01

    The evolutionary significance of molecular variation is still contentious, with much current interest focusing on the relative contribution of structural changes in proteins versus regulatory variation in gene expression. We present a population genetic and biochemical study of molecular variation at the malic enzyme locus (Men) in Drosophila melanogaster. Two amino acid polymorphisms appear to affect substrate-binding kinetics, while only one appears to affect thermal stability. Interestingly, we find that enzyme activity differences previously assigned to one of the polymorphisms may, instead, be a function of linked regulatory differences. These results suggest that both regulatory and structural changes contribute to differences in protein function. Our examination of the Men coding sequences reveals no evidence for selection acting on the polymorphisms, but earlier work on this enzyme indicates that the biochemical variation observed has physiological repercussions and therefore could potentially be under natural selection.

  12. TCDD Induces the Hypoxia-Inducible Factor (HIF)-1α Regulatory Pathway in Human Trophoblastic JAR Cells

    PubMed Central

    Liao, Tien-Ling; Chen, Su-Chee; Tzeng, Chii-Reuy; Kao, Shu-Huei

    2014-01-01

    The exposure to dioxin can compromise pregnancy outcomes and increase the risk of preterm births. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) has been demonstrated to induce placental hypoxia at the end of pregnancy in a rat model, and hypoxia has been suggested to be the cause of abnormal trophoblast differentiation and placental insufficiency syndromes. In this study, we demonstrate that the non-hypoxic stimulation of human trophoblastic cells by TCDD strongly increased hypoxia inducible factor-1 alpha (HIF-1α) stabilization. TCDD exposure induced the generation of reactive oxygen species (ROS) and nitric oxide. TCDD-induced HIF-1α stabilization and Akt phosphorylation was inhibited by pretreatment with wortmannin (a phosphatidylinositol 3-kinase (PI3K) inhibitor) or N-acetylcysteine (a ROS scavenger). The augmented HIF-1α stabilization by TCDD occurred via the ROS-dependent activation of the PI3K/Akt pathway. Additionally, a significant increase in invasion and metallomatrix protease-9 activity was found in TCDD-treated cells. The gene expression of vascular endothelial growth factor and placental growth factor was induced upon TCDD stimulation, whereas the protein levels of peroxisome proliferator-activated receptor γ (PPARγ), PPARγ coactivator-1α, mitochondrial transcription factor, and uncoupling protein 2 were decreased. Our results indicate that an activated HIF-1α pathway, elicited oxidative stress, and induced metabolic stress contribute to TCDD-induced trophoblastic toxicity. These findings may provide molecular insight into the TCDD-induced impairment of trophoblast function and placental development. PMID:25272228

  13. Low edge safety factor operation and passive disruption avoidance in current carrying plasmas by the addition of stellarator rotational transform

    NASA Astrophysics Data System (ADS)

    Pandya, M. D.; ArchMiller, M. C.; Cianciosa, M. R.; Ennis, D. A.; Hanson, J. D.; Hartwell, G. J.; Hebert, J. D.; Herfindal, J. L.; Knowlton, S. F.; Ma, X.; Massidda, S.; Maurer, D. A.; Roberds, N. A.; Traverso, P. J.

    2015-11-01

    Low edge safety factor operation at a value less than two ( q (a )=1 /ι̷tot(a )<2 ) is routine on the Compact Toroidal Hybrid device with the addition of sufficient external rotational transform. Presently, the operational space of this current carrying stellarator extends down to q (a )=1.2 without significant n = 1 kink mode activity after the initial plasma current rise phase of the discharge. The disruption dynamics of these low edge safety factor plasmas depend upon the fraction of helical field rotational transform from external stellarator coils to that generated by the plasma current. We observe that with approximately 10% of the total rotational transform supplied by the stellarator coils, low edge q disruptions are passively suppressed and avoided even though q(a) < 2. When the plasma does disrupt, the instability precursors measured and implicated as the cause are internal tearing modes with poloidal, m, and toroidal, n, helical mode numbers of m /n =3 /2 and 4/3 observed on external magnetic sensors and m /n =1 /1 activity observed on core soft x-ray emissivity measurements. Even though the edge safety factor passes through and becomes much less than q(a) < 2, external n = 1 kink mode activity does not appear to play a significant role in the disruption phenomenology observed.

  14. Role of Na+/H+ exchanger regulatory factor 1 in forward trafficking of the type IIa Na+-Pi cotransporter.

    PubMed

    Ketchem, Corey J; Khundmiri, Syed J; Gaweda, Adam E; Murray, Rebecca; Clark, Barbara J; Weinman, Edward J; Lederer, Eleanor D

    2015-07-15

    Na+/H+ exchanger regulatory factor (NHERF1) plays a critical role in the renal transport of phosphate by binding to Na+-Pi cotransporter (NpT2a) in the proximal tubule. While the association between NpT2a and NHERF1 in the apical membrane is known, the role of NHERF1 to regulate the trafficking of NpT2a has not been studied. To address this question, we performed cell fractionation by sucrose gradient centrifugation in opossum kidney (OK) cells placed in low-Pi medium to stimulate forward trafficking of NpT2a. Immunoblot analysis demonstrated expression of NpT2a and NHERF1 in the endoplasmic reticulum (ER)/Golgi. Coimmunoprecipitation demonstrated a NpT2a-NHERF1 interaction in the ER/Golgi. Low-Pi medium for 4 and 8 h triggered a decrease in NHERF1 in the plasma membrane with a corresponding increase in the ER/Golgi. Time-lapse total internal reflection fluorescence imaging of OK cells placed in low-Pi medium, paired with particle tracking and mean square displacement analysis, indicated active directed movement of NHERF1 at early and late time points, whereas NpT2a showed active movement only at later times. Silence of NHERF1 in OK cells expressing green fluorescent protein (GFP)-NpT2a resulted in an intracellular accumulation of GFP-NpT2a. Transfection with GFP-labeled COOH-terminal (TRL) PDZ-binding motif deleted or wild-type NpT2a in OK cells followed by cell fractionation and immunoprecipitation confirmed that the interaction between NpT2a and NHERF1 was dependent on the TRL motif of NpT2a. We conclude that appropriate trafficking of NpT2a to the plasma membrane is dependent on the initial association between NpT2a and NHERF1 through the COOH-terminal TRL motif of NpT2a in the ER/Golgi and requires redistribution of NHERF1 to the ER/Golgi.

  15. Cloning and expression analyses of interferon regulatory factor (IRF) 3 and 7 genes in European eel, Anguilla anguilla with the identification of genes involved in IFN production.

    PubMed

    Huang, Bei; Huang, Wen Shu; Nie, P

    2014-04-01

    Interferon regulatory factor (IRF) 3 and IRF7 have been identified as regulators of type I interferon (IFN) gene expression in mammals. In the present study, the two genes were cloned and characterized in the European eel, Anguilla anguilla. The full-length cDNA sequence of IRF3 and IRF7 in the European eel, named as AaIRF3 and AaIRF7 consists of 2879 and 2419 bp respectively. Multiple alignments showed that the two IRFs have a highly conserved DNA binding domain (DBD) in the N terminus, with the characteristic motif containing five tryptophan residues, which is a feature present in their mammalian homologues. But, IRF7 has only four of the five residues in other species of fish. The expression of AaIRF3 and AaIRF7 both displayed an obvious dose-dependent manner following polyinosinic:polycytidylic acid (PolyI:C) challenge. In vivo expression analysis showed that the mRNA level of AaIRF3 and AaIRF7 was significantly up-regulated in response to PolyI:C stimulation in all examined tissues/organs except in muscle, with a lower level of increase observed in response to lipopolysaccharide (LPS) challenge and Edwardsiella tarda infection, indicating that AaIRF3 and AaIRF7 may be more likely involved in antiviral immune response. In addition, some pattern recognition receptors genes related with the production of type I IFNs and those genes in response to type I IFNs were identified in the European eel genome database, indicating a relatively conserved system in the production of type I IFN and its signalling in the European eel.

  16. Cloning and expression analyses of interferon regulatory factor (IRF) 3 and 7 genes in European eel, Anguilla anguilla with the identification of genes involved in IFN production.

    PubMed

    Huang, Bei; Huang, Wen Shu; Nie, P

    2014-04-01

    Interferon regulatory factor (IRF) 3 and IRF7 have been identified as regulators of type I interferon (IFN) gene expression in mammals. In the present study, the two genes were cloned and characterized in the European eel, Anguilla anguilla. The full-length cDNA sequence of IRF3 and IRF7 in the European eel, named as AaIRF3 and AaIRF7 consists of 2879 and 2419 bp respectively. Multiple alignments showed that the two IRFs have a highly conserved DNA binding domain (DBD) in the N terminus, with the characteristic motif containing five tryptophan residues, which is a feature present in their mammalian homologues. But, IRF7 has only four of the five residues in other species of fish. The expression of AaIRF3 and AaIRF7 both displayed an obvious dose-dependent manner following polyinosinic:polycytidylic acid (PolyI:C) challenge. In vivo expression analysis showed that the mRNA level of AaIRF3 and AaIRF7 was significantly up-regulated in response to PolyI:C stimulation in all examined tissues/organs except in muscle, with a lower level of increase observed in response to lipopolysaccharide (LPS) challenge and Edwardsiella tarda infection, indicating that AaIRF3 and AaIRF7 may be more likely involved in antiviral immune response. In addition, some pattern recognition receptors genes related with the production of type I IFNs and those genes in response to type I IFNs were identified in the European eel genome database, indicating a relatively conserved system in the production of type I IFN and its signalling in the European eel. PMID:24565894

  17. The Streptococcus pyogenes serotype M49 Nra-Ralp3 transcriptional regulatory network and its control of virulence factor expression from the novel eno ralp3 epf sagA pathogenicity region.

    PubMed

    Kreikemeyer, Bernd; Nakata, Masanobu; Köller, Thomas; Hildisch, Hendrikje; Kourakos, Vassilios; Standar, Kerstin; Kawabata, Shigetada; Glocker, Michael O; Podbielski, Andreas

    2007-12-01

    Many Streptococcus pyogenes (group A streptococcus [GAS]) virulence factor- and transcriptional regulator-encoding genes cluster together in discrete genomic regions. Nra is a central regulator of the FCT region. Previous studies exclusively described Nra as a transcriptional repressor of adhesin and toxin genes. Here transcriptome and proteome analysis of a serotype M49 GAS strain and an isogenic Nra mutant of this strain revealed the complete Nra regulon profile. Nra is active in all growth phases tested, with the largest regulon in the transition phase. Almost exclusively, virulence factor-encoding genes are repressed by Nra; these genes include the GAS pilus operon, the capsule synthesis operon, the cytolysin-mediated translocation system genes, all Mga region core virulence genes, and genes encoding other regulators, like the Ihk/Irr system, Rgg, and two additional RofA-like protein family regulators. Surprisingly, our experiments revealed that Nra additionally acts as a positive regulator, mostly for genes encoding proteins and enzymes with metabolic functions. Epidemiological investigations revealed strong genetic linkage of one particular Nra-repressed regulator, Ralp3 (SPy0735), with a gene encoding Epf (extracellular protein factor from Streptococcus suis). In a serotype-specific fashion, this ralp3 epf gene block is integrated, most likely via transposition, into the eno sagA virulence gene block, which is present in all GAS serotypes. In GAS serotypes M1, M4, M12, M28, and M49 this novel discrete genetic region is therefore designated the eno ralp3 epf sagA (ERES) pathogenicity region. Functional experiments showed that Epf is a novel GAS plasminogen-binding protein and revealed that Ralp3 activity counteracts Nra and MsmR regulatory activity. In addition to the Mga and FCT regions, the ERES region is the third discrete chromosomal pathogenicity region. All of these regions are transcriptionally linked, adding another level of complexity to the known

  18. Regulatory skill as a resilience factor for adults with a history of foster care: a pilot study.

    PubMed

    Johnson, Angela J; Tottenham, Nim

    2015-01-01

    Individuals with a history of foster care (FC) are at elevated risk for emotion regulation-related mental illness. The purpose of the current study was to characterize regulatory function in a group of adults with a history of FC (N = 26) relative to those without a history of FC (N = 27) and how regulatory function moderates adverse caregiving-related outcomes (daily cortisol production and trait anxiety). Self-report items (anxiety, emotion regulation strategies, inhibitory control, caregiving history) were collected along with more objective measures (computerized task and salivary cortisol). Inhibitory control was assessed via self-report and a computerized task (emotional face go/nogo). Results showed that for adults with a history of FC, higher levels of inhibitory control were associated with higher accuracy on the emotional face go/nogo task and greater reported use of the emotion regulation strategy cognitive reappraisal. Greater use of cognitive reappraisal in turn was associated with healthier stress-related outcomes (decreased trait anxiety and steeper sloped cortisol production throughout the day). Dose-response associations were observed between self-reported regulatory skills and FC experiences (i.e., number of placements and age when exited foster care). These findings suggest that adverse caregiving can have long-term influences on mental health that extend into adulthood; however, individual differences in regulatory skills moderate these outcomes and may be an important target for intervention following caregiving adversity. PMID:25270099

  19. The Protein Phosphatase 2A Regulatory Subunit B56γ Mediates Suppression of T Cell Receptor (TCR)-induced Nuclear Factor-κB (NF-κB) Activity*

    PubMed Central

    Breuer, Rebecca; Becker, Michael S.; Brechmann, Markus; Mock, Thomas; Arnold, Rüdiger; Krammer, Peter H.

    2014-01-01

    NF-κB is an important transcription factor in the immune system, and aberrant NF-κB activity contributes to malignant diseases and autoimmunity. In T cells, NF-κB is activated upon TCR stimulation, and signal transduction to NF-κB activation is triggered by a cascade of phosphorylation events. However, fine-tuning and termination of TCR signaling are only partially understood. Phosphatases oppose the role of kinases by removing phosphate moieties. The catalytic activity of the protein phosphatase PP2A has been implicated in the regulation of NF-κB. PP2A acts in trimeric complexes in which the catalytic subunit is promiscuous and the regulatory subunit confers substrate specificity. To understand and eventually target NF-κB-specific PP2A functions it is essential to define the regulatory PP2A subunit involved. So far, the regulatory PP2A subunit that mediates NF-κB suppression in T cells remained undefined. By performing a siRNA screen in Jurkat T cells harboring a NF-κB-responsive luciferase reporter, we identified the PP2A regulatory subunit B56γ as negative regulator of NF-κB in TCR signaling. B56γ was strongly up-regulated upon primary human T cell activation, and B56γ silencing induced increased IκB kinase (IKK) and IκBα phosphorylation upon TCR stimulation. B56γ silencing enhanced NF-κB activity, resulting in increased NF-κB target gene expression including the T cell cytokine IL-2. In addition, T cell proliferation was increased upon B56γ silencing. These data help to understand the physiology of PP2A function in T cells and the pathophysiology of diseases involving PP2A and NF-κB. PMID:24719332

  20. Exocytosis of CTLA-4 is dependent on phospholipase D and ADP ribosylation factor-1 and stimulated during activation of regulatory T cells.

    PubMed

    Mead, Karen I; Zheng, Yong; Manzotti, Claire N; Perry, Laura C A; Liu, Michael K P; Burke, Fiona; Powner, Dale J; Wakelam, Michael J O; Sansom, David M

    2005-04-15

    CTLA-4 is an essential protein in the regulation of T cell responses that interacts with two ligands found on the surface of APCs (CD80 and CD86). CTLA-4 is itself poorly expressed on the T cell surface and is predominantly localized to intracellular compartments. We have studied the mechanisms involved in the delivery of CTLA-4 to the cell surface using a model Chinese hamster ovary cell system and compared this with activated and regulatory human T cells. We have shown that expression of CTLA-4 at the plasma membrane (PM) is controlled by exocytosis of CTLA-4-containing vesicles and followed by rapid endocytosis. Using selective inhibitors and dominant negative mutants, we have shown that exocytosis of CTLA-4 is dependent on the activity of the GTPase ADP ribosylation factor-1 and on phospholipase D activity. CTLA-4 was identified in a perinuclear compartment overlapping with the cis-Golgi marker GM-130 but did not colocalize strongly with lysosomal markers such as CD63 and lysosome-associated membrane protein. In regulatory T cells, activation of phospholipase D was sufficient to trigger release of CTLA-4 to the PM but did not inhibit endocytosis. Taken together, these data suggest that CTLA-4 may be stored in a specialized compartment in regulatory T cells that can be triggered rapidly for deployment to the PM in a phospholipase D- and ADP ribosylation factor-1-dependent manner.

  1. 25 CFR 39.1101 - Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in...

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 25 Indians 1 2012-04-01 2011-04-01 true Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in fiscal year 1982. 39.1101 Section 39.1101 Indians BUREAU OF INDIAN... Programs § 39.1101 Addition of pre-kindergarten as a weight factor to the Indian School...

  2. 25 CFR 39.1101 - Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in...

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 25 Indians 1 2011-04-01 2011-04-01 false Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in fiscal year 1982. 39.1101 Section 39.1101 Indians BUREAU OF INDIAN... Programs § 39.1101 Addition of pre-kindergarten as a weight factor to the Indian School...

  3. 25 CFR 39.1101 - Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in...

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 25 Indians 1 2014-04-01 2014-04-01 false Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in fiscal year 1982. 39.1101 Section 39.1101 Indians BUREAU OF INDIAN... Programs § 39.1101 Addition of pre-kindergarten as a weight factor to the Indian School...

  4. 25 CFR 39.1101 - Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in...

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 25 Indians 1 2010-04-01 2010-04-01 false Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in fiscal year 1982. 39.1101 Section 39.1101 Indians BUREAU OF INDIAN... Programs § 39.1101 Addition of pre-kindergarten as a weight factor to the Indian School...

  5. 25 CFR 39.1101 - Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in...

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 25 Indians 1 2013-04-01 2013-04-01 false Addition of pre-kindergarten as a weight factor to the Indian School Equalization Formula in fiscal year 1982. 39.1101 Section 39.1101 Indians BUREAU OF INDIAN... Programs § 39.1101 Addition of pre-kindergarten as a weight factor to the Indian School...

  6. Working with Policy and Regulatory Factors to Implement Universal Design in the Built Environment: The Australian Experience.

    PubMed

    Larkin, Helen; Hitch, Danielle; Watchorn, Valerie; Ang, Susan

    2015-07-01

    Built environments that are usable by all provide opportunities for engagement in meaningful occupations. However, enabling them in day to day design processes and practice is problematic for relevant professions. The purpose of this phenomenological study was to gain greater understanding of the policy and regulatory influences that promote or hinder the uptake of universal design in built environments, to inform better future design. Focus groups or telephone interviews were undertaken with 28 key building industry and disability stakeholders in Australia. Four themes were identified: the difficulties of definition; the push or pull of regulations and policy; the role of formal standards; and, shifting the focus of design thinking. The findings highlight the complexity of working within policy and regulatory contexts when implementing universal design. Occupational therapists working with colleagues from other professions must be aware of these influences, and develop the skills to work with them for successful practice. PMID:26184278

  7. Working with Policy and Regulatory Factors to Implement Universal Design in the Built Environment: The Australian Experience.

    PubMed

    Larkin, Helen; Hitch, Danielle; Watchorn, Valerie; Ang, Susan

    2015-07-15

    Built environments that are usable by all provide opportunities for engagement in meaningful occupations. However, enabling them in day to day design processes and practice is problematic for relevant professions. The purpose of this phenomenological study was to gain greater understanding of the policy and regulatory influences that promote or hinder the uptake of universal design in built environments, to inform better future design. Focus groups or telephone interviews were undertaken with 28 key building industry and disability stakeholders in Australia. Four themes were identified: the difficulties of definition; the push or pull of regulations and policy; the role of formal standards; and, shifting the focus of design thinking. The findings highlight the complexity of working within policy and regulatory contexts when implementing universal design. Occupational therapists working with colleagues from other professions must be aware of these influences, and develop the skills to work with them for successful practice.

  8. Working with Policy and Regulatory Factors to Implement Universal Design in the Built Environment: The Australian Experience

    PubMed Central

    Larkin, Helen; Hitch, Danielle; Watchorn, Valerie; Ang, Susan

    2015-01-01

    Built environments that are usable by all provide opportunities for engagement in meaningful occupations. However, enabling them in day to day design processes and practice is problematic for relevant professions. The purpose of this phenomenological study was to gain greater understanding of the policy and regulatory influences that promote or hinder the uptake of universal design in built environments, to inform better future design. Focus groups or telephone interviews were undertaken with 28 key building industry and disability stakeholders in Australia. Four themes were identified: the difficulties of definition; the push or pull of regulations and policy; the role of formal standards; and, shifting the focus of design thinking. The findings highlight the complexity of working within policy and regulatory contexts when implementing universal design. Occupational therapists working with colleagues from other professions must be aware of these influences, and develop the skills to work with them for successful practice. PMID:26184278

  9. Regulatory Skill as a Resilience Factor for Adults With a History of Foster Care: A Pilot Study

    PubMed Central

    Johnson, Angela J; Tottenham, Nim

    2015-01-01

    Individuals with a history of foster care (FC) are at elevated risk for emotion regulation-related mental illness. The purpose of the current study was to characterize regulatory function in a group of adults with a history of FC (N = 26) relative to those without a history of FC (N = 27) and how regulatory function moderates adverse caregiving-related outcomes (daily cortisol production and trait anxiety). Self-report items (anxiety, emotion regulation strategies, inhibitory control, caregiving history) were collected along with more objective measures (computerized task and salivary cortisol). Inhibitory control was assessed via self-report and a computerized task (emotional face go/nogo). Results showed that for adults with a history of FC, higher levels of inhibitory control were associated with higher accuracy on the emotional face go/nogo task and greater reported use of the emotion regulation strategy cognitive reappraisal. Greater use of cognitive reappraisal in turn was associated with healthier stress-related outcomes (decreased trait anxiety and steeper sloped cortisol production throughout the day). Dose-response associations were observed between self-reported regulatory skills and FC experiences (i.e., number of placements and age when exited foster care). These findings suggest that adverse caregiving can have long-term influences on mental health that extend into adulthood; however, individual differences in regulatory skills moderate these outcomes and may be an important target for intervention following caregiving adversity. © 2014 The Authors. Developmental Psychobiology published by Wiley Periodicals, Inc. Dev Psychobiol 57: 1–16, 2015. PMID:25270099

  10. Distinct Regulatory Changes Underlying Differential Expression of TEOSINTE BRANCHED1-CYCLOIDEA-PROLIFERATING CELL FACTOR Genes Associated with Petal Variations in Zygomorphic Flowers of Petrocosmea spp. of the Family Gesneriaceae1[OPEN

    PubMed Central

    Yang, Xia; Zhao, Xiao-Ge; Li, Chao-Qun; Liu, Jing; Qiu, Zhi-Jing; Dong, Yang; Wang, Yin-Zheng

    2015-01-01

    CYCLOIDEA (CYC)-like genes, belonging to the plant-specific TCP transcription factor family that is named after TEOSINTE BRANCHED1 (TB1) from maize (Zea mays), CYC from Antirrhinum majus, and the PROLIFERATING CELL FACTORS (PCF) from rice (Oryza sativa), have conserved dorsal identity function in patterning floral zygomorphy mainly through specific expression in dorsal petals of a flower. Their expression changes are usually related to morphological diversity of zygomorphic flowers. However, it is still a challenge to elucidate the molecular mechanism underlying their expression differentiation. It is also unknown whether CINCINNATA (CIN)-like TCP genes, locally controlling cell growth and proliferation, are involved in the evolution of floral zygomorphy. To address these questions, we selected two closely related species, i.e. Petrocosmea glabristoma and Petrocosmea sinensis, with distinct petal morphology to conduct expression, hybridization, mutant, and allele-specific expression analyses. The results show that the size change of the dorsal petals between the two species is mainly mediated by the expression differentiation of CYC1C and CYC1D, while the shape variation of all petals is related to the expression change of CIN1. In reciprocal F1 hybrids, the expression of CYC1C, CYC1D, and CIN1 conforms to an additive inheritance mode, consistent with the petal phenotypes of hybrids. Through allele-specific expression analyses, we find that the expression differentiation of these TCP genes is underlain by distinctly different types of regulatory changes. We suggest that highly redundant paralogs with identical expression patterns and interspecific expression differentiation may be controlled by remarkably different regulatory pathways because natural selection may favor different regulatory modifications rather than coding sequence changes of key developmental genes in generating morphological diversity. PMID:26351309

  11. Distinct Regulatory Changes Underlying Differential Expression of TEOSINTE BRANCHED1-CYCLOIDEA-PROLIFERATING CELL FACTOR Genes Associated with Petal Variations in Zygomorphic Flowers of Petrocosmea spp. of the Family Gesneriaceae.

    PubMed

    Yang, Xia; Zhao, Xiao-Ge; Li, Chao-Qun; Liu, Jing; Qiu, Zhi-Jing; Dong, Yang; Wang, Yin-Zheng

    2015-11-01

    CYCLOIDEA (CYC)-like genes, belonging to the plant-specific TCP transcription factor family that is named after TEOSINTE BRANCHED1 (TB1) from maize (Zea mays), CYC from Antirrhinum majus, and the PROLIFERATING CELL FACTORS (PCF) from rice (Oryza sativa), have conserved dorsal identity function in patterning floral zygomorphy mainly through specific expression in dorsal petals of a flower. Their expression changes are usually related to morphological diversity of zygomorphic flowers. However, it is still a challenge to elucidate the molecular mechanism underlying their expression differentiation. It is also unknown whether CINCINNATA (CIN)-like TCP genes, locally controlling cell growth and proliferation, are involved in the evolution of floral zygomorphy. To address these questions, we selected two closely related species, i.e. Petrocosmea glabristoma and Petrocosmea sinensis, with distinct petal morphology to conduct expression, hybridization, mutant, and allele-specific expression analyses. The results show that the size change of the dorsal petals between the two species is mainly mediated by the expression differentiation of CYC1C and CYC1D, while the shape variation of all petals is related to the expression change of CIN1. In reciprocal F1 hybrids, the expression of CYC1C, CYC1D, and CIN1 conforms to an additive inheritance mode, consistent with the petal phenotypes of hybrids. Through allele-specific expression analyses, we find that the expression differentiation of these TCP genes is underlain by distinctly different types of regulatory changes. We suggest that highly redundant paralogs with identical expression patterns and interspecific expression differentiation may be controlled by remarkably different regulatory pathways because natural selection may favor different regulatory modifications rather than coding sequence changes of key developmental genes in generating morphological diversity. PMID:26351309

  12. Regulatory Physiology

    NASA Technical Reports Server (NTRS)

    Lane, Helen W.; Whitson, Peggy A.; Putcha, Lakshmi; Baker, Ellen; Smith, Scott M.; Stewart, Karen; Gretebeck, Randall; Nimmagudda, R. R.; Schoeller, Dale A.; Davis-Street, Janis

    1999-01-01

    As noted elsewhere in this report, a central goal of the Extended Duration Orbiter Medical Project (EDOMP) was to ensure that cardiovascular and muscle function were adequate to perform an emergency egress after 16 days of spaceflight. The goals of the Regulatory Physiology component of the EDOMP were to identify and subsequently ameliorate those biochemical and nutritional factors that deplete physiological reserves or increase risk for disease, and to facilitate the development of effective muscle, exercise, and cardiovascular countermeasures. The component investigations designed to meet these goals focused on biochemical and physiological aspects of nutrition and metabolism, the risk of renal (kidney) stone formation, gastrointestinal function, and sleep in space. Investigations involved both ground-based protocols to validate proposed methods and flight studies to test those methods. Two hardware tests were also completed.

  13. The isolation and characterization of growth regulatory factors produced by a herpes simplex virus Type 2 transformed mouse tumor cell line, H238

    SciTech Connect

    Stagg, R.B.

    1988-01-01

    This study was performed in an attempt to associate HSV-2-transformation with specific growth factors in order to develop a testable model for HSV-2-transformation. We report here the isolation and characterization of four growth regulatory factors produced by H238, an HSV-2-transformed mouse tumor cell line. These factors were separated from the H238-CM by heparin-sepharose affinity chromatography into three peaks of mitogenic activity and a fourth containing inhibitory activity for splenocytes. The three peaks of mitogenic activity have been identified based on physiochemical characteristics: the first supported the anchorage-independent growth of EGF treated NRK-c-49 cells and resembles transforming growth factor-{beta} (TGF-{beta}); the second bound to lectin-coated sepharose beads and was sensitive to trypsin, neuroaminidase, and the reducing agent dithiothreitol (DTT) and, resembled a platelet-derived growth factor (PDGF)-like factor; and the third displaced ({sup 125}I)-labeled basic fibroblast growth factor (bFGF) in a dose-dependent fashion when tested with a radioimmune assay. The fourth peak was inhibitory for a variety of splenocyte function assays. A model for the interaction of these factors in vivo is presented with an emphasis on testability.

  14. The Pseudomonas aeruginosa PAO1 Two-Component Regulator CarSR Regulates Calcium Homeostasis and Calcium-Induced Virulence Factor Production through Its Regulatory Targets CarO and CarP

    PubMed Central

    Guragain, Manita; King, Michelle M.; Williamson, Kerry S.; Pérez-Osorio, Ailyn C.; Akiyama, Tatsuya; Khanam, Sharmily

    2016-01-01

    ABSTRACT Pseudomonas aeruginosa is an opportunistic human pathogen that causes severe, life-threatening infections in patients with cystic fibrosis (CF), endocarditis, wounds, or artificial implants. During CF pulmonary infections, P. aeruginosa often encounters environments where the levels of calcium (Ca2+) are elevated. Previously, we showed that P. aeruginosa responds to externally added Ca2+ through enhanced biofilm formation, increased production of several secreted virulence factors, and by developing a transient increase in the intracellular Ca2+ level, followed by its removal to the basal submicromolar level. However, the molecular mechanisms responsible for regulating Ca2+-induced virulence factor production and Ca2+ homeostasis are not known. Here, we characterized the genome-wide transcriptional response of P. aeruginosa to elevated [Ca2+] in both planktonic cultures and biofilms. Among the genes induced by CaCl2 in strain PAO1 was an operon containing the two-component regulator PA2656-PA2657 (here called carS and carR), while the closely related two-component regulators phoPQ and pmrAB were repressed by CaCl2 addition. To identify the regulatory targets of CarSR, we constructed a deletion mutant of carR and performed transcriptome analysis of the mutant strain at low and high [Ca2+]. Among the genes regulated by CarSR in response to CaCl2 are the predicted periplasmic OB-fold protein, PA0320 (here called carO), and the inner membrane-anchored five-bladed β-propeller protein, PA0327 (here called carP). Mutations in both carO and carP affected Ca2+ homeostasis, reducing the ability of P. aeruginosa to export excess Ca2+. In addition, a mutation in carP had a pleotropic effect in a Ca2+-dependent manner, altering swarming motility, pyocyanin production, and tobramycin sensitivity. Overall, the results indicate that the two-component system CarSR is responsible for sensing high levels of external Ca2+ and responding through its regulatory targets that

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

    PubMed

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

    2015-11-01

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

  16. Additional cytosine inside mitochondrial C-tract D-loop as a progression risk factor in oral precancer cases

    PubMed Central

    Pandey, Rahul; Mehrotra, Divya; Mahdi, Abbas Ali; Sarin, Rajiv; Kowtal, Pradnya

    2014-01-01

    Introduction Alterations inside Polycytosine tract (C-tract) of mitochondrial DNA (mtDNA) have been described in many different tumor types. The Poly-Cytosine region is located within the mtDNA D-loop region which acts as point of mitochondrial replication origin. A suggested pathogenesis is that it interferes with the replication process of mtDNA which in turn affects the mitochondrial functioning and generates disease. Methodology 100 premalignant cases (50 leukoplakia & 50 oral submucous fibrosis) were selected and the mitochondrial DNA were isolated from the lesion tissues and from the blood samples. Polycytosine tract in mtDNA was sequenced by direct capillary sequencing. Results 40 (25 leukoplakia & 15 oral submucous fibrosis) patients harbored lesions that displayed one additional cytosine after nucleotide thymidine (7CT6C) at nt position 316 in C-tract of mtDNA which were absent in corresponding mtDNA derived from blood samples. Conclusion Our results show an additional cytosine in the mtDNA at polycytosine site in oral precancer cases. It is postulated that any increase/decrease in the number of cytosine residues in the Poly-Cytosine region may affect the rate of mtDNA replication by impairing the binding of polymerase and other transacting factors. By promoting mitochondrial genomic instability, it may have a central role in the dysregulation of mtDNA functioning, for example alterations in energy metabolism that may promote tumor development. We, therefore, report and propose that this alteration may represent the early development of oral cancer. Further studies with large number of samples are needed in to confirm the role of such mutation in carcinogenesis. PMID:25737911

  17. MAGIA²: from miRNA and genes expression data integrative analysis to microRNA-transcription factor mixed regulatory circuits (2012 update).

    PubMed

    Bisognin, Andrea; Sales, Gabriele; Coppe, Alessandro; Bortoluzzi, Stefania; Romualdi, Chiara

    2012-07-01

    MAGIA(2) (http://gencomp.bio.unipd.it/magia2) is an update, extension and evolution of the MAGIA web tool. It is dedicated to the integrated analysis of in silico target prediction, microRNA (miRNA) and gene expression data for the reconstruction of post-transcriptional regulatory networks. miRNAs are fundamental post-transcriptional regulators of several key biological and pathological processes. As miRNAs act prevalently through target degradation, their expression profiles are expected to be inversely correlated to those of the target genes. Low specificity of target prediction algorithms makes integration approaches an interesting solution for target prediction refinement. MAGIA(2) performs this integrative approach supporting different association measures, multiple organisms and almost all target predictions algorithms. Nevertheless, miRNAs activity should be viewed as part of a more complex scenario where regulatory elements and their interactors generate a highly connected network and where gene expression profiles are the result of different levels of regulation. The updated MAGIA(2) tries to dissect this complexity by reconstructing mixed regulatory circuits involving either miRNA or transcription factor (TF) as regulators. Two types of circuits are identified: (i) a TF that regulates both a miRNA and its target and (ii) a miRNA that regulates both a TF and its target.

  18. Regulatory approaches to obesity prevention: A systematic overview of current laws addressing diet-related risk factors in the European Union and the United States.

    PubMed

    Sisnowski, Jana; Handsley, Elizabeth; Street, Jackie M

    2015-06-01

    High prevalence of overweight and obesity remains a significant international public health problem. Law has been identified as a tool for obesity prevention and selected high-profile measures have been reported. However, the nature and extent of enacted legislation internationally are unclear. This research provides an overview of regulatory approaches enacted in the United States, the European Union, and EU Member States since 2004. To this end, relevant databases of primary and secondary legislation were systematically searched to identify and explore laws addressing dietary risk factors for obesity. Across jurisdictions, current regulatory approaches to obesity prevention are limited in reach and scope. Target groups are rarely the general population, but instead sub-populations in government-supported settings. Consumer information provision is preferred over taxation and marketing restrictions other than the regulation of health and nutrition claims. In the EU in particular, product reformulation with industry consent has also emerged as a popular small-scale measure. While consistent and widespread use of law is lacking, governments have employed a range of regulatory measures in the name of obesity prevention, indicating that there is, in principle, political will. Results from this study may serve as a starting point for future research and policy development. PMID:25963556

  19. Regulatory approaches to obesity prevention: A systematic overview of current laws addressing diet-related risk factors in the European Union and the United States.

    PubMed

    Sisnowski, Jana; Handsley, Elizabeth; Street, Jackie M

    2015-06-01

    High prevalence of overweight and obesity remains a significant international public health problem. Law has been identified as a tool for obesity prevention and selected high-profile measures have been reported. However, the nature and extent of enacted legislation internationally are unclear. This research provides an overview of regulatory approaches enacted in the United States, the European Union, and EU Member States since 2004. To this end, relevant databases of primary and secondary legislation were systematically searched to identify and explore laws addressing dietary risk factors for obesity. Across jurisdictions, current regulatory approaches to obesity prevention are limited in reach and scope. Target groups are rarely the general population, but instead sub-populations in government-supported settings. Consumer information provision is preferred over taxation and marketing restrictions other than the regulation of health and nutrition claims. In the EU in particular, product reformulation with industry consent has also emerged as a popular small-scale measure. While consistent and widespread use of law is lacking, governments have employed a range of regulatory measures in the name of obesity prevention, indicating that there is, in principle, political will. Results from this study may serve as a starting point for future research and policy development.

  20. MAGIA2: from miRNA and genes expression data integrative analysis to microRNA–transcription factor mixed regulatory circuits (2012 update)

    PubMed Central

    Bisognin, Andrea; Sales, Gabriele; Coppe, Alessandro; Bortoluzzi, Stefania; Romualdi, Chiara

    2012-01-01

    MAGIA2 (http://gencomp.bio.unipd.it/magia2) is an update, extension and evolution of the MAGIA web tool. It is dedicated to the integrated analysis of in silico target prediction, microRNA (miRNA) and gene expression data for the reconstruction of post-transcriptional regulatory networks. miRNAs are fundamental post-transcriptional regulators of several key biological and pathological processes. As miRNAs act prevalently through target degradation, their expression profiles are expected to be inversely correlated to those of the target genes. Low specificity of target prediction algorithms makes integration approaches an interesting solution for target prediction refinement. MAGIA2 performs this integrative approach supporting different association measures, multiple organisms and almost all target predictions algorithms. Nevertheless, miRNAs activity should be viewed as part of a more complex scenario where regulatory elements and their interactors generate a highly connected network and where gene expression profiles are the result of different levels of regulation. The updated MAGIA2 tries to dissect this complexity by reconstructing mixed regulatory circuits involving either miRNA or transcription factor (TF) as regulators. Two types of circuits are identified: (i) a TF that regulates both a miRNA and its target and (ii) a miRNA that regulates both a TF and its target. PMID:22618880

  1. “Pressured to prescribe” The impact of economic and regulatory factors on South-Eastern ED physicians when managing the drug seeking patient

    PubMed Central

    Kelly, Sharon; Johnson, Giffe T.; Harbison, Raymond D.

    2016-01-01

    Objective: The purpose of this study was to elicit the opinions of Emergency Department (ED) physicians, currently practicing in the United States, regarding the impact of economic and regulatory factors on their management of patients exhibiting “drug seeking” behavior. Methods: A descriptive, cross-sectional study, utilizing a convenience sample of ED physicians located in Florida and Georgia was conducted for a period of 2 months. The inclusion criteria specified that any ED physician, currently practicing within the United States, could participate. Results: Of the ED physicians surveyed (n = 141), 71% reported a perceived pressure to prescribe opioid analgesics to avoid administrative and regulatory criticism and 98% related patient satisfaction scores as being too highly emphasized by reimbursement entities as a means of evaluating their patient management. Rising patient volumes and changes in the healthcare climate were cited by ED physicians as impacting their management of patients exhibiting “drug seeking” behavior. Conclusions: The ED physician faces unique challenges in changing healthcare and economic climates. Requirements to address pain as the “fifth vital sign,” patient satisfaction based reimbursement metrics and an economically driven rise in ED patient volume, may have inadvertently created an environment conducive to exploitation by prescription opioid abusers. There is an identified need for the development of continuing medical education and standardized regulatory and legislative protocols to assist ED physicians in the appropriate management of patients exhibiting “drug seeking” behavior. PMID:27162437

  2. Overexpression of E2F mRNAs associated with gastric cancer progression identified by the transcription factor and miRNA co-regulatory network analysis.

    PubMed

    Zhang, XiaoTian; Ni, ZhaoHui; Duan, ZiPeng; Xin, ZhuoYuan; Wang, HuaiDong; Tan, JiaYi; Wang, GuoQing; Li, Fan

    2015-01-01

    Gene expression is regulated at the transcription and translation levels; thus, both transcription factors (TFs) and microRNAs (miRNA) play roles in regulation of gene expression. This study profiled differentially expressed mRNAs and miRNAs in gastric cancer tissues to construct a TF and miRNA co-regulatory network in order to identify altered genes in gastric cancer progression. A total of 70 cases gastric cancer and paired adjacent normal tissues were subjected to cDNA and miRNA microarray analyses. We obtained 887 up-regulated and 93 down-regulated genes and 41 down-regulated and 4 up-regulated miRNAs in gastric cancer tissues. Using the Transcriptional Regulatory Element Database, we obtained 105 genes that are regulated by the E2F family of genes and using Targetscan, miRanda, miRDB and miRWalk tools, we predicted potential targeting genes of these 45 miRNAs. We then built up the E2F-related TF and miRNA co-regulatory gene network and identified 9 hub-genes. Furthermore, we found that levels of E2F1, 2, 3, 4, 5, and 7 mRNAs associated with gastric cancer cell invasion capacity, and has associated with tumor differentiation. These data showed Overexpression of E2F mRNAs associated with gastric cancer progression.

  3. A large-scale, in vivo transcription factor screen defines bivalent chromatin as a key property of regulatory factors mediating Drosophila wing development

    PubMed Central

    Schertel, Claus; Albarca, Monica; Rockel-Bauer, Claudia; Kelley, Nicholas W.; Bischof, Johannes; Hens, Korneel

    2015-01-01

    Transcription factors (TFs) are key regulators of cell fate. The estimated 755 genes that encode DNA binding domain-containing proteins comprise ∼5% of all Drosophila genes. However, the majority has remained uncharacterized so far due to the lack of proper genetic tools. We generated 594 site-directed transgenic Drosophila lines that contain integrations of individual UAS-TF constructs to facilitate spatiotemporally controlled misexpression in vivo. All transgenes were expressed in the developing wing, and two-thirds induced specific phenotypic defects. In vivo knockdown of the same genes yielded a phenotype for 50%, with both methods indicating a great potential for misexpression to characterize novel functions in wing growth, patterning, and development. Thus, our UAS-TF library provides an important addition to the genetic toolbox of Drosophila research, enabling the identification of several novel wing development-related TFs. In parallel, we established the chromatin landscape of wing imaginal discs by ChIP-seq analyses of five chromatin marks and RNA Pol II. Subsequent clustering revealed six distinct chromatin states, with two clusters showing enrichment for both active and repressive marks. TFs that carry such “bivalent” chromatin are highly enriched for causing misexpression phenotypes in the wing, and analysis of existing expression data shows that these TFs tend to be differentially expressed across the wing disc. Thus, bivalently marked chromatin can be used as a marker for spatially regulated TFs that are functionally relevant in a developing tissue. PMID:25568052

  4. Natural Disruption of Two Regulatory Networks in Serotype M3 Group A Streptococcus Isolates Contributes to the Virulence Factor Profile of This Hypervirulent Serotype

    PubMed Central

    Cao, Tram N.; Liu, Zhuyun; Cao, Tran H.; Pflughoeft, Kathryn J.; Treviño, Jeanette; Danger, Jessica L.; Beres, Stephen B.; Musser, James M.

    2014-01-01

    Despite the public health challenges associated with the emergence of new pathogenic bacterial strains and/or serotypes, there is a dearth of information regarding the molecular mechanisms that drive this variation. Here, we began to address the mechanisms behind serotype-specific variation between serotype M1 and M3 strains of the human pathogen Streptococcus pyogenes (the group A Streptococcus [GAS]). Spatially diverse contemporary clinical serotype M3 isolates were discovered to contain identical inactivating mutations within genes encoding two regulatory systems that control the expression of important virulence factors, including the thrombolytic agent streptokinase, the protease inhibitor-binding protein-G-related α2-macroglobulin-binding (GRAB) protein, and the antiphagocytic hyaluronic acid capsule. Subsequent analysis of a larger collection of isolates determined that M3 GAS, since at least the 1920s, has harbored a 4-bp deletion in the fasC gene of the fasBCAX regulatory system and an inactivating polymorphism in the rivR regulator-encoding gene. The fasC and rivR mutations in M3 isolates directly affect the virulence factor profile of M3 GAS, as evident by a reduction in streptokinase expression and an enhancement of GRAB expression. Complementation of the fasC mutation in M3 GAS significantly enhanced levels of the small regulatory RNA FasX, which in turn enhanced streptokinase expression. Complementation of the rivR mutation in M3 GAS restored the regulation of grab mRNA abundance but did not alter capsule mRNA levels. While important, the fasC and rivR mutations do not provide a full explanation for why serotype M3 strains are associated with unusually severe invasive infections; thus, further investigation is warranted. PMID:24516115

  5. Effects of Factor XIII Deficiency on Thromboelastography. Thromboelastography with Calcium and Streptokinase Addition is more Sensitive than Solubility Tests

    PubMed Central

    Martinuzzo, M.; Barrera, L.; Altuna, D.; Baña, F. Tisi; Bieti, J.; Amigo, Q.; D’Adamo, M.; López, M.S.; Oyhamburu, J.; Otaso, J.C.

    2016-01-01

    Background Homozygous or double heterozygous factor XIII (FXIII) deficiency is characterized by soft tissue hematomas, intracranial and delayed spontaneous bleeding. Alterations of thromboelastography (TEG) parameters in these patients have been reported. The aim of the study was to show results of TEG, TEG Lysis (Lys 60) induced by subthreshold concentrations of streptokinase (SK), and to compare them to the clot solubility studies results in samples of a 1-year-old girl with homozygous or double heterozygous FXIII deficiency. Case A year one girl with a history of bleeding from the umbilical cord. During her first year of life, several hematomas appeared in soft upper limb tissue after punctures for vaccination and a gluteal hematoma. One additional sample of a heterozygous patient and three samples of acquired FXIII deficiency were also evaluated. Materials and Methods Clotting tests, von Willebrand factor (vWF) antigen and activity, plasma FXIII-A subunit (pFXIII-A) were measured by an immunoturbidimetric assay in a photo-optical coagulometer. Solubility tests were performed with Ca2+-5 M urea and thrombin-2% acetic acid. Basal and post-FXIII concentrate infusion samples were studied. TEG was performed with CaCl2 or CaCl2 + SK (3.2 U/mL) in a Thromboelastograph. Results Prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time, fibrinogen, factor VIIIc, vWF, and platelet aggregation were normal. Antigenic pFXIII-A subunit was < 2%. TEG, evaluated at diagnosis and post FXIII concentrate infusion (pFXIII-A= 37%), presented a normal reaction time (R), 8 min, prolonged k (14 and 11min respectively), a low Maximum-Amplitude (MA) ( 39 and 52 mm respectively), and Clot Lysis (Lys60) slightly increased (23 and 30% respectively). In the sample at diagnosis, clot solubility was abnormal, 50 and 45 min with Ca-Urea and thrombin-acetic acid, respectively, but normal (>16 hours) 1-day post-FXIII infusion. Analysis of FXIII deficient and normal

  6. Effects of Factor XIII Deficiency on Thromboelastography. Thromboelastography with Calcium and Streptokinase Addition is more Sensitive than Solubility Tests

    PubMed Central

    Martinuzzo, M.; Barrera, L.; Altuna, D.; Baña, F. Tisi; Bieti, J.; Amigo, Q.; D’Adamo, M.; López, M.S.; Oyhamburu, J.; Otaso, J.C.

    2016-01-01

    Background Homozygous or double heterozygous factor XIII (FXIII) deficiency is characterized by soft tissue hematomas, intracranial and delayed spontaneous bleeding. Alterations of thromboelastography (TEG) parameters in these patients have been reported. The aim of the study was to show results of TEG, TEG Lysis (Lys 60) induced by subthreshold concentrations of streptokinase (SK), and to compare them to the clot solubility studies results in samples of a 1-year-old girl with homozygous or double heterozygous FXIII deficiency. Case A year one girl with a history of bleeding from the umbilical cord. During her first year of life, several hematomas appeared in soft upper limb tissue after punctures for vaccination and a gluteal hematoma. One additional sample of a heterozygous patient and three samples of acquired FXIII deficiency were also evaluated. Materials and Methods Clotting tests, von Willebrand factor (vWF) antigen and activity, plasma FXIII-A subunit (pFXIII-A) were measured by an immunoturbidimetric assay in a photo-optical coagulometer. Solubility tests were performed with Ca2+-5 M urea and thrombin-2% acetic acid. Basal and post-FXIII concentrate infusion samples were studied. TEG was performed with CaCl2 or CaCl2 + SK (3.2 U/mL) in a Thromboelastograph. Results Prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time, fibrinogen, factor VIIIc, vWF, and platelet aggregation were normal. Antigenic pFXIII-A subunit was < 2%. TEG, evaluated at diagnosis and post FXIII concentrate infusion (pFXIII-A= 37%), presented a normal reaction time (R), 8 min, prolonged k (14 and 11min respectively), a low Maximum-Amplitude (MA) ( 39 and 52 mm respectively), and Clot Lysis (Lys60) slightly increased (23 and 30% respectively). In the sample at diagnosis, clot solubility was abnormal, 50 and 45 min with Ca-Urea and thrombin-acetic acid, respectively, but normal (>16 hours) 1-day post-FXIII infusion. Analysis of FXIII deficient and normal

  7. Distinctive Expression of Bcl-2 Factors in Regulatory T Cells Determines a Pharmacological Target to Induce Immunological Tolerance

    PubMed Central

    Gabriel, Sarah Sharon; Bon, Nina; Chen, Jin; Wekerle, Thomas; Bushell, Andrew; Fehr, Thomas; Cippà, Pietro Ernesto

    2016-01-01

    Distinctive molecular characteristics of functionally diverse lymphocyte populations may represent novel pharmacological targets for immunotherapy. The intrinsic apoptosis pathway is differently regulated among conventional and regulatory T cells (Tregs). Targeted pharmacological modulation of this pathway with a small molecule Bcl-2/Bcl-xL inhibitor (ABT-737) caused a selective depletion of effector T cells and a relative enrichment of Tregs in vivo. Treatment with ABT-737 resulted in a tolerogenic milieu, which was exploited to alleviate graft-versus-host disease, to prevent allograft rejection in a stringent fully MHC-mismatched skin transplantation model and to induce immunological tolerance in combination with bone marrow transplantation. This concept has the potential to find various applications for immunotherapy, since it allows pharmacologic exploitation of the immunomodulatory properties of Tregs without the need for cell manipulation ex vivo. PMID:26973650

  8. Sigma Factor N, Liaison to an ntrC and rpoS Dependent Regulatory Pathway Controlling Acid Resistance and the LEE in Enterohemorrhagic Escherichia coli

    PubMed Central

    Mitra, Avishek; Fay, Pamela A.; Morgan, Jason K.; Vendura, Khoury W.; Versaggi, Salvatore L.; Riordan, James T.

    2012-01-01

    Enterohemorrhagic Escherichia coli (EHEC) is dependent on acid resistance for gastric passage and low oral infectious dose, and the locus of enterocyte effacement (LEE) for intestinal colonization. Mutation of rpoN, encoding sigma factor N (σN), dramatically alters the growth-phase dependent regulation of both acid resistance and the LEE. This study reports on the determinants of σN-directed acid resistance and LEE expression, and the underlying mechanism attributable to this phenotype. Glutamate-dependent acid resistance (GDAR) in TW14359ΔrpoN correlated with increased expression of the gadX-gadW regulatory circuit during exponential growth, whereas upregulation of arginine-dependent acid resistance (ADAR) genes adiA and adiC in TW14359ΔrpoN did not confer acid resistance by the ADAR mechanism. LEE regulatory (ler), structural (espA and cesT) and effector (tir) genes were downregulated in TW14359ΔrpoN, and mutation of rpoS encoding sigma factor 38 (σS) in TW14359ΔrpoN restored acid resistance and LEE genes to WT levels. Stability, but not the absolute level, of σS was increased in TW14359ΔrpoN; however, increased stability was not solely attributable to the GDAR and LEE expression phenotype. Complementation of TW14359ΔrpoN with a σN allele that binds RNA polymerase (RNAP) but not DNA, did not restore WT levels of σS stability, gadE, ler or GDAR, indicating a dependence on transcription from a σN promoter(s) and not RNAP competition for the phenotype. Among a library of σN enhancer binding protein mutants, only TW14359ΔntrC, inactivated for nitrogen regulatory protein NtrC, phenocopied TW14359ΔrpoN for σS stability, GDAR and ler expression. The results of this study suggest that during exponential growth, NtrC-σN regulate GDAR and LEE expression through downregulation of σS at the post-translational level; likely by altering σS stability or activity. The regulatory interplay between NtrC, other EBPs, and σN–σS, represents a mechanism by

  9. Mexican American First-Generation Students' Perceptions of Siblings and Additional Factors Influencing Their College Choice Process

    ERIC Educational Resources Information Center

    Elias McAllister, Dora

    2012-01-01

    The purpose of this study was to understand the factors influencing the college choice process of Mexican American first-generation students who had an older sibling with college experience. While a considerable amount of research exists on factors influencing the college choice process of first-generation college students, and a few studies…

  10. Specific cis-acting sequence for PHO8 expression interacts with PHO4 protein, a positive regulatory factor, in Saccharomyces cerevisiae.

    PubMed Central

    Hayashi, N; Oshima, Y

    1991-01-01

    The PHO8 gene of Saccharomyces cerevisiae encodes repressible alkaline phosphatase (rALPase; EC 3.1.3.1). The rALPase activity of the cells is two to three times higher in medium containing a low concentration of Pi than in high-Pi medium due to transcription of PHO8. The Pi signals are conveyed to PHO8 by binding of PHO4 protein, a positive regulatory factor, to a promoter region of PHO8 (PHO8p) under the influence of the PHO regulatory circuit. Deletion analysis of PHO8p DNA revealed two separate regulatory regions required for derepression of rALPase located at nucleotide positions -704 to -661 (distal region) and -548 to -502 (proximal region) and an inhibitory region located at -421 to -289 relative to the translation initiation codon. Gel retardation experiments showed that a beta-galactosidase-PHO4 fusion protein binds to a 132-bp PHO8p fragment bearing the proximal region but not to a 226-bp PHO8 DNA bearing the distal region. The fusion protein also binds to a synthetic oligonucleotide having the same 12-bp nucleotide sequence as the PHO8p DNA from positions -536 to -525. The 132-bp PHO8p fragment, connected at position -281 of the 5' upstream region of a HIS5'-'lacZ fused gene, could sense Pi signals in vivo, but a 20-bp synthetic oligonucleotide having the same sequence from -544 to -525 of the PHO8p DNA could not. Linker insertions in the PHO8p DNA indicated that the 5-bp sequence 5'-CACGT-3' from positions -535 to -531 is essential for binding the beta-galactosidase-PHO4 fusion protein and for derepression of rALPase. Images PMID:1990283

  11. Lovastatin insensitive 1, a Novel pentatricopeptide repeat protein, is a potential regulatory factor of isoprenoid biosynthesis in Arabidopsis.

    PubMed

    Kobayashi, Keiko; Suzuki, Masashi; Tang, Jianwei; Nagata, Noriko; Ohyama, Kiyoshi; Seki, Hikaru; Kiuchi, Reiko; Kaneko, Yasuko; Nakazawa, Miki; Matsui, Minami; Matsumoto, Shogo; Yoshida, Shigeo; Muranaka, Toshiya

    2007-02-01

    Higher plants have two metabolic pathways for isoprenoid biosynthesis: the cytosolic mevalonate (MVA) pathway and the plastidal non-mevalonate (MEP) pathway. Despite the compartmentalization of these two pathways, metabolic flow occurs between them. However, little is known about the mechanisms that regulate the two pathways and the metabolic cross-talk. To identify such regulatory mechanisms, we isolated and characterized the Arabidopsis T-DNA insertion mutant lovastatin insensitive 1 (loi1), which is resistant to lovastatin and clomazone, inhibitors of the MVA and MEP pathways, respectively. The accumulation of the major products of these pathways, i.e. sterols and chlorophyll, was less affected by lovastatin and clomazone, respectively, in loi1 than in the wild type. Furthermore, the 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) activity analysis showed higher activity of HMGR in loi1-1 treated with lovastatin than that in the WT. We consider that the lovastatin-resistant phenotype of loi1-1 was derived from this post-transcriptional up-regulation of HMGR. The LOI1 gene encodes a novel pentatricopeptide repeat (PPR) protein. PPR proteins are thought to regulate the expression of genes encoded in organelle genomes by post-transcriptional regulation in mitochondria or plastids. Our results demonstrate that LOI1 is predicted to localize in mitochondria and has the ability to bind single-stranded nucleic acids. Our investigation revealed that the post-transcriptional regulation of mitochondrial RNA may be involved in isoprenoid biosynthesis in both the MVA and MEP pathways.

  12. Stress-dependent and -independent expression of the myogenic regulatory factors and the MARP genes after eccentric contractions in rats

    PubMed Central

    Hentzen, Eric R; Lahey, Michele; Peters, David; Mathew, Liby; Barash, Ilona A; Fridén, Jan; Lieber, Richard L

    2006-01-01

    The relationship between muscle mechanical conditions and gene expression was investigated by varying both stress and contraction mode imposed upon rat dorsiflexors (n = 25), activating them at high or low frequencies (150 Hz or 40 Hz) either eccentrically or isometrically. Muscle physiological, immunohistochemical and gene expression changes were then measured 24 h after the exercise bout. Peak stress was the best predictor of muscle injury, independent of contraction mode (i.e. eccentric or isometric). When peak stresses were matched, no physiological or immunohistochemical differences were detected between isometric and eccentric contractions. The expression of certain myogenic regulatory and muscle ankyrin repeat protein (MARP) genes (myoD, myogenin, MLP and CARP) depended both on peak muscle stress achieved during contraction and contraction mode. In contrast, Arpp/Ankrd2 was dramatically upregulated only by eccentric contractions, but not by isometric contractions, even though the stress level of the eccentric contractions varied over a three-fold range and overlapped with that of the isometric group. The role that Arpp/Ankrd2 upregulation plays in the biological response to eccentric contraction remains to be determined, as does the control mechanism whereby the expression of certain genes (such as myoD, myogenin, MLP and CARP) is sensitive to muscle stress while another (Arpp/Ankrd2) is sensitive only to contraction mode. PMID:16239282

  13. Construction and analysis of regulatory genetic networks in cervical cancer based on involved microRNAs, target genes, transcription factors and host genes.

    PubMed

    Wang, Ning; Xu, Zhiwen; Wang, Kunhao; Zhu, Minghui; Li, Yang

    2014-04-01

    Over recent years, genes and microRNA (miRNA/miR) have been considered as key biological factors in human carcinogenesis. During cancer development, genes may act as multiple identities, including target genes of miRNA, transcription factors and host genes. The present study concentrated on the regulatory networks consisting of the biological factors involved in cervical cancer in order to investigate their features and affect on this specific pathology. Numerous raw data was collected and organized into purposeful structures, and adaptive procedures were defined for application to the prepared data. The networks were therefore built with the factors as basic components according to their interacting associations. The networks were constructed at three levels of interdependency, including a differentially-expressed network, a related network and a global network. Comparisons and analyses were made at a systematic level rather than from an isolated gene or miRNA. Critical hubs were extracted in the core networks and notable features were discussed, including self-adaption feedback regulation. The present study expounds the pathogenesis from a novel point of view and is proposed to provide inspiration for further investigation and therapy.

  14. The identification of transcription factors expressed in the notochord of Ciona intestinalis adds new potential players to the Brachyury gene regulatory network

    PubMed Central

    José-Edwards, Diana S.; Kerner, Pierre; Kugler, Jamie E.; Deng, Wei; Jiang, Di; Di Gregorio, Anna

    2013-01-01

    The notochord is the distinctive characteristic of chordates; however, the knowledge of the complement of transcription factors governing the development of this structure is still incomplete. Here we present the expression patterns of seven transcription factor genes detected in the notochord of the ascidian Ciona intestinalis at various stages of embryonic development. Four of these transcription factors, Fos-a, NFAT5, AFF and Klf15, have not been directly associated with the notochord in previous studies, while the others, including Spalt-like-a, Lmx-like and STAT5/6-b, display evolutionarily conserved expression in this structure as well as in other domains. We examined the hierarchical relationships between these genes and the transcription factor Brachyury, which is necessary for notochord development in all chordates. We found that Ciona Brachyury regulates the expression of most, although not all, of these genes. These results shed light on the genetic regulatory program underlying notochord formation in Ciona and possibly other chordates. PMID:21594950

  15. Dependence on nuclear factor of activated T-cells (NFAT) levels discriminates conventional T cells from Foxp3+ regulatory T cells.

    PubMed

    Vaeth, Martin; Schliesser, Ulrike; Müller, Gerd; Reissig, Sonja; Satoh, Kazuki; Tuettenberg, Andrea; Jonuleit, Helmut; Waisman, Ari; Müller, Martin R; Serfling, Edgar; Sawitzki, Birgit S; Berberich-Siebelt, Friederike

    2012-10-01

    Several lines of evidence suggest nuclear factor of activated T-cells (NFAT) to control regulatory T cells: thymus-derived naturally occurring regulatory T cells (nTreg) depend on calcium signals, the Foxp3 gene harbors several NFAT binding sites, and the Foxp3 (Fork head box P3) protein interacts with NFAT. Therefore, we investigated the impact of NFAT on Foxp3 expression. Indeed, the generation of peripherally induced Treg (iTreg) by TGF-β was highly dependent on NFAT expression because the ability of CD4(+) T cells to differentiate into iTreg diminished markedly with the number of NFAT family members missing. It can be concluded that the expression of Foxp3 in TGF-β-induced iTreg depends on the threshold value of NFAT rather than on an individual member present. This is specific for iTreg development, because frequency of nTreg remained unaltered in mice lacking NFAT1, NFAT2, or NFAT4 alone or in combination. Different from expectation, however, the function of both nTreg and iTreg was independent on robust NFAT levels, reflected by less nuclear NFAT in nTreg and iTreg. Accordingly, absence of one or two NFAT members did not alter suppressor activity in vitro or during colitis and transplantation in vivo. This scenario emphasizes an inhibition of high NFAT activity as treatment for autoimmune diseases and in transplantation, selectively targeting the proinflammatory conventional T cells, while keeping Treg functional.

  16. cIRF-3, a new member of the interferon regulatory factor (IRF) family that is rapidly and transiently induced by dsRNA.

    PubMed Central

    Grant, C E; Vasa, M Z; Deeley, R G

    1995-01-01

    In mammals, some of the effects of interferon (IFN) on gene transcription are known to be mediated by a family of IFN-inducible DNA-binding proteins, the IFN regulatory factor (IRF) family, which includes both activators and repressors of transcription. Although IFN activities have been described in many vertebrates, little is known about regulation of IFN- or IFN-stimulated genes in species other than human and mouse. Here, we report the cloning of a chicken cDNA, cIRF-3, encoding a protein with a DNA-binding domain similar to that found in the mammalian IRF family of proteins. Similarity between cIRF-3 and the mammalian IRFs is comparable with that between known members of the family. It is most similar to the IRF proteins ICSBP and ISGF3 gamma but is equally divergent from both. Gel mobility shift assays indicate that cIRF-3 is capable of binding a known IFN-stimulated response element that is conserved between the mammalian and chicken Mx genes. Expression of the cIRF-3 gene can be induced to high levels by poly(I).poly(C). Induction is rapid and transient with no requirement for protein synthesis. Co-treatment of cells with cycloheximide results in superinduction of cIRF-3 mRNA. The structural and regulatory characteristics of cIRF-3 indicate that it is the first example of a non-mammalian IRF protein. Images PMID:7541908

  17. Specific interactions between transcription factors and the promoter-regulatory region of the human cytomegalovirus major immediate-early gene

    SciTech Connect

    Ghazal, P.; Lubon, H.; Hennighausen, L. )

    1988-03-01

    Repeat sequence motifs as well as unique sequences between nucleotides {minus}150 and {minus}22 of the human cytomegalovirus immediate-early 1 gene interact in vitro with nuclear proteins. The authors show that a transcriptional element between nucleotides {minus}91 and {minus}65 stimulated promoter activity in vivo and in vitro by binding specific cellular transcription factors. Finally, a common sequence motif, (T)TGG/AC, present in 15 of the determined binding sites suggests a particular class of nuclear factors associated with the immediate-early 1 gene.

  18. Expansion of effector memory regulatory T cells represents a novel prognostic factor in lower risk myelodysplastic syndrome.

    PubMed

    Mailloux, Adam W; Sugimori, Chiharu; Komrokji, Rami S; Yang, Lili; Maciejewski, Jaroslaw P; Sekeres, Mikkael A; Paquette, Ronald; Loughran, Thomas P; List, Alan F; Epling-Burnette, Pearlie K

    2012-09-15

    Myelodysplastic syndromes are premalignant diseases characterized by cytopenias, myeloid dysplasia, immune dysregulation with association to autoimmunity, and variable risk for acute myeloid leukemia transformation. Studies of FOXP3(+) regulatory T cells (Tregs) indicate that the number and/or activation state may influence cancer progression in these patients. Focusing on patients with a lower risk for leukemia transformation, 18 (34.6%) of 52 patients studied displayed an altered Treg compartment compared with age-matched controls. Delineation of unique Treg subsets revealed that an increase in the absolute number of CD4(+)FOXP3(+)CD25(+)CD127(low)CD45RA(-)CD27(-) Tregs (effector memory Tregs [Treg(EM)]) was significantly associated with anemia (p = 0.046), reduced hemoglobin (p = 0.038), and blast counts ≥5% (p = 0.006). In healthy donors, this Treg(EM) population constitutes only 2% of all Tregs (one to six Tregs per microliter) in peripheral blood but, when isolated, exhibit greater suppressive activity in vitro. With a median follow-up of 3.1 y (range 2.7-4.9 y) from sample acquisition, increased numbers of Treg(EM) cells proved to have independent prognostic importance in survival estimates, suggesting that enumeration of this Treg subset may be a more reliable indicator of immunological escape than FOXP3(+) T cells as a whole. Based on multivariate analyses, Treg(EM) impacted survival independently from myeloblast characteristics, cytopenias, karyotype, and comorbidities. Based on these findings, Treg(EM) cell expansion may be synonymous with human Treg activation and indicate microenvironmental changes conducive to transformation in myelodysplastic syndromes.

  19. TFmiR: a web server for constructing and analyzing disease-specific transcription factor and miRNA co-regulatory networks.

    PubMed

    Hamed, Mohamed; Spaniol, Christian; Nazarieh, Maryam; Helms, Volkhard

    2015-07-01

    TFmiR is a freely available web server for deep and integrative analysis of combinatorial regulatory interactions between transcription factors, microRNAs and target genes that are involved in disease pathogenesis. Since the inner workings of cells rely on the correct functioning of an enormously complex system of activating and repressing interactions that can be perturbed in many ways, TFmiR helps to better elucidate cellular mechanisms at the molecular level from a network perspective. The provided topological and functional analyses promote TFmiR as a reliable systems biology tool for researchers across the life science communities. TFmiR web server is accessible through the following URL: http://service.bioinformatik.uni-saarland.de/tfmir.

  20. The Regulatory T Cell Lineage Factor Foxp3 Regulates Gene Expression through Several Distinct Mechanisms Mostly Independent of Direct DNA Binding

    PubMed Central

    Andersen, Kristian G.; Hebenstreit, Daniel; Teichmann, Sarah A.; Betz, Alexander G.

    2015-01-01

    The lineage factor Foxp3 is essential for the development and maintenance of regulatory T cells, but little is known about the mechanisms involved. Here, we demonstrate that an N-terminal proline-rich interaction region is crucial for Foxp3’s function. Subdomains within this key region link Foxp3 to several independent mechanisms of transcriptional regulation. Our study suggests that Foxp3, even in the absence of its DNA-binding forkhead domain, acts as a bridge between DNA-binding interaction partners and proteins with effector function permitting it to regulate a large number of genes. We show that, in one such mechanism, Foxp3 recruits class I histone deacetylases to the promoters of target genes, counteracting activation-induced histone acetylation and thereby suppressing their expression. PMID:26107960

  1. TFmiR: a web server for constructing and analyzing disease-specific transcription factor and miRNA co-regulatory networks.

    PubMed

    Hamed, Mohamed; Spaniol, Christian; Nazarieh, Maryam; Helms, Volkhard

    2015-07-01

    TFmiR is a freely available web server for deep and integrative analysis of combinatorial regulatory interactions between transcription factors, microRNAs and target genes that are involved in disease pathogenesis. Since the inner workings of cells rely on the correct functioning of an enormously complex system of activating and repressing interactions that can be perturbed in many ways, TFmiR helps to better elucidate cellular mechanisms at the molecular level from a network perspective. The provided topological and functional analyses promote TFmiR as a reliable systems biology tool for researchers across the life science communities. TFmiR web server is accessible through the following URL: http://service.bioinformatik.uni-saarland.de/tfmir. PMID:25943543

  2. Identification of temozolomide resistance factors in glioblastoma via integrative miRNA/mRNA regulatory network analysis.

    PubMed

    Hiddingh, Lotte; Raktoe, Rajiv S; Jeuken, Judith; Hulleman, Esther; Noske, David P; Kaspers, Gertjan J L; Vandertop, W Peter; Wesseling, Pieter; Wurdinger, Thomas

    2014-01-01

    Drug resistance is a major issue in the treatment of glioblastoma. Almost all glioblastomas are intrinsically resistant to chemotherapeutic temozolomide (TMZ) or develop resistance during treatment. The interaction networks of microRNAs (miRNAs) and mRNAs likely regulate most biological processes and can be employed to better understand complex processes including drug resistance in cancer. In this study, we examined if integrative miRNA/mRNA network analysis using the web-service tool mirConnX could be used to identify drug resistance factors in glioblastoma. We used TMZ-resistant glioblastoma cells and their integrated miRNA/mRNA networks to identify TMZ-sensitizing factors. TMZ resistance was previously induced in glioblastoma cell lines U87, Hs683, and LNZ308. miRNA/mRNA expression profiling of these cells and integration of the profiles using mirConnX resulted in the identification of plant homeodomain (PHD)-like finger 6 (PHF6) as a potential TMZ-sensitizing factor in resistant glioblastoma cells. Analysis of PHF6 expression showed significant upregulation in glioblastoma as compared to normal tissue. Interference with PHF6 expression in three TMZ-resistant subclones significantly enhanced TMZ-induced cell kill in two of these cell lines. Altogether, these results demonstrate that mirConnX is a feasible and useful tool to investigate miRNA/mRNA interactions in TMZ-resistant cells and has potential to identify drug resistance factors in glioblastoma.

  3. Identification of temozolomide resistance factors in glioblastoma via integrative miRNA/mRNA regulatory network analysis

    PubMed Central

    Hiddingh, Lotte; Raktoe, Rajiv S.; Jeuken, Judith; Hulleman, Esther; Noske, David P.; Kaspers, Gertjan J. L.; Vandertop, W. Peter; Wesseling, Pieter; Wurdinger, Thomas

    2014-01-01

    Drug resistance is a major issue in the treatment of glioblastoma. Almost all glioblastomas are intrinsically resistant to chemotherapeutic temozolomide (TMZ) or develop resistance during treatment. The interaction networks of microRNAs (miRNAs) and mRNAs likely regulate most biological processes and can be employed to better understand complex processes including drug resistance in cancer. In this study, we examined if integrative miRNA/mRNA network analysis using the web-service tool mirConnX could be used to identify drug resistance factors in glioblastoma. We used TMZ-resistant glioblastoma cells and their integrated miRNA/mRNA networks to identify TMZ-sensitizing factors. TMZ resistance was previously induced in glioblastoma cell lines U87, Hs683, and LNZ308. miRNA/mRNA expression profiling of these cells and integration of the profiles using mirConnX resulted in the identification of plant homeodomain (PHD)-like finger 6 (PHF6) as a potential TMZ-sensitizing factor in resistant glioblastoma cells. Analysis of PHF6 expression showed significant upregulation in glioblastoma as compared to normal tissue. Interference with PHF6 expression in three TMZ-resistant subclones significantly enhanced TMZ-induced cell kill in two of these cell lines. Altogether, these results demonstrate that mirConnX is a feasible and useful tool to investigate miRNA/mRNA interactions in TMZ-resistant cells and has potential to identify drug resistance factors in glioblastoma. PMID:24919120

  4. CONREAL: conserved regulatory elements anchored alignment algorithm for identification of transcription factor binding sites by phylogenetic footprinting.

    PubMed

    Berezikov, Eugene; Guryev, Victor; Plasterk, Ronald H A; Cuppen, Edwin

    2004-01-01

    Prediction of transcription-factor target sites in promoters remains difficult due to the short length and degeneracy of the target sequences. Although the use of orthologous sequences and phylogenetic footprinting approaches may help in the recognition of conserved and potentially functional sequences, correct alignment of the short transcription-factor binding sites can be problematic for established algorithms, especially when aligning more divergent species. Here, we report a novel phylogenetic footprinting approach, CONREAL, that uses biologically relevant information, that is, potential transcription-factor binding sites as represented by positional weight matrices, to establish anchors between orthologous sequences and to guide promoter sequence alignment. Comparison of the performance of CONREAL with the global alignment programs LAGAN and AVID using a reference data set, shows that CONREAL performs equally well for closely related species like rodents and human, and has a clear added value for aligning promoter elements of more divergent species like human and fish, as it identifies conserved transcription-factor binding sites that are not found by other methods. CONREAL is accessible via a Web interface at http://conreal.niob.knaw.nl/.

  5. An Additional Potential Factor for Kidney Stone Formation during Space Flights: Calcifying Nanoparticles (Nanobacteria): A Case Report

    NASA Technical Reports Server (NTRS)

    Jones, Jeffrey A.; Ciftcioglu, Neva; Schmid, Joseph; Griffith, Donald

    2007-01-01

    Spaceflight-induced microgravity appears to be a risk factor for the development of urinary calculi due to skeletal calcium liberation and other undefined factors, resulting in stone disease in crewmembers during and after spaceflight. Calcifying nanoparticles, or nanobacteria, reproduce at a more rapid rate in simulated microgravity conditions and create external shells of calcium phosphate in the form of apatite. The questions arises whether calcifying nanoparticles are niduses for calculi and contribute to the development of clinical stone disease in humans, who possess environmental factors predisposing to the development of urinary calculi and potentially impaired immunological defenses during spaceflight. A case of a urinary calculus passed from an astronaut post-flight with morphological characteristics of calcifying nanoparticles and staining positive for a calcifying nanoparticle unique antigen, is presented.

  6. Compound mouse mutants of bZIP transcription factors Mafg and Mafk reveal a regulatory network of non-crystallin genes associated with cataract.

    PubMed

    Agrawal, Smriti A; Anand, Deepti; Siddam, Archana D; Kakrana, Atul; Dash, Soma; Scheiblin, David A; Dang, Christine A; Terrell, Anne M; Waters, Stephanie M; Singh, Abhyudai; Motohashi, Hozumi; Yamamoto, Masayuki; Lachke, Salil A

    2015-07-01

    Although majority of the genes linked to early-onset cataract exhibit lens fiber cell-enriched expression, our understanding of gene regulation in these cells is limited to function of just eight transcription factors and largely in the context of crystallins. We report on small Maf transcription factors Mafg and Mafk as regulators of several non-crystallin human cataract-associated genes in fiber cells and establish their significance to this disease. We applied a bioinformatics tool for cataract gene discovery iSyTE to identify Mafg and its co-regulators in the lens, and generated various null-allelic combinations of Mafg:Mafk mouse mutants for phenotypic and molecular analysis. By age 4 months, Mafg-/-:Mafk+/- mutants exhibit lens defects that progressively develop into cataract. High-resolution phenotypic characterization of Mafg-/-:Mafk+/- mouse lens reveals severely disorganized fiber cells, while microarray-based expression profiling identifies 97 differentially regulated genes (DRGs). Integrative analysis of Mafg-/-:Mafk+/- lens-DRGs with (1) binding motifs and genomic targets of small Mafs and their regulatory partners, (2) iSyTE lens expression data, and (3) interactions between DRGs in the String database, unravel a detailed small Maf regulatory network in the lens, several nodes of which are linked to cataract. This approach identifies 36 high-priority candidates from the original 97 DRGs. Significantly, 8/36 (22%) DRGs are associated with cataracts in human (GSTO1, MGST1, SC4MOL, UCHL1) or mouse (Aldh3a1, Crygf, Hspb1, Pcbd1), suggesting a multifactorial etiology that includes oxidative stress and misregulation of sterol synthesis. These data identify Mafg and Mafk as new cataract-associated candidates and define their function in regulating largely non-crystallin genes linked to human cataract. PMID:25896808

  7. Suppression of the pancreatic duodenal homeodomain transcription factor-1 (Pdx-1) promoter by sterol regulatory element-binding protein-1c (SREBP-1c).

    PubMed

    Amemiya-Kudo, Michiyo; Oka, Junko; Takeuchi, Yoshinori; Okazaki, Hiroaki; Yamamoto, Takashi; Yahagi, Naoya; Matsuzaka, Kaori; Okazaki, Sachiko; Osuga, Jun-ichi; Yamada, Nobuhiro; Murase, Toshio; Shimano, Hitoshi

    2011-08-12

    Overexpression of sterol regulatory element-binding protein-1c (SREBP-1c) in β cells causes impaired insulin secretion and β cell dysfunction associated with diminished pancreatic duodenal homeodomain transcription factor-1 (PDX-1) expression in vitro and in vivo. To identify the molecular mechanism responsible for this effect, the mouse Pdx-1 gene promoter (2.7 kb) was analyzed in β cell and non-β cell lines. Despite no apparent sterol regulatory element-binding protein-binding sites, the Pdx-1 promoter was suppressed by SREBP-1c in β cells in a dose-dependent manner. PDX-1 activated its own promoter. The E-box (-104/-99 bp) in the proximal region, occupied by ubiquitously expressed upstream stimulatory factors (USFs), was crucial for the PDX-1-positive autoregulatory loop through direct PDX-1·USF binding. This positive feedback activation was a prerequisite for SREBP-1c suppression of the promoter in non-β cells. SREBP-1c and PDX-1 directly interact through basic helix-loop-helix and homeobox domains, respectively. This robust SREBP-1c·PDX-1 complex interferes with PDX-1·USF formation and inhibits the recruitment of PDX-1 coactivators. SREBP-1c also inhibits PDX-1 binding to the previously described PDX-1-binding site (-2721/-2646 bp) in the distal enhancer region of the Pdx-1 promoter. Endogenous up-regulation of SREBP-1c in INS-1 cells through the activation of liver X receptor and retinoid X receptor by 9-cis-retinoic acid and 22-hydroxycholesterol inhibited PDX-1 mRNA and protein expression. Conversely, SREBP-1c RNAi restored Pdx-1 mRNA and protein levels. Through these multiple mechanisms, SREBP-1c, when induced in a lipotoxic state, repressed PDX-1 expression contributing to the inhibition of insulin expression and β cell dysfunction.

  8. Intra-articular nuclear factor-κB blockade ameliorates collagen-induced arthritis in mice by eliciting regulatory T cells and macrophages

    PubMed Central

    Min, S-Y; Yan, M; Du, Y; Wu, T; Khobahy, E; Kwon, S-R; Taneja, V; Bashmakov, A; Nukala, S; Ye, Y; Orme, J; Sajitharan, D; Kim, H-Y; Mohan, C

    2013-01-01

    Nuclear factor (NF)-κB is a transcription factor implicated in the pathogenesis of autoimmune disorders such as rheumatoid arthritis (RA). Here we have examined the effect of intra-articular administration of the IKK inhibitor, NEMO-binding domain peptide (NBD), on the severity of collagen-induced arthritis (CIA). NBD peptides were injected intra-articularly into the knee joints of DBA/1J mice after the onset of disease. Collagen-injected mice given a scrambled peptide served as controls. Arthritis severity was determined by visual examination of paws. Intra-articular NBD injection reduced the arthritis score and ameliorated morphological signs of bone destruction compared to the controls. Serum levels of type-II collagen-specific immunoglobulin (Ig)G2a antibodies were lower in NBD-treated mice versus the control mice, whereas the levels of type-II collagen-specific IgG1 antibodies were increased by NBD treatment. NBD treatment diminished the proinflammatory cytokines interleukin (IL)-17 and interferon (IFN)-γ in serum, but increased the regulatory cytokine IL-10. NBD-treated CIA mice exhibited significantly higher percentages and numbers of forkhead box protein 3 (FoxP3+)CD4+CD25+ regulatory T cells than controls. Immunofluorescence analysis of NBD-treated mice revealed that FoxP3 and Ym1, a marker of alternatively activated macrophages, were juxtaposed to each other within draining inguinal lymph nodes. Intra-articular administration of NBD peptide is effective as an experimental therapy in a murine model of RA. Nevertheless, the intra-articular treatment modality is still associated with systemic effects on the immune system. PMID:23574318

  9. The role of internal and external factors in the implementation of disruptive court orders by the Federal Energy Regulatory Commission

    NASA Astrophysics Data System (ADS)

    Nagler, Peter Andrew

    A court, ordering an administrative agency to alter its practices, begins a sequence of events that may involve not only the agency but also others in its environment. Two main questions are considered in this research. The first is how does the agency actually respond to this disturbance: what does it do and why? The second is what role, if any, do influential outsiders including members of Congress, representatives of interest groups and other officials from the executive branch have in shaping the agency response to the court order? This study concentrates on the Federal Energy Regulatory Commission (FERC) and its natural gas regulations. Five court orders, issued by the District of Columbia and Fifth Circuits during the Reagan administration (1981--89), are examined in separate case studies. Five independent variables, agency commitment to its existing program, the disruptiveness of the court order, the likelihood and severity of penalties for noncompliance with the court order, agency views of the court, and external pressures on the agency, are analyzed with respect to their relative influence on the dependent variable of agency response to the court. The analysis is qualitative, marshalling facts and evidence gathered from several major data sources: FERC orders, policy statements and newsletters, trade press accounts, congressional hearings, and personal and telephone interviews conducted by the author with FERC staff and commissioners, interest group representatives, and congressional staff. The research shows that the FERC generally implemented the court orders although some evasion due at least partly to the Commission's commitment to its programs was found in four of the five cases. Agency view of the court as a legitimate authority to be obeyed was a major counter to any tendencies to evade the judicial mandates. External pressure was important only in those cases with widespread impact on the industry while disruptiveness became significant when

  10. Employing Lead Thiocyanate Additive to Reduce the Hysteresis and Boost the Fill Factor of Planar Perovskite Solar Cells.

    PubMed

    Ke, Weijun; Xiao, Chuanxiao; Wang, Changlei; Saparov, Bayrammurad; Duan, Hsin-Sheng; Zhao, Dewei; Xiao, Zewen; Schulz, Philip; Harvey, Steven P; Liao, Weiqiang; Meng, Weiwei; Yu, Yue; Cimaroli, Alexander J; Jiang, Chun-Sheng; Zhu, Kai; Al-Jassim, Mowafak; Fang, Guojia; Mitzi, David B; Yan, Yanfa

    2016-07-01

    Lead thiocyanate in the perovskite precursor can increase the grain size of a perovskite thin film and reduce the conductivity of the grain boundaries, leading to perovskite solar cells with reduced hysteresis and enhanced fill factor. A planar perovskite solar cell with grain boundary and interface passivation achieves a steady-state efficiency of 18.42%.

  11. CArG boxes in the human cardiac. cap alpha. -actin gene are core binding sites for positive trans-acting regulatory factors

    SciTech Connect

    Miwa, T.; Boxer, L.M.; Kedes, L.

    1987-10-01

    Positively acting, rate-limiting regulatory factors that influence tissue-specific expression of the human cardiac ..cap alpha..-actin gene in a mouse muscle cell line are shown by in vivo competition and gel mobility-shift assays to bind to upstream regions of its promoter but to neither vector DNA not a ..beta..-globin promoter. Although the two binding regions are distinctly separated, each corresponds to a cis region required for muscle-specific transcriptional stimulation, and each contains a core CC(A+T-rich)/sub 6/GC sequence (designated CArG box), which is found in the promoter regions of several muscle-associated genes. Each site has an apparently different binding affinity for trans-acting factors, which may explain the different transcriptional stimulation activities of the two cis regions. Therefore, the authors conclude that the two CArG box regions are responsible for muscle-specific transcriptional activity of the cardiac ..cap alpha..-actin gene through a mechanism that involves their binding of a positive trans-acting factor in muscle cells.

  12. A conserved alternative form of the purple sea urchin HEB/E2-2/E2A transcription factor mediates a switch in E-protein regulatory state in differentiating immune cells.

    PubMed

    Schrankel, Catherine S; Solek, Cynthia M; Buckley, Katherine M; Anderson, Michele K; Rast, Jonathan P

    2016-08-01

    E-proteins are basic helix-loop-helix (bHLH) transcription factors with essential roles in animal development. In mammals, these are encoded by three loci: E2-2 (ITF-2/ME2/SEF2/TCF4), E2A (TCF3), and HEB (ME1/REB/TCF12). The HEB and E2-2 paralogs are expressed as alternative (Alt) isoforms with distinct N-terminal sequences encoded by unique exons under separate regulatory control. Expression of these alternative transcripts is restricted relative to the longer (Can) forms, suggesting distinct regulatory roles, although the functions of the Alt proteins remain poorly understood. Here, we characterize the single sea urchin E-protein ortholog (SpE-protein). The organization of the SpE-protein gene closely resembles that of the extended HEB/E2-2 vertebrate loci, including a transcript that initiates at a homologous alternative transcription start site (SpE-Alt). The existence of an Alt form in the sea urchin indicates that this feature predates the emergence of the vertebrates. We present additional evidence indicating that this transcript was present in the common bilaterian ancestor. In contrast to the widely expressed canonical form (SpE-Can), SpE-Alt expression is tightly restricted. SpE-Alt is expressed in two phases: first in aboral non-skeletogenic mesenchyme (NSM) cells and then in oral NSM cells preceding their differentiation and ingression into the blastocoel. Derivatives of these cells mediate immune response in the larval stage. Inhibition of SpE-Alt activity interferes with these events. Notably, although the two isoforms are initially co-expressed, as these cells differentiate, SpE-Can is excluded from the SpE-Alt(+) cell population. This mutually exclusive expression is dependent on SpE-Alt function, which reveals a previously undescribed negative regulatory linkage between the two E-protein forms. Collectively, these findings reorient our understanding of the evolution of this transcription factor family and highlight fundamental properties of E

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

    PubMed

    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

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

    PubMed Central

    Dai, Qi; Ren, Aiming; Westholm, Jakub O.; Duan, Hong; Patel, Dinshaw J.

    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

  15. Factors affecting the microbial and chemical composition of silage. III. Effect of urea additions on maize silage.

    PubMed

    Mahmoud, S A; Abd-el-Hafez, A; Zaki, M M; Saleh, E A

    1978-01-01

    The effect of urea additions on the microbiological and chemical properties of silage, produced from young maize plants (Darawa stage), was studied. Urea treatments, i.e., 0.25%, 0.50%, 0.75%, and 1.00%, stimulated higher densities of the desired microorganisms than the control, while undesired organisms showed lower counts (proteolytic and saccharolytic anaerobes). Addition of 0.25 to 0.50% or urea resulted in the production of high quality silage with pleasant small and high nutritive value, as confirmed by the various microbiological and chemical analyses conducted. Higher levels (0.75 and 1.00%) of urea decreased the quality of the product. PMID:29417

  16. Zinc-rich inhibitor of apoptosis proteins (IAPs) as regulatory factors in the epithelium of normal and inflamed airways.

    PubMed

    Roscioli, Eugene; Hamon, Rhys; Lester, Susan; Murgia, Chiara; Grant, Janet; Zalewski, Peter

    2013-04-01

    Integrity of the airway epithelium (AE) is important in the context of inhaled allergens and noxious substances, particularly during asthma-related airway inflammation where there is increased vulnerability of the AE to cell death. Apoptosis involves a number of signaling pathways which activate procaspases leading to cleavage of critical substrates. Understanding the factors which regulate AE caspases is important for development of strategies to minimize AE damage and airway inflammation, and therefore to better control asthma. One such factor is the essential dietary metal zinc. Zinc deficiency results in enhanced AE apoptosis, and worsened airway inflammation. This has implications for asthma, where abnormalities in zinc homeostasis have been observed. Zinc is thought to suppress the steps involved in caspase-3 activation. One target of zinc is the family of inhibitor of apoptosis proteins (IAPs) which are endogenous regulators of caspases. More studies are needed to identify the roles of IAPs in regulating apoptosis in normal and inflamed airways and to study their interaction with labile zinc ions. This new information will provide a framework for future clinical studies aimed at monitoring and management of airway zinc levels as well as minimising airway damage and inflammation in asthma.

  17. c-Fos: an AP-1 transcription factor with an additional cytoplasmic, non-genomic lipid synthesis activation capacity.

    PubMed

    Caputto, Beatriz L; Cardozo Gizzi, Andrés M; Gil, Germán A

    2014-09-01

    The mechanisms that co-ordinately activate lipid synthesis when high rates of membrane biogenesis are needed to support cell growth are largely unknown. c-Fos, a well known AP-1 transcription factor, has emerged as a unique protein with the capacity to associate to specific enzymes of the pathway of synthesis of phospholipids at the endoplasmic reticulum and activate their synthesis to accompany genomic decisions of growth. Herein, we discuss this cytoplasmic, non-genomic effect of c-Fos in the context of other mechanisms that have been proposed to regulate lipid synthesis.

  18. Decay-accelerating factor (CD55), a glycosylphosphatidylinositol-anchored complement regulatory protein, is a receptor for several echoviruses.

    PubMed Central

    Bergelson, J M; Chan, M; Solomon, K R; St John, N F; Lin, H; Finberg, R W

    1994-01-01

    Echoviruses are human pathogens belonging to the picornavirus family. Decay-accelerating factor (DAF) is a glycosylphosphatidylinositol (GPI)-anchored surface protein that protects cells from lysis by autologous complement. Anti-DAF monoclonal antibodies prevented echovirus 7 attachment to susceptible cells and protected cells from infection. HeLa cells specifically lost the capacity to bind echovirus 7 when treated with phosphatidylinositol-specific phospholipase C, an enzyme that releases GPI-anchored proteins from the cell surface, indicating that the virus receptor, like DAF, is a GPI-anchored protein. Although Chinese hamster ovary cells do not bind echovirus 7, transfectants expressing human DAF bound virus efficiently, and binding was prevented by pretreatment with an anti-DAF monoclonal antibody. Anti-DAF antibodies prevented infection by at least six echovirus serotypes. These results indicate that DAF is the receptor mediating attachment and infection by several echoviruses. Images PMID:7517044

  19. Interferon regulatory factor 5 (IRF5) gene variants are associated with multiple sclerosis in three distinct populations

    PubMed Central

    Kristjansdottir, G; Sandling, J K; Bonetti, A; Roos, I M; Milani, L; Wang, C; Gustafsdottir, S M; Sigurdsson, S; Lundmark, A; Tienari, P J; Koivisto, K; Elovaara, I; Pirttilä, T; Reunanen, M; Peltonen, L; Saarela, J; Hillert, J; Olsson, T; Landegren, U; Alcina, A; Fernández, O; Leyva, L; Guerrero, M; Lucas, M; Izquierdo, G; Matesanz, F; Syvänen, A-C

    2008-01-01

    Background: IRF5 is a transcription factor involved both in the type I interferon and the toll-like receptor signalling pathways. Previously, IRF5 has been found to be associated with systemic lupus erythematosus, rheumatoid arthritis and inflammatory bowel diseases. Here we investigated whether polymorphisms in the IRF5 gene would be associated with yet another disease with features of autoimmunity, multiple sclerosis (MS). Methods: We genotyped nine single nucleotide polymorphisms and one insertion-deletion polymorphism in the IRF5 gene in a collection of 2337 patients with MS and 2813 controls from three populations: two case–control cohorts from Spain and Sweden, and a set of MS trio families from Finland. Results: Two single nucleotide polymorphism (SNPs) (rs4728142, rs3807306), and a 5 bp insertion-deletion polymorphism located in the promoter and first intron of the IRF5 gene, showed association signals with values of p<0.001 when the data from all cohorts were combined. The predisposing alleles were present on the same common haplotype in all populations. Using electrophoretic mobility shift assays we observed allele specific differences in protein binding for the SNP rs4728142 and the 5 bp indel, and by a proximity ligation assay we demonstrated increased binding of the transcription factor SP1 to the risk allele of the 5 bp indel. Conclusion: These findings add IRF5 to the short list of genes shown to be associated with MS in more than one population. Our study adds to the evidence that there might be genes or pathways that are common in multiple autoimmune diseases, and that the type I interferon system is likely to be involved in the development of these diseases. PMID:18285424

  20. The CarD/CarG regulatory complex is required for the action of several members of the large set of Myxococcus xanthus extracytoplasmic function σ factors.

    PubMed

    Abellón-Ruiz, Javier; Bernal-Bernal, Diego; Abellán, María; Fontes, Marta; Padmanabhan, S; Murillo, Francisco J; Elías-Arnanz, Montserrat

    2014-08-01

    Extracytoplasmic function (ECF) σ factors are critical players in signal transduction networks involved in bacterial response to environmental changes. The Myxococcus xanthus genome reveals ∼45 putative ECF-σ factors, but for the overwhelming majority, the specific signals or mechanisms for selective activation and regulation remain unknown. One well-studied ECF-σ, CarQ, binds to its anti-σ, CarR, and is inactive in the dark but drives its own expression from promoter P(QRS) on illumination. This requires the CarD/CarG complex, the integration host factor (IHF) and a specific CarD-binding site upstream of P(QRS). Here, we show that DdvS, a previously uncharacterized ECF-σ, activates its own expression in a CarD/CarG-dependent manner but is inhibited when specifically bound to the N-terminal zinc-binding anti-σ domain of its cognate anti-σ, DdvA. Interestingly, we find that the autoregulatory action of 11 other ECF-σ factors studied here depends totally or partially on CarD/CarG but not IHF. In silico analysis revealed possible CarD-binding sites that may be involved in direct regulation by CarD/CarG of target promoter activity. CarD/CarG-linked ECF-σ regulation likely recurs in other myxobacteria with CarD/CarG orthologous pairs and could underlie, at least in part, the global regulatory effect of the complex on M. xanthus gene expression.

  1. Mutational analysis of the [4Fe-4S]-cluster converting iron regulatory factor from its RNA-binding form to cytoplasmic aconitase.

    PubMed Central

    Hirling, H; Henderson, B R; Kühn, L C

    1994-01-01

    The control of cellular iron homeostasis involves the coordinate post-transcriptional regulation of ferritin mRNA translation and transferring receptor mRNA stability. These regulatory events are mediated by a soluble cytoplasmic protein, iron regulatory factor (IRF), which binds specifically to mRNA hairpin structures, termed iron-responsive elements (IREs), in the respective mRNAs. IRF is modulated by variations of cellular iron levels and exists as either an apo-protein or a [4Fe-4S]-cluster protein. The two conformations show distinct, mutually exclusive functions. High-affinity IRE binding is observed with the apo-form induced by iron deprivation, but is lost under high iron conditions when IRF is converted to the [4Fe-4S]-cluster form which shows cytoplasmic aconitase activity. Moreover, IRE binding is inactivated by the sulfhydryl-oxidizing agent diamide and fully activated in vitro by 2% 2-mercapto-ethanol, whereas alkylation of IRF inhibits IRE binding. In the present study, we analyzed each of the above features using site-directed mutants of recombinant human IRF. The results support the bifunctional nature of IRF. We conclude that cysteines 437, 503 and 506 anchor the [4Fe-4S]-cluster, and are essential to the aconitase activity. Mutagenesis changing any of the cysteines to serine leads to constitutive RNA binding in 0.02% 2-mercaptoethanol. Cysteine 437 is particularly critical to the RNA-protein interaction. The spontaneous or diamide-induced formation of disulfide bonds between cysteines 437 and 503 or 437 and 506, in apo-IRF, as well as its alkylation by N-ethylmaleimide, inhibit binding to the IRE.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:7508861

  2. Ubiquitin-associated Domain-containing Ubiquitin Regulatory X (UBX) Protein UBXN1 Is a Negative Regulator of Nuclear Factor κB (NF-κB) Signaling*

    PubMed Central

    Wang, Yu-Bo; Tan, Bo; Mu, Rui; Chang, Yan; Wu, Min; Tu, Hai-Qing; Zhang, Yu-Cheng; Guo, Sai-Sai; Qin, Xuan-He; Li, Tao; Li, Wei-Hua; Li, Ai-Ling; Zhang, Xue-Min; Li, Hui-Yan

    2015-01-01

    Excessive nuclear factor κB (NF-κB) activation should be precisely controlled as it contributes to multiple immune and inflammatory diseases. However, the negative regulatory mechanisms of NF-κB activation still need to be elucidated. Various types of polyubiquitin chains have proved to be involved in the process of NF-κB activation. Many negative regulators linked to ubiquitination, such as A20 and CYLD, inhibit IκB kinase activation in the NF-κB signaling pathway. To find new NF-κB signaling regulators linked to ubiquitination, we used a small scale siRNA library against 51 ubiquitin-associated domain-containing proteins and screened out UBXN1, which contained both ubiquitin-associated and ubiquitin regulatory X (UBX) domains as a negative regulator of TNFα-triggered NF-κB activation. Overexpression of UBXN1 inhibited TNFα-triggered NF-κB activation, although knockdown of UBXN1 had the opposite effect. UBX domain-containing proteins usually act as valosin-containing protein (VCP)/p97 cofactors. However, knockdown of VCP/p97 barely affected UBXN1-mediated NF-κB inhibition. At the same time, we found that UBXN1 interacted with cellular inhibitors of apoptosis proteins (cIAPs), E3 ubiquitin ligases of RIP1 in the TNFα receptor complex. UBXN1 competitively bound to cIAP1, blocked cIAP1 recruitment to TNFR1, and sequentially inhibited RIP1 polyubiquitination in response to TNFα. Therefore, our findings demonstrate that UBXN1 is an important negative regulator of the TNFα-triggered NF-κB signaling pathway by mediating cIAP recruitment independent of VCP/p97. PMID:25681446

  3. Genome-wide characterization and comparative analysis of R2R3-MYB transcription factors shows the complexity of MYB-associated regulatory networks in Salvia miltiorrhiza

    PubMed Central

    2014-01-01

    Background MYB is the largest plant transcription factor gene family playing vital roles in plant growth and development. However, it has not been systematically studied in Salvia miltiorrhiza, an economically important medicinal plant. Results Here we report the genome-wide identification and characterization of 110 R2R3-MYBs, the largest subfamily of MYBs in S. miltiorrhiza. The MYB domain and other motifs of SmMYBs are largely conserved with Arabidopsis AtMYBs, whereas the divergence of SmMYBs and AtMYBs also exists, suggesting the conservation and diversity of plant MYBs. SmMYBs and AtMYBs may be classified into 37 subgroups, of which 31 include proteins from S. miltiorrhiza and Arabidopsis, whereas 6 are specific to a species, indicating that the majority of MYBs play conserved roles, while others may exhibit species-specialized functions. SmMYBs are differentially expressed in various tissues of S. miltiorrhiza. The expression profiles are largely consistent with known functions of their Arabidopsis counterparts. The expression of a subset of SmMYBs is regulated by microRNAs, such as miR159, miR319, miR828 and miR858. Based on functional conservation of MYBs in a subgroup, SmMYBs potentially involved in the biosynthesis of bioactive compounds were identified. Conclusions A total of 110 R2R3-MYBs were identified and analyzed. The results suggest the complexity of MYB-mediated regulatory networks in S. miltiorrhiza and provide a foundation for understanding the regulatory mechanism of SmMYBs. PMID:24725266

  4. Fas-Associated Factor 1 Negatively Regulates the Antiviral Immune Response by Inhibiting Translocation of Interferon Regulatory Factor 3 to the Nucleus

    PubMed Central

    Song, Soonhwa; Lee, Jae-Jin; Kim, Hee-Jung; Lee, Jeong Yoon; Chang, Jun

    2016-01-01

    This study is designed to examine the cellular functions of human Fas-associated factor 1 (FAF1) containing multiple ubiquitin-related domains. Microarray analyses revealed that interferon-stimulated genes related to the antiviral response are significantly increased in FAF1-knockdown HeLa cells. Silencing FAF1 enhanced the poly(I·C)- and respiratory syncytial virus (RSV)-induced production of type I interferons (IFNs), the target genes of interferon regulator factor 3 (IRF3). IRF3 is a key transcription factor in IFN-β signaling responsible for the host innate immune response. This study also found that FAF1 and IRF3 physically associate with IPO5/importin-β3 and that overexpression of FAF1 reduces the interaction between IRF3 and IPO5/importin-β3. These findings suggest that FAF1 negatively regulates IRF3-mediated IFN-β production and the antiviral innate immune response by regulating nuclear translocation of IRF3. We conclude that FAF1 plays a novel role in negatively regulating virus-induced IFN-β production and the antiviral response by inhibiting the translocation of active, phosphorylated IRF3 from the cytosol to the nucleus. PMID:26811330

  5. Murine elongation factor 1 alpha (EF-1 alpha) is posttranslationally modified by novel amide-linked ethanolamine-phosphoglycerol moieties. Addition of ethanolamine-phosphoglycerol to specific glutamic acid residues on EF-1 alpha

    SciTech Connect

    Whiteheart, S.W.; Shenbagamurthi, P.; Chen, L.; Cotter, R.J.; Hart, G.W. )

    1989-08-25

    Elongation Factor 1 alpha (EF-1 alpha), an important eukaryotic translation factor, transports charged aminoacyl-tRNA from the cytosol to the ribosomes during poly-peptide synthesis. Metabolic radiolabeling with ({sup 3}H) ethanolamine shows that, in all cells examined, EF-1 alpha is the major radiolabeled protein. Radiolabeled EF-1 alpha has an apparent Mr = 53,000 and a basic isoelectric point. It is cytosolic and does not contain N-linked oligosaccharides. Trypsin digestion of murine EF-1 alpha generated two major ({sup 3}H)ethanolamine-labeled peptides. Three peptides were sequenced and were identical to two distinct regions of the human EF-1 alpha protein. Blank sequencing cycles coinciding with glutamic acid in the human cDNA-derived sequence were also found to release ({sup 3}H)ethanolamine, and compositional analysis of these peptides confirmed the presence of glutamic acid. Dansylation analysis demonstrates that the amine group of the ethanolamine is blocked. These results indicate that EF-1 alpha is posttranslationally modified by the covalent attachment of ethanolamine via an amide bond to at least two specific glutamic acid residues (Glu-301 and Glu-374). The hydroxyl group of the attached ethanolamine was shown by mass spectrometry and compositional analysis, to be further modified by the addition of a phosphoglycerol unit. This novel posttranslational modification may represent an important alteration of EF-1 alpha, comparable to the regulatory effects of posttranslational methylation of EF-1 alpha lysine residues.

  6. 42 CFR 136.408 - What are other factors, in addition to the minimum standards of character, that may be considered...

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 42 Public Health 1 2012-10-01 2012-10-01 false What are other factors, in addition to the minimum standards of character, that may be considered in determining placement of an individual in a position that involves regular contact with or control over Indian children? 136.408 Section 136.408 Public Health PUBLIC HEALTH SERVICE, DEPARTMENT...

  7. 42 CFR 136.408 - What are other factors, in addition to the minimum standards of character, that may be considered...

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 42 Public Health 1 2014-10-01 2014-10-01 false What are other factors, in addition to the minimum standards of character, that may be considered in determining placement of an individual in a position that involves regular contact with or control over Indian children? 136.408 Section 136.408 Public Health PUBLIC HEALTH SERVICE, DEPARTMENT...

  8. Ameliorative effects of telmisartan on the inflammatory response and impaired spatial memory in a rat model of Alzheimer's disease incorporating additional cerebrovascular disease factors.

    PubMed

    Shindo, Taro; Takasaki, Kotaro; Uchida, Kanako; Onimura, Rika; Kubota, Kaori; Uchida, Naoki; Irie, Keiichi; Katsurabayashi, Shutaro; Mishima, Kenichi; Nishimura, Ryoji; Fujiwara, Michihiro; Iwasaki, Katsunori

    2012-01-01

    Telmisartan, an angiotensin type 1 receptor blocker, is used in the management of hypertension to control blood pressure. In addition, telmisartan has a partial agonistic effect on peroxisome proliferator activated receptor γ (PPARγ). Recently, the effects of telmisartan on spatial memory or the inflammatory response were monitored in a mouse model of Alzheimer's disease (AD). However, to date, no studies have investigated the ameliorative effects of telmisartan on impaired spatial memory and the inflammatory response in an AD animal model incorporating additional cerebrovascular disease factors. In this study, we examined the effect of telmisartan on spatial memory impairment and the inflammatory response in a rat model of AD incorporating additional cerebrovascular disease factors. Rats were subjected to cerebral ischemia and an intracerebroventricular injection of oligomeric or aggregated amyloid-β (Aβ). Oral administration of telmisartan (0.3, 1, 3 mg/kg/d) seven days after ischemia and Aβ treatment resulted in better performance in the eight arm radial maze task in a dose-dependent manner. Telmisartan also reduced tumor necrosis factor α mRNA expression in the hippocampal region of rats with impaired spatial memory. These effects of telmisartan were antagonized by GW9662, an antagonist of PPARγ. These results suggest that telmisartan has ameliorative effects on the impairment of spatial memory in a rat model of AD incorporating additional cerebrovascular disease factors via its anti-inflammatory effect.

  9. Breeding site selection by coho salmon (Oncorhynchus kisutch) in relation to large wood additions and factors that influence reproductive success

    USGS Publications Warehouse

    Clark, Steven M.; Dunham, Jason B.; McEnroe, Jeffery R.; Lightcap, Scott W.

    2014-01-01

    The fitness of female Pacific salmon (Oncorhynchus spp.) with respect to breeding behavior can be partitioned into at least four fitness components: survival to reproduction, competition for breeding sites, success of egg incubation, and suitability of the local environment near breeding sites for early rearing of juveniles. We evaluated the relative influences of habitat features linked to these fitness components with respect to selection of breeding sites by coho salmon (Oncorhynchus kisutch). We also evaluated associations between breeding site selection and additions of large wood, as the latter were introduced into the study system as a means of restoring habitat conditions to benefit coho salmon. We used a model selection approach to organize specific habitat features into groupings reflecting fitness components and influences of large wood. Results of this work suggest that female coho salmon likely select breeding sites based on a wide range of habitat features linked to all four hypothesized fitness components. More specifically, model parameter estimates indicated that breeding site selection was most strongly influenced by proximity to pool-tail crests and deeper water (mean and maximum depths). Linkages between large wood and breeding site selection were less clear. Overall, our findings suggest that breeding site selection by coho salmon is influenced by a suite of fitness components in addition to the egg incubation environment, which has been the emphasis of much work in the past.

  10. Structural basis for the requirement of additional factors for MLL1 SET domain activity and recognition of epigenetic marks.

    PubMed

    Southall, Stacey M; Wong, Poon-Sheng; Odho, Zain; Roe, S Mark; Wilson, Jon R

    2009-01-30

    The mixed-lineage leukemia protein MLL1 is a transcriptional regulator with an essential role in early development and hematopoiesis. The biological function of MLL1 is mediated by the histone H3K4 methyltransferase activity of the carboxyl-terminal SET domain. We have determined the crystal structure of the MLL1 SET domain in complex with cofactor product AdoHcy and a histone H3 peptide. This structure indicates that, in order to form a well-ordered active site, a highly variable but essential component of the SET domain must be repositioned. To test this idea, we compared the effect of the addition of MLL complex members on methyltransferase activity and show that both RbBP5 and Ash2L but not Wdr5 stimulate activity. Additionally, we have determined the effect of posttranslational modifications on histone H3 residues downstream and upstream from the target lysine and provide a structural explanation for why H3T3 phosphorylation and H3K9 acetylation regulate activity. PMID:19187761

  11. Epidermal growth factor (EGF)-receptor is phosphorylated at threonine-654 in A431 cells following EGF addition

    SciTech Connect

    Whiteley, B.; Glaser, L.

    1986-05-01

    It has been shown that activation of protein kinase C by tumor-promoting phorbol diesters causes phorphorylation of the EGF-receptor at threonine-654 and is believed to thereby regulate the EGF receptor tyrosine kinase and EGF binding activity. In their present studies, /sup 32/P-labeled A431 cells were treated with and without 10 nM phorbol 12-myristate 13-acetate (PMA), or with 200 ng/ml EGF. Analysis of /sup 32/P-labeled EGF receptor tryptic phosphopeptides by reverse-phase HPLC confirmed the known effects of PMA and revealed that EGF caused phosphorylation at threonine-654 as well as various tyrosine residues. This effect occurred as early as 1 minute after EGF addition and was maximal after 5 minutes. The magnitude of the response appears to be 50% of a 15 minute treatment with 10 nM PMA. Direct measurement of diacylglycerol using an E. coli diacylglycerol kinase confirmed that EGF-stimulated phosphoinositide turnover could cause very rapid activation of protein kinase C. These results imply that protein kinase C is playing a role in negative modulation of EGF-receptor activity following EGF addition to A431 cells.

  12. The Third Intron of the Interferon Regulatory Factor-8 Is an Initiator of Repressed Chromatin Restricting Its Expression in Non-Immune Cells

    PubMed Central

    Barnea-Yizhar, Ofer; Ram, Sigal; Kovalev, Ekaterina; Azriel, Aviva; Rand, Ulfert; Nakayama, Manabu; Hauser, Hansjörg; Gepstein, Lior; Levi, Ben-Zion

    2016-01-01

    Interferon Regulatory Factor-8 (IRF-8) serves as a key factor in the hierarchical differentiation towards monocyte/dendritic cell lineages. While much insight has been accumulated into the mechanisms essential for its hematopoietic specific expression, the mode of restricting IRF-8 expression in non-hematopoietic cells is still unknown. Here we show that the repression of IRF-8 expression in restrictive cells is mediated by its 3rd intron. Removal of this intron alleviates the repression of Bacterial Artificial Chromosome (BAC) IRF-8 reporter gene in these cells. Fine deletion analysis points to conserved regions within this intron mediating its restricted expression. Further, the intron alone selectively initiates gene silencing only in expression-restrictive cells. Characterization of this intron’s properties points to its role as an initiator of sustainable gene silencing inducing chromatin condensation with suppressive histone modifications. This intronic element cannot silence episomal transgene expression underlining a strict chromatin-dependent silencing mechanism. We validated this chromatin-state specificity of IRF-8 intron upon in-vitro differentiation of induced pluripotent stem cells (iPSCs) into cardiomyocytes. Taken together, the IRF-8 3rd intron is sufficient and necessary to initiate gene silencing in non-hematopoietic cells, highlighting its role as a nucleation core for repressed chromatin during differentiation. PMID:27257682

  13. Quantitative trait mapping reveals a regulatory axis involving peroxisome proliferator-activated receptors, PRDM16, transforming growth factor-β2 and FLT3 in hematopoiesis.

    PubMed

    Avagyan, Serine; Aguilo, Francesca; Kamezaki, Kenjiro; Snoeck, Hans-Willem

    2011-12-01

    Hematopoiesis is the process whereby BM HSCs renew to maintain their number or to differentiate into committed progenitors to generate all blood cells. One approach to gain mechanistic insight into this complex process is the investigation of quantitative genetic variation in hematopoietic function among inbred mouse strains. We previously showed that TGF-β2 is a genetically determined positive regulator of hematopoiesis. In the presence of unknown nonprotein serum factors TGF-β2, but not TGF-β1 or -β3, enhances progenitor proliferation in vitro, an effect that is subject to mouse strain-dependent variation mapping to a locus on chr.4, Tb2r1. TGF-β2-deficient mice show hematopoietic defects, demonstrating the physiologic role of this cytokine. Here, we show that TGF-β2 specifically and predominantly cell autonomously enhances signaling by FLT3 in vitro and in vivo. A coding polymorphism in Prdm16 (PR-domain-containing 16) underlies Tb2r1 and differentially regulates transcriptional activity of peroxisome proliferator-activated receptor-γ (PPARγ), identifying lipid PPAR ligands as the serum factors required for regulation of FLT3 signaling by TGF-β2. We furthermore show that PPARγ agonists play a FLT3-dependent role in stress responses of progenitor cells. These observations identify a novel regulatory axis that includes PPARs, Prdm16, and TGF-β2 in hematopoiesis.

  14. Transforming Growth Factor (TGF)-β1–producing Regulatory T Cells Induce Smad-mediated Interleukin 10 Secretion That Facilitates Coordinated Immunoregulatory Activity and Amelioration of TGF-β1–mediated Fibrosis

    PubMed Central

    Kitani, Atsushi; Fuss, Ivan; Nakamura, Kazuhiko; Kumaki, Fumiyuki; Usui, Takashi; Strober, Warren

    2003-01-01

    Interleukin (IL)-10 and transforming growth factor (TGF)-β1 are suppressor cytokines that frequently occur together during a regulatory T cell response. Here we used a one gene doxycycline (Dox)-inducible plasmid encoding TGF-β1 to analyze this association and test its utility. In initial studies, we showed that intranasal administration of this plasmid (along with Dox) led to the appearance of TGF-β1–producing cells (in spleen and lamina propria) and the almost concomitant appearance of IL-10–producing cells. Moreover, we showed that these cells exert Dox-regulated suppression of the T helper cell (Th)1-mediated inflammation in trinitrobenzene sulfonic acid colitis. In subsequent in vitro studies using retroviral TGF-β1 expression, we established that IL-10 production by Th1 cells occurs after exposure to TGF-β1 from either an endogenous or exogenous source. In addition, using a self-inactivating retrovirus luciferase reporter construct we showed that TGF-β1 induces Smad4, which then binds to and activates the IL-10 promoter. Furthermore, intranasal TGF-β1 plasmid administration ameliorates bleomycin-induced fibrosis in wild-type but not IL-10–deficient mice, strongly suggesting that the amelioration is IL-10 dependent and that IL-10 protects mice from TGF-β1–mediated fibrosis. Taken together, these findings suggest that the induction of IL-10 by TGF-β1 is not fortuitous, but instead fulfills important requirements of TGF-β1 function after its secretion by regulatory T cells. PMID:14557415

  15. Epstein-Barr virus transcription factor Zta acts through distal regulatory elements to directly control cellular gene expression.

    PubMed

    Ramasubramanyan, Sharada; Osborn, Kay; Al-Mohammad, Rajaei; Naranjo Perez-Fernandez, Ijiel B; Zuo, Jianmin; Balan, Nicolae; Godfrey, Anja; Patel, Harshil; Peters, Gordon; Rowe, Martin; Jenner, Richard G; Sinclair, Alison J

    2015-04-20

    Lytic replication of the human gamma herpes virus Epstein-Barr virus (EBV) is an essential prerequisite for the spread of the virus. Differential regulation of a limited number of cellular genes has been reported in B-cells during the viral lytic replication cycle. We asked whether a viral bZIP transcription factor, Zta (BZLF1, ZEBRA, EB1), drives some of these changes. Using genome-wide chromatin immunoprecipitation coupled to next-generation DNA sequencing (ChIP-seq) we established a map of Zta interactions across the human genome. Using sensitive transcriptome analyses we identified 2263 cellular genes whose expression is significantly changed during the EBV lytic replication cycle. Zta binds 278 of the regulated genes and the distribution of binding sites shows that Zta binds mostly to sites that are distal to transcription start sites. This differs from the prevailing view that Zta activates viral genes by binding exclusively at promoter elements. We show that a synthetic Zta binding element confers Zta regulation at a distance and that distal Zta binding sites from cellular genes can confer Zta-mediated regulation on a heterologous promoter. This leads us to propose that Zta directly reprograms the expression of cellular genes through distal elements. PMID:25779048

  16. Epstein–Barr virus transcription factor Zta acts through distal regulatory elements to directly control cellular gene expression

    PubMed Central

    Ramasubramanyan, Sharada; Osborn, Kay; Al-Mohammad, Rajaei; Naranjo Perez-Fernandez, Ijiel B.; Zuo, Jianmin; Balan, Nicolae; Godfrey, Anja; Patel, Harshil; Peters, Gordon; Rowe, Martin; Jenner, Richard G.; Sinclair, Alison J.

    2015-01-01

    Lytic replication of the human gamma herpes virus Epstein-Barr virus (EBV) is an essential prerequisite for the spread of the virus. Differential regulation of a limited number of cellular genes has been reported in B-cells during the viral lytic replication cycle. We asked whether a viral bZIP transcription factor, Zta (BZLF1, ZEBRA, EB1), drives some of these changes. Using genome-wide chromatin immunoprecipitation coupled to next-generation DNA sequencing (ChIP-seq) we established a map of Zta interactions across the human genome. Using sensitive transcriptome analyses we identified 2263 cellular genes whose expression is significantly changed during the EBV lytic replication cycle. Zta binds 278 of the regulated genes and the distribution of binding sites shows that Zta binds mostly to sites that are distal to transcription start sites. This differs from the prevailing view that Zta activates viral genes by binding exclusively at promoter elements. We show that a synthetic Zta binding element confers Zta regulation at a distance and that distal Zta binding sites from cellular genes can confer Zta-mediated regulation on a heterologous promoter. This leads us to propose that Zta directly reprograms the expression of cellular genes through distal elements. PMID:25779048

  17. Francisella tularensis Modulates a Distinct Subset of Regulatory Factors and Sustains Mitochondrial Integrity to Impair Human Neutrophil Apoptosis.

    PubMed

    McCracken, Jenna M; Kinkead, Lauren C; McCaffrey, Ramona L; Allen, Lee-Ann H

    2016-01-01

    Tularemia is a disease characterized by profound neutrophil accumulation and tissue destruction. The causative organism, Francisella tularensis, is a facultative intracellular bacterium that replicates in neutrophil cytosol, inhibits caspase activation and profoundly prolongs cell lifespan. Here, we identify unique features of this infection and provide fundamental insight into the mechanisms of apoptosis inhibition. Mitochondria are critical regulators of neutrophil apoptosis. We demonstrate that F. tularensis significantly inhibits Bax translocation and Bid processing during 24-48 h of infection, and in this manner sustains mitochondrial integrity. Downstream of mitochondria, X-linked inhibitor of apoptosis protein (XIAP) and proliferating cell nuclear antigen (PCNA) inhibit caspase-9 and caspase-3 by direct binding. Notably, we find that PCNA disappeared rapidly and selectively from infected cells, thereby demonstrating that it is not essential for neutrophil survival, whereas upregulation of calpastatin correlated with diminished calpain activity and reduced XIAP degradation. In addition, R-roscovitine is a cyclin-dependent kinase inhibitor developed for the treatment of cancer; it also induces neutrophil apoptosis and can promote the resolution of several infectious and inflammatory disorders. We confirm the ability of R-roscovitine to induce neutrophil apoptosis, but also demonstrate that its efficacy is significantly impaired by F. tularensis. Collectively, our findings advance the understanding of neutrophil apoptosis and its capacity to be manipulated by pathogenic bacteria.

  18. Regulatory effect of nerve growth factor in α9β1 integrin–dependent progression of glioblastoma

    PubMed Central

    Brown, Meghan C.; Staniszewska, Izabela; Lazarovici, Philip; Tuszynski, George P.; Del Valle, Luis; Marcinkiewicz, Cezary

    2008-01-01

    In the present study we described the role of α9β1 integrin in glioblastoma progression following its interaction with nerve growth factor (NGF). The level of expression of α9β1 on astrocytomas is correlated with increased grade of this brain tumor and is highest on glioblastoma, whereas normal astrocytes do not express this integrin. Two glioblastoma cell lines, LN229 and LN18, that are α9β1 integrin positive and negative, respectively, were used for α9β1 integrin–dependent NGF-induced tumor progression. NGF was a significant promoter of promigratory and pro-proliferative activities of glioblastoma cells through direct interaction with α9β1 integrin and activation of MAPK Erk1/2 pathway. The level of NGF increases approximately threefold in the most malignant glioma tissue when compared with normal brain. This increase is related to secretion of NGF by tumor cells. Specific inhibitors of α9β1 integrin or gene silencing inhibited NGF-induced proliferation of LN229 cell line to the level shown by LN18 cells. VLO5 promoted α9β1-dependent programmed cell death by induction of intrinsic apoptosis pathway in cancer cells. LN229 cells were rescued from proapoptotic effect of VLO5 by the presence of NGF. This disintegrin significantly inhibited tumor growth induced by implantation of LN229 cells to the chorioallantoic membrane (CAM) of quail embryonic model, and this inhibitory effect was significantly abolished by the presence of NGF. α9β1 integrin appears to be an interesting target for blocking the progression of malignant gliomas, especially in light of the stimulatory effect of NGF on the development of these tumors and its ability to transfer proapoptotic signals in cancer cells. PMID:19074980

  19. Regulatory Action of Calcium Ion on Cyclic AMP-Enhanced Expression of Implantation-Related Factors in Human Endometrial Cells

    PubMed Central

    Kusama, Kazuya; Yoshie, Mikihiro; Tamura, Kazuhiro; Imakawa, Kazuhiko; Isaka, Keiichi; Tachikawa, Eiichi

    2015-01-01

    Decidualization of human endometrial stroma and gland development is mediated through cyclic AMP (cAMP), but the role of intracellular calcium ion (Ca2+) on cAMP mediated-signaling in human endometrial stroma and glandular epithelia has not been well-characterized. The present study was designed to investigate the role of intracellular Ca2+ on cAMP mediated-decidualization and gland maturation events, which can be identified by the up-regulation of prolactin and IGF-binding protein (IGFBP)1 in human endometrial stromal cells (ESCs), and cyclooxygenase 2 (COX2) and prostaglandin E2 (PGE2) and glandular epithelial EM-1 cells. Increases in decidual prolactin and IGFBP-1 transcript levels, induced by cAMP-elevating agents forskolin or dibutyryl cyclic AMP, were inhibited by Ca2+ influx into ESCs with Ca2+ ionophores (alamethicin, ionomycin) in a dose-dependent manner. Conversely, inhibitors of Ca2+ influx through L-type voltage-dependent Ca2+ channel (VDCC), nifedipine and verapamil, enhanced the decidual gene expression. Furthermore, dantrolene, an inhibitor of Ca2+ release from the intracellular Ca2+ store, up-regulated prolactin and IGFBP-1 expression. Ca2+ ionophores decreased intracellular cAMP concentrations, whereas nifedipine, verapamil or dantrolene increased cAMP concentrations in ESCs. In glandular epithelial cells, similar responses in COX2 expression and PGE2 production were found when intracellular cAMP levels were up-regulated by decreases in Ca2+ concentrations. Thus, a marked decrease in cytosolic Ca2+ levels caused the elevation of cAMP concentrations, resulting in enhanced expression of implantation-related factors including decidual markers. These findings suggest that fluctuation in cytosolic Ca2+ concentrations alters intracellular cAMP levels, which then regulate differentiation of endometrial stromal and glandular epithelial cells. PMID:26161798

  20. Mammals of Australia's Tropical Savannas: A Conceptual Model of Assemblage Structure and Regulatory Factors in the Kimberley Region

    PubMed Central

    Radford, Ian J.; Dickman, Christopher R.; Start, Antony N.; Palmer, Carol; Carnes, Karin; Everitt, Corrin; Fairman, Richard; Graham, Gordon; Partridge, Thalie; Thomson, Allan

    2014-01-01

    We construct a state-and-transition model for mammals in tropical savannas in northern Australia to synthesize ecological knowledge and understand mammalian declines. We aimed to validate the existence of alternative mammal assemblage states similar to those in arid Australian grasslands, and to speculate on transition triggers. Based on the arid grassland model, we hypothesized that assemblages are partitioned across rainfall gradients and between substrates. We also predicted that assemblages typical of arid regions in boom periods would be prevalent in savannas with higher and more regular rainfall. Data from eight mammal surveys from the Kimberley region, Western Australia (1994 to 2011) were collated. Survey sites were partitioned across rainfall zones and habitats. Data allowed us to identify three assemblage states: State 0:- low numbers of mammals, State II:- dominated by omnivorous rodents and State III:- dominated by rodents and larger marsupials. Unlike arid grasslands, assemblage dominance by insectivorous dasyurids (State I) did not occur in savannas. Mammal assemblages were partitioned across rainfall zones and between substrates as predicted, but—unlike arid regions—were not related strongly to yearly rainfall. Mammal assemblage composition showed high regional stability, probably related to high annual rainfall and predictable wet season resource pulses. As a consequence, we speculate that perpetually booming assemblages in savannas allow top-down control of the ecosystem, with suppression of introduced cats by the dingo, the region's top predator. Under conditions of low or erratic productivity, imposed increasingly by intense fire regimes and introduced herbivore grazing, dingoes may not limit impacts of cats on native mammals. These interacting factors may explain contemporary declines of savanna mammals as well as historical declines in arid Australia. The cat-ecosystem productivity hypothesis raised here differs from the already

  1. Mammals of Australia's tropical savannas: a conceptual model of assemblage structure and regulatory factors in the Kimberley region.

    PubMed

    Radford, Ian J; Dickman, Christopher R; Start, Antony N; Palmer, Carol; Carnes, Karin; Everitt, Corrin; Fairman, Richard; Graham, Gordon; Partridge, Thalie; Thomson, Allan

    2014-01-01

    We construct a state-and-transition model for mammals in tropical savannas in northern Australia to synthesize ecological knowledge and understand mammalian declines. We aimed to validate the existence of alternative mammal assemblage states similar to those in arid Australian grasslands, and to speculate on transition triggers. Based on the arid grassland model, we hypothesized that assemblages are partitioned across rainfall gradients and between substrates. We also predicted that assemblages typical of arid regions in boom periods would be prevalent in savannas with higher and more regular rainfall. Data from eight mammal surveys from the Kimberley region, Western Australia (1994 to 2011) were collated. Survey sites were partitioned across rainfall zones and habitats. Data allowed us to identify three assemblage states: State 0:--low numbers of mammals, State II:--dominated by omnivorous rodents and State III:--dominated by rodents and larger marsupials. Unlike arid grasslands, assemblage dominance by insectivorous dasyurids (State I) did not occur in savannas. Mammal assemblages were partitioned across rainfall zones and between substrates as predicted, but-unlike arid regions-were not related strongly to yearly rainfall. Mammal assemblage composition showed high regional stability, probably related to high annual rainfall and predictable wet season resource pulses. As a consequence, we speculate that perpetually booming assemblages in savannas allow top-down control of the ecosystem, with suppression of introduced cats by the dingo, the region's top predator. Under conditions of low or erratic productivity, imposed increasingly by intense fire regimes and introduced herbivore grazing, dingoes may not limit impacts of cats on native mammals. These interacting factors may explain contemporary declines of savanna mammals as well as historical declines in arid Australia. The cat-ecosystem productivity hypothesis raised here differs from the already-articulated cat

  2. Mammals of Australia's tropical savannas: a conceptual model of assemblage structure and regulatory factors in the Kimberley region.

    PubMed

    Radford, Ian J; Dickman, Christopher R; Start, Antony N; Palmer, Carol; Carnes, Karin; Everitt, Corrin; Fairman, Richard; Graham, Gordon; Partridge, Thalie; Thomson, Allan

    2014-01-01

    We construct a state-and-transition model for mammals in tropical savannas in northern Australia to synthesize ecological knowledge and understand mammalian declines. We aimed to validate the existence of alternative mammal assemblage states similar to those in arid Australian grasslands, and to speculate on transition triggers. Based on the arid grassland model, we hypothesized that assemblages are partitioned across rainfall gradients and between substrates. We also predicted that assemblages typical of arid regions in boom periods would be prevalent in savannas with higher and more regular rainfall. Data from eight mammal surveys from the Kimberley region, Western Australia (1994 to 2011) were collated. Survey sites were partitioned across rainfall zones and habitats. Data allowed us to identify three assemblage states: State 0:--low numbers of mammals, State II:--dominated by omnivorous rodents and State III:--dominated by rodents and larger marsupials. Unlike arid grasslands, assemblage dominance by insectivorous dasyurids (State I) did not occur in savannas. Mammal assemblages were partitioned across rainfall zones and between substrates as predicted, but-unlike arid regions-were not related strongly to yearly rainfall. Mammal assemblage composition showed high regional stability, probably related to high annual rainfall and predictable wet season resource pulses. As a consequence, we speculate that perpetually booming assemblages in savannas allow top-down control of the ecosystem, with suppression of introduced cats by the dingo, the region's top predator. Under conditions of low or erratic productivity, imposed increasingly by intense fire regimes and introduced herbivore grazing, dingoes may not limit impacts of cats on native mammals. These interacting factors may explain contemporary declines of savanna mammals as well as historical declines in arid Australia. The cat-ecosystem productivity hypothesis raised here differs from the already-articulated cat

  3. Regulatory Action of Calcium Ion on Cyclic AMP-Enhanced Expression of Implantation-Related Factors in Human Endometrial Cells.

    PubMed

    Kusama, Kazuya; Yoshie, Mikihiro; Tamura, Kazuhiro; Imakawa, Kazuhiko; Isaka, Keiichi; Tachikawa, Eiichi

    2015-01-01

    Decidualization of human endometrial stroma and gland development is mediated through cyclic AMP (cAMP), but the role of intracellular calcium ion (Ca2+) on cAMP mediated-signaling in human endometrial stroma and glandular epithelia has not been well-characterized. The present study was designed to investigate the role of intracellular Ca2+ on cAMP mediated-decidualization and gland maturation events, which can be identified by the up-regulation of prolactin and IGF-binding protein (IGFBP)1 in human endometrial stromal cells (ESCs), and cyclooxygenase 2 (COX2) and prostaglandin E2 (PGE2) and glandular epithelial EM-1 cells. Increases in decidual prolactin and IGFBP-1 transcript levels, induced by cAMP-elevating agents forskolin or dibutyryl cyclic AMP, were inhibited by Ca2+ influx into ESCs with Ca2+ ionophores (alamethicin, ionomycin) in a dose-dependent manner. Conversely, inhibitors of Ca2+ influx through L-type voltage-dependent Ca2+ channel (VDCC), nifedipine and verapamil, enhanced the decidual gene expression. Furthermore, dantrolene, an inhibitor of Ca2+ release from the intracellular Ca2+ store, up-regulated prolactin and IGFBP-1 expression. Ca2+ ionophores decreased intracellular cAMP concentrations, whereas nifedipine, verapamil or dantrolene increased cAMP concentrations in ESCs. In glandular epithelial cells, similar responses in COX2 expression and PGE2 production were found when intracellular cAMP levels were up-regulated by decreases in Ca2+ concentrations. Thus, a marked decrease in cytosolic Ca2+ levels caused the elevation of cAMP concentrations, resulting in enhanced expression of implantation-related factors including decidual markers. These findings suggest that fluctuation in cytosolic Ca2+ concentrations alters intracellular cAMP levels, which then regulate differentiation of endometrial stromal and glandular epithelial cells. PMID:26161798

  4. Vault-poly-ADP-ribose polymerase in the Octopus vulgaris brain: a regulatory factor of actin polymerization dynamic.

    PubMed

    De Maio, Anna; Natale, Emiliana; Rotondo, Sergio; Di Cosmo, Anna; Faraone-Mennella, Maria Rosaria

    2013-09-01

    Our previous behavioural, biochemical and immunohistochemical analyses conducted in selected regions (supra/sub oesophageal masses) of the Octopus vulgaris brain detected a cytoplasmic poly-ADP-ribose polymerase (more than 90% of total enzyme activity). The protein was identified as the vault-free form of vault-poly-ADP-ribose polymerase. The present research extends and integrates the biochemical characterization of poly-ADP-ribosylation system, namely, reaction product, i.e., poly-ADP-ribose, and acceptor proteins, in the O. vulgaris brain. Immunochemical analyses evidenced that the sole poly-ADP-ribose acceptor was the octopus cytoskeleton 50-kDa actin. It was present in both free, endogenously poly-ADP-ribosylated form (70kDa) and in complex with V-poly-ADP-ribose polymerase and poly-ADP-ribose (260kDa). The components of this complex, alkali and high salt sensitive, were purified and characterized. The kind and the length of poly-ADP-ribose corresponded to linear chains of 30-35 ADP-ribose units, in accordance with the features of the polymer synthesized by the known vault-poly-ADP-ribose polymerase. In vitro experiments showed that V-poly-ADP-ribose polymerase activity of brain cytoplasmic fraction containing endogenous actin increased upon the addition of commercial actin and was highly reduced by ATP. Anti-actin immunoblot of the mixture in the presence and absence of ATP showed that the poly-ADP-ribosylation of octopus actin is a dynamic process balanced by the ATP-dependent polymerization of the cytoskeleton protein, a fundamental mechanism for synaptic plasticity.

  5. Processed engineered fuels derived from paper and plastics - techno-economic factors and regulatory issues in a competitive market

    SciTech Connect

    Fisher, M.M.; Singhania, A.; Sosa, J.; Ohlsson, O.O.

    1997-12-01

    Plastics have gained a prominent position in modern society because their unique properties, cost-performance and versatility. They contribute to almost every aspect of modern life including transportation, recreation, medicine, packaging, appliances, furniture, textiles and building and construction. The diverse properties of plastics allow them to play a major role in providing solutions for technical progress as well as solutions for everyday life. Plastics also make a significant contribution to effective resource management and conservation, reducing the use of natural resources and helping to minimize waste. By choosing plastics, manufacturers, designers, distributors and retailers in every sector have been able to respond to the challenge of preventing excess use of resources and, at the same time, bring increasingly sophisticated products to the market. Resource conservation seeks to maximize efficiency and minimize waste from a product`s manufacture, use and disposal. It involves more than managing and minimizing waste, however; it also includes taking advantage of effective life-cycle resource management at local and global levels. There are many ways that plastics contribute to effective resource management. From manufacture to use, to recovery, to waste management, plastics help conserve resources. Not only are plastics energy efficient, but they often create less waste than other materials. They can be recovered, recycled and safely landfilled. This paper reviews the energy recovery option for post-use plastics. It discusses plastics` role in modern waste-to-energy systems and then reviews some recent work exploring the use of plastics as alternative fuels. The emphasis is on solid fuels, often referred to as process engineered fuel or PEF. The liquefaction and gasification of plastics offers additional opportunities for resource recovery, but this subject is only briefly discussed in this paper.

  6. Lactobacilli Modulate Hypoxia-Inducible Factor (HIF)-1 Regulatory Pathway in Triple Negative Breast Cancer Cell Line

    PubMed Central

    Abedin-Do, Atieh; Mirfakhraie, Reza; Shirzad, Mahdieh; Ghafouri-Fard, Soudeh; Motevaseli, Elahe

    2016-01-01

    Objective Hypoxia-Inducible Factor (HIF)-1 plays an essential role in the body’s response to low oxygen concentrations and regulates expression of several genes implicated in homeostasis, vascularization, anaerobic metabolism as well as immunological responses. Increased levels of HIF-1α are associated with increased proliferation and more aggressive breast tumor development. Lactobacilli have been shown to exert anti-cancer effects on several malignancies including breast cancer. However, the exact mechanism of such effect is not clear yet. The aim of this study was to analyze the expression of selected genes from HIF pathway in a triple negative breast cancer cell line (expressing no estrogen and progesterone receptors as well as HER-2/Neu), MDA-MB-231, following treatment with two lactobacilli culture supernatants. Materials and Methods In this experimental study, we analyzed the expression of HIF-1α, SLC2A1, VHL, HSP90, XBP1 and SHARP1 genes from HIF pathway in MDA-MB-231 cells, before and after treatment with Lactobacillus crispatus and Lactobacillus rhamnosus culture supernatants (LCS and LRS, respectively) by means of quantitative reverse-transcription polymerase chain reaction (qRT-PCR). Results Both LRS and LCS had cytotoxic effects on MDA-MB-231 cells, while the former type was more cytotoxic. LRS dramatically down-regulated expression levels of the HIF-1α, HSP90 and SLC2A1 in the MDA-MB-231 cells. LCS had similar effect on the expression of HSP90, to what was observed in the LRS treatment. The expression level of tumor suppressor genes VHL and SHARP1 were also decreased in LCS treated cells. Conclusion Although both LCS and LRS had cytotoxic effects on the MDA-MB-231 cells, it is proposed that LRS could be more appropriate for pathway directed treatment modalities, as it did not decrease expression of tumor suppressor genes involved in HIF pathway. Down-regulation of HIF pathway mediated oncogenes by LRS suggests that the cytotoxic effects of this

  7. Influence of physico-chemical factors on leaching of chemical additives from aluminium foils used for packaging of food materials.

    PubMed

    Ojha, Priyanka; Ojha, C S; Sharma, V P

    2007-01-01

    In recent years, the use of aluminium foils to wrap foodstuff and commodities has been increased to a great extent. Aluminium was found to leach out from the foil in different simulants particularly in distilled water, acidic and alkaline medium at 60 +/- 2 degrees C for 2 hours and 40 +/- 2 degrees C for 24 hours. The migration was found to be above the permissible limit as laid down by WHO guidelines, that is of 0.2 mg/L of water. The protocol used for this study was based on the recommendation of Bureau of Indian Standard regarding the migration of chemical additives from packaging materials used to pack food items. Migration of the aluminium metal was found significantly higher in acidic and aqueous medium in comparison to alcoholic and saline medium. Higher temperature conditions also enhanced the rate of migration of aluminium in acidic and aqueous medium. Leaching of aluminium metal occurred in double distilled water, acetic acid 3%, normal saline and sodium carbonate, except ethanol 8%, in which aluminium migration was below the detection limit of the instrument where three brands of the aluminium foil samples studied.

  8. A new single nucleotide polymorphism in the insulin-like growth factor I regulatory region associates with colorectal cancer risk in singapore chinese.

    PubMed

    Wong, Hui-Lee; Delellis, Katherine; Probst-Hensch, Nicole; Koh, Woon-Puay; Van Den Berg, David; Lee, Hin-Peng; Yu, Mimi C; Ingles, Sue A

    2005-01-01

    Elevated levels of plasma insulin-like growth factor I (IGF-I) are a potential risk factor for several cancers, including colorectal cancer. Physiologic levels of plasma IGF-I vary greatly; this variation may be in part genetically determined. We identified two single nucleotide polymorphisms (SNP) in perfect linkage disequilibrium with each other and in partial linkage disequilibrium with a previously studied cytosine-adenine microsatellite [-969(CA)(n)]. We investigated one of the SNPs, -533T/C,and the 969(CA)(n) in relation to the risk of colorectal cancer in a case-control study nested within a cohort of Singapore Chinese (cases/controls = 290:873). The (CA)(21) allele, rather than the previously implicated (CA)(19) allele, was associated with a reduced risk of colorectal cancer (odds ratio for 21/21 versus all other genotypes, 0.48; 95% confidence interval, 0.28-0.84). For the -533C/T SNP, persons carrying one or more copies of the C allele had a decreased in risk of colorectal cancer compared with noncarriers (odds ratio for CC/CT versus TT, 0.58; 95% confidence interval, 0.41-0.82). This association was specific for colon, as opposed to rectal cancer and was modified by age. We also examined a functional insulin-like growth factor binding protein (IGFBP3) promoter SNP, -202 A/C, previously reported to predict serum IGFBP3 levels. Although we were able to confirm this genotype-phenotype association, the -202A/C IGFBP3 SNP was not significantly associated with colorectal cancer risk. In conclusion, we report a novel SNP in the IGF-I regulatory region that is associated with colorectal cancer risk.

  9. Apoptosis Induced by Mammalian Reovirus Is Beta Interferon (IFN) Independent and Enhanced by IFN Regulatory Factor 3- and NF-κB-Dependent Expression of Noxa

    PubMed Central

    Knowlton, Jonathan J.; Dermody, Terence S.

    2012-01-01

    A variety of signal transduction pathways are activated in response to viral infection, which dampen viral replication and transmission. These mechanisms involve both the induction of type I interferons (IFNs), which evoke an antiviral state, and the triggering of apoptosis. Mammalian orthoreoviruses are double-stranded RNA viruses that elicit apoptosis in vitro and in vivo. The transcription factors interferon regulatory factor 3 (IRF-3) and nuclear factor kappa light-chain enhancer of activated B cells (NF-κB) are required for the expression of IFN-β and the efficient induction of apoptosis in reovirus-infected cells. However, it is not known whether IFN-β induction is required for apoptosis, nor have the genes induced by IRF-3 and NF-κB that are responsible for apoptosis been identified. To determine whether IFN-β is required for reovirus-induced apoptosis, we used type I IFN receptor-deficient cells, IFN-specific antibodies, and recombinant IFN-β. We found that IFN synthesis and signaling are dispensable for the apoptosis of reovirus-infected cells. These results indicate that the apoptotic response following reovirus infection is mediated directly by genes responsive to IRF-3 and NF-κB. Noxa is a proapoptotic BH3-domain-only protein of the Bcl-2 family that requires IRF-3 and NF-κB for efficient expression. We found that Noxa is strongly induced at late times (36 to 48 h) following reovirus infection in a manner dependent on IRF-3 and NF-κB. The level of apoptosis induced by reovirus is significantly diminished in cells lacking Noxa, indicating a key prodeath function for this molecule during reovirus infection. These results suggest that prolonged innate immune response signaling induces apoptosis by eliciting Noxa expression in reovirus-infected cells. PMID:22090144

  10. Transforming growth factor-β2 increases the capacity of retinal pigment epithelial cells to induce the generation of regulatory T cells.

    PubMed

    Yan, Feng; He, Jin; Tang, Li; Kong, Yi; Shi, Yuhua; Chen, Suihua; Huang, Zhenping

    2016-02-01

    The present study investigated the underlying mechanism of the induction of regulatory T cells (Tregs) by retinal pigment epithelial (RPE) cells and the characteristics of these Tregs. Human RPE cells were cultured in the presence or absence of transforming growth factor-β 2 (TGF-β2), and reverse-transcription quantitative PCR was performed to determine the mRNA expression of indoleamine 2,3-dioxygenase (IDO) and nuclear factor erythroid 2-related factor (Nrf2). Supernatants of RPE cell cultures were added to CD4+ T cells to induce Tregs. The RPE-induced Tregs were purified by two-step magnetic cell sorting. The natural Tregs were isolated from the peripheral blood mononuclear cells of healthy volunteers. Purified CD4+ CD25- T cells (2 x 10(5)/well) were cultured alone or with Tregs (various densities, natural or RPE-induced). The proliferation of CD4+ CD25- T cells was determined by 3H-thymidine incorporation. After 24 h of stimulation with TGF-β2, the mRNA expression of IDO in RPE cells was upregulated. The highest level of IDO mRNA expression was reached after 72 h of stimulation with TGF-β2. However, the Nrf2 mRNA expression was slightly decreased after 24 h of stimulation with TGF-β2 and significantly increased after 48-72 h of TGF-β2 stimulation. Increased levels of CD25 expression were observed on CD4+ T cells exposed to supernatants of RPE cell cultures treated with TGF-β2 and recombinant interleukin-2. The RPE-induced Tregs were more effective at suppressing the proliferation of CD4+ CD25- T cells compared with native Tregs. These findings suggested that IDO may be a signaling protein in RPE cells which is implicated in the induction of Tregs. RPE-induced Tregs have the potential to be applied for immunotherapy for ocular inflammatory diseases.

  11. The severity of retinal pathology in homozygous Crb1rd8/rd8 mice is dependent on additional genetic factors.

    PubMed

    Luhmann, Ulrich F O; Carvalho, Livia S; Holthaus, Sophia-Martha Kleine; Cowing, Jill A; Greenaway, Simon; Chu, Colin J; Herrmann, Philipp; Smith, Alexander J; Munro, Peter M G; Potter, Paul; Bainbridge, James W B; Ali, Robin R

    2015-01-01

    Understanding phenotype-genotype correlations in retinal degeneration is a major challenge. Mutations in CRB1 lead to a spectrum of autosomal recessive retinal dystrophies with variable phenotypes suggesting the influence of modifying factors. To establish the contribution of the genetic background to phenotypic variability associated with the Crb1(rd8/rd8) mutation, we compared the retinal pathology of Crb1(rd8/rd8)/J inbred mice with that of two Crb1(rd8/rd8) lines backcrossed with C57BL/6JOlaHsd mice. Topical endoscopic fundal imaging and scanning laser ophthalmoscopy fundus images of all three Crb1(rd8/rd8) lines showed a significant increase in the number of inferior retinal lesions that was strikingly variable between the lines. Optical coherence tomography, semithin, ultrastructural morphology and assessment of inflammatory and vascular marker by immunohistochemistry and quantitative reverse transcriptase-polymerase chain reaction revealed that the lesions were associated with photoreceptor death, Müller and microglia activation and telangiectasia-like vascular remodelling-features that were stable in the inbred, variable in the second, but virtually absent in the third Crb1(rd8/rd8) line, even at 12 months of age. This suggests that the Crb1(rd8/rd8) mutation is necessary, but not sufficient for the development of these degenerative features. By whole-genome SNP analysis of the genotype-phenotype correlation, a candidate region on chromosome 15 was identified. This may carry one or more genetic modifiers for the manifestation of the retinal pathology associated with mutations in Crb1. This study also provides insight into the nature of the retinal vascular lesions that likely represent a clinical correlate for the formation of retinal telangiectasia or Coats-like vasculopathy in patients with CRB1 mutations that are thought to depend on such genetic modifiers.

  12. Physiological basis of tolerance to complete submergence in rice involves genetic factors in addition to the SUB1 gene.

    PubMed

    Singh, Sudhanshu; Mackill, David J; Ismail, Abdelbagi M

    2014-01-01

    1 lines. This suggests the possibility of further improvements in submergence tolerance by incorporating additional traits present in FR13A or other similar landraces. PMID:25281725

  13. Symbolic integration of a product of two spherical Bessel functions with an additional exponential and polynomial factor

    NASA Astrophysics Data System (ADS)

    Gebremariam, B.; Duguet, T.; Bogner, S. K.

    2010-06-01

    We present a Mathematica package that performs the symbolic calculation of integrals of the form ∫0∞exj(x)j(x)dx where j(x) and j(x) denote spherical Bessel functions of integer orders, with ν⩾0 and μ⩾0. With the real parameter u>0 and the integer n, convergence of the integral requires that n+ν+μ⩾0. The package provides analytical result for the integral in its most simplified form. In cases where direct Mathematica implementations succeed in evaluating these integrals, the novel symbolic method implemented in this work obtains the same result and in general, it takes a fraction of the time required for the direct implementation. We test the accuracy of such analytical expressions by comparing the results with their numerical counterparts. Program summaryProgram title: SymbBesselJInteg Catalogue identifier: AEFY_v1_0 Program summary URL:http://cpc.cs.qub.ac.uk/summaries/AEFY_v1_0.html Program obtainable from: CPC Program Library, Queen's University, Belfast, N. Ireland Licensing provisions: Standard CPC licence, http://cpc.cs.qub.ac.uk/licence/licence.html No. of lines in distributed program, including test data, etc.: 275 934 No. of bytes in distributed program, including test data, etc.: 399 705 Distribution format: tar.gz Programming language: Mathematica 7.1 Computer: Any computer running Mathematica 6.0 and later versions. Operating system: Windows Xp, Linux/Unix. RAM: 256 Mb Classification: 5. Nature of problem: Integration, both analytical and numerical, of products of two spherical Bessel functions with an exponential and polynomial multiplying factor can be a very complex task depending on the orders of the spherical Bessel functions. The Mathematica package discussed in this paper solves this problem using a novel symbolic approach. Solution method: The problem is first cast into a related limit problem which can be broken into two related subproblems involving exponential and exponential integral functions. Solving the cores of each

  14. Genome-Wide Mapping of the Binding Sites and Structural Analysis of Kaposi's Sarcoma-Associated Herpesvirus Viral Interferon Regulatory Factor 2 Reveal that It Is a DNA-Binding Transcription Factor

    PubMed Central

    Hu, Haidai; Dong, Jiazhen; Liang, Deguang; Gao, Zengqiang; Bai, Lei; Sun, Rui; Hu, Hao; Zhang, Heng

    2015-01-01

    ABSTRACT The oncogenic herpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV) is known to encode four viral interferon regulatory factors (vIRF1 to -4) to subvert the host antiviral immune response, but their detailed DNA-binding profiles as transcription factors in the host remain uncharacterized. Here, we first performed genome-wide vIRF2-binding site mapping in the human genome using chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq). vIRF2 was capable of binding to the promoter regions of 100 putative target genes. Importantly, we confirmed that vIRF2 can specifically interact with the promoters of the genes encoding PIK3C3, HMGCR, and HMGCL, which are associated with autophagosome formation or tumor progression and metastasis, and regulate their transcription in vivo. The crystal structure of the vIRF2 DNA-binding domain (DBD) (referred to here as vIRF2DBD) showed variable loop conformations and positive-charge distributions different from those of vIRF1 and cellular IRFs that are associated with DNA-binding specificities. Structure-based mutagenesis revealed that Arg82 and Arg85 are required for the in vitro DNA-binding activity of vIRF2DBD and can abolish the transcription regulation function of vIRF2 on the promoter reporter activity of PIK3C3, HMGCR, and HMGCL. Collectively, our study provided unique insights into the DNA-binding potency of vIRF2 and suggested that vIRF2 could act as a transcription factor of its target genes in the host antiviral immune response. IMPORTANCE The oncogenic herpesvirus KSHV is the etiological agent of Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman's disease. KSHV has developed a unique mechanism to subvert the host antiviral immune responses by encoding four homologues of cellular interferon regulatory factors (vIRF1 to -4). However, none of their DNA-binding profiles in the human genome have been characterized until now, and the structural basis for their diverse

  15. Differentiated evolutionary relationships among chordates from comparative alignments of multiple sequences of MyoD and MyoG myogenic regulatory factors.

    PubMed

    Oliani, L C; Lidani, K C F; Gabriel, J E

    2015-10-16

    MyoD and MyoG are transcription factors that have essential roles in myogenic lineage determination and muscle differentiation. The purpose of this study was to compare multiple amino acid sequences of myogenic regulatory proteins to infer evolutionary relationships among chordates. Protein sequences from Mus musculus (P10085 and P12979), human Homo sapiens (P15172 and P15173), bovine Bos taurus (Q7YS82 and Q7YS81), wild pig Sus scrofa (P49811 and P49812), quail Coturnix coturnix (P21572 and P34060), chicken Gallus gallus (P16075 and P17920), rat Rattus norvegicus (Q02346 and P20428), domestic water buffalo Bubalus bubalis (D2SP11 and A7L034), and sheep Ovis aries (Q90477 and D3YKV7) were searched from a non-redundant protein sequence database UniProtKB/Swiss-Prot, and subsequently analyzed using the Mega6.0 software. MyoD evolutionary analyses revealed the presence of three main clusters with all mammals branched in one cluster, members of the order Rodentia (mouse and rat) in a second branch linked to the first, and birds of the order Galliformes (chicken and quail) remaining isolated in a third. MyoG evolutionary analyses aligned sequences in two main clusters, all mammalian specimens grouped in different sub-branches, and birds clustered in a second branch. These analyses suggest that the evolution of MyoD and MyoG was driven by different pathways.

  16. Differentiated evolutionary relationships among chordates from comparative alignments of multiple sequences of MyoD and MyoG myogenic regulatory factors.

    PubMed

    Oliani, L C; Lidani, K C F; Gabriel, J E

    2015-01-01

    MyoD and MyoG are transcription factors that have essential roles in myogenic lineage determination and muscle differentiation. The purpose of this study was to compare multiple amino acid sequences of myogenic regulatory proteins to infer evolutionary relationships among chordates. Protein sequences from Mus musculus (P10085 and P12979), human Homo sapiens (P15172 and P15173), bovine Bos taurus (Q7YS82 and Q7YS81), wild pig Sus scrofa (P49811 and P49812), quail Coturnix coturnix (P21572 and P34060), chicken Gallus gallus (P16075 and P17920), rat Rattus norvegicus (Q02346 and P20428), domestic water buffalo Bubalus bubalis (D2SP11 and A7L034), and sheep Ovis aries (Q90477 and D3YKV7) were searched from a non-redundant protein sequence database UniProtKB/Swiss-Prot, and subsequently analyzed using the Mega6.0 software. MyoD evolutionary analyses revealed the presence of three main clusters with all mammals branched in one cluster, members of the order Rodentia (mouse and rat) in a second branch linked to the first, and birds of the order Galliformes (chicken and quail) remaining isolated in a third. MyoG evolutionary analyses aligned sequences in two main clusters, all mammalian specimens grouped in different sub-branches, and birds clustered in a second branch. These analyses suggest that the evolution of MyoD and MyoG was driven by different pathways. PMID:26505406

  17. A Novel Sucrose-Regulatory MADS-Box Transcription Factor GmNMHC5 Promotes Root Development and Nodulation in Soybean (Glycine max [L.] Merr.).

    PubMed

    Liu, Wei; Han, Xiangdong; Zhan, Ge; Zhao, Zhenfang; Feng, Yongjun; Wu, Cunxiang

    2015-08-31

    The MADS-box protein family includes many transcription factors that have a conserved DNA-binding MADS-box domain. The proteins in this family were originally recognized to play prominent roles in floral development. Recent findings, especially with regard to the regulatory roles of the AGL17 subfamily in root development, have greatly broadened their known functions. In this study, a gene from soybean (Glycine max [L.] Merr.), GmNMHC5, was cloned from the Zigongdongdou cultivar and identified as a member of the AGL17 subfamily. Real-time fluorescence quantitative PCR analysis showed that GmNMHC5 was expressed at much higher levels in roots and nodules than in other organs. The activation of expression was first examined in leaves and roots, followed by shoot apexes. GmNMHC5 expression levels rose sharply when the plants were treated under short-day conditions (SD) and started to pod, whereas low levels were maintained in non-podding plants under long-day conditions (LD). Furthermore, overexpression of GmNMHC5 in transgenic soybean significantly promoted lateral root development and nodule building. Moreover, GmNMHC5 is upregulated by exogenous sucrose. These results indicate that GmNMHC5 can sense the sucrose signal and plays significant roles in lateral root development and nodule building.

  18. A Novel Sucrose-Regulatory MADS-Box Transcription Factor GmNMHC5 Promotes Root Development and Nodulation in Soybean (Glycine max [L.] Merr.)

    PubMed Central

    Liu, Wei; Han, Xiangdong; Zhan, Ge; Zhao, Zhenfang; Feng, Yongjun; Wu, Cunxiang

    2015-01-01

    The MADS-box protein family includes many transcription factors that have a conserved DNA-binding MADS-box domain. The proteins in this family were originally recognized to play prominent roles in floral development. Recent findings, especially with regard to the regulatory roles of the AGL17 subfamily in root development, have greatly broadened their known functions. In this study, a gene from soybean (Glycine max [L.] Merr.), GmNMHC5, was cloned from the Zigongdongdou cultivar and identified as a member of the AGL17 subfamily. Real-time fluorescence quantitative PCR analysis showed that GmNMHC5 was expressed at much higher levels in roots and nodules than in other organs. The activation of expression was first examined in leaves and roots, followed by shoot apexes. GmNMHC5 expression levels rose sharply when the plants were treated under short-day conditions (SD) and started to pod, whereas low levels were maintained in non-podding plants under long-day conditions (LD). Furthermore, overexpression of GmNMHC5 in transgenic soybean significantly promoted lateral root development and nodule building. Moreover, GmNMHC5 is upregulated by exogenous sucrose. These results indicate that GmNMHC5 can sense the sucrose signal and plays significant roles in lateral root development and nodule building. PMID:26404246

  19. Dissection of the regulatory mechanism of a heat-shock responsive promoter in Haloarchaea: a new paradigm for general transcription factor directed archaeal gene regulation

    PubMed Central

    Lu, Qiuhe; Han, Jing; Zhou, Ligang; Coker, James A.; DasSarma, Priya; DasSarma, Shiladitya; Xiang, Hua

    2008-01-01

    Multiple general transcription factors (GTFs), TBP and TFB, are present in many haloarchaea, and are deemed to accomplish global gene regulation. However, details and the role of GTF-directed transcriptional regulation in stress response are still not clear. Here, we report a comprehensive investigation of the regulatory mechanism of a heat-induced gene (hsp5) from Halobacterium salinarum. We demonstrated by mutation analysis that the sequences 5′ and 3′ to the core elements (TATA box and BRE) of the hsp5 promoter (Phsp5) did not significantly affect the basal and heat-induced gene expression, as long as the transcription initiation site was not altered. Moreover, the BRE and TATA box of Phsp5 were sufficient to render a nonheat-responsive promoter heat-inducible, in both Haloferax volcanii and Halobacterium sp. NRC-1. DNA–protein interactions revealed that two heat-inducible GTFs, TFB2 from H. volcanii and TFBb from Halobacterium sp. NRC-1, could specifically bind to Phsp5 likely in a temperature-dependent manner. Taken together, the heat-responsiveness of Phsp5 was mainly ascribed to the core promoter elements that were efficiently recognized by specific heat-induced GTFs at elevated temperature, thus providing a new paradigm for GTF-directed gene regulation in the domain of Archaea. PMID:18390887

  20. The cellular distribution of Na+/H+ exchanger regulatory factor 1 is determined by the PDZ-I domain and regulates the malignant progression of breast cancer

    PubMed Central

    Du, Guifang; Gu, Yanan; Hao, Chengcheng; Yuan, Zhu; He, Junqi; Jiang, Wen G.; Cheng, Shan

    2016-01-01

    The oncogenic role of ectopic expression of Na+/H+ exchanger regulatory factor 1 (NHERF1) was recently suggested. Here, we show that NHERF1 was upregulated in high grades compared with low grades. Increased NHERF1 expression was correlated with poor prognosis and poor survival. NHERF1 expression was higher in the nucleus of cancer cells than in contiguous non- mammary epithelial cells. A novel mutation, namely NHERF1 Y24S, was identified in human breast cancer tissues and shown to correspond to a conserved residue in the PDZ-I domain of NHERF1. Truncation and mutation of the PDZ-I domain of NHERF1 increased the nuclear distribution of the NHERF1 protein, and this redistribution was associated with the malignant phenotype of breast cancer cells, including growth, migration, and adhesion. The present results suggest a role for NHERF1 in the progression of breast cancer mediated by the nuclear distribution of the NHERF1 protein, as determined by the truncation or key site mutation of the PDZ-I domain. PMID:27097111

  1. Sterol regulatory element-binding factor 2 (SREBF-2) predicts 7-year NAFLD incidence and severity of liver disease and lipoprotein and glucose dysmetabolism.

    PubMed

    Musso, Giovanni; Cassader, Maurizio; Bo, Simona; De Michieli, Franco; Gambino, Roberto

    2013-04-01

    We prospectively assessed the impact of a sterol regulatory element-binding factor-2 (SREBF-2) polymorphism on the risk of developing nonalcoholic fatty liver disease (NAFLD) and on liver histology and lipoprotein and glucose metabolism in biopsy-proven NAFLD. In a population-based study, we followed 175 nonobese, nondiabetic participants without NAFLD or metabolic syndrome at baseline, characterized for the SREBF-2 rs133291 C/T polymorphism, dietary habits, physical activity, adipokines, C-reactive protein (CRP), and endothelial adhesion molecules. A comparable cohort of NAFLD patients underwent liver biopsy, an oral glucose tolerance test with minimal model analysis to yield glucose homeostasis parameters, and an oral fat tolerance test with measurement of plasma lipoproteins, adipokines, and cytokeratin-18 fragments. After 7 years, 27% of subjects developed NAFLD and 5% developed diabetes. SREBF-2 predicted incident NAFLD and diabetes and CRP and endothelial adhesion molecule changes. In biopsy-proven NAFLD patients, SREBF-2 predicted nonalcoholic steatohepatitis (odds ratio 2.92 [95% CI 2.08-4.18], P = 0.002) and the severity of tissue insulin resistance, β-cell dysfunction, and oral fat intolerance (characterized by higher postprandial lipemia, cholesterol enrichment of triglyceride-rich lipoproteins and oxidized LDLs, HDL cholesterol fall, adipokine imbalance, and postprandial apoptosis activation). An SREBF-2 polymorphism predisposes individuals to NAFLD and associated cardiometabolic abnormalities and affects liver histology and glucose and lipid metabolism in biopsy-proven NAFLD.

  2. Poly I:C enhances production of nitric oxide in response to interferon-γ via upregulation of interferon regulatory factor 7 in vascular endothelial cells.

    PubMed

    Mori, Daiki; Koide, Naoki; Tsolmongyn, Bilegtsaikhan; Nagata, Hiroshi; Sano, Tsuyoshi; Nonami, Toshiaki; Yokochi, Takashi

    2015-03-01

    The effect of poly I:C on interferon (IFN)-γ-induced nitric oxide (NO) production in vascular endothelial cells was examined using murine aortic endothelial END-D cells. Poly I:C augmented IFN-γ-induced NO production although it alone did not induce the NO production. Poly I:C augmented the NO production via enhanced expression of an inducible NO synthase protein. Poly I:C did not affect the activation of Janus kinase (JAK) 1/2, and signal transducer and activator of transcription (STAT) 1 in IFN-γ signaling. Moreover, there was no significant difference in the IFN-γ-induced interferon regulatory factor (IRF) 1 expression between the presence and absence of poly I:C. Poly I:C led to the activation of IRF7 in END-D cells. Inhibition of poly I:C signaling by amlexanox, an inhibitor of TANK-binding kinase (TBK) 1 and IκB kinase (IKK) ε, abolished the augmentation of IFN-γ-induced NO production. Therefore, poly I:C was suggested to augment IFN-γ-induced NO production at the transcriptional level via enhanced IRF7 activation.

  3. A Novel Sucrose-Regulatory MADS-Box Transcription Factor GmNMHC5 Promotes Root Development and Nodulation in Soybean (Glycine max [L.] Merr.).

    PubMed

    Liu, Wei; Han, Xiangdong; Zhan, Ge; Zhao, Zhenfang; Feng, Yongjun; Wu, Cunxiang

    2015-01-01

    The MADS-box protein family includes many transcription factors that have a conserved DNA-binding MADS-box domain. The proteins in this family were originally recognized to play prominent roles in floral development. Recent findings, especially with regard to the regulatory roles of the AGL17 subfamily in root development, have greatly broadened their known functions. In this study, a gene from soybean (Glycine max [L.] Merr.), GmNMHC5, was cloned from the Zigongdongdou cultivar and identified as a member of the AGL17 subfamily. Real-time fluorescence quantitative PCR analysis showed that GmNMHC5 was expressed at much higher levels in roots and nodules than in other organs. The activation of expression was first examined in leaves and roots, followed by shoot apexes. GmNMHC5 expression levels rose sharply when the plants were treated under short-day conditions (SD) and started to pod, whereas low levels were maintained in non-podding plants under long-day conditions (LD). Furthermore, overexpression of GmNMHC5 in transgenic soybean significantly promoted lateral root development and nodule building. Moreover, GmNMHC5 is upregulated by exogenous sucrose. These results indicate that GmNMHC5 can sense the sucrose signal and plays significant roles in lateral root development and nodule building. PMID:26404246

  4. Rapidly elevated levels of PGC-1α-b protein in human skeletal muscle after exercise: exploring regulatory factors in a randomized controlled trial.

    PubMed

    Gidlund, Eva-karin; Ydfors, Mia; Appel, Susanna; Rundqvist, Helene; Sundberg, Carl Johan; Norrbom, Jessica

    2015-08-15

    Individuals with high skeletal muscle mitochondrial content have a lower risk to acquire cardiovascular and metabolic disease, obesity, and type II diabetes. Regular endurance training increases mitochondrial density through a complex network of transcriptional regulators that in an accumulated way are affected by each single exercise bout. The aim of the present study was to investigate the effect of a single exercise bout on the levels of PGC-1α and related regulatory factors important for the initial phase of skeletal muscle adaptation. Ten men and ten women were randomized to either an exercise group (60 min cycling at a work load corresponding to 70% of peak oxygen uptake) or a nonexercising control group. Skeletal muscle biopsies were taken before, at 30 min, and at 2, 6, and 24 h after the intervention. Twenty-two mRNA transcripts and five proteins were measured. With exercise, protein levels of PGC-1α-ex1b increased, and this elevation occurred before that of total PGC-1α protein. We also demonstrated the existence and postexercise expression pattern of two LIPIN-1 (LIPIN-1α and LIPIN-1β) and three NCoR1 (NCoR1-1, NCoR1-2, and NCoR1-3) isoforms in human skeletal muscle. The present study contributes new insights into the initial signaling events following a single bout of exercise and emphasizes PGC-1α-ex1b as the most exercise-responsive PGC-1α isoform.

  5. Contributions of Two-Component Regulatory Systems, Alternative σ Factors, and Negative Regulators to Listeria monocytogenes Cold Adaptation and Cold Growth

    PubMed Central

    Chan, Yvonne C.; Hu, Yuewei; Chaturongakul, Soraya; Files, Kali D.; Bowen, Barbara M.; Boor, Kathryn J.; Wiedmann, Martin

    2011-01-01

    The ability of Listeria monocytogenes to grow at refrigeration temperatures is critical for transmission of this foodborne pathogen. We evaluated the contributions of different transcriptional regulators and two-component regulatory systems to L. monocytogenes cold adaptation and cold growth. L. monocytogenes parent strain 10403S and selected isogenic null mutants in genes encoding four alternative σ factors (sigB, sigH, sigC, and sigL), two regulators of σB (rsbT and rsbV), two negative regulators (ctsR and hrcA), and 15 two-component response regulators were grown in brain heart infusion broth at 4°C with (i) a high-concentration starting inoculum (108 CFU/ml), (ii) a low-concentration starting inoculum (102 CFU/ml), and (iii) a high-concentration starting inoculum of cold-adapted cells. With a starting inoculum of 108 CFU/ml, null mutants in genes encoding selected alternative σ factors (ΔsigH, ΔsigC, and ΔsigL), a negative regulator (ΔctsR), regulators of σB (ΔrsbT and ΔrsbV), and selected two-component response regulators (ΔlisR, Δlmo1172, and Δlmo1060) had significantly reduced growth (P < 0.05) compared with the parent strain after 12 days at 4°C. The growth defect for ΔsigL was limited and was not confirmed by optical density (OD600) measurement data. With a starting inoculum of 102 CFU/ml and after monitoring growth at 4°C over 84 days, only the ΔctsR strain had a consistent but limited growth defect; the other mutant strains had either no growth defects or limited growth defects apparent at only one or two of the nine sampling points evaluated during the 84-day growth period (ΔsigB, ΔsigC, and Δlmo1172). With a 108 CFU/ml starting inoculum of cold-adapted cells, none of the mutant strains that had a growth defect when inoculation was performed with cells pregrown at 37°C had reduced growth as compared with the parent strain after 12 days at 4°C, suggesting a specific defect in the ability of these mutant strains to adapt to 4

  6. Lysosomal-associated Transmembrane Protein 4B (LAPTM4B) Decreases Transforming Growth Factor β1 (TGF-β1) Production in Human Regulatory T Cells.

    PubMed

    Huygens, Caroline; Liénart, Stéphanie; Dedobbeleer, Olivier; Stockis, Julie; Gauthy, Emilie; Coulie, Pierre G; Lucas, Sophie

    2015-08-14

    Production of active TGF-β1 is one mechanism by which human regulatory T cells (Tregs) suppress immune responses. This production is regulated by glycoprotein A repetitions predominant (GARP), a transmembrane protein present on stimulated Tregs but not on other T lymphocytes (Th and CTLs). GARP forms disulfide bonds with proTGF-β1, favors its cleavage into latent inactive TGF-β1, induces the secretion and surface presentation of GARP·latent TGF-β1 complexes, and is required for activation of the cytokine in Tregs. We explored whether additional Treg-specific protein(s) associated with GARP·TGF-β1 complexes regulate TGF-β1 production in Tregs. We searched for such proteins by yeast two-hybrid assay, using GARP as a bait to screen a human Treg cDNA library. We identified lysosomal-associated transmembrane protein 4B (LAPTM4B), which interacts with GARP in mammalian cells and is expressed at higher levels in Tregs than in Th cells. LAPTM4B decreases cleavage of proTGF-β1, secretion of soluble latent TGF-β1, and surface presentation of GARP·TGF-β1 complexes by Tregs but does not contribute to TGF-β1 activation. Therefore, LAPTM4B binds to GARP and is a negative regulator of TGF-β1 production in human Tregs. It may play a role in the control of immune responses by decreasing Treg immunosuppression.

  7. Assessment of the frequency of regulatory T cells (CD4+CD25+CD127-) in children with hemophilia A: relation to factor VIII inhibitors and disease severity.

    PubMed

    El-Asrar, Mohamed Abo; Hamed, Ahmed El-Saeed; Darwish, Yasser Wagih; Ismail, Eman Abdel Rahman; Ismail, Noha Ali

    2016-01-01

    A rapidly growing evidence showed that regulatory T cells (Tregs) play a crucial role in tolerance to coagulation factors and may be involved in the pathogenesis of inhibitor formation in patients with hemophilia. We determined the percentage of Tregs (CD4CD25CD127) in 45 children with hemophilia A compared with 45 healthy controls, and assessed their relation to the clinical characteristics of patients and factor VIII (FVIII) inhibitors. Patients were studied stressing on frequency of bleeding attacks, joint pain, history of viral hepatitis, and the received therapy (FVIII precipitate/cryotherapy). FVIII activity and FVIII inhibitors were assessed with flow cytometric analysis of CD4CD25CD127 Tregs. According to residual FVIII activity levels, 30 patients (66.7%) had mild/moderate hemophilia A, whereas 15 (33.3%) patients had severe hemophilia A. The frequency of Tregs was significantly lower among all patients with hemophilia A compared with controls (2.59 ± 1.1 versus 3.73 ± 1.12%; P = 0.002). Tregs were significantly decreased among patients with FVIII inhibitors compared with the inhibitor-negative group (P < 0.001). Patients with hematuria or severe hemophilia A had lower Tregs levels than those without (P = 0.34 and P = 0.011, respectively). A significant positive correlation was found between the percentage of Tregs and FVIII among hemophilia A patients. ROC curve analysis revealed that the cut-off value of Tregs at 1.91% could differentiate patients with and without FVIII inhibitors, with a sensitivity of 100% and a specificity of 91.3%. We suggest that alteration in the frequency of Tregs in young patients with hemophilia A may contribute to inhibitor formation and disease severity.

  8. Multiple myeloma oncogene 1 (MUM1)/interferon regulatory factor 4 (IRF4) upregulates monokine induced by interferon-gamma (MIG) gene expression in B-cell malignancy.

    PubMed

    Uranishi, M; Iida, S; Sanda, T; Ishida, T; Tajima, E; Ito, M; Komatsu, H; Inagaki, H; Ueda, R

    2005-08-01

    MUM1 (multiple myeloma oncogene 1)/IRF4 (interferon regulatory factor 4) is a transcription factor that is activated as a result of t(6;14)(p25;q32) in multiple myeloma. MUM1 expression is seen in various B-cell lymphomas and predicts an unfavorable outcome in some lymphoma subtypes. To elucidate its role in B-cell malignancies, we prepared MUM1-expressing Ba/F3 cells, which proliferated until higher cellular density than the parental cells, and performed cDNA microarray analysis to identify genes whose expression is regulated by MUM1. We found that the expression of four genes including FK506-binding protein 3 (FKBP3), the monokine induced by interferon-gamma(MIG), Fas apoptotic inhibitory molecule (Faim) and Zinc-finger protein 94 was altered in the MUM1-expressing cells. We then focused on MIG since its expression was immediately upregulated by MUM1. In reporter assays, MUM1 activated the MIG promoter in cooperation with PU.1, and the interaction between MUM1 and the MIG promoter sequence was confirmed. The expression of MIG was correlated with that of MUM1 in B-CLL cell lines, and treatment with neutralizing antibodies against MIG and its receptor, CXCR3, slightly inhibited the proliferation of two MUM1-expressing lines. These results suggest that MUM1 plays roles in the progression of B-cell lymphoma/leukemia by regulating the expression of various genes including MIG. Leukemia (2005) 19, 1471-1478. doi:10.1038/sj.leu.2403833; published online 16 June 2005.

  9. Characterization and expression analyses of five interferon regulatory factor transcripts (Irf4a, Irf4b, Irf7, Irf8, Irf10) in Atlantic cod (Gadus morhua).

    PubMed

    Inkpen, Sabrina M; Hori, Tiago S; Gamperl, A Kurt; Nash, Gordon W; Rise, Matthew L

    2015-05-01

    The interferon regulatory factor (IRF) family of genes encodes a group of transcription factors that have important roles not only in regulating the expression of Type I interferons (IFNs) and other genes in the IFN pathway, but also in growth, development and the regulation of oncogenesis. In this study, several IRF family members (Irf4a, Irf4b, Irf7, Irf8, Irf10) in Atlantic cod (Gadus morhua) were characterized at the cDNA and putative amino acid levels, allowing for phylogenetic analysis of these proteins in teleost fish, as well as the development of gene-specific primers used in RT-PCR and quantitative PCR (QPCR) analyses. Two Atlantic cod Irf10 splice variants were identified and their presence confirmed by sequencing of the Irf10 genomic region. RT-PCR showed that Irf7, Irf8 and both Irf10 transcripts were expressed in all 15 cod tissues tested, while Irf4a and Irf4b were absent in some tissues. QPCR analysis of spleen expression expanded upon this, and upon previous work. All IRF transcripts in the study were responsive to stimulation by the viral mimic poly(I:C), and all except Irf4a were responsive to exposure to formalin-killed Aeromonas salmonicida (ASAL). These IRF genes, thus, are likely important in the cod immune response to both viral and bacterial infections. Increased temperature (10 °C to 16 °C) was also observed to modulate the antibacterial responses of all IRF transcripts, and the antiviral responses of Irf4b and Irf10-v2. This research supports earlier studies which reported that elevated temperature modulates the expression of many immune genes in Atlantic cod. PMID:25731920

  10. Gene structures and promoter characteristics of interferon regulatory factor 1 (IRF-1), IRF-2 and IRF-7 from snakehead Channa argus.

    PubMed

    Jia, Weizhang; Guo, Qionglin

    2008-04-01

    Three interferon regulatory factor (IRF) genes, CaIRF-1, CaIRF-2 and CaIRF-7, and their promoters of snakehead (Channa argus) were cloned and characterized. The CaIRF-1 gene consists of ten exons, spans 4.3 kb and encodes a putative peptide of 299 aa. The CaIRF-2 gene consists of nine exons, spans 8 kb and encodes a putative peptide of 328 aa. The gene organizations of CaIRF-1 and CaIRF-2 are very similar to that of human IRF-1 and IRF-2 except more compact. Comparison of exon-intron organization of the two genes indicated a common evolutionary structure, notably within the exons encoding the DNA binding domain (DBD) of the two factors. The CaIRF-7 gene spans 4.1 kb and encodes a putative peptide of 437 aa. However, the gene organization of CaIRF-7 consisting of ten exons is different to human IRF-7a gene which has an intron in 5' UTR. Three CaIRFs share homology in N-terminal encompassing the DBD that contains a characteristic repeat of tryptophan residues. The promoters of CaIRF-1 and CaIRF-2 genes contain the conserved sites for NF-kappaB and Sp1. The gamma-IFN activation sites (GAS) were found in the promoters of CaIRF-1 and CaIRF-7. The promoter of CaIRF-7 contains conserved interferon stimulating response element (ISRE) which is characteristic of IFN-induced gene promoter, and suggests that there also exist intracellular amplifier circuit in fish IFN signal pathway. Moreover, the element GAAANN oriented in both directions is repeated in CaIRF promoter regions, which confers to further inducibility by IFN. The constitutive expression of CaIRF genes were found to increase obviously in response to induction by the known IFN-inducer poly I:C.

  11. Differential Regulation of Interferon Regulatory Factor (IRF)-7 and IRF-9 Gene Expression in the Central Nervous System during Viral Infection

    PubMed Central

    Ousman, Shalina S.; Wang, Jianping; Campbell, Iain L.

    2005-01-01

    Interferon regulatory factors (IRFs) are a family of transcription factors involved in the regulation of the interferons (IFNs) and other genes that may have an essential role in antiviral defense in the central nervous system, although this is currently not well defined. Therefore, we examined the regulation of IRF gene expression in the brain during viral infection. Several IRF genes (IRF-2, -3, -5, -7, and -9) were expressed at low levels in the brain of uninfected mice. Following intracranial infection with lymphocytic choriomeningitis virus (LCMV), expression of the IRF-7 and IRF-9 genes increased significantly by day 2. IRF-7 and IRF-9 gene expression in the brain was widespread at sites of LCMV infection, with the highest levels in infiltrating mononuclear cells, microglia/macrophages, and neurons. IRF-7 and IRF-9 gene expression was increased in LCMV-infected brain from IFN-γ knockout (KO) but not IFN-α/βR KO animals. In the brain, spleen, and liver or cultured glial and spleen cells, IRF-7 but not IRF-9 gene expression increased with delayed kinetics in the absence of STAT1 but not STAT2 following LCMV infection or IFN-α treatment, respectively. The stimulation of IRF-7 gene expression by IFN-α in glial cell culture was prevented by cycloheximide. Thus, (i) many of the IRF genes were expressed constitutively in the mouse brain; (ii) the IRF-7 and IRF-9 genes were upregulated during viral infection, a process dependent on IFN-α/β but not IFN-γ; and (iii) IRF-7 but not IRF-9 gene expression can be stimulated in a STAT1-independent but STAT2-dependent fashion via unidentified indirect pathways coupled to the activation of the IFN-α/β receptor. PMID:15919906

  12. Mapping the Regulatory Network for Salmonella enterica Serovar Typhimurium Invasion

    PubMed Central

    Smith, Carol; Stringer, Anne M.; Mao, Chunhong; Palumbo, Michael J.

    2016-01-01

    ABSTRACT Salmonella enterica pathogenicity island 1 (SPI-1) encodes proteins required for invasion of gut epithelial cells. The timing of invasion is tightly controlled by a complex regulatory network. The transcription factor (TF) HilD is the master regulator of this process and senses environmental signals associated with invasion. HilD activates transcription of genes within and outside SPI-1, including six other TFs. Thus, the transcriptional program associated with host cell invasion is controlled by at least 7 TFs. However, very few of the regulatory targets are known for these TFs, and the extent of the regulatory network is unclear. In this study, we used complementary genomic approaches to map the direct regulatory targets of all 7 TFs. Our data reveal a highly complex and interconnected network that includes many previously undescribed regulatory targets. Moreover, the network extends well beyond the 7 TFs, due to the inclusion of many additional TFs and noncoding RNAs. By comparing gene expression profiles of regulatory targets for the 7 TFs, we identified many uncharacterized genes that are likely to play direct roles in invasion. We also uncovered cross talk between SPI-1 regulation and other regulatory pathways, which, in turn, identified gene clusters that likely share related functions. Our data are freely available through an intuitive online browser and represent a valuable resource for the bacterial research community. PMID:27601571

  13. Influence of the season on vitamin D levels and regulatory T cells in patients with polymorphic light eruption† †Electronic supplementary information (ESI) available. See DOI: 10.1039/c5pp00398a Click here for additional data file. Click here for additional data file.

    PubMed Central

    Schweintzger, N. A.; Gruber-Wackernagel, A.; Shirsath, N.; Quehenberger, F.; Obermayer-Pietsch, B.

    2016-01-01

    The exact mechanisms of photohardening in polymorphic light eruption (PLE) are still unknown, but medical photohardening was shown to increase regulatory T cell (Treg) numbers in the blood of PLE patients, similar to natural hardening. Furthermore, oral vitamin D supplementation increased peripheral Tregs in healthy individuals. We herein report on a post hoc analysis of 26 screened PLE patients of a clinical trial (ClinicalTrials.gov No. NCT01595893), in which the influence of the progressing season was investigated on baseline CD4+CD25+FoxP3+CD127– Treg numbers by flow cytometry and Treg suppressive function by co-culture assays with T effector cells as a secondary endpoint, together with 25-hydroxy vitamin D (25(OH)D) serum levels at the study's screening visit, taking place in the period from January to June. The mean 25(OH)D serum level of all patients was 33.2 ng ml–1. Ten of those patients (38.5%) were identified with low 25(OH)D levels (<30 ng ml–1). Significantly higher baseline 25(OH)D serum levels (plus 34.4%; P = 0.0182) as well as higher relative Treg percentages in CD4+ population (plus 62.8%; P = 0.0157) and in total lymphocyte population (plus 59.6%; P = 0.0372) and higher absolute Treg numbers (plus 100.2%; P = 0.0042) were observed in the late spring/early summer period (April to June) compared to the winter period (January to February). No significant relationship was observed when Treg numbers and function were correlated with 25(OH)D levels. These data indicate that in PLE patients Treg numbers and their suppressive function are independent of vitamin D serum levels and suggest that UV light and/or other seasonal factors may affect these cells via the non-vitamin D related pathway(s). PMID:26911519

  14. Optimization of codon composition and regulatory elements for expression of human insulin like growth factor-1 in transgenic chloroplasts and evaluation of structural identity and function

    PubMed Central

    Daniell, Henry; Ruiz, Gricel; Denes, Bela; Sandberg, Laurence; Langridge, William

    2009-01-01

    Background Transgenic chloroplasts are potential bioreactors for recombinant protein production, especially for achievement of high levels of protein expression and proper folding. Production of therapeutic proteins in leaves provides transgene containment by elimination of reproductive structures. Therefore, in this study, human Insulin like Growth Factor-1 is expressed in transgenic chloroplasts for evaluation of structural identity and function. Results Expression of the synthetic Insulin like Growth Factor 1 gene (IGF-1s, 60% AT) was observed in transformed E. coli. However, no native IGF-1 gene (IGF-1n, 41% AT) product was detected in the western blots in E. coli. Site-specific integration of the transgenes into the tobacco chloroplast genome was confirmed after transformation using PCR. Southern blot analysis confirmed that the transgenic lines were homoplasmic. The transgenic plant lines had IGF-1s expression levels of 11.3% of total soluble protein (TSP). The IGF-1n plants contained 9.5% TSP as IGF-1n, suggesting that the chloroplast translation machinery is more flexible than E. coli in codon preference and usage. The expression of IGF-1 was increased up to 32% TSP under continuous illumination by the chloroplast light regulatory elements. IgG-Sepharose affinity column chromatographic separation of Z domain containing chloroplast derived IGF-1 protein, single and two dimensional electrophoresis methods and mass spectrometer analysis confirmed the identity of human IGF-1 in transgenic chloroplasts. Two spots analyzed from 2-D focusing/phoresis acrylamide gel showed the correct amino acid sequence of human IGF-1 and the S. aureus Z-tag. Cell proliferation assays in human HU-3 cells demonstrated the biological activity of chloroplast derived IGF-1 even in the presence of the S. aureus Z tag. Conclusion This study demonstrates that the human Insulin like Growth Factor-1 expressed in transgenic chloroplasts is identical to the native protein and is fully

  15. Conservation and evolution of cis-regulatory systems in ascomycete fungi

    SciTech Connect

    Gasch, Audrey P.; Moses, Alan M.; Chiang, Derek Y.; Fraser, Hunter B.; Berardini, Mark; Eisen, Michael B.

    2004-03-15

    Relatively little is known about the mechanisms through which gene expression regulation evolves. To investigate this, we systematically explored the conservation of regulatory networks in fungi by examining the cis-regulatory elements that govern the expression of coregulated genes. We first identified groups of coregulated Saccharomyces cerevisiae genes enriched for genes with known upstream or downstream cis-regulatory sequences. Reasoning that many of these gene groups are coregulated in related species as well, we performed similar analyses on orthologs of coregulated S. cerevisiae genes in 13 other ascomycete species. We find that many species-specific gene groups are enriched for the same flanking regulatory sequences as those found in the orthologous gene groups from S. cerevisiae, indicating that those regulatory systems have been conserved in multiple ascomycete species. In addition to these clear cases of regulatory conservation, we find examples of cis-element evolution that suggest multiple modes of regulatory diversification, including alterations in transcription factor-binding specificity, incorporation of new gene targets into an existing regulatory system, and cooption of regulatory systems to control a different set of genes. We investigated one example in greater detail by measuring the in vitro activity of the S. cerevisiae transcription factor Rpn4p and its orthologs from Candida albicans and Neurospora crassa. Our results suggest that the DNA binding specificity of these proteins has coevolved with the sequences found upstream of the Rpn4p target genes and suggest that Rpn4p has a different function in N. crassa.

  16. The regulatory mechanism of fruit ripening revealed by analyses of direct targets of the tomato MADS-box transcription factor RIPENING INHIBITOR.

    PubMed

    Fujisawa, Masaki; Ito, Yasuhiro

    2013-06-01

    The developmental process of ripening is unique to fleshy fruits and a key factor in fruit quality. The tomato (Solanum lycopersicum) MADS-box transcription factor RIPENING INHIBITOR (RIN), one of the earliest-acting ripening regulators, is required for broad aspects of ripening, including ethylene-dependent and -independent pathways. However, our knowledge of direct RIN target genes has been limited, considering the broad effects of RIN on ripening. In a recent work published in The Plant Cell, we identified 241 direct RIN target genes by chromatin immunoprecipitation coupled with DNA microarray (ChIP-chip) and transcriptome analysis. Functional classification of the targets revealed that RIN participates in the regulation of many biological processes including well-known ripening processes such as climacteric ethylene production and lycopene accumulation. In addition, we found that ethylene is required for the full expression of RIN and several RIN-targeting transcription factor genes at the ripening stage. Here, based on our recently published findings and additional data, we discuss the ripening processes regulated by RIN and the interplay between RIN and ethylene.

  17. Smad7 protein induces interferon regulatory factor 1-dependent transcriptional activation of caspase 8 to restore tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis.

    PubMed

    Hong, Suntaek; Kim, Hye-Youn; Kim, Jooyoung; Ha, Huyen Trang; Kim, Young-Mi; Bae, Eunjin; Kim, Tae Hyung; Lee, Kang Choon; Kim, Seong-Jin

    2013-02-01

    Smad7 has been known as a negative regulator for the transforming growth factor-β (TGF-β) signaling pathway through feedback regulation. However, Smad7 has been suspected to have other biological roles through the regulation of gene transcription. By screening differentially regulated genes, we found that the caspase 8 gene was highly up-regulated in Smad7-expressing cells. Smad7 was able to activate the caspase 8 promoter through recruitment of the interferon regulatory factor 1 (IRF1) transcription factor to the interferon-stimulated response element (ISRE) site. Interaction of Smad7 on the caspase 8 promoter was confirmed with electrophoretic mobility shift assay and chromatin immunoprecipitation experiment. Interestingly, Smad7 did not directly interact with the ISRE site, but it increased the binding activity of IRF1 with ISRE. These results support that Smad7 recruits IRF1 protein on the caspase 8 promoter and functions as a transcriptional coactivator. To confirm the biological significance of caspase 8 up-regulation, we tested tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL)-mediated cell death assay in breast cancer cells. Smad7 in apoptosis-resistant MCF7 cells markedly sensitized the cells to TRAIL-induced cell death by restoring the caspase cascade. Furthermore, restoration of caspase 8-mediated apoptosis pathway repressed the tumor growth in the xenograft model. In conclusion, we suggest a novel role for Smad7 as a transcriptional coactivator for caspase 8 through the interaction with IRF1 in regulation of the cell death pathway. PMID:23255602

  18. Analysis of Air Toxics From NOAA WP-3 Aircraft Measurements During the TexAQS 2006 Campaign: Comparison With Emission Inventories and Additive Inhalation Risk Factors

    NASA Astrophysics Data System (ADS)

    Del Negro, L. A.; Warneke, C.; de Gouw, J. A.; Atlas, E.; Lueb, R.; Zhu, X.; Pope, L.; Schauffler, S.; Hendershot, R.; Washenfelder, R.; Fried, A.; Richter, D.; Walega, J. G.; Weibring, P.

    2007-12-01

    Benzene and nine other air toxics classified as human carcinogens by the International Agency for Research on Cancer (IARC) were measured from the NOAA WP-3 aircraft during the TexAQS 2006 campaign. In-situ measurements of benzene, measured with a PTR-MS instrument, are used to estimate emission fluxes for comparison with point source emission inventories developed by the Texas Commission on Environmental Quality. Mean and median mixing ratios for benzene, acetaldehyde, formaldehyde, 1,3-butadiene, carbon tetrachloride, chloroform, 1,2-dichloroethane, dibromoethane, dichloromethane, and vinyl chloride, encountered over the city of Houston during the campaign, are combined with inhalation unit risk factor values developed by the California Environmental Protection Agency and the United States Environmental Protection Agency to estimate the additive inhalation risk factor. This additive risk factor represents the risk associated with lifetime (70 year) exposure at the levels measured and should not be used as an absolute indicator of risk to individuals. However, the results are useful for assessments of changing relative risk over time, and for identifying dominant contributions to the overall air toxic risk.

  19. Interferon regulatory factor 4b (IRF4b) in Japanese flounder, Paralichthys olivaceus: Sequencing, ubiquitous tissue distribution and inducible expression by poly(I:C) and DNA virus.

    PubMed

    Liu, Dahai; Chen, Jinjing; Zhang, Haiyan; Hu, Mengzhu; Lou, Huimin; Liu, Qiuming; Zhang, Shicui; Hu, Guobin

    2016-09-01

    Interferon regulatory factor 4 (IRF4) in mammals is known to be critical in regulation of development and functions of lymphomyeloid cell lineages. Recent studies have demonstrated its involvement in immune responses to bacterial and viral challenges in teleosts. In this study, an IRF4 gene was cloned from Japanese flounder (Paralichthys olivaceus) and its expression in response to polyinosinic:polycytidylic acid [poly(I:C)] and lymphocystis disease virus (LCDV) stimulations was studied in vivo. The cloned gene spans over 5.9 kb, comprises eight exons and seven introns and encodes a putative protein of 456 amino acids. The deduced amino acid sequence possesses a conserved DNA-binding domain (DBD), an IRF-association domain (IAD) and a nuclear localization signal (NLS). Phylogenetic analysis clustered it into the teleost IRF4b clade and, thus, it was named Paralichthys olivaceus (Po)IRF4b. The constitutive expression of PoIRF4b transcripts was detectable in all examined organs, with highest levels found in lymphomyeloid-rich tissues. They were induced by both poly(I:C) and LCDV with a similar inducibility in immune or non-immune organs. Two waves of induced expression of PoIRF4b were observed with the two stimuli during a 7-day time course in the immune organs, with the early-phase induction being stronger. The maximum increases of PoIRF4b transcript levels ranged from 1.3 to 4.0-fold and appeared at day 1-5 post-injection depending on different organs and stimuli. In both stimulation cases, the strongest induction was detected in spleen and the weakest in muscle. These results indicate that PoIRF4b may participate in regulation of immune responses of flounders to both RNA and DNA virus infections. PMID:27084058

  20. Muscle hypertrophy in heavy weight Japanese quail line: delayed muscle maturation and continued muscle growth with prolonged upregulation of myogenic regulatory factors.

    PubMed

    Choi, Y M; Suh, Y; Ahn, J; Lee, K

    2014-09-01

    The objective of this study was to compare the temporal expression of myosin heavy chain (MyHC) isoforms, Pax7, and myogenic regulatory factors (MRF) between heavy weight (HW) and random bred control (RBC) Japanese quail lines during muscle development to better understand the mechanisms leading to increased skeletal muscle mass in the HW quail line selected for a greater BW at 4 wk of age separated from RBC quail. Expression of neonatal MyHC isoform began at 3 and 7 d posthatch in RBC and HW quail lines, respectively. In the RBC quail line, adult MyHC isoform, as a marker for muscle maturation, was expressed at 28 d posthatch with sustained expression through 75 d posthatch, whereas this protein was detected only at 75 d posthatch in the HW quail line. Moreover, Pax7 expression continued from embryonic ages to 14 d posthatch in the HW quail line and to 7 d posthatch in the RBC quail line. These expression patterns of MyHC isoforms and Pax7 in the HW quail line were accompanied by delayed muscle maturation and prolonged growth compared with the RBC quail line. Temporal expressions of the primary MRF showed that higher expression levels of MyoD and Myf-5 were observed at 9 and 11 d embryo in the HW quail line compared with the RBC quail line (P < 0.05). The HW quail line exhibited approximately 2 times greater average levels of myogenin expression from 7 to 75 d posthatch (P < 0.05) than the RBC quail line. Prolonged upregulation of these primary and secondary MRF during muscle development is associated with delayed maturation and continued muscle growth, which consequently would permit muscle hypertrophic potentials in the HW quail line compared with the RBC quail line.

  1. Phytoalexin-Deficient Mutants of Arabidopsis Reveal That Pad4 Encodes a Regulatory Factor and That Four Pad Genes Contribute to Downy Mildew Resistance

    PubMed Central

    Glazebrook, J.; Zook, M.; Mert, F.; Kagan, I.; Rogers, E. E.; Crute, I. R.; Holub, E. B.; Hammerschmidt, R.; Ausubel, F. M.

    1997-01-01

    We are working to determine the role of the Arabidopsis phytoalexin, camalexin, in protecting the plant from pathogen attack by isolating phytoalexin-deficient (pad) mutants in the accession Columbia (Col-0) and examining their response to pathogens. Mutations in PAD1, PAD2, and PAD4 caused enhanced susceptibility to the bacterial pathogen Pseudomonas syringae pv. maculicola strain ES4326 (PsmES4326), while mutations in PAD3 or PAD5 did not. Camalexin was not detected in any of the double mutants pad1-1 pad2-1, pad1-1 pad3-1 or pad2-1 pad3-1. Growth of PsmES4326 in pad1-1 pad2-1 was greater than that in pad1-1 or pad2-1 plants, while growth in pad1-1 pad3-1 and pad2-1 pad3-1 plants was similar to that in pad1-1 and pad2-1 plants, respectively. The pad4-1 mutation caused reduced camalexin synthesis in response to PsmES4326 infection, but not in response to Cochliobolus carbonum infection, indicating that PAD4 has a regulatory function. PAD1, PAD2, PAD3 and PAD4 are all required for resistance to the eukaryotic biotroph Peronospora parasitica. The pad4-1 mutation caused the most dramatic change, exhibiting full susceptibility to four of six Col-incompatible parasite isolates. Interestingly, each combination of double mutants between pad1-1, pad2-1 and pad3-1 exhibited additive shifts to moderate or full susceptibility to most of the isolates. PMID:9136026

  2. Regulatory RNAs in photosynthetic cyanobacteria.

    PubMed

    Kopf, Matthias; Hess, Wolfgang R

    2015-05-01

    Regulatory RNAs play versatile roles in bacteria in the coordination of gene expression during various physiological processes, especially during stress adaptation. Photosynthetic bacteria use sunlight as their major energy source. Therefore, they are particularly vulnerable to the damaging effects of excess light or UV irradiation. In addition, like all bacteria, photosynthetic bacteria must adapt to limiting nutrient concentrations and abiotic and biotic stress factors. Transcriptome analyses have identified hundreds of potential regulatory small RNAs (sRNAs) in model cyanobacteria such as Synechocystis sp. PCC 6803 or Anabaena sp. PCC 7120, and in environmentally relevant genera such as Trichodesmium, Synechococcus and Prochlorococcus. Some sRNAs have been shown to actually contain μORFs and encode short proteins. Examples include the 40-amino-acid product of the sml0013 gene, which encodes the NdhP subunit of the NDH1 complex. In contrast, the functional characterization of the non-coding sRNA PsrR1 revealed that the 131 nt long sRNA controls photosynthetic functions by targeting multiple mRNAs, providing a paradigm for sRNA functions in photosynthetic bacteria. We suggest that actuatons comprise a new class of genetic elements in which an sRNA gene is inserted upstream of a coding region to modify or enable transcription of that region.

  3. Identification of orange-spotted grouper (Epinephelus coioides) interferon regulatory factor 3 involved in antiviral immune response against fish RNA virus.

    PubMed

    Huang, Youhua; Huang, Xiaohong; Cai, Jia; OuYang, Zhengliang; Wei, Shina; Wei, Jingguang; Qin, Qiwei

    2015-02-01

    Interferon regulatory factor 3 (IRF3) is an important transcription factor which regulates the expression of interferon (IFN) and IFN-stimulated genes (ISGs) following virus recognition. In this study, a novel IRF3 gene was cloned from grouper Epinephelus coioides (EcIRF3) and its effects against Singapore grouper iridovirus (SGIV) and red spotted grouper nervous necrosis virus (RGNNV) was investigated. The full-length of EcIRF3 cDNA was composed of 2513 bp and encoded a polypeptide of 458 amino acids which shared 82% identity with European seabass (Dicentrarchus labrax). EcIRF3 contained three conserved domains including a DNA-binding domain (DBD), an IRF associated domain (IAD) and a serine-rich domain. Expression profile analysis revealed that EcIRF3 was abundant in head kidney, kidney, spleen and gill. Upon different stimuli in vitro, the transcript of EcIRF3 was significantly up-regulated after RGNNV infection or treatment with polyinosin-polycytidylic acid (poly I:C). During SGIV infection, the increase of the EcIRF3 transcription was only detected at the late stage, suggesting that EcIRF3 was differently regulated by different stimuli. Immune fluorescence assay indicated that the fluorescence signal of EcIRF3 was increased significantly after infection with RGNNV or treatment with poly I:C, but moderately at the late stage of SGIV infection. Reporter gene assay showed that EcIRF3 activated zebrafish type I IFN and type III IFN promoter in vitro. The viral gene transcription and virus production of RGNNV were significantly decreased in EcIRF3 overexpressing cells. However, the ectopic expression of EcIRF3 did not affect the gene transcription and virus production of SGIV. Moreover, the mRNA expression levels of type I IFN and IFN-inducible genes (MxI, ISG15 and ISG56) were increased in RGNNV infected EcIRF3 overexpressing cells compared to empty vector transfected cells. Together, our results demonstrated that IFN immune response mediated by grouper IRF3 was