scholarly journals Somatostatin gene upstream enhancer element activated by a protein complex consisting of CREB, Isl-1-like, and alpha-CBF-like transcription factors.

1992 ◽  
Vol 267 (18) ◽  
pp. 12876-12884 ◽  
Author(s):  
M Vallejo ◽  
L Penchuk ◽  
J.F. Habener
Keyword(s):  
Transcription Factors ◽  
Protein Complex ◽  
Enhancer Element

scholarly journals Transcription of the dominant-negative helix-loop-helix protein Id1 is regulated by a protein complex containing the immediate-early response gene Egr-1.

1996 ◽  
Vol 16 (5) ◽  
pp. 2418-2430 ◽  
Author(s):  
O Tournay ◽  
R Benezra
Keyword(s):  
Cell Cycle ◽  
Transcription Factors ◽  
Protein Complex ◽  
Early Response ◽  
Dominant Negative ◽  
Upstream Region ◽  
Response Gene ◽  
Enhancer Element ◽  
Early Response Gene ◽  
Helix Loop Helix

The expression of Id1, a helix-loop-helix protein which inhibits the activity of basic helix-loop-helix transcription factors, is down-regulated during cellular differentiation and cell cycle withdrawal both in tissue culture models and in mouse embryos. In order to study the mechanism of control of Idl expression, we have isolated a 210-bp enhancer element in the upstream region of the Id1 gene whose activity recapitulates Id1 expression in C2C12 muscle cells and C3H10T1/2 fibroblasts: i.e., this element is active in proliferating cells in the presence of serum and completely inactivated upon mitogen depletion, cell cycle withdrawal, and (in the case of C2C12) induced myoblast differentiation. Using linker-scanning mutations and site-directed mutagenesis in transient transfection experiments, we have identified two functional elements within the 210-bp enhancer which are required for proper serum responsiveness. One element (A) contains a consensus Egr-1 binding site and additional flanking sequences required for optimal activity, and the other element (B) fits no known consensus. Gel shift experiments demonstrate that the protein complex binding to the A site contains Egr-1 and other proteins. This complex as well as a protein complex that binds to the B site is lost within 24 h of serum depletion, correlating with the down-regulation of Id1 expression. On the basis of these findings, we propose that the regulation of the Id1 response to serum is mediated in part by the early response gene Egr-1 and as such provides a signaling link between the early-growth-response transcription factors and dominant-negative helix-loop-helix proteins.

scholarly journals Multi-omic profiling of pituitary thyrotropic cells and progenitors

BMC Biology ◽  
2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Alexandre Z. Daly ◽  
Lindsey A. Dudley ◽  
Michael T. Peel ◽  
Stephen A. Liebhaber ◽  
Stephen C. J. Parker ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Pituitary Gland ◽  
Cell Lines ◽  
Binding Sites ◽  
Critical Factor ◽  
Regulatory Elements ◽  
Thyroid Stimulating Hormone ◽  
Neuroendocrine Cells ◽  
Bzip Transcription Factor ◽  
Enhancer Element

Abstract Background The pituitary gland is a neuroendocrine organ containing diverse cell types specialized in secreting hormones that regulate physiology. Pituitary thyrotropes produce thyroid-stimulating hormone (TSH), a critical factor for growth and maintenance of metabolism. The transcription factors POU1F1 and GATA2 have been implicated in thyrotrope fate, but the transcriptomic and epigenomic landscapes of these neuroendocrine cells have not been characterized. The goal of this work was to discover transcriptional regulatory elements that drive thyrotrope fate. Results We identified the transcription factors and epigenomic changes in chromatin that are associated with differentiation of POU1F1-expressing progenitors into thyrotropes using cell lines that represent an undifferentiated Pou1f1 lineage progenitor (GHF-T1) and a committed thyrotrope line that produces TSH (TαT1). We compared RNA-seq, ATAC-seq, histone modification (H3K27Ac, H3K4Me1, and H3K27Me3), and POU1F1 binding in these cell lines. POU1F1 binding sites are commonly associated with bZIP transcription factor consensus binding sites in GHF-T1 cells and Helix-Turn-Helix (HTH) or basic Helix-Loop-Helix (bHLH) factors in TαT1 cells, suggesting that these classes of transcription factors may recruit or cooperate with POU1F1 binding at unique sites. We validated enhancer function of novel elements we mapped near Cga, Pitx1, Gata2, and Tshb by transfection in TαT1 cells. Finally, we confirmed that an enhancer element near Tshb can drive expression in thyrotropes of transgenic mice, and we demonstrate that GATA2 enhances Tshb expression through this element. Conclusion These results extend the ENCODE multi-omic profiling approach to the pituitary gland, which should be valuable for understanding pituitary development and disease pathogenesis. Graphical abstract

scholarly journals Signalling assemblies: the odds of symmetry

2017 ◽  
Vol 45 (3) ◽  
pp. 599-611 ◽  
Author(s):  
Gábor Maksay ◽  
Joseph A. Marsh
Keyword(s):  
Transcription Factors ◽  
Structure Determination ◽  
Protein Complex ◽  
Protein Complexes ◽  
Asymmetric Structures ◽  
Traditional Structure ◽  
Dominant Feature ◽  
Transmembrane Channels ◽  
Determination Methods

The assembly of proteins into complexes is fundamental to nearly all biological signalling processes. Symmetry is a dominant feature of the structures of experimentally determined protein complexes, observed in the vast majority of homomers and many heteromers. However, some asymmetric structures exist, and asymmetry also often forms transiently, intractable to traditional structure determination methods. Here, we explore the role of protein complex symmetry and asymmetry in cellular signalling, focusing on receptors, transcription factors and transmembrane channels, among other signalling assemblies. We highlight a recurrent tendency for asymmetry to be crucial for signalling function, often being associated with activated states. We conclude with a discussion of how consideration of protein complex symmetry and asymmetry has significant potential implications and applications for pharmacology and human disease.

scholarly journals The epigenetic signature of CFTR expression is co-ordinated via chromatin acetylation through a complex intronic element

2007 ◽  
Vol 408 (3) ◽  
pp. 317-326 ◽  
Author(s):  
Thankam Paul ◽  
SiDe Li ◽  
Sanjeev Khurana ◽  
Neal S. Leleiko ◽  
Martin J. Walsh
Keyword(s):  
Transcription Factors ◽  
Cultured Cells ◽  
Therapeutic Potential ◽  
Gastrointestinal Disease ◽  
Regulatory Element ◽  
Cell Types ◽  
Intestinal Cell ◽  
Enhancer Element ◽  
Histone Deacetylase Inhibition ◽  
Regulatory Factors

The CFTR (cystic fibrosis transmembrane conductance regulator) gene is a tightly regulated and differentially expressed transcript in many mucosal epithelial cell types. It appears that DNA sequence variations alone do not explain CFTR-related gastrointestinal disease patterns and that epigenetic modifiers influence CFTR expression. Our aim was to characterize the native chromatin environment in cultured cells for intestinal CFTR expression by determining the relationship between histone acetylation and occupation of CFTR by multiple transcription factors, through a common regulatory element. We used HDAC (histone deacetylase) inhibition and ChIP (chromatin immunoprecipitation) analyses to define regions associated with acute acetylation of histone at the CFTR locus. We identified a region within the first intron associated with acute acetylation of histone H4 as an epigenetic signature corresponding to an intestine-specific enhancer element for CFTR. DHS (DNase I-hypersensitivity) assays and ChIP were used to specify control elements and occupation by regulatory factors. Quantitative ChIP procedures indicate that HNF1α (hepatic nuclear factor 1α) and Cdx2 (caudal homeobox protein 2) occupy and regulate through a novel intronic enhancer element of CFTR and that Tcf4 (T-cell factor 4) overlaps the same DNA element. RNAi (RNA interference) of Tcf4 and HNF1α decreased intestinal cell CFTR expression, identifying these as positive regulatory factors and CFTR as a target for Wnt signalling. We have linked the acetylation signature of nucleosomal histones to active intestinal CFTR expression and occupation by transcription factors HNF1α, Cdx2 and Tcf4 which converge to modify chromatin architecture. These studies suggest the therapeutic potential of histone modification strategies, such as inhibition of HDAC activity, to treat CFTR-associated disease by selectively enhancing CFTR expression.

scholarly journals The embryonic enhancer-binding protein SSAP contains a novel DNA-binding domain which has homology to several RNA-binding proteins.

1995 ◽  
Vol 15 (3) ◽  
pp. 1254-1264 ◽  
Author(s):  
D J DeAngelo ◽  
J DeFalco ◽  
L Rybacki ◽  
G Childs
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Sea Urchin ◽  
Rna Processing ◽  
Histone H1 ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Blastula Stage ◽  
Enhancer Element ◽  
Specific Manner

Stage-specific activator protein (SSAP) is a 43-kDa polypeptide that binds to an enhancer element of the sea urchin late histone H1 gene. This enhancer element mediates the transcriptional activation of the late histone H1 gene in a temporally specific manner at the mid-blastula stage of embryogenesis. We have cloned cDNAs encoding SSAP by using polyclonal antibodies raised against purified SSAP to screen expression libraries. SSAP is unrelated to previously characterized transcription factors; however, it exhibits striking homology to a large family of proteins involved in RNA processing. The protein is a sequence-specific DNA-binding protein that recognizes both single- and double-stranded DNA. The DNA-binding domain of the protein was localized to the conserved RNA recognition motif (RRM). In addition to tandem copies of this conserved domain, SSAP contains a central domain that is rich in glutamine and glycine and a C-terminal domain that is enriched in serine, threonine, and basic amino acids. Overexpression of SSAP in sea urchin embryos by microinjection of either synthetic mRNA or an SSAP expression vector results in four- to eightfold transactivation of target reporter genes that contain the enhancer sequence. Transactivation occurs beginning only at the mid-blastula stage of development, suggesting that SSAP must be modified in a stage-specific manner in order to activate transcription. In addition, there are a number of other RRM-containing proteins that contain glutamine-rich regions which are postulated to function in the regulation of RNA processing. Instead, we suggest that SSAP is a member of a family of glutamine-rich RRM proteins which constitute a novel class of transcription factors.

scholarly journals MON-723 Identification of Thyrotrope Signature Genes and Regulatory Elements

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Alexandre Daly ◽  
Leonard Cheung ◽  
Michelle Brinkmeier ◽  
Sally Ann Camper
Keyword(s):  
Gene Expression ◽  
Risk Factors ◽  
Transcription Factors ◽  
Cell Line ◽  
Cell Lines ◽  
Binding Sites ◽  
Regulatory Elements ◽  
Genome Wide Association Studies ◽  
Expression Data ◽  
Enhancer Element

Abstract Recent genome wide association studies have begun to identify loci that are risk factors for sporadic pituitary adenomas, but the genes associated with these loci are unknown. In general, ~90% of GWAS hits are in noncoding regions, making it difficult to transition from genetic mapping to a biological understanding of risk factors. Recent studies that identify enhancer regions by undertaking large scale functional genomic annotation of non-coding elements like Encyclopedia of DNA Elements (ENCODE) have begun to yield a better understanding of some complex diseases. Dense molecular profiling maps of the transcriptome and epigenome have been generated for more than 250 cell lines and 150 tissues, but pituitary cell lines or tissues were not included. Epigenetic and gene expression data are emerging for somatotropes, gonadotropes and corticotropes, but there is very little available data on thyrotropes. We identified the transcription factors and epigenetic changes in chromatin that are associated with differentiation of POU1F1-expressing progenitors into thyrotropes using cell lines that represent an early, undifferentiated Pou1f1 lineage progenitor (GHF-T1) and a committed thyrotrope (TαT1). TαT1 is an excellent cell line for this purpose because it responds to TRH, retinoids, and secretes TSH in response to diurnal cues. We have also used genetic labeling and fluorescence activated cell sorting to purify thyrotropes from pituitaries of young mice and analyzed gene expression using single cell transcriptomics. We used the Assay for TransposaseAccessible Chromatin with sequencing (ATACseq) and Cleavage Under Target and Release Using Nuclease (CUT&RUN) to identify POU1F1 binding sites and histone marks associated with active enhancers, H3K27Ac and H3K4Me1, or inactive regions, H3K27Me3, in GHF-T1 and TαT1 cells. We integrated DNA accessibility, histone modification patterns, transcription factor binding and RNA expression data to identify regulatory elements and candidate transcriptional regulators. We identified POU1F1 binding sites that were unique to each cell line. For example, POU1F1 binds sites in and around Cga and Tshb only in TαT1 cells and Twist1 and Gli3 only in GHFT1 cells. POU1F1 binding sites are commonly associated with bZIP factor consensus binding sites in GHFT1 cells and Helix-Turn-Helix or basic Helix-Loop-Helix in TαT1 cells, suggesting classes of transcription factors that may recruit POU1F1 to unique sites. We validated enhancer function of novel elements we mapped near Tshb, Gata2, and Pitx1 by transfection in TαT1 cells. Finally, we confirmed that an enhancer element near Tshb can drive expression in thyrotropes of transgenic mice. These data extend the ENCODE analysis to an organ that is critical for growth and metabolism. This information could be valuable for understanding pituitary development and disease pathogenesis.

Sox2 regulatory sequences direct expression of a (beta)-geo transgene to telencephalic neural stem cells and precursors of the mouse embryo, revealing regionalization of gene expression in CNS stem cells

Development ◽  
2000 ◽  
Vol 127 (11) ◽  
pp. 2367-2382 ◽  
Author(s):  
M.V. Zappone ◽  
R. Galli ◽  
R. Catena ◽  
N. Meani ◽  
S. De Biasi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Spinal Cord ◽  
Stem Cells ◽  
Transcription Factors ◽  
Neural Stem Cells ◽  
Ventricular Zone ◽  
Specific Gene ◽  
Regulatory Sequences ◽  
Enhancer Element ◽  
Dorsal Telencephalon

Sox2 is one of the earliest known transcription factors expressed in the developing neural tube. Although it is expressed throughout the early neuroepithelium, we show that its later expression must depend on the activity of more than one regionally restricted enhancer element. Thus, by using transgenic assays and by homologous recombination-mediated deletion, we identify a region upstream of Sox2 (−5.7 to −3.3 kb) which can not only drive expression of a (beta)-geo transgene to the developing dorsal telencephalon, but which is required to do so in the context of the endogenous gene. The critical enhancer can be further delimited to an 800 bp fragment of DNA surrounding a nuclease hypersensitive site within this region, as this is sufficient to confer telencephalic expression to a 3.3 kb fragment including the Sox2 promoter, which is otherwise inactive in the CNS. Expression of the 5.7 kb Sox2(beta)-geo transgene localizes to the neural plate and later to the telencephalic ventricular zone. We show, by in vitro clonogenic assays, that transgene-expressing (and thus G418-resistant) ventricular zone cells include cells displaying functional properties of stem cells, i.e. self-renewal and multipotentiality. We further show that the majority of telencephalic stem cells express the transgene, and this expression is largely maintained over two months in culture (more than 40 cell divisions) in the absence of G418 selective pressure. In contrast, stem cells grown in parallel from the spinal cord never express the transgene, and die in G418. Expression of endogenous telencephalic genes was similarly observed in long-term cultures derived from the dorsal telencephalon, but not in spinal cord-derived cultures. Thus, neural stem cells of the midgestation embryo are endowed with region-specific gene expression (at least with respect to some networks of transcription factors, such as that driving telencephalic expression of the Sox2 transgene), which can be inherited through multiple divisions outside the embryonic environment.

Transcriptional Regulation of Thrombopoiesis.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. sci-38-sci-38
Author(s):  
Alan B. Cantor ◽  
Hui Huang ◽  
Andrew Woo ◽  
James Mann ◽  
Ming Yu ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Cell Signaling ◽  
Signaling Pathways ◽  
Transcriptional Activation ◽  
Protein Complex ◽  
Molecular Mechanisms ◽  
Control Cell ◽  
Cell Fate Decisions ◽  
Platelet Disorder ◽  
Cell Signaling Pathways

Abstract Over the past two decades, a number of key transcription factors have been identified that play essential roles in megakaryocyte development. These include GATA-1, GATA-2, Friend of GATA-1 (FOG-1), Runx-1, Cbf-β, Fli-1, GABPα, TEL, NF-E2 p45, Gfi-1b, and SCL/TAL. Importantly, mutations in genes encoding several of these have been linked to human disorders of thrombopoieisis. Germline GATA-1 mutations that disrupt binding to FOG-1 cause X-linked macrothrombocytopenia and dyserythropoietic anemia. Acquired GATA-1 mutations that lead to exclusive production of a short isoform (GATA-1s) play initiating roles in Down Syndrome Transient Myeloproliferative Disorder (DS-TMD) and subsequent Acute Megakaryoblastic Leukemia (DS-AMKL). Haploinsufficiency of Runx-1 causes Familial Platelet Disorder with Propensity to Develop AML (FPD/ AML). Heterozygous loss of the Fli-1 gene leads to the macrothrombocytopenia seen in Jacobsen’s (Paris-Trousseau) syndrome. Important outstanding questions include: how these transcription factors act together to control megakaryocyte terminal maturation; how they differentially act as activators or repressors depending on gene context; how they intersect with cell signaling pathways; how they may coordinate terminal megakaryocyte maturation with spatial location within the bone marrow; how they may control cell fate decisions of bipotential erythroid/megakaryocytic progenitor cells; and whether additional key transcription factors exist. Application of proteomic approaches involving multi-protein complex purification has provided novel insights into some of these questions. We have isolated GATA-1 containing complexes from megakaryocytic cells and identified the Krüppel-type zinc finger transcription factor ZBP-89 as a novel regulator of megakaryocyte and erythroid development. Knockdown of ZBP-89 expression in zebrafish embryos and mice results in blocked early megakaryopoiesis and definitive erythropoiesis, phenocopying aspects of GATA-1- and FOG-1-deficient animals. We have also found that the focal adhesion component Kindlin-3 co-localizes to the nucleus and interacts with FOG-1, suggesting a possible link between integrin signaling and megakaryocyte transcriptional control. Runx-1 multi-protein complex purifications have led to the identification of Fli-1 as a direct binding partner. This interaction results in synergistic transcriptional activation of megakaryocyte-specific genes. Interestingly, the interaction between Runx-1 and Fli-1 occurs preferentially in cells that are differentiating, even though both proteins are expressed abundantly in undifferentiated megakaryoblastic cells. This binding event correlates with assembly of a large complex containing Runx-1/ Fli-1/GATA-1/FOG-1 based on gel filtration chromatography experiments. These factors may, therefore, act as a megakaryocyte-specific enhancesome. Key future directions are aimed at elucidating the molecular mechanisms that regulate these protein-protein interactions and how cell signaling pathways may modulate them.

MS4A1 (CD20) Gene Expression Is Down-Regulated by Recruiting the Histone Deacetylase Protein Complex to the Promoter in the CD20-Negative B-Lymphoma Cells After Treatment with Rituximab.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1286-1286
Author(s):  
Takumi Sugimoto ◽  
Akihiro Tomita ◽  
Junji Hiraga ◽  
Kazuyuki Shimada ◽  
Hitoshi Kiyoi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Protein Expression ◽  
B Cell ◽  
Protein Complex ◽  
Research Funding ◽  
Gene Promoter ◽  
B Lymphoma ◽  
B Lymphoma Cells ◽  
Cd20 Expression

Abstract Abstract 1286 Poster Board I-308 Background Rituximab, a mouse/human chimeric-monoclonal antibody, is now one of the critical molecular targeting drugs for treatment of CD20-positive B-cell lymphomas. Although the survival benefit of rituximab has been proved for several types of CD20-positive B-cell malignancies, resistance to rituximab has also become a considerable problem. Very recently, we reported that down-modulation of CD20 protein expression in CD20-positive B-cell lymphoma patients after treatment with rituximab-containing combination chemotherapies had been observed in 26.3% of re-biopsied patients under relapsed/progress disease (RD/PD) condition (Hiraga et al., 2009, Blood). Interestingly, CD20 expression and the rituximab sensitivity were partially restored by some epigenetic drugs in vitro, suggesting that aberrant down-regulation of MS4A1 gene expression by epigenetic mechanisms may be related to the loss of CD20 protein expression. Aims Analyses of the molecular mechanisms of MS4A1 gene down-regulation after treatment with rituximab-containing chemotherapies. Results Primary B-lymphoma cells and RRBL1 cells (Hiraga et al., 2009, Blood; Tomita et al., 2007, Int J Hematol.), that showed CD20-negative phenotype after using rituximab, were analyzed in these assays. CD20 mRNA and protein expression was partially stimulated by decitabine (DAC), a DNA methyltransferase (DNMT) inhibitor, and the expression was enhanced by trichostation A (TSA), a histone deacetylase (HDAC) inhibitor. Immunoblot analysis indicated that DNMT1 expression was once down-regulated one day after treatment with DAC, and reversed within 3 days. On the other hand, IRF4/Pu.1, the transcription regulators of MS4A1 gene expression, were consistently present with or without DAC. Bisulfite sequencing was performed to check the CpG methylation status of MS4A1 gene promoter, with the result that no significant methylation was confirmed in CD20-negative transformed cells without DAC. Chromatin immunoprecipitation (ChIP) assay indicated that Sin3-HDAC1 co-repressor complex was recruited to MS4A1 gene promoter without DAC/TSA. In the presence of those drugs, Sin3-HDAC1 recruitment was dissociated from the promoter and the histone acetylation of the promoter was confirmed. Under these conditions with/without DAC/TSA, IRF4 and Pu.1 were constantly recruited to the promoter. Immunoprecipitation using whole cell lysate of RRBL1 cells indicated that endogenous Sin3-HDAC1 forms a protein complex, but IRF4 and/or Pu.1 interaction with the complex was not confirmed under this condition. To explore the critical factors for CD20 transcription regulation, expression-profiling assay using cDNA micro array was performed. mRNA from RRBL1 cell with/without DAC/TSA was harvested, and expression profiles were compared. In the presence of DAC or DAC+TSA, 0.7% and 7.0%, respectively, of genes were significantly activated. We are now analyzing some candidates that are critical for the transcription regulation of MS4A1 gene expression. Conclusions Our data indicate that the transcription repression of MS4A1 gene in the CD20-negative phenotypic change after treatment with rituximab is, in part, introduced by the recruitment of Sin3-HDAC1 co-repressor protein complex, not by CpG methylation of the promoter. However, the direct interaction of the complex with IRF4/Pu.1 transcription factors was not confirmed in our assay, and the existence of other transcription factors that interact with Sin3-HDAC1 complex was suggested. To confirm the key regulators of CD20 expression is quite useful for exploring some strategies to overcome the rituximab resistance through CD20-negative transformation after using rituximab. Disclosures Kiyoi: Novartis Pharma Co. Ltd.: Research Funding; Kyowa Hakko Kirin Co. Ltd.: Consultancy. Naoe:Kyowa Hakko Kirin Co., Ltd.: Research Funding; Chugai Pharmaceutical Co., Ltd.: Research Funding; Wyeth K.K.: Research Funding.

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