scholarly journals A common intermediary factor (p52/54) recognizing "acidic blob"-type domains is required for transcriptional activation by the Jun proteins.

1992 ◽  
Vol 12 (12) ◽  
pp. 5508-5515 ◽  
Author(s):  
T Oehler ◽  
P Angel
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Target Genes ◽  
Serum Response Factor ◽  
Limiting Factor ◽  
Regulation Of Transcription ◽  
Physical Interaction ◽  
Main Component

The ability of the c-Jun protein, the main component of the transcription factor AP1, to interact directly or indirectly with the RNA polymerase II-initiation complex to activate transcription was investigated by in vivo transcription interference ("squelching") experiments. Coexpression of a Jun mutant lacking its DNA binding domain strongly represses the activity of wild-type c-Jun. Repression depends on the presence of the transactivation domains (TADs), suggesting that a limiting factor interacting with the TADs is essential to link Jun and the components of the transcriptional machinery. The activity of this intermediary factor(s) is restricted to TADs characterized by an abundance of negatively charged amino acids, as demonstrated by the abilities of the TADs of JunB, GAL4, and VP16 to repress c-Jun activity. Depending on the presence of the TADs of Jun, we found physical interaction between Jun and a cluster of three proteins with molecular masses of 52, 53, and 54 kDa (p52/54). Association between Jun and p52/54 is strongly reduced in the presence of VP16, suggesting that the two proteins compete for binding to p52/54. Transcription factors containing a different type of TAD (e.g., GHF1, estrogen receptor, or serum response factor) fail to inhibit Jun activity, suggesting that these proteins act through a different mechanism. We consider the requirement of Jun to interact with p52/54 utilized by other transcription factors a new mechanism in the regulation of transcription of Jun-dependent target genes.

A common intermediary factor (p52/54) recognizing "acidic blob"-type domains is required for transcriptional activation by the Jun proteins

1992 ◽  
Vol 12 (12) ◽  
pp. 5508-5515
Author(s):  
T Oehler ◽  
P Angel
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Target Genes ◽  
Serum Response Factor ◽  
Limiting Factor ◽  
Regulation Of Transcription ◽  
Physical Interaction ◽  
Main Component

The ability of the c-Jun protein, the main component of the transcription factor AP1, to interact directly or indirectly with the RNA polymerase II-initiation complex to activate transcription was investigated by in vivo transcription interference ("squelching") experiments. Coexpression of a Jun mutant lacking its DNA binding domain strongly represses the activity of wild-type c-Jun. Repression depends on the presence of the transactivation domains (TADs), suggesting that a limiting factor interacting with the TADs is essential to link Jun and the components of the transcriptional machinery. The activity of this intermediary factor(s) is restricted to TADs characterized by an abundance of negatively charged amino acids, as demonstrated by the abilities of the TADs of JunB, GAL4, and VP16 to repress c-Jun activity. Depending on the presence of the TADs of Jun, we found physical interaction between Jun and a cluster of three proteins with molecular masses of 52, 53, and 54 kDa (p52/54). Association between Jun and p52/54 is strongly reduced in the presence of VP16, suggesting that the two proteins compete for binding to p52/54. Transcription factors containing a different type of TAD (e.g., GHF1, estrogen receptor, or serum response factor) fail to inhibit Jun activity, suggesting that these proteins act through a different mechanism. We consider the requirement of Jun to interact with p52/54 utilized by other transcription factors a new mechanism in the regulation of transcription of Jun-dependent target genes.

scholarly journals Stepwise Recruitment of Components of the Preinitiation Complex by Upstream Activators In Vivo

1998 ◽  
Vol 18 (5) ◽  
pp. 2876-2883 ◽  
Author(s):  
Song He ◽  
Steven Jay Weintraub
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Mechanism Of Action ◽  
Transcriptional Activation ◽  
Tata Binding Protein ◽  
Preinitiation Complex ◽  
Functional Assays ◽  
Rna Polymerase Ii Holoenzyme

ABSTRACT Recently, it was found that if either the TATA binding protein or RNA polymerase II holoenzyme is artificially tethered to a promoter, transcription is activated. This finding provided presumptive evidence that upstream activating proteins function by recruiting components of the preinitiation complex (PIC) to the promoter. To date, however, there have been no studies demonstrating that upstream factors actually recruit components of the PIC to the promoter in vivo. Therefore, we have studied the mechanism of action of two disparate transactivating domains. We present a series of in vivo functional assays that demonstrate that each of these proteins targets different components of the PIC for recruitment. We show that, by targeting different components of the PIC for recruitment, these activating domains can cooperate with each other to activate transcription synergistically and that, even within one protein, two different activating subdomains can activate transcription synergistically by cooperating to recruit different components of the PIC. Finally, considering our work together with previous studies, we propose that certain transcription factors both recruit components of the PIC and facilitate steps in transcriptional activation that occur subsequent to recruitment.

scholarly journals Ets-dependent Regulation of Target Gene Expression during Megakaryopoiesis

2004 ◽  
Vol 279 (50) ◽  
pp. 52183-52190 ◽  
Author(s):  
Pascale Jackers ◽  
Gabor Szalai ◽  
Omar Moussa ◽  
Dennis K. Watson
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase Ii ◽  
Chromatin Immunoprecipitation ◽  
Target Genes ◽  
Hematopoietic Stem ◽  
Exogenous Expression ◽  
Direct Target ◽  
Transcriptional Complex

Megakaryopoiesis is the process by which hematopoietic stem cells in the bone marrow differentiate into mature megakaryocytes. The expression of megakaryocytic genes during megakaryopoiesis is controlled by specific transcription factors. Fli-1 and GATA-1 transcription factors are required for development of megakaryocytes and promoter analysis has definedin vitrofunctional binding sites for these factors in several megakaryocytic genes, includingGPIIb,GPIX, andC-MPL. Herein, we utilize chromatin immunoprecipitation to examine the presence of Ets-1, Fli-1, and GATA-1 on these promotersin vivo. Fli-1 and Ets-1 occupy the promoters ofGPIIb,GPIX, andC-MPLgenes in both Meg-01 and CMK11-5 cells. WhereasGPIIbis expressed in both Meg-01 and CMK11-5 cells,GPIXandC-MPLare only expressed in the more differentiated CMK11–5 cells. Thus,in vivooccupancy by an Ets factor is not sufficient to promote transcription of some megakaryocytic genes. GATA-1 and Fli-1 are both expressed in CMK11-5 cells and co-occupy theGPIXandC-MPLpromoters. Transcription of all three megakaryocytic genes is correlated with the presence of acetylated histone H3 and phosphorylated RNA polymerase II on their promoters. We also show that exogenous expression of GATA-1 in Meg-01 cells leads to the expression of endogenous c-mpl and gpIX mRNA. WhereasGPIIb,GPIX, andC-MPLare direct target genes for Fli-1, both Fli-1 and GATA-1 are required for formation of an active transcriptional complex on theC-MPLandGPIXpromotersin vivo. In contrast,GPIIbexpression appears to be independent of GATA-1 in Meg-01 cells.

New Approaches for Mapping Epigenome Dynamics

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. SCI-21-SCI-21
Author(s):  
Steven Henikoff
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase Ii ◽  
Conflicts Of Interest ◽  
Regulation Of Transcription ◽  
Base Pairs ◽  
Simple Method ◽  
Genome Wide ◽  
Resolution Mapping

Abstract The protein complexes that package our genomes must be mobilized for active processes to occur, including replication and transcription, but until recently we have only had a static, low resolution view of the "epigenome". Genomes are packaged into nucleosomes, octamers of four core histones wrapped by 147 base pairs of DNA. Nucleosomes present obstacles to transcription, which over genes is the RNA Polymerase II (RNAPII) complex, and one current challenge is to understand what happens to a nucleosome when RNAPII transcribes through the DNA that it occupies. We study this process by developing methods for following nucleosomes as they are evicted and replaced. Among the factors that we have implicated in the process is torsional stress, which we can now measure genome-wide. RNAPII movement can unwrap nucleosomes and thus destabilize them, causing them to be occasionally evicted and replaced. Interestingly, we find that destabilization of nucleosomes during transcription is enhanced by anthracycline compounds, widely used chemotherapeutic drugs that intercalate between DNA base pairs, thus suggesting a new mechanism for cell killing during chemotherapy. We are also interested in what happens to RNAPII during its encounter with a nucleosomes. In vitro, RNAPII cannot transcribe completely through a nucleosome, but rather stalls as it tries to unwrap the DNA from around the core. We have been studying this process in vivo, and have developed a simple method for precisely mapping RNAPII genome-wide. We have used this method to show exactly where RNAPII stalls as it unwraps a nucleosome in vivo, surprisingly in a different place in vivo from where it stalls in vitro. We also have discovered that a variant histone, H2A.Z, which is found in essentially all eukaryotes, helps to reduce the nucleosome barrier to transcription, and in this way may modulate transcription. Other protein components of the epigenome involved in dynamic processes are nucleosome remodelers, which use the energy of ATP to slide or even evict nucleosomes from DNA. Some remodelers help RNAPII get started and others help it overcome the nucleosome barrier to transcription, and by mapping them at base-pair resolution, we can gain insight into how they act. We have also applied our high-resolution mapping tools to transcription factors, which bind DNA at specific sites to regulate transcription and other processes. Our ability to achieve high spatial and temporal resolution mapping of the binding and action of nucleosomes, transcription factors, remodelers and RNAPII provides us with a detailed picture of epigenome dynamics. By using these tools we are beginning to understand how DNA sequence and conformation are recognized for regulation of transcription and other epigenomic processes. Disclosures No relevant conflicts of interest to declare.

scholarly journals Regulation of Thermotolerance by Stress-Induced Transcription Factors in Saccharomyces cerevisiae

2008 ◽  
Vol 7 (5) ◽  
pp. 783-790 ◽  
Author(s):  
Noritaka Yamamoto ◽  
Yuka Maeda ◽  
Aya Ikeda ◽  
Hiroshi Sakurai
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Factors ◽  
High Temperature ◽  
Heat Shock ◽  
Cell Growth ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Target Genes ◽  
Recovery Period ◽  
Before And After

ABSTRACT The heat shock transcription factor Hsf1 and the general stress transcription factors Msn2 and Msn4 (Msn2/4) are major regulators of the heat shock response in Saccharomyces cerevisiae. Here, we show that transcriptional activation of their target genes, including HSP104, an antistress chaperone gene, is obligatory for thermotolerance. Although Hsf1 activity might be necessary before the exposure of cells to high temperature, severe heat shock induced the binding of hyperphosphorylated Hsf1 to its target promoters. However, promoter-bound, phosphorylated Hsf1 was inactive for transcription because RNA polymerase II was inactive at high temperatures. Rather, our results suggest that Hsf1 activates the transcription of most of its target genes during the recovery period following severe heat shock. This delayed upregulation by Hsf1, which would be induced by misfolded proteins that accumulate in severely heat-shocked cells, is required for the resumption of normal cell growth. In contrast, the factors Msn2/4 were not involved in the delayed upregulation of genes and were dispensable for cell growth during the recovery period, suggesting that they play a role before the exposure to high temperature. These results show that Hsf1 and Msn2/4 act differentially before and after exposure to extreme temperatures to ensure cell survival and growth.

scholarly journals HMGN1 Modulates Estrogen-Mediated Transcriptional Activation through Interactions with Specific DNA-Binding Transcription Factors

2007 ◽  
Vol 27 (24) ◽  
pp. 8859-8873 ◽  
Author(s):  
Nan Zhu ◽  
Ulla Hansen
Keyword(s):  
Transcription Factors ◽  
Protein Interactions ◽  
Transcriptional Activation ◽  
Serum Response Factor ◽  
Estrogen Receptor Α ◽  
Order Structure ◽  
Specific Gene ◽  
Estrogen Induction

ABSTRACT HMGN1, an abundant nucleosomal binding protein, can affect both the chromatin higher order structure and the modification of nucleosomal histones, but it alters the expression of only a subset of genes. We investigated specific gene targeting by HMGN1 in the context of estrogen induction of gene expression. Knockdown and overexpression experiments indicated that HMGN1 limits the induction of several estrogen-regulated genes, including TFF1 and FOS, which are induced by estrogen through entirely distinct mechanisms. HMGN1 specifically interacts with estrogen receptor α (ERα), both in vitro and in vivo. At the TFF1 promoter, estrogen increases HMGN1 association through recruitment by the ERα. HMGN1 S20E/S24E, although deficient in binding nucleosomal DNA, still interacts with ERα and, strikingly, still represses estrogen-driven activation of the TFF1 gene. On the FOS promoter, which lacks the ERα binding sites, constitutively bound serum response factor (SRF) mediates estrogen stimulation. HMGN1 also interacts specifically with SRF, but HMGN1 S20E/S24E does not. Consistent with the protein interactions, only wild-type HMGN1 significantly inhibits the estrogen-driven activation of the FOS gene. Mechanistically, the inhibition of estrogen induction of several ERα-associated genes, including TFF1, by HMGN1 correlates with decreased levels of acetylation of Lys9 on histone H3. Together, these findings indicate that HMGN1 regulates the expression of particular genes via specific protein-protein interactions with transcription factors at target gene regulatory regions.

Orientation-Dependent Regulation of RUNX1 and GATA-1 Transcriptional Function by the p300/CBP Coactivators.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1184-1184
Author(s):  
Ivailo S. Mihaylov ◽  
Kamaleldin E. Elagib ◽  
Lorrie L. Delehanty ◽  
Sara L. Gonias ◽  
Jill F. Caronia ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Transcriptional Activation ◽  
Transcriptional Activity ◽  
Target Genes ◽  
Serine Phosphorylation ◽  
Amino Terminus ◽  
Physical Interaction ◽  
Wild Type ◽  
Carboxy Terminus ◽  
Target Sites

Abstract The transcription factors RUNX1 and GATA-1, as well as the coactivators p300/CBP, have been implicated in the regulation of primary megakaryopoiesis through studies of knockout mice. In particular, p300/CBP has previously been shown to serve as a coactivator for both RUNX1 and GATA-1 in the transactivation of hematopoietic target genes. Coactivator orientation within transactivating complexes is generally not considered to influence the degree of transcriptional activity, reflecting an inherent flexibility in the spatial requirements for coactivator function. Experiments to further explore this issue showed coexpression of p300 to enhance cooperative transcriptional activation by wild type RUNX1 and GATA-1. Enforced recruitment of p300/CBP to GATA-1 through fusion of GATA-1 with the p300/CBP docking module of adenoviral E1A, E1A(1–89), enhanced GATA-1 activity alone, regardless of whether the fusion was to the amino or carboxy terminal of GATA-1. However, enforced recruitment of p300/CBP to the amino terminus of GATA-1 completely eliminated cooperation with RUNX1, and enforced recruitment of p300/CBP to the carboxy terminus of GATA-1 diminished cooperation with RUNX1. Both GATA-1 fusions retained physical interaction with RUNX1. Similarly, fusion of E1A(1–89) directly to the amino terminus of RUNX1 completely eliminated its transcriptional activity, while fusion to the carboxy terminus diminished RUNX1 transcriptional activity. For both the GATA-1 and the RUNX1 fusions, these repressive effects were attributable to the ectopic recruitment of p300/CBP because a mutation within E1A(1–89) known to specifically diminish p300/CBP recruitment, R2G, rescued the transcriptional activities. Addressing the mechanism of repression by ectopic p300/CBP, we found that E1A(1–89)-GATA-1 caused diminished serine phosphorylation within RUNX1, an effect opposite to that of wild type GATA-1 which enhanced RUNX1 phosphorylation. Similarly, E1A(1–89)-RUNX1 showed complete loss of phosphorylation on cdk target sites, as compared with wild type RUNX1. RUNX1-E1A(1–89) showed diminished phosphorylation on cdk target sites, as compared with wild type RUNX1. From these results, we conclude that p300/CBP may function as a coactivator for the RUNX1-GATA-1 complex when recruited to endogenous, wild type domains. By contrast, ectopic recruitment of p300/CBP to RUNX1, particularly to the amino terminus, targets RUNX1 for repression through inhibition or reversal of phosphorylation. Our results thus offer a novel paradigm in which the function of p300/CBP, coactivator versus repressor, may be determined by its mode of recruitment to certain transcriptional complexes. Notably, some transcription factors, such as GATA-1, have relaxed requirements for the topology of coactivator recruitment, whereas other transcription factors, such as RUNX1, have stringent requirements in this regard.

Activation Functions 1 and 2 of Nuclear Receptors: Molecular Strategies for Transcriptional Activation

2003 ◽  
Vol 17 (10) ◽  
pp. 1901-1909 ◽  
Author(s):  
Anette Wärnmark ◽  
Eckardt Treuter ◽  
Anthony P. H. Wright ◽  
Jan-Åke Gustafsson
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase Ii ◽  
Nuclear Receptors ◽  
Transcriptional Activation ◽  
Molecular Mechanisms ◽  
Target Genes ◽  
Current Knowledge ◽  
Preinitiation Complex ◽  
Activation Domains ◽  
Terminal Domain

Abstract Nuclear receptors (NRs) comprise a family of ligand inducible transcription factors. To achieve transcriptional activation of target genes, DNA-bound NRs directly recruit general transcription factors (GTFs) to the preinitiation complex or bind intermediary factors, so-called coactivators. These coactivators often constitute subunits of larger multiprotein complexes that act at several functional levels, such as chromatin remodeling, enzymatic modification of histone tails, or modulation of the preinitiation complex via interactions with RNA polymerase II and GTFs. The binding of NR to coactivators is often mediated through one of its activation domains. Many NRs have at least two activation domains, the ligand-independent activation function (AF)-1, which resides in the N-terminal domain, and the ligand-dependent AF-2, which is localized in the C-terminal domain. In this review, we summarize and discuss current knowledge regarding the molecular mechanisms of AF-1- and AF-2-mediated gene activation, focusing on AF-1 and AF-2 conformation and coactivator binding.

scholarly journals Activator Gcn4p and Cyc8p/Tup1p Are Interdependent for Promoter Occupancy at ARG1 In Vivo

2005 ◽  
Vol 25 (24) ◽  
pp. 11171-11183 ◽  
Author(s):  
Soon-ja Kim ◽  
Mark J. Swanson ◽  
Hongfang Qiu ◽  
Chhabi K. Govind ◽  
Alan G. Hinnebusch
Keyword(s):  
Amino Acid ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Target Genes ◽  
Direct Role ◽  
Valine Biosynthesis ◽  
Promoter Occupancy ◽  
Yeast Deletion Library ◽  
Deletion Library

ABSTRACT The Cyc8p/Tup1p complex mediates repression of diverse genes in Saccharomyces cerevisiae and is recruited by DNA binding proteins specific for the different sets of repressed genes. By screening the yeast deletion library, we identified Cyc8p as a coactivator for Gcn4p, a transcriptional activator of amino acid biosynthetic genes. Deletion of CYC8 confers sensitivity to an inhibitor of isoleucine/valine biosynthesis and impairs activation of Gcn4p-dependent reporters and authentic amino acid biosynthetic target genes. Deletion of TUP1 produces similar but less severe activation defects in vivo. Although expression of Gcn4p is unaffected by deletion of CYC8, chromatin immunoprecipitation assays reveal a strong defect in binding of Gcn4p at the target genes ARG1 and ARG4 in cyc8Δ cells and to a lesser extent in tup1Δ cells. The defects in Gcn4p binding and transcriptional activation in cyc8Δ cells cannot be overcome by Gcn4p overexpression but are partially suppressed in tup1Δ cells. The impairment of Gcn4p binding in cyc8Δ and tup1Δ cells is severe enough to reduce recruitment of SAGA, Srb mediator, TATA binding protein, and RNA polymerase II to the ARG1 and ARG4 promoters, accounting for impaired transcriptional activation of these genes in both mutants. Cyc8p and Tup1p are recruited to the ARG1 and ARG4 promoters, consistent with a direct role for this complex in stimulating Gcn4p occupancy of the upstream activation sequence (UAS). Interestingly, Gcn4p also stimulates binding of Cyc8p/Tup1p at the 3′ ends of these genes, raising the possibility that Cyc8p/Tup1p influences transcription elongation. Our findings reveal a novel coactivator function for Cyc8p/Tup1p at the level of activator binding and suggest that Gcn4p may enhance its own binding to the UAS by recruiting Cyc8p/Tup1p.

scholarly journals Interaction of Serum Response Factor (SRF) with the Elk-1 B Box Inhibits RhoA-Actin Signaling to SRF and Potentiates Transcriptional Activation by Elk-1

2002 ◽  
Vol 22 (20) ◽  
pp. 7083-7092 ◽  
Author(s):  
Kasumi Murai ◽  
Richard Treisman
Keyword(s):  
Binding Site ◽  
Transcriptional Activation ◽  
Rho Gtpases ◽  
Target Genes ◽  
Serum Response Factor ◽  
Relative Sensitivity ◽  
Actin Dynamics ◽  
Physical Interaction ◽  
Response Factor ◽  
Serum Response

ABSTRACT Serum response factor (SRF) is a transcription factor which regulates many immediate-early genes. Rho GTPases regulate SRF activity through changes in actin dynamics, but some SRF target genes, such as c-fos, are insensitive to this pathway. At the c-fos promoter, SRF recruits members of the ternary complex factor (TCF) family of Ets domain proteins through interactions with the TCF B-box region. Analysis of c-fos promoter mutations demonstrates that the TCF and ATF/AP1 sites adjoining the SRF binding site inhibit activation of the promoter by RhoA-actin signaling. The presence of the TCF binding site is sufficient for inhibition, and experiments with an altered-specificity Elk-1 derivative demonstrate that inhibition can be mediated by the Elk-1 TCF. Using Elk-1 fusion proteins that can bind DNA autonomously, we show that inhibition of RhoA-actin signaling requires physical interaction between the Elk-1 B box and SRF. These results account for the insensitivity of c-fos to RhoA-actin signaling. Interaction of the B box with SRF also potentiates transcriptional activation by the Elk-1 C-terminal activation domain. Combinatorial interactions between SRF and TCF proteins are thus likely to play an important role in determining the relative sensitivity of SRF target genes to Ras- and Rho-controlled signal transduction pathways.

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