Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Activation of gene transcription in metazoans is a multistep process that is triggered by factors that recognize transcriptional enhancer sites in DNA. These factors work with co-activators to direct transcriptional initiation by the RNA polymerase II apparatus. One class of co-activator, the TAF(II) subunits of transcription factor TFIID, can serve as targets of activators and as proteins that recognize core promoter sequences necessary for transcription initiation. Transcriptional activation by enhancer-binding factors such as Sp1 requires TFIID, but the identity of other necessary cofactors has remained unknown. Here we describe a new human factor, CRSP, that is required together with the TAF(II)s for transcriptional activation by Sp1. Purification of CRSP identifies a complex of approximate relative molecular mass 700,000 (M(r) approximately 700K) that contains nine subunits with M(r) values ranging from 33K to 200K. Cloning of genes encoding CRSP subunits reveals that CRSP33 is a homologue of the yeast mediator subunit Med7, whereas CRSP150 contains a domain conserved in yeast mediator subunit Rgr1. CRSP p200 is identical to the nuclear hormone-receptor co-activator subunit TRIP2/PBP. CRSPs 34, 77 and 130 are new proteins, but the amino terminus of CRSP70 is homologous to elongation factor TFIIS. Immunodepletion studies confirm that these subunits have an essential cofactor function. The presence of common subunits in distinct cofactor complexes suggests a combinatorial mechanism of co-activator assembly during transcriptional activation.
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PMID:The transcriptional cofactor complex CRSP is required for activity of the enhancer-binding protein Sp1. 998 12

The Mediator of transcriptional regulation is the central coactivator that enables a response of RNA polymerase II (Pol II) to activators and repressors. We present the 3.0-A crystal structure of a highly conserved part of the Mediator, the MED7.MED21 (Med7.Srb7) heterodimer. The structure is very extended, spanning one-third of the Mediator length and almost the diameter of Pol II. It shows a four-helix bundle domain and a coiled-coil protrusion connected by a flexible hinge. Four putative protein binding sites on the surface allow for assembly of the Mediator middle module and for binding of the conserved subunit MED6, which is shown to bridge to the Mediator head module. A flexible MED6 bridge and the MED7.MED21 hinge could account for changes in overall Mediator structure upon binding to Pol II or activators. Our results support the idea that transcription regulation involves conformational changes within the general machinery.
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PMID:A conserved mediator hinge revealed in the structure of the MED7.MED21 (Med7.Srb7) heterodimer. 1571 Jun 19

The human thyroid hormone receptor-associated proteins (TRAP)/Mediator and related complexes mediate transcription through regulatory factors. To further understand the structural and functional diversity of these complexes we established three HeLa cell lines each expressing one of three epitope-tagged human TRAP/Mediator subunits, MED6, MED7, and CDK8 and isolated the complexes in which these subunits were contained by affinity and HPLC-gel filtration chromatography. The largest complexes from each cell line had a molecular mass of 1.5 MDa and possessed almost identical subunit compositions; we designated these complexes TRAP/Mediator-like complex 1 (TMLC1). Two potential subcomplexes were additionally observed: a 1-MDa complex from the CDK8-cell line (TMLC2) and a 600-kDa complex from the MED6-cell line (TMLC3). All three complexes regulated transcription in vitro; TMLC1 and TMLC3 augmented transcriptional activation, whereas TMLC2 repressed it. TMLC1 and TMLC2 phosphorylated RNA polymerase II (Pol II), but TMLC3 did not. Furthermore, TMLC1 predominantly interacted with the general transcription factors TFIIE, TFIIF, and TFIIH, which function during transcription initiation and the transition to elongation. In a final experiment, knockdown of CDK8 using RNA interference prevented transcriptional activation by Gal4-VP16 in a luciferase-assay. This, together with the effect of TMLC1 on transcription in vitro, suggests that CDK8 play positive roles in transcriptional activation.
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PMID:A kinase subunit of the human mediator complex, CDK8, positively regulates transcriptional activation. 1721 59

Mediator is a conserved multisubunit complex that acts as a functional interface between regulatory transcription factors and the general RNA polymerase II initiation apparatus. MED1 is a pivotal component of the complex that binds to nuclear receptors and a broad array of other gene-specific activators. Paradoxically, MED1 is found in only a fraction of the total cellular Mediator complexes, and the mechanisms regulating its binding to the core complex remain unclear. Here, we report that phosphorylation of MED1 by mitogen-activated protein kinase-extracellular signal-regulated kinase (MAPK-ERK) promotes its association with Mediator. We show that MED1 directly binds to the MED7 subunit and that ERK phosphorylation of MED1 enhances this interaction. Interestingly, we found that both thyroid and steroid hormones stimulate MED1 phosphorylation in vivo and that MED1 phosphorylation is required for its nuclear hormone receptor coactivator activity. Finally, we show that MED1 phosphorylation by ERK enhances thyroid hormone receptor-dependent transcription in vitro. Our findings suggest that ERK phosphorylation of MED1 is a regulatory mechanism that promotes MED1 association with Mediator and, as such, may facilitate a novel feed-forward action of nuclear hormones.
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PMID:MED1 phosphorylation promotes its association with mediator: implications for nuclear receptor signaling. 1839 Oct 15

HIV-1 exploits multiple host proteins during infection. siRNA-based screenings have identified new proteins implicated in different pathways of the viral cycle that participate in a broad range of cellular functions. The human Mediator complex (MED) is composed of 28 elements and represents a fundamental component of the transcription machinery, interacting with the RNA polymerase II enzyme and regulating its ability to express genes. Here, we provide an evaluation of the MED activity on HIV replication. Knockdown of 9 out of 28 human MED proteins significantly impaired viral replication without affecting cell viability, including MED6, MED7, MED11, MED14, MED21, MED26, MED27, MED28, and MED30. Impairment of viral replication by MED subunits was at a post-integration step. Inhibition of early HIV transcripts was observed by siRNA-mediated knockdown of MED6, MED7, MED11, MED14, and MED28, specifically affecting the transcription of the nascent viral mRNA transactivation-responsive element. In addition, MED14 and MED30 were shown to have special relevance during the formation of unspliced viral transcripts (p < 0.0005). Knockdown of the selected MED factors compromised HIV transcription induced by Tat, with the strongest inhibitory effect shown by siMED6 and siMED14 cells. Co-immunoprecipitation experiments suggested physical interaction between MED14 and HIV-1 Tat protein. A better understanding of the mechanisms and factors controlling HIV-1 transcription is key to addressing the development of new strategies required to inhibit HIV replication or reactivate HIV-1 from the latent reservoirs.
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PMID:Characterization of the influence of mediator complex in HIV-1 transcription. 2510 Jul 19

Mediator plays an integral role in activation of RNA polymerase II (Pol II) transcription. A key step in activation is binding of Mediator to Pol II to form the Mediator-Pol II holoenzyme. Here, we exploit a combination of biochemistry and macromolecular EM to investigate holoenzyme assembly. We identify a subset of human Mediator head module subunits that bind Pol II independent of other subunits and thus probably contribute to a major Pol II binding site. In addition, we show that binding of human Mediator to Pol II depends on the integrity of a conserved "hinge" in the middle module MED21-MED7 heterodimer. Point mutations in the hinge region leave core Mediator intact but lead to increased disorder of the middle module and markedly reduced affinity for Pol II. These findings highlight the importance of Mediator conformation for holoenzyme assembly.
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PMID:Role for the MED21-MED7 Hinge in Assembly of the Mediator-RNA Polymerase II Holoenzyme. 2782 93