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)

The X protein of hepatitis B virus (HBV) coactivates activators bearing potent (mostly acidic) activation domains. Here, we investigated the molecular mechanisms of this coactivation. We show that pX interacts with general transcription factors TFIIB and TFIIH, as well as with the potent activation domain of VP16. TFIIB interacts with both pX and VP16 simultaneously. In addition, the RNA polymerase II enzyme itself binds to pX. By reducing the activity of cellular coactivators, through squelching, we intensify the dependence of the activator on pX-mediated coactivation. Squelching is essentially diminished in the presence of pX, both in vivo and in vitro. The target of pX in this activity is the template-bound activator, and not the squelcher. Furthermore, by following transcription in a TAF-deprived reaction, we demonstrate absolute dependence of the activator on the activity of pX. We propose that pX coactivates transcription by substituting cellular coactivators in activator-preinitiation complex interactions.
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PMID:pX, the HBV-encoded coactivator, interacts with components of the transcription machinery and stimulates transcription in a TAF-independent manner. 867 Aug 43

The control of gene expression during development, differentiation and maintenance of cellular function is governed by a complex array of transcription factors. We have undertaken a molecular dissection of the regulatory factors that direct transcription of protein coding genes by RNA polymerase II. Our early studies identified sequence-specific transcriptional activators that bind to enhancer and promoter sequences to modulate the transcriptional initiation event. However, the mechanism by which activators enhance transcription and mediate promoter selectivity remained unknown. Combining biochemical purification and in vitro assays, we have recently identified an essential class of transcription factors called TAFs that are tightly associated with the basal factor TBP (TATA-binding protein). We have found that TAFs are responsible for at least two regulatory functions. Some TAFs serve as coactivators capable of binding activators and mediating enhancing function. Other TAFs have been shown to confer template selectivity by binding directly to core DNA elements of the promoter. Thus different subunits of TBP/TAF complexes perform a variety of functions critical for transcriptional regulation in animal cells.
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PMID:The biochemistry of transcription in eukaryotes: a paradigm for multisubunit regulatory complexes. 873 71

TFIID is the DNA binding component of the RNA polymerase II transcriptional machinery and is composed of the TATA binding protein (TBP) and TBP-associated factors (TAFIIs). Here we report the characterization of a new human TAF, hTAFII100, which is the human homologue of Drosophila TAFII80 and yeast TAFII90. hTAFII100 interacts strongly with hTAFII250, hTAFII55 and hTAFII28, less with hTAFII20 and hTAFII18, weakly with TBP and not at all with delta NTAFII135 and hTAFII30. Deletion analysis revealed that the C-terminal half of hTAFII100, which contains six WD-40 repeats, is not required for incorporation into the TFIID complex. Our results suggest that hTAFII100 can be divided into two domains, the N-terminal region responsible for interactions within the TFIID complex and the C-terminal WD repeat-containing half responsible for interactions between hTAFII100 and other factors. An anti-hTAFII100 antibody, raised against a C-terminal epitope, selectively inhibited basal TFIID-dependent in vitro transcription and the specific interaction between hTAFII100 and the 30 kDa subunit of TFIIF (RAP30). We demonstrate that the hTAFII100-TFIIF interaction supports pre-initiation complex formation in the presence of TFIID. Thus, this is the first demonstration that a TAFII functionally interacts with a basal transcription factor in vitro.
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PMID:Distinct domains of hTAFII100 are required for functional interaction with transcription factor TFIIF beta (RAP30) and incorporation into the TFIID complex. 875 37

The RNA polymerase II general transcription factor TFIID is a multisubunit complex comprising TATA-box binding protein and associated factors (TAFIIs). In vitro experiments have suggested that TAFIIs are essential coactivators required for RNA polymerase II-directed transcription activation. Here, for the first time, we analyze systematically the in vivo function of a specific TAFII, yeast TAFII90 (yTAFII90). We show that functional inactivation of yTAFII90 by temperature-sensitive mutations or depletion leads to arrest at the G2/M phase of the cell cycle. Unexpectedly, in the absence of functional yTAFII90, a variety of endogenous yeast genes were all transcribed normally, including those driven by well-characterized activators. Taken together, our results indicate that yTAFII90 is not required for transcription activation in general, and reveal linkages between TAF function and cell-cycle progression.
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PMID:Yeast TAF(II)90 is required for cell-cycle progression through G2/M but not for general transcription activation. 882 95

The core promoters for mammalian protein-coding genes often contain a TATA box, an initiator (Inr) element, or both of these control elements. The TFIID complex is essential both for TATA activity and for the activity of a common class of Inr elements characterized by an approximate consensus sequence PyPyA+1NT/APyPy. Although the complete set of proteins required for basal TATA-mediated transcription has been established, the requirements for TFIID-dependent Inr activity remain undefined. In this study we set out to reconstitute Inr activity with purified and recombinant general transcription factors. For this analysis, Inr activity was measured as the ability of an Inr to enhance the strength of a core promoter containing an upstream TATA box. Inr activity was not detected in reactions containing TFIIB, RAP30, RAP74, RNA polymerase II, and either TBP or TFIID, even though these factors were sufficient for TATA-mediated transcription from supercoiled templates. By use of a complementation assay, a factor that imparts Inr activity was identified. This factor, named CIF, stimulated Inr activity in reactions containing the TFIID complex, but activity was not detected with TBP. Further characterization of CIF suggested that it contains multiple components. Functional and immunological experiments demonstrated that one of the CIF components is the mammalian homolog of Drosophila TAF(II)150, which is not tightly associated with mammalian TFIID. These results reveal significant differences in the factor requirements for basal TATA and Inr activity. Further elucidation of these differences is likely to explain the need for the core promoter heterogeneity found within protein-coding genes.
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PMID:CIF, an essential cofactor for TFIID-dependent initiator function. 884 23

TFIID is the main sequence-specific DNA-binding component of the RNA polymerase II (Pol II) transcriptional machinery. It is a multiprotein complex composed of the TATA-binding protein (TBP) and TBP-associated factors (TAF(II)s). Here we report the cloning and characterization of a novel human TBP-associated factor, hTAF(II)68. It contains a consensus RNA-binding domain (RNP-CS) and binds not only RNA, but also single stranded (ss) DNA. hTAF(II)68 shares extensive sequence similarity with TLS/FUS and EWS, two human nuclear RNA-binding pro-oncoproteins which are products of genes commonly translocated in human sarcomas. Like hTAF(II)68, TLS/FUS is also associated with a sub-population of TFIID complexes chromatographically separable from those containing hTAF(II)68. Therefore, these RNA and/or ssDNA-binding proteins may play specific roles during transcription initiation at distinct promoters. Moreover, we demonstrate that hTAF(II)68 co-purifies also with the human RNA polymerase II and can enter the preinitiation complex together with Pol II.
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PMID:hTAF(II)68, a novel RNA/ssDNA-binding protein with homology to the pro-oncoproteins TLS/FUS and EWS is associated with both TFIID and RNA polymerase II. 889 Jan 75

In this report we describe the cloning and initial characterization of TAF40, a gene that encodes a yeast TATA-binding protein-associated factor (yTAF) of Mr = approximately 40,000. This gene has many similarities to other yTAFs described thus far in that it is present at a single copy per haploid genome, it is essential for viability, and the deduced protein sequence of yTAF40 exhibits similarity to previously described human and Drosophila TAFIIs. Immunological studies confirm that yTAF40 protein is a subunit of a large multiprotein TATA-binding protein-TAF complex that contains a subset of the total number of the yTAFs present in yeast cell extracts. Transcription reactions performed using yeast whole cell extracts reveal that of the three nuclear RNA polymerases only RNA polymerase II function is abrogated when yTAF40 and associated proteins are immunodepleted from solution, indicating that the functionality of the multiprotein complex containing yTAF40 is RNA polymerase II-specific. By these criteria yTAF40 appears to encode a bona fide RNA polymerase II-specific TAF, and thus the protein that it encodes has been termed yTAFII40.
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PMID:Cloning and characterization of an essential Saccharomyces cerevisiae gene, TAF40, which encodes yTAFII40, an RNA polymerase II-specific TATA-binding protein-associated factor. 908 82

SV40 large T antigen is a multifunctional regulatory protein that plays a key role in the viral life cycle and can stimulate cell proliferation. To accomplish this, large T antigen has to control the expression of cellular genes involved in cell cycle progression and cell growth. rRNA synthesis by RNA polymerase I (Pol I) is tightly associated with cell growth and proliferation, and previous studies indicated that large T antigen up-regulates RNA Pol I transcription in SV40-infected cells. How this process occurs is currently unclear. To investigate the mechanisms of large T antigen stimulation of RNA Pol I transcription, we have established an in vitro transcription system that is responsive to large T antigen. Here, we show that recombinant large T antigen stimulates Pol I transcription reconstituted with purified RNA Pol I, UBF, and the TBP/TAF complex SL1. Immunoprecipitation experiments revealed that large T antigen directly binds to SL1, in vitro, as well as in SV40-infected cells. In addition, our data indicate that this interaction occurs by direct association with three SL1 subunits, namely TBP, TAF(I)48, and TAF(I)110. Transcription studies with large T antigen deletion mutants show that the 538-amino-acid amino-terminal domain is necessary for full stimulation of Pol I transcription. Importantly, mutants that no longer bind to SL1 are also unable to stimulate Pol I transcription. This indicates that recruitment of large T antigen to the rRNA promoter by SL1 constitutes a crucial step in the activation process. Taken together with recent studies on large T antigen activation of RNA Pol II transcription, these results suggest that viral modulation of genes involved in cell proliferation involves direct targeting of promoter-specific TBP/TAF complexes (i.e., SL1 or TFIID) by large T antigen.
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PMID:SV40 large T antigen binds to the TBP-TAF(I) complex SL1 and coactivates ribosomal RNA transcription. 920 86

In eukaryotes, TFIIIB is required for proper initiation by RNA polymerase III. In the yeast Saccharomyces cerevisiae a single form of TFIIIB (gammaTFIIIB) is sufficient for transcription of all pol III genes, whereas in extracts derived from human cells two different hTFIIIB complexes exist which we have previously designated as hTFIIIB-alpha and hTFIIIB-beta. Human TFIIIB-alpha is a TBP-free entity and must be complemented by TBP for transcription of pol III genes driven by gene external promoters, whereas hTFIIIB-beta is a TBP-TAF complex which governs transcription from internal pol III promoters. We show that hTFIIIB-beta cannot be replaced by yeast TFIIIB for transcription of tRNA genes, but that the B" component of gammaTFIIIB can substitute for hTFIIIB-alpha activity in transcription of the human U6 gene. Moreover, hTFIIIB-alpha can be chromatographically divided into activities which are functionally related to gammaTFIIIE and recombinant yB"90, suggesting that hTFIIIB-alpha is a human homolog of yeast TFIIIB". In addition, we show that yeast TBP can only be exchanged against human TBP for in vitro transcription of the human and yeast U6 gene but virtually not for that of the yeast tRNA4Sup gene. This deficiency can be counteracted by a mutant of human TBP (R231K) which is able to replace yeast TBP for transcription of yeast tRNA genes in vitro.
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PMID:Functional interchangeability of TFIIIB components from yeast and human cells in vitro. 930 15

Initiation of transcription by RNA polymerase II from a promoter region on DNA requires the assembly of several initiation factors to form a preinitiation complex. Assembly of this complex is initiated by the binding of the transcription factor TFIID, composed of the TATA-box binding protein (TBP) and TBP-associated factors (TAF[II]s), to the promoter. We have now characterized an immunopurified TFIID complex which we unexpectedly find contains the cleavage-polyadenylation specificity factor (CPSF), one of the factors required for formation of the 3' end of messenger RNA. CPSF is brought to the preinitiation complex by TFIID, but after transcription starts, CPSF dissociates from TFIID and becomes associated with the elongating polymerase. We also show that overexpression of recombinant TBP in HeLa cells decreases polyadenylation without affecting the correct initiation of transcription of the reporter gene. This indicates that, owing to incomplete assembly of TFIID on recombinant TBP, CPSF is not brought to the promoter and therefore polyadenylation becomes less efficient. Our observations have thus revealed a link between transcription initiation and elongation by RNA polymerase II and processing of the 3' end of mRNA.
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PMID:Transcription factor TFIID recruits factor CPSF for formation of 3' end of mRNA. 931 84


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