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)

RNA polymerase II-dependent transcription requires the assembly of a multi-protein, preinitiation complex on core promoter elements. Transcription factor IID (TFIID) comprising the TATA box-binding protein (TBP) and TBP-associated factors (TAFs) is responsible for promoter recognition in this complex. Subsequent association of TFIIA and TFIIB provides enhanced complex stability. TFIIA is required for transcriptional stimulation by certain viral and cellular activators, and favors formation of the preinitiation complex in the presence of repressor NC2. The X-ray structures of human and yeast TBP/TFIIA/DNA complexes at 2.1A and 1.9A resolution, respectively, are presented here and seen to resemble each other closely. The interactions made by human TFIIA with TBP and DNA within and upstream of the TATA box, including those involving water molecules, are described and compared to the yeast structure. Of particular interest is a previously unobserved region of TFIIA that extends the binding interface with TBP in the yeast, but not in the human complex, and that further elucidates biochemical and genetic results.
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PMID:Novel interactions between the components of human and yeast TFIIA/TBP/DNA complexes. 1297 51

TATA binding protein (TBP) is a key regulator of RNA polymerase transcription. It binds to core promoters, often in large multiprotein complexes, and nucleates RNA polymerase II (Pol II) transcription initiation. In addition to the previously described TBP-like factor present in metazoans (TLF/TRF2/TRP/TLP), we describe a third, vertebrate-specific member of the TBP protein family from zebrafish, called TBP2. Evolutionary conserved TBP2 homologs were also found in human, mouse, frog, and pufferfish. The N-terminal domains of TBP2s are divergent amongst themselves and different from those of TBPs; however, the core domain of TBP2s and TBPs are almost identical. TBP2 binds the TATA box, interacts with TFIIA and TFIIB (similarly to TBP), and can mediate Pol II transcription initiation. However, TBP2 shows contrasting expression patterns in the gonads and during embryonic development in comparison to TBP, suggesting differential function. Knockdown of zebrafish TBP2 results in specific reduction of the protein level, leading to a phenotype, which indicates the requirement of TBP2 for embryonic patterning. The presence of three different TBP family members in vertebrates suggests the existence of developmental stage- and tissue-specific preinitiation complexes with specific requirements for different TBP family members.
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PMID:TBP2, a vertebrate-specific member of the TBP family, is required in embryonic development of zebrafish. 1506

The TATA-binding protein (TBP) is a critical general transcription factor that associates with the core promoter and acts as a nexus for gene regulation through its interactions with other factors. A large number of proteins recognize the relatively small yet highly conserved C-terminal domain of TBP. One subset of these proteins (general transcription factors) interacts with the TBP.TATA complex and RNA polymerase II to create the preinitiation complex. To study TBP functions in preinitiation complex and other complexes, we generated a set of RNA aptamers with high affinity to yeast TBP. These aptamers act on TBP in different ways: all of them bind TBP competitively with DNA bearing the TATA element, and some can actively disrupt the TBP.TATA interaction in preformed, higher-order complexes containing the additional general transcription factors TFIIB and TFIIA. In crude cell extracts, the aptamers inhibit transcription in ways that reveal the dynamic nature of TBP interactions during initiation and reinitiation.
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PMID:Probing TBP interactions in transcription initiation and reinitiation with RNA aptamers that act in distinct modes. 1510 22

RNA polymerase II (RNAPII) requires a set of general transcription factors - TFIIA, TFIIB, TFIID, TFIIE, TFIIF and TFIIH - to initiate transcription from a gene promoter in vitro. General transcription factors have been isolated from Saccharomyces cerevisiae, rat, human and Drosophila, and their corresponding cDNAs have been cloned. In this report, we describe a reconstituted in vitro transcription system that consists of the following preparations of factors from the yeast Schizosaccharomyces pombe: affinity-purified RNAPII, TFIIH, and recombinant TBP, TFIIB, TFIIE and TFIIF. We show that this system can support basal transcription from the adenovirus major late promoter when purified RNAPII is used and activated transcription when the RNAPII holoenzyme (RNAPII plus the Mediator proteins) is included in the reaction. In contrast, the TATA binding protein-associated factors had no effect on transcriptional activation in our Sc. pombe system. These results indicate that Sc. pombe uses the same set of general transcription factors as other eukaryotes and that the Mediator is involved in activated transcription.
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PMID:Mediator is required for activated transcription in a Schizosaccharomyces pombe in vitro system. 1518 71

The Saccharomyces cerevisiae Nhp6 protein is related to the high-mobility-group B family of architectural DNA-binding proteins that bind DNA nonspecifically but bend DNA sharply. Nhp6 is involved in transcriptional activation by both RNA polymerase II (Pol II) and Pol III. Our previous genetic studies have implicated Nhp6 in facilitating TATA-binding protein (TBP) binding to some Pol II promoters in vivo, and we have used a novel genetic screen to isolate 32 new mutations in TBP that are viable in wild-type cells but lethal in the absence of Nhp6. The TBP mutations that are lethal in the absence of Nhp6 cluster in three regions: on the upper surface of TBP that may have a regulatory role, near residues that contact Spt3, or near residues known to contact either TFIIA or Brf1 (in TFIIIB). The latter set of mutations suggests that Nhp6 becomes essential when a TBP mutant compromises its ability to interact with either TFIIA or Brf1. Importantly, the synthetic lethality for some of the TBP mutations is suppressed by a multicopy plasmid with SNR6 or by an spt3 mutation. It has been previously shown that nhp6ab mutants are defective in expressing SNR6, a Pol III-transcribed gene encoding the U6 splicing RNA. Chromatin immunoprecipitation experiments show that TBP binding to SNR6 is reduced in an nhp6ab mutant. Nhp6 interacts with Spt16/Pob3, the yeast equivalent of the FACT elongation complex, consistent with nhp6ab cells being extremely sensitive to 6-azauracil (6-AU). However, this 6-AU sensitivity can be suppressed by multicopy SNR6 or BRF1. Additionally, strains with SNR6 promoter mutations are sensitive to 6-AU, suggesting that decreased SNR6 RNA levels contribute to 6-AU sensitivity. These results challenge the widely held belief that 6-AU sensitivity results from a defect in transcriptional elongation.
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PMID:TATA-binding protein mutants that are lethal in the absence of the Nhp6 high-mobility-group protein. 1522 42

We have investigated the role of phosphorylation by vertebrate protein kinase CK2 on the activity of the General Transcription Factors TFIIA, TFIIE, TFIIF, and RNAPII. The largest subunits of TFIIA, TFIIE, and TFIIF were phosphorylated by CK2 holoenzyme. Also, RNA polymerase II was phosphorylated by CK2 in the 214,000 and 20,500 daltons subunits. Our results show that phosphorylation of TFIIA, TFIIF, and RNAPII increase the formation of complexes on the TATA box of the Ad-MLP promoter. Also, phosphorylation of TFIIF increases the formation of transcripts, where as phosphorylation of RNA polymerase II dramatically inhibits transcript formation. Furthermore, we demonstrate that CK2 beta directly interacts with RNA polymerase II, TFIIA, TFIIF, and TBP. These results strongly suggest that CK2 may play a role in regulating transcription of protein coding genes.
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PMID:Effects of phosphorylation by protein kinase CK2 on the human basal components of the RNA polymerase II transcription machinery. 1535 56

7,8-Dihydro-8-oxoguanine (8-oxoG) is the most frequent mutagenic lesion caused by oxidative stress. Eukaryotic cells use a specific DNA glycosylase, OGG1, to excise 8-oxoG from DNA. The mild phenotype of OGG1 null mice has been attributed to the existence of alternative pathways, including Cockayne syndrome B (CSB)-dependent transcription coupled repair (TCR), for removal of 8-oxoG. We have studied repair and transcription activities at 8-oxoG lesions with a reconstituted transcription system (RTS; RNA polymerase II, TBP, TFIIA, TFIIB, TFIIE, TFIIF and TFIIH), as well as in cellular extracts and in vivo. All measurable repair activity at 8-oxoG lesions takes place in the 3'-direction from the lesion, indicating base excision repair (BER) activity and negligible role of nucleotide excision repair (NER). Although 8-oxoG has been shown to be preferentially removed from the transcribed strand, in vitro experiments with purified transcription factors failed to identify a definite block for RNA polymerase II at the lesion. However, a weak block was observed at the lesion during transcription carried out with RTS as well as with cellular extracts. RNA polymerase II was identified at the site of the lesion on obstructed templates. Wild-type cells, as well as cells carrying targeted mutations of genes required for removal of 8-oxoG, were transfected with a luciferase expression vector containing an 8-oxoG lesion. No significant obstruction at 8-oxoG lesions was observed by this in vivo approach. In control experiments transcription elongation was completely blocked by cisplatin.
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PMID:Transcription activities at 8-oxoG lesions in DNA. 1538 Jan 1

Negative cofactor 2 (NC2) forms a stable complex with TATA-binding protein (TBP) on promoters. This prevents the assembly of transcription factor (TF) IIA and TFIIB and leads to repression of RNA polymerase II transcription. Here we have revisited the interactions of NC2.TBP with DNA. We show that NC2.TBP complexes exhibit a significantly reduced preference for TATA box sequences compared with TBP and TBP.TFIIA complexes. In chromatin immunoprecipitations, NC2 is found on a variety of human TATA-containing and TATA-less promoters. Substantial amounts of NC2 are present in a complex with TBP in bulk chromatin. A complex of NC2.TBP displays a K(D) for DNA of approximately 2 x 10(-9) m for a 35-bp major late promoter oligonucleotide. While preferentially recognizing promoter-bound TBP, NC2 also accelerates TBP binding to promoters and stabilizes TBP.DNA complexes. Our data suggest that NC2 controls TBP binding and maintenance on DNA that is largely independent of a canonical TATA sequence.
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PMID:Efficient binding of NC2.TATA-binding protein to DNA in the absence of TATA. 1557 13

The BTAF1 transcription factor interacts with TATA-binding protein (TBP) to form the B-TFIID complex, which is involved in RNA polymerase II transcription. Here, we present an extensive mapping study of TBP residues involved in BTAF1 interaction. This shows that residues in the concave, DNA-binding surface of TBP are important for BTAF1 binding. In addition, BTAF1 interacts with residues in helix 2 on the convex side of TBP as assayed in protein-protein and in DNA-binding assays. BTAF1 drastically changes the TATA-box binding specificity of TBP, as it is able to recruit DNA-binding defective TBP mutants to both TATA-containing and TATA-less DNA. Interestingly, other helix 2 interacting factors, such as TFIIA and NC2, can also stabilize mutant TBP binding to DNA. In contrast, TFIIB which interacts with a distinct surface of TBP does not display this activity. Since many proteins contact helix 2 of TBP, this provides a molecular basis for mutually exclusive TBP interactions and stresses the importance of this structural element for eukaryotic transcription.
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PMID:Mutational analysis of BTAF1-TBP interaction: BTAF1 can rescue DNA-binding defective TBP mutants. 1617 47

Our previous work suggests that the Nhp6 HMGB protein stimulates RNA polymerase II transcription via the TATA-binding protein TBP and that Nhp6 functions in the same functional pathway as the Gcn5 histone acetyltransferase. In this report we examine the genetic relationship between Nhp6 and Gcn5 with the Mot1 and Ccr4-Not complexes, both of which have been implicated in regulating DNA binding by TBP. We find that combining either a nhp6ab or a gcn5 mutation with mot1, ccr4, not4, or not5 mutations results in lethality. Combining spt15 point mutations (in TBP) with either mot1 or ccr4 also results in either a growth defect or lethality. Several of these synthetic lethalities can be suppressed by overexpression of TFIIA, TBP, or Nhp6, suggesting that these genes facilitate formation of the TBP-TFIIA-DNA complex. The growth defect of a not5 mutant can be suppressed by a mot1 mutant. HO gene expression is reduced by nhp6ab, gcn5, or mot1 mutations, and the additive decreases in HO mRNA levels in nhp6ab mot1 and gcn5 mot1 strains suggest different modes of action. Chromatin immunoprecipitation experiments show decreased binding of TBP to promoters in mot1 mutants and a further decrease when combined with either nhp6ab or gcn5 mutations.
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PMID:Genetic interactions between Nhp6 and Gcn5 with Mot1 and the Ccr4-Not complex that regulate binding of TATA-binding protein in Saccharomyces cerevisiae. 1627 10


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