Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

Yeast transcription factor TFIIH (also known as factor b) is a component of the RNA polymerase II initiation complex. Several TFIIH subunits (RAD3, SSL2 and SSL1) have also been implicated in DNA repair. Ssl1 interacts directly with another TFIIH subunit, Tfb1, which has not previously been shown to have a role in DNA repair. We isolated mutations in TFB1 that lead to a temperature sensitive phenotype. These mutations result in C-terminal truncations of the Tfb1 protein and disrupt its interaction with Ssl1. The C-terminal 114 amino acids of Tfb1 are necessary and sufficient for this interaction. Interestingly, cells carrying these truncations in Tfb1 cause sensitivity to ultraviolet (UV) light induced DNA damage, as previously observed for mutations in RAD3, SSL1 and SSL2. Many other mutations in RNA polymerase II basal factors were tested and found not to cause an increase in UV sensitivity, indicating that this phenotype is not due to a general defect in transcription.
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PMID:An interaction between the Tfb1 and Ssl1 subunits of yeast TFIIH correlates with DNA repair activity. 770 91

Nucleotide excision repair (NER) functions to remove DNA damage caused by ultraviolet light and by other agents that distort the DNA helix. The NER machinery has been conserved in structure and function from yeast to humans, and in humans, defective NER is the underlying cause of the cancer-prone disease xeroderma pigmentosum. Here, we reconstitute the incision reaction of NER in Saccharomyces cerevisiae using purified protein factors. The Rad14 protein, the Rad4-Rad23 complex, the Rad2 nuclease, the Rad1-Rad10 nuclease, replication protein A, and the RNA polymerase II transcription factor TFIIH were purified to near homogeneity from yeast. We show that these protein factors are both necessary and sufficient for dual incision of DNA damaged by either ultraviolet light or N-acetoxy-2-aminoacetylfluorene. Incision in the reconstituted system requires ATP, which cannot be substituted by adenosine 5'-O-(3-thiotriphosphate), suggesting that the hydrolysis of ATP is indispensable for the incision reaction. The excision DNA fragments formed as a result of dual incision are in the 24-27-nucleotide range.
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PMID:Reconstitution of yeast nucleotide excision repair with purified Rad proteins, replication protein A, and transcription factor TFIIH. 776 86

The essential TFB1 and SSL1 genes of the yeast Saccharomyces cerevisiae encode two subunits of the RNA polymerase II transcription factor TFIIH (factor b). Here we show that extracts of temperature-sensitive mutants carrying mutations in both genes (tfb1-101 and ssl1-1) are defective in nucleotide excision repair (NER) and RNA polymerase II transcription but are proficient for base excision repair. RNA polymerase II-dependent transcription at the CYC1 promoter was normal at permissive temperatures but defective in extracts preincubated at a restrictive temperature. In contrast, defective NER was observed at temperatures that are permissive for growth. Additionally, both mutants manifested increased sensitivity to UV radiation at permissive temperatures. The extent of this sensitivity was not increased in a tfb1-101 strain and was only slightly increased in a ssl1-1 strain at temperatures that are semipermissive for growth. Purified factor TFIIH complemented defective NER in both tfb1-101 and ssl1-1 mutant extracts. These results define TFB1 and SSL1 as bona fide NER genes and indicate that, as is the case with the yeast Rad3 and Ss12 (Rad25) proteins, Tfb1 and Ssl1 are required for both RNA polymerase II basal transcription and NER. Our results also suggest that the repair and transcription functions of Tfb1 and Ssl1 are separable.
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PMID:The yeast TFB1 and SSL1 genes, which encode subunits of transcription factor IIH, are required for nucleotide excision repair and RNA polymerase II transcription. 789 22

In yeast and humans, nucleotide excision repair (NER) of ultraviolet (UV)-damaged DNA requires a large number of highly conserved protein factors, which include the multisubunit RNA polymerase II transcription factor TFIIH. Here, we examine whether NER occurs by sequential assembly of different repair factors at the site of DNA damage or by the placement there of a "preformed" repairosome containing TFIIH and all the other essential NER factors. Contrary to the recent report (Svejstrup, J. Q., Wang, Z., Feaver, W. J., Wu, X., Bushnell, D. A., Donahue, T. F., Friedberg, E. C., and Kornberg, R. D. (1995) Cell 80, 21-28), our results provide no evidence for a pre-assembled repairosome; instead, they support the sequential assembly model. By several independent criteria, including co-purification, immunoprecipitation, and gel filtration of homogeneous proteins, we show that the damage recognition factor Rad14 exists in a ternary complex with the Rad1-Rad10 nuclease. We also find that Rad14 interacts directly with Rad1, but only slightly with Rad10, and that it interacts with the Rad1-Rad10 complex much more efficiently than with Rad1 alone. In the reconstituted NER system, a higher level of incision of UV-damaged DNA is achieved with the Rad1-Rad10-Rad14 complex, which we designate as nucleotide excision repair factor-1, NEF-1.
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PMID:Nucleotide excision repair in yeast is mediated by sequential assembly of repair factors and not by a pre-assembled repairosome. 862 33

The human basal transcription factor TFIIH plays a central role in two distinct processes. TFIIH is an obligatory component of the RNA polymerase II (RNAP II) transcription initiation complex. Additionally, it is believed to be the core structure around which some if not all the components of the nucleotide excision repair (NER) machinery assemble to constitute a nucleotide excision repairosome. At least two of the subunits of TFIIH (XPB and XPD proteins) are implicated in the disease xeroderma pigmentosum (XP). We have exploited the availability of the cloned XPB, XPD, p62, p44, and p34 genes (all of which encode polypeptide subunits of TFIIH) to examine interactions between in vitro-translated polypeptides by co-immunoprecipitation. Additionally we have examined interactions between TFIIH components, the human NER protein XPG, and the CSB protein which is implicated in Cockayne syndrome (CS). Our analyses demonstrate that the XPB, XPD, p44, and p62 proteins interact with each other. XPG protein interacts with multiple subunits of TFIIH and with CSB protein.
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PMID:Interactions involving the human RNA polymerase II transcription/nucleotide excision repair complex TFIIH, the nucleotide excision repair protein XPG, and Cockayne syndrome group B (CSB) protein. 865 57

The transcription factor TFIIH continues to be a subject of interest. In addition to its function as a repair and transcription factor, TFIIH includes a cyclin-dependent kinase and a cyclin, which raises the possibility that nucleotide excision repair (NER), RNA polymerase II transcription and cell cycle control are connected. Progress in mechanistic studies of NER include the identification of dual incision activities operating on either side of base damage and the isolation of a repairosome supercomplex in yeast. Additionally, NER has been demonstrated in reconstituted human and yeast systems, both of which include TFIIH.
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PMID:DNA repair and transcription. 872 69

The protein Tat is encoded by the HIV-1 genome and is essential for viral replication because of its activation of viral transcription. Tat enhances the ability of RNA polymerase II (Pol II) to move long distances down the DNA through a poorly understood mechanism that involves its binding the to the 5' end of the nascent HIV-1 transcript. It has been suggested that the stimulation of transcript elongation by conventional DNA-binding activators may involve phosphorylation of the carboxy-terminal domain (CTD) of Pol II by the transcription factor TFIIH through the associated CAK kinase. Here we show that Tat-enhanced HIV-1 transcription in vitro requires both TFIIH and the CTD of Pol II. In addition, Tat, through its activation domain, both interacts with a functional TFIIH-containing complex and stimulates phosphorylation of a CTD-containing substrate by the TFIIH kinase. Under conditions that jointly restrict transcriptional elongation and TFIIH-mediated CTD phosphorylation, Tat stimulates both these activities. Furthermore, RNA synthesis is required for Tat to stimulate phosphorylation of the CTD when it is part of an initiation complex, as expected from Tat's interaction with viral transcripts. Thus, stimulation of Pol II elongation by Tat may involve direct effects on TFIIH-mediated CTD phosphorylation.
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PMID:Enhanced processivity of RNA polymerase II triggered by Tat-induced phosphorylation of its carboxy-terminal domain. 893 26

It has recently been reported that the XPD (ERCC2) gene is an integral component of the basal transcription factor TFIIH. We have studied the direct role of this repair gene on the fine structure of DNA repair in hamster cells. The gene and strand specific DNA repair of UV induced pyrimidine dimers was determined in wild-type hamster cells, in hamster cells harboring a mutation in the gene homologous to the XPD gene and in mutant cells transfected with the human XPD gene. In the mutant cells, strand specific repair was severely deficient. In the transfected cells, preferential and strand specific gene repair were restored to wild-type levels. The results of the current study clearly demonstrate a direct role for the XPD gene product both in the preferential repair and bulk repair of pyrimidine dimers as well as its high functional conservation between rodent and human cells. An in vitro transcription assay was employed to investigate whether RNA polymerase II mediated transcription was also affected by the transfection with the XPD gene. No change in transcription between the mutant and transfected cells was observed. This suggests that the role of XPD in repair can be distinguished from its role in TFIIH dependent transcription initiation. Different functional domains of XPD appear to be necessary for repair versus transcription.
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PMID:Restoration of preferential and strand specific gene repair in group 2 Chinese hamster ovary mutants (UV5) by the XPD (ERCC2) gene. 911 Dec

The molecular mechanism of transcription-coupled nucleotide excision repair in eukaryotes is poorly understood. The identification of the dual role of basal transcription factor TFIIH in DNA repair and transcription provided a plausible link between both processes. However, TFIIH is not part of the elongating transcription complex, suggesting that additional components are required to recruit TFIIH when RNA polymerase II (RNAPII) stalls at the site of DNA damage. Previously, we have shown that the yeast Rad26 protein is involved in transcription-coupled DNA repair. This paper describes the differential contribution of the Rad26 protein to efficient removal of UV-induced cyclobutane pyrimidine dimers (CPDs) from transcribed DNA. Two distinct regions within the transcribed strand of RNAPII-transcribed genes are identified that differ in their requirement for the RAD26 gene product. Using high-resolution repair analysis, we determined the in vivo repair kinetics of cyclobutane pyrimidine dimers positioned around the transcription initiation site of RNAPII-transcribed genes RPB2 and URA3. Although transcription-coupled repair is severely reduced in rad26 mutants, lesions positioned in a small region immediately downstream of transcription initiation are efficiently removed in the absence of Rad26. The observed transition in repair characteristics is abrupt and in excellent agreement with the region where TFIIH dissociates from RNAPII in vitro, strongly suggesting an inverse correlation between TFIIH association and Rad26 requirement. These data suggest that a transcription repair coupling factor (Rad26/CSB) is required for efficient repair only during the elongating stages of RNAPII transcription.
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PMID:Transitions in the coupling of transcription and nucleotide excision repair within RNA polymerase II-transcribed genes of Saccharomyces cerevisiae. 922 8

Kin28/Cell, a cyclin-dependent kinase, is essential for the in vivo phosphorylation of the C-terminal domain of the largest subunit of RNA polymerase II in Saccharomyces cerevisiae. In a search for mutations co-lethal with a thermosensitive kin28 mutation, we have identified genes whose products interact functionally with Kin28. In the present work, we have studied a new complementation group of synthetic lethal mutations. The corresponding gene, RIG2, encodes a predicted RING finger protein. Rig2 is likely to be a homolog of MAT1 of higher eukaryotes which forms a ternary complex with MO15(cdk7) and cyclin H. Our genetic data suggest that Rig2 is a component of transcription factor TFIIH. Transcription activity in a rig2-ts mutant is impeded at restrictive temperature. However, none of the rig2-ts mutants obtained was UV sensitive, suggesting that Rig2 is dispensable for nucleotide excision repair.
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PMID:Rig2, a RING finger protein that interacts with the Kin28/Ccl1 CTD kinase in yeast. 929 30


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