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 gene coding for the largest subunit (RPA1) of RNA polymerase I (A) of Drosophila melanogaster (DmRPA1) was cloned and sequenced. The gene is interrupted by seven small introns and the cDNA reveals an open reading frame of 4932 nucleotides. The deduced polypeptide consists of 1644 amino acids with a calculated molecular weight of 185 kDa. Although the protein sequence exhibits the specific pattern of conserved regions found in all RNA polymerase largest subunits characterized so far, the overall sequence similarity among the RPA1 subunits of different species is much lower than seen with the corresponding subunits of RNA polymerases II and III. Two highly divergent hydrophilic domains characteristic for RPA1 separate the conserved blocks a and b in the N-terminal region and blocks g and h in the C-terminal section, respectively. In both cases the distance between the homologous blocks is enlarged by about 70 amino acids relative to the largest subunits of RNA polymerases II and III, and the corresponding subunit of the archaebacterial enzyme. Compared with RPA1 sequences of lower eukaryotes, the C-terminal hydrophilic domain in DmRPA1 is similar in length and acidity whereas the N-terminal domain is slightly shorter but retains the same basicity. The sequence insertions do not feature common motifs, suggesting a role for them in the interaction of RNA polymerase I with proteins required for the species-specific transcription of rDNA. The RPA1 subunits of Drosophila melanogaster and lower eukaryotes share an additional Zn-binding motif at the N-terminus with archaebacterial and RPC1 subunits, testifying to the complex evolutionary relationships among the RNA polymerases.
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PMID:Identification of the gene coding for the largest subunit of RNA polymerase I (A) of Drosophila melanogaster. 906 85

We describe the cloning and analysis of mRPA1, the cDNA encoding the largest subunit (RPA194) of murine RNA polymerase I. The coding region comprises an open reading frame of 5151 bp that encodes a polypeptide of 1717 amino acids with a calculated molecular mass of 194 kDa. Alignment of the deduced protein sequence reveals homology to the beta' subunit of Escherichia coli RNA polymerase in the conserved regions a-h present in all large subunits of RNA polymerases. However, the overall sequence homology among the conserved regions of RPA1 from different species is significantly lower than that observed in the corresponding beta'-like subunits of class II and III RNA polymerase. We have raised two types of antibodies which are directed against the conserved regions c and f of RPA194. Both antibodies are monospecific for RPA194 and do not cross-react with subunits of RNA polymerase II or III. Moreover, these antibodies immunoprecipitate RNA polymerase I both from murine and human cell extracts and, therefore, represent an invaluable tool for the identification of RNA polymerase I-associated proteins.
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PMID:Molecular cloning and characterization of the cDNA encoding the largest subunit of mouse RNA polymerase I. 923 75

Nucleotide excision repair (NER) of DNA in the yeast Saccharomyces cerevisiae and in human cells has been shown to be a biochemically complex process involving multiple gene products. In yeast, the involvement of the DNA replication accessory proteins, replication protein A (RPA1) and proliferating cell nuclear antigen (PCNA) in NER has not been demonstrated genetically. In this study we have generated temperature-degradable rfa1 and pcna mutants and show that these mutants are defective in NER in vitro under conditions that promote degradation of the RFA1 and PCNA gene products. We also demonstrate a physical interaction between RPA1 protein and subunits of the RNA polymerase II basal transcription factor IIH (TFIIH).
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PMID:The N-degron protein degradation strategy for investigating the function of essential genes: requirement for replication protein A and proliferating cell nuclear antigen proteins for nucleotide excision repair in yeast extracts. 980 17

Trypanosoma brucei harbors a unique multifunctional RNA polymerase (pol) I which transcribes, in addition to ribosomal RNA genes, the gene units encoding the major cell surface antigens variant surface glycoprotein and procyclin. In consequence, this RNA pol I is recruited to three structurally different types of promoters and sequestered to two distinct nuclear locations, namely the nucleolus and the expression site body. This versatility may require parasite-specific protein-protein interactions, subunits or subunit domains. Thus far, data mining of trypanosomatid genomes have revealed 13 potential RNA pol I subunits which include two paralogous sets of RPB5, RPB6, and RPB10. Here, we analyzed a cDNA library prepared from procyclic insect form T. brucei and found that all 13 candidate subunits are co-expressed. Moreover, we PTP-tagged the largest subunit TbRPA1, tandem affinity-purified the enzyme complex to homogeneity, and determined its subunit composition. In addition to the already known subunits RPA1, RPA2, RPC40, 1RPB5, and RPA12, the complex contained RPC19, RPB8, and 1RPB10. Finally, to evaluate the absence of RPB6 in our purifications, we used a combination of epitope-tagging and reciprocal coimmunoprecipitation to demonstrate that 1RPB6 but not 2RPB6 binds to RNA pol I albeit in an unstable manner. Collectively, our data strongly suggest that T. brucei RNA pol I binds a distinct set of the RPB5, RPB6, and RPB10 paralogs.
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PMID:Purification of an eight subunit RNA polymerase I complex in Trypanosoma brucei. 1673 80

African trypanosomes are the only organisms known to use RNA polymerase I (pol I) to transcribe protein-coding genes. These genes include VSG, which is essential for immune evasion and is transcribed from an extranucleolar expression site body (ESB). Several trypanosome pol I subunits vary compared to their homologues elsewhere, and the question arises as to how these variations relate to pol I function. A clear example is the N-terminal extension found on the second-largest subunit of pol I, RPA2. Here, we identify an essential role for this region. RPA2 truncation leads to nuclear exclusion and a growth defect which phenocopies single-allele knockout. The N terminus is not a general nuclear localization signal (NLS), however, and it fails to accumulate unrelated proteins in the nucleus. An ectopic NLS is sufficient to reinstate nuclear localization of truncated RPA2, but it does not restore function. Moreover, NLS-tagged, truncated RPA2 has a different subnuclear distribution to full-length protein and is unable to build stable pol I complexes. We conclude that the RPA2 N-terminal extension does not have a role exclusive to the expression of protein-coding genes, but it is essential for all pol I functions in trypanosomes because it directs trypanosomatid-specific interactions with RPA1.
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PMID:The trypanosomatid-specific N terminus of RPA2 is required for RNA polymerase I assembly, localization, and function. 2238 85

Transcription factor access to regulatory elements is prevented by the nucleosome. Heat shock factor 1 (HSF1) is a winged helix transcription factor that plays roles in control and stressed conditions by gaining access to target elements, but mechanisms of HSF1 access are not well known in mammalian cells. Here, we show the physical interaction between the wing motif of human HSF1 and replication protein A (RPA), which is involved in DNA metabolism. Depletion of RPA1 abolishes HSF1 access to the promoter of HSP70 in unstressed condition and delays its rapid activation in response to heat shock. The HSF1-RPA complex leads to preloading of RNA polymerase II and opens the chromatin structure by recruiting a histone chaperone, FACT. Furthermore, this interaction is required for melanoma cell proliferation. These results provide a mechanism of constitutive HSF1 access to nucleosomal DNA, which is important for both basal and inducible gene expression.
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PMID:RPA assists HSF1 access to nucleosomal DNA by recruiting histone chaperone FACT. 2294 Feb 45