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)

RNA polymerase II subunit composition, stoichiometry, and phosphorylation were investigated in Saccharomyces cerevisiae by attaching an epitope coding sequence to a well-characterized RNA polymerase II subunit gene (RPB3) and by immunoprecipitating the product of this gene with its associated polypeptides. The immunopurified enzyme catalyzed alpha-amanitin-sensitive RNA synthesis in vitro. The 10 polypeptides that immunoprecipitated were identical in size and number to those previously described for RNA polymerase II purified by conventional column chromatography. The relative stoichiometry of the subunits was deduced from knowledge of the sequence of the subunits and from the extent of labeling with [35S]methionine. Immunoprecipitation from 32P-labeled cell extracts revealed that three of the subunits, RPB1, RPB2, and RPB6, are phosphorylated in vivo. Phosphorylated and unphosphorylated forms of RPB1 could be distinguished; approximately half of the RNA polymerase II molecules contained a phosphorylated RPB1 subunit. These results more precisely define the subunit composition and phosphorylation of a eucaryotic RNA polymerase II enzyme.
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PMID:RNA polymerase II subunit composition, stoichiometry, and phosphorylation. 218 13

RNA polymerases I, II, and III share three subunits that are immunologically and biochemically indistinguishable. The Saccharomyces cerevisiae genes that encode these subunits (RPB5, RPB6, and RPB8) were isolated and sequenced, and their transcriptional start sites were deduced. RPB5 encodes a 25-kD protein, RPB6, an 18-kD protein, and RPB8, a 16-kD protein. These genes are single copy, reside on different chromosomes, and are essential for viability. The fact that the genes are single copy, corroborates previous evidence suggesting that each of the common subunits is identical in RNA polymerases I, II, and III. Furthermore, immunoprecipitation of RPB6 coprecipitates proteins whose sizes are consistent with RNA polymerase I, II, and III subunits. Sequence similarity between the yeast RPB5 protein and a previously characterized human RNA polymerase subunit demonstrates that the common subunits of the nuclear RNA polymerases are well conserved among eukaryotes. The presence of these conserved and essential subunits in all three nuclear RNA polymerases and the absence of recognizable sequence motifs for DNA and nucleoside triphosphate-binding indicate that the common subunits do not have a catalytic role but are important for a function shared by the RNA polymerases such as transcriptional efficiency, nuclear localization, enzyme stability, or coordinate regulation of rRNA, mRNA, and tRNA synthesis.
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PMID:Subunits shared by eukaryotic nuclear RNA polymerases. 218 66

Four cDNAs encoding human polypeptides hRPB7.0, hRPB7.6, hRPB17, and hRPB14.4 (referred to as Hs10 alpha, Hs10 beta, Hs8, and Hs6, respectively), homologous to the ABC10 alpha, ABC10 beta, ABC14.5, and ABC23 RNA polymerase subunits (referred to as Sc10 alpha, Sc10 beta, Sc8, and Sc6, respectively) of Saccharomyces cerevisiae, were cloned and characterized for their ability to complement defective yeast mutants. Hs10 alpha and the corresponding Sp10 alpha of Schizosaccharomyces pombe can complement an S. cerevisiae mutant (rpc10-delta::HIS3) defective in Sc10 alpha. The peptide sequences are highly conserved in their carboxy-terminal halves, with an invariant motif CX2CX12RCX2CGXR corresponding to a canonical zinc-binding domain. Hs10 beta, Sc10 beta, and the N subunit of archaeal RNA polymerase are homologous. An invariant CX2CGXnCCR motif presumably forms an atypical zinc-binding domain. Hs10 beta, but not the archaeal subunit, complemented an S. cerevisiae mutant (rpb10-delta 1::HIS3) lacking Sc10 beta. Hs8 complemented a yeast mutant (rpb8-delta 1::LYS2) defective in the corresponding Sc8 subunit, although with a strong thermosensitive phenotype. Interspecific complementation also occurred with Hs6 and with the corresponding Dm6 cDNA of Drosophila melanogaster. Hs6 cDNA and the Sp6 cDNA of S. pombe are dosage-dependent suppressors of rpo21-4, a mutation generating a slowly growing yeast defective in the largest subunit of RNA polymerase II. Finally, a doubly chimeric S. cerevisiae strain bearing the Sp6 cDNA and the human Hs10 beta cDNA was also viable. No interspecific complementation was observed for the human hRPB25 (Hs5) homolog of the yeast ABC27 (Sc5) subunit.
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PMID:Four subunits that are shared by the three classes of RNA polymerase are functionally interchangeable between Homo sapiens and Saccharomyces cerevisiae. 765 87

The cDNA of a small subunit (hRPB14.4) of RNA polymerase II (or B) from HeLa cells has been cloned. A 127 residue peptide sequence (calculated molecular weight of 14,478; isoelectric point of 3.7) was deduced and compared to that of the homologous subunit of Saccharomyces cerevisiae polymerase (ABC23, encoded by the RPB6/RPO26 gene). About 50% of the total residues were found to be conserved between yeast and man, with the C-terminal two-third being the most conserved (72% identity). A putative leucine-zipper comprising four properly spaced leucine residues, but not preceded by a basic domain, was identified near the C-terminal end of both proteins.
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PMID:A 14.4 KDa acidic subunit of human RNA polymerase II with a putative leucine-zipper. 780 19

The diverse functions of Saccharomyces cerevisiae RNA polymerase II are partitioned among its 12 subunits, designated RPB1-RPB12. Although multiple functions have been assigned to the three largest subunits, RPB1, RPB2, and RPB3, the functions of the remaining smaller subunits are unknown. We have determined the function of one of the smaller subunits, RPB9, by demonstrating that it is necessary for accurate start site selection. Transcription in the absence of RPB9 initiates farther upstream at new and previously minor start sites both at the CYC1 promoter in vitro and at the CYC1, ADH1, HIS4, H2B-1, and RPB6 promoters in vivo. Immunoprecipitation of RNA polymerase II from cells lacking the RPB9 gene revealed that all of the remaining 11 subunits are assembled into the enzyme, suggesting that the start site defect is attributable solely to the absence of RPB9. In support of this hypothesis, we have shown that addition of wild-type recombinant RPB9 completely corrects for the start site defect seen in vitro. A mutated recombinant RPB9 protein, with an alteration in a metal-binding domain required for high temperature growth and accurate start site selection in vivo, was at least 10-fold less effective at correcting the start site defect in vitro. RPB9 appears to play a unique role in transcription initiation, as the defects revealed in its absence are distinct from those seen with mutants in RNA polymerase subunit RPB1 and factor e (TFIIB), two other yeast proteins also involved in start site selection.
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PMID:RNA polymerase II subunit RPB9 is required for accurate start site selection. 788 69

One key component of the eukaryotic transcriptional apparatus is the multisubunit enzyme RNA polymerase II. We have discovered that two of the subunits shared by the three nuclear RNA polymerases in the yeast Saccharomyces cerevisiae, RPB6 and RPB10, have counterparts among the Archaea.
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PMID:Halobacterial S9 operon contains two genes encoding proteins homologous to subunits shared by eukaryotic RNA polymerases I, II, and III. 804 7

A single-copy gene, homologous to the RPB6 gene from Saccharomyces cerevisiae, encoding a small phosphorylated subunit common to all three forms of nuclear DNA-dependent RNA polymerase was isolated from the fission yeast Schizosaccharomyces pombe. Its cDNA copy consists of an open reading frame of 142 codons and encodes an acidic protein (predicted pI 4.1) with a M(r) of 15,730. The genomic copy of Sz. pombe rpb6 contains an intron (219 nucleotides) located at codon 92, a position which does not correspond to the single intron of the S. cerevisiae gene. The sequencing of both genomic and cDNA copies of rpb6 allowed us to determine the probable positions of the start and stop of rpb6 transcription and to identify a putative TATA box. The primary structures of the Sz. pombe and S. cerevisiae Rpb6 proteins have 60.7% identity, with the same general organization: a highly acidic N-terminal region followed by a short basic region and a C terminus featuring a putative heptad Leu repeat. The C-terminal half of the sequence is particularly well conserved and, therefore, probably contains the most important functional domain. Moreover, a heterospecific complementation test showed that rpb6 from Sz. pombe fully complements a complete deletion of its S. cerevisiae homologue.
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PMID:The fission yeast Schizosaccharomyces pombe rpb6 gene encodes the common phosphorylated subunit of RNA polymerase and complements a mutation in the corresponding gene of Saccharomyces cerevisiae. 808 49

We isolated the cDNA encoding the homolog of the Saccharomyces cerevisiae nuclear RNA polymerase common subunit RPB6 from hamster CHO cells. Alignment of yeast RPB6 with its mammalian counterpart revealed that the subunits have nearly identical carboxy-terminal halves and a short acidic region at the amino terminus. Remarkably, the length and amino acid sequence of the hamster RPB6 are identical to those of the human RPB6 subunit. The conservation in sequence from lower to higher eukaryotes also reflects conservation of function in vivo, since hamster RPB6 supports normal wild-type yeast cell growth in the absence of the essential gene encoding RPB6.
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PMID:Functional substitution of an essential yeast RNA polymerase subunit by a highly conserved mammalian counterpart. 819 53

Cajal bodies (coiled bodies) are nuclear organelles that contain a variety of components required for transcription and processing of RNA. Cajal bodies in amphibian oocytes are stained by mAb H14, which recognizes the carboxy-terminal domain (CTD) of the largest subunit of RNA polymerase II when the heptapeptide repeat is phosphorylated on serine-5. Oocytes were treated with the transcription inhibitor 5, 6-dichloro-1-beta-d-ribofuranosylbenzimidazole (DRB), which prevents phosphorylation of the CTD. Cajal bodies from oocytes that had been treated for 2-3 h with DRB no longer stained with mAb H14, but staining reappeared when the inhibitor was washed out. Epitope-tagged transcripts of two small subunits of polymerase II, RPB6 and RPB9, were injected into the cytoplasm of Xenopus and Triturus oocytes. Newly translated RPB6 and RPB9 were specifically targeted to Cajal bodies within 4 h, and Cajal bodies remained the site of highest concentration of tagged protein during the next 2 days. These data suggest that polymerase subunits pass through the Cajal bodies with a transit time no greater than a few hours. We discuss the possibility that Cajal bodies are sites of assembly or modification of the transcription machinery of the nucleus.
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PMID:RNA polymerase II in Cajal bodies of amphibian oocytes. 1080 76

Bacterial DNA-dependent RNA polymerase (RNAP) has subunit composition beta'betaalpha(I)alpha(II)omega. The role of omega has been unclear. We show that omega is homologous in sequence and structure to RPB6, an essential subunit shared in eukaryotic RNAP I, II, and III. In Escherichia coli, overproduction of omega suppresses the assembly defect caused by substitution of residue 1362 of the largest subunit of RNAP, beta'. In yeast, overproduction of RPB6 suppresses the assembly defect caused by the equivalent substitution in the largest subunit of RNAP II, RPB1. High-resolution structural analysis of the omega-beta' interface in bacterial RNAP, and comparison with the RPB6-RPB1 interface in yeast RNAP II, confirms the structural relationship and suggests a "latching" mechanism for the role of omega and RPB6 in promoting RNAP assembly.
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PMID:Bacterial RNA polymerase subunit omega and eukaryotic RNA polymerase subunit RPB6 are sequence, structural, and functional homologs and promote RNA polymerase assembly. 1115 66


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