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)

We comment on the current understanding of transcriptional initiation by RNA polymerases I and III, and look for common modes of operation of these enzymes, emphasizing selected recent developments. These include definitive experiments on the constitution of the human RNA polymerase I transcription factor SL1/TIF-IB, the development of a genetic system for analyzing the function of RNA polymerase I in yeast, the elucidation of the structure of the human snRNA gene transcription factor SNAPc, and initial stages of mapping the protein-protein interactions involved in the assembly of transcriptional initiation complexes.
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PMID:Comparing transcriptional initiation by RNA polymerases I and III. 766 64

A new gene, RRN11, has been defined by certain rrn mutants of Saccharomyces cerevisiae which are defective specifically in the transcription of 35 S rRNA gene by RNA polymerase I (pol I). We have cloned the gene and found that it encodes a protein of 507 amino acids. We have used a strain with the chromosomal RRN11 deleted and carrying HA1 epitope-tagged RRN11 on a plasmid to isolate a protein complex containing the protein encoded by RRN11. This protein complex complemented rrn6 mutant extracts, which were previously shown to be deficient in the essential pol I transcription factor called Rrn6/7 complex or core factor (CF). The CF complex was previously shown to consist of three proteins, the 102- and 60-kDa subunits encoded by RRN6 and RRN7, respectively, and the 66-kDa subunit. The results of the above complementation experiments combined with mobility of Rrn11p in SDS-polyacrylamide gel electrophoresis analysis relative to Rrn6p and Rrn7p led to the conclusion that RRN11 encodes the 66-kDa subunit of CF. Glutathione S-transferase-Rrn11p fusion protein was found to bind strongly to 35S-labeled Rrn6p and Rrn7p but only weakly to 35S-labeled TATA-binding protein. Similarly, glutathione S-transferase-Rrn7p fusion protein bound strongly to 35S-labeled Rrn6p and Rrn11p but only weakly to 35S-labeled TATA-binding protein. These results are consistent with the fact that one can purify CF consisting of Rrn6p, Rrn7p, and Rrn11p from yeast cell extracts, but the purified complex does not contain TATA-binding protein. RRN11 was shown to be an essential gene, and [3H]uridine pulse experiments demonstrated directly that RRN11 is essential for rDNA transcription by pol I in vivo. Thus all three subunits of CF are essential for rDNA transcription. Because of the resemblance of CF to mammalian essential pol I transcription factor SL1, the amino acid sequences of Rrn11p and the other two subunits of CF were compared with those of the three TATA-binding protein-associated factors (TAFs) in the human SL1, TAFI48, TAFI63, and TAFI110. No significant similarity was detected between two sets of the proteins. Similarity as well as differences between CF and SL1 are discussed.
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PMID:RRN11 encodes the third subunit of the complex containing Rrn6p and Rrn7p that is essential for the initiation of rDNA transcription by yeast RNA polymerase I. 870 72

A crucial step in transcription is the recruitment of RNA polymerase to promoters. In the transcription of human rRNA genes by RNA Polymerase I (Pol I), transcription factor SL1 has a role as the essential core promoter binding factor. Little is known about the mechanism by which Pol I is recruited. We provide evidence for an essential role for hRRN3, the human homologue of a yeast Pol I transcription factor, in this process. We find that whereas the bulk of human Pol I complexes (I alpha) are transcriptionally inactive, hRRN3 defines a distinct subpopulation of Pol I complexes (I beta) that supports specific initiation of transcription. Human RRN3 interacts directly with TAF(I)110 and TAF(I)63 of promoter-selectivity factor SL1. Blocking this connection prevents recruitment of Pol I beta to the rDNA promoter. Furthermore, hRRN3 can be found in transcriptionally autonomous Pol I holoenzyme complexes. We conclude that hRRN3 functions to recruit initiation-competent Pol I to rRNA gene promoters. The essential role for hRRN3 in linking Pol I to SL1 suggests a mechanism for growth control of Pol I transcription.
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PMID:hRRN3 is essential in the SL1-mediated recruitment of RNA Polymerase I to rRNA gene promoters. 1125 Sep 3

The protein complex Selectivity Factor 1, composed of TBP, TAF(I)48, TAF(I)63 and TAF(I)110, is required for rRNA transcription by RNA polymerase I in the nucleolus. The steps involved in targeting Selectivity Factor 1 will be dependent on the transport pathways that are used and the localization signals that direct this trafficking. In order to investigate these issues, we characterized human TAF(I)48, a subunit of Selectivity Factor 1. By domain analysis of TAF(I)48, the carboxyl-terminal 51 residues were found to be required for the localization of TAF(I)48, as well as sufficient to direct Green Fluorescent Protein to the nucleus and nucleolus. The carboxyl-terminus of TAF(I)48 also has the ability to associate with multiple members of the beta-karyopherin family of nuclear import receptors, including importin beta (karyopherin beta1), transportin (karyopherin beta2) and RanBP5 (karyopherin beta3), in a Ran-dependent manner. This property of interacting with multiple beta-karyopherins has been previously reported for the nuclear localization signals of some ribosomal proteins that are likewise directed to the nucleolus. This study identifies the first nuclear import sequence identified within the TBP-Associated Factor subunits of Selectivity Factor 1.
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PMID:The carboxyl-terminus directs TAF(I)48 to the nucleus and nucleolus and associates with multiple nuclear import receptors. 1511 42

RNA polymerase I transcription in human cells requires Selectivity Factor 1, a multisubunit complex composed of the TATA-box-binding protein (TBP) and three TBP-associated factors (TAFs) called TAF(I)48, TAF(I)63 and TAF(I)110. Each of the Selectivity Factor 1 subunits binds directly to the other three components, but these interactions have not been characterized. This study is the initial identification and analysis of a TBP-binding domain within a Selectivity Factor 1 TAF. The interaction between human TBP and human TAF(I)48 was initially examined using the yeast two-hybrid assay, and a TBP-binding domain was identified in the carboxyl-terminus of human (h)TAF(I)48. Consistent with this result, the hTAF(I)48 carboxyl-terminus was able to bind directly to TBP in protein-protein interaction assays. When mutations were introduced into the hTAF(I)48 carboxyl-terminus, we identified changes in uncharged and positive residues that affect its interaction with TBP. By examining TBP mutants, residues within and adjacent to helix 2 of TBP, previously demonstrated to interact with subunits of other TBP-containing complexes [Transcription Factor IID (TFIID) and TFIIIB] were also found to diminish its affinity for the carboxyl-terminus of hTAF(I)48. The regions of hTAF(I)48 and TBP that interact are compared to those identified within other complexes containing TBP.
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PMID:Identification of a domain within human TAF(I)48, a subunit of Selectivity Factor 1, that interacts with helix 2 of TBP. 1531 21

Knowledge of the role of components of the RNA polymerase I transcription machinery is paramount to understanding regulation of rDNA expression. We describe key findings for the roles of essential transcription factor SL1 and activator upstream binding factor (UBF). We demonstrate that human SL1 can direct accurate Pol I transcription in the absence of UBF and can interact with the rDNA promoter independently and stably, consistent with studies of rodent SL1 but contrary to previous reports of human SL1. UBF itself does not bind stably to rDNA but rapidly associates and dissociates. We show that SL1 significantly reduces the rate of dissociation of UBF from the rDNA promoter. Our findings challenge the idea that UBF activates transcription through recruitment of SL1 at the rDNA promoter and suggest that the rate of pre-initiation complex (PIC) formation is primarily determined by the rate of association of SL1, rather than UBF, with the promoter. Therefore, we propose that SL1 directs PIC formation, functioning in core promoter binding, RNA polymerase I recruitment, and UBF stabilization and that SL1-promoter complex formation is a necessary prerequisite to the assembly of functional and stable PICs that include the UBF activator in mammalian cells.
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PMID:TBP-TAF complex SL1 directs RNA polymerase I pre-initiation complex formation and stabilizes upstream binding factor at the rDNA promoter. 1597 May 93

Lacking an RNA-dependent RNA polymerase, hepatitis delta virus (HDV), which contains a circular RNA of 1.7 kilobases, is nonetheless able to replicate its RNA by use of cellular transcription machineries. Previously, we have shown that the replications of genomic- and antigenomic-strand HDV RNAs have different sensitivities to alpha-amanitin, suggesting that these two strands are synthesized in different transcription machineries in the cells, but the nature of these transcription machineries is not clear. In this study, we performed metabolic labeling and immunofluorescence staining of newly synthesized HDV RNA with bromouridine after HDV RNA transfection into hepatocytes and confirmed that HDV RNA synthesis had both alpha-amanitin-sensitive and -resistant components. The antigenomic RNA labeling was alpha-amanitin resistant and localized to the nucleolus. The genomic RNA labeling was alpha-amanitin sensitive and more diffusely localized in the nucleoplasm. Most of the genomic RNA labeling appeared to colocalize with the PML nuclear bodies. Furthermore, promyelocytic leukemia protein, RNA polymerase II (Pol II), and the Pol I-associated transcription factor SL1 could be precipitated together with hepatitis delta antigen, suggesting the association of HDV replication complex with the Pol I and Pol II transcription machineries. This conclusion was further confirmed by an in vitro replication assay. These findings provide additional evidence that HDV RNA synthesis occurs in the Pol I and Pol II transcription machineries, thus extending the capability of the cellular DNA-dependent RNA polymerases to utilizing RNA as templates.
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PMID:RNA-templated replication of hepatitis delta virus: genomic and antigenomic RNAs associate with different nuclear bodies. 1677 35

Mitotic repression of rRNA synthesis requires inactivation of the RNA polymerase I (Pol I)-specific transcription factor SL1 by Cdk1/cyclin B-dependent phosphorylation of TAF(I)110 (TBP-associated factor 110) at a single threonine residue (T852). Upon exit from mitosis, T852 is dephosphorylated by Cdc14B, which is sequestered in nucleoli during interphase and is activated upon release from nucleoli at prometaphase. Mitotic repression of Pol I transcription correlates with transient nucleolar enrichment of the NAD(+)-dependent deacetylase SIRT1, which deacetylates another subunit of SL1, TAFI68. Hypoacetylation of TAFI68 destabilizes SL1 binding to the rDNA promoter, thereby impairing transcription complex assembly. Inhibition of SIRT1 activity alleviates mitotic repression of Pol I transcription if phosphorylation of TAF(I)110 is prevented. The results demonstrate that reversible phosphorylation of TAF(I)110 and acetylation of TAFI68 are key modifications that regulate SL1 activity and mediate fluctuations of pre-rRNA synthesis during cell cycle progression.
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PMID:Cooperative Action of Cdk1/cyclin B and SIRT1 Is Required for Mitotic Repression of rRNA Synthesis. 2602 73

In Saccharomyces cerevisiae, Core Factor (CF) is a key evolutionarily conserved transcription initiation factor that helps recruit RNA polymerase I (Pol I) to the ribosomal DNA (rDNA) promoter. Upregulated Pol I transcription has been linked to many cancers, and targeting Pol I is an attractive and emerging anti-cancer strategy. Using yeast as a model system, we characterized how CF binds to the Pol I promoter by electrophoretic mobility shift assays (EMSA). Synthetic DNA competitors along with anti-tumor drugs and nucleic acid stains that act as DNA groove blockers were used to discover the binding preference of yeast CF. Our results show that CF employs a unique binding mechanism where it prefers the GC-rich minor groove within the rDNA promoter. In addition, we show that yeast CF is able to bind to the human rDNA promoter sequence that is divergent in DNA sequence and demonstrate CF sensitivity to the human specific Pol I inhibitor, CX-5461. Finally, we show that the human Core Promoter Element (CPE) can functionally replace the yeast Core Element (CE) in vivo when aligned by conserved DNA structural features rather than DNA sequence. Together, these findings suggest that the yeast CF and the human ortholog Selectivity Factor 1 (SL1) use an evolutionarily conserved, structure-based mechanism to target DNA. Their shared mechanism may offer a new avenue in using yeast to explore current and future Pol I anti-cancer compounds.
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PMID:DNA binding preferences of S. cerevisiae RNA polymerase I Core Factor reveal a preference for the GC-minor groove and a conserved binding mechanism. 3138 53