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 I requires at least two nucleolar transcription factors, UBF and SL-1, for ribosomal RNA gene (rDNA) transcription. UBF requires SL-1 for the formation of a stable initiation complex on the rDNA promoter region. We have determined the region of mouse UBF (mUBF) required for nucleolar targeting. Although mUBF has a nuclear localization sequence, this sequence alone is not sufficient for mUBF to accumulate in the nucleolus. Deletion analyses show that mUBF requires a wide region except for the N-terminal 101 amino acids for nucleolar targeting. Deletion of either the HMG-box1, a region crucial for rDNA binding, or the acidic tail, a region that may interact with SL-1, results in the loss of nucleolar targeting. We show by DNA affinity analysis that the HMG-box1 is absolutely necessary for mUBF to bind to the upstream control element of the rDNA. We also show that mUBFs with various internal deletions retain both nucleolar targeting and DNA binding ability. A clear correlation was demonstrated between the DNA binding and nucleolar targeting ability. These results suggest that UBF is transferred to the nucleus by its NLS and is sequestered in the nucleolus by its specific and stable binding to the rDNA promoter via HMG-boxes and the acidic tail.
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PMID:Mouse rRNA gene transcription factor mUBF requires both HMG-box1 and an acidic tail for nucleolar accumulation: molecular analysis of the nucleolar targeting mechanism. 139 65

Two transcription factors, rat UBF (rUBF) and rat SL-1 are required for the efficient transcription of the rat promoter in vitro. In vitro studies have established that two broadly defined cis-acting domains, the core promoter element and the upstream promoter element, cooperate to direct correct transcription by RNA polymerase I. The ability of UBF to bind to two linker-scanning mutants of the upstream promoter element, which did not respond to the addition of UBF in in vitro transcription assays, was assessed by DNase footprinting. UBF protected the same region of the promoter in the linker-scanning mutant in BSM 129/124 as it did in the wild-type, but did not yield a typical footprint over the promoter in the linker-scanning mutant BSM 106/101. Previously we reported that promoters with mutant core promoters elements, either the guanine at -16 or -7 substituted by an adenine, were inactive in vitro unless the assays were supplemented with UBF. Those results suggested that the binding of UBF upstream of the core was required for the promotion of transcription. The interactions between the core and upstream promoter elements were assessed by constructing double mutants of the promoter. In two constructs the conserved guanines at either -16 or -7 were altered in a deletion mutant (-86) that did not respond to UBF. In a third construct the guanine at -16 in BSM 129/124 was changed to an adenine. These bidomain mutant constructs did not respond to the addition of UBF in an in vitro transcription assay, confirming that the rescue of the core promoter mutants requires an intact and functional upstream promoter element.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Complementary in vivo and in vitro analyses of the interactions between the cis-acting elements of the rat rDNA promoter. 192 91

The protein components that direct and activate accurate transcription by rat RNA polymerase I were studied in extracts of Novikoff hepatoma ascites cells. A minimum of at least two components, besides RNA polymerase I, that are necessary for efficient utilization of templates were identified. The first factor, rat SL-1, is required for species-specific recognition of the rat RNA polymerase I promoter and may be sufficient to direct transcription by pure RNA polymerase I. Rat SL-1 directed the transcription of templates deleted to -31, the 5' boundary of the core promoter element (+1 being the transcription initiation site). The second factor, rUBF, increased the efficiency of template utilization. Transcription of deletion mutants indicated that the 5' boundary of the domain required for rUBF lay between -137 and -127. Experiments using block substitution mutants confirmed and extended these observations. Transcription experiments using those mutants demonstrated that two regions within the upstream promoter element were required for optimal levels of transcription in vitro. The first region was centered on nucleotides -129 and -124. The 5' boundary of the second domain mapped to between nucleotides -106 and -101. DNase footprint experiments using highly purified rUBF indicated that rUBF bound between -130 and -50. However, mutation of nucleotides -129 and -124 did not affect the rUBF footprint. These results indicate that basal levels of transcription by RNA polymerase I may require only SL-1 and the core promoter element. However, higher transcription levels are mediated by additional interactions of rUBF, and possibly SL-1, bound to distal promoter elements.
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PMID:Characterization of factors that direct transcription of rat ribosomal DNA. 234 70

For efficient transcription from the rat ribosomal DNA (rDNA) promoter by RNA polymerase I in vitro, at least two transcription factors, rat UBF and rat SL-1, are required. Transcription cannot take place in vitro in the absence of SL-1. On the other hand, there is considerable difference of opinion concerning the necessity for UBF in in vitro transcription mediated by RNA polymerase 1, and the requirement for UBF is not clear. Mammalian cells code for UBF1 and UBF2, two forms of UBF that differ in HMG box-2, one of four HMG boxes or DNA-binding domains. We have used a monospecific antibody raised to recombinant rat UBF to determine whether UBF1 and UBF2 are required for RNA polymerase I-mediated transcription. This antibody can detect as little as 1.35 x 10(-15) moles of UBF1 or UBF2 in an immunoblot. Fractionated extracts that were competent for transcription had no detectable UBF1 or UBF2 when assayed in immunoblots with this antiserum. This evidence supports the hypothesis that UBF is not required for transcription of the rat rDNA promoter in vitro and most likely functions as an auxillary transcription factor. In addition, we have fractionated rat UBF1 from UBF2 and tested each of them in in vitro transcription assays in which the 45S or spacer rDNA promoter template is limiting. UBF1 can activate transcription from either the 45S or spacer promoter under these conditions, whereas UBF2 cannot. This implies that there is a functional difference in the transactivation of RNA polymerase I by UBF1 and UBF2 in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Transcription from the rat 45S ribosomal DNA promoter does not require the factor UBF. 801 25

Transcription of the 45S rRNA genes is carried out by RNA polymerase I and at least two trans-acting factors, upstream binding factor (UBF) and SL-1. We have examined the hypothesis that SL-1 and UBF interact. Coimmunoprecipitation studies using an antibody to UBF demonstrated that TATA-binding protein, a subunit of SL-1, associates with UBF in the absence of DNA. Inclusion of the detergents sodium dodecyl sulfate and deoxycholate disrupted this interaction. In addition, partially purified UBF from rat cell nuclear extracts and partially purified SL-1 from human cells coimmunoprecipitated with the anti-UBF antibody after mixing, indicating that the UBF-SL-1 complex can re-form. Treatment of UBF-depleted extracts with the anti-UBF antibody depleted the extracts of SL-1 activity only if UBF was added to the extract prior to the immunodepletion reaction. Furthermore, SL-1 activity could be recovered in the immunoprecipitate. Interestingly, these immunoprecipitates did not contain RNA polymerase I, as a monospecific antibody to the 194-kDa subunit of RNA polymerase I failed to detect that subunit in the immunoprecipitates. Treatment of N1S1 cell extracts with the anti-UBF antibody depleted the extracts of SL-1 activity but not TFIIIB activity, suggesting that the binding of UBF to SL-1 is specific and not solely mediated by an interaction between UBF and TATA-binding protein, which is also a component of TFIIIB. These data provide evidence that UBF and SL-1 interact.
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PMID:The species-specific RNA polymerase I transcription factor SL-1 binds to upstream binding factor. 855 83

Two genes encoding manganese superoxide dismutase (sod-2 and sod-3) have been identified in the nematode Caenorhabditis elegans. Each gene is composed of five exons, and intron positions are identical; however, intron sizes and sequences are not the same. The predicted protein sequences are 86.3% homologous (91.8% conservative), and the cDNAs are only 75.2% homologous. Both deduced protein sequences contain the expected N-terminal mitochondrial transit peptides. Reverse transcriptase polymerase chain reaction analysis shows that both genes are expressed under normal growth conditions and that their RNA transcripts are trans-spliced to the SL-1 leader sequence. The latter result together with Northern blot analysis indicate that both genes have mono-cistronic transcripts. The sod-3 gene was mapped to chromosome X, and the location of sod-2 was confirmed to be chromosome I. Polymerase chain reaction was used to amplify the cDNA regions encoding the predicted mature manganese superoxide dismutase proteins and each was cloned and expressed to high levels in Escherichia coli cells deficient in cytosolic superoxide dismutases. Both proteins were shown to be active in E. coli, providing similar protection against methyl viologen-induced oxidative stress. The expressed enzymes, which were not inhibited by hydrogen peroxide or cyanide, are dimeric, show quite different electrophoretic mobilities and isoelectric points, but exhibit comparable specific activities.
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PMID:Cloning, expression, and characterization of two manganese superoxide dismutases from Caenorhabditis elegans. 935 32

The genes that code for 45S rRNA, the precursor of 18S, 5.8S and 28S rRNA, are transcribed by RNA polymerase I. In many eukaryotes the genes are arranged as tandem repeats in discrete chromosomal clusters. rDNA transcription and rRNA processing occur in the nucleolus. In vertebrates, at least two factors, SL-1 and UBF, specific for transcription by RNA polymerase I cooperate in the formation of the initiation complex. Interestingly, there are proteins analogous to SL-1 in unicellular eukaryotes, but the requirement for a UBF-like factor appears to vary. Recent advances in our understanding of the rDNA transcription system and its regulation have demonstrated overlap with the other nuclear transcription systems (RNA polymerase II and III). This is exemplified by the utilization of TBP as a component of SL-1 and the role of Rb in regulatory rDNA transcription.
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PMID:Transcription by RNA polymerase I. 951 85

Traditional models for transcription initiation by RNA polymerase I include a stepwise assembly of basic transcription factors/regulatory proteins on the core promoter to form a preinitiation complex. In contrast, we have identified a preassembled RNA polymerase I (RPI) complex that contains all the factors necessary and sufficient to initiate transcription from the rDNA promoter in vitro. The purified RPI holoenzyme contains the RPI homolog of TFIID, SL-1 and the rDNA transcription terminator factor (TTF-1), but lacks UBF, an activator of rDNA transcription. Certain components of the DNA repair/replication system, including Ku70/80, DNA topoisomerase I and PCNA, are also associated with the RPI complex. We have found that the holo-enzyme supported specific transcription and that specific transcription was stimulated by the RPI transcription activator UBF. These results support the hypothesis that a fraction of the RPI exists as a preassembled, transcriptionally competent complex that is readily recruited to the rDNA promoter, i.e. as a holoenzyme, and provide important new insights into the mechanisms governing initiation by RPI.
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PMID:Identification of a mammalian RNA polymerase I holoenzyme containing components of the DNA repair/replication system. 1047 42

Changes in masticatory muscle structure and function are either developmental, as seen in anomalies of facial form, or adaptive, as seen during procedures such as orthognathic surgery and functional-appliance orthodontic therapy. Remodelling of muscle extracellular matrix is pivotal in these processes. This turnover is mediated via members of the family of enzymes known as matrix metalloproteinases (MMP) and inhibited by the tissue inhibitors of metalloproteinases (TIMP). The aim here was to investigate the in vivo pattern of expression and distribution of MMPs and TIMPs in masseter muscle of humans with both normal and abnormal facial forms. Masseter muscle biopsies were taken from 10 patients, four with long-face syndrome and six normal controls as confirmed by cephalometry. Immunohistochemical techniques were used to show the pattern and distribution of MMPs and TIMP proteins in the muscle. Zymography of tissue extracts was used to determine the presence of MMP activity. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the presence of MMP and TIMP-2 mRNA. MMP-1 was expressed around the individual muscle fibres, especially in those fibre surfaces in contact with the interstices of the connective tissue and around blood vessels. MMP-9 staining was less intense and was expressed in the interstices of the connective tissue and around blood vessels. Zymography of protein extracts confirmed that MMP-9 activity was present. MMP-2 and MMP-3 were not expressed in the samples, although MMP-2 mRNA could be detected by RT-PCR and its activity could be detected by zymography. Intense TIMP-1 staining was present around each muscle fibre, in the interstices of the connective tissue and surrounding blood vessels; TIMP-2 mRNA could be detected in all samples. These staining patterns were seen in all biopsies examined and were irrespective of the facial form of the donor. These findings provide evidence that the mechanisms required for matrix remodelling are present in the human masseter muscle.
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PMID:Identification of matrix metalloproteinases and their tissue inhibitors type 1 and 2 in human masseter muscle. 1077 72

We have previously demonstrated that the protein encoded by the retinoblastoma susceptibility gene (Rb) functions as a regulator of transcription by RNA polymerase I (rDNA transcription) by inhibiting UBF-mediated transcription. In the present study, we have examined the mechanism by which Rb represses UBF-dependent rDNA transcription and determined if other Rb-like proteins have similar effects. We demonstrate that authentic or recombinant UBF and Rb interact directly and this requires a functional A/B pocket. DNase footprinting and band-shift assays demonstrated that the interaction between Rb and UBF does not inhibit the binding of UBF to DNA. However, the formation of an UBF/Rb complex does block the interaction of UBF with SL-1, as indicated by using the 48 kDa subunit as a marker for SL-1. Additional evidence is presented that another pocket protein, p130 but not p107, can be found in a complex with UBF. Interestingly, the cellular content of p130 inversely correlated with the rate of rDNA transcription in two physiological systems, and overexpression of p130 inhibited rDNA transcription. These results suggest that p130 may regulate rDNA transcription in a similar manner to Rb.
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PMID:Rb and p130 regulate RNA polymerase I transcription: Rb disrupts the interaction between UBF and SL-1. 1104 86


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