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)

A new class of fluorescent nucleotide analogs which contain the fluorophore 1-aminonaphthalene-5-sulfonate attached via a gamma-phosphoamidate bond has been synthesized. Both the purine and pyrimidine analogs have fluorescence emission maxima at 460 nm. Cleavage of the alpha-beta-phosphoryl bond produces change in both the absorption and fluorescence emission spectra. The fluorescence of the pyrimidine analogs is quenched; cleavage of the alpha-beta-phosphoryl bond of the UTP analog produces about a 14-fold increase in fluorescence intensity at 500 nm. Under the same conditions the fluorescence of the CTP analog increases about 8-fold, whereas the fluorescence of the purine analogs shows only a slight change. These derivatives are good substrates for Escherichia coli RNA polymerase with only slightly increased Km values and with Vmax values about 50 to 70% that of the normal nucleotides. They are used less efficiently by wheat germ RNA polymerase II. The ATP analog can be used by E. coli RNA polymerase to initiate RNA chains.
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PMID:Synthesis and properties of fluorescent nucleotide substrates for DNA-dependent RNA polymerases. 38 81

An attempt was made to elucidate the possible participation of low molecular weight nuclear RNAs in the transcription process. The parameters of the RNA synthesis cell-free system in isolated rat liver nuclei were studied for this purpose. Tissue homogenization in isotonic sucrose during isolation of the nuclei resulted in a loss of 4S nuclear RNA. The dependence of RNA synthesis on the salt concentration in the incubation medium is indicative of possible initiation of the RNA synthesis in this system. At salt concentration of 0.08 M 50-75% of incorporation is due to the activity of RNA polymerase II. Under these conditions the electrophoretically homogenous individual fractions of low molecular weight nuclear RNAs do not affect specifically the activity of RNA polymerase II. No evidence for the participation of low molecular weight nuclear RNAs in the mechanisms of heterogeneous nuclear RNA synthesis were obtained.
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PMID:[Effects of low molecular weight nuclear RNAs on the activity of RNA polymerase II in isolated rat liver nuclei]. 42 Aug 76

In an attempt to establish which RNA polymerase catalyzes the synthesis of the low molecular weight RNA components A, C and D, Ama 1 cells (mutant Chinese hamster cells) were used in experiments with addition of alpha-amanitin. Ama 1 cells contain an altered RNA polymerase II which is 800 times more resistant towards inhibition by alpha-amanitin than the wild type enzyme. Alpha-amanitin (up to 200 microgram/ml) added to these cells does not affect the synthesis of the low molecular weight RNAs A, C and D. These data together with our previous data showing that alpha-amanitin (0.5 - 5.0 microgram/ml) preferentially inhibits the synthesis of A, C and D in normal cells indicate that RNA polymerase II catalyzes the synthesis of the low molecular weight RNA components A, C and D.
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PMID:Synthesis of low molecular weight RNA components A, C and D by polymerase II in alpha-amanitin-resistant hamster cells. 42 95

L cell nuclear preparations were shown to transcribe RNA for periods up to 1 h at 37 degrees C. Nearly 70% of the transcription products were sensitive to inhibition by 1 microgram/mL of alpha-amanitin, indicating that they were transcribed by RNA polymerase II. Analysis of polyphosphorylated termini of in vitro synthesized RNA showed the presence of a phosphatase activity which prevents quantitative recovery of these termini. The finding of in vitro labeled polyphosphorylated termini in RNA greater than 12 S after short periods of incubation shows initiation in vitro for this size class. The labeling of these polyphosphorylated termini is decreased in the presence of rifamycin AF/013. The use of two apparent inhibitors of initiation, rifamycin AF/013 and 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB), has allowed detection of in vitro initiated transcripts of heterogeneous nuclear RNA. Both of these inhibitors act primarily at later times of incubation, in contrast to alpha-amanitin which acts on elongation and inhibits in vitro RNA synthesis immediately. The selective pattern of DRB inhibition on hnRNA is retained in vitro and some accumulation of large-size molecules is observed. It can be estimated that about 30% of the greater than 12S hnRNA sequences transcribed in vitro are sensitive to DRB and 48% of greater than 12S RNA are sensitive to rifamycin AF/013 inhibition.
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PMID:Nuclear transcription in vitro. Sensitivity to inhibition by ribosyldichlorobenzimidazole and rifamycin AF/013. 42 32

In vitro RNA synthesis by isolated RNA polymerase II of chicken myeloblastosis cells was shown to be highly sensitive to adriamycin inhibition. The template activity of the single-stranded DNA, purified by chromatography of denatured calf thymus DNA through hydroxylapatite columns, was found to be equally as sensitive to the inhibition as denatured calf thymus DNA. However, contrary to denatured DNA, the single-stranded DNA thus purified showed no significant binding to adriamycin as analyzed by cosedimentation of the drug and DNA through a sucrose gradient. This indicated that inhibition of RNA synthesis on a single-stranded DNA template might involve a mechanism other than DNA intercalation. Kinetic studies of the inhibition showed that the inhibition of RNA synthesis by adriamycin could not be reversed by increasing the concentrations of RNA polymerase and four nucleoside triphosphates, but it could be reversed by increasing DNA concentrations. Analysis of the size of RNA synthesized indicated that the ultimate size of the product RNA was not altered by adriamycin, suggesting that the drug may inhibit RNA synthesis by reducing RNA chain initiation.
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PMID:Inhibition of chicken myeloblastosis RNA polymerase II activity by adriamycin. 43 65

A heat-stable protein (HSF) that stimulates the activity of lamb thymus RNA polymerase II has been purified 2500-fold and partially characterized. This factor stimulates the activity of RNA polymerase II up to 13 times and retains complete activity when heated at 90 degrees C for 5 min. Stimulation is observed only in the presence of RNA polymerase II and requires native DNA as template. The stimulatory factor has a sedimentation coefficient of 2.7 S, a diffusion coefficient of 9.55 x 10(-7) cm2/s, and an isoelectric point of 8.0. Calculated from the sedimentation and diffusion data, the factor has a molecular weight of about 24,000. Electrophoresis of the purified factor on polyacrylamide gels in the presence of sodium dodecyl sulfate results in a single band corresponding to a molecular weight of 25,000. The number-average length of the RNA synthesized by RNA polymerase II is increased in the presence of the factor. Sedimentation velocity and exclusion chromatography experiments suggest that the stimulatory factor interacts with RNA polymerase II. These results suggest that the factor stimulates RNA synthesis through a direct interaction with RNA polymerase II. The stoichiometry of the HSF-RNA polymerase binding appears to be about 1:1. HSF is located in the nucleus, as determined by cell fractionation studies.
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PMID:Purification and partial characterization of a stimulatory factor for lamb thymus RNA polymerase II. 43 87

RNA polymerase II polypeptides present in [35S]methionine-labeled Chinese hamster ovary (CHO) cell extracts have been quantitatively immunoprecipitated with an anti-calf thymus RNA polymerase II serum. Analyses of the immunoprecipitates on sodium dodecyl sulfate polyacrylamide gels indicated that the immunoprecipitated polymerase II of both wild type CHO cells and the alpha-amanitin-resistant mutant Ama1 had polypeptides of molecular weight 214,000, 140,000, 34,000, 25,000, 23,000, 20,500, and 16,500. In heterozygous alpha-amanitin-resistant/alpha-amanitin-sensitive hybrid CHO cells, growth in the presence of alpha-amanitin results in the inactivation of the alpha-amanitin-sensitive RNA polymerase II activity and a compensating increase in the activity of the alpha-amanitin-resistant enzyme. Determination of the rates of synthesis and degradation of RNA polymerase II polypeptides using [35S]methionine labeling and polymerase II immunoprecipitation demonstrated that this increase in activity of alpha-amanitin-resistant polymerase II resulted from a co-ordinate increase in the rate of synthesis of at least three polypeptides of RNA polymerase II. At the same time, there was an enhanced rate of degradation of the alpha-amanitin-inactivated RNA polymerase II polypeptides.
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PMID:Regulated synthesis of RNA polymerase II polypeptides in Chinese hamster ovary cell lines. 43 82

Transcription of Adenovirus 2 DNA (Ad 2 DNA) by wheat germ RNA polymerase II in vitro satisfies criteria that have been used to establish that Escherichia coli or coliphage transcription in vitro is initiated at true promoters. (1) Wheat germ RNA polymerase forms highly stable complexes at specific sites on Ad 2 DNA, with a Kassoc of (4--5) X 10(10) M-1. (2) Electron microscopic visualization of enzyme bound to Ad 2 DNA reveals the location of eight strong binding sites, at least five of which appear to correspond to promoters that have been identified in studies of Ad 2 transcription in vivo [Evans, R. M., Fraser, N., Ziff, E., Weber, J., Wilson, M., & Darnell, J.E. (1977) Cell 12, 733--739; Berk, A.J., & Sharp, P.A. (1977) Cell 12, 45--55; Weinmann, R., & Aiello, L. O. (1978) Proc. Natl. Acad. Sci. U.S.A. 75, 1662--1666]. (3) Transcription of Ad 2 DNA from preformed complexes with wheat germ polymerase is capable of escaping the action of rifamycin AF/013 and is relatively resistant to polyriboinosinic acid. In addition, our results are consistent with a two-state model for the interaction of wheat germ RNA polymerase with Ad 2 DNA, indicating that the mechansisms of transcription initiation and promoter-site selection in eucaryotes may be very similar to mechanisms elucidated in procaryotic systems.
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PMID:Interaction between wheat germ RNA polymerase II and adenovirus 2 DNA. Evidence for two types of stable binary complexes. 46 76

Transcription of the genome of the nondefective parvovirus BPV was examined in nuclei isolated from synchronized bovine fetal spleen cells. The relative levels of total RNA polymerase and RNA polymerase I, II, and III activities in nuclei isolated from BPV-infected and mock-infected cells were found to be similar throughout the course of infection. Hybridization of RNA synthesized in isolated nuceli indicated that BPV-specific RNA synthesis began during the period of 8 to 12 h postinfection and proceeded linearly until at least 20 h postinfection. By 20 h postinfection, 5% of the total RNA synthesized in nuclei from infected cells was virus specific. BPV-specific RNA synthesis was inhibited by 95% in the presence of 0.1 microgram of alpha-amanitin per ml, suggesting that the viral genome is transcribed by cellular RNA polymerase II.
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PMID:Transcription of the bovine parvovirus genome in isolated nuclei. 48 Apr 72

We have developed a system for the in vitro transcription of specific genes in rooster liver chromatin by endogenous RNA polymerase II that maintains the specificity of transcription in vivo. Radioactive transcripts synthesized in vitro were identified and quantitated by hybridization to a vast excess of cloned cDNA. The cDNA preparations employed corresponded to vitellogenin mRNA, the synthesis of which is responsive to estrogen stimulation in vivo, and chicken serum albumin mRNA, the synthesis of which is not significantly affected by estrogen stimulation in vivo. Comparing the pattern of transcription of the albumin and vitellogenin genes in chromatin from the liver of the normal rooster with the pattern in chromatin from the liver of the estrogen-stimulated rooster, we found that prior estrogen treatment of the rooster is attended by a slight decrease in the differential rate of transcription of the albumin gene and approximately a 10-fold increase in the differential rate of transcription of the vitellogenin gene. Because this pattern of transcription reflects the estrogen-induced changes in transcription observed in vivo, chromatin preparations from the livers of normal and estrogen-stimulated roosters can be used to investigate regulation of specific gene transcription at the molecular level in vitro.
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PMID:Specific transcription in chicken liver chromatin by endogenous RNA polymerase II. Comparison of an estrogen-inducible gene with a constitutively expressed gene. 48 77


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