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)

Bovine lymphosarcoma tissue has been extracted with low- and high-salt buffers [0.05 M Tris-C1 plus or minus 0.3 M (NH4) 2S04]. Diethylaminoethyl-Sephadex chromatography of both the high-salt and low-salt extracts yields RNA polymerases I and II, although low-salt extraction releases only one-third as much activity. Extraction by high salt of the residue from the low-salt extract, followed by diethylaminoethyl-Sephadex chromatography, yields additional enzyme activity with properties of Form II. Purification of the low-salt extract by protamine precipitation, elution with sodium succinate, and phosphocellulose chromatography yields a preparation of RNA polymerase (RNAP) with hybrid properties, combining the salt optimum of Form I, diethylaminoethyl-Sephadex elution pattern of form II, and alpha-amanitin sensitivity of Form III. RNAP. transcribes native D,A and chromatin efficiently. More RNAPL is recovered from lymphosarcoma tissue than from calf thymus.
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PMID:RNA polymerase isolated from bovine lymphosarcoma by sequential low- and high-salt extraction. 117 19

Poly(A) polymerase has been extensively purified from low-salt extracts of bovine lymphosarcoma. The enzyme is Mn2+ dependent, requires an oligonucleotide or RNA primer, incorporates only adenosine triphosphate, and is inhibited by other ribonucleotides or deoxynucleotides. Oligoadenylate and ribosomal RNA are good primers for the enzyme; transfer RNA and poly(A) are poor. RNA transcribed in vitro by homologous RNA polymerase is an efficient primer. The properties of the enzyme are similar to the properties of the Mn2+ -activated poly(A) polymerase of calf thymus. Approximately the same amount of enzyme appears to be present in lymphosarcoma and calf thymus.
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PMID:Poly (A) polymerase of bovine lymphosarcoma. 117 55

DNA-dependent RNA polymerase was solubilized from nuclei of Ehrlich ascites carcinoma (EAC) cells by sonic disruption in the presence of 0.3 M (NH4)2 SO4, and the multiple RNA polymerases were separated by chromatography on DEAE-Sephadex A-25. Elution with a nine-step gradient of (NH4)2 SO4 yielded five peaks of activity designated RNA polymerases Ia, Ib, IIa, IIb, and III, of which IIb was the most prominent. Linear-gradient elusion also yielded five peaks of the same designation, but Ia and Ib, as well as IIa and IIb, were not well separated. IIa and IIb were inhibited completely by 0.1 mug alpha-amanitin/ml, whereas the other forms were not. EAC RNA polymerases Ia, Ib, IIa, and IIb possessed Mg2+ ion, Mn2+ ion, and (NH4)2 SO4 optima, molecular weights, and thermal sensitivities similar to those reported for other mammalian DNA-dependent RNA polymerases. As measured by relative ribonucleoside monophosphate incorporation, with native calf thymus DNA template, EAC RNA polymerases Ia and Ib synthesized ribosomal RNA-like products, whereas forms IIa, IIb, and the parent enzyme mixture synthesized compounds that were more similar to DNA. No species specificity was found for DNA templates, and denatured DNA was consistently preferred to the native template by RNA polymerases IIa and IIb; the two kinds of template were about equally efficient for RNA polymerases Ia and Ib. Although EAC RNA polymerases Ia, IIa, and IIb were inhibited by daunomycin, form IIa was preferentially affected. 3',5'-Cyclic AMP, 3',5'-Cyclic GMP, and gibberellic acid, implicated as RNA polymerase regulators in other systems, were generally ineffective. The levels of nuclear RNA polymerase activities, per mg DNA, of 3 mouse ascites tumors (EAC, 6C3HED lymphosarcoma, and TA3 adenocarcinoma) were compared with those from 3 normal mouse tissues (kidney, liver, and spleen). On the average, the tumor cell nuclei contained (per mg of DNA) 8.9, 1.5, 2.7, 20.0, and 3.8 times the activities of RNA polymerases Ia, Ib, IIa, IIb, and III, respectively, as the normal cells, but the difference was significantly only for IIb.
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PMID:DNA-dependent RNA polymerases of Ehrlich carcinoma, other murine ascites tumors, and murine normal tissues. 117 42

A simple and rapid method to copurify RNA polymerase I and the glucocorticoid-regulated transcription factor, TFIC, is described. This protocol results in 1262-fold purification and 15% total recovery of the enzyme and factors needed to support faithful transcription in vitro from cloned mouse rRNA gene (rDNA). Using this method, proteins involved in rDNA transcription were purified from exponentially growing lymphosarcoma P1798 cells as well as cells treated with 0.1 microM dexamethasone. A combination of transcription and reconstitution assays using G-free cassette-containing constructs and polyacrylamide gel electrophoretic analysis upon silver staining were used to detect TFIC activity as well as the characteristic TFIC polypeptides in control and dexamethasone-treated cell extracts. Treatment of P1798 cells with 0.1 microM dexamethasone for 24 h results in an over 95% reduction of TFIC activity, but no significant differences in the amount of TFIC polypeptides in the final product purified from control and glucocorticoid-treated cells could be detected. Our data indicate that glucocorticoid regulation of transcription of rDNA is mediated via post-translational modulation of the activity of TFIC.
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PMID:Copurification of RNA polymerase I and the glucocorticoid-regulated transcription factor IC. 145 55

Glucocorticoids reversibly inhibit transcription of ribosomal RNA genes in murine lymphosarcoma P1798 cells in culture. Inhibition of rDNA transcription is due to reduction in the amount or activity of an RNA polymerase I transcription factor called transcription factor IC (TFIC). TFIC has been purified over 100,000-fold. The highly purified preparation contains neither RNA polymerase I activity nor any of the conventional RNA polymerase I subunits. TFIC activity co-purifies with three polypeptides of approximately 55, 50, and 42 kDa molecular mass. These polypeptides are present in a stoichiometric ration of 1:1:1.
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PMID:Hormonal regulation of transcription of rDNA. Purification and characterization of the hormone-regulated transcription factor IC. 239 50

Effects of cyclosporin A (CsA) on rRNA synthesis in vivo and in vitro were studied using lymphosarcoma P1798 in culture. Pulse labeling with [3H]uridine indicated that treatment of P1798 cells with 1 microgram/ml of CsA for 24-h reduced rRNA levels by 50-60%, whereas rRNA levels of cells rescued from CsA and grown for 24 h were similar to those of controls. Transcription experiments using nuclei from control, treated, and rescued cells indicated that the reduction in rRNA synthesis in treated cells was due to reversible inhibition of transcription of rDNA. Transcription studies in vitro indicated that S100 extracts from CsA-treated cells were unable to carry out faithful transcription of cloned mouse rDNA, even though RNA polymerase I levels of control and treated cell extracts were similar. Mixing experiments indicated that the inability of the CsA-treated cell extract to transcribe cloned rDNA in vitro was not due to the presence of inhibitor(s) or nuclease(s) in such extracts. Supplementation of CsA-treated cell extract with partially or highly purified preparations of a transcription initiation factor for RNA polymerase I, obtained from control cell extracts, conferred transcriptional ability on the CsA-treated cell extract. Extracts from cells treated with cyclosporin H, an inactive analogue of CsA, faithfully transcribed rDNA, indicating the specificity of CsA action. These data indicate that CsA-treated cells lack the ability to initiate rDNA transcription in vivo and in vitro, due to specific, reversible reduction in the amount or activity of transcription factor IC. Significance of these results in understanding the mechanisms of the lymphostatic activity of CsA is discussed.
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PMID:Cyclosporin A inhibits rDNA transcription in lymphosarcoma P1798 cells. 368 Feb 46

Cell-free extracts of lymphosarcoma P1798 cell culture lines support faithful initiation upon the cloned mouse DNA encoding rRNA (rDNA) promoter, whereas extracts from cells treated for 16 hr with 0.1 microM dexamethasone cannot. Extracts from both sources transcribe the cloned 5S RNA gene in vitro and mixing experiments further demonstrate that inhibition of transcription of rDNA in vitro is not due to nucleases or inhibitors of transcription present in extracts from glucocorticoid-treated cells. Incubation of extracts from control cells at 45 degrees C for 15 min inactivates RNA polymerase I and abolishes transcription. Activity can be restored by the addition of partially purified RNA polymerase I from control cells and hormone-treated cells. Moreover, extracts from hormone treated cells can be reconstituted by the addition of a partially purified, heat-stable transcription factor from control cells.
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PMID:Glucocorticoid inhibition of initiation of transcription of the DNA encoding rRNA (rDNA) in lymphosarcoma P1798 cells. 632 67

A lymphosarcoma cell line designated P1798.S6M has been characterized with respect to glucocorticoid responsiveness in culture. Cells ceased to proliferate in the presence of 10(-7) M dexamethasone. Cell viability remained high and glucocorticoid-sensitive cells could be rescued from cultures treated with dexamethasone for 24 or 48 h. These data indicate that P1798.S6M undergoes reversible arrest in the presence of dexamethasone. The system was used to study the effects of mitotic arrest upon transcription of rDNA. Incorporation of [methyl-3H]methionine into rRNA was rapidly inhibited and pulse-chase experiments indicated that 28 S RNA was not synthesized after 24 h of exposure to dexamethasone. Hybridization studies indicated that the amount of pre-rRNA was reduced by 90 to 95% in cells treated for 24 h. Transcription studies were carried out in isolated nuclei. Twenty-four hours after addition of dexamethasone, template-bound RNA polymerase I activity decreased by 89 to 96%. Total RNA polymerase I activity did not decrease, whereas disengaged nuclear polymerase I activity increased dramatically. Filter hybridization studies indicated that labeling of nascent pre-rRNA chains in vitro was inhibited 93%. These data are interpreted as follows: Dexamethasone reversibly inhibits proliferation of P1798.S6M cells and transcription of rDNA. Total RNA polymerase I activity does not decrease, but the amount of template-bound enzyme is reduced with a concomitant increase in the amount of disengaged polymerase I. This indicates that initiation of transcription is inhibited in cells undergoing mitotic arrest in the presence of dexamethasone.
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PMID:Hormonal regulation of transcription of rDNA. Inhibition of transcription during glucocorticoid-mediated inhibition of proliferation of lymphosarcoma P1798 cells in culture. 668 17

Lymphosarcoma P1798 cells undergo growth arrest when exponentially growing cultures are exposed to 1 micrograms/ml of Rapamycin (Rapa). This growth arrest is accompanied by inhibition of RNA biosynthesis as measured by incorporation of 3H-uridine into the newly synthesized RNA. Approximately 50% inhibition of 3H-uridine incorporation was observed, upon exposure of P1798 cells to 1 microgram/ml Rapa for 24 h. Run-on transcription experiments using nuclei from Rapa-treated cells indicated a dose-dependent inhibition of transcription or rRNA genes. Cells were relieved from this inhibition of transcription when Rapa was removed from the medium. Under similar conditions, transcriptions of U3 snRNA genes remained unaffected. Cytoplasmic extracts prepared from P1798 cells treated with 1 microgram/ml Rapa for 24 h failed to support transcription from cloned mouse rRNA promoter. This treatment does not affect the RNA polymerase I activity of P1798 cells. Addition of a highly purified murine transcription initiation factor specific for RNA polymerase I reconstitutes the extracts from Rapa-treated P1798 cells. Our data indicate that this new immunosuppressive agent modulates transcription of rRNA genes via regulation of specific transcription factor function.
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PMID:Modulation of transcription of rRNA genes by rapamycin. 752 36

The effect of heat shock on RNA polymerase I (pol I)-directed transcription of the rRNA gene was studied in S-100 extract derived from mouse lymphosarcoma cells, and by in vivo labelling of rRNA. Exposure of cells to 42 degrees C for 2 h resulted in complete inhibition of rRNA synthesis in vivo. Pol I transcription was inhibited by 50% within 2 h of heat shock and was abolished after 3 h exposure at 42 degrees C. Under this condition, the core-promoter-binding activity of the factor (CPBF) that modulates pol I transcription was unaffected. In contrast, the promoter-binding activity of enhancer-1-binding factor, a protein related to the Ku autoantigen, which is involved in pol I transcription initiation, was reduced by 50 and 90% after 2 and 3 h of heat shock respectively. Western-blot analysis with antibodies specific for the two subunits of Ku protein showed the absence of p72 subunit after 3 h of heat shock. Under this condition, pol II transcription from the adenovirus major late promoter and pol III transcription of 5 S RNA gene remained unaffected. Mixing experiments ruled out the possibility that the inhibition of transcription was due to activation of nucleases or other inhibitors. This is the first report to show selective down-regulation of pol I transcription in vitro by heat shock and of the potential involvement of a pol I transcription factor in this process.
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PMID:Heat shock selectively inhibits ribosomal RNA gene transcription and down-regulates E1BF/Ku in mouse lymphosarcoma cells. 876 Mar 51


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