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)

Reverse transcriptase (RT; RNA-dependent DNA nucleotidyltransferase) from Rauscher leukemia virus is synthesized in infected cells by way of a read-through poly- rotein of 200,000 molecular weight. This polyprotein (Pr200(gag-pol)) was precipitated by antiserum to RT; in a previous study all the monospecific antisera to gag proteins recognized Pr200(gag-pol). Pr200(gag-pol) contains both p30 and RT peptide sequences. Intermediate RT-related precursors of 145,000 (Pr145(pol)), 135,000 (Pr135(pol)), and 125,000 (Pr125(pol)) molecular weights were specifically recognized by precipitation from infected cell extracts by antiserum to RT. These proteins shared methionine-containing tryptic peptide sequences with a virion polypeptide of 80,000 molecular weight (p80(pol)) precipitate by antiserum to RT. Purification of active RT enzyme from virions labeled with [(3)H]methionine showed that p80(pol) was the major component, based on analysis by gel electrophoresis and tryptic peptide mapping experiments. A polypeptide (Pr80(pol)), similar in size to mature viral p80(pol), was also precipitated from infected cells by antiserum to RT. Its peptide map was nearly identical to that of virion p80(pol). Pulse-chase studies showed that Pr80(pol), Pr125(pol), and Pr135(pol) were stable polypeptides, whereas Pr200(gag-pol) and Pr145(pol) were unstable precursors. Pulse-chase studies with the protein synthesis inhibitor, cycloheximide, showed that the processing of Pr200(gag-pol) occurred for a short time in the absence of protein synthesis.
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PMID:Biosynthesis of reverse transcriptase from Rauscher murine leukemia virus by synthesis and cleavage of a gag-pol read-through viral precursor polyprotein. 7 22

We have demonstrated by recombination of two highly pathogenic avian influenza viruses [A/FPV/Rostock (Hav1N1) x A/turkey/England/63 (Hav1Nav3)] that recombinants can be isolated which are pathogenic as well as non-pathogenic for chickens. They carried the glycoproteins of either parent strains, and all are produced in infectious form in chick embryo cells. Genetic analysis revealed that the non-pathogenic recombinants possess a mixed RNA polymerase complex, consisting of pol 1, pol 2, ptra and NP gene products, while, with one exception, the pathogenic recombinants have the genes coding for the polymerase activity from one or other parent virus. The biological properties of the recombinant viruses did not correlate with their pathogenicity for chickens.
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PMID:Correlation of pathogenicity and gene constellation of influenza A viruses. III. Non-pathogenic recombinants derived from highly pathogenic parent strains. 52 99

In vitro inhibitions by coumermycin A1 of DNA and RNA synthesis in toluenized cells were studied. In a sensitive strain, 50% inhibitions of replication and transcription were observed at 0.035 and 0.600 mug/ml, respectively. DNA synthesis in a toluenized-resistant mutant was 50% inhibited at 0.140 mug/ml of coumermycin A1, whereas RNA synthesis was unaffected at all concentrations tested. Studies with a mixture of toluenized-sensitive and -resistant bacteria ruled out the presence of a diffusable activator or inhibitor of coumermycin A1 action. Density label studies with toluenized pol A+ and pol A- strains indicated that replicative DNA synthesis was specifically inhibited, in agreement with the in vivo studies in the preceding paper of this issue (Ryan, M. J. (1976), Biochemistry 15). Highly purified Escherichia coli DNA polymerase III and RNA polymerase both were inhibited by this antibiotic. However, the high concentrations necessary for these inhibitions suggest that they are not biologically relevant. No interaction between DNA and coumermycin A1 was observed with the following analytical procedures: ultraviolet difference spectra, DNA absorbance-temperature transitions, equilibrium buoyant density centrifugation, and DNA cross-linking determinations.
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PMID:Coumerimycin A1: A preferential inhibitor of replicative DNA synthesis in Escherichia coli. II. In vivo characterization. 78 23

The objective of the present study was to compare the data of in situ hybridization (ISH), RNA polymerase chain reaction (PCR/RNA) and p24 core antigen (p24 Ag) enzyme immunoassay (EIA) for the detection of HIV-1 expression in peripheral blood mononuclear cells (PBMCs) and in plasma of infected patients at various CDC stages. PBMCs of 24 patients mostly of CDC stage II were obtained from heparinized blood samples, cytocentrifuged and hybridized with a (35S) labelled single-stranded RNA probe specific for gag-pol of LAVBru HIV-1 allowing the detection of genomic and/or messenger RNA. The corresponding plasma samples were used for the determination of p24 Ag by EIA and detection of HIV-1 genomic RNA by RT-PCR using specific primers in the LTR, gag and env regions. Whereas p24 was detected in only six out of 24 patients, both ISH and PCR/RNA enabled the detection of viral RNAs in more than 60% of the patients; cumulation of positive results of ISH and RT-PCR showed that 100% of patients at stage IV and 83% of patients at stages II/III have molecular signs of HIV expression therefore indicating that transcription of the provirus is a highly frequent event, even in the early stages of the disease, and, pleading for undertaking a very early antiviral chemotherapy.
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PMID:Analysis of HIV-1 expression in vivo with in situ hybridization and the polymerase chain reaction. 135 48

Premature termination of transcription by RNA polymerase II (pol II) occurs in the 5' region of many viral and cellular genes. Modulation of this process, or attenuation, is an important means of transcriptional control, but its mechanism is unknown. Using injected Xenopus oocytes, the efficiency of the mouse c-myc attenuator was tested when it was placed at various distances from the transcription initiation site. The attenuator functioned with each of six different pol II promoters tested; however, termination efficiency declined markedly when it was placed more than approximately 400 bases from the start site. This decline in attenuator function with distance from the start site coincided with increased sensitivity to the pol II inhibitor 5,6-dichloro-1-beta-D-ribofuranosyl benzimidazole (DRB). Thus transcription complexes situated further from the promoter appear to have a lower ability to recognize the attenuator and a greater sensitivity to DRB. Furthermore, polymerases which have read through one attenuation site have a reduced ability to terminate at a second site. The results imply that a discrete subset of elongation complexes is capable of premature termination, and that this subset exists only within the first few hundred bases of the transcription unit. Regulation of termination efficiency may be effected by changing the balance between the two modes of transcription committed either to read through or to terminate prematurely.
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PMID:Distinct modes of transcription read through or terminate at the c-myc attenuator. 137 47

Transcription elongation in a nuclear extract in vitro is efficiently blocked by Sarkosyl at a specific site downstream of the adenovirus major late (ML) promoter at which regulated transcription arrest has also been observed in vivo. In the experiments reported here, we examined the response of the polymerase to the ML attenuation site in two assay systems: 1) purified RNA polymerase II (pol II) transcribing tailed templates and 2) elongation complexes formed on immobilized templates and then depleted of elongation factors by extensive washing. Efficient site-specific arrest occurred in both systems, demonstrating that recognition of the site is an intrinsic property of the polymerase. However, the elongation properties of washed elongation complexes and purified pol II were not equivalent. In particular, the efficiency of arrest of washed elongation complexes was influenced both by the promoter from which transcription was initiated and by DNA sequences upstream from the attenuation site that did not contribute to the arrest of purified pol II. The polymerase and washed elongation complexes both remained in stable ternary complexes at the ML site with a lifetime of hours; addition of the elongation factor SII to these complexes promoted resumption of elongation. The efficiency of arrest in both systems was dependent on the solution concentration of the nucleotide incorporated at +187 (just beyond the attenuation site), indicating that pausing is an important part of the arrest mechanism. Based on this and other findings, we argue that the polymerase assumes an altered, elongation-incompetent conformation when arrest occurs.
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PMID:Mechanistic studies of transcription arrest at the adenovirus major late attenuation site. Comparison of purified RNA polymerase II and washed elongation complexes. 137 37

We investigated whether changes in expression of RNA polymerase III (pol III) or heterodisperse RNA polymerase II (pol II) transcripts hybridizing to Alu could be detected in Alzheimer's disease (AD). RNA samples obtained from AD and control brain tissues were examined by Northern analysis for Alu and 7SL RNA expression. All RNA samples contained a prominent band of approximately 300 nucleotides which corresponds to 7SL RNA, the Alu-homologous RNA component of the signal recognition particle. In addition, three small (i.e. less than 300 nucleotide) 7SL/Alu-hybridizing transcripts were detected. The two larger of the low molecular weight transcripts hybridized preferentially to the 7SL RNA probe, while the smallest transcript hybridized to the Alu probe. These transcripts and the heterodisperse RNA were variable in quantity and displayed a lack of correlation with AD.
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PMID:Expression of Alu and 7SL RNA in Alzheimer's and control brains. 137 21

The nucleotide sequence of 1731, a retrotransposon cloned from the genome of Drosophila melanogaster, reveals a structural similarity with the proviral form of the retroviruses including a pol-like gene containing a putative reverse-transcriptase(RT)-coding sequence. Diverse parts of that sequence were subcloned and expressed in Escherichia coli. It has been demonstrated that the expression of the RT-like sequence, when translated, gives rise to peptides displaying enzyme activity characteristic of a true RT enzyme. In addition, rabbit antisera directed against such recombinant proteins allowed us to detect an immunoreactive protein of around 110 kDa, which was only present in D. melanogaster cell lines, but not in cells derived from Drosophila virilis or Drosophila hydei, whose genomes do not bear the 1731 element. This protein is expected to correspond to a non-processed pol-gene translated product and cosediments with virus-like particles exhibiting RT activity.
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PMID:Characterization of the reverse transcriptase of 1731, a Drosophila melanogaster retrotransposon. 138 19

We have developed and characterized an efficient in vitro system from the protozoan, Acanthamoeba castellanii, that accurately initiates transcription from the adenovirus-2 major late promoter (AdMLP). Transcription by A. castellanii RNA polymerase II (pol II) is initiated at the same nucleotide (nt) that is used by HeLa extracts and is dependent upon adenovirus sequences located between nt -51 and the region around the transcription start point (tsp). The results suggest that the A. castellanii transcription factors for pol II which determine the tsp and the promoter elements that they recognize have been functionally conserved during evolution.
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PMID:Acanthamoeba castellanii RNA polymerase II transcription in vitro: accurate initiation at the adenovirus major late promoter. 139 30

Previously we have shown that nuclear extracts from mouse cells contain a heterogeneous group of polypeptides (p65, p80, p90, p100) which form distinct DNA-protein complexes on the 18 base-pair sequence element (termed Sal-box), which constitutes the murine rDNA transcription termination signal. These distinct proteins mediate cessation of RNA polymerase I (pol I) transcription elongation and release of the nascent RNA chains, indicating that they function as termination factor(s). Here, we report the biochemical analysis of the pol I-specific transcription termination factor TTFI. We show that the heterogeneity of TTFI is due to limited proteolysis of a larger, 130 kDa precursor protein (p130). The DNA-binding activity of p130 is strongly reduced as compared to the proteolytic derivatives, indicating that the DNA-binding domain is repressed within the full-length molecule. We have used limited proteolysis to purify and functionally characterize a TTFI core polypeptide (p50) which still specifically binds to the Sal-box target sequence and directs rDNA transcription termination. The equilibrium constant of purified p50 to bind specifically to DNA is 9 x 10(9) M-1. Additionally, we demonstrate that TTFI binds to DNA as a monomer and that binding induces DNA bending. This observation suggests that not only specific DNA-protein and protein-protein interactions but also conformational alterations of DNA may play a role in the termination process.
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PMID:Limited proteolysis unmasks specific DNA-binding of the murine RNA polymerase I-specific transcription termination factor TTFI. 140 80


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