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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The characteristics of Bacillus subtilis dnaF, a mutation specifying a temperature sensitive phenotype, were examined to determine its relationship to polC, the gene specifying the structure of DNA polymerase III (pol III). Exposure of growing cells bearing dnaF to non-permissive temperature inhibited replicative DNA synthesis and specifically depressed the expression of pol III activity in crude extracts. Highly purified pol III derived from cells bearing dnaF was temperature.sensitive in its polymerase activity, indicating that dnaF is a specific, polC mutation which specifies a structurally altered enzyme.
Mol Gen Genet 1978 Sep 08
PMID:Bacillus subtilis dnaF: a mutation of the gene specifying the structure of DNA polymerase III. 10 68

The ultraviolet (UV) sensitivity of Escherichia coli mutants deficient in the 5' leads to 3' exonuclease activity of DNA polymerase I is intermediate between that of pol+ strains and mutants which are deficient in the polymerizing activity of pol I (polA1). Like polA1 mutants, the 5'-exonclease deficient mutants exhibit increased UV-induced DNA degradation and increased repair synthesis compared to a pol+ strain, although the increase is not as great as in polA1 or in the conditionally lethal mutant BT4113ts deficient in both polymerase I activities. When dimer excision was measured at UV doses low enough to avoid interference from extensive DNA degradation, all three classes of polymerase I deficient mutants were found to remove dimers efficiently from their DNA. We conclude that enzymes alternative to polymerase I can operate in both the excision and resynthesis steps of excision repair and that substitution for either of the polymerase I functions results in longer patches of repair. A model is proposed detailing the possible events in the alternative pathways.
Mol Gen Genet 1977 Jan 07
PMID:Excision-repair in mutants of Escherichia coli deficient in DNA polymerase I and/or its associated 5' leads to 3' exonuclease. 31 38

The role of the three E. coli DNA polymerases (pol I, II, and III) in the replication of Col E1 DNA and other small plasmids with similar replicative properties was investigated in a soluble in vitro system prepared by freeze-thaw lysis of chloramphenicol-treated cells (Staudenbauer, 1976). Extracts from isogenic mutants of the polA, polB and polC gene loci deficient in pol I, II, and III respectively were examined for their replicative capacity. It was found that polA and polC extracts are deficient in the synthesis of supercoiled plasmid DNA, whereas the polB mutation has not effect. Deficient extracts could be complemented by addition of purified pol I and pol III holoenzyme. Analysis of the in vitro synthesized DNA by alkaline gradient centrifugation indicates that pol I is involved in an early step of the replication cycle whereas pol III is required at a later stage. These conclusions are confirmed by inhibition studies employing arabionsylcytosine triphosphate (aCTP) which is shown to interfere with pol III as well as pol II. The strong inhibitory effect of aCTP on plasmid replication is not influenced by the polB mutation and mimicks the effects of thermal inactivation of polC extracts. It is suggested that aCTP blocks plasmid ENA replication in vitro by interfering with pol III function
Mol Gen Genet 1976 Dec 08
PMID:REPLICAtion of small plasmids in extracts of Escherichia coli: requirement for both DNA polymerases I and II. 79 76

The amount and type of residual DNA synthesis was determined in eight temperature-sensitive mutants of the smut fungus Ustilago maydis after incubation at the restrictive temperature (32 degrees C) for eight hours. Mutants ts-220, ts-207, ts-432 and ts-346 were found to have an overall reduction in the synthesis of both nuclear and mitochondrial DNA comparison to the wild-type. In mutants ts-20, tsd 1-1, ts-84 and pol 1-1 nuclear DNA synthesis was depressed relative to mitochondrial synthesis. The DNA-polymerase mutant pol 1-1 had persistent nuclear synthesis at about 50% of the rate of synthesis of mitochondrial DNA and similar behavior was observed in a diploid homozygous strain. Mutant ts-84 had an initial burst of DNA synthesis which was reduced for nuclear but not mitochondrial synthesis after three hours preincubation at 32 degrees C. tsd 1-1 and ts-20 had nuclear residual synthesis amounting to about 25% of the relative rate of mitochondrial synthesis which correlates to increasing UV sensitivity of these strains on incubation at 32 degrees C. A pol 1-1 ts-84 double mutant had an additive loss of nuclear DNA synthesis which indicates that the steps of replication involved may be sequential.
Mol Gen Genet 1977 Jan 07
PMID:Differential chromosomal and mitochondrial DNA synthesis in temperature-sensitive mutants of Ustilago maydis. 83 76

The major DNA polymerase activity of wild-type U. maydis has been extensively purified. It possesses a molecular weight of about 150,000 daltons and appears to require a DNA primer with a 3'-hydroxyl terminus as well as a template. The polymerase activity has also been purified from the pol 1-1 strain, which is temperature sensitive fro growth and DNA synthesis, and which at the restrictive temperature contains only 10-25% levels of the DNA polymerase activity obtained from wild-type strains. It was similar in all properties studied, except that the activity was thermolabile at 40 degrees C compared to that from the wild-type strain. Physiological studies on the mutant showed that it was only slightly sensitive to UV, ionising radiation and nitrosoguanidine at the permissive temperature, and was proficient in genetic recombination. The results suggest that the pol 1-1 gene product does not play an important role in repair and recombination processes within the cell, and that its primary function lies in replication.
Mol Gen Genet 1975 Dec 30
PMID:DNA polymerase of Ustilago maydis: partial characterization of the enzyme and a pol 1 mutation. 122 4

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.
Mol Cell Probes 1992 Jun
PMID:Analysis of HIV-1 expression in vivo with in situ hybridization and the polymerase chain reaction. 135 48

The rat osteosarcoma cell line UMR 106-01 is a commonly used model system for the study of osteoblast function. However, it also expresses a phenotype characteristic of transformed cells. To test whether the latter could be accounted for by aberrant oncogene expression, we probed Northern blots of UMR and other osteoblastic cells with a panel of oncogene probes. These blots, when probed with a cDNA specific for v-H-ras, revealed a 7.0-kilobase (kb) H-ras-related transcript (designated HRRT) in UMR 106-01 cells that was not expressed in other osteoblastic cells. Osteoblast-enriched calvarial cells expressed the typical 1.1-kb H-ras mRNA, which was absent in UMR cells. Additionally, Western blots of lysates of UMR cells documented the presence of three proteins immunologically related to H-rasp21. To determine whether HRRT represented a recombinant retrovirus product, Northern blots were probed with a cDNA specific for the highly conserved gag-pol region of Moloney murine leukemia virus. These blots showed parallel cross-reactivity with an apparently identical transcript of 7.0 kb. The 7.0-kb transcripts detected by both v-H-ras and gag-pol probes declined to the same extent after treatment with concentrations of PTH known to inhibit proliferation of these cells. PTH regulated the abundance of HRRT in a time- and dose-dependent manner, with greatest repression of the transcript after 8 h of treatment with 10(-8) M PTH. The decrease in HRRT could not be completely accounted for by changes in transcriptional activity, as determined by nuclear run-on assays.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Endocrinol 1992 Sep
PMID:Regulation of an H-ras-related transcript by parathyroid hormone in rat osteosarcoma cells. 135 1

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.
J Mol Biol 1992 Oct 05
PMID:Limited proteolysis unmasks specific DNA-binding of the murine RNA polymerase I-specific transcription termination factor TTFI. 140 80

An RNA polymerase activity has been purified from pea (Pisum sativum) chloroplast extracts with a distinct transcriptional specificity for a chloroplast messenger gene. This activity (ms-RNA pol) differs from the pea RNA polymerase preparation reported by Sun, Shapiro, Wu & Tewari [(1986) Plant Mol. Biol. 6, 429-439], which specifically transcribes only the rRNA gene (rb-RNA pol). The specificity of transcription has been assessed by the synthesis in vitro of discrete transcripts of predicted sizes using cloned promoter regions of the chloroplast psbA and 16 S rRNA genes. The ms-RNA pol preparation, with polypeptides ranging in apparent molecular mass from 22 to 180 kDa, correctly initiates transcription from recombinant plasmids containing the psbA promoter and does not support 16 S rRNA promoter-directed transcription. The two activities differ also in their response to Mn2+ ions. To investigate whether the two transcriptional activities share common functional polypeptides, monoclonal antibodies were developed against the rb-RNA pol preparation. Three clones were selected on the basis of their ability to inhibit transcription in vitro of the 16 S rRNA gene by rb-RNA pol. The antibodies from these clones independently recognized three polypeptides with molecular masses of 27, 90 and 95 kDa on immunoblots. Antibodies cross-reacting with the 90 kDa polypeptide completely eliminated the specific retardation of an end-labelled 16 S rRNA promoter fragment in a mobility-shift assay, whereas the antibodies against the 95 kDa polypeptide resulted in the formation of a ternary complex (enzyme-DNA-antibody). The antibodies cross-reacting with the 27 kDa polypeptide, however, did not alter the mobility of the retarded DNA-enzyme complex on the gel. These antibodies also inhibited transcription in vitro of the psbA gene by ms-RNA pol and recognized polypeptides of identical molecular masses in the ms-RNA pol. These results show that the three polypeptides are functional components of the chloroplast transcriptional complex and appear to be involved in the transcription of both rRNA and mRNA genes. Transcriptional specificity is probably conferred by ancillary transcription factor(s) which remain to be identified.
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PMID:Two distinct transcriptional activities of pea (Pisum sativum) chloroplasts share immunochemically related functional polypeptides. 141 45

We have demonstrated recently that the genes encoding the U3 small nuclear RNA (snRNA) in dicot plants are transcribed by RNA polymerase III (pol III), and not RNA polymerase II (pol II) as in all other organisms studied to date. The U3 gene was the first example of a gene transcribed by different polymerases in different organisms. Based on phylogenetic arguments we proposed that a polymerase specificity change of the U3 snRNA gene promoter occurred during plant evolution. To map such an event we are examining the U3 gene polymerase specificity in other plant species. We report here the characterization of a U3 gene from wheat, a monocot plant. This gene contains the conserved promoter elements, USE and TATA, in a pol III-specific spacing seen also in a wheat U6 snRNA gene characterized in this report. Both the U3 and the U6 genes possess typical pol III termination signals but lack the cis element, responsible for 3'-end formation, found in all plant pol II-specific snRNA genes. In addition, expression of the U3 gene in transfected maize protoplasts is less sensitive to alpha-amanitin than a pol II-transcribed U2 gene. Based on these data we conclude that the wheat U3 gene is transcribed by pol III. This observation suggests that the postulated RNA polymerase specificity switch of the U3 gene took place prior to the divergence of angiosperm plants into monocots and dicots.
Plant Mol Biol 1992 Sep
PMID:Characterization of the U3 and U6 snRNA genes from wheat: U3 snRNA genes in monocot plants are transcribed by RNA polymerase III. 151 Nov 42


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