Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous studies have shown that alkylation of MS2 RNA by certain derivatives of polycyclic aromatic hydrocarbons renders it noninfectious. Since phage RNA serves as a template for translation and transcription, either of these RNA-directed processes, or both, could be responsible in vivo for the inhibition of phage replication by metabolically activated hydrocarbons. The present study correlates the degree of inhibition of MS2 RNA infectivity, at various levels of alkylation by (+/-)-trans, 7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10-tetrahydrobenzol[a]pyrene, with the translation efficiency in vitro of the same alkylated RNA for the synthesis of viral synthetase and of maturation and coat proteins. The results indicate that dihydroxyepoxy-tetrahydrobenzo[a]pyrene modification of MS2 RNA impairs its template capacity for the synthesis of phage-specific proteins; this inhibition is insufficient, however, to account for the loss of RNA infectivity at lower molar ratios of alkylation. For the three viral proteins synthesized in vitro, the translation of RNA synthetase is much more sensitive to MS2 RNA modification than either coat or maturation protein synthesis. Our results also indicate that the loss of viral RNA infectivity follows a single-hit inactivation mechanism, whereas several alkylation events in the viral RNA synthetase cistron may be necessary to block translation of this gene product.
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PMID:Effect of benzo[a]pyrene-diolepoxide on infectivity and in vitro translation of phage MS2 RNA. 28 86

Mutagenic heterocyclic amines are metabolized to mutagens which act directly on Salmonella typhimurium by P-448 forms of cytochrome P-450. These direct mutagens are N-hydroxylated heterocyclic amines, such as N-hydroxy-Trp-P-1, N-hydroxy-Trp-P-2, N-hydroxy-Glu-P-1, N-hydroxy-Glu-P-2, N-hydroxy-IQ, N-hydroxy-2-amino-alpha-carboline (N-hydroxy-A alpha C), and N-hydroxy-2-amino-3-methyl-alpha-carboline (N-hydroxy-MeA alpha C). The treatment of rats with polychlorinated biphenyl stimulated N-hydroxylation of heterocyclic amines about 10- to 260-fold depending on the substrates used. The N-hydroxylation activities of purified cytochrome P-448-H and P-448-L were markedly different. P-448-H, which had very low activity for benzo[a] pyrene metabolic activation, showed high N-hydroxylation activity. The activity ratio P-448-H:P-448-L was markedly different depending on the amines used. This ratio was 45, 22, 3, and 0.02, respectively, for Glu-P-1, IQ, Trp-P-2, and benzo[a] pyrene. On the other hand, N-acetylation of the heterocyclic amines was very low. Although marked species differences in the N-acetylation were observed, the activities of the heterocyclic amines were about 1/100 of that of 2-aminofluorene. N-Hydroxy-Trp-P-2 could react directly to DNA, but N-hydroxy-Glu-P-1 could not. Therefore we need to consider the presence of a further activating system in mammalian and bacterial cells. We observed that N-hydroxy-Trp-P-2 was activated by prolyl-t-RNA synthetase, but N-hydroxy-Glu-P-1 was not activated by the same system. In the bacterial cells, both N-hydroxy-Trp-P-2 and N-hydroxy-Glu-P-1 were not activated by prolyl-t-RNA synthetase. However, both hydroxylamines were activated by the acetyl-CoA-dependent mechanism in mammalian and bacterial cells. These results indicated that the O-acetylation is an important pathway for DNA damage by heterocyclic amines in chemical carcinogenesis.
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PMID:Metabolic activation of mutagenic heterocyclic aromatic amines from protein pyrolysates. 351 87

Binding of small RNAs by the RNA-dependent RNA polymerase of coliphage Qbeta was studied utilizing a fluorometric assay. A DNA oligonucleotide probe of sequence 5'-d(TTTTTCC) was 5'-end-labeled with pyrene. In this construct, the proximal thymine residues efficiently quench the fluorophore emission in solution. Upon stoichiometric binding of one probe per polymerase molecule, the pyrene steady-state fluorescence increases by two orders of magnitude, the fluorescence anisotropy increases, and a long fluorescence lifetime component of 140 ns appears. With addition of replicable RNA, steady-state fluorescence decreases in a concentration dependent manner and the long lifetime component is lost. This observation most likely reflects displacement of the pyrene-labeled probe from the proposed nucleic acid binding site II of Qbeta replicase. The effect was utilized to access binding affinities of different RNAs to this site in a reverse titration assay format. In 10 mM sodium phosphate (pH 7.0), 100 mM NaCl, at 16 degrees C, equilibrium dissociation constants for different template midi- and minivariant RNAs were calculated to be in the nanomolar range. In general, the minus and plus strands, concomitantly synthesized by Qbeta replicase during replication, exhibited discriminative affinities, while their hybrid bound less efficiently than either of the single strands. Different non-replicable tRNAs also bound to the polymerase with comparable dissociation constants. By titration with DNA homo-oligonucleotides it was shown that the probed site on Qbeta replicase does not require a 2' hydroxyl group for binding nucleic acids, but recognizes pyrimidine residues. Its interaction with thymine is lost in an A.T base-pair, while that with cytosine is retained after Watson-Crick base-pairing. These findings can explain the affinities of RNA-Qbeta replicase interactions reported here and in earlier investigations. The sensitivity of the described fluorometric assay allows detection of RNA amplification by Qbeta replicase in real-time.
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PMID:Probing RNA-protein interactions using pyrene-labeled oligodeoxynucleotides: Qbeta replicase efficiently binds small RNAs by recognizing pyrimidine residues. 935 49

Molecular analysis of mutations at the hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus in peripheral blood T-lymphocytes can provide information on mechanisms of somatic in vivo mutation in populations exposed to exogenous carcinogens and in individuals with inherent susceptibility to cancer and other diseases. To study possible mutational changes associated with smoking as a risk factor for lung cancer, we analyzed HPRT mutations in T-cells of newly diagnosed, nonsmoking and smoking lung cancer patients before treatment. Reverse transcriptase polymerase chain reaction (RT-PCR) and DNA sequencing methods were used to identify 146 independent mutations, 73 each from 32 nonsmoking and 31 smoking cases. In 35 T-cell mutants, the HPRT cDNA showed loss of an entire exon, indicating a splicing mutation. Among the remaining 111 fully characterized mutations in the coding region, single base pair (bp) substitutions predominated with 79% (48/61) in nonsmokers and 90% (45/50) in smokers. Frameshift and small deletion (1-24 bp) mutations were found in 18 mutants. The distribution of base pair substitutions was nonrandom, with significant clustering at previously identified hotspot positions 143, 197 and 617 in the HPRT coding sequence (P< or =0.008). One additional hotspot, GC-->TA at position 606, was observed only in smokers (P=0.006). The frequency of GC>TA transversions was higher in smokers (13%) than in nonsmokers (6%). Conversely, smokers had a lower frequency of GC>AT transitions (24%) than nonsmokers (35%). This smoking-associated shift of the HPRT mutational spectrum, although not statistically significant, is consistent with the in vitro mutagenicity of benzo(a)pyrene (BaP), a prominent carcinogen of tobacco smoke, and with known differences in the TP53 mutational spectrum in lung tumors of smokers and nonsmokers. Among nonsmokers, the HPRT mutational spectra in healthy population controls and lung cancer patients were similar, but there was a marginally significant difference (P=0.07) in the distribution of base pair substitutions between smoking controls and patients. These results suggest that (i) general mechanisms of somatic mutagenesis in individuals with possible predisposition to cancer (e.g. nonsmoking lung cancer patients) are not different from those in normal healthy individuals, and (ii) the HPRT gene in T-cells is a useful reporter locus for smoking-associated somatic in vivo mutations occurring early in lung cancer development.
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PMID:Mutational spectra at the hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus in T-lymphocytes of nonsmoking and smoking lung cancer patients. 1086 57

Metallothioneins (MTs) are a family of low molecular weight, cysteine-rich, inducible, intracellular proteins that bind heavy metals with high affinity. MT-1 is known as a stress-inducible protein and functions as an antioxidant enzyme. Areca quid chewing is a major risk factor in the development and further progression of oral squamous cell carcinoma (OSCC). The aim of this study was to compare MT-1 expression in normal human oral epithelium and OSCC and further explore the potential mechanism that may lead to induce MT-1 expression. Thirty four OSCC and 10 normal epithelium specimens were examined by immunohistochemistry and analyzed by the clinico-pathological profiles. The oral epithelial cell line GMN cells were challenged with arecoline, a major areca nut alkaloid, by reverse-transcriptase polymerase chain reaction. Furthermore, tobacco smoke carcinogen benzo[a]pyrene (BaP) and glutathione (GSH) precursor N-acetyl-l-cysteine (NAC) were added to find the possible regulatory mechanisms. The results from immunohistochemistry demonstrated that MT-1 expression was significantly higher in OSCC specimens (p<0.05). No significant difference in MT-1 expression was observed with respect to age, sex, T category, and stage (p>0.05). The high MT-1 expression was associated with lymph node metastasis (p=0.012). In addition, arecoline was found to elevate MT-1 mRNA in a dose-dependent manner (p<0.05). Furthermore, the addition of BaP enhanced the arecoline-induced MT-1 expression (p<0.05). The addition of NAC markedly inhibited the arecoline-induced MT-1 expression (p<0.05). These results lead to the conclusion that MT-1 expression is significantly upregulated in areca quid chewing associated-OSCC. The expression profile suggests MT-1 could be used clinically as a marker for tumors possessing the potential for lymph node metastasis. The compounds of tobacco products may act synergistically in the pathogenesis of OSCC in areca quid chewers. The regulation of MT-1 expression induced by arecoline is critically dependent on the intracellular GSH concentration.
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PMID:The upregulation of metallothionein-1 expression in areca quid chewing-associated oral squamous cell carcinomas. 1741 20