Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.4.2.8 (hypoxanthine-guanine phosphoribosyltransferase)
 2,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The induction of DNA single-strand breaks in C3H10T1/2 mouse fibroblasts and Chinese hamster ovary (CHO) cells by N-nitroso-N-2-fluorenylacetamide (N-NO-2-FAA) was demonstrated by the alkaline elution technique. Without metabolic activating system (i.e., rat liver S9 fraction), N-NO-2-FAA exhibits more direct and strong damaging effects on DNA than its parent compound, 2-FAA, at equal concentration in both cell lines. To compare the DNA-damaging potency of N-NO-2-FAA with other well-known carcinogens, such as benzo[a]pyrene, 2-nitrofluorene, and N-methyl-N'-nitrosoguanidine (MNNG), the order of potency is as follows: MNNG (5 microM) greater than N-NO-2-FAA (150 microM) greater than benzo[a]pyrene (20 microM) at equitoxic concentrations, LD37, in the same cell system. Another parallel experiment indicated that N-NO-2-FAA could disrupt the superhelicity of circular plasmid DNA (pBR 322) at a dose range of 0.1-50 mM; however, a complete conversion to form III linear DNA was found at the highest concentration (50 mM). After treatment with various concentrations of N-NO-2-FAA, ouabain resistance (ouar) was induced in C3H10T1/2 cells, while both ouar and 6-thioguanine resistance (6-TGr) were induced in CHO cells. The mutation frequency in the Na+/K+-ATPase locus in CHO cells (1.5 X 10(-6) mutants/microM) is higher than that in C3H10T1/2 cells (1.0 X 10(-6) mutants/microM). The maximal mutation frequency at the Na+/K+-ATPase gene locus was attained with 30 min of exposure in C3H10T1/2 cells, whereas the mutation frequency in CHO cells continued to increase up to 80 min of treatment. Similarly, the maximal mutation frequency at the HPRT locus also continued to increase up to 80 min of treatment. Finally, a linear plot of alkali-labile lesions versus 6-TGr mutations was obtained; but the same relationship was not observed in the case of ouar mutation.
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PMID:The relationship between DNA damage and mutation frequency in mammalian cell lines treated with N-nitroso-N-2-fluorenylacetamide. 273 14

The usefulness of the 32P-post-labeling/t.l.c. method for quantitative DNA adduct dosimetry was evaluated. 2-Acetylaminofluorene (2-AAF)-DNA adducts from three systems were characterized qualitatively and quantitatively by the 3H-radiolabeled technique with subsequent analysis by h.p.l.c. (pre-labeling method) and by the 32P-post-labeling method. Both methods showed N-acetoxyacetylaminofluorene (N-OAc-AAF) reaction products with calf thymus DNA were predominantly N-(deoxyguanosin-8-yl)-2-acetylaminofluorene (dG-C8-AAF) with some N-(deoxyguanosin-8-yl)-2-aminofluorene (dG-C8-AF) and N-(deoxyguanosin-N2-yl)-2-acetylaminofluorene (dG-N2-AAF). In contrast, Chinese hamster ovary (CHO) cells treated with [3H]N-OAc-AAF gave 80 or 90% dG-C8-AF adducts and 20 or 10% dG-C8-AAF adducts with the post- or pre-labeling method, respectively. Likewise in CHO cells treated with 2-AAF in the presence of rat liver homogenate, approximately 90% dG-C8-AF and 10% dG-C8-AAF adducts were detected using the 32P-post-labeling method. In Salmonella typhimurium strain TA1538 treated with 2-AAF or [3H]2-AAF in the presence of a rat liver homogenate, one adduct, dG-C8-AF, was identified. Similar quantitative results were also obtained with the two methods. However, the 32P-post-labeling method was more sensitive and also eliminated the use of radiolabeled-mutagen treatments. Quantitative DNA adduct dosimetry was applied to AAF-induced mutagenesis in the S. typhimurium and CHO/HPRT mutation assays. A linear and reproducible relationship existed between dG-C8-AF levels and AAF-induced mutants in both systems.
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PMID:The 32P-post-labeling method in quantitative DNA adduct dosimetry of 2-acetylaminofluorene-induced mutagenicity in Chinese hamster ovary cells and Salmonella typhimurium TA1538. 354 38

The mutagenic specificities of ethylnitrosourea (ENU), X-rays (+/-)7 beta,8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7, 8,9,10-tetrahydrobenzo[a]pyrene (BPDE), ICR-191, and N-acetoxy-2-acetylaminofluorene (N-AcO-AAF) were analyzed and compared in diploid human fibroblasts and Salmonella typhimurium. In the human fibroblasts, we compared the frequency of diphtheria toxin (DT)-resistant mutants, presumably induced in the gene coding for elongation factor-2, with the frequency of 6-thioguanine (TG) resistance induced by mutations in the gene coding for hypoxanthine(guanine)phosphoribosyltransferase (HPRT). Recovery of DT-resistant (DTr) cells requires that the mutant EF-2 retain the ability to carry on protein synthesis since the normal EF-2 will be inactivated by DT selection. Therefore, the DTr mutation cannot involve major changes in the gene. In contrast, cells can acquire TG resistance by any mechanism which eliminates HPRT activity, e.g., base substitution, frameshift, deletion, loss of chromosomes. Each agent was assessed by calculating the ratio of the slopes of the dose-response plots (induced variant frequency as a function of dose of the agent used) for the two markers (DTr/TGr variants.). In S. typhimurium we examined the reversion frequency in four histidine-requiring strains bearing forward mutations of the frameshift (TA1538, TA98) or missense (TA1535, TA100) type. ENU, which was predominantly a base substitution mutagen in the bacteria, gave a ratio of DTr to TGr variants of 1.5. As expected of an agent inducing gross chromosomal changes, X-rays induced no revertants in bacteria and in human cells gave a ratio of 0.1. ICR-191 which was predominantly a frameshift mutagen in bacteria gave a ratio of 0.15. In the set of bacterial strains containing the plasmid pKM101, BPDE reverted both frameshift and base substitution mutations. It did not cause reversions in the other set of strains. In human cells BPDE gave a response similar to ENU, i.e., a ratio of DTr/TGr variants of 1.5. As reported by others, N-AcO-AAF was predominantly a frameshift mutagen in bacteria. However, in the human cells it gave a ratio of DTr/TGr variants of 1.5, similar to ENU and BPDE. These results suggest that in human cells, BPDE and N-AcO-AAF, like ENU, yield predominantly base substitutions, while ICR-191 and X-rays largely produce mutations by mechanisms which result in more extensive alterations in the gene.
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PMID:Comparison of the frequency of diphtheria toxin and thioguanine resistance induced by a series of carcinogens to analyze their mutational specificities in diploid human fibroblasts. 636 45

Isolated rat and human hepatocytes in primary culture were shown to metabolize AAF to reactive intermediates which damaged hepatocyte DNA. A significant increase in unscheduled DNA synthesis was detectable by autoradiography in rat and human hepatocytes exposed to concentrations of AAF as low as 1 microM. When rat hepatocytes were plated over confluent monolayers of human fibroblasts and exposed to 3H-AAF, significant binding of AAF to the DNA of the fibroblasts as well as the hepatocytes was measured. In other experiments with hepatocyte-fibroblast cocultures, nonradioactive AAF, at concentrations greater than 40 microM, induced a significant increase in the HPRT- mutation frequency in the human fibroblasts. These results demonstrate that hepatocytes can be used to assess genotoxicity of carcinogenic compounds and are useful for interspecies comparisons in chemical carcinogenesis.
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PMID:Genotoxic effects of 2-acetylaminofluorene on rat and human hepatocytes. 683 90

The model that transcription-coupled excision repair reflects the interference of DNA damage with the transcription process predicts that the rate of such excision repair will be related to the degree to which a particular type of lesion blocks transcription. We tested this by measuring the rate of excision repair of guanine adducts formed in the HPRT gene of diploid human fibroblasts and in the overall genome by two structurally related polycyclic carcinogens, 1-nitrosopyrene (1-NOP) and N-acetoxy-2-acetylaminofluorene (N-AcO-AAF) and comparing the results with those we found previously using benzo[a]pyrene diol epoxide (BPDE). We also measured the degree of interference with in vitro transcription by these adducts. Our results showed that, although BPDE adducts are four times more effective than 1-NOP adducts in blocking transcription, the preferential and strand-specific repair of 1-NOP adducts was twice as fast as that of BPDE adducts. Excision repair of N-AcO-AAF adducts was significantly slower than that of BPDE adducts and was not strand-specific. The efficiency of blocking of transcription by deacetylated N-AcO-AAF adducts was similar to 1-NOP adducts. Therefore, the extent to which a particular lesion blocks transcription in vitro does not predict its rate of preferential or transcription-coupled excision repair.
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PMID:Lack of correlation between degree of interference with transcription and rate of strand specific repair in the HPRT gene of diploid human fibroblasts. 759 80

The cytotoxic and mutagenic effect of 1-nitrosopyrene (1-NOP) and N-acetoxy-2-acetylaminofluorene (N-AcO-AAF) were compared with that of (+/-)-7 beta, 8 alpha-dihydroxy-9 alpha, 10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) as a function of the initial frequency of adducts formed in the DNA of repair-proficient diploid human fibroblasts and the fraction remaining at the time the cells replicate their DNA. The principal adducts of all three agents involve guanine. The initial level of BPDE-, 1-NOP-, or N-AcO-AAF-induced adducts per 10(6) nucleotides required to lower the survival of these cells to 37% of the control was 8, 25, and 50, respectively. The frequency of mutants per 10(6) clonable cells induced at those levels of initial adduct formation was 160, 80, and 40, respectively. We determined the rate of excision repair of these adducts from the overall genome, from the individual strands of the hypoxanthine phosphoribosyltransferase (HPRT) gene, and in the case of 1-NOP and BPDE, at the level of individual nucleotides in the nontranscribed strand of exon 3 of that gene, a region where mutations induced by those agents are particularly frequent. 1-NOP-induced adducts were excised from the overall genome and from the individual strands of HPRT at a rate 2-3 times faster than BPDE-induced adducts. The average rate of repair of 1-NOP-induced adducts in exon 3 was also 2-3 times faster than the average rate of repair of BPDE-induced adducts. However, at particular nucleotides 1-NOP-induced adducts were repaired much faster, or slower, or in some cases at a rate equal to that of BPDE-induced adducts. Excision repair of N-AcO-AAF-induced adducts (i.e., deacetylated aminofluorene residues) was significantly slower than that of BPDE- or 1-NOP-induced adducts, and was not strand-specific. In an in vitro assay, BPDE adducts were four times more effective in blocking transcription than were 1-NOP or N-AcO-AAF-induced adducts. We conclude that the cytotoxic and mutagenic effect of these carcinogens reflect a complex interplay of adduct conformation, ability of adducts to block replication and transcription, and variation in the rate of excision repair, even at the nucleotide level.
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PMID:Relationship between adduct formation, rates of excision repair and the cytotoxic and mutagenic effects of structurally-related polycyclic aromatic carcinogens. 920 50

The lymphocyte hypoxanthine-guanine phosphoribosyltransferase (Hprt) assay is frequently used as a biomarker for the exposure of both humans and laboratory animals to potentially carcinogenic agents. To obtain information concerning the sensitivity of the rat Hprt lymphocyte assay toward aromatic amine carcinogens, male F344 rats were fed 0.02% 2-acetylaminofluorene (2-AAF) for 1 month and then returned to control diet for 2 months. At 4, 27, 48, 62, and 90 days after the initiation of 2-AAF-feeding, the frequency of mutants in the Hprt gene was determined. In addition, DNA was isolated from liver nuclei, spleen lymphocytes, bone marrow, and thymus, and DNA adducts were analyzed by 32P-postlabeling. 2-AAF feeding resulted in a significant induction of 6-thioguanine-resistant T-lymphocytes and the mutant frequency continued to increase after the 2-AAF feeding was stopped. The same major DNA adduct, N-(deoxyguanosin-8-yl)-2-aminofluorene, was detected in liver, spleen lymphocytes, bone marrow, and thymus. DNA adduct levels were greatest in the tumor target tissue (liver) but occurred in all T-lymphocyte compartments, being highest in spleen lymphocytes. The DNA adduct levels were highest at the end of the 1-month 2-AAF feeding period and decreased rapidly in all tissues. The data indicate that the Hprt lymphocyte mutagenesis assay detects arylamine carcinogens, but with relatively low sensitivity.
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PMID:Hprt lymphocyte mutant frequency in relation to DNA adduct formation in rats fed the hepatocarcinogen 2-acetylaminofluorene. 1050 13