UNIPROT:P00492 - FACTA Search

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Query: UNIPROT:P00492 (hypoxanthine-guanine phosphoribosyltransferase)
2,385 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hyperbaric oxygen (HBO) treatment as used therapeutically (i.e., exposure to 100% oxygen at a pressure of 1.5 bar for a total of 60 min) has been shown to induce DNA damage in the alkaline comet assay with leukocytes from test subjects. Under these conditions, HBO did not lead to an induction of gene- and chromosome mutations. Due to known toxic effects, exposure of humans to HBO is limited and possible genetic consequences of HBO could not be completely evaluated in vivo. We thus established an in vitro HBO model, where human blood cells or V79 cells were exposed to hyperbaric oxygen (98% O(2) and 2% CO(2) at a pressure of either 1.5 or 3 bar) for up to 3 hr in a temperature-controlled hyperbaric chamber. Using the comet assay, we found exposure-related genotoxic effects in V79 cells, whole blood, and isolated lymphocytes. V79 cells showed the highest sensitivity toward HBO-induced DNA damage, and the exposure conditions applied to blood in vitro, to induce DNA migration, had to be higher than those used in vivo. We could also show that prolonged HBO treatment clearly increased the frequency of micronuclei in V79 cells, whereas it exerted only a marginal effect on the frequency of hprt mutations. These results demonstrate that HBO treatment of cell cultures is a well-suited model for investigating the biological significance of oxidative stress. The relationship between oxygen-induced DNA lesions and the formation of gene- and chromosome mutations is discussed.
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PMID:Evaluation of mutagenic effects of hyperbaric oxygen (HBO) in vitro. 1061 78

Mutatect MN-11 is a tumor line that can be grown subcutaneously in syngeneic C57BL/6 mice. The frequency of spontaneously arising mutants at the hypoxanthine phosphoribosyltransferase (Hprt) locus was observed to be elevated as a result of in vivo growth. The objective of the present study was to identify factors in the tumor microenvironment that might explain this increase in mutant frequency (MF). When tumors were examined histologically, neutrophils were found to be the predominant infiltrating cell type. Quantitative estimates of the number of neutrophils and MF of tumors in different animals revealed a statistically significant correlation (r = 0.63, P < 0.0001). Immunohistochemical analysis for inducible nitric oxide synthase (iNOS) demonstrated its presence, mainly in neutrophils. Biochemical analysis of tumor homogenates for nitric oxide synthase (NOS) activity indicated a statistically significant correlation with MF (r = 0.77, P < 0.0001). Nitrotyrosine was detected throughout the tumor immunohistochemically; both cytoplasmic and nuclear staining was seen. To increase the number of infiltrating neutrophils, tumors were injected with chemoattractant interleukin-8 and prostaglandin E2. This produced a statistically significant increase in neutrophil content (P = 0.005) and MF (P = 0.0002). As in control MN-11 tumors, neutrophil content and MF were strongly correlated (r = 0.63, P = 0. 003). Because neutrophils are a potential source of genotoxic reactive oxygen and/or nitrogen species, our results support the notion that these tumor-infiltrating cells may be mutagenic and contribute to the burden of genetic abnormalities associated with tumor progression.
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PMID:Neutrophils, nitric oxide synthase, and mutations in the mutatect murine tumor model. 1066 80

We recently showed that treatment of V79 cells with hyperbaric oxygen (HBO) efficiently induced DNA effects in the comet assay and chromosomal damage in the micronucleus test (MNT), but did not lead to gene mutations at the hprt locus. Using the comet assay in conjunction with bacterial formamidopyrimidine DNA glycosylase (FPG protein), we now provide indirect evidence that the same treatment leads to the induction of 8-oxoguanine, a premutagenic oxidative DNA base modification in V79 and mouse lymphoma (L5178Y) cells. We also demonstrate that HBO efficiently induces mutations in the mouse lymphoma assay (MLA). Exposure of L5178Y cells to HBO (98% O(2); 3bar) for 2h caused a clear mutagenic effect in the MLA, which was further enhanced after a 3h exposure. As this mutagenic effect was solely due to the strong increase of small colony (SC) mutants, we suggest that HBO causes mutations by induction of chromosomal alterations. Molecular characterization of induced SC mutants by loss of heterozygosity (LOH) analysis showed an extensive loss of functional tk sequences similar to the pattern found in spontaneous SC mutants. This finding confirmed that the majority of HBO-induced mutants is actually produced by a clastogenic mechanism. The induction of point mutations as a consequence of induced oxidative DNA base damage seems to be of minor importance.
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PMID:Evaluation of mutagenic effects of hyperbaric oxygen (HBO) in vitro. II. Induction of oxidative DNA damage and mutations in the mouse lymphoma assay. 1108 Jun 64

Neutrophils represent a potential source of genotoxic reactive oxygen and nitrogen species in the tumor microenvironment. Using Mutatect cell lines, which can form subcutaneous tumors in syngeneic C57BL/6 mice, we have previously established that the number of spontaneously infiltrating neutrophils correlates with the number of mutations at the hypoxanthine phosphoribosyltransferase (Hprt) locus. We now describe the properties of four lines that express different levels of the neutrophil chemokine, interleukin-8 (IL-8), from a tetracycline (TET)-responsive promoter. In a series involving 45 animals, IL-8-expressing lines produced tumors with a higher neutrophil content than the control line. Analysis of the 45 tumors revealed that the neutrophil level again strongly correlated with hprt mutant frequency (MF) (P<.0001, r=0.88). Administration of TET was effective in lowering the neutrophil content of low IL-8-expressing tumors, but not high IL-8-expressing tumors. Although the IL-8 transgene was stable in all lines in vitro, high IL-8-expressing lines completely lost the transgene in vivo whereas low IL-8-expressing lines showed no evidence of transgene instability. These results provide further evidence, based on the study of an endogenous gene (hprt) and an IL-8 transgene, that neutrophils may contribute to genetic instability in tumors.
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PMID:Expression of interleukin-8 promotes neutrophil infiltration and genetic instability in mutatect tumors. 1122 49

Oxidation of the methyl group of thymine yields 5-(hydroxymethyl)uracil (5-hmU) and 5-formyluracil (5-foU) as major products. Whereas 5-hmU appears to have normal base pairing properties, the biological effects of 5-foU are rather poorly characterised. Here, we show that the colony forming ability of Chinese hamster fibroblast (CHF) cells is greatly reduced by addition of 5-foU, 5-formyluridine (5-foUrd) and 5-formyl-2'-deoxyuridine (5-fodUrd) to the growth medium. There are no toxic effects of 5-fodUrd on cells defective in thymidine kinase or thymidylate synthetase, suggesting that the toxicity may be caused by 5-fodUrd phosphorylation and subsequent inhibition of thymidylate synthetase. Whereas 5-fodUrd was the most effective 5-foU derivative causing cell growth inhibition, the corresponding ribonucleoside 5-foUrd was more effective in inhibiting [3H]uridine incorporation in non-dividing rat nerve cells in culture, suggesting that 5-foUrd exerts its toxicity through interference with RNA rather than DNA synthesis. Addition of 5-foU and 5-fodUrd was also found to promote mutagenicity at the hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus of CHF cells; 5-fodUrd being three orders of magnitude more potent than 5-foU. In contrast, neither 5-hmU nor 5-(hydroxymethyl)-2'-deoxyuridine induced HPRT mutations. The mutation induction indicates that 5-foU will be incorporated into DNA and has base pairing properties different from that of thymine. These results suggest that 5-foU residues, originating from incorporation of oxidised bases, nucleosides or nucleotides or by oxidation of DNA, may contribute significantly to the damaging effects of oxygen radical species in mammalian cells.
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PMID:5-Formyluracil and its nucleoside derivatives confer toxicity and mutagenicity to mammalian cells by interfering with normal RNA and DNA metabolism. 1127 23

During autoxidation of the pentachlorophenol (PCP) metabolite tetrachlorohydroquinone (TCHQ) the semiquinone is formed as well as reactive oxygen species (ROS). It was examined if *OH or the semiquinone are the cause of TCHQ-induced genotoxicity by direct comparison of TCHQ- and H(2)O(2)-induced DNA damage in human cells. All endpoints tested (DNA damage, DNA repair, and mutagenicity) revealed a greater genotoxic potential for TCHQ than for H(2)O(2). In the comet assay, TCHQ induced DNA damage at lower concentrations than H(2)O(2). The damaging rate by TCHQ (tail moment (tm)/concentration) was 10-fold greater than by H(2)O(2). DNA repair was lower for TCHQ than for H(2)O(2) treatment. This was shown by measuring DNA repair in the unscheduled DNA synthesis (UDS) assay and the persistence of the DNA damage in the comet assay. In contrast to H(2)O(2), TCHQ in non-toxic concentrations was mutagenic in the hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus of V79 cells. Finally, there were also differences observed in cytotoxicity (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay) of TCHQ and H(2)O(2). Whereas the TCHQ cytotoxicity was enhanced during a 21h recovery phase, the H(2)O(2) cytotoxicity did not change. The results demonstrated that the pronounced genotoxic properties of TCHQ in human cells were not caused by *OH radicals but more likely by the tetrachlorosemiquinone (TCSQ) radical.
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PMID:Differences in genotoxicity of H(2)O(2) and tetrachlorohydroquinone in human fibroblasts. 1171 1

Rheumatoid arthritis (RA) is an inflammatory disease in which high levels of reactive nitrogen oxygen species (RNOS) may be present in the affected joints. RNOS are known to produce small-scale mutational events (transitions, transversions, small insertions, and small deletions) but the ability of these compounds to cause deletion of large segments of genomic DNA has not been previously determined. To address this question, a human lymphoblastoid cell line (WIL2-NS) was exposed to nitric oxide (NO)-donating drugs and hypoxanthine phosphoribosyltransferase (hprt)-negative clones were selected and analyzed by multiplex-PCR. Large-scale deletions accounted for 60-80% of hprt mutations arising in drug-treated cultures compared to 12% in untreated cultures (P-values of 0.006 and 0.0001, respectively, in two experiments). Deletion mutations in untreated cultures affected exon 9, whereas 75% of drug-induced deletion mutations affected exons 2, 3, and 9, and the remainder were very large, ranging from 26 to 1200 kbp. To compare this spectrum of NO-induced mutations in a lymphoblastoid line to that arising in vivo in arthritis patients, T-cells from RA patients, osteoarthritis (OA) patients, and controls were cloned and similarly analyzed. We previously showed that the overall frequency of Hprt mutant clones from patients is appreciably elevated compared to that of control subjects. Large-scale hprt deletions (0.5 to >26 kb) were detected in mutant T-cell clones from both RA and OA patients and also from control subjects. A total of 54 mutant clones from 16 RA patients and 19 mutant clones from 6 OA patients were studied. Of these, 6 clones (from 3 RA and 1 OA patient) had suffered large-scale deletions. A total of 9 control subjects were studied and 62 mutant clones were obtained. Of these, 19 had suffered large-scale deletions, arising in 7 of 9 control subjects. In conclusion, (1) RNOS are capable of inducing large-scale deletion mutations in a human lymphoblastoid cell line and (2) large-scale deletion mutations were found in 10-30% of T-cell clones from RA and OA patients and controls, which we hypothesize may be induced by RNOS.
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PMID:Nitric oxide donors induce large-scale deletion mutations in human lymphoblastoid cells: implications for mutations in T-lymphocytes from arthritis patients. 1177 57

Crystal structures have been determined for free Escherichia coli hypoxanthine phosphoribosyltransferase (HPRT) (2.9 A resolution) and for the enzyme in complex with the reaction products, inosine 5'-monophosphate (IMP) and guanosine 5'-monophosphate (GMP) (2.8 A resolution). Of the known 6-oxopurine phosphoribosyltransferase (PRTase) structures, E. coli HPRT is most similar in structure to that of Tritrichomonas foetus HGXPRT, with a rmsd for 150 Calpha atoms of 1.0 A. Comparison of the free and product bound structures shows that the side chain of Phe156 and the polypeptide backbone in this vicinity move to bind IMP or GMP. A nonproline cis peptide bond, also found in some other 6-oxopurine PRTases, is observed between Leu46 and Arg47 in both the free and complexed structures. For catalysis to occur, the 6-oxopurine PRTases have a requirement for divalent metal ion, usually Mg(2+) in vivo. In the free structure, a Mg(2+) is coordinated to the side chains of Glu103 and Asp104. This interaction may be important for stabilization of the enzyme before catalysis. E. coli HPRT is unique among the known 6-oxopurine PRTases in that it exhibits a marked preference for hypoxanthine as substrate over both xanthine and guanine. The structures suggest that its substrate specificity is due to the modes of binding of the bases. In E. coli HPRT, the carbonyl oxygen of Asp163 would likely form a hydrogen bond with the 2-exocyclic nitrogen of guanine (in the HPRT-guanine-PRib-PP-Mg(2+) complex). However, hypoxanthine does not have a 2-exocyclic atom and the HPRT-IMP structure suggests that hypoxanthine is likely to occupy a different position in the purine-binding pocket.
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PMID:Crystal structures of free, IMP-, and GMP-bound Escherichia coli hypoxanthine phosphoribosyltransferase. 1207 Mar 15

Methamphetamine (METH) is a widely abused psychomotor stimulant. Although numerous studies have examined METH-induced neurotoxicity, its ability to produce genotoxic effects has not been evaluated. In this article, we report on the genotoxicity of METH in vitro and in human METH abusers. METH induced his(+) revertants in Salmonella typhimurium strains TA98 and TA100, and increased the frequency of hprt mutants, micronuclei, and sister chromatid exchange (SCE) in cultured Chinese hamster ovary K1 (CHO-K1) cells. These METH-induced genotoxic effects were eliminated if METH exposure was conducted in the presence of rat liver S9, indicating that the genotoxicity was caused by METH, and not by metabolites of METH. In addition, reactive oxygen species (ROS) scavengers inhibited the METH-induced micronuclei in CHO-K1 cells. Further investigation with 76 human long-term METH abusers and 98 unexposed controls demonstrated that total METH exposure correlated with micronucleus and SCE frequencies in cultured lymphocytes. The results of this study indicate that METH is a genotoxic agent and that ROS may play a role in METH-induced genotoxicity.
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PMID:Genetic toxicity of methamphetamine in vitro and in human abusers. 1467 68

Aspirin has been proposed as a possible chemopreventive agent. On the other hand, a recent cohort study showed that aspirin may increase the risk for pancreatic cancer. To clarify whether aspirin is potentially carcinogenic, we investigated the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), which is correlated with the incidence of cancer, in cultured cells treated with 2,3-dihydroxybenzoic acid (2,3-DHBA), a metabolite of aspirin. 2,3-DHBA induced 8-oxodG formation in the PANC-1 human pancreatic cancer cell line. 2,3-DHBA-induced DNA single-strand breaks were also revealed by comet assay using PANC-1 cells. Flow cytometric analyses showed that 2,3-DHBA increased the levels of intracellular reactive oxygen species (ROS) in PANC-1 cells. The 8-oxodG formation and ROS generation were also observed in the HL-60 leukemia cell line, but not in the hydrogen peroxide (H(2)O(2))-resistant clone HP100 cells, suggesting the involvement of H(2)O(2). In addition, an hprt mutation assay supported the mutagenicity of 2,3-DHBA. We investigated the mechanism underlying the 2,3-DHBA-induced DNA damage using (32)P-labeled DNA fragments of human tumor suppressor genes. 2,3-DHBA induced DNA damage in the presence of Cu(II) and NADH. DNA damage induced by 2,3-DHBA was enhanced by the addition of histone peptide-6 [AKRHRK]. Interestingly, 2,3-DHBA and histone peptide-6 caused base damage in the 5'-ACG-3' and 5'-CCG-3' sequences, hotspots of the p53 gene. Bathocuproine, a Cu(I) chelator, and catalase inhibited the DNA damage. Typical hydroxyl radical scavengers did not inhibit the DNA damage. These results suggest that ROS derived from the reaction of H(2)O(2) with Cu(I) participate in the DNA damage. In conclusion, 2,3-DHBA induces oxidative DNA damage and mutations, which may result in carcinogenesis.
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PMID:Damage to cellular and isolated DNA induced by a metabolite of aspirin. 1910 73

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