Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.4.2.8 (hypoxanthine-guanine phosphoribosyltransferase)
2,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chinese hamster cells were treated with ethyl methanesulfonate or N-methyl-N'-nitro-N-nitrosoguanidine, and mutants resistant to 8-azaguanine were selected and characterized. Hypoxanthine-guanine phosphoribosyltransferase activity of sixteen mutants is extremely negative, making them suitable for reversion to HGPRTase(+). Ten of the extremely negative mutants revert at a frequency higher than 10(-7) suggesting their point mutational character. The remaining mutants have demonstrable HGPRTase activity and are not useful for reversion analysis. Five of these mutants have < 2% HGPRTase and are presumably also HGPRTase point mutants. The remaining 14 mutants utilize exogenous hypoxanthine for nucleic acid synthesis poorly, and possess 20-150% of wild-type HGPRTase activity in in vitro. Their mechanism of 8-azaguanine resistance is not yet defined.
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PMID:8-Azaguanine resistance in mammalian cells. I. Hypoxanthine-guanine phosphoribosyltransferase. 434 96

The influence of inhibitors of poly(ADP-ribose) polymerase such as 3-aminobenzamide (3AB) and benzamide (B) on the spontaneously occurring as well as mutagen induced chromosomal aberrations, sister chromatid exchanges (SCEs) and point mutations has been studied. In addition, we have measured the influence of 3AB on DNA repair following treatment with physical and chemical mutagens. Post treatment of X-irradiated mammalian cells with 3AB increases the frequencies of induced chromosomal aberrations by a factor of 2 to 3. Both acentric fragments and exchanges increase indicating that the presence of 3AB slows down the repair of DNA strand breaks (probably DNA double strand breaks), thus making breaks available for interaction with each other to give rise to exchanges. 3AB, when present in the medium containing bromodeoxyuridine(BrdUrd) during two cell cycles, increases the frequencies of SCEs in Chinese hamster ovary cells (CHO) in a concentration dependent manner leading to about a 10-fold increase at 10 mM concentration. Most 3AB induced SCEs occur during the second cell cycle, in which DNA containing bromouridine (BU) is used as template for replication. BU containing DNA appears to be prone to errors during replication. The extent of increase in the frequencies of SCEs by 3AB is correlated with the amount of BU incorporated in the DNA of the cells. The frequencies of spontaneously occurring DNA single strand breaks in cells grown in BrdUrd containing medium are higher than in the cells grown in normal medium and this increase depends on the amount of BU incorporated in the DNA of these cells. We have studied the extent of increase in the frequencies of SCEs due to 1 mM 3AB in several human cell lines, including those derived from patients suffering from genetic diseases such as ataxia telangiectasia (A-T), Fanconi's anemia (FA), and Huntington's chorea. None of these syndromes showed any increased response when compared to normal cells. 3AB, however, increased the frequencies of spontaneously occurring chromosomal aberrations in A-T and FA cells. 3AB does not influence the frequencies of SCEs induced by UV or mitomycin C (MMC) in CHO cells. However, it increases the frequencies of SCEs induced by ethyl methanesulfonate (EMS) and methyl methanesulfonate (MMS). Under the conditions in which 3AB increases the frequencies of spontaneously occurring as well as induced SCEs, it does not increase the frequencies of point mutations in hypoxanthine-guanine phosphoribosyltransferase (HGPRT) locus. 3AB does not influence the amount of repair replication following dimethylsulphate (DMS) treatment of human fibroblasts, or UV irradiated human lymphocytes.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Influence of inhibitors of poly(ADP-ribose) polymerase on DNA repair, chromosomal alterations, and mutations. 631 38

Mutants of Chinese hamster ovary cells (CHO-K1 Pro-), resistant to the proline transport antagonist 2-(methylamino)-isobutyrate (MeAIB) were isolated by a single-step procedure. Mutation rates to Pro+ and to Pro- MeAIB resistance (MeAIBr) are 1.7 X 10(-6) and 2.4 X 10(-5), respectively. Several Pro- MeAIBr mutants were tested by measuring the uptake of 0.05 mM proline through the various amino acid transport systems: some showed increases in one transport system only; others revealed pleiotropic changes affecting two or more systems; still others had no apparent change in proline transport. One Pro- MeAIBr mutant analyzed in detail (MeAIBr22) was isolated after EMS treatment as resistant to 5 mM MeAIB, is Pro-, stable, and shows a 1.6-fold increase in the initial velocity of transport of 0.05 mM proline. There appears to be no change in the velocity of proline transport through the amino acid transport systems A, P, and L, and the "glutamine inhibitable fraction." In contrast, there is a 5.5-fold increase in the velocity of transport of 0.05 mM proline through the ASC system. Kinetic studies reveal a sixfold increase in the Vm and a slight increase in the Km of the transport of serine through the ASC system. Hybrids between MeAIBr22 and CHO-K1 Pro-, OUAr, HPRT- showed the parental phenotype. These results indicate that the mutant ASC phenotype of MeAIBr22 is recessive and is probably the result of a regulatory gene mutation.
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PMID:Recessive 2-(methylamino)-isobutyrate (MeAIB)-resistant mutant of Chinese hamster ovary cells (CHO-K1) with increased transport through ASC system. 642 46

As a first step in the development of a multiple-marker, mammalian cell mutagenesis assay system, we have isolated a Chinese hamster ovary (CHO) cell line that is heterozygous for both the adenine phosphoribosyltransferase (aprt) and thymidine kinase (tk) loci. Presumptive aprt+/- heterozygotes with intermediate levels of APRT activity were selected from unmutagenized CHO cell populations on the basis of resistance to low concentrations of the adenine analog, 8-azaadenine. a functional aprt+/ heterozygote with approximately 50% wild-type APRT activity was subsequently used to derive sublines that were also heterozygous for the tk locus. Biochemical and genetic characterization of one such subline, CHO-AT3-2, indicated that it was indeed heterozygous at both the aprt and tk loci. CHO-AT3-2 cells permitted single-step selection of mutants resistant for 8-azaadenine or 5-fluorodeoxyuridine, allowing quantitation and direct comparison of mutation induction at the autosomal aprt or tk loci, as well as in the gene involved in ouabain resistance or at the X-linked, hypoxanthine--guanine phosphoribosyltransferase (hgprt) locus. Significant dose-dependent increases in mutation frequency were observed for all 4 genetic markers after treatment of CHO-AT3-2 cells with ethyl methanesulfonate.
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PMID:Mutagenicity testing in mammalian cells. I. Derivation of a Chinese hamster ovary cell line heterozygous for the adenine phosphoribosyltransferase and thymidine kinase loci. 644 63

Conditions for reliable and efficient assay of mutations affecting the activity of HPRT (hypoxanthine phosphoribosyltransferase EC 2.4.2.8) and APRT (adenine phosphoribosyltransferase EC 2.4.2.7) have been determined for a strain of CHO (Chinese hamster ovary) cells that has been adapted for rapid growth both in suspension culture and in monolayer. To facilitate measurement of mutation at the aprt locus, clones were derived that are presumptively heterozygous at that locus. At a limiting concentration of 8 microgram/ml of azaadenine, 14/16 of the resistant clones picked and tested had approximately 1/2 of the APRT activity of the wild-type cells. One such clone, strain AA8, was chosen for further studies and found to be readily mutable to resistance to 80 microgram/ml azaadenine. Most of the highly resistant colonies isolated (21/24) had very low in vitro APRT activity. The optimal conditions for detection of TGr and AAr mutations were determined for two critical parameters, expression time and cell density. Cultures treated with mutagen either in monolayer or in suspension were allowed to express mutations in suspension. The expression of mutations induced by UV light, EMS, and ICR-191 was complete by 3 days for AAr and by 4-5 days for TGr. The time required to reach a maximal frequency of mutants was essentially independent of the type of mutagen and the level of survival after treatment. Induced mutation frequencies for both loci were notably stable during the time intervals examined. With respect to cell-density conditions, both markers were detected at frequencies that were independent of the cell inocula over the range of 1 x 10(5) to 1 x 10(6) cells per 100-mm petri dish (i.e. 1.6 x 10(3) to 1.6 x 10(4) cells/cm2) containing 20 ml of medium. These results were obtained with both mutagenized populations and with reconstructed mixtures obtained by adding drug-resistant cells to varying numbers of wild-type cells. The rapid expression of mutations for both markers, particularly AAr, combined with the advantage that large inocula can be plated for selection of mutants, make this CHO strain an attractive system for the simultaneous measurement of mutations at the autosomal aprt and X-linked hprt loci.
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PMID:Validation of conditions for efficient detection of HPRT and APRT mutations in suspension-cultured Chinese hamster ovary cells. 736 Jan 55

Cells from the human lymphoblastoid cell line, AHH-1, were exposed to two direct-acting mutagens, ethyl methanesulfonate (EMS) and ethyl nitrosourea (ENU), and to three carcinogens that require metabolic activation to an electrophile, benzo[a]pyrene (B(a)P), 6-aminochrysene (6-AC), and 6-nitrochrysene (6-NC); mutation induction at the HPRT locus was quantified by resistance to 6-thioguanine (6-TGr). Exposure of AHH-1 cells to either EMS or ENU resulted in a concentration-dependent increase in mutant frequency at the HPRT locus. When AHH-1 cells were exposed to B(a)P, the increase in mutant frequency at the HPRT locus was marginally significant linearly and significant quadratically. The 32P-postlabeling assay revealed the formation of DNA adducts derived from (+/-)anti-benzo[a]pyrene-trans-7,8-dihydrodiol-9,10-epoxide which may account for the increase in 6-TGr clones. Although DNA adducts could be detected by the 32P-postlabeling assay in both 6-NC- and 6-AC-treated AHH-1 cells, exposure to 6-AC or 6-NC did not result in a concentration-dependent increase in mutant frequency at the HPRT locus. Our results are consistent with the results of previous studies which indicate that EMS and ENU are effective inducers of 6-TGr clones as is B9(a)P when activated to an electrophile. In 6-NC- and 6-AC-exposed cells, low levels of N-hydroxy-6-aminochrysene-derived adducts were detected in only 6-NC-exposed cells. No 6-aminochrysene-1,2-dihydrodiol-derived adducts were detected following 6-NC or 6-AC exposure. Minimal metabolic activation of 6-NC or 6-AC by AHH-1 cells may account for the lack of a positive mutagenic response for either 6-AC or 6-NC.
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PMID:Induction of mutations at the hypoxanthine phosphoribosyl transferase (HPRT) locus in AHH-1 human lymphoblastoid cells. 752 83

Spontaneous and induced mutations at the HPRT locus were analyzed in one normal (MRC5CV1) and one ataxia telangiectasia (AT5BIVA) SV40-transformed cell line derived from male donors. Multiplex PCR and Southern analyses revealed a high frequency of spontaneous deletion mutations that may be a consequence of the SV40 transformation. Four mutagens (ethyl methanesulfonate, bromodeoxyuridine, bleomycin, adriamycin), which differ in their types of primary DNA lesions, caused specific patterns of mutations. By using fluorescence in situ hybridization (FISH) techniques, we were able to show that more than 90% of the AT5BIVA cells contained two X chromosomes with HPRT alleles, while in more than 90% of the MRC5CV1 cells genomic hemizygosity for the HPRT gene was found. Taking into account these findings we found that the AT5BIVA cell line possesses spontaneous hypermutability as well as hypersensitivity and hypermutability to bleomycin (BLM) and adriamycin (AM). Both mutagens induced deletion mutations in both cell lines, but more complex mutations and larger deletions were found in AT5BIVA cells.
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PMID:Characterization of spontaneous and induced mutations in SV40-transformed normal and ataxia telangiectasia cell lines. 753 43

The rapid developments in the molecular genetics of Toxoplasma gondii have far reaching implications in treatment and vaccination strategies for this as well as closely related pathogens such as Plasmodium. Although stable transformation of this parasite through homologous and illegitimate genomic integration has provided many of the tools necessary for genetic analysis, subsequent manipulations of the DNA have proven laborious. This report describes the selection and subsequent characterization of a Toxoplasma sequence that permits the episomal maintenance of bacterial plasmids in this parasite. This sequence was isolated from the Toxoplasma genome through selection for episomal stability of a pUC19-based library in the absence of a selectable marker. A 500-base pair fragment was determined to possess the stabilization activity. Transformations of Toxoplasma using vectors possessing this fragment, referred to as EMS (episomal maintenance sequence), demonstrated an elevated stable transformation frequency compared with the vector alone. Mutants deficient in hypoxanthine-xanthine-guanine phosphoribosyltransferase activity were used as a test to see if this gene could be selected from a genomic library using a vector containing the EMS. The success of this test demonstrates the utility of EMS-containing vectors in complementation strategies and the ability of such constructs bearing large fragments of the Toxoplasma genome to be maintained episomally.
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PMID:Development of a stable episomal shuttle vector for Toxoplasma gondii. 946 85

Centchroman (CC), a non-steroidal oral contraceptive and a candidate drug for breast cancer, has been reported to exhibit partial to complete remission of lesions in 40.5% of breast cancer patients. The potent anti-oestrogenic activity, negligible side-effects and anti-breast cancer activity of CC prompted us to evaluate the antimutagenic effects of this compound in a bacterial mutagenicity assay and CHO/HPRT and AS52/GPT mutation assays in vitro and in vivo in female Swiss albino mice as measured by both sister chromatid exchange (SCE) and chromosome aberrations (CA) against three known positive mutagen compounds, dimethylbenz[a]anthracene (DMBA), cyclophosphamide (CP) and mitomycin C (MMC). Antimutagenicity assays in Salmonella strains TA97a, TA100, TA98 and TA102 were carried out against commonly used known positive mutagens, sodium azide, 4-nitro-o-phenylenediamine, cumine hydroperoxide, 2-aminofluorene and danthron. A significantly reduced number of bacterial histidine revertant colonies was observed in the plates treated with 0.1, 1, 5 and 10 microg/plate CC and a positive compound when compared with bacterial plates treated with the respective positive compound alone. Ethyl methanesulfonate (EMS), a commonly used positive mutagen for CHO/HPRT and AS52/GPT gene mutation assays, was used for antimutagenicity assay in these cells. CC exhibited protective effects against the mutagenicity of EMS in these two mammalian cell mutation assays, CHO/HPRT and AS52/GPT. In the in vivo studies, pretreatment with CC reduced DMBA-induced SCE and CA and CP- and MMC-induced CA when compared with the group treated only with the positive compounds. These results indicate that CC can reduce the mutagenic effects of known genotoxic compounds.
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PMID:Antimutagenic effects of centchroman--a contraceptive and a candidate drug for breast cancer in multiple mutational assays. 1056 37

Folic acid deficiency acts synergistically with alkylating agents to increase genetic damage at the HPRT locus in Chinese hamster ovary cells in vitro and in rat splenocytes in vivo. The present studies extend these observations to human cells and, in addition, investigate the role of p53 activity on mutation induction. The human lymphoblastoid cell lines TK6 and WTK1 are derived from the same parental cell line (WI-L2), but WTK1 expresses mutant p53. Treatment of folate-replete or deficient WTK1 and TK6 cells with increasing concentrations (0-50microg/ml) of ethyl methanesulfonate (EMS) resulted in significantly different HPRT mutation dose-response relationships (P<0.01), indicating that folate deficiency increased the EMS-induced mutant frequency in both cell lines, but with a greater effect in TK6 cells. Molecular analyses of 152 mutations showed that the predominant mutation (65%) in both cell types grown in the presence or absence of folic acid was a G>A transition on the non-transcribed strand. These transitions were mainly at non-CpG sites, particularly when these bases were flanked 3' by a purine or on both sides by G:C base pairs. A smaller number of G>A transitions occurred on the transcribed strand (C>T=14%), resulting in 79% total G:C>A:T transitions. There were more genomic deletions in folate-deficient (15%) as compared to replete cells (4%) of both cell types. Mutations that altered RNA splicing were common in both cell types and under both folate conditions, representing 33% of the total mutations. These studies indicate that cells expressing p53 activity exhibit a higher rate of mutation induction but are more sensitive to the toxic effects of alkylating agents than those lacking p53 activity. Folate deficiency tends to reduce toxicity but increase mutation induction after EMS treatment. The p53 gene product did not have a major influence on the molecular spectrum after treatment with EMS, while folate deficiency increased the frequency of deletions in both cell types.
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PMID:The effect of folate deficiency on the cytotoxic and mutagenic responses to ethyl methanesulfonate in human lymphoblastoid cell lines that differ in p53 status. 1116 26


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