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)

We have analysed the recovery of individual CHO-derived mutants during the generations immediately following their induction. This characteristic, which we call persistence, was measured by propagating mutagenized cultures in non-selective medium after subdivision into many very small populations, each containing either zero or one mutant. The recovery of most hypoxanthine phosphoribosyltransferase (hprt)-deficient mutants induced by ethyl methanesulphonate was low, and we have previously shown that this was usually due to an apparent rapid loss of the mutant phenotype with continued culture in non-selective medium (Bradley, 1980). A minority of about 15% manifest high persistence. We now show that most adenine phosphoribosyltransferase (aprt)-deficient mutants and some ouabain-resistant mutants had low persistence. Mutants induced by UV irradiation also generally exhibited low persistence but those induced by X-irradiation had significantly higher persistence than what was seen among EMS-induced mutants. Among various sublines of CHO cells which were tested for persistence of induced mutants, only one group consistently yielded mutants of high persistence. These were lines which carried glucose-6-phosphate dehydrogenase mutations which themselves had been originally induced by EMS.
 ...
PMID:Low persistence of the induced mutant phenotype in Chinese hamster cells. 253 33

Recent reports by several laboratories indicate that not all non-essential target loci are equally capable of detecting chromosomal mutations. The present study was undertaken to determine if both the tk locus in mouse lymphoma cells and the hgprt locus in Chinese hamster ovary (CHO) cells can be used to quantitate chromosomal mutations. Seven known mutagens for the tk locus were selected. These compounds were evaluated in the mouse lymphoma assay and in a suspension adapted CHO assay for their mutagenicity. In addition to the specific locus mutagenesis analysis, mouse lymphoma and CHO cells were evaluated for the frequency of gross chromosome aberrations. From these investigations, it appears that only those compounds [2-methoxy-6-chloro-9-(3-[ethyl-2-chloroethyl] aminopropylamino)-acridine-dihydrochloride (ICR 170), ethyl methanesulfonate (EMS) and methyl methanesulfonate (MMS)] that induce significant numbers of large-colony thymidine kinase (TK) mutants also induce significant numbers of hypoxanthine-guanine phosphoribosyltransferase (HGPRT) mutants. The four acrylates evaluated (methyl acrylate, ethyl acrylate, trimethylolpropane triacrylate and tetraethyleneglycol diacrylate) induced almost exclusively small-colony TK mutants and very few if any HGPRT mutants. Aberration analysis revealed that both the mouse lymphoma and CHO cells responded to the clastogenicity of the compounds (except for ICR 170 which was not positive in CHO cells) and that neither cell line was clearly more sensitive than the other to the clastogens tested. It is significant that the four acrylates give little or no evidence of genotoxicity when evaluated using selection for HGPRT-deficient mutants, yet are clearly clastogenic to the same cells in the same experiment. These results are consistent with the hypothesis that the hgprt locus may not be useful as a marker to evaluate the clastogenic component of a genotoxic compound. The present study adds to the increasing number of studies that support the view that the hemizygous nature of the hgprt locus permits the recovery of mutations primarily affecting the function of a single gene; whereas the heterozygous nature of the tk locus permits the recovery of both single gene and chromosomal mutations.
 ...
PMID:Differential mutant quantitation at the mouse lymphoma tk and CHO hgprt loci. 268 35

In LLC-PK1 cells, a cyclic AMP (cAMP)-elevating peptide hormone, calcitonin, induces urokinase-type plasminogen activator (uPA) gene transcription without concomitant protein synthesis. To understand the molecular mechanism of the uPA gene regulation by cAMP, we developed a system which allows us to obtain mutant cells with modified regulatory proteins. A uPA-gpt hybrid gene was constructed, in which the regulatory region of the uPA gene was linked to a bacterial xanthine-guanine phosphoribosyltransferase gene (gpt), and it was transfected into LLC-PK1 cells. A stably transformed cell line, which expressed gpt only in the presence of calcitonin, was obtained, and then these cells were treated with a chemical mutagen, ethyl methanesulfonate. Cells were screened for constitutive gpt expression and, as mutations in regulatory proteins should affect the two genes at the same time, cells were further screened for an increased basal uPA mRNA level. Several such clones were obtained and none of them had modified cAMP-dependent protein kinase activity, suggesting that mutations were in the post-protein kinase step in the pathway of hormone action. Five clones were fused with the parent LLC-PK1 cells, and all of the fusion cells showed reduced basal uPA mRNA levels, indicating that they were recessive mutants. One clone was analyzed further for sensitivity to calcitonin in the induction of uPA mRNA, and it showed a significantly different dose-response pattern compared with parent cells. These results suggest that the uPA gene is regulated, at least partly, by a negatively regulating factor and that the action of cAMP is linked to this factor.
 ...
PMID:A new genetic approach for studying hormonal regulation of urokinase-type plasminogen activator gene expression in LLC-PK1 cells. 283 Apr 99

A series of stable mutants bearing nuclear genetic markers were developed from the established chicken cell line DU24. The mutants were obtained after mutagenesis of DU24 cells with ethyl methanesulfonate (EMS) or arose spontaneously when plated in the appropriate selective medium. Clones resistant to 5-bromodeoxyuridine (BrdU) were obtained following a two-step selection procedure and analyzed. The BrdUr cells were found to be deficient in thymidine kinase activity and were HAT sensitive. Molecular characterization of these mutants revealed no deletions or other rearrangements, but methylation of some cytosine residues was decreased in the mutants. A similar restriction profile was seen in a series of mutants made resistant to BrdU after cultivation of DU24 cells in increasing concentrations of the drug over a period of six months. Selection of EMS-treated BrdUr cells in 10 microM ouabain gave rise to a clone resistant to both drugs and which was still HAT sensitive. Clones resistant to 6-thioguanine were also isolated, but showed wild-type hypoxanthine phosphoribosyltransferase activity and were HAT resistant. A number of the cell lines isolated were found to be suitable for fusion experiments with both chicken cells and cells from other vertebrate species.
 ...
PMID:Development and characterization of mutant chicken cell lines for somatic cell genetics studies. 316 27

The molecular mechanisms of ethyl methanesulfonate-induced reversion in mammalian cells were studied by using as a target a gpt gene that was integrated chromosomally as part of a shuttle vector. Murine cells containing mutant gpt genes with single base changes were mutagenized with ethyl methanesulfonate, and revertant colonies were isolated. Ethyl methanesulfonate failed to increase the frequency of revertants for cell lines with mutant gpt genes carrying GC----AT transitions or AT----TA transversions, whereas it increased the frequency 50-fold to greater than 800-fold for cell lines with mutant gpt genes carrying AT----GC transitions and for one cell line with a GC----CG transversion. The gpt genes of 15 independent revertants derived from the ethyl methanesulfonate-revertible cell lines were recovered and sequenced. All revertants derived from cell lines with AT----GC transitions had mutated back to the wild-type gpt sequence via GC----AT transitions at their original sites of mutation. Five of six revertants derived from the cell line carrying a gpt gene with a GC----CG transversion had mutated via GC----AT transition at the site of the original mutation or at the adjacent base in the same triplet; these changes generated non-wild-type DNA sequences that code for non-wild-type amino acids that are apparently compatible with xanthine-guanine phosphoribosyltransferase activity. The sixth revertant had mutated via CG----GC transversion back to the wild-type sequence. The results of this study define certain amino acid substitutions in the xanthine-guanine phosphoribosyltransferase polypeptide that are compatible with enzyme activity. These results also establish mutagen-induced reversion analysis as a sensitive and specific assay for mutagenesis in mammalian cells.
 ...
PMID:Molecular analysis of ethyl methanesulfonate-induced reversion of a chromosomally integrated mutant shuttle vector gene in mammalian cells. 318 46

The aim of the study was to establish enzyme-deficient mutants of the human permanent cell line U-937. Following chemical mutagenesis with the use of ethyl methanesulfonate, this cell line was chronically exposed to increasing concentrations of the toxic hypoxanthine analogue 6-thioguanine. Cells surviving hypoxanthine-aminopterin-thymidine selective media were separated by glass adherence with the use of 12-O-tetradecanoyl-phorbol-13-acetate. Three mutant clones were established, which have remained hypoxanthine phosphoribosyltransferase (HPRT) deficient for a period of 7 months, as shown by indirect measurements with the use of autoradiography and scintillation counting of cells exposed to [3H]hypoxanthine. Since the phenotypic properties and growth behavior of U-937 cells have remained unaltered after the induced mutation, a highly restricted chromosomal segment coding for HPRT seems to have been mutated.
 ...
PMID:Induction of hypoxanthine phosphoribosyltransferase deficiency in human U-937 cells. 345 59

Two protocols have been developed, both of which utilize the thymidine analog 5-bromodeoxyuridine (BrdUrd) to induce mutations in mammalian cells in culture (E. R. Kaufman and R. L. Davidson, Proc. Natl. Acad. Sci. USA 75:4982-4986, 1978; E. R. Kaufman, Mol. Cell. Biol. 4:2449-2454, 1984). The first protocol, termed incorporational (INC) mutagenesis, utilizes high concentrations of BrdUrd in the culture medium to generate a high intracellular ratio of BrdUTP/dCTP. The second protocol, termed replicational (REP) mutagenesis, entails the incorporation of BrdUrd into DNA under nonmutagenic conditions, the removal of all BrdUrd from the culture medium, and the subsequent replication of the bromouracil-containing DNA in the presence of high intracellular levels of dTTP and dGTP. Genetic studies using reversion analysis at the hypoxanthine-guanine phosphoribosyltransferase locus were used to determine whether the mechanisms of these two BrdUrd mutagenesis protocols had enough specificity to be distinguishable by their ability to revert various mutants. The results of these studies indicated that (i) mutants induced by INC mutagenesis were induced to revert only by REP mutagenesis and not by INC mutagenesis, (ii) mutants induced by REP mutagenesis were more efficiently reverted by INC mutagenesis than by REP mutagenesis, and (iii) both spontaneous mutants and mutants induced by the chemical mutagen ethyl methanesulfonate showed a high degree of specificity when tested for reversion by the BrdUrd mutagenesis protocols.
 ...
PMID:Reversion analysis of mutations induced by 5-bromodeoxyuridine mutagenesis in mammalian cells. 383 41

DNA-adduct formation and induction of gene mutations were determined simultaneously after treatment with the four ethylating agents, ethyl methanesulfonate (EMS), ethylnitrosourea (ENU), diethyl sulfate (DES), and N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG). Both, in E. coli K-12 (NAL-resistance) and in V79 Chinese hamster cells in culture (HPRT-deficiency), the frequencies of mutation induction by all chemicals were the same when plotted against the amount of O6-ethylguanine formed in DNA, suggesting that this DNA adduct can be used as a common dosimeter for the comparisons of the frequencies of gene mutations induced by ethylating agents in various mutagenicity assay systems. Using ENU, such a comparison was performed between mutation induction in V79 cells in vitro and in the specific-locus assay in the mouse. The data indicate that at equal levels of O6-ethylguanine in the DNA of V79 cells and in testicular DNA from male mice treated with ENU, the frequencies of induced mutants in both assay systems were quite similar. These results support the concept that the determination of premutagenic DNA adducts in vivo can be used to monitor exposure to chemical mutagens and that genetic risk estimations may ultimately be performed on the basis of such measurements and of comparative mutagenesis in vitro and in vivo.
 ...
PMID:Quantitative comparison of genetic effects of ethylating agents on the basis of DNA adduct formation. Use of O6-ethylguanine as molecular dosimeter for extrapolation from cells in culture to the mouse. 391 Apr 16

The nuclear enzyme, poly(ADP-ribose) synthetase is involved in the repair of damaged DNA. We report here the results obtained with 3-aminobenzamide (3AB), an inhibitor of this enzyme, on induced biological effects. 3AB increases the frequency of chromosomal aberrations induced by DMS, EMS, ENU, bleomycin and CldUrd. The magnitude of the effect is dependent on the type of chemical used, the combinations with DMS and EMS being the most potent ones. No potentiation was observed after treatment of cells with MMC. Mutation frequencies were determined on the HPRT locus and showed that 3AB did not increase the frequency of gene mutations induced by EMS, ENU and CldUrd. Cell-cycle progression is affected when cells are grown in medium containing CldUrd and 3AB, primarily when the inhibitor is present during the second cell cycle when substituted DNA becomes replicated. The extent of the effect depends on the amount of analogue incorporated and is independent of the presence of the analogue in the medium during the second cell cycle. Analysis of chromosomal aberrations in delayed G2 cells with the aid of the premature chromosome-condensation technique revealed numerous aberrations after incorporation of CldUrd and treatment with 3AB.
 ...
PMID:Effects of 3-aminobenzamide on Chinese hamster cells treated with thymidine analogues and DNA-damaging agents. Chromosomal aberrations, mutations and cell-cycle progression. 392 78

Inhibition of poly (ADP-ribose) synthesis by agents such as 3-aminobenzamide (3-AB) potentiates the cytotoxic, carcinogenic, and clastogenic effects of certain DNA-damaging agents. Experiments were carried out in Chinese hamster ovary cells to compare chromosome aberration production and cytotoxicity with the induction of somatic mutations at the hypoxanthine-guanine phosphoribosyltransferase (HGPRT) and sodium-potassium ATPase loci after treatment with 3-AB in combination with certain monofunctional alkylating agents. On its own, 1 to 10 mM concentrations of 3-AB were not mutagenic, reduced plating efficiencies only slightly, and produced a small elevation in the frequency of chromatid aberrations. In combination with ethyl methanesulfonate (EMS), 3-AB increased cytotoxicity and the frequency of alkylation-induced chromatid aberrations. 3-AB also increased the frequency of EMS and N-methyl-N'-nitro-N-nitrosoguanidine-induced 6-thioguanine-resistant cells (a marker for the HGPRT- phenotype). It had no effect on the frequency of EMS-induced ouabain-resistant cells (a marker for ATPase mutations). All the effects were dose dependent. Larger absolute increases were found with 10 mM 3-AB as compared with 1 mM 3-AB and with 2 mM EMS as compared to 1 mM EMS. The 3-AB-mediated increases in 6-thioguanine-resistant cells, which are often deletion mutations, and the lack of any increase in the frequency of ouabain-resistant cells, which can only arise through point mutation induction, along with the increases in chromosome aberration frequency, suggests that 3-AB increases the frequency of deletion mutations by increasing the frequency and duration of DNA strand breaks.
 ...
PMID:Comutagenic effects of 3-aminobenzamide in Chinese hamster ovary cells. 397 24


<< Previous 1 2 3 4 Next >>