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:2.4.2.7 (adenine phosphoribosyltransferase)
692 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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

Chinese hamster ovary (CHO) cell lines heterozygous at both the adenine phosphoribosyltransferase (aprt) and thymidine kinase (tk) loci were used for single-step selection of spontaneous and induced mutants resistant to 8-azaadenine (AAr), 6-thioguanine (TGr), ouabain (OUAR), or 5-fluorodeoxyuridine (FUdRr). Mutation data are reported for direct mutagens (EMS, ethyl methanesulfonate; MNNG, N-methyl-N'-nitro-N-nitrosoguanidine; NQO, 4-nitroquinoline 1-oxide) and promutagens (DMN, dimethylnitrosamine; BP, benzo[a]-pyrene) activated by rat-liver homogenates. Optimal plating densities were established for AAr, TGr, OUAR and FUdRr. The induced mutant frequencies as a function of relative cell survival after treatment with EMS, DMN or BP were 2--4 d for AAr, 6--8 d for TGr, 3 d for OUAR, and 1--3 d for FUdRr. The induced mutant frequencies as a function of relative cell survival after treatment with EMS, DMN or BP showed locus-specific differences in sensitivity. Of 61 clonal isolates resistant to AA and assayed for APRT activity, 87% had less than or equal to 5% wild-type activity; of 30 TGr clones assayed, 83% had less than or equal to 5% wild-type HGPRT activity. Of 42 FUdRr clones assayed, 98% had less than or equal to 1% wild-type TK activity. 50 clones selected in medium containing FUdR displayed cross-resistance to 5-bromodeoxyuridine (BUdR) and trifluorothymidine (TFT) and all were sensitive to HAT (hypoxanthine--amethopterin--thymidine) medium. The tk locus showed the largest mutational response as a function of cell survival after mutagen treatment. The rapid expression kinetics for FUdRr and the possibility that the locus detects a broader spectrum of genetic lesions than the other drug-resistance markers are discussed in terms of a sensitive screening assay for detecting potential mutagens.
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PMID:Mutagenicity testing in mammalian cells. II. Validation of multiple drug-resistance markers having practical application for screening potential mutagens. 644 64

The effect of DNA methylation on the expression of the hamster adenine phosphoribosyltransferase (aprt) gene in mouse cells has been examined. This gene was methylated in vitro at all of its C-C-G-G sites by using Hpa II methylase and was inserted into mouse Ltk- aprt- L cells by cotransformation, with the herpes virus thymidine kinase gene as a selectable vector. Whereas clones carrying unmethylated aprt sequences were found to have an aprt+ phenotype as shown by their ability to grow in azaserine-containing medium, almost all clones carrying methylated aprt sequences were shown to be phenotypically aprt-. Blot hybridization analysis demonstrated that both the methylated and unmethylated aprt sequences were integrated into the cellular genome to the same extent and that the in vitro modification was stably maintained in these cells for many generations. When clones containing methylated aprt genes were exposed to conditions that select for the expression of the aprt gene, a low frequency of reversion to the aprt+ phenotype was observed. In all of these clones, this reversion was accompanied by reorganization and undermethylation of the aprt sequences. These results show that the expression of certain genes may be inhibited by site-specific methylation of these sequences and suggest that methylation may play a direct role in the regulation of gene expression.
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PMID:In vitro methylation of the hamster adenine phosphoribosyltransferase gene inhibits its expression in mouse L cells. 695 87

Transformation, or DNA-mediated gene transfer, permits the introduction of new genetic information into a cell and frequently results in a change in phenotype. The transforming DNA is ultimately integrated into a recipient cell chromosome. No unique chromosomal locations are apparent, different lines contain the transforming DNA on different chromosomes. Expression of transformed genes frequently results in the synthesis of new polypeptide products which restore appropriate mutant cells to the wild-type phenotype. Thus transformation provides an in vivo assay for the functional role of DNA sequence organization about specific genes. Transforming genes coding for selectable functions, such as adenine phosphoribosyltransferase or thymidine kinase, have now been isolated by utilizing transformation in concert with molecular cloning. Finally, transformation may provide a general approach to the analysis of complex heritable phenotypes by permitting the distinction between phenotypic changes without concomitant changes in DNA and functional genetic rearrangements.
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PMID:Altering genotype and phenotype by DNA-mediated gene transfer. 741 20

Chromosomal aberrations in human gliomas are principally numerical. In tumours of low malignancy, karyotypes are frequently normal, but occasionally an excess of chromosome 7 and a loss of sex chromosome are observed. In highly malignant tumours, the most frequent aberrations are gain of chromosome 7, loss of chromosome 10 and less frequently losses or deletions of chromosomes 9, 22, 6, 13 and 14 or gains of chromosomes 19 and 20. To understand the meaning of these chromosome imbalances, the relationships between chromosome abnormalities and metabolic disturbances were studied. The losses or deletions observed affected principally chromosomes carrying genes encoding enzymes involved in purine metabolism. The activities of ten enzymes were measured: adenosine kinase, adenine phosphoribosyltransferase, adenylate kinase, methylthioadenosine phosphorylase, hypoxanthine phosphoribosyltransferase, adenylosuccinate lyase, inosine monophosphate dehydrogenase, adenosine deaminase, nucleoside phosphorylase and adenosine monophosphate deaminase. In parallel, two enzymes involved in pyrimidine metabolism, thymidine kinase and thymidylate synthase (TS), were studied. The activities of all these enzymes were measured on samples from 30 human primary glial tumours with low or high malignancy, six xenografted tumours at different passages, four portions of normal brain tissue and four non-glial brain neoplasms. As suggested by cytogenetic data, the enzymatic results showed a relatively low activity of purine metabolism in glial tumours when compared with normal brain and non-glial brain neoplasms. Considering the two enzymes involved in pyrimidine metabolism, only TS had higher activity in glial tumours of high malignancy than in normal brain. In comparison with normal brain, the balance between salvage and de novo pathways changes in gliomas, and even more in grafted tumours, in favour of de novo synthesis. The relation between chromosomes and metabolic imbalances does not correspond to a simple gene dosage effect in these tumours. These data suggest that the decrease of adenosine metabolism occurs before chromosomal aberrations appear, since it is observed in tumours of low malignancy when most karyotypes are still normal, and that the de novo pathway increases with tumour progression.
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PMID:Purine and pyrimidine metabolism in human gliomas: relation to chromosomal aberrations. 805 68

Previous experiments in our research group showed that 3'-azido-3'-deoxythymidine (AZT) caused increased mutant frequencies (Mfs) at the X-linked hypoxanthine-guanine phosphoribosyltransferase (HPRT) and the autosomal thymidine kinase (TK) genes in human lymphoblastoid cells and that there was a significant positive correlation between AZT incorporation into cellular DNA and AZT-induced TK Mfs. In the current study, the mutagenicity of AZT was further evaluated at the autosomal adenine phosphoribosyltransferase (APRT) gene. AZH1 cells, a human lymphoblastoid cell line heterozygous at the APRT locus, were exposed to 300 microM AZT for 0, 1, 3 or 6 days or to 0, 33, 100, 300 or 900 microM AZT for 3 days (n = 5 flasks/group). A cell cloning assay was used to quantitate APRT Mfs. AZT-induced APRT Mf increased with extended duration and with incremental concentrations of AZT exposure. There was a positive correlation (P = 0.022, coefficient = 0.93) between AZT incorporation into DNA and AZT-induced APRT Mfs. RFLP analyses indicated that AZT exclusively induced loss of heterozygosity in APRT mutants. These results, which are consistent with findings on the mutagenicity of AZT at the HPRT and TK genes, indicate the need for further investigations on the potential long-term side effects of AZT on humans, especially those who receive AZT for a prophylactic reason.
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PMID:Mutagenicity and loss of heterozygosity at the APRT locus in human lymphoblastoid cells exposed to 3'-azido-3'-deoxythymidine. 1097 Apr 46

Nucleotide pool imbalances have been reported to affect the fidelity of DNA replication and repair in prokaryotic and eukaryotic cells. We have reported previously that the mutagen-hypersensitive thymidine kinase (TK)-deficient Friend erythroleukemia (FEL) cells (subclones 707BUF and 707BUE), have a more than sixfold increase in the dCTP:dTTP pool ratio when compared to that of wild-type, TK-positive (TK(+)) clone 707 cells. In this study we present the results of an investigation of the effect of the dCTP:dTTP pool imbalance on the accuracy of DNA replication within 707BUF cells. We examined the spontaneous mutation spectra occurring at the adenine phosphoribosyltransferase (aprt) locus within clone 707 (TK(+)) and 707BUF (TK(-)) FEL cells. Mutations recovered at the aprt locus in FEL cells comprised: base substitutions (43:73), frameshifts (14:13.5), and deletions (43:13.5) [clone 707 (TK(+)):707BUF (TK(-)), respectively, expressed as percentages]. A comparison of the mutation spectra obtained for the two cell lines did not reveal any significant increase in misincorporation of dCTP, the nucleotide in excess, in 707BUF (TK(-)) cells, during DNA replication synthesis. These data suggest that the dCTP:dTTP pool imbalance does not alter the fidelity of DNA replication synthesis in 707BUF (TK(-)) FEL cells. Rather, the predominance of GC --> AT transitions (53%) in the 707BUF (TK(-)) spectrum may reflect a reduced efficiency of repair by uracil DNA glycosylase of uracil residues within these cells.
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PMID:Effect of a dCTP:dTTP pool imbalance on DNA replication fidelity in Friend murine erythroleukemia cells. 1101 6


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