Molecular analysis of O6-substituted guanine-induced mutagenesis of ras oncogenes.

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作者：

G Mitra，GT Pauly，R Kumar，GK Pei，SH Hughes，RC Moschel，M Barbacid

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摘要：

We have designed an Ha-ras/thymidine kinase (TK) cassette that permits the incorporation of chemically synthesized adducts within specific domains of the rat Ha-ras protooncogene. This cassette has been used to evaluate the mutagenicity of O6-substituted guanine residues, including O6-methylguanine and O6-benzylguanine, incorporated within the 12th codon of this locus. Mutations were monitored by the ability of these modified Ha-ras DNAs to transform Rat4 TK- cells. Our results indicate that both types of O6-substituted guanines are substantially mutagenic, although the methyl substituent induced a 2-fold higher percentage of transformed Rat4 TK+ colonies than its bulkier benzyl analogue. Interestingly, the mutagenicity of both O6-substituted guanines was found to be independent of their relative position within codon 12, therefore suggesting that the specific activation of Ha-ras oncogenes by GGA → GAA mutations in tumors induced by methylating carcinogens might be due to differences in the accessibility of these guanine residues to the carcinogen rather than to a differential rate of repair. Molecular analysis of the mutations induced by these O6-substituted guanines indicated that O6-methylguanine exclusively induced G → A transitions. In contrast, O6-benzylguanine produced G → C and G → T transversions in addition to G → A transitions. These results suggest that O6-methylguanine and its bulkier analogue O6-benzylguanine may induce mutagenesis by different mechanisms.

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关键词：

Biochemistry O6-methylguanine O6-benzylguanine BspMI Cassette Gene Transfer Polymerase Chain Reaction

DOI：

10.1073/pnas.86.22.8650

被引量：

712

年份：

1989