Mechanism for control of mutagenesis in mammals

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Reactive oxygen species produced during normal cellular matabolism damage DNA and its precursors. An oxidized form of guanine base, 8-oxoguanine, is regarded as most critical in terms of mutagenesis as well as carcinogenesis. An enzyme 8-oxo-dGTPase hydrolyzes 8-oxo-dGTP, an oxidized form of dGTP, to 8-oxo-dGMP, thereby preventing the occurrence of A : T to C : G transversion, caused by rnisincorporation of 8-oxo-dGTP into DNA.To elucidate the role of 8-oxo-dGTPase in carcinogenesis, it is necessary to construct cell or mouse lines with altered levels of the enzyme activity. We isolated the genomic sequence for mouse 8-oxo-dGTPase protein, identified the exon / intron region of the gene MTH1, and characterized the promoter in relation to the regulation of expression of the gene. Based on these data, we performed an gene targeting experiment using ES cells. The ES cells defective in both alleles of the MTH1 gene are viable but yield a moderately higher frequency of spontaneous mutation. By sing MTH1-defective ES cells, we constructed mouse lines defective in both alleles of the MTH1 gene. The mice are viable and apparently normal in appearance but produced a significantly large number of tumores in their organs, especially in stomach.The mouse MTH1 gene has 5 exons, among which the coding sequence resides on the third through the fifth exon, and spans approximately 9 kb. Part of the human and rat genes for 8-oxo-dGTPase has been isolated, and the human gene is composed of five exons while the rat gene has three exons. A comparison of the gross structures of the genes revealed that the overall structures of the human and rat genes are similar to that of the mouse gene.