Methylated CpG dinucleotides are the preferential targets for G-to-T transversion mutations induced by benzo[a]pyrene diol epoxide in mammalian cells: Similarities with the p53 mutation spectrum in smoking-associated lung cancers

A large fraction of the p53 mutations in lung cancers from smokers are G-to -T transversions, a type of mutation that is infrequent in lung cancers fro m nonsmokers and in most other tumors. Previous studies have indicated that there is an association between G-to-T transversion hotspots in lung cance rs and sites of preferential formation of polycyclic aromatic hydrocarbon a dducts along the p53 gene. p53 codons containing methylated CpG sequences a re preferential targets for formation of adducts by (+/-) anti-7 beta ,8 al pha -dihydroxy-9 alpha 10 alpha -epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (B PDE). To assess the role of CpG methylation in induction of mutations by BP DE, we analyzed BPDE mutagenesis in three CpG methylated target genes: a su pF shuttle vector and the ell and lacI transgenes in embryonic mouse fibrob lasts. After methylation of the shuttle vector at all CpG sequences, 42% of all G-to-T transversions were at CpG sites compared with 23% in unmethylat ed DNA. In the cII transgene, which is methylated at CpG sequences in vivo, 83 of 147 (56%) of the BPDE-induced mutations were G-to-T transversions, a nd 58% (48 of 83) of all G-to-T transversions occurred at methylated CpG se quences. In the lacI gene, 68% (75 of 111) of the BPDE-induced mutations we re G-to-T events, and 58 of 75 (77%) of these occurred at methylated CpG se quences. The occurrence of transversion hotspots at methylated CpGs correla ted with high levels of BPDE adducts formed at such sites. This situation m irrors the one in the p53 gene in lung cancers from smokers where 236 of 46 5 (51%) of the G-to-T transversions occurred at methylated CpG sites. These findings further strengthen a link between polycyclic aromatic hydrocarbon s present in cigarette smoke and lung cancer mutations and provide evidence that mutational processes other than C-to-T transition mutations can occur selectively at methylated CpG sequences.