Mutation and formation of methyl- and hydroxylguanine adducts in DNA caused by N-nitrosodimethylamine and N-nitrosodiethylamine with UVA irradiation

Abstract

Previously we reported that when Escherichia coli was treated with N-nitrosodialkylamine under irradiation with near UV light, mutagenesis of the bacteria took place: there was no requirement for metabolic activation. We have now studied the spectra of mutations caused by N-nitrosodimethylamine (NDMA) and N-nitrosodiethylamine (NDEA) with UVA (320-400 nm) irradiation, using standard tester strains for identifying types of mutations. Induced mutations by NDMA + UVA were the transition GC-->AT and transversions GC-->CG, GC-->TA and AT-->TA. NDEA + UVA induced mainly the GC-->CG transversion. In both cases no frameshift mutations were observed. When O6-alkylguanine-DNA alkyltransferase-deficient strains of E. coli and Salmonella typhimurium were used, the mutation levels with both NDMA + UVA and NDEA + UVA became remarkably higher than those observed with the proficient strains. We measured the O6-methylguanine (O6-meG) level in calf thymus DNA treated with NDMA + UVA. The O6-meG level was increased as a function of NDMA concentration and irradiation time. We also detected N7-methylguanine in DNA treated with NDMA + UVA. In our previous work we found formation of 8-oxodeoxyguanosine (8-oxodG) in DNA treated with N-nitrosomorpholine + UVA. The 8-oxodG/dG ratio in DNA treated with NDMA + UVA increased up to 42-fold over that of the untreated control and that in DNA treated with NDEA + UVA increased up to 67-fold. 8-OxodG formation was not affected by replacing H2O in the reaction mixture with D2O, suggesting that singlet oxygen is not the rate limiting factor in this photoactivation. We conclude that both alkylation and oxidation are involved in mutations induced by NDMA + UVA and NDEA + UVA.