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

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.