CONTRIBUTION OF DNA METHYLATION AND BENZYLATION TO N-NITROSO-N-BENZYL-METHYLAMINE-INDUCED MUTAGENESIS IN BACTERIA - EFFECTS OF RAT-LIVER CYTOCHROME-P450 ISOZYMES AND GLUTATHIONE TRANSFERASES

The mutagenicity of N-nitroso-Nbenzyl-methylamine (NBzMA), N-benzyl-N-nitrosourea (BzNU) and N-methyl-N-nitrosourea (MNU) in Salmonella typhimurium strains was Investigated. BzNU selectively mutated TA100 strain as compared to TA1535, whereas MNU showed an inverse strain response, an effect probably related to the fact that benzylation of DNA is a stronger inducerof SOS DNA repair than methylation, as indicated by the higher activity of BzNU in the SOS chromotest. Benzylation of bacterial DNA by NBzMA, as deduced from the differential strain res ponsiveness, contributed predominantly to its mutagenicityin the presence of liver preparation from untreated, Aroclor or ethanol-treated rats. Since benzyl alcohol, a metabolite of NIBzMA, was not mutagenic in S.typhimurium, it appears that benzyl carbonium cations responsible for the mutagenicity of NBzMA in TA100 are formed via cytochrome P450-mediated hydroxylation of the methyl group. Neither femc-EDTA nor desferrioxainine altered the mutagenicity of NIBzMA, suggesting that activation occurs mainly within the catalyticsite of P450. Experiments with isozyme-specific monodonal antibodies showed that P450IIE1 did not contribute to N-demethylation of NBzMA at either low or high substrate concentrations and that P450IA contributed only wealdy. Debenzylation was catalysed predominantly by P4SOIA at high NIBzMA concentration. Antibodies against rat liver P450IIB enhanced NBzMA mutagenicity in S.typhimurium TA1535 strain up to 17-fold at low substrate concentration, but were without effect at high concentration. lii liquid incubation assays, a 100% GSH-dependent reduction of NIBzMA mutagenicity was found with liver S9 from untreated Wistar rats. The reducing effect of GSH was less pronounced in the presence of liver S9 from BIIWII or Fischer 344 rats. © 1990 Oxford University Press.