Resistance of Salmonella typhimurium TA 1535 to O6-guanine methylation and mutagenesis induced by low doses of N-methyl-N'-nitro-N-nitrosoguanidine: an apparent constitutive repair activity

Salmonella tester strains which are reverted by base-pair substitutionmutagens are relatively insensitive to the mutagenic effectsof N-methyl-N-nitroso compounds. One reason for this insensitivityis the ability of these strains to withstand low doses of thesecompounds before they become sensitive to their mutagenic effects.In this report it is shown that mutagenesis induced by treatmentof Salmonella typhimurium TA 1535 with N-methyl-N'-nitro-N-nitrosoguanidine(MNNG) in buffer is biphasic with a low sensitivity range atlow doses where little mutagenesis occurs, followed by a highsensitivity range whose onset begins after an apparent thresholddose has been exceeded. Levels of O⁶-methylguanine (O⁶-MeG)in the DNA extracted from the bacteria follow a similar dose-responsecurve suggesting a dependency of mutagenesis on O⁶-MeG. In contrast,levels of 7-methylguanine (7-MeG) in the DNA increase linearlywith dose. O⁶-MeG was undetectable at the lowest dose of MNNGwhereas 7-MeG was readily detectable. Although such resistanceto O⁶-alkylation has been demonstrated in MNNG-pretreated (adapted)E. coli, it has not been reported in unpretreated cells. Whenisolated DNA was treated with MNNG a linear dose-response inthe generation of O⁶-MeG was observed. The lack of O⁶-MeG inDNA isolated from MNNG treated cells after low doses is attributedto a saturable, constitutive repair activity in the bacteria.An attempt to observe the removal of O⁶-MeG from the bacteriaafter exposure to a short challenge dose of N-nitroso-N-methylurea(NMU) followed by a subsequent incubation in buffer was unsuccessful,probably because all the repair occurred within the time necessaryto treat and lyse the cells.