N-Methyl-N′-nitro-N-nitrosoguanidine-resistant HeLa S3 cells still have little O6-methylguanine-DNA methyltransferase activity and are hypermutable by alkylating agents

Abstract

To clarify the involvement of O⁶ -methylguanine ( O⁶ -MeG) in mutagenesis, we isolated N -methyl- N ′-nitro- N -nitrosoguanidine (MNNG)-resistant cells, MR10-1 from HeLa S3 mer ⁻ cells. MR10-1 cells were 40 times more resistant to MNNG than the parental cells. MR10-1 cells were also significantly more resistant to N -methyl- N -nitrosourea and slightly more resistant to methyl methanesulfonate and dimethyl sulfate than parental cells. However, we found that MR10-1 cells had still little O⁶ -MeG-DNA methyltransferase activity and were sensitive to 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride, like HeLa mer ⁻ cells, thereby showing that MR10-1 cells are still mer ⁻ . When induced 6-thioguanine (6TG)-resistant colonies were plotted as a function of the corresponding percentage survival, the resistant colonies of MR10-1 cells were induced much more frequently than in the case of HeLa mer ⁻ cells. However, induction of 6TG-resistant cells in the both cell lines did not differ significantly in terms of mutant cells per 0.1 μM MNNG. On the contrary, MR10-1 cells (mer ⁻ ) and two HeLa S3 mer ⁺ cells lines differed in the induction of mutation as a function of MNNG concentration. The HeLa mer ⁺ cell lines were not mutable, while MR10-1 cells were highly mutable. These above results clearly show that the HeLa mer ⁻ cell has at least two defects in the repair of the alkylated adducts which are related to cell killing and mutation, and also suggest that O ⁶ -MeG is involved in the induction of mutation.