Interplay of DNA Repair Pathways Controls Methylation Damage Toxicity in Saccharomyces cerevisiae

Abstract

Abstract Methylating agents of SN1 type are widely used in cancer chemotherapy, but their mode of action is poorly understood. In particular, it is unclear how the primary cytotoxic lesion, O6-methylguanine (MeG), causes cell death. One hypothesis stipulates that binding of mismatch repair (MMR) proteins to MeG/T mispairs arising during DNA replication triggers cell-cycle arrest and cell death. An alternative hypothesis posits that MeG cytotoxicity is linked to futile processing of MeG-containing base pairs by the MMR system. In this study, we provide compelling genetic evidence in support of the latter hypothesis. Treatment of 4644 deletion mutants of Saccharomyces cerevisiae with the prototypic SN1-type methylating agent N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) identified MMR as the only pathway that sensitizes cells to MNNG. In contrast, homologous recombination (HR), postreplicative repair, DNA helicases, and chromatin maintenance factors protect yeast cells against the cytotoxicity of this chemical. Notably, DNA damage signaling proteins played a protective rather than sensitizing role in the MNNG response. Taken together, this evidence demonstrates that MeG-containing lesions in yeast must be processed to be cytotoxic.