The Preference for Error-Free or Error-Prone Postreplication Repair in Saccharomyces cerevisiae Exposed to Low-Dose Methyl Methanesulfonate Is Cell Cycle Dependent

Abstract

Cells employ error-free or error-prone postreplication repair (PRR) processes to tolerate DNA damage. Here, we present a genome-wide screen for sensitivity to 0.001% methyl methanesulfonate (MMS). This relatively low dose is of particular interest because wild-type cells exhibit no discernible phenotypes in response to treatment, yet PRR mutants are unique among repair mutants in their exquisite sensitivity to 0.001% MMS; thus, low-dose MMS treatment provides a distinctive opportunity to study postreplication repair processes. We show that upon exposure to low-dose MMS, a PRR-defective rad18Δ mutant stalls into a lengthy G₂ arrest associated with the accumulation of single-stranded DNA (ssDNA) gaps. Consistent with previous results following UV-induced damage, reactivation of Rad18, even after prolonged G₂ arrest, restores viability and genome integrity. We further show that PRR pathway preference in 0.001% MMS depends on timing and context; cells preferentially employ the error-free pathway in S phase and do not require MEC1-dependent checkpoint activation for survival. However, when PRR is restricted to the G₂ phase, cells utilize REV3-dependent translesion synthesis, which requires a MEC1-dependent delay and results in significant hypermutability.

ACKNOWLEDGMENTS

We thank Brenda Andrews and Charles Boone for strains and plasmids used in this study and anonymous reviewers for helpful suggestions.

B.D.P. was supported by a U.S. Department of Defense Breast Cancer Research Program predoctoral fellowship. This work was supported by NIH grant R01 CA 129604.