Novel Checkpoint Pathway Organization Promotes Genome Stability in Stationary-Phase Yeast Cells

Abstract

Most DNA alterations occur during DNA replication in the S phase of the cell cycle. However, the majority of eukaryotic cells exist in a nondividing, quiescent state. Little is known about the factors involved in preventing DNA instability within this stationary-phase cell population. Previously, we utilized a unique assay system to identify mutations that increased minisatellite alterations specifically in quiescent cells in Saccharomyces cerevisiae. Here we conducted a modified version of synthetic genetic array analysis to determine if checkpoint signaling components play a role in stabilizing minisatellites in stationary-phase yeast cells. Our results revealed that a subset of checkpoint components, specifically MRC1, CSM3, TOF1, DDC1, RAD17, MEC3, TEL1, MEC1, and RAD53, prevent stationary-phase minisatellite alterations within the quiescent cell subpopulation of stationary-phase cells. Pathway analysis revealed at least three pathways, with MRC1, CSM3, and TOF1 acting in a pathway independent of MEC1 and RAD53. Overall, our data indicate that some well-characterized checkpoint components maintain minisatellite stability in stationary-phase cells but are regulated differently in those cells than in actively growing cells. For the MRC1-dependent pathway, the checkpoint itself may not be the important element; rather, it may be loss of the checkpoint proteins' other functions that contributes to DNA instability.

ACKNOWLEDGMENTS

We thank P. Jauert for technical assistance. We also thank D. Clarke, S. Elledge, D. Koepp, R. Kolodner, and L. Symington for yeast strains or plasmids. We are grateful to C. Boone and R. Wright for yeast haploid sets.

This work was funded by a grant from the National Institutes of Health (5RO1-GM072598) to David T. Kirkpatrick.