Rpd3L Contributes to the DNA Damage Sensitivity of Saccharomyces cerevisiae Checkpoint Mutants.

Belén Gómez-González,Anne Early,John F X Diffley,Harshil Patel

doi:10.1534/genetics.118.301817

Abstract

DNA replication forks that are stalled by DNA damage activate an S-phase checkpoint that prevents irreversible fork arrest and cell death. The increased cell death caused by DNA damage in budding yeast cells lacking the Rad53 checkpoint protein kinase is partially suppressed by deletion of the EXO1 gene. Using a whole-genome sequencing approach, we identified two additional genes, RXT2 and RPH1, whose mutation can also partially suppress this DNA damage sensitivity. We provide evidence that RXT2 and RPH1 act in a common pathway, which is distinct from the EXO1 pathway. Analysis of additional mutants indicates that suppression works through the loss of the Rpd3L histone deacetylase complex. Our results suggest that the loss or absence of histone acetylation, perhaps at stalled forks, may contribute to cell death in the absence of a functional checkpoint.

Highlights

DNA replication forks that are stalled by DNA damage activate an S-phase checkpoint that prevents irreversible fork arrest and cell death
Mutation/loss of both EXO1 and RXT2 contributed to the suppression of lethality in methyl methanesulfonate (MMS) previously seen in YMS6 (Segurado and Diffley 2008)
Our previous conclusion that deletion of EXO1 suppresses irreversible fork arrest in rad53D cells is likely unaffected by this discovery, since the strains used in the replication fork stability experiments were derived separately from YMS6 and showed MMS sensitivity similar to YGDP939

Summary

Introduction

DNA replication forks that are stalled by DNA damage activate an S-phase checkpoint that prevents irreversible fork arrest and cell death. Checkpoint kinases regulate many aspects of cell metabolism in response to DNA damage (Zegerman and Diffley 2009) These include blocking cell cycle progression (Allen et al 1994), suppressing late origin firing (Santocanale and Diffley 1998; Shirahige et al 1998; Santocanale et al 1999; Lopez-Mosqueda et al 2010; Zegerman and Diffley 2010), altering global gene expression patterns (Zhou and Elledge 1993; Allen et al 1994; Huang et al 1998), preventing irreversible DNA replication fork arrest (Lopes et al 2001; Tercero and Diffley 2001; Tercero et al 2003), and upregulating deoxyribonucleoside triphosphate (dNTP) levels (Desany et al 1998; Zhao et al 1998). We have taken an unbiased genetic approach to further understand why checkpoint mutant cells are hypersensitive to DNA damage, revealing a connection between chromatin state and replication fork stability

Methods

Results

Conclusion