The SUMO Isopeptidase Ulp2p Is Required to Prevent Recombination-Induced Chromosome Segregation Lethality following DNA Replication Stress

Abla A Bakir,Kristen N Nguyen,Jeff Bachant,Ming-Ta Lee

doi:10.1371/journal.pgen.1001355

Abla A Bakir, Kristen N Nguyen + Show 2 more

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https://doi.org/10.1371/journal.pgen.1001355

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Journal: PLoS Genetics	Publication Date: Mar 31, 2011
Citations: 45	License type: CC BY 4.0

Affiliation: University of California, Riverside

Abstract

SUMO conjugation is a key regulator of the cellular response to DNA replication stress, acting in part to control recombination at stalled DNA replication forks. Here we examine recombination-related phenotypes in yeast mutants defective for the SUMO de-conjugating/chain-editing enzyme Ulp2p. We find that spontaneous recombination is elevated in ulp2 strains and that recombination DNA repair is essential for ulp2 survival. In contrast to other SUMO pathway mutants, however, the frequency of spontaneous chromosome rearrangements is markedly reduced in ulp2 strains, and some types of rearrangements arising through recombination can apparently not be tolerated. In investigating the basis for this, we find DNA repair foci do not disassemble in ulp2 cells during recovery from the replication fork-blocking drug methyl methanesulfonate (MMS), corresponding with an accumulation of X-shaped recombination intermediates. ulp2 cells satisfy the DNA damage checkpoint during MMS recovery and commit to chromosome segregation with similar kinetics to wild-type cells. However, sister chromatids fail to disjoin, resulting in abortive chromosome segregation and cell lethality. This chromosome segregation defect can be rescued by overproducing the anti-recombinase Srs2p, indicating that recombination plays an underlying causal role in blocking chromatid separation. Overall, our results are consistent with a role for Ulp2p in preventing the formation of DNA lesions that must be repaired through recombination. At the same time, Ulp2p is also required to either suppress or resolve recombination-induced attachments between sister chromatids. These opposing defects may synergize to greatly increase the toxicity of DNA replication stress.

Highlights

As part of the DNA damage response, homologous recombination (HR), template switch recombination through the post-replication DNA repair pathway (PRR), provides an important mechanism for restarting stalled replication forks and filling in un-replicated gaps in DNA
Our data indicate that cells defective for Ulp2p develop a love/hate relationship with recombination, requiring recombination for viability while failing to resolve chromosome attachments induced by recombination repair
For rad52D, we examined this apparent synthetic lethality further by isolating ulp2 deletion mutant (ulp2D) rad52D segregants harboring a wild type (WT) copy of RAD52 on a URA3 minichromosome. ulp2D rad52D/pRAD52 mutants grew weakly, if at all, on media containing 5-FOA, a drug that only allows growth if cells are capable of losing pRAD52 (Figure 1A)

Summary

Introduction

As part of the DNA damage response, homologous recombination (HR), template switch recombination through the post-replication DNA repair pathway (PRR), provides an important mechanism for restarting stalled replication forks and filling in un-replicated gaps in DNA (reviewed in [1,2]). These recombination events must be managed carefully, . Following replication fork stalling at MMSinduced DNA lesions, PRR proteins catalyze either mono- or poly-ubiquitinylation of PCNA K164 [5] These modifications recruit trans-lesion bypass polymerases or induce template switching HR, respectively, providing alternative mechanisms to bypass the lesion and restart replication [5,11,12,13]. They resemble mutants defective for the Sgs1p/Top3p/Rmi1p complex, which, through concerted heli-

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