DNA Interstrand Cross-Link Repair in theSaccharomyces cerevisiaeCell Cycle: Overlapping Roles forPSO2(SNM1) with MutS Factors andEXO1during S Phase

Abstract

Pso2/Snm1 is a member of the ␤-CASP metallo-␤-lactamase family of proteins that include the V(D)J recombination factor Artemis. Saccharomyces cerevisiae pso2 mutants are specifically sensitive to agents that induce DNA interstrand cross-links (ICLs). Here we establish a novel overlapping function for PSO2 with MutS mismatch repair factors and the 5-3 exonuclease Exo1 in the repair of DNA ICLs, which is confined to S phase. Our data demonstrate a requirement for NER and Pso2, or Exo1 and MutS factors, in the processing of ICLs, and this is required prior to the repair of ICL-induced DNA double-strand breaks (DSBs) that form during replication. Using a chromosomally integrated inverted-repeat substrate, we also show that loss of both pso2 and exo1/msh2 reduces spontaneous homologous recombination rates. Therefore, PSO2, EXO1, and MSH2 also appear to have overlapping roles in the processing of some forms of endogenous DNA damage that occur at an irreversibly collapsed replication fork. Significantly, our analysis of ICL repair in cells synchronized for each cell cycle phase has revealed that homologous recombination does not play a major role in the direct repair of ICLs, even in G 2 , when a suitable template is readily available. Rather, we propose that recombination is primarily involved in the repair of DSBs that arise from the collapse of replication forks at ICLs. These findings have led to considerable clarification of the complex genetic relationship between various ICL repair pathways.

A mutation in EXO1 defines separable roles in DNA mismatch repair and post-replication repair

“…However, based on the exo1Δ mutational spectra and genetic interaction with REV3, we previously hypothesized that EXO1 participates in at least one MMR-independent mutation avoidance pathway [12]. In agreement with our supposition, recent studies have identified a potential role for EXO1 in the maintenance and repair of stalled replication forks [20][21][22].…”

“…In addition to the role of EXO1 in PRR proposed here, EXO1 has been implicated to function in cell cycle checkpoint, fork maintenance and fork repair pathways [20][21][22]51,52]. EXO1 has been shown to play subtle roles in end-processing for mitotic recombination and function redundantly with at least one other unidentified nuclease.…”

“…Thus, the requirement for Exo1p in the absence of Sgs1p suggests an important role for Exo1p in the maintenance, repair or restart of stalled replication forks. Finally, EXO1 and PSO2 appear to have overlapping roles in the processing of collapsed replication forks due to some forms of endogenous DNA damage and from nitrogen mustard induced interstrand cross-links [21].…”

The Pso4 mRNA Splicing and DNA Repair Complex Interacts with WRN for Processing of DNA Interstrand Cross-links

“…Consistent with these biochemical findings, it has been shown that Msh2 and Ercc1-Xpf act cooperatively in human cells to remove cisplatin ICLs and that these proteins interact as determined by co-immunoprecipitation experiments (22). Furthermore, recent findings in budding yeast have implicated a role for Msh2 and Exo I in repair of nitrogen mustard ICLs (23). Taken together these findings suggest that mismatch repair factors and Ercc1-Xpf are involved in a novel pathway of ICL repair.…”

DNA polymerase η reduces the γ-H2AX response to psoralen interstrand crosslinks in human cells

“…In yeast and mammalian cells, nucleotide excision repair (NER), recombination, and translesion synthesis, as well as certain mismatch repair proteins, have been reported to participate in ICL repair [2,[5][6][7][8][9][10]. Recombinational repair is in part thought to be a response to formation of double-strand breaks (DSBs) at ICL that appear to be unique to eukaryotes and that arise either as a result of collapsed replication forks blocked at ICL, or as true repair intermediates [8,9,[11][12][13].…”