The Slx5-Slx8 Complex Affects Sumoylation of DNA Repair Proteins and Negatively Regulates Recombination

Abstract

Recombination is important for repairing DNA lesions, yet it can also lead to genomic rearrangements. This process must be regulated, and recently, sumoylation-mediated mechanisms were found to inhibit Rad51-dependent recombination. Here, we report that the absence of the Slx5-Slx8 complex, a newly identified player in the SUMO (small ubiquitin-like modifier) pathway, led to increased Rad51-dependent and Rad51-independent recombination. The increases were most striking during S phase, suggesting an accumulation of DNA lesions during replication. Consistent with this view, Slx8 protein localized to replication centers. In addition, like SUMO E2 mutants, slx8Δ mutants exhibited clonal lethality, which was due to the overamplification of 2μm, an extrachromosomal plasmid. Interestingly, in both SUMO E2 and slx8Δ mutants, clonal lethality was rescued by deleting genes required for Rad51-independent recombination but not those involved in Rad51-dependent events. These results suggest that sumoylation negatively regulates Rad51-independent recombination, and indeed, the Slx5-Slx8 complex affected the sumoylation of several enzymes involved in early steps of Rad51-independent recombination. We propose that, during replication, the Slx5-Slx8 complex helps prevent DNA lesions that are acted upon by recombination. In addition, the complex inhibits Rad51-independent recombination via modulating the sumoylation of DNA repair proteins.

SUPPLEMENTAL MATERIAL

We thank Gunter Blobel, in whose lab this project was initiated. We appreciate Steve Brill and Mark Hochstrasser for communication of their data before publication. We thank the following people for reagents: Mark Gartenberg, Gerald Fink, Kevin Verstrepen, Grezegorz Ira, and Lorraine Symington. We thank David Alvaro for help with the initial recombination assays, Kevin Verstrepen for help with the FLO1 recombination assay, and Gene Bryant and Daniel Spagna in the Ptashne lab for their help with real-time PCR. We thank Peter Thorpe, Ken Marians, Lorraine Symington, and Maria Jasin for critical reading of the manuscript.

This work was supported by Cancer Center support grant NCI P30 CA-08478-41 (X.Z.) and Kirschstein NRSA predoctoral fellowships GM073567 (R.C.B.) and GM050237 and GM067055 (R.R.).