Saccharomyces cerevisiae Mus81-Mms4 and Rad52 can cooperate in the resolution of recombination intermediates.

Mus81 is a well-conserved DNA structure-specific endonuclease which belongs to the XPF/Rad1 family of proteins that are involved in DNA nucleotide excision repair. Mus81 forms a heterodimer with a non-catalytic subunit, Mms4, in Saccharomyces cerevisiae (Eme1/EME1 in Schizosaccharomyces pombe and mammals). Recent evidence shows that Mus81 functions redundantly with Sgs1, a member of the ubiquitous RecQ family of DNA helicases, to process toxic recombinant intermediates. In budding yeast, homologous recombination is regulated by the Rad52 epistasis group of proteins, including Rad52, which stimulates the main steps of DNA sequence-homology searching. Mus81 was proven to act in the Rad52-dependent pathway. Here, we demonstrate that Rad52 and Mus81-Mms4 possesses a functional interaction; the presence of Rad52 significantly enhances the endonuclease activity of Mus81-Mms4 on a broad range of its preferred synthetic substrates. Furthermore, this functional interaction is demonstrated to be species specific. We fragmented Rad52 and found that the N-terminal fragment from the 86th to 169th amino acid residue, which belongs to DNA-binding and self-association domains, can stimulate Mus81-Mms4 endonuclease. These results strongly support the notion that Rad52 and Mus81-Mms4 collaborate and work jointly in processing of homologous recombination intermediates.

The possible function of Flp1 in homologous recombination repair in Saccharomyces cerevisiae.

Saccharomyces cerevisiae Mus81 is a structure-selective endonuclease which constitutes an alternative pathway in parallel with the helicase-topoisomerase Sgs1-Top3-Rmi1 complex to resolve a number of DNA intermediates during DNA replication, repair, and homologous recombination. Previously, it was showed that the N-terminal region of Mus81 was required for its in vivo function in a redundant manner with Sgs1; mus81Δ120N mutant that lacks the first 120 amino acid residues at the N-terminus exhibited synthetic lethality in combination with the loss of SGS1. In this study, the physiologically important role of the N-terminal region of Mus81 in processing toxic intermediates was further investigated. We examined the cellular defect of sgs1Δmus81Δ100N cells and observed that although viable, the cells became very sensitive to DNA damaging agents. A single-copy suppressor screening to seek for a factor(s) that could rescue the drug sensitivity of sgs1Δmus81Δ100N cells was performed and revealed that Flp1, a site-specific recombinase 1 encoded on the 2-micron plasmid was a suppressor. Moreover, Flp1 overexpression could partially suppress the drug sensitivity of mus81Δ cells at 37 °C. Our findings suggest a possible function of Flp1 in coordination with Mus81 and Sgs1 to jointly resolve the branched-DNA structures generated in cells attempting to repair DNA damages.