We have identified two C. elegans proteins containing ERCC4-like domains, which are responsible for incising damaged DNA: the XPF orthologue (C47D12.8) and a gene that we have named
mus-81 (C43E11.2) because it appears to be an orthologue of yeast Mus81. Yeast Mus81 (MMS and UV sensitive 81) was identified through 2-hybrid screens; in S. pombe Mus81 interacts with Cds1, a cell cycle checkpoint kinase, and in S. cerevisiae Mus81 interacts with Rad54, a DNA repair protein. Mus81 is reported to participate in repair of DNA damage caused by UV irradiation or MMS. The
mus81 orthologue in C. elegans is likely to be the terminal gene in an operon. We have shown that a 1.3 kb
mus-81 transcript is produced and trans-spliced to SL2. All known Mus81 proteins contain two non-sequence specific DNA-binding hairpin-helix-hairpin motifs flanking the ERCC4 domain. The predicted C. elegans MUS-81 is no exception and shares 50% similarity to Mus81 proteins from other phyla. Depletion of
mus-81 by RNAi causes hypersensitivity to UV irradiation; treated animals display a decrease in brood size, a reduction in viability and an increase in germ cell apoptosis when compared to similarly treated N2 worms. In addition, continuous depletion of
mus-81 by RNAi through multiple generations eventually leads to sterility and an increased frequency of abnormal karyotypes. Together these results suggest that
mus-81 plays a role in maintaining genome stability. Yeast that lack Mus81 are defective in meiosis and in vitro analysis suggests that Mus81 might be a Holliday Junction (HJ) resolvase. We have failed to detect meiotic defects in
mus-81 (RNAi) worms, so it is unlikely that MUS-81 functions as a HJ resolvase in C. elegans. We are investigating the possibility that the C. elegans XPF endonuclease orthologue is the HJ resolvase. Presently we are analysing the expression patterns of
mus-81 gene products by Northern analysis and through the use of a MUS-81::GFP reporter. The fusion is expressed in the nucleus and is enriched in the nucleolus. We are also pursuing protein interaction studies to identify MUS-81 partners and have generated an anti-MUS-81 polyclonal antibody to aid in our analysis.