Fission Yeast Rad51 and Dmc1, Two Efficient DNA Recombinases Forming Helical Nucleoprotein Filaments

Abstract

Homologous recombination is important for the repair of double-strand breaks during meiosis. Eukaryotic cells require two homologs of Escherichia coli RecA protein, Rad51 and Dmc1, for meiotic recombination. To date, it is not clear, at the biochemical level, why two homologs of RecA are necessary during meiosis. To gain insight into this, we purified Schizosaccharomyces pombe Rad51 and Dmc1 to homogeneity. Purified Rad51 and Dmc1 form homo-oligomers, bind single-stranded DNA preferentially, and exhibit DNA-stimulated ATPase activity. Both Rad51 and Dmc1 promote the renaturation of complementary single-stranded DNA. Importantly, Rad51 and Dmc1 proteins catalyze ATP-dependent strand exchange reactions with homologous duplex DNA. Electron microscopy reveals that both S. pombe Rad51 and Dmc1 form nucleoprotein filaments. Rad51 formed helical nucleoprotein filaments on single-stranded DNA, whereas Dmc1 was found in two forms, as helical filaments and also as stacked rings. These results demonstrate that Rad51 and Dmc1 are both efficient recombinases in lower eukaryotes and reveal closer functional and structural similarities between the meiotic recombinase Dmc1 and Rad51. The DNA strand exchange activity of both Rad51 and Dmc1 is most likely critical for proper meiotic DNA double-strand break repair in lower eukaryotes.

ACKNOWLEDGMENTS

We are grateful to Isabelle Brodeur, Steve West, and Rhea Utley for helpful comments on the manuscript. We also thank Steve West and Jacques Dubochet for support and interest, Alexander Mazin for the communication of unpublished data, and Hiroyuki Yamano for the gift of S. pombe cDNA and genomic libraries.

A.S. is supported by Swiss National Foundation grant 3100A0-103962. J.-Y.M. is a Canadian Institutes of Health New Investigator, and this research is supported by funds from the National Cancer Institute of Canada and the Natural Sciences and Engineering Research Council of Canada.