Assembly of RecA-like recombinases: Distinct roles for mediator proteins in mitosis and meiosis

Members of the RecA family of recombinases from bacteriophage T4, Escherich ia coil, yeast, and higher eukaryotes function in recombination as higher-o rder oligomers assembled on tracts of single-strand DNA (ssDNA). Biochemica l studies have shown that assembly of recombinase involves accessory factor s. These studies have identified a class of proteins, called recombination mediator proteins, that act by promoting assembly of recombinase on ssDNA t racts that are bound by ssDNA-binding protein (ssb). In the absence of medi ators, ssb inhibits recombination reactions by competing with recombinase f or DNA-binding sites. Here we briefly review mediated recombinase assembly and present results of new in vivo experiments. Immuno-double-staining expe riments in Saccharomyces cerevisiae suggest that Rad51, the eukaryotic reco mbinase, can assemble at or near sites containing ssb (replication protein A, RPA) during the response to DNA damage, consistent with a need for media tor activity. Correspondingly, mediator gene mutants display defects in Rad 51 assembly after DNA darhage and during meiosis, although the requirements for assembly are distinct in the two cases. In meiosis, both Rad52 and Rad 55/57 are required, whereas either Rad52 or Rad55/57 is sufficient to promo te assembly of Rad51 in irradiated mitotic cells. Rad52 promotes normal amo unts of Rad51 assembly in the absence of Rad55 at 30 degreesC but not 20 de greesC, accounting for the cold sensitivity of rad55 null mutants, Finally, we show that assembly of Rad51 is induced by radiation during S phase but not during G(1), consistent with the role of Rad51 in repairing the spontan eous damage that occurs during DNA replication.