Specific complex formation between proteins encoded by the yeast DNA repair and recombination genes RAD1 and RAD10

Abstract

The RAD1 and RAD10 genes of Saccharomyces cerevisiae are required for excision repair of ultraviolet light-damaged DNA, and they also function in a mitotic recombination pathway that is distinct from the double-strand-break recombination pathway controlled by RAD52. Here, we show that the RAD1 and RAD10 proteins are complexed with each other in vivo. Immunoprecipitation of yeast cell extracts with either anti-RAD1 antibody or anti-RAD10 antibody coprecipitated quantitative amounts of both RAD1 and RAD10 proteins. The level of coprecipitable RAD1 and RAD10 increased when both proteins were overproduced together, but not if only one of the proteins was overproduced. The RAD1/RAD10 complex is highly stable, being refractory to 1 M NaCl and to low concentrations of SDS. By hydroxylamine mutagenesis, we have identified a rad1 mutant allele whose encoded protein fails to complex with RAD10. The interaction-defective rad1 mutant resembles the rad1 or rad10 null mutant in defective DNA repair and recombination, implying that complex formation is essential for the expression of biological activities controlled by RAD1 and RAD10.