Livingston 9603515 The research investigated in this project utilizes classical genetic and molecular genetic techniques to examine the yeast RAD52 gene. This gene is important to protect yeast cells from ionizing radiation and to aid them in recombining their chromosomes. By mutagenizing a cloned copy of the gene, rad52 mutants with varying properties can be recovered. Temperature sensitive mutations have already been made and characterized. These mutants are conditionally sensitive to ionizing radiation but can recombine their chromosomes, suggesting that the protein product of the RAD52 gene (Rad52p) may have more than one role. The current project intends to examine three aspects of the biology of RAD52. First, why do mutations altering amino acid residues in the N-terminal domain of Rad52p have a drastically different effect from those that change C-terminal residues? Second, what genes encode proteins that interact either directly by physical interaction with Rad52p or indirectly by circumvention of the RAD52 recombinational repair pathway? Third, what role does Rad52p play in processing of double strand breaks made to initiate meiotic recombination? The experimental plan begins with a focus on cold sensitive rad52 mutations and extragenic suppressors of these mutations. Another set of experiments will utilize in vitro mutagenesis to explore the interaction between Rad52p and Rad51p, the protein product of another gene important in repair and recombination. Finally, rad52 mutants will be examined for their inability to process meiotic double strand breaks in recombinant chromosomes. The need of all organisms to defend themselves from natural radiation and to properly disjoin their chromosomes during gamete production has been evidenced by the recent discovery of RAD52 genes in may other organisms including humans.
