Role of the Rad52 amino-terminal DNA binding activity in DNA strand capture in homologous recombination.
Saccharomyces cerevisiae Rad52 protein promotes homologous recombination by nucleating the Rad51 recombinase onto replication protein A-coated single-stranded DNA strands and also by directly annealing such strands. We show that the purified rad52-R70A mutant protein, with a compromised amino-terminal DNA binding domain, is capable of Rad51 delivery to DNA but is deficient in DNA annealing. Results from chromatin immunoprecipitation experiments find that rad52-R70A associates with DNA double-strand breaks and promotes recruitment of Rad51 as efficiently as wild-type Rad52. Analysis of gene conversion intermediates reveals that rad52-R70A cells can mediate DNA strand invasion but are unable to complete the recombination event. These results provide evidence that DNA binding by the evolutionarily conserved amino terminus of Rad52 is needed for the capture of the second DNA end during homologous recombination.
Shi
Idina
I
Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Hallwyl
Swee C L
SC
Seong
Changhyun
C
Mortensen
Uffe
U
Rothstein
Rodney
R
Sung
Patrick
P
D019943:Amino Acid Substitution; D001665:Binding Sites; D047369:Chromatin Immunoprecipitation; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004277:DNA, Single-Stranded; D004591:Electrophoresis, Polyacrylamide Gel; D008854:Microscopy, Electron; D009154:Mutation; D011485:Protein Binding; D051135:Rad51 Recombinase / Q000235:genetics / Q000378:metabolism; D051721:Rad52 DNA Repair and Recombination Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D004277:DNA, Single-Stranded; D051721:Rad52 DNA Repair and Recombination Protein; D029701:Saccharomyces cerevisiae Proteins; D051135:Rad51 Recombinase