Interaction with Rad51 is indispensable for recombination mediator function of Rad52.
In the yeast Saccharomyces cerevisiae, the RAD52 gene is indispensable for homologous recombination and DNA repair. Rad52 protein binds DNA, anneals complementary ssDNA strands, and self-associates to form multimeric complexes. Moreover, Rad52 physically interacts with the Rad51 recombinase and serves as a mediator in the Rad51-catalyzed DNA strand exchange reaction. Here, we examine the functional significance of the Rad51/Rad52 interaction. Through a series of deletions, we have identified residues 409-420 of Rad52 as being indispensable and likely sufficient for its interaction with Rad51. We have constructed a four-amino acid deletion mutation within this region of Rad52 to ablate its interaction with Rad51. We show that the rad52delta409-412 mutant protein is defective in the mediator function in vitro even though none of the other Rad52 activities, namely, DNA binding, ssDNA annealing, and protein oligomerization, are affected. We also show that the sensitivity of the rad52delta409-412 mutant to ionizing radiation can be complemented by overexpression of Rad51. These results thus demonstrate the significance of the Rad51-Rad52 interaction in homologous recombination.
Krejci
Lumir
L
Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, San Antonio, Texas 78245-3207, USA. krejci@uthscsa.edu
Song
Binwei
B
Bussen
Wendy
W
Rothstein
Rodney
R
Mortensen
Uffe H
UH
Sung
Patrick
P
D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D004307:Dose-Response Relationship, Radiation; D004591:Electrophoresis, Polyacrylamide Gel; D005720:Gamma Rays; D017353:Gene Deletion; D005982:Glutathione Transferase / Q000378:metabolism; D007716:Kluyveromyces / Q000378:metabolism; D008938:Mitosis; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D051135:Rad51 Recombinase; D051721:Rad52 DNA Repair and Recombination Protein; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011995:Recombination, Genetic; D029701:Saccharomyces cerevisiae Proteins; D013997:Time Factors
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C497197:RAD52 protein, S cerevisiae; D051721:Rad52 DNA Repair and Recombination Protein; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D005982:Glutathione Transferase; D051135:Rad51 Recombinase