Recruitment of the recombinational repair machinery to a DNA double-strand break in yeast.
Repair of DNA double-strand breaks (DSBs) by homologous recombination requires members of the RAD52 epistasis group. Here we use chromatin immunoprecipitation (ChIP) to examine the temporal order of recruitment of Rad51p, Rad52p, Rad54p, Rad55p, and RPA to a single, induced DSB in yeast. Our results suggest a sequential, interdependent assembly of Rad proteins adjacent to the DSB initiated by binding of Rad51p. ChIP time courses from various mutant strains and additional biochemical studies suggest that Rad52p, Rad55p, and Rad54p each help promote the formation and/or stabilization of the Rad51p nucleoprotein filament. We also find that all four Rad proteins associate with homologous donor sequences during strand invasion. These studies provide a near comprehensive view of the molecular events required for the in vivo assembly of a functional Rad51p presynaptic filament.
Wolner
Branden
B
Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA.
van Komen
Stephen
S
Sung
Patrick
P
Peterson
Craig L
CL
D002478:Cells, Cultured; D004247:DNA / Q000235:genetics*; D004249:DNA Damage / Q000235:genetics*; D004265:DNA Helicases; D004260:DNA Repair / Q000235:genetics*; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins / Q000235:genetics*; D051135:Rad51 Recombinase; D051721:Rad52 DNA Repair and Recombination Protein; D011995:Recombination, Genetic / Q000235:genetics*; D051716:Replication Protein A; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D015003:Yeasts / Q000235:genetics* / Q000378:metabolism
D004268:DNA-Binding Proteins; C497197:RAD52 protein, S cerevisiae; C087191:RAD55 protein, S cerevisiae; D051721:Rad52 DNA Repair and Recombination Protein; D051716:Replication Protein A; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA; D051135:Rad51 Recombinase; C075468:RAD54 protein, S cerevisiae; D004265:DNA Helicases; D045643:DNA Repair Enzymes