Rad52 recruitment is DNA replication independent and regulated by Cdc28 and the Mec1 kinase.
Recruitment of the homologous recombination machinery to sites of double-strand breaks is a cell cycle-regulated event requiring entry into S phase and CDK1 activity. Here, we demonstrate that the central recombination protein, Rad52, forms foci independent of DNA replication, and its recruitment requires B-type cyclin/CDK1 activity. Induction of the intra-S-phase checkpoint by hydroxyurea (HU) inhibits Rad52 focus formation in response to ionizing radiation. This inhibition is dependent upon Mec1/Tel1 kinase activity, as HU-treated cells form Rad52 foci in the presence of the PI3 kinase inhibitor caffeine. These Rad52 foci colocalize with foci formed by the replication clamp PCNA. These results indicate that Mec1 activity inhibits the recruitment of Rad52 to both sites of DNA damage and stalled replication forks during the intra-S-phase checkpoint. We propose that B-type cyclins promote the recruitment of Rad52 to sites of DNA damage, whereas Mec1 inhibits spurious recombination at stalled replication forks.
Barlow
Jacqueline H
JH
Department of Genetics & Development, Columbia University Medical Center, New York, NY 10032-2704, USA.
Rothstein
Rodney
R
D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism; D017489:CDC28 Protein Kinase, S cerevisiae / Q000235:genetics / Q000378:metabolism*; D002110:Caffeine / Q000378:metabolism; D002453:Cell Cycle / Q000502:physiology; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D064447:Checkpoint Kinase 2; D004249:DNA Damage; D004261:DNA Replication; D004791:Enzyme Inhibitors / Q000378:metabolism; D044783:F-Box Proteins / Q000235:genetics / Q000378:metabolism; D006918:Hydroxyurea / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D051721:Rad52 DNA Repair and Recombination Protein / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism
C080443:CDC4 protein, S cerevisiae; C066503:CDC6 protein, S cerevisiae; D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D044783:F-Box Proteins; D047908:Intracellular Signaling Peptides and Proteins; C497197:RAD52 protein, S cerevisiae; D051721:Rad52 DNA Repair and Recombination Protein; D029701:Saccharomyces cerevisiae Proteins; D002110:Caffeine; D044767:Ubiquitin-Protein Ligases; D064447:Checkpoint Kinase 2; C088937:MEC1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; D017489:CDC28 Protein Kinase, S cerevisiae; C067022:RAD53 protein, S cerevisiae; D006918:Hydroxyurea