Rhp51-dependent recombination intermediates that do not generate checkpoint signal are accumulated in Schizosaccharomyces pombe rad60 and smc5/6 mutants after release from replication arrest.
The Schizosaccharomyces pombe rad60 gene is essential for cell growth and is involved in repairing DNA double-strand breaks. Rad60 physically interacts with and is functionally related to the structural maintenance of chromosomes 5 and 6 (SMC5/6) protein complex. In this study, we investigated the role of Rad60 in the recovery from the arrest of DNA replication induced by hydroxyurea (HU). rad60-1 mutant cells arrested mitosis normally when treated with HU. Significantly, Rad60 function is not required during HU arrest but is required on release. However, the mutant cells underwent aberrant mitosis accompanied by irregular segregation of chromosomes, and DNA replication was not completed, as revealed by pulsed-field gel electrophoresis. The deletion of rhp51 suppressed the aberrant mitosis of rad60-1 cells and caused mitotic arrest. These results suggest that Rhp51 and Rad60 are required for the restoration of a stalled or collapsed replication fork after release from the arrest of DNA replication by HU. The rad60-1 mutant was proficient in Rhp51 focus formation after release from the HU-induced arrest of DNA replication or DNA-damaging treatment. Furthermore, the lethality of a rad60-1 rqh1Delta double mutant was suppressed by the deletion of rhp51 or rhp57. These results suggest that Rad60 is required for recombination repair at a step downstream of Rhp51. We propose that Rhp51-dependent DNA structures that cannot activate the mitotic checkpoints accumulate in rad60-1 cells.
Miyabe
Izumi
I
Genome Dynamics Group, Research Institute for Microbial Disease, Osaka University, 3-1 Yamada-oka, Suita, Osaka 565-0871, Japan.
Morishita
Takashi
T
Hishida
Takashi
T
Yonei
Shuji
S
Shinagawa
Hideo
H
D000970:Antineoplastic Agents / Q000494:pharmacology; D002453:Cell Cycle / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics; D064447:Checkpoint Kinase 2; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000502:physiology*; D020090:Chromosome Segregation; D015825:Chromosomes, Fungal; D004265:DNA Helicases / Q000235:genetics; D004260:DNA Repair / Q000235:genetics*; D004261:DNA Replication / Q000187:drug effects / Q000235:genetics*; D004271:DNA, Fungal / Q000378:metabolism; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D017353:Gene Deletion; D006918:Hydroxyurea / Q000494:pharmacology; D008938:Mitosis / Q000235:genetics; D009154:Mutation; D009690:Nucleic Acid Conformation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics; D051135:Rad51 Recombinase / Q000235:genetics / Q000502:physiology*; D011995:Recombination, Genetic / Q000235:genetics*; D012568:Schizosaccharomyces / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics / Q000502:physiology*
D000970:Antineoplastic Agents; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D004271:DNA, Fungal; C084232:RHP51 protein, S pombe; C455501:Rad60 protein, S pombe; D029702:Schizosaccharomyces pombe Proteins; C486628:Smc5 protein, S pombe; C486630:smc6 protein, S pombe; D064447:Checkpoint Kinase 2; C578636:Cds1 protein, S pombe; D017346:Protein Serine-Threonine Kinases; D051135:Rad51 Recombinase; D004265:DNA Helicases; C106462:Rqh1 protein, S pombe; D006918:Hydroxyurea