G2/M checkpoint genes of Saccharomyces cerevisiae: further evidence for roles in DNA replication and/or repair.
We have cloned, sequenced and disrupted the checkpoint genes RAD17, RAD24 and MEC3 of Saccharomyces cerevisiae. Mec3p shows no strong similarity to other proteins currently in the database. Rad17p is similar to Rec1 from Ustilago maydis, a 3' to 5' DNA exonuclease/checkpoint protein, and the checkpoint protein Rad1p from Schizosaccharomyces pombe (as we previously reported). Rad24p shows sequence similarity to replication factor C (RFC) subunits, and the S. pombe Rad17p checkpoint protein, suggesting it has a role in DNA replication and/or repair. This hypothesis is supported by our genetic experiments which show that overexpression of RAD24 strongly reduces the growth rate of yeast strains that are defective in the DNA replication/repair proteins Rfc1p (cdc44), DNA pol alpha (cdc17) and DNA pol delta (cdc2) but has much weaker effects on cdc6, cdc9, cdc15 and CDC4 strains. The idea that RAD24 overexpression induces DNA damage, perhaps by interfering with replication/repair complexes, is further supported by our observation that RAD24 overexpression increases mitotic chromosome recombination in CDC4 strains. Although RAD17, RAD24 and MEC3 are not required for cell cycle arrest when S phase is inhibited by hydroxyurea (HU), they do contribute to the viability of yeast cells grown in the presence of HU, possibly because they are required for the repair of HU-induced DNA damage. In addition, all three are required for the rapid death of cdc13 rad9 mutants. All our data are consistent with models in which RAD17, RAD24 and MEC3 are coordinately required for the activity of one or more DNA repair pathways that link DNA damage to cell cycle arrest.
Lydall
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Department of Molecular and Cellular Biology, University of Arizona, Tucson 85721, USA. Lydall@u.arizona.edu
Weinert
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D000477:Alkylating Agents / Q000494:pharmacology; D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000235:genetics / Q000502:physiology*; D003001:Cloning, Molecular; D004249:DNA Damage; D004260:DNA Repair / Q000235:genetics*; D004261:DNA Replication / Q000235:genetics*; D004271:DNA, Fungal / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins; D004843:Epistasis, Genetic; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000502:physiology*; D016195:G2 Phase / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal; D006918:Hydroxyurea / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D008677:Metaphase / Q000235:genetics*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D051818:Replication Protein C; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D000477:Alkylating Agents; D018797:Cell Cycle Proteins; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; C099938:MEC3 protein, S cerevisiae; D009687:Nuclear Proteins; C501395:RAD17 protein, S cerevisiae; C088385:RAD24 protein, S cerevisiae; C084903:RFC1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; C501114:rad24 protein, S pombe; D005090:Exodeoxyribonucleases; D051818:Replication Protein C; D006918:Hydroxyurea