The role of the Saccharomyces cerevisiae Cdc7-Dbf4 complex in the replication checkpoint.
The Cdc7-Dbf4 complex is a conserved serine/threonine protein kinase essential for the initiation of eukaryotic DNA replication. Although an mcm5-bob1 mutation bypasses lethality conferred by mutations in CDC7 or DBF4, the Deltacdc7 mcm5-bob1 mutant is sensitive to hydroxyurea (HU), which induces replication stress. To elucidate the reasons for HU sensitivity conferred by deletion of CDC7, we examined the role of Cdc7-Dbf4 in the replication checkpoint. We found that in Cdc7-Dbf4-deficient cells exposed to replication stress, Rad53 remains in a hypophosphorylated form, anaphase spindle is elongated, and checkpoint-specific transcription is not induced. The hypophosphorylated Rad53 exhibits a low autophosphorylation activity, and recombinant Cdc7-Dbf4 phosphorylates Rad53 in vitro. These results suggest that Cdc7-Dbf4 is required for full activation of Rad53 in response to replication stress.
Ogi
Hiroo
H
Laboratories for Biomolecular Networks, Graduate School of Frontier Biosciences, Osaka University, Suita, Osaka, Japan. oogi@fbs.osaka-u.ac.jp
Wang
Cheng-Zhong
CZ
Nakai
Wataru
W
Kawasaki
Yasuo
Y
Masumoto
Hiroshi
H
D000705:Anaphase; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D064447:Checkpoint Kinase 2; D004261:DNA Replication; D004271:DNA, Fungal / Q000502:physiology*; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal; D006918:Hydroxyurea / Q000494:pharmacology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D016196:S Phase / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D001616:beta-Galactosidase / Q000378:metabolism
D018797:Cell Cycle Proteins; D004271:DNA, Fungal; C088096:Dbf4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C489404:CDC7 protein, S cerevisiae; D064447:Checkpoint Kinase 2; D017346:Protein Serine-Threonine Kinases; C067022:RAD53 protein, S cerevisiae; D001616:beta-Galactosidase; D006918:Hydroxyurea