The Cdc23 (Mcm10) protein is required for the phosphorylation of minichromosome maintenance complex by the Dfp1-Hsk1 kinase.
Previous studies in Saccharomyces cerevisiae have defined an essential role for the Dbf4-Cdc7 kinase complex in the initiation of DNA replication presumably by phosphorylation of target proteins, such as the minichromosome maintenance (Mcm) complex. We have examined the phosphorylation of the Mcm complex by the Dfp1-Hsk1 kinase, the Schizosaccharomyces pombe homologue of Dbf4-Cdc7. In vitro, the purified Dfp1-Hsk1 kinase efficiently phosphorylated Mcm2p. In contrast, Mcm2p, present in the six-subunit Mcm complex, was a poor substrate of this kinase and required Cdc23p (homologue of Mcm10p) for efficient phosphorylation. In the presence of Cdc23p, Dfp1-Hsk1 phosphorylated the Mcm2p and Mcm4p subunits of the Mcm complex. Cdc23p interacted with both the Mcm complex and Dfp1-Hsk1 by selectively binding to the Mcm467 subunits and Dfp1p, respectively. The N terminus of Cdc23p was found to interact directly with Dfp1-Hsk1 and was essential for phosphorylation of the Mcm complex. Truncated derivatives of Cdc23p that complemented the temperature-sensitive phenotype of cdc23 mutant cells also stimulated the phosphorylation of Mcm complex, implying that this activity might be a critical role of Cdc23p in vivo. These results suggest that Cdc23p participates in the activation of prereplicative complex by recruiting the Dfp1-Hsk1 kinase and stimulating the phosphorylation of the Mcm complex.
Lee
Joon-Kyu
JK
Program in Molecular Biology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Seo
Yeon-Soo
YS
Hurwitz
Jerard
J
D064173:Anaphase-Promoting Complex-Cyclosome; D064196:Apc8 Subunit, Anaphase-Promoting Complex-Cyclosome; D002262:Carboxy-Lyases / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism* / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D005816:Genetic Complementation Test; D026341:Minichromosome Maintenance 1 Protein / Q000378:metabolism; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000378:metabolism*; D010453:Peptide Synthases / Q000378:metabolism*; D010641:Phenotype; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism / Q000502:physiology*; D012568:Schizosaccharomyces / Q000201:enzymology / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors; D043743:Ubiquitin-Protein Ligase Complexes
D064196:Apc8 Subunit, Anaphase-Promoting Complex-Cyclosome; C577441:CDC23 protein, S cerevisiae; D018797:Cell Cycle Proteins; D026341:Minichromosome Maintenance 1 Protein; D009097:Multienzyme Complexes; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; C094761:HSK1 protein, S pombe; D017346:Protein Serine-Threonine Kinases; D002262:Carboxy-Lyases; D010453:Peptide Synthases