Premium
Ordered assembly of Sld3, GINS and Cdc45 is distinctly regulated by DDK and CDK for activation of replication origins
Author(s) -
Yabuuchi Hayato,
Yamada Yoshiki,
Uchida Tomonori,
Sunathvanichkul Tul,
Nakagawa Takuro,
Masukata Hisao
Publication year - 2006
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1038/sj.emboj.7601347
Subject(s) - biology , origin recognition complex , pre replication complex , replication factor c , control of chromosome duplication , origin of replication , dna replication , microbiology and biotechnology , cyclin dependent kinase , eukaryotic dna replication , licensing factor , dna replication factor cdt1 , genetics , dna , cell cycle , gene
Initiation of chromosome DNA replication in eukaryotes is tightly regulated through assembly of replication factors at replication origins. Here, we investigated dependence of the assembly of the initiation complex on particular factors using temperature‐sensitive fission yeast mutants. The psf3‐1 mutant, a GINS component mutant, arrested with unreplicated DNA at the restrictive temperature and the DNA content gradually increased, suggesting a defect in DNA replication. The mutation impaired GINS complex formation, as shown by pull‐down experiments. Chromatin immunoprecipitation assays indicated that GINS integrity was required for origin loading of Psf2, Cut5 and Cdc45, but not Sld3. In contrast, loading of Psf2 onto origins depended on Sld3 and Cut5 but not on Cdc45. These results suggest that Sld3 functions furthest upstream in initiation complex assembly, followed by GINS and Cut5, then Cdc45. Consistent with this conclusion, Cdc7‐Dbf4 kinase (DDK) but not cyclin‐dependent kinase (CDK) was required for Sld3 loading, whereas recruitment of the other factors depended on both kinases. These results suggest that DDK and CDK regulate distinct steps in activation of replication origins in fission yeast.