Regulation of initiation of S phase, replication checkpoint signaling, and maintenance of mitotic chromosome structures during S phase by Hsk1 kinase in the fission yeast.
Hsk1, Saccharomyces cerevisiae Cdc7-related kinase in Shizosaccharomyces pombe, is required for G1/S transition and its kinase activity is controlled by the regulatory subunit Dfp1/Him1. Analyses of a newly isolated temperature-sensitive mutant, hsk1-89, reveal that Hsk1 plays crucial roles in DNA replication checkpoint signaling and maintenance of proper chromatin structures during mitotic S phase through regulating the functions of Rad3 (ATM)-Cds1 and Rad21 (cohesin), respectively, in addition to expected essential roles for initiation of mitotic DNA replication through phosphorylating Cdc19 (Mcm2). Checkpoint defect in hsk1-89 is indicated by accumulation of cut cells at 30 degrees C. hsk1-89 displays synthetic lethality in combination with rad3 deletion, indicating that survival of hsk1-89 depends on Rad3-dependent checkpoint pathway. Cds1 kinase activation, which normally occurs in response to early S phase arrest by nucleotide deprivation, is largely impaired in hsk1-89. Furthermore, Cds1-dependent hyperphosphorylation of Dfp1 in response to hydroxyurea arrest is eliminated in hsk1-89, suggesting that sufficient activation of Hsk1-Dfp1 kinase is required for S phase entry and replication checkpoint signaling. hsk1-89 displays apparent defect in mitosis at 37 degrees C leading to accumulation of cells with near 2C DNA content and with aberrant nuclear structures. These phenotypes are similar to those of rad21-K1 and are significantly enhanced in a hsk1-89 rad21-K1 double mutant. Consistent with essential roles of Rad21 as a component for the cohesin complex, sister chromatid cohesion is partially impaired in hsk1-89, suggesting a possibility that infrequent origin firing of the mutant may affect the cohesin functions during S phase.
Takeda
T
T
Departments of Molecular and Developmental Biology, Institute of Medical Science, University of Tokyo, Tokyo 108-8639, Japan. ttakeda@ims.u-tokyo.ac.jp
Ogino
K
K
Tatebayashi
K
K
Ikeda
H
H
Arai Ki
Masai
H
H
D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D015825:Chromosomes, Fungal / Q000378:metabolism*; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication / Q000187:drug effects; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D017930:Genes, Reporter; D018816:Genes, cdc / Q000502:physiology; D005816:Genetic Complementation Test; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology; D015151:Immunoblotting; D008938:Mitosis / Q000235:genetics / Q000502:physiology; D009154:Mutation / Q000235:genetics; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases; D016196:S Phase / Q000235:genetics / Q000502:physiology*; D012568:Schizosaccharomyces / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D029702:Schizosaccharomyces pombe Proteins; D013696:Temperature; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D029702:Schizosaccharomyces pombe Proteins; D015534:Trans-Activators; C045264:replication initiator protein; C094761:HSK1 protein, S pombe; D017346:Protein Serine-Threonine Kinases; D004265:DNA Helicases; C085710:mcm2 protein, S pombe; D006918:Hydroxyurea