CDC7-dependent protein kinase activity in yeast replicative-complex preparations.
A protein kinase activity was identified in preparations of DNA-replicative complex from the budding yeast Saccharomyces cerevisiae. The activity phosphorylated only a few of the endogenous proteins in the replicative fraction, and it displayed a marked preference for a 48-kDa polypeptide. Despite this relative specificity, the protein kinase activity was capable of utilizing exogenously added histone as substrate. The 48-kDa polypeptide was phosphorylated on serine residue(s) exclusively by the endogenous activity in the replicative-complex preparation. The activity was not stimulated by cAMP, cGMP, Ca2+/phosphatidylserine/diacylglycerol, or Ca2+/calmodulin. It did not utilize Ca2+ or Zn2+ in the place of Mg2+, and Mn2+ was only 22% as effective in fulfilling the divalent-cation requirement. Most importantly, the protein kinase activity was heat-sensitive in replicative fractions from the cell division cycle 7 (cdc7) mutant, which arrests at or close to the G1/S boundary of the cell cycle at restrictive temperature. Thus, the activity is CDC7-dependent. An effect of heat treatment on replicating activity in the replicative fraction from cdc7 cells was also found. This result and the finding that the protein kinase activity copurified with replicating activity in the preparations suggest that the CDC7 gene product and the protein kinase activity, whether or not they are the same entity, interact with yeast replicative complex. All of these results raise the possibility that phosphorylation of components of the replication machinery may play a role in the control of initiation of DNA replication during the cell cycle. It is possible that the phosphorylation observed is part of a protein kinase cascade that regulates progress through the G1 phase of the cell cycle.
Jazwinski
S M
SM
Department of Biochemistry and Molecular Biology, Louisiana State University Medical Center, New Orleans 70112.
D004261:DNA Replication; D005656:Fungal Proteins / Q000378:metabolism*; D007399:Interphase; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*
D005656:Fungal Proteins; D011494:Protein Kinases