Cell cycle control of Cdc7p kinase activity through regulation of Dbf4p stability.
In Saccharomyces cerevisiae, the heteromeric kinase complex Cdc7p-Dbf4p plays a pivotal role at replication origins in triggering the initiation of DNA replication during the S phase. We have assayed the kinase activity of endogenous levels of Cdc7p kinase by using a likely physiological target, Mcm2p, as a substrate. Using this assay, we have confirmed that Cdc7p kinase activity fluctuates during the cell cycle; it is low in the G1 phase, rises as cells enter the S phase, and remains high until cells complete mitosis. These changes in kinase activity cannot be accounted for by changes in the levels of the catalytic subunit Cdc7p, as these levels are constant during the cell cycle. However, the fluctuations in kinase activity do correlate with levels of the regulatory subunit Dbf4p. The regulation of Dbf4p levels can be attributed in part to increased degradation of the protein in G1 cells. This G1-phase instability is cdc16 dependent, suggesting a role of the anaphase-promoting complex in the turnover of Dbf4p. Overexpression of Dbf4p in the G1 phase can partially overcome this elevated turnover and lead to an increase in Cdc7p kinase activity. Thus, the regulation of Dbf4p levels through the control of Dbf4p degradation has an important role in the regulation of Cdc7p kinase activity during the cell cycle.
Oshiro
G
G
Molecular Biology Program, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA.
Owens
J C
JC
Shellman
Y
Y
Sclafani
R A
RA
Li
J J
JJ
D064194:Apc6 Subunit, Anaphase-Promoting Complex-Cyclosome; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity
D064194:Apc6 Subunit, Anaphase-Promoting Complex-Cyclosome; C577397:CDC16 protein, S cerevisiae; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; C088096:Dbf4 protein, S cerevisiae; D005656:Fungal Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C489404:CDC7 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; C069199:MCM2 protein, S cerevisiae