Phosphorylation of Cdc28 and regulation of cell size by the protein kinase CKII in Saccharomyces cerevisiae.
The CDK (cyclin-dependent kinase) family of enzymes is required for the G(1)-to-S-phase and G(2)-to-M-phase transitions during the cell-division cycle of eukaryotes. We have shown previously that the protein kinase CKII catalyses the phosphorylation of Ser-39 in Cdc2 during the G(1) phase of the HeLa cell-division cycle [Russo, Vandenberg, Yu, Bae, Franza and Marshak (1992) J. Biol. Chem. 267, 20317-20325]. To identify a functional role for this phosphorylation, we have studied the homologous enzymes in the budding yeast Saccharomyces cerevisiae. The S. cerevisiae homologue of Cdc2, Cdc28, contains a consensus CKII site (Ser-46), which is homologous with that of human Cdc2. Using in vitro kinase assays, metabolic labelling, peptide mapping and phosphoamino acid analysis, we demonstrate that this site is phosphorylated in Cdc28 in vivo as well in vitro. In addition, S. cerevisiae cells in which Ser-46 has been mutated to alanine show a decrease in both cell volume and protein content of 33%, and this effect is most pronounced in the stationary phase. Because cell size in S. cerevisiae is regulated primarily at the G(1) stage, we suggest that CKII contributes to the regulation of the cell cycle in budding yeast by phosphorylation of Cdc28 as a checkpoint for G(1) progression.
Russo
G L
GL
Istituto di Scienze dell'Alimentazione, Consiglio Nazionale delle Ricerche, Avellino 83100, Italy. glrusso@isa.av.cnr.it
van den Bos
C
C
Sutton
A
A
Coccetti
P
P
Baroni
M D
MD
Alberghina
L
L
Marshak
D R
DR
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D015153:Blotting, Western; D017489:CDC28 Protein Kinase, S cerevisiae / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D047390:Casein Kinase II; D002455:Cell Division; D005434:Flow Cytometry; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010449:Peptide Mapping; D010766:Phosphorylation / Q000187:drug effects; D010768:Phosphoserine / Q000032:analysis / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D016415:Sequence Alignment
D010768:Phosphoserine; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D017489:CDC28 Protein Kinase, S cerevisiae