Cak1 is required for Kin28 phosphorylation and activation in vivo.
Complete activation of most cyclin-dependent protein kinases (CDKs) requires phosphorylation by the CDK-activating kinase (CAK). In the budding yeast, Saccharomyces cerevisiae, the major CAK is a 44-kDa protein kinase known as Cak1. Cak1 is required for the phosphorylation and activation of Cdc28, a major CDK involved in cell cycle control. We addressed the possibility that Cak1 is also required for the activation of other yeast CDKs, such as Kin28, Pho85, and Srb10. We generated three new temperature-sensitive cak1 mutant strains, which arrested at the restrictive temperature with nonuniform budding morphology. All three cak1 mutants displayed significant synthetic interactions with loss-of-function mutations in CDC28 and KIN28. Loss of Cak1 function reduced the phosphorylation and activity of both Cdc28 and Kin28 but did not affect the activity of Pho85 or Srb10. In the presence of the Kin28 regulatory subunits Ccl1 and Tfb3, Kin28 was phosphorylated and activated when coexpressed with Cak1 in insect cells. We conclude that Cak1 is required for the activating phosphorylation of Kin28 as well as that of Cdc28.
Espinoza
F H
FH
Departments of Physiology and Biochemistry & Biophysics, University of California, San Francisco, California 94143-0444, USA.
Farrell
A
A
Nourse
J L
JL
Chamberlin
H M
HM
Gileadi
O
O
Morgan
D O
DO
D000818:Animals; D017489:CDC28 Protein Kinase, S cerevisiae / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000502:physiology*; D002453:Cell Cycle / Q000502:physiology; D002455:Cell Division / Q000235:genetics; D018844:Cyclin-Dependent Kinases / Q000502:physiology; D029721:Drosophila Proteins; D004789:Enzyme Activation / Q000502:physiology; D005656:Fungal Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010641:Phenotype; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins; D018411:Spodoptera / Q000235:genetics
D029721:Drosophila Proteins; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; D017346:Protein Serine-Threonine Kinases; C099190:CASK protein, Drosophila; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D017489:CDC28 Protein Kinase, S cerevisiae; D018844:Cyclin-Dependent Kinases; C084210:Kin28 protein kinase, S cerevisiae