Phosphorylation by Cak1 regulates the C-terminal domain kinase Ctk1 in Saccharomyces cerevisiae.
Ctk1 is a Saccharomyces cerevisiae cyclin-dependent protein kinase (CDK) that assembles with Ctk2 and Ctk3 to form an active protein kinase complex, CTDK-I. CTDK-I phosphorylates Ser2 within the RNA polymerase II C-terminal domain, an activity that is required for efficient transcriptional elongation and 3' RNA processing. Ctk1 contains a conserved T loop, which undergoes activating phosphorylation in other CDKs. We show that Ctk1 is phosphorylated on Thr-338 within the T loop. Mutation of this residue abolished Ctk1 kinase activity in vitro and resulted in a cold-sensitive phenotype. As with other yeast CDKs undergoing T-loop phosphorylation, Ctk1 phosphorylation on Thr-338 was dependent on the Cak1 protein kinase. Ctk1 isolated from cak1Delta cells was unphosphorylated and exhibited low protein kinase activity. Moreover, Cak1 directly phosphorylated Ctk1 in vitro. Unlike wild-type cells, cells expressing Ctk1(T338A) delayed growth at early stationary phase, did not show the increase in Ser2 phosphorylation that normally accompanies the transition from rapid growth to stationary phase, and had compromised transcriptional activation of two stationary-phase genes, CTT1 and SPI1. Therefore, Ctk1 phosphorylation on Thr-338 is carried out by Cak1 and is required for normal gene transcription during the transition into stationary phase.
Ostapenko
Denis
D
Yale University School of Medicine, Department of Molecular Biophysics and Biochemistry, 333 Cedar Street, New Haven, CT 06520-8024, USA.
Solomon
Mark J
MJ
D000595:Amino Acid Sequence; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D004789:Enzyme Activation; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal / Q000235:genetics; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism*; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D013912:Threonine / Q000378:metabolism; D015533:Transcriptional Activation / Q000235:genetics; D000095043:Cyclin-Dependent Kinase-Activating Kinase
C118305:CTDK-I protein complex, S cerevisiae; D012331:RNA, Fungal; D029701:Saccharomyces cerevisiae Proteins; D013912:Threonine; D011494:Protein Kinases; D018844:Cyclin-Dependent Kinases; D012319:RNA Polymerase II; D000095043:Cyclin-Dependent Kinase-Activating Kinase