CELL CYCLE-DEPENDENT AND CLN2P-CDC28P-DEPENDENT PHOSPHORYLATION OF THE YEAST STE20P PROTEIN-KINASE

Ste20p from Saccharomyces cerevisiae is a member of the Ste20/p21-acti vated protein kinase family of protein kinases. The Ste20p kinase is p ost-translationally modified by phosphorylation in a cell cycle-depend ent manner, as judged by the appearance of phosphatase-sensitive speci es with reduced mobility on SDS-polyacrylamide gel electrophoresis. Th is modification is maximal during S phase, and correlates with the acc umulation of Ste20p fused to green fluorescent protein at the site of bud emergence. Overexpression of Cln2p, but not Clb2p or Clb5p, causes a quantitative shift of Ste20p to the reduced mobility form, and this shift is dependent on Cdc28p activity. The post-translational mobilit y shift can be generated in vitro by incubation of Ste20p with immunop recipitated Cln2p kinase complexes, but not by immunoprecipitated Clb2 p or Clb5p kinase complexes. Ste20p is therefore a substrate for the C dc28p kinase, and undergoes a Cln2p-Cdc28p mediated mobility shift as cells initiate budding and DNA replication. In cells that express only the Cln2p G(1) cyclin, minor overexpression of Ste20p causes aberrant morphology, suggesting a proper coordination of Ste20p and Cln-Cdc28p activity may be required for the control of cell shape.