FUNCTIONAL-CHARACTERIZATION OF THE CDC42P BINDING DOMAIN OF YEAST STE20P PROTEIN-KINASE

Ste20p from Saccharomyces cerevisiae belongs to the Ste20p/p65(PAK) fa mily of protein kinases which are highly conserved from yeast to man a nd regulate conserved mitogen-activated protein kinase pathways. Ste20 p fulfills multiple roles in pheromone signaling, morphological switch ing and vegetative growth and binds Cdc42p, a Rho-like small GTP bindi ng protein required for polarized morphogenesis. We have analyzed the functional consequences of mutations that prevent binding of Cdc42p to Ste20p. The complete amino-terminal, non-catalytic half of Ste20p, in cluding the conserved Cdc42p binding domain, was dispensable for heter otrimeric G-protein-mediated pheromone signaling. However, the Cdc42p binding domain was necessary for filamentous growth in response to nit rogen starvation and for an essential function that Ste20p shares with its isoform Cla4p during vegetative growth. Moreover, the Cdc42p bind ing domain was required for cell-cell adhesion during conjugation. Sub cellular localization of wild-type and mutant Ste20p fused to green fl uorescent protein showed that the Cdc42p binding domain is needed to d irect localization of Ste20p to regions of polarized growth. These res ults suggest that Ste20p is regulated in different developmental pathw ays by different mechanisms which involve heterotrimeric and small GTP binding proteins.