Proteins involved in sterol synthesis interact with Ste20 and regulate cell polarity

The Saccharomyces cerevisiae p21- activated kinase ( PAK) Ste20 regulates various aspects of cell polarity during vegetative growth, mating and filamentous growth. To gain further insight into the mechanisms of Ste20 action, we screened for interactors of Ste20 using the split- ubiquitin system. Among the identified proteins were Erg4, Cbr1 and Ncp1, which are all involved in sterol biosynthesis. The interaction between Ste20 and Erg4, as well as between Ste20 and Cbr1, was confirmed by pull- down experiments. Deletion of either ERG4 or NCP1 resulted in various polarity defects, indicating a role for these proteins in bud site selection, apical bud growth, cell wall assembly, mating and invasive growth. Interestingly, Erg4 was required for the polarized localization of Ste20 during mating. Lack of CBR1 produced no detectable phenotype, whereas the deletion of CBR1 in the absence of NCP1 was lethal. Using a conditional lethal mutant we demonstrate that both proteins have overlapping functions in bud morphology.