Septin organization is regulated by the Gpa1 ubiquitination domain and endocytic machinery during the yeast pheromone response.

Abstract

Saccharomyces cerevisiae detect and respond to mating pheromone using a G-protein-coupled receptor signaling pathway (GPCR) to initiate polarized growth toward mating partners. Septins form structures at the base of the mating projection to control morphogenesis in a manner that is dependent upon desensitization of the large G-protein Gpa1. We sought to identify the pathway through which Gpa1 regulates septin organization using gene deletions in the presence of a hyperactive Gpa1 mutant, live cell imaging, and computational approaches. We found that the Cdc42 effector Gic1, the Cdc42 GAP Bem3, and the endocytic proteins Ent1, Ent2, and End3 are all involved in Gpa1 organization of septins. Mathematical modeling indicates that changes in the location of endocytic events could be responsible for the observed septin localization in the varied genetic backgrounds. Both the GPCR Ste2 and Gpa1 undergo endocytosis. We found that while the Ste2 c-terminus impacts septin organization in the presence of WT Gpa1, hyperactive Gpa1 drives septin organization through its ubiquitin domain that is required for its endocytosis. These data support a model where cargo recruitment of epsins (Ent1/2) and Cdc42 GAPs couples endocytosis with septin organization.