2 ACTIVE STATES OF THE RAS-RELATED BUD1 RSR1 PROTEIN BIND TO DIFFERENT EFFECTORS TO DETERMINE YEAST-CELL POLARITY/

Cells of budding yeast organize their cytoskeleton in a highly polariz ed manner during vegetative growth, Selection of a site for polarizati on requires a group of proteins including a Ras-like GTPase, Bud1, and its regulators. Another group of proteins, which includes a Rho-like GTPase (Cdc42), its guanine nucleotide exchange factor (Cdc24), and Be m1, is necessary for organization of the actin cytoskeleton and for ce ll polarization. We have proposed previously that the Bud1 protein, th rough its GTPase cycle, determines the localization of one or more of the cell polarity proteins to the bud site. Herein we demonstrate that Bud1 directly interacts with Cdc24 and Bem1: Bud1 in its GTP-bound fo rm associates preferentially with Cdc24, whereas the GDP-bound form of Bud1 associates with Bem1. We also present subcellular fractionation data for Bud1 that is consistent with the idea that Bud1 can travel be tween the site for budding on the plasma membrane and the cytosol. We propose that Budl can exist in two active states for association with different partners and that the switch from Bud1-GTP to Bud1-GDP provi des a regulatory device for ordered assembly of a macromolecular compl ex at the bud site.