Regulation of formins and cell polarity in yeast

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DESCRIPTION (provided by applicant): Our long-term goal is to gain a highly mechanistic understanding of how the actin cytoskeleton directs cell polarity and cell morphogenesis. All living cells have internal and external structures tailored to and critical for their distinctive physiological functions. Further, cell architecture can be changed rapidly in response to various cues. The mechanisms underlying these events remain poorly understood and represent a major challenge for cell biologists to define. Recently, a conserved family of proteins called formins has emerged as crucial regulators of actin assembly and remodeling in cells, often functioning directly downstream of Rho GTPases. Formins are large multi-domain proteins that play essential roles in cell polarity, cell division, cell migration, endocytosis, and cell adhesion in a wide range of organisms. Formins directly nucleate actin assembly by a novel mechanism and remain processively attached to the growing end of the filament, protecting the end from capping proteins while guiding insertion of new actin subunits. While the last five years have seen rapid progress in elucidating formin protein structure, mechanism and function, comparatively little is known about how formin activities are regulated spatially and temporally in cells. In this proposal, we will address this question using the budding yeast Saccharomyces cerevisiae as a model organism. Whereas mammals have 15 different formin genes, S. cerevisiae has only two (Bni1 and Bnr1), and hence offers a simplified model to dissect formin regulation. Yeast also allows us to take a multidisciplinary approach, combining genetics, biochemistry, and live cell imaging. Bni1 and Bnr1 have distinct localization and dynamics, and assemble two distinct sets of actin cables. These cables serve as polarized tracks required for targeted secretion and polarized cell growth. We will determine how one of these formins (Bnr1) is regulated in vivo, which will provide key mechanistic insights into the molecular basis of cell polarity and cell morphogenesis. The Specific Aims of the proposal are: (1) How is Bnr1 anchored at the bud neck, activated/released from an autoinhibited state, and then retrieved from actin filament ends for new rounds of actin assembly? (2) How are the activities and cellular functions of a novel Bnr1-regulator (Bud14) controlled by its in vivo binding partners (Kel1 and Kel2)? Defining the molecular basis of these events is critical not only for understanding normal human cell biology and physiology, but also for determining how mutations in the genes encoding morphogenetic determinants give rise to disease states including cancer, birth defects, and neurodegenerative disorders.