Analysis of Unregulated Formin Activity Reveals How Yeast Can Balance F-Actin Assembly between Different Microfilament-based Organizations

Abstract

Formins are regulated actin-nucleating proteins that are widespread among eukaryotes. Overexpression of unregulated formins in budding yeast is lethal and causes a massive accumulation of disorganized cable-like filaments. To explore the basis of this lethality, a cDNA library was screened to identify proteins whose overexpression could rescue the lethality conferred by unregulated Bnr1p expression. Three classes of suppressors encoding actin-binding proteins were isolated. One class encodes proteins that promote the assembly of actin cables (TPM1, TPM2, and ABP140), suggesting that the lethality was rescued by turning disorganized filaments into functional cables. The second class encodes proteins that bind G-actin (COF1, SRV2, and PFY1), indicating that reduction of the pool of actin available for cable formation may also rescue lethality. Consistent with this, pharmacological or genetic reduction of available actin also protected the cell from overproduction of unregulated Bnr1p. The third class consists of Las17p, an activator of the formin-independent Arp2/3p-dependent actin nucleation pathway. These results indicate that proper assembly of actin cables is sensitive to the appropriate balance of their constituents and that input into one pathway for actin filament assembly can affect another. Thus, cells must have a way of ensuring a proper balance between actin assembly pathways.