Yeast formins regulate cell polarity by controlling the assembly of actin cables

Abstract

Formins are conserved Rho-GTPase effectors that communicate Rho-GTPase signals to the cytoskeleton. We found that formins were required for the assembly of one of the three budding yeast actin structures: polarized arrays of actin cables. A dominant-active formin induced the assembly of actin cables. The activation and localization of the formin Bni1p required components of the polarisome complex. These findings potentially define the cellular function of formins in budding yeast and explain their involvement in the generation of cell polarity. A requirement for formins in constructing specific actin structures might be the basis for the diverse activities of formins in development.

Stable and Dynamic Axes of Polarity Use Distinct Formin Isoforms in Budding Yeast

“…Assembly of these cables depends on two formin homologues, Bni1p and Bnr1p (Evangelista et al, 2002;Sagot et al, 2002a), members of a family of cytoskeletal regulatory proteins defined by conserved formin homology (FH)1 and FH2 domains (for reviews, see Evangelista et al, 2003;Wallar and Alberts, 2003). In vitro, the FH2 domain nucleates actin filaments from monomers Sagot et al, 2002b;Kovar et al, 2003;Li and Higgs, 2003;Harris et al, 2004;Kobielak et al, 2004), but distinct from other nucleators, it remains associated with the growing barbed end to promote elongation, even in the presence of barbed-end capping proteins Kovar et al, 2003;Li and Higgs, 2003;Pring et al, 2003;Harris et al, 2004;Kobielak et al, 2004;Moseley et al, 2004).…”

“…The FH1 region is a proline-rich stretch that recruits profilin, an actin-monomer binding protein essential for formin function in vivo , to participate in FH2-mediated nucleation in vitro (Sagot et al, 2002b;Kovar et al, 2003;Pring et al, 2003). Other regions present in Bni1p and Bnr1p are a regulatory NH 2 -terminal rho-GTPase-binding domain that subjects many formins to rho-dependent activation (Kohno et al, 1996;Evangelista et al, 1997;Imamura et al, 1997;Dong et al, 2003), and an FH3 motif that helps localize formins within the cell (Petersen et al, 1998).With loss of formin function, actin cables disassemble in 2 min, and a cytokinetic ring is unable to assemble, but actin patches remain intact Sagot et al, 2002a;Tolliday et al, 2002). Other actin nucleators, particularly the Arp2/3 complex, play no apparent role in cable or cytokinetic ring assembly in budding yeast (Winter et al, 1999;Evangelista et al, 2002;Tolliday et al, 2002), suggesting the formins might be in vivo nucleators for these actin filaments.…”

“…We wished to examine whether these localizations impart differences in the distribution of the actin filaments assembled by the two isoforms. However, overexpression of either formin alters actin organization and cell morphology (Kamei et al, 1998;Evangelista et al, 2002;Sagot et al, 2002a). To avoid this complication, we observed the distribution of the formins and formin-dependent actin filaments in cells that expressed Bni1p and Bnr1p at endogenous levels.…”

“…A COOHterminal GFP-fusion of full-length Bnr1p expressed from the BNR1 locus also displayed a localization in agreement with previous overexpression studies ( Kamei et al, 1998), showing a polarized distribution in very few unbudded cells (5% of unbudded cells), but a ring of fluorescence around the neck of cells with buds of all sizes ( Figures 1C and S1A). In yeast, formin-dependent actin filaments can be distinguished from the Arp2/3 complex-dependent filaments by their association with the F-actin binding protein tropomyosin, encoded by the TPM1 and TPM2 genes (Liu and Bretscher, 1989;Pruyne et al, 1998;Evangelista et al, 2002;Sagot et al, 2002a;Tolliday et al, 2002). To determine whether Bni1p or Bnr1p make distinct contributions to assembling these filaments, we examined Tpm1p distribution in wild-type yeast and strains lacking Bni1p (bni1⌬/bni1⌬) or Bnr1p (bnr1⌬/bnr1⌬) (Figure 1, C and D).…”

Actin bundling in plants

“…In contrast, cells of uninfected root tissue predominantly exhibit longitudinal AF arrays . The potential ability of AtFH6 to trigger a cytoskeletal reorganization was assessed by functional complementation of a yeast mutant deficient for the BNI1p and BNR1p formins, which both control the assembly of yeast actin bundles [Evangelista et al, 2002;Sagot et al, 2002]. Based on its ability to rescue the bin1Dbnr1D yeast mutant phenotype and on its localization at the plasma membrane, AtFH6 was proposed to be, at least partially, responsible for the assembly of the cortical actin bundles required for vesicle trafficking during the extensive plasma membrane and cell wall biogenesis [Favery et al, 2004].…”