Stephanie Brody

Stephanie BrodyBrandeis / Biology and Mathematics
Hosted by Goode's Lab

“Role of the Bud14 Formin Effector Elongation Domain in polarized growth and cell morphology”

Stephanie Brody, Alison Wirshing, and Bruce Goode


Formins assemble diverse linear actin networks that drive cell motility, cytokinesis and intracellular transport. This family of highly conserved actin assembly proteins is characterized by domains that work in concert to rapidly build actin networks by first nucleating and then processivly elongating actin filaments. Purified formins in vitro produce long (>50 μm) actin filaments. However, most in vivo actin structures are composed of relatively short (<1 μm) filaments, highlighting the importance of regulating formin activity to produce cellular actin structures of a precise length and architecture.

Here we use actin cables in budding yeast as a model system to investigate mechanisms of formin regulation in vivo. Actin cables are linear actin bundles that are assembled by the two yeast formins, Bnr1, and Bni1. These cables serve as tracks for myosin-based transport of vesicles and organelles during polarized growth. In 2009, the Goode Lab identified Bud14 as a binding partner and inhibitor of the formin Bnr1 (with no effect on Bni1). Loss of Bud14 results in overgrown and tortuous actin cables that are defective in vesicle transport. Additionally, Bud14 regulates bud morphology and functions as a regulatory subunit of the PP1/Glc7 phosphatase to regulate dynein activity. To investigate if the roles of Bud14 in formin inhibition and bud morphology regulation are separable or integrated we used CRISPR to target the Bud14 Formin Effector Elongation Domain (FEED). The FEED motif is conserved in formin binding and regulatory proteins and is predicted to mediate the interaction between Bud14 and the FH2 domain of Bnr1.

We find that disruption of the FEED motif does not alter the expression or dynamics of Bud14 bud does disrupt Bud14 regulation of Bnr1. Cells expressing Bud14(FEED*) have synthetic growth defects in a genetic background where Bnr1 is the only formin. Interestingly, these cells have WT bud morphology indicating that Bud14(FEED*) is a separation of function mutant that disrupts formin regulation but leaves other functions of Bud14 intact.

With this tool in hand we are next characterizing the effects of FEED mutation on actin cable dynamics and architecture. This work will add to our understanding of the cellular mechanisms tuning formin activity to build specific actin networks.

Support: SMURF (Summer MRSEC Undergrad Research Fellowship)