Characterization of Novel Molecular Mechanisms Favoring Rac1 Membrane Translocation.

Antonio Castro-Castro,Xosé R Bustelo,Miguel A Del Pozo,Olivia Muriel,Mirjam M Zegers

doi:10.1371/journal.pone.0166715

Abstract

The Rac1 GTPase plays key roles in cytoskeletal organization, cell motility and a variety of physiological and disease-linked responses. Wild type Rac1 signaling entails dissociation of the GTPase from cytosolic Rac1-Rho GDP dissociation inhibitor (GDI) complexes, translocation to membranes, activation by exchange factors, effector binding, and activation of downstream signaling cascades. Out of those steps, membrane translocation is the less understood. Using transfections of a expression cDNA library in cells expressing a Rac1 bioreporter, we previously identified a cytoskeletal feedback loop nucleated by the F-actin binding protein coronin 1A (Coro1A) that promotes Rac1 translocation to the plasma membrane by facilitating the Pak-dependent dissociation of Rac1-Rho GDI complexes. This screening identified other potential regulators of this process, including WDR26, basigin, and TMEM8A. Here, we show that WDR26 promotes Rac1 translocation following a Coro1A-like and Coro1A-dependent mechanism. By contrast, basigin and TMEM8A stabilize Rac1 at the plasma membrane by inhibiting the internalization of caveolin-rich membrane subdomains. This latter pathway is F-actin-dependent but Coro1A-, Pak- and Rho GDI-independent.

Highlights

Rac1 is a Rho GTPase subfamily member that plays key roles in biological processes such as cytoskeletal structure, cell motility, adhesion, axon guidance, and cell proliferation [1,2,3]
During a genome-wide functional screening previously described by us [7, 35], we identified four cDNAs clones that could trigger the translocation of an enhanced green fluorescent protein (EGFP)-Rac1 chimera from the cytosol to the plasma membrane when expressed in HEK293T cells
These clones encoded the β-propeller domain-containing proteins known as coronin 1A (Coro1A) [35] and WD repeat domain 26 (WDR26) as well as two membrane-localized proteins, basigin (Bsg) and transmembrane protein 8A (TMEM8A) (Fig 1A)

Summary

Introduction

Rac is a Rho GTPase subfamily member that plays key roles in biological processes such as cytoskeletal structure, cell motility, adhesion, axon guidance, and cell proliferation [1,2,3]. Rac remains sequestered in the cytosol in non-stimulated cells due to the formation of inhibitory complexes with Rho GDIs [7, 8]. Rac is released from those inhibitory complexes, moves to the plasma membrane, and undergoes exchange of GDP by GTP molecules to acquire full signaling competence. This latter step is favored by the catalytic

Methods

Results

Conclusion