Abstract
The contribution of the actin cytoskeleton to the unique architecture of the Golgi complex is manifold. An important player in this process is Coronin7 (CRN7), a Golgi-resident protein that stabilizes F-actin assembly at the trans-Golgi network (TGN) thereby facilitating anterograde trafficking. Here, we establish that CRN7-mediated association of F-actin with the Golgi apparatus is distinctly modulated via the small Rho GTPase Cdc42 and N-WASP. We identify N-WASP as a novel interaction partner of CRN7 and demonstrate that CRN7 restricts spurious F-actin reorganizations by repressing N-WASP ‘hyperactivity’ upon constitutive Cdc42 activation. Loss of CRN7 leads to increased cellular F-actin content and causes a concomitant disruption of the Golgi structure. CRN7 harbours a Cdc42- and Rac-interactive binding (CRIB) motif in its tandem β-propellers and binds selectively to GDP-bound Cdc42N17 mutant. We speculate that CRN7 can act as a cofactor for active Cdc42 generation. Mutation of CRIB motif residues that abrogate Cdc42 binding to CRN7 also fail to rescue the cellular defects in fibroblasts derived from CRN7 KO mice. Cdc42N17 overexpression partially rescued the KO phenotypes whereas N-WASP overexpression failed to do so. We conclude that CRN7 spatiotemporally influences F-actin organization and Golgi integrity in a Cdc42- and N-WASP-dependent manner.
Highlights
The actin cytoskeleton is an indispensable machinery of all eukaryotic cells
We propose a signalling mechanism by which CRN7 influences F-actin organization and thereby provides structure to the Golgi apparatus in a Cdc42- and neural WASP (N-WASP)-dependent manner
The Golgi apparatus in interphase cells is often present as a compact ribbon-like structure that is localised in a juxtanuclear, pericentriolar position
Summary
The actin cytoskeleton is an indispensable machinery of all eukaryotic cells. In the secretory pathway, actin filaments and actin binding proteins (ABPs) are required in organizing the Golgi complex[1]. Cdc[42] GTPase cooperates in actin assembly by inducing filopodia formation and an active Golgi pool of Cdc[42] regulates cell polarity and protein trafficking[3,4,5,6,7,8]. Cdc[42] funnels several signals through GTP-dependent binding to effector proteins containing a short stretch of ~15 amino acids referred to as Cdc42/Rac-interactive-binding (CRIB) motif. Proteins with partially conserved CRIB motifs can interact with Rho GTPases in a GTP- or GDP-dependent manner[11,12]. The activity of N-WASP on Arp2/3 complex is auto-inhibited by an intra-molecular interaction between its B-GBD/CRIB (Basic motif-GTPase binding domain) and VCA (verprolin homology, cofilin homology, and acidic region) domain, which is released upon GTP-Cdc[42] binding to the CRIB domain[23]. We propose a signalling mechanism by which CRN7 influences F-actin organization and thereby provides structure to the Golgi apparatus in a Cdc42- and N-WASP-dependent manner
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