Cell migration: GAPs between membrane traffic and the cytoskeleton

Abstract

During cell migration, coordination between membrane traffic, cell substrate adhesion and actin reorganization is required for protrusive activity to occur at the leading edge. Actin organization is regulated by Rho family GTPases and, with a contribution from the endocytic cycle, serves to extend the cell front. The details of the molecular mechanisms that direct membrane traffic at sites of adhesion and rearrange actin at the cell edge are still unknown. However, recent findings show that a number of multi-domain proteins characterized by an ArfGAP domain interact with both actin-regulating and integrin-binding proteins, as well as affecting Rac-mediated protrusive activity and cell migration. Some of these proteins have been shown to localize to endocytic compartments and to have a role in regulating endocytosis. Given the participation of Arf proteins in regulating membrane traffic, one appealing hypothesis is that the ArfGAPs act as molecular devices that coordinate membrane traffic and cytoskeletal reorganization during cell motility.

Fig. 1. Schematic representation of three recently identified groups of multi-domain proteins characterized by the presence of an ArfGAP domain (GAP) and by ankyrin repeats (Ank). (A) ASAP1 and two PAP proteins are characterized by the presence of a pleckstrin homology domain (PH) and a C-terminal Src homology type 3 (SH3) domain. (B) ACAP proteins, include a PH domain. (C) Four members of the GIT family are characterized by a Spa2 homology 1 domain (SHD1) (Sheu et al.,1998), and a C-terminal paxillin binding subdomain (PBS). Proteins in (A) and (B) are members of the centaurin family, which is characterized by ArfGAP domain, ankyrin repeats and PH domain.

Fig. 2. Model for the intermolecular interactions and functional connections proposed for members of the GIT family (Figure 1C). Double-ended arrows point to known direct intermolecular interactions; single-ended arrows indicate functional connections. P, Proline-rich region; crib, Cdc42/Rac interactive binding motif; SH3, Src homology type 3; DH, Dbl homology; PH, pleckstrin homology; GAP, ArfGAP domain; Ank, ankyrin repeat; SHD1, Spa2 homology domain type 1; PBS, paxillin binding subdomain; p85/PI3K, 85 kD regulatory subunit of the phosphatidylinositol-3 kinase; ArfGAP = multi-domain ArfGAP protein. Members of the GIT family of ArfGAP proteins (indicated as ArfGAP) can stably interact with PIX and PAK (Bagrodia et al., 1999; Turner et al., 1999; Di Cesare et al., 2000), which mediate the interaction of the complex with active Rac and Cdc42 at the membrane. The complex may regulate actin remodelling at the cell surface by controlling Rac/Cdc42 activity via PAK and PIX. The interaction of ArfGAP proteins with paxillin (Turner et al., 1990, 1999) and FAK (Zhao et al., 2000b) functionally links the complex to integrin-mediated adhesion. Finally, the ArfGAPs are localized to the endosomal compartment by both PIX-dependent and PIX-independent mechanisms.

Fig. 3. Proposed model for the role of multi-domain ArfGAPs of the GIT family during cell migration. The ArfGAP protein is able to colocalize with Arf6 in the recycling endosomal compartment via its ankyrin repeats and the interaction with PIX. There the internalized membranes converge before recycling. In the proposed model it is speculated that wild-type Arf proteins (including Arf6) and the GAP activity of the ArfGAP are required for the formation of recycling vesicles. Once formed, vesicles may be recruited to Rac-enriched sites of the leading edge of the migrating cell. By recruiting paxillin to the complex, ArfGAP proteins would induce the redistribution of paxillin away from established focal adhesions to the leading edge. Here, paxillin would contribute to the formation of membrane protrusions, by participating in the formation of focal complexes in which paxillin is required for the anchorage of the Rac-induced actin filaments to the sites of substrate adhesion. Regulatory mechanisms must exist to direct distinct pools of the complex to endosomal membranes or to the cell surface.

Ivan de Curtis