Regulation of actin cytoskeleton dynamics by Arf-family GTPases

Abstract

Small GTPases of the Arf family are best known for their role in vesicular transport, wherein they nucleate the assembly of coat proteins at sites of carrier vesicle formation. However, accumulating evidence indicates that the Arfs are also important regulators of actin cytoskeleton dynamics and are involved in a variety of actin-based processes, including cell adhesion, migration and neurite outgrowth. The mechanisms of this regulation are remarkably diverse, ranging from the integration of vesicular transport with cytoskeleton assembly to the direct regulation of Rho-family GTPase function. Here, we review recent progress in our understanding of how Arfs and their interacting proteins function to integrate membrane and cytoskeletal dynamics.

Figure 1

Regulation of actin assembly by Arf1. (a) Arf1 acts both upstream and downstream of Cdc42 to promote vesicle formation in the Golgi. Cdc42 interacts with the γCOP subunit of the COPI coatomer complex and is brought to Golgi membranes by Arf1-mediated coatomer recruitment. The identity of the GEF that activates Golgi-associated Cdc42 is not known. Activated Cdc42 then stimulates N-WASP, which activates the Arp2/3 complex, leading to local actin assembly, and it is thought that this promotes vesicle scission. It is not known if Arf1 remains associated with the Cdc42-N-WASP complex. In addition, Arf1 also downregulates Cdc42 activity by recruiting the Cdc42 GAP, ARHGAP21, to Golgi membranes. In the absence of ARHGAP21, unregulated actin assembly leads to disruption of the Golgi complex. How the upstream and downstream functions of Arf1 are coordinated is not currently understood. (b) Arf1, Cdc42, and ARHGAP21 also cooperate at the plasma membrane to mediate clathrin-independent endocytosis. Cdc42 activation leads to the formation of endocytic vesicles containing fluid phase markers and GPI-anchored membrane proteins (vesicle budding) [22]. It is not known if this pathway also uses N-WASP to drive actin assembly. Activation of Arf1 on the plasma membrane recruits ARHGAP21, leading to downregulation of Cdc42 activity. Depletion of either Arf1 or ARHGAP21 by RNAi results in accumulation of Cdc42 in puncta at the plasma membrane [22]. (c) In the TGN, Arf1 activity promotes the assembly of a cortactin–dynamin complex at sites of vesicle formation through a mechanism that remains unknown. In a similar fashion to N-WASP, cortactin can activate the Arp2/3 complex and presumably acts in concert with dynamin to mediate vesicle scission.

Figure 2

Induction of lamellipodia by activation of Arf in the cell periphery. MDCK cells were infected with recombinant adenoviruses encoding the Arf GEF, ARNO. Epithelial cells typically cluster together and do not form lamellipodia even at subconfluent density (a). Expression of wild type ARNO induces the formation of large lamellipodia (b), whereas a catalytically inactive mutant (E156K) does not (c), indicating that activation of Arf is sufficient to drive lamellipodium assembly. Cells were stained to identify exogenous ARNO (green), and F-actin (red). Arrows indicate sites of lamellipodia formation. Images courtesy of Lorraine Santy.

Figure 3

Models for the regulation of Rac activity by Arf6. (a) GEF recruitment. The activation of Arf6 by ARNO triggers the recruitment of the Rac GEF, Dock180–ELMO, to the plasma membrane, where it then stimulates Rac activation. In this model, cytosolic Rac reaches the membrane by diffusion. (b) Endosomal translocation. In this model, Arf6 regulates the vesicular transport of Rac from perinuclear endosomes to the plasma membrane, where it is available to be activated by local GEFs. (c) Lipid raft translocation. Arf6 can control the trafficking of lipid raft components from endosomal compartments to the plasma membrane. Incorporation of Rac into lipid rafts at the plasma membrane is thought to be important for both its activation and its coupling to downstream effector proteins. In this model, cytosolic Rac reaches the plasma membrane by diffusion, but lipid raft components are transported in vesicular carriers whose formation requires Arf6.

Figure 4

Domain organization of the ARAP, ASAP and GIT families of Arf GAPs, and their known binding partners. Abbreviations: ANK, ankyrin repeat; BAR, Bin1–amphiphysin–Rvs167 domain; CC, coiled-coil domain; PBS, paxillin-binding sequence; PH, pleckstrin homology domain; PR, proline-rich domain; RA, Ras-association domain; SAM, sterile α-motif domain; SH3, Src homology 3 domain; SHD, Spa2 homology domain.