ReviewHow WASP-family proteins and the Arp2/3 complex convert intracellular signals into cytoskeletal structures
Introduction
Under the microscope cytoskeletal structures, such as the mitotic spindle or the leading edge of a motile cell, convey an overwhelming sense of order and plan. How are such large and complex structures assembled? And how do cellular signaling pathways control this process? Recent work on regulation of actin filament assembly provides clues about how one type of intracellular signal is converted into a three-dimensional structure.
Directed motility of almost all motile eukaryotic cells requires the assembly of a large, specialized network of actin filaments at the forward or leading edge of the cell; polymerization of these actin filaments in this network drives membrane protrusion [1]. To change direction in response to external cues, such as chemoattractants, actin polymerization must somehow be controlled by cellular signaling pathways. At present, we can trace one pathway from activation of small Rho-family G-proteins to de novo nucleation and crosslinking of actin filaments (Figure 1). The following review outlines the work that has allowed this pathway to be described and the molecular details of how G-protein activation is converted into actin assembly is then discussed.
Section snippets
Rho-family G-proteins regulate actin assembly and organization
The Rho-family of small G-proteins includes Rho, Rac and Cdc42. Activation of these proteins in vivo alters the structure of the actin cytoskeleton. Microinjection of activated Rho induces the formation of actin stress fibers [2], activated Rac induces actin-dependent membrane ruffling [3] and activated Cdc42 induces protrusion of actin-rich microspikes from the cell surface [4]. Co-injection of fluorescently-labeled actin, along with each of the G-proteins, reveals that Rac and Cdc42 induce
WASP-family proteins are downstream effectors of Rho-family proteins
The WASP-family of proteins consists of WASP, N-WASP and SCAR (Figure 2a). The available data suggest that, in order to initiate actin polymerization, Cdc42 acts through WASP or N-WASP and Rac acts through SCAR. WASP was originally identified as a protein that was altered in patients with Wiskott–Aldrich Syndrome, a genetic immunodeficiency disorder characterized by reduced motility of lymphoid immune cells [8]. N-WASP is a related and more ubiquitous protein, first discovered in the brain [9].
The Arp2/3 complex is a downstream effector of WASP-family proteins
The Arp2/3 complex is a stable complex of seven-subunits — two actin-related proteins (Arp2 and Arp3) and five novel proteins (p40, p35, p19, p18, and p14). The Arp2/3 complex was first discovered by affinity chromatography on the actin-binding protein profilin in protozoans. It is abundant [14] and conserved [15] across eukaryotic phyla. One of the subunits, p40, is essential in S. cerevisiae. Individual knockouts of other subunits are conditionally lethal and produce a profoundly disrupted
The Arp2/3 complex is a multifunctional protein complex that nucleates actin filaments and crosslinks them into orthogonal networks
Actin filaments are polarized. In vitro monomers add preferentially to the fast growing (barbed) end of the filament and this is the major site of filament elongation in vivo. The Arp2/3 complex is the only known cellular factor that nucleates formation of actin filaments that grow from their fast-growing (barbed) ends. After nucleation, the Arp2/3 complex remains attached to the slow-growing (pointed) end of the filament [22••]. The Arp2/3 complex also binds to the sides of actin filaments and
A general model for regulated actin polymerization
The above results can be integrated into a simple model [22••] of regulated actin polymerization (Figure 1) that may apply generally to any cellular process in which de novo actin polymerization drives membrane protrusion (e.g. macropinocytosis or reorganization of synaptic butons).
The first step is localization and activation of the nucleation machinery. Prenylated, GTP-bound Cdc42 localizes to the membrane where it recruits and activates WASP. Following this, activated WASP recruits the
Coincidence and polarity
Polarity is essential for directed motion. But is cell polarity established by signaling systems, by the cytoskeleton, or by both? In chemotactic cells the activity of chemoattractant receptors is polarized, even in the absence of polymerized actin 41•, 42. The mechanism behind the ability of the signaling system to establish polarity is not well understood but is thought to involve production of two signals, one that locally enhances receptor activity and another that globally inhibits it [41•]
Conclusion
With the discovery of the connection between Rho-family G proteins, WASP-family proteins and the Arp2/3 complex, we are now beginning to understand the signaling pathways that control actin assembly. It is clear, however, that much more work will be required before we understand precisely how cells establish polarity and the molecular details of how they convert signaling events into ordered three-dimensional structures.
References and recommended reading
Papers of particular interest, published within the annual period of review, have been highlighted as:
• of special interest
•• of outstanding interest
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