WASP and WAVE proteins belong to the Wiskott-Aldrich Syndrome protein family, with recessive mutations in the founding member WASP being responsible for the X-linked recessive immunodeficieny known as the Wiskott-Aldrich Syndrome. WASP proteins include WASP and WASL (N-WASP). WAVE proteins include WASF1 (WAVE1), WASF2 (WAVE2) and WASF3 (WAVE3). WASPs and WAVEs contain a VCA domain (consisting of WH2 and CA subdomains) at the C-terminus, responsible for binding to G-actin (WH2 subdomain) and the actin-associated ARP2/3 complex (CA subdomain). WASPs contain a WH1 (WASP homology 1) domain at the N-terminus, responsible for binding to WIPs (WASP-interacting proteins). A RHO GTPase binding domain (GBD) is located in the N-terminal half of WASPs and C-terminally located in WAVEs. RHO GTPases activate WASPs by disrupting the autoinhibitory interaction between the GBD and VCA domains, which allows WASPs to bind actin and the ARP2/3 complex and act as nucleation promoting factors in actin polymerization. WAVEs have the WAVE/SCAR homology domain (WHD/SHD) at the N-terminus, which binds ABI, NCKAP1, CYFIP2 and BRK1 to form the WAVE regulatory complex (WRC). Binding of the RAC1:GTP to the GBD of WAVEs most likely induces a conformational change in the WRC that allows activating phosphorylation of WAVEs by ABL1, thus enabling them to function as nucleation promoting factors in actin polymerization through binding G-actin and the ARP2/3 complex (Reviewed by Lane et al. 2014).
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Rohatgi R, Nollau P, Ho HY, Kirschner MW, Mayer BJ.; ''Nck and phosphatidylinositol 4,5-bisphosphate synergistically activate actin polymerization through the N-WASP-Arp2/3 pathway.''; PubMedEurope PMCScholia
Nimmerjahn F, Ravetch JV.; ''Fcgamma receptors: old friends and new family members.''; PubMedEurope PMCScholia
Le Clainche C, Carlier MF.; ''Regulation of actin assembly associated with protrusion and adhesion in cell migration.''; PubMedEurope PMCScholia
Wu X, Suetsugu S, Cooper LA, Takenawa T, Guan JL.; ''Focal adhesion kinase regulation of N-WASP subcellular localization and function.''; PubMedEurope PMCScholia
Amann KJ, Pollard TD.; ''The Arp2/3 complex nucleates actin filament branches from the sides of pre-existing filaments.''; PubMedEurope PMCScholia
Nakanishi O, Suetsugu S, Yamazaki D, Takenawa T.; ''Effect of WAVE2 phosphorylation on activation of the Arp2/3 complex.''; PubMedEurope PMCScholia
Kato M, Miki H, Kurita S, Endo T, Nakagawa H, Miyamoto S, Takenawa T.; ''WICH, a novel verprolin homology domain-containing protein that functions cooperatively with N-WASP in actin-microspike formation.''; PubMedEurope PMCScholia
Lane J, Martin T, Weeks HP, Jiang WG.; ''Structure and role of WASP and WAVE in Rho GTPase signalling in cancer.''; PubMedEurope PMCScholia
Bompard G, Caron E.; ''Regulation of WASP/WAVE proteins: making a long story short.''; PubMedEurope PMCScholia
Callebaut I, Cossart P, Dehoux P.; ''EVH1/WH1 domains of VASP and WASP proteins belong to a large family including Ran-binding domains of the RanBP1 family.''; PubMedEurope PMCScholia
Cory GO, Garg R, Cramer R, Ridley AJ.; ''Phosphorylation of tyrosine 291 enhances the ability of WASp to stimulate actin polymerization and filopodium formation. Wiskott-Aldrich Syndrome protein.''; PubMedEurope PMCScholia
GarcÃa-GarcÃa E, Rosales C.; ''Signal transduction during Fc receptor-mediated phagocytosis.''; PubMedEurope PMCScholia
Le Clainche C, Pantaloni D, Carlier MF.; ''ATP hydrolysis on actin-related protein 2/3 complex causes debranching of dendritic actin arrays.''; PubMedEurope PMCScholia
Park H, Cox D.; ''Cdc42 regulates Fc gamma receptor-mediated phagocytosis through the activation and phosphorylation of Wiskott-Aldrich syndrome protein (WASP) and neural-WASP.''; PubMedEurope PMCScholia
Ho HY, Rohatgi R, Ma L, Kirschner MW.; ''CR16 forms a complex with N-WASP in brain and is a novel member of a conserved proline-rich actin-binding protein family.''; PubMedEurope PMCScholia
Zalevsky J, Lempert L, Kranitz H, Mullins RD.; ''Different WASP family proteins stimulate different Arp2/3 complex-dependent actin-nucleating activities.''; PubMedEurope PMCScholia
Higgs HN, Pollard TD.; ''Activation by Cdc42 and PIP(2) of Wiskott-Aldrich syndrome protein (WASp) stimulates actin nucleation by Arp2/3 complex.''; PubMedEurope PMCScholia
Egile C, Rouiller I, Xu XP, Volkmann N, Li R, Hanein D.; ''Mechanism of filament nucleation and branch stability revealed by the structure of the Arp2/3 complex at actin branch junctions.''; PubMedEurope PMCScholia
Rohatgi R, Ma L, Miki H, Lopez M, Kirchhausen T, Takenawa T, Kirschner MW.; ''The interaction between N-WASP and the Arp2/3 complex links Cdc42-dependent signals to actin assembly.''; PubMedEurope PMCScholia
Kim AS, Kakalis LT, Abdul-Manan N, Liu GA, Rosen MK.; ''Autoinhibition and activation mechanisms of the Wiskott-Aldrich syndrome protein.''; PubMedEurope PMCScholia
Leng Y, Zhang J, Badour K, Arpaia E, Freeman S, Cheung P, Siu M, Siminovitch K.; ''Abelson-interactor-1 promotes WAVE2 membrane translocation and Abelson-mediated tyrosine phosphorylation required for WAVE2 activation.''; PubMedEurope PMCScholia
Aspenström P.; ''The WASP-binding protein WIRE has a role in the regulation of the actin filament system downstream of the platelet-derived growth factor receptor.''; PubMedEurope PMCScholia
Takenawa T, Miki H.; ''WASP and WAVE family proteins: key molecules for rapid rearrangement of cortical actin filaments and cell movement.''; PubMedEurope PMCScholia
Mendoza MC, Er EE, Zhang W, Ballif BA, Elliott HL, Danuser G, Blenis J.; ''ERK-MAPK drives lamellipodia protrusion by activating the WAVE2 regulatory complex.''; PubMedEurope PMCScholia
Millard TH, Sharp SJ, Machesky LM.; ''Signalling to actin assembly via the WASP (Wiskott-Aldrich syndrome protein)-family proteins and the Arp2/3 complex.''; PubMedEurope PMCScholia
Moreau V, Frischknecht F, Reckmann I, Vincentelli R, Rabut G, Stewart D, Way M.; ''A complex of N-WASP and WIP integrates signalling cascades that lead to actin polymerization.''; PubMedEurope PMCScholia
Carlier MF, Nioche P, Broutin-L'Hermite I, Boujemaa R, Le Clainche C, Egile C, Garbay C, Ducruix A, Sansonetti P, Pantaloni D.; ''GRB2 links signaling to actin assembly by enhancing interaction of neural Wiskott-Aldrich syndrome protein (N-WASp) with actin-related protein (ARP2/3) complex.''; PubMedEurope PMCScholia
Mullins RD, Heuser JA, Pollard TD.; ''The interaction of Arp2/3 complex with actin: nucleation, high affinity pointed end capping, and formation of branching networks of filaments.''; PubMedEurope PMCScholia
Miki H, Suetsugu S, Takenawa T.; ''WAVE, a novel WASP-family protein involved in actin reorganization induced by Rac.''; PubMedEurope PMCScholia
Suetsugu S, Kurisu S, Oikawa T, Yamazaki D, Oda A, Takenawa T.; ''Optimization of WAVE2 complex-induced actin polymerization by membrane-bound IRSp53, PIP(3), and Rac.''; PubMedEurope PMCScholia
Indik ZK, Park JG, Hunter S, Schreiber AD.; ''The molecular dissection of Fc gamma receptor mediated phagocytosis.''; PubMedEurope PMCScholia
Phagocytosis is one of the important innate immune responses that function to eliminate invading infectious agents. Monocytes, macrophages, and neutrophils are the professional phagocytic cells. Phagocytosis is a complex process involving the recognition of invading foreign particles by specific types of phagocytic receptors and the subsequent internalization of the particles. Fc gamma receptors (FCGRs) are among the best studied phagocytic receptors that bind to Fc portion of immunoglobulin G (IgG). Through their antigen binding F(ab) end, antibodies bind to specific antigen while their constant (Fc) region binds to FCGRs on phagocytes. The clustering of FCGRs by IgG antibodies on the phagocyte initiates a variety of signals, which lead, through the reorganisation of actin cytoskeleton and membrane remodelling, to the formation of pseudopod and phagosome. Fc gamma receptors are classified into three classes: FCGRI, FCGRII and FCGRIII. Each class of these FCGRs consists of several individual isoforms. Among all these isoforms FCGRI, FCGRIIA and FCGRIIIA, are able to mediate phagocytosis (Joshi et al. 2006, Garcia Garcia & Rosales 2002, Nimmerjahn & Ravetch 2006).
WASP family verprolin-homologous proteins (WAVEs) function downstream of RAC1 and are involved in activation of the ARP2/3 complex. The resulting actin polymerization mediates the projection of the plasma membrane in lamellipodia and membrane ruffles. WAVEs exist as a pentameric hetero-complex called WAVE Regulatory Complex (WRC). The WRC consists of a WAVE family protein (WASF1, WASF2 or WASF3 - commonly known as WAVE1, WAVE2 or WAVE3), ABI (Abelson-interacting protein), NCKAP1 (NAP1, p125NAP1), CYFIP1 (SRA1) or the closely related CYFIP2 (PIR121), and BRK1 (HSPC300, BRICK). Of the three structurally conserved WAVEs in mammals, the importance of WAVE2 in activation of the ARP2/3 complex and the consequent formation of branched actin filaments is best established. WAVEs in the WRC are intrinsically inactive and are stimulated by RAC1 GTPase and phosphatidylinositols (PIP3). The C-terminal VCA domain of WAVE2 (and likely WAVE1 and WAVE3) which can bind both the ARP2/3 complex and actin monomers (G-actin) is masked in the inactive state. After PIP3 binds to the polybasic region of WAVE2 (and likely WAVE1 and WAVE3) and RAC1:GTP binds to the CYFIP1 (or CYFIP2) subunit of the WRC, allosteric changes most likely occur which allow WAVEs to interact with the ARP2/3 complex. The interactions between WAVEs and RAC1 are indirect. BAIAP2/IRSp53, an insulin receptor substrate, acts as a linker, binding both activated RAC1 and the proline-rich region of WAVE2 (and likely WAVE1 and WAVE3) and forming a trimolecular complex. CYFIP1 (or CYFIP2) in the WAVE regulatory complex binds directly to RAC1:GTP and links it to WAVE2 (and likely WAVE1 and WAVE3) (Derivery et al. 2009, Yamazaki et al. 2006, Takenawa & Suetsugu 2007, Chen et al. 2010, Pollard 2007, Lebensohn & Kirschner 2009).
Once activated, the ARP2/3 complex nucleates new actin filaments that extend from the sides of pre-existing mother actin filaments at a 70-degree angle to form Y-branched networks (Firat-Karalar & Welch 2010). These branched actin filaments push the cell membrane forward to form a pseudopod. The ARP2/3 complex is composed of two Arps (actin-related proteins), ARP2 and ARP3, and five unique proteins ARPC1, ARPC2, ARPC3, ARPC4 and ARPC5 (Gournier et al. 2001). Both ARP2 and ARP3 subunits bind ATP. There are two proposed models to explain the process of actin nucleation by ARP2/3 complex: the barbed-end branching model and the dendritic nucleation/side branching model (Le Clainche & Carlier 2008). In barbed-end branching model, the branching/ternary complex (G-actin-WASP/WAVE-Arp2/3 complex) binds to the barbed end of the mother filament. G-actin bound to VCA domain or one of the Arp subunits incorporates into the mother filament at the barbed end, thus positioning ARP2/3 complex to initiate the daughter branch on the side of the mother filament. ARP2/3 nucleates the formation of new actin filament branches, which elongate at the barbed ends (Le Clainche & Carlier 2008, Pantaloni et al 2000, Le Clainche et al. 2003, Egile et al. 2005). In side branching model, the branching complex binds to the side of the mother actin filament mimicking an actin nucleus and initiates a lateral branch (Le Clainche & Carlier 2008, Amann & Pollard 2001).
The ARP2/3 complex shows higher affinity for the phosphorylated VCA domain of WAVE2 than for the unphosphorylated VCA domain. WAVE proteins can be phosphorylated by various kinases. Active ERK (Mitogen activated protein kinase 3) phosphorylates the WAVE regulatory complex (WRC) on multiple serine/threonine sites within the proline-rich domains (PRDs) of WAVE2 and ABI1. Phosphorylation of the PRDs would disrupt their interaction with SH3 and PLP binding domains, potentially altering WRC activation. ERK phosphorylates both S343 and T346 in WAVE2 and S183, S216, S225, S392, and S410 in ABI1. Cumulatively, the phosphorylation of both WAVE2 and ABI in the WAVE regulatory complex (WRC) contributes to the RAC-induced WRC conformational change that exposes the VCA domain, leading to binding and activation of ARP2/3 (Mendoza et al. 2011, Nakanishi et al. 2007). ERK phosphorylation sites in WAVE2 are not strictly conserved in WAVE1 and WAVE3 but, based on the amino acid sequence, other potential ERK phosphorylation sites exist.
ATP bound G-actin monomers are added to the fast growing barbed ends of both mother and daughter filaments. The polymerization of these filaments drives membrane protrusion. In the process of phagocytosis, pseudopodia extend around the antibody-bound particle to form the phagocytic cup. This elongation continues until the filament reaches steady state equilibrium with free G-actin monomers (Millard et al. 2004, Le Clainche et al. 2008).
Abelson interactor-1 (ABL) tyrosine kinase phosphorylates the strictly conserved tyrosine 150 in WAVE2 (Y151 in WAVE1 and WAVE3) (Leng et al. 2003, Chen et al. 2010).
After incorporation at the branch, the actin bound to VCA domain of WASP/WAVE undergoes ATP hydrolysis and this destabilizes its interaction with WASP/WAVE. This dissociates the branched junction from the membrane-bound WASP/WAVE (Kovar 2006).
WASP interacting proteins (WIP) family includes WIPF1 (WIP), WIPF2 (WIRE,WICH) and WIPF3 (CR16, corticosteroids and regional expression-16). WIPs share a specific proline rich sequence that interacts with the WH1 domain of WASP and N-WASP (WASL). WIPs form heterocomplexes with WASPs and may contribute to the WASP protein stability (Aspenstrom 2002, Kato et al. 2002, Ho et al. 2001, Moreau et al. 2000). SH3 domain containing adaptor proteins like GRB2 (Carlier et al. 2000), NCK (Rohatgi et al. 2001) and WISH (DIP/SPIN90) (Fukuoka et al. 2001) bind to the proline rich domain in WASPs and activate the ARP2/3 complex. By binding simultaneously to N-WASP and the ARP2/3 complex, GRB2 works synergistically with CDC42 in the activation of ARP2/3 complex-mediated actin assembly (Carlier et al. 2000).
WASP is phosphorylated on Tyr291 (Cory et al. 2002) and N-WASP (WASL) on Tyr256 (Wu et al. 2004) by Src family of tyrosine kinases and this phosphorylation may release the autoinhibitory intramolecular interactions. The phosphorylation seems to be enhanced by the activation of CDC42. WASP phosphorylation and binding of CDC42 have a synergistic effect on the activation of the ARP2/3 complex (Takenawa & Suetsugu 2007). In N-WASP, the phosphorylation may reduce its nuclear translocation and may sustain it in its functional site in the cytoplasm (Wu et al. 2004).
Wiskott-Aldrich syndrome protein (WASP) and Neural-WASP (N-WASP, WASL) proteins are scaffolds that transduce signals from cell surface receptors to the activation of the ARP2/3 complex and actin polymerization. WASP and N-WASP possess a central GTPase binding domain (GBD) and an NH2-terminal WASP homology domain 1 (WH1) followed by a basic region (B), and a C-terminal VCA region that contains: a V domain (verprolin homology/WASP homology 2), a C domain (connecting), and an A motif (acidic). The VCA region is responsible for binding to and activating the ARP2/3 complex (Bompard & Caron 2004, Callebaut et al 1998). Under resting conditions, WASP and N-WASP are maintained in an autoinhibited state via interaction of the GBD and the VCA domains. This prevents access of the ARP2/3 complex and G-actin to the VCA region. Activated CDC42 binds to the GBD region of WASPs and this interaction releases the VCA region from autoinhibition, enabling binding of the ARP2/3 complex and stimulating actin polymerization (Kim et al 2000, Park & Cox 2009). Phosphoinositides (PtdIns(4,5)P2) interact with the basic (B) region in WASPs and this interaction is important for activation of the WASPs and the ARP2/3 complex (Higgs & Pollard 2000).
Once WASPs (WASP and N-WASP) and WAVEs (WAVE2 and probably WAVE1 and WAVE3) are activated, their VCA region becomes available for binding to the ARP2/3 complex and actin monomer (G-actin). The actin monomer binds to the V domain and ARP2/3 complex binds to the CA domain. The simultaneous binding of G-actin and the ARP2/3 complex to the VCA region contributes to the activation of the ARP2/3-complex-mediated actin polymerization. The VCA module acts as a platform on which an actin monomer binds to the ARP2/3 complex to trigger actin polymerization (Takenawa & Suetsugu 2007).
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DataNodes
(FCGR) dependent
phagocytosisfilament:branching complex:daughter
filamentfilament:branching
complexN-WASP:ARP2/3
complex:G-actinAnnotated Interactions
filament:branching complex:daughter
filamentfilament:branching
complexfilament:branching
complexIn barbed-end branching model, the branching/ternary complex (G-actin-WASP/WAVE-Arp2/3 complex) binds to the barbed end of the mother filament. G-actin bound to VCA domain or one of the Arp subunits incorporates into the mother filament at the barbed end, thus positioning ARP2/3 complex to initiate the daughter branch on the side of the mother filament. ARP2/3 nucleates the formation of new actin filament branches, which elongate at the barbed ends (Le Clainche & Carlier 2008, Pantaloni et al 2000, Le Clainche et al. 2003, Egile et al. 2005). In side branching model, the branching complex binds to the side of the mother actin filament mimicking an actin nucleus and initiates a lateral branch (Le Clainche & Carlier 2008, Amann & Pollard 2001).
SH3 domain containing adaptor proteins like GRB2 (Carlier et al. 2000), NCK (Rohatgi et al. 2001) and WISH (DIP/SPIN90) (Fukuoka et al. 2001) bind to the proline rich domain in WASPs and activate the ARP2/3 complex. By binding simultaneously to N-WASP and the ARP2/3 complex, GRB2 works synergistically with CDC42 in the activation of ARP2/3 complex-mediated actin assembly (Carlier et al. 2000).
N-WASP:ARP2/3
complex:G-actinN-WASP:ARP2/3
complex:G-actin