IQGAP1 Stimulates Actin Assembly through the N-Wasp-Arp2/3 Pathway

Abstract

IQGAP1 is a conserved modular protein overexpressed in cancer and involved in organizing actin and microtubules in motile processes such as adhesion, migration, and cytokinesis. A variety of proteins have been shown to interact with IQGAP1, including the small G proteins Rac1 and Cdc42, actin, calmodulin, ␤-catenin, the microtubule plus end-binding proteins CLIP170 (cytoplasmic linker protein) and adenomatous polyposis coli. However, the molecular mechanism by which IQGAP1 controls actin dynamics in cell motility is not understood. Quantitative co-localization analysis and down-regulation of IQGAP1 revealed that IQGAP1 controls the co-localization of N-WASP with the Arp2/3 complex in lamellipodia. Co-immunoprecipitation supports an in vivo link between IQGAP1 and N-WASP. Pull-down experiments and kinetic assays of branched actin polymerization with N-WASP and Arp2/3 complex demonstrated that the C-terminal half of IQGAP1 activates N-WASP by interacting with its BR-CRIB domain in a Cdc42-like manner, whereas the N-terminal half of IQGAP1 antagonizes this activation by association with a C-terminal region of IQGAP1. We propose that signal-induced relief of the autoinhibited fold of IQGAP1 allows activation of N-WASP to stimulate Arp2/3-dependent actin assembly.Directional cell migration results from the coordination of protrusion formation and cell adhesion. Although the concerted re-organization of actin and microtubules establishes and maintains cell polarization during directional movement, little is known about the molecular mechanisms underlying signal-mediated crosstalk between the two different cytoskeletal arrays (1). In this context, the modular IQGAP1 protein has received intense interest in the past years (2). The multiple partners of IQGAP1, including signaling molecules like Cdc42 or Rac1, calmodulin (3-6), and adhesion/cytoskeletal proteins like ␤-catenin, E-cadherin, actin filaments, and microtubule plus end-tracking proteins (CLIP170 and adenomatous polyposis coli (APC)) strongly suggest that IQGAP1 is an important player in coordinating cell polarity, adhesion, and migration (7-13). Concrete support to this view was brought by evidence showing that IQGAP1 is overexpressed in cancer (14, 15), controls cytokinesis (16 -21), and cell-cell adhesion (22-24). In addition, recent reports showed that IQGAP1 localizes in lamellipodia of motile cells (4,25,26) where it may link microtubule ends to the actin cytoskeleton (12,27) and that overexpression of IQGAP1 increases cell motility, whereas knockdown of the protein reduces cell migration and inhibits the formation of a protrusive actin meshwork at the leading edge (25). Finally, IQGAP1 regulates E-cadherinmediated cell-cell adhesion both positively and negatively (11). However, the functional and molecular link between IQGAP1 and the actin cytoskeleton in cell-cell adhesions and in lamellipodia has remained elusive.Extension of lamellipodia is driven by stimulus-responsive WASP family proteins (N-WASP, WASP, and Scar/WAVE) Cortactin and CARMIL, which act...

Endothelial IQGAP1 regulates efficient lymphocyte transendothelial migration

“…In our experiments, the IQGAP1-deficient HUVEC were plated at confluence, then maintained in complete media with 20% FBS for 48 h to promote junction maturation. Hence, in the current experiments, effects of IQGAP1 knockdown on cell migration or repopulation at subconfluent densities were minimized.IQGAP1 is known to associate with E-cadherin, regulate actin assembly, and coordinate the tethering of MT during polarized cell migration through interaction with MT plus-end binding proteins [21,22,24,35,36]. Our data implicate the participation of MT in endothelial IQGAP1-dependent junction remodeling during lymphocyte diapedesis.…”

A dual role for IQGAP1 in regulating exocytosis

“…By contrast, IQGAP1-N, which binds the exocyst-septin complex (this study), was reported to self-associate and to inhibit CDC42 activation, perhaps by associating with C-terminus sequences of endogenous IQGAP1 (Mataraza et al, 2002;Le Clainche et al, 2007), which would present a synergistic mechanism for enhancing secretion. Together, these data affirm the idea that IQGAP1 modulates CDC42 activity, but, at present, it remains unclear how, because purified recombinant IQGAP1 failed to exhibit a direct GEF activity on CDC42.…”

“…IQGAP1, the most studied, binds calmodulin, cross-links actin filaments, integrates signaling networks (reviewed in Mateer et al, 2003;Brown and Sacks, 2006), and regulates cell-cell contacts (Fukata et al, 1999;Fukata et al, 2001) and the capture of microtubule plus-ends via association with CLIP-170 (Fukata et al, 2002). The current paradigm is that IQGAP1 regulates actin assembly in cooperation with the ARP2/3 complex and the Rho GTPases in different cell types to regulate cell outgrowth and migration (Watanabe et al, 2004;Bensenor et al, 2007;Le Clainche et al, 2007;Wang et al, 2007). This role, however, does not contradict with an IQGAP1 essential function in secretion, because the mammalian exocyst subunit EXO70 (EXOC7) has also Polarized secretion is a tightly regulated event generated by conserved, asymmetrically localized multiprotein complexes, and the mechanism(s) underlying its temporal and spatial regulation are only beginning to emerge.…”

Protein 4.1R regulates cell migration and IQGAP1 recruitment to the leading edge

“…IQGAP1 also binds to Dia1, N-WASP and adenomatous polyposis coli (APC), a multifunctional protein that links IQGAP1 to microtubules . APC and Dia1 directly mediate actin filament assembly (Okada et al, 2010), whereas IQGAP1 stimulates actin polymerization through the N-WASP-Arp2/3 complex (Le Clainche et al, 2007). It has been demonstrated that IQGAP1 and APC or CLIP-170 recruitment to the leading edge are interdependent (Watanabe et al, 2004).…”