Regulation of Actin Polymerization and Adhesion-Dependent Cell Edge Protrusion by the Abl-Related Gene (Arg) Tyrosine Kinase and N-WASp

Abstract

Extracellular cues stimulate the Abl family nonreceptor tyrosine kinase Arg to promote actin-based cell edge protrusions. Several Arg-interacting proteins are potential links to the actin cytoskeleton, but exactly how Arg stimulates actin polymerization and cellular protrusion has not yet been fully elucidated. We used affinity purification to identify N-WASp as a novel binding partner of Arg. N-WASp activates the Arp2/3 complex and is an effector of Abl. We find that the Arg SH3 domain binds directly to N-WASp. Arg phosphorylates N-WASp on Y256, modestly increasing the affinity of Arg for N-WASp, an interaction that does not require the Arg SH2 domain. The Arg SH3 domain stimulates N-WASp-dependent actin polymerization in vitro, and Arg phosphorylation of N-WASp weakly stimulates this effect. Arg and N-WASp colocalize to adhesion-dependent cell edge protrusions in vivo. The cell edge protrusion defects of arg−/− fibroblasts can be complemented by re-expression of an Arg-yellow fluorescent protein (YFP) fusion, but not by an N-WASp binding-deficient Arg SH3 domain point mutant. These results suggest that Arg promotes actin-based protrusions in response to extracellular stimuli through phosphorylation of and physical interactions with N-WASp.

Cdc42 and the RhoGEF Intersectin-1 collaborate with Nck to promote N-WASP-dependent actin polymerisation

“…It should, however, not be forgotten that the SH3 domains in ITSN1, as well as those in Src and Abl family kinases, might also contribute activating inputs into N-WASP in the absence of Cdc42 and Nck. Indeed, the SH3 domain of the tyrosine kinase Arg, which is recruited by vaccinia (Reeves et al, 2005;Newsome et al, 2006), is capable of activating N-WASP in vitro (Miller et al, 2010). Nevertheless, our analysis suggests that in the absence of Grb2 recruitment, Arp2/3-dependent actin based motility of vaccinia is principally driven by Cdc42-and Nck-dependent activation of N-WASP.…”

Abl2/Abl-related Gene Stabilizes Actin Filaments, Stimulates Actin Branching by Actin-related Protein 2/3 Complex, and Promotes Actin Filament Severing by Cofilin

“…Abl family kinases use both kinase-dependent and kinase-independent mechanisms to regulate the formation of actin-based cellular structures. Previous work described how Arg phosphorylates discrete cytoskeletal regulatory proteins to control changes in cell structure and function (8,11,(15)(16)(17)(41)(42)(43)(44)(45).…”

The Src kinases Hck, Fgr, and Lyn activate Abl2/Arg to facilitate IgG-mediated phagocytosis andLeishmaniainfection

“…Receptor engagement stimulates these kinases to bind and phosphorylate Arp2/3 complex activators (Lapetina et al, 2009;Miller et al, 2010), yielding dynamic cell edge protrusions that resemble phagocytic intermediates. Abl and Arg also facilitate endocytosis (Jacob et al, 2009;Pendergast, 2006, 2007), autophagy (Yogalingam and Pendergast, 2008), viral (Reeves et al, 2005(Reeves et al, , 2011Swimm et al, 2010) and bacterial uptake (Burton et al, 2003;Elwell et al, 2008;Ly and Casanova, 2009;Napier et al, 2011), and IgG-mediated phagocytosis (Greuber and Pendergast, 2012).…”