title: Mechanism of synergistic activation of Arp2/3 complex by cortactin and N-WASP.
abstract: Nucleation promoting factors (NPFs) initiate branched actin network assembly by activating Arp2/3 complex, a branched actin filament nucleator. Cellular actin networks contain multiple NPFs, but how they coordinately regulate Arp2/3 complex is unclear. Cortactin is an NPF that activates Arp2/3 complex weakly on its own, but with WASP/N-WASP, another class of NPFs, potently activates. We dissect the mechanism of synergy and propose a model in which cortactin displaces N-WASP from nascent branches as a prerequisite for nucleation. Single-molecule imaging revealed that unlike WASP/N-WASP, cortactin remains bound to junctions during nucleation, and specifically targets junctions with a ∼160-fold increased on rate over filament sides. N-WASP must be dimerized for potent synergy, and targeted mutations indicate release of dimeric N-WASP from nascent branches limits nucleation. Mathematical modeling shows cortactin-mediated displacement but not N-WASP recycling or filament recruitment models can explain synergy. Our results provide a molecular basis for coordinate Arp2/3 complex regulation. DOI:http://dx.doi.org/10.7554/eLife.00884.001.
authors.affiliation: Institute of Molecular Biology , University of Oregon , Eugene , United States ; Department of Chemistry and Biochemistry , University of Oregon , Eugene , United States.
authors.forename: Luke A
authors.initials: LA
authors.lastname: Helgeson
authors.forename: Brad J
authors.initials: BJ
authors.lastname: Nolen
mesh_terms: D051376:Actin-Related Protein 2-3 Complex / Q000378:metabolism*; D051356:Cortactin / Q000378:metabolism*; D058105:Polymerization; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal / Q000378:metabolism*
chemical_list: D051376:Actin-Related Protein 2-3 Complex; D051356:Cortactin; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal
keywords: Arp2/3; Mouse; N-WASP; WASP; actin; cortactin; single molecule