TY - JOUR TI - Mechanism of synergistic activation of Arp2/3 complex by cortactin and N-WASP AU - Helgeson, Luke A AU - Nolen, Brad J A2 - Sundquist, Wesley VL - 2 PY - 2013 DA - 2013/09/03 SP - e00884 C1 - eLife 2013;2:e00884 DO - 10.7554/eLife.00884 UR - https://doi.org/10.7554/eLife.00884 AB - 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. KW - Arp2/3 KW - actin KW - WASP KW - cortactin KW - single molecule KW - N-WASP JF - eLife SN - 2050-084X PB - eLife Sciences Publications, Ltd ER -