Direct observation of dendritic actin filament networks nucleated by Arp2/3 complex and WASP/Scar proteins

Most nucleated cells crawl about by extending a pseudopod that is driven by the polymerization of actin filaments in the cytoplasm behind the leading edge of the plasma membrane(1,2). These actin filaments are linked into a n etwork by Y-branches, with the pointed end of each filament attached to the side of another filament and the rapidly growing barbed end facing forward (3). Because Arp2/3 complex nucleates actin polymerization and links the po inted end to the side of another filament in vitro, a dendritic nucleation model has been proposed 4 in which Arp2/3 complex initiates filaments from the sides of older filaments. Here we report, by using a light microscopy a ssay, many new features of the mechanism. Branching occurs during, rather t han after, nucleation by Arp2/3 complex activated by the Wiskott-Aldrich sy ndrome protein (WASP) or Scar protein; capping protein and profilin act syn ergistically with Arp2/3 complex to favour branched nucleation; phosphate r elease from aged actin filaments favours dissociation of Arp2/3 complex fro m the pointed ends of filaments; and branches created by Arp2/3 complex are relatively rigid. These properties result in the automatic assembly of the branched actin network after activation by proteins of the WASP/ Scar fami ly and favour the selective disassembly of proximal regions of the network.