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. These actin filaments are linked into a network 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. Because Arp2/3 complex nucleates actin polymerization and links the pointed end to the side of another filament in vitro, a dendritic nucleation model has been proposed in which Arp2/3 complex initiates filaments from the sides of older filaments. Here we report, by using a light microscopy assay, many new features of the mechanism. Branching occurs during, rather than after, nucleation by Arp2/3 complex activated by the Wiskott-Aldrich syndrome protein (WASP) or Scar protein; capping protein and profilin act synergistically with Arp2/3 complex to favour branched nucleation; phosphate release from aged actin filaments favours dissociation of Arp2/3 complex from 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 family and favour the selective disassembly of proximal regions of the network.
Blanchoin
L
L
Structural Biology Laboratory, Salk Institute for Biological Studies, La Jolla, California 92037, USA.
Amann
K J
KJ
Higgs
H N
HN
Marchand
J B
JB
Kaiser
D A
DA
Pollard
T D
TD
D000048:Acanthamoeba; D051339:Actin Depolymerizing Factors; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D003285:Contractile Proteins; D003598:Cytoskeletal Proteins; D051345:Destrin; D006801:Humans; D008840:Microfilament Proteins / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism; D009711:Nucleotides / Q000378:metabolism; D051304:Profilins; D011506:Proteins / Q000378:metabolism*; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D051316:Wiskott-Aldrich Syndrome Protein; D051300:Wiskott-Aldrich Syndrome Protein Family
C495926:ACTR2 protein, human; C495928:ACTR3 protein, human; D051339:Actin Depolymerizing Factors; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; D003285:Contractile Proteins; D003598:Cytoskeletal Proteins; D051345:Destrin; D008840:Microfilament Proteins; D009711:Nucleotides; C495738:PFN1 protein, human; D051304:Profilins; D011506:Proteins; D011994:Recombinant Proteins; C495767:WAS protein, human; C114864:WASF1 protein, human; D051316:Wiskott-Aldrich Syndrome Protein; D051300:Wiskott-Aldrich Syndrome Protein Family