The Arp2/3 complex nucleates actin filament branches from the sides of pre-existing filaments.
Regulated assembly of actin-filament networks provides the mechanical force that pushes forward the leading edge of motile eukaryotic cells and intracellular pathogenic bacteria and viruses. When activated by binding to actin filaments and to the WA domain of Wiskott-Aldrich-syndrome protein (WASP)/Scar proteins, the Arp2/3 complex nucleates new filaments that grow from their barbed ends. The Arp2/3 complex binds to the sides and pointed ends of actin filaments, localizes to distinctive 70 degrees actin-filament branches present in lamellae, and forms similar branches in vitro. These observations have given rise to the dendritic nucleation model for actin-network assembly, in which the Arp2/3 complex initiates branches on the sides of older filaments. Recently, however, an alternative mechanism for branch formation has been proposed. In the 'barbed-end nucleation' model, the Arp2/3 complex binds to the free barbed end of a filament and two filaments subsequently grow from the branch. Here we report the use of kinetic and microscopic experiments to distinguish between these models. Our results indicate that the activated Arp2/3 complex preferentially nucleates filament branches directly on the sides of pre-existing filaments.
Amann
K J
KJ
Structural Biology Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines Road, La Jolla, California 92037, USA.
Pollard
T D
TD
D051339:Actin Depolymerizing Factors; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001704:Biopolymers / Q000378:metabolism; D003598:Cytoskeletal Proteins; D051345:Destrin; D005456:Fluorescent Dyes / Q000378:metabolism; D018260:Gelsolin / Q000378:metabolism; D007700:Kinetics; D008840:Microfilament Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism*; D051316:Wiskott-Aldrich Syndrome Protein; D051300:Wiskott-Aldrich Syndrome Protein Family
D051339:Actin Depolymerizing Factors; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; D001704:Biopolymers; D003598:Cytoskeletal Proteins; D051345:Destrin; D005456:Fluorescent Dyes; D018260:Gelsolin; D008840:Microfilament Proteins; D011506:Proteins; C114864:WASF1 protein, human; D051316:Wiskott-Aldrich Syndrome Protein; D051300:Wiskott-Aldrich Syndrome Protein Family; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate