title: Critical conformational changes in the Arp2/3 complex are induced by nucleotide and nucleation promoting factor.
abstract: Actin nucleation and branching by the Arp2/3 complex is tightly regulated by activating factors. However, the mechanism of Arp2/3 complex activation remains unclear. We used fluorescence resonance energy transfer (FRET) to probe the conformational dynamics of the Arp2/3 complex accompanying its activation. We demonstrate that nucleotide binding promotes a substantial conformational change in the complex, with distinct conformations depending on the bound nucleotide. Nucleotide binding to each Arp is critical for activity and is coupled to nucleation promoting factor (NPF) binding. The binding of Wiskott-Aldrich syndrome protein (WASP) family NPFs induces further conformational reorganization of the Arp2/3 complex, and the ability to promote this conformational reorganization correlates with activation efficiency. Using an Arp2/3 complex that is fused to the actin binding domain of WASP, we confirm that the NPF-induced conformational change is critical for activation, and that the actin and Arp2/3 binding activities of WASP are separable, but are independently essential for activity.
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
authors.forename: Erin D
authors.initials: ED
authors.lastname: Goley
authors.forename: Stacia E
authors.initials: SE
authors.lastname: Rodenbusch
authors.forename: Adam C
authors.initials: AC
authors.lastname: Martin
authors.forename: Matthew D
authors.initials: MD
authors.lastname: Welch
mesh_terms: D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D008087:Listeria / Q000737:chemistry / Q000378:metabolism; D009711:Nucleotides / Q000378:metabolism; D011487:Protein Conformation; D011506:Proteins / Q000378:metabolism; D051316:Wiskott-Aldrich Syndrome Protein
chemical_list: D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; D003598:Cytoskeletal Proteins; D009711:Nucleotides; D011506:Proteins; D051316:Wiskott-Aldrich Syndrome Protein