title: Interactions of ADF/cofilin, Arp2/3 complex, capping protein and profilin in remodeling of branched actin filament networks.
abstract: BACKGROUND
  Cellular movements are powered by the assembly and disassembly of actin filaments. Actin dynamics are controlled by Arp2/3 complex, the Wiskott-Aldrich syndrome protein (WASp) and the related Scar protein, capping protein, profilin, and the actin-depolymerizing factor (ADF, also known as cofilin). Recently, using an assay that both reveals the kinetics of overall reactions and allows visualization of actin filaments, we showed how these proteins co-operate in the assembly of branched actin filament networks. Here, we investigated how they work together to disassemble the networks.
  
  
  RESULTS
  Actin filament branches formed by polymerization of ATP-actin in the presence of activated Arp2/3 complex were found to be metastable, dissociating from the mother filament with a half time of 500 seconds. The ADF/cofilin protein actophorin reduced the half time for both dissociation of gamma-phosphate from ADP-Pi-actin filaments and debranching to 30 seconds. Branches were stabilized by phalloidin, which inhibits phosphate dissociation from ADP-Pi-filaments, and by BeF3, which forms a stable complex with ADP and actin. Arp2/3 complex capped pointed ends of ATP-actin filaments with higher affinity (Kd approximately 40 nM) than those of ADP-actin filaments (Kd approximately 1 microM), explaining why phosphate dissociation from ADP-Pi-filaments liberates branches. Capping protein prevented annealing of short filaments after debranching and, with profilin, allowed filaments to depolymerize at the pointed ends.
  
  
  CONCLUSIONS
  The low affinity of Arp2/3 complex for the pointed ends of ADP-actin makes actin filament branches transient. By accelerating phosphate dissociation, ADF/cofilin promotes debranching. Barbed-end capping proteins and profilin allow dissociated branches to depolymerize from their free pointed ends.
authors.affiliation: Structural Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA.
authors.forename: L
authors.initials: L
authors.lastname: Blanchoin
authors.forename: T D
authors.initials: TD
authors.lastname: Pollard
authors.forename: R D
authors.initials: RD
authors.lastname: Mullins
mesh_terms: D000048:Acanthamoeba; D008841:Actin Cytoskeleton / Q000378:metabolism* / Q000648:ultrastructure; D051339:Actin Depolymerizing Factors; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure*; D000818:Animals; D003598:Cytoskeletal Proteins; D003712:Dendrites / Q000502:physiology / Q000648:ultrastructure; D007700:Kinetics; D008840:Microfilament Proteins / Q000378:metabolism*; D015800:Protozoan Proteins / Q000378:metabolism
chemical_list: D051339:Actin Depolymerizing Factors; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; D003598:Cytoskeletal Proteins; D008840:Microfilament Proteins; D015800:Protozoan Proteins; C047604:actophorin protein, Acanthamoeba