Activation by Cdc42 and PIP(2) of Wiskott-Aldrich syndrome protein (WASp) stimulates actin nucleation by Arp2/3 complex.
We purified native WASp (Wiskott-Aldrich Syndrome protein) from bovine thymus and studied its ability to stimulate actin nucleation by Arp2/3 complex. WASp alone is inactive in the presence or absence of 0.5 microM GTP-Cdc42. Phosphatidylinositol 4,5 bisphosphate (PIP(2)) micelles allowed WASp to activate actin nucleation by Arp2/3 complex, and this was further enhanced twofold by GTP-Cdc42. Filaments nucleated by Arp2/3 complex and WASp in the presence of PIP(2) and Cdc42 concentrated around lipid micelles and vesicles, providing that Cdc42 was GTP-bound and prenylated. Thus, the high concentration of WASp in neutrophils (9 microM) is dependent on interactions with both acidic lipids and GTP-Cdc42 to activate actin nucleation by Arp2/3 complex. The results also suggest that membrane binding increases the local concentrations of Cdc42 and WASp, favoring their interaction.
Higgs
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The Salk Institute for Biological Studies, La Jolla, California 92037, USA.
Pollard
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D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000378:metabolism*; D000818:Animals; D000906:Antibodies; D002417:Cattle; D003598:Cytoskeletal Proteins; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D050356:Lipid Metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D018482:Muscle, Skeletal / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D009504:Neutrophils / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism*; D017368:Protein Prenylation; D011817:Rabbits; D013950:Thymus Gland / Q000378:metabolism; D014923:Wiskott-Aldrich Syndrome / Q000378:metabolism*; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism*
C495926:ACTR2 protein, human; C495928:ACTR3 protein, human; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; D000906:Antibodies; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; C495771:WASL protein, human; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; D019204:GTP-Binding Proteins; D020764:cdc42 GTP-Binding Protein