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J. Cell Biol. 150 (6): 1311-1320

Copyright © 2000 by the Rockefeller University Press.


Original Article

Activation by Cdc42 and Pip2 of Wiskott-Aldrich Syndrome Protein (Wasp) Stimulates Actin Nucleation by Arp2/3 Complex

Henry N. Higgsa, and Thomas D. Pollarda

a The Salk Institute for Biological Studies, La Jolla, California 92037
The Salk Institute for Biological Studies, 10010 North Torrey Pines Road, La Jolla, CA 92037.(858) 452-3683(858) 453-4100 x1716

Abstract: 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 µM GTP-Cdc42. Phosphatidylinositol 4,5 bisphosphate (PIP2) 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 PIP2 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 µM) 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.

Key Words: prenylation • membrane • GBD • neutrophil • thymus



Abbreviations used in this paper: CF-PE, carboxyfluorescein-labeled PE; GBD, GTPase binding domain; GST, glutathione-S-transferase; HSS, high speed supernatant; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PIP2, phosphatidylinositol 4,5 bisphosphate; PS, phosphatidylserine; PMN, polymorphonuclear leukocytes; WA, WASp/Scar protein COOH-terminal 70–100 amino acids; WASp, Wiskott-Aldrich Syndrome protein.


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   Abstract »    Full Text »    PDF »
Phosphorylation of Tyrosine 291 Enhances the Ability of WASp to Stimulate Actin Polymerization and Filopodium Formation.
G. O. C. Cory, R. Garg, R. Cramer, and A. J. Ridley (2002)
J. Biol. Chem. 277, 45115-45121
   Abstract »    Full Text »    PDF »
Motility Determinants in WASP Family Proteins.
D. Yarar, J. A. D'Alessio, R. L. Jeng, and M. D. Welch (2002)
Mol. Biol. Cell 13, 4045-4059
   Abstract »    Full Text »    PDF »
Lateral Sequestration of Phosphatidylinositol 4,5-Bisphosphate by the Basic Effector Domain of Myristoylated Alanine-rich C Kinase Substrate Is Due to Nonspecific Electrostatic Interactions.
J. Wang, A. Gambhir, G. Hangyas-Mihalyne, D. Murray, U. Golebiewska, and S. McLaughlin (2002)
J. Biol. Chem. 277, 34401-34412
   Abstract »    Full Text »    PDF »
Remodeling of organelle-bound actin is required for yeast vacuole fusion.
G. Eitzen, L. Wang, N. Thorngren, and W. Wickner (2002)
J. Cell Biol. 158, 669-679
   Abstract »    Full Text »    PDF »
Normal Arp2/3 complex activation in platelets lacking WASp.
H. Falet, K. M. Hoffmeister, R. Neujahr, and J. H. Hartwig (2002)
Blood 100, 2113-2122
   Abstract »    Full Text »    PDF »

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