Abstract
The acidic (A) region of WASp family proteins is thought to represent a high-affinity binding site for Arp2/3 complex without activating properties. Here we show that GST-fused WASp-A and N-WASP-A, but not a WASP-A/W500S mutant, several truncated WASp-A constructs or WAVE1-A can pull down Arp2/3 complex from cell lysates. Significantly, WASp-A and N-WASP-A synergistically trigger formation of filopodia or lamellipodia when coinjected with sub-effective concentrations of V12CDC42Hs or V12Rac1, respectively, into macrophages. The ability of WASp family A region constructs to induce this effect is closely correlated with their ability to bind Arp2/3 complex in vitro. These results imply that (i) Arp2/3 complex is critically involved in filopodia and lamellipodia formation in macrophages and (ii) acidic regions of WASp and N-WASP are not simply binding sites for Arp2/3 complex but can prime it for RhoGTPase-triggered signals leading to actin nucleation.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Actin-Related Protein 2
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Actin-Related Protein 3
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Actins / metabolism
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Cells, Cultured
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Cyclin-Dependent Kinases / genetics
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Cyclin-Dependent Kinases / metabolism*
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Cyclin-Dependent Kinases / pharmacology
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Cytoskeletal Proteins*
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Fluorescence Polarization
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Glutathione Transferase / genetics
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Humans
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Macromolecular Substances
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Macrophages / cytology
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Macrophages / drug effects
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Macrophages / metabolism
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Microfilament Proteins / genetics
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Microfilament Proteins / metabolism
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Microinjections
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Microscopy, Fluorescence
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Mutagenesis, Site-Directed
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Nerve Tissue Proteins / genetics
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Nerve Tissue Proteins / metabolism*
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Nerve Tissue Proteins / pharmacology
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Protein Binding / physiology
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Proteins / genetics
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Proteins / metabolism*
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Proteins / pharmacology
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Pseudopodia / drug effects
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Pseudopodia / metabolism*
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Recombinant Fusion Proteins / administration & dosage
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Wiskott-Aldrich Syndrome Protein
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Wiskott-Aldrich Syndrome Protein Family
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Wiskott-Aldrich Syndrome Protein, Neuronal
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rac1 GTP-Binding Protein / metabolism*
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rac1 GTP-Binding Protein / pharmacology
Substances
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ACTR2 protein, human
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ACTR3 protein, human
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Actin-Related Protein 2
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Actin-Related Protein 3
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Actins
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Cytoskeletal Proteins
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Macromolecular Substances
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Microfilament Proteins
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Nerve Tissue Proteins
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Proteins
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Recombinant Fusion Proteins
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WAS protein, human
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WASL protein, human
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Wiskott-Aldrich Syndrome Protein
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Wiskott-Aldrich Syndrome Protein Family
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Wiskott-Aldrich Syndrome Protein, Neuronal
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Glutathione Transferase
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Cyclin-Dependent Kinases
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rac1 GTP-Binding Protein