N-WASP, a novel actin-depolymerizing protein, regulates the cortical cytoskeletal rearrangement in a PIP2-dependent manner downstream of tyrosine kinases.
Here we identify a 65 kDa protein (N-WASP) from brain that binds the SH3 domains of Ash/Grb2. The sequence is homologous to Wiskott-Aldrich syndrome protein (WASP). N-WASP has several functional motifs, such as a pleckstrin homology (PH) domain and cofilin-homologous region, through which N-WASP depolymerizes actin filaments. When overexpressed in COS 7 cells, the wild-type N-WASP causes several surface protrusions where N-WASP co-localizes with actin filaments. Epidermal growth factor (EGF) treatment induces the complex formation of EGF receptors and N-WASP, and produces microspikes. On the other hand, two mutants, C38W (a point mutation in the PH domain) and deltaVCA (deletion of the actin binding domain), localize predominantly in the nucleus and do not cause a change in the cytoskeleton, irrespective of EGF treatment. Interestingly, the C38W PH domain binds less effectively to phosphatidylinositol 4,5-bisphosphate (PIP2) than the wild-type PH domain. These results suggest the importance of the PIP2 binding ability of the PH domain and the actin binding for retention in membranes. Collectively, we conclude that N-WASP transmits signals from tyrosine kinases to cause a polarized rearrangement of cortical actin filaments dependent on PIP2.
Department of Biochemistry, Institute of Medical Science, University of Tokyo, Shirokanedai, Japan.
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D000199:Actins / Q000187:drug effects / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001923:Brain Chemistry; D019556:COS Cells; D002147:Calmodulin / Q000378:metabolism; D002417:Cattle; D002462:Cell Membrane / Q000737:chemistry; D003001:Cloning, Molecular; D003599:Cytoskeleton / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism; D051380:GRB2 Adaptor Protein; D008969:Molecular Sequence Data; D009154:Mutation; D009419:Nerve Tissue Proteins; D009928:Organ Specificity; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000502:physiology; D051316:Wiskott-Aldrich Syndrome Protein; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D002147:Calmodulin; D051380:GRB2 Adaptor Protein; D009419:Nerve Tissue Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D011506:Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D051316:Wiskott-Aldrich Syndrome Protein; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases