title: Domain analysis of supervillin, an F-actin bundling plasma membrane protein with functional nuclear localization signals.
abstract: A growing number of actin-associated membrane proteins have been implicated in motile processes, adhesive interactions, and signal transduction to the cell nucleus. We report here that supervillin, an F-actin binding protein originally isolated from bovine neutrophil plasma membranes, contains functional nuclear targeting signals and localizes at or near vinculin-containing focal adhesion plaques in COS7-2 and CV1 cells. Overexpression of full-length supervillin in these cells disrupts the integrity of focal adhesion plaques and results in increased levels of F-actin and vinculin. Localization studies of chimeric proteins containing supervillin sequences fused with the enhanced green fluorescent protein indicate that: (1) the amino terminus promotes F-actin binding, targeting to focal adhesions, and limited nuclear localization; (2) the dominant nuclear targeting signal is in the center of the protein; and (3) the carboxy-terminal villin/gelsolin homology domain of supervillin does not, by itself, bind tightly to the actin cytoskeleton in vivo. Overexpression of chimeras containing both the amino-terminal F-actin binding site(s) and the dominant nuclear targeting signal results in the formation of large nuclear bundles containing F-actin, supervillin, and lamin. These results suggest that supervillin may contribute to cytoarchitecture in the nucleus, as well as at the plasma membrane.
authors.affiliation: Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA 01605, USA.
authors.forename: J D
authors.initials: JD
authors.lastname: Wulfkuhle
authors.forename: I E
authors.initials: IE
authors.lastname: Donina
authors.forename: N H
authors.initials: NH
authors.lastname: Stark
authors.forename: R K
authors.initials: RK
authors.lastname: Pope
authors.forename: K N
authors.initials: KN
authors.lastname: Pestonjamasp
authors.forename: M L
authors.initials: ML
authors.lastname: Niswonger
authors.forename: E J
authors.initials: EJ
authors.lastname: Luna
mesh_terms: D000199:Actins / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D019556:COS Cells; D002417:Cattle; D002448:Cell Adhesion; D002460:Cell Line; D003599:Cytoskeleton / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D034882:Lamins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008840:Microfilament Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins / Q000378:metabolism; D010641:Phenotype; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016596:Vinculin / Q000378:metabolism
chemical_list: D000199:Actins; D017931:DNA Primers; D034882:Lamins; D008164:Luminescent Proteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D016596:Vinculin; D049452:Green Fluorescent Proteins