title: Binding of activated alpha2-macroglobulin to its cell surface receptor GRP78 in 1-LN prostate cancer cells regulates PAK-2-dependent activation of LIMK.
abstract: Two characteristics of highly malignant cells are their increased motility and secretion of proteinases allowing these cells to penetrate surrounding basement membranes and metastasize. Activation of 21-kDa activated kinases (PAKs) is an important mechanism for increasing cell motility. Recently, we reported that binding of receptor-recognized forms of the proteinase inhibitor alpha2-macroglobulin (alpha2M*) to GRP78 on the cell surface of 1-LN human prostate cancer cells induces mitogenic signaling and cellular proliferation. In the current study, we have examined the ability of alpha2M* to activate PAK-1 and PAK-2. Exposure of 1-LN cells to alpha2M* caused a 2- to 3-fold increase in phosphorylated PAK-2 and a similar increase in its kinase activity toward myelin basic protein. By contrast, the phosphorylation of PAK-1 was only negligibly affected. Silencing the expression of the GRP78 gene, using either of two different mRNA sequences, greatly attenuated the appearance of phosphorylated PAK-2 in alpha2M*-stimulated cells. Treatment of 1-LN cells with alpha2M* caused translocation of PAK-2 in association with NCK to the cell surface as evidenced by the co-immunoprecipitation of PAK-2 and NCK in the GRP78 immunoprecipitate from plasma membranes. alpha2M*-induced activation of PAK-2 was inhibited by prior incubation of the cells with specific inhibitors of tyrosine kinases and phosphatidylinositol 3-kinase. PAK-2 activation was accompanied by significant increases in the levels of phosphorylated LIMK and phosphorylated cofilin. Silencing the expression of the PAK-2 gene greatly attenuated the phosphorylation of LIMK. In conclusion, we show for the first time the activation of PAK-2 in 1-LN prostate cancer cells by a proteinase inhibitor, alpha2-macroglobulin. These studies suggest a mechanism by which alpha2M* enhances the metastatic potential of these cells.
authors.affiliation: Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: Uma Kant
authors.initials: UK
authors.lastname: Misra
authors.forename: Rohit
authors.initials: R
authors.lastname: Deedwania
authors.forename: Salvatore Vincent
authors.initials: SV
authors.lastname: Pizzo
mesh_terms: D051339:Actin Depolymerizing Factors; D000199:Actins / Q000737:chemistry; D002352:Carrier Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism*; D002465:Cell Movement; D049109:Cell Proliferation; D003599:Cytoskeleton / Q000378:metabolism; D000091342:Endoplasmic Reticulum Chaperone BiP; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D054461:Lim Kinases; D008297:Male; D008840:Microfilament Proteins / Q000378:metabolism; D008954:Models, Biological; D018832:Molecular Chaperones / Q000378:metabolism; D004676:Myelin Basic Protein / Q000378:metabolism; D009362:Neoplasm Metastasis; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D020033:Protein Isoforms; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D021381:Protein Transport; D012330:RNA, Double-Stranded / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D000511:alpha-Macroglobulins / Q000737:chemistry / Q000378:metabolism*; D051025:bcl-Associated Death Protein; D054462:p21-Activated Kinases; D020744:rac GTP-Binding Proteins / Q000737:chemistry
chemical_list: D051339:Actin Depolymerizing Factors; D000199:Actins; D002352:Carrier Proteins; D000091342:Endoplasmic Reticulum Chaperone BiP; D004791:Enzyme Inhibitors; D006360:Heat-Shock Proteins; D054461:Lim Kinases; D008840:Microfilament Proteins; D018832:Molecular Chaperones; D004676:Myelin Basic Protein; D019869:Phosphatidylinositol 3-Kinases; D020033:Protein Isoforms; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D000511:alpha-Macroglobulins; D051025:bcl-Associated Death Protein; D054462:p21-Activated Kinases; D020744:rac GTP-Binding Proteins; C494819:BAD protein, human; C000717247:HSPA5 protein, human; C515892:LIMK1 protein, human; C515907:PAK2 protein, human