Binding of activated alpha2-macroglobulin to its cell surface receptor GRP78 in 1-LN prostate cancer cells regulates PAK-2-dependent activation of LIMK.
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.
Misra
Uma Kant
UK
Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Deedwania
Rohit
R
Pizzo
Salvatore Vincent
SV
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
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