Functional suppression of E-cadherin by protein kinase Cdelta.
Protein kinase C (PKC) delta, a member of the novel PKC subfamily, has been shown to have an important role in cell proliferation, differentiation, apoptosis and cell motility. In this study, we investigated the effect of green fluorescent protein (GFP)-PKCdelta and GFP-PKCalpha on cell-cell junctions of Madin-Darby canine kidney (MDCK) cells and found that only GFP-PKCdelta suppressed the homophilic interactions between the ectodomains of E-cadherins, accompanied by a weaker cell-cell adhesion. The kinase-deficient mutant of GFP-PKCdelta retained its localization at cell-cell junctions but failed to suppress the function of E-cadherin. In addition, we demonstrated that the hinge region (residues 280-347) that links the regulatory domain and the catalytic domain of PKCdelta is essential for both its kinase activity and the targeting of cell-cell junctions. A PKCdelta mutant with the deletion of amino acids 280-323 within the hinge region, which is catalytically active but defective in the targeting of cell-cell junctions, failed to suppress the function of E-cadherin. Moreover, expression of GFP-PKCdelta in MDCK cells expedited the detachment of cells from their neighbors and facilitated cell scatter induced by hepatocyte growth factor. By contrast, the GFP-PKCdelta mutants including the kinase-deficient mutant and the truncated mutant lacking residues 280-323 suppressed hepatocyte-growth-factor-induced cell scattering. Finally, siRNA-mediated knockdown of endogenous PKCdelta in MDCK cells was found to delay the onset of cell-cell detachment and cell scattering induced by hepatocyte growth factor. Taken together, our results demonstrate that the catalytic activity of PKCdelta and its localization to cell-cell junctions are necessary for PKCdelta to suppress the function of E-cadherin, which thereby facilitates scattering of epithelial cells in response to extracellular cues.
Chen
Chien-Lin
CL
Department of Life Science and Graduate Institute of Biomedical Sciences, National Chung Hsing University, Taichung 40227, Taiwan.
Chen
Hong-Chen
HC
D000818:Animals; D015820:Cadherins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D020134:Catalytic Domain / Q000502:physiology; D002448:Cell Adhesion; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D004285:Dogs; D049452:Green Fluorescent Proteins; D017228:Hepatocyte Growth Factor / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects / Q000201:enzymology; D007365:Intercellular Junctions / Q000201:enzymology*; D051571:Protein Kinase C-alpha / Q000378:metabolism; D051745:Protein Kinase C-delta / Q000737:chemistry / Q000378:metabolism*; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins
D015820:Cadherins; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D017228:Hepatocyte Growth Factor; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta