Inhibition of junction assembly in cultured epithelial cells by hepatocyte growth factor/scatter factor is concomitant with increased stability and altered phosphorylation of the soluble junctional molecules.
Hepatocyte growth factor/scatter factor (HGF/SF) is a mesenchymally derived glycoprotein with a strong scattering effect on epithelial cells. A receptor tyrosine kinase encoded by the met proto-oncogene has been identified as the cellular receptor for HGF/SF. Following stimulation with HGF/SF, cell scattering occurs concurrent with decreased cell-cell adhesion and disassembly of junctional components. In culture, junction formation is cell-cell contact dependent and can be regulated by modulating the Ca2+ concentrations of the growth media. Decreasing the Ca2+ concentrations below 50 microM causes rapid disassembly of junctions, whereas increasing the Ca2+ concentrations to 1.8 mM induces cell-cell contact and junction assembly. Although associated with decreased cell-cell adhesion and disassembly of the junctional complex, HGF/SF-induced scattering occurs under high extracellular Ca2+ concentrations. To gain insight into the mechanisms of HGF/SF-induced scattering of epithelial cells, we have studied the effect(s) of HGF/SF on junction assembly by examining the solubility, stability, phosphorylation, and subcellular localization of the major components of the adhering junctions, plakoglobin (Pg) and E-cadherin, in Madin-Darby canine kidney (MDCK) epithelial cells and in a MDCK cell line expressing an exogenous chimeric met receptor (CSF-MET) that scatters in response to colony-stimulating factor 1 (CSF-1). The results have shown that in HGF/SF-stimulated MDCK cells, adhering junctions were not assembled upon induction of cell-cell contact. Immunofluorescence analyses showed that larger amounts of Pg and E-cadherin were Triton X-100 extractable, and more significantly, these proteins were homogeneously distributed along the membrane and were not concentrated at the areas of cell-cell contact. Similar results were obtained for CSF-MET expressing MDCK cells in response to CSF-1. In contrast, none of the above effects were detected in MDCK cells expressing a mutant CSF-MET chimera containing a phenylalanine substitution at tyrosine 1356 in met, which fails to scatter in response to CSF-1. When compared with the unstimulated cells, the inhibition of cell adhesion promoted by HGF/SF correlated with an increased stability of the newly synthesized soluble E-cadherin and Pg and an altered phosphorylation pattern of E-cadherin, as determined by partial proteolytic peptide mapping.
Pasdar
M
M
Department of Anatomy and Cell Biology, University of Alberta, Edmonton, Canada. mpasdar@anat.med.ualberta.ca
Li
Z
Z
Marreli
M
M
Nguyen
B T
BT
Park
M
M
Wong
K
K
D000818:Animals; D015820:Cadherins / Q000378:metabolism; D002448:Cell Adhesion / Q000187:drug effects; D015815:Cell Adhesion Molecules / Q000378:metabolism; D003115:Colony-Stimulating Factors / Q000235:genetics / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D051180:Desmoplakins; D003896:Desmosomes / Q000378:metabolism; D004285:Dogs; D004847:Epithelial Cells; D004848:Epithelium / Q000187:drug effects; D017228:Hepatocyte Growth Factor / Q000494:pharmacology*; D007365:Intercellular Junctions / Q000187:drug effects*; D016173:Macrophage Colony-Stimulating Factor / Q000378:metabolism; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D019859:Proto-Oncogene Proteins c-met; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D051185:gamma Catenin
D015820:Cadherins; D015815:Cell Adhesion Molecules; D003115:Colony-Stimulating Factors; D003598:Cytoskeletal Proteins; D051180:Desmoplakins; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D051185:gamma Catenin; D017228:Hepatocyte Growth Factor; D016173:Macrophage Colony-Stimulating Factor; D019859:Proto-Oncogene Proteins c-met; D020794:Receptor Protein-Tyrosine Kinases