Characterization of hepatocyte-growth-factor receptors on Meth A cells.
Hepatocyte growth factor (HGF) is a heparin-binding polypeptide mitogen for a variety of cell types including hepatocytes. HGF also has cytotoxic activity on some tumor cell lines as well as scattering activity on epithelial cells. In this study, recombinant human HGF was used to identify HGF-binding cell surface receptors on Meth A cells, whose growth is inhibited by HGF. Scatchard analysis of binding data indicated that there were two classes of binding sites with high affinity (Kd = 17 pM) and low affinity (Kd = 6.7 nM) and the average numbers were 6600 and 2,600,000 per cell, respectively. Affinity cross-linking of 125I-HGF to Meth A cells resulted in a major and a minor specifically labeled complex. Competition analysis followed by cross-linking indicated that the HGF-binding proteins were involved in the formation of the high-affinity binding. The existence of the two HGF-binding surface proteins was confirmed by HGF-dependent immunoprecipitation of the binding proteins with an anti-HGF polyclonal antibody. The molecular masses of the major and the minor surface proteins were 160 kDa and 130 kDa, respectively. The 160-kDa protein was autophosphorylated in vitro on tyrosine residue and was immunoprecipitated with an antiserum against the c-met proto-oncogene product. These results indicate that the 160-kDa HGF-binding surface protein on Meth A cells is the c-met protein. Furthermore, tyrosine phosphorylation of the c-met protein was stimulated by HGF treatment of Meth A cells, suggesting that it may be involved in the signal transduction of the growth inhibition of Meth A cells by HGF.
Komada
M
M
Institute for Liver Research, Kansai Medical University, Osaka, Japan.
Miyazawa
K
K
Ishii
T
T
Kitamura
N
N
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001345:Autoradiography; D001665:Binding Sites; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D015180:Electrophoresis, Gel, Two-Dimensional; D006133:Growth Substances / Q000378:metabolism; D017228:Hepatocyte Growth Factor; D006801:Humans; D051379:Mice; D010766:Phosphorylation; D011233:Precipitin Tests; D000090063:Proto-Oncogene Mas; D019859:Proto-Oncogene Proteins c-met; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012513:Sarcoma, Experimental / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000378:metabolism*
D000596:Amino Acids; D003432:Cross-Linking Reagents; D006133:Growth Substances; C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D017228:Hepatocyte Growth Factor; D019859:Proto-Oncogene Proteins c-met