Constitutive activation of c-Met in liver metastatic B16 melanoma cells depends on both substrate adhesion and cell density and is regulated by a cytosolic tyrosine phosphatase activity

Serial selection in vivo for liver colonization of B16 murine melanoma cells consistently resulted in cell lines expressing elevated amounts of the hepatocyte growth factor/scatter factor receptor (c-Met), which is constitutively activated in the absence of its cognate ligand, In this paper we present evidence suggesting that c-Met constitutive activation in liver specific B16 melanoma cells depends on both receptor concentration on the cell surface and a cytosolic tyrosine phosphatase activity. In fact, c-Met constitutive activation is suddenly lost upon detachment of the cells from the substrate and is dramatically decreased in adherent cells plated at low density, The loss of tyrosine phosphorylation of c-Met in suspension appears to depend, at least partly, on an increased cytosolic tyrosine phosphatase activity, Instead, lower activation of c-Met at low density mostly results from a decrease in receptor concentration on the membrane, Moreover, we show that c-Met activation does not occur homogeneously on the surface of adherent cells, In fact, receptor concentration and activation appear to be higher on the ventral surface (adherent to the substrate) than on the apical surface, Upon detachment, compartmentalization is lost, leading to a decrease in average receptor density on the plasma membrane and hence to a lower activation.