Branching tubulogenesis but not scatter of Madin-Darby canine kidney cells requires a functional Grb2 binding site in the met receptor tyrosine kinase

Hepatocyte growth factor is a multifunctional cytokine that induces mitogenesis, motility, invasion, and branching tubulogenesis of several epithelial and endothelial cell lines in culture. The receptor for hepatocyte growth factor has been identified as the Met tyrosine kinase, To investigate the signaling pathways that are involved in these events, we have generated chimeric receptors containing the colony stimulating factor-1 receptor fused to the transmembrane and intracellular domains of the Met receptor. Madin-Darby canine kidney epithelial cells expressing the Met chimera dissociate scatter and form branching tubules in response to colony stimulating factor-1. From structure-function analyses, tyrosine residue 1356 within the carboxyl terminus of the Met receptor is critical for these events. The amino acid sequence downstream from tyrosine 1356 represents a consensus binding site for the Grb2 adaptor protein and forms a multisubstrate binding site for the p85 subunit of phosphatidylinositol 3-kinase, phospholipase C gamma, and the Shc adaptor protein. To distinguish which of these signaling pathways are required, we generated a mutant receptor that selectively fails to associate with the Grb2 adaptor protein. Cells expressing this mutant receptor scattered but were unable to form branching tubules, indicating that a Grb2 binding site in the Met receptor is critical for morphogenic responses.