Hepatocyte growth factor/scatter factor stimulates the Ras-guanine nucleotide exchanger

Hepatocyte growth factor/scatter factor (HGF/SF) induces mitogenesis and cell dissociation upon binding to the protein-tyrosine kinase receptor encoded by the MET proto-oncogene (p190^MET). The signal transduction pathways downstream from the receptor activation are largely unknown. We show that HGF/SF activates Ras protein. HGF/SF stimulation of metabolically labeled A549 cells raised the amount of Ras-bound radiolabeled guanine nucleotides by over 5-fold. Furthermore, following HGF/SF stimulation of these cells, 50% of Ras was in the GTP-bound active state. The uptake by Ras of radiolabeled GTP was also increased by 5-fold following HGF/SF stimulation in digitonin-permeabilized A549 cells. Moreover, HGF/SF treatment of A549 cells leads to stimulation of the cytosolic Ras-guanine nucleotide exchange activity, measured as accelerated release of [³H]GDP from purified recombinant Ras protein in vitro, in a dose- and time-dependent manner. Likewise, treatment with the protein-tyrosine kinase inhibitor 3-(1′,4′dihydroxytetralyl)methylene-2-oxindole of GTL-16 cells (featuring a p190^MET receptor constitutively active) significantly decreased the cytosolic Ras-guanine nucleotide exchange activity. These data demonstrate that HGF/ SF activates Ras protein by shifting the equilibrium toward the GTP-bound state and increases the uptake of guanine nucleotides by Ras, through mechanism(s) including the activation of a Ras-guanine nucleotide exchanger.