STRUCTURAL AND FUNCTIONAL DOMAINS CRITICAL FOR CONSTITUTIVE ACTIVATION OF THE HGF-RECEPTOR (MET)

The MET gene, encoding the tyrosine kinase receptor for Hepatocyte Gro wth Factor, is a potentially harmful oncogene overexpressed ina signif icant fraction of human cancers. To study the molecular mechanisms res ponsible for oncogenic activation, the biochemical and biological prop erties of a number of MET constructs were analysed. The native heterod imeric receptor (alpha beta), the beta chain alone, as well as a kinas e defective mutant did not transform rodent fibroblasts upon transfect ion. The cytoplasmic domain, truncated immediately below the transmemb rane region, acquired constitutive tyrosine kinase activity in vivo, p roduced foci of transformation, and was tumorigenic in nude mice. Remo val of the first 39 amino acids of the juxtamembrane domain resulted i n loss of constitutive activation in vivo and transforming potential, without impairment of the in vitro kinase activity. Replacement of the juxtamembrane domain with 5' TPR sequences restored constitutive kina se activation and transforming properties. Site-directed mutagenesis o f either of the two tyrosine residues involved in the positive regulat ion of the catalytic activity upon phosphorylation (Y-1234 or Y-1235 i n the kinase domain of the HGF receptor), strongly impaired TPR-MET tr ansforming potential. These data show that: (1) the truncated cytoplas mic HGF receptor has constitutive kinase activity and is oncogenic; (2 ) the first 39 amino acids of the juxtamembrane domain and (3) the reg ulatory tyrosines in the catalytic domain are required to unleash its transforming potential.