c phosphorylation and activation of the IGF-I receptor in src-transformed cells.

Using a panel of src mutants partially defective for malignant transformation, our laboratory has previously identified the insulin-like growth factor (IGF-I) receptor as a protein whose tyrosine phosphorylation correlates with transformation by src in embryonic chick cells (Kozma et al., 1990; Kozma and Weber, 1990). It has not been clear, however, whether src-induced phosphorylation altered the enzymatic or signaling properties of the IGF-I receptor and thus whether the IGF-I receptor could be a functionally significant target for pp60v-src. To examine the effect of src expression on the activity of the IGF-I receptor, the human IGF-I receptor was expressed in Rat-1 fibroblasts co-expressing the temperature-sensitive v-src mutant, tsLA29. The IGF-I receptor exhibited an elevated level of tyrosine phosphorylation in src transformed cells even in the absence of IGF-I treatment. Increased receptor phosphorylation occurred rapidly when cells expressing a temperature-conditional src mutant were shifted from the restrictive to the permissive temperature. Src-induced phosphorylation of the receptor was correlated with an increase in the in vitro tyrosine kinase activity of the receptor, both toward itself and exogenous substrates. The src-induced increase in receptor activity was shown to be dependent on tyrosine phosphorylation, as treatment with a tyrosine-specific phosphatase lowered receptor activity. A kinase-defective mutant of the IGF-I receptor also became constitutively phosphorylated in src-transformed cells, ruling out a possible autocrine mechanism for this phosphorylation. Collectively these data indicate that pp60v-src induces ligand-independent phosphorylation and activation of the IGF-I receptor by an intracellular mechanism, consistent with the possibility that receptor phosphorylation could contribute to the genesis of the transformed phenotype.