FUNCTIONAL-CHARACTERIZATION OF HYBRID RECEPTORS COMPOSED OF A TRUNCATED INSULIN-RECEPTOR AND WILD-TYPE INSULIN-LIKE GROWTH-FACTOR-1 OR INSULIN-RECEPTORS

To assess the characteristics of hybrid receptors composed of one kinase-inactive alpha beta-insulin half-receptor and one endogenous alpha beta-insulin-like growth factor I (IGF-1) or insulin half-receptor, a cell Line expressing an insulin receptor truncated by 365 amino acids (HIR Delta 978) was studied, which lacks most of the cytoplasmic beta-subunit. Analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis under nonreducing conditions revealed four distinct receptor species: endogenous receptors, the more rapidly migrating HIR Delta 978 homodimer, and two intermediate species representing HIR Delta 978/IGF-1 hybrid receptors and HIR Delta 978/IR hybrid receptors. In vivo ligand-binding affinity of the hybrid receptors was studied by receptor-ligand cross-linking, and the Delta 978/IGF-1R hybrid receptor was found to have a high affinity for IGF-1, whereas its affinity for insulin was low. Autophosphorylation studies of lectin-purified receptors revealed that neither the HIR Delta 978 holoreceptor nor the hybrid receptors underwent autophosphorylation in response to either ligand, despite the presence of intact IGF-1 or insulin half-receptors in the hybrids. Neither hybrid receptor underwent Ligand-induced endocytosis, as assessed with the bioactive photoaffinity probes B2(2-nitro-4-azidophenylacetyl)-des-Phe(B1) -insulin and N-epsilon B28-monoazidobenzoyl-IGF-1. In conclusion, the HIR Delta 978/IGF-1R hybrid receptor has a high in vivo affinity for IGF-1 but not for insulin. Neither the Delta 978/IGF-1R nor the Delta 978/IR hybrids undergo autophosphorylation or ligand-induced endocytosis in response to either ligand, indicating that intramolecular trans-, rather than cis; signal transduction is important in mediating autophosphorylation and endocytosis.