MUTATION OF A CONSERVED AMINO-ACID RESIDUE (TRYPTOPHAN-1173) IN THE TYROSINE KINASE DOMAIN OF THE IGF-I RECEPTOR ABOLISHES AUTOPHOSPHORYLATION BUT DOES NOT ELIMINATE BIOLOGIC FUNCTION

The amino acid sequence of the tyrosine kinase domain of the insulin-l ike growth factor-I (IGF-I) receptor is 84% identical to the sequence of the analogous region of the insulin receptor, A naturally occurring mutation of the tryptophan residue at position 1200 of the insulin re ceptor to serine results in impaired beta subunit autophosphorylation of wheat germ agglutinin-purified receptors, severely impaired thymidi ne incorporation and moderately reduced glycogen synthesis; however, g lucose uptake was unaffected. To study the importance of this residue in IGF-I receptor function, we mutated the analogous tryptophan residu e at position 1173 of the IGF-I receptor to serine and overexpressed t he mutant receptor in NIH-3T3 cells, In cell Lines overexpressing this mutant IGF-I receptor, beta subunit autophosphorylation was severely reduced. Additionally, the overexpressed mutant receptors exhibited a dominant-negative effect on IGF-I-stimulated autophosphorylation of en dogenous mouse IGF-I receptors, Phosphorylation of insulin receptor su bstrate (IRS)-1 in intact cells by the mutant IGF-I receptors was simi lar to the level of IRS-1 phosphorylation seen in the parental NIH-3T3 cells, but there was no obvious dominant-negative effect on IRS-1 pho sphorylation. Wheat germ agglutinin-purified mutant receptors were as active in phosphorylating poly(Glu,Tyr) 4:1 as wild-type IGF-I recepto rs, suggesting that, in intact cells, additional factors are necessary in order for the IGF-I receptor to phosphorylate IRS-1, Thymidine inc orporation was severely reduced in one clone overexpressing the mutant IGF-I receptor and abolished in a second clone, Glucose uptake in bot h clones was reduced to about half of that seen in a cell line overexp ressing mild-type IGF-I receptors. Thus, we propose that the tryptopha n residue at position 1173 of the IGF-I receptor is important in the r egulation of autophosphorylation in vivo. This study again confirms th at high levels of autophosphorylation are not required for mediation o f all of the biologic activities of the IGF-I receptor.