Differential phosphorylation of IRS-1 by insulin and insulin-like growth factor I receptors in Chinese hamster ovary cells

Insulin receptor (IR) and insulin-like growth factor I receptor (IGF-IR) ar e closely related receptor tyrosine kinases. Despite their high degree of h omology, recent evidence suggests that the two receptors have distinct biol ogical roles. In several recent studies, the cytoplasmic tyrosine kinase do mains of the two receptors have been shown to possess different signalling specificities. In this study, we examine the hypothesis that differential p hosphorylation of insulin receptor substrate 1 (IRS-1) may contribute to th ese differences in signalling between the two receptors. Using Chinese hams ter ovary (CHO) cells stably expressing human IR or IGF-IR and activated by their respective ligands, we show that there are differences between the t wo receptors with regard to the complement of SH2-containing proteins recru ited to IRS-1. In particular, IGF-IR appears to couple IRS-1 preferentially to Grb2 whereas, in contrast, IP, appears to couple IRS-1 preferentially t o the p85 subunit of phosphatidyl inositol 3-kinase (PI3-kinase) and to Nck . The two receptors couple IRS-1 equally to the tyrosine phosphatase SHP2. We have also generated phosphospecific antibodies to three important tyrosi ne phosphorylation sites on IRS-1 (pY608, pY895 and pY1172). We used these antibodies to probe the phosphorylation status of these sites in intact CHO /IR and CHO/IGF-IR cells, In the case of pY608, these results also show evi dence for differential phosphorylation of IRS-1 by the two receptors, Taken together, the results presented here support the notion that the cytoplasm ic domains of IR and IGF-IR have differences in their intrinsic signalling potentials.