The insulin-like growth factor I receptor-induced interaction of insulin receptor substrate-4 and Crk-II

Stimulation of the insulin or insulin-like growth factor (IGF)-I receptor r esults in activation of several signaling pathways. Proteins of the insulin receptor substrate (IRS) family play important roles in mediating these si gnaling cascades. To date, four members of the IRS family of docking protei ns have been characterized. Recently, we have reported that stimulation of the IGF-I receptor in 293 HEK cells regulates interaction of the newly disc overed IRS-4 molecule with the Crk. family of proteins. In the present stud y, we characterize the molecular basis of these interactions. C- and N term ini truncation analysis of IRS-4 demonstrated that the region between amino acids 678 and 800 of the IRS-4 molecule is involved in this interaction. T his region contains a cluster of four tyrosines (Y-700, Y-717, Y-743, and Y -779). We hypothesize that one or more of these tyrosines are involved in t he interaction between the SH2 domain of the Crk-II molecule when IRS-4 is phosphorylated upon IGF-I receptor activation. Additional mutational analys es confirmed this hypothesis. Interestingly, none of these four tyrosines w as individually critical for the interaction between Crk-II and IRS-4, but when all four tyrosines were simultaneously mutated to phenylalanine, the I GF-I induced interaction between these molecules was abolished. Taken toget her, these results suggest a novel mechanism of Crk-II binding to tyrosine phosphorylated proteins.