DISTINCT MODES OF INTERACTION OF SHC AND INSULIN-RECEPTOR SUBSTRATE-1WITH THE INSULIN-RECEPTOR NPEY REGION VIA NON-SH2 DOMAINS

Insulin receptor substrate 1 (IRS-1) and src homology and collagen pro tein (SHC) are signaling proteins which are rapidly phosphorylated on tyrosines after insulin receptor (IR) activation. We have recently sho wn that both SHC and IRS-1 interact with the tyrosine-phosphorylated N PEY motif of the PR and insulin-like growth factor I receptor via non- SH2 domains (Gustafson, T. A., He, W., Craparo, A., Schaub, C. D., and O'Neill, T. J. (1995) Mel. Cell. Biol. 15, 2500-2508; O'Neill, T. J., Craparo, A., and Gustafson, T. A. (1994) Mel. Cell. Biol. 14, 6433-64 42; Craparo, A., O'Neill, T. J., and Gustafson, T. A. (1995) J. Biol. Chem. 270, 15639-15643). In this study we characterize these interacti ons by examining the effects of 18 amino acid substitutions within and around the IR NPEY motif upon interaction with SHC and IRS 1. We conf irm that Tyr-960 within the NPEY motif of the IR is essential for both IRS-1 and SHC interaction and that Asn 957 and Pro-958 are essential for IRS-1 interaction and important but not critical for SHC interacti on. Additional mutations surrounding the NPEY motif revealed completel y distinct patterns of interaction for SHC and IRS-1. Specifically, mu tation of Leu-952 or Tyr-953 (at positions -7 and -8 from Tyr-960) mar kedly reduced IRS-1 interaction but had no effect upon SHC interaction . Likewise, mutation of Ala-963 (+3) reduced IRS-1 but not SHC interac tion. Conversely, substitution of Leu-961 (+1) with either Ala or Arg reduced SHC interaction by 70 and 90%, respectively, yet had no effect upon interaction with IRS-1. Our data show that the sequences within and surrounding the NPEY contribute differentially to either SHC or IR S-1 recognition. Our findings suggest mechanisms by which the differen tial interaction of known receptors with IRS-1 and SHC may be mediated .