THE PLECKSTRIN HOMOLOGY DOMAIN IN INSULIN-RECEPTOR SUBSTRATE-1 SENSITIZES INSULIN SIGNALING

The NH2 terminus of insulin receptor substrate-1 (IRS-1) contains a pleckstrin homology (PH) domain. We deleted the PH domain in IRS-1 (IRS-1(Delta PH)) and expressed the mutant in Chinese hamster ovary and 32D cells. During insulin stimulation, IRS-1(Delta PH) is poorly tyrosine-phosphorylated in CHO cells, but undergoes serine/threonine phosphorylation. Similarly, IRS-1(Delta PH) fails to undergo insulin-stimulated tyrosine phosphorylation in 32D cells, which uncouples the activation of phosphatidylinositol 3'-kinase and p70(s6k) from the endogenous insulin receptors. Overexpression of the insulin receptor in 32D(IR) cells, however, restores tyrosine phosphorylation of IRS-1(Delta PH) and rescues insulin responses including mitogenesis. Thus, while the PH domain is not required for the engagement of downstream signals, it is one of the elements in the NH2 terminus of IRS-1 that is needed for a sensitive coupling to insulin receptors, especially at ordinary receptor levels found in most cells and tissues.