GROWTH-FACTORS AND INSULIN STIMULATE TYROSINE PHOSPHORYLATION OF THE 51C SHIP2 PROTEIN/

Antibodies raised against the 51C/SHIP2 inositol polyphosphate 5'-phos phatase were used to examine the effects of growth factors and insulin on the metabolism of this protein. Immunoblot analysis revealed that the 51C/SHIP2 protein was widely expressed in fibroblast and nonhemato poietic tumor cell lines, unlike the SHIP protein, which was found onl y in cell lines of hematopoietic origin. The 51C/SHIP2 antiserum preci pitated a protein of approximately 145 kDa along with an activity whic h hydrolyzed phosphatidylinositol 3,4,5-trisphosphate to phosphatidyli nositol 3,4-bisphosphate. Tyrosine phosphorylation of the 51C/SHIP2 pr otein occurred in response to treatment of cells with epidermal growth (EGF), platelet-derived growth factor (PDGF), nerve growth factor (NG F), insulin-like growth factor-1 (IGF-1), or insulin. EGF and PDGF ind uced transient tyrosine phosphorylation of 51C/SHIP2, with maximal tyr osine phosphorylation occurring at 5-10 min following treatment and re turning to near basal levels within 20 min, In contrast, treatment of cells with NGF, IGF-1, or insulin resulted in prolonged tyrosine phosp horylation of 51C/SHIP2 protein, with 40-80% maximal phosphorylation s ustained for up to 2 h following agonist treatment. The kinetics of ac tivation of the Akt/PKB protein kinase by the various factors correlat ed well with the kinetics of tyrosine phosphorylation of 51C/SHIP2, EG F, NGF, and PDGF stimulated the association of 51C/SHIP2 protein with the Shc adapter protein; however, no Shc could be detected in 51C/SHIP 2-immune precipitates from cells treated with IGF-1 or insulin. The da ta suggest that 51C/SHIP2 may play a significant role in regulation of phosphatidylinositol 3'-kinase signaling by growth factors and insuli n.