Impact of the regulation of PI3-kinase product on insulin action and its role in the disease state.

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PI3-kinase plays an important role in various metabolic actions of insulin including glucose uptake and glycogen synthesis. Although PI3-kinase primarily functions as a lipid kinase which preferentially phosphorylates the D-3 position of phospholipids, the effect of hydrolysis of the key PI3-kinase product PI (3, 4, 5) P3 on these biological responses is unknown. We have cloned rat SHIP2 cDNA which possesses the 5'-phosphatase activity to hydrolyze PI (3, 4, 5) P3 to PI (3, 4) P2 and which is mainly expressed in the target tissues of insulin. To study the role of SHIP2 in insulin signaling, wild-type (WT)- and 5'-phosphatase defective SHIP2 (ΔIP-SHIP2) were overexpressed in 3T3-L1 adipocytes and L6 myocytes by means of adenovirus mediated gene transfer. Early events of insulin signaling including insulin-induced tyrosine phosphorylation of the insulin receptor p-subunit and IRS-1, IRS-1 association with p85 subunit, and PI3-kinase activity were not affected by expression of either WT- … More or ΔIP-SHIP2. As expected from possessing the 5'-phosphatase catalytic region, insulin-induced PI (3, 4, 5) P3 production was markedly decreased by overexpression of WT-SHIP2. In contrast, the amount of PI (3, 4, 5) P3 was oppositely increased by expression of ΔIP-SHIP2, indicating that ΔIP-SHIP2 functions in a dominant negative manner in 3T3-L1 adipocytes. Both PI (3, 4, 5) P3 and PtdIns (3, 4) P2 were known to possibly activate downstream targets Akt and PKCλ in vitro. Importantly, expression of WT-SHIP2 inhibited insulin-induced activation of Akt and PKCλ, whereas these activations were increased by expression of ΔIP-SHIP2 in vivo. Consistent with the regulation of downstream molecules of PI3-kinase, insulin-induced 2-deoxyglucose uptake and Glut4 translocation were decreased by expression of WT-SHIP2 and increased by expression of ΔIP-SHIP2. In addition, insulin-induced phosphorylation of GSK-3β and activation of PP1 followed by activation of glycogen synthase and glycogen synthesis were decreased by expression of WT-SHIP2, and these insulin actions were increased by the expression of ΔIP-SHIP2. These results indicate that SHIP2 negatively regulates metabolic signaling of insulin via the 5'-phosphatase activity. Less