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

pi3激酶在胰岛素的多种代谢活动中起重要作用，包括葡萄糖摄取和糖原合成。虽然pi3 -激酶主要作为一种脂质激酶，优先磷酸化磷脂的D-3位点，但pi3 -激酶关键产物PI (3,4,5) P3的水解对这些生物反应的影响尚不清楚。我们克隆了大鼠SHIP2 cDNA，该cDNA具有将PI (3,4,5) P3水解为PI (3,4) P2的5′-磷酸酶活性，主要表达于胰岛素靶组织中。为了研究SHIP2在胰岛素信号传导中的作用，通过腺病毒介导的基因转移，野生型（WT）和5'-磷酸酶缺陷型SHIP2 （ΔIP-SHIP2）在3T3-L1脂肪细胞和L6肌细胞中过表达。胰岛素信号的早期事件，包括胰岛素诱导的胰岛素受体p亚基和IRS-1的酪氨酸磷酸化，IRS-1与p85亚基的关联，以及pi3激酶活性，不受WT-…More或ΔIP-SHIP2表达的影响。正如预期的那样，由于拥有5'-磷酸酶催化区域，胰岛素诱导的PI (3,4,5) P3的产生通过过表达WT-SHIP2显着减少。相反，通过ΔIP-SHIP2的表达，PI (3,4,5) P3的数量相反增加，表明ΔIP-SHIP2在3T3-L1脂肪细胞中以显性负向方式发挥作用。已知PI (3,4,5) P3和PtdIns (3,4) P2可能在体外激活下游靶点Akt和PKCλ。重要的是，WT-SHIP2的表达抑制胰岛素诱导的Akt和PKCλ的激活，而这些激活在体内通过ΔIP-SHIP2的表达而增加。与pi3激酶下游分子的调控一致，胰岛素诱导的2-去氧葡萄糖摄取和Glut4易位通过WT-SHIP2的表达而减少，通过ΔIP-SHIP2的表达而增加。此外，胰岛素诱导的GSK-3β磷酸化、PP1活化、糖原合成酶活化和糖原合成通过WT-SHIP2的表达而减弱，而这些胰岛素作用通过ΔIP-SHIP2的表达而增强。这些结果表明SHIP2通过5'-磷酸酶活性负调控胰岛素代谢信号。少

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