Phosphatidylinositol (PI) 3-kinase plays an important role in various metabolic actions of insulin including glucose uptake and glycogen synthesis. Although PI 3-kinase primarily functions as a lipid kinase which preferentially phosphorylates the D-3 position of phospholipids, the effect of hydrolysis of the key PI 3-kinase product PI 3,4,5-triphosphate [PI(3,4,5)P3] on these biological responses is unknown. We recently cloned rat SH2-containing inositol phosphatase 2 (SHIP2) cDNA which possesses the 5-phosphatase activity to hydrolyze PI(3,4,5)P3 to PI 3,4-bisphosphate [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 SHIP2 (WT-SHIP2) and 5-phosphatase-defective SHIP2 (⌬IP-SHIP2) were overexpressed in 3T3-L1 adipocytes by means of adenovirus-mediated gene transfer. Early events of insulin signaling including insulin-induced tyrosine phosphorylation of the insulin receptor ␤ subunit and IRS-1, IRS-1 association with the p85 subunit, and PI 3-kinase activity were not affected by expression of either WT-SHIP2 or ⌬IP-SHIP2. Because WT-SHIP2 possesses the 5-phosphatase catalytic region, its overexpression marked by decreased insulin-induced PI(3,4,5)P3 production, as expected. In contrast, the amount of PI(3,4,5)P3 was increased by the expression of ⌬IP-SHIP2, indicating that ⌬IP-SHIP2 functions in a dominant-negative manner in 3T3-L1 adipocytes. Both PI(3,4,5)P3 and PI(3,4)P2 were known to possibly activate downstream targets Akt and protein kinase C in vitro. Importantly, expression of WT-SHIP2 inhibited insulin-induced activation of Akt and protein kinase C, whereas these activations were increased by expression of ⌬IP-SHIP2 in vivo. Consistent with the regulation of downstream molecules of PI 3-kinase, insulin-induced 2-deoxyglucose uptake and Glut4 translocation were decreased by expression of WT-SHIP2 and increased by expression of ⌬IP-SHIP2. In addition, insulin-induced phosphorylation of GSK-3␤ and activation of PP1 followed by activation of glycogen synthase and glycogen synthesis were decreased by expression of WT-SHIP2 and increased by the expression of ⌬IP-SHIP2. These results indicate that SHIP2 negatively regulates metabolic signaling of insulin via the 5-phosphatase activity and that PI(3,4,5)P3 rather than PI(3,4)P2 is important for in vivo regulation of insulin-induced activation of downstream molecules of PI 3-kinase leading to glucose uptake and glycogen synthesis.Insulin binding to the extracellular ␣ subunit of the insulin receptor activates the intrinsic tyrosine kinase activity of the intracellular ␤ subunit. The activated insulin receptor phosphorylates the insulin receptor substrate (IRS) family of proteins on the tyrosine residues. IRS proteins propagate insulin signals to the p85 regulatory subunit of phosphatidylinositol (PI) 3-kinase, which activates the p110 catalytic subunit. Insulin-induced PI 3-kinase activation is shown to be extremely important for the subsequent performance of a variety of insulin...

OBJECTIVEWe evaluated the clinical usefulness of a genetic risk score (GRS) based on 14 well-established variants for type 2 diabetes.RESEARCH DESIGN AND METHODSWe analyzed 14 SNPs at HHEX, CDKAL1, CDKN2B, SLC30A8, KCNJ11, IGF2BP2, PPARG, TCF7L2, FTO, KCNQ1, IRS-1, GCKR, UBE2E2, and C2CD4A/B in 1,487 Japanese individuals (724 patients with type 2 diabetes and 763 control subjects). A GRS was calculated according to the number of risk alleles by counting all 14 SNPs (T-GRS) as well as 11 SNPs related to β-cell function (β-GRS) and then assessing the association between each GRS and the clinical features.RESULTSAmong the 14 SNPs, 4 SNPs were significantly associated with type 2 diabetes in the present Japanese sample (P < 0.0036). The T-GRS was significantly associated with type 2 diabetes (P = 5.9 × 10−21). Among the subjects with type 2 diabetes, the β-GRS was associated with individuals receiving insulin therapy (β = 0.0131, SE = 0.006, P = 0.0431), age at diagnosis (β = −0.608, SE = 0.204, P = 0.0029), fasting serum C-peptide level (β = −0.032, SE = 0.0140, P = 0.022), and C-peptide index (β = −0.031, SE = 0.012, P = 0.0125).CONCLUSIONSOur data suggest that the β-GRS is associated with reduced β-cell functions and may be useful for selecting patients who should receive more aggressive β-cell–preserving therapy.

SH-2؊containing inositol 5-phosphatase 2 (SHIP-2) is a physiologically important lipid phosphatase that functions to hydrolyze phosphatidylinositol (PI) 3-kinase product PI(3,4,5)P3 to PI(3,4)P2 in the negative regulation of insulin signaling. We investigated whether SHIP-2 is associated with the insulin resistance of diabetic db/db mice. The amount of SHIP-2 protein was elevated in quadriceps muscle and epididymal fat tissue, but not in the liver, of db/db mice relative to that in control db/؉m mice. In accordance with the enhanced expression of SHIP-2, its localization at the membrane preparation was increased in the skeletal muscle and fat tissue of db/db mice. Insulin stimulation of PI 3-kinase activity was modestly decreased in skeletal muscle, fat tissue, and liver of db/db mice compared with that of db/؉m mice. In addition to the modest decrease at the level of PI 3-kinase, the activity of Akt and protein kinase C (PKC)-/, which are downstream molecules of PI 3-kinase, was more severely reduced in the skeletal muscle and fat tissue, but not in liver of db/db mice. Treatment with the insulin-sensitizing agent rosiglitazone decreased the elevated expression of SHIP-2 in the skeletal muscle and fat tissue of db/db mice. Insulininduced Akt activation and PKC-/ phosphorylation were restored to the control level, although insulinstimulated PI 3-kinase activation was minimally affected in the skeletal muscle and fat tissue of db/db mice. These results indicate that SHIP-2 is a novel molecule associated with insulin resistance in the skeletal muscle and fat tissue, and that insulin-induced activity of the downstream molecules of PI 3-kinase is decreased, at least in part, by the elevated expression of SHIP-2 in diabetic db/db mice. Diabetes 51:2387-2394, 2002

SH2-containing inositol phosphatase 2 (SHIP2) is a physiologically important negative regulator of insulin signaling by hydrolyzing the phosphatidylinositol (PI)3-kinase product PI 3,4,5-trisphosphate in the target tissues of insulin. Targeted disruption of the SHIP2 gene in mice resulted in increased insulin sensitivity without affecting biological systems other than insulin signaling. Therefore, we investigated the molecular mechanisms by which SHIP2 specifically regulates insulin-induced metabolic signaling in 3T3-L1 adipocytes. Insulin-induced phosphorylation of Akt, one of the molecules downstream of PI3-kinase, was inhibited by expression of wild-type SHIP2, whereas it was increased by expression of 5-phosphatase-defective (⌬IP) SHIP2 in whole cell lysates. The regulatory effect of SHIP2 was mainly seen in the plasma membrane (PM) and low density microsomes but not in the cytosol. In this regard, following insulin stimulation, a proportion of Akt2, and not Akt1, appeared to redistribute from the cytosol to the PM. Thus, insulin-induced phosphorylation of Akt2 at the PM was predominantly regulated by SHIP2, whereas the phosphorylation of Akt1 was only minimally affected. Interestingly, insulin also elicited a subcellular redistribution of both wild-type and ⌬IP-SHIP2 from the cytosol to the PM. The degree of this redistribution was inhibited in part by pretreatment with PI3-kinase inhibitor. Although the expression of a constitutively active form of PI3-kinase myr-p110 also elicited a subcellular redistribution of SHIP2 to the PM, expression of SHIP2 appeared to affect the myr-p110-induced phosphorylation, and not the translocation, of Akt2. Furthermore, insulin-induced phosphorylation of Akt was effectively regulated by SHIP2 in embryonic fibroblasts derived from knockout mice lacking either insulin receptor substrate-1 or insulin receptor substrate-2. These results indicate that insulin specifically stimulates the redistribution of SHIP2 from the cytosol to the PM independent of 5-phosphatase activity, thereby regulating the insulin-induced translocation and phosphorylation of Akt2 at the PM. Phosphatidylinositol (PI)1 3-kinase plays a central role in the metabolic actions of insulin. PI(3,4,5)P 3 produced by activated PI3-kinase is thought to function as a key lipid second messenger for signaling to further downstream molecules including Akt and atypical PKC (1-4). We and others (5, 6) have recently cloned SH2-containing inositol phosphatase 2 (SHIP2), which has 5Ј-phosphatase activity toward the PI3-kinase product, PI(3,4,5)P 3 , in the target tissues of insulin. Overexpression of SHIP2 inhibited insulin-induced metabolic signaling leading to glucose uptake and glycogen synthesis via 5Ј-phosphatase activity hydrolyzing the PI3-kinase product PI(3,4,5)P 3 to phosphatidylinositol 3,4-diphosphate in 3T3-L1 adipocytes and L6 myotubes (7,8). Importantly, targeted disruption of the SHIP2 gene in mice increased insulin sensitivity without affecting other biological systems (9). These reports indicate that SHIP...

Src homology 2-containing 5'-inositol phosphatase 2 (SHIP2) is known to be one of lipid phosphatases converting PI(3,4,5)P3 to PI(3,4)P2 in the negative regulation of insulin signaling with the fundamental impact on the state of insulin resistance. To clarify the possible involvement of SHIP2 in the pathogenesis of human type 2 diabetes, we examined the relation of human SHIP2 gene polymorphisms to type 2 diabetes in a Japanese population. We identified 10 polymorphisms including four missense mutations. Among them, single nucleotide polymorphism (SNP)3 (L632I) was located in the 5'-phosphatase catalytic region, and SNP5 (N982S) was adjacent to the phosphotyrosine binding domain binding consensus motif in the C terminus. SNP3 was found more frequently in control subjects than in type 2 diabetic patients, suggesting that this mutation might protect from insulin resistance. Transfection study showed that expression of SNP3-SHIP2 inhibited insulin-induced PI(3,4,5)P3 production and Akt2 phosphorylation less potently than expression of wild-type SHIP2 in CHO-IR cells. Insulin-induced tyrosine phosphorylation of SNP5-SHIP2 was decreased compared with that of wild-type SHIP2, resulting in increased Shc/Grb2 association and MAPK activation. These results indicate that the polymorphisms of SHIP2 are implicated, at least in part, in type 2 diabetes, possibly by affecting the metabolic and/or mitogenic insulin signaling in the Japanese population.