Findings suggest that the T allele at nucleotide 29 in the TGF-beta1 gene is a risk factor for genetic susceptibility to MI, at least in middle-aged Japanese men.

Adipose differentiation-related protein (ADRP) is a lipid droplet-associated protein that is expressed early during adipose differentiation. The present study was undertaken to reveal the role of ADRP in adipose differentiation. In murine fibroblasts infected with green fluorescent protein (GFP)-ADRP fusion protein expression adenovirus vector, confocal microscopic analysis showed the number and size of lipid droplets apparently increased comparing with those of control cells. Overexpressed GFP-ADRP were mainly located at the surface of lipid droplets and appeared to be “ring-shaped.” Triacylglycerol content was also significantly ( P < 0.001) increased in GFP-ADRP-overexpressed cells compared with control cells. ADRP-induced lipid accumulation did not depend on adipocyte-specific gene induction, such as peroxisome proliferator-activated receptor-γ, lipoprotein lipase, or other lipogenic genes, including acyl-CoA synthetase, fatty acid-binding protein, and fatty acid transporter. In conclusion, ADRP stimulated lipid accumulation and lipid droplet formation without induction of other adipocyte-specific genes or other lipogenic genes in murine fibroblasts. The detailed molecular mechanisms of ADRP on lipid accumulation remain to be elucidated.

We evaluated the role of the G alpha-q (G␣q) subunit of heterotrimeric G proteins in the insulin signaling pathway leading to GLUT4 translocation. We inhibited endogenous G␣q function by single cell microinjection of anti-G␣q/11 antibody or RGS2 protein (a GAP protein for G␣q), followed by immunostaining to assess GLUT4 translocation in 3T3-L1 adipocytes. G␣q/11 antibody and RGS2 inhibited insulin-induced GLUT4 translocation by 60 or 75%, respectively, indicating that activated G␣q is important for insulin-induced glucose transport. We then assessed the effect of overexpressing wild-type G␣q (WT-G␣q) or a constitutively active G␣q mutant (Q209L-G␣q) by using an adenovirus expression vector. In the basal state, Q209L-G␣q expression stimulated 2-deoxy-D-glucose uptake and GLUT4 translocation to 70% of the maximal insulin effect. This effect of Q209L-G␣q was inhibited by wortmannin, suggesting that it is phosphatidylinositol 3-kinase (PI3-kinase) dependent. We further show that Q209L-G␣q stimulates PI3-kinase activity in p110␣ and p110␥ immunoprecipitates by 3-and 8-fold, respectively, whereas insulin stimulates this activity mostly in p110␣ by 10-fold. Nevertheless, only microinjection of anti-p110␣ (and not p110␥) antibody inhibited both insulin-and Q209L-G␣q-induced GLUT4 translocation, suggesting that the metabolic effects induced by Q209L-G␣q are dependent on the p110␣ subunit of PI3-kinase. In summary, (i) G␣q appears to play a necessary role in insulin-stimulated glucose transport, (ii) G␣q action in the insulin signaling pathway is upstream of and dependent upon PI3-kinase, and (iii) G␣q can transmit signals from the insulin receptor to the p110␣ subunit of PI3-kinase, which leads to GLUT4 translocation.

OBJECTIVETo identify novel type 2 diabetes gene variants and confirm previously identified ones, a three-staged genome-wide association study was performed in the Japanese population.RESEARCH DESIGN AND METHODSIn the stage 1 scan, we genotyped 519 case and 503 control subjects with 482,625 single nucleotide polymorphism (SNP) markers; in the stage 2 panel comprising 1,110 case subjects and 1,014 control subjects, we assessed 1,456 SNPs (P < 0.0025, stage 1); additionally to direct genotyping, 964 healthy control subjects formed the in silico control panel. Along with genome-wide exploration, we aimed to replicate the disease association of 17 SNPs from 16 candidate loci previously identified in Europeans. The associated and/or replicated loci (23 SNPs; P < 7 × 10–5 for genome-wide exploration and P < 0.05 for replication) were examined in the stage 3 panel comprising 4,000 case subjects and 12,569 population-based samples, from which 4,889 nondiabetic control subjects were preselected. The 12,569 subjects were used for overall risk assessment in the general population.RESULTSFour loci—1 novel with suggestive evidence (PEPD on 19q13, P = 1.4 × 10–5) and three previously reported—were identified; the association of CDKAL1, CDKN2A/CDKN2B, and KCNQ1 were confirmed (P < 10–19). Moreover, significant associations were replicated in five other candidate loci: TCF7L2, IGF2BP2, SLC30A8, HHEX, and KCNJ11. There was substantial overlap of type 2 diabetes susceptibility genes between the two populations, whereas effect size and explained variance tended to be higher in the Japanese population.CONCLUSIONSThe strength of association was more prominent in the Japanese population than in Europeans for more than half of the confirmed type 2 diabetes loci.

PTEN is a tumor suppressor with sequence homology to protein-tyrosine phosphatases and the cytoskeleton protein tensin. PTEN is capable of dephosphorylating phosphatidylinositol 3,4,5-trisphosphate in vitro and down-regulating its levels in insulin-stimulated 293 cells. To study the role of PTEN in insulin signaling, we overexpressed PTEN in 3T3-L1 adipocytes ϳ30-fold above uninfected or control virus (green fluorescent protein)-infected cells, using an adenovirus gene transfer system. PTEN overexpression inhibited insulin-induced 2-deoxy-glucose uptake by 36%, GLUT4 translocation by 35%, and membrane ruffling by 50%, all of which are phosphatidylinositol 3-kinase-dependent processes, compared with uninfected cells or cells infected with control virus. Microinjection of an anti-PTEN antibody increased basal and insulin stimulated GLUT4 translocation, suggesting that inhibition of endogenous PTEN function led to an increase in intracellular phosphatidylinositol 3,4,5-trisphosphate levels, which stimulates GLUT4 translocation. Further, insulin-induced phosphorylation of downstream targets Akt and p70S6 kinase were also inhibited significantly by overexpression of PTEN, whereas tyrosine phosphorylation of the insulin receptor and IRS-1 or the phosphorylation of mitogen-activated protein kinase were not affected, suggesting that the Ras/mitogen-activated protein kinase pathway remains fully functional. Thus, we conclude that PTEN may regulate phosphatidylinositol 3-kinasedependent insulin signaling pathways in 3T3-L1 adipocytes.Insulin binding to its receptor stimulates receptor autophosphorylation activating its intrinsic tyrosine kinase activity leading to phosphorylation of cellular substrates such as Shc, IRS-1, IRS-2, IRS-3, IRS-4, and other proteins (1-5). Tyrosine phosphorylation of the IRS and Shc proteins allows them to serve as docking proteins that interact with signaling molecules containing Src homology 2 domains (6), including PI 1 3-kinase. PI 3-kinase is a dual protein and lipid kinase composed of a heterodimer of a 110-kDa catalytic subunit (p110) and an 85-kDa regulatory subunit (p85) that contains two Src homology 2 domains (7). Binding of the p85 Src homology 2 domains to phosphotyrosines of the IRSs or directly to the carboxyl terminus of the IR, leads to activation of the associated p110 (8 -10), which preferentially phosphorylates the D-3 position of phosphatidylinositol (PtdIns), PtdIns 4-phosphate, and PtdIns 4,5-bisphosphate producing PtdIns 3-phosphate, PtdIns 3,4-bisphosphate, and PtdIns 3,4,5-P 3 , respectively. These PtdIns can serve as lipid second messengers that play a crucial role in the biologic actions of growth factors (11). However, the exact function of each of these PtdIns in hormone signaling is not fully defined.PI 3-kinase activation is necessary for a number of insulinstimulated effects, ranging from stimulation of glucose transport, glycogen synthesis, and membrane ruffling to mitogenesis, although the precise function of PtdIns products in eliciting these responses is cu...

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These findings indicate that the domain including the hydrophobic region of ATGL was essential for association with LDs.