Deficiency in the extracellular signal-regulated kinase 1 (ERK1) protects leptin-deficient mice from insulin resistance without affecting obesity.

AIMS/HYPOTHESIS: Extracellular signal-regulated kinase (ERK) activity is increased in adipose tissue in obesity and type 2 diabetes mellitus and strong evidences suggests that it is implicated in the downregulation of insulin signalling and action in the insulin-resistant state. To determine the role of ERK1 in obesity-associated insulin resistance in vivo, we inactivated Erk1 (also known as Mapk3) in obese leptin-deficient mice (ob/ob). METHODS: Mice of genotype ob/ob-Erk1⁻(/)⁻ were obtained by crossing Erk1⁻(/)⁻ mice with ob/ob mice. Glucose tolerance and insulin sensitivity were studied in 12-week-old mice. Tissue-specific insulin sensitivity, insulin signalling, liver steatosis and adipose tissue inflammation were determined. RESULTS: While ob/ob-Erk1⁻(/)⁻ and ob/ob mice exhibited comparable body weight and adiposity, ob/ob-Erk1⁻(/)⁻ mice did not develop hyperglycaemia and their glucose tolerance was improved. Hyperinsulinaemic-euglycaemic clamp studies demonstrated an increase in whole-body insulin sensitivity in the ob/ob-Erk1⁻(/)⁻ mice associated with an increase in both insulin-stimulated glucose disposal in skeletal muscles and adipose tissue insulin sensitivity. This occurred in parallel with improved insulin signalling in both tissues. The ob/ob-Erk1⁻(/)⁻ mice were also partially protected against hepatic steatosis with a strong reduction in acetyl-CoA carboxylase level. These metabolic improvements were associated with reduced expression of mRNA encoding inflammatory cytokine and T lymphocyte markers in the adipose tissue. CONCLUSIONS/INTERPRETATION: Our results demonstrate that the targeting of ERK1 could partially protect obese mice against insulin resistance and liver steatosis by decreasing adipose tissue inflammation and by increasing muscle glucose uptake. Our results indicate that deregulation of the ERK1 pathway could be an important component in obesity-associated metabolic disorders.

The inflammatory receptor CD40 is expressed on human adipocytes: contribution to crosstalk between lymphocytes and adipocytes.

AIMS/HYPOTHESIS: Obesity is associated with adipose tissue inflammation. The CD40 molecule, TNF receptor superfamily member 5 (CD40)/CD40 ligand (CD40L) pathway plays a role in the onset and maintenance of the inflammatory reaction, but has not been studied in human adipose tissue. Our aim was to examine CD40 expression by human adipocytes and its participation in adipose tissue inflammation. METHODS: CD40 expression was investigated in human whole adipose tissue and during adipocyte differentiation by real-time PCR, Western blot and immunohistochemistry. The CD40/CD40L pathway was studied using recombinant CD40L (rCD40L) in adipocyte culture and neutralising antibodies in lymphocyte/adipocyte co-culture. RESULTS: CD40 mRNA levels in subcutaneous adipose tissue were higher in the adipocyte than in the stromal-vascular fraction. CD40 expression was upregulated during adipocyte differentiation. Addition of rCD40L to adipocytes induced mitogen activated protein kinase (MAPK) activation, stimulated inflammatory adipocytokine production, and decreased insulin-induced glucose transport in parallel with a downregulation of IRS1 and GLUT4 (also known as SCL2A4). rCD40L decreased the expression of lipogenic genes and increased lipolysis. CD40 mRNA levels were significantly higher in subcutaneous adipose tissue than in visceral adipose tissue of obese patients and were positively correlated with BMI, and with IL6 and leptin mRNA levels. Lymphocyte/adipocyte co-culture led to an upregulation of proinflammatory adipocytokines and a downregulation of leptin and adiponectin. Physical separation of the two cell types attenuated these effects, suggesting the involvement of a cell-cell contact. Blocking the CD40/CD40L interaction with neutralising antibodies reduced IL-6 secretion from adipocytes. CONCLUSIONS/INTERPRETATION: Adipocyte CD40 may contribute to obesity-related inflammation and insulin resistance. T lymphocytes regulate adipocytokine production through both the release of soluble factor(s) and heterotypic contact with adipocytes involving CD40.

Involvement of TNF-alpha in abnormal adipocyte and muscle sortilin expression in obese mice and humans.

AIMS/HYPOTHESIS: Insulin resistance is caused by numerous factors including inflammation. It is characterised by defective insulin stimulation of adipocyte and muscle glucose transport, which requires the glucose transporter GLUT4 translocation towards the plasma membrane. Defects in insulin signalling can cause insulin resistance, but alterations in GLUT4 trafficking could also play a role. Our goal was to determine whether proteins controlling GLUT4 trafficking are altered in insulin resistance linked to obesity. METHODS: Using real-time RT-PCR, we searched for selected transcripts that were differentially expressed in adipose tissue and muscle in obese mice and humans. Using various adipocyte culture models and in vivo mice treatment, we searched for the involvement of TNF-alpha in these alterations in obesity. RESULTS: Sortilin mRNA and protein were downregulated in adipose tissue from obese db/db and ob/ob mice, and also in muscle. Importantly, sortilin mRNA was also decreased in morbidly obese human diabetic patients. Sortilin and TNF-alpha (also known as TNF) mRNA levels were inversely correlated in mice and human adipose tissues. TNF-alpha decreased sortilin mRNA and protein levels in cultured mouse and human adipocytes, an effect partly prevented by the peroxisome proliferator-activated receptor gamma activator rosiglitazone. TNF-alpha also inhibited adipocyte and muscle sortilin mRNA when injected to mice. CONCLUSIONS/INTERPRETATION: Sortilin, an essential player in adipocyte and muscle glucose metabolism through the control of GLUT4 localisation, is downregulated in obesity and TNF-alpha is likely to be involved in this defect. Chronic low-grade inflammation in obesity could thus contribute to insulin resistance by modulating proteins that control GLUT4 trafficking.

p38MAP Kinase activity is required for human primary adipocyte differentiation.

Little is known about the role of p38MAPK in human adipocyte differentiation. Here we showed that p38MAPK activity increases during human preadipocytes differentiation. Pharmacological inhibition of p38MAPK during adipocyte differentiation of primary human preadipocytes markedly reduced triglycerides accumulation and adipocyte markers expression. Cell cycle arrest or proliferation was not affected by p38MAPK inhibition. Although induction of C/EBPbeta was not altered by the p38MAPK inhibitor, its phosphorylation on Threonine(188) was decreased as well as PPARgamma expression. These results indicate that p38MAPK plays a positive role in human adipogenesis through regulation of C/EBPbeta and PPARgamma factors.

Defect in skeletal muscle phosphatidylinositol-3-kinase in obese insulin-resistant mice.

Activation of phosphatidylinositol-3-kinase (PI3K) is one of the earliest postreceptor events in the insulin signaling pathway. Incubation of soleus muscles from lean mice with 50 nM insulin caused a 3-10-fold increase in antiphosphotyrosine-immunoprecipitable PI3K (antiPTyr-PI3K) activity within 2 min in muscle homogenates as well as both the cytosolic and membrane fractions. Insulin did not affect total PI3K activity. Both the antiPTyr-PI3K stimulation and activation of insulin receptor tyrosine kinase were dependent on hormone concentration. In muscles from obese, insulin-resistant mice, there was a 40-60% decrease in antiPTyr-PI3K activity after 2 min of insulin that was present equally in the cytosolic and membrane fractions. A significant reduction in insulin sensitivity was also observed. The defect appears to result from alterations in both insulin receptor and postreceptor signaling. Starvation of obese mice for 48 h, which is known to reverse insulin resistance, normalized the insulin response of both PI3K and the receptor tyrosine kinase. The results demonstrate that: (a) antiPTyr-PI3K activity is responsive to insulin in mouse skeletal muscle, (b) both the insulin responsiveness and sensitivity of this activity are blunted in insulin-resistant muscles from obese mice, (c) these alterations result from a combination of insulin receptor and postreceptor defects, and (d) starvation restores normal insulin responses.

Energy disruptors: rising stars in anticancer therapy?

The metabolic features of tumor cells diverge from those of normal cells. Otto Warburg was the first to observe that cancer cells dramatically increase their glucose consumption to generate ATP. He also claimed that cancer cells do not have functional mitochondria or oxidative phosphorylation (OXPHOS) but simply rely on glycolysis to provide ATP to the cell, even in the presence of oxygen (aerobic glycolysis). Several studies have revisited this observation and demonstrated that most cancer cells contain metabolically efficient mitochondria. Indeed, to sustain high proliferation rates, cancer cells require functional mitochondria to provide ATP and intermediate metabolites, such as citrate and cofactors, for anabolic reactions. This difference in metabolism between normal and tumors cells causes the latter to be more sensitive to agents that can disrupt energy homeostasis. In this review, we focus on energy disruptors, such as biguanides, 2-deoxyglucose and 5-aminoimidazole-4-carboxamide ribonucleotide, that interfere with the main metabolic pathways of the cells, OXPHOS, glycolysis and glutamine metabolism. We discuss the preclinical data and the mechanisms of action of these disruptors at the cellular and molecular levels. Finally, we consider whether these drugs can reasonably contribute to the antitumoral therapeutic arsenal in the future.

Inhibition of the MAP3 kinase Tpl2 protects rodent and human ß-cells from apoptosis and dysfunction induced by cytokines and enhances anti-inflammatory actions of exendin-4.

Proinflammatory cytokines exert cytotoxic effects on ß-cells, and are involved in the pathogenesis of type I and type II diabetes and in the drastic loss of ß-cells following islet transplantation. Cytokines induce apoptosis and alter the function of differentiated ß-cells. Although the MAP3 kinase tumor progression locus 2 (Tpl2) is known to integrate signals from inflammatory stimuli in macrophages, fibroblasts and adipocytes, its role in ß-cells is unknown. We demonstrate that Tpl2 is expressed in INS-1E ß-cells, mouse and human islets, is activated and upregulated by cytokines and mediates ERK1/2, JNK and p38 activation. Tpl2 inhibition protects ß-cells, mouse and human islets from cytokine-induced apoptosis and preserves glucose-induced insulin secretion in mouse and human islets exposed to cytokines. Moreover, Tpl2 inhibition does not affect survival or positive effects of glucose (i.e., ERK1/2 phosphorylation and basal insulin secretion). The protection against cytokine-induced ß-cell apoptosis is strengthened when Tpl2 inhibition is combined with the glucagon-like peptide-1 (GLP-1) analog exendin-4 in INS-1E cells. Furthermore, when combined with exendin-4, Tpl2 inhibition prevents cytokine-induced death and dysfunction of human islets. This study proposes that Tpl2 inhibitors, used either alone or combined with a GLP-1 analog, represent potential novel and effective therapeutic strategies to protect diabetic ß-cells.

Brown adipose tissue in lean and obese mice. Insulin-receptor binding and tyrosine kinase activity.

Insulin-receptor binding and tyrosine kinase activity have been studied in brown adipose tissue from lean and obese mice. Brown adipose tissue carries functional insulin receptors comparable with those of conventional insulin target tissues. The alpha-subunit (Mr, 130,000) was labeled with photoreactive insulin; the beta-subunit (Mr, 95,000) was phosphorylated in a cell-free system, and its level of phosphorylation was increased in a dose-dependent manner by insulin. Two types of obese mice, mice rendered obese by gold thioglucose injection (GTG obese) and genetically obese ob/ob mice, were used. Insulin-receptor number was decreased by 60-70% in obese mice, when expressed per milligram of plasma membrane protein or per microgram of glycoprotein, whereas only a 30-40% diminution was observed in skeletal muscle, indicating that insulin receptors from brown adipose tissue are greatly affected by the downregulation process. Insulin-stimulated autophosphorylation of the insulin-receptor beta-subunit was decreased by 60-70% in preparations of obese mice compared with lean mice in direct proportion to the diminished level of insulin-receptor number. Similarly, the ability of receptors to catalyze the phosphorylation of a synthetic substrate (copolymer glutamate-tyrosine) was reduced. These results suggest that the decrease in insulin-receptor number and in associated tyrosine kinase activity could explain the insulin-resistant glucose uptake and the alteration in diet-induced thermogenesis described in obese animals.

Isolation and characterization of a T lymphocyte mutant defective in the protein kinase C signal transduction pathway.

The phorbol ester TPA is a potent protein kinase C (PKC) activator and a cofactor in the activation of the human Jurkat leukemic T cell line. We have studied the implication of the PKC signaling pathway in the process of T cell activation by generating TPA resistant mutants of Jurkat. These mutants were obtained by recovery of cells that survived a growth arrest induced by TPA. Several cellular phenomena dependent on TPA were dramatically altered in the mutated cells. The mutants were unable to form homoaggregates upon TPA stimulation. Moreover, they did not produce interleukin-2 after activation through engagement of the T cell receptor, in the presence of TPA. These results suggest that the PKC signaling pathway activated by TPA is defective in these cells. In an attempt to define and locate the defect present in the mutants, we have analysed the biochemical properties of PKC, the cellular receptor of TPA. The increase in kinase activity and the translocation of the enzyme to the plasma membrane after stimulation by TPA appeared to be normal in the mutants. We hypothesize that a metabolic step, critical for the completion of T cell activation, distinct from protein kinase C, is impaired in the mutant cells.

Potential involvement of the carboxy-terminus of the Glut 1 transporter in glucose transport.

The role of the carboxy-terminal domain of the Glut 1 glucose transporter was investigated using an antipeptide antibody to the C-terminal part of the molecule. The study was performed in fibroblasts transfected with the cDNA coding for the human insulin receptor. These cells acutely respond to insulin for glucose transport. Using antipeptide antibodies to Glut 1 and Glut 4, we first established that these cells expressed only Glut 1. Then, to define the role of the C-terminal part of Glut 1 in glucose transport, the antibodies were loaded into the cells by electroporation. When anti-Glut 1 immunoglobulins were introduced into the cells, a 60% increase in basal deoxyglucose and 3-O-methylglucose transport was observed compared to that in cells electroporated with nonimmune immunoglobulins. The stimulatory action of the antipeptide was not due to an increase in the total amount of transporters. It was found only at low glucose concentrations, suggesting that the affinity of the transporter, rather than its maximal capacity, was changed. Finally, the effect of antibody was additive to that of insulin. The interaction between the anti-Glut 1 antibody and the carboxy-tail of the transporter seems to lead to an increase in the intrinsic activity of the transporter, suggesting that this part of the molecule could be implicated in the regulation of glucose uptake.

Subcellular distribution of low molecular weight guanosine triphosphate-binding proteins in adipocytes: colocalization with the glucose transporter Glut 4.

Insulin stimulation of glucose transport involves the translocation of vesicles containing the glucose transporter Glut 4 to the plasma membrane. Since low mol wt GTP-binding proteins (LMW-GTP-binding proteins) have been implicated in the regulation of vesicular trafficking, we have analyzed these proteins in adipocytes. Isolated adipocytes were incubated in the absence or presence of insulin before separation of plasma membranes (PM) and low density microsomes (LDM). [alpha-32P]GTP binding to proteins transferred to nitrocellulose after sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed specific and distinct subsets of proteins in the PM and LDM; those proteins were more abundant in PM than in LDM. [alpha-32P]GTP binding to these proteins was specific for the guanylnucleotides, since it was competed for by GTP and guanosine 5'-O-(3-thiotriphosphate), but not by ATP or adenosine 5'-O-(3-thiotriphosphate). The LMW-GTP-binding proteins were tightly associated with the membranes, as treatment with 1.5 M KCl did not modify this association. The distribution of the LMW-GTP-binding proteins in the fractions and their affinity for guanylnucleotides were the same in control and insulin-treated adipocytes. When the presence of Gi alpha subunits was looked for with a specific antibody, Gi alpha 1 and Gi alpha 2 were found almost exclusively in PM. By contrast, the same antibody revealed the presence of a 100 kDa band in the LDM. Insulin treatment of adipocytes did not modify the amounts of those G-proteins in PM or LDM fractions, although it promoted the translocation of Glut 4 proteins from LDM to PM. LDM fractions contain a specific subset of vesicles markedly enriched in Glut 4 molecules. When those vesicles were isolated from the total LDM fraction by immunoadsorption on highly specific antibodies to Glut 4 protein, LMW-GTP-binding proteins were found in the immune pellet. Those proteins were absent when immunoprecipitation was performed after solubilization of the vesicles with 1% Triton X-100. Our results strongly suggest that the vesicles containing the Glut 4 protein also contained LMW-GTP-binding proteins and indicate that these GTP-binding proteins could play a role in the exocytosis of the Glut 4-containing vesicles.

Potential role of protein kinase B in glucose transporter 4 translocation in adipocytes.

Phosphatidylinositol 3-kinase (PI 3-kinase) activation promotes glucose transporter 4 (Glut 4) translocation in adipocytes. In this study, we demonstrate that protein kinase B, a serine/threonine kinase stimulated by PI 3-kinase, is activated by both insulin and okadaic acid in isolated adipocytes, in parallel with their effects on Glut 4 translocation. In 3T3-L1 adipocytes, platelet-derived growth factor activated PI 3-kinase as efficiently as insulin but was only half as potent as insulin in promoting protein kinase B (PKB) activation. To look for a potential role of PKB in Glut 4 translocation, adipocytes were transfected with a constitutively active PKB (Gag-PKB) together with an epitope tagged transporter (Glut 4 myc). Gag-PKB was associated with all membrane fractions, whereas the endogenous PKB was mostly cytosolic. Expression of Gag-PKB led to an increase in Glut 4 myc amount at the cell surface. Our results suggest that PKB could play a role in promoting Glut 4 appearance at the cell surface following exposure of adipocytes to insulin and okadaic acid stimulation.

Glucose transporter in insulin sensitive tissues of lean and obese mice. Effect of the thermogenic agent BRL 26830A.

Glucose transport is decreased in skeletal muscle and adipose tissues of obese, hyperglycemic, insulin-resistant animals. Here we have characterized the glucose transporter(s) in muscle and adipose tissues from normal and obese mice, and we have studied the effect of a treatment with the thermogenic agent BRL 26830A. Glucose transporters were examined in crude tissue membrane fractions (microsomal + plasma membranes) by Western blot analysis using antipeptide antibodies specific for the erythroid (Glut 1) or muscle/fat (Glut 4) glucose transporters. In these insulin sensitive tissues, only Glut 4 was detected. In membranes from obese animals, the Glut 4 number was decreased by 40% +/- 4% in brown adipose tissue (mean +/- SEM of 9 preparations, P less than 0.001), whether the results were expressed per total tissue or per mg of protein. By contrast, Glut 4 number was unchanged in skeletal muscle. In white adipose tissue of obese animals, Glut 4 number per total fat pad was increased. However, due to the enlarged fat pad size, Glut 4 content was diminished when expressed per mg of white adipose tissue membrane protein in obese compared to lean animals. After a 18 day-treatment with BRL 26830A (1 or 2 mg/kg.day), glycemia of obese mice, which was slightly elevated compared to lean animals, was normalized, while insulinemia remained markedly above control values. In brown adipose tissue, the total number of Glut 4 returned to normal at 1 mg of the drug, or increased by 63% +/- 14% at 2 mg. Since membrane protein content was increased by the treatment, when results were expressed per mg of membrane protein, Glut 4 was similar in lean and BRL 26830A (1 or 2 mg) treated obese mice. BRL 26830A treatment did not modify Glut 4 in skeletal muscle, and it increased Glut 4 number in white adipose tissue in a dose-dependent manner. In conclusion, in obese mice, the glucose transporter number was reduced mainly in brown adipose tissue, a defect which could contribute to the hyperglycemic syndrome. Treatment with the thermogenic agent BRL 26830A normalized in parallel glycemia and glucose transporter number in brown adipose tissue, suggesting that this tissue could play a role in glucose homeostasis in rodents.

Parallel changes in Glut 4 and Rab4 movements in two insulin-resistant states.

Insulin-induced Glut 4 and Rab4 movements were studied in two insulin-resistant states. In adipocytes from streptozotocin diabetic rats, the amount of Glut 4 was decreased by 60%. The remaining Glut 4 molecules were translocated in response to insulin, and in parallel, Rab4 left the intracellular compartment. In contrast, in 3T3-L1 adipocytes rendered insulin-resistant by a prolonged insulin treatment, both Rab4 and Glut 4 remained in the intracellular compartment following an acute insulin stimulation. Those results illustrate a similar behavior of Glut 4 and Rab4 in two situations where insulin resistance results from different mechanisms, and add further support for a role of Rab4 in Glut 4 translocation.

Alteration of insulin receptor kinase in obese, insulin-resistant mice.

We have studied the properties of muscle insulin receptors obtained from genetically or experimentally-induced obese mice that are both insulin-resistant. Insulin receptors, partially purified by wheat germ agglutinin--agarose chromatography, were studied in a cell-free system for autophosphorylation, for their ability to phosphorylate a synthetic glutamate--tyrosine copolymer and for their binding characteristics. Insulin receptor number was decreased by 25% in muscles from obese mice without any change in their binding affinity. The insulin stimulatory action on its beta-subunit receptor phosphorylation was diminished in preparations from genetically- or experimentally-induced obese mice to a higher degree than the decrease in insulin receptor number. HPLC analysis of the phosphopeptides generated by trypsin treatment of the labeled receptor beta-subunit was identical in lean and obese mice. Similar alteration of the kinase activity was found in obese mice when the phosphorylation of casein or polyglutamate--tyrosine was measured. Trypsin treatment of the receptor preparations was less effective in stimulating the kinase activity in obese mice than in lean mice. These results suggest that the defect in insulin receptor kinase activity reflects an alteration in the transmission of the message from the alpha- to the beta-subunit or an impairment of the enzyme functioning by environmental conditions.

Insulin-like growth factor-I-stimulated glucose transport in myotubes derived from chicken muscle satellite cells.

The effects of insulin and insulin-like growth factor-I (IGF-I) on glucose transport were compared in myotubes derived from chicken breast muscle satellite cells in vitro. Myotubes were incubated (for 0.5 or 4 h) with or without glucose in the presence or absence of insulin or IGF-I. Glucose uptake was subsequently measured by the incorporation of 2-[1,2-3H(N)] deoxy-D-glucose ([3H]2DG) in glucose-free medium (10 min at 20 degrees C). Glucose uptake was almost completely abolished by the addition of cytochalasin B or phloretin. It was increased by a decrease in glucose concentration in the incubation medium. Insulin (5 mg/l) stimulated [3H]2DG uptake to a maximum of 43 +/- 10% above basal after 30-min incubation and 101 +/- 15% after 4-h incubation. IGF-I and insulin at equimolar concentrations (25 micrograms/l and 20 micrograms/l respectively) were almost equipotent after 0.5 h but after 4-h incubation IGF-I was 17-fold more potent, suggesting that this 'late' effect was mediated through the IGF-I receptor. Incubation with cycloheximide suggested that the effect of IGF-I involved increased protein synthesis. The results suggest that chicken myotubes express a glucose transporter which is regulated by IGF-I and glucose concentration. However, they do not appear to express a typical insulin-responsive transport system.

Expression of guanine-nucleotide-binding proteins in lean and obese insulin-resistant mice.

To examine whether G protein were affected in the obese insulin-resistant state, the level of various G proteins (alpha i1, alpha i2, alpha i3, alpha o and alpha s) was assessed by immunodetection in lean and experimentally induced obese mice. Crude membranes were prepared from adipose tissues, muscle, liver, kidney and brain. G alpha-subunits were similar in lean and obese animals in brain, kidney, skeletal or heart muscle. Hepatic G alpha s, G alpha i2 and G alpha i3 subunits were markedly elevated in obese mice. When total tissue contents were considered, interscapular brown adipose tissue and epididymal fat pads from obese animals contained more alpha i2 than the lean tissues, while alpha i1, alpha i3 and alpha s were similar in both groups. However, when expressed per mg of membrane protein, alpha i1, alpha i3 and alpha s were decreased and alpha i2 was normal in white adipose tissue of obese animals. Thus the expression of the G protein alpha-subunits seems to be regulated by tissue-specific factors rather than by circulating factors.

Characterization of 6-deoxy-6-iodo-D-glucose: a potential new tool to assess glucose transport.

6-deoxy-6-iodo-D-glucose (6-DIG) was rapidly taken up by adipocytes. Insulin increased 6-DIG transport in adipocytes isolated from both rats and mice. This stimulation was more important in rat than in mouse adipocytes, in agreement with their respective amount of Glut 4 transporters. In two insulin resistant states, the biological behavior of 6-DIG and 3-O-methyl-D-glucose was similar. These results indicated that 6-DIG, which was transported into the cells via the glucose transporters, could be potentially useful to measure modifications of glucose transport.

Alteration in insulin action: role of IRS-1 serine phosphorylation in the retroregulation of insulin signalling.

Insulin resistance, when combined with impaired insulin secretion, contributes to the development of type 2 diabetes. Insulin resistance is characterised by a decrease in insulin effect on glucose transport in muscle and adipose tIssue. Tyrosine phosphorylation of insulin receptor substrate 1 (IRS-1) and its binding to phosphatidylinositol 3-kinase (PI 3-kinase) are critical events in the insulin signalling cascade leading to insulin-stimulated glucose transport. Modification of IRS-1 by serine phosphorylation could be one of the mechanisms leading to a decrease in IRS-1 tyrosine phosphorylation, PI 3-kinase activity and glucose transport. Recent findings demonstrate that "diabetogenic" factors such as FFA, TNFalpha, hyperinsulinemia and cellular stress, increase the serine phosphorylation of IRS-1 and identified Ser307/612/632 as phosphorylated sites. Moreover, several kinases able to phosphorylate these serine residues have been identified. These exciting results suggest that serine phosphorylation of IRS-1 is a possible hallmark of insulin resistance in biologically insulin responsive cells or tIssues. Identifying the pathways by which "diabetogenic" factors activate IRS-1 kinases and defining the precise role of serine phosphorylation events in IRS-1 regulation represent important goals. Such studies may enable rational drug design to selectively inhibit the activity of the relevant enzymes and generate a novel class of therapeutic agents for type 2 diabetes.

Sestrin2 integrates Akt and mTOR signaling to protect cells against energetic stress-induced death.

The phosphoinositide-3 kinase/Akt (PI3K/Akt) pathway has a central role in cancer cell metabolism and proliferation. More importantly, it is one of the cardinal pro-survival pathways mediating resistance to apoptosis. The role of Akt in response to an energetic stress is presently unclear. Here, we show that Sestrin2 (Sesn2), also known as Hi95, a p53 target gene that protects cells against oxidative and genotoxic stresses, participates in the protective role of Akt in response to an energetic stress induced by 2-deoxyglucose (2-DG). Sesn2 is upregulated in response to an energetic stress such as 2-DG and metformin, and mediates the inhibition of mammalian target of rapamycin (mTOR), the major cellular regulator of energy metabolism. The increase of Sesn2 is independent of p53 but requires the anti-apoptotic pathway, PI3K/Akt. Inhibition of Akt, as well as loss of Sesn2, sensitizes cells to 2-DG-induced apoptosis. In addition, the rescue of Sesn2 partially reverses the pro-apoptotic effects of 2-DG. In conclusion, we identify Sesn2 as a new energetic stress sensor, which appears to be protective against energetic stress-induced apoptosis that integrates the pro-survival function of Akt and the negative regulation of mTOR.