The interaction of insulin and growth factors with their receptors on the outside surface of a cell, leads to the activation of phosphatidylinositol 3-kinase (PI 3-kinase) and generation of the phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P 3 ) second messenger at the inner surface of the cell membrane. One of the most studied signalling events controlled by PtdIns(3,4,5)P 3 , comprises the activation of a group of AGC family protein kinases, including isoforms of protein kinase B (PKB)/Akt, p70 ribosomal S6 kinase (S6K), serum-and glucocorticoid-induced protein kinase (SGK) and protein kinase C (PKC), which play crucial roles in regulating physiological processes relevant to metabolism, growth, proliferation and survival. Here, we review recent biochemical, genetic and structural studies on the 3-phosphoinositide-dependent protein kinase-1 (PDK1), which phosphorylates and activates the AGC kinase members regulated by PI 3-kinase. We also discuss whether inhibitors of PDK1 might have chemotherapeutic potential in the treatment of cancers in which the PDK1-regulated AGC kinases are constitutively activated.

A high-fat diet causes activation of the regulatory protein cJun NH 2 -terminal kinase 1 (JNK1) and triggers the development of insulin resistance.

PDK1 functions as a master kinase, phosphorylating and activating PKB/Akt, S6K and RSK. To learn more about the roles of PDK1, we generated mice that either lack PDK1 or possess PDK1 hypomorphic alleles, expressing only approximately 10% of the normal level of PDK1. PDK1(-/-) embryos die at embryonic day 9.5, displaying multiple abnormalities including lack of somites, forebrain and neural crest derived tissues; however, development of hind- and midbrain proceed relatively normally. In contrast, hypomorphic PDK1 mice are viable and fertile, and insulin injection induces the normal activation of PKB, S6K and RSK. Nevertheless, these mice are 40-50% smaller than control animals. The organ volumes from the PDK1 hypomorphic mice are reduced proportionately. We also establish that the volume of a number of PDK1-deficient cells is reduced by 35-60%, and show that PDK1 deficiency does not affect cell number, nuclear size or proliferation. We provide genetic evidence that PDK1 is essential for mouse embryonic development, and regulates cell size independently of cell number or proliferation, as well as insulin's ability to activate PKB, S6K and RSK.

Human arginase I is a potential target for therapeutic intervention in diseases linked to compromised L-arginine homeostasis. Here, we report high-affinity binding of the reaction coordinate analogue inhibitors 2(S)-amino-6-boronohexanoic acid (ABH, K d ‫؍‬ 5 nM) and S-(2-boronoethyl)-L-cysteine (BEC, Kd ‫؍‬ 270 nM) to human arginase I, and we report x-ray crystal structures of the respective enzyme-inhibitor complexes at 1.29-and 1.94-Å resolution determined from crystals twinned by hemihedry. The ultrahighresolution structure of the human arginase I-ABH complex yields an unprecedented view of the binuclear manganese cluster and illuminates the structural basis for nanomolar affinity: bidentate inner-sphere boronate-manganese coordination interactions and fully saturated hydrogen bond networks with inhibitor ␣-amino and ␣-carboxylate groups. These interactions are therefore implicated in the stabilization of the transition state for L-arginine hydrolysis. Electron density maps also reveal that active-site residue H141 is protonated as the imidazolium cation. The location of H141 is such that it could function as a general acid to protonate the leaving amino group of L-ornithine during catalysis, and this is a revised mechanistic proposal for arginase. This work serves as a foundation for studying the structural and chemical biology of arginase I in the immune response, and we demonstrate the inhibition of arginase activity by ABH in human and murine myeloid cells.boronic acid ͉ metalloenzyme ͉ protein crystallography A rginase is a trimeric binuclear manganese metalloenzyme that catalyzes the hydrolysis of L-arginine to form L-ornithine and urea (1-3). Two isozymes have been identified in mammals: arginase I catalyzes the final cytosolic step of the urea cycle in liver, and arginase II is a mitochondrial enzyme that functions in Larginine homeostasis in nonhepatic tissues. Notably, arginase I is also expressed in certain nonhepatic tissues where it, too, can function in L-arginine homeostasis. For example, arginase I may regulate substrate L-arginine bioavailability to NO synthase in the immune response. Macrophage arginase I and NO synthase are reciprocally regulated at the level of transcription: NO synthase is induced by T-helper type 1 (TH1) cytokines, and arginase I is induced by T-helper type 2 (TH2) cytokines (4-7). As a modulator of NO-dependent macrophage cytotoxicity, arginase I is implicated in the regulation of macrophage activity in wound healing (8) and the suppression of the tumoricidal activity of macrophages (9) and T cells (10). Notably, arginase I is very highly up-regulated in the murine spinal cord during experimental autoimmune encephalomyelitis, an animal model for human multiple sclerosis (11), and it is up-regulated in the inflammatory regions of the asthmatic lung (12)(13)(14).Arginase I in the immune response is also implicated in cancer biology: arginase I is significantly up-regulated and promotes tumor cell growth in breast cancer (15, 16) and colorectal cancer (17). Rodriguez et a...

Non-alcoholic steatosis (fatty liver) is a major cause of liver dysfunction that is associated with insulin resistance and metabolic syndrome. The cJun NH2-terminal kinase 1 (JNK1) signaling pathway is implicated in the pathogenesis of hepatic steatosis and drugs that target JNK1 may be useful for treatment of this disease. Indeed, mice with defects in JNK1 expression in adipose tissue are protected against hepatic steatosis. Here we report that mice with specific ablation of Jnk1 in hepatocytes exhibit glucose intolerance, insulin resistance, and hepatic steatosis. JNK1 therefore serves opposing actions in liver and adipose tissue to both promote and prevent hepatic steatosis. This finding has profound implications for the design of JNK1-selective drugs for the treatment of metabolic syndrome.

The cJun N-terminal kinase 1 (JNK1) is implicated in diet-induced obesity. Indeed, germline ablation of the murine Jnk1 gene prevents diet-induced obesity. Here we demonstrate that selective deficiency of JNK1 in the murine nervous system is sufficient to suppress diet-induced obesity. The failure to increase body mass is mediated, in part, by increased energy expenditure that is associated with activation of the hypothalamicpituitary-thyroid axis. Disruption of thyroid hormone function prevents the effects of nervous system JNK1 deficiency on body mass. These data demonstrate that the hypothalamic-pituitary-thyroid axis represents an important target of metabolic signaling by JNK1.[Keywords: JNK1; obesity; insulin resistance; thyroid hormone] Supplemental material is available at http://www.genesdev.org. Human obesity represents a serious world-wide health problem. One consequence of obesity is the development of insulin resistance, hyperglycemia, and metabolic syndrome that can lead to b-cell dysfunction and type 2 diabetes (Kahn et al. 2006). It is therefore important that we gain an understanding of the physiology and pathophysiology of the development of obesity, because this knowledge represents a basis for the design of potential therapeutic interventions.The cJun N-terminal kinase 1 (JNK1) represents one signaling pathway that has been implicated in dietinduced obesity (Weston and Davis 2007). JNK1 is activated when mice are fed a high-fat diet (HFD) (Hirosumi et al. 2002). Moreover, Jnk1 À/À mice are protected against HFD-induced weight gain (Hirosumi et al. 2002). The mechanism that accounts for the effect of germline JNK1 deficiency to control body weight is unclear. Tissue-specific deficiency of JNK1 in fat, muscle, liver, and myeloid cells does not affect HFD-induced weight gain (Sabio et al. 2008(Sabio et al. , 2009(Sabio et al. , 2010. A different organ must therefore play a major role in the diet-induced regulation of body weight by JNK1. The brain represents a possible site of JNK1 function because the hypothalamus and pituitary gland are known to regulate metabolism, including feeding behavior, physical activity, and energy expenditure (Lenard and Berthoud 2008).The purpose of this study was to investigate the role of JNK1 in the brain. Our approach was to examine the effect of selective ablation of the Jnk1 gene in the mouse nervous system. We found that HFD-fed control (N WT ) mice gained substantially greater body weight than JNK1-deficient (N KO ) mice. The decreased weight gain by N KO mice was accounted for by decreased food intake, increased physical activity, and increased energy expenditure. These changes were associated with increased amounts of thyroid hormone in the blood and increased expression of thyroid hormone-responsive genes in target tissues. Importantly, pharmacological inhibition of thyroid hormone markedly attenuated N KO phenotypes. These data demonstrate that the hypothalamic-pituitary-thyroid axis is a major target of the JNK1 signaling pathway that regulates metabolis...

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