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

The binding of tumor necrosis factor α (TNFα) to cell surface receptors engages multiple signal transduction pathways, including three groups of mitogen-activated protein (MAP) kinases: extracellular-signal-regulated kinases (ERKs); the cJun NH2-terminal kinases (JNKs); and the p38 MAP kinases. These MAP kinase signalling pathways induce a secondary response by increasing the expression of several inflammatory cytokines (including TNFα) that contribute to the biological activity of TNFα. MAP kinases therefore function both upstream and down-stream of signalling by TNFα receptors. Here we review mechanisms that mediate these actions of MAP kinases during the response to TNFα.

Glycogen synthase kinase 3β (GSK3β) is involved in metabolism, neurodegeneration, and cancer. Inhibition of GSK3β activity is the primary mechanism that regulates this widely expressed active kinase. Although the protein kinase Akt inhibits GSK3β by phosphorylation at the N terminus, preventing Akt-mediated phosphorylation does not affect the cell-survival pathway activated through the GSK3β substrate β-catenin. Here, we show that p38 mitogen-activated protein kinase (MAPK) also inactivates GSK3β by direct phosphorylation at its C terminus, and this inactivation can lead to an accumulation of β-catenin. p38 MAPK-mediated phosphorylation of GSK3β occurs primarily in the brain and thymocytes. Activation of β-catenin-mediated signaling through GSK3β inhibition provides a potential mechanism for p38 MAPK-mediated survival in specific tissues.The p38 mitogen-activated protein kinase (MAPK) is activated through phosphorylation primarily by MAPK kinase 3 (MKK3) and MKK6 in response to cellular stress and cytokines. The p38 MAPK pathway functions in the control of differentiation, the blockade of proliferation, and in the induction of apoptosis (1). It is also activated in response to DNA double-stranded breaks (DSBs) induced by ionizing irradiation or chemotherapeutic drugs, and it participates in the induction of a G 2 /M cell-cycle checkpoint (2,3). p38 MAPK can also promote survival (4-6) by unknown mechanisms. During T cell receptor β (TCRβ) rearrangement, V(D)J recombination-mediated DSBs also activate p38 MAPK in immature thymocytes at the double negative 3 (DN3) stage of development (7,8). The expression of a constitutively active mutant of MKK6 [MKK6(Glu)] in thymocytes of transgenic mice (MKK6 transgenic mice) activates a p53-mediated G 2 /M phase cell-cycle checkpoint (8). Like recombination-activating gene (Rag) deficiency, persistent activation of p38 MAPK interferes with the differentiation of thymocytes beyond the DN3 stage. However, MKK6 transgenic thymocytes (but not Rag -/-thymocytes) survive and accumulate in vivo (8), suggesting that

SummaryThyroid hormones (THs) act in the brain to modulate energy balance. We show that central triiodothyronine (T3) regulates de novo lipogenesis in liver and lipid oxidation in brown adipose tissue (BAT) through the parasympathetic (PSNS) and sympathetic nervous system (SNS), respectively. Central T3 promotes hepatic lipogenesis with parallel stimulation of the thermogenic program in BAT. The action of T3 depends on AMP-activated protein kinase (AMPK)-induced regulation of two signaling pathways in the ventromedial nucleus of the hypothalamus (VMH): decreased ceramide-induced endoplasmic reticulum (ER) stress, which promotes BAT thermogenesis, and increased c-Jun N-terminal kinase (JNK) activation, which controls hepatic lipid metabolism. Of note, ablation of AMPKα1 in steroidogenic factor 1 (SF1) neurons of the VMH fully recapitulated the effect of central T3, pointing to this population in mediating the effect of central THs on metabolism. Overall, these findings uncover the underlying pathways through which central T3 modulates peripheral metabolism.

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...

Activation of the p38 MAP kinase pathways is crucial for the adaptation of mammalian cells to changes in the osmolarity of the environment. Here we identify SAP97/ hDlg, the mammalian homologue of the Drosophila tumour suppressor Dlg, as a physiological substrate for the p38c MAP kinase (SAPK3/p38c) isoform. SAP97/hDlg is a scaffold protein that forms multiprotein complexes with a variety of proteins and is targeted to the cytoskeleton by its association with the protein guanylate kinase-associated protein (GKAP). The SAPK3/p38c-catalysed phosphorylation of SAP97/hDlg triggers its dissociation from GKAP and therefore releases it from the cytoskeleton. This is likely to regulate the integrity of intercellular-junctional complexes, and cell shape and volume in response to osmotic stress.

The compound BIRB796 inhibits the stress-activated protein kinases p38␣ and p38␤ and is undergoing clinical trials for the treatment of inflammatory diseases. Here we report that BIRB796 also inhibits the activity and the activation of SAPK3/p38␥. This occurs at higher concentrations of BIRB796 than those that inhibit p38␣ and p38␤ and at lower concentrations than those that inhibit the activation of JNK isoforms. We also show that at these concentrations, BIRB796 blocks the stressinduced phosphorylation of the scaffold protein SAP97, further establishing that this is a physiological substrate of SAPK3/p38␥. Our results demonstrate that BIRB796, in combination with SB203580, a compound that inhibits p38␣ and p38␤, but not the other p38 isoforms, can be used to identify physiological substrates of SAPK3/p38␥ as well as those of p38␣ and p38␤.The stress-activated protein kinase (SAPK) 1 p38 isoforms are mitogen-activated protein kinase (MAPK) family members that are activated by changes in the cellular environment, such as alterations in the concentration of nutrients, cytokines, celldamaging agents, and changes in osmolarity of the surrounding medium (1). They comprise p38␣, p38␤, SAPK3/p38␥ (also known as ERK6), and SAPK4/p38␦. Each p38 isoform may have different biological functions and different physiological substrates, but they all phosphorylate substrates containing the minimal consensus sequence Ser/Thr-Pro. A major challenge of current research in this field is to identify the downstream physiological substrates and processes that each p38 MAPK regulates in the cell, as well as determining which "upstream" components regulate their activities. One of the most successful aids to the identification of physiological substrates has been the use of small cell-permeable compounds that are specific inhibitors of particular protein kinases. These compounds enter cells within minutes and act rapidly to suppress the activity of a particular kinase so that indirect effects caused, for example, by changes in gene expression or protein activity, a potential risk when cells deficient in a particular kinase are used, are excluded. Moreover, the use of protein kinase inhibitors avoids the need for transfection-based approaches, which have the potential to give misleading results since the fidelity of signaling can break down when components are overexpressed.Identification of physiological substrates for p38␣ and p38␤ has been greatly facilitated by the availability of specific inhibitors of these enzymes, such as the cell-permeant pyridinyl imidazole SB203580 and related compounds (2, 3). Substrates for p38␣ and p38␤ include other protein kinases, as well as several transcription factors and metabolic protein (4, 5). However, little is known about the physiological substrates for SAPK3/p38␥ and SAPK4/p38␦ as they are not inhibited by SB203580 (6, 7), and so far there are not any commercially available inhibitors for these kinases. Nevertheless, we have recently demonstrated that the synapse-associated proteins SAP90 and...

The cJun NH2-terminal kinase (JNK) stress signaling pathway is implicated in the metabolic response to the consumption of a high fat diet, including the development of obesity and insulin resistance. These metabolic adaptations involve altered liver function. Here we demonstrate that hepatic JNK potently represses the nuclear hormone receptor peroxisome proliferator-activated receptor α (PPARα). JNK therefore causes decreased expression of PPARα target genes that increase fatty acid oxidation / ketogenesis and promote the development of insulin resistance. We show that the PPARα target gene fibroblast growth factor 21 (Fgf21) plays a key role in this response because disruption of the hepatic PPARα - FGF21 hormone axis suppresses the metabolic effects of JNK-deficiency. This analysis identifies the hepatokine FGF21 as a critical mediator of JNK signaling in the liver.