The fructose-2,6-bisphosphatase TIGAR suppresses NF-κB signaling by directly inhibiting the linear ubiquitin assembly complex LUBAC.

The systems integration of whole-body metabolism and immune signaling are central homeostatic mechanisms necessary for maintenance of normal physiology, and dysregulation of these processes leads to a variety of chronic disorders. However, the intracellular mechanisms responsible for cell-autonomous cross-talk between the inflammatory signaling pathways and metabolic flux have remained enigmatic. In this study, we discovered that the fructose-2,6-bisphosphatase TIGAR (Tp53-induced glycolysis and apoptosis regulator) critically regulates NF-κB activation. We found that TIGAR potently inhibits NF-κB-dependent gene expression by suppressing the upstream activation of IKKß phosphorylation and kinase activation. This inhibition occurred through a direct binding competition between NEMO and TIGAR for association with the linear ubiquitination assembly complex (LUBAC). This competition prevented linear ubiquitination of NEMO, which is required for activation of IKKß and other downstream targets. Furthermore, a TIGAR phosphatase activity-deficient mutant was equally effective as WT TIGAR in inhibiting NEMO linear ubiquitination, IKKß phosphorylation/activation, and NF-κB signaling, indicating that TIGAR's effect on NF-κB signaling is due to its interaction with LUBAC. Physiologically, TIGAR knockout mice displayed enhanced adipose tissue NF-κB signaling, whereas adipocyte-specific overexpression of TIGAR suppressed adipose tissue NF-κB signaling. Together, these results demonstrate that TIGAR has a nonenzymatic molecular function that modulates the NF-κB signaling pathway by directly inhibiting the E3 ligase activity of LUBAC.

Identification of Atypical Peri-Nuclear Multivesicular Bodies in Oxidative and Glycolytic Skeletal Muscle of Aged and Pompe's Disease Mouse Models.

Muscle wasting that occurs during aging or from disease pathology presents with an accumulation of lipid species termed ceroid or lipofuscin. This unique species of lipid has been characterized in various cell types but its properties and organization in skeletal muscle remains unclear. Using immunofluorescence and transmission electron microscopy, we were able to visualize and characterize an atypical lipid storing organelle in skeletal muscle. White myofibers contain two organelles at each pole of the myonuclei and red myofibers contain many of these structures in and around the perinuclear space. These organelles contain markers for late endosomes, are morphologically similar to multivesicular bodies, store lipid, and hypertrophy in aged muscle and a model of muscle wasting with an accumulation of large amounts of lipofuscin. Rapamycin treatment reduces the multivesicular body hypertrophy, restores late endosomal protein markers, and also increases the number and intensity of lipofuscin deposits. Together, these data demonstrate for the first time a perinuclear organelle in skeletal muscle that hypertrophies in muscle wasting phenotypes and is involved in endocytic lipid storage.

Dietary exposure to the endocrine disruptor tolylfluanid promotes global metabolic dysfunction in male mice.

Environmental endocrine disruptors are implicated as putative contributors to the burgeoning metabolic disease epidemic. Tolylfluanid (TF) is a commonly detected fungicide in Europe, and previous in vitro and ex vivo work has identified it as a potent endocrine disruptor with the capacity to promote adipocyte differentiation and induce adipocytic insulin resistance, effects likely resulting from activation of glucocorticoid receptor signaling. The present study extends these findings to an in vivo mouse model of dietary TF exposure. After 12 weeks of consumption of a normal chow diet supplemented with 100 parts per million TF, mice exhibited increased body weight gain and an increase in total fat mass, with a specific augmentation in visceral adipose depots. This increased adipose accumulation is proposed to occur through a reduction in lipolytic and fatty acid oxidation gene expression. Dietary TF exposure induced glucose intolerance, insulin resistance, and metabolic inflexibility, while also disrupting diurnal rhythms of energy expenditure and food consumption. Adipose tissue endocrine function was also impaired with a reduction in serum adiponectin levels. Moreover, adipocytes from TF-exposed mice exhibited reduced insulin sensitivity, an effect likely mediated through a specific down-regulation of insulin receptor substrate-1 expression, mirroring effects of ex vivo TF exposure. Finally, gene set enrichment analysis revealed an increase in adipose glucocorticoid receptor signaling with TF treatment. Taken together, these findings identify TF as a novel in vivo endocrine disruptor and obesogen in mice, with dietary exposure leading to alterations in energy homeostasis that recapitulate many features of the metabolic syndrome.

Skeletal muscle autophagy: a new metabolic regulator.

Autophagy classically functions as a physiological process to degrade cytoplasmic components, protein aggregates, and/or organelles, as a mechanism for nutrient breakdown, and as a regulator of cellular architecture. Proper autophagic flux is vital for both functional skeletal muscle, which controls the support and movement of the skeleton, and muscle metabolism. The role of autophagy as a metabolic regulator in muscle has been previously studied; however, the underlying molecular mechanisms that control autophagy in skeletal muscle have only recently begun to emerge. We review recent literature on the molecular pathways controlling skeletal muscle autophagy and discuss how they connect autophagy to metabolic regulation. We also focus on the implications these studies hold for understanding metabolic and muscle-wasting diseases.

The endocrine disrupting chemical tolylfluanid alters adipocyte metabolism via glucocorticoid receptor activation.

Glucocorticoid signaling plays a critical role in regulating energy metabolism. Emerging data implicate environmental endocrine-disrupting chemicals as contributors to the obesity and diabetes epidemics. Previous studies have shown that the phenylsulfamide fungicide tolylfluanid (TF) augments glucocorticoid receptor (GR)-dependent luciferase expression in 3T3-L1 preadipocytes while modulating insulin action in primary murine and human adipocytes. Studies were performed to interrogate glucocorticoid signaling in primary adipocytes exposed to TF. TF mimicked the gene transcription profile of the murine glucocorticoid corticosterone (Cort). Cellular fractionation assays demonstrated that TF treatment promoted the activating serine phosphorylation of GR, augmenting its cytoplasmic-to-nuclear translocation as well as its enrichment at glucocorticoid response elements on the glucocorticoid-induced leucine zipper gene promoter. After acute treatment, Cort or TF promoted insulin receptor substrate-1 (IRS-1) gene and protein expression. Either treatment also enriched GR binding at an identified glucocorticoid response element in the IRS-1 gene. TF or Cort each increased insulin-stimulated lipogenesis, an effect resulting from increased lipogenic gene expression and enhanced insulin-stimulated dephosphorylation of acetyl-coenzyme A carboxylase. The augmentation of insulin-stimulated lipogenesis was mediated through a specific enhancement of Akt phosphorylation at T308. These findings support modulation of IRS-1 levels as a mechanism for glucocorticoid-mediated changes in insulin action in primary adipocytes. Albeit with less affinity than Cort, in silico analysis suggests that TF can interact with the ligand binding pocket of GR. Collectively, these studies identify TF as a structurally unique environmental glucocorticoid. Glucocorticoid signaling may thus represent a novel pathway by which environmental toxicants promote the development of metabolic diseases.

The novel endocrine disruptor tolylfluanid impairs insulin signaling in primary rodent and human adipocytes through a reduction in insulin receptor substrate-1 levels.

Emerging data suggest that environmental endocrine disrupting chemicals may contribute to the pathophysiology of obesity and diabetes. In a prior work, the phenylsulfamide fungicide tolylfluanid (TF) was shown to augment adipocyte differentiation, yet its effects on mature adipocyte metabolism remain unknown. Because of the central role of adipose tissue in global energy regulation, the present study tested the hypothesis that TF modulates insulin action in primary rodent and human adipocytes. Alterations in insulin signaling in primary mammalian adipocytes were determined by the phosphorylation of Akt, a critical insulin signaling intermediate. Treatment of primary murine adipose tissue in vitro with 100nM TF for 48h markedly attenuated acute insulin-stimulated Akt phosphorylation in a strain- and species-independent fashion. Perigonadal, perirenal, and mesenteric fat were all sensitive to TF-induced insulin resistance. A similar TF-induced reduction in insulin-stimulated Akt phosphorylation was observed in primary human subcutaneous adipose tissue. TF treatment led to a potent and specific reduction in insulin receptor substrate-1 (IRS-1) mRNA and protein levels, a key upstream mediator of insulin's diverse metabolic effects. In contrast, insulin receptor-ß, phosphatidylinositol 3-kinase, and Akt expression were unchanged, indicating a specific abrogation of insulin signaling. Additionally, TF-treated adipocytes exhibited altered endocrine function with a reduction in both basal and insulin-stimulated leptin secretion. These studies demonstrate that TF induces cellular insulin resistance in primary murine and human adipocytes through a reduction of IRS-1 expression and protein stability, raising concern about the potential for this fungicide to disrupt metabolism and thereby contribute to the pathogenesis of diabetes.