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The t-SNARE protein SNAP23 conventionally functions as a component of the cellular machinery required for intracellular transport vesicle fusion with target membranes and has been implicated in the regulation of fasting glucose levels, BMI, and type 2 diabetes. Surprisingly, we observed that adipocyte-specific KO of SNAP23 in mice resulted in a temporal development of severe generalized lipodystrophy associated with adipose tissue inflammation, insulin resistance, hyperglycemia, liver steatosis, and early death. This resulted from adipocyte cell death associated with an inhibition of macroautophagy and lysosomal degradation of the proapoptotic regulator BAX, with increased BAX activation. BAX colocalized with LC3- positive autophagic vacuoles and was increased upon treatment with lysosome inhibitors. Moreover, BAX deficiency suppressed the lipodystrophic phenotype in the adipocyte-specific SNAP23-KO mice and prevented cell death. In addition, ATG9 deficiency phenocopied SNAP23 deficiency, whereas ATG7 deficiency had no effect on BAX protein levels, BAX activation, or apoptotic cell death. These data demonstrate a role for SNAP23 in the control of macroautophagy and programmed cell death through an ATG9-dependent, but ATG7-independent, pathway regulating BAX protein levels and BAX activation., Introduction SNARE complexes are a large group of membrane-bound proteins that regulate the fusion of intracellular transport vesicles with their target membranes (1-3). All SNAREs contain a characteristic coiled-coil SNARE [...]

Altered lipid metabolism underlies several major human diseases, including obesity and type 2 diabetes. However, lipid metabolism pathophysiology remains poorly understood at the molecular level. Insulin is the primary stimulator of hepatic lipogenesis through activation of the SREBP-1c transcription factor. Here we identified cyclin-dependent kinase 8 (CDK8) and its regulatory partner cyclin C (CycC) as negative regulators of the lipogenic pathway in Drosophila, mammalian hepatocytes, and mouse liver. The inhibitory effect of CDK8 and CycC on de novo lipogenesis was mediated through CDK8 phosphorylation of nuclear SREBP-1c at a conserved threonine residue. Phosphorylation by CDK8 enhanced SREBP-1c ubiquitination and protein degradation. Importantly, consistent with the physiologic regulation of lipid biosynthesis, CDK8 and CycC proteins were rapidly downregulated by feeding and insulin, resulting in decreased SREBP-1c phosphorylation. Moreover, overexpression of CycC efficiently suppressed insulin and feeding-induced lipogenic gene expression. Taken together, these results demonstrate that CDK8 and CycC function as evolutionarily conserved components of the insulin signaling pathway in regulating lipid homeostasis., Introduction Dysregulation of lipid metabolism is closely associated with major human diseases such as obesity, type 2 diabetes, and cardiovascular disease (1-4). However, the physiologic and pathophysiologic regulation of lipid [...]

Although supraphysiological concentrations of urea are known to increase oxidative stress in cultured cells, it is generally thought that the elevated levels of urea in chronic renal failure patients have negligible toxicity. We previously demonstrated that ROS increase intracellular protein modification by O-linked β-N-acetyl-glucosamine (O-GlcNAc), and others showed that increased modification of insulin signaling molecules by O-GlcNAc reduces insulin signal transduction. Because both oxidative stress and insulin resistance have been observed in patients with end-stage renal disease, we sought to determine the role of urea in these phenotypes. Treatment of 3T3-L1 adipocytes with urea at disease-relevant concentrations induced ROS production, caused insulin resistance, increased expression of adipokines retinol binding protein 4 (RBP4) and resistin, and increased O-GlcNAc-modified insulin signaling molecules. Investigation of a mouse model of surgically induced renal failure (uremic mice) revealed increased ROS production, modification of insulin signaling molecules by O-GlcNAc, and increased expression of RBP4 and resistin in visceral adipose tissue. Uremic mice also displayed insulin resistance and glucose intolerance, and treatment with an antioxidant SOD/catalase mimetic normalized these defects. The SOD/catalase mimetic treatment also prevented the development of insulin resistance in normal mice after urea infusion. These data suggest that therapeutic targeting of urea-induced ROS may help reduce the high morbidity and mortality caused by end-stage renal disease., Introduction Elevated urea in chronic renal failure (CRF) is considered to have negligible toxicity because Johnson et al. found in 1972 that in patients with far-advanced renal failure, blood urea [...]

The relative balance between the quantity of white and brown adipose tissue can profoundly affect lipid storage and whole-body energy homeostasis. However, the mechanisms regulating the formation, expansion, and interconversion of these 2 distinct types of fat remain unknown. Recently, the lysosomal degradative pathway of macroautophagy has been identified as a regulator of cellular differentiation, suggesting that autophagy may modulate this process in adipocytes. The function of autophagy in adipose differentiation was therefore examined in the current study by genetic inhibition of the critical macroautophagy gene autophagy-related 7 (Atg7). Knockdown of Atg7 in 3T3-L1 preadipocytes inhibited lipid accumulation and decreased protein levels of adipocyte differentiation factors. Knockdown of Atg5 or pharmacological inhibition of autophagy or lysosome function also had similar effects. An adipocyte-specific mouse knockout of Atg7 generated lean mice with decreased white adipose mass and enhanced insulin sensitivity. White adipose tissue in knockout mice had increased features of brown adipocytes, which, along with an increase in normal brown adipose tissue, led to an elevated rate of fatty acid, β-oxidation, and a lean body mass. Autophagy therefore functions to regulate body lipid accumulation by controlling adipocyte differentiation and determining the balance between white and brown fat., Introduction Obesity is characterized by an expansion of adipose tissue mass resulting from increased adipocyte number and/or size. Lipids in the form of triglycerides (TG) accumulate in various anatomical locations [...]

Concerns were recently raised with regard to the immunoblot images in Figure 5B, and it was unclear to the Editorial Board whether the position of the bands in lane 2 [...]

The t-SNARE protein SNAP23 conventionally functions as a component of the cellular machinery required for intracellular transport vesicle fusion with target membranes and has been implicated in the regulation of fasting glucose levels, BMI, and type 2 diabetes. Surprisingly, we observed that adipocyte-specific KO of SNAP23 in mice resulted in a temporal development of severe generalized lipodystrophy associated with adipose tissue inflammation, insulin resistance, hyperglycemia, liver steatosis, and early death. This resulted from adipocyte cell death associated with an inhibition of macroautophagy and lysosomal degradation of the proapoptotic regulator BAX, with increased BAX activation. BAX colocalized with LC3-positive autophagic vacuoles and was increased upon treatment with lysosome inhibitors. Moreover, BAX deficiency suppressed the lipodystrophic phenotype in the adipocyte-specific SNAP23-KO mice and prevented cell death. In addition, ATG9 deficiency phenocopied SNAP23 deficiency, whereas ATG7 deficiency had no effect on BAX protein levels, BAX activation, or apoptotic cell death. These data demonstrate a role for SNAP23 in the control of macroautophagy and programmed cell death through an ATG9-dependent, but ATG7-independent, pathway regulating BAX protein levels and BAX activation. [ABSTRACT FROM AUTHOR]