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1.
Mol Cell ; 57(2): 235-46, 2015 Jan 22.
Article in English | MEDLINE | ID: mdl-25578880

ABSTRACT

Uncoupling protein 1 (UCP1) mediates nonshivering thermogenesis and, upon cold exposure, is induced in brown adipose tissue (BAT) and subcutaneous white adipose tissue (iWAT). Here, by high-throughput screening using the UCP1 promoter, we identify Zfp516 as a transcriptional activator of UCP1 as well as PGC1α, thereby promoting a BAT program. Zfp516 itself is induced by cold and sympathetic stimulation through the cAMP-CREB/ATF2 pathway. Zfp516 directly binds to the proximal region of the UCP1 promoter, not to the enhancer region where other transcription factors bind, and interacts with PRDM16 to activate the UCP1 promoter. Although ablation of Zfp516 causes embryonic lethality, knockout embryos still show drastically reduced BAT mass. Overexpression of Zfp516 in adipose tissue promotes browning of iWAT even at room temperature, increasing body temperature and energy expenditure and preventing diet-induced obesity. Zfp516 may represent a future target for obesity therapeutics.


Subject(s)
Adipose Tissue, Brown/physiology , Adipose Tissue, White/physiology , Ion Channels/genetics , Mitochondrial Proteins/genetics , Trans-Activators/physiology , Adipogenesis , Adipose Tissue, Brown/cytology , Adipose Tissue, White/cytology , Animals , Cold-Shock Response , DNA-Binding Proteins/metabolism , HEK293 Cells , Humans , Ion Channels/metabolism , Mice, Inbred C57BL , Mice, Transgenic , Mitochondrial Proteins/metabolism , Muscle Development , Phenotype , Promoter Regions, Genetic , Protein Binding , Thermogenesis , Transcription Factors/metabolism , Transcription, Genetic , Transcriptional Activation , Uncoupling Protein 1
2.
PLoS One ; 6(11): e26415, 2011.
Article in English | MEDLINE | ID: mdl-22073164

ABSTRACT

Endocannabinoids regulate energy balance and lipid metabolism by stimulating the cannabinoid receptor type 1 (CB1). Genetic deletion and pharmacological antagonism have shown that CB1 signaling is necessary for the development of obesity and related metabolic disturbances. However, the sufficiency of endogenously produced endocannabinoids to cause hepatic lipid accumulation and insulin resistance, independent of food intake, has not been demonstrated. Here, we show that a single administration of isopropyl dodecylfluorophosphonate (IDFP), perhaps the most potent pharmacological inhibitor of endocannabinoid degradation, increases hepatic triglycerides (TG) and induces insulin resistance in mice. These effects involve increased CB1 signaling, as they are mitigated by pre-administration of a CB1 antagonist (AM251) and in CB1 knockout mice. Despite the strong physiological effects of CB1 on hepatic lipid and glucose metabolism, little is known about the downstream targets responsible for these effects. To elucidate transcriptional targets of CB1 signaling, we performed microarrays on hepatic RNA isolated from DMSO (control), IDFP and AM251/IDFP-treated mice. The gene for the secreted glycoprotein lipocalin 2 (lcn2), which has been implicated in obesity and insulin resistance, was among those most responsive to alterations in CB1 signaling. The expression pattern of IDFP mice segregated from DMSO mice in hierarchal cluster analysis and AM251 pre-administration reduced (>50%) the majority (303 of 533) of the IDFP induced alterations. Pathway analysis revealed that IDFP altered expression of genes involved in lipid, fatty acid and steroid metabolism, the acute phase response, and amino acid metabolism in a CB1-dependent manner. PCR confirmed array results of key target genes in multiple independent experiments. Overall, we show that acute IDFP treatment induces hepatic TG accumulation and insulin resistance, at least in part through the CB1 receptor, and identify novel cannabinoid responsive genes.


Subject(s)
Cannabinoid Receptor Modulators/metabolism , Endocannabinoids , Fatty Liver/metabolism , Glucose Tolerance Test , Receptor, Cannabinoid, CB1/metabolism , Signal Transduction , Animals , Insulin Resistance , Mice , Polymerase Chain Reaction
3.
Cell Metab ; 13(6): 739-48, 2011 Jun 08.
Article in English | MEDLINE | ID: mdl-21641555

ABSTRACT

While fatty acids (FAs) released by white adipose tissue (WAT) provide energy for other organs, lipolysis is also critical in brown adipose tissue (BAT), generating FAs for oxidation and UCP-1 activation for thermogenesis. Here we show that adipose-specific ablation of desnutrin/ATGL in mice converts BAT to a WAT-like tissue. These mice exhibit severely impaired thermogenesis with increased expression of WAT-enriched genes but decreased BAT genes, including UCP-1 with lower PPARα binding to its promoter, revealing the requirement of desnutrin-catalyzed lipolysis for maintaining a BAT phenotype. We also show that desnutrin is phosphorylated by AMPK at S406, increasing TAG hydrolase activity, and provide evidence for increased lipolysis by AMPK phosphorylation of desnutrin in adipocytes and in vivo. Despite adiposity and impaired BAT function, desnutrin-ASKO mice have improved hepatic insulin sensitivity with lower DAG levels. Overall, desnutrin is phosphorylated/activated by AMPK to increase lipolysis and brings FA oxidation and UCP-1 induction for thermogenesis.


Subject(s)
Adenylate Kinase/metabolism , Lipase/genetics , Protein Processing, Post-Translational , Adipose Tissue, Brown/metabolism , Adipose Tissue, White/metabolism , Adiposity/genetics , Animals , Cells, Cultured , Enzyme Assays , Gene Knockout Techniques , Ion Channels/genetics , Ion Channels/metabolism , Lipase/metabolism , Lipolysis/genetics , Male , Mice , Mice, Obese , Mitochondrial Proteins/genetics , Mitochondrial Proteins/metabolism , Phosphorylation , Thermogenesis , Uncoupling Protein 1
4.
Cell ; 136(6): 1056-72, 2009 Mar 20.
Article in English | MEDLINE | ID: mdl-19303849

ABSTRACT

Fatty acid synthase (FAS) is a central enzyme in lipogenesis and transcriptionally activated in response to feeding and insulin signaling. The transcription factor USF is required for the activation of FAS transcription, and we show here that USF phosphorylation by DNA-PK, which is dephosphorylated by PP1 in response to feeding, triggers a switch-like mechanism. Under fasting conditions, USF-1 is deacetylated by HDAC9, causing promoter inactivation. In contrast, feeding induces the recruitment of DNA-PK to USF-1 and its phosphorylation, which then allows recruitment of P/CAF, resulting in USF-1 acetylation and FAS promoter activation. DNA break/repair components associated with USF induce transient DNA breaks during FAS activation. In DNA-PK-deficient SCID mice, feeding-induced USF-1 phosphorylation/acetylation, DNA breaks, and FAS activation leading to lipogenesis are impaired, resulting in decreased triglyceride levels. Our study demonstrates that a kinase central to the DNA damage response mediates metabolic gene activation.


Subject(s)
DNA-Activated Protein Kinase/metabolism , Insulin/metabolism , Acetylation , Animals , Cell Line, Tumor , Eating/physiology , Fasting/metabolism , Fatty Acid Synthases/metabolism , Histone Deacetylases/metabolism , Humans , Mice , Mice, SCID , Phosphorylation , Promoter Regions, Genetic , Repressor Proteins/metabolism , Upstream Stimulatory Factors/metabolism
5.
Proc Natl Acad Sci U S A ; 105(38): 14561-6, 2008 Sep 23.
Article in English | MEDLINE | ID: mdl-18794527

ABSTRACT

The endocannabinoid (EC) system regulates food intake and energy metabolism. Cannabinoid receptor type 1 (CB1) antagonists show promise in the treatment of obesity and its metabolic consequences. Although the reduction in adiposity resulting from therapy with CB1 antagonists may not account fully for the concomitant improvements in dyslipidemia, direct effects of overactive EC signaling on plasma lipoprotein metabolism have not been documented. The present study used a chemical approach to evaluate the direct effects of increased EC signaling in mice by inducing acute elevations of endogenously produced cannabinoids through pharmacological inhibition of their enzymatic hydrolysis by isopropyl dodecylfluorophosphonate (IDFP). Acute IDFP treatment increased plasma levels of triglyceride (TG) (2.0- to 3.1-fold) and cholesterol (1.3- to 1.4-fold) in conjunction with an accumulation in plasma of apolipoprotein (apo)E-depleted TG-rich lipoproteins. These changes did not occur in either CB1-null or apoE-null mice, were prevented by pretreatment with CB1 antagonists, and were not associated with reduced hepatic apoE gene expression. Although IDFP treatment increased hepatic mRNA levels of lipogenic genes (Srebp1 and Fas), there was no effect on TG secretion into plasma. Instead, IDFP treatment impaired clearance of an intravenously administered TG emulsion, despite increased postheparin lipoprotein lipase activity. Therefore, overactive EC signaling elicits an increase in plasma triglyceride levels associated with reduced plasma TG clearance and an accumulation in plasma of apoE-depleted TG-rich lipoproteins. These findings suggest a role of CB1 activation in the pathogenesis of obesity-related hypertriglyceridemia and underscore the potential efficacy of CB1 antagonists in treating metabolic disease.


Subject(s)
Apolipoproteins E/metabolism , Cannabinoid Receptor Modulators/metabolism , Endocannabinoids , Lipoproteins/metabolism , Signal Transduction , Triglycerides/metabolism , Animals , Arachidonic Acids/metabolism , Cholesterol/blood , Enzyme Inhibitors/pharmacology , Gene Expression Regulation/drug effects , Glycerides/metabolism , Lipase/metabolism , Lipid Metabolism/drug effects , Lipids/blood , Lipoproteins/blood , Liver/drug effects , Liver/enzymology , Liver/metabolism , Male , Mice , Organophosphonates/pharmacology , Receptor, Cannabinoid, CB1/metabolism , Signal Transduction/drug effects , Triglycerides/blood
6.
Bioorg Med Chem Lett ; 18(22): 5875-8, 2008 Nov 15.
Article in English | MEDLINE | ID: mdl-18752948

ABSTRACT

The structure-activity relationships of organophosphorus (OP) and organosulfur compounds were examined in vitro and in vivo as inhibitors of mouse brain monoacylglycerol lipase (MAGL) hydrolysis of 2-arachidonoylglycerol (2-AG) and agonist binding at the CB1 receptor. Several compounds showed exceptional potency toward MAGL activity with IC(50) values of 0.1-10 nM in vitro and high inhibition at 10mg/kg intraperitoneally in mice. We find for the first time that MAGL activity is a major in vivo determinant of 2-AG and arachidonic acid levels not only in brain but also in spleen, lung, and liver. Apparent direct OP inhibition of CB1 agonist binding may be due instead to metabolic stabilization of 2-AG in brain membranes as the actual inhibitor.


Subject(s)
Arachidonic Acid/analysis , Arachidonic Acids/analysis , Brain/drug effects , Glycerides/analysis , Monoacylglycerol Lipases/antagonists & inhibitors , Animals , Arachidonic Acid/metabolism , Arachidonic Acids/metabolism , Brain/enzymology , Cell Membrane/drug effects , Endocannabinoids , Glycerides/metabolism , Inhibitory Concentration 50 , Mice , Molecular Structure , Monoacylglycerol Lipases/metabolism , Organophosphorus Compounds/chemistry , Organophosphorus Compounds/pharmacology , Receptor, Cannabinoid, CB1/metabolism , Structure-Activity Relationship , Sulfur Compounds/chemistry , Sulfur Compounds/pharmacology
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