Coactivator Comparative Gene Identification-58 Research Articles

Adipose Lipolysis Regulates Cardiac Glucose Uptake and Function in Mice under Cold Stress.

The heart primarily uses fatty acids as energy substrates. Adipose lipolysis is a major source of fatty acids, particularly under stress conditions. In this study, we showed that mice with selective inactivation of the lipolytic coactivator comparative gene identification-58 (CGI-58) in adipose tissue (FAT-KO mice), relative to their littermate controls, had lower circulating FA levels in the fed and fasted states due to impaired adipose lipolysis. They preferentially utilized carbohydrates as energy fuels and were more insulin sensitive and glucose tolerant. Under cold stress, FAT-KO versus control mice had >10-fold increases in glucose uptake in the hearts but no increases in other tissues examined. Plasma concentrations of atrial natriuretic peptide and cardiac mRNAs for atrial and brain-type natriuretic peptides, two sensitive markers of cardiac remodeling, were also elevated. After one week of cold exposure, FAT-KO mice showed reduced cardiac expression of several mitochondrial oxidative phosphorylation proteins. After one month of cold exposure, hearts of these animals showed depressed functions, reduced SERCA2 protein, and increased proteins for MHC-β, collagen I proteins, Glut1, Glut4 and phospho-AMPK. Thus, CGI-58-dependent adipose lipolysis critically regulates cardiac metabolism and function, especially during cold adaptation. The adipose-heart axis may be targeted for the management of cardiac dysfunction.

ATGL/CGI-58-Dependent Hydrolysis of a Lipid Storage Pool in Murine Enterocytes

SummaryAs circulating lipid levels are balanced by the rate of lipoprotein release and clearance from the plasma, lipid absorption in the small intestine critically contributes to the maintenance of whole-body lipid homeostasis. Within enterocytes, excessive triglycerides are transiently stored as cytosolic lipid droplets (cLDs), and their mobilization sustains lipid supply during interprandial periods. Using mice lacking adipose triglyceride lipase (ATGL) and its coactivator comparative gene identification-58 (CGI-58) exclusively in the intestine (intestine-specific double KO [iDKO]), we show that ATGL/CGI-58 are not involved in providing substrates for chylomicron synthesis. Massive intestinal cLD accumulation in iDKO mice independent of dietary lipids together with inefficient lipid incorporation into cLDs in the early absorption phase demonstrate the existence of a secretion/re-uptake cycle, corroborating the availability of two diverse cLD pools. This study identified ATGL/CGI-58 as critical players in the catabolism of basolaterally (blood) derived lipids and highlights the necessity to modify the current model of intestinal lipid metabolism.

Adipocyte STAT5 deficiency promotes adiposity and impairs lipid mobilisation in mice

Aims/hypothesisDysfunction of lipid metabolism in white adipose tissue can substantially interfere with health and quality of life, for example in obesity and associated metabolic diseases. Therefore, it is important to characterise pathways that regulate lipid handling in adipocytes and determine how they affect metabolic homeostasis. Components of the Janus kinase (JAK)–signal transducer and activator of transcription (STAT) pathway are involved in adipocyte physiology and pathophysiology. However, the exact physiological importance of the STAT family member STAT5 in white adipose tissue is yet to be determined. Here, we aimed to delineate adipocyte STAT5 functions in the context of lipid metabolism in white adipose tissue.MethodsWe generated an adipocyte specific knockout of Stat5 in mice using the Adipoq-Cre recombinase transgene followed by in vivo and in vitro biochemical and molecular studies.ResultsAdipocyte-specific deletion of Stat5 resulted in increased adiposity, while insulin resistance and gluconeogenic capacity was decreased, indicating that glucose metabolism can be improved by interfering with adipose STAT5 function. Basal lipolysis and fasting-induced lipid mobilisation were diminished upon STAT5 deficiency, which coincided with reduced levels of the rate-limiting lipase of triacylglycerol hydrolysis, adipose triglyceride lipase (ATGL, encoded by Pnpla2) and its coactivator comparative gene identification 58 (CGI-58). In a mechanistic analysis, we identified a functional STAT5 response element within the Pnpla2 promoter, indicating that Pnpla2 is transcriptionally regulated by STAT5.Conclusions/interpretationOur findings reveal an essential role for STAT5 in maintaining lipid homeostasis in white adipose tissue and provide a rationale for future studies into the potential of STAT5 manipulation to improve outcomes in metabolic diseases.

Skin Barrier Development Depends on CGI-58 Protein Expression during Late-Stage Keratinocyte Differentiation

Adipose triglyceride lipase (ATGL) and its coactivator comparative gene identification-58 (CGI-58) are limiting in cellular triglyceride catabolism. Although ATGL deficiency is compatible with normal skin development, mice globally lacking CGI-58 die postnatally and exhibit a severe epidermal permeability barrier defect, which may originate from epidermal and/or peripheral changes in lipid and energy metabolism. Here, we show that epidermis-specific disruption of CGI-58 is sufficient to provoke a defect in the formation of a functional corneocyte lipid envelope linked to impaired ω-O-acylceramide synthesis. As a result, epidermis-specific CGI-58-deficient mice show severe skin dysfunction, arguing for a tissue autonomous cause of disease development. Defective skin permeability barrier formation in global CGI-58-deficient mice could be reversed via transgenic restoration of CGI-58 expression in differentiated but not basal keratinocytes suggesting that CGI-58 is essential for lipid metabolism in suprabasal epidermal layers. The compatibility of ATGL deficiency with normal epidermal function indicated that CGI-58 may stimulate an epidermal triglyceride lipase beyond ATGL required for the adequate provision of fatty acids as a substrate for ω-O-acylceramide synthesis. Pharmacological inhibition of ATGL enzyme activity similarly reduced triglyceride-hydrolytic activities in wild-type and CGI-58 overexpressing epidermis implicating that CGI-58 participates in ω-O-acylceramide biogenesis independent of its role as a coactivator of epidermal triglyceride catabolism.

Pronounced expression of the lipolytic inhibitor G0/G1 Switch Gene 2 (G0S2) in adipose tissue from brown bears (Ursus arctos) prior to hibernation

Prior to hibernation, the brown bear (Ursus arctos) exhibits unparalleled weight gain. Unlike humans, weight gain in bears is associated with lower levels of circulating free fatty acids (FFA) and increased insulin sensitivity. Understanding how free‐ranging brown bears suppress lipolysis when gaining weight may therefore provide novel insight toward the development of human therapies. Blood and subcutaneous adipose tissue were collected from immobilized free‐ranging brown bears (fitted with GPS‐collars) during hibernation in winter and from the same bears during the active period in summer in Dalarna, Sweden. The expression of lipid droplet‐associated proteins in adipose tissue was examined under the hypothesis that bears suppress lipolysis during summer while gaining weight by increased expression of negative regulators of lipolysis. Adipose triglyceride lipase (ATGL) expression did not differ between seasons, but in contrast, the expression of ATGL coactivator Comparative gene identification‐58 (CGI‐58) was lower in summer. In addition, the expression of the negative regulators of lipolysis, G0S2 and cell‐death inducing DNA fragmentation factor‐a‐like effector (CIDE)C markedly increased during summer. Free‐ranging brown bears display potent upregulation of inhibitors of lipolysis in adipose tissue during summer. This is a potential mechanism for increased insulin sensitivity during weight gain and G0S2 may serve as a target to modulate insulin sensitivity.

Fasting-induced G0/G1 switch gene 2 and FGF21 expression in the liver are under regulation of adipose tissue derived fatty acids

Adipose tissue (AT)-derived fatty acids (FAs) are utilized for hepatic triacylglycerol (TG) generation upon fasting. However, their potential impact as signaling molecules is not established. Herein we examined the role of exogenous AT-derived FAs in the regulation of hepatic gene expression by investigating mice with a defect in AT-derived FA supply to the liver. Plasma FA levels, tissue TG hydrolytic activities and lipid content were determined in mice lacking the lipase co-activator comparative gene identification-58 (CGI-58) selectively in AT (CGI-58-ATko) applying standard protocols. Hepatic expression of lipases, FA oxidative genes, transcription factors, ER stress markers, hormones and cytokines were determined by qRT-PCR, Western blotting and ELISA. Impaired AT-derived FA supply upon fasting of CGI-58-ATko mice causes a marked defect in liver PPARα-signaling and nuclear CREBH translocation. This severely reduced the expression of respective target genes such as the ATGL inhibitor G0/G1 switch gene-2 (G0S2) and the endocrine metabolic regulator FGF21. These changes could be reversed by lipid administration and raising plasma FA levels. Impaired AT-lipolysis failed to induce hepatic G0S2 expression in fasted CGI-58-ATko mice leading to enhanced ATGL-mediated TG-breakdown strongly reducing hepatic TG deposition. On high fat diet, impaired AT-lipolysis counteracts hepatic TG accumulation and liver stress linked to improved systemic insulin sensitivity. AT-derived FAs are a critical regulator of hepatic fasting gene expression required for the induction of G0S2-expression in the liver to control hepatic TG-breakdown. Interfering with AT-lipolysis or hepatic G0S2 expression represents an effective strategy for the treatment of hepatic steatosis.

Deletion of CGI-58 or adipose triglyceride lipase differently affects macrophage function and atherosclerosis

Cellular TG stores are efficiently hydrolyzed by adipose TG lipase (ATGL). Its coactivator comparative gene identification-58 (CGI-58) strongly increases ATGL-mediated TG catabolism in cell culture experiments. To investigate the consequences of CGI-58 deficiency in murine macrophages, we generated mice with a targeted deletion of CGI-58 in myeloid cells (macCGI-58−/− mice). CGI-58−/− macrophages accumulate intracellular TG-rich lipid droplets and have decreased phagocytic capacity, comparable to ATGL−/− macrophages. In contrast to ATGL−/− macrophages, however, CGI-58−/− macrophages have intact mitochondria and show no indications of mitochondrial apoptosis and endoplasmic reticulum stress, suggesting that TG accumulation per se lacks a significant role in processes leading to mitochondrial dysfunction. Another notable difference is the fact that CGI-58−/− macrophages adopt an M1-like phenotype in vitro. Finally, we investigated atherosclerosis susceptibility in macCGI-58/ApoE-double KO (DKO) animals. In response to high-fat/high-cholesterol diet feeding, DKO animals showed comparable plaque formation as observed in ApoE−/− mice. In agreement, antisense oligonucleotide-mediated knockdown of CGI-58 in LDL receptor−/− mice did not alter atherosclerosis burden in the aortic root. These results suggest that macrophage function and atherosclerosis susceptibility differ fundamentally in these two animal models with disturbed TG catabolism, showing a more severe phenotype by ATGL deficiency.

Lipolysis meets inflammation: arachidonic acid mobilization from fat

Lipolysis meets inflammation: arachidonic acid mobilization from fat

IL-6 and epinephrine have divergent fiber type effects on intramuscular lipolysis

IL-6 is an exercise-regulated myokine that has been suggested to increase lipolysis in fast-twitch skeletal muscle. However, it is not known if a similar effect is present in slow-twitch muscle. Furthermore, epinephrine increases IL-6 secretion from skeletal muscle, suggesting that IL-6 could play a role in mediating the lipolytic effects of catecholamines. The purpose of this study was to determine whether IL-6 stimulates skeletal muscle lipolysis in a fiber type dependent manner and is required for epinephrine-stimulated lipolysis in murine skeletal muscle. Soleus and extensor digitorum longus (EDL) muscles from male C57BL/6J wild-type and IL-6(-/-) mice were incubated with 1 μM (183 ng/ml) epinephrine or 75 ng/ml recombinant IL-6 (rIL-6) for 60 min. IL-6 treatment increased 5'-AMP-activated protein kinase and signal transducer and activator of transcription 3 phosphorylation and glycerol release in isolated EDL but not soleus muscles from C57BL/6J mice. Conversely, epinephrine increased glycerol release in soleus but not EDL muscles from C57BL/6J mice. Basal lipolysis was elevated in soleus muscle from IL-6(-/-) mice, and this was associated with increases in adipose triglyceride lipase (ATGL) and its coactivator comparative gene identification-58 (CGI-58). The increase in ATGL content does not appear to be due to a loss of IL-6's direct effects, because ex vivo treatment with IL-6 failed to alter the expression of ATGL mRNA in soleus muscle. In summary, IL-6 stimulates lipolysis in glycolytic but not oxidative muscle, whereas the opposite fiber type effect is seen with epinephrine. The absence of IL-6 indirectly upregulates lipolysis, and this is associated with increases in ATGL and its coactivator CGI-58.

Cardiac-specific overexpression of perilipin 5 provokes severe cardiac steatosis via the formation of a lipolytic barrier

Cardiac triacylglycerol (TG) catabolism critically depends on the TG hydrolytic activity of adipose triglyceride lipase (ATGL). Perilipin 5 (Plin5) is expressed in cardiac muscle (CM) and has been shown to interact with ATGL and its coactivator comparative gene identification-58 (CGI-58). Furthermore, ectopic Plin5 expression increases cellular TG content and Plin5-deficient mice exhibit reduced cardiac TG levels. In this study we show that mice with cardiac muscle-specific overexpression of perilipin 5 (CM-Plin5) massively accumulate TG in CM, which is accompanied by moderately reduced fatty acid (FA) oxidizing gene expression levels. Cardiac lipid droplet (LD) preparations from CM of CM-Plin5 mice showed reduced ATGL- and hormone-sensitive lipase-mediated TG mobilization implying that Plin5 overexpression restricts cardiac lipolysis via the formation of a lipolytic barrier. To test this hypothesis, we analyzed TG hydrolytic activities in preparations of Plin5-, ATGL-, and CGI-58-transfected cells. In vitro ATGL-mediated TG hydrolysis of an artificial micellar TG substrate was not inhibited by the presence of Plin5, whereas Plin5-coated LDs were resistant toward ATGL-mediated TG catabolism. These findings strongly suggest that Plin5 functions as a lipolytic barrier to protect the cardiac TG pool from uncontrolled TG mobilization and the excessive release of free FAs.

Skeletal muscle PLIN proteins, ATGL and CGI-58, interactions at rest and following stimulated contraction

Evidence indicates that skeletal muscle lipid droplet-associated proteins (PLINs) regulate lipolysis through protein-protein interactions on the lipid droplet surface. In adipocytes, PLIN1 is thought to regulate lipolysis by directly interacting with comparative gene identification-58 (CGI-58), an activator of adipose triglyceride lipase (ATGL). Upon lipolytic stimulation, PLIN1 is phosphorylated, releasing CGI-58 to fully activate ATGL and initiate triglyceride breakdown. The absence of PLIN1 in skeletal muscle leads us to believe that other PLIN family members undertake this role. Our purpose was to examine interactions between PLIN2, PLIN3, and PLIN5, with ATGL and its coactivator CGI-58 at rest and following contraction. Isolated rat solei were incubated for 30 min at rest or during 30 min of intermittent tetanic stimulation [150-ms volleys at 60 Hz with a train rate of 20 tetani/min (25°C)] to maximally stimulate intramuscular lipid breakdown. Results show that the interaction between ATGL and CGI-58 increased 128% following contraction (P = 0.041). Further, ATGL interacts with PLIN2, PLIN3, and PLIN5 at rest and following contraction. The PLIN2-ATGL interaction decreased significantly by 21% following stimulation (P = 0.013). Both PLIN3 and PLIN5 coprecipitated with CGI-58 at rest and following contraction, while there was no detectable interaction between PLIN2 and CGI-58 in either condition. Therefore, our findings indicate that in skeletal muscle, during contraction-induced muscle lipolysis, ATGL and CGI-58 strongly associate and that the PLIN proteins work together to regulate lipolysis, in part, by preventing ATGL and CGI-58 interactions at rest.

Distinct Mechanisms Regulate ATGL-Mediated Adipocyte Lipolysis by Lipid Droplet Coat Proteins

Adipose triglyceride lipase (ATGL) is the key triacylglycerol hydrolase in adipocytes. The precise mechanisms by which ATGL action is regulated by lipid droplet (LD) coat proteins and responds to hormonal stimulation are incompletely defined. By combining usage of loss- and gain-of-function approaches, we sought to determine the respective roles of perilipin 1 and fat-specific protein 27 (FSP27) in the control of ATGL-mediated lipolysis in adipocytes. Knockdown of endogenous perilipin 1 expression resulted in elevated basal lipolysis that was less responsive to β-adrenergic agonist isoproterenol. In comparison, depletion of FSP27 protein increased both basal and stimulated lipolysis with no significant impact on the overall response of cells to isoproterenol. In vitro assays showed that perilipin but not FSP27 was able to inhibit the triacylglycerol hydrolase activity of ATGL. Perilipin 1 also attenuated dose-dependent activation of ATGL by its Coactivator Comparative Gene identification-58. Accordingly, depletion of perilipin 1 and CGI-58 in adipocytes inversely affected basal lipolysis specifically mediated by overexpressed ATGL. Moreover, although depletion of perilipin 1 abolished the LD translocation of ATGL stimulated by isoproterenol, absence of FSP27 resulted in multilocularization of LDs along with increased LD presence of ATGL under both basal and stimulated conditions. Interestingly, knockdown of ATGL expression increased LD size and decreased LD number in FSP27-depeleted cells. Together, our results demonstrate that although FSP27 acts to constitutively limit the LD presence of ATGL, perilipin 1 plays an essential role in mediating the response of ATGL action to β-adrenergic hormones.

B56α/Protein Phosphatase 2A Inhibits Adipose Lipolysis in High-Fat Diet-Induced Obese Mice

Lipolysis and lipogenesis are two opposite processes that control lipid storage in adipocytes. Impaired adipose lipolysis has been observed in both obese human subjects and animal models. This study investigated the mechanisms underlying impaired adipose lipolysis in a high-fat diet-induced obese (DIO) mouse model. DIO models were created using male C57BL/6 mice. Our results show that beta3 adrenergic receptor-specific agonist BRL37344 induced adipose lipolysis was significantly blunted in DIO mice. The levels of Ser660 phosphorylation of hormone-sensitive lipase (HSL) were significantly decreased in the epididymal fat of DIO mice. However, protein levels of HSL, adipose triglyceride lipase and its coactivator comparative gene identification-58 were similar between DIO and control mice. It is known that upon lipolytic hormone stimulation, protein kinase A phosphorylates HSL Ser660 and activates HSL, whereas protein phosphatase 2A (PP2A) dephosphorylates and inactivates HSL. Interestingly, our study shows that high-fat feeding did not alter epididymal fat cAMP and protein kinase A protein levels but significantly increased the expression of the alpha-isoform of PP2A regulatory subunit B' (B56alpha). To study the role of B56alpha in obesity-associated lipolytic defect, B56alpha was overexpressed or knocked down by adenovirus-mediated gene transduction in cultured 3T3-L1CARDelta1 adipocytes. Overexpression of B56alpha significantly decreased HSL Ser660 phosphorylation. In contrast, knocking down B56alpha increased hormone-stimulated HSL activation and lipolysis in mature 3T3-L1CARDelta1 adipocytes. These results strongly suggest that elevated B56alpha/PP2A inhibits HSL and lipolysis in white adipose tissue of DIO mice.