Robert A. Zimmermann

Adipose Triglyceride Lipase and Hormone-sensitive Lipase Are the Major Enzymes in Adipose Tissue Triacylglycerol Catabolism

The mobilization of free fatty acids from adipose triacylglycerol (TG) stores requires the activities of triacylglycerol lipases. In this study, we demonstrate that adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) are the major enzymes contributing to TG breakdown in in vitro assays and in organ cultures of murine white adipose tissue (WAT). To differentiate between ATGL- and HSL-specific activities in cytosolic preparations of WAT and to determine the relative contribution of these TG hydrolases to the lipolytic catabolism of fat, mutant mouse models lacking ATGL or HSL and a mono-specific, small molecule inhibitor for HSL (76-0079) were used. We show that 76-0079 had no effect on TG catabolism in HSL-deficient WAT but, in contrast, essentially abolished free fatty acid mobilization in ATGL-deficient fat. CGI-58, a recently identified coactivator of ATGL, stimulates TG hydrolase activity in wild-type and HSL-deficient WAT but not in ATGL-deficient WAT, suggesting that ATGL is the sole target for CGI-58-mediated activation of adipose lipolysis. Together, ATGL and HSL are responsible for more than 95% of the TG hydrolase activity present in murine WAT. Additional known or unknown lipases appear to play only a quantitatively minor role in fat cell lipolysis.

Lipolysis: pathway under construction.

Purpose of review The lipolytic catabolism of stored fat in adipose tissue supplies tissues with fatty acids as metabolites and energy substrates during times of food deprivation. This review focuses on the function of recently discovered enzymes in adipose tissue lipolysis and fatty acid mobilization. Recent findings The characterization of hormone-sensitive lipase-deficient mice provided compelling evidence that hormone-sensitive lipase is not uniquely responsible for the hydrolysis of triacylglycerols and diacylglycerols of stored fat. Recently, three different laboratories independently discovered a novel enzyme that also acts in this capacity. We named the enzyme ‘adipose triglyceride lipase’ in accordance with its predominant expression in adipose tissue, its high substrate specificity for triacylglycerols, and its function in the lipolytic mobilization of fatty acids. Two other research groups showed that adipose triglyceride lipase (named desnutrin and Ca-independent phospholipase A2ζ, respectively) is regulated by the nutritional status and that it might exert acyl-transacylase activity in addition to its activity as triacylglycerol hydrolase. Adipose triglyceride lipase represents a novel type of ‘patatin domain-containing’ triacylglycerol hydrolase that is more closely related to plant lipases than to other known mammalian metabolic triacylglycerol hydrolases. Summary Although the regulation of adipose triglyceride lipase and its physiological function remain to be determined in mouse lines that lack or overexpress the enzyme, present data permit the conclusion that adipose triglyceride lipase is involved in the cellular mobilization of fatty acids, and they require a revision of the concept that hormone-sensitive lipase is the only enzyme involved in the lipolysis of adipose tissue triglycerides.

Obese Yeast: Triglyceride Lipolysis Is Functionally Conserved from Mammals to Yeast

Storage and degradation of triglycerides are essential processes to ensure energy homeostasis and availability of precursors for membrane lipid synthesis. Recent evidence suggests that an emerging class of enzymes containing a conserved patatin domain are centrally important players in lipid degradation. Here we describe the identification and characterization of a major triglyceride lipase of the adipose triglyceride lipase/Brummer family, Tgl4, in the yeast Saccharomyces cerevisiae. Elimination of Tgl4 in a tgl3 background led to fat yeast, rendering growing cells unable to degrade triglycerides. Tgl4 and Tgl3 lipases localized to lipid droplets, independent of each other. Serine 315 in the GXSXG lipase active site consensus sequence of the patatin domain of Tgl4 is essential for catalytic activity. Mouse adipose triglyceride lipase (which also contains a patatin domain but is otherwise highly divergent in primary structure from any yeast protein) localized to lipid droplets when expressed in yeast, and significantly restored triglyceride breakdown in tgl4 mutants in vivo. Our data identify yeast Tgl4 as a functional ortholog of mammalian adipose triglyceride lipase.

Lipoprotein lipase: the regulation of tissue specific expression and its role in lipid and energy metabolism.

Purpose of review The aim of this review is to summarize and discuss recent advances in the understanding of the physiological role of lipoprotein lipase in lipid and energy metabolism. Recent findings Studies on the transcriptional and the posttranscriptional level of lipoprotein lipase expression have provided new insights into the complex mechanisms that are involved in the regulation of the enzyme. Additionally a large body of evidence from both human studies and animal models suggests that the level of lipoprotein lipase expression in a given tissue is the rate limiting process for the uptake of triglyceride derived fatty acids. Imbalances in the partitioning of fatty acids among peripheral tissues have major metabolic consequences. For example, in mice both decreased lipoprotein lipase activities in adipose tissue and increased activity in muscle are associated with resistance to obesity; lack of lipoprotein lipase activity in macrophages is correlated with a decreased susceptibility to develop atherosclerotic lesions and overexpression of the enzyme in muscle is associated with increased blood glucose levels and insulin resistance. Summary Considering the central role of lipoprotein lipase in energy metabolism it is a reasonable goal to discover and develop new drugs that affect the tissue specific expression pattern of the enzyme.

Neutral lipid storage disease: genetic disorders caused by mutations in adipose triglyceride lipase/PNPLA2 or CGI-58/ABHD5

Neutral lipid storage disease (NLSD) is a group of autosomal recessive disorders characterized by the excessive accumulation of neutral lipids in multiple tissues. Recently, two genes, adipose triglyceride lipase (ATGL/PNPLA2) and comparative gene identification-58 (CGI-58/ABHD5), have been shown to cause NLSD. ATGL specifically hydrolyzes the first fatty acid from triacylglycerols (TG) and CGI-58/ABHD5 stimulates ATGL activity by a currently unknown mechanism. Mutations in both the ATGL and the CGI-58 genes are associated with systemic TG accumulation, yet the resulting clinical manifestations are not identical. Patients with defective ATGL function suffer from more severe myopathy (NLSDM) than patients with defective CGI-58 function. On the other hand, CGI-58 mutations are always associated with ichthyosis (NLSDI), which was not observed in patients with defective ATGL function. These observations indicate an ATGL-independent function of CGI-58. This review summarizes recent findings with the goal of relating structural variants of ATGL and CGI-58 to functional consequences in lipid metabolism.

Growth Retardation, Impaired Triacylglycerol Catabolism, Hepatic Steatosis, and Lethal Skin Barrier Defect in Mice Lacking Comparative Gene Identification-58 (CGI-58)

Comparative gene identification-58 (CGI-58), also designated as α/β-hydrolase domain containing-5 (ABHD-5), is a lipid droplet-associated protein that activates adipose triglyceride lipase (ATGL) and acylates lysophosphatidic acid. Activation of ATGL initiates the hydrolytic catabolism of cellular triacylglycerol (TG) stores to glycerol and nonesterified fatty acids. Mutations in both ATGL and CGI-58 cause “neutral lipid storage disease” characterized by massive accumulation of TG in various tissues. The analysis of CGI-58-deficient (Cgi-58−/−) mice, presented in this study, reveals a dual function of CGI-58 in lipid metabolism. First, systemic TG accumulation and severe hepatic steatosis in newborn Cgi-58−/− mice establish a limiting role for CGI-58 in ATGL-mediated TG hydrolysis and supply of nonesterified fatty acids as energy substrate. Second, a severe skin permeability barrier defect uncovers an essential ATGL-independent role of CGI-58 in skin lipid metabolism. The neonatal lethal skin barrier defect is linked to an impaired hydrolysis of epidermal TG. As a consequence, sequestration of fatty acids in TG prevents the synthesis of acylceramides, which are essential lipid precursors for the formation of a functional skin permeability barrier. This mechanism may also underlie the pathogenesis of ichthyosis in neutral lipid storage disease patients lacking functional CGI-58.

The C-terminal Region of Human Adipose Triglyceride Lipase Affects Enzyme Activity and Lipid Droplet Binding

Adipose triglyceride lipase (ATGL) catalyzes the first step in the hydrolysis of triacylglycerol (TG) generating diacylglycerol and free fatty acids. The enzyme requires the activator protein CGI-58 (or ABHD5) for full enzymatic activity. Defective ATGL function causes a recessively inherited disorder named neutral lipid storage disease that is characterized by systemic TG accumulation and myopathy. In this study, we investigated the functional defects associated with mutations in the ATGL gene that cause neutral lipid storage disease. We show that these mutations lead to the expression of either inactive enzymes localizing to lipid droplets (LDs) or enzymatically active lipases with defective LD binding. Additionally, our studies assign important regulatory functions to the C-terminal part of ATGL. Truncated mutant ATGL variants lacking ∼220 amino acids of the C-terminal protein region do not localize to LDs. Interestingly, however, these mutants exhibit substantially increased TG hydrolase activity in vitro (up to 20-fold) compared with the wild-type enzyme, indicating that the C-terminal region suppresses enzyme activity. Protein-protein interaction studies revealed an increased binding of truncated ATGL to CGI-58, suggesting that the C-terminal part interferes with CGI-58 interaction and enzyme activation. Compared with the human enzyme, the C-terminal region of mouse ATGL is much less effective in suppressing enzyme activity, implicating species-dependent differences in enzyme regulation. Together, our results demonstrate that the C-terminal region of ATGL is essential for proper localization of the enzyme and suppresses enzyme activity.

Adipose triglyceride lipase in human skeletal muscle is upregulated by exercise training

Mobilization of fatty acids from stored triacylglycerol (TG) in adipose tissue and skeletal muscle [intramyocellular triacylglycerol (IMTG)] requires activity of lipases. Although exercise training increases the lipolytic capacity of skeletal muscle, the expression of hormone-sensitive lipase (HSL) is not changed. Recently, adipose triglyceride lipase (ATGL) was identified as a TG-specific lipase in various rodent tissues. To investigate whether human skeletal muscle ATGL protein is regulated by endurance exercise training, 10 healthy young men completed 8 wk of supervised endurance exercise training. Western blotting analysis on lysates of skeletal muscle biopsy samples revealed that exercise training induced a twofold increase in skeletal muscle ATGL protein content. In contrast to ATGL, expression of comparative gene identification 58 (CGI-58), the activating protein of ATGL, and HSL protein was not significantly changed after the training period. The IMTG concentration was significantly decreased by 28% at termination of the training program compared with before. HSL-phoshorylation at Ser(660) was increased, HSL-Ser(659) phosporylation was unchanged, and HSL-phoshorylation at Ser(565) was decreased altogether, indicating an enhanced basal activity of this lipase. No change was found in the expression of diacylglycerol acyl transferase 1 (DGAT1) after training. Inhibition of HSL with a monospecific, small molecule inhibitor (76-0079) and stimulation of ATGL with CGI-58 revealed that significant ATGL activity is present in human skeletal muscle. These results suggest that ATGL in addition to HSL may be important for human skeletal muscle lipolysis.

Structure of the L1 protuberance in the ribosome.

The L1 protuberance of the 50S ribosomal subunit is implicated in the release/disposal of deacylated tRNA from the E site. The apparent mobility of this ribosomal region has thus far prevented an accurate determination of its three-dimensional structure within either the 50S subunit or the 70S ribosome. Here we report the crystal structure at 2.65 Å resolution of ribosomal protein L1 from Sulfolobus acidocaldarius in complex with a specific 55-nucleotide fragment of 23S rRNA from Thermus thermophilus. This structure fills a major gap in current models of the 50S ribosomal subunit. The conformations of L1 and of the rRNA fragment differ dramatically from those within the crystallographic model of the T. thermophilus 70S ribosome. Incorporation of the L1–rRNA complex into the structural models of the T. thermophilus 70S ribosome and the Deinococcus radiodurans 50S subunit gives a reliable representation of most of the L1 protuberance within the ribosome.

Adipose triglyceride lipase plays a key role in the supply of the working muscle with fatty acids

FAs are mobilized from triglyceride (TG) stores during exercise to supply the working muscle with energy. Mice deficient for adipose triglyceride lipase (ATGL-ko) exhibit defective lipolysis and accumulate TG in adipose tissue and muscle, suggesting that ATGL deficiency affects energy availability and substrate utilization in working muscle. In this study, we investigated the effect of moderate treadmill exercise on blood energy metabolites and liver glycogen stores in mice lacking ATGL. Because ATGL-ko mice exhibit massive accumulation of TG in the heart and cardiomyopathy, we also investigated a mouse model lacking ATGL in all tissues except cardiac muscle (ATGL-ko/CM). In contrast to ATGL-ko mice, these mice did not accumulate TG in the heart and had normal life expectancy. Exercise experiments revealed that ATGL-ko and ATGL-ko/CM mice are unable to increase circulating FA levels during exercise. The reduced availability of FA for energy conversion led to rapid depletion of liver glycogen stores and hypoglycemia. Together, our studies suggest that ATGL-ko mice cannot adjust circulating FA levels to the increased energy requirements of the working muscle, resulting in an increased use of carbohydrates for energy conversion. Thus, ATGL activity is required for proper energy supply of the skeletal muscle during exercise.