Hormone-sensitive lipase
2002; Elsevier BV; Volume: 43; Issue: 10 Linguagem: Inglês
10.1194/jlr.r200009-jlr200
ISSN1539-7262
AutoresFredric B. Kraemer, Wen‐Jun Shen,
Tópico(s)Muscle metabolism and nutrition
ResumoHormone-sensitive lipase (HSL) is an intracellular neutral lipase that is capable of hydrolyzing triacylglycerols, diacylglycerols, monoacylglycerols, and cholesteryl esters, as well as other lipid and water soluble substrates. HSL activity is regulated post-translationally by phosphorylation and also by pretranslational mechanisms. The enzyme is highly expressed in adipose tissue and steroidogenic tissues, with lower amounts expressed in cardiac and skeletal muscle, macrophages, and islets. Studies of the structure of HSL have identified several amino acids and regions of the molecule that are critical for enzymatic activity and regulation of HSL. This has led to important insights into its function, including the interaction of HSL with other intracellular proteins, such as adipocyte lipid binding protein. Accumulating evidence has defined important functions for HSL in normal physiology, affecting adipocyte lipolysis, steroidogenesis, spermatogenesis, and perhaps insulin secretion and insulin action; however, direct links between abnormal expression or genetic variations of HSL and human disorders, such as obesity, insulin resistance, type 2 diabetes, and hyperlipidemia, await further clarification.The published reports examining the regulation, and function of HSL in normal physiology and disease are reviewed in this paper. Hormone-sensitive lipase (HSL) is an intracellular neutral lipase that is capable of hydrolyzing triacylglycerols, diacylglycerols, monoacylglycerols, and cholesteryl esters, as well as other lipid and water soluble substrates. HSL activity is regulated post-translationally by phosphorylation and also by pretranslational mechanisms. The enzyme is highly expressed in adipose tissue and steroidogenic tissues, with lower amounts expressed in cardiac and skeletal muscle, macrophages, and islets. Studies of the structure of HSL have identified several amino acids and regions of the molecule that are critical for enzymatic activity and regulation of HSL. This has led to important insights into its function, including the interaction of HSL with other intracellular proteins, such as adipocyte lipid binding protein. Accumulating evidence has defined important functions for HSL in normal physiology, affecting adipocyte lipolysis, steroidogenesis, spermatogenesis, and perhaps insulin secretion and insulin action; however, direct links between abnormal expression or genetic variations of HSL and human disorders, such as obesity, insulin resistance, type 2 diabetes, and hyperlipidemia, await further clarification. The published reports examining the regulation, and function of HSL in normal physiology and disease are reviewed in this paper. Hormone-sensitive lipase (HSL) activity was first identified as an epinephrine-sensitive lipolytic activity in adipose tissue. Its name was coined to reflect the ability of hormones such as catecholamines, ACTH, and glucagon to stimulate the activity of this intracellular neutral lipase (1Vaughan M. Berger J.E. Steinberg D. Hormone-sensitive lipase and monoglycerol lipase activities in adipose tissue.J. Biol. Chem. 1964; 239: 401-409Google Scholar). Hormonal activation of HSL occurs via cyclic AMP dependent protein kinase (PKA), which phosphorylates HSL (2Yeaman S.J. Hormone-sensitive lipase - a multipurpose enzyme in lipid metabolism.Biochim. Biophys. Acta. 1990; 1052: 128-132Google Scholar). As the enzyme responsible for the release of free fatty acids (FFA) from adipose tissue, HSL is felt to play a pivotal role in providing the major source of energy for most tissues. Although its expression is highest in adipose tissue, HSL is also expressed in adrenal, ovary, testis, and to a lesser extent in skeletal and cardiac muscle and macrophages (3Holm C. Kirchgessner T.G. Svenson K.L. Fredrikson G. Nilsson S. Miller C.G. Shively J.E. Heinzmann C. Sparkes R.S. Mohandas T. Lusis A.J. Belfrage P. Schotz M.C. Hormone-sensitive lipase: sequence, expression, and chromosomal localization to 19 cent-q13.3.Science. 1988; 241: 1503-1506Google Scholar, 4Kraemer F.B. Patel S. Saedi M.S. Sztalryd C. Detection of hormone-sensitive lipase in various tissues. I. Expression of an HSL/bacterial fusion protein and generation of anti-HSL antibodies.J. Lipid Res. 1993; 34: 663-671Google Scholar). Following the purification of the enzyme and the cloning of the cDNA encoding HSL, many research efforts have focused on understanding the activity, regulation, expression and function of this protein. The HSL gene is located on chromosome 19q13.3 (3Holm C. Kirchgessner T.G. Svenson K.L. Fredrikson G. Nilsson S. Miller C.G. Shively J.E. Heinzmann C. Sparkes R.S. Mohandas T. Lusis A.J. Belfrage P. Schotz M.C. Hormone-sensitive lipase: sequence, expression, and chromosomal localization to 19 cent-q13.3.Science. 1988; 241: 1503-1506Google Scholar) and was initially described to contain 9 exons spanning approximately 11 and 10 kB in human (5Langin D. Laurell H. Holst L.S. Belfrage P. Holm C. Gene organization and primary structure of human hormone-sensitive lipase: possible significance of a sequence homology with a lipase of Moraxella TA144, an antarctic bacterium.Proc. Natl. Acad. Sci. USA. 1993; 90: 4897-4901Google Scholar) and mouse (6Li Z. Sumida M. Birchbauer A. Schotz M.C. Reue K. Isolation and characterization of the gene for mouse hormone-sensitive lipase.Genomics. 1994; 24: 259-265Google Scholar), respectively, that encode an mRNA of ∼2.8 kB (7Grober J. Laurell H. Blaise R. Fabry B. Schaak S. Holm C. Langin D. Characterization of the promoter of human adipocyte hormone-sensitive lipase.Biochem. J. 1997; 328: 453-461Google Scholar). Subsequently, two additional exons (termed A and B) that differentially encode 170 and 70 nt 5′ untranslated regions were identified approximately 12.5 and 1.5 kB upstream of exon 1, respectively (7Grober J. Laurell H. Blaise R. Fabry B. Schaak S. Holm C. Langin D. Characterization of the promoter of human adipocyte hormone-sensitive lipase.Biochem. J. 1997; 328: 453-461Google Scholar). Only the smaller HSL mRNA product is expressed in human adipose tissue. In contrast, five different exons have been reported within 7 kB of the translation start site of exon 1 in mouse HSL, each of which can be alternatively utilized and expressed in mouse adipose tissue to varying degrees (8Laurin N.N. Wang S.P. Mitchell G.A. The hormone-sensitive lipase gene is transcibed from at least five alternative first exons in mouse adipose tissue.Mamm. Genome. 2000; 11: 972-978Google Scholar). In addition to these alternative exons encoding 5′ untranslated regions, several isoforms of HSL have been reported. A testis specific exon 15.5 kB upstream of exon 1 of human adipocyte HSL yields a 3.9 kB testicular HSL mRNA and encodes a larger protein (9Stenson Holst L. Langin D. Mulder H. Laurell H. Grober J. Bergh A. Mohrenweiser H.W. Edgren G. Holm C. Molecular cloning, genomic organization, and expression of a testicular isoform of hormone-sensitive lipase.Genomics. 1996; 35: 441-447Google Scholar). A second testis specific exon was identified ∼12 kB upstream of exon 1 and encodes a protein identical to adipocyte HSL (10Mairal A. Melaine N. Laurell H. Grober J. Holst L. Guillaudeux T. Holm C. Jegou B. Langin D. Characterization of a novel testicular form of human hormone-sensitive lipase.Biochem. Biophys. Res. Commun. 2002; 291: 286-290Google Scholar). β cells may have a specific exon (11Mulder H. Holst L. Svensson H. Degerman E. Sundler F. Ahren B. Rorsman P. Holm C. Hormone-sensitive lipase, the rate-limiting enzyme in triglyceride hydrolysis, is expressed and active in beta-cells.Diabetes. 1999; 48: 228-232Google Scholar) or an alternate translation start site may be 7 kB upstream of exon 1 (8Laurin N.N. Wang S.P. Mitchell G.A. The hormone-sensitive lipase gene is transcibed from at least five alternative first exons in mouse adipose tissue.Mamm. Genome. 2000; 11: 972-978Google Scholar). The purified rat enzyme has a molecular weight of approximately 84,000 Da on SDS-PAGE, corresponding to the 768 amino acid protein with a molecular size of 82,820 Da predicted from the primary translation product of rat HSL cDNA (3Holm C. Kirchgessner T.G. Svenson K.L. Fredrikson G. Nilsson S. Miller C.G. Shively J.E. Heinzmann C. Sparkes R.S. Mohandas T. Lusis A.J. Belfrage P. Schotz M.C. Hormone-sensitive lipase: sequence, expression, and chromosomal localization to 19 cent-q13.3.Science. 1988; 241: 1503-1506Google Scholar). The human HSL cDNA encodes a 775 amino acid protein with a molecular size of 84,032 Da, which corresponds to an 88 kDa immunoreactive protein seen on SDS-PAGE (5Langin D. Laurell H. Holst L.S. Belfrage P. Holm C. Gene organization and primary structure of human hormone-sensitive lipase: possible significance of a sequence homology with a lipase of Moraxella TA144, an antarctic bacterium.Proc. Natl. Acad. Sci. USA. 1993; 90: 4897-4901Google Scholar). However, a truncated, catalytically inactive form of HSL due to alternative splicing which eliminates exon 6 has been described in human, but not rat, adipose tissue (12Laurell H. Grober J. Vindis C. Lacome T. Dauzats M. Holm C. Langin D. Species-specific alternative splicing generates a catalytically inactive form of human hormone-sensitive lipase.Biochem. J. 1997; 328: 137-143Google Scholar). At least three additional isoforms of HSL have been reported. The testis appears to express two isoforms, one encoded by the larger testicular mRNA produces a protein with an additional 300 (rat) or 301 (human) amino acids N-terminal to the normal adipose form (9Stenson Holst L. Langin D. Mulder H. Laurell H. Grober J. Bergh A. Mohrenweiser H.W. Edgren G. Holm C. Molecular cloning, genomic organization, and expression of a testicular isoform of hormone-sensitive lipase.Genomics. 1996; 35: 441-447Google Scholar). So in addition to an 84 kDa protein that is similar to adipose HSL, a second larger isoform of ∼120–130 kDa is encoded by a unique testis mRNA (10Mairal A. Melaine N. Laurell H. Grober J. Holst L. Guillaudeux T. Holm C. Jegou B. Langin D. Characterization of a novel testicular form of human hormone-sensitive lipase.Biochem. Biophys. Res. Commun. 2002; 291: 286-290Google Scholar). Islets and β cells have an HSL isoform that contains an additional 43 amino acids N-terminal to the normal adipose form (11Mulder H. Holst L. Svensson H. Degerman E. Sundler F. Ahren B. Rorsman P. Holm C. Hormone-sensitive lipase, the rate-limiting enzyme in triglyceride hydrolysis, is expressed and active in beta-cells.Diabetes. 1999; 48: 228-232Google Scholar). HSL exists as a functional dimer composed of homologous subunits; dimeric HSL has greater hydrolytic activity when compared with monomeric HSL but no difference in substrate affinity (13Shen W-J. Patel S. Kraemer F.B. Hormone-sensitive lipase functions as an oligomer.Biochemistry. 2000; 39: 2392-2398Google Scholar). The primary sequence of HSL is unrelated to any of the other known mammalian lipases; however, it shares some sequence homology with lipase 2 of an antarctic bacterium, Moraxella TA144 (5Langin D. Laurell H. Holst L.S. Belfrage P. Holm C. Gene organization and primary structure of human hormone-sensitive lipase: possible significance of a sequence homology with a lipase of Moraxella TA144, an antarctic bacterium.Proc. Natl. Acad. Sci. USA. 1993; 90: 4897-4901Google Scholar). This homology aided in locating a G-X-S-X-G motif, which represents a consensus lipid binding sequence that contains the active site serine in other lipases, such as pancreatic lipase. This region was proposed (14Hemilä H. Koivula T.T. Palva I. Hormone-sensitive lipase is closely related to several bacterial proteins, and distantly related to acetylcholinesterase and lipoprotein lipase: identification of a superfamily of esterases and lipases.Biochim. Biophys. Acta. 1994; 1210: 249-253Google Scholar), and later shown by site-directed mutagenesis (15Holm C. Davis R.C. Østerlund T. Schotz M.C. Fredrikson G. Identification of the active site serine of hormone-sensitive lipase by site-directed mutagenesis.FEBS Lett. 1994; 344: 234-238Google Scholar), to contain the catalytically active serine at position 423 in rat HSL. Limited proteolysis and denaturation studies (16Østerlund T. Danielsson B. Degerman E. Contreras J.A. Edgren G. Davis R.C. Schotz M.C. Holm C. Domain-structure analysis of recombinant rat hormone-sensitive lipase.Biochem. J. 1996; 319: 411-420Google Scholar) suggested that HSL, like other lipases (17Wang H. Schotz M.C. The lipase gene family.J. Lipid Res. 2002; 43: 993-999Google Scholar) con-tains two major domains (Fig. 1). The N-terminal 320 amino acid domain is encoded by exons 1–4 and has no primary or secondary structural similarity with known proteins; however, the N-terminal domain has been shown to interact with adipocyte lipid-binding protein (ALBP) and has been proposed to function as a docking domain for protein-protein interactions (18Shen W-J. Sridhar K. Bernlohr D.A. Kraemer F.B. Interaction of rat hormone-sensitive lipase with adipocyte lipid-binding protein.Proc. Natl. Acad. Sci. USA. 1999; 96: 5528-5532Google Scholar). The C-terminal portion of HSL is similar to acetylcholinesterase, bile salt-stimulated lipase and several fungal lipases, and is composed of α/β-hydrolase folds that accommodate the catalytic site. A significant advance in understanding the structure of HSL was made when it was observed that, even in the absence of primary sequence homology, the organization of the secondary structure predicted for the C-terminal ∼450 amino acids of HSL was similar to the secondary structure of acetylcholinesterase and of two fungal lipases from Geotrichum candidum and Candida rugosa (19Contreras J.A. Karlsson M. Østerlund T. Laurell H. Svensson A. Holm C. Hormone-sensitive lipase is structurally related to acetylcholinesterase, bile salt-stimulated lipase, and several fungal lipases: building of a three-dimensional model for the catalytic domain of hormone-sensitive lipase.J. Biol. Chem. 1996; 271: 31426-31430Google Scholar). Using molecular modeling, it was proposed, and later confirmed by site-directed mutagenesis, that Ser-423, Asp-703, and His-733 (numbered for rat HSL) constitute the catalytic triad for HSL and are found within this C-terminal portion (20Østerlund T. Contreras J.A. Holm C. Identification of essential aspartic acid and histidine residues of hormone-sensitive lipase: apparent residues of the catalytic triad.FEBS Lett. 1997; 403: 259-262Google Scholar). Also located within the C-terminal portion is a 150 amino acid stretch that is not predicted to be composed of α helices or β sheets, but contains known phosphorylation sites and has been termed the regulatory module. HSL has broad substrate specificity; in addition to triacylglycerol, HSL can also catalyze the hydrolysis of diacylglycerol, 1(3) monoacylglycerol, cholesteryl esters, lipoidal esters of steroid hormones, and retinyl esters in adipose tissue, as well as water-soluble butyrate substrates (21Fredrikson G. Stralfors P. Nilsson N.O. Belfrage P. Hormone-sensitive lipase of rat adipose tissue: purification and some properties.J. Biol. Chem. 1981; 256: 6311-6320Google Scholar, 22Cook K.G. Yeaman S.J. Stralfors P. Fredrikson G. Belfrage P. Direct evidence that cholesteryl ester hydrolase from adrenal cortex is the same enzyme as hormone-sensitive lipase from adipose tissue.Eur. J. Biochem. 1982; 125: 245-249Google Scholar, 23Lee F.T. Adams J.B. Garton A.J. Yeaman S.J. Hormone-sensitive lipase is involved in the hydrolysis of lipoidal derivatives of estrogens and other steroid hormones.Biochim. Biophys. Acta. 1988; 963: 258-264Google Scholar, 24Wei S. Lai K. Patel S. Piantedosi R. Shen H. Colantuoni V. Kraemer F.B. Blaner W.S. Retinyl ester hydrolysis and retinol efflux from BFC-1β adipocytes.J. Biol. Chem. 1997; 272: 14159-14165Google Scholar); however, in contrast to many other lipases, HSL has no phospholipase activity. The activity against diacylglycerol is about 10-fold and 5-fold higher than the activity against triacylglycerol and monoacylglycerol, respectively, whereas the activity against cholesteryl esters is about twice the activity toward triacylglycerol. The esterase activity against water-soluble substrates is more than 20-fold that of triacylglycerols. HSL shows a preference for the sn 1- or 3-ester bond over the sn 2-ester bond as its substrate, with the relative activity against the sn 3-ester bond three to four times higher than the sn 2-ester bond. Although fatty acids appear to be more readily mobilized from adipose cells as their chain length shortens (between 12–24 carbons) and as their degree of unsaturation increases, examination of the ability of recombinant HSL to release individual fatty acids from triacylglycerol substrates in vitro does not support a large contribution of HSL to this selective mobilization (25Raclot T. Holm C. Langin D. Fatty acid specificity of hormone-sensitive lipase: implication in the selective hydrolysis of triacylglycerols.J. Lipid Res. 2001; 42: 2049-2057Google Scholar). Nonetheless, the relative hydrolysis of 12–24 carbon atom saturated fatty acids by HSL does increase with decreasing chain length (26Raclot T. Holm C. Langin D. A role for hormone-sensitive lipase in the selective mobilization of adipose tissue fatty acids.Biochim. Biophys. Acta. 2001; 1532: 88-96Google Scholar), and there is a tendency for a decrease in release as the number of unsaturated bonds increases, except for C20 fatty acids (25Raclot T. Holm C. Langin D. Fatty acid specificity of hormone-sensitive lipase: implication in the selective hydrolysis of triacylglycerols.J. Lipid Res. 2001; 42: 2049-2057Google Scholar). HSL may preferentially hydrolyze oxidized cholesteryl esters (at least 13-HODE cholesteryl ester) compared with cholesteryl linoleate (27Belkner J. Stender H. Holzhutter H. Holm C. Kuhn H. Macrophage cholesteryl ester hydrolases and hormone-sensitive lipase prefer specifically oxidized cholesteryl esters as substrates over their non-oxidized counterparts.Biochem. J. 2000; 352: 125-133Google Scholar). One of the unique features of HSL that differentiates it from most other lipases is that its activity against triacylglycerol and cholesteryl ester substrates appears to be regulated by reversible phosphorylation; however, hydrolytic activity against diacylglycerol, monoacylglycerol and water-soluble substrates is unaffected by phosphorylation (2Yeaman S.J. Hormone-sensitive lipase - a multipurpose enzyme in lipid metabolism.Biochim. Biophys. Acta. 1990; 1052: 128-132Google Scholar). PKA increases the hydrolytic activity of HSL by phosphorylation of a single site that was initially identified as S563 in rat HSL (2Yeaman S.J. Hormone-sensitive lipase - a multipurpose enzyme in lipid metabolism.Biochim. Biophys. Acta. 1990; 1052: 128-132Google Scholar) and is located within the regulatory module (Fig. 1). Although evidence to support the phosphorylation of S563 by PKA has been provided from mutagenesis experiments (28Shen W-J. Patel S. Natu V. Kraemer F.B. Mutational analysis of structural features of rat hormone-sensitive lipase.Biochemistry. 1998; 37: 8973-8979Google Scholar), other investigators have reported that S659 and S660 were phosphorylated by PKA in vitro and were required for the phosphorylation-induced increase in hydrolytic activity against triacylglycerol substrate (29Anthonsen M.W. Rönnstrandt L. Wernstedt C. Degerman E. Holm C. Identification of novel phosphorylation sites in hormone-sensitive lipase that are phosphorylated in response to isoproterenol and govern activation properties in vitro.J. Biol. Chem. 1998; 273: 215-221Google Scholar). Additionally, lipolytic hormones not only can activate PKA, but also the mitogen activated protein kinase pathway and extracellular signal-regulated kinase (ERK). Activation of the ERK pathway appears to be able to regulate adipocyte lipolysis by phosphorylating HSL on S600 and increasing the activity of HSL (30Greenberg A.S. Shen W-J. Muliro K. Patel S. Souza S.C. Roth R.A. Kraemer F.B. Stimulation of lipolysis and hormone-sensitive lipase via the extracellular signal-regulated kinase pathway.J. Biol. Chem. 2001; 276: 45456-45461Google Scholar). In contrast to activation of activity seen with PKA or ERK phosphorylation, other kinases such as glycogen synthase kinase-4, Ca++/calmodulin-dependent protein kinase II, and AMP-activated protein kinase phosphorylate HSL at a secondary basal site S565 in rat HSL (2Yeaman S.J. Hormone-sensitive lipase - a multipurpose enzyme in lipid metabolism.Biochim. Biophys. Acta. 1990; 1052: 128-132Google Scholar). Phosphorylation at S565 impairs the phosphorylation of S563 by PKA (2Yeaman S.J. Hormone-sensitive lipase - a multipurpose enzyme in lipid metabolism.Biochim. Biophys. Acta. 1990; 1052: 128-132Google Scholar). HSL activity can be inactivated by protein phosphatases. The most active phosphatases against S563 are phosphatase 2A and 2C, while S565 is predominately dephosphorylated by phosphatase 2A (31Wood S.L. Emmison N. Borthwick A.C. Yeaman S.J. The protein phosphatases responsible for dephosphorylation of hormone-sensitive lipase in isolated rat adipocytes.Biochem. J. 1993; 295: 531-535Google Scholar). Thus, several different kinases phosphorylate HSL at unique serines within the regulatory module and modulate HSL activity. The primary action attributed to HSL is hydrolysis of stored triacylglycerols in adipose tissue, i.e., lipolysis. The control of lipolysis is complex and involves multiple mechanisms (32Londos C. Brasaemle D.L. Schultz C.J. Adler-Wailes D.C. Levin D.M. Kimmel A.R. Rondinone C.M. On the control of lipolysis in adipocytes.Ann. N. Y. Acad. Sci. 1999; 892: 155-168Scopus (220) Google Scholar, 33Holm C. Osterlund T. Laurell H. Contreras J.A. Molecular mechanisms regulating hormone-sensitive lipase and lipolysis.Annu. Rev. Nutr. 2000; 20: 365-393Google Scholar). These include lipolytic (β-adrenergic agonists, ACTH, etc.) and anti-lipolytic (insulin, adenosine, etc.) hormones, their cognate receptors and signaling pathways, lipid droplet-associated proteins such as perilipins, as well as HSL or other as yet unidentified lipases. In addition to the activation of HSL hydrolytic activity, other mechanisms involving HSL have been suggested to account for lipolysis. Evidence has been provided using subcellular fractionation to show that catecholamine-induced stimulation of lipolysis in vitro in 3T3-L1 adipocytes or in rat adipose cells is due to the translocation of phosphorylated HSL from an aqueous cytosolic compartment to the lipid droplet (34Egan J.J. Greenberg A.S. Chang M-K. Wek S.A. Moos Jr., M.C. Londos C. Mechanism of hormone-stimulated lipolysis in adipocytes: translocation of hormone-sensitive lipase to the lipid storage droplet.Proc. Natl. Acad. Sci. USA. 1992; 89: 8537-8541Google Scholar, 35Clifford G.M. Londos C. Kraemer F.B. Vernon R.G. Yeaman S.J. Translocation of hormone-sensitive lipase and perilipin upon lipolytic stimulation of rat adipocytes.J. Biol. Chem. 2000; 275: 5011-5015Google Scholar). Using immunofluorescence microscopy, HSL was observed diffusely distributed throughout the cytosol of 3T3-L1 adipocytes and, upon catecholamine stimulation, HSL translocated from the cytosol to the surfaces of intracellular lipid droplets concomitant with the onset of lipolysis (36Brasaemle D.L. Levin D.M. Adler-Wailes D.C. Londos C. The lipolytic stimulation of 3T3–L1 adipocytes promotes the translocation of hormone-sensitive lipase to the surfaces of lipid storage droplets.Biochim. Biophys. Acta. 2000; 1483: 251-262Google Scholar). It has been suggested that translocation of HSL to the lipid droplet is the critical event in regulating lipolysis induced by a variety of lipolytic agents, such as isoproterenol, forskolin, cyclic AMP, theophylline, and okadaic acid (37Morimoto C. Kiyama A. Kameda K. Ninomiya H. Tsujita T. Okuda H. Mechanism of the stimulatory action of okadaic acid on lipolysis in rat fat cells.J. Lipid Res. 2000; 41: 199-204Google Scholar, 38Morimoto C. Kameda K. Tsujita T. Okuda H. Relationships between lipolysis induced by various lipolytic agents and hormone-sensitive lipase in rat fat cells.J. Lipid Res. 2001; 42: 120-127Google Scholar); however, this is not true for all physiological conditions, since translocation of HSL was not observed with lipolytic stimulation in adipocytes from old (35Clifford G.M. Londos C. Kraemer F.B. Vernon R.G. Yeaman S.J. Translocation of hormone-sensitive lipase and perilipin upon lipolytic stimulation of rat adipocytes.J. Biol. Chem. 2000; 275: 5011-5015Google Scholar) or lactating rats (39Clifford G.M. Kraemer F.B. Yeaman S.J. Vernon R.G. Translocation of hormone-sensitive lipase and perilipin upon lipolytic stiumlation during the lactation cycle of the rat.Metabolism. 2001; 50: 1264-1269Google Scholar). Although the mechanisms mediating the translocation of HSL have not been well studied, disruption of microtubules or microfilaments appears to have minimal effects on isoproterenol-stimulated glycerol release and no visible effects on the translocation of HSL determined by immunofluorescence light microscopy (36Brasaemle D.L. Levin D.M. Adler-Wailes D.C. Londos C. The lipolytic stimulation of 3T3–L1 adipocytes promotes the translocation of hormone-sensitive lipase to the surfaces of lipid storage droplets.Biochim. Biophys. Acta. 2000; 1483: 251-262Google Scholar). However, a protein (lipotransin) was identified in a yeast two-hybrid screen that interacts with HSL and was proposed to be a potential participant in the process of the translocation of HSL to the lipid droplet (40Syu L-J. Saltiel A.R. Lipotransin, a novel docking protein for hormone-sensitive lipase.Mol. Cell. 1999; 4: 109-115Google Scholar). Lipotransin is homologous to p60 katanin and is a member of the AAA protein superfamily, possessing ATPase and microtubule severing activities (41Hartman J.J. Mahr J. McNally K. Okawa K. Iwamatsu A. Thomas S. Cheesman S. Heuser J. Vale R.D. McNally F.J. Katanin, a microtubule-severing protein, is a novel AAA ATPase that targets to the centrosome using a WD40-containing subunit.Cell. 1998; 93: 277-287Google Scholar). The function of lipotransin in interacting with HSL and influencing lipolysis has yet to be elucidated. A proposed model for hormone-induced lipolysis is depicted in Fig. 2. Fatty acids and monoacylglycerol exert product inhibition on HSL activity. This is interesting in light of the observation that HSL specifically interacts with ALBP, a member of the family of intracellular lipid-binding proteins which bind fatty acids and other hydrophobic ligands (18Shen W-J. Sridhar K. Bernlohr D.A. Kraemer F.B. Interaction of rat hormone-sensitive lipase with adipocyte lipid-binding protein.Proc. Natl. Acad. Sci. USA. 1999; 96: 5528-5532Google Scholar). Mutational analysis has identified several amino acids within the N-terminal domain of HSL (H194 and E199) as critical for mediating the interaction of HSL with ALBP (42Shen W-J. Liang Y. Hong R. Patel S. Natu V. Sridhar K. Jenkins A. Bernlohr D.A. Kraemer F.B. Characterization of the functional interaction of adipocyte lipid-binding protein with hormone-sensitive lipase.J. Biol. Chem. 2001; 276: 49443-49448Google Scholar). Incubation or co-expression of ALBP with HSL increased substrate hydrolysis, which was lost when the binding of ALBP to HSL was disrupted by mutagenesis. In addition, the ability of fatty acids to inhibit HSL hydrolytic activity was attenuated by co-incubation with ALBP. These observations suggest that ALBP and HSL constitute a lipolytic complex that increases the hydrolytic activity of HSL through the physical interaction of HSL with ALBP, and because ALBP sequesters fatty acids and prevents product inhibition. This is consistent with experiments in ALBP null mice where basal and isoproterenol-stimulated lipolysis are decreased ∼40% (43Coe N.R. Simpson M.A. Bernlohr D.A. Targeted disruption of the adipocyte lipid binding protein (aP2 protein) gene impairs fat cell lipolysis and increases cellular fatty acid levels.J. Lipid Res. 1999; 40: 967-972Google Scholar). Indirect evidence has suggested that HSL is the rate limiting enzyme in intracellular lipolysis; overexpression of HSL in 3T3-F442A cells prevents differentiated adipocytes from accumulating triglyceride (44Sztalryd C. Komaromy M.C. Kraemer F.B. Overexpression of hormone-sensitive lipase prevents triglyceride accumulation in adipocytes.J. Clin. Invest. 1995; 95: 2652-2661Google Scholar). Recently, the functional significance of HSL in adipose tissue metabolism has begun to be clarified in studies using HSL null mice (45Osuga J-i. Ishibashi S. Oka T. Yagyu H. Tozawa R. Fujimoto A. Shionoira F. Yahagi N. Kraemer F.B. Tsutsumi O. Yamada N. Targeted disruption of hormone-sensitive lipase results in male sterility and adipocyte hypertrophy, but not in obesity.Proc. Natl. Acad. Sci. USA. 2000; 97: 787-792Google Scholar, 46Wang S.P. Laurin N. Himms-Hagen J. Rudnicki M.A. Levy E. Robert M-F. Pan L. Oligny L. Mitchell G.A. The adipose tissue phenotype of hormone-sensitive lipase deficiency in mice.Obes. Res. 2001; 9: 119-128Google Scholar, 47Haemmerle G. Zimmermann R. Hayn M. Theussl C. Waeg G. Wagner E. Sattler W. Magin T.M. Wagner E.F. Zechner R. Hormone-sensitive lipase deficiency in mice causes diglyceride accumulation in adipose tissue, muscle, and testis.J. Biol. Chem. 2002; 277: 4806-4815Google Scholar). Inactivation of HSL by homologous recombination resulted in the complete absence of neutral cholesteryl ester hydrolase activity in adipose tissue (both white and brown); however, triacylglycerol lipase activity in white adipose tissue was reduced by only 40% and triacylglycerol lipase activity in brown adipose tissue was similar to wild-type mice (45Osuga J-i. Ishibashi S. Oka T. Yagyu H. Tozawa R. Fu
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