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Adipocyte hormone-sensitive lipase: a major regulator of lipid metabolism

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Adipocyte hormone-sensitive lipase: a major regulator of lipid metabolism
  Proceedings ofthe Nutrition Society zyxwvut 1996), zyxwv 5, 93-109 93 z Adipocyte hormone-sensitive ipase: a major regulator of lipid metabolism BY DOMINIQUE LANGIN’, CECILIA HOLM2 AND MAX LAFONTANl Unite‘ INSERM 31 7, Institut Louis Bugnard, Faculte‘ de Mkdecine, Universite‘ Paul Sabatier, CHR Rangueil, BBt. L3, zyxwv 1054 Toulouse Cedex, France 2Section for Molecular Signalling, Department of Cell and Molecular Biology, Lund University, PO Box 94, 221 00 Lund, Sweden Lipase hormono-sensible de l’adipocyte: un rCgulateur important du mCtabolisme lipidique RESUMG Le tissu adipeux joue un r81e important dans le contrde de la balance CnergCtique. La mobilisation des triacylglycCrols par la lipase hormono-sensible EC; LHS) est soumis a un contrBle direct par les hormones et neurotransmetteurs qui modulent les concentrations intracellulaires d’AMP cyclique (AMPc). L’hydrolyse des triacylglycCrols par la LHS constitue l’etape limitante de la lipolyse. La LHS est phosphorylCe sur le site rkgulateur (Ser552 dans la LHS humaine) par la prothe kinase dependante de 1’AMPc EC lorsque les concentrations intra- cellulaires d’ AMPc augmentent. Cette phosphorylation conduit l’activation de l’enzyme. Une deuxikme site de phosphorylation (Ser5.54 dans la LHS humaine) dCnommC site basal est la cible de la protCine kinase activCe par I’AMP. La phosphorylation du site basal ne conduit pas l’activation de la LHS et empkche la phosphorylation sur le site regulateur. La phosphorylation et I’activation de la protCine kinase activke par 1’AMP constitue donc un mecanisme antilipolytique qui est fonctionnel sur cellules isolCes mais dont l’importance physiologique n’est pas connue. Les ADN complkmentaires de la LHS de plusieurs espkces ont CtC clones et la structure des gbnes de LHS de l’homme et de la souris sont connus. Differents domaines fonctionnels de la protCine ont CtC proposes. Une rCgion d’homologie de sequences en amont de la serine 424 du site catalytique avec cinq enzymes d’organismes procaryotes et une enzyme humaine a ete decrite. La localisation chromosomique du gbne de la LHS est connue chez l’homme (chromo- some 19, region q13-1+13.2), le porc et la souris. La mise en evidence de marqueurs polymorphiques dans le g&ne devrait permettre de tester l’hypothkse d’une implication de la LHS dans certaines maladies hereditaires du metabolisme lipidique. L’expression de la LHS varie selon la localisation anatomique du tissu adipeux chez le rat. Cette expression subit Cgalement des variations durant la gestation chez le rat et le cycle annuel chez les mammifbres hibernants. Chez l’homme, les taux d’ARN messagers de la LHS sont diminuCs dans le tissu adipeux de certains patients atteints de cancer. L‘activitC enzymatique totale est diminuee chez les patients atteints d’hyperlipidemie familiale combinCe mais pas chez les patients atteints du syndrome metabolique bien que, dans les deux cas, la lipolyse adipocytaire maximale soit diminuke. Les mkcanismes molCculaires de contr6le de l’expression de la LHS sont pratiquement inconnus. http:/ from http:/ IP address:, on 29 Sep 2016 at 12:37:32, subject to the Cambridge Core terms of use, available at  94 D. LANGIN AND OTHERS All animals feed and fast intermittently. During evolution it has been necessary to develop precisely-regulated mechanisms to control the storage and release of metabolic fuels. In mammals, short-term fluctuations in energy balance are to some extent buffered by the glycogen stores. However, the capacity to store glycogen is limited. Longer-term imbalances between energy intake and expenditure are translated into changes in the body’s store of lipid, mainly in the form of intracellular triacylglycerol (TAG) in adipose tissue. Thus, a highly developed adipose tissue is characteristic of all mammalian species. The vast majority of the body’s TAG (>95 ) is found in adipose-tissue stores. Lipolysis refers to the hydrolysis of TAG, via di- and monoacylglycerol intermediates, to fatty acids and glycerol. Adipose-tissue lipolysis is the major regulator of the supply of lipid energy because it controls the release of fatty acids into the plasma. The rate-limiting step of adipose-tissue lipolysis is the hydrolysis of TAG by hormone-sensitive lipase z EC; HSL). Adipose tissue HSL is thus one of the enzymes determining whole-body lipid fuel availability. In the post-absorptive state, HSL activity accounts for most of the detectable lipolysis (Frayn et al. 1995). The present review focuses mainly on recent advances in the understanding of HSL function and regulation in the adipocyte. Cellular aspects of lipid mobilization such as substrate selection have been recently reviewed (Lafontan zyx   Langin, 1995). MOLECULAR CONTROL OF HORMONE-SENSITIVE LIPASE ACTIVITY It is generally accepted that lipolysis is controlled mainly by sympathetic nervous system activity and plasma insulin levels. Basically, the lipolytic response of the fat cell depends on the balanced action of stimulatory and inhibitory pathways on HSL activity. The different steps of the lipolytic process leading to the activation of HSL are quite well defined (Fig. 1). The first cellular action of catecholamines and of a number of endocrine and/or paracrine regulators of lipolysis (e.g. adenosine and prostaglandins) is their binding to plasma membrane receptors. The stimulatory effect on lipolysis is strictly connected to the receptor-controlled increment of intracellular cAMP concentrations which in turn promotes activation of CAMP-dependent protein kinase EC; CAMP-PK; Honnor zyxwvut t al. 1985) which phosphorylates HSL. The first step leading to activation of the lipolytic cascade, involves the multi-regulated enzyme, adenylate cyclase EC, which produces CAMP. Detailed mechanistic considerations have been reviewed recently by Lafontan Berlan (1993). Cate- cholamines are the most sophisticated regulators of fat cell function since they operate through five separate adrenergic receptors. They are able to stimulate three subtypes of P-adrenoceptors which are positively coupled to adenylate cyclase by Gs proteins, and an a2-adrenoceptor negatively coupled to the enzyme by a Gi protein. An important point in the metabolic actions initiated by endocrine and paracrine regulators concerns the functional significance of intracellular cAMP elevations promoted by receptor-mediated adenylate cyclase control. In fat cells, the lipolytic agents promote cAMP increments which largely overcome the concentrations required for maximal activation of CAMP-PK and lipolysis (Fain Garcia-Sainz, 1983; Honnor et al. 1985). Biphasic regulation of lipolysis by catecholamines has been demonstrated clearly in human fat cells (Berlan Lafontan, 1985; Mauri2ge et al. 1987). The interplay between a2- and P-adrenoceptors plays a key role in the triggering of cAMP increments in fat cells; important species- specific differences exist. http:/ from http:/ IP address:, on 29 Sep 2016 at 12:37:32, subject to the Cambridge Core terms of use, available at  LIPID ABSORPTION AND METABOLISM 95 Noradrenaline Insulin drenaline Plasma membrane k q-Pzj33-ARs TAR zyxwv P-Tyr-P HI zyxwvu   dipoc yte Inactive zyxwv   zyx c ATP- CAMP zyxwvuts   t Active Inactive S Fat droplet Fig. 1. Adipose-tissue lipolysis showing catecholamine receptors and insulin receptor (IR), G proteins (Gs and Gi), cGMP-inhibited low zyxwvutsr ,,, cAMP phosphodiesterase EC; cGI-PDE) and the catalyst moieties of adenylate cyclase EC; AC). Three stimulatory P-adrenoceptors PI, p and P3-ARS), coupled to Gs, and one inhibitory a2-adrenoceptor (cuz-AR), oupled to Gi, exert antagonistic actions on AC activity, cAMP production, and CAMP-dependent protein kinase EC; CAMP-PK) activity. Insulin promotes the degradation of cAMP via phosphorylation and activation of cGI-PDE which is associated with intracellular membranes. Phosphorylation of hormone-sensitive lipase EC; HSL) by CAMP-PK s followed by HSL enzymic activation and lipolysis. Monoacylglycerol lipase zyxwv EC; MGL) hydrolyses the breakdown of monoacylglycerol into fatty acid and glycerol. Ser-PK, serine kinase. Insulin is the physiologically important anti-lipolytic hormone. Initially insulin was thought to cause dephosphorylation of HSL and its deactivation by an effect which could involve phosphatase activation. However, its action is probably linked to a decrease in cellular cAMP levels. Insulin-induced reduction of cAMP could result from the inhibition of adenylate cyclase and/or from a stimulation of cGMP-inhibited low-K, CAMP-phosphodiesterase EC; cGI-PDE). Effects through adenylate cyclase inhibition are still largely questionable and Gi proteins do not play a role in the transduction of the insulin signal in the adipocytes (Wesslau et al. 1993). Activation of cGI-PDE by insulin is believed to be the major mechanism whereby insulin reduces cellular cAMP levels. This activation is the result of serine phosphorylation of cGI-PDE (Degerman et al. 1990; Smith ef al. 1991). A synergistic activation and phosphorylation of cGI-PDE was seen in response to insulin and catecholamines (Smith Manganielio, 1988; Smith et al. 1991). Results obtained in intact adipocytes are consistent with the notion that insulin mediates the phosphorylation of serine site(s) on cGI-PDE and promotes its activation (Eriksson et al. 1995). The insulin-induced activation- phosphorylation of cGI-PDE is catalysed by a cGI-PDE serine kinase (Lopez-Aparicio et al. 1993). The identity of this protein is unknown. Very recently, it was shown that http:/ from http:/ IP address:, on 29 Sep 2016 at 12:37:32, subject to the Cambridge Core terms of use, available at  96 D. LANGIN AND OTHERS zyx Horrnone-sensitive lipase zyxwvu EC Monoacylglycerol lipase X3.1.1.23) zy  \ triacylglycerol dipocyte - 1.2-Diacylglycerol zyxwv   -Monoacylglycerol FFA FFA FFA Fig. 2. Hydrolysis of triacylglycerols stored in adipocytes. The breakdown of triacylglycerols into diacyl- glycerols is the rate-limiting step in adipose tissue lipolysis. FFA, free fatty acids. phosphatidyl inositol 3-kinase, an important mediator of insulin-dependent metabolic effects, was involved in mediating the anti-lipolytic effect of insulin at a step upstream from the activation of the cGI-PDE serine kinase. Further investigations should lead in the near future to a complete characterization of signal transduction steps involved in insulin-mediated anti-lipolysis. zyxwvu n vivo, HSL catalyses the hydrolysis of TAG to diacylglycerol, and then to monoacylglycerol (Fig. 2). The hydrolysis of the monoacylglycerol-fatty acid bond is assured by monoacylglycerol lipase EC The abundance of this enzyme, which is not under hormonal control, is sufficient to avoid accumulation of intermediary products of lipolysis (Fredrikson et al. 1986). HSL exhibits positional specificity for the l(3)-ester bond, although this specificity is less pronounced than that for lipoprotein lipase EC; LPL) or pancreatic lipase EC TAG are hydrolysed at a much lower rate than diacylglycerol. Therefore, the first step of lipolysis is rate-limiting. Moreover, phosphorylation of HSL by CAMP-PK is paralleled by an enhanced TAG lipase EC activity, whereas the activity against diacylglycerol is unchanged. The hallmark of HSL, which distinguishes this enzyme from all other known lipases, is the control of its activity through phosphorylation (Fig. 3). zyx   single serine residue (regulatory site) is phosphorylated by CAMP-PK, 1 mol phosphate being incorporated per mol subunit (Strhlfors Belfrage, 1983). The reversible phosphorylation of the regulatory site controls the active state of the enzyme. However, the mechanism of activation is still unclear. HSL activity is increased 2-3-fold by CAMP-PK-mediated phosphorylation in vitro, while a more than 20-fold increase in lipolytic rate is measured in intact fat cells in response to hormonal stimulation. The TAG emulsion used as substrate in vitro provides a much larger interfacial area than the larger TAG droplet present in the adipocyte (Strhlfors et al. 1987). Hence, a high basal activity of HSL in the dephosphorylated form might be caused in vitro by favourable conditions for enzyme- substrate interaction, phosphorylation promoting a small increase in an already-high submaximal activity. The relationship between the subcellular distribution of HSL and its phosphorylation and activation may be important. HSL has some properties of an intrinsic membrane protein, for example it associates strongly with phospholipids, requires detergents for solubilization and exhibits an amphiphilic character (Holm et al. 1986). Redistribution of HSL may be a major event associated with phosphorylation and activation. On lipolytic stimulation, translocation of HSL from the cytosol to a particulate fraction was found in 3T3-Ll adipocytes (Hirsch Rosen, 1984). This view was recently strengthened. Using mild disruption of rat adipocytes and polyclonal antiserum directed against HSL, a translocation of phosphorylated HSL at the surface of http:/ from http:/ IP address:, on 29 Sep 2016 at 12:37:32, subject to the Cambridge Core terms of use, available at  LIPID ABSORPTION AND METABOLISM 97 z   z MP: ATPt -kKinase kinase pp-2~ Insulin zyxwv z atecholamines t zy 1 Acyl-CoA 4 Fatty acids TAG Export Fig. 3. Short-term regulation zyxwvuts f hormone-sensitive lipase zyxwv EC; HSL). The diagram illustrates the phosphorylation of the regulatory site of human HSL Ser552) by CAMP-dependent protein kinase EC; CAMP-PK) and the hypothesis regarding the phosphorylation of the basal site Ser.554) by 5’- AMP-activated protein kinase S’AMP-PK). PP-2A, PP-2C, protein phosphatases 2A and 2C EC; TAG, triacylglycerol. the lipid droplet was demonstrated (Egan et al. 1992). The nature of the binding of the enzyme to the lipid droplet is not known. The primary structure of HSL does not show any highly hydrophobic regions, which could explain the propensity of HSL to bind to lipids. Early reports (Wise Jungas, 1978) have brought up the question as to whether-a protein component located at the surface of the lipid droplet may undergo some kind of ‘substrate activation’ occurring concomitantly with HSL activation and governing droplet-driven translocation of HSL. It could be speculated that perilipins, specific adipocyte lipid-droplet-associated proteins, are possible candidates as ‘docking’ proteins for HSL (Greenberg et al. 1991). These proteins, probably contribute to the organization of lipid droplets and lipid vacuoles found in mature adipocytes (Hare zy t al. 1994). Perilipins are phosphorylated by CAMP-PK in parallel with activation of lipolysis. This event could represent the ‘substrate activation’ process. The recent isolation of cDNA for perilipins will allow detailed analysis of their role in fat cells (Greenberg et al. 1993). In addition to the regulatory site, another serine residue (basal site) can be phosphorylated (Fig. 3). Three protein kinases i.e. Ca*+/calmodulin-dependent protein kinase I1 EC, glycogen synthase kinase-4 EC and the 5’-AMP- activated protein kinase have been shown to phosphorylate site 2 in vitro. Phosphory- lation of site 2 does not directly alter HSL activity, but can exert a regulatory role since phosphorylation of site 1 and site 2 on HSL are mutually exclusive. Using a synthetic peptide, based on the sequence surrounding sites 1 and 2 of HSL, it was shown that phosphorylation of the peptide at site 2 totally prevents the subsequent phosphorylation of site 1 and vice versa (Garton et al. 1989; Garton Yeaman, 1990). Evidence for a role of 5’-AMP-activated kinase in adipocytes was provided by use of a cell-permeable precursor of 5-amino-imidazole 4-carboxamide ribonucleoside monophosphate (ZMP) http:/ from http:/ IP address:, on 29 Sep 2016 at 12:37:32, subject to the Cambridge Core terms of use, available at
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