The Perilipins are a family of intracellular neutral lipid droplet storage proteins that are responsive to acute protein kinase A-mediated, hormonal stimulation. Perilipin (Peri) expression appears to be limited to adipocytes and steroidogenic cells, in which intracellular neutral lipid hydrolysis is regulated by protein kinase A. cDNA sets and overlapping genomic fragments of the murine Peri locus were isolated and chromosomal location, transcription start sites, polyadenylylation sites, and intron/exon junctions were mapped. Data confirm that the Perilipins are encoded by a single-copy gene, with alternative and tissue-specific, mRNA splicing and polyadenylylation yielding four different protein species. The Perilipin proteins have identical (approximately 22-kDa) amino termini with distinct carboxyl terminal sequences of varying lengths. These genomic and transcriptional maps of murine Perilipin are also essential for evaluating presumptive endogenous and targeted mutations within the locus. The N-terminal identity region of the Perilipins defines a sequence motif, termed PAT, that is shared with the ADRP and TIP47 proteins; additionally, the PAT domain may represent a novel, conserved pattern for lipid storage droplet (LSD) proteins of vertebrates and invertebrates alike. Comparative genomics suggest the presence of related LSD genes in species as diverse as Drosophila and Dictyostelium (Lu, 2002).
The perilipins are the most abundant proteins coating the surfaces of lipid droplets in adipocytes and are found at lower levels surrounding lipid droplets in steroidogenic cells. Perilipins drive triacylglycerol storage in adipocytes by regulating the rate of basal lipolysis and are also required to maximize hormonally stimulated lipolysis. To map the domains that target and anchor perilipin A to lipid droplets, fragments of perilipin A were stabily expressed in 3T3-L1 fibroblasts. Immunofluorescence microscopy and immunoblotting of proteins from isolated lipid droplets revealed that neither the amino nor the carboxyl terminus is required to target perilipin A to lipid droplets; however, there are multiple, partially redundant targeting signals within a central domain including 25% of the primary amino acid sequence. A peptide composed of the central domain of perilipin A directs a fused green fluorescent protein to the surfaces of lipid droplets. Full-length perilipin A associates with lipid droplets via hydrophobic interactions, as shown by the persistence of perilipins on lipid droplets after centrifugation through an alkaline carbonate solution. Results of the mutagenesis studies indicate that the sequences responsible for anchoring perilipin A to lipid droplets are most likely domains of moderately hydrophobic amino acids located within the central 25% of the protein. Thus, it is concluded that the central 25% of the perilipin A sequence contains all of the amino acids necessary to target and anchor the protein to lipid droplets (Garcia, 2003).
Perilipin A is the most abundant lipid droplet-associated protein in adipocytes and serves important functions in regulating triacylglycerol levels by reducing rates of basal lipolysis and facilitating hormonally stimulated lipolysis. The central region of perilipin A targets and anchors it to lipid droplets, at least in part via three moderately hydrophobic sequences that embed the protein into the hydrophobic core of the droplet. The current study examines the roles of the amino and carboxyl termini of perilipin A in facilitating triacylglycerol storage. Amino- and carboxyl-terminal truncation mutations of mouse perilipin A were stably expressed in 3T3-L1 preadipocytes, which lack perilipins. Triacylglycerol content of the cells was quantified as a measure of perilipin function and was compared with that of cells expressing full-length perilipin A or control cells lacking perilipins. The amino-terminal sequence between amino acids 122 and 222, including four 10-11-amino acid sequences predicted to form amphipathic beta-strands and a consensus site for cAMP-dependent protein kinase, and the carboxyl terminus of 112 amino acids that is unique to perilipin A are critical to facilitate triacylglycerol storage. The precocious expression of full-length perilipin A in 3T3-L1 preadipocytes aided more rapid storage of triacylglycerol during adipose differentiation. By contrast, the expression of highly truncated amino- or carboxyl-terminal mutations of perilipin failed to serve a dominant negative function in lowering triacylglycerol storage during adipose differentiation. It has been concluded that the amino and carboxyl termini are critical to the function of perilipin A in facilitating triacylglycerol storage (Garciaw, 2004).
Perilipin coats the lipid droplets of adipocytes and is thought to have a role in regulating triacylglycerol hydrolysis. To study the role of perilipin in vivo, a perilipin knockout mouse was created. Perilipin null (peri-/-) and wild-type (peri+/+) mice consume equal amounts of food, but the adipose tissue mass in the null animals is reduced to approximately 30% of that in wild-type animals. Isolated adipocytes of perilipin null mice exhibit elevated basal lipolysis because of the loss of the protective function of perilipin. They also exhibit dramatically attenuated stimulated lipolytic activity, indicating that perilipin is required for maximal lipolytic activity. Plasma leptin concentrations in null animals were greater than expected for the reduced adipose mass. The peri-/- animals have a greater lean body mass and increased metabolic rate but they also show an increased tendency to develop glucose intolerance and peripheral insulin resistance. When fed a high-fat diet, the perilipin null animals are resistant to diet-induced obesity but not to glucose intolerance. The data reveal a major role for perilipin in adipose lipid metabolism and suggest perilipin as a potential target for attacking problems associated with obesity (Tansey, 2001).
Obesity is a major risk factor for diabetes and heart disease. Inactivation of the gene for perilipin (plin), an adipocyte lipid droplet surface protein, produces lean and obesity-resistant mice. To dissect the underlying mechanisms involved, oligonucleotide microarrays were used to analyze the gene-expression profile of white adipose tissue (WAT), liver, heart, skeletal muscle, and kidney of plin-/- and plin+/+ mice. As compared with wild-type littermates, the WAT of plin-/- mice had 270 and 543 transcripts that were significantly up- or down-regulated. There was a coordinated upregulation of genes involved in beta-oxidation, the Krebs cycle, and the electron transport chain concomitant with a downregulation of genes involved in lipid biosynthesis. There was also a significant downregulation of the stearoyl CoA desaturase-1 gene, which has been associated with obesity resistance. Thus, in response to the constitutive activation of lipolysis associated with absence of perilipin, WAT activates pathways to rid itself of the products of lipolysis and activates pathways of energy expenditure that contribute to the observed obesity resistance. The biochemical pathways involved in obesity resistance in plin-/- mice identified in this study may represent potential targets for the treatment of obesity (Castro-Chavez, 2003).
Targeted disruption of the lipid droplet protein, perilipin, in mice, leads to constitutional lipolysis associated with marked reduction in white adipose tissue as a result of unbridled lipolysis. To investigate the metabolic adaptations in response to the constitutive lipolysis, perilipin-null (plin-/-) mice were studied in terms of their fatty acid oxidation and glycerol and glucose metabolism homeostasis by using dynamic biochemical testing and clamp and tracer infusion methods. plin-/- mice show increased beta-oxidation in muscle, liver, and adipose tissue resulting from a coordinated regulation of the enzymes and proteins involved in beta-oxidation. The increased beta-oxidation helped remove the extra free fatty acids created by the constitutive lipolysis. An increase in the expression of the transcripts for uncoupling proteins-2 and -3 also accompany this increase in fatty acid oxidation. Adult plin-/- mice have normal plasma glucose but a reduced basal hepatic glucose production (46% that of plin+/+). Insulin infusion during low dose hyperinsulinemic-euglycemic clamp further lowers the glucose production in plin-/- mice, but plin-/- mice also show a 36% decrease in glucose disposal rate during the low dose insulin clamp, indicating peripheral insulin resistance. However, compared with plin+/+ mice, 14-week-old plin-/- mice show no significant difference in glucose disposal rate during the high dose hyperinsulinemic clamp, whereas 42-week-old plin-/- mice display significant insulin resistance on high dose hyperinsulinemic clamp. Despite increasing insulin resistance with age, plin-/- mice at different ages maintain a normal glucose response during an intraperitoneal glucose tolerance curve, being compensated by the increased beta-oxidation and reduced hepatic glucose production. These experiments uncover the metabolic adaptations associated with the constitutional lipolysis in plin-/- mice that allow the mice to continue to exhibit normal glucose tolerance in the presence of peripheral insulin resistance (Saha, 2004).
In a systematic search for peroxisome proliferator-activated receptor-gamma (PPAR-gamma) target genes, S3-12 and perilipin were identified as novel direct PPAR-gamma target genes. Together with adipophilin and tail-interacting protein of 47 kDa, these genes are lipid droplet-associating proteins with distinct expression pattern but overlapping expression in adipose tissue. The expression of S3-12 and perilipin is tightly correlated to the expression and activation of PPAR-gamma in adipocytes, and promoter characterization revealed that the S3-12 and the perilipin promoters contain three and one evolutionarily conserved PPAR response elements, respectively. The expression of S3-12 and perilipin is reduced in obese compared with lean Zucker rats, whereas the expression of adipophilin is increased. Perilipin has been shown to be an essential factor in the hormonal regulation of lipolysis of stored triglycerides within adipose tissue. The direct regulation of perilipin and S3-12 by PPAR-gamma therefore is likely to be an important mediator of the in vivo effects of prolonged treatment with PPAR-gamma activators: insulin sensitization, fatty acid trapping in adipose tissue, reduced basal adipose lipolysis, and weight gain (Dalen, 2004).
Most cis-acting regulatory elements have generally been assumed to activate a single nearby gene. However, many genes are clustered together, raising the possibility that they are regulated through a common element. A single peroxisome proliferator response element (PPRE), located between the mouse PEX11 alpha and perilipin genes, confers on both genes activation by peroxisome proliferator-activated receptor alpha (PPAR alpha) and PPAR gamma. A functional PPRE 8.4 kb downstream of the promoter of PEX11 alpha, a PPAR alpha target gene, was identified by a gene transfection study. This PPRE is positioned 1.9 kb upstream of the perilipin gene and also functions with the perilipin promoter. In addition, this PPRE, when combined with the natural promoters of the PEX11 alpha and perilipin genes, confers subtype-selective activation by PPAR alpha and PPAR gamma 2. The PPRE sequence specifically binds to the heterodimer of RXR alpha and PPAR alpha or PPAR gamma 2, as assessed by electrophoretic gel mobility shift assays. Furthermore, tissue-selective binding of PPAR alpha and PPAR gamma to the PPRE was demonstrated in hepatocytes and adipocytes, respectively, by chromatin immunoprecipitation assay. Hence, the expression of these genes is induced through the same PPRE in the liver and adipose tissue, where the two PPAR subtypes are specifically expressed (Shimizu, 2004).
Recent studies have shown that lipid droplets are covered with a proteinaceous coat, although the functions and identities of the component proteins have not yet been well elucidated. The first identified lipid droplet-specific proteins are the perilipins, a family of proteins coating the surfaces of lipid droplets of adipocytes. The generation of perilipin-null mice has revealed that although they consume more food than control mice, they have normal body weight and are resistant to diet-induced obesity. In one study it was reported that in an animal model obesity was reversible by breeding perilipin -/- alleles into Lepr db/db obese mice, ostensibly by increasing the metabolic rate of the mice. To understand the exact mechanisms that drive the exclusive expression of the perilipin gene in adipocytes, the 5'-flanking region of the mouse gene was examined. Treatment of differentiating 3T3-L1 adipocytes with an agonist of proliferator-activated receptor (PPAR) gamma, the putative 'master regulator' of adipocyte differentiation, significantly augments perilipin gene expression. Reporter assays using the -2.0-kb promoter revealed that this region contains a functional PPARgamma-responsive element. Gel mobility shift and chromatin immunoprecipitation assays showed that endogenous PPARgamma protein binds to the perilipin promoter. PPARgamma2, an isoform exclusively expressed in adipocytes, was found to be the most potent regulator from among the PPAR family members including PPARalpha and PPARgamma1. These results make evident the fact that perilipin gene expression in differentiating adipocytes is crucially regulated by PPARgamma2, providing new insights into the adipogenic action of PPARgamma2 and adipose-specific gene expression, as well as potential anti-obesity pharmaceutical agents targeted to a reduction of the perilipin gene product (Arimura, 2004).
Expression of FoxC2 blocks the capacity of 3T3-L1 preadipocytes to undergo adipogenesis in the presence of dexamethasone, isobutylmethylxanthine, and insulin. This block is characterized by an extensive decrease in the expression of proteins associated with the function of the mature fat cell, most notably C/EBPalpha, adiponectin, perilipin, and the adipose-specific fatty acid-binding protein, FABP4/aP2. Since the expression of these proteins lies downstream of PPARgamma, overexpressed PPARgamma was overexpressed in Swiss mouse fibroblasts to promote adipocyte differentiation. FoxC2 blocks the ability of PPARgamma to induce adipogenic gene expression in response to exposure of the cells to dexamethasone, isobutylmethylxanthine, insulin, and a PPARgamma ligand. Interestingly, the expression of aP2 escapes the inhibitory action of FoxC2 under conditions that promote maximum PPARgamma activity. In contrast, FoxC2 inhibits the expression of C/EBPalpha, perilipin, and adiponectin even in the presence of potent PPARgamma ligands. Finally, it has been shown that FoxC2 does not affect the ability of PPARgamma to bind to or transactivate from a PPARgamma response element. These data suggest that FoxC2 blocks adipogenesis by inhibiting the capacity of PPARgamma to promote the expression of a subset of adipogenic genes (Davis, 2004).
Perilipin, a family of phosphoproteins located around lipid droplets in adipocytes, is essential for enlargement of lipid droplets and lipolytic reaction by hormone-sensitive lipase. Thiazolidinediones, peroxisome proliferator-activated receptor (PPAR) gamma agonists, have been shown to increase perilipin expression in fully differentiated adipocytes. However, the precise mechanism of transcriptional regulation of murine perilipin gene heretofore remains unclear. The transcription start site of murine perilipin gene was determined by RNA ligase-mediated rapid amplification of the cDNA ends method. Luciferase reporter gene constructs containing various lengths of the 5'-flanking region of the murine perilipin gene were generated and promoter/enhancer activities were assayed using differentiated 3T3-L1 adipocytes. A functional PPAR-responsive element (PPRE) was identified in the murine perilipin promoter, and this was confirmed by gel EMSAs using nuclear extracts from differentiated 3T3-L1 adipocytes. Furthermore, point mutations of the identified functional PPRE markedly reduce both the reporter gene activity in differentiated 3T3-L1 adipocytes and PPARgamma/thiazolidinedione-induced transactivation in NIH-3T3 fibroblasts. Real-time RT-PCR reveals that thiazolidinedione up-regulates endogenous perilipin mRNA levels. It is proposed that PPARgamma plays a significant role in the transcriptional regulation of murine perilipin gene via the PPRE in its promoter (Nagai, 2004).
Perilipin (Peri) A is a phosphoprotein located at the surface of intracellular lipid droplets in adipocytes. Activation of cyclic AMP-dependent protein kinase (PKA) results in the phosphorylation of Peri A and hormone-sensitive lipase (HSL), the predominant lipase in adipocytes, with concurrent stimulation of adipocyte lipolysis. To investigate the relative contributions of Peri A and HSL in basal and PKA-mediated lipolysis, NIH 3T3 fibroblasts were used lacking Peri A and HSL but stably overexpressing acyl-CoA synthetase 1 (ACS1) and fatty acid transport protein 1 (FATP1). When incubated with exogenous fatty acids, ACS1/FATP1 cells accumulate 5 times more triacylglycerol (TG) as compared with NIH 3T3 fibroblasts. Adenoviral-mediated expression of Peri A in ACS1/FATP1 cells enhances TG accumulation and inhibits lipolysis, whereas expression of HSL fused to green fluorescent protein (GFPHSL) reduces TG accumulation and enhances lipolysis. Forskolin treatment induces Peri A hyperphosphorylation and abrogates the inhibitory effect of Peri A on lipolysis. Expression of a mutated Peri A Delta 3 (Ser to Ala substitutions at PKA consensus sites Ser-81, Ser-222, and Ser-276) reduces Peri A hyperphosphorylation and blocks constitutive and forskolin-stimulated lipolysis. Thus, perilipin expression and phosphorylation state are critical regulators of lipid storage and hydrolysis in ACS1/FATP1 cells (Souza, 2002).
Perilipin A coats the lipid storage droplets in adipocytes and is polyphosphorylated by protein kinase A (PKA); the fact that PKA activates lipolysis in adipocytes suggests a role for perilipins in this process. To assess whether perilipins participate directly in PKA-mediated lipolysis, constructs coding for native and mutated forms of the two major splice variants of the perilipin gene, perilipins A and B, were expressed in Chinese hamster ovary fibroblasts. Perilipins localize to lipid droplet surfaces and displace the adipose differentiation-related protein that normally coats the droplets in these cells. Perilipin A inhibits triacylglycerol hydrolysis by 87% when PKA is quiescent, but activation of PKA and phosphorylation of perilipin A engenders a 7-fold lipolytic activation. Mutation of PKA sites within the N-terminal region of perilipin abrogates the PKA-mediated lipolytic response. In contrast, perilipin B exerts only minimal protection against lipolysis and is unresponsive to PKA activation. Since Chinese hamster ovary cells contain no PKA-activated lipase, it is concluded that the expression of perilipin A alone is sufficient to confer PKA-mediated lipolysis in these cells. Moreover, the data indicate that the unique C-terminal portion of perilipin A is responsible for its protection against lipolysis and that phosphorylation at the N-terminal PKA sites attenuates this protective effect (Tansey, 2003).
Perilipin (Peri) A is a lipid droplet-associated phosphoprotein that acts dually as a suppressor of basal (constitutive) lipolysis and as an enhancer of cyclic AMP-dependent protein kinase (PKA)-stimulated lipolysis by both hormone-sensitive lipase (HSL) and non-HSL(s). To identify domains of Peri A that mediate these multiple actions, adenoviruses expressing truncated or mutated Peri A and HSL were introduced into NIH 3T3 fibroblasts lacking endogenous perilipins and HSL but overexpressing acyl-CoA synthetase 1 and fatty acid transporter 1. Two lipase-selective functional domains were identified: (1) Peri A (amino acids 1-300), which inhibits basal lipolysis and promotes PKA-stimulated lipolysis by HSL, and (2) Peri A (amino acids 301-517), which inhibits basal lipolysis by non-HSL and promotes PKA-stimulated lipolysis by both HSL and non-HSL. PKA site mutagenesis reveals that PKA-stimulated lipolysis by HSL requires phosphorylation of one or more sites within Peri 1-300 (Ser81, Ser222, and Ser276). PKA-stimulated lipolysis by non-HSL additionally requires phosphorylation of one or more PKA sites within Peri 301-517 (Ser433, Ser492, and Ser517). Peri 301-517 promotes PKA-stimulated lipolysis by HSL yet does not block HSL-mediated basal lipolysis, indicating that an additional region(s) within Peri 301-517 promotes hormone-stimulates lipolysis by HSL. These results suggest a model of Peri A function in which (1) lipase-specific 'barrier' domains block basal lipolysis by HSL and non-HSL, (2) differential PKA site phosphorylation allows PKA-stimulated lipolysis by HSL and non-HSL, respectively, and (3) additional domains within Peri A further facilitate PKA-stimulated lipolysis, again with lipase selectivity (Zhang, 2003).
Perilipins, the major structural proteins coating the surfaces of mature lipid droplets of adipocytes, play an important role in the regulation of triacylglycerol storage and hydrolysis. Proteomic analysis was used to identify CGI-58, a member of the alpha/beta-hydrolase fold family of enzymes, as a component of lipid droplets of 3T3-L1 adipocytes. CGI-58 mRNA is highly expressed in adipose tissue and testes, tissues that also express perilipins, and at lower levels in liver, skin, kidney, and heart. Both endogenous CGI-58 and an ectopic CGI-58-GFP chimera show diffuse cytoplasmic localization in 3T3-L1 preadipocytes, but localize almost exclusively to the surfaces of lipid droplets in differentiated 3T3-L1 adipocytes. The localization of endogenous CGI-58 was investigated in 3T3-L1 cells stably expressing mutated forms of perilipin using microscopy. CGI-58 binds to lipid droplets coated with perilipin A or mutated forms of perilipin with an intact C-terminal sequence from amino acid 382 to 429, but not to lipid droplets coated with perilipin B or mutated perilipin A lacking this sequence. Immunoprecipitation studies confirmed these findings, but also showed co-precipitation of perilipin B and CGI-58. Remarkably, activation of cAMP-dependent protein kinase by the incubation of 3T3-L1 adipocytes with isoproterenol and isobutylmethylxanthine disperses CGI-58 from the surfaces of lipid droplets to a cytoplasmic distribution. This shift in subcellular localization can be reversed by the addition of propanolol to the culture medium. Thus, CGI-58 binds to perilipin A-coated lipid droplets in a manner that is dependent upon the metabolic status of the adipocyte and the activity of cAMP-dependent protein kinase (Subramanian. 2004).
Lipid droplets (LDs) are a class of ubiquitous cellular organelles that are involved in lipid storage and metabolism. Although the mechanisms of the biogenesis of LDs are still unclear, a set of proteins called the PAT domain family have been characterized as factors associating with LDs. Perilipin, a member of this family, is expressed exclusively in the adipose tissue and regulates the breakdown of triacylglycerol in LDs via its phosphorylation. A yeast two-hybrid system was used to examine the potential function of perilipin. Direct interaction was found between perilipin and CGI-58, a deficiency of which correlated with the pathogenesis of Chanarin-Dorfman syndrome (CDS). Endogenous CGI-58 is distributed predominantly on the surface of LDs in differentiated 3T3-L1 cells, and its expression increases during adipocyte differentiation. Overexpressed CGI-58 tagged with GFP gathers at the surface of LDs and colocalizes with perilipin. This interaction seems physiologically important because CGI-58 mutants carrying an amino acid substitution identical to that found in CDS lost the ability to be recruited to LDs. These mutations significantly weakened the binding of CGI-58 with perilipin, indicating that the loss of this interaction is involved in the etiology of CDS. Furthermore, CGI-58 was identified as a binding partner of ADRP, another PAT domain protein expressed ubiquitously. GFP-CGI-58 expressed in non-differentiated 3T3-L1 or CHO-K1 cells colocalizes with ADRP, and CGI-58 mutants are not recruited to LDs carrying ADRP, indicating that CGI-58 may also cooperate with ADRP (Yamaguchi, 2004).
Adipocyte lipolysis was compared with hormone-sensitive lipase (HSL)/perilipin subcellular distribution and perilipin phosphorylation using Western blot analysis. Under basal conditions, HSL resides predominantly in the cytosol and unphosphorylated perilipin resides predominantly on the surface of the lipid droplet. Upon lipolytic stimulation of adipocytes isolated from young rats with the beta-adrenergic agonist, isoproterenol, HSL translocates from the cytosol to the lipid droplet, but there was no movement of perilipin from the droplet to the cytosol; however, perilipin phosphorylation was observed. By contrast, upon lipolytic stimulation and perilipin phosphorylation in cells from more mature rats, there was no HSL translocation but a significant movement of perilipin away from the lipid droplet. Adipocytes from younger rats have markedly greater rates of lipolysis than those from the older rats. Thus high rates of lipolysis require translocation of HSL to the lipid droplet and translocation of HSL and perilipin can occur independently of each other. A loss of the ability to translocate HSL to the lipid droplet probably contributes to the diminished lipolytic response to catecholamines with age (Clifford, 2000).
The perilipins are the most abundant proteins at the surfaces of lipid droplets in adipocytes and are also found in steroidogenic cells. To investigate perilipin function, perilipin A, the predominant isoform, was ectopically expressed in fibroblastic 3T3-L1 pre-adipocytes that normally lack the perilipins. In control cells, fluorescent staining of neutral lipids with Bodipy 493/503 show a few minute and widely dispersed lipid droplets, while in cells stably expressing perilipin A, the lipid droplets were more numerous and tightly clustered in one or two regions of the cytoplasm. Immunofluorescence microscopy revealed that the ectopic perilipin A localizes to the surfaces of the tiny clustered lipid droplets; subcellular fractionation of the cells using sucrose gradients confirmed that the perilipin A localizes exclusively to lipid droplets. Cells expressing perilipin A store 6- to 30-fold more triacylglycerol than control cells due to reduced lipolysis of triacylglycerol stores. The lipolysis of stored triacylglycerol is 5 times slower in lipid-loaded cells expressing perilipin A than in lipid-loaded control cells, when triacylglycerol synthesis is blocked with 6 microm triacsin C. This stabilization of triacylglycerol is not due to the suppression of triacylglycerol lipase activity by the expression of perilipin A. It is concluded that perilipin A increases the triacylglycerol content of cells by forming a barrier that reduces the access of soluble lipases to stored lipids, thus inhibiting triacylglycerol hydrolysis. These studies suggest that perilipin A plays a major role in the regulation of triacylglycerol storage and lipolysis in adipocytes (Brasaemle, 2000).
A key step in lipolytic activation of adipocytes is the translocation of hormone-sensitive lipase (HSL) from the cytosol to the surface of the lipid storage droplet. Adipocytes from perilipin-null animals have an elevated basal rate of lipolysis compared with adipocytes from wild-type mice, but fail to respond maximally to lipolytic stimuli. This defect is downstream of the beta-adrenergic receptor-adenylyl cyclase complex. HSL is basally associated with lipid droplet surfaces at a low level in perilipin nulls, but stimulated translocation from the cytosol to lipid droplets is absent in adipocytes derived from embryonic fibroblasts of perilipin-null mice. The HSL translocation reaction was reconstructed in the nonadipocyte Chinese hamster ovary cell line by introduction of GFP-tagged HSL with and without perilipin A. On activation of protein kinase A, HSL-GFP translocates to lipid droplets only in cells that express fully phosphorylatable perilipin A, confirming that perilipin is required to elicit the HSL translocation reaction. Moreover, in Chinese hamster ovary cells that express both HSL and perilipin A, these two proteins cooperate to produce a more rapidly accelerated lipolysis than do cells that express either of these proteins alone, indicating that lipolysis is a concerted reaction mediated by both protein kinase A-phosphorylated HSL and perilipin A (Sztalryd, 2003).
Transgenic mice overexpressing leptin (LepTg) exhibit substantial reductions in adipose mass. Since the binding of leptin to its receptor activates the sympathetic nervous system, it was reasoned that the lean state of the LepTg mice could be caused by chronic lipolysis. Instead, the LepTg mice exhibit a low basal lipolysis state and their lean phenotype is not dependent on the presence of beta3-adrenergic receptors. In their white adipose tissue, protein levels of protein kinase A, hormone-sensitive lipase, and ADRP are not impaired. However, compared to normal mice, perilipin, perilipin mRNA, and cAMP-stimulated PKA activity are significantly attenuated. Overall, it is demonstrated that the lean phenotype of the LepTg mice does not result in a chronically elevated lipolytic state, but instead in a low basal lipolysis state characterized by a decrease in perilipin and PKA activity in white fat (Ke, 2003).
Obesity is a disorder of energy balance. Hormone-sensitive lipase (HSL) mediates the hydrolysis of triacylglycerol, the major form of stored energy in the body. Perilipin (encoded by the gene Plin), an adipocyte protein, has been postulated to modulate HSL activity. Targeted disruption of Plin results in healthy mice that have constitutively activated fat-cell HSL. Plin-/- mice consume more food than control mice, but have normal body weight. They are much leaner and more muscular than controls, have 62% smaller white adipocytes, show elevated basal lipolysis that is resistant to beta-adrenergic agonist stimulation, and are cold-sensitive except when fed. They are also resistant to diet-induced obesity. Breeding the Plin-/- alleles into Leprdb/db mice reverses the obesity by increasing the metabolic rate of the mice. These results demonstrate a role for perilipin in reining in basal HSL activity and regulating lipolysis and energy balance; thus, agents that inactivate perilipin may prove useful as anti-obesity medications (Martinez-Botas, 2000).
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