LSDP5在肝脏脂肪和脂滴代谢中的作用及机制研究
|
摘要
研究背景：
脂肪肝（liver steatosis or fatty liver）是各种病因造成脂肪代谢异常时最为常见的一种肝脏疾病，在我国约20%~30%的人患有脂肪肝。脂肪肝的典型病理变化是肝细胞内过多中性脂肪（neutral lipid）储积，并出现脂滴（lipiddroplet，LD）。目前对脂肪肝的病因研究较多，但对肝细胞内脂滴代谢和形成的研究较少。脂滴（LD）早期被认为仅仅是一个类似于糖原的能量储存结构，但是目前研究表明，脂滴并非细胞内一个简单的能量贮存器，而是一个复杂的、活动旺盛、动态变化的多功能细胞成分。脂滴可能在脂类代谢与存储、膜转运、蛋白降解，以及信号传导过程中均起着重要的作用。脂滴表面镶嵌着多种脂滴蛋白（lipid droplet proteins，LDP），其对脂滴的稳定性和代谢过程均具有非常重要的作用。目前，在所有脂滴蛋白中，PAT家族成员是最为重要，也是研究最多的一组分子。PAT家族主要包括perilipin、adipophilin、TIP47（tail-interacting protein of47kDa）、S3-12以及LSDP5（lipid storage droplet protein5）。其中LSDP5是PAT家族第五个成员，主要分布于脂肪分解代谢速度较快的组织中，如肝脏、心肌、骨骼肌等，因此又称为OXPAT（oxidative tissue-enriched PAT protein）/PAT-1或MLDP（myocardial lipid droplet protein），但是其具体作用及机制目前尚不明确。
研究目的：
本研究拟确定LSDP5的亚细胞定位；通过构建LSDP5的截短体，分析LSDP5各个结构域的功能；确定LSDP5对肝细胞脂滴数量和大小、甘油三酯的合成和分解的影响，明确LSDP5对肝细胞脂肪代谢的作用；确定LSDP5的相互作用分子，分析LSDP5对脂滴代谢相关分子表达的影响，阐明LSDP5对肝脂肪代谢影响可能的分子机制。
研究方法：
1、利用免疫荧光的方法明确LSDP5在肝细胞内的分布与亚细胞定位；
2、在肝细胞系和原代肝细胞中改变LSDP5的表达，观察其对肝细胞脂滴形态和脂代谢的影响。
3、以LSDP5为诱饵蛋白，通过酵母双杂交系统来筛选与之相互作用分子。并进一步通过免疫共沉淀和免疫荧光共定位验证其相互作用，以相互作用的分子为线索，最终阐明LSDP5的分子功能。
研究结果：
1、LSDP5是一个脂滴表面蛋白（LDP）。使用免疫荧光技术，我们发现LSDP5定位在脂滴表面。构建LSDP5截短体，进一步研究发现LSDP5的N端，含有PAT家族保守性的PAT-1区和11-mer α-螺旋的重复序列，是LSDP5脂滴定位的关键部位。同时，细胞内甘油三酯含量测定结果提示，LSDP5的N端也是影响脂滴储积的关键区域。
2、LSDP5增加肝细胞内中性脂肪的储积。
在肝细胞内过表达LSDP5，利用Nile Red荧光染料显示肝细胞内脂滴的形态和数量。结果表明LSDP5能够明显增加肝细胞内大脂滴的比例，提高细胞内甘油三酯的含量。LSDP5的沉默使细胞内的脂滴增大受到明显的抑制，胞浆内脂滴明显变小，肝细胞中甘油三酯含量也明显降低。
3、 LSDP5可以影响脂肪分解和脂肪酸的氧化，并与具有脂肪分解作用的蛋白ES1（esterase1）存在相互作用。
LSDP5的沉默刺激脂肪分解，上调甘油三酯分解速率和脂肪分解相关酶的转录，增加脂肪酸β氧化速率和线粒体的数目。通过酵母双杂交，我们筛选出了与LSDP5相互作用的分子ES1。ES1是一个在肝脏高表达的65KD的糖蛋白，它能够水解许多的酯类，包括甘油三酯和其他的脂肪酸酯。进一步应用免疫共沉淀和免疫荧光共定位，我们证明LSDP5能够与ES1相互作用。
结论：
本研究证实LSDP5是一个定位于脂滴表面的蛋白质，可以促进肝细胞内脂滴的增大和甘油三酯的储积。其机制可能是通过LSDP5与ES1的相互作用，抑制ES1对于甘油三酯的水解，并进一步减少脂肪酸的β氧化来实现的。抑制LSDP5的表达，可以有效减少肝细胞内甘油三酯的储积，因此LSDP5有望成为治疗脂肪肝的新靶点。
Background：
Liver steatosis or fatty liver which is believed to be caused by abnormalmetabolism of fats is one of the most common metabolic disorders in China.Nearly Chinese20-30%people have fatty livers. The typical pathological featureof liver steatosis is the lipid deposition in liver cells in the form of lipid droplets(LDs). With regard to etiology analysis, the researches on metabolism andformation of lipid droplets are involved little. Lipid droplets are neutral lipidstorage surrounded by a phospholipid monolayer, which are now recognized tobe functional subcellular organelles rather than metabolically inactive lipiddepots. They are involved in multiple intracellular processes including lipidmetabolism, vesicle traffic, and cell signaling through interactions with otherorganelles so that important roles in lipid homeostasis are likely. In mammaliancells, LDs contain a class of proteins in their surface layers that contribute to thelipolysis, synthesis and fusion of LDs and share a homologous sequence calledthe PAT domain, including perilipin, adipophilin/adipose differentiation-related protein (ADRP), a tail-interacting protein of47kDa (TIP47), and S3-12. Lipidstorage droplet protein5(LSDP5) is a newly identified member of PAT family. Initialidentifications and characterizations of LSDP5as a lipid droplet binding protein werereported by three independent groups as myocardial lipid droplet protein (MLDP),oxidative tissue-enriched PAT protein (OXPAT), and LSDP5respectively. All thesereports are in agreement that LSDP5is expressed in tissues that exhibit high levels offatty acid oxidation, including heart, skeletal muscle, and liver. However, as a newmember of PAT family, the exact function of LSDP5has not been elucidated so far.
Objective:
We initiate the current study to determine the subcellular localization ofLSDP5protein and to further characterize the LD-localization signal sequenceswithin LSDP5. We investigate the effects of LSDP5on the numbers and sizes ofLD, synthesis and hydrolysis of triglycerides (TG). We indentify theLSDP5-interacting proteins and facilitate the illustration of the molecularmechanism of LSDP5.
Methods:
1) Immunofluorescence was used to detect the the distribution and subcellularlocalization of LSDP5protein.
2) The biological functions of LSDP5were observed by upregulating anddownregulating the expression of LSDP5in hepatic cell line AML12and
primary mouse hepatocytes.
3) The interacting-proteins of LSDP5were screened through the mouse livercDNA library with the prey protein LSDP5using yeast two-hybrid.Co-immunoprecipitation and subcellular localization of LSDP5and the interacting-protein were employed to confirm the specificity of theinteractions.
Results:
1) LSDP5protein was located in the surface of lipid droplets.By using immunofluorescence assay, we observed that LSDP5was localizedto the surface of lipid droplets in hepatocytes. Serial deletion analyses showedthat the lipid droplet targeting domain and the domain directing lipid dropletclustering were overlapped and localized to the188amino acid residues at theN-terminal region of LSDP5.
2) LSDP5enhances triglyceride storage in hepatocytes.Overexpression of LSDP5could enhance lipid accumulation in primaryhepatocytes and hepatic cell line AML12. Moreover, knock-down of LSDP5significantly decreased the size of lipid droplets and reduced the contents oftriglyceride.
3) LSDP5could inhibit lipolysis and interacted with esterase1(ES1).Knock-down of LSDP5significantly stimulated lipolysis in cells, and furtherincreased fatty acid β-oxidation and mitochondrial number. Real-time PCRanalysis revealed that the depletion of LSDP5was associated with increasedmRNA level of enzymes in lipolysis and fatty acid oxidation. In addition, weperformed yeast two-hybrid using LSDP5as a bait protein. Among all thesepositive clones we got, ES1was choosed for the further investigation. ES1isa65-kDa glycoprotein present in liver able to hydrolyze a variety of estersincluding triglycerides, cholesteryl esters and fatty acid esters.Co-immunoprecipitation and lmmunofluorescent co-localization furtherlyproved that LSDP5and ES1possessed physical interactions.
Conclusions
Our data clearly demonstrate that LSDP5, a novel lipid droplet protein,contributes to triglyceride accumulation by negatively regulating lipolysis andfatty acid oxidation in hepatocytes may be through mechanisms of the interactionof LSDP5and ES1. This study makes an experimental foundation for the role andmechanism of LSDP5in lipid metabolism and may provide new thinking for thetreatment of fatty liver.
脂肪肝（liver steatosis or fatty liver）是各种病因造成脂肪代谢异常时最为常见的一种肝脏疾病，在我国约20%~30%的人患有脂肪肝。脂肪肝的典型病理变化是肝细胞内过多中性脂肪（neutral lipid）储积，并出现脂滴（lipiddroplet，LD）。目前对脂肪肝的病因研究较多，但对肝细胞内脂滴代谢和形成的研究较少。脂滴（LD）早期被认为仅仅是一个类似于糖原的能量储存结构，但是目前研究表明，脂滴并非细胞内一个简单的能量贮存器，而是一个复杂的、活动旺盛、动态变化的多功能细胞成分。脂滴可能在脂类代谢与存储、膜转运、蛋白降解，以及信号传导过程中均起着重要的作用。脂滴表面镶嵌着多种脂滴蛋白（lipid droplet proteins，LDP），其对脂滴的稳定性和代谢过程均具有非常重要的作用。目前，在所有脂滴蛋白中，PAT家族成员是最为重要，也是研究最多的一组分子。PAT家族主要包括perilipin、adipophilin、TIP47（tail-interacting protein of47kDa）、S3-12以及LSDP5（lipid storage droplet protein5）。其中LSDP5是PAT家族第五个成员，主要分布于脂肪分解代谢速度较快的组织中，如肝脏、心肌、骨骼肌等，因此又称为OXPAT（oxidative tissue-enriched PAT protein）/PAT-1或MLDP（myocardial lipid droplet protein），但是其具体作用及机制目前尚不明确。
研究目的：
本研究拟确定LSDP5的亚细胞定位；通过构建LSDP5的截短体，分析LSDP5各个结构域的功能；确定LSDP5对肝细胞脂滴数量和大小、甘油三酯的合成和分解的影响，明确LSDP5对肝细胞脂肪代谢的作用；确定LSDP5的相互作用分子，分析LSDP5对脂滴代谢相关分子表达的影响，阐明LSDP5对肝脂肪代谢影响可能的分子机制。
研究方法：
1、利用免疫荧光的方法明确LSDP5在肝细胞内的分布与亚细胞定位；
2、在肝细胞系和原代肝细胞中改变LSDP5的表达，观察其对肝细胞脂滴形态和脂代谢的影响。
3、以LSDP5为诱饵蛋白，通过酵母双杂交系统来筛选与之相互作用分子。并进一步通过免疫共沉淀和免疫荧光共定位验证其相互作用，以相互作用的分子为线索，最终阐明LSDP5的分子功能。
研究结果：
1、LSDP5是一个脂滴表面蛋白（LDP）。使用免疫荧光技术，我们发现LSDP5定位在脂滴表面。构建LSDP5截短体，进一步研究发现LSDP5的N端，含有PAT家族保守性的PAT-1区和11-mer α-螺旋的重复序列，是LSDP5脂滴定位的关键部位。同时，细胞内甘油三酯含量测定结果提示，LSDP5的N端也是影响脂滴储积的关键区域。
2、LSDP5增加肝细胞内中性脂肪的储积。
在肝细胞内过表达LSDP5，利用Nile Red荧光染料显示肝细胞内脂滴的形态和数量。结果表明LSDP5能够明显增加肝细胞内大脂滴的比例，提高细胞内甘油三酯的含量。LSDP5的沉默使细胞内的脂滴增大受到明显的抑制，胞浆内脂滴明显变小，肝细胞中甘油三酯含量也明显降低。
3、 LSDP5可以影响脂肪分解和脂肪酸的氧化，并与具有脂肪分解作用的蛋白ES1（esterase1）存在相互作用。
LSDP5的沉默刺激脂肪分解，上调甘油三酯分解速率和脂肪分解相关酶的转录，增加脂肪酸β氧化速率和线粒体的数目。通过酵母双杂交，我们筛选出了与LSDP5相互作用的分子ES1。ES1是一个在肝脏高表达的65KD的糖蛋白，它能够水解许多的酯类，包括甘油三酯和其他的脂肪酸酯。进一步应用免疫共沉淀和免疫荧光共定位，我们证明LSDP5能够与ES1相互作用。
结论：
本研究证实LSDP5是一个定位于脂滴表面的蛋白质，可以促进肝细胞内脂滴的增大和甘油三酯的储积。其机制可能是通过LSDP5与ES1的相互作用，抑制ES1对于甘油三酯的水解，并进一步减少脂肪酸的β氧化来实现的。抑制LSDP5的表达，可以有效减少肝细胞内甘油三酯的储积，因此LSDP5有望成为治疗脂肪肝的新靶点。
Background：
Liver steatosis or fatty liver which is believed to be caused by abnormalmetabolism of fats is one of the most common metabolic disorders in China.Nearly Chinese20-30%people have fatty livers. The typical pathological featureof liver steatosis is the lipid deposition in liver cells in the form of lipid droplets(LDs). With regard to etiology analysis, the researches on metabolism andformation of lipid droplets are involved little. Lipid droplets are neutral lipidstorage surrounded by a phospholipid monolayer, which are now recognized tobe functional subcellular organelles rather than metabolically inactive lipiddepots. They are involved in multiple intracellular processes including lipidmetabolism, vesicle traffic, and cell signaling through interactions with otherorganelles so that important roles in lipid homeostasis are likely. In mammaliancells, LDs contain a class of proteins in their surface layers that contribute to thelipolysis, synthesis and fusion of LDs and share a homologous sequence calledthe PAT domain, including perilipin, adipophilin/adipose differentiation-related protein (ADRP), a tail-interacting protein of47kDa (TIP47), and S3-12. Lipidstorage droplet protein5(LSDP5) is a newly identified member of PAT family. Initialidentifications and characterizations of LSDP5as a lipid droplet binding protein werereported by three independent groups as myocardial lipid droplet protein (MLDP),oxidative tissue-enriched PAT protein (OXPAT), and LSDP5respectively. All thesereports are in agreement that LSDP5is expressed in tissues that exhibit high levels offatty acid oxidation, including heart, skeletal muscle, and liver. However, as a newmember of PAT family, the exact function of LSDP5has not been elucidated so far.
Objective:
We initiate the current study to determine the subcellular localization ofLSDP5protein and to further characterize the LD-localization signal sequenceswithin LSDP5. We investigate the effects of LSDP5on the numbers and sizes ofLD, synthesis and hydrolysis of triglycerides (TG). We indentify theLSDP5-interacting proteins and facilitate the illustration of the molecularmechanism of LSDP5.
Methods:
1) Immunofluorescence was used to detect the the distribution and subcellularlocalization of LSDP5protein.
2) The biological functions of LSDP5were observed by upregulating anddownregulating the expression of LSDP5in hepatic cell line AML12and
primary mouse hepatocytes.
3) The interacting-proteins of LSDP5were screened through the mouse livercDNA library with the prey protein LSDP5using yeast two-hybrid.Co-immunoprecipitation and subcellular localization of LSDP5and the interacting-protein were employed to confirm the specificity of theinteractions.
Results:
1) LSDP5protein was located in the surface of lipid droplets.By using immunofluorescence assay, we observed that LSDP5was localizedto the surface of lipid droplets in hepatocytes. Serial deletion analyses showedthat the lipid droplet targeting domain and the domain directing lipid dropletclustering were overlapped and localized to the188amino acid residues at theN-terminal region of LSDP5.
2) LSDP5enhances triglyceride storage in hepatocytes.Overexpression of LSDP5could enhance lipid accumulation in primaryhepatocytes and hepatic cell line AML12. Moreover, knock-down of LSDP5significantly decreased the size of lipid droplets and reduced the contents oftriglyceride.
3) LSDP5could inhibit lipolysis and interacted with esterase1(ES1).Knock-down of LSDP5significantly stimulated lipolysis in cells, and furtherincreased fatty acid β-oxidation and mitochondrial number. Real-time PCRanalysis revealed that the depletion of LSDP5was associated with increasedmRNA level of enzymes in lipolysis and fatty acid oxidation. In addition, weperformed yeast two-hybrid using LSDP5as a bait protein. Among all thesepositive clones we got, ES1was choosed for the further investigation. ES1isa65-kDa glycoprotein present in liver able to hydrolyze a variety of estersincluding triglycerides, cholesteryl esters and fatty acid esters.Co-immunoprecipitation and lmmunofluorescent co-localization furtherlyproved that LSDP5and ES1possessed physical interactions.
Conclusions
Our data clearly demonstrate that LSDP5, a novel lipid droplet protein,contributes to triglyceride accumulation by negatively regulating lipolysis andfatty acid oxidation in hepatocytes may be through mechanisms of the interactionof LSDP5and ES1. This study makes an experimental foundation for the role andmechanism of LSDP5in lipid metabolism and may provide new thinking for thetreatment of fatty liver.
引文
1. Meigs, J. B.2002. Epidemiology of the metabolic syndrome,2002. Am JManag Care8: S283-292; quiz S293-286.
2. WHO.2000. Obesity: preventing and managing the global epidemic. Reportof a WHO consultation. World Health Organ Tech Rep Ser894: i-xii,1-253.
3. Rudolph, B., and Y. Rivas.2010. Nonalcoholic fatty liver disease. PediatrRev31: e52-53.
4. Brunt, E. M., and D. G. Tiniakos.2010. Histopathology of nonalcoholic fattyliver disease. World J Gastroenterol16:5286-5296.
5. Zafrani, E. S.2004. Non-alcoholic fatty liver disease: an emergingpathological spectrum. Virchows Arch444:3-12.
6. Digel, M., R. Ehehalt, and J. Fullekrug.2010. Lipid droplets lighting up:insights from live microscopy. FEBS Lett584:2168-2175.
7. Guo, Y., K. R. Cordes, R. V. Farese, Jr., and T. C. Walther.2009. Lipiddroplets at a glance. J Cell Sci122:749-752.
8. Tauchi-Sato, K., S. Ozeki, T. Houjou, R. Taguchi, and T. Fujimoto.2002. Thesurface of lipid droplets is a phospholipid monolayer with a unique Fatty Acidcomposition. J Biol Chem277:44507-44512.
9. Murphy, D. J., and J. Vance.1999. Mechanisms of lipid-body formation.Trends Biochem Sci24:109-115.
10. Murphy, D. J.2001. The biogenesis and functions of lipid bodies in animals,plants and microorganisms. Prog Lipid Res40:325-438.
11. Thiele, C., and J. Spandl.2008. Cell biology of lipid droplets. Curr Opin CellBiol20:378-385.
12. Bartz, R., W. H. Li, B. Venables, J. K. Zehmer, M. R. Roth, R. Welti, R. G.Anderson, P. Liu, and K. D. Chapman.2007. Lipidomics reveals that adiposomesstore ether lipids and mediate phospholipid traffic. J Lipid Res48:837-847.
13. Bartz, R., J. K. Zehmer, M. Zhu, Y. Chen, G. Serrero, Y. Zhao, and P. Liu.
2007. Dynamic activity of lipid droplets: protein phosphorylation andGTP-mediated protein translocation. J Proteome Res6:3256-3265.
14. Bostrom, P., L. Andersson, M. Rutberg, J. Perman, U. Lidberg, B. R.Johansson, J. Fernandez-Rodriguez, J. Ericson, T. Nilsson, J. Boren, and S. O.Olofsson.2007. SNARE proteins mediate fusion between cytosolic lipid dropletsand are implicated in insulin sensitivity. Nat Cell Biol9:1286-1293.
15. Brasaemle, D. L.2007. Thematic review series: adipocyte biology. Theperilipin family of structural lipid droplet proteins: stabilization of lipid dropletsand control of lipolysis. J Lipid Res48:2547-2559.
16. Kuerschner, L., C. Moessinger, and C. Thiele.2008. Imaging of lipidbiosynthesis: how a neutral lipid enters lipid droplets. Traffic9:338-352.
17. Martin, S., and R. G. Parton.2006. Lipid droplets: a unified view of adynamic organelle. Nat Rev Mol Cell Biol7:373-378.
18. Stone, S. J., M. C. Levin, and R. V. Farese, Jr.2006. Membrane topology andidentification of key functional amino acid residues of murineacyl-CoA:diacylglycerol acyltransferase-2. J Biol Chem281:40273-40282.
19. Zehmer, J. K., R. Bartz, P. Liu, and R. G. Anderson.2008. Identification of anovel N-terminal hydrophobic sequence that targets proteins to lipid droplets. JCell Sci121:1852-1860.
20. Robenek, H., M. J. Robenek, and D. Troyer.2005. PAT family proteinspervade lipid droplet cores. J Lipid Res46:1331-1338.
21. Ehehalt, R., J. Fullekrug, J. Pohl, A. Ring, T. Herrmann, and W. Stremmel.
2006. Translocation of long chain fatty acids across the plasma membrane--lipidrafts and fatty acid transport proteins. Mol Cell Biochem284:135-140.
22. Ploegh, H. L.2007. A lipid-based model for the creation of an escape hatchfrom the endoplasmic reticulum. Nature448:435-438.
23. Walther, T. C., and R. V. Farese, Jr.2009. The life of lipid droplets. BiochimBiophys Acta1791:459-466.
24. Fei, W., G. Shui, B. Gaeta, X. Du, L. Kuerschner, P. Li, A. J. Brown, M. R.Wenk, R. G. Parton, and H. Yang.2008. Fld1p, a functional homologue of humanseipin, regulates the size of lipid droplets in yeast. J Cell Biol180:473-482.
25. Szymanski, K. M., D. Binns, R. Bartz, N. V. Grishin, W. P. Li, A. K. Agarwal,A. Garg, R. G. Anderson, and J. M. Goodman.2007. The lipodystrophy proteinseipin is found at endoplasmic reticulum lipid droplet junctions and is importantfor droplet morphology. Proc Natl Acad Sci U S A104:20890-20895.
26. Fujimoto, T., Y. Ohsaki, J. Cheng, M. Suzuki, and Y. Shinohara.2008. Lipiddroplets: a classic organelle with new outfits. Histochem Cell Biol130:263-279.
27. Brown, D. A.2001. Lipid droplets: proteins floating on a pool of fat. CurrBiol11: R446-449.
28. Guo, Y., T. C. Walther, M. Rao, N. Stuurman, G. Goshima, K. Terayama, J. S.Wong, R. D. Vale, P. Walter, and R. V. Farese.2008. Functional genomic screenreveals genes involved in lipid-droplet formation and utilization. Nature453:657-661.
29. Olofsson, S. O., P. Bostrom, L. Andersson, M. Rutberg, M. Levin, J. Perman,and J. Boren.2008. Triglyceride containing lipid droplets and lipiddroplet-associated proteins. Curr Opin Lipidol19:441-447.
30. Ducharme, N. A., and P. E. Bickel.2008. Lipid droplets in lipogenesis andlipolysis. Endocrinology149:942-949.
31. Zechner, R., J. G. Strauss, G. Haemmerle, A. Lass, and R. Zimmermann.
2005. Lipolysis: pathway under construction. Curr Opin Lipidol16:333-340.
32. Yamaguchi, T., N. Omatsu, E. Morimoto, H. Nakashima, K. Ueno, T. Tanaka,K. Satouchi, F. Hirose, and T. Osumi.2007. CGI-58facilitates lipolysis on lipiddroplets but is not involved in the vesiculation of lipid droplets caused byhormonal stimulation. J Lipid Res48:1078-1089.
33. Igal, R. A., and R. A. Coleman.1998. Neutral lipid storage disease: a geneticdisorder with abnormalities in the regulation of phospholipid metabolism. J LipidRes39:31-43.
34. Lass, A., R. Zimmermann, G. Haemmerle, M. Riederer, G. Schoiswohl, M.Schweiger, P. Kienesberger, J. G. Strauss, G. Gorkiewicz, and R. Zechner.2006.Adipose triglyceride lipase-mediated lipolysis of cellular fat stores is activated byCGI-58and defective in Chanarin-Dorfman Syndrome. Cell Metab3:309-319.
35. Marcinkiewicz, A., D. Gauthier, A. Garcia, and D. L. Brasaemle.2006. Thephosphorylation of serine492of perilipin a directs lipid droplet fragmentationand dispersion. J Biol Chem281:11901-11909.
36. Goodman, J. M.2008. The gregarious lipid droplet. J Biol Chem283:28005-28009.
37. Murphy, S., S. Martin, and R. G. Parton.2009. Lipid droplet-organelleinteractions; sharing the fats. Biochim Biophys Acta1791:441-447.
38. Liu, P., R. Bartz, J. K. Zehmer, Y. Ying, and R. G. Anderson.2008.Rab-regulated membrane traffic between adiposomes and multipleendomembrane systems. Methods Enzymol439:327-337.
39. Ohsaki, Y., J. Cheng, A. Fujita, T. Tokumoto, and T. Fujimoto.2006.Cytoplasmic lipid droplets are sites of convergence of proteasomal andautophagic degradation of apolipoprotein B. Mol Biol Cell17:2674-2683.
40. Welte, M. A.2007. Proteins under new management: lipid droplets deliver.Trends Cell Biol17:363-369.
41. Cocchiaro, J. L., Y. Kumar, E. R. Fischer, T. Hackstadt, and R. H. Valdivia.
2008. Cytoplasmic lipid droplets are translocated into the lumen of theChlamydia trachomatis parasitophorous vacuole. Proc Natl Acad Sci U S A105:9379-9384.
42. Kumar, Y., J. Cocchiaro, and R. H. Valdivia.2006. The obligate intracellularpathogen Chlamydia trachomatis targets host lipid droplets. Curr Biol16:1646-1651.
43. Miyanari, Y., K. Atsuzawa, N. Usuda, K. Watashi, T. Hishiki, M. Zayas, R.Bartenschlager, T. Wakita, M. Hijikata, and K. Shimotohno.2007. The lipiddroplet is an important organelle for hepatitis C virus production. Nat Cell Biol9:1089-1097.
44. Greenberg, A. S., J. J. Egan, S. A. Wek, N. B. Garty, E. J. Blanchette-Mackie,and C. Londos.1991. Perilipin, a major hormonally regulated adipocyte-specificphosphoprotein associated with the periphery of lipid storage droplets. J BiolChem266:11341-11346.
45. Miura, S., J. W. Gan, J. Brzostowski, M. J. Parisi, C. J. Schultz, C. Londos, B.Oliver, and A. R. Kimmel.2002. Functional conservation for lipid storage dropletassociation among Perilipin, ADRP, and TIP47(PAT)-related proteins inmammals, Drosophila, and Dictyostelium. J Biol Chem277:32253-32257.
46. Brasaemle, D. L., T. Barber, N. E. Wolins, G. Serrero, E. J.Blanchette-Mackie, and C. Londos.1997. Adipose differentiation-related proteinis an ubiquitously expressed lipid storage droplet-associated protein. J Lipid Res38:2249-2263.
47. Brasaemle, D. L., G. Dolios, L. Shapiro, and R. Wang.2004. Proteomicanalysis of proteins associated with lipid droplets of basal and lipolyticallystimulated3T3-L1adipocytes. J Biol Chem279:46835-46842.
48. Cermelli, S., Y. Guo, S. P. Gross, and M. A. Welte.2006. The lipid-dropletproteome reveals that droplets are a protein-storage depot. Curr Biol16:1783-1795.
49. Cho, S. Y., E. S. Shin, P. J. Park, D. W. Shin, H. K. Chang, D. Kim, H. H.Lee, J. H. Lee, S. H. Kim, M. J. Song, I. S. Chang, O. S. Lee, and T. R. Lee.2007.Identification of mouse Prp19p as a lipid droplet-associated protein and itspossible involvement in the biogenesis of lipid droplets. J Biol Chem282:2456-2465.
50. Umlauf, E., E. Csaszar, M. Moertelmaier, G. J. Schuetz, R. G. Parton, and R.Prohaska.2004. Association of stomatin with lipid bodies. J Biol Chem279:23699-23709.
51. Jiang, H., J. He, S. Pu, C. Tang, and G. Xu.2007. Heat shock protein70istranslocated to lipid droplets in rat adipocytes upon heat stimulation. BiochimBiophys Acta1771:66-74.
52. Litvak, V., Y. D. Shaul, M. Shulewitz, R. Amarilio, S. Carmon, and S. Lev.
2002. Targeting of Nir2to lipid droplets is regulated by a specific threonineresidue within its PI-transfer domain. Curr Biol12:1513-1518.
53. Bickel, P. E., J. T. Tansey, and M. A. Welte.2009. PAT proteins, an ancientfamily of lipid droplet proteins that regulate cellular lipid stores. Biochim BiophysActa1791:419-440.
54. Wolins, N. E., B. K. Quaynor, J. R. Skinner, M. J. Schoenfish, A. Tzekov,and P. E. Bickel.2005. S3-12, Adipophilin, and TIP47package lipid inadipocytes. J Biol Chem280:19146-19155.
55. Olofsson, S. O., P. Bostrom, L. Andersson, M. Rutberg, J. Perman, and J.Boren.2009. Lipid droplets as dynamic organelles connecting storage and effluxof lipids. Biochim Biophys Acta1791:448-458.
56. Londos, C., C. Sztalryd, J. T. Tansey, and A. R. Kimmel.2005. Role of PATproteins in lipid metabolism. Biochimie87:45-49.
57. Lu, X., J. Gruia-Gray, N. G. Copeland, D. J. Gilbert, N. A. Jenkins, C.Londos, and A. R. Kimmel.2001. The murine perilipin gene: the lipiddroplet-associated perilipins derive from tissue-specific, mRNA splice variantsand define a gene family of ancient origin. Mamm Genome12:741-749.
58. Londos, C., J. Gruia-Gray, D. L. Brasaemle, C. M. Rondinone, T. Takeda, N.K. Dwyer, T. Barber, A. R. Kimmel, and E. J. Blanchette-Mackie.1996. Perilipin:possible roles in structure and metabolism of intracellular neutral lipids inadipocytes and steroidogenic cells. Int J Obes Relat Metab Disord20Suppl3:S97-101.
59. Miyoshi, H., J. W. Perfield,2nd, S. C. Souza, W. J. Shen, H. H. Zhang, Z. S.Stancheva, F. B. Kraemer, M. S. Obin, and A. S. Greenberg.2007. Control ofadipose triglyceride lipase action by serine517of perilipin A globally regulatesprotein kinase A-stimulated lipolysis in adipocytes. J Biol Chem282:996-1002.
60. Li, D., Q. Kang, and D. M. Wang.2007. Constitutive coactivator ofperoxisome proliferator-activated receptor (PPARgamma), a novel coactivator ofPPARgamma that promotes adipogenesis. Mol Endocrinol21:2320-2333.
61. Akter, M. H., T. Yamaguchi, F. Hirose, and T. Osumi.2008. Perilipin, acritical regulator of fat storage and breakdown, is a target gene of estrogenreceptor-related receptor alpha. Biochem Biophys Res Commun368:563-568.
62. Sztalryd, C., G. Xu, H. Dorward, J. T. Tansey, J. A. Contreras, A. R. Kimmel,and C. Londos.2003. Perilipin A is essential for the translocation ofhormone-sensitive lipase during lipolytic activation. J Cell Biol161:1093-1103.
63. Miyoshi, H., J. W. Perfield,2nd, M. S. Obin, and A. S. Greenberg.2008.Adipose triglyceride lipase regulates basal lipolysis and lipid droplet size inadipocytes. J Cell Biochem105:1430-1436.
64. Brasaemle, D. L., B. Rubin, I. A. Harten, J. Gruia-Gray, A. R. Kimmel, and C.Londos.2000. Perilipin A increases triacylglycerol storage by decreasing the rateof triacylglycerol hydrolysis. J Biol Chem275:38486-38493.
65. Castro-Chavez, F., V. K. Yechoor, P. K. Saha, J. Martinez-Botas, E. C.Wooten, S. Sharma, P. O'Connell, H. Taegtmeyer, and L. Chan.2003.Coordinated upregulation of oxidative pathways and downregulation of lipidbiosynthesis underlie obesity resistance in perilipin knockout mice: a microarraygene expression profile. Diabetes52:2666-2674.
66. Tansey, J. T., C. Sztalryd, J. Gruia-Gray, D. L. Roush, J. V. Zee, O. Gavrilova,M. L. Reitman, C. X. Deng, C. Li, A. R. Kimmel, and C. Londos.2001. Perilipinablation results in a lean mouse with aberrant adipocyte lipolysis, enhanced leptinproduction, and resistance to diet-induced obesity. Proc Natl Acad Sci U S A98:6494-6499.
67. Gandotra, S., C. Le Dour, W. Bottomley, P. Cervera, P. Giral, Y. Reznik, G.Charpentier, M. Auclair, M. Delepine, I. Barroso, R. K. Semple, M. Lathrop, O.Lascols, J. Capeau, S. O'Rahilly, J. Magre, D. B. Savage, and C. Vigouroux.2011.Perilipin deficiency and autosomal dominant partial lipodystrophy. N Engl J Med364:740-748.
68. Xu, G., C. Sztalryd, X. Lu, J. T. Tansey, J. Gan, H. Dorward, A. R. Kimmel,and C. Londos.2005. Post-translational regulation of adiposedifferentiation-related protein by the ubiquitin/proteasome pathway. J Biol Chem280:42841-42847.
69. Imamura, M., T. Inoguchi, S. Ikuyama, S. Taniguchi, K. Kobayashi, N.Nakashima, and H. Nawata.2002. ADRP stimulates lipid accumulation and lipiddroplet formation in murine fibroblasts. Am J Physiol Endocrinol Metab283:E775-783.
70. Chang, B. H., L. Li, A. Paul, S. Taniguchi, V. Nannegari, W. C. Heird, and L.Chan.2006. Protection against fatty liver but normal adipogenesis in micelacking adipose differentiation-related protein. Mol Cell Biol26:1063-1076.
71. Magnusson, B., L. Asp, P. Bostrom, M. Ruiz, P. Stillemark-Billton, D.Linden, J. Boren, and S. O. Olofsson.2006. Adipocyte differentiation-relatedprotein promotes fatty acid storage in cytosolic triglycerides and inhibitssecretion of very low-density lipoproteins. Arterioscler Thromb Vasc Biol26:1566-1571.
72. Buechler, C., M. Ritter, C. Q. Duong, E. Orso, M. Kapinsky, and G. Schmitz.
2001. Adipophilin is a sensitive marker for lipid loading in human bloodmonocytes. Biochim Biophys Acta1532:97-104.
73. Wang, X., T. J. Reape, X. Li, K. Rayner, C. L. Webb, K. G. Burnand, and P. G.Lysko.1999. Induced expression of adipophilin mRNA in human macrophagesstimulated with oxidized low-density lipoprotein and in atherosclerotic lesions.FEBS Lett462:145-150.
74. Larigauderie, G., C. Furman, M. Jaye, C. Lasselin, C. Copin, J. C. Fruchart, G.Castro, and M. Rouis.2004. Adipophilin enhances lipid accumulation andprevents lipid efflux from THP-1macrophages: potential role in atherogenesis.Arterioscler Thromb Vasc Biol24:504-510.
75. Larigauderie, G., C. Cuaz-Perolin, A. B. Younes, C. Furman, C. Lasselin, C.Copin, M. Jaye, J. C. Fruchart, and M. Rouis.2006. Adipophilin increasestriglyceride storage in human macrophages by stimulation of biosynthesis andinhibition of beta-oxidation. Febs J273:3498-3510.
76. Dalen, K. T., S. M. Ulven, B. M. Arntsen, K. Solaas, and H. I. Nebb.2006.PPARalpha activators and fasting induce the expression of adiposedifferentiation-related protein in liver. J Lipid Res47:931-943.
77. Wolins, N. E., D. L. Brasaemle, and P. E. Bickel.2006. A proposed model offat packaging by exchangeable lipid droplet proteins. FEBS Lett580:5484-5491.
78. Beller, M., D. Riedel, L. Jansch, G. Dieterich, J. Wehland, H. Jackle, and R. P.Kuhnlein.2006. Characterization of the Drosophila lipid droplet subproteome.Mol Cell Proteomics5:1082-1094.
79. Wolins, N. E., J. R. Skinner, M. J. Schoenfish, A. Tzekov, K. G. Bensch, andP. E. Bickel.2003. Adipocyte protein S3-12coats nascent lipid droplets. J BiolChem278:37713-37721.
80. Li, F., Y. Gu, W. Dong, H. Li, L. Zhang, N. Li, W. Li, L. Zhang, Y. Song, L.Jiang, J. Ye, and Q. Li.2010. Cell death-inducing DFF45-like effector, a lipiddroplet-associated protein, might be involved in the differentiation of humanadipocytes. Febs J277:4173-4183.
81. Matsusue, K.2010. A physiological role for fat specific protein27/celldeath-inducing DFF45-like effector C in adipose and liver. Biol Pharm Bull33:346-350.
82. Gong, J., Z. Sun, and P. Li.2009. CIDE proteins and metabolic disorders.Curr Opin Lipidol20:121-126.
83. Dalen, K. T., T. Dahl, E. Holter, B. Arntsen, C. Londos, C. Sztalryd, and H. I.Nebb.2007. LSDP5is a PAT protein specifically expressed in fatty acidoxidizing tissues. Biochim Biophys Acta1771:210-227.
84. Yamaguchi, T., S. Matsushita, K. Motojima, F. Hirose, and T. Osumi.2006.MLDP, a novel PAT family protein localized to lipid droplets and enriched in theheart, is regulated by peroxisome proliferator-activated receptor alpha. J BiolChem281:14232-14240.
85. Wolins, N. E., B. K. Quaynor, J. R. Skinner, A. Tzekov, M. A. Croce, M. C.Gropler, V. Varma, A. Yao-Borengasser, N. Rasouli, P. A. Kern, B. N. Finck, andP. E. Bickel.2006. OXPAT/PAT-1is a PPAR-induced lipid droplet protein thatpromotes fatty acid utilization. Diabetes55:3418-3428.
86. Hall, A. M., E. M. Brunt, Z. Chen, N. Viswakarma, J. K. Reddy, N. E. Wolins,and B. N. Finck.2010. Dynamic and differential regulation of proteins that coatlipid droplets in fatty liver dystrophic mice. J Lipid Res51:554-563.
87. Takahashi, Y., A. Shinoda, J. Inoue, and R. Sato.2010. The gene expressionof the myocardial lipid droplet protein is highly regulated by PPARgamma inadipocytes differentiated from MEFs or SVCs. Biochem Biophys Res Commun399:209-214.
88. Granneman, J. G., H. P. Moore, E. P. Mottillo, and Z. Zhu.2009. Functionalinteractions between Mldp (LSDP5) and Abhd5in the control of intracellularlipid accumulation. J Biol Chem284:3049-3057.
89. Yamaguchi, T.2010. Crucial role of CGI-58/alpha/beta hydrolasedomain-containing protein5in lipid metabolism. Biol Pharm Bull33:342-345.
90. Granneman, J. G., H. P. Moore, E. P. Mottillo, Z. Zhu, and L. Zhou.2011.Interactions of perilipin-5(Plin5) with adipose triglyceride lipase. J Biol Chem286:5126-5135.
91. Wang, H., M. Bell, U. Sreenevasan, H. Hu, J. Liu, K. Dalen, C. Londos, T.Yamaguchi, M. A. Rizzo, R. Coleman, D. Gong, D. Brasaemle, and C. Sztalryd.
2011. Unique regulation of adipose triglyceride lipase (ATGL) by perilipin5, alipid droplet-associated protein. J Biol Chem.
92. Grasselli, E., A. Voci, C. Pesce, L. Canesi, E. Fugassa, G. Gallo, and L.Vergani.2010. PAT protein mRNA expression in primary rat hepatocytes: Effectsof exposure to fatty acids. Int J Mol Med25:505-512.
93. Nakamura, N., and T. Fujimoto.2003. Adipose differentiation-related proteinhas two independent domains for targeting to lipid droplets. Biochem Biophys ResCommun306:333-338.
94. Ohsaki, Y., T. Maeda, M. Maeda, K. Tauchi-Sato, and T. Fujimoto.2006.Recruitment of TIP47to lipid droplets is controlled by the putative hydrophobiccleft. Biochem Biophys Res Commun347:279-287.
95. Bulankina, A. V., A. Deggerich, D. Wenzel, K. Mutenda, J. G. Wittmann, M.G. Rudolph, K. N. Burger, and S. Honing.2009. TIP47functions in thebiogenesis of lipid droplets. J Cell Biol185:641-655.
96. Livak, K. J., and T. D. Schmittgen.2001. Analysis of relative geneexpression data using real-time quantitative PCR and the2(-Delta Delta C(T))Method. Methods25:402-408.
97. Meex, S. J., U. Andreo, J. D. Sparks, and E. A. Fisher.2011. Huh-7orHepG2cells: which is the better model for studying human apolipoprotein-B100assembly and secretion? J Lipid Res52:152-158.
98. Guillou, H., P. G. Martin, and T. Pineau.2008. Transcriptional regulation ofhepatic fatty acid metabolism. Subcell Biochem49:3-47.
99. Wang, S., K. G. Soni, M. Semache, S. Casavant, M. Fortier, L. Pan, and G. A.Mitchell.2008. Lipolysis and the integrated physiology of lipid energymetabolism. Mol Genet Metab95:117-126.
100. Ye, J., J. Z. Li, Y. Liu, X. Li, T. Yang, X. Ma, Q. Li, Z. Yao, and P. Li.2009.Cideb, an ER-and lipid droplet-associated protein, mediates VLDL lipidation andmaturation by interacting with apolipoprotein B. Cell Metab9:177-190.
101. Dolinsky, V. W., D. Gilham, M. Alam, D. E. Vance, and R. Lehner.2004.Triacylglycerol hydrolase: role in intracellular lipid metabolism. Cell Mol Life Sci61:1633-1651.
102. Becker-Follmann, J., A. Gaa, E. Bausch, E. Natt, G. Scherer, and O. vonDeimling.1997. High-resolution mapping of a linkage group on mousechromosome8conserved on human chromosome16Q. Mamm Genome8:172-177.
103. Kadner, S. S., J. Katz, and T. H. Finlay.1992. Esterase-1: developmentalexpression in the mouse and distribution of related proteins in other species. ArchBiochem Biophys296:435-441.
104. Chahinian, H., J. Fantini, N. Garmy, G. Manco, and L. Sarda.2010.Non-lipolytic and lipolytic sequence-related carboxylesterases: a comparativestudy of the structure-function relationships of rabbit liver esterase1and bovinepancreatic bile-salt-activated lipase. Biochim Biophys Acta1801:1195-1204.
105. Rao, M. S., and J. K. Reddy.2004. PPARalpha in the pathogenesis of fattyliver disease. Hepatology40:783-786.
106. Ko, K. W., B. Erickson, and R. Lehner.2009. Es-x/Ces1preventstriacylglycerol accumulation in McArdle-RH7777hepatocytes. Biochim BiophysActa1791:1133-1143.
107. Sanghani, S. P., P. C. Sanghani, M. A. Schiel, and W. F. Bosron.2009.Human carboxylesterases: an update on CES1, CES2and CES3. Protein PeptLett16:1207-1214.
108. Imai, T., M. Imoto, H. Sakamoto, and M. Hashimoto.2005. Identification ofesterases expressed in Caco-2cells and effects of their hydrolyzing activity inpredicting human intestinal absorption. Drug Metab Dispos33:1185-1190.
109. Wei, E., M. Alam, F. Sun, L. B. Agellon, D. E. Vance, and R. Lehner.2007.Apolipoprotein B and triacylglycerol secretion in human triacylglycerolhydrolase transgenic mice. J Lipid Res48:2597-2606.
110. Wei, E., Y. Ben Ali, J. Lyon, H. Wang, R. Nelson, V. W. Dolinsky, J. R.Dyck, G. Mitchell, G. S. Korbutt, and R. Lehner.2010. Loss of TGH/Ces3inmice decreases blood lipids, improves glucose tolerance, and increases energyexpenditure. Cell Metab11:183-193.
111.Wang, H., E. Wei, A. D. Quiroga, X. Sun, N. Touret, and R. Lehner.2010.Altered lipid droplet dynamics in hepatocytes lacking triacylglycerol hydrolaseexpression. Mol Biol Cell21:1991-2000.
112.Gilham, D., M. Alam, W. Gao, D. E. Vance, and R. Lehner.2005.Triacylglycerol hydrolase is localized to the endoplasmic reticulum by an unusualretrieval sequence where it participates in VLDL assembly without utilizingVLDL lipids as substrates. Mol Biol Cell16:984-996.
113.Soni, K. G., R. Lehner, P. Metalnikov, P. O'Donnell, M. Semache, W. Gao, K.Ashman, A. V. Pshezhetsky, and G. A. Mitchell.2004. Carboxylesterase3(EC
3.1.1.1) is a major adipocyte lipase. J Biol Chem279:40683-40689.
2. WHO.2000. Obesity: preventing and managing the global epidemic. Reportof a WHO consultation. World Health Organ Tech Rep Ser894: i-xii,1-253.
3. Rudolph, B., and Y. Rivas.2010. Nonalcoholic fatty liver disease. PediatrRev31: e52-53.
4. Brunt, E. M., and D. G. Tiniakos.2010. Histopathology of nonalcoholic fattyliver disease. World J Gastroenterol16:5286-5296.
5. Zafrani, E. S.2004. Non-alcoholic fatty liver disease: an emergingpathological spectrum. Virchows Arch444:3-12.
6. Digel, M., R. Ehehalt, and J. Fullekrug.2010. Lipid droplets lighting up:insights from live microscopy. FEBS Lett584:2168-2175.
7. Guo, Y., K. R. Cordes, R. V. Farese, Jr., and T. C. Walther.2009. Lipiddroplets at a glance. J Cell Sci122:749-752.
8. Tauchi-Sato, K., S. Ozeki, T. Houjou, R. Taguchi, and T. Fujimoto.2002. Thesurface of lipid droplets is a phospholipid monolayer with a unique Fatty Acidcomposition. J Biol Chem277:44507-44512.
9. Murphy, D. J., and J. Vance.1999. Mechanisms of lipid-body formation.Trends Biochem Sci24:109-115.
10. Murphy, D. J.2001. The biogenesis and functions of lipid bodies in animals,plants and microorganisms. Prog Lipid Res40:325-438.
11. Thiele, C., and J. Spandl.2008. Cell biology of lipid droplets. Curr Opin CellBiol20:378-385.
12. Bartz, R., W. H. Li, B. Venables, J. K. Zehmer, M. R. Roth, R. Welti, R. G.Anderson, P. Liu, and K. D. Chapman.2007. Lipidomics reveals that adiposomesstore ether lipids and mediate phospholipid traffic. J Lipid Res48:837-847.
13. Bartz, R., J. K. Zehmer, M. Zhu, Y. Chen, G. Serrero, Y. Zhao, and P. Liu.
2007. Dynamic activity of lipid droplets: protein phosphorylation andGTP-mediated protein translocation. J Proteome Res6:3256-3265.
14. Bostrom, P., L. Andersson, M. Rutberg, J. Perman, U. Lidberg, B. R.Johansson, J. Fernandez-Rodriguez, J. Ericson, T. Nilsson, J. Boren, and S. O.Olofsson.2007. SNARE proteins mediate fusion between cytosolic lipid dropletsand are implicated in insulin sensitivity. Nat Cell Biol9:1286-1293.
15. Brasaemle, D. L.2007. Thematic review series: adipocyte biology. Theperilipin family of structural lipid droplet proteins: stabilization of lipid dropletsand control of lipolysis. J Lipid Res48:2547-2559.
16. Kuerschner, L., C. Moessinger, and C. Thiele.2008. Imaging of lipidbiosynthesis: how a neutral lipid enters lipid droplets. Traffic9:338-352.
17. Martin, S., and R. G. Parton.2006. Lipid droplets: a unified view of adynamic organelle. Nat Rev Mol Cell Biol7:373-378.
18. Stone, S. J., M. C. Levin, and R. V. Farese, Jr.2006. Membrane topology andidentification of key functional amino acid residues of murineacyl-CoA:diacylglycerol acyltransferase-2. J Biol Chem281:40273-40282.
19. Zehmer, J. K., R. Bartz, P. Liu, and R. G. Anderson.2008. Identification of anovel N-terminal hydrophobic sequence that targets proteins to lipid droplets. JCell Sci121:1852-1860.
20. Robenek, H., M. J. Robenek, and D. Troyer.2005. PAT family proteinspervade lipid droplet cores. J Lipid Res46:1331-1338.
21. Ehehalt, R., J. Fullekrug, J. Pohl, A. Ring, T. Herrmann, and W. Stremmel.
2006. Translocation of long chain fatty acids across the plasma membrane--lipidrafts and fatty acid transport proteins. Mol Cell Biochem284:135-140.
22. Ploegh, H. L.2007. A lipid-based model for the creation of an escape hatchfrom the endoplasmic reticulum. Nature448:435-438.
23. Walther, T. C., and R. V. Farese, Jr.2009. The life of lipid droplets. BiochimBiophys Acta1791:459-466.
24. Fei, W., G. Shui, B. Gaeta, X. Du, L. Kuerschner, P. Li, A. J. Brown, M. R.Wenk, R. G. Parton, and H. Yang.2008. Fld1p, a functional homologue of humanseipin, regulates the size of lipid droplets in yeast. J Cell Biol180:473-482.
25. Szymanski, K. M., D. Binns, R. Bartz, N. V. Grishin, W. P. Li, A. K. Agarwal,A. Garg, R. G. Anderson, and J. M. Goodman.2007. The lipodystrophy proteinseipin is found at endoplasmic reticulum lipid droplet junctions and is importantfor droplet morphology. Proc Natl Acad Sci U S A104:20890-20895.
26. Fujimoto, T., Y. Ohsaki, J. Cheng, M. Suzuki, and Y. Shinohara.2008. Lipiddroplets: a classic organelle with new outfits. Histochem Cell Biol130:263-279.
27. Brown, D. A.2001. Lipid droplets: proteins floating on a pool of fat. CurrBiol11: R446-449.
28. Guo, Y., T. C. Walther, M. Rao, N. Stuurman, G. Goshima, K. Terayama, J. S.Wong, R. D. Vale, P. Walter, and R. V. Farese.2008. Functional genomic screenreveals genes involved in lipid-droplet formation and utilization. Nature453:657-661.
29. Olofsson, S. O., P. Bostrom, L. Andersson, M. Rutberg, M. Levin, J. Perman,and J. Boren.2008. Triglyceride containing lipid droplets and lipiddroplet-associated proteins. Curr Opin Lipidol19:441-447.
30. Ducharme, N. A., and P. E. Bickel.2008. Lipid droplets in lipogenesis andlipolysis. Endocrinology149:942-949.
31. Zechner, R., J. G. Strauss, G. Haemmerle, A. Lass, and R. Zimmermann.
2005. Lipolysis: pathway under construction. Curr Opin Lipidol16:333-340.
32. Yamaguchi, T., N. Omatsu, E. Morimoto, H. Nakashima, K. Ueno, T. Tanaka,K. Satouchi, F. Hirose, and T. Osumi.2007. CGI-58facilitates lipolysis on lipiddroplets but is not involved in the vesiculation of lipid droplets caused byhormonal stimulation. J Lipid Res48:1078-1089.
33. Igal, R. A., and R. A. Coleman.1998. Neutral lipid storage disease: a geneticdisorder with abnormalities in the regulation of phospholipid metabolism. J LipidRes39:31-43.
34. Lass, A., R. Zimmermann, G. Haemmerle, M. Riederer, G. Schoiswohl, M.Schweiger, P. Kienesberger, J. G. Strauss, G. Gorkiewicz, and R. Zechner.2006.Adipose triglyceride lipase-mediated lipolysis of cellular fat stores is activated byCGI-58and defective in Chanarin-Dorfman Syndrome. Cell Metab3:309-319.
35. Marcinkiewicz, A., D. Gauthier, A. Garcia, and D. L. Brasaemle.2006. Thephosphorylation of serine492of perilipin a directs lipid droplet fragmentationand dispersion. J Biol Chem281:11901-11909.
36. Goodman, J. M.2008. The gregarious lipid droplet. J Biol Chem283:28005-28009.
37. Murphy, S., S. Martin, and R. G. Parton.2009. Lipid droplet-organelleinteractions; sharing the fats. Biochim Biophys Acta1791:441-447.
38. Liu, P., R. Bartz, J. K. Zehmer, Y. Ying, and R. G. Anderson.2008.Rab-regulated membrane traffic between adiposomes and multipleendomembrane systems. Methods Enzymol439:327-337.
39. Ohsaki, Y., J. Cheng, A. Fujita, T. Tokumoto, and T. Fujimoto.2006.Cytoplasmic lipid droplets are sites of convergence of proteasomal andautophagic degradation of apolipoprotein B. Mol Biol Cell17:2674-2683.
40. Welte, M. A.2007. Proteins under new management: lipid droplets deliver.Trends Cell Biol17:363-369.
41. Cocchiaro, J. L., Y. Kumar, E. R. Fischer, T. Hackstadt, and R. H. Valdivia.
2008. Cytoplasmic lipid droplets are translocated into the lumen of theChlamydia trachomatis parasitophorous vacuole. Proc Natl Acad Sci U S A105:9379-9384.
42. Kumar, Y., J. Cocchiaro, and R. H. Valdivia.2006. The obligate intracellularpathogen Chlamydia trachomatis targets host lipid droplets. Curr Biol16:1646-1651.
43. Miyanari, Y., K. Atsuzawa, N. Usuda, K. Watashi, T. Hishiki, M. Zayas, R.Bartenschlager, T. Wakita, M. Hijikata, and K. Shimotohno.2007. The lipiddroplet is an important organelle for hepatitis C virus production. Nat Cell Biol9:1089-1097.
44. Greenberg, A. S., J. J. Egan, S. A. Wek, N. B. Garty, E. J. Blanchette-Mackie,and C. Londos.1991. Perilipin, a major hormonally regulated adipocyte-specificphosphoprotein associated with the periphery of lipid storage droplets. J BiolChem266:11341-11346.
45. Miura, S., J. W. Gan, J. Brzostowski, M. J. Parisi, C. J. Schultz, C. Londos, B.Oliver, and A. R. Kimmel.2002. Functional conservation for lipid storage dropletassociation among Perilipin, ADRP, and TIP47(PAT)-related proteins inmammals, Drosophila, and Dictyostelium. J Biol Chem277:32253-32257.
46. Brasaemle, D. L., T. Barber, N. E. Wolins, G. Serrero, E. J.Blanchette-Mackie, and C. Londos.1997. Adipose differentiation-related proteinis an ubiquitously expressed lipid storage droplet-associated protein. J Lipid Res38:2249-2263.
47. Brasaemle, D. L., G. Dolios, L. Shapiro, and R. Wang.2004. Proteomicanalysis of proteins associated with lipid droplets of basal and lipolyticallystimulated3T3-L1adipocytes. J Biol Chem279:46835-46842.
48. Cermelli, S., Y. Guo, S. P. Gross, and M. A. Welte.2006. The lipid-dropletproteome reveals that droplets are a protein-storage depot. Curr Biol16:1783-1795.
49. Cho, S. Y., E. S. Shin, P. J. Park, D. W. Shin, H. K. Chang, D. Kim, H. H.Lee, J. H. Lee, S. H. Kim, M. J. Song, I. S. Chang, O. S. Lee, and T. R. Lee.2007.Identification of mouse Prp19p as a lipid droplet-associated protein and itspossible involvement in the biogenesis of lipid droplets. J Biol Chem282:2456-2465.
50. Umlauf, E., E. Csaszar, M. Moertelmaier, G. J. Schuetz, R. G. Parton, and R.Prohaska.2004. Association of stomatin with lipid bodies. J Biol Chem279:23699-23709.
51. Jiang, H., J. He, S. Pu, C. Tang, and G. Xu.2007. Heat shock protein70istranslocated to lipid droplets in rat adipocytes upon heat stimulation. BiochimBiophys Acta1771:66-74.
52. Litvak, V., Y. D. Shaul, M. Shulewitz, R. Amarilio, S. Carmon, and S. Lev.
2002. Targeting of Nir2to lipid droplets is regulated by a specific threonineresidue within its PI-transfer domain. Curr Biol12:1513-1518.
53. Bickel, P. E., J. T. Tansey, and M. A. Welte.2009. PAT proteins, an ancientfamily of lipid droplet proteins that regulate cellular lipid stores. Biochim BiophysActa1791:419-440.
54. Wolins, N. E., B. K. Quaynor, J. R. Skinner, M. J. Schoenfish, A. Tzekov,and P. E. Bickel.2005. S3-12, Adipophilin, and TIP47package lipid inadipocytes. J Biol Chem280:19146-19155.
55. Olofsson, S. O., P. Bostrom, L. Andersson, M. Rutberg, J. Perman, and J.Boren.2009. Lipid droplets as dynamic organelles connecting storage and effluxof lipids. Biochim Biophys Acta1791:448-458.
56. Londos, C., C. Sztalryd, J. T. Tansey, and A. R. Kimmel.2005. Role of PATproteins in lipid metabolism. Biochimie87:45-49.
57. Lu, X., J. Gruia-Gray, N. G. Copeland, D. J. Gilbert, N. A. Jenkins, C.Londos, and A. R. Kimmel.2001. The murine perilipin gene: the lipiddroplet-associated perilipins derive from tissue-specific, mRNA splice variantsand define a gene family of ancient origin. Mamm Genome12:741-749.
58. Londos, C., J. Gruia-Gray, D. L. Brasaemle, C. M. Rondinone, T. Takeda, N.K. Dwyer, T. Barber, A. R. Kimmel, and E. J. Blanchette-Mackie.1996. Perilipin:possible roles in structure and metabolism of intracellular neutral lipids inadipocytes and steroidogenic cells. Int J Obes Relat Metab Disord20Suppl3:S97-101.
59. Miyoshi, H., J. W. Perfield,2nd, S. C. Souza, W. J. Shen, H. H. Zhang, Z. S.Stancheva, F. B. Kraemer, M. S. Obin, and A. S. Greenberg.2007. Control ofadipose triglyceride lipase action by serine517of perilipin A globally regulatesprotein kinase A-stimulated lipolysis in adipocytes. J Biol Chem282:996-1002.
60. Li, D., Q. Kang, and D. M. Wang.2007. Constitutive coactivator ofperoxisome proliferator-activated receptor (PPARgamma), a novel coactivator ofPPARgamma that promotes adipogenesis. Mol Endocrinol21:2320-2333.
61. Akter, M. H., T. Yamaguchi, F. Hirose, and T. Osumi.2008. Perilipin, acritical regulator of fat storage and breakdown, is a target gene of estrogenreceptor-related receptor alpha. Biochem Biophys Res Commun368:563-568.
62. Sztalryd, C., G. Xu, H. Dorward, J. T. Tansey, J. A. Contreras, A. R. Kimmel,and C. Londos.2003. Perilipin A is essential for the translocation ofhormone-sensitive lipase during lipolytic activation. J Cell Biol161:1093-1103.
63. Miyoshi, H., J. W. Perfield,2nd, M. S. Obin, and A. S. Greenberg.2008.Adipose triglyceride lipase regulates basal lipolysis and lipid droplet size inadipocytes. J Cell Biochem105:1430-1436.
64. Brasaemle, D. L., B. Rubin, I. A. Harten, J. Gruia-Gray, A. R. Kimmel, and C.Londos.2000. Perilipin A increases triacylglycerol storage by decreasing the rateof triacylglycerol hydrolysis. J Biol Chem275:38486-38493.
65. Castro-Chavez, F., V. K. Yechoor, P. K. Saha, J. Martinez-Botas, E. C.Wooten, S. Sharma, P. O'Connell, H. Taegtmeyer, and L. Chan.2003.Coordinated upregulation of oxidative pathways and downregulation of lipidbiosynthesis underlie obesity resistance in perilipin knockout mice: a microarraygene expression profile. Diabetes52:2666-2674.
66. Tansey, J. T., C. Sztalryd, J. Gruia-Gray, D. L. Roush, J. V. Zee, O. Gavrilova,M. L. Reitman, C. X. Deng, C. Li, A. R. Kimmel, and C. Londos.2001. Perilipinablation results in a lean mouse with aberrant adipocyte lipolysis, enhanced leptinproduction, and resistance to diet-induced obesity. Proc Natl Acad Sci U S A98:6494-6499.
67. Gandotra, S., C. Le Dour, W. Bottomley, P. Cervera, P. Giral, Y. Reznik, G.Charpentier, M. Auclair, M. Delepine, I. Barroso, R. K. Semple, M. Lathrop, O.Lascols, J. Capeau, S. O'Rahilly, J. Magre, D. B. Savage, and C. Vigouroux.2011.Perilipin deficiency and autosomal dominant partial lipodystrophy. N Engl J Med364:740-748.
68. Xu, G., C. Sztalryd, X. Lu, J. T. Tansey, J. Gan, H. Dorward, A. R. Kimmel,and C. Londos.2005. Post-translational regulation of adiposedifferentiation-related protein by the ubiquitin/proteasome pathway. J Biol Chem280:42841-42847.
69. Imamura, M., T. Inoguchi, S. Ikuyama, S. Taniguchi, K. Kobayashi, N.Nakashima, and H. Nawata.2002. ADRP stimulates lipid accumulation and lipiddroplet formation in murine fibroblasts. Am J Physiol Endocrinol Metab283:E775-783.
70. Chang, B. H., L. Li, A. Paul, S. Taniguchi, V. Nannegari, W. C. Heird, and L.Chan.2006. Protection against fatty liver but normal adipogenesis in micelacking adipose differentiation-related protein. Mol Cell Biol26:1063-1076.
71. Magnusson, B., L. Asp, P. Bostrom, M. Ruiz, P. Stillemark-Billton, D.Linden, J. Boren, and S. O. Olofsson.2006. Adipocyte differentiation-relatedprotein promotes fatty acid storage in cytosolic triglycerides and inhibitssecretion of very low-density lipoproteins. Arterioscler Thromb Vasc Biol26:1566-1571.
72. Buechler, C., M. Ritter, C. Q. Duong, E. Orso, M. Kapinsky, and G. Schmitz.
2001. Adipophilin is a sensitive marker for lipid loading in human bloodmonocytes. Biochim Biophys Acta1532:97-104.
73. Wang, X., T. J. Reape, X. Li, K. Rayner, C. L. Webb, K. G. Burnand, and P. G.Lysko.1999. Induced expression of adipophilin mRNA in human macrophagesstimulated with oxidized low-density lipoprotein and in atherosclerotic lesions.FEBS Lett462:145-150.
74. Larigauderie, G., C. Furman, M. Jaye, C. Lasselin, C. Copin, J. C. Fruchart, G.Castro, and M. Rouis.2004. Adipophilin enhances lipid accumulation andprevents lipid efflux from THP-1macrophages: potential role in atherogenesis.Arterioscler Thromb Vasc Biol24:504-510.
75. Larigauderie, G., C. Cuaz-Perolin, A. B. Younes, C. Furman, C. Lasselin, C.Copin, M. Jaye, J. C. Fruchart, and M. Rouis.2006. Adipophilin increasestriglyceride storage in human macrophages by stimulation of biosynthesis andinhibition of beta-oxidation. Febs J273:3498-3510.
76. Dalen, K. T., S. M. Ulven, B. M. Arntsen, K. Solaas, and H. I. Nebb.2006.PPARalpha activators and fasting induce the expression of adiposedifferentiation-related protein in liver. J Lipid Res47:931-943.
77. Wolins, N. E., D. L. Brasaemle, and P. E. Bickel.2006. A proposed model offat packaging by exchangeable lipid droplet proteins. FEBS Lett580:5484-5491.
78. Beller, M., D. Riedel, L. Jansch, G. Dieterich, J. Wehland, H. Jackle, and R. P.Kuhnlein.2006. Characterization of the Drosophila lipid droplet subproteome.Mol Cell Proteomics5:1082-1094.
79. Wolins, N. E., J. R. Skinner, M. J. Schoenfish, A. Tzekov, K. G. Bensch, andP. E. Bickel.2003. Adipocyte protein S3-12coats nascent lipid droplets. J BiolChem278:37713-37721.
80. Li, F., Y. Gu, W. Dong, H. Li, L. Zhang, N. Li, W. Li, L. Zhang, Y. Song, L.Jiang, J. Ye, and Q. Li.2010. Cell death-inducing DFF45-like effector, a lipiddroplet-associated protein, might be involved in the differentiation of humanadipocytes. Febs J277:4173-4183.
81. Matsusue, K.2010. A physiological role for fat specific protein27/celldeath-inducing DFF45-like effector C in adipose and liver. Biol Pharm Bull33:346-350.
82. Gong, J., Z. Sun, and P. Li.2009. CIDE proteins and metabolic disorders.Curr Opin Lipidol20:121-126.
83. Dalen, K. T., T. Dahl, E. Holter, B. Arntsen, C. Londos, C. Sztalryd, and H. I.Nebb.2007. LSDP5is a PAT protein specifically expressed in fatty acidoxidizing tissues. Biochim Biophys Acta1771:210-227.
84. Yamaguchi, T., S. Matsushita, K. Motojima, F. Hirose, and T. Osumi.2006.MLDP, a novel PAT family protein localized to lipid droplets and enriched in theheart, is regulated by peroxisome proliferator-activated receptor alpha. J BiolChem281:14232-14240.
85. Wolins, N. E., B. K. Quaynor, J. R. Skinner, A. Tzekov, M. A. Croce, M. C.Gropler, V. Varma, A. Yao-Borengasser, N. Rasouli, P. A. Kern, B. N. Finck, andP. E. Bickel.2006. OXPAT/PAT-1is a PPAR-induced lipid droplet protein thatpromotes fatty acid utilization. Diabetes55:3418-3428.
86. Hall, A. M., E. M. Brunt, Z. Chen, N. Viswakarma, J. K. Reddy, N. E. Wolins,and B. N. Finck.2010. Dynamic and differential regulation of proteins that coatlipid droplets in fatty liver dystrophic mice. J Lipid Res51:554-563.
87. Takahashi, Y., A. Shinoda, J. Inoue, and R. Sato.2010. The gene expressionof the myocardial lipid droplet protein is highly regulated by PPARgamma inadipocytes differentiated from MEFs or SVCs. Biochem Biophys Res Commun399:209-214.
88. Granneman, J. G., H. P. Moore, E. P. Mottillo, and Z. Zhu.2009. Functionalinteractions between Mldp (LSDP5) and Abhd5in the control of intracellularlipid accumulation. J Biol Chem284:3049-3057.
89. Yamaguchi, T.2010. Crucial role of CGI-58/alpha/beta hydrolasedomain-containing protein5in lipid metabolism. Biol Pharm Bull33:342-345.
90. Granneman, J. G., H. P. Moore, E. P. Mottillo, Z. Zhu, and L. Zhou.2011.Interactions of perilipin-5(Plin5) with adipose triglyceride lipase. J Biol Chem286:5126-5135.
91. Wang, H., M. Bell, U. Sreenevasan, H. Hu, J. Liu, K. Dalen, C. Londos, T.Yamaguchi, M. A. Rizzo, R. Coleman, D. Gong, D. Brasaemle, and C. Sztalryd.
2011. Unique regulation of adipose triglyceride lipase (ATGL) by perilipin5, alipid droplet-associated protein. J Biol Chem.
92. Grasselli, E., A. Voci, C. Pesce, L. Canesi, E. Fugassa, G. Gallo, and L.Vergani.2010. PAT protein mRNA expression in primary rat hepatocytes: Effectsof exposure to fatty acids. Int J Mol Med25:505-512.
93. Nakamura, N., and T. Fujimoto.2003. Adipose differentiation-related proteinhas two independent domains for targeting to lipid droplets. Biochem Biophys ResCommun306:333-338.
94. Ohsaki, Y., T. Maeda, M. Maeda, K. Tauchi-Sato, and T. Fujimoto.2006.Recruitment of TIP47to lipid droplets is controlled by the putative hydrophobiccleft. Biochem Biophys Res Commun347:279-287.
95. Bulankina, A. V., A. Deggerich, D. Wenzel, K. Mutenda, J. G. Wittmann, M.G. Rudolph, K. N. Burger, and S. Honing.2009. TIP47functions in thebiogenesis of lipid droplets. J Cell Biol185:641-655.
96. Livak, K. J., and T. D. Schmittgen.2001. Analysis of relative geneexpression data using real-time quantitative PCR and the2(-Delta Delta C(T))Method. Methods25:402-408.
97. Meex, S. J., U. Andreo, J. D. Sparks, and E. A. Fisher.2011. Huh-7orHepG2cells: which is the better model for studying human apolipoprotein-B100assembly and secretion? J Lipid Res52:152-158.
98. Guillou, H., P. G. Martin, and T. Pineau.2008. Transcriptional regulation ofhepatic fatty acid metabolism. Subcell Biochem49:3-47.
99. Wang, S., K. G. Soni, M. Semache, S. Casavant, M. Fortier, L. Pan, and G. A.Mitchell.2008. Lipolysis and the integrated physiology of lipid energymetabolism. Mol Genet Metab95:117-126.
100. Ye, J., J. Z. Li, Y. Liu, X. Li, T. Yang, X. Ma, Q. Li, Z. Yao, and P. Li.2009.Cideb, an ER-and lipid droplet-associated protein, mediates VLDL lipidation andmaturation by interacting with apolipoprotein B. Cell Metab9:177-190.
101. Dolinsky, V. W., D. Gilham, M. Alam, D. E. Vance, and R. Lehner.2004.Triacylglycerol hydrolase: role in intracellular lipid metabolism. Cell Mol Life Sci61:1633-1651.
102. Becker-Follmann, J., A. Gaa, E. Bausch, E. Natt, G. Scherer, and O. vonDeimling.1997. High-resolution mapping of a linkage group on mousechromosome8conserved on human chromosome16Q. Mamm Genome8:172-177.
103. Kadner, S. S., J. Katz, and T. H. Finlay.1992. Esterase-1: developmentalexpression in the mouse and distribution of related proteins in other species. ArchBiochem Biophys296:435-441.
104. Chahinian, H., J. Fantini, N. Garmy, G. Manco, and L. Sarda.2010.Non-lipolytic and lipolytic sequence-related carboxylesterases: a comparativestudy of the structure-function relationships of rabbit liver esterase1and bovinepancreatic bile-salt-activated lipase. Biochim Biophys Acta1801:1195-1204.
105. Rao, M. S., and J. K. Reddy.2004. PPARalpha in the pathogenesis of fattyliver disease. Hepatology40:783-786.
106. Ko, K. W., B. Erickson, and R. Lehner.2009. Es-x/Ces1preventstriacylglycerol accumulation in McArdle-RH7777hepatocytes. Biochim BiophysActa1791:1133-1143.
107. Sanghani, S. P., P. C. Sanghani, M. A. Schiel, and W. F. Bosron.2009.Human carboxylesterases: an update on CES1, CES2and CES3. Protein PeptLett16:1207-1214.
108. Imai, T., M. Imoto, H. Sakamoto, and M. Hashimoto.2005. Identification ofesterases expressed in Caco-2cells and effects of their hydrolyzing activity inpredicting human intestinal absorption. Drug Metab Dispos33:1185-1190.
109. Wei, E., M. Alam, F. Sun, L. B. Agellon, D. E. Vance, and R. Lehner.2007.Apolipoprotein B and triacylglycerol secretion in human triacylglycerolhydrolase transgenic mice. J Lipid Res48:2597-2606.
110. Wei, E., Y. Ben Ali, J. Lyon, H. Wang, R. Nelson, V. W. Dolinsky, J. R.Dyck, G. Mitchell, G. S. Korbutt, and R. Lehner.2010. Loss of TGH/Ces3inmice decreases blood lipids, improves glucose tolerance, and increases energyexpenditure. Cell Metab11:183-193.
111.Wang, H., E. Wei, A. D. Quiroga, X. Sun, N. Touret, and R. Lehner.2010.Altered lipid droplet dynamics in hepatocytes lacking triacylglycerol hydrolaseexpression. Mol Biol Cell21:1991-2000.
112.Gilham, D., M. Alam, W. Gao, D. E. Vance, and R. Lehner.2005.Triacylglycerol hydrolase is localized to the endoplasmic reticulum by an unusualretrieval sequence where it participates in VLDL assembly without utilizingVLDL lipids as substrates. Mol Biol Cell16:984-996.
113.Soni, K. G., R. Lehner, P. Metalnikov, P. O'Donnell, M. Semache, W. Gao, K.Ashman, A. V. Pshezhetsky, and G. A. Mitchell.2004. Carboxylesterase3(EC
3.1.1.1) is a major adipocyte lipase. J Biol Chem279:40683-40689.