Proteomic analysis of proteins associated with lipid droplets of basal and lipolytically stimulated 3T3-L1 adipocytes
Proteomic analysis of proteins associated with lipid droplets of basal and lipolytically stimulated 3T3-L1 adipocytes
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DOI：
10.1074/jbc.m409340200
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发表时间：
2004-11-05
影响因子：
4.8
通讯作者：
Wang, R
中科院分区：
文献类型：
--
作者：
Brasaemle, DL;Dolios, G;Wang, R
脂肪细胞作为三酰甘油包裹在大的脂滴中，是人体主要的能量储备。周脂蛋白是这些脂滴上最丰富的蛋白质，在促进三酰甘油储存和水解方面起着关键作用。β-肾上腺素能激动剂对脂解的刺激触发了脂周蛋白的快速磷酸化和脂滴敏感性脂肪酶向脂滴表面的易位以及微脂滴的更逐渐的碎裂和分散。由于很少脂滴相关蛋白已被确定在脂肪细胞中，我们分离脂滴从基础和脂解刺激的3 T3-L1脂肪细胞，并确定了组成蛋白质的质谱。在两种制剂中鉴定的结构蛋白包括周脂蛋白、S3-12、波形蛋白和TIP 47;相反，在脂解刺激的细胞中，亲脂蛋白、小窝蛋白-1和微管蛋白选择性地定位于液滴。在两种制剂中鉴定的脂质代谢酶包括脂肪酶敏感性脂肪酶、羊毛甾醇合酶、NAD（P）依赖性类固醇脱氢酶样蛋白、酰基辅酶A合成酶、长链家族成员（ACSL）1和CGI-58。仅在基础制剂中鉴别出7型17-β-羟基类固醇脱氢酶，而在脂解刺激细胞的液滴中鉴别出ACSL 3和4以及两种短链还原酶/脱氢酶。此外，两种制剂均含有FSP 27、核糖核酸酶I、EHD 2、心肌黄酶I和古老的普遍存在的蛋白质。基础制剂含有CGI-49，而来自脂解刺激细胞的脂滴含有几种Rab GTP酶和肿瘤蛋白D54。线粒体与脂滴的密切联系，建议丙酮酸羧化酶，抑制素，和ATP合酶的亚基在制剂中的鉴定。因此，脂肪细胞脂滴含有特定的结构蛋白以及脂质代谢酶;响应脂解的激素刺激的脂滴的结构重组伴随着几种蛋白质的相对质量的增加和额外蛋白质的募集。
Adipocytes hold the body's major energy reserve as triacylglycerols packaged in large lipid droplets. Perilipins, the most abundant proteins on these lipid droplets, play a critical role in facilitating both triacylglycerol storage and hydrolysis. The stimulation of lipolysis by beta-adrenergic agonists triggers rapid phosphorylation of perilipin and translocation of hormone-sensitive lipase to the surfaces of lipid droplets and more gradual fragmentation and dispersion of micro-lipid droplets. Because few lipid droplet-associated proteins have been identified in adipocytes, we isolated lipid droplets from basal and lipolytically stimulated 3T3-L1 adipocytes and identified the component proteins by mass spectrometry. Structural proteins identified in both preparations include perilipin, S3-12, vimentin, and TIP47; in contrast, adipophilin, caveolin-1, and tubulin selectively localized to droplets in lipolytically stimulated cells. Lipid metabolic enzymes identified in both preparations include hormone-sensitive lipase, lanosterol synthase, NAD(P)-dependent steroid dehydrogenase-like protein, acyl-CoA synthetase, long chain family member (ACSL) 1, and CGI-58. 17-beta-Hydroxysteroid dehydrogenase, type 7, was identified only in basal preparations, whereas ACSL3 and 4 and two short-chain reductase/dehydrogenases were identified on droplets from lipolytically stimulated cells. Additionally, both preparations contained FSP27, ribophorin I, EHD2, diaphorase I, and ancient ubiquitous protein. Basal preparations contained CGI-49, whereas lipid droplets from lipolytically stimulated cells contained several Rab GTPases and tumor protein D54. A close association of mitochondria with lipid droplets was suggested by the identification of pyruvate carboxylase, prohibitin, and a subunit of ATP synthase in the preparations. Thus, adipocyte lipid droplets contain specific structural proteins as well as lipid metabolic enzymes; the structural reorganization of lipid droplets in response to the hormonal stimulation of lipolysis is accompanied by increases in the relative mass of several proteins and the recruitment of additional proteins.