Localization of Mammalian NAD(P)H Steroid Dehydrogenase-like Protein on Lipid Droplets*
Localization of Mammalian NAD(P)H Steroid Dehydrogenase-like Protein on Lipid Droplets*
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DOI：
10.1074/jbc.m301408200
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发表时间：
2003-09
影响因子：
4.8
通讯作者：
M. Ohashi;N. Mizushima;Y. Kabeya;T. Yoshimori
中科院分区：
文献类型：
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作者：
M. Ohashi;N. Mizushima;Y. Kabeya;T. Yoshimori
哺乳动物胆固醇合成晚期的酶通过酶法均匀分布在内质网上。我们在这里报告了第一个哺乳动物胆固醇生物合成酶，明确地定位在细胞内脂存储液滴的表面。NAD(P)H类固醇脱氢酶类蛋白(Nsdhl)是一种哺乳动物C-3类固醇脱氢酶，参与羊毛甾醇向胆固醇的转化，通过免疫荧光显微镜和亚细胞分级技术定位于脂滴。即使细胞的生长完全依赖于这种酶介导的胆固醇生物合成，Nsdhl也定位于脂滴上。培养基中脂肪酸的耗尽减少了脂滴的形成，并导致Nsdhl重新分布到内质网。在培养液中增加油酸可以诱导发育良好的Nsdhl阳性脂滴，同时导致细胞内羊毛甾醇转化为胆固醇的减少。操纵人类NSDHL的错义突变(G205S)导致人类胚胎发育障碍，先天性半侧发育不良伴鱼鳞状痣和肢体缺陷综合征(CHILD)，不能再定位在脂滴上。虽然野生型NSDHL的表达可以恢复CHO胆固醇缺陷型营养缺陷型细胞LEX2的生长，但NSDHL(G205S)的表达不能恢复该缺陷生长。这些结果表明Nsdh1在脂滴上的定位具有功能意义。Nsdh1与TIP47在脂滴上的共同定位也表明了其功能意义。TIP47是一种货物选择蛋白，可将甘露糖6-磷酸受体从晚期内吞体转至反式高尔基体网络。这些结果增加了越来越多的观点，即脂滴是一种细胞器，在各种生物现象中具有更复杂的作用。
Mammalian enzymes in late cholesterol biosynthesis have been localized uniformly over the endoplasmic reticulum by enzymatic methods. We report here the first mammalian cholesterol biosynthetic enzyme unequivocally localized at the surface of intracellular lipid storage droplets. NAD(P)H steroid dehydrogenase-like protein (Nsdhl), a mammalian C-3 sterol dehydrogenase involved in the conversion of lanosterol into cholesterol, was localized on lipid droplets by immunofluorescence microscopy and subcellular fractionation. Nsdhl was localized on lipid droplets even when cell growth exclusively depended on cholesterol biosynthesis mediated by this enzyme. Depletion of fatty acids in culture medium reduced the development of lipid droplets and caused Nsdhl redistribution to the endoplasmic reticulum. Elevating oleic acid in medium induced well developed, Nsdhl-positive lipid droplets, and simultaneously caused a reduction in cellular conversion of lanosterol into cholesterol. Manipulated human NSDHL with a missense mutation (G205S) causing a human embryonic developmental disorder, congenital hemidysplasia with ichthyosiform nevus and limb defects (CHILD) syndrome, could no longer be localized on lipid droplets. Although the expression of wild-type NSDHL could restore the defective growth of a CHO cholesterol auxotroph, LEX2 in cholesterol-deficient medium, the expression of NSDHL(G205S) failed to do so. These results point to functional significance of the localization of Nsdhl on lipid droplets. Functional significance was also suggested by the colocalization of Nsdhl on lipid droplets with TIP47, a cargo selection protein for mannose 6-phosphate receptors from late endosomes to the trans-Golgi network. These results add to the growing notion that the lipid droplet is an organelle endowed with more complex roles in various biological phenomena.