Niemann-Pick C2 protein regulates sterol transport between plasma membrane and late endosomes in human fibroblasts
Niemann-Pick C2 protein regulates sterol transport between plasma membrane and late endosomes in human fibroblasts
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DOI：
10.1016/j.chemphyslip.2018.03.006
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发表时间：
2018-07-01
影响因子：
3.4
通讯作者：
Wustner, Daniel
中科院分区：
文献类型：
--
作者：
Berzina, Zane;Solanko, Lukasz M.;Wustner, Daniel
C2型尼曼-匹克病是一种脂质储存障碍，其中NPC 2蛋白的突变导致脂蛋白衍生的胆固醇在晚期内体和溶酶体（LE/LYSs）中积累。目前尚不清楚通过其他方式递送至NPC2缺陷细胞的胆固醇是否也在LE/LYSs中积累。我们表明，接近胆固醇类似物脱氢麦角固醇（DHE），当交付到质膜（PM）积累在LE/LYSs的人成纤维细胞缺乏功能NPC 2。我们测量了甾醇扩散的两个不同时间尺度;当富含DHE的LE/LYSs在疾病细胞中通过缓慢的异常扩散移动时（D 4.6.10（-4）μ m（2）/sec; α类似于0.76），一小部分甾醇可以在LE/LYSs之间快速交换，D类似于3 μ m（2）/s，如光漂白后荧光恢复（FRAP）所示。通过定量脂质质谱，我们发现，与对照细胞相比，疾病成纤维细胞中C-13标记的胆固醇的酯化而不是DHE的酯化减少了10倍。内化的NPC2挽救了固醇储存表型，并强烈扩大了FRAP实验中观察到的动态固醇库。总之，我们的研究表明，胆固醇酯化和PM和LE/LYSs之间的固醇运输取决于功能性NPC2蛋白。NPC2可能在LE/LYSs内起作用，从那里它增加与其他细胞器的非囊泡固醇交换。
Niemann-Pick disease type C2 is a lipid storage disorder in which mutations in the NPC2 protein cause accumulation of lipoprotein-derived cholesterol in late endosomes and lysosomes (LE/LYSs). Whether cholesterol delivered by other means to NPC2 deficient cells also accumulates in LE/LYSs is currently unknown. We show that the close cholesterol analog dehydroergosterol (DHE), when delivered to the plasma membrane (PM) accumulates in LE/LYSs of human fibroblasts lacking functional NPC2. We measured two different time scales of sterol diffusion; while DHE rich LE/LYSs moved by slow anomalous diffusion in disease cells (D 4.6.10(-4) mu m(2)/ sec; alpha similar to 0.76), a small pool of sterol could exchange rapidly with D similar to 3 mu m(2)/s between LE/LYSs, as shown by fluorescence recovery after photobleaching (FRAP). By quantitative lipid mass spectrometry we found that esterification of C-13-labeled cholesterol but not of DHE is reduced 10-fold in disease fibroblasts compared to control cells. Internalized NPC2 rescued the sterol storage phenotype and strongly expanded the dynamic sterol pool seen in FRAP experiments. Together, our study shows that cholesterol esterification and trafficking of sterols between the PM and LE/LYSs depends on a functional NPC2 protein. NPC2 likely acts inside LE/LYSs from where it increases non-vesicular sterol exchange with other organelles.