Distribution and transport of cholesterol-rich membrane domains monitored by a membrane-impermeant fluorescent polyethylene glycol-derivatized cholesterol
Distribution and transport of cholesterol-rich membrane domains monitored by a membrane-impermeant fluorescent polyethylene glycol-derivatized cholesterol
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DOI：
10.1074/jbc.m313568200
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发表时间：
2004-05-28
影响因子：
4.8
通讯作者：
Kobayashi, T
中科院分区：
文献类型：
--
作者：
Sato, SB;Ishii, K;Kobayashi, T
富含胆固醇的膜结构域在各种膜事件中起作用，如信号转导和膜交通。我们研究了荧光素酯的聚乙二醇衍生的胆固醇（fPEG-Chol）与富含胆固醇的细胞膜和模型膜的相互作用。与菲律宾和其他胆固醇探针不同，这种分子可以作为水分散体应用于各种样品。当添加到活细胞中时，fPEG-Chol仅分布在外质膜小叶中，并富集在通过受体信号传导的激活而动态聚集的微区中。表面结合的fPEG-Chol通过网格蛋白非依赖性途径与脂筏标记物一起缓慢内化到内体中。值得注意的是，fPEG-Chol可以显微注射到活细胞中，我们发现高尔基体作为唯一的主要细胞器被标记。因此，PEG-Chol提供了一种新的，灵敏的探针，用于解开活细胞中富含胆固醇的微区的动力学。
Cholesterol-rich membrane domains function in various membrane events as diverse as signal transduction and membrane traffic. We studied the interaction of a fluorescein ester of polyethylene glycol-derivatized cholesterol (fPEG-Chol) with cholesterol-rich membranes both in cells and in model membranes. Unlike filipin and other cholesterol probes, this molecule could be applied as an aqueous dispersion to various samples. When added to live cells, fPEG-Chol distributed exclusively in the outer plasma membrane leaflet and was enriched in microdomains that dynamically clustered by the activation of receptor signaling. The surface-bound fPEG-Chol was slowly internalized via clathrin-independent pathway into endosomes together with lipid raft markers. Noteworthy, fPEG-Chol could be microinjected in the living cells in which we found Golgi apparatus as the sole major organelle to be labeled. PEG-Chol, thus, provides a novel, sensitive probe for unraveling the dynamics of cholesterol-rich microdomains in living cells.