Detection of cholesterol-rich microdomains in the inner leaflet of the plasma membrane
Detection of cholesterol-rich microdomains in the inner leaflet of the plasma membrane
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DOI：
10.1016/j.bbrc.2006.10.088
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发表时间：
2006-12-22
影响因子：
3.1
通讯作者：
Ohno-Iwashita, Yoshiko
中科院分区：
文献类型：
--
作者：
Hayashi, Masami;Shimada, Yukiko;Ohno-Iwashita, Yoshiko
产气荚膜梭菌溶素0的C末端结构域（D4）选择性地结合富含胆固醇的微结构域中的胆固醇。为了解决富含胆固醇的微区是否存在于质膜的内小叶中的问题，我们在MEF细胞中表达D4作为与EGFP的融合蛋白。在稳定的细胞克隆中表达的EGFP-D4的一半以上结合到筏级分中的膜上。用β-环糊精耗尽膜胆固醇减少了位于筏组分中的EGFP-D4的量，证实EGFP-D4与富含胆固醇的微区结合。筏馏分的亚分级显示大多数的EGFP-D4结合到质膜，而不是细胞内膜。两者合计，这些结果强烈表明存在的富含胆固醇的微区的内小叶的质膜。(c)2006年爱思唯尔公司All rights reserved.
The C-terminal domain (D4) of perfringolysin 0 binds selectively to cholesterol in cholesterol-rich microdomains. To address the issue of whether cholesterol-rich microdomains exist in the inner leaflet of the plasma membrane, we expressed D4 as a fusion protein with EGFP in MEF cells. More than half of the EGFP-D4 expressed in stable cell clones was bound to membranes in raft fractions. Depletion of membrane cholesterol with beta-cyclodextrin reduced the amount of EGFP-D4 localized in raft fractions, confirming EGFP-D4 binding to cholesterol-rich microdomains. Subfractionation of the raft fractions showed most of the EGFP-D4 bound to the plasma membrane rather than to intracellular membranes. Taken together, these results strongly suggest the existence of cholesterol-rich microdomains in the inner leaflet of the plasma membrane. (c) 2006 Elsevier Inc. All rights reserved.