Microdomains of GPI-anchored proteins in living cells revealed by crosslinking
Microdomains of GPI-anchored proteins in living cells revealed by crosslinking
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DOI：
10.1038/29570
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发表时间：
1998-08-20
期刊：
影响因子：
64.8
通讯作者：
Kurzchalia, TV
中科院分区：
文献类型：
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作者：
Friedrichson, T;Kurzchalia, TV
关于糖基磷脂酰肌醇(GPI)锚定蛋白是否存在于细胞膜的微区中有一些讨论(1，2)这些假定的微区参与了极化细胞的分选(3-5)和信号转导(6-8)，富含GPI锚定蛋白的复合体，胆固醇和神经节苷脂已通过非离子去污剂从细胞膜中分离：这些复合体被认为代表GPI锚定蛋白的簇状排列(9，10)。然而，当抗体或洗涤剂诱导的GPI锚定蛋白的聚集被阻止时，所获得的结果支持稳定状态下GPI锚定蛋白的分散表面分布的想法(11-13)。在这里，我们使用化学交联法来证明GPI锚定蛋白的膜微域存在于活细胞的表面。这种聚集是GPI锚定形式所特有的，因为两种带有相同胞外结构域的跨膜形式不会形成低聚物。膜胆固醇的耗尽会导致GPI锚定蛋白的聚集破裂，而用洗涤剂处理细胞则大大增加了复合体的大小。我们发现，在活细胞中，这些GPI锚定蛋白存在于至少由15个分子组成的微域中，这些微域比洗涤剂提取后看到的要小得多。
There is some discussion as to whether glycosyl-phosphatidylinositol(GPI)-anchored proteins occur in microdomains in the cell membrane(.1,2) These putative microdomains have been implicated in processes such as sorting in polarized cells(3-5) and signal transduction(6-8), Complexes enriched in GPI-anchored proteins, cholesterol and glycosphingolipids have been isolated from cell membranes by using non-ionic detergents: these complexes were thought to represent a clustered arrangement of GPI-anchored proteins(9,10). However, results obtained when clustering of GPI-anchored proteins induced by antibodies or by detergents was prevented support the idea of a dispersed surface distribution of GPI-anchored proteins at steady state(11-13). Here we use chemical crosslinking to show that membrane microdomains of a GPI-anchored protein exist at the surface in living cells. This clustering is specific for the GPI-anchored form, as two transmembrane forms bearing the same ectodomain do not form oligomers. Depletion of membrane cholesterol causes the clustering of GPI-anchored proteins to break up, whereas treatment of cells with detergent substantially increases the size of the complexes. We find that in living cells these GPI-anchored proteins reside in microdomains consisting of at least 15 molecules, which are much smaller than those seen after detergent extraction.