Association of γ-secretase with lipid rafts in post-golgi and endosome membranes
Association of γ-secretase with lipid rafts in post-golgi and endosome membranes
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DOI：
10.1074/jbc.m407986200
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发表时间：
2004-10-22
影响因子：
4.8
通讯作者：
Thinakaran, G
中科院分区：
文献类型：
--
作者：
Vetrivel, KS;Cheng, HP;Thinakaran, G
阿尔茨海默病相关的β -淀粉样蛋白肽（Abeta）是由β和γ分泌酶对淀粉样蛋白前体蛋白（APP）的顺序蛋白水解加工产生的。越来越多的证据表明，富含胆固醇和鞘脂的膜微域参与调控APP的运输和加工。BACE1是神经元中主要的β -分泌酶，是一种存在于脂筏中的棕榈酰化跨膜蛋白。APP的一个子集在脂筏中受到BACE1的淀粉样变性加工，这一过程取决于脂筏的完整性。在这里，我们描述了γ -分泌酶复合物的所有四种成分，即早老素1 （PS1）衍生片段、成熟nicastrin、APH-1和PEN-2，与非神经元细胞和符合脂筏标准的神经元的富含胆固醇的洗涤不溶性膜（DIM）结构域的关联。在PS1(-/-)/PS2（-/-）和NCT-/-成纤维细胞中，仍然存在的γ -分泌酶成分无法抵抗洗涤剂，这表明筏状结合需要γ -分泌酶复合物的组装。生化证据表明，γ -分泌酶复合物的亚基和三个TGN/内核体居住的SNAREs在蔗糖密度梯度中分解，并在不同的非离子和两性离子洗涤剂中表现出相似的溶解性或不溶解性特征，表明它们以相似的蛋白质-脂质组成共同居住在膜微域中。采用Lubrol WX提取或超声处理后梯度离心，从具有相似浮力密度的膜混合物中对PS1-或syntaxin 6阳性膜片进行磁免疫分离，证实了这一观点。这些发现与γ -分泌酶定位于高尔基体和内体的脂筏微域一致，内体是以前与APP淀粉样变性过程有关的细胞器。
Alzheimer's disease-associated beta-amyloid peptides (Abeta) are generated by the sequential proteolytic processing of amyloid precursor protein (APP) by beta- and gamma-secretases. There is growing evidence that cholesterol- and sphingolipid-rich membrane microdomains are involved in regulating trafficking and processing of APP. BACE1, the major beta-secretase in neurons is a palmitoylated transmembrane protein that resides in lipid rafts. A subset of APP is subject to amyloidogenic processing by BACE1 in lipid rafts, and this process depends on the integrity of lipid rafts. Here we describe the association of all four components of the gamma-secretase complex, namely presenilin 1 (PS1)-derived fragments, mature nicastrin, APH-1, and PEN-2, with cholesterol- rich detergent insoluble membrane (DIM) domains of non-neuronal cells and neurons that fulfill the criteria of lipid rafts. In PS1(-/-)/PS2(-/-) and NCT-/- fibroblasts, gamma-secretase components that still remain fail to become detergent-resistant, suggesting that raft association requires gamma-secretase complex assembly. Biochemical evidence shows that subunits of the gamma-secretase complex and three TGN/endosome-resident SNAREs cofractionate in sucrose density gradients, and show similar solubility or insolubility characteristics in distinct non-ionic and zwitterionic detergents, indicative of their co-residence in membrane microdomains with similar protein-lipid composition. This notion is confirmed using magnetic immunoisolation of PS1- or syntaxin 6-positive membrane patches from a mixture of membranes with similar buoyant densities following Lubrol WX extraction or sonication, and gradient centrifugation. These findings are consistent with the localization of gamma-secretase in lipid raft microdomains of post-Golgi and endosomes, organelles previously implicated in amyloidogenic processing of APP.