Uncoupled packaging of amyloid precursor protein and presenilin 1 into coat protein complex II vesicles
Uncoupled packaging of amyloid precursor protein and presenilin 1 into coat protein complex II vesicles
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DOI：
10.1074/jbc.m411091200
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发表时间：
2005-03-04
影响因子：
4.8
通讯作者：
Schekman, R
中科院分区：
文献类型：
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作者：
Kim, J;Hamamoto, S;Schekman, R
早老素（PS） 1和2以及淀粉样前体蛋白（APP）的突变形式导致家族性阿尔茨海默病。一些报道表明，PS可能调节内质网（ER）的APP输出。为了验证这种可能性，我们重建了渗透培养细胞内质网膜形成的COPII（外壳蛋白复合物II）囊泡中APP和PS1的捕获。哺乳动物COPII蛋白的重组形式在化学定义的合成脂质体上测量涂层亚基组装和包被囊泡出芽的反应中具有活性。然而，重组COPII蛋白在微粒体膜的货物捕获和囊泡出芽中没有活性。相比之下，大鼠肝细胞质能积极刺激APP、PS1和p58（一种流动的内质网到高尔基体标记蛋白）从供体内质网膜分选和包装到运输囊泡中。在稀释的细胞质中加入重组COPII蛋白刺激出芽。细胞质溶胶的分离表明，除了COPII亚基外，还有一种或多种其他蛋白质可能对哺乳动物内质网膜的货物选择或囊泡形成至关重要。重组Sec24C特异性识别APP c端区域进行包装。Sarla滴定区分了APP和PS1的包装要求。此外，APP的包装不受PS1或PS1和2的删除的影响，这表明APP和PS1从ER的贩运通常是不耦合的。
Mutant forms of presenilin (PS) 1 and 2 and amyloid precursor protein (APP) lead to familial Alzheimer's disease. Several reports indicate that PS may modulate APP export from the endoplasmic reticulum (ER). To develop a test of this possibility, we reconstituted the capture of APP and PS1 in COPII (coat protein complex II) vesicles formed from ER membranes in permeabilized cultured cells. The recombinant forms of mammalian COPII proteins were active in a reaction that measures coat subunit assembly and coated vesicle budding on chemically defined synthetic liposomes. However, the recombinant COPII proteins were not active in cargo capture and vesicle budding from microsomal membranes. In contrast, rat liver cytosol was active in stimulating the sorting and packaging of APP, PS1, and p58 (an itinerant ER to Golgi marker protein) into transport vesicles from donor ER membranes. Budding was stimulated in dilute cytosol by the addition of recombinant COPII proteins. Fractionation of the cytosol suggested one or more additional proteins other than the COPII subunits may be essential for cargo selection or vesicle formation from the mammalian ER membrane. The recombinant Sec24C specifically recognized the APP C-terminal region for packaging. Titration of Sarla distinguished the packaging requirements of APP and PS1. Furthermore, APP packaging was not affected by deletion of PS1 or PS1 and 2, suggesting APP and PS1 trafficking from the ER are normally uncoupled.