Aβ46 Is Processed to Aβ40 and Aβ43, but Not to Aβ42, in the Low Density Membrane Domains*
Aβ46 Is Processed to Aβ40 and Aβ43, but Not to Aβ42, in the Low Density Membrane Domains*
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DOI：
10.1074/jbc.m707103200
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发表时间：
2008-01
影响因子：
4.8
通讯作者：
Sosuke Yagishita;M. Morishima-kawashima;S. Ishiura;Y. Ihara
中科院分区：
文献类型：
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作者：
Sosuke Yagishita;M. Morishima-kawashima;S. Ishiura;Y. Ihara
γ-分泌酶在称为γ-、γ-和γ-切割位点的多个位点切割β-淀粉样前体蛋白的跨膜结构域。我们先前发现，N-[N-（3，5-二氟苯乙酰基）-l-丙氨酰]-S-苯基甘氨酸叔丁酯（DAPT），一种有效的二肽γ-分泌酶抑制剂，导致较长的淀粉样β蛋白（Aβ）在共表达β C-末端片段和野生型早老素1的中国仓鼠卵巢细胞（C99/wtPS 1细胞）中的差异蓄积。在这项研究中，我们使用蔗糖密度梯度离心破碎C99/wtPS 1细胞的膜，已经用DAPT预处理。我们发现Aβ46的聚集只局限于低密度膜（LDM）结构域。在37 °C下孵育Aβ46累积LDM结构域产生Aβ40、Aβ42、Aβ43和β-淀粉样前体蛋白胞内结构域。添加L 685，458完全阻止了β-淀粉样前体蛋白胞内结构域的生成，并导致Aβ46水平大幅降低，同时出现Aβ40和Aβ43，但不出现Aβ42。进一步添加DAPT抑制了Aβ40/43的产生，并消除了Aβ46量的减少。这些数据表明，预先积累的Aβ46被γ-分泌酶加工成Aβ40/43，而不是LDM结构域中的Aβ42。新产生的Aβ40和Aβ43的量大致相当于Aβ46量的减少。时间曲线未显示Aβ43的最大浓度，表明Aβ46通过非连续过程加工为Aβ40和Aβ43。
γ-Secretase cleaves the transmembrane domain of β-amyloid precursor protein at multiple sites referred to as γ-, ϵ-, and ζ-cleavage sites. We previously showed that N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT), a potent dipeptide γ-secretase inhibitor, causes differential accumulation of longer amyloid β-proteins (Aβs) within Chinese hamster ovary cells co-expressing β C-terminal fragment and wild-type presenilin 1 (C99/wtPS1 cells). In this study, we used sucrose density gradient centrifugation to fractionate the membranes from C99/wtPS1 cells that had been pretreated with DAPT. We found that accumulating Aβ46 localized exclusively to low density membrane (LDM) domains. Incubating the Aβ46-accumulating LDM domains at 37 °C produced Aβ40, Aβ42, Aβ43, and β-amyloid precursor protein intracellular domain. The addition of L685,458 completely prevented β-amyloid precursor protein intracellular domain generation and resulted in a large decrease in the level of Aβ46 and the concomitant appearance of Aβ40 and Aβ43 but not Aβ42. Further addition of DAPT suppressed the production of Aβ40/43 and abolished the decrease in the amount of Aβ46. These data indicate that preaccumulated Aβ46 is processed by γ-secretase to Aβ40/43 but not to Aβ42 in the LDM domains. The amount of newly produced Aβ40 and Aβ43 was roughly equivalent to the decrease in the amount of Aβ46. Temporal profiles did not show a maximal concentration for Aβ43, suggesting that Aβ46 is processed to Aβ40 and Aβ43 through a nonsuccessive process.