Removal of a single pore subcomplex results in vertebrate nuclei devoid of nuclear pores
Removal of a single pore subcomplex results in vertebrate nuclei devoid of nuclear pores
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DOI：
10.1016/s1097-2765(03)00116-3
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发表时间：
2003-04-01
期刊：
影响因子：
16
通讯作者：
Forbes, DJ
中科院分区：
文献类型：
--
作者：
Harel, A;Orjalo, AV;Forbes, DJ
脊椎动物核孔复合体的大小是核糖体的30倍，由可溶性亚基和两种膜蛋白组成。利用非洲爪蟾核重构提取物的免疫消耗，以前有可能组装缺乏与蛋白质输入、输出或mRNA输出相关的孔亚基的细胞核。但这些蚀变孔隙仍具有孔隙结构的主体。在这里，我们使用Nup85和Nup133抗体（Nup85和Nup133是Nup107-160复合体的两个组成部分）免疫消耗单个亚基。重组后的细胞核在NLS导入和DNA复制方面存在严重缺陷。引人注目的是，它们显示出每一种被测试的核孔蛋白都有深刻的缺陷。即使是完整的膜蛋白POM121和gp210也不存在或无组织。扫描电镜显示无孔核，而添加Nup407-160复合体恢复功能孔。我们得出结论，Nup107-160复合物是脊椎动物核孔复合物组装的关键决定因素。
The vertebrate nuclear pore complex, 30, times the size of a ribosome, assembles from a library of soluble subunits and two membrane proteins. Using immunodepletion of Xenopus nuclear reconstitution extracts, it has previously been possible to assemble nuclei lacking pore subunits tied to protein import, export, or mRNA export. However, these altered pores all still possessed the bulk of pore structure. Here, we immunodeplete a single subunit, the Nup107-160 complex, using antibodies to Nup85 and Nup133, two of its components. The resulting reconstituted nuclei are severely defective for NLS import and DNA replication. Strikingly, they show a profound defect for every tested nucleoporin. Even the integral membrane proteins POM121 and gp210 are absent or unorganized. Scanning electron microscopy reveals pore-free nuclei, while addback of the Nup407-160 complex restores functional pores. We conclude that the Nup107-160 complex is a pivotal determinant for vertebrate nuclear pore complex assembly.