Dissecting the roles of Cse1 and Nup2 in classical NLS-cargo release in vivo.
Dissecting the roles of Cse1 and Nup2 in classical NLS-cargo release in vivo.
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DOI：
10.1111/tra.12759
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发表时间：
2020-10
期刊：
影响因子：
--
通讯作者：
Corbett AH
中科院分区：
文献类型：
--
作者：
Lange A;Fasken MB;Stewart M;Corbett AH
输入素α/β转运机制介导携带经典核定位序列（cNLS）的货物蛋白的核输入。这些货物蛋白通过importin-α衔接子连接到主要的核蛋白输入因子importin-β，之后货物/载体复合物通过核孔进入细胞核。在细胞核中，货物通过RanGTP和核孔蛋白Nup 2的作用释放，之后，输入蛋白再循环到细胞质中进行进一步的运输循环。importin-α的核输出由Cse 1/CAS介导。在这里，我们利用功能上重要的复合物的结构来识别对这些相互作用至关重要的残基，并使用酿酒酵母模型深入了解蛋白质输入和importin-α再循环的循环如何在体内发生。我们研究这些分子相互作用如何影响蛋白质定位，货物进口，功能和复合物的形成。我们发现，逆转importin-α（Arg 44）或Cse 1（Asp 220）中关键残基的电荷会导致相应蛋白质功能的丧失，并在体外和体内损害复合物的形成。为了扩展这些结果，我们表明Nup 2 N-末端的碱性残基是Nup 2与importin-α相互作用和Nup 2功能所必需的。这些结果提供了一个更全面的机制模型，说明Cse 1、RanGTP和Nup 2如何协同作用介导cNLS-货物在细胞核中的释放。细胞核和细胞质之间的货物定向运输由受体介导，所述受体在一个隔室中结合货物并将货物释放到目的地隔室中。含有cNLS的货物被细胞质中的importin-α识别。释放因子包括输入素-α输出受体Cse 1和核孔复合蛋白Nup 2，确保有效的货物输送到细胞核。先前结构研究定义的相互作用是importin-α、Cse 1和Nup 2之间在体内发生有效相互作用所必需的。
The importin α/β transport machinery mediates the nuclear import of cargo proteins that bear a classical nuclear localization sequence (cNLS). These cargo proteins are linked to the major nuclear protein import factor, importin‐β, by the importin‐α adapter, after which cargo/carrier complexes enter the nucleus through nuclear pores. In the nucleus, cargo is released by the action of RanGTP and the nuclear pore protein Nup2, after which the importins are recycled to the cytoplasm for further transport cycles. The nuclear export of importin‐α is mediated by Cse1/CAS. Here, we exploit structures of functionally important complexes to identify residues that are critical for these interactions and provide insight into how cycles of protein import and recycling of importin‐α occur in vivo using a Saccharomyces cerevisiae model. We examine how these molecular interactions impact protein localization, cargo import, function and complex formation. We show that reversing the charge of key residues in importin‐α (Arg44) or Cse1 (Asp220) results in loss of function of the respective proteins and impairs complex formation both in vitro and in vivo. To extend these results, we show that basic residues in the Nup2 N‐terminus are required for both Nup2 interaction with importin‐α and Nup2 function. These results provide a more comprehensive mechanistic model of how Cse1, RanGTP and Nup2 function in concert to mediate cNLS‐cargo release in the nucleus. Directional transport of cargoes between the nucleus and cytoplasm is mediated by receptors that bind cargo in one compartment and release cargo into a destination compartment. Cargoes that contain a cNLS are recognized by importin‐α in the cytoplasm. Release factors including the importin‐α export receptor, Cse1, and a nuclear pore complex protein, Nup2, ensure efficient cargo delivery into the nucleus. Interactions defined by previous structural studies are required for productive interactions between importin‐α, Cse1, and Nup2 to occur in vivo.
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DOI：
10.1083/jcb.130.5.1017
发表时间：
1995-09
期刊：
The Journal of cell biology
影响因子：
--
作者：
Corbett AH;Koepp DM;Schlenstedt G;Lee MS;Hopper AK;Silver PA
通讯作者：
Silver PA
DOI：
10.1083/jcb.138.1.65
发表时间：
1997-07-14
期刊：
The Journal of cell biology
影响因子：
--
作者：
Görlich D;Dabrowski M;Bischoff FR;Kutay U;Bork P;Hartmann E;Prehn S;Izaurralde E
通讯作者：
Izaurralde E
DOI：
10.1038/7625
发表时间：
1999-04-01
期刊：
NATURE STRUCTURAL BIOLOGY
影响因子：
--
作者：
Kobe, B
通讯作者：
Kobe, B
影响因子：
3.3
作者：
Duina, Andrea A.;Miller, Mary E.;Keeney, Jill B.
通讯作者：
Keeney, Jill B.
影响因子：
64.5
作者：
Kutay, U;Bischoff, FR;Gorlich, D
通讯作者：
Gorlich, D