RanGTP-mediated nuclear export of karyopherin alpha involves its interaction with the nucleoporin Nup153
RanGTP-mediated nuclear export of karyopherin alpha involves its interaction with the nucleoporin Nup153
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DOI：
10.1073/pnas.94.18.9699
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发表时间：
1997-09-02
影响因子：
11.1
通讯作者：
Radu, A
中科院分区：
文献类型：
--
作者：
Moroianu, J;Blobel, G;Radu, A
通过结合分析，我们发现了核粘蛋白α2和核孔蛋白Nup153之间的相互作用，并绘制了它们的相互作用结构域。我们还分离了一个15 kDa的核粘附素β1胰酶片段，称为β1*，它包含一个与含有核孔蛋白Nup98的多肽重复序列结合的决定簇。在体外实验中，用抗核粘附素α抗体的间接免疫荧光定量监测了洋地黄素通透性细胞的细胞核内源性核粘附素a的输出，我们发现核粘附素α的输出被添加的GTP酶RAN刺激，需要GTP水解，而被小麦胚凝集素抑制。代表Nup153和核粘附素α2相互作用结构域的多肽抑制了RanGTP介导的核粘附素α的输出，表明体外检测到的结合反应是生理相关的，并验证了我们的作图数据。此外，β1*虽然抑制了核粘附素的进口，但并不抑制核粘附素α的出口。因此，核粘附素α进出细胞核是不对称的过程。
Using binding assays, we discovered an interaction between karyopherin alpha 2 and the nucleoporin Nup153 and mapped their interacting domains. We also isolated a 15-kDa tryptic fragment of karyopherin beta 1, termed beta 1*, that contains a determinant for binding to the peptide repeat containing nucleoporin Nup98, In an in vitro assay in which export of endogenous nuclear karyopherin a from nuclei of digitonin-permeabilized cells was quantitatively monitored by indirect immunofluorescence with anti-karyopherin alpha antibodies, we found that karyopherin alpha export was stimulated by added GTPase Ran, required GTP hydrolysis, and was inhibited by wheat germ agglutinin. RanGTP-mediated export of karyopherin alpha was inhibited by peptides representing the interacting domains of Nup153 and karyopherin alpha 2, indicating that the binding reactions detected in vitro are physiologically relevant and verifying our mapping data. Moreover, beta 1*, although it inhibited import, did not inhibit export of karyopherin alpha. Hence, karyopherin alpha import into and export from nuclei are asymmetric processes.