Crystal structure of the karyopherin Kap121p bound to the extreme C-terminus of the protein phosphatase Cdc14p
Crystal structure of the karyopherin Kap121p bound to the extreme C-terminus of the protein phosphatase Cdc14p
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DOI：
10.1016/j.bbrc.2015.05.060
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发表时间：
2015-07-31
影响因子：
3.1
通讯作者：
Matsuura, Yoshiyuki
中科院分区：
文献类型：
--
作者：
Kobayashi, Junya;Hirano, Hidemi;Matsuura, Yoshiyuki
在酿酒酵母中，蛋白磷酸酶CDC14P是有丝分裂周期蛋白依赖性蛋白激酶的拮抗剂，是晚期有丝分裂事件的关键调节因子，如染色体分离、纺锤体分解和胞质分裂。CDC14p的活性受其与核仁中竞争抑制物Net1p(也称为Cfi1p)结合的细胞周期依赖性变化的控制。在直到中期的大部分细胞周期中，CDC14P以不活跃的状态被隔离在核仁中。在后期，CDC14P从Net1p释放，扩散到细胞核和细胞质，并使关键的有丝分裂靶点去磷酸化。尽管CDC14p的核质穿梭被认为是退出有丝分裂的重要因素，但其核转运的机制仍然知之甚少。我们发现，在体内，CDC14p的C末端区域(残基517-551)可以作为核定位信号(NLS)，也可以在体外以Gsp1p-GTP依赖的方式与Kap121p(也称为Pse1p)结合，Kap121p(也称为Pse1p)是酵母中重要的核输入载体。此外，我们还报道了一种2.4埃分辨率的Kap121P晶体结构，该结构结合在CDC14P的C-末端区域。结构和基于结构的突变分析表明，CDc14p末端的最后5个残基(残基547-551；Gly-Ser-Ile-Lys-Lys)或邻近序列相似的残基(残基540-544；Gly-Gly-Ile-Arg-Lys)可以与Kap121p的NLS结合位点结合，其中CDc14p的两个残基(中间的Ile和5个残基的末端的Lys)对结合特异性起关键作用。在与Kap121P-配体络合物的其他结构进行比较的基础上，我们建议将“IK-NLS”作为Kap121P特异的NLS的合适术语。(C)2015 Elsevier Inc.保留所有权利。
In Saccharomyces cerevisiae, the protein phosphatase Cdc14p is an antagonist of mitotic cyclin-dependent kinases and is a key regulator of late mitotic events such as chromosome segregation, spindle disassembly and cytokinesis. The activity of Cdc14p is controlled by cell-cycle dependent changes in its association with its competitive inhibitor Net1p (also known as Cfi1p) in the nucleolus. For most of the cell cycle up to metaphase, Cdc14p is sequestered in the nucleolus in an inactive state. During anaphase, Cdc14p is released from Net1p, spreads into the nucleus and cytoplasm, and dephosphorylates key mitotic targets. Although regulated nucleocytoplasmic shuttling of Cdc14p has been suggested to be important for exit from mitosis, the mechanism underlying Cdc14p nuclear trafficking remains poorly understood. Here we show that the C-terminal region (residues 517-551) of Cdc14p can function as a nuclear localization signal (NLS) in vivo and also binds to Kap121p (also known as Pse1p), an essential nuclear import carrier in yeast, in a Gsp1p-GTP-dependent manner in vitro. Moreover we report a crystal structure, at 2.4 angstrom resolution, of Kap121p bound to the C-terminal region of Cdc14p. The structure and structure-based mutational analyses suggest that either the last five residues at the extreme C-terminus of Cdc14p (residues 547-551; Gly-Ser-Ile-Lys-Lys) or adjacent residues with similar sequence (residues 540-544; Gly-Gly-Ile-Arg-Lys) can bind to the NLS-binding site of Kap121p, with two residues (Ile in the middle and Lys at the end of the five residues) of Cdc14p making key contributions to the binding specificity. Based on comparison with other structures of Kap121p-ligand complexes, we propose "IK-NLS" as an appropriate term to refer to the Kap121p-specific NLS. (C) 2015 Elsevier Inc. All rights reserved.