Phosphoproteomics reveals new ERK MAP kinase targets and links ERK to nucleoporin-mediated nuclear transport
Phosphoproteomics reveals new ERK MAP kinase targets and links ERK to nucleoporin-mediated nuclear transport
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DOI：
10.1038/nsmb.1656
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发表时间：
2009-10-01
影响因子：
16.8
通讯作者：
Hattori, Seisuke
中科院分区：
文献类型：
--
作者：
Kosako, Hidetaka;Yamaguchi, Nozomi;Hattori, Seisuke
许多细胞外信号调节激酶（ERK）有丝分裂原活化蛋白（MAP）激酶底物已被确定，但ERK介导的过程的多样性表明存在其他目标。使用磷酸化蛋白质组学方法结合类固醇受体融合系统，IMAC，2D-DIGE和磷酸化特异性抗体，我们检测到38个蛋白质显示ERK激活和ERK抑制样品之间的可重复磷酸化变化，包括24个新的候选ERK靶点。ERK在体外直接磷酸化至少13种蛋白质。其中，Nup 50被验证为真正的ERK底物。值得注意的是，Nup 50的FG重复区的ERK磷酸化降低了其对输入蛋白-β家族蛋白、输入蛋白-β和转运蛋白的亲和力。其他FG核孔蛋白在ERK介导的磷酸化后表现出类似的功能变化。在ERK激活的毛地黄皂苷透化细胞中，由于Nup 50的ERK磷酸化，importin-β和转运蛋白的核迁移受损。因此，我们认为ERK磷酸化各种核孔蛋白来调节核质转运。
Many extracellular signal-regulated kinase (ERK) mitogen-activated protein ( MAP) kinase substrates have been identified, but the diversity of ERK-mediated processes suggests the existence of additional targets. Using a phosphoproteomic approach combining the steroid receptor fusion system, IMAC, 2D-DIGE and phosphomotif-specific antibodies, we detected 38 proteins showing reproducible phosphorylation changes between ERK-activated and ERK-inhibited samples, including 24 new candidate ERK targets. ERK directly phosphorylated at least 13 proteins in vitro. Of these, Nup50 was verified as a bona fide ERK substrate. Notably, ERK phosphorylation of the FG repeat region of Nup50 reduced its affinity for importin-beta family proteins, importin-beta and transportin. Other FG nucleoporins showed a similar functional change after ERK-mediated phosphorylation. Nuclear migration of importin-beta and transportin was impaired in ERK-activated, digitonin-permeabilized cells, as a result of ERK phosphorylation of Nup50. Thus, we propose that ERK phosphorylates various nucleoporins to regulate nucleocytoplasmic transport.