Nuclear translocation of mitogen-activated protein kinase kinase (MEK1) in response to mitogenic stimulation
Nuclear translocation of mitogen-activated protein kinase kinase (MEK1) in response to mitogenic stimulation
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DOI：
10.1073/pnas.94.8.3742
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发表时间：
1997-04-15
影响因子：
11.1
通讯作者：
Seger, R
中科院分区：
文献类型：
--
作者：
Jaaro, H;Rubinfeld, H;Seger, R
丝裂原活化蛋白激酶激酶（MEK）是一种双特异性蛋白激酶，主要位于细胞胞质溶胶中，在促有丝分裂刺激之前和之后。在MEK的N-末端结构域中存在核输出信号[Fukuda，M.，后藤岛戈托岛，巴西-地& Nishida，E.（1996）J.Biol.Chem.271，20024-20028]表明存在MEK进入细胞核并且必须输出的情况。使用突变体的MEK，我们表明，从组成型活性MEK的核输出信号序列的缺失引起的COS 7和HEK-293 T细胞的细胞核中的MEK组成型定位。然而，当从催化失活的MEK中删除相同的区域时，在静息细胞中观察到细胞质定位，其在刺激时变成核。表达上述突变体的COS 7细胞的共聚焦显微镜显示活性MEK在核膜中以及在细胞外周中的定位。野生型和突变型MEK之间细胞定位的差异并不是因为特异性的严重变化，因为缺乏N末端区域的重组组成型活性MEK在体外和完整细胞中都表现出与野生型MEK相同的底物特异性。总而言之，我们的结果表明，在促有丝分裂刺激后，MEK与细胞外信号响应激酶和p90（RSK）一样，被大量转移到细胞核。由核输出信号介导的从核的快速输出可能是用野生型MEK观察到的细胞质分布的原因。
Mitogen-activated protein kinase kinase (MEK) is a dual-specificity protein kinase that is located primarily in the cellular cytosol, both prior to and upon mitogenic stimulation. The existence of a nuclear export signal in the N-terminal domain of MEK [Fukuda, M., Gotoh, I., Gotoh, Y. & Nishida, E. (1996) J. Biol. Chem. 271, 20024-20028] suggests that there are circumstances under which MEK enters the nucleus and must be exported. Using mutants of MEK, we show that the deletion of the nuclear export signal sequence from constitutively active MEK caused constitutive localization of MEK in the nucleus of COS7 and HEK-293T cells. However, when the same region was deleted from a catalytically inactive MEK, cytoplasmic localization was observed in resting cells, which turned nuclear upon stimulation. Confocal microscopy of COS7 cells expressing the above mutants showed localization of the active MEK in the nuclear envelope and also in the cell periphery. The differences in cellular localization between the wild-type and mutant MEKs are not due to severe changes in specificity because the recombinant, constitutively active MEK that lacked its N-terminal region exhibited the same substrate specificity as the wild-type MEK, both in vitro and in intact cells. Taken together, our results indicate that upon mitogenic stimulation, MEK, like extracellular signal responsive kinase and p90(RSK), is massively translocated to the nucleus. Rapid export from the nucleus, which is mediated by the nuclear export signal, is probably the cause for the cytoplasmic distribution observed with wild-type MEK.