Ranganathan Aarati, Yazicioglu Mustafa N, Cobb Melanie H
Department of Pharmacology, The University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75390-9041, USA.
J Biol Chem. 2006 Jun 9;281(23):15645-52. doi: 10.1074/jbc.M513866200. Epub 2006 Apr 4.
The mitogen-activated protein (MAP) kinases ERK1 and ERK2 often accumulate in the nuclei of stimulated cells to mediate changes in transcription. The mechanisms underlying stimulus-dependent redistribution of these kinases remain unclear. We have used a permeabilized cell reconstitution assay in HeLa cells and human foreskin fibroblasts to explore the processes by which ERK2 enters and exits the nucleus. We previously reported that entry of unphosphorylated ERK2 into the nucleus occurs by facilitated diffusion not requiring cytosolic transport factors. We find that export, like import, can occur by an energy- and carrier-independent mechanism. An energy-dependent mechanism of ERK2 export can also be distinguished, mediated at least in part through the exportin CRM1. We have also examined import and export of thiophosphorylated, active ERK2. Import of active ERK2 is significantly enhanced by the addition of exogenous transport factors and an energy regeneration system. These studies support a model in which multiple constitutive and regulated processes control the subcellular distribution of ERK2.
丝裂原活化蛋白（MAP）激酶ERK1和ERK2常常在受刺激细胞的细胞核中积累，以介导转录变化。这些激酶依赖刺激的重新分布背后的机制仍不清楚。我们利用人宫颈癌细胞系（HeLa细胞）和人包皮成纤维细胞的透化细胞重建试验，来探究ERK2进出细胞核的过程。我们之前报道过，未磷酸化的ERK2进入细胞核是通过易化扩散实现的，不需要胞质转运因子。我们发现，与入核一样，出核也可以通过一种不依赖能量和载体的机制发生。还可以区分出一种ERK2出核的能量依赖机制，至少部分是由核输出蛋白CRM1介导的。我们也研究了硫代磷酸化的活性ERK2的入核和出核。添加外源性转运因子和能量再生系统后，活性ERK2的入核显著增强。这些研究支持了一个模型，即多个组成性和调节性过程控制着ERK2的亚细胞分布。
