Mandl Margret, Slack David N, Keyse Stephen M
Cancer Research UK, Molecular Pharmacology Unit, Biomedical Research Centre, Level 5, Ninewells Hospital and Medical School, Dundee DD1 9SY, Scotland, United Kingdom.
Mol Cell Biol. 2005 Mar;25(5):1830-45. doi: 10.1128/MCB.25.5.1830-1845.2005.
The mechanisms which determine the nuclear accumulation and inactivation of the extracellular signal-regulated kinase 1 (ERK1) or ERK2 mitogen-activated protein (MAP) kinases are poorly understood. Here we demonstrate that DUSP5, an inducible nuclear phosphatase, interacts specifically with ERK2 via a kinase interaction motif (KIM) within its amino-terminal noncatalytic domain. This binding determines the substrate specificity of DUSP5 in vivo, as it inactivates ERK2 but not Jun N-terminal protein kinase or p38 MAP kinase. Using green fluorescent protein fusions, we identify within this same domain of DUSP5 a functional nuclear localization signal (NLS) which functions independently of the KIM. Moreover, we demonstrate that the expression of DUSP5 causes both nuclear translocation and sequestration of inactive ERK2. Nuclear anchoring is ERK2 specific and requires both interactions between the DUSP5 KIM and the common docking site of ERK2 and a functional NLS within DUSP5. Finally, the expression of a catalytically inactive mutant of DUSP5 also tethers ERK2 within the nucleus. Furthermore, this nuclear ERK2 is phosphorylated by MAP kinase kinase in response to growth factors and also activates transcription factor Elk-1. We conclude that DUSP5 is an inducible nuclear ERK-specific MAP kinase phosphatase that functions as both an inactivator of and a nuclear anchor for ERK2 in mammalian cells. In addition, our data indicate that the cytoplasm may not be an exclusive site of MAP kinase activation.
目前对于决定细胞外信号调节激酶1（ERK1）或ERK2丝裂原活化蛋白（MAP）激酶的核内积累及失活的机制仍知之甚少。在此我们证明，诱导型核磷酸酶DUSP5通过其氨基末端非催化结构域内的激酶相互作用基序（KIM）与ERK2特异性相互作用。这种结合决定了DUSP5在体内的底物特异性，因为它能使ERK2失活，但不能使Jun氨基末端蛋白激酶或p38 MAP激酶失活。利用绿色荧光蛋白融合体技术，我们在DUSP5的同一结构域内鉴定出一个功能性核定位信号（NLS），其功能独立于KIM。此外，我们证明DUSP5的表达会导致无活性ERK2的核转位和隔离作用。核锚定作用具有ERK2特异性，并且需要DUSP5的KIM与ERK2的共同对接位点之间的相互作用以及DUSP5内的一个功能性NLS。最后，DUSP5催化失活突变体的表达也能将ERK2束缚在细胞核内。此外，这种核内ERK2会在生长因子的作用下被MAP激酶激酶磷酸化，并且还能激活转录因子Elk-1。我们得出结论，DUSP5是一种诱导型核ERK特异性MAP激酶磷酸酶 在哺乳动物细胞中，它既是ERK2的失活剂，也是ERK2的核锚定物。此外，我们的数据表明细胞质可能并非MAP激酶激活的唯一部位。
