Kuwahara Hiroshi, Nishizaki Michihiko, Kanazawa Hiroshi
Department of Biological Sciences, Graduate School of Science, Osaka University, Machikaneyama-cho, Toyonaka City, Osaka, Japan.
J Biochem. 2008 Mar;143(3):349-58. doi: 10.1093/jb/mvm236. Epub 2007 Dec 15.
DAP kinase-related apoptosis-inducing kinase 2 (DRAK2) is a serine/threonine kinase of the death-associated protein kinase family. DRAK2 mediates apoptosis induced by extracellular stimuli, including UV irradiation and interleukin-2, and also regulates T-cell receptor sensitivity in developing thymocytes. During these events, the subcellular localization of DRAK2 changes between the nucleus and cytoplasm. We found that DRAK2 has a putative nuclear-localization signal (NLS) sequence. Mutations in this sequence interfered with DRAK2 localization to the nucleus. Furthermore, green fluorescence protein fused to the putative NLS accumulated in the nucleus, indicating that the putative sequence functions as an NLS. We also found that the function of the NLS was regulated by phosphorylation. Phorbol myristate acetate (PMA) induced the accumulation of DRAK2 in the cytoplasm of NIH3T3 cells, whereas in the absence of PMA, DRAK2 was localized to the nucleus. Ectopic expression of PKC-gamma induced cytoplasmic localization of DRAK2 and PKC-gamma phosphorylated Ser350 flanking the NLS. DRAK2, but not the Ser350Asp mutant, accumulated in the nuclei of ACL-15 cells in response to UV-irradiation. These results suggest that phosphorylation of Ser350 plays an essential role in regulating translocation of DRAK2 to the nucleus from the cytoplasm, possibly by affecting the activity of the NLS.
死亡相关蛋白激酶相关凋亡诱导激酶2（DRAK2）是死亡相关蛋白激酶家族的一种丝氨酸/苏氨酸激酶。DRAK2介导由细胞外刺激诱导的凋亡，包括紫外线照射和白细胞介素-2，并且还调节发育中的胸腺细胞中的T细胞受体敏感性。在这些过程中，DRAK2的亚细胞定位在细胞核和细胞质之间变化。我们发现DRAK2具有一个假定的核定位信号（NLS）序列。该序列中的突变干扰了DRAK2向细胞核的定位。此外，与假定的NLS融合的绿色荧光蛋白在细胞核中积累，表明该假定序列起到NLS的作用。我们还发现NLS的功能受磷酸化调节。佛波酯肉豆蔻酸酯乙酸盐（PMA）诱导DRAK2在NIH3T3细胞的细胞质中积累，而在没有PMA的情况下，DRAK2定位于细胞核。PKC-γ的异位表达诱导DRAK2的细胞质定位并且PKC-γ使NLS侧翼的Ser350磷酸化。响应紫外线照射，DRAK2而非Ser350Asp突变体在ACL-15细胞的细胞核中积累。这些结果表明，Ser350的磷酸化可能通过影响NLS的活性，在调节DRAK2从细胞质向细胞核的转运中起重要作用。
