Akiyama Shoko, Yonezawa Takayuki, Kudo Tada-Aki, Li Ming Guang, Wang Hong, Ito Michihiko, Yoshioka Katsuji, Ninomiya-Tsuji Jun, Matsumoto Kunihiro, Kanamaru Ryunosuke, Tamura Shinri, Kobayashi Takayasu
Departments of Biochemistry and Clinical Oncology, Institute of Development, Aging and Cancer, Tohoku University, 4-1 Seiryo-machi, Aoba-ku, Sendai 980-8575, Japan.
J Biol Chem. 2004 Aug 27;279(35):36616-20. doi: 10.1074/jbc.M406610200. Epub 2004 Jun 24.
P19 embryonic carcinoma cells, a model system for studying early development and differentiation, can differentiate into neurons and primitive endoderm-like cells depending on the culture conditions. We have previously reported that the activation of c-Jun amino-terminal kinase (JNK) is required for the retinoic acid-induced neural differentiation of P19 cells. However, the signaling pathway(s) responsible for the activation of JNK has not been known. In this study, we demonstrated that activities of MAPK kinase 4 (MKK4) and TAK1, one of the upstream kinases of MKK4, were enhanced in the neurally differentiating cells. Inhibition of the neural differentiation by an overexpression of protein phosphatase 2Cepsilon, an inactivator of TAK1, suggested a critical role of the TAK1 signaling pathway during the differentiation. Confocal microscopic analysis indicated that TAK1, phospho-MKK4, and phospho-JNK were colocalized with tubulin in the neurites and localized also in the nuclei of the differentiating cells. In contrast, two TAK1-binding proteins, TAB1 and TAB2, which are involved in the activation of TAK1, were localized in the neurites and the nuclei of the differentiating cells, respectively. These results suggest that two distinct TAK1-MKK4-JNK signaling pathways are independently activated at the different intracellular locations and may participate in the regulation of the neural differentiation of P19 cells.
P19胚胎癌细胞是一种用于研究早期发育和分化的模型系统，根据培养条件可分化为神经元和原始内胚层样细胞。我们之前报道过，c-Jun氨基末端激酶（JNK）的激活是视黄酸诱导P19细胞神经分化所必需的。然而，负责JNK激活的信号通路尚不清楚。在本研究中，我们证明了丝裂原活化蛋白激酶激酶4（MKK4）和TAK1（MKK4的上游激酶之一）的活性在神经分化细胞中增强。通过过表达TAK1的失活剂蛋白磷酸酶2Cε来抑制神经分化，表明TAK1信号通路在分化过程中起关键作用。共聚焦显微镜分析表明，TAK1、磷酸化MKK4和磷酸化JNK与微管蛋白在神经突中共定位，并且也定位于分化细胞的细胞核中。相比之下，参与TAK1激活的两个TAK1结合蛋白TAB1和TAB2分别定位于分化细胞的神经突和细胞核中。这些结果表明，两条不同的TAK1-MKK4-JNK信号通路在不同的细胞内位置被独立激活，并可能参与P19细胞神经分化的调控。
