国家重点基础研究发展规划“恶性肿瘤发生发展的基础研究”项目(973)(G1998051201)和国家自然科学基金项目(30000087).
This work was supported by grants from Special Funds for Major State Basic Research of China (1998051201) and The National Natural Sciences Fundation of China (30000087).
为了探讨在鼻咽癌细胞中EB病毒编码的潜伏膜蛋白1(LMP1)激活c-Jun氨基端激酶(JNK)信号途径的分子机制,利用可调控表达LMP1的鼻咽癌细胞系L7,蛋白质印迹检测,发现LMP1能够促进JNK的活化;利用稳定表达LMP1的鼻咽癌细胞系HNE2-LMP1及其三种突变体HNE2-LMP1ΔCTAR1、HNE2-LMP1ΔCTAR2、HNE2-LMP1ΔCTAR1,2及LMP1阴性的HNE2为材料,采用蛋白质印迹和报告基因法分析JNK和活化蛋白1(AP1)活化情况,结果显示HNE2-LMP1和HNE2-LMP1ΔCTAR1中磷酸化JNK蛋白表达量和AP1活性都无显著差异,而与HNE2-LMP1ΔCTAR2、HNE2-LMP1ΔCTAR1,2、阴性对照HNE2及空白载体转染细胞的JNK蛋白表达和AP1活性具有显著差异;进一步比较转染TRAF、TRADD显性负性突变体鼻咽癌细胞系HNE2-LMP1中磷酸化的JNK量和AP1活性,结果显示:TRAF-DN和TRADD-DN的导入使活化的JNK蛋白和AP-1活性显著降低,二者间无显著差异,提示TRAF和TRADD可能参与了LMP1对JNK和AP-1的活化.以上结果提示在鼻咽癌细胞系中LMP1功能结构域CTAR2通过结合TRAF/TRADD激活JNK从而活化重要的转录因子AP1.
In order to illustrate the mechanism of AP1 signalling pathway mediated by latent membrane protein 1 (LMP1) encoded by EB virus, the expression of phospho-JNK in Tet-on-LMP1-HNE2 cell line (L7) was determined at different time by Western blotting, JNK activation increased with the induction of LMP1 in a time dependant manner was abserved. The expression of phospho-JNK and AP1 activity were analyzed by Western blotting and reporter gene in nasopharyngeal carcinoma cell lines which stably expressing LMP1 and the three kinds of its mutants containing different mutation in carboxyterminal activating region. The result showed that no difference existed between HNE2-LMP1 and HNE2-LMP1ΔCTAR1, but significant difference between HNE2-LMP1ΔCTAR2 and HNE2-LMP1, HNE2-LMP1ΔCTAR1,2. The phospho-JNK expression and AP1 activity of HNE2-LMP1 cell lines were individually transfected by dominant negative TRAF(TRAF-DN) and dominant negative TRADD(TRADD-DN). The results showed that the transfection of TRAF-DN or TRADD-DN made JNK expression and AP1 activity decreased significantly. The results suggested that the functional domain CTAR2 of LMP1 encoded by EB virus can mediate JNK signalling pathway through cooperation with TRAF/TRADD complex.
胡智,曾亮,陶永光,唐发清,王海,罗非君,易薇,曹亚. EB病毒潜伏膜蛋白1通过TRAF/TRADD激活JNK信号途径[J].生物化学与生物物理进展,2002,29(4):562-566
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