TAK1赖氨酸158位点发生Lys63连接多聚泛素化在辐射诱导NF-κB激活过程中的作用  

Roei of Lys63-1inked TAK1 poly-ubiquitination at lysine 158 in radiation-induced NF-κB activity

作　　者：朱翔[1] 曹宝山[2] 张照辉[2] 马力文[2] 苏旭[3] 梁莉[2] 

机构地区：[1]北京大学第三医院病理科,100191 [2]北京大学第三医院肿瘤化疗与放射病科,100191 [3]中国疾病预防控制中心辐射防护与核安全医学所

出　　处：《中华放射医学与防护杂志》2013年第3期235-238,共4页Chinese Journal of Radiological Medicine and Protection

摘　　要：目的研究转化生长因子激酶1（TAK1）Lys63连接多聚泛素化在辐射导致的NF-κB激活过程中起到的作用及其作用位点。方法通过基因瞬时转染技术将FLAG—TAK1和HA-Ub—K63质粒共同转染至HEK-293T细胞，应用免疫沉淀和免疫印迹技术研究辐射对TAK1泛素化的影响；用稳定转染TAK1表达的HeLa细胞，证实辐射对TAK1泛素化的影响。通过免疫印迹技术，比较分析辐射对稳定转染TAK1K158R的MEF细胞与TAK1野生型MEF细胞比较分析辐射对它表达1KKs、p38及JNK的影响，以证明辐射诱导的TAK1-Lys63多聚泛素化的位点是否在TAKI赖氨酸158位点。结果瞬时转染的HEK-293T细胞接受辐射后1h开始出现明显的TAK1泛素化表现，2h更明显。在稳定表达FLAG—TAKI的HeLa细胞系中得到证实。与TAK1野生型MEF细胞相比，TAK1K158R突变的MEF细胞中辐射诱导的IKKs和p38磷酸化作用减低。结论TAK1多聚泛素化在辐射导致的NF—κB激活过程中起到关键作用，辐射所致TAK1-Lys63连接多聚泛素化发生在TAK1的赖氨酸158位点。Objective To study the role of the transforming growth factor-13-activating kinase 1 （TAK1） poly-ubiquitination in radiation-induced NF-κB activation. Methods FLAG-TAK1 and HA-Ub- K63 plasmids were transiently transfected into HEK-293T cells. The radiation effect on TAK1 ubiquitination was detected by immunoprecipitation and Western blot, which was further confirmed in FLAG-TAK1 HeLa stable cell line. Results At 1 h after irradiation, Lys 63-1ink TAK1 ubiquitination was induced in the HEK-293T cells with plasmids transfection. This result was further confirmed by using a FLAG-TAK1 stably expressed HeLa cell line. Radiation-induced IKKs and p38 phosphorylation were greatly impaired in MEFs reconstituted with TAKI K158R mutant compared to ones with wild-type TAK1. Conclusions Lys63-1inked TAK1 poly-ubiquitination at Lys-58 plays a key role in the process of radiation-induced NF-κB activation.

关 键 词：辐射损伤 转化生长因子β激酶 多聚泛素化 NF-ΚB 信号通路 

分 类 号：R[医药卫生]