Zhang Zhi-Wei, He Zhi-Min, Zhou Min, Zhang Qiong, Yu Yan-Hui, Chen Zhu-Chu
Cancer Research Institute, Xiangya Medical College, Central South University, Changsha, Hunan, PR China.
Ai Zheng. 2007 Feb;26(2):118-22.
BACKGROUND & OBJECTIVE: Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) is an oncoprotein coded by EBV genome. This study was to investigate the effects of EBV LMP1 on transformation and tumorigenesis of Rat-1 cells.
Retrovirus plasmids pLNSX-LMP1 and pBabe-IkappaBalpha, constructed by gene recombination technique, were cotransfected respectively with nuclear factor-kappaB luciferase reporter (pNF-kappaB-luc) into 293 cells. The actions of LMP1 in activating NF-kappaB and IkappaBalpha in inhibiting NF-kappaB were measured by luciferase activity assay. Moreover, pLNSX-LMP1 and pBabe-IkappaBalpha were transfected respectively into the ecotropic retrovirus packaging cell line PA317 to generate LMP1 retrovirus (RV-LMP1) and IkappaBalpha retrovirus (RV-IkappaBalpha). After Rat-1 cells were infected by RV-LMP1 alone or RV-LMP1 combined RV-IkappaBalpha, their malignant transformation phenotype was detected by colony forming assay and nude mice tumorigenicity assay.
When pLNSX-LMP1 and pBabe-IkappaB were cotransfected at a ratio of 1:1, IkappaBalpha inhibited LMP1-mediated NF-kappaB activation by 75%û and at a ratio of 3:1, it almost completely inhibited LMP1-mediated NF-kappaB activation. IkappaBalpha obviously inhibited LMP1-mediated malignant phenotype of Rat-1 cells:colony formation number on plates were significantly decreased from (368+/-7)/well and (287+/-17)/well to (59+/-6)/well and (8+/-2)/well (P<0.001). Foci in soft agarose were decreased from (477+/-13)/well and (347+/-10)/well to (61+/-15)/well and (95+/-7)/well (P<0.001). The ability of tumorigenicity in nude mice was markedly decreased: tumor volume was decreased from (1.61+/-0.23) cm3 to (0.20+/-0.08) cm3 (P<0.001).
EBV-LMP1 could lead to transformation and tumorigenesis of Rat-1 cells by activating NF-kappaB.
爱泼斯坦-巴尔病毒（EBV）潜伏膜蛋白1（LMP1）是由EBV基因组编码的一种癌蛋白。本研究旨在探讨EBV LMP1对Rat-1细胞转化和肿瘤发生的影响。
采用基因重组技术构建的逆转录病毒质粒pLNSX-LMP1和pBabe-IκBα，分别与核因子-κB荧光素酶报告基因（pNF-κB-luc）共转染293细胞。通过荧光素酶活性测定法检测LMP1激活NF-κB的作用以及IκBα抑制NF-κB的作用。此外，将pLNSX-LMP1和pBabe-IκBα分别转染到嗜亲性逆转录病毒包装细胞系PA317中，以产生LMP1逆转录病毒（RV-LMP1）和IκBα逆转录病毒（RV-IκBα）。Rat-1细胞单独感染RV-LMP1或RV-LMP1与RV-IκBα联合感染后，通过集落形成试验和裸鼠致瘤性试验检测其恶性转化表型。
当pLNSX-LMP1和pBabe-IκB以1:1的比例共转染时，IκBα抑制LMP1介导的NF-κB激活达75%，以3:1的比例共转染时，几乎完全抑制LMP1介导的NF-κB激活。IκBα明显抑制LMP1介导的Rat-1细胞恶性表型：平板上的集落形成数从（368±7）/孔和（287±17）/孔显著降至（59±6）/孔和（8±2）/孔（P<0.001）。软琼脂中的集落数从（477±13）/孔和（347±10）/孔降至（61±15）/孔和（95±7）/孔（P<0.001）。裸鼠中的致瘤能力明显降低：肿瘤体积从（1.61±0.23）cm3降至（0.20±0.08）cm3（P<0.001）。
EBV-LMP1可通过激活NF-κB导致Rat-1细胞转化和肿瘤发生。
Zotero 插件