EB病毒LMP1 C-端序列缺失对细胞增殖的影响  

The Effect of 30-Base Pair Deletion of Latent Membrane Protein 1 C Terminus of Epstein-Barr Virus on Cell Proliferation

作　　者：杜海军[1] 赵健[2] 周玲[1] 王琦[1] 李红霞[1] 曾毅[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所传染病预防控制国家重点实验室,北京100052 [2]北京大学附属第一医院妇产科,北京100034

出　　处：《病毒学报》2006年第6期440-444,共5页Chinese Journal of Virology

基　　金：国家自然科学基金(30270520);国家科技攻关项目(2003BA712A01)

摘　　要：潜伏膜蛋白1(Latent membrane protein 1,LMP1)是由γ疱疹病毒亚科的EB病毒基因编码的能够使正常细胞发生恶性转化的跨膜信号蛋白,C-端为LMP1的主要功能区。通过PCR从B95-8细胞和鼻咽癌活检组织中获得lmp1和lmp1 C-端30bp缺失(lmp1 C-terminal deleted,lmp1-ctd)基因;利用体外基因重组技术,将lmp1和lmp1-ctd分别插入AAV质粒载体中;应用Lipofectamine介导转染HEK-293包装细胞,获得含lmp1和lmp1-ctd的rAAV;再以rAAV感染猴肾上皮细胞。利用RT-PCR检测lmp1和lmp1-ctd在猴肾上皮细胞中的转录;MTT法、流式细胞仪技术检测细胞生长和DNA合成,探讨lmp1和lmp1-ctd对其在细胞中活性的影响。结果显示:lmp1在转化细胞中转录,导致细胞生长速度加快,其中lmp1-ctd转化细胞的生长速度和处于增殖期比例高于lmp1转化细胞。提示lmp1-ctd较lmp1具有更强的促细胞增殖作用。Latent membrane protein 1 （LMP1） encoded by Epstein-Barr virus gene is a kind of transmembrane signal protein causing cell malignancy. LMP1 C terminus is essential for its transforming activities. Both Lmp 1 and lmp 1 C-terminal 30-base pair deleted （lmp 1-ctd） genes were respectively extracted from B95-8 cell and nasopharyngeal carcinoma biopsy by polymerase chain reaction （PCR）. These genes were separately inserted into adenovirus-associated virus （AAV） plasmid vector with DNA recombinant technique. The recombinant AAVs （rAAVs） each including lmp1 and lmp1-ctd were assembled in HEK-293 package cell line under the help of lipofectamine transfection reagents. The epithelial cells of monkey kidney were infected by rAAVs. The transcription of lmp1 and lmp1-ctd in monkey kidney cells was determined by RT-PCR. The growth and DNA synthesis of infected epithelial cells of monkey kidney were examined with MTT method and flow cytometry technology. The effect of lmp1-ctd on cell proliferation was discussed. Our data suggested mRNAs of lmp1 and lmp1-ctd genes could be detected in the infected epithelial cells of monkey kidney. The growth of infected monkey kidney epithelial cells was strongly promoted by lmp1 and lmp1-ctd. More cells infected by rAAV including lmp 1-ctd transited quiescent phase to proliferation phase than cells including lmp1 did. These results indicated lmp 1-ctd has stronger activity to promote cell propagation.

关 键 词：γ疱疹病毒亚科 EB病毒潜伏膜蛋白1 C-端缺失 细胞增殖 

分 类 号：R373.3[医药卫生—病原生物学]