受miRNA-205调控的重组柯萨奇病毒B3的构建及其复制  

Construction of miRNA-205-regulated recombinant Coxsackievirus B3 and its replication

作　　者：戴潇逸 钟江 DAI Xiaoyi;ZHONG Jiang(Department of Microbiology and Microbial Engineering,School of Life Sciences,Fudan University,Shanghai 200438,China)

机构地区：[1]复旦大学生命科学学院微生物学与微生物工程系

出　　处：《微生物与感染》2019年第4期209-215,共7页Journal of Microbes and Infections

基　　金：国家自然科学基金(31370183)

摘　　要：本研究在柯萨奇病毒B3(coxsackievirus B3,CVB3)基因组P1编码区与P2编码区之间插入一段has-miRNA-205-3p和has-miRNA-205-5p(简称miR-205)的靶序列,得到重组病毒v205T,并比较分析了它在人宫颈癌细胞系HeLa细胞(miR-205低水平表达)和非小细胞肺癌细胞系A549细胞(miR-205高水平表达)中的复制情况。结果表明,插入的miR-205靶序列不影响病毒在HeLa细胞中的复制水平,但抑制了病毒在A549细胞中的复制,病毒滴度为对照的1%以下。为探讨v205T在2株细胞中复制差异的原因,进一步加入miR-205的类似物和抑制物。miR-205类似物可抑制v205T在HeLa细胞中复制和杀伤细胞的水平,而miR-205抑制物可提高v205T在A549细胞中的复制和杀伤细胞的水平。结果表明,v205T的复制确实受miR-205的调控。本研究为开发基于CVB3载体的溶瘤病毒和针对CVB3的减毒活疫苗提供了依据。A recombinant Coxsackievirus B3(CVB3),v205T,was constructed by inserting repeats of a target sequence for miR-205 between the coding sequence of P1 and P2.Its replication was analyzed in Hela and A549 cells,in which the level of miRNA-205 was known to be low and high,respectively.The results showed that the insertion of miRNA-205 targets in the selected site did not affect the infectivity of the recombinant virus in Hela cells,but greatly reduced its replication in A549 cells,resulting in more than 100-fold reduction in virus titer.miR-205 mimics and inhibitors were used to study the mechanism of differential replication of v205T.miR-205 mimics inhibited the replication and cytotoxicity of v205T in Hela cells,whereas miRNA-205 inhibitors increased the replication and cytotoxicity of v205T in A549.These results showed that the replication of v205T was regulated by cellular miR-205.The work provided basis for using CVB3 vectors for cancer viral therapy and vaccine development.

关 键 词：柯萨奇病毒B3 miRNA-205 感染性克隆载体 细胞趋性 

分 类 号：R73[医药卫生—肿瘤]