柞蚕核型多角体病毒对Tn-Hi5细胞的感染性及抗细胞凋亡分析  

Analysis of Infectivity and Anti-apoptotic Function of Antheraea pernyi Nucleopolyhedrovirus to Tn- Hi5 Cells

作　　者：赵振军[1] 王林美[1] 叶博[1] 岳冬梅[1] 张波[1] 李树英[1] 都兴范[1] 范琦[1] 

机构地区：[1]辽宁省农业科学院大连生物技术研究所辽宁省野蚕重点实验室,辽宁大连116024

出　　处：《蚕业科学》2015年第2期292-299,共8页ACTA SERICOLOGICA SINICA

基　　金：国家自然科学基金项目(No.31072081);辽宁省科技厅农业攻关计划项目(No.2011208001)

摘　　要：细胞凋亡是昆虫宿主细胞抗病毒感染及控制病毒复制增殖的一个复杂的分子生物学机制。前期研究发现柞蚕核型多角体病毒(Ap NPV)的重组病毒Ap NPV-Δph/egfp+能够感染非特异性宿主细胞Tn-Hi5,并表达EGFP蛋白。利用实时荧光定量PCR方法进一步检测分析Ap NPV-Δph/egfp+在Tn-Hi5细胞中的复制增殖特点以及子代病毒的感染性;通过检测细胞凋亡信号通路中类caspase-3蛋白酶活性,分析Ap NPV-Δph/egfp+感染Tn-Hi5细胞后对抗细胞凋亡的作用途径。结果显示,随着病毒感染时间的延长,Ap NPV-Δph/egfp+在Tn-Hi5细胞中的DNA拷贝数增加,被感染的Tn-Hi5细胞能够产生子代病毒,但病毒DNA拷贝数逐代减少;Ap NPV-Δph/egfp+感染Tn-Hi5细胞后可抑制放线菌素D诱导的细胞类caspase-3蛋白酶活性。研究结果证实Ap NPV能够在Tn-Hi5细胞中复制增殖,并具有抑制由放线菌素D诱导的细胞凋亡的作用。Apoptosis is a complicated antiviral mechanism in host insect cells,which can inhibit viral replication and proliferation. We previously found that the recombinant Antheraea pernyi nucleopolyhedrovirus Ap NPV-Δph / egfp＋was able to infect and express EGFP protein in non-specific host cell Tn-Hi5. In this study,we analyzed the characteristic of Ap NPV-Δph /egfp＋replication and proliferation and the infectivity of progeny virus in Tn-Hi5 cells by real-time fluorescent quantitative PCR and the anti-apoptotic function of Ap NPV-Δph / egfp＋on Tn-Hi5 cells after infection by detecting caspase-3-like protease activity involved in apoptosis signal pathway. The results showed that the DNA copy number of Ap NPV-Δph /egfp＋in Tn-Hi5 cells increased with infection time. The infectious progeny virus of Ap NPV-Δph / egfp＋in Tn-Hi5 cells was generated,but the viral DNA copy numbers were decreased with the generations. Caspase-3-like protease activity induced by actinomycin-D in Tn-Hi5 cells was inhibited by Ap NPV-Δph / egfp＋infection. These results demonstrated that Ap NPV was able to replicate in Tn-Hi5 cells and inhibit apoptosis induced by actinomycin-D.

关 键 词：柞蚕核型多角体病毒 Tn-Hi5细胞 复制增殖 细胞凋亡 

分 类 号：Q939.4[生物学—微生物学]