Zhao Kuijun, Cui Liwang
Department of Entomology, The Pennsylvania State University, 501 ASI Building, University Park, PA 16802, USA.
Virus Genes. 2003 Aug;27(1):93-102. doi: 10.1023/a:1025132720885.
Ascoviruses (AVs) belong to a family of double-stranded DNA viruses that infect Lepidoptera insects and cause the unique pathology of forming virion-containing vesicles in the hemolymph of infected hosts. Virions of AVs are large and contain more than 12 polypeptides. A gene, TnAV-CP, encoding the major structural protein of the Trichoplusia ni ascovirus 2a (TnAV-2a) was cloned by immunoscreening an expression library with antibodies against total TnAV virion proteins. TnAV-CP is an intronless gene with an open reading frame encoding a protein of 455 amino acids. Southern blot showed that it is a single copy gene. A 3.8 kb BamHI fragment containing the complete TnAV-CP gene was cloned and sequenced. Northern analysis detected the transcription of the 1.4 kb TnAV-CP mRNA from 24 h after infection. The predicted TnAV-CP protein was expressed in bacterial expression system and purified to homogeneity. The recombinant protein was used to affinity-purify specific antibodies from the antiserum. The purified antibodies reacted strongly with a single protein of approximately 52 kDa from the total TnAV virion proteins in a Western blot. This protein corresponds to the most abundant structural protein present in the virions of several AVs. Sequence comparison showed that TnAV-CP is most homologous to the putative capsid proteins from AVs infecting noctuid insects, less homologous to that of Diadromus pulchellus ascovirus 4a (DpAV-4a), further supporting the distinction of two subgroups within the family Ascoviridae. Phylogenetic analysis using the putative capsid protein suggested that AVs were closely related to members of Iridoviridae, which corroborated the result based on DNA polymerase delta sequences. The apparent differences between Ascoviridae and Iridoviridae in host range, virion morphology, and genome configuration, and the similarities in genes and methylation of genomic DNA were discussed.
无包涵体病毒（AVs）属于双链DNA病毒家族，可感染鳞翅目昆虫，并在受感染宿主的血淋巴中引起形成含病毒粒子囊泡的独特病理现象。AVs的病毒粒子较大，包含12种以上的多肽。通过用抗TnAV病毒粒子总蛋白的抗体免疫筛选表达文库，克隆了一个编码粉纹夜蛾无包涵体病毒2a（TnAV-2a）主要结构蛋白的基因TnAV-CP。TnAV-CP是一个无内含子基因，其开放阅读框编码一个455个氨基酸的蛋白质。Southern杂交显示它是一个单拷贝基因。克隆并测序了一个包含完整TnAV-CP基因的3.8 kb BamHI片段。Northern分析检测到感染后24小时开始转录1.4 kb的TnAV-CP mRNA。预测的TnAV-CP蛋白在细菌表达系统中表达并纯化至同质。该重组蛋白用于从抗血清中亲和纯化特异性抗体。纯化的抗体在Western印迹中与TnAV病毒粒子总蛋白中一条约52 kDa的单一蛋白强烈反应。该蛋白对应于几种AVs病毒粒子中最丰富的结构蛋白。序列比较表明，TnAV-CP与感染夜蛾科昆虫的AVs的假定衣壳蛋白最同源，与秀丽盘绒茧蜂无包涵体病毒4a（DpAV-4a）的同源性较低，进一步支持了无包涵体病毒科内两个亚组的区分。使用假定衣壳蛋白的系统发育分析表明，AVs与虹彩病毒科成员密切相关，这与基于DNA聚合酶δ序列的结果一致。讨论了无包涵体病毒科和虹彩病毒科在宿主范围、病毒粒子形态和基因组结构方面的明显差异，以及基因和基因组DNA甲基化方面的相似性。
