近期，中科院武汉病毒所石正丽研究员领导的科研团队采用第二代高通量测序技术，对蝙蝠腺病毒BtAdV-TJM感染蝙蝠细胞系的转录谱做了系统的分析，提供了一套完整的蝙蝠腺病毒复制过程中病毒基因表达的动态分布。结果发表在2013年第1期Journal of Virology上。
腺病毒广泛存在于脊椎动物包括人和非人类灵长动物，能引起人呼吸道和肠道疾病。由于其良好的免疫原性，腺病毒被开发为疫苗和基因治疗的载体。但由于人体已有腺病毒的感染限制了人类腺病毒作为基因载体的广泛应用。因此，需要开发其它动物腺病毒作为新型载体。
该课题组前期用蝙蝠原代细胞分离到一株新型蝙蝠腺病毒（BtAdV-TJM）（Li et al., JVI, 2010）。本研究利用蝙蝠腺病毒感染大卫鼠耳蝠细胞系，采用高通量测序技术对病毒及宿主细胞的转录组进行系统研究。结果表明：随着感染时间的增加，蝙蝠腺病毒在蝙蝠细胞中转录复制加快，在感染后18h达到顶峰。和人腺病毒感染类似，蝙蝠腺病毒复制分为明显早，中，晚期三个阶段。早期基因主要编码和宿主相互作用的一些蛋白，中期基因编码主要编码与自身复制相关的蛋白，晚期基因编码病毒装配和结构蛋白。RT-PCR确认有7个蝙蝠腺病毒基因转录复制不同于人腺病毒。这预示着，蝙蝠腺病毒的转录复制机制可能与人和其他哺乳动物来源的腺病毒不同。同时，该研究对病毒感染后宿主细胞上调及下调基因进行功能分类，主要分为细胞免疫反应、转录、翻译、细胞复制及修复等，其中有79个宿主免疫基因在病毒感染后发生明显上调。上述研究结果为蝙蝠腺病毒载体的开发和蝙蝠免疫系统抗病毒免疫反应的研究提供了基础。
doi: 10.1128/JVI.02332-12
PMC:
PMID:
Deep RNA Sequencing Reveals Complex Transcriptional Landscape of a Bat Adenovirus
Lijun Wu, Peng Zhou, Xingyi Ge, Lin-Fa Wang, Michelle L. Baker and Zhengli Shi
Bat adenoviruses are a group of recently identified adenoviruses (AdVs) which are highly prevalent in bats yet share low similarity to known AdVs from other species. In this study, deep RNA sequencing was used to analyze the transcriptome at five time points following the infection of a bat AdV in a kidney cell line derived from a myotis bat species. Evidence of AdV replication was observed with the proportion of viral RNAs ranging from 0.01% at 6 h to 1.3% at 18 h. Further analysis of viral temporal gene expression revealed three replication stages, the early-stage genes encoding mainly host interaction proteins, the intermediate-stage genes for the DNA replication and assembly proteins, and the late-stage genes for most structural proteins. Several bat AdV genes were expressed at stages that differed from those of their counterpart genes previously reported for human AdV type 2. In addition, single-base resolution splice sites of several genes and promoter regions of all 30 viral genes were fully determined. Simultaneously, the temporal cellular gene expression profiles were identified. The most overrepresented functional categories of the differentially expressed genes were related to cellular immune response, transcription, translation, and DNA replication and repair. Taken together, the deep RNA sequencing provided a global, transcriptional profile of the novel bat AdV and the virus-host interactions which will be useful for the understanding and investigation of AdV replication, pathogenesis, and specific virus-bat interactions in future research.
