Design of Chimeric Alphaviruses with a Programmed, Attenuated, Cell Type-Restricted Phenotype
Design of Chimeric Alphaviruses with a Programmed, Attenuated, Cell Type-Restricted Phenotype
复制标题
DOI：
10.1128/jvi.00065-11
复制
发表时间：
2011-05-01
影响因子：
5.4
通讯作者：
Frolov, Ilya
中科院分区：
文献类型：
--
作者：
Kim, Dal Young;Atasheva, Svetlana;Frolov, Ilya
披膜病毒科中的甲病毒属包含许多人类和动物病原体。甲病毒的重要性一直被严重低估;然而，基孔肯雅病毒（CHIKV）的流行病在印度次大陆引起了数百万例严重且经常持续的关节炎，近年来已经提高了它们的知名度。尽管持续存在公共卫生威胁，但迄今为止尚未开发出用于甲病毒感染的许可疫苗。在这项研究中，我们已经应用了积累的知识，甲病毒复制和蛋白质功能的病毒-宿主相互作用的机制，介绍了一种新的方法，在设计减毒甲病毒。这些变体是由来自不同地理隔离病毒的基因构建的。所得的活变体编码CHIKV包膜，并且与天然循环病毒相反，缺乏病毒发病机制的重要贡献者：编码在抑制细胞转录和下调细胞抗病毒应答中起作用的蛋白质的基因。为了使这些病毒不能通过蚊子载体传播并差异调节病毒结构蛋白的表达，使它们的复制依赖于来自其他正极性RNA（RNA（+））病毒的内部核糖体进入位点。基因组的合理设计得到了选择程序的补充，该程序使病毒适应组织培养中的复制，并产生变异体，这些变异体（i）表现出不同水平的复制和单个结构蛋白的产生，（ii）有效诱导感染细胞的抗病毒反应，（iii）无法在蚊子起源的细胞中复制，以及（iv）在维罗细胞中有效复制。这种基因组设计的模块化方法适用于构建具有程序化的不可逆减毒表型的其他甲病毒。
The Alphavirus genus in the Togaviridae family contains a number of human and animal pathogens. The importance of alphaviruses has been strongly underappreciated; however, epidemics of chikungunya virus (CHIKV), causing millions of cases of severe and often persistent arthritis in the Indian subcontinent, have raised their profile in recent years. In spite of a continuous public health threat, to date no licensed vaccines have been developed for alphavirus infections. In this study, we have applied an accumulated knowledge about the mechanism of alphavirus replication and protein function in virus-host interactions to introduce a new approach in designing attenuated alphaviruses. These variants were constructed from genes derived from different, geographically isolated viruses. The resulting viable variants encoded CHIKV envelope and, in contrast to naturally circulating viruses, lacked the important contributors to viral pathogenesis: genes encoding proteins functioning in inhibition of cellular transcription and downregulation of the cellular antiviral response. To make these viruses incapable of transmission by mosquito vectors and to differentially regulate expression of viral structural proteins, their replication was made dependent on the internal ribosome entry sites, derived from other positive-polarity RNA (RNA(+)) viruses. The rational design of the genomes was complemented by selection procedures, which adapted viruses to replication in tissue culture and produced variants which (i) demonstrated different levels of replication and production of the individual structural proteins, (ii) efficiently induced the antiviral response in infected cells, (iii) were incapable of replication in cells of mosquito origin, and (iv) efficiently replicated in Vero cells. This modular approach to genome design is applicable for the construction of other alphaviruses with a programmed, irreversibly attenuated phenotype.