乙型脑炎病毒3'UTR短重复序列对病毒种属特异性的影响及机制研究

乙型脑炎病毒（JEV）在脊椎动物与蚊虫中循环传播，是重要的人类病原体。黄病毒3ʹUTR在病毒的宿主差异复制及跨种传播中发挥重要作用，蚊传黄病毒中 SL/DB重复结构与种属特异性相关，但短重复序列DRs的功能尚不清楚。申请人前期研究发现黄病毒DRs影响sfRNA的产生，并且在不同种属细胞中产生不同形式的sfRNA，提示DRs可能发挥种属特异性功能。本项目通过比较JEV短重复序列缺失及点突变在不同种属细胞中的复制和适应性特征，明确短重复序列如何发挥种属特异性。通过检测突变体RNA与宿主蛋白的相互作用及在不同种属细胞中sfRNA的产生，阐明短重复序列发挥种属特异性的机制，继而通过蚊虫感染实验证实其对蚊虫感染和传播的影响。本项目旨在明确3ʹUTR短重复序列对蚊传黄病毒种属特异性的影响及作用机制，为减毒疫苗研发、控制虫媒病毒的传播提供理论依据。

Japanese encephalitis virus (JEV) is a serious pathogen to public health, which circulates between vertebrates and mosquitoes. Flavivirus 3ʹUTR plays an important role in virus host species-specific replication and cross-species transmission. Repeated stem loop (SL) and dumbbell (DB) structures in mosquito-borne flavivirus (MBFV) are key structures in host species-specific replication, while the function of short direct repeats (DRs) are still unclear. Our recent study found that flavivirus DRs were involved in the production of sfRNA and DRs deletion mutants produced different sfRNA patterns in mammalian and mosquito cells, suggesting that DRs maybe have host species-specific function. In this project, by characterizing replication and adaptability of JEV DRs deletion or site mutants in different species cells, we will confirm that whether and how DRs play a role in species specificity. Further, the mechanism of DRs on species specificity will be identified by detecting the interaction of mutant 3’UTR RNA and host specific proteins and the production of sfRNA pattern of mutants in different species cells. In addition, the efficiency of infection and transmission of the mutant virus in mosquitoes will be detected by mosquito infection assay. This project aims to address the effect and mechanism of 3ʹUTR DRs on MBFV species specificity, which will provide theoretical basis for the development of attenuated vaccine and controlling the spread of arboviruses.