媒介伊蚊内源性病毒序列对外源性病毒感染的作用及分子机制研究

Mosquito-borne diseases such as zika, dengue, and yellow fever constitute a major public health problem, because they are responsible for a considerable amount of morbidity and mortality. Vector control is a key means of combating mosquito-borne diseases. The dynamic interactions between vector and pathogen play a key role in viral persistent and non-fatal infection in vector, which is an necessary for vector-to-human transmission. An Endogenous viral elements(EVEs) is a DNA sequence derived from a virus, and present within the genome of a non-viral organism. EVEs widespread appear in the animal genome, parts of them even are involved in host antiviral immunity. We previously identified endogenous Dengue virus (DENV) elements in Aedes albopictus Foshan strain genome, as well as endogenous Mosquito densovirus (MDV) elements in Aedes aegypti Aag2 cell genome. Furthermore, reads of transcriptome and small RNA were derived from these EVEs. Whether these EVEs are involved in mosquito antiviral immunity, and the major molecular mechanisms still need the further exploration. So, base on ours research foundation and scientific questions, the main research objective of our proposal focus on (1) Analyze the expression profile of the EVEs derived genes before and post virus infection base on RNA-sequencing technology. (2) Knocking in and/or knocking out candidate EVEs derived genes to verify their functions in viruses infected mosquito vector. (3) Exploration of the major molecular mechanisms of EVEs derived genes in antiviral immunity using ChIP-Seq, RIP and Co-IP methods. Results from this study will allow us to detect the key factors in mosquito antiviral immunity, and provide new strategy for mosquito control.

媒介伊蚊能传播许多重要的病毒性疾病，病毒与蚊媒之间的相互作用是病毒在蚊媒中的持久、非致命性感染，并最终传染给人的重要条件。内源性病毒序列（EVEs）不仅在动物基因组中广泛存在，部分EVEs还可以通过多种方式参与宿主抗病毒免疫途径。申请者前期在白纹伊蚊与埃及伊蚊Aag2细胞基因组中鉴定到内源性的登革病毒与蚊浓核病毒序列，且这两种EVEs存在对应的mRNA与小RNA转录。这些EVES序列是否参与到蚊虫与病毒的相互作用中，其机制如何？根据以上科学问题和研究基础本项目拟：①利用转录组与小RNA测序，分析外源病毒感染蚊媒前后EVEs基因的表达特征；②利用基因敲除或干扰的方法，探索EVEs基因对外源病毒感染的影响；③通过ChIP-Seq、RIP、Co-IP等方法，阐明EVEs基因参与抗病毒免疫的分子机制。研究结果将有助于寻找媒介伊蚊抗病毒感染的关键因子，为其传染病的防治提供新的靶点和思路。

病毒与媒介两者存在着复杂相互作用，互相博弈，协同进化，其作用结果决定了病毒是否能吸附、穿入媒介细胞，并在其中脱壳、转录、翻译、复制、装配、成熟、释放最终完成生活史，也决定了媒介宿主的媒介能量、媒介效能、并影响了蚊的寿命、摄食行为等。内源性病毒序列（EVEs）不仅在动物基因组中广泛存在，部分EVEs还可以通过多种方式参与宿主抗病毒免疫途径。申请者发现研究病原体与媒介关系的最常用蚊虫细胞Aag2存在着几乎完整的蚊浓核病毒序列，本项目通过研究进而发现：①我国野外蚊虫广泛感染浓核病毒。②野生浓核病毒在蚊虫细胞内与登革病毒产生相互影响，并在体内对白纹伊蚊的登革病毒的媒介效能产生影响。③对于Aag2细胞，其内源性病毒序列可以转录与表达NS1蛋白 ④利用干扰内源性病毒mRNA的方法，发现感染登革病毒与寨卡病毒后，细胞内或培养上清中病毒的基因拷贝数DENV-2显著上升，ZIKV则下降，即产生不同的影响。本研究阐明浓核病毒可以参与蚊虫的异源的病毒相互作用，从而影响虫媒黄病毒，也同时提供参考Aag2细胞是否是研究病毒与宿主相互作用的最优选择细胞系值得深入评估。研究结果将有助于寻找媒介伊蚊抗病毒感染的关键因子，为其传染病的防治提供新的靶点和思路。