水稻黑条矮缩病毒外壳蛋白与灰飞虱传毒介体间的分子互作机制

Rice black-streaked dwarf disease caused by Rice black-streaked dwarf virus (RBSDV) has brought severe damage to rice and corn production in China. Elucidation of the interaction mechanism of RBSDV and its vector is significant for the prevention and control of RBSDV.Because there are no any reported references about the molecular interaction mechanism between RBSDV and small brown planthopper vector and the viral capsid protein is important for infecting its vector, this project plans to screen vector interaction factors, analyze subcellular localization and interaction domains of RBSDV CP and 3-5 vector factors, study functions of 3-5 vector factors on obtaining and transmitting viruses of its vector, explore biochemical mechanism of interaction between RBSDV CP and vector factors, resolve molecular mechanism of interaction between RBSDV and its vector using technologies such as conventional and split-ubiquitin yeast two-hybrid assay,Co-IP, 2-D electrophoresis, Far western blot, GST pull-down, mass spectrometry, gene silencing and bioinformatics. Sequentially, the research results will provide theory bases for elucidating the disaster change mechanism of occurrence and prevalence of rice black-streaked dwarf disease, developing new strategies for preventing and controlling the disease. If this project is successfully finished, it is hopeful to have a new discovery in the research of interaction mechanism of RBSDV and its vector, and the research results also will supply the lack in this research field of rice black-streaked dwarf virus.

水稻黑条矮缩病毒（RBSDV）引起的水稻黑条矮缩病严重影响我国水稻和玉米生产，解析病毒与昆虫介体互作机制是制定新的病毒病防控策略的关键。鉴于目前未见任何有关RBSDV病毒与介体灰飞虱间分子互作机理文献报道及无包膜病毒衣壳蛋白对于病毒入侵介体的重要性，本项目拟通过常规及分裂泛素化酵母双杂交、2-D电泳、免疫共沉淀、Far western blot、GST pull-down、蛋白质谱、基因沉默、生物信息学等技术筛选与RBSDV CP蛋白互作的灰飞虱介体因子、研究CP蛋白与3-5个介体因子的亚细胞定位及互作结构域，分析3-5个介体因子对介体获毒、传毒的作用，探索病毒与介体因子互作的生化机制，解析RBSDV与灰飞虱之间的分子互作机制，从而为阐明水稻黑条矮缩病发生流行的灾变机制及防治该水稻病毒病提供新的理论依据。本项目的开展有望在RBSDV与介体互作研究方面有全新的发现，可填补该研究领域的空白。

水稻黑条矮缩病毒（RBSDV）引起的水稻黑条矮缩病严重影响我国水稻和玉米生产，解析病毒与传毒介体互作机制是制定新的病毒病防控策略的关键。本项目通过常规及分裂泛素化酵母双杂交、免疫共沉淀、GST pull-down、基因沉默、生物信息学等技术筛选获得与RBSDV CP蛋白互作的灰飞虱介体因子14个，研究了RBSDV CP蛋白与LsRACK1、LsVg、LsVgR 和 Lsp5四个介体因子的亚细胞定位及互作结构域，分析了LsRACK1、LsVg、LsVgR 和 Lsp5介体因子对介体获毒、传毒的作用，探索了病毒与这些介体因子互作的生化机制。主要研究结果表明，PKC途径参与灰飞虱抗RBSDV的天然免疫反应，而RBSDV CP与LsRACK1的互作改变了LsRACK1蛋白的亚细胞定位进而抑制PKC天然免疫途径，导致灰飞虱体内RBSDV的积累量增加并有利于RBSDV的传播；LsVg与RBSDV CP互作进而将其带到卵巢组织附近，而通过VgR介导RBSDV CP进入卵巢管；发现带毒灰飞虱体内LsP5基因的表达水平显著上调，沉默LsP5后灰飞虱体内RBSDV病毒的积累量明显下降，表明LsP5 促进RBSDV在灰飞虱体内的复制增殖并有利于RBSDV的传播。这些研究结果为阐明水稻黑条矮缩病发生流行的灾变机制及防治该水稻病毒病提供新的理论依据。