传染性法氏囊病病毒蛋白VP3调控病毒复制的自噬调控机制研究

Infectious bursal disease virus (IBDV) causes a severe immunosuppressive disease with immunological cell damages in poultry industry worldwide. The research of preventive techniques against infectious bursal disease (IBD) caused by IBDV has been never stopped since IBD was discovered. However, how IBDV replication antagonize and evade host responses still unclear. Based on that the original data of proteome and transcriptome of IBDV-infected cells and tissue obtained by ourselves, in forthcoming study, using the monoclonal antibodies to IBDV-encoded proteins as tools, by some techniques including virology, molecular biology, gene engineering, molecular cell biology as well as biochemistry, our proposal shall try to identify viral protein interacted with host factors under different signaling pathways of innate immunity, to map the fine localization of binding domain between IBDV protein and different host proteins, and to analyse the mechanism that the interaction binding domain between IBDV protein and different host proteins regulates the signaling pathways of pattern recognized receptor, mTOR, PKR/NFB and API5/Apaf-1 relevant to autophagy, and to discover the potential anti-IBDV targets and diagnostic markers. The forthcoming research data will.uncover three-dimensional regulating mechanisms that IBDV antagonize host innate immunity to replication and provides an important clue to the innovations of novel IBDV control techniques against IBDV. Therefore our proposal is very important to control IBD prevalence.

传染性法氏囊病毒(IBDV)是以诱导免疫细胞损伤为特征、对养鸡业危害严重的病原体，虽经几十年的控制研究实践，但IBDV复制如何躲避、拮抗宿主应答的控制基础理论研究进展缓慢。本申请课题是我们在国际上首次系统获得IBDV感染宿主比较蛋白组、转录组原始数据和部分揭示宿主固有PRR免疫应答反应机制的基础上，站在病毒蛋白结合宿主因子的全新立体视角，以我们创制的IBDV及宿主因子单克隆抗体为工具，鉴定、发现宿主固有免疫不同路径元件与IBDV VP3进行互作的调控因子，定位IBDV VP3靶向宿主蛋白的互作功能域，分析病毒蛋白VP3互作宿主蛋白功能域通过宿主模式识别受体途径、MTOR通路、PKR/NFB通路、API5/Apaf-1通路调控自噬的机制，阐释IBDV复制的自噬调控机制，揭示潜在的抗IBDV靶标、检测标记，为创新IBDV控制技术研究策略提供理论依据，具有重要意义。

传染性法氏囊病毒发现于上世纪50年代，其引起的鸡传染性法氏囊病严重危害养禽生产，虽然长期使用疫苗免疫，但至今未能得到彻底控制。病毒的复制机制不清是该病长期得不到彻底清除的根本原因。本研究针对IBDV编码蛋白VP3,从宿主防御反应-自噬的角度，开展了以下内容研究：VP3与细胞模式识别受体(PRR)信号途径元件互作的自噬调控，VP3与细胞MTOR信号通路元件互作的自噬调控，VP3与细胞PKR/NFB信号通路元件互作对自噬的调控，VP3与细胞API5/Apaf-1凋亡途径元件互作的自噬调控。通过五年的研究，发现并证明宿主凋亡抑制因子5、3-磷酸肌醇依赖蛋白激酶1的SUMO化修饰特征；发现并证明病毒蛋白VP3的磷酸化修饰位点、泛素化修饰特征；发现并证明了VP3蛋白结合PIK3C3-PDPK1复合物、活化AKT-MTOR信号通路抑制自噬的机制。发现并证明PDPK1结合PIK3C3调控自噬小体生物合成的机制。发现并证明传染性法氏囊病毒VP3蛋白通过靶向TRAF6阻断NF-κB介导的干扰素生产的自噬机制。发现并证明传染性法氏囊病毒VP3蛋白结合API5诱导API5去SUMO化促进病毒复制的机制。传染性法氏囊病毒VP3蛋白通过阻断TBK1-TRAF3复合物的形成和IRF3的激活抑制抗病毒天然免疫。阐明了API5通过自噬受体SQSTM1/P62去泛素修饰而抑制自噬的机制。阐明了EIF4A2通过介导MTOR-ULK1轴诱导的自噬抑制IBDV感染的机制。解析了解旋酶DDX42促进IBDV复制的机制。这些研究结果发现了潜在的药物靶点，为抗病毒药物和疫苗设计提供了理论依据。