Asia Ⅱ7和MEAM1烟粉虱传播木尔坦棉花曲叶病毒能力差异的分子机理

Cotton leaf curl Multan virus (CLCuMuV) is one of pathogen causing the fatal cotton leaf curl disease in the world. The disease is transmitted by the whitefly, Bemisia tabaci in a persistent and circulative manner. The virus has invaded, colonized and spread in China. The applicant team has found that whitefly Asia Ⅱ7 cryptic species could transmit CLCuMuV, but MEAM1 cryptic species could not. Its molecule mechanism is unclear. In the study, we will use CLCuMuV isolate GD01, Asia Ⅱ7 and MEAM1 as materials, to screen the protein of Asia Ⅱ 7 interacting with CLCuMuV coat protein by affinity chromatograph, mass spectrometry, DUAL membrane yeast two hybrid technology, and to identify the specific protein of Asia Ⅱ7 whitefly with bimolecular fluorescence complementation (BiFC), Co- Immunoprecipitation, GST pull down test and RNAi technology. We also will analyze the gene express dynamic of the specific protein during the CLCuMuV circulation in Asia Ⅱ7 whitefly, and detect the interaction between CLCuMuV and the protein of Asia Ⅱ7 by immunofluorescence method. And then it would be revealed that the molecule mechanism of ability difference transmitting CLCuMuV between whitefly Asia Ⅱ7 cryptic species and MEAM1 cryptic species. It will provide the scientific basis for exploring the new way to control CLCuMuV. Therefore, the study has extremely important scientific value and application prospect.

木尔坦棉花曲叶病毒（CLCuMuV）是引起棉花毁灭性病害棉花曲叶病的病原之一，由烟粉虱以持久循回方式传播。该病毒已入侵我国，并定殖和扩散。本团队发现，AsiaⅡ7烟粉虱传 CLCuMuV，而MEAM1烟粉虱不传，其分子机理不明。本项目将以CLCuMuV分离物GD01、AsiaⅡ7和 MEAM1烟粉虱为研究对象，应用亲和层析-质谱分析、膜体系酵母双杂交技术，筛选出AsiaⅡ7烟粉虱与CLCuMuV外壳蛋白互作的蛋白；利用BiFC、免疫共沉淀、GST pull down和基因沉默技术，鉴定出AsiaⅡ7烟粉虱传CLCuMuV的特异受体蛋白；分析该蛋白基因在AsiaⅡ7烟粉虱传毒循回中的表达，检测CLCuMuV与受体互作，进而解析AsiaⅡ7和MEAM1烟粉虱传播CLCuMuV能力差异的分子机理，为探索防控该病毒病新途径提供科学依据。因此，本项目具有十分重要的科学价值和应用前景。

木尔坦棉花曲叶病毒（CLCuMuV）是引起棉花毁灭性病害棉花曲叶病的病原之一，由烟粉虱以持久循回方式传播。本团队前期发现AsiaⅡ7烟粉虱传CLCuMuV，而MEAM1烟粉虱不传，其分子机理不明。本项目应用膜体系酵母双杂交从烟粉虱文库中筛选出与CLCuMuV-AV1存在互作的候选蛋白21个；通过生物信息学分析、Y2H和BiFC鉴定出烟粉虱先天免疫相关蛋白(BTB/POZ)和翻译起始因子(eIF1A)与病毒外壳蛋白(AV1)之间存在互作；进一步分析了BTB/POZ对烟粉虱传毒的影响，发现沉默BTB/POZ后可提高AsiaⅡ7烟粉虱获取和积累CLCuMuV的能力，而MEAM1烟粉虱获毒能力呈下降趋势。通过对AsiaⅡ7和MAEM1烟粉虱获取CLCuMuV初期的转录组分析发现，不能有效传毒MAEM1烟粉虱获毒后体内免疫相关反应明显升高；同时对获毒48h后烟粉虱中其他抗病毒免疫信号通路相关基因进行测定，结果同样证实CLCuMuV能诱导MAEM1烟粉虱体内Toll、Imd、Jnk和Jak/STAT等多个抗病毒免疫信号通路相关基因表达。由此得出，CLCuMuV通过靶向抑制介体烟粉虱先天免疫相关BTB/POZ基因的转录和其他抗病毒免疫反应，以隐种专化性方式增强了介体AsiaⅡ7烟粉虱体内病毒的高效积累能力，从而导致介体AsiaⅡ7烟粉虱能有效传播CLCuMuV。本研究从免疫角度解析了AsiaⅡ7和MAEM1烟粉虱差异性传播CLCuMuV的分子机理，为有效控制虫传病毒提供新的分子靶点，为探索防控该病毒病新途径提供科学依据。