Illumination of the transmission mechanism of Soil-borne wheat mosaic virus by its vector, Polymyxa graminis and identification of novel resistance factors in barley

谷粒多粘菌传播的土传病毒在世界范围内造成严重的谷物损失。由于病毒在多粘菌休眠孢子内保持数十年的传染性，因此在受污染的土壤上种植谷物只有通过种植抗性品种才经济。不幸的是，对这些病毒的抗性很容易被病毒克服，因此需要创新的抗性策略。确定新的抗性策略的一种方法是阐明多粘菌传播的土壤传播病毒之间相互作用的分子机制，从而确定通过靶向调节参与感染的宿主过程来控制病毒的候选病毒。因此，本研究的目的是确定决定禾粒小麦-土传小麦花叶病毒(SBWMV) -大麦互作的关键因素。由于病毒的传播和运动是影响感染的关键过程，本研究特别阐明了SBWMV传播因子CP-RT和SBWMV运动蛋白（MP）的功能和相互作用网络。预览工作包括对病毒传播过程和病毒在根细胞之间运动的详细分析。特别是，我们将通过高分辨率体内荧光显微镜研究禾本科小麦和大麦根细胞之间的相互作用，特别关注在感染过程和病毒传播过程中发生的膜和细胞骨架重排。此外，我们将揭示病毒MP和CP-RT蛋白用于在疟原虫膜和根细胞之间运输病毒感染性RNA的细胞内和细胞间运输途径。最后，我们将使用生命细胞显微镜和免疫定位结合病毒定量方法来研究载体内是否存在病毒粒子或病毒成分，以及病毒是否能够在多黏菌游动孢子内复制。与这些细胞生物学方法并行，我们将识别和功能表征MP-和cp - rt相互作用的宿主因子在抗病毒抗性中的作用。与SBWMV MP和CP-RT蛋白相互作用的候选宿主因子将通过共免疫沉淀和质谱鉴定，候选宿主因子在病毒感染中的作用将通过在候选基因表达减少的植物中量化载体和病毒水平来确定。这项工作将大大促进我们对禾本科小麦、SBWMV和大麦之间相互作用的分子机制的理解，以及对多粘菌传播的土壤传播病毒的传播和感染机制的理解。通过在天然的SBWMV宿主大麦中识别新的抗性因子，该项目将直接将识别的抗性因子纳入抗性育种计划，从而直接促进农业可持续解决方案的发展。

Polymyxa graminis-transmitted soil-borne viruses cause severe losses in cereals worldwide. As the viruses remain infectious inside polymyxa resting spores for decades, cereal cultivation on contaminated soil is only economic by growing resistant cultivars. Unfortunately, resistances to these viruses are readily overcome by the viruses, thus necessitating innovative resistance strategies. One way to identify novel resistance strategies is the illumination of the molecular mechanisms underlying the interaction between polymyxa-transmitted soil-borne viruses, thereby identifying candidates for virus control by the targeted modulation of host processes involved in infection. Therefore, the objective of the proposed work is the identification of key factors determining the P. graminis - Soil-borne wheat mosaic virus (SBWMV) – barley interaction. As virus transmission and movement are key processes influencing infection, the work specifically elucidates the function and interaction network of the SBWMV transmission factor CP-RT and the SBWMV movement protein (MP). The previewed work includes a detailed analysis of the virus transmission process and virus movement between root cells. In particular, we will investigate interaction between P. graminis and barley root cells by high-resolution in vivo fluorescence microscopy with specific attention on membrane and cytoskeletal rearrangements occurring during the infection process and during virus transmission. Moreover, we will uncover the intra- and intercellular transport pathways the viral MP and CP-RT proteins use to transport viral infectious RNA across the plasmodium membrane and between root cells. Finally, we will use life cell microscopy and immunolocalization in combination with methods for virus quantification to investigate whether virions or viral components are present inside the vector and whether the virus is able to replicate inside polymyxa zoospores. In parallel to these cell biological approaches, we will identify and functionally characterize MP- and CP-RT-interacting host factors in regard to a role in antiviral resistance. Candidate host factors interacting with SBWMV MP and CP-RT proteins will be identified by co-immunoprecipitation followed by mass spectroscopy and a role of the candidate host factors in virus infection will be determined by quantification of vector and virus levels in plants with reduced candidate gene expression. The proposed work will significantly advance our understanding of the molecular mechanisms underlying the interaction between P. graminis, SBWMV and barley in particular and the transmission and infection mechanisms of polymyxa-transmitted soil borne viruses in general. By identifying novel resistance factors in the natural SBWMV host barley, the project will directly contribute to the development of sustainable solutions for agriculture by allowing direct inclusion of the identified resistance factors into resistance breeding programs.