类病毒交叉保护的分子机理研究

Cross-protection is an important strategy to control plant diseases caused by virus and viroid infection.One mechanism of cross-protection involved the coat protein (CP) of attenuated virus strains.The CP of the protecting strain present in the cell would encapsidate the RNA of the challenging strain before it could replicate.However,the involvement of CP in cross-protection could not explain case where viroids were used as protective strains,because viorids cannot encode any proteins. In this work, Potato spindle tuber viroid and will be used to study the mechanism(s) of viorid cross-protection. Tagging viroids with different fluorescent proteins (e.g. green fluorescent protein) will be applied to determine viroid distribution in systemically infected N. benthamiana tissues mixly infected by closely related viroids.This could reveal mutual exclusion of closely related viroids.Furthermore, the constructs concluding hairpin structures of genes associated with RNA silencing will be transformed into N.benthamiana to obtain transgenic plants which are defective in RNA silencing.Thus,combining tagged viroids with the use of plants compromised in the RNA silencing machinery (e.g. dcl mutants) may answer the involvement of RNA silencing with cross-protection, based on detections of viroids in tissues of both transgenic and wild plants.The results of this research will determine the mechanism of viroid cross-protecion and help to further understand molecular mechanism of phenomenon of plant viruses cross-protection. Furthermore, they will be useful for improving application of cross-protection to control plant viral diseases.

交叉保护是防控植物病毒和类病毒病害的重要手段。虽然已经证实病毒的弱毒株系可以通过抑制强毒株系的脱壳实现交叉保护，但是由于类病毒不能编码任何蛋白质，该机制难以解释类病毒的交叉保护现象。本项目以马铃薯纺锤块茎类病毒(PSTVd)为研究对象，首先，应用荧光标记技术比较分析类病毒的不同株系在组织水平上的分布，进而明确类病毒不同株系间的相互排斥作用；其次，制备含有与RNA沉默路径相关的蛋白质DCLs和RdRp6的基因的发夹结构的载体，将其转入本生烟(N.benthamiana)，获得RNA沉默被抑制的转基因植株；最后，利用已获得的转基因植株，比较RNA沉默被抑制前后类病毒不同株系在组织水平上分布的异同以及交叉保护现象的变化，从而明确RNA沉默在交叉保护中的作用。研究结果不仅可以揭示类病毒交叉保护的分子机理，有利于进一步解释植物病毒交叉保护现象，而且对利用交叉保护防治植物病毒病害具有重要的现实意义。

交叉保护是防控植物病毒和类病毒病害的重要手段。虽然已经证实病毒的弱毒株系可以通过抑制强毒株系的脱壳实现交叉保护，但是由于类病毒不能编码任何蛋白质，该机制难以解释类病毒的交叉保护现象。本项目以马铃薯纺锤块茎类病毒(PSTVd)为研究对象，首先，应用荧光标记技术比较分析类病毒的不同株系在组织水平上的分布，进而明确类病毒不同株系间的相互排斥作用；其次，制备含有与RNA沉默路径相关的蛋白质DCLs和RdRps的基因的发夹结构的载体，将其转入本生烟(N.benthamiana)，获得RNA沉默被抑制的转基因植株；最后，利用已获得的转基因植株，比较RNA沉默被抑制前后类病毒不同株系在组织水平上分布的异同以及交叉保护现象的变化，从而明确RNA沉默在交叉保护中的作用。研究结果不仅可以揭示类病毒交叉保护的分子机理，有利于进一步解释植物病毒交叉保护现象，而且对利用交叉保护防治植物病毒病害具有重要的现实意义。