RNA沉默是植物重要的抗病毒防御机制，DCL、DRB和AGO都是RNA沉默信号途径中的重要蛋白。在前期研究中首次发现拟南芥存在一条DCL4介导、但不依赖于DRB4的抗病毒RNA沉默途径，但其分子机制尚不明确。本项目以TCV病毒诱导沉默载体和拟南芥为研究材料，采用分子生物学和遗传学相结合的方法，首先研究DRB1是否与DCL4相互作用及在体内能否结合TCV，其次，研究同时缺少DCL2和DRB4时DRB1、AGO1、AGO2或AGO7蛋白分别被互补前后产生vsiRNA的变化，第三，研究同时缺少DCL2和DRB4时AGO1、AGO2、AGO7所结合的vsiRNA，从而明确参与该抗病毒RNA沉默途径的DRB蛋白及上述3个AGO蛋白是否参与该途径。研究结果将有助于解析这条新发现的抗病毒RNA沉默分子机制，加深对病毒-寄主之间复杂的相互作用分子机制的认识，亦为探索更有效的病毒防治新策略提供理论依据。

RNA silencing is an important antiviral mechanism in plants. In Arabidopsis Dicer-like (DCL), dsRNA-binding proteins (DRBs), and Argonaute proteins (AGOs) are important proteins in RNA silencing signaling pathway. Our preliminary data shows a novel DCL4-initiated antiviral RNA silencing pathway which is independent of DRB4 protein in Arabidopsis while the molecular details of this novel RNA silencing pathway is still unclear. This specific objective of the proposed research is to identify DRB proteins that function together with DCL4 in the absence of DRB4 and whether AGO1, AGO2, AGO7 participate in the novel DCL4-initiated antiviral RNA silencing pathway which is independent of DRB4 protein. To achieve the objective the proposal employs molecular biology methods as well as molecular genetics research process to determine the following questions: ① whether DRB1 protein interact with DCL4 protein as well as whether it can recruit Turnip crinkle virus （TCV）in vivo; ② what kind of vsiRNA could be recruited by AGO1, AGO2 and AGO7 respectively in Aribidopsis thaliana which is absent of DCL2 and DRB4; ③ the viral small interference RNA triggered by virus induced gene silencing vector CPB-CC-PDS in Aribidopsis thaliana which is in absent of DCL2, DRB4 as well as DRB1, AGO1, AGO2 or AGO7 respectively; ④ the viral small interference RNA triggered by virus induced gene silencing vector CPB-CC-PDS in Aribidopsis thaliana whose DRB1, AGO1, AGO2 or AGO7 has been complemented through gene transformation respectively while remain in absent of DCL2 and DRB4.The proposed research is innovative because it aspires to unravel a unique antiviral defense mechanism in plants that is initiated by DCL4 while independent of DRB4 protein. Our proposed studies are significant because, upon their completion, we will have learned the molecular mechanism of antiviral silencing pathway initiated by DCL4 while independent of DRB4 protein. It will not only deepen our fundamental understanding of the regulation of RNA silencing, but also provide for a more in-depth grasp of the complex host defense mechanisms directed against viral pathogens.