[en] Using a transient plant system, it was previously found that the suppression of Cucumber mosaic virus (CMV) 2b protein relies on its double-strand (ds) RNA binding capacity, but it is independent of its interaction with ARGONAUTE (AGO) proteins. Thus, the biological meaning of the 2b-AGO interaction in the context of virus infection remains elusive. In this study, we created infectious clones of CMV mutants that expressed the 2b functional domains of dsRNA or AGO binding and tested the effect of these CMV mutants on viral pathogenicity. We found that the mutant CMV2b(1-76) expressing the 2b dsRNA-binding domain exhibited the same virulence as wild-type CMV in infection with either wild-type Arabidopsis or rdr1/6 plants with RDR1- and RDR6-deficient mutations. However, remarkably reduced viral RNA levels and increased virus (v)siRNAs were detected in CMV2b(1-76)-infected Arabidopsis in comparison to CMV infection, which demonstrated that the 2b(1-76) deleted AGO-binding domain failed to suppress the RDR1/RDR6-dependent degradation of viral RNAs. The mutant CMV2b(8-111) expressing mutant 2b, in which the N-terminal 7 amino acid (aa) was deleted, exhibited slightly reduced virulence, but not viral RNA levels, in both wild-type and rdr1/6 plants, which indicated that 2b retained the AGO-binding activity acquired the counter-RDRs degradation of viral RNAs. The deletion of the N-terminal 7 aa of 2b affected virulence due to the reduced affinity for long dsRNA. The mutant CMV2b(18-111) expressing mutant 2b lacked the N-terminal 17 aa but retained its AGO-binding activity greatly reduced virulence and viral RNA level. Together with the instability of both 2b(18-111)-EGFP and RFP-AGO4 proteins when co-expressed in Nicotiana benthamiana leaves, our data demonstrates that the effect of 2b-AGO interaction on counter-RDRs antiviral defense required the presence of 2b dsRNA-binding activity. Taken together, our findings demonstrate that the dsRNA-binding activity of the 2b was essential for virulence, whereas the 2b-AGO interaction was necessary for interference with RDR1/6-dependent antiviral silencing in Arabidopsis.
Fang, Yuan-Yuan; State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences Beijing, China
Zhao, Jian-Hua; State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences Beijing, China
Liu, Shangwu ; State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of SciencesBeijing, China, Virus-free Seedling Research Institute, Heilongjiang Academy of Agricultural SciencesHarbin, China
Wang, Sheng; State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences Beijing, China
Duan, Cheng-Guo; State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences Beijing, China
Guo, Hui-Shan; State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of SciencesBeijing, China, College of Life Sciences, University of Chinese Academy of SciencesBeijing, China
Language :
English
Title :
CMV2b-AGO Interaction Is Required for the Suppression of RDR-Dependent Antiviral Silencing in Arabidopsis.
Publication date :
2016
Journal title :
Frontiers in Microbiology
eISSN :
1664-302X
Publisher :
Frontiers Media S.A., Switzerland
Volume :
7
Issue :
AUG
Pages :
1329
Peer reviewed :
Peer Reviewed verified by ORBi
Funders :
Ministry of Science and Technology of the People's Republic of China [CN] NSCF - National Natural Science Foundation of China [CN]
Funding text :
This study was supported by the Ministry of Science and Technology (2014CB138402) and the Natural Science Foundation of China (No.91519327)
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