Profiling of small RNAs derived from cucumber mosaic virus in infected Nicotiana benthamiana plants by deep sequencing

In plants, RNA silencing is a conserved mechanism underlying antiviral immunity. To investigate antiviral responses in Nicotiana benthamiana, we analyzed the profiles of the virus-derived small RNAs (vsRNAs) in wild-type N. benthamiana and NbRDR6 mutant plants infected with the cucumber mosaic virus (CMV) 2b-deficient mutant. We observed that NbRDR6 regulates RNA silencing by producing vsRNAs that trigger an effective antiviral response, while NbRDR1 may nonredundantly and synergistically function with NbRDR6 to mediate immune responses. The vsRNAs in N. benthamiana and NbRDR6 mutant plants mainly comprised 21 or 22 nucleotides, and mostly consisted of a 5′-terminal adenine. Additionally, NbAGO2 expression was significantly up-regulated in N. benthamiana and NbRDR6 mutant plants, suggesting that NbAGO2 is closely associated with the antiviral activities of vsRNAs. The distribution of vsRNAs in the CMV genome was biased toward RNA sense strands in both N. benthamiana and NbRDR6 mutant plants. These findings indicate the specific and conserved antiviral immunity in Nicotiana benthamiana.