Comparative analysis of RNA interference and pattern-triggered immunity induced by dsRNA in response to virus infection
The main goal of this study was to compare the differential efficiency of the antiviral activity triggered by externally delivered virus-specific and nonspecific dsRNAs in plants. RNAi mediated by virus-specific dsRNA is the main antiviral defense mechanism in plants, but nonspecific dsRNA-triggered responses have been documented to play a role in antiviral defense. Using PVX as a model, we have determined that co-inoculation with either virus-specific or nonspecific dsRNA reduced virus accumulation in both inoculated and systemic leaves although at different extent. While the administration of dsRNA specific for the targeted virus induced a potent RNAi-based antiviral response that resulted in highly effective control of viral disease, the degree of interference with PVX-GFP infection afforded by nonspecific dsRNA (PTI) was limited. To identify early biological processes and pathways associated with dsRNA-based immunity, we conducted a global transcriptome RNAseq assay from N. benthamiana leaves inoculated with four different treatments, i.e., dsGFP alone (dsG), PVX-GFP combined with dsGFP (dsG_VG), wild-type (Wt) PVX combined with dsGFP (dsG_V) and bacterial nucleic acid extracts not expressing dsRNA as a control (Ctr). KEGG analysis of differentially expressed genes showed a significant enrichment of terms related to plant-pathogen signaling pathways (KEGG terms Plant-pathogen interaction and MAPK signaling) in all the three treatments with dsRNA. Our results further indicate that the transcriptomic response triggered by dsRNA alone includes canonical immune pathways or genes known to be involved in defense responses, i.e., Ca+2 signaling, ethylene signaling, MAPK signaling, WRKY transcription factors, PR transcriptional factors, NBS-LRR resistance genes, EDS1, and LRR receptor-like kinases, many of which are typical of antimicrobial PTI . Moreover, the transcriptomic response to the homologous combination (dsGFP plus PVX-GFP) had a greater overrepresentation of genes involved in plant-pathogen signaling pathways than the heterologous combination (dsGFP plus Wt PVX), highlighting a quantitative difference between RNAi and PTI immune responses. Overall design: Comparative gene expression profiling analysis of RNA-seq data from Nicotiana bentamina leaves inoculated with four different treatments, i.e., dsGFP alone (dsG), PVX-GFP combined with dsGFP (dsG_VG), PVX combined with dsGFP (dsG_V) and bacterial nucleic acid extracts not expressing dsRNA as a control (Ctr).  Three biological replicates were analyzed for each treatment.