Potato virus Y infection alters small RNA metabolism and immune responses in tomato

Potato virus Y (PVY) is an important pathogen of solanaceous crops. The PVYC-to isolate induces stunting and foliar symptoms on tomato, but new vegetation displays symptom recovery at a later stage. In order to investigate the role of micro(mi)RNA-regulated mechanisms in tomato defenses against PVY, we performed small RNA sequencing from healthy and PVYC-to infected tomato plants at 21 and 30 days-post-inoculation (dpi). A total of 756 unique miRNA sequences were obtained, among which 118 canonical miRNA sequences, many isomiR variants and 22 novel miRNAs. 46 mature miRNAs were overexpressed in infected vs. healthy plants, and showed a consistent alteration at both 21 and 30 dpi, whereas only a few miRNAs were underexpressed. Increased accumulation of isomiRs was correlated with viral infection. Among miRNA-targets, enriched gene ontology categories included resistance (R) gene families such as NBS-LRR receptors, RLPs and RLKs that are post-transcriptionally controlled by sly-miR482, sly-miR6023, sly-miR6024, sly-miR6027. These 22-nt miRNAs were shown to target R genes and trigger the production of 21-nt secondary phased sRNAs (phasiRNAs). 500 phasiRNA-generating loci were identified, mostly active in PVY-infected tissues. Our data demonstrate that sRNA-regulated host responses, encompassing quantitative miRNA alteration, diversification within miRNA families and miRNA-driven phasiRNA accumulation, regulate R genes, suggesting a functional role in the recovery phenotype.