Genome-wide analysis of phasiRNAs in tea plant reveals a miR482k-PHAS regulatory pathway in response to gray blight

The tea plant (Camellia sinensis) is an important beverage crop, with substantial economic value and proven health benefits. Its predominant distribution in tropical and subtropical regions exposes it to numerous pathogens, making disease resistance crucial for tea cultivation. Phased secondary small interfering RNAs (phasiRNAs) are a class of small non-coding RNAs, generated in a phased manner from specific genomic regions termed phasiRNA-producing loci (PHAS loci). PhasiRNAs have been demonstrated to participate in disease response across various plant species. However, the disease resistance function of phasiRNAs in tea plants remains unexplored. Here, we present a genome-wide characterization of PHAS loci and phasiRNA biogenesis pathways in Camellia sinensis. Using sRNAminer, we analyzed all publicly available tea small RNA sequencing data, identifying 305 PHAS loci and four highly conserved phasiRNA biogenesis pathways in tea plants. Among these, a miR482/2118-triggered 21-nt phasiRNA biogenesis pathway was investigated in detail. We identified 16 members of the miR482/2118 family in tea plants, with miR482k exhibiting significantly higher expression levels. Pathogen infection studies showed that a miR482k-targeted PHAS locus regulated the disease resistance gene CsCAR through phasiRNA production. Further analysis across tea cultivars revealed that the activity of this phasiRNA biogenesis pathway correlates with their disease resistance levels. These findings highlight the critical role of the miR482k-PHAS pathways in disease resistance mechanisms, providing new insights into how phasiRNAs contribute to tea plant immunity and defense responses.