Heat-responsive microRNAs and phased small interfering RNAs in reproductive development of flax

Plants will face increased heat stress due to rising global temperatures. Heat stress affects plant reproductive development and decreases productivity; however, the underlying molecular mechanisms of these processes are poorly characterized. Plant small RNAs (sRNAs) have important regulatory roles in plant reproductive development following abiotic stress responses. We generated sRNA transcriptomes of reproductive bud stages at three different time points to identify sRNA-mediated pathways responsive to heat stress in flax (Linum usitatissimum). With added sRNA transcriptomes of vegetative tissues, we comprehensively annotated miRNA and phasiRNA-encoding genes (PHAS) in flax. We identified 173 miRNA genes, of which 42 are newly annotated. Our analysis revealed that 141 miRNA genes were differentially accumulated between tissue types, while 18 miRNA genes were differentially accumulated in reproductive tissues following heat stress, including members of miR482/2118 and miR2275 families, known triggers of reproductive phasiRNAs. Furthermore, we identified 68 21-PHAS flax loci from protein-coding and noncoding regions, four 24-PHAS loci triggered by miR2275, and 658 24-PHAS-like loci with unknown triggers, derived mostly from noncoding regions. The reproductive phasiRNAs are mostly downregulated in response to heat stress. Overall, we found that several previously unreported miRNAs and phasiRNAs are responsive to heat stress in flax reproductive tissues.