Genome-Wide Identification and Characterization of TAS1/2-like Loci Associated with the miR173–TAS1/2 Regulatory Pathway in Camelina sativa

Trans-acting small interfering RNAs (tasiRNAs) produced from TAS1/2 loci regulate HEAT-INDUCED TAS1 TARGET transcripts and mRNAs encoding pentatricopeptide repeat proteins in Arabidopsis thaliana (A. thaliana). However, their genomic organization in polyploid crops is unknown. Here, we identified and characterized TAS1/2-like loci (TAS1L/2L) in 12 cultivars of the allohexaploid crop Camelina sativa (C. sativa) using conserved miR173/TAS1c 3′D6(–) trigger sites with TAS1/2-derived tasiRNAs. Through phasing analysis with publicly available sRNA-Seq dataset from one of 12 cultivars, we confirmed that these loci tend to produce small RNAs in a phased manner. We also identified that TAS2–TAS1 arrangement in A. thaliana is also conserved in all identified TAS1L/2L loci across the genome of 12 C. sativa’s cultivars. Furthermore, for C. sativa cv. Ames1043, identified TAS2L–TAS1L loci showed a distinct miR173-triggered 21-nucleotide-length phasing, whereas one TAS2L–TAS1L with a disrupted miR173-binding site exhibited weak aberrant phasing. Our findings suggest that the copy number variations in TAS1L-derived tasiR255-like and TAS1c 3′D6(–)-like elements may contribute to cultivar-specific differences in the potential regulation of their target genes. Together, our results suggest a significant conservation of TAS1L/2L and their relative genomic organization of TAS1/2 loci across C. sativa cultivars and provide functioning of miR173-triggered phasing mechanism on TAS1L/2Ls, offering a concise framework for dissecting small-RNA regulatory networks in polyploid crops.