Evolution of Arabidopsis MIR genes generates novel microRNA classes

In Arabidopsis, canonical 21-nt miRNAs are generated by Dicer-like (DCL) 1 from hairpin precursors. We have identified a novel class of functional 23- to 25-nt long-miRNAs that is generated independently from the same miRNA precursors by DCL3. Long-miRNAs are developmentally regulated and in some cases have been conserved during evolution implying that they have biological functions. Plant microRNA genes (MIR) have been proposed to evolve by inverted duplication of the target gene. We found that recently evolved MIR genes consistently give rise to long-miRNAs, while ancient MIR genes give rise predominantly to canonical miRNAs. Transcripts from inverted repeats representing evolving proto-MIR genes were processed by DCL3 into long-miRNAs and also by DCL1, DCL2 or DCL4 depending on hairpin stem length to produce different sizes of miRNAs. Our results suggest that evolution of MIR genes is associated with gradual, overlapping changes in DCL usage resulting in specific size classes of miRNAs.