The MIR157-SPL15 module regulates flowering and inflorescence development in Arabidopsis thaliana under short days...

The plant life cycle progresses through distinct phases defined by the morphology of the organs formed on the shoot. In Arabidopsis, age-dependent reduction in the related microRNAs miR156 and miR157 controls transitions from juvenile to adult vegetative phase and from adult to reproductive phase. However, whether these miRNA isoforms have specific contributions remains unclear. To compare their roles, we used Trans-kingdom, rapid, affordable Purification of RISCs (TraPR) for small RNA sequencing, CRISPR-Cas9, and confocal imaging. We show that in shoot apices, levels of miR156 in RNA-induced silencing complexes (RISCs) decline more rapidly than those of miR157, so that miR157 is more abundant than miR156 in RISCs of older plants undergoing floral transition and inflorescence development. Accordingly, confocal microscopy analysis showed that MIR156A and MIR156C are not detectably expressed in shoot apices of older plants, whereas at this stage MIR157C is expressed in upper stems, and MIR157D is expressed in axils of inflorescence leaves. Arabidopsis flowers much earlier under long days (LDs) than short days (SDs). CRISPR-induced mir157c mutations but not mir156ac mutations accelerated flowering under SDs, and altered inflorescence leaf morphology. Notably, mir157c mutations also caused early flowering in Arabis alpina, a perennial relative of Arabidopsis, indicating that the repression of flowering by this paralogue is evolutionarily conserved. SPL15 transcription factor promotes flowering under SDs and its mRNA is a target of miR156/miR157. SPL15 abundance was higher in apices of mir157 cd mutants under SDs, and spl15 mutations partially suppressed the early flowering of mir157c mutants and this effect was enhanced by spl4 mutation. We show by genetic analysis that the florigen FLOWERING LOCUS T overcomes the requirement for SPL15 in LDs but not SDs, contributing to the increased importance of the MIR157C-SPL15 module under SDs. We conclude that MIR157 genes have important evolutionarily conserved roles in repressing floral transition and modulating inflorescence development of older plants under SDs.