PomiR164 targets PoLBD25 to promote early flowering in tree peony

Tree peony (Paeonia Section Moutan DC.), a highly valued ornamental plant, has a short and concentrated blooming period, which limits its economic and aesthetic value. MicroRNAs play a crucial role in regulating the flowering time in peony. In this study, we profiled small RNAs across sequential floral stages of tree peony (Paeonia suffruticosa) and identified miR164 as a putative flowering-time regulator. Dual-luciferase reporter assays coupled with co-expression in Nicotiana benthamiana demonstrated that miR164 directly recognizes and promotes the degradation of PoLBD25 transcripts. Mining of the peony genome recovered 25 LBD homologues (PoLBD1–25) that are non-randomly distributed on five chromosomes and were systematically designated according to their physicochemical properties. qRT-PCR revealed that PoLBDs were expressed throughout the flowering stages, with the highest expression during the color-exposure stage. Using homologous transient transformation and heterologous stable transformation, we obtained transgenic peony with altered expression of PomiR164 and PoLBD25 genes and transgenic tobacco overexpressing these genes. PomiR164a overexpression promoted early flowering, and silencing caused delayed flowering. In contrast, PoLBD25 overexpression delayed flowering, and its silencing resulted in early flowering. Taken together, our study elucidates the critical role of the PomiR164–PoLBD25 module in regulating the flowering time in peony, providing a theoretical basis for flowering control and genetic breeding in peony.