MiR172-APETALA2-like genes integrate vernalization and plant age to control flowering time in wheat

Plants possess regulatory mechanisms that allow them to flower under conditions that maximize reproductive success. Selection of natural variants affecting those mechanisms has been critical in agriculture to modulate the flowering response of crops to specific environments and to increase yield. In the temperate cereals, wheat and barley, the photoperiod and vernalization pathways explain most of the natural variation in flowering time. However, other pathways also participate in fine-tuning the flowering response. In this work, we integrate the conserved microRNA miR172 and its targets APETALA2-like (AP2L) genes into the temperate grass flowering network involving VERNALIZATION 1 (VRN1), VRN2 and FLOWERING LOCUS T 1 (FT1 = VRN3) genes. Using mutants, transgenics and different growing conditions, we show that miR172 promotes flowering in wheat, while its target genes AP2L1 (TaTOE1) and AP2L5 (Q) act as flowering repressors. Moreover, we reveal that the miR172-AP2L pathway regulates FT1 expression in the leaves, and that this regulation is independent of VRN2 and VRN1. In addition, we show that the miR172-AP2L module and flowering are both controlled by plant age through miR156 in spring cultivars. However, in winter cultivars, flowering and the regulation of AP2L1 expression are decoupled from miR156 downregulation with age, and induction of VRN1 by vernalization is required to repress AP2L1 in the leaves and promote flowering. Interestingly, the levels of miR172 and both AP2L genes modulate the flowering response to different vernalization treatments in winter cultivars. In summary, our results show that conserved and grass specific gene networks interact to modulate the flowering response, and that natural or induced mutations in AP2L genes are useful tools for fine-tuning wheat flowering time in a changing environment.Author summary: Reproductive success is essential for species survival, and in cultivated crops to maximize yield. Plants can sense and integrate different internal and environmental signals to ensure that flowering occurs under optimal conditions. In the temperate cereals, wheat and barley, specific mechanisms have evolved that guarantee flowering is promoted by the longer days of spring only after the plants have been exposed to the cold days of winter, a process called vernalization. In this work, we characterized the interactions between the vernalization requirement and a conserved pathway that integrates plant age into flowering regulation. This pathway involves the sequential action of two microRNAs, miR156 and miR172. In spring wheat cultivars, miR156 expression decreases with plant age, while miR172 expression increases. This results in the downregulation of its targets, the APETALA2-like (AP2L) flowering repressors, and the induction of flowering. In winter wheat cultivars, however, the induction of miR172 and the downregulation of AP2L1 is decoupled from miR156, and induction of the VERNALIZATION1 gene by vernalization is required to repress AP2L1 and promote flowering. Our results show that natural or induced mutations in the AP2L genes are useful tools for fine-tuning wheat flowering time in a changing environment.