Wheat TILLING Mutants Show That the Vernalization Gene VRN1 Down-Regulates the Flowering Repressor VRN2 in Leaves but Is Not Essential for Flowering

Abstract

Most of the natural variation in wheat vernalization response is determined by allelic differences in the MADS-box transcription factor VERNALIZATION1 (VRN1). Extended exposures to low temperatures during the winter (vernalization) induce VRN1 expression and promote the transition of the apical meristem to the reproductive phase. In contrast to its Arabidopsis homolog (APETALA1), which is mainly expressed in the apical meristem, VRN1 is also expressed at high levels in the leaves, but its function in this tissue is not well understood. Using tetraploid wheat lines with truncation mutations in the two homoeologous copies of VRN1 (henceforth vrn1-null mutants), we demonstrate that a central role of VRN1 in the leaves is to maintain low transcript levels of the VRN2 flowering repressor after vernalization. Transcript levels of VRN2 were gradually down-regulated during vernalization in both mutant and wild-type genotypes, but were up-regulated after vernalization only in the vrn1-null mutants. The up-regulation of VRN2 delayed flowering by repressing the transcription of FT, a flowering-integrator gene that encodes a mobile protein that is transported from the leaves to the apical meristem to induce flowering. The role of VRN2 in the delayed flowering of the vrn1-null mutant was confirmed using double vrn1-vrn2-null mutants, which flowered two months earlier than the vrn1-null mutants. Both mutants produced normal flowers and seeds demonstrating that VRN1 is not essential for wheat flowering, which contradicts current flowering models. This result does not diminish the importance of VRN1 in the seasonal regulation of wheat flowering. The up-regulation of VRN1 during winter is required to maintain low transcript levels of VRN2, accelerate the induction of FT in the leaves, and regulate a timely flowering in the spring. Our results also demonstrate the existence of redundant wheat flowering genes that may provide new targets for engineering wheat varieties better adapted to changing environments. Crop yields are strongly associated with flowering time, therefore a precise understanding of the mechanisms involved in the regulation of flowering is required to engineer varieties adapted to new or changing environments. In wheat, most of the natural variation in flowering time is determined by VERNALIZATION1 (VRN1), a gene responsible for the transition of the apical meristem from the vegetative to the reproductive phase. Extended exposures to low temperatures during winter (vernalization) induce VRN1 expression, which promotes flowering in the spring. VRN1 is expressed in the apices and in the leaves, but its role in the leaves is not well understood. Using two sets of VRN1 knock-out mutants, we demonstrate that a central role of VRN1 in the leaves is to maintain low transcript levels of the VRN2 flowering repressor, which allows the production of the mobile FT protein (florigen) required to initiate flowering. Both sets of VRN1 knock-out mutants flowered very late but, eventually, produced normal flowers and seeds, which demonstrates that VRN1 is not essential for wheat flowering. This last result also demonstrates the existence of redundant flowering genes that could provide new targets for engineering flowering time in wheat.