In winter wheat (Triticum spp.) and barley (Hordeum vulgare) varieties, long exposures to nonfreezing cold temperatures accelerate flowering time (vernalization) and improve freezing tolerance (cold acclimation). However, when plants initiate their reproductive development, freezing tolerance decreases, suggesting a connection between the two processes. To better understand this connection, we used two diploid wheat (Triticum monococcum) mutants, maintained vegetative phase (mvp), that carry deletions encompassing VRN-1, the major vernalization gene in temperate cereals. Homozygous mvp/mvp plants never flower, whereas plants carrying at least one functional VRN-1 copy (Mvp/-) exhibit normal flowering and high transcript levels of VRN-1 under long days. The Mvp/- plants showed reduced freezing tolerance and reduced transcript levels of several cold-induced C-REPEAT BINDING FACTOR transcription factors and COLD REGULATED genes (COR) relative to the mvp/mvp plants. Diploid wheat accessions with mutations in the VRN-1 promoter, resulting in high transcript levels under both long and short days, showed a significant down-regulation of COR14b under long days but not under short days. Taken together, these studies suggest that VRN-1 is required for the initiation of the regulatory cascade that down-regulates the cold acclimation pathway but that additional genes regulated by long days are required for the down-regulation of the COR genes. In addition, our results show that allelic variation in VRN-1 is sufficient to determine differences in freezing tolerance, suggesting that quantitative trait loci for freezing tolerance previously mapped on this chromosome region are likely a pleiotropic effect of VRN-1 rather than the effect of a separate closely linked locus (FROST RESISTANCE-1), as proposed in early freezing tolerance studies.

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Wheat chromosome 5A plays a key role in cold acclimation and frost tolerance. The major frost tolerance gene Fr-A1 (formerly Fr1) and two loci that regulate the transcription of cold-regulated genes (Cor) have previously been mapped on the long arm of this chromosome. In this study we report the discovery of a new locus for frost tolerance designated Fr-A2. This new locus was mapped on the long arm of chromosome 5A of diploid wheat (T. monococcum), 40 cM from the centromere and 30 cM proximal to the major frost tolerance locus Fr-A1. We found also that frost-tolerant and frost-susceptible T. monococcum parental lines differed in the transcription level of the cold induced gene Cor14b when plants were grown at 15°C. Transcription levels of this gene were measured in each of the recombinant inbred lines and mapped as a QTL that perfectly overlapped the QTL for frost survival at the Fr-A2 locus. This result suggested that frost tolerance in this cross was mediated by differential regulation of the expression of the Cor genes. In a previous study in hexaploid wheat (T. aestivum) we had shown that Cor14b was regulated by two loci located on chromosome 5A, one in the same chromosome region as the T. monococcum Fr-A2 locus and the other one closely linked to Fr-A1. Since CBF transcriptional activators in Arabidopsis regulate Cor genes and are involved in frost tolerance, we decided to localize the cold-regulated CBF-like barley gene Cbf3 on the T. monococcum map. This gene was mapped on the peak of the Fr-A2 QTL for frost tolerance. This result suggests that the observed differential regulation of Cor14b at the Fr-A2 locus is due to allelic variation at the XCbf3 locus, and that this transcriptional activator gene might be a candidate gene for the Fr-A2 frost tolerance locus on wheat chromosome 5A.