Functional characterization of DHN-5, a dehydrin showing a differential phosphorylation pattern in two Tunisian durum wheat (Triticum durum Desf.) varieties with marked differences in salt and drought tolerance

Water-deficit stress caused by drought and soil salinization adversely affects plant growth and crop productivity. Dehydrins are involved in the adaptation to water and osmotic stress. We have identified a wheat dehydrin named DHN-5 that is closely related to the maize RAB17. The full-length cDNA of Dhn-5 gene encodes a putative protein of 227 amino acids and contains 2 conserved lysine-rich-K-segment (EKKGIMDKIKEKLPG) repeats preceded by a stretch of eight serine residues, characteristic of group 2 LEA family. The Northern blot analyses showed a strong accumulation of Dhn-5 transcript in mature wheat embryos and to a lesser extent in ABA and salt-treated seedlings.Interestingly, DHN-5 protein accumulated differentially in two Tunisian durum wheat (Triticum durum Desf.) varieties with marked differences in salt and drought tolerance. By using specific dehydrin antibodies and 2D immunoblot analysis on proteins extracted from mature embryos in these two varieties, a differential phosphorylation pattern of DHN-5 was observed. In the resistant variety (R), beside a basic protein spot, a series of acidic spots were detected whereas in the sensitive variety (S) the acidic spots were weakly detectable. These acidic forms correspond to highly phosphorylated forms of DHN-5, which can be removed by alkaline phosphatase treatment. Accumulation of phosphorylated DHN-5 mainly in the R variety suggests a role of P-DHN-5 in preservation of cell integrity during late embryogenesis and desiccation. Subcellular localization of the DHN-5:GFP fusion protein indicated that DHN-5 would be primarily nuclear, suggesting a nuclear role in wheat osmotic stress response. The observed differential phosphorylation pattern of DHN-5 in the resistant and sensitive wheat varieties could be used as a basis for a molecular screen of tolerance/sensitivity to drought and salt stresses in wheat germplasm.