MicroRNAs in durum wheat seedlings under chronic and short-term nitrogen stress
Nitrogen is an essential macronutrient for plant growth and reproduction. In durum wheat, an appropriate nitrogen soil availability is essential for an optimal seed development. miRNAs contribute to the environmental change adaptation of plants through the regulation of important genes involved in stress processes. In this work, nitrogen stress response was evaluated in durum wheat seedlings of Ciccio and Svevo cultivars. Eight small RNA libraries from leaves and roots of chronically stressed plants were sequenced to detect conserved and novel miRNAs. A total of 294 miRNAs were identified, 7 of which were described here for the first time. The expression level of selected miRNAs and target genes was analyzed by qPCR in seedlings subjected to chronic (Ciccio and Svevo, leaves and roots) or short-term (Svevo roots) stress conditions. Some miRNAs showed an immediate stress response, and their level of expression was either maintained or returned to a basal level during a long-term stress. Other miRNAs showed a gradual up- or downregulation during the short-term stress. The newly identified miRNA ttu-novel-106 showed an immediate strongly downregulation after nitrogen stress, which was negatively correlated with the expression of MYB-A, its putative target gene. PHO2 gene was significantly upregulated after 24–48-h stress, corresponding to a downregulation of miR399b. Ttu-miR399b putative binding sites in the 5′ UTR region of the Svevo PHO2 gene were identified in the A and B genomes. Both MYB-A and PHO2 genes were validated for their cleavage site using 5′ RACE assay.
KeywordsAbiotic stress Triticum turgidum subsp. durum seedlings miRNA expression Target genes Leaves and roots
We thank Donatella Danzi, Domenico De Paola, and Anita Morgese for their assistance.
This research was supported by the MIUR projects PON01_01145 ISCOCEM “Sviluppo tecnologico e innovazione per la sostenibilità e competitività della cerealicoltura meridionale”; PON PONa3_00025 prot. no. 1424/29 BIOforIU “Infrastruttura multidisciplinare per lo studio e la valorizzazione della Biodiversità marina e terrestre nella prospettiva della Innovation Union”; PRIN 2010–2011 DD 23/10/2012 n. 719 “Identificazione e caratterizzazione di geni utili ad incrementare la produttività e sostenibilità del frumento duro”; and by the Puglia Region D.G.R. n. 1719 of 02.02.2011, Project Code 73 BioNet-PTP “Biodiversità per la valorizzazione e sicurezza delle produzioni alimentari tipiche pugliesi”.
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