Transcriptional regulation of genes involved in Zn transport after foliar Zn application to Medicago sativa

Zinc (Zn) is an essential micronutrient for both plants and animals, and Zn deficiency is one of the most widespread problems for agricultural production. Although many studies have been performed on the biofortification of staple crops with Zn, few studies have focused on forage crops. In this study the molecular mechanisms of Zn transport-related in Medicago sativa L. were investigated following foliar Zn applications aimed at increasing the accumulation of Zn in edible tissues. Zinc uptake and redistribution between shoot and root were determined following the application of six Zn doses to leaves (0, 0.01, 0.1, 0.5, 1, 10 mg Zn plant-1). Twelve putative genes encoding proteins involved in Zn transport (MsZIP1-7, MsZIF1, MsMTP1, MsYSL1, MsHMA4 and MsNAS1) were identified and the changes in their expression following foliar Zn application were quantified using newly designed RT-qPCR assays. Shoot and root Zn concentration was increased following foliar Zn applications [≥] 0.1 mg plant-1. Increased expression of MsZIP2, MsHMA4 and MsNAS1 in shoots, and of MsZIP2 and MsHMA4 in roots, was observed with the largest Zn dose. By contrast, MsZIP3 was downregulated in shoots at Zn doses [≥] 0.1 mg plant-1. Three functional modules were identified in the M. sativa response to foliar Zn application: genes involved in Zn uptake by cells, genes involved in vacuolar Zn sequestration and genes involved in Zn redistribution within the plant. These results will inform genetic engineering strategies aimed at increasing the efficiency of crop Zn biofortification. One-sentence summaryUpregulation of ZIP2, NASI and HMA4 and downregulation of ZIP3 are associated with Zn sequestration and shoot-to-root translocation in Medicago sativa following foliar Zn biofortification