Final Report Summary - MENOMED (Metal homeostasis in nodulated Medicago truncatula) It is estimated that the overuse of nitrogen fertilizers costs the European Union €70-300 billion/yr, and that the industrial synthesis of these chemicals represents 15 % of the global greenhouse emissions. In order to avoid this, sustainable agriculture strategies are being encouraged, crop rotation with legumes among them. Legumes are one of the most important crops worldwide and the main source of vegetal protein for animal and human diet. They are used in crop rotation cycles due to their ability to enrich soils in nitrogen, diminishing the need for fertilizers. Nitrogen fixation occurs at differentiated root organs, the nodules, where an endosymbiosis with bacteria generally known as rhizobia is established. The rhizobia within the nodule cells differentiate in bacteroids and express the enzymatic machinery necessary for nitrogen fixation. Many of these enzymes are metalloproteins. In this research project we are interested on how legume metal homeostasis is affected by nodulation and on how metals are exchanged between the symbionts, since metal deficiency is prevalent in crops and has severe effects on nodulation. By understanding how this process is regulated in the nodule and which elements are involved in it, we expect to improve legume metal nutrition and consequently, increase nitrogen fixation rates and productivity.Towards this goal we have determined the path that metals follow from soil to reach the nodule. This has been achieved using synchrotron-based X-ray fluorescence in collaboration with researchers at Argonne National Laboratory (USA). Our results indicate that iron, and likely other transition metals, are released from the vasculature into the apoplast of the infection/differentiation zone of the nodule.This would ensure a maximum availability of the essential metal oligonutrients at the same time and place as the apoproteins involved in nitrogen fixation are being synthesized. To determine the transporters involved in metal uptake and delivery to the symbiosome, we have used a transcriptomic approach identifying ten genes involved in metal homeostasis upregulated more than seven fold in the nodule, some of them nodule-specific. Several of these transporters, such as MtNramp1, are located in the plasma membrane of cells in the infection/differentiation zone of the nodule.Consequently with their putative importance in symbiotic nitrogen fixation, mutation of these transporters result in loss of nitrogen fixation capabilities due to essential metal cofactors not reaching their designated target apoproteins. All these results open a new venue to improve nitrogen fixation capabilities in legumes by optimizing metal delivery to the nodules.