Medicago truncatula Natural Resistance-Associated Macrophage Protein1 Is Required for Iron Uptake by Rhizobia-Infected Nodule Cells

30 páginas, 9 figuras, 1 tabla, 13 páginas de explicaciones suplementarias de las figuras, más otras 10 figuras suplementarias,Iron is critical for symbiotic nitrogen fixation (SNF) as a key component of multiple ferroproteins involved in this biological process. In the model legume Medicago truncatula, iron is delivered by the vasculature to the infection/maturation zone (zone II) of the nodule, where it is released to the apoplast. From there, plasma membrane iron transporters move it into rhizobia-containing cells, where iron is used as the cofactor of multiple plant and rhizobial proteins (e.g. plant leghemoglobin and bacterial nitrogenase). MtNramp1 (Medtr3g088460) is the M. truncatula Natural Resistance-Associated Macrophage Protein family member, with the highest expression levels in roots and nodules. Immunolocalization studies indicate that MtNramp1 is mainly targeted to the plasma membrane. A loss-of-function nramp1 mutant exhibited reduced growth compared with the wild type under symbiotic conditions, but not when fertilized with mineral nitrogen. Nitrogenase activity was low in the mutant, whereas exogenous iron and expression of wild-type MtNramp1 in mutant nodules increased nitrogen fixation to normal levels. These data are consistent with a model in which MtNramp1 is the main transporter responsible for apoplastic iron uptake by rhizobia-infected cells in zone II.This work was supported by the Spanish Ministry of Economy and Competitiveness (grant no. AGL–2012–32974 to M.G.-G. and Formación del Personal Investigador fellowship no. BES–2013– 062674 to R.C.-R.), a Marie Curie International Reintegration grant (grant no. IRG–2010–276771 to M.G.-G.), a European Research Council Starting grant (grant no. ERC–2013–StG–335284 to M.G.-G.), and a Ramón y Cajal fellowship (no. RYC–2010–06363 to M.G.-G.).ACKNOWLEDGMENTS We thank Dr. Alonso Rodríguez-Navarro (Universidad Politécnica deMadrid) for providing plasmid pYPGE15, Dr. David Eide (University of Wisconsin, Madison) for the strain fet3/fet4 and its parental strain, Dr. Benoit Lefebvre (Laboratoire des Interactions Plantes Microorganismes) for the pBI101 vector, Dr. Florian Frugier (Institut des Sciences du Vegétal) for the pFRN vector, Dr. Rosario Haro (Universidad Politécnica de Madrid) for the organelle-specific markers, Dr. Regla Bustos (Instituto Nacional de Investigaciones Agroalimentarias) for the agroinfiltration protocol, Dr. Larry Peterson for the Casparian strip visualization method, and Dr. Tsuyoshi Nakagawa (Shiimane University) for the pGWB vectors.Peer reviewe