Abstract
Iron is a universal essential micronutrient for primordial metabolism (respiration, photosynthesis, electron transfer etc ...). Although abundant, it is not bio-available in aerobic environments and can generate ROS. Its acquisition and transport require the control of its redox status to limit its toxicity. In addition to iron reduction to mobilize it, eukaryotic models have ferroxydases associated with transporters to transport iron in the form of Fe(III), less toxic than Fe(II). In higher plants, iron reduction has received particular attention in the context of the fight against iron deficiency in culture soil for crops, but the study of ferroxydation has remained superficial until very recently. Ferroxydases exist in Arabidopsis but their role, if any, in iron homeostasis has not yet been elucidated. In Arabidopsis, there are several potential ferroxydases including a small subfamily of Multicopper-oxidases (MCO1,2,3) which has a great similarity with the ferroxydase model FET3 of S.cerevisiae yeast. In my work, I was able to highlight that among them, MCO3 has a ferroxydase activity and complements the fet3fet4 mutant; MCO1 and MCO3 are apoplastic proteins. To analyze the physiological role of these genes, I generated a triple mutant mco1,2,3 by CRISPR/Cas9. Using iron imaging approaches, I was able to show that the mesophyll cells of the triple mutant accumulate more iron than the wild. Specifically, more iron is observed in chloroplasts and in vacuoles. These observations were confirmed by analysis of the expression of genes related to intracellular iron transport. In the triple mutant mco1,2,3, the VTL2 and VTL5 genes, encoding vacuolar iron influx transporters, are induced while the vacuolar efflux transporter, NRAMP4, is repressed. This over-accumulation of iron could lead to some toxicity, also suggested by a higher production of reactive oxygen species in the triple mutant. I propose that these AtMCOs would form an additional barrier in the management of excess iron at mesophyll cells surface, thus influencing iron distribution at the cellular level by limiting its entry into leaf cells. AtMCOs thus enlarge the small cohort of Arabidopsis known ferroxydases (with LPR1 and LAC12 (Muller et al 2015; Bernal et al 2021)) whose integration into Arabidopsis iron homeostasis remains to be clarified.