Studies of vacuolar transporters controlling nutrient storage and ion detoxification in Arabidopsis thaliana

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The vacuole and TGN/EE (trans-Golgi network/early endosome) are intracellular compartments essential for nutrient accumulation and cytosolic ion detoxification by compartmentalizing the different cellular components. The multiple roles of these compartments are tightly linked to the activities of transporters present in their membranes. It is assumed that transporters use the energy of the electrochemical gradient generated by proton pumps such as the V-type H+-Adenosintriphosphatases (V-ATPase). The studies on V-ATPase suggest a specificity of action: a vha-a2 vha-a3 knock-out Arabidopsis mutant affected in vacuolar V-ATPase activity specifically reduces the vacuolar storage of nitrate but not the sequestration of chloride whereas the sensitivity to NaCl salt stress increases in a knock-down mutant of vha-a1, a mutant with reduced V-ATPase activity at the TGN/EE. The project aims at understanding the molecular basis of this functional specification through the analysis of interaction between V-ATPase and CLC anion transporters in Arabidopsis. This latter protein family is characterized by interesting features: (1) they can be channel (passive transporter) or anion/proton exchangers (active transporter) although all the members are structurally very close, (2) they can be either more selective for chloride or nitrate depending of a presence of a serine/proline in their selectivity filter and, (3) members are present on both TGN/EE and vacuolar membranes. The project will be divided in three Tasks: Task 1 will investigate the specificities/redundancies of AtCLCs localised in the vacuole and the TGN/EE. The two other Tasks will analyze the co-regulation between V-ATPase and AtCLCs in the vacuolar (Task 2) and TGN/EE membrane (Task 3). The project will take advantage of the expertise of two expert groups in the field of primary and secondary transport in Arabidopsis thaliana. The French partner will provide knowledge in molecular physiology and electrophysiology which will be complemented by cell biological and omics approaches of the German partner. The cooperation between the two laboratories will allow an in-depth analyses of the interaction between V-ATPase and AtCLC at the molecular, cellular and whole-plant level, taking different environmental fluctuations (nitrate concentrations, drought and NaCl stress) into account. Overall, this project will gain new insights into the coordination of primary and secondary transport processes at the tonoplast and TGN/EE and holds the potential for developing future strategies to improve crops for nutrient assimilation and tolerance to abiotic stress.