Proton exchange by the vacuolar nitrate transporter CLCa is required for plant growth and nitrogen use efficiency

Abstract

Nitrate is a major nutrient and osmoticum for plants. To deal with fluctuating nitrate availability in soils, plants store this nutrient in their vacuoles. CLCa, a 2NO3-/1H+ exchanger localized to the vacuole in Arabidopsis (Arabidopsis thaliana), ensures this storage process. CLCa belongs to the CLC family, which includes anion/proton exchangers and anion channels. A mutation in a glutamate residue conserved across CLC exchangers is likely responsible for the conversion of exchangers to channels. Here, we show that CLCa with a mutation in glutamate 203 (E203) behaves as an anion channel in its native membrane. To investigate its physiological importance, we introduced the CLCaE203A point mutation into the Arabidopsis clca knockout mutant. These CLCaE203A mutants displayed a growth deficit linked to the disruption of water homeostasis. Additionally, CLCaE203A expression failed to complement the defect in nitrate accumulation of clca and favored higher N-assimilation at the vegetative stage. Further analyses at the post-flowering stages indicated that CLCaE203A expression results in an increase in N uptake allocation to seeds, leading to a higher nitrogen use efficiency compared to the wild type. Altogether, these results point to the critical function of the CLCa exchanger on the vacuole for plant metabolism and development.

Overexpressing NRT2.7 induces nitrate export from the vacuole and increases growth of Arabidopsis

“…In this scenario, a transitory elevated cytosolic nitrate level would decrease nitrate uptake, increase the nitrate efflux from the cells and/or the assimilation pathways through the nitrate reductase (NR). This was confirmed in a recent study demonstrating that the inhibition of nitrate storage in the vacuole leads to an increase of the NR activity and a better NUE (Hodin et al, 2023).…”

Differential Morpho-Physiological, Ionomic, and Phytohormone Profiles, and Genome-Wide Expression Profiling Involving the Tolerance of Allohexaploid Wheat (Triticum aestivum L.) to Nitrogen Limitation

“…CLCa and CLCb are important transporters responsible for vacuolar NO 3 – influx and efflux. , Under low N, reduced CLCa expression and increased CLCb expression contributed to efficient long-distance NO 3 – translocation from roots to shoots, which was undertaken by the elevated expression of NPF7.3 / NRT1.5. Therefore, we investigated the expression of TaCLCa and TaCLCb in the roots of ZM578 and XM26; we found that the low-N-tolerant cultivar ZM578 presented lower expression of TaCLCa-6A and higher expression of TaCLCb-6D than the low-N-sensitive cultivar XM26, particularly under N limitation (Figure B).…”

“…In terms of the molecular mechanisms underlying efficient N use in the three major cereal crop species, namely, rice, maize, and wheat, although great progress has been made in rice 40,41 and maize, 42,43 few noticeable achievements have been obtained in wheat. 35 Plants tend to retain N in source organs when N nutrients are sufficient; when the N status is relatively low, plants remobilize N more efficiently. 44 Therefore, NUE is usually significantly higher under low N conditions than under high N conditions.…”

Chloride Channel Family in the Euhalophyte Suaeda altissima (L.) Pall: Cloning of Novel Members SaCLCa2 and SaCLCc2, General Characterization of the Family

“…Proteins of the CLC (ChLoride Channel) family are found in representatives of all kingdoms [ 17 , 18 , 19 , 20 , 21 ] and play important roles in the transport of Cl − and NO 3 − in plants [ 20 , 22 ]. Contrary to animal CLCs, all of the plant CLCs whose subcellular localization has been studied have been found so far in intracellular membranes, including the tonoplast [ 20 , 22 , 23 ]. The plant CLC family includes Cl − /H + - and NO 3 − /H + -antiporters along with chloride channels [ 20 , 23 ].…”

“…Along with Cl − transport into the organelles, the Cl − /H + antiport stimulates acidification of the organellar lumen through V-ATPase activation mediated by neutralization of positive charges entering the lumen when the ATPase operates [ 26 ]. NO 3 − /H + -antiporter AtCLCa was shown to be involved in NO 3 − storage in vacuoles and regulation of NO 3 − concentrations in the cytosol of Arabidopsis thaliana cells [ 22 , 27 , 28 ]. It was shown that, in glycophytes, salt-sensitive plants, the CLC proteins participated in vesicular trafficking, response to medium salinization, and nitrogen nutrition.…”