Chloride (Cl⁻) has traditionally been considered a micronutrient largely excluded by plants due to its ubiquity and abundance in nature, its antagonism with nitrate (NO₃⁻), and its toxicity when accumulated at high concentrations. In recent years, there has been a paradigm shift in this regard since Cl⁻ has gone from being considered a harmful ion, accidentally absorbed through NO₃⁻ transporters, to being considered a beneficial macronutrient whose transport is finely regulated by plants. As a beneficial macronutrient, Cl⁻ determines increased fresh and dry biomass, greater leaf expansion, increased elongation of leaf and root cells, improved water relations, higher mesophyll diffusion to CO₂, and better water- and nitrogen-use efficiency. While optimal growth of plants requires the synchronic supply of both Cl⁻ and NO₃⁻ molecules, the NO₃⁻/Cl⁻ plant selectivity varies between species and varieties, and in the same plant it can be modified by environmental cues such as water deficit or salinity. Recently, new genes encoding transporters mediating Cl⁻ influx (ZmNPF6.4 and ZmNPF6.6), Cl⁻ efflux (AtSLAH3 and AtSLAH1), and Cl⁻ compartmentalization (AtDTX33, AtDTX35, AtALMT4, and GsCLC2) have been identified and characterized. These transporters have proven to be highly relevant for nutrition, long-distance transport and compartmentalization of Cl⁻, as well as for cell turgor regulation and stress tolerance in plants.

Chloride (Cl^–) has traditionally been considered harmful to agriculture because of its toxic effects in saline soils and its antagonistic interaction with nitrate (NO₃^–), which impairs NO₃^– nutrition. It has been largely believed that Cl^– antagonizes NO₃^– uptake and accumulation in higher plants, reducing crop yield. However, we have recently uncovered that Cl^– has new beneficial macronutrient, functions that improve plant growth, tissue water balance, plant water relations, photosynthetic performance, and water-use efficiency. The increased plant biomass indicates in turn that Cl^– may also improve nitrogen use efficiency (NUE). Considering that N availability is a bottleneck for the plant growth, the excessive NO₃^– fertilization frequently used in agriculture becomes a major environmental concern worldwide, causing excessive leaf NO₃^– accumulation in crops like vegetables and, consequently, a potential risk to human health. New farming practices aimed to enhance plant NUE by reducing NO₃^– fertilization should promote a healthier and more sustainable agriculture. Given the strong interaction between Cl^– and NO₃^– homeostasis in plants, we have verified if indeed Cl^– affects NO₃^– accumulation and NUE in plants. For the first time to our knowledge, we provide a direct demonstration which shows that Cl^–, contrary to impairing of NO₃^– nutrition, facilitates NO₃^– utilization and improves NUE in plants. This is largely due to Cl^– improvement of the N–NO₃^– utilization efficiency (NU_TE), having little or moderate effect on N–NO₃^– uptake efficiency (NU_PE) when NO₃^– is used as the sole N source. Clear positive correlations between leaf Cl^– content vs. NUE/NU_TE or plant growth have been established at both intra- and interspecies levels. Optimal NO₃^– vs. Cl^– ratios become a useful tool to increase crop yield and quality, agricultural sustainability and reducing the negative ecological impact of NO₃^– on the environment and on human health.

Chloride (Cl⁻) and nitrate (NO3−) are closely related anions involved in plant growth. Their similar physical and chemical properties make them to interact in cellular processes like electrical balance and osmoregulation. Since both anions share transport mechanisms, Cl⁻ has been considered to antagonize NO3− uptake and accumulation in plants. However, we have recently demonstrated that Cl⁻ provided at beneficial macronutrient levels improves nitrogen (N) use efficiency (NUE). Biochemical mechanisms by which beneficial Cl⁻ nutrition improves NUE in plants are poorly understood. First, we determined that Cl⁻ nutrition at beneficial macronutrient levels did not impair the NO3− uptake efficiency, maintaining similar NO3− content in the root and in the xylem sap. Second, leaf NO3− content was significantly reduced by the treatment of 6 mM Cl⁻ in parallel with an increase in NO3− utilization and NUE. To verify whether Cl⁻ nutrition reduces leaf NO3− accumulation by inducing its assimilation, we analysed the content of N forms and the activity of different enzymes and genes involved in N metabolism. Chloride supply increased transcript accumulation and activity of most enzymes involved in NO3− assimilation into amino acids, along with a greater accumulation of organic N (mostly proteins). A reduced glycine/serine ratio and a greater ammonium accumulation pointed to a higher activity of the photorespiration pathway in leaves of Cl⁻-treated plants. Chloride, in turn, promoted higher transcript levels of genes encoding enzymes of the photorespiration pathway. Accordingly, microscopy observations suggested strong interactions between different cellular organelles involved in photorespiration. Therefore, in this work we demonstrate for the first time that the greater NO3− utilization and NUE induced by beneficial Cl⁻ nutrition is mainly due to the stimulation of NO3− assimilation and photorespiration, possibly favouring the production of ammonia, reductants and intermediates that optimize C-N re-utilization and plant growth. This work demonstrates new Cl⁻ functions and remarks on its relevance as a potential tool to manipulate NUE in plants.

Chloride (Cl^-) is traditionally categorized as an antagonist of nitrate (NO₃^-) because Cl^- hinders plant NO₃^- transport and accumulation. However, we have recently defined Cl^- as a beneficial macronutrient for higher plants, due to specific functions that lead to more efficient use of water, nitrogen (N) and CO₂ under optimal N and water supply. When accumulated in leaves at macronutrient levels, Cl^- promotes growth through osmotic, physiological, metabolic, anatomical and cellular changes that improve plant performance under optimal NO₃^- nutrition. Nitrate over-fertilization in agriculture can adversely affect crop yield and nature, while its deficiency limits plant growth. To study the relationship between Cl^- nutrition and NO₃^- availability, we have characterized different physiological responses such as growth and yield, N-use efficiency, water status, photosynthesis, leaf anatomy, pigments and antioxidants in tomato plants treated with or without 5 mM Cl^- salts and increasing NO₃^- treatments (3-15 mM). First, we have demonstrated that 5 mM Cl^- application can reduce the use of NO₃^- in the nutrient solution by up to half without detriment to plant growth and yield in tomato and other horticultural plants. Second, Cl^- application reduced stress symptoms and improved plant growth under low-NO₃^- conditions. The Cl^--dependent resistance to low-N stress resulted from: more efficient use of the available NO₃^-; improved plant osmotic and water status regulation; improved stomatal conductance and photosynthetic rate; and better antioxidant response. We proposed that beneficial Cl^- levels increase the crop ability to grow better with lower NO₃^- requirements and withstand N deficiency, promoting a more sustainable and resilient agriculture.

SUMMARY: Despite being an essential micronutrient and its recent classification as a beneficial macronutrient, chloride (Cl⁻) has traditionally been considered of limited agricultural relevance and a potentially toxic saline ion. This study provides the first comprehensive demonstration of the quantitative and qualitative importance of Cl⁻ during early vegetative development (EVD) of tobacco and Arabidopsis thaliana plants. During this developmental stage, these and other species (including celery, lettuce, Swiss chard, spinach, squash, tomato, chili pepper, eggplant, and perennial ryegrass) exhibit the highest demand and transport rate of this non‐assimilable mineral nutrient to maximise growth of these herbaceous and also woody (such as citrus and olive) species. While Cl⁻ promotes cell expansion across all growth stages, its particularly pronounced stimulation of plant growth during EVD is associated with enhanced photosynthetic performance and PSII activity. This enhancement is in turn linked to a reduction in non‐regulated energy dissipation in PSII and an increase in the electron transport rate, along with ultrastructural changes in chloroplasts, underscoring that Cl⁻ is specifically required during EVD to drive the maturation of the photosynthetic apparatus. Unlike adult plants, the growth deficiencies caused by sub‐macronutrient Cl⁻ levels during EVD cannot be mitigated by equivalent nitrate (NO₃⁻) supplementation. As EVD concludes, plant demand for Cl⁻ gradually decreases, accompanied by a reduced growth response to Cl⁻ and an increased reliance on NO₃⁻, emphasising stage‐specific nutrient needs. The relevance of Cl⁻ as a morphogenic driver during a critical stage of development has significant implications for optimizing agronomic practices, particularly by reducing dependence on nitrogen fertilisers.

Sea level rise (SLR) imposes increasing salinity and inundation stresses in salt marshes which simultaneously face invasions by exotic plant species. We aimed to improve and apply knowledge on the ecophysiological responses of halophytes to SLR to conservation management of salt marshes. In a mesocosm experiment, we measured and compared phosphoenolpyruvate carboxylase (PEPC) activity and related functional traits of the California-native Spartina foliosa, invasive S. densiflora and their hybrid S. densiflora × foliosa in response to increasing levels of salinity and inundation. S. foliosa was moderately sensitive to salinity, showing a 57% reduction in PEPC specific activity from freshwater to hypersalinity. This native species compensated for the reduction of PEPC activity with increased salinity through 80% higher enzyme activation by phosphorylation. PEPC functional trait responses of S. foliosa were mostly independent of inundation depth. In view of these results, managers should conserve undeveloped lands for accommodation space above current high tide lines to facilitate colonization of stress-tolerant S. foliosa. Our results on functional responses of PEPC traits recorded high sensitivity to salinity for S. densiflora. This was reflected by 65% lower PEPC specific activity together with increasing accumulation of free proline (+96%) and total proteins (+23%) with elevated salinity. These results suggest prioritized eradication of S. densiflora populations in brackish habitats. Measured PEPC responses support the high stress tolerance of the S. densiflora × foliosa hybrid. PEPC traits for the hybrid were mostly independent of salinity and inundation. The hybrid showed higher PEPC-specific activity than S. foliosa (+70%) and S. densiflora (+15%) in freshwater under intermediate inundation. Results suggest that eradication of the hybrid should be the highest management priority. Our study shows that the responses of key functional physiological traits to environmental stresses serve as biological indicators that can guide ecosystem management practices in a scenario of climate change.