Nitrate Transport Research Articles

Multiomic Approaches Reveal Hormonal Modulation and Nitrogen Uptake and Assimilation in the Initial Growth of Maize Inoculated with Herbaspirillum seropedicae

Herbaspirillum seropedicae is an endophytic bacterium that can fix nitrogen and synthesize phytohormones, which can lead to a plant growth-promoting effect when used as a microbial inoculant. Studies focused on mechanisms of action are crucial for a better understanding of the bacteria-plant interaction and optimization of plant growth-promoting response. This work aims to understand the underlined mechanisms responsible for the early stimulatory growth effects of H. seropedicae inoculation in maize. To perform these studies, we combined transcriptomic and proteomic approaches with physiological analysis. The results obtained eight days after inoculation (d.a.i) showed increased root biomass (233 and 253%) and shoot biomass (249 and 264%), respectively, for the fresh and dry mass of maize-inoculated seedlings and increased green content and development. Omics data analysis, before a positive biostimulation phenotype (5 d.a.i.) revealed that inoculation increases N-uptake and N-assimilation machinery through differentially expressed nitrate transporters and amino acid pathways, as well carbon/nitrogen metabolism integration by the tricarboxylic acid cycle and the polyamine pathway. Additionally, phytohormone levels of root and shoot tissues increased in bacterium-inoculated-maize plants, leading to feedback regulation by the ubiquitin-proteasome system. The early biostimulatory effect of H. seropedicae partially results from hormonal modulation coupled with efficient nutrient uptake-assimilation and a boost in primary anabolic metabolism of carbon-nitrogen integrative pathways.

Rice miR168a-5p regulates seed length, nitrogen allocation and salt tolerance by targeting OsOFP3, OsNPF2.4 and OsAGO1a, respectively

Rice microRNA168a (osa-miR168a) plays important roles in mediating flowering time, grain yield and vigor, seeding growth, and immunity by targeting the RNA-induced silencing complex component Argonaute 1 (AGO1). However, the functions of miR168a exerted by targeting other genes require further clarification before it could be used in rice molecular breeding. In this study, we identified a new target gene of osa-miR168a-5p (miR168a-5p) in rice called OsOFP3 (ovate family protein 3) and investigated the roles of miR168a-5p in response to brassinosteroids (BRs), salt stress, and nitrogen allocation. Up- and downregulated miR168a-5p expression respectively decreased and increased the expression of the BR-negative regulator OsOPF3. The results of RNA ligase-mediated rapid amplification of cDNA ends (5′RLM-RACE) revealed cleavage sites in OsOPF3 and OsNPF2.4 mRNAs. The phenotype of miR168a-5p transgenic rice was BR-associated and included the lamina bending response to BR, short seeds, and low 1000-grain weight. MicroRNA 168a-5p also regulated the expression of the nitrate transporter, OsNPF2.4, which affected nitrogen allocation, and regulated OsAGO1a expression in response to salt stress. Taken together, rice miR168a-5p regulates BR-associated pathways, nitrogen transport, and stress by targeting OsOFP3, OsNPF2.4, and OsAGO1a, respectively, resulting in a series of important agronomic traits for rice breeding.

Light-Dependent Nitrate Removal Capacity of Green Microalgae

In the present study, Chlamydomonas sp. MACC-216 was used to investigate total nitrate removal in TAP medium with sodium nitrate as the sole nitrogen source under several light conditions made up of permuted combinations of three light colors (referred to as blue, red, and white light) and three light intensities (50 µmol m-2 s-1, 100 µmol m-2 s-1, and 250 µmol m-2 s-1). It was observed that nitrate removal efficiency is influenced by light color as well as light intensity. Additionally, Chlamydomonas sp. MACC-216 was cultivated in synthetic wastewater under four light conditions, namely, Blue 250, Blue 125 + Red 125, Red 250, and White 250, where it showed the highest nitrate removal efficiency and nitrate reductase activity under the Blue 125 + Red 125 light condition. To observe the impact of light color on the nitrate removal capacity of Chlamydomonas sp. MACC-216, the expression of five genes participating in nitrate transport and reduction (NRT1, NRT2.1, NRT2.2, NIA, and MCP) was also analyzed; these genes showed the highest expression under the Blue 125 + Red 125 light condition. Based on the above-mentioned findings, the blue + red light combination emerged as a promising light combination for nitrate removal. Hence, our study suggests the importance of the blue + red light combination together with high light intensity, as the optimal light condition for nitrate removal from synthetic wastewater in comparison to other monochromatic lights with high light intensity.

Hillslope Hydrology in a Deeply Weathered Saprolite and Associated Nitrate Transport to a Valley Bottom Wetland in Central Uganda

While interflow from the slopes can be crucial for water and nutrient availability in low-input farming systems in wetlands in East Africa, very little data exist on hillslope hydrology and associated nutrient transport in deeply weathered saprolites over crystalline rocks. This study aims for a better process understanding of interflow generation and routing in this environment and its contribution to water and nitrate availability at the wetland fringe of a valley bottom wetland in central Uganda. The study was set up as a plot study following a multi-method approach, including interflow trenches, soil analysis, and geo-electrical measurements. We found that interflow generation was related to the undulating subsurface topography and the conductivity of the upper saprolite, while interflow was conducted to the slope toe via small, perched aquifers and preferential flow paths within the saprolite, which are connected during the rainy season. Interflow volumes and nitrate transport were strongly related to the land-use type and rainfall characteristics. As the nitrate delivered from the slopes was quickly lost in the anaerobic environment of the wetland fringe, sustainable agricultural management should focus on the slope toe and the upland positions.

Hydrological regulation of nitrate sources, transformation and transport pathway in a karstic river

A comprehensive understanding of nitrate dynamics in rivers is vital for nitrogen management, especially for the karst zone with high leaching of nitrate producing potential risk to environmental quality. However, identification of the source-transformation-pathway of nitrate is limited in the karst rivers. Thus, a suit of hydrogeochemical parameters, including δ15N-NO3−, δ18O-NO3−, δ13C-DIC, δ18O-H2O, soil-related parameters (DOC) and geogenic ions (Mg2+, DIC), were used to identify nitrate sources and dynamics in the Wujiang River, a typical karstic river as a tributary of Changjiang River. The combined use of a conceptual model of normalized NO3− and DOC, and other hydrogeochemical parameters (δ13C-DIC, Mg2+, DIC) suggested that most nitrates were transported through the soil/epikarst pathway in the wet season, whereas >70 % of nitrates were transported through groundwater pathway in the dry season for Wujiang River. Nitrification and mixing were the dominant processes, meanwhile ∼31 % of nitrate was denitrified in the wet season in this study. The reanalysis of δ15N-NO3− and δ18O-NO3− values for reported karstic rivers in China and worldwide also revealed the predominant nitrification and denitrification in karst regions. The Bayesian model indicated that chemical fertilizer (CF, 42 %) and soil organic nitrogen (SON, 32 %) contributed the most in the wet season, while ∼60 % of nitrate originated from SON (31 %) and manure and sewage wastes (M&S, 30 %) in the dry season in the Wujiang River. The sustained contribution of non-point sources from nitrification through the soil/epikarst pathway might be related to the widespread porosities in the karst rivers. Thus, measures to reduce nitrogen leaching and improve fertilizer use efficiency should be considered, like water and soil conservation, slow-release fertilizer and nitrification inhibitors. Additionally, a suit of hydrogeochemical indicators proves to be helpful for identification of source, transformation and transport pathway of nitrate in the karst basin.

Response Characteristics of Growth Properties and Yield of Strong-gluten Winter Wheat Upon the Water-saving Supply Condition

Identification of the wheat cultivars with excellent strong-gluten property and the related stress resistance-associated physiological mechanism can provide practical basis for guiding the high-quality wheat cultivation under water-saving conditions. With an aim to understand the growth characteristics and yield traits of strong-gluten wheat in Hebei plain, this study investigated the photosynthetic characteristics, contents of osmotic regulatory substances (proline and soluble sugar), nitrogen uptake and accumulation, population characteristics and yield of strong-gluten wheat varieties (Gaoyou 5218, Shiyou 20 and Shishi Luan 02-1), three ones with demonstration potential, in response to reduction of water supply (irrigated prior to seed sowing and joint stage over the whole growth period). Results showed that compared with control (a common-gluten type cultivar Shimai 22), the photosynthetic parameters (chlorophyll content and Pn and gs), proline and soluble sugar contents, plant biomass, and the yield traits (grain number per spike, 1000 grain weight, and yield) were alleviated to different degrees. In contrast, the plant nitrogen content, population stem numbers, and spike amounts per planting unit area were shown to be superior to the control. According to the physiological parameters, population plant growth traits, yield and protein yield, the tested strong-gluten wheat cultivars were ranked in an order of Gaoyou 5218, Shiyou 20, and Shiluan 02-1, with the first one displaying the best on behaviors of photosynthetic parameters, osmotic regulation substance content and yield and to be similar to those shown in the control cultivar. Moreover, the grain protein yield of Gaoyou 5218 plants was higher than that of the control. Regression analysis showed that the expression level of <SUP>Δ</SUP>1-pyrrolin-5-carboxylate synthase (P5CS) gene <i>TaP5CS1</i> was highly positively correlated with the proline content of the tested varieties during growth stages, and the transcript abundance of nitrate transporter gene <i>TaNRT2.1</i> was significantly positively correlated with the nitrogen accumulative amount of plants during the same growth period. These results indicated that above genes played an important role in regulating the osmotic regulation ability and nitrogen uptake and accumulation of the strong-gluten and control cultivars under water-saving cultivation conditions. Gaoyou 5218 showed relatively elite yield and protein yield per unit area, indicating that this cultivar has an important application prospective for strong-gluten wheat cultivation.

Genome-wide identification and characterization of the NPF genes provide new insight into low nitrogen tolerance in Setaria.

Nitrogen (N) is essential for plant growth and yield production and can be taken up from soil in the form of nitrate or peptides. The NITRATE TRANSPORTER 1/PEPTIDE TRANSPORTER family (NPF) genes play important roles in the uptake and transportation of these two forms of N. Bioinformatic analysis was used to identify and characterize the NPF genes in Setaria. RNA-seq was employed to analyze time-series low nitrate stress response of the SiNPF genes. Yeast and Arabidopsis mutant complementation were used to test the nitrate transport ability of SiNRT1.1B1 and SiNRT1.1B2. We identified 92 and 88 putative NPF genes from foxtail millet (Setaria italica L.) and its wild ancestor green foxtail (Setaria viridis L.), respectively. These NPF genes were divided into eight groups according to their sequence characteristics and phylogenetic relationship, with similar intron-exon structure and motifs in the same subfamily. Twenty-six tandem duplication and 13 segmental duplication events promoted the expansion of SiNPF gene family. Interestingly, we found that the tandem duplication of the SiNRT1.1B gene might contribute to low nitrogen tolerance of foxtail millet. The gene expression atlas showed that the SiNPFs were divided into two major clusters, which were mainly expressed in root and the above ground tissues, respectively. Time series transcriptomic analysis further revealed the response of these SiNPF genes to short- and long- time low nitrate stress. To provide natural variation of gene information, we carried out a haplotype analysis of these SiNPFs and identified 2,924 SNPs and 400 InDels based on the re-sequence data of 398 foxtail millet accessions. We also predicted the three-dimensional structure of the 92 SiNPFs and found that the conserved proline 492 residues were not in the substrate binding pocket. The interactions of SiNPF proteins with were analyzed using molecular docking and the pockets were then identified. We found that the SiNPFs- binding energy ranged from -3.8 to -2.7 kcal/mol. Taken together, our study provides a comprehensive understanding of the NPF gene family in Setaria and will contribute to function dissection of these genes for crop breeding aimed at improving high nitrogen use efficiency.

A Haplotype-Based GWAS Identified Trait-Improving QTL Alleles Controlling Agronomic Traits under Contrasting Nitrogen Fertilization Treatments in the MAGIC Wheat Population WM-800.

The multi-parent-advanced-generation-intercross (MAGIC) population WM-800 was developed by intercrossing eight modern winter wheat cultivars to enhance the genetic diversity present in breeding populations. We cultivated WM-800 during two seasons in seven environments under two contrasting nitrogen fertilization treatments. WM-800 lines exhibited highly significant differences between treatments, as well as high heritabilities among the seven agronomic traits studied. The highest-yielding WM-line achieved an average yield increase of 4.40 dt/ha (5.2%) compared to the best founder cultivar Tobak. The subsequent genome-wide-association-study (GWAS), which was based on haplotypes, located QTL for seven agronomic traits including grain yield. In total, 40, 51, and 46 QTL were detected under low, high, and across nitrogen treatments, respectively. For example, the effect of QYLD_3A could be associated with the haplotype allele of cultivar Julius increasing yield by an average of 4.47 dt/ha (5.2%). A novel QTL on chromosome 2B exhibited pleiotropic effects, acting simultaneously on three-grain yield components (ears-per-square-meter, grains-per-ear, and thousand-grain-weight) and plant-height. These effects may be explained by a member of the nitrate-transporter-1 (NRT1)/peptide-family, TaNPF5.34, located 1.05 Mb apart. The WM-800 lines and favorable QTL haplotypes, associated with yield improvements, are currently implemented in wheat breeding programs to develop advanced nitrogen-use efficient wheat cultivars.

Imidacloprid Disturbs the Nitrogen Metabolism and Triggers an Overall Stress Response in Maize Seedlings

Imidacloprid (IMI) protects crops from pests; however, its potential toxicity to plants and underlying mechanisms are still poorly understood. We investigated the effects of IMI on maize seedlings under different nitrogen sufficiency conditions. Our measurement of the maize seedlings’ growth traits and physiological indicators found that a 5 ppm IMI treatment stunted the maize’s growth and enhanced membrane lipid peroxidation under a nitrogen-supplied condition, but that it promoted an increase in biomass and alleviated chlorosis in maize shoots under nitrogen deficiency. These results suggest that IMI causes serious toxicity in maize seedlings under nitrogen-sufficient conditions. The content of IMI indicated that the leaf was the main site of IMI accumulation in maize, and that NO3− was beneficial for the transportation of IMI from the roots to the leaves. The three groups of seedlings, which received 0 (−N), 4 (N) or 10 mmol L−1 NO3− (NN), were either treated or not treated with 5 ppm IMI. The six sets of transcriptome profiles from the shoots and roots were compared using Illumina sequencing. Transcriptome analysis revealed that IMI treatment led to changes in the expression of the genes involved in multiple biological processes, including nitrate transporter, nitrogen assimilation, nitrogen-regulatory factors, detoxification-related genes and several antioxidant-related genes in maize roots. The above results and the data for the nitrate content, glutamine synthetase activities and nitrate reductase activities showed that IMI disturbed nitrogen absorption and metabolism in maize seedlings. Glutathione S-transferase genes, C-type ATP-binding cassette (ABC) transporter 4, anthocyanins and lignin may play an important role in the detoxification of IMI in maize. These findings have helped us to elaborate the underlying detoxification mechanisms of IMI in plants, which is highly important in the cultivation of anti-pesticide crop varieties.

Disentangling scatter in long-term concentration–discharge relationships: the role of event types

Abstract. Relationships between nitrate concentrations and discharge rates (C–Q) at the catchment outlet can provide insights into sources, mobilization and biogeochemical transformations of nitrate within the catchment. Nitrate C–Q relationships often exhibit considerable scatter that might be related to variable hydrologic conditions during runoff events at sampling time, corresponding to variable sources and flow paths despite similar discharge (Q) rates. Although previous studies investigated the origins of this scatter in individual or in a few catchments, the role of different runoff event types across a large set of catchments is not yet fully understood. This study combines a hydrological runoff event classification framework with low-frequency nitrate samples in 184 catchments to explore the role of different runoff events in shaping long-term C–Q relationships and their variability across contrasting catchments. In most of the catchments, snow-impacted events produce positive deviations of concentrations, indicating an increased nitrate mobilization compared to the long-term pattern. In contrast, negative deviations occur mostly for rainfall-induced events with dry antecedent conditions, indicating the occurrence of lower nitrate concentrations (C) in river flows than their long-term pattern values during this type of event. Pronounced differences in event runoff coefficients among different event types indicate their contrasting levels of hydrologic connectivity that in turn might play a key role in controlling nitrate transport due to the activation of faster flow paths between sources and streams. Using long-term, low-frequency nitrate data, we demonstrate that runoff event types shape observed scatter in long-term C–Q relationships according to their level of hydrologic connectivity. In addition, we hypothesize that the level of biogeochemical attenuation of catchments can partially explain the spatial variability of the scatter during different event types.

Soil-to-fruit nitrogen flux mediates the onset of fruit-nitrogen remobilization and color change in citrus

In non-climacteric fruit tree species, color change is primarily controlled by the interplay between environmental conditions, nutritional factors (nitrogen and sugars) and hormones, mainly abscisic acid and gibberellins (GA), through a complex mechanism which is not completely understood. Nitrogen has a strong impact on color change, influenced by environmental changes, either locally or at the whole tree level. We use Citrus trees, as a non-climacteric model species, to understand the long-distance nitrogen signaling mechanism delaying the chloroplast-to-chromoplast transition at the molecular level. It is unknown whether nitrogen regulates the branch point of geranylgeranyl diphosphate (GGPP), the shared precursor for gibberellin, chlorophyll and carotenoid synthesis. We used 15 N to trace the root-to-fruit nitrogen flux in trees grown under soils differing in nitrogen content and temperature, and treated with NH 4 NO 3 and GA. Key genes involved in nitrogen signaling, transport and metabolism, and those from the GGPP branch point were analyzed in the fruit. Results explain how soil temperature modifies 15 N transport to play a key role in signaling citrus color change, and show that fruit-nitrogen remobilization is required for triggering degreening. Nitrogen content in the fruit modulates the onset of glutamate deamination, asparagine synthesis, nitrite assimilation and GA 1 depletion. Expression of the nitrate transporter NRT1.2, glutamate dehydrogenase and asparagine synthetase genes was high right before the start of degreening, together with NH 4 + concentration. Nitrogen delayed carotenoid synthesis (phytoene synthase gene expression) without modifying gibberellin synthesis ( ent -kaurene synthase and oxidase) at the chloroplast level. • N and GA 1 depletion from the fruit is a prerequisite for citrus color change • Cold-soil temperatures reduce 15 N translocation and trigger fruit color change • Low N flux correlates fruit protein hydrolysis, upregulating GDH, AS and NRT1.2 • GA treatment delays protein hydrolysis, NRT1.2 expression and N depletion • The first steps of synthesis of GA in the chloroplast are not modified by N

The small peptide CEP1 and the NIN-like protein NLP1 regulate NRT2.1 to mediate root nodule formation across nitrate concentrations.

Legumes acquire fixed nitrogen (N) from the soil and through endosymbiotic association with diazotrophic bacteria. However, establishing and maintaining N2-fixing nodules are expensive for the host plant, relative to taking up N from the soil. Therefore, plants suppress symbiosis when N is plentiful and enhance symbiosis when N is sparse. Here, we show that the nitrate transporter MtNRT2.1 is required for optimal nodule establishment in Medicago truncatula under low-nitrate conditions and the repression of nodulation under high-nitrate conditions. The NIN-like protein (NLP) MtNLP1 is required for MtNRT2.1 expression and regulation of nitrate uptake/transport under low- and high-nitrate conditions. Under low nitrate, the gene encoding the C-terminally encoded peptide (CEP) MtCEP1 was more highly expressed, and the exogenous application of MtCEP1 systemically promoted MtNRT2.1 expression in a compact root architecture 2 (MtCRA2)-dependent manner. The enhancement of nodulation by MtCEP1 and nitrate uptake were both impaired in the Mtnrt2.1 mutant under low nitrate. Our study demonstrates that nitrate uptake by MtNRT2.1 differentially affects nodulation at low- and high-nitrate conditions through the actions of MtCEP1 and MtNLP1.

Take it to the bank: A numerical examination of the effects of soil pipes on bypass of riparian buffer nitrate removal capacity

Streams are vital landscape ecosystems, and urbanization and modern agriculture have introduced pollutants (including excess nitrate) which threaten these waterways. Riparian zones have been shown to attenuate pollutants, and riparian buffers are commonly implemented to capture these benefits. Nevertheless, preferential flowpaths such as soil pipes have potential to bypass such attenuation. We used a MODFLOW groundwater model with the conduit flow package (CFP), along with the groundwater transport code MT3D-USGS, to simulate flow of water and nitrate through riparian groundwater to a gaining stream. We conducted a numerical sensitivity analysis to examine the impact of soil pipe presence and characteristics, as wells as soil matrix characteristics, on nitrate transport and uptake by denitrification within the saturated zone. We found that in systems with 2.0-m-long soil pipes, low head gradient (∇h less than 0.0035), low hydraulic conductivity (K < 10−4.75 m/s), and high reaction-constant (k > 4 day−1), soil pipes increased riparian bypass of nitrate by several orders of magnitude. Yet soil pipes increased the volumetric flowrate of water across the riparian zone only by up to 3.9 times. This comparatively greater effect on transport occurred because the accelerated advection of nitrate through the soil pipes decreased the residence time available for denitrification. We created two non-dimensional parameters that show when bypass will be important, the riparian bypass potential (ψ) that accounts for key governing factors such as volumetric flowrate and Damköhler number, and riparian predictive index (ζ), which is calculated from easily measurable parameters. Nitrate bypass starts to increase by orders of magnitude above ψ ≈ 2.75 and ζ ≈ 3.4. Our results emphasize the importance of accounting for soil pipes when constructing riparian buffers and predicting their effects on dissolved pollutant transport.

Overexpression of Nitrate Transporter 1/Peptide Gene OsNPF7.6 Increases Rice Yield and Nitrogen Use Efficiency.

Overuse of nitrogen fertilizer in fields has raised production costs, and caused environmental problems. Improving nitrogen use efficiency (NUE) of rice is essential for sustainable agriculture. Here we report the cloning, characterization and roles for rice of OsNPF7.6, a member of the nitrate transporter 1/peptide transporter family (NPF). The OsNPF7.6 protein is located in the plasma membrane, expressed in each tissue at all stages and is significantly regulated by nitrate in rice. Our study shows that the overexpression of OsNPF7.6 can increase the nitrate uptake rate of rice. Additionally, field experiments showed that OsNPF7.6 overexpression increased the total tiller number per plant and the grain weight per panicle, thereby improving grain yield and agronomic NUE in rice. Thus, OsNPF7.6 can be applied to be a novel target gene for breeding rice varieties with high NUE, and provide a reference for breeding higher yielding rice.

Land use as a major factor of riverine nitrate in a semi-arid farming-pastoral ecotone: New insights from multiple environmental tracers and molecular signatures of DOM

The nitrogen contamination in rivers has become significant concern in arid and semiarid areas due to water resource shortage and extensive anthropogenic activities in relation to land-use changes in China. As a major nitrogen species, identifying driving factors, transformation and sources of nitrate is crucial for managing nitrogen pollution in rivers. In this study, nitrate sources and transformations were deciphered using physicochemical variables, molecular signature of dissolved organic matter and coupled isotopes of nitrate under different land use types in the Yang River, a typical farming-pastoral ecotone in the semi-arid area of North China. The results of river water showed a significant positive correlation between NO3− concentrations, δ15N-NO3− values and percentage of urban land and cropland, which confirmed the critical role of land use in the variations of riverine nitrate. The correlation between dissolved organic matter composition (aliphatic and lignin-like compounds) and NO3−/Cl− ratios as well as Cl− concentrations verified the effect of agricultural activities on nitrate source and transport. The variation in water chemical variables and dual isotopes of nitrate in river and soil extracts (δ15N-NO3− and δ18O-NO3−) was indicative of the concurrence of in-soil nitrification process and assimilation, whereas denitrification was inhibited under aerobic conditions in the semiarid area. The Bayesian model revealed that about 60% of nitrate was derived from non-point sources (manure, soil organic nitrogen and chemical fertilizer) and 36% from sewage. Although urban is not the major land-use type in the farming-pastoral ecotone, sewage contributed to about 36% of nitrate. The source identification of nitrate stresses the importance of the management of non-point pollution and demand for sewage treatment facilities in the farming-pastoral ecotone. This multiple-tracer approach will help gain deeper insights into nitrogen management in semi-arid areas with extensive human disturbance.

High-temperature adaptation of an OsNRT2.3 allele is thermoregulated by small RNAs.

Climate change negatively affects crop yield, which hinders efforts to reach agricultural sustainability and food security. Here, we show that a previously unidentified allele of the nitrate transporter gene OsNRT2.3 is required to maintain high yield and high nitrogen use efficiency under high temperatures. We demonstrate that this tolerance to high temperatures in rice accessions harboring the HTNE-2 (high temperature resistant and nitrogen efficient-2) alleles from enhanced translation of the OsNRT2.3b mRNA isoform and the decreased abundance of a unique small RNA (sNRT2.3-1) derived from the 5' untranslated region of OsNRT2.3. sNRT2.3-1 binds to the OsNRT2.3a mRNA in a temperature-dependent manner. Our findings reveal that allelic variation in the 5' untranslated region of OsNRT2.3 leads to an increase in OsNRT2.3b protein levels and higher yield during high-temperature stress. Our results also provide a breeding strategy to produce rice varieties with higher grain yield and lower N fertilizer input suitable for a sustainable agriculture that is resilient against climate change.

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

Nitrate is a major nutrient and osmoticum for plants. To deal with fluctuating nitrate availability in soils, plants store this nutrient in their vacuoles. Chloride channel a (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. We introduced the CLCaE203A point mutation to investigate its physiological importance 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.

Understanding plant-microbe interaction of rice and soybean with two contrasting diazotrophic bacteria through comparative transcriptome analysis.

Understanding the beneficial plant-microbe interactions is becoming extremely critical for deploying microbes imparting plant fitness and achieving sustainability in agriculture. Diazotrophic bacteria have the unique ability to survive without external sources of nitrogen and simultaneously promote host plant growth, but the mechanisms of endophytic interaction in cereals and legumes have not been studied extensively. We have studied the early interaction of two diazotrophic bacteria, Gluconacetobacter diazotrophicus (GAB) and Bradyrhizobium japonicum (BRH), in 15-day-old seedlings of rice and soybean up to 120 h after inoculation (hai) under low-nitrogen medium. Root colonization of GAB in rice was higher than that of BRH, and BRH colonization was higher in soybean roots as observed from the scanning electron microscopy at 120 hai. Peroxidase enzyme was significantly higher at 24 hai but thereafter was reduced sharply in soybean and gradually in rice. The roots of rice and soybean inoculated with GAB and BRH harvested from five time points were pooled, and transcriptome analysis was executed along with control. Two pathways, "Plant pathogen interaction" and "MAPK signaling," were specific to Rice-Gluconacetobacter (RG), whereas the pathways related to nitrogen metabolism and plant hormone signaling were specific to Rice-Bradyrhizobium (RB) in rice. Comparative transcriptome analysis of the root tissues revealed that several plant-diazotroph-specific differentially expressed genes (DEGs) and metabolic pathways of plant-diazotroph-specific transcripts, viz., chitinase, brassinosteroid, auxin, Myeloblastosis (MYB), nodulin, and nitrate transporter (NRT), were common in all plant-diazotroph combinations; three transcripts, viz., nitrate transport accessory protein (NAR), thaumatin, and thionin, were exclusive in rice and another three transcripts, viz., NAC (NAM: no apical meristem, ATAF: Arabidopsis thaliana activating factor, and CUC: cup-shaped cotyledon), ABA (abscisic acid), and ammonium transporter, were exclusive in soybean. Differential expression of these transcripts and reduction in pathogenesis-related (PR) protein expression show the early interaction. Based on the interaction, it can be inferred that the compatibility of rice and soybean is more with GAB and BRH, respectively. We propose that rice is unable to identify the diazotroph as a beneficial microorganism or a pathogen from an early response. So, it expressed the hypersensitivity-related transcripts along with PR proteins. The molecular mechanism of diazotrophic associations of GAB and BRH with rice vis-à-vis soybean will shed light on the basic understanding of host responses to beneficial microorganisms.

Identification and Expression Analysis of the NPF Genes in Cotton.

The NPF (NITRATE TRANSPORTER 1/PEPTIDE TRANSPORTER FAMILY) transports various substrates, including nitrogen (N), which is essential for plant growth and development. Although many NPF homologs have been identified in various plants, limited studies on these proteins have been reported in cotton. This study identified 75, 71, and 150 NPF genes in Gossypium arboreum, G. raimondii, and G. hirsutum, respectively, via genome-wide analyses. The phylogenetic tree indicated that cotton NPF genes are subdivided into eight subgroups, closely clustered with Arabidopsis orthologues. The chromosomal location, gene structure, motif compositions, and cis-elements have been displayed. Moreover, the collinearity analysis showed that whole-genome duplication event has played an important role in the expansion and diversification of the NPF gene family in cotton. According to the transcriptome and qRT-PCR analyses, several GhNPFs were induced by the nitrogen deficiency treatment. Additional functional experiments revealed that virus-induced silencing (VIGS) of the GhNPF6.14 gene affects the growth and N absorption and accumulation in cotton. Thus, this study lays the foundation for further functional characterization of NPF genes in cotton.

Rice glutamate receptor-like channel OsGLR3.4 modulates the root tropism growth towards amino acids via plasma membrane depolarization and ROS generation

Understanding how plants determine root directional growth towards nutrient sources is important to reveal optimal strategies during plant growth. GLUTAMATE RECEPTOR-LIKE (GLR) channels play essential roles in various Ca2+-mediated plant-specific physiological processes, but how GLRs are involved in the root tropism growth towards surrounding nutrients remains largely unknown. Here, we report that OsGLR3.4, a rice Ca2+-permeable channel, specifically modulates the root growth and tropism towards Glycine (Gly) via Ca2+-mediated electrical activity and reactive oxygen species (ROS) production. Mutation of OsGLR3.4 significantly reduced the inhibitory effect of Gly on the primary root growth, coupled with the decrease of Gly-induced membrane depolarization and ROS levels at the elongation zone in the root tip. More importantly, our results showed that OsGLR3.4 was required for root nutritropism growth towards Gly. Mechanistic study showed that, when grown in the agar with unequal Gly application, the bilaterally asymmetrical distribution of membrane depolarization and ROS evoked by OsGLR3.4-mediated [Ca2+]cyt fluctuation across the root resulted in the differential cell elongation at two sides of the root tip, which therefore led to the root nutritropism growth. Nevertheless, addition of Ca2+ channel blocker eliminated the Gly-induced asymmetrical distribution of depolarization and ROS across the root tip. Notably, OsGLR3.4 promoted the nitrate uptake in the root by enhancing expression of nitrate transporter genes in response to Gly treatment, suggesting its potential roles in crop yield. Our findings established a mechanism by which OsGLR3.4 modulates rice root growth and tropism towards nutrient sources, and shed light on the root behavioral property used by plants to acquire nutrients efficiently.