High-affinity Transport System Research Articles

Physiological, molecular, and environmental insights into plant nitrogen uptake, and metabolism under abiotic stresses.

Nitrogen (N) as an inorganic macronutrient is inevitable for plant growth, development, and biomass production. Many external factors and stresses, such as acidity, alkalinity, salinity, temperature, oxygen, and rainfall, affect N uptake and metabolism in plants. The uptake of ammonium (NH4 +) and nitrate (NO3 -) in plants mainly depends on soil properties. Under the sufficient availability of NO3 - (>1mM), low-affinity transport system is activated by gene network NRT1, and under low NO3 - availability (<1mM), high-affinity transport system starts functioning encoded by NRT2 family of genes. Further, under limited N supply due to edaphic and climatic factors, higher expression of the AtNRT2.4 and AtNRT2.5T genes of the NRT2 family occur and are considered as N remobilizing genes. The NH4 + ion is the final form of N assimilated by cells mediated through the key enzymes glutamine synthetase and glutamate synthase. The WRKY1 is a major transcription factor of the N regulation network in plants. However, the transcriptome and metabolite profiles show variations in N assimilation metabolites, including glycine, glutamine, and aspartate, under abiotic stresses. The overexpression of NO3 - transporters (OsNRT2.3a and OsNRT1.1b) can significantly improve the biomass and yield of various crops. Altering the expression levels of genes could be a valuable tool to improve N metabolism under the challenging conditions of soil and environment, such as unfavorable temperature, drought, salinity, heavy metals, and nutrient stress.

Glutamine as sole nitrogen source prevents induction of nitrate transporter gene NRT2.4 and affects amino acid metabolism in Arabidopsis.

Plants assimilate inorganic nitrogen (N) to glutamine. Glutamine is the most abundant amino acid in most plant species, the N-supplying precursor of all N-containing compounds in the cell and the first organic nitrogen molecule formed from inorganic nitrogen taken up by the roots. In addition to its role in plant nutrition, glutamine most likely also has a function as a signaling molecule in the regulation of nitrogen metabolism. We investigated whether glutamine influences the high-affinity transporter system for nitrate uptake. Therefore, we analyzed the expression of the nitrate transporter NRT2.4, which is inducible by N deficiency, in Arabidopsis thaliana grown under different nitrogen starvation scenarios, comparing nitrate or glutamine as the sole nitrogen source. Using the reporter line ProNRT2.4:GFP and two independent knockout lines, nrt2.4-1 and nrt2.4-2, we analyzed gene expression and amino acid profiles. We showed that the regulation of NRT2.4 expression depends on available nitrogen in general, for example on glutamine as a nitrogen source, and not specifically on nitrate. In contrast to high nitrate concentrations, amino acid profiles changed to an accumulation of amino acids containing more than one nitrogen during growth in high glutamine concentrations, indicating a switch to nitrogen storage metabolism. Furthermore, we demonstrated that the nrt2.4-2 line shows unexpected effects on NRT2.5 gene expression and the amino acids profile in shoots under high glutamine supply conditions compared to Arabidopsis wild type and nrt2.4-1, suggesting non-NRT2.4-related metabolic consequences in this knockout line.

Integrated genomics provides insights into the evolution of the polyphosphate accumulation trait of Ca. Accumulibacter

Candidatus Accumulibacter, a prominent polyphosphate-accumulating organism (PAO) in wastewater treatment, plays a crucial role in enhanced biological phosphorus removal (EBPR). The genetic underpinnings of its polyphosphate accumulation capabilities, however, remain largely unknown. Here, we conducted a comprehensive genomic analysis of Ca. Accumulibacter-PAOs and their relatives within the Rhodocyclaceae family, identifying 124 core genes acquired via horizontal gene transfer (HGT) at its least common ancestor. Metatranscriptomic analysis of an enrichment culture of Ca. Accumulibacter revealed active transcription of 44 of these genes during an EBPR cycle, notably including the polyphosphate kinase 2 (PPK2) gene instead of the commonly recognized polyphosphate kinase 1 (PPK1) gene. Intriguingly, the phosphate regulon (Pho) genes showed minimal transcriptions, pointing to a distinctive fact of Pho dysregulation, where PhoU, the phosphate signaling complex protein, was not regulating the high-affinity phosphate transport (Pst) system, resulting in continuous phosphate uptake. To prevent phosphate toxicity, Ca. Accumulibacter utilized the laterally acquired PPK2 to condense phosphate into polyphosphate, resulting in the polyphosphate-accumulating feature. This study provides novel insights into the evolutionary emergence of the polyphosphate-accumulating trait in Ca. Accumulibacter, offering potential advancements in understanding the PAO phenotype in the EBPR process.

Identification, expression and DNA variation analysis of high affinity nitrate transporter NRT2/3 gene family in Sorghum bicolor

Nitrate is the main form of inorganic nitrogen that crop absorbs, and nitrate transporter 2 (NRT2) is a high affinity transporter using nitrate as a specific substrate. When the available nitrate is limited, the high affinity transport systems are activated and play an important role in the process of nitrate absorption and transport. Most NRT2 cannot transport nitrates alone and require the assistance of a helper protein belonging to nitrate assimilation related family (NAR2) to complete the absorption or transport of nitrates. Crop nitrogen utilization efficiency is affected by environmental conditions, and there are differences between varieties, so it is of great significance to develop varieties with high nitrogen utilization efficiency. Sorghum bicolor has high stress tolerance and is more efficient in soil nitrogen uptake and utilization. The S. bicolor genome database was scanned to systematically analyze the gene structure, chromosomal localization, physicochemical properties, secondary structure and transmembrane domain, signal peptide and subcellular localization, promoter region cis-acting elements, phylogenetic evolution, single nucleotide polymorphism (SNP) recognition and annotation, and selection pressure of the gene family members. Through bioinformatics analysis, 5 NRT2 gene members (designated as SbNRT2-1a, SbNRT2-1b, SbNRT2-2, SbNRT2-3, and SbNRT2-4) and 2 NAR2 gene members (designated as SbNRT3-1 and SbNRT3-2) were identified, the number of which was less than that of foxtail millet. SbNRT2/3 were distributed on 3 chromosomes, and could be divided into four subfamilies. The genetic structure of the same subfamilies was highly similar. The average value of SbNRT2/3 hydrophilicity was positive, indicating that they were all hydrophobic proteins, whereas α-helix and random coil accounted for more than 70% of the total secondary structure. Subcellular localization occurred on plasma membrane, where SbNRT2 proteins did not contain signal peptides, but SbNRT3 proteins contained signal peptides. Further analysis revealed that the number of transmembrane domains of the SbNRT2s family members was greater than 10, while that of the SbNRT3s were 2. There was a close collinearity between NRT2/3s of S. bicolor and Zea mays. Protein domains analysis showed the presence of MFS_1 and NAR2 protein domains, which supported executing high affinity nitrate transport. Phylogenetic tree analysis showed that SbNRT2/3 were more closely related to those of Z. mays and Setaria italic. Analysis of gene promoter cis-acting elements indicated that the promoter region of SbNRT2/3 had several plant hormones and stress response elements, which might respond to growth and environmental cues. Gene expression heat map showed that SbNRT2-3 and SbNRT3-1 were induced by nitrate in the root and stem, respectively, and SbNRT2-4 and SbNRT2-3 were induced by low nitrogen in the root and stem. Non-synonymous SNP variants were found in SbNRT2-4 and SbNRT2-1a. Selection pressure analysis showed that the SbNRT2/3 were subject to purification and selection during evolution. The expression of SbNRT2/3 gene and the effect of aphid infection were consistent with the expression analysis results of genes in different tissues, and SbNRT2-1b and SbNRT3-1 were significantly expressed in the roots of aphid lines 5-27sug, and the expression levels of SbNRT2-3, SbNRT2-4 and SbNRT3-2 were significantly reduced in sorghum aphid infested leaves. Overall, genome-wide identification, expression and DNA variation analysis of NRT2/3 gene family of Sorghum bicolor provided a basis for elucidating the high efficiency of sorghum in nitrogen utilization.

Influence of nitrate starvation on transcript levels of nitrate transporter (Nrt2) and β-subunit of phycoerythrin (Cpeb) genes in Rhodomonas sp.

Rhodomonas species are promising aquaculture feeds that provide high nutritional value. However, the degradation of intracellular phycoerythrin (PE) under nitrogen starvation negatively affects their nutritional value and growth. The present study aimed to identify the Nrt2 and Cpeb genes, which encode the high-affinity nitrate transporter system and the β-subunit of PE in Rhodomonas sp., respectively, and to investigate the effect of nitrate starvation on their expression. First, we screened RhNrt2 and RhCpeb by analysis of sequencing data from the transcriptome library of Rhodomonas sp. Blast analysis revealed that the amino acid sequences of RhNRT2 and RhCPEB with other organisms shared 29.5–53.3 % and 84.3–100 % identity, respectively. Additionally, their amino acid sequences exhibited distinctive features in their primary structure. Secondly, a batch culture experiment was designed under nitrate-replete and -starved conditions. Consistent with previous findings, nitrate-starved conditions had a detrimental effect on cell growth and intracellular PE content. The maximum cell density was 3.4 × 106 cells mL−1 under nitrate-starved condition, whereas a higher maximum cell density of 4.4 × 106 cells mL−1 was achieved under the nitrate-replete condition. Nitrate-starved conditions resulted in an over 80 % reduction in intracellular PE content. Relative RhNrt2 expression was consistently higher under nitrate-starved conditions compared to under nitrate-replete conditions, increasing 60.9-fold from the beginning to the end of the test. Conversely, relative RhCpeb expression was significantly suppressed and barely detectable under nitrate-starved conditions. Notably, the intracellular PE content decreases following RhCpeb suppression. The regulated expression of RhNrt2 and RhCpeb may allow these genes to be used as molecular indicators of nitrate status and intracellular PE biosynthesis. This study represents the first identification of RhNrt2 and RhCpeb genes and characterizes their expression in Rhodomonas species.

Zinc Supply Affects Cadmium Uptake and Translocation in the Hyperaccumulator Sedum Plumbizincicola as Evidenced by Isotope Fractionation.

This study employs stable isotope analysis to investigate the mechanisms of cadmium (Cd) and zinc (Zn) interaction in the metal hyperaccumulating plant species Sedum plumbizincicola. To this end, the Cd and Zn isotope compositions of root, stem, leaf, and xylem sap samples were determined during metal uptake and translocation at different Cd and Zn concentrations. The enrichment of light isotopes of both elements in plants during uptake was less pronounced at low metal supply levels, likely reflecting the switch from a low-affinity to a high-affinity transport system at lower levels of external metal supply. The lower δ114/110Cd values of xylem sap when treated with a metabolic inhibitor decreasing the active Cd uptake further supports the preference of heavier Cd isotopes during high-affinity transport. The Δ66Znplant-initialsolution or Δ66Znplant-finalsolution values were similar at different Cd concentrations, indicating negligible interaction of Cd in the Zn uptake process. However, decreasing Zn supply levels significantly increased the enrichment of light Cd isotopes in plants (Δ114/110Cd = -0.08‰) in low-Cd treatments but reduced the enrichment of light Cd isotopes in plants (Δ114/110Cd = 0.08‰) under high Cd conditions. A systematic enrichment of heavy Cd and light Zn isotopes was found in root-to-shoot translocation of the metals. The Cd concentrations of the growth solutions thereby had no significant impact on Zn isotope fractionation during root-to-shoot translocation. However, the Δ114/110Cdtranslocation values hint at possible competition between Cd and Zn for transporters during root-to-shoot transfer and this may impact the transport pathway of Cd. The stable isotope data demonstrate that the interactions between the two metals influenced the uptake and transport mechanisms of Cd in S. plumbizincicola but had little effect on those of Zn.

Rapid microbial uptake and mineralization of 14C-labelled cysteine and methionine along a grassland productivity gradient

Cysteine (Cys) and methionine (Met) are central to terrestrial S cycling because they are sources of carbon (C), nitrogen (N), and sulphur (S) for plant nutrition and microbial growth. However, soil microorganisms are expected to compete for the C, N and S in these S-amino acids. We hypothesized that microbial competition would be greater in soils with low plant productivity due to lower C inputs from plants. Here we added 14C-labelled Cys and Met to 5 soils collected from an altitude-driven primary grassland productivity gradient, we then measured microbial uptake with a centrifugal drainage procedure over 60 min, and the subsequent mineralization with NaOH traps over 48 h. Our results revealed that both Cys and Met were rapidly assimilated by soil microbes, with half-lives ranging from 0.34 to 2.14 min, which is an order of magnitude (or more) faster than when determined from measurement of 14CO2 evolution. This considerable delay between microbial 14C removal from soil solution and subsequent 14CO2 evolution indicates that the degradation of Cys and Met in grassland soils occurred mainly through biological processes. Soil microbial uptake of Cys and Met was dominated by a high-affinity transport system (0.01–0.1 mM), while a lower affinity transport system became more important at higher substrate concentrations (1–100 mM). In addition, microbial uptake and mineralization rates of Cys and Met declined in less productive, higher elevation sites, suggesting that the turnover of organic N and S, and subsequent availability for plant uptake is likely to be controlled by soil fertility. We conclude that although Cys and Met may represent a minor component of DON and DOS pools in soil, their importance for soil microbes and plant nutrition may have been underestimated due to their fast turnover and replenishment rates in grassland soils.

Copper toxicity compromises root acquisition of nitrate in the high affinity range.

The application of copper (Cu)-based fungicides for crop protection plans has led to a high accumulation of Cu in soils, especially in vineyards. Copper is indeed an essential micronutrient for plants, but relatively high concentrations in soil or other growth substrates may cause toxicity phenomena, such as alteration of the plant's growth and disturbance in the acquisition of mineral nutrients. This last aspect might be particularly relevant in the case of nitrate , whose acquisition in plants is finely regulated through the transcriptional regulation of transporters and plasma membrane H+-ATPase in response to the available concentration of the nutrient. In this study, cucumber plants were grown hydroponically and exposed to increasing concentrations of Cu (i.e., 0.2, 5, 20, 30, and 50 µM) to investigate their ability to respond to and acquire . To this end, the kinetics of substrate uptake and the transcriptional modulation of the molecular entities involved in the process have been assessed. Results showed that the inducibility of the high-affinity transport system was significantly affected by increasing Cu concentrations; at Cu levels higher than 20 µM, plants demonstrated either strongly reduced or abolished uptake activity. Nevertheless, the transcriptional modulation of both the nitrate transporter CsNRT2.1 and the accessory protein CsNRT3.1 was not coherent with the hindered uptake activity. On the contrary, CsHA2 was downregulated, thus suggesting that a possible impairment in the generation of the proton gradient across the root PM could be the cause of the abolishment of uptake.

Growth stage and nitrate limiting response of NRT2 and NAR2 gene families of bread wheat, and complementation and retrieval of nitrate uptake of atnrt2.1 mutant by a wheat NRT2 gene

The Nitrate Transporter 2 (NRT2), together with accessory protein (NAR), form high-affinity nitrate transport systems (HATS) responsible for nitrate uptake and translocation under its limited availability in plants. In the present study, we aimed to identify members of these two families’ genes in the bread wheat genome on a genome-wide scale, the evolutionary relationship of these families to major cereals, and their expression dynamics. A total of 46 and 8 NRT2 and NAR2 family genes, respectively, have been identified, which showed tissue and growth-stage specific nitrate limiting responses in all major tissues with a tendency of homeolog expression bias in two wheat genotypes. The high-affinity transport capacity of four wheat genotypes using 15N influx at three major physiological stages also showed significant roles of HATS at tissue and growth stage levels. The complementation of atnrt2.1 mutant with one of the high-affinity nitrate transporter genes, i.e., TaNRT2.1-B6, caused recovery of about 1.5 times 15N influx in N + and N- conditions in Arabidopsis mutant than wild type. The present study contributes to the genomic and functional understanding of NRT2 and NAR2 genes of bread wheat, providing a platform for further learning the underlying mechanism of nitrate uptake in bread wheat.

Identification of potassium transport proteins in algae and determination of their role under salt and saline-alkaline stress

K+ is an essential macronutrient in life activities of organisms. Compared with the natural environment (high Na+ and low K+), living cells contain high concentration of K+ and low concentration of Na+, which is one of the three major relics of the original birth of life. As the beginning of green plants evolution, algae are the most suitable species for understanding the efficient absorption of K+ through systematical analysis the potassium transport systems. In this study, we found that four algal species namely Chlorella variabilis (C. variabilis), Chlamydomonas reinhardtii (C. reinhardtii), Chlorella sacchrarophila (C. sacchrarophila), and Nannochloris sp. JB17 (JB17) displayed adaption to low K+ condition. Further, K+ concentrations in the growth media affected the growth of algae under the salt and saline-alkali stresses. Moreover, JB17, previously isolated from extremely saline-alkali soil, was found to maintain the high K+/Na+ ratio under the condition with high Na+ and low K+. According to bioinformatics data, there were great differences in the number, conserved motif and secondary structure of K+ transport proteins among algae, animals and plants, including the high affinity potassium transporter (HAK), Na+/K+-ATPase (NaK) and H+-ATPase (AHA). The expression of HAKs and NaK1 was significantly induced by low K+ condition, but most of them were decreased under salt or saline-alkali stress in C. reinhardtii, while the expression of those were significantly induced by saline-alkali stress in JB17. Ouabain, a specific inhibitor of Na+/K+-ATPase, impaired the saline-alkali tolerance of JB17 and C. variabilis by affecting the Na+ and K+ balance. Overexpressing JbAHA from JB17 improved the tolerance to saline-alkali stress in C. reinhardtii. All above indicating that Na+/K+-ATPase, H+-ATPase and K+/H+ high affinity transport systems in algae played important role in tolerance to high Na+ and low K+ environment.

LjNRT2.3 plays a hierarchical role in the control of high affinity transport system for root nitrate acquisition in Lotus japonicus.

Nitrate is a key mineral nutrient required for plant growth and development. Plants have evolved sophisticated mechanisms to respond to changes of nutritional availability in the surrounding environment and the optimization of root nitrate acquisition under nitrogen starvation is crucial to cope with unfavoured condition of growth. In this study we present a general description of the regulatory transcriptional and spatial profile of expression of the Lotus japonicus nitrate transporter NRT2 family. Furthermore, we report a phenotypic characterization of two independent Ljnrt2.3 knock out mutants indicating the involvement of the LjNRT2.3 gene in the root nitrate acquisition and lateral root elongation pathways occurring in response to N starvation conditions. We also report an epistatic relationship between LjNRT2.3 and LjNRT2.1 suggesting a combined mode of action of these two genes in order to optimize the Lotus response to a prolonged N starvation.

Effects of the Fractionation of the Nitrogen Fertilization on Root Nitrate Uptake and Vine Agronomic Performance in Pinot Gris Grapevines in a Temperate Climate

The study aims to investigate at a physiological and molecular level the root absorption of nitrogen (N) during the annual cycle of grapevine. The study was performed on potted Pinot Gris plants grafted on Kober 5BB and grown for two consecutive years in a semi-controlled environment (Northeast, Italy). The study compared the response of plants N-fertilized in spring (T1), or in spring and in post-harvest (T2) or no-N fertilized (C). Results showed that under our climate conditions nitrate was taken up by the grapevines when applied both in spring or in spring and post-harvest. The nitrate acquisition in T1 roots is mediated at molecular and physiological level by a higher activation of high-affinity nitrate-transport system to take up nitrate in comparison to no-fertilized plants. Comparing the two N fertilization managements, the dynamic of nitrate uptake rates showed different patterns with an overall late response of High-Affinity Transport System when the application of part of N-fertilization was delayed to post-harvest (T2) in comparison to T1. Nevertheless, during the 2 years of the trial the fractionation of N fertilization applied in spring and post-harvest did not negatively affect the yield and quality parameters of vines. An increase of N concentration in T2 roots after two consecutive growing seasons may suggest that the fractionation can lead to beneficial effects on long period. Results of this work contribute to improve the comprehension of N acquisition in grapevine in order to optimize the use of N inputs in vineyard.

Root nitrate uptake in sugarcane (Saccharum spp.) is modulated by transcriptional and presumably posttranscriptional regulation of the NRT2.1/NRT3.1 transport system.

Nitrate uptake in sugarcane roots is regulated at the transcriptional and posttranscriptional levels based on the physiological status of the plant and is likely a determinant mechanism for discrimination against nitrate. Sugarcane (Saccharum spp.) is one of the most suitable energy crops for biofuel feedstock, but the reduced recovery of nitrogen (N) fertilizer by sugarcane roots increases the crop carbon footprint. The low nitrogen use efficiency (NUE) of sugarcane has been associated with the significantly low nitrate uptake, which limits the utilization of the large amount of nitrate available in agricultural soils. To understand the regulation of nitrate uptake in sugarcane roots, we identified the major canonical nitrate transporter genes (NRTs-NITRATE TRANSPORTERS) and then determined their expression profiles in roots under contrasting N conditions. Correlation of gene expression with 15N-nitrate uptake revealed that under N deprivation or inorganic N (ammonium or nitrate) supply in N-sufficient roots, the regulation of ScNRT2.1 and ScNRT3.1 expression is the predominant mechanism for the modulation of the activity of the nitrate high-affinity transport system. Conversely, in N-deficient roots, the induction of ScNRT2.1 and ScNRT3.1 transcription is not correlated with the marked repression of nitrate uptake in response to nitrate resupply or high N provision, which suggested the existence of a posttranscriptional regulatory mechanism. Our findings suggested that high-affinity nitrate uptake is regulated at the transcriptional and presumably at the posttranscriptional levels based on the physiological N status and that the regulation of NRT2.1 and NRT3.1 activity is likely a determinant mechanism for the discrimination against nitrate uptake observed in sugarcane roots, which contributes to the low NUE in this crop species.

Genome-wide identification and in silico analysis of NPF, NRT2, CLC and SLAC1/SLAH nitrate transporters in hexaploid wheat (Triticum aestivum)

Nitrogen transport is one of the most important processes in plants mediated by specialized transmembrane proteins. Plants have two main systems for nitrogen uptake from soil and its transport within the system—a low-affinity transport system and a high-affinity transport system. Nitrate transporters are of special interest in cereal crops because large amount of money is spent on N fertilizers every year to enhance the crop productivity. Till date four gene families of nitrate transporter proteins; NPF (nitrate transporter 1/peptide transporter family), NRT2 (nitrate transporter 2 family), the CLC (chloride channel family), and the SLAC/SLAH (slow anion channel-associated homologues) have been reported in plants. In our study, in silico mining of nitrate transporter genes along with their detailed structure, phylogenetic and expression analysis was carried out. A total of 412 nitrate transporter genes were identified in hexaploid wheat genome using HMMER based homology searches in IWGSC Refseq v2.0. Out of those twenty genes were root specific, 11 leaf/shoot specific and 17 genes were grain/spike specific. The identification of nitrate transporter genes in the close proximity to the previously identified 67 marker-traits associations associated with the nitrogen use efficiency related traits in nested synthetic hexaploid wheat introgression library indicated the robustness of the reported transporter genes. The detailed crosstalk between the genome and proteome and the validation of identified putative candidate genes through expression and gene editing studies may lay down the foundation to improve nitrogen use efficiency of cereal crops.

Bacterial chemotaxis to saccharides is governed by a trade-off between sensing and uptake

To swim up gradients of nutrients, E. coli senses nutrient concentrations within its periplasm. For small nutrient molecules, periplasmic concentrations typically match extracellular concentrations. However, this is not necessarily the case for saccharides, such as maltose, which are transported into the periplasm via a specific porin. Previous observations have shown that, under various conditions, E. coli limits maltoporin abundance so that, for extracellular micromolar concentrations of maltose, there are predicted to be only nanomolar concentrations of free maltose in the periplasm. Thus, in the micromolar regime, the total uptake of maltose from the external environment into the cytoplasm is limited not by the abundance of cytoplasmic transport proteins but by the abundance of maltoporins. Here, we present results from experiments and modeling suggesting that this porin-limited transport enables E. coli to sense micromolar gradients of maltose despite having a high-affinity ABC transport system that is saturated at these micromolar levels. We used microfluidic assays to study chemotaxis of E. coli in various gradients of maltose and methyl-aspartate and leveraged our experimental observations to develop a mechanistic transport-and-sensing chemotaxis model. Incorporating this model into agent-based simulations, we discover a trade-off between uptake and sensing: although high-affinity transport enables higher uptake rates at low nutrient concentrations, it severely limits the range of dynamic sensing. We thus propose that E. coli may limit periplasmic uptake to increase its chemotactic sensitivity, enabling it to use maltose as an environmental cue.

Oilseed Rape Cultivars Show Diversity of Root Morphologies with the Potential for Better Capture of Nitrogen

The worldwide demand for vegetable oils is rising. Oilseed rape (Brassica napus) diversifies cereal dominated crop rotations but requires important nitrogen input. Yet, the root organ is offering an untapped opportunity to improve the nitrogen capture in soil. This study evaluates three culture systems in controlled environment, to observe root morphology and to identify root attributes for superior biomass production and nitrogen use. The phenotypic diversity in a panel of 55 modern winter oilseed rape cultivars was screened in response to two divergent nitrate supplies. Upon in vitro and hydroponic cultures, a large variability for root morphologies was observed. Root biomass and morphological traits positively correlated with shoot biomass or leaf area. The activities of high-affinity nitrate transport systems correlated negatively with the leaf area, while the combined high- and low-affinity systems positively with the total root length. The X-ray computed tomography permitted to visualize the root system in pipes filled with soil. The in vitro root phenotype at germination stage was indicative of lateral root deployment in soil-grown plants. This study highlights great genetic potential in oilseed rape, which could be manipulated to optimize crop root characteristics and nitrogen capture with substantial implications for agricultural production.

Phylogenomic and Microsynteny Analysis Provides Evidence of Genome Arrangements of High-Affinity Nitrate Transporter Gene Families of Plants.

Nitrate transporter 2 (NRT2) and NRT3 or nitrate-assimilation-related 2 (NAR2) proteins families form a two-component, high-affinity nitrate transport system, which is essential for the acquisition of nitrate from soils with low N availability. An extensive phylogenomic analysis across land plants for these families has not been performed. In this study, we performed a microsynteny and orthology analysis on the NRT2 and NRT3 genes families across 132 plants (Sensu lato) to decipher their evolutionary history. We identified significant differences in the number of sequences per taxonomic group and different genomic contexts within the NRT2 family that might have contributed to N acquisition by the plants. We hypothesized that the greater losses of NRT2 sequences correlate with specialized ecological adaptations, such as aquatic, epiphytic, and carnivory lifestyles. We also detected expansion on the NRT2 family in specific lineages that could be a source of key innovations for colonizing contrasting niches in N availability. Microsyntenic analysis on NRT3 family showed a deep conservation on land plants, suggesting a high evolutionary constraint to preserve their function. Our study provides novel information that could be used as guide for functional characterization of these gene families across plant lineages.

Increasing Temperature Alters the Effects of Extracellular Copper on Thalassiosira Pseudonana Physiology and Transcription

Copper (Cu) is essential for many physiological processes in phytoplankton, including electron-transfer reactions and high-affinity Fe transport systems. However, at high concentrations, Cu can have a toxic effect on phytoplankton. Phytoplankton’s tolerance to certain toxicants in marine ecosystems cannot always be solely attributed to the presence of compounds, as various environmental factors including temperature can also indirectly influence their effects on organisms. In this study, we investigated the effects of different Cu concentrations (0, 19.6, 160, 800, and 8000 nM) on the growth and physiological changes in the centric diatom Thalassiosira pseudonana (CCMP 1335) when simultaneously applied with temperature cultivation of 20 °C and 25 °C. At low (0 nM) and high (8000 nM) Cu concentrations, the growth rate of T. pseudonana was inhibited, though an increase in temperature lessened this inhibition. There were no significant changes in the POC:PON ratio during all of the treatments. However, increasing the temperature significantly decreased the POC: POP, PON: POP and BSi: POP ratios of T. pseudonana. The intracellular Cu content of T. pseudonana varied from 0.13 to 13.28 fg cell−1 in response to increases in ambient Cu concentrations. Lastly, an increase in the Cu concentration decreased the transcriptional expression of CTR (copper transporter), 3Hfcp (photosynthetic protein), and Sit1 (silica shell formation) encoding genes. In conclusion, our results indicated that T. pseudonana can adapt to physiological processes and molecular mechanisms in response to varying Cu concentrations and ambient temperatures.

GARP transcription factors repress Arabidopsis nitrogen starvation response via ROS-dependent and -independent pathways.

Plants need to cope with strong variations of nitrogen availability in the soil. Although many molecular players are being discovered concerning how plants perceive NO3- provision, it is less clear how plants recognize a lack of nitrogen. Following nitrogen removal, plants activate their nitrogen starvation response (NSR), which is characterized by the activation of very high-affinity nitrate transport systems (NRT2.4 and NRT2.5) and other sentinel genes involved in N remobilization such as GDH3. Using a combination of functional genomics via transcription factor perturbation and molecular physiology studies, we show that the transcription factors belonging to the HHO subfamily are important regulators of NSR through two potential mechanisms. First, HHOs directly repress the high-affinity nitrate transporters, NRT2.4 and NRT2.5. hho mutants display increased high-affinity nitrate transport activity, opening up promising perspectives for biotechnological applications. Second, we show that reactive oxygen species (ROS) are important to control NSR in wild-type plants and that HRS1 and HHO1 overexpressors and mutants are affected in their ROS content, defining a potential feed-forward branch of the signaling pathway. Taken together, our results define the relationships of two types of molecular players controlling the NSR, namely ROS and the HHO transcription factors. This work (i) up opens perspectives on a poorly understood nutrient-related signaling pathway and (ii) defines targets for molecular breeding of plants with enhanced NO3- uptake.

Genome-wide analysis in response to nitrogen and carbon identifies regulators for root AtNRT2 transporters.

In Arabidopsis (Arabidopsis thaliana), the High-Affinity Transport System (HATS) for root nitrate (NO3-) uptake depends mainly on four NRT2 NO3- transporters, namely NRT2.1, NRT2.2, NRT2.4, and NRT2.5. The HATS is the target of many regulations to coordinate nitrogen (N) acquisition with the N status of the plant and with carbon (C) assimilation through photosynthesis. At the molecular level, C and N signaling pathways control gene expression of the NRT2 transporters. Although several regulators of these transporters have been identified in response to either N or C signals, the response of NRT2 gene expression to the interaction of these signals has never been specifically investigated, and the underlying molecular mechanisms remain largely unknown. To address this question we used an original systems biology approach to model a regulatory gene network targeting NRT2.1, NRT2.2, NRT2.4, and NRT2.5 in response to N/C signals. Our systems analysis of the data identified three transcription factors, TGA3, MYC1, and bHLH093. Functional analysis of mutants combined with yeast one-hybrid experiments confirmed that all three transcription factors are regulators of NRT2.4 or NRT2.5 in response to N or C signals. These results reveal a role for TGA3, MYC1, and bHLH093 in controlling the expression of root NRT2 transporter genes.