Ammonium Transport Research Articles

SpiAMT1d: molecular characterization, localization, and potential role in coral calcification of an ammonium transporter in Stylophora pistillata

SpiAMT1d: molecular characterization, localization, and potential role in coral calcification of an ammonium transporter in Stylophora pistillata

Harnessing root-foraging capacity to improve nutrient-use efficiency for sustainable maize production

Producing more food with less input is imperative in feeding the increasing world population. Here, for the first time we characterize the multipartite responses of maize to nutrient-rich patches and demonstrate significant synergistic effects between root morphological and physiological responses on improving nutrient-use efficiency and yield. Our results showed that maize root length, lateral root formation, acid phosphatase (APase) secretion, and expression of ammonium transporter genes increased in the ammonium-containing patches; moreover, ammonium-induced rhizosphere acidification enhanced activity of APase. These root responses were associated with improved maize nutrition and growth even with reduced fertilizer input, suggesting an adaptive benefit of synergistic root foraging strategies. These new findings improve our understanding of root-foraging responses to nutrient-specific cues and are crucial for engineering root and rhizosphere properties to improve nutrient-use efficiency for sustainable crop production. • Producing more food with less input is imperative for sustainable intensification of agriculture. • Rhizosphere dynamics is key to controlling nutrient-use efficiency (NUE). • Localized nutrient-supply maximizes root/rhizosphere efficiency for improving NUE. • Harnessing root foraging provides a new approach for sustainable maize production.

Molecular spectroscopy and docking simulation revealed the binding mechanism of phenol onto anammox sludge extracellular polymeric substances

The rapid development of chemical industry has induced to the large amount of phenolic wastewater production. When the promising anaerobic ammonium oxidation (anammox) was employed to treat the industrial wastewater, phenolic compounds would possibly inhibit the microbial performance. Extracellular polymeric substances (EPSs) play an essential role in protecting cells from being intoxicated by phenolic compound while the distinct mechanism remains elusive. In this work, the interaction of phenol with anammox sludge EPSs and transmembrane ammonium transport (Amt) domain was explored at molecular level by using spectral method and molecular docking simulation. It was found that phenol statically quenched the fluorescent components of EPSs and the protein component dominated the interaction between EPSs and phenol. The overall interaction was an entropy-driven process with hydrophobic interaction as the main driving force, and the CO vibration responded preferentially. As phenol continued to penetrate into the cell surface, there were hydrogen bond, hydrophobic interaction force and π-π base-stacking forces between the Amt domain and phenol. The interaction between phenol and amino acid residues of the Amt domain would interfere the NH4+ transport and further affect the activity of anammox sludge. This work is beneficial for in-depth understanding the role of EPSs in protecting anammox sludge from inhibiting by phenolic pollutants.

Impact of microplastics from polyethylene and biodegradable mulch films on rice (Oryza sativa L.)

Extensive use of plastic film mulch has resulted in accumulation of a large amount of residual plastic, which will eventually fragment into microplastics in agricultural soils. However, it is unclear how microplastics from plastic mulch film affect crops. To address this issue, rice plants exposed to microplastics derived from poly(butyleneadipate-co-terephthalate) (PBAT)-based biodegradable mulch film (BM) and polyethylene (PE) mulch film (PM) were investigated for plant growth, physio-biochemical processes, and gene expressions. Both types of microplastics significantly reduced the height and dry weight of rice plant. Oxidative stress was induced by microplastics in rice shoot and root, with levels of ROS relatively higher under treatment PM than that under treatment BM. Transcriptomic data showed that more genes were down-regulated by treatment PM than that by treatment BM. Genes encoding ammonium and nitrate transporters were down-regulated by both types of microplastics in rice roots at vegetative stage, whereas up-regulated at reproductive stage, as compared to their respective treatment with no microplastics (CK). Similar results regarding phenylpropanoid biosynthesis pathway and lignin content were also observed in rice roots. Net photosynthetic rate and SPAD value were significantly inhibited by treatments BM and PM in rice shoot, and the expression of genes involved in light reaction was reduced at vegetative stage, whereas there were no differences of them at reproductive stage, as compared to their respective treatment CK. Our study suggests that microplastics from BM and PM both affect the growth of rice plants via nitrogen metabolism and photosynthesis. The negative effects imposed by both types of microplastics on rice plant can be mitigated with the growth of plants, and the negative effects of microplastics from PE mulch film on rice plant are relatively stronger than that from the PBAT-based biodegradable film.

Comparative Proteomic Analysis of Fucosylated Glycoproteins Produced by Bacteroides thetaiotaomicron Under Different Polysaccharide Nutrition Conditions.

Bacteroides thetaiotaomicron, one of the most eminent representative gut commensal Bacteroides species, is able to use the L-fucose in host-derived and dietary polysaccharides to modify its capsular polysaccharides and glycoproteins through a mammalian-like salvage metabolic pathway. This process is essential for the colonization of the bacteria and for symbiosis with the host. However, despite the importance of fucosylated proteins (FGPs) in B. thetaiotaomicron, their types, distribution, and functions remain unclear. In this study, the effects of different polysaccharide (corn starch, mucin, and fucoidan) nutrition conditions on newly synthesized FGPs expressions and fucosylation are investigated using a chemical biological method based on metabolic labeling and bioorthogonal reaction. According to the results of label-free quantification, 559 FGPs (205 downregulated and 354 upregulated) are affected by the dietary conditions. Of these differentially expressed proteins, 65 proteins show extremely sensitive to polysaccharide nutrition conditions (FGPs fold change/global protein fold change ≥2.0 or ≤0.5). Specifically, the fucosylation of the chondroitin sulfate ABC enzyme, Sus proteins, and cationic efflux system proteins varies significantly upon the addition of mucin, corn starch, or fucoidan. Moreover, these polysaccharides can trigger an appreciable increase in the fucosylation level of the two-component system and ammonium transport proteins. These results highlight the efficiency of the combined metabolic glycan labeling and bio-orthogonal reaction in enriching the intestinal Bacteroides glycoproteins. Moreover, it emphasizes the sensitivity of Bacteroides fucosylation to polysaccharide nutrition conditions, which allows for the regulation of bacterial growth.

Effects of enhancing leaf nitrogen output on tiller growth and carbon metabolism in rice

<p id="C3">Nitrogen fertilizer application is one of the main cultivation measures to raise the yield, and high nitrogen level has limited contribution to grain yield due to limited nitrogen translocation in rice. To clarify the effects of nitrogen allocation on rice growth, we constructed<italic> pOsSUT1::AtAMT1.2 </italic>transgenic rice, the ammonium transporter gene <italic>AtAMT1.2</italic> specific expression in phloem to promote leaf nitrogen output. The growth and yield of transgenic plants were measured under HN (high nitrogen) and LN (low nitrogen) conditions. Compared to WT plants, more tillers and higher grain yield were detected in transgenic plants in response to HN condition. The sugar output in leaves was increased, and the relative expression levels of the strigolactone pathway related genes<italic> OsTB1</italic> and <italic>OsD14</italic> in tiller buds were down-regulated. Our results indicated that the increase of leaf nitrogen export by overexpressing <italic>AtAMT1.2</italic> gene could promote sugar translocation from leaves to tillering buds, which improved the growth of tiller, increased the effective tiller number and nitrogen use efficiency.

Ammonium (NH4+) transport processes in the riverbank under varying hydrologic conditions

Attenuation of groundwater ammonium (NH4+) is expected to occur through redox reaction and adsorption of the riverbank. Previous studies determined that NH4+ mostly degraded through nitrification along subsurface flow, however, the adsorption capacities of riverbanks were always ignored in the NH4+ reduction processes. In this study, field experiments were conducted in the Fuliji section of the Xiaosuixin River, China, to understand NH4+ transport and attenuation under rainfall events-induced river and groundwater interactions. The results indicated that the NH4+ concentration in river water increased significantly after heavy rainfall events and reached a peak of about 5.88 mg L−1, and the lag time was more than 2 weeks compared with the river peak stage. Adsorption plays a dominant role in attenuation of NH4+ in riverbank with high amounts of organic materials and clay minerals, reducing its concentration to less than 0.05 mg L−1. A two-dimensional lateral exchange and transport model of NH4+ was developed and calibrated against observations in the aquifer, and an exponential reduction pattern of NH4+ was identified. The model's possible implications about the effects of varying hydrologic changes (i.e., peak stage and lag time differences between river and groundwater) on NH4+ transport were also discussed. Thus, the effects of river-groundwater interactions on nitrogen pollution should be taken into consideration in river regulation strategies in order to ensure proper hydrogeochemical functioning of river–aquifer interfaces and related ecosystems.

Ammonium transporter 1 increases rice resistance to sheath blight by promoting nitrogen assimilation and ethylene signalling.

SummarySheath blight (ShB) significantly threatens rice yield production. However, the underlying mechanism of ShB defence in rice remains largely unknown. Here, we identified a highly ShB‐susceptible mutant Ds‐m which contained a mutation at the ammonium transporter 1;1 (AMT1;1) D358N. AMT1;1 D358N interacts with AMT1;1, AMT1;2 and AMT1;3 to inhibit the ammonium transport activity. The AMT1 RNAi was more susceptible and similar to the AMT1;1 D358N mutant; however, plants with higher NH4 + uptake activity were less susceptible to ShB. Glutamine synthetase 1;1 (GS1;1) mutant gs1;1 and overexpressors (GS1;1 OXs) were more and less susceptible to ShB respectively. Furthermore, AMT1;1 overexpressor (AMT1;1 OX)/gs1;1 and gs1;1 exhibited a similar response to ShB, suggesting that ammonium assimilation rather than accumulation controls the ShB defence. Genetic and physiological assays further demonstrated that plants with higher amino acid or chlorophyll content promoted rice resistance to ShB. Interestingly, the expression of ethylene‐related genes was higher in AMT1;1 OX and lower in RNAi mutants than in wild‐type. Also, ethylene signalling positively regulated rice resistance to ShB and NH4 + uptake, suggesting that ethylene signalling acts downstream of AMT and also NH4 + uptake is under feedback control. Taken together, our data demonstrated that the AMT1 promotes rice resistance to ShB via the regulation of diverse metabolic and signalling pathways.

LbAMT3-1, an ammonium transporter induced by arbuscular mycorrhizal in Lycium barbarum, confers tobacco with higher mycorrhizal levels and nutrient uptake.

An ammonium transporter LbAMT3-1 overexpression increases the arbuscular abundance of mycorrhizal that opens the possibility of using LbAMT3-1 in breeding programs to improve symbiotic nutrient uptake in Lycium barbarum. Nitrogen (N) is one of the most essential nutrients required by plants and limits net primary production much of the time in most terrestrial ecosystems. Arbuscular mycorrhizal (AM) fungi can enhance plant nutrient uptake and improve plant productivity in nutrient limit ecosystems. Here, we identified an ammonia transporter, LbAMT3-1, specifically induced by AM fungi in Lycium barbarum. To understand the expression characteristics and biological functions, LbAMT3-1 was cloned, characterized, and overexpressed in Nicotiana tabacum (tobacco). A BLAST search identified the coding sequence for LbAMT3-1 with an open-reading frame of 1473bp. Reverse transcription polymerase chain reaction (RT-PCR) analysis indicated that, besides mycorrhizal roots, LbAMT3-1 were barely detectable in other tissues, including stems and leaves. Promoter-GUS assay showed that GUS staining was detected in mycorrhizal roots, and GUS activity driven by the LbAMT3-1 promoter was exclusively confined to root cells containing arbuscules. LbAMT3-1 functionally complemented the yeast mutant efficiently, and yeast expressing LbAMT3-1 showed well growth on the agar medium with 0.02, 0.2, and 2mM NH4+ supply. Moreover, overexpression of LbAMT3-1 in N. tabacum resulted a significant increase in arbuscular abundance and enhanced the nutrient acquisition capacity of mycorrhizal plants. Based on the results of our study, we propose that overexpression of LbAMT3-1 can promote P and N uptake of host plants through the mycorrhizal pathway, and increase the colonization intensity and arbuscular abundance, which opens the possibility of using LbAMT3-1 in breeding programs.

Transcriptional regulation of metal metabolism- and nutrient absorption-related genes in Eucalyptus grandis by arbuscular mycorrhizal fungi at different zinc concentrations

BackgroundEucalyptus spp. are candidates for phytoremediation in heavy metal (HM)-polluted soils as they can adapt to harsh environments, grow rapidly, and have good economic value. Arbuscular mycorrhizal fungi (AMF) are the most widely distributed plant symbiotic fungi in nature, and they play an important role in promoting the phytoremediation of HM-polluted soils. However, few studies have evaluated the HM detoxification mechanism of E. spp. in symbiosis with AMF, and thus, the molecular mechanism remains unclear.ResultsThe gene transcription and metabolic pathways of E. grandis were studied with and without inoculation with AMF and at different zinc (Zn) concentrations. Here, we focused on the transcript level of six HM-related gene families (ZNT, COPT/Ctr, YSL, ZIFL and CE). Under high-Zn conditions, thirteen genes (ZNT:2, COPT/Ctr:5, YSL:3, ZIFL:1, CE:2) were upregulated, whereas ten genes (ZNT:3, COPT/Ctr:2, YSL:3, ZIFL:1, CE:1) were downregulated. With AMF symbiosis under high-Zn conditions, ten genes (ZNT:4, COPT/Ctr:2, YSL:3, CE:1) were upregulated, whereas nineteen genes (ZNT:9, COPT/Ctr:2, YSL:3, ZIFL:4, CE:1) were downregulated. Under high-Zn conditions, genes of three potassium-related transporters, six phosphate transporters (PHTs), and two nitrate transporters (NRTs) were upregulated, whereas genes of four potassium-related transporters,four PHTs, and four nitrogen-related transporters were downregulated. With AMF symbiosis under high-Zn conditions, genes of two potassium-related transporters, six ammonium transporters (AMTs) and five PHTs were upregulated, whereas genes of six potassium-related transporters, two AMTs and five PHTs were downregulated.ConclusionsOur results indicates that AMF increases the resistance of E. grandis to high-Zn stress by improving nutrients uptake and regulating Zn uptake at the gene transcription level. Meanwhile, our findings provide a genome-level resource for the functional assignments of key genes regulated by Zn treatment and AM symbiosis in six HM-associated gene families and macromineral nutrient-related gene families of E. grandis. This may contribute to the elucidation of the molecular mechanisms of the response to Zn stress in E. grandis with AM symbiosis at the aspect of the interaction between HM tolerance and nutrient acquisition.

Collective Behavior of Urease pH Clocks in Nano- and Microvesicles Controlled by Fast Ammonia Transport.

The transmission of chemical signals via an extracellular solution plays a vital role in collective behavior in cellular biological systems and may be exploited in applications of lipid vesicles such as drug delivery. Here, we investigated chemical communication in synthetic micro- and nanovesicles containing urease in a solution of urea and acid. We combined experiments with simulations to demonstrate that the fast transport of ammonia to the external solution governs the pH–time profile and synchronizes the timing of the pH clock reaction in a heterogeneous population of vesicles. This study shows how the rate of production and emission of a small basic product controls pH changes in active vesicles with a distribution of sizes and enzyme amounts, which may be useful in bioreactor or healthcare applications.

Ammonium transporters cooperatively regulate rice crown root formation responding to ammonium nitrogen.

Crown roots (CRs) are major components of the rice root system. They form at the basal node of the shoot, and their development is greatly influenced by environmental factors. Ammonium nitrogen is known to impact plant root development through ammonium transporters (AMTs), but it remains unclear whether ammonium and AMTs play roles in rice CR formation. In this study, we revealed a significant role of ammonium, rather than nitrate, in regulating rice CR development. High ammonium supply increases CR formation but inhibits CR elongation. Genetic evidence showed that ammonium regulation of CR development relies on ammonium uptake mediated jointly by ammonium transporters OsAMT1;1, OsAMT1;2; OsAMT1;3, and OsAMT2;1, but not on root acidification which was the result of ammonium uptake. OsAMTs are also needed for glutamine-induced CR formation. Furthermore, we showed that polar auxin transport dependent on the PIN auxin efflux carriers acts downstream of ammonium uptake and assimilation to activate local auxin signaling at CR primordia, in turn promoting CR formation. Taken together, our results highlight a critical role for OsAMTs in cooperatively regulating CR formation through regulating auxin transport under nitrogen-rich conditions.

Integrated application of multi-omics approach and biochemical assays provides insights into physiological responses to saline-alkaline stress in the gills of crucian carp (Carassius auratus)

Given the decline of freshwater resources in recent years, the accessible space for freshwater aquaculture is rapidly shrinking, and aquaculture in saline-alkaline water has become a critical approach to meet the rising demand. However, the molecular mechanism behind the adverse effects of saline-alkaline water on fish and the regulatory mechanism in fish tolerance remains unclear. Here, adult crucian carp (Carassius auratus) were exposed to 60 mmol/L NaHCO3 for 30 days. It was observed that long-term carbonate alkalinity (CA) exposure not only caused gill oxidative stress but also changed the levels of several physiological parameters associated with ammonia transport, including blood ammonia, urea nitrogen (BUN), glutamine (Gln), and glutamine synthetase (GS). According to the metabolomics study, differential metabolites (DMs) engaged in various metabolic pathways, such as glycerophospholipid metabolism, sphingolipid metabolism, and arachidonic acid metabolism. In addition, transcriptomics data showed that differentially expressed genes (DEGs) were closely related to ammonia transport, apoptosis, and immunological response. In general, comprehensive multi-omics and biochemical analysis revealed that crucian carp might adopt Rh glycoprotein as a carrier to mediate ammonia transport and increase glutamine and urea synthesis under long-term high saline-alkaline stress to mitigate the adverse effects of blocked ammonia excretion. Simultaneously, saline-alkaline stress caused the destruction of the antioxidant system and the disorder of lipid metabolism in the crucian carp gills, which induced apoptosis and immunological response. To our knowledge, this is the first study to investigate fish's molecular and metabolic mechanisms under saline-alkaline stress using integrated metabolomics, transcriptomics, and biochemical assays. Overall, the results of this study provided new insights into the molecular mechanism behind the adverse effects of saline-alkaline water on fish and the regulatory mechanism in fish tolerance.

Structural and molecular dynamics of ammonia transport in Staphylococcus aureus NH3-dependent NAD synthetase

Ammonia dependent NAD+ synthetase from multi drug resistance Staphylococcus aureus catalyzes ATP dependent formation of NAD+ from deamido-NAD+ and ammonia at the synthetase active site. Binding of ATP accompanies a large movement of flexible loop region (205-225) acting as a lid to the catalytic core. A 17 Å long ammonia tunnel with an entry and exit radius of 3.5 Å and 3.2 Å respectively allows transfer of ammonia from surface to the active site of the enzyme in each monomer to attack the C7N=O7N linkage of transient intermediate NAD-adenylate thus releasing NAD+. In this study, we report structural details of ammonia transport tunnel in Staphylococcus aureus NH3-dependent NAD synthetase and compared their architecture and dynamics with other bacterial and eukaryotic enzymes. Tunnel shows conformational variations in apo and substrate complexes and is less intricate compared to glutamine dependent counterparts. We have also performed steered molecular dynamic simulations of ammonia transport across the tunnel in enzyme-intermediate complex which reveals critical bottleneck residues and structural determinants during ammonium migration. Ordered water molecules and conserved charged residues form a network of hydrogen bonds and electrostatic interaction which facilitate the ammonium movement towards the active center. Analysis of the sMD simulated structural snapshots delineates the conformational reshaping of ammonia tunnel at the different step of the enzymatic reaction. Tunnel architecture and environment could offer the new target site to design novel small molecule inhibitors for the development of more efficient therapeutics against multi drug resistant S. aureus strains.

First Author Spotlight.

First Author Spotlight.

Single cell‐type transcriptome profiling reveals genes that promote nitrogen fixation in the infected and uninfected cells of legume nodules

Single cell‐type transcriptome profiling reveals genes that promote nitrogen fixation in the infected and uninfected cells of legume nodules

Transcriptome and Proteomics Analysis of Wheat Seedling Roots Reveals That Increasing NH4 +/NO3 - Ratio Induced Root Lignification and Reduced Nitrogen Utilization.

Wheat growth and nitrogen (N) uptake gradually decrease in response to high NH4+/NO3– ratio. However, the mechanisms underlying the response of wheat seedling roots to changes in NH4+/NO3– ratio remain unclear. In this study, we investigated wheat growth, transcriptome, and proteome profiles of roots in response to increasing NH4+/NO3– ratios (Na: 100/0; Nr1: 75/25, Nr2: 50/50, Nr3: 25/75, and Nn: 0/100). High NH4+/NO3– ratio significantly reduced leaf relative chlorophyll content, Fv/Fm, and ΦII values. Both total root length and specific root length decreased with increasing NH4+/NO3– ratios. Moreover, the rise in NH4+/NO3– ratio significantly promoted O2– production. Furthermore, transcriptome sequencing and tandem mass tag-based quantitative proteome analyses identified 14,376 differentially expressed genes (DEGs) and 1,819 differentially expressed proteins (DEPs). The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis indicated that glutathione metabolism and phenylpropanoid biosynthesis were the main two shared enriched pathways across ratio comparisons. Upregulated DEGs and DEPs involving glutathione S-transferases may contribute to the prevention of oxidative stress. An increment in the NH4+/NO3– ratio induced the expression of genes and proteins involved in lignin biosynthesis, which increased root lignin content. Additionally, phylogenetic tree analysis showed that both A0A3B6NPP6 and A0A3B6LM09 belong to the cinnamyl-alcohol dehydrogenase subfamily. Fifteen downregulated DEGs were identified as high-affinity nitrate transporters or nitrate transporters. Upregulated TraesCS3D02G344800 and TraesCS3A02G350800 were involved in ammonium transport. Downregulated A0A3B6Q9B3 is involved in nitrate transport, whereas A0A3B6PQS3 is a ferredoxin-nitrite reductase. This may explain why an increase in the NH4+/NO3– ratio significantly reduced root NO3–-N content but increased NH4+-N content. Overall, these results demonstrated that increasing the NH4+/NO3– ratio at the seedling stage induced the accumulation of reactive oxygen species, which in turn enhanced root glutathione metabolism and lignification, thereby resulting in increased root oxidative tolerance at the cost of reducing nitrate transport and utilization, which reduced leaf photosynthetic capacity and, ultimately, plant biomass accumulation.

Correcting Differential Gene Expression Analysis for Cyto-Architectural Alterations in Substantia Nigra of Parkinson's Disease Patients Reveals Known and Potential Novel Disease-Associated Genes and Pathways.

Several studies have analyzed gene expression profiles in the substantia nigra to better understand the pathological mechanisms causing Parkinson’s disease (PD). However, the concordance between the identified gene signatures in these individual studies was generally low. This might have been caused by a change in cell type composition as loss of dopaminergic neurons in the substantia nigra pars compacta is a hallmark of PD. Through an extensive meta-analysis of nine previously published microarray studies, we demonstrated that a big proportion of the detected differentially expressed genes was indeed caused by cyto-architectural alterations due to the heterogeneity in the neurodegenerative stage and/or technical artefacts. After correcting for cell composition, we identified a common signature that deregulated the previously unreported ammonium transport, as well as known biological processes such as bioenergetic pathways, response to proteotoxic stress, and immune response. By integrating with protein interaction data, we shortlisted a set of key genes, such as LRRK2, PINK1, PRKN, and FBXO7, known to be related to PD, others with compelling evidence for their role in neurodegeneration, such as GSK3β, WWOX, and VPC, and novel potential players in the PD pathogenesis. Together, these data show the importance of accounting for cyto-architecture in these analyses and highlight the contribution of multiple cell types and novel processes to PD pathology, providing potential new targets for drug development.

Genome-wide identification, characterization, and expression analysis of the ammonium transporter gene family in tea plants (Camellia sinensis L.).

As a preferred nitrogen form, ammonium (NH4 + ) transport via specific transporters is particularly important for the growth and development of tea plants (Camellia sinensis L.). However, our understanding of the functions of the AMT family in tea plants is limited. We identified and named 16 putative AMT genes according to phylogenetic analysis. All CsAMT genes were divided into three groups, distributed on 12 chromosomes with only one segmental duplication repetition. The CsAMT genes showed different expression levels in different organs, and most of them were expressed mainly in the apical buds and roots. Complementation analysis of yeast mutants showed that CsAMTs restored the uptake of NH4 + . This study provides insights into the genome-wide distribution and spatial expression of AMT genes in tea plants.

Nitrogen nutrition modifies the susceptibility of Arabidopsis thaliana to the necrotrophic fungus, Alternaria brassicicola.

The impact of the form of nitrogen (N) source (nitrate versus ammonium) on the susceptibility to Alternaria brassicicola, a necrotrophic fungus, has been examined in Arabidopsis thaliana at the rosette stage. Nitrate nutrition was found to increase fungal lesions considerably. There was a similar induction of defence gene expression following infection under both N nutritions, except for the phytoalexin deficient 3 gene, which was overexpressed with nitrate. Nitrate also led to a greater nitric oxide production occurring in planta during the saprophytic growth and lower nitrate reductase (NIA1) expression 7 days after inoculation. This suggests that nitrate reductase-dependent nitric oxide production had a dual role, whereby, despite its known role in the generic response to pathogens, it affected plant metabolism, and this facilitated fungal infection. In ammonium-grown plants, infection with A. brassicicola induced a stronger gene expression of ammonium transporters and significantly reduced the initially high ammonium content in the leaves. There was a significant interaction between N source and inoculation (presence versus absence of the fungus) on the total amino acid content, while N nutrition reconfigured the spectrum of major amino acids. Typically, a higher content of total amino acid, mainly due to a stronger increase in asparagine and glutamine, is observed under ammonium nutrition while, in nitrate-fed plants, glutamate was the only amino acid which content increased significantly after fungal inoculation. N nutrition thus appears to control fungal infection via a complex set of signalling and nutritional events, shedding light on how nitrate availability can modulate disease susceptibility.