Impact of low light intensity on biomass partitioning and genetic diversity in a chickpea mapping population.

With recent climatic changes, the reduced access to solar radiation has become an emerging threat to chickpeas' drought tolerance capacity under rainfed conditions. This study was conducted to assess, and understand the effects of reduced light intensity and quality on plant morphology, root development, and identifying resistant sources from a Sonali/PBA Slasher mapping population. We evaluated 180 genotypes, including recombinant inbred lines (RILs), parents, and commercial checks, using a split-block design with natural and low light treatments. Low light conditions, created by covering one of the two benches inside two growth chambers with a mosquito net, reduced natural light availability by approximately 70%. Light measurements encompassed photosynthetic photon flux density, as well as red, and far-red light readings taken at various stages of the experiment. The data, collected from plumule emergence to anthesis initiation, encompassed various indices relevant to root, shoot, and carbon gain (biomass). Statistical analysis examined variance, treatment effects, heritability, correlations, and principal components (PCs). Results demonstrated significant reductions in root biomass, shoot biomass, root/shoot ratio, and plant total dry biomass under suboptimal light conditions by 52.8%, 28.2%, 36.3%, and 38.4%, respectively. Plants also exhibited delayed progress, taking 9.2% longer to produce their first floral buds, and 19.2% longer to commence anthesis, accompanied by a 33.4% increase in internodal lengths. A significant genotype-by-environment interaction highlighted differing genotypic responses, particularly in traits with high heritability (> 77.0%), such as days to anthesis, days to first floral bud, plant height, and nodes per plant. These traits showed significant associations with drought tolerance indicators, like root, shoot, and plant total dry biomass. Genetic diversity, as depicted in a genotype-by-trait biplot, revealed contributions to PC1 and PC2 coefficients, allowing discrimination of low-light-tolerant RILs, such as 1_52, 1_73, 1_64, 1_245, 1_103, 1_248, and 1_269, with valuable variations in traits of interest. These RILs could be used to breed desirable chickpea cultivars for sustainable production under water-limited conditions. This study concludes that low light stress disrupts the balance between root and shoot morphology, diverting photosynthates to vegetative structures at the expense of root development. Our findings contribute to a better understanding of biomass partitioning under limited-light conditions, and inform breeding strategies for improved drought tolerance in chickpeas.

Arbuscular-Mycorrhizal Symbiosis in Medicago Regulated by the Transcription Factor MtbHLHm1;1 and the Ammonium Facilitator Protein MtAMF1;3.

Root systems of most land plants are colonised by arbuscular mycorrhiza fungi. The symbiosis supports nutrient acquisition strategies predominantly associated with plant access to inorganic phosphate. The nutrient acquisition is enhanced through an extensive network of external fungal hyphae that extends out into the soil, together with the development of fungal structures forming specialised interfaces with root cortical cells. Orthologs of the bHLHm1;1 transcription factor, previously described in soybean nodules (GmbHLHm1) and linked to the ammonium facilitator protein GmAMF1;3, have been identified in Medicago (Medicago truncatula) roots colonised by AM fungi. Expression studies indicate that transcripts of both genes are also present in arbuscular containing root cortical cells and that the MtbHLHm1;1 shows affinity to the promoter of MtAMF1;3. Both genes are induced by AM colonisation. Loss of Mtbhlhm1;1 expression disrupts AM arbuscule abundance and the expression of the ammonium transporter MtAMF1;3. Disruption of Mtamf1;3 expression reduces both AM colonisation and arbuscule development. The respective activities of MtbHLHm1;1 and MtAMF1;3 highlight the conservation of putative ammonium regulators supporting both the rhizobial and AM fungal symbiosis in legumes.

The Metabolic Profile of Young, Watered Chickpea Plants Can Be Used as a Biomarker to Predict Seed Number under Terminal Drought.

Chickpea is the second-most-cultivated legume globally, with India and Australia being the two largest producers. In both of these locations, the crop is sown on residual summer soil moisture and left to grow on progressively depleting water content, finally maturing under terminal drought conditions. The metabolic profile of plants is commonly, correlatively associated with performance or stress responses, e.g., the accumulation of osmoprotective metabolites during cold stress. In animals and humans, metabolites are also prognostically used to predict the likelihood of an event (usually a disease) before it occurs, e.g., blood cholesterol and heart disease. We sought to discover metabolic biomarkers in chickpea that could be used to predict grain yield traits under terminal drought, from the leaf tissue of young, watered, healthy plants. The metabolic profile (GC-MS and enzyme assays) of field-grown chickpea leaves was analysed over two growing seasons, and then predictive modelling was applied to associate the most strongly correlated metabolites with the final seed number plant-1. Pinitol (negatively), sucrose (negatively) and GABA (positively) were significantly correlated with seed number in both years of study. The feature selection algorithm of the model selected a larger range of metabolites including carbohydrates, sugar alcohols and GABA. The correlation between the predicted seed number and actual seed number was R2 adj = 0.62, demonstrating that the metabolic profile could be used to predict a complex trait with a high degree of accuracy. A previously unknown association between D-pinitol and hundred-kernel weight was also discovered and may provide a single metabolic marker with which to predict large seeded chickpea varieties from new crosses. The use of metabolic biomarkers could be used by breeders to identify superior-performing genotypes before maturity is reached.

Balancing nitrate acquisition strategies in symbiotic legumes.

MAIN CONCLUSION: Legumes manage both symbiotic (indirect) and non-symbiotic (direct) nitrogen acquisition pathways. Understanding and optimising the direct pathway for nitrate uptake will support greater legume growth and seed yields. Legumes have multiple pathways to acquire reduced nitrogen to grow and set seed. Apart from the symbiotic N2-fixation pathway involving soil-borne rhizobia bacteria, the acquisition of nitrate and ammonia from the soil can also be an important secondary nitrogen source to meet plant N demand. The balance in N delivery between symbiotic N (indirect) and inorganic N uptake (direct) remains less clear over the growing cycle and with the type of legume under cultivation. In fertile, pH balanced agricultural soils, NO3- is often the predominant form of reduced N available to crop plants and will be a major contributor to whole plant N supply if provided at sufficient levels. The transport processes for NO3- uptake into legume root cells and its transport between root and shoot tissues involves both high and low-affinity transport systems called HATS and LATS, respectively. These proteins are regulated by external NO3- availability and by the N status of the cell. Other proteins also play a role in NO3- transport, including the voltage dependent chloride/nitrate channel family (CLC) and the S-type anion channels of the SLAC/SLAH family. CLC's are linked to NO3- transport across the tonoplast of vacuoles and the SLAC/SLAH's with NO3- efflux across the plasma membrane and out of the cell. An important step in managing the N requirements of a plant are the mechanisms involved in root N uptake and the subsequent cellular distribution within the plant. In this review, we will present the current knowledge of these proteins and what is understood on how they function in key model legumes (Lotus japonicus, Medicago truncatula and Glycine sp.). The review will examine their regulation and role in N signalling, discuss how post-translational modification affects NO3- transport in roots and aerial tissues and its translocation to vegetative tissues and storage/remobilization in reproductive tissues. Lastly, we will present how NO3-influences the autoregulation of nodulation and nitrogen fixation and its role in mitigating salt and other abiotic stresses.

The preceding root system drives the composition and function of the rhizosphere microbiome.

BACKGROUND: The soil environment is responsible for sustaining most terrestrial plant life, yet we know surprisingly little about the important functions carried out by diverse microbial communities in soil. Soil microbes that inhabit the channels of decaying root systems, the detritusphere, are likely to be essential for plant growth and health, as these channels are the preferred locations of new root growth. Understanding the microbial metagenome of the detritusphere, and how it responds to agricultural management such as crop rotations and soil tillage, is vital for improving global food production. RESULTS: This study establishes an in-depth soil microbial gene catalogue based on the living-decaying rhizosphere niches in a cropping soil. The detritusphere microbiome regulates the composition and function of the rhizosphere microbiome to a greater extent than plant type: rhizosphere microbiomes of wheat and chickpea were homogenous (65-87% similarity) in the presence of decaying root (DR) systems but were heterogeneous (3-24% similarity) where DR was disrupted by tillage. When the microbiomes of the rhizosphere and the detritusphere interact in the presence of DR, there is significant degradation of plant root exudates by the rhizosphere microbiome, and genes associated with membrane transporters, carbohydrate and amino acid metabolism are enriched. CONCLUSIONS: The study describes the diversity and functional capacity of a high-quality soil microbial metagenome. The results demonstrate the contribution of the detritusphere microbiome in determining the metagenome of developing root systems. Modifications in root microbial function through soil management can ultimately govern plant health, productivity and food security.

Effect of N supply on the carbon economy of barley when accounting for plant size.

Nitrogen availability and ontogeny both affect the relative growth rate (RGR) of plants. In this study of barley (Hordeum vulgare L.) we determined which growth parameters are affected by nitrate (N) availability, and whether these were confounded by differences in plant size, reflecting differences in growth. Plants were hydroponically grown on six different nitrate (N) concentrations for 28 days, and nine harvests were performed to assess the effect of N on growth parameters. Most growth parameters showed similar patterns of responses to N supply whether compared at common time points or common plant sizes. N had a significant effect on the biomass allocation: increasing N increased leaf mass ratio (LMR) and decreased root mass ratio (RMR). Specific leaf area (SLA) was not significantly affected by N. RGR increased with increasing N supply up to 1 mM, associated with increases in both LMR and net assimilation rate (NAR). Increases in N supply above 1 mM did not increase RGR as increases in LMR were offset by decreases in NAR. The high RGR at suboptimal N supply suggest a higher nitrogen use efficiency (biomass/N supply). The reasons for the homeostasis of growth under suboptimal N levels are discussed.

Tissue and nitrogen-linked expression profiles of ammonium and nitrate transporters in maize.

BACKGROUND: In order to grow, plants rely on soil nutrients which can vary both spatially and temporally depending on the environment, the soil type or the microbial activity. An essential nutrient is nitrogen, which is mainly accessible as nitrate and ammonium. Many studies have investigated transport genes for these ions in Arabidopsis thaliana and recently in crop species, including Maize, Rice and Barley. However, in most crop species, an understanding of the participants in nitrate and ammonium transport across the soil plant continuum remains undefined. RESULTS: We have mapped a non-exhaustive set of putative nitrate and ammonium transporters in maize. The selected transporters were defined based on previous studies comparing nitrate transport pathways conserved between Arabidopsis and Zea mays (Plett D et. al, PLOS ONE 5:e15289, 2010). We also selected genes from published studies (Gu R et. al, Plant and Cell Physiology, 54:1515-1524, 2013, Garnett T et. al, New Phytol 198:82-94, 2013, Garnett T et. al, Frontiers in Plant Sci 6, 2015, Dechorgnat J et. al, Front Plant Sci 9:531, 2018). To analyse these genes, the plants were grown in a semi-hydroponic system to carefully control nitrogen delivery and then harvested at both vegetative and reproductive stages. The expression patterns of 26 putative nitrogen transporters were then tested. Six putative genes were found not expressed in our conditions. Transcripts of 20 other genes were detected at both the vegetative and reproductive stages of maize development. We observed the expression of nitrogen transporters in all organs tested: roots, young leaves, old leaves, silks, cobs, tassels and husk leaves. We also followed the gene expression response to nitrogen starvation and resupply and uncovered mainly three expression patterns: (i) genes unresponsiveness to nitrogen supply; (ii) genes showing an increase of expression after nitrogen starvation; (iii) genes showing a decrease of expression after nitrogen starvation. CONCLUSIONS: These data allowed the mapping of putative nitrogen transporters in maize at both the vegetative and reproductive stages of development. No growth-dependent expression was seen in our conditions. We found that nitrogen transporter genes were expressed in all the organs tested and in many cases were regulated by the availability of nitrogen supplied to the plant. The gene expression patterns in relation to organ specificity and nitrogen availability denote a speciality of nitrate and ammonium transporter genes and their probable function depending on the plant organ and the environment.

Unraveling the Functional Role of NPF6 Transporters.

The nitrate transporter 1/peptide transporter (NPF) family represents a growing list of putative nitrate permeable transport proteins expressed within multiple cell types and tissues across a diverse range of plant species. Their designation as nitrate permeable and/or selective transporters is slowly being defined as more genes are characterized and their functional activities tested both in planta and in vitro. The most notable of the NPF family has been the Arabidopsis thaliana homolog, AtNPF6.3, previously known as AtNRT1.1 or CHL1. AtNPF6.3 has traditionally been characterized as a dual-affinity nitrate transporter contributing to root nitrate uptake in Arabidopsis. It has also been identified as a nitrate sensor which regulates the expression of high-affinity nitrate transport proteins NRT2s and lateral root development as a part of the primary nitrate response in plants. The sensor function of AtNPF6.3 has also been attributed to its auxin transport activity. Other homologs of AtNPF6.3 are now being described highlighting the variability in their functional capabilities (alternative substrates and kinetics) linking to structural aspects of the proteins. This review focusses on NPF6.3-like transport proteins and the knowledge that has been gained since their initial discovery over two decades ago. The review will investigate from a structural point of view how NPF6.3-like proteins may transport nitrate as well as other ions and what can be learned from structural uniqueness about predicted activities in plants.

Root Ideotype Influences Nitrogen Transport and Assimilation in Maize.

Maize (Zea mays, L.) yield is strongly influenced by external nitrogen inputs and their availability in the soil solution. Overuse of nitrogen-fertilizers can have detrimental ecological consequences through increased nitrogen pollution of water and the release of the potent greenhouse gas, nitrous oxide. To improve yield and overall nitrogen use efficiency (NUE), a deeper understanding of nitrogen uptake and utilization is required. This study examines the performance of two contrasting maize inbred lines, B73 and F44. F44 was selected in Florida on predominantly sandy acidic soils subject to nitrate leaching while B73 was selected in Iowa on rich mollisol soils. Transcriptional, enzymatic and nitrogen transport analytical tools were used to identify differences in their N absorption and utilization capabilities. Our results show that B73 and F44 differ significantly in their genetic, enzymatic, and biochemical root nitrogen transport and assimilatory pathways. The phenotypes show a strong genetic relationship linked to nitrogen form, where B73 showed a greater capacity for ammonium transport and assimilation whereas F44 preferred nitrate. The contrasting phenotypes are typified by differences in root system architecture (RSA) developed in the presence of both nitrate and ammonium. F44 crown roots were longer, had a higher surface area and volume with a greater lateral root number and density than B73. In contrast, B73 roots (primary, seminal, and crown) were more abundant but lacked the defining features of the F44 crown roots. An F1 hybrid between B73 and F44 mirrored the B73 nitrogen specificity and root architecture phenotypes, indicating complete dominance of the B73 inbred. This study highlights the important link between RSA and nitrogen management and why both variables need to be tested together when defining NUE improvements in any selection program.

Plant roots redesign the rhizosphere to alter the three-dimensional physical architecture and water dynamics.

The mechanisms controlling the genesis of rhizosheaths are not well understood, despite their importance in controlling the flux of nutrients and water from soil to root. Here, we examine the development of rhizosheaths from drought-tolerant and drought-sensitive chickpea varieties; focusing on the three-dimensional characterization of the pore volume (> 16 µm voxel spatial resolution) obtained from X-ray microtomography, along with the characterization of mucilage and root hairs, and water sorption. We observe that drought-tolerant plants generate a larger diameter root, and a greater and more porous mass of rhizosheath, which also has a significantly increased water sorptivity, as compared with bulk soil. Using lattice Boltzmann simulations of soil permeability, we find that the root activity of both cultivars creates an anisotropic structure in the rhizosphere, in that its ability to conduct water in the radial direction is significantly higher than in the axial direction, especially in the drought-tolerant cultivar. We suggest that significant differences in rhizosheath architectures are sourced not only by changes in structure of the volumes, but also from root mucilage, and further suggest that breeding for rhizosheath architectures and function may be a potential future avenue for better designing crops in a changing environment.

A Comparison of Petiole Hydraulics and Aquaporin Expression in an Anisohydric and Isohydric Cultivar of Grapevine in Response to Water-Stress Induced Cavitation.

We report physiological, anatomical and molecular differences in two economically important grapevine (Vitis vinifera L.) cultivars cv. Grenache (near-isohydric) and Chardonnay (anisohydric) in their response to water-stress induced cavitation. The aim of the study was to compare organ vulnerability (petiole and stem) to cavitation by measuring ultrasonic acoustic emissions (UAE) and percent loss of conductance of potted grapevines subject to the onset of water-stress. Leaf (ψL) and stem water potential (ψS), stomatal conductance (gs), transpiration (E), petiole hydraulics (KPet), and xylem diameter were also measured. Chardonnay displayed hydraulic segmentation based on UAE, with cavitation occurring at a less negative ψL in the petiole than in the stem. Vulnerability segmentation was not observed in Grenache, with both petioles and stems equally vulnerable to cavitation. Leaf water potential that induced 50% of maximum UAE was significantly different between petioles and stems in Chardonnay (ψ50Petiole = -1.14 and ψ50Stem = -2.24 MPa) but not in Grenache (ψ50Petiole = -0.73 and ψ50Stem = -0.78 MPa). Grenache stems appeared more susceptible to water-stress induced cavitation than Chardonnay stems. Grenache displayed (on average) a higher KPet likely due to the presence of larger xylem vessels. A close relationship between petiole hydraulic properties and vine water status was observed in Chardonnay but not in Grenache. Transcriptional analysis of aquaporins in the petioles and leaves (VvPIP1;1, VvPIP2;1, VvPIP2;2 VvPIP2;3, VvTIP1;1, and VvTIP2;1) showed differential regulation diurnally and in response to water-stress. VvPIP2;1 showed strong diurnal regulation in the petioles and leaves of both cultivars with expression highest predawn. Expression of VvPIP2;1 and VvPIP2;2 responded to ψL and ψS in both cultivars indicating the expression of these two genes are closely linked to vine water status. Expression of several aquaporin genes correlated with gas exchange measurements, however, these genes differed between cultivars. In summary, the data shows two contrasting responses in petiole hydraulics and aquaporin expression between the near-isohydric cultivar, Grenache and anisohydric cultivar, Chardonnay.

Maize NPF6 Proteins Are Homologs of Arabidopsis CHL1 That Are Selective for Both Nitrate and Chloride.

Nitrate uptake by plant cells requires both high- and low-affinity transport activities. Arabidopsis thaliana nitrate transporter 1/peptide transporter family (NPF) 6.3 is a dual-affinity plasma membrane transport protein that has both high- and low-affinity functions. At-NPF6.3 imports and senses nitrate and is regulated by phosphorylation at Thr-101 (T101). A detailed functional analysis of two maize (Zea mays) homologs of At-NPF6.3 (Zm-NPF6.6 and Zm-NPF6.4) showed that Zm-NPF6.6 was a pH-dependent nonbiphasic high-affinity nitrate-specific transport protein. By contrast, maize NPF6.4 was a low-affinity nitrate transporter with efflux activity. When supplied chloride, NPF6.4 switched to a high-affinity chloride selective transporter, while NPF6.6 had only a low-affinity chloride transport activity. Structural predictions identified a nitrate binding His (H362) in NPF6.6 but not in NPF6.4. Mutation of NPF6.4 Tyr-370 to His (Y370H) resulted in saturable high-affinity nitrate transport activity and nitrate selectivity. Loss of H362 in NPF6.6 (H362Y) eliminated both nitrate and chloride transport. Furthermore, alterations to Thr-104, a conserved phosphorylation site in NPF6.6, resulted in a similar high-affinity nitrate transport activity with increased Km, whereas equivalent changes in NPF6.4 (T106) disrupted high-affinity chloride transport activity. NPF6 proteins exhibit different substrate specificity in plants and regulate nitrate transport affinity/selectivity using a conserved His residue.

The response of mesophyll conductance to nitrogen and water availability differs between wheat genotypes.

Increased mesophyll conductance (gm) has been suggested as a target for selection for high productivity and high water-use efficiency in crop plants, and genotypic variability in gm has been reported in several important crop species. However, effective selection requires an understanding of how gm varies with growth conditions, to ensure that the ranking of genotypes is consistent across environments. We assessed the genotypic variability in gm and other leaf gas exchange traits, as well as growth and biomass allocation for six wheat genotypes under different water and nitrogen availabilities. The wheat genotypes differed in their response of gm to growth conditions, resulting in genotypic differences in the mesophyll limitation to photosynthesis and a significant increase in the mesophyll limitation to photosynthesis under drought. In this experiment, leaf intrinsic water-use efficiency was more closely related to stomatal conductance than to mesophyll conductance, and stomatal limitation to photosynthesis increased more in some genotypes than in others in response to drought. Screening for gm should be carried out under a range of growth conditions.

Nitrogen assimilation system in maize is regulated by developmental and tissue-specific mechanisms.

KEY MESSAGE: We found metabolites, enzyme activities and enzyme transcript abundances vary significantly across the maize lifecycle, but weak correlation exists between the three groups. We identified putative genes regulating nitrate assimilation. Progress in improving nitrogen (N) use efficiency (NUE) of crop plants has been hampered by the complexity of the N uptake and utilisation systems. To understand this complexity we measured the activities of seven enzymes and ten metabolites related to N metabolism in the leaf and root tissues of Gaspe Flint maize plants grown in 0.5 or 2.5 mM NO3 (-) throughout the lifecycle. The amino acids had remarkably similar profiles across the lifecycle except for transient responses, which only appeared in the leaves for aspartate or in the roots for asparagine, serine and glycine. The activities of the enzymes for N assimilation were also coordinated to a certain degree, most noticeably with a peak in root activity late in the lifecycle, but with wide variation in the activity levels over the course of development. We analysed the transcriptional data for gene sets encoding the measured enzymes and found that, unlike the enzyme activities, transcript levels of the corresponding genes did not exhibit the same coordination across the lifecycle and were only weakly correlated with the levels of various amino acids or individual enzyme activities. We identified gene sets which were correlated with the enzyme activity profiles, including seven genes located within previously known quantitative trait loci for enzyme activities and hypothesise that these genes are important for the regulation of enzyme activities. This work provides insights into the complexity of the N assimilation system throughout development and identifies candidate regulatory genes, which warrant further investigation in efforts to improve NUE in crop plants.

VAMP721a and VAMP721d are important for pectin dynamics and release of bacteria in soybean nodules.

In root nodules rhizobia enter host cells via infection threads. The release of bacteria to a host cell is possible from cell wall-free regions of the infection thread. We hypothesized that the VAMP721d and VAMP721e exocytotic pathway, identified before in Medicago truncatula, has a role in the local modification of cell wall during the release of rhizobia. To clarify the role of VAMP721d and VAMP721e we used Glycine max, a plant with a determinate type of nodule. The localization of the main polysaccharide compounds of primary cell walls was analysed in control vs nodules with partially silenced GmVAMP721d. The silencing of GmVAMP721d blocked the release of rhizobia. Instead of rhizobia-containing membrane compartments - symbiosomes - the infected cells contained big clusters of bacteria embedded in a matrix of methyl-esterified and de-methyl-esterified pectin. These clusters were surrounded by a membrane. We found that GmVAMP721d-positive vesicles were not transporting methyl-esterified pectin. We hypothesized that they may deliver the enzymes involved in pectin turnover. Subsequently, we found that GmVAMP721d is partly co-localized with pectate lyase. Therefore, the biological role of VAMP721d may be explained by its action in delivering pectin-modifying enzymes to the site of release.

Neglecting legumes has compromised human health and sustainable food production.

The United Nations declared 2016 as the International Year of Pulses (grain legumes) under the banner 'nutritious seeds for a sustainable future'. A second green revolution is required to ensure food and nutritional security in the face of global climate change. Grain legumes provide an unparalleled solution to this problem because of their inherent capacity for symbiotic atmospheric nitrogen fixation, which provides economically sustainable advantages for farming. In addition, a legume-rich diet has health benefits for humans and livestock alike. However, grain legumes form only a minor part of most current human diets, and legume crops are greatly under-used. Food security and soil fertility could be significantly improved by greater grain legume usage and increased improvement of a range of grain legumes. The current lack of coordinated focus on grain legumes has compromised human health, nutritional security and sustainable food production.

Maize maintains growth in response to decreased nitrate supply through a highly dynamic and developmental stage-specific transcriptional response.

Elucidation of the gene networks underlying the response to N supply and demand will facilitate the improvement of the N uptake efficiency of plants. We undertook a transcriptomic analysis of maize to identify genes responding to both a non-growth-limiting decrease in NO3- provision and to development-based N demand changes at seven representative points across the life cycle. Gene co-expression networks were derived by cluster analysis of the transcript profiles. The majority of NO3--responsive transcription occurred at 11 (D11), 18 (D18) and 29 (D29) days after emergence, with differential expression predominating in the root at D11 and D29 and in the leaf at D18. A cluster of 98 probe sets was identified, the expression pattern of which is similar to that of the high-affinity NO3- transporter (NRT2) genes across the life cycle. The cluster is enriched with genes encoding enzymes and proteins of lipid metabolism and transport, respectively. These are candidate genes for the response of maize to N supply and demand. Only a few patterns of differential gene expression were observed over the entire life cycle; however, the composition of the classes of the genes differentially regulated at individual time points was unique, suggesting tightly controlled regulation of NO3--responsive gene expression.