Efforts to end hunger are as old as human civilization, and they have affected the history of humanity. The concern for global food security results primarily from saturated crop yields and also an imbalance between the supply and demand of the major food crops (wheat, rice, and maize) (DeFries et al., 2015; Wood et al., 2018). Increasing production on a sustained basis is an essential component of ensuring food security. Food production needs to increase 50% by 2030 and double by 2050 to meet projected demands (Pastor et al., 2019). Effective nutrient use especially nitrogen (N) and phosphorous (P) are important for sustained food security. A reliable supply of N and other nutrients vital for plant growth has allowed farmers to improve productivity over the past century, thus promoting economic development (Drechsel et al., 2015; McLaughlin and Kinzelbach, 2015; Reis et al., 2016).

Nitrogen (N) plays a prominent role in the plant metabolic system (Shah et al., 2021a,b). Nitrogen, is a principal component of many organic compounds (protein, nucleic acids, alkaloids enzyme) and is also associated with energy transfer molecules like adenosine diphosphate (ADP) and adenosine triphosphate (ATP) (ATP). It consists the 16% of the total protein biomass present in plants. N is also found in nucleic acids (deoxyribonucleic acid and ribonucleic acid), which play an important role in plant genetics and heredity. It is also a component of chlorophyll, which serves as a photosynthesis factory. N also has a major role in photosynthetic processes, leaf area production, leaf area duration as well as net assimilation rate that is directly related to yield enhancement (Leghari et al., 2016). More than half of the world’s population is fed by crops cultivated using synthetic nitrogen (N) fertilizers, which were made feasible by the advent of the Haber-Bosch process in the early twentieth century, which converts atmospheric nitrogen gas (N₂) to active forms of nitrogen. Total global consumption of N fertilizer was 112.5 million tons in 2015, 118.2 million tons in 2019, and likely to reach 7.9–10.5 billion by 2050 (Zhang X. et al., 2015). The demand for huge amounts of N fertilizers is a major concern for farmers as it increases the cost of cultivation and also degrades the inherent fertility of the soil. Consequently, the use of excess N fertilizers causes the problem of N pollution, which is now considered a new threat to environmental sustainability (Kanter et al., 2020).

Nitrogen use efficiency (NUE) is defined as the yield of grain achieved per unit of N available to the crop from soil and applied fertilizer, and it can be divided into two biological components, N uptake efficiency (NUpE) which is the efficiency of absorption/uptake of supplied N, and N utilization efficiency (NUtE) which is the efficiency of assimilation and remobilization of plant N to ultimately produce grain (Han et al., 2015; Yadav et al., 2017; Congreves et al., 2021). It has been documented that the NUE value is decreased with the application of N fertilizers. For instance, in developed countries, NUE increased because of the adaption of best agronomical and fertilizer management practices, while it reduced in developing countries like India as the use of N fertilizer dramatically increased (Heffer and Prud’homme, 2016). It was reported that the world average NUE was nearly 47 and 42% in 2009 and 2010, respectively, and the major reason behind the NUE decreasing is the adoption of agronomical practices (nutrient management and varieties) associated with low NUE (Zhang X. et al., 2015). Therefore, this study primarily focused on the advanced agronomical and molecular strategies, which reduce the N fertilizer demand and enhance NUE.

Several morphological and physiological traits such as root length, lateral roots, root architecture, light capture photosynthesis, canopy height, flowering time, carbohydrate partitioning, storage, and the remobilization of N, (stem remobilization, leaf blade remobilization, and total remobilization) are linked with higher NUE in plants (Mosleth et al., 2015; Carmo-Silva et al., 2017; Guttieri et al., 2017; Tabbita et al., 2017; Hawkesford and Griffiths, 2019). Improving nitrogen use efficiency is essential for tackling the triple threat of environmental degradation, climate change, and food security. The efficiency of applied nitrogenous fertilizers is very low due to their various losses i.e., volatilization, leaching, surface runoff, and denitrification from the soil-plant system (Zhang X. et al., 2015). Nitrogen use efficiency (NUE) is a convoluted phenomenon governed by various edaphic, climatic, and management factors; it has to be managed through an amalgamated approach involving agronomical, physiological, and molecular aspects. Among these, the agronomical approach is economically feasible, ecologically viable, and sustainable. The modern fertilizers (customized fertilizer, nano fertilizers, bio-fertilizers, biochar, and zeolite) and deep fertigation helps in improving NUE in sustainable ways. Likely, the precision farming (Leaf color chart, chlorophyll meter, and remote sensing) and modern agronomical practices (Diversified crop rotation, 4R nutrient stewardship, and crop modeling) have been developed for enhancing NUE and can support good plant performance and better crop output. New technological developments need to achieve further gains by new varieties, slow-release fertilizers, nitrification and urease inhibitors, fertigation, and high-tech approaches to precision agriculture (Pires et al., 2015; Cao et al., 2017; Mylonas et al., 2020). The identification and employment of QTLs/genes/transporters which are responsible for NUE at the molecular level is now a more feasible and robust technique with the availability of efficient molecular tools (Ali et al., 2018). Previously, Zhang X. et al. (2021) employed the RNA sequencing (RNA-seq) technique to investigate the genotypic difference in response to N deficiency between two wheat NILs (1Y, high-NUE, and 1W, low-NUE). The high- and low-NUE wheat NILs showed different patterns of gene expression under N-deficient conditions, and these N-responsive genes were classified into two major classes, including “frontloaded genes” and “relatively upregulated genes.” In total, 103 and 45 genes were identified as frontloaded genes in high-NUE and low-NUE wheat, respectively. Moreover, the expression of TaPT4, TaNRT2.2, and TaAMT1.2 was down-regulated by arbuscular mycorrhizal (AM) colonization only when roots of host plants received phosphate or nitrogen nutrient signals. However, the expression of TaPHT1.2, TaNRT2.1, and TaNRT2.3 was down-regulated by AM colonization, regardless of whether there was nutrient transfer from AM hyphae. The expression of TaNRT1.2 was also down-regulated by AM colonization even when there was no nutrient transfer from AM hyphae (Zhang X. et al., 2021). Nitrogen limitation adaptation (NLA) was involved in source-to-sink remobilization of nitrate by mediating the degradation of NRT1.7 in Arabidopsis (Liu et al., 2017). In the last decade, numerous studies suggested that the use of advanced soil and agronomical practices with modern molecular techniques could be the best way to enhance NUE swiftly under future climate change. Therefore, the objective of the present study is to discuss the role of advancement in agronomy, breeding, and molecular biology to enhance NUE and this information might be helpful in reducing N pollution.

In healthy plants, nitrogen concentrations range from 3 to 4%. When compared to other nutrients, this is a substantially higher amount. In plant cells, nitrogen is a significant source of various morphological, genetic, and metabolic components. The biological combination of nitrogen with C, H, O, and S to form amino acids, the building blocks of proteins, makes nitrogen a key component of protein (Okumoto and Pilot, 2011). Protoplasm, the site of cellular division and consequently of growth and development of plants, is formed using amino acids. The N increases the leaf area (AF) and leaf area index (LAI), promoting the synthesis of proteins involved in cell development, cell proliferation, and the development of the cell wall and cytoskeleton (Luo et al., 2020; Sun et al., 2020). Plant development and grain output are fueled by nitrogen’s role in the chlorophyll molecule, which allows the plant to absorb sunlight energy through photosynthesis (Evans and Clarke, 2019). Compounds that transport energy, such as ATP (adenosine triphosphate), include nitrogen. Adenosine triphosphate (ATP) helps cells store and utilize the energy supplied during metabolism (Evans and Clarke, 2019). As an osmotic agent N also plays an important role in water retention in plant vacuoles found to be essential toward its nutrition function. Water is the key limiting element in plant development since it is the main factor that can regulate cell growth and metabolism (Ding et al., 2018). Nitrogen raises the protein concentration of forage crops, improves the quality of the fruit, and speeds up the growth of green vegetables. Potassium and phosphorous are absorbed and utilized more efficiently by plants that are fed this nutrient (Chand et al., 2022). Nitrogen is essential for plants’ healthy growth and development. Nitrogen deficiency greatly impacts crop productivity, whereas extra N might have detrimental consequences on the plant. This topic is always being discussed in crop production. Therefore, this concern is constantly being addressed in agricultural output (Mu and Chen, 2021).

As of today, the primary strategy for preserving and increasing agriculture production is the administration of mineral fertilizers, such as nitrogen. Commercial fertilizers contain N which is highly soluble, making it easy for plants to absorb and assimilate. Only 30–40% of the applied nitrogen is utilized by crops since the N compounds are often available in the form of nitrate and ammonium and are highly mobile in the soil (Omara et al., 2019). NUE can be described in a variety of approaches, but the most fundamental is the yield (grain, fruit, or forage) per unit of nitrogen present in the soil. The utilization of nitrogen by plants occurs in two distinct stages (Dobermann, 2005). In the first step, the quantity of N taken in, stored, and converted into amino acids and other essential nitrogenous molecules is a factor in the production of biomass. The eventual yield is determined by the amount of nitrogen (N) given to the seed at the second phase. Plants use the nitrate acquisition pathway to take in and synthesize inorganic nitrogen and store amino acids in immature leaves and roots. Their use in protein and enzyme biosynthesis, as well as the photosynthetic machinery that controls plant growth, architecture, and development, is extensive. For optimal flowering and grain development, nitrogenous molecules must be supplemented during the reproductive stage. Nitrogen absorption and remobilization are crucial, with leaves and shoots serving as a source of amino acids for the reproductive and storage organs (Chen et al., 2020). As a result, improving NUE necessitates an understanding of the processes by which N is taken up, assimilated, and remobilized throughout the plant life cycle. But over 60% of soil nitrogen is lost via leaching, runoff, denitrification, evaporation, and microorganism utilization (Ding et al., 2021). The intricacy of NUE lies in the multiple N sources that contribute to crop production, including soil N availability, conversion, accumulation, transport, and depletion, microclimatic conditions, crop genetics, and the effect of administration, environment, and climate (Chen et al., 2020). This leads to N losses in diverse types to the soil, air, and water, which is both economically and environmentally problematic.

Nitrate is a key source of nitrogen for plants; in fact, the majority of plants spend a substantial percentage of their carbon and energy reserves on its uptake and assimilation (Kant, 2018). Nitrate is a nutrient and a signaling molecule that has a dramatic influence on plant metabolism and development. Plants have evolved complex methods for detecting nitrate and integrating its absorption into their transport systems (Vega et al., 2019). Nitrate assimilation begins with its absorption into the cell. Normally, nitrate is taken up from the soil solution through the root’s apopalsm and then absorbed into the epidermal and cortical cells. Once inside the symplast, it is reduced or mobilized into the xylem for transport to the shoots through the Casparian strip (Islam, 2021).

Plants utilize both high affinity and low-affinity transporters to import nitrate. These import pathways allow plants to accommodate a wide range of external nitrate concentration without experiencing severe deficiency or toxicity. A wide array of genes are associated with the uptake of nitrogen in plants. The nitrate uptake by plant roots appears to be a complex process involving at least four distinct transport systems: (a) constitutive high-affinity transporters (cHATS); (b) nitrate-inducible high-affinity transporters (iHATS); (c) constitutive low-affinity transporters (cLATS); and (d) nitrate-inducible low-affinity transporters (iLATS) (O’Brien et al., 2016). In plants, two families of genes producing nitrate transporters have been identified: NRT1 and NRT2. The NRT 2 family encodes transporters to the high-affinity uptake system whereas NRT 1 family is more complex, including nitrate transporters with dual affinity or low affinity. The nitrate uptake is driven by the proton gradient against the plasma membrane and its uptake is driven by ATPases (Wang et al., 2020; You et al., 2022). In comparison to nitrate, plants generally do not collect excessive amounts of ammonium ions. Toxicity symptoms usually occur when agricultural plants are cultivated in ammonium deficient in nitrate (Duan et al., 2018). Initially, a family of five ammonium transporter genes named AMT1;1–AMT1;5 was found in Saccharomyces cerevisiae and Escherichia coli (Loqué and von Wirén, 2004). However, the scenario is somewhat different in rice, where 10 distinct genes have been found (Suenaga et al., 2003). Attempts have been made further to identify transporters of nitrogen uptake in plants and overexpression of nitrogen transporters genes will lead to enhanced nitrogen uptake efficiency of the roots from the soil. Some of the nitrogen and ammonium transporters and their function are highlighted in Figure 1.

Nitrogen is the major essential nutrient required by the plants and it interacts with almost all the other essential nutrients. N has synergistic interaction with most of the nutrients that leads to enhanced NUE. However, the kind of interaction depends on the number and amount of interacting nutrients present in the soil and their application rate. If both the interacting nutrients have an imbalanced amount in the soil it could cause antagonistic interaction (Datta and Meena, 2015). Jiang J. et al. (2019) reported that P x N have positive interaction (synergistic) because the application of P improved the N content in the plant. Dwivedi et al. (2003) also observed that combined fertilization of N and P improved the NUE by 28% in wheat crop. Conversely, Aulakh and Malhi (2005) observed that in P deficient soil, sole application of N led to a reduction in grain yield and reported antagonistic interaction. Barker and Pilbeam (2015) found that fertilization of K enhanced the N uptake and assimilation in plants, and eventually NUE, by many folds. However, they also reported that when K is low in the soil, it competes with NH4 for selective binding sites in the adsorption process and eventually caused an antagonistic effect. Sulfur is the most important secondary nutrient, and Jamal et al. (2010) revealed that S application has an instrumental role in enhancing the recovery efficiency and NUE of applied N. However, Rietra et al. (2017) reported that in S deficiency condition, application of N leads to abundant accumulation of a pernicious level of N metabolites in plants. Wilkinson et al. (2000) concluded that Ca and Mg enhanced solubility of fixed N in the acidic soils, and improved N translocation in plant. However, in Mg deficient soil, sole application of N fertilizers causes grass tetany in livestock caused by low Mg concentration in forage. N fertilization improved the Zn absorption, translocation, and assimilation in crops and led to increased zinc use efficiency and vice versa. Antagonistic effect of N fertilization on Zn nutrition either due to dilution effect (decrease or dilution in plant nutrient concentration due to increase in biomass yield) effect or poor translocation of Zn-protein complex in the roots. The application of N fertilizers enhanced the availability of Fe and Mn by increasing acidity which further led to the conversion of unavailable forms of Mn and Fe into available forms (Aulakh and Malhi, 2005). In optimum conditions, N has a positive interaction with Cu but in the case of Cu deficient soil application of N resulted in crystal clear symptoms of Cu deficiency in plants (Barker and Pilbeam, 2015). The interaction effect of B, Cl, Mo, and Ni with N has not been reported so far. However, some studies reported that these nutrients are helpful in increasing NUE. In acidic conditions Al and Ce reduced the uptake and assimilation of N due to reduced growth of roots ultimately decreased NUE (Hille et al., 2011; Zhao and Shen, 2020). The cross-talks of different mineral elements with N are highlighted in Table 1.

Physiological attributes play a pivotal role in improving NUE in plant species. Among those attributes, the root system is crucial in the acquisition of water and nutrients through its ability of soil exploration which is the main determinant of NUE (Li X. et al., 2016). NUE depends upon the root distribution in soil, root size, and root/shoot (R/S) ratio (Garnett et al., 2009). These factors maximize the uptake and interception of nitrogen as well as reduce the nitrogen losses to groundwater and deeper layers of soil (Lynch, 2013). The important plant attributes related to N uptake, acquisition, assimilation, remobilization, and portioning are highlighted in Figure 3. Various research studies have been conducted to show that root system architect (RSA) is closely related to the NUE e.g., plants having deeper and steeper roots can uptake nitrogen effectively from deeper soils (Zhan and Lynch, 2015). A research study on two varieties of Chinese (XY335 and ZD958) and one variety of US (P32D79) maize was conducted to analyze the effect of the root system on NUE. Root analysis with respect to NUE revealed the positive correlation of NUE with dry weight (RDW) and root/shoot (R/S) biomass ratio. While RDW and R/S ratio of western variety (P32D79) were greater than Chinese varieties (XY335 and ZD958) similarly, it was also observed that P32D79 had a better root system which leads to higher N-uptake and removal of more N-minerals than Chinese varieties from the soil. Thus, it was concluded that maize variety with a better root system and higher stress resistance underlines higher NUE (Yu et al., 2015).

Due to the major role of the root system in NUE, its genetic basis is also subjected to study. A research study was conducted to understand the genetic association between root traits of maize seedlings and NUE through QTL analysis. Recombinant inbred lines of maize were sown under low and high nitrogen conditions in which 9 traits of RSA and 10 traits of NUE were evaluated in three hydroponic and four field conditions, respectively. A significant correlation was observed between uptake efficiency of nitrogen and RSA (especially in the crown and seminal roots). Almost 331 QTLs in total were identified including 147 QTLs of RSA and 184 QTLs of NUE. Almost 70% of NUE-QTLs coincided with the QTL clusters of RSA which indicated the significant genetic association between traits of NUE and RSA. This association can aid in the selection of RSA-traits for improvement of NUE in the maize (Li et al., 2015).

In a research study, recombinant inbred rapeseed “BnaZNRIL” was studied to investigate traits related to NUE and root morphology (RM) under high and low nitrogen hydroponic conditions. Results indicated the significant correlation of root size (trait related to RM) with N-uptake and dry biomass of plant while no correlation was observed with the efficiency of nitrogen utilization (NUtE). Approximately 23 stable QTLs out of a total of 129 were detected in both high and low nitrogen conditions. Most of the stable QTLs (20/23) were observed to be related to RM traits under both high and low nitrogen conditions which lead to the suggestion of regulating the root morphology through NUE is more feasible than through regulating the nitrogen efficiency in rapeseed (Wang et al., 2017). Another genome-wide association study was conducted to understand the effect of low nitrogen levels on root growth and root traits at seedling emergence level. A panel of 461 inbred lines of maize was assayed for the root growth under low nitrogen (0.05 mml^–1) and high nitrogen level (5 mml^–1). The panel was further genotyped with an SNP marker of 542,796 high density. Various root traits were observed to be increased under low nitrogen conditions such as total root length, dry weight of root, root-to-shoot ratio, lateral and axial root length of the primary root. Furthermore, a wider heritability range of 0.43–0.82 was observed under low nitrogen conditions than the range of 0.25–0.55 under high nitrogen conditions. After marker-assisted analysis, it was observed that under low nitrogen conditions, gene encoding DELLA protein was in association with the lateral root-zone of primary root and protoporphyrinogen IX oxidase-2 gene was in association with the surface area of the root. While under high nitrogen conditions the histone-deacetylase gene was in association with the plant height which can be further used to exploit genetic improvement in root traits through better NUE of maize (Sun et al., 2021). The important root traits and associated genes in response to NUE are highlighted in Table 2.

Cereals, such as rice, wheat, and maize, are the most important sources of calories and nutrition for humans. Modern varieties differ in key NUE characteristics (Cormier et al., 2013; Barraclough et al., 2014). However, it is clear that a broader germplasm base has a much higher potential for variation (Monostori et al., 2017). The efficient use of resources, including fertilizers such as nitrogen, is critical to long-term sustainability. According to Raun and Johnson (1999), globally only 33% of applied nitrogen fertilizer is recovered in harvested grain. The main problem with landraces and relatives is that while biomass may be high, yields and in particular HI are normally low, making traditional NUE measurements less useful. Height, which is influenced by dwarfing genes, is the most important architectural influence. While Rht genes may have a negative effect on height, they may also have other pleiotropic effects, such as decreased root proliferation (Gooding et al., 2012; Bai et al., 2013). The efficiency of absorption will be aided by variations in root architecture and function. Root traits have been deconstructed as quantitative trait loci (QTL) in a number of studies. The variation in root proliferation, length, lateral profusion, and spread or angle of roots has been identified (Gooding et al., 2012; Atkinson et al., 2015). Wheat has a large number of nitrate transporter genes that are involved in both initial uptake and internal translocation mechanisms.

NUE is a polygenic trait with a number of factors. As a result, identifying individual genetic effects involves quantitative genetics strategies, and such effects have been usually characterized as QTL. There are several methods for identifying genes that control NUE and utilizing them in breeding. Associations genetics could be based on genetic panels that are well-suited to the researcher’s test environment, sample multiple alleles, and provide extremely high genetic resolution. Furthermore, association genetics requires a good balance of alleles at each location analyzed, with low-frequency alleles (usually less than 10%) being excluded from the research. Segregating populations derived from two diverse parents is the best-established way of QTL identification. Recombinant Inbred Lines and Single Seed Descent or the doubled haploids are statistically strong because only the two parental alleles are segregating at anyone locus and the population is comprised 50% of each allelic class.

A number of studies on NUE QTLs have been well known by this approach for example in rice (Wei et al., 2011). Multi Parent Advanced Generation Intercross (MAGIC) permits the concurrent analysis of several alleles and the mapping resolution afforded by recombination of the population as well as historical recombination (Mackay et al., 2014). Wingen et al. (2017) produced a publicly available NAM population that represents more than 90 landrace parents and over 10,000 recombinant inbred lines. Preceding studies of cereal crops have searched for novel NUE traits and alleles in adapted breeding materials (Fontaine et al., 2009), landraces (Pozzo et al., 2018; Van Deynze et al., 2018), and wheat wild relatives (Hu et al., 2015). While these authors have successfully identified QTLs, genes, and genotypes conferring high NUE, additional sources of genetic variation likely still exist within the currently unexplored germplasm.

Recently the development of cultivars with improved NUE is of utmost essential as after decades of green revolution application of N fertilizer is increased tremendously to meet grain yield demands. Simultaneously, its negative impact may be witnessed in the form of the increased cost of cultivation and hazardous effect on the ecosystem. The transcription factors, allosteric control, and post-transcriptional modification all play a vital role in the expression of a complex trait like NUE (Basu and Jenkins, 2020). Because quantitative traits like NUE are influenced by a large number of genes with minor effects and environmental influence, identifying QTLs for such traits necessitates a larger mapping population with phenotyping at a variety of locations and environmental conditions, as well as a sufficiently large coverage of the genome by the markers. Though, major QTLs with major and minor effects that can easily be incorporated in the breeding cycle are very limited in number. However, minor QTLs with minor effects are more in number but they require proper validation across the population. The molecular information on genes involved in various stages of the N metabolic process, from protein to final metabolites can aid in the identification of QTLs associated with NUE or its components. The important molecular biology tools for improving NUE are highlighted in Figure 4.

Global food security with nutritional security is the most crucial task for future generations. However, the efforts of scientists or agricultural researchers can achieve this task and somehow they succeed in finding numerous technologies and ways through which they are trying to complete. Although, in modern times several new problems such as climate change, abiotic stresses, and problematic soils cause the loss of natural resources, and together amplify our food security target. In this scenario, there is an emergency need to reduce the cost of cultivation and degradation of natural resources. Therefore, increasing the NUE in economically important crops is a great challenge to secure environmental sustainability. However, the advancement in agronomical approaches has achieved some milestones in the last decades such as precision farming and nano-fertilizers. On the other hand, recent advancements in molecular biology and tools speed up the process and have a bright future in developing higher NUE efficient crops. However, still there is a huge gap in targeted NUE and also reduced during the last decades because of higher use of N fertilizers, especially urea. Therefore, it is recommended that the new fertilizer policy is based on the alternatives to urea for N sources such as customized, bio, and nano-fertilizers, which might be a breakthrough in enhancing NUE and reducing N pollution. Also, there might be a decision support system for best management practices at a local basis, which can directly benefit to the farmer in enhancing NUE and yield. Therefore, we combined these two crucial approaches in respect to improving NUE and might be game-changing for enhancing NUE in near future. Moreover, future trends and expectations should be aimed at cracking the current main hurdles to crop plants. Modern genome-editing tools can provide a permanent solution by developing varieties with enhanced NUE, although further studies are needed to reach these goals. The integration of all these approaches will lead to the sustainable production of crops, through the effective management of environmental stresses under the present scenario of changing climate. In addition, plant epigenetics, which is a conserved gene expression regulatory mechanism including histone modification, DNA methylation, non-coding RNA, and chromatin remodeling, represents an emerging and efficient tool to better understand biological processes in response to nitrogen enhancement.

TJ and RKS: conceptualization. II, TJ, RKS, RS, and AS: writing—original draft preparation. PK, AS, DJ, MAS, PD, DSa, HA, RA, and DSi: writing—review and editing. DSi and RA: funding acquisition. All authors have read and agreed to the published version of the manuscript.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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