Exploiting the self-regulatory circuit of nitrate assimilation in plants for improved nitrogen use efficiency and crop sustainability.

The Food and Agriculture Organization of the United Nations projects that in the next 30-40 years the world will be one third more populous. This increase in the global population will put extraordinary pressure on agricultural systems. A concurrent boost in agricultural production will be required to meet already overburdened food, fiber and fuel demands. However, for major cropping systems, the actual average yield range between 20% and 80% of potential. Thus, the development of new strategies to reduce the yield gap is of great economic and social importance. Currently, crop productivity relies heavily on the use of commercial fertilizers, with attributable yield percentage reaching values as high as 90%. Particularly, supply of the inorganic ion nitrate, the primary source of nitrogen for land plants, represents a major bottleneck in crop yield. Due to its high mobility in water, nitrate ions are often runoff from the soil, eventually leading to environmental impact. Thus, current fertilization strategies often offer limited efficacy and are potentially hazardous to human health and the environment, while still leading to economic losses of billions every year. Conversely, genetic and biochemical improvement of plant primary metabolism represents a safe and sustainable alternative to increasing crop yield while making more efficient use of natural resources.
To cope with fluctuations in its availability in time and space, plants have evolved the ability to modulate nitrate acquisition according to their N status and nitrate concentration in soil. Plants actively transport nitrate across the plasma membrane of the roots through the sophisticated transport systems. Under limiting availability, nitrate acquisition relies on high-affinity transporters, which recruitment and activity in response to nitrate is mediated by post-translational modifications. Once taken up by roots, nitrate is mainly transported to shoots for further incorporation of N atoms into carbon skeleton through sequential assimilatory reactions to form N-containing organic molecules, such as amino acids, proteins and nucleotides. As one of the most energy-consuming biochemical pathways in nature, nitrate assimilation is tightly controlled to ensure proper plant development and growth.
Several lines of evidence indicate that flux in nitrate assimilation pathway is associated with production of reactive nitrogen species. Particularly, we have recently shown that the redox active molecule nitric oxide (NO), one of the end products of nitrogen metabolism, feedback regulates flux through nitrate assimilation pathway. NO bioactivity is mediated mainly through the protein post-translational modification S-nitrosylation, i.e. the covalent attachment of a NO moiety form protein-SNO. Our findings revealed that intracellular protein-SNO accumulation is associated with reduced expression of the nitrate transporters and inhibition of assimilatory reactions. Thus, a feedback loop mechanism associated with nitrate assimilation limits nutrient assimilation in plants. Remarkably, genetic manipulation of protein-SNO levels markedly impacted plant vigour, suggesting that this feedback mechanism can be harnessed to improve plant productivity.
It remains unclear, however, the identity of the redox-responsive nodes in nitrogen assimilation pathway and how they operate to control plant fitness. Here I propose to use a innovative, genetic, genomic and inter-disciplinary imaging techniques to identify and synthetically manipulate metabolic nodes that feedback nitrate assimilation in plants. Moreover, the proposed genetic and biochemical management of NO-mediated redox signalling has the potential to ultimately reveal novel chemical and genetic targets that can be used in future crop improvement strategies.

Continued rapid growth of the global population is raising concerns over future food security. With agricultural lands in decline an attractive way of increasing crop yield per unit of land is to improve nutrient use efficiency and prevent significant economical losses and environmental impact caused by the excessive use of fertilizers. The inorganic ion nitrate is the primary source of nitrogen for land plants, and the availability of this nutrient in the soil represents a bottleneck in crop yield. To assimilate nitrate, plants employ a variety of transporters (e.g. NRT1.1/NPF6.3 and NRT2.1) and reductases (e.g. Nitrate reductase) expressed in different tissues and organs to transport and catalyse the sequential reduction of assimilates. Importantly, assimilation of nitrate is linked to generation of the redox signal nitric oxide (NO). NO regulates plant development and stress responses through S-nitrosylation, i.e. the covalent attachment of NO to cysteines residues to form protein-SNO. Recently, we showed that protein-SNO signalling suppresses both nitrate uptake and reduction by transporters and reductases, respectively, to fine-tune nitrogen homeostasis. It remains unclear, however, the identity of the redox-responsive nodes in nitrate assimilation pathway and how they operate to control plant fitness. Here I propose to use innovative, genetic, genomic and inter-disciplinary imaging techniques to identify and synthetically manipulate targets of redox regulation that feedback nitrate assimilation. Moreover, the proposed management of NO-mediated redox signalling by emerging denitrosylases (GSNOR1 and TRX-h5) has the potential to ultimately reveal novel chemical and genetic targets that can be harnessed in future crop improvement strategies.

To support maximum yield, current crop management relies heavily on the production and application of nitrogen (N)-containing fertilizers. Due to the massive energy input required and fertilizer runoff, this strategy is associated with significant environmental impact and economic losses. By increasing our understanding at the molecular level of plant nutrient uptake and assimilation, we may ultimately be able to genetically improve nitrogen use efficiency (NUE) and breed high-NUE cultivars for increased crop yield.
With nearly 5 million hectares of agricultural land, the UK accounts for almost 10% of total EU crop production worth billions of pounds. Each year tons of fertilizers are applied in the UK as an attempt to boost crop productivity, driving commodities market price whilst leading to significant environmental impact. Genetic and biochemical improvement of NUE represents a safe and environmentally sustainable alternative to improve crop yield. Preventing current and future losses in the UK by the synthetically management of crop NUE will increase economic competitiveness. Nevertheless, the UK imports nearly half of its vegetable crops and about 90 percent of fresh fruits. Thus, improvement of crop performance will be required to meet food security with global increasing demand.
Outcomes from the proposed research may substantiate genetic-based, translational works to improve crop yield. Breeding of cultivars with economic interest often relies on the development of genetically modified organisms (GMO). GMO have been on the agenda for extensive discussions by the wider public and policy makers. The research program and development strategy proposed here will promote engagement of a variety of sectors of society in science divulgation to address shared interests.
Redox-mediated control of metabolism reprogramming is a conspicuous feature found throughout the Eukarya domain. For instance, the fundamental and comprehensive understanding of how specificity in redox-mediated signalling is achieved is a prerequisite to the development of strategies to address diseases that are caused by unsuitable redox signalling, such as oncogenesis and Parkinson disease. In addition to the improvement of crop productivity, the outcomes of this research programme will bring novel advances in cellular and molecular biochemistry and open new avenues in applied biotechnology.