The ability of plants to compete effectively for nitrogen (N) resources is critical to plant survival. However, controversy surrounds the importance of organic and inorganic sources of N in plant nutrition because of our poor ability to visualize and understand processes happening at the root-microbial-soil interface. Using high-resolution nano-scale secondary ion mass spectrometry stable isotope imaging (NanoSIMS-SII), we quantified the fate of ¹⁵N over both space and time within the rhizosphere. We pulse-labelled the soil surrounding wheat (Triticum aestivum) roots with either ¹⁵NH₄⁺ or ¹⁵N-glutamate and traced the movement of ¹⁵N over 24 h. Imaging revealed that glutamate was rapidly depleted from the rhizosphere and that most ¹⁵N was captured by rhizobacteria, leading to very high ¹⁵N microbial enrichment. After microbial capture, approximately half of the ¹⁵N-glutamate was rapidly mineralized, leading to the excretion of NH₄⁺, which became available for plant capture. Roots proved to be poor competitors for ¹⁵N-glutamate and took up N mainly as ¹⁵NH₄⁺. Spatial mapping of ¹⁵N revealed differential patterns of ¹⁵N uptake within bacteria and the rapid uptake and redistribution of ¹⁵N within roots. In conclusion, we demonstrate the rapid cycling and transformation of N at the soil-root interface and that wheat capture of organic N is low in comparison to inorganic N under the conditions tested.© 2013 The Authors. New Phytologist © 2013 New Phytologist Trust.

Oilseed rape (Brassica napus L.) is commonly grown for oil or bio-fuel production, while the seed residues can be used for animal feed. It can also be grown as a catch crop because of its efficiency in extracting mineral N from the soil profile. However, the N harvest index is usually low, due in part to a low ability to remobilize N from leaves and to the fall of N-rich leaves which allows a significant amount of N to return to the environment. In order to understand how N filling of pods occurs, experiments were undertaken to quantify N flows within the plant by (15)N labelling and to follow the changes in soluble protein profiles of tissues presumed to store and subsequently to remobilize N. Whereas N uptake increased as a function of growth, N uptake capacity decreased at flowering to a non-significant level during pod filling. However, large amounts of endogenous N were transferred from the leaves to the stems and to taproots which acted as a buffering storage compartment later used to supply the reproductive tissue. About 15% of the total N cycling through the plant were lost through leaf fall and 48%, nearly all of which had been remobilized from vegetative tissues, were finally recovered in the mature pods. SDS-PAGE analysis revealed that large amounts of a 23 kDa polypeptide accumulated in the taproots during flowering and was later fully hydrolysed. Its putative function of storage protein is further supported by the fact that when plants were grown at lower temperature, both flowering, its accumulation and further mobilization were delayed. The overall results are discussed in relation to plant strategies which optimize N cycling to reproductive sinks by means of buffering vegetative tissues such as stems and taproots.

Wheat leaves (Triticum aestivum L.) at the moment of their maximum expansion were detached and put in darkness. Their protein, RNA and DNA contents, as well as their rates of protein synthesis and degradation, were measured at different times from 0 to 5 days after detachment. Rates of protein synthesis were measured by incorporation into proteins of large amounts of [(3)H]leucine. Fractional rates of protein degradation were estimated either from the difference between the rates of synthesis and the net protein change or by the disappearance of radioactivity from proteins previously labeled with [(3)H]leucine or [(14)C]proline.Protein loss reached a value of 20% during the first 48 hours of the process. RNA loss paralleled that of protein, whereas DNA content proved to be almost constant during the first 3 days and decreased dramatically thereafter.Measurements of protein synthesis and degradation indicate that, in spite of a slowdown in rate of protein synthesis, an increased rate of protein breakdown is mainly responsible for the observed rapid protein loss.

Senescence is a highly organized and well-regulated process. As much as 75% of total cellular nitrogen may be located in mesophyll chloroplasts of C(3)-plants. Proteolysis of chloroplast proteins begins in an early phase of senescence and the liberated amino acids can be exported to growing parts of the plant (e.g. maturing fruits). Rubisco and other stromal enzymes can be degraded in isolated chloroplasts, implying the involvement of plastidial peptide hydrolases. Whether or not ATP is required and if stromal proteins are modified (e.g. by reactive oxygen species) prior to their degradation are questions still under debate. Several proteins, in particular cysteine proteases, have been demonstrated to be specifically expressed during senescence. Their contribution to the general degradation of chloroplast proteins is unclear. The accumulation in intact cells of peptide fragments and inhibitor studies suggest that multiple degradation pathways may exist for stromal proteins and that vacuolar endopeptidases might also be involved under certain conditions. The breakdown of chlorophyll-binding proteins associated with the thylakoid membrane is less well investigated. The degradation of these proteins requires the simultaneous catabolism of chlorophylls. The breakdown of chlorophylls has been elucidated during the last decade. Interestingly, nitrogen present in chlorophyll is not exported from senescencing leaves, but remains within the cells in the form of linear tetrapyrrolic catabolites that accumulate in the vacuole. The degradation pathways for chlorophylls and chloroplast proteins are partially interconnected.

Crop nutrient and especially nitrogen use efficiency (NUE) is both an economically and an environmentally highly desirable trait. It has been estimated that only a third of nitrogen inputs to cereal crop worldwide are recovered in grain for consumption, resulting in a huge waste of resource with major negative impacts on the environment. Most measures of NUE in wheat and other cereals are based on field assessments of crop yields at given N inputs, performance responses to added N fertilizer, or by quantifying N fertilizer recovery rates. However, NUE is a complex trait comprising two key major components, N uptake and N utilization efficiency, both also complex traits in themselves, each involving many physiological processes and biochemical pathways. A deeper understanding of the processes involved in NUE has been a target of the UK Wheat Genetic Improvement Network project (http://www.wgin.org.uk/). This has enabled the breakdown of characteristics contributing to NUE and an assessment of the variation present in those characteristics, predominantly in modern cultivars; a total of 13 years of data have been obtained to date. Significant but limited variation suggests a requirement for broader germplasm screening such as older varieties, landraces, and wild relatives.© The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

As a result of climate changes, land use and agriculture have to adapt to new demands. Agriculture is responsible for a large part of the greenhouse gas (GHG) emissions that have to be urgently reduced in order to protect the environment. At the same time, agriculture has to cope with the challenges of sustainably feeding a growing world population. Reducing the use of the ammonia-nitrate fertilizers that are responsible for a large part of the GHGs released and that have a negative impact on carbon balance is one of the objectives of precision agriculture. One way to reduce N fertilizers without dramatically affecting grain yields is to improve the nitrogen recycling and remobilization performances of plants. Mechanisms involved in nitrogen recycling, such as autophagy, are essential for nutrient remobilization at the whole-plant level and for seed quality. Studies on leaf senescence and nutrient recycling provide new perspectives for improvement. The aim of this review is to give an overview of the mechanisms involved in nitrogen recycling and remobilization during leaf senescence and to present the different approaches undertaken to improve nitrogen remobilization efficiency using both model plants and crop species.© The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Oilseed rape (Brassica napus L.) is a crop plant characterized by a poor nitrogen (N) use efficiency that is mainly due to low N remobilization efficiency during the sequential leaf senescence of the vegetative stage. As a high leaf N remobilization efficiency was strongly linked to a high remobilization of proteins during leaf senescence of rapeseed, our objective was to identify senescence-associated protease activities implicated in the protein degradation. To reach this goal, leaf senescence processes and protease activities were investigated in a mature leaf becoming senescent in plants subjected to ample or low nitrate supply. The characterization of protease activities was performed by using in vitro analysis of RuBisCO degradation with or without inhibitors of specific protease classes followed by a protease activity profiling using activity-dependent probes. As expected, the mature leaf became senescent regardless of the nitrate treatment, and nitrate limitation enhanced the senescence processes associated with an enhanced degradation of soluble proteins. The characterization of protease activities revealed that: (i) aspartic proteases and the proteasome were active during senescence regardless of nitrate supply, and (ii) the activities of serine proteases and particularly cysteine proteases (Papain-like Cys proteases and vacuolar processing enzymes) increased when protein remobilization associated with senescence was accelerated by nitrate limitation. Short statement: Serine and particularly cysteine proteases (both PLCPs and VPEs) seem to play a crucial role in the efficient protein remobilization when leaf senescence of oilseed rape was accelerated by nitrate limitation. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.