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In the near future, field-grown winter oilseed rape may be required to frequently withstand both restricted N availability due to the introduction of low N input crop management practices and limited water supply due to forecast climatic changes. Current oilseed rape breeding programmes are seeking better NUE genotypes adapted to limited N conditions but do not necessarily take into account the impact that such adaptations might have on the plant tolerance to other environmental constraints (drought, high or low temperatures, diseases,…). Nitrogen metabolism is by nature highly mobilized in the process of nutrient recycling between source and sink organs and seed filling. Nitrogen metabolism is also highly mobilized in response to abiotic stresses. The objective is to investigate how metabolic adjustments under stress can both participate to tolerance and interfere with the trophic processes and efficiency of nitrogen use in plants.

Research and main results

The functional value of metabolism known to be induced under stress conditions is sought. These include the metabolic pathways involved in mobilizing glutamate and directed to glutamine, proline and gamma-aminobutyric acid (Gaba) production and utilization. Proteases and protease inhibitors are also being studied as key effectors of nitrogen remobilization whose regulation by stress is investigated. Besides these targeted investigations more comprehensive metabolomic approaches are performed in oilseed rape under stress and in related species more or less adapted to adverse conditions with the prospect of discovering metabolic markers of tolerance.

1) Proteolysis regulation under stress

The effective recycling of the N compounds from source leaves to sink growing tissues requires a fine coordination between sink demand and the process of proteolysis. Involvement of protease inhibitors and proteases in the control of proteolysis during the remobilisation process is investigated by identification of proteases and protease inhibitors as potential regulators of natural or drought induced senescence. Thus, BnD22, a protease inhibitor, may have an important role in plant resistance to diverse forms of stress and may contribute to a better utilization of recycling N from sources, a physiological trait that improves N-use efficiency.

2) Stress induced glutamate-derived pathways

The regulation of glutamate (Glu) metabolism appears to be of considerable importance in the N economy of plants. Using aminoacid pool released via proteolysis, series of transamination reactions lead to an increase in Glu pool that could serve immediately as a substrate for Glutamine Synthetase 1 (GS1) and Glutamate dehydrogenase (GDH). Therefore gene coding GS1 enzymes are candidate genes studied for their potential involvement in N remobilisation during seed filling under both optimal and stress conditions. Gaba (g-aminobutyric acid), is also derived from Glu via the activity of Glu decarboxylase (GAD). This molecule is known to accumulate in response to a wide range of environmental stimuli. It has been recently shown that GABA may act as a putative long-distance signal molecule in up-regulation of nitrate uptake in B. napus. Moreover, GABA-transaminase (GABA-T), the first enzyme of the GABA catabolism, is up-regulated during leaf senescence and under osmotic stress conditions. We performed a functional analysis of the GABA-T gene in Arabidopsis thaliana and demonstrated that the previously isolated loss-of-function GABA-T mutant is affected in root development, salt stress tolerance and C/N ratio regulation.

Oversensitive phenotype of pop2-1 (Gaba transaminase) mutant in response to NaCl (from Renault et al., 2010)

Under osmotic stresses, Glu becomes a predominant precursor for proline accumulation as a compatible solute in oilseed rape via P5CS (Pyroline 5-carboxylate synthetase) and PDH (Proline dehydrogenase) activity regulations. Proline can be also considered as a storage compound since it becomes readily available during the post-stress period. PDH is under focus as a major contributor to proline metabolism regulation and nitrogen use efficiency under osmotic stress.

3) Functional metabolomic of stress tolerance

We consider both the studies of targeted biochemical families of functional importance (e.g. osmoprotectants, anti-oxidants, growth regulators,...) and the non-supervised metabolomic investigations dedicated to the characterization of novel metabolic attributes of drought and salt stress tolerance. A close attention is given to the exploration of metabolic fingerprinting reports done on oilseed rape grown under controlled conditions with combined nitrogen and drought stress or Arabidopsis and related species. Metabolic signatures of extremophile taxons recently proposed as model species for abiotic stress tolerance investigations (e.g. Thellungiella sp.; Arabis sp.) are researched.

Metabolic signatures of oilseed rape leaves submitted to both nitrogen and water stresses (Albert et al., in prep.)

Main references

Lugan R., Niogret M.F., Kervazo L., Larher F.L., Kopka J., Bouchereau A., 2009 - Metabolome and water status phenotyping of Arabidopsis under abiotic stress cues reveals new insight into ESK1 function. Plant Cell Env., 32, 95–108.

Lugan R., Niogret MF., Leport L.,  Guégan JP.,  Larher F., Savouré A.,  Kopka J.,  Bouchereau, A., 2010 - Metabolome and water homeostasis analysis of Thellungiella salsuginea suggests that dehydration tolerance is a key response to osmotic stress in this halophyte. Plant J. 64, 215-229.

Renault H., Roussel V., El Amrani A., Arzel M., Renault D., Bouchereau A., Deleu C., 2010 – The Arabidopsis pop2-1 mutant reveals the involvement of GABA transaminase in salt stress tolerance. BMC Plant Biology, 10, 1-16.

Renault H., El Amrani A., Palanivelu R., Updegraff EP., Yu A., Renou JP., Preuss D., Bouchereau A., Deleu C., 2011 – GABA accumulation causes cell elongation defects and a decrease in expression of genes encoding secreted and cell-wall-related proteins in Arabidopsis thaliana.

Plant Cell Physiol., 52(5), 894-908.Albert B., Le Cahérec F., Niogret M.F., Faes P., Avice J.C., Leport L., Bouchereau A. (2012) Nitrogen availability impacts oilseed rape (Brassica napus L.) plant water status and proline production efficiency under water-limited conditions. Planta, 236, 659-676.

Renault H., El Amrani A., Berger A., Mouille G., Soubigou-Taconnat L., Bouchereau A., Deleu C., 2013 - γ-Aminobutyric acid transaminase deficiency impairs central carbon metabolism and leads to cell wall defects during salt stress in Arabidopsis roots. Plant Cell Environ. DOI

Collaborations

• Unité "Physiologie Cellulaire et Moléculaire des Plantes" , UPMC, Paris, France
• UMR950 Ecophysiologie Végétale, Agronomie et Nutritions N, C, S (EVA), INRA-UCBN, France.
• IJPB,  INRA Versailles, France
• IRSTEA, UR TERE, Rennes, France
• UMR 5553, LECA, CNRS-UJF, Grenoble, France
• Max Planck Institute of Molecular Plant Physiology, Potsdam, Germany
• Institute of Molecular Physiology and Biotechnology of Plants, University of Bonn, Germany