Metabolic and physiological adjustment of Suaeda maritima to combined salinity and hypoxia

BACKGROUND AND AIMS: Suaeda maritima is a halophyte commonly found on coastal wetlands in the intertidal zone. Due to its habitat S. maritima has evolved tolerance to high salt concentrations and hypoxic conditions in the soil caused by periodic flooding. In the present work, the adaptive mechanisms of S. maritima to salinity combined with hypoxia were investigated on a physiological and metabolic level. METHODS: To compare the adaptive mechanisms to deficient, optimal and stressful salt concentrations, S. maritima plants were grown in a hydroponic culture under low, medium and high salt concentrations. Additionally, hypoxic conditions were applied to investigate the impact of hypoxia combined with different salt concentrations. A non-targeted metabolic approach was used to clarify the biochemical pathways underlying the metabolic and physiological adaptation mechanisms of S. maritima . KEY RESULTS: Roots exposed to hypoxic conditions showed an increased level of tricarboxylic acid (TCA)-cycle intermediates such as succinate, malate and citrate. During hypoxia, the concentration of free amino acids increased in shoots and roots. Osmoprotectants such as proline and glycine betaine increased in concentrations as the external salinity was increased under hypoxic conditions. CONCLUSIONS: The combination of high salinity and hypoxia caused an ionic imbalance and an increase of metabolites associated with osmotic stress and photorespiration, indicating a severe physiological and metabolic response under these conditions. Disturbed proline degradation in the roots induced an enhanced proline accumulation under hypoxia. The enhanced alanine fermentation combined with a partial flux of the TCA cycle might contribute to the tolerance of S. maritima to hypoxic conditions

U-13C-glucose incorporation into source leaves of Brassica napus highlights light-dependent regulations of metabolic fluxes within central carbon metabolism

International audiencePlant central carbon metabolism comprises several important metabolic pathways acting together to support plant growth and yield establishment. Despite the emergence of 13 C-based dynamic approaches, the regulation of metabolic fluxes between light and dark conditions has not yet received sufficient attention for agronomically relevant plants. Here, we investigated the impact of light/dark conditions on carbon allocation processes within central carbon metabolism of Brassica napus after U-13 C-glucose incorporation into leaf discs. Leaf gas-exchanges and metabolite contents were weakly impacted by the leaf disc method and the incorporation of glucose. 13 Canalysis by GC-MS showed that U-13 C-glucose was converted to fructose for de novo biosynthesis of sucrose at similar rates in both light and dark conditions. However, light conditions led to a reduced commitment of glycolytic carbons towards respiratory substrates (pyruvate, alanine, malate) and TCA cycle intermediates compared to dark conditions. Analysis of 13 C-enrichment at the isotopologue level and metabolic pathway isotopic tracing reconstructions identified the contribution of multiple pathways to serine biosynthesis in light and dark conditions. However, the direct contribution of the glucose-6-phosphate shunt to serine biosynthesis was not observed. Our results also provided isotopic evidences for an active metabolic connection between the TCA cycle, glycolysis and photorespiration in light conditions through a rapid reallocation of TCA cycle decarboxylations back to the TCA cycle through photorespiration and glycolysis. Altogether, these results suggest the active coordination of core metabolic pathways across multiple compartments to reorganize C-flux modes

Impact of climate warming on carbon metabolism and on morphology of invasive and native aquatic plant species varies between spring and summer

International audienceThe rise of global surface temperature by between 1.2 °C and 4 °C by 2100 is expected to affect freshwater ecosystems and the growth of aquatic plants. By extending the distribution range of invasive macrophytes, climate warming could increase their management costs. The aim of this study was to test the impact of a 3 °C warming in spring and in summer on the morphology and physiology of two native species (Mentha aquatica, Myosotis scorpioides) and two invasive species (Ludwigia hexapetala, Myriophyllum aquaticum) under controlled conditions. Our study showed that the increase of spring temperature induced morphological modifications for all species, while a 3 °C warming induced changes in carbohydrates composition for native species in spring, and modification of carbohydrate content for invasive species at both seasons. Patterns of carbohydrate content group the two invasive species together, possibly highlighting common physiological mechanisms. Moreover, the increase of spring temperature favoured the apical and/or lateral growth solely for invasive species. Hence, the invasive species specific response to warming suggests that higher temperature may favour their growth in spring, which might allow them to colonise the water column earlier than natives. This competitive advantage could affect aquatic ecosystems functioning and biodiversity in the coming years

Using 13 C metabolic probes to assess carbon allocation processes within central metabolism of Brassica napus source leaves

International audiencePlant central metabolism comprises several essential metabolic pathways acting together to support plant growth and yield establishment. The deployment of 13C-tracing dynamic approaches in the model brassica specie Arabidopsis allowed to improve our understanding of this complex metabolic network by identifying major leaf carbon allocation processes. However, some questions remained open for Brassica napus leaves, an economically relevant oleaginous crop experiencing sequential senescence during its vegetative growth. Notably, the functioning of the tricarboxylic acid (TCA) cycle in the light in relation to glycolysis, Calvin cycle, photorespiration and branched-chain amino acid catabolism could be substantially different in source leaves in order to support senescence-associated remobilization processes. Here, we investigated these carbon fluxes by performing short-term incorporations of different fully 13C-labelled metabolic probes into Brassica napus leaf discs. Indeed, this experimental setup showed relatively similar photosynthetic and respiratory capacities compared to attached leaves. Evaluation of 13C-enrichments at molecular and isotopologue levels by GC-MS identified light/dark regulation of key carbon fluxes and separated competing metabolic contributions to metabolite biosynthesis. Notably, the results supported: i) the occurrence of both cyclic and non-cyclic flux modes of TCA cycle in the light; ii) the contribution of glycolysis rather than stored citrate to TCA cycle functioning in the light; iii) the reallocation of TCA cycle decarboxylations back to the TCA cycle through photorespiration and glycolysis; iv) The multiple contributions of branched-chain amino acid catabolism to TCA cycle in the light. Overall, our results improved our understanding of central metabolism in source leaves of Brassica napus

Characterisation of the salt stress vulnerability of three invasive freshwater plant species using a metaolic profiling approach

the Corsaire P2M2 Platform of BiogenouestInternational audienceThe effects of salt stress on freshwater plants has been little studied up to now, despite the fact that they are expected to present different levels of salt sensitivity or salt resistance depending on the species. The aim of this work was to assess the effect of NaCl at two concentrations on three invasive freshwater species, Elodea canadensis, Myriophyllum aquaticum and Ludwigia grandiflora, by examining morphological and physiological parameters and using metabolic profiling. The growth rate (biomass and stem length) was reduced for all species, whatever the salt treatment, but the response to salt differed between the three species, depending on the NaCl concentration. For E. canadensis, the physiological traits and metabolic profiles were only slightly modified in response to salt, whereas M. aquaticum and L. grandiflora showed great changes. In both of these species, root number, photosynthetic pigment content, amino acids and carbohydrate metabolism were affected by the salt treatments. Moreover, we are the first to report the salt-induced accumulation of compatible solutes in both species. Indeed, in response to NaCl, L. grandiflora mainly accumulated sucrose. The response of M. aquaticum was more complex, because it accumulated not only sucrose and myo-inositol whatever the level of salt stress, but also amino acids such as proline and GABA, but only at high NaCl concentrations. These responses are the metabolic responses typically found in terrestrial plants

Validation of carbon isotopologue distribution measurements by GC-MS and application to 13 C-metabolic flux analysis of the tricarboxylic acid cycle in Brassica napus leaves

International audienceThe estimation of metabolic fluxes in photosynthetic organisms represents an important challenge that has gained interest over the last decade with the development of 13C-Metabolic Flux Analysis at isotopically non-stationary steady-state. This approach requires a high level of accuracy for the measurement of Carbon Isotopologue Distribution in plant metabolites. But this accuracy has still not been evaluated at the isotopologue level for GC-MS, leading to uncertainties for the metabolic fluxes calculated based on these fragments. Here, we developed a workflow to validate the measurements of CIDs from plant metabolites with GC-MS by producing tailor-made E. coli standard extracts harboring a predictable binomial CID for some organic and amino acids. Overall,most of our TMS-derivatives mass fragments were validated with these standards and at natural isotope abundance in plant matrices. Then, we applied this validated MS method to investigate the light/ dark regulation of plant TCA cycle by incorporating U-13C-pyruvate to Brassica napus leaf discs. We took advantage of pathway-specific isotopologues/ isotopomers observed between two and six hours of labeling to show that the TCA cycle can operate in a cyclic manner under both light and dark conditions. Interestingly, this forward cyclic flux mode has a nearly four-fold higher contribution for pyruvate-to-citrate and pyruvate-to-malate fluxes than the phosphoenolpyruvate carboxylase (PEPc) flux reassimilating carbon derived from some mitochondrial enzymes. The contribution of stored citrate to the mitochondrial TCA cycle activity was also questioned based on dynamics of 13C-enrichment in citrate, glutamate and succinate and variations of citrate total amounts under light and dark conditions. Interestingly, there was a light-dependent 13C-incorporation into glycine and serine showing that decarboxylations from pyruvate dehydrogenase complex and TCA cycle enzymes were actively reassimilated and could represent up to 5% to net photosynthesis

Manipulating feeding stimulation to protect crops against insect pests?

International audienceEnhancing natural mechanisms of plant defense against herbivores is one of the possible strategies to protect cultivated species against insect pests. Host plant feeding stimulation, which results from phagostimulant and phagodeterrent effects of both primary and secondary metabolites, could play a key role in levels of damage caused to crop plants. We tested this hypothesis by comparing the feeding intensity of the pollen beetle Meligethes aeneus on six oilseed rape (Brassica napus) genotypes in a feeding experiment, and by assessing the content of possible phagostimulant and phagodeterrent compounds in tissues targeted by the insect (flower buds). For this purpose, several dozens of primary and secondary metabolites were quantified by a set of chromatographic techniques. Intergenotypic variability was found both in the feeding experiment and in the metabolic profile of plant tissues. Biochemical composition of the perianth was in particular highly correlated with insect damage. Only a few compounds explained this correlation, among which was sucrose, known to be highly phagostimulating. Further testing is needed to validate the suggested impact of the specific compounds we have identified. Nevertheless, our results open the way for a crop protection strategy based on artificial selection of key determinants of insect feeding stimulation

Overexpression of ALDH10A8 and ALDH10A9 Genes Provides Insight into Their Role in Glycine Betaine Synthesis and Affects Primary Metabolism in Arabidopsis thaliana

International audienceBetaine aldehyde dehydrogenases oxidize betaine aldehyde to glycine betaine in species that accumulate glycine betaine as a compatible solute under stress conditions. In contrast, the physiological function of betaine aldehyde dehydrogenase genes is at present unclear in species that do not accumulate glycine betaine, such as Arabidopsis thaliana. To address this question, we overexpressed the Arabidopsis ALDH10A8 and ALDH10A9 genes, which were identified to code for betaine aldehyde dehydrogenases, in wild-type A. thaliana. We analysed changes in metabolite contents of transgenic plants in comparison with the wild type. Using exogenous or endogenous choline, our results indicated that ALDH10A8 and ALDH10A9 are involved in the synthesis of glycine betaine in Arabidopsis. Choline availability seems to be a factor limiting glycine betaine synthesis. Moreover, the contents of diverse metabolites including sugars (glucose and fructose) and amino acids were altered in fully developed transgenic plants compared with the wild type. The plant metabolic response to salt and the salt stress tolerance were impaired only in young transgenic plants, which exhibited a delayed growth of the seedlings early after germination. Our results suggest that a balanced expression of the betaine aldehyde dehydrogenase genes is important for early growth of A. thaliana seedlings and for salt stress mitigation in young seedling

Polyphenol composition and antioxidant activity of Searsia tripartita and Limoniastrum guyonianum growing in Southeastern Algeria

International audienceDue to the notable medicinal value of Searsia tripartita and Limoniastrum guyonianum, it was of great interest to conduct phytochemical and antioxidant investigations. Polyphenol extracts of the dried powder of plants were prepared by maceration in 70% of methanol. The concentrated extracts were successively fractioned with distilled water, ethyl acetate and n-butanol. Obtained extracts were analysed for their phenol compositions by colorimetric processes and were evaluated for their total antiradical capacity by ABTS, DPPH and ORAC assays. Additionally, the phytochemical components were characterized in the methanolic extracts by HPLC–DAD–ESI-MS/MS in the negative ionisation mode. S. tripartita exhibited the higher quantity of phenolic compounds. The ethyl acetate extract of S. tripartita had the greatest quantities of polyphenols (55.5 ± 4.9 mg gallic acid equivalent per gram of dry residue (GAE/g DR)), flavonoids (44.2 ± 0.8 mg rutin equivalent per gram of dry residue (RE/g DR)), and tannins (12.5 ± 2.5 mg GAE/g DR). In L. guyonianum, the aqueous fraction had the highest quantity of polyphenols and tannins (10.2 ± 1.4 and 3.2 ± 1.9 mg GAE/g DR, respectively), whereas flavonoids (1.8 ± 0.2 mg RE/g DR) were higher in the ethyl acetate portion. The antioxidant capacity of the hydromethanolic extract of S. tripartita was found to be 3–16 times more effective than that of L. guyonianum using ORAC, DPPH, ABTS tests. HPLC analysis of hydromethanolic extracts provided tentative identification of four flavonoid glycosides in S. tripartita (myricetin-3-O-glucoside, myricetin-3-O-rhamnoside, quercetin-3-O-glucoside, and quercetin-3-O-rhamnoside) and five phenolic metabolites of the flavonoid class in L. guyonianum (myricetin-3-O-rhamnosylglucoside, myricetin-3-O-glucuronide, myricetin-3-O-pentoside, quercetin-3-O-glucuronide, and eriodictyol-7-O-rutinoside). Our findings revealed that these plants could be used as a potent source of health molecules