Do nitrogen- and sulphur-remobilization-related parameters measured at the onset of the reproductive stage provide early indicators to adjust N and S fertilization in oilseed rape (Brassica napus L.) grown under N- and/or S-limiting supplies?

MAIN CONCLUSION: Specific combinations of physiological and molecular parameters associated with N and S remobilization measured at the onset of flowering were predictive of final crop performances in oilseed rape. Oilseed rape (Brassica napus L.) is a high nitrogen (N) and sulphur (S) demanding crop. Nitrogen- and S-remobilization processes allow N and S requirements to reproductive organs to be satisfied when natural uptake is reduced, thus ensuring high yield and seed quality. The quantification of physiological and molecular indicators of early N and S remobilization could be used as management tools to correct N and S fertilization. However, the major limit of this corrective strategy is to ensure the correlation between final performances-related variables and early measured parameters. In our study, four genotypes of winter oilseed rape (OSR) were grown until seed maturity under four nutritional modalities combining high and/or low N and S supplies. Plant final performances, i.e., seed production, N- and S-harvest indexes, seed N and S use efficiencies, and early parameters related to N- or S-remobilization processes, i.e., photosynthetic leaf area, N and S leaf concentrations, leaf soluble protein and leaf sulphate concentrations, and leaf RuBisCO abundance at flowering, were measured. We demonstrated that contrasting final performances existed according to the N and S supplies. An optimal N:S ratio supply could explain the treatment-specific crop performances, thus justifying N and S concurrent managements. Specific combinations of early measured plant parameters could be used to predict final performances irrespective of the nutritional supply and the genotype. This work demonstrates the potential of physiological and molecular indicators measured at flowering to reflect the functioning of N- and S-compound remobilization and to predict yield and quality penalties. However, because the predictive models are N and S independent, instant N and S leaf analyses are required to further adjust the adequate fertilization. This study is a proof of a concept which opens prospects regarding instant diagnostic tools in the context of N and S mineral fertilization management.

Germinative and Post-Germinative Behaviours of Brassica napus Seeds Are Impacted by the Severity of S Limitation Applied to the Parent Plants.

In oilseed rape (Brassica napus L.), sulphur (S) limitation leads to a reduction of seed yield and nutritional quality, but also to a reduction of seed viability and vigour. S metabolism is known to be involved in the control of germination sensu stricto and seedling establishment. Nevertheless, how the germination and the first steps of plant growth are impacted in seeds produced by plants subjected to various sulphate limitations remains largely unknown. Therefore, this study aimed at determining the impact of various S-limited conditions applied to the mother plants on the germination indexes and the rate of viable seedlings in a spring oilseed rape cultivar (cv. Yudal). Using a 34S-sulphate pulse method, the sulphate uptake capacity during the seedling development was also investigated. The rate of viable seedlings was significantly reduced for seeds produced under the strongest S-limited conditions. This is related to a reduction of germination vigour and to perturbations of post-germinative events. Compared to green seedlings obtained from seeds produced by well-S-supplied plants, the viable seedlings coming from seeds harvested on plants subjected to severe S-limitation treatment showed nonetheless a higher dry biomass and were able to enhance the sulphate uptake by roots and the S translocation to shoots.

Heat stress during seed filling interferes with sulfur restriction on grain composition and seed germination in oilseed rape (Brassica napus L.).

In coming decades, increasing temperatures are expected to impact crop yield and seed quality. To develop low input systems, the effects of temperature and sulfur (S) nutrition in oilseed rape, a high S demanding crop, need to be jointly considered. In this study, we investigated the effects of temperatures [High Temperature (HT), 33°C/day, 19°C/night vs. Control Temperature (Ctrl T), 20°C/day, 15°C/day] and S supply [High S (HS), 500 µm SO(2-) 4 vs. Low S (LS), 8.7 µM SO(2-) 4] during seed filling on (i) yield components [seed number, seed dry weight (SDW) and seed yield], (ii) grain composition [nitrogen (N) and S contents] and quality [fatty acid (FA) composition and seed storage protein (SSP) accumulation] and (iii) germination characteristics (pre-harvest sprouting, germination rates and abnormal seedlings). Abscisic acid (ABA), soluble sugar contents and seed conductivity were also measured. HT and LS decreased the number of seeds per plant. SDW was less affected due to compensatory effects since the number of seeds decreased under stress conditions. While LS had negative effects on seed composition by reducing the FA contents and increasing the ratio S-poor SSPs (12S globulins)/S-rich SSPs (2S albumins) ratio, HT had positive effects by increasing S and FA contents and decreasing the C18:2/C18:3 ratio and the 12S/2S protein ratio. Seeds produced under HT showed high pre-harvest sprouting rates along with decreased ABA contents and high rates of abnormal seedlings. HT and LS restriction significantly accelerated germination times. High conductivity, which indicates poor seed storage capacity, was higher in HT seeds. Consistently, the lower ratio of (raffinose + stachyose)/sucrose in HT seeds indicated low seed storage capacity. We demonstrated the effects of HT and LS on grain and on germination characteristics. These results suggest that hormonal changes might control several seed characteristics simultaneously.

Corrigendum: Heat Stress during Seed Filling Interferes with Sulfur Restriction on Grain Composition and Seed Germination in Oilseed Rape (Brassica napus L.).

[This corrects the article on p. 213 in vol. 6, PMID: 25914702.].

Evidence for proteomic and metabolic adaptations associated with alterations of seed yield and quality in sulfur-limited Brassica napus L.

In Brassica napus, seed yield and quality are related to sulfate availability, but the seed metabolic changes in response to sulfate limitation remain largely unknown. To address this question, proteomics and biochemical studies were carried out on mature seeds obtained from plants grown under low sulfate applied at the bolting (LS32), early flowering (LS53), or start of pod filling (LS70) stage. The protein quality of all low-sulfate seeds was reduced and associated with a reduction of S-rich seed storage protein accumulation (as Cruciferin Cru4) and an increase of S-poor seed storage protein (as Cruciferin BnC1). This compensation allowed the protein content to be maintained in LS70 and LS53 seeds but was not sufficient to maintain the protein content in LS32 seeds. The lipid content and quality of LS53 and LS32 seeds were also affected, and these effects were primarily associated with a reduction of C18-derivative accumulation. Proteomics changes related to lipid storage, carbohydrate metabolism, and energy (reduction of caleosins, phosphoglycerate kinase, malate synthase, ATP-synthase ß-subunit, and thiazole biosynthetic enzyme THI1 and accumulation of ß-glucosidase and citrate synthase) provide insights into processes that may contribute to decreased oil content and altered lipid composition (in favor of long-chain fatty acids in LS53 and LS32 seeds). These data indicate that metabolic changes associated with S limitation responses affect seed storage protein composition and lipid quality. Proteins involved in plant stress response, such as dehydroascorbate reductase and Cu/Zn-superoxide dismutase, were also accumulated in LS53 and LS32 seeds, and this might be a consequence of reduced glutathione content under low S availability. LS32 treatment also resulted in (i) reduced germination vigor, as evidenced by lower germination indexes, (ii) reduced seed germination capacity, related to a lower seed viability, and (iii) a strong decrease of glyoxysomal malate synthase, which is essential for the use of fatty acids during seedling establishment.

A specific method of 34S labelling provides evidence that sulphate assimilation occurs in developing seeds and pod walls of Brassica napus L. subjected to ample or limited S nutrition.

RATIONALE: Seeds from different species actively assimilate sulphur (S) from sulphate. This has never been proved for Brassica napus L., a high S demanding plant, especially with regard to S limitation. The role of pod walls in the assimilation and allocation of S in well-fed and sulphate-limited conditions also needs to be clarified. METHODS: Freshly harvested seeds and pod walls from plants well-supplied (HS) or limited with sulphate (LS) from the 'visible buds' stage were subjected to a nutrient solution containing (34)S-sulphate (10 atom% excess) for 24 h. The (34)S labelling of the sulphate and protein fractions was determined with an elemental analyser connected to a continuous flow isotope ratio mass spectrometer. The amino acid profiles of seeds and pod walls were also determined by ion-exchange chromatography. RESULTS: The 24 h of (34)S-sulphate feeding treatment leads to an important production of proteins in HS and LS seeds, associated with a decrease in numerous amino acid contents. The treatment also leads to an incorporation of (34)S in seeds and pod walls proteins in both HS and LS conditions. The incorporation of (34)S in proteins was not different between HS and LS seeds, but was lower in LS than in HS pod walls, related to a higher incorporation in the other organic S compounds. CONCLUSIONS: This study provides evidence that Brassica napus seeds and pod walls are able to assimilate sulphate in HS and LS conditions, and that the LS condition leads to enhancement of the sulphate assimilation capacity of pod walls, which may be of crucial importance for the allocation of S to developing seeds.

Sulphur limitation provokes physiological and leaf proteome changes in oilseed rape that lead to perturbation of sulphur, carbon and oxidative metabolisms.

BACKGROUND: The decline in industrial emissions of sulphur (S) has led to a sulphate depletion in soil resulting in an alteration of crop performance. In oilseed rape, an S deficiency dramatically reduced the seed yield and/or quality. Paradoxically, little is known about the impact of sulphate limitation on oilseed rape leaf metabolism, despite it being a key determinant of growth. In order to identify the metabolic processes involved in the oilseed rape response to S restriction, an analysis of the young leaf proteome combined with a physiological study was carried out at the vegetative stage. RESULTS: S limitation does not significantly reduce the total shoot biomass but inhibits growth and photosynthesis of young leaves. This photosynthesis decline is not due to a decrease in chlorophyll content, which remains similar to Control. The increase in anthocyanins and H(2)O(2) content in young leaves of S-limited plants suggests that S restriction leads to an oxidative stress. Proteomic analysis at 35 d of S limitation also revealed the induction of 12-oxophitodienoate reductase and ACC synthase, respectively involved in jasmonate and ethylene biosynthesis, two phytohormones that could be implicated in oxidative stress. Proteins involved in photosynthesis and carbon metabolism were also modulated by S restriction. In particular, the decrease in plastocyanin and ferredoxin-NADP reductase suggests that H(2)O(2) accumulation is associated with perturbation of the photosynthetic electron transport chain. The accumulation of chloroplastic Cu-Zn SOD reinforces the idea that an oxidative stress probably occurs in the chloroplast. Proteomic results suggest that the maintenance of chlorophyll in S-limited conditions is related to an accumulation of Water Soluble Chlorophyll binding Proteins, involved in the protection of chlorophyll against ROS. The accumulation of the catalytic α-subunit of chloroplastic ATP synthase suggests that energy production is maintained. CONCLUSION: S limitation leads to photosynthesis and carbon metabolism disturbances that could be responsible for the oxidative stress observed in the young leaves of oilseed rape. Despite this, induction of proteins involved in oxidative stress resistance and energy production shows that the leaf capacity to capture and use photosynthetic active radiations for ATP production remains efficient for as long as possible.