Light enables a very high efficiency of carbon storage in developing embryos of rapeseed

Abstract

The conversion of photosynthate to seed storage reserves is crucial to plant fitness and agricultural production, yet quantitative information about the efficiency of this process is lacking. To measure metabolic efficiency in developing seeds, rapeseed (Brassica napus) embryos were cultured in media in which all carbon sources were [U-14C]-labeled and their conversion into CO2, oil, protein, and other biomass was determined. The conversion efficiency of the supplied carbon into seed storage reserves was very high. When provided with 0, 50, or 150 micromol m(-2) s(-1) light, the proportion of carbon taken up by embryos that was recovered in biomass was 60% to 64%, 77% to 86%, and 85% to 95%, respectively. Light not only improved the efficiency of carbon storage, but also increased the growth rate, the proportion of 14C recovered in oil relative to protein, and the fixation of external 14CO2 into biomass. Embryos grown at 50 micromol m(-2) s(-1) in the presence of 5 microM 1,1-dimethyl-3-(3,4-dichlorophenyl) urea (an inhibitor of photosystem II) were reduced in total biomass and oil synthesis by 3.2-fold and 2.8-fold, respectively, to the levels observed in the dark. To explore if the reduced growth and carbon conversion efficiency in dark were related to oxygen supplied by photosystem II, embryos and siliques were cultured with increased oxygen. The carbon conversion efficiency of embryos remained unchanged when oxygen levels were increased 3-fold. Increasing the O2 levels surrounding siliques from 21% to 60% did not increase oil synthesis rates either at 1,000 micromol m(-2) s(-1) or in the dark. We conclude that light increases the growth, efficiency of carbon storage, and oil synthesis in developing rapeseed embryos primarily by providing reductant and/or ATP.

Figure 1.

Distribution of ¹⁴C-labeling among biomass fractions and CO₂ (100% = ¹⁴C biomass + ¹⁴CO₂): (A) 0 μmol m⁻² s⁻¹, (B) 50 μmol m⁻² s⁻¹, and (C) 150 μmol m⁻² s⁻¹. Embryos were cultured at 21°C for 14 d (at 150 and 50 μmol m⁻² s⁻¹) and 21 d (at 0 μmol m⁻² s⁻¹) in the presence of U-¹⁴C carbon supplies. After culture, the ¹⁴CO₂ was collected by flushing the flasks through a trapping system, and the embryos were frozen in liquid N₂ for analysis of ¹⁴C-labeling in biomass components. The percent values shown represent the percentage of ¹⁴C recovered in the respective fractions. Because oil, protein, and carbohydrate contain, respectively, 77%, 53%, and 40% carbon by weight, these values differ from weight percent values such as presented in Table I.

Figure 2.

Effect of light irradiance on ¹⁴CO₂ incorporation into biomass. Embryos were grown under a 2% ¹⁴CO₂ atmosphere (specific activity of C = 44.9 mCi/mol) for 3 d at 150, 50, and 0 μmol m⁻² s⁻¹. After culture, the embryos were rapidly frozen in liquid N₂, and the ¹⁴C-labeling in biomass compounds was measured. A, Total biomass ¹⁴C-labeling (microcuries). B, ¹⁴C-labeling in oil (microcuries). C, ¹⁴C-labeling in protein (microcuries). D, ¹⁴C-labeling in starch + cell wall (microcuries). Columns and error bars correspond to the mean ± sd of three independent replicates. Means with the same letter are not significantly different at the P < 0.05 level (Tukey's studentized range test).

Figure 3.

Effect of inhibition of PSII by DCMU on biomass production, oil synthesis, carbon conversion efficiency, and the oil to CO₂ ratio under light and dark conditions. Embryos were cultured in the presence of U-¹⁴C carbon sources for 3 d at 50 μmol m⁻² s⁻¹ or in the dark with or without DCMU. After culture, the ¹⁴C-labeling in the CO₂ and biomass compounds was determined. A, Total biomass production expressed in milligrams dry weight (seven embryos). B, Oil synthesis expressed in milligrams (seven embryos). C, CCE, expressed as percent of the sum of ¹⁴C biomass and ¹⁴CO₂. D, Oil to CO₂ ratio, calculated from the ratio of ¹⁴C-labeling between oil and CO₂. Columns and error bars correspond to the mean ± sd of three independent replicates. Means with the same letter are not significantly different at the P < 0.05 level (Tukey's studentized range test).

Figure 4.

Effect of oxygen on biomass production, oil synthesis, CCE, and oil to CO₂ ratio under dark conditions. Embryos were cultured in the dark for 3 d under approximately 1%, 21% (ambient oxygen), and 64% (v/v) O₂ in the presence of U-¹⁴C carbon sources. A, Total biomass production expressed in milligrams dry weight (seven embryos). B, Oil synthesis, expressed in milligrams (seven embryos). C, CCE expressed as percent of the sum of ¹⁴C biomass and ¹⁴CO₂. D, Oil to CO₂ ratio, calculated from the ratio of ¹⁴C-labeling between oil and CO₂. Columns and error bars correspond to the mean ± sd of three independent replicates. Means with the same letter are not significantly different at the P < 0.05 level (Tukey's studentized range test).

Figure 5.

Incorporation of radiolabel from (A) ³H₂O and (B) [U-¹⁴C]Suc into lipids in seeds inside siliques as influenced by light and oxygen levels. A, detached siliques were prelabeled for 24 h (initial) in a buffered solution containing ³H₂O. They were then incubated at 1,000 μmol (light treatment) for further 24 h either at ambient air or at 60% oxygen with one-half of the siliques being loosely covered with aluminum foil (dark treatment). After incubation, four seeds from the central region of each silique were removed, the lipids extracted, and the ³H incorporation into fatty acids determined. Columns and error bars correspond to the mean ± se of 12 independent replicates. Means with the same letter are not significantly different at the P < 0.05 level (Tukey's studentized range test). B, Detached siliques were cultured in the dark in the presence of [U-¹⁴C]Suc for 4 d either at ambient air or at 60% oxygen. After incubation, the seeds from each silique were removed, the lipids extracted, and the ¹⁴C incorporation into oil determined. Columns and error bars correspond to the mean ± sd of 12 independent replicates. Means with the same letter are not significantly different at the P < 0.05 level (Tukey's studentized range test).

Figure 6.

Diagram of the culture system used to perform 3-d carbon balances of rapeseed embryos under 2% CO₂.