Contribution of carbon fixed by Rubisco and PEPC to phloem export in the Crassulacean acid metabolism plant Kalanchoë daigremontiana

Abstract

Crassulacean acid metabolism (CAM) plants exhibit a complex interplay between CO2 fixation by phosphoenolpyruvate carboxylase (PEPC) and ribulose-1,5-bisphosphate carboxylase oxygenase (Rubisco), and carbon demand for CAM maintenance and growth. This study investigated the flux of carbon from PEPC and direct Rubisco fixation to different leaf carbon pools and to phloem sap over the diurnal cycle. Concentrations and carbon isotope compositions of starch, soluble sugars, and organic acids were determined in leaves and phloem exudates of Kalanchoë daigremontiana Hamet et Perr., and related to CO2 fixation by PEPC and Rubisco. Three types of leaf carbon pools could be distinguished. (i) Starch and malate pools were dominant and showed a pattern of reciprocal mobilization and accumulation (85/54 and 13/48 mg C g−1 DW, respective, at the beginning/end of phase I). The carbon isotope composition of these pools was compatible with predominant PEPC fixation (δ13C values of –13 and –11‰ for starch and malate compared to –11‰ of PEPC fixed carbon). (ii) Isotopic composition (–17‰ and –14‰) and concentration of glucose and fructose (2 and 3 mg C g−1 DW, respectively) were not affected by diurnal metabolism, suggesting a low turnover. (iii) Sucrose (1–3 mg C g−1 DW), in contrast, exhibited large diurnal changes in δ13C values (from –17‰ in the evening to –12‰ in the morning), which were not matched by net changes in sucrose concentration. This suggests a high sucrose turnover, fed by nocturnal starch degradation and direct Rubisco fixation during the day. A detailed dissection of the carbon fixation and mobilization pattern in K. daigremontiana revealed that direct fixation of Rubisco during the light accounted for 30% of phloem sucrose, but only 15% of fixed carbon, indicating that carbon from direct Rubisco fixation was preferentially used for leaf export.

Phosphorylation of Phosphoenolpyruvate Carboxylase Is Essential for Maximal and Sustained Dark CO2 Fixation and Core Circadian Clock Operation in the Obligate Crassulacean Acid Metabolism Species Kalanchoë fedtschenkoi

“…CAM leaves of the closely related obligate species Kalanchoë daigremontiana have been reported previously to display a characteristic postdawn peak of sucrose peaking at the end of phase II of CAM (Wild et al, 2010). At the beginning of the light period, wild-type K. fedtschenkoi had a peak of sucrose phased to 2 h after dawn that was reduced to around half the wild-type level in line rPPCK1-3 ( Figure 2E).…”

“…Sucrose has been demonstrated to be a product of CAM-derived starch degradation during the hours at the end of the phase I dark period, although sucrose was also shown to accumulate further during phase II after dawn as a direct result of Rubisco-mediated fixation of atmospheric CO 2 , as is classically observed in the late afternoon in C 3 plants such as Arabidopsis (Wild et al, 2010;Haydon et al, 2013). Both of the KfCCA1 genes and one of the two KfTOC1 genes were downregulated and displayed dampening rhythmicity in the rPPCK1-3 plants under LL conditions (Figure 6), and this may be due to the rPPCK1-3 plants being in a state of metabolic starvation after dawn.…”

Host-compound foraging by intestinal microbiota revealed by single-cell stable isotope probing

“…For compound-specific carbon isotope analysis of acetate, propionate, and butyrate, an HPLC system (Dionex Corporation) coupled to a Delta V Advantage Mass Spectrometer by a Liquid Chromatography (LC) IsoLink Interface (Thermo Fisher Scientific) was used (22). Samples were acidified with 1 M phosphoric acid to remove bicarbonate, and subsequently separated on a Macherey-Nagel Nucleogel Sugar 810H column at 75°C with 0.5 mL·min −1 20 mM phosphoric acid as eluent.…”

“…Peaks were converted to CO 2 at 99°C with 50 μL·min −1 of each 0.5 M sodium persulfate and 1.7 M phosphoric acid. Concentrations and isotopic compositions of individual compounds were calibrated against external standards as described elsewhere (22).…”

Compound-Specific Carbon Isotopes and Concentrations of Carbohydrates and Organic Acids as Indicators of Tree Decline in Mountain Pine

“…More detailed information can be derived from the individual carbohydrates (sucrose, glucose, and fructose), and sugar alcohols (e.g., pinitol), but also from compounds like organic acids (malate and citrate) that are involved in more "downstream metabolic processes" leading to systematic but varying 13 C-enrichment of sink organs (such as stem) in comparison to leaves [20]. Thus far, there have been few investigations analyzing δ 13 C of individual carbohydrates and organic acids [19][20][21][22][23][24][25][26] using high-performance liquid chromatography (HPLC). However, even fewer investigations were applied in natural forest ecosystems to address questions of tree eco-physiology [19,22,23,26,27].…”