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When isotopes of carbon are fed to photosynthesizing leaves, metabolites of the Calvin–Benson cycle (CBC) are rapidly labeled initially, but then the rate of labeling slows considerably, raising questions about the integration of the CBC within leaf metabolism. We have used 2-h time courses of labeling of Camelina sativa leaf metabolites to test models of 12 C washout when the CO 2 source is rapidly switched to 13 CO 2 . Fitting exponential functions to the time course of CBC metabolites, we found evidence for three temporally distinct processes contributing to the labeling but none for metabolically inactive pools. We next modeled the data of all metabolites by 13 C isotopically nonstationary metabolic flux analysis, testing a variety of flux networks. In the model that best explains measured data, three processes determine CBC metabolite labeling. First is fixation of incoming 13 CO 2 ; second is dilution by weakly labeled carbon in cytosolic glucose reentering the CBC following oxidative pentose phosphate pathway reactions, which forms a shunt bypassing much of the CBC. Third, very weakly labeled carbon from the vacuole further dilutes the labeling. This model predicts the shunt proceeds at about 5% of the rate of net CO 2 fixation and explains the three phases of labeling. In showing the interconnection of three compartments, we have drawn a more complete picture of how carbon moves through photosynthetic metabolism in a way that integrates the CBC, cytosolic sugar pools, glucose-6-phosphate shunt, and vacuolar sugars into a single system.
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This content will become publicly available on June 9, 2026
In vitro demonstration and in planta characterization of a condensed, reverse TCA (crTCA) cycle
IntroductionPlants employ the Calvin-Benson cycle (CBC) to fix atmospheric CO2for the production of biomass. The flux of carbon through the CBC is limited by the activity and selectivity of Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase (RuBisCO). Alternative CO2fixation pathways that do not use RuBisCO to fix CO2have evolved in some anaerobic, autotrophic microorganisms. MethodsRather than modifying existing routes of carbon metabolism in plants, we have developed a synthetic carbon fixation cycle that does not exist in nature but is inspired by metabolisms of bacterial autotrophs. In this work, we build and characterize a condensed, reverse tricarboxylic acid (crTCA) cyclein vitroandin planta. ResultsWe demonstrate that a simple, synthetic cycle can be used to fix carbon in vitro under aerobic and mesophilic conditions and that these enzymes retain activity whenexpressed transientlyin planta. We then evaluate stable transgenic lines ofCamelina sativathat have both phenotypic and physiologic changes. TransgenicC. sativaare shorter than controls with increased rates of photosynthetic CO2assimilation and changes in photorespiratory metabolism. DiscussionThis first iteration of a build-test-learn phase of the crTCA cycle provides promising evidence that this pathway can be used to increase photosynthetic capacity in plants.
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- Award ID(s):
- 2414048
- PAR ID:
- 10614247
- Publisher / Repository:
- Frontiers
- Date Published:
- Journal Name:
- Frontiers in Plant Science
- Volume:
- 16
- ISSN:
- 1664-462X
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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