Dynamic metabolic flux analysis of plant cell wall synthesis

highlights
• A dynamic metabolic flux model was developed based on pulse labeling experiments.
• Results map carbon flow from sucrose to plant cell wall precursors.
• Sucrose invertase controls the entry of sucrose into Arabidopsis heterotrophic metabolism.
• UDP-Glucose-4-epimerase plays a dominant role in plant UDP-Gal synthesis.
• Candidates for engineering cell wall synthesis rates and composition are identified.

The regulation of plant cell wall synthesis pathways remains poorly understood. This has become a bottleneck in designing bioenergy crops. The goal of this study was to analyze the regulation of plant cell wall precursor metabolism using metabolic flux analysis based on dynamic labeling experiments. Arabidopsis T87 cells were cultured heterotrophically with 13C labeled sucrose. The time course of 13C labeling patterns in cell wall precursors and related sugar phosphates was monitored using liquid chromatography tandem mass spectrometry until steady state labeling was reached. A kinetic model based on mass action reaction mechanisms was developed to simulate the carbon flow in the cell wall synthesis network. The kinetic parameters of the model were determined by fitting the model to the labeling time course data, cell wall composition, and synthesis rates. A metabolic control analysis was performed to predict metabolic regulations that may improve plant biomass composition for biofuel production. Our results describe the routes and rates of carbon flow from sucrose to cell wall precursors. We found that sucrose invertase is responsible for the entry of sucrose into metabolism and UDP-glucose-4-epimerase plays a dominant role in UDP-Gal synthesis in heterotrophic Aradidopsis cells under aerobic conditions. We also predicted reactions that exert strong regulatory influence over carbon flow to cell wall synthesis and its composition.