Strategies for investigating the plant metabolic network with steady-state metabolic flux analysis: lessons from an Arabidopsis cell culture and other systems.

  title={Strategies for investigating the plant metabolic network with steady-state metabolic flux analysis: lessons from an Arabidopsis cell culture and other systems.},
  author={Nicholas J. Kruger and Shyam Kumar Masakapalli and R. George Ratcliffe},
  journal={Journal of experimental botany},
  volume={63 6},
Steady-state (13)C metabolic flux analysis (MFA) is currently the experimental method of choice for generating flux maps of the compartmented network of primary metabolism in heterotrophic and mixotrophic plant tissues. While statistically robust protocols for the application of steady-state MFA to plant tissues have been developed by several research groups, the implementation of the method is still far from routine. The effort required to produce a flux map is more than justified by the… 

Figures from this paper

Fluxes through plant metabolic networks: measurements, predictions, insights and challenges.
The discussion focuses on how a system-wide view of plant metabolism has increased the understanding of network structure, metabolic perturbations and the provision of reducing power and energy for cell function.
Pathways and fluxes: exploring the plant metabolic network.
Current developments in the genomic reconstruction of metabolic networks, the application of flux-balance analysis to such networks, kinetic modelling, and both steady-state-and non-steady state isotope-based measurements of multiple fluxes in the network of central carbon metabolism are described.
Green pathways: Metabolic network analysis of plant systems.
Cell-Type Specific Metabolic Flux Analysis: A Challenge for Metabolic Phenotyping and a Potential Solution in Plants
A protocol for the immunopurification of ectopically expressed green fluorescent protein (GFP) from Arabidopsis thaliana seedlings using a GFP-binding nanobody is developed, and through GC-MS analysis of protein hydrolysates it is established that constitutively expressed GFP reports accurately on the labelling of total protein in root tissues.
The metabolic flux phenotype of heterotrophic Arabidopsis cells reveals a complex response to changes in nitrogen supply.
St steady-state metabolic flux analysis was used to define the metabolic phenotype of a heterotrophic Arabidopsis thaliana cell culture grown in Murashige and Skoog and ammonium-free media, treatments that influenced growth and biomass composition.
Towards high throughput metabolic flux analysis in plants.
Improvements on various sub-steps are proposed which will move flux analysis to the medium-throughput range and closer to established methods such as metabolite profiling, which may enable novel applications of MFA, for example screening plant populations for traits related to the flux phenotype.
The metabolic flux phenotype of heterotrophic Arabidopsis cells reveals a flexible balance between the cytosolic and plastidic contributions to carbohydrate oxidation in response to phosphate limitation.
It is argued that plant cells respond to phosphate deprivation by reconfiguring the flux distribution through the pathways of carbohydrate oxidation to take advantage of better phosphate homeostasis in the plastid.
A detailed view on sulphur metabolism at the cellular and whole-plant level illustrates challenges in metabolite flux analyses.
Sulphur metabolism was taken to illustrate current challenges of metabolic flux analyses and can be expanded to the whole-plant level by considering long-distance transport and, thus, the interaction of roots and the shoot in metabolite fluxes.
Methods and advances in metabolic flux analysis: a mini-review
  • M. Antoniewicz
  • Biology, Engineering
    Journal of Industrial Microbiology & Biotechnology
  • 2015
This mini-review discusses and classify the various methods of MFA that have been developed, which include stoichiometric MFA, 13C metabolic flux analysis, isotopic non-stationary 13Cmetabolic fluxAnalysis, dynamic metabolic fluxanalysis, and 13C dynamic metabolism flux analysis.
Insights into metabolic efficiency from flux analysis.
Insight into the basis of efficiency provided by (13)C-based metabolic flux analysis (MFA) as well as the uses and limitations of efficiency in predictive flux balance analysis (FBA) are highlighted.


Metabolic flux analysis in plants: coping with complexity.
An overview of different MFA approaches, the measurements required to implement them and the information they yield are given, and the challenges that plant metabolism poses for MFA are discussed together with ways that these can be addressed.
Metabolic Network Fluxes in Heterotrophic Arabidopsis Cells: Stability of the Flux Distribution under Different Oxygenation Conditions1[W][OA]
The results add to the emerging picture of the stability of the central metabolic network and its capacity to respond to physiological perturbations with the minimum of rearrangement and highlight a potential difficulty in the interpretation of metabolomic data.
Subcellular Flux Analysis of Central Metabolism in a Heterotrophic Arabidopsis Cell Suspension Using Steady-State Stable Isotope Labeling1[W][OA]
Fluxes through the pathways of central metabolism in a heterotrophic Arabidopsis cell suspension culture were deduced from the redistribution of label in steady-state 13C-labeling experiments, highlighting the need for caution when predicting flux through metabolic networks using assumptions based on the energetics of resource utilization.
Quantitative approaches for analysing fluxes through plant metabolic networks using NMR and stable isotope labelling
It is concluded that metabolic flux analysis provides a powerful method for defining the metabolic phenotype of wild type, mutant and transgenic plants and that its development should be pursued.
Understanding flux in plant metabolic networks.
Strategies for metabolic flux analysis in plants using isotope labelling.
Metabolic flux analysis in plants: from intelligent design to rational engineering.
This review brings together the recent developments in quantitative MFA and predictive modeling and the application of predictive tools to high quality flux maps in particular promises to be important in the rational metabolic engineering of plants.