SE162:/S1/M1

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Sample Set Information

ID TSE1323
Title A U-system approach for predicting metabolic behaviors and responses based on an alleged metabolic reaction network.
Description Background

Progress in systems biology offers sophisticated approaches toward a comprehensive understanding of biological systems. Yet, computational analyses are held back due to difficulties in determining suitable model parameter values from experimental data which naturally are subject to biological fluctuations. The data may also be corrupted by experimental uncertainties and sometimes do not contain all information regarding variables that cannot be measured for technical reasons.

Results
We show here a streamlined approach for the construction of a coarse model that allows us to set up dynamic models with minimal input information. The approach uses a hybrid between a pure mass action system and a generalized mass action (GMA) system in the framework of biochemical systems theory (BST) with rate constants of 1, normal kinetic orders of 1, and -0.5 and 0.5 for inhibitory and activating effects, named Unity (U)-system. The U-system model does not necessarily fit all data well but is often sufficient for predicting metabolic behavior of metabolites which cannot be simultaneously measured, identifying inconsistencies between experimental data and the assumed underlying pathway structure, as well as predicting system responses to a modification of gene or enzyme. The U-system approach was validated with small, generic systems and implemented to model a large-scale metabolic reaction network of a higher plant, Arabidopsis. The dynamic behaviors obtained by predictive simulations agreed with actually available metabolomic time-series data, identified probable errors in the experimental datasets, and estimated probable behavior of unmeasurable metabolites in a qualitative manner. The model could also predict metabolic responses of Arabidopsis with altered network structures due to genetic modification.

Conclusions
The U-system approach can effectively predict metabolic behaviors and responses based on structures of an alleged metabolic reaction network. Thus, it can be a useful first-line tool of data analysis, model diagnostics and aid the design of next-step experiments.

Authors Sriyudthsak K, Sawada Y, Chiba Y, Yamashita Y, Kanaya S, Onouchi H, Fujiwara T, Naito S, Voit EO, Shiraishi F, Hirai MY.
Reference BMC Syst Biol. 2014;8 Suppl 5:S4. doi: 10.1186/1752-0509-8-S5-S4. Epub 2014 Dec 12.
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Sample Information

ID S1
Title Arabidopsis callus
Organism - Scientific Name Arabidopsis thaliana
Organism - ID NCBI taxonomy:3702
Compound - ID
Compound - Source
Preparation Arabidopsis thaliana liquid callus culture derived from accession Col-0 was prepared as described in Murota et al. with slight modifications. For callus induction, minced seedlings were incubated in RM28 medium under constant light. The medium was changed every 6 days. For a metabolic perturbation experiment, RM28 medium supplemented with 10 mM L-lysine and 1 mM L-threonine was used at the third medium change. For a control experiment, RM28 without supplementation was used. Sucrose in RM28 medium was a sole carbon source for callus culture. The experiments were carried out in triplicate.

For both metabolome and amino acid analyses, calli were collected prior to lysine and threonine treatment (0 h), and 2, 6, 12, 24, 36, 48, 60, 72, 84 and 96 h after the treatment. The calli were immediately frozen in liquid nitrogen and stored at -80°C. Prior to analyses, the frozen samples were lyophilized using a freeze dryer (FDU-2100, EYELA) in a vacuum.

Sample Preparation Details ID
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Analytical Method Information

ID M1
Title Metabolome analysis (LC-MS)
Method Details ID MS1
Sample Amount 1 μL
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Analytical Method Details Information

ID MS1
Title Metabolome analysis (LC-MS)
Instrument UPLC-TQD system (Waters)
Instrument Type UPLC-QTOF-MS
Ionization ESI
Ion Mode positive and negative
Description Metabolites were extracted by homogenizing lyophilized callus in 500 uL 80% methanol in 0.1% formic acid per 2 mg dry weight callus with 5 mm zirconia beads (no.5-4060-13, AS ONE Co. Ltd.) in 2.0 mL sampling tubes (no.132-620C, WATSON Co., LTD) for 5 min using shake master NEO (Bio Medical Science, Tokyo, Japan). After centrifugation using a high speed refrigerated micro centrifuge (TOMY MX-300) at 14,000 r.p.m. at 4°C, 250 uL supernatant was dried up in 96 well plate and the residue was dissolved in 120 uL ultrapure water (no. 210-01303, Wako Pure Chemical Industries, Ltd.). One uL of the solution was subjected to widely targeted metabolome analysis by LC-MS using UPLC-TQD system (Waters, Milford, MA, USA).
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