A systems biology approach to analyse leaf carbohydrate metabolism in Arabidopsis thaliana

Henkel Sebastian, Nägele Thomas, Hörmiller Imke, Sauter Thomas, Sawodny Oliver, Ederer Michael, Heyer

Description

Plant carbohydrate metabolism comprises numerous metabolite interconversions, some of which form cycles of metabolite degradation and re-synthesis and are thus referred to as futile cycles. In this study, we present a systems biology approach to analyse any possible regulatory principle that operates such futile cycles based on experimental data for sucrose (Scr) cycling in photosynthetically active leaves of the model plant Arabidopsis thaliana . Kinetic parameters of enzymatic steps in Scr cycling were identified by fitting model simulations to experimental data. A statistical analysis of the kinetic parameters and calculated flux rates allowed for estimation of the variability and supported the predictability of the model. A principal component analysis of the parameter results revealed the identifiability of the model parameters. We investigated the stability properties of Scr cycling and found that feedback inhibition of enzymes catalysing metabolite interconversions at different steps of the cycle have differential influence on stability. Applying this observation to futile cycling of Scr in leaf cells points to the enzyme hexokinase as an important regulator, while the step of Scr degradation by invertases appears subordinate.

Sujets

• Systems biology
• Carbohydrate metabolism
• Arabidopsis thaliana

Informations

• Publié par: biomed
• Publié le: 01 janvier 2011
• Nombre de lectures: 5
• Langue: English

Extrait

Henkel et al. EURASIP Journal on Bioinformatics and Systems Biology 2011, 2011:2
http://bsb.eurasipjournals.com/content/2011/1/2
RESEARCH Open Access
A systems biology approach to analyse leaf
carbohydrate metabolism in Arabidopsis thaliana
1† 2*† 2 3 1 1Sebastian Henkel , Thomas Nägele , Imke Hörmiller , Thomas Sauter , Oliver Sawodny , Michael Ederer and
2Arnd G Heyer
Abstract
Plant carbohydrate metabolism comprises numerous metabolite interconversions, some of which form cycles of
metabolite degradation and re-synthesis and are thus referred to as futile cycles. In this study, we present a
systems biology approach to analyse any possible regulatory principle that operates such futile cycles based on
experimental data for sucrose (Scr) cycling in photosynthetically active leaves of the model plant Arabidopsis
thaliana. Kinetic parameters of enzymatic steps in Scr cycling were identified by fitting model simulations to
experimental data. A statistical analysis of the kinetic parameters and calculated flux rates allowed for estimation of
the variability and supported the predictability of the model. A principal component analysis of the parameter
results revealed the identifiability of the model parameters. We investigated the stability properties of Scr cycling
and found that feedback inhibition of enzymes catalysing metabolite interconversions at different steps of the
cycle have differential influence on stability. Applying this observation to futile cycling of Scr in leaf cells points to
the enzyme hexokinase as an important regulator, while the step of Scr degradation by invertases appears
subordinate.
Keywords: Systems biology, carbohydrate metabolism, Arabidopsis thaliana, kinetic modelling, stability analysis,
sucrose cycling
Introduction involving phosphorylation of hexoses (Hex) allows the
Plant metabolic pathways are highly complex, compris- cell to balance deflections of metabolic homeostasis dur-
ing various branch points and crosslinks, and thus ing light-dark cycles.
kinetic modelling turns up as an adequate tool to inves- In this study, we investigate the structural and stability
tigate regulatory principles. Recently, we presented a properties of a model derived from the Scr cycling part
kinetic modelling approach to investigate core reactions of the metabolic pathway described in [1]. Based on the
of primary carbohydrate metabolism in photosyntheti- existing model structure, model parameters were repeat-
cally active leaves of the model plant Arabidopsis thali- edly adjusted in an automated process applying a para-
ana [1] with an emphasis on the physiological role of meter identification algorithm to match the measured
vacuolar invertase, an enzyme that is involved in degra- and simulated data. A method for statistical evaluation
dation of sucrose (Scr). This model was developed in an of the parameters and simulation results is introduced,
iterative process of modelling and validation. A final which allows for the estimation of parameter variability.
parameter set was identified allowing for simulation of Statistical evaluation demonstrates that the same nom-
the main carbohydrate fluxes and interpretation of the inal concentration courses are predicted for different
system behaviour over diurnal cycles. We found that Scr identification runs, while small variability in fluxes and
degradation by vacuolar invertase and re-synthesis larger variability in parameters can be observed. Further,
the parameter identification results were analysed apply-
ing a principal component analysis (PCA). This leads to
* Correspondence: [e-mail] a more extensive investigation with respect to the exten-
† Contributed equally
2 sion and alignment of the parameter values in the para-Biologisches Institut, Abteilung Pflanzenbiotechnologie, Universität Stuttgart,
Pfaffenwaldring 57, D-70550 Stuttgart, Germany meter space. In addition, this allows for conclusions
Full list of author information is available at the end of the article
© 2011 Henkel et al; licensee Springer. This is an Open Access article distributed under the terms of the Creative Commons Attribution
License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium,
provided the original work is properly cited.Henkel et al. EURASIP Journal on Bioinformatics and Systems Biology 2011, 2011:2 Page 2 of 10
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concerning the identifiability of the parameters and the sum of photosynthesis and respiration. Carbon
confirmation that the cost function is sensitive along exchange with the environment and intracellular inter-
parameter combinations. An investigation of structural conversions are linked through the pool of SP. This
stability properties of Scr cycling showed feedback inhi- pool is predominantly constituted by the phosphorylated
bition of Hex on invertase and sugar phosphates (SP) on intermediates glucose-6-phosphate and fructose-6-phos-
hexokinase likely to be involved in stabilisation of the phate. SP can reversibly be converted to St through the
metabolic pathway under consideration. Feedback inhi- reaction v . The reaction v ® represents a set ofSt SP Scr
bition of hexokinase was more efficient in stabilising Scr reactions leading to Scr synthesis. Among them, the
cycling than inhibition of invertase, indicating that, at reaction of Scr phosphate synthase is considered the
this step of the cycle, a superior contribution to stabili- rate-limiting step [2]. Scr can either be exported, for
sation of homeostasis can be achieved. example, by a transport to sinks v ® , or cleavedSP Sinks
intoGlcandFrcbyinvertases, v . The free Hex canInv
The central carbohydrate metabolism in leaves of be phosphorylated by v ® and v ® , respectively.Glc SP Frc SP
A. thaliana These reactions are catalysed by the enzymes glucoki-
Within a 24-h light/dark cycle, two principal modes of nase and fructokinase.
metabolism can be distinguished for plant leaves: photo-
synthesis (day), and respiration (night). During the day, Mathematical model structure
carbon dioxide is taken up, and storage compounds like Time-dependent changes of metabolite concentrations
starch (St) accumulate, while this stock is in part during a diurnal cycle can be described by a system of
respired during the night. Under normal conditions, a ordinary differential equations (ODE). With c being the
certain proportion of carbon is fixed as new plant bio- m-dimensional vector of metabolite concentrations, N
mass. However, typical source leaves as considered here being the m × r stoichiometric matrix and v being the
are mature, and thus carbon use for growth can be r-dimensional vector of fluxes, the biochemical reaction
neglected. Therefore, the carbon balance is completely network can be described as follows:
determined by photosynthesis, respiration and carbon
dc
allocation to associated pathways or heterotrophic tis- (1)=Nv(c,p),
dtsues that are not able to assimilate carbon on their own.
Based on this information and known biochemical reac- with v(c,p) indicating that the fluxes are dependent on
tions, a simplified model structurefortheinterconver- both, metabolite concentrations c and kinetic parameters
sion of central metabolites was created (Figure 1). p. Thus, based on the model structure (Figure 1) of our
The compounds SP, St, Scr, glucose (Glc) and fruc- system, the concentration changes of the five-state vari-
tose (Frc) are derived from photosynthetic carbon fixa- ables: SP, St, sucrose, Glc and Frc are defined as:
tion and linked by interconverting reactions. The flux
1
vCO represents the rate of net photosynthesis, i.e. the ˙c = v −v −v +v +v ,2 SP CO SP→Scr St Glc→SP Frc→SP2
6
c˙ = v ,St St
1 1 (2)
c˙ = ·v − ·v −v ,Scr SP→Scr Scr→Sinks Inv
2 2
c˙ = v −v ,Glc Inv Glc→SP
v vCO ScrSinks2
v c˙ = v −v .SPScr Frc Inv Frc→SP
The stoichiometric coefficients account for the inter-vGlcSP
vInv conversions of species with a different number of carbon
vSt
atoms. For example, the reaction ν ® has a stoichio-SP Scr
vFrcSP metric coefficient value of 1 in the SP state equation,
while in the Scr state equation, this value is 0.5 because
SP contains 6 carbon atoms and Scr contains 12 carbonLeaf Cell
atoms. The stoichiometric coefficients for the reactionEnvironment
catalysed by invertase are 1 in all the respective state
equations because this reaction represents the cleavage
of the disaccharide Scr into two monosaccharides: Glc,Figure 1 Model structure of the central carbohydrate
and Frc. St content is expressed in Glc units, i.e. a car-metabolism in leaves of A. thaliana. SP, sugar phosphates; St,
starch; Scr, sucrose; Glc, glucose; Frc, fructose. v represent rates of bohydrate with six carbon atoms. The rates of the ODE
metabolite interconversion. system (Equation 2) are determined in three ways: byHenkel et al. EURASIP Journal on Bioinformatics and Systems Biology 2011, 2011:2 Page 3 of 10
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measurements (model inputs), carbon balancing and for product inhibition (Equations 6-8) as described in
kinetic rate laws. [3] and [4]:
V (t) ·cmax,Inv ScrModel input and carbon balancing v = ,Inv
c