Metabolic Modelling
We adapted a metabolic model by Arnold and Nikoloski (2014) to allow for
the diurnal accumulation of carbon compounds. Specifically, we ensured
that cytosolic fumarate could be produced from cytosolic malate and
added “export reactions” to the model (describing diurnal storage
pools) for malate, fumarate and starch in addition to the already
existing sucrose export. We generated four models: Wild type Col-0 in
20oC and 4oC conditions andfum2 in 20oC and 4oC
conditions. We constrained the models using metabolite assays such that
the beginning of day concentrations of fumarate, malate, and starch
subtracted from their respective end of day concentrations equated to
the diurnal flux over the eight-hour photoperiod. Furthermore, we
assumed a constant rate of photosynthesis (Dyson et al. , 2016)
throughout the day and converted the measured rates of photosynthesis to
diurnal fluxes of carbon intake. We then used proteomics data to further
constrain the upper bounds of the flux reactions (Ramon, Gollub, &
Stelling, 2018). For each metabolic reaction we checked whether all of
the corresponding proteins were available in the data set; if so, then
those reactions were given an upper bounds of additive value of all of
the identified proteins in case multiple isoforms exist. Given that the
proteomics data is relative and not quantitative we scaled all of the
proteomics constraints such that we were able to obtain model solutions
across all four models. We used single flux solution, from a flux
balance analysis maximizing carbon assimilation via Rubisco within
feasible model constraints, in order to eliminate non-essential
reactions which generate loops within the model, using the loopless
function in the cobra (version 0.10.1) package. We then conducted
flux sampling using the CHRR algorithm in the MATLAB toolbox as outlined
in Herrmann, Dyson, Vass, Johnson, and Schwartz (2019).
All models, code and data used to conduct the computational analyses are
available at https://github.com/HAHerrmann/FluxSamplingCol0Fum2 and have
been archived in Zenodo (DOI: 10.5281/zenodo.3366934).