C4GEM, a genome-scale metabolic model to study C4 plant metabolism

Abstract

Leaves of C(4) grasses (such as maize [Zea mays], sugarcane [Saccharum officinarum], and sorghum [Sorghum bicolor]) form a classical Kranz leaf anatomy. Unlike C(3) plants, where photosynthetic CO(2) fixation proceeds in the mesophyll (M), the fixation process in C(4) plants is distributed between two cell types, the M cell and the bundle sheath (BS) cell. Here, we develop a C(4) genome-scale model (C4GEM) for the investigation of flux distribution in M and BS cells during C(4) photosynthesis. C4GEM, to our knowledge, is the first large-scale metabolic model that encapsulates metabolic interactions between two different cell types. C4GEM is based on the Arabidopsis (Arabidopsis thaliana) model (AraGEM) but has been extended by adding reactions and transporters responsible to represent three different C(4) subtypes (NADP-ME [for malic enzyme], NAD-ME, and phosphoenolpyruvate carboxykinase). C4GEM has been validated for its ability to synthesize 47 biomass components and consists of 1,588 unique reactions, 1,755 metabolites, 83 interorganelle transporters, and 29 external transporters (including transport through plasmodesmata). Reactions in the common C(4) model have been associated with well-annotated C(4) species (NADP-ME subtypes): 3,557 genes in sorghum, 11,623 genes in maize, and 3,881 genes in sugarcane. The number of essential reactions not assigned to genes is 131, 135, and 156 in sorghum, maize, and sugarcane, respectively. Flux balance analysis was used to assess the metabolic activity in M and BS cells during C(4) photosynthesis. Our simulations were consistent with chloroplast proteomic studies, and C4GEM predicted the classical C(4) photosynthesis pathway and its major effect in organelle function in M and BS. The model also highlights differences in metabolic activities around photosystem I and photosystem II for three different C(4) subtypes. Effects of CO(2) leakage were also explored. C4GEM is a viable framework for in silico analysis of cell cooperation between M and BS cells during photosynthesis and can be used to explore C(4) plant metabolism.

Figure 1.

Schematic representation of the photosynthetic metabolism of three C₄ subtypes distinguished according to the decarboxylating enzyme. Numbers refer to enzymes: (1) PEPC, (2) NADP-malate dehydrogenase, (3) NADP-ME, (4) pyruvate-Pi dikinase, (5) Rubisco, (6) PCK, (7) Ala aminotransferase, (8) Asp aminotransferase, (9) NAD-malate dehydrogenase, (10) NAD-ME. Some steps were hidden for the sake of simplicity. 3PGA, 3-Phosphoglycerate; DHAP, dihydroxyacetonephosphate; OAA, oxaloacetate; RuBP, ribulose-1,5-bisphosphate.

Figure 2.

Elements of the reconstructed C4GEM network and the two-tissue model. ORFs, Open reading frames.

Figure 3.

Flux map of the central metabolism. Relative flux distributions (BS/M) are shown between BS and M during C₄ photosynthesis. Green and red lines represent increased and decreased flux in BS, respectively. Gray lines represent flux values that do not differ significantly between cells.

Figure 4.

Flux simulation for each C₄ subtype with no leakage of CO₂ and considering 50% of CO₂ leakage out of BS. LET, Linear electron transport.

Figure 5.

C₄ genome-scale model reconstruction process.