Metabolic regulation of malic enzyme of two Panicoideae clade members, Setaria italica and Panicum virgatum

GERRARD WHEELER, M. C.; ANDREO, C. S.; CALACE, P.; SAIGO, M.

Salta

Congreso; LV Reunión Anual SAIB XIV PABMB Congress; 2019

SAIB - PABMB

C4 photosynthesis has evolved to increase the photosynthetic efficiency in conditions where photorespiration would be enhanced, such as high temperatures and drought. This was achieved through the introduction of a series of anatomical and biochemical features that allow the concentration of CO2 around Rubisco. In most C4 species, this is done through compartmentalizing the initial and definitive CO2 assimilation processes into two discrete cell types, namely mesophyll (M) and bundle sheath (BS) cells. As part of this mechanism a compound of four carbon atoms is transported from M to BS cells. Setaria italica (foxtail millet) and Panicum virgatum (switchgrass) are two members of the Panicoideae clade used for feed and biofuel production. Despite their evolutionary closeness, they have been classified into different C4 subtypes. S. italica belong to the C4-NADP-ME subtype as it posseses a NADP-dependent malic enzyme (NADP-ME) as major malate decarboxylase within chloroplasts and transport mainly malate from MC to BSC. In P. virgatum (C4-NAD-ME subtype) the major decarboxylase is a mitocondrial NAD-ME and aspartate is the main C4 acid transported to BSC from MC. In this study we made progresses in the kinetic characterization that distinguishes each decarboxylase enzyme operating in both grasses. Furthemore, we carried out a comparative analysis in order to identify the isoforms involved in the C4 cycle and to characterize the differences between the photosynthetic and the non-photosynthetic versions. We purified recombinant NAD-MEs (α and β) and cloroplastic NADP-MEs from both species. To gain insight into the physiological roles of NAD and NADP- MEs, the activity of recombinant enzymes in the malate decarboxylation direction was evaluated in the presence of different metabolites. The general analysis of the NAD-MEs responses showed some similarities with those reported for the enzymes from Arabidopsis, the C3 model species. The NAD-ME type β (Seita.2G322000 and Pavir.2KG446000) responded positively to the presence of Krebs cycle intermediates such as citrate, fumarate, succinate and oxaloacetate and the NAD-ME type α (Seita.9G200600 and Pavir.9KG132400) were strongly activated by both acetyl-CoA and CoA and by fructose-1,6-bP and PEP, both metabolites markers of the decrease in pyruvate glycolytic supply. These regulations agree with the functioning of NAD-ME as a pyruvate-contributing enzyme when there is a decrease in the glycolysis and a high content of the Krebs cycle intermediates. On the other hand, both NADP-MEs showed inhibition by the substrate malate at pH 7, a regulatory characteristic linked to the photosynthetic function. In addition, the ATP activation and the aspartate inhibition exhibited by NADP-ME from P. virgatum would indicate that in this NAD-ME subtype species the NADP-ME decarboxylation could also contribute to the C4 photosynthesis.