METABOLIC REGULATION OF MALIC ENZYMES OF TWO PANICOIDEAE CLADE MEMBERS, SETARIA ITALICA AND PANICUM VIRGATUM

CALACE, PAULA; ANDREO, CARLOS SANTIAGO; GERRARD WHEELER, MARIEL CLAUDIA; SAIGO, MARIANA

Congreso; SAIB LV-XIV PABMB; 2019

C4 photosynthesis has evolved to increase the photosynthetic efficiency in conditions where photorespiration would be enhanced, such as hightemperatures and drought. This was achieved through the introduction of a series of anatomical and biochemical features that allow theconcentration of CO2 around Rubisco. In most C4 species, this is done through compartmentalizing the initial and definitive CO2 assimilationprocesses into two discrete cell types, namely mesophyll (M) and bundle sheath (BS) cells. As part of this mechanism a compound of fourcarbon atoms is transported from M to BS cells. Setaria italica (foxtail millet) and Panicum virgatum (switchgrass) are two members of thePanicoideae clade used for feed and biofuel production. Despite their evolutionary closeness, they have been classified into different C4subtypes. S. italica belong to the C4-NADP-ME subtype as it posseses a NADP-dependent malic enzyme (NADP-ME) as major malatedecarboxylase within chloroplasts and transport mainly malate from MC to BSC. In P. virgatum (C4-NAD-ME subtype) the majordecarboxylase is a mitocondrial NAD-ME and aspartate is the main C4 acid transported to BSC from MC. In this study we made progresses inthe kinetic characterization that distinguishes each decarboxylase enzyme operating in both grasses. Furthemore, we carried out a comparativeanalysis in order to identify the isoforms involved in the C4 cycle and to characterize the differences between the photosynthetic and the nonphotosyntheticversions. We purified recombinant NAD-MEs (α and β) and cloroplastic NADP-MEs from both species. To gain insight into thephysiological roles of NAD and NADP- MEs, the activity of recombinant enzymes in the malate decarboxylation direction was evaluated in thepresence of different metabolites. The general analysis of the NAD-MEs responses showed some similarities with those reported for the enzymesfrom Arabidopsis, the C3 model species. The NAD-ME type β (Seita.2G322000 and Pavir.2KG446000) responded positively to the presence ofKrebs cycle intermediates such as citrate, fumarate, succinate and oxaloacetate and the NAD-ME type α (Seita.9G200600 andPavir.9KG132400) were strongly activated by both acetyl-CoA and CoA and by fructose-1,6-bP and PEP, both metabolites markers of thedecrease in pyruvate glycolytic supply. These regulations agree with the functioning of NAD-ME as a pyruvate-contributing enzyme when thereis a decrease in the glycolysis and a high content of the Krebs cycle intermediates. On the other hand, both NADP-MEs showed inhibition by thesubstrate malate at pH 7, a regulatory characteristic linked to the photosynthetic function. In addition, the ATP activation and the aspartateinhibition exhibited by NADP-ME from P. virgatum would indicate that in this NAD-ME subtype species the NADP-ME decarboxylation couldalso contribute to the C4 photosynthesis.