RT Journal Article SR Electronic T1 PEPc-mediated CO2 assimilation provides carbons to gluconeogenesis and the TCA cycle in both dark-exposed and illuminated guard cells JF bioRxiv FD Cold Spring Harbor Laboratory SP 2021.11.03.467183 DO 10.1101/2021.11.03.467183 A1 Valéria F. Lima A1 David B. Medeiros A1 Silvio A. Cândido-Sobrinho A1 Francisco Bruno S. Freire A1 Nicole P. Porto A1 Alexander Erban A1 Joachim Kopka A1 Markus Schwarzländer A1 Alisdair R. Fernie A1 Danilo M. Daloso YR 2021 UL http://biorxiv.org/content/early/2021/11/04/2021.11.03.467183.abstract AB Evidence suggests that guard cells have higher rate of phosphoenolpyruvate carboxylase (PEPc)-mediated dark CO2 assimilation than mesophyll cells. However, it is unknown which metabolic pathways are activated following dark CO2 assimilation in guard cells. Furthermore, it remains unclear how the metabolic fluxes throughout the tricarboxylic acid (TCA) cycle and associated pathways are regulated in illuminated guard cells. Here we used 13C-HCO3 labelling of tobacco guard cells harvested under continuous dark or during the dark-to-light transition to elucidate principles of metabolic dynamics downstream of CO2 assimilation. Most metabolic changes were similar between dark-exposed and illuminated guard cells. However, illumination increased the 13C-enrichment in sugars and metabolites associated to the TCA cycle. Sucrose was labelled in the dark, but light exposure increased the 13C-labelling into this metabolite. Fumarate was strongly labelled under both dark and light conditions, while illumination increased the 13C-enrichment in pyruvate, succinate and glutamate. Only one 13C was incorporated into malate and citrate in either dark or light conditions. Our results collectively suggest that the PEPc-mediated CO2 assimilation provides carbons for gluconeogenesis, the TCA cycle and glutamate synthesis and that previously stored malate and citrate are used to underpin the specific metabolic requirements of illuminated guard cells.Highlight PEPc-mediated CO2 assimilation provides carbons for gluconeogenesis and the TCA cycle, whilst previously stored malate and citrate are used to underpin the specific metabolic requirements of illuminated guard cells.Competing Interest StatementThe authors have declared no competing interest.