Photosynthetic activity triggers pH and NAD redox signatures across different plant cell compartments

Abstract

A characteristic feature of most plants is their ability to perform photosynthesis, which ultimately provides energy and organic substrates to most life. Photosynthesis dominates chloroplast physiology but represents only a fraction of the tightly interconnected metabolic network that spans the entire cell. Here, we explore how photosynthetic activity affects the energy physiological status in cell compartments beyond the chloroplast. We develop precision live monitoring of subcellular energy physiology under illumination to investigate pH, MgATP2- and NADH/NAD+ dynamics at dark-light transitions by confocal imaging of genetically encoded fluorescent protein biosensors in Arabidopsis thaliana leaf mesophyll. We resolve the in vivo signature of stromal alkalinisation resulting from photosynthetic proton pumping and observe a similar pH signature also in the cytosol and the mitochondria suggesting that photosynthesis triggers an 'alkalinisation wave' that affects the pH landscape of large parts of the cell. MgATP2- increases in the stroma at illumination, but no major effects on MgATP2- concentrations in the cytosol were resolved. Photosynthetic activity triggers a signature of substantial NAD reduction in the cytosol that is driven by photosynthesis-derived electron export. Strikingly, cytosolic NAD redox status was deregulated in mutants of chloroplastic NADP- and mitochondrial NAD-dependent malate dehydrogenases even at darkness, pinpointing the participation of the chloroplasts and mitochondria in shaping cytosolic redox metabolism in vivo with a dominant function of malate metabolism. Our data illustrate how profoundly and rapidly changes in photosynthetic activity affect the physiological and metabolic landscape throughout green plant cells.

Effects of chloroplast–cytoplasm exchange and lateral mass transfer on slow induction of chlorophyll fluorescence in Characeae

“…These data suggest that the export of photometabolites by source chloroplasts starts much earlier after the onset of illumination than the metabolite accumulation by recipient chloroplasts. According to Elsässer et al (2020), the accumulation of cytoplasmic NADH in Arabidopsis mesophyll begins within less than 30 s of light exposure.…”

Measuring ROS and redox markers in plant cells

“…with an NADH binding affinity of 1.2 mM and 31.4 mM respectively (in the presence of 500 mM NAD + ). 159 Peredox has already been applied to explore the role of the nucleotide transporter NDT in stomatal development, 160 the importance of malate dehydrogenase for redox shuttling during photosynthesis 161 and a possible link between pathogen elicitation and the cytosolic NADH : NAD + ratio. 159 SoNar and iNap have been used together with subcellular pH and ATP sensors to investigate photorespiration, supporting the hypothesis that reductant in excess of the capacity of the mitochondrial electron transport chain is produced during photosynthesis.…”

cROStalk for Life: Uncovering ROS Signaling in Plants and Animal Systems, from Gametogenesis to Early Embryonic Development

“…For instance, recent studies with NADH sensors [ 74 , 75 ] reveal that (i) there are large variations in the NADH/NAD + ratio among tissues and conditions (for instance, upon sugar supplementation, elicitor exposure, and illumination of seedlings/leaves); and (ii) inhibiting plastidial or mitochondrial ETCs is quite readily reverberated in a change of the ratio in the cytosol, either directly through external NADH dehydrogenases or indirectly through the various metabolite shuttles. A parallel analysis of cytosolic NADH/NAD + , ATP, and pH revealed that there is a rapid transfer of reducing equivalents and protons, but not of ATP, from the chloroplast to cytosol upon illumination [ 76 ]. Altering the capacity of the malate valves by knocking out the mitochondrial malate dehydrogenases (mMDH1 or mMDH2), or the NADP + -dependent plastidic isoform (cpNADP-MDH), did not change the capacity of exporting reducing equivalents or its kinetics in the light.…”

“…Altering the capacity of the malate valves by knocking out the mitochondrial malate dehydrogenases (mMDH1 or mMDH2), or the NADP + -dependent plastidic isoform (cpNADP-MDH), did not change the capacity of exporting reducing equivalents or its kinetics in the light. However, the cytosolic NAD pool was more reduced in the dark in all three mutants [ 76 ]. It is easy to understand how a reduction in capacity for malate oxidation in mitochondria may lead to a higher NADH accumulation in the cytosol.…”