Suppression of metabolite shuttles for export of chloroplast and mitochondrial ATP and NADPH increases the cytosolic NADH:NAD+ ratio in tobacco leaves in the dark

The communication between chloroplasts and mitochondria, which depends on the inter-organellar exchange of carbon skeletons, energy, and reducing equivalents, is essential for maintaining efficient respiratory metabolism and photosynthesis. We devised a multi-transgene approach to manipulate the lea...

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Published in:Journal of plant physiology 2022-01, Vol.268, p.153578-153578, Article 153578
Main Authors: Moreno-García, Beatriz, López-Calcagno, Patricia E., Raines, Christine A., Sweetlove, Lee J.
Format: Article
Language:English
Subjects:
Adenosine Triphosphate - metabolism
Arabidopsis
Arabidopsis - genetics
Arabidopsis - metabolism
Biosensors
Chlorophyll
Chloroplasts
Chloroplasts - metabolism
Darkness
Dehydrogenase
Dehydrogenases
Energy balance
Equivalence
Fluorescence
Glyceraldehyde
Glyceraldehyde 3-phosphate
Glyceraldehyde-3-phosphate dehydrogenase
Greenhouses
Ketoglutaric acid
Leaves
Malate dehydrogenase
Malates - metabolism
Metabolism
Metabolites
Mitochondria
Mitochondria - metabolism
Multi-transgene transformation
NAD
NAD - metabolism
NADH
NADP - metabolism
Nicotiana - metabolism
Nicotiana tabacum
Nicotinamide adenine dinucleotide
Organelle communication
Oxidation-Reduction
Photosynthesis
Plant growth
Plant Leaves - metabolism
Plant metabolic engineering
Principal components analysis
Redox metabolism
Tobacco
Transcription
Transgenic plants
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Summary:The communication between chloroplasts and mitochondria, which depends on the inter-organellar exchange of carbon skeletons, energy, and reducing equivalents, is essential for maintaining efficient respiratory metabolism and photosynthesis. We devised a multi-transgene approach to manipulate the leaf energy and redox balance in tobacco (Nicotiana tabacum) while monitoring the in vivo cytosolic redox status of NAD(H) using the biosensor c-Peredox-mCherry. Our strategy involved altering the shuttling capacity of the chloroplast by (1) increasing the chloroplast malate valve capacity by overexpression of the chloroplast malate valve transporter pOMT from Arabidopsis (AtpOMT1) while (2) reducing the activity of the chloroplast triose-phosphate/3-phosphoglycerate shuttle by knocking down the cytosolic NAD-dependent glyceraldehyde 3-phosphate dehydrogenase (NtGAPC). This was accompanied by (3) alterations to the export of reducing equivalents in the mitochondria by knocking down the mitochondrial malate dehydrogenase (NtmMDH) and (4) an increased expression of the mitochondrial fission regulator FIS1A from Arabidopsis (AtFIS1A). The multi-transgene tobacco plants were analysed in glasshouse conditions and showed significant increases in the cytosolic NADH:NAD+ in the dark when transcript levels for NtGAPC or NtmMDH were knocked down. In addition, principal component analysis and Spearman correlation analyses showed negative correlations between average transcript levels for the gene targets and parameters related to chlorophyll fluorescence and plant growth. Our results highlight the importance of the shuttling of energy and reducing equivalents from chloroplasts and mitochondria to support photosynthesis and growth and suggest an important role for the dual 2-oxoglutarate/malate and oxaloacetate/malate transporter (pOMT). •A multi-transgene construct targeted tobacco leaf redox and energy metabolism.•Average transcript levels correlated negatively with plant growth and photosynthesis.•Overexpression of chloroplast malate valve transporter pOMT could explain correlations.•The redox biosensor c-Peredox-mCherry was used to report cytosolic NADH/NAD+ ratios.•Defects in mitochondrial and chloroplasts shuttles increase dark NADH/NAD+ ratios.
ISSN:0176-1617
1618-1328
DOI:10.1016/j.jplph.2021.153578