ABSTRACT
Ascorbic acid (AA) is a major redox buffer in plant cells. The role of ethylene in the redox signaling pathways that influence photosynthesis and growth was explored in two independent AA deficient Arabidopsis thaliana mutants (vtc2-1 and vtc2-4). Both mutants, which are defective in the AA biosynthesis gene GDP-L-galactose phosphorylase, produce higher amounts of ethylene than wt plants. In contrast to the wt, the inhibition of ethylene signaling increased leaf conductance, photosynthesis and dry weight in both vtc2 mutant lines. The AA-deficient mutants showed altered expression of genes encoding proteins involved in the synthesis/responses to phytohormones that control growth, particularly auxin, cytokinins, abscisic acid, brassinosterioids, ethylene and salicylic acid. These results demonstrate that AA deficiency modifies hormone signaling in plants, redox-ethylene interactions providing a regulatory node controlling shoot biomass accumulation.
Subject(s)
Arabidopsis Proteins/genetics , Ascorbic Acid/metabolism , Ethylenes/metabolism , Phosphoric Monoester Hydrolases/genetics , Plant Growth Regulators/metabolism , Abscisic Acid/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , Ascorbic Acid/genetics , Biomass , Gene Expression Regulation, Plant , Mutation , Oxidation-Reduction , Phosphoric Monoester Hydrolases/metabolism , Photosynthesis/genetics , Plant Growth Regulators/genetics , Salicylic Acid/metabolism , Signal Transduction/geneticsABSTRACT
Ascorbic acid (AA) is synthesized in plant mitochondria through the oxidation of l-galactono-1,4-lactone (l-GalL) and then distributed to different cell compartments. AA-deficient Arabidopsis thaliana mutants (vtc2) and exogenous applications of l-GalL were used to generate plants with different AA content in their leaves. This experimental approach allows determining specific AA-dependent effects on carbon metabolism. No differences in O2 uptake, malic and citric acid and NADH content suggest that AA synthesis or accumulation did not affect mitochondrial activity; however, l-GalL treatment increased CO2 assimilation and photosynthetic electron transport rate in vtc2 (but not wt) leaves demonstrating a stimulation of photosynthesis after l-GalL treatment. Increased CO2 assimilation correlated with increased leaf stomatal conductance observed in l-GalL-treated vtc2 plants.
Subject(s)
Arabidopsis/physiology , Ascorbic Acid/biosynthesis , Mitochondria/metabolism , Photosynthesis , Arabidopsis/drug effects , Cell Respiration/drug effects , Dehydroascorbic Acid/metabolism , Glutathione/metabolism , Lactones/pharmacology , Mitochondria/drug effects , Photosynthesis/drug effects , Plant Stomata/drug effects , Plant Stomata/physiology , Pyridines/pharmacology , Ribulose-Bisphosphate Carboxylase/metabolism , Sugar Acids/pharmacologyABSTRACT
Ascorbic acid is synthesized from galactono-gamma-lactone (GL) in plant tissues. An improved extraction procedure involving ammonium sulfate precipitation of membrane proteins from crude leaf homogenates yielded a simple, quick method for determining tissue activities of galactono-gamma-lactone dehydrogenase (GLDH). Total foliar ascorbate and GLDH activity decreased with leaf age. Subcellular fractionation experiments using marker enzymes demonstrated that 80% of the total GLDH activity was located on the inner mitochondrial membrane, and 20% in the microsomal fraction. Specific antibody raised against potato (Solanum tuberosum L.) tuber GLDH recognized a 56-kD polypeptide in extracts from the mitochondrial membranes but failed to detect the equivalent polypeptide in microsomes. We demonstrate that isolated intact mitochondria synthesize ascorbate in the presence of GL. GL stimulated mitochondrial electron transport rates. The respiration inhibitor antimycin A stimulated ascorbate biosynthesis, while cyanide inhibited both respiration and ascorbate production. GL-dependent oxygen uptake was observed in isolated intact mitochondria. This evidence suggests that GLDH delivers electrons to the mitochondrial electron transport chain between complexes III and IV.