Metabolism of Organic Acids, Nitrogen and Amino Acids in Chlorotic Leaves of ‘Honeycrisp' Apple with Excessive Accumulation of Carbohydrates

Metabolite profiles and activities of key enzymes in the metabolism of organic acids, nitrogen and amino acids were compared between chlorotic leaves and normal leaves of ‘Honeycrisp’ apple to understand how accumulation of non-structural carbohydrates affects the metabolism of organic acids, nitrogen and amino acids. Higher concentrations of non-structural carbohydrates, particularly starch, fructose, sucrose, maltose, maltitol, and trehalose were accumulated in chlorotic leaves, and correspondingly much lower CO₂ assimilation was observed in chlorotic leaves than in normal leaves, confirming feedback inhibition of photosynthesis in chlorotic leaves. Dark respiration and activities of several key enzymes in glycolysis and tricarboxylic acid (TCA) cycle, ATP-phosphofructokinase, pyruvate kinase, citrate synthase, aconitase and isocitrate dehydrogenase, were significantly higher in chlorotic leaves than in normal leaves. However, concentrations of most organic acids including pyruvate, phosphoenolpyruvate (PEP), oxaloacetate, 2-oxoglutarate, malate and fumarate, and activities of key enzymes involved in the anapleurotic pathway including PEP carboxylase, NAD-malate dehydrogenase and NAD-malic enzyme were significantly lower in chlorotic leaves than in normal leaves. Concentrations of total soluble proteins and most free amino acids were significantly lower in chlorotic leaves than in normal leaves. Activities of key enzymes in nitrogen assimilation and amino acid synthesis, including nitrate reductase, glutamine synthetase, ferredoxin and NADH-dependent glutamate synthase, and glutamate pyruvate transaminase were significantly lower whereas activities of glutamate dehydrogenase and glutamate oxaloacetate transaminase were significantly higher in chlorotic leaves than in normal leaves. It was concluded that, in response to excessive accumulation of non-structural carbohydrates, glycolysis and TCA cycle are up-regulated to “consume” the excess carbon available whereas the anapleurotic pathway, nitrogen assimilation and amino acid synthesis are down-regulated to reduce the overall rate of amino acid and protein synthesis.