Organic Acid Metabolism and Nitrogen Metabolism In the Leaves of Transgenic Apple Trees with Decreased Sorbitol Synthesis

Sorbitol is a main photosynthetic end-product and a primary translocated form of carbon in apple and many other tree fruits of the Rosaceae family. Sorbitol synthesis shares the same hexose-phosphate pool with sucrose synthesis in the cytosol of source leaves and the reaction catalyzed by aldose-6-phosphate (A6PR) is the limiting step. When ‘Greensleeves’ apple was transformed with the cDNA of A6PR in the antisense orientation, A6PR activity in mature leaves was decreased to approximately 15–30% of the untransformed control, and consequently sorbitol synthesis was significantly decreased. As a result, both glucose 6-phosphate and fructose 6-phosphate accumulated in the cytosol at the expense of inorganic phosphate, leading to up-regulation of starch synthesis without altering CO₂ assimilation. The objective of this study was to determine how the metabolism of organic acids and amino acids responds to decreased sorbitol synthesis in the leaves of the transgenic trees. The leaves of the transgenic plants were found to have higher concentrations of phosphoenolpyruvate, pyruvate, citrate, 2-oxoglutarate, succinate and oxaloacetate, higher activities of key enzymes in glycolysis and TCA cycle such as ATP-phosphofructokinase, PPi- phosphofructokinase, phosphoenolpyruvate carboxylase, pyruvate kinase, aconitase, isocitrate dehydrogenase and NAD-malic enzyme and higher concentrations of 12 amino acids (e.g. Asp, Asn, Glu, Gln, Ser, etc.) out of the 20 free amino acids measured. Additionally, when the detached leaves of the untransformed control were fed with 10 mM mannose to lower the inorganic phosphate pool, some key enzymes in organic acid metabolism such as phosphoenolpyruvate carboxylase, ATP-phosphofructokinase, PPi-phosphofructokinase, isocitrate dehydrogenase, NAD-malic enzyme showed a similar trend as found in the transgenic plants. These results support our hypothesis that both organic acid metabolism and amino acid metabolism are up-regulated in the transgenic plants with decreased sorbitol synthesis.