Effects of Altered Fructose 2,6-Bisphosphate Levels on Carbohydrate Metabolism in Carnation

The aim of this work was to examine the role of fructose 2,6-bisphosphate (fru 2,6P2) in the carbohydrate metabolism in carnation (Dianthus caryophyllus L.). For this purpose, transgenic plants harboring two modified bifunctional enzyme complementary DNAs of rat liver origin (6-phosphofructo-2-kinase/fructose 2,6-biphosphatase) were generated. Transformation with the kinase construct resulted in a 45% to 85% increase in fru 2,6P2 concentrations compared with the wild type. Transformation with the phosphatase construct reduced the fru 2,6P2 contents by 45% and 70%. These alterations in fru 2,6P2 amounts affected the key enzyme activities of sucrose and starch metabolism. Accordingly, plants with elevated fru 2,6P2 concentrations had high levels of starch, fructose, and triose phosphates, and low levels of sucrose, glucose, and hexose phosphates. In plants with reduced amounts of fru 2,6P2 different results could be observed in major carbohydrate compounds.

Mechanism of Up-regulation of Starch Synthesis in Mature Leaves of Transgenic Apple Trees with Decreased Sorbitol Synthesis

The reaction catalyzed by ADP-glucose pyrophosphorylase (AGPase) to form ADP-glucose is a regulatory and rate-limiting step in starch synthesis in plants. In response to decreased sorbitol synthesis, starch synthesis was up-regulated in the transgenic apple plants. In this study, we examined both redox and metabolite regulation of AGPase to understand the mechanism responsible for the up-regulation of starch synthesis. No difference in the monomerization/dimerization of apple leaf AGPase small subunits was observed between the transgenic plants and the untransformed control. NADP-dependent malate dehydrogenase, indicative of chloroplastic redox status, did not show significant change in the transgenic plants either. Determination of key metabolites with nonaqueous fractionation indicated that concentrations of hexose phosphates (mainly glucose-6-phosphate and fructose-6-phosphate) were higher in both the cytosol and chloroplasts of the transgenic plants than in the control, whereas 3-phosphoglycerate (PGA) concentration in the chloroplast was not higher in the transgenic plants. We conclude that accumulation of hexose-phosphates results in a decrease in inorganic phosphate (Pi) concentration and an increase in PGA/Pi ratio in the chloroplast, leading to up-regulation of starch synthesis via activating AGPase.

Control of glycolysis in contracting skeletal muscle. II. Turning it off

Glycolytic flux in muscle declines rapidly after exercise stops, indicating that muscle activation is a key controller of glycolysis. The mechanism underlying this control could be 1) a Ca2+-mediated modulation of glycogenolysis, which supplies substrate (hexose phosphates, HP) to the glycolytic pathway, or 2) a direct effect on glycolytic enzymes. To distinguish between these possibilities, HP levels were raised by voluntary 1-Hz exercise, and glycolytic flux was measured after the exercise ceased. Glycolytic H+ and ATP production were quantified from changes in muscle pH, phosphocreatine concentration, and Pi concentration as measured by 31P magnetic resonance spectroscopy. Substrate (HP) and metabolite (Pi, ADP, and AMP) levels remained high when exercise stopped because of the occlusion of blood flow with a pressure cuff. Glycolytic flux declined to basal levels within ∼20 s of the end of exercise despite elevated levels of HP and metabolites. Therefore, this flux does not subside because of insufficient HP substrate; rather, glycolysis is controlled independently of glycogenolytic HP production. We conclude that the inactivation of glycolysis after exercise reflects the cessation of contractile activity and is mediated within the glycolytic pathway rather than via the control of glycogen breakdown.