These experiments investigate events involved in triggering sugar accumulation in the cold in tubers of Solanum tuberosum L. cv. Desiree. Sugar content, C-14-glucose metabolism, metabolite levels and activities of sucrose phosphate synthase (SPS) and starch-degrading enzymes were followed after transfer to 4 degrees C. (i) Net sucrose accumulation began between 2 and 4 d. By 10 d, reducing sugars were also increasing. From 20 d onwards, sugar accumulation slowed. Sucrose fell, but reducing sugars continued to increase. (ii) To measure unidirectional sucrose synthesis, U-[C-14]glucose was injected into tubers after various times at 4 degrees C. The tubers were then incubated for 6 h. After 1 d at 4 degrees C, both the absolute and the relative (expressed as a percentage of the metabolized label) rates of sucrose synthesis decreased compared to those at 20 degrees C. Between 2 and 4 d at 4 degrees C, labelling of sucrose increased 3-fold, to over 60% of the metabolized label. This high rate was maintained for up to 50 d in cold storage. When tissue slices were incubated with 2.5 mol m(-3) U-[C-14]glucose, the rate of labelling of sucrose in slices from 6 d cold-stored material was higher than in slices from warm-stored material, irrespective of whether the incubation occurred at 4 degrees C or at 20 degrees C. (iii) Hexosephosphates increased during the first day after transfer to 4 degrees C. Their levels fell during the next 3 d, as sucrose synthesis increased. They then rose (until 20 d) and fell, in parallel with the rise and decline of sucrose levels. UDPglucose remained unaltered during the first 4 d, and then increased and decreased in parallel with sucrose, (iv) SPS activity assayed in optimal conditions and the total amount of SPS protein did not change. However, when assayed in the presence of phosphate and limiting substrate concentrations, activity rose 3-5-fold between 2 and 4 d. (v) Amylases and phosphorylases were investigated using zymograms to separate isoforms. Phosphorylases did not change. Between 2 and 4 d at 4 degrees C, a new amylolytic activity appeared. (vi) Estimates of the specific activity of the phosphorylated intermediates and the absolute rate of sucrose synthesis (calculated from the C-14-labelling data and metabolite analysis) showed that changed kinetic properties of SPS and decreased levels of hexose-phosphate are accompanied by a 6-8-fold stimulation of sucrose synthesis. They also show that the final level of sugar is partly determined by a cycle of sugar synthesis and degradation. (vii) It is concluded that the onset of sugar accumulation in cold-stored tubers is initiated by a change in the kinetic properties of SPS and the appearance of a new amylolytic activity. It is discussed how other factors, including hexose-phosphate levels and subcellular compartmentalization, could also influence the final levels of sugars by altering the balance of sugar synthesis and remobilization.
