Library

Notes: These experiments investigate events involved in triggering sugar accumulation in the cold in tubers of Solanum tuberosum L. cv. Desirée. Sugar content, 14C-glucose metabolism, metabolite levels and activities of sucrose phosphate synthase (SPS) and starch-degrading enzymes were followed after transfer to 4°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-[14C]glucose was injected into tubers after various times at 4°C. The tubers were then incubated for 6 h. After 1 d at 4°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°C. Between 2 and 4 d at 4°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-[14C]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°C or at 20°C. (iii) Hexose-phosphates increased during the first day after transfer to 4°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°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 14C-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.

Notes: Transfer of potato tubers to low temperature leads after 2–4 d to a stimulation of sucrose synthesis, a decline of hexose-phosphates and a change in the kinetic properties, and the appearance of a new form of sucrose phosphate synthase (SPS). Antisense and co-suppression transformants with a 70–80% reduction in SPS expression have been used to analyse the contribution of SPS to the control of cold sweetening. The rate of sucrose synthesis in cold-stored tubers was investigated by measuring the accumulation of sugars, by injecting labelled glucose of high specific activity into intact tubers, and by providing 50 mol m–3 labelled glucose to fresh tuber slices from cold-stored tubers. A 70–80% decrease of SPS expression resulted in a reproducible but non-proportional (10–40%) decrease of soluble sugars in cold-stored tubers, and a non-proportional (about 25%) inhibition of label incorporation into sucrose, increased labelling of respiratory intermediates and carbon dioxide, and increased labelling of glucans. The maximum activity of SPS is 50-fold higher than the net rate of sugar accumulation in wild-type tubers, and decreased expression of SPS in the transformants was partly compensated for increased levels of hexose-phosphates. It is concluded that SPS expression per se does not control sugar synthesis. Rather, a comparison of the in vitro properties of SPS with the estimated in vivo concentrations of effectors shows that SPS is strongly substrate limited in vivo. Alterations in the kinetic properties of SPS, such as occur in response to low temperature, will provide a more effective way to stimulate sucrose synthesis than changes of SPS expression.

Notes: Abstract Large spoils of metal-rich filtercake from the Port Kembla BHP steelworks were characterised by certain physical, chemical and biological parameters. The vegetation was assessed for potential metal hyperaccumulators and the presence of arbuscular mycorrhizae in the rhizospheres. Fresh filtercake (〈2 yr old) was charcoal in colour and had a fine (clay loam) texture. Its composition was distinctly different to the older filtercake (3–4 yr) which was similar to the reference soil, being lighter in colour, loamy textured, more aggregated and with higher organic matter. Variability between the two filtercake types also occurred for electrical conductivity and pH. Total trace metal concentrations (Cd, Cu, Pb and Zn) were high and although only minor proportions were in bioavailable forms, the greater percentages in the older filtercake suggested temporal changes in metal speciation. Fresh filtercake was devoid of any plant life and fungal communities. The older material, however, was able to sustain a reasonable environment for plant, bacterial and fungal growth. The vegetation was dominated by Ricinus communis, along with species belonging to the families Asteraceae, Solanaceae, Malvaceae and Apiaceae. Rhizosphere of all these plants harboured mycorrhizae belonging to the Gigaspora and Glomus strains and while trace metal accumulation was observed no plant was a hyperaccumulator. The reference site was principally inhabited by certain Poaceae, Asteraceae and Verbenaceae species. All were mycorrhizal (principally Glomus strains), but no plant was represented on the contaminated site. Here also trace metal accumulation was taking place. The absence of hyperaccumulation in the filtercake was considered a consequence of low levels of bioavailable trace metals. However, as such metal forms may increase over time, particularly for copper and lead, cultivation of known hyperaccumulators could become worthwhile after prolonged exposure (〉4 yr). Among the naturally colonising plants, Ricinus communis and Sonchus oleraceus are regarded the most suitable options for zinc and cadmium removal, respectively. Arbuscular mycorrhizal infection (particularly with Glomus and Gigaspora strains) should be promoted for optimum soil reclamation.