Expression of a UDPglucose pyrophosphorylase cDNA during fruit ripening of banana (Musa acuminata)

We report the isolation of a banana cDNA, designated MWUGPA, encoding uridine diphosphoryl (UDP)-glucose pyrophosphorylase (UGPase, EC.2.7.7.9) that catalyses the reversible conversion between glucose 1-phosphate and UDPglucose in plants and animals. Furthermore, UGPase expression in fruit during ripening and in response to exogenous ethylene and sugars was also investigated. MWUGPA encodes a polypeptide of 467 amino acid residues and shares a high degree of sequence similarity (85–90%) with other plant UGPase homologs. In northern blot analysis, a 1.7-kb UGPase transcript was detected in both the vegetative and reproductive organs, but the former was considerably less abundant than the latter. In fruit, the level of accumulated transcripts was higher in pulp than peel at all ripening stages. Transcript abundance in both fruit tissues was relatively constant during ripen-ing, but pulp transcripts surged in the ‘more green than yellow’ category fruit when ethylene also increased. Further analysis revealed that UGPase expression in fruit was ethylene-inducible, but the response was tissue-specific, as evidenced by the promoting effect of exogenous ethylene on accumulation of UGPase transcripts in pulp but not peel. Exogenous application of sucrose and fructose also increased UGPase transcript abundance in leaf and fruit tissues, especially pulp, whereas exogenous glucose had little or no effect. The results of this study indicate that ethy-lene and soluble sugars may play a regulatory role in UGPase expression during ripening in banana fruit.

Characterization of transgenic potato (Solanum tuberosum) tubers with increased ADPglucose pyrophosphorylase*

The aim of the work described in this paper was to characterize the tubers of potato (Solanum tuberosum var. Prairie) plants that had been transformed with the Escherichiacoli ADPglucose pyrophosphorylase (EC 2.7.7.27) gene, glgC-16, under the control of a patatin promoter. Over 30 lines of transformed plants with increased ADPglucose pyrophosphorylase activity were obtained. The tubers of six of these lines were compared with those of control plants expressing the gene for β-glucuronidase. The average increase in pyrophosphorylase activity was 200%, and the highest was 400%. Western immunoblotting of tuber extracts showed that the amounts of glgC-16 protein were linearly related to the extractable activity of the ADPglucose pyrophosphorylase. Cell fractionation studies showed that the increased activity of the pyrophosphorylase in the glgC-16 tubers had a similar intracellular location, the amyloplast fraction, to that found in the control tubers. No pleiotropic changes in the maximum catalytic activities of the following enzymes could be detected in the glgC-16 tubers: sucrose synthase, fructokinase, UDPglucose pyrophosphorylase, phosphofructokinase, soluble starch synthase, starch branching enzyme, phosphoglucomutase and alkaline inorganic pyrophosphatase. The glgC-16 tubers are held to be suitable for the study of the role of ADPglucose pyrophosphorylase in the control of starch synthesis.

High Temperature Affects the Activity of Enzymes in the Committed Pathway of Starch Synthesis in Developing Wheat Endosperm

Ears of wheat were exposed for up to 10 days during the grain-filling stage to high temperature (35�C) and activities of five enzymes in the sucrose to starch pathway were compared to those in ears maintained at lower temperature (21�C day/16�C night). Two cultivars of wheat known to differ in their post-anthesis tolerance of high temperature were compared. On a per grain basis, the activity of sucrose synthase and of ADPglucose pyrophosphorylase in ears maintained at 21/16�C throughout did not change greatly between days 16 and 32 after anthesis, whereas UDPglucose pyrophosphorylase and soluble starch synthase activities declined with advancing development. Soluble starch synthase activity in grains of heated ears was decreased within 1 day to about one- half of the value in unheated grains, and 3 days' additional heating did not reduce the activity much further. Insoluble starch synthase activity was not significantly reduced by heating. Compared to soluble starch synthase, ADPglucose pyrophosphorylase activity was more slowly affected and decreased to a lesser extent by heat. Sucrose synthase and UDPglucose pyrophosphorylase activities were either not affected or only slightly reduced; part of this reduction could be due to advanced development at the higher temperature. In recovery experiments ears were heated for brief periods and then returned to 21/16�C for a few days. ADPglucose pyrophosphorylase and soluble starch synthase activities recovered in the cooler conditions but the other two enzymes generally only maintained or lost further activity. From a comparison of the activities of these enzymes with the rate of starch deposition, and by taking into account the effects of heating, it is proposed that the influence of heating on final grain dry weight is attributable to the observed reductions of soluble starch synthase activity.

Effects of humic substances on carbohydrate metabolism of maize leaves

Maize plants were grown in nutrient solution with or without 100 mg organic carbon L−1 of humic substances (HS) (MW > 12 000) extracted from a cambisol. To evaluate the effect of HS on leaf carbohydrate metabolism, after 14 d of growth apical and basal segments from the third leaf were collected and analyzed for sugar content and for the activities of enzymes involved in the synthesis and breakdown of starch and sucrose. Leaf starch content decreased in treated plants, whereas the level of soluble sugars increased. The decrease of starch was accompanied by an enhanced activity of amylase, whereas the activity of ADPglucose pyrophosphorylase was not affected. Activities of invertases and sucrose synthase were stimulated by HS treatment in apical tissue, and inhibited in basal one. Activities of enzymes involved in sucrose biosynthesis (sucrose phosphate synthase, cytoplasmic fructose-1,6- bisphosphatase, UDPglucose pyrophosphorylase) were stimulated or inhibited depending on the enzyme and on the tissue examined. Our results show that HS may affect both the level and percent distribution of sugars of maize leaves; these effects seem to be mediated by changes in the activities of enzymes involved in carbohydrate metabolism. Key words: Humic substances, enzymes, carbohydrate metabolism, maize

Towards understanding oscillations: a mathematical model of the biochemistry of photosynthesis

A general model of electron transport, carbon assimilation, starch and sucrose synthesis was built on the basis of two partial models. Individual reactions were described by their Δ G' 0 , V m and K m values for substrates and products. The system of 33 differential equations was solved on a personal computer programmed in Turbo-PASCAL 3.0. The rate of cytosolic fructose bisphosphatase (FBPase) is modelled to be dependent on the concentration of fructose 2,6-bisphosphate (F2,6BP). The synthesis of the latter is activated by inorganic phosphate and inactivated by triose phosphates. The quantum efficiency of PSII is depressed at high proton charge in thylakoids and at high redox states of the electron transport chain. One of the aims of the model was to check whether these regulatory systems could cause oscillations in photosynthesis. Transients calculated from a low to high photon flux density and from a low to high CO 2 concentration revealed an overshoot but no oscillations. Therefore, it has not been sufficiently proved whether cytosolic FBPase and PSII activity control oscillations in photosynthesis. The phosphate-limited photosynthesis is stable in cases where UDPglucose pyrophosphorylase and ADPglucose pyrophosphorylase have greater affinity for ATP (UTP) than CO 2 assimilation. In a phosphate-limited state high ΔpH is not generated, as electron transport becomes limited by the low concentration of 1,3-diphosphoglycerate.