Soluble Acid Invertase Activity Research Articles

Analysis of carbohydrate metabolism enzymes and cellular contents of sugars and proteins during spruce somatic embryogenesis suggests a regulatory role of exogenous sucrose in embryo development

Carbohydrate metabolism was investigated during spruce somatic embryogenesis. During the period of maintenance corresponding to the active phase of embryogenic tissue growth, activities of soluble acid invertase and alkaline invertase increased together with cellular glucose and fructose levels. During the same time, sucrose phosphate synthase (SPS) activity increased while sucrose synthase (SuSy) activity stayed constant together with the cellular sucrose level. Therefore, during maintenance, invertases were thought to generate the hexoses necessary for embryogenic tissue growth while SuSy and SPS would allow cellular sucrose to be kept at a constant level. During maturation on sucrose-containing medium, SuSy and SPS activities stayed constant whereas invertase activities were high during the early stage of maturation before declining markedly from the second to the fifth week. This decrease of invertase activities resulted in a decreased hexose:sucrose ratio accompanied by starch and protein deposition. Additionally, carbohydrate metabolism was strongly modified when sucrose in the maturation medium was replaced by equimolar concentrations of glucose and fructose. Essentially, during the first 2 weeks, invertase activities were low in tissues growing on hexose-containing medium while cellular glucose and fructose levels increased. During the same period, SuSy activity increased while the SPS activity stayed constant together with the cellular sucrose level. This metabolism reorganization on hexose-containing medium affected cellular protein and starch levels resulting in a decrease of embryo number and quality. These results provide new knowledge on carbohydrate metabolism during spruce somatic embryogenesis and suggest a regulatory role of exogenous sucrose in embryo development.

Activity and Stability of a Soluble Acid Invertase from Sugarbeet Roots

Soluble acid invertase catalyzes the hydrolysis of sucrose to the invert sugars, glucose and fructose, and has been implicated in sucrose loss during sugarbeet root development, postharvest storage and processing. To better understand the ability of this enzyme to degrade sucrose during sugarbeet root production and processing, the activity and stability of the major sugarbeet root soluble acid invertase were determined under different physical conditions. Maximum enzyme velocity was observed at sucrose concentrations above 20 mM, with a K m for sucrose of 8.9 mM. The enzyme exhibited a plateau of activity at pH 5.0 to 5.5, a pH range in which most plant soluble acid invertases exhibit high activity. Enzyme activity increased at pH values less than 5.0, possibly due to the loss of an inhibitor. Irreversible inactivation of the enzyme occurred at pH values of 7.5 or greater. Enzyme activity was greatest at 35°C and declined rapidly at temperatures above or below this optimum. At 5°C, the enzyme retained 16% of its maximal activity. Partial inactivation of the enzyme occurred at 40° to 50°C; complete inactivation was observed at 55°C and above. Inactivation at elevated temperatures was rapid and irreversible. This study indicates that the major soluble acid invertase is likely to operate at or near its maximum velocity in sugarbeet roots, is capable of hydrolyzing sucrose at the temperatures typical of sugarbeet root storage, and is active at the pH conditions typical of sugarbeet root processing.

Manipulation of fruit sugar composition but not content in Lycopersicon esculentum fruit by introgression of an acid invertase gene from Lycopersicon pimpinellifolium

Summary • The effects are shown here of genetic manipulation of invertase on fruit sugar metabolism in tomato. • Introgression of the acid invertase gene on chromosome 3 from Lycopersicon pimpinellifolium (an accession containing high invertase activity) into Lycopersicon esculentum (with relatively low activity) was used to study effects on fruit development and both sugar composition and content. • The L. pimpinellifolium parent fruit had higher fruit invertase activities, greater hexose contents and lower sucrose accumulation that the L. esculentum parent. A strong correlation between fruit invertase activity and soluble sugar content was observed only in the L. pimpinellifolium parent and not in the homozygous fruit from either cross. In all cases high soluble acid invertase activities prevented sucrose accumulation and led to the build-up of hexoses. • The introgression of the invertase locus from L. pimpinellifolium into L. esculentum did not result in higher soluble solids in the progeny than in the L. esculentum parent, but it did modify their relative composition. Changes in the invertase gene composition alone are not sufficient to increase fruit soluble solids in tomato.

Sucrose metabolism in domesticated cherry tomato, Lycopersicon esculentum var. cerasiforme Alef., and purification of sucrose synthase

SummarySucrose metabolism was followed in developing fruit of domesticated cherry tomato. The high activities of soluble acid invertase (EC 3.2.1.26) were consistently linked with high reducing sugars, whereas Sucrose synthase (SuSy) (EC.2.4.1.13) reached a peak of activity during early stage of maturation and then decreased to near nil in mature non-growing fruits. Sucrose phosphate synthase (SPS) (EC 2.4.1.14) activity remains relatively constant throughout development, indicate it may not be an indication of cherry tomato fruit sink strength. Thus, like the large fruited types, SuSy and acid invertase, rather than SPS, could play a central role in regulating sucrose accumulation in cherry tomatoes. The result also suggested that SuSy is an indicator of sink strength in growing tomato fruit. Cultivated cherry tomato SuSy (UDP-glucose: D-fructose 2-glucosyltransferase) was purified (687-fold) to homogeneity by ammonium sulfate precipitation, anion exchange chromatography on DEAE-Toyopreal 650 and gel filtration on Sephadex G-200. Further purification to homogeneity resulted from a single band from sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme was a homotetramer with a total molecular mass of 370 kDa and subunits of 92 kDa with maximum activity for the cleavage and synthesis of sucrose was at pH 7.0 and 8.0, respectively. The optimum temperature was 408C for both directions in HEPES-KOH buffer and an isoelectric point of 5.8. The enzymatic reaction followed typical Michaelis-Menten kinetics, with the following parameters: Km (fructose), 7.84; (UDP-glucose), 0.2612; (sucrose), 33.24; (UDP), 0.0946. The enzyme was very sensitive to inhibition by heavy metals.

Loss of Freezing Tolerance Associated with Decrease in Sugar Concentrations by Short-term Deacclimation in Cabbage Seedlings

Cabbage seedlings cold-acclimated for 8 days at 5°C were transferred to 15, 20 or 25°C in the dark or light. The acquired freezing tolerance was reversed during deacclimation at all temperatures ; the higher the deacclimation temperature, the more accelerated was the loss of freezing tolerance. Concentrations of soluble sugars, particularly sucrose in the leaves, rapidly decreased within deacclimation for 1 to 3 hr at 20°C in the dark. This reduction was accompanied with a decrease in the freezing tolerance of leaves. Activities of soluble acid invertase in leaves were not affected by short-term deacclimation.

Muskmelon Fruit Soluble Acid Invertase and Sucrose Phosphate Synthase Activity and Polypeptide Profiles during Growth and Maturation

Muskmelon [Cucumis melo L. (Reticulatus Group)] fruit sugar content is the single most important consumer preference attribute. During fruit ripening, sucrose accumulates when soluble acid invertase (AI) activity is less then sucrose phosphate synthase (SPS) activity. To genetically heighten fruit sugar content, knowledge of sugar accumulation during fruit development in conjunction with AI and SPS enzyme activities and their peptide immunodetection profiles, is needed. Two netted muskmelon cultivars, Valley Gold a high sugar accumulator, and North Star a low sugar accumulator, with identical maturity indices were assayed for fruit sugars, AI and SPS activity, and immunodetection of AI and SPS polypeptides 2, 5, 10, 15, 20, 25, 30, 35, or 40 (abscission) days after anthesis (DAA). Both cultivars, grown in the Fall, 1998 and Spring, 1999, showed similar total sugar accumulation profiles. Total sugars increased 1.5 fold, from 2 through 5 DAA, then remained unchanged until 30 DAA. From 30 DAA until abscission, total sugar content increased, with `Valley Gold' accumulating significantly more than `North Star'. During both seasons, sucrose was detected at 2 DAA, which coincided with SPS activity higher than AI activity, at 5 through 25 DAA, no sucrose was detected which coincided with SPS activity less than AI activity. At 30 DAA when SPS activity was greater than AI activity, increased sucrose accumulation occurred. `Valley Gold' at abscission had higher total sugar content and SPS activity, and lower AI activity than `North Star'. `North Star' had AI isoforms at 75, 52, 38, and 25 kDa (ku) that generally decreased with maturation, although the isoform at 52 ku remained detectable up to anthesis (40 DAA). `Valley Gold' had the same four AI isoforms, all decreased with maturation and became undetectable by 20 DAA. Both `Valley Gold' and `North Star' had one SPS band at 58 ku that increased with DAA, and coincided with SPS activity. `Valley Gold' had a more intense SPS polypeptide band at abscission than `North Star'. Thus, netted muskmelon fruit sugar accumulation may be increased, either by genetic manipulation or by selecting for cultivars with a specific number of down-regulated AI isoforms, and higher SPS activity during fruit ripening.

Fructan accumulation induced by nitrogen deficiency in barley leaves correlates with the level of sucrose:fructan 6-fructosyltransferase mRNA.

Hydroponically cultivated barley plants were exposed to nitrogen (N)-deficiency followed by N-resupply. Metabolic and genetic regulation of fructan accumulation in the leaves were investigated. Fructan accumulated in barley leaves under N-deficiency was mobilized during N-resupply. The enhanced total activity of fructan-synthesizing enzymes, sucrose:sucrose 1-fructosyltransferase (EC 2.4.1.99) and sucrose:fructan 6-fructosyltransferase (6-SFT; EC 2.4. 1.10) caused by N-deficiency decreased with the mobilization of fructan during N-resupply. The activity of the barley fructan-degrading enzyme, fructan exohydrolyase (EC 3.2.1.80) was less affected by the N status. The low level of foliar soluble acid invertase activity under N-deficiency conditions was maintained during the commencement of N-resupply but increased subsequently. Further analyses by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, western blot and northern blot demonstrated that the fructan accumulation and the total activity of fructan-synthesizing enzymes correlated with the 6-SFT mRNA level. We suggest that the changes in fructan levels under N stress are intimately connected with the regulation of fructan synthetic rate which is mostly controlled by 6-SFT.

174 Muskmelon Fruit Soluble Acid Invertase and Sucrose Phosphate Synthase Activity and Polypeptide Profiles during Growth and Maturation

Muskmelon [Cucumis melo L. (Reticulatus Group)] fruit sugar content is the single most important consumer preference attribute. During fruit ripening, sucrose accumulates when soluble acid invertase (AI) activity is less then sucrose phosphate synthase (SPS) activity. To genetically heighten fruit sugar content, knowledge of sugar accumulation during fruit development in conjunction with AI and SPS enzyme activities and their peptide immunodetection profiles is needed. Two netted muskmelon cultivars [`Valley Gold' (VG), a high sugar accumulator, and `North Star' (NS), a low sugar accumulator] with similar maturity indices were assayed for fruit sugars, AI, and SPS activity and immunodetection of AI and SPS polypeptides following 2, 5, 10, 15, 20, 25, 30, 35, and 40 (abscission) days after anthesis (DAA). Both cultivars, grown in spring and fall, showed similar total sugar accumulation profiles. Total sugars increased 1.5 fold, from 2 through 5 DAA and then remained unchanged until 30 DAA. From 30 DAA until abscission, total sugar content increased, with VG accumulating significantly more sugar then NS. In both cultivars, during both seasons, sucrose was detected at 2 DAA, which coincided with higher SPS activity than AI activity. At 5 through 25 DAA, SPS activity was less then AI activity resulting in little or no sucrose detection. It was not until 30 DAA that SPS activity was greater than AI activity resulting in increased sucrose accumulation. VG at abscission had higher total sugar content and SPS activity and lower AI activity than NS. Total polypeptides from both cultivars 2 through 40 DAA, were immunodetected with antibodies: anti-AI and anti-SPS. NS had Al isoforms bands at 75, 52, 38, and 25 kDa that generally decreased wtih DAA. One isoform at 52 kDa remained detectable up to anthesis (40 DAA) VG had the same four Al isoforms, all decreased with DAA and became undetectable by 20 DAA. It is unclear if one or all AI isoforms correspond with detected enzyme activity. VG and NS had one SPS band at 58 kDa that increased with DAA and concomitantly with SPS activity. VG had a more intense SPS polypeptide band at abscission then did NS. Thus, netted muskmelon sugar accumulation may be increased by selecting for cultivars with a specific number of AI isoforms, which are down-regulated, and with high SPS activity during fruit ripening.

Differential expression of soluble acid invertase genes in the shoots of high-sucrose and low-sucrose species of Saccharum and their hybrids

The hydrolytic activity of soluble acid invertase (SAI) is strongly correlated to sucrose accumulation in sugarcane (Saccharum spp.). Plants exhibiting SAI activity above a low threshold level do not accumulate high concentrations of sucrose. The present work investigates the basis for the difference in SAI activity observed between high- and low-sucrose-accumulating sugarcane lines. SAI-encoding cDNAs were isolated from two high- and one low-sucrose lines. All of these cDNAs were highly similar, with deduced proteins at least 98% identical. Expression of SAI in the stem of sugarcane was developmentally regulated, with relatively larger pools of SAI protein and mRNA in the apex and young internodes, which declined rapidly in the maturing internodes where sucroseaccumulation occurs. This developmental pattern, while qualitatively similar, was quantitatively quite different between low- and high-sucrose lines. SAI protein and mRNA pools started substantially higher, declined later, and stabilized at a significantly higher level in a low-sucrose line than in a high-sucrose line. These data indicate that differences in SAI activity between high- and low-sucrose sugarcane lines are due, at least in part, to differences inthe level of expression of essentially identical SAI genes.

Metabolic engineering of invertase activities in different subcellular compartments affects sucrose accumulation in sugarcane cells

Transgenic sugarcane (Saccharum officinarum L.) lines were created to express altered invertase isoform activity to elucidate the role(s) of invertase in the sucrose accumulation process. A sugarcane soluble acid invertase cDNA (SCINVm, AF062734) in the antisense orientation was used to decrease invertase activity. The Saccharomyces cerevisiae invertase gene (SUC2), fused with appropriate targeting elements, was used to increase invertase activity in the apoplast, cytoplasm and vacuole. A callus/liquid culture system was established to evaluate change in invertase activity and sugar concentration in the transgenic lines. Increased invertase activity in the apoplast led to rapid hydrolysis of sucrose and rapid increase of hexose in the medium. The cellular hexose content increased dramatically and the sucrose level decreased. Cells with higher cytoplasmic invertase activity did not show a significant change in the sugar composition in the medium, but did significantly reduce the sucrose content in the cells. Transformation with the sugarcane antisense acid invertase gene produced a cell line with moderate inhibition of soluble acid invertase activity and a 2-fold increase in sucrose accumulation. Overall, intracellular and extracellular sugar composition was very sensitive to the change in invertase activities. Lowering acid invertase activity increased sucrose accumulation.

Sucrose cycling, Rubisco expression, and prediction of photosynthetic acclimation to elevated atmospheric CO2

Photosynthetic acclimation to elevated CO2 cannot presently be predicted due to our limited understanding of the molecular mechanisms and metabolic signals that regulate photosynthetic gene expression. We have examined acclimation by comparing changes in the leaf content of RuBP carboxylase/oxygenase (Rubisco) with changes in the transcripts of Rubisco subunit genes and with leaf carbohydrate metabolism. When grown at 1000 mm3 dm–3 CO2, 12 of 16 crop species at peak vegetative growth had a 15–44% decrease in leaf Rubisco protein, but with no specific association with changes in transcript levels measured at midday. Species with only modest reductions in Rubisco content (10–20%) often had a large reduction in Rubisco small subunit gene mRNAs (> 30%), with no reduction in large subunit gene mRNAs. However, species with a very large reduction in Rubisco content generally had only small reductions in transcript mRNAs. Photosynthetic acclimation also was not specifically associated with a change in the level of any particular carbohydrate measured at midday. However, a threshold relationship was found between the reduction in Rubisco content at high CO2 and absolute levels of soluble acid invertase activity measured in plants grown at ambient or high CO2. This relationship was valid for 15 of the 16 species examined. There also occurred a similar, albeit less robust, threshold relationship between the leaf hexose/sucrose ratio at high CO2 and a reduced photosynthetic capacity ≥ 20%. These data indicate that carbohydrate repression of photosynthetic gene expression at elevated CO2 may involve leaf sucrose cycling through acid invertase and hexokinase.

Soluble acid invertase activity in leaves is independent of species differences in leaf carbohydrates, diurnal sugar profiles and paths of phloem loading

Leaf sucrose, starch, hexose and maximum extractable soluble acid invertase activity were compared throughout the day in source leaves of 13 plant species chosen for their putative phloem‐loading type (apoplastic or symplastic). Four species which represent the different phloem‐loading types (tomato, barley, maize and Fuchsia) were studied in detail. Using this information we wished to determine whether a positive correlation between foliar carbohydrates and acid invertase activity exists in leaves from different species and, furthermore, whether this relationship is determined by phloem‐loading type. Acid invertase activity was relatively constant throughout the day in all species. The extent of sucrose, hexose and starch accumulation and the sucrose: starch ratio measured at a given time were species‐dependent. No correlations were found between foliar soluble acid invertase activity and the hexose, sucrose or starch content of the leaves in any of the species, regardless of phloem‐loading type. The species examined could be divided into three distinct groups: (1) high sucrose, low invertase; (2) low sucrose, low invertase; and (3) low sucrose, high invertase. The absence of an inverse relationship between leaf sucrose, hexose or starch contents and endogenous soluble acid invertase suggests that this enzyme is not directly involved in carbon partitioning in leaves but serves an auxiliary function.

Sucrose metabolism during apical hook opening in sunflower hypocotyls

The relationships between cell elongation and changes in the activities of enzymes of sucrose metabolism were investigated in the hypocotyl hook of sunflower seedlings ( Helianthus annuus L.) that were grown either in darkness or irradiated with continuous white light (WL). Hook opening was promoted by white light (WL). In etiolated seedlings, endogenous hook opening occurred between days 5 and 6 after sowing (darkness). This response is due to a large increase in the rate of cell elongation in the inner side and a decrease in cell length on the outer surface of the organ. After WL irradiation and between days 5 and 6 after sowing (darkness) an enhancement in the specific activity of the enzyme soluble acid invertase (EC 3.2.1.26) was measured, whereas that of sucrose synthase (EC 2.4.1.13) was unaffected. The concentration of sucrose decreased in WL, and to a lesser extent, in darkness. The results demonstrate that endogenous and WL-induced apical hook opening is accompanied by sucrolysis via acid invertase. We suggest that apical hook opening promotes invertase activity and thus sucrose catabolism.

Antioxidant defences of the apoplast

The apoplast of barley and oat leaves contained superoxide dismutase (SOD), catalase, ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase, and glutathione reductase activities. The activities of these enzymes in the apoplastic extracts were greatly modified 24 h after inoculation with the biotrophic fungal pathogenBlumeria graminis. The quantum efficiency of photosystem II, which is related to photosynthetic electron transport flux, was comparable in inoculated and healthy leaves during this period. Apoplastic soluble acid invertase activity was also modified in inoculated leaves. Inoculation-dependent increases in apoplastic SOD activity were observed in all lines. Major bands of SOD activity, observed in apoplastic protein extracts by activity staining of gels following isoelectric focusing, were similar to those observed in whole leaves but two additional minor bands were found in the apoplastic fraction. The apoplastic extracts contained substantial amounts of dehydroascorbate (DHA) but little or no glutathione (GSH). Biotic stress decreased apoplastic ascorbate and DHA but increased apoplastic GSH in resistant lines. The antioxidant cycle enzymes may function to remove apoplastic H2O2 with ascorbate and GSH derived from the cytoplasm. DHA and oxidized glutathione may be reduced in the apoplast or returned to the cytosol for rereduction.

Effects of drought on non‐mycorrhizal and mycorrhizal maize: changes in the pools of non‐structural carbohydrates, in the activities of invertase and trehalase, and in the pools of amino acids and imino acids

To study the response of non‐mycorrhizal and mycorrhizal maize plants to drought, the changes in the pools of non‐structural carbohydrates and amino acids were analysed in leaves and roots of two maize cvs. Plants well colonized by the arbuscular mycorrhizal fungus Glomus mosseae (Nicol. & Gerd.) (60% of root length infected) and comparable non‐mycorrhizal plants were subjected to moderate drought stress by reducing the water supply. This stress induced a conspicuous increase in the trehalose pool in the mycorrhizal roots, probably because it was accumulated by the fungal symbiont. Furthermore, glucose and fructose were accumulated in leaves and roots of non‐mycorrhizal plants but not in the mycorrhizal ones. Starch disappeared completely from the leaves of both mycorrhizal and non‐mycorrhizal plants in response to drought. Activities of soluble acid invertase and trehalase were also measured. Acid invertase activity increased during drought in the leaves of both non‐mycorrhizal and mycorrhizal plants whilst in the roots it was unaffected in non‐mycorrhizal plants and decreased in the mycorrhizal ones. Without drought stress, trehalase activity was considerably higher in the leaves and roots of mycorrhizal plants than in those of non‐mycorrhizal plants. It increased conspicuously during drought, primarily in the leaves of non‐mycorrhizal plants. A drought‐induced accumulation of amino acids as well as imino acids was found in roots and leaves of both mycorrhizal and non‐mycorrhizal plants; leaves of mycorrhizal plants accumulated more imino acids than those of non‐mycorrhizal ones. Our results show that drought stress and the presence of a mycorrhizal fungus have a considerable effect on carbon partitioning, imino acid and amino acid accumulation in maize plants.

Re-Examination of the solute-import hypothesis of gibberellin action in developing pea internodes

Exogenous gibberellic acid (GA 3 ) strongly stimulated the growth of internodes 1, 2 and 3 in light-grown pea seedlings ( Pisum sativum L. var. Senator). In all three internodes, rapid GA 3 -induced elongation was accompanied by a rise in the specific catalytic activity of the sucrolysis enzyme soluble acid invertase. The activity of the second sucrose-metabolizing enzyme, soluble sucrose synthase, was not affected by GA 3 . Cessation of growth was associated with a corresponding decrease in acid invertase and an increase in sucrose synthase activity. In all three internodes, GA 3 -induced growth was associated with an increase in the level of glucose. The osmotic concentration of the tissue sap from the rapidly growing internodes of GA 3 -treated seedlings was as large as in the water control. As a result of GA 3 -treatment, the concentration of osmotica increased in the mature region of the stem. We conclude that GA 3 -induced growth of pea internodes is in part attributable to a hormone-mediated increase in acid invertase activity, which results in the provision of hexoses for energy metabolism, biosynthesis of cell walls and the maintenance of intracellular osmotic concentration during rapid elongation of the stem.

Sucrose Accumulation in the Sugarcane Stem Is Regulated by the Difference between the Activities of Soluble Acid Invertase and Sucrose Phosphate Synthase.

To assess the relative importance of morphological and biochemical factors in the regulation of sucrose (Suc) accumulation in the sugarcane (Saccharum spp. hybrids) stem, we investigated morphological and biochemical correlates of Suc accumulation among parents and progeny of a family segregating for differences. In contrast to the parents, no relationship was observed between morphology and the level of Suc accumulation among the progeny. The level and timing of Suc accumulation in the whole stalk and within individual internodes was correlated with the down-regulation of soluble acid invertase (SAI) activity. High SAI activity prevented most, but not all, Suc accumulation. There was a critical threshold of SAI activity above which high concentrations of Suc did not accumulate. This low level of SAI activity was always exceeded in the internodes of the lower-Suc-storing genotypes. However, low activity of SAI was not sufficient by itself to account for the Suc accumulation in the higher-Suc-storing genotypes. Major differences in Suc accumulation among the population were attributed to the difference between activities of SAI and Suc phosphate synthase, provided SAI is below the critical threshold concentration. This result is not unexpected, since the pathway of Suc transport for storage involves Suc hydrolysis and resynthesis.

Developmental changes of enzymes involved in conversion of sucrose to hexose-phosphate during early tuberisation of potato

A highly synchronised in-vitro tuberisation system, based on single-node cuttings containing an axillary bud, was used to investigate the activity patterns of enzymes involved in the conversion of sucrose to hexose-phosphates during stolon-to-tuber transition of potato (Solanum tuberosum L.). Two different non-tuberising systems were included to distinguish between changes that are or are not tuber-specific. At tuberisation the activity of soluble acid invertase decreased (13-fold) and of sucrose synthase increased (12-fold). The activity of both enzymes remained unchanged in the non-tuberising treatments. Based on the opposite patterns and large difference in activity of these two sucrolytic enzymes, we conclude that sucrose synthase constitutes the predominant route of sucrose breakdown after tuber initiation. During the period before tuberisation, the activity of cell-wall-bound invertase and of hexokinase showed a highly positive correlation (r 2 = 0.96 in all the three treatments, suggesting coordinated coarse control of both enzyme activities. After the onset of tuberisation cell-wall-bound invertase activity decreased to a very low level, a change not observed in the non-tuberising systems, indicating that cell-wall-bound invertase is presumably not involved in the unloading mechanism and/or short-distance transport of sucrose within the perimedulla of growing tubers. The overall activity of fructokinase and of hexokinase both showed a fourfold increase after tuber initiation, but remained unchanged in the non-tuberising systems. The increase of fructokinase suggests that the phosphorylation of fructose by fructokinase down-regulates the cytosolic fructose content in order to maintain a high sucrose-synthase-catalysed net flux of sucrose to phosphorylated hexoses during rapid tuber growth. The increase of total glucose-phosphorylating potential could be a response to the tuberisation-related starch accumulation process. The activity of UDP-glucose pyrophosphorylase showed no developmental change. The level of UDP-glucose pyrophosphorylase activity is very likely the result of metabolic regulation.

Carbohydrate Content and Enzyme Metabolism in Developing Canola Siliques.

Little biochemical information is available on carbohydrate metabolism in developing canola (Brassica napus L.) silique (pod) wall and seed tissues. This research examines the carbohydrate contents and sucrose (Suc) metabolic enzyme activities in different aged silique wall and seed tissues during oil filling. The silique wall partitioned photosynthate into Suc over starch and predominantly accumulated hexose. The silique wall hexose content and soluble acid invertase activity rapidly fell as embryos progressed from the early- to late-cotyledon developmental stages. A similar trend was not evident for alkaline invertase, Suc synthase (SuSy), and Suc-phosphate synthase. Silique wall SuSy activities were much higher than source leaves at all times and may serve to supply the substrate for secondary cell wall thickening. In young seeds starch was the predominant accumulated carbohydrate over the sampled developmental range. Seed hexose levels dropped as embryos developed from the early- to midcotyledon stage. Hexose and starch were localized to the testa or liquid endosperm, whereas Suc was evenly distributed among seed components. With the switch to oil accumulation, seed SuSy activity increased by 3.6-fold and soluble acid invertase activity decreased by 76%. These data provide valuable baseline knowledge for the genetic manipulation of canola seed carbon partitioning.

Sucrose metabolism and fruit growth in parthenocarpic vs seeded blueberry (Vaccinium ashei) fruits

Although fruit set and development are induced by applications of gibberellins, final fruit weight of gibberellin‐induced parthenocarpic fruit is often less than that of pollinated fruit. We examined changes in the activities of sucrose‐metabolizing enzymes and sugar accumulation in developing fruits of cultivated blueberry (Vaccinium ashei Reade) and their correlation with fruit growth upon pollination or exogenous applications of gibberellic acid (GA3). The objective was to determine if differences in fruit growth could be attributed to differences in enzyme activities and subsequent sugar accumulation in fruits. The fruit development period of GA3‐treated fruits was 15 days longer than that of pollinated fruits. At maturity, GA3‐treated fruit accumulated an average of 180 mg dry weight while pollinated fruit accumulated 390 mg dry weight. Dry weight accumulation in nonpollinated fruits was negligible and these fruits abscised by 45 days after bloom (DAB). The total carbon (C) cost (dry weight C + respiratory C) for fruit development was 109 and 244 mg C fruit‐1 for GA3‐treated and pollinated fruits, respectively. Hexose concentration increased to 100 mg (g fresh weight)‐1 at ripening in both GA3‐treated and pollinated fruits. Nonpollinated fruits reached a maximum hexose concentration at 45 DAB. Sucrose phosphate synthase (EC 2.4.1.14) and sucrose synthase (EC 2.4.1.13) activities reached a maximum of ≤5.0 μmol (g fresh weight)‐1 h‐1 in both GA3‐treated and pollinated fruits. Soluble acid invertase (EC 3.2.1.26) activity increased to about 60 μmol (g fresh weight)‐1 h‐1 in both GA3‐treated and pollinated fruits at ripening, while in nonpollinated fruits, a maximum soluble acid invertase activity of 0.12 μmol (g fresh weight)‐1 h‐1 was measured at 24 DAB. Insoluble acid invertase activity declined during the early stages of fruit growth and remained relatively low throughout fruit development. Neutral invertase activity was low throughout development, increasing to 5 μmol (g fresh weight)‐1 h‐1 at ripening in GA3‐treated and pollinated fruits. Our studies demonstrate that blueberry fruit development does not appear to be limited by sucrose metabolizing enzyme activity and/or the ability to accumulate sugars in either GA3‐treated or pollinated fruits.