Invertase Activity, Carbohydrate Metabolism and Cell Expansion in the Stem of Phaseolus vulgaris L.

In the stem of Phaseolus vulgaris L. the specific activity ofacid invertase was highest in the most rapidly elongating internode.Activity of the enzyme was very low in internodes which hadcompleted their elongation, in young internodes before the onsetof rapid elongation, and in the apical bud. From shortly afterits emergence from the apical bud the elongation of internode3 was attributable mainly to cell expansion. Total and specificactivities of acid invertase in this internode rose to a maximumat the time of most rapid elongation and then declined. Transferof plants to complete darkness, or treatment of plants withgibberellic acid (GA₃), increased the rate of internode elongationand final internode length by stimulating cell expansion. Bothtreatments rapidly increased the total and specific activitiesof acid invertase in the responding internodes; peak activitiesof the enzyme occurred at the time of most rapid cell expansion. In light-grown plants, including those treated with GA₃, rapidcell and internode elongation and high specific activities ofacid invertase were associated with high concentrations of hexosesugar and low concentrations of sucrose. As cell growth ratesand invertase activities declined, the concentration of hexosefell and that of sucrose rose. In plants transferred to darkness,stimulated cell elongation was accompanied by a rapid decreasein hexose concentration and the disappearance of sucrose, indicatingrapid utilization of hexose. No sucrose was detected in theapical tissues of light-grown plants. The results are discussed in relation to the role of acid invertasein the provision of carbon substrates for cell growth. Key words: Cell expansion, Acid invertase, Hexose, Sucrose, Phaseolus     

Soluble and insoluble invertase activity in elongating Tulipa gesneriana flower stalks

Previously 'frozen' Tulipa gesneriana L. bulbs cv. Apeldoorn, were planted and grown at higher temperatures to study the role of invertase (EC 3.2.1.26) in the cold-induced elongation of the flower stalk internodes. After planting, flower stalks were left intact, or, the leaves and flower bud were both removed to inhibit internode elongation. In intact flower stalks, elongation of the internodes was accompanied by an accumulation of glucose and an initial decrease in the sucrose content g,⁻¹ dry weight. Insoluble invertase activity g,⁻¹ dry weight hardly changed, but soluble invertase activity showed a peak pattern, that was related, at least for the greater part, to the changes in the sugar contents. Peak activities of soluble invertase were found during (lower- and uppermost internodes) or around the onset of the rapid phase of internode elongation (middle internodes). Internode elongation and glucose accumulation immediately ceased when the leaves and flower bud were removed. Insoluble invertase activity g,⁻¹ dry weight remained at its initial level (lowermost internode) or increased more towards the upper internodes. Soluble invertase activity did not further increase (uppermost internode) or decreased abruptly to a low level. It is concluded that soluble invertase may be one of the factors contributing to glucose accumulation and internode elongation in the tulip flower stalk.     

Sucrose accumulation in sweet sorghum stem internodes in relation to growth

Sweet sorghum (Sorghum bicolor L. Moench) stems of different cultivars (NK 405. Keller and Tracy) reveal a different pattern of sucrose accumulation with respect to in-ternodal sugar content and distribution. The onset of sucrose storage is not necessarily associated with the reproductive stage of the plant, as was hitherto assumed, but obviously occurs after cessation of internodai elongation as was postulated for the sugarcane stem. For at least two of the three cultivars, ripening is an internode to internode process beginning at the lowermost culm parts. Intensive growth of the internodes, combined with a high hexose content in stern parenchyma, shows a strong positive correlation (r |Mg 0.94) to the activity of sucrose synthase (SuSy; EC 2.4.13), but not to invertase (EC 3.2.1.26) which is not present as soluble (neutral and acid) or cell wall-bound, salt-extractable enzyme in the three culsivars investigated. Sucrose synthase measured in sucrose cleavage and synthesis direction reveals divergent activity rates and sensitivity towards exogenously applied Mg²⁺ ions and pH. SuSy activity is connected to the increase of internodai sucrose content in so far as (1) its decline is a prerequisite for the onset of sucrose accumulation and (2) it remains at a constant low level during sucrose storage. Sucrose phosphate synthase (SPS; EC 2.4.1.14) activity in the sorghum stem is low compared to SuSy and uniformly distributed over all inter-nodes. Only source leaves of sorghum show a considerable SPS activity, but neither stem nor leaf SPS reveal a positive correlation to the increase of internodai sucrose content. Sucrose phosphate phosphatase (SPP; EC 3.1.3.24) amounts lo only 24–30% of the respective SPS activity but follows the same distribution pattern. None of the enzymes under study proves to be responsible for the extent of sucrose storage in the stem, so other phenomena such as transport processes within the stern tissue require further investigation.     

Carbohydrate Status of Tulip Bulbs during Cold-Induced Flower Stalk Elongation and Flowering

The effect of a cold treatment on the carbohydrate status of the scales and flower stalk of Tulipa gesneriana L. cv Apeldoorn bulbs during growth after planting was studied and compared with bulbs not given cold treatment. Bulbs were stored dry for 12 weeks at 5[deg]C (precooled) or 17[deg]C (noncooled). Only the 5[deg]C treatment led to rapid flower stalk elongation and flowering following planting at higher temperatures. Precooling enhanced mobilization of starch, fructans, and sucrose in the scales. The cold-stimulated starch breakdown was initially accompanied by increased [alpha]-amylase activity per scale. In noncooled bulbs, [alpha]-amylase activity slightly decreased or remained more or less constant. Cold-induced flower stalk elongation was partially accompanied by a decrease in the sucrose content and an increase in the glucose content and invertase activity per g dry weight. The starch content in internodes initially decreased and subsequently increased; [alpha]-amylase activity per g dry weight of the lowermost internode showed a peak pattern during starch breakdown and increased thereafter. The internodes of noncooled bulbs, on the contrary, accumulated sucrose. Their glucose content and invertase activity per g dry weight remained low. Starch breakdown was not found and [alpha]-amylase activity per g dry weight of the lowermost internode remained at a low level. Precooling of tulip bulbs thus favors reserve mobilization in the scales and flower stalk and glucose accumulation in the elongating internodes.     

Plant density influences fiber sucrose metabolism in relation to cotton fiber quality

Planting density plays an important role in improving cotton yield and regulating fiber quality. A 2-year experiment was conducted to investigate the effects of plant density on sucrose metabolism in relation to fiber quality of field-grown cotton. The results showed that lint yield increased with increasing plant density, fiber micronaire, fiber maturity ratio, and fiber fineness decreased with the increasing of plant density, whereas fiber length, fiber uniformity index, fiber strength, and fiber elongation were little affected by plant density. Increased plant density decreased sucrose synthase (SuSy) activity, sucrose content, and cellulose content in cotton fiber, but increased invertase activity. Increased invertase activity would restrain SuSy activity in cotton fiber: therefore, SuSy activity was the most severely affected enzyme in fiber sucrose metabolism by cotton plant density during fiber development. Abundant sucrose content in fiber after 24 days post anthesis (DPA) and high activities of SuSy and sucrose phosphate synthase (SPS) at 38 DPA were beneficial for cellulose synthesis, and were propitious to optimize the fiber maturity properties. The results also showed that fiber micronaire, maturity ratio, and fineness decreased 0.11, 0.02, and 5.89 mtex, respectively, with each increase of 10,000 plants per hectare. It was concluded that high plant density decreased SuSy activity, sucrose content, and cellulose content, but increased invertase activity in sucrose metabolism, resulting in low fiber micronaire, fiber maturity ratio, and fiber fineness.     

Effect of Night Temperature on the Activity of Sucrose Phosphate Synthase, Acid Invertase, and Sucrose Synthase in Source and Sink Tissues of Rosa hybrida cv Golden Times

Sucrose phosphate synthase and acid invertase activities in the mature leaves of roses (Rosa hybrida cv Golden Times) were greater in plants grown under a higher night temperature than under a lower temperature regime. In young shoots, the activity of acid invertase was promoted by the lower temperature while that of sucrose synthase was increased at the higher temperature. At both temperatures benzyladenine when applied to the axillary bud stimulated sucrose phosphate synthase activity and advancement of its peak of activity in the leaf subtending to the bud, and also stimulated sucrose synthase activity in the young shoot. At the lower temperature, application of benzyladenine to the axillary bud stimulated acid invertase activity in the young shoot but not in the leaves.     

Expression analysis of genes associated with sucrose accumulation in sugarcane (Saccharum spp. hybrids) varieties differing in content and time of peak sucrose storage

Sucrose synthesis/accumulation in sugarcane is a complex process involving many genes and regulatory sequences that control biochemical events in source–sink tissues. Among these, sucrose synthase (SuSy), sucrose phosphate synthase (SPS), soluble acid (SAI) and cell wall (CWI) invertases are important. Expression of these enzymes was compared in an early (CoJ64) and late (BO91) maturing sugarcane variety using end‐point and qRT‐PCR. Quantitative RT‐PCR at four crop stages revealed high CWI expression in upper internodes of CoJ64, which declined significantly in both top and bottom internodes with maturity. In BO91, CWI expression was high in top and bottom internodes and declined significantly only in top internodes as the crop matured. Overall, CWI expression was higher in CoJ64 than in BO91. During crop growth, there was no significant change in SPS expression in bottom internodes in CoJ64, whereas in BO91 it decreased significantly. Apart from a significant decrease in expression of SuSy in mature bottom internodes of BO91, there was no significant change. Similar SAI expression was observed with both end‐point and RT‐PCR, except for significantly increased expression in top internodes of CoJ64 with maturity. SAI, being a major sucrose hydrolysing enzyme, was also monitored with end‐point PCR expression in internode tissues of CoJ64 and BO91, with higher expression of SAI in BO91 at early crop stages. Enzyme inhibitors, e.g. manganese chloride (Mn⁺⁺), significantly suppressed expression of SAI in both early‐ and late‐maturing varieties. Present findings enhance understanding of critical sucrose metabolic gene expression in sugarcane varieties differing in content and time of peak sucrose storage. Thus, through employing these genes, improvement of sugarcane sucrose content is possible.     

Sucrose Metabolism at Three Leaf Development Stages in Bean Plants

Sucrose metabolism was studied at three leaf development stages in two Phaseolus vulgaris L. cultivars, Tacarigua and Montalban. The changes of enzyme activities involved in sucrose metabolism at the leaf development stages were: (1) Sink (9-11 % full leaf expansion, FLE): low total sucrose phosphate synthase (SPS) activity, and higher acid invertase (AI) activity accompanied by low sucrose synthase (SuSy) synthetic and sucrolytic activities. (2) Sink to source transition (40-47 % FLE): increase in total SPS and SuSy activities, decrease in AI activity. (3) Source (96-97 % FLE): high total SPS activity, increased SuSy activities, decreased AI activity. The hexose/sucrose ratio decreased from sink to source leaves in both bean cultivars. The neutral invertase activity was lower than that of AI; it showed an insignificant decrease during the sink-source transition.     

Sucrose Metabolism at Three Leaf Development Stages in Bean Plants

Sucrose metabolism was studied at three leaf development stages in two Phaseolus vulgaris L. cultivars, Tacarigua and Montalban. The changes of enzyme activities involved in sucrose metabolism at the leaf development stages were: (1) Sink (9-11 % full leaf expansion, FLE): low total sucrose phosphate synthase (SPS) activity, and higher acid invertase (AI) activity accompanied by low sucrose synthase (SuSy) synthetic and sucrolytic activities. (2) Sink to source transition (40-47 % FLE): increase in total SPS and SuSy activities, decrease in AI activity. (3) Source (96-97 % FLE): high total SPS activity, increased SuSy activities, decreased AI activity. The hexose/sucrose ratio decreased from sink to source leaves in both bean cultivars. The neutral invertase activity was lower than that of AI; it showed an insignificant decrease during the sink-source transition. This revised version was published online in June 2006 with corrections to the Cover Date.     

Biochemical and genetic analysis of carbohydrate accumulation in Allium cepa L

Onion and shallot (Allium cepa L.) exhibit wide variation in bulb fructan content, and the Frc locus on chromosome 8 conditions much of this variation. To understand the biochemical basis of Frc, we conducted biochemical and genetic analyses of Allium fistulosum (FF)-shallot (A. cepa Aggregatum group) alien monosomic addition lines (AALs; FF+1A-FF+8A) and onion mapping populations. Sucrose and fructan levels in leaves of FF+2A were significantly lower than in FF throughout the year, and the springtime activity of acid invertase was also lower. FF+8A showed significantly higher winter sucrose accumulation and sucrose phosphate synthase (SPS) activity. Inbred high fructan (Frc_) lines from the 'W202Ax Texas Grano 438' onion population exhibited significantly higher sucrose levels prior to bulbing than low fructan (frcfrc) lines. Sucrose synthase (SuSy) activity in these lines was correlated with leaf hexose content but not with Frc phenotype. Markers for additional candidate genes for sucrose metabolism were obtained by cloning a major SPS expressed in onion leaf and exhaustively mining onion expressed sequence tag resources. SPS and SuSy loci were assigned to chromosome 8 and 6, respectively, using AALs and linkage mapping. Further loci were assigned, using AALs, to chromosomes 1 (sucrose phosphate phosphatase), 2 (SuSy and three invertases) and 8 (neutral invertase). The concordance between chromosome 8 localization of SPS and elevated leaf sucrose levels conditioned by high fructan alleles at the Frc locus in bulb onion or alien monosomic additions of chromosome 8 in A. fistulosum suggest that the Frc locus may condition variation in SPS activity.     

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.     

Exogenous Application of <Emphasis Type="Italic">Ethrel</Emphasis> and Gibberellic Acid Stimulates Physiological Growth of Late Planted Sugarcane with Short Growth Period in Sub-tropical India

Physiological growth of late planted sugarcane crop is restricted by high temperature and a short growth period. This causes considerable reduction in crop and sucrose yields. Improving physiological growth within the short period is, therefore, highly desirable. Two field experiments were undertaken to determine the effect of exogenous applications of Ethrel and gibberellic acid (GA₃) on sprouting, shoot population and physiological growth. Sugarcane setts were soaked overnight in Ethrel before planting. Foliar application of GA₃ was performed at 90, 120 and 150 days after planting (DAP). Ethrel soaking led to 100% sprouting and high settling population at 20 DAP, due to a significant increase in bud moisture and activities of acid invertase (AI), indole acetic acid oxidase (IAAO), adenosine triphosphatase (ATPase), superoxide dismutase (SOD) and nitrate reductase (NR) activity in vivo. Early sprouting increased the growth period to 245 days compared to 220 days in the unsoaked setts. The applications increased leaf area (57%), leaf area index (76%), leaf area ratio (71%), leaf area duration (48%), biomass duration (52%) and net assimilation rate (69.64%) at the grand growth stage. The changes led to increased shoot numbers (26.3%), internodal numbers stalk^?1 (40.74%), internodal length (40%), internodal girth (46.15%) and stalk length (42%) at the harvest stage. The stimulated physiological growth augmented dry matter content, ^oBrix and purity of cane juice by 24.2, 3 and 0.3%, respectively. The study demonstrates that the induction of higher shoot numbers together with increased leaf area index (LAI) and stalk elongation within a short growth period through Ethrel soaking and gibberellic acid applications is positively associated with enhanced dry matter and sucrose contents.     

A role for 'futile cycles' involving invertase and sucrose synthase in sucrose metabolism of tomato fruit.

Current concepts of the factors determining sink strength and the subsequent regulation of carbohydrate metabolism in tomato fruit are based upon an understanding of the relative roles of sucrose synthase, sucrose phosphate synthase and invertase, derived from studies in mutants and transformed plants. These enzymes participate in at least four futile cycles that involve sugar transport between the cytosol, vacuole and apoplast. Key reactions are (1) the continuous rapid degradation of sucrose in the cytosol by sucrose synthase (SuSy), (2) sucrose re-synthesis via either SuSy or sucrose phosphate synthase (SPS), (3) sucrose hydrolysis in the vacuole or apoplast by acid invertase, (4) subsequent transport of hexoses to the cytosol where they are once more converted into sucrose, and (5) rapid synthesis and breakdown of starch in the amyloplast. In this way futile cycles of sucrose/hexose interchange govern fruit sugar content and composition. The major function of the high and constant invertase activity in red tomato fruit is, therefore, to maintain high cellular hexose concentrations, the hydrolysis of sucrose in the vacuole and in the intercellular space allowing more efficient storage of sugar in these compartments. Vacuolar sugar storage may be important in sustaining fruit cell growth at times when less sucrose is available for the sink organs because of exhaustion of the carbohydrate pools in source leaves.     

Partial purification and characterisation of sucrose synthase in sugarcane

Three sucrose synthase (SuSy) (EC 2.4.1.13) forms were isolated from sugarcane leaf roll tissue. During anion exchange chromatography, one peak of activity (SuSyA) eluted during the wash step and the other peak (SuSyB) during the salt gradient phase at 180mM KCl concentration. A third form of activity (SuSyC), which also eluted at 180mM KCl, was also present in the leaf roll and replaced SuSyB depending on the season of the year. Substrate Km values, as well as sucrose breakdown/synthesis ratios, differed between these forms. For SuSyA, SuSyB, and SuSyC, respectively, Km values+/-SE (mM) were: 41.8+/-3.4, 109+/-23, and 35.9+/-2.3 for sucrose, 1.07+/-0.08, 0.214+/-0.039, and 0.00191+/-0.00019 for UDP, 6.62+/-1.55, 11.7+/-2.6, and 6.49+/-0.61 for fructose, and 3.59+/-0.37, 0.530+/-0.142, and 0.234+/-0.025 for UDP-glucose. Sucrose breakdown/synthesis ratios+/-SE were 0.0791+/-0.0199, 0.330+/-0.180, and 0.426+/-0.069 for SuSyA, SuSyB, and SuSyC, respectively. The ratio of the area of peak 1 (low breakdown/synthesis ratio) to the area of peak 2 (high breakdown/synthesis ratio) in sucrose accumulating tissue (internode 9) was 0.88, while in non-accumulating (leaf roll) tissue it was 14.5 at the same time of year. The molecular mass of the denatured subunits of all three forms was 94kDa by SDS-PAGE. A polyclonal antiserum raised against SuSyB cross-reacted with all three forms on an immunoblot, but only SuSyA and SuSyB were immunoinactivated by this serum.     

Effect of water deficit on carbohydrate status and enzymes of carbohydrate metabolism in seedlings of wheat cultivars

The effect of water deficit on carbohydrate status and enzymes of carbohydrate metabolism (alpha and beta amylases, sucrose phosphate synthase, sucrose synthase, acid and alkaline invertases) in wheat (Triticum aestivum L.) was investigated in the seedlings of drought-sensitive (PBW 343) and drought-tolerant (C 306) cultivars. The water deficit was induced by adding 6% mannitol (water potential -0.815 Mpa) in the growth medium. The water deficit reduced starch content in the shoots of tolerant seedlings as compared to the sensitive ones, but increased sucrose content in the shoots and roots of tolerant seedlings, indicating their protective role during stress conditions. It also decreased the alpha-amylase activity in the endosperm of seedlings of both the cultivars, but increased alpha and beta amylase activities in the shoots of tolerant ones. Sucrose phosphate synthase (SPS) activity showed a significant increase at 6 days of seedling growth (DSG) in the shoots of stressed seedlings of tolerant cultivar. However, SPS activity in the roots of stressed seedlings of sensitive cultivar was very low at 4 DSG and appeared significantly only at day 6. Sucrose synthase (SS) activity was lower in the shoots and roots of stressed seedlings of tolerant cultivar than sensitive ones at early stage of seedling growth. Higher acid invertase activity in the shoots of seedlings of tolerant cultivar appeared to be a unique characteristic of this cultivar for stress tolerance. Alkaline invertase activity, although affected under water deficit conditions, but was too low as compared to acid invertase activity to cause any significant affect on sucrose hydrolysis. In conclusion, higher sucrose content with high SPS and low acid invertase and SS activities in the roots under water deficit conditions could be responsible for drought tolerance of C 306.     

Effects of potassium deficiency on the enzymatic changes in developing cotton fibers

Although effects of potassium (K) on cotton growth have been explored extensively, the effects of K deficiency on the physiological changes closely related to cotton fiber development are lacking. Thus, a 2-year field experiment was conducted with two cotton cultivars (Simian 3 and Siza 3) under 0 kg K₂O ha^?1 (K deficiency) and 300 kg K₂O ha^?1 (K sufficiency). The results showed that tonoplast adenosine triphosphatase (V-ATPase), pyrophosphatase (PPase), plasma membrane H⁺-ATPase (PM H⁺-ATPase), phosphoenolpyruvate carboxylase (PEPC), sucrose synthesis (SuSy) and vacuolar invertase (V-INV) were highly sensitive to K deficiency. The decreases in those enzymes resulted in low malate and soluble sugar contents, which together with low K concentration declined the driving force for fiber elongation, leading to significantly lower fiber length in the 0 kg K₂O ha^?1 treatment. The activity of sucrose phosphate synthase (SPS) was obviously increased by K deficiency before 20 days post anthesis (DPA), which could partly explain the acceleration of fiber cellulose synthesis and the increase in fiber strength in the 0 kg K₂O ha^?1 treatment in the early stage. However, SPS activity was decreased by K deficiency after 20 DPA and SuSy activity was reduced by K deficiency at any sampling date, resulting in low fiber strength in the end. Compared with Simian 3, the enzymes V-ATPase, PPase, PM H⁺-ATPase, PEPC and SuSy during fiber elongation stage were more sensitive to K deficiency in Siza 3, and the enzymes SuSy and SPS during fiber-thickening stage were more sensitive to K deficiency in Siza 3, which were the important reasons causing greater decreases in final fiber length and final fiber strength for Siza 3 than Simian 3 under K deficiency.     

Specificity of sugar cane trehalase

An extract containing trehalase and invertase was prepared from apical internodes of sugar cane. The extract hydrolysed three glucosides: maltose, trehalose and sucrose. By reprecipitation with ammonium sulphate, maltase and trehalase activities appear to be due to different enzymes. As was also shown by differential inhibition and activation and by studies on the behaviour of both enzymes during growth, invertase and trehalase activities are attributed to different enzymes whose activities do not overlap. Invertase-free preparations confirm these results. Sucrose is a simple competitive inhibitor of sugar cane trehalase, excluding a regulatory role for this sugar. Sucrose was found at inhibitory levels in the first four apical internodes. A close correlation between sugar cane growth and invertase and trehalase levels was found in the apical internodes. Invertase has the greatest activity during growing, and trehalase reaches a maximum at maturity, prior to the flowering process. The high levels of trehalase in the flower suggest that the enzyme is involved in flowering or in related processes linked to seed formation.