Suc Phosphate Synthase Research Articles

Effects of phosphorus on fruit soluble sugar and citric acid accumulations in citrus

Phosphorus (P) is one of the essential macro-elements for plants. Sugar and organic acid are important factors affecting sensory characteristics of citrus fruit quality. The aim of this study was to investigate how P fertilizer affects quality improvement particularly sucrose (Suc), fructose (Fru), glucose (Glu) and citric acid (CA) accumulations in Cara Cara navel. P fertilizer improved fruit quality of Cara Cara navel, as supported by decreasing titratable acid (TA), CA and increasing soluble solid (TSS), sugars and the ratio of TSS and TA. At the early stage of fruit development, P fertilizer had greater roles in degrading Suc into Fru and Glu due to the increased activities of Suc-degrading enzymes including acid invertase, neutral invertase and Suc synthase-cleavage activity. Coversely, at the mid and late stages of fruit development, P fertilizer had greater roles in re-synthesizing Suc due to the increased activities of Suc-synthesizing enzymes including Suc phosphate synthase and Suc synthase-synthetic activity. These results indicated that application of P fertilizer increased soluble sugars concentrations by improving Suc metabolism and sink strength in fruit conferred by the upregulations of the activities of Suc-degrading and Suc-synthesizing enzymes. P fertilizer decreased CA accumulations at least partially by inhibiting synthesis of CA due to the decreased activities of CA-synthesizing enzymes including citrate synthetase and phosphoenolpyruvate carboxylase. This study suggested that P fertilizer, particularly fertilized with 0.40 kg/plant, increased soluble sugars but decreased CA accumulations in citrus fruit.

Sucrose metabolism in developing oil-rich tubers of Cyperus esculentus: comparative transcriptome analysis

BackgroundCyperus esculentus is unique in that it can accumulate significant amounts of oil, starch and sugar as major storage reserves in tubers with high tuber yield and therefore considered as a novel model to study carbon allocation into different storage reserves in underground sink tissues such as tubers and roots. Sucrose (Suc) plays a central role in control of carbon flux toward biosynthesis of different storage reserves; however, it remains unclear for the molecular mechanism underlying Suc metabolism in underground oil-rich storage tissues. In the present study, a comprehensive transcriptome analysis of C. esculentus oil tuber compared to other plant oil- or carbohydrate-rich storage tissues was made for the expression patterns of genes related to the Suc metabolism.ResultsThe results revealed some species-specific features of gene transcripts in oil tuber of C. esculentus, indicating that: (i) the expressions of genes responsible for Suc metabolism are developmentally regulated and displayed a pattern dissimilar to other plant storage tissues; (ii) both of Suc breakdown and biosynthesis processes might be the major pathways associated with Suc metabolism; (iii) it was probably that Suc degradation could be primarily through the action of Suc synthase (SUS) other than invertase (INV) during tuber development. The orthologs of SUS1, SUS3 and SUS4 are the main SUS isoforms catalyzing Suc breakdown while the vacuolar INV (VIN) is the leading determinant controlling sugar composition; (iv) cytosolic hexose phosphorylation possibly relies more on fructose as substrate and uridine diphosphate glucose pyrophosphorylase (UGP) plays an important role in this pathway; (v) it is Suc-phosphate synthase (SPS) B- and C-family members rather than SPS A that are the principal contributors to SPS enzymes and play crucial roles in Suc biosynthesis pathway.ConclusionsWe have successfully identified the Suc metabolic pathways in C. esculentus tubers, highlighting several conserved and distinct expressions that might contribute to sugar accumulation in this unique underground storage tissue. The specific and differential expression genes revealed in this study might indicate the special molecular mechanism and transcriptional regulation of Suc metabolism occurred in oil tubers of C. esculentus.

Loss of the two major leaf isoforms of sucrose-phosphate synthase in Arabidopsis thaliana limits sucrose synthesis and nocturnal starch degradation but does not alter carbon partitioning during photosynthesis

Sucrose (Suc)-phosphate synthase (SPS) catalyses one of the rate-limiting steps in the synthesis of Suc in plants. The Arabidopsis genome contains four annotated SPS genes which can be grouped into three different families (SPSA1, SPSA2, SPSB, and SPSC). However, the functional significance of this multiplicity of SPS genes is as yet only poorly understood. All four SPS isoforms show enzymatic activity when expressed in yeast although there is variation in sensitivity towards allosteric effectors. Promoter-reporter gene analyses and quantitative real-time reverse transcription-PCR studies indicate that no two SPS genes have the same expression pattern and that AtSPSA1 and AtSPSC represent the major isoforms expressed in leaves. An spsa1 knock-out mutant showed a 44% decrease in leaf SPS activity and a slight increase in leaf starch content at the end of the light period as well as at the end of the dark period. The spsc null mutant displayed reduced Suc contents towards the end of the photoperiod and a concomitant 25% reduction in SPS activity. In contrast, an spsa1/spsc double mutant was strongly impaired in growth and accumulated high levels of starch. This increase in starch was probably not due to an increased partitioning of carbon into starch, but was rather caused by an impaired starch mobilization during the night. Suc export from excised petioles harvested from spsa1/spsc double mutant plants was significantly reduced under illumination as well as during the dark period. It is concluded that loss of the two major SPS isoforms in leaves limits Suc synthesis without grossly changing carbon partitioning in favour of starch during the light period but limits starch degradation during the dark period.

First evidence of sucrose biosynthesis by single cyanobacterial bimodular proteins

The net synthesis of sucrose (Suc) is catalysed by the sequential action of Suc-phosphate synthase (SPS) and Suc-phosphate phosphatase (SPP). SPS and SPP from Anabaena sp. PCC 7120 (7120-SPS and 7120-SPP) define minimal catalytic units. Bidomainal SPSs, where both units are fused, occur in plants and cyanobacteria, but they display only SPS activity. Using recombinant proteins that have fused 7120-SPS and 7120-SPP, we demonstrated that they are bifunctional chimeras and that the arrangement 7120-SPS/SPP is the most efficient to catalyse the sequential reactions to yield Suc. Moreover, we present the first evidence of a bidomainal SPS present in the cyanobacterium Synechococcus elongatus PCC 7942 with both, SPS and SPP activity.

Differential regulation of sorbitol and sucrose loading into the phloem of Plantago major in response to salt stress.

Several plant families generate polyols, the reduced form of monosaccharides, as one of their primary photosynthetic products. Together with sucrose (Suc) or raffinose, these polyols are used for long-distance allocation of photosynthetically fixed carbon in the phloem. Many species from these families accumulate these polyols under salt or drought stress, and the underlying regulation of polyol biosynthetic or oxidizing enzymes has been studied in detail. Here, we present results on the differential regulation of genes that encode transport proteins involved in phloem loading with sorbitol and Suc under salt stress. In the Suc- and sorbitol-translocating species Plantago major, the mRNA levels of the vascular sorbitol transporters PmPLT1 and PmPLT2 are rapidly up-regulated in response to salt treatment. In contrast, mRNA levels for the phloem Suc transporter PmSUC2 stay constant during the initial phase of salt treatment and are down-regulated after 24 h of salt stress. This adaptation in phloem loading is paralleled by a down-regulation of mRNA levels for a predicted sorbitol dehydrogenase (PmSDH1) in the entire leaf and of mRNA levels for a predicted Suc phosphate synthase (PmSPS1) in the vasculature. Analyses of Suc and sorbitol concentrations in leaves, in enriched vascular tissue, and in phloem exudates of detached leaves revealed an accumulation of sorbitol and, to a lesser extent, of Suc within the leaves of salt-stressed plants, a reduced rate of phloem sap exudation after NaCl treatment, and an increased sorbitol-to-Suc ratio within the phloem sap. Thus, the up-regulation of PmPLT1 and PmPLT2 expression upon salt stress results in a preferred loading of sorbitol into the phloem of P. major.

Light and Metabolic Signals Control the Selective Degradation of Sucrose Synthase in Maize Leaves during Deetiolation

The content and activity of Suc (Suc) synthase (SUS) protein is high in sink organs but low in source organs. In this report, we examined light and metabolic signals regulating SUS protein degradation in maize (Zea mays) leaves during deetiolation. We found that SUS protein accumulated in etiolated leaves of the dark-grown seedlings but was rapidly degraded upon exposure to white, blue, or red light. This occurred concurrent with the accumulation of photosynthetic enzymes, such as Rubisco and Rubisco activase, and enzymes of Suc biosynthesis such as Suc-phosphate synthase. Deetiolation-induced SUS degradation was not inhibited by the proteasome inhibitor MG132. Moreover, neither full-length nor truncated SUS phosphorylated at the serine-170 site was found in the crude 26S proteasome fraction (150,000g postmicrosomal pellet) isolated in the presence of MG132. However, SUS degradation was strongly inhibited by feeding cycloheximide or amino acids to detached leaves, while Suc feeding had no effect. Of the amino acids tested, exogenous glutamate had the greatest effect. Collectively, these results demonstrate that SUS protein degradation during deetiolation: (1) is selective; (2) can be triggered by either blue- or red light-mediated signaling pathways; (3) does not involve the 26S proteasome; and (4) is inhibited by free amino acids. These findings suggest that SUS degradation is important to supply residues for the synthesis of other proteins required for autotrophic metabolism.

Differential expression of sucrose-phosphate synthase isoenzymes in tobacco reflects their functional specialization during dark-governed starch mobilization in source leaves.

Sucrose (Suc)-phosphate synthase (SPS) plays a crucial role in the synthesis of Suc in photosynthetic and nonphotosynthetic tissues. Several isoforms of SPS exist in dicotyledonous plants that can be grouped into the different families A, B, and C. To explore whether functional differences between the SPS gene families might exist, we characterized a representative for each family from tobacco (Nicotiana tabacum). RNA-blot analysis revealed a distinct expression pattern for each of the three SPS genes. While the A-family member (NtSPSA) was found to be expressed in all tissues examined, expression of the B isoform (NtSPSB) was mainly confined to the reproductive organs and NtSPSC mRNA was exclusively detected in mature source leaves. We used RNA interference to assess the in planta function of NtSPSA and C. While silencing of NtSPSA had no detectable influence on leaf carbohydrate metabolism, reduction of NtSPSC led to an increase in leaf starch content by a factor of 3 to 8. Further analysis revealed that starch accumulation in NtSPSC-silenced plants was not due to an increased partitioning of carbon into starch, but rather showed that starch mobilization was impaired. The transgenic plants were unable to efficiently mobilize their transitory leaf starch during a prolonged period of darkness and accumulated maltose as a major intermediate of starch breakdown. NtSPSC mRNA level increased appreciably during the dark period while transcript levels of the other isoforms showed no diurnal changes. Together, these results suggest that NtSPSC is specifically involved in the synthesis of Suc during starch mobilization in the dark. The roles of the other SPS isoforms are discussed.

Evolution and function of the sucrose-phosphate synthase gene families in wheat and other grasses.

Suc-phosphate synthase (SPS) is a key regulatory enzyme in the pathway of Suc biosynthesis and has been linked to quantitative trait loci controlling plant growth and yield. In dicotyledonous plants there are three SPS gene families: A, B, and C. Here we report the finding of five families of SPS genes in wheat (Triticum aestivum) and other monocotyledonous plants from the family Poaceae (grasses). Three of these form separate subfamilies within the previously described A, B, and C gene families, but the other two form a novel and distinctive D family, which on present evidence is only found in the Poaceae. The D-type SPS proteins lack the phosphorylation sites associated with 14-3-3 protein binding and osmotic stress activation, and the linker region between the N-terminal catalytic glucosyltransferase domain and the C-terminal Suc-phosphatase-like domain is 80 to 90 amino acid residues shorter than in the A, B, or C types. The D family appears to have arisen after the divergence of mono- and dicotyledonous plants, with a later duplication event resulting in the two D-type subfamilies. Each of the SPS gene families in wheat showed different, but overlapping, spatial and temporal expression patterns, and in most organs at least two different SPS genes are expressed. Analysis of expressed sequence tags indicated similar expression patterns to wheat for each SPS gene family in barley (Hordeum vulgare) but not in more distantly related grasses. We identified an expressed sequence tag from rice (Oryza sativa) that appears to be derived from an endogenous antisense SPS gene, and this might account for the apparently low level of expression of the related OsSPS11 sense gene, adding to the already extensive list of mechanisms for regulating the activity of SPS in plants.

Sucrose Phosphate Synthase Activity Rises in Correlation with High-Rate Cellulose Synthesis in Three Heterotrophic Systems

Based on work with cotton fibers, a particulate form of sucrose (Suc) synthase was proposed to support secondary wall cellulose synthesis by degrading Suc to fructose and UDP-glucose. The model proposed that UDP-glucose was then channeled to cellulose synthase in the plasma membrane, and it implies that Suc availability in cellulose sink cells would affect the rate of cellulose synthesis. Therefore, if cellulose sink cells could synthesize Suc and/or had the capacity to recycle the fructose released by Suc synthase back to Suc, cellulose synthesis might be supported. The capacity of cellulose sink cells to synthesize Suc was tested by analyzing the Suc phosphate synthase (SPS) activity of three heterotrophic systems with cellulose-rich secondary walls. SPS is a primary regulator of the Suc synthesis rate in leaves and some Suc-storing, heterotrophic organs, but its activity has not been previously correlated with cellulose synthesis. Two systems analyzed, cultured mesophyll cells of Zinnia elegans L. var. Envy and etiolated hypocotyls of kidney beans (Phaseolus vulgaris), contained differentiating tracheary elements. Cotton (Gossypium hirsutum L. cv Acala SJ-1) fibers were also analyzed during primary and secondary wall synthesis. SPS activity rose in all three systems during periods of maximum cellulose deposition within secondary walls. The Z. elegans culture system was manipulated to establish a tight linkage between the timing of tracheary element differentiation and rising SPS activity and to show that SPS activity did not depend on the availability of starch for degradation. The significance of these findings in regard to directing metabolic flux toward cellulose will be discussed.

Function and dynamics of auxin and carbohydrates during earlywood/latewood transition in scots pine.

In temperate regions the annual pattern of wood development is characterized by the formation of radially narrow and thick walled latewood cells. This takes place at the later part of the growing season when cambial cell division declines. To gain new insight into the regulation of this process, micro-analytical techniques were used to visualize the distribution of indole-3-acetic acid (IAA), soluble carbohydrates, and activities of sucrose (Suc)-metabolizing enzymes across the cambial region tissues in Scots pine (Pinus sylvestris). The total amount of IAA in the cambial region did not change with latewood initiation. But its radial distribution pattern was altered, resulting in an increased concentration in the cambial meristem and its recent derivatives. Thus, initiation of latewood formation and cessation of cambial cell division is not a consequence of decreased IAA concentrations in dividing and expanding cells. Rather, IAA most likely has a role in defining the altered developmental pattern associated with latewood formation. Carbohydrates and enzyme activities showed distinctive radial distribution patterns. Suc peaked in the phloem and decreased sharply to low levels across the cambial zone, whereas fructose and glucose reached their highest levels in the maturing tracheids. Suc synthase was the dominating Suc cleaving enzyme with a peak in the secondary wall-forming tracheids and in the phloem. Soluble acid invertase peaked in dividing and expanding cells. Suc-phosphate synthase had its highest activities in the phloem. Activities of cell wall bound invertase were low. The absence of major seasonal variations indicates that carbohydrate availability is not a trigger for latewood initiation. However, steep concentration gradients of the sugars suggest a role for sugar signaling in vascular development.

High-temperature perturbation of starch synthesis is attributable to inhibition of ADP-glucose pyrophosphorylase by decreased levels of glycerate-3-phosphate in growing potato tubers

To investigate the short-term effect of elevated temperatures on carbon metabolism in growing potato (Solanum tuberosum L.) tubers, developing tubers were exposed to a range of temperatures between 19 degreesC and 37 degreesC. Incorporation of [14C]glucose (Glc) into starch showed a temperature optimum at 25 degreesC. Increasing the temperature from 23 degreesC or 25 degreesC up to 37 degreesC led to decreased labeling of starch, increased labeling of sucrose (Suc) and intermediates of the respiratory pathway, and increased respiration rates. At elevated temperatures, hexose-phosphate levels were increased, whereas the levels of glycerate-3-phosphate (3PGA) and phosphoenolpyruvate were decreased. There was an increase in pyruvate and malate, and a decrease in isocitrate. The amount of adenine diphosphoglucose (ADPGlc) decreased when tubers were exposed to elevated temperatures. There was a strong correlation between the in vivo levels of 3PGA and ADPGlc in tubers incubated at different temperatures, and the decrease in ADPGlc correlated very well with the decrease in the labeling of starch. In tubers incubated at temperatures above 30 degreesC, the overall activities of Suc synthase and ADPGlc pyrophosphorylase declined slightly, whereas soluble starch synthase and pyruvate kinase remained unchanged. Elevated temperatures led to an activation of Suc phosphate synthase involving a change in its kinetic properties. There was a strong correlation between Suc phosphate synthase activation and the in vivo level of Glc-6-phosphate. It is proposed that elevated temperatures lead to increased rates of respiration, and the resulting decline of 3PGA then inhibits ADPGlc pyrophosphorylase and starch synthesis.

Drought-induced effects on nitrate reductase activity and mRNA and on the coordination of nitrogen and carbon metabolism in maize leaves

Maize (Zea mays L.) plants were grown to the nine-leaf stage. Despite a saturating N supply, the youngest mature leaves (seventh position on the stem) contained little NO3- reserve. Droughted plants (deprived of nutrient solution) showed changes in foliar enzyme activities, mRNA accumulation, photosynthesis, and carbohydrate and amino acid contents. Total leaf water potential and CO2 assimilation rates, measured 3 h into the photoperiod, decreased 3 d after the onset of drought. Starch, glucose, fructose, and amino acids, but not sucrose (Suc), accumulated in the leaves of droughted plants. Maximal extractable phosphoenolpyruvate carboxylase activities increased slightly during water deficit, whereas the sensitivity of this enzyme to the inhibitor malate decreased. Maximal extractable Suc phosphate synthase activities decreased as a result of water stress, and there was an increase in the sensitivity to the inhibitor orthophosphate. A correlation between maximal extractable foliar nitrate reductase (NR) activity and the rate of CO2 assimilation was observed. The NR activation state and maximal extractable NR activity declined rapidly in response to drought. Photosynthesis and NR activity recovered rapidly when nutrient solution was restored at this point. The decrease in maximal extractable NR activity was accompanied by a decrease in NR transcripts, whereas Suc phosphate synthase and phosphoenolpyruvate carboxylase mRNAs were much less affected. The coordination of N and C metabolism is retained during drought conditions via modulation of the activities of Suc phosphate synthase and NR commensurate with the prevailing rate of photosynthesis.

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.

Physical and Kinetic Evidence for an Association between Sucrose-Phosphate Synthase and Sucrose-Phosphate Phosphatase.

The possible formation of a multienzyme complex between sucrose (Suc)-phosphate synthase (SPS) and Suc-phosphate phosphatase (SPP) was examined by measuring the rates of Suc-6-phosphate (Suc-6-P) synthesis and hydrolysis in mixing experiments with partially purified enzymes from spinach (Spinacia oleracea) and rice (Oryza sativa) leaves. The addition of SPP to SPS stimulated the rate of Suc-6-P synthesis. SPS inhibited the hydrolysis of exogenous Suc-6-P by SPP when added in the absence of its substrate (i.e. UDP-glucose) but stimulated SPP activity when the SPS substrates were present and used to generate Suc-6-P directly in the reaction. Results from isotope-dilution experiments suggest that Suc-6-P was channeled between SPS and SPP. A portion of the SPS activity comigrated with SPP during native polyacrylamide gel electrophoresis, providing physical evidence for an enzyme-enzyme interaction. Taken together, these results strongly suggest that SPS and SPP associate to form a multienzyme complex.

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.

Elevated Levels of Both Sucrose-Phosphate Synthase and Sucrose Synthase in Vicia Guard Cells Indicate Cell-Specific Carbohydrate Interconversions.

A long series of reports correlate larger stomatal aperture size with elevated concentration of sucrose (Suc) in guard cells. To assess the role and autonomy of guard cells with respect to these changes, we have determined quantitatively the cellular distribution of the synthetic enzyme, Suc-phosphate synthase (SPS) and the degradative enzyme Suc synthase (SS) in Vicia leaflet. As expected for Suc-exporting cells, the photosynthetic parenchyma had a high SPS:SS ratio of approximately 45. Also as expected, in epidermal cells, which had only few and rudimentary plastids, the SPS:SS ratio was low (0.4). Of all cells and tissues measured, those that had the highest specific activity of SPS (about 4.8 [mu]mol mg-1 of protein h-1) were guard cells. Guard cells also had a very high relative specific activity of SS.