Hormonal Control of Sucrose Phosphate Synthase and Sucrose Synthase in Sugar Beet

We studied the effects of synthetic analogs of phytohormones (benzyladenine, IAA, and GA) on the activities of the enzymes catalyzing sucrose synthesis and metabolism, sucrose phosphate synthase (SPS, EC 2.4.1.14) and sucrose synthase (SS, EC 2.4.1.13), and on the content of chlorophyll and protein during the sugar-beet (Beta vulgaris L.) ontogeny. Plant spraying with phytohormonal preparations activated SPS in leaves; direct interaction between phytohormones and the enzyme also increased its activity. The degree of this activation differed during the ontogeny and in dependence on the compound used for treatment. Analogs of phytohormones maintained high protein level in leaves, retarded chlorophyll breakdown, and, thus, prolonged leaf functional activity during development. Phytohormonal preparations practically did not affect the SS activity both after plant treatment and at their direct interaction with the enzyme. It is supposed that the SS activity in sugar-beet roots is controlled by sucrose synthesized in leaves rather than by phytohormones. The effects of hormones on leaf metabolism were mainly manifested in growth activation.     

蔗糖磷酸合成酶研究的新进展

蔗糖磷酸合成酶(sucrose phosphate synthase,SPS)是高等植物体内控制蔗糖合成的关键酶之一,它主要通过异构调节和磷酸化修饰在酶水平调节蔗糖合成。本文简要介绍SPS家族的成员、SPS蛋白上的3个磷酸化位点,以及SPS的生物学功能、SPS与磷酸蔗糖磷酸酶的关系等。     

alpha-Amylase Isozymes in Gibberellic Acid-treated Barley Half-seeds

The presence of multiple forms of α-amylase in gibberellic acid-treated embryoless barley half-seeds was demonstrated by separation on diethylaminoethyl-Sephadex and isoelectric focusing polyacrylamide gel disc electrophoresis. Two major α-amylase fractions (A and B), each consisting of two to three isozyme components, were purified. α-Amylase fractions A and B were distinguishable in their reaction patterns. The optimal pH of fraction A α-amylase was found to reside in the acidic side (pH 5.0), as was determined by analyzing the reducing sugars formed as well as the paper chromatographic detection of reaction products. At neutral pH, 6.9, fraction A exhibited weak amylolytic activity in forming maltose. The α-amylase activity in fraction A was markedly stimulated by heat treatment (70 C/15 minutes). Fraction B, constituting a major part of amylases in the endosperm extract, was also found to be composed of α-amylase, as evidenced by the loss of enzyme activity upon allowing fractions A and B to stand at pH 3.3 for a prolonged period. The possible physiological function of the two different types of α-amylase in the carbohydrate breakdown of barley seeds is discussed.     

植物蔗糖磷酸合成酶研究进展

蔗糖磷酸合成酶(Sucrose Phosphate Synthase,以下简称SPS)是植物体内控制蔗糖合成的关键酶。植物体内蔗糖的积累与SPS活性正相关,SPS还参与植物的生长和产量形成,并在植物的抗逆过程中起重要作用。高等植物中至少存在A、B、C三个家族的SPS,而禾本科植物至少存在A、B、C、DIII和DIV五个家族的SPS。不同植物体内不同家族的SPS基因的表达特性不同,它们所发挥的功能也存在差异。SPS的活性在基因表达调控和SPS蛋白磷酸化共价修饰作用两个层面受到植物生长发育、光照、代谢产物、外源物质如激素和糖类等多种因素的复杂调控。转基因研究表明,转SPS基因是提高作物产量和品质、增强作物抗逆性的有效途径,值得深入研究。全面总结了国内外在植物蔗糖磷酸合成酶方面的研究进展,并提出问题与研究展望,期望为进一步研究并利用植物SPS基因改良作物品种提供参考。     

The Role of Sucrose Synthase in Sink Organs of Woody Plants

Russian Journal of Plant Physiology - Sucrose synthase (EC 2.4.1.13) (SS) is the enzyme that regulates participation of sucrose in metabolism of the cells of the cambial zone of woody plants,...     

Sucrose Synthase: Expanding Protein Function

Sucrose synthase (SUS: EC 2.4.1.13), a key enzyme in plant sucrose catabolism, is uniquely able to mobilize sucrose into multiple pathways involved in metabolic, structural, and storage functions. Our research indicates that the biological function of SUS may extend beyond its catalytic activity. This inference is based on the following observations: (a) tissue-specific, isoform-dependent and metabolically-regulated association of SUS with mitochondria and (b) isoform-specific and anoxia-responsive interaction of SUS with the voltage-dependent anion channel (VDAC), the major outer mitochondrial membrane protein. More recent work shows that both VDAC and SUS are also localized to the nucleus in maize seedling tissues. Their intricate regulation under anoxia indicates that these two proteins may have a role in inter-compartmental signaling.Key Words: sucrose synthase, mitochondria, nucleus, localization, voltage-dependent anion channel (VDAC), non-catalytic rolesThe biochemical function of a protein is encoded within its primary sequence and can often be deciphered by simple in vitro assays. The cellular or organismal function of a protein is frequently the same as its biochemical activity. However, for many proteins, the biological function cannot be easily derived based on its biochemical function. This appears to be particularly true when the gene encoding the protein has a history of duplication and is represented by a family of paralogs. In maize and other species, sucrose synthase (SUS) isoforms are almost identical in their catalytic properties.^1,2 However, the characteristic phenotypes of mutants in specific isoforms suggest that the isoforms contribute to vastly different organismal functions.^2–4 Our interest is to identify the range of functions that maize SUS isoforms may have and elucidate the molecular basis of this functional diversity. Although expression divergence and consequent variation in their cellular abundance significantly contributes to this diversity,⁵ other factors such as intracellular distribution, post-translational modifications and interacting partners,^3,4,6,7 seem to be equally critical for the functional diversification of different SUS isoforms.Our study, spurred by a bioinformatics prediction, opened up a new facet of SUS biology, in that the protein may have organelle-based functions.⁸ Our analysis indicated that two of the three maize SUS isoforms (SH1 and SUS1) partly localize to mitochondria and nuclei, compartments not related to sucrose metabolism. In addition to this isoform-specificity, the compartmentation of SUS isoforms is influenced by developmental as well as environmental cues. Furthermore, its isoform-specific interaction with the voltage-dependent anion channel (VDAC) and an apparent conservation of SUS mitochondrial targeting across plant species suggest that SUS may have novel, noncatalytic biological functions. Our recent work shows that along with SUS, VDAC is also localized to the nucleus and these two proteins are inversely regulated in these two compartments under anoxic stress, indicating SUS-VDAC interaction may play a role in inter-compartmental signaling (Fig. 1).Open in a separate windowFigure 1Current working model of SUS-VDAC interactions in maize root tip cells. Prolonged anoxia leads to de-oligomerization of VDAC and the release of SUS from mitochondria, resulting in the migration of SUS to the nucleus. We hypothesize that the nuclear accumulation of SUS signals the induction of cell death pathway leading to the death of the root tip in anoxic maize seedlings. The insets show the primary root tip and a part of the axis from aerobic and anoxic seedlings. The root tip death is indicated by Evans Blue staining pattern of the anoxic root. ≠ = SUS. □ = VDAC.SUS mitochondrial localization also provided us an opportunity to reinterpret the phylogeny of sucrose metabolism. The proposed origin of sucrose metabolism is equivocal between the proteobacterial and cyanobacterial lineages.^9,10 Our discovery of SUS inside mitochondria, absence of plastid-bound SUS or plastid-targeting information in any of the plant SUS proteins and occurrence of mitochondrial targeting information in proteobacterial SUS orthologs strongly support a proteobacterial origin of plant sucrose synthases.⁸ Based on a genome-wide analysis of E. coli proteins, Lucattini et al.¹¹ proposed that mitochondrial targeting information may have been derived from the preexisting sequences of the endosymbiont proteins. We hypothesize that, in addition to the structural features needed for mitochondrial association, the functional basis of SUS-VDAC interaction may have been recruited by plants from the prokaryotic SUS genes. Based on striking similarities between bacterial and mitochondrial porins in their structure as well as regulation by purine nucleotides and their role in the host-cell death as modulated by cellular ATP levels, Frade and Michaelidis¹² speculated that the eukaryotic programmed cell death may have been a consequence of acquiring aerobic metabolism via the endosymbiotic process. Is organellar SUS a part of this acquisition?     

Sucrose Transport in Isolated Immature Barley Embryos

Techniques have been devised to select immature barley embryosat various stages in their development, and to study their accumulationof sucrose in vitro. Isolated embryos accumulate sucrose overa period of several hours of which some 80 per cent is conservedas a pool of free sucrose and the remainder utilized in macromolecularsynthesis. The rate of sucrose uptake increases with embryodevelopment, however the specific activity of uptake remainsconstant, indicating that the transport processes are fullyoperative early in embryogenesis. From the kinetics of sucroseuptake it is deduced that facilitated transport predominatesat sucrose concentrations of 50 mM, while at higher concentrationspassive diffusion makes an increasing contribution to sucroseaccumulation. The substrate specificity and the sensitivityof sucrose transport to uncoupling agents, in addition to thestability of the pool of accumulated sucrose, are all indicativeof active transport playing a major role in the sucrose assimilationof developing barley embryos. Hordeum distichum (L.) Lam, barley, embryo, sucrose transport     

Sucrose-Phosphate Synthase,Sucrose Synthase,and Invertase in Sugar Beet Leaves

The activity of sucrose-phosphate synthase (SPS) in sugar beet (Beta vulgaris L.) leaves was shown to exceed considerably the synthesizing activity of sucrose synthase (SS). The rise in SPS activity was related to the daylight period; i.e., it was associated with the rate of photosynthesis. The highest SPS activity was characteristic of fully expanded source leaves. In young developing leaves (leaves expanded to less than half of their final size), which represent the sink organs, the SPS activity was 2.5 times lower. At all stages of leaf development, the synthesizing SS activity was rather low. The diurnal change of SS activity was independent of photosynthesis and showed a slight rise from 6:00–8:00 p.m. Under field conditions, the highest SPS activity was found in leaves in the terminal stage of their development (105-day-old plants); the synthesizing activity of SS showed little changes during this period. The activity of soluble acid invertase was characteristic of young leaves. In mature leaves, the activity of this enzyme correlated with the daylight period. These changes occurred on the background of low sucrose content in leaves. The regulation of SPS, SS, and invertase activity is discussed. It is supposed that compartmentation of these enzymes in the photosynthesizing cell is important for transport, metabolism, and the osmotic function of sucrose in leaves.     

Sucrose Synthase Expression during Cold Acclimation in Wheat

When wheat (Triticum aestivum) seedlings are exposed to a cold temperature (2-4°C) above 0°C, sucrose accumulates and sucrose synthase activity increases. The effect of a cold period on the level of sucrose synthase (SS) was investigated. Using antibodies against wheat germ SS, Western blots studies showed that the amount of the SS peptide increased during 14 days in the cold, when plants were moved from 23°C to 4°C. The level of SS diminished when plants were moved back to 23°C. Northern blots of poly(A)⁺ RNA, confirmed a five- to sixfold induction of SS in wheat leaves during cold acclimation. These results indicate that SS is involved in the plant response to a chilling stress.     

Sucrose Phosphate Synthase, Sucrose Synthase, and Invertase Activities in Developing Fruit of Lycopersicon esculentum Mill. and the Sucrose Accumulating Lycopersicon hirsutum Humb. and Bonpl

The green-fruited Lycopersicon hirsutum Humb. and Bonpl. accumulated sucrose to concentrations of about 118 micromoles per gram fresh weight during the final stages of development. In comparison, Lycopersicon esculentum Mill. cultivars contained less than 15 micromoles per gram fresh weight of sucrose at the ripe stage. Glucose and fructose levels remained relatively constant throughout development in L. hirsutum at 22 to 50 micromoles per gram fresh weight each. Starch content was low even at early stages of development, and declined further with development. Soluble acid invertase (EC 3.2. 1.26) activity declined concomitant with the rise in sucrose content. Acid invertase activity, which was solubilized in 1 molar NaCl (presumably cell-wall bound), remained constant throughout development (about 3 micromoles of reducing sugars (per gram fresh weight) per hour. Sucrose phosphate synthase (EC 2.4.1.14) activity was present at about 5 micromoles of sucrose (per gram fresh weight) per hour even at early stages of development, and increased sharply to about 40 micromoles of sucrose (per gram fresh weight) per hour at the final stages of development studied, parallel to the rise in sucrose content. In comparison, sucrose phosphate synthase activity in L. esculentum remained low throughout development. The possible roles of the sucrose metabolizing enzymes in determining sucrose accumulation are discussed.     

Sucrose Synthase in Legume Nodules Is Essential for Nitrogen Fixation

The role of sucrose synthase (SS) in the fixation of N was examined in the rug4 mutant of pea (Pisum sativum L.) plants in which SS activity was severely reduced. When dependent on nodules for their N supply, the mutant plants were not viable and appeared to be incapable of effective N fixation, although nodule formation was essentially normal. In fact, N and C resources invested in nodules were much greater in mutant plants than in the wild-type (WT) plants. Low SS activity in nodules (present at only 10% of WT levels) resulted in lower amounts of total soluble protein and leghemoglobin and lower activities of several enzymes compared with WT nodules. Alkaline invertase activity was not increased to compensate for reduced SS activity. Leghemoglobin was present at less than 20% of WT values, so O₂ flux may have been compromised. The two components of nitrogenase were present at normal levels in mutant nodules. However, only a trace of nitrogenase activity was detected in intact plants and none was found in isolated bacteroids. The results are discussed in relation to the role of SS in the provision of C substrates for N fixation and in the development of functional nodules.     

Photorespiratory Amino Donors, Sucrose Synthesis and the Induction of CO2 Fixation in Barley Deficient in Glutamine Synthetase and/or Glutamate Synthase

Murray, A. J. S., Black well, R. D., Lea, P. J. and Joy, K.W. 1988. Photorespiratory amino donors, sucrose synthesis andthe induction of CO₂ fixation in barley deficient in glutaminesynthetase and/or glutamate synthase.—J. exp. Bot. 39:845–858. A number of mutants of barley have been produced which lackboth chloroplastic glutamine synthetase and ferredoxin-dependentglutamate synthase activities. The plants accumulated ammoniato the same extent as mutants deficient in only glutamine synthetasebut shared the gas-exchange characteristics of the glutamatesynthase deficient parent. These mutants have been used to demonstratedirectly the ability of alanine to ameliorate the dramatic dropin fixation rate normally exhibited by glutamate synthase deficientmutants on transfer to photorespiratory conditions. Immediatelyafter transfer to air, the mutants deficient in glutamate synthaseactivity demonstrated a reduced ability to incorporate ¹⁴C derivedfrom ¹⁴CO₂ into sucrose. This effect was, however, dependenton the previous induction of CO₂ fixation. Use of ¹⁴CO₂ revealedthat the induction phase of CO₂ fixation was altered in allthree mutants. Neither of the parents nor the double mutantaccumulated sucrose in air under conditions which promote sucroseaccumulation by the wild type. The implications of these resultsfor photosynthesis and the control of sucrose synthesis arediscussed. Key words: Photorespiratory barley mutant, amino donors, sucrose, GS, glutamate synthase.     

Activities of Sucrose Synthase and Acid Invertase in Pea Seedling Organs

The organ topography of sucrose synthase and soluble acid invertase in pea seedlings at heterotrophic stage (3–14 days) was studied. Sucrose synthase was most active in the roots, with the highest activity on the 6–8th days. In the leaves, its activity decreased from day 3 to day 14. In the stems, sucrose synthase activity was at an invariantly low level. The patterns of sucrose synthase activity in etiolated and green plants were similar. As distinct from sucrose synthase, invertase activity was the highest in the stem, especially in etiolated plants. The peak of its activity was observed on the 6-8th days. In the leaves, invertase activity was lower but its pattern was the same. In the roots, acid invertase activity decreased from the 3rd day and did not depend on illumination. The conclusion is that differences in sucrose synthase and acid invertase activities in roots, leaves, and stem are determined by differences in the import of hydrolytic products of stored compound from the cotyledons as well as by different demands of these organs for these products for the processes of organ expansion and for the maintenance of organ metabolism.     

苹果中磷酸蔗糖合酶家族基因的表达特性及其与蔗糖含量的关系

磷酸蔗糖合酶(sucrose phosphate synthase,SPS)是植物中蔗糖合成的主要限速酶,影响植物的生长发育和果实中蔗糖的含量。为探明苹果中SPS基因家族特性及其在蔗糖合成中的作用,该研究从苹果基因组中分离了MdSPS家族基因,分析了它们的进化关系以及mRNA表达特性与酶活性和蔗糖含量的关系。结果显示:(1)在苹果基因组中有8个SPS家族基因表达,它们分别属于双子叶植物的3个SPS亚家族。(2)荧光定量PCR分析显示,苹果C类的MdSPS6基因和A类的MdSPS1a/b基因是苹果中表达丰度最高的SPS基因成员,其中MdSPS6在苹果成熟果中表达丰度最高,其次是成熟叶片,而MdSPS1a/b在不积累蔗糖的幼果中表达丰度最高。(3)在果实发育过程中,除MdSPS1a/b之外,其它5个苹果MdSPS家族基因均随果实的生长表达丰度增加,与SPS活性和蔗糖含量明显呈正相关关系。研究表明,C类家族MdSPS6是苹果果实发育后期和叶片中蔗糖合成的主要SPS基因。     

植物水溶性蔗糖合成酶生物信息学分析初探

用生物信息学方法对已在GenBank上注册的黑麦草、绿竹、菜豆、马铃薯、颤杨等植物水溶性蔗糖合成酶基因的核苷酸序列以及推导的氨基酸序列、组成成分、氨基酸翻译后修饰、导肽、跨膜拓朴结构域、疏水性／亲水性、蛋白质二级结构以及功能结构域等进行分析预测和推断的结果表明,这些植物的水溶性蔗糖合成酶位于线粒体中,是非跨膜的亲水性蛋白,α-螺旋和不规则卷曲是其蛋白质二级结构的主要结构元件,β-转角和延伸链散布于整个蛋白质中,包含2个功能结构域,即蔗糖合成功能域和糖基化合物转移功能域。     

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.