Development of Sink Capacity of the "Storage Root" in a Radish Variety with a Low Ratio of "Storage Root" to Shoot

Controling mechanisms of sink capacity are poorly understood.Previously we suggested that sucrose synthase (SuSy), but notinvertase, plays an important role for sink capacity of theradish "storage root" in a variety, Raphanus sativus L. (cv.White Cherish) [plant Cell Physiol. (1999) 40: 369]. With thisvariety about 50% of the total dry weight (DW) was in the "storageroot" at 21 d after sowing (DAS). We investigated the sink capacityof another radish variety, R. sativus L. (cv. Kosena) with alow ratio of "storage root" to shoot. With the latter varietyonly 3% of the total DW was in the "storage root" at 21 DAS.Sink activity (increase in DW of the "storage root" per unitof DW present per unit of time) of the "storage root" in Kosenaas well as White Cherish was strongly related to the level andactivity of SuSy but not to the activity of invertase. Theseresults confirmed that SuSy rather than invertase may be criticalfor the development of the sink activity of the radish "storageroot" and that the reaction products of UDP-glucose and fructoseare utilized for sink growth including biosynthesis of the cellwall. In Kosena photosynthates seemed to be partitioned mainlyinto developing leaves and fibrous roots. Differences in partitioningof photosynthates among various sinks with these two varietiesare discussed including anatomical considerations. (Received July 19, 1999; Accepted September 30, 1999)     

Development of Sink Capacity of the "Storage Root" in a Radish Cultivar with a High Ratio of "Storage Root" to Shoot

The mechanisms that control sink capacity are poorly understood.in radish, a major sink is the "storage root", which beginsto thicken early in development, mainly as a result of thickeningof the hypocotyl. We investigated changes in the accumulationof dry matter, sink activity (increase in dry weight of thehypocotyl per unit of dry weight present per unit of time),carbohydrate content, levels of metabolites, activities of enzymesrelated to the breakdown of sucrose, and the profile of solubleproteins, as well as changes in anatomy, using hypocotyls ofa cultivar with a high ratio of "storage root" to shoot. Wefound that sink activity was strongly related to the level andactivity of sucrose synthase but not to the activity of invertase.We also found a significant correlation between sucrose contentand the level and activity of sucrose synthase. Our resultssuggest that sucrose synthase, but not invertase, might be criticalfor the development of the sink activity of the radish hypocotyland that the level of sucrose might regulate the expressionof sucrose synthase. A discussion of sink capacity is presentedthat includes consideration of structural changes in the hypocotyl. (Received December 14, 1998; Accepted January 27, 1999)     

Storage Organ Development in Radish (Raphanus sativus L.). 2. Effects of Growth Promoters on Cambial Activity in Cultured Roots, Decapitated Seedlings and Intact Plants

The radish varieties Cherry Belle and Long White Icicle wereused to investigate the role of the shoot and the effects ofsynthetic growth promoters in controlling cambial activity inthe seedling axis. Development was compared in excised roots, roots with hypocotylsattached and intact seedlings cultured aseptically on a nutrientmedium. No cambial divisions were seen in isolated radicleswhich had been cultured for ten days following excision butretention of hypocotyl tissue or the entire shoot resulted incambial activity and the production of secondary vascular tissues.Enriching the culture medium by raising the sucrose conantrationto 8% and including 10^–5 M indol-3yl acetic acid (IAA)5 x 10^–6 M 6-benzylaminopurine (BA) and 5 x 10^–4Minositol enhanced root thickening, increasing stele and xylemdiameters in roots cultured both with and without attached shoottissues. The effects of shoot tissues and enrichment of themedium were additive. The effects of auxin, cytokinin and gibberellin (gibberellicacid, GA²) were also studied on daxpitated seedlings. BA wasmuch more effective in inducing cell divisions in the hypocotylthan either IAA or GA supplied separately but a mixture of IAA+GAalso produced clearly defined arcs of cambial tissue. Littlesecondary tissue had been produced after seven days' treatment,and stelar enlargement was due to the development of a cambialzone and cell expansion in the primary tissues. Only minor differencesin response were observed between the two varieties. No stimulation of storage organ development occurred when auxin,cytokinin or inositol was inwrporated into the inorganic culturesolution in which plants of Cherry Belle were grown. Rnphanus sarivus, radish, storage organ, cambial activity, growth promoters, indol-3-ylacetic acid, 6-benzylaminopurine, gibberellic acid     

Changes in thiocyanate content in some radish Raphanus sativus L. cultivars during hypocotyl-root growth

Changes in the thiocyanate content in hypocotyl-roots and leaves of radish were observed in a two-year field experiment. Six cultivars were tested: early radish (Rex and Ostergruss Różowa), Japanese radish (Tokinashi and Minowase Summer Cross F₁), and winter radish (Monachijska Biała and Murzynka). A significant diversification in thiocyanate content among cultivars, plant parts, harvest dates and observation years was found. Early cultivars contained the least amount of these compounds, Murzynka — the greatest. The content of thiocyanates in leaves was 3 – 5 times higher than that in hypocotyl-roots. The changes in the thiocyanate content during root growth showed a constant rising tendency in the case of the leaves of all cultivars and the storage organs of Murzynka.     

Storage Organ Development in Radish (Raphanus sativus L.). 1. A Comparision of Development in Seedlings and Rooted Cuttings of Two Contrasting Varieties

Development was compared in two contrasting varieties of radishgrown in nutrient solution. Differences in shape of the long,tapering radish of Long White Icicle and the smaller, roundCherry Belle were associated with differences in the growthof the upper part of the tap root. Thickening was the resultof , cambial activity and transverse scctions through the upperroot showed differences in both cell number and a l l size.Long White Icicle contained more cells in the secondary xylemand phloem/secondary cortex than Cherry Belle, and the non-lignifiedxylem parenchyma cells were also larger in Icicle. The upperpart of the tap root also elongated in Icicle but not in CherryBelle, and this was entirely due to an increase in cell numberin the longitudinal plane. Removing the distal part of the root system had little effecton storage organ size unless most of the root system was removed.Adventitious roots on seedling and shoot-tip cuttings of bothvarieties were capable of thickening and the single root developingon shoot-tip cuttings of Cherry Belle achieved a diameter greaterthan that of the radicle of the intact plant. Rooted cotyledonsof Cherry Belle cxrasionally produced small storage organs. Raphanus sativus, radish storage organ, root thickening, cambial activity     

Localization of sucrose synthase in wheat roots: increased in situ activity of sucrose synthase correlates with cell wall thickening by cellulose deposition under hypoxia

Sucrose synthase (SuSy; EC 2.4.1.13) plays a prominent role in O(2) deficiency and functions at a branch point, partitioning sucrose between cell wall biosynthesis and glycolysis. The cleavage of sucrose by SuSy was localized in wheat ( Triticum aestivum L. cv. Alcedo) roots subjected to 4 days of hypoxia. Increased SuSy activity was observed by in situ activity staining in the tip region and in the stele of root axes. The pattern of cellulose deposition correlated with regions of high SuSy activity. Cellulose accounted for more than 30% of root dry weight and the cellulose content increased substantially under hypoxia. The strongest accumulation of cellulose occurred in the base and mid-regions of the roots where the content rose to 163% and 182% of controls, respectively. In the root axis, cellulose deposition occurred in the endodermis and walls of pith cells. In root tips, cellulose was primarily deposited in developing xylem and phloem. The marker enzyme for O(2) shortage, pyruvate decarboxylase (EC 4.1.1.17), exhibited a 14-fold increase in the root apex, whereas in basal root tissues, which contained more aerenchyma, pyruvate decarboxylase activity was only doubled. The root apex also contained the highest concentration of sucrose and hexoses. The elevated sugar content in all root zones was partially used to synthesize cellulose for secondary wall thickening.     

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.     

Ectopic over-expression of the maize beta-glucosidase Zm-p60.1 perturbs cytokinin homeostasis in transgenic tobacco

The activity of the phytohormone cytokinin depends on a complex interplay of factors such as its metabolism, transport, stability, and cellular/tissue localization. O-glucosides of zeatin-type cytokinins are postulated to be storage and/or transport forms, and are readily deglucosylated. Transgenic tobacco (Nicotiana tabacum L. cv. Petit Havana SR1) plants were constructed over-expressing Zm-p60.1, a maize beta-glucosidase capable of releasing active cytokinins from O- and N3-glucosides, to analyse its potential to perturb zeatin metabolism in planta. Zm-p60.1 in chloroplasts isolated from transgenic leaves has an apparent K(m) more than 10-fold lower than the purified enzyme in vitro. Adult transgenic plants grown in the absence of exogenous zeatin were morphologically indistinguishable from the wild type although differences in phytohormone levels were observed. When grown on medium containing zeatin, inhibition of root elongation was apparent in all seedlings 14 d after sowing (DAS). Between 14 and 21 DAS, the transgenic seedlings accumulated fresh weight leading later (28-32 DAS) to ectopic growths at the base of the hypocotyl. The development of ectopic structures correlated with the presence of the enzyme as demonstrated by histochemical staining. Cytokinin quantification showed that transgenic seedlings grown on medium containing zeatin accumulate active metabolites like zeatin riboside and zeatin riboside phosphate and this might lead to the observed changes. The presence of the enzyme around the base of the hypocotyl and later, in the ectopic structures themselves, suggests that the development of these structures is due to the perturbance in zeatin metabolism caused by the ectopic presence of Zm-p60.1.     

Sucrose Utilization via Invertase and Sucrose Synthase with Respect to Accumulation of Cellulose and Callose Synthesis in Wheat Roots under Oxygen Deficiency

The sucrose cleavage by sucrose synthase (SuSy) and neutral invertase was studied in wheat roots (Triticum aestivum L.) subjected to hypoxia or anoxia for 4 days. By in situ activity staining, increased SuSy activity was observed in the tip region and stele of root axes while the activity of invertase decreased. Cellulose content significantly increased in hypoxically treated roots. The cellulose deposition was correlated with regions of high SuSy activity, being mainly located in the pericycle and endodermis. Invertase activity was distributed along the root without clear difference between cortex and stele. Under root hypoxia, a significant increase in the structural carbohydrates, callose and especially cellulose, was shown. Increasing levels of soluble carbohydrates were partially used to synthesize cellulose for secondary wall thickening and callose to counteract the tissue injury following low-oxygen stress. Under strict anoxia, the roots were much more injured but sustained a high level of cellulose and callose while the soluble carbohydrates almost disappeared.     

The impact of red and blue light-emitting diode illumination on radish physiological indices

The objective was to evaluate the effect of different combinations of red (638 nm) and blue (455 nm) light produced by solid-state light-emitting diodes (LEDs) on physiological indices (net assimilation rate, hypocotyl-to-leaf ratio, leaf area, leaf dry weight, hypocotyl length and diameter, plant length, developed leaves), variation of photosynthetic pigments and non-structural carbohydrates in radish (Raphanus sativus L., var. ‘Faraon’). Lighting experiments were performed under controlled conditions (total PPFD - 200 μmol m⁻² s⁻¹; 16 h photoperiod; 14/18°C night/day temperature). The LED conditions: 638 nm; 638 + 5% 455 nm; 638 + 10% 455 nm; 638 + 10% 455 + 731 nm; 638 + 10% 455 + 731 + 669 nm. Our results showed that radishes grown under red (638 nm) alone were elongated, and the formation of hypocotyl was weak. The net assimilation rate, hypocotyl-to-leaf ratio, and leaf dry weight also were low due to the low accumulation of photosynthetic pigments and non-structural carbohydrates in leaves. The supplemented blue (455 nm) light was necessary for the non-structural carbohydrates distribution between radish storage organs and leaves which resulted in hypocotyl thickening. Red alone (638 nm) or in combination with far-red (731 nm), or red669 for radish generative development was required.     

白及蔗糖合成酶基因的克隆及表达分析

为揭示白及蔗糖合成酶基因与生长发育的关系,该研究以白及为材料,利用RT-PCR技术同源克隆白及蔗糖合成酶的关键基因SuSy,对SuSy基因的生物学特性及表达特征进行了分析,并利用实时荧光定量PCR检测SuSy基因在不同组织中的表达规律。结果表明:(1)白及SuSy基因长度为2 215 bp,编码737个氨基酸,与铁皮石斛、文心兰和蝴蝶兰的蛋白质氨基酸序列的相似性分别为97%、92%和95%。(2)生物信息学分析表明,SuSy蛋白质序列具有较高的亲水性,与拟南芥SuSy蛋白质氨基酸三级结构一致性为75.2%;系统进化树分析发现,白及SuSy蛋白与铁皮石斛处于同一个分支上。(3) qRT-PCR结果表明,SuSy基因在叶片中的表达量最高,块茎中的表达量最低;成熟叶片的表达量高于未成熟叶片的表达量;数据差异性分析显示,SuSy基因在根、块茎中表达量具有极显著性差异,但在一年生叶和二年生叶中的表达量无显著性差异,幼苗叶和一、二年生叶中表达量具有极显著性差异。由此推测,SuSy基因可能受生长发育的诱导,是调控白及生长发育关键基因。     

Sucrose synthase activity does not restrict glycolysis in roots of transgenic potato plants under hypoxic conditions

The effect of hypoxia on root development and carbon metabolism was studied using potato (Solanum tuberosum L.) plants as a model system. Hypoxia led to a cessation of root elongation, and finally to the death of meristematic cells. These changes were accompanied by a 4- to 5-fold accumulation of hexoses, suggesting that insufficient carbohydrate supply was not the cause of cell death. In addition, prolonged hypoxia (96 h) resulted in a 50% increase in activity of most glycolytic enzymes studied and the accumulation of glycerate-3-phosphate and phosphoenolpyruvate. This indicates that endproduct utilisation may restrict metabolic flux through glycolysis. As expected, the activities of alcohol dehydrogenase (EC 1.1.1.1) and pyruvate decarboxylase (EC 4.1.1.17) increased during hypoxia. Apart from the enzymes of ethanolic fermentation the activity of sucrose synthase (SuSy; EC 2.4.1.13) was enhanced. To investigate the in-vivo significance of this increase, transgenic plants with reduced SuSy activity were analysed. Compared to untransformed controls, transgenic plants showed a reduced ability to resume growth after re-aeration, emphasising the crucial role of SuSy in the toleration of hypoxia. Surprisingly, analysis of glycolytic intermediates in root extracts from SuSy antisense plants revealed no change as compared to wildtype plants. Therefore, limitation of glycolysis is most likely not responsible for the observed decreased ability for recovery after prolonged oxygen starvation. We assume that the function of SuSy during hypoxia might be to channel excess carbohydrates into cell wall polymers for later consumption rather than fuelling glycolysis. Received: 17 February 1999 / Accepted: 10 June 1999     

Growth patterns and gravitropic curvature of radish hypocotyls

The rapid growth rate of radish cells makes the hypocotyl particularly valuable in research on growth phenomena and gravitropism. An examination of mean data regarding the growth of aetiolated radish seedlings not subjected to gravitropic stimulation showed that there was an increase in the growth rate from day 3 to day 5. When the hypocotyl was placed horizontally all zones showed an increase in growth of the lower surface of the hypocotyl and generally a decrease in the growth rate in the upper surface. The upper apical part of the gravistimulated hypocotyl grew almost the same as controls. In some cases, in both 4- and 5-day-old seedlings, in the upper median part there was a lower growth rate than in controls. The zones of growth during the development of the hypocotyl were different. Analysis of curvature showed that the growth rate of the different zones was a function of their location in the hypocotyl and that the rate of curvature was different in various parts of the hypocotyl.     

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.     

Absolute quantification of Medicago truncatula sucrose synthase isoforms and N-metabolism enzymes in symbiotic root nodules and the detection of novel nodule phosphoproteins by mass spectrometry

Mass spectrometry (MS) has become increasingly important for tissue specific protein quantification at the isoform level, as well as for the analysis of protein post-translational regulation mechanisms and turnover rates. Thanks to the development of high accuracy mass spectrometers, peptide sequencing without prior knowledge of the amino acid sequence--de novo sequencing--can be performed. In this work, absolute quantification of a set of key enzymes involved in carbon and nitrogen metabolism in Medicago truncatula 'Jemalong A17' root nodules is presented. Among them, sucrose synthase (SuSy; EC 2.4.1.13), one of the central enzymes in sucrose cleavage in root nodules, has been further characterized and the relative phosphorylation state of the three most abundant isoforms has been quantified. De novo sequencing provided sequence information of a so far unidentified peptide, most probably belonging to SuSy2, the second most abundant isoform in M. truncatula root nodules. TiO(2)-phosphopeptide enrichment led to the identification of not only a phosphorylation site at Ser11 in SuSy1, but also of several novel phosphorylation sites present in other root nodule proteins such as alkaline invertase (AI; EC 3.2.1.26) and an RNA-binding protein.     

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.     

Involvement of nitrogen in storage root growth and related gene expression in sweet potato (Ipomoea batatas)

• Nitrogen (N) could affect storage root growth and development of sweet potato. To manage external N concentration fluctuations, plants have developed a wide range of strategies, such as growth changes and gene expression.
• Five sweet potato cultivars were used to analyse the functions of N in regulating storage root growth. Growth responses and physiological indicators were measured to determine the physiological changes regulated by different N concentrations. Expression profiles of related genes were analysed via microarray hybridization data and qRT‐PCR analysis to reveal the molecular mechanisms of storage root growth regulated by different N concentrations.
• The growth responses and physiological indicators of the five cultivars were changed by N concentration. The root fresh weight of two of the sweet potato cultivars, SS19 and GS87, was higher under low N concentrations compared with the other cultivars. SS19 and GS87 were found to be having greater tolerance to low N concentration. The expression of N metabolism and storage root growth related genes was regulated by N concentration in sweet potato.
• These results reveal that N significantly regulated storage root growth. SS19 and GS87 were more tolerant to low N concentration and produced greater storage root yield (at 30 days). Furthermore, several N response genes were involved in both N metabolism and storage root growth.

     

Laboratory methods for studying non-preference resistance to cabbage root fly in cruciferous crops

A laboratory method for testing cruciferous plants for their non-preference resistance to cabbage root fly is described. Test plants were fully randomized on a turntable inside a large chamber containing cabbage root flies. The apparatus was housed in a controlled environment room. The root-fly eggs, laid in the sand surrounding test plants, were extracted using a flask flotation method which was quicker and more efficient than the stirring technique which it superseded. Two batches of plants were tested alternately, one being exposed to the flies, while the other was sampled for eggs. Each batch of test plants received three 1-day exposures to egg-laying, their positions within the test chamber being changed for each exposure to ensure complete randomization. This test method gave highly consistent results and indicated that there were differences in the flies' preference both between cultivars of radish and cauliflower and for individual plants within cultivars. Plants representing the extremes of preferences discovered in the two crops were saved for further study. It was not possible to correlate seed weight, time of seedling emergence, foliage surface area, or hypocotyl attitude in relation to soil level, with the egg-laying preference of the cabbage root flies.     

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.