Suppression of Calcium Transport to Shoots by Root Restriction in Tomato Plants

Restriction of tomato roots by growth in small containers stronglysuppressed transport of ⁴⁵Ca²⁺ ions to new leaves and apices.Water transport, expressed on a leaf area basis, was marginallyreduced by root restriction, an indication that calcium transportwas more severely limited than water transport. (Received October 11, 1996; Accepted January 27, 1997)     

Influence of Brassinosteroids on Antioxidant Enzymes Activity in Tomato Under Different Temperatures

The effect of brassinosteroids (BRs) on catalase (EC 1.11.1.6), peroxidase (EC 1.11.1.7) and superoxide dismutase (SOD, EC 1.15.1.1) activity in tomato leaf discs was analyzed at 25 and 40 °C. Tomato leaf discs were preincubated for 24 h in Petri dishes with 24-epibrassinolide (EBR) or a polyhydroxylated spirostanic analogue of brassinosteroids (MH5). Both concentrations (10.60 and 2.12 nM) of EBR and MH5 stimulated the activity of SOD at 25 and 40 °C, the MH5-stimulated increase of this enzyme activity was greater. Peroxidase activity was unaffected at 25 °C, while at 40 °C this activity was enhanced by both compounds. The changes in catalase activity markedly depended on the structure BRs, doses and temperature. The results suggest a possible role of EBR and MH5 in the reduction of cell damage produced by heat stress due to induction of enzymatic antioxidants.     

Sulphatation Does Not Appear to Be a Protective Mechanism to Prevent Oxysterol Accumulation in Humans and Mice

24S- and 27-hydroxycholesterol (24OHC and 27OHC) are potent regulators of different biochemical systems in vitro and are the major circulating oxysterols. A small fraction of these oxysterols has been reported to be sulphated but there are no detailed studies. We considered the possibility that sulphatation is a protective mechanism preventing accumulation of free oxysterols. Using an accurate assay we found the sulphated fraction of 24OHC and 27OHC in circulation of adults to be less than 15% of total. In two patients with a mutation in CYP7B1 and markedly increased levels of 27OHC the sulphated fraction was 8% and 10% respectively. Infants with severe neonatal cholestasis had however markedly increased sulphate fraction of the above oxysterols. In untreated mice the degree of sulphatation of 24OHC and 27OHC in serum varied between 0 and 16%. Similar degree of sulphatation was found in two mouse models with markedly increased levels of 27OHC and 24OHC respectively. Bile duct ligated mice had higher levels of oxysterols than sham-operated controls but the sulphate fraction was not increased. We conclude that a primary increase in the levels of the oxysterols due to increased synthesis or reduced metabolism in adults and mice does not induce increased sulphatation.     

Brassinosteroids Positively Modulate Growth: Physiological,Biochemical and Anatomical Evidence Using Two Tomato Genotypes Contrasting to Dwarfism

Growth and development are vital processes in the life cycles of plants. Brassinosteroids (BRs) are steroids that when exogenously applied can regulate several biological responses. The aim of this research was to investigate the possible interferences caused by the exogenous application of BR on growth and metabolism using two genotypes of the DWARF gene, MT-d and MT-D, that are BR-deficient and BR-efficient, respectively. The experiment had four treatments with two genotypes (BR-efficient and BR-deficient) and two levels of brassinosteroids (0 and 100 nM BR, described as ??BR and +?BR, respectively). This study revealed that the exogenous application of BR promoted improvement in growth, inducing increases in all variables of both genotypes evaluated. In general, BR-deficient plants sprayed with BR had effects more intense, confirming the benefits of this steroid on photosynthetic apparatus and gas exchange. The changes in the anatomical characteristics of the leaf are related to the contribution of BR on the influx and consequent fixation of CO₂. In addition, modifications related to root anatomy occurred as a result of the BR action with the purpose of increasing the root protection and absorption of water and nutrients. Increases in photosynthetic pigments suggest that the role of BR is linked with chlorophyll biosynthesis and the maintenance of chloroplast integrity, resulting from associations with the increases found in the activities of antioxidant enzymes that modulate the accumulation of reactive oxygen species.     

Phytohormones Accumulation and Distribution in Shoots and Roots of Haploid,Diploid and Tetraploid Barley Seedlings Derived from Microspore Culture

Phytohormones play important roles in plant growth and development, and polyploids are thought to be an important method for plant breeding. However, the relationship between ploidy and phytohormone is still unclear. In this study, barley at three ploidy levels were produced by microspore culture. Therefore, we further analyzed the phytohormone content in the shoots and roots of the three kinds of barley materials to study the effect of ploidy on phytohormones accumulation and distribution. The results showed that Abscisic acid (ABA), gibberellin (GA), jasmonic acid (JA), auxin (IAA), salicylic acid (SA) and cytokinin (CTK) were successfully determined in shoots and roots using LC-MS (liquid chromatography mass spectrometry). By comparing the shoots of the haploid and diploid plants, it was found that the distribution trend of the six phytohormones was consistent, and another consistent trend was found in the roots of the diploid and tetraploid plants. In addition, we further analyzed the shoot/root ratio of the different phytohormones to identify the potential differences for haploid, diploid and tetraploid. Here, the relationship between ploidy and phytohormone we provided would provide new insights into understanding the new phenotypes that occur in polyploid species.     

乳抗氧化肽的分离纯化与结构鉴定

本文检测了不同分子量范围的WPI(乳清分离蛋白)酶解物的抗氧化活性,结果表明各个组分显示出不同的抗氧化活性,其中分子量小于5 kDa的组分最强。采用凝胶过滤色谱对分子量小于5 kDa的WPI酶解物进行分离,抗氧化活性强的组分继续采用RP-HPLC进行纯化。通过MALDI-TOF-MS与氨基酸组成分析鉴定该活性肽为His-Ile-Arg。     

Contrasting Incorporation of 2H from 2H2O into ABA, Xanthoxin and Carotenoids in Tomato Shoots

Nonhebel, H. M. and Milborrow, B. V. 1987. Contrasting incorporationof ²H from ²H²O into ABA, xanthoxin and carotenoids in tomatoshoots.—J. exp. Bot. 38: 980–991. The incorporation of ²H into abscisie acid, xanthoxin, ß-carotene,lutein, lutein epoxide and violaxanthin in tomato shoots incubatedfor 6 d in 70% ²H₂O was compared to investigate whether thesecompounds are precursors of abscisie acid. On average, 5% ofabscisie acid molecules became labelled with a single ²H atomand 21% with from 3 to 14 atoms of ²H. However, mass spectralanalysis of endogenous xanthoxin extracted from the same plants,in darkness, under nitrogen, and derivatized to the pentafluorobenzyloximeshowed incorporation of only single ²H atoms, ruling out xanthoxinas an abscisie acid precursor. Normal-phase HPLC analysis oftomato shoot extracts showed four major carotenoid peaks; ß-carotene,lutein, lutein epoxide and violaxanthin. Calculations basedon the measured carotenoid pool sizes and on the calculatedminimum pool size of the ABA precursor predicted that at least6·8% of violaxanthin molecules or 7·9% of luteinepoxide molecules should become labelled with from 3 to 14 ²Hatoms if these molecules are precursors of abscisie acid. However,mass spectral analysis of xanthoxin derived from purified violaxanthinand lutein epoxide showed no molecules with more than a single²H atom, with detection limits of less than 1% and 0·2%respectively. Similarly, mass spectra of ß-caroteneand lutein did not show any ²H. We conclude that these carotenoidsare not precursors of abscisie acid. Key words: Abscisic acid, xanthoxin, carotenoids     

Biostimulants Derived from Moroccan Seaweeds: Seed Germination Metabolomics and Growth Promotion of Tomato Plant

Polysaccharides extracted from seaweeds can function as plant biostimulants. The aim of this study was to assess the effects of Polysaccharide Enriched Extracts (PEEs) obtained from 17 Moroccan seaweeds, on tomato seed germination and plant growth. Three concentrations (0.02, 0.05 and 0.1 mg mL⁻¹) of PEEs were applied to tomato seeds to evaluate their effect on 3 germination parameters: germination percentage (GP), germination speed (GS) and mean germination time (MGT). Metabolomic analysis by GC–MS was subsequently performed on seedlings. In the second experiment, four PEEs concentrations (0.02, 0.05, 0.1 and 0.2 mg mL⁻¹) were used as foliar spray or as soil application to tomato plants. Their growth parameters (number of leaves, shoot length, fresh and dry weight of stem and roots) and biochemical parameters (chlorophyll a and b) were measured. Results indicated a significant increase of GP and GS associated with a significant reduction of MGT of tomato seeds treated with 0.02 mg mL⁻¹ of PEEs obtained from Gigartina sp., Gigartina pistillata, Chondracanthus acicularis, Gelidium crinale, Schizymenia dubyi, Cystoseira. foeniculacea and Fucus spiralis. Similar results were also obtained by application of higher PEEs concentration (0.1 mg mL⁻¹) extracted from Ulva rigida, Codium tomentosum, Codium decorticatum and Bifurcaria bifurcata. Metabolomic analysis on seedlings detected the presence of some metabolites which could possibly be involved in seed germination enhancement or inhibition. The results of the second experiment showed that the same PEEs cited above at the same concentrations enhanced plant dry weight and chlorophyll a content except Gigartina sp., C. foeniculacea and C. decorticatum. Furthermore, soil application of PEEs was more effective in improving plant growth parameters than foliar application. The study shows the potential of PEEs from Moroccan seaweed to be used as biostimulants for a sustainable agriculture.

     

利用多重PCR反应同时筛选番茄Cf-9和Tm-1基因

利用同一PCR反应体系,对分别与番茄抗叶霉病的Cf-9基因和抗番茄烟草花叶病毒病的Tm-1基因紧密连锁的PCR标记进行了同时扩增筛选,扩增的特异性片段与单引物扩增片段吻合。其中与Cf-9基因紧密连锁的CAPs标记在抗感试材均可扩增出560bp的特异片段,且都存在TaqⅠ酶切位点,抗病基因型酶切后分别产生了450bp、330bp和290bp的不同特异性片段,而感病基因型试材酶切后产生450bp和290bp的特异性片段;与Tm-1基因紧密连锁的SCAR标记为显性标记,只有抗病试材产生750bp的特异片段,不能被TaqⅠ酶切。经反复验证,结果稳定准确,可用于在同一PCR反应体系中对两个抗病基因进行同时筛选鉴定。该体系的建立不仅省时、省工、节省费用,而且可用于苗期辅助选育,加快番茄抗病育种进程。     

Signaling Activities of the Drosophila Wingless Gene Are Separately Mutable and Appear to Be Transduced at the Cell Surface

The Drosophila segment polarity gene wingless encodes an intercellular signaling molecule that transmits positional information during development of the embryonic epidermis. We have explored the mechanism of wg signal transduction by perturbing cellular processes genetically and by performing structure/function analysis of the Wg protein. We present evidence that Wingless protein may transduce signal at the cell surface and that Wg may bind to its cell surface receptor without necessarily activating it. We demonstrate that two specific signaling activities of the Wg molecule can be disrupted independently by mutation. Sequence analysis indicates that these different signaling activities are not promoted by discrete functional domains, but rather that the overall conformation of the molecule may control distinct signaling functions. We conclude that wg signaling may involve complex interactions between the Wg ligand and its cell surface receptor molecule(s) and that some of this complexity resides within the Wg ligand itself.     

Culture-Independent Identification of Microorganisms That Respond to Specified Stimuli

A new approach that permits culture-independent identification of microorganisms that respond to specified stimuli was developed. This approach was illustrated by examination of microorganisms that grew in response to various nutrient supplements added to soil. A thymidine nucleotide analog, bromodeoxyuridine (BrdU), and supplements were added to soil and incubated for 3 days. DNA was extracted from the soil, and the newly synthesized DNA was isolated by immunocapture of the BrdU-labeled DNA. The unique perspective this approach offers was demonstrated by comparing the microbial community structures obtained from total soil DNA and the BrdU-labeled fraction in an rRNA gene (rDNA) analysis. The traditional total DNA analysis revealed no notable differences between the treatments, whereas the BrdU-labeled DNA showed significantly different banding patterns between the nutrient supplement treatments and compared with total DNA banding patterns. PCR primers were developed to specifically amplify the intergenic region of an rDNA sequence unique to the BrdU analysis of a phosphate supplement treatment. Amplification of DNA from all treatments using these primers showed that it was unique to the phosphate treatment and that it was present in both the total DNA and BrdU-labeled DNA fractions. This result demonstrates the promise of this new strategy, because it was able to permit identification of a sequence from a phosphate-responsive organism that was not discernable in the traditional total DNA community structure analysis.     

Isolation of Temperature-Sensitive Mutants of Arabidopsis thaliana That Are Defective in the Redifferentiation of Shoots

Three temperature-sensitive mutants of Arabidopsis thaliana that were defective in the redifferentiation of shoots were isolated as tools for the study of organogenesis. M3 lines were constructed by harvesting M3 seeds separately from each M2 plant. Comparative examination of shoot redifferentiation in root explants of 2700 M3 lines at 22[deg]C (permissive temperature) and at 27[deg]C (restrictive temperature) led to the identification of seven temperature-sensitive mutant lines. Genetic tests of three of the seven mutant lines indicated that temperature-sensitive redifferentiation of shoots in these three lines resulted from single, nuclear, recessive mutations in three different genes, designated SRD1, SRD2, and SRD3. The morphology of root explants of srd mutants cultured at the restrictive temperature suggests that the products of these SRD genes function at different stages of the redifferentiation of shoots.     

抗CD47胞外区骆驼多克隆抗体的制备及鉴定

验证人CD47蛋白能否在新疆双峰驼中产生高滴度抗体;为制备高亲和力的抗CD47纳米抗体提供实验依据。构建CD47胞外区原核表达载体,诱导表达及纯化CD47蛋白,免疫新疆双峰驼。通过酶联免疫吸附试验(ELISA)和Western blot分别检测骆驼多克隆抗体的效价及与CD47蛋白特异性结合活性。结果显示,成功构建CD47胞外区原核表达载体,获得纯度大于90%的CD47蛋白,表达纯化的CD47重组蛋白可与抗CD47单抗B6H12.2特异结合;ELISA测定CD47重组蛋白免疫骆驼7次后,抗CD47多克隆抗血清的效价至少达到1∶200 000,免疫印迹检测表明抗CD47骆驼抗血清与CD47重组蛋白、Jurkat和Raji细胞表面表达的CD47均能特异结合。重组人CD47蛋白可以在骆驼体内激发高滴度抗体反应。     

Conversion of Exogenous Cholesterol into Glycoalkaloids in Potato Shoots,Using Two Methods for Sterol Solubilisation

Steroidal glycoalkaloids (SGA) are toxic secondary metabolites naturally occurring in the potato, as well as in certain other Solanaceous plant species, such as tomato, eggplant and pepper. To investigate the steroidal origin of SGA biosynthesis, cut potato shoots were fed cholesterol labelled with deuterium (D) in the sterol ring structure (D₅- or D₆-labelled), or side chain (D₇-labelled), and analysed after three or five weeks. The labelled cholesterol and presence of D-labelled SGA were analysed by GC-MS and LC-MS/MS, respectively. When feeding D-labelled cholesterol solubilised in Tween-80, labelled cholesterol in free form became present in both leaves and stems, although the major part was recovered as steryl esters. Minor amounts of D-labelled SGA (α-solanine and α-chaconine) were identified in cholesterol-treated shoots, but not in blank controls, or in shoots fed D₆-27-hydroxycholesterol. Solubilising the labelled cholesterol in methyl-β-cyclodextrin instead of Tween-80 increased the levels of labelled SGA up to 100-fold, and about 1 mole% of the labelled cholesterol was recovered as labelled SGA in potato leaves. Both side chain and ring structure D labels were retained in SGA, showing that the entire cholesterol molecule is converted to SGA. However, feeding side chain D₇-labelled cholesterol resulted in D₅-labelled SGA, indicating that two hydrogen atoms were released during formation of the SGA nitrogen-containing ring system. Feeding with D₇-sitosterol did not produce any labelled SGA, indicating that cholesterol is a specific SGA precursor. In conclusion, we have demonstrated a superior performance of methyl-β-cyclodextrin for delivery of cholesterol in plant tissue feeding experiments, and given firm evidence for cholesterol as a specific sterol precursor of SGA in potato.     

Studies on two Gibberellin-like Substances in Young Shoots of Tomato (Lycopersicon esculentum Mill.)

Two gibberellin-like substances were found in the acidic fractionof shoot extracts of the tomato (Lycopersicon esculentum Mill.,cultivar Potentate). These were resolved by paper chromotographywith iso-propanol/ammonia/water (10:1:1) as the developing solventbut not with n-butanol/1.5 N ammonia (3:1). Both substanceswere active in the dwarf maize bioassay on mutants d-1, d-2,d-3, and d-5, and appeared to be more active on d-5 than d-1.Neither was active in the Meteor Pea assay. Neutral and basicfractions were inactive. The relative amounts of these two substances varied accordingto the age of the tissues from which they were extracted andthis feature is discussed in relation to future studies on thephysiology of gibberellin-like substances in vivo.     

Genome-Wide Identification and Expression Analysis of Aquaporins in Tomato

The family of aquaporins, also called water channels or major intrinsic proteins, is characterized by six transmembrane domains that together facilitate the transport of water and a variety of low molecular weight solutes. They are found in all domains of life, but show their highest diversity in plants. Numerous studies identified aquaporins as important targets for improving plant performance under drought stress. The phylogeny of aquaporins is well established based on model species like Arabidopsis thaliana, which can be used as a template to investigate aquaporins in other species. In this study we comprehensively identified aquaporin encoding genes in tomato (Solanum lycopersicum), which is an important vegetable crop and also serves as a model for fleshy fruit development. We found 47 aquaporin genes in the tomato genome and analyzed their structural features. Based on a phylogenetic analysis of the deduced amino acid sequences the aquaporin genes were assigned to five subfamilies (PIPs, TIPs, NIPs, SIPs and XIPs) and their substrate specificity was assessed on the basis of key amino acid residues. As ESTs were available for 32 genes, expression of these genes was analyzed in 13 different tissues and developmental stages of tomato. We detected tissue-specific and development-specific expression of tomato aquaporin genes, which is a first step towards revealing the contribution of aquaporins to water and solute transport in leaves and during fruit development.