Protein Phosphorylation in Plastids from Ripening Tomato Fruits {Lycopersicon esculentum)

Incubation of plastids isolated from ripening tomato fruits(Lycopersicon esculentum) with [     

Acetyl-CoA Carboxylase Inhibitors from Avocado (Persea americana Mill) Fruits

A methanol extract of avocado fruits showed potent inhibitory activity against acetyl-CoA carboxylase, a key enzyme in fatty acid biosynthesis. The active principles were isolated and identified as (5E,12Z,15Z)-2-hydroxy-4-oxoheneicosa-5,12,15-trienyl (1), (2R,12Z,15Z)-2-hydroxy-4-oxoheneicosa-12,15-dienyl (2), (2R^*,4R^*)-2,4-dihydroxyheptadec-16-enyl (3) and (2R^*,4R^*)-2,4-dihydroxyheptadec-16-ynyl (4) acetates by instrumental analyses. The IC₅₀ of the compounds were 4.0×10^-6, 4.9×10^-6, 9.4×10^-6, and 5.1×10^-6M, respectively.     

Cell Wall Metabolism in Ripening Fruit (VII. Biologically Active Pectin Oligomers in Ripening Tomato (Lycopersicon esculentum Mill.) Fruits)

A water-soluble, ethanol-insoluble extract of autolytically inactive tomato (Lycopersicon esculentum Mill.) pericarp tissue contains a series of galacturonic acid-containing (pectic) oligosaccharides that will elicit a transient increase in ethylene biosynthesis when applied to pericarp discs cut from mature green fruit. The concentration of these oligosaccharides in extracts (2.2 [mu]g/g fresh weight) is in excess of that required to promote ethylene synthesis. Oligomers in extracts of ripening fruits were partially purified by preparative high-performance liquid chromatography, and their compositions are described. Pectins were extracted from cell walls prepared from mature green fruit using chelator and Na2CO3 solutions. These pectins are not active in eliciting ethylene synthesis. However, treatment of the Na2CO3-soluble, but not the chelator-soluble, pectin with pure tomato polygalacturonase 1 generates oligomers that are similar to those extracted from ripening fruit (according to high-performance liquid chromatography analysis) and are active as elicitors. The possibility that pectin-derived oligomers are endogenous regulators of ripening is discussed.     

Reduction in Pectin Methylesterase Activity Modifies Tissue Integrity and Cation Levels in Ripening Tomato (Lycopersicon esculentum Mill.) Fruits

Pectin methylesterase (PME, EC 3.1.1.11) is an ubiquitous enzyme in the plant kingdom; however, its role in plant growth and development is not yet understood. Using transgenic tomato (Lycopersicon esculentum Mill.) fruits that show more than 10-fold reduction in PME activity because of expression of an antisense PME gene, we have investigated the role of PME in tomato fruit ripening. Our results show that reduced PME activity causes an almost complete loss of tissue integrity during fruit senescence but shows little effect on fruit firmness during ripening. Low PME activity in the transgenic fruit pericarp modified both accumulation and partitioning of cations between soluble and bound forms and selectively impaired accumulation of Mg2+ over other major cations. Decreased PME activity was associated with a 30 to 70% decrease in bound Ca2+ and Mg2+ in transgenic pericarp. Levels of soluble Ca2+ increase 10 to 60%, whereas levels of soluble Mg2+ and Na+ are reduced by 20 to 60% in transgenic pericarp. Changes in cation levels associated with lowered PME activity do not affect the rate of respiration or membrane integrity of fruit during ripening. Overall, these results suggest that PME plays a role in determining tissue integrity during fruit senescence, perhaps by regulating cation binding to the cell wall.     

Mannosyl- and Xylosyl-Containing Glycans Promote Tomato (Lycopersicon esculentum Mill.) Fruit Ripening

The oligosaccharide glycans mannosylα1-6(mannosylα1-3)mannosylα1-6(mannosylα1-3) mannosylβ1-4-N-acetylglucosamine and mannosylα1-6(mannosylα1-3)(xylosylβ1-2) mannosylβ1-4-N-acetylglucosaminyl(fucosylα1-3) N-acetylglucosamine were infiltrated into mature green tomato fruit (Lycopersicon esculentum Mill., cv Rutgers). Coinfiltration of 1 nanogram per gram fresh weight of the glycans with 40 micrograms per gram fresh weight galactose, a level of galactose insufficient to promote ripening, stimulated ripening as measured by red coloration and ethylene production.     

Interactions of Avocado (Persea americana) Cytochrome P-450 with Monoterpenoids

The microsomal fraction of avocado (Persea americana) mesocarp is a rich source of cytochrome P-450 active in the demethylation of xenobiotics. Cytochrome P-450 from this tissue has been purified and well characterized at the molecular level (DP O'Keefe, KJ Leto [1989] Plant Physiol 89: 1141-1149; KR Bozak, H Yu, R Sirevag, RE Christoffersen [1990] Proc Natl Acad Sci USA 87: 3904-3908). Despite this extensive characterization, the role of the enzyme in vivo was not established. Optical and electron paramagnetic resonance binding studies described here suggest that the monoterpenoids, nerol and geraniol, are substrates of avocado cytochrome P-450 (spectral dissociation constant of 7.2 and 35 micromolar, respectively). Avocado microsomes have been shown to catalyze the hydroxylation of these monoterpenoids, and both nerol and geraniol have been shown to inhibit the activity of avocado cytochrome P-450 toward the artificial substrate 7-ethoxycoumarin, with nerol a competitive inhibitor of this activity.     

Cytochrome P-450 from the Mesocarp of Avocado (Persea americana)

The microsomal fraction from the mesocarp of avocado (Persea americana) is one of few identified rich sources of plant cytochrome P-450. Cytochrome P-450 from this tissue has been solubilized and purified. Enzymatic assays (p-chloro-N-methylaniline demethylase) and spectroscopic observations of substrate binding suggest a low spin form of the cytochrome, resembling that in the microsomal membrane, can be recovered. However, this preparation of native protein is a mixture of nearly equal proportions of two cytochrome P-450 polypeptides that have been resolved only under denaturing conditions. Overall similarities between these polypeptides include indistinguishable amino acid compositions, similar trypsin digest patterns, and cross reactivity with the same antibody. The amino terminal sequences of both polypeptides are identical, with the exception that one of them lacks a methionine residue at the amino terminus. This sequence exhibits some similarities with the membrane targeting signal found at the amino terminus of most mammalian cytochromes P-450.     

Ripening Physiology of Fruit from Transgenic Tomato (Lycopersicon esculentum) Plants with Reduced Ethylene Synthesis

The physiological effects of reduced ethylene synthesis in a transgenic tomato (Lycopersicon esculentum) line expressing 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase enzyme have been examined. Fruit from the transgenic line 5673 ripen significantly slower than control fruit when removed from the vine early in ripening. In contrast, fruit that remain attached to the plants ripen much more rapidly, exhibiting little delay relative to the control. Ethylene determinations on attached fruit revealed that there was significantly more internal ethylene in attached than detached fruit. The higher ethylene content can fully account for the observed faster on-the-vine ripening. All of the data are consistent with a catalytic role for ethylene in promoting many, although not all, aspects of fruit ripening. Biochemical analyses of transgenic fruit indicated no significant differences from controls in the levels of ACC oxidase or polygalacturonase. Because transgenic fruit are significantly firmer than controls, this last result indicates that other enzymes may have a significant role in fruit softening.     

Descriptive Physiology and Biochemistry of the Abnormally Ripening Tomato Fruit (Lycopersicon esculentum) cv. Snowball

Aspergillus tamarii Kita grew and sporulated best at 30°C. The best pH for growth and sporulation were 5.5 and 6.5, respectively. Among the carbon sources employed, glucose supported the highest growth and sporulation. Best growth was obtained with sodium nitrate as nitrogen source and best sporulation with ammonium tar-tarate. When glucose was used as carbon source, the carbohydrates found in the mycelium included myoinositol. dulcitol, fructose, arabinose and ribose.     

Expression of a Ripening-Related Avocado (Persea americana) Cytochrome P450 in Yeast

One of the mRNAs that accumulates during the ripening of avocado (Persea americana Mill. cv Hass) has been previously identified as a cytochrome P450 (P450) monooxygenase and the corresponding gene designated CYP71A1. In this report we demonstrate that during ripening the accumulation of antigenically detected CYP71A1 gene product (CYP71A1) correlates with increases in total P450 and two P450-dependent enzyme activities: para-chloro-N-methylaniline demethylase, and trans-cinnamic acid hydroxylase (tCAH). To determine whether both of these activities are derived from CYP71A1, we have expressed this protein in yeast (Saccharomyces cerevisiae) using a galactose-inducible yeast promoter. Following induction, the microsomal fraction of transformed yeast cells undergoes a large increase in P450 level, attributable almost exclusively to the plant CYP71A1 protein. These membranes exhibit NADPH-dependent para-chloro-N-methylaniline demethylase activity at a rate comparable to that in avocado microsomes but have no detectable tCAH. These results demonstrate both that the CYP71A1 protein is not a tCAH and that a plant P450 is fully functional upon heterologous expression in yeast. These findings also indicate that the heterologous P450 protein can interact with the yeast NADPH:P450 reductase to produce a functional complex.     

Compartments and Permeability for Potassium in Developing Fruits of Tomato (Lycopersicon esculentum Mill.)

Permeabilities of plasmalemma and tonoplast, and the distributionof potassium between free space, cytoplasm, and vacuole, wereestimated from the kinetics of the efflux of potassium fromslices of tomato pericarp. The permeabilities of plasmalemmaand tonoplast did not change during development, but the proportionof potassium in the cytoplasm increased during ripening. Itis suggested that the effects of ethylene, and the changes takingplace during ripening are not caused by changing permeabilitiesof membranes. Increasing activities of ions in the cytoplasm,rather than increasing membrane permeabilities, may explainprevious observations that the efflux of solutes from fruittissues increases during ripening.     

Tissue Culture Studies of Tomato (Lycopersicon esculentum)

Tomato is a major vegetable crop that has achieved tremendous popularity over the last century. It is grown in almost every country of the world. Development of protocols for in vitro selection can provide new advances for the production of stress tolerant cultivars. Techniques have been optimised for the production of haploids and somatic hybrids. Attempts have also been made to transfer the higher regenerative ability of wild varieties to cultivated tomatoes. Although, some information is available on the morphogenesis of tomato, the techniques have not been developed to a level at which they can be utilised in large-scale multiplication of commercially important cultivars. The morphogenesis response seems to be highly dependent PGRs used in the media, which is again cultivar and genotypic specific. Somatic embryogenesis in tomato is still at its infancy, and efficient procedures for large-scale production via somatic embryogenesis are yet to be developed. Genetic stability of the tissue culture raised tomato plants also needs to be addressed. The use of a combination of molecular and conventional breeding techniques could be the option for the development of cultivars resistant to biotic and abiotic stresses. This paper reviews the advances made in various aspects of tissue culture in tomato. It also discusses the issues that still need to be addressed to utilise the full potential of plant tissue culture techniques in genetic improvement and mass propagation of tomato.     

On the Function of Seedcoats in Gibberellin-induced Hydrolysis of Starch Reserves during Avocado (Persea americana) Germination

Of all seed components investigated, the endogenous levels ofboth bound and free gibberellin (GA) are highest in embryo tissueof young immature avocado seeds. Experimentation with the gelatinaceousendosperm tissue of these seeds indicated that hydrolysis ofstarch reserves may be triggered by the embryo-contained GAacting on the ‘endosperm sheath’, a thin layer ofcells encompassing the endosperm. In the mature fruit when theendosperm and the encompassing layer are no longer present andin which starch reserves are present in the two stony cotyledons,the papery bi-integumental layer, commonly regarded as biologicallyinert, retains the ability to trigger starch hydrolysis uponcontact with GA. From these data and also based upon the findingsthat during germination levels of endogenous GA increase inembryo tissue and that exogenously applied GA₃ is effectivein induction of dormancy break it is proposed that in the avocadoseed an analogy, physiological but not anatomical, to the cerealendosperm system exists.     

Relationship between Fruit Softening and Wall Polysaccharides in Avocado (Persea americana Mill) Mesocarp Tissues

Changes in avocado (Persea americana) fruit texture during ripeningwere evaluated by stress-relaxation analysis. A conical probewas imposed into the mesocarp tissue to a depth of 0.6 mm andthe initial stress and the stress relaxation over 60 s weredetermined. The initial stress, an elastic parameter, was substantiallyreduced within one day when ripening was initiated by transferringthe fruit from 15 to 25°C. The minimum and maximum relaxationtime, parameters which reflect viscosity, were also reducedwithin one day. Mesocarp cell walls were fractionated into water-soluble(WS), hot EDTA-soluble (EDTA), alkaline soluble (hemicellulose)and the residual (cellulose) fractions. The amount of cellulosedid not change during ripening. Rhamnose, arabinose and uronicacids in the WS fraction increased during the initial day ofripening; those same components decreased in the EDTA fraction.A molecular weight downshift in the WS acidic polysaccharideswas detected within one day, while only slight changes wereobserved in the molecular weights of the EDTA fraction. Thequantities of individual sugar components of major hemicellulosefraction were unchanged, but there was a prominent molecularweight downshift in the xyloglucan components within one day.These results clearly revealed that both elastic and viscousproperties of avocado mesocarp tissues were substantially alteredduring ripening, and that the solubility changes in acidic polysaccharidesand decreases in the average molecular weight of cell wall xyloglucancomponents were associated with significant changes in fruittexture. (Received December 13, 1996; Accepted March 5, 1997)     

1-甲基环丙烯采后处理对樱桃番茄果实成熟过程的影响

研究了不同浓度(0、0.035、0.07和0.11μL/L)的乙烯受体竞争性抑制剂1-甲基环丙烯(1-MCP)采后处理对绿熟期樱桃番茄的乙烯合成、果实软化、果实色素(叶绿素、茄红素、β-胡萝卜素)含量消长的影响.0.07 μL/L及其以上浓度的1-MCP降低了前期乙烯合成,同时推迟了乙烯释放高峰,但0.035 μL/L浓度的1-MCP处理并不能抑制内源乙烯合成.1-MCP显著延迟了果实软化和叶绿素降解,但并不影响这两个过程的启动.茄红素合成的启动和积累均受到了1-MCP抑制,而1-MCP并不推迟β-胡萝卜素合成的启动,只抑制其积累.这些结果提示了乙烯调节成熟生理过程的不同机制.对于绿熟期的樱桃番茄,0.07～0.11μL/L的1-MCP是实用的有效处理浓度.1-MCP有效浓度可能用于了解果实的乙烯受体水平和乙烯敏感性.     

Pistil Starch Reserves at Anthesis Correlate with Final Flower Fate in Avocado (Persea americana)

A common observation in different plant species is a massive abscission of flowers and fruitlets even after adequate pollination, but little is known as to the reason for this drop. Previous research has shown the importance of nutritive reserves accumulated in the flower on fertilization success and initial fruit development but direct evidence has been elusive. Avocado (Persea americana) is an extreme case of a species with a very low fruit to flower ratio. In this work, the implications of starch content in the avocado flower on the subsequent fruit set are explored. Firstly, starch content in individual ovaries was analysed from two populations of flowers with a different fruit set capacity showing that the flowers from the population that resulted in a higher percentage of fruit set contained significantly more starch. Secondly, in a different set of flowers, the style of each flower was excised one day after pollination, once the pollen tubes had reached the base of the style, and individually fixed for starch content analysis under the microscope once the fate of its corresponding ovary (that remained in the tree) was known. A high variability in starch content in the style was found among flowers, with some flowers having starch content up to 1,000 times higher than others, and the flowers that successfully developed into fruits presented significantly higher starch content in the style at anthesis than those that abscised. The relationship between starch content in the ovary and the capacity of set of the flower together with the correlation found between the starch content in the style and the fate of the ovary support the hypothesis that the carbohydrate reserves accumulated in the flower at anthesis are related to subsequent abscission or retention of the developing fruit.