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Inhibitory action of five tannins on growth induced by several gibberellins.

The following tannins, Chinese gallotannin, 1,2,3,4,6-pentagalloyl glucose, chebulinic acid, procyanidin dimers, and procyanidin trimers were tested and found to be antagonists of seven gibberellins (GAs). Each tannin inhibited the growth induced by any of the gibberellins GA(1), GA(3), GA(4), GA(7), GA(9), GA(13), and GA(14) in the dwarf pea assay. Endogenous growth was not affected. The highest ratio of tannin to gibberellin tested (1000:1 by weight) inhibited from 60 to 95% of the induced growth for all tannins and all gibberellins tested. The tannins were particularly inhibitory against GA(4) and GA(14) where a ratio of 10:1 (tannins: GA by weight) resulted in up to 85% growth reduction. Inhibition could be completely reversed by increasing the amount of gibberellin in all combinations studied. The procyanidin dimers and trimers were the first purified components of condensed tannins to be tested in this system and were potent inhibitors particularly against growth induced by GA(4) and GA(14). Inhibition by these compounds along with similar inhibition by previously tested hydrolyzable tannins demonstrates that the effect is general to tannins of all classes.

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Gibberellins promote flowering of arabidopsis by activating the LEAFY promoter

The gibberellin class of plant hormones has been implicated in the control of flowering in several species. In Arabidopsis, severe reduction of endogenous gibberellins delays flowering in long days and prevents flowering in short days. We have investigated how the differential effects of gibberellins on flowering correlate with expression of LEAFY, a floral meristem identity gene. We have found that the failure of gibberellin-deficient ga1-3 mutants to flower in short days was paralleled by the absence of LEAFY promoter induction. A causal connection between these two events was confirmed by the ability of a constitutively expressed LEAFY transgene to restore flowering to ga1-3 mutants in short days. In contrast to short days, impairment of gibberellin biosynthesis caused merely a reduction of LEAFY expression when plants were grown in long days or with sucrose in the dark. As a first step toward identifying other small molecules that might regulate flowering, we have developed a rapid in vitro assay for LEAFY promoter activity.

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GIBBERELLIN PRODUCTION IN PEA SEEDS DEVELOPING IN EXCISED PODS: EFFECT OF GROWTH RETARDANT AMO-1618.

The gibberellin content of pea seeds developing in excised pea pods cultured on a medium containing the plant growth retardant AMO-1618 was reduced in comparison with pea seeds cultured on retardant-free medium. The reduction increased with increasing concentrations of AMO-1618. However, at the lowest concentration tested (5 milligrams per liter) the growth of the seeds was not affected whereas their gibberellin content was significantly reduced. In conjunction with earlier work on the effect of growth retardants on gibberellin biosynthesis in the fungus Fusarium moniliforme, these results indicate that AMO-1618 inhibits the biosynthesis of gibberellins in tissues of higher plants in much the same manner it does in Fusarium.

Gibberellins↗

Action of carbamate biocides on sterols, gibberellin and aflatoxin formation.

An interrelationship between primary and secondary metabolites of a toxigenic mould (Aspergillus parasiticus var. globosus IMI 120920) was obtained in liquid broth containing various concentrations of dithane Z-78 and propoxur. The results show that a stimulatory response was recorded in gibberellin, protein, sterol and aflatoxin yield in cultures treated with 15, 30, 50 and 100 ppm a.i. of dithane Z-78. On the other hand, propoxur reduced the yield of gibberellin, amino acids, protein, sterol and aflatoxin. In comparison to its effect on the mycelial growth, gibberellin and aflatoxin activity is inhibited more severely than dry weight increase, thus leading to the assumption that there is a specific effect of the inhibitor on both gibberellin and aflatoxin synthesis.

Aflatoxins↗

Computer modeling of gibberellin-DNA binding.

Computer modeling and molecular mechanics performed on the intercalation complexes of selected gibberellins or biosynthetic precursors with DNA dinucleotides revealed that under appropriate conditions the ligands insert (intercalate) between the base-paired double-stranded dinucleotide, 5'-dTdA-3'. Stabilization of the double-stranded dinucleotide after docking of a gibberellin between base pairs is inferred by the sum negative energy of hydrogen bonding and van der Waals contacts and the entropic changes which accompany the formation of each ligand-dinucleotide complex. In addition, the interactions of the gibberellins and dinucleotides, with the gibberellic acid-dinucleotide complex serving as the prototype, show optimum geometry and stereochemical hydrogen bonding recognition which are dependent upon the complementary chirality and stereochemistry of the individual components. Whether or not the gibberellins directly influence the uncoiling of DNA or gene expression at the transcriptional level via an intercalation mechanism is a matter of conjecture, albeit one that warrants intensive investigation.

Computer Simulation↗

Gibberellin metabolism: new insights revealed by the genes.

The identification of most of the genes involved in the metabolic pathways for gibberellin hormones has helped us to understand these pathways and their regulation. Many of these enzymes are multifunctional and therefore fewer enzymes than might be expected are required to synthesize the various gibberellin structures. However, several of the enzymes are encoded by multiple genes that are regulated differently, adding unexpected genetic complexity. Several endogenous and environmental factors modify the expression of gibberellin biosynthesis genes, including developmental stage, hormonal status and light. A future challenge will be to dissect the complex, interacting pathways that mediate the regulation of gibberellin metabolism.

Amino Acid Sequence↗

The CYP88A cytochrome P450, ent-kaurenoic acid oxidase, catalyzes three steps of the gibberellin biosynthesis pathway.

We have shown that ent-kaurenoic acid oxidase, a member of the CYP88A subfamily of cytochrome P450 enzymes, catalyzes the three steps of the gibberellin biosynthetic pathway from ent-kaurenoic acid to GA(12). A gibberellin-responsive barley mutant, grd5, accumulates ent-kaurenoic acid in developing grains. Three independent grd5 mutants contain mutations in a gene encoding a member of the CYP88A subfamily of cytochrome P450 enzymes, defined by the maize Dwarf3 protein. Mutation of the Dwarf3 gene gives rise to a gibberellin-responsive dwarf phenotype, but the lesion in the gibberellin biosynthesis pathway has not been identified. Arabidopsis thaliana has two CYP88A genes, both of which are expressed. Yeast strains expressing cDNAs encoding each of the two Arabidopsis and the barley CYP88A enzymes catalyze the three steps of the GA biosynthesis pathway from ent-kaurenoic acid to GA(12). Sequence comparison suggests that the maize Dwarf3 locus also encodes ent-kaurenoic acid oxidase.

Arabidopsis↗

Separate compartments for the production of sterols, carotenoids and gibberellins in Gibberella fujikuroi.

Substrate flows in the sterol, carotenoid and gibberellin pathways of Gibberella fujikuroi were examined by isotope-dilution experiments. The wild type and two carotenoid mutants of this fungus were grown in minimal medium with abundant glucose, limiting ammonium nitrate and a radioactively labelled precursor (either acetate, mevalonate or leucine). The precursors did not affect growth or terpenoid production, with two exceptions; leucine allowed additional growth, as expected from the nitrogen limitation in the medium, and mevalonate inhibited the accumulation of gibberellins, but only if added before the onset of gibberellin production. The relative contributions of glucose, mevalonate, leucine and acetate as terpenoid precursors, calculated from the specific radioactivities of ergosterol, neurosporaxanthin and phytoene, were different for different products and different precursors. We conclude that the biosyntheses of sterols, gibberellins and carotenoids in Gibberella are physically separated in different subcellular compartments with independent substrate pools. The same results were obtained with the three strains, except for carotenoid production, indicating that this pathway is regulated independently from other terpenoid pathways.

Acetates↗

Identification of a peptide mimic of bioactive gibberellins with affinity to GA 2-oxidase.

Previously we reported the first example of peptide mimics of a small hydrophobic molecule, a phytohormone gibberellin. The second peptide mimic of gibberellin has been identified from random peptide libraries by its affinity to a type of catalyzing enzyme of gibberellins, which specifically recognizes bioactive gibberellins. These results suggest that even hydrophobic compounds can be mimicked by peptides.

Catalysis↗

Aloe vera and gibberellin. Anti-inflammatory activity in diabetes.

Aloe vera inhibits inflammation and adjuvant-induced arthritis. The authors' laboratory has shown that A. vera improves wound healing, which suggests that it does not act like an adrenal steroid. Diabetic animals were used in this study because of their poor wound healing and anti-inflammatory capabilities. The anti-inflammatory activity of A. vera and gibberellin was measured in streptozotocin-induced diabetic mice by measuring the inhibition of polymorphonuclear leukocyte infiltration into a site of gelatin-induced inflammation over a dose range of 2 to 100 mg/kg. Both Aloe and gibberellin similarly inhibited inflammation in a dose-response manner. These data tend to suggest that gibberellin or a gibberellin-like substance is an active anti-inflammatory component in A. vera.

Aloe↗

The Effect of Gibberellins on Flowering in Roses.

The gibberellins A(1), A(3), A(5), A(8), A(19), A(20), and A(29) were identified in vegetative shoot tips of Rosa canina by comparing their mass spectra and Kovats retention indices with those of standards. Most wild roses have a short flowering season of 2-4 weeks in spring, whereas most modern cultivars flower recurrently. 'Félicité et Perpétue' is a short-season hybrid from a cross between a wild rose and a recurrent-flowering rose, whereas its sport, 'Little White Pet,' flowers recurrently. The concentrations of gibberellins (GAs) were measured in shoot apices of both cultivars. In March (before floral initiation in spring) the concentrations of GA(1) and GA(3) were respectively threefold and twofold higher in 'Félicité et Perpétue' than in 'Little White Pet.' In April (after floral initiation) the concentrations of both gibberellins were substantially greater than in March, and concentrations of GA(1) and GA(3) were, respectively, 17-fold and 12-fold greater in 'Félicité et Perpétue' than in 'Little White Pet.' It is postulated that, in 'Félicité et Perpétue,' floral initiation occurs when concentrations of GAs are low and is inhibited when concentrations of GAs are high, whereas in 'Little White Pet' concentrations of GAs remain at permissive levels throughout the growing season. Applications of GA(1) and GA(3) to axillary shoots in March inhibited floral development in 'Félicité et Perpétue' but not in 'Little White Pet.' This suggests that the combined concentration of exogenous and endogenous gibberellins might have been raised to inhibitory levels in the former but not in the latter cultivar.

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The Deoxyxylulose Phosphate Pathway for the Biosynthesis of Plastidic Isoprenoids: Early Days in Our Understanding of the Early Stages of Gibberellin Biosynthesis.

The identification of a novel pathway for isopentenyl diphosphate synthesis by Rohmer, Arigoni and colleagues in the early 1990's has led to a reappraisal of terpenoid biosynthesis in many organisms. It is now apparent that in plants there are two biosynthetic routes to isopentenyl diphosphate-the classical mevalonate pathway in the cytosol and the deoxyxylulose phosphate pathway in plastids. Sesquiterpenoids and sterols are predominantly synthesized in the cytosol by the mevalonate pathway whereas monoterpenoids, diterpenoids, the phytol side-chain of chlorophyll, carotenoids, and the nonaprenyl side-chain of plastoquinone-9 are synthesized within plastids by the deoxyxylulose phosphate pathway. Our assumptions that the early stages of gibberellin biosynthesis are plastid-localized has led to several attempts to demonstrate that the deoxyxylulose phosphate pathway is the biosynthetic route to gibberellins. Although definitive evidence is still not available there is a growing body of evidence, mostly from transgenic plants and from the use of the inhibitor, fosmidomycin, that gibberellins are synthesized from deoxyxylulose phosphate-derived isopentenyl diphosphate. However, there is evidence that a small amount of cross-talk between the two pathways may occur, implying that the pathways are not totally autonomous. Implications for the regulation of the early stages of gibberellin biosynthesis are discussed.

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In vitro binding of gibberellin A(4) to extracts of cucumber measured by using DEAE-cellulose filters.

A rapid method for assaying [(3)H]gibberellin A(4) bound to a soluble protein from cucumber hypocotyls by using DEAE-cellulose filter discs is described. The binding is saturable, reversible with unlabeled gibberellin A(4), and has a half-life of association under nonequilibrium conditions at 0-4 degrees C of 6-7 min. By using this assay, the dissociation constant (K(d)) was estimated to be 70 nM and the number of binding sites, 0.4 pmol mg(-1) of soluble protein or 0.69 pmol g(-1) (fresh weight) of hypocotyls. The speed and reliability of the assay make it invaluable for kinetic studies involving competitors. Thus, it has been possible to show that gibberellins that are biologically active in a cucumber bioassay compete for binding to the same protein and their calculated affinity constants bear a direct relationship to their known activities in the cucumber bioassay. Gibberellins that are inactive in this bioassay and other plant hormones, such as indoleacetic acid, abscisic acid, and kinetin, show a noncompetitive interaction.

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Reduction of the Gibberellin Content of Pharbitis Seeds by CCC and After-Effects in the Progeny.

Plants of Pharbitis nil were treated with the growth retardant (2-chloroethyl) trimethylammonium chloride (CCC) shortly before and after anthesis. Fresh and dry weight of immature seeds were not affected by the CCC treatment.The level of gibberellin-like activity in Pharbitis seeds as compared to control seeds was strongly reduced by CCC application. The progenies of the treated plants also had a much reduced GA content in the seedling stage. These results are interpreted to indicate that CCC blocks gibberellin biosynthesis in higher plants, as it does in the fungus Fusarium.CCC applied via the roots accumulated in the immature seeds and was carried over to the following generation. Consequently, growth of CCC progenies was dwarfed and flower formation inhibited. Both phenomena were overcome by application of gibberellin A(3).Three gibberellin-like substances (called fractions I, II, and III) were present in Pharbitis seeds and could be separated by thin-layer chromatography. All 3 fractions were also present in seeds treated with CCC. Fractions II and III were present in much higher quantities than fraction I. Both fractions II and III promoted growth of d5 corn but only fraction II was active in dwarf peas grown under red light.

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Gibberellin-auxin interaction in pea stem elongation.

Joint application of gibberellic acid and indole-3-acetic acid to excised stem sections, terminal cuttings, and decapitated plants of a green dwarf pea results in a markedly synergistic growth response to these hormones. Synergism in green tall pea stem sections is comparatively small, although growth is kinetically indistinguishable from similarly treated dwarf sections.Gibberellin-induced growth does not appear to be mediated through its effect on auxin synthesis, since gibberellin pretreatment of dwarf cuttings fails to elicit an enhanced tryptophan-induced growth response of sections, whereas auxin-induced growth is strongly enhanced. Also, tryptophan-gibberellin synergism is not significant in sections and cuttings of green dwarf peas, while auxin-gibberellin synergism is.Administration of gibberellic acid prior to indole-3-acetic acid results in greatly increased growth. In reversed order, the application fails to produce any synergistic interaction. This indicates that gibberellin action must precede auxin action in growth regulation.

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Physiological effects of gibberellic Acid. X. The release of gibberellin-like substances by germinating barley embryos.

Barley embryos, completely free from endosperm, were excised from germinating grain at various times and allowed to diffuse into an aqueous medium for varying lengths of time. At the end of this time, the embryos and ambient solutions were separately extracted. Gibberellin-like activity in the extracts was determined with the barley endosperm bioassay using seed from the same variety, harvest and treatment schedule as was employed for the embryo diffusion experiments. Gibberellin-like substances were released by embryos throughout the 60 hour germination period, though at no time during this period could sufficient activity be extracted from the embryos themselves to account for the observed release. Solvent partitioning and chromatography identified at least one major acidic component migrating at an Rf similar to that of GA(3).It is concluded that the endogenous gibberellin-like substance(s) originates within the embryo during germination, and that the release of this substance(s) is temporally consistent with, and quantitatively sufficient to account for the in vivo endosperm mobilization response syndrome. A gibberellin-like substance is undoubtedly the endosperm mobilizing hormone.

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Gibberellin-like Activity in Berries of Seeded and Seedless Tokay Grapes.

Gibberellin-like activity in berries of both ;Tokay' (seeded) and ;Seedless Tokay' grapes (Vitis vinifera L.) at different stages of development was determined. Activity in both seeded and seedless berries was mainly in the acidic ethyl acetate fraction. In seeded berries there was very high activity beginning at the early fruit-set stage and persisting for about 3 weeks after which the activity fell to low level and vanished by the middle of July. On August 1 there was another activity peak. The activity in ;Seedless Tokay' was similar except the decline was considerably more rapid and a secondary peak was reached in mid-June instead of August. The gibberellin-like substances in the acidic ethyl acetate fraction occurred mainly between R(F) 0.3 and 0.6 when the chromatogram was developed with ammoniacal isopropanol.The gibberellin-like substances from both types of berries were active in dwarf pea, dwarf corn, and lettuce hypocotyl bioassays, but not in the cucumber hypocotyl test. The role of gibberellin-like substances in berry development is discussed.

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Antagonism of some gibberellin actions by a substituted pyrimidine.

From a comparison of the effects of seven growth retardants and abscisic acid (ABA) on various growth systems, it was found that the gibberellin-regulated growth of lettuce hypocotyls was uniquely inhibited by the growth retardant, alpha-cyclopropyl-alpha-(4-methoxyphenyl)-5-pyrimidine methanol (EL-531). Auxin-regulated growth of coleoptile sections was inhibited by Phosfon and only slightly by EL-531 and Alar. Cytokinin-regulated growth of Xanthium cotyledons showed little or no inhibition by any of the retardants. ABA was inhibitory in all three types of tests. The distinctive effects of EL-531 against gibberellin-stimulated growth and the general ability of gibberellic acid to relieve EL-531 inhibition suggest that this retardant acts in part against the gibberellin-stimulated growth system, but at a locus which discriminates between growth and nongrowth functions of gibberellic acid. It shows little or no antagonism of gibberellin actions which do not involve growth: the barley endosperm test and the Rumex leaf senescence test.

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