Distribution of Radioactivity from Exogenously Supplied [1-14C]Indol-3-yl-acetic Acid and [3, 4-3H (N)] Gibberellin A, in Geotropically-stimulated Picea abies (L.) Karst. Roots

The distribution of exogenously-supplied radioactive labelledindol-3-yl-acetic acid (IAA) and gibberellin A₁ (GA₁) in geotropicallystimulated roots of Norway spruce (Picea abies (L.) Karst.)has been demonstrated. Seedlings were positioned with theirroot tips in 2.1 x 10^–6 M [¹⁴C]IAA or 1.3 x 10^–8m ³H-GA₁ for 4 and 20 h, respectively. After geotropic stimulationfor 90 min in the horizontal position the root tips were cutlongitudinally in 50 µm thick sections, using a freeze-microtome.The radioactivity in the ¹⁴C-IAA treated roots occurred in higherconcentration in the lower than in the upper halves (ratio 1.25:1). A similar trend was observed in the [³H]GA₁-treated rootswhere the ratio lower: upper halves was 2.04: 1. The ratio ofradioactivity in right and left halves of vertical roots wasapproximately the same in roots supplied with [¹⁴C]IAA and [³H]GA₁(1.09: 1). The supplied radioactive compounds were analysed chromatographicallyafter extraction in methanol of 6 mm apical root segments. Onlya small fraction (7–8 per cent) of the supplied [¹⁴C]IAAwas revealed unchanged in the segments. The major part of thechromatographed, labelled compound has not been identified,but on basis of its R_F value it is suggested that it may beindol-3-acetyl-aspartic acid (IAAasp). The chromatographic analysis of the [³H]GA,-treated segmentsshowed that only small fractions of this gibberellin has beenconverted to other compounds. These results have been discussed and correlated with knowledgeof plant growth regulators and their participation in root geotropism. Picea abies, spruce, geotropism, gibberellin A₁, indol-3-yl-acetic acid, growth regulators, redistribution in roots     

Abscisic Acid and Apical Dominance in Phaseolus coccineus L. The Role of Tissue Age

Abscisic acid (ABA) applied to the decapitated second internodeof runner bean plants enhanced outgrowth of lateral buds onlywhen internode stumps were no longer elongating. Applied toelongating internodes of slightly younger plants, ABA causesinhibition of bud outgrowth. Together with 10^–4 M indol-3-ylacetic acid (IAA), ABA stimulated internode elongation and interactedadditively in the inhibition of bud outgrowth. A mixture of10^–5 M ABA and 10^–6 M gibberellic acid (GA₃ ) causedsimilar effects on internode growth as IAA + ABA, but was mutuallyantagonistic in effect on growth of the lateral buds. Abscisic acid, apical dominance, gibberellic acid, indol-3yl acetic acid, Phaseolus coccineus, bean     

Effects Produced on Tomato Plants, Lycopersicum esculentum, by Seed or Root Treatment with Gibberellic Acid and Indol-3yl-Acetic Acid

Growth in lengths of tomato stems and leaves was acceleratedby 5.0 µg gibberellic acid (GA₂) applied to the seed,or by 5.0, 0.5, and 0.05 µg given to the roots. Treatmentwith 5.0 µg also decreased bud number and lengthened thetime between bud appearance and fruit formation on the firsttruss by 1–8 days. Smaller amounts applied to roots shortenedthis time by 1–4 days. Indol-3yl-acetic acid at 0.5 µghad no effect, nor was simultaneous application of GA₃ and IAAto the roots more effective than GA₃, alone. Single applicationsof very small amounts of GA₃ to seeds or seedling roots thusproved capable of changing growth-rates of stems, leaves, andtrusses.The effects of treating tomatoes with GA₂ and with culturesof Azotobacter chroococcum, which contain small amounts of GA₃,and IAA, are compared.     

Effects of Auxin and Gibberellin on Conidial Germination and Elongation of Young Hyphae in Gibberella fujikuroi and Penicillium notatum

In Gibberella fujikuroi and Penicillium notatum, IAA, 2,4-Dand GA₃ promoted conidial germination and the elongation ofyoung hyphae. The promotive effects of IAA and GA₃ were additive.In both fungi, the concentrations of endogenous auxin and gibberellinin the culture media were 10^–10 to 610^–12M. (Received April 27, 1985; Accepted August 12, 1985)     

Effects of Gibberellic Acid on the Activation, Synthesis, and Release of Acid Phosphatase in Barley Seed

Acid phosphatase activity was present in unimbibed barley seed,but rose during incubation of embryoless half-seeds and isolatedaleurone layers, and was further increased by 10^–6 M gibberellicacid (GA₃). Release of total acid phosphatase activity fromhalf-seeds and aleurone layers was markedly enhanced by GA₃.Inhibitor studies with cycloheximide and actinomycin D suggestedthat de novo synthesis of acid phosphatase occurred followingimbibition. Gel nitration, electrophoresis, and [¹⁴C]leucineincorporation studies revealed that a single molecular formof acid phosphatase was present in dry seed, whereas on incubationtwo further forms arose. A proportion of the three molecularforms of the enzyme was synthesized de novo. Gibberellic acidstimulated activation, but not de novo synthesis, of all threemolecular forms of acid phosphatase. Although a small amountof one of the molecular forms was secreted in the absence ofGA₃, the presence of gibberellin greatly increased secretionof the same form of acid phosphatase.     

Identification and Quantification of Cambial Region Hormones of Eucalyptus globulus

Gibberellins GA₁, GA₄, GA₈, GA₉, GA₁₉, GA₂₀, GA₂₉, GA₄₄, GA₈₁,indole-3-acetic acid (IAA) and abscisic acid (ABA) were identifiedin cambial region tissues of Eucalyptus globulus by comparingmass spectra and Kovats retention indices with those of authenticstandards. Using stable isotope labelled internal standardsGA₁₉, GA₂₀ and GA₄₄ were quantified at levels of 2–7 ng(g fr wt)^-1, other GAs were present at levels < 1 ng (g frwt)^-1. Levels of IAA and ABA ranged from 417–1, 140 ng(g fr wt)^-1 and 86–305 ng (g fr wt)^-1 respectively. Thepresence of brassinosteroid-like substances was also indicatedbased on activity in the rice seedling leaf inclination assay. (Received April 28, 1995; Accepted June 20, 1995)     

Effects of Auxin and Gibberellin on Elongation of Young Hyphae in Neurospora crassa

IAA, 2,4-D and GA₃ promoted the elongation of young hyphae inNeurospora crassa at the optimum concentrations of 10^–6,10^–6 and 10^–4 M, respectively. The effects of IAAand GA₃ were additive. (Received June 17, 1983; Accepted December 22, 1983)     

Potassium Transport in Non-Growing Corn Root Tissue as Affected by IAA and GA3

Experiments were done to determine if the spontaneous recoveryof non-growing segments of corn root (Zea mays L.) from excisioninjury is dependent on auxin. Washing the segments with 5 runindoleacetic acid (IAA) for 2 to 4 hours gave a small but significantincrease in K⁺ (⁸⁶Rb) influx, used here as a parameter reflectingrecovery of electrogenie H⁺-efflux pumping. This promotive effectwas obtained only after an hour of washing, and was sustainedby 100 nm gibberellic acid (GA₃). Any early responses to auxinwere obscured by an adverse reaction of the root cells to externalIAA which resulted in a transitory inhibition of H⁺ pumpingand K⁺ influx. Pretreatment of excised root tips with 10 µM IAA in thegrinding medium protected a plasmalemma-enriched fraction ofthe microsomes during isolation, giving increased uncoupler-sensitiveATPase activity. Non-growing root tissue thus shows three responses to auxin:an adverse reaction at the outer surface of the plasmalemmawhich blocks H⁺ pumping; a protective or restorative effecton the H⁺-ATPase; an increased capacity for K⁺ influx duringthe developmental phase of washing, which is augmented by thepresence of GA₃. (Received March 31, 1986; Accepted September 8, 1986)     

Effects of Plant Growth Regulators on Grain-filling and Yield of Rice

Effects of three phytohormones (IAA₁, GA₃ and kinetin) on grain-fillingand the pattern of ³²P translocation from individual leavesto grains were studied at intervals of 7 days during the progressof reproductive development of rice (Oryza saliva L. cv. Jaya).The plants were sprayed with 100 µg ml^–1 aqueoussolutions of the hormones at 100 days, when the plants wereentering the reproductive stage. Kinetin produced a pronouncedeffect on grain-filling as well as on ³²P mobilization fromindividual leaf to grains and increased yield, possibly by increasingleaf longevity. GA₃ and IAA also increased the grain-fillingand ³²P mobilization significantly over control but the effectswere less marked than those of kinetin. Oryza sativa L., rice, grain yield, translocation, growth regulators, gibberellic acid, indol-3-yl acetic acid, kinetin     

Effects of a Plant-Growth Regulator, Prohexadione, on the Biosynthesis of Gibberellins in Cell-Free Systems Derived from Immature Seeds

Inhibition of the biosynthesis of gibberellins by prohexadione,3,5-dioxo-4-propionylcyclo-hexanecarboxylic acid, was studiedwith cell-free systems derived from immature seeds of Cucur-bitamaxima, Phaseolus vulgaris and Pisum sativum. Prohexadione,at a concentration of 10–⁴ M, inhibited C-7 oxidationof GA₁₂-aldehyde, C-20 oxidation of GA₁₅, conversion of C₂₀-gib-berellinsto C₁₉-gibberellins, 3ß-hydroxylation, 2,3-dehydrogenationof GA²⁰, 2,3-epoxidation of GA₅ and 2ß-hydroxylationof GA₉ and GA₂₀. The 3ß-hydroxylase activity appearedto be more sensitive to prohexadione than were the C-20 oxygenaseand the 2ß-hydroxylase activities. The conversionof mevalonic acid to GA₁₂-aldehyde and the 13-hydroxylationof GA₁₂ were not affected by prohexadione at a concentrationof 3 ? 10^–4 M. All of the steps inhibited by prohexadioneare oxidation steps catalyzed by soluble enzymes that require2-oxoglutarate, Fe²⁺ and oxygen, and all of them occur distalto the synthesis of GA₁₂-aldehyde in the biosynthesis of gibberellins. (Received April 4, 1990; Accepted September 14, 1990)     

Partial Purification and Characterization of Gibberellin 3{beta}-hydroxylase from Immature Seeds of Phaseolus vulgaris L.

Gibberellin 3/ß-hydroxylase,a 2-oxoglutarate-dependentdioxygenase that catalyzes the hydroxylation of GA₂₀ to GA₁,was purified 313-fold from immature seeds of Phaseolus vulgarisL. The mol wt of the enzyme was estimated to be 42,000 by gelfiltration HPLC and SDS-polyacrylamide gel electrophoresis.The enzyme exhibited maximum activity at pH 7.7. The Km valuesfor [2,3-³H]GA20 and [2,3-³H]GA, were 0.29µu and 0.33µm, respectively. The enzyme requires 2-oxoglutarate asa cosubstrate; the K_m value for 2-oxoglutarate was 250µMusing [³H]- GA₂₀ as a substrate. Fe²⁺ and ascorbate significantlyactivated the enzyme at all purification steps, while catalaseand BSA activated the purified enzyme only. The enzyme was inhibitedby divalent cations Mn²⁺, Co²⁺, Ni²⁺, Cu²⁺, Zn²⁺, Cd²⁺ and Hg²⁺.3ß-Hydroxylation of [³H]- GA₂₀ was also inhibitedby non-radioactive GA₅, GA₉,GA₁₅, GA₂₀ and GA₄₄. The possiblesite of 3ß-hydroxylation in gibberellin biosynthesisis discussed in terms of the substrate specificity of partiallypurified gibberellin 3ß-hydroxylase. (Received February 29, 1988; Accepted June 3, 1988)     

Inoculation with Azospirillum lipoferum Affects Growth and Gibberellin Status of Corn Seedling Roots

Maize (Zea mays L., hybrid Cargill 147) seedlings, grown inaseptic conditions, were inoculated with three strains of Azospirillumlipoferum (Al op 33, Al iaa 320, and ATCC 29708) or culturedin different concentrations of indol-3-acetic acid (IAA) orgibberellin A₃ (GA₃). After 48 h, root length, root surfacearea, root dry weight, and shoot dry weight were measured inall treatments. Gibberellin content was evaluated in selectedroots of control and Azospirillum inoculated seedlings by gaschromatography-mass spectrometry-selected ion monitoring withthe use of deuterated gibberellins as internal standards. Thethree strains of A. lipoferum, IAA (2 ng ml^–1), and GA₃(40 to 400 pg ml^–1) significantly enhanced root growth.Improvement of root hair growth and density was obtained mainlywith A. lipoferum strain Al op 33 and GA₃ 40 pg ml^–1.GA₃ was identified by gas chromatography-mass spectrometry (byboth, full scan and selected ion monitoring) in a free acidfraction from roots of the seedlings inoculated with A. lipoferum.In the roots of the non inoculated seedlings GA₃ was found afterhydrolysis of a fraction expected to contain glucosyl conjugates. (Received April 26, 1993; Accepted September 27, 1993)     

The Effect of Gibberellic Acid on Starch Breakdown in Sprouting Tubers of Solanum tuberosum L.

The treatment of potato tubers with 150 µmol dm^–3gibberellic acid (GA₃) stimulated starch breakdown and hexoseaccumulation in tuber tissues and the transfer of dry matterto stems. These effects could not be accounted for by enhancedactivities of starch phosphorylase, amylase and acid invertase.Indeed enzyme activities either declined or remained relativelyconstant as starch degradation and hexose accumulation proceeded.Changes in the rate of starch depletion were related to changesin sink strength and sink type, the onset of tuber initiationin controls causing the rate of starch degradation to exceedthat in GA₃-treated tissues, in which tuberization was inhibited. Solanum tuberosum L., gibberellic acid, starch breakdown     

Interaction of a Brassinosteroid with IAA and GA3 in the Elongation of Cucumber Hypocotyl Sections

A synthetic brassinosteroid, 22,23(S,S)-homobrassinolide (hBR),was examined for its interaction with IAA and GA₃ in the elongationof hypocotyl sections of light-grown cucumber (Cucumis salivusL. cv. Aonagajibai) seedlings. hBR alone was less active thanIAA. Its optimal concentration was around 10 µM and thelowest effective concentration between 10 and 100 µM,which is more than 100 times higher than that of brassinolide.hBR was more active in sections from younger seedlings. Itsgrowth-promoting effect was negated or greatly reduced by inhibitorsof auxin-induced elongation such as p-chlorophenoxyisobutyricacid and kinetin. hBR acted synergistically with IAA and 2,4-Dbut not with GA₃ showing only an additive effect. Sequentialtreatment of sections with hBR and then with IAA also resultedin synergistic enhancement of auxininduced elongation, but whenthe order of treatment was reversed, hBR was inactive. The synergisticeffect was obtained with 1 h pretreatment with hBR and couldbe reduced by subsequent washing with water. There was no sequentialinteraction between hBR and GA₃. The synergistic pretreatmenteffects of hBR and GA₃ were simply additive to each other. Amembrane-bound ATPase inhibitor, dicyclohexylcarbodiimide, inhibitedthe hBR-induced elongation, but did not affect GA₃-induced elongation.The findings led to the conclusion that brassinosteroids enhanceauxin action and possess growth-promoting activity which isindependent of that of gibberellin. (Received November 9, 1984; Accepted February 18, 1985)     

Differential modulation of caffeine- and IP3-induced calcium release in cultured arterial tissue

To investigatethe Ca²⁺-dependent plasticity ofsarcoplasmic reticulum (SR) function in vascular smooth muscle,transient responses to agents releasing intracellularCa²⁺ by either ryanodine(caffeine) orD-myo-inositol1,4,5-trisphosphate [IP₃;produced in response to norepinephrine (NE),5-hydroxytryptamine (5-HT), arginine vasopressin (AVP)] receptorsin rat tail arterial rings were evaluated after 4 days of organculture. Force transients induced by all agents were increased comparedwith those induced in fresh rings. Stimulation by 10% FCSduring culture further potentiated the force andCa²⁺ responses to caffeine (20 mM)but not to NE (10 µM), 5-HT (10 µM), or AVP (0.1 µM). The effectwas persistent, and SR capacity was not altered after reversibledepletion of stores with cyclopiazonic acid. The effects of serum couldbe mimicked by culture in depolarizing medium (30 mMK⁺) and blocked by the additionof verapamil (1 µM) or EGTA (1 mM) to the medium, loweringintracellular Ca²⁺ concentration([Ca²⁺]_i)during culture. These results show that modulation of SR function canoccur in vitro by a mechanism dependent on long-term levels of basal[Ca²⁺]_iand involving ryanodine- but notIP₃ receptor-mediatedCa²⁺release.     

Effect of Hormones on Sucrose Uptake and on ATPase Activity of Citrus sinensis L. Osbeck Leaves

Carbohydrate accumulation in young, fully expanded leaves ofCitrus sinensis L. Osbeck is affected by the presence of thefruitlet on the shoot. Previous work gave evidence that gibberellinsmay be involved in this 'fruit effect'. In the present workwe have studied the effect of gibberellic acid (GA₃) on ¹⁴C-sucroseuptake by leaf discs and whether its action could be due toa modulation of the plasma membrane ATPase, which maintainsthe H⁺ gradient that drives H⁺/sucrose co-transport. The effect of GA₃ on ¹⁴C-sucrose uptake depended on the osmolarityof the assay medium. At 300 mOsm a reduction in the uptake ratewas observed. The inhibitory effect of the hormone disappearedafter preincubating the leaf discs with para-chloromercuri-phenylsulphonicacid (PCMPS), a sulphydril binding inhibitor. ATPase activityof isolated plasma membrane vesicles was inhibited by IAA treatments,while GA₃ or ABA did not affect this enzyme, even after a 3h preincubation period. However, in the absence of a surfactantin the assay medium, GA₃, together with turgor pressure, modulatedplasma membrane ATPase activity, possibly through modificationsof membrane permeability. The hormone effect on ¹⁴ C-sucroseuptake may involve action on the sucrose carrier.Copyright 1994,1999 Academic Press Abscisic acid, Citrus sinensis, gibberellic acid, indoleacetic acid, orange, osmotic pressure, plasma membrane ATPase, ¹⁴C-sucrose uptake     

Effects of Some Growth Regulators and Benzoic Acid Derivatives on Flower Initiation and Root Elongation of Pharbitis nil, Strain Kidachi

Seedlings of Pharbitis nil, strain Kidachi, were grown undercontinuous light at 20°C in vessels containing 5,000-mlnutrient solution, 24 plants per vessel. NAA (0.005–0.5µM), GA₃ (0.1–0.5 µM), kinetin (0.5–5µM), benzyladenine (0.05–5 µM) or abscisicacid (4 µM) added to the nutrient solution induced long-dayflowering, and the flowering was always accompanied by suppressionof root elongation. 3,4-Dichlorobenzoic acid (0.05–10µM) and some other benzoic acid derivatives which arehighly effective for the induction of flowering in Lemna paucicostataalso showed similar effects. Neither NAA, kinetin nor 3,4-dichlorobenzoicacid applied via the apical part of the hypocotyl could causeflowering or suppression of root elongation. Thus, the flower-inducingeffect of the above substances was presumed to be secondaryto the suppression of root elongation. Ethrel (1–50 µM)added to the nutrient solution suppressed root elongation, butdid not induce flowering probably because it has flower-inhibitingactivity. ¹ This paper is dedicated to the memory of Dr. Joji Ashida,the first president of the Japanese Society of Plant Physiologists. (Received December 15, 1982; Accepted February 25, 1983)     

SH oxidation coordinates subunits of rat brain ryanodine receptor channels activated by calcium and ATP

We have reported that ryanodine receptor (RyR) channels display three different responses to cytoplasmic free Ca²⁺ concentration ([Ca²⁺]) depending on their redox state (Marengo JJ, Hidalgo C, and Bull R. Biophys J 74: 1263–1277, 1998), with low, moderate, and high maximal fractional open times (P_o). Activation by ATP of single RyR channels from rat brain cortex was tested in planar lipid bilayers with 10 or 0.1 µM cytoplasmic [Ca²⁺]. At 10 µM [Ca²⁺], low-P_o channels presented lower apparent affinity to activation by ATP [[ATP] for half-maximal activation (K_aATP) = 422 µM] than moderate-P_o channels (K_aATP = 82 µM). Oxidation of low-P_o channels with thimerosal or 2,2'-dithiodipyridine (DTDP) gave rise to moderate-P_o channels and decreased K_aATP from 422 to 82 µM. At 0.1 µM cytoplasmic [Ca²⁺], ATP induced an almost negligible activation of low-P_o channels. After oxidation to high-P_o behavior, activation by ATP was markedly increased. Noise analysis of single-channel fluctuations of low-P_o channels at 10 µM [Ca²⁺] plus ATP revealed the presence of subconductance states, suggesting a conduction mechanism that involves four independent subchannels. On oxidation the subchannels opened and closed in a concerted mode. subconductance states; calcium ion release channels; calcium ion regulation; thimerosal; 2,2'-dithiodipyridine     

Involvement of cellulose synthesis in actions of gibberellin and kinetin on cell expansion. Gibberellin-coumarin and kinetin-coumarin interactions on stem elongation

In azuki bean (Azukia angularis = Vignia angularis) epicotylsections, 5 ? 10^–4 M coumarin inhibited the incorporationof radioactivity from [U^–14C]glucose into the cellulosefraction by 35% in the absence of indole-3-acetic acid (IAA)and by 40% in the presence of 1 ? 10^–4 M IAA. There wasno inhibitory effect on the incorporation of radioactivity intothe other fractions. Coumarin at 5 ? 10^–4 M reversed thepromoting effect of 1 ? 10^–5 M gibberellin A₃ (GA) andthe inhibitory effect of 1 ? 10^–5 M kinetin on IAA-inducedelongation of sections with no significant effects on IAA-inducedelongation. Neither GA nor kinetin had any appreciable effectson cellulose synthesis. No inhibition of cellulose syntheiswas observed with 1 ? 10^–3 M colchichine, which has beenreported to have effects similar to those of coumarin on GA-or kinetin-affected stem elongation. Coumarin at 5 ? 10^–4M was ineffectual in breaking up wall microtubules, while adisrupting effect on wall microtubules was clearly demonstratedwith 3 ? 10^–4M colchicine. From these results, the possible involvement of cellulose synthesisin cell expansion controlled by GA or kinetin was suggested. (Received August 3, 1973; )     

Ethylene Biosynthesis and Xylogenesis in Lactuca Pith Explants Cultured In Vitro in the Presence of Auxin and Cytokinin: The Effect of Ethylene Precursors and Inhibitors

The possible involvement of ethylene in the induction of xylemdifferentiation was studied in lettuce (Lactuca saliva L. cv.Romaine) pith parenchyma explants. The addition of the ethyleneprecursors L-methionine (0.25 µM), S-adenosylmethionine(25 µM) and 1-aminocyclopropane-l-carboxylic acid (0.01µM), or the ethylene-releasing agent 2-chloroethylphosphonicacid (1.0 µM), to a standard IAA-kinetin-containing mediumenhanced xylogenesis compared to control explants cultured inthe absence of these compounds. In the presence of the ethyleneinhibitors aminoethoxyvinylglycine, Co(NO₃)₂ and AgNO₃, xylogenesiswas inhibited. Inhibition of xylogenesis by aminoethoxyvinylglycine(75 µM), Co(NO₃)₂ (50 µM) and AgNO₃ (6.0 µM)was reversed by exogenous 1-aminocyclopropane-l-carboxylic acid(0.01 µM), 2-chloroethylphosphonic acid (5.0 µM)and L-methionine (0.25 µM), respectively. Ethylene productionby explants cultured on media containing L-methionine or 1-aminocyclopropane-l-carboxylicacid was greater than the biosynthesis of ethylene by explantscultured in the absence of these compounds. The incorporationof 2-chloroethylphosphonic acid into the culture medium resultedin higher rates of ethylene production compared to explantscultured on the IAA-kinetin medium. The presence of either aminoethoxyvinylglycineor Co(NO₃)₂ inhibited ethylene production by explants culturedon the IAA-kinetin medium. The data support the hypothesis thatethylene plays a positive role in the initiation of xylem differentiation. Key words: Xylogenesis, Differentiation, Ethylene, IAA, Kinetin, Lactuca sativa