Influence of gibberellic acid on auxin biosynthesis and their effects on coleoptile elongation in garlic

To demonstrate the effect of auxin on intact coleoptile growth, garlic (Allium sativum L.) cloves were inoculated in agar supplemented with DW (control), GA₃ and GA₃+tryptophan (a precursor of IAA, GA₃+T). The coleoptiles were harvested at 24 h intervals to measure growth in terms of length, activities of IAAld DH (which convert tryptophan to IAA) and peroxidase (that oxidizes IAA). Contents of endogenous IAA and PAA were also measured by indirect ELISA. Peroxidase activity was suppressed by GA₃ treatment and increased by GA₃+T treatment. Although endogenous contents of IAA were increased by the addition of GA₃ and even more by GA₃+T in the media, there was no further increase in coleoptile length, suggesting that garlic coleoptiles are sufficient in their production of IAA.     

Interaction of Gibberellic Acid and Indole-3-acetic Acid in the Growth of Excised Cuscuta Shoot Tips in Vitro

Gibberellic acid (GA₃) induced a marked elongation of 2.5-centimeter shoot tips of Cuscuta chinensis Lamk. cultured in vitro. In terms of the absolute amount of elongation, this growth may be the largest reported for an isolated plant system. The response to hormone was dependent on an exogenous carbohydrate supply. The hormone-stimulated growth was due to both cell division and cell elongation. The growth response progressively decreased if GA₃ was given at increasingly later times after culturing, but the decreased growth response could be restored by the application of indole-3-acetic acid (IAA) to the apex. Explants deprived of GA₃ gradually lost their ability to transport IAA basipetally, but this ability was also restored by auxin application. The observations are explained on the basis that: (a) the growth of Cuscuta shoot tip in vitro requires, at least, both an auxin and a gibberellin; and (b) in the absence of gibberellin the cultured shoot tip explants lose the ability to produce and/or transport auxin.     

Effects of Auxin Transport Inhibitors on Gibberellins in Pea

The effects of the auxin transport inhibitors 2,3,5-triiodobenzoic acid (TIBA), 9-hydroxyfluorene-9-carboxylic acid (HFCA), and 1-N-naphthylphthalamic acid (NPA) on gibberellins (GAs) in the garden pea (Pisum sativum L.) were studied. Application of these compounds to elongating internodes of intact wild type plants reduced markedly the endogenous level of the bioactive gibberellin A₁ (GA₁) below the application site. Indole-3-acetic acid (IAA) levels were also reduced, as was internode elongation. The auxin transport inhibitors did not affect the level of endogenous GA₁ above the application site markedly, nor that of GA₁ precursors above or below it. When plants were treated with [¹³C,³H]GA₂₀, TIBA reduced dramatically the level of [¹³C,³H]GA₁ recovered below the TIBA application site. The internodes treated with auxin transport inhibitors appeared to be still in the phase where endogenous GA₁ affects elongation, as indicated by the strong response to applied GA₁ by internodes of a GA₁-deficient line at the same stage of expansion. On the basis of the present results it is suggested that caution be exercised when attributing the developmental effects of auxin transport inhibitors to changes in IAA level alone. Received April 13, 1998; accepted April 14, 1998     

Short-term effects of plant hormones on membrane potential and membrane permeability of dwarf maize coleoptile cells (Zea mays L. d 1) in comparison with growth responses

The membrane potential difference of dwarf maize coleoptile cells is increased by both 10^-5moll^-1 gibberellic acid (GA₃) and indoleacetic acid (IAA) a few minutes after application. A final level is reached after 10–20 min. The membrane permeability ratio P _Na:P _K is altered by both hormones during the first 15 min after application, indicating a rapid effect on the membrane. Elongation growth of coleoptile segments, however, is only stimulated by IAA. The auxin-induced growth as well as the auxin effect on membrane permeability depends on the calcium ion concentration of the medium. It is concluded that IAA acts via a proton extrusion pump that is electrically balanced by a potassium ion uptake, driven by the electromotive force of the pump. The mode of action of GA₃ on elongation growth is assumed to involve a process that depends on the physiologic state of the tissue and/or metabolic energy.Abbreviations IAA indoleacetic acid - GA₃ gibberellic acid - FC fusicoccin - PD electric potential difference between the vacuole and the external medium     

In vivo evidence for the involvement of phospholipase A and protein kinase in the signal transduction pathway for auxin-induced corn coleoptile elongation

Auxin-induced elongation of com coleoptiles is accompanied by cell wall acidification, which depends upon H⁺-pump activity. We tested the hypothesis that phospholipase A and a protein kinase are involved in the pathway of auxin signal transduction leading to H⁺ secretion, and elongation of corn coleoptiles. Initially, the pH of the bath solution at 50–100 μm from the surface of a coleoptile segment (pH_o) ranged between 4.8 and 6.6 when measured with an H⁺-sensitive microelectrode. Twenty or 50 μM lysophosphatidylcholine, 50 μM linolenic acid or 50 μM arachidonic acid induced a decline in pH_o by 0.3 to 2.1 units. The effect was blocked by 1 mM vanadate, suggesting that lysophosphatidylcholine or linolenic acid induced acidification of the apoplast by activating the H⁺-pump. Lysophosphatidylcholine and linolenic acid also accelerated the elongation rate of the coleoptiles. While linolenic acid and arachidonic acid, highly unsaturated fatty acids, promoted pH_o decrease and coleoptile elongation, linoleic acid, oleic acid, and stearic acid, fatty acids with a lesser extent of unsaturation, had no such effects. The effects of lysophosphatidylcholine, linolenic acid, and arachidonic acid on H⁺ secretion were not additive to that of indoleacetic acid (IAA), suggesting that lysophospholipids, fatty acids and auxin use similar pathways for the activation of the H⁺-pump. The phospholipase A₂ inhibitors, aristolochic acid and manoalide, inhibited the IAA-induced pH_o decrease and coleoptile elongation. The general protein kinase inhibitors, H-7 or staurosporine, blocked the IAA- or lysophosphatidylcholine-induced decrease in pH_o. H-7 also inhibited the coleoptile elongation induced by IAA or lysophosphatidylcholine. These results support the hypothesis that phospholipase A is activated by auxin, and that the products of the enzyme, lysophospholipids and fatty acids, induce acidification of the apoplast by activating the H⁺-pump through a mechanism involving a protein kinase, which in turn promotes com coleoptile elongation.     

Biogenesis of auxin in the coleptile of a semi-brachytic barley, uzu

The coleoptile of a semi-brachytic barley, uzu(Hordeum vulgareL. cv. Akashinriki), elongated ca 1/2 as much as the coleoptileof the contrasting normal form which is isogenic excepting theuz gene. This retarded growth of the uzu coleoptile, as comparedto the normal coleoptile, is not due to changes in the rateof basipetal transport of auxin, neither to the destructionof auxin during transport, nor to sensitivity to auxin. Furthermore,less of the extractable and bound auxins were found in the uzu(uzuz)coleoptile than in the normal(UZUZ)coleoptile, suggesting thatthe retarded growth of uzu coleoptile may be due to less auxinproduction. Apical tips of the normal coleoptile grown under sterile conditionsresponded to both tryptophan and tryptamine, but uzu coleoptiletips responded only to tryptamine. Thus, growth retardationof the uzu coleoptile may be due to lower activity of the enzymewhich converts tryptophan to tryptamine in the uzu coleoptile. (Received August 20, 1973; )     

Auxin-Gibberellin Interactions in Pea: Integrating the Old with the New

Recent findings on auxin-gibberellin interactions in pea are reviewed, and related to those from studies conducted in the 1950s and 1960s. It is now clear that in elongating internodes, auxin maintains the level of the bioactive gibberellin, GA₁, by promoting GA₁ biosynthesis and by inhibiting GA₁ deactivation. These effects are mediated by changes in expression of key GA biosynthesis and deactivation genes. In particular, auxin promotes the step GA₂₀ to GA₁, catalyzed by a GA 3-oxidase encoded by Mendel’s LE gene. We have used the traditional system of excised stem segments, in which auxin strongly promotes elongation, to investigate the importance for growth of auxin-induced GA₁. After excision, the level of GA₁ in wild-type (LE) stem segments rapidly drops, but the auxin indole-3-acetic acid (IAA) prevents this decrease. The growth response to IAA was greater in internode segments from LE plants than in segments from the le-1 mutant, in which the step GA₂₀ to GA₁ is impaired. These results indicate that, at least in excised segments, auxin partly promotes elongation by increasing the content of GA₁. We also confirm that excised (light-grown) segments require exogenous auxin in order to respond to GA. On the other hand, decapitated internodes typically respond strongly to GA₁ application, despite being auxin-deficient. Finally, unlike the maintenance of GA₁ content by auxin, other known relationships among the growth-promoting hormones auxin, brassinosteroids, and GA do not appear to involve large changes in hormone level.     

PHYTOCHROME ACTION IN ORYZA SATIVA L. III. THE SEPARATION OF PHOTOPERCEPTIVE SITE AND GROWING ZONE IN COLEOPTILES, AND AUXIN TRANSPORT AS EFFECTOR SYSTEM

• 1 In 4-day-old etiolated rice seedlings, 3 mm of the coleoptile tip did mainly perceive the photostimulus to cause the phytochrome-dependent inhibition of coleoptile elongation. At this age, cell elongation occurred most in the middle portion of coleoptiles in the dark, and was reversibly controlled by a brief exposure of the tip to red and far-red light. Thus, the photoperceptive site was evidently separated from the growing zone in intact rice coleoptiles.
• 2 The red-light-induced inhibition of coleoptile elongation was nullified by the removal of tip followed by the exogenous application of IAA. The sensitivity of thus treated coleoptiles to IAA was gradually lost during intervening darkness between the irradiation and the decapitation, and a 50% loss was obtained at ca. 6th hour at 26°C.
• 3 Polar auxin transport from coleoptile tips was remarkably prevented at the period between, at least, 2nd and 4th hour after red irradiation, and it recovered to the level of dark control by the 6th hour. Far-red light given immediately after red irradiation reversed the yield of diffusible auxin up to that of far-red control.

     

The mode of action of a morphactin,fluoren-9-carboxylic acid

Fluoren-9-carboxylic acid acts not only as an auxin but also as an gibberellin-antagonist. In the standard pea straight test (S₅ section) for auxin it stimulated elongation, the optimum concentration being 10 mg/l. On the other hand, it inhibited elongation at 0.1 mg/l. This inhibitory effect was more marked when younger tissue (S₁ section) which also responds to gibberellin was used. Interaction of FCA and IAA in the S₅ section has shown that at higher concentration of IAA there seemed to be a suppraoptimal effect, indicating that FCA acted as an auxin. However, in the S₁ section, the stimulating effect of GA₃ was markedly inhibited by 0.1 mg/l FCA; 10 mg/l FCA was either additive or less than additive to GA₃. In the cucumber hypocotyl test FCA itself was inactive up to 100 μg/plant, but it inhibited the GA₃-induced elongation. This inhibition was overcome by increasing the dosage of GA₃. In the same material, the IAA-induced elongation was not affected by FCA. These results indicate that whether FCA acts as an auxin or a gibberellin-antagonist depends on whether the tissue is sensitive to gibberellin and/or auxin.     

Stem elongation and changes in the levels of gibberellins in shoot tips induced by differential photoperiodic treatments in the long-day plant Silene armeria

The effects of differential photoperiodic treatments applied to shoot tips and mature leaves of the long-day (LD) plant Silene armeria L. on growth and flowering responses, and on the levels of endogenous gibberellins (GAs), were investigated. Gibberellins were analyzed by gaschromatography-mass spectrometry and the use of internal standards. Exposure of mature leaves to LD, regardless of the photoperiodic conditions of the shoot tips, short days (SD), LD, or darkness, promoted elongation of the stems and of the immature leaves. Long-day treatment of the mature leaves modified the levels of endogenous GAs in shoot tips kept under LD, SD, or darkness. In shoot tips kept in LD or darkness the levels of GA₅₃ were reduced, whereas the levels of GA₁₉ and GA₂₀ were increased. The contents of GA₁ were increased in all three types of shoots: SD twofold, LD fivefold, and darkness eightfold. Dark treatment of the shoot tips on plants of which the mature leaves were grown in SD promoted elongation of the immature etiolated leaves and increased the GA₁ content of the shoot tips threefold. However, this treatment did not cause stem elongation. The different photoperiodic treatments applied to the shoot tips did not change the levels of GAs in mature leaves. These results indicate that both LD and dark treatments result in an increase in GA₁ in shoot tips. In addition, it is proposed that LD treatment induces the formation of a signal that is transmitted from mature leaves to shoot tips where it enhances the effect of GA on stem elongation.Abbreviations GA_n gibberellin A_n - LD long day(s) - SD short day(s) We thank Dr. L.N. Mander, Australian National University, Canberra, for providing [²H]-gibberellins and Dr. D.A. Gage, MSU-NIH Mass Spectrometry Facility, East Lansing, for advice with mass spectrometry. This work was supported, in part, by a fellowship from the Spanish Ministry of Agriculture (Instituto Nacional de Investigaciones Agrarias) to M.T., by the U.S. Department of Energy grant No. DE-FG02-91ER20021, and by the U.S. Department of Agriculture grant No. 88-37261-3434 to J.A.D.Z.     

Role of auxin and gibberellin in differentiation of primary Phloem fibers

The hypothesis that auxin and gibberellic acid (GA₃) control the differentiation of primary phloem fibers is confirmed for the stem of Coleus blumei Benth. Indoleacetic acid (IAA) alone sufficed to cause the differentiation of a few primary phloem fibers. In long term experiments auxin induced a considerable number of fibers in mature internodes. GA₃ by itself did not exert any effect on fiber differentiation. Combinatiosn of IAA with GA₃ completely replaced the role of the leaves in primary phloem fiber differentiation qualitatively and quantitatively. Although the combined effect of the two growth hormones diminished considerably with increasing distance from the source of induction, auxin with GA₃ or IAA alone induced fibers in a few internodes below the application site. When various combinations of both hormones were applied, high concentrations of IAA stimulated rapid differentiation of fibers with thick secondary walls, while high levels of GA₃ resulted in long fibers with thin walls. The size of the primary phloem fibers correlated with the dimensions of the differentiating internode, thereby providing evidence that both growth regulators figure in the control of stem extension. High IAA/low GA₃ concentrations have an inhibitory effect on internode elongation, whereas low IAA/high GA₃ concentrations promote maximal stem elongation.     

Effects of light, temperature, gibberellin (GA3) and their interaction on coleoptile and leaf elongation of tall, semi-dwarf and dwarf wheat

Near-isogenic wheat lines differing in height-reducing (Rht) alleles, in each of two cultivars, were used to investigate the effects of light intensity and of their interaction with temperature and GA₃ application, on the elongation of the coleoptile and the first seedling leaf. Darkness caused a conspicuous increase in the lengths of the coleoptile and of the sheath and lamina of the first leaf, in GA₃ treated and untreated seedlings of all genotypes grown at 11 and 25°C. The genotype effects and the effects of light intensity and GA₃ application on leaf length were ascribed entirely to their effects on the rate of leaf elongation since the duration of leaf elongation was not affected by these factors. Temperature elevation from 11 to 25°C caused a 55% shortening of the duration of leaf elongation and a concomitant increase in elongation rate, which diminished with increased genotypic dwarfness. Accordingly, temperature elevation resulted in a significant reduction in leaf-length of the light-grown dwarf genotypes and the dark-grown dwarf and semi-dwarf genotypes. It is suggested that this temperature × light × genotype interaction effect is due to environmental dependent upper limits of elongation rate set by the Rht alleles.Abbreviations PAR Photosynthetic Active Radiation     

Effects of inorganic solutes on the binding of auxin

The binding of α-naphthaleneacetic acid (¹⁴C-NAA) to pelletable particulates from corn (Zea mays) coleoptiles was found to be influenced by inorganic solutes. La³⁺, Ca²⁺, and Mg²⁺ increased the binding whereas monovalent cations did not. The concentrations of CaCl₂ which increased auxin binding were similar to those which inhibited coleoptile elongation in the presence of auxin. These results are interpreted as suggesting that the alteration of hormonal effectiveness by some inorganic solutes involves alterations in the attachment of the hormone to binding sites in the cell.     

Effect of Ecdysterone on Morphological and Physiological Processes in Plants

In order to elucidate the physiological role of phytoecdysteroids in plants, we investigated the effects of exogenous ecdysterone (ECD) and phytohormones (IAA, GA₃, and 24-epibrassinolide (EBL)) on the growth of wheat coleoptiles and Arabidopsis thaliana seedlings (wild-type ecotype Columbia (Col) and its det2 mutant), on -amylase activity in the barley aleurone layer, and on the pigment content in the kidney bean senescent leaves. The range of effective ECD concentrations depended on the type of a reaction to be regulated. The regulation of growth processes was affected by a wide range of ECD concentrations (10^–13–10^–5 M), whereas some metabolic processes, such as the activation of -amylase and the retardation of leaf yellowing, by a narrow range, that is, 10^–9–10^–7 M and 10^–9–10^–8 M, respectively. We noted the synergetic effect of ECD and IAA on coleoptile elongation, the antagonistic effect of ECD and EBL on coleoptile elongation, as well as the antagonistic action of ECD and GA₃ on coleoptile elongation and -amylase activity. The data obtained demonstrate that ECD is a physiologically active compound. ECD might be supposed to act as a source of sterols or a regulator of IAA and protein synthesis. The effects of this regulator seems to be brought about by its interaction with the EBL and GA₃ receptors.     

The action of penicillin in mungbean (Phaseolus aureus L.) seedlings and wheat (Triticum vulgare L.) coleoptile sections

Penicillin caused elongation inhibition of mungbean (Phaseolus aureus L.) seedlings at concentrations above 100 mgl^-1. Inhibition of hypocotyl could be reduced to different degrees by manganese, pyruvate, succinate, fumarate, malate, GA₃, purines, pyrimidines and nucleosides, whereas the amino acids except cysteine were not effective. Penicillin inhibited neither elongation of wheat coleoptile sections nor the cell enlargement induced by IAA in this tissue. Thus penioillin inhibits intact seedling elongation probably through an Effect on cell division apparently without inhibiting cell enlargement.     

Effects of Brassin-Complex on Auxin and Gibberellin Mediated Events in the Morphogenesis of the Etiolated Bean Hypocotyl

The excised, hooked bean hypocotyl was the system used to determine wheiher the ‘auxin- and gibberellin like’ effect of the lipoidal pollen extract, Brass in-complex (Br), were mediated through, or independent of, auxin and gibberellin. The morphogenetic events of hook opening and hypocotyl elongation in this system are regulated by auxin and gibberellin, respectively. Brassin complex, like IAA, elicited a book closure in (he dark and retarded its opening in red light. This effect was synergized by T1BA, IAA and the presence of the auxin-producing organs, the epicotyl and cotyledons. Br-elicited hook closure was inhibited by the antiauxin. PCIB. Both GA₃ and Br totally reversed the light inhibition of hypocotyl elongation. The GA₃-effect, but nol the Br elicited elongation, was overcome by Ancymidol. Hypocotyl elongation was partially inhibited by TIBA and PCIB. suggesting a possible auxin involvement also in this effect of Br. Br may elicit its growth responses through an effect on endogenous auxin levels, In this way it is different from other lipoidat growth regulators, such as the oleanimins which require the presence of exogenous growth regulators for activity.     

Interactions between Plants and Epiphytic Bacteria Regarding Their Auxin Metabolism

More “diffusible” auxin is received from nonsterile than from sterile corn coleoptile tips. An artificial reinfection of sterile coleoptiles with epiphytic, IAA-producing bacteria strains does, a superinfection of nonsterile coleoptiles does not increase the auxin amount. The difference between sterile and nonsterile tips persists if diffusion from the coleoptile surface is excluded by covering the surface with a paraffin layer. The greater the distance from the apex, the higher becomes the superiority of nonsterile tips. An artificial bacterial contamination of the contact face between tip and receiver agar block, or addition of glucose and tryptophan to the agar block, do not influence the received auxin amount. Consequently the additional, bacteria-produced auxin delivered by the nonsterile tip is not produced at the cut surface or in the agar but is present in the tissues of the coleoptile tip.     

Effects of the steroidal alkaloid tomatine in auxin bioassays and its interaction with indole-3-acetic acid

Summary The steroidal alkaloid tomatine did not enhance elongation of oat coleoptile and first internode sections, or of wheat coleoptile sections. Higher concentrations of the alkaloid inhibited elongation and interacted antagonistically with IAA. Although 10^-4 M tomatine alone did not influence elongation of oat coleoptile sections, it did reduce growth response to exogenous IAA. Tomatine concentrations less than 10^-4 M did not influence response to IAA. The auxin activity of tomatine, reported by Vendrig, was therefore not confirmed.     

Estimation of endogenous contents of phytohormones during internode development in <Emphasis Type="Italic">Merremia emarginata</Emphasis>

During the entire period of internode growth of Merremia emarginata contents of gibberellic acid (GA₃), phenyl-acetic acid (PAA), indole-3-acetic acid (IAA, free and conjugated) and abscisic acid (ABA, free and conjugated) were estimated by ELISA using polyclonal antibodies raised against each hormones. At the time of internode elongation free auxin content was low and increased with the decrease in the rate of elongation. In contrast, conjugated IAA showed declining trend where free IAA content was remarkably high, suggesting thereby that conjugated IAA might have mobilized during the later phase of internode development. The endogenous GA₃ contents were high as compared to other hormones; however, no significant role of GA₃ was discernible in elongation growth. Conjugated ABA contents remained very low during the elongation growth and increased thereafter.