Auxin-stimulated ATPase in membrane fractions from pumpkin hypocotyls (Cucurbita maxima L.)

Membrane fractions from Cucurbita maxima hypocotyls were isolated in a medium which inhibits the action of endogenous phospholipases. After removal of soluble phosphatases by Sepharose 2B-CL column chromatography, an auxin-stimulated ATPase activity was found in membrane fractions from linear sucrose gradients. In the presence of 10^-4 M phenylacetic acid (PAA), the stimulation by indol-3-acetic acid (IAA) exhibited a bimodal concentration dependence with maximal stimulation of about 50% at 10^-6 M IAA. Without PAA, only a high concentration of 10^-4 M IAA was stimulatory, whereas 10^-6 M IAA had no apparent effect and 10^-8 M IAA exhibited weak inhibition. PAA alone had only weak or no effects. The effects of IAA must be considered as hormone-specific. The ATPase activity in the presence of 10^-4 M PAA was activated only by 2,4-dichlorophenoxyacetic acid (2,4-D), an active auxin analogue, but not by the inactive stereoisomers, 2,3-D and 3,5-D. Comparison with marker enzyme profiles suggested that part of the auxin-stimulated ATPase was localized on plasma membranes as well as other compartments. Thus, the auxin-stimulated ATPase may become a useful tool in the investigation of the mechanism of action of auxin.Abbrevations 2,4-D 2,4-dichlorophenoxyacetic acid - 2,3-D 2,3-dichlorophenoxyacetic acid - 3,5-D 3,5-dichlorophenoxyacetic acid - IAA indol-3-acetic acid - PAA phenylacetic acid - MES (2-(N-morpholino))-ethanesulfonic acid - EDTA ethylenediamine tetraacetic acid     

In vitro binding affinities of 4-chloro-, 2-methyl-, 4-methyl-, and 4-ethylindoleacetic acid to auxin-binding protein 1 (ABP1) correlate with their growth-stimulating activities

4-Chlorindole-3-acetic acid (4-CI-IAA), an endogenous auxin in certain plant species of Fabaceae, has a higher efficiency in stimulating cell elongation of grass coleoptiles compared with indole-3-acetic acid (IAA), particularly at low concentrations. However, some investigations reported a 1,000-fold discrepancy between growth stimulation and binding affinity of 4-CI-IAA to auxin-binding protein 1 (ABP1) from maize. Here we report binding data of 4-CI-IAA and three alkylated IAA derivatives using purified ABP1 in equilibrium dialysis. There is a clear correlation between the growth-promoting effects and the binding affinity to ABP1 of the different IAA analogues measured by competition of [³H]naphthalene-1-acetic acid binding. Our data are consistent with the hypothesis that ABP1 mediates auxin-induced cell elongation.Abbreviations ABP1 auxin-binding protein 1 - 4-CI-IAA 4-chloroindole-3-acetic acid - NAA naphthalene-1-acetic acid - ER endoplasmic reticulum - IAA indole-3-acetic acid - 2-Me-IAA 2-methylindole-3-acetic acid - 4-Me-IAA 4-methylindole-3-acetic acid - 4-Et-IAA 4-ethylindole-3-acetic acid - MES 4-morpholineethanesulfonic acid - PAA phenylacetic acid     

Regulation of auxin transport in pea (Pisum sativum L.) by phenylacetic acid: inhibition of polar auxin transport in intact plants and stem segments

The transport of [¹⁴C]phenylacetic acid (PAA) in intact plants and stem segments of light-grown pea (Pisum sativum L. cv. Alderman) plants was investigated and compared with the transport of [¹⁴C]indiol-3yl-acetic acid (IAA). Although PAA was readily taken up by apical tissues, unlike IAA it did not undergo long-distance transport in the stem. The absence of PAA export from the apex was shown not to be the consequence of its failure to be taken up or of its metabolism. Only a weak diffusive movement of PAA was observed in isolated stem segments which readily transported IAA. When [1-¹⁴C]PAA was applied to a mature foliage leaf in light, only 5.4% of the ¹⁴C recovered in ethanol extracts (89.6% of applied ¹⁴C) had been exported from the leaf after 6.0 h. When applied to the corresponding leaf, [¹⁴C]sucrose was readily exported (46.4% of the total recovered ethanol-soluble ¹⁴C after 6.0 h). [1-¹⁴C]phenylacetic acid applied to the root system was readily taken up but, after 5.0 h, 99.3% of the recovered ¹⁴C was still in the root system.When applied to the stem of intact plants (either in lanolin at 10 mg·g^-1, or as a 10^-4 M solution), unlabelled PAA blocked the transport through the stem of [1-¹⁴C]IAA applied to the apical bud, and caused IAA to accumulate in the PAA-treated region of the stem. Applications of PAA to the stem also inhibited the basipetal polar transport of [1-¹⁴C]IAA in isolated stem segments. These results are consistent with recent observations (C.F. Johnson and D.A. Morris, 1987, Planta 172, 400–407) that no carriers for PAA occur in the plasma membrane of the light-grown pea stem, but that PAA can inhibit the carrier-mediated efflux of IAA from cells. The possible functions of endogenous PAA are discussed and its is suggested that an important role of the compound may be to modulate the polar transport and-or accumulation by cells of IAA.Abbreviations IAA indol-3yl-acetic acid - NPA N-1-naphthylphthalamic acid - PAA phenylacetic acid - IIBA 2,3,5-triiodobenzoic acid     

Continuous measurement of initial curvature of maize coleoptiles induced by lateral auxin application

To analyze early effects of auxin application, an apparatus was developed which continuously and simultaneously registered the curvature of 10 individual maize (Zea mays L.) coleoptiles. Resolution was less than 5 m over a range of ±0.5 mm. The data were evaluated and plotted via paper tape and Hewlett-Packard-computer. Unilateral application of 3×10^-5 M indoleacetic acid (IAA) resulted in a transient inhibition of growth on the side of application for ca. 10 min (Phase I), followed by a strong stimulation (Phase II). The phytotoxin fusicoccin (FC) caused an immediate stimulation of elongation. The initial negative reaction of Phase I is auxin-specific. Only active auxins such as IAA and 1-naphtaleneacetic acid produced this initial inhibition; chemical analogs-inhibitory or neutral in long-term growth tests, e.g. phenylacetic acid-did not show any significant effects on Phase I. When the coleoptiles were symmetrically preloaded with different levels of auxin, only a large step-up of subsequent unilateral auxin application resulted in a negative phase I; a small step-up led to an immediate positive reaction. The results are discussed in context with the parallel kinetics for various other auxin-induced reactions of coleoptile cells which have already been published.Abbreviations FC fusicoccin - IAA indole-3-acetic acid - NAA -naphthaleneacetic acid - PAA phenylacetic acid     

The effects of antibiotics and their breakdown products on the in vitro growth of Antirrhinum majus

Summary The effects of various antibiotics on the development of hypocotyls of Antirrhinum majus in tissue culture have been studied. The penicillins, carbenicillin and penicillin G, have been shown to stimulate callus growth, have little impact on shoot production and may stimulate root formation. The cephalosporins, cephotaxime and cephalosporin, have no effect on callus production and reduce shoot and root formation. HPLC, GC and GC-MS analyses have shown that concentrations of carbenicillin and penicillin G, commonly used in plant tissue culture, break down to give physiologically active levels of the auxin phenylacetic acid. This offers a mechanism for the stimulation of growth caused by these two antibiotics.Abbreviations Amp ampicillin - 6APA 6 aminopenicillanic acid - BAP benzyl amino purine - Cb carbenicillin - Cb_n and Cb_o 1 month and 1 year old samples respectively of carbenicillin stored at 4°C as powder - Cph cephalothin - Cx cephotaxime - IAA indoleacetic acid - MS Murashige and Skoog medium - NOA -naphthoxyacetic acid - PAA phenylacetic acid - PenG benzyl penicillin - PMA phenylmalonic acid     

Effect of plant growth substances on the growth of axillary buds in cultured stem segments of Phaseolus vulgaris L.

The hormonal control of axillary bud growth was investigated in cultured stem segments of Phaseolus vulgaris L. When the stem explants were excised and implanted with their apical end in a solid nutrient medium, outgrowth of the axillary buds-located at the midline of the segment-was induced. However, if indoleacetic acid (IAA) or naphthaleneacetic acid (NAA) was included in the medium, bud growth was inhibited. The exposure of the apical end to IAA also caused bud abscission and prevented the appearance of new lateral buds.In contrast to apically inserted segments, those implanted in the control medium with their basal end showed much less bud growth. In these segments, the auxin added to the medium either had no effect or caused a slight stimulation of bud growth.The IAA transport inhibitor N-1-naphthylphthalamic acid (NPA) relieved bud growth inhibition by IAA. This suggests that the effect of IAA applied at the apical end requires the transport of IAA itself rather than a second factor. With the apical end of the segment inserted into the IAA-containing medium, simultaneous basal application of IAA relieved to some extent the inhibitory effect of the apical IAA treatment. These results, together with data presented in a related article [Lim R and Tamas I (1989) Plant Growth Regul 8: 151–164], show that the polarity of IAA transport is a critical factor in the control of axillary bud growth.Of the IAA conjugates tested for their effect on axillary bud growth, indoleacetyl alanine, indoleacetic acid ethyl ester, indoleacetyl-myo-inositol and indoleacetyl glucopyranose were strongly inhibitory when they were applied to the apical end of the stem explants. There was a modest reduction of growth by indoleacetyl glycine and indoleacetyl phenylalanine. Indoleacetyl aspartic acid and indoleglyoxylic acid had no effect.In addition to IAA and its conjugates, a number of other plant growth substances also affected axillary bud growth when applied to the apical end of stem segments. Myo-inositol caused some increase in the rate of growth, but it slightly enhanced the inhibitory effect of IAA when the two substances were added together. Gibberellic acid (GA₃) caused some stimulation of bud growth when the explants were from younger, rather than older plants. The presence of abscisic acid (ABA) in the medium had no effect on axillary bud growth. Both kinetin and zeatin caused some inhibition of axillary buds from younger plants but had the opposite effect on buds from older ones. Kinetin also enhanced the inhibitory effect of IAA when the two were applied together.In conclusion, axillary buds of cultured stem segments showed great sensitivity to auxins and certain other substances. Their growth responded to polarity effects and the interaction among different substances. Therefore, the use of cultured stem segments seems to offer a convenient, sensitive and versatile test system for the study of axillary bud growth regulation.     

Auxin-Promoted Transport of Metabolites in Stems of Phaseolus vulgaris L.: Auxin Dose-Response Curves and Effects of Inhibitors of Polar Auxin Transport

Transport of ¹⁴C-photosynthate in decapitated stems of Phaseolusvulgaris explants was dependent on the concentration of indole-3-aceticacid (IAA) applied to the cut surfaces of the stem stumps. Thephysiological age of the stem influenced the nature of the transportresponse to IAA with stems that had ceased elongation exhibitinga more pronounced response with a distinct optimum. Increasednutrient status of the explants had little influence on theshape of the IAA dose-response curve but increased, by two ordersof magnitude, the IAA concentration that elicited the optimalresponse. Applications of the inhibitor of polar auxin transport,1-(2-carboxyphenyl)-3-phenylpropane-1, 3-dione (CPD), affectedIAA-promoted transport of ¹⁴C-photosynthates. At sub-optimalIAA concentrations, CPD inhibited transport, whereas at supra-optimalIAA concentrations, ¹⁴C-photosynthate transport was marginallystimulated by CPD. Treatment with CPD resulted in a significantreduction in stem levels of [¹⁴C]IAA below the site of inhibitorapplication, while above this point, levels of [¹⁴C]1AA remainedunaltered. The divergent responses of auxin-promoted transportto CPD treatment are most consistent with a remote action ofIAA on photosynthate transport in the decapitated stems. Key words: Auxin, photosynthate, transport     

Inhibition of Lettuce Seed Germination and Root Elongation by Derivatives of Auxin and Reversal by Derivatives of Cycocel

Lettuce seed germination or lettuce root elongation after germination in water was inhibited by 5.7 × 10^-6M indoleacetic acid (IAA) and designated other IAA derivatives. These IAA-inhibited growth responses were reversed by 10^-5 to 10^-6M Cycocel, (2-chloroethyl) trimethylammonium chloride, which alone was without effect. Only those Cycocel analogs, which have previously been shown to be active as plant growth retardants, were effective in reversing IAA inhibition of germination or root elongation. These results are consistent with the concept that Cycocel at low concentrations acts as an auxin antagonist. However. Cycocel did not reverse the inhibitory effects from indole-3-propionic acid or indole-3-butyric acid.     

Auxin transport and the regulation of petiole abscission in cotton (Gossypium hirsutum L.): A comparison of indol-3yl-acetic acid and phenylacetic acid

Distal applications of indol-3yl-acetic acid (IAA) to debladed cotyledonary petioles of cotton (Gossypium hirsutum L.) seedlings greatly delayed petiole abscission, but similar applications of phenylacetic acid (PAA) slightly accelerated abscission compared with untreated controls. Both compounds prevented abscission for at least 91 h when applied directly to the abscission zone at the base of the petiole. The contrasting effects of distal IAA and PAA on abscission were correlated with their polar transport behaviour-[1-¹⁴C]IAA underwent typical polar (basipetal) transport through isolated 30 mm petiole segments, but only a weak diffusive movement of [1-¹⁴C]PAA occurred.Removal of the shoot tip substantially delayed abscission of subtending debladed cotyledonary petioles. The promotive effect of the shoot tip on petiole abscission could be replaced in decapitated shoots by applications of either IAA or PAA to the cut surface of the stem. Following the application of [1-¹⁴C]IAA or [1-¹⁴C]PAA to the cut surface of decapitated shoots, only IAA was transported basipetally through the stem. Proximal applications of either compound stimulated the acropetal transport of [¹⁴C]sucrose applied to a subtending intact cotyledonary leaf and caused label to accumulate at the shoot tip. However, PAA was considerably less active than IAA in this response.It is concluded that whilst the inhibition of petiole abscission by distal auxin is mediated by effects of auxin in cells of the abscission zone itself, the promotion of abscission by the shoot tip (or by proximal exogenous auxin) is a remote effect which does not require basipetal auxin transport to the abscission zone. Possible mechanisms to explain this indirect effect of proximal auxin on abscission are discussed.     

Gas chromatography-mass spectrometry evidence for several endogenous auxins in pea seedling organs

Qualitative analysis by gas chromatography-mass spectrometry (GC-MS) of the auxins present in the root, cotyledons and epicotyl of 3-dold etiolated pea (Pisum sativum L., cv. Alaska) seedlings has shown that all three organs contain phenylacetic acid (PAA), 3-indoleacetic acid (IAA) and 4-chloro-3-indoleacetic acid (4Cl-IAA). In addition, 3-indolepropionic acid (IPA) was present in the root and 3-indolebutyric acid (IBA) was detected in both root and epicotyl. Phenylacetic acid, IAA and IPA were measured quantitatively in the three organs by GC-MS-single ion monitoring, using deuterated internal standards. Levels of IAA were found to range from 13 to 115 pmol g^-1 FW, while amounts of PAA were considerably higher (347–451 pmol g^-1 FW) and the level of IPA was quite low (5 pmol g^-1 FW). On a molar basis the PAA:IAA ratio in the whole seedling was approx. 15:1.Abbreviations IAA 3-indoleacetic acid - 4Cl-IAA 4-chloro-3-indoleacetic acid - IBA 3-indolebutyric acid - IPA 3-indolepropionic acid - PAA phenylacetic acid - GC-MS gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - PFB pentafluorobenzyl ester - PFBBr pentafluorobenzyl bromide - SIM single-ion monitoring - TMSI trimethylsilyl ester     

Indole-3-butyric acid in Arabidopsis thaliana III. In vivo biosynthesis

Indole-3-butyric acid (IBA) was identified by HPLC and GC-MS as one of the reaction products after incubation of sterile cultures of Arabidopsis thaliana seedlings with labeled indole-3-acetic acid (IAA). This is the first demonstration of IBA biosynthesis in a dicotyledonous plant. After 1 h of incubation most of the IBA was found in the free form, while after longer periods of incubation most of it was detected in conjugated forms. Formation of IBA conjugates was inhibited by the addition of unlabeled IBA. The biosynthesis of IBA and its conjugates was followed throughout the development of the seedlings and at different pH values. All parts of the plant (isolated roots, leaves, shoots and flowers) were able to convert IAA to IBA to the same extent.IAA was more readily transported than IBA in mature Arabidopsis plants. Feeding of labeled phenylacetic acid (PAA) and -naphthylacetic acid (NAA) to Arabidopsis seedlings resulted in a new small peak which was hydrolyzed by 7N NaOH, but the formation of compounds with longer side chains (analogous to IBA) could not be detected.Abbreviations IAA indole-3-acetic acid - IBA indole-3-butyric acid - NAA -naphthylacetic acid - PAA phenylacetic acid     

Regulation of auxin transport in pea (Pisum sativum L.) by phenylacetic acid: effects on the components of transmembrane transport of indol-3yl-acetic acid

Phenylacetic acid (PAA), a naturally-occurring acidic plant growth substance, was readily taken up by pea (Pisum sativum L. cv. Alderman) stem segments from buffered external solutions by a pH-dependent, non-mediated diffusion. Net uptake from a 0.2 M solution at pH 4.5 proceeded at a constant rate for at least 60 min and, up to approx. 100 M, the rate of uptake was directly proportional to the external concentration of the compound. The net rate of uptake of PAA was not affected by the inclusion of indol-3yl-acetic acid (IAA) in the uptake medium (up to approx. 30 M) and, unlike the net uptake of IAA, was not stimulated by N-1-naphthylphthalamic acid (NPA) or 2,3,5-triiodobenzoic acid. At an external concentration of 0.2 M and pH 4.5, the net rate of uptake of PAA was about twice that of IAA. It was concluded that the uptake of PAA did not involve the participation of carriers and that PAA was not a transported substrate for the carriers involved in the uptake and polar transport of IAA. Nevertheless, the inclusion of 3–100 M unlabelled PAA in the external medium greatly stimulated the uptake by pea stem segments of [1-¹⁴C]IAA (external concentration 0.2 M). It was concluded that whilst PAA was not a transported substrate for the NPA-sensitive IAA efflux carrier, it interacted with this carrier to inhibit IAA efflux from cells. Over the concentration range 3–100 M, PAA progressively reduced the stimulatory effect of NPA on IAA uptake, indicating that PAA also inhibited carrier-mediated uptake of IAA. The consequences of these observations for the regulation of polar auxin transport are discussed.Abbreviations IAA indol-3yl-acetic acid - DMO 5,5-dimethyloxazolidine-2,4-dione - NPA N-1-naphthylphthalamic acid - PAA phenylacetic acid - TIBA 2,3,5-triiodobenzoic acid     

On the Physiological Effects of Phloridzin, Phloretin and some Related Substances upon Higher Plants

Some dihydrochalcones and related flavonoids were tested Cor their effects upon indoleacetic acid oxidase (mainly from pea roots) and upon oxidative phosphorylation in mitochondria from cucumber hypocotyls. Also their influence upon the growth of wheat roots and upon the absorption of ¹⁴C-labelled sugars and 2, 4-dichloro-phenoxyacetic acid in wlieat roots was studied. The dihydrochalcone glucoside phloridzin known for its strong inhibitory effects upon sugar transport in some animal tissues gives no corresponding specific effects in higher plants. Phloridzin and some related dihvdrochalcone glycosides as well as the structurally similar compounds naringenin and 2′, 4, 4′-trihydroxychalcone are potent stimulators of indoleacetic acid oxidase. Sieboldin, which compared to phloridzin has an additional hydroxyl group (giving an ortho-diphenolic substitution in the B-ring) is a strong inhibitor of indoleacetic acid oxidase (10^?6M gives about 50 per cent Inhibition). Phloridzin and sieboldin replace each other in different groups of species in the genus Malus. The aglycones phloretin, naringenin and 2′, 4, 4′-trihydroxychalcone stimulate wheat root growth (especially in the presence of auxins or growth-inhibilory sugars) and inhibit the absorption of sugars and of 2, 4-dichlorophenoxyacetic acid. They also inhibit oxidative phosphorylalion and give a distinct uncoupling effect. The glycosides are as a rule less active in all these respects and sieboldin is almost inactive. There is a rather good correlation between uncoupling of oxidative phosphorylation and effects upon root growth, whereas there is no agreement in the details between the effects upon root growth and upon IAA oxidase.     

Evidence supporting a model of voltage-dependent uptake of auxin into Cucurbita vesicles

The accumulation of [¹⁴C]indole-3-acetic acid (IAA), of [³H]tetra-phenyl phosphonium ion as a membrane potential probe, and of [¹⁴C]butyric acid as probe for pH gradients was studied with membrane vesicles from etiolated hypocotyls of Cucurbita pepo. Ion gradients (K⁺, H⁺) were applied in the presence and absence of specific ionophores e.g. valinomycin or carbonylcyanide m-chlorophenylhydrazone. In all cases tested, the accumulation of [¹⁴C]IAA equals neither potential probe nor pH-probe accumulation, but represents. an intermediate between the two. Auxin molecules seem to be taken up as positively charged ions and a pH gradient is required for accumulation. The uptake mechanism thus appears to be a specific, carrier-mediated cotransport of the anion of IAA and no less than two protons. The initial rates of auxin uptake by the saturable influx carrier, of permeation through the membrane, and of efflux by the phytotropin-affected efflux carrier were analysed.Abbreviations BA butyric acid - CCCP carbonylcyanid-3-chlorophenylhydrazone - CPD 2-carboxylphenyl-3-phenylpropan-1,3-dion - IAA indole-3-acetic acid - IAA anion of IAA - IAAH undissociated form of IAA - 2-NAA 2-naphthaleneacetic acid - NPA 1-N-naphthylphthalamic acid - TPP⁺ tetra-phenyl phosphonium ion     

A tracheid-differentiation factor from pine needles

Exogenous indol-3yl-acetic acid (IAA), alone and together with several cytokinins, was ineffective in promoting the complete differentiation into tracheids of cambial derivatives of Pinus contorta Dougl.; IAA alone promoted cambial cell division and primary-wall growth in cambial derivatives throughout the stem's length. In contrast, a single pair of needles on a stem cutting in light promoted neither cambial cell division nor primary-wall growth in cambial derivatives but did promote complete differentiation of cambial cells into tracheids; tracheids differentiated only near the junction of the foliated short shoot with the stem. Clear inter-and intracellular differences in the extent of tracheid differentiation occurred in response to a single needle pair and have suggested the hypothesis that a specific tracheid-differentiation factor regulates the differentiation of cells into proto-, meta-, or secondary-xylem tracheary elements through an interaction with IAA.Abbreviations IAA indol-3yl acetic acid - K kinetin - BAP 6-benzylaminopurine - Z zeatin - ZR zeatin riboside - FAA formalin: acetic acid: ethanol: water (10:5:50:35, by vol.)     

Facile Preparation of Deuterium-Labeled Standards of Indole-3-Acetic Acid (IAA) and Its Metabolites to Quantitatively Analyze the Disposition of Exogenous IAA in Arabidopsis thaliana

[2′,2′-²H₂]-indole-3-acetic acid ([2′,2′-²H₂]IAA) was prepared in an easy and efficient manner involving base-catalyzed hydrogen/deuterium exchange. 1-O-([2′,2′-²H₂]-indole-3-acetyl)-β-D-glucopyranose, [2′,2′-²H₂]-2-oxoindole-3-acetic acid, and 1-O-([2′,2′-²H₂]-2-oxoindole-3-acetyl)-β-D-glucopyranose were also successfully synthesized from deuterated IAA, and effectively utilized as internal standards in the quantitative analysis of IAA and its metabolites in Arabidopsis thaliana by using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). The use of this technique shows that these metabolites were accumulated in the roots of Arabidopsis seedlings. Dynamic changes in the metabolites of IAA were observed in response to exogenous IAA, revealing that each metabolic action was regulated differently to contribute to the IAA homeostasis in Arabidopsis.     

Applicability of the chemiosmotic polar diffusion theory to the transport of indol-3yl-acetic acid in the intact pea (Pisum sativum L.)

The transport of exogenous indol-3yl-acetic acid (IAA) from the apical tissues of intact, light-grown pea (Pisum sativum L. cv. Alderman) shoots exhibited properties identical to those associated with polar transport in isolated shoot segments. Transport in the stem of apically applied [1-¹⁴C]-or [5-³H]IAA occurred at velocities (approx. 8–15 mm·h^-1) characteristic of polar transport. Following pulse-labelling, IAA drained from distal tissues after passage of a pulse and the rate characteristics of a pulse were not affected by chases of unlabelled IAA. However, transport of [1-¹⁴C]IAA was inhibited through a localised region of the stem pretreated with a high concentration of unlabelled IAA or with the synthetic auxins 1-napthaleneacetic acid and 2,4-dichlorophenoxyacetic acid, and label accumulated in more distal tissues. Transport of [1-¹⁴C]IAA was also completely prevented through regions of the intact stem treated with N-1-naphthylphthalamic acid (NPA) and 2,3,5-triiodobenzoic acid.Export of IAA from the apical bud into the stem increased with total concentration of IAA applied (labelled+unlabelled) but approached saturation at high concentrations (834 mmol·m^-3). Transport velocity increased with concentration up to 83 mmol·m^-3 IAA but fell again with further increase in concentration.Stem segments (2 mm) cut from intact plants transporting apically applied [1-¹⁴C]IAA effluxed 93% of their initial radioactivity into buffer (pH 7.0) in 90 min. The half-time for efflux increased from 32.5 to 103.9 min when 3 mmol·m^-3 NPA was included in the efflux medium. Long (30 mm) stem sections cut from immediately below an apical bud 3.0 h after the apical application of [1-¹⁴C]IAA effluxed IAA when their basal ends, but not their apical ends, were immersed in buffer (pH 7.0). Addition of 3 mmol·m^-3 NPA to the external medium completely prevented this basal efflux.These results support the view that the slow long-distance transport of IAA from the intact shoot apex occurs by polar cell-to-cell transport and that it is mediated by the components of IAA transmembrane transport predicted by the chemiosmotic polar diffusion theory.Abbreviations IAA indol-3yl-acetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - NAA 1-naphthaleneacetic acid - NPA N-1-naphthylphthalamic acid - TIBA 2,3,5-triiodobenzoic acid     

Antifungal and sprout regulatory bioactivities of phenylacetic acid, indole-3-acetic acid, and tyrosol isolated from the potato dry rot suppressive bacterium Enterobacter cloacae S11:T:07

Enterobacter cloacae S11: T:07 (NRRL B-21050) is a promising biological control agent that has significantly reduced both fungal dry rot disease and sprouting in laboratory and pilot potato storages. The metabolites phenylacetic acid (PAA), indole-3-acetic acid (IAA), and tyrosol (TSL) were isolated from S11:T:07 liquid cultures provided with three different growth media. The bioactivities of these metabolites were investigated via thin-layer chromatography bioautography of antifungal activity, wounded potato assays of dry rot suppressiveness, and cored potato eye assays of sprout inhibition. Relative accumulations of PAA, IAA, and TSL in cultures were nutrient dependent. For the first time, IAA, TSL, and PAA were shown to have antifungal activity against the dry rot causative pathogen Gibberella pulicaris, and to suppress dry rot infection of wounded potatoes. Disease suppression was optimal when all three metabolites were applied in combination. Dosages of IAA that resulted in disease suppression also resulted in sprout inhibition. These results suggest the potential for designing culture production and formulation conditions to achieve a dual purpose biological control agent able to suppress both dry rot and sprouting of stored potatoes.     

Characterization of the growth and auxin physiology of roots of the tomato mutant,diageotropica

Roots of the tomato (Lycopersicon esculentum, Mill.) mutant diageotropica (dgt) exhibit an altered phenotype. These roots are agravitropic and lack lateral roots. Relative to wild-type (VFN8) roots, dgt roots are less sensitive to growth inhibition by exogenously applied IAA and auxin transport inhibitors (phytotropins), and the roots exhibit a reduction in maximal growth inhibition in response to ethylene. However, IAA transport through roots, binding of the phytotropin, tritiated naphthylphthalamic acid ([³H]NPA), to root microsomal membranes, NPA-sensitive IAA uptake by root segments, and uptake of [³H]NPA into root segments are all similar in mutant and wild-type roots. We speculate that the reduced sensitivity of dgt root growth to auxin-transport inhibitors and ethylene is an indirect result of the reduction in sensitivity to auxin in this single gene, recessive mutant. We conclude that dgt roots, like dgt shoots, exhibit abnormalities indicating they have a defect associated with or affecting a primary site of auxin perception or action.Abbreviations BCA bicinchoninic acid - IAA indole 3-acetic acid - dgt diageotropica - IC₅₀ concentration for 50% inhibition of growth - NPA N-1-naphthylphthalamic acid - SCB-1 semicarbazone 1 This research was supported by grants from Sandoz Agro, Inc. (G.K.M), the National Aeronautics and Space Administration (NASA) and the National Science Foundation (T.L.L), and NASA (D.L.R.).     

Stems of the <Emphasis Type="Italic">Arabidopsis pin1-1</Emphasis> mutant are not deficient in free indole-3-acetic acid

The pin1-1 mutant of Arabidopsis thaliana has been pivotal for studies on auxin transport and on the role of auxin in plant development. It was reported previously that when whole shoots were analysed, levels of the major auxin, indole-3-acetic acid (IAA) were dramatically reduced in the mutant, compared with the WT (Okada et al. 1991). The cloning of PIN1, however, provided evidence that this gene encodes a facilitator of auxin efflux, raising the question of how the pin1-1 mutation might reduce overall IAA levels as well as IAA transport. We therefore re-examined IAA levels in individual parts of pin1-1 and WT plants, focusing on inflorescence stems. Our data show that there is in fact no systemic IAA deficiency in the mutant. The previously reported difference between mutant and WT may have been due to the inclusion of reproductive structures in the WT harvest: we show here that the inflorescence itself contains high levels of IAA. We reconcile the normal IAA levels of pin1-1 inflorescence stems with their (previously-reported) reduced ability to transport IAA by presenting evidence that the auxin in mutant stems is not imported from their apical portion. Our data also indicate that levels of another auxin, indole-3-butyric acid (IBA), are very low in stems of the genotypes used in this study.