The effect of ethylene and auxin on cell wall extensibility of the Semi-aquatic fern,Regnellidium diphyllum

Ethylene and auxin both enhance cell elongation growth in the rachis of the frond of Regnellidium diphyllum. Measurements of the stress relaxation modulus of the walls of methanol-killed rachis segments show that both auxin and ethylene cause an increase in cell wall extensibility, that the effects are additive, and that they occur in the presence of hypertonic solutions of mannitol that preclude cell elongation. The results are taken as evidence for the operation of two separate mechanisms for cell wall loosening.Abbreviation IAA indol-3yl-acetic acid     

The positional differentiation of abscission zones during the development of leaves of Sambucus nigra and the response of the cells to auxin and ethylene

Summary Abscission in the leaf rachis of Sambucus nigra L. is preceded by a positional differentiation of zone cells that enlarge and separate in response to ethylene but not to auxin. These cells are absent from youngest leaves, and such leaves do not abscind even in ethylene; other cells of the immature rachii will enlarge in response to auxin. These two classes of target cells are always recognisable by their opposing responses to auxin and ethylene. Prior to separation zone cells exposed to ethylene show considerable activation of the cytoplasm, many polysomes, elongate endoplasmic reticulum and highly dilated dictyosomes with many associated vesicles. Treatment with auxin precludes these changes, and abscission is always retarded: high levels of ethylene must be added to overcome the auxin inhibition. The differentiation of zone cells and their ethylene-stimulated growth and activation are prerequisites for rachis abscission in Sambucus. Such cell development may be of general occurrence prior to organ abscission in plants.Abbreviation IAA indole-3yl-acetic acid     

The Auxins IAA and 4-Cl-IAA Differentially Modify Gibberellin Action via Ethylene Response in Developing Pea Fruit

This study explores the unique growth-regulatory roles of two naturally occurring auxins, indole-3-acetic acid (IAA) and 4-chloroindole-3-acetic acid (4-Cl-IAA), and their interactions with gibberellin (GA) during early pea (Pisum sativum L.) fruit development. We have previously shown that 4-Cl-IAA can replace the seed requirement in pea pericarp growth (length and fresh weight), whereas IAA had no effect or was inhibitory. When applied simultaneously, gibberellin (GA₃ or GA₁) and 4-Cl-IAA had a synergistic effect on pericarp growth. In the present study, we found that simultaneous application of IAA and GA₃ to deseeded pericarps inhibited GA₃-stimulated growth. The inhibitory effect of IAA on GA-stimulated growth was mimicked by treatment with ethephon (ethylene releasing agent), and the inhibitory effects of IAA and ethylene on GA-mediated growth were reversed by silver thiosulfate (STS), an ethylene action inhibitor. Although pretreatment with STS could retard senescence of IAA-treated pericarps, STS pretreatment did not lead to IAA-induced pericarp growth. Although 4-Cl-IAA stimulated growth whereas IAA was ineffective, both auxins induced similar levels of ethylene evolution. However, only 4-Cl-IAA-stimulated growth was insensitive to the effects of ethylene. Gibberellin treatment did not influence the amount of ethylene released from pericarps in the presence or absence of either auxin. We propose a growth regulatory role for 4-Cl-IAA through induction of GA biosynthesis and inhibition of ethylene action. Additionally, ethylene (IAA-induced or IAA-independent) may inhibit GA responses under physiological conditions that limit fruit growth.     

The positional differentiation of ethylene-responsive cells in rachis abscission zones in leaves of Sambucus nigra and their growth and ultrastructural changes at senescence and separation

Summary In leaves of S. nigra, fragmentation of the rachis follows the autumnal abscission of leaflets and the high levels of ethylene produced by the senescing blades. Fragmentation is accompanied by cell growth and ultrastructural changes in a zone of cells precisely differentiated at the separation zone. Studies with explants from the rachis show that those that contain an abscission zone increase in freshweight by as much as 50% before and during cell separation. Cell growth changes are induced by ethylene but not by auxin, and are restricted to explants that contain the separation zone cells. In ethylene, enlarging cells of the zone show cytoplasmic activation indicated by dilated dictyosomes, enhanced production of Golgi vesicles, elongated profiles of rough endoplasmic reticulum, a crenellated plasmalemma, and the apparent discharge and accumulation of cytoplasmic vesicles within the desmotubules of the branched plasmodesmata. Degradation of the middle lamella and cell wall matrix could be associated with the release of hydrolytic enzymes on the disruption of the vesicles. Although ultrastructural changes of a similar but limited nature occur in all cells of the rachis in response to ethylene, only those that are morphologically delimited as zone cells exhibit the growth and separation that leads to rachis fragmentation. It is proposed that abscission can occur only at the sites of the positional differentiation of these special ethylene-responsive target cells.Abbreviation IAA indole-acetic acid (auxin)     

Endogenous Auxin is Required but Supraoptimal for Rapid Growth of Rice (Oryza sativa L.) Seminal Roots, and Auxin Inhibition of Rice Seminal Root Growth is Not Caused by Ethylene

The dual effects of auxin and ethylene on rice seminal root growth were investigated in this study. Low concentrations of exogenous indole-3-acetic acid (IAA) had no effect on rice seminal root growth, whereas higher concentrations (≥0.003 μM) were inhibitory. In contrast, low concentrations of the auxin action inhibitor p-chlorophenoxyisobutyric acid (PCIB), ranging from 0.5 to 50 μM, promoted rice seminal root growth, whereas high concentrations of PCIB (≥500 μM) and the polar auxin transport inhibitor 2,3,5-triiodobenzoic acid (TIBA) inhibited rice seminal root growth. These results suggest that endogenous auxin is required but supraoptimal for rapid growth of rice seminal roots. In addition, although rice seminal root growth was inhibited by the exogenous ethylene-releasing compound ethephon or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) as well as exogenous IAA, the 50% inhibition of growth (I₅₀) caused by ethephon or ACC was weakened by certain concentrations of the ethylene action inhibitor Ag⁺ (0.016-0.4 μM). However, the I₅₀ caused by exogenous IAA was strengthened by Ag⁺ or the ethylene biosynthetic inhibitor aminoethoxyvinylglycine (AVG) and weakened by certain concentrations of PCIB (0.5-50 μM). Together, the inhibitory mechanisms of auxin and ethylene on rice seminal root growth should be different, and auxin inhibition of rice seminal root growth should not be caused by ethylene. Furthermore, our results indicated that a certain threshold level of ethylene was required to maintain rice seminal root growth, and that ethylene within the threshold may antagonize auxin inhibition of rice seminal root growth.     

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.).     

Cytokinin and Ethylene Affect Auxin Transport-Dependent Rhizogenesis in Hypocotyls of Common Ice Plant (<Emphasis Type="Italic">Mesembryanthemum crystallinum</Emphasis> L<Emphasis Type="Italic">.</Emphasis>)

Hypocotyl explants of Mesembryanthemum crystallinum regenerated roots when cultured vertically with either the apical end (AE) or basal end (BE) in media containing indole-3-acetic acid (IAA). IAA alone induced roots regularly from the basal end of the explants, either from the cut surface immersed in the medium or from the opposite side. The inhibitors of auxin efflux carriers, α-naphthylphthalamic acid (NPA) and 2,3,5-triiodobenzoic acid (TIBA), inhibited rhizogenesis only from AE-cultured explants, indicating the role of polar auxin transport in root regeneration in this system. Cytokinin (zeatin, kinetin, BAP) added to auxin-containing medium reduced rhizogenesis from the explants maintained with BE and AE and additionally changed the IAA-induced pattern of rooting in AE-cultured explants by favoring rooting from the apical end and middle part of the hypocotyl with its concomitant reduction from the basal end. The addition of kinetin did not influence the content of IAA in the explants maintained with AE, suggesting that the cytokinin effect on root patterning was not dependent on auxin biosynthesis. Kinetin, however, strongly enhanced ethylene production. The importance of ethylene in regulating PAT-dependent rhizogenesis was tested by using an ethylene antagonist AgNO₃, an inhibitor of ethylene synthesis aminoethoxyvinylglycine (AVG), and a precursor of ethylene, 1-aminocyclopropane-1-carboxylic acid (ACC). AgNO₃ applied together with IAA or with IAA and kinetin strongly reduced the production of ethylene, inhibited rhizogenesis, and induced nonregenerative callus from BE, suggesting the need for ethylene signaling to elicit the rhizogenic action of auxin. A reduction of rhizogenesis and decrease of ethylene biosynthesis was also caused by AVG. In addition, AVG at 10 μM reversed the effect of cytokinin on root patterning, resulting in roots emerging only from BE on the medium with IAA and kinetin. Conversely, ACC at 200 μM markedly enhanced the production of ethylene and partly mimicked the effect of cytokinin when applied with IAA alone, thus confirming that in cultured hypocotyls of ice plant, cytokinin affects IAA-induced rhizogenesis through an ethylene-dependent pathway.     

Cell specificity in auxin- and ethylene-induced ‘supergrowth’ in Riella helicophylla

Summary In gemmalings of Riella helicophylla, auxin and ethylene stimulate elongation growth, especially of pillar cells. When the two hormones are supplied simultaneously, the effects are additive, i.e. the result is supergrowth. In the cells of the meristem, elongation is enhanced by auxin, but not by ethylene when given alone. However, these cells also respond with supergrowth to a combined treatment with auxin and ethylene. The antiauxin p-chlorophenoxyisobutyric acid suppresses both the ethylene stimulation of cell growth and the additive supergrowth. The results support the concept that auxin pre-conditions the cells to the ethylene-dependent growth event. We suggest that the response elicited by the specific cell types could be related to differences in their level of endogenous auxin.Abbreviations IAA indole-3-acetic acid - PCIB p-chlorophenoxyisobutyric acid     

Auxin and Ethylene Regulation of Petiole Epinasty in Two Developmental Mutants of Tomato, diageotropica and Epinastic

The epinastic growth responses of petioles to auxin and ethylene were quantified in two developmental mutants of tomato (Lycopersicon esculentum Mill.). In the wild type parent line, cultivar VFN8, the epinastic response of excised petiole sections was approximately log-linear between 0.1 and 100 micromolar indole-3-acetic acid (IAA) and 2,4-dichlorophenoxyacetic acid (2,4-D) concentrations, with a greater response to 2,4-D at any concentration. When ethylene synthesis was inhibited by aminoethoxyvinylglycine (AVG), epinasty was no longer induced by auxin, but could be restored by the addition of ethylene gas. In the auxin-insensitive mutant, diageotropica (dgt), no epinastic response to IAA was observed at IAA concentrations that effectively induced epinasty in VFN8. In the absence of added IAA, epinastic growth of dgt petioles in 1.3 microliters per liter exogenous ethylene gas was more than double that of VFN8 petioles. IAA had little additional effect in dgt, but promoted epinasty in VFN8. These results confirm that tomato petiole cells respond directly to ethylene and make it unlikely that the differential growth responsible for epinasty results from lateral auxin redistribution. The second mutant, Epinastic (Epi), exhibits constitutively epinasty, cortical swelling, and root branching symptomatic of possible alternation in auxin or ethylene regulation of growth. Only minor quantitative differences were observed between the epinastic responses to auxin and ethylene of VFN8 and Epi. However, in contrast to VFN8, when ethylene synthesis or action was inhibited in Epi, auxin still induced 40 to 50% of the epinastic response observed in the absence of inhibitors. This indicates that the target cells for epinastic growth in Epi are qualitatively different from those of VFN8, having gained the ability to grow differentially in response to auxin alone. The dgt and Epi mutants provide useful systems in which to study the genetic determination of target cell specificity for hormone action.     

Characterization of 1-aminocyclopropane-1-carboxylate (ACC) deaminase containing <Emphasis Type="Italic">Methylobacterium oryzae</Emphasis> and interactions with auxins and ACC regulation of ethylene in canola (<Emphasis Type="Italic">Brassica campestris</Emphasis>)

The possible interaction of the plant hormones auxin and ethylene and the role of 1-aminocyclopropane-1-carboxylate (ACC) deaminase containing bacteria on ethylene production in canola (Brassica campestris) in the presence of inhibitory concentrations of growth regulators were investigated. The effects of auxin (indole-3-acetic acid and 2,4-dichlorophenoxy acetic acid), auxin transport inhibitor 2-(p-chlorophenoxy)-2-methylpropionic acid, ethylene precursor 1-aminocyclopropane-1-carboxylate and ethylene synthesis inhibitor l-α-(2-aminoethoxyvinyl)glycine hydrochloride on root elongation were concentration dependent. Exogenous addition of growth regulators influences the enzyme activities of ethylene production and we have presented here evidences that support the hypothesis that inhibitory effects of auxin on root elongation are independent of ethylene. Additionally, we have proved that inoculation of ACC deaminase containing Methylobacterium oryzae sequester ACC exuded from roots and hydrolyze them lowering the concentration of ACC in root exudates. However, the inhibitory actions of exogenous additions of auxins could not be ameliorated by bacterial inoculation that reduces ethylene concentration in canola seedlings.     

The outer epidermis of Avena and maize coleoptiles is not a unique target for auxin in elongation growth

A controversy exists as to whether or not the outer epidermis in coleoptiles is a unique target for auxin in elongation growth. The following evidence indicates that the outer epidermis is not the only auxin-responsive cell layer in either Avena sativa L. or Zea mays L. coleoptiles. Coleoptile sections from which the epidermis has been removed by peeling elongate in response to auxin. The magnitude of the response is similar to that of intact sections provided the incubation solution contains both auxin and sucrose. The amount of elongation is independent of the amount of epidermis removed. Sections of oat coleoptiles from which the epidermis has been removed from one side are nearly straight after 22 h in auxin and sucrose, despite extensive growth of the sections. These data indicate that the outer epidermis is not a unique target for auxin in elongation growth, at least in Avena and maize coleoptiles.Abbreviations IAA indole-3-acetic acid - PCIB p-chlorophenoxyiso-butyric This research was supported by grants from the National Aeronautics and Space Administration and from the U.S. Department of Energy. The help of S. Ann Dreyer is gratefully acknowledged.     

Effect of ethylene on auxin transport and metabolism in midrib sections in relation to leaf abscission of woody plants

Abstract The relationship between ethylene-induced leaf abscission and ethylene-induced inhibition of auxin transport in midrib sections of the leaf blade of Citrus sinensis L. Osbeck, Populus deltoides Bart, and Eucalyptus camaldulensis Dehn. was studied. These species differed greatly in their abscission response to ethylene. The kinetic trend of abscission resembled that of the inhibition of auxin transport in all three species. It is suggested that one of the main actions of ethylene in the leaf blade is to inhibit auxin transport in the veinal tissues, thus reducing the amount of auxin transported from the leaf blade to the abscission zone. Ethylene inhibited transport of both IAA (indole-3-acetic acid) and NAA (α-naphthaleneacetic acid) in the midrib sections. However, while ethylene enhanced the conjugation of IAA with aspartic acid and glucose in the apical (absorbing) segment of the midrib sections, it had little effect on the conjugation of NAA. The data indicate that auxin destruction through conjugation does not play a major role in the inhibition of auxin transport by ethylene.     

A dominant mutation in Arabidopsis confers resistance to auxin, ethylene and abscisic acid

Summary We have screened a large population of M₂ seeds ofArabidopsis thaliana for plants which are resistant to exogenously applied indole-acetic acid (IAA). One of the resistant lines identified in this screen carries a dominant mutation which we have namedaxr2. Linkage analysis indicates that theaxr2 gene lies on chromosome 3. Plants carrying theaxr2 mutation are severe dwarfs and display defects in growth orientation of both the shoot and root suggesting that the mutation affects some aspect of gravitropic growth. In addition, the roots ofaxr2 plants lack root hairs. Growth inhibition experiments indicate that the roots ofaxr2 plants are resistant to ethylene and abscisic acid as well as auxin.     

Restart of Lignification in Micropropagated Walnut Shoots Coincides with Rooting Induction

The lignin content of walnut shoots did not change during in vitro shoot multiplication. Lignin content started to increase as soon as shoots were passed to a rooting medium with auxin. Exogenous auxin (applied for rooting) caused a transient elevation of the endogenous free indoleacetic acid (IAA) content with a simultaneous decrease of peroxidase activity. These events typically marked the completion of the rooting inductive phase (before any visible histological event, that is before the cell divisions beginning the rooting initiation phase). This meant that either the given exogenous auxin or the endogenous IAA has served as signal for the stimulation of lignification. Continued increase of lignification in the shoots required completion of root formation; this increase indeed was slown down when root emergence did not occur. It was further shown that lignification varied conversely to the content of the soluble phenol content, itself apparently being related to the activity of phenylalanine ammonia-lyase activity. This revised version was published online in July 2006 with corrections to the Cover Date.     

Inhibitory action of auxin on root elongation not mediated by ethylene

The inhibitory effects of indole-3-acetic acid (IAA) and 1-aminocyclopropane-1-carboxylic acid (ACC) on elongation growth of pea (Pisum sativum L.) seedling roots were investigated in relation to the effects of these compounds on ethylene production by the root tips. When added to the growth solution both compounds caused a progressively increasing inhibition of growth within the concentration range of 0.01 to 1 micromolar. However, only ACC increased ethylene production in root tips excised from the treated seedlings after 24 hours. High auxin concentrations caused a transitory increase of ethylene production during a few hours in the beginning of the treatment period, but even in 1 micromolar IAA this increase was too low to have any appreciable effect on growth. ACC, but not IAA, caused growth curvatures, typical of ethylene treatment, in the root tips. IAA caused conspicuous swelling of the root tips while ACC did not. Cobalt and silver ions reversed the growth inhibitory effects induced by ACC but did not counteract the inhibition of elongation or swelling caused by IAA. The growth effects caused by the ACC treatments were obviously due to ethylene production. We found no evidence to indicate that the growth inhibition or swelling caused by IAA is mediated by ethylene. It is concluded that the inhibitory action of IAA on root growth is caused by this auxin per se.     

Control of the glutathione S-transferase and mas1' promoter-driven GUS activity in auxin heterotrophic and autotrophic tobacco calli by exogenous 2,4-d-induced ethylene

Auxin autotrophic and heterotrophic lines of tobacco calli may differ not only in their indoleacetic acid (IAA) synthetizing abilities and sensitivities to exogenous auxins, but also in their gene expression patterns. Auxin autotrophic callus tissues from the leaf protoplasts of transgenic Nicotiana tabacum SR1 plants involving mas1′::GUS gene fusion were generated and the growth of cultures was compared with that of the heterotrophic lines of the same transgenic tissues on MS medium containing different concentrations of IAA or 2,4‐d . The mas1′::GUS gene fusion expression was investigated, together with the glutathione S‐transferase activities (GST, EC 2.5.1.18) in auxin autotrophic and heterotrophic tobacco calli. Both the mas1′ promoter and GST gene promoters contain ocs or ocs‐like elements, responsible for both auxin and ethylene/wound inducibility. The mas1′ promoter exhibited a much higher expression activity in the heterotrophic cultures growing on IAA than in the autotrophic ones, but in contrast with the natural auxin, the mas1′::GUS activity decreased at elevated 2,4‐d concentrations in the heterotrophic tissues and increased with increasing 2,4‐d concentrations in the autotrophic lines. The induction of GST activity by different exogenous auxin concentrations was much higher in the autotrophic lines, especially in the case of 2,4‐d . Higher concentrations of external 2,4‐d resulted in increased ethylene production, which displayed different kinetics in the two types of calli. The ethylene‐inducing 2,4‐d concentrations increased the growth of the heterotrophic, but decreased that of the autotrophic lines. Blocking the ethylene receptors and hence the signal perception by 2,5‐norbornadiene (NBD) in the heterotrophic tissues increased the 2,4‐d ‐induced mas1′ promoter and GST activities, suggesting that the gaseous hormone counteracted the auxin response pathway. This was not found in the autotrophic tissues, where NBD decreased the mas1′‐driven GUS activity. The GST activities were slightly decreased, or almost independent of the action of ethylene. It is suggested that the cross‐talk between the auxin‐ and ethylene‐induced signal transduction pathways may differ in the auxin autotrophic and heterotrophic lines.     

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.     

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.     

An auxin-responsive 1-aminocyclopropane-1-carboxylate synthase is responsible for differential ethylene production in gravistimulated<Emphasis Type="Italic"> Antirrhinum majus</Emphasis> L. flower stems

The regulation of gravistimulation-induced ethylene production and its role in gravitropic bending was studied in Antirrhinum majus L. cut flower stems. Gravistimulation increased ethylene production in both lower and upper halves of the stems with much higher levels observed in the lower half. Expression patterns of three different 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS) genes, an ACC oxidase (ACO) and an ethylene receptor (ETR/ERS homolog) gene were studied in the bending zone of gravistimulated stems and in excised stem sections following treatment with different chemicals. One of the ACS genes (Am-ACS3) was abundantly expressed in the bending zone cortex at the lower side of the stems within 2 h of gravistimulation. Am-ACS3 was not expressed in vertical stems or in other parts of (gravistimulated) stems, leaves or flowers. Am-ACS3 was strongly induced by indole-3-acetic acid (IAA) but not responsive to ethylene. The Am-ACS3 expression pattern strongly suggests that Am-ACS3 is responsible for the observed differential ethylene production in gravistimulated stems; its responsiveness to IAA suggests that Am-ACS3 expression reflects changes in auxin signalling. Am-ACS1 also showed increased expression in gravistimulated and IAA-treated stems although to a much lesser extent than Am-ACS3. In contrast to Am-ACS3, Am-ACS1 was also expressed in non-bending regions of vertical and gravistimulated stems and in leaves, and Am-ACS1 expression was not confined to the lower side cortex but evenly distributed over the diameter of the stem. Am-ACO and Am-ETR/ERS expression was increased in both the lower and upper halves of gravistimulated stems. Expression of both Am-ACO and Am-ETR/ERS was responsive to ethylene, suggesting regulation by IAA-dependent differential ethylene production. Am-ACO expression and in vivo ACO activity, in addition, were induced by IAA, independent of the IAA-induced ethylene. IAA-induced growth of vertical stem sections and bending of gravistimulated flowering stems were little affected by ethylene or 1-methylcyclopropene treatments, indicating that the differential ethylene production plays no pivotal role in the kinetics of gravitropic bending.