Abscisic acid antagonizes ethylene-induced hyponastic growth in Arabidopsis

Ethylene induces enhanced differential growth in petioles of Arabidopsis (Arabidopsis thaliana), resulting in an upward movement of the leaf blades (hyponastic growth). The amplitude of this effect differs between accessions, with Columbia-0 (Col-0) showing a large response, while in Landsberg erecta (Ler), hyponastic growth is minimal. Abscisic acid (ABA) was found to act as an inhibitory factor of this response in both accessions, but the relationship between ethylene and ABA differed between the two; the ability of ABA to inhibit ethylene-induced hyponasty was significantly more pronounced in Col-0. Mutations in ABI1 or ABI3 induced a strong ethylene-regulated hyponastic growth in the less responsive accession Ler, while the response was abolished in the ABA-hypersensitive era1 in Col-0. Modifications in ABA levels altered petiole angles in the absence of applied ethylene, indicating that ABA influences petiole angles also independently from ethylene. A model is proposed whereby the negative effect of ABA on hyponastic growth is overcome by ethylene in Col-0 but not in Ler. However, when ABA signaling is artificially released in Ler, this regulatory mechanism is bypassed, resulting in a strong hyponastic response in this accession.     

A kinetic analysis of hyponastic growth and petiole elongation upon ethylene exposure in Rumex palustris

Background and Aims

Complete submergence is an important stress factor for many terrestrial plants, and a limited number of species have evolved mechanisms to deal with these conditions. Rumex palustris is one such species and manages to outgrow the water, and thus restore contact with the atmosphere, through upward leaf growth (hyponasty) followed by strongly enhanced petiole elongation. These responses are initiated by the gaseous plant hormone ethylene, which accumulates inside plants due to physical entrapment. This study aimed to investigate the kinetics of ethylene-induced leaf hyponasty and petiole elongation.

Methods

Leaf hyponasty and petiole elongation was studied using a computerized digital camera set-up followed by image analyses. Linear variable displacement transducers were used for fine resolution monitoring and measurement of petiole growth rates.

Key Results

We show that submergence-induced hyponastic growth and petiole elongation in R. palustris can be mimicked by exposing plants to ethylene. The petiole elongation response to ethylene is shown to depend on the initial angle of the petiole. When petiole angles were artificially kept at 0°, rather than the natural angle of 35°, ethylene could not induce enhanced petiole elongation. This is very similar to submergence studies and confirms the idea that there are endogenous, angle-dependent signals that influence the petiole elongation response to ethylene.

Conclusions

Our data suggest that submergence and ethylene-induced hyponastic growth and enhanced petiole elongation responses in R. palustris are largely similar. However, there are some differences that may relate to the complexity of the submergence treatment as compared with an ethylene treatment.     

Ethylene-induced hyponastic growth in Arabidopsis thaliana is controlled by ERECTA

Plants can respond quickly and profoundly to detrimental changes in their environment. For example, Arabidopsis thaliana can induce an upward leaf movement response through differential petiole growth (hyponastic growth) to outgrow complete submergence. This response is induced by accumulation of the phytohormone ethylene in the plant. Currently, only limited information is available on how this response is molecularly controlled. In this study, we utilized quantitative trait loci (QTL) analysis of natural genetic variation among Arabidopsis accessions to isolate novel factors controlling constitutive petiole angles and ethylene-induced hyponastic growth. Analysis of mutants in various backgrounds and complementation analysis of naturally occurring mutant accessions provided evidence that the leucin-rich repeat receptor-like Ser/Thr kinase gene, ERECTA , controls ethylene-induced hyponastic growth. Moreover, ERECTA controls leaf positioning in the absence of ethylene treatment. Our data demonstrate that this is not due to altered ethylene production or sensitivity.     

The roles of ethylene, auxin, abscisic acid, and gibberellin in the hyponastic growth of submerged Rumex palustris petioles

Rumex palustris responds to complete submergence with upward movement of the younger petioles. This so-called hyponastic response, in combination with stimulated petiole elongation, brings the leaf blade above the water surface and restores contact with the atmosphere. We made a detailed study of this differential growth process, encompassing the complete range of the known signal transduction pathway: from the cellular localization of differential growth, to the hormonal regulation, and the possible involvement of a cell wall loosening protein (expansin) as a downstream target. We show that hyponastic growth is caused by differential cell elongation across the petiole base, with cells on the abaxial (lower) surface elongating faster than cells on the adaxial (upper) surface. Pharmacological studies and endogenous hormone measurements revealed that ethylene, auxin, abscisic acid (ABA), and gibberellin regulate different and sometimes overlapping stages of hyponastic growth. Initiation of hyponastic growth and (maintenance of) the maximum petiole angle are regulated by ethylene, ABA, and auxin, whereas the speed of the response is influenced by ethylene, ABA, and gibberellin. We found that a submergence-induced differential redistribution of endogenous indole-3-acetic acid in the petiole base could play a role in maintenance of the response, but not in the onset of hyponastic growth. Since submergence does not induce a differential expression of expansins across the petiole base, it is unlikely that this cell wall loosening protein is the downstream target for the hormones that regulate the differential cell elongation leading to submergence-induced hyponastic growth in R. palustris.     

Modulation of ethylene- and heat-controlled hyponastic leaf movement in Arabidopsis thaliana by the plant defence hormones jasmonate and salicylate

Upward leaf movement (hyponastic growth) is adopted by several plant species including Arabidopsis thaliana, as a mechanism to escape adverse growth conditions. Among the signals that trigger hyponastic growth are, the gaseous hormone ethylene, low light intensities, and supra-optimal temperatures (heat). Recent studies indicated that the defence-related phytohormones jasmonic acid (JA) and salicylic acid (SA) synthesized by the plant upon biotic infestation repress low light-induced hyponastic growth. The hyponastic growth response induced by high temperature (heat) treatment and upon application of the gaseous hormone ethylene is highly similar to the response induced by low light. To test if these environmental signals induce hyponastic growth via parallel pathways or converge downstream, we studied here the roles of Methyl-JA (MeJA) and SA on ethylene- and heat-induced hyponastic growth. For this, we used a time-lapse camera setup. Our study includes pharmacological application of MeJA and SA and biological infestation using the JA-inducing caterpillar Pieris rapae as well as mutants lacking JA or SA signalling components. The data demonstrate that MeJA is a positive, and SA, a negative regulator of ethylene-induced hyponastic growth and that both hormones repress the response to heat. Taking previous studies into account, we conclude that SA is the first among many tested components which is repressing hyponastic growth under all tested inductive environmental stimuli. However, since MeJA is a positive regulator of ethylene-induced hyponastic growth and is inhibiting low light- and heat-induced leaf movement, we conclude that defence hormones control hyponastic growth by affecting stimulus-specific signalling pathways.     

Ethylene-induced differential petiole growth in Arabidopsis thaliana involves local microtubule reorientation and cell expansion

? Hyponastic growth is an upward petiole movement induced by plants in response to various external stimuli. It is caused by unequal growth rates between adaxial and abaxial sides of the petiole, which bring rosette leaves to a more vertical position. The volatile hormone ethylene is a key regulator inducing hyponasty in Arabidopsis thaliana. Here, we studied whether ethylene-mediated hyponasty occurs through local stimulation of cell expansion and whether this involves the reorientation of cortical microtubules (CMTs). ? To study cell size differences between the two sides of a petiole in ethylene and control conditions, we analyzed epidermal imprints. We studied the involvement of CMT orientation in epidermal cells using the tubulin marker line as well as genetic and pharmacological means of CMT manipulation. ? Our results demonstrate that ethylene induces cell expansion at the abaxial side of the- petiole and that this can account for the observed differential growth. At the abaxial side, ethylene induces CMT reorientation from longitudinal to transverse, whereas, at the adaxial side, it has an opposite effect. The inhibition of CMTs disturbed ethylene-induced hyponastic growth. ? This work provides evidence that ethylene stimulates cell expansion in a tissue-specific manner and that it is associated with tissue-specific changes in the arrangement of CMTs along the petiole.     

New perspectives in flooding research: the use of shade avoidance and Arabidopsis thaliana

BACKGROUND: Complete submergence of Rumex palustris leads to hyponastic (upward) petiole growth followed by enhanced petiole elongation. Previous pharmacological experiments have provided insights into the signal transduction pathway leading to this combined 'escape' response. It will, however, be difficult to gain further knowledge using these methods. Consequently, new approaches are required. SCOPE: Here we propose that different environmental signals resulting in similar phenotypes can help to understand better the submergence response. In this review, we show that both ethylene and shade induce similar growth responses in R. palustris and Arabidopsis thaliana. We illustrate how this can be exploited to unravel novel signalling components in submergence-induced elongation growth. Furthermore, we illustrate the potential of arabidopsis as a useful model in submergence research based on similarities with submergence-tolerant species such as R. palustris and the molecular opportunities it presents. This is illustrated by examples of current work exploring this concept. CONCLUSIONS: Incorporating different model systems, such as arabidopsis and shade avoidance, into submergence research can be expected to create powerful tools to unravel signal transduction routes determining submergence tolerance.     

Auxin perception and polar auxin transport are not always a prerequisite for differential growth

Using time-lapse photography, we studied the response kinetics of low light intensity-induced upward leaf-movement, called hyponastic growth, in Arabidopsis thaliana. This response is one of the traits of shade avoidance and directs plant organs to more favorable light conditions. Based on mutant- and pharmacological data we demonstrated that among other factors, functional auxin perception and polar auxin transport (PAT) are required for the amplitude of hyponastic growth and for maintenance of the high leaf angle, upon low light treatment. Here, we present additional data suggesting that auxin and PAT antagonize the hyponastic growth response induced by ethylene treatment. We conclude that ethylene- and low light-induced hyponastic growth occurs at least partly via separate signaling routes, despite their strong similarities in response kinetics.Key words: hyponastic growth, petiole, Arabidopsis, ethylene, low light, auxin, polar auxin transport, differential growthUpward leaf movement (hyponastic growth) is a trait of several plant species to escape from growth-limiting conditions.^1,2 Interestingly, Arabidopsis thaliana induces a marked hyponastic growth response triggered by various environmental stimuli, including complete submergence, high temperature, canopy shade and spectral neutral low light intensities (Fig. 1).^3–6 The paper of Millenaar et al. in the New Phytologist 2009,⁷ provides a detailed analysis of low light intensity-induced hyponastic growth and components of the signal transduction are characterized using time-lapse photography. Low light intensity-induced hyponastic growth is a component of the so-called shade avoidance syndrome. Light-spectrum manipulations and mutant analyses indicated that predominantly the blue light wavelength region affects petiole movement and fast induction of hyponastic growth to low light conditions involves the photoreceptor proteins Cryptochrome 1 (Cry1), Cry2, Phytochrome-A (PhyA) and PhyB. Moreover, we show that also photosynthesis-derived signals can induce differential growth.Open in a separate windowFigure 1Typical hyponastic growth phenotype of Arabidopsis thaliana. Side view of Columbia-0 plants treated 10 h with ethylene (5 µl l⁻¹) or low light (20 µmol m⁻² s⁻¹). Plants in control light conditions were in 200 µmol m⁻² s⁻¹. Both stimuli induce a clear leaf inclination (hyponasty) relative to the horizontal by differential growth of the petioles. Plants kept in control conditions only show modest diurnal leaf movement and leaf angles gradually decline over time due to maturation of the leaves. Note that the paint droplets were applied to facilitate quantitative measurement of leaf angle kinetics in a time-lapse camera setup.⁷The hyponastic growth response to low light intensity was not affected in several ethylene-insensitive mutant lines. Moreover, low light did not affect expression of ethylene inducible marker genes nor differences in ethylene release were noted. Therefore, we concluded that low light-induced hyponastic growth is independent of ethylene signaling. This is perhaps surprising, because ethylene is the main trigger of hyponastic growth induced by complete submergence in several species. Interestingly, both ethylene and low light can induce hyponastic growth in Arabidopsis with similar kinetics.³We showed that plants mutant in auxin perception components (transport inhibitor response1 (tir1) and tir1 afb1 afb2 afb3 quadruple, containing additional mutant alleles of TIR1 homologous F-box proteins) and plants mutant in (polar) auxin transport (tir3-1, pin-formed3 (pin3) and pin7) components had a lower hyponastic growth amplitude in low light conditions.⁷ Moreover, these mutants were less able to maintain the high leaf angles after the response maximum. Both characteristics were also noted in plants pre-treated with the polar auxin transport (PAT) inhibitor 2,3,5-triiodobenzoic acid (TIBA). We therefore concluded that auxin perception and PAT are involved in the regulation of low light-induced hyponastic growth.⁷ Interestingly, we observed that TIBA pretreatment did not inhibit ethylene-induced hyponastic growth. In fact, the response upon ethylene treatment was even modestly enhanced. In agreement with this observation, we show here that the above mentioned auxin perception and PAT mutants also showed a slightly enhanced hyponastic growth response upon ethylene treatment (Fig. 2).Open in a separate windowFigure 2Auxin involvement in ethylene induced hyponasty. Effect of exposure to ethylene (5 µl l⁻¹) on the kinetics of hyponastic petiole growth (A) in Arabidopsis thaliana Columbia-0 plants treated with 50 µm TIBa (open circles) or a mock treatment (line) adapted from Supporting Information Figure S3 of Millenaar et al. (2009)⁷ and (B–F) in Arabidopsis auxin signaling and polar auxin transport mutants (closed circles), compared to the wild type response to low light (lines). Petiole angles are pair wise subtracted, which corrects for diurnal petiole movement in control conditions. For details on this procedure, growth conditions, treatments, data acquirement and analysis see.^7,13 Error bars represent standard errors; n ≥ 12. mutants were obtained from the Nottingham Arabidopsis Stock Center (accession numbers are shown between brackets) or from the authors describing the lines. tir1-1 (n3798,¹⁴), tir1-1 afb1-1 afb2-1 (in a mixed Columbia/Wassilewskija background),¹⁵ tir3-1,¹⁴ pin3-4 (n9363,¹⁶) and pin7-1 (n9365,¹⁰).Despite that auxin and PAT are required for many differential growth responses such as phototropism and gravitropism,^8,11 these data indicate that auxin perception and PAT are not obligatory for ethylene-induced hyponasty in Arabidopsis per se. In fact, one might even conclude that auxin and PAT antagonizes ethylene-induced hyponasty. These results are partly in agreement with observations on the wetland species Rumex palustris, were pretreatment with the auxin-efflux carrier 1-naphthylphthalamic acid (NPA) resulted in doubling of the lag-phase for hyponastic growth under water, but hardly affected the amplitude of the response.¹²Together, this indicates that auxin is not always a prerequisite for differential growth responses. Based on the apparent contrasting effects of auxin perception and PAT in low light- and ethylene-induced hyponastic growth, we conclude that ethylene and low light induce hyponastic growth, at least partly, via separate signaling routes.     

Submergence research using Rumex palustris as a model; looking back and going forward

Flooding is a phenomenon that destroys many crops worldwide. During evolution several plant species evolved specialized mechanisms to survive short- or long-term waterlogging and even complete submergence. One of the plant species that evolved such a mechanism is Rumex palustris. When flooded, this plant species is capable to elongate its petioles to reach the surface of the water. Thereby it restores normal gas exchange which leads to a better survival rate. Enhanced levels of ethylene, due to physical entrapment, is the key signal for the plant that its environment has changed from air to water. Subsequently, a signal transduction cascade involving at least four (classical) plant hormones, ethylene, auxin, abscisic acid, and gibberellic acid, is activated. This results in hyponastic growth of the leaves accompanied by a strongly enhanced elongation rate of the petioles enabling them to reach the surface. Other factors, among them cell wall loosening enzymes have been shown to play a role as well.     

Ethylene Promotes Elongation Growth and Auxin Promotes Radial Growth in Ranunculus sceleratus Petioles

Submergence induces elongation in the petioles of Ranunculus sceleratus L., after a rise in endogenous ethylene levels in the tissue. Petioles of isolated leaves also elongate 100% in 24 hours when treated with ethylene gas, without a change in the radius. Application of silver thiosulfate, aminoethoxyvinylglycine (AVG), abscisic acid (ABA), or methyl jasmonate inhibits this elongation response. Gibberellic acid treatment promotes ethylene-induced elongation, without an effect on the radius. Indoelastic acid (IAA) induces radial growth in the petioles, irrespective of the presence or absence of added ethylene. High concentrations of IAA will also induce elongation growth, but this is largely due to auxin-induced ethylene synthesis; treatment with silver thiosulfate, AVG, ABA, or methyl jasmonate inhibit this auxin-promoted elongation growth. However, the radial growth induced by IAA is not affected by gibberellic acid, and not specifically inhibited by ABA, methyl jasmonate, silver thiosulfate, or AVG. These results support the idea that petiole cell elongation during “accommodation growth” can be separated from radial expansion. The radial expansion may well be regulated by IAA. However, effects of high levels of IAA are probably anomalous, since they do not mimic normal developmental patterns.     

ERECTA controls low light intensity-induced differential petiole growth independent of phytochrome B and cryptochrome 2 action in Arabidopsis thaliana

Plants can respond quickly and profoundly to changes in their environment. Several species, including Arabidopsis thaliana, are capable of differential petiole growth driven upward leaf movement (hyponastic growth) to escape from detrimental environmental conditions. Recently, we demonstrated that the leucine-rich repeat receptor-like Ser/Thr kinase gene ERECTA, explains a major effect Quantitative Trait Locus (QTL) for ethylene-induced hyponastic growth in Arabidopsis. Here, we demonstrate that ERECTA controls the hyponastic growth response to low light intensity treatment in a genetic background dependent manner. Moreover, we show that ERECTA affects low light-induced hyponastic growth independent of Phytochrome B and Cryptochrome 2 signaling, despite that these photoreceptors are positive regulators of low light-induced hyponastic growth.Key words: hyponastic growth, petiole, Arabidopsis, low light, ERECTA, differential growth, phytochrome B, cryptochrome 2Plants must adjust growth and reproduction to adverse environmental conditions. Among the strategies that plants employ to escape from unfavorable conditions is differential petiole growth-driven upward leaf movement, called hyponastic growth. Arabidopsis thaliana is able to exhibit a marked hyponastic response upon flooding, which is triggered by endogenous accumulation of the gaseous phytohormone ethylene.¹ Moreover, a similar response is triggered upon low light intensity perception and in response to supra-optimal temperatures.^2–5 By tilting the leaves to a more vertical position during submergence and shading, the plants restore contact with the atmosphere and light, respectively. The kinetics of the hyponastic growth response induced by the various stimuli is remarkably similar. This led to the hypothesis that shared functional genetic components may be employed to control hyponastic growth. Yet, at least part of the signaling cascades is parallel, as the hormonal control of the response differs between the stimuli. Low light-induced hyponastic growth for example does not require ethylene action.² Whereas the response to heat is antagonized by this hormone.⁵ The abiotic stress hormone abscisic acid (ABA) antagonizes ethylene-induced hyponastic growth and stimulates heat-induced hyponastic growth.^5,6 Moreover, ethylene-induced hyponasty does not involve auxin action⁷ whereas both heat- and low light-induced hyponasty require functional auxin signaling and transport components.^2,5In our recent paper, published in The Plant Journal,⁸ we employed Quantitative Trait Locus (QTL) analysis to identify loci involved in the control of ethylene-induced hyponastic petiole growth. By analyzing induced mutants and by complementation analysis of naturally occurring mutant accessions, we found that the leucine-rich repeat receptor-like Ser/Thr kinase gene ERECTA (ER) is a positive regulator of ethylene-induced hyponastic growth and most likely is causal to one of the identified QTLs. In addition, we demonstrated that the ER dependency is not via ER mediated control of ethylene production or sensitivity.Since low light-induced hyponasty does not require ethylene action,² ER may be part of the proposed shared signaling cascade leading to hyponastic growth where ethylene and low light signals meet. Therefore, we studied low light intensity-induced hyponasty in various erecta mutants. Moreover, natural occurring er mutant accessions complemented with a functional, Col-0 derived, ER allele were tested. The response of Lan-0 (Lan-0; with functional ER) to low light was indistinguishable from the response of Landsberg erecta (Ler) (Fig. 1A). However, complemented Ler (ER-Ler) showed an enhanced response compared to Ler (Fig. 1B). The response of mutant er105 was slightly attenuated compared to the wild type Columbia-0 (Fig. 1C). Mutant er104, however, showed an indistinguishable hyponastic growth phenotype to low light compared to the wild type Wassilewskija-2 (Ws-2) (Fig. 1D). Complementation of the natural occurring erecta mutant accession Vancouver-0 (Van-0) resulted in an enhanced hyponastic growth response to low light (Fig. 1E), whereas this was not the case for Hiroshima-1 (Hir-1) (Fig. 1F). Together, these data suggest that ER acts as positive regulator of low light-induced hyponastic growth and therefore may be part of the shared signaling cascade towards differential petiole growth. Yet, the effect is strongly dependent on the genetic background since the effects were not observed in every accession tested.Open in a separate windowFigure 1ERECTA involvement in low light-induced hyponasty. Effect of exposure to low light (spectral neutral reduction in light intensity from 200 to 20 µmol m⁻² s⁻¹) on the kinetics of hyponastic petiole growth in Arabidopsis thaliana. (A) mutant (circles) Ler and wild type (dashed line) Lan-0, (B) Ler and Ler complemented (ER-; squares) with the Col-0 ERECTA allele (ER-Ler), (C) er105 and Col-0 wild type, (D) er104 and Ws-2 wild type, (E) natural mutant Van-0 and Van-0 complemented with the Col-0 ER allele (ER-Van-0), (F) natural mutant Hir-1 and Hir-1 complemented with the Col-0 ER allele (ER-Hir-1). Petiole angles were measured using time-lapse photography and subsequent image analysis. Data is pairwise subtracted, which corrects for diurnal petiole movement in control conditions. For details on this procedure, growth conditions and materials, transformation protocol, treatments, data acquirement and all analyses see.^1,8 Error bars represent standard errors; n ≥ 12.Phytochrome B (PhyB) and Cryptochrome 2 (Cry2) photoreceptor proteins are required for a full induction of low light-induced hyponastic growth.² We transformed the phyb5 cry2 mutant9 (Ler genetic background) with Col-0 derived ER. This complementation did not restore the ability of phyb5 cry2 to induce hyponastic growth to neither ethylene (data not shown) nor low light conditions (Fig. 2A). Mutant phyb5 cry2 plants have a typical constitutive shade avoidance phenotype, reflected by severely elongated organs. This includes enhanced inflorescence and silique length and thin inflorescences (Fig. 2B-D). Complementation with ER resulted in a significant additional effect on these parameters (Fig. 2B-D). Together, this suggests that ER is not an integral part of PhyB nor Cry2 signaling with respect to (hyponastic) growth. Moreover, PhyB and Cry2 control of plant architecture does not require ER action. Rather, ER seems to mediate growth via genetic interaction with light-reliant growth mechanisms, instead of being downstream of photoreceptor action. Studies on the effects of ER on shade avoidance responses and various hormone responses, including cytokinin and auxin, led to the similar conclusion, suggesting a possible role for ER as a molecular hub coordinating light- and hormone-mediated plant growth.^10,11 One could speculate that ER fine-tunes other (than light) environmental clues with light signaling components. A comparable conclusion was drawn previously for gibberellin (GA) reliant growth mechanisms, as er enhanced the negative effect on plant size of the short internode (shi) mutation12 and er represses the positive effect of the spindly mutation in a GA independent manner.¹³Open in a separate windowFigure 2Effects of ERECTA on light signaling. (A) Effect of exposure to low light (spectral neutral reduction in light intensity from 200 to 20 µmol m⁻² s⁻¹) on the kinetics of hyponastic petiole growth of Ler (dashed lines), the photoreceptor double mutant phyb5 cry2 (circles) and this mutant complemented with the Col-0 ERECTA (ER-phyb cry2; squares). For details see legend Figure 1. (B) Plant height, (C) silique length and (D) inflorescence stem thickness of the above mentioned lines. These parameters were measured when the last flower on the plant developed a silique. Plant height was measured from root/shoot junction to inflorescence top. Stem thickness was measured ∼1 cm above the root/shoot junction with a caliper and silique lengths were measured from representative pedicels in the top ∼10 cm of the main inflorescence stem. Error bars represent standard errors; n ≥ 12. Significance levels; *p < 0.05; **p < 0.01; ***p < 0.001; ns = non significant, by Students t-test.     

Brassinosteroid signaling modulates submergence-induced hyponastic growth in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

The role of brassinosteroids (BRs) in hyponastic growth induced by submergence was investigated in Arabidopsis thaliana. Under flooding conditions, exogenously applied BRs increased hyponastic growth of rosette leaves. This hyponastic growth was reduced in a BR insensitive mutant (bri1-5), while it was increased in a BR dominant mutant (bes1-D). Further, expression of hypoxia marker genes, HRE1 and HRE2, was elevated in submerged bes1-D. These results indicate that BRs exert a positive action on hyponastic growth of submerged Arabidopsis leaves. Expression of ethylene biosynthetic genes, such as ACS6, ACS8 and ACO1, which are up-regulated by submergence, was also activated by application of BRs and in bes1-D. The enhanced hyponastic growth in submerged bes1-D was significantly reduced by application of cobalt ion, suggesting that BRs control hyponastic growth via ethylene, which seems to be synthesized by ACO6 and ACO8 followed by ACO1 in submerged leaves. A double mutant, bes1-Dxaco1-1, showed hyponastic growth activity similar to that seen in aco1-1, demonstrating that the BR signaling for regulation of hyponastic growth seems to be an upstream event in ethylene-induced hyponastic growth under submergence in Arabidopsis.     

Genetic and Physiological Analysis of a New Locus in Arabidopsis That Confers Resistance to 1-Aminocyclopropane-1-Carboxylic Acid and Ethylene and Specifically Affects the Ethylene Signal Transduction Pathway

A population of M2 seedlings of Arabidopsis thaliana was screened for mutants that were insensitive to the ethylene precursor 1-aminocyclopropane-1-carboxylate (ACC). Several independent lines were obtained and proved insensitive to both ACC and ethylene. Two lines were identified as alleles of a single recessive mutation, designated ain1. Linkage analysis indicated that the ain1 gene is located on chromosome 1, adjacent to the cer5 marker and, therefore, genetically distinct from previously identified ethylene resistance loci. General phenotypic aspects of ain1 mutants were similar to wild type. For both alleles, the level of insensitivity to ethylene at the seedling stage was indistinguishable in terms of elongation growth. In contrast, the gravitropic response of ain1-1 seedlings was slower than that of wild-type and ain1-2 seedlings. At the adult stage, stress responses of mutants were similar to wild type. However, ethylene-induced leaf senescence was delayed in both mutants. In addition, we observed significant interallelic variation in ethylene production rates. Growth inhibition experiments showed that the ain1 mutation does not confer resistance to other hormones. Thus, ain1 most probably affects a step specific for the ethylene signal transduction pathway.     

THE INTERACTION OF AUXIN AND ETHYLENE IN THE MAINTENANCE OF LEAF BLADE FORM IN PHASEOLUS VULGARIS L. VAR. PINTO

Auxin treatment results in hyponastic curvature of the primary leaves of Phaseolus vulgaris L. var pinto. Ethylene production by hyponastic leaves is detected within 1 hr after treatment with IAA in concentrations at or above 1 μm. The amount of ethylene detected is proportional to the concentration of auxin applied. Untreated control leaves and leaves treated with 2,3,5-tri-iodobenzoic acid or gibberellic acid did not produce ethylene detectable by our equipment. The hyponastic curvature induced by auxin treatment can be inhibited by exogenous application of ethylene or ethylene-generating compounds, and these treatments produce epinasty in auxin-treated leaves. Treatment with inhibitors of ethylene synthesis or action, such as aminoethoxy-vinylglycine, carbon dioxide, or heat treatment, prolong hyponasty. The planar form, therefore, appears to be affected by both hyponastic auxin effect and an epinastic ethylene effect.     

GROWTH AND GRAVITATIONAL RESPONSE IN THE DEVELOPMENT OF LEAF BLADE HYPONASTY

Rapid (4 hr) auxin-induced hyponastic curvature of primary leaves of Phaseolus vulgaris is shown to depend on a positive increase in growth of the lower portion of the blade. The curvature involves laminar growth as well as vein growth and is not due to simple turgor changes. The response is sensitive to gravitational orientation, as inversion and horizontal rotation reduce the auxin-induced curvature. The ethylene-generating compound, 2-chloroethylphosphonic acid, had no hyponastic effect on the leaves when applied to either the upper or lower surface and it inhibited auxin-induced hyponasty. This inhibition was additive to that of inversion. Long-term (24–48 hr) effects of 1 mM auxin depend on the surface of the leaf treated. Application to the upper surface results in epinasty, lower surface application in hyponasty, although the initial response in each case is a hyponastic curvature. A dorsi-ventral auxin transport system and differential auxin sensitivity of upper and lower portions of the leaf blade are postulated to account for these responses.     

Auxin and Ethylene Regulate Elongation Responses to Neighbor Proximity Signals Independent of Gibberellin and DELLA Proteins in Arabidopsis

Plants modify growth in response to the proximity of neighbors. Among these growth adjustments are shade avoidance responses, such as enhanced elongation of stems and petioles, that help plants to reach the light and outgrow their competitors. Neighbor detection occurs through photoreceptor-mediated detection of light spectral changes (i.e. reduced red:far-red ratio [R:FR] and reduced blue light intensity). We recently showed that physiological regulation of these responses occurs through light-mediated degradation of nuclear, growth-inhibiting DELLA proteins, but this appeared to be only part of the full mechanism. Here, we present how two hormones, auxin and ethylene, coregulate DELLAs but regulate shade avoidance responses through DELLA-independent mechanisms in Arabidopsis (Arabidopsis thaliana). Auxin appears to be required for both seedling and mature plant shoot elongation responses to low blue light and low R:FR, respectively. Auxin action is increased upon exposure to low R:FR and low blue light, and auxin inhibition abolishes the elongation responses to these light cues. Ethylene action is increased during the mature plant response to low R:FR, and this growth response is abolished by ethylene insensitivity. However, ethylene is also a direct volatile neighbor detection signal that induces strong elongation in seedlings, possibly in an auxin-dependent manner. We propose that this novel ethylene and auxin control of shade avoidance interacts with DELLA abundance but also controls independent targets to regulate adaptive growth responses to surrounding vegetation.     

Auxin is a positive regulator for ethylene-mediated response in the growth of Arabidopsis roots

The requirement of auxin for the ethylene-mediated growth response in the root of Arabidopsis thaliana seedlings was investigated using two ethylene-resistant mutants, aux1-7 and eir1-1, whose roots have been shown to have a defect in the auxin influx and efflux carriers, respectively. A 50% inhibition of growth (I(50)) was achieved with 0.84 microl liter(-1) ethylene in wild-type roots, but 71.3 microl liter( -1) ethylene was required to induce I(50) in eir1-1 roots. In aux1-7 roots, I(50) was not obtained even at 1,000 microl liter(-1) ethylene. By contrast, in the presence of 10 nM 1-naphthaleneacetic acid (NAA), the concentrations of ethylene required to induce I(50) in eir1-1 and aux1-7 roots were greatly reduced nearly to the level required in wild-type roots. Since the action of NAA to restore the ethylene response in aux1-7 roots was not replaced by IAA, an increase in the intracellular level of auxin is likely to be the cause for the restoration of ethylene response. NAA at 10 nM did not inhibit root growth when applied solely, but it was the optimum concentration to recover the ethylene response in the mutant roots. These results suggest that auxin is a positive regulator for ethylene-induced inhibition in root elongation.