Local application of indole-3-acetic acid,by resin beads to intact growing maize roots

Five types of anion-exchanger resin beads which had adsorbed indole-3-acetic acid (IAA) were tested as IAA donors. The rate of IAA-uptake by beads was a function of time and pH. The release was relatively steady during 6 h application on vertical maize roots. No IAA degradation occurred in the beads (Amberlite IRA 400 type) but 45.8% was metabolised in the roots during treatment. Beads loaded with IAA and placed on one side of the root (at 2.20±0.03 mm from the tip) induced a curvature towards and above the bead (23.3±1.1 degrees after 5.25 h application). In contrast, control beads (without IAA) did not change the axial growth rate. Applied IAA seemed to move differently from endogenous IAA. The use of resin beads loaded with IAA offers a technique to study the effects of local IAA application on intact growing roots.Abbreviations 3,3-DGA 3,3 dimethyl-glutaric acid - HPLC high-performance liquid chromatography - IAA indole-3-acetic acid - Ox-IAA oxindole-3-acetic acid     

NPH3- and PGP-like genes are exclusively expressed in the apical tip region essential for blue-light perception and lateral auxin transport in maize coleoptiles

Phototropic curvature results from differential growth on two sides of the elongating shoot, which is explained by asymmetrical indole-3-acetic acid (IAA) distribution. Using 2 cm maize coleoptile segments, 1st positive phototropic curvature was confirmed here after 8 s irradiation with unilateral blue light (0.33 μmol m(-2) s(-1)). IAA was redistributed asymmetrically by approximately 20 min after photo-stimulation. This asymmetric distribution was initiated in the top 0-3 mm region and was then transmitted to lower regions. Application of the IAA transport inhibitor, 1-N-naphthylphthalamic acid (NPA), to the top 2 mm region completely inhibited phototropic curvature, even when auxin was simultaneously applied below the NPA-treated zone. Thus, lateral IAA movement occurred only within the top 0-3 mm region after photo-stimulation. Localized irradiation experiments indicated that the photo-stimulus was perceived in the apical 2 mm region. The results suggest that this region harbours key components responsible for photo-sensing and lateral IAA transport. In the present study, it was found that the NPH3- and PGP-like genes were exclusively expressed in the 0-2 mm region of the tip, whereas PHOT1 and ZmPIN1a, b, and c were expressed relatively evenly along the coleoptile, and ZmAUX1, ZMK1, and ZmSAURE2 were strongly expressed in the elongation zone. These results suggest that the NPH3-like and PGP-like gene products have a key role in photo-signal transduction and regulation of the direction of auxin transport after blue light perception by phot1 at the very tip region of maize coleoptiles.     

Indole-3-acetic Acid (IAA) and IAA Conjugates Applied to Bean Stem Sections: IAA Content and the Growth Response

High resolution growth recording techniques and reverse isotope dilution analysis were used to study the relationship between indole-3-acetic acid (IAA) concentration and curvature of excised bean (Phaseolus vulgaris L. cv Bush Burpee Stringless) first internode sections unilaterally treated with hormone. The maximum rate of curvature occurred rapidly (within 25 minutes) and was proportional to the log of the amount of applied IAA recovered in the tissue. The rate of curvature decreased after 30 minutes although little or no lateral migration of applied IAA occurred and tissue levels of IAA increased. The biologic activity of IAA-amino acid conjugates was found to be directly related to the amount of free IAA, resulting from their hydrolysis, which could be recovered from the tissue.     

The Movement and Physiological Effect of Indoleacetic Acid Following Point Applications to Root Tips of Zea mays

Pinpoint applications of labeled and non-labeled indoleacetic acid (IAA) on resin beads were made, without injury, to vertical roots of intact seedlings of Zea mays. Points of application were at the extreme tip of the root, 0.5, 2 and 5 mm from the root tip. The movement of label and bending of the roots was recorded. Radiolabel was found to move basipetally from the extreme tip and 0.5 mm applications to a similar extent, reaching 8 mm from the tip. The level of label in the growing zone after 4 h was 10% of that found in the extreme tip. Movement from 2 and 5 mm applications was equal in both directions. Higher amounts of non-labeled IAA caused bending towards the point of application if applied at 0.5 or 2 mm but not at 5 mm from the tip. It is proposed that any endogenous IAA in the root cap could move to the growing zone and cause a unilateral inhibition of growth, provided that it was in the same transport pool as the exogenously applied IAA.     

Auxin, transport and growth in intact roots of Vicia faba

Transport of IAA applied to the intact root of Vicia seedlingswas investigated in relation to root growth. The root was treatedat 3–4, 4–5 or 7–8 mm from the tip with anarrow ring of lanolin paste containing IAA or IAA-2-¹⁴C ingrowth or transport experiments, respectively. The growth processalong the root axis was examined in every 1-mm part from thetip at 30 min, 1 or 4 hr intervals. The elongation zone of thecontrol root was 1–9 mm from the tip. IAA treatment broughtabout no significant change in the growth of the region apicalto the treated site, whereas distinct inhibition of growth occurredin the region basal to the treated site within 1 hr. The transportof radioactivity was observed in both acropetal and basipetaldirections within 1 hr, but the latter predominated for 8 hror more; the nearer to the tip the treatment site, the longerthe predominance lasted. The velocities of acropetal and basipetaltransport were estimated at about 4 and 8 mm/hr, respectively.Autoradiographs of transverse section of roots showed that basipetaltransport occurred mainly through the outer part of the root,whereas acropetal transport occurred mainly through the innerpart, the central cylinder. It may be concluded that the basipetallytransported IAA which passed through the outer part of the rootinhibited the elongation of the intact root. (Received November 25, 1975; )     

IAA-Induced Growth Responses of Decapitated Corn Seedlings: Indications of Two Apparent Adaptations with a Possible Role in Gravitropism

The vertical growth responses of corn seedlings (Zea mays L. Mo17 × B73) were determined over an 8-hour period. When seedlings were decapitated 3 millimeters from the coleoptile's tip and supplied with indole-3-acetic acid (IAA) in 1.5% agar blocks, the response was dependent both on time and IAA concentration. The dose-response curves changed in shape and magnitude depending on the total time of IAA application. High concentrations (>3.2 × 10⁻⁶ molar) initially produced high relative growth rates that decreased back to the intact rate (0.03 millimeter per hour per millimeter) after 3 hours. Low concentrations (<1.0 × 10⁻⁶ molar), or agar blocks without IAA, resulted in a rapid decrease from the intact rate to a level that stabilized at 0.01 millimeter per hour per millimeter until the growth rate began to recover after 3 to 4 hours. Intermediate concentrations produced responses similar to that of the intact organ, though some features of these responses were unique.

The coleoptile curvature in response to gravity depended upon whether the coleoptiles were intact, decapitated, or decapitated and supplied with IAA. Coleoptiles decapitated and not supplied wth IAA showed little or no curvature for 3 hours after decapitation. By this time an adaptation, evoked by the low IAA level, had developed and the coleoptiles began to curve steadily. When 1.0 or 3.2 × 10⁻⁶ molar IAA was supplied, curvature was initiated within the first 30 minutes and reached a maximum rate before decreasing and stopping after 3 to 4 hours. The sequence of events in response to these concentrations was similar to the intact sequence but the curvature rate was reduced to one-third to one-half. A model for the autotropic response involving an auxin concentration-dependent, growth-modulating mechanism capable of two modes of adaptation is described.

     

Can Lateral Redistribution of Auxin Account for Phototropism of Maize Coleoptiles?

Elongation growth of intact, red-light grown maize (Zea mays L.) coleoptiles was studied by applying a small spot of an indole acetic acid (IAA)-lanolin mixture to the coleoptile tip. We report that: (a) endogenous auxin is limiting for growth, (b) an approximately linear relation holds between auxin concentration and growth rate over a range which spans those rates occurring in phototropism, and (c) an auxin gradient established at the coleoptile tip is well sustained during its basipetal transport. We argue that the growth differential underlying coleoptile phototropism (first-positive curvature) can be explained by redistribution of auxin at the coleoptile tip.     

Mediation of tropisms by lateral translocation of endogenous indole-3-acetic acid in maize coleoptiles

Abstract. The hypothesis that tropic responses result from lateral auxin gradients was examined in coleoptiles of red-light-grown maize ( Zea mays L.) by measuring endogenous IAA (indole-3-acetic acid) using a physicochemical method. Phototropic stimulation (unilateral blue light; 8s at 0.33 μmol m⁻²s⁻¹) was found to induce a lateral gradient of solvent-extractable IAA in a subapical zone (2-7mm from the tip). The gradient occurred in advance of the bending response, with a decrease of IAA in the irradiated half and a compensatory increase in the shaded half. The maximal gradient measured was about 1:2 (irradiated: shaded). Diffusible IAA, obtained from the cut end of an excised coleoptile tip (3mm long, with its base split by 1mm), was similarly redistributed between the two sides, indicating that IAA is laterally translocated in the tip and that the resulting IAA gradient migrates to the subapical zone. A smaller gradient was induced in a basal zone (12-17mm from the tip). This gradient was initiated about 20 min later than that at the subapical zone, in agreement with a similar delay of bending observed in this zone. Gravitropic stimulation (60° from the vertical) also resulted in a lateral gradient of extractable IAA in the subapical zone, the gradient preceding the bending response. It is concluded that the tropisms of maize coleoptiles are mediated by IAA gradients, which are most likely caused by lateral IAA transport as the Cholodny-Went theory of tropisms describes. From IAA measurement data, the mean velocity of basipetally-polar transport of endogenous IAA was estimated to be 12 mm h⁻¹.     

Transport and Degradation of Auxin in Relation to Geotropism in Roots of Phaseolus vulgaris

The movement of auxin in Phaseolus vulgaris roots has been examined after injection of IAA^?3H into the basal root/hypocotyl region of intact, dark-grown seedlings. Only a portion of the applied IAA^?3H was transported unchanged to the root tip. The major part of the chromatographed, labelled compounds translocated to the roots was indole-3-acetylaspartic acid (IAAsp) and an unidentified compound running near the front in isopropanol, ammonia, water. The velocity of the auxin transport (7.2 mm per hour) was calculated from scintillation countings of methanol extracts from serial sections of the root. An accumulation of radioactive compounds in the extreme root tip, was observed 5 h after the injection of IAA. The influence of exogenous IAA on the geotropical behaviour of the bean seedling roots was examined. Pretreated roots were stimulated for 5 min in the horizontal position and then rotated parallel to the horizontal axis of the klinostat for 60 or 90 min. The resulting geotropic curvature of IAA-injected and control roots showed significantly different patterns of development. When the stimulation was started 5 h after application of the auxin, the geotropic curvature became larger in roots of the injected plants than in the controls. If, however, the translocation period was extended to 20 h the geotropic curvature was significantly smaller in the roots of the injected plants. The auxin injection did not significally affect the rate of root elongation. The change in geotropical behaviour of the roots is interpreted as a result of the influence of the conversion products of the applied IAA on the geotropical responsiveness.     

Altered Growth Response to Exogenous Auxin and Gibberellic Acid by Gravistimulation in Pulvini of Avena sativa

Pulvini of excised segments from oats (Avena sativa L. cv Victory) were treated unilaterally with indoleacetic acid (IAA) or gibberellic acid (GA₃) with or without gravistimulation to assess the effect of gravistimulation on hormone action. Optimum pulvinus elongation growth (millimeters) and segment curvature (degrees) over 24 hours were produced by 100 micromolar IAA in vertical segments. The curvature response to IAA at levels greater than 100 micromolar, applied to the lower sides of gravistimulated (90°) pulvini, was significantly less than the response to identical levels in vertical segments. Furthermore, the bending response of pulvini to 100 micromolar IAA did not vary significantly over a range of presentation angles between 0 and 90°. In contrast, the response to IAA at levels less than 10 micromolar, with gravistimulation, was approximately the sum of the responses to gravistimulation alone and to IAA without gravistimulation. This was observed over a range of presentation angles. Also, GA₃ (0.3-30 micromolar) applied to the lower sides of horizontal segments significantly enhanced pulvinus growth and segment curvature, although exogenous GA₃ over a range of concentrations had no effect on pulvinus elongation growth or segment curvature in vertical segments. The response to GA₃ (10 micromolar) plus IAA (1.0 or 100 micromolar) was additive for either vertical or horizontal segments. These results indicate that gravistimulation produces changes in pulvinus responsiveness to both IAA and GA₃ and that the changes are unique for each growth regulator. It is suggested that the changes in responsiveness may result from processes at the cellular level other than changes in hormonal sensitivity.     

Transport of [3H]IAA label in gravistimulated primary roots of maize

The curvature of roots in response to gravity is attributed to the development of a differential concentration gradient of IAA in the top and bottom of the elongation region of roots. The development of the IAA gradient has been attributed to the redistribution of IAA from the stele to cortical tissues in the elongation region. The gravistimulated redistribution of IAA was investigated by applying [³H]IAA to the cut surface of 5 mm apical primary root segments. The movement of label from the stele-associated [³H]IAA into the root, tip, root cap, and cortical tissues on the top and bottom of the elongation region was determined in vertically growing roots and gravistimulated roots. Label from the stele moved into the region of cell differentiation (root tip) prior to accumulating in the elongation region. Little label was observed in the root cap. Gravistimulation did not increase the amount of label moving from the stele; but gravistimulation did increase the amount of label accumulating in cortical tissues on the lower side of the elongation region, and decreased the amount of label accumulating in cortical tissues on the upper side of the elongation region. Removal of the cap prior to or immediately following gravity stimulation rendered the roots partially insensitive to gravity and also prevented gravity-induced asymmetric redistribution of label. However, removal of the root cap following 30 min of gravistimulation did not alter root curvature or the establishment of an IAA asymmetry across the region of root elongation. These results suggest that a signal originating in the root cap directs auxin redistribution in tissues behind the root cap, leading to the development of an asymmetry of IAA concentration in the elongation region that in turn causes the differential growth rate in the elongation region of a graviresponding root.     

Transport of [3H]IAA label in gravistimulated primary roots of maize

The curvature of roots in response to gravity is attributed to the development of a differential concentration gradient of IAA in the top and bottom of the elongation region of roots. The development of the IAA gradient has been attributed to the redistribution of IAA from the stele to cortical tissues in the elongation region. The gravistimulated redistribution of IAA was investigated by applying [³H]IAA to the cut surface of 5 mm apical primary root segments. The movement of label from the stele-associated [³H]IAA into the root, tip, root cap, and cortical tissues on the top and bottom of the elongation region was determined in vertically growing roots and gravistimulated roots. Label from the stele moved into the region of cell differentiation (root tip) prior to accumulating in the elongation region. Little label was observed in the root cap. Gravistimulation did not increase the amount of label moving from the stele; but gravistimulation did increase the amount of label accumulating in cortical tissues on the lower side of the elongation region, and decreased the amount of label accumulating in cortical tissues on the upper side of the elongation region. Removal of the cap prior to or immediately following gravity stimulation rendered the roots partially insensitive to gravity and also prevented gravity-induced asymmetric redistribution of label. However, removal of the root cap following 30 min of gravistimulation did not alter root curvature or the establishment of an IAA asymmetry across the region of root elongation. These results suggest that a signal originating in the root cap directs auxin redistribution in tissues behind the root cap, leading to the development of an asymmetry of IAA concentration in the elongation region that in turn causes the differential growth rate in the elongation region of a graviresponding root.     

Geotropic curvature of decapitated Avena coleoptiles after application of tips of maize roots,tips of Avena coleoptiles,indoleacetic acid,or abscisic acid

Isolated Avena coleoptiles were decapitated at different distances from the tip and then placed horizontally, after which the geotropic curvature was measured. No geotropic curvature could be detected during the first 3 h. Later, upward curvature occurred which was found to depend inversely on the length of the decapitated tips. When the tips of maize roots or Avena coleoptiles were placed on the cut surface of decapitated Avena coleoptiles, the coleoptiles showed a significantly stronger upward curvature as compared to controls which had been provided with agar blocks on the cut surface. The same upward curvature was found with decapitated coleoptiles provided with agar blocks containing 10^-6 or 10^-7 M indoleacetic acid (IAA). After application of abscisic acid (ABA) at concentrations of 10^-6 and 10^-8 M to the decapitated coleoptiles, the curvature observed was not different from that of the controls; at higher concentrations of ABA the curvature was found to be lower than that of the controls. It is concluded that root tips secrete a substance which may replace the effect of IAA in coleoptiles. The results are discussed in view of the validity of the Cholodny-Went hypothesis for the geotropic reaction of roots.Abbreviations ABA abscisic acid - IAA 3-indoleacetic acid     

Analysis of growth patterns during gravitropic curvature in roots ofZea mays by use of a computer-based video digitizer

A computer-based video digitizer system is described which allows automated tracking of markers placed on a plant surface. The system uses customized software to calculate relative growth rates at selected positions along the plant surface and to determine rates of gravitropic curvature based on the changing pattern of distribution of the surface markers. The system was used to study the time course of gravitropic curvature and changes in relative growth rate along the upper and lower surface of horizontally-oriented roots of maize (Zea mays L.). The growing region of the root was found to extend from about 1 mm behind the tip to approximately 6 mm behind the tip. In vertically-oriented roots the relative growth rate was maximal at about 2.5 mm behind the tip and declined smoothly on either side of the maximum. Curvature was initiated approximately 30 min after horizontal orientation with maximal (50°) curvature being attained in 3 h. Analysis of surface extension patterns during the response indicated that curvature results from a reduction in growth rate along both the upper and lower surfaces with stronger reduction along the lower surface.     

Effect of indoleacetic acid,abscisic acid,root tips and coleoptile tips on growth and curvature of maize roots

The effect of externally applied indoleacetic acid (IAA) and abscisic acid (ABA) on the growth of roots of Zea mays L. was measured. Donor blocks of agar with IAA or ABA were placed laterally on the roots and root curvature was measured. When IAA was applied to vertical roots, a curvature directed toward the donor block was observed. This curvature corresponded to a growth inhibition at the side of the root where the donor was applied. When IAA was applied to horizontal roots from the upper side, normal geotropic downward bending was delayed or totally inhibited. The extent of retardation and the inhibition of curvature were found to depend on the concentration of IAA in the donor block. ABA neither induced curvature in vertical roots nor inhibited geotropic curvature in horizontal roots; thus the growth of roots was not inhibited by ABA. However, when, instead of donor blocks, root tips or coleoptile tips were placed onto vertical roots, a curvature of the roots was observed.Abbreviations ABA abscisic acid - IAA 3-indoleacetic acid     

Effect of Indoleacetic Acid and Related Indoles on Lactobacillus sp. Strain 11201 Growth, Indoleacetic Acid Catabolism, and 3-Methylindole Formation

A study was conducted to determine the activity of the 3-methylindole (3MI)-forming enzyme in Lactobacillus sp. strain 11201. Cells were incubated anaerobically with 17 different indolic and aromatic compounds. Indoleacetic acid (IAA), 5-hydroxyindoleacetic acid, 5-methoxy-3-indoleacetic acid, indole-3-pyruvate, or indole-3-propionic acid induced 3MI-forming activity. The highest total enzyme activity induced by IAA was observed in cells incubated with an initial concentration of 1.14 mM IAA. Peak activity of the 3MI-forming enzyme occurred 4 h after bacteria were incubated with either 0.114 or 1.14 mM IAA. Enzyme activity peaked earlier (2 h) and disappeared more rapidly at 5.7 mM IAA than at other concentrations of IAA. The effects of IAA and 3MI on the growth of Lactobacillus sp. strain 11201 and formation of 3MI from IAA also were determined. Bacterial growth and 3MI formation from IAA were reduced in medium containing exogenous 3MI. The growth depression observed in medium containing 5.7 mM IAA appears to be due to the toxicity of 3MI rather than IAA. The formation of 3MI in this ruminal Lactobacillus sp. is mediated by an inducible enzyme, and as 3MI accumulates, bacterial growth and rates of 3MI formation from IAA are reduced.     

Effects of γ-irradiation on elongation and indole-3-acetic acid level of maize (Zea mays) coleoptiles

The effects of γ-irradiation on elongation and the level of indole-3-acetic acid (IAA) of maize (Zea mays) coleoptiles were investigated. When 3-day-old seedlings of maize were exposed to γ-radiation lower than 1 kGy, a temporal retardation of coleoptile elongation was induced. This retardation was at least partly ascribed to a temporal decrease in the amount of free IAA in coleoptile tips on the basis of the following facts: (1) the reactivity to IAA of the elongating coleoptile cells was not altered by irradiation; (2) endogenous IAA level in the tip of irradiated coleoptiles was at first unchanged, but then declined before returning to nearly the same level as that of the non-irradiated control; and (3) the amount of IAA that diffused from coleoptile tip sections showed a similar pattern to that of endogenous IAA. The rate of conversion between free and conjugated IAA was not significantly affected by irradiation. These results suggest that a temporal inhibition of maize coleoptile elongation induced by γ-irradiation can be ascribed to the reduction of endogenous IAA level in the coleoptile tip, and this may originate from the modulation in the rate of IAA biosynthesis or catabolism.     

The response to auxin of rapeseed (Brassica napus L.) roots displaying reduced gravitropism due to transformation by Agrobacterium rhizogenes

It has recently been documented that, compared to untransformed controls, the roots of oilseed rape (Brassica napus L. CV CrGC5) seedlings transformed by Agrobacterium rhizogenes A4 show a reduced gravitropic reaction (Legué et al. 1994, Physiol Plant 91: 559–566). After stimulation at 90°C or 135°, the transformed root tips curve, but never reach a vertical orientation. In the present study, we investigated the causes of reduced gravitropic bending observed in stimulated transformed root tips. First, we localized the gravitropic curvature in normal and in transformed roots after 1.5 h of stimulation. The cells involved in root curvature (target cells) corresponded at the cellular level to the apical part of the zone of increasing cell length. In transformed roots grown in the vertical position, these cells showed a reduction in cell length compared to controls. Because auxin is considered to be the gravitropic mediator, the response of normal and transformed roots to exogenous auxin was studied. Indole-3-acetic acid (IAA) was applied along the first 3 mm using resin beads loaded with the hormone. In comparison to normal roots, transformed roots showed reduced bending toward the bead at all points of bead application. Moreover, the cells which responded to IAA corresponded to the target cells involved in the gravitropic reaction. The level of endogenous IAA was lower in transformed roots. Thus, it was concluded that the modified behavior of transformed roots during gravitropic stimulation could be due to differences either in IAA levels or in reactivity of the target cells to the message from the cap.Abbreviations DEZ distal elongation zone - ELISA enzymelinked immunosorbent assay - T-DNA DNA transferred from Agrobacterium rhizogenes to the plant genome This work was supported by the Centre National d'Etudes Spatiales.     

Inhibitors from Carob (Ceratonia siliqua L.): II. Effect on Growth Induced by Indoleacetic Acid or Gibberellins A(1), A(4), A(5), and A(7)

Two inhibitory fractions (B₁ and C) from extracts of immature fruit of carob were tested for their ability to inhibit the action of indoleacetic acid (IAA) in three bioassays. There was no reduction of IAA-induced reactions in the Avena curvature test, abscission of debladed coleus petioles, or growth of cucumber hypocotyls. The highest ratio of inhibitor to IAA was 10,000 times greater than the ratio necessary to inhibit by 50% the growth caused by an equivalent amount of gibberellin A₃ in pea seedlings. At the highest concentration used, fraction C alone caused curvature of Avena coleoptiles. The inhibitory fractions appeared to enhance the effect of IAA in the cucumber test.