Interactions of microtubule disorganizers, plant hormones, and red light in wheat coleoptile segment growth

Growth response of coleoptile segments excised from 3-day-old seedlings of wheat (Triticum vulgare cv. Baart) to gibberellic acid, indoleacetic acid, and 2,4-dichlorophenoxyacetic acid, to red light, and to several microtubule disorganizers depends on the initial position of the excised segment in the intact coleoptile. Red light, 660 nm, stimulates the growth of the apical cells, but inhibits markedly the growth of the cells in the basal region of the coleoptile. The effects of red light are independent of sucrose, gibberellic acid, indoleacetic acid, and 2,4-dichlorophenoxyacetic acid, even though these substances themselves markedly affect the growth of the coleoptile segments. Concentractions of the microtubule disorganizers, vinblastine sulfate, cupric chloride, urea, and colchicine, which do not alter significantly the growth of the dark control apical segments, substantially repress the promotive effects of red light or auxin on the increase in length of the apical cells of the coleoptile. This suggests that stimulation by red light and by auxin involves microtubule production. Microtubule disorganizers repress the growth of elongating cells of the coleoptile, yet on the other hand, auxin and irradiation do not alter significantly the response of basal cells to the microtubule disorganizing agents. We hypothesized that light and growth regulators induce polymerization of nonaggregated microtubule subunits, resulting in faster growth.     

Response of excised Avena coleoptile segments to red and far-red light

Summary In seeking a simple, red light-promoted straight growth test in which phytochrome assays may be conducted without interference by protochlorophyll, the response of excised Avena coleoptile segments to red and far-red light was re-examined. The elongation of apical (non-decapitated) segments is promoted by a brief exposure to red light, and this effect is almost completely nullified by an immediately subsequent exposure to far-red light. Although growth promotion by red light occurs in distilled water alone, the effect is greater on a medium consisting of 0.02 M phosphate buffer, pH 6.2 to 6.4, with 1 to 2% sucrose. Over the pH range 4.5 to 7.4, dark-growth decreases with increasing pH, but the absolute increment brought about by red light is nearly constant. Elongation appears to be entirely the result of increased cell size.Contrary to previous reports, similar results can be obtained with subapical (decapitated) coleoptile segments, although the absolute magnitude of the response is reduced.Research carried out at Brookhaven National Laboratory under the auspices of the U.S. Atomic Energy Commision.     

Phytochrome-regulated growth of excised coleoptile tips of Triticum aestivum induced by blue, red and far-red light

The elongation growth of 5-mm tip sections of 3-day-old etiolatedwheat coleoptiles was promoted by light flashes of 436, 650and 750 nm in comparison with the dark control. As short as7/1,000 sec red light (R) of 14,420 W/m² led to saturation ofthe R-induced growth response. At quantum-identical far-redlight (FR) irradiation, a plateau developed at about 9/1,000sec which, however, was interrupted at longer irradiation periods.Varying dark periods between R or FR activation of growth anda second FR flash, a light-independent period with a half-lifeof 45 to 60 sec was found. Only after this light-independentperiod could R or FR activation of growth be reversed by FR.A second light-independent period was found by the followingirradiation schedule: 1 sec R/100 sec darknees(D)/l sec FR/varied D/l sec R. Our experimental conditions enabled us (a) to distinguish betweenactivation and inactivation by FR, (b) to clarify the reactionchain of different phytochrome forms and (c) to determine thehalf-life of light-independent phytochrome reactions.     

Influence of red light and acetylcholine on 45Ca2+ uptake by oat coleoptile cells

Influence of red light and acetylcholine on ⁴⁵Ca²⁺ uptake by oat coleoptile cells was examined. It was found that the uptake is passive in darkness, while short, 10–15 min. exposure of coleoptile sections to red light or treatment with acetylcholine solution increases the rate of ⁴⁵Ca²⁺ uptake from the medium. Calcium channel blockers, La³⁺ and Verapamil, hinder ⁴⁵Ca²⁺ uptake in darkness and neutralize the stimulative influence of red light and acetylcholine.     

Effect of auxinhormone lanthanide complexes on the growth of wheat coleoptile

Naphthalene-1-acetic acid (HNAA), dichlorophenoxy acetic acid (HDAA), and indole-3-acetic acid (HIAA) are auxinhormones that can affect the growth of plants. The lanthanide complexes of the above auxinhormone LnA₃-3H₂O (Ln = La³⁺, Ce³⁺, Sm³⁺, Er³⁺, Yb³⁺; HA = HNAA, HDAA, HIAA; A = NAA^-, DAA^-, IAA^-) were synthesized and are characterized in this paper. The solubility and IR spectra of these complexes were also studied. Experiments of the effects of LnCl₃-nH₂O, HA, and LnA₃-3H₂O on the growth rate of wheat coleoptile sections show, that LnCl₃-nH₂O promotes the growth of wheat coleoptile when this compounds concentration is lower than 2 x 10^-5 M and the promotion is very significant when the concentration of Ln³⁺ is lower than 8 x 10^-6 M. It was also found that the effect of LnA₃· 3H₂O on the growth of wheat coleoptile is stronger than that of LnCl₃·nH₂O and HA, which indicates that the combination of Ln³⁺ with HA act synergistically.     

Effect of light and gibberellic acid on coleoptile and first-foliage-leaf growth in durum wheat (Triticum durum Desf.)

A comparison has been made of the relative effectiveness of light quality and quantity and gibberellic acid (GA₃) treatment on the elongation growth of the coleoptile and the first foliage leaf in durum wheat (Triticum durum Desf. cvs. Cappelli and Creso). The cultivar Creso is a shortstrawed variety carrying the Gai 1 gene on chromosome 4A, which influences both plant height and insensitivity to applied gibberellins. The main conclusions are as follows: 1) coleoptile elongation growth appears to be modulated via the fluencerate-dependent action of a blue-light receptor and via a low energy response of phytochrome; 2) the inhibition of first-foliage-leaf growth depends on the operation of a single blue-light-responsive photoreceptor; 3) high energy blue light produces the same inhibitory effect on the two wheat cultivars, whereas at relatively low fluences of white and blue light, the cultivar Creso is more sensitive; 4) the insensitivity to applied GA₃ exerted by the gene Gai 1 in Creso is independent of light; 5) in Cappelli, the action of light on coleoptiles appears to be independent of the applied GA₃, whereas the hormone is able to change the pattern of growth inhibition of the first-foliage-leaf.Abbreviations BL blue light - FR far-red light - GA gibberellin - GA₃ gibberellic acid - R red light - WL white light     

Effect of IAA and 4-Cl-IAA on growth rate in maize coleoptile segments

The dose-response curves for IAA and 4-Cl-IAA-induced growth of Zea mays L. coleoptile segments were studied as a function of time. Moreover, some characteristic growth parameters for both auxins were compared. The dose-response curve of growth rate measured after IAA or 4-Cl-IAA application was bell-shaped in all experiments. The optimum concentration was 10⁻⁶ M for 4-Cl-IAA and was found not to depend on the time of the growth measurement. However, in the case of IAA the optimum shifted from 10⁻⁶ M at the time of maximal growth rate to 10⁻⁵ M or even 10⁻⁴ M, when growth measured 3–4 hours after auxin application was analysed. The relative activity of 4-Cl-IAA-induced growth rate (as compared to IAA) increased significantly with increasing time from addition of this auxin to the medium. For both auxins the time needed to reach the maximal growth rate was clearly related to their concentrations. These data provided further evidence that 4-Cl-IAA is much more active auxin than IAA and can also suggest that IAA is more rapidly metabolized in comparison to 4-Cl-IAA.     

Effect of red light on geotropism in pea epicotyls

Dose response curves were determined for phytochrome phototransformation and for a phytochrome-controlled decrease in geotropic curvature in epicotyls of dark-grown Pisum sativum L. cv. Alaska. Ten times as much light was required to produce a spectrophotometrically detectable transformation of phytochrome as was required to produce a significant change in the geotropic response. The red light energy required for a 50% phytochrome transformation caused a 90% change in the physiological response.     

Transient effects of light on auxin transport in the Avena coleoptile

The transport of indole-3-acetic acid-1-¹⁴C (IAA) through 4 mm segments of etiolated Avena coleoptiles was studied as a function of time by applying IAA in apical agar blocks and measuring the basal IAA export rate at 5-minute intervals. The transport velocity found in this way is at least 15 mm/hour at 26°. Following a 30-minute equilibration period, the export rate is nearly constant for at least 50 minutes at physiological donor concentrations. Exposure to about 3 × 10⁵ ergs/cm² blue light for 15 minutes leads to a transient reduction in the export. The export rate reaches a minimum about 25 minutes after the onset of illumination, then rises to reach a maximum by 35 minutes, and subsequently declines again. The result is a net export depression during the first 80 minutes, amounting to some 12 to 17%. Its timing closely matches the timing of the light growth response elicited by the same light dosage.

At higher IAA concentrations (0.5 and 1.8 mg/l), the export rate reaches a peak about 60 minutes after the initial application of auxin, and thereafter declines rapidly. Light increases this decline in export rate, without causing peaks and troughs, and even at 0.25 mg/l IAA the transient changes in export appear to be superimposed on a gradual decline in export rate after illumination. Blue light is effective in all these phenomena; the red far-red system appears to exert no effect. The results are discussed in relation to the mechanism of action of light both in the light growth response and in phototropism.

     

Optimization of red light-induced elongation in Avena coleoptile sections and properties of the phytochrome-mediated growth response*

Abstract Optimal conditions for studying the elongation response to a 1 mmol m^?2, 2-min pulse of red light in subapical coleoptile sections from dark-grown oat (Avena sativa L. ev. Lodi) seedlings have been determined. A technique for obtaining standard-length coleoptile sections without exposing either seedlings or sections to any light has been developed, and is described. The optimal conditions found were: sampling time, 12 h after irradiation; buffer conditions, 5 mol m^?3 potassium phosphate with 5% (w/v) sucrose (pH 5.9). The optima were determined by obtaining the time course for light-induced growth under various conditions. The red light-induced growth response is linear until 12 h after irradiation, when it undergoes an interruption. Optimal incubation conditions were determined by varying the buffer contents systematically and measuring the responses at the optimal lime determined. The results indicate a distinct difference between auxin-induced and light-induced growth responses. Even with variations of basal growth rate and several incubation conditions, the red light-induced elongation appears to be of a constant magnitude, to persist for a constant time period. and to exhibit a constant lag period between irradiation and the onset of response. The use of sections that were produced and handled in complete darkness yielded an unusual response to fusicoccin. A linear, high growth rate in response to I mmol m^?3 FC was observed for more than 12 h, both in the irradiated sections and in the dark controls.     

Stimulation of rice coleoptile growth by ethylene

Summary The growth rate of rice coleoptiles is increased by low concentrations of ethylene, especially in oxygen concentrations lower than air; carbon dioxide enhanced this response. C₂H₄ is produced by rice seedlings, and this production is also enhanced by carbon dioxide. Ethane and propane were produced in trace amounts but were inactive in growth stimulation as were also methane, propylene, and butane.This investigation was supported in part by research grants (FD-00071 and GM-12885) from the U. S. Public Health Service.     

Effect of cadmium on growth, proton extrusion and membrane potential in maize coleoptile segments

Cd accumulation, its effects on elongation growth of maize coleoptile segments, pH changes of their incubation medium and the membrane potential of parenchymal cells were studied. The Cd content increased significantly with exposure to increasing cadmium concentrations. Coleoptile segments accumulated the metal more efficiently in the range 10–100 μM Cd, than in the range 100–1000 μM Cd. Cd at concentrations higher than 1.0 μM produced a significant inhibition of both growth and proton extrusion. 100 μM Cd caused depolarization of the plasma membrane (PM) potential in parenchymal cells. The simultaneous treatment of maize coleoptile segments by indole-3-acetic acid (IAA) and Cd, counteracted the toxic effect of Cd on growth. Moreover, our data also showed that 100 μM Cd suppressed the characteristic IAA-induced hyperpolarization of the membrane potential, causing membrane depolarization. These results indicate that the toxic effect of Cd on growth of maize coleoptile segments might be, at least in part, caused via reduced PM H⁺-ATPase activity.     

Effects of red light and ethylene on growth of etiolated lettuce seedlings

Low concentrations of ethylene inhibit hypocotyl elongation of etiolated lettuce seedlings (Lactuca sativa cv. Grand Rapids), whereas red light does not inhibit it. The plumular hook tightens in response to either ethylene or red light. A combination of these two factors gives an additive response. Red light has no effect on ethylene production and red light will cause hook closure even under hypobaric pressure which removes endogenous ethylene. This suggests that ethylene and red light act independently in causing hook closure.     

Physiological mechanism of the auxin-induced increase in light sensitivity of phytochrome-mediated growth responses in Avena coleoptile sections

The physiology of the auxin-induced 10,000-fold increase in light sensitivity of a phytochrome-mediated growth response (Shinkle and Briggs, 1984 Proc Natl Acad Sci USA 81: 3742-3746) has been characterized in subapical coleoptile sections from dark-grown oat (Avena sativa L. cv Lodi) seedlings. Six micromolar indole-3-acetic acid (IAA) must be present for 1 hour before to 2 hour after irradiation in order to confer maximal sensitivity to light. The direct effect of IAA on growth can be separated from its effect on light sensitivity. Several classes of synthetic auxins will substitute for IAA in inducing an increase in sensitivity to light, as will both the phytotoxin fusicoccin and treatment of sections with pH 4.5 buffer. The increase in sensitivity to light induced by 6 micromolar IAA is completely inhibited by buffering the sections at pH 5.9 with 30 millimolar 2-(N-morpholino)ethanesulfonic acid. These findings suggest that the capacity to respond to very low fluences of light is regulated by extracellular pH.     

Dual effect of phytochrome A on hypocotyl growth under continuous red light

The role of phytochrome A in the control of hypocotyl growth under continuous red light (Rc) was investigated using phyA and phyB mutants of Arabidopsis thaliana, which lack phytochrome A (phyA) or phytochrome B (phyB), respectively, and transgenic seedlings of Nicotiana tabacum overexpressing Avena phyA, compared to the corresponding wild type (WT). In WT seedlings of A. thaliana, hypocotyl growth inhibition showed a biphasic response to the fluence rate of Rc, with a brake at 10^?2μmol m^?2 s^?1. At equal total fluence rate, hourly pulses of red light caused slightly more inhibition than Rc. The response to very low fluences of continuous or pulsed red light was absent in the phyA and phyA phyB mutants and present in the phyB mutant. The second part of the response was steeper in the phyA mutant than in the WT but was absent in the phyB mutant. In WT tobacco the response to Rc was biphasic. Overexpression of Avena phyA enhanced the response only at very low fluence rates of Rc (< 10^?2μmol m^?2 s^?1). In both species, the effect of hourly pulses of far-red light was similar to the maximum inhibition observed in the first phase of the response to Rc. Using reciprocity failure (i.e. higher inhibition under continuous than pulsed light) as the operational criterion, a ‘true’ high-irradiance reaction occurred under continuous far-red light but not under Rc or red plus far-red light mixtures. Native and overexpressed phyA are proposed to mediate very low fluence responses under Rc. In WT A. thaliana, this effect is counteracted by a negative action of phyA on phyB-mediated low-fluence responses.     

Accelerated endogenous growth in Avena coleoptile segments

Summary An unexpected acceleration in elongation of Avena coleoptile segments has been observed with a position sensor transducer 2 to 2.5 h after excision when the segments are immersed in 5 mM succinate buffer (pH 6). The cause of the accelerated growth is unknown.Paper No. 869 from the Department of Botany, Ohio State University.     

Effect of red light on the phototropic sensitivity of corn coleoptiles

The effect of red light in alteration of the phototropic sensitivity of corn coleoptiles (Zea mays L., cultivar Burpee Barbecue Hybrid) is investigated. Phototropic dosage-response curves for etiolated coleoptiles are compared with those for coleoptiles receiving 1 hour of continuous red light immediately prior to phototropic induction. In the former case, only curvature comparable to the first positive curvature of oat coleoptiles is obtained. There is no evidence for first negative curvature and only minimal second positive curvature. The reciprocity law proved valid for all curvatures obtained. With red light, the sensitivity of the first positive curvature was decreased over ten-fold and there was clear appearance of second positive curvature for which the reciprocity law was not valid. Once again there was no evidence for negative curvature. Time course studies indicated that within 1 hour of the beginning of red light treatment at 25°, reactions leading to the decrease in phototropic sensitivity of the first positive component had gone to completion whether the red light was continuous or consisted of a single 1 second exposure followed by a 1 hour dark period. An action spectrum for the red-induced change in phototropic sensitivity showed a marked peak near 660 mμ with a small broad shoulder between 610 and 630 mμ, characteristic of phytochrome-mediated responses. The effect of red light could be fully reversed by low dosages of far-red light, but longer doses of far red were less effective. Large dosages of far-red light alone induced the same alteration in phototropic sensitivity as did red light.     

The growth promoting effect of red light on internode elongation of Progress seedlings

Red light increased elongation of the apical 10 mm of epicotylsexcised from 7-day-old dark grown Progress seedlings. Removalof the basal portion of the plant appears to render the tissueinsensitive to the inhibitory influences of light. Additionof gibberellins A₁, A₃ or A₅ further increased elongation. Thered light growth response was independent of the gibberellinresponse; therefore, it was considered to be unrelated to anincrease in gibberellin biosynthesis. A study of the time course of growth in the presence of thegibberellins revealed that a 6–8 hr lag period was requiredbefore A₁ and A₅ became effective, while no lag period was associatedwith A₃. It was suggested that A₁ and A₅ were converted to anA₃-like gibberellin. (Received July 29, 1972; )