Exogenous Jasmonic and Abscisic Acids Act Differentially in Elongating Tissues from Oat Stem Segments

Segments can be cut from the peducular-1 internode of oat (Avena sativa L.) shoots so as to contain the graviresponsive, auxin-sensitive leaf sheath pulvinus, and the gibberellin-sensitive internodal tissue. These two growth-capable tissues were used to study the effects and interactions of jasmonic acid (JA) and abscisic acid (ABA) in regulating cell elongation. When supplied alone at physiologic concentrations (10⁻⁵, 10⁻⁴ m), JA promoted growth and cell wall synthesis in the internodal tissue, whereas by itself, ABA inhibited internodal elongation and even inhibited JA-promoted growth. When gibberellic acid (GA₃) was used to stimulate internodal elongation, JA and ABA caused similar levels of inhibition and, at certain concentrations, were synergistic. Inhibition by ABA was initiated several hours earlier than inhibition by JA, and only the ABA effect could be partially overcome by 10⁻³ m aminoethoxyvinylglycine. Both JA and ABA inhibited elongation of pulvinar tissue that was induced to grow by gravistimulus or auxin, although here JA was more potent than ABA at equimolar concentrations. When 10⁻⁵ m fusicoccin was used as a general nonphysiologic growth stimulus, JA had no effect on the internode but inhibited the pulvinus, whereas ABA had no effect on the pulvinus but inhibited the internode. These results provide strong physiologic evidence that JA and ABA act by different mechanisms in the regulation of elongation, at least in this representative grass. Received May 28, 1996; accepted November 7, 1996     

The Role of Abscisic Acid in Induction of Androgenesis: A Comparative Study Between Hordeum vulgare L. Cvs. Igri and Digger

Under the same mannitol pretreatment and culture conditions, regeneration efficiency in the barley cultivar (cv.) Igri was about 10 times higher than in the cv. Digger, a difference only partially reflected by a difference in viable microspores after anther pretreatment. Therefore, a comparative study between cvs. Igri and Digger was carried out under various pretreatment conditions. For both cultivars, under water, CPW buffer and mannitol pretreatment conditions, there was a positive correlation between microspore viability and regeneration efficiency in that mannitol > CPW buffer >> water. Mannitol pretreatment of cv. Igri produced a much higher endogenous abscisic acid (ABA) level than as to Digger. Addition of ABA stimulated both percentages of viability and regeneration efficiency except in the case of mannitol pretreatment. Under CPW buffer pretreatment conditions, addition of ABA significantly stimulated regeneration efficiency and was ABA concentration dependent. However, cv. Digger was less responsive to ABA than cv. Igri. In both cultivars, under less optimal pretreatment conditions (e.g., water and CPW buffer), the effect of ABA was to stimulate increased percentages of viability and/or to reduce the number of binucleate microspores. Moreover, in cv. Igri, direct culture of anthers for 4 days without pretreatment caused an increased number of binucleate microspores compared with microspores with pretreatment for 4 days. These binucleate microspores showed DNA degradation in the nuclei. However, with mannitol pretreatment binucleate microspores and DNA fragmentation in the nuclei of microspores was rarely observed. On the basis of our observations, we suggest that the difference in regeneration efficiency in cv. Igri and cv. Digger is related to the differences in endogenous ABA production levels under mannitol pretreatment and responsiveness to ABA. One of the effects of ABA is likely due to an inhibition of cell death. Received May 21, 1999; accepted October 5, 1999     

Involvement of Abscisic Acid in Mesocotyl Growth in Etiolated Seedlings of a Foxtail Millet Dwarf Mutant

Etiolated seedlings of foxtail millet (Setaria italica Beauv.) dwarf mutant CH84113 were treated with various concentrations of abscisic acid (ABA), mefluidide, mannitol, or polyethylene glycol (PEG) 6000. It was found that these chemicals, at suitable concentrations, could increase mesocotyl length significantly, whereas these chemicals at higher concentrations had an inhibitory effect. Endogenous levels of ABA in mesocotyl were measured by enzyme-linked immunosorbent assay. It was found that endogenous ABA increased progressively in a chemical (ABA, mefluidide, mannitol, or PEG 6000) concentration-dependent manner, indicating that the effects of these chemicals on mesocotyl growth may be mediated by increased endogenous ABA levels. On the other hand, S-3307, an inhibitor of the oxidative reactions in gibberellin (GA) biosynthesis, inhibited the elongation of mesocotyl significantly. When ABA and GA₃ were applied simultaneously, the effect on mesocotyl growth was additive. These results imply that ABA and GA may control different processes in the regulation of mesocotyl growth. Received October 27, 1997; accepted May 11, 1998     

Role of the Ring Methyl Groups in Abscisic Acid Activity in Erucic Acid Accumulation in Oilseed Rape (Brassica napus L.)

Modification of the structure of abscisic acid (ABA) has been reported to result in modification of its physiologic activity. In this study we tested the effect of removing methyl groups from the ring and of chirality of ABA on activity in microspore-derived embryos of oilseed rape (Brassica napus L.). The natural (+)-ABA molecule induced growth inhibition and an increase in the amount of erucic acid accumulated in the oil at medium concentrations less than 1 μm. (−)-ABA showed similar effects. Removing the 7′-methyl group resulted in a dramatic decrease in activity: (+)-7′-demethyl-ABA retained some activity as a growth inhibitor; a 10–100 μm concentration of this compound was needed for a response, and (−)-7′-demethyl-ABA was almost completely inactive. Similar effects were observed with regard to elongase activity, which catalyzes erucic acid biosynthesis from oleic acid. Removal of the 8′- and 9′-methyl groups resulted in a more complex response. These compounds all showed intermediate activity; for growth inhibition, the presence of the 9′-methyl was the more important determinant, whereas chirality dominated the response on erucic acid accumulation, with the (+)-enantiomers being more active. Received July 25, 1997; accepted October 31, 1997     

Effects of Exogenously Applied Jasmonates on Growth and Intracellular pH in Maize Coleoptile Segments

The effects of jasmonic acid (JA) on elongation growth of coleoptile segments from etiolated maize (Zea mays L.) were investigated in the presence and absence of auxin. When supplied alone, at physiological concentrations (10⁻⁹, 10⁻⁸, and 10⁻⁵ m), JA (or methyl-JA) inhibited growth. JA at a similar range of concentrations also inhibited auxin-induced elongation growth. To determine whether this effect on growth depended on endogenous abscisic acid (ABA), we grew maize coleoptiles in the presence of norflurazon (an inhibitor of carotenoid biosynthesis) that results in reduced endogenous ABA levels. Growth of etiolated coleoptile segments from these plants was inhibited by JA (or methyl-JA) in both the absence and presence of auxin. Previously, we have observed a correlation between elongation growth and cytosolic pH (pH_i), in which auxin lowers pH_i, and growth inhibitors such as ABA raise pH_i. We examined the effect of low concentrations of methyl-JA on pH_i with dual emission dye, carboxy seminaphthorhodafluor-1, and confocal microscopy. To confirm these studies, we also used in vivo ³¹P NMR spectrometry to ascertain the changes in pH_i after addition of jasmonate to maize coleoptiles. Coleoptiles grown in either the absence or presence of norflurazon responded to methyl-JA or JA by increases in pH_i of approximately 0.2 pH unit. This response occurs over a period of 15–20 min and appears to be independent of endogenous ABA. This alkalization induced by JA is likely to form a permissive environment for JA signal transduction pathway(s). Received February 5, 1999; accepted August 25, 1999     

New Plant Growth Regulators Protect Photosynthesis and Enhance Growth Under Drought of Jack Pine Seedlings

To determine whether natural plant growth regulators (PGRs) can enhance drought tolerance and the competitive ability of transplanted seedlings, 1.5-year-old jack pine (Pinus banksana Lamb.) seedlings were treated with homobrassinolide, salicylic acid, and two polyamines, spermine and spermidine, triacontanol, abscisic acid (ABA), and the synthetic antioxidant, Ambiol. PGRs were fed into the xylem for 7 days and plants were droughted by withholding water for 12 days. ABA, Ambiol, spermidine, and spermine at a concentration of 10 μg L⁻¹ stimulated elongation growth under drought, whereas ABA, Ambiol, and spermidine maintained higher photosynthetic rates, higher water use efficiency, and lower Ci/Ca ratio under drought compared with control plants. The damaging effects of drought on membrane leakage was reversed by Ambiol, ABA, triacontanol, spermidine, and spermine. Because ABA, Ambiol, and both polyamines enhanced elongation growth and also reduced membrane damage in jack pine under drought, they show promise as treatments to harden seedlings against environmental stress. The protective action of these compounds on membrane integrity was associated with an inhibition of ethylene evolution, with a reduction in transpiration rate and an enhancement of photosynthesis, which together increased water use efficiency under drought. Although most of the tested compounds acted as antitranspirants, the inhibition in membrane leakage in ABA-, Ambiol-, and polyamine-treated plants appeared more closely related to the antiethylene action. Received December 30, 1998; accepted October 14, 1999     

Potential Role of Abscisic Acid in Cotton Fiber and Ovule Development

Fibers and ovules of a cotton cultivar (Gossypium hirsutum L. Trambak-108) were analyzed for growth and free abscisic acid (ABA) content by indirect enzyme immunoassay. An inverse correlation between fiber elongation and ABA content was observed. In the seed, accumulation of ABA was observed during secondary thickening and the maturation phase. The potential role of ABA in fiber and seed development is discussed. Received June 25, 1997; accepted October 15, 1997     

Involvement of Abscisic Acid and Indole-3-acetic Acid in the Flowering of Pharbitis nil

The involvement of abscisic acid (ABA) and indole-3-acetic acid (IAA) in the regulation of flowering of Pharbitis nil was investigated through exogenous applications and analyses of endogenous levels. Both hormones inhibited the flowering of P. nil when they were applied before or after a single 15-h dark treatment. The inhibitory effect of ABA and IAA was significant when they were applied before the dark treatment, and the application to plumules was more effective than that to cotyledons. In all applications, the inhibitory effect of IAA was stronger than that of ABA. Endogenous levels of ABA and IAA in the plumules were compared between flower-inductive (15-h dark treatment) and noninductive (continuous light) light conditions. There was no significant difference in the ABA level between light and dark conditions, whereas the level of IAA was decreased by the dark treatment. These results suggest that biosynthesis and/or catabolism of IAA is affected by the light treatment and therefore may be involved in the regulation of early flowering processes in the apex. The inhibitory effects of ABA and IAA were reversed by an application of gibberellin A₃, indicating that gibberellin A₃ counteracts the flowering processes affected by ABA and IAA. Application of aminoethoxyvinylglycine restored the flowering response inhibited by IAA, which suggests the possibility that the inhibitory effect of IAA is the result of enhanced ethylene biosynthesis. Received November 22, 1996; accepted February 17, 1997     

Influence of 2,3,5-Triiodobenzoic Acid and 1-N-Naphthylphthalamic Acid on Indoleacetic Acid Transport in Carnation Cuttings: Relationship with Rooting

³H-IAA transport in excised sections of carnation cuttings was studied by using two receiver systems for recovery of transported radioactivity: agar blocks (A) and wells containing a buffer solution (B). When receivers were periodically renewed, transport continued for up to 8 h and ceased before 24 h. If receivers were not renewed, IAA transport decreased drastically due to immobilization in the base of the sections. TIBA was as effective as NPA in inhibiting the basipetal transport irrespective of the application site (the basal or the apical side of sections). The polarity of IAA transport was determined by measuring the polar ratio (basipetal/acropetal) and the inhibition caused by TIBA or NPA. The polar ratio varied with receiver, whereas the inhibition by TIBA or NPA was similar. Distribution of immobilized radioactivity along the sections after a transport period of 24 h showed that the application of TIBA to the apical side or NPA to the basal side of sections, increased the radioactivity in zones further from the application site, which agrees with a basipetal and acropetal movement of TIBA and NPA, respectively. The existence of a slow acropetal movement of the inhibitor was confirmed by using ³H-NPA. From the results obtained, a methodological approach is proposed to measure the variations in polar auxin transport. This method was used to investigate whether the variations in rooting observed during the cold storage of cuttings might be related to changes in polar auxin transport. As the storage period increased, a decrease in intensity and polarity of auxin transport occurred, which was accompanied by a delay in the formation and growth of adventitious roots, confirming the involvement of polar auxin transport in supplying the auxin for rooting. Received April 19, 1999; accepted December 2, 1999     

Polyamines in Helianthus annuus L. during Germination under Salt Stress

The level of the three main polyamines putrescine, spermidine, and spermine and the biosynthetic enzyme arginine decarboxylase (ADC) decreased in Helianthus annuus L. seedlings subjected to increasing (50, 100, and 150 mm) NaCl concentrations. The pattern of polyamines in control plants increased during the initial 72 h and then reached a plateau. The putrescine level showed an increase of 370% after 72 h of development. The lower salt treatment slightly diminished the overall polyamine content. The highest NaCl concentration (150 mm) induced a strong putrescine diminution (from 381 to 78.9 nmol g⁻¹ FW) at 72 h whereas a small decrease in ADC activity was detected. ODC was detected in neither control nor treated plantlets during the experimental period. The level of spermidine also decreased, but the magnitude of the decay was less pronounced than putrescine. The fact that ODC was not detected and ADC activity followed a pattern similar to that of putrescine led us to suppose that the variation in putrescine content could be attributed entirely to the decrease in ADC activity. α-Difluoromethylarginine and α-difluoromethylornithine (ADC and ODC inhibitor, respectively) did not inhibit but delayed the onset of germination of sunflower seeds, and α-difluoromethylornithine increased the content of spermidine and spermine. The present data suggest that polyamines could be involved in the germination process of H. annuus seeds and in response to salt stress. Received April 14, 1997; accepted July 10, 1997     

Gibberellic Acid Treatment Reduces the Tolerance of Field-Grown Common Bean to Leaf Removal

I studied the influence of gibberellic acid (GA₃) treatment in a field population of common bean on plant tolerance to leaf removal. Individual bean seedlings were treated with a foliar application of 10 μM GA₃ on day 7 and day 14 after emergence, which led to a significant increase in height in GA₃-treated plants. Twenty-eight days after emergence, either zero, one, two, or three leaflets from each trifoliate leaf were removed from each of 20 GA₃-treated and 20 control plants. All pods were harvested from each plant after plants became senescent 6 weeks later. Multivariate analyses revealed that leaf removal produced significant reductions in several yield components in both GA₃-treated and control plants, although the effects were not pronounced until at least two leaflets from each trifoliate leaf (67% of the total leaf area) were removed. However, GA₃-treated plants suffered greater reductions in total pod wall mass and total seed number than control plants after 33 and 67% leaf area removal. These results indicate that GA₃ treatment may have altered the assimilatory capacity or resource allocation pattern of treated plants in such a way as to decrease their ability to tolerate leaf removal, a negative consequence of the hormonal alteration of traits important to plant compensation for biotic stressors. Received December 6, 1996; accepted March 5, 1997     

Screening for Barley yellow dwarf virus-Resistant Barley Genotypes by Assessment of Virus Content in Inoculated Seedlings

The content of Barley yellow dwarf virus (BYDV) in roots and leaves of barley seedling plants differing in their level of resistance was assessed by quantitative ELISA 1–42 days after inoculation with the strain of BYDV (PAV). High virus accumulation in roots and low concentration in leaves was characteristic of the period 9–15 days after inoculation. In leaves, the differences in virus content between resistant and susceptible genotypes became significant after 15 days and resistance to virus accumulation was better expressed 30–39 days after inoculation. Roots of resistant materials exhibited evident retardation of virus accumulation and the greatest difference in virus content between resistant and susceptible plants was detected 9 days after inoculation. By these criteria, the selected winter and spring barley cultivars and lines (in total 44 materials) fell in to five groups according to field reactions and the presence or absence of the Yd2 resistance gene. There were highly significant and positive relations between ELISA values and 5‐year field data on symptomatic reactions and grain‐yield reductions due to infection. Using the described method, resistant and moderately resistant genotypes (both Yd2 and non‐Yd2) were significantly differentiated from susceptible genotypes. The possible use of this method in screening for BYDV resistance is discussed.     

Lepidimoide Promotes Light-Induced Chlorophyll Accumulation in Cotyledons of Sunflower Seedlings

The effect of disaccharide lepidimoide on light-induced chlorophyll accumulation was studied in cotyledons of sunflower (Helianthus annuus L.) seedlings and detached cucumber (Cucumis sativus L.) cotyledons. From studies on the structure-activity relationships of lepidimoide, its analogs, and sucrose with respect to light-induced chlorophyll accumulation in the cotyledons of sunflower seedlings, both lepidimoide and the free carboxylic acid of lepidimoide (lepidimoic acid) showed the highest promoting activity, whereas the hydrogenated lepidimoide, which lacks a double bond in the C4, 5 position in uronic acid, showed lower activity than lepidimoide; however, sucrose exhibited very weak activity. These results suggest that lepidimoide acts as a new type of plant growth regulator, not simply as a carbon source providing energy. Lepidimoide promoted not only light-induced chlorophyll accumulation in sunflower cotyledons but also light-induced 5-aminolevulinic acid content, which is considered to be a rate-limiting step in chlorophyll biosynthesis. Lepidimoide with cytokinin stimulated the accumulation of chlorophyll and 5-aminolevulinic acid additively. In detached cucumber cotyledons, lepidimoide also promoted light-induced chlorophyll accumulation. These results indicate that lepidimoide, in cooperation with cytokinin, causes light-induced chlorophyll accumulation in the cotyledons of several dicot plant species by affecting the level of 5-aminolevulinic acid. Received April 4, 1997; accepted September 28, 1998     

Possible Involvement of Altered Gibberellin Metabolism in the Induction of Tomato Irregular Ripening in Dwarf Cherry Tomato by Silverleaf Whitefly

External and internal tomato irregular ripening (TIR) symptoms were associated with the feeding of silverleaf whitefly (SLW), Bemisia argentifolii Bellows and Perring. Four experiments consisting of various soil drench applications of GA₃ (100 ppm) and cycocel (CCC, an inhibitor of GA biosynthesis; 1,000 and 2,000 ppm) were applied to dwarf cherry tomato cv. Florida Petite in the presence and absence of SLW in an attempt to mimic the disorders induced by the SLW. The application of GA₃ induced external and internal TIR symptoms similar to the SLW-induced disorder. Minimal TIR-like symptoms also occurred in the control and CCC treatments. Internal TIR symptoms in GA₃, GA₃ plus SLW, and GA₃ plus CCC treatments ranged from 66% to 97% throughout the experiments. The incidence of external TIR symptoms was highest in the GA₃ plus SLW treatment compared with the other treatments. CCC reduced the incidence of external TIR symptoms induced by GA₃ or GA₃ plus SLW treatments. However, CCC-treated plants also attracted more oviposition and higher populations of SLW and consequently induced a greater incidence of TIR symptoms than SLW treatment alone. Furthermore, although low SLW populations may be associated with low external TIR symptoms, internal TIR symptoms almost always remained high in infested plants. The results suggest that the TIR disorder in dwarf cherry tomato which is induced by the SLW may be a gibberellin-regulated disorder. Received May 27, 1997; accepted September 26, 1997     

Effect of Pot Size and Timing of Plant Growth Regulator Treatments on Growth and Tuber Yield in Greenhouse-Grown Norland and Russet Burbank Potatoes

The effects of pot size, timing of the application of paclobutrazol (PTZ) and gibberellic acid (GA₃), and the counteractive effect of these two compounds on growth and tuber yield of greenhouse-grown Norland and Russet Burbank potatoes were investigated. Plants were grown either in 1.5-liter pots (15 cm deep) or 3.0-liter pots (18 cm deep) and received a foliar application of either 1.5 mm PTZ or 9 × 10⁻³ mm GA₃ at early or late stolon initiation. Some plants that had been foliar treated with 1.5 mm PTZ at early stolon initiation were foliar treated with 9 × 10⁻³ mm GA₃ at late stolon initiation. PTZ reduced haulm length in both cultivars significantly, particularly when the treatment was applied at early stolon initiation, but the late treatment reduced haulm length only when growing in 3.0-liter pots. Irrespective of the timing of treatment, GA₃ increased haulm length in Norland growing in both pot sizes, but the treatment increased haulm length in Russet Burbank only when applied at late stolon initiation. GA₃ applied after PTZ did not overcome the growth-inhibiting effect of the PTZ treatment. The PTZ treatment effectively increased usable tuber number/plant (UTN) in Norland, but PTZ had no effect on UTN in Russet Burbank. PTZ reduced usable tuber weight/plant (UTW) only in Norland growing in 1.5-liter pots. By contrast, GA₃ increased UTN only when treated at late stolon initiation of 1.5-liter pot-grown Norland, whereas the same treatment was effective when applied only at early stolon initiation for Russet Burbank. For Norland, the increase in UTN by early applied PTZ was reduced by the subsequent application of GA₃. The use of 3.0-liter pots for minituber production in both Norland and Russet Burbank appears to have no advantage over growing in 1.5-liter pots, particularly when PTZ or GA₃ is used to enhance tuberization. Received May 30, 1997; accepted February 3, 1998     

Uptake, Distribution, and Metabolism of 1-Naphthaleneacetic Acid and Indole-3-Acetic Acid During Callus Initiation From Actinidia deliciosa Tissues

1-Naphthaleneacetic acid (NAA) and 6-benzyladenine (BA) were required for in vitro callus formation at the basal edge of kiwifruit (Actinidia deliciosa [A. Chev] Liang and Ferguson, cv. Hayward) petioles. The uptake, metabolism, and concentration of NAA and indole-3-acetic acid (IAA) content were examined in the explants during the callus initiation period. After 1, 6, 12, 24, 48, and 96 h of culture in the presence of [H³]NAA, petioles were divided into apical, middle, and basal portions and analyzed. Except for a high IAA level measured at 12 h, IAA content decreased in tissues during a culture period of 96 h. NAA uptake was higher in petiolar edges than in the middle portion, and NAA was rapidly conjugated with sugars and aspartic acid inside the tissues. The amide conjugation was triggered in apical and basal portions from 12 h and in the middle part from 48 h, with α-naphthylacetylaspartic acid being the major metabolite. Free-NAA concentration in cultured petioles achieved an equilibrium with the exogenously applied NAA (0.27 μm) from 12 h, and it remained constant thereafter. The relationships between the role attributed to NAA and BA in the initiation and the maintenance of disorganized growth of callus in kiwifruit cultures are discussed. Received December 21, 1998; accepted July 20, 1999     

Drought Stress Syndrome in Wheat Is Provoked by Ethylene Evolution Imbalance and Reversed by Rewatering, Aminoethoxyvinylglycine, or Sodium Benzoate

In this work we present evidence that the drought stress syndrome in the flag leaves and ears of wheat plants, provoked by the production of ethylene (shortening the grain filling period and lowering the grain weight) is reversed by the application of a free radical scavenger, sodium benzoate or the ethylene synthesis inhibitor, aminoethoxyvinylglycine. Rehydration by watering also attenuated the detrimental effect of the water deficit. Consequently, the grain filling period was longer, the grain weight increased, and the total protein content was higher than that in plants watered regularly. Received June 11, 1998; accepted March 10, 1999