Stimulatory effect of ethephon,ACC, gibberellin A3 and A4+7 on germination of methyl jasmonate inhibited Amaranthus caudatus L. seeds

Methyl jasmonate (JA-Me) inhibited or retarded germination of Amaranthus caudatus seeds in darkness at 24°C, Ethephon, ACC and gibberellins (GA₃ or GA₄₊₇) partially or completely reversed this inhibition depending on the concentration of JA-Me applied. Both ethephon and the gibberellins were more effective than ACC. Both GA₃ and GA₄₊₇ enhanced the stimulatory effect of ethephon or ACC on germination of seeds inhibited by JA-Me.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - JA jasmonic acid - JA-Me methyl jasmonate     

Influence of ACC and Ethephon on cell growth in etiolated lupin hypocotyls. dependence on cell growth state

The possible implication of ethylene on the growth regulation of etiolated lupin hypocotyls was investigated. Excised hypocotyl sections from actively growing seedlings produced ethylene at a rate of 3 nmol h^-1 g^-1 min^-1. The rate of ethylene production was increased about 7 times when sections were treated with 10 mM 1-aminocyclopropane-1-carboxylic acid (ACC). Measurement of endogenous ACC showed that 95 % of total ACC (64.2 nmol g^-1 min^-1) corresponded to conjugated ACC. Treatments to intact seedlings with the ethylene precursor ACC, and the ethylene generating compound, 2-chloroethyl phosphonic acid (ethephon) during the cell elongation phase of the hypocotyl (from 7 to 21 dage), modified the cell growth of the organ. ACC (1 or 5 mM) or low concentrations of ethephon (0.66 mM) produced a transient decrease in the growth rate without modifying the final length of the hypocotyls. Higher concentrations of ethephon reduced the final length; the younger the seedlings were, the greater the reduction. Simultaneously to inhibition of cell elongation, ethephon produced stimulation of the radial expansion of cells in pith and cortex. The growth inhibition period, which lasted for 2 days after the treatments, was followed by another period in which the growth rate of treated plants surpassed that of the control. In both cases differences were observed along the hypocotyls due to the different growth status of the cells. It is suggested that the sensitivity to ethylene and the metabolism of ethylene depend on the growth status of the cells.     

Alleviation of Salinity-Induced Dormancy in Perennial Grasses

All seeds of Aeluropus lagopoides and Urochondra setulosa germinated under non-saline conditions except for Sporobolus ioclados which showed only 40 % germination. Increase in salinity substantially inhibited germination and few seeds germinated at 400 mM NaCl. Germination at 200 mM NaCl was alleviated in U. setulosa by the application of gibberellic acid and fusicoccin, in A. lagopoides by thiourea, betaine, kinetin, fusicoccin and ethephon, and in S. ioclados by gibberellin and ethephon. High salinity (400 mM NaCl) induced germination inhibition was alleviated by proline, kinetin, fusicoccin and ethephon only in A. lagopoides.     

Jasmonic Acid and Methyl Jasmonate in Pollens and Anthers of Three Camellia Species

Jasmonic acid (JA), which showed a nontoxic inhibitory effecton pollen germination in Camellia sinensis, was identified inpollens and anthers of C. sinensis, C. japonica and C. sasanquatogether with its methyl ester (JA-Me); the possibility thatJA is an endogenous pollen germination regulator is suggested.As JA-Me showed no effect on pollen germination, it may be formof JA inactive in pollen germination regulation. (Received April 15, 1982; Accepted June 15, 1982)     

Requirement for the action of endogenous ethylene during germination of non-dormant seeds of Amaranthus caudatus

The role of endogenous ethylene during germination of non-dormant seeds of Amaranthus caudatus L. was investigated. The seeds readily germinated in water and darkness at 24°C. Application of ethylene or of its precursor I-aminocyclopropane-I-carboxylic acid (ACC) slightly increased the rate of germination. Both compounds effectively antagonized osmotic inhibition by polyethyleneglycol. Application of aminoethoxyvinylglycine (AVG) reduced ethylene production by 90% but did not inhibit germination. However, germination was inhibited by 2,5-norbornadiene, a competitive inhibitor of ethylene action. This inhibition was counteracted by ethylene, ethephon or ACC and enforced by AVG. It is concluded that the action of endogenous ethylene is an indispensable factor during germination of non-dormant seeds of A. caudatus. Ethylene action is required from the start of imbibition on. In water, low levels of endogenous ethylene are sufficient for this action. PEG increased the ethylene requirement considerably.     

Inhibition of Amaranthus caudatus seed germination by polyethylene glycol-6000 and abscisic acid and its reversal by ethephon or 1-aminocyclopropane-1-carboxylic acid

The effect of polyethylene glycol (PEG-6000), abscisic acid (ABA), 2-chloroethylphosphonic acid (ethephon) and 1-aminocyclopropane-1-carboxylic acid (ACC) on the germination of Amaranthus caudatus L. seeds was examined. Both PEG-6000 and ABA inhibited the rate and percentage of seed germination. ABA potentiated the effect of PEG. Ethephon was highly effective in reversing the inhibitory effect of PEG and ABA or combinations of both. ACC relieved inhibition by ABA and the combined effect of ABA and PEG. Aminoethoxyvinylglycine (AVG) increased the inhibition of seed germination caused by ABA. The inhibition of seed germination by ABA seems to be related more or less to ethylene biosynthesis or is associated with a change of tissue sensitivity to ethylene. The possibility of ethylene control of water uptake by seeds is also considered.     

Germination strategy of Striga hermonthica involves regulation of ethylene biosynthesis

Ethylene involvement in germination of Striga hermonthica (Del.) Benth., an important root parasitic weed on poaceous crops, was investigated at the physiological and molecular levels. Seeds, conditioned at 30°C for 14 days, were treated with ethylene, ethephon or 1-aminocyclopropane-1-carboxylic acid (ACC). Ethylene consistently induced low germination. Ethephon and ACC effectively stimulated germination at concentrations of 0.01 and 1 m M , respectively. In contrast to ethylene, both ethephon and ACC acted in a concentration-dependent manner. Germination induced by the synthetic strigolactone GR24 was inhibited by aminoethoxyvinylglycine (AVG) and 1-methylcyclopropene. ACC reversed the inhibition caused by AVG. When seeds were treated with GR24 in sealed vials, ethylene concentration in headspace gas increased prior to the onset of germination. Total RNA extracted from germinating seeds 12 h after GR24 treatment was used for PCR-based amplification of cDNA fragments encoding the ACC synthase- and oxidase-active site domains. Two distinct cDNA fragments encoding ACC synthase ( SHACS1 and SHACS2 ) and one encoding ACC oxidase ( SHACO1 ) were cloned and sequenced. Southern analysis suggested that each of the cloned genes was present as a single copy in the genome of S. hermonthica . Northern analyses showed that SHACS1 exhibited a temporal change in expression peaking at 10 h after GR24 treatment, which coincided with a steady increase in ethylene concentration. SHACS2 was expressed at a low level with a similar trend. SHACO1 exhibited a temporal change in expression peaking at 15 days during conditioning, when seed response to GR24 was maximal. In summary, expression of ACC synthase and ACC oxidase genes was found to be responsive to a germination stimulant and to conditioning, respectively. The implications of these findings with respect to germination of S. hermonthica under field conditions are discussed.     

Reversing the inhibitory effect of paclobutrazol on seed germination of Amaranthus paniculatus by GA3, ethephon or ACC

The germination of Amaranthus paniculatus seeds was inhibited by applying paclobutrazol, a specific inhibitor of gibberellin biosynthesis. This inhibition was markedly counteracted by gibberellin A₃ (GA₃), suggesting that endogenous gibberellins are required for germination in this species. The inhibitory effect of paclobutrazol was also overcome by ethephon (2-chloroethylphosphonic acid) or the precursor of ethylene biosynthesis, ACC (1-aminocyclopropane-l-carboxylic acid). Thus the physiological effect of gibberellin can be mimicked by ethylene released from ethephon or synthesised from exogenous ACC. It is suggested, that endogenous gibberellins are involved in germination of Amaranthus paniculatus seeds and that action of GA₃ can be substituted by ethylene.Abbreviations ACC 1-aminocyclopropane-l-carboxylic acid - AMO-1618 (2-isopropyl-5methyl-4-trimethylammoniumchloride)-phenyl-l-piperidinium-carboxylate - ancymidol -cyclopropyl--(4-methoxyphenyl)-5-pyrimidine methanol - chloromequat chloride (2-chloroethyl)trimethylammoniumchloride - ethephon 2-chloroethylphosphonic acid - GA gibberellin A₃ - paclobutrazol (2RS, 3RS)-1-(4-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-lyl)pentan-3-ol - Phosphon D 2,4,dichlorobenzyl-tributhylphosphoniumchloride - tetcyclacis 5,(4-chlorophenyl)-3,4,5,9,10-pentaaza-tetracyclo)5,4,1,0,Z,6,0^8,11 dodeca-3,9-diene     

Hormonal regulation of tomato seed germination at a supraoptimal temperature

Germination of tomato cv. New Yorker seed is inhibited at 35°C. This thermoinhibition was partially counteracted by application of GA₄₊₇ alone, the compound applied in combination with ACC or ethephon markedly enhancing the process. The latter compound alone was not able to induce germination at 35 °C. Thermoinhibition of seeds at 35 °C was also counteracted by fluridone, an inhibitor of ABA biosynthesis. At 25 °C, an optimal temperature, ABA inhibited germination of New Yorker seeds. Although another known growth inhibitor MeJA, when applied at an optimal temperature (25 °C), had also a slightly inhibitory effect on germination of those seeds and clearly delayed the process, inhibitors of its biosynthetic pathway (ibuprofen, indoprofen, antypiryne and salicylic acid) did not remove thermoinhibition at 35 °C. An increase in endo-β-mannanase activity after 24 hours of incubation at 35 °C was observed in the seeds incubated in the presence of gibberellins, ACC, ethephon, fluridone used alone and in combinations, but it was not clearly correlated with the effects of these compounds on alleviation of seed germination. However, fluridone present in the same incubation medium at 35 °C with ABA was able to counteract the inhibitory effect of ABA on endo-β-mannanase activity. The results of our study suggest that gibberellins, ethylene (produced from ACC or ethephon) and ABA, but not jasmonates, regulate tomato seed germination at supraoptimal temperatures. Alleviation of thermoinhibition of New Yorker seed germination by plant growth regulators and fluridone is partially associated with their controlling endo-β-mannanase activity.     

Involvement of Ca2+ in 1-Aminocyclopropane-1-Carboxylic Acid-Stimulated Pollen Germination

The role of 1-aminocyclopropane-1-carboxylic acid (ACC) in pollen germination was investigated in several plant species. It was found that ACC stimulated in vitro pollen germination in all five species of plants tested. EGTA and phenothiazine inhibited the increase in the germination rate induced by ACC. Free Ca²⁺ levels in the cytosol ([Ca²⁺]_cyt) in ungerminated and germinated pollen were 136 and 287 nm, respectively. Adding 0.25 mm ACC to the germination medium increased the [Ca²⁺]_cyt in germinated pollen up to 450 nm. When pollen was treated with both 0.25 mm ACC and 3.6 μm inositol 1,4,5-trisphosphate, the [Ca²⁺]_cyt increased to 850 nm, and pollen germination was also stimulated. In the presence of Li⁺, an inhibitor of inositol monophosphatase, the [Ca²⁺]_cyt was reduced to 155 nm, and the ACC-stimulated pollen germination was inhibited. The data provided evidence for the involvement of Ca²⁺ as a messenger in the stimulative effect of ACC on pollen germination. Received December 1, 1995; accepted February 18, 1998     

Effects of gibberellic acid,ethephon, and 1-aminocyclopropane-1-carboxylic acid on germination ofAmaranthus caudatus seeds inhibited by paclobutrazol

The specific inhibitor of gibberellin biosynthesis, (2RS,3RS)-1-(4-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)pentan-3-ol (paclobutrazol), inhibited germination ofAmaranthus caudatus L. seeds. Addition of gibberellic acid (GA₃), 2-chloroethylphosphonic acid (ethephon), or 1-aminocyclopropane-1-carboxylic acid (ACC) effectively antagonized inhibition. Ethephon was found to be the most efficient antagonist. The transfer of seeds after 1 day's incubation in paclobutrazol to solutions of GA₃ or ethephon reversed the inhibition, the effect increasing with increasing concentration of GA₃ or ethephon. Seeds incubated in paclobutrazol for 5 days decreased sensitivity to GA₃ and ethephon.     

Effects of Jasmonic Acid and Ethephon on Tillering to Maturity in Spring Barley

Spring barley cultivated in Mitcherlich pots under both climaticchamber and field conditions were treated at developmental stageFeekes 2 with a foliar application of jasmonic acid (JA), itsmethyl ester (JAMe) and ethephon. Tillering was determined atFeekes 5 by counting all emerged tillers. The number of ear-bearingtillers, yield, and shoot length were ascertained at harvest.A combination of either JA or JAMe together with ethephon, appliedsimultaneously or successively (within 24 h), led to a significantincrease in tiller production. However, the additional tillerswere not sustained to maturity. Neither JA or JAMe nor ethephonalone were able to cause these effects at Feekes 2. JA seemsto increase the sensitivity to ethylene, possibly by influencingthe level of other native regulators. The JA/ethephon applicationat Feekes 2 did not affect the yield and the final shoot length. Spring barley, Hordeum vulgare L., tillering, apical dominance, plant growth regulators, ethephon, jasmonic acid     

In vitro maturation and germination of <Emphasis Type="Italic">Orychophragmus violaceus</Emphasis> microspores

Microspores cultured in vitro can be regarded as a system to study gene regulation, cell fate determination and cell differentiation during pollen development as well as an alternative method of genetic transformation in plants. In our study, pollen development and viability in Orychophragmus violaceus in vivo were determined and then pollen from the late unicellular stage was cultured in vitro. MS liquid medium + White vitamins + 2% (V/V) coconut milk + 0.5 M maltose, pH = 7.0 was the most appropriate for in vitro culture of Orychophragmus violaceus microspores. With this medium, the rates of in maturation and germination were 19.3% and 4.7%, respectively. Liquid medium with 0.6 M maltose + 1.6 mM boric acid + 2.9 mM Ca(NO₃)₂ + 29.6 μM vitamin B1, pH = 7.0 was optimal for germination of pollen matured in vivo. The rate of germination was 70.7%. Pollen matured in vitro cultured in similar medium exhibited a rate of germination of 62.7%. Hence, the experimental study showed that in vitro maturation of microspores is feasible and this experimental system can be applied to further theoretical and practical research.     

In vitro Arabidopsis pollen germination and characterization of the inward potassium currents in Arabidopsis pollen grain protoplasts

The focus of this study is to investigate the regulatory role of K(+) influx in Arabidopsis pollen germination and pollen tube growth. Using agar-containing media, in vitro methods for Arabidopsis pollen germination have been successfully established for the first time. The pollen germination percentage was nearly 75% and the average pollen tube length reached 135 microm after a 6 h incubation. A decrease in external K(+) concentration from 1 mM to 35 microM resulted in 30% inhibition of pollen germination and 40% inhibition of pollen tube growth. An increase in external K(+) concentration from 1 mM to 30 mM stimulated pollen tube growth but inhibited pollen germination. To study how K(+) influx is associated with pollen germination and tube growth, regulation of the inward K(+) channels in the pollen plasma membrane was investigated by conducting patch-clamp whole-cell recording with pollen protoplasts. K(+) currents were first identified in Arabidopsis pollen protoplasts. The inward K(+) currents were insensitive to changes in cytoplasmic Ca(2+) but were inhibited by a high concentration of external Ca(2+). A decrease of external Ca(2+) concentration from 10 mM (control) to 1 mM had no significant effect on the inward K(+) currents, while an increase of external Ca(2+) concentration from 10 mM to 50 mM inhibited the inward K(+) currents by 46%. Changes in external pH significantly affected the magnitude, conductance, voltage-independent maximal conductance, and activation kinetics of the inward K(+) currents. The physiological importance of potassium influx mediated by the inward K(+)-channels during Arabidopsis pollen germination and tube growth is discussed.     

Kinetin Enhanced 1-Aminocyclopropane-1-Carboxylic Acid Utilization during Alleviation of High Temperatures Stress in Lettuce Seeds

The thermoinhibition at 35 and 32°C of pregermination ethylene production and germination in lettuce (Lactuca sativa L. cv Mesa 659) seeds was synergistically or additively alleviated by 0.05 millimolar kinetin (KIN) and 10 millimolar 1-aminocyclopropane-1-carboxylic acid (ACC). The synergistic effect of KIN + ACC on ethylene production and germination at 35°C was inhibited by Co²⁺ (44-46%) but not by aminoethoxyvinyl glycine (AVG). The uptake of ACC by the seed was not influenced by KIN. Upon slitting of the seed coats (composed of pericarp, testa and endosperm), following the uptake of chemicals, ACC was readily converted into ethylene at all temperatures, and the synergistic effects of KIN + ACC at 35°C were lost. At 35°C, KIN acted synergistically with ACC or ethephon (ETH) in alleviating the osmotic restraint. At 25°C, ETH was more active than KIN or KIN + ACC in overcoming the osmotic restraint. Thus, the integrity of the seed coats, the KIN-enhanced ACC utilization, and an interaction of KIN with the ethylene produced may be the basis for the synergistic or additive effects of KIN + ACC at high temperature.     

Ethylene in seed dormancy and germination

The role of ethylene in the release of primary and secondary dormancy and the germination of non-dormant seeds under normal and stressed conditions is considered. In many species, exogenous ethylene, or ethephon – an ethylene-releasing compound - stimulates seed germination that may be inhibited because of embryo or coat dormancy, adverse environmental conditions or inhibitors (e.g. abscisic acid, jasmonate). Ethylene can either act alone, or synergistically or additively with other factors. The immediate precursor of ethylene biosynthesis, 1-aminocyclopropane-1-carboxylic acid (ACC), may also improve seed germination, but usually less effectively. Dormant or non-dormant inhibited seeds have a lower ethylene production ability, and ACC and ACC oxidase activity than non-dormant, uninhibited seeds. Aminoethoxyvinyl-glycine (AVG) partially or markedly inhibits ethylene biosynthesis in dormant or non-dormant seeds, but does not affect seed germination. Ethylene binding is required in seeds of many species for dormancy release or germination under optimal or adverse conditions. There are examples where induction of seed germination by some stimulators requires ethylene action. However, the mechanism of ethylene action is almost unknown.
The evidence presented here shows that ethylene performs a relatively vital role in dormancy release and seed germination of most plant species studied.     

Role of ethylene in the control of gametophyte-sporophyte interactions in the course of the progamic phase of fertilization

We investigated dynamics of the content of 1-aminocyclopropane-1-carboxylic acid (ACC) and ethylene production in male gametophyte development and germination in fertile (self-compatible and selfincompatible) and sterile clones of petunia. Fertile male gametophyte development was accompanied by two peaks of ethylene production by anther tissues. The first peak occurred during the microspore development simultaneously with the degeneration of both the tapetal tissues and the middle layers of the anther wall. The second peak coincided with dehydration and maturation of pollen grains. In the anther tissues of the sterile line of petunia, tenfold higher ethylene production was observed at the meiosis stage compared with that in fertile male gametophytes. This fact correlated with the degeneration of both microsporocytes and tapetal tissues. Exogenously applied ethylene (1–100 ppm) induced a degradation of the gametophytic generation at the meiosis stage. According to the obtained data, ethylene synthesis in germinating male gametophyte is provided by a 100-fold ACC accumulation in mature pollen grains. The male gametophyte germination, both in vitro, on the culture medium, and in vivo, on the stigma surface, was accompanied by an increase in ethylene production. Depending on the type of pollination, germination of pollen on the stigma surface and the pollen tube growth in the tissues of style were accompanied by various levels of ACC and ethylene release. The male gametophyte germination after self-compatible pollination was accompanied by higher content of ACC as compared with the self-incompatible clone, whereas, after the self-incompatible pollination, we observed a higher level of ethylene production compared with compatible pollination. For both types of pollination, ACC and ethylene were predominantly produced in the stigma tissues. Inhibitor of ethylene action, 2,5-norbornadiene (NBN), blocked both the development and germination of the male gametophyte. These results suggest that ethylene is an important factor in male gametophyte development, germination, and growth at the progamic phase of fertilization.     

Influence of growth retardants on the internode and ethylene production of sunflower plants

The growth-retarding activity of the norbornenodiazetine tetcyclacis and the di-oxanylalkenyl triazole LAB 150 978 as well as the ethylene-forming compounds 2-chloroethyl-phosphonic acid (ethephon) and 1-amino-cyclopropane-l-carboxylic acid (ACC) on stem histogenesis and ethylene production of sunflower plants ( He-lianthus annuus L. cv.Spanners Allzweck) has been studied. The shoot growth of plants hydroponically grown and treated was reduced by the compounds. The shortening in the length of the 1st internode caused by tetcyclacis and LAB 150 978 was mainly induced by inhibition of cell division (the internode possessed fewer cortical cells per cell file). In contrast, ethephon and ACC decreased internode elongation mainly by reducing the rate of cell enlargement.
The ethylene production of sunflower seedlings cultivated on agar nutrient medium rose with increasing concentrations of ethephon and ACC, the shoot length of the plants being progressively reduced.
Tetcyclacis and LAB 150 978 inhibited both the formation of ethylene and shoot growth. It is suggested that in contrast to ethephon and ACC, tetcyclacis and LAB 150 978 do not achieve their growth-retarding effect by influencing the production of ethylene.     

Bis(guanylhydrazones) negatively affect in vitro germination of kiwifruit pollen and alter the endogenous polyamine pool

Bis(guanylhydrazones) are a class of compounds known to interfere with the metabolism of polyamines (PAs). Among them, the methylglyoxal derivative (MGBG) has been studied most thoroughly. Because PAs and their biosynthetic enzymes are strongly involved in pollen tube organization, emergence and elongation, a number of these inhibitors have been studied in the present work for their effects on the in vitro performance of kiwifruit (Actinidia deliciosa) pollen. Increasing concentrations of several bis(guanylhydrazones) in the range 0.05-1 mM were checked for their effect on pollen germination. Most of the compounds tested showed a dose-dependent inhibitory effect on tube emergence, which was established very early during incubation. At 0.5 mM, the methylpropylglyoxal derivative (MPGBG) had a stronger inhibitory effect than MGBG. To verify whether the inhibitors reached their metabolic target, PA levels and S-adenosylmethionine decarboxylase (SAMDC) activity were determined in pollen germinated in the presence or absence (controls) of 0.5 mM bis(guanylhydrazones). Spermidine (Spd) content was significantly reduced in the treated pollen, and this effect was more pronounced after treatment with MGBG than with MPGBG. An early and strong reduction in SAMDC activity was observed after exposure to either inhibitor. Inhibition of pollen germination by MGBG or MPGBG could not be reversed by the addition of exogenous Spd, which per se was inhibitory. Taken together, our results suggest that bis(guanylhydrazones) alter PA metabolism and negatively affect kiwifruit pollen germination, even though a strict cause-effect relationship could not be established, and other mechanisms, unrelated to PA activity, must be involved.