Inhibition of ethylene biosynthesis by aminoethoxyvinylglycine and by polyamines shunts label from 3,4-[C]methionine into spermidine in aged orange peel discs

The flux of radioactivity from 3,4-[(14)C]methionine into S-adenosyl-l-methionine (SAM), 1-aminocyclopropane-1-carboxylic acid (ACC), spermine, and spermidine while inhibiting conversion of ACC to ethylene by 100 millimolar phosphate and 2 millimolar Co(2+) was studied in aged peel discs of orange (Citrus sinensis L. Osbeck) fruit. Inhibition up to 80% of ethylene production by phosphate and cobalt was accompanied by a 3.3 times increase of label in ACC while the radioactivity in SAM was only slightly reduced. Aminoethoxyvinylglycine (AVG) increased the label in SAM by 61% and reduced it in ACC by 47%. Different combinations of standard solution, in which putrescine or spermidine were administered alone or with AVG, demonstrated clearly that inhibition of ethylene biosynthesis-at the conversion of SAM to ACC-by AVG, exogenous putrescine or exogenous spermidine, stimulated the incorporation of 3,4-[(14)C]methionine into spermidine.     

Auxin-induced Ethylene Production and Its Inhibition by Aminoethyoxyvinylglycine and Cobalt Ion

Auxin is known to stimulate greatly both C₂H₄ production and the conversion of methionine to ethylene in vegetative tissues, while amino-ethoxyvinylglycine (AVG) or Co²⁺ ion effectively block these processes. To identify the step in the ethylene biosynthetic pathway at which indoleacetic acid (IAA) and AVG exert their effects, [3-¹⁴C]methionine was administered to IAA or IAA-plus-AVG-treated mung bean hypocotyls, and the conversion of methionine to S-adenosylmethionine (SAM), 1-amino-cyclopropane-1-carboxylic acid (ACC), and C₂H₄ was studied. The conversion of methionine to SAM was unaffected by treatment with IAA or IAA plus AVG, but active conversion of methionine to ACC was found only in tissues which were treated with IAA and which were actively producing ethylene. AVG treatment abolished both the conversion of methionine to ACC and ethylene production. These results suggest that in the ethylene biosynthetic pathway (methionine → SAM → ACC → C₂H₄) IAA stimulates C₂H₄ production by inducing the synthesis or activation of ACC synthase, which catalyzes the conversion of SAM to ACC. Indeed, ACC synthase activity was detected only in IAA-treated tissues and its activity was completely inhibited by AVG. This conclusion was supported by the observation that endogenous ACC accumulated after IAA treatment, and that this accumulation was completely eliminated by AVG treatment. The characteristics of Co²⁺ inhibition of IAA-dependent and ACC-dependent ethylene production were similar. The data indicate that Co²⁺ exerts its effect by inhibiting the conversion of ACC to ethylene. This conclusion was further supported by the observation that when Co²⁺ was administered to IAA-treated tissues, endogenous ACC accumulated while ethylene production declined.     

Effects of 1-Aminocyclopropane-1-carboxylic Acid Production on the Changes in the Polyamine Levels in Hiproly Barley Callus after Auxin Withdrawal

Contents of polyamines and 1-aminocyclopropane-1-carboxylic acid (ACC) in Hiproly barley callus were examined under different culture conditions. After auxin withdrawal, the contents of free polyamines changed conversely to the contents of ACC. In the absence of auxin, incorporation of l-[3,4–¹⁴C]methionine into polyamines and the activity of S-adenosylmethionine decarboxylase (SAMDCase) in the callus increased, then remained stable, but incorporation of l-[3,4- ¹⁴C]methionine into ACC, precursor of ethylene and ACC synthase activity once declined and increased again.

Aminooxyacetic acid (AOA) affected the increase in the levels of polyamines in the callus. 1- Aminoisobutyric acid (AIB) had a slight effect on the polyamine production. The incorporation of l-[3,4–¹⁴C]methionine into ACC and ACC synthase activity were inhibited by AOA, but not by « 4 AIB. AOA stimulated the activity of SAMDCase, and also enhanced the incorporation of l-[3,4- ¹⁴C]methionine into polyamines in the callus. Methylglyoxal-bis(guanylhydrazone) (MGBG) greatly enhanced the ACC production. The rate of incorporation of l-[3,4–¹⁴C]methionine into ACC and ACC synthase activity in the callus were significantly enhanced by MGBG. MGBG strongly inhibited SAMDCase activity and the incorporation of l-[3,4–¹⁴C]methionine into polyamines. Moreover, the synthesis of polyamines was inhibited by MGBG.

These results suggested that in Hiproly barley callus ACC production has an important effect on changes in the polyamine levels, and that polyamine and ethylene biosynthetic pathways are regulated by competition against each other.     

Carbohydrates Stimulate Ethylene Production in Tobacco Leaf Discs : II. Sites of Stimulation in the Ethylene Biosynthesis Pathway

Galactose, sucrose, and glucose (50 millimolar) applied to tobacco leaf discs (Nicotiana tabacum L. cv `Xanthi') during a prolonged incubation (5-6 d) markedly stimulated ethylene production which, in turn, could be inhibited by aminoethoxyvinylglycine (2-amino-4-(2′-aminoethoxy)-trans-3-butenoic acid) (AVG) or Co²⁺ ions. These three tested sugars also stimulated the conversion of l-[3,4-¹⁴C]methionine to [¹⁴C]1-amino-cyclopropane-1-carboxylic acid (ACC) and to [¹⁴C]ethylene, thus indicating that the carbohydrates-stimulated ethylene production proceeds from methionine via the ACC pathway. Sucrose concentrations above 25 mm considerably enhanced ACC-dependent ethylene production, and this enhancement was related to the increased respiratory carbon dioxide. However, sucrose by itself could directly promote the step of ACC conversion to ethylene, since low sucrose concentrations (1-25 mm) enhanced ACC-dependent ethylene production also in the presence of 15% CO₂.     

Daminozide inhibits ethylene production in apple fruit by blocking the conversion of methionine to aminocyclopropane-1-carboxylic acid (ACC)

At harvest, fruit from apple trees sprayed with daminozide (+daminozide) had lower levels of aminocyclopropane-1-carboxylic acid (ACC) and produced significantly lower amounts of ethylene than untreated (–daminozide) fruit. Flesh discs from the fruit of +daminozide and –daminozide trees were fed precursors of ethylene to determine how daminozide inhibits ethylene production. ACC was metabolized to ethylene regardless of treatment. Methionine (MET), however, was only converted to ethylene by –daminozide fruit, and only after the fruit had been maintained at 4 °C for 5 months. +Daminozide fruit failed to convert MET to ethylene at harvest, as well as after cold storage. When daminozide was added to the incubation media of flesh discs it did not inhibit ethylene production or the conversion of ACC to ethylene. The addition of daminozide did, however, inhibit the metabolism of exogenous MET to ethylene. Aminooxyacetate acid (AOA) blocked both the endogenous production of ethylene and that from MET feeds. Daminozide inhibits ethylene production by preventing the conversion of MET to ACC, but it does not appear to act as a simple competitive inhibitor of ACC synthase activity.Abbreviations ACC aminocyclopropane-1-carboxylic acid - AVG aminoethoxyvinylglycine - AOA aminooxyacetic acid - CH cycloheximide - MET methionine - PUT putrescine Author for correspondence     

Inhibition of ethylene production in fruit slices by a rhizobitoxine analog and free radical scavengers

The rhizobitoxine analog, L-2-amino-4-(2-aminoethoxy)-trans-3-butenoic acid (Ro), which effectively inhibits ethylene production in apple (Malus domestica Borkh.) and other tissues at concentrations at about 68 micromolar, inhibited ethylene production by about 50 to 70% in green tomato (Lycopersicon esculentum Mill.) fruit slices but only by about 15% in pink and ripe tomato tissue slices. Ethylene production in climacteric-rise and postclimacteric avocado slices was likewise relatively insensitive to 68 micromolar Ro. At 340 micromolar Ro, inhibition of ethylene production increased up to 50% in pink tomato slices, whereas 680 micromolar Ro was required to inhibit ethylene production by 30% in avocado slices. Incorporation of ¹⁴C from [¹⁴C]methionine into ethylene in green and pink tomato tissues was inhibited by Ro to about the same extent as inhibition of total ethylene production. Results thus far are inconclusive as to the mechanism of Ro resistance in tomato and avocado tissues. At 1 millimolar, free radical scavengers such as benzoate, propyl gallate, nordihydroguaiaretic acid, and to a lesser extent, eugenol, inhibited ethylene production in both Ro-sensitive (green tomato and apple) tissues and Ro-resistant (pink tomato and avocado) tissues. Therefore, free radical steps are suggested in the ethylene-forming systems.     

Rapidly induced ethylene formation after wounding is controlled by the regulation of 1-aminocyclopropane-1-carboxylic acid synthesis

Bean leaves from Phaseolus vulgaris L. var. Pinto 111 react to mechanical wounding with the formation of ethylene. The substrate for wound ethylene is 1-aminocyclopropane-1-carboxylic acid (ACC). It is not set free by decompartmentation but is newly synthesized. ACC synthesis starts 8 to 10 min after wounding at 28°C, and 15 to 20 min after wounding at 20°C. Aminoethoxyvinylglycine (AVG), a potent inhibitor of ethylene formation from methionine via ACC, inhibits wound ethylene synthesis by about 95% when applied directly after wounding (incubations at 20°C). AVG also inhibits the accumulation of ACC in wounded tissue. AVG does not inhibit conversion of ACC to ethylene. Wound ethylene production is also inhibited by cycloheximide, n-propyl gallate, and ethylenediaminetetraacetic acid.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - AVG ammoethoxyvinylglycine - EDTA ethylenediaminetetraacetic acid     

Ethylene as an effector of wound-induced resistance to cellulase in oat leaves

Peeling the abaxial epidermis from oat leaves (Avena sativa var. Victory) induces the formation of wound ethylene and the development of resistance to cellulolytic digestion of mesophyll cell walls. Ethylene release begins between 1 and 2 hours after peeling in the light or dark. Aminoethoxyvinylglycine (AVG, 0.1 millimolar), CoCl₂ (1.0 millimolar), propyl gallate (PG, 1.0 millimolar) or aminooxyacetic acid (AOA, 1.0 millimolar) inhibits, whereas AgNO₃ stimulates wound ethylene formation. Incubation on inhibitors of ethylene biosynthesis (AVG, CoCl₂, PG, AOA) or action (AgNO₃, hypobaric pressure or the trapping of ethylene with HgClO₄) also prevents the development of wound-induced resistance to enzymic cell wall digestion. 1-Aminocyclopropane-1-carboxylic acid (ACC, 1.0 millimolar) reverses AVG (0.1 millimolar) inhibition of the development of resistance. Exogenous ethylene partially induces the development of resistance in unwounded oat leaves.

These results suggest that peeling of oat leaves induces ethylene biosynthesis, which in turn effects changes in the mesophyll cells resulting in the development of resistance to cellulolytic digestion.

     

Auxin-induced ethylene biosynthesis in subapical stem sections of etiolated seedlings of Pisum sativum L.

1-Aminocyclopropane-1-carboxylic acid (ACC) stimulated the production of ethylene in subapical stem sections of etiolated pea (cv. Alaska) seedlings in the presence and absence of indole-3-acetic acid (IAA). No lag period was evident following application of ACC, and the response was saturated at a concentration of 1 mM ACC. Levels of endogenous ACC paralleled the increase in ethylene production in sections treated with different concentrations of IAA and with selenoethionine or selenomethionine plus IAA. The IAA-induced formation of both ACC and ethylene was blocked by the rhizobitoxine analog aminoethoxyvinylglycine (AVG). Labelling studies with L-[U-¹⁴C]methionine showed an increase in the labelling of ethylene and ACC after treatment with IAA. IAA had no specific effect on the incorporation of label into S-methylmethionine or homoserine. The specific radioactivity of ethylene was similar to the specific radioactivity of carbon atoms 2 and 3 of ACC after treatment with IAA, indicating that all of the ethylene was derived from ACC. The activity of the ACC-forming enzyme was higher in sections incubated with IAA than in sections incubated with water alone. These results support the hypothesis that ACC is the in-vivo precursor of ethylene in etiolated pea tissue and that IAA stimulates ethylene production by increasing the activity of the ACC-forming enzyme.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - AVG aminoethoxyvinylglycine, the aminoethoxy analog of rhizobitoxine - IAA indole-3-acetic acid - SAM S-adenosylmethionine - SMM S-methylmethionine     

Biosynthesis of auxin-induced ethylene in mung bean hypocotyls

The pathway of ethylene biosynthesis in auxin-treated mung beanhypocotyls was investigated by comparing the specific radioactivitiesof ethylene produced and S-adenosylmethionine (SAM) in the tissuefollowing the administration of 3,4-¹⁴C-methionine, and by analyzingthe methionine metabolites. When the rate of auxin-induced ethyleneproduction was low due to a low concentration of auxin, thespecific radioactivity of ethylene released was always higherthan that of SAM in the tissue. When the tissue was treatedwith auxin, the tissue produced and accumulated a methioninemetabolite which was converted into ethylene more efficientlythan methionine. The metabolite was identified as 1-aminocyclopropane-l-carboxylicacid (ACC) by means of paper and thin-layer chromatography,high voltage paper electrophoresis and co-crystallization. ACCformation was neither inhibited by low oxygen nor by the inhibitoryprotein of ethylene synthesis, but inhibited by aminoethoxyvinylglycine(AVG). ACC application to the tissue greatly reduced incorporationof 3,4-¹⁴C-methionine into ethylene. The control tissue thatwas not treated with auxin also converted ACC into ethyleneindicating that the enzyme which converts ACC into ethyleneis already present in the tissue and that auxin induced productionof the enzymatic system responsible for the conversion of methionineinto ACC. Ethylene synthesis from ACC was not inhibited by AVG,abscisic acid, cycloheximide or actinomycin D, but inhibitedby low oxygen and the inhibitory protein. (Received November 21, 1979; )     

Role of Ethylene on de Novo Shoot Regeneration from Cotyledonary Explants of Brassica campestris ssp. pekinensis (Lour) Olsson in Vitro

The promotive effect of AgNO₃ and aminoethoxyvinylglycine (AVG) on in vitro shoot regeneration from cotyledons of Brassica campestris ssp. pekinensis in relation to endogenous 1-amino-cyclopropane-1-carboxylic acid (ACC) synthase, ACC, and ethylene production was investigated. AgNO₃ enhanced ACC synthase activity and ACC accumulation, which reached a maximum after 3 to 7 days of culture. ACC accumulation was concomitant with increased emanation of ethylene which peaked after 14 days. In contrast, AVG was inhibitory to endogenous ACC synthase activity and reduced ACC and ethylene production. The promotive effect of AVG on shoot regeneration was reversed by 2-chloroethylphosphonic acid at 50 micromolar or higher concentrations, whereas explants grown on AgNO₃ medium were less affected by 2-chloroethylphosphonic acid. The distinctive effect of AgNO₃ and AVG on endogenous ACC synthase, ACC, and ethylene production and its possible mechanisms are discussed.     

Synergistic enhancement of ethylene production and germination with kinetin and 1-aminocyclopropane-1-carboxylic Acid in lettuce seeds exposed to salinity stress

Relief of salt (0.1 molar NaCl) stress on germination of lettuce (Lactuca sativa L., cv Mesa 659) seeds occurred with applications of 0.05 millimolar kinetin (KIN) and 1 to 10 millimolar 1-aminocyclopropane 1-carboxylic acid (ACC). Treatment with KIN enhanced the pregermination ethylene production under saline condition. A synergistic or an additive enhancement of pregermination ethylene production and germination occurred under saline condition in the presence of KIN and a saturating dose (10 millimolar) of ACC. No KIN-ACC synergism was noted in ethylene production or germination under nonsaline condition. Addition of 1 millimolar aminoethoxyvinylglycine (AVG) inhibited the KIN-enhanced pregermination ethylene production (85 to 89%) and germination (58%) under saline condition but not the synergistic effect of KIN + ACC on ethylene production. Under nonsaline condition, AVG had no effect on germination even though ethylene production was strongly inhibited. Alleviation of salt stress by KIN was inhibited in a competitive manner by 2,5-norbornadiene (NBD) (0.02-0.2 milliliter per liter), and the addition of ACC and/or ethylene reduced this inhibition. An increase in the pregermination ethylene production and germination occurred also by cotylenin E (CN) under saline condition. However, neither AVG (1 millimolar) nor NBD (0.02 to 0.2 milliliter per liter) prevented the relief of salt stress by CN. Thus, KIN may alleviate salt stress on germination by promoting both ACC production and its conversion to ethylene. Rapid utilization of ACC may be the basis for the synergistic or the additive effect of KIN plus ACC. The need for ethylene production and action for the relief of salt stress is circumvented by a treatment with CN.     

Inhibition of ethylene synthesis in tomato plants subjected to anaerobic root stress

Enhanced ethylene production and leaf epinasty are characteristic responses of tomato (Lycopersicon esculentum Mill.) to waterlogging. It has been proposed (Bradford, Yang 1980 Plant Physiol 65: 322-326) that this results from the synthesis of the immediate precursor of ethylene, 1-aminocyclopropane-1-carboxylic acid (ACC), in the waterlogged roots, its export in the transpiration stream to the shoot, and its rapid conversion to ethylene. Inhibitors of the ethylene biosynthetic pathway are available for further testing of this ACC transport hypothesis: aminooxyacetic acid (AOA) or aminoethoxyvinylglycine (AVG) block the synthesis of ACC, whereas CO²⁺ prevents its conversion to ethylene. AOA and AVG, supplied in the nutrient solution, were found to inhibit the synthesis and export of ACC from anaerobic roots, whereas Co²⁺ had no effect, as predicted from their respective sites of action. Transport of the inhibitors to the shoot was demonstrated by their ability to block wound ethylene synthesis in excised petioles. All three inhibitors reduced petiolar ethylene production and epinasty in anaerobically stressed tomato plants. With AOA and AVG, this was due to the prevention of ACC import from the roots as well as inhibition of ACC synthesis in the petioles. With Co²⁺, conversion of both root- and petiole-synthesized ACC to ethylene was blocked. Collectively, these data support the hypothesis that the export of ACC from low O₂ roots to the shoot is an important factor in the ethylene physiology of waterlogged tomato plants.     

Regulation of Auxin-induced Ethylene Biosynthesis. Repression of Inductive Formation of 1-Aminocyclopropane-1-carboxylate Synthase by Ethylene

Changes in the 1-aminocyclopropane-1-carboxylate (ACC) synthaseactivity which regulates auxin-induced ethylene production werestudied in etiolated mung bean hypocotyl segments. Increasesboth in ethylene production and ACC synthase activity in tissuetreated with IAA and BA were severely inhibited by cycloheximide(CHI), 2-(4-methyl-2,6-dinitroanilino)-N-methylpropionamide,actinomycin D and -amanitin. Aminoethoxyvinylglycine (AVG),a potent inhibitor of the ACC synthase reaction, increased theactivity of the enzyme in the tissue 3- to 4-fold. This stimulationalso was severely inhibited by the above inhibitors. Stimulationof the increase in the enzyme content by AVG was partially suppressedby an exogenous supply of ACC or ethylene. Suppression of theincrease in the enzyme took place with 0.3 µl/liter ethylene,and inhibition was increased to 10 µl/liter, which caused65% suppression. Air-flow incubation of the AVG-treated tissue,which greatly decreased the ethylene concentration surroundingthe tissue, further increased the amount of enzyme. Thus, oneeffect of AVG is to decrease the ethylene concentration insidethe tissue. The apparent half life of ACC synthase activity,measured by the administration of CHI, was estimated as about25 min. AVG lengthened the half life of the activity about 2-fold.Feedback repression by ethylene in the biosynthetic pathwayof auxin-induced ethylene is discussed in relation to the effectof AVG. (Received January 22, 1982; Accepted March 26, 1982)     

Effect of plant growth regulators on ethylene production, 1-aminocyclopropane-1-carboxylic acid oxidase activity, and initiation of inflorescence development of pineapple

With the development of pineapple [Ananas comosus (L.) Merr.] as a fresh fruit crop, it became common to force inflorescence development with ethephon [(2-chloroethyl)phosphonic acid] or ethylene throughout the year. Environmental induction (EI) of inflorescence development disrupts scheduling of fruit harvest and may cause significant losses if small plants are induced, resulting in fruits that are too small to be marketable. Our objective was to identify plant growth regulators (PGRs) that could inhibit EI. Because circumstantial evidence indicates that EI occurs in response to naturally produced ethylene or changes in plant sensitivity to it, most work was done with PGRs that inhibit ethylene biosynthesis or block ethylene action. The synthetic auxin 2-(3-chlorophenoxy)propionic acid (CPA) was included because in one study it reduced the percentage of EI. GA₃, aminooxyacetic acid (AOA), aminoethoxyvinylglycine (AVG), daminozide [butanedioic acid mono-(2,2-dimethylhydrazide)], and silver thiosulfate (STS) had no effect on EL CPA, paclobutrazol [(2RS,3RS)-1-(4-chlorophenyl)methyl-4,4-dimethyl-2(1h-1,2,4-triazol-1-yl)penten-3-ol], and uniconazole [(E)-(p-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)-1-penten-3-ol] delayed or inhibited EI of pot-grown pineapple plants. Uniconazole and paclobutrazol inhibited growth and ethylene production by leaf basal-white tissue, and either or both effects could account for the inhibition of EI. Production of 1-aminocyclopropane-1-carboxylic acid (ACC) was unaffected by these compounds, but the activity of ACC oxidase, which converts ACC to ethylene, was inhibited and probably accounts for the reduced ethylene production by leaf basal-white tissue. CPA stimulated ethylene production by stem apical tissue approximately fourfold relative to the control. ACC oxidase activity and the malonyl-ACC (MACC) content in stem apical tissue were also greater than in the control, indicating that CPA greatly stimulated the production of ACC and its sequestration into MACC. The mechanism by which CPA delayed or inhibited EI is not known. CPA, paclobutrazol, and uniconazole appear to have some potential for inhibiting EI of pineapple. Their effect on yield needs to be determined.Abbreviations ACC oxidase 1-aminocyclopropane-1-carboxylic acid oxidase - CPA 2-(3-chlorophenoxy)propionic acid - AOA aminooxyacetic acid - AVG aminoethoxyvinylglycine - daminozide butanedioic acid mono-(2,2-dimethylhydrazide) - DM dry mass - ethephon [(2-chloroethyl)phosphonic acid] - FM fresh mass - GA gibberellin - EI environmental induction of inflorescence development - IA inflorescence appearance - LSD Fisher's protected least significant difference - MACC malonyl-ACC - NAA naphthaleneacetic acid - PGR plant growth regulator - paclobutrazol (2RS,3RS)-1-(4-chlorophenyl)methyl-4,4-dimethyl-2-(1h-1,2,4-triazol-1-yl)penten-3-ol] - uniconazole (E)-(p-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)-1-penten-3-ol - STS silver thiosulfate - M-leaf fourth leaf - Ml-L first leaf younger than M-leaf     

Stimulation of somatic embryogenesis in carrot by ethylene: Effects of modulators of ethylene biosynthesis and action

Endogenous levels of ethylene appeared to he suhoptimal for somatic embryogenesis in a suspension culture of carrot. Low concentrations of 1-aminocyclopropane-1-carboxylic acid (ACC). 2-chloroethylphosphonic acid (ethephon) and elhylene stimulated embryogenesis whereas higher concentrations were inhibitory. The stimulation by ACC was through its conversion to ethylene. whereas the inhibition by ACC was not. Low concentrations of AgNO₃. an inhibitor of ethylene action, inhibited embryo-genesis but stimulated ethylene production. Aminoethoxyvinylglycine (AVG) and aminooxyacetic acid (AOA). commonly used inhibitors of ACC synthase. inhibited both embryogenesis and ethylene production. However, the inhibition of embryogenesis was not related to the inhibition ote ethylene production. Very low concentrations of AVG stimulated embryo production in a way unrelated to its effect on ethylene production. Salicylic acid and CoCl₂. inhibitors of ACC oxidase in other systems, inhibited embryogenesis but. again, in way(s) unrelated to their inhibition of ethylene production. In fact, low concentrations of salicylic acid stimulated rather than inhibited ethylene production. The results show that in suspension-cultured cells, caution is warranted in the interpretation of results obtained with agents presumed to inhibit ethylene biosynthesis. The stimulation of somatic embryogenesis by ethylene unequivocally shows that the inhibition of embryo development by 2.4-dichlorophenoxyacetic acid (2.4-D) and other auxins cannot be through their stimulatory effect on ethylene production.     

Ethylene-Enhanced 1-Aminocyclopropane-1-carboxylic Acid Synthase Activity in Ripening Apples

Apples (Malus sylvestris Mill, cv Golden Delicious) were treated before harvest with aminoethoxyvinylglycine (AVG). AVG is presumed to reversibly inhibit 1-aminocyclopropane-1-carboxylic acid (ACC) activity, but not the formation of ACC synthase. AVG treatment effectively blocked initiation of autocatalytic ethylene production and ripening of harvested apples. Exogenous ethylene induced extractable ACC synthase activity and ripening in AVG-treated apples. Removal of exogenous ethylene caused a rapid decline in ACC synthase activity and in CO₂ production. The results with ripened, AVG-treated apples indicate (a) a dose-response relationship between ethylene and enhancement of ACC synthase activity with a half-maximal response at approximately 0.8 μl/l ethylene; (b) reversal of ethylene-enhanced ACC synthase activity by CO₂; (c) enhancement of ACC synthase activity by the ethylene-activity analog propylene.

Induction of ACC synthase activity, autocatalytic ethylene production, and ripening of preclimacteric apples not treated with AVG were delayed by 6 and 10% CO₂, but not by 1.25% CO₂. However, each of these CO₂ concentrations reduced the rate of increase of ACC synthase activity.

     

Methionine metabolism and ethylene formation in etiolated pea stem sections

Stem sections of etiolated pea seedlings (Pisum sativum L. cv. Alaska) were incubated overnight on tracer amounts of l-[U-(14)C]methionine and, on the following morning, on 0.1 millimolar indoleacetic acid to induce ethylene formation. Following the overnight incubation, over 70% of the radioactivity in the soluble fraction was shown to be associated with S-methylmethionine (SMM). The specific radioactivity of the ethylene evolved closely paralleled that of carbon atoms 3 and 4 of methionine extracted from the tissue and was always higher than that determined for carbon atoms 3 and 4 of extracted SMM.Overnight incubation of pea stem sections on 1 millimolar methionine enhanced indoleacetic acid-induced ethylene formation by 5 to 10%. Under the same conditions, 1 millimolar homocysteine thiolactone increased ethylene synthesis by 20 to 25%, while SMM within a concentration range of 0.1 to 10 millimolar did not influence ethylene production. When unlabeled methionine or homocysteine thiolactone was applied to stem sections which had been incubated overnight in l-[U-(14)C]methionine, the specific radioactivity of the ethylene evolved was considerably lowered. Application of unlabeled SMM reduced the specific radioactivity of ethylene only slightly.     

Biosynthesis of stress ethylene in soybean seedlings: Similarities to endogenous ethylene biosynthesis

The similarity of stress ethylene biosynthesis in whole plants to endogenous ethylene biosynthesis was investigated using two inhibitors of ethylene biosynthesis, aminoethoxyvinylglycine (AVG) and cobalt chloride (Co²⁺); and the intermediates, methionine, S -adenosylmethionine (SAM), and 1-aminocyclopropane-1-carboxylic acid (ACC), of basal ethylene biosynthesis. Stress ethylene production induced by ozone, cadmium, or 2,4-dichlorophenoxyacetic acid was inhibited in hydroponically-grown soybean seedlings in a concentration-dependent manner by both AVG and CO²⁺. The ethylene intermediates evoked responses in intact seedlings similar to that described for endogenous ethylene production in isolated vegetative tissue. The addition of SAM to the hydroponic system relieved AVG inhibition of stress ethylene production. Feeding ACC to the seedlings resulted in increased ethylene production independent of stress application or prior AVG inhibition. Cobalt inhibition of stress ethylene production was relieved by increasing concentrations of ACC. A short lag period of 12–18 min was observed in stress ethylene production following a 30-min ozone exposure. Addition of cycloheximide partially inhibited ozone-induced ethylene production.
These results suggest a common pathway in whole plants for stress ethylene production and endogenous ethylene biosynthesis.     

Ethylene biosynthesis in Regnellidium diphyllum and Marsilea quadrifolia

The pathway of ethylene biosynthesis was examined in two lower plants, the semi-aquatic ferns Regnellidium diphyllum Lindm. and Marsilea quadrifolia L. As a positive control for the ethylene-biosynthetic pathway of higher plants, leaves of Arabidopsis thaliana (L.) Heynh. were included in each experiment. Ethylene production by Regnellidium and Marsilea was not increased by treatment of leaflets with 1-aminocyclopropane-1-carboxylic acid (ACC), the precursor of ethylene in higher plants. Similarly, ethylene production was not inhibited by application of aminoethoxyvinylglycine and -aminoisobutyric acid, inhibitors of the ethylene biosynthetic enzymes ACC synthase and ACC oxidase, respectively. However, ACC was present in both ferns, as was ACC synthase. Compared to leaves of Arabidopsis, leaflets of Regnellidium and Marsilea incorporated little [¹⁴C]ACC and [¹⁴C]methionine into [¹⁴C]ethylene. From these data, it appears that the formation of ethylene in both ferns occurs mainly, if not only, via an ACC-independent route, even though the capacity to synthesize ACC is present in these lower plants.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - AdoMet S-adenosyl-l-methionine - AIB -aminoisobutyric acid - AVG aminoethoxyvinylglycine This research was supported by the U.S. Department of Energy through grant No. DE-FG02-91ER20021 and, in part, by a fellowship of the National Engineering and Research Council of Canada to Jacqueline Chernys.