Ethylene production and involvement during the first steps of durum wheat (Triticum durum) anther culture

The role of ethylene in anther culture of durum wheat ( Triticum durum Desf. cv. Ardente) was analyzed by testing the effects of 2-chloroethylphosphonic acid (ethrel) silver thiosulfate (Ag⁺), a -aminooxyacetic acid (AOA) and 1-aminocyclopropane-l-carboxylic acid (ACC) on microspore division observed after 21 days of culture and on development of calli estimated at day 45. The use of ethrel and Ag⁺ indicated a positive effect of ethylene on microspore division, whereas the use of AOA, and to a lesser extent ACC, snowed a negative effect. In contrast, the addition of ethrel or Ag⁺ indicated that ethylene inhibits the development of microspore-derived calli. AOA gave contradictory results. Ethylene production by anthers was about 7 pl anther⁻¹h⁻¹ and decreased during culture. ACC content in the anthers was maximal at day 9, whereas malonyl ACC (MACC) increased sharply from day 0 to day 3 and then decreased. The addition of AOA or ACC to the culture medium decreased or increased, respectively, ethylene production of anthers and the ACC and/or MACC content, but at concentrations higher than those that modified the formation of calli. This formation seems to occur in two successive phases: induction and initiation of microspore division, which was promoted by ethylene, followed by callus development, which was inhibited by ethylene.     

Chilling-Induced Ethylene Production in Relation to Chill-Sensitivity in Phaseolus spp.

Guye, M. G, Vigh, L. and Wilson, J. M. 1987. Chilling-inducedethylene production in relation to chill-sensitivity in Phaseolusspp.—J. exp. Bot. 38: 680–690. Ethylene production from the primary leaves of six bean (Phaseolusspp.) cultivars known to differ in chill-sensitivity, was monitoredat 23 ?C following chilling of whole plants at 5 ?C for 24 h.The more chill-tolerant cultivars produced greater amounts ofchilling-induced ethylene than the chill-sensitive cultivars.The onset of maximum ethylene production rates and the followingdecline in rates was more rapid in chill-tolerant cultivars.This pattern of ethylene production was also similar when chill-tolerancewas chemically enhanced by choline treatment. The low levelsof ethylene production in chill-sensitive genotypes was alsoreflected by their poor ability to convert the exogenously appliedethylene precursor, 1-aminocyclopropane-l-carboxylic acid (ACC),to ethylene. Moderate levels of leaf water deficit induced by chilling chill-tolerantcultivars and choline treated plants appeared to stimulate chilling-inducedethylene production. High levels of leaf wilt, shown by morechill-sensitive cultivars, reduced this stimulatory effect.Ethylene production was slightly greater when warming was carriedout in the light rather than in the dark. Key words: Ethylene, ACC, choline, chill-sensitivity, Phaseolus     

Ethylene precursors and antagonists increase embryogenesis of Hordeum vulgare L. anther culture

The role of ethylene in microspore embryogenesis and regeneration was analyzed by studying the effects of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) and the ethylene antagonists silver nitrate and silver thiosulphate on the androgenic response of in vitro cultured anthers of seven genotypes of barley. Incorporation of either ACC or silver salts in the culture medium lead to a significant increase in callus induction for five of the seven genotypes tested. The treatment that increased callus induction depended upon genotype. Only anthers cultured on 1 mg l^–1 silver thiosulphate gave rise to fertile plants in all seven genotypes tested.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - IAA indole acetic acid - PAA phenyl acetic acid - STS silver thiosulphate - ACC 1-aminocyclopropane-1-carboxylic acid     

Endogenous ethylene in indirect somatic embryogenesis of <Emphasis Type="Italic">Medicago sativa</Emphasis> L.

Ethylene biosynthesis during different phases of somatic embryogenesis in Medicago sativa L. cv. Rangelander using two regeneration protocols, RPI and RPII, was studied. The highest ethylene production was detected during callus growth on induction medium in both regeneration protocols. Significantly less ethylene was produced by embryogenic suspension than by callus (RPII). Developing embryos synthesized higher amounts of ethylene than mature embryos. Production of ethylene was strongly limited by the availability of 1-aminocyclopropane-1-carboxylic acid and also by ACC-oxidase activity. However, removal of ethylene from culture vessels’ atmosphere using KMnO₄ or HgClO₄ had no significant effect on callus growth, somatic embryo induction and development. Reducing of ethylene biosynthesis by aminoethoxyvinylglycine substantially decreased somatic embryo production and adversely affected their development, indicating ethylene requirement during proliferation and differentiation but not induction.     

The effect of AOA on ethylene and polyamine metabolism during early phases of somatic embryogenesis in Medicago sativa

Changes in the levels of ethylene, 1-aminocyclopropane-1-carboxylic acid (ACC), 1-(malonylamino)cyclopropane-1-carboxylic acid (MACC) and polyamines were simultaneously investigated during the early phases of alfalfa somatic embryogenesis. These included the period of induction and subculture of callus, and 3- and 7-day suspension cultures for the induction of somatic embryogenesis. The polyamines contained in the embryogenic callus were found to include putrescine (Put), spermidine (Spd) and spermine (Spm), but the level of Spm was much less than that of Put and Spd. There was a dramatic increase in MACC after induction of embryogenesis, and ACC levels were lower in somatic embryos than in embryogenic callus. Induction of embryogenesis for 3 days increased the levels of ACC and polyamines to a maximum level, and these then reduced as the embryogenesis proceeded. The ratios of Put/Spd and ACC/MACC were decreased during the induction. This indicated that both high levels of ACC and polyamines might be a prerequisite for early differentiation during the induction of the embryogenesis. Thus, there appears not to be competition between polyamine biosynthesis and ethylene biosynthesis at least during the induction of somatic embryogenesis, because both the polyamines and ACC were simultaneously increased during the induction period. Conversion of ACC into MACC and the maintenance of a relatively high level of polyamines, especially Spd, appear to be important for further development of the embryos.
When aminooxylvinylglycine (AOA) was added at the initiation of the callus subculture, it had no significant effect on the callus growth, the ethylene production and ACC level of the callus. However, AOA increased the numbers of the embryos accompanying an increase in Spd level and S-adenosylmethionine decarboxylase (SAMDC) activity. Thus, the AOA effect could be associated with Spd increase rather than with the effect of ethylene biosynthesis.     

Endogenous Levels and Transport of 1-Aminocyclopropane-1-Carboxylic Acid in Stamens of Ipomoea nil (Convolvulaceae)

Filament and corolla growth in flowers of Ipomoea nil are inhibited by ethylene production. Anthers inhibited filament growth in vitro during younger stages of development even in the presence of the growth promoter gibberellic acid (GA₃). To test whether the anthers could be sources of 1-aminocyclopropane-1-carboxylic acid (ACC) endogenous levels of ACC and ethylene production were monitored using gas chromatography. To also test whether the filaments could be transport vectors for ACC the movement of [¹⁴C]ACC was assessed by scintillation counting from donor agarose blocks, through filament sections, and into receiver agarose blocks. While ACC levels fluctuated in anthers 87 to 21 h before anthesis, anthers contained increased levels of ACC from 15 to 6 hours before anthesis. Ethylene production also fluctuated but peak levels were shifted about 6 hours closer to anthesis than ACC levels within the anthers. Both ACC and ethylene levels in filaments showed fluctuations similar to those in the anthers. [¹⁴C]ACC movement became increasingly basipetal during development. Older stages showed greater polar [¹⁴C]ACC efflux rates, while all stages showed constant polar influx rates. Low levels of endogenous ACC were transported basipetally from the anther through the filament into agarose blocks at all stages of development. Corresponding levels of endogenous ethylene production remained constant between the various stages during ACC transport. We have evidence that stamens of I. nil have a role as source tissues and transport vectors for ACC, to stimulate corolla growth, such as corolla unfolding and senescence.     

Somatic cell genetic studies inBrassica species

In vitro culture ofBrassica alba anthers on a growth medium containing inorganics of KB5 and organics, iron, sucrose and hormones of B5 resulted in a very high response of anthers (93.75%) towards callus induction. All the calli transferred to regeneration media responded favourably even after six months of callus induction. Numerous torpedo-shaped embryoids developed in clusters at many sites from each callus mass. Secondary embryogenesis and multiple shoot formation was also observed in many cases. The number of embryoids and plantlets produced by one embryogenic anther were as high as 169.8 and 17 respectively. 87% of the regenerated plants were haploids.     

Differential physiological and morphological responses of inbred lines to the ethylene precursor 1-aminocyclopropane-1-carboxylic acid by cultured Helianthus annuus (sunflower) shoot tips

The four Helianthus annuus (sunflower) inbred lines examined showed different abilities to convert 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene and different morphological responses to exogenous ACC, however, ACC had no effect on precocious flowering. The greatest effect of ACC was seen with inbred SS405B where it suppressed shoot growth and induced hypocotyl enlargement and callus induction. The greatest response did not correlate with the highest ethylene production. Although each inbred responded differently, callus induction and hypocotyl enlargement observed in hypocotyl segments treated with naphthalene acetic acid and benzyladenine could be partially explained as ethylene-mediated effects of the two hormones. It is suggested that inbred differences could be due to different endogenous hormone levels and/or different sensitivities to them.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - AVG aminoethoxyvinylglycine - BA benzyladenine - NAA naphthalene acetic acid     

Studies on lavandin callus cultures: Ethylene production in relation to the growth

Ethylene production and growth of callus cultures of lavandin (Lavandula offidnalis Cham x Lavandula latifolia Villars) cv. Grosso were examined. Callus lines, derived from various kinds of primary expiants (shoot tip, leaf and calyx), exhibited differences in ethylene production patterns independent of callus growth. Moreover these differences could not be ascribed to the expiant source. Within a line, ethylene pattern paralleled callus growth curve. Variations in ethylene evolution were induced in shoot tip callus by means of ACC, AVG and varied amounts of 2,4-D in the culture medium. Following all these treatments callus growth was not altered. Hie decrease in 2,4-D concentration caused changes in Chl a and water content of the tissues.     

Biotechniques for improving acid aluminum tolerance in alfalfa

Alfalfa (Medicago sativa L.), cultivars ARC, Regen Y and Saranac were selected in vitro in a recently developed acid/aluminum toxic media. The new media produced higher initiation rates and higher fresh callus weights than those obtainable with the media described by Meredith and Connor for the selection of aluminum resistant variants in Nicotiana plumbaginofolia. Both rescue and direct initiation yielded adequate amounts of healthy callus for the initiation of embryogenesis. The toxic effect of the acid/aluminum media is expressed in both the percent of explants initiating callus and in the fresh-weights obtained during initiation and two subsequent subcultures.     

Ethylene production during anther culture of Brussels sprouts (Brassica oleracea var gemmifera) and its relationship with factors that affect embryo production

The ethylene inhibitor silver nitrate (AgNO₃) is known to overcome the poor response of the Brussels sprouts cultivar Hal to anther culture. Ethylene production by Hal anthers after 6 h of culture at 35°C was on average 10- and 20-fold greater than from anthers of the highly responsive cultivars Gower and GA1 x RDF2. The initial 24 h period at 35°C necessary for embryogenesis in anther culture of Brussels sprouts generally reduced ethylene production by the anthers after 6, 24, 48 and 72 h of culture, although the effect was not seen in 2 out of 3 Hal experiments until 24 h, and after 6 h was only found with 1 of 3 GA1 x RDF2 experiments. Embryo production was inhibited by the inclusion of the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid (ACC) or the ethylene-releasing compound, ethephon in the media. Silver nitrate (AgNO₃) and the ethylene biosynthesis inhibitor aminoethoxyvinylglycine (AVG) promoted embryogenesis but did not substitute for the high temperature treatment. The relevance of ethylene production during anther culture to the effects of genotype and high temperature on anther culture embryogenesis is discussed.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - AVG aminoethoxyvinylglycine     

Effects of 1-aminocyclopropane-1-carboxylic acid,aminoethoxyvinylglycine, methylglyoxal bis-(guanylhydrazone) and dicyclohexylammonium sulfate on induction of embryogenic competence of mango nucellar explants

The effects of aminoethoxyvinylglycine (AVG),1-aminocyclopropane- 1 -carboxylic acid (ACC), dicycyclohexylammonium sulfate (DCHA) and methylglyoxal bis-(guanylhydrazone) (MGBG) on induction of embryogenic competence from the nucellus of two mango genotypes, ‘Tutehau’ (polyembryonic) and ‘Tommy Atkins’ (monoembryonic), were compared. Induction of embryogenic competence in the explanted nucellus of ‘Tommy Atkins’ was more sensitive to AVG and DCHA than ‘Tutehau’. MGBG had no effect on induction of embryogenic competence of either genotype, and ACC suppressed somatic embryogenesis with both cultivars. Ethylene biosynthesis was greater from explanted ‘Tommy Atkins’ cultures. Reducing ethylene biosynthesis with AVG marginally increased the number of embryogenic cultures of ‘Tutehau’, but completely inhibited somatic embryogenesis in ‘Tommy Atkins’ cultures. Ethylene biosynthesis was stimulated by ACC in nonembryogenic cultures of both genotypes and in ‘Tommy Atkins’ embryogenic cultures, in which ethylene biosynthesis was ca 7 X greater than in ‘Tutegau’ embryogenic cultures. The greater sensitivity of ‘Tommy Atkins’ to the ethylene antagonist, AVG, may be due to its greater sensitivity to disturbance of ethylene biosynthesis and spermidine synthesis. This revised version was published online in June 2006 with corrections to the Cover Date.     

Abscisic acid and methyl jasmonate as regulators of ethylene biosynthesis during somatic embryogenesis of Medicago sativa L.

Effects of abscisic acid (ABA) and methyl jasmonate (MeJA) on ethylene production, ACC oxidase (ACO) activity, and content of 1-aminocyclopropane-1-carboxylic acid (ACC) during indirect somatic embryogenesis (SE) of Medicago sativa L. were studied. ABA and MeJA, at 50 μM, were applied during the induction, proliferation, or differentiation phase. ABA decreased ethylene production at the beginning of callus and SE induction and during the differentiation of somatic embryos. The hormone inhibited ACO activity in explants with overgrowing callus during the first two weeks of induction, in embryogenic suspension and also in differentiating embryos. The ACC content was reduced by ABA in callus at the end of SE induction, in embryogenic suspension and in globular embryos, but elevated in cotyledonary embryos. MeJA had no significant effect on ethylene production during M. sativa SE, despite the fact, that it inhibited ACO activity during the first two weeks of induction and in torpedo and cotyledonary embryos. The ACC content was increased by MeJA in 14-day-old callus and embryogenic suspension but was inhibited in globular embryos. Both ABA and MeJA seem to be involved in the regulation of ethylene biosynthesis during distinct phases of SE in M. sativa. It might be considered that exogenous ABA, more probably than MeJA, exerts its inhibitory effect on M. sativa somatic embryo formation by modifying ethylene production.     

Effect of MGBG on Growth of the Callus Tissue,Induction of Somatic Embryogenesis of Alfalfa and Their Ethylene Biosynthesis

MGBG [methylglyoxal bis(guanylhydrazone)], when added to the initial of callus subculture, promoted ethylene production in callus grawn in Bsh medium, and enhanced the ACC (1-aminocyclopropane-l-carboxylic acid) level and the ACC synthase activity both in the callus grown in Bsh medimn, and in the suspension cultures in Bsg liquid medium for induction of somatic embryogenesis. However, MGBG reduced the malonyl-ACC (MACC) level of the suspension tissues. The treatments of MGBG caused growth of the cultured tissues and induction efficiency of somatic embryogenesis to reduce. Moreover, the effects of MGBG was enhanced when it was added at induction medium of somatic embryogenesis. It could be concluded that this inhibitory effect of MGBG could be due to its promotion of ethylene, ACC levels and ACC synthase activity which has shown to inhibit somatic embryogenesis. The reduction of MACC level could be also involved in the MGBG effect on induction of the embryogenesis.     

Enhancement of wound-induced ethylene synthesis by ethylene in preclimacteric cantaloupe

Although intact fruits of unripe cantaloupe (Cucumis melo L.) produce very little ethylene, a massive increase in ethylene production occurred in response to excision. The evidence indicates that this wound ethylene is produced from methionine via 1-aminocyclopropanecarboxylic acid (ACC) as in ripening fruits. Excision induced an increase in both ACC synthase and the enzyme converting ACC to ethylene. Ethylene further increased the activity of the enzyme system converting ACC to ethylene. The induction by ethylene required a minimum exposure of 1 hour; longer exposure had increasingly larger effect. The response was saturated at approximately 3 microliters per liter ethylene and was inhibited by Ag⁺. Neither ethylene nor ACC had a promotive or inhibitory effect on ACC synthase beyond the effect attributable to wounding.     

Effect of l-aminocyclopropane-l-carboxylic acid,silver nitrate,and norbornadiene on plant regeneration from maize callus cultures

The effect of the ethylene antagonists norbornadiene and silver nitrate and the ethylene precursor l-aminocyclopropane-l-carboxylic acid (ACC) on Zea mays plant regeneration was studied. A 12-fold increase in plant regeneration, as measured by number of plants obtained per gram fresh weight from callus cultures of maize inbreds Pa91 and H99, was obtained by 250 M norbornadiene and 100 M silver nitrate treatments. An increase in amout of nonregenerable tissue and a 68% decrease in plant regeneration were associated with callus treated with 1 mM ACC. Ethylene emanation from 1 mM ACC treated callus reached a maximum of 170 nl g^–1 h^–1 after 3 days compared to 7 nl g^–1 h^–1 for the control. The free proline content was up to 80% lower in 1 mM ACC treated callus grown for 30 days on medium with or without 12 mM proline, respectively, as compared to each control. These studies indicate that ethylene action inhibitors such as norbornadiene and silver nitrate can be used to increase plant regeneration efficiency from maize callus cultures.Abbreviations ACC l-aminocyclopropane-l-carboxylic acid - gfw gram fresh weight     

Wound-induced Ethylene Formation in Albedo Tissue of Citrus Fruit

Excised albedo tissue of citrus fruit (Citrus unshiu and Citrus hassaku) produced ethylene at an increasing rate in response to wounding and aging. The application of 1-aminocyclopropane-1-carboxylic acid (ACC) enhanced ethylene production in both the fresh and aged tissues, but this increase was greater in the aged tissue than in the fresh tissue. ACC content was very low in fresh tissue but increased greatly in aging tissue, paralleling the rise in ethylene production. Aminoethoxyvinylglycine (AVG) strongly inhibited ethylene production in the aged tissue. In the presence of ACC, however, ethylene production was not inhibited by AVG. These results suggest that ACC is an intermediate in the pathway of ethylene biosynthesis in the albedo tissue and that both steps of ACC formation and ACC conversion to ethylene are enhanced by wounding and aging. Inhibitors of protein synthesis, cycloheximide and 2-(4-methyl-2,6-dinitroanilino)-N-methyl propionamide, strongly inhibited ethylene production in the albedo tissue, implying that protein synthesis is required to maintain the continuous evolution of ethylene. The stimulation of ethylene production by ACC was reduced by the addition of l-methionine, whereas d-methionine had very little inhibitory effect. Ethylene production in the albedo tissue was also inhibited by the addition of n-propyl gallate and 3,5-dibromo-4-hydroxybenzoic acid.     

Effects of ethylene on somatic embryogenesis and galanthamine content in <Emphasis Type="Italic">Leucojum aestivum</Emphasis> L. cultures

The influence of ethylene on in vitro morphogenesis of Leucojum aestivum and galanthamine accumulation was studied. Calli were cultivated on Murashige and Skoog (MS) medium supplemented with 25 μM 4-amino-3,5,6-trichloropicolinic acid (picloram) and 0.5 μM benzyladenine (BA). During incubation under these conditions, callus cultures produced ethylene (9.5 nL/g fresh weight: F.W.) whereas no ethylene was found in somatic embryos cultivated on medium supplemented with 0.5 μM α-naphthalene acetic acid (NAA) and 5 μM zeatin. Application of the precursor of ethylene 1-aminocyclopropane-1-carboxylic acid (ACC) increased ethylene production in both cultures, and decreased callus growth by a factor of 1.2, whereas callus growth was enhanced by a factor of 1.1 in the presence of an inhibitor of ethylene silver nitrate (AgNO₃) or by a factor of 1.2 with an absorbent potassium permanganate (KMnO₄). ACC enhanced the induction of somatic embryos and the development of globular embryos. Removal of ethylene by KMnO₄ during somatic embryogenesis led to the development of plants with greater length. Silver thiosulphate (STS) induced galanthamine production in callus cultures (0.1% dry weight), whereas ACC induced galanthamine production in somatic embryo cultures (2% dry weight).     

Changes in 1-Aminocyclopropane-1-carboxylic Acid Content and Ethylene Production in Hiproly Barley Callus during Differentiation

During differentiation after auxin withdrawal, the change in the ethylene production of Hiproly barley callus paralleled the change in 1-aminocyclopropane-1-carboxylic acid (ACC) content. The levels of ACC and ethylene production decreased rapidly, and then increased in Hiproly barley callus.

Aminooxyacetic acid (AOA) prevented the ACC and ethylene production of the callus. Moreover, aminoisobutyric acid (AIB) also inhibited the ethylene production, but did not prevent the ACC synthesis of the callus. On the other hand, methylglyoxal-bis(guanylhydrazone) (MGBG) greatly enhanced the ACC and ethylene production. Formation of adventitious roots in Hiproly barley callus was enhanced by the cultivation in the medium containing AIB or AOA. However, differentiation of the callus was strongly inhibited by MGBG.

Thus, prevention of ethylene production may be significant for differentiation of Hiproly barley callus.     

The Role of 1-Aminocyclopropane-1-carboxylic acid in the Control of Low Temperature Induced Ethylene Production in Leaf Tissue of Phaseolus vulgaris L .

Ethylene production from leaf discs of dwarf bean (Phaseolausvulgaris L.) was less than 02 nl g^–1 h^–1 at 5 Cbut rapidly increased tenfold on transfer to 25 C. The lowethylene production at 5 C and the potential for overshootproduction on transfer to 25C were not associated with accumulationof the ethylene synthesis intermediate 1-aminocyclopropane-1-carboxylicacid (ACC). Addition of exogenous ACC to leaf discs incubatedat 5C increased ethylene production, while similarly incubatedleaf discs did not synthesize increasing amounts of endogenousACC until they were transferred to 25 C. The basis for theovershoot in ethylene production when leafdiscs were transferredfrom 5 to 25 C appears to reside in changes to the pathwayleading to the synthesis of ACC or an earlier intermediate inthe pathway of ethylene biosynthesis. Ethylene, 1-aminocyclopropane-l-carboxylic acid, Phuseolru vulgaris L., dwarf bean, temperature