Phytohormonal regulation of S-adenosylmethionine synthetase and S-adenosylmethionine levels in dwarf pea epicotyls.

A significant stimulation (2- to 2.5-fold) of AdoMet synthetase was witnessed in glibberellicd acid (GA3, 1 microM)-treated epicotyls of the dwarf pea (Pisum sativum). This was accompanied by a 2.4-fold increase in the endogenous pool of S-adenosylmethionine. Both abscisic acid (10 microM) and cycloheximide (20 micrograms/ml) inhibited the GA3-mediated enhancement of AdoMet synthetase activity. Three isozymes of AdoMet synthetase were detected in GA3-treated epicotyls, whereas a single activity peak was observed in controls. Thus, GA3 seems to control the induction of two new isozymes of AdoMet synthetase in the dwarf pea. By contrast, the tall pea exhibited three isozymes of AdoMet synthetase even in the absence of GA3 treatment. High concentration of L-methionine (2 mM) mimicked the GA3-elicited induction of two new isozymes of AdoMet synthetase in dwarf pea epicotyls.     

The biological and structural similarity between lunularic acid and abscisic acid

Lunularic acid (LA) inhibited not only the germination and the growth of cress and lettuce at 1 mM but also the gibberellic acid (GA3)-induced alpha-amylase induction in embryoless barley seeds at 120 microM, which was recognized as a specific activity of abscisic acid (ABA). Moreover LA and ABA equally inhibited the growth of Lunularia cruciata A18 strain callus at 40 and 120 microM. A computational analysis revealed that the stable conformers of LA could be superimposed on the stable ABA conformers. In addition, the antibody raised against the conjugate of C1-ABA-bovine serum albumin (ABA-BSA) reacted with LA-horse-radish peroxidase (LA-HRP) conjugate as well as ABA-HRP conjugate, apparently. These results can explain why LA has ABA-like activity in higher plants. Moreover the results suggest that LA and ABA bind to the same receptor in higher plants.     

Effect of exogenous gibberellic acid,abscisic acid,and benzylaminopurine on epicotyl dormancy of cultured herbaceous peony embryos

Epicotyl dormancy was broken in cultured peony (Paeonia lactiflora Pall.) embryos after topical application of agarose gels containing gibberellic acid, with optimum growth at 1.5 mM gibberellic acid. Addition of 100 M abscisic acid to the medium resulted in complete inhibition of gibberellic acid-stimulated promotion of dormant epicotyls. Epicotyl dormancy was also broken in embryos by culture on media containing 1 or 10 M benzylaminopurine. A highly significant increase in leaf number occurred when embryos were both cultured on medium containing benzylaminopurine and treated topically with gibberellic acid. Anatomical and morphological studies indicated that the increase in shoot growth was due to the development and growth of 1) buds formed at the cotyledonary node, 2) axillary buds, and 3) adventitious meristems originating from subepidermal parenchymatous tissue.Abbreviations ABA abscisic acid - BA N⁶-benzylaminopurine - DMSO dimethyl sulfoxide - GA₃ gibberellic acid - LS Linsmaier and Skoog     

Reversal of induced dormancy in lettuce by ethylene, kinetin, and gibberellic Acid

The germination of lettuce seeds (Lactuca sativa L., cv. Premier Great Lakes) was significantly inhibited by high temperature (32 C), 0.1 mM abscisic acid or 0.4 M mannitol. Ethylene (16 μl/1 of air) partially reversed the dormancy induced by all three inhibitors but only in the presence of 1 mM gibberellic acid (GA) or light. Neither ethylene plus GA nor ethylene plus light were able to promote germination when thermal inhibition was imposed at 36 C. Addition of 0.01 mM kinetin to the ethylene plus GA or light reversed thermodormancy at 36 C. The dormancy imposed by abscisic acid was also reversed by kinetin. Kinetin was unable to reverse the osmotic dormancy imposed by mannitol. The reversal of osmotic dormancy by ethylene or ethylene plus GA was actually inhibited by kinetin but only in the light. Kinetin apparently stimulates cotyledonary growth in the presence of light, and this growth may compete for certain metabolites critical to radicle growth and subsequent germination. Kinetin and ethylene, as demonstrated primarily in the thermodormancy at 36 C and in osmotic dormancy, appear to regulate a common event(s) leading to germination but through mechanisms unique to each respective growth regulator. The regulation of germination by ethylene is absolutely dependent upon an interaction with GA and/or light.     

Induction and development of somatic embryos from spinach (Spinacia oleracea) leaf segments

Somatic embryogenesis was induced in embryogenic calli of Spinacia oleracea L., on a Murashige and Skoog (1962) medium, containing 4.6 μM kinetin as the sole growth regulator. Abscisic acid, gibberellic acid, and indole-3-acetic acid were supplemented to kinetin-containing medium and their effects on the initiation of somatic embryos was studied. Abscisic acid at a particular concentration (4 μM) dramatically increased the number of embryos per g fresh weight of callus, while both gibberellic acid and indole-3-acetic acid suppressed the embryo initiation. It is suggested that the promoting effect of abscisic acid on the embryo initiation may be explained as a stress response of the tissue. The relative number of globular embryos vs. the embryos in heart/torpedo and cotyledonary stages was increased at 4 μM abscisic acid and at all gibberellic acid concentrations (0.3–10 μM). In contrast, the ratio of globular to polar embryos was lower than in controls at 1 μM abscisic acid and indole-3-acetic acid (1 and 10 μM). The effects of growth regulators on the ratio of globular to polar embryos indicate that they interfere with the normal distribution of cell division and cell expansion during early embryogenesis. This revised version was published online in June 2006 with corrections to the Cover Date.     

Inhibition of cottonseed germination with abscisic Acid and its reversal

Germination of cottonseed (Gossypium hirsutum L.) was inhibited by abscisic acid. Inhibition was greater when seeds were soaked in abscisic acid for 5 hours and dried prior to germination than when abscisic acid was applied in the germination medium. (2-Chloroethyl)phosphonic acid, gibberellic acid, and kinetin partially overcame the inhibitory action of abscisic acid. Combinations of (2-chloroethyl)phosphonic acid with gibberellic acid or kinetin were more effective than the individual substances. Germination also was partially restored by removal of seed coats. Fusicoccin completely restored germination of abscisic acidtreated seeds.     

Effect of phytohormones on fiber initiation of cotton ovule

In order to study the effect of phytohormones on cotton fiber initiation, contents of four endogenous phytohormones and activities of four related enzymes in ovules (in vivo) of a fuzzless–lintless mutant (fl) and its wild-type (FL) line were measured from 4 days before anthesis (day −4) to 4 days after anthesis (day 4). The results showed that contents of indole-3-acetic acid, gibberellic acid (GA), and zeatin riboside in fl ovules were lower than those in FL ovules. Therefore, indole-3-acetic acid, GA, and zeatin riboside were thought to be the promoters of fiber initiation. Although abscisic acid (ABA) content in fl ovule was slightly higher than that in FL ovule on day 0, which might imply that ABA inhibited fiber initiation. Fiber initiation could also be influenced by enzyme through regulating synthesis and degradation of related phytohormones since fl ovules were significantly higher in activities of indole-3-acetic acid oxidase, cytokinin oxidase and peroxidase, but lower in activity of tryptophan synthetase than those in FL ovules. To test the above hypothesis, exogenous plant growth regulators were also applied for the culture of ovules from fl and FL in vitro. When no regulators were added, no fiber was induced on fl ovule, but a few fibers were induced in FL ovule. Higher total fiber units (TFU) were observed when indole-3-acetic acid and gibberellic acid (GA₃) were applied either separately or in combination to media. TFU did not increased with zeatin riboside alone, but the highest TFU was achieved when zeatin riboside was applied together with indole-3 acetic acid and GA₃, which implied that fiber initiation could be promoted by them as additive.     

Growth and graviresponsiveness of primary roots of Zea mays seedlings deficient in abscisic acid and gibberellic acid

The objective of this research was to determine if gibberellic acid (GA) and/or abscisic acid (ABA) are necessary for graviresponsiveness by primary roots of Zea mays. To accomplish this objective we measured the growth and graviresponsiveness of primary roots of seedlings in which the synthesis of ABA and GA was inhibited collectively and individually by genetic and chemical means. Roots of seedlings treated with Fluridone (an inhibitor of ABA biosynthesis) and Ancymidol (an inhibitor of GA biosynthesis) were characterized by slower growth rates but not significantly different gravicultures as compared to untreated controls. Gravicurvatures of primary roots of d-5 mutants (having undetectable levels of GA) and vp-9 mutants (having undectable levels of ABA) were not significantly different from those of wild-type seedlings. Roots of seedlings in which the biosynthesis of ABA and GA was collectively inhibited were characterized by gravicurvatures not significantly different for those of controls. These results (1) indicate that drastic reductions in the amount of ABA and GA in Z. mays seedlings do not significantly alter root graviresponsiveness, (2) suggest that neither ABA nor GA is necessary for root gravicurvature, and (3) indicate that root gravicurvature is not necessarily proportional to root elongation.     

Effect of phytohormones on the transmembrane translocation of protons in plasma membrane vesicles from tubers of Solanum tuberosum L

Effects of phytohormones gibberellic acid (GA) and abscisic acid (ABA) on the ATP-dependent transmembrane transport of protons were studied in plasma membrane vesicles (PMVs) from non-dormant potato tubers. The uptake of H+ into PMVs was assessed by the fluorescence quenching of acridine orange (AO) after the addition of ATP to the incubation medium. Addition of ATP to the incubation medium led to the instantaneous rise of the AO fluorescence intensity followed by its decrease. The fluorescence quenching was not observed in the presence of either protonophore CCCP or inhibitors of the membrane-bound H+-ATPase. It is concluded that the ATP-induced quenching of the AO fluorescence resulted from the accumulation of protons in PMVs due to the function of the plasma membrane-bound H+-ATPase. Depending on their concentrations, GA and ABA either inhibited or stimulated the ATP-driven H+ translocation across the vesicle membrane. The growth-stimulating hormone GA at concentrations of 10(-9)-10(-5) M increased the initial rate of the fluorescence quenching, whereas 10(-4) M GA slightly inhibited the H+ translocation. The growth inhibitor ABA at a concentration of 10(-9) M slightly increased the rate of the proton accumulation in PMVs; at higher concentrations (10(-8)-10(-4) M), ABA inhibited the H+ translocation. Acetic acid, which has pK similar to pK of GA and ABA, did not influence the ATP-dependent H+ accumulation in PMVs, suggesting the hormone-specific action of GA and ABA on the H+-ATPase activity. In the presence of DCC, which completely inhibited the accumulation of H+, GA and ABA did not affect the passive proton efflux from PMVs. It is proposed that the mechanisms of the regulatory effects of phytohormones may involve modification of H+-ATPase activity leading to changes in the electrochemical gradient of H+ across the plasma membrane.     

Influence of culture conditions on embryo formation and maturation in auxin-induced embryogenic cultures of Digitalis obscura

Somatic embryogenesis was induced in hypocotyls of Digitalis obscura using indoleacetic acid or 2,4-dichlorophenoxyacetic acid with different culture and subculture conditions. Indoleacetic acid-induced embryogenic cultures were used to investigate the effects of amino acids, polyamines and growth regulators on embryo differentiation and maturation. Supplementation of the media with amino acids, polyamines or abscisic acid did not influence or had an adverse effect on embryogenic response. Gibberellic acid at 1.4 M in either culture (30 days) or subculture medium was effective in promoting both differentiation and normal embryo development. The efficiency of somatic embryogenesis was greatly enhanced when isolated indoleacetic acid-induced proembryogenic masses were subcultured in liquid medium with reduced auxin content.Abbreviations ABA abscisic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - GA gibberellic acid - IAA indoleacetic acid - Ptr putrescine - Spd spermidine - Spn spermine     

Gibberellic acid (GA) increases fibre cell differentiation and secondary cell-wall deposition in spring wheat (Triticum aestivum L.) culms

The role of gibberellic acid (GA) in differentiation and secondary cell-wall deposition of fibre cells of spring wheat (Triticum aestivum) culms was studied using applications of GA and chlormequat (a GA biosynthesis inhibitor). In certain genotypes, higher GA levels may increase the number of cortical fibre cell files by changing cell fate from parenchyma to fibre, and induce thicker secondary cell-walls.     

Abscisic acid in the thermoinhibition of lettuce seed germination and enhancement of its catabolism by gibberellin

Germination of lettuce (Lactuca sativa L. cv. 'Grand Rapids') seeds was inhibited at high temperatures (thermoinhibition). Thermoinhibition at 28 degrees C was prevented by the application of fluridone, an inhibitor of abscisic acid (ABA) biosynthesis. At 33 degrees C, the sensitivity of the seeds to ABA increased, and fluridone on its own was no longer effective. However, a combined application of fluridone and gibberellic acid (GA3) was able to restore the germination. Exogenous GA3 lowered endogenous ABA content in the seeds, enhancing catabolism of ABA and export of the catabolites from the intact seeds. The fluridone application also decreased the ABA content. Consequently, the combined application of fluridone and GA3 decreased the ABA content to a sufficiently low level to allow germination at 33 degrees C. There was no significant temperature-dependent change in endogenous GA1 contents. It is concluded that ABA is an important factor in the regulation of thermoinhibition of lettuce seed germination, and that GA affects the temperature responsiveness of the seeds through ABA metabolism.     

Influences of ascorbic acid and gibberellic acid in alleviating effects of salinity in Petunia under in vitro

Salinity is one of the abiotic stresses that limits the growth and productivity of many crops. A possible survival strategy for plant under saline conditions is to use compounds that could minimize the harmful effects of salt stress on the plant development. The objective of the presented study was to investigate the effect of exogenous ascorbic acid (ASA) with or without gibberellic acid (GA3) on key growth and biochemical parameters in two petunia cultivars ‘Prism Rose’ and ‘Prism White’ under saline (150 mM NaCl) and non-saline in vitro condition. Nodal cutting with an axillary buds were used as explants. Application of 1 mM ascorbic acid with or without 0.05 mM gibberellic acid into the MS medium stimulated the length of shoots and the number of new shoots of ‘Prism Rose’; whereas, it decreased the root length and the number of roots of both ‘Prism Rose’ and ‘Prism White’ under non-saline condition. The addition of ascorbic acid with or without gibberellic acid into the MS medium under saline condition, increased the length of plants and the number of new shoots, but did not affect their root number and length. NaCl treatments increased the proline content and lipid peroxidation which was indicated by the accumulation of malondialdehyde (MDA). The study revealed a correlation between chlorophylls a and b content and the leaf pigmentation intensity – parameter a*. Addition of 1 mM ascorbic acid with 0.05 mM gibberellic acid into the MS medium plays a protective role in salinity tolerance by improving the shoot growth and the development as well as increasing the activities of the antioxidant enzymes and other antioxidant substances.     

Correlation of the induction of various enzymes concerned with phenylpropanoid and lignin synthesis during differentiation of bean callus (Phaseolus vulgaris L.)

Summary The activities of phenylalanine ammonia lyase and caffeate-0-methyl transferase in bean callus tissue, during differentiation, were induced in a coordinated manner over a period of 21 days. Neither enzyme activity could be induced to increase when differentiation was repressed by the addition of abscisic acid to the induction medium. The formation of soluble phenols was also increased during the induction of differentiation but it was also increased by the transfer to induction medium of an old callus which could no longer differentiate. PAL activity was more directly correlated with xylogenesis and nodule induction than with the formation of soluble phenols. Peroxidase activity in the callus tissue was always high but the level decreased during the induction of differentiation. A new isoenzyme of peroxidase was induced during callus formation and this was probably a response to the presence of auxins in the medium used to grow the callus.     

Mercury inhibition of avian fatty acid synthetase complex.

(1) Subcutaneous or intra-abdominal injections of 8 mg of HgCl2/100 g body weight markedly depressed hepatic fatty acid synthetase activity of chicks at 1 h post-injection. The depression occurred despite the fact that the chicks continued to eat up until the time they were killed. Under these same conditions, the hepatic activity of acetyl-CoA carboxylase (EC 6.4.1.2) was not affected by HgCl2, while the activity of the mitochondrial system of fatty acid elongation was stimulated. (2) When 2-mercaptoethanol was included in the incubation medium for a highly purified preparation of fatty acid synthetase, 500 muM HgCl2 was required to show definite inhibition of the enzyme. When 2-mercaptoethanol was omitted, 50 muM HgCl2 was inhibitory and 100 muM HgCl2 abolished enzyme activity. (3) 2 mM dithiothreitol completely protected the purified fatty acid synthetase preparation from inhibition by 100 muM HgCl2. When dithiothreitol was added after the addition of enzyme to the mercury-containing medium, protection of the enzyme was not complete. (4) Dialysis of cytosol fractions from chicks injected with HgCl2 against 500 vol. of 0.2 M potassium phosphate buffer (pH 7.0) containing 1 mM EDTA and 10 mM dithiothreitol for 4 h at 4 degrees stimulated the fatty acid synthetase activity of the fractions. Dialysis of cytosol fractions from noninjected chicks under the same conditions was without effect on fatty acid synthetase activity. (5) These data support the hypothesis that the inhibitory effect of HgCl2 administered in vivo on hepatic fatty acid synthetase activity in chicks is mediated through the interaction of mercury with the sulfhydryl groups of the enzyme.     

A pleiotropic mutation results in cross-resistance to auxin, abscisic acid and paclobutrazol

Summary Mutant lines of Nicotiana plumbaginifolia resistant to the synthetic auxins 1-naphthaleneacetic acid (NAA) and indole-3-butyric acid (IBA) were isolated as germinating seedlings on selective medium. In each case, resistance was conferred by a single recessive nuclear mutation at one of 3 loci designated iba1, iba2 and iba3. Labelling studies with ¹⁴C NAA suggest that resistance was not due to changes in the uptake or metabolism of NAA. Plants homozygous for the iba1 mutation exhibit a syndrome of atypical germination and growth suggestive of a defect in the biosynthesis, metabolism or localization of abscisic acid. Wild-type seeds treated with gibberellin exhibit the same syndrome, including resistance to NAA and IBA. On the basis of these observations, we propose that auxin toxicity in seeds may be mediated by a block in gibberellin biosynthesis.Abbreviations ABA abscisic acid - GA gibberellic acid - IBA indole-3-butyric acid - NAA 1-naphthaleneacetic acid - p-cell protoplast-derived cell - 2,4-D 2,4-dichlorophenoxyacetic acid     

Gibberellic acid regulation of adventitious shoot formation from tuber discs of potato

Summary The formation of adventitious shoots from potato tuber discs explanted onto a modified Murashige and Skoog (MS) medium containingN ⁶-benzylaminopurine (BAP) (3.0 mg/l), and α-naphthaleneacetic, acid (NAA) (0.01 mg/l), was affected by gibberellic acid (GA). The presence of GA in the explant medium was required for shoot formation and 3×10⁻¹⁰ M GA appeared optimum. However, microscopic examination of the tissue protuberances on the surface of the tuber discs from which shoots arose revealed that GA inhibited the formation of shoot meristems. Tuber discs cultured for 6 wk on MS medium containing BAP and NAA without GA did not initiate adventitious shoots that could be determined visually, but microscopic examination of the tissue protuberances revealed the presence of numerous shoot meristems. Subsequent transfer of these tuber discs to medium with GA but without BAP or NAA resulted in the formation of shoots from 100% of the recultrued dises. Thus it appears that although GA inhibits shoot meristem initiation from potato tuber discs, it is required for shoot development once meristems are initiated. This is Journal Paper 8297 of the Purdue University Agricultural Experiment Station. The research was supported by Purdue University Agricultural Experiment Station Program Improvement Funds. Potato tubers were supplied by Wm. Gehring Farms, Inc., Rensselaer, Indiana.     

Exogenous phytohormones and the induction of plant galls by insects

The mechanism of gall induction by insects has remained elusive. Previous studies have met with limited success in attempting to induce galls by injection or application of chemical compounds. To determine whether an exogenous source of phytohormones plays a role in gall induction, we injected cytokinin (CK), auxin (IAA), gibberellic acid (GA), and abscisic acid (ABA) in various concentrations, ratios, and combinations into leaf petioles of Capsicum annuum L. cv. California Wonder (Solanaceae). We found that CK + IAA injections lead to gall-like growth in C. annuum. GA enhanced and ABA inhibited gall growth except in the presence of GA. Isopentenyl adenine (IP) was the most effective type of CK at inducing growth. Our work is consistent with the hypothesis that exogenous CK + IAA produced and supplied by insects leads to gall induction. We hypothesize that insects have obtained the capability to induce galls via acquisition of the biosynthetic pathways to produce IAA and trans-zeatin family CKs through microbial symbiosis or lateral gene transfer.     

Shoot growth in willow (Salix viminalis) in relation to abscisic acid, plant water status and photoperiod

The decline in growth rate of field-grown willow trees in Aberystwyth, U.K., began in mid-summer and was followed by the senescence and abortion of shoot tips. These events were not triggered by a decline in the length of the natural photoperiod but were coincident with low leaf water potentials that developed in summer. Transient increases in the abscisic acid (ABA) content of shoot tips were observed during the period of declining water potential. These increases were roughly coincident with the onset of growth decline and preceded abortion and senescence of shoot tips. Under controlled conditions growth of both rooted cuttings and potted plants was arrested by short days (8 h) without any increase in tip ABA levels. Growth of rooted cuttings under long days (16 h) was inhibited by exogenous ABA; this inhibition could be relieved by addition of gibberellic acid (GA3) to the nutrient solution. Growth of aseptically cultured apices was also inhibited by ABA; this inhibition was relieved by joint application of GA9 and zeatin riboside.