Role of plant growth regulators in host-pathogen relationships

The effect of indol-3-ylacetic acid, gibberellie acid, kinetin, abscisic acid and Ethrel on the growth of mycelium, sporulation and germination of spores ofFusarium culmorum of different pathogenicity to wheat seedlings was studied. The production of gibberellins, auxins, cytokinins, ethylene and growth inhibitors by these isolates was determined as well. It has been found that most pronounced and explicit effect on growth and development in fungi was produced by Ethrel which strongly inhibited these processes. ABA proved to be a strong growth and development stimulator, though to a different extent in different isolates. GA₃ strongly stimulated sporulation and spore germination in some isolates. The effect of IAA and K on growth and development in fungi was slight. More sensitive to growth regulators were the fungi in earlier stages of growth. No correlation between the pathogenicity of the isolates and their ability to produce growth regulators as well as between their susceptibility to exogenous growth substances in the processes of fungal growth and development was stated. Presented at the International Symposium “Plant Growth Regulators” held on June 18–22, 1984 at Liblice, Czechoslovakia.     

Cytokinins as inhibitors of root growth

The elongation of roots of wheat ( Triticum aestivum L. cv. Diamant II), flax ( Linum usitatissimum L. cv. Concurrent) and cucumber ( Cucumis sativus L. cv. Favör) seedlings in the dark was strongly inhibited by various native and synthetic cytokinins (kinetin, benzyladenine, isopentenyladenine, zeatin and their corresponding 9-ribosides). An inhibition of 50% was obtained for wheat roots with 3 · 10⁻⁹ M zeatin and for flax roots with 6 · 10⁻⁹ M isopentenyladenine. The ribosides were in all cases less inhibitory. The inhibition was reversed by various types of 'antiauxins' and 'antiethylenes' (such as structural auxin analogues, uncouplers, specific inhibitors of ethylene synthesis, free radical scavengers, inhibitors of ethylene action). These substances as a rule counteract also inhibitions caused by auxins. Auxins and cytokinins stimulate ethylene production synergistically, and the similar inhibitory effects of these two types of hormone can be understood if it is assumed that their effect is at least partly mediated through ethylene. The cytokinins must be considered as possible natural inhibitors and regulators of root growth.     

Effect of growth regulators on microspore embryogenesis in coconut anthers

The effect of growth regulators on induction of androgenesis in coconut was investigated using seven different growth regulators at various concentrations and combinations. Three auxins (1-naphthalene acetic acid—NAA, indoleacetic acid—IAA, picloram) and three cytokinins (2-isopentyl adenine-2-iP, kinetin, zeatin) were tested either alone or in combination with 2,4-dichlorophenoxyacetic acid (2,4-D), using modified Eeuwens Y3 liquid medium as the basal medium. Among the tested auxins, 100 μM NAA in combination with 100 μM 2,4-D enhanced the production of calli/embryos (123) whereas IAA and picloram showed negative and detrimental effects, respectively, for androgenesis induction over 100 μM 2,4-D alone. Kinetin and 2-iP enhanced the production of calli/embryos when 100 μM 2,4-D was present in the culture medium. Both cytokinins at 10 μM yielded the highest frequencies of embryos (113 and 93, respectively) whereas zeatin (1 or 2.5 μM) had no impact on microspore embryogenesis. When calli/embryos (produced from different treatments in different experiments) were sub-cultured in somatic embryo induction medium (Y₃ medium containing 66 μM 2,4-D), followed by maturation medium (Y₃ medium without growth regulators) and germination medium (Y₃ medium containing 5 μM-6-benzyladenine—BA and 0.35 μM gibberellic acid—GA₃), plantlets were regenerated at low frequencies (in most treatments ranging from 0% to 7%).     

Mixture screening and mixture-amount designs to determine plant growth regulator effects on shoot regeneration from grapefruit (<Emphasis Type="Italic">Citrus paradisi</Emphasis> macf.) epicotyls

The objective of this study was to improve shoot regeneration from grapefruit. Because many commercially grown citrus types are apomictic, important in vitro applications such as Agrobacterium-mediated transformation commonly use epicotyl explants from in vitro seedlings; thus, adequate adventitious shoot production is an important prerequisite for efficient use of these applications. Eight plant growth regulators were studied—six cytokinins (6-benzylaminopurine, kinetin, zeatin trans-isomer, 6-[γ,γ-dimethylallylamino] purine, zeatin riboside trans-isomer and meta-topolin) and two auxins (α-naphthalene acetic acid and indole-3-acetic acid). An iterative design strategy was followed that included mixture and mixture-amount experimental designs suitable for resolving proportional and concentration effects; in vitro effects of cytokinins and auxins are affected by both proportion and concentration. One-centimeter-long explants were excised from the epicotyl of etiolated, in vitro-grown seedlings. Explants were placed onto experimental formulations and cultured in growth cabinets at 27°C over 6 wk, which included 2 wk in the dark followed by 4 wk in the light. The results indicated that (1) 6-benzylaminopurine or zeatin riboside were the most effective cytokinins for inducing shoot regeneration in citrus; (2) zeatin riboside singly or in combination with indole-3-acetic acid resulted in the highest quality, the greatest number of explants with buds/shoots, and the greatest shoot number; and (3) 6-benzylaminopurine and indole-3-acetic acid improved shoot regeneration vs. 6-benzylaminopurine at a considerably lesser cost than zeatin riboside and indole-3-acetic acid.     

The influence of growth regulators on membrane permeability in cultures of winter wheat cells

The effect of plant growth substances (IAA, 2,4-D, zeatin, kinetin, zearalenone) were studied on membrane properties of the cells of embryogenic (E) and non-embryogenic (NE) calli derived from immature inflorescences (inf) or embryos (emb) of winter wheat. Calli initiated from inflorescences show higher permeability. The ion leakage from cells of E calli was higher than from cells of NE calli. Growth regulators were used in concentrations of 2-30 mg/l (about 10-140 microM). All tested growth substances increased ion leakage from NE emb cells, IAA, zeatin and kinetin being most effective. In NE inf cells the effect of growth substances was similar as in NE emb, but much weaker. In E cells of both types (inf and emb) growth substances decreased ion leakage. Changes in the leakage of potassium and calcium ions were similar to those in total ion leakage. The uptake of labelled auxins (IAA and 2,4-D) was higher in NE cells (especially in NE inf) than in E cells. The endogenous level of IAA was higher in E cells than in NE cells and in inf cells than in emb cells. The importance of auxin in determining permeability of cell membranes is discussed.     

Some effects of plant growth regulators on tissue cultures of the marine red alga Agardhiella subulata (Gigartinales,Rhodophyta)

We examined whether auxins and cytokinins, either singly or in combination, stimulate cell division in tissue cultures of a red seaweed. Our experimental model consisted of filamentous and callus-like growths that developed from cross-sectional discs cut from young branches of Agardhiella subulata. Plant growth regulators were added to the medium to give combinations of an auxin with a cytokinin over a range of concentrations (1 µg L^–1 –10 mg L^–1). Several mixtures of auxins and cytokinins, as well as some single auxins, cytokinins and phenolics, stimulated cell division and growth in the tissue cultures beyond that of controls. The treatments that were effective included: phenylacetic acid/zeatin; phenylacetic acid/6-benzylaminopurine; -naphthaleneacetic acid/zeatin; 2,4,5-trichlorophenoxyacetic acid/6-benzylaminopurine; and indoleacetic acid/kinetin. High concentrations of cytokinins (i.e. 10 mg L^–1) inhibited the regeneration of plants in some of the cell cultures. These results provide further evidence that growth regulators can be used for the tissue culture of seaweeds and for the study of developmental phenomena in these plants.     

Changes in the content of endogenous auxins in apical buds ofchenopodium rubrum L. Induced with respect to the endogenous rhythm in capacity to flower

The content of endogenous auxins was examined in apical buds ofChenopodium rubrum plants induced by a photoperiodic cycle of 16h darkness and 8h light followed by a dark period of various duration so as to correspond with either maximal or minimal flowering response in the endogenous rhythm in capacity to flower initiated by the photoperiodic treatment. Apical buds of potentially generative plants contained less auxins than apical buds of plants which remained in the vegetative state. Apical buds from plants treated with kinetin (1. 10^-3 M) and therefore remaining in the vegetative state showed an auxin level comparable to that of untreated plants exhibiting minimal flowering response irrespective of the duration of the second dark period. Plants cultivated on a sucrose solution (0.6 M) during the second dark period became generative even at the normal minimum of flowering. The auxin content of the apical buds was low, similarly as in untreated plants induced for a period leading to maximal flowering response. On the other hand, apical buds from plants grown on sucrose solution during a dark period leading to the manifestation of maximal flowering response showed a relatively high auxin content comparable to that found in untreated plants which had obtained a more extended induction by three photoperiodic cycles. The results are discussed with respect to the possible role of endogenous auxins in the regulation of the changes in growth correlations occurring in the shoot apex during photoperiodic induction and in the expression of the competence to flower.     

Exogenous regulators in the mycelium ofPleurotus ostreatus after exogenous application

The mycelium of the fungusPleurotus ostreatus was cultured on a synthetic medium with growth regulators. Best growth was observed on media with 100 ppm IAA, 200 ppm GA₃ and 200 ppm of kinetin. Growth of terminal hyphae and the appearance of colonies differed characteristically. Auxin bio-assays showed that the content of substances in younger mycelium slowly increased during cultivation without conspicuous changes in the medium. After exogenous application of IAA the content of auxins in younger mycelium increased and the content of IAA decreased gradually in the culture medium. Gibberellin bio-assays showed their content in the mycelium fell during cultivation and while it increased in the medium. Application of GA₃ to the medium leads to a significant increase in these substances in the young mycelium and in the medium their level remains the same. Cytokinin bio-assays showed their content to vary during cultivation. An addition of kinetin to the nutrient medium results in an increase of cytokinins in the mycelium while in the medium the level of cytokinins is maintained. Addition of one substance thus influences the levels of other endogenous regulators.     

High-frequency in vitro propagation of the endangered species <Emphasis Type="Italic">Tuberaria major</Emphasis>

A novel protocol suitable for the micropropagation of the endangered species Tuberaria major using seedlings as explants is reported. Using this protocol, we studied the effects of explant type (apical shoots and nodal segments) and cytokinins [6-benzyladenine (BA), kinetin, and zeatin (ZEA)] on shoot proliferation. Explant type significantly influenced the proliferation frequency and mean number of shoots, with nodal segments showing a higher proliferation capacity. The mean number of shoots was significantly higher when the explants were cultured in half-strength (1/2) MS medium supplemented with 0.2 mg l⁻¹ BA (6.83 ± 0.77 shoots) or ZEA (6.55 ± 0.71 shoots). The shoots showed a great rooting capacity that was significantly influenced by the concentration of MS macronutrients but not by the concentration of auxins. The highest rooting frequencies (97–100%) were obtained in 1/2 MS medium with or without plant growth regulators. The plants obtained were easily acclimatized to ex vitro conditions, with 97% surviving after 6 weeks. The micropropagated plants were successfully reintroduced into their natural habitat and exhibited normal development. In conclusion, our culture protocol, with efficient seed germination, subsequent multiplication of nodal explants using ZEA at 0.2 mg l⁻¹, and successful ex vitro establishment of well-rooted plantlets on 1/2 MS medium, provides a simple and reliable methodology for the large-scale propagation of T. major, thereby contributing to germplasm preservation of this endangered species.     

Stimulation of the formation of fruiting bodies of the fungusLentinus tigrinus (Bull.) Fr. by growth regulators

Growth regulators, indole-3-acetic acid (IAA), gibberellic acid (GA₃), and kinetin (KIN), were used in different concentrations to stimulate the initiation and further development of the fruiting bodies of the fungusLentinus tigrinus. Vegetative mycelium of the fungus was cultivated on cellulose cylinders soaked with a synthetic nutrient solution or with a 3% malt extract. When the mycelium covered the surface of the cylinders, further cultivation was carried out in graduated concentrations of the growth regulators mentioned above. The number of developed fruiting bodies showed that the optimum IAA and GA₃ concentrations were in both media 300 p.p.m. The optimum concentration of kinetin was 400 p.p.m. The individual growth regulators influenced characteristically also the shape of the fruiting bodies and changed in them the natural level of endogenous growth regulators. The addition of IAA into the medium raised the level of endogenous auxins in the cap. The presence of gibberellic acid and of kinetic in the medium resulted in an increase in the level of these regulators in the fruiting bodies.     

Changes in the level of endogenous gibberellins and auxins in apical buds ofChenopodium rubrum L. after application of growth substances reversing the effect of (2-chlorethyl)-trimethylammonium chloride (CCC) on flowering

The application of CCC at concentrations inhibiting flowering ofChenopodium rubrum reduces the level of endogenous gibberellins in the apical buds of the plants. The effect of CCC may be reversed by appropriate concentrations of gibberellin (GA-), indole acetic acid (IAA) or kinetin. Kinetin applied to the apical bud during floral induction reduced the level of endogenous gibberellins similarly as CCC and if both CCC and kinetin were applied simultaneously their action was additive. On the other hand IAA applied under the same conditions increased the level of endogenous gibberellins and after joint application of CCC and IAA their level was the same as in untreated control plants. After application of CCC during floral induction the level of endogenous auxins did not change markedly but an active substance “x” appeared on the chromatograms of indole compounds. This substance was found also after simultaneous application of GA- and CCC but not after joint application of CCC and kinetin. If follows from our results that the same morphological phenomenon (flowering) can take place in plants considerably differing as to their level of endogenous growth substances. The ratio of different growth substances is obviously more important than the actual level of the single substances.     

Effect of exogenous plant growth regulators on in vitro seed growth,embryo development,and haploid production in a cross between H. vulgare (L.) and H. bulbosum (L.)

Exogenous plant growth regulators are known to increase the efficiency of interspecific and intergeneric crosses. In vitro floret culture provides a defined system for assessing the importance of various plant growth regulators on the determinants of haploid production efficiency (seed set, embryos per seeds, and plants per embryos) in Hordeum vulgare × Hordeum bulbosum crosses. The individual and combined effects of three plant growth regulators (2,4-D, GA₃ and kinetin) on in vitro seed growth, embryo development and haploid production efficiency were tested in floret culture of the cross H. vulgare, cultivar Klages × H. bulbosum. All treatments, except kinetin alone, produced larger seeds and more embryos/100 seeds than the control (no plant growth regulator). 2,4-D alone was superior to GA₃ alone in haploid production efficiency (70.6 vs. 51.5) as measured by the number of plants regenerated/100 florets pollinated. Although kinetin +2,4-D+GA₃ produced the largest seeds and embryos, no advantage over 2,4-D alone was observed in haploid production efficiency. 2,4-D alone or kinetin +2,4-D are recommended for the purpose of barley haploid production in floret culture using the bulbosum method.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid     

Effect of gibberellin and kinetin on the regeneration ability ofFucus vesiculosus L.

The influence of gibberellic acid (GA³) and kinetin (6-furfurylaminopurine) on the regeneration ability of the basal and apical thallus fragments ofFucus vesiculosus L. was examined. The naturally occurring gibberellin and kinetin-like substances in these thallus fragments were also studied. It was found that exogenously applied GA³ markedly increased the number of adventitious branches formed on the cut surface of the thallus fragments taken from the apical parts of plants. The concentration of 0.001 mg GA³ I-1 proved to be the most effective. The growth promoting effect of GA³ was increased by simultaneous action with kinetin. In experiments in which the fragments of the basal parts of the thallus were treated with GA³, as a rule a slight growth inhibition was observed. The growth responses of the investigated plant tissues to gibberellin and kinetin varied according to season. Usually their susceptibility to the applied plant hormones was greater in spring than is summer. The shifts in growth reaction were related to the seasonal changes in the content of endogenous gibberellin and kinetin-like substances in the investigated parts of the thallus. It is suggested that growth regulators of the gibberellin and cytokinin type are involved in the regeneration processes inFucus.     

Evaluation of the effect of three growth regulators in the germination of <Emphasis Type="Italic">comparettia falcata</Emphasis> seeds under <Emphasis Type="Italic">in vitro</Emphasis> conditions

Summary The effect of 3-indoleacetic acid (IAA), 6-furfurylaminopurine (kinetin), and gibberellic acid (GA₃) on germination of the orchid Comparettia falcata was evaluated in a factorial experiment (4×4×4) with Murashige and Skoog (1962) basal medium. It was established that seeds of this orchid could be maintained under aseptic conditions as long as the necessary nutrients and appropriate concentrations of growth regulators were provided. Of the three growth regulators used, IAA significantly decreased seed germination of Comparettia falcata. There was a synergistic effect in the kinetin:GA₃ combination that produced a positive response in both percentage seed germination and development of seedlings. This study describes a single medium-based protocol able to achieve more than 160000 seedlings within 21 wk, starting from a single capsule, sufficient for both large-scale propagation and in vitro conservation of this threatened orchid.     

Effect of growth regulators onVicia faba plants irrigated by sea water Leaf area,pigment content and photosynthetic activity

The antagonistic effects of some growth regulators [i.e. indol-3-yl-acetic acid (IAA), gibberellic acid (GA₃) or kinetin] on stress imposed by sea water on leaf area, pigment and photosynthetic activity in leaves of broad bean plants at different stages of development were investigated. Seed priming with GA₃ alleviated either partially or completely the effects induced by the two levels of sea water (10 and 25 %) used on leaf area at all experimental stages. However, IAA, GA₃ and kinetin inhibited leaf growth by themselves in almost all measurements. Seed pretreatment with kinetin alleviated the inhibition of pigment production in sea water-irrigated plants. Furthermore, GA₃ or kinetin nullified the deleterious effects imposed by irrigation with sea water particularly the high level (25 %) on photosynthetic¹⁴CO₂ fixation.     

Growth and pigment content of wheat as influenced by the combined effects of salinity and growth regulators

A field experiment was conducted to investigate the effects of presoaking the wheat grains (Triticum aestivum L.) in different levels of salinity (33 or 66 mM) and in growth regulators (indolyl-3-acetic acid, IAA at SO g m^-3, gibberellic acid, GA₃ at 100 g m^-3, or kinetin at 100 g m_-3) on the shoot growth and pigment content of the developing wheat flag leaf. Salinity at 33 or 66 mM led to an insignificant increase in the fresh and dry masses as well as in the shoot diameter and shoot length, but it attenuated the flag leaf area. In the majority of cases, salinity increased the chlorophyll (Chla, Chlb) and carotenoid contents as well as the number of chloroplasts per a mesophyll cell. The growth in the wheat shoot of the saline-treated plants was, in general, stimulated in response to presoaking the grains in kinetin or GA₃. On the other hand, IAA + salinity led to a negligible effect on the growth in the wheat plants particularly at the early stages of growth. The presoaking of grains in NaCl at 33 mM + IAA or 66 mM + kinetin induced a marked increase in the pigment content of the wheat flag leaf particularly at the early stages of growth. The interaction between salinity and phytohormones increased the number of chloroplasts; kinetin was the most effective.     

Flower formation in excised tobacco stem segments; I. Methodology and effects of plant hormones

The formation of flowers has been studied in stem tissue excised from flowering plants of Nicotiana tabacum variety Wisconsin No. 38, and cultured in vitro on Murashige and Skoog nutrient medium. A procedure for quantitative evaluation of factors influencing floral expression has been developed and effects of the growth substances, indole-3-acetic acid (IAA), kinetin and gibberellic acid (GA₃), on the process are reported.     

Effect of Vernalization and Red Light Illumination of Seedlings of Bread Wheat (Triticum aestivum L.) on the Temperature Profile of cAMP Phosphodiesterase Activity

Phenotypic manifestations of Vrn(vernalization) and Ppd (photoperiod) genes responsible for transition of bread wheat Triticum aestivumL. to generative growth (flowering) are mutually related. Since the mechanism of phytochrome-induced photoperiodism involves the enzymes of cyclic adenosine monophosphate metabolism, and phosphodiesterase in particular, we tested involvement of phosphodiesterase in the process of winter wheat vernalization and formation of flowering competence in alternate wheat requiring a long photoperiod but no vernalization for its transition to flowering. We studied temperature dependence of phosphodiesterase activity in vernalized and unvernalized winter wheat on the one hand and in etiolated and red light illuminated seedlings of alternate wheat on the other hand. Short-term experiments demonstrated that red light illumination is similar to long photoperiod by the effect on the long-day plants. Both influences induced a pronounced inversion of the temperature profile of phosphodiesterase activity in the 28–45°C range. We propose that phosphodiesterase is involved in vernalization and can serve as a receptor of low temperature in winter wheat. Changed temperature profile is a radical control mechanism of phosphodiesterase activity in response to the influences (red light and vernalizing temperatures) responsible for competence of various bread wheat forms for generative growth.     

Indole auxins in spinach plants grown in long and short days

The endogenous auxin-like substances were analyzed in the shoot extracts of young spinach seedlings, exposed to photoperiodic induction. At least eight indole auxins were found. One of them was identified as tryptophan, the other one is most probably IAA. The plants grown in long days had a higher level of ether soluble auxins than the controls in short days. Separate extractions of plants after each of the eight inductive days showed that the auxin content was not constant, but subjected to irregular oscillations. However, parallel oscillations were also found in control plants grown in short days. Staminate plants were found to contain more endogenous auxins than the pistillate ones. It is concluded that the quantitative changes in auxins during the photoperiodic induction are probably not related to flowering, but to some other growth process, common to all plants in that phase of growth. The higher level of auxins in staminate plants may be the cause of their faster elongation before the onset of flowering.     

Influence of growth regulators on direct embryo formation from leaf explants of Phalaenopsis orchids

Leaf explants of two Phalaenopsis, P. amabilis and P. Nebula, were used to test the effects of auxins (2,4-D, IAA, IBA, NAA), cytokinins (2iP, BA, kinetin, TDZ, zeatin), GA₃, ancymidol, polyamines (putrescine, spermine, spermidine), ACC, AgNO₃ and CoCl₂ on the amount of direct embryo formation on different leaf locations (the cut end, the adaxial side, the abaxial side and the leaf tip). The results showed that there was a genotypic effect on direct embryo formation induced by cytokinins that 13.32 μM BA and 4.92 μM 2iP was the most effective in P. amabilis and P. Nebula, respectively. Besides, explant position highly affected embryogenic competence of leaf cells in both species that the cut end showed highest embryogenic response, the adaxial side was the second, and then the abaxial side and the leaf tip. Altogether, cytokinins tested were all effective in both species, and ACC at 20 μM had 35% of embryogenic response in P. amabilis. However, auxins, GA₃, ancymidol and polyamines were inhibitory in both species.