Gibberellic acid-induced cell elongation in cotton suspension cultures

Gibberellic acid (GA₃) causes cell elongation in cotton suspension cultures derived from cotton ovule callus tissue of both auxin-dependent and-independent lines. Cell elongation was more pronounced in auxin-dependent cultures. Cells were cultured for a period of 14 days but differences in cell lengths could be detected after 6 days in culture. Cell elongation took place in cultures in which GA₃ was present throughout the culture period or only for the first 3 days. Auxins and cytokinin alone or in the presence of GA₃ did not promote cotton cell elongation above the value for the treatment with GA₃ alone.Names of companies of commercial products are given solely for the purpose of providing specific information; their mention does not imply recommendation or endorsement by the U.S. Department of Agriculture over others not mentioned.     

Essai de remplacement de la réfrigération vernalisante par des applications d’acide gibbérellique chez la jusquiame noire bisannuelle (hyoscyamus niger L.)

Applications of various amounts of GA₃ onto the shoot apex of biennial rosette-plantHyoscyamus niger L., exposed to long days and non vernalized, caused internodes formation and stem elongation. The stem length was proportional to the amount of GA₃ applied. Number and length of developed internodes and the whole length of the shoot were maximal in plants treated with a greater amount of GA₃. In the end, stem elongation stopped and plants formed perchedrosettes without flowering. Hence, gibberellic acid participates in mechanisms of flowering only by indirect effect on stem elongation and not directly on flower formation itself.     

Effect of gibberellic acid on the cyanobacterium <Emphasis Type="Italic">Nostoc linckia</Emphasis>

We investigated the influence of gibberellic acid (GA₃; 0, 1, 10, and 100 μM) on Nostoc linckia culture at 7, 14, and 21 days. The fresh and dry weight of N. linckia was increased considerably by the 10 and 100 μM GA₃ treatments. A reduction in heterocyst frequency was observed in cultures treated with 1 and 10 μM GA₃. Adding GA₃ to N. linckia culture had a little effect on cell size. The amount of chlorophyll a and carotenoids decreased at all concentrations of GA₃. The amount of phycocyanin increased up to twofold in 7-day-old culture treated with 1 μM GA₃, and similar changes were observed for allophycocyanin and phycoerythrin content after 7 days. The effect of GA₃ on reducing sugar content was different and was dependent on the growth period. A reduction in soluble sugar content was detected after GA₃ application in 7- and 14-day-old cyanobacteria. Cultures treated with GA₃ had a higher protein content after 14 days and a lower protein content after 7 and 21 days, and reduced nitrogenase activity after 7, 14, and 21 days. Our data show that GA₃ application can be a suitable and inexpensive way to increase N. linckia biomass and phycobiliprotein production.     

Enhanced plumbagin accumulation in embryogenic cell suspension cultures of Plumbago rosea L. following elicitation

This study focused on enhancing the production of plumbagin, an anticancer compound, in embryogenic cell suspension cultures of Plumbago rosea. Elicitation techniques have been reported to enhance plumbagin production. Cell suspension cultures raised from embryogenic calli induced from in vitro leaf explants were exposed to different concentrations of jasmonic acid, yeast extract and different auxin combinations. Influence of these on cell growth, biomass and plumbagin production was studied. To our knowledge this is the first report on elicitation of embryogenic cell suspension cultures of P. rosea for enhanced plumbagin production. Elicitor treated suspension cultures exhibited decreased culture viability and increased plumbagin synthesis. A maximum of 5.59-fold enhancement of plumbagin production was observed in cultures added with 1 mg L^?1 naphthalene acetic acid after 6 days of incubation. Viability of cultures decreased with increased concentration of elicitors and prolonged incubation period. Application of elicitors in cell suspension cultures induces defense related responses which lead to increased secondary metabolite production for making the cells adapt to the situation. If the stressed condition persists or is in intolerable level this will eventually lead to programmed cell death and loss of culture viability.     

The action of exogenous gibberellic acid on polysome formation and translation of mRNA in germinating castor-bean seeds

The amount of protein synthesis in germinating castor-bean seeds has been estimated by the quantitative and qualitative exmainatin of polysomes from the seeds in the presence and absence of gibberellic acid (GA₃). Careful optimisation of polysome extraction procedures was required to minimise the ribonuclease activity in the extracts. Ribonuclease activity in seed extracts increased fourfold over the first 5 d of germination. Gibberellic acid stimulated polysome formation about twofold during the first 4 d of germination. It also stimulated the amount of mRNA associated with polysomes by about twofold during the first 3 d of germination. Between days 1 and 5 of germination, polysome formation was primarily limited by mRNA availability. During the period 0–24 h, polysome formation was independent of mRNA levles. The increase in enzyme activities stimulated by GA₃ was probably the result of an increase in the amount of cellular mRNA. No evidence was obtained for an action of GA₃ on translation other than on the increased production of RNA. Examination of the recruitment of isocitrate-lyase mRNA into polysomes showed that GA₃ did not specifically stimulate production of this enzyme.     

Effects of concentration and treatment duration upon dwarf pea response to gibberellic acid root treatments in solution culture

Gibberellic acid (GA₃) root treatments stimulated internode elongation of hydroponically grown dwarf pea seedlings (Pisum sativum L.,cv. Little Marvel) When the GA₃ concentration in the solution was at least 2.9 M.Both GA₃ concentration and the duration of the root-treatment period significantly affected internode elongation. This is attributed to a limited availability or saturation of active sites for gibberellin-induced cell elongation. The amount of GA₃ taken up through the roots in 1 day from a 29 M GA₃ solution apparently equaled or exceeded the amount which could be metabolized during the first four days after treatment, although higher concenrations and longer treatment periods produced a more prolonged response, conceivably due to 1) initial saturation of gibberellin active sites, 2) storage of surplus gibberellin in the plant, and 3) subsequent utilization of the stored gibberellin. GA₃-induced stem elongation in hydroponically grown Little Marvel peas seemed to be limited initially by apparent saturation of active sites when the GA₃ concentration exceeded 29 M.     

Formation of GA20 glucosyl conjugates in seedlings ofVicia faba

[³H]GA₂₀ (1)¹, fed toVicia faba seedlings, was converted to [³H]GA₂₀ glucosyl ester (5) and [³H]GA₂₀-13-0-glucoside (6). The GA₂₀ glucosyl ester (5) was identified by HPLC-RC and by GC-MS of GA₂₀-Me formed by transesterification of (5). The [³H]GA₂₀-Me was crystallized to constant specific radioactivity with authentic GA₂₀-Me. On HPLC-RC the GA₂₀-13-0-glucoside (6) was shown to have the same retention time as an authentic sample. Subsequent enzymic hydrolysis gave a product with an HPLC retention time identical to that of authentic GA₂₀ (1).     

Gibberellic-acid-regulated expression of α-amylase and six other genes in wheat aleurone layers

The effect of gibberellic acid (GA₃) on gene expression in wheat aleurone cells has been characterised. In-vitro translation of polyadenylated RNA indicated that α-amylase and other messenger-RNA (mRNA) species increase in relative concentration in GA₃-treated tissue. At least one mRNA species declines in relative level in response to GA₃. There is also a GA₃-dependent, four-fold increase in the level of polyadenylated RNA. This effect is largely the result of increased levels of many mRNA species which are also present in untreated tissue. Seven GA₃-induced polyadenylated RNA species including the Amyl α-amylase gene product have been cloned as complementary DNA in the plasmid pBR322. These cloned DNAs have been used as hybridisation probes to show that the GA₃-induced increase in α-amylase mRNA is more prolonged than the accumulation of the other GA₃-regulated mRNA species. A polyadenylated-RNA sequence showing reduced concentration in GA₃-treated tissue has also been cloned.     

Enzymic glucosylation of gibberellins

Starting from the well-known conversion of exogenously applied free gibberellic acid (GA₃) to its 3(O)-glucoside by intact immature fruits of runner beans (Phaseolus coccineus L.), a protein fraction has been prepared from this plant material possessing glucosylating activity towards GAs. This glucosyltransferase is located in the pericarp only and utilizes preferably UDP-glucose as a sugar donor. The product formed enzymically from GA₃ and UDP-glucose could be identified by derivatization and comparison with the authentic compound to be GA₃-3(O)-glucoside. Among 15 native or chemically modified GAs, the enzyme glucosylates only GA₃ and to a lower extent GA₇ and GA₃₀, indicating a high enzyme specificity with regard to the A ring of gibberellins. The physiological significance of the enzymic GA₃-3(O)-glucoside formation inPhaseolus coccineus is not clear, since this glucoside is not known to be endogenous in this plant. The enzyme preparation did not glucosylate substances of phenolic structure, such as hydroquinone, aesculetin, and quercetin. Glucosylation of GA₃ was achieved also by enzyme preparations fromVigna sinensis and from cell suspension cultures ofDigitalis purpurea. A number of other plant materials showed no activity.     

Cell elongation and cell division in elongating lettuce hypocotyl sections

The roles of cell division and cell elongation in the growth of sections excised from hypocotyls of lettuce (Lactuca sativa L. cv. Arctic) were investigated. Elongation of sections incubated in the light is inhibited compared to dark-grown sections and this inhibition is reversed by gibberellic acid (GA₃). The elongation of both dark-grown and GA₃-treated, light-grown sections can be enhanced by 10mM KCl. Under all conditions of incubation, elongation growth is greatest in the uppermost quarter of the hypocotyl section while the basal quarter does not elongate. In darkness the two apical segments of sections marked into four equal parts grow at the same rate, while in light, growth of the apical segment exceeds that of the second segment. Cell division in cortical or epidermal cells, as measured by mitotic index or cell number, is not affected by illumination conditions nor by GA₃ or KCl treatments. Although -irradiation and FUDR pretreatment eliminate or cause a marked reduction in cell division in the excised hypocotyl, sections from seeds irradiated with -rays or incubated in 5-fluorodeoxyuridine elongate in response to GA₃ and KCl treatment as do sections from non-pretreated controls. Therefore, since neither GA₃ nor darkness affect celldivision activity and since treatments which eliminate or significantly reduce cell division do not affect growth, we conclude that the effect of GA₃ and darkness in this material is to increase cell elongation.Abbreviations FUDR 5-fluorodeoxyuridine - GA(s) gibberellin(s) - GA₃ gibberellic acid     

The biosynthesis of the gibberellin precursorent-kaurene in cell-free extracts and the endogenous gibberellins of Japanese morning glory in relation to seed development

The endogenous levels of gibberellins (GAs) determined by a combined HPLC-bioassay procedure and the formation ofent-kaurene, an immediate GA precursor, in cell-free extracts were studied in relation to seed development inPharbitis nil Choisy cv. Violet. Three biologically active GA fractions were obtained, tentatively identified as GA₃, GA₅/ GA₂₀, and a GA fraction, possibly GA₁₉ and/or GA₄₄, which all increased in activity during early seed development and subsequently declined during maturation of the seeds. The total endogenous GA level reached its maximum at 19 days after anthesis, just before the seeds had attained their maximum fresh weight at about 23 days after anthesis. Similarly, theent-kaurene synthesizing capacity showed a rapid increase during the period of rapid growth of the seeds, followed by a decline during maturation. A direct relationship between the endogenous GA levels and theent-kaurene synthesizing capacity of a particular tissue was indicated.     

Gibberellin biosynthetic pathway and the physiologically active gibberellin in the shoot ofCucumis sativus L.

[²H₂]Gibberellin A₂₄ (GA₂₄) and [²H₄]-GA₉ were applied to the apices of normal-type cucumber (Cucumis sativus L. cv. Yomaki) seedlings treated with uniconazole, an inhibitor of GA biosynthesis. The metabolites from these feeds were identified by full-scan gas chromatography-mass spectrometry (GC-MS) to confirm the conversions of [²H₂]GA₂₄ to [²H₂]GA₉ and of [²H₄]GA₉ to [²H₄]GA₄. The results show that GA₄ is biosynthesized from GA₂₄ via GA₉. In a cucumber hypocotyl elongation bioassay using cv. Yomaki, prohexadione (DOCHC), an inhibitor of 2-oxoglutaratedependent dioxygenase, inhibited the hypocotyl elongation caused by application of GA₉, while DOCHC enhanced the elongation caused by application of GA₄. These results indicate that GA₄ is a physiologically active GA and that the activity of GA₉ is due to its conversion to GA₄ in cucumber shoots.     

Identification of gibberellins from sugarcane plants

Five GAs, GA₁, GA₃, GA₁₉, GA₂₀, and GA₂₉, were identified in extracts from mature leaf and shoot apical meristem of flowering and non-flowering sugarcane (Saccharum spp. hybrids) by combined GC/MS. The presence of ABA was also confirmed.     

Endogenous gibberellins and kauranoids identified from developing and germinating barley grain

Several gibberellins (GAs) and kauranoids were identified in extracts of barley (Hordeum vulgare) by combined capillary gas chromatography-mass spectrometry (GC-MS). A partially purified acidic ethyl acetate extract from 21-day postanthesis developing barley grain (cv. Proctor) contained GA₁ (trace), GA₄ (trace), GA₈ (trace), GA₁₂, GA₁₇, GA₂₀ (tentative) (trace), GA₂₅, GA₃₄, GA₄₈, 18-hydroxy-GA₄, 12β-hydroxy-GA₉, and 18-hydroxy-GA₃₄ (tentative). A hydrolyzed butanol extract contained GA₁₇, GA₂₀, GA₄₈, and 18-hydroxy-GA₃₄ (tentative). An acidic ethyl acetate extract from 3-day-old germinating barley grain (cv. Maris Otter) contained GA₁, GA₃ (possibly a contaminant), GA₁₇, GA₁₉, GA₂₀, GA₃₄, GA₄₈, and 18-hydroxy-GA₃₄ (tentative). A hydrolyzed butanol extract contained GA₃₄, GA₄₈, and 18-hydroxy-GA₃₄ (tentative). In germinating grain, levels of all GAs were very low. Two hydroxylated kauranoic acids and a number of other kauranoids were also detected in the above extracts. 1β-Hydroxylated GAs previously found in wheat were not found in barley in this study.     

Gibberellin structure and florigenic activity in Lolium temulentum,a long-day plant

Structural requirements for florigenic activity among gibberellins (GAs) and GA derivatives, including several new ones, applied once to leaves of Lolium temulentum, were examined. The compounds were applied to plants kept either in non-inductive short days (SD) or exposed to one inductive long day (LD). Inflorescence initiation and stem-elongation responses were assessed three weeks later. Among the GAs used, the range in effective dose for inflorescence initiation was more than 1000-fold, but substantially less for stem elongation. Some GAs promoted both stem elongation and inflorescence initiation, some promoted one without the other, and some affected neither. The structural features enhancing florigenic activity were often different from those enhancing stem elongation. Except in the case of 2,2-dimethyl GA₄, a double bond in the A ring at either C-1,2 or C-2,3 was essential for high florigenic activity, though not for stem elongation. A free carboxy group was needed for both. Inflorescence initiation in Lolium was enhanced by hydroxylation at C-12, ?13 and ?15, whereas hydroxylation at C-3 reduced the effect on inflorescence initiation but increased that on stem elongation. A 12β-hydroxyl was more effective than the α epimer for inflorescence initiation whereas the reverse was true for stem elongation. Although such differential effectiveness of GAs for inflorescence initiation and for stem elongation could reflect differences in uptake, transport or metabolism, we suggest that it is indicative of specific structural requirements for inflorescence initiation.     

Relationship between gibberellic acid and water amount in the cotton seed

The role of endogenous GA₃ and its application to seed development in two cotton genotypes Hybrid-6 (H-6) (big seeds) and Gujarat cotton 13 (G. Cot) (small seeds) was studied. Kernel and seed coat were subjected to growth analysis in terms of dry weight, water amount, and rates of dry matter accumulation and water uptake. H-6 kernel had manifold higher dry weight and water amount than G. Cot. Seed coat of both genotypes had similar dry weight at maturity, but the maximum rates of dry matter accumulation and water uptake were distinctly higher in H-6. According to growth analysis, development of seed kernel and coat was subdivided into four phases, i.e., cell division, cell elongation, dry matter accumulation and maturation. Endogenous GA₃ level was estimated in kernel and seed coat by indirect ELISA using antibodies raised against GA₃. GA₃ amount per seed components was higher in the seed kernel of H-6 than of G. Cot, except 33 and 36 days after anthesis in kernel. H-6 seed coat had the higher amount of GA₃ during cell division phase than that of G. Cot. Close correlation between in vivo GA₃ level and water amount was recorded in both seed components. With GA₃ or GA₃ + NAA treatments in ovule culture, higher promotion in dry weight, water amount and seed size was noted in G. Cot than in H-6 suggesting that G. Cot is more deficient in endogenous GA₃. The greatest stimulation of parameters studied was obtained in ovule culture with GA₃ + NAA. When GA₃ or GA₃ + NAA was applied, initial significant difference in water amount and seed size was nullified. Data presented in this study indicated that GA₃ regulates cell expansion through the water uptake by cotton seed.     

Stem elongation of ornamental bromeliad in tissue culture depends on the temperature even in the presence of gibberellic acid

Acanthostachys strobilacea Link, Klotzsch, & Otto is an ornamental bromeliad native to Brazilian Atlantic Forest that naturally exhibits a rosette growth pattern. According to the temperature conditions of the in vitro culture, this species can exhibit stem elongation, facilitating the isolation of the nodal segments to be applied in its micropropagation. The rosette morphology is reestablished when this species is maintained under low temperature, thus allowing the maintenance of a germplasm collection (slow growth storage). Gibberellins (GA) are usually applied to stimulate stem elongation in micropropagated plants. Thus, our aim here was to verify the influence of temperature over the stem elongation of A. strobilacea when GA₃ is applied to the medium, thus estimating the use of this phytoregulator in slow growth cultures at low temperatures. Physiological and anatomical studies were performed on plants obtained from nodal segments maintained at 10, 15, 20, and 25 °C. Regardless of the applied treatment, no segments developed at 10 °C. Stem elongation occurred at 25 and 30 °C, and was not seen for plants grown under 15 and 20 °C. The application of 50 µM of GA₃ restored stem elongation in plants at 20 but not at 15 °C. The influence of gibberellins on stem elongation of this tropical bromeliad depends on the cultivation temperature, in which low temperature preponderates over the stem elongation effects of GA₃. In addition, the optimum temperature for the slow growth of this species depends on the starting temperature of the explant used in the micropropagation.     

The control by gibberellic acid of stem elongation and flowering in biennial plants of Centaurium minus Moench

Summary Application of a small amount of gibberellic acid (GA₃) to unvernalized rosettes of a biennial strain of Centaurium minus Moench brings about immediate stem elongation under both long days (LD) and short days (SD), but the rate of stem elongation falls after the cessation of treatment. Under LD, but not SD, a second period of rapid and prolonged stem elongation may subsequently take place, associated with flowering. Extended GA₃ treatment under SD leads to the formation of a long stem but not to flowering; after the treatment the plants revert to vegetative aerial rosettes unless transferred to LD prior to the cessation of stem elongation; after such a transfer, rapid stem elongation and flowering may be initiated. If flower primordia are initiated under LD, stem elongation and formation of flower primordia continue after transfer to SD, though flowers do not develop fully. It is suggested that under LD but not SD applied GA₃ may bring about the production of endogenous gibberellin, and that this synthesis of endogenous gibberellin occurs in the flower primordia.     

Der Einfluß von Wachstumsregulatoren auf Protein- und Enzym-Spektren kultivierter Explantate aus Rüben von Cichorium intybus L.

Summary On a simple nutrient medium in explants from roots of Cichorium intybus form shoots visible after about 14 days. Gibberellic acid (GA₃) does not influence the spontaneous development of the chicory explants. GA₃ in combination with kinetin inhibits shoot formation whereas kinetin alone promotes the process. On the other hand high concentrations of IAA inhibit the regeneration of shoots.The soluble proteins of chicory cultures treated with growth regulators were examined by disc-electrophoresis. It was shown that the proteins detected by staining with Amido black, phosphatases, esterases and glutamate dehydrogenase (GDH) present in the original root tissue remained constant under the different culture conditions during a period of 12 days. The quantitative changes of some of the proteins, phosphatases and esterases observed during the culture period were identical for all the different cultures in spite of the different morphogenetic behaviour. Only the activities of GDH and peroxidase changed after treatment with different growth regulators; however, in these cases, there was also no direct connection with the morphogenetic responses of the cultures.The specific activity of the GDH-band was promoted by IAA and at the same time the formation of peroxidases was inhibited. Kinetin delayed the formation of peroxidases during the first days of the culture period but promoted it later on. There was a repression by IAA of a specific kationic peroxidase. In the tissues treated with GA₃ the activity of peroxidases was always higher than in the control tissue. This effect of GA₃ can be partly explained by the fact that GA₃ inhibits the release of peroxidases of the explants into the nutrient medium.     

Growth retarding effects of the herbicide difenzoquat

The growth-regulating properties of the herbicide Difenzoquat (1,2-dimethyl-3,5-diphenyl-1-H-pyrazolium methyl sulfate) were investigated in seedlings and cell suspension cultures of sunflower (Helianthus annuus L.). Application of 10 g or more Difenzoquat to the apex of seedlings resulted in a transient inhibition of internode elongation. Application of GA₃ to treated seedlings resulted in enhanced internode elongation but did not reverse the degree of growth inhibition elicited by Difenzoquat. Endogenous gibberellin levels were estimated by bioassay and were qualitatively and quantitatively similar in extracts from control and treated seedlings. Treatment of suspension cultures of sunflower cells with 1 M or more Difenzoquat resulted in an inhibition of cell division (dry-matter accumulation). Neither GA₃ nor a mixture of sterols (cholesterol, -sitosterol, and stigmasterol) alone or in combination was able to overcome this inhibition of cell division. It was concluded that the growth-retarding activity of Difenzoquat was the result of its action at the cellular level and was not mediated by inhibition of gibberellin biosynthesis.