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PubMed1975

Endogenous Gibberellins of Pine Pollen: II. Changes during Germination of Pinus attenuata, P. coulteri, and P. ponderosa Pollen.

The endogenous gibberellins (GAs) of pollen of Pinus attenuata, P. coulteri, and P. ponderosa were bioassayed at hour 0, 3, 15, 24, 48 and 72 of germination. Dormant pollen showed relatively high GA activity throughout the elution spectrum (i.e. ranging from relatively nonpolar to highly polar). The maximum GA activity was obtained at hour 15 in more polar regions and especially in the zone corresponding to GA(3) (for P. attenuata estimated as 250 micrograms of GA(3)/kilogram pollen). It is probable that the "nonpolar" GAs present in high quantities in dormant pollen and in early stages of germination were converted to "more polar" GAs as germination progressed. The amount of all GAs decreased after hour 15 of germination and by hour 72 no GAs could be detected. Among the species tested P. attenuata showed the highest over-all GA activity.

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PubMed1976

Effects of Light, Abscisic Acid, and N-Benzyladenine on the Metabolism of [H]Gibberellin A(4) in Seeds and Seedlings of Lettuce, cv. Grand Rapids.

Gibberellin A(4) (GA(4)) can substitute for light in the germination of Grand Rapids lettuce seeds. Seeds imbibed in [(3)H]GA(4) do not convert this to other GAs prior to, or immediately following, visible germination: thus GA(4) alone can promote radicle expansion. Abscisic acid inhibited [(3)H]GA(4)-induced germination, but did not significantly affect [(3)H]GA(4) uptake or metabolism during germination. (6)N-benzyladenine overcame the inhibitory effect of abscisic acid and increased [(3)H]GA(4) uptake, although radicle emergence was delayed somewhat.During hypocotyl extension there was a large conversion of [(3)H]GA(4) to [(3)H]GA(1) in light or darkness, the major conversion site being the growing root. Hypocotyls of dark-grown seedlings contained more [(3)H]GA(1) than those of light-grown seedlings. The apparent inability of exogenous GA(1) to promote greater hypocotyl extension than GA(4) is related to its poorer uptake. Abacisic acid markedly inhibited hypocotyl expansion, root growth, and the conversion of [(3)H]GA(4) to [(3)H]GA(1).

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PubMed1976

Analysis of native gibberellins in the internode, nodes, leaves, and inflorescence of developing Avena plants.

The native gibberellins (GAs) of various organs of the Avena plant were analyzed by bioassay and gas chromatography-mass spectrometry (GC-MS) after silicic acid partition column chromatography. The major GA of the inflorescence was identified as GA(3) by GC-MS, and this GA also forms the major component of the nodes, p-1 internode, and roots as determined by GLC or chromatography/bioassay. The inflorescence and nodes are the major sources of native GAs, the last two leaves, internode, and roots having significantly lower amounts of GA-like substances. In the internode, less polar GAs predominated at the lag stage of development, whereas by the log and plateau stages, the more polar GAs increased significantly.Since less polar GAs are early in oxidative interconversion sequences, this finding indicates sequential conversion to more polar and probably more active GAs, during log phase growth of the p-1 internode.

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PubMed1976

Metabolism of [H]Gibberellin A(20) in Light- and Dark-grown Tobacco Callus Cultures.

The growth of tobacco callus in culture (previously shown to contain gibberellin [GA]-like substances), and its ability to metabolize [(3)H]-GA(20) were examined. Growth rates, in the presence and absence of exogenously applied GA, were examined in light and dark conditions. Dark-grown callus grew at a much faster rate than light-grown and [(3)H]GA(20) was metabolized much more rapidly in darkness than in light.[(3)H]GA(1) was identified by combined gas-liquid chromatography/mass spectrometry as a major product of [(3)H]GA(20), and was found to be a more potent promoter of tobacco callus growth than GA(20).

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PubMed1977

Roles of Extensibility and Turgor in Gibberellin- and Dark-stimulated Growth.

The elongation response elicited by incubating excised hypocotyl sections of lettuce (Lactuca sativa L.) in light in gibberellin (GA) can be enhanced by the addition of Cl(-), Br(-), and NO(3) (-) salts of K(+) and Na(+). Sections incubated in light in the absence of GA do not elongate in response to the addition of salts. In contrast, excised hypocotyls incubated in darkness elongate equally in both GA and water, and their elongation can also be enhanced by KCl treatment. Growth stimulation by the salts of K(+) and Na(+) occurs optimally at 10 mm and the magnitude of the response is proportional to the duration of salt treatment. Although the growth of sections incubated in light in the absence of GA is not enhanced by various salts of K(+) and Na(+), the concentration of these cations exceeds that in GA-treated sections. In dark-grown tissue, uptake of K(+) also occurs in both GA- and H(2)O-treated sections incubated in 10 mm KCl. Since increased osmotic potential resulting from cation uptake does not correlate with growth stimulation resulting from salt treatments, we conclude that increased cell turgor is not the principal driving force for growth in hypocotyl sections. Changes in the extensibility of GA-treated, light-grown tissue and dark-grown tissue incubated with and without GA correlate with the increased growth rate of these sections. Incubation of sections in KCl results only in changes in water potential of sections without having a significant effect on extensibility. When changes in water potential are accompanied by increased extensibility, however, a marked increase in growth rate is observed.

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PubMed1977

Differential counteraction of ethylene effects by gibberellin a(3) and n(6)-benzyladenine in senescing citrus peel.

Treatment of mature citrus fruit (Citrus sinensis) with ethylene induced rapid chlorophyll destruction, a rise in respiration, a release of free amino acids, an accumulation of reducing sugars, and an appearance of phenylalanine ammonia lyase activity. Gibberellin A(3) (GA(3)) and N(6)-benzyladenine (BA) opposed the effects of ethylene on chlorophyll, amino acids, and to a lesser extent, reducing sugar levels. The ethylene-induced respiratory rise was only slightly modified by GA(3) and BA. Phenylalanine ammonia lyase activity was not affected by GA(3).The antagonism between ethylene and the senescence-delaying regulators GA(3) and BA seems to operate mainly within the chloroplast, but might not be confined to this compartment. The accumulation of reducing sugars exhibits the antagonism although it is not apparently related to the chloroplast.

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PubMed1977

Effect of ethylene on the endogenous cytokinin and gibberellin levels in tuberizing potatoes.

High concentrations of 2-chloroethylphosphonic acid inhibited tuberization on aged potato tubers (Solanum tuberosum) that had been predisposed to the little tuber disorder. As a result of this treatment sprouts developed which contained relatively high levels of endogenous gibberellins and which elongated normally. The endogenous cytokinin levels in the different treatments did not change appreciably. It is suggested that tuberization is prevented by ethylene either as a direct inhibition of cell division or that it prevents the endogenous cytokinins from functioning. Irrespective of the mode of action of ethylene, cell division apparently is the primary process affected, the result being that storage tissue required for the accumulation of starch is not formed.

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PubMed1978

Effect of gibberellin a(3) on the endoplasmic reticulum and on the formation of glyoxysomes in the endosperm of germinating castor bean.

Seedlings of castor bean (Ricinus communis cv. Hale) were exposed to a range of concentrations of gibberellin A(3) (GA(3)). Treatments for 20 hours with GA(3) concentrations of 0.5 muM or higher resulted in increased levels of NADH-cytochrome c reductase, phosphorylcholine glyceride transferase, and malate synthase in endoplasmic reticulum (ER) isolated from endosperm on linear sucrose gradients. GA(3) treatment also resulted in increased RNA associated with ER. Malate synthase and catalase in crude homogenates were enhanced by 1 to 100 muM GA(3) concentrations. Isocitrate lyase, citrate synthase, malate synthase, catalase, and glycolate oxidase in isolated glyoxysomes were enhanced by 60, 20, 18, 40, and 28%, respectively, over controls. Treatment with abscisic acid led to decreased levels of glyoxysomal enzymes and reduced glyoxysomal protein. The effect of GA(3) and abscisic acid on the specific activities of glyoxysomes of different densities suggests that GA(3) influences enzyme levels and glyoxysome assembly.

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PubMed1979

Temperature Interactions with Growth Regulators and Endogenous Gibberellin-like Activity during Seedstalk Elongation in Carrots.

Stecklings (roots) of three cultivars of carrots (Daucus carota L.) were vernalized 10 weeks at 5 C and subsequently grown at each of three greenhouse night/day temperature regimes: high (27/32 C), medium (21/27 C), and low (15/21 C). Floral differentiation occurred first in the easy bolting cv. Scarlet Nantes, intermediate in cv. Danvers 126, and last in cv. Royal Chantenay. Stem elongation arising from the subapical meristematic region always preceded floral differentiation. Extractable gibberellin-like activity in carrot stem apices increased from harvest during the 10-week vernalization period, then remained constant even though floral differentiation and stem elongation occurred during an additional 20-week cold storage period. Low temperature had both an inductive and a direct effect on reproductive development depending on length of low temperature exposure.After 10 weeks vernalization at 5 C, high greenhouse temperature severely reduced ultimate seedstalk height and the endogenous gibberellinlike activity decreased rapidly during the first 3 weeks in the greenhouse. At the low greenhouse temperature, activity remained fairly constant during the 10-week sampling period. Changes in endogenous gibberellinlike activity were related with stem elongation, but not with floral initiation. Exogenous gibberellic acid (GA(3)) applied following vernalization prevented the inhibitory effect of high greenhouse temperature on seedstalk elongation and resulted in seedstalk heights comparable to untreated controls grown at the low greenhouse temperature. Exogenous applications of succinic acid-2,2-dimethylhydrazide and chlormequat reduced seedstalk height of carrot plants grown at the medium and low greenhouse temperatures to that of untreated controls grown at high temperature. Exogenous growth regulators and greenhouse temperature affected seedstalk elongation, but did not affect the number of plants that flowered.

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PubMed1979

Metabolism of Tritiated Gibberellin A(20) in Immature Seeds of Dwarf Pea, cv. Meteor.

Tritium-labeled gibberellin A(20) ([(3)H]GA(20)) was applied via the pedicel to immature pods and seeds of dwarf peas and three harvests were made at days 5, 10, and 23 (mature) after application. Of the five metabolites of [(3)H]GA(20), the three in highest yield were GA(29), an alpha,beta-unsaturated ketone, and a compound (B), whose structure was only tentatively assigned. The metabolic sequence GA(20) --> GA(29) --> compound B --> the ketone was indicated. The amount of [(3)H]GA(29) in both seeds and pods was highest at day 5 and declined to its lowest level at maturity. The amount of the [(3)H]ketone in the seed increased with time to its highest level at maturity. It is suggested that compound B and the ketone represent the major pathway of catabolism of GA(29), a 2beta-hydroxylated GA of low biological activity, and that the ketone is not metabolized, or only slowly metabolized, during seed maturation.

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PubMed1980

Gibberellin-sensitive Suspension Cultures.

Suspension cultures were incubated in the presence and absence of gibberellic acid (GA(3)) in an attempt to define a new experimental system for study of the molecular action of gibberellins upon growth. Unlike many suspension cultures, an auxin-independent green clone from spinach (Spinacia oleracea L.) and an auxin-dependent line of "Paul's Scarlet" rose (Rosa sp.) were promoted in expansion growth by GA(3) at 10(-11) to 10(-6) molar. In Rosa the cells also elongated upon GA(3) treatment whereas in Spinacia they remained isodiametric.Attempts were made to intensify the response. The effect of GA(3) in Spinacia was stronger when gas exchange between the culture and the laboratory air was facilitated. The response of Rosa was dependent on the presence of a cytokinin, although this culture did not require exogenous cytokinin for serial subculture. GA(3) antagonized the growth retardant AMO-1618 in Rosa but not in Spinacia. In general, conditions that enhanced growth also rendered GA(3) a more effective promoter; conversely, GA(3) tended to become inhibitory under conditions that permitted only slow growth.

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PubMed1980

Identification of six endogenous gibberellins in spinach shoots.

Analysis of highly purified extracts from spinach shoots by combined gas chromatography-mass spectrometry has demonstrated the presence of six 13-C-hydroxylated gibberellins (GAs): GA(53), GA(44), GA(19), GA(17), GA(20), and GA(29). The major GAs were GA(17), GA(19), and GA(20), whereas the other three GAs occurred in trace amounts. Structural considerations suggest that the six GAs identified in spinach are related in the following metabolic sequence: GA(53) --> GA(44) --> GA(19) --> GA(17) --> GA(20) --> GA(29).

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PubMed1980

Uptake and subcellular compartmentation of gibberellin a(1) applied to leaves of barley and cowpea.

The uptake and subcellular accumulation of gibberellin A(1) (GA(1)) by leaves and protoplasts of barley (cv. Numar) and cowpea (cv. Blackeye pea No. 5) were investigated.Uptake of GA(1) by cowpea leaves is optimal at pH 5.8 and occurs by a saturable, probably carrier-mediated process having a half-maximal velocity at 10 to 20 micromolar. Uptake by both barley and cowpea leaves is inhibited by low temperature (+4 C) and the metabolic inhibitors 2,4-dinitrophenol and azide and is stimulated by ATP. Mesophyll protoplasts isolated from leaves fed radioactive GA(1) retain 20 to 80% of the radioactivity incorporated by excised leaves.The subcellular localization of the [(3)H]GA was determined by lysing protoplasts and separating subcellular organelles by density gradient centrifugation. Less than 5% of the incorporated [(3)H]GA was found associated with chloroplasts, mitochondria, nuclei, or other organelles or membranes with densities in sucrose gradients greater than 1.15 grams per cubic centimeter. Fifty to 100% of the [(3)H]GA was found in vacuoles. Isolated vacuoles were judged to be free of contamination by cytoplasm using phosphoenolpyruvate carboxylase as a marker enzyme. Osmotic breakage of vacuoles or protoplasts released > 95% of the [(3)H]GA, suggesting that GA is associated with the vacuolar sap rather than with the tonoplast membrane.

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PubMed1980

Comparison of the levels of six endogenous gibberellins in roots and shoots of spinach in relation to photoperiod.

This communication describes the distribution of gibberellins (GAs) in roots and shoots of spinach in relation to photoperiod. From previous work (Metzger, Zeevaart 1980 Plant Physiol 65: 623-626) shoots were known to contain GA(53), GA(44), GA(19), GA(17), GA(20), and GA(29). We now show by combined gas chromatography-mass spectrometry that roots contain GA(44), GA(19), and GA(29). Trace amounts of GA(53) were detected by combined gas chromatography-selected ion current monitoring. Neither GA(17) nor GA(20) were detected in root extracts. Analysis by the d-5 corn bioassay also showed no effect of photoperiodic treatment on the levels of GA-like substances in root extracts. Both phloem and xylem exudates had patterns of GA-like activity similar to those found in shoots and roots, respectively. Moreover, foliar application of [(3)H]GA(20) resulted in the transport of label from the shoot to the roots. Over half of the label in the roots represented unmetabolized [(3)H]GA(20), indicating that part of the GA(20) in the phloem is transported to the roots. Consequently, if GA(20) is made in, or transported to the roots, it is rapidly metabolized in that organ. This is a clear indication that regulation of GA metabolism is greatly different in roots and shoots.

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PubMed1980

Changes of Endogenous Gibberellin-like Substances with Sex Reversal of the Apical Inflorescence of Corn.

In developing apical meristems of corn, the level of acidic, ethyl acetate-soluble gibberellin (GA)-like substances increased to a maximum of 108 micrograms GA(3)-equivalents per kilogram dry weight of tissue at inflorescence initiation, and then fell rapidly. At anthesis, only a trace (0.2 microgram per kilogram) of GA-like activity remained in the apical (male) inflorescences, whereas moderate activity (32 micrograms per kilogram), mostly of a nonpolar nature, was present in lateral, female, inflorescences.A sex reversal of the apical inflorescence, from male to female, was elicited by reducing the ambient light intensity. Higher levels of GA-like substances, particularly those eluting from a SiO(2) partition column in the nonpolar region, were observed at all harvests in the reverting meristems; levels increased to 180 micrograms per kilogram at inflorescence initiation, then dropped to 122 micrograms per kilogram in the apical (female), reverted meristems. This increase in endogenous GA-like activity with reversion to the female inflorescence is consistent with observations that (a) reversion can be obtained with exogenous application of GA(3) and (b) maleness is enhanced in GA-deficient mutants of maize. Endogenous GAs may thus play a key role in the control of sexuality of corn.

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PubMed1980

Effect of Photoperiod on the Levels of Endogenous Gibberellins in Spinach as Measured by Combined Gas Chromatography-selected Ion Current Monitoring.

The changes in the levels of five endogenous gibberellins (GAs) in spinach in relation to photoperiodic treatment have been examined by combined gas chromatography-selected ion current monitoring. Long-day treatment caused a 5-fold decline in the level of GA(19) while the levels of GA(20) and GA(29) increased dramatically during the same period. In absolute terms, the level of GA(20) increased from 0.8 microgram per 100 grams dry weight in short days to 5.5 micrograms per 100 grams dry weight after 14 long days. The levels of GA(17) and GA(44) did not change significantly with long-day treatment. These results are consistent with the hypothesis that GA(19) is converted to GA(20) and that this conversion is under photoperiodic control. Since stem growth in spinach is correlated with an increase in the level of GA(20), one major aspect of photoperiodic control of stem growth might be the availability of GA(20) through regulation of the conversion of GA(19) to GA(20).

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PubMed1980

In vivo Binding of Gibberellin A(1) in Dwarf Pea Epicotyls.

Binding of [(3)H]gibberellin A(1) (GA(1)) to extracts of dwarf pea epicotyls was investigated using sliced pea epicotyls (0.5-1.0 millimeter thick) that had been incubated in a solution containing [(3)H]GA(1) at 0 C for 3 days. Gel filtration of a 100,000g supernatant indicated binding to a high (HMW) and an intermediate molecular weight (IMW) fraction with estimated molecular weights of 6 x 10(5) daltons and 4 to 7 x 10(4) daltons, respectively. The bound (3)H-activity was [(3)H]GA(1) and not a metabolite as deduced by thin layer chromatography. The bound label did not sediment during centrifugation at 100,000g for 2 hours; also, binding was not disrupted after treatment of a combined HMW and IMW fraction with DNase, RNase, or phospholipase A or C, but it was disrupted by protease or heat treatment. These facts suggest that binding of [(3)H]GA(1) was occurring to a soluble protein(s). [(3)H]GA(1) bound to a combined HMW and IMW fraction was not susceptible to changes in pH, nor could it be exchanged with a variety of GAs tested under in vitro conditions. Under in vivo equilibrium conditions, biologically active GAs, such as GA(1), GA(3), GA(4), GA(5), GA(7), and keto GA(1), could reduce the level of [(3)H]GA(1) binding, whereas inactive GAs, such as iodo GA(1) methyl ester, GA(8), GA(13), GA(26), and non-GAs, such as (+/-)abscisic acid, had no effect. By varying the concentration of [(3)H]GA(1) in the incubation medium, the specific binding of [(3)H]GA(1) appeared to be due to two classes of binding sites having estimated K(d) of 6 x 10(-8) molar and 1.4 x 10(-6) molar. The concentrations of the two sites were estimated to be 0.45 picomole per gram and 4.04 picomoles per gram on a fresh weight and 0.1 picomole per milligram and 0.9 picomole per milligram on a soluble protein basis, respectively.

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PubMed1981

Changes in Endogenous Gibberellins and the Metabolism of [H]GA(4) after Geostimulation in Shoots of the Oat Plant (Avena sativa).

The recovery from "lodging," or bending over, by shoots of 42-day-old Avena sativa plants is controlled primarily by a negatively geotropic differential growth of the lower halves of the p-1 node-pulvinus and the base of the p-1 internode, relative to the upper halves. Although geostimulation causes a significant reduction in p-1 internode length, dry matter accumulation in the p-1 node-pulvinus is increased, apparently at the expense of the sheath. Recovery to an angle of 30 degrees is associated with changes in endogenous gibberellin-like substances (GAs), and in differential metabolism of applied [(3)H]GA(4) (1.4 Curie per millimole). Although geostimulation depressed total GAs (relative to upright plant parts) to 0.40 and 0.13 for node-pulvini and sheaths, respectively, it increased them 2-fold for internodes. Within the plant part geostimulation increased GAs (relative to upper halves) 29- and 7-fold in lower halves of node-pulvini and internodes, respectively, but reduced GAs to 0.3 in lower halves of sheaths. At age 42 days a GA(4/7)-like (nonpolar) substance predominates, with lesser amounts of a GA(3)-like (polar) substance. Native GAs of Avena include GA(3), GA(4), and GA(7). Geostimulation enhanced the ratio of nonpolar to polar GAs for both halves of internodes, but tended to depress it for sheaths and nodepulvini.The disposition and metabolism of applied [(3)H]GA(4) confirmed the above trends for endogenous GAs regarding localization (e.g. up to 2-fold increases in [(3)H]GA(4) and acidic (3)H-metabolites in the lower halves, relative to upper halves). Also, metabolism into highly water-soluble (3)H-metabolites (biologically inactive conjugates?) was greater (up to 1.8-fold) in upper than in lower halves. The end result of such metabolic trends would be to reduce acidic (biologically active?) GAs in the upper half, while retaining them for a longer time in the lower half.Geotropically stimulated Avena shoots thus increase, within 24 hours, the levels of acidic GAs in the lower halves of the p-1 node-pulvinus and p-1 internode, the two plant parts responsible for the geostimulated growth.

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