Dwarf mutants ofBrassica: Responses to applied gibberellins and gibberellin content

Eight rapid-cyclingBrassica genotypes differing in height were treated with gibberellins (GAs) by syringe application to the shoot tip. The height of two genotypes ofBrassica napus, Bn5-2 and Bn5-8, andB. rapa mutants,dwarf 1 (dwf1) anddwarf 2 (dwf2), was unaffected by exogenous GA₃ at dosages up to 0.1 μg/plant, a level which increased shoot elongation of normal genotypes. Thus, these dwarf mutants are “GA-insensitive.” In contrast to theB. napus dwarfs, twoB. rapa mutants,rosette (ros), anddormant (dor), elongated following GA₃ application. The dwarfros was most sensitive, responding to applications as low as 1 ng GA₃/plant. Furthermore,ros also responded to GA₁ and some of its precursors with decreasing efficacy: GA₃>ent-kaurenoic acid ≥GA₁>GA₂₀≥GA₁₉=GA₄₄≥GA₅₃. Endogenous GAs were measured by gas chromatography-selected ion monitoring using [²H₂]GA internal standards for calibration, from shoots of the GA-insensitive genotypes Bn5-2, Bn5-8 which contained theB. napus mutantdwarf 1, and from a normal genotype Bn5-1. Concentrations of GA₁ and GA₂₀ averaged 3.2- and 4.6-fold higher, respectively, and GA₁₉ levels also tended to be higher in the dwarfs than in the normal genotype.     

The endogenous gibberellins of dwarf mutants of lettuce

The gibberellin (GA) content of E-1, a tall genotype of early flowering lettuce (Lactuca sativa L.), and of three selected GA-responsive dwarfs, dwf1, dwf2, and dwf2¹, has been determined using ¹³C-labeled internal standards and gas chromatographymass spectrometry (GC-MS). In the shoots of the E-1 parent, GA₁, 3-epi-GA₁, GA₃, GA₅, GA₈, GA₁₉, GA₂₀, GA₂₉, and GA₅₃ were identified by full scan GC-MS and Kovats retention indices. Purification by immunoaffinity chromatography selective for 13-hydroxy GAs, was necessary for GA identification. Relative to the parent E-1, the concentrations of GA₁, GA₈, GA₂₀, and GA₂₉ in the shoots of dwf2 plants were reduced to about 10% and in shoots of dwf2¹ plants to less than 50%. In dwf1 the levels of GA₁, GA₈, and GA₂₉ were also reduced to less than 50% of the parent E-1, but the level of GA₂₀ was fivefold higher than in E-1. Plant height was correlated with the endogenous levels of GA₁ and GA₈.     

Dwarf mutants ofBrassica: Responses to applied gibberellins and gibberellin content

Eight rapid-cyclingBrassica genotypes differing in height were treated with gibberellins (GAs) by syringe application to the shoot tip. The height of two genotypes ofBrassica napus, Bn5-2 and Bn5-8, andB. rapa mutants,dwarf 1 (dwf1) anddwarf 2 (dwf2), was unaffected by exogenous GA₃ at dosages up to 0.1 g/plant, a level which increased shoot elongation of normal genotypes. Thus, these dwarf mutants are GA-insensitive. In contrast to theB. napus dwarfs, twoB. rapa mutants,rosette (ros), anddormant (dor), elongated following GA₃ application. The dwarfros was most sensitive, responding to applications as low as 1 ng GA₃/plant. Furthermore,ros also responded to GA₁ and some of its precursors with decreasing efficacy: GA₃>ent-kaurenoic acid GA₁>GA₂₀GA₁₉=GA₄₄GA₅₃. Endogenous GAs were measured by gas chromatography-selected ion monitoring using [²H₂]GA internal standards for calibration, from shoots of the GA-insensitive genotypes Bn5-2, Bn5-8 which contained theB. napus mutantdwarf 1, and from a normal genotype Bn5-1. Concentrations of GA₁ and GA₂₀ averaged 3.2- and 4.6-fold higher, respectively, and GA₁₉ levels also tended to be higher in the dwarfs than in the normal genotype.     

Comparison of endogenous gibberellins and of the fate of applied radioactive gibberellin a(1) in a normal and a dwarf strain of Japanese morning glory

The effect of application of GA₃ on hypocotyl growth, the endogenous GAs, and the metabolism of applied ³H-GA₁ were investigated in relation to dwarfism and light-mediated growth inhibition in the normal (tall) strain Violet and the dwarf strain Kidachi of Japanese morning glory (Pharbitis nil). GA₃ applied in a wide concentration range (10⁻⁹ to 10⁻³m) to 4-day-old seedlings caused great extension of the hypocotyls in light-grown plants of both the normal and the dwarf strain. However, the dwarf strain did not attain the same length as the normal one at any given GA₃ concentration, even when saturation was reached. Dark-grown plants of the dwarf strain responded to GA₃, although relatively much less than light-grown ones; dark-grown plants of the normal strain showed no GA₃ response at all.     

Response of seedlings of a dwarf and a normal strain of watermelon to gibberellins

Hypocotyl and root elongation in a dwarf and a normal strain of watermelon (Citrullus lanatus [Thunb.] Matsu.) in the absence or presence of different gibberellins was investigated in seedlings grown under gold fluorescent light or in darkness. The normal strain, “Sugar Baby,” responded only slightly to the gibberellic acids employed. At appropriate concentrations all of the gibberellic acids were capable of normalizing growth in the monorecessive dwarf strain, WB-2, in darkness or in light. Gibberellins A₄₊₇ and A₇ were effective in stimulating hypocotyl elongation at concentrations 10 to 15 times lower than that needed for a response to GA₁ or GA₃. Dark-grown dwarfs responded to about a 3-fold lower concentration of GA₄₊₇ than those grown in light.     

Growth and gibberellin-A1 metabolism in normal and gibberellin-insensitive (Rht3) wheat (Triticum aestivum L.) seedlings

Growth parameters were determined for tall (rht3) and dwarf (Rht3) seedlings of wheat (Triticum aestivum L.). Plant statures and leaf length were reduced by 50% in dwarfs but root and shoot dry weights were less affected. Leaves of dwarf seedlings had shorter epidermal cells and the numbers of cells per rank in talls and dwarfs matched the observed relationships in overall length. Talls grew at twice the rate of dwarfs (2.3 compared with 1.2 mm h^-1). [³H]Gibberellin A₁ ([³H]GA₁) was fed to seedlings via the third leaf and metabolism was followed over 12 h. Immature leaves of tall seedlings transferred radioactivity rapidly to compounds co-chromatographing with [³H]gibberellin A₈ ([³H]GA₈) and a conjugate of [³H]GA₈, whereas leaves of dwarf seedlings metabolised [³H]GA₁ more slowly. Roots of both genotypes produced [³H]GA₈-like material at similar rates. Isotopic dilution studies indicated a reduced 2-hydroxylation capacity in dwarfs, but parallel estimates of the endogenous GA pool size, obtained by radioimmunoassay, indicated a 12–15 times higher level of GA in the dwarf immature leaves. Dwarfing by the Rht3 gene does not appear to operate through enhanced, or abnormal metabolism of active gibberellins and the act of GA metabolism does not bear an obligate relationship to the growth response.Abbreviations GA_n gibberellin A_n - HPLC high-performance liquid chromatography     

Narrow-band light regulation of ethylene and gibberellin levels in hydroponically-grown <Emphasis Type="Italic">Helianthus annuus</Emphasis> hypocotyls and roots

Both hypocotyl and root growth of sunflower (Helianthus annuus) were examined in response to a range of narrow-band width light treatments. Changes in two growth-regulating hormones, ethylene and gibberellins (GAs) were followed in an attempt to better understand the interaction of light and hormonal signaling in the growth of these two important plant organs. Hydroponically-grown 6-day-old sunflower seedlings had significantly elongated hypocotyls and primary roots when grown under far-red (FR) light produced by light emitting diodes (LEDs), compared to narrow-band red (R) and blue (B) light. However, hypocotyl and primary root lengths of seedlings given FR light were still shorter than was seen for dark-grown seedlings. Light treatment in general (compared to dark) increased lateral root formation and FR light induced massive lateral root formation, relative to treatment with R or B light. Levels of ethylene evolution (roots and hypocotyls) and concentrations of endogenous GAs (hypocotyls) were assessed from both 6-day-old sunflower plants either grown in the dark, or treated with FR, R or B light. Both R and B light had similar effects on hypocotyl and root growth as well as on ethylene and on hypocotyl GA levels. Dark treatment resulted in the highest ethylene levels, whereas FR treatment significantly reduced ethylene evolution for both hypocotyls and roots. R- and B-light treatments elevated ethylene evolution relative to FR light. Endogenous GA₅₃ and GA₁₉ levels in hypocotyls were significantly higher and GA₄₄, GA₂₀ and GA₁ levels significantly lower, for dark and FR light treatments compared to R and B light-treatments. The patterns seen for changes in GA concentrations indicate FR-, R- and B-light-mediated effects [differences] in the metabolism of the early C₂₀ GAs, GA₅₃ → GA₄₄ → GA_19. Surprisingly, GA₂₀, GA₁ and GA₈ levels in hypocotyls were very much reduced by treatment of the plants with FR light, relative to B and R-light treatments, e.g. the increased hypocotyl elongation induced by FR light was correlated with reduced levels of all three of the downstream C₁₉ GAs. The best explanation, albeit speculative, is that a more rapid metabolism, i.e. GA₂₀ → GA₁ → GA₈ → GA₈ conjugates occurs under FR light. Although this study provided no evidence that elevated ethylene evolution by roots or hypocotyls of sunflower is controlling growth via endogenous GA biosynthesis, there are differences between soil-grown and hydroponically-grown sunflower seedlings with regard to trends seen for hypocotyl GA concentrations and both root and hypocotyl ethylene evolution in response to narrow band width R and FR light signaling.     

Exogenous GA3 Application Can Compensate the Morphogenetic Effects of the GA-Responsive Dwarfing Gene Rht12 in Bread Wheat

The most common dwarfing genes in wheat, Rht-B1b and Rht-D1b, classified as gibberellin-insensitive (GAI) dwarfing genes due to their reduced response to exogenous GA, have been verified as encoding negative regulators of gibberellin signaling. In contrast, the response of gibberellin-responsive (GAR) dwarfing genes, such as Rht12, to exogenous GA is still unclear and the role of them, if any, in GA biosynthesis or signaling is unknown. The responses of Rht12 to exogenous GA₃ were investigated on seedling vigour, spike phenological development, plant height and other agronomic traits, using F_2∶3 and F_3∶4 lines derived from a cross between Ningchun45 and Karcagi-12 in three experiments. The application of exogenous GA₃ significantly increased coleoptile length and seedling leaf 1 length and area. While there was no significant difference between the dwarf and the tall lines at the seedling stage in the responsiveness to GA₃, plant height was significantly increased, by 41 cm (53%) averaged across the three experiments, in the GA₃-treated Rht12 dwarf lines. Plant height of the tall lines was not affected significantly by GA₃ treatment (<10 cm increased). Plant biomass and seed size of the GA₃-treated dwarf lines was significantly increased compared with untreated dwarf plants while there was no such difference in the tall lines. GA₃-treated Rht12 dwarf plants with the dominant Vrn-B1 developed faster than untreated plants and reached double ridge stage 57 days, 11 days and 50 days earlier and finally flowered earlier by almost 7 days while the GA₃-treated tall lines flowering only 1–2 days earlier than the untreated tall lines. Thus, it is clear that exogenous GA₃ can break the masking effect of Rht12 on Vrn-B1 and also restore other characters of Rht12 to normal. It suggested that Rht12 mutants may be deficient in GA biosynthesis rather than in GA signal transduction like the GA-insensitive dwarfs.     

Comparison of growth and gibberellin concentrations in shoots from orchard-grown standard and thermosensitive dwarf apple trees

Apple (Malus domestica Borkh.) trees were propagated by budding from selected fully grown hybrids that ranged in height from 1.5 to 8 m. The growth and development of the selected budded trees after 7 years in the orchard was similar to that of the parent trees. Additional grafting studies showed that the dwarfism was not associated with the roots. Differences in photosynthetic activity and associated processes were not related to the size difference between tissue culture-propagated orchard-grown standard cv. Golden Delicious and dwarf hybrid trees. Applications of GA₃ did not stimulate elongation of shoots of dwarf trees. Shoots of both standard and dwarf trees started to develop in mid-April when they contained nearly the same amounts of GA₁, GA₃ and GA₈, but standard shoots contained higher concentrations of GA₁₉, GA₂₀ and GA₂₉. On 2 June standard shoots were almost three times the length of dwarf shoots, but the number of leaves and area per leaf were nearly the same. The relative amounts of GAs on 12 May and 2 June for both plant types were similar to those on 20 April, except that GA₁₉, GA₂₀, GA₁ and GA₂₉ levels had declined. Gibberellin levels in standard shoots declined further between 2 and 22 June, after which there was no further shoot elongation or production of new leaves. Between 2 June and the end of the growing season, when summer temperatures were high, dwarf shoots continued to elongate slowly and to develop new leaves, which expanded little. During this time, the GA₁₉ content of dwarf shoots nearly doubled, whereas the amounts of GA₂₀, GA₁, GA₂₉ and GA₈ declined. By the end of the season, standard shoots were 40 cm in length with 20 leaves and dwarf shoots were 28 cm in length, but with 36 leaves. High summer temperatures appear to induce loss of GA-responsiveness in orchard-grown dwarf trees and to cause a reduced rate of conversion of GA₁₉ to GA₂₀ in these genotypes.     

Interaction of Brassinosteroids with Light Quality and Plant Hormones in Regulating Shoot Growth of Young Sunflower and <Emphasis Type="Italic">Arabidopsis</Emphasis> Seedlings

Sunflower hypocotyls elongate as light quality changes from the normal red to far-red (R/FR) ratio of sunlight to a lower R/FR ratio. This low R/FR ratio-induced elongation significantly increases endogenous concentrations of indole-3-acetic acid (IAA) and also of three gibberellins (GAs): GA₂₀, GA₁, and GA₈. Of these, it is likely GA₁ that drives low R/FR-induced growth. Brassinosteroids are also involved in shoot growth. Here we tested three R/FR ratios: high, normal, and low. Significant hypocotyl elongation occurred with this stepwise reduction in R/FR ratio, but endogenous castasterone concentrations in the hypocotyls remained unchanged. Brassinolide was also applied to the seedlings and significantly increased hypocotyl growth, though one that was uniform across all three R/FR ratios. Applied brassinolide increased hypocotyl elongation while significantly reducing (usually) levels of IAA, GA₂₀, and GA₈, but not that of GA₁, which remained constant. Given the above, we conclude that endogenous castasterone does not mediate the hypocotyl growth that is induced by enriching FR light, relative to R light. Similarly, we conclude that the hypocotyl growth that is induced by applied brassinolide does not result from an interaction of brassinolide with changes in light quality. The ability of applied brassinolide to influence IAA, GA₂₀, and GA₈ content, yet have no significant effect on GA₁, is hard to explain. One speculative hypothesis, though, could involve the brassinolide-induced reductions that occurred for endogenous IAA, given IAA’s known ability to differentially influence the expression levels of GA20ox, GA3ox, and GA2ox, key genes in GA biosynthesis.     

Ethylene-Mediated Regulation of Gibberellin Content and Growth in Helianthus annuus L

Elongation of hypocotyls of sunflower can be promoted by gibberellins (GAs) and inhibited by ethylene. The role of these hormones in regulating elongation was investigated by measuring changes in both endogenous GAs and in the metabolism of exogenous [³H]- and [²H₂]GA₂₀ in the hypocotyis of sunflower (Helianthus annuus L. cv Delgren 131) seedlings exposed to ethylene. The major biologically active GAs identified by gas chromatography-mass spectrometry were GA₁, GA₁₉, GA₂₀, and GA₄₄. In hypocotyls of seedlings exposed to ethylene, the concentration of GA₁, known to be directly active in regulating shoot elongation in a number of species, was reduced. Ethylene treatment reduced the metabolism of [³H]GA₂₀ and less [²H₂]GA₁ was found in the hypocotyls of those seedlings exposed to the higher ethylene concentrations. However, it is not known if the effect of ethylene on GA₂₀ metabolism was direct or indirect. In seedlings treated with exogenous GA₁ or GA₃, the hypocotyls elongated faster than those of controls, but the GA treatment only partially overcame the inhibitory effect of ethylene on elongation. We conclude that GA content is a factor which may limit elongation in hypocotyls of sunflower, and that while exposure to ethylene results in reduced concentration of GA₁ this is not sufficient per se to account for the inhibition of elongation caused by ethylene.     

Comparison of Biological Activities of Gibberellins and Gibberellin-Precursors Native to Thlaspi arvense L

Field pennycress (Thlaspi arvense L.) is a winter annual weed with a cold requirement for stem elongation and flowering. The relative abilities of several native gibberellins (GAs) and GA-precursors to elicit stem growth were compared. Of the eight compounds tested, gibberellin A₁, (GA₁), GA₉, and GA₂₀ caused stem growth in noninduced (no cold treatment) plants. No stem growth was observed in plants treated with ent-kaurene, ent-kaurenol, ent-kaurenoic acid, GA₅₃, or GA₈. Moreover, of the biologically active compounds, GA₉ was the most active followed closely by GA₁. In thermoinduced plants (4-week cold treatment at 6°C) that were continuously treated with 2-chlorocholine chloride to reduce endogenous GA production, GA₉ was the most biologically active compound. However, the three kaurenoid GA precursors also promoted stem growth in thermoinduced plants, and were almost as active as GA₂₀. No such increase in activity was observed for either GA_[unk] or GA₅₃. The results are discussed in relation to thermoinductive regulation of GA metabolism and its significance to the initiation of stem growth in field pennycress. It is proposed that thermoinduction results in increased conversion of ent-kaurenoic acid to GAs through the C-13 desoxy pathway and that GA₉ is the endogenous mediator of thermoinduced stem growth in field pennycress.     

Comparative effects of gibberellic acid and N6-benzyladenine on dry matter partitioning and osmotic and water potentials in seedling organs of dwarf watermelon

Apical applications of 0.2 μg N⁶-benzyladenine (BA), a synthetic cytokinin, or 5 μg of gibberellic acid (GA₃) significantly enhanced hypocotyl elongation in intact dwarf watermelon seedlings over a 48-h period. Accompanying the increase in hypocotyl length was marked expansion of cotyledons in BA-treated seedlings and inhibition of root growth by both compounds. A study on dry matter partitioning indicated that both growth regulators caused a preferential accumulation of dry matter in hypocotyls at the expense of the roots; however, GA₃ elicited a more rapid and greater change than did BA. In comparison to untreated seedlings, BA decreased total translocation of metabolites out of the cotyledons. Water potentials of cotyledons and hypocotyls were determined by allowing organs to equilibrate for 2 h in serial concentrations of polyethylene glycol 4000. Osmotic potentials were determined by thermocouple psychrometry. During periods of rapid growth in cotyledons and hypocotyls of BA-treated seedlings and in hypocotyls of GA-treated seedlings, the osmotic potential increased and the turgor pressure decreased in relation to untreated seedlings, indicating that cell wall extensibility was being increased. Osmotic potentials were lower in hypocotyls of GA-treated than in those of BA-treated seedlings, even though growth rates were higher in GA-treated seedlings, indicating that the latter treatment was generating more osmotically active solutes in hypocotyls.     

Endogenous Gibberellins from Sporophytes of Two Tree Ferns, Cibotium glaucum and Dicksonia antarctica

Gibberellin A₁ (GA₁), 3-epi-GA₁, GA₄, GA₉, 11α-hydroxyGA₁₂, 12α-hydroxyGA₁₂, GA₁₅, GA₁₇, GA₁₉, GA₂₀, GA₂₅, GA₃₇, GA₄₀, GA₅₈, GA₆₉, GA₇₀, and GA₇₁ have been identified from Kovats retention indices and full scan mass spectra by capillary GC-MS analyses of purified extracts from sporophytes of the tree fern, Cibotium glaucum. Abscisic acid, dihydrophaseic acid, an epimer of 4′-dihydrophaseic acid, and the epimeric ent-6α, 7α, 16α, 17-(OH)₄ and ent-6α, 7α, 16β, 17-(OH)₄ derivatives of ent16, 17-dihydrokaurenoic acid, in addition to the epimeric 16α, 17- and 16β, 17-dihydroxy-16, 17-dihydro derivatives of GA₁₂, were also identified in extracts of C. glaucum. An oxodihydrophaseic acid and a hydroxydihydrophaseic acid were also detected. In extracts of sporophytes of Dicksonia antarctica, GA₄, GA₉, 12α- and 12β-hydroxyGA₁₂, GA₁₅, GA₂₅, and GA₃₇ were identified by the same criteria, as well as abscisic acid, phaseic acid, 8′-hydroxymethylabscisic acid and dihydrophaseic acid. This is the first time that GA₄₀ has been identified in a higher plant; it is also the first report of the natural occurrence of the two gibberellins, 11α- and 12β-hydroxyGA₁₂. The total gibberellin (GA) content in C. glaucum (tall) was at least one order of magnitude greater than that of D. antarctica (dwarf) based on total ion current response in GC-MS and bioassay data. Abscisic acid was a major component of D. antarctica and the oxodihydrophaseic acid was a major component of C. glaucum.     

Microbiological conversion of 12-oxygenated and other derivatives of ent-kaur-16-en-19-oic acid by Gibberella Fujikuroi, mutant B1-41a

The metabolism of several ring C and D-functionalized ent-kaur-16-en-19-oic acids by cultures of Gibberella fujikuroi, mutant B1-41a, to the corresponding derivatives of the normal fungal gibberellins (GAs) and ent-kaurenoids is described. A range of 12α- and 12β-hydroxyGAs and ent-kaurenoids are characterized by their mass spectra and GC Kovats retention indices. The mass spectral and GC data are used to identify the 12α-hydroxy derivatives of GA₁₂, GA₁₄, GA₃₇ and GA₄ (GA₅₈), and of the 12β-hydroxy derivatives of ent-7α-hydroxy- and ent-6α, 7α-dihydroxykaurenoic acids, in seeds of Cucurbita maxima. Similarly the metabolites of GA₉, formed in seeds of Pisum sativum and cultures of G.fujikuroi, mutant B1-41a, are identified as 12α-hydroxyGA₉. ent-11β-Hydroxy- and ent-11-oxo-kaurenoic acids are metabolized by the fungus to the corresponding 11-oxygenated derivatives of the normal fungal ent-kaurenoids and some C₂₀-GAs; no 11-oxygenated C₁₉-GAs are formed. Grandiflorenic acid, 11β-hydroxygrandiflorenic acid, attractyligen and ent-15β-hydroxykaurenoic acid are metabolized to unidentified products.     

Effect of temperature regimes on gibberellin levels in thermosensitive dwarf apple trees

Orchard-grown dwarf apple (Malus domestica Borkh.) trees selected from a hybrid population were propagated by tissue culture but had a growth pattern similar to standard cv. Golden Delicious plants when grown at constant 27°C instead of the expected dwarf pattern of growth. Shoot elongation was markedly reduced, with or without gibberellin A₁ (GA₁) or GA₄ treatment, when trees were grown in an environment where day temperature was maintained at 35°C for 2 h in a ramped regime (night 20°C day ramped to 35°C, held for 2 h and ramped down to 20°C night over a 14-h photoperiod). Application of GA₁ or GA₄ partially overcame growth retardation resulting from prior paclobutrazol treatment of both standard and dwarf trees grown at constant 27°C and of standard trees grown in the ramped environment. However, these GAs had no effect on paclobutrazol-treated or untreated dwarfs grown in the ramped regime. Gas chromatography-mass spectrometry with labelled internal standards was used to quantify GA₁, GA₃, GA₈, GA₁₉, GA₂₀ and GA₂₉ in extracts from standard and dwarf plants grown either at a constant 27°C or in a 20-30-20°C ramped temperature regime. Standard plants, which elongate quite rapidly in either environment, had similar levels of these GAs in both temperature regimes. The slowly growing dwarfs in the ramped temperature environment contained three times more GA₁₉ than the rapidly elongating dwarfs grown at 27°C. The concentrations of the other GAs were reduced to ca 40% or less in plants grown in the ramped temperature regime compared with those grown at 27°C. These data suggest that shoot elongation of dwarf plants is sensitive to elevated temperatures both as a result of reduced responsiveness to GAs and because of a reduction in the concentration of GA₁, apparently as a result of a lower rate of conversion of GA₁₉ to GA₂₀. It is possible that the altered GA metabolism may be a consequence of the change in GA sensitivity.     

Metabolism of ent-kaurenol-[17-14C], ent-kaurenal-[17-14C] and ent-kaurenoic acid-[17-14C] by germinating Hordeum distichon grains

Subcellular fractions from germinated barley embryos, chloroplast preparations and whole germinating barley grains are able to carry out the conversions ent-kaurenol → ent-kaurenal → ent-kaurenoic acid → ent-hydroxykaurenoic acid, the initial steps of the biosynthetic pathway to gibberellins. Whole grains, and chloroplasts to a slight extent, incorporate radioactivity from ent-kaurenol-[17-¹⁴C] and ent-kaurenoic acid-[17-¹⁴C] into materials with similar but distinct properties from the gibberellins GA₁, GA₃, GA₄ and GA₇.     

Detection of endogenous gibberellins and their relationship to hypocotyl elongation in soybean seedlings

Four gibberellins, GA₅₃, GA₁₉, GA₂₀, and GA₁, were detected by bioassay, chromatography in two HPLC systems, and combined gas chromatography-mass spectroscopy-selected ion monitoring (GC-MS-SIM) in etiolated soybean (Glycine max [L.] Merr.) hypocotyls. GC-MS-SIM employed [²H₂]-labeled standards for each endogenous gibberellin detected, and quantities estimated from bioassays and GC-MS-SIM were similar. This result plus the tentative detection of GA₄₄ and GA₈ (standards not available) indicates that the early-C-13-hydroxylation pathway for gibberellin biosynthesis predominates in soybean hypocotyls. Other gibberellins were not detected. Growth rates decreased after transfer to low water potential (ψ_w) vermiculite and were completely arrested 24 hours after transfer. The GA₁ content in the elongating region of hypocotyls had declined to 38% of the 0 time value at 24 hours after transfer to low ψ_w vermiculite, a level which was only 13% of the GA₁ content in control seedlings at the same time (24 hours posttransfer). Rewatering seedlings following 24 hours growth in low ψ_w vermiculite resulted in a complete recovery in elongation rate, an increase in GA₁ (20% at 2 hours, two-fold at 8 hours, eightfold at 24 hours), and a decrease in ABA levels (tenfold at 2 hours). Treatment of well-watered seedlings with the GA-synthesis inhibitor tetcyclacis (TCY) resulted in lowered GA₁ levels and increased ABA levels. When seedlings grown 24 hours in low ψ_w vermiculite were rewatered with TCY, recovery of the elongation rate was delayed and reduced, and the decline in ABA levels was slowed. Addition of GA₃ restored the elongation rate inhibited by TCY. Seedlings were growth responsive to exogenous GA₃, and this GA₃-promoted growth was inhibited by exogenous ABA. The data are consistent with the hypothesis that changes in GA₁ and ABA levels play a role in adjusting hypocotyl elongation rates. However, the changes observed are not of sufficient magnitude nor do they occur rapidly enough to suggest they are the primary regulators of elongation rate responses to rapidly changing plant water status.     

Native gibberellins and the metabolism of [14C]gibberellin A53 and of [17-13C, 17-3H2]gibberellin A20 in tassels of Zea mays

The native hormones from tassels of maize (Zea mays) were re-investigated. The previous identification by GC/SIM of GA₁, GA₈ and GA₂₉ in normal tassels was confirmed by full GC/MS scans at the correct Kovats retention indices. In tassels of dwarf-1 mutants, GA₄₄,^?GA₁₉, GA₁₇, GA₂₀ and the 16,17-dihydro, 7β,16α,17-trihydroxy derivative of ent-kaurenoic acid were identified by GC/MS. Gibberellin A₁ was not found in the mutant tassels. [¹⁴C]Gibberellin A₅₃ was fed to tassels of the dwarf-5 mutant. In the ethyl acetate-soluble acidic fraction from the feeds, [¹⁴C]GA₄₄ was identified by GC/MS; [¹⁴C]GA₁₉ and [¹⁴C]GA₂₉ were identified by GC/SIM. The GA₂₉ is probably a metabolite of the feeds because the dwarf-5 mutant is known to control the step copalyl pyrophosphate to ent-kaurene in the maize GA-biosynthetic pathway and because GA₂₉ was not identified in a control experiment. The n-butanol fractions obtained from the feeds were shown, by GC/MS, to contain [¹⁴C]GA₅₃ after hydrolysis, suggesting that conjugated [¹⁴C]GA₅₃ is a major metabolite from GA₅₃ feeds. [17-¹³C, 17-³H₂]Gibberellin A₂₀ was fed to normal, dwarf-1 and dwarf-5 tassels. In each case, analysis of the purified ethyl acetate-soluble acidic extracts by GC/MS led to the identification of [¹³C]GA₂₉ and unmetabolized [¹³C]GA₂₀ in which no ¹³C-isotope dilution was observed.