Quantitative Analysis of Endogenous Gibberellins in Normal and Dwarf Cultivars of Rice

Endogenous levels of gibberellins in shoots and ears of twodwarf rice (Oryza sativa L.) cultivars, Tan-ginbozu (dx mutant)and Waito-C (dy mutant), were analyzed and compared with thoseof normal rice cultivar, Nihonbare. The endogenous levels of13-hydroxylated gibberellins in Tan-ginbozu were much lowerthan those in Nihonbare. In Waito-C, the levels of GA₁₉ andGA₂₀ in the shoots were higher but that of GA₁ was lower thanthe levels of these gibberellins in Nihonbare. These resultssupport the hypothesis that the dy gene controls the 3ß-hydroxylationof GA₂₀ to GA₁ while the dx gene controls a much earlier stepin the gibberellin biosynthesis. Our results indicate that GA₁is the active gibberellin that regulates the vegetative growthof rice. The endogenous levels of GA₄ in the ears of the twodwarf cultivars of rice were higher than the level of GA₄ inthe ears of the normal cultivar, Nihonbare suggesting that thebiosynthesis of gibberellin is not blocked in the anthers ofthe dwarf rice. (Received April 27, 1989; Accepted July 12, 1989)     

Identification and Quantification of Cambial Region Hormones of Eucalyptus globulus

Gibberellins GA₁, GA₄, GA₈, GA₉, GA₁₉, GA₂₀, GA₂₉, GA₄₄, GA₈₁,indole-3-acetic acid (IAA) and abscisic acid (ABA) were identifiedin cambial region tissues of Eucalyptus globulus by comparingmass spectra and Kovats retention indices with those of authenticstandards. Using stable isotope labelled internal standardsGA₁₉, GA₂₀ and GA₄₄ were quantified at levels of 2–7 ng(g fr wt)^-1, other GAs were present at levels < 1 ng (g frwt)^-1. Levels of IAA and ABA ranged from 417–1, 140 ng(g fr wt)^-1 and 86–305 ng (g fr wt)^-1 respectively. Thepresence of brassinosteroid-like substances was also indicatedbased on activity in the rice seedling leaf inclination assay. (Received April 28, 1995; Accepted June 20, 1995)     

NaCl-induced Senescence in Leaves of Rice (Oryza sativa L.) Cultivars Differing in Salinity Resistance

The influence of NaCl on senescence-related parameters (proteinand chlorophyll concentrations, membrane permeability and chlorophyllfluorescence) was investigated in young and old leaves of fiverice cultivars differing in salt resistance. NaCl hastened thenaturally-occurring senescence of rice leaves which normallyappears during leaf ontogeny: it decreased chlorophyll and proteinconcentrations and increased membrane permeability and malondialdehydesynthesis. Such an acceleration of deteriorative processes affectedall leaves in salt-sensitive cultivars while it was more markedin oldest than in youngest leaves of salt-resistant genotypes.NaCl-induced senescence also involved specific modifications,such as an increase in basal non-variable chlorophyll fluorescence(F ₀) recorded in all cultivars or a transient increase in solubleprotein concentration recorded in salt-resistant genotypes only.Alteration of membrane permeability appeared as one of the firstsymptoms of senescence in rice leaves and allowed discriminationamong cultivars after only 7 d of stress. In contrast, F _v/F _mratio (variable fluorescence/maximal fluorescence) was thesame for all cultivars during the first 18 d of stress and thuscould not be used for identifying salt-resistant rice exposedto normal light conditions. Relationships between parametersinvolved in leaf senescence are discussed in relation to salinityresistance of rice cultivars. Chlorophyll concentration; chlorophyll fluorescence; electrolyte leakage; magnesium; malondialdehyde; membrane permeability; NaCl; Oryza sativa L.; protein; rice; salinity resistance; senescence; UV absorbing substances     

Metabolism of Gibberellins in Cell-Free Extracts of Anthers from Normal and Dwarf Rice

Cell-free extracts were prepared from anthers of normal anddwarf rice (Oryza sativa L.), and the metabolism of radioisotope-labeledgibberellins in the extracts was analyzed by HPLC and gas chromatography-massspectrometry (GC/MS). GA₁₂ was converted to GA₁₅ and GA₃₄ inthe extracts. GA₂₀ was converted to GA¹, GA₈ and GA₂₉, but GA₉was converted only to GA₃₄. The extracts of the dwarf cultivar,Waito-C (dy mutant), showed the same 3ß-hydroxylationactivity as did those of the normal cultivar, Nihonbare, indicatingthat the dy gene is not expressed in the anthers. These resultssuggest that the regulation of the biosynthesis of gibberellinsin rice is organ-specific. (Received November 9, 1989; Accepted January 10, 1990)     

The Role of Gibberellins in Early Growth and Development of Sugar Beet

Two gibberellin(GA)-like compounds were found in both rootsand shoots of sugar beet plants using the barley endosperm bioassay.One had chromatographic properties similar to GA₃ and GA₁; theother was highly non-polar, relatively inactive in the endospermassay, and may be a new gibberellin. Presence of the GA_3/1-likecompound was confirmed with the dwarf rice bio-assay. The quantityof this GA was relatively high in the root compared with theshoot at the 3–4 leaf stage when the first supernumerarycambia are being formed in the root. As plants developed throughthe 8–9 leaf stage and the 15–16 leaf stage thequantity of GA per unit fresh weight of material decreased. Application of gibberellic acid (GA₃) to the roots of youngsugar beet plants caused a significant increase in root dryweight shortly after treatment and the rate at which supernumerarycambia were produced was increased. Application of GA₃ to asingle petiole caused a significant increase in both root andshoot dry weight. GA₃ applied to either root or shoot causeda reduction in the rate of leaf formation although total leafarea per plant and shoot dry weight were unaffected. The probablerole of GA-like substances in controlling the growth and developmentof young sugar beet plants is discussed.     

Seasonal Changes of GA1, GA19 and Abscisic Acid in Three Rice Cultivars

The levels of endogenous gibberellin A₁ and A₁₉ (GA₁ and GA₁₉)and abscisic acid (ABA) in three rice (Oryza sativa L.) cultivars,Nihonbare (normal), Tan-ginbozu (semid-warf) and Tong-il (semi-dwarf),were measured at various stages of internode elongation andear development. GA₁₉ was the main GA in Nihonbare and Tong-ilthroughout the life cycle but was not detected in Tan-ginbozu.The levels of GA₁ in the ears of all three cultivars were lowand reached their maxima after anthesis. Similarly, the earsof all three cultivars contained high ABA levels which peakedafter anthesis. Shoots contained low quantities of ABA throughoutthe life cycle. The roles of GAs and ABA are discussed withrespect to physiological phenomena, such as internode elongation,ear development and dwarfism. (Received May 9, 1981; Accepted July 18, 1981)     

Effects of a Plant-Growth Regulat or, Prohexadione-Calcium (BX-112), on the Endogenous Levels of Gibberellins in Rice

The effects of the plant-growth regulator, prohexadione-calcium,on the levels of the endogenous gibberellins in rice shootswere measured by GC-SIM using ²H-labeled gibberellins as internalstandards. The compound was applied at the 4-leaf stage andshoots were harvested 5 and 12 days after treatment. Plant lengthwas reduced to 78% and 66%, respectively, relative to controlby application of the compound at 1 and 30 mg m^–2, whenplant length was measured 12 days after treatment. The levelof GA, was reduced to 36% and 18%, respectively, relative tocontrol in the treated plants. The levels of GA₁₉and GA₂₀ increasedbut that of GA₄₄ was reduced in the treated plants. The levelof GA₅₃ was unchanged. These results suggest that primary modeof action of the compound in vivo is the inhibition of the 3ß-hydroxylationof GA₂₀ to GA₁ and further support the hypothesis that GA₁ notGA₁₉ nor GA₂₀ is active in promoting shoot elongation in rice. ³Present address: Frontier Research Program, RIKEN, Wakoshi,Saitama, 351-01 Japan.     

A Crucial Role for Gibberellins in Stress Protection of Plants

The hypothesis that there is an intimate relationship betweengibberellin levels and plant stress protection has been testedusing near-isogenic lines of a normal and dwarf barley (Hordeumvulgare L.). Application of paclobutrazol (a triazole), inducedstress protection in the normal line and application of GA₃to the dwarf reversed the inherent stress tolerance. Reversalof the dwarf phenotype by specific gibberellins (GAs) suggeststhat the conversion of GA₂₀ to GA₁ and GA₉ to GA₄ has been compromised.These observations indicate that modulation of specific GAsplays a key role in stress protection. The preferential useof non-chemical technologies including enzyme regulation, phytochromeA overexpression, and breeding for induction of stress tolerancein plants are discussed. (Received November 26, 1998; Accepted March 11, 1999)     

The role of gibberellins in the growth of floral organs of Pharbitis nil

Evidence that the synthesis of GA₃ is involved in the growthof floral orga'ns of Pharbitis nil is presented. GAs in floralorgans at different developmental stages were surveyed usingTLC followed by the bioassay with two dwarf rice seedlings,‘Tanginbozu’ and ‘Waito-C’. The amountof GAs in the petal and stamen increased rapidly after the petalemerged from calyx, reached a maximum 12 hr before anthesis,then declined markedly thereafter. The GA content in the calyxremained unchanged before and after anthesis, and that in thepistil increased after anthesis. Pharbitis flowers containedat least two active GAs, one of which was probably GA₃, theother appeared to be GA₁₉. GA₃ was detected in relatively largeamounts in both the petal and stamen during their rapid elongation.In the calyx, which showed little increase in fresh weight duringrapid flower growth, GA₉ was the dominant GA. Exogenously suppliedGA₃ promoted elongation of sections in excised young filaments.Sucrose was necessary for definite growth promotion by GA₃.GA₁₉ had little effect on filament elongation, and IAA was ratherinhibitive. (Received July 29, 1972; )     

Biological Activities of the Methyl Ester of Gibberellin A73, a Novel and Principal Antheridiogen in Lygodium japonicum

The synthetic methyl ester of GA₇₃ (GA₇₃-Me) and the naturalantheridiogen of Lygodium japonicum showed almost the same activityto induce the formation of antheridia in dark-grown protonemataof L. japonicum at concentrations of 10^-14 M and higher. Thus,it appears that the principal antheridiogen in L. japonicumis GA₇₃-Me. GA₇₃-Me inhibited formation of ar-chegonia in light-grownprothallia of L. japonicum at concentrations of 10^-11 M andhigher and induced germination of spores in the dark in thisspecies at the same range of concentrations. GA₇₃(free acidform) promoted growth of seedlings of dwarf rice and hypocotylsof cucumber seedlings at dosages of and above 1 and 100 ng/plant,respectively. Eight compounds related to GA₇₃-Me, includingantheridiogens of Anemia phyllitidis and Anemia mexicana, wereactive in inducing an-theridial formation in L. japonicum, althoughtheir activities were considerably lower than that of GA₇₃-Me. (Received August 24, 1988; Accepted November 28, 1988)     

Internode Length in Pisum. A New Allele at the le Locus

In Pisum sativum L. a third, more severe, allele at the internodelength locus le is identified and named le^d. Plants homozygousfor le^d possess shorter internodes and appear relatively lessresponsive to GA₂₀ than comparable le (dwarf) plants. Gene le^dmay act by reducing the 3ß-hydroxylation of GA₂₀ tothe highly active GA₁ more effectively than does gene le. Theresults indicate that le is a leaky mutant and therefore thatendogenous GA₁ influences internode elongation in dwarf (le)plants. Pisum sativum, peas, internode length, genetics, gibberellin, dwarf elongation     

A lack of correlation between the biological activity and rate of metabolism of ent-(3H)-17-kaurenoic acid by seedlings of dwarf rice cv. Tan-ginbozu

Significant leaf sheath elongation occurred within 24 hr afterapplication of 10 µg (0.67, µCi) of ent-(³H)-17-kaurenoicacid (KA) to individual seedlings of dwarf rice cv. Tan-ginbozu,but this growth was unaccompanied by production of significantlevels of radioactivity in more polar, acidic, ethyl acetate-solublemetabolites of (³H)-KA. However modest levels of radioactivityappeared in the highly water-soluble fraction by hour 24, subsequentto the most rapid phase of KA-induced growth. Growth continuedand by hour 48 was accompanied by the appearance of small amountsof radioactivity in polar, acidic products. It would appearthat KA per se, and not its metabolic products, may be responsiblefor the leaf sheath elongation noted at hour 24. On the speculation that it might be a metabolite of KA, gibberellinA₁₄ (GA₁₄) was applied simultaneously with (³H)-KA to individualrice seedlings. Several changes in the metabolism of ³H-KA inthe presence of GA₁₄ were noted, and GA₁₄ antagonized the KA-inducedsheath elongation. ¹Present address: Botany Department, Rhodes University, Grahamstown,6140, South Africa. ²Present address: Crop Science Department, University of Saskatchewan,Saskatoon, Sask. S7N OWO, Canada. (Received May 12, 1975; )     

Endogenous Gibberellins in the Vegetative Shoots of Tall and Dwarf Cultivars of Phaseolus vulgaris L.

A series of 13-hydroxygibberellins, gibberellin A₁ (GA₁), GA₁₉,GA₂₀, GA₄₄ and GA₅₃, were identified by GC/MS (full scan) fromvegetative shoots of tall (cv. Kentucky Wonder) and dwarf (cv.Masterpiece) Phaseolus vulgaris L. It is suggested that GA₁is active per se in the control of shoot elongation of P. vulgarisL., and that dwarfism in Masterpiece is not due to shortageof the active GA, but to its low ability to respond to the bioactiveGA. (Received August 29, 1988; Accepted November 21, 1988)     

Effects of Gibberellins on Seed Germination of Phytochrome-Deficient Mutants of Arabidopsis thaliana

Experiments were carried out to explore the involvement of gibberellins(GAs) in the light-induced germination of Arabidopsis thaliana(L.) Heynh, using wild type (WT) and phytochrome-deficient mutants(phyA, phyB and phyAphyB deficient in phytochrome A, B and Aplus B, respectively). Seed germination of WT and phytochrome-deficientmutants was inhibited by uniconazole (an inhibitor of an earlystep in biosynthesis of GA, the oxidation of ent-kaurene) andprohexadione (an inhibitor of late steps, namely, 2rß-and 3rß-hydroxylation). This inhibition was overcomeby simultaneous application of 10^-5 M GA₄. The relative activityof GAs for promoting germination of uniconazole-treated seedswas GA₄>GA₁=GA₉>GA₂₀. The wild type and the phyA and phyBmutants had an increased response to a red light pulse in thepresence of GA₁, GA₄, GA₉, GA₂₀ and GA₂₄ but there were no significantdifferences in activity of each GA between the mutants. Therefore,neither phytochrome A nor hytochrome B appears to regulate GAbiosynthesis from GA₁₂ to GA₄ during seed germination, sincethe conversion of GA₁₂ to GA₉ is regulated by one enzyme (GA20-oxidase). However, GA responsiveness appears to be regulatedby phytochromes other than phytochromes A and B, since the phyAphyBdouble mutant retains the photoreversible increased responseto GAs after a red light pulse. (Received February 13, 1995; Accepted July 11, 1995)     

Effects of Plant Growth Regulators on Grain-filling and Yield of Rice

Effects of three phytohormones (IAA₁, GA₃ and kinetin) on grain-fillingand the pattern of ³²P translocation from individual leavesto grains were studied at intervals of 7 days during the progressof reproductive development of rice (Oryza saliva L. cv. Jaya).The plants were sprayed with 100 µg ml^–1 aqueoussolutions of the hormones at 100 days, when the plants wereentering the reproductive stage. Kinetin produced a pronouncedeffect on grain-filling as well as on ³²P mobilization fromindividual leaf to grains and increased yield, possibly by increasingleaf longevity. GA₃ and IAA also increased the grain-fillingand ³²P mobilization significantly over control but the effectswere less marked than those of kinetin. Oryza sativa L., rice, grain yield, translocation, growth regulators, gibberellic acid, indol-3-yl acetic acid, kinetin     

Peroxidase Ontogeny in a Dwarf Pea Stem as Affected by Gibberellin and Decapitation

The ontogeny of peroxidase activity and isoenzyme pattern wasinvestigated in the stem of dwarf pea plants. Peroxidase activityper unit soluble protein was a given internode is highest inthe youngest growth stage, drops during elongation, remainsconstant upon cessation of growth, and increase at senescence.The lower the internode on the stem the higher is its peroxidaseactivity. These developmental differences are already apparentat the youngest growth stage of the internodes adn increaseduring elongation. Several anodic and five cathodic isoperoxidasesare apparent after starch gel electrophoresis. This patternis constant for all internodes at all growth stages, but therelative importance of particular isoenzymes changes with time. Gibberellic acid (GA₃) treatment causes greatly elongated internodes,decreased soluble protein, and inhibition of the rise in peroxidaseactivity within 4–8 h. Application of GA₃ to young internodesleads to a persistent depression in peroxidase activity, whiletreated older internodes suffer only a temporary depression.GA₃ causes no qualitative changes in the isoenzyme pattern butproduces some quantitative alterations in internodes in whichits influence on peroxidase activity is persistent. Decapitation of untreated and GA₃-treated dwarfs has littleinfluence on internode elongation, causes an increase in peroxidaseactivity, especially in the upper internodes, and alters therelative activity of particular isoenzymes. By contrast, decapitationinhibits elongation of young internodes in genetically tallpea plants.     

The role of gibberellins in the growth of floral organs of Mirabilis jalapa

Evidence that the anther is a source of GAs which may be a mainfactor limiting the growth of floral organs of Mirabilis jalapais presented. GAs in floral tubes at different developmentalstages were surveyed using TLC followed by bioassay with ‘Tan-ginbozu’dwarf rice seedlings. The amount of GA_s in the floral tube increasedrapidly after the calyx became visible above the bract 8 daysbefore anthesis, reached a maximum 5 days before anthesis, thendeclined markedly thereafter. The stamen had relatively largeamounts of GAs and mainly accounted for the change of GA_s duringthe flower development. The GA content in the anther was 3 timesas much as that in the filament. Exogenously supplied GA3 promotedelongation of sections in excised flower organs. Extracts ofanthers also showed definite growth promotion of style sections.Removal of the anthers greatly reduced the final length of thefloral tube. (Received December 7, 1974; )     

Effects of Gibberellins and Prohexadione on the Activities of Oryzain and {alpha}-Amylase in Rice Seeds

Oryzains, cysteine proteinases of rice seeds, are induced byGA₃ in germinating rice seeds [Abe et al. (1987) Agric. Biol.Chem. 51: 1509]. The effects of GA₁, GA₃, GA₄, GA₉, and GA₂₀on the production of oryzain and -amylase were investigatedin embryoless half- and whole-seeds of rice (cv. Nipponbare).When gibberellins (GAs) were incubated with embryoless half-seeds,GA₁, GA₃ and GA₄ induced oryzain and -amylase, but GA₉, andGA₂₀ did not. GA₉ and GAM induced oryzain and -amylase productionin whole seeds, but this production was inhibited by the simultaneousapplication of prohexadione, an inhibitor of 2ß- and3ß-hydroxylation of GAs. Prohexadione did not inhibitthe activities of oryzain and -amylase induced by GA₁. Theseresults suggest that GAs possessing the 3ß-hydroxylgroup induce activities of oryzain and -amylase in rice seedsand that GA₉ and GA₂₀ have activity only after they are convertedmetabolically to active GAs, probably GA₄ and GA₁, respectively.GA₁, was more active than GA₄ in both half seeds and wholeseeds incubation. Oryzain and -amylase activities induced byGA₄ were significantly inhibited in the presence of 10^–4M prohexadione. This suggests that the conversion of GA₁, toGA₄ (13-hydroxylation) might be inhibited at a high dose ofprohexadione in whole seeds. ⁴Present address: Institute of Food Development, Kyung Hee University,Suwon 449-701, Korea     

The regulation of leaf elongation and xyloglucan endotransglycosylase by gibberellin in 'Himalaya' barley (Hordeum vulgare L.)

The potential role of xyloglucan endotransglycosylase (XET)in GA-stimulated cell elongation was investigated during leafexpansion in barley (Hordeum vulgare L.). XET activity in aqueousextracts of leaves was detected in all segments along the elongatingblade of leaf 1 of seedlings, but was at highest levels in basalsegments. Leaf 1 elongation rates of gibberellin (GA)-responsivedwarf mutants were lower than the wild type, and accompaniedby reduced levels of XET activity. Leaf elongation rates ofthe dwarfs increased following treatment with gibberellic acid(GA₃) associated with higher levels of XET activity. The slendermutant, crossed into a dwarfing background, exhibited high ratesof leaf 1 elongation and high levels of XET activity withoutadded GA₃. The elongation of leaf 3 in a GA-responsive dwarfmutant was also studied. Treatment with GA₃ resulted in bladeand sheath lengths being 5-fold and 7-fold (respectively) thelengths of controls, and again there were increases in bladeand sheath XET activities. To investigate the basis for changesin XET activity levels two XET-related cDNA clones were isolated.RNAs detected by the two clones occurred at the highest levelsin basal segments of rapidly elongating leaves, but they haddifferent distribution patterns along the leaf. Overall, thedata indicate that an XET-like activity is detectable in barleyleaves, that the activity level and related. Key words: Gibberellin (GA), leaf elongation, Hordeum vulgare, xyloglucan endotransglycosylase (XET)     

THE INDEPENDENT ACTION OF MORPHACTINS AND GIBBERELLIC ACID ON HIGHER PLANTS

The comparative activity of three morphactins, n-butyl-9-hydroxyfluorene-9-carboxylate (IT 3233), 9-hydroxy-fluorene-9-carboxylicacid (IT 3235) and methyl-2-chloro-9-hydroxy fluorene-9-carboxylate(IT 3456) on the action of gibberellic acid (GA₃) were examinedutilizing dwarf pea (Progress No. 9), dwarf corn (strain d-5),CCC dwarfed Alaska pea, and embryoless barley half-seeds. When applied either prior to or simultaneously with GA₃ themorphactins were without effect in reducing the response ofthe dwarf peas to the gibberellin. In fact IT 3233 and IT 3235in combination with GA₃ produced taller plants than the GA₃controls. Similar results were observed using CCC dwarfed Alaskapeas. The morphactins did, however, produce several morphologicaleffects on the peas. In addition, they were effective in breakingapical dominance in the peas. The morphactins were also withouteffect on the dwarf corn bioassay and no morphological changeswere observed. In the embryoless barley half-seeds the morphactins did notinhibit the response of the tissue to the GA₃. Also, the morphactinsdid not exhibit gibberellin-like properties. The results suggest that morphactins do not compete with gibberellinfor similar sites of action. ¹Published with the approval of the Director of the MichiganAgricultural Experiment Station as Journal Article Number 3917.