Inhibition of stem growth and gibberellin production in Agrostemma githago L. by the growth retardant tetcyclacis

The effects of the new growth retardant tetcyclacis (TCY) on stem growth and endogenous gibberellin (GA) levels were investigated in the long-day rosette plant Agrostemma githago. Application of TCY (10 ml of a 5·10^-5M solution daily) to the soil suppressed stem elongation in Agrostemma grown under long-day conditions. A total of 10 g GA₁ (1 g applied on alternate days) per plant overcame the growth retardation caused by TCY.Control plants and plants treated with TCY were analyzed for endogenous GAs after exposure to nine long days. The acidic extracts were fractionated by high-performance liquid chromatography. Part of each fraction was tested in the d-5 maize bioassay, while the remainder was analyzed by combined gas chromatography-selected ion monitoring. The bioassay results indicated that the GA content of plants treated with TCY was much lower than that of untreated plants. The data obtained by gas chromatography-selected ion monitoring confirmed that the levels of seven GAs present in Agrostemma were much reduced in TCY-treated plants when compared with the levels in control plants: GA₅₃ (13%), GA₄₄ (0%), GA₁₉ (1%), GA₁₇ (33%), GA₂₀ (15%), GA₁ (4%), and epi-GA₁ (13%). These results provide evidence that TCY inhibits stem growth in Agrostemma by blocking GA biosynthesis and thus lowering the levels of endogenous GAs.Abbreviations AMO-1618 2-isopropyl-4-dimethylamino-5-methylphenyl-1-piperidine-carboxylate methyl chloride - GA(s) gibberellin(s) - HPLC high-performance liquid chromatography - TCY Tetcyclacis (5-[4-chlorophenyl]-3,4,5,9,10-pentaaza-tetracyclo-5,4,1,0^2,6,0^8,11-dodeca-3,9-diene)     

Rice inositol polyphosphate kinase gene (<Emphasis Type="Italic">OsIPK2</Emphasis>), a putative new player of gibberellic acid signaling,involves in modulation of shoot elongation and fertility

Gibberellic acid (GA) is an important plant hormone mediating plant growth and development throughout the life span. Although many GA biosynthesis genes and signaling components have been revealed, the signal transduction mechanisms from GA perception to physiological actions are still largely unclear. In this study, we investigated the functions of a rice (Oryza sativa) inositol polyphosphate kinase gene (OsIPK2) in rice growth and development, showing that OsIPK2 is a putative new player in GA signaling. OsIPK2 is widely expressed in rice with high accumulation in tender and rapidly dividing tissues. The OsIPK2 protein is mainly localized in the nucleus and plasma membrane. To study the biological roles of OsIPK2 in rice, RNA interference and overexpression transgenic plants were generated. OsIPK2 antisense plants exhibited taller seedling height and lower fertility rate than the wild type, while overexpression lines showed reduced plant height. Microarray and qRT-PCR assays showed that expression levels of several GA-related genes were altered in transgenic plants. Besides, down-regulation of OsIPK2 resulted in hypersensitivity to paclobutrazol (PAC), a GA biosynthesis inhibitor. We also described that the expression of OsIPK2 could be either induced by GA or repressed by PAC. Taken together, these findings suggested that OsIPK2 is likely a negative regulator of GA signaling and involves in modulating shoot elongation and fertility.     

A role of <Emphasis Type="Italic">OsGA20ox1</Emphasis>, encoding an isoform of gibberellin 20-oxidase,for regulation of plant stature in rice

Gibberellin (GA) 20-oxidase (GA20ox) is a key enzyme that normally catalyzes the penultimate steps in GA biosynthesis. One of the GA20ox genes in rice (Oryza sativaL.), OsGA20ox2 (SD1), is well known as the Green Revolution gene, and loss-of function mutation in this locus causes semi-dwarfism. Another GA20ox gene, OsGA20ox1, has also been identified, but its contribution to plant stature has remained unclear because no suitable mutants have been available. We isolated a mutant, B142, tagged with a T-DNA containing three CaMV 35S promoters, which showed a tall, GA-overproduction phenotype. The final stature of the B142 mutant reflects internode overgrowth and is approximately twice that of its wild-type parent. This mutant responds to application of both GA3 and a GA biosynthesis inhibitor, indicating that it is a novel tall mutant of rice distinct from GA signaling mutants such as slr1. The integrated T-DNAs, which contain three CaMV 35S promoters, are located upstream of the OsGA20ox1 open reading frame (ORF) in the B142 mutant genome. Analysis of mRNA and the endogenous GAs reveal that biologically active GA level is increased by up-regulation of the OsGA20ox1 gene in B142. Introduction of OsGA20ox1 cDNA driven by 35S promoter into the wild type phenocopies the morphological characteristics of B142. These results indicate that the elongated phenotype of the B142 mutant is caused by up-regulation of the OsGA20ox1 gene. Moreover, the final stature of rice was reduced by specific suppression of the OsGA20ox1 gene expression. This result indicates that not only OsGA20ox2 but also OsGA20ox1 affects plant stature.     

Characterization of cis-regulatory regions responsible for developmental regulation of the gibberellin biosynthetic gene GA1 in Arabidopsis thaliana

The Arabidopsis GA1 gene encodes copalyl diphosphate synthase, which catalyzes the first committed step in the gibberellin biosynthetic pathway. Previous studies indicated that the expression pattern of the GA1 gene is tissue-specific and cell-type-specific during development. Here we showed that expression of GA1 cDNA driven by the 2.4 kb 5-upstream sequence plus the GA1 genomic coding region into the third exon was able to rescue the ga1-3 mutant phenotype. To understand the mechanism controlling GA1 gene expression, cis-regulatory regions in the GA1 promoter were identified by promoter deletion analysis with the GA1--glucuronidase (GUS) gene fusion system. The second intron and the region from –1391 to –997, with respect to the translation initiation site, positively regulate overall GA1-GUS expression level in all tissues examined. Several additional regulatory regions are involved in GA1-GUS expression in all the stages except in seeds: two positive regulatory regions in the first intron and the sequence between –425 and –207, and a negative regulatory region between –1848 and –1391. We also found that the region between –997 and –796 is essential for a high level of GA1 expression in developing seeds.     

Potato purple top phytoplasma‐induced disruption of gibberellin homeostasis in tomato plants

Phytoplasmas are phloem‐inhabiting, cell wall‐less bacteria that cause numerous plant diseases worldwide. Plants infected by phytoplasmas often exhibit various symptoms indicative of hormonal imbalance. In this study, we investigated the effects of potato purple top (PPT) phytoplasma infection on gibberellin homeostasis in tomato plants. We found that PPT phytoplasma infection caused a significant reduction in endogenous levels of gibberellic acid (GA₃). The decrease in GA₃ content in diseased plants was correlated with down regulation of genes responsible for biosynthesis of bioactive GAs ( GA20ox1 and GA3ox1) and genes involved in formation of GA precursors [geranyl diphosphate synthase (GPS) and copalyldiphosphate synthase (CPS)]. Exogenous application of GA₃ at 200 µmol L^?1 was able to restore the GA content in infected plants to levels comparable to those in healthy controls, and to attenuate the characteristic ‘big bud’ symptoms induced by the phytoplasma. The interesting observation that PPT phytoplasma‐infected plants had prolonged low expression of key GA biosynthesis genes GA20ox1 and GA3ox1 under GA deficiency conditions led us to hypothesise that there was a diminished sensitivity of the GA metabolism feedback regulation, especially GA biosynthesis negative feedback regulation, in those affected plants, and such diminished sensitization in early stages of infection may represent a central element of the phytoplasma‐induced disruption of GA homeostasis and pathogenesis.     

Role of the gibberellin receptors GID1 during fruit‐set in Arabidopsis

Gibberellins (GAs) play a critical role in fruit‐set and fruit growth. Gibberellin is perceived by its nuclear receptors GA INSENSITIVE DWARF1s (GID1s), which then trigger degradation of downstream repressors DELLAs. To understand the role of the three GA receptor genes (GID1A, GID1B and GID1C) in Arabidopsis during fruit initiation, we have examined their temporal and spatial localization, in combination with analysis of mutant phenotypes. Distinct expression patterns are revealed for each GID1: GID1A is expressed throughout the whole pistil, while GID1B is expressed in ovules, and GID1C is expressed in valves. Functional study of gid1 mutant combinations confirms that GID1A plays a major role during fruit‐set and growth, whereas GID1B and GID1C have specific roles in seed development and pod elongation, respectively. Therefore, in ovules, GA perception is mediated by GID1A and GID1B, while GID1A and GID1C are involved in GA perception in valves. To identify tissue‐specific interactions between GID1s and DELLAs, we analyzed spatial expression patterns of four DELLA genes that have a role in fruit initiation (GAI, RGA, RGL1 and RGL2). Our data suggest that GID1A can interact with RGA and GAI in all tissues, whereas GID1C–RGL1 and GID1B–RGL2 interactions only occur in valves and ovules, respectively. These results uncover specific functions of each GID1–DELLA in the different GA‐dependent processes that occur upon fruit‐set. In addition, the distribution of GA receptors in valves along with lack of expression of GA biosynthesis genes in this tissue, strongly suggests transport of GAs from the developing seeds to promote fruit growth.     

Over-expression of a gibberellin 2-oxidase gene from Phaseolus coccineus L. enhances gibberellin inactivation and induces dwarfism in Solanum species

Gibberellins (GAs) are endogenous hormones that play a predominant role in regulating plant stature by increasing cell division and elongation in stem internodes. The product of the GA 2-oxidase gene from Phaseolus coccineus (PcGA2ox1) inactivates C₁₉-GAs, including the bioactive GAs GA₁ and GA₄, by 2β-hydroxylation, reducing the availability of these GAs in plants. The PcGA2ox1 gene was introduced into Solanum melanocerasum and S. nigrum (Solanaceae) by Agrobacterium-mediated transformation with the aim of decreasing the amounts of bioactive GA in these plants and thereby reducing their stature. The transgenic plants exhibited a range of dwarf phenotypes associated with a severe reduction in the concentrations of the biologically active GA₁ and GA₄. Flowering and fruit development were unaffected. The transgenic plants contained greater concentrations of chlorophyll b (by 88%) and total chlorophyll (11%), although chlorophyll a and carotenoid contents were reduced by 8 and 50%, respectively. This approach may provide an alternative to the application of chemical growth retardants for reducing the stature of plants, particularly ornamentals, in view of concerns over the potential environmental and health hazards of such compounds. C. Dijkstra, E. Adams, A. Bhattacharya and A. F. Page contributed equally to this paper.     

Accelerated shoot overgrowth of rice mutant <Emphasis Type="Italic">ao-1</Emphasis> is epistatic to gibberellin-sensitive and -insensitive dwarf mutants

 A shoot overgrowth mutant of rice (Oryza sativa L.), accelerated internode overgrowth-1 (ao-1), is marked by accelerated longitudinal elongation of aerial parts and overgrowth of internodes at the vegetative stage. The physiological properties of ao-1 were similar to those of wild plants treated with a saturating level of exogenous gibberellins (GAs), except for the internode-overgrowth phenotype, which was not mimicked by GA-treated wild plants. The ao-1 mutant was less sensitive to a GA biosynthesis inhibitor, Uniconazole-P, than the wild type. Dwarf alleles of three loci, including two GA-sensitive and one GA-insensitive mutation, were introduced to produce double-mutants with ao-1, but the overgrowth phenotype was not suppressed in double-homozygous mutants. These results suggest that the overgrowth phenotype of ao-1 is caused by abolition of GA signaling rather than by GA overproduction. It is likely that a part of the shoot regulation system of ao-1 is saturated with the GA signal. As a possible model consistent with the results, we propose that AO-1 protein acts as a negative regulator in GA signal transduction. Received: August 14, 2001 / Accepted: February 8, 2002     

Identification of gibberellins in the rice plant and quantitative changes of gibberellin A19 throughout its life cycle

The major endogenous gibberellin (GA) in shoots, roots and ears of the rice plant, Oryza sativa L. japonica cv. Nihonbare, was identified as GA₁₉ by combined gas liquid chromatography-mass spectrometry (GC-MS) and GC-selected ion current monitoring (GC-SICM). Another GA present in these tissues in small quantity was tentatively identified as GA₁ by GC-SICM, and GA₄ may be present in the seeds (kernels) of 3rd-leaf-stage seedlings. Using GC-SICM, the GA₁₉ content was quantified throughout the life cycle of rice plants. It was found to reach high levels (ca. 10–15 g/kg fresh weight) in 3rd-leaf seedlings, at panicle initiation (shoots), and during heading and anthesis (ears). The levels of GA₁₉ in Oryza sativa indica cv. T-136 underwent changes closely similar to those found in Nihonbare. The growth-promoting activity in rice of exogenous GA₁₉ is generally considerably less than that of GA₁. It therefore seems possible that GA₁₉ functions as a pool GA. The level of active GAs such as GA₁ may be regulated by the rate of biosynthesis of GA₁₉ or its metabolic conversions.Abbreviations GA(s) gibberellin(s) - GA_n gibberellin A_n - GA_n-MeTMS trimethylsilyl ether of GA_n methyl ester - GC-MS combined gas liquid chromatography-mass spectrometry - GC-SICM combined gas liquid chromatography-selected ion current monitoring - TLC thin-layer chromatography     

Rice <Emphasis Type="Italic">PLASTOCHRON</Emphasis> genes regulate leaf maturation downstream of the gibberellin signal transduction pathway

Rice PLASTOCHRON 1 (PLA1) and PLA2 genes regulate leaf maturation and plastochron, and their loss-of-function mutants exhibit small organs and rapid leaf emergence. They encode a cytochrome P450 protein CYP78A11 and an RNA-binding protein, respectively. Their homologs in Arabidopsis and maize are also associated with plant development/organ size. Despite the importance of PLA genes in plant development, their molecular functions remain unknown. Here, we investigated how PLA1 and PLA2 genes are related to phytohormones. We found that gibberellin (GA) is the major phytohormone that promotes PLA1 and PLA2 expression. GA induced PLA1 and PLA2 expression, and conversely the GA-inhibitor uniconazole suppressed PLA1 and PLA2 expression. In pla1-4 and pla2-1 seedlings, expression levels of GA biosynthesis genes and the signal transduction gene were similar to those in wild-type seedlings. GA treatment slightly down-regulated the GA biosynthesis gene GA20ox2 and up-regulated the GA-catabolizing gene GA2ox4, whereas the GA biosynthesis inhibitor uniconazole up-regulated GA20ox2 and down-regulated GA2ox4 both in wild-type and pla mutants, suggesting that the GA feedback mechanism is not impaired in pla1 and pla2. To reveal how GA signal transduction affects the expression of PLA1 and PLA2, PLA expression in GA-signaling mutants was examined. In GA-insensitive mutant, gid1 and less-sensitive mutant, Slr1-d1, PLA1 and PLA2 expression was down-regulated. On the other hand, the expression levels of PLA1 and PLA2 were highly enhanced in a GA-constitutive-active mutant, slr1-1, causing ectopic overexpression. These results indicate that both PLA1 and PLA2 act downstream of the GA signal transduction pathway to regulate leaf development.     

SPYing on GA Signaling and Plant Development

It has been ten years since the SPINDLY (SPY) locus was first identified from a screen of mutagenized wild type Arabidopsis seeds by selecting for germination in the presence of a gibberellin (GA) biosynthesis inhibitor (Jacobsen and Olszewski 1993). Since then research into this novel protein, an O-GlcNAc transferase (OGT), has revealed some fascinating and surprising results. SPY was originally described as a negative regulator specific to the GA signal transduction pathway, but recent research suggests that SPY is involved in additional aspects of plant development. SPY is also being investigated in barley, petunia and rice, adding to the complex story that is SPY.     

Isolation and expression profiles of gibberellin metabolism genes in developing male and female cones of Pinus tabuliformis

Gibberellins (GAs) are important in the floral regulatory networks of angiosperm plants. Several lines of evidence suggest that GAs also play a pivotal role in conifer male and female cone development. To gain new insights into the GA metabolism pathway in conifer trees and the role of GA metabolism in male and female cone development, we identified GA metabolism genes in Pinus tabuliformis. These included one PtCPS gene, one PtKS gene, one PtKO gene, TWO PtKAO genes, one PtGA20ox gene, two PtGA3ox genes and 12 PtGA2ox genes. According to phylogenetic analysis, the GA biosynthesis pathway evolved after the divergence of mosses from ferns, but the GA-deactivating gene family underwent divided expansion after divergence of the angiosperms from gymnosperms. However, the active sites of all GA metabolism enzymes were conserved during the evolution of land plants. During male and female cone development of P. tabuliformis, the expression of most of the PtGA2ox genes, especially PtGA2ox10, was higher than GA biosynthesis genes. However, the expression of PtKAO1 in cones peaked at a very early developmental stage. The expression pattern of GA metabolism genes indicated that GAs play different roles at the early and late stages of cone development.     

高等植物赤霉素生物合成及其调节研究进展

主要介绍近年来高等植物中生物活性GAs的生物合成,拟南芥GA生物合成途径中关键酶基因（GA1－GA5）的克隆和GA3基因CYP701A3的母（Saccharomyces cerevisiae）中的成功表达。评述了活性GAs对赤霉不生物合成的反馈抑制作用和反馈调节中信号的传递和接收问题。高等植物中光周期对GA生物合成的调节主要是在20－氧化和／或3β－羟基化步骤。     

Physiology of gibberellin-induced elongation of epicotyl explants from Vigna sinensis

The physiological characteristics of the response of excised cowpea (Vigna sinensis cv Blackeye pea No. 5) epicotyls to gibberellins (GAs) were studied. Epicotyl explants, retaining the petioles and a 2-cm portion of hypocotyl, were placed upright in small vials containing water. Plant growth substances were injected into the subapical tissues as ethanol solutions.Epicotyl elongation resulting from treatment with 0.5 g of GA ranged between 5 and 13 times that of the control, depending on the GA applied. With GA₁, no differences were obtained with explants prepared from 5 to 9-day-old seedlings. The increase in elongation could be detected within 6 h of treatment, and the stimulus of a single application lasted at least 4 days. Final elongation was proportional to the logarithm of the amount of GA, applied, 0.01 to lug. The response to GA treatment was limited to the upper part, the most sensitive zone being located between 2 to 4 mm below the apex of the epicotyl; this effect was entirely due to cell elongation.The induction of epicotyl elongation by GAs seems to be specific and independent of the effect of auxin. IAA had no effect on elongation and 4-chloro-phenoxyisobutyric acid (PCIB) did not affect the response to GA₁ Abbreviations ABA abscisic acid - GA gibberellin - IAA Indole-3-acetic acid - TIBA 2,3,5-triiodobenzoic acid - PCIB 4-chloro-phenoxyisobutyric acid     

赤霉素信号转导的中间组分研究

具有生物活性的赤霉素(Gibberellins GAs)影响着植物生长和发育的许多进程,包括种子的萌发、叶的生长、茎的伸长,花的分化和果实的形成。随着GA信号转导途径中越来越多的中间组分的分离和鉴定,GA调控植物发育的分子机理也越来越清楚。就近年来GA信号转导途径的中间组分的研究做一综述。     

Chlorophyll antenna size adjustments by irradiance in Dunaliella salina involve coordinate regulation of chlorophyll a oxygenase (CAO) and Lhcb gene expression

To elucidate the mechanism of irradiance-dependent adjustments in the chlorophyll antenna size of photosynthesis, we addressed the regulation of expression of genes encoding a variety of chlorophyll biosynthesis enzymes and that of the Lhcb genes in the model organism Dunaliella salina. Among the chlorophyll biosynthesis enzymes tested, only the chlorophyll a oxygenase (CAO) gene responded to changes in the level of irradiance with substantial mRNA level and kinetics of change that were similar to those of the Lhcb genes. Evidence is presented for the operation of a cytosolic signal transduction pathway for the rapid (order of minutes) regulation of both CAO and Lhcb gene expression by irradiance. Inhibitor studies and transient activation of Ca²⁺-dependent kinase suggested phopholipase-C activation to Ca²⁺ release, and activation of a specific Ca²⁺/CaM-dependent protein kinase in this cytosolic signal transduction pathway. The redox state of the plastoquinone pool also serves to regulate CAO and Lhcb gene expression on a slower time scale (hours) and probably serves as a plastidic-origin signal that acts coordinately with the cytosolic signal transduction pathway. It is proposed that irradiance-dependent adjustments in the chlorophyll antenna size occur by coordinate regulation of CAO and Lhcb gene expression via two distinct signal transduction pathways in photosynthetic organisms.