Gibberellins promote vegetative phase change and reproductive maturity in maize.

Postembryonic shoot development in maize (Zea mays L.) is divided into a juvenile vegetative phase, an adult vegetative phase, and a reproductive phase that differ in the expression of many morphological traits. A reduction in the endogenous levels of bioactive gibberellins (GAs) conditioned by any one of the dwarf1, dwarf3, dwarf5, or another ear1 mutations in maize delays the transition from juvenile vegetative to adult vegetative development and from adult vegetative to reproductive development. Mutant plants cease producing juvenile traits (e.g. epicuticular wax) and begin producing adult traits (e.g. epidermal hairs) later than wild-type plants. They also cease producing leaves and begin producing reproductive structures later than wild-type plants. These mutations greatly enhance most aspects of the phenotype of Teopod1 and Teopod2, suggesting that GAs suppress part but not all of the Teopod phenotype. Application of GA3 to Teopod2 mutants and Teopod1, dwarf3 double mutants confirms this result. We conclude that GAs act in conjunction with several other factors to promote both vegetative and reproductive maturation but affect different developmental phases unequally. Furthermore, the GAs that regulate vegetative and reproductive maturation, like those responsible for stem elongation, are downstream of GA20 in the GA biosynthetic pathway.     

Genome‐wide identification and characterization of microRNAs responding to ABA and GA in maize embryos during seed germination

• MicroRNAs (miRNAs) are an important class of non‐coding small RNAs that regulate the expression of target genes through mRNA cleavage or translational inhibition. Previous studies have revealed their roles in regulating seed dormancy and germination in model plants such as Arabidopsis thaliana, rice (Oryza sativa) and maize (Zea mays). However, the miRNA response to exogenous gibberellic acid (GA) and abscisic acid (ABA) during seed germination in maize has yet to be explored.
• In this study, small RNA libraries were generated and sequenced from maize embryos treated with GA, ABA or double‐distilled water as control.
• A total of 247 miRNAs (104 known and 143 novel) were identified, of which 45 known and 53 novel miRNAs were differentially expressed in embryos in the different treatment groups. In total, 74 (37 up‐regulated and 37 down‐regulated) and 55 (23 up‐regulated and 32 down‐regulated) miRNAs were expressed in response to GA and to ABA, respectively, and a total of 18 known and 38 novel miRNAs displayed differential expression between the GA‐ and ABA‐treated groups. Using bioinformatics tools, we predicted the target genes of the differentially expressed miRNAs. Using GO enrichment and KEGG pathway analysis of these targets, we showed that miRNAs differentially expressed in our samples affect genes encoding proteins involved in the peroxisome, ribosome and plant hormonal signalling pathways.
• Our results indicate that miRNA‐mediated gene expression influences the GA and ABA signalling pathways during seed germination.

     

Inositol phosphate signaling and gibberellic acid

To respond to physical signals and endogenous hormones, plants use specific signal transduction pathways. We and others have previously shown that second messenger inositol 1,4,5-trisphosphate [Ins(1,4,5)P₃] is used in abscisic acid (ABA) signaling, and that some mutants with altered Ins(1,4,5)P₃ have altered responses to ABA. Specifically, mutants defective in the myo-inositol polyphosphate 5-phosphatases (5PTases) 1 and 2 genes that hydrolyze 5-phosphates from Ins(1,4,5)P₃ and other PtdInsP and InsP substrates, have elevated Ins (1,4,5)P₃, and are ABA-hypersensitive. Given the antagonistic relationship between ABA and gibberellic acid (GA), we tested the response of these same mutants to a GA synthesis inhibitor, paclobutrazol (PAC). We report here that 5ptase1, 5ptase2 and 5ptase11 mutants are hypersensitive to PAC, suggesting a relationship between elevated Ins(1,4,5)P₃ and decreased GA signal transduction. These data provide insight into signaling cross-talk between ABA and GA pathways.Key words: inositol, phosphatidylinositol phosphate, paclobutrazol, gibberellic acid, inositol trisphosphate, paclobutrazol     

Augmented Phytoextraction of Lead (Pb2+)-Polluted Soils: A Comparative Study of the Effectiveness of Plant Growth Regulators,EDTA, and Plant Growth–Promoting Rhizobacteria

Abstract

The role of gibberellic acid (GA3), indole-3-acetic acid (IAA), plant growth–promoting bacteria (Rhizobium and Azotobacter), and a synthetic chelator (EDTA; ethylenediaminetetraacetic acid) in lead (Pb) phytoextraction was evaluated using Parthenium hysterophorus (dicot, unpalatable noncrop) and Zea mays (monocot food/forage crop) plants at the flowering stage. Various plant parts were analyzed by atomic absorption/flame spectrophotometer for their Pb content. Both plant growth regulators and both growth-promoting bacteria significantly increased the plant growth in Pb-polluted soils, whereas EDTA significantly decreased growth and biomass of both plants. EDTA increased the Pb uptake (μg g^?1 dry biomass), but the total plant Pb accumulation was decreased. GA3 and IAA significantly increased both uptake and translocation, and the maximum total Pb in the entire plant of Parthenium was found with GA3 foliar spray, whereas in Z. mays the total Pb was maximum in the plant treated with GA3 in combination with EDTA, followed by the GA3 foliar spray treatment. Overall, the GA3 foliar application showed superior response compared with all other treatments. Further research is recommended to observe the role of endogenous GA3 levels in correlation with metal phytoextraction in different plants.     

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.     

Overexpression of GA20‐OXIDASE1 impacts plant height,biomass allocation and saccharification efficiency in maize

Increased biomass yield and quality are of great importance for the improvement of feedstock for the biorefinery. For the production of bioethanol, both stem biomass yield and the conversion efficiency of the polysaccharides in the cell wall to fermentable sugars are of relevance. Increasing the endogenous levels of gibberellic acid (GA) by ectopic expression of GA20‐OXIDASE1 (GA20‐OX1), the rate‐limiting step in GA biosynthesis, is known to affect cell division and cell expansion, resulting in larger plants and organs in several plant species. In this study, we examined biomass yield and quality traits of maize plants overexpressing GA20‐OX1 (GA20‐OX1). GA20‐OX1 plants accumulated more vegetative biomass than control plants in greenhouse experiments, but not consistently over two years of field trials. The stems of these plants were longer but also more slender. Investigation of GA20‐OX1 biomass quality using biochemical analyses showed the presence of more cellulose, lignin and cell wall residue. Cell wall analysis as well as expression analysis of lignin biosynthetic genes in developing stems revealed that cellulose and lignin were deposited earlier in development. Pretreatment of GA20‐OX1 biomass with NaOH resulted in a higher saccharification efficiency per unit of dry weight, in agreement with the higher cellulose content. On the other hand, the cellulose‐to‐glucose conversion was slower upon HCl or hot‐water pretreatment, presumably due to the higher lignin content. This study showed that biomass yield and quality traits can be interconnected, which is important for the development of future breeding strategies to improve lignocellulosic feedstock for bioethanol production.     

Gibberellin retards chlorophyll degradation during senescence of Paris polyphylla

Chlorophyll (Chl) degradation was found to be related to the endogenous gibberellin (GA) content in shoots during senescence in the perennial plant Paris polyphylla var. yunnanensis (Franch.). Treatment with gibberellic acid (GA₃) significantly increased the content of endogenous GAs (GA₄ + GA₇), retarded the senescence of shoots, and the degradation of proteins and Chl. Chlorophyllase, Mg-dechelation and peroxidase activities increased more in control plants than in those treated with GA₃. GA₃ treatment also protected lipoxygenase activity, which decreased significantly in control plants.     

Hormonal content and sensitivity of transgenic tobacco and potato plants expressing single rol genes of Agrobacterium rhizogenes T-DNA

The expression of single rol genes of the T_L-DNA of Agrobacterium rhizogenes strain A4 in transgenic tobacco (Nicotiana tabacum L.) and potato (Solanum tuberosum L.) plants alters the internal concentrations of, and the sensitivity to, several plant hormones. The levels of immunoreactive cytokinins, abscisic acid, gibberellins and indole-3-acetic acid were analysed in tissues of the apical shoots, stems, leaves, roots and undifferentiated callus tissue. The addition of the dominant and morphogenetically active rolA, rolB, or rolC genes resulted in alterations in the content of several hormones. rolC overexpression in particular led to an up to fourfold increase in the content of isopentenyladenosine, dihydrozeatin riboside and trans-zeatin riboside-type cytokinins in potato plants. This increase correlated well with different levels of expression of the rolC gene in different transgenic plants. Furthermore it was shown that the dwarfism of P_35s-rolC transgenic tobacco and potato plants is correlated with a 28–60% reduction of gibberellic acid A₁ concentration in apical shoots. Exogenous addition of gibberellic acid completely restored stem elongation in P_35s-_rolC transgenic plants. Apical shoots of dwarf rolA transgenic tobacco plants also contained 22% less gibberellic acid A₁ than control plants, but growth cannot be restored completely by exogenously added gibberellic acid. Similarly, the sensitivity of transgenic tobacco seedlings or callus tissues towards different phytohormone concentrations can be altered by the expression of single rol genes. The overexpression of the rolC gene in seedlings led to an altered response to auxins, cytokinins, abscisic acid, gibberellic acid and the ethylene precursor 1-aminocyclopropane-carboxylic acid. The overexpression of the rolB gene in tobacco calli led to necrosis at lower auxin concentrations than in the wild-type, while other parameters of auxin action, like the induction of cell growth, remained unchanged.     

Characterization of the first dominant dwarf maize mutant carrying a single amino acid insertion in the VHYNP domain of the <Emphasis Type="Italic">dwarf8</Emphasis> gene

The “green revolution” involving mainly wheat and rice was based on the use by breeders of semidominant mutations involved in the signal transduction pathway of Gibberellin (GA). In particular, mutations in the Reduced height (Rht) gene of wheat have been used to reduce plant height and consequently to avoid storm damage and lodging. These genes have been cloned and they encode for DELLA proteins which contain an N-terminal DELLA and a VHYNP domain essential for GA-dependent degradation of these proteins. In maize several mutations have been isolated which affect gibberellin biosynthesis and perception and in particular, mutations in Dwarf8 (D8) gene cause a severe dwarfing phenotype. D8 gene has been identified as an orthologue of Rht (Reduced height), Slr1(Slender rice 1) and Gibberellic Acid Insensitive (GAI) genes, this latter is a negative regulator of GA response in Arabidopsis. In this work, for the first time, we isolated and characterized a single amino acid insertion in the VHYNP domain of D8 maize gene causing the appearance of a dominant dwarf mutation. This spontaneous mutation, named D8-1023, showed a phenotype which is less severe in comparison with the other D8 mutants previously isolated which have modifications in the DELLA domain. This mutant appears to be an useful tool either to study the mechanism of GA-modulated growth in plants or to lower the height of maize tropical germplasm for breeding purposes.     

An endogenous RNA-synthesis-promoting oligopeptide from Cucurbita pepto var. patissonina

The effect of CPPTI (Cucurbita pepo patissonina trypsin inhibitor) on the growth of Cucurbita pepo var patissonina (White bush) was examined. Plants treated with CPPTI grew faster than untreated plants and similarly to those treated with gibberellic acid. Isolated cell nuclei from plants treated with CPPTI showed that of the three DNA-dependent RNA polymerases assayed, RNA polymerase I (EC2.7.7.6) activity was preferentially elevated.Abbreviations CPPTI Cucurbita pepo patissonina trypsin inhibitor - GA₃ gibberellic acid - IAA indole-3-acetic acid - kDa kilodalton - RP I DNA-dependent RNA polymerase I     

Gibberellic acid improves water deficit tolerance in maize plants

The combination effects of water stress and gibberellic acid (GA₃) on physiological attributes and nutritional status of maize (Zea mays L. cv., DK 647 F1) were studied in a pot experiment. Maize plants were grown in the control (well watered WW) and water stress subjected to treated both water stress and two concentrations of gibberellic acid (GA₃ 25 mg L⁻¹, 50 mg L⁻¹). WS was imposed by maintaining the moisture level equivalent to 50 % pot capacity whereas the WW pots were maintained at full pot capacity. Water stress reduced the total dry weight, chlorophyll concentration, and leaf relative water content (RWC), but it increased proline accumulation and electrolyte leakage in maize plants and appears to affect shoots more than roots. Both concentrations of GA₃ (25 and 50 mg L⁻¹) largely enhanced the above physiological parameters to levels similar to control. WS reduced leaf Ca²⁺ and K⁺ concentrations, but exogenous application of GA₃ increased those nutrient levels similar or close to control. Exogenous application of GA₃ improved the water stress tolerance in maize plants by maintaining membrane permeability, enhancing chlorophyll concentration, leaf relative water content (LRWC) and some macro-nutrient concentrations in leaves.     

Interrelations between Gibberellic Acid, Cytokinins and Abscisic Acid in Retarding Leaf Senescence

The interrelation between the effects of abscisic acid (ABA) and the effects of cytokinins and gibberellic acid in retarding leaf senescence was investigated. Leaf discs from plants of Taraxacum megallorrhizon, Rumex pulcber and Tropaeolum majus were floated on solutions of cytokinin or GA to which given amounts of ABA were added. After five days, chlorophyll was extracted and the amount estimated spectrophoto-metrically. The interrelation between the effects of abscisic acid and cytokinins differed from that between the effects of ABA and gibberellic acid. Abscisic acid reduced the senescence retarding effect of GA more than that of cytokinins. A high concentration of cytokinins nullified the senescence enhancing effect of low concentrations of ABA. GA did not reverse the effects of ABA.     

Physiological genetics of the dominant gibberellin-nonresponsive maize dwarfs,Dwarf8 and Dwarf9

Maize (Zea mays L.) Dwarf8-1 (D8-1) is an andromonoecious dwarf mutant proposed to be involved in gibberellin (GA) reception (Fujioka et al. 1988b; Harberd and Freeling 1989). The mutant D8-1 is dominant and GA-nonresponsive (Phinney 1956). We show by map position and similarity of phenotype that five additional dwarf mutants are D8 alleles. We show by map position and similarity of phenotype that a second andromonoecious dwarf mutant, D9-1, defines a duplicate gene. Maize D9-1 and each dominant D8 allele specify a different plant stature, from very mild to very severe dwarfism. Plants of D9-1 and all dominant D8 alleles, except D8-1591, were GA-nonresponsive when treated with 7500 nmol GA₃. The behavior of the mild dwarf D8-1591 was unique in that a small but significant growth response was detected (37% for D8-1591 vs. 130% for the wild type) when treated with 7500 nmol GA₃. These results establish that all dwarf genotypes, except D8-1591, in one dose set a maximum limit on plant growth and block the normal response to GA. When treated with the GA-synthesis inhibitor paclobutrazol, plants of all dwarf genotypes and wild-type siblings were severely dwarfed. Plants of all dwarf genotypes treated with the GA-synthesis inhibitor paclobutrazol and GA₃ were returned to their normal dwarf phenotype. Dominant dwarfing, delayed flowering, increased tillering, and anther development in the ear are characteristic features of D9-1 and all D8 alleles. The GA-synthesis-deficient dwarfs also have these characteristic features. We discuss the function of the wild-type gene product in the context of the observed results.Abbreviations D8 Dwarf8 - D9 Dwarf9 - GA_(n) gibberellin A_(n) - GA₃ gibberellic acid - MNL Maize Genetics Cooperation Newsletter - NIL near-isogenic lines - RFLP restriction fragment length polymorphism - WT wild type This work was supported, in part, by a National Science Foundation Plant Postdoctoral Fellowship to R.G.W., by grants from NIH and ICI Seeds to M.F., the NSF Center for Plant Developmental Biology and the California Agriculture Experiment Station. Much of the work was done in the laboratory of Tim Helentjaris and was supported by a grant from Pioneer Hi-Bred Int'l. The generous gifts of the dominant dwarfing mutants from M.G. Neuffer and O.E. Nelson Jr. are gratefully acknowledged.     

Effect of gibberellic acid treatments, environmental conditions, and genetic background on the expression of theparthenocarpic fruit mutation in tomato

Summary Theparthenocarpic fruit (pat) allele causes a complex syndrome affecting different aspects of tomato reproductive development. This mutation affects stamen (reduced length and carpelloidy), ovule (arrested integument growth and unviability), and ovary (autonomous growth, i.e., parthenocarpy) development;pat mutant plants therefore have reduced male and female fertility. We studied the phenotypic expression patterns of thepat gene after treatments with gibberellic acid (GA₃) and under different growth seasons (late spring and autumn) and genetic backgrounds (backcross [BC] population after interspecific cross). GA₃ treatments were only effective in restoring carpelloid anthers to the wild-type phenotype. Compared to late spring, mutant plants grown in autumn had a lower frequency of carpelloid anthers and aberrant ovules and a higher seed set. Inflorescence position also affected thepat expression; upper inflorescences had low frequency of short anthers and aberrant ovules and an increased tendency to set seeds,pat expressivity was more variable in BC₁ plants segregating after interspecific cross withLycopersicon pennellii than in the originalL. esculentum line. Therefore, a role for minor genes that modify the quantitative expression of thepat mutation is postulated and discussed.Abbreviations ANOVA analysis of variance - BC backcross - GA gibberellic acid     

REGULATORY ROLE OF INDOLE-3-ACETIC ACID AND GIBBERELLIC ACID IN VEGETATIVE DEVELOPMENT OF XANTHIUM PENNSYLVANICUM

A single application of gibberellic acid to young internodes significantly accelerated the rate of internode growth and the rate of leaf production in shoots of Xanthium pennsylvanicum Wallr. The average duration of one plastochron in treated plants was reduced to 43% of control levels. Gibberellic acid also had a pronounced morphogenetic effect on leaves so that the area and leaf length of treated plants were both significantly reduced. Depending upon concentration, auxin had both inhibitory and promotive effects on Xanthium shoots. Indole-3-acetic acid markedly altered the response of the gibberellic acid-treated internodes and those located above and below the site of application. In addition, high auxin concentrations induced the formation of adventitious roots in treated internodes. Auxin also brought about significant reductions in the length and area of leaves developed under the influence of this hormone.     

Gibberellin biosynthesis in fungi: genes, enzymes, evolution, and impact on biotechnology

Gibberellins (GAs) constitute a large family of tetracyclic diterpenoid carboxylic acids, some members of which function as growth hormones in higher plants. As well as being phytohormones, GAs are also present in some fungi and bacteria. In recent years, GA biosynthetic genes from Fusarium fujikuroi and Arabidopsis thaliana have been cloned and well characterised. Although higher plants and the fungus both produce structurally identical GAs, there are important differences indicating that GA biosynthetic pathways have evolved independently in higher plants and fungi. The fact that horizontal gene transfer of GA genes from the plant to the fungus can be excluded, and that GA genes are obviously missing in closely related Fusarium species, raises the question of the origin of fungal GA biosynthetic genes. Besides characterisation of F. fujikuroi GA pathway genes, much progress has been made in the molecular analysis of regulatory mechanisms, especially the nitrogen metabolite repression controlling fungal GA biosynthesis. Basic research in this field has been shown to have an impact on biotechnology. Cloning of genes, construction of knock-out mutants, gene amplification, and regulation studies at the molecular level are powerful tools for improvement of production strains. Besides increased yields of the final product, GA₃, it is now possible to produce intermediates of the GA biosynthetic pathway, such as ent-kaurene, ent-kaurenoic acid, and GA₁₄, in high amounts using different knock-out mutants. This review concentrates mainly on the fungal biosynthetic pathway, the genes and enzymes involved, the regulation network, the biotechnological relevance of recent studies, and on evolutionary aspects of GA biosynthetic genes.     

Mutation induced by ethylmethanesulphonate (EMS), in vitro screening for salt tolerance and plant regeneration of sweet potato (<Emphasis Type="Italic">Ipomoea</Emphasis><Emphasis Type="Italic">batatas</Emphasis> L.)

Salt tolerant cultivars of sweet potato (Ipomoea batatas L.) can be obtained from induced mutation. The objective of the present study was to induce mutation for salt tolerance using ethylmethanesulphonate (EMS) in calli of sweet potato, followed by cell line selection and subsequent plant regeneration. Calli initiated from leaf explants were treated with 0.5% EMS for 0, 1, 1.5, 2, 2.5 and 3 h, followed by rinsing with sterile distilled water for four times. Preliminary experiments showed that 200 mM NaCl could be used as selection pressure. Salt tolerant calli were sub-cultured on medium supplemented with 200 mM NaCl for selection of mutant cell lines and this process repeated 5 times (20 days each). The selected calli were transferred onto somatic embryo formation medium, which was Murashige and Skoog (MS) medium supplemented with 4 mg l⁻¹ abscisic acid (ABA), 10 mg l⁻¹ gibberellic acid (GA). After 15 days, somatic embryos were transferred onto MS medium supplemented with 0.05 mg l⁻¹ ABA, 0.2 mg l⁻¹ zeatin (ZT) for regeneration. Plants designated as ML1, ML2 and ML3 were regenerated from the somatic embryos formed by calli treated with 0.5% EMS for 2 and 2.5 h. After propagation, salt tolerance of these mutants was investigated. Data suggested the mutants were more salt tolerant than control plants.     

The gibberellin biosynthetic genes AtKAO1 and AtKAO2 have overlapping roles throughout Arabidopsis development

Ent‐kaurenoic acid oxidase (KAO), a class of cytochrome P450 monooxygenases of the subfamily CYP88A, catalyzes the conversion of ent‐kaurenoic acid (KA) to gibberellin (GA) GA₁₂, the precursor of all GAs, thereby playing an important role in determining GA concentration in plants. Past work has demonstrated the importance of KAO activity for growth in various plant species. In Arabidopsis, this enzyme is encoded by two genes designated KAO1 and KAO2. In this study, we used various approaches to determine the physiological roles of KAO1 and KAO2 throughout plant development. Analysis of gene expression pattern reveals that both genes are mainly expressed in germinating seeds and young developing organs, thus suggesting functional redundancy. Consistent with this, kao1 and kao2 single mutants are indistinguishable from wild‐type plants. By contrast, the kao1 kao2 double mutant exhibits typical non‐germinating GA‐dwarf phenotypes, similar to those observed in the severely GA‐deficient ga1‐3 mutant. Phenotypic characterization and quantitative analysis of endogenous GA contents of single and double kao mutants further confirm an overlapping role of KAO1 and KAO2 throughout Arabidopsis development.     

Plant regeneration through somatic embryogenesis from spear callus culture of Asparagus cooperi Baker

Somatic embryogenesis and plantlet formation were obtained from callus derived from the subapical region of spears of Asparagus cooperi Baker. Callus was obtained in Murashige and Skoog's medium supplemented with 1-naphthaleneacetic acid and kinetin. Increase in the concentration of potassium nitrate in subsequent subcultures resulted in the formation of embryos. Rapid multiplication of embryos was secured on transfer to a medium containing a different source of nitrogen and a low level (0.01 mg/1) of gibberellic acid. Media containing zeatin or gibberellic acid led to the formation of complete plantlets from embryos. Regenerated plants were cytologically and phenotypically stable.Abbreviations IBA Indole-3-butyric acid - NAA 1-Naphthaleneacetic acid - 2ip (2-Isopentenyl) adenine - BAP 6-Benzylaminopurine, GA,, Gibberellin - ABA Abscisic acid - MS Murashige and Skoog (1962) medium