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Sample records for 1-aminocyclopropane-1-carboxylate acc deaminase

  1. New Insights into 1-Aminocyclopropane-1-Carboxylate (ACC) Deaminase Phylogeny, Evolution and Ecological Significance

    PubMed Central

    Nascimento, Francisco X.; Rossi, Márcio J.; Soares, Cláudio R. F. S.; McConkey, Brendan J.; Glick, Bernard R.

    2014-01-01

    The main objective of this work is the study of the phylogeny, evolution and ecological importance of the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, the activity of which represents one of the most important and studied mechanisms used by plant growth–promoting microorganisms. The ACC deaminase gene and its regulatory elements presence in completely sequenced organisms was verified by multiple searches in diverse databases, and based on the data obtained a comprehensive analysis was conducted. Strain habitat, origin and ACC deaminase activity were taken into account when analyzing the results. In order to unveil ACC deaminase origin, evolution and relationships with other closely related pyridoxal phosphate (PLP) dependent enzymes a phylogenetic analysis was also performed. The data obtained show that ACC deaminase is mostly prevalent in some Bacteria, Fungi and members of Stramenopiles. Contrary to previous reports, we show that ACC deaminase genes are predominantly vertically inherited in various bacterial and fungal classes. Still, results suggest a considerable degree of horizontal gene transfer events, including interkingdom transfer events. A model for ACC deaminase origin and evolution is also proposed. This study also confirms the previous reports suggesting that the Lrp-like regulatory protein AcdR is a common mechanism regulating ACC deaminase expression in Proteobacteria, however, we also show that other regulatory mechanisms may be present in some Proteobacteria and other bacterial phyla. In this study we provide a more complete view of the role for ACC deaminase than was previously available. The results show that ACC deaminase may not only be related to plant growth promotion abilities, but may also play multiple roles in microorganism's developmental processes. Hence, exploring the origin and functioning of this enzyme may be the key in a variety of important agricultural and biotechnological applications. PMID:24905353

  2. Differentiation of 1-aminocyclopropane-1-carboxylate (ACC) deaminase from its homologs is the key for identifying bacteria containing ACC deaminase.

    PubMed

    Li, Zhengyi; Chang, Siping; Ye, Shuting; Chen, Mingyue; Lin, Li; Li, Yuanyuan; Li, Shuying; An, Qianli

    2015-10-01

    1-Aminocyclopropane-1-carboxylate (ACC) deaminase-mediated reduction of ethylene generation in plants under abiotic stresses is a key mechanism by which bacteria can promote plant growth. Misidentification of ACC deaminase and the ACC deaminase structure gene (acdS) can lead to overestimation of the number of bacteria containing ACC deaminase and their function in ecosystems. Previous non-specific amplification of acdS homologs has led to an overestimation of the horizontal transfer of acdS genes. Here, we designed consensus-degenerate hybrid oligonucleotide primers (acdSf3, acdSr3 and acdSr4) based on differentiating the key residues in ACC deaminases from those of homologs for specific amplification of partial acdS genes. PCR amplification, sequencing and phylogenetic analysis identified acdS genes from a wide range of proteobacteria and actinobacteria. PCR amplification and a genomic search did not find the acdS gene in bacteria belonging to Pseudomonas stutzeri or in the genera Enterobacter, Klebsiella or Bacillus. We showed that differentiating the acdS gene and ACC deaminase from their homologs was crucial for the molecular identification of bacteria containing ACC deaminase and for understanding the evolution of the acdS gene. We provide an effective method for screening and identifying bacteria containing ACC deaminase.

  3. 1-Aminocyclopropane-1-carboxylate (ACC) deaminases from Methylobacterium radiotolerans and Methylobacterium nodulans with higher specificity for ACC.

    PubMed

    Fedorov, Dmitry N; Ekimova, Galina A; Doronina, Nina V; Trotsenko, Yuri A

    2013-06-01

    The 1-aminocyclopropane-1-carboxylate (ACC) deaminases (EC 3.4.99.7), the key enzymes of degradation of the precursor of the phytohormone ethylene, have not been well studied despite their great importance for plant-bacterial interactions. Using blast, the open reading frames encoding ACC deaminases were found in the genomes of epiphytic methylotroph Methylobacterium radiotolerans JCM2831 and nodule-forming endosymbiont Methylobacterium nodulans ORS2060. These genes were named acdS and cloned; recombinant proteins were expressed and purified from Escherichia coli. The enzyme from M. nodulans displayed the highest substrate specificity among all of the characterized ACC deaminases (Km 0.80 ± 0.04 mM), whereas the enzyme from M. radiotolerans had Km 1.8 ± 0.3 mM. The kcat values were 111.8 ± 0.2 and 65.8 ± 2.8 min(-1) for the enzymes of M. nodulans and M. radiotolerans, respectively. Both enzymes are homotetramers with a molecular mass of 144 kDa, as was demonstrated by size exclusion chromatography and native PAGE. The purified enzymes displayed the maximum activity at 45-50 °C and pH 8.0. Thus, the priority data have been obtained, extending the knowledge of biochemical properties of bacterial ACC deaminases.

  4. Improved method for effective screening of ACC (1-aminocyclopropane-1-carboxylate) deaminase producing microorganisms.

    PubMed

    Patil, Chandrashekhar; Suryawanshi, Rahul; Koli, Sunil; Patil, Satish

    2016-12-01

    Aminocyclopropane-1-carboxylate deaminase (ACCD) producing microorganisms support plant growth under a variety of biotic and abiotic stress conditions such as drought, soil salinity, flooding, heavy metal pollution and phyto-pathogen attack. Available screening methods for ACCD give idea only about its primary microbial ACCD activity than the actual potential. In the present investigation, we have simply improved screening method by incorporating pH indicator dyes (phenol red and bromothymol blue) in ACC containing medium. This modification is based on the basic principle that ACCD action releases ammonia which can be detected by color change and zone around the bacterial colony. High color intensity and zone around the colony indicates most potent producer, colony showing only a color change indicates moderate potential and no change in colony color indicates least efficiency. Enzymatic bioassays as well as root elongation studies revealed that ACC-deaminase activity of Pseudomonas aeruginosa, Stenotrophomonas maltophilia and Bacillus subtilis clearly corresponds to their growth on dye incorporated ACC medium. This method could be used to complement the existing screening methods and to speed up the targeted isolation of agriculturally important microorganisms.

  5. The cloned 1-aminocyclopropane-1-carboxylate (ACC) deaminase gene from Sinorhizobium sp. strain BL3 in Rhizobium sp. strain TAL1145 promotes nodulation and growth of Leucaena leucocephala.

    PubMed

    Tittabutr, Panlada; Awaya, Jonathan D; Li, Qing X; Borthakur, Dulal

    2008-06-01

    The objective of this study was to determine the role of 1-aminocyclopropane-1-carboxylate (ACC) deaminase of symbionts in nodulation and growth of Leucaena leucocephala. The acdS genes encoding ACC deaminase were cloned from Rhizobium sp. strain TAL1145 and Sinorhizobium sp. BL3 in multicopy plasmids, and transferred to TAL1145. The BL3-acdS gene greatly enhanced ACC deaminase activity in TAL1145 compared to the native acdS gene. The transconjugants of TAL1145 containing the native or BL3 acdS gene could grow in minimal media containing 1.5mM ACC, whereas BL3 could tolerate up to 3mM ACC. The TAL1145 acdS gene was inducible by mimosine and not by ACC, while the BL3 acdS gene was highly inducible by ACC and not by mimosine. The transconjugants of TAL1145 containing the native- and BL3-acdS genes formed nodules with greater number and sizes, and produced higher root mass on L. leucocephala than by TAL1145. This study shows that the introduction of multiple copies of the acdS gene increased ACC deaminase activities of TAL1145 and enhanced its symbiotic efficiency on L. leucocephala.

  6. 1-Aminocyclopropane-1-carboxylic acid (ACC) deaminase-containing rhizobacteria protect Ocimum sanctum plants during waterlogging stress via reduced ethylene generation.

    PubMed

    Barnawal, Deepti; Bharti, Nidhi; Maji, Deepamala; Chanotiya, Chandan Singh; Kalra, Alok

    2012-09-01

    Ocimum sanctum grown as rain-fed crop, is known to be poorly adapted to waterlogged conditions. Many a times the crop suffers extreme damages because of anoxia and excessive ethylene generation due to waterlogging conditions present under heavy rain. The usefulness of 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase-containing plant growth promoting rhizobacteria was investigated under waterlogging stress. The comparison of herb yield and stress induced biochemical changes of waterlogged and non-waterlogged plants with and without ACC deaminase-containing microbiological treatments were monitored in this study. Ten plant growth promoting rhizobacteria strains containing ACC-deaminase were isolated and characterized. Four selected isolates Fd2 (Achromobacter xylosoxidans), Bac5 (Serratia ureilytica), Oci9 (Herbaspirillum seropedicae) and Oci13 (Ochrobactrum rhizosphaerae) had the potential to protect Ocimum plants from flood induced damage under waterlogged glass house conditions. Pot experiments were conducted to evaluate the potential of these ACC deaminase-containing selected strains for reducing the yield losses caused by waterlogging conditions. Bacterial treatments protected plants from waterlogging induced detrimental changes like stress ethylene production, reduced chlorophyll concentration, higher lipid peroxidation, proline concentration and reduced foliar nutrient uptake. Fd2 (A. xylosoxidans) induced maximum waterlogging tolerance as treated waterlogged plants recorded maximum growth and herb yield (46.5% higher than uninoculated waterlogged plants) with minimum stress ethylene levels (53% lower ACC concentration as compared to waterlogged plants without bacterial inoculation) whereas under normal non-waterlogged conditions O. rhizosphaerae was most effective in plant growth promotion.

  7. Southern blight disease of tomato control by 1-aminocyclopropane-1-carboxylate (ACC) deaminase producing Paenibacillus lentimorbus B-30488.

    PubMed

    Dixit, Ritu; Agrawal, Lalit; Gupta, Swati; Kumar, Manoj; Yadav, Sumit; Chauhan, Puneet Singh; Nautiyal, Chandra Shekhar

    2016-01-01

    Tomato cultivation is highly susceptible for soil born diseases and among them southern blight disease caused by Scelerotium rolfsii is very common. For its management use of chemical fungicides is not very successful as their spores are able to survive for many years in the soil. As an alternative eco-friendly approach to control the disease antagonistic microbes are being characterized.Among them plant growth promoting rhizobacteria Paenibacillus lentimorbus B-30488 (B-30488) with antagonistic properties, multiple PGP attributes stress tolerance and ACC deaminase enzyme activity is characterized to decipher its mode of action against S. rolfsii under in vitro and in vivo conditions. In vitro results obtained from this study clearly demonstrate that B-30488 has ability to show antagonistic properties under different abiotic stresses against S. rolfsii. Similar results were also obtained from in vivo experiments where B-30488 inoculation has efficiently controlled the disease caused by S. rolfsii and improve the plant growth. Deleterious enhanced ethylene level in S. rolfsii infected plants was also ameliorated by inoculation of ACC deaminase producing B-30488. The ACC accumulation, ACO and ACS activities were also modulated in S. rolfsii infected plants. Results from defense enzymes and other biochemical attributes were also support the role of B-30488 inoculation in ameliorating the biotic stress caused by S. rolfsii in tomato plants. These results were further validated by pathogen related gene expression analysis by real time PCR. Overall results from the present study may be concluded that ACC deaminase producing B-30488 has ability to control the southern blight disease caused by S. rolfsii and commercial bioinoculant package may be developed.

  8. Southern blight disease of tomato control by 1-aminocyclopropane-1-carboxylate (ACC) deaminase producing Paenibacillus lentimorbus B-30488

    PubMed Central

    Dixit, Ritu; Agrawal, Lalit; Gupta, Swati; Kumar, Manoj; Yadav, Sumit; Chauhan, Puneet Singh; Nautiyal, Chandra Shekhar

    2016-01-01

    abstract Tomato cultivation is highly susceptible for soil born diseases and among them southern blight disease caused by Scelerotium rolfsii is very common. For its management use of chemical fungicides is not very successful as their spores are able to survive for many years in the soil. As an alternative eco-friendly approach to control the disease antagonistic microbes are being characterized.Among them plant growth promoting rhizobacteria Paenibacillus lentimorbus B-30488 (B-30488) with antagonistic properties, multiple PGP attributes stress tolerance and ACC deaminase enzyme activity is characterized to decipher its mode of action against S. rolfsii under in vitro and in vivo conditions. In vitro results obtained from this study clearly demonstrate that B-30488 has ability to show antagonistic properties under different abiotic stresses against S. rolfsii. Similar results were also obtained from in vivo experiments where B-30488 inoculation has efficiently controlled the disease caused by S. rolfsii and improve the plant growth. Deleterious enhanced ethylene level in S. rolfsii infected plants was also ameliorated by inoculation of ACC deaminase producing B-30488. The ACC accumulation, ACO and ACS activities were also modulated in S. rolfsii infected plants. Results from defense enzymes and other biochemical attributes were also support the role of B-30488 inoculation in ameliorating the biotic stress caused by S. rolfsii in tomato plants. These results were further validated by pathogen related gene expression analysis by real time PCR. Overall results from the present study may be concluded that ACC deaminase producing B-30488 has ability to control the southern blight disease caused by S. rolfsii and commercial bioinoculant package may be developed. PMID:26825539

  9. Possible Role of 1-Aminocyclopropane-1-Carboxylate (ACC) Deaminase Activity of Sinorhizobium sp. BL3 on Symbiosis with Mung Bean and Determinate Nodule Senescence

    PubMed Central

    Tittabutr, Panlada; Sripakdi, Sudarat; Boonkerd, Nantakorn; Tanthanuch, Waraporn; Minamisawa, Kiwamu; Teaumroong, Neung

    2015-01-01

    Sinorhizobium sp. BL3 forms symbiotic interactions with mung bean (Vigna radiata) and contains lrpL-acdS genes, which encode the 1-aminocyclopropane-1-carboxylate (ACC) deaminase enzyme that cleaves ACC, a precursor of plant ethylene synthesis. Since ethylene interferes with nodule formation in some legumes and plays a role in senescence in plant cells, BL3-enhancing ACC deaminase activity (BL3+) and defective mutant (BL3−) strains were constructed in order to investigate the effects of this enzyme on symbiosis and nodule senescence. Nodulation competitiveness was weaker in BL3− than in the wild-type, but was stronger in BL3+. The inoculation of BL3− into mung bean resulted in less plant growth, a lower nodule dry weight, and smaller nodule number than those in the wild-type, whereas the inoculation of BL3+ had no marked effects. However, similar nitrogenase activity was observed with all treatments; it was strongly detected 3 weeks after the inoculation and gradually declined with time, indicating senescence. The rate of plant nodulation by BL3+ increased in a time-dependent manner. Nodules occupied by BL3− formed smaller symbiosomes, and bacteroid degradation was more prominent than that in the wild-type 7 weeks after the inoculation. Changes in biochemical molecules during nodulation were tracked by Fourier Transform Infrared (FT-IR) microspectroscopy, and the results obtained confirmed that aging processes differed in nodules occupied by BL3 and BL3−. This is the first study to show the possible role of ACC deaminase activity in senescence in determinate nodules. Our results suggest that an increase in ACC deaminase activity in this strain does not extend the lifespan of nodules, whereas the lack of this activity may accelerate nodule senescence. PMID:26657304

  10. 1-aminocyclopropane-1-carboxylic acid (ACC) in plants: more than just the precursor of ethylene!

    PubMed Central

    Van de Poel, Bram; Van Der Straeten, Dominique

    2014-01-01

    Ethylene is a simple two carbon atom molecule with profound effects on plants. There are quite a few review papers covering all aspects of ethylene biology in plants, including its biosynthesis, signaling and physiology. This is merely a logical consequence of the fascinating and pleiotropic nature of this gaseous plant hormone. Its biochemical precursor, 1-aminocyclopropane-1-carboxylic acid (ACC) is also a fairly simple molecule, but perhaps its role in plant biology is seriously underestimated. This triangularly shaped amino acid has many more features than just being the precursor of the lead-role player ethylene. For example, ACC can be conjugated to three different derivatives, but their biological role remains vague. ACC can also be metabolized by bacteria using ACC-deaminase, favoring plant growth and lowering stress susceptibility. ACC is also subjected to a sophisticated transport mechanism to ensure local and long-distance ethylene responses. Last but not least, there are now a few exciting studies where ACC has been reported to function as a signal itself, independently from ethylene. This review puts ACC in the spotlight, not to give it the lead-role, but to create a picture of the stunning co-production of the hormone and its precursor. PMID:25426135

  11. Studies of 1-Amino-2,2-difluorocyclopropane-1-carboxylic Acid: Mechanism of Decomposition and Inhibition of 1-Aminocyclopropane-1-carboxylic Acid Deaminase.

    PubMed

    Liu, Cheng-Hao; Wang, Shao-An; Ruszczycky, Mark W; Chen, Huawei; Li, Keqiang; Murakami, Kazuo; Liu, Hung-wen

    2015-07-02

    1-Amino-2,2-difluorocyclopropane-1-carboxylic acid (DFACC) is of interest in the study of 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase due to the increased reactivity of its cyclopropyl functionality. It is shown that DFACC is unstable under near-physiological conditions where it primarily decomposes via specific-base catalysis to 3-fluoro-2-oxobut-3-enoic acid with a rate constant of 0.18 ± 0.01 min(-1). Upon incubation with ACC deaminase, DFACC is found to be a slow-dissociating inhibitor of ACC deaminase with submicromolar affinity.

  12. Increased 1-aminocyclopropane-1-carboxylate deaminase activity enhances Agrobacterium tumefaciens-mediated gene delivery into plant cells.

    PubMed

    Someya, Tatsuhiko; Nonaka, Satoko; Nakamura, Kouji; Ezura, Hiroshi

    2013-10-01

    Agrobacterium-mediated transformation is a useful tool for the genetic modification in plants, although its efficiency is low for several plant species. Agrobacterium-mediated transformation has three major steps in laboratory-controlled experiments: the delivery of T-DNA into plant cells, the selection of transformed plant cells, and the regeneration of whole plants from the selected cells. Each of these steps must be optimized to improve the efficiency of Agrobacterium-mediated plant transformation. It has been reported that increasing the number of cells transformed by T-DNA delivery can improve the frequency of stable transformation. Previously, we demonstrated that a reduction in ethylene production by plant cells during cocultivation with A. tumefaciens-expressing 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase resulted in increased T-DNA delivery into the plant cells. In this study, to further improve T-DNA delivery by A. tumefaciens, we modified the expression cassette of the ACC deaminase gene using vir gene promoter sequences. The ACC deaminase gene driven by the virD1 promoter was expressed at a higher level, resulting in a higher ACC deaminase activity in this A. tumefaciens strain than in the strain with the lac promoter used in a previous study. The newly developed A. tumefaciens strain improves the delivery of T-DNA into Solanum lycopersicum (tomato) and Erianthus ravennae plants and thus may be a powerful tool for the Agrobacterium-mediated genetic engineering of plants.

  13. Characterization of plant growth promoting rhizobacteria isolated from polluted soils and containing 1-aminocyclopropane-1-carboxylate deaminase.

    PubMed

    Belimov, A A; Safronova, V I; Sergeyeva, T A; Egorova, T N; Matveyeva, V A; Tsyganov, V E; Borisov, A Y; Tikhonovich, I A; Kluge, C; Preisfeld, A; Dietz, K J; Stepanok, V V

    2001-07-01

    Fifteen bacterial strains containing 1-aminocyclopropane-1-carboxylate (ACC) deaminase were isolated from the rhizoplane of pea (Pisum sativum L.) and Indian mustard (Brassica juncea L.) grown in different soils and a long-standing sewage sludge contaminated with heavy metals. The isolated strains were characterized and assigned to various genera and species, such as Pseudomonas brassicacearum, Pseudomonas marginalis, Pseudomonas oryzihabitans, Pseudomonas putida, Pseudomonas sp., Alcaligenes xylosoxidans, Alcaligenes sp., Variovorax paradoxus, Bacillus pumilus, and Rhodococcus sp. by determination of 16S rRNA gene sequences. The root elongation of Indian mustard and rape (Brassica napus var. oleifera L.) germinating seedlings was stimulated by inoculation with 8 and 13 isolated strains, respectively. The bacteria were tolerant to cadmium toxicity and stimulated root elongation of rape seedlings in the presence of 300 microM CdCl2 in the nutrient solution. The effect of ACC-utilising bacteria on root elongation correlated with the impact of aminoethoxyvinylglycine and silver ions, chemical inhibitors of ethylene biosynthesis. A significant improvement in the growth of rape caused by inoculation with certain selected strains was also observed in pot experiments, when the plants were cultivated in cadmium-supplemented soil. The biomass of pea cv. Sparkle and its ethylene sensitive mutant E2 (sym5), in particular, was increased through inoculation with certain strains of ACC-utilising bacteria in pot experiments in quartz sand culture. The beneficial effect of the bacteria on plant growth varied significantly depending on individual bacterial strains, plant genotype, and growth conditions. The results suggest that plant growth promoting rhizobacteria containing ACC deaminase are present in various soils and offer promise as a bacterial inoculum for improvement of plant growth, particularly under unfavourable environmental conditions.

  14. Mechanistic studies of 1-aminocyclopropane-1-carboxylate deaminase: characterization of an unusual pyridoxal 5'-phosphate-dependent reaction.

    PubMed

    Thibodeaux, Christopher J; Liu, Hung-Wen

    2011-03-22

    1-Aminocyclopropane-1-carboxylic acid (ACC) deaminase (ACCD) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that cleaves the cyclopropane ring of ACC, to give α-ketobutyric acid and ammonia as products. The cleavage of the C(α)-C(β) bond of an amino acid substrate is a rare event in PLP-dependent enzyme catalysis. Potential chemical mechanisms involving nucleophile- or acid-catalyzed cyclopropane ring opening have been proposed for the unusual transformation catalyzed by ACCD, but the actual mode of cyclopropane ring cleavage remains obscure. In this report, we aim to elucidate the mechanistic features of ACCD catalysis by investigating the kinetic properties of ACCD from Pseudomonas sp. ACP and several of its mutant enzymes. Our studies suggest that the pK(a) of the conserved active site residue, Tyr294, is lowered by a hydrogen bonding interaction with a second conserved residue, Tyr268. This allows Tyr294 to deprotonate the incoming amino group of ACC to initiate the aldimine exchange reaction between ACC and the PLP coenzyme and also likely helps to activate Tyr294 for a role as a nucleophile to attack and cleave the cyclopropane ring of the substrate. In addition, solvent kinetic isotope effect (KIE), proton inventory, and (13)C KIE studies of the wild type enzyme suggest that the C(α)-C(β) bond cleavage step in the chemical mechanism is at least partially rate-limiting under k(cat)/K(m) conditions and is likely preceded in the mechanism by a partially rate-limiting step involving the conversion of a stable gem-diamine intermediate into a reactive external aldimine intermediate that is poised for cyclopropane ring cleavage. When viewed within the context of previous mechanistic and structural studies of ACCD enzymes, our studies are most consistent with a mode of cyclopropane ring cleavage involving nucleophilic catalysis by Tyr294.

  15. 1-Aminocyclopropane-1-Carboxylate Deaminase from Pseudomonas stutzeri A1501 Facilitates the Growth of Rice in the Presence of Salt or Heavy Metals.

    PubMed

    Han, Yunlei; Wang, Rui; Yang, Zhirong; Zhan, Yuhua; Ma, Yao; Ping, Shuzhen; Zhang, Liwen; Lin, Min; Yan, Yongliang

    2015-07-01

    1-Aminocyclopropane-1-carboxylate (ACC) deaminase, which is encoded by some bacteria, can reduce the amount of ethylene, a root elongation inhibitor, and stimulate the growth of plants under various environmental stresses. The presence of ACC deaminase activity and the regulation of ACC in several rhizospheric bacteria have been reported. The nitrogen-fixing Pseudomonas stutzeri A1501 is capable of endophytic association with rice plants and promotes the growth of rice. However, the functional identification of ACC deaminase has not been performed. In this study, the proposed effect of ACC deaminase in P. stutzeri A1501 was investigated. Genome mining showed that P. stutzeri A1501 carries a single gene encoding ACC deaminase, designated acdS. The acdS mutant was devoid of ACC deaminase activity and was less resistant to NaCl and NiCl2 compared with the wild-type. Furthermore, inactivation of acdS greatly impaired its nitrogenase activity under salt stress conditions. It was also observed that mutation of the acdS gene led to loss of the ability to promote the growth of rice under salt or heavy metal stress. Taken together, this study illustrates the essential role of ACC deaminase, not only in enhancing the salt or heavy metal tolerance of bacteria but also in improving the growth of plants, and provides a theoretical basis for studying the interaction between plant growth-promoting rhizobacteria and plants.

  16. Accumulation and Transport of 1-Aminocyclopropane-1-Carboxylic Acid (ACC) in Plants: Current Status, Considerations for Future Research and Agronomic Applications

    PubMed Central

    Vanderstraeten, Lisa; Van Der Straeten, Dominique

    2017-01-01

    1-aminocyclopropane-1-carboxylic acid (ACC) is a non-protein amino acid acting as the direct precursor of ethylene, a plant hormone regulating a wide variety of vegetative and developmental processes. ACC is the central molecule of ethylene biosynthesis. The rate of ACC formation differs in response to developmental, hormonal and environmental cues. ACC can be conjugated to three derivatives, metabolized in planta or by rhizobacteria using ACC deaminase, and is transported throughout the plant over short and long distances, remotely leading to ethylene responses. This review highlights some recent advances related to ACC. These include the regulation of ACC synthesis, conjugation and deamination, evidence for a role of ACC as an ethylene-independent signal, short and long range ACC transport, and the identification of a first ACC transporter. Although unraveling the complex mechanism of ACC transport is in its infancy, new questions emerge together with the identification of a first transporter. In the light of the future quest for additional ACC transporters, this review presents perspectives of the novel findings and includes considerations for future research toward applications in agronomy. PMID:28174583

  17. Rhizosphere bacteria containing 1-aminocyclopropane-1-carboxylate deaminase increase yield of plants grown in drying soil via both local and systemic hormone signalling.

    PubMed

    Belimov, Andrey A; Dodd, Ian C; Hontzeas, Nikos; Theobald, Julian C; Safronova, Vera I; Davies, William J

    2009-01-01

    Decreased soil water availability can stimulate production of the plant hormone ethylene and inhibit plant growth. Strategies aimed at decreasing stress ethylene evolution might attenuate its negative effects. An environmentally benign (nonchemical) method of modifying crop ethylene relations - soil inoculation with a natural root-associated bacterium Variovorax paradoxus 5C-2 (containing the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase that degrades the ethylene precursor ACC), was assessed with pea (Pisum sativum) plants grown in drying soil. Inoculation with V. paradoxus 5C-2, but not with a transposome mutant with massively decreased ACC deaminase activity, improved growth, yield and water-use efficiency of droughted peas. Systemic effects of V. paradoxus 5C-2 included an amplified soil drying-induced increase of xylem abscisic acid (ABA) concentration, but an attenuated soil drying-induced increase of xylem ACC concentration. A local bacterial effect was increased nodulation by symbiotic nitrogen-fixing bacteria, which prevented a drought-induced decrease in nodulation and seed nitrogen content. Successfully deploying a single bacterial gene in the rhizosphere increased yield and nutritive value of plants grown in drying soil, via both local and systemic hormone signalling. Such bacteria may provide an easily realized, economic means of sustaining crop yields and using irrigation water more efficiently in dryland agriculture.

  18. Novel Rhizosphere Soil Alleles for the Enzyme 1-Aminocyclopropane-1-Carboxylate Deaminase Queried for Function with an In Vivo Competition Assay.

    PubMed

    Jin, Zhao; Di Rienzi, Sara C; Janzon, Anders; Werner, Jeff J; Angenent, Largus T; Dangl, Jeffrey L; Fowler, Douglas M; Ley, Ruth E

    2015-12-04

    Metagenomes derived from environmental microbiota encode a vast diversity of protein homologs. How this diversity impacts protein function can be explored through selection assays aimed to optimize function. While artificially generated gene sequence pools are typically used in selection assays, their usage may be limited because of technical or ethical reasons. Here, we investigate an alternative strategy, the use of soil microbial DNA as a starting point. We demonstrate this approach by optimizing the function of a widely occurring soil bacterial enzyme, 1-aminocyclopropane-1-carboxylate (ACC) deaminase. We identified a specific ACC deaminase domain region (ACCD-DR) that, when PCR amplified from the soil, produced a variant pool that we could swap into functional plasmids carrying ACC deaminase-encoding genes. Functional clones of ACC deaminase were selected for in a competition assay based on their capacity to provide nitrogen to Escherichia coli in vitro. The most successful ACCD-DR variants were identified after multiple rounds of selection by sequence analysis. We observed that previously identified essential active-site residues were fixed in the original unselected library and that additional residues went to fixation after selection. We identified a divergent essential residue whose presence hints at the possible use of alternative substrates and a cluster of neutral residues that did not influence ACCD performance. Using an artificial ACCD-DR variant library generated by DNA oligomer synthesis, we validated the same fixation patterns. Our study demonstrates that soil metagenomes are useful starting pools of protein-coding-gene diversity that can be utilized for protein optimization and functional characterization when synthetic libraries are not appropriate.

  19. Novel Rhizosphere Soil Alleles for the Enzyme 1-Aminocyclopropane-1-Carboxylate Deaminase Queried for Function with an In Vivo Competition Assay

    PubMed Central

    Jin, Zhao; Di Rienzi, Sara C.; Janzon, Anders; Werner, Jeff J.; Angenent, Largus T.; Dangl, Jeffrey L.; Fowler, Douglas M.

    2015-01-01

    Metagenomes derived from environmental microbiota encode a vast diversity of protein homologs. How this diversity impacts protein function can be explored through selection assays aimed to optimize function. While artificially generated gene sequence pools are typically used in selection assays, their usage may be limited because of technical or ethical reasons. Here, we investigate an alternative strategy, the use of soil microbial DNA as a starting point. We demonstrate this approach by optimizing the function of a widely occurring soil bacterial enzyme, 1-aminocyclopropane-1-carboxylate (ACC) deaminase. We identified a specific ACC deaminase domain region (ACCD-DR) that, when PCR amplified from the soil, produced a variant pool that we could swap into functional plasmids carrying ACC deaminase-encoding genes. Functional clones of ACC deaminase were selected for in a competition assay based on their capacity to provide nitrogen to Escherichia coli in vitro. The most successful ACCD-DR variants were identified after multiple rounds of selection by sequence analysis. We observed that previously identified essential active-site residues were fixed in the original unselected library and that additional residues went to fixation after selection. We identified a divergent essential residue whose presence hints at the possible use of alternative substrates and a cluster of neutral residues that did not influence ACCD performance. Using an artificial ACCD-DR variant library generated by DNA oligomer synthesis, we validated the same fixation patterns. Our study demonstrates that soil metagenomes are useful starting pools of protein-coding-gene diversity that can be utilized for protein optimization and functional characterization when synthetic libraries are not appropriate. PMID:26637602

  20. Inducing salt tolerance in mung bean through coinoculation with rhizobia and plant-growth-promoting rhizobacteria containing 1-aminocyclopropane-1-carboxylate deaminase.

    PubMed

    Ahmad, Maqshoof; Zahir, Zahir A; Asghar, H Naeem; Asghar, M

    2011-07-01

    Twenty-five strains of plant-growth-promoting rhizobacteria (PGPR) containing 1-aminocyclopropane-1-carboxylate (ACC) deaminase and 10 strains of rhizobia were isolated from rhizosphere soil samples and nodules of mung bean. They were screened in separate trials under salt-stressed axenic conditions. The three most effective strains of PGPR (Mk1, Pseudomonas syringae ; Mk20, Pseudomonas fluorescens ; and Mk25, Pseudomonas fluorescens biotype G) and Rhizobium phaseoli strains M1, M6, and M9 were evaluated in coinoculation for their growth-promoting activity at three salinity levels (original, 4 dS·m(-1), and 6 dS·m(-1)) under axenic conditions. The results showed that salinity stress significantly reduced plant growth but inoculation with PGPR containing ACC deaminase and rhizobia enhanced plant growth, thus reducing the inhibitory effect of salinity. However, their combined application was more effective under saline conditions, and the combination Mk20 × M6 was the most efficient for improving seedling growth and nodulation. The effect of high ethylene concentrations on plant growth and the performance of these strains for reducing the negative impact of saline stress was also evaluated by conducting a classical triple-response bioassay. The intensity of the classical triple response decreased owing to inoculation with these strains, with the root and shoot lengths of inoculated mung bean seedlings increasing and stem diameter decreasing, which is a typical response to the dilution in a classical triple response bioassay. Thus, coinoculation with PGPR containing ACC deaminase and Rhizobium spp. could be a useful approach for inducing salt tolerance and thus improving growth and nodulation in mung bean under salt-affected conditions.

  1. Variovorax guangxiensis sp. nov., an aerobic, 1-aminocyclopropane-1-carboxylate deaminase producing bacterium isolated from banana rhizosphere.

    PubMed

    Gao, Jun-lian; Yuan, Mei; Wang, Xu-ming; Qiu, Tian-lei; Li, Ji-wei; Liu, Hong-can; Li, Xiu-ai; Chen, Jian; Sun, Jian-guang

    2015-01-01

    A 1-aminocyclopropane-1-carboxylate deaminase producing bacterium, designated GXGD002(T), was isolated from the rhizosphere of banana plants cultivated in Guangxi province, China. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain GXGD002(T) is a member of the genus Variovorax. High levels of 16S rRNA gene sequence similarity are found between strain GXGD002(T) and Variovorax paradoxus DSM 30034(T) (99.4 %), Variovorax ginsengisoli KCTC 12583(T) (99.1 %), Variovorax boronicumulans KCTC 22010(T) (99.0 %), Variovorax soli DSM18216(T) (98.7 %), Variovorax defluvii DSM 27259(T) (98.1 %) and Variovorax dokdonensis KCTC 12544(T) (97.4 %) respectively. However, the DNA-DNA hybridization values between strain GXGD002(T) and its closely related species V. paradoxus DSM 30034(T), V. ginsengisoli KCTC 12583(T) and V. boronicumulans KCTC 22010(T) were found to be 40.7, 30.9 and 23.7 %, respectively. The DNA G + C content of strain GXGD002(T) was found to be 67.8 mol%. The major fatty acids of strain GXGD002(T) are C16:0 (20.3 %), C10:0 3OH (18.4 %), C17:0 cyclo (18.9 %), C18:1w7c (12.3 %) and summed feature 3 (13.9 %). The predominant respiratory quinone was identified as ubiquinone-8 (Q-8) and the major polar lipids as phosphatidylethanolamine and phosphatidylglycerol. The results of polyphasic taxonomic study including physiological and biochemical tests, whole-cell SDS-PAGE profiles and chemotaxonomic analysis allowed a clear differentiation of strain GXGD002(T) from the other species in the genus Variovorax. Based on these results, a new species, Variovorax guangxiensis, is proposed. The type strain is GXGD002(T) (=DSM 27352(T) = ACCC 05911(T)).

  2. Expression of an exogenous 1-aminocyclopropane-1-carboxylate deaminase gene in psychrotolerant bacteria modulates ethylene metabolism and cold induced genes in tomato under chilling stress.

    PubMed

    Subramanian, Parthiban; Krishnamoorthy, Ramasamy; Chanratana, Mak; Kim, Kiyoon; Sa, Tongmin

    2015-04-01

    The role of stress induced ethylene under low temperature stress has been controversial and hitherto remains unclear. In the present study, 1-aminocyclopropane-1-carboxylate deaminase (ACCD) gene, acdS expressing mutant strains were generated from ACCD negative psychrotolerant bacterial strains Flavobacterium sp. OR306 and Pseudomonas frederiksbergensis OS211, isolated from agricultural soil during late winter. After transformation with plasmid pRKACC which contained the acdS gene, both the strains were able to exhibit ACCD activity in vitro. The effect of this ACCD under chilling stress with regards to ethylene was studied in tomato plants inoculated with both acdS expressing and wild type bacteria. On exposing the plants to one week of chilling treatment at 12/10 °C, it was found that stress ethylene, ACC accumulation and ACO activity which are markers of ethylene stress, were significantly reduced in plants inoculated with the acdS gene transformed mutants. In case of plants inoculated with strain OS211-acdS, ethylene emission, ACC accumulation and ACO activity was significantly reduced by 52%, 75.9% and 23.2% respectively compared to uninoculated control plants. Moreover, expression of cold induced LeCBF1 and LeCBF3 genes showed that these genes were significantly induced by the acdS transformed mutants in addition to reduced expression of ethylene-responsive transcription factor 13 (ETF-13) and ACO genes. Induced expression of LeCBF1 and LeCBF3 in plants inoculated with acdS expressing mutants compared to wild type strains show that physiologically evolved stress ethylene and its transcription factors play a role in regulation of cold induced genes as reported earlier in the literature.

  3. 1-Aminocyclopropane-1-carboxylic acid (ACC) concentration and ACC synthase expression in soybean roots, root tips, and soybean cyst nematode (Heterodera glycines)-infected roots.

    PubMed

    Tucker, Mark L; Xue, Ping; Yang, Ronghui

    2010-01-01

    Colonization of plant roots by root knot and cyst nematodes requires a functional ethylene response pathway. However, ethylene plays many roles in root development and whether its role in nematode colonization is direct or indirect, for example lateral root initiation or root hair growth, is not known. The temporal requirement for ethylene and localized synthesis of ethylene during the life span of soybean cyst nematode (SCN) on soybean roots was further investigated. Although a significant increase in ethylene evolution was not detected from SCN-colonized roots, the concentration of the immediate precursor to ethylene, 1-aminocyclopropane-1-carboxylic acid (ACC), was higher in SCN-colonized root pieces and root tips than in other parts of the root. Moreover, expression analysis of 17 ACC synthase (ACS) genes indicated that a select set of ACS genes is expressed in SCN-colonized root pieces that is clearly different from the set of genes expressed in non-colonized roots or root tips. Semi-quantitative real-time PCR indicated that ACS transcript accumulation correlates with the high concentration of ACC in root tips. In addition, an ACS-like sequence was found in the public SCN nucleotide database. Acquisition of a full-length sequence for this mRNA (accession GQ389647) and alignment with transcripts for other well-characterized ACS proteins indicated that the nematode sequence is missing a key element required for ACS activity and therefore probably is not a functional ACS. Moreover, no significant amount of ACC was found in any growth stage of SCN that was tested.

  4. Inoculation of Soil with Plant Growth Promoting Bacteria Producing 1-Aminocyclopropane-1-Carboxylate Deaminase or Expression of the Corresponding acdS Gene in Transgenic Plants Increases Salinity Tolerance in Camelina sativa

    PubMed Central

    Heydarian, Zohreh; Yu, Min; Gruber, Margaret; Glick, Bernard R.; Zhou, Rong; Hegedus, Dwayne D.

    2016-01-01

    Camelina sativa (camelina) is an oilseed crop touted for use on marginal lands; however, it is no more tolerant of soil salinity than traditional crops, such as canola. Plant growth-promoting bacteria (PGPB) that produce 1-aminocyclopropane-1-carboxylate deaminase (ACC deaminase) facilitate plant growth in the presence of abiotic stresses by reducing stress ethylene. Rhizospheric and endophytic PGPB and the corresponding acdS- mutants of the latter were examined for their ability to enhance tolerance to salt in camelina. Stimulation of growth and tolerance to salt was correlated with ACC deaminase production. Inoculation of soil with wild-type PGPB led to increased shoot length in the absence of salt, and increased seed production by approximately 30–50% under moderately saline conditions. The effect of ACC deaminase was further examined in transgenic camelina expressing a bacterial gene encoding ACC deaminase (acdS) under the regulation of the CaMV 35S promoter or the root-specific rolD promoter. Lines expressing acdS, in particular those using the rolD promoter, showed less decline in root length and weight, increased seed production, better seed quality and higher levels of seed oil production under salt stress. This study clearly demonstrates the potential benefit of using either PGPB that produce ACC deaminase or transgenic plants expressing the acdS gene under the control of a root-specific promoter to facilitate plant growth, seed production and seed quality on land that is not normally suitable for the majority of crops due to high salt content. PMID:28018305

  5. Inoculation of Soil with Plant Growth Promoting Bacteria Producing 1-Aminocyclopropane-1-Carboxylate Deaminase or Expression of the Corresponding acdS Gene in Transgenic Plants Increases Salinity Tolerance in Camelina sativa.

    PubMed

    Heydarian, Zohreh; Yu, Min; Gruber, Margaret; Glick, Bernard R; Zhou, Rong; Hegedus, Dwayne D

    2016-01-01

    Camelina sativa (camelina) is an oilseed crop touted for use on marginal lands; however, it is no more tolerant of soil salinity than traditional crops, such as canola. Plant growth-promoting bacteria (PGPB) that produce 1-aminocyclopropane-1-carboxylate deaminase (ACC deaminase) facilitate plant growth in the presence of abiotic stresses by reducing stress ethylene. Rhizospheric and endophytic PGPB and the corresponding acdS- mutants of the latter were examined for their ability to enhance tolerance to salt in camelina. Stimulation of growth and tolerance to salt was correlated with ACC deaminase production. Inoculation of soil with wild-type PGPB led to increased shoot length in the absence of salt, and increased seed production by approximately 30-50% under moderately saline conditions. The effect of ACC deaminase was further examined in transgenic camelina expressing a bacterial gene encoding ACC deaminase (acdS) under the regulation of the CaMV 35S promoter or the root-specific rolD promoter. Lines expressing acdS, in particular those using the rolD promoter, showed less decline in root length and weight, increased seed production, better seed quality and higher levels of seed oil production under salt stress. This study clearly demonstrates the potential benefit of using either PGPB that produce ACC deaminase or transgenic plants expressing the acdS gene under the control of a root-specific promoter to facilitate plant growth, seed production and seed quality on land that is not normally suitable for the majority of crops due to high salt content.

  6. Rapid identification of 1-aminocyclopropane-1-carboxylate (ACC) synthase genotypes in cultivars of Japanese pear (Pyrus pyrifolia Nakai) using CAPS markers.

    PubMed

    Itai, A; Kotaki, T; Tanabe, K; Tamura, F; Kawaguchi, D; Fukuda, M

    2003-05-01

    In Japanese pear (Pyrus pyrifolia Nakai), fruit storage potential is closely related to the amount of ethylene produced. We have developed a rapid and accurate method for analyzing genes involved in high ethylene production during fruit ripening in Japanese pear. This involves cleaved-amplified polymorphic sequences (CAPS) of two 1-aminocyclopropane-1-carboxylate (ACC) synthase genes (PPACS1 and PPACS2). Two CAPS markers (A for PPACS1 and B for PPACS2), associated with the amount of ethylene produced, were identified. Marker A was associated with high ethylene producers and marker B with moderate ethylene producers. The absence of these two markers enabled the identification of low ethylene producers. Using these markers, we have identified ethylene genotypes for 40 Japanese pear cultivars and two Chinese pear (P. bretschneideri) cultivars that are commercially important and used in breeding programs. Furthermore, we performed linkage analysis of these two genes in the F(2) population, which revealed that the recombination frequency between the two markers was 20.8 +/- 3.6%. This information is critical to the selection of parents and in breeding strategies to improve storage ability of Japanese pears.

  7. Enhancement of growth and salt tolerance of red pepper seedlings (Capsicum annuum L.) by regulating stress ethylene synthesis with halotolerant bacteria containing 1-aminocyclopropane-1-carboxylic acid deaminase activity.

    PubMed

    Siddikee, Md Ashaduzzaman; Glick, Bernard R; Chauhan, Puneet S; Yim, Woo jong; Sa, Tongmin

    2011-04-01

    Three 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase-producing halotolerant bacteria were isolated from West Coast soil of Yellow Sea, Incheon, South Korea and evaluated for their efficiency in improving red pepper plant growth under salt stress. The strains RS16, RS656 and RS111 were identified by 16S rRNA gene sequencing as Brevibacterium iodinum, Bacillus licheniformis and Zhihengliuela alba, respectively. Two hour exposure of 100, 150 and 200 mM NaCl stress on 8 day old red pepper seedlings caused 44, 64 and 74% increase ethylene production, while at 150 mM NaCl stress, inoculation of B. licheniformis RS656, Z. alba RS111, and Br. iodinum RS16 reduces ethylene production by 44, 53 and 57%, respectively. Similarly, 3 week old red pepper plants were subjected to salt stress for two weeks and approximately ∼50% reduction in growth recorded at 150 mM NaCl stress compared to negative control whereas bacteria inoculation significantly increase the growth compared to positive control. Salt stress also caused 1.3-fold reduction in the root/shoot dry weight ratio compared to the absence of salt while bacteria inoculation retained the biomass allocation similar to control plants. The salt tolerance index (ratio of biomass of salt stressed to non-stressed plant) was also significantly increased in inoculated plants compared to non-inoculated. Increase nutrient uptakes under salt stress by red pepper further evident that bacteria inoculation ameliorates salt stress effect. In summary, this study indicates that the use of ACC deaminase-producing halotolerant bacteria mitigates the salt stress by reducing salt stress-induced ethylene production on growth of red pepper plants.

  8. A strategy for promoting astaxanthin accumulation in Haematococcus pluvialis by 1-aminocyclopropane-1-carboxylic acid application.

    PubMed

    Lee, Changsu; Choi, Yoon-E; Yun, Yeoung-Sang

    2016-10-20

    The green algae Haematococcus pluvialis is a freshwater unicellular microalga belonging to Chlorophyceae. It is one of the best natural sources of astaxanthin, a secondary metabolite commonly used as an antioxidant and anti-inflammatory agent. Due to the importance of astaxanthin, various efforts have been made to increase its production. In this study, we attempted to develop a strategy for promoting astaxanthin accumulation in H. pluvialis using 1-aminocyclopropane-1-carboxylic acid (ACC), a precursor of ethylene (normally known as an aging hormone in plants). Our results demonstrated that ACC could enhance the growth of H. pluvialis, thereby promoting astaxanthin accumulation. Therefore, ACC has an indirect influence on astaxanthin production. We further verified the effect of ACC with a direct treatment of ethylene originated from banana peels. These results indicate that ethylene could be applied as an indirect method for enhancing growth and astaxanthin biosynthesis in H. pluvialis.

  9. In silico structural and functional analysis of Mesorhizobium ACC deaminase.

    PubMed

    Pramanik, Krishnendu; Soren, Tithi; Mitra, Soumik; Maiti, Tushar Kanti

    2017-02-11

    Nodulation is one of the very important processes of legume plants as it is the initiating event of fixing nitrogen. Although ethylene has essential role in normal plant metabolism but it has also negative impact on plants particularly in nodule formation in legume plants. It is also produced due to a variety of biotic or abiotic stresses. 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase is a rhizobial enzyme which cleaves ACC (immediate precursor of ethylene) into α-ketobutyrate and ammonia. As a result, the level of ethylene from the plant cells is decreased and the negative impact of ethylene on nodule formation is reduced. ACC deaminase is widely studied in several plant growth promoting rhizobacterial (PGPR) strains including many legume nodulating bacteria like Mesorhizobium sp. It is an important symbiotic nitrogen fixer belonging to the class - alphaproteobacteria under the order Rhizobiales. ACC deaminase has positive role in Legume-rhizobium symbiosis. Rhizobial ACC deaminase has the potentiality to reduce the adverse effects of ethylene, thereby triggering the nodulation process. The present study describes an in silico comparative structural (secondary structure prediction, homology modeling) and functional analysis of ACC deaminase from Mesorhizobium spp. to explore physico-chemical properties using a number of bio-computational tools. M. loti was selected as a representative species of Mesorhizobium genera for 3D modelling of ACC deaminase protein. Correlation by the phylogenetic relatedness on the basis of both ACC deaminase enzymes and respective acdS genes of different strains of Mesorhizobium has also studied.

  10. Molecular cloning of an 1-aminocyclopropane-1-carboxylate synthase from senescing carnation flower petals.

    PubMed

    Park, K Y; Drory, A; Woodson, W R

    1992-01-01

    Synthetic oligonucleotides based on the sequence of 1-aminocyclopropane-1-carboxylate (ACC) synthase from tomato were used to prime the synthesis and amplification of a 337 bp tomato ACC synthase cDNA by polymerase chain reaction (PCR). This PCR product was used to screen a cDNA library prepared from mRNA isolated from senescing carnation flower petals. Two cDNA clones were isolated which represented the same mRNA. The longer of the two clones (CARACC3) contained a 1950 bp insert with a single open reading frame of 516 amino acids encoding a protein of 58 kDa. The predicted protein from the carnation ACC synthase cDNA was 61%, 61%, 64%, and 51% identical to the deduced proteins from zucchini squash, winter squash, tomato, and apple, respectively. Genomic DNA gel blot analysis indicated the presence of at least a second gene in carnation which hybridized to CARACC3 under conditions of low stringency. ACC synthase mRNA accumulates during senescence of carnation flower petals concomitant with the increase in ethylene production and ACC synthase enzyme activity. Ethylene induced the accumulation of ACC synthase mRNA in presenescent petals. Wound-induced ethylene production in leaves was not associated with an increase in ACC synthase mRNA represented by CARACC3. These results indicate that CARACC3 represents an ACC synthase transcript involved in autocatalytic ethylene production in senescing flower petals.

  11. Transport and Metabolism of 1-Aminocyclopropane-1-carboxylic Acid in Sunflower (Helianthus annuus L.) Seedlings 1

    PubMed Central

    Finlayson, Scott A.; Foster, Kenneth R.; Reid, David M.

    1991-01-01

    Transport and metabolism of [2,3-14C] 1-aminocyclopropane-1-carboxylic acid (ACC) from roots to shoots in 4-day-old sunflower (Helianthus annuus L.) seedlings were studied. [14C]ACC was detected in, and 14C2H4 was evolved from, shoots 0.5 hours after [14C]ACC was supplied to roots. Ethylene emanation from the shoots returned to normal levels after 6 hours. The roots showed a similar pattern, although at 24 hours ethylene emanation was still slightly higher than in those plants that did not receive ACC. [14C]N-malonyl-ACC (MACC) was detected in both tissues at all times sampled. [14C]MACC levels surpassed [14C]ACC levels in the shoot at 2 hours, whereas [14C]MACC levels in the root remained below [14C]ACC levels until 6 hours, after which they were higher. Thin-layer chromatography analysis identified [14C] ACC in 1-hour shoot extracts, and [14C]MACC was identified in root tissues at 1 and 12 hours after treatment. [14C]ACC and [14C] MACC in the xylem sap of treated seedlings were identified by thin-layer chromatography. Xylem transport of [14C]ACC in treated seedlings, and transport of ACC in untreated seedlings, was confirmed by gas chromatography-mass spectrometry. Some evidence for the presence of [14C]MACC in xylem sap in [14C]ACC-treated seedlings is presented. A substantial amount of radioactivity in both ACC and MACC fractions was detected leaking from the roots over 24 hours. A second radiolabeled volatile compound was trapped in a CO2-trapping solution but not in mercuric perchlorate. Levels of this compound were highest after the peak of ACC levels and before peak MACC levels in both tissues, suggesting that an alternate pathway of ACC metabolism was operating in this system. PMID:16668342

  12. 1-Aminocyclopropane-1-Carboxylate Oxidase Activity Limits Ethylene Biosynthesis in Rumex palustris during Submergence

    PubMed Central

    Vriezen, Wim H.; Hulzink, Raymond; Mariani, Celestina; Voesenek, Laurentius A.C.J.

    1999-01-01

    Submergence strongly stimulates petiole elongation in Rumex palustris, and ethylene accumulation initiates and maintains this response in submerged tissues. cDNAs from R. palustris corresponding to a 1-aminocyclopropane-1-carboxylate (ACC) oxidase gene (RP-ACO1) were isolated from elongating petioles and used to study the expression of the corresponding gene. An increase in RP-ACO1 messenger was observed in the petioles and lamina of elongating leaves 2 h after the start of submergence. ACC oxidase enzyme activity was measured in homogenates of R. palustris shoots, and a relevant increase was observed within 12 h under water with a maximum after 24 h. We have shown previously that the ethylene production rate of submerged shoots does not increase significantly during the first 24 h of submergence (L.A.C.J. Voesenek, M. Banga, R.H. Thier, C.M. Mudde, F.M. Harren, G.W.M. Barendse, C.W.P.M. Blom [1993] Plant Physiol 103: 783–791), suggesting that under these conditions ACC oxidase activity is inhibited in vivo. We found evidence that this inhibition is caused by a reduction of oxygen levels. We hypothesize that an increased ACC oxidase enzyme concentration counterbalances the reduced enzyme activity caused by low oxygen concentration during submergence, thus sustaining ethylene production under these conditions. Therefore, ethylene biosynthesis seems to be limited at the level of ACC oxidase activity rather than by ACC synthase in R. palustris during submergence. PMID:10482674

  13. Purification and characterization of 1-aminocyclopropane-1-carboxylate synthase from etiolated mung bean hypocotyls.

    PubMed

    Tsai, D S; Arteca, R N; Bachman, J M; Phillips, A T

    1988-08-01

    1-Aminocyclopropane-1-carboxylate (ACC) synthase, EC 4.4.1.14, was purified to homogeneity from etiolated mung bean hypocotyl segments. This was made possible by the ability to elevate the enzyme level markedly through hormone treatments and by stabilization of the enzyme with high phosphate concentrations. The four-step procedure resulted in 1050-fold purification with 25% yield, and consisted of stepwise elution from hydroxylapatite, chromatography on phenyl-Sepharose CL-4B, gradient elution from hydroxylapatite, and fast protein liquid chromatography (FPLC) on a MonoQ anion-exchange column. FPLC-purified ACC synthase migrated as a single band of Mr 65,000 on denaturing polyacrylamide gel electrophoresis. The molecular weight of native enzyme by Bio-Gel A-0.5 M chromatography was 125,000, indicating that the enzyme probably exists as a dimer of identical 65,000 Mr subunits. The mung bean ACC synthase exhibited a pH optimum of 8.0 for activity and a Km for S-adenosylmethionine (AdoMet) of 55 microM at 30 degrees C. It exhibited an Arrhenius activation energy of 12 kcal mol-1 degree-1 and was inactivated at temperatures in excess of 40 degrees C. The specific activity for pure ACC synthase was 21 mumol of ACC formed/mg protein/h when determined under optimal conditions with 400 microM AdoMet.

  14. Cloning and expression of the 1-aminocyclopropane-1-carboxylic oxidase gene from Agrostis stolonifera.

    PubMed

    Xiao, G Z; Li, L J; Teng, K; Chao, Y H; Han, L B

    2016-11-03

    A gene encoding 1-aminocyclopropane-1-carboxylic oxidase (ACO), which catalyzes the terminal step in ethylene biosynthesis, was isolated from Agrostis stolonifera. The AsACO gene is composed of 975 bp, encoding 324 amino acids. Three exons interspersed by two introns form AsACO gDNA. A BLAST search of the nucleotide sequence revealed a high level of similarity (79-91%) between AsACO and ACO genes of other plants. A phylogenetic tree was constructed via BLAST in the NCBI, and revealed the highest homology with wheat TaACO. The calculated molecular mass and predicted isoelectric point of AsACO were 36.25 and 4.89 kDa, respectively. Analysis of subcellular localization revealed that AsACO is located in the nucleus and cytoplasm. The Fe(II)-binding cofactors and cosubstrate were identified, pertaining to the ACO family. The expression patterns of AsACO were determined by quantitative real time PCR. AsACO expression was highest in the stem, and was strongly up-regulated in response to ethephon, methyl jasmonate, salicylic acid, and cold temperature, but down-regulated in response to drought and NaCl treatment. The protein encoded by AsACO exhibited ACC oxidase activity in vitro. Taken together, these findings suggest that AsACO contains domains common to the ACO family, and is induced in response to exogenous hormones. Conversely, some abiotic stress conditions can inhibit AsACO expression.

  15. Dynamic 1-Aminocyclopropane-1-Carboxylate-Synthase and -Oxidase Transcript Accumulation Patterns during Pollen Tube Growth in Tobacco Styles1

    PubMed Central

    Weterings, Koen; Pezzotti, Mario; Cornelissen, Marc; Mariani, Celestina

    2002-01-01

    In flowering plants, pollination of the stigma sets off a cascade of responses in the distal flower organs. Ethylene and its biosynthetic precursor 1-aminocyclopropane-1-carboxylate (ACC) play an important role in regulating these responses. Because exogenous application of ethylene or ACC does not invoke the full postpollination syndrome, the pollination signal probably consists of a more complex set of stimuli. We set out to study how and when the pollination signal moves through the style of tobacco (Nicotiana tabacum) by analyzing the expression patterns of pistil-expressed ACC-synthase and -oxidase genes. Results from this analysis showed that pollination induces high ACC-oxidase transcript levels in all cells of the transmitting tissue. ACC-synthase mRNA accumulated only in a subset of transmitting tract cells and to lower levels as compared with ACC-oxidase. More significantly, we found that although ACC-oxidase transcripts accumulate to uniform high levels, the ACC-synthase transcripts accumulate in a wave-like pattern in which the peak coincides with the front of the ingrowing pollen tube tips. This wave of ACC-synthase expression can also be induced by incongruous pollination and (partially) by wounding. This indicates that wounding-like features of pollen tube invasion might be part of the stimuli evoking the postpollination response and that these stimuli are interpreted differently by the regulatory mechanisms of the ACC-synthase and -oxidase genes. PMID:12427986

  16. Cell wall integrity controls root elongation via a general 1-aminocyclopropane-1-carboxylic acid-dependent, ethylene-independent pathway.

    PubMed

    Tsang, Dat L; Edmond, Clare; Harrington, Jennifer L; Nühse, Thomas S

    2011-06-01

    Cell expansion in plants requires cell wall biosynthesis and rearrangement. During periods of rapid elongation, such as during the growth of etiolated hypocotyls and primary root tips, cells respond dramatically to perturbation of either of these processes. There is growing evidence that this response is initiated by a cell wall integrity-sensing mechanism and dedicated signaling pathway rather than being an inevitable consequence of lost structural integrity. However, the existence of such a pathway in root tissue and its function in a broader developmental context have remained largely unknown. Here, we show that various types of cell wall stress rapidly reduce primary root elongation in Arabidopsis (Arabidopsis thaliana). This response depended on the biosynthesis of 1-aminocyclopropane-1-carboxylic acid (ACC). In agreement with the established ethylene signaling pathway in roots, auxin signaling and superoxide production are required downstream of ACC to reduce elongation. However, this cell wall stress response unexpectedly does not depend on the perception of ethylene. We show that the short-term effect of ACC on roots is partially independent of its conversion to ethylene or ethylene signaling and that this ACC-dependent pathway is also responsible for the rapid reduction of root elongation in response to pathogen-associated molecular patterns. This acute response to internal and external stress thus represents a novel, noncanonical signaling function of ACC.

  17. Glutathione Regulates 1-Aminocyclopropane-1-Carboxylate Synthase Transcription via WRKY33 and 1-Aminocyclopropane-1-Carboxylate Oxidase by Modulating Messenger RNA Stability to Induce Ethylene Synthesis during Stress.

    PubMed

    Datta, Riddhi; Kumar, Deepak; Sultana, Asma; Hazra, Saptarshi; Bhattacharyya, Dipto; Chattopadhyay, Sharmila

    2015-12-01

    Glutathione (GSH) plays a fundamental role in plant defense-signaling network. Recently, we have established the involvement of GSH with ethylene (ET) to combat environmental stress. However, the mechanism of GSH-ET interplay still remains unexplored. Here, we demonstrate that GSH induces ET biosynthesis by modulating the transcriptional and posttranscriptional regulations of its key enzymes, 1-aminocyclopropane-1-carboxylate synthase (ACS) and 1-aminocyclopropane-1-carboxylate oxidase (ACO). Transgenic Arabidopsis (Arabidopsis thaliana) plants with enhanced GSH content (AtECS) exhibited remarkable up-regulation of ACS2, ACS6, and ACO1 at transcript as well as protein levels, while they were down-regulated in the GSH-depleted phytoalexin deficient2-1 (pad2-1) mutant. We further observed that GSH induced ACS2 and ACS6 transcription in a WRKY33-dependent manner, while ACO1 transcription remained unaffected. On the other hand, the messenger RNA stability for ACO1 was found to be increased by GSH, which explains our above observations. In addition, we also identified the ACO1 protein to be a subject for S-glutathionylation, which is consistent with our in silico data. However, S-glutathionylation of ACS2 and ACS6 proteins was not detected. Further, the AtECS plants exhibited resistance to necrotrophic infection and salt stress, while the pad2-1 mutant was sensitive. Exogenously applied GSH could improve stress tolerance in wild-type plants but not in the ET-signaling mutant ethylene insensitive2-1, indicating that GSH-mediated resistance to these stresses occurs via an ET-mediated pathway. Together, our investigation reveals a dual-level regulation of ET biosynthesis by GSH during stress.

  18. Perspective of plant growth promoting rhizobacteria (PGPR) containing ACC deaminase in stress agriculture.

    PubMed

    Saleem, Muhammad; Arshad, Muhammad; Hussain, Sarfraz; Bhatti, Ahmad Saeed

    2007-10-01

    Ethylene is a gaseous plant growth hormone produced endogenously by almost all plants. It is also produced in soil through a variety of biotic and abiotic mechanisms, and plays a key role in inducing multifarious physiological changes in plants at molecular level. Apart from being a plant growth regulator, ethylene has also been established as a stress hormone. Under stress conditions like those generated by salinity, drought, waterlogging, heavy metals and pathogenicity, the endogenous production of ethylene is accelerated substantially which adversely affects the root growth and consequently the growth of the plant as a whole. Certain plant growth promoting rhizobacteria (PGPR) contain a vital enzyme, 1-aminocyclopropane-1-carboxylate (ACC) deaminase, which regulates ethylene production by metabolizing ACC (an immediate precursor of ethylene biosynthesis in higher plants) into alpha-ketobutyrate and ammonia. Inoculation with PGPR containing ACC deaminase activity could be helpful in sustaining plant growth and development under stress conditions by reducing stress-induced ethylene production. Lately, efforts have been made to introduce ACC deaminase genes into plants to regulate ethylene level in the plants for optimum growth, particularly under stressed conditions. In this review, the primary focus is on giving account of all aspects of PGPR containing ACC deaminase regarding alleviation of impact of both biotic and abiotic stresses onto plants and of recent trends in terms of introduction of ACC deaminase genes into plant and microbial species.

  19. Biosynthesis of 1-aminocyclopropane-1-carboxylic acid and ethylene from delta-aminolevulinic acid in ripening tomato fruits

    SciTech Connect

    El-Rayes, D.E.D.A.

    1987-01-01

    A new pathway for ethylene (C/sub 2/H/sub 4/) biosynthesis, which utilizes delta-aminolevulinic acid (ALA) as a precursor of 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor of C/sub 2/H/sub 4/, is presented. ALA enhanced ACC accumulation to 410% and C/sub 2/H/sub 4/ production to 232% of the control. The C/sub 2/H/sub 4/ production rate varied with the ALA concentration and the stage of tomato fruit development. As the ALA concentration increased from zero to 40 mM, the C/sub 2/H/sub 4/ production rate increased. Both treated and untreated pericarp discs from fruits at the pink stage of development yielded the largest C/sub 2/H/sub 4/ production rate. Radioactivity from (2,3-/sup 3/H)ALA was detected in both ACC and C/sub 2/H/sub 4/, and radioactivity from (4-/sup 14/C)ALA was detected in ACC and CO/sub 2/ but not in C/sub 2/H/sub 4/. However, radioactivity from (5-/sup 14/C)ALA was detected in CO/sub 2/, and its amount was greater than that obtained from (4-/sup 14/C)ALA. Neither ACC nor C/sub 2/H/sub 4/ showed any radioactivity when (5-/sup 14/C)ALA was supplied to the fruit discs. In addition, when (2,3-/sup 3/H)ALA or (4-/sup 14/C)ALA was supplied to the fruit discs, radioactivity was detected in other metabolites such as fumarate, succinate, malate, glutamate, glutamine, ..cap alpha..-ketoglutarate, and methionine, but the amount of radioactivity was insignificant as compared with the amount of radioactivity found in C/sub 2/H/sub 4/ and ACC.

  20. Expression of 1-Aminocyclopropane-1-Carboxylate Oxidase during Leaf Ontogeny in White Clover1

    PubMed Central

    Hunter, Donald A.; Yoo, Sang Dong; Butcher, Stephen M.; McManus, Michael T.

    1999-01-01

    We examined the expression of three distinct 1-aminocyclopropane-1-carboxylic acid oxidase genes during leaf ontogeny in white clover (Trifolium repens). Significant production of ethylene occurs at the apex, in newly initiated leaves, and in senescent leaf tissue. We used a combination of reverse transcriptase-polymerase chain reaction and 3′-rapid amplification of cDNA ends to identify three distinct DNA sequences designated TRACO1, TRACO2, and TRACO3, each with homology to 1-aminocyclopropane-1-carboxylic acid oxidase. Southern analysis confirmed that these sequences represent three distinct genes. Northern analysis revealed that TRACO1 is expressed specifically in the apex and TRACO2 is expressed in the apex and in developing and mature green leaves, with maximum expression in developing leaf tissue. The third gene, TRACO3, is expressed in senescent leaf tissue. Antibodies were raised to each gene product expressed in Escherichia coli, and western analysis showed that the TRACO1 antibody recognizes a protein of approximately 205 kD (as determined by gradient sodium dodecyl sulfate-polyacylamide gel electrophoresis) that is expressed preferentially in apical tissue. The TRACO2 antibody recognizes a protein of approximately 36.4 kD (as determined by gradient sodium dodecyl sulfate-polyacylamide gel electrophoresis) that is expressed in the apex and in developing and mature green leaves, with maximum expression in mature green tissue. No protein recognition by the TRACO3 antibody could be detected in senescent tissue or at any other stage of leaf development. PMID:10318691

  1. Isolation and characterization of ACC deaminase-producing fluorescent pseudomonads, to alleviate salinity stress on canola (Brassica napus L.) growth.

    PubMed

    Jalili, Farzad; Khavazi, Kazem; Pazira, Ebrahim; Nejati, Alireza; Rahmani, Hadi Asadi; Sadaghiani, Hasan Rasuli; Miransari, Mohammad

    2009-04-01

    Salinity stress is of great importance in arid and semi-arid areas of the world due to its impact in reducing crop yield. Under salinity stress, the amount of 1-aminocyclopropane-1-carboxylate (ACC), a precursor for ethylene production in plants, increases. Here, we conducted research under the hypothesis that isolated ACC deaminase-producing Pseudomonas fluorescens and Pseudomonas putida can alleviate the stressful effects of salinity on canola (Brassica napus L.) growth. The experiments were conducted in the Soil and Water Research Institute, Tehran, Iran. Seven experimental stages were conducted to isolate and characterize ACC deaminase-producing Pseudomonas fluorescens strains and to determine factors enhancing their growth and, consequently, their effects on the germination of canola seeds. Under salinity stress, in 14% of the isolates, ACC deaminase activity was observed, indicating that they were able to utilize ACC as the sole N-source. Bacterial strains differed in their ability to synthesize auxin and hydrogen cyanide compounds, as well as in their ACC deaminase activity. Under salinity stress, the rate of germinating seeds inoculated with the strains of ACC deaminase-producing Pseudomonas fluorescens and Pseudomonas putida, and seedling growth was significantly higher. These results indicate the significance of soil biological activities, including the activities of plant growth-promoting bacteria, in the alleviation of soil stresses such as salinity on plant growth.

  2. Ethylene Evolution following Treatment with 1-Aminocyclopropane-1-carboxylic Acid and Ethephon in an in Vitro Olive Shoot System in Relation to Leaf Abscission

    PubMed Central

    Lavee, S.; Martin, George C.

    1981-01-01

    1-Aminocyclopropane-1-carboxylic acid (ACC) supplied via the cut base of detached olive shoots caused a burst of ethylene from leaves, but other cyclopropanes tested did not exhibit this effect. Ethephon (ET) and another ethylene-releasing compound caused a prolonged increase in ethylene evolution. ACC had only a very limited effect on leaf abscission regardless of concentration, whereas shoots placed with cut bases in ET for 60 to 80 minutes exhibited 100% leaf abscission within 90 hours. Shoots with inflorescences treated with ET just prior to anthesis began to wilt in vitro within 20 to 30 hours and failed to exhibit leaf abscission. At earlier stages of development, ET induced more leaf abscission on reproductive shoots than on vegetative shoots. It is suggested that the duration of ethylene evolution from the leaves governs their potential for abscission and that bursts of ethylene evolution even though large in amount may not induce abscission. Images PMID:16661837

  3. 1-Aminocyclopropane-1-Carboxylic Acid Transported from Roots to Shoots Promotes Leaf Abscission in Cleopatra Mandarin (Citrus reshni Hort. ex Tan.) Seedlings Rehydrated after Water Stress.

    PubMed

    Tudela, D; Primo-Millo, E

    1992-09-01

    The effect of water stress and subsequent rehydration on 1-aminocyclopropane-1-carboxylic acid (ACC) content, ACC synthase activity, ethylene production, and leaf abscission was studied in Cleopatra mandarin (Citrus reshni Hort. ex Tan.) seedlings. Leaf abscission occurred when drought-stressed plants were allowed to rehydrate, whereas no abscission was observed in plants under water stress conditions. In roots of water-stressed plants, a high ACC accumulation and an increase in ACC synthase activity were observed. Neither increase in ACC content nor significant ethylene production were detected in leaves of water-stressed plants. After rehydration, a sharp rise in ACC content and ethylene production was observed in leaves of water-stressed plants. Content of ACC in xylem fluid was 10-fold higher in plants rehydrated for 2 h after water stress than in nonstressed plants. Leaf abscission induced by rehydration after drought stress was inhibited when roots or shoots were treated before water stress with aminooxyacetic acid (AOA, inhibitor of ACC synthase) or cobalt ion (inhibitor of ethylene-forming enzyme), respectively. However, AOA treatments to shoots did not suppress leaf abscission. The data indicate that water stress promotes ACC synthesis in roots of Cleopatra mandarin seedlings. Rehydration of plants results in ACC transport to the shoots, where it is oxidized to ethylene. Subsequently, this ethylene induces leaf abscission.

  4. Isolation of an 1-aminocyclopropane-1-carboxylate oxidase gene from mulberry (Morus alba L.) and analysis of the function of this gene in plant development and stresses response.

    PubMed

    Pan, Gang; Lou, Chengfu

    2008-07-31

    Mulberry (Morus alba) is an important crop tree involved in sericulture and pharmaceuticals. To further understand the development and the environmental adaptability mechanism of mulberry, a cDNA of the gene MaACO1 encoding 1-aminocyclopropane-1-carboxylate oxidase was isolated from mulberry. This was used to investigate stress-responsive expression in mulberry. Developmental expression of ACC oxidase in mulberry leaves and spatial expression in mulberry flowers were also investigated. Damage and low-temperature treatment promoted the expression of MaACO1 in mulberry. In leaves, expression of the MaACO1 gene increased in cotyledons and the lowest leaves with leaf development, but showed reduced levels in emerging leaves. In flowers, the pollinated stigma showed the highest expression level, followed by the unpollinated stigma, ovary, and immature flowers. These results suggest that high MaACO1 expression may be predominantly associated with tissue aging or senescence in mulberry.

  5. Bacteria with ACC deaminase can promote plant growth and help to feed the world.

    PubMed

    Glick, Bernard R

    2014-01-20

    To feed all of the world's people, it is necessary to sustainably increase agricultural productivity. One way to do this is through the increased use of plant growth-promoting bacteria; recently, scientists have developed a more profound understanding of the mechanisms employed by these bacteria to facilitate plant growth. Here, it is argued that the ability of plant growth-promoting bacteria that produce 1-aminocyclopropane-1-carboxylate (ACC) deaminase to lower plant ethylene levels, often a result of various stresses, is a key component in the efficacious functioning of these bacteria. The optimal functioning of these bacteria includes the synergistic interaction between ACC deaminase and both plant and bacterial auxin, indole-3-acetic acid (IAA). These bacteria not only directly promote plant growth, they also protect plants against flooding, drought, salt, flower wilting, metals, organic contaminants, and both bacterial and fungal pathogens. While a considerable amount of both basic and applied work remains to be done before ACC deaminase-producing plant growth-promoting bacteria become a mainstay of plant agriculture, the evidence indicates that with the expected shift from chemicals to soil bacteria, the world is on the verge of a major paradigm shift in plant agriculture.

  6. Various effects of fluorescent bacteria of the genus Pseudomonas containing ACC deaminase on wheat seedling growth.

    PubMed

    Magnucka, Elżbieta G; Pietr, Stanisław J

    2015-12-01

    The study evaluates the effect of rhizobacteria having 1-aminocyclopropane-1-carboxylate deaminase (ACCd) on the development of wheat seedlings. This enzyme has been proposed to play a key role in microbe-plant association. Three fluorescent pseudomonads containing this deaminase were selected from 70 strains of pseudomonads isolated from rhizosphere of wheat (Triticum aestivum L.) and rape (Brassica napus L.). These bacteria, varied significantly in the ability to both biosynthesize auxins and hydrolyze ACC. Among them, Pseudomonas brassicacearum subsp. brassicacearum strain RZ310 presented the highest activities of ACC deaminase during 96h of growth in liquid Dworkin and Foster (DF) salt medium. Additionally, this rape rhizosphere strain did not produce indoles. Two other isolates, Pseudomonas sp. PO283 and Pseudomonas sp. PO366, secreted auxins only in the presence of their precursor. Phylogenetic analysis of the 16S rRNA gene and four other protein-encoding genes indicated that these wheat rhizosphere isolates belonged to the fluorescent Pseudomonas group. Moreover, the effects of these strains on wheat seedling growth under in vitro conditions were markedly dependent on both their cell suspensions used to grain inoculation and nutrient conditions. Strains tested had beneficial influence on wheat seedlings mainly at low cell densities. In addition, access to nutrients markedly changed bacteria action on cereal growth. Their presence generally favored the positive effects of pseudomonads on length and the estimated biomasses of wheat coleoptiles. Despite these general rules, impacts of each isolate on the growth parameters of cereal seedlings were unique.

  7. Amelioration of high salinity stress damage by plant growth-promoting bacterial endophytes that contain ACC deaminase.

    PubMed

    Ali, Shimaila; Charles, Trevor C; Glick, Bernard R

    2014-07-01

    Plant growth and productivity is negatively affected by soil salinity. However, it is predicted that plant growth-promoting bacterial (PGPB) endophytes that contain 1-aminocyclopropane-1-carboxylate (ACC) deaminase (E.C. 4.1.99.4) can facilitate plant growth and development in the presence of a number of different stresses. In present study, the ability of ACC deaminase containing PGPB endophytes Pseudomonas fluorescens YsS6, Pseudomonas migulae 8R6, and their ACC deaminase deficient mutants to promote tomato plant growth in the absence of salt and under two different levels of salt stress (165 mM and 185 mM) was assessed. It was evidence that wild-type bacterial endophytes (P. fluorescens YsS6 and P. migulae 8R6) promoted tomato plant growth significantly even in the absence of stress (salinity). Plants pretreated with wild-type ACC deaminase containing endophytic strains were healthier and grew to a much larger size under high salinity stress compared to plants pretreated with the ACC deaminase deficient mutants or no bacterial treatment (control). The plants pretreated with ACC deaminase containing bacterial endophytes exhibit higher fresh and dry biomass, higher chlorophyll contents, and a greater number of flowers and buds than the other treatments. Since the only difference between wild-type and mutant bacterial endophytes was ACC deaminase activity, it is concluded that this enzyme is directly responsible for the different behavior of tomato plants in response to salt stress. The use of PGPB endophytes with ACC deaminase activity has the potential to facilitate plant growth on land that is not normally suitable for the majority of crops due to their high salt contents.

  8. Tissue Localization of a Submergence-Induced 1-Aminocyclopropane-1-Carboxylic Acid Synthase in Rice1

    PubMed Central

    Zhou, Zhongyi; de Almeida Engler, Janice; Rouan, Dominique; Michiels, Frank; Van Montagu, Marc; Van Der Straeten, Dominique

    2002-01-01

    At least two 1-aminocyclopropane-1-carboxylic acid synthase genes (ACS) are implicated in the submergence response of rice (Oryza sativa). Previously, the OS-ACS5 gene has been shown to be induced during short- as well as long-term complete submergence of seedlings and to be controlled by a balance of gibberellin and abscisic acid in both lowland and deepwater rice. This study demonstrates that OS-ACS5 mRNA is localized in specific tissues and cells both during normal development and in response to complete submergence. The temporal and spatial regulation of OS-ACS5 expression is presented by in situ hybridization and histochemical analysis of β-glucuronidase (GUS) activity in transgenic rice carrying an OS-ACS5-gus fusion. Whole-mount in situ hybridization revealed that in air-grown rice seedlings, OS-ACS5 was expressed at a low level in the shoot apex, meristems, leaf, and adventitious root primordia, and in vascular tissues of nonelongated stems and leaf sheaths. In response to complete submergence, the expression in vascular bundles of young stems and leaf sheaths was strongly induced. The results of histochemical GUS assays were consistent with those found by whole-mount in situ hybridization. Our findings suggest that OS-ACS5 plays a role in vegetative growth of rice under normal conditions and is also recruited for enhanced growth upon complete submergence. The possible implication of OS-ACS5 in root-shoot communication during submergence stress and its putative role in aerenchyma formation upon low-oxygen stress are discussed. PMID:12011339

  9. Involvement of ethylene and 1-aminocyclopropane-1-carboxylate synthase gene in regulation of programmed cell death during rose (Rosa x hybrida) flower development.

    PubMed

    Pan, Hai-Chun; Li, Ji-Hong; Wang, Xian-Ze

    2005-08-01

    Programmed cell death (PCD) is an integral part of plant development. Flower petal usually has the shortest lifetime among all plant organs. There must be a sensitive, tightly controlled PCD in the life cycle of the flower. To understand its mechanism, the ethylene production rate of petals and its correlation with degree of senescence, 1-aminocyclopropane-1-carboxylate (ACC) synthase gene expression, ACC synthase activity and ACC content were determined through the whole flower development period which was arbitrarily divided into five stages depending on appearance of the flower. The results showed that ethylene was not detectable at stages 1 and 2, appeared at stage 3 and increased at stage 5. Transcript of ACC synthase gene did not accumulate at stages 1 and 2, but did so at stages 3-5, and increased gradually at stage 5. ACC synthase activity and ACC content changed in similar way to ethylene production. Ethylene plays a critical role in initiation of rose flower senescence through regulating petal PCD.

  10. Deletion of the carboxyl-terminal region of 1-aminocyclopropane-1-carboxylic acid synthase, a key protein in the biosynthesis of ethylene, results in catalytically hyperactive, monomeric enzyme.

    PubMed

    Li, N; Mattoo, A K

    1994-03-04

    1-Aminocyclopropane-1-carboxylic acid (ACC) synthase is a key enzyme regulating biosynthesis of the plant hormone ethylene. The expression of an enzymatically active, wound-inducible tomato (Lycopersicon esculentum L. cv Pik-Red) ACC synthase (485 amino acids long) in a heterologous Escherichia coli system allowed us to study the importance of hypervariable COOH terminus in enzymatic activity and protein conformation. We constructed several deletion mutants of the gene, expressed these in E. coli, purified the protein products to apparent homogeneity, and analyzed both conformation and enzyme kinetic parameters of the wild-type and truncated ACC syntheses. Deletion of the COOH terminus through Arg429 results in complete inactivation of the enzyme. Deletion of 46-52 amino acids from the COOH terminus results in an enzyme that has nine times higher affinity for the substrate S-adenosylmethionine than the wild-type enzyme. The highly efficient, truncated ACC synthase was found to be a monomer of 52 +/- 1.8 kDa as determined by gel filtration, whereas the wild-type ACC synthase, analyzed under similar conditions, is a dimer. These results demonstrate that the non-conserved COOH terminus of ACC synthase affects its enzymatic function as well as dimerization.

  11. 1-Aminocyclopropane-1-carboxylic acid and abscisic acid during the germination of sugar beet (Beta vulgaris L.): a comparative study of fruits and seeds.

    PubMed

    Hermann, Katrin; Meinhard, Juliane; Dobrev, Peter; Linkies, Ada; Pesek, Bedrich; Hess, Barbara; Machácková, Ivana; Fischer, Uwe; Leubner-Metzger, Gerhard

    2007-01-01

    The control of sugar beet (Beta vulgaris L.) germination by plant hormones was studied by comparing fruits and seeds. Treatment of sugar beet fruits and seeds with gibberellins, brassinosteroids, auxins, cytokinins, and jasmonates or corresponding hormone biosynthesis inhibitors did not appreciably affect radicle emergence of fruits or seeds. By contrast, treatment with ethylene or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) promoted radicle emergence of fruits and seeds. Abscisic acid (ABA) acted as an antagonist of ethylene and inhibited radicle emergence of seeds, but not appreciably of fruits. High endogenous contents of ACC and of ABA were evident in seeds and pericarps of dry mature fruits, but declined early during imbibition. ABA-treatment of seeds and fruits induced seed ACC accumulation while ACC-treatment did not affect the seed ABA content. Transcripts of ACC oxidase (ACO, ethylene-forming enzyme) and ABA 8'-hydroxylase (CYP707A, ABA-degrading enzyme) accumulate in fruits and seeds upon imbibition. ABA and ACC and the pericarp did not affect the seed CYP707A transcript levels. By contrast, seed ACO transcript accumulation was promoted by ABA and by pericarp removal, but not by ACC. Quantification of the endogenous ABA and ACC contents, ABA and ACC leaching, and ethylene evolution, demonstrate that an embryo-mediated active ABA extrusion system is involved in keeping the endogenous seed ABA content low by 'active ABA leaching', while the pericarp restricts ACC leaching during imbibition. Sugar beet radicle emergence appears to be controlled by the pericarp, by ABA and ACC leaching, and by an ABA-ethylene antagonism that affects ACC biosynthesis and ACO gene expression.

  12. Simultaneous analysis of apolar phytohormones and 1-aminocyclopropan-1-carboxylic acid by high performance liquid chromatography/electrospray negative ion tandem mass spectrometry via 9-fluorenylmethoxycarbonyl chloride derivatization.

    PubMed

    Ziegler, Jörg; Qwegwer, Jakob; Schubert, Melvin; Erickson, Jessica L; Schattat, Martin; Bürstenbinder, Katharina; Grubb, C Douglas; Abel, Steffen

    2014-10-03

    A strategy to detect and quantify the polar ethylene precursor 1-aminocyclopropan-1-carboxylic acid (ACC) along with the more apolar phytohormones abscisic acid (ABA), indole-3-acetic acid (IAA), jasmonic acid (JA), jasmonic acid-isoleucine conjugate (JA-Ile), 12-oxo-phytodienoic acid (OPDA), trans-zeatin, and trans-zeatin 9-riboside using a single extraction is presented. Solid phase resins commonly employed for extraction of phytohormones do not allow the recovery of ACC. We circumvent this problem by attaching an apolar group to ACC via derivatization with the amino group specific reagent 9-fluorenylmethoxycarbonyl chloride (Fmoc-Cl). Derivatization in the methanolic crude extract does not modify other phytohormones. The derivatized ACC could be purified and detected together with the more apolar phytohormones using common solid phase extraction resins and reverse phase HPLC/electrospray negative ion tandem mass spectrometry. The limit of detection was in the low nanomolar range for all phytohormones, a sensitivity sufficient to accurately determine the phytohormone levels from less than 50mg (fresh weight) of Arabidopsis thaliana and Nicotiana benthamiana tissues. Comparison with previously published phytohormone levels and the reported changes in phytohormone levels after stress treatments confirmed the accuracy of the method.

  13. Bio-inspired amino acid oxidation by a non-heme iron catalyst modeling the action of 1-aminocyclopropane-1-carboxylic acid oxidase.

    PubMed

    Baráth, Gábor; Kaizer, József; Pap, József Sándor; Speier, Gábor; El Bakkali-Taheri, Nadia; Simaan, A Jalila

    2010-10-21

    In this communication we describe the first example of a biomimetic mononuclear iron complex, [Fe(III)(Salen)Cl] (Salen = N,N'-bis(salicylidene)-ethylenediaminato), that highly selectively and efficiently catalyzes the oxidation of 1-aminocyclopropane-1-carboxylic acid (ACCH), α-aminoisobutyric acid (AIBH), and alanine (ALAH) to ethylene or the corresponding carbonyl compounds, mimicking the action of the non-heme iron enzyme 1-aminocyclopropane-1-carboxylic acid oxidase (ACCO).

  14. Radioisotope assay for 1-aminocyclopropane-1-carboxylic acid synthase: s-adenosylhomocysteine analogs as inhibitors of the enzyme involved in plant senescence

    SciTech Connect

    Miura, G.A.; Chiang, P.K.

    1985-01-01

    A simple and rapid radioisotopic assay for 1-aminocyclopropane-1-carboxylic acid (ACC) synthase was developed, an enzyme involved in the biosynthesis of the plant hormone ethylene. The assay utilizes an AG50-X4(NH4 (+)) column which separates S-adenosyl-L-(carboxyl-/sup 14/C)methionine (AdoMet) from the product (/sup 14/C)acc, since the latter is not bound to the resin while (/sup 14/C)adoMet is. As opposed to other assays, this procedure measures ACC directly and does not require further conversion to ethylene. When an enzyme preparation from ripe-tomato fruits (Lycopersicon esculentum Mill) was assayed, an I/sub 50/ of 2.5 + or - 0.8 micrometers for sinefungin and a K/sub m/ of 27 + or - 2 micrometers for AdoMet were obtained; these values were in good agreement with previous previous determinations made with a gas-chromatographic assay. When other nucleosides were tested as inhibitors the following order of decreasing activity was found: sinefungin, S-adenosylhomocysteine (AdoHcy), AdoHcy sulfoxide, S-n-butyladenosine, 3-deaza-adenosylhomocysteine, S-isobutyladenosine, S-isobutyladenosine, S-isobutyl-l-deazaadenosine. In contrast, S-isobutyl-3-deazaadenosine, S-isobutyl-7-deazaadenosine, 3-deazaadenosine, and adenodine were not inhibitory.

  15. Biochemistry and genetics of ACC deaminase: a weapon to “stress ethylene” produced in plants

    PubMed Central

    Singh, Rajnish P.; Shelke, Ganesh M.; Kumar, Anil; Jha, Prabhat N.

    2015-01-01

    1-aminocyclopropane-1-carboxylate deaminase (ACCD), a pyridoxal phosphate-dependent enzyme, is widespread in diverse bacterial and fungal species. Owing to ACCD activity, certain plant associated bacteria help plant to grow under biotic and abiotic stresses by decreasing the level of “stress ethylene” which is inhibitory to plant growth. ACCD breaks down ACC, an immediate precursor of ethylene, to ammonia and α-ketobutyrate, which can be further metabolized by bacteria for their growth. ACC deaminase is an inducible enzyme whose synthesis is induced in the presence of its substrate ACC. This enzyme encoded by gene AcdS is under tight regulation and regulated differentially under different environmental conditions. Regulatory elements of gene AcdS are comprised of the regulatory gene encoding LRP protein and other regulatory elements which are activated differentially under aerobic and anaerobic conditions. The role of some additional regulatory genes such as AcdB or LysR may also be required for expression of AcdS. Phylogenetic analysis of AcdS has revealed that distribution of this gene among different bacteria might have resulted from vertical gene transfer with occasional horizontal gene transfer (HGT). Application of bacterial AcdS gene has been extended by developing transgenic plants with ACCD gene which showed increased tolerance to biotic and abiotic stresses in plants. Moreover, distribution of ACCD gene or its homolog's in a wide range of species belonging to all three domains indicate an alternative role of ACCD in the physiology of an organism. Therefore, this review is an attempt to explore current knowledge of bacterial ACC deaminase mediated physiological effects in plants, mode of enzyme action, genetics, distribution among different species, ecological role of ACCD and, future research avenues to develop transgenic plants expressing foreign AcdS gene to cope with biotic and abiotic stressors. Systemic identification of regulatory circuits

  16. Characterization of ACC deaminase gene in Pseudomonas entomophila strain PS-PJH isolated from the rhizosphere soil.

    PubMed

    Kamala-Kannan, Seralathan; Lee, Kui-Jae; Park, Seung-Moon; Chae, Jong-Chan; Yun, Bong-Sik; Lee, Yong Hoon; Park, Yool-Jin; Oh, Byung-Taek

    2010-04-01

    The enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase cleaves the ethylene precursor ACC into alpha-ketobutyrate and ammonia. The decreased level of ethylene allows the plant to be more resistant to a wide environmental stress including plant pathogens. In the present study, we characterized the ACC deaminase activity of a Pseudomonas entomophila strain PS-PJH isolated from the red pepper rhizosphere region of red pepper grown at Jinan, Korea. The isolate produced 23.8 +/- 0.4 micromol of alpha-ketobutyrate/mg of protein/h during ACC deamination under in vitro conditions. Polymerase chain reaction for acdS gene showed that the isolated P. entomophila strain PS-PJH carry sequences similar to the known acdS genes. Results of the multiple sequence alignment revealed >99% identity (nucleotide and amino acid) with acdS gene of Pseudomonas putida strains AM15 and UW4. The isolated bacteria promoted 43.3 and 34.1% of growth in Raphanus sativus and Lactuca sativa plants, respectively. Based on the 16S-23S internal transcribed spacer region sequences, the isolate was identified as P. entomophila. To the best of our knowledge this is the first study to report the acdS gene in P. entomophila.

  17. Glutathione Regulates 1-Aminocyclopropane-1-Carboxylate Synthase Transcription via WRKY33 and 1-Aminocyclopropane-1-Carboxylate Oxidase by Modulating Messenger RNA Stability to Induce Ethylene Synthesis during Stress1[OPEN

    PubMed Central

    Kumar, Deepak; Hazra, Saptarshi; Chattopadhyay, Sharmila

    2015-01-01

    Glutathione (GSH) plays a fundamental role in plant defense-signaling network. Recently, we have established the involvement of GSH with ethylene (ET) to combat environmental stress. However, the mechanism of GSH-ET interplay still remains unexplored. Here, we demonstrate that GSH induces ET biosynthesis by modulating the transcriptional and posttranscriptional regulations of its key enzymes, 1-aminocyclopropane-1-carboxylate synthase (ACS) and 1-aminocyclopropane-1-carboxylate oxidase (ACO). Transgenic Arabidopsis (Arabidopsis thaliana) plants with enhanced GSH content (AtECS) exhibited remarkable up-regulation of ACS2, ACS6, and ACO1 at transcript as well as protein levels, while they were down-regulated in the GSH-depleted phytoalexin deficient2-1 (pad2-1) mutant. We further observed that GSH induced ACS2 and ACS6 transcription in a WRKY33-dependent manner, while ACO1 transcription remained unaffected. On the other hand, the messenger RNA stability for ACO1 was found to be increased by GSH, which explains our above observations. In addition, we also identified the ACO1 protein to be a subject for S-glutathionylation, which is consistent with our in silico data. However, S-glutathionylation of ACS2 and ACS6 proteins was not detected. Further, the AtECS plants exhibited resistance to necrotrophic infection and salt stress, while the pad2-1 mutant was sensitive. Exogenously applied GSH could improve stress tolerance in wild-type plants but not in the ET-signaling mutant ethylene insensitive2-1, indicating that GSH-mediated resistance to these stresses occurs via an ET-mediated pathway. Together, our investigation reveals a dual-level regulation of ET biosynthesis by GSH during stress. PMID:26463088

  18. Silicon-mediated changes in polyamine and 1-aminocyclopropane-1-carboxylic acid are involved in silicon-induced drought resistance in Sorghum bicolor L.

    PubMed

    Yin, Lina; Wang, Shiwen; Liu, Peng; Wang, Wenhua; Cao, Dan; Deng, Xiping; Zhang, Suiqi

    2014-07-01

    The fact that silicon application alleviates drought stress has been widely reported, but the mechanism it underlying remains unclear. Here, morphologic and physiological changes were investigated in sorghum (Sorghum bicolor L.) seedlings treated with silicon and exposed to PEG-simulated drought stress for seven days. Drought stress dramatically decreased growth parameters (biomass, root/shoot ratio, leaf area, chlorophyll concentration and photosynthetic rate), while silicon application reduced the drought-induced decreases in those parameters. Leaf relative water content and transpiration rate were maintained at high levels compared to those in seedlings without silicon. The soluble sugar contents were increased, but the proline contents and the osmotic potential were decreased, showing that osmotic adjustment did not contribute to the silicon induced-drought resistance. Furthermore, levels of both free and conjugated polyamines (PAs) levels, including putrescine, spermidine and spermine, were all found to be increased by silicon under drought stress both in leaf and root. Meanwhile, 1-aminocyclopropane-1-carboxylic acid (ACC), the precursor of ethylene, was markedly decreased by supplemental silicon. Several key PA synthesis genes were upregulated by silicon under drought stress. These results suggest that silicon improves sorghum drought resistance by mediating the balance of PAs and ethylene levels. In leaf, the increased PAs and decreased ACC help to retard leaf senescence. In root, the balance between PAs and ACC participates in the modulation of root plasticity, increases the root/shoot ratio, and contributes to an increase in water uptake. These results suggest that silicon increases drought resistance through regulating several important physiological processes in plants.

  19. Dissecting the role of climacteric ethylene in kiwifruit (Actinidia chinensis) ripening using a 1-aminocyclopropane-1-carboxylic acid oxidase knockdown line.

    PubMed

    Atkinson, Ross G; Gunaseelan, Kularajathevan; Wang, Mindy Y; Luo, Luke; Wang, Tianchi; Norling, Cara L; Johnston, Sarah L; Maddumage, Ratnasiri; Schröder, Roswitha; Schaffer, Robert J

    2011-07-01

    During climacteric fruit ripening, autocatalytic (Type II) ethylene production initiates a transcriptional cascade that controls the production of many important fruit quality traits including flavour production and softening. The last step in ethylene biosynthesis is the conversion of 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene by the enzyme ACC oxidase (ACO). Ten independent kiwifruit (Actinidia chinensis) lines were generated targeting suppression of fruit ripening-related ACO genes and the fruit from one of these lines (TK2) did not produce detectable levels of climacteric ethylene. Ripening behaviour in a population of kiwifruit at harvest is asynchronous, so a short burst of exogenous ethylene was used to synchronize ripening in TK2 and control fruit. Following such a treatment, TK2 and control fruit softened to an 'eating-ripe' firmness. Control fruit produced climacteric ethylene and softened beyond eating-ripe by 5 d. In contrast, TK2 fruit maintained an eating-ripe firmness for >25 d and total volatile production was dramatically reduced. Application of continuous exogenous ethylene to the ripening-arrested TK2 fruit re-initiated fruit softening and typical ripe fruit volatiles were detected. A 17 500 gene microarray identified 401 genes that changed after ethylene treatment, including a polygalacturonase and a pectate lyase involved in cell wall breakdown, and a quinone oxidoreductase potentially involved in volatile production. Many of the gene changes were consistent with the softening and flavour changes observed after ethylene treatment. However, a surprisingly large number of genes of unknown function were also observed, which could account for the unique flavour and textural properties of ripe kiwifruit.

  20. Pear ACO genes encoding putative 1-aminocyclopropane-1-carboxylate oxidase homologs are functionally expressed during fruit ripening and involved in response to salicylic acid.

    PubMed

    Shi, Hai-Yan; Zhang, Yu-Xing

    2012-10-01

    1-Aminocyclopropane-1-carboxylate (ACC) oxidase catalyzes the final reaction of the ethylene biosynthetic pathway, converting ACC into ethylene. Past studies have shown a possible link between ACC oxidase and salicylic acid during fruit ripening in pear, but the relationship has received no more than modest study at the gene expression level. In this study, two cDNA clones encoding putative ACC oxidase, PpACO1 and PpACO2, were isolated from a cDNA library constructed by our own laboratory and produced using mRNA from mesocarp of pear (Pyrus pyrifolia Nakai. cv.Whangkeumbae). One cDNA clone, designated PpACO1 (GenBank accession No. JN807390), comprised an open reading frame of 945 bp encoding a protein of 314 amino acids. The other cDNA, designated PpACO2 (GenBank accession No. JN807392), encodes a protein with 322 amino acids that shares high similarity with the known plant ACOs. Using PCR amplification techniques, two genomic clones corresponding to PpACO1 and PpACO2 were isolated and shown to contain independently three introns with typical GT/AG boundaries defining the splice junctions. The PpACO1 gene product shared 99 % identity with an ACC oxidase from pear (Pyrus × bretschneideri Rehd.cv.Yali), and phylogenetic analyses clearly placed the gene product in the ACC oxidase cluster of the pear 2-oxoglutarate-dependent dioxygenase superfamily tree. Quantitative RT-PCR analysis indicated that the two PpACO genes are differentially expressed in pear tissues. PpACO1 and PpACO2 were predominantly expressed in fruit. The transcripts of PpACO1 were accumulated at relatively low levels in early fruit, but strongly high levels in fruit ripening and senescence stages, while the transcripts of PpACO2 were accumulated at higher levels in early fruit and much lower levels with further fruit cell development than the transcripts of PpACO1. In addition, PpACO1 gene was down-regulated in fruit by salicylic acid (SA). Nevertheless, PpACO2 gene was dramatically up-regulated in

  1. Studies on Plant Growth Promoting Properties of Fruit-Associated Bacteria from Elettaria cardamomum and Molecular Analysis of ACC Deaminase Gene.

    PubMed

    Jasim, B; Anish, Mathew Chacko; Shimil, Vellakudiyan; Jyothis, Mathew; Radhakrishnan, E K

    2015-09-01

    Endophytic microorganisms have been reported to have diverse plant growth promoting mechanisms including phosphate solubilization, N2 fixation, production of phyto-hormones and ACC (1-aminocyclopropane-1-carboxylate) deaminase and antiphyto-pathogenic properties. Among these, ACC deaminase production is very important because of its regulatory effect on ethylene which is a stress hormone with precise role in the control of fruit development and ripening. However, distribution of these properties among various endophytic bacteria associated with fruit tissue and its genetic basis is least investigated. In the current study, 11 endophytic bacteria were isolated and identified from the fruit tissue of Elettaria cardamomum and were studied in detail for various plant growth promoting properties especially ACC deaminase activity using both culture-based and PCR-based methods. PCR-based screening identified the isolates EcB 2 (Pantoea sp.), EcB 7 (Polaromonas sp.), EcB 9 (Pseudomonas sp.), EcB 10 (Pseudomonas sp.) and EcB 11 (Ralstonia sp.) as positive for ACC deaminase. The PCR products were further subjected to sequence analysis which proved the similarity of the sequences identified in the study with ACC deaminase sequences reported from other sources. The detailed bioinformatic analysis of the sequence including homology-based modelling and molecular docking confirmed the sequences to have ACC deaminase activity. The docking of the modelled proteins was done using patch dock, and the detailed scrutiny of the protein ligand interaction revealed conservation of key amino acids like Lys51, Ser78, Tyr268 and Tyr294 which play important role in the enzyme activity. These suggest the possible regulatory effect of these isolates on fruit physiology.

  2. The rhizobacterium Variovorax paradoxus 5C-2, containing ACC deaminase, promotes growth and development of Arabidopsis thaliana via an ethylene-dependent pathway.

    PubMed

    Chen, Lin; Dodd, Ian C; Theobald, Julian C; Belimov, Andrey A; Davies, William J

    2013-04-01

    Many plant-growth-promoting rhizobacteria (PGPR) associated with plant roots contain the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase and can metabolize ACC, the immediate precursor of the plant hormone ethylene, thereby decreasing plant ethylene production and increasing plant growth. However, relatively few studies have explicitly linked ethylene emission and/or action to growth promotion in these plant-microbe interactions. This study examined effects of the PGPR Variovorax paradoxus 5C-2 containing ACC deaminase on the growth and development of Arabidopsis thaliana using wild-type (WT) plants and several ethylene-related mutants (etr1-1, ein2-1, and eto1-1). Soil inoculation with V. paradoxus 5C-2 promoted growth (leaf area and shoot biomass) of WT plants and the ethylene-overproducing mutant eto1-1, and also enhanced floral initiation of WT plants by 2.5 days. However, these effects were not seen in ethylene-insensitive mutants (etr1-1 and ein2-1) even though bacterial colonization of the root system was similar. Furthermore, V. paradoxus 5C-2 decreased ACC concentrations of rosette leaves of WT plants by 59% and foliar ethylene emission of both WT plants and eto1-1 mutants by 42 and 37%, respectively. Taken together, these results demonstrate that a fully functional ethylene signal transduction pathway is required for V. paradoxus 5C-2 to stimulate leaf growth and flowering of A. thaliana.

  3. Methyl jasmonate-induced defense responses are associated with elevation of 1-aminocyclopropane-1-carboxylate oxidase in Lycopersicon esculentum fruit.

    PubMed

    Yu, Mengmeng; Shen, Lin; Zhang, Aijun; Sheng, Jiping

    2011-10-15

    It has been known that methyl jasmonate (MeJA) interacts with ethylene to elicit resistance. In green mature tomato fruits (Lycopersicon esculentum cv. Lichun), 0.02mM MeJA increased the activity of 1-aminocyclopropane-1-carboxylate oxidase (ACO), and consequently influenced the last step of ethylene biosynthesis. Fruits treated with a combination of 0.02 MeJA and 0.02 α-aminoisobutyric acid (AIB, a competitive inhibitor of ACO) exhibited a lower ethylene production comparing to that by 0.02mM MeJA alone. The increased activities of defense enzymes and subsequent control of disease incidence caused by Botrytis cinerea with 0.2mM MeJA treatment was impaired by AIB as well. A close relationship (P<0.05) was found between the activity alterations of ACO and that of chitinase (CHI) and β-1,3-glucanase (GLU). In addition, this study further detected the changes of gene expressions and enzyme kinetics of ACO to different concentrations of MeJA. LeACO1 was found the principal member from the ACO gene family to respond to MeJA. Accumulation of LeACO1/3/4 transcripts followed the concentration pattern of MeJA treatments, where the largest elevations were reached by 0.2mM. For kinetic analysis, K(m) values of ACO stepped up during the experiment and reached the maximums at 0.2mM MeJA with ascending concentrations of treatments. V(max) exhibited a gradual increase from 3h to 24h, and the largest induction appeared with 1.0mM MeJA. The results suggested that ACO is involved in MeJA-induced resistance in tomato, and the concentration influence of MeJA on ACO was attributable to the variation of gene transcripts and enzymatic properties.

  4. Characterization of ACC deaminase-producing endophytic bacteria isolated from copper-tolerant plants and their potential in promoting the growth and copper accumulation of Brassica napus.

    PubMed

    Zhang, Yan-Feng; He, Lin-Yan; Chen, Zhao-Jin; Wang, Qing-Ya; Qian, Meng; Sheng, Xia-Fang

    2011-03-01

    One hundred Cu-resistant-endophytic bacteria were isolated from Cu-tolerant plants grown on Cu mine wasteland, of which, eight Cu-resistant and 1-aminocyclopropane-1-carboxylate (ACC) deaminase-producing endophytic bacteria were obtained based on the ACC deaminase activity of the bacteria and characterized with respect to metal resistance, production of ACC deaminase, indole-3-acetic acid (IAA) as well as siderophores and mineral phosphate solubilization. Ralstonia sp. J1-22-2, Pantoea agglomerans Jp3-3, and Pseudomonas thivervalensis Y1-3-9 with higher ACC deaminase activity (ranging from 213 to 370 μM α-ketobutyrate mg(-1)h(-1)) were evaluated for promoting plant growth and Cu uptake of rape grown in quartz sand containing 0, 2.5, and 5 mg kg(-1) of Cu in pot experiments. The eight bacteria were found to exhibit different multiple heavy metal resistance characteristics, to show different levels of ACC deaminase activity and to produce indole acetic acid. Seven bacteria produced siderophores and solubilized inorganic phosphate. Pot experiments showed that inoculation with the strains (J1-22-2, Jp3-3, and Y1-3-9) was found to increase the biomass of rape. Increases in above-ground tissue Cu contents of rape cultivated in 2.5 and 5 mg kg(-1) of Cu-contaminated substrates varied from 9% to 31% and from 3 to 4-fold respectively in inoculated-rape plants compared to the uninoculated control. The maximum Cu uptake of rape was observed after inoculation with P. agglomerans Jp3-3. The results show that metal-resistant and plant growth promoting endophytic bacteria play an important role in plant growth and Cu uptake which may provide a new endophytic bacterial-assisted phytoremediation of Cu-contaminated environment.

  5. Bacterial community compositions of tomato (Lycopersicum esculentum Mill.) seeds and plant growth promoting activity of ACC deaminase producing Bacillus subtilis (HYT-12-1) on tomato seedlings.

    PubMed

    Xu, Mingshuang; Sheng, Jiping; Chen, Lin; Men, Yejun; Gan, Lin; Guo, Shuntang; Shen, Lin

    2014-03-01

    Study of endophytic bacteria within plant seeds is very essential and meaningful on account of their heritability and versatility. This study investigated Bacillus bacterial communities within the seeds of four commercial tomato varieties, by 16S rRNA gene PCR-RFLP (restriction fragment length polymorphism). Phylogenetic analysis of 16S rRNA gene sequences indicated that the 22 representative isolates belonged to five species of genus Bacillus and the bacterial compositions showed remarkable differences among tomato varieties. Isolates exhibited multiple plant growth promoting (PGP) traits: 37 % of indole-3-acetic acid production; 37 % of phosphate solubilization; 24 % of siderophores production; 85 % of potential nitrogen fixation and 6 % of 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. Isolate HYT-12-1 was shown to have highest ACC deaminase activity (112.02 nmol α-ketobutyrate mg⁻¹ protein h⁻¹) among the five ACC deamiase producing strains. 16S rRNA gene sequencing indicated that the isolate HYT-12-1 shared the highest sequence similarity (100 %) with B. subtilis. PGP experiments under gnotobiotic and greenhouse conditions revealed the ability of strain HYT-12-1 to enhance the growth of tomato seedlings. This is the first study to describe endophytic Bacillus communities within tomato seeds, and the results suggest that B. subtilis strain HYT-12-1 would have a great potential for industrial application as biofertilizer in the future.

  6. 1-Aminocyclopropane-1-Carboxylate Oxidase Induction in Tomato Flower Pedicel Phloem and Abscission Related Processes Are Differentially Sensitive to Ethylene

    PubMed Central

    Chersicola, Marko; Kladnik, Aleš; Tušek Žnidarič, Magda; Mrak, Tanja; Gruden, Kristina; Dermastia, Marina

    2017-01-01

    Ethylene has impact on several physiological plant processes, including abscission, during which plants shed both their vegetative and reproductive organs. Cell separation and programmed cell death are involved in abscission, and these have also been correlated with ethylene action. However, the detailed spatiotemporal pattern of the molecular events during abscission remains unknown. We examined the expression of two tomato ACO genes, LeACO1, and LeACO4 that encode the last enzyme in ethylene biosynthesis, 1-aminocyclopropane-1-carboxylate oxidase (ACO), together with the expression of other abscission-associated genes involved in cell separation and programmed cell death, during a period of 0–12 h after abscission induction in the tomato flower pedicel abscission zone and nearby tissues. In addition, we determined their localization in specific cell layers of the flower pedicel abscission zone and nearby tissues obtained by laser microdissection before and 8 h after abscission induction. The expression of both ACO genes was localized to the vascular tissues in the pedicel. While LeACO4 was more uniformly expressed in all examined cell layers, the main expression site of LeACO1 was in cell layers just outside the abscission zone in its proximal and distal part. We showed that after abscission induction, ACO1 protein was synthesized in phloem companion cells, in which it was localized mainly in the cytoplasm. Samples were additionally treated with 1-methylcyclopropene (1-MCP), a competitive inhibitor of ethylene actions, and analyzed 8 h after abscission induction. Cell-layer-specific changes in gene expression were observed together with the specific localization and ethylene sensitivity of the hallmarks of cell separation and programmed cell death. While treatment with 1-MCP prevented separation of cells through inhibition of the expression of polygalacturonases, which are the key enzymes involved in degradation of the middle lamella, this had less impact on

  7. Expression and regulation of pear 1-aminocyclopropane-1-carboxylic acid synthase gene (PpACS1a) during fruit ripening, under salicylic acid and indole-3-acetic acid treatment, and in diseased fruit.

    PubMed

    Shi, Hai-Yan; Zhang, Yu-Xing

    2014-06-01

    In plants, the level of ethylene is determined by the activity of the key enzyme 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (ACS). A gene encoding an ACC synthase protein was isolated from pear (Pyrus pyrifolia). This gene designated PpACS1a (GenBank accession no. KC632526) was 1488 bp in length with an open reading frame (ORF) encoding a protein of 495 amino acids that shared high similarity with other pear ACC synthase proteins. The PpACS1a was grouped into type-1 subfamily of plant ACS based on its conserved domain and phylogenetic status. Real-time quantitative PCR indicated that PpACS1a was differentially expressed in pear tissues and predominantly expressed in anthers. The expression signal of PpACS1a was also detected in fruit and leaves, but no signal was detected in shoots and petals. Furthermore, the PpACS1a expression was regulated during fruit ripening. In addition, the PpACS1a gene expression was regulated by salicylic acid (SA) and indole-3-acetic acid (IAA) in fruit. Moreover, the expression of the PpACS1a was up-regulated in diseased pear fruit. These results indicated that PpACS1a might be involved in fruit ripening and response to SA, IAA and disease.

  8. Ethylene emission and PR protein synthesis in ACC deaminase producing Methylobacterium spp. inoculated tomato plants (Lycopersicon esculentum Mill.) challenged with Ralstonia solanacearum under greenhouse conditions.

    PubMed

    Yim, Woojong; Seshadri, Sundaram; Kim, Kiyoon; Lee, Gillseung; Sa, Tongmin

    2013-06-01

    Bacteria of genus Methylobacterium have been found to promote plant growth and regulate the level of ethylene in crop plants. This work is aimed to test the induction of defense responses in tomato against bacterial wilt by stress ethylene level reduction mediated by the ACC deaminase activity of Methylobacterium strains. Under greenhouse conditions, the disease index value in Methylobacterium sp. inoculated tomato plants was lower than control plants. Plants treated with Methylobacterium sp. challenge inoculated with Ralstonia solanacearum (RS) showed significantly reduced disease symptoms and lowered ethylene emission under greenhouse condition. The ACC and ACO (1-aminocyclopropane-1-carboxylate oxidase) accumulation in tomato leaves were significantly reduced with Methylobacterium strains inoculation. While ACC oxidase gene expression was found higher in plants treated with R. solanacearum than Methylobacterium sp. treatment, PR proteins related to induced systemic resistance like β-1,3-glucanase, PAL, PO and PPO were increased in Methylobacterium sp. inoculated plants. A significant increase in β-1,3-glucanase and PAL gene expression was found in all the Methylobacterium spp. treatments compared to the R. solanacearum treatment. This study confirms the activity of Methylobacterium sp. in increasing the defense enzymes by modulating the ethylene biosynthesis pathway and suggests the use of methylotrophic bacteria as potential biocontrol agents in tomato cultivation.

  9. RNA interference of 1-aminocyclopropane-1-carboxylic acid oxidase (ACO1 and ACO2) genes expression prolongs the shelf life of Eksotika (Carica papaya L.) papaya fruit.

    PubMed

    Sekeli, Rogayah; Abdullah, Janna Ong; Namasivayam, Parameswari; Muda, Pauziah; Abu Bakar, Umi Kalsom; Yeong, Wee Chien; Pillai, Vilasini

    2014-06-19

    The purpose of this study was to evaluate the effectiveness of using RNA interference in down regulating the expression of 1-aminocyclopropane-1-carboxylic acid oxidase gene in Eksotika papaya. One-month old embryogenic calli were separately transformed with Agrobacterium strain LBA 4404 harbouring the three different RNAi pOpOff2 constructs bearing the 1-aminocyclopropane-1-carboxylic acid oxidase gene. A total of 176 putative transformed lines were produced from 15,000 calli transformed, selected, then regenerated on medium supplemented with kanamycin. Integration and expression of the targeted gene in putatively transformed lines were verified by PCR and real-time RT-PCR. Confined field evaluation of a total of 31 putative transgenic lines planted showed a knockdown expression of the targeted ACO1 and ACO2 genes in 13 lines, which required more than 8 days to achieve the full yellow colour (Index 6). Fruits harvested from lines pRNAiACO2 L2-9 and pRNAiACO1 L2 exhibited about 20 and 14 days extended post-harvest shelf life to reach Index 6, respectively. The total soluble solids contents of the fruits ranged from 11 to 14° Brix, a range similar to fruits from non-transformed, wild type seed-derived plants.

  10. Effect of the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid on different growth stages of Haematococcus pluvialis.

    PubMed

    Vo, Thi-Thao; Lee, Changsu; Han, Sang-Il; Kim, Jee Young; Kim, Sok; Choi, Yoon-E

    2016-11-01

    In this study, we explored the effects of ACC on other stages of H. pluvialis. Interestingly, even though ACC displayed a dose-dependent effect on astaxanthin production, it is evident that astaxanthin production could be facilitated whenever the cells were treated at the early red stage. The transcriptional levels of BKT, CHY, SOD, and CAT genes supported enhanced astaxanthin biosynthesis upon ACC treatment at the early red stage. The combinatorial synergistic effect of ACC and light intensity was also confirmed. Finally, two-step application of ACC at the vegetative phase to increase biomass production and at the early-red stage to promote astaxanthin biosynthesis was proposed to maximize the efficiency of ACC treatment.

  11. Isolation, characterization, and use for plant growth promotion under salt stress, of ACC deaminase-producing halotolerant bacteria derived from coastal soil.

    PubMed

    Siddikee, Md Ashaduzzaman; Chauhan, Puneet S; Anandham, R; Han, Gwang-Hyun; Sa, Tongmin

    2010-11-01

    In total, 140 halotolerant bacterial strains were isolated from both the soil of barren fields and the rhizosphere of six naturally growing halophytic plants in the vicinity of the Yellow Sea, near the city of Incheon in the Republic of Korea. All of these strains were characterized for multiple plant growth promoting traits, such as the production of indole acetic acid (IAA), nitrogen fixation, phosphorus (P) and zinc (Zn) solubilization, thiosulfate (S2O3) oxidation, the production of ammonia (NH3), and the production of extracellular hydrolytic enzymes such as protease, chitinase, pectinase, cellulase, and lipase under in vitro conditions. From the original 140 strains tested, on the basis of the latter tests for plant growth promotional activity, 36 were selected for further examination. These 36 halotolerant bacterial strains were then tested for 1- aminocyclopropane-1-carboxylic acid (ACC) deaminase activity. Twenty-five of these were found to be positive, and to be exhibiting significantly varying levels of activity. 16S rRNA gene sequencing analyses of the 36 halotolerant strains showed that they belong to 10 different bacterial genera: Bacillus, Brevibacterium, Planococcus, Zhihengliuella, Halomonas, Exiguobacterium, Oceanimonas, Corynebacterium, Arthrobacter, and Micrococcus. Inoculation of the 14 halotolerant bacterial strains to ameliorate salt stress (150 mM NaCl) in canola plants produced an increase in root length of between 5.2% and 47.8%, and dry weight of between 16.2% and 43%, in comparison with the uninoculated positive controls. In particular, three of the bacteria, Brevibacterium epidermidis RS15, Micrococcus yunnanensis RS222, and Bacillus aryabhattai RS341, all showed more than 40% increase in root elongation and dry weight when compared with uninoculated saltstressed canola seedlings. These results indicate that certain halotolerant bacteria, isolated from coastal soils, have a real potential to enhance plant growth under saline stress

  12. Kinetic and mutagenic evidence for the role of histidine residues in the Lycopersicon esculentum 1-aminocyclopropane-1-carboxylic acid oxidase.

    PubMed

    Tayeh, M A; Howe, D L; Salleh, H M; Sheflyan, G Y; Son, J K; Woodard, R W

    1999-01-01

    The ACCO gene from Lycopersicon esculentum (tomato) has been cloned into the expression vector PT7-7. The highly expressed protein was recovered in the form of inclusion bodies. ACCO is inactivated by diethyl pyrocarbonate (DEPC) with a second-order rate constant of 170 M(-1) min(-1). The pH-inactivation rate data imply the involvement of an amino acid residue with a pK value of 6.05. The difference UV spectrum of the the DEPC-inactivated versus native ACCO showed a single peak at 242 nm indicating the modification of histidine residues. The inactivation was reversed by the addition of hydroxylamine to the DEPC-inactivated ACCO. Substrate/cofactor protection studies indicate that both iron and ACC bind near the active site, which contains histidine residues. Four histidines of ACCO were individually mutated to alanine and glycine. H39A is catalytically active, while H177A, H177G, H211A, H211G, H234A, and H234G are basically inactive. The results indicate that histidine residues 177, 211, and 234 may serve as ligands for the active-site iron of ACCO and/or may play some important structural or catalytic role.

  13. Gibberellic acid, synthetic auxins, and ethylene differentially modulate alpha-L-Arabinofuranosidase activities in antisense 1-aminocyclopropane-1-carboxylic acid synthase tomato pericarp discs.

    PubMed

    Sozzi, Gabriel O; Greve, L Carl; Prody, Gerry A; Labavitch, John M

    2002-07-01

    Alpha-L-Arabinofuranosidases (alpha-Afs) are plant enzymes capable of releasing terminal arabinofuranosyl residues from cell wall matrix polymers, as well as from different glycoconjugates. Three different alpha-Af isoforms were distinguished by size exclusion chromatography of protein extracts from control tomatoes (Lycopersicon esculentum) and an ethylene synthesis-suppressed (ESS) line expressing an antisense 1-aminocyclopropane-1-carboxylic synthase transgene. alpha-Af I and II are active throughout fruit ontogeny. alpha-Af I is the first Zn-dependent cell wall enzyme isolated from tomato pericarp tissues, thus suggesting the involvement of zinc in fruit cell wall metabolism. This isoform is inhibited by 1,10-phenanthroline, but remains stable in the presence of NaCl and sucrose. alpha-Af II activity accounts for over 80% of the total alpha-Af activity in 10-d-old fruit, but activity drops during ripening. In contrast, alpha-Af III is ethylene dependent and specifically active during ripening. alpha-Af I released monosaccharide arabinose from KOH-soluble polysaccharides from tomato cell walls, whereas alpha-Af II and III acted on Na(2)CO(3)-soluble pectins. Different alpha-Af isoform responses to gibberellic acid, synthetic auxins, and ethylene were followed by using a novel ESS mature-green tomato pericarp disc system. alpha-Af I and II activity increased when gibberellic acid or 2,4-dichlorophenoxyacetic acid was applied, whereas ethylene treatment enhanced only alpha-Af III activity. Results suggest that tomato alpha-Afs are encoded by a gene family under differential hormonal controls, and probably have different in vivo functions. The ESS pericarp explant system allows comprehensive studies involving effects of physiological levels of different growth regulators on gene expression and enzyme activity with negligible wound-induced ethylene production.

  14. Gibberellic Acid, Synthetic Auxins, and Ethylene Differentially Modulate α-l-Arabinofuranosidase Activities in Antisense 1-Aminocyclopropane-1-Carboxylic Acid Synthase Tomato Pericarp Discs1

    PubMed Central

    Sozzi, Gabriel O.; Greve, L. Carl; Prody, Gerry A.; Labavitch, John M.

    2002-01-01

    α-l-Arabinofuranosidases (α-Afs) are plant enzymes capable of releasing terminal arabinofuranosyl residues from cell wall matrix polymers, as well as from different glycoconjugates. Three different α-Af isoforms were distinguished by size exclusion chromatography of protein extracts from control tomatoes (Lycopersicon esculentum) and an ethylene synthesis-suppressed (ESS) line expressing an antisense 1-aminocyclopropane-1-carboxylic synthase transgene. α-Af I and II are active throughout fruit ontogeny. α-Af I is the first Zn-dependent cell wall enzyme isolated from tomato pericarp tissues, thus suggesting the involvement of zinc in fruit cell wall metabolism. This isoform is inhibited by 1,10-phenanthroline, but remains stable in the presence of NaCl and sucrose. α-Af II activity accounts for over 80% of the total α-Af activity in 10-d-old fruit, but activity drops during ripening. In contrast, α-Af III is ethylene dependent and specifically active during ripening. α-Af I released monosaccharide arabinose from KOH-soluble polysaccharides from tomato cell walls, whereas α-Af II and III acted on Na2CO3-soluble pectins. Different α-Af isoform responses to gibberellic acid, synthetic auxins, and ethylene were followed by using a novel ESS mature-green tomato pericarp disc system. α-Af I and II activity increased when gibberellic acid or 2,4-dichlorophenoxyacetic acid was applied, whereas ethylene treatment enhanced only α-Af III activity. Results suggest that tomato α-Afs are encoded by a gene family under differential hormonal controls, and probably have different in vivo functions. The ESS pericarp explant system allows comprehensive studies involving effects of physiological levels of different growth regulators on gene expression and enzyme activity with negligible wound-induced ethylene production. PMID:12114586

  15. A Ser/Thr protein kinase phosphorylates MA-ACS1 (Musa acuminata 1-aminocyclopropane-1-carboxylic acid synthase 1) during banana fruit ripening.

    PubMed

    Choudhury, Swarup Roy; Roy, Sujit; Sengupta, Dibyendu N

    2012-08-01

    1-Aminocyclopropane-1-carboxylic acid synthase (ACS) catalyzes the rate-limiting step in ethylene biosynthesis during ripening. ACS isozymes are regulated both transcriptionally and post-translationally. However, in banana, an important climacteric fruit, little is known about post-translational regulation of ACS. Here, we report the post-translational modification of MA-ACS1 (Musa acuminata ACS1), a ripening inducible isozyme in the ACS family, which plays a key role in ethylene biosynthesis during banana fruit ripening. Immunoprecipitation analyses of phospholabeled protein extracts from banana fruit using affinity-purified anti-MA-ACS1 antibody have revealed phosphorylation of MA-ACS1, particularly in ripe fruit tissue. We have identified the induction of a 41-kDa protein kinase activity in pulp at the onset of ripening. The 41-kDa protein kinase has been identified as a putative protein kinase by MALDI-TOF/MS analysis. Biochemical analyses using partially purified protein kinase fraction from banana fruit have identified the protein kinase as a Ser/Thr family of protein kinase and its possible involvement in MA-ACS1 phosphorylation during ripening. In vitro phosphorylation analyses using synthetic peptides and site-directed mutagenized recombinant MA-ACS1 have revealed that serine 476 and 479 residues at the C-terminal region of MA-ACS1 are phosphorylated. Overall, this study provides important novel evidence for in vivo phosphorylation of MA-ACS1 at the molecular level as a possible mechanism of post-translational regulation of this key regulatory protein in ethylene signaling pathway in banana fruit during ripening.

  16. Role and Regulation of ACC Deaminase Gene in Sinorhizobium meliloti: Is It a Symbiotic, Rhizospheric or Endophytic Gene?

    PubMed Central

    Checcucci, Alice; Azzarello, Elisa; Bazzicalupo, Marco; De Carlo, Anna; Emiliani, Giovanni; Mancuso, Stefano; Spini, Giulia; Viti, Carlo; Mengoni, Alessio

    2017-01-01

    Plant-associated bacteria exhibit a number of different strategies and specific genes allow bacteria to communicate and metabolically interact with plant tissues. Among the genes found in the genomes of plant-associated bacteria, the gene encoding the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase (acdS) is one of the most diffused. This gene is supposed to be involved in the cleaving of plant-produced ACC, the precursor of the plant stress-hormone ethylene toning down the plant response to infection. However, few reports are present on the actual role in rhizobia, one of the most investigated groups of plant-associated bacteria. In particular, still unclear is the origin and the role of acdS in symbiotic competitiveness and on the selective benefit it may confer to plant symbiotic rhizobia. Here we present a phylogenetic and functional analysis of acdS orthologs in the rhizobium model-species Sinorhizobium meliloti. Results showed that acdS orthologs present in S. meliloti pangenome have polyphyletic origin and likely spread through horizontal gene transfer, mediated by mobile genetic elements. When acdS ortholog from AK83 strain was cloned and assayed in S. meliloti 1021 (lacking acdS), no modulation of plant ethylene levels was detected, as well as no increase in fitness for nodule occupancy was found in the acdS-derivative strain compared to the parental one. Surprisingly, AcdS was shown to confer the ability to utilize formamide and some dipeptides as sole nitrogen source. Finally, acdS was shown to be negatively regulated by a putative leucine-responsive regulator (LrpL) located upstream to acdS sequence (acdR). acdS expression was induced by root exudates of both legumes and non-leguminous plants. We conclude that acdS in S. meliloti is not directly related to symbiotic interaction, but it could likely be involved in the rhizospheric colonization or in the endophytic behavior. PMID:28194158

  17. Role and Regulation of ACC Deaminase Gene in Sinorhizobium meliloti: Is It a Symbiotic, Rhizospheric or Endophytic Gene?

    PubMed

    Checcucci, Alice; Azzarello, Elisa; Bazzicalupo, Marco; De Carlo, Anna; Emiliani, Giovanni; Mancuso, Stefano; Spini, Giulia; Viti, Carlo; Mengoni, Alessio

    2017-01-01

    Plant-associated bacteria exhibit a number of different strategies and specific genes allow bacteria to communicate and metabolically interact with plant tissues. Among the genes found in the genomes of plant-associated bacteria, the gene encoding the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase (acdS) is one of the most diffused. This gene is supposed to be involved in the cleaving of plant-produced ACC, the precursor of the plant stress-hormone ethylene toning down the plant response to infection. However, few reports are present on the actual role in rhizobia, one of the most investigated groups of plant-associated bacteria. In particular, still unclear is the origin and the role of acdS in symbiotic competitiveness and on the selective benefit it may confer to plant symbiotic rhizobia. Here we present a phylogenetic and functional analysis of acdS orthologs in the rhizobium model-species Sinorhizobium meliloti. Results showed that acdS orthologs present in S. meliloti pangenome have polyphyletic origin and likely spread through horizontal gene transfer, mediated by mobile genetic elements. When acdS ortholog from AK83 strain was cloned and assayed in S. meliloti 1021 (lacking acdS), no modulation of plant ethylene levels was detected, as well as no increase in fitness for nodule occupancy was found in the acdS-derivative strain compared to the parental one. Surprisingly, AcdS was shown to confer the ability to utilize formamide and some dipeptides as sole nitrogen source. Finally, acdS was shown to be negatively regulated by a putative leucine-responsive regulator (LrpL) located upstream to acdS sequence (acdR). acdS expression was induced by root exudates of both legumes and non-leguminous plants. We conclude that acdS in S. meliloti is not directly related to symbiotic interaction, but it could likely be involved in the rhizospheric colonization or in the endophytic behavior.

  18. Cloning and expression of 1-aminocyclopropane-1-carboxylate oxidase cDNA induced by thidiazuron during somatic embryogenesis of alfalfa (Medicago sativa).

    PubMed

    Feng, Bi-Hong; Wu, Bei; Zhang, Chun-Rong; Huang, Xia; Chen, Yun-Feng; Huang, Xue-Lin

    2012-01-15

    Embryogenic callus (EC) induced from petioles of alfalfa (Medicago sativa L. cv. Jinnan) on B5h medium turned green, compact and non-embryogenic when the kinetin (KN) in the medium was replaced partially or completely by thidiazuron (TDZ). The application of CoCl₂, which is an inhibitor of 1-aminocyclopropane-1-carboxylate oxidase (ACO), counteracted the effect of TDZ. Ethylene has been shown to be involved in the modulation of TDZ-induced morphogenesis responses. However, very little is known about the genes involved in ethylene formation during somatic embryogenesis (SE). To investigate whether ethylene mediated by ACO is involved in the effect of TDZ on inhibition of embryogenic competence of the alfalfa callus. In this study we cloned full-length ACO cDNA from the alfalfa callus, named MsACO, and observed changes in this gene expression during callus formation and induction of SE under treatment with TDZ or TDZ plus CoCl₂. RNA blot analysis showed that during the EC subcultural period, the expression level of MsACO in EC was significantly increased on the 2nd day, rose to the highest level on the 8th day and remained at this high level until the 21st day. However, the ACO expression in the TDZ (0.93 μM)-treated callus was higher than in the EC especially on the 8th day. Moreover the ACO expression level increased with increasing TDZ concentration during the subcultural/maintenance period of the callus. It is worth noting that comparing the treatment with TDZ alone, the treatment with 0.93 μM TDZ plus 50 μM CoCl₂ reduced both of the ACO gene expressions and ACO activity in the treated callus. These results indicate that the effect of TDZ could be counteracted by CoCl₂ either on the ACO gene expression level or ACO activity. Thus, a TDZ inhibitory effect on embryogenic competence of alfalfa callus could be mediated by ACO gene expression.

  19. Comparative effectiveness of ACC-deaminase and/or nitrogen-fixing rhizobacteria in promotion of maize (Zea mays L.) growth under lead pollution.

    PubMed

    Hassan, Waseem; Bano, Rizwana; Bashir, Farhat; David, Julie

    2014-09-01

    Lead (Pb) pollution is appearing as an alarming threat nowadays. Excessive Pb concentrations in agricultural soils result in minimizing the soil fertility and health which affects the plant growth and leads to decrease in crop production. Plant growth promoting rhizobacteria (PGPR) are beneficial bacteria which can protect the plants against many abiotic stresses, and enhance the growth. The study aimed to identify important rhizobacterial strains by using the 1-aminocyclopropane-1-carboxylate (ACC) enrichment technique and examine their inoculation effects in the growth promotion of maize, under Pb pollution. A pot experiment was conducted and six rhizobacterial isolates were used. Pb was added to 2 kg soil in each pot (with 4 seeds/pot) using Pb(NO3)2 at the rate of 0, 100, 200, 300, and 400 mg kg(-1) Pb with three replications in completely randomized design. Rhizobacterial isolates performed significantly better under all Pb levels, i.e., 100 to 400 Pb mg kg(-1) soil, compared to control. Comparing the efficacy of the rhizobacterial isolates under different Pb levels, rhizobacterial isolates having both ACC-deaminase and nitrogen-fixing activities (AN8 and AN12) showed highest increase in terms of the physical, chemical and enzymatic growth parameters of maize, followed by the rhizobacterial isolates having ACC-deaminase activity only (ACC5 and ACC8), and then the nitrogen-fixing rhizobia (Azotobacter and RN5). However, the AN8 isolate showed maximum efficiency, and highest shoot and root length (14.2 and 6.1 cm), seedling fresh and dry weights (1.91 and 0.14 g), chlorophyll a, b, and carotenoids (24.1, 30.2 and 77.7 μg/l), protein (0.82 mg/g), proline (3.42 μmol/g), glutathione S-transferase, peroxidase and catalase (12.3, 4.2 and 7.2 units/mg protein), while the lowest Pb uptake in the shoot and root (0.83 and 0.48 mg/kg) were observed under this rhizobial isolate at the highest Pb level (i.e., 400 Pb mg kg(-1) soil). The results revealed that PGPR

  20. The formation of ACC and competition between polyamines and ethylene for SAM

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ethylene biosynthesis involves the conversion of S-adenosylmethionine (SAM) to 1-aminocyclopropane-1-carboxylic acid (ACC) by ACC synthase (ACS). ACC is then converted to ethylene. The genes that encode enzymes in this pathway all belong to a family of genes. Differential transcriptional regulation ...

  1. Analysis of genomic DNA of DcACS1, a 1-aminocyclopropane-1-carboxylate synthase gene, expressed in senescing petals of carnation (Dianthus caryophyllus) and its orthologous genes in D. superbus var. longicalycinus.

    PubMed

    Harada, Taro; Murakoshi, Yuino; Torii, Yuka; Tanase, Koji; Onozaki, Takashi; Morita, Shigeto; Masumura, Takehiro; Satoh, Shigeru

    2011-04-01

    Carnation (Dianthus caryophyllus) flowers exhibit climacteric ethylene production followed by petal wilting, a senescence symptom. DcACS1, which encodes 1-aminocyclopropane-1-carboxylate synthase (ACS), is a gene involved in this phenomenon. We determined the genomic DNA structure of DcACS1 by genomic PCR. In the genome of 'Light Pink Barbara', we found two distinct nucleotide sequences: one corresponding to the gene previously shown as DcACS1, designated here as DcACS1a, and the other novel one designated as DcACS1b. It was revealed that both DcACS1a and DcACS1b have five exons and four introns. These two genes had almost identical nucleotide sequences in exons, but not in some introns and 3'-UTR. Analysis of transcript accumulation revealed that DcACS1b is expressed in senescing petals as well as DcACS1a. Genomic PCR analysis of 32 carnation cultivars showed that most cultivars have only DcACS1a and some have both DcACS1a and DcACS1b. Moreover, we found two DcACS1 orthologous genes with different nucleotide sequences from D. superbus var. longicalycinus, and designated them as DsuACS1a and DsuACS1b. Petals of D. superbus var. longicalycinus produced ethylene in response to exogenous ethylene, accompanying accumulation of DsuACS1 transcripts. These data suggest that climacteric ethylene production in flowers was genetically established before the cultivation of carnation.

  2. Assessing the effects of heavy metals in ACC deaminase and IAA production on plant growth-promoting bacteria.

    PubMed

    Carlos, Mendoza-Hernández José; Stefani, Perea-Vélez Yazmin; Janette, Arriola-Morales; Melani, Martínez-Simón Sara; Gabriela, Pérez-Osorio

    2016-01-01

    This study poses a methodology in order to simultaneously quantify ACC deaminase and IAA levels in the same culture medium. Ten bacterial strains isolated from plant rhizosphere naturally settled in mining residues were chosen. These bacterial strains were characterized as PGPB, and all of them showed at least three characteristics (indole-3 acetic acid and siderophore production, ACC deaminase enzyme activity, and inorganic phosphate solubilization). Taxonomic identification showed that the strains belong to Enterobacter, Serratia, Klebsiella, and Escherichia genera. Similarly, both the ACC deaminase enzyme activity and the IAA synthesis in the presence of Cu, As, Pb, Ni, Cd, and Mn were measured. The results showed that both the ACC deaminase enzyme activity and the IAA synthesis were higher with the Pb, As, and Cu treatments than with the Escherichia N16, Enterobacter K131, Enterobacter N9, and Serratia K120 control treatments. On the other hand, Ni, Cd, and Mn negatively affected both the ACC deaminase enzyme activity and the IAA production on every bacterium except on the Klebsiella Mc173 strain. Serratia K120 bacterium got a positive correlation between ACC deaminase and IAA in the presence of every heavy metal, and it also promoted Helianthus annuus plant growth, showing a potential use in phytoremediation systems.

  3. Real time expression of ACC oxidase and PR-protein genes mediated by Methylobacterium spp. in tomato plants challenged with Xanthomonas campestris pv. vesicatoria.

    PubMed

    Yim, W J; Kim, K Y; Lee, Y W; Sundaram, S P; Lee, Y; Sa, T M

    2014-07-15

    Biotic stress like pathogenic infection increases ethylene biosynthesis in plants and ethylene inhibitors are known to alleviate the severity of plant disease incidence. This study aimed to reduce the bacterial spot disease incidence in tomato plants caused by Xanthomonas campestris pv. vesicatoria (XCV) by modulating stress ethylene with 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity of Methylobacterium strains. Under greenhouse condition, Methylobacterium strains inoculated and pathogen challenged tomato plants had low ethylene emission compared to pathogen infected ones. ACC accumulation and ACC oxidase (ACO) activity with ACO related gene expression increased in XCV infected tomato plants over Methylobacterium strains inoculated plants. Among the Methylobacterium spp., CBMB12 resulted lowest ACO related gene expression (1.46 Normalized Fold Expression), whereas CBMB20 had high gene expression (3.42 Normalized Fold Expression) in pathogen challenged tomato. But a significant increase in ACO gene expression (7.09 Normalized Fold Expression) was observed in the bacterial pathogen infected plants. In contrast, Methylobacterium strains enhanced β-1,3-glucanase and phenylalanine ammonia-lyase (PAL) enzyme activities in pathogen challenged tomato plants. The respective increase in β-1,3-glucanase related gene expressions due to CBMB12, CBMB15, and CBMB20 strains were 66.3, 25.5 and 10.4% higher over pathogen infected plants. Similarly, PAL gene expression was high with 0.67 and 0.30 Normalized Fold Expression, in pathogen challenged tomato plants inoculated with CBMB12 and CBMB15 strains. The results suggest that ethylene is a crucial factor in bacterial spot disease incidence and that methylobacteria with ACC deaminase activity can reduce the disease severity with ultimate pathogenesis-related protein increase in tomato.

  4. A functional tomato ACC synthase expressed in Escherichia coli demonstrates suicidal inactivation by its substrate S-adenosylmethionine.

    PubMed

    Li, N; Wiesman, Z; Liu, D; Mattoo, A K

    1992-07-20

    1-Aminocyclopropane-1-carboxylate (ACC) synthase is a key enzyme in the biosynthesis of the plant hormone, ethylene. We have isolated, sequenced and expressed a functional tomato (cv Pik-Red) ACC synthase gene in Escherichia coli. ACC synthase expressed in E. coli was inactivated by incubation with S-adenosylmethionine (SAM), the half-time of which was concentration dependent. Mixing the tomato fruit protein extract with the cell-free extract from transformed E. coli did not affect SAM-dependent inactivation of ACC synthase activity. Thus, single isoforms of the ACC synthase enzyme, which demonstrate the biochemical features expected of the tomato fruit enzyme, can be expressed in E. coli and their structure-function relationships investigated.

  5. Genome Sequence of the Banana Plant Growth-Promoting Rhizobacterium Pseudomonas fluorescens PS006

    PubMed Central

    Gamez, Rocío M.; Rodríguez, Fernando; Ramírez, Sandra; Gómez, Yolanda; Agarwala, Richa; Landsman, David

    2016-01-01

    Pseudomonas fluorescens is a well-known plant growth-promoting rhizobacterium (PGPR). We report here the first whole-genome sequence of PGPR P. fluorescens evaluated in Colombian banana plants. The genome sequences contains genes involved in plant growth and defense, including bacteriocins, 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, and genes that provide resistance to toxic compounds. PMID:27151797

  6. Isoelectric focusing of wound-induced tomato ACC synthase

    SciTech Connect

    White, J.A.; Kende, H. )

    1990-05-01

    Several techniques of electrofocusing have been used to determine whether 1-aminocyclopropane-1-carboxylate (ACC) synthase isolated from wounded tomato pericarp tissue exists in different isoforms, each with its characteristic isoelectric point (pI). The pI of the native enzyme was found to be 6.0 {plus minus} 0.2. When radiolabeled, denatured ACC synthase was electrofocused by non-equilibrium pH gradient electrophoresis (NEpHGE), the enzyme separated into four discernible spots which, upon reaching equilibrium, ranged in pI from 6.6 to 6.9. Immunopurified ACC synthase from four tomato cultivars (Duke, Cornell, Mountain Pride and Pik Red) migrated in each case as a 50-kDa protein on sodium dodecyl sulfate polyacrylamide gels (SDS-PAGE). We propose that native ACC synthase in extracts of tomato pericarp tissue exists in one single form and that the charge heterogeneities observed upon electrofocusing of denatured enzyme result from modifications of preexisting protein.

  7. Accumulation of wound-inducible ACC synthase transcript in tomato fruit is inhibited by salicylic acid and polyamines.

    PubMed

    Li, N; Parsons, B L; Liu, D R; Mattoo, A K

    1992-02-01

    Regulation of wound-inducible 1-aminocyclopropane-1-carboxylic acid (ACC) synthase expression was studied in tomato fruit (Lycopersicon esculentum cv. Pik-Red). A 70 base oligonucleotide probe homologous to published ACC synthase cDNA sequences was successfully used to identify and analyze regulation of a wound-inducible transcript. The 1.8 kb ACC synthase transcript increased upon wounding the fruit as well as during fruit ripening. Salicylic acid, an inhibitor of wound-responsive genes in tomato, inhibited the wound-induced accumulation of the ACC synthase transcript. Further, polyamines (putrescine, spermidine and spermine) that have anti-senescence properties and have been shown to inhibit the development of ACC synthase activity, inhibited the accumulation of the wound-inducible ACC synthase transcript. The inhibition by spermine was greater than that caused by putrescine or spermidine. The transcript level of a wound-repressible glycine-rich protein gene and that of the constitutively expressed rRNA were not affected as markedly by either salicylic acid or polyamines. These data suggest that salicylic acid and polyamines may specifically regulate ethylene biosynthesis at the level of ACC synthase transcript accumulation.

  8. A type III ACC synthase, ACS7, is involved in root gravitropism in Arabidopsis thaliana.

    PubMed

    Huang, Shih-Jhe; Chang, Chia-Lun; Wang, Po-Hsun; Tsai, Min-Chieh; Hsu, Pang-Hung; Chang, Ing-Feng

    2013-11-01

    Ethylene is an important plant hormone that regulates developmental processes in plants. The ethylene biosynthesis pathway is a highly regulated process at both the transcriptional and post-translational level. The transcriptional regulation of these ethylene biosynthesis genes is well known. However, post-translational modifications of the key ethylene biosynthesis enzyme 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS) are little understood. In vitro kinase assays were conducted on the type III ACS, AtACS7, fusion protein and peptides to determine whether the AtACS7 protein can be phosphorylated by calcium-dependent protein kinase (CDPK). AtACS7 was phosphorylated at Ser216, Thr296, and Ser299 by AtCDPK16 in vitro. To investigate further the function of the ACS7 gene in Arabidopsis, an acs7-1 loss-of-function mutant was isolated. The acs7-1 mutant exhibited less sensitivity to the inhibition of root gravitropism by treatment with the calcium chelator ethylene glycol tetraacetic acid (EGTA). Seedlings were treated with gradient concentrations of ACC. The results showed that a certain concentration of ethylene enhanced the gravity response. Moreover, the acs7-1 mutant was less sensitive to inhibition of the gravity response by treatment with the auxin polar transport inhibitor 1-naphthylphthalamic acid, but exogenous ACC application recovered root gravitropism. Altogether, the results indicate that AtACS7 is involved in root gravitropism in a calcium-dependent manner in Arabidopsis.

  9. Differential Expression and Internal Feedback Regulation of 1-Aminocyclopropane-1-Carboxylate Synthase, 1-Aminocyclopropane-1-Carboxylate Oxidase, and Ethylene Receptor Genes in Tomato Fruit during Development and Ripening1

    PubMed Central

    Nakatsuka, Akira; Murachi, Shiho; Okunishi, Hironori; Shiomi, Shinjiro; Nakano, Ryohei; Kubo, Yasutaka; Inaba, Akitsugu

    1998-01-01

    We investigated the feedback regulation of ethylene biosynthesis in tomato (Lycopersicon esculentum) fruit with respect to the transition from system 1 to system 2 ethylene production. The abundance of LE-ACS2, LE-ACS4, and NR mRNAs increased in the ripening fruit concomitant with a burst in ethylene production. These increases in mRNAs with ripening were prevented to a large extent by treatment with 1-methylcyclopropene (MCP), an ethylene action inhibitor. Transcripts for the LE-ACS6 gene, which accumulated in preclimacteric fruit but not in untreated ripening fruit, did accumulate in ripening fruit treated with MCP. Treatment of young fruit with propylene prevented the accumulation of transcripts for this gene. LE-ACS1A, LE-ACS3, and TAE1 genes were expressed constitutively in the fruit throughout development and ripening irrespective of whether the fruit was treated with MCP or propylene. The transcripts for LE-ACO1 and LE-ACO4 genes already existed in preclimacteric fruit and increased greatly when ripening commenced. These increases in LE-ACO mRNA with ripening were also prevented by treatment with MCP. The results suggest that in tomato fruit the preclimacteric system 1 ethylene is possibly mediated via constitutively expressed LE-ACS1A and LE-ACS3 and negatively feedback-regulated LE-ACS6 genes with preexisting LE-ACO1 and LE-ACO4 mRNAs. At the onset of the climacteric stage, it shifts to system 2 ethylene, with a large accumulation of LE-ACS2, LE-ACS4, LE-ACO1, and LE-ACO4 mRNAs as a result of a positive feedback regulation. This transition from system 1 to system 2 ethylene production might be related to the accumulated level of NR mRNA. PMID:9847103

  10. Bacterial Modulation of Plant Ethylene Levels

    PubMed Central

    Gamalero, Elisa; Glick, Bernard R.

    2015-01-01

    A focus on the mechanisms by which ACC deaminase-containing bacteria facilitate plant growth.Bacteria that produce the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, when present either on the surface of plant roots (rhizospheric) or within plant tissues (endophytic), play an active role in modulating ethylene levels in plants. This enzyme activity facilitates plant growth especially in the presence of various environmental stresses. Thus, plant growth-promoting bacteria that express ACC deaminase activity protect plants from growth inhibition by flooding and anoxia, drought, high salt, the presence of fungal and bacterial pathogens, nematodes, and the presence of metals and organic contaminants. Bacteria that express ACC deaminase activity also decrease the rate of flower wilting, promote the rooting of cuttings, and facilitate the nodulation of legumes. Here, the mechanisms behind bacterial ACC deaminase facilitation of plant growth and development are discussed, and numerous examples of the use of bacteria with this activity are summarized. PMID:25897004

  11. Characterization of three members of the ACC synthase gene family in Solanum tuberosum L.

    PubMed

    Destéfano-Beltrán, L J; van Caeneghem, W; Gielen, J; Richard, L; van Montagu, M; van der Straeten, D

    1995-02-20

    Two genomic clones corresponding to three members of the 1-aminocyclopropane-1-carboxylic acid (ACC) synthase gene family in potato (Solanum tuberosum L.) have been isolated and sequenced. Two highly homologous genes, ST-ACS1A and ST-ACS1B, transcribed in opposite directions were found in an 8.9 kb region. Their coding sequences are interrupted by two introns at identical positions. Their closest relative in tomato is the LE-ACS3 gene. The third gene in potato, ST-ACS2, was found in a 4 kb region and shows a gene structure similar to that of the tomato LE-ACS4 gene and to the mung bean VR-ACS4 and VR-ACS5 genes. Based on its lack of significant homology to the tomato gene family and its closeness to the VR-ACS4 and VR-ACS5 genes, we propose that LE-ACS7 represents an additional isoform in the tomato genome. Moreover, in a phylogenetic comparison of known ACC synthases, the ST-ACS2 isoform was grouped in a separate lineage together with the mung bean VR-ACS4 and VR-ACS5, and the moth orchid DS-ACS1A and DS-ACS1B gene products. Expression of the three potato genes was studied by reverse transcription-polymerase chain reaction on total RNA. The twin genes are positively regulated by indole-3-acetic acid in hypocotyls and expression is modulated by wounding in the leaves. The third gene is responsive to ethylene and wounding mainly in tubers. The roles of these three genes and of other members of the ACC synthase gene family in vegetative processes of potato such as tuberization, dormancy, and sprouting have yet to be determined.

  12. ACC oxidase genes expressed in the wood-forming tissues of loblolly pine (Pinus taeda L.) include a pair of nearly identical paralogs (NIPs).

    PubMed

    Yuan, S; Wang, Y; Dean, J F D

    2010-03-15

    1-Aminocyclopropane-1-carboxylate (ACC) oxidase catalyzes the final reaction of the ethylene biosynthetic pathway, converting the unusual cyclic amino acid, ACC, into ethylene. Past studies have shown a possible link between ethylene and compression wood formation in conifers, but the relationship has received no more than modest study at the gene expression level. In this study, a cDNA clone encoding a putative ACC oxidase, PtACO1, was isolated from a cDNA library produced using mRNA from lignifying xylem of loblolly pine (Pinus taeda) trunk wood. The cDNA clone comprised an open reading frame of 1461 bp encoding a protein of 333 amino acids. Using PCR amplification techniques, a genomic clone corresponding to PtACO1 was isolated and shown to contain three introns with typical GT/AG boundaries defining the splice junctions. The PtACO1 gene product shared 70% identity with an ACC oxidase from European white birch (Betula pendula), and phylogenetic analyses clearly placed the gene product in the ACC oxidase cluster of the Arabidopsis thaliana 2-oxoglutarate-dependent dioxygenase superfamily tree. The PtACO1 sequence was used to identify additional ACC oxidase clones from loblolly pine root cDNA libraries characterized as part of an expressed sequence tag (EST) discovery project. The PtACO1 sequence was also used to recover additional paralogous sequences from genomic DNA, one of which (PtACO2) turned out to be >98% identical to PtACO1 in the nucleotide coding sequence, leading to its classification as a "nearly identical paralog" (NIP). Quantitative PCR analyses showed that the expression level of PtACO1-like transcripts varied in different tissues, as well as in response to hormonal treatments and bending. Possible roles for PtACO1 in compression wood formation in loblolly pine and the discovery of its NIP are discussed in light of these results.

  13. Bacteria in combination with fertilizers promote root and shoot growth of maize in saline-sodic soil.

    PubMed

    Zafar-Ul-Hye, Muhammad; Farooq, Hafiz Muhammad; Hussain, Mubshar

    2015-03-01

    Salinity is the leading abiotic stress hampering maize ( Zea mays L.) growth throughout the world, especially in Pakistan. During salinity stress, the endogenous ethylene level in plants increases, which retards proper root growth and consequent shoot growth of the plants. However, certain bacteria contain the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, which converts 1-aminocyclopropane-1-carboxylic acid (an immediate precursor of ethylene biosynthesis in higher plants) into ammonia and α-ketobutyrate instead of ethylene. In the present study, two Pseudomonas bacterial strains containing ACC-deaminase were tested separately and in combinations with mineral fertilizers to determine their potential to minimize/undo the effects of salinity on maize plants grown under saline-sodic field conditions. The data recorded at 30, 50 and 70 days after sowing revealed that both the Pseudomonas bacterial strains improved root and shoot length, root and shoot fresh weight, and root and shoot dry weight up to 34, 43, 35, 71, 55 and 68%, respectively, when applied without chemical fertilizers: these parameter were enhanced up to 108, 95, 100, 131, 100 and 198%, respectively, when the strains were applied along with chemical fertilizers. It can be concluded that ACC-deaminase Pseudomonas bacterial strains applied alone and in conjunction with mineral fertilizers improved the root and shoot growth of maize seedlings grown in saline-sodic soil.

  14. Global Transcriptomic Analysis of Targeted Silencing of Two Paralogous ACC Oxidase Genes in Banana

    PubMed Central

    Xia, Yan; Kuan, Chi; Chiu, Chien-Hsiang; Chen, Xiao-Jing; Do, Yi-Yin; Huang, Pung-Ling

    2016-01-01

    Among 18 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase homologous genes existing in the banana genome there are two genes, Mh-ACO1 and Mh-ACO2, that participate in banana fruit ripening. To better understand the physiological functions of Mh-ACO1 and Mh-ACO2, two hairpin-type siRNA expression vectors targeting both the Mh-ACO1 and Mh-ACO2 were constructed and incorporated into the banana genome by Agrobacterium-mediated transformation. The generation of Mh-ACO1 and Mh-ACO2 RNAi transgenic banana plants was confirmed by Southern blot analysis. To gain insights into the functional diversity and complexity between Mh-ACO1 and Mh-ACO2, transcriptome sequencing of banana fruits using the Illumina next-generation sequencer was performed. A total of 32,093,976 reads, assembled into 88,031 unigenes for 123,617 transcripts were obtained. Significantly enriched Gene Oncology (GO) terms and the number of differentially expressed genes (DEGs) with GO annotation were ‘catalytic activity’ (1327, 56.4%), ‘heme binding’ (65, 2.76%), ‘tetrapyrrole binding’ (66, 2.81%), and ‘oxidoreductase activity’ (287, 12.21%). Real-time RT-PCR was further performed with mRNAs from both peel and pulp of banana fruits in Mh-ACO1 and Mh-ACO2 RNAi transgenic plants. The results showed that expression levels of genes related to ethylene signaling in ripening banana fruits were strongly influenced by the expression of genes associated with ethylene biosynthesis. PMID:27681726

  15. Global Transcriptomic Analysis of Targeted Silencing of Two Paralogous ACC Oxidase Genes in Banana.

    PubMed

    Xia, Yan; Kuan, Chi; Chiu, Chien-Hsiang; Chen, Xiao-Jing; Do, Yi-Yin; Huang, Pung-Ling

    2016-09-26

    Among 18 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase homologous genes existing in the banana genome there are two genes, Mh-ACO1 and Mh-ACO2, that participate in banana fruit ripening. To better understand the physiological functions of Mh-ACO1 and Mh-ACO2, two hairpin-type siRNA expression vectors targeting both the Mh-ACO1 and Mh-ACO2 were constructed and incorporated into the banana genome by Agrobacterium-mediated transformation. The generation of Mh-ACO1 and Mh-ACO2 RNAi transgenic banana plants was confirmed by Southern blot analysis. To gain insights into the functional diversity and complexity between Mh-ACO1 and Mh-ACO2, transcriptome sequencing of banana fruits using the Illumina next-generation sequencer was performed. A total of 32,093,976 reads, assembled into 88,031 unigenes for 123,617 transcripts were obtained. Significantly enriched Gene Oncology (GO) terms and the number of differentially expressed genes (DEGs) with GO annotation were 'catalytic activity' (1327, 56.4%), 'heme binding' (65, 2.76%), 'tetrapyrrole binding' (66, 2.81%), and 'oxidoreductase activity' (287, 12.21%). Real-time RT-PCR was further performed with mRNAs from both peel and pulp of banana fruits in Mh-ACO1 and Mh-ACO2 RNAi transgenic plants. The results showed that expression levels of genes related to ethylene signaling in ripening banana fruits were strongly influenced by the expression of genes associated with ethylene biosynthesis.

  16. ACC synthase genes are polymorphic in watermelon (Citrullus spp.) and differentially expressed in flowers and in response to auxin and gibberellin.

    PubMed

    Salman-Minkov, Ayelet; Levi, Amnon; Wolf, Shmuel; Trebitsh, Tova

    2008-05-01

    The flowering pattern of watermelon species (Citrullus spp.) is either monoecious or andromonoecious. Ethylene is known to play a critical role in floral sex determination of cucurbit species. In contrast to its feminizing effect in cucumber and melon, in watermelon ethylene promotes male flower development. In cucumber, the rate-limiting enzyme of ethylene biosynthesis, 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS), regulates unisexual flower development. To investigate the role of ethylene in flower development, we isolated four genomic sequences of ACS from watermelon (CitACS1-4). Both CitACS1 and CitACS3 are expressed in floral tissue. CitACS1 is also expressed in vegetative tissue and it may be involved in cell growth processes. Expression of CitACS1 is up-regulated by exogenous treatment with auxin, gibberellin or ACC, the immediate precursor of ethylene. No discernible differential floral sex-dependent expression pattern was observed for this gene. The CitACS3 gene is expressed in open flowers and in young staminate floral buds (male or hermaphrodite), but not in female flowers. CitACS3 is also up-regulated by ACC, and is likely to be involved in ethylene-regulated anther development. The expression of CitACS2 was not detected in vegetative or reproductive organs but was up-regulated by auxin. CitACS4 transcript was not detected under our experimental conditions. Restriction fragment length polymorphism (RFLP) and sequence tagged site (STS) marker analyses of the CitACS genes showed polymorphism among and within the different Citrullus groups, including watermelon cultivars, Citrullus lanatus var. lanatus, the central subspecies Citrullus lanatus var. citroides, and the desert species Citrullus colocynthis (L).

  17. Plant Growth-Promoting Rhizobacteria Enhance Salinity Stress Tolerance in Okra through ROS-Scavenging Enzymes.

    PubMed

    Habib, Sheikh Hasna; Kausar, Hossain; Saud, Halimi Mohd

    2016-01-01

    Salinity is a major environmental stress that limits crop production worldwide. In this study, we characterized plant growth-promoting rhizobacteria (PGPR) containing 1-aminocyclopropane-1-carboxylate (ACC) deaminase and examined their effect on salinity stress tolerance in okra through the induction of ROS-scavenging enzyme activity. PGPR inoculated okra plants exhibited higher germination percentage, growth parameters, and chlorophyll content than control plants. Increased antioxidant enzyme activities (SOD, APX, and CAT) and upregulation of ROS pathway genes (CAT, APX, GR, and DHAR) were observed in PGPR inoculated okra plants under salinity stress. With some exceptions, inoculation with Enterobacter sp. UPMR18 had a significant influence on all tested parameters under salt stress, as compared to other treatments. Thus, the ACC deaminase-containing PGPR isolate Enterobacter sp. UPMR18 could be an effective bioresource for enhancing salt tolerance and growth of okra plants under salinity stress.

  18. Plant Growth-Promoting Rhizobacteria Enhance Salinity Stress Tolerance in Okra through ROS-Scavenging Enzymes

    PubMed Central

    Habib, Sheikh Hasna; Kausar, Hossain; Saud, Halimi Mohd

    2016-01-01

    Salinity is a major environmental stress that limits crop production worldwide. In this study, we characterized plant growth-promoting rhizobacteria (PGPR) containing 1-aminocyclopropane-1-carboxylate (ACC) deaminase and examined their effect on salinity stress tolerance in okra through the induction of ROS-scavenging enzyme activity. PGPR inoculated okra plants exhibited higher germination percentage, growth parameters, and chlorophyll content than control plants. Increased antioxidant enzyme activities (SOD, APX, and CAT) and upregulation of ROS pathway genes (CAT, APX, GR, and DHAR) were observed in PGPR inoculated okra plants under salinity stress. With some exceptions, inoculation with Enterobacter sp. UPMR18 had a significant influence on all tested parameters under salt stress, as compared to other treatments. Thus, the ACC deaminase-containing PGPR isolate Enterobacter sp. UPMR18 could be an effective bioresource for enhancing salt tolerance and growth of okra plants under salinity stress. PMID:26951880

  19. Plant–Agrobacterium interaction mediated by ethylene and super-Agrobacterium conferring efficient gene transfer

    PubMed Central

    Nonaka, Satoko; Ezura, Hiroshi

    2014-01-01

    Agrobacterium tumefaciens has a unique ability to transfer genes into plant genomes. This ability has been utilized for plant genetic engineering. However, the efficiency is not sufficient for all plant species. Several studies have shown that ethylene decreased the Agrobacterium-mediated transformation frequency. Thus, A. tumefaciens with an ability to suppress ethylene evolution would increase the efficiency of Agrobacterium-mediated transformation. Some studies showed that plant growth-promoting rhizobacteria (PGPR) can reduce ethylene levels in plants through 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, which cleaves the ethylene precursor ACC into α-ketobutyrate and ammonia, resulting in reduced ethylene production. The whole genome sequence data showed that A. tumefaciens does not possess an ACC deaminase gene in its genome. Therefore, providing ACC deaminase activity to the bacteria would improve gene transfer. As expected, A. tumefaciens with ACC deaminase activity, designated as super-Agrobacterium, could suppress ethylene evolution and increase the gene transfer efficiency in several plant species. In this review, we summarize plant–Agrobacterium interactions and their applications for improving Agrobacterium-mediated genetic engineering techniques via super-Agrobacterium. PMID:25520733

  20. Enhancement of drought stress tolerance in crops by plant growth promoting rhizobacteria.

    PubMed

    Vurukonda, Sai Shiva Krishna Prasad; Vardharajula, Sandhya; Shrivastava, Manjari; SkZ, Ali

    2016-03-01

    Drought is one of the major constraints on agricultural productivity worldwide and is likely to further increase. Several adaptations and mitigation strategies are required to cope with drought stress. Plant growth promoting rhizobacteria (PGPR) could play a significant role in alleviation of drought stress in plants. These beneficial microorganisms colonize the rhizosphere/endo-rhizosphere of plants and impart drought tolerance by producing exopolysaccharides (EPS), phytohormones, 1-aminocyclopropane- 1-carboxylate (ACC) deaminase, volatile compounds, inducing accumulation of osmolytes, antioxidants, upregulation or down regulation of stress responsive genes and alteration in root morphology in acquisition of drought tolerance. The term Induced Systemic Tolerance (IST) was coined for physical and chemical changes induced by microorganisms in plants which results in enhanced tolerance to drought stresses. In the present review we elaborate on the role of PGPR in helping plants to cope with drought stress.

  1. Native Bacterial Endophytes Promote Host Growth in a Species-Specific Manner; Phytohormone Manipulations Do Not Result in Common Growth Responses

    PubMed Central

    Long, Hoang Hoa; Schmidt, Dominik D.; Baldwin, Ian T.

    2008-01-01

    Background All plants in nature harbor a diverse community of endophytic bacteria which can positively affect host plant growth. Changes in plant growth frequently reflect alterations in phytohormone homoeostasis by plant-growth-promoting (PGP) rhizobacteria which can decrease ethylene (ET) levels enzymatically by 1-aminocyclopropane-1-carboxylate (ACC) deaminase or produce indole acetic acid (IAA). Whether these common PGP mechanisms work similarly for different plant species has not been rigorously tested. Methodology/ Principal Findings We isolated bacterial endophytes from field-grown Solanum nigrum; characterized PGP traits (ACC deaminase activity, IAA production, phosphate solubilization and seedling colonization); and determined their effects on their host, S. nigrum, as well as on another Solanaceous native plant, Nicotiana attenuata. In S. nigrum, a majority of isolates that promoted root growth were associated with ACC deaminase activity and IAA production. However, in N. attenuata, IAA but not ACC deaminase activity was associated with root growth. Inoculating N. attenuata and S. nigrum with known PGP bacteria from a culture collection (DSMZ) reinforced the conclusion that the PGP effects are not highly conserved. Conclusions/ Significance We conclude that natural endophytic bacteria with PGP traits do not have general and predictable effects on the growth and fitness of all host plants, although the underlying mechanisms are conserved. PMID:18628963

  2. Adaptive Cruise Control (ACC)

    NASA Astrophysics Data System (ADS)

    Reif, Konrad

    Die adaptive Fahrgeschwindigkeitsregelung (ACC, Adaptive Cruise Control) ist eine Weiterentwicklung der konventionellen Fahrgeschwindigkeitsregelung, die eine konstante Fahrgeschwindigkeit einstellt. ACC überwacht mittels eines Radarsensors den Bereich vor dem Fahrzeug und passt die Geschwindigkeit den Gegebenheiten an. ACC reagiert auf langsamer vorausfahrende oder einscherende Fahrzeuge mit einer Reduzierung der Geschwindigkeit, sodass der vorgeschriebene Mindestabstand zum vorausfahrenden Fahrzeug nicht unterschritten wird. Hierzu greift ACC in Antrieb und Bremse ein. Sobald das vorausfahrende Fahrzeug beschleunigt oder die Spur verlässt, regelt ACC die Geschwindigkeit wieder auf die vorgegebene Sollgeschwindigkeit ein (Bild 1). ACC steht somit für eine Geschwindigkeitsregelung, die sich dem vorausfahrenden Verkehr anpasst.

  3. Fertilizer-dependent efficiency of Pseudomonads for improving growth, yield, and nutrient use efficiency of wheat (Triticum aestivum L.).

    PubMed

    Shaharoona, Baby; Naveed, Muhammad; Arshad, Muhammad; Zahir, Zahir A

    2008-05-01

    Acquisition of nutrients by plants is primarily dependent on root growth and bioavailability of nutrients in the rooting medium. Most of the beneficial bacteria enhance root growth, but their effectiveness could be influenced by the nutrient status around the roots. In this study, two 1-aminocyclopropane-1-carboxylate (ACC)-deaminase containing plant-growth-promoting rhizobacteria (PGPR), Pseudomonas fluorescens and P. fluorescens biotype F were tested for their effect on growth, yield, and nutrient use efficiency of wheat under simultaneously varying levels of all the three major nutrients N, P, and K (at 0%, 25%, 50%, 75%, and 100% of recommended doses). Results of pot and field trials revealed that the efficacy of these strains for improving growth and yield of wheat reduced with the increasing rates of NPK added to the soil. In most of the cases, significant negative linear correlations were recorded between percentage increases in growth and yield parameters of wheat caused by inoculation and increasing levels of applied NPK fertilizers. It is highly likely that under low fertilizer application, the ACC-deaminase activity of PGPR might have caused reduction in the synthesis of stress (nutrient)-induced inhibitory levels of ethylene in the roots through ACC hydrolysis into NH(3) and alpha-ketobutyrate. The results of this study imply that these Pseudomonads could be employed in combination with appropriate doses of fertilizers for better plant growth and savings of fertilizers.

  4. Investigating the ability of Pseudomonas fluorescens UW4 to reduce cadmium stress in Lactuca sativa via an intervention in the ethylene biosynthetic pathway.

    PubMed

    Albano, Lucas J; Macfie, Sheila M

    2016-12-01

    A typical plant response to any biotic or abiotic stress, including cadmium (Cd), involves increased ethylene synthesis, which causes senescence of the affected plant part. Stressed plants can experience reduced ethylene and improved growth if they are inoculated with bacteria that have the enzyme ACC deaminase, which metabolizes the ethylene precursor ACC (1-aminocyclopropane-1-carboxylate). We investigated whether one such bacterium, Pseudomonas fluorescens UW4, reduces the production of ethylene and improves the growth of lettuce (Lactuca sativa) sown in Cd-contaminated potting material (PRO-MIX® BX). Plants were inoculated with the wild-type P. fluorescens UW4 or a mutant strain that cannot produce ACC deaminase. Cadmium-treated plants contained up to 50 times more Cd than did control plants. In noninoculated plants, Cd induced a 5-fold increase in ethylene concentration. The wild-type bacterium prevented Cd-induced reductions in root biomass but there was no relationship between Cd treatment and ethylene production in inoculated plants. In contrast, when the concentration of ethylene was plotted against the extent of bacterial colonization of the roots, increased colonization with wild-type P. fluorescens UW4 was associated with 20% less ethylene production. Ours is the first study to show that the protective effect of this bacterium is proportional to the quantity of bacteria on the root surface.

  5. Genetic and functional diversity among fluorescent pseudomonads isolated from the rhizosphere of banana.

    PubMed

    Naik, Popavath Ravindra; Sahoo, Nirakar; Goswami, Devrishi; Ayyadurai, Niraikulam; Sakthivel, Natarajan

    2008-10-01

    Fluorescent pseudomonads from banana rhizospheric soil were isolated and screened for the production of enzymes and hormones such as phosphatase, indole-3-acetic acid (IAA), 1-aminocyclopropane-1-carboxylate (ACC) deaminase, protease, and antifungal metabolites. Of 95 isolates, 50 (52%) isolates solubilized tri-calcium phosphate (TCP), 63 (66%) isolates produced plant growth hormone IAA, 10 (11%) isolates exhibited ACC deaminase, and 23 (24%) isolates produced protease. Isolates were screened for antifungal activity toward phytopathogenic fungi. Gene-specific primers have identified the putative antibiotic producing isolates. These putative isolates were grown in the production media and production of antibiotics was confirmed by thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC). Genotypic analysis by BOX (bacterial repetitive BOX element)-polymerase chain reaction (PCR) resulted into three distinct genomic clusters at a 50% similarity level and 62 distinct BOX profiles. Based on the sequence similarity of 16S rRNA and construction of subsequent phylogenetic tree analysis, isolates were designated as Pseudomonas monteilii, P. plecoglossicida, P. fluorescens, P. fulva, P. mosselii, P. aeruginosa, P. alcaligenes, and P. pseudoalcaligenes. Present study revealed the genetic and functional diversity among isolates of fluorescent pseudomonads associated with rhizospheric soil of banana and also identified P. monteilii as dominant species. The knowledge on genetic and functional diversity of fluorescent pseudomonads associated with banana rhizosphere is useful to understand their ecological role and for their utilization in sustainable agriculture.

  6. [Characterization of growth-promoting rhizobacteria in Eucalyptus nitens seedlings].

    PubMed

    Angulo, Violeta C; Sanfuentes, Eugenio A; Rodríguez, Francisco; Sossa, Katherine E

    2014-01-01

    Rhizospheric and endophytic bacteria were isolated from the rizosphere and root tissue of Eucalyptus nitens. The objective of this work was to evaluate their capacity to promote growth in seedlings of the same species under greenhouse conditions. The isolates that improved seedling growth were identified and characterized by their capacity to produce indoleacetic acid (IAA), solubilize phosphates and increase 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. One hundred and five morphologically different strains were isolated, 15 of which promoted E. nitens seedling growth, significantly increasing the height (50%), root length (45%) as well as the aerial and root dry weight (142% and 135% respectively) of the plants. Bacteria belonged to the genus Arthrobacter, Lysinibacillus, Rahnella and Bacillus. Isolates A. phenanthrenivorans 21 and B. cereus 113 improved 3.15 times the emergence of E. nitens after 12 days, compared to control samples. Among isolated R. aquatilis, 78 showed the highest production of IAA (97.5±2.87 μg/ml) in the presence of tryptophan and the highest solubilizer index (2.4) for phosphorus, while B. amyloliquefaciens 60 isolate was positive for ACC deaminase activity. Our results reveal the potential of the studied rhizobacteria as promoters of emergence and seedling growth of E. nitens, and their possible use as PGPR inoculants, since they have more than one mechanism associated with plant growth promotion.

  7. Induction of drought tolerance in cucumber plants by a consortium of three plant growth-promoting rhizobacterium strains.

    PubMed

    Wang, Chun-Juan; Yang, Wei; Wang, Chao; Gu, Chun; Niu, Dong-Dong; Liu, Hong-Xia; Wang, Yun-Peng; Guo, Jian-Hua

    2012-01-01

    Our previous work showed that a consortium of three plant growth-promoting rhizobacterium (PGPR) strains (Bacillus cereus AR156, Bacillus subtilis SM21, and Serratia sp. XY21), termed as BBS for short, was a promising biocontrol agent. The present study investigated its effect on drought tolerance in cucumber plants. After withholding watering for 13 days, BBS-treated cucumber plants had much darker green leaves and substantially lighter wilt symptoms than control plants. Compared to the control, the BBS treatment decreased the leaf monodehydroascorbate (MDA) content and relative electrical conductivity by 40% and 15%, respectively; increased the leaf proline content and the root recovery intension by 3.45-fold and 50%, respectively; and also maintained the leaf chlorophyll content in cucumber plants under drought stress. Besides, in relation to the control, the BBS treatment significantly enhanced the superoxide dismutase (SOD) activity and mitigated the drought-triggered down-regulation of the expression of the genes cAPX, rbcL, and rbcS encoding cytosolic ascorbate peroxidase, and ribulose-1,5-bisphosphate carboxy/oxygenase (Rubisco) large and small subunits, respectively, in cucumber leaves. However, 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity was undetected in none of the culture solutions of three BBS constituent strains. These results indicated that BBS conferred induced systemic tolerance to drought stress in cucumber plants, by protecting plant cells, maintaining photosynthetic efficiency and root vigor and increasing some of antioxidase activities, without involving the action of ACC deaminase to lower plant ethylene levels.

  8. Isolation and characterization of endophytic plant growth-promoting bacteria from date palm tree (Phoenix dactylifera L.) and their potential role in salinity tolerance.

    PubMed

    Yaish, Mahmoud W; Antony, Irin; Glick, Bernard R

    2015-06-01

    Endophytic bacteria were isolated from date palm (Phoenix dactylifera L.) seedling roots, characterized and tested for their ability to help plants grow under saline conditions. Molecular characterization showed that the majority of these strains belonged to the genera Bacillus and Enterobacter and had different degrees of resistance to various antibiotics. Some of these strains were able to produce the enzyme 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase and the plant growth regulatory hormone indole-3-acetic acid (IAA). Some strains were also able to chelate ferric iron (Fe(3+)) and solubilize potassium (K(+)), phosphorus (PO 4 (3-) ) and zinc (Zn(2+)), and produce ammonia. The results also showed that ACC deaminase activity and IAA production was slightly increased in some strains in response to an increase in NaCl concentration in the growth media. Consistent with these results, selected strains such as PD-R6 (Paenibacillus xylanexedens) and PD-P6 (Enterobacter cloacae) were able to enhance canola root elongation when grown under normal and saline conditions as demonstrated by a gnotobiotic root elongation assay. These results suggest that the isolated and characterized endophytic bacteria can alter ethylene and IAA levels and also facilitate nutrient uptake in roots and therefore have the potential role to promote the growth and development of date palm trees growing under salinity stress.

  9. Relationship between in vitro characterization and comparative efficacy of plant growth-promoting rhizobacteria for improving cucumber salt tolerance.

    PubMed

    Nadeem, Sajid Mahmood; Ahmad, Maqshoof; Naveed, Muhammad; Imran, Muhammad; Zahir, Zahir Ahmad; Crowley, David E

    2016-05-01

    Phosphate solubilization, 1-aminocyclopropane-1-carboxylic acid (ACC)-deaminase activity and production of siderophores and indole acetic acid (IAA) are well-known traits of plant growth-promoting rhizobacteria (PGPR). Here we investigated the expression of these traits as affected by salinity for three PGPR strains (Pseudomonas fluorescens, Bacillus megaterium and Variovorax paradoxus) at two salinity levels [2 and 5 % NaCl (w/v)]. Among the three strains, growth of B. megaterium was the least affected by high salinity. However, P. fluorescens was the best strain for maintaining ACC-deaminase activity, siderophore and IAA production under stressed conditions. V. paradoxus was the least tolerant to salts and had minimal growth and low PGPR trait expression under salt stress. Results of experiment examining the impact of bacterial inoculation on cucumber growth at three salinity levels [1 (normal), 7 and 10 dS m(-1)] revealed that P. fluorescens also had good rhizosphere competence and was the most effective for alleviating the negative impacts of salinity on cucumber growth. The results suggest that in addition to screening the PGPR regarding their effect on growth under salinity, PGPR trait expression is also an important aspect that may be useful for selecting the most promising PGPR bacterial strains for improving plant tolerance to salinity stress.

  10. Isolation and characterization of endophytic plant growth-promoting (PGPB) or stress homeostasis-regulating (PSHB) bacteria associated to the halophyte Prosopis strombulifera.

    PubMed

    Sgroy, Verónica; Cassán, Fabricio; Masciarelli, Oscar; Del Papa, María Florencia; Lagares, Antonio; Luna, Virginia

    2009-11-01

    This study was designed to isolate and characterize endophytic bacteria from halophyte Prosopis strombulifera grown under extreme salinity and to evaluate in vitro the bacterial mechanisms related to plant growth promotion or stress homeostasis regulation. Isolates obtained from P. strombulifera were compared genotypically by BOX-polymerase chain reaction, grouped according to similarity, and identified by amplification and partial sequences of 16S DNAr. Isolates were grown until exponential growth phase to evaluate the atmospheric nitrogen fixation, phosphate solubilization, siderophores, and phytohormones, such as indole-3-acetic acid, zeatin, gibberellic acid and abscisic acid production, as well as antifungal, protease, and 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. A total of 29 endophytic strains were grouped into seven according to similarity. All bacteria were able to grow and to produce some phytohormone in chemically defined medium with or without addition of a nitrogen source. Only one was able to produce siderophores, and none of them solubilized phosphate. ACC deaminase activity was positive for six strains. Antifungal and protease activity were confirmed for two of them. In this work, we discuss the possible implications of these bacterial mechanisms on the plant growth promotion or homeostasis regulation in natural conditions.

  11. Characterization of Mn-resistant endophytic bacteria from Mn-hyperaccumulator Phytolacca americana and their impact on Mn accumulation of hybrid penisetum.

    PubMed

    Zhang, Wen-Hui; Chen, Wei; He, Lin-Yan; Wang, Qi; Sheng, Xia-Fang

    2015-10-01

    Three hundred Mn-resistant endophytic bacteria were isolated from the Mn-hyperaccumulator, Phytolacca americana, grown at different levels of Mn (0, 1, and 10mM) stress. Under no Mn stress, 90%, 92%, and 11% of the bacteria produced indole acetic acid (IAA), siderophore, and 1-aminocyclopropane-1-carboxylate (ACC) deaminase, respectively. Under Mn stress, 68-94%, 91-92%, and 21-81% of the bacteria produced IAA, siderophore, and ACC deaminase, respectively. Greater percentages of ACC deaminase-producing bacteria were found in the Mn-treated P. americana. Furthermore, the ratios of IAA- and siderophore-producing bacteria were significantly higher in the Mn treated plant leaves, while the ratio of ACC deaminase-producing bacteria was significantly higher in the Mn treated-roots. Based on 16S rRNA gene sequence analysis, Mn-resistant bacteria were affiliated with 10 genera. In experiments involving hybrid penisetum grown in soils treated with 0 and 1000mgkg(-1) of Mn, inoculation with strain 1Y31 was found to increase the root (ranging from 6.4% to 18.3%) and above-ground tissue (ranging from 19.3% to 70.2%) mass and total Mn uptake of above-ground tissues (64%) compared to the control. Furthermore, inoculation with strain 1Y31 was found to increase the ratio of IAA-producing bacteria in the rhizosphere and bulk soils of hybrid penisetum grown in Mn-added soils. The results showed the effect of Mn stress on the ratio of the plant growth-promoting factor-producing endophytic bacteria of P. americana and highlighted the potential of endophytic bacterium as an inoculum for enhanced phytoremediation of Mn-polluted soils by hybrid penisetum plants.

  12. Ethylene limits abscisic acid- or soil drying-induced stomatal closure in aged wheat leaves.

    PubMed

    Chen, Lin; Dodd, Ian C; Davies, William J; Wilkinson, Sally

    2013-10-01

    The mechanism of age-induced decreased stomatal sensitivity to abscisic acid (ABA) and soil drying has been explored here. Older, fully expanded leaves partly lost their ability to close stomata in response to foliar ABA sprays, and soil drying which stimulated endogenous ABA production, while young fully expanded leaves closed their stomata more fully. However, ABA- or soil drying-induced stomatal closure of older leaves was partly restored by pretreating plants with 1-methylcyclopropene (1-MCP), which can antagonize ethylene receptors, or by inoculating soil around the roots with the rhizobacterium Variovorax paradoxus 5C-2, which contains 1-aminocyclopropane-1-carboxylic acid (ACC)-deaminase. ACC (the immediate biosynthetic precursor of ethylene) sprays revealed higher sensitivity of stomata to ethylene in older leaves than younger leaves, despite no differences in endogenous ACC concentrations or ethylene emission. Taken together, these results indicate that the relative insensitivity of stomatal closure to ABA and soil drying in older leaves is likely due to altered stomatal sensitivity to ethylene, rather than ethylene production. To our knowledge, this is the first study to mechanistically explain diminished stomatal responses to soil moisture deficit in older leaves, and the associated reduction in leaf water-use efficiency.

  13. Bioprospecting of Plant Growth Promoting Bacilli and Related Genera Prevalent in Soils of Pristine Sacred Groves: Biochemical and Molecular Approach

    PubMed Central

    Lyngwi, Nathaniel A.; Nongkhlaw, Macmillan; Kalita, Debajit; Joshi, Santa Ram

    2016-01-01

    Bacillus spp. and related genera native to soils of the pristine sacred groves from Meghalaya, India were characterized using biochemical and 16S rRNA gene analysis which revealed dominance of Bacillus, Paenibacillus, Lysinibacillus and Viridibacillus in the groves. Biochemical estimation was carried out for in vitro testing of plant growth promoting traits present in these isolates. PCR screening were performed for plant growth-promoting related genes involved in the biosynthesis of acid phosphatase (AcPho), indolepyruvate decarboxylase (ipdC), 1-aminocyclopropane-1-carboxylate deaminase (accd) and siderophore biosynthesis protein (asbA). 76% of the sacred grove isolates gave an amplified fragment for AcPho. Three of the isolates gave an amplified fragment for IpdC gene. Apart from 2 isolates, all the other isolates including the reference strains were positive for the amplification of the accd gene indicating their potential to produce ACC deaminase enzyme. 42% of the isolates gave an amplified fragment for asbA gene indicating the potential ability of these isolates to produce the catechol type siderophore, petrobactin. Overall findings indicated multiple PGP genetic traits present in these isolates which suggested that these isolates are capable of expressing multiple PGP traits. Phylogenetic and sequence analysis of accd and asbA genes from the isolates revealed that asbA genes from Paenibacillus taichungiensis SG3 and Paenibacillus tylopili SG24 indicated the occurrence of intergeneric horizontal transfer between Paenibacillus and Bacillus. PMID:27111883

  14. Bioprospecting of Plant Growth Promoting Bacilli and Related Genera Prevalent in Soils of Pristine Sacred Groves: Biochemical and Molecular Approach.

    PubMed

    Lyngwi, Nathaniel A; Nongkhlaw, Macmillan; Kalita, Debajit; Joshi, Santa Ram

    2016-01-01

    Bacillus spp. and related genera native to soils of the pristine sacred groves from Meghalaya, India were characterized using biochemical and 16S rRNA gene analysis which revealed dominance of Bacillus, Paenibacillus, Lysinibacillus and Viridibacillus in the groves. Biochemical estimation was carried out for in vitro testing of plant growth promoting traits present in these isolates. PCR screening were performed for plant growth-promoting related genes involved in the biosynthesis of acid phosphatase (AcPho), indolepyruvate decarboxylase (ipdC), 1-aminocyclopropane-1-carboxylate deaminase (accd) and siderophore biosynthesis protein (asbA). 76% of the sacred grove isolates gave an amplified fragment for AcPho. Three of the isolates gave an amplified fragment for IpdC gene. Apart from 2 isolates, all the other isolates including the reference strains were positive for the amplification of the accd gene indicating their potential to produce ACC deaminase enzyme. 42% of the isolates gave an amplified fragment for asbA gene indicating the potential ability of these isolates to produce the catechol type siderophore, petrobactin. Overall findings indicated multiple PGP genetic traits present in these isolates which suggested that these isolates are capable of expressing multiple PGP traits. Phylogenetic and sequence analysis of accd and asbA genes from the isolates revealed that asbA genes from Paenibacillus taichungiensis SG3 and Paenibacillus tylopili SG24 indicated the occurrence of intergeneric horizontal transfer between Paenibacillus and Bacillus.

  15. A Halotolerant Bacterium Bacillus licheniformis HSW-16 Augments Induced Systemic Tolerance to Salt Stress in Wheat Plant (Triticum aestivum)

    PubMed Central

    Singh, Rajnish P.; Jha, Prabhat N.

    2016-01-01

    Certain plant growth promoting bacteria can protect associated plants from harmful effects of salinity. We report the isolation and characterization of 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase bacterium Bacillus licheniformis HSW-16 capable of ameliorating salt (NaCl) stress in wheat plants. The bacterium was isolated from the water of Sambhar salt lake, Rajasthan, India. The presence of ACC deaminase activity was confirmed by enzyme assay and analysis of AcdS gene, a structural gene for ACC deaminase. Inoculation of B. licheniformis HSW-16 protected wheat plants from growth inhibition caused by NaCl and increased plant growth (6-38%) in terms of root length, shoot length, fresh weight, and dry weight. Ionic analysis of plant samples showed that the bacterial inoculation decreased the accumulation of Na+ content (51%), and increased K+ (68%), and Ca2+ content (32%) in plants at different concentration of NaCl. It suggested that bacterial inoculation protected plants from the effect of NaCl by decreasing the level of Na+ in plants. Production of exopolysaccharide by the B. licheniformis HSW-16 can also protect from Na+ by binding this ion. Moreover, application of test isolate resulted in an increase in certain osmolytes such as total soluble sugar, total protein content, and a decrease in malondialdehyde content, illustrating their role in the protection of plants. The ability of B. licheniformis HSW-16 to colonize plant root surface was examined by staining the bacterium with acridine orange followed by fluorescence microscopy and polymerase chain reaction-based DNA finger printing analysis. These results suggested that B. licheniformis HSW-16 could be used as a bioinoculant to improve the productivity of plants growing under salt stress. PMID:28018415

  16. Increased growth and root Cu accumulation of Sorghum sudanense by endophytic Enterobacter sp. K3-2: Implications for Sorghum sudanense biomass production and phytostabilization.

    PubMed

    Li, Ya; Wang, Qi; Wang, Lu; He, Lin-Yan; Sheng, Xia-Fang

    2016-02-01

    Endophytic bacterial strain K3-2 was isolated from the roots of Sorghum sudanense (an bioenergy plant) grown in a Cu mine wasteland soils and characterized. Strain K3-2 was identified as Enterobacter sp. based on 16S rRNA gene sequence analysis. Strain K3-2 exhibited Cu resistance and produced 1-aminocyclopropane-1-carboxylate (ACC) deaminase, indole-3-acetic acid (IAA), siderophores, and arginine decarboxylase. Pot experiments showed that strain K3-2 significantly increased the dry weight and root Cu accumulation of Sorghum sudanense grown in the Cu mine wasteland soils. Furthermore, increase in total Cu uptake (ranging from 49% to 95%) of the bacterial inoculated-Sorghum sudanense was observed compared to the control. Notably, most of Cu (83-86%) was accumulated in the roots of Sorghum sudanense. Furthermore, inoculation with strain K3-2 was found to significantly increase Cu bioconcentration factors and the proportions of IAA- and siderophore-producing bacteria in the root interiors and rhizosphere soils of Sorghum sudanense compared with the control. Significant decrease in the available Cu content was also observed in the rhizosphere soils of the bacterial-inoculated Sorghum sudanense. The results suggest that the endophytic bacterial strain K3-2 may be exploited for promoting Sorghum sudanense biomass production and Cu phytostabilization in the Cu mining wasteland soils.

  17. The effect of plant growth-promoting rhizobacteria on the growth, physiology, and Cd uptake of Arundo donax L.

    PubMed

    Sarathambal, Chinnathambi; Khankhane, Premraj Jagoji; Gharde, Yogita; Kumar, Bhumesh; Varun, Mayank; Arun, Sellappan

    2017-04-03

    In this study, plant growth-promoting potential isolates from rhizosphere of 10 weed species grown in heavy metal-contaminated areas were identified and their effect on growth, antioxidant enzymes, and cadmium (Cd) uptake in Arundo donax L. was explored. Plant growth-promoting traits of isolates were also analyzed. These isolates were found to produce siderophores and enzymes such as 1-aminocyclopropane-1-carboxylate (ACC) deaminase, and aid in solubilization of mineral nutrients and modulate plant growth and development. Based on the presence of multiple plant growth-promoting traits, isolates were selected for molecular characterization and inoculation studies. Altogether, 58 isolates were obtained and 20% of them were able to tolerate Cd up to 400 ppm. The sequence analysis of the 16S rRNA genes indicates that the isolates belong to the phylum Firmicutes. Bacillus sp. along with mycorrhizae inoculation significantly improves the growth, the activity of antioxidants enzymes, and the Cd uptake in A. donax than Bacillus alone. Highly significant correlations were observed between Cd uptake, enzymatic activities, and plant growth characteristics at 1% level of significance. The synergistic interaction effect between these organisms helps to alleviate Cd effects on soil. Heavy metal-tolerant isolate along with arbuscular mycorrhizae (AM) could be used to improve the phytoremedial potential of plants.

  18. Detection of biosynthetic gene and phytohormone production by endophytic actinobacteria associated with Solanum lycopersicum and their plant-growth-promoting effect.

    PubMed

    Passari, Ajit Kumar; Chandra, Preeti; Zothanpuia; Mishra, Vineet Kumar; Leo, Vincent Vineeth; Gupta, Vijai Kumar; Kumar, Brijesh; Singh, Bhim Pratap

    2016-10-01

    In the present study, fifteen endophytic actinobacterial isolates recovered from Solanum lycopersicum were studied for their antagonistic potential and plant-growth-promoting (PGP) traits. Among them, eight isolates showed significant antagonistic and PGP traits, identified by amplification of the 16S rRNA gene. Isolate number DBT204, identified as Streptomyces sp., showed multiple PGP traits tested in planta and improved a range of growth parameters in seedlings of chili (Capsicum annuum L.) and tomato (S. lycopersicum L.). Further, genes of indole acetic acid (iaaM) and 1-aminocyclopropane-1-carboxylate (ACC) deaminase (acdS) were successively amplified from five strains. Six antibiotics (trimethoprim, fluconazole, chloramphenicol, nalidixic acid, rifampicin and streptomycin) and two phytohormones [indole acetic acid (IAA) and kinetin (KI)] were detected and quantified in Streptomyces sp. strain DBT204 using UPLC-ESI-MS/MS. The study indicates the potential of these PGP strains for production of phytohormones and shows the presence of biosynthetic genes responsible for production of secondary metabolites. It is the first report showing production of phytohormones (IAA and KI) by endophytic actinobacteria having PGP and biosynthetic potential. We propose Streptomyces sp. strain DBT204 for inoculums production and development of biofertilizers for enhancing growth of chili and tomato seedlings.

  19. Enhancement of heavy metal phytoremediation by Alnus firma with endophytic Bacillus thuringiensis GDB-1.

    PubMed

    Babu, A Giridhar; Kim, Jong-Dae; Oh, Byung-Taek

    2013-04-15

    Phytoremediation shows potential for remediating mine tailing sites contaminated with heavy metals. Our aim was to isolate, characterize, and assess the potential of endophytic bacteria to enhance growth and metal accumulation by the hyperaccumulator Alnus firma. A bacterial strain isolated from roots of Pinus sylvestris had the capacity to remove heavy metals from mine tailing and was identified as Bacillus thuringiensis GDB-1 based on 16S ribosomal DNA sequencing. GDB-1 exhibited plant growth-promoting traits, including 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity, indole acetic acid (IAA) and siderophore production, and P solubilization. The efficiency of GDB-1 to remove heavy metals was influenced by pH and initial metal concentration. Removal capacity (mg/l) was 77% for Pb (100), 64% for Zn (50), 34% for As (50), 9% for Cd (10), 8% for Cu (10), and 8% for Ni (10) during the active growth cycle in heavy metal-amended, mine tailing extract medium. Inoculating soil with GDB-1 significantly increased biomass, chlorophyll content, nodule number, and heavy metal (As, Cu, Pb, Ni, and Zn) accumulation in A. firma seedlings. Results indicate that inoculating the native plant A. firma with B. thuringiensis GDB-1 improves its efficiency for phytoremediation of soil containing mine tailings contaminated with heavy metals.

  20. Genetic diversity of plant growth promoting rhizobacteria isolated from rhizospheric soil of wheat under saline condition.

    PubMed

    Upadhyay, Sudhir K; Singh, Devendra P; Saikia, Ratul

    2009-11-01

    In this study, a total of 130 rhizobacteria was isolated from a saline infested zone of wheat rhizosphere, and screened for plant growth promoting (PGP) traits at higher salt (NaCl) concentrations (2, 4, 6, and 8%). The results revealed that 24 rhizobacterial isolates were tolerant at 8% NaCl. Although all the 24 salt tolerable isolates produced indole-3-acetic acid (IAA), while 10 isolates solubilized phosphorus, eight produced siderophore, and six produced gibberellin. However, only three isolates showed the production of 1-aminocyclopropane-1-carboxylate (ACC) deaminase. Diversity was analyzed through 16S rDNA-RFLP, and of these isolates with three tetra cutter restriction enzymes (HaeIII, AluI, and MspI), the representative cluster groups were identified by 16S rDNA sequencing. Bacillus and Bacillus-derived genera were dominant which showed PGP attributes at 8% NaCl concentration. Out of 24 isolates, nitrogen fixing ability (nif H gene) was detected in the two isolates, SU18 (Arthrobacter sp.) and SU48.

  1. Native rhizobia from Zn mining soil promote the growth of Leucaena leucocephala on contaminated soil.

    PubMed

    Rangel, Wesley M; Thijs, Sofie; Janssen, Jolien; Oliveira Longatti, Silvia M; Bonaldi, Daiane S; Ribeiro, Paula R A; Jambon, Inge; Eevers, Nele; Weyens, Nele; Vangronsveld, Jaco; Moreira, Fatima M S

    2017-02-01

    Plants on contaminated mining soils often show a reduced growth due to nutrient depletion as well as trace elements (TEs) toxicity. Since those conditions threat plant's survival, plant growth-promoting rhizobacteria (PGPRs), such as rhizobia, might be of crucial importance for plant colonization on TE-contaminated soils. Native rhizobia from mining soils are promising candidates for bioaugmented phytoremediation of those soils as they are adapted to the specific conditions. In this work, rhizobia from Zn- and Cd-contaminated mining soils were in vitro screened for their PGP features [organic acids, indole-3-acetic acid (IAA), and siderophore (SID) production; 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity; and Ca3(PO4)2 solubilization] and Zn and Cd tolerance. In addition, some type and reference rhizobia strains were included in the study as well. The in vitro screening indicated that rhizobia and other native genera have great potential for phytoremediation purposes, by exerting, besides biological N2 fixation, other plant growth-promoting traits. Leucaena leucocephala-Mesorhizobium sp. (UFLA 01-765) showed multielement tolerance and an efficient symbiosis on contaminated soil, decreasing the activities of antioxidative enzymes in shoots. This symbiosis is a promising combination for phytostabilization.

  2. Stimulation of the growth of Jatropha curcas by the plant growth promoting bacterium Enterobacter cancerogenus MSA2.

    PubMed

    Jha, Chaitanya Kumar; Patel, Baldev; Saraf, Meenu

    2012-03-01

    A novel Enterobacter cancerogenus MSA2 is a plant growth promoting gamma-proteobacterium that was isolated from the rhizosphere of Jatropha cucas a potentially important biofuel feed stock plant. Based on phenotypic, physiological, biochemical and phylogenetic studies, strain MSA2 could be classified as a member of E. cancerogenus. However, comparisons of characteristics with other known species of the genus Enterobacter suggested that strain MSA2 could be a novel PGPB strain. In vitro studies were carried for the plant growth promoting attribute of this culture. It tested positive for ACC (1-aminocyclopropane-1-carboxylic acid) deaminase production, phytase, phosphate solubilization, IAA (Indole acetic acid) production, siderophore, and ammonia production. The isolate was then used as a inoculant for the vegetative study of Jatropha curcas plant. Enterobacter cancerogenus MSA2 supplemented with 1% carboxymethylcellulose showed overall plant growth promotion effect resulting in enhanced root length (124.14%), fresh root mass (81%), fresh shoot mass (120.02%), dry root mass (124%), dry shoot mass (105.54%), number of leaf (30.72%), chlorophyll content (50.41%), and biomass (87.20%) over control under the days of experimental observation. This study was designed for 120 days and was in triplicate and the data was collected at every 30 days.

  3. Plant growth-promoting rhizobacteria (PGPR): emergence in agriculture.

    PubMed

    Bhattacharyya, P N; Jha, D K

    2012-04-01

    Plant growth-promoting rhizobacteria (PGPR) are the rhizosphere bacteria that can enhance plant growth by a wide variety of mechanisms like phosphate solubilization, siderophore production, biological nitrogen fixation, rhizosphere engineering, production of 1-Aminocyclopropane-1-carboxylate deaminase (ACC), quorum sensing (QS) signal interference and inhibition of biofilm formation, phytohormone production, exhibiting antifungal activity, production of volatile organic compounds (VOCs), induction of systemic resistance, promoting beneficial plant-microbe symbioses, interference with pathogen toxin production etc. The potentiality of PGPR in agriculture is steadily increased as it offers an attractive way to replace the use of chemical fertilizers, pesticides and other supplements. Growth promoting substances are likely to be produced in large quantities by these rhizosphere microorganisms that influence indirectly on the overall morphology of the plants. Recent progress in our understanding on the diversity of PGPR in the rhizosphere along with their colonization ability and mechanism of action should facilitate their application as a reliable component in the management of sustainable agricultural system. The progress to date in using the rhizosphere bacteria in a variety of applications related to agricultural improvement along with their mechanism of action with special reference to plant growth-promoting traits are summarized and discussed in this review.

  4. Trichoderma virens PDR-28: a heavy metal-tolerant and plant growth-promoting fungus for remediation and bioenergy crop production on mine tailing soil.

    PubMed

    Babu, A Giridhar; Shim, Jaehong; Bang, Keuk-Soo; Shea, Patrick J; Oh, Byung-Taek

    2014-01-01

    A heavy metal-tolerant fungus, Trichoderma virens PDR-28, was isolated from rhizosphere soil and evaluated for use in remediating mine tailing soil and for plant biomass production. PDR-28 exhibited plant growth-promoting traits, including 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, acid phosphatase and phytase activity, siderophore production, and P solubilization. HMs were more available in mine tailing soil inoculated soil with PDR-28 than in uninoculated soil; the order of HM bioleaching was Cd > As > Zn > Pb > Cu. PDR-28 effectively removed HMs in the order of Pb > Cd > As > Zn > Cu from liquid media containing 100 mg HM L(-1). Inoculating HM-contaminated mine tailing soil with the fungus significantly increased the dry biomass of maize roots (64%) and shoots (56%). Chlorophyll, total soluble sugars (reducible and nonreducible), starch, and protein contents increased by 46%, 28%, 30%, and 29%, respectively, compared to plants grown in uninoculated soil. Inoculation increased heavy metal concentrations in maize roots by 25% (Cu) to 62% (Cd) and in shoots by 35% (Cu) to 64% (Pb) compared to uninoculated plants. Results suggest that PDR-28 would be beneficial for phytostabilization and plant biomass production as a potential source of biofuel in the quest for renewable energy.

  5. Rhizosphere bacteria of Costularia spp. from ultramafic soils in New Caledonia: diversity, tolerance to extreme edaphic conditions, and role in plant growth and mineral nutrition.

    PubMed

    Gonin, Mathieu; Gensous, Simon; Lagrange, Alexandre; Ducousso, Marc; Amir, Hamid; Jourand, Philippe

    2013-03-01

    Rhizosphere bacteria were isolated from Costularia spp., pioneer sedges from ultramafic soils in New Caledonia, which is a hotspot of biodiversity in the South Pacific. Genus identification, ability to tolerate edaphic constraints, and plant-growth-promoting (PGP) properties were analysed. We found that 10(5) colony-forming units per gram of root were dominated by Proteobacteria (69%) and comprised 21 genera, including Burkholderia (28%), Curtobacterium (15%), Bradyrhizobium (9%), Sphingomonas (8%), Rhizobium (7%), and Bacillus (5%). High proportions of bacteria tolerated many elements of the extreme edaphic conditions: 82% tolerated 100 μmol·L(-1) chromium, 70% 1 mmol·L(-1) nickel, 63% 10 mmol·L(-1) manganese, 24% 1 mmol·L(-1) cobalt, and 42% an unbalanced calcium/magnesium ratio (1/16). These strains also exhibited multiple PGP properties, including the ability to produce ammonia (65%), indole-3-acetic acid (60%), siderophores (52%), and 1-aminocyclopropane-1-carboxylate (ACC) deaminase (39%); as well as the capacity to solubilize phosphates (19%). The best-performing strains were inoculated with Sorghum sp. grown on ultramafic substrate. Three strains significantly enhanced the shoot biomass by up to 33%. The most successful strains influenced plant nutrition through the mobilization of metals in roots and a reduction of metal transfer to shoots. These results suggest a key role of these bacteria in plant growth, nutrition, and adaptation to the ultramafic constraints.

  6. Evaluation of Varying Biochars as Carrier Materials for Bacterial Soil Inoculants

    NASA Astrophysics Data System (ADS)

    Hale, Lauren; Crowley, David

    2014-05-01

    The incorporation of biochar into agricultural soils for carbon sequestration and improved soil fertility creates an opportunity to simultaneously deliver plant-growth promoting rhizobacteria (PGPR). Many characteristics of biochar materials indicate that these particles could be conducive as inoculum carriers. This could provide a value-added component for biochar marketing and has an advantage over traditional carrier materials, which can be unsustainable or expensive to produce. Here, we assessed the suitability of 10 biochar types, made from 5 feedstocks at 2 pyrolysis temperatures (300°C and 600°C), to serve as carriers for 2 model PGPR strains, Enterobacter cloacae UW5 and Pseudomonas putida UW4. All biochars were characterized based on BET specific surface area, C-N content, pH, EC, and their abilities to adsorb bacterial cells from a liquid inoculum. Further studies incorporated qPCR to quantify the survival of inoculants after introduction into soils via biochar carriers. The biochars that performed well were further assayed for their influence on PGPR traits, 1-aminocyclopropane-1-carboxylate (ACC) deaminase and auxin production. Peat and vermiculite served as traditional carrier materials to which we compared the biochars. Our findings indicated that biochars varied in their interactions with our model PGPR strains. Based on our analysis several biochar types were able to serve as carriers which were as good, if not better than, the traditional carrier materials. Future work should seek to assess shelf life and varying inoculation methods for the biochar-inoculant complexes.

  7. Endophytic Cultivable Bacteria of the Metal Bioaccumulator Spartina maritima Improve Plant Growth but Not Metal Uptake in Polluted Marshes Soils

    PubMed Central

    Mesa, Jennifer; Mateos-Naranjo, Enrique; Caviedes, Miguel A.; Redondo-Gómez, Susana; Pajuelo, Eloisa; Rodríguez-Llorente, Ignacio D.

    2015-01-01

    Endophytic bacterial population was isolated from Spartina maritima tissues, a heavy metal bioaccumulator cordgrass growing in the estuaries of Tinto, Odiel, and Piedras River (south west Spain), one of the most polluted areas in the world. Strains were identified and ability to tolerate salt and heavy metals along with plant growth promoting and enzymatic properties were analyzed. A high proportion of these bacteria were resistant toward one or several heavy metals and metalloids including As, Cu, and Zn, the most abundant in plant tissues and soil. These strains also exhibited multiple enzymatic properties as amylase, cellulase, chitinase, protease and lipase, as well as plant growth promoting properties, including nitrogen fixation, phosphates solubilization, and production of indole-3-acetic acid (IAA), siderophores and 1-aminocyclopropane-1-carboxylate (ACC) deaminase. The best performing strains (Micrococcus yunnanensis SMJ12, Vibrio sagamiensis SMJ18, and Salinicola peritrichatus SMJ30) were selected and tested as a consortium by inoculating S. maritima wild plantlets in greenhouse conditions along with wild polluted soil. After 30 days, bacterial inoculation improved plant photosynthetic traits and favored intrinsic water use efficiency. However, far from stimulating plant metal uptake, endophytic inoculation lessened metal accumulation in above and belowground tissues. These results suggest that inoculation of S. maritima with indigenous metal-resistant endophytes could mean a useful approach in order to accelerate both adaption and growth of this indigenous cordgrass in polluted estuaries in restorative operations, but may not be suitable for rhizoaccumulation purposes. PMID:26733985

  8. Brevundimonas diminuta mediated alleviation of arsenic toxicity and plant growth promotion in Oryza sativa L.

    PubMed

    Singh, Namrata; Marwa, Naina; Mishra, Shashank K; Mishra, Jyoti; Verma, Praveen C; Rathaur, Sushma; Singh, Nandita

    2016-03-01

    Arsenic (As), a toxic metalloid adversely affects plant growth in polluted areas. In the present study, we investigated the possibility of improving phytostablization of arsenic through application of new isolated strain Brevundimonas diminuta (NBRI012) in rice plant [Oryza sativa (L.) Var. Sarju 52] at two different concentrations [10ppm (low toxic) and 50ppm (high toxic)] of As. The plant growth promoting traits of bacterial strains revealed the inherent ability of siderophores, phosphate solubilisation, indole acetic acid (IAA), 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase production which may be associated with increased biomass, chlorophyll and MDA content of rice and thereby promoting plant growth. The study also revealed the As accumulation property of NBRI012 strain which could play an important role in As removal from contaminated soil. Furthermore, NBRI012 inoculation significantly restored the hampered root epidermal and cortical cell growth of rice plant and root hair elimination. Altogether our study highlights the multifarious role of B. diminuta in mediating stress tolerance and modulating translocation of As in edible part of rice plant.

  9. Poplar and its bacterial endophytes: coexistence and harmony

    SciTech Connect

    van der Lelie, D.; Taghavi, S.; Monchy, S.; Schwender, J.; Miller, L.; Ferrieri, R.; Rogers, A.; Zhu, W.; Weyens, N.; Vangronsveld, J.; Newman, L.

    2009-09-01

    Associations between plants and microorganisms are very complex and are the subject of an increasing number of studies. Here, we specifically address the relationship between poplar and its endophytic bacteria. The role and importance of endophytic bacteria in growth and development of their host plants is still underestimated. However, since many endophytes have a beneficial effect on their host, an improved understanding of the interaction between poplar and its endophytic bacteria has the potential to provide major breakthroughs that will improve the productivity of poplar. Endophytic bacteria can improve plant growth and development in a direct or indirect way. Direct plant growth promoting mechanisms may involve nitrogen fixation, production of plant growth regulators such as auxins, cytokinins and gibberellins, and suppression of stress ethylene synthesis by 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. Endophytic bacteria can indirectly benefit the plant by preventing the growth or activity of plant pathogens through competition for space and nutrients, antibiosis, production of hydrolytic enzymes, inhibition of pathogen-produced enzymes or toxins, and through systemic induction of plant defense mechanisms. Examples of applications for custom endophyte-host partnerships include improved productivity and establishment of poplar trees on marginal soils and the phytoremediation of contaminated soils and groundwater. A systems biology approach to understand the synergistic interactions between poplar and its beneficial endophytic bacteria represents an important field of research, which is facilitated by the recent sequencing of the genomes of poplar and several of its endophytic bacteria.

  10. Epiphytic and endophytic bacteria that promote growth of ethnomedicinal plants in the subtropical forests of Meghalaya, India.

    PubMed

    Nongkhlaw, Fenella Mary War; Joshi, S R

    2014-12-01

    The present study was aimed to investigate the endophytic and epiphytic bacteria associated with selected ethnomedicinal plants from the pristine subtropical forests of Meghalaya and analyse them for plant growth promotion and antagonistic ability. This study is an attempt to explore plant associated bacteria which are beneficial to host plants, and thus aid in the conservation of ethnomedicinal plants of the studied subtropical forests, which are dwindling due to exploitation. The plant growth promotion parameters like indole acetic acid (IAA) production, mineral phosphate solubilisation, acid phosphatase activity, presence of 1-aminocyclopropane-1-carboxylic acid deaminase (ACC) gene, nitrogen fixation, cellulose digestion, chitin and pectin degrada- tion were screened among the isolates. The study revealed significant differences in bacterial population not only between the epiphytic and endophytic microhabitats, but also amongst the host plants. Out of the 70 isolated plant associated bacteria, Bacillus sp., Serratia sp., Pseudomonas sp., Pantoea sp., and Lysinibacillus sp. showed potent plant growth promotion properties. Bacillus siamensis C53 and B. subtilis cenB showed significant antagonistic activity against the tested pathogens. This study indicated the isolates inhabiting the plants prevalent in the subtropical sacred forests could be explored for use as plant growth promoters while practising the cultiva- tion and conservation of ethnomedicinal plants.

  11. Genetics Home Reference: adenosine monophosphate deaminase deficiency

    MedlinePlus

    ... that can affect the muscles used for movement ( skeletal muscles ). In many affected individuals, AMP deaminase deficiency does ... called AMP deaminase. This enzyme is found in skeletal muscles , where it plays a role in producing energy. ...

  12. Cloning, identification and expression analysis of ACC oxidase gene involved in ethylene production pathway.

    PubMed

    Jafari, Zohreh; Haddad, Raheem; Hosseini, Ramin; Garoosi, Ghasemali

    2013-02-01

    1-aminocyclopropane-1-carboxylic acid oxidase (ACO) enzyme is a member of the Fe II-dependent family of oxidases/oxygenases which require Fe(2+) as a cofactor, ascorbate as a cosubstrate and CO(2) as an activator. This enzyme catalyses the terminal step in the plant signaling of ethylene biosynthetic pathway. A 948 bp fragment of the ACO1 gene cDNA sequence was cloned from tomato (Lycopersicon esculentum) fruit tissues by using reverse transcriptase-polymerase chain reaction (RT-PCR) with two PCR primers designed according to the sequence of a tomato cDNA clone (X58273). The BLAST search showed a high level of similarity (77-98 %) between ACO1 and ACO genes of other plants. The calculated molecular mass and predicted isoelectric point of LeACO1 were 35.8 kDa and 5.13, respectively. The three-dimensional structure studies illustrated that the LeACO1 protein folds into a compact jelly-roll motif comprised of 8 α-helices, 12 β-strands and several long loops. The cosubstrate was located in a cofactor-binding pocket referred to as a 2-His-1-carboxylate facial triad. Semi-quantitative RT-PCR analysis of gene expression revealed that the LeACO1 was expressed in fruit tissues at different ripening stages.

  13. Bioprospecting in potato fields in the Central Andean Highlands: screening of rhizobacteria for plant growth-promoting properties.

    PubMed

    Ghyselinck, Jonas; Velivelli, Siva L S; Heylen, Kim; O'Herlihy, Eileen; Franco, Javier; Rojas, Mercy; De Vos, Paul; Prestwich, Barbara Doyle

    2013-03-01

    The Central Andean Highlands are the center of origin of the potato plant (Solanum tuberosum). Ages of mutualism between potato plants and soil bacteria in this region support the hypothesis that Andean soils harbor interesting plant growth-promoting (PGP) bacteria. Therefore, the aim of this study was to isolate rhizobacteria from Andean ecosystems, and to identify those with PGP properties. A total of 585 bacterial isolates were obtained from eight potato fields in the Andes and they were screened for suppression of Phytophthora infestans and Rhizoctonia solani. Antagonistic mechanisms were determined and antagonistic isolates were further tested for phosphate solubilization, 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity, and production of NH3- and indole-3-acetic acid (IAA). PGP was studied in healthy and R. solani diseased plantlets under growth room conditions. Performance was compared to the commercial strain B. subtilis FZB24(®) WG. Isolates were dereplicated with matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS), and identified with 16S rRNA gene sequencing and multi locus sequence analysis (MLSA). A total of 10% of the isolates were effective antagonists, of which many were able to solubilize phosphate, and produce IAA, ACC deaminase, NH3 and hydrogen cyanide (HCN). During growth room experiments, 23 antagonistic isolates were associated with plant growth-promotion and/or disease suppression. Ten isolates had a statistically significant impact on test parameters compared to the uninoculated control. Three isolates significantly promoted plant growth in healthy plantlets compared to the commercial strain, and seven isolates outperformed the commercial strain in in vitro R. solani diseased plantlets.

  14. Metagenomic Analysis of the Bacterial Community Associated with the Taproot of Sugar Beet

    PubMed Central

    Tsurumaru, Hirohito; Okubo, Takashi; Okazaki, Kazuyuki; Hashimoto, Megumi; Kakizaki, Kaori; Hanzawa, Eiko; Takahashi, Hiroyuki; Asanome, Noriyuki; Tanaka, Fukuyo; Sekiyama, Yasuyo; Ikeda, Seishi; Minamisawa, Kiwamu

    2015-01-01

    We analyzed a metagenome of the bacterial community associated with the taproot of sugar beet (Beta vulgaris L.) in order to investigate the genes involved in plant growth-promoting traits (PGPTs), namely 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, indole acetic acid (IAA), N2 fixation, phosphate solubilization, pyrroloquinoline quinone, siderophores, and plant disease suppression as well as methanol, sucrose, and betaine utilization. The most frequently detected gene among the PGPT categories encoded β-1,3-glucanase (18 per 105 reads), which plays a role in the suppression of plant diseases. Genes involved in phosphate solubilization (e.g., for quinoprotein glucose dehydrogenase), methanol utilization (e.g., for methanol dehydrogenase), siderophore production (e.g. isochorismate pyruvate lyase), and ACC deaminase were also abundant. These results suggested that such PGPTs are crucially involved in supporting the growth of sugar beet. In contrast, genes for IAA production (iaaM and ipdC) were less abundant (~1 per 105 reads). N2 fixation genes (nifHDK) were not detected; bacterial N2 -fixing activity was not observed in the 15N2 -feeding experiment. An analysis of nitrogen metabolism suggested that the sugar beet microbiome mainly utilized ammonium and nitroalkane as nitrogen sources. Thus, N2 fixation and IAA production did not appear to contribute to sugar beet growth. Taxonomic assignment of this metagenome revealed the high abundance of Mesorhizobium, Bradyrhizobium, and Streptomyces, suggesting that these genera have ecologically important roles in the taproot of sugar beet. Bradyrhizobium-assigned reads in particular were found in almost all categories of dominant PGPTs with high abundance. The present study revealed the characteristic functional genes in the taproot-associated microbiome of sugar beet, and suggest the opportunity to select sugar beet growth-promoting bacteria. PMID:25740621

  15. Induction of Drought Tolerance in Cucumber Plants by a Consortium of Three Plant Growth-Promoting Rhizobacterium Strains

    PubMed Central

    Wang, Chao; Gu, Chun; Niu, Dong-Dong; Liu, Hong-Xia; Wang, Yun-Peng; Guo, Jian-Hua

    2012-01-01

    Our previous work showed that a consortium of three plant growth-promoting rhizobacterium (PGPR) strains (Bacillus cereus AR156, Bacillus subtilis SM21, and Serratia sp. XY21), termed as BBS for short, was a promising biocontrol agent. The present study investigated its effect on drought tolerance in cucumber plants. After withholding watering for 13 days, BBS-treated cucumber plants had much darker green leaves and substantially lighter wilt symptoms than control plants. Compared to the control, the BBS treatment decreased the leaf monodehydroascorbate (MDA) content and relative electrical conductivity by 40% and 15%, respectively; increased the leaf proline content and the root recovery intension by 3.45-fold and 50%, respectively; and also maintained the leaf chlorophyll content in cucumber plants under drought stress. Besides, in relation to the control, the BBS treatment significantly enhanced the superoxide dismutase (SOD) activity and mitigated the drought-triggered down-regulation of the expression of the genes cAPX, rbcL, and rbcS encoding cytosolic ascorbate peroxidase, and ribulose-1,5-bisphosphate carboxy/oxygenase (Rubisco) large and small subunits, respectively, in cucumber leaves. However, 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity was undetected in none of the culture solutions of three BBS constituent strains. These results indicated that BBS conferred induced systemic tolerance to drought stress in cucumber plants, by protecting plant cells, maintaining photosynthetic efficiency and root vigor and increasing some of antioxidase activities, without involving the action of ACC deaminase to lower plant ethylene levels. PMID:23285089

  16. Trichoderma-plant root colonization: escaping early plant defense responses and activation of the antioxidant machinery for saline stress tolerance.

    PubMed

    Brotman, Yariv; Landau, Udi; Cuadros-Inostroza, Álvaro; Tohge, Takayuki; Takayuki, Tohge; Fernie, Alisdair R; Chet, Ilan; Viterbo, Ada; Willmitzer, Lothar

    2013-03-01

    Trichoderma spp. are versatile opportunistic plant symbionts which can colonize the apoplast of plant roots. Microarrays analysis of Arabidopsis thaliana roots inoculated with Trichoderma asperelloides T203, coupled with qPCR analysis of 137 stress responsive genes and transcription factors, revealed wide gene transcript reprogramming, proceeded by a transient repression of the plant immune responses supposedly to allow root colonization. Enhancement in the expression of WRKY18 and WRKY40, which stimulate JA-signaling via suppression of JAZ repressors and negatively regulate the expression of the defense genes FMO1, PAD3 and CYP71A13, was detected in Arabidopsis roots upon Trichoderma colonization. Reduced root colonization was observed in the wrky18/wrky40 double mutant line, while partial phenotypic complementation was achieved by over-expressing WRKY40 in the wrky18 wrky40 background. On the other hand increased colonization rate was found in roots of the FMO1 knockout mutant. Trichoderma spp. stimulate plant growth and resistance to a wide range of adverse environmental conditions. Arabidopsis and cucumber (Cucumis sativus L.) plants treated with Trichoderma prior to salt stress imposition show significantly improved seed germination. In addition, Trichoderma treatment affects the expression of several genes related to osmo-protection and general oxidative stress in roots of both plants. The MDAR gene coding for monodehydroascorbate reductase is significantly up-regulated and, accordingly, the pool of reduced ascorbic acid was found to be increased in Trichoderma treated plants. 1-Aminocyclopropane-1-carboxylate (ACC)-deaminase silenced Trichoderma mutants were less effective in providing tolerance to salt stress, suggesting that Trichoderma, similarly to ACC deaminase producing bacteria, can ameliorate plant growth under conditions of abiotic stress, by lowering ameliorating increases in ethylene levels as well as promoting an elevated antioxidative capacity.

  17. Evaluating Performance Portability of OpenACC

    SciTech Connect

    Sabne, Amit J; Sakdhnagool, Putt; Lee, Seyong; Vetter, Jeffrey S

    2015-01-01

    Accelerator-based heterogeneous computing is gaining momentum in High Performance Computing arena. However, the increased complexity of the accelerator architectures demands more generic, high-level programming models. OpenACC is one such attempt to tackle the problem. While the abstraction endowed by OpenACC offers productivity, it raises questions on its portability. This paper evaluates the performance portability obtained by OpenACC on twelve OpenACC programs on NVIDIA CUDA, AMD GCN, and Intel MIC architectures. We study the effects of various compiler optimizations and OpenACC program settings on these architectures to provide insights into the achieved performance portability.

  18. Bacterial Ammeline Metabolism via Guanine Deaminase

    PubMed Central

    Seffernick, Jennifer L.; Dodge, Anthony G.; Sadowsky, Michael J.; Bumpus, John A.; Wackett, Lawrence P.

    2010-01-01

    Melamine toxicity in mammals has been attributed to the blockage of kidney tubules by insoluble complexes of melamine with cyanuric acid or uric acid. Bacteria metabolize melamine via three consecutive deamination reactions to generate cyanuric acid. The second deamination reaction, in which ammeline is the substrate, is common to many bacteria, but the genes and enzymes responsible have not been previously identified. Here, we combined bioinformatics and experimental data to identify guanine deaminase as the enzyme responsible for this biotransformation. The ammeline degradation phenotype was demonstrated in wild-type Escherichia coli and Pseudomonas strains, including E. coli K12 and Pseudomonas putida KT2440. Bioinformatics analysis of these and other genomes led to the hypothesis that the ammeline deaminating enzyme was guanine deaminase. An E. coli guanine deaminase deletion mutant was deficient in ammeline deaminase activity, supporting the role of guanine deaminase in this reaction. Two guanine deaminases from disparate sources (Bradyrhizobium japonicum USDA 110 and Homo sapiens) that had available X-ray structures were purified to homogeneity and shown to catalyze ammeline deamination at rates sufficient to support bacterial growth on ammeline as a sole nitrogen source. In silico models of guanine deaminase active sites showed that ammeline could bind to guanine deaminase in a similar orientation to guanine, with a favorable docking score. Other members of the amidohydrolase superfamily that are not guanine deaminases were assayed in vitro, and none had substantial ammeline deaminase activity. The present study indicated that widespread guanine deaminases have a promiscuous activity allowing them to catalyze a key reaction in the bacterial transformation of melamine to cyanuric acid and potentially contribute to the toxicity of melamine. PMID:20023034

  19. Airborne Cloud Computing Environment (ACCE)

    NASA Technical Reports Server (NTRS)

    Hardman, Sean; Freeborn, Dana; Crichton, Dan; Law, Emily; Kay-Im, Liz

    2011-01-01

    Airborne Cloud Computing Environment (ACCE) is JPL's internal investment to improve the return on airborne missions. Improve development performance of the data system. Improve return on the captured science data. The investment is to develop a common science data system capability for airborne instruments that encompasses the end-to-end lifecycle covering planning, provisioning of data system capabilities, and support for scientific analysis in order to improve the quality, cost effectiveness, and capabilities to enable new scientific discovery and research in earth observation.

  20. Potential of Pseudomonas putida PCI2 for the Protection of Tomato Plants Against Fungal Pathogens.

    PubMed

    Pastor, Nicolás; Masciarelli, Oscar; Fischer, Sonia; Luna, Virginia; Rovera, Marisa

    2016-09-01

    Tomato is one of the most economically attractive vegetable crops due to its high yields. Diseases cause significant losses in tomato production worldwide. We carried out Polymerase Chain Reaction studies to detect the presence of genes encoding antifungal compounds in the DNA of Pseudomonas putida strain PCI2. We also used liquid chromatography-electrospray tandem mass spectrometry to detect and quantify the production of compounds that increase the resistance of plants to diseases from culture supernatants of PCI2. In addition, we investigated the presence of 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase in PCI2. Finally, PCI2 was used for inoculation of tomato seeds to study its potential biocontrol activity against Fusarium oxysporum MR193. The obtained results showed that no fragments for the encoding genes of hydrogen cyanide, pyoluteorin, 2,4-diacetylphloroglucinol, pyrrolnitrin, or phenazine-1-carboxylic acid were amplified from the DNA of PCI2. On the other hand, PCI2 produced salicylic acid and jasmonic acid in Luria-Bertani medium and grew in a culture medium containing ACC as the sole nitrogen source. We observed a reduction in disease incidence from 53.33 % in the pathogen control to 30 % in tomato plants pre-inoculated with PCI2 as well as increases in shoot and root dry weights in inoculated plants, as compared to the pathogenicity control. This study suggests that inoculation of tomato seeds with P. putida PCI2 increases the resistance of plants to root rot caused by F. oxysporum and that PCI2 produces compounds that may be involved at different levels in increasing such resistance. Thus, PCI2 could represent a non-contaminating management strategy potentially applicable in vegetable crops such as tomato.

  1. ACC Effectiveness Review, 1999-2002.

    ERIC Educational Resources Information Center

    Wallace, Roslyn, Ed.

    2002-01-01

    These newsletters on Institutional Effectiveness (IE) at Austin Community College (ACC) in Texas include the following articles: (1) "The 'Fast Track'...Students Say It Works!" (2) "Are Students Successfully Completing Distance Learning Courses at ACC?" (3) "Tracking Transfers"; (4) "Math Pilot: Study Skills…

  2. Rescue of the Orphan Enzyme Isoguanine Deaminase

    PubMed Central

    Hitchcock, Daniel S.; Fedorov, Alexander A.; Fedorov, Elena V.; Dangott, Lawrence J.; Almo, Steven C.; Raushel, Frank M.

    2011-01-01

    Cytosine deaminase (CDA) from Escherichia coli was shown to catalyze the deamination of isoguanine (2-oxoadenine) to xanthine. Isoguanine is an oxidation product of adenine in DNA that is mutagenic to the cell. The isoguanine deaminase activity in E. coli was partially purified by ammonium sulfate fractionation, gel filtration and anion exchange chromatography. The active protein was identified by peptide mass fingerprint analysis as cytosine deaminase. The kinetic constants for the deamination of isoguanine at pH 7.7 are kcat = 49 s-1, Km = 72 μM, and kcat/Km = 6.7 × 105 M-1 s-1. The kinetic constant for the deamination of cytosine are kcat = 45 s-1, Km = 302 μM, and kcat/Km = 1.5 × 105 M-1 s-1. Under these reaction conditions isoguanine is the better substrate for cytosine deaminase. The three dimensional structure of CDA was determined with isoguanine in the active site. PMID:21604715

  3. Genetics Home Reference: adenosine deaminase 2 deficiency

    MedlinePlus

    ... This Page Bras J, Guerreiro R, Santo GC. Mutant ADA2 in vasculopathies. N Engl J Med. 2014 ... M, Anikster Y, King MC, Levy-Lahad E. Mutant adenosine deaminase 2 in a polyarteritis nodosa vasculopathy. ...

  4. Radioimmunochemical quantitation of human adenosine deaminase.

    PubMed Central

    Daddona, P E; Frohman, M A; Kelley, W N

    1979-01-01

    Markedly reduced or absent adenosine deaminase activity in man is associated with an autosomal recesive form of severe conbined immunodeficiency disease. To further define the genetic nature of this enzyme defect, we have quantitated immunologically active adenosine deaminase (CRM) in the hemolysate of homozygous deficient patients and their heterozygous parents. A highly specific radioimmunoassay was developed capable of detecting 0.05% of normal erythrocyte adenosine deaminase. Hemolysates from nine heterozygotes (five families) showed a wide range in CRM (32--100% of normal) and variable absolute specific activities with several being at least 1 SD BELOW THE NORMAL MEAN. Hemolysates from four unrelated patients showed less than 0.09% adenosine deaminase activity with CRM ranging from less than 0.06 to 5.6% of the normal mean. In conclusion, heterozygote and homozygote hemolysates from five of the eight families analyzed revealed variable levels of CRM suggesting heterogeneous genetic alteration or expression of the silent or defective allele(s) of adenosine deaminase. PMID:468994

  5. Maintaining Genome Stability: The Role of Helicases and Deaminases

    DTIC Science & Technology

    2007-07-01

    of Helicases and Deaminases PRINCIPAL INVESTIGATOR: XiaoJiang Chen CONTRACTING ORGANIZATION: University of Southern...SUBTITLE 5a. CONTRACT NUMBER Maintaining Genome Stability: The Role of Helicases and Deaminases 5b. GRANT NUMBER W81XWH-05-1-0391 5c... deaminases . We will focus on AID and APOBEC3G to obtain purified deaminase proteins for the in vitro biochemical, functional, and structural

  6. Maintaining Genome Stability: The Role of Helicases and Deaminases

    DTIC Science & Technology

    2006-07-01

    W81XWH-05-1-0391 TITLE: Maintaining Genome Stability: The Role of Helicases and Deaminases PRINCIPAL INVESTIGATOR: Xiaojiang Chen...Helicases and Deaminases 5b. GRANT NUMBER W81XWH-05-1-0391 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d. PROJECT NUMBER Xiaojiang Chen 5e...crystallize the proteins of deaminases . We will focus on AID and APOBEC3G to obtain purified deaminase proteins for the in vitro biochemical

  7. Whole Genome Sequencing and Analysis of Plant Growth Promoting Bacteria Isolated from the Rhizosphere of Plantation Crops Coconut, Cocoa and Arecanut

    PubMed Central

    Thomas, George V.; Manikandan, Vinu; Gajewski, John; Thomas, George; Seshagiri, Somasekar; Schuster, Stephan C.

    2014-01-01

    Coconut, cocoa and arecanut are commercial plantation crops that play a vital role in the Indian economy while sustaining the livelihood of more than 10 million Indians. According to 2012 Food and Agricultural organization's report, India is the third largest producer of coconut and it dominates the production of arecanut worldwide. In this study, three Plant Growth Promoting Rhizobacteria (PGPR) from coconut (CPCRI-1), cocoa (CPCRI-2) and arecanut (CPCRI-3) characterized for the PGP activities have been sequenced. The draft genome sizes were 4.7 Mb (56% GC), 5.9 Mb (63.6% GC) and 5.1 Mb (54.8% GB) for CPCRI-1, CPCRI-2, CPCRI-3, respectively. These genomes encoded 4056 (CPCRI-1), 4637 (CPCRI-2) and 4286 (CPCRI-3) protein-coding genes. Phylogenetic analysis revealed that both CPCRI-1 and CPCRI-3 belonged to Enterobacteriaceae family, while, CPCRI-2 was a Pseudomonadaceae family member. Functional annotation of the genes predicted that all three bacteria encoded genes needed for mineral phosphate solubilization, siderophores, acetoin, butanediol, 1-aminocyclopropane-1-carboxylate (ACC) deaminase, chitinase, phenazine, 4-hydroxybenzoate, trehalose and quorum sensing molecules supportive of the plant growth promoting traits observed in the course of their isolation and characterization. Additionally, in all the three CPCRI PGPRs, we identified genes involved in synthesis of hydrogen sulfide (H2S), which recently has been proposed to aid plant growth. The PGPRs also carried genes for central carbohydrate metabolism indicating that the bacteria can efficiently utilize the root exudates and other organic materials as energy source. Genes for production of peroxidases, catalases and superoxide dismutases that confer resistance to oxidative stresses in plants were identified. Besides these, genes for heat shock tolerance, cold shock tolerance and glycine-betaine production that enable bacteria to survive abiotic stress were also identified. PMID:25162593

  8. Volatile-Mediated Effects Predominate in Paraburkholderia phytofirmans Growth Promotion and Salt Stress Tolerance of Arabidopsis thaliana.

    PubMed

    Ledger, Thomas; Rojas, Sandy; Timmermann, Tania; Pinedo, Ignacio; Poupin, María J; Garrido, Tatiana; Richter, Pablo; Tamayo, Javier; Donoso, Raúl

    2016-01-01

    Abiotic stress has a growing impact on plant growth and agricultural activity worldwide. Specific plant growth promoting rhizobacteria have been reported to stimulate growth and tolerance to abiotic stress in plants, and molecular mechanisms like phytohormone synthesis and 1-aminocyclopropane-1-carboxylate deamination are usual candidates proposed to mediate these bacterial effects. Paraburkholderia phytofirmans PsJN is able to promote growth of several plant hosts, and improve their tolerance to chilling, drought and salinity. This work investigated bacterial determinants involved in PsJN stimulation of growth and salinity tolerance in Arabidopsis thaliana, showing bacteria enable plants to survive long-term salinity treatment, accumulating less sodium within leaf tissues relative to non-inoculated controls. Inactivation of specific bacterial genes encoding ACC deaminase, auxin catabolism, N-acyl-homoserine-lactone production, and flagellin synthesis showed these functions have little influence on bacterial induction of salinity tolerance. Volatile organic compound emission from strain PsJN was shown to reproduce the effects of direct bacterial inoculation of roots, increasing plant growth rate and tolerance to salinity evaluated both in vitro and in soil. Furthermore, early exposure to VOCs from P. phytofirmans was sufficient to stimulate long-term effects observed in Arabidopsis growth in the presence and absence of salinity. Organic compounds were analyzed in the headspace of PsJN cultures, showing production of 2-undecanone, 7-hexanol, 3-methylbutanol and dimethyl disulfide. Exposure of A. thaliana to different quantities of these molecules showed that they are able to influence growth in a wide range of added amounts. Exposure to a blend of the first three compounds was found to mimic the effects of PsJN on both general growth promotion and salinity tolerance. To our knowledge, this is the first report on volatile compound-mediated induction of plant abiotic

  9. Whole genome sequencing and analysis of plant growth promoting bacteria isolated from the rhizosphere of plantation crops coconut, cocoa and arecanut.

    PubMed

    Gupta, Alka; Gopal, Murali; Thomas, George V; Manikandan, Vinu; Gajewski, John; Thomas, George; Seshagiri, Somasekar; Schuster, Stephan C; Rajesh, Preeti; Gupta, Ravi

    2014-01-01

    Coconut, cocoa and arecanut are commercial plantation crops that play a vital role in the Indian economy while sustaining the livelihood of more than 10 million Indians. According to 2012 Food and Agricultural organization's report, India is the third largest producer of coconut and it dominates the production of arecanut worldwide. In this study, three Plant Growth Promoting Rhizobacteria (PGPR) from coconut (CPCRI-1), cocoa (CPCRI-2) and arecanut (CPCRI-3) characterized for the PGP activities have been sequenced. The draft genome sizes were 4.7 Mb (56% GC), 5.9 Mb (63.6% GC) and 5.1 Mb (54.8% GB) for CPCRI-1, CPCRI-2, CPCRI-3, respectively. These genomes encoded 4056 (CPCRI-1), 4637 (CPCRI-2) and 4286 (CPCRI-3) protein-coding genes. Phylogenetic analysis revealed that both CPCRI-1 and CPCRI-3 belonged to Enterobacteriaceae family, while, CPCRI-2 was a Pseudomonadaceae family member. Functional annotation of the genes predicted that all three bacteria encoded genes needed for mineral phosphate solubilization, siderophores, acetoin, butanediol, 1-aminocyclopropane-1-carboxylate (ACC) deaminase, chitinase, phenazine, 4-hydroxybenzoate, trehalose and quorum sensing molecules supportive of the plant growth promoting traits observed in the course of their isolation and characterization. Additionally, in all the three CPCRI PGPRs, we identified genes involved in synthesis of hydrogen sulfide (H2S), which recently has been proposed to aid plant growth. The PGPRs also carried genes for central carbohydrate metabolism indicating that the bacteria can efficiently utilize the root exudates and other organic materials as energy source. Genes for production of peroxidases, catalases and superoxide dismutases that confer resistance to oxidative stresses in plants were identified. Besides these, genes for heat shock tolerance, cold shock tolerance and glycine-betaine production that enable bacteria to survive abiotic stress were also identified.

  10. Multiple impacts of the plant growth-promoting rhizobacterium Variovorax paradoxus 5C-2 on nutrient and ABA relations of Pisum sativum

    PubMed Central

    Dodd, Ian C.

    2012-01-01

    Resolving the physiological mechanisms by which rhizobacteria enhance plant growth is difficult, since many such bacteria contain multiple plant growth-promoting properties. To understand further how the 1-aminocyclopropane-1-carboxylate (ACC) deaminase (ACCd)-containing rhizobacterium Variovorax paradoxus 5C-2 affects plant growth, the flows and partitioning of mineral nutrients and abscisic acid (ABA) and ABA metabolism were studied in pea (Pisum sativum) plants following rhizosphere bacterial inoculation. Although root architecture was not affected, inoculation increased root and shoot biomass, and stomatal conductance, by 20, 15, and 24%, respectively, and increased N, P, K, Ca, and Mg uptake by 16, 81, 50, 46, and 58%, respectively. P deposition in inoculated plant roots was 4.9 times higher than that in uninoculated controls. Rhizobacterial inoculation increased root to shoot xylem flows and shoot to root phloem flows of K by 1.8- and 2.1-fold, respectively. In control plants, major sinks for K deposition were the roots and upper shoot (43% and 49% of total uptake, respectively), while rhizobacterial inoculation increased K distribution to the lower shoot at the expense of other compartments (xylem, phloem, and upper shoot). Despite being unable to metabolize ABA in vitro, V. paradoxus 5C-2 decreased root ABA concentrations and accumulation by 40–60%. Although inoculation decreased xylem ABA flows, phloem ABA flows increased. Whether bacterial ACCd attenuates root to shoot ABA signalling requires further investigation, since ABA is critical to maintain growth of droughted plants, and ACCd-containing organisms have been advocated as a means of minimizing growth inhibition of plants in drying soil. PMID:23136167

  11. A new insight to adsorption and accumulation of high lead concentration by exopolymer and whole cells of lead-resistant bacterium Acinetobacter junii L. Pb1 isolated from coal mine dump.

    PubMed

    Kushwaha, Anamika; Rani, Radha; Kumar, Sanjay; Thomas, Tarence; David, Arun Alfred; Ahmed, Meraz

    2017-03-11

    A lead-resistant bacterial strain was isolated from coal mine dump and identified as Acinetobacter junii Pb1 on basis of 16S rRNA (ribosomal ribonucleic acid) gene sequencing. The minimum inhibitory concentration of lead for the strain was 16,000 mg l(-1) and it showed antibiotic and multi metal resistance. In aqueous culture, at an initial lead (Pb(II)) concentration of 100 and 500 mg l(-1), lead adsorption and accumulation by the isolate was 100 and 60%, at pH 7 at 30 °C after 48 and 120 h, respectively. The two fractions of exopolysaccharide (EPS), loosely associated EPS (laEPS) and bound EPS (bEPS), and whole cells (devoid of EPS) showed high binding affinity towards Pb(II). The binding affinity of laEPS towards Pb(II) (1071 mg Pb g(-1)) was three times higher than that of bEPS (321.5 mg Pb g(-1)) and 6.5 times higher than that of whole cells (165 mg Pb g(-1)). The binding affinity of EPS and whole cells with Pb(II), reported in the current study, is considerably higher as compared to that reported in the literature, till date. SEM analysis, showed an increase in thickness of cells on exposure to Pb(II) and TEM analysis, revealed its accumulation (interior of cell) and its adsorption (with the external cell surface). The isolate was also found to be positive for indole acetic acid (IAA) and 1-aminocyclopropane-1-carboxylate (ACC) deaminase production which helps in promoting plant growth. Thus, this study provides a new understanding towards Pb(II) uptake by A. junii Pb1, highlighting its potential on the restoration of Pb(II) contaminated repositories.

  12. Effect of IAA produced by Klebsiella oxytoca Rs-5 on cotton growth under salt stress.

    PubMed

    Liu, Yan; Shi, Zaiqiang; Yao, Lixia; Yue, Haitao; Li, Hui; Li, Chun

    2013-01-01

    Klebsiella oxytoca Rs-5 isolated with ACC (1-aminocyclopropane-1-carboxylate) deaminase activity as the sole nitrogen source could obviously promote cotton seedling growth under salt stress and produce phytohormone indole-3-acetic acid (IAA). The amount of IAA produced by the strain Rs-5 was measured, and the effect of IAA on cotton growth under salt stress was studied. Different treatments were set to treat cotton seeds with fermentation broth containing strain Rs-5 (FB), strain Rs-5, fermentation broth with bacteria removed (FB-NB), fermentation broth without bacteria or IAA (FB-NB-NI) and single IAA solutions (SI) according to the IAA concentration after strain Rs-5 culturing of 48, 72 and 120 h. The germination rate, dry weight, plant height, root length and malondialdehyde (MDA), proline and endogenous IAA content in roots were determined. The results showed that both IAA produced by strain Rs-5 and the strain were effective in promoting cotton growth under salt stress. The growth and ability to resist salt stress of cotton seedlings were increased with the enhancement of IAA concentration. The treatment of FB containing bacteria and IAA at 120 h obtained the best state of cotton growth, when the IAA content was the highest in the fermentation broth (42.14 μg·L(-1)). The germination rate, dry weight, plant height and root length were increased by 29.4%, 24.3%, 27.2% and 27.2% , respectively, compared to the saline control. The strain Rs-5 and/or IAA could obviously reduce the MDA and proline content and increase the endogenous IAA content in cotton seedlings. However, the efficacy of other components in the fermentation broth was inconspicuous.

  13. Characterization of a versatile rhizospheric organism from cucumber identified as Ochrobactrum haematophilum.

    PubMed

    Zhao, Lei; Teng, Songshan; Liu, Yanping

    2012-04-01

    Several rhizobacteria play a vital role in promoting plant growth and protecting plants against fungal diseases and degrading pesticides in the environment. In this study, a bacterial strain, designated H10, was isolated from the rhizosphere at Laixi in Shandong Province, China, and was identified as Ochrobactrum haematophilum based on API 20 NE tests and 16S rRNA gene sequence analysis. The plant growth-promoting characteristics of the strain were further characterized, and the results showed that strain H10 produces siderophore, indol-3-acetic (IAA) and solubilized phosphate but lacks 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. Inoculation with the strain was found to significantly increase (p < 0.05) the growth of cucumber in pot experiments. Strain H10 was assessed in vitro for antagonism against several pathogenic fungi and showed high antifungal activity. The cell-free culture filtrates, which had high extracellular chitinase, β-1,3-glucanase and protease activities, could inhibit the growth of all pathogenic fungi tested, indicating that growth suppression was partly due to extracellular antifungal metabolites present in the culture filtrates. Changes in hyphal morphology were observed in phytopathogenic fungi after treatment with the culture filtrates. Additionally, strain H10 was able to degrade 80%, 85% and 58% of the pesticides chlorpyrifos, β-cypermethrin and imidacloprid, respectively, within 60 h in liquid culture. The inoculation of strain H10 into soil treated with 100 mg kg(-1) of the three pesticides accordingly resulted in a higher degradation rate than in noninoculated soils. These results highlight the potential of this bacterium for use as a biofertilizer and biopesticide and suggest that it may provide an alternative to the use of chemical fertilizers and pesticides in agriculture. Additionally, it may represent a bioremediation agent that can remove contaminating chemical pesticide residues from the environment.

  14. Multiple impacts of the plant growth-promoting rhizobacterium Variovorax paradoxus 5C-2 on nutrient and ABA relations of Pisum sativum.

    PubMed

    Jiang, Fan; Chen, Lin; Belimov, Andrey A; Shaposhnikov, Alexander I; Gong, Fan; Meng, Xu; Hartung, Wolfram; Jeschke, Dieter W; Davies, William J; Dodd, Ian C

    2012-11-01

    Resolving the physiological mechanisms by which rhizobacteria enhance plant growth is difficult, since many such bacteria contain multiple plant growth-promoting properties. To understand further how the 1-aminocyclopropane-1-carboxylate (ACC) deaminase (ACCd)-containing rhizobacterium Variovorax paradoxus 5C-2 affects plant growth, the flows and partitioning of mineral nutrients and abscisic acid (ABA) and ABA metabolism were studied in pea (Pisum sativum) plants following rhizosphere bacterial inoculation. Although root architecture was not affected, inoculation increased root and shoot biomass, and stomatal conductance, by 20, 15, and 24%, respectively, and increased N, P, K, Ca, and Mg uptake by 16, 81, 50, 46, and 58%, respectively. P deposition in inoculated plant roots was 4.9 times higher than that in uninoculated controls. Rhizobacterial inoculation increased root to shoot xylem flows and shoot to root phloem flows of K by 1.8- and 2.1-fold, respectively. In control plants, major sinks for K deposition were the roots and upper shoot (43% and 49% of total uptake, respectively), while rhizobacterial inoculation increased K distribution to the lower shoot at the expense of other compartments (xylem, phloem, and upper shoot). Despite being unable to metabolize ABA in vitro, V. paradoxus 5C-2 decreased root ABA concentrations and accumulation by 40-60%. Although inoculation decreased xylem ABA flows, phloem ABA flows increased. Whether bacterial ACCd attenuates root to shoot ABA signalling requires further investigation, since ABA is critical to maintain growth of droughted plants, and ACCd-containing organisms have been advocated as a means of minimizing growth inhibition of plants in drying soil.

  15. Volatile-Mediated Effects Predominate in Paraburkholderia phytofirmans Growth Promotion and Salt Stress Tolerance of Arabidopsis thaliana

    PubMed Central

    Ledger, Thomas; Rojas, Sandy; Timmermann, Tania; Pinedo, Ignacio; Poupin, María J.; Garrido, Tatiana; Richter, Pablo; Tamayo, Javier

    2016-01-01

    Abiotic stress has a growing impact on plant growth and agricultural activity worldwide. Specific plant growth promoting rhizobacteria have been reported to stimulate growth and tolerance to abiotic stress in plants, and molecular mechanisms like phytohormone synthesis and 1-aminocyclopropane-1-carboxylate deamination are usual candidates proposed to mediate these bacterial effects. Paraburkholderia phytofirmans PsJN is able to promote growth of several plant hosts, and improve their tolerance to chilling, drought and salinity. This work investigated bacterial determinants involved in PsJN stimulation of growth and salinity tolerance in Arabidopsis thaliana, showing bacteria enable plants to survive long-term salinity treatment, accumulating less sodium within leaf tissues relative to non-inoculated controls. Inactivation of specific bacterial genes encoding ACC deaminase, auxin catabolism, N-acyl-homoserine-lactone production, and flagellin synthesis showed these functions have little influence on bacterial induction of salinity tolerance. Volatile organic compound emission from strain PsJN was shown to reproduce the effects of direct bacterial inoculation of roots, increasing plant growth rate and tolerance to salinity evaluated both in vitro and in soil. Furthermore, early exposure to VOCs from P. phytofirmans was sufficient to stimulate long-term effects observed in Arabidopsis growth in the presence and absence of salinity. Organic compounds were analyzed in the headspace of PsJN cultures, showing production of 2-undecanone, 7-hexanol, 3-methylbutanol and dimethyl disulfide. Exposure of A. thaliana to different quantities of these molecules showed that they are able to influence growth in a wide range of added amounts. Exposure to a blend of the first three compounds was found to mimic the effects of PsJN on both general growth promotion and salinity tolerance. To our knowledge, this is the first report on volatile compound-mediated induction of plant abiotic

  16. Human adenosine deaminase. Distribution and properties.

    PubMed

    Van der Weyden, M B; Kelley, W N

    1976-09-25

    Adenosine deaminase exists in multiple molecular forms in human tissue. One form of the enzyme appears to be "particulate". Three forms of the enzyme are soluble and interconvertible with apparent molecular weights of approximately 36,000, 114,000, and 298,000 (designated small, intermediate, and large, respectively). The small form of adenosine deaminase is convertible to the large form only in the presence of a protein, which has an apparent molecular weight of 200,000 and has no adenosine deaminase activity. This conversion of the small form of the enzyme to the large form occurs at 4 degrees, exhibits a pH optimum of 5.0 to 8.0, and is associated with a loss of conversion activity. The small form of the enzyme predominates in tissue preparations exhibiting the higher enzyme-specific activities and no detectable conversion activity. The large form of adenosine deaminase predominates in tissue extracts exhibiting the lower enzyme specific activities and abundant conversion activity. The small form of adenosine deaminase shows several electrophoretic variants by isoelectric focusing. The electrophoretic heterogeneity observed with the large form of the enzyme is similar to that observed with the small form, with the exception that several additional electrophoretic variants are uniformly identified. No organ specificity is demonstrable for the different electrophoretic forms. The kinetic characteristics of the three soluble molecular species of adenosine deaminase are identical except for pH optimum, which is 5.5 for the intermediate species and 7.0 to 7.4 for the large and small forms.

  17. GENERAL: The effect of ACC vehicles to mixed traffic flow consisting of manual and ACC vehicles

    NASA Astrophysics Data System (ADS)

    Xie, Dong-Fan; Gao, Zi-You; Zhao, Xiao-Mei

    2008-12-01

    This paper studies the effect of adaptive cruise control (ACC) system on traffic flow by using simulations. The multiple headway and velocity direrence (MHVD) model is used to depict the motion of ACC vehicles, and the simulation results are compared with the optimal velocity (OV) model which is used to depict the motion of manual vehicles. Compared the cases between the manual and the ACC vehicle flow, the fundamental diagram can be classified into four regions: I, II, III, IV. In low and high density the flux of the two models is the same; in region II the free flow region of the MHVD model is enlarged, and the flux of the MHVD model is larger than that of the OV model; in region III serious jams occur in the OV model while the ACC system suppresses the jams in the MHVD model and the traffic flow is in order, but the flux of the OV model is larger than that of the MHVD model. Similar phenomena also appeared in mixed traffic flow which consists of manual and ACC vehicles. The results indicate that ACC vehicles have significant effect on traffic flow. The improvement induced by ACC vehicles decreases with the increasing proportion of ACC vehicles.

  18. Rescue of the Orphan Enzyme Isoguanine Deaminase

    SciTech Connect

    D Hitchcock; A Fedorov; E Fedorov; L Dangott; S Almo; F Raushel

    2011-12-31

    Cytosine deaminase (CDA) from Escherichia coli was shown to catalyze the deamination of isoguanine (2-oxoadenine) to xanthine. Isoguanine is an oxidation product of adenine in DNA that is mutagenic to the cell. The isoguanine deaminase activity in E. coli was partially purified by ammonium sulfate fractionation, gel filtration, and anion exchange chromatography. The active protein was identified by peptide mass fingerprint analysis as cytosine deaminase. The kinetic constants for the deamination of isoguanine at pH 7.7 are as follows: k{sub cat} = 49 s{sup -1}, K{sub m} = 72 {micro}M, and k{sub cat}/K{sub m} = 6.7 x 10{sup 5} M{sup -1} s{sup -1}. The kinetic constants for the deamination of cytosine are as follows: k{sub cat} = 45 s{sup -1}, K{sub m} = 302 {micro}M, and k{sub cat}/K{sub m} = 1.5 x 10{sup 5} M{sup -1} s{sup -1}. Under these reaction conditions, isoguanine is the better substrate for cytosine deaminase. The three-dimensional structure of CDA was determined with isoguanine in the active site.

  19. Rescue of the orphan enzyme isoguanine deaminase.

    PubMed

    Hitchcock, Daniel S; Fedorov, Alexander A; Fedorov, Elena V; Dangott, Lawrence J; Almo, Steven C; Raushel, Frank M

    2011-06-28

    Cytosine deaminase (CDA) from Escherichia coli was shown to catalyze the deamination of isoguanine (2-oxoadenine) to xanthine. Isoguanine is an oxidation product of adenine in DNA that is mutagenic to the cell. The isoguanine deaminase activity in E. coli was partially purified by ammonium sulfate fractionation, gel filtration, and anion exchange chromatography. The active protein was identified by peptide mass fingerprint analysis as cytosine deaminase. The kinetic constants for the deamination of isoguanine at pH 7.7 are as follows: k(cat) = 49 s(-1), K(m) = 72 μM, and k(cat)/K(m) = 6.7 × 10(5) M(-1) s(-1). The kinetic constants for the deamination of cytosine are as follows: k(cat) = 45 s(-1), K(m) = 302 μM, and k(cat)/K(m) = 1.5 × 10(5) M(-1) s(-1). Under these reaction conditions, isoguanine is the better substrate for cytosine deaminase. The three-dimensional structure of CDA was determined with isoguanine in the active site.

  20. ACC forum looks at 'burning' questions

    SciTech Connect

    Carter, R.

    2005-06-01

    The American Coal Council's (ACC) Spring Coal Forum had as its theme: Coal's renaissance: prospects for regenerating coal generation'. It explored US coal demand, supply, end-user technology and market trends. The article gives an overview of the conference, highlighting several presentations. 2 figs., 1 tab.

  1. Adrenocortical carcinoma (ACC): diagnosis, prognosis, and treatment

    PubMed Central

    Libé, Rossella

    2015-01-01

    Adrenocortical carticnoma (ACC) is a rare malignancy with an incidence of 0.7–2.0 cases/million habitants/year. The diagnosis of malignancy relies on careful investigations of clinical, biological, and imaging features before surgery and pathological examination after tumor removal. Most patients present with steroid hormone excess or abdominal mass effects, but 15% of patients with ACC is initially diagnosed incidentally. After the diagnosis, in order to assess the ACC prognosis and establish an adequate basis for treatment decisions different tools are proposed. The stage classification proposed by the European Network for the Study of Adrenal Tumors (ENSAT) is recommended. Pathology reports define the Weiss score, the resection status and the proliferative index, including the mitotic count and the Ki67 index. As far as the treatment is concerned, in case of tumor limited to the adrenal gland, the complete resection of the tumor is the first option. Most patients benefit from adjuvant mitotane treatment. In metastatic disease, mitotane is the cornerstone of initial treatment, and cytotoxic drugs should be added in case of progression. Recently, the First International Randomized (FIRM-ACT) Trial in metastatic ACC reported the association between mitotane and etoposide/doxorubicin/cisplatin (EDP) as the new standard in first line treatment of ACC. In last years, new targeted therapies, including the IGF-1 receptor inhibitors, have been investigated, but their efficacy remains limited. Thus, new treatment concepts are urgently needed. The ongoing “omic approaches” and next-generation sequencing will improve our understanding of the pathogenesis and hopefully will lead to better therapies. PMID:26191527

  2. [Adenosine deaminase in experimental trypanosomiasis: future implications].

    PubMed

    Pérez-Aguilar, Mary Carmen; Rondón-Mercado, Rocío

    2015-09-01

    The adenosine deaminase represents a control point in the regulation of extracellular adenosine levels, thus playing a critical role in the modulation of purinergic responses to certain pathophysiological events. Several studies have shown that serum and plasma enzyme levels are elevated in some diseases caused by microorganisms, which may represent a compensatory mechanism due to the elevated levels of adenosine and the release of inflammatory mediators. Recent research indicates that adenosine deaminase activity decreases and affects hematological parameters of infected animals with Trypanosoma evansi, so that such alterations could have implications in the pathogenesis of the disease. In addition, the enzyme has been detected in this parasite; allowing the inference that it could be associated with the vital functions of the same, similar to what occurs in mammals. This knowledge may be useful in the association of chemotherapy with specific inhibitors of the enzyme in future studies.

  3. Purine metabolism in adenosine deaminase deficiency.

    PubMed Central

    Mills, G C; Schmalstieg, F C; Trimmer, K B; Goldman, A S; Goldblum, R M

    1976-01-01

    Purine and pyrimidine metabolites were measured in erythrocytes, plasma, and urine of a 5-month-old infant with adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4) deficiency. Adenosine and adenine were measured using newly devised ion exchange separation techniques and a sensitive fluorescence assay. Plasma adenosine levels were increased, whereas adenosine was normal in erythrocytes and not detectable in urine. Increased amounts of adenine were found in erythrocytes and urine as well as in the plasma. Erythrocyte adenosine 5'-monophosphate and adenosine diphosphate concentrations were normal, but adenosine triphosphate content was greatly elevated. Because of the possibility of pyrimidine starvation, pyrimidine nucleotides (pyrimidine coenzymes) in erythrocytes and orotic acid in urine were measured. Pyrimidine nucleotide concentrations were normal, while orotic acid was not detected. These studies suggest that the immune deficiency associated with adenosine deaminase deficiency may be related to increased amounts of adenine, adenosine, or adenine nucleotides. PMID:1066699

  4. Ethylene is Involved in Brassinosteroids Induced Alternative Respiratory Pathway in Cucumber (Cucumis sativus L.) Seedlings Response to Abiotic Stress

    PubMed Central

    Wei, Li-Jie; Deng, Xing-Guang; Zhu, Tong; Zheng, Ting; Li, Peng-Xu; Wu, Jun-Qiang; Zhang, Da-Wei; Lin, Hong-Hui

    2015-01-01

    Effects of brassinosteroids (BRs) on cucumber (Cucumis sativus L.) abiotic stresses resistance to salt, polyethylene glycol (PEG), cold and the potential mechanisms were investigated in this work. Previous reports have indicated that BRs can induce ethylene production and enhance alternative oxidase (AOX) pathway. The mechanisms whether ethylene is involved as a signal molecule which connected BR with AOX in regulating stress tolerance are still unknown. Here, we found that pretreatment with 1 μM brassinolide (BL, the most active BRs) relieved stress-caused oxidative damage in cucumber seedlings and clearly enhanced the capacity of AOX and the ethylene biosynthesis. Furthermore, transcription level of ethylene signaling biosynthesis genes including ripening-related ACC synthase1 (CSACS1), ripening-related ACC synthase2 (CSACS2), ripening-related ACC synthase3 (CSACS3), 1-aminocyclopropane-1-carboxylate oxidase1 (CSACO1), 1-aminocyclopropane-1-carboxylate oxidase2 (CSACO2), and CSAOX were increased after BL treatment. Importantly, the application of the salicylhydroxamic acid (SHAM, AOX inhibitor) and ethylene biosynthesis inhibitor aminooxyacetic acid (AOA) decreased plant resistance to environmental stress by blocking BRs-induced alternative respiration. Taken together, our results demonstrated that ethylene was involved in BRs-induced AOX activity which played important roles in abiotic stresses tolerance in cucumber seedlings. PMID:26617622

  5. Effect of the defoliant thidiazuron on ethylene evolution from mung bean hypocotyl segments.

    PubMed

    Suttle, J C

    1984-08-01

    The effect of the defoliant thidiazuron (N-phenyl-N'1,2,3-thiadiazol-5-ylurea) on ethylene evolution from etiolated mung bean hypocotyl segments was examined. Treatment of hypocotyl segments with concentrations of thidiazuron equal to or greater than 30 nanomolar stimulated ethylene evolution. Increased rates of ethylene evolution from thidiazuron-treated tissues could be detected within 90 minutes of treatment and persisted up to 30 hours after treatment. Radioactive methionine was readily taken up by thidiazuron-treated tissues and was converted to ethylene, 1-aminocyclopropane-1-carboxylic acid (ACC) and an acidic conjugate of ACC. Aminoethoxyvinylglycine, aminooxyacetic acid, cobalt chloride, and alpha-aminoisobutyric acid reduced ethylene evolution from treated tissues. An increase in the endogenous content of free ACC coincided with the increase in ethylene evolution following thidiazuron treatment. Uptake and conversion of exogenous ACC to ethylene were not affected by thidiazuron treatment. No increases in the extractable activities of ACC synthase were detected following thidiazuron treatment.

  6. Engineering and optimising deaminase fusions for genome editing

    PubMed Central

    Yang, Luhan; Briggs, Adrian W.; Chew, Wei Leong; Mali, Prashant; Guell, Marc; Aach, John; Goodman, Daniel Bryan; Cox, David; Kan, Yinan; Lesha, Emal; Soundararajan, Venkataramanan; Zhang, Feng; Church, George

    2016-01-01

    Precise editing is essential for biomedical research and gene therapy. Yet, homology-directed genome modification is limited by the requirements for genomic lesions, homology donors and the endogenous DNA repair machinery. Here we engineered programmable cytidine deaminases and test if we could introduce site-specific cytidine to thymidine transitions in the absence of targeted genomic lesions. Our programmable deaminases effectively convert specific cytidines to thymidines with 13% efficiency in Escherichia coli and 2.5% in human cells. However, off-target deaminations were detected more than 150 bp away from the target site. Moreover, whole genome sequencing revealed that edited bacterial cells did not harbour chromosomal abnormalities but demonstrated elevated global cytidine deamination at deaminase intrinsic binding sites. Therefore programmable deaminases represent a promising genome editing tool in prokaryotes and eukaryotes. Future engineering is required to overcome the processivity and the intrinsic DNA binding affinity of deaminases for safer therapeutic applications. PMID:27804970

  7. Role of oxidative stress and the activity of ethylene biosynthetic enzymes on the formation of spongy tissue in 'Alphonso' mango.

    PubMed

    Nagamani, J E; Shivashankara, K S; Roy, T K

    2010-06-01

    Spongy tissue formation in 'Alphonso' mangoes (Mangifera indica L) is a major national problem leading to loss for farmers and traders. Spongy tissue is whitish sponge like tissue formed near the seed with insipid taste and off odour. Lipid peroxidation of membranes as studied by malondialdehyde formation was significantly higher in spongy tissue. Activities of antioxidative enzymes like superoxide dismutase, catalase, peroxidase and polyphenol oxidase were lower in spongy tissue. Among the antioxidative enzymes, activities of catalase and peroxidases were severely reduced leading to membrane damage in spongy tissue. A significant reduction in 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase and accumulation of ACC was also observed in spongy tissue. However, ACC synthase activity in spongy tissue was more compared to healthy tissue. Results indicate that the membrane peroxidation leading to lower activity of ACC oxidase might lead to the formation of spongy tissue in 'Alphonso' mango.

  8. Multivalent Induction of Biodegradative Threonine Deaminase

    PubMed Central

    Yui, Yoshiki; Watanabe, Yasuyoshi; Ito, Seiji; Shizuta, Yutaka; Hayaishi, Osamu

    1977-01-01

    To determine the inducer(s) of the biodegradative threonine deaminase in Escherichia coli, the effects of various amino acids on the synthesis of this enzyme were investigated. The complex medium used hitherto for the enzyme induction can be completely replaced by a synthetic medium composed of 18 natural amino acids. In this synthetic medium, the omission of each of the seven amino acids threonine, serine, aspartic acid, methionine, valine, leucine, and arginine resulted in the greatest loss of enzyme formation. These seven amino acids did not significantly influence the uptake of other amino acids into the cells. Furthermore, they did not stimulate the conversion of inactive enzyme into an active form, since they did not affect the enzyme level in cells in which protein synthesis was inhibited by chloramphenicol. Threonine, serine, aspartic acid, and methionine failed to stimulate enzyme production in cells in which messenger ribonucleic acid synthesis was arrested by rifampin, whereas valine, leucine, and arginine stimulated enzyme synthesis under the same conditions. Therefore, the first four amino acids appear to act as inducers of the biodegradative threonine deaminase in E. coli and the last three amino acids appear to be amplifiers of enzyme production. The term “multivalent induction” has been proposed for this type of induction, i.e., enzyme induction only by the simultaneous presence of several amino acids. PMID:334736

  9. Neuroprotective effects of adenosine deaminase in the striatum

    PubMed Central

    Tamura, Risa; Satoh, Yasushi; Nonoyama, Shigeaki; Nishida, Yasuhiro; Nibuya, Masashi

    2016-01-01

    Adenosine deaminase (ADA) is a ubiquitous enzyme that catabolizes adenosine and deoxyadenosine. During cerebral ischemia, extracellular adenosine levels increase acutely and adenosine deaminase catabolizes the increased levels of adenosine. Since adenosine is a known neuroprotective agent, adenosine deaminase was thought to have a negative effect during ischemia. In this study, however, we demonstrate that adenosine deaminase has substantial neuroprotective effects in the striatum, which is especially vulnerable during cerebral ischemia. We used temporary oxygen/glucose deprivation (OGD) to simulate ischemia in rat corticostriatal brain slices. We used field potentials as the primary measure of neuronal damage. For stable and efficient electrophysiological assessment, we used transgenic rats expressing channelrhodopsin-2, which depolarizes neurons in response to blue light. Time courses of electrically evoked striatal field potential (eFP) and optogenetically evoked striatal field potential (optFP) were recorded during and after oxygen/glucose deprivation. The levels of both eFP and optFP decreased after 10 min of oxygen/glucose deprivation. Bath-application of 10 µg/ml adenosine deaminase during oxygen/glucose deprivation significantly attenuated the oxygen/glucose deprivation-induced reduction in levels of eFP and optFP. The number of injured cells decreased significantly, and western blot analysis indicated a significant decrease of autophagic signaling in the adenosine deaminase-treated oxygen/glucose deprivation slices. These results indicate that adenosine deaminase has protective effects in the striatum. PMID:26746865

  10. Lattice QCD simulations using the OpenACC platform

    NASA Astrophysics Data System (ADS)

    Majumdar, Pushan

    2016-10-01

    In this article we will explore the OpenACC platform for programming Graphics Processing Units (GPUs). The OpenACC platform offers a directive based programming model for GPUs which avoids the detailed data flow control and memory management necessary in a CUDA programming environment. In the OpenACC model, programs can be written in high level languages with OpenMP like directives. We present some examples of QCD simulation codes using OpenACC and discuss their performance on the Fermi and Kepler GPUs.

  11. Acc homoeoloci and the evolution of wheat genomes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We analyzed the DNA sequences of BACs from many wheat libraries containing the Acc-1 and Acc-2 loci, encoding the plastid and cytosolic forms of the enzyme acetyl-CoA carboxylase, to gain understanding of the evolution of these genes and the origin of the three genomes in modern hexaploid wheat. Mor...

  12. Enhanced ethylene emissions from red and Norway spruce exposed to acidic mists

    SciTech Connect

    Chen, Yimin; Wellburn, A.R. )

    1989-09-01

    Acidic cloudwater is believed to cause needle injury and to decrease winter hardiness in conifers. During simulations of these adverse conditions, rates of ethylene emissions from and levels of 1-aminocyclopropane-1-carboxylic acid (ACC) in both red and Norway spruce needles increased as a result of treatment with acidic mists but amounts of 1-malonyl(amino)cyclopropane-1-carboxylic acid remained unchanged. However, release of significant quantities of ethylene by another mechanism independent of ACC was also detected from brown needles. Application of exogenous plant growth regulators such as auxin, kinetic, abscisic acid and gibberellic acid (each 0.1 millimolar) had no obvious effects on the rates of basal or stress ethylene production from Norway spruce needles. The kinetics of ethylene formation by acidic mist-stressed needles suggest that there is no active inhibitive mechanism in spruce to prevent stress ethylene being released once ACC has been formed.

  13. Recycling of 5'-methylthioadenosine-ribose carbon atoms into methionine in tomato tissue in relation to ethylene production.

    PubMed

    Wang, S Y; Adams, D O; Lieberman, M

    1982-07-01

    The ribose moiety of 5'-methylthioadenosine (MTA) is metabolized to form the four-carbon unit (2-aminobutyrate) of methionine in tomato tissue (Lycopersicon esculentum Mill., cv. Pik Red). When [U-(14)C-adenosine] MTA was administered to tomato tissue slices, label was recovered in 5-methylthioribose (MTR), methionine, 1-aminocyclopropane-1-carboxylic acid (ACC), C(2)H(4) and other unidentified compounds. However, when [U-(14)C-ribose]MTR was administered, radioactivities were recovered in methionine, ACC and C(2)H(4), but not MTA. This suggests that C(2)H(4) formed in tomato pericarp tissue may be derived from the ribose portion of MTA via MTR, methionine and ACC. The conversion of MTR to methionine is not inhibited by aminoethoxyvinylglycine (AVG), but is O(2) dependent. These data present a new salvage pathway for methionine biosynthesis which may be important in relation to polyamine and ethylene biosynthesis in tomato tissue.

  14. Arachidicoccus rhizosphaerae gen. nov., sp. nov., a plant-growth-promoting bacterium in the family Chitinophagaceae isolated from rhizosphere soil.

    PubMed

    Madhaiyan, Munusamy; Poonguzhali, Selvaraj; Senthilkumar, Murugaiyan; Pragatheswari, Dhandapani; Lee, Jung-Sook; Lee, Keun-Chul

    2015-02-01

    Three novel bacterial strains, designated Vu-144(T), Vu-7 and Vu-35, were isolated on minimal medium from rhizosphere soil of field-grown cowpea and subjected to a taxonomic study using a polyphasic approach. Cells of the strains were Gram-stain-negative, non-motile, non-spore-forming, coccoid rods, and formed non-pigmented colonies. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain Vu-144(T) was affiliated with an uncultivated lineage of the phylum Bacteroidetes. Its closest phylogenetic neighbour was the recently described species Niastella populi, a member of the family Chitinophagaceae, with just 90.7 % sequence similarity to the type strain. The only isoprenoid quinone detected was menaquinone 7 (MK-7). The fatty acid profiles showed large amounts of iso-C15 : 0, iso-C17 : 0 3-OH and iso-C15 : 1 G and minor amounts of summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH), C16 : 0 and other fatty acids, allowing the differentiation of the strains from other genera. The G+C content of the genomic DNA of the three strains ranged from 43.1 to 44.3 mol%. In addition to phosphatidylethanolamine, the major polar lipids were three unidentified aminophospholipids (APL1-APL3), two unidentified phospholipids (PL1, PL2) and three unidentified lipids (UL1-UL3). Biochemical test patterns also differed from those of Niastella populi and members of other genera. All three isolates showed plant-growth-promoting properties, e.g. the ability to produce indole-3-acetic acid and NH3 and to solubilize phosphate, utilized 1-aminocyclopropane 1-carboxylate (ACC) as a sole source of nitrogen and possessed the ACC deaminase enzyme. The novel isolates readily colonized roots and stimulated growth of tomato and cowpea under glasshouse conditions. Inoculated plants showed a 45-60 % increase in dry matter weight with respect to uninoculated controls. On the basis of the evidence from our polyphasic study, isolate Vu-144(T) represents a novel genus and species in

  15. Studies on guanine deaminase and its inhibitors in rat tissue

    PubMed Central

    Kumar, S.; Josan, V.; Sanger, K. C. S.; Tewari, K. K.; Krishnan, P. S.

    1967-01-01

    1. In kidney, but not in rat whole brain and liver, guanine-deaminase activity was localized almost exclusively in the 15000g supernatant fraction of iso-osmotic sucrose homogenates. However, as in brain and liver, the enzymic activity recovered in the supernatant was higher than that in the whole homogenate. The particulate fractions of kidney, especially the heavy mitochondria, brought about powerful inhibition of the supernatant guanine-deaminase activity. 2. In spleen, as in kidney, guanine-deaminase activity was localized in the 15000g supernatant fraction of iso-osmotic sucrose homogenates. However, the particulate fractions did not inhibit the activity of the supernatant. 3. Guanine-deaminase activity in rat brain was absent from the cerebellum and present only in the cerebral hemispheres. The inhibitor of guanine deaminase was located exclusively in the cerebellum, where it was associated with the particles sedimenting at 5000g from sucrose homogenates. 4. Homogenates of cerebral hemispheres, the separated cortex or the remaining portion of the hemispheres had significantly higher guanine-deaminase activity than homogenates of whole brain. The enzymic activity of the subcellular particulate fractions was nearly the same. 5. Guanine deaminase was purified from the 15000g supernatant of sucrose homogenates of whole brain. The enzyme separated as two distinct fractions, A and B, on DEAE-cellulose columns. 6. The guanine-deaminase activity of the light-mitochondrial fraction of whole brain was fully exposed and solubilized by treatment with Triton X-100, and partially purified. 7. Tested in the form of crude preparations, the inhibitor from kidney did not act on the brain and liver supernatant enzymes and the inhibitor from cerebellum did not act on kidney enzyme, but the inhibitor from liver acted on both brain and kidney enzyme. 8. The inhibitor of guanine deaminase was purified from the heavy mitochondria of whole brain and liver and the 5000g residue of

  16. The catalase activity of diiron adenine deaminase

    SciTech Connect

    Kamat S. S.; Swaminathan S.; Holmes-Hampton, G. P.; Bagaria, A.; Kumaran, D.; Tichy, S. E.; Gheyi, T.; Zheng, X.; Bain, K.; Groshong, C.; Emtage, S.; Sauder, J. M.; Burley, S. K.; Lindahl, P. A.; Raushel, F. M.

    2011-12-01

    Adenine deaminase (ADE) from the amidohydrolase superfamily (AHS) of enzymes catalyzes the conversion of adenine to hypoxanthine and ammonia. Enzyme isolated from Escherichia coli was largely inactive toward the deamination of adenine. Molecular weight determinations by mass spectrometry provided evidence that multiple histidine and methionine residues were oxygenated. When iron was sequestered with a metal chelator and the growth medium supplemented with Mn{sup 2+} before induction, the post-translational modifications disappeared. Enzyme expressed and purified under these conditions was substantially more active for adenine deamination. Apo-enzyme was prepared and reconstituted with two equivalents of FeSO{sub 4}. Inductively coupled plasma mass spectrometry and Moessbauer spectroscopy demonstrated that this protein contained two high-spin ferrous ions per monomer of ADE. In addition to the adenine deaminase activity, [Fe{sup II}/Fe{sup II}]-ADE catalyzed the conversion of H{sub 2}O{sub 2} to O{sub 2} and H{sub 2}O. The values of k{sub cat} and k{sub cat}/K{sub m} for the catalase activity are 200 s{sup -1} and 2.4 x 10{sup 4} M{sup -1} s{sup -1}, respectively. [Fe{sup II}/Fe{sup II}]-ADE underwent more than 100 turnovers with H{sub 2}O{sub 2} before the enzyme was inactivated due to oxygenation of histidine residues critical for metal binding. The iron in the inactive enzyme was high-spin ferric with g{sub ave} = 4.3 EPR signal and no evidence of anti-ferromagnetic spin-coupling. A model is proposed for the disproportionation of H{sub 2}O{sub 2} by [Fe{sup II}/Fe{sup II}]-ADE that involves the cycling of the binuclear metal center between the di-ferric and di-ferrous oxidation states. Oxygenation of active site residues occurs via release of hydroxyl radicals. These findings represent the first report of redox reaction catalysis by any member of the AHS.

  17. 24 CFR 969.105 - Extension of ACC upon payment of operating subsidy.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... for which Operating Subsidy is paid with respect to the project. (b) Consolidated ACC. Where a single... Consolidated ACC. In any event, no Operating Subsidy payable under a Consolidated ACC or otherwise shall...

  18. [Gene therapy for adenosine deaminase deficiency].

    PubMed

    Sakiyama, Yukio; Ariga, Tadashi; Ohtsu, Makoto

    2005-03-01

    A four year-old boy with adenosine deaminase (ADA-) deficient severe combined immunodeficiency(SCID) receiving PEG-ADA was treated under a gene therapy protocol targeting peripheral blood lymphocytes (PBLs) in 1995. After eleven infusions of autologous PBLs transduced with retroviral vector LASN encoding ADAcDNA, he exhibited increased levels of the CD8+ T lymphocytes, serum immunoglobulin, specific antibodies and delayed type hypersensitivity skin tests. Follow-up studies also provided evidence of long-term persistence and function of transduced PBLs with improvement in the immune function. However, the therapeutic effect of this gene therapy has been difficult to assess because of the concomitant treatment of PEG-ADA. Two ADA-SCID patients have been currently treated with autologous bone marrow CD34+ cells engineered with a retroviral vector GCsapM-ADA after discontinuation of PEG-ADA. The restoration of intracellular ADA enzymatic activity in lymphocytes and granulocytes resulted in correction of the systemic toxicity and liver function in the absence of PEG-ADA treatment. Both patients are at home where they are clinically well, and they do not experience adversed effect, with follow up being 12 months after CD34+ cells gene therapy.

  19. Altered AMP deaminase activity may extend postmortem glycolysis.

    PubMed

    England, E M; Matarneh, S K; Scheffler, T L; Wachet, C; Gerrard, D E

    2015-04-01

    Postmortem energy metabolism drives hydrogen accumulation in muscle and results in a fairly constant ultimate pH. Extended glycolysis results in adverse pork quality and may be possible with greater adenonucleotide availability postmortem. We hypothesized that slowing adenonucleotide removal by reducing AMP deaminase activity would extend glycolysis and lower the ultimate pH of muscle. Longissimus muscle samples were incorporated into an in vitro system that mimics postmortem glycolysis with or without pentostatin, an AMP deaminase inhibitor. Pentostatin lowered ultimate pH and increased lactate and glucose 6-phosphate with time. Based on these results and that AMPK γ3(R200Q) mutated pigs (RN⁻) produce low ultimate pH pork, we hypothesized AMP deaminase abundance and activity would be lower in RN⁻ muscle than wild-type. RN⁻ muscle contained lower AMP deaminase abundance and activity. These data show that altering adenonucleotide availability postmortem can extend postmortem pH decline and suggest that AMP deaminase activity may, in part, contribute to the low ultimate pH observed in RN⁻ pork.

  20. Adrenocortical cancer (ACC) - literature overview and own experience.

    PubMed

    Dworakowska, Dorota; Drabarek, Agata; Wenzel, Ingrid; Babińska, Anna; Świątkowska-Stodulska, Renata; Sworczak, Krzysztof

    2014-01-01

    Adrenocortical carcinoma (ACC) is a malignant endocrine tumour. The rarity of the disease has stymied therapeutic development. Age distribution shows two peaks: the first and fifth decades of life, with children and women more frequently affected. Although 60-70% of ACCs are biochemically found to overproduce hormones, it is not clinically apparent in many cases. If present, endocrine symptoms include signs of hypercortisolaemia, virilisation or gynaecomastia. ACC carries a poor prognosis, and a cure can be achieved only by complete surgical resection. Mitotane is used both as an adjuvant treatment and also in non-operative patients. The role of radio- and chemotherapy is still controversial. The post-operative disease free survival is low and oscillates around 30% due to high tumour recurrence rate. The diagnosis is based on tumour histological assessment with the use of the Weiss score, however urinary steroid profiling (if available) can serve to differentiate between ACC and other adrenal tumours. Conventional prognostic markers in ACC include stage and grade of disease, and, as currently reported, the presence of hypercortisolaemia. Molecular analysis has had a significant impact on the understanding of the pathogenetic mechanism of ACC development and the evaluation of prognostic and predictive markers, among which alterations of the IGF system, the Wnt pathway, p53 and molecules involved in cancer cell invasion properties and angiogenesis seem to be very promising. We here summarise our own experience related to the management of ACC and present a literature overview. We have not aimed to include a detailed summary of the molecular alterations biology described in ACC, as this has already been addressed in other papers.

  1. Identification, expression, and characterization of Escherichia coli guanine deaminase.

    PubMed

    Maynes, J T; Yuan, R G; Snyder, F F

    2000-08-01

    Using the human cDNA sequence corresponding to guanine deaminase, the Escherichia coli genome was scanned using the Basic Local Alignment Search Tool (BLAST), and a corresponding 439-residue open reading frame of unknown function was identified as having 36% identity to the human protein. The putative gene was amplified, subcloned into the pMAL-c2 vector, expressed, purified, and characterized enzymatically. The 50.2-kDa protein catalyzed the conversion of guanine to xanthine, having a K(m) of 15 microM with guanine and a k(cat) of 3.2 s(-1). The bacterial enzyme shares a nine-residue heavy metal binding site with human guanine deaminase, PG[FL]VDTHIH, and was found to contain approximately 1 mol of zinc per mol of subunit of protein. The E. coli guanine deaminase locus is 3' from an open reading frame which shows homology to a bacterial purine base permease.

  2. Identification, Expression, and Characterization of Escherichia coli Guanine Deaminase

    PubMed Central

    Maynes, Jason T.; Yuan, Richard G.; Snyder, Floyd F.

    2000-01-01

    Using the human cDNA sequence corresponding to guanine deaminase, the Escherichia coli genome was scanned using the Basic Local Alignment Search Tool (BLAST), and a corresponding 439-residue open reading frame of unknown function was identified as having 36% identity to the human protein. The putative gene was amplified, subcloned into the pMAL-c2 vector, expressed, purified, and characterized enzymatically. The 50.2-kDa protein catalyzed the conversion of guanine to xanthine, having a Km of 15 μM with guanine and a kcat of 3.2 s−1. The bacterial enzyme shares a nine-residue heavy metal binding site with human guanine deaminase, PG[FL]VDTHIH, and was found to contain approximately 1 mol of zinc per mol of subunit of protein. The E. coli guanine deaminase locus is 3′ from an open reading frame which shows homology to a bacterial purine base permease. PMID:10913105

  3. Unique properties of Plasmodium falciparum porphobilinogen deaminase.

    PubMed

    Nagaraj, Viswanathan Arun; Arumugam, Rajavel; Gopalakrishnan, Bulusu; Jyothsna, Yeleswarapu Sri; Rangarajan, Pundi N; Padmanaban, Govindarajan

    2008-01-04

    The hybrid pathway for heme biosynthesis in the malarial parasite proposes the involvement of parasite genome-coded enzymes of the pathway localized in different compartments such as apicoplast, mitochondria, and cytosol. However, knowledge on the functionality and localization of many of these enzymes is not available. In this study, we demonstrate that porphobilinogen deaminase encoded by the Plasmodium falciparum genome (PfPBGD) has several unique biochemical properties. Studies carried out with PfPBGD partially purified from parasite membrane fraction, as well as recombinant PfPBGD lacking N-terminal 64 amino acids expressed and purified from Escherichia coli cells (DeltaPfPBGD), indicate that both the proteins are catalytically active. Surprisingly, PfPBGD catalyzes the conversion of porphobilinogen to uroporphyrinogen III (UROGEN III), indicating that it also possesses uroporphyrinogen III synthase (UROS) activity, catalyzing the next step. This obviates the necessity to have a separate gene for UROS that has not been so far annotated in the parasite genome. Interestingly, DeltaPfP-BGD gives rise to UROGEN III even after heat treatment, although UROS from other sources is known to be heat-sensitive. Based on the analysis of active site residues, a DeltaPfPBGDL116K mutant enzyme was created and the specific activity of this recombinant mutant enzyme is 5-fold higher than DeltaPfPBGD. More interestingly, DeltaPfPBGDL116K catalyzes the formation of uroporphyrinogen I (UROGEN I) in addition to UROGEN III, indicating that with increased PBGD activity the UROS activity of PBGD may perhaps become rate-limiting, thus leading to non-enzymatic cyclization of preuroporphyrinogen to UROGEN I. PfPBGD is localized to the apicoplast and is catalytically very inefficient compared with the host red cell enzyme.

  4. Adenosine deaminase from Streptomyces coelicolor: recombinant expression, purification and characterization.

    PubMed

    Pornbanlualap, Somchai; Chalopagorn, Pornchanok

    2011-08-01

    The sequencing of the genome of Streptomyces coelicolor A3(2) identified seven putative adenine/adenosine deaminases and adenosine deaminase-like proteins, none of which have been biochemically characterized. This report describes recombinant expression, purification and characterization of SCO4901 which had been annotated in data bases as a putative adenosine deaminase. The purified putative adenosine deaminase gives a subunit Mr=48,400 on denaturing gel electrophoresis and an oligomer molecular weight of approximately 182,000 by comparative gel filtration. These values are consistent with the active enzyme being composed of four subunits with identical molecular weights. The turnover rate of adenosine is 11.5 s⁻¹ at 30 °C. Since adenine is deaminated ∼10³ slower by the enzyme when compared to that of adenosine, these data strongly show that the purified enzyme is an adenosine deaminase (ADA) and not an adenine deaminase (ADE). Other adenine nucleosides/nucleotides, including 9-β-D-arabinofuranosyl-adenine (ara-A), 5'-AMP, 5'-ADP and 5'-ATP, are not substrates for the enzyme. Coformycin and 2'-deoxycoformycin are potent competitive inhibitors of the enzyme with inhibition constants of 0.25 and 3.4 nM, respectively. Amino acid sequence alignment of ScADA with ADAs from other organisms reveals that eight of the nine highly conserved catalytic site residues in other ADAs are also conserved in ScADA. The only non-conserved residue is Asn317, which replaces Asp296 in the murine enzyme. Based on these data, it is suggested here that ADA and ADE proteins are divergently related enzymes that have evolved from a common α/β barrel scaffold to catalyze the deamination of different substrates, using a similar catalytic mechanism.

  5. Optimatization of transient transformation methods to study gene expression in Musa acuminata (AAA group) cultivar Ambon Lumut

    NASA Astrophysics Data System (ADS)

    Prayuni, Kinasih; Dwivany, Fenny M.

    2015-09-01

    Banana is classified as a climateric fruit, whose ripening is regulated by ethylene. Ethylene is synthesized from ACC (1-aminocyclopropane-1-carboxylic acid) by ACC oxidase enzyme which is encoded by ACO gene. Controling an important gene expression in ethylene biosynthesis pathway has became a target to delay the ripening process. Therefore in the previous study we have designed a MaACO-RNAi construct to control MaACO gene expression. In this research, we study the effectiveness of different transient transformation methods to deliver the construct. Direct injection, with or no vaccum infiltration methods were used to deliver MaACO-RNAi construct. All of the methods succesfully deliver the construct into banana fruits based on RT-PCR result.

  6. Selenium delays tomato fruit ripening by inhibiting ethylene biosynthesis and enhancing the antioxidant defense system.

    PubMed

    Zhu, Zhu; Chen, Yanli; Shi, Guoqing; Zhang, Xueji

    2017-03-15

    The antioxidant activity of selenium (Se) detoxifies reactive oxygen species (ROS) in plants and animals. In the present study, we elucidated the mechanism underlying Se induced fruit development and ripening. Our study showed that foliar pretreatment with 1mgL(-1) sodium selenate effectively delayed fruit ripening and maintained fruit quality. Gene expression studies revealed that the repression of ethylene biosynthetic genes 1-aminocyclopropane-1-carboxylic acid (ACC) synthase and ACC oxidase decreased ethylene production and respiration rate. Moreover, Se treatment probably boosted the antioxidant defense system to reduce ROS generation and membrane damage. The enhanced antioxidative effect was attributed to higher glutathione content and increased activity of enzymes such as glutathione peroxidase and glutathione reductase. The upregulation of respiratory burst oxidase homologue genes in tomato fruit may also contribute to the enhanced antioxidative effect. Selenium treatment represents a promising strategy for delaying ripening and extending the shelf life of tomato fruit.

  7. Guanine Deaminase Functions as Dihydropterin Deaminase in the Biosynthesis of Aurodrosopterin, a Minor Red Eye Pigment of Drosophila*

    PubMed Central

    Kim, Jaekwang; Park, Sang Ick; Ahn, Chiyoung; Kim, Heuijong; Yim, Jeongbin

    2009-01-01

    Dihydropterin deaminase, which catalyzes the conversion of 7,8-dihydropterin to 7,8-dihydrolumazine, was purified 5850-fold to apparent homogeneity from Drosophila melanogaster. Its molecular mass was estimated to be 48 kDa by gel filtration and SDS-PAGE, indicating that it is a monomer under native conditions. The pI value, temperature, and optimal pH of the enzyme were 5.5, 40 °C, and 7.5, respectively. Interestingly the enzyme had much higher activity for guanine than for 7,8-dihydropterin. The specificity constant (kcat/Km) for guanine (8.6 × 106 m−1·s−1) was 860-fold higher than that for 7,8-dihydropterin (1.0 × 104 m−1·s−1). The structural gene of the enzyme was identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis as CG18143, located at region 82A1 on chromosome 3R. The cloned and expressed CG18143 exhibited both 7,8-dihydropterin and guanine deaminase activities. Flies with mutations in CG18143, SUPor-P/Df(3R)A321R1 transheterozygotes, had severely decreased activities in both deaminases compared with the wild type. Among several red eye pigments, the level of aurodrosopterin was specifically decreased in the mutant, and the amount of xanthine and uric acid also decreased considerably to 76 and 59% of the amounts in the wild type, respectively. In conclusion, dihydropterin deaminase encoded by CG18143 plays a role in the biosynthesis of aurodrosopterin by providing one of its precursors, 7,8-dihydrolumazine, from 7,8-dihydropterin. Dihydropterin deaminase also functions as guanine deaminase, an important enzyme for purine metabolism. PMID:19567870

  8. 24 CFR 905.302 - Timely submission of the CF ACC amendment by the PHA.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 24 Housing and Urban Development 4 2014-04-01 2014-04-01 false Timely submission of the CF ACC... Requirements § 905.302 Timely submission of the CF ACC amendment by the PHA. Upon being provided with a CF ACC Amendment from HUD, the PHA must sign and date the CF ACC Amendment and return it to HUD by the...

  9. Plant growth-promoting traits of epiphytic and endophytic yeasts isolated from rice and sugar cane leaves in Thailand.

    PubMed

    Nutaratat, Pumin; Srisuk, Nantana; Arunrattiyakorn, Panarat; Limtong, Savitree

    2014-08-01

    A total of 1035 yeast isolates, obtained from rice and sugar cane leaves, were screened primarily for indole-3-acetic acid (IAA) production. Thirteen isolates were selected, due to their IAA production ranging from 1.2 to 29.3 mg g(-)(1) DCW. These isolates were investigated for their capabilities of calcium phosphate and ZnO(3) solubilisation, and also for production of NH(3), polyamine, and siderophore. Their 1-aminocyclopropane-1-carboxylate (ACC) deaminase, catalase and fungal cell wall-degrading enzyme activities were assessed. Their antagonism against rice fungal pathogens was also evaluated. Strain identification, based on molecular taxonomy, of the thirteen yeast isolates revealed that four yeast species - i.e. Hannaella sinensis (DMKU-RP45), Cryptococcus flavus (DMKU-RE12, DMKU-RE19, DMKU-RE67, and DMKU-RP128), Rhodosporidium paludigenum (DMKU-RP301) and Torulaspora globosa (DMKU-RP31) - were capable of high IAA production. Catalase activity was detected in all yeast strains tested. The yeast R. paludigenum DMKU-RP301 was the best IAA producer, yielding 29.3 mg g(-)(1) DCW, and showed the ability to produce NH3 and siderophore. Different levels of IAA production (7.2-9.7 mg g(-)(1) DCW) were found in four strains of C. flavus DMKU-RE12, DMKU-RE19, and DMKU-RE67, which are rice leaf endophytes, and strain DMKU-RP128, which is a rice leaf epiphyte. NH(3) production and carboxymethyl cellulase (CMCase) activity was also detected in these four strains. Antagonism to fungal plant pathogens and production of antifungal volatile compounds were exhibited in T. globosa DMKU-RP31, as well as a moderate level of IAA production (4.9 mg g(-)(1) DCW). The overall results indicated that T. globosa DMKU-RP31 might be used in two ways: enhancing plant growth and acting as a biocontrol agent. In addition, four C. flavus were also found to be strains of interest for optimal IAA production.

  10. Quantitative changes in adenosine deaminase isoenzymes in human colorectal adenocarcinomas.

    PubMed

    ten Kate, J; Wijnen, J T; van der Goes, R G; Quadt, R; Griffioen, G; Bosman, F T; Khan, P M

    1984-10-01

    Several reports have suggested that a decrease or absence of adenosine deaminase complexing protein (ADCP) is consistently associated with cancer. However, in other studies, decreased as well as increased ADCP levels were found. In the present study, we investigated ADCP levels in 37 colorectal adenocarcinomas and correlated the results with clinicopathological characteristics in individual carcinomas. The levels of adenosine deaminase (EC 3.5.4.4) and soluble ADCP were determined in tissue samples by, respectively, a spectrophotometric assay and an ADCP specific radioimmunoassay. The values in the individual tumors were compared with their histological characteristics, such as degree of differentiation, nuclear grading, and the preoperative plasma carcinoembryonic antigen levels in the patients. It was found that ADCP was decreased in about a third of the tumors but unaltered or even increased in others. However, there was an overall 40% increase of the adenosine deaminase activity in the tumors compared to normal tissue. There seems to be no simple correlation between any of the clinicopathological parameters and the ADCP or adenosine deaminase levels. Methods detecting ADCP at single cell level might be helpful in exploring its potential use as a cancer-associated marker.

  11. Enterobacter asburiae KUNi5, a Nickel Resistant Bacterium for Possible Bioremediation of Nickel Contaminated Sites.

    PubMed

    Paul, Anirudha; Mukherjee, Samir Kumar

    2016-01-01

    Nickel resistant bacterial strain Enterobacter asburiae KUNi5 was isolated and showed resistance up to 15 mM and could remove Ni optimally better at 37 degrees C and pH 7. Maximum removal was found at initial concentration of 0.5 to 2 mM, however, growth and Ni removal were affected by other heavy metals. Major amount of the metal was accumulated in the membrane fractions and certain negatively charged groups were found responsible for Ni binding. KUNi5 could also produce 1-aminocyclopropane-1-carboxylate deaminase, indole-acetic acid and siderophore. It seems that KUNi5 could be a possible candidate for Ni detoxification and plant growth promotion in Ni-contaminated field.

  12. Plant growth-promoting and rhizosphere-competent Acinetobacter rhizosphaerae strain BIHB 723 from the cold deserts of the Himalayas.

    PubMed

    Gulati, Arvind; Vyas, Pratibha; Rahi, Praveen; Kasana, Ramesh Chand

    2009-04-01

    A phosphate-solubilizing bacterial strain BIHB 723 isolated from the rhizosphere of Hippophae rhamnoides was identified as Acinetobacter rhizosphaerae on the basis of phenotypic characteristics, carbon source utilization pattern, fatty acid methyl esters analysis, and 16S rRNA gene sequence. The strain exhibited the plant growth-promoting attributes of inorganic and organic phosphate solubilization, auxin production, 1-aminocyclopropane-1-carboxylate deaminase activity, ammonia generation, and siderophore production. A significant increase in the growth of pea, chickpea, maize, and barley was recorded for inoculations under controlled conditions. Field testing with the pea also showed a significant increment in plant growth and yield. The rifampicin mutant of the bacterial strain effectively colonized the pea rhizosphere without adversely affecting the resident microbial populations.

  13. Plant growth-promoting traits of yeasts isolated from the phyllosphere and rhizosphere of Drosera spatulata Lab.

    PubMed

    Fu, Shih-Feng; Sun, Pei-Feng; Lu, Hsueh-Yu; Wei, Jyuan-Yu; Xiao, Hong-Su; Fang, Wei-Ta; Cheng, Bai-You; Chou, Jui-Yu

    2016-03-01

    Microorganisms can promote plant growth through direct and indirect mechanisms. Compared with the use of bacteria and mycorrhizal fungi, the use of yeasts as plant growth-promoting (PGP) agents has not been extensively investigated. In this study, yeast isolates from the phyllosphere and rhizosphere of the medicinally important plant Drosera spatulata Lab. were assessed for their PGP traits. All isolates were tested for indole-3-acetic acid-, ammonia-, and polyamine-producing abilities, calcium phosphate and zinc oxide solubilizing ability, and catalase activity. Furthermore, the activities of siderophore, 1-aminocyclopropane-1-carboxylate deaminase, and fungal cell wall-degrading enzymes were assessed. The antagonistic action of yeasts against pathogenic Glomerella cingulata was evaluated. The cocultivation of Nicotiana benthamiana with yeast isolates enhanced plant growth, indicating a potential yeast-plant interaction. Our study results highlight the potential use of yeasts as plant biofertilizers under controlled and field conditions.

  14. Discovery and optimization of antibacterial AccC inhibitors

    SciTech Connect

    Cheng, Cliff C.; Shipps, Jr., Gerald W.; Yang, Zhiwei; Sun, Binyuan; Kawahata, Noriyuki; Soucy, Kyle A.; Soriano, Aileen; Orth, Peter; Xiao, Li; Mann, Paul; Black, Todd

    2010-09-17

    The biotin carboxylase (AccC) is part of the multi-component bacterial acetyl coenzyme-A carboxylase (ACCase) and is essential for pathogen survival. We describe herein the affinity optimization of an initial hit to give 2-(2-chlorobenzylamino)-1-(cyclohexylmethyl)-1H-benzo[d]imidazole-5-carboxamide (1), which was identified using our proprietary Automated Ligand Identification System (ALIS). The X-ray co-crystal structure of 1 was solved and revealed several key interactions and opportunities for further optimization in the ATP site of AccC. Structure Based Drug Design (SBDD) and parallel synthetic approaches resulted in a novel series of AccC inhibitors, exemplified by (R)-2-(2-chlorobenzylamino)-1-(2,3-dihydro-1H-inden-1-yl)-1H-imidazo[4,5-b]pyridine-5-carboxamide (40). This compound is a potent and selective inhibitor of bacterial AccC with an IC{sub 50} of 20 nM and a MIC of 0.8 {micro}g/mL against a sensitized strain of Escherichia coli (HS294 E. coli).

  15. Attenuation of exercise vasodilatation by adenosine deaminase in anaesthetized dogs.

    PubMed Central

    Goonewardene, I P; Karim, F

    1991-01-01

    1. In dogs anaesthetized with sodium pentobarbitone and artificially ventilated, the gracilis muscles were vascularly isolated and perfused at a constant flow of 28.4 +/- 4.6 ml min-1 (100 g muscle tissue)-1 (99.8 +/- 4.5% of maximum free flow, means +/- standard error of the mean (S.E.M.), n = 9). 2. Three to five minutes of electrical stimulation of the cut peripheral end of the obturator nerve (4 Hz, 6 V, 0.2 ms) resulted in muscle contraction (0.61 +/- 0.14 kg (100 g)-1 during solvent infusion and 0.56 +/- 0.10 kg (100 g)-1 during intra-arterial adenosine deaminase infusion (50 U min-1) and an immediate decrease in arterial perfusion pressure from 184.5 +/- 8.1 mmHg to 148.2 +/- 5.7 mmHg (18.7 +/- 3.4% decrease) during solvent infusion, and from 193.5 +/- 7.16 to 142.0 +/- 10.2 mmHg (25.4 +/- 6.1% decrease) during adenosine deaminase infusion 10 s after the commencement of muscle stimulation. After about 5 min of muscle contractions, the arterial perfusion pressure decreased to 120.8 +/- 7.8 mmHg (32.9 +/- 5.8% decrease) during solvent infusion, and to 152.8 +/- 11.2 mmHg (20.9 +/- 5.3% decrease) during adenosine deaminase infusion (i.e. 37.9 +/- 6.2% attenuation of the fall in arterial perfusion pressure). The time taken for 90% recovery of the arterial perfusion pressure was 72.1 +/- 10.9 s during solvent infusion, and 51.5 +/- 9.3 s during adenosine deaminase infusion (P less than 0.05). 3. Adenosine (2 x 10(-3) mol l-1) infusion in the resting muscle during solvent infusion (final concentration in arterial blood 1.3 x 10(-4) +/- 6.0 x 10(-5) mol l-1) resulted in a 34.8 +/- 7.2% fall in arterial perfusion pressure but a fall of only 7.2 +/- 1.8% during adenosine deaminase infusion (50 U min-1; P less than 0.05; n = 5) indicating that adenosine deaminase infused at 50 U min-1 was more than adequate to metabolize endogenous adenosine produced during muscle contractions. 4. These data suggest that adenosine contributes about 40% to the sustained

  16. Catalytic Mechanism and Three-Dimensional Structure of Adenine Deaminase

    SciTech Connect

    S Kamat; A Bagaria; D Kumaran; G Holmes-Hampton; H Fan; A Sali; J Sauder; S Burley; P Lindahl; et. al.

    2011-12-31

    Adenine deaminase (ADE) catalyzes the conversion of adenine to hypoxanthine and ammonia. The enzyme isolated from Escherichia coli using standard expression conditions was low for the deamination of adenine (k{sub cat} = 2.0 s{sup -1}; k{sub cat}/K{sub m} = 2.5 x 10{sup 3} M{sup -1} s{sup -1}). However, when iron was sequestered with a metal chelator and the growth medium was supplemented with Mn{sup 2+} prior to induction, the purified enzyme was substantially more active for the deamination of adenine with k{sub cat} and k{sub cat}/K{sub m} values of 200 s{sup -1} and 5 x 10{sup 5} M{sup -1} s{sup -1}, respectively. The apoenzyme was prepared and reconstituted with Fe{sup 2+}, Zn{sup 2+}, or Mn{sup 2+}. In each case, two enzyme equivalents of metal were necessary for reconstitution of the deaminase activity. This work provides the first example of any member of the deaminase subfamily of the amidohydrolase superfamily to utilize a binuclear metal center for the catalysis of a deamination reaction. [Fe{sup II}/Fe{sup II}]-ADE was oxidized to [Fe{sup III}/Fe{sup III}]-ADE with ferricyanide with inactivation of the deaminase activity. Reducing [Fe{sup III}/Fe{sup III}]-ADE with dithionite restored the deaminase activity, and thus, the diferrous form of the enzyme is essential for catalytic activity. No evidence of spin coupling between metal ions was evident by electron paramagnetic resonance or Moessbauer spectroscopy. The three-dimensional structure of adenine deaminase from Agrobacterium tumefaciens (Atu4426) was determined by X-ray crystallography at 2.2 {angstrom} resolution, and adenine was modeled into the active site on the basis of homology to other members of the amidohydrolase superfamily. On the basis of the model of the adenine-ADE complex and subsequent mutagenesis experiments, the roles for each of the highly conserved residues were proposed. Solvent isotope effects, pH-rate profiles, and solvent viscosity were utilized to propose a chemical reaction

  17. Catalytic Mechanism and Three-Dimensional Structure of Adenine Deaminase

    SciTech Connect

    Kamat, S.S.; Swaminathan, S.; Bagaria, A.; Kumaran, D.; Holmes-Hampton, G. P.; Fan, H.; Sali, A.; Sauder, J. M.; Burley, S. K.; Lindahl, P. A.; Raushel, F. M.

    2011-03-22

    Adenine deaminase (ADE) catalyzes the conversion of adenine to hypoxanthine and ammonia. The enzyme isolated from Escherichia coli using standard expression conditions was low for the deamination of adenine (k{sub cat} = 2.0 s{sup -1}; k{sub cat}/K{sub m} = 2.5 x 10{sup 3} M{sup -1} s{sup -1}). However, when iron was sequestered with a metal chelator and the growth medium was supplemented with Mn{sup 2+} prior to induction, the purified enzyme was substantially more active for the deamination of adenine with kcat and kcat/Km values of 200 s{sup -1} and 5 x 10{sup 5} M{sup -1} s{sup -1}, respectively. The apoenzyme was prepared and reconstituted with Fe{sup 2+}, Zn{sup 2+}, or Mn{sup 2+}. In each case, two enzyme equivalents of metal were necessary for reconstitution of the deaminase activity. This work provides the first example of any member of the deaminase subfamily of the amidohydrolase superfamily to utilize a binuclear metal center for the catalysis of a deamination reaction. [Fe{sup II}/Fe{sup II}]-ADE was oxidized to [Fe{sup III}/Fe{sup III}]-ADE with ferricyanide with inactivation of the deaminase activity. Reducing [Fe{sup III}/Fe{sup III}]-ADE with dithionite restored the deaminase activity, and thus, the diferrous form of the enzyme is essential for catalytic activity. No evidence of spin coupling between metal ions was evident by electron paramagnetic resonance or Moessbauer spectroscopy. The three-dimensional structure of adenine deaminase from Agrobacterium tumefaciens (Atu4426) was determined by X-ray crystallography at 2.2 {angstrom} resolution, and adenine was modeled into the active site on the basis of homology to other members of the amidohydrolase superfamily. On the basis of the model of the adenine-ADE complex and subsequent mutagenesis experiments, the roles for each of the highly conserved residues were proposed. Solvent isotope effects, pH-rate profiles, and solvent viscosity were utilized to propose a chemical reaction mechanism and the

  18. Expression of human adenosine deaminase in murine hematopoietic cells.

    PubMed Central

    Belmont, J W; MacGregor, G R; Wager-Smith, K; Fletcher, F A; Moore, K A; Hawkins, D; Villalon, D; Chang, S M; Caskey, C T

    1988-01-01

    Multiple replication-defective retrovirus vectors were tested for their ability to transfer and express human adenosine deaminase in vitro and in vivo in a mouse bone marrow transplantation model. High-titer virus production was obtained from vectors by using both a retrovirus long terminal repeat promoter and internal transcriptional units with human c-fos and herpes virus thymidine kinase promoters. After infection of primary murine bone marrow with one of these vectors, human adenosine deaminase was detected in 60 to 85% of spleen colony-forming units and in the blood of 14 of 14 syngeneic marrow transplant recipients. This system offers the opportunity to assess methods for increasing efficiency of gene transfer, for regulation of expression of foreign genes in hematopoietic progenitors, and for long-term measurement of the stability of expression in these cells. Images PMID:3072474

  19. Adenosine Deaminases Acting on RNA, RNA Editing, and Interferon Action

    PubMed Central

    George, Cyril X.; Gan, Zhenji; Liu, Yong

    2011-01-01

    Adenosine deaminases acting on RNA (ADARs) catalyze adenosine (A) to inosine (I) editing of RNA that possesses double-stranded (ds) structure. A-to-I RNA editing results in nucleotide substitution, because I is recognized as G instead of A both by ribosomes and by RNA polymerases. A-to-I substitution can also cause dsRNA destabilization, as I:U mismatch base pairs are less stable than A:U base pairs. Three mammalian ADAR genes are known, of which two encode active deaminases (ADAR1 and ADAR2). Alternative promoters together with alternative splicing give rise to two protein size forms of ADAR1: an interferon-inducible ADAR1-p150 deaminase that binds dsRNA and Z-DNA, and a constitutively expressed ADAR1-p110 deaminase. ADAR2, like ADAR1-p110, is constitutively expressed and binds dsRNA. A-to-I editing occurs with both viral and cellular RNAs, and affects a broad range of biological processes. These include virus growth and persistence, apoptosis and embryogenesis, neurotransmitter receptor and ion channel function, pancreatic cell function, and post-transcriptional gene regulation by microRNAs. Biochemical processes that provide a framework for understanding the physiologic changes following ADAR-catalyzed A-to-I ( = G) editing events include mRNA translation by changing codons and hence the amino acid sequence of proteins; pre-mRNA splicing by altering splice site recognition sequences; RNA stability by changing sequences involved in nuclease recognition; genetic stability in the case of RNA virus genomes by changing sequences during viral RNA replication; and RNA-structure-dependent activities such as microRNA production or targeting or protein–RNA interactions. PMID:21182352

  20. Severe combined immunodeficiency due to adenosine deaminase deficiency.

    PubMed

    Hussain, Waqar; Batool, Asma; Ahmed, Tahir Aziz; Bashir, Muhammad Mukarram

    2012-03-01

    Severe Combined Immunodeficiency is the term applied to a group of rare genetic disorders characterised by defective or absent T and B cell functions. Patients usually present in first 6 months of life with respiratory/gastrointestinal tract infections and failure to thrive. Among the various types of severe combined immunodeficiency, enzyme deficiencies are relatively less common. We report the case of a 6 years old girl having severe combined immunodeficiency due to adenosine deaminase deficiency.

  1. Polymorphous crystallization and diffraction of threonine deaminase from Escherichia coli.

    PubMed

    Gallagher, D T; Eisenstein, E; Fisher, K E; Zondlo, J; Chinchilla, D; Yu, H D; Dill, J; Winborne, E; Ducote, K; Xiao, G; Gilliland, G L

    1998-05-01

    The biosynthetic threonine deaminase from Escherichia coli, an allosteric tetramer with key regulatory functions, has been crystallized in several crystal forms. Two distinct forms, both belonging to either space group P3121 or P3221, with different sized asymmetric units that both contain a tetramer, grow under identical conditions. Diffraction data sets to 2.8 A resolution (native) and 2. 9 A resolution (isomorphous uranyl derivative) have been collected from a third crystal form in space group I222.

  2. The influence of AccD5 on AccD6 carboxyltransferase essentiality in pathogenic and non-pathogenic Mycobacterium

    PubMed Central

    Pawelczyk, Jakub; Viljoen, Albertus; Kremer, Laurent; Dziadek, Jaroslaw

    2017-01-01

    Malonyl-coenzyme A (CoA) is a crucial extender unit for the synthesis of mycolic and other fatty acids in mycobacteria, generated in a reaction catalyzed by acetyl-CoA carboxylase. We previously reported on the essentiality of accD6Mtb encoding the functional acetyl-CoA carboxylase subunit in Mycobacterium tuberculosis. Strikingly, the homologous gene in the fast-growing, non-pathogenic Mycobacterium smegmatis - (accD6Msm) appeared to be dispensable, and its deletion did not influence the cell lipid content. Herein, we demonstrate that, despite the difference in essentiality, accD6Msm and accD6Mtb encode proteins of convergent catalytic activity in vivo. To identify an alternative, AccD6-independent, malonyl-CoA synthesis pathway in M. smegmatis, a complex genetic approach combined with lipid analysis was applied to screen all five remaining carboxyltransferase genes (accD1-accD5) with respect to their involvement in mycolic acid biosynthesis and ability to utilize acetyl-CoA as the substrate for carboxylation. This approach revealed that AccD1Msm, AccD2Msm and AccD3Msm are not essential for mycolic acid biosynthesis. Furthermore, we confirmed in vivo the function of AccD4Msm as an essential, long-chain acyl-CoA carboxyltransferase, unable to carboxylate short-chain substrate. Finally, our comparative studies unambiguously demonstrated between-species difference in in vivo ability of AccD5 carboxyltransferase to utilize acetyl-CoA that influences AccD6 essentiality in pathogenic and non-pathogenic mycobacteria. PMID:28205597

  3. Kinetics of shoot inversion-induced ethylene production in Pharbitis nil

    NASA Technical Reports Server (NTRS)

    Prasad, T. K.; Cline, M. G.

    1986-01-01

    Shoot inversion promotes a significant increase in ethylene production in the inverted part of the Pharbitis nil main shoot. The latent period for shoot inversion-induced ethylene production is ca. 2.75 h. Our results indicate that the shoot-inversion ethylene response is not persistent and can be terminated and rapidly reinitiated by appropriate alteration of the orientation of the main shoot regardless of prolonged previous exposures of the shoot to various orientations. The time course of the production of ACC (1-aminocyclopropane-1-carboxylic acid), the immediate precursor of ethylene, follows a pattern similar to that of ethylene during the various alterations of shoot orientation. Excised stem segments and intact stems are capable of induction, inhibition, and reinduction of ethylene evolution. Ethylene production reported here for shoot inversion does not result from segmenting (wounding) of the tissue.

  4. Burst of ethylene upon horizontal placement of tomato seedlings

    NASA Technical Reports Server (NTRS)

    Harrison, M.; Pickard, B. G.

    1984-01-01

    Seedlings of Lycopersicon esculentum Mill. cv Rutgers emit a pulse of ethylene during the first 2 to 4 minutes following horizontal placement. Because this burst appears too rapid and brief to be mediated by increase in net activity of 1-aminocyclopropane-1-carboxylic acid synthase, it might result form accelerated transformation of vacuolar 1-aminocyclopropane-1-carboxylic acid to ethylene.

  5. In vitro synthesis and stabilization of amorphous calcium carbonate (ACC) nanoparticles within liposomes

    SciTech Connect

    Tester, Chantel C.; Brock, Ryan E.; Wu, Ching-Hsuan; Krejci, Minna R.; Weigand, Steven; Joester, Derk

    2012-02-07

    We show that amorphous calcium carbonate (ACC) can be synthesized in phospholipid bilayer vesicles (liposomes). Liposome-encapsulated ACC nanoparticles are stable against aggregation, do not crystallize for at least 20 h, and are ideally suited to investigate the influence of lipid chemistry, particle size, and soluble additives on ACC in situ.

  6. 24 CFR 905.304 - CF ACC term and covenant to operate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 24 Housing and Urban Development 4 2014-04-01 2014-04-01 false CF ACC term and covenant to operate... URBAN DEVELOPMENT THE PUBLIC HOUSING CAPITAL FUND PROGRAM General Program Requirements § 905.304 CF ACC... operate all public housing projects in accordance with the CF ACC, as amended, and applicable...

  7. Adenosine deaminase deficiency with normal immune function. An acidic enzyme mutation.

    PubMed Central

    Daddona, P E; Mitchell, B S; Meuwissen, H J; Davidson, B L; Wilson, J M; Koller, C A

    1983-01-01

    In most instances, marked deficiency of the purine catabolic enzyme adenosine deaminase results in lymphopenia and severe combined immunodeficiency disease. Over a 2-yr period, we studied a white male child with markedly deficient erythrocyte and lymphocyte adenosine deaminase activity and normal immune function. We have documented that (a) adenosine deaminase activity and immunoreactive protein are undetectable in erythrocytes, 0.9% of normal in lymphocytes, 4% in cultured lymphoblasts, and 14% in skin fibroblasts; (b) plasma adenosine and deoxyadenosine levels are undetectable and deoxy ATP levels are only slightly elevated in lymphocytes and in erythrocytes; (c) no defect in deoxyadenosine metabolism is present in the proband's cultured lymphoblasts; (d) lymphoblast adenosine deaminase has normal enzyme kinetics, absolute specific activity, S20,w, pH optimum, and heat stability; and (e) the proband's adenosine deaminase exhibits a normal apparent subunit molecular weight but an abnormal isoelectric pH. In contrast to the three other adenosine deaminase-deficient healthy subjects who have been described, the proband is unique in demonstrating an acidic, heat-stable protein mutation of the enzyme that is associated with less than 1% lymphocyte adenosine deaminase activity. Residual adenosine deaminase activity in tissues other than lymphocytes may suffice to metabolize the otherwise lymphotoxic enzyme substrate(s) and account for the preservation of normal immune function. Images FIGURE 1 FIGURE 2 FIGURE 3 PMID:6603477

  8. ACTIVATION OF A CRYPTIC D-SERINE DEAMINASE (DSD) GENE FROM PSEUDOMONAS CEPACIA 17616

    EPA Science Inventory

    D-serine inhibits growth of P. cepacia 17616; however, resistant mutants able to express an ordinarily cryptic D-serine deaminase (dsd) gene were isolated readily. The resistant strains formed high levels of a D-serine deaminase active on D-threonine as well as D-serine. IS eleme...

  9. Reward salience and risk aversion underlie differential ACC activity in substance dependence

    PubMed Central

    Alexander, William H.; Fukunaga, Rena; Finn, Peter; Brown, Joshua W.

    2015-01-01

    The medial prefrontal cortex, especially the dorsal anterior cingulate cortex (ACC), has long been implicated in cognitive control and error processing. Although the association between ACC and behavior has been established, it is less clear how ACC contributes to dysfunctional behavior such as substance dependence. Evidence from neuroimaging studies investigating ACC function in substance users is mixed, with some studies showing disengagement of ACC in substance dependent individuals (SDs), while others show increased ACC activity related to substance use. In this study, we investigate ACC function in SDs and healthy individuals performing a change signal task for monetary rewards. Using a priori predictions derived from a recent computational model of ACC, we find that ACC activity differs between SDs and controls in factors related to reward salience and risk aversion between SDs and healthy individuals. Quantitative fits of a computational model to fMRI data reveal significant differences in best fit parameters for reward salience and risk preferences. Specifically, the ACC in SDs shows greater risk aversion, defined as concavity in the utility function, and greater attention to rewards relative to reward omission. Furthermore, across participants risk aversion and reward salience are positively correlated. The results clarify the role that ACC plays in both the reduced sensitivity to omitted rewards and greater reward valuation in SDs. Clinical implications of applying computational modeling in psychiatry are also discussed. PMID:26106528

  10. Leaf Abscission Induced by Ethylene in Water-Stressed Intact Seedlings of Cleopatra Mandarin Requires Previous Abscisic Acid Accumulation in Roots.

    PubMed

    Gomez-Cadenas, A.; Tadeo, F. R.; Talon, M.; Primo-Millo, E.

    1996-09-01

    The involvement of abscisic acid (ABA) in the process of leaf abscission induced by 1-aminocyclopropane-1-carboxylic acid (ACC) transported from roots to shoots in Cleopatra mandarin (Citrus reshni Hort. ex Tan.) seedlings grown under water stress was studied using norflurazon (NF). Water stress induced both ABA (24-fold) and ACC (16-fold) accumulation in roots and arrested xylem flow. Leaf bulk ABA also increased (8-fold), although leaf abscission did not occur. Shortly after rehydration, root ABA and ACC returned to their prestress levels, whereas sharp and transitory increases of ACC (17-fold) and ethylene (10-fold) in leaves and high percentages of abscission (up to 47%) were observed. NF suppressed the ABA and ACC accumulation induced by water stress in roots and the sharp increases of ACC and ethylene observed after rewatering in leaves. NF also reduced leaf abscission (7-10%). These results indicate that water stress induces root ABA accumulation and that this is required for the process of leaf abscission to occur. It was also shown that exogenous ABA increases ACC levels in roots but not in leaves. Collectively, the data suggest that ABA, the primary sensitive signal to water stress, modulates the levels of ethylene, which is the hormonal activator of leaf abscission. This assumption implies that root ACC levels are correlated with root ABA amounts in a dependent way, which eventually links water status to an adequate, protective response such as leaf abscission.

  11. Recombination and horizontal transfer of nodulation and ACC deaminase (acdS) genes within Alpha- and Betaproteobacteria nodulating legumes of the Cape Fynbos biome.

    PubMed

    Lemaire, Benny; Van Cauwenberghe, Jannick; Chimphango, Samson; Stirton, Charles; Honnay, Olivier; Smets, Erik; Muasya, A Muthama

    2015-11-01

    The goal of this work is to study the evolution and the degree of horizontal gene transfer (HGT) within rhizobial genera of both Alphaproteobacteria (Mesorhizobium, Rhizobium) and Betaproteobacteria (Burkholderia), originating from South African Fynbos legumes. By using a phylogenetic approach and comparing multiple chromosomal and symbiosis genes, we revealed conclusive evidence of high degrees of horizontal transfer of nodulation genes among closely related species of both groups of rhizobia, but also among species with distant genetic backgrounds (Rhizobium and Mesorhizobium), underscoring the importance of lateral transfer of symbiosis traits as an important evolutionary force among rhizobia of the Cape Fynbos biome. The extensive exchange of symbiosis genes in the Fynbos is in contrast with a lack of significant events of HGT among Burkholderia symbionts from the South American Cerrado and Caatinga biome. Furthermore, homologous recombination among selected housekeeping genes had a substantial impact on sequence evolution within Burkholderia and Mesorhizobium. Finally, phylogenetic analyses of the non-symbiosis acdS gene in Mesorhizobium, a gene often located on symbiosis islands, revealed distinct relationships compared to the chromosomal and symbiosis genes, suggesting a different evolutionary history and independent events of gene transfer. The observed events of HGT and incongruence between different genes necessitate caution in interpreting topologies from individual data types.

  12. Laser photobleaching leads to a fluorescence grade adenosine deaminase.

    PubMed

    Parola, A H; Caiolfa, V R; Bar, I; Rosenwaks, S

    1989-09-01

    The enzyme adenosine deaminase (adenosine aminohydrolase EC 3.5.4.4) from calf intestinal mucosa is commercially available at high purity grade yet, at the sensitivity at which fluorescence studies may be undertaken, a nonpeptidic fluorescence is detectable at lambda exmax = 350 nm and lambda emmax = 420 nm. A sevenfold decrease of this nonpeptidic fluorescence was obtained upon irradiation by the third harmonic (355 nm) of a Nd:YAG laser for 16 min, at 5 mJ/pulse, with a pulse width of 6 ns at a repetition rate of 10 Hz. The decline of fluorescence was accompanied by a negligible loss of enzymatic activity. Moreover, the integrity of the protein was ascertained by (i) its fluorescence (lambda exmax = 305 nm, lambda emmax = 335 nm) and lifetime distribution and (ii) its kinetics in the presence of the substrate adenosine and two inhibitors, all of which remained essentially unaltered. Laser photobleaching is a simple way to achieve a fluorescence grade adenosine deaminase.

  13. Melamine Deaminase and Atrazine Chlorohydrolase: 98 Percent Identical but Functionally Different

    PubMed Central

    Seffernick, Jennifer L.; de Souza, Mervyn L.; Sadowsky, Michael J.; Wackett, Lawrence P.

    2001-01-01

    The gene encoding melamine deaminase (TriA) from Pseudomonas sp. strain NRRL B-12227 was identified, cloned into Escherichia coli, sequenced, and expressed for in vitro study of enzyme activity. Melamine deaminase displaced two of the three amino groups from melamine, producing ammeline and ammelide as sequential products. The first deamination reaction occurred more than 10 times faster than the second. Ammelide did not inhibit the first or second deamination reaction, suggesting that the lower rate of ammeline hydrolysis was due to differential substrate turnover rather than product inhibition. Remarkably, melamine deaminase is 98% identical to the enzyme atrazine chlorohydrolase (AtzA) from Pseudomonas sp. strain ADP. Each enzyme consists of 475 amino acids and differs by only 9 amino acids. AtzA was shown to exclusively catalyze dehalogenation of halo-substituted triazine ring compounds and had no activity with melamine and ammeline. Similarly, melamine deaminase had no detectable activity with the halo-triazine substrates. Melamine deaminase was active in deamination of a substrate that was structurally identical to atrazine, except for the substitution of an amino group for the chlorine atom. Moreover, melamine deaminase and AtzA are found in bacteria that grow on melamine and atrazine compounds, respectively. These data strongly suggest that the 9 amino acid differences between melamine deaminase and AtzA represent a short evolutionary pathway connecting enzymes catalyzing physiologically relevant deamination and dehalogenation reactions, respectively. PMID:11274097

  14. Inhibition of Acetyl-CoA Carboxylase 1 (ACC1) and 2 (ACC2) Reduces Proliferation and De Novo Lipogenesis of EGFRvIII Human Glioblastoma Cells

    PubMed Central

    Jones, Jessica E. C.; Esler, William P.; Patel, Rushi; Lanba, Adhiraj; Vera, Nicholas B.; Pfefferkorn, Jeffrey A.; Vernochet, Cecile

    2017-01-01

    Tumor cell proliferation and migration processes are regulated by multiple metabolic pathways including glycolysis and de novo lipogenesis. Since acetyl-CoA carboxylase (ACC) is at the junction of lipids synthesis and oxidative metabolic pathways, we investigated whether use of a dual ACC inhibitor would provide a potential therapy against certain lipogenic cancers. The impact of dual ACC1/ACC2 inhibition was investigated using a dual ACC1/ACC2 inhibitor as well as dual siRNA knock down on the cellular viability and metabolism of two glioblastoma multiform cancer cell lines, U87 and a more aggressive form, U87 EGFRvIII. We first demonstrated that while ACCi inhibited DNL in both cell lines, ACCi preferentially blunted the U87 EGFRvIII cellular proliferation capacity. Metabolically, chronic treatment with ACCi significantly upregulated U87 EGFRvIII cellular respiration and extracellular acidification rate, a marker of glycolytic activity, but impaired mitochondrial health by reducing maximal respiration and decreasing mitochondrial ATP production efficiency. Moreover, ACCi treatment altered the cellular lipids content and increased apoptotic caspase activity in U87 EGFRvIII cells. Collectively these data indicate that ACC inhibition, by reducing DNL and increasing cellular metabolic rate, may have therapeutic utility for the suppression of lipogenic tumor growth and warrants further investigation. PMID:28081256

  15. Adenine arabinoside inhibition of adenovirus replication enhanced by an adenosine deaminase inhibitor.

    PubMed

    Wigand, R

    1979-01-01

    The inhibition of adenovirus multiplication by adenine arabinoside was determined by yield reduction in one-step multiplication cycle. Inhibition was greatly enhanced by an adenosine deaminase inhibitor (2-deoxycoformycin) in concentrations down to 10 ng/ml. Adenovirus types from four subgroups showed similar results. However, the enhancing effect of adenosine deaminase inhibitor was great in HeLa cells, moderate in human fibroblasts, and negligible in Vero cells. This difference could be explained by different concentrations of adenosine deaminase found in cell homogenates.

  16. Further studies of auxin and ACC induced feminization in the cucumber plant using ethylene inhibitors

    NASA Technical Reports Server (NTRS)

    Takahashi, H.; Jaffe, M. J.

    1984-01-01

    The present study was designed to establish the role of an essential hormone controlling sex expression in cucumber. A potent anti-ethylene agent, AgNO3, completely inhibited pistillate flower formation caused by IAA, ACC or ethephon. Inhibitors of ethylene biosynthesis, AVG and CoCl2 also suppressed feminization due to exogenous IAA or ACC. Though AVG also suppressed ethephon-induced feminization, this may be due to the second effect of AVG rather than the effect on ACC biosynthesis. These results confirm that ethylene is a major factor regulating feminization and that exogenous auxin induces pistillate flower formation through its stimulation of ethylene production, rather than ACC production.

  17. AMP-deaminase from thymus of patients with myasthenia gravis.

    PubMed

    Rybakowska, I; Szydłowska, M; Szrok, S; Bakuła, S; Kaletha, K

    2015-01-01

    Myasthenia gravis (MG) is characterized clinically by skeletal muscle fatigue following the excessive exercise. Interestingly most of MG patients manifest parallely also some abnormalities of the thymus.AMP-deaminase (AMPD) from human thymus was not a subject of studies up to now. In this paper, mRNA expression and some physico-chemical and immunological properties of AMPD purified from the thymus of MG patients were described. Experiments performed identified the liver isozyme (AMPD2) as the main isoform of AMPD expressed in this organ. The activity of AMPD found in this organ was higher than in other human non-(skeletal) muscle tissues indicating on role the enzyme may play in supplying of guanylates required for the intensive multiplication of thymocytes.

  18. Late-onset adenosine deaminase deficiency presenting with Heck's disease.

    PubMed

    Artac, Hasibe; Göktürk, Bahar; Bozdemir, Sefika Elmas; Toy, Hatice; van der Burg, Mirjam; Santisteban, Ines; Hershfield, Michael; Reisli, Ismail

    2010-08-01

    Focal epithelial hyperplasia, also known as Heck's disease, is a rare but distinctive entity of viral etiology with characteristic clinical and histopathological features. It is a benign, asymptomatic disease of the oral mucosa caused by human papilloma viruses (HPV). Previous studies postulated an association between these lesions and immunodeficiency. Genetic deficiency of adenosine deaminase (ADA) results in varying degrees of immunodeficiency, including neonatal onset severe combined immunodeficiency (ADA-SCID), and milder, later onset immunodeficiency. We report a 12-year-old girl with the late onset-ADA deficiency presenting with Heck's disease. Our case report should draw attention to the possibility of immunodeficiency in patients with HPV-induced focal epithelial hyperplasia.

  19. Photodynamic therapy-driven induction of suicide cytosine deaminase gene.

    PubMed

    Bil, Jacek; Wlodarski, Pawel; Winiarska, Magdalena; Kurzaj, Zuzanna; Issat, Tadeusz; Jozkowicz, Alicja; Wegiel, Barbara; Dulak, Jozef; Golab, Jakub

    2010-04-28

    Photodynamic therapy (PDT) of tumors is associated with induction of hypoxia that results in activation of hypoxia-inducible factors (HIFs). Several observations indicate that increased HIFs transcriptional activity in tumor cells is associated with cytoprotective responses that limit cytotoxic effectiveness of PDT. Therefore, we decided to examine whether this cytoprotective mechanism could be intentionally used for designing more efficient tumor cell cytotoxicity. To this end we transfected tumor cells with a plasmid vector carrying a suicide cytosine deaminase gene driven by a promoter containing hypoxia response elements (HRE). The presence of such a genetic molecular beacon rendered tumor cells sensitive to cytotoxic effects of a non-toxic prodrug 5-fluorocytosine (5-FC). The results of this study provides a proof of concept that inducible cytoprotective mechanisms can be exploited to render tumor cells more susceptible to cytotoxic effects of prodrugs activated by products of suicide genes.

  20. Oxygen control of ethylene biosynthesis during seed development in Arabidopsis thaliana (L.) Heynh

    NASA Technical Reports Server (NTRS)

    Ramonell, K. M.; McClure, G.; Musgrave, M. E.

    2002-01-01

    An unforeseen side-effect on plant growth in reduced oxygen is the loss of seed production at concentrations around 25% atmospheric (50 mmol mol-1 O2). In this study, the model plant Arabidopsis thaliana (L.) Heynh. cv. 'Columbia' was used to investigate the effect of low oxygen on ethylene biosynthesis during seed development. Plants were grown in a range of oxygen concentrations (210 [equal to ambient], 160, 100, 50 and 25 mmol mol-1) with 0.35 mmol mol-1 CO2 in N2. Ethylene in full-sized siliques was sampled using gas chromatography, and viable seed production was determined at maturity. Molecular analysis of ethylene biosynthesis was accomplished using cDNAs encoding 1-aminocyclopropane-1-carboxylic acid (ACC) synthase and ACC oxidase in ribonuclease protection assays and in situ hybridizations. No ethylene was detected in siliques from plants grown at 50 and 25 mmol mol-1 O2. At the same time, silique ACC oxidase mRNA increased three-fold comparing plants grown under the lowest oxygen with ambient controls, whereas ACC synthase mRNA was unaffected. As O2 decreased, tissue-specific patterning of ACC oxidase and ACC synthase gene expression shifted from the embryo to the silique wall. These data demonstrate how low O2 modulates the activity and expression of the ethylene biosynthetic pathway during seed development in Arabidopsis.

  1. ETHY. A Theory of Fruit Climacteric Ethylene Emission1

    PubMed Central

    Génard, Michel; Gouble, Barbara

    2005-01-01

    A theory of fruit climacteric ethylene emission was developed and used as the basis of a simulation model called ETHY. According to the theory, the biosynthetic pathway of ethylene is supplied by ATP and is regulated by 1-aminocyclopropane-1-carboxylic acid (ACC) synthase and ACC oxidase. The conjugation of ACC with malonate to form MACC was taken into account as a way to decrease the availability of ACC. Because of the seasonal increase of fruit volume, the dilution of biochemical compounds used in ETHY was taken into account. Finally, the ethylene diffusion across the skin was considered. The theory took into account the effect of temperature and O2 and CO2 internal concentrations on ethylene. The model was applied to peach (Prunus persica) fruit over 3 years, several leaf:fruit ratios, and irrigation conditions. An adequate ethylene increase was predicted without considering any increase in respiration during the ripening period, which suggests that the respiratory climacteric may not be required for ripening. Another important result of this study is the high sensitivity of ETHY to the parameters involved in the calculation of ACC oxidase and ACC synthase activities, ATP production, and skin surface and permeability. ETHY was also highly sensitive to changes in fruit growth and temperature. PMID:16143642

  2. Inhibitory Action of Auxin on Root Elongation Not Mediated by Ethylene

    PubMed Central

    Eliasson, Lennart; Bertell, Gertrud; Bolander, Eva

    1989-01-01

    The inhibitory effects of indole-3-acetic acid (IAA) and 1-aminocyclopropane-1-carboxylic acid (ACC) on elongation growth of pea (Pisum sativum L.) seedling roots were investigated in relation to the effects of these compounds on ethylene production by the root tips. When added to the growth solution both compounds caused a progressively increasing inhibition of growth within the concentration range of 0.01 to 1 micromolar. However, only ACC increased ethylene production in root tips excised from the treated seedlings after 24 hours. High auxin concentrations caused a transitory increase of ethylene production during a few hours in the beginning of the treatment period, but even in 1 micromolar IAA this increase was too low to have any appreciable effect on growth. ACC, but not IAA, caused growth curvatures, typical of ethylene treatment, in the root tips. IAA caused conspicuous swelling of the root tips while ACC did not. Cobalt and silver ions reversed the growth inhibitory effects induced by ACC but did not counteract the inhibition of elongation or swelling caused by IAA. The growth effects caused by the ACC treatments were obviously due to ethylene production. We found no evidence to indicate that the growth inhibition or swelling caused by IAA is mediated by ethylene. It is concluded that the inhibitory action of IAA on root growth is caused by this auxin per se. PMID:16667017

  3. Ethylene biosynthesis and cadmium toxicity in leaf tissue of beans (Phaseolus vulgaris L. )

    SciTech Connect

    Fuhrer, J.

    1982-07-01

    Stress ethylene production in bean (Phaseolus vulgaris L., ev. Taylor's Horticultural) leaf tissue was stimulated by Cd/sup 2 +/ at concentrations above 1 micromolar. Cd/sup 2 +/-induced ethylene biosynthesis was dependent upon synthesis of 1-aminocyclopropane-1-carboxylic acid (ACC) by ACC synthase. Activity of ACC synthase and ethylene production rate peaked at 8 h of treatment. The subsequent decline in enzyme activity was most likely due to inactivation of the enzyme by Cd/sup 2 +/, which inhibited ACC synthase activity in vitro at concentrations as low as 0.1 micromolar. Decrease in ethylene production rate was accompanied by leakage of solutes and increasing inhibition of ACC-dependent ethylene production. Ca/sup 2 +/, present during a 2-hour preincubation, reduced the effect of Cd/sup 2 +/ on leakage and ACC conversion. This suggests that Cd/sup 2 +/ exerts its toxicity through membrane damage and inactivation of enzymes. The possibility of an indirect stimulation of ethylene biosynthesis through a wound signal from injured cells is discussed.

  4. Effect of silver ions on ethylene biosynthesis by tomato fruit tissue.

    PubMed

    Atta-Aly, M A; Saltveit, M E; Hobson, G E

    1987-01-01

    Mature-green tomato fruit (Lycopersicon esculentum Mill.) were treated asymmetrically with 2 millimolar silver thiosulfate (STS) through a cut portion of the peduncle while still attached to the plant. One-half of the fruit received silver and remained green while the other half ripened normally and was silver-free (less than 0.01 parts per billion). Harvested mature-green fruit were also treated with STS through the cut pedicel. Green tissue from silver-treated fruit had levels of 1-aminocyclopropane-1-carboxylic acid (ACC, the immediate ethylene precursor) slightly less or similar to that of turning or red-ripe tissue from the same fruit, and similar to that of mature-green tissue from control fruit. Ethylene production was higher in green tissue from silver-treated fruit than from either red tissue from the same fruit, or mature-green tissue from control fruit. By inhibiting ACC synthesis with aminoethoxyvinyl glycine, and by applying ACC +/- silver to excised disks of pericarp tissue from control or silver-treated tomatoes, we showed that short-term silver treatment did not affect the biological conversion of ACC to ethylene, while long-term treatment stimulated both the conversion of ACC to ethylene and the synthesis of ACC.

  5. Opposing activity changes in AMP deaminase and AMP-activated protein kinase in the hibernating ground squirrel.

    PubMed

    Lanaspa, Miguel A; Epperson, L Elaine; Li, Nanxing; Cicerchi, Christina; Garcia, Gabriela E; Roncal-Jimenez, Carlos A; Trostel, Jessica; Jain, Swati; Mant, Colin T; Rivard, Christopher J; Ishimoto, Takuji; Shimada, Michiko; Sanchez-Lozada, Laura Gabriela; Nakagawa, Takahiko; Jani, Alkesh; Stenvinkel, Peter; Martin, Sandra L; Johnson, Richard J

    2015-01-01

    Hibernating animals develop fatty liver when active in summertime and undergo a switch to a fat oxidation state in the winter. We hypothesized that this switch might be determined by AMP and the dominance of opposing effects: metabolism through AMP deaminase (AMPD2) (summer) and activation of AMP-activated protein kinase (AMPK) (winter). Liver samples were obtained from 13-lined ground squirrels at different times during the year, including summer and multiples stages of winter hibernation, and fat synthesis and β-fatty acid oxidation were evaluated. Changes in fat metabolism were correlated with changes in AMPD2 activity and intrahepatic uric acid (downstream product of AMPD2), as well as changes in AMPK and intrahepatic β-hydroxybutyrate (a marker of fat oxidation). Hepatic fat accumulation occurred during the summer with relatively increased enzymes associated with fat synthesis (FAS, ACL and ACC) and decreased enoyl CoA hydratase (ECH1) and carnitine palmitoyltransferase 1A (CPT1A), rate limiting enzymes of fat oxidation. In summer, AMPD2 activity and intrahepatic uric acid levels were high and hepatic AMPK activity was low. In contrast, the active phosphorylated form of AMPK and β-hydroxybutyrate both increased during winter hibernation. Therefore, changes in AMPD2 and AMPK activity were paralleled with changes in fat synthesis and fat oxidation rates during the summer-winter cycle. These data illuminate the opposing forces of metabolism of AMP by AMPD2 and its availability to activate AMPK as a switch that governs fat metabolism in the liver of hibernating ground squirrel.

  6. Freeze-drying yields stable and pure amorphous calcium carbonate (ACC).

    PubMed

    Ihli, Johannes; Kulak, Alexander N; Meldrum, Fiona C

    2013-04-18

    A simple synthetic method is presented for the precipitation of high purity, dry amorphous calcium carbonate (ACC) based on freeze-drying saturated, counter ion free CaCO3 solutions, where the ACC produced shows an extended atmospheric stability. Translation of the methodology to amorphous calcium phosphate demonstrates the generality of the approach.

  7. ACCE/ACS National Educator and Leader of the Year Winners: AEC Congratulates These Outstanding Educators

    ERIC Educational Resources Information Center

    Australian Educational Computing, 2012

    2012-01-01

    This article presents the ACCE/ACS National Educator and Leader of the Year winners. Anne Mirtschin is the recipient of the ACCE/ACS 2012 Educator of the Year Award. Mirtschin is an innovative teacher at Hawkesdale P-12 College a small rural school that is isolated culturally and geographically. She uses online tools and technology to create…

  8. 24 CFR 882.805 - HA application process, ACC execution, and pre-rehabilitation activities.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    .... (4) The owner is responsible for selecting a competent contractor to undertake the rehabilitation...; (ii) Assure that the owner has selected a contractor in accordance with paragraph (c)(4) of this... ACC for an additional 10 years. (3) Section 882.403(a) (Maximum Total ACC Commitments) applies to...

  9. Effect of Lithium on Thigmomorphogenesis in Bryonia dioica Ethylene Production and Sensitivity 1

    PubMed Central

    Boyer, Nicole; Desbiez, Marie-Odile; Hofinger, Michel; Gaspar, Thomas

    1983-01-01

    Rubbing internodes of Bryonia dioica plants reduced their ethylene production but increased their capacity to convert 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene. These results were explained by the previously shown rubbing-induced decrease of indoleacetic acid, which controls the level of ACC synthase, and by the increase of membrane-associated peroxidases which would participate in the conversion of ACC-ethylene. Pretreatment of the plants with Li had no significant effect on control plants but counteracted the rubbing-induced decrease of ethylene production and diminished the capacity of the internodes to convert ACC to ethylene. Exogenously applied ethylene induced an increase of peroxidase activity similar to that caused by rubbing. Inasmuch as both effects were reduced by Li, it was concluded that Li inhibition of thigmomorphogenetic processes was essentially due to a Li inhibition of the effect of ethylene formed in response to mechanical stimuli. The decreased ethylene production and ACC conversion capacity in the presence of Li were explained by a cellular redistribution of peroxidases. PMID:16663035

  10. Defense Responses in Aspen with Altered Pectin Methylesterase Activity Reveal the Hormonal Inducers of Tyloses1[OPEN

    PubMed Central

    Leśniewska, Joanna; Krzesłowska, Magdalena; Kushwah, Sunita; Sundberg, Björn; Moritz, Thomas

    2017-01-01

    Tyloses are ingrowths of parenchyma cells into the lumen of embolized xylem vessels, thereby protecting the remaining xylem from pathogens. They are found in heartwood, sapwood, and in abscission zones and can be induced by various stresses, but their molecular triggers are unknown. Here, we report that down-regulation of PECTIN METHYLESTERASE1 (PtxtPME1) in aspen (Populus tremula × tremuloides) triggers the formation of tyloses and activation of oxidative stress. We tested whether any of the oxidative stress-related hormones could induce tyloses in intact plantlets grown in sterile culture. Jasmonates, including jasmonic acid (JA) and methyl jasmonate, induced the formation of tyloses, whereas treatments with salicylic acid (SA) and 1-aminocyclopropane-1-carboxylic acid (ACC) were ineffective. SA abolished the induction of tyloses by JA, whereas ACC was synergistic with JA. The ability of ACC to stimulate tyloses formation when combined with JA depended on ethylene (ET) signaling, as shown by a decrease in the response in ET-insensitive plants. Measurements of internal ACC and JA concentrations in wild-type and ET-insensitive plants treated simultaneously with these two compounds indicated that ACC and JA regulate each other’s concentration in an ET-dependent manner. The findings indicate that jasmonates acting synergistically with ethylene are the key molecular triggers of tyloses. PMID:27923986

  11. The Arabidopsis mutant alh1 illustrates a cross talk between ethylene and auxin.

    PubMed

    Vandenbussche, Filip; Smalle, Jan; Le, Jie; Saibo, Nelson José Madeira; De Paepe, Annelies; Chaerle, Laury; Tietz, Olaf; Smets, Raphael; Laarhoven, Lucas J J; Harren, Frans J M; Van Onckelen, Harry; Palme, Klaus; Verbelen, Jean-Pierre; Van Der Straeten, Dominique

    2003-03-01

    Ethylene or its precursor 1-aminocyclopropane-1-carboxylic acid (ACC) can stimulate hypocotyl elongation in light-grown Arabidopsis seedlings. A mutant, designated ACC-related long hypocotyl 1 (alh1), that displayed a long hypocotyl in the light in the absence of the hormone was characterized. Etiolated alh1 seedlings overproduced ethylene and had an exaggerated apical hook and a thicker hypocotyl, although no difference in hypocotyl length was observed when compared with wild type. Alh1 plants were less sensitive to ethylene, as reflected by reduction of ACC-mediated inhibition of hypocotyl growth in the dark and delay in flowering and leaf senescence. Alh1 also had an altered response to auxin, whereas auxin levels in whole alh1 seedlings remained unaffected. In contrast to wild type, alh1 seedlings showed a limited hypocotyl elongation when treated with indole-3-acetic acid. Alh1 roots had a faster response to gravity. Furthermore, the hypocotyl elongation of alh1 and of ACC-treated wild type was reverted by auxin transport inhibitors. In addition, auxin up-regulated genes were ectopically expressed in hypocotyls upon ACC treatment, suggesting that the ethylene response is mediated by auxins. Together, these data indicate that alh1 is altered in the cross talk between ethylene and auxins, probably at the level of auxin transport.

  12. Discovery and Structure Determination of the Orphan Enzyme Isoxanthopterin Deaminase

    SciTech Connect

    Hall, R.S.; Swaminathan, S.; Agarwal, R.; Hitchcock, D.; Sauder, J. M.; Burley, S. K.; Raushel, F. M.

    2010-05-25

    Two previously uncharacterized proteins have been identified that efficiently catalyze the deamination of isoxanthopterin and pterin 6-carboxylate. The genes encoding these two enzymes, NYSGXRC-9339a (gi|44585104) and NYSGXRC-9236b (gi|44611670), were first identified from DNA isolated from the Sargasso Sea as part of the Global Ocean Sampling Project. The genes were synthesized, and the proteins were subsequently expressed and purified. The X-ray structure of Sgx9339a was determined at 2.7 {angstrom} resolution (Protein Data Bank entry 2PAJ). This protein folds as a distorted ({beta}/{alpha}){sub 8} barrel and contains a single zinc ion in the active site. These enzymes are members of the amidohydrolase superfamily and belong to cog0402 within the clusters of orthologous groups (COG). Enzymes in cog0402 have previously been shown to catalyze the deamination of guanine, cytosine, S-adenosylhomocysteine, and 8-oxoguanine. A small compound library of pteridines, purines, and pyrimidines was used to probe catalytic activity. The only substrates identified in this search were isoxanthopterin and pterin 6-carboxylate. The kinetic constants for the deamination of isoxanthopterin with Sgx9339a were determined to be 1.0 s{sup -1}, 8.0 {micro}M, and 1.3 x 10{sup 5} M{sup -1} s{sup -1} (k{sub cat}, K{sub m}, and k{sub cat}/K{sub m}, respectively). The active site of Sgx9339a most closely resembles the active site for 8-oxoguanine deaminase (Protein Data Bank entry 2UZ9). A model for substrate recognition of isoxanthopterin by Sgx9339a was proposed on the basis of the binding of guanine and xanthine in the active site of guanine deaminase. Residues critical for substrate binding appear to be conserved glutamine and tyrosine residues that form hydrogen bonds with the carbonyl oxygen at C4, a conserved threonine residue that forms hydrogen bonds with N5, and another conserved threonine residue that forms hydrogen bonds with the carbonyl group at C7. These conserved active site

  13. OpenARC: Extensible OpenACC Compiler Framework for Directive-Based Accelerator Programming Study

    SciTech Connect

    Lee, Seyong; Vetter, Jeffrey S

    2014-01-01

    Directive-based, accelerator programming models such as OpenACC have arisen as an alternative solution to program emerging Scalable Heterogeneous Computing (SHC) platforms. However, the increased complexity in the SHC systems incurs several challenges in terms of portability and productivity. This paper presents an open-sourced OpenACC compiler, called OpenARC, which serves as an extensible research framework to address those issues in the directive-based accelerator programming. This paper explains important design strategies and key compiler transformation techniques needed to implement the reference OpenACC compiler. Moreover, this paper demonstrates the efficacy of OpenARC as a research framework for directive-based programming study, by proposing and implementing OpenACC extensions in the OpenARC framework to 1) support hybrid programming of the unified memory and separate memory and 2) exploit architecture-specific features in an abstract manner. Porting thirteen standard OpenACC programs and three extended OpenACC programs to CUDA GPUs shows that OpenARC performs similarly to a commercial OpenACC compiler, while it serves as a high-level research framework.

  14. Three-dimensional structure and catalytic mechanism of cytosine deaminase.

    PubMed

    Hall, Richard S; Fedorov, Alexander A; Xu, Chengfu; Fedorov, Elena V; Almo, Steven C; Raushel, Frank M

    2011-06-07

    Cytosine deaminase (CDA) from E. coli is a member of the amidohydrolase superfamily. The structure of the zinc-activated enzyme was determined in the presence of phosphonocytosine, a mimic of the tetrahedral reaction intermediate. This compound inhibits the deamination of cytosine with a K(i) of 52 nM. The zinc- and iron-containing enzymes were characterized to determine the effect of the divalent cations on activation of the hydrolytic water. Fe-CDA loses activity at low pH with a kinetic pK(a) of 6.0, and Zn-CDA has a kinetic pK(a) of 7.3. Mutation of Gln-156 decreased the catalytic activity by more than 5 orders of magnitude, supporting its role in substrate binding. Mutation of Glu-217, Asp-313, and His-246 significantly decreased catalytic activity supporting the role of these three residues in activation of the hydrolytic water molecule and facilitation of proton transfer reactions. A library of potential substrates was used to probe the structural determinants responsible for catalytic activity. CDA was able to catalyze the deamination of isocytosine and the hydrolysis of 3-oxauracil. Large inverse solvent isotope effects were obtained on k(cat) and k(cat)/K(m), consistent with the formation of a low-barrier hydrogen bond during the conversion of cytosine to uracil. A chemical mechanism for substrate deamination by CDA was proposed.

  15. The ONIOM molecular dynamics method for biochemical applications: cytidine deaminase

    SciTech Connect

    Matsubara, Toshiaki; Dupuis, Michel; Aida, Misako

    2007-03-22

    Abstract We derived and implemented the ONIOM-molecular dynamics (MD) method for biochemical applications. The implementation allows the characterization of the functions of the real enzymes taking account of their thermal motion. In this method, the direct MD is performed by calculating the ONIOM energy and gradients of the system on the fly. We describe the first application of this ONOM-MD method to cytidine deaminase. The environmental effects on the substrate in the active site are examined. The ONIOM-MD simulations show that the product uridine is strongly perturbed by the thermal motion of the environment and dissociates easily from the active site. TM and MA were supported in part by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan. MD was supported by the Division of Chemical Sciences, Office of Basic Energy Sciences, and by the Office of Biological and Environmental Research of the U.S. Department of Energy DOE. Battelle operates Pacific Northwest National Laboratory for DOE.

  16. ADA (adenosine deaminase) gene therapy enters the competition

    SciTech Connect

    Culliton, B.J.

    1990-08-31

    Around the world, some 70 children are members of a select and deadly club. Born with an immune deficiency so severe that they will die of infection unless their immune systems can be repaired, they have captured the attention of would-be gene therapists who believe that a handful of these kids--the 15 or 20 who lack functioning levels of the enzyme adenosine deaminase (ADA)--could be saved by a healthy ADA gene. A team of gene therapists is ready to put the theory to the test. In April 1987, a team of NIH researchers headed by R. Michael Blaese and W. French Anderson came up with the first formal protocol to introduce a healthy ADA gene into an unhealthy human. After 3 years of line-by-line scrutiny by five review committees, they have permission to go ahead. Two or three children will be treated in the next year, and will be infused with T lymphocytes carrying the gene for ADA. If the experiment works, the ADA gene will begin producing normal amounts of ADA. An interesting feature of ADA deficiency, that makes it ideal for initial gene studies, is that the amount of ADA one needs for a healthy immune system is quite variable. Hence, once inside a patient's T cells, the new ADA gene needs only to express the enzyme in moderate amounts. No precise gene regulation is necessary.

  17. Adenosine deaminase in cell transformation. Biophysical manifestation of membrane dynamics.

    PubMed

    Porat, N; Gill, D; Parola, A H

    1988-10-15

    Cell transformation is associated with a dramatic collapse of a graphic fingerprint characteristic of normal cells, as measured by phase fluorimetry. This is demonstrated on adenosine deaminase (ADA, EC 3.5.4.4), an established malignancy marker. ADA activity is known to decrease markedly in chick embryo fibroblasts (CEF) transformed by Rous sarcoma virus. The high affinity between the catalytic small subunit ADA (SS-ADA) and its membranal complexing protein (ADCP) (which abounds on the plasma membrane of CEF) allowed the hybridization of fluorescent labeled SS-ADA with native ADCP on CEF. Multifrequency differential phase fluorimetry responded remarkably to the state of this hybrid membrane protein. The transformation process is shown to have led to increased membrane fluidity and rotational mobility of ADCP as well as to its reduced availability to SS-ADA binding. The hypothesis of protein vertical sinking into the lipid core of the membrane is now given support by our spectroscopic data. Additional models are considered. A regulatory role is thus suggested for the complexing protein, which may also account for (a) reduced ADA activity in transformed cells and (b) detachment, exclusive to normal cells, upon addition of SS-ADA in excess.

  18. Three-Dimensional Structure and Catalytic Mechanism of Cytosine Deaminase

    SciTech Connect

    R Hall; A Fedorov; C Xu; E Fedorov; S Almo; F Raushel

    2011-12-31

    Cytosine deaminase (CDA) from E. coli is a member of the amidohydrolase superfamily. The structure of the zinc-activated enzyme was determined in the presence of phosphonocytosine, a mimic of the tetrahedral reaction intermediate. This compound inhibits the deamination of cytosine with a K{sub i} of 52 nM. The zinc- and iron-containing enzymes were characterized to determine the effect of the divalent cations on activation of the hydrolytic water. Fe-CDA loses activity at low pH with a kinetic pKa of 6.0, and Zn-CDA has a kinetic pKa of 7.3. Mutation of Gln-156 decreased the catalytic activity by more than 5 orders of magnitude, supporting its role in substrate binding. Mutation of Glu-217, Asp-313, and His-246 significantly decreased catalytic activity supporting the role of these three residues in activation of the hydrolytic water molecule and facilitation of proton transfer reactions. A library of potential substrates was used to probe the structural determinants responsible for catalytic activity. CDA was able to catalyze the deamination of isocytosine and the hydrolysis of 3-oxauracil. Large inverse solvent isotope effects were obtained on k{sub cat} and k{sub cat}/K{sub m}, consistent with the formation of a low-barrier hydrogen bond during the conversion of cytosine to uracil. A chemical mechanism for substrate deamination by CDA was proposed.

  19. Functions and Regulation of RNA Editing by ADAR Deaminases

    PubMed Central

    Nishikura, Kazuko

    2010-01-01

    One type of RNA editing converts adenosines to inosines (A→I editing) in double-stranded RNA (dsRNA) substrates. A→I RNA editing is mediated by adenosine deaminase acting on RNA (ADAR) enzymes. A→I RNA editing of protein-coding sequences of a limited number of mammalian genes results in recoding and subsequent alterations of their functions. However, A→I RNA editing most frequently targets repetitive RNA sequences located within introns and 5′ and 3′ untranslated regions (UTRs). Although the biological significance of noncoding RNA editing remains largely unknown, several possibilities, including its role in the control of endogenous short interfering RNAs (esiRNAs), have been proposed. Furthermore, recent studies have revealed that the biogenesis and functions of certain microRNAs (miRNAs) are regulated by the editing of their precursors. Here, I review the recent findings that indicate new functions for A→I editing in the regulation of noncoding RNAs and for interactions between RNA editing and RNA interference mechanisms. PMID:20192758

  20. Adenosine Deaminase Deficiency – More Than Just an Immunodeficiency

    PubMed Central

    Whitmore, Kathryn V.; Gaspar, Hubert B.

    2016-01-01

    Adenosine deaminase (ADA) deficiency is best known as a form of severe combined immunodeficiency (SCID) that results from mutations in the gene encoding ADA. Affected patients present with clinical and immunological manifestations typical of a SCID. Therapies are currently available that can target these immunological disturbances and treated patients show varying degrees of clinical improvement. However, there is now a growing body of evidence that deficiency of ADA has significant impact on non-immunological organ systems. This review will outline the impact of ADA deficiency on various organ systems, starting with the well-understood immunological abnormalities. We will discuss possible pathogenic mechanisms and also highlight ways in which current treatments could be improved. In doing so, we aim to present ADA deficiency as more than an immunodeficiency and suggest that it should be recognized as a systemic metabolic disorder that affects multiple organ systems. Only by fully understanding ADA deficiency and its manifestations in all organ systems can we aim to deliver therapies that will correct all the clinical consequences. PMID:27579027

  1. Adenosine deaminase complexing protein (ADCP) immunoreactivity in colorectal adenocarcinoma.

    PubMed

    ten Kate, J; van den Ingh, H F; Khan, P M; Bosman, F T

    1986-04-15

    Immunoreactive adenosine deaminase complexing protein (ADCP) was studied in 91 human colorectal adenocarcinomas. The expression of ADCP was correlated with that of secretory component (SC) and carcinoembryonic antigen (CEA), with the histological grade and the Dukes' stage of the carcinomas. The histological grade was scored semi-quantitatively according to 5 structural and 4 cytological variables. ADCP expression was observed in 3 different staining patterns, namely: (1) diffuse cytoplasmic (77% of the carcinomas); (2) granular cytoplasmic (13%); and (3) membrane-associated (66%). These patterns were observed alone or in combination. Eleven percent of the carcinomas exhibited no ADCP immunoreactivity. Linear regression analysis showed that the expression of ADCP correlates with that of SC and CEA. However, no significant correlation emerged between the histological parameters or the Dukes' stage and any of the immunohistological parameters. Comparison of the histological characteristics of carcinomas exhibiting little or no ADCP immunoreactivity with those showing extensive immunoreactivity, showed that membranous ADCP immunoreactivity occurs more frequently in well-differentiated carcinomas. Structural parameters showed a better correlation with membranous ADCP expression than the cytological variables. It is concluded that membranous expression of ADCP and CEA are indicators of a high level of differentiation as reflected primarily in the structural characteristics of the tumor.

  2. Distribution of adenosine deaminase complexing protein (ADCP) in human tissues.

    PubMed

    Dinjens, W N; ten Kate, J; van der Linden, E P; Wijnen, J T; Khan, P M; Bosman, F T

    1989-12-01

    The normal distribution of adenosine deaminase complexing protein (ADCP) in the human body was investigated quantitatively by ADCP-specific radioimmunoassay (RIA) and qualitatively by immunohistochemistry. In these studies we used a specific rabbit anti-human ADCP antiserum. In all 19 investigated tissues, except erythrocytes, ADCP was found by RIA in the soluble and membrane fractions. From all tissues the membrane fractions contained more ADCP (expressed per mg protein) than the soluble fractions. High membrane ADCP concentrations were found in skin, renal cortex, gastrointestinal tract, and prostate. Immunoperoxidase staining confirmed the predominant membrane-associated localization of the protein. In serous sweat glands, convoluted tubules of renal cortex, bile canaliculi, gastrointestinal tract, lung, pancreas, prostate gland, salivary gland, gallbladder, mammary gland, and uterus, ADCP immunoreactivity was found confined to the luminal membranes of the epithelial cells. These data demonstrate that ADCP is present predominantly in exocrine glands and absorptive epithelia. The localization of ADCP at the secretory or absorptive apex of the cells suggests that the function of ADCP is related to the secretory and/or absorptive process.

  3. Recalibration of the ACC/AHA Risk Score in Two Population-Based German Cohorts

    PubMed Central

    de las Heras Gala, Tonia; Geisel, Marie Henrike; Peters, Annette; Thorand, Barbara; Baumert, Jens; Lehmann, Nils; Jöckel, Karl-Heinz; Moebus, Susanne; Erbel, Raimund; Meisinger, Christine

    2016-01-01

    Background The 2013 ACC/AHA guidelines introduced an algorithm for risk assessment of atherosclerotic cardiovascular disease (ASCVD) within 10 years. In Germany, risk assessment with the ESC SCORE is limited to cardiovascular mortality. Applicability of the novel ACC/AHA risk score to the German population has not yet been assessed. We therefore sought to recalibrate and evaluate the ACC/AHA risk score in two German cohorts and to compare it to the ESC SCORE. Methods We studied 5,238 participants from the KORA surveys S3 (1994–1995) and S4 (1999–2001) and 4,208 subjects from the Heinz Nixdorf Recall (HNR) Study (2000–2003). There were 383 (7.3%) and 271 (6.4%) first non-fatal or fatal ASCVD events within 10 years in KORA and in HNR, respectively. Risk scores were evaluated in terms of calibration and discrimination performance. Results The original ACC/AHA risk score overestimated 10-year ASCVD rates by 37% in KORA and 66% in HNR. After recalibration, miscalibration diminished to 8% underestimation in KORA and 12% overestimation in HNR. Discrimination performance of the ACC/AHA risk score was not affected by the recalibration (KORA: C = 0.78, HNR: C = 0.74). The ESC SCORE overestimated by 5% in KORA and by 85% in HNR. The corresponding C-statistic was 0.82 in KORA and 0.76 in HNR. Conclusions The recalibrated ACC/AHA risk score showed strongly improved calibration compared to the original ACC/AHA risk score. Predicting only cardiovascular mortality, discrimination performance of the commonly used ESC SCORE remained somewhat superior to the ACC/AHA risk score. Nevertheless, the recalibrated ACC/AHA risk score may provide a meaningful tool for estimating 10-year risk of fatal and non-fatal cardiovascular disease in Germany. PMID:27732641

  4. Report of the American College of Cardiology (ACC) Scientific Sessions 2015, San Diego.

    PubMed

    Murohara, Toyoaki

    2015-01-01

    The 64th Annual Scientific Sessions and Exposition of the American College of Cardiology (ACC) were held at the San Diego Convention Center from March 14-16, 2015. The ACC Scientific Sessions are 1 of 2 major scientific cardiology meetings in the United States, with nearly 20,000 attendees, including 15,000 cardiovascular professionals. There were over 2,100 oral and poster abstracts, and more than 15 late-breaking clinical trials (LBCTs) abstructs. This report presents the highlights and several key presentations, especially the LBCTs, from the ACC Scientific Sessions 2015. I hope this review will help cardiologists update to the latest information.

  5. Report of the American College of Cardiology (ACC) Scientific Sessions 2016, Chicago.

    PubMed

    Mano, Toshiaki; Yamamoto, Kazuhiro

    2016-05-25

    The 65(th)Annual Scientific Sessions of the American College of Cardiology (ACC) were held at McCormick Place, Chicago, from April 2-4, 2016. The ACC Scientific Sessions are one of the 2 major scientific cardiology meetings in the USA and one of the major scientific meetings of cardiology in the world. It had an attendance of 18,769 and over 2,000 oral and poster abstracts, including 8 late-breaking clinical trials. This report presents the key presentations and the highlights from the ACC Scientific Sessions 2016 in Chicago. (Circ J 2016; 80: 1308-1313).

  6. Adenosine deaminase production by an endophytic bacterium (Lysinibacillus sp.) from Avicennia marina.

    PubMed

    Kathiresan, Kandasamy; Saravanakumar, Kandasamy; Sahu, Sunil Kumar; Sivasankaran, Muthu

    2014-06-01

    The present study was carried out with the following objectives: (1) to isolate the endophytic bacilli strains from the leaves of mangrove plant Avicennia marina, (2) to screen the potential strains for the production of adenosine deaminase, (3) to statistically optimize the factors that influence the enzyme activity in the potent strain, and (4) to identify the potent strain using 16S rRNA sequence and construct its phylogenetic tree. The bacterial strains isolated from the fresh leaves of a mangrove A. marina were assessed for adenosine deaminase activity by plating method. Optimization of reaction process was carried out using response surface methodology of central composite design. The potent strain was identified based on 16S rRNA sequencing and phylogeny. Of five endophytic strains, EMLK1 showed a significant deaminase activity over other four strains. The conditions for maximum activity of the isolated adenosine deaminase are described. The potent strain EMLK1 was identified as Lysinibacillus sp. (JQ710723) being the first report as a mangrove endophyte. Mangrove-derived endophytic bacillus strain Lysinibacillus sp. EMLK1 is proved to be a promising source for the production of adenosine deaminase and this enzyme deserves further studies for purification and its application in disease diagnosis.

  7. Glucosamine-6-phosphate deaminase from beef kidney is an allosteric system of the V-type.

    PubMed

    Lara-Lemus, R; Calcagno, M L

    1998-10-14

    The enzyme glucosamine-6-phosphate deaminase from beef kidney has been purified to homogeneity by allosteric-site affinity chromatography. Its amino acid composition and the N-terminal sequence (1-42), were obtained. The amino acid sequence of this segment is essentially identical to the corresponding regions of the human and hamster glucosamine-6-phosphate deaminases. The beef enzyme is a hexamer of 32.5 kDa subunits; this is nearly 2.5 kDa higher than the molecular mass of the homologous enzyme from Escherichia coli. Beef kidney deaminase exhibits a notable difference from the bacterial enzyme in its allosteric activation by N-acetylglucosamine 6-phosphate This metabolite, which is also is the allosteric activator of the bacterial glucosamine-6-phosphate deaminase, activates the enzyme by increasing its kcat without any change in the Km values for glucosamine 6-phosphate, over a wide range of activator concentration. This observation places beef kidney deaminase in the class of V-type allosteric systems.

  8. IMPACC: A Tightly Integrated MPI+OpenACC Framework Exploiting Shared Memory Parallelism

    SciTech Connect

    Lee, Seyong; Vetter, Jeffrey S

    2016-01-01

    We propose IMPACC, an MPI+OpenACC framework for heterogeneous accelerator clusters. IMPACC tightly integrates MPI and OpenACC, while exploiting the shared memory parallelism in the target system. IMPACC dynamically adapts the input MPI+OpenACC applications on the target heterogeneous accelerator clusters to fully exploit target system-specific features. IMPACC provides the programmers with the unified virtual address space, automatic NUMA-friendly task-device mapping, efficient integrated communication routines, seamless streamlining of asynchronous executions, and transparent memory sharing. We have implemented IMPACC and evaluated its performance using three heterogeneous accelerator systems, including Titan supercomputer. Results show that IMPACC can achieve easier programming, higher performance, and better scalability than the current MPI+OpenACC model.

  9. Technology Awareness Workshop on Active Combustion Control (ACC) in Propulsion Systems: JANNAF Combustion Subcommittee Workshop

    NASA Technical Reports Server (NTRS)

    Fry, Ronald S. (Editor); Gannaway, Mary T. (Editor)

    1997-01-01

    A JANNAF Combustion Subcommittee Technology Awareness Seminar on Active Combustion Control (ACC) in Propulsion Systems' was held 12 November 1997 at the NASA Lewis Research Center (LeRC), Cleveland, Ohio. The objectives of the seminar were: 1) Define the need and potential of ACC to meet future requirements for gas turbines and ramjets; 2) Explain general principles of ACC and discuss recent successes to suppress combustion instabilities, increase combustion efficiency, reduce emission, and extend flammability limits; 3) Identify R&D barriers/needs for practical implementation of ACC; 4) Explore potential for improving coordination of future R&D activities funded by various government agencies. Over 40 individuals representing senior management from over 20 industry and government organizations participated. This document summarizes the presentations and findings of this seminar.

  10. OpenACC programs of the Swendsen-Wang multi-cluster spin flip algorithm

    NASA Astrophysics Data System (ADS)

    Komura, Yukihiro

    2015-12-01

    We present sample OpenACC programs of the Swendsen-Wang multi-cluster spin flip algorithm. OpenACC is a directive-based programming model for accelerators without requiring modification to the underlying CPU code itself. In this paper, we deal with the classical spin models as with the sample CUDA programs (Komura and Okabe, 2014), that is, two-dimensional (2D) Ising model, three-dimensional (3D) Ising model, 2D Potts model, 3D Potts model, 2D XY model and 3D XY model. We explain the details of sample OpenACC programs and compare the performance of the present OpenACC implementations with that of the CUDA implementations for the 2D and 3D Ising models and the 2D and 3D XY models.

  11. Expression of ripening-related genes in prickly pear (Opuntia sp.) fruits.

    PubMed

    Collazo-Siqués, P; Valverde, M E; Paredes-López, O; Guevara-Lara, F

    2003-01-01

    To throw light on the expression of ripening-related genes in prickly pear (Opuntia sp.) fruits and on the possible role of the gaseous hormone ethylene in nonclimacteric fruit ripening, cDNA fragments that showed high homologies with 1-aminocyclopropane-1-carboxylate (ACC) synthase and ACC oxidase cDNAs from other plants were cloned and partially characterized. Thus, the corresponding genes were accordingly named opaccs-1 and opacco-1, after Opuntia ACC synthase-1 and Opuntia ACC oxidase-1, respectively. Southern analysis suggests the presence of at least one copy of both genes, as well as other related homologous sequences in the Opuntia genome. Northern analysis of the opaccs-1 gene shows an enhanced expression in ripening fruit tissues, whereas opacco-1 expression is highly induced in ripe tissues with respect to the green fruits and mature cladodes. These results are in agreement with an active metabolic role of ethylene during nonclimacteric prickly pear fruit ripening. This is the first report on the analysis at the molecular level of ripening-related genes of the Opuntia genus.

  12. A role for ethylene in the yellowing of broccoli after harvest

    SciTech Connect

    Tian, M.S.; Downs, C.G.; Lill, R.E.; King, G.A. . Levin Research Center)

    1994-03-01

    Ethylene production from florets of Shogun harvested broccoli (Brassica oleracea L. var. italica) held at 20C in darkness increased as the sepal tissues yellowed. The pattern of respiration rate and ethylene production from branchlets or entire heads was similar, although the magnitude of ethylene and carbon dioxide production appeared to be diluted by the other fleshy stem tissues. The reproductive structures, stamens and pistil, may have a role in determining the rate of sepal degreening, since removing them from florets reduced the yellowing rate. The pistil and stamens also had 7-fold higher levels of 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase activity and more than double the ethylene production of other tissues within the floret. Stamen ACC oxidase activity was high on the first day after harvest, before yellowing became obvious. Changes in ACC oxidase activity of the pistil and stamens mirrored changes in ACC content in these tissues. The climacteric status of harvested broccoli was confirmed by exposure to 0.5% propylene. Propylene stimulated respiration and ethylene production and accelerated yellowing. Broccoli tissues did not respond to propylene immediately after harvest. In tissues aged in air before treatment, the time for response to propylene was shorter, a result suggesting a change in tissue sensitivity. Ethylene exposure induced a dose-dependent decline in hue angle, with 1 ppm ethylene giving the maximum response.

  13. Synthesis and characterization of a novel chitosan based E. coli cytosine deaminase nanocomposite for potential application in prodrug enzyme therapy.

    PubMed

    Yata, Vinod Kumar; Ghosh, Siddhartha Sankar

    2011-01-01

    Cytosine deaminase is a non-mammalian enzyme of widespread interest for prodrug enzyme therapy due to its ability to convert prodrug 5-fluorocytosine into anticancer drug 5-fluorouracil. Cytosine deaminase enzyme has been purified to homogeneity from E. coli K-12 MTCC 1302 strain. K(m) values for cytosine and 5-fluorocytosine were found to be 0.26 mM and 1.82 mM, respectively. We developed a chitosan-entrapped cytosine deaminase nanocomposite. Atomic force microscopy and transmission electron microscopy images showed an elongated sphere shape nanocomposite with an average size of 80 nm diameter. Fourier transform infrared spectroscopy and X-ray diffraction results confirmed gel formation and entrapment of cytosine deaminase within the nanocomposite. Sustained release of cytosine deaminase from the nanocomposite up to one week depicted its potential implication in prodrug inducted enzyme therapy.

  14. Effect of dexamethasone and ACC on bacteria-induced mucin expression in human airway mucosa.

    PubMed

    Hauber, Hans-Peter; Goldmann, Torsten; Vollmer, Ekkehard; Wollenberg, Barbara; Zabel, Peter

    2007-11-01

    Gram-negative bacteria can stimulate mucin production, but excessive mucus supports bacterial infection and consequently leads to airway obstruction. Therefore, the effect of dexamethasone (DEX) and the antioxidant acetyl-cysteine (ACC) on bacteria-induced mucus expression was investigated. Explanted human airway mucosa and mucoepidermoid cells (Calu-3) were stimulated with lipopolysaccharide (LPS) or PAM3 (a synthetic lipoprotein). DEX or ACC were added to either LPS- or PAM3-stimulated airway mucosa or Calu-3 cells. Mucin mRNA expression (MUC5AC) and total mucus glycoconjugates (mucin protein) were quantified using real-time PCR and periodic acid Schiff staining. LPS and PAM3 significantly increased mucin expression in airway mucosa and Calu-3 cells (P < 0.05). DEX alone had no significant effect on mucin expression in airway mucosa or Calu-3 cells (P > 0.05). In contrast, DEX significantly reduced LPS- and PAM3-induced mucin expression in explanted mucosal tissue and mucin expression in Calu-3 cells (P < 0.05). In explanted human airway mucosa ACC alone significantly increased mucin expression (P < 0.05). In contrast, ACC significantly decreased LPS- and PAM3-induced mucin expression (P < 0.05). In Calu-3 cells ACC alone had no significant effect on mucin expression (P > 0.05). ACC decreased LPS- and PAM3-induced mucin expression, but this effect was not significant (P > 0.05). These data suggest that DEX can effectively reduce bacteria-induced mucin expression in the airways. ACC alone may increase mucin expression in noninfected mucosa, but it decreased bacteria-induced mucin expression. Further studies are warranted to evaluate whether the effect of DEX or ACC is clinically relevant.

  15. Autoimmune dysregulation and purine metabolism in adenosine deaminase deficiency.

    PubMed

    Sauer, Aisha Vanessa; Brigida, Immacolata; Carriglio, Nicola; Aiuti, Alessandro

    2012-01-01

    Genetic defects in the adenosine deaminase (ADA) gene are among the most common causes for severe combined immunodeficiency (SCID). ADA-SCID patients suffer from lymphopenia, severely impaired cellular and humoral immunity, failure to thrive, and recurrent infections. Currently available therapeutic options for this otherwise fatal disorder include bone marrow transplantation (BMT), enzyme replacement therapy with bovine ADA (PEG-ADA), or hematopoietic stem cell gene therapy (HSC-GT). Although varying degrees of immune reconstitution can be achieved by these treatments, breakdown of tolerance is a major concern in ADA-SCID. Immune dysregulation such as autoimmune hypothyroidism, diabetes mellitus, hemolytic anemia, and immune thrombocytopenia are frequently observed in milder forms of the disease. However, several reports document similar complications also in patients on long-term PEG-ADA and after BMT or GT treatment. A skewed repertoire and decreased immune functions have been implicated in autoimmunity observed in certain B-cell and/or T-cell immunodeficiencies, but it remains unclear to what extent specific mechanisms of tolerance are affected in ADA deficiency. Herein we provide an overview about ADA-SCID and the autoimmune manifestations reported in these patients before and after treatment. We also assess the value of the ADA-deficient mouse model as a useful tool to study both immune and metabolic disease mechanisms. With focus on regulatory T- and B-cells we discuss the lymphocyte subpopulations particularly prone to contribute to the loss of self-tolerance and onset of autoimmunity in ADA deficiency. Moreover we address which aspects of immune dysregulation are specifically related to alterations in purine metabolism caused by the lack of ADA and the subsequent accumulation of metabolites with immunomodulatory properties.

  16. Inhibition of AMP deaminase as therapeutic target in cardiovascular pathology.

    PubMed

    Zabielska, Magdalena A; Borkowski, Tomasz; Slominska, Ewa M; Smolenski, Ryszard T

    2015-08-01

    AMP deaminase (AMPD; EC 3.5.4.6) catalyzes hydrolysis of the amino group from the adenine ring of AMP resulting in production of inosine 5'-monophosphate (IMP) and ammonia. This reaction helps to maintain healthy cellular energetics by removing excess AMP that accumulates in energy depleted cells. Furthermore, AMPD permits the synthesis of guanine nucleotides from the larger adenylate pool. This enzyme competes with cytosolic 5'-nucleotidases (c5NT) for AMP. Adenosine, a product of c5NT is a vasodilator, antagonizes inotropic effects of catecholamines and exerts anti-platelet, anti-inflammatory and immunosuppressive activities. The ratio of AMPD/c5NT defines the amount of adenosine produced in adenine nucleotide catabolic pathway. Inhibition of AMPD could alter this ratio resulting in increased adenosine production. Besides the potential effect on adenosine production, elevation of AMP due to inhibition of AMPD could also lead to activation of AMP regulated protein kinase (AMPK) with myriad of downstream events including enhanced energetic metabolism, mitochondrial biogenesis and cytoprotection. While the benefits of these processes are well appreciated in cells such as skeletal or cardiac myocytes its role in protection of endothelium could be even more important. Therapeutic use of AMPD inhibition has been limited due to difficulties with obtaining compounds with adequate characteristics. However, endothelium seems to be the easiest target as effective inhibition of AMPD could be achieved at much lower concentration than in the other types of cells. New generation of AMPD inhibitors has recently been established and its testing in context of endothelial and organ protection could provide important basic knowledge and potential therapeutic tools.

  17. Structural and Metabolic Specificity of Methylthiocoformycin for Malarial Adenosine Deaminases

    SciTech Connect

    Ho, M.; Cassera, M; Madrid, D; Ting, L; Tyler, P; Kim, K; Almo, S; Schramm, V

    2009-01-01

    Plasmodium falciparum is a purine auxotroph requiring hypoxanthine as a key metabolic precursor. Erythrocyte adenine nucleotides are the source of the purine precursors, making adenosine deaminase (ADA) a key enzyme in the pathway of hypoxanthine formation. Methylthioadenosine (MTA) is a substrate for most malarial ADAs, but not for human ADA. The catalytic site specificity of malarial ADAs permits methylthiocoformycin (MT-coformycin) to act as a Plasmodium-specific transition state analogue with low affinity for human ADA. The structural basis for MTA and MT-coformycin specificity in malarial ADAs is the subject of speculation. Here, the crystal structure of ADA from Plasmodium vivax (PvADA) in a complex with MT-coformycin reveals an unprecedented binding geometry for 5?-methylthioribosyl groups in the malarial ADAs. Compared to malarial ADA complexes with adenosine or deoxycoformycin, 5?-methylthioribosyl groups are rotated 130 degrees. A hydrogen bonding network between Asp172 and the 3?-hydroxyl of MT-coformycin is essential for recognition of the 5?-methylthioribosyl group. Water occupies the 5?-hydroxyl binding site when MT-coformycin is bound. Mutagenesis of Asp172 destroys the substrate specificity for MTA and MT-coformycin. Kinetic, mutagenic, and structural analyses of PvADA and kinetic analysis of five other Plasmodium ADAs establish the unique structural basis for its specificity for MTA and MT-coformycin. Plasmodium gallinaceum ADA does not use MTA as a substrate, is not inhibited by MT-coformycin, and is missing Asp172. Treatment of P. falciparum cultures with coformycin or MT-coformycin in the presence of MTA is effective in inhibiting parasite growth.

  18. Demonstration of adenosine deaminase activity in human fibroblast lysosomes.

    PubMed Central

    Lindley, E R; Pisoni, R L

    1993-01-01

    Human fibroblast lysosomes, purified on Percoll density gradients, contain an adenosine deaminase (ADA) activity that accounts for approximately 10% of the total ADA activity in GM0010A human fibroblasts. In assays of lysosomal ADA, the conversion of [3H]adenosine into [3H]inosine was proportional to incubation time and the amount of lysosomal material added to reaction mixtures. Maximal activity was observed between pH 7 and 8, and lysosomal ADA displayed a Km of 37 microM for adenosine at 25 degrees C and pH 5.5. Lysosomal ADA was completely inhibited by 2.5 mM Cu2+ or Hg2+ salts, but not by other bivalent cations (Ba2+, Cd2+, Ca2+, Fe2+, Mg2+, Mn2+ and Zn2+). Coformycin (2.5 mM), deoxycoformycin (0.02 mM), 2'-deoxyadenosine (2.5 mM), 6-methylaminopurine riboside (2.5 mM), 2'-3'-isopropylidene-adenosine (2.5 mM) and erythro-9-(2-hydroxy-3-nonyl)adenine (0.2 mM) inhibited lysosomal ADA by > 97%. In contrast, 2.5 mM S-adenosyl-L-homocysteine and cytosine were poor inhibitors. Nearly all lysosomal ADA activity is eluted as a high-molecular-mass protein (> 200 kDa) just after the void volume on a Sephacryl S-200 column, and is very heat-stable, retaining 70% of its activity after incubation at 65 degrees C for 80 min. We speculate that compartmentalization of ADA within lysosomes would allow deamination of adenosine to occur without competition by adenosine kinase, which could assist in maintaining cellular energy requirements under conditions of nutritional deprivation. PMID:8452534

  19. [Conformation of adenosine deaminase in complexes with inhibitors: application of selective quenching of fluorescence emission].

    PubMed

    Vermishian, I G; Sharoian, S G; Antonian, A A; Grigorian, N A; Mardanian, S S; Khoetsian, A V; Markarian, Sh A

    2008-01-01

    The effect of inhibitors, 1-deazaadenosine (1-dAdo) and erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), on the conformation of adenosine deaminase was studied using the method of selective quenching of fluorescence emission by acrylamide, I- and Cs+. Both in free adenosine deaminase and in its complexes with the inhibitors, the wavelength maxima and half-width of the emission characterize the environment of fluorescing tryptophan residues in adenosine deaminase as weak polar with limited access to solvent. The formation of complexes with the ground state inhibitors used did not quench or change the main emission characteristics of tryptophan fluorescence in adenosine deaminase. Small blue shifts of emission maxima were observed upon quenching in all three samples. The Stern-Volmer parameters of tryptophan fluorescence quenching by acrylamide were not essentially influenced by complex formation of the enzyme with the inhibitors: in general, the folding of the enzyme molecule in the complexes is not perturbed. On the contrary, the emission quenching by charged heavy ions, I- and Cs+, in the complexes was hindered in comparison with free adenosine deaminase. In the complex with 1-deazaadenosine, the parameters for quenching by both ions evidence the essential worsening of their interaction with tryptophans. In the complex with erythro-9-(2-hydroxy-3-nonyl)adenine, along with the worse quenching by I-, complete prohibition of quenching by Cs+ was observed. These data indicate that the local environments of fluorescing tryptophan residues is substantially distorted compared with free adenosine deaminase, which leads to their screening from charged heavy ions.

  20. Ozone stress induces the expression of ACC synthase in potato plants

    SciTech Connect

    Schlagnhaufer, C.D.; Arteca, R.N.; Pell, E.J. )

    1993-05-01

    When potato plants (Solanum tuberosum L. cv Norland) are subjected to oxone stress ethylene is emitted. Increases in ethylene production are often the result of increased expression of the enzyme ACC synthase. We used the polymerase chain reaction (PCR) to clone a cDNA encoding an ozone-induced ACC synthase. After treating potato plants with 300 ppb ozone for 4 h, RNA was extracted using a guanidinium isothiocyanate method. Using degenerate oligonucleotides corresponding to several conserved regions of ACC synthase sequences reported from different plant tissues as primers, we were able to reverse transcribe the RNA and amplify a cDNA for ACC synthase. The clone is 1098 bp in length encoding for 386 amino acids comprising [approximately]80% of the protein. Computer analysis of the deduced amino acid sequence showed that our clone is 50-70% homologous with ACC synthase genes cloned from other plant tissues. Using the cDNA as a probe in northern analysis we found that there is little or no expression in control tissue: however there is a large increase in the expression of the ACC synthase message in response to ozone treatment.

  1. Abundance and diversity of archaeal accA gene in hot springs in Yunnan Province, China.

    PubMed

    Song, Zhao-Qi; Wang, Li; Wang, Feng-Ping; Jiang, Hong-Chen; Chen, Jin-Quan; Zhou, En-Min; Liang, Feng; Xiao, Xiang; Li, Wen-Jun

    2013-09-01

    It has been suggested that archaea carrying the accA gene, encoding the alpha subunit of the acetyl CoA carboxylase, autotrophically fix CO2 using the 3-hydroxypropionate/4-hydroxybutyrate pathway in low-temperature environments (e.g., soils, oceans). However, little new information has come to light regarding the occurrence of archaeal accA genes in high-temperature ecosystems. In this study, we investigated the abundance and diversity of archaeal accA gene in hot springs in Yunnan Province, China, using DNA- and RNA-based phylogenetic analyses and quantitative polymerase chain reaction. The results showed that archaeal accA genes were present and expressed in the investigated Yunnan hot springs with a wide range of temperatures (66-96 °C) and pH (4.3-9.0). The majority of the amplified archaeal accA gene sequences were affiliated with the ThAOA/HWCG III [thermophilic ammonia-oxidizing archaea (AOA)/hot water crenarchaeotic group III]. The archaeal accA gene abundance was very close to that of AOA amoA gene, encoding the alpha subunit of ammonia monooxygenase. These data suggest that AOA in terrestrial hot springs might acquire energy from ammonia oxidation coupled with CO2 fixation using the 3-hydroxypropionate/4-hydroxybutyrate pathway.

  2. The Effect of Acute Exercise upon Adenosin Deaminase Oxidant and Antioxidant Activity

    ERIC Educational Resources Information Center

    Kafkas, M. Emin; Karabulut, Aysun Bay; Sahin, Armagan; Otlu, Onder; Savas, Seyfi; Aytac, Aylin

    2012-01-01

    The purpose of this study was to determine the changes of MDA, glutation (GSH), Adenozine deaminase (ADA) and superoxidase dismutaze (SOD) levels with exercise training in obese middle-aged women (body mass index, MMI [greater than or equal to] 30.0). Twelve obese middle-aged women participated in this study. The descriptive statistics of some of…

  3. Improved cytotoxic effects of Salmonella-producing cytosine deaminase in tumour cells.

    PubMed

    Mesa-Pereira, Beatriz; Medina, Carlos; Camacho, Eva María; Flores, Amando; Santero, Eduardo

    2015-01-01

    In order to increase the cytotoxic activity of a Salmonella strain carrying a salicylate-inducible expression system that controls cytosine deaminase production, we have modified both, the vector and the producer bacterium. First, the translation rates of the expression module containing the Escherichia coli codA gene cloned under the control of the Pm promoter have been improved by using the T7 phage gene 10 ribosome binding site sequence and replacing the original GUG start codon by AUG. Second, to increase the time span in which cytosine deaminase may be produced by the bacteria in the presence of 5-fluorocytosine, a 5-fluorouracyl resistant Salmonella strain has been constructed by deleting its upp gene sequence. This new Salmonella strain shows increased cytosine deaminase activity and, after infecting tumour cell cultures, increased cytotoxic and bystander effects under standard induction conditions. In addition, we have generated a purD mutation in the producer strain to control its intracellular proliferation by the presence of adenine and avoid the intrinsic Salmonella cell death induction. This strategy allows the analysis and comparison of the cytotoxic effects of cytosine deaminase produced by different Salmonella strains in tumour cell cultures.

  4. Intracellular localization of human cytidine deaminase. Identification of a functional nuclear localization signal.

    PubMed

    Somasekaram, A; Jarmuz, A; How, A; Scott, J; Navaratnam, N

    1999-10-01

    The cytidine deaminases belong to the family of multisubunit enzymes that catalyze the hydrolytic deamination of their substrate to a corresponding uracil product. They play a major role in pyrimidine nucleoside and nucleotide salvage. The intracellular distribution of cytidine deaminase and related enzymes has previously been considered to be cytosolic. Here we show that human cytidine deaminase (HCDA) is present in the nucleus. A highly specific, affinity purified polyclonal antibody against HCDA was used to analyze the intracellular localization of native HCDA in a variety of mammalian cells by in situ immunochemistry. Native HCDA was found to be present in the nucleus as well as the cytoplasm in several cell types. Indirect immunofluorescence microscopy indicated a predominantly nuclear localization of FLAG-tagged HCDA overexpressed in these cells. We have identified an amino-terminal bipartite nuclear localization signal that is both necessary and sufficient to direct HCDA and a non-nuclear reporter protein to the nucleus. We also show HCDA binding to the nuclear import receptor, importin alpha. Similar putative bipartite nuclear localization sequences are found in other cytidine/deoxycytidylate deaminases. The results presented here suggest that the pyrimidine nucleotide salvage pathway may operate in the nucleus. This localization may have implications in the regulation of nucleoside and nucleotide metabolism and nucleic acid biosynthesis.

  5. Efficient, low-cost protein factories: expression of human adenosine deaminase in baculovirus-infected insect larvae.

    PubMed Central

    Medin, J A; Hunt, L; Gathy, K; Evans, R K; Coleman, M S

    1990-01-01

    Human adenosine deaminase (EC 3.5.4.4), a key purine salvage enzyme essential for immune competence, has been overproduced in Spodoptera frugiperda cells and in Trichoplusia ni (cabbage looper) larvae infected with recombinant baculovirus. The coding sequence of human adenosine deaminase was recombined into a baculovirus immediately downstream from the strong polyhedrin gene promoter. Approximately 60 hr after infection of insect cells with the recombinant virus, maximal levels of intracellular adenosine deaminase mRNA, protein, and enzymatic activity were detected. The recombinant human adenosine deaminase represented 10% of the total cellular protein and exhibited a specific activity of 70 units/mg of protein in crude homogenate. This specific activity is 70-350 times greater than that exhibited by the enzyme in homogenates of the two most abundant natural sources of human adenosine deaminase, thymus and leukemic cells. When the recombinant virus was injected into insect larvae, the maximum recombinant enzyme was produced 4 days postinfection and represented about 2% of the total insect protein with a specific activity of 10-25 units/mg of protein. The recombinant human adenosine deaminase was purified to homogeneity from both insect cells and larvae and demonstrated to be identical to native adenosine deaminase purified from human cells with respect to molecular weight, interaction with polyclonal anti-adenosine deaminase antibody, and enzymatic properties. A pilot purification yielded 8-9 mg of homogeneous enzyme from 22 larvae. The production of large quantities of recombinant human adenosine deaminase in insect larvae is inexpensive and rapid and eliminates the need for specialized facilities for tissue culture. This method should be applicable to large-scale production of many recombinant proteins. Images PMID:2181448

  6. Positive coping styles and perigenual ACC volume: two related mechanisms for conferring resilience?

    PubMed

    Holz, Nathalie E; Boecker, Regina; Jennen-Steinmetz, Christine; Buchmann, Arlette F; Blomeyer, Dorothea; Baumeister, Sarah; Plichta, Michael M; Esser, Günter; Schmidt, Martin; Meyer-Lindenberg, Andreas; Banaschewski, Tobias; Brandeis, Daniel; Laucht, Manfred

    2016-05-01

    Stress exposure has been linked to increased rates of depression and anxiety in adults, particularly in females, and has been associated with maladaptive changes in the anterior cingulate cortex (ACC), which is an important brain structure involved in internalizing disorders. Coping styles are important mediators of the stress reaction by establishing homeostasis, and may thus confer resilience to stress-related psychopathology. Anatomical scans were acquired in 181 healthy participants at age 25 years. Positive coping styles were determined using a self-report questionnaire (German Stress Coping Questionnaire, SVF78) at age 22 years. Adult anxiety and depression symptoms were assessed at ages 22, 23 and 25 years with the Young Adult Self-Report. Information on previous internalizing diagnoses was obtained by diagnostic interview (2-19 years). Positive coping styles were associated with increased ACC volume. ACC volume and positive coping styles predicted anxiety and depression in a sex-dependent manner with increased positive coping and ACC volume being related to lower levels of psychopathology in females, but not in males. These results remained significant when controlled for previous internalizing diagnoses. These findings indicate that positive coping styles and ACC volume are two linked mechanisms, which may serve as protective factors against internalizing disorders.

  7. OpenACC to FPGA: A Framework for Directive-based High-Performance Reconfigurable Computing

    SciTech Connect

    Lee, Seyong; Kim, Jungwon; Vetter, Jeffrey S

    2016-01-01

    This paper presents a directive-based, high-level programming framework for high-performance reconfigurable computing. It takes a standard, portable OpenACC C program as input and generates a hardware configuration file for execution on FPGAs. We implemented this prototype system using our open-source OpenARC compiler; it performs source-to-source translation and optimization of the input OpenACC program into an OpenCL code, which is further compiled into a FPGA program by the backend Altera Offline OpenCL compiler. Internally, the design of OpenARC uses a high- level intermediate representation that separates concerns of program representation from underlying architectures, which facilitates portability of OpenARC. In fact, this design allowed us to create the OpenACC-to-FPGA translation framework with minimal extensions to our existing system. In addition, we show that our proposed FPGA-specific compiler optimizations and novel OpenACC pragma extensions assist the compiler in generating more efficient FPGA hardware configuration files. Our empirical evaluation on an Altera Stratix V FPGA with eight OpenACC benchmarks demonstrate the benefits of our strategy. To demonstrate the portability of OpenARC, we show results for the same benchmarks executing on other heterogeneous platforms, including NVIDIA GPUs, AMD GPUs, and Intel Xeon Phis. This initial evidence helps support the goal of using a directive-based, high-level programming strategy for performance portability across heterogeneous HPC architectures.

  8. Inhibition of ethylene production by putrescine alleviates aluminium-induced root inhibition in wheat plants

    PubMed Central

    Yu, Yan; Jin, Chongwei; Sun, Chengliang; Wang, Jinghong; Ye, Yiquan; Zhou, Weiwei; Lu, Lingli; Lin, Xianyong

    2016-01-01

    Inhibition of root elongation is one of the most distinct symptoms of aluminium (Al) toxicity. Although putrescine (Put) has been identified as an important signaling molecule involved in Al tolerance, it is yet unknown how Put mitigates Al-induced root inhibition. Here, the possible mechanism was investigated by using two wheat genotypes differing in Al resistance: Al-tolerant Xi Aimai-1 and Al-sensitive Yangmai-5. Aluminium caused more root inhibition in Yangmai-5 and increased ethylene production at the root apices compared to Xi Aimai-1, whereas the effects were significantly reversed by ethylene biosynthesis inhibitors. The simultaneous exposure of wheat seedlings to Al and ethylene donor, ethephon, or ethylene biosynthesis precursor, 1-aminocyclopropane-1-carboxylic acid (ACC), increased ethylene production and aggravated root inhibition, which was more pronounced in Xi Aimai-1. In contrast, Put treatment decreased ethylene production and alleviated Al-induced root inhibition in both genotypes, and the effects were more conspicuous in Yangmai-5. Furthermore, our results indicated that Al-induced ethylene production was mediated by ACC synthase (ACS) and ACC oxidase, and that Put decreased ethylene production by inhibiting ACS. Altogether, these findings indicate that ethylene is involved in Al-induced root inhibition and this process could be alleviated by Put through inhibiting ACS activity. PMID:26744061

  9. Antioxidative properties of the essential oil from Pinus mugo.

    PubMed

    Grassmann, Johanna; Hippeli, Susanne; Vollmann, Renate; Elstner, Erich F

    2003-12-17

    The essential oil from Pinus mugo (PMEO) was tested on its antioxidative capacity. For this purpose, several biochemical test systems were chosen (e.g., the Fenton System, the xanthine oxidase assay, or the copper-induced oxidation of low-density lipoprotein (LDL)). The results show that there is moderate or weak antioxidative activity when tested in aqueous environments, like in the Fenton system, xanthine oxidase induced superoxide radical formation, or in the HOCl driven fragmentation of 1-aminocyclopropane-1-carboxylic acid (ACC). In contrast, when tested in more lipophilic environments (e.g., the ACC-cleavage by activated neutrophils in whole blood) the PMEO exhibits good antioxidative activity. PMEO does also show good antioxidative capacity in another lipophilic test system (i.e., the copper induced oxidation of LDL). Some components of PMEO (i.e., Delta(3)-carene, camphene, alpha-pinene, (+)-limonene and terpinolene) were also tested. As the PMEO, they showed weak or no antioxidant activity in aqueous environments, but some of them were effective antioxidants regarding ACC-cleavage by activated neutrophils in whole blood or copper-induced LDL-oxidation. Terpinolene, a minor component of PMEO, exhibited remarkable protection against LDL-oxidation.

  10. Downregulation of Ethylene Production Increases Mycelial Growth and Primordia Formation in the Button Culinary-Medicinal Mushroom, Agaricus bisporus (Agaricomycetes).

    PubMed

    Zhang, Chaohui; Huang, Tao; Shen, Chaohui; Wang, Xiaoting; Qi, Yuancheng; Shen, Jinwen; Song, Andong; Qiu, Liyou; Ai, Yuncan

    2016-01-01

    Ethylene biosynthesis and function in Agaricus bisporus (the button mushroom) remain uncertain. The enzyme activities of 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS) and ACC oxidase (ACO) were detectable in A. bisporus AS2796 and inhibited by α-aminooxyacetic acid and Co2+. We cloned and sequenced 2 ACS genes (Ab-ACS1 and Ab-ACS2) and 1 ACO gene (Ab-ACO) from the mushroom strain. Ab-ACS1 and Ab-ACS2 demonstrated low amino acid sequence similarity. Ab-ACO demonstrated an amino acid sequence completely identical to that of ACO1_AGABI from A. bisporus. Antisense ACO significantly reduced ACO gene expression level, ACO enzyme activity, and ethylene production in the mushroom transformants. The transformants grew faster than the wild-type strain in sterilized compost and normally formed primordia when cultivated in sterilized compost with the sterilized casing vermiculite, but the wild-type strain did not. Our results show that ethylene is synthesized in button mushrooms via the ACC pathway. Ethylene inhibited button mushroom mycelial growth and development.

  11. Inhibition of ethylene production by putrescine alleviates aluminium-induced root inhibition in wheat plants.

    PubMed

    Yu, Yan; Jin, Chongwei; Sun, Chengliang; Wang, Jinghong; Ye, Yiquan; Zhou, Weiwei; Lu, Lingli; Lin, Xianyong

    2016-01-08

    Inhibition of root elongation is one of the most distinct symptoms of aluminium (Al) toxicity. Although putrescine (Put) has been identified as an important signaling molecule involved in Al tolerance, it is yet unknown how Put mitigates Al-induced root inhibition. Here, the possible mechanism was investigated by using two wheat genotypes differing in Al resistance: Al-tolerant Xi Aimai-1 and Al-sensitive Yangmai-5. Aluminium caused more root inhibition in Yangmai-5 and increased ethylene production at the root apices compared to Xi Aimai-1, whereas the effects were significantly reversed by ethylene biosynthesis inhibitors. The simultaneous exposure of wheat seedlings to Al and ethylene donor, ethephon, or ethylene biosynthesis precursor, 1-aminocyclopropane-1-carboxylic acid (ACC), increased ethylene production and aggravated root inhibition, which was more pronounced in Xi Aimai-1. In contrast, Put treatment decreased ethylene production and alleviated Al-induced root inhibition in both genotypes, and the effects were more conspicuous in Yangmai-5. Furthermore, our results indicated that Al-induced ethylene production was mediated by ACC synthase (ACS) and ACC oxidase, and that Put decreased ethylene production by inhibiting ACS. Altogether, these findings indicate that ethylene is involved in Al-induced root inhibition and this process could be alleviated by Put through inhibiting ACS activity.

  12. An ACC Design Method for Achieving Both String Stability and Ride Comfort

    NASA Astrophysics Data System (ADS)

    Yamamura, Yoshinori; Seto, Yoji; Nishira, Hikaru; Kawabe, Taketoshi

    An investigation was made of a method for designing adaptive cruise control (ACC) so as to achieve a headway distance response that feels natural to the driver while at the same time obtaining high levels of both string stability and ride comfort. With this design method, the H∞ norm is adopted as the index of string stability. Additionally, two norms are introduced for evaluating ride comfort and natural vehicle behavior. The relationship between these three norms and headway distance response characteristics was analyzed, and an evaluation method was established for achieving high levels of the various performance characteristics required of ACC. An ACC system designed with this method was evaluated in driving tests conducted on a proving ground course, and the results confirmed that it achieved the targeted levels of string stability, ride comfort and natural vehicle behavior.

  13. Precipitation of ACC in liposomes-a model for biomineralization in confined volumes

    SciTech Connect

    Tester, Chantel C; Wu, Ching-Hsuan; Weigand, Steven; Joester, Derk

    2013-01-10

    Biomineralizing organisms frequently precipitate minerals in small phospholipid bilayer-delineated compartments. We have established an in vitro model system to investigate the effect of confinement in attoliter to femtoliter volumes on the precipitation of calcium carbonate. In particular, we analyze the growth and stabilization of liposome-encapsulated amorphous calcium carbonate (ACC) nanoparticles using a combination of in situ techniques, cryo-transmission electron microscopy (Cryo-TEM), and small angle X-ray scattering (SAXS). Herein, we discuss ACC nanoparticle growth rate as a function of liposome size, carbon dioxide flux across the liposome membrane, pH, and osmotic pressure. Based on these experiments, we argue that the stabilization of ACC nanoparticles in liposomes is a consequence of a low nucleation rate (high activation barrier) of crystalline polymorphs of calcium carbonate.

  14. A feasibility study on porting the community land model onto accelerators using OpenACC

    SciTech Connect

    Wang, Dali; Wu, Wei; Winkler, Frank; Ding, Wei; Hernandez, Oscar R.

    2014-01-01

    As environmental models (such as Accelerated Climate Model for Energy (ACME), Parallel Reactive Flow and Transport Model (PFLOTRAN), Arctic Terrestrial Simulator (ATS), etc.) became more and more complicated, we are facing enormous challenges regarding to porting those applications onto hybrid computing architecture. OpenACC appears as a very promising technology, therefore, we have conducted a feasibility analysis on porting the Community Land Model (CLM), a terrestrial ecosystem model within the Community Earth System Models (CESM)). Specifically, we used automatic function testing platform to extract a small computing kernel out of CLM, then we apply this kernel into the actually CLM dataflow procedure, and investigate the strategy of data parallelization and the benefit of data movement provided by current implementation of OpenACC. Even it is a non-intensive kernel, on a single 16-core computing node, the performance (based on the actual computation time using one GPU) of OpenACC implementation is 2.3 time faster than that of OpenMP implementation using single OpenMP thread, but it is 2.8 times slower than the performance of OpenMP implementation using 16 threads. On multiple nodes, MPI_OpenACC implementation demonstrated very good scalability on up to 128 GPUs on 128 computing nodes. This study also provides useful information for us to look into the potential benefits of “deep copy” capability and “routine” feature of OpenACC standards. In conclusion, we believe that our experience on the environmental model, CLM, can be beneficial to many other scientific research programs who are interested to porting their large scale scientific code using OpenACC onto high-end computers, empowered by hybrid computing architecture.

  15. A feasibility study on porting the community land model onto accelerators using OpenACC

    DOE PAGES

    Wang, Dali; Wu, Wei; Winkler, Frank; ...

    2014-01-01

    As environmental models (such as Accelerated Climate Model for Energy (ACME), Parallel Reactive Flow and Transport Model (PFLOTRAN), Arctic Terrestrial Simulator (ATS), etc.) became more and more complicated, we are facing enormous challenges regarding to porting those applications onto hybrid computing architecture. OpenACC appears as a very promising technology, therefore, we have conducted a feasibility analysis on porting the Community Land Model (CLM), a terrestrial ecosystem model within the Community Earth System Models (CESM)). Specifically, we used automatic function testing platform to extract a small computing kernel out of CLM, then we apply this kernel into the actually CLM dataflowmore » procedure, and investigate the strategy of data parallelization and the benefit of data movement provided by current implementation of OpenACC. Even it is a non-intensive kernel, on a single 16-core computing node, the performance (based on the actual computation time using one GPU) of OpenACC implementation is 2.3 time faster than that of OpenMP implementation using single OpenMP thread, but it is 2.8 times slower than the performance of OpenMP implementation using 16 threads. On multiple nodes, MPI_OpenACC implementation demonstrated very good scalability on up to 128 GPUs on 128 computing nodes. This study also provides useful information for us to look into the potential benefits of “deep copy” capability and “routine” feature of OpenACC standards. In conclusion, we believe that our experience on the environmental model, CLM, can be beneficial to many other scientific research programs who are interested to porting their large scale scientific code using OpenACC onto high-end computers, empowered by hybrid computing architecture.« less

  16. 5th International ACC Symposium: Future and Current Therapeutic Trials in Adrenocortical Carcinoma.

    PubMed

    Hoff, Ana O; Berruti, Alfredo

    2016-02-01

    Adrenocortical carcinoma (ACC) is a rare and complex disease associated with a high mortality rate. Despite intensive translational and clinical research, prognosis remains poor. Over the past decade, a significant effort has been made to develop multinational, collaborative studies to better understand the pathogenesis and clinical features of this rare disease in attempt to improve the therapeutic strategies and patient outcome. The results of both standard and newer treatments are discussed in this review as well as the recent discovery of pathways involved in ACC pathogenesis that provide the rationale to introduce new molecular target therapies. Finally, remaining issues regarding how to improve available therapies in adjuvant setting are raised and addressed.

  17. Synthesis of stable ACC using mesoporous silica gel as a support

    PubMed Central

    2014-01-01

    Stable amorphous calcium carbonate supported by mesoporous silica gel was successfully synthesized. The silica gel support is prepared through the hydrolytic polycondensation of ethyl silicate under suitable conditions. Laser scanning confocal microscopy (LSCM) observations reveal that the morphology of the products is branched with cruciform-like and flower-like structure. Raman spectroscopic analysis and scanning electron microscopy (SEM) observation of the products confirm the combination of stable amorphous calcium carbonate (ACC) nanoparticles and mesoporous silica gel. A possible growth mechanism for the branched structure has been proposed. Results indicate potential application of this work to ACC storage, crystal engineering, biomimetic synthesis, etc. PMID:25246865

  18. An OpenACC-Based Unified Programming Model for Multi-accelerator Systems

    SciTech Connect

    Kim, Jungwon; Lee, Seyong; Vetter, Jeffrey S

    2015-01-01

    This paper proposes a novel SPMD programming model of OpenACC. Our model integrates the different granularities of parallelism from vector-level parallelism to node-level parallelism into a single, unified model based on OpenACC. It allows programmers to write programs for multiple accelerators using a uniform programming model whether they are in shared or distributed memory systems. We implement a prototype of our model and evaluate its performance with a GPU-based supercomputer using three benchmark applications.

  19. L-Serine deaminase activity is induced by exposure of Escherichia coli K-12 to DNA-damaging agents.

    PubMed Central

    Newman, E B; Ahmad, D; Walker, C

    1982-01-01

    The synthesis of L-serine deaminase in Escherichia coli K-12 was induced after exposure of cells to a variety of DNA-damaging agents, including UV irradiation, nalidixic acid, and mitomycin C. Synthesis was also induced during growth at high temperature. A mutant constitutive for SOS functions showed an elevated level of L-serine deaminase activity. The response to DNA-damaging agents thus may be mediated via the SOS system. PMID:6813312

  20. Expression and characterization of a second L-amino acid deaminase isolated from Proteus mirabilis in Escherichia coli.

    PubMed

    Baek, Jin-Oh; Seo, Jeong-Woo; Kwon, Ohsuk; Seong, Su-Il; Kim, Ik-Hwan; Kim, Chul Ho

    2011-04-01

    L-amino acid deaminases catalyze the deamination of natural L-amino acids. Two types of L-amino acid deaminase have been identified in Proteus species. One exhibits high levels of activity toward a wide range of aliphatic and aromatic L-amino acids, typically L-phenylalanine, whereas the other acts on a relatively narrow range of basic L-amino acids, typically L-histidine. In this study, we cloned, expressed, and characterized a second amino acid deaminase, termed Pm1, from P. mirabilis KCTC 2566. Homology alignment of the deduced amino acid sequence of Pm1 demonstrated that the greatest similarity (96%) was with the L-amino acid deaminase (LAD) of P. vulgaris, and that homology with Pma was relatively low (72%). Also, similar to LAD, Pm1 was most active on L-histidine, indicating that Pm1 belongs to the second type of amino acid deaminase. In agreement with this conclusion, the V(max) and K(m) values of Pm1 were 119.7 (μg phenylpyruvic acid/mg/min) and 31.55 mM phenylalanine, respectively, values lower than those of Pma. The Pml deaminase will be very useful industrially in the preparation of commercially valuable materials including urocanic acid and α-oxoglutarate.

  1. GPER agonist G-1 decreases adrenocortical carcinoma (ACC) cell growth in vitro and in vivo

    PubMed Central

    Zolea, Fabiana; Rizza, Pietro; Avena, Paola; Malivindi, Rocco; De Luca, Arianna; Campana, Carmela; Martire, Emilia; Domanico, Francesco; Fallo, Francesco; Carpinelli, Giulia; Cerquetti, Lidia; Amendola, Donatella; Stigliano, Antonio; Pezzi, Vincenzo

    2015-01-01

    We have previously demonstrated that estrogen receptor (ER) alpha (ESR1) increases proliferation of adrenocortical carcinoma (ACC) through both an estrogen-dependent and -independent (induced by IGF-II/IGF1R pathways) manner. Then, the use of tamoxifen, a selective estrogen receptor modulator (SERM), appears effective in reducing ACC growth in vitro and in vivo. However, tamoxifen not only exerts antiestrogenic activity, but also acts as full agonist on the G protein-coupled estrogen receptor (GPER). Aim of this study was to investigate the effect of a non-steroidal GPER agonist G-1 in modulating ACC cell growth. We found that G-1 is able to exert a growth inhibitory effect on H295R cells both in vitro and, as xenograft model, in vivo. Treatment of H295R cells with G-1 induced cell cycle arrest, DNA damage and cell death by the activation of the intrinsic apoptotic mechanism. These events required sustained extracellular regulated kinase (ERK) 1/2 activation. Silencing of GPER by a specific shRNA partially reversed G-1-mediated cell growth inhibition without affecting ERK activation. These data suggest the existence of G-1 activated but GPER-independent effects that remain to be clarified. In conclusion, this study provides a rational to further study G-1 mechanism of action in order to include this drug as a treatment option to the limited therapy of ACC. PMID:26131713

  2. Usability of AcceSS for Web Site Accessibility. Research Report

    ERIC Educational Resources Information Center

    Hackett, Stephanie; Parmanto, Bambang

    2006-01-01

    The standard display of web pages is inadequate for users who are visually impaired. Most visually impaired people obtain information from a web page in a linear fashion via a screen reader, whereas sighted users can immediately obtain a bird's-eye view of a web page's organization and content by quickly scanning the page. AcceSS (which stands for…

  3. AccesSports: A Model for Adapting Mainstream Sports Activities for Individuals with Visual Impairments.

    ERIC Educational Resources Information Center

    Ponchilla, Paul E.

    1995-01-01

    The AccesSports Model allows professionals with basic knowledge of visual impairments and mainstream sports to analyze any sports activity and design adaptations needed for targets or goals, boundaries, and rules to enable individuals with visual impairments to participate. Suggestions for modifying baseball, table tennis, swim racing, wrestling,…

  4. A Cytidine Deaminase Edits C to U in Transfer RNAs in Archaea

    PubMed Central

    Randau, Lennart; Stanley, Bradford J.; Kohlway, Andrew; Mechta, Sarah; Xiong, Yong; Söll, Dieter

    2010-01-01

    All canonical transfer RNAs (tRNAs) have a uridine at position 8, involved in maintaining tRNA tertiary structure. However, the hyperthermophilic archaeon Methanopyrus kandleri harbors 30 (out of 34) tRNA genes with cytidine at position 8. Here, we demonstrate C-to-U editing at this location in the tRNA’s tertiary core, and present the crystal structure of a tRNA-specific cytidine deaminase, CDAT8, which has the cytidine deaminase domain linked to a tRNA-binding THUMP domain. CDAT8 is specific for C deamination at position 8, requires only the acceptor stem hairpin for activity, and belongs to a unique family within the “cytidine deaminase–like” superfamily. The presence of this C-to-U editing enzyme guarantees the proper folding and functionality of all M. kandleri tRNAs. PMID:19407206

  5. Adenosine deaminase complexing protein (ADCP): a transformation sensitive protein with potentials of a cancer marker.

    PubMed

    Herbschleb-Voogt, E; Ten Kate, J; Meera Khan, P

    1983-01-01

    Several observations by independent investigators in the past have indicated that adenosine deaminase complexing protein (ADCP), present in considerable quantities in certain human tissues, was absent or decreased in the cancers originated from them. During the present study, electrophoretic analysis of adenosine deaminase (ADA) isozymes and radioimmunoassay for ADCP in the primary fibroblasts and the transformed as well as certain tumor derived cell lines have demonstrated that ADCP present in large quantities in the primary cells was absent or nearly absent in the transformed or tumor-derived cell lines. Though the mechanisms involved are not yet clear, the above observations indicate that ADCP has the potentials of a useful marker in the studies on transformed cells and cancer tissues.

  6. Targeted base editing in rice and tomato using a CRISPR-Cas9 cytidine deaminase fusion.

    PubMed

    Shimatani, Zenpei; Kashojiya, Sachiko; Takayama, Mariko; Terada, Rie; Arazoe, Takayuki; Ishii, Hisaki; Teramura, Hiroshi; Yamamoto, Tsuyoshi; Komatsu, Hiroki; Miura, Kenji; Ezura, Hiroshi; Nishida, Keiji; Ariizumi, Tohru; Kondo, Akihiko

    2017-03-27

    We applied a fusion of CRISPR-Cas9 and activation-induced cytidine deaminase (Target-AID) for point mutagenesis at genomic regions specified by single guide RNAs (sgRNAs) in two crop plants. In rice, we induced multiple herbicide-resistance point mutations by multiplexed editing using herbicide selection, while in tomato we generated marker-free plants with homozygous heritable DNA substitutions, demonstrating the feasibility of base editing for crop improvement.

  7. Gene therapy for severe combined immunodeficiency due to adenosine deaminase deficiency.

    PubMed

    Montiel-Equihua, Claudia A; Thrasher, Adrian J; Gaspar, H Bobby

    2012-02-01

    The severe combined immunodeficiency caused by the absence of adenosine deaminase (SCID-ADA) was the first monogenic disorder for which gene therapy was developed. Over 30 patients have been treated worldwide using the current protocols, and most of them have experienced clinical benefit; importantly, in the absence of any vector-related complications. In this document, we review the progress made so far in the development and establishment of gene therapy as an alternative form of treatment for ADA-SCID patients.

  8. Alanine-scanning mutagenesis reveals a cytosine deaminase mutant with altered substrate preference.

    PubMed

    Mahan, Sheri D; Ireton, Greg C; Stoddard, Barry L; Black, Margaret E

    2004-07-20

    Suicide gene therapy of cancer is a method whereby cancerous tumors can be selectively eradicated while sparing damage to normal tissue. This is accomplished by delivering a gene, encoding an enzyme capable of specifically converting a nontoxic prodrug into a cytotoxin, to cancer cells followed by prodrug administration. The Escherichia coli gene, codA, encodes cytosine deaminase and is introduced into cancer cells followed by administration of the prodrug 5-fluorocytosine (5-FC). Cytosine deaminase converts 5-FC into cytotoxic 5-fluorouracil, which leads to tumor-cell eradication. One limitation of this enzyme/prodrug combination is that 5-FC is a poor substrate for bacterial cytosine deaminase. The crystal structure of bacterial cytosine deaminase (bCD) reveals that a loop structure in the active site pocket of wild-type bCD comprising residues 310-320 undergoes a conformational change upon cytosine binding, making several contacts to the pyrimidine ring. Alanine-scanning mutagenesis was used to investigate the structure-function relationship of amino acid residues within this region, especially with regard to substrate specificity. Using an E. coli genetic complementation system, seven active mutants were identified (F310A, G311A, H312A, D314A, V315A, F316A, and P318A). Further characterization of these mutants reveals that mutant F316A is 14-fold more efficient than the wild-type at deaminating cytosine to uracil. The mutant D314A enzyme demonstrates a dramatic decrease in cytosine activity (17-fold) as well as a slight increase in activity toward 5-FC (2-fold), indicating that mutant D314A prefers the prodrug over cytosine by almost 20-fold, suggesting that it may be a superior suicide gene.

  9. Yeast Cytosine Deaminase Mutants with Increased Thermostability Impart Sensitivity to 5-Fluorocytosine

    PubMed Central

    Stolworthy, Tiffany S.; Korkegian, Aaron M.; Willmon, Candice L.; Ardiani, Andressa; Cundiff, Jennifer; Stoddard, Barry L.; Black, Margaret E.

    2008-01-01

    SUMMARY Prodrug gene therapy (PGT) is a treatment strategy in which tumor cells are transfected with a 'suicide' gene that encodes a metabolic enzyme capable of converting a nontoxic prodrug into a potent cytotoxin. One of the most promising PGT enzymes is cytosine deaminase (CD), a microbial salvage enzyme that converts cytosine to uracil. CD also converts 5-fluorocytosine (5FC) to 5-fluorouracil (5FU), an inhibitor of DNA synthesis and RNA function. Over 150 studies of cytosine deaminase-mediated PGT applications have been reported since 2000, all using wild-type enzymes. However, various forms of cytosine deaminase are limited by inefficient turnover of 5FC and/or limited thermostability. In a previous study we stabilized and extended the half-life of yeast cytosine deaminase (yCD) by repacking of its hydrophobic core at several positions distant from the active site. Here we report that random mutagenesis of residues selected based on alignment with similar enzymes, followed by selection for enhanced sensitization to 5FC, also produces an enzyme variant (yCD-D92E) with elevated Tm values and increased activity half-life. The new mutation is located at the enzyme's dimer interface, indicating that independent mutational pathways can lead to an increase in the temperature that induces protein unfolding and aggregation in thermal denaturation experiments measured by circular dichroism spectroscopy, and an increase in the half-life of enzyme activity at physiological temperature, as well as more subtle effect on enzyme kinetics. Each independently derived set of mutations significantly improves the enzyme's performance in PGT assays both in cell culture and in animal models. PMID:18291415

  10. Real-time dynamic optical imaging of ACC-M tumor cells killed by HSV-tk/ACV system.

    PubMed

    Xiong, Tao; Li, Yongjin; Li, Zhiyang; Xie, Xiangmo; Lu, Lisha

    2013-01-01

    HSV-tk/ACV induced and killed human adenoid cystic carcinoma cell (ACC-M) in vivo and in vitro, which were observed through optical imaging and green fluorescence protein (GFP) tagging technique. ACC-M was transfected with TK-GFP, and the single clone cell ACC-M-TK-GFP was selected by G418. With fluorescent stereomicroscope, whole-body fluorescent imaging system and fluorescent microscope, we could observe ACV treated ACC-M-TK-GFP cells in cell level and nude mice. The therapies of tumor were visualized clearly with optical imaging. This study proves that optical imaging is a very good approach for studying the effect of HSV-tk/ACV on the ACC-M tumor cells and decreasing the amount of vessel about tumors cell. Optical imaging will become a visual groundwork for monitoring tumor growth and evaluating in vivo curative effect of antitumor drugs.

  11. The Multifaceted Roles of RNA Binding in APOBEC Cytidine Deaminase Functions

    PubMed Central

    Prohaska, Kimberly M.; Bennett, Ryan P.; Salter, Jason D.; Smith, Harold C.

    2014-01-01

    Cytidine deaminases have important roles in the regulation of nucleoside/deoxynucleoside pools for DNA and RNA synthesis. The APOBEC family of cytidine deaminases (named after the first member of the family that was described, Apolipoprotein B mRNA Editing Catalytic Subunit 1, a.k.a. APOBEC1 or A1) is a fascinating group of mutagenic proteins that use RNA and single stranded DNA (ssDNA) as substrates for their cytidine or deoxycytidine deaminase activities. APOBEC proteins and base-modification nucleic acid editing have been the subject of numerous publications, reviews and speculation. These proteins play diverse roles in host cell defense, protecting cells from invading genetic material, enabling the acquired immune response to antigens and changing protein expression at the level of the genetic code in mRNA or DNA. The amazing power these proteins have for interphase cell functions relies on structural and biochemical properties that are beginning to be understood. At the same time, the substrate selectivity of each member in the family and their regulation remains to be elucidated. This review of the APOBEC family will focus on an open question in regulation, namely what role the interactions of these proteins with RNA have in editing substrate recognition or allosteric regulation of DNA mutagenic and host defense activities. PMID:24664896

  12. Evaluation of adenosine deaminase assay for analyzing T-lymphocyte density in vitro.

    PubMed

    Kainthla, Rani Poonam; Kashyap, Rajpal Singh; Prasad, Sweta; Purohit, Hemant J; Taori, Giridhar M; Daginawala, Hatim F

    2006-01-01

    The proliferative capacity of T cells in response to various stimuli is commonly determined by radioactive assay based on incorporation of [3H]thymidine ([3H]TdR) into newly synthesized DNA. In order to assess techniques for application in laboratories where radioactive facilities are not present, an alternative method was tested. As an alternative, T-cell proliferation was measured by spectrophotometrically analyzing the presence of an enzyme adenosine deaminase in lymphocytes and also using a standard XTT assay. Jurkat (human) T-cell line (clone E6.1) was used for lymphocyte population. The Jurkat cell concentration was adjusted according to different cell densities and enzyme activity was determined. Cells were also seeded in complete medium up to 72 h and harvested for estimation of enzyme activity. A significant correlation between the standard cell-proliferation assay and adenosine deaminase assay was observed. The present study indicates that the assay of adenosine deaminase is a reliable and accurate method for measuring proliferation of T lymphocytes.

  13. Cytosine deaminase MX cassettes as positive/negative selectable markers in Saccharomyces cerevisiae.

    PubMed

    Hartzog, Phillip E; Nicholson, Bradly P; McCusker, John H

    2005-07-30

    We describe positive/negative selectable cytosine deaminase MX cassettes for use in Saccharomyces cerevisiae. The basis of positive selection for cytosine deaminase (Fcy1) activity is that (a) fcy1 strains are unable to grow on medium containing cytosine as a sole nitrogen source and (b) fcy1 ura3 strains are unable to grow on medium containing cytosine as the sole pyrimidine source. Conversely, as 5-fluorocytosine (5FC) is toxic to cytosine deaminase-producing cells, fcy1 strains are resistant to 5FC. FCY1MX and FCA1MX cassettes, containing open reading frames (ORFs) of S. cerevisiae FCY1 and Candida albicans FCA1, respectively, were constructed and used to disrupt targeted genes in S. cerevisiae fcy1 strains. In addition, new direct repeat cassettes, kanPR, FCA1PR, FCY1PR and CaURA3PR, were developed to allow efficient deletion of target genes in cells containing MX3 repeats. Finally, the FCY1- and FCA1MX3 or PR direct repeat cassettes can be readily recycled after 5FC counter-selection on both synthetic and rich media.

  14. Magnetic nanoparticle hyperthermia induced cytosine deaminase expression in microencapsulated E. coli for enzyme-prodrug therapy.

    PubMed

    Nemani, Krishnamurthy V; Ennis, Riley C; Griswold, Karl E; Gimi, Barjor

    2015-06-10

    Engineered bacterial cells that are designed to express therapeutic enzymes under the transcriptional control of remotely inducible promoters can mediate the de novo conversion of non-toxic prodrugs to their cytotoxic forms. In situ cellular expression of enzymes provides increased stability and control of enzyme activity as compared to isolated enzymes. We have engineered Escherichia coli (E. coli), designed to express cytosine deaminase at elevated temperatures, under the transcriptional control of thermo-regulatory λpL-cI857 promoter cassette which provides a thermal switch to trigger enzyme synthesis. Enhanced cytosine deaminase expression was observed in cultures incubated at 42°C as compared to 30°C, and enzyme expression was further substantiated by spectrophotometric assays indicating enhanced conversion of 5-fluorocytosine to 5-fluorouracil. The engineered cells were subsequently co-encapsulated with magnetic iron oxide nanoparticles in immunoprotective alginate microcapsules, and cytosine deaminase expression was triggered remotely by alternating magnetic field-induced hyperthermia. The combination of 5-fluorocytosine with AMF-activated microcapsules demonstrated tumor cell cytotoxicity comparable to direct treatment with 5-fluorouracil chemotherapy. Such enzyme-prodrug therapy, based on engineered and immunoisolated E. coli, may ultimately yield an improved therapeutic index relative to monotherapy, as AMF mediated hyperthermia might be expected to pre-sensitize tumors to chemotherapy under appropriate conditions.

  15. Genome-wide target specificities of CRISPR RNA-guided programmable deaminases.

    PubMed

    Kim, Daesik; Lim, Kayeong; Kim, Sang-Tae; Yoon, Sun-Heui; Kim, Kyoungmi; Ryu, Seuk-Min; Kim, Jin-Soo

    2017-04-10

    Cas9-linked deaminases, also called base editors, enable targeted mutation of single nucleotides in eukaryotic genomes. However, their off-target activity is largely unknown. Here we modify digested-genome sequencing (Digenome-seq) to assess the specificity of a programmable deaminase composed of a Cas9 nickase (nCas9) and the deaminase APOBEC1 in the human genome. Genomic DNA is treated with the base editor and a mixture of DNA-modifying enzymes in vitro to produce DNA double-strand breaks (DSBs) at uracil-containing sites. Off-target sites are then computationally identified from whole genome sequencing data. Testing seven different single guide RNAs (sgRNAs), we find that the rAPOBEC1-nCas9 base editor is highly specific, inducing cytosine-to-uracil conversions at only 18 ± 9 sites in the human genome for each sgRNA. Digenome-seq is sensitive enough to capture off-target sites with a substitution frequency of 0.1%. Notably, off-target sites of the base editors are often different from those of Cas9 alone, calling for independent assessment of their genome-wide specificities.

  16. Adenosine Deaminases Acting on RNA (ADARs) are both Antiviral and Proviral Dependent upon the Virus

    PubMed Central

    Samuel, Charles E.

    2010-01-01

    A-to-I RNA editing, the deamination of adenosine (A) to inosine (I) that occurs in regions of RNA with double-stranded character, is catalyzed by a family of Adenosine Deaminases Acting on RNA (ADARs). In mammals there are three ADAR genes. Two encode proteins that possess demonstrated deaminase activity: ADAR1, which is interferon-inducible, and ADAR2 which is constitutively expressed. ADAR3, by contrast, has not yet been shown to bean active enzyme. The specificity of the ADAR1 and ADAR2 deaminases ranges from highly site-selective to non-selective, dependent on the duplex structure of the substrate RNA. A-to-I editing is a form of nucleotide substitution editing, because I is decoded as guanosine (G) instead of A by ribosomes during translation and by polymerases during RNA-dependent RNA replication. Additionally, A-to-I editing can alter RNA structure stability as I:U mismatches are less stable than A:U base pairs. Both viral and cellular RNAs are edited by ADARs. A-to-I editing is of broad physiologic significance. Among the outcomes of A-to-I editing are biochemical changes that affect how viruses interact with their hosts, changes that can lead to either enhanced or reduced virus growth and persistence dependent upon the specific virus. PMID:21211811

  17. Beyond SHM and CSR: AID and related cytidine deaminases in the host response to viral infection.

    PubMed

    Rosenberg, Brad R; Papavasiliou, F Nina

    2007-01-01

    As the primary effector of immunoglobulin somatic hypermutation (SHM) and class switch recombination (CSR), activation-induced cytidine deaminase (AID) serves an important function in the adaptive immune response. Recent advances have demonstrated that AID and a group of closely related cytidine deaminases, the APOBEC3 proteins, also act in the innate host response to viral infection. Antiviral activity was first attributed to APOBEC3G as a potent inhibitor of HIV. It is now apparent that the targets of the APOBEC3 proteins extend beyond HIV, with family members acting against a wide variety of viruses as well as host-encoded retrotransposable genetic elements. Although it appears to function through a different mechanism, AID also possesses antiviral properties. Independent of its antibody diversification functions, AID protects against transformation by Abelson murine leukemia virus (Ab-MLV), an oncogenic retrovirus. Additionally, AID has been implicated in the host response to other pathogenic viruses. These emerging roles for the AID/APOBEC cytidine deaminases in viral infection suggest an intriguing evolutionary connection of innate and adaptive immune mechanisms.

  18. Silicon-mediated changes in polyamines participate in silicon-induced salt tolerance in Sorghum bicolor L.

    PubMed

    Yin, Lina; Wang, Shiwen; Tanaka, Kiyoshi; Fujihara, Shinsuke; Itai, Akihiro; Den, Xiping; Zhang, Suiqi

    2016-02-01

    Silicon (Si) is generally considered a beneficial element for the growth of higher plants, especially under stress conditions, but the mechanisms remain unclear. Here, we tested the hypothesis that Si improves salt tolerance through mediating important metabolism processes rather than acting as a mere mechanical barrier. Seedlings of sorghum (Sorghum bicolor L.) growing in hydroponic culture were treated with NaCl (100 mm) combined with or without Si (0.83 mm). The result showed that supplemental Si enhanced sorghum salt tolerance by decreasing Na(+) accumulation. Simultaneously, polyamine (PA) levels were increased and ethylene precursor (1-aminocyclopropane-1-carboxylic acid: ACC) concentrations were decreased. Several key PA synthesis genes were up-regulated by Si under salt stress. To further confirm the role of PA in Si-mediated salt tolerance, seedlings were exposed to spermidine (Spd) or a PA synthesis inhibitor (dicyclohexylammonium sulphate, DCHA) combined with salt and Si. Exogenous Spd showed similar effects as Si under salt stress whereas exogenous DCHA eliminated Si-enhanced salt tolerance and the beneficial effect of Si in decreasing Na(+) accumulation. These results indicate that PAs and ACC are involved in Si-induced salt tolerance in sorghum and provide evidence that Si plays an active role in mediating salt tolerance.

  19. The interaction with arbuscular mycorrhizal fungi or Trichoderma harzianum alters the shoot hormonal profile in melon plants.

    PubMed

    Martínez-Medina, Ainhoa; Roldán, Antonio; Albacete, Alfonso; Pascual, Jose A

    2011-02-01

    Arbuscular mycorrhizal fungi (AMF) and Trichoderma harzianum are known to affect plant growth and disease resistance through interaction with phytohormone synthesis or transport in the plant. Cross-talk between these microorganisms and their host plants normally occurs in nature and may affect plant resistance. Simultaneous quantification in the shoots of melon plants revealed significant changes in the levels of several hormones in response to inoculation with T. harzianum and two different AMF (Glomus intraradices and Glomus mosseae). Analysis of zeatin (Ze), indole-3-acetic acid (IAA), 1-aminocyclopropane-1-carboxylic acid (ACC), salicylic acid (SA), jasmonic acid (JA) and abscisic acid (ABA) in the shoot showed common and divergent responses of melon plants to G. intraradices and G. mosseae. T. harzianum effected systemic increases in Ze, IAA, ACC, SA, JA and ABA. The interaction of T. harzianum and the AMF with the plant produced a characteristic hormonal profile, which differed from that produced by inoculation with each microorganism singly, suggesting an attenuation of the plant response, related to the hormones SA, JA and ethylene. These results are discussed in relation to their involvement in biomass allocation and basal resistance against Fusarium wilt.

  20. Endogenous hormones response to cytokinins with regard to organogenesis in explants of peach (Prunus persica L. Batsch) cultivars and rootstocks (P. persica × Prunus dulcis).

    PubMed

    Pérez-Jiménez, Margarita; Cantero-Navarro, Elena; Pérez-Alfocea, Francisco; Cos-Terrer, José

    2014-11-01

    Organogenesis in peach (Prunus persica L. Batsch) and peach rootstocks (P. persica × Prunus dulcis) has been achieved and the action of the regeneration medium on 7 phytohormones, zeatin (Z), zeatin riboside (ZR), indole-3-acetic acid (IAA), abscisic acid (ABA), ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), salicylic acid (SA), and jasmonic acid (JA), has been studied using High performance liquid chromatography - mass spectrometry (HPLC-MS/MS). Three scion peach cultivars, 'UFO-3', 'Flariba' and 'Alice Bigi', and the peach × almond rootstocks 'Garnem' and 'GF677' were cultured in two different media, Murashige and Skoog supplemented with plant growth regulators (PGRs) (regeneration medium) and without PGRs (control medium), in order to study the effects of the media and/or genotypes in the endogenous hormones content and their role in organogenesis. The highest regeneration rate was obtained with the peach × almond rootstocks and showed a lower content of Z, IAA, ABA, ACC and JA. Only Z, ZR and IAA were affected by the action of the culture media. This study shows which hormones are external PGRs-dependent and what is the weight of the genotype and hormones in peach organogenesis that provide an avenue to manipulate in vitro organogenesis in peach.

  1. Effects of plant growth-promoting bacteria isolated from copper tailings on plants in sterilized and non-sterilized tailings.

    PubMed

    Liu, Weiqiu; Yang, Chao; Shi, Si; Shu, Wensheng

    2014-02-01

    Ten strains of Cu-tolerant bacteria with potential plant growth-promoting ability were isolated by selecting strains with the ability to use 1-aminocyclopropane-1-carboxylate as a sole nitrogen source (designated ACC-B) or fix nitrogen (designated FLN-B) originating from the rhizosphere of plants growing on copper tailings. All 10 strains proved to have intrinsic ability to produce indole acetic acid and siderophores, and most of them could mobilize insoluble phosphate. In addition, a greenhouse study showed that ACC-B, FLN-B and a mixture of both had similar, potent ability to stimulate growth of Pennisetum purpureum, Medicago sativa and Oenothera erythrosepala plants grown on sterilized tailings. For instance, above-ground biomass of P. purpureum was 278-357% greater after 60d growth on sterilized tailings in their presence. They could also significantly promote the growth of the plants grown on non-sterilized tailings, though the growth-promoting effects were much weaker. So, strategies for using of the plant growth-promoting bacteria in the practice of phytoremediation deserve further studies to get higher growth-promoting efficiency.

  2. Ethylene Production is Associated with Germination but not Seed Dormancy in Red Rice

    PubMed Central

    Gianinetti, Alberto; Laarhoven, Lucas J. J.; Persijn, Stefan T.; Harren, Frans J. M.; Petruzzelli, Luciana

    2007-01-01

    Background and Aims The relationship between ethylene production and both seed dormancy and germination was investigated using red rice (weedy rice) as a model species. Methods Both fully dormant and after-ripened (non-dormant) naked caryopses were incubated with or without inhibitors of ethylene synthesis [aminoethoxyvinylglycine (AVG)] and perception [silver thiosulfate (STS)], or in the presence of the natural ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). The kinetics of ethylene emissions were measured with a sensitive laser–photoacoustic system. Key Results Dormant red rice caryopses did not produce ethylene. In non-dormant caryopses, ethylene evolution never preceded the first visible stage of germination (pericarp splitting), and ethylene inhibitors completely blocked ethylene production, but not pericarp splitting. Accordingly, endogenous ACC appeared to be lacking before pericarp splitting. However, early seedling growth (radicle or coleoptile attaining the length of 1 mm) followed ethylene evolution and was delayed by the inhibitors. Wounding the dormant caryopses induced them to germinate and produce ethylene, but their germination was slow and pericarp splitting could be speeded up by ethylene. Conclusions The findings suggest that, in red rice, endogenous ethylene stimulates the growth of the nascent seedling, but does not affect seed dormancy or germination inception. Correspondingly, this phytohormone does not play a role in the dormancy breakage induced by wounding, but accelerates germination after such breakage has occurred. PMID:17347162

  3. Relationship between endogenous hormonal content and somatic organogenesis in callus of peach (Prunus persica L. Batsch) cultivars and Prunus persica×Prunus dulcis rootstocks.

    PubMed

    Pérez-Jiménez, Margarita; Cantero-Navarro, Elena; Pérez-Alfocea, Francisco; Le-Disquet, Isabel; Guivarc'h, Anne; Cos-Terrer, José

    2014-05-01

    The relationship between endogenous hormones content and the induction of somatic peach plant was studied. To induce multiple shoots from callus derived from the base of stem explants of the scion cultivars 'UFO-3', 'Flariba' and 'Alice Bigi', and the peach×almond rootstocks 'Garnem' and 'GF677', propagated plants were cultured on Murashige and Skoog salts augmented with 0.1mgL(-1) of indolebutyric acid, 1mgL(-1) of 6-benzylaminopurine and 3% sucrose. The highest regeneration rate was obtained with the peach×almond rootstocks. Endogenous levels of abscisic acid (ABA), indole-3-acetic acid (IAA), zeatin (Z), zeatin riboside (ZR), ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), salicylic acid (SA), and jasmonic acid (JA) were analyzed in the organogenic callus. Lower levels of several hormones, namely Z, ZR, ABA, and ACC were found in the peach×almond rootstock compared to peach cultivars, while IAA and SA presented inconclusive returns. These results suggest that the difference in somatic organogenesis capacity observed in peach and peach×almond hybrids is markedly affected by the endogenous hormonal content of the studied genotypes.

  4. Anthropogenic increase in carbon dioxide compromises plant defense against invasive insects

    SciTech Connect

    Zavala, J.; Casteel, C.; DeLucia, E.; Berenbaum, M.

    2008-04-01

    Elevated levels of atmospheric carbon dioxide (CO{sub 2}), a consequence of anthropogenic global change, can profoundly affect the interactions between crop plants and insect pests and may promote yet another form of global change: the rapid establishment of invasive species. Elevated CO{sub 2} increased the susceptibility of soybean plants grown under field conditions to the invasive Japanese beetle (Popillia japonica) and to a variant of western corn rootworm (Diabrotica virgifera virgifera) resistant to crop rotation by down-regulating gene expression related to defense signaling [lipoxygenase 7 (lox7), lipoxygenase 8 (lox8), and 1-aminocyclopropane-1-carboxylate synthase (acc-s)]. The down-regulation of these genes, in turn, reduced the production of cysteine proteinase inhibitors (CystPIs), which are specific deterrents to coleopteran herbivores. Beetle herbivory increased CystPI activity to a greater degree in plants grown under ambient than under elevated CO{sub 2}. Gut cysteine proteinase activity was higher in beetles consuming foliage of soybeans grown under elevated CO{sub 2} than in beetles consuming soybeans grown in ambient CO{sub 2}, consistent with enhanced growth and development of these beetles on plants grown in elevated CO{sub 2}. These findings suggest that predicted increases in soybean productivity under projected elevated CO{sub 2} levels may be reduced by increased susceptibility to invasive crop pests.

  5. Is a cysteine proteinase inhibitor involved in the regulation of petal wilting in senescing carnation (Dianthus caryophyllus L.) flowers?

    PubMed

    Sugawara, Hiroaki; Shibuya, Kenichi; Yoshioka, Toshihito; Hashiba, Teruyoshi; Satoh, Shigeru

    2002-03-01

    Senescence of carnation petals is accompanied by autocatalytic ethylene production and wilting of the petals; the former is caused by the expression of 1-aminocyclopropane-1-carboxylate (ACC) synthase and ACC oxidase genes and the latter is related to the expression of a cysteine proteinase (CPase) gene. CPase is probably responsible for the degradation of proteins, leading to the decomposition of cell components and resultant cell death during the senescence of petals. The carnation plant also has a gene for the CPase inhibitor (DC-CPIn) that is expressed abundantly in petals at the full opening stage of flowers. In the present study, DC-CPIn cDNA was cloned and expressed in E. coli. The recombinant DC-CPIn protein completely inhibited the activities of a proteinase (CPase) extracted from carnation petals and papain. Northern blot analysis showed that the mRNA for CPase (DC-CP1) accumulated in large amounts, whereas that for DC-CPIn disappeared, corresponding to the onset of petal wilting in flowers undergoing natural senescence and exogenous ethylene-induced senescence. Based on these findings, a role of DC-CPIn in the regulation of petal wilting is suggested; DC-CPIn acts as a suppressor of petal wilting, which probably functions to fine-tune petal wilting in contrast to coarse tuning, the up-regulation of CPase activity by gene expression.

  6. Altered cultivar resistance of kimchi cabbage seedlings mediated by salicylic Acid, jasmonic Acid and ethylene.

    PubMed

    Lee, Young Hee; Kim, Sang Hee; Yun, Byung-Wook; Hong, Jeum Kyu

    2014-09-01

    Two cultivars Buram-3-ho (susceptible) and CR-Hagwang (moderate resistant) of kimchi cabbage seedlings showed differential defense responses to anthracnose (Colletotrichum higginsianum), black spot (Alternaria brassicicola) and black rot (Xanthomonas campestris pv. campestris, Xcc) diseases in our previous study. Defense-related hormones salicylic acid (SA), jasmonic acid (JA) and ethylene led to different transcriptional regulation of pathogenesis-related (PR) gene expression in both cultivars. In this study, exogenous application of SA suppressed basal defenses to C. higginsianum in the 1st leaves of the susceptible cultivar and cultivar resistance of the 2nd leaves of the resistant cultivar. SA also enhanced susceptibility of the susceptible cultivar to A. brassicicola. By contrast, SA elevated disease resistance to Xcc in the resistant cultivar, but not in the susceptible cultivar. Methyl jasmonate (MJ) treatment did not affect the disease resistance to C. higginsianum and Xcc in either cultivar, but it compromised the disease resistance to A. brassicicola in the resistant cultivar. Treatment with 1-aminocyclopropane-1-carboxylic acid (ACC) ethylene precursor did not change resistance of the either cultivar to C. higginsianum and Xcc. Effect of ACC pretreatment on the resistance to A. brassicicola was not distinguished between susceptible and resistant cultivars, because cultivar resistance of the resistant cultivar was lost by prolonged moist dark conditions. Taken together, exogenously applied SA, JA and ethylene altered defense signaling crosstalk to three diseases of anthracnose, black spot and black rot in a cultivar-dependent manner.

  7. Tissue specific analysis reveals a differential organization and regulation of both ethylene biosynthesis and E8 during climacteric ripening of tomato

    PubMed Central

    2014-01-01

    Background Solanum lycopersicum or tomato is extensively studied with respect to the ethylene metabolism during climacteric ripening, focusing almost exclusively on fruit pericarp. In this work the ethylene biosynthesis pathway was examined in all major tomato fruit tissues: pericarp, septa, columella, placenta, locular gel and seeds. The tissue specific ethylene production rate was measured throughout fruit development, climacteric ripening and postharvest storage. All ethylene intermediate metabolites (1-aminocyclopropane-1-carboxylic acid (ACC), malonyl-ACC (MACC) and S-adenosyl-L-methionine (SAM)) and enzyme activities (ACC-oxidase (ACO) and ACC-synthase (ACS)) were assessed. Results All tissues showed a similar climacteric pattern in ethylene productions, but with a different amplitude. Profound differences were found between tissue types at the metabolic and enzymatic level. The pericarp tissue produced the highest amount of ethylene, but showed only a low ACC content and limited ACS activity, while the locular gel accumulated a lot of ACC, MACC and SAM and showed only limited ACO and ACS activity. Central tissues (septa, columella and placenta) showed a strong accumulation of ACC and MACC. These differences indicate that the ethylene biosynthesis pathway is organized and regulated in a tissue specific way. The possible role of inter- and intra-tissue transport is discussed to explain these discrepancies. Furthermore, the antagonistic relation between ACO and E8, an ethylene biosynthesis inhibiting protein, was shown to be tissue specific and developmentally regulated. In addition, ethylene inhibition by E8 is not achieved by a direct interaction between ACO and E8, as previously suggested in literature. Conclusions The Ethylene biosynthesis pathway and E8 show a tissue specific and developmental differentiation throughout tomato fruit development and ripening. PMID:24401128

  8. Ethylene regulates lateral root formation and auxin transport in Arabidopsis thaliana

    PubMed Central

    Negi, Sangeeta; Ivanchenko, Maria G; Muday, Gloria K

    2008-01-01

    Lateral root branching is a genetically defined and environmentally regulated process. Auxin is required for lateral root formation, and mutants that are altered in auxin synthesis, transport or signaling often have lateral root defects. Crosstalk between auxin and ethylene in root elongation has been demonstrated, but interactions between these hormones in the regulation of Arabidopsis lateral root formation are not well characterized. This study utilized Arabidopsis mutants altered in ethylene signaling and synthesis to explore the role of ethylene in lateral root formation. We find that enhanced ethylene synthesis or signaling, through the eto1-1 and ctr1-1 mutations, or through the application of 1-aminocyclopropane-1-carboxylic acid (ACC), negatively impacts lateral root formation, and is reversible by treatment with the ethylene antagonist, silver nitrate. In contrast, mutations that block ethylene responses, etr1-3 and ein2-5, enhance root formation and render it insensitive to the effect of ACC, even though these mutants have reduced root elongation at high ACC doses. ACC treatments or the eto1-1 mutation significantly enhance radiolabeled indole-3-acetic acid (IAA) transport in both the acropetal and the basipetal directions. ein2-5 and etr1-3 have less acropetal IAA transport, and transport is no longer regulated by ACC. DR5-GUS reporter expression is also altered by ACC treatment, which is consistent with transport differences. The aux1-7 mutant, which has a defect in an IAA influx protein, is insensitive to the ethylene inhibition of root formation. aux1-7 also has ACC-insensitive acropetal and basipetal IAA transport, as well as altered DR5-GUS expression, which is consistent with ethylene altering AUX1-mediated IAA uptake, and thereby blocking lateral root formation. PMID:18363780

  9. A genome-wide identification and analysis of the DYW-deaminase genes in the pentatricopeptide repeat gene family in cotton (Gossypium spp.)

    PubMed Central

    Liu, Guoyuan; Li, Xue; Guo, Liping; Zhang, Xuexian; Qi, Tingxiang; Wang, Hailin; Tang, Huini; Qiao, Xiuqin; Zhang, Jinfa; Xing, Chaozhu; Wu, Jianyong

    2017-01-01

    The RNA editing occurring in plant organellar genomes mainly involves the change of cytidine to uridine. This process involves a deamination reaction, with cytidine deaminase as the catalyst. Pentatricopeptide repeat (PPR) proteins with a C-terminal DYW domain are reportedly associated with cytidine deamination, similar to members of the deaminase superfamily. PPR genes are involved in many cellular functions and biological processes including fertility restoration to cytoplasmic male sterility (CMS) in plants. In this study, we identified 227 and 211 DYW deaminase-coding PPR genes for the cultivated tetraploid cotton species G. hirsutum and G. barbadense (2n = 4x = 52), respectively, as well as 126 and 97 DYW deaminase-coding PPR genes in the ancestral diploid species G. raimondii and G. arboreum (2n = 26), respectively. The 227 G. hirsutum PPR genes were predicted to encode 52–2016 amino acids, 203 of which were mapped onto 26 chromosomes. Most DYW deaminase genes lacked introns, and their proteins were predicted to target the mitochondria or chloroplasts. Additionally, the DYW domain differed from the complete DYW deaminase domain, which contained part of the E domain and the entire E+ domain. The types and number of DYW tripeptides may have been influenced by evolutionary processes, with some tripeptides being lost. Furthermore, a gene ontology analysis revealed that DYW deaminase functions were mainly related to binding as well as hydrolase and transferase activities. The G. hirsutum DYW deaminase expression profiles varied among different cotton tissues and developmental stages, and no differentially expressed DYW deaminase-coding PPRs were directly associated with the male sterility and restoration in the CMS-D2 system. Our current study provides an important piece of information regarding the structural and evolutionary characteristics of Gossypium DYW-containing PPR genes coding for deaminases and will be useful for characterizing the DYW deaminase gene

  10. Assessing driver's mental representation of Adaptive Cruise Control (ACC) and its possible effects on behavioural adaptations.

    PubMed

    Piccinini, Giulio Francesco; Simões, Anabela; Rodrigues, Carlos Manuel; Leitão, Miguel

    2012-01-01

    The introduction of Adaptive Cruise Control (ACC) could be very helpful for making the longitudinal driving task more comfortable for the drivers and, as a consequence, it could have a global beneficial effect on road safety. However, before or during the usage of the device, due to several reasons, drivers might generate in their mind incomplete or flawed mental representations about the fundamental operation principles of ACC; hence, the resulting usage of the device might be improper, negatively affecting the human-machine interaction and cooperation and, in some cases, leading to negative behavioural adaptations to the system that might neutralise the desirable positive effects on road safety. Within this context, this paper will introduce the methodology which has been developed in order to analyse in detail the topic and foresee, in the future, adequate actions for the recovery of inaccurate mental representations of the system.

  11. Quorum sensing and indole-3-acetic acid degradation play a role in colonization and plant growth promotion of Arabidopsis thaliana by Burkholderia phytofirmans PsJN.

    PubMed

    Zúñiga, Ana; Poupin, María Josefina; Donoso, Raúl; Ledger, Thomas; Guiliani, Nicolás; Gutiérrez, Rodrigo A; González, Bernardo

    2013-05-01

    Although not fully understood, molecular communication in the rhizosphere plays an important role regulating traits involved in plant-bacteria association. Burkholderia phytofirmans PsJN is a well-known plant-growth-promoting bacterium, which establishes rhizospheric and endophytic colonization in different plants. A competent colonization is essential for plant-growth-promoting effects produced by bacteria. Using appropriate mutant strains of B. phytofirmans, we obtained evidence for the importance of N-acyl homoserine lactone-mediated (quorum sensing) cell-to-cell communication in efficient colonization of Arabidopsis thaliana plants and the establishment of a beneficial interaction. We also observed that bacterial degradation of the auxin indole-3-acetic acid (IAA) plays a key role in plant-growth-promoting traits and is necessary for efficient rhizosphere colonization. Wildtype B. phytofirmans but not the iacC mutant in IAA mineralization is able to restore promotion effects in roots of A. thaliana in the presence of exogenously added IAA, indicating the importance of this trait for promoting primary root length. Using a transgenic A. thaliana line with suppressed auxin signaling (miR393) and analyzing the expression of auxin receptors in wild-type inoculated plants, we provide evidence that auxin signaling in plants is necessary for the growth promotion effects produced by B. phytofirmans. The interplay between ethylene and auxin signaling was also confirmed by the response of the plant to a 1-aminocyclopropane-1-carboxylate deaminase bacterial mutant strain.

  12. Multifarious beneficial traits and plant growth promoting potential of Serratia marcescens KiSII and Enterobacter sp. RNF 267 isolated from the rhizosphere of coconut palms (Cocos nucifera L.).

    PubMed

    George, Priya; Gupta, Alka; Gopal, Murali; Thomas, Litty; Thomas, George V

    2013-01-01

    Two plant growth promoting bacteria designated as KiSII and RNF 267 isolated from the rhizosphere of coconut palms were identified as Serratia marcescens and Enterobacter sp. based on their phenotypic features, BIOLOG studies and 16S rRNA gene sequence analysis. Both bacteria exhibited phosphate solubilization, ammonification, and production of indole acetic acid, β-1, 3 glucanase activities and 1-aminocyclopropane-1-carboxylate-deaminase activity. They could also tolerate a range of pH conditions, low temperature and salinity (NaCl). In addition, S. marcescens KiSII exhibited N- fixation potential, chitinase activity, siderophore production and antibiotics production. Seed bacterization with these bacteria increased the growth parameters of test plants such as paddy and cowpea over uninoculated control in green house assay. In coconut seedlings, significant increase in growth and nutrient uptake accompanied with higher populations of plant beneficial microorganisms in their rhizospheres were recorded on inoculation with both the PGPRs. The present study clearly revealed that PGPRs can aid in production of healthy and vigorous seedlings of coconut palm which are hardy perennial crops. They offer a scope to be developed into novel PGPR based bioinoculants for production of elite seedlings that can benefit the coconut farming community and the coconut based ecology.

  13. Bioaccumulation of nickel by E. sativa and role of plant growth promoting rhizobacteria (PGPRs) under nickel stress.

    PubMed

    Kamran, Muhammad Aqeel; Eqani, Syed Ali Musstjab Akber Shah; Bibi, Sadia; Xu, Ren-kou; Amna; Monis, Muhammad Farooq Hussain; Katsoyiannis, Athanasios; Bokhari, Habib; Chaudhary, Hassan Javed

    2016-04-01

    Phytoremediation potential of plants can be enhanced in association with microbes. Further, many plant growth-promoting rhizobacteria can improve growth under stress. The present study was conducted to investigate the effect of Pseudomonas putida (P. putida) on nickel (Ni) uptake and on growth of Eruca sativa (E. sativa). Three different levels of Ni (low; 150 ug/g, medium; 250 ug/g and high; 500 ug/g) were applied to the soil containing E. sativa seedlings, with or without P. putida. Ni-toxicity was measured by metamorphic parameters including shoot length, root length, biomass, chlorophyll and proline and Ni contents. Inoculation with P. putida increased 34% and 41% in root and shoot length and 38% and 24% in fresh, dry weight respectively, as compared to non-inoculated plants. Similarly, Ni uptake increased by up to 46% following P. putida inoculation as compared to non-inoculated plants. Indole acetic acid, siderophore and 1-aminocyclopropane-1-carboxylate deaminase (ACCD) activity in the growing media enhanced growth and Ni uptake in E. sativa. The present results offer insight on Plant Growth Promoting Rhizobacteria (PGPR), such as P. putida, for the potential to enhance the plant growth by inhibiting the adverse effects of Ni in E. sativa.

  14. Biocontrol and plant growth-promoting activity of rhizobacteria from Chinese fields with contaminated soils

    PubMed Central

    Wang, Xuefei; Mavrodi, Dmitri V; Ke, Linfeng; Mavrodi, Olga V; Yang, Mingming; Thomashow, Linda S; Zheng, Na; Weller, David M; Zhang, Jibin

    2015-01-01

    The aim of this study was to inventory the types of plant growth-promoting rhizobacteria (PGPR) present in the rhizosphere of plants grown in soils contaminated with heavy metals, recalcitrant organics, petroleum sewage or salinity in China. We screened 1223 isolates for antifungal activity and about 24% inhibited Rhizoctonia solani or Sclerotinia sclerotiorum. Twenty-four strains inhibitory to R. solani, Gaeumannomyces graminis var. tritici and/or S. sclerotiorum and representing the dominant morphotypes were assayed for PGPR activity. Seven strains contained phlD, prnD, pltC or phzF genes and produced the antibiotics 2,4-diacetylphloroglucinol, pyrrolnitrin, pyoluteorin and phenazines respectively. Six strains contained acdS, which encodes 1-aminocyclopropane-1-carboxylic acid deaminase. Phylogenetic analysis of 16S rDNA and phlD, phzF and acdS genes demonstrated that some strains identified as Pseudomonas were similar to model PGPR strains Pseudomonas protegens Pf-5, Pseudomonas chlororaphis subsp. aureofaciens 30–84 and P. brassicacearum Q8r1-96. Pseudomonas protegens- and P. chlororaphis-like strains had the greatest biocontrol activity against Rhizoctonia root rot and take-all of wheat. Pseudomonas protegens and P. brassicacearum-like strains showed the greatest promotion of canola growth. Our results indicate that strains from contaminated soils are similar to well-described PGPR found in agricultural soils worldwide. Growth-promoting rhizobacteria in polluted soils PMID:25219642

  15. Isolation and characterization of a heavy metal-resistant Burkholderia sp. from heavy metal-contaminated paddy field soil and its potential in promoting plant growth and heavy metal accumulation in metal-polluted soil.

    PubMed

    Jiang, Chun-yu; Sheng, Xia-fang; Qian, Meng; Wang, Qing-ya

    2008-05-01

    A heavy metal-resistant bacterial strain was isolated from heavy metal-contaminated soils and identified as Burkholderia sp. J62 based on the 16S rDNA gene sequence analysis. The heavy metal- and antibiotic resistance, heavy metal solubilization of the isolate were investigated. The isolate was also evaluated for promoting plant growth and Pb and Cd uptakes of the plants from heavy metal-contaminated soils in pot experiments. The isolate was found to exhibit different multiple heavy metal and antibiotic resistance characteristics. Atomic absorption spectrometer analysis showed increased bacterial solubilization of lead and cadmium in solution culture and in soils. The isolate produced indole acetic acid, siderophore and 1-aminocyclopropane-1-carboxylate deaminase. The isolate also solubilized inorganic phosphate. Inoculation with the isolate was found to significantly (p<0.05) increase the biomass of maize and tomato plants. Increase in tissue Pb and Cd contents varied from 38% to 192% and from 5% to 191% in inoculated plants growing in heavy metal-contaminated soils compared to the uninoculated control, respectively. These results show that heavy metal-solubilizing and plant growth promoting bacteria are important for plant growth and heavy metal uptake which may provide a new microbial enhanced-phytoremediation of metal-polluted soils.

  16. Inoculation of Brassica oxyrrhina with plant growth promoting bacteria for the improvement of heavy metal phytoremediation under drought conditions.

    PubMed

    Ma, Ying; Rajkumar, Mani; Zhang, Chang; Freitas, Helena

    2016-12-15

    The aim of this study was to investigate the effects of drought resistant serpentine rhizobacteria on plant growth and metal uptake by Brassica oxyrrhina under drought stress (DS) condition. Two drought resistant serpentine rhizobacterial strains namely Pseudomonas libanensis TR1 and Pseudomonas reactans Ph3R3 were selected based on their ability to stimulate seedling growth in roll towel assay. Further assessment on plant growth promoting (PGP) parameters revealed their ability to produce indole-3-acetic acid, siderophore and 1-aminocyclopropane-1-carboxylate deaminase. Moreover, both strains exhibited high resistance to various heavy metals, antibiotics, salinity and extreme temperature. Inoculation of TR1 and Ph3R3 significantly increased plant growth, leaf relative water and pigment content of B. oxyrrhina, whereas decreased concentrations of proline and malondialdehyde in leaves under metal stress in the absence and presence of DS. Regardless of soil water conditions, TR1 and Ph3R3 greatly improved organ metal concentrations, translocation and bioconcentration factors of Cu and Zn. The successful colonization and metabolic activities of P. libanensis TR1 and P. reactans Ph3R3 represented positive effects on plant development and metal phytoremediation under DS. These results indicate that these strains could be used as bio-inoculants for the improvement of phytoremediation of metal polluted soils under semiarid conditions.

  17. Potential plant growth-promoting strain Bacillus sp. SR-2-1/1 decolorized azo dyes through NADH-ubiquinone:oxidoreductase activity.

    PubMed

    Mahmood, Faisal; Shahid, Muhammad; Hussain, Sabir; Shahzad, Tanvir; Tahir, Muhammad; Ijaz, Muhammad; Hussain, Athar; Mahmood, Khalid; Imran, Muhammad; Babar, Shahid Ali Khan

    2017-03-22

    In this study, a bacterial strain SR-2-1/1 was isolated from textile wastewater-irrigated soil for its concurrent potential of plant growth promotion and azo-dye decolorization. Analysis of 16S rRNA gene sequence confirmed its identity as Bacillus sp. The strain tolerated high concentrations (i.e. up to 1000mgL(-1)) of metals (Ni(2+), Cd(2+), Co(2+), Zn(2+), and Cr(6+)) and efficiently decolorized the azo dyes (i.e. reactive black-5, reactive red-120, direct blue-1 and congo red). It also demonstrated considerable in vitro phosphate solubilizing and 1-aminocyclopropane-1-carboxylic acid deaminase abilities at high metal and salt levels. Bioinformatics analysis of its 537bp azoreductase gene and deduced protein revealed that it decolorized azo dyes through NADH-ubiquinone:oxidoreductase enzyme activity. The deduced protein was predicted structurally and functionally different to those of its closely related database proteins. Thus, the strain SR-2-1/1 is a powerful bioinoculant for bioremediation of textile wastewater contaminated soils in addition to stimulation of plant growth.

  18. Cold-adapted and rhizosphere-competent strain of Rahnella sp. with broad-spectrum plant growth-promotion potential.

    PubMed

    Vyas, Pratibha; Joshi, Robin; Sharma, K C; Rahi, Praveen; Gulati, Ashu; Gulati, Arvind

    2010-12-01

    A phosphate-solubilizing bacterial strain isolated from Hippophae rhamnoides rhizosphere was identified as Rahnella sp. based on its phenotypic features and 16S rRNA gene sequence. The bacterial strain showed the growth characteristics of a cold-adapted psychrotroph, with the multiple plant growth-promoting traits of inorganic and organic phosphate solubilization, 1-aminocyclopropane-1- carboxylate-deaminase activity, ammonia generation, and siderophore production. The strain also produced indole- 3-acetic acid, indole-3-acetaldehyde, indole-3-acetamide, indole-3-acetonitrile, indole-3-lactic acid, and indole-3- pyruvic acid in tryptophan-supplemented nutrient broth. Gluconic, citric and isocitric acids were the major organic acids detected during tricalcium phosphate solubilization. A rifampicin-resistant mutant of the strain exhibited high rhizosphere competence without disturbance to the resident microbial populations in pea rhizosphere. Seed bacterization with a charcoal-based inoculum significantly increased growth in barley, chickpea, pea, and maize under the controlled environment. Microplot testing of the inoculum at two different locations in pea also showed significant increase in growth and yield. The attributes of coldtolerance, high rhizosphere competence, and broad-spectrum plant growth-promoting activity exhibited the potential of Rahnella sp. BIHB 783 for increasing agriculture productivity.

  19. Combined endophytic inoculants enhance nickel phytoextraction from serpentine soil in the hyperaccumulator Noccaea caerulescens

    PubMed Central

    Visioli, Giovanna; Vamerali, Teofilo; Mattarozzi, Monica; Dramis, Lucia; Sanangelantoni, Anna M.

    2015-01-01

    This study assesses the effects of specific bacterial endophytes on the phytoextraction capacity of the Ni-hyperaccumulator Noccaea caerulescens, spontaneously growing in a serpentine soil environment. Five metal-tolerant endophytes had already been selected for their high Ni tolerance (6 mM) and plant growth promoting ability. Here we demonstrate that individual bacterial inoculation is ineffective in enhancing Ni translocation and growth of N. caerulescens in serpentine soil, except for specific strains Ncr-1 and Ncr-8, belonging to the Arthrobacter and Microbacterium genera, which showed the highest indole acetic acid production and 1-aminocyclopropane-1-carboxylic acid-deaminase activity. Ncr-1 and Ncr-8 co-inoculation was even more efficient in promoting plant growth, soil Ni removal, and translocation of Ni, together with that of Fe, Co, and Cu. Bacteria of both strains densely colonized the root surfaces and intercellular spaces of leaf epidermal tissue. These two bacterial strains also turned out to stimulate root length, shoot biomass, and Ni uptake in Arabidopsis thaliana grown in MS agar medium supplemented with Ni. It is concluded that adaptation of N. caerulescens in highly Ni-contaminated serpentine soil can be enhanced by an integrated community of bacterial endophytes rather than by single strains; of the former, Arthrobacter and Microbacterium may be useful candidates for future phytoremediation trials in multiple metal-contaminated sites, with possible extension to non-hyperaccumulator plants. PMID:26322074

  20. Bioprospecting of plant growth promoting psychrotrophic Bacilli from the cold desert of north western Indian Himalayas.

    PubMed

    Yadav, Ajar Nath; Sachan, Shashwati Ghosh; Verma, Priyanka; Saxena, Anil Kumar

    2016-02-01

    The plant growth promoting psychrotrophic Bacilli were investigated from different sites in north western Indian Himalayas. A total of 247 morphotypes were obtained from different soil and water samples and were grouped into 43 clusters based on 16S rDNA-RFLP analysis with three restriction endonucleases. Sequencing of representative isolates has revealed that these 43 Bacilli belonged to different species of 11 genera viz., Desemzia, Exiguobacterium, Jeotgalicoccus, Lysinibacillus, Paenibacillus, Planococcus, Pontibacillus, Sinobaca, Sporosarcina, Staphylococcus and Virgibacillus. With an aim to develop microbial inoculants that can perform efficiently at low temperatures, all representative isolates were screened for different plant growth promoting traits at low temperatures (5-15 degrees C). Among the strains, variations were observed for production (%) of indole-3-acetic acid (20), ammonia (19), siderophores (11), gibberellic acid (4) and hydrogen cyanide (2); solubilisation (%) of zinc (14), phosphate (13) and potassium (7); 1-aminocyclopropane-1-carboxylate deaminase activity (6%) and biocontrol activity (4%) against Rhizoctonia solani and Macrophomina phaseolina. Among all the strains, Bacillus licheniformis, Bacillus muralis, Desemzia incerta, Paenibacillus tylopili and Sporosarcina globispora were found to be potent candidates to be developed as inoculants as they exhibited multiple PGP traits at low temperature.

  1. Bacterial communities associated with Brassica napus L. grown on trace element-contaminated and non-contaminated fields: a genotypic and phenotypic comparison

    PubMed Central

    Croes, S; Weyens, N; Janssen, J; Vercampt, H; Colpaert, JV; Carleer, R; Vangronsveld, J

    2013-01-01

    Summary Cultivable bacterial strains associated with field-grown Brassica napus L. (soil, rhizosphere and roots) from a trace elements (Cd, Zn and Pb) contaminated field and a non-contaminated control field were characterized genotypically and phenotypically. Correspondence analysis of the genotypic data revealed a correlation between soil and rhizosphere communities isolated from the same field, indicating that local conditions play a more important role in influencing the composition of (rhizosphere) soil bacterial communities than root exudates. In contrast, endophytic communities of roots showed a correlation between fields, suggesting that plants on the two fields contain similar obligate endophytes derived from a common seed endophytic community and/or can select bacteria from the rhizosphere. The latter seemed not very likely since, despite the presence of several potential endophytic taxa in the rhizosphere, no significant correlation was found between root and rhizosphere communities. The majority of Cd/Zn tolerant strains capable of phosphorus solubilization, nitrogen fixation, indole-3-acetic acid production and showing 1-aminocyclopropane-1-carboxylate deaminase capacity were found in the rhizosphere and roots of plants growing on the contaminated field. PMID:23594409

  2. Alleviation of salt stress by halotolerant and halophilic plant growth-promoting bacteria in wheat (Triticum aestivum).

    PubMed

    Orhan, Furkan

    2016-01-01

    In the current study, 18 halotolerant and halophilic bacteria have been investigated for their plant growth promoting abilities in vitro and in a hydroponic culture. The bacterial strains have been investigated for ammonia, indole-3-acetic acid and 1-aminocyclopropane-1-carboxylate-deaminase production, phosphate solubilisation and nitrogen fixation activities. Of the tested bacteria, eight were inoculated with Triticum aestivum in a hydroponic culture. The investigated bacterial strains were found to have different plant-growth promoting activities in vitro. Under salt stress (200mM NaCl), the investigated bacterial strains significantly increased the root and shoot length and total fresh weight of the plants. The growth rates of the plants inoculated with bacterial strains ranged from 62.2% to 78.1%. Identifying of novel halophilic and halotolerant bacteria that promote plant growth can be used as alternatives for salt sensitive plants. Extensive research has been conducted on several halophilic and halotolerant bacterial strains to investigate their plant growth promoting activities. However, to the best of my knowledge, this is the first study to inoculate these bacterial strains with wheat.

  3. Effect of heavy metal-solubilizing microorganisms on zinc and cadmium extractions from heavy metal contaminated soil with Tricholoma lobynsis.

    PubMed

    Ji, Ling-yun; Zhang, Wei-wei; Yu, Dong; Cao, Yan-ru; Xu, Heng

    2012-01-01

    The macrofungus, Tricholoma lobynsis, was chosen to remedy Zn-Cd-Pb contaminated soil. To enhance its metal-extracting efficiency, two heavy metal resistant microbes M6 and K1 were applied owing to their excellent abilities to solubilize heavy metal salts. The two isolated microbial strains could also produce indole acetic acid (IAA), siderophore and solubilize inorganic phosphate, but neither of them showed 1-aminocyclopropane-1-carboxylate deaminase activity. The strains M6 and K1 were identified as Serratia marcescens and Rhodotorula mucilaginosa based on 16S rDNA and ITS sequence analysis respectively. Pot experiment showed that spraying to T. lobynsis-inoculated soil with M6 and K1 respectively could increase total Cd accumulations of this mushroom by 216 and 61%, and Zn by 153 and 49% compared to the uninoculated control. Pb accumulation however, was too low (<1 mg kg(-1)) to be determined. The results illustrated that special microbes and macrofungi can work together to remedy polluted soil as plant and plant growth promoting microbes do, probably because of excellent metal-accumulating abilities of macrofungi and IAA-siderophore production, phosphate solubilization abilities of the assisted-microbes. This kind of macrofungi-microbe interaction can be developed into a novel bioremediation strategy.

  4. Significance of diazotrophic plant growth-promoting Herbaspirillum sp. GW103 on phytoextraction of Pband Zn by Zea mays L.

    PubMed

    Praburaman, Loganathan; Park, Sung-Hee; Cho, Min; Lee, Kui-Jae; Ko, Jeong-Ae; Han, Sang-Sub; Lee, Sang-Hyun; Kamala-Kannan, Seralathan; Oh, Byung-Taek

    2017-01-01

    Microbe-assisted phytoremediation has been considered a promising measure for the remediation of heavy metal-polluted soil. The aim of this study was to assess the effect of diazotrophic plant growth-promoting Herbaspirillum sp. GW103 on growth and lead (Pb) and zinc (Zn) accumulation in Zea mays L. The strain GW103 exhibited plant growth-promoting traits such as indole-3-acetic acid, siderophores, and 1-aminocyclopropane-1-carboxylic deaminase. Treatment of Z. mays L. plants with GW103 significantly increased 19, 31, and 52% of plant biomass and 10, 50, and 126% of chlorophyll a contents in Pb, Zn, and Pb + Zn-amended soils, respectively. Similarly, the strain GW103 significantly increased Pb and Zn accumulation in shoots and roots of Z. mays L., which were 77 and 25% in Pb-amended soil, 42 and 73% in Zn-amended soil, and 27 and 84% in Pb + Zn-amended soil. Furthermore, addition of GW103 increased 8, 12, and 7% of total protein content, catalase, and superoxide dismutase levels, respectively, in Z. mays L. plants. The results pointed out that isolate GW103 could potentially reduce the phytotoxicity of metals and increase Pb and Zn accumulation in Z. mays L. plant.

  5. A plant growth-promoting bacterium that decreases nickel toxicity in seedlings

    SciTech Connect

    Burd, G.I.; Dixon, D.G.; Glick, B.R.

    1998-10-01

    A plant growth-promoting bacterium, Kluyvera ascorbata SUD165, that contained high levels of heavy metals was isolated from soil collected near Sudbury, Ontario, Canada. The bacterium was resistant to the toxic effects of Ni{sup 2+}, Pb{sup 2+}, Zn{sup 2+}, and CrO{sub 4}{sup {minus}}, produced a siderophore(s), and displayed 1-aminocyclopropane-1-carboxylic acid deaminase activity. Canola seeds inoculated with this bacterium and then grown under gnotobiotic conditions in the presence of high concentrations of nickel chloride were partially protected against nickel toxicity. In addition, protection by the bacterium against nickel toxicity was evident in pot experiments with canola and tomato seeds. The presence of K. ascorbata SUD165 had no measurable influence on the amount of nickel accumulated per milligram (dry weight) of either roots or shoots of canola plants. Therefore, the bacterial plant growth-promoting effect in the presence of nickel was probably not attributable to the reduction of nickel uptake by seedlings. Rather, it may reflect the ability of the bacterium to lower the level of stress ethylene induced by the nickel.

  6. Biocontrol and plant growth-promoting activity of rhizobacteria from Chinese fields with contaminated soils.

    PubMed

    Wang, Xuefei; Mavrodi, Dmitri V; Ke, Linfeng; Mavrodi, Olga V; Yang, Mingming; Thomashow, Linda S; Zheng, Na; Weller, David M; Zhang, Jibin

    2015-05-01

    The aim of this study was to inventory the types of plant growth-promoting rhizobacteria (PGPR) present in the rhizosphere of plants grown in soils contaminated with heavy metals, recalcitrant organics, petroleum sewage or salinity in China. We screened 1223 isolates for antifungal activity and about 24% inhibited Rhizoctonia solani or Sclerotinia sclerotiorum. Twenty-four strains inhibitory to R. solani, Gaeumannomyces graminis var. tritici and/or S. sclerotiorum and representing the dominant morphotypes were assayed for PGPR activity. Seven strains contained phlD, prnD, pltC or phzF genes and produced the antibiotics 2,4-diacetylphloroglucinol, pyrrolnitrin, pyoluteorin and phenazines respectively. Six strains contained acdS, which encodes 1-aminocyclopropane-1-carboxylic acid deaminase. Phylogenetic analysis of 16S rDNA and phlD, phzF and acdS genes demonstrated that some strains identified as Pseudomonas were similar to model PGPR strains Pseudomonas protegens Pf-5, Pseudomonas chlororaphis subsp. aureofaciens 30-84 and P. brassicacearum Q8r1-96. Pseudomonas protegens- and P. chlororaphis-like strains had the greatest biocontrol activity against Rhizoctonia root rot and take-all of wheat. Pseudomonas protegens and P. brassicacearum-like strains showed the greatest promotion of canola growth. Our results indicate that strains from contaminated soils are similar to well-described PGPR found in agricultural soils worldwide.

  7. Biochemical and Molecular Mechanisms of Plant-Microbe-Metal Interactions: Relevance for Phytoremediation

    PubMed Central

    Ma, Ying; Oliveira, Rui S.; Freitas, Helena; Zhang, Chang

    2016-01-01

    Plants and microbes coexist or compete for survival and their cohesive interactions play a vital role in adapting to metalliferous environments, and can thus be explored to improve microbe-assisted phytoremediation. Plant root exudates are useful nutrient and energy sources for soil microorganisms, with whom they establish intricate communication systems. Some beneficial bacteria and fungi, acting as plant growth promoting microorganisms (PGPMs), may alleviate metal phytotoxicity and stimulate plant growth indirectly via the induction of defense mechanisms against phytopathogens, and/or directly through the solubilization of mineral nutrients (nitrogen, phosphate, potassium, iron, etc.), production of plant growth promoting substances (e.g., phytohormones), and secretion of specific enzymes (e.g., 1-aminocyclopropane-1-carboxylate deaminase). PGPM can also change metal bioavailability in soil through various mechanisms such as acidification, precipitation, chelation, complexation, and redox reactions. This review presents the recent advances and applications made hitherto in understanding the biochemical and molecular mechanisms of plant–microbe interactions and their role in the major processes involved in phytoremediation, such as heavy metal detoxification, mobilization, immobilization, transformation, transport, and distribution. PMID:27446148

  8. Changes in the population of seed bacteria of transgenerationally Cd-exposed Arabidopsis thaliana.

    PubMed

    Truyens, S; Weyens, N; Cuypers, A; Vangronsveld, J

    2013-11-01

    Plant-associated bacteria can have beneficial effects on the growth and health of their host. Nevertheless, the role of endophytic bacteria present in seeds has not been investigated in depth. In this study, the cultivable endophytic population of seeds from Arabidopsis thaliana exposed to 2 μm cadmium for several generations (Cd seeds) was compared with a population isolated from seeds of plants that were never exposed to Cd (control seeds). We observed obvious differences between the two types of seed concerning genera present and phenotypic characteristics of the different isolates. Sinorhizobium sp. and Micrococcus sp. were only found in control seeds, while Pseudomonas sp., Bosea sp. and Paenibacillus sp. were only found in Cd seeds. Sphingomonas sp., Rhizobium sp., Acidovorax sp., Variovorax sp., Methylobacterium sp., Bacillus sp. and Staphylococcus sp. occurred in varying numbers in both types of seed. Metal tolerance and 1-aminocyclopropane-1-carboxylate deaminase activity were predominantly found in strains isolated from Cd seeds, while the production of siderophores, indole-3-acetic acid and organic acids was more prevalent in endophytes isolated from control seeds. These data support the hypothesis that certain endophytes are selected for transfer to the next generation and that their presence might be important for subsequent germination and early seedling development.

  9. Characterization of bacteria in the rhizosphere soils of Polygonum pubescens and their potential in promoting growth and Cd, Pb, Zn uptake by Brassica napus.

    PubMed

    Jing, Yuan Xiao; Yan, Jun Lan; He, Huai Dong; Yang, Dan Jing; Xiao, Li; Zhong, Ting; Yuan, Ming; Cai, Xin De; Li, Shu Bin

    2014-01-01

    Microbe-enhanced phytoremediation has been considered as a promising measure for the remediation of metal-contaminated soils. In this study, two bacterial strains JYX7 and JYX10 were isolated from rhizosphere soils of Polygonum pubescens grown in metal-polluted soil and identified as of Enterobacter sp. and Klebsiella sp. based on 16S rDNA sequences, respectively. JYX7 and JYX10 showed high Cd, Pb and Zn tolerance and increased water-soluble Cd, Pb and Zn concentrations in culture solution and metal-added soils. Two isolates produced plant growth-promoting substances such as indole acetic acid, siderophore, 1-aminocyclopropane-1-carboxylic deaminase, and solubilized inorganic phosphate. Based upon their ability in metal tolerance and solubilization, two isolates were further studied for their effects on growth and accumulation of Cd, Pb, and Zn in Brassica napus (rape) by pot experiments. Rapes inoculated with JYX7 and JYX10 had significantly higher dry weights, concentrations and uptakes of Cd, Pb, Zn in both above-ground and root tissues than those without inoculation grown in soils amended with Cd (25 mg kg(-1)), Pb (200 mg kg(-1)) or Zn (200 mg kg(-1)). The present results demonstrated that JYX7 and JYX10 are valuable microorganism, which can improve the efficiency of phytoremediation in soils polluted by Cd, Pb, and Zn.

  10. Characterization of plant-growth-promoting effects and concurrent promotion of heavy metal accumulation in the tissues of the plants grown in the polluted soil by Burkholderia strain LD-11.

    PubMed

    Huang, Gui-Hai; Tian, Hui-Hui; Liu, Hai-Ying; Fan, Xian-Wei; Liang, Yu; Li, You-Zhi

    2013-01-01

    Plant-growth-promoting (PGP) bacteria especially with the resistance to multiple heavy metals are helpful to phytoremediation. Further development of PGP bacteria is very necessary because of the extreme diversity of plants, soils, and heavy metal pollution. A Burkholderia sp. strain, numbered LD-11, was isolated, which showed resistances to multiple heavy metals and antibiotics. It can produce indole-3-acetic acid, 1-aminocyclopropane-1-carboxylic acid deaminase and siderophores. Inoculation with the LD-11 improved germination of seeds of the investigated vegetable plants in the presence of Cu, promoted elongation of roots and hypocotyledonary axes, enhanced the dry weights of the plants grown in the soils polluted with Cu and/or Pb, and increased activity of the soil urease and the rhizobacteria diversity. Inoculation with the LD-11 significantly enhanced Cu and/or Pb accumulation especially in the roots of the plants grown in the polluted soils. Notably, LD-11 could produce siderophores in the presence of Cu. Conclusively, the PGP effects and concurrent heavy metal accumulation in the plant tissues results from combined effects of the above-mentioned multiple factors. Cu is an important element that represses production of the siderophore by the bacteria. Phytoremediation by synergistic use of the investigated plants and the bacterial strain LD-11 is a phytoextraction process.

  11. Diversity of bacterial endophytes in 3 and 15 year-old grapevines of Vitis vinifera cv. Corvina and their potential for plant growth promotion and phytopathogen control.

    PubMed

    Andreolli, Marco; Lampis, Silvia; Zapparoli, Giacomo; Angelini, Elisa; Vallini, Giovanni

    2016-02-01

    This study represents the first investigation on ecology of endophytic bacteria isolated from 3 and 15 year-old vine stems of Vitis vinifera cv. Corvina. The analysis was performed by means of culture-dependent techniques. The obtained results showed that new grapevine endophytic genera are being discovered. Moreover, Bacilli and Actinobacteria are frequently isolated from 3 year-old plants, whereas Alpha- and Gamma- Proteobacteria classes are more prevalent in the 15 year-old plants. Shannon-Wiener (H) index and analysis of rarefaction curves revealed greater genus richness in young grapevine plants. Furthermore, results evidenced an increase of genotypic group number within specific genera (e.g., Rhizobium and Pantoea). Among isolated strains from 3 and 15 year-old stems, respectively, 34 and 39% produce siderophores; 22 and 15% secrete ammonia; 22 and 21% produce indole-3-acetic acid; 8.7 and 41% solubilize phosphate. Besides, two strains isolated from 15 year-old grapevines showed 1-aminocyclopropane-1-carboxylate deaminase activity. Antifungal activity analysis evidenced that two Bacillus strains possess growth antagonistic effect toward all the tested fungal strains. Therefore, the present study extends our knowledge of the diversity of the endophytic bacteria by providing new insights into the complexity of the grapevine microbiome.

  12. Isolation of Endophytic Plant Growth-Promoting Bacteria Associated with the Halophyte Salicornia europaea and Evaluation of their Promoting Activity Under Salt Stress.

    PubMed

    Zhao, Shuai; Zhou, Na; Zhao, Zheng-Yong; Zhang, Ke; Wu, Guo-Hua; Tian, Chang-Yan

    2016-10-01

    Several reports have highlighted that many plant growth-promoting endophytic bacteria (PGPE) can assist their host plants in coping with various biotic and abiotic stresses. However, information about the PGPE colonizing in the halophytes is still scarce. This study was designed to isolate and characterize PGPE from salt-accumulating halophyte Salicornia europaea grown under extreme salinity and to evaluate in vitro the bacterial mechanisms related to plant growth promotion. A total of 105 isolates were obtained from the surface-sterilized roots, stems, and assimilation twigs of S. europaea. Thirty-two isolates were initially selected for their ability to produce 1-aminocyclopropane-1-carboxylate deaminase as well as other properties such as production of indole-3-acetic acid and phosphate-solubilizing activities. The 16S rRNA gene-sequencing analysis revealed that these isolates belong to 13 different genera and 19 bacterial species. For these 32 strains, seed germination and seedling growth in axenically grown S. europaea seedlings at different NaCl concentrations (50-500 mM) were quantified. Five isolates possessing significant stimulation of the host plant growth were obtained. The five isolates were identified as Bacillus endophyticus, Bacillus tequilensis, Planococcus rifietoensis, Variovorax paradoxus, and Arthrobacter agilis. All the five strains could colonize and can be reisolated from the host plant interior tissues. These results demonstrate that habitat-adapted PGPE isolated from halophyte could enhance plant growth under saline stress conditions.

  13. Efficiency of plant growth-promoting P-solubilizing Bacillus circulans CB7 for enhancement of tomato growth under net house conditions.

    PubMed

    Mehta, Preeti; Walia, Abhishek; Kulshrestha, Saurabh; Chauhan, Anjali; Shirkot, Chand Karan

    2015-01-01

    P-solubilizing bacterial isolate CB7 isolated from apple rhizosphere soil of Himachal Pradesh, India was identified as Bacillus circulans on the basis of phenotypic characteristics, biochemical tests, fatty acid methyl esters analysis, and 16S rRNA gene sequence. The isolate exhibited plant growth-promoting traits of P-solubilization, auxin, 1-aminocyclopropane-1-carboxylate deaminase activity, siderophore, nitrogenase activity, and antagonistic activity against Dematophora necatrix. In vitro studies revealed that P-solubilization and other plant growth-promoting traits were dependent on the presence of glucose in PVK medium and removal of yeast extract had no significant effect on plant growth-promoting traits. Plant growth-promoting traits of isolate CB7 were repressed in the presence of KH2 PO4 . P-solubilization activity was associated with the release of organic acids and a drop in the pH of the Pikovskaya's medium. HPLC analysis detected gluconic and citric acid as major organic acids in the course of P-solubilization. Remarkable increase was observed in seed germination (22.32%), shoot length (15.91%), root length (25.10%), shoot dry weight (52.92%) and root dry weight (31.4%), nitrogen (18.75%), potassium (57.69%), and phosphorus (22.22%) content of shoot biomass over control. These results demonstrate that isolate CB7 has the promising PGPR attributes to be developed as a biofertilizer to enhance soil fertility and promote plant growth.

  14. The ACC strategy in biomineralization: the case of earthworm's amorphous spherulites

    NASA Astrophysics Data System (ADS)

    Briones, Maria J. I.; Alvarez-Otero, Rosa; Méndez, Jesús; Gago Duport, Luis

    2010-05-01

    The occurrence of amorphous calcium carbonate (ACC), an hydrated and highly soluble form of solid CaCO3, seems to be a common feature in all carbonate forming organisms such as mollusks, corals, echinoderms and crustaceans. The ubiquity of ACC in these Ca-carbonate biomineralizing systems, as a precursor of further crystalline phases, has recently open the interesting question about if the formation of an amorphous phase is a necessary step in the calcification process of all organisms and consequently, whether it would be possible to define the "amorphous precursor estategy" as a general mechanism in biomineralization. Neverthelees, although ACC appears to be widespread in these organisms very little is known about its particular role in the biomineralization scheme of the different phyla. The formation of CaCO3 spherulites in the calciferous glands of earthworms is a particular case of calcareous biomineralization involving the presence of ACC as a transient precursor phase [2]. Interestingly, the formation of crystalline carbonates via ACC in these organisms is not connected with skeleton building so it must play another functional role. In addition, the transient transformation stages can be followed by in situ spectrometric techniques and therefore, earthworms provide and adequate model to analyse the mutual interactions between ACC-solvent-and crystalline phases. In this study, we have analysed the morphological and structural transformations from the initial ACC spherulites until the formation of the crystalline phases: vaterite (and/or aragonite) and finally calcite, is accomplished. The characterization of ACC was done by performing in situ FT-IR, together with and HREM and Debye scherrer -XRD. The structural results were interpreted in the light of the histological study of the gland. The geometry of the secretory epithelium of the calciferous gland, as evidenced by TEM [2], shows the presence of irregulary shaped cells with their apical surface

  15. The kinetics and mechanisms of amorphous calcium carbonate (ACC) crystallization to calcite, via vaterite.

    PubMed

    Rodriguez-Blanco, Juan Diego; Shaw, Samuel; Benning, Liane G

    2011-01-01

    The kinetics and mechanisms of nanoparticulate amorphous calcium carbonate (ACC) crystallization to calcite, via vaterite, were studied at a range of environmentally relevant temperatures (7.5-25 °C) using synchrotron-based in situ time-resolved Energy Dispersive X-ray Diffraction (ED-XRD) in conjunction with high-resolution electron microscopy, ex situ X-ray diffraction and infrared spectroscopy. The crystallization process occurs in two stages; firstly, the particles of ACC rapidly dehydrate and crystallize to form individual particles of vaterite; secondly, the vaterite transforms to calcite via a dissolution and reprecipitation mechanism with the reaction rate controlled by the surface area of calcite. The second stage of the reaction is approximately 10 times slower than the first. Activation energies of calcite nucleation and crystallization are 73±10 and 66±2 kJ mol(-1), respectively. A model to calculate the degree of calcite crystallization from ACC at environmentally relevant temperatures (7.5-40 °C) is also presented.

  16. Ethylene-auxin interactions regulate lateral root initiation and emergence in Arabidopsis thaliana.

    PubMed

    Ivanchenko, Maria G; Muday, Gloria K; Dubrovsky, Joseph G

    2008-07-01

    Plant root systems display considerable plasticity in response to endogenous and environmental signals. Auxin stimulates pericycle cells within elongating primary roots to enter de novo organogenesis, leading to the establishment of new lateral root meristems. Crosstalk between auxin and ethylene in root elongation has been demonstrated, but interactions between these hormones in root branching are not well characterized. We find that enhanced ethylene synthesis, resulting from the application of low concentrations of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), promotes the initiation of lateral root primordia. Treatment with higher doses of ACC strongly inhibits the ability of pericycle cells to initiate new lateral root primordia, but promotes the emergence of existing lateral root primordia: behaviour that is also seen in the eto1 mutation. These effects are correlated with decreased pericycle cell length and increased lateral root primordia cell width. When auxin is applied simultaneously with ACC, ACC is unable to prevent the auxin stimulation of lateral root formation in the root tissues formed prior to ACC exposure. However, in root tissues formed after transfer to ACC, in which elongation is reduced, auxin does not rescue the ethylene inhibition of primordia initiation, but instead increases it by several fold. Mutations that block auxin responses, slr1 and arf7 arf19, render initiation of lateral root primordia insensitive to the promoting effect of low ethylene levels, and mutations that inhibit ethylene-stimulated auxin biosynthesis, wei2 and wei7, reduce the inhibitory effect of higher ethylene levels, consistent with ethylene regulating root branching through interactions with auxin.

  17. Dormancy removal in apple embryos by nitric oxide or cyanide involves modifications in ethylene biosynthetic pathway.

    PubMed

    Gniazdowska, Agnieszka; Krasuska, Urszula; Bogatek, Renata

    2010-11-01

    The connection between classical phytohormone-ethylene and two signaling molecules, nitric oxide (NO) and hydrogen cyanide (HCN), was investigated in dormancy removal and germination "sensu stricto" of apple (Malus domestica Borkh.) embryos. Deep dormancy of apple embryos was removed by short-term (3-6 h) pre-treatment with NO or HCN. NO- or HCN-mediated stimulation of germination was associated with enhanced emission of ethylene by the embryos, coupled with transient increase in ROS concentration in embryos. Ethylene vapors stimulated germination of dormant apple embryos and eliminated morphological anomalies characteristic for young seedlings developed from dormant embryos. Inhibitors of ethylene receptors completely impeded beneficial effect of NO and HCN on embryo germination. NO- and HCN-induced ethylene emission by apple embryo was only slightly reduced by inhibitor of 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase activity during first 4 days of germination. Short-term pre-treatment of the embryos with NO and HCN modified activity of both key enzymes of ethylene biosynthetic pathway: ACC synthase and ACC oxidase. Activity of ACC synthase declined during first 4 days of germination, while activity of ACC oxidase increased markedly at that time. Additional experiments point to non-enzymatic conversion of ACC to ethylene in the presence of ROS (H(2)O(2)). The results indicate that NO and HCN may alleviate dormancy of apple embryos "via" transient accumulation of ROS, leading to enhanced ethylene emission which is required to terminate germination "sensu stricto". Therefore, ethylene seems to be a trigger factor in control of apple embryo dormancy removal and germination.

  18. Red light regulation of ethylene biosynthesis and gravitropism in etiolated pea stems

    NASA Technical Reports Server (NTRS)

    Steed, C. L.; Taylor, L. K.; Harrison, M. A.

    2004-01-01

    During gravitropism, the accumulation of auxin in the lower side of the stem causes increased growth and the subsequent curvature, while the gaseous hormone ethylene plays a modulating role in regulating the kinetics of growth asymmetries. Light also contributes to the control of gravitropic curvature, potentially through its interaction with ethylene biosynthesis. In this study, red-light pulse treatment of etiolated pea epicotyls was evaluated for its effect on ethylene biosynthesis during gravitropic curvature. Ethylene biosynthesis analysis included measurements of ethylene; the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC); malonyl-conjugated ACC (MACC); and expression levels of pea ACC oxidase (Ps-ACO1) and ACC synthase (Ps-ACS1, Ps-ACS2) genes by reverse transcriptase-polymerase chain reaction analysis. Red-pulsed seedlings were given a 6 min pulse of 11 micromoles m-2 s-1 red-light 15 h prior to horizontal reorientation for consistency with the timeline of red-light inhibition of ethylene production. Red-pulse treatment significantly reduced ethylene production and MACC levels in epicotyl tissue. However, there was no effect of red-pulse treatment on ACC level, or expression of ACS or ACO genes. During gravitropic curvature, ethylene production increased from 60 to 120 min after horizontal placement in both control and red-pulsed epicotyls. In red-pulsed tissues, ACC levels increased by 120 min after horizontal reorientation, accompanied by decreased MACC levels in the lower portion of the epicotyl. Overall, our results demonstrate that ethylene production in etiolated epicotyls increases after the initiation of curvature. This ethylene increase may inhibit cell growth in the lower portion of the epicotyl and contribute to tip straightening and reduced overall curvature observed after the initial 60 min of curvature in etiolated pea epicotyls.

  19. Identification of two pentatricopeptide repeat genes required for RNA editing and zinc binding by C-terminal cytidine deaminase-like domains.

    PubMed

    Hayes, Michael L; Giang, Karolyn; Berhane, Beniam; Mulligan, R Michael

    2013-12-20

    Many transcripts expressed from plant organelle genomes are modified by C-to-U RNA editing. Nuclear encoded pentatricopeptide repeat (PPR) proteins are required as RNA binding specificity determinants in the RNA editing mechanism. Bioinformatic analysis has shown that most of the Arabidopsis PPR proteins necessary for RNA editing events include a C-terminal portion that shares structural characteristics with a superfamily of deaminases. The DYW deaminase domain includes a highly conserved zinc binding motif that shares characteristics with cytidine deaminases. The Arabidopsis PPR genes, ELI1 and DOT4, both have DYW deaminase domains and are required for single RNA editing events in chloroplasts. The ELI1 DYW deaminase domain was expressed as a recombinant protein in Escherichia coli and was shown to bind two zinc atoms per polypeptide. Thus, the DYW deaminase domain binds a zinc metal ion, as expected for a cytidine deaminase, and is potentially the catalytic component of an editing complex. Genetic complementation experiments demonstrate that large portions of the DYW deaminase domain of ELI1 may be eliminated, but the truncated genes retain the ability to restore editing site conversion in a mutant plant. These results suggest that the catalytic activity can be supplied in trans by uncharacterized protein(s) of the editosome.

  20. Transcriptional pausing and stalling causes multiple clustered mutations by human activation-induced deaminase

    PubMed Central

    Canugovi, Chandrika; Samaranayake, Mala; Bhagwat, Ashok S.

    2009-01-01

    Transcription of the rearranged immunoglobulin gene and expression of the enzyme activation-induced deaminase (AID) are essential for somatic hypermutations of this gene during antibody maturation. While AID acts as a single-strand DNA-cytosine deaminase creating U · G mispairs that lead to mutations, the role played by transcription in this process is less clear. We have used in vitro transcription of the kan gene by the T7 RNA polymerase (RNAP) in the presence of AID and a genetic reversion assay for kanamycin-resistance to investigate the causes of multiple clustered mutations (MCMs) during somatic hypermutations. We find that, depending on transcription conditions, AID can cause single-base substitutions or MCMs. When wild-type RNAP is used for transcription at physiologically relevant concentrations of ribonucleoside triphosphates (NTPs), few MCMs are found. In contrast, slowing the rate of elongation by reducing the NTP concentration or using a mutant RNAP increases several-fold the percent of revertants containing MCMs. Arresting the elongation complexes by a quick removal of NTPs leads to formation of RNA-DNA hybrids (R-loops). Treatment of these structures with AID results in a high percentage of KanR revertants with MCMs. Furthermore, selecting for transcription elongation complexes stalled near the codon that suffers mutations during acquisition of kanamycin-resistance results in an overwhelming majority of revertants with MCMs. These results show that if RNAP II pauses or stalls during transcription of immunoglobulin gene, AID is likely to promote MCMs. As changes in physiological conditions such as occurrence of certain DNA primary or secondary structures or DNA adducts are known to cause transcriptional pausing and stalling in mammalian cells, this process may cause MCMs during somatic hypermutation.—Canugovi, C., Samaranayake, M., Bhagwat, A. S. Transcriptional pausing and stalling causes multiple clustered mutations by human activation

  1. Significance of the D-serine-deaminase and D-serine metabolism of Staphylococcus saprophyticus for virulence.

    PubMed

    Korte-Berwanger, Miriam; Sakinc, Türkan; Kline, Kimberly; Nielsen, Hailyn V; Hultgren, Scott; Gatermann, Sören G

    2013-12-01

    Staphylococcus saprophyticus is the only species of Staphylococcus that is typically uropathogenic and possesses a gene coding for a D-serine-deaminase (DsdA). As D-serine is prevalent in urine and toxic or bacteriostatic to many bacteria, it is not surprising that the D-serine-deaminase gene is found in the genome of uropathogens. It has been suggested that D-serine-deaminase or the ability to respond to or to metabolize D-serine is important for virulence. For uropathogenic Escherichia coli (UPEC), a high intracellular D-serine concentration affects expression of virulence factors. S. saprophyticus is able to grow in the presence of high D-serine concentrations; however, its D-serine metabolism has not been described. The activity of the D-serine-deaminase was verified by analyzing the formation of pyruvate from D-serine in different strains with and without D-serine-deaminase. Cocultivation experiments were performed to show that D-serine-deaminase confers a growth advantage to S. saprophyticus in the presence of D-serine. Furthermore, in vivo coinfection experiments showed a disadvantage for the ΔdsdA mutant during urinary tract infection. Expression analysis of known virulence factors by reverse transcription-quantitative PCR (RT-qPCR) showed that the surface-associated lipase Ssp is upregulated in the presence of D-serine. In addition, we show that S. saprophyticus is able to use D-serine as the sole carbon source, but interestingly, D-serine had a negative effect on growth when glucose was also present. Taken together, D-serine metabolism is associated with virulence in S. saprophyticus, as at least one known virulence factor is upregulated in the presence of D-serine and a ΔdsdA mutant was attenuated in virulence murine model of urinary tract infection.

  2. Adenosine Deaminase Inhibition Prevents Clostridium difficile Toxin A-Induced Enteritis in Mice ▿

    PubMed Central

    de Araújo Junqueira, Ana Flávia Torquato; Dias, Adriana Abalen Martins; Vale, Mariana Lima; Spilborghs, Graziela Machado Gruner Turco; Bossa, Aline Siqueira; Lima, Bruno Bezerra; Carvalho, Alex Fiorini; Guerrant, Richard Littleton; Ribeiro, Ronaldo Albuquerque; Brito, Gerly Anne

    2011-01-01

    Toxin A (TxA) is able to induce most of the classical features of Clostridium difficile-associated disease in animal models. The objective of this study was to determine the effect of an inhibitor of adenosine deaminase, EHNA [erythro-9-(2-hydroxy-3-nonyl)-adenine], on TxA-induced enteritis in C57BL6 mice and on the gene expression of adenosine receptors. EHNA (90 μmol/kg) or phosphate-buffered saline (PBS) was injected intraperitoneally (i.p.) 30 min prior to TxA (50 μg) or PBS injection into the ileal loop. A2A adenosine receptor agonist (ATL313; 5 nM) was injected in the ileal loop immediately before TxA (50 μg) in mice pretreated with EHNA. The animals were euthanized 3 h later. The changes in the tissue were assessed by the evaluation of ileal loop weight/length and secretion volume/length ratios, histological analysis, myeloperoxidase assay (MPO), the local expression of inducible nitric oxide synthase (NOS2), pentraxin 3 (PTX3), NF-κB, tumor necrosis factor alpha (TNF-α), and interleukin-1β (IL-1β) by immunohistochemistry and/or quantitative reverse transcription-PCR (qRT-PCR). The gene expression profiles of A1, A2A, A2B, and A3 adenosine receptors also were evaluated by qRT-PCR. Adenosine deaminase inhibition, by EHNA, reduced tissue injury, neutrophil infiltration, and the levels of proinflammatory cytokines (TNF-α and IL-1β) as well as the expression of NOS2, NF-κB, and PTX3 in the ileum of mice injected with TxA. ATL313 had no additional effect on EHNA action. TxA increased the gene expression of A1 and A2A adenosine receptors. Our findings show that the inhibition of adenosine deaminase by EHNA can prevent Clostridium difficile TxA-induced damage and inflammation possibly through the A2A adenosine receptor, suggesting that the modulation of adenosine/adenosine deaminase represents an important tool in the management of C. difficile-induced disease. PMID:21115723

  3. Maize haplotype with a helitron-amplified cytidine deaminase gene copy

    PubMed Central

    Xu, Jian-Hong; Messing, Joachim

    2006-01-01

    Background Genetic maps are based on recombination of orthologous gene sequences between different strains of the same species. Therefore, it was unexpected to find extensive non-collinearity of genes between different inbred strains of maize. Interestingly, disruption of gene collinearity can be caused among others by a rolling circle-type copy and paste mechanism facilitated by Helitrons. However, understanding the role of this type of gene amplification has been hampered by the lack of finding intact gene sequences within Helitrons. Results By aligning two haplotypes of the z1C1 locus of maize we found a Helitron that contains two genes, one encoding a putative cytidine deaminase and one a hypothetical protein with part of a 40S ribosomal protein. The cytidine deaminase gene, called ZmCDA3, has been copied from the ZmCDA1 gene on maize chromosome 7 about 4.5 million years ago (mya) after maize was formed by whole-genome duplication from two progenitors. Inbred lines contain gene copies of both progenitors, the ZmCDA1 and ZmCDA2 genes. Both genes diverged when the progenitors of maize split and are derived from the same progenitor as the rice OsCDA1 gene. The ZmCDA1 and ZmCDA2 genes are both transcribed in leaf and seed tissue, but transcripts of the paralogous ZmCDA3 gene have not been found yet. Based on their protein structure the maize CDA genes encode a nucleoside deaminase that is found in bacterial systems and is distinct from the mammalian RNA and/or DNA modifying enzymes. Conclusion The conservation of a paralogous gene sequence encoding a cytidine deaminase gene over 4.5 million years suggests that Helitrons could add functional gene sequences to new chromosomal positions and thereby create new haplotypes. However, the function of such paralogous gene copies cannot be essential because they are not present in all maize strains. However, it is interesting to note that maize hybrids can outperform their inbred parents. Therefore, certain haplotypes may

  4. Annexin V-targeted enzyme prodrug therapy using cytosine deaminase in combination with 5-fluorocytosine.

    PubMed

    Van Rite, Brent D; Harrison, Roger G

    2011-08-01

    A fusion protein, consisting of cytosine deaminase (CD) linked to human annexin V, was created for use in an enzyme prodrug therapy targeted to the tumor vasculature and associated cancer cells in the primary tumor and distant metastases. The major finding of this study is that the CD-annexin V fusion protein in combination with the prodrug 5-fluorocytosine has significant cytotoxic activity against endothelial cells and two breast cancer cells lines in vitro that expose phosphatidylserine on their surface. The cytotoxicity experiments verified this novel enzyme prodrug system has the ability to produce therapeutic levels of 5-fluorouracil and thus appears promising.

  5. Synthesis of conformationally locked carbocyclic 1,3-diazepinone nucleosides as inhibitors of cytidine deaminase

    PubMed Central

    Ludek, Olaf R.; Schroeder, Gottfried K.; Wolfenden, Richard; Marquez, Victor E.

    2009-01-01

    We synthesized a series of carbocyclic nucleoside inhibitors of cytidine deaminase (CDA) based on a seven-membered 1,3-diazepin-2-one moiety. In the key step, the seven-membered ring was formed by a ringclosing- metathesis reaction. Therefore, the bis-allylurea moiety had to be protected by benzoylation in order to obtain an orientation suitable for ring closure. To our surprise, the analogue built on a flexible sugar template (4) showed a 100-fold stronger inhibition of CDA than the derivative with the preferred southconformation. PMID:18776552

  6. Threonine deaminase from extremely halophilic bacteria - Cooperative substrate kinetics and salt dependence.

    NASA Technical Reports Server (NTRS)

    Lieberman, M. M.; Lanyi, J. K.

    1972-01-01

    The effect of salt on the activity, stability, and allosteric properties of catabolic threonine deaminase from Halobacterium cutirubrum was studied. The enzyme exhibits sigmoidal kinetics with the substrate, threonine. The Hill slope is 1.55 at pH 10. The enzyme is activated by ADP at low substrate concentrations. In the presence of this effector, sigmoidal kinetics are no longer observed. At pH 10, in the absence of ADP, enzyme activity increases with increasing NaCl concentration from 0 to 4 M.

  7. Long-term expression of human adenosine deaminase in vascular smooth muscle cells of rats: A model for gene therapy

    SciTech Connect

    Lynch, C.M.; Miller, A.D. ); Clowes, M.M.; Osborne, W.R.A.; Clowes, A.W. )

    1992-02-01

    Gene transfer into vascular smooth muscle cells in animals was examined by using recombinant retroviral vectors containing an Escherichia coli {beta}-galactosidase gene or a human adenosine deaminase gene. Direct gene transfer by infusion of virus into rat carotid arteries was not observed. However, gene transfer by infection of smooth muscle cells in culture and seeding of the transduced cells onto arteries that had been denuded of endothelial cells was successful. Potentially therapeutic levels of human adenosine deaminase activity were detected over 6 months of observation, indicating the utility of vascular smooth muscle cells for gene therapy in humans.

  8. Improving Production of Malonyl Coenzyme A-Derived Metabolites by Abolishing Snf1-Dependent Regulation of Acc1

    PubMed Central

    Shi, Shuobo; Chen, Yun; Siewers, Verena

    2014-01-01

    ABSTRACT Acetyl coenzyme A (acetyl-CoA) carboxylase (ACCase) plays a central role in carbon metabolism and has been the site of action for the development of therapeutics or herbicides, as its product, malonyl-CoA, is a precursor for production of fatty acids and other compounds. Control of Acc1 activity in the yeast Saccharomyces cerevisiae occurs mainly at two levels, i.e., regulation of transcription and repression by Snf1 protein kinase at the protein level. Here, we demonstrate a strategy for improving the activity of ACCase in S. cerevisiae by abolishing posttranslational regulation of Acc1 via site-directed mutagenesis. It was found that introduction of two site mutations in Acc1, Ser659 and Ser1157, resulted in an enhanced activity of Acc1 and increased total fatty acid content. As Snf1 regulation of Acc1 is particularly active under glucose-limited conditions, we evaluated the effect of the two site mutations in chemostat cultures. Finally, we showed that our modifications of Acc1 could enhance the supply of malonyl-CoA and therefore successfully increase the production of two industrially important products derived from malonyl-CoA, fatty acid ethyl esters and 3-hydroxypropionic acid. PMID:24803522

  9. The human ACC2 CT-domain C-terminus is required for full functionality and has a novel twist

    SciTech Connect

    Madauss, Kevin P.; Burkhart, William A.; Consler, Thomas G.; Cowan, David J.; Gottschalk, William K.; Miller, Aaron B.; Short, Steven A.; Tran, Thuy B.; Williams, Shawn P.

    2009-05-01

    The use of biophysical assays permitted the identification of a specific human ACC2 carboxyl transferase (CT) domain mutant that binds inhibitors and crystallizes in their presence. This mutant led to determination of the human ACC2 CT domain–CP-640186 complex crystal structure, which revealed differences in the inhibitor conformation from the yeast protein complex that are caused by differing residues in the binding pocket. Inhibition of acetyl-CoA carboxylase (ACC) may prevent lipid-induced insulin resistance and type 2 diabetes, making the enzyme an attractive pharmaceutical target. Although the enzyme is highly conserved amongst animals, only the yeast enzyme structure is available for rational drug design. The use of biophysical assays has permitted the identification of a specific C-terminal truncation of the 826-residue human ACC2 carboxyl transferase (CT) domain that is both functionally competent to bind inhibitors and crystallizes in their presence. This C-terminal truncation led to the determination of the human ACC2 CT domain–CP-640186 complex crystal structure, which revealed distinctions from the yeast-enzyme complex. The human ACC2 CT-domain C-terminus is comprised of three intertwined α-helices that extend outwards from the enzyme on the opposite side to the ligand-binding site. Differences in the observed inhibitor conformation between the yeast and human structures are caused by differing residues in the binding pocket.

  10. Molecular cloning, expression, and stress response of the estrogen-related receptor gene (AccERR) from Apis cerana cerana

    NASA Astrophysics Data System (ADS)

    Zhang, Weixing; Zhu, Ming; Zhang, Ge; Liu, Feng; Wang, Hongfang; Guo, Xingqi; Xu, Baohua

    2016-04-01

    Estrogen-related receptor (ERR), which belongs to the nuclear receptor superfamily, has been implicated in diverse physiological processes involving the estrogen signaling pathway. However, little information is available on ERR in Apis cerana cerana. In this report, we isolated the ERR gene and investigated its involvement in antioxidant defense. Quantitative real-time polymerase chain reaction (qPCR) revealed that the highest mRNA expression occurred in eggs during different developmental stages. The expression levels of AccERR were highest in the muscle, followed by the rectum. The predicted transcription factor binding sites in the promoter of AccERR suggested that AccERR potentially functions in early development and in environmental stress responses. The expression of AccERR was induced by cold (4 °C), heat (42 °C), ultraviolet light (UV), HgCl2, and various types of pesticides (phoxim, deltamethrin, triadimefon, and cyhalothrin). Western blot was used to measure the expression levels of AccERR protein. These data suggested that AccERR might play a vital role in abiotic stress responses.

  11. Metabolic and functional consequences of inhibiting adenosine deaminase during renal ischemia in rats.

    PubMed Central

    Stromski, M E; van Waarde, A; Avison, M J; Thulin, G; Gaudio, K M; Kashgarian, M; Shulman, R G; Siegel, N J

    1988-01-01

    The concentrations of renal ATP have been measured by 31P-nuclear magnetic resonance (NMR) before, during, and after bilateral renal artery occlusion. Using in vivo NMR, the initial postischemic recovery of ATP increased with the magnitude of the residual nucleotide pool at the end of ischemia. ATP levels after 120 min of reflow correlated with functional recovery at 24 h. In the present study the effect of blocking the degradation of ATP during ischemia upon the postischemic restoration of ATP was investigated. Inhibition of adenosine deaminase by 80% with the tight-binding inhibitor 2'-deoxycoformycin led to a 20% increase in the residual adenine nucleotide pool. This increased the ATP initial recovery after 45 min of ischemia from 52% (in controls) to 62% (in the treated animals), as compared to the basal levels. The inhibition also caused an accelerated postischemic restoration of cellular ATP so that at 120 min it was 83% in treated rats vs. 63% in untreated animals. There was a corresponding improvement in the functional recovery from the insult (increase of 33% in inulin clearance 24 h after the injury). Inhibition of adenosine deaminase during ischemia results in a injury similar to that seen after a shorter period of insult. PMID:3263396

  12. The Role of Cytidine Deaminases on Innate Immune Responses against Human Viral Infections

    PubMed Central

    Vieira, Valdimara C.; Soares, Marcelo A.

    2013-01-01

    The APOBEC family of proteins comprises deaminase enzymes that edit DNA and/or RNA sequences. The APOBEC3 subgroup plays an important role on the innate immune system, acting on host defense against exogenous viruses and endogenous retroelements. The role of APOBEC3 proteins in the inhibition of viral infection was firstly described for HIV-1. However, in the past few years many studies have also shown evidence of APOBEC3 action on other viruses associated with human diseases, including HTLV, HCV, HBV, HPV, HSV-1, and EBV. APOBEC3 inhibits these viruses through a series of editing-dependent and independent mechanisms. Many viruses have evolved mechanisms to counteract APOBEC effects, and strategies that enhance APOBEC3 activity constitute a new approach for antiviral drug development. On the other hand, novel evidence that editing by APOBEC3 constitutes a source for viral genetic diversification and evolution has emerged. Furthermore, a possible role in cancer development has been shown for these host enzymes. Therefore, understanding the role of deaminases on the immune response against infectious agents, as well as their role in human disease, has become pivotal. This review summarizes the state-of-the-art knowledge of the impact of APOBEC enzymes on human viruses of distinct families and harboring disparate replication strategies. PMID:23865062

  13. How We Manage Adenosine Deaminase-Deficient Severe Combined Immune Deficiency (ADA SCID).

    PubMed

    Kohn, Donald B; Gaspar, H Bobby

    2017-02-14

    Adenosine deaminase-deficient severe combined immune deficiency (ADA SCID) accounts for 10-15% of cases of human SCID. From what was once a uniformly fatal disease, the prognosis for infants with ADA SCID has improved greatly based on the development of multiple therapeutic options, coupled with more frequent early diagnosis due to implementation of newborn screening for SCID. We review the various treatment approaches for ADA SCID including allogeneic hematopoietic stem cell transplantation (HSCT) from a human leukocyte antigen-matched sibling or family member or from a matched unrelated donor or a haplo-identical donor, autologous HSCT with gene correction of the hematopoietic stem cells (gene therapy-GT), and enzyme replacement therapy (ERT) with polyethylene glycol-conjugated adenosine deaminase. Based on growing evidence of safety and efficacy from GT, we propose a treatment algorithm for patients with ADA SCID that recommends HSCT from a matched family donor, when available, as a first choice, followed by GT as the next option, with allogeneic HSCT from an unrelated or haplo-identical donor or long-term ERT as other options.

  14. A 24-Year Enzyme Replacement Therapy in an Adenosine-deaminase-Deficient Patient.

    PubMed

    Tartibi, Hana M; Hershfield, Michael S; Bahna, Sami L

    2016-01-01

    Severe combined immunodeficiency (SCID) is a fatal childhood disease unless immune reconstitution is performed early in life, with either hematopoietic stem cell transplantation or gene therapy. One of its subtypes is caused by adenosine deaminase (ADA) enzyme deficiency, which leads to the accumulation of toxic metabolites that impair lymphocyte development and function. With the development of polyethylene glycol-conjugated adenosine deaminase (PEG-ADA) enzyme replacement therapy, many ADA-deficient children with SCID who could not receive a hematopoietic stem cell transplantation or gene therapy survived and had longer and healthier lives. We report a 24-year course of treatment in a patient who was diagnosed with ADA deficiency at 4 months of age. The patient was treated with PEG-ADA, which was the only therapy available for him. The patient's plasma ADA level was regularly monitored and the PEG-ADA dose adjusted accordingly. This treatment has resulted in near-normalization of lymphocyte counts, and his clinical course has been associated with only minor to moderate infections. Thus far, he has had no manifestations of autoimmune or lymphoproliferative disorders. This patient is among the longest to be maintained on PEG-ADA enzyme replacement therapy.

  15. Expression of a functional human adenosine deaminase in transgenic tobacco plants.

    PubMed

    Singhabahu, Sanjeewa; George, John; Bringloe, David

    2013-06-01

    An inherited disorder, adenosine deaminase deficiency is a form of severe combined immunodeficiency, which is ultimately caused by an absence of adenosine deaminase (ADA), a key enzyme of the purine salvage pathway. The absence of ADA-activity in sufferers eventually results in a dysfunctional immune system due to the build-up of toxic metabolites. To date, this has been treated with mixed success, using PEG-ADA, made from purified bovine ADA coupled to polyethylene glycol. It is likely, however, that an enzyme replacement therapy protocol based on recombinant human ADA would be a more effective treatment for this disease. Therefore, as a preliminary step to produce biologically active human ADA in transgenic tobacco plants a human ADA cDNA has been inserted into a plant expression vector under the control of the CaMV 35S promoter and both human and TMV 5' UTR control regions. Plant vector expression constructs have been used to transform tobacco plants via Agrobacterium-mediated transformation. Genomic DNA, RNA and protein blot analyses have demonstrated the integration of the cDNA construct into the plant nuclear genome and the expression of recombinant ADA mRNA and protein in transgenic tobacco leaves. Western blot analysis has also revealed that human and recombinant ADA have a similar size of approximately 41 kDa. ADA-specific activities of between 0.001 and 0.003 units per mg total soluble protein were measured in crude extracts isolated from transformed tobacco plant leaves.

  16. Ab Initio ONIOM-Molecular Dynamics (MD) Study on the Deamination Reaction by Cytidine Deaminase

    SciTech Connect

    Matsubara, Toshiaki; Dupuis, Michel; Aida, Misako

    2007-08-23

    We applied the ONIOM-molecular dynamics (MD) method to the hydrolytic deamination of cytidine by cytidine deaminase, which is an essential step of the activation process of the anticancer drug inside the human body. The direct MD simulations were performed for the realistic model of cytidine deaminase calculating the energy and its gradient by the ab initio ONIOM method on the fly. The ONIOM-MD calculations including the thermal motion show that the neighboring amino acid residue is an important factor of the environmental effects and significantly affects not only the geometry and energy of the substrate trapped in the pocket of the active site but also the elementary step of the catalytic reaction. We successfully simulate the second half of the catalytic cycle, which has been considered to involve the rate-determining step, and reveal that the rate-determing step is the release of the NH3 molecule. TM and MA were supported in part by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan. MD was supported by the Division of Chemical Sciences, Office of Basic Energy Sciences, and by the Office of Biological and Environmental Research of the U.S. Department of Energy DOE. Battelle operates Pacific Northwest National Laboratory for DOE.

  17. Diagnostic Value of Adenosine Deaminase and Its Isoforms in Type II Diabetes Mellitus

    PubMed Central

    Larijani, Bagher; Heshmat, Ramin; Ebrahimi-Rad, Mina; Khatami, Shohreh; Valadbeigi, Shirin

    2016-01-01

    Background and Aims. In the present study, we have investigated the activity of adenosine deaminase (ADA) as a diagnostic marker in type 2 (or II) diabetes mellitus (T2DM). Design and Methods. The deaminase activity of ADA1 and ADA2 was determined in serum from 33 patients with type 2 (or II) diabetes mellitus and 35 healthy controls. We also determined the proportion of glycated hemoglobin (HbA1c). Results. Our results showed significant differences between total serum ADA (tADA) and ADA2 activities in the diabetic groups with HbA1c < 8 (%) and HbA1c ≥ 8 (%) with respect to the values in healthy individuals (p < 0.001). ADA2 activity in patients with high HbA1c was found to be much higher than that in patients with low HbA1c (p = 0.0001). In addition, total ADA activity showed a significant correlation with HbA1c (r = 0.6, p < 0.0001). Conclusions. Total serum ADA activity, specially that due to ADA2, could be useful test for the diagnosis of type 2 (or II) diabetes mellitus. PMID:28050278

  18. Ethylene promotes germination of Arabidopsis seed under salinity by decreasing reactive oxygen species: evidence for the involvement of nitric oxide simulated by sodium nitroprusside.

    PubMed

    Lin, Yingchao; Yang, Lei; Paul, Matthew; Zu, Yuangang; Tang, Zhonghua

    2013-12-01

    Both ethylene and nitric oxide (NO) are involved in modulating seed germination in adverse environments. However, the mechanisms by which they interact and affect germination have not been explained. In this study, the relationship between ethylene and NO during germination of Arabidopsis seed under salinity was analysed. Application of exogenous 1-aminocyclopropane-1-carboxylate (ACC, a precursor of ethylene biosynthesis) or sodium nitroprusside (SNP, an NO donor) largely overcame the inhibition of germination induced by salinity. The effects of ACC and SNP were decreased by 2-phenyl-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide (cPTIO), a specific NO scavenger, or by aminoisobutyric acid (AIB), an inhibitor of ethylene biosynthesis, indicating that ethylene and NO interact during germination under salinity. Further, we demonstrated that ACC increased NO production and that SNP greatly induced the expression of the ACS2 gene involved in ethylene synthesis in Arabidopsis seeds germinating under salinity stress, suggesting that each substance influences the production of the other. Application of exogenous ACC increased germination under oxidative stress induced by hydrogen peroxide (H2O2) while SNP had a much smaller effect on wild-type Arabidopsis (Col-0) and no effect on the ethylene insensitive mutant (ein3-1) seeds, respectively. This shows that NO increased germination under salinity indirectly through H2O2 acting via the ethylene pathway. The endogenous concentration of H2O2 was increased by salinity in germinating seeds but was decreased by exogenous ACC, which stimulated germination ultimately. To explain all these results and the regulation of germination of Arabidopsis seed under salinity we propose a model involving ethylene, NO and H2O2 interaction.

  19. Contribution of Ethylene Biosynthesis for Resistance to Blast Fungus Infection in Young Rice Plants1[OA

    PubMed Central

    Iwai, Takayoshi; Miyasaka, Atsushi; Seo, Shigemi; Ohashi, Yuko

    2006-01-01

    The role of ethylene (ET) in resistance to infection with blast fungus (Magnaporthe grisea) in rice (Oryza sativa) is poorly understood. To study it, we quantified ET levels after inoculation, using young rice plants at the four-leaf stage of rice cv Nipponbare (wild type) and its isogenic plant (IL7), which contains the Pi-i resistance gene to blast fungus race 003. Small necrotic lesions by hypersensitive reaction (HR) were formed at 42 to 72 h postinoculation (hpi) in resistant IL7 leaves, and whitish expanding lesions at 96 hpi in susceptible wild-type leaves. Notable was the enhanced ET emission at 48 hpi accompanied by increased 1-aminocyclopropane-1-carboxylic acid (ACC) levels and highly elevated ACC oxidase (ACO) activity in IL7 leaves, whereas only an enhanced ACC increase at 96 hpi in wild-type leaves. Among six ACC synthase (ACS) and seven ACO genes found in the rice genome, OsACS2 was transiently expressed at 48 hpi in IL7 and at 96 hpi in wild type, and OsACO7 was expressed at 48 hpi in IL7. Treatment with an inhibitor for ACS, aminooxyacetic acid, suppressed enhanced ET emission at 48 hpi in IL7, resulting in expanding lesions instead of HR lesions. Exogenously supplied ACC compromised the aminooxyacetic acid-induced breakdown of resistance in IL7, and treatment with 1-methylcyclopropene and silver thiosulfate, inhibitors of ET action, did not suppress resistance. These findings suggest the importance of ET biosynthesis and, consequently, the coproduct, cyanide, for HR-accompanied resistance to blast fungus in young rice plants and the contribution of induced OsACS2 and OsACO7 gene expression to it. PMID:17012402

  20. Capteurs monopodes pour mesures accélérométriques

    NASA Astrophysics Data System (ADS)

    Delaite, R.; Valentin, J.-P.

    1993-08-01

    A new design for accelerometric measurements sensors is described. It uses a plate vibrating in thickness shear mode, maintained by the means of a single holder located at the crystal edge. This mounting does cancel the mechanical and thermal stresses which generally modify the sensor output signal. So the ratio signal/noise of a thickness shear accelerometer is improved and the intrinsic sensitivity is multiplied by a factor 40, by comparison with the sensitivity of a thickness shear plate bonded by the means of two opposite holders. Un nouveau dispositif destiné aux mesures d'accélération est présenté. Il met en œuvre une lame vibrant en cisaillement d'épaisseur, fixée à sa structure de maintien par l'intermédiaire d'une unique liaison. Ce montage permet d'éliminer les contraintes mécaniques et thermiques qui perturbent habituellement le signal de mesure, et qui sont liées soit au montage des éléments du capteur, soit aux variations rapides de température qui interviennent lors de la mise en fonctionnement du capteur. Le rapport signal/bruit d'un accéléromètre à lame vibrant en cisaillement d'épaisseur s'en trouve amélioré et la sensibilité à l'accélération est multipliée par un facteur 40, comparée à celle d'un capteur qui serait constitué d'une lame vibrant en cisaillement d'épaisseur, fixée par deux liaisons diamétralement opposées.

  1. Dissociating contributions of ACC and vmPFC in reward prediction, outcome, and choice.

    PubMed

    Vassena, Eliana; Krebs, Ruth M; Silvetti, Massimo; Fias, Wim; Verguts, Tom

    2014-07-01

    Acting in an uncertain environment requires estimating the probability and the value of potential outcomes. These computations are typically ascribed to various parts of the medial prefrontal cortex (mPFC), but the functional architecture of this region remains debated. The anterior cingulate cortex (ACC) encodes reward prediction and outcome (i.e. win vs lose, Silvetti, Seurinck, & Verguts, 2013. Cortex, 49(6), 1627-35. doi:10.1016/j.cortex.2012.05.008). An outcome-related value signal has also been reported in the ventromedial Prefrontal Cortex (vmPFC, Rangel & Hare, 2010. Current Opinion in Neurobiology, 20(2), 262-70. doi:10.1016/j.conb.2010.03.001). Whether a functional dissociation can be traced in these regions with respect to reward prediction and outcome has been suggested but not rigorously tested. Hence an fMRI study was designed to systematically examine the contribution of ACC and vmPFC to reward prediction and outcome. A striking dissociation was identified, with ACC coding for positive prediction errors and vmPFC responding to outcome, irrespective of probability. Moreover, ACC has been assigned a crucial role in the selection of intentional actions (decision-making) and computing the value associated to these actions (action-based value). Conversely, vmPFC seems to implement stimulus-based value processing (Rudebeck et al., 2008. Journal of Neuroscience, 28(51), 13775-85. doi:10.1523/JNEUROSCI.3541-08.2008; Rushworth, Behrens, Rudebeck, & Walton, 2007. Trends in Cognitive Sciences, 11(4), 168-76. doi:10.1016/j.tics.2007.01.004). Therefore, a decision-making factor (choice vs. no choice condition) was also implemented in the present paradigm to distinguish stimulus-based versus action-based value coding in the mPFC during both decision and outcome phase. We found that vmPFC was more activated during the outcome phase in the no-choice than in the choice condition, potentially confirming the role of this area in stimulus-based (more than action

  2. A recessive mutation in the RUB1-conjugating enzyme, RCE1, reveals a requirement for RUB modification for control of ethylene biosynthesis and proper induction of basic chitinase and PDF1.2 in Arabidopsis.

    PubMed

    Larsen, Paul B; Cancel, Jesse D

    2004-05-01

    By screening etiolated Arabidopsis seedlings for mutants with aberrant ethylene-related phenotypes, we identified a mutant that displays features of the ethylene-mediated triple response even in the absence of ethylene. Further characterization showed that the phenotype observed for the dark-grown seedlings of this mutant is reversible by prevention of ethylene perception and is dependent on a modest increase in ethylene production correlated with an increase in 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase (ACO) activity in the hypocotyl. Molecular characterization of leaves of the mutant revealed severely impaired induction of basic chitinase (chiB) and plant defensin (PDF)1.2 following treatment with jasmonic acid and/or ethylene. Positional cloning of the mutation resulted in identification of a 49-bp deletion in RCE1 (related to ubiquitin 1 (RUB1)-conjugating enzyme), which has been demonstrated to be responsible for covalent attachment of RUB1 to the SCF (Skpl Cdc 53 F-box) ubiquitin ligase complex to modify its activity. Our analyses with rce1-2 demonstrate a previously unknown requirement for RUB1 modification for regulation of ethylene biosynthesis and proper induction of defense-related genes in Arabidopsis.

  3. The effect of plant growth regulators, nitric oxide, nitrate, nitrite and light on the germination of dimorphic seeds of Suaeda salsa under saline conditions.

    PubMed

    Li, Weiqiang; Liu, Xiaojing; Ajmal Khan, M; Yamaguchi, Shinjiro

    2005-06-01

    Suaeda salsa, a leaf succulent shrub in the family Chenopodiaceae, is one of the most important halophytes in China. Suaeda salsa produces dimorphic seeds (soft brown seeds and hard black seeds). Seeds of S. salsa were collected from the coastal salt flats near Huanghua City, China. Experiments were conducted to determine the salinity-alleviating effect of plant growth regulators, nitric oxide, nitrate, nitrite and light on the germination of dimorphic seeds of S. salsa. Brown seeds had a higher germination rate than black seeds in all experiments. Black seeds were more sensitive to salt in the absence of light in comparison to brown seeds. Brown seeds absorbed water more quickly in comparison to black seeds and were found to be more tolerant of salt stress. Our results showed that 1-aminocyclopropane-1-carboxylate (ACC, the immediate precursor of ethylene), nitrite, GA(4) and BA improved seed germination in the presence of salt. However, nitrate, GA(1), GA(3) failed to alleviate salt stress. ABA inhibited seed germination and seedling growth. Possible mechanisms involved in the alleviation of salt stress in S. salsa seeds and the ecological adaptation of the seeds to the environment are discussed.

  4. A Factor Linking Floral Organ Identity and Growth Revealed by Characterization of the Tomato Mutant unfinished flower development (ufd)

    PubMed Central

    Poyatos-Pertíñez, Sandra; Quinet, Muriel; Ortíz-Atienza, Ana; Yuste-Lisbona, Fernando J.; Pons, Clara; Giménez, Estela; Angosto, Trinidad; Granell, Antonio; Capel, Juan; Lozano, Rafael

    2016-01-01

    Floral organogenesis requires coordinated interactions between genes specifying floral organ identity and those regulating growth and size of developing floral organs. With the aim to isolate regulatory genes linking both developmental processes (i.e., floral organ identity and growth) in the tomato model species, a novel mutant altered in the formation of floral organs was further characterized. Under normal growth conditions, floral organ primordia of mutant plants were correctly initiated, however, they were unable to complete their development impeding the formation of mature and fertile flowers. Thus, the growth of floral buds was blocked at an early stage of development; therefore, we named this mutant as unfinished flower development (ufd). Genetic analysis performed in a segregating population of 543 plants showed that the abnormal phenotype was controlled by a single recessive mutation. Global gene expression analysis confirmed that several MADS-box genes regulating floral identity as well as other genes participating in cell division and different hormonal pathways were affected in their expression patterns in ufd mutant plants. Moreover, ufd mutant inflorescences showed higher hormone contents, particularly ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) and strigol compared to wild type. Such results indicate that UFD may have a key function as positive regulator of the development of floral primordia once they have been initiated in the four floral whorls. This function should be performed by affecting the expression of floral organ identity and growth genes, together with hormonal signaling pathways. PMID:27872633

  5. Effects of dynamic controlled atmosphere by respiratory quotient on some quality parameters and volatile profile of 'Royal Gala' apple after long-term storage.

    PubMed

    Both, Vanderlei; Thewes, Fabio Rodrigo; Brackmann, Auri; de Oliveira Anese, Rogerio; de Freitas Ferreira, Daniele; Wagner, Roger

    2017-01-15

    The effects of dynamic controlled atmosphere (DCA) storage based on chlorophyll fluorescence (DCA-CF) and respiratory quotient (DCA-RQ) on the quality and volatile profile of 'Royal Gala' apple were evaluated. DCA storage reduces ACC (1-aminocyclopropane-1-carboxylate) oxidase activity, ethylene production and respiration rate of apples stored for 9months at 1.0°C plus 7days at 20°C, resulting in higher flesh firmness, titratable acidity and lesser physiological disorders, and provided a higher proportion of healthy fruit. Storage in a regular controlled atmosphere gave higher levels of key volatiles (butyl acetate, 2-methylbutyl acetate and hexyl acetate), as compared to fruit stored under DCA-CF, but fruit stored under DCA-RQ 1.5 and RQ 2.0 also showed higher amounts of key volatile compounds, with increment in ethanol and ethyl acetate, but far below the odour threshold. Storage in DCA-CF reduces fruit ester production, especially 2-methylbutyl acetate, which is the most important component of 'Royal Gala' apple flavour.

  6. Ethylene response pathway is essential for ARABIDOPSIS A-FIFTEEN function in floral induction and leaf senescence.

    PubMed

    Chen, Guan-Hong; Chan, Yuan-Li; Liu, Chia-Ping; Wang, Long-Chi

    2012-04-01

    ARABIDOPSIS A-FIFTEEN (AAF) encodes a plastid protein and was originally identified as a SENESCENCE-ASSOCIATED GENE. Previously, we found that overexpression of AAF (AAF-OX) in Arabidopsis led to accumulated reactive oxygen species and promoted leaf senescence induced by oxidative stress, which was suppressed by a null mutant, ein2-5, in ethylene response pathway. Whether AAF function is involved in ethylene biosynthesis and/or the response pathway remained unknown. Here we show that neither overexpression (AAF-OX) nor a null mutant (aaf-KO) of AAF generates a higher level of ethylene than the wild type and display a typical triple-response phenotype in etiolated seedlings treated with 1-aminocyclopropane-1-carboxylic acid (ACC). Nevertheless, ein2-5 suppresses the phenotypes of early flowering and age-dependent leaf senescence in AAF-OX plants. We reveal that a functional ethylene response is essential for AAF function in leaf senescence and floral induction, but AAF is unlikely a regulatory component integral to the ethylene pathway.

  7. Identification and characterization of a novel water-deficit-suppressed gene OsARD encoding an aci-reductone-dioxygenase-like protein in rice.

    PubMed

    Lin, Tao; He, Xiaowei; Yang, Ling; Shou, Huixia; Wu, Ping

    2005-10-24

    The aci-reductone dioxygenase (ARD) family common to bacteria, plants and animals is involved in the methionine salvage pathway. A water-deficit-suppressed gene, OsARD encoding an aci-reductone-dioxygenase-like protein, was identified from rice (Oryza sativa L.). Northern blot and reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed that the OsARD expression is regulated by abiotic stresses and phytohormones. OsARD was mainly expressed in roots under flood conditions. It was suppressed by abiotic stresses including water deficit, high salinity and low temperature, and induced by ethylene and gibberellin acid (GA). Our results showed that the genes for S-adenosylmethionine (SAM) synthase and 1-aminocyclopropane-1-carboxylic acid (ACC) synthase were upregulated in RNA-interference (RNAi) transgenic rice plants with a significant reduction of OsARD expression. Furthermore, the expression of two genes for ethylene signal transduction, ETR2 and EIN3, increased in these RNAi transgenic plants, whereas the expression of ERF3 was suppressed. These results suggest that OsARD may play a role in the metabolism of methionine and ethylene in response to abiotic stresses.

  8. A Factor Linking Floral Organ Identity and Growth Revealed by Characterization of the Tomato Mutant unfinished flower development (ufd).

    PubMed

    Poyatos-Pertíñez, Sandra; Quinet, Muriel; Ortíz-Atienza, Ana; Yuste-Lisbona, Fernando J; Pons, Clara; Giménez, Estela; Angosto, Trinidad; Granell, Antonio; Capel, Juan; Lozano, Rafael

    2016-01-01

    Floral organogenesis requires coordinated interactions between genes specifying floral organ identity and those regulating growth and size of developing floral organs. With the aim to isolate regulatory genes linking both developmental processes (i.e., floral organ identity and growth) in the tomato model species, a novel mutant altered in the formation of floral organs was further characterized. Under normal growth conditions, floral organ primordia of mutant plants were correctly initiated, however, they were unable to complete their development impeding the formation of mature and fertile flowers. Thus, the growth of floral buds was blocked at an early stage of development; therefore, we named this mutant as unfinished flower development (ufd). Genetic analysis performed in a segregating population of 543 plants showed that the abnormal phenotype was controlled by a single recessive mutation. Global gene expression analysis confirmed that several MADS-box genes regulating floral identity as well as other genes participating in cell division and different hormonal pathways were affected in their expression patterns in ufd mutant plants. Moreover, ufd mutant inflorescences showed higher hormone contents, particularly ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) and strigol compared to wild type. Such results indicate that UFD may have a key function as positive regulator of the development of floral primordia once they have been initiated in the four floral whorls. This function should be performed by affecting the expression of floral organ identity and growth genes, together with hormonal signaling pathways.

  9. Higher order Arabidopsis 14-3-3 mutants show 14-3-3 involvement in primary root growth both under control and abiotic stress conditions

    PubMed Central

    van Kleeff, P. J. M.; Jaspert, N.; Li, K. W.; Rauch, S.; Oecking, C.; de Boer, A. H.

    2014-01-01

    Arabidopsis 14-3-3 proteins are a family of conserved proteins that interact with numerous partner proteins in a phospho-specific manner, and can affect the target proteins in a number of ways; e.g. modification of enzymatic activity. We isolated T-DNA insertion lines in six 14-3-3 genes within the non-epsilon group that phylogenetically group in three closely related gene pairs. In total, 6 single, 3 double, 12 triple, and 3 quadruple mutants were generated. The mutants were phenotyped for primary root growth on control plates: single and double mutants were indistinguishable from WT, whereas six triples and all quadruples showed a shorter primary root. In addition, length of the first epidermal cell with a visible root hair bulge (LEH) was used to determine primary root elongation on medium containing mannitol and 1-aminocyclopropane-1-carboxylic acid (ACC). This analysis showed clear differences depending on the stress and 14-3-3 gene combinations. Next to the phenotypic growth analyses, a 14-3-3 pull-down assay on roots treated with and without mannitol showed that mannitol stress strongly affects the 14-3-3 interactome. In conclusion, we show gene specificity and functional redundancy among 14-3-3 proteins in primary root elongation under control and under abiotic stress conditions and changes in the 14-3-3 interactome during the onset of stress adaptation. PMID:25189593

  10. Apple SEPALLATA1/2-like genes control fruit flesh development and ripening.

    PubMed

    Ireland, Hilary S; Yao, Jia-Long; Tomes, Sumathi; Sutherland, Paul W; Nieuwenhuizen, Niels; Gunaseelan, Kularajathevan; Winz, Robert A; David, Karine M; Schaffer, Robert J

    2013-03-01

    Flowering plants utilize different floral structures to develop flesh tissue in fruits. Here we show that suppression of the homeologous SEPALLATA1/2-like genes MADS8 and MADS9 in the fleshy fruit apple (Malus x domestica) leads to sepaloid petals and greatly reduced fruit flesh. Immunolabelling of cell-wall epitopes and differential staining showed that the developing hypanthium (from which the apple flesh develops) of MADS8/9-suppressed apple flowers lacks a tissue layer, and the remaining flesh tissue of fully developed apples has considerably smaller cells. From these observations, it is proposed that MADS8 and MADS9 control the development of discrete zones within the hypanthium tissue, and therefore fruit flesh, and also act as foundations for development of different floral organs. At fruit maturity, the MADS8/9-suppressed apples do not ripen in terms of both developmentally controlled ripening characters, such as starch degradation, and ethylene-modulated ripening traits. Transient assays suggest that, like the RIN gene in tomato, the MADS9 gene acts as a transcriptional activator of the ethylene biosynthesis enzyme, 1-aminocyclopropane-1-carboxylate (ACC) synthase 1. The existence of a single class of genes that regulate both flesh formation and ripening provides an evolutionary tool for controlling two critical aspects of fleshy fruit development.

  11. Metabolic profiling reveals ethylene mediated metabolic changes and a coordinated adaptive mechanism of 'Jonagold' apple to low oxygen stress.

    PubMed

    Bekele, Elias A; Beshir, Wasiye F; Hertog, Maarten L A T M; Nicolai, Bart M; Geeraerd, Annemie H

    2015-11-01

    Apples are predominantly stored in controlled atmosphere (CA) storage to delay ripening and prolong their storage life. Profiling the dynamics of metabolic changes during ripening and CA storage is vital for understanding the governing molecular mechanism. In this study, the dynamics of the primary metabolism of 'Jonagold' apples during ripening in regular air (RA) storage and initiation of CA storage was profiled. 1-Methylcyclopropene (1-MCP) was exploited to block ethylene receptors and to get insight into ethylene mediated metabolic changes during ripening of the fruit and in response to hypoxic stress. Metabolic changes were quantified in glycolysis, the tricarboxylic acid (TCA) cycle, the Yang cycle and synthesis of the main amino acids branching from these metabolic pathways. Partial least square discriminant analysis of the metabolic profiles of 1-MCP treated and control apples revealed a metabolic divergence in ethylene, organic acid, sugar and amino acid metabolism. During RA storage at 18°C, most amino acids were higher in 1-MCP treated apples, whereas 1-aminocyclopropane-1-carboxylic acid (ACC) was higher in the control apples. The initial response of the fruit to CA initiation was accompanied by an increase of alanine, succinate and glutamate, but a decline in aspartate. Furthermore, alanine and succinate accumulated to higher levels in control apples than 1-MCP treated apples. The observed metabolic changes in these interlinked metabolites may indicate a coordinated adaptive strategy to maximize energy production.

  12. A novel role of ethephon in controlling the noxious weed Ipomoea cairica (Linn.) Sweet.

    PubMed

    Sun, Zhong-Yu; Zhang, Tai-Jie; Su, Jin-Quan; Chow, Wah Soon; Liu, Jia-Qin; Chen, Li-Ling; Li, Wei-Hua; Peng, Shao-Lin; Peng, Chang-Lian

    2015-06-18

    Several auxin herbicides, such as 2, 4-D and dicamba, have been used to eradicate an exotic invasive weed Ipomoea cairica in subtropical China, but restraining the re-explosion of this weed is still a challenge. Since ethylene is one of the major intermediate functioning products during the eradication process, we explored the possibility, mechanism and efficiency of using ethephon which can release ethylene to control Ipomoea cairica. The results of the pot experiment showed that 7.2 g /L ethephon could totally kill Ipomoea cairica including the stems and roots. The water culture experiment indicated that ethephon released an abundance of ethylene directly in leaves and caused increases in electrolyte leakage, 1-aminocyclopropane-1-carboxylic acid (ACC), abscisic acid (ABA) and H2O2 and decreases in chlorophyll content and photosynthetic activity, finally leading to the death of Ipomoea cairica. The field experiment showed that the theoretical effective concentration of ethephon for controlling Ipomoea cairica (weed control efficacy, WCE = 98%) was 4.06 g/L and the half inhibitory concentration (I50) was 0.56 g/L. More than 50% of the accompanying species were insensitive to the phytotoxicity of ethephon. Therefore, ethephon is an excellent alternative herbicide for controlling Ipomoea cairica.

  13. ABA- and ethylene-mediated responses in osmotically stressed tomato are regulated by the TSS2 and TOS1 loci.

    PubMed

    Rosado, Abel; Amaya, Iraida; Valpuesta, Victoriano; Cuartero, Jesús; Botella, Miguel A; Borsani, Omar

    2006-01-01

    The study of mutants impaired in the sensitivity or synthesis of abscisic acid (ABA) has become a powerful tool to analyse the interactions occurring between the ABA and ethylene signalling pathways, with potential to change the traditional view of the role of ABA as just being involved in growth inhibition. The tss2 tomato mutant, which is hypersensitive to NaCl and osmotic stress, shows enhanced growth inhibition in the presence of exogenous ABA. The tos1 tomato mutant is also hypersensitive to osmotic stress, but in contrast to tss2, shows decreased sensitivity to ABA. Surprisingly, blocking ethylene signalling suppresses the growth defect of tss2 seedlings on ABA, NaCl, and osmotic stress, but not the osmotic hypersensitivity of tos1. The ethylene production of tss2 seedlings is increased compared with that of control seedlings under osmotic stress. In addition, the tss2 plants are hypersensitive to root growth inhibition by the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). This suggests that, in addition to ABA regulation, TSS2 acts as a negative regulator of endogenous ethylene accumulation. As previously shown in Arabidopsis, it is shown here that extensive cross-talk occurs between the ABA and ethylene signalling pathways in tomato and that the TSS2 and TOS1 loci appear as regulators of this cross-talk.

  14. Role of the gynoecium in natural senescence of carnation (Dianthus caryophyllus L.) flowers.

    PubMed

    Shibuya, K; Yoshioka, T; Hashiba, T; Satoh, S

    2000-12-01

    Although the role of the gynoecium in natural senescence of the carnation flower has long been suggested, it has remained a matter of dispute because petal senescence in the cut carnation flower was not delayed by the removal of gynoecium. In this study, the gynoecium was snapped off by hand, in contrast to previous investigations where removal was achieved by forceps or scissors. The removal of the gynoecium by hand prevented the onset of ethylene production and prolonged the vase life of the flower, demonstrating a decisive role of the gynoecium in controlling natural senescence of the carnation flower. Abscisic acid (ABA) and indole-3-acetic acid (IAA), which induced ethylene production and accelerated petal senescence in carnation flowers, did not stimulate ethylene production in the flowers with gynoecia removed (-Gyn flowers). Application of 1-aminocyclopropane-1-carboxylate (ACC), the ethylene precursor, induced substantial ethylene production and petal wilting in the flowers with gynoecia left intact, but was less effective at stimulating ethylene production in the -Gyn flowers and negligible petal in-rolling was observed. Exogenous ethylene induced autocatalytic production of the gas and petal wilting in the -Gyn flowers. These results indicated that ethylene generated in the gynoecium triggers the onset of ethylene production in the petals of carnation during natural senescence.

  15. The gravitropic setpoint angle of dark-grown rye seedlings and the role of ethylene.

    PubMed

    Edelmann, Hans G; Gudi, Gennadi; Kühnemann, Frank

    2002-07-01

    The orientation growth of coleoptiles of dark-grown seedlings of rye (Secale cereale L. cv. Marder II), when grown under various conditions, was analysed with respect to the gravivector ('gravitropic setpoint angle', GSA). Coleoptiles growing through moist vermiculite attain and maintain a GSA with an average of about 180 degrees, i.e. a vertical orientation. Seedlings growing uncovered either on the surface of vermiculite or positionally fixed on filter paper attain and maintain a GSA of 140-150 degrees (i.e. deviating from the vertical by an average of 30-40 degrees ). Changing the position of the embryo relative to the horizontally fixed seed kernel or of the angle of the seed with respect to gravity during germination (+/-40 degrees relative to the horizontal) had no significant effect on the subsequent GSA of both covered and uncovered seedlings. The GSA of uncovered coleoptiles could be restored close to 180 degrees by treatment of the seedlings with ethylene, either applied via ethephon or 1-aminocyclopropane-1-carboxylic acid (ACC) as well as by fruit-released ethylene. The results are discussed with respect to the mechanism of the regulation of gravitropic growth of grass seedlings.

  16. New chiral didehydroamino acid derivatives from a cyclic glycine template with 3,6-dihydro-2H-1,4-oxazin-2-one structure: applications to the asymmetric synthesis of nonproteinogenic alpha-amino acids.

    PubMed

    Chinchilla, R; Falvello, L R; Galindo, N; Nájera, C

    2000-05-19

    New chiral (Z)-alpha,beta-didehydroamino acid (DDAA) derivatives with 3,5-dihydro-2H-1,4-oxazin-2-one structure 11a-f have been stereoselectively prepared after condensation of chiral glycine equivalent 7 with aldehydes in the presence of K(2)CO(3) under mild solid-liquid phase-transfer catalysis reaction conditions. These new systems have been used in diastereoselective cyclopropanation reactions using Corey's ylide for the asymmetric synthesis of 1-aminocyclopropane-1-carboxylic acids (ACCs) such as allo-corononamic and allo-norcoronamic acids. The hydrogenation reaction of these systems at ambient pressure in the presence of formaldehyde affords saturated oxazinones and N-methylated oxazinones which have been transformed into the N-methyl-alpha-amino acids (N-MAAs) (S)-2-(methylamino)butanoic acid and (S)-N-methylleucine. In addition, the parent alpha, beta-didehydroalanine derivative 11g has been prepared by a direct aminomethylation-elimination sequence from 7 and Eschenmoser's salt and has been used in Diels-Alder cycloaddition with endo selectivity for the synthesis of the enantiomerically pure bicyclic alpha-amino acids (-)-2-aminobicyclo[2.2.1]heptane-2-carboxylic and (-)-2-aminobicyclo[2.2.2]octane-2-carboxylic acids.

  17. A novel role of ethephon in controlling the noxious weed Ipomoea cairica (Linn.) Sweet

    PubMed Central

    Sun, Zhong-Yu; Zhang, Tai-Jie; Su, Jin-Quan; Soon Chow, Wah; Liu, Jia-Qin; Chen, Li-Ling; Li, Wei-Hua; Peng, Shao-Lin; Peng, Chang-Lian

    2015-01-01

    Several auxin herbicides, such as 2, 4-D and dicamba, have been used to eradicate an exotic invasive weed Ipomoea cairica in subtropical China, but restraining the re-explosion of this weed is still a challenge. Since ethylene is one of the major intermediate functioning products during the eradication process, we explored the possibility, mechanism and efficiency of using ethephon which can release ethylene to control Ipomoea cairica. The results of the pot experiment showed that 7.2 g /L ethephon could totally kill Ipomoea cairica including the stems and roots. The water culture experiment indicated that ethephon released an abundance of ethylene directly in leaves and caused increases in electrolyte leakage, 1-aminocyclopropane-1-carboxylic acid (ACC), abscisic acid (ABA) and H2O2 and decreases in chlorophyll content and photosynthetic activity, finally leading to the death of Ipomoea cairica. The field experiment showed that the theoretical effective concentration of ethephon for controlling Ipomoea cairica (weed control efficacy, WCE = 98%) was 4.06 g/L and the half inhibitory concentration (I50) was 0.56 g/L. More than 50% of the accompanying species were insensitive to the phytotoxicity of ethephon. Therefore, ethephon is an excellent alternative herbicide for controlling Ipomoea cairica. PMID:26087386

  18. Participation of Ethylene in Two Modes of Gravistimulation of Shoots

    NASA Technical Reports Server (NTRS)

    Harrison, M.

    1985-01-01

    In order to elucidate the role of ehtylene in gravitropism, detailed time courses for ethylene production in horizontal and upright plants were measured. Tomato and pea were chosen as examples of plants which exhibit different patterns of gravitropic curvature. Tomato seedlings were placed in gas-tight lucite boxes from which air was sampled and analyzed for ethylene. During the first 2 min interval after one set of plants was turned horizontal ethylene production was double the baseline. Similarly, plants rotated 3 rpm about a vertical axis transiently doubled ethylene production when the axis was shifted 90 deg. In order to clarify the role of this 2-min burst, the effect of exogenous ethylene was studied. In peas, epicotyls were excised, equilibrated until wound ethylene had subsided to a low stable level, and ethylene production was measured in vertical and horizontal segments. As for tomatoes, excised pea epicotyls increased their rate of ethylene production during the first 2 min of gravistimulation. Also, very low concentrations of exogenous ethylene slightly enhance curvature. On the other hand, higher levels of ethylene and 1-aminocyclopropane-1-carboxylic acid (ACC) inhibit overall curvature.

  19. The retroviral hypermutation specificity of APOBEC3F and APOBEC3G is governed by the C-terminal DNA cytosine deaminase domain.

    PubMed

    Haché, Guylaine; Liddament, Mark T; Harris, Reuben S

    2005-03-25

    The human proteins APOBEC3F and APOBEC3G restrict retroviral infection by deaminating cytosine residues in the first cDNA strand of a replicating virus. These proteins have two putative deaminase domains, and it is unclear whether one or both catalyze deamination, unlike their homologs, AID and APOBEC1, which are well characterized single domain deaminases. Here, we show that only the C-terminal cytosine deaminase domain of APOBEC3F and -3G governs retroviral hypermutation. A chimeric protein with the N-terminal cytosine deaminase domain from APOBEC3G and the C-terminal cytosine deaminase domain from APOBEC3F elicited a dinucleotide hypermutation preference nearly indistinguishable from that of APOBEC3F. This 5'-TC-->TT mutational specificity was confirmed in a heterologous Escherichia coli-based mutation assay, in which the 5'-CC-->CT dinucleotide hypermutation preference of APOBEC3G also mapped to the C-terminal deaminase domain. An N-terminal APOBEC3G deletion mutant displayed a preference indistinguishable from that of the full-length protein, and replacing the C-terminal deaminase domain of APOBEC3F with AID resulted in an AID-like mutational signature. Together, these data indicate that only the C-terminal domain of APOBEC3F and -3G dictates the retroviral minus strand 5'-TC and 5'-CC dinucleotide hypermutation preferences, respectively, leaving the N-terminal domain to perform other aspects of retroviral restriction.

  20. Cytosine deaminase as a negative selection marker for gene disruption and replacement in the genus Streptomyces and other actinobacteria.

    PubMed

    Dubeau, Marie-Pierre; Ghinet, Mariana Gabriela; Jacques, Pierre-Etienne; Clermont, Nancy; Beaulieu, Carole; Brzezinski, Ryszard

    2009-02-01

    We developed a novel negative selection system for actinobacteria based on cytosine deaminase (CodA). We constructed vectors that include a synthetic gene encoding the CodA protein from Escherichia coli optimized for expression in Streptomyces species. Gene disruption and the introduction of an unmarked in-frame deletion were successfully achieved with these vectors.

  1. Isolation and properties of AMP deaminase from jumbo squid (Dosidicus gigas) mantle muscle from the Gulf of California, Mexico.

    PubMed

    Marquez-Rios, E; Pacheco-Aguilar, R; Castillo-Yañez, F J; Figueroa-Soto, C G; Ezquerra-Brauer, J M; Gollas-Galvan, T

    2008-09-01

    Adenosine monophosphate (AMP) deaminase was purified from jumbo squid mantle muscle by chromatography in cellulose phosphate, Q-Fast and 5'-AMP sepharose. Specific activity of 2.5U/mg protein, 4.5% recovery and 133.68 purification fold were obtained at the end of the experiment. SDS-PAGE showed a single band with 87kDa molecular mass, native PAGE proved a band of 178kDa, whereas gel filtration detected a 180kDa protein, suggesting the homodimeric nature of this enzyme, in which subunits are not linked by covalent forces. Isoelectric focusing of this enzyme showed a pI of 5.76, which agrees with pI values of AMP deaminase from other invertebrate organisms. AMP deaminase presented a kinetic sigmoidal plot with Vmax of 1.16μM/min/mg, Km of 13mM, Kcat of 3.48μM.s(-1) and a Kcat/Km of 267 (mol/L)(-1).s(-1). The apparent relative low catalytic activity of jumbo squid muscle AMP deaminase in the absence of positive effectors is similar to that reported for homologous enzymes in other invertebrate organisms.

  2. A curving ACC system with coordination control of longitudinal car-following and lateral stability

    NASA Astrophysics Data System (ADS)

    Zhang, Dezhao; Li, Keqiang; Wang, Jianqiang

    2012-07-01

    The paper presents a curving adaptive cruise control (ACC) system that is coordinated with a direct yaw-moment control (DYC) system and gives consideration to both longitudinal car-following capability and lateral stability on curved roads. A model including vehicle longitudinal and lateral dynamics is built first, which is as discrete as the predictive model of the system controller. Then, a cost function is determined to reflect the contradictions between vehicle longitudinal and lateral dynamics. Meanwhile, some I/O constraints are formulated with a driver permissible longitudinal car-following range and the road adhesion condition. After that, desired longitudinal acceleration and desired yaw moment are obtained by a linear matrix inequality based robust constrained state feedback method. Finally, driver-in-the-loop tests on a driving simulator are conducted and the results show that the developed control system provides significant benefits in weakening the impact of DYC on ACC longitudinal car-following capability while also improving lateral stability.

  3. Preparation of ethylene gas and comparison of ethylene responses induced by ethylene, ACC, and ethephon.

    PubMed

    Zhang, Wei; Wen, Chi-Kuang

    2010-01-01

    Ethylene is a gaseous plant hormone used in many physiological studies examining its role in plant growth and development. However, ethylene gas may not be conveniently available to many laboratories for occasional use, and therefore several chemicals can be used as replacements. Here we report that the kinetics of the ethylene response induced by ethylene and two widely-used ethylene replacements are different. ACC failed to efficiently replace prolonged ethylene treatments, while the decomposition products of ethephon may cause non-specific responses and the efficiency of ethephon conversion to ethylene was relatively low. A cost-effective method to prepare ethylene gas was developed. Analyzed by gas chromatography, the chemically produced ethylene exhibited an identical chromatogram to that from the commercial source. Our synthetic ethylene gave the same dose-response curve in Arabidopsis as gaseous ethylene. Our study shows that the use of the ethylene gas is essential to experiments that are sensitive to treatment duration and dosage. When ACC and ethephon are used as replacements, caution should be taken in the experimental design. For laboratories that do not have an ethylene tank, ethylene gas can be easily prepared by a chemical approach without further purification.

  4. Efficient retrovirus-mediated transfer and expression of a human adenosine deaminase gene in diploid skin fibroblasts from an adenosine deaminase-deficient human

    SciTech Connect

    Palmer, T.D.; Hock, R.A.; Osborne, W.R.A.; Miller, A.D.

    1987-02-01

    Skin fibroblasts might be considered suitable recipients for therapeutic genes to cure several human genetic diseases; however, these cells are resistant to gene transfer by most methods. The authors studied the ability of retroviral vectors to transfer genes into normal human diploid skin fibroblasts. Retroviruses carrying genes for neomycin or hygromycin B resistance conferred drug resistance to greater than 50% of the human fibroblasts after a single exposure to virus-containing medium. This represents at least a 500-fold increase in efficiency over other methods. Transfer was achieved in the absence of helper virus by using amphotropic retrovirus-packaging cells. A retrovirus vector containing a human adenosine deaminase (ADA) cDNA was constructed and used to infect ADA/sup -/ fibroblasts from a patient with ADA deficiency. The infected cells produced 12-fold more ADA enzyme than fibroblasts from normal individuals and were able to rapidly metabolize exogenous deoxyadenosine and adenosine, metabolites that accumulate in plasma in ADA-deficient patients and are responsible for the severe combined immunodeficiency in these patients. These experiments indicate the potential of retrovirus-mediated gene transfer into human fibroblasts for gene therapy.

  5. Effect of alginate microencapsulation on the catalytic efficiency and in vitro enzyme-prodrug therapeutic efficacy of cytosine deaminase and of recombinant E. coli expressing cytosine deaminase.

    PubMed

    Funaro, Michael G; Nemani, Krishnamurthy V; Chen, Zhihang; Bhujwalla, Zaver M; Griswold, Karl E; Gimi, Barjor

    2016-02-01

    Cytosine deaminase (CD) catalyses the enzymatic conversion of the non-toxic prodrug 5-fluorocytosine (5-FC) to the potent chemotherapeutic form, 5-fluorouracil (5-FU). Intratumoral delivery of CD localises chemotherapy dose while reducing systemic toxicity. Encapsulation in biocompatible microcapsules immunoisolates CD and protects it from degradation. We report on the effect of alginate encapsulation on the catalytic and functional activity of isolated CD and recombinant E. coli engineered to express CD (E. coli(CD)). Alginate microcapsules containing either CD or Escherichia coli(CD) were prepared using ionotropic gelation. Conversion of 5-FC to 5-FU was quantitated in unencapsulated and encapsulated CD/E. coli(CD) using spectrophotometry, with a slower rate of conversion observed following encapsulation. Both encapsulated CD/5-FC and E. coli(CD)/5-FC resulted in cell kill and reduced proliferation of 9 L rat glioma cells, which was comparable to direct 5-FU treatment. Our results show that encapsulation preserves the therapeutic potential of CD and E. coli(CD) is equally effective for enzyme-prodrug therapy.

  6. 5th International ACC Symposium: Classification of Adrenocortical Cancers from Pathology to Integrated Genomics: Real Advances or Lost in Translation?

    PubMed

    de Krijger, Ronald E; Bertherat, Jérôme

    2016-02-01

    For the clinician, despite its rarity, adrenocortical cancer is a heterogeneous tumor both in term of steroid excess and tumor evolution. For patient management, it is crucial to have an accurate vision of this heterogeneity, in order to use a correct tumor classification. Pathology is the best way to classify operated adrenocortical tumors: to recognize their adrenocortical nature and to differentiate benign from malignant tumors. Among malignant tumors pathology also aims at prognosis assessment. Although progress has being made for prognosis assessment, there is still a need for improvement. Recent studies have established the value of Ki67 for adrenocortical cancer (ACC) prognostication, aiming also at standardization to reduce variability. The use of genomics to study adrenocortical tumors gives a very new insight in their pathogenesis and molecular classification. Genomics studies of ACC give now a clear description of the mRNA (transcriptome) and miRNA expression profile, as well as chromosomal and methylation alterations. Exome sequencing also established firmly the list of the main ACC driver genes. Interestingly, genomics study of ACC also revealed subtypes of malignant tumors with different pattern of molecular alterations, associated with different outcome. This leads to a new vision of adrenocortical tumors classification based on molecular analysis. Interestingly, these molecular classifications meet also the results of pathological analysis. This opens new perspectives on the development and use of various molecular tools to classify, along with pathological analysis, ACC, and guides patient management at the area of precision medicine.

  7. Monitoring apoptosis of TK-GFP-expressing ACC-M cells induced by ACV using FRET technique

    NASA Astrophysics Data System (ADS)

    Xiong, Tao; Zhang, Zhihong; Lin, Juqiang; Yang, Jie; Zeng, Shaoqun; Luo, Qingming

    2006-05-01

    Apoptosis is an evolutionary conserved cellular process that plays an important role during development, but it is also involved in tissue homeostasis and in many diseases. To study the characteristics of suicide gene system of the herpes simplex virus thymidine kinase (HSV-tk) gene in tumor cells and explore the apoptosis phenomena in this system and its effect on the human adenoid cystic carcinoma line ACC-M cell, we detected apoptosis of CD3- (ECFP-CRS-DsRed) and TK-GFP-expressing ACC-M (ACC-M-TK-GFP-CD3) cells induced by acyclovir (ACV) using fluorescence resonance energy transfer (FRET) technique. CD3 is a FRET-based indicator for activity of caspase-3, which is composed of an enhanced cyan fluorescent protein, a caspase-3 sensitive linker, and a red fluorescent protein from Discosoma with efficient maturation property. FRET from ECFP to DsRed could be detected in normal ACC-M-TK-GFP-CD3 cells, and the FRET efficient was remarkably decreased and then disappeared during the cells apoptosis induced by ACV. It was due to the activated caspase-3 cleaved the CD3 fusion protein. In this study, the results suggested that the ACV-induced apoptosis of ACC-M-TK-GFP-CD3 cells was through caspase-3 pathway.

  8. Monitoring apoptosis of TK-GFP-expressing ACC-M cells induced by ACV using FRET technique

    NASA Astrophysics Data System (ADS)

    Xiong, Tao; Zhang, Zhihong; Lin, Juqiang; Yang, Jie; Zeng, Shaoqun; Luo, Qingming

    2006-09-01

    Apoptosis is an evolutionary conserved cellular process that plays an important role during development, but it is also involved in tissue homeostasis and in many diseases. To study the characteristics of suicide gene system of the herpes simplex virus thymidine kinase (HSV-tk) gene in tumor cells and explore the apoptosis phenomena in this system and its effect on the human adenoid cystic carcinoma line ACC-M cell, we detected apoptosis of CD3- (ECFP-CRS-DsRed) and TK-GFP-expressing ACC-M (ACC-M-TK-GFP-CD3) cells induced by acyclovir (ACV) using fluorescence resonance energy transfer (FRET) technique. CD3 is a FRET-based indicator for activity of caspase-3, which is composed of an enhanced cyan fluorescent protein, a caspase-3 sensitive linker, and a red fluorescent protein from Discosoma with efficient maturation property. FRET from ECFP to DsRed could be detected in normal ACC-M-TK-GFP-CD3 cells, and the FRET efficient was remarkably decreased and then disappeared during the cells apoptosis induced by ACV. It was due to the activated caspase-3 cleaved the CD3 fusion protein. In this study, the results suggested that the AVC-induced apoptosis of ACC-M-TK-GFP-CD3 cells was through caspase-3 pathway.

  9. Thermoperiodic control of hypocotyl elongation depends on auxin-induced ethylene signaling that controls downstream PHYTOCHROME INTERACTING FACTOR3 activity.

    PubMed

    Bours, Ralph; Kohlen, Wouter; Bouwmeester, Harro J; van der Krol, Alexander

    2015-02-01

    We show that antiphase light-temperature cycles (negative day-night temperature difference [-DIF]) inhibit hypocotyl growth in Arabidopsis (Arabidopsis thaliana). This is caused by reduced cell elongation during the cold photoperiod. Cell elongation in the basal part of the hypocotyl under -DIF was restored by both 1-aminocyclopropane-1-carboxylic acid (ACC; ethylene precursor) and auxin, indicating limited auxin and ethylene signaling under -DIF. Both auxin biosynthesis and auxin signaling were reduced during -DIF. In addition, expression of several ACC Synthase was reduced under -DIF but could be restored by auxin application. In contrast, the reduced hypocotyl elongation of ethylene biosynthesis and signaling mutants could not be complemented by auxin, indicating that auxin functions upstream of ethylene. The PHYTOCHROME INTERACTING FACTORS (PIFs) PIF3, PIF4, and PIF5 were previously shown to be important regulators of hypocotyl elongation. We now show that, in contrast to pif4 and pif5 mutants, the reduced hypocotyl length in pif3 cannot be rescued by either ACC or auxin. In line with this, treatment with ethylene or auxin inhibitors reduced hypocotyl elongation in PIF4 overexpressor (PIF4ox) and PIF5ox but not PIF3ox plants. PIF3 promoter activity was strongly reduced under -DIF but could be restored by auxin application in an ACC Synthase-dependent manner. Combined, these results show that PIF3 regulates hypocotyl length downstream, whereas PIF4 and PIF5 regulate hypocotyl length upstream of an auxin and ethylene cascade. We show that, under -DIF, lower auxin biosynthesis activity limits the signaling in this pathway, resulting in low activity of PIF3 and short hypocotyls.

  10. Development of gene therapy: potential in severe combined immunodeficiency due to adenosine deaminase deficiency

    PubMed Central

    Montiel-Equihua, Claudia A; Thrasher, Adrian J; Gaspar, H Bobby

    2010-01-01

    The history of stem cell gene therapy is strongly linked to the development of gene therapy for severe combined immunodeficiencies (SCID) and especially adenosine deaminase (ADA)-deficient SCID. Here we discuss the developments achieved in over two decades of clinical and laboratory research that led to the establishment of a protocol for the autologous transplant of retroviral vector-mediated gene-modified hematopoietic stem cells, which has proved to be both successful and, to date, safe. Patients in trials in three different countries have shown long-term immunological and metabolic correction. Nevertheless, improvements to the safety profile of viral vectors are underway and will undoubtedly reinforce the position of stem cell gene therapy as a treatment option for ADA-SCID. PMID:24198507

  11. Development of gene therapy: potential in severe combined immunodeficiency due to adenosine deaminase deficiency.

    PubMed

    Montiel-Equihua, Claudia A; Thrasher, Adrian J; Gaspar, H Bobby

    2009-12-22

    The history of stem cell gene therapy is strongly linked to the development of gene therapy for severe combined immunodeficiencies (SCID) and especially adenosine deaminase (ADA)-deficient SCID. Here we discuss the developments achieved in over two decades of clinical and laboratory research that led to the establishment of a protocol for the autologous transplant of retroviral vector-mediated gene-modified hematopoietic stem cells, which has proved to be both successful and, to date, safe. Patients in trials in three different countries have shown long-term immunological and metabolic correction. Nevertheless, improvements to the safety profile of viral vectors are underway and will undoubtedly reinforce the position of stem cell gene therapy as a treatment option for ADA-SCID.

  12. Myoadenylate deaminase deficiency. Functional and metabolic abnormalities associated with disruption of the purine nucleotide cycle.

    PubMed Central

    Sabina, R L; Swain, J L; Olanow, C W; Bradley, W G; Fishbein, W N; DiMauro, S; Holmes, E W

    1984-01-01

    To assess the role of the purine nucleotide cycle in human skeletal muscle function, we evaluated 10 patients with AMP deaminase deficiency (myoadenylate deaminase deficiency; MDD). 4 MDD and 19 non-MDD controls participated in an exercise protocol. The latter group was composed of a patient cohort (n = 8) exhibiting a constellation of symptoms similar to those of the MDD patients, i.e., postexertional aches, cramps, and pains; as well as a cohort of normal, unconditioned volunteers (n = 11). The individuals with MDD fatigued after performing only 28% as much work as their non-MDD counterparts. Muscle biopsies were obtained from the four MDD patients and the eight non-MDD patients at rest and following exercise to the point of fatigue. Creatine phosphate content fell to a comparable extent in the MDD (69%) and non-MDD (52%) patients at the onset of fatigue. Following exercise the 34% decrease in ATP content of muscle from the non-MDD subjects was significantly greater than the 6% decrease in ATP noted in muscle from the MDD patients (P = 0.048). Only one of four MDD patients had a measurable drop in ATP compared with seven of eight non-MDD patients. At end-exercise the muscle content of inosine 5'-monophosphate (IMP), a product of AMP deaminase, was 13-fold greater in the non-MDD patients than that observed in the MDD group (P = 0.008). Adenosine content of muscle from the MDD patients increased 16-fold following exercise, while there was only a twofold increase in adenosine content of muscle from the non-MDD patients (P = 0.028). Those non-MDD patients in whom the decrease in ATP content following exercise was measurable exhibited a stoichiometric increase in IMP, and total purine content of the muscle did not change significantly. The one MDD patient in whom the decrease in ATP was measurable, did not exhibit a stoichiometric increase in IMP. Although the adenosine content increased 13-fold in this patient, only 48% of the ATP catabolized could be accounted for

  13. Hematopoietic stem cell gene therapy for adenosine deaminase deficient-SCID.

    PubMed

    Aiuti, Alessandro; Brigida, Immacolata; Ferrua, Francesca; Cappelli, Barbara; Chiesa, Robert; Marktel, Sarah; Roncarolo, Maria-Grazia

    2009-01-01

    Gene therapy is a highly attractive strategy for many types of inherited disorders of the immune system. Adenosine deaminase (ADA) deficient-severe combined immunodeficiency (SCID) has been the target of several clinical trials based on the use of hematopoietic stem/progenitor cells engineered with retroviral vectors. The introduction of a low intensity conditioning regimen has been a crucial factor in achieving stable engrafment of hematopoietic stem cells and therapeutic levels of ADA-expressing cells. Recent studies have demonstrated that gene therapy for ADA-SCID has favorable safety profile and is effective in restoring normal purine metabolism and immune functions. Stem cell gene therapy combined with appropriate conditioning regimens might be extended to other genetic disorders of the hematopoietic system.

  14. Reaction mechanism of zinc-dependent cytosine deaminase from Escherichia coli: a quantum-chemical study.

    PubMed

    Manta, Bianca; Raushel, Frank M; Himo, Fahmi

    2014-05-29

    The reaction mechanism of cytosine deaminase from Escherichia coli is studied using density functional theory. This zinc-dependent enzyme catalyzes the deamination of cytosine to form uracil and ammonia. The calculations give a detailed description of the catalytic mechanism and establish the role of important active-site residues. It is shown that Glu217 is essential for the initial deprotonation of the metal-bound water nucleophile and the subsequent protonation of the substrate. It is also demonstrated that His246 is unlikely to function as a proton shuttle in the nucleophile activation step, as previously proposed. The steps that follow are nucleophilic attack by the metal-bound hydroxide, protonation of the leaving group assisted by Asp313, and C-N bond cleavage. The calculated overall barrier is in good agreement with the experimental findings. Finally, the calculations reproduce the experimentally determined inverse solvent deuterium isotope effect, which further corroborates the suggested reaction mechanism.

  15. Mycoplasma hyorhinis-encoded cytidine deaminase efficiently inactivates cytosine-based anticancer drugs.

    PubMed

    Vande Voorde, Johan; Vervaeke, Peter; Liekens, Sandra; Balzarini, Jan

    2015-01-01

    Mycoplasmas may colonize tumor tissue in patients. The cytostatic activity of gemcitabine was dramatically decreased in Mycoplasma hyorhinis-infected tumor cell cultures compared with non-infected tumor cell cultures. This mycoplasma-driven drug deamination could be prevented by exogenous administration of the cytidine deaminase (CDA) inhibitor tetrahydrouridine, but also by the natural nucleosides or by a purine nucleoside phosphorylase inhibitor. The M. hyorhinis-encoded CDAHyor gene was cloned, expressed as a recombinant protein and purified. CDAHyor was found to be more catalytically active than its human equivalent and efficiently deaminates (inactivates) cytosine-based anticancer drugs. CDAHyor expression at the tumor site may result in selective drug inactivation and suboptimal therapeutic efficiency.

  16. Markerless Gene Deletion with Cytosine Deaminase in Thermus thermophilus Strain HB27.

    PubMed

    Wang, Lei; Hoffmann, Jana; Watzlawick, Hildegard; Altenbuchner, Josef

    2015-12-11

    We developed a counterselectable deletion system for Thermus thermophilus HB27 based on cytosine deaminase (encoded by codA) from Thermaerobacter marianensis DSM 12885 and the sensitivity of T. thermophilus HB27 to the antimetabolite 5-fluorocytosine (5-FC). The deletion vector comprises the pUC18 origin of replication, a thermostable kanamycin resistance marker functional in T. thermophilus HB27, and codA under the control of a constitutive putative trehalose promoter from T. thermophilus HB27. The functionality of the system was demonstrated by deletion of the bglT gene, encoding a β-glycosidase, and three carotenoid biosynthesis genes, CYP175A1, crtY, and crtI, from the genome of T. thermophilus HB27.

  17. Stabilization of Aspergillus parasiticus cytosine deaminase by immobilization on calcium alginate beads improved enzyme operational stability.

    PubMed

    Zanna, H; Nok, A J; Ibrahim, S; Inuwa, H M

    2013-12-01

    Cytosine deaminase (CD) from Aspergillus parasiticus, which has half-life of 1.10 h at 37°C, was stabilized by immobilization on calcium alginate beads. The immobilized CD had pH and temperature optimum of 5 and 50°C respectively. The immobilized enzyme also stoichiometrically deaminated Cytosine and 5-fluorocytosine (5-FC) with the apparent K(M) values of 0.60 mM and 0.65 mM respectively, displaying activation energy of 10.72 KJ/mol. The immobilization of native CD on calcium alginate beads gave the highest yield of apparent enzymatic activity of 51.60% of the original activity and the enzymatic activity was lost exponentially at 37°C over 12 h with a half-life of 5.80 h. Hence, the operational stability of native CD can be improved by immobilization on calcium alginate beads.

  18. Genetic immunotherapy for hepatocellular carcinoma by endothelial progenitor cells armed with cytosine deaminase.

    PubMed

    Chen, Rong; Yu, Hui; An, Yan-Li; Yu-Jia, Zhen; Teng, Gao-Jun

    2014-02-01

    Endothelial progenitor cells (EPCs) serve as cellular vehicles for targeting cancer cells and are a powerful tool for delivery of therapeutic genes. Cytosine deaminase (CD), a kind of frequent suicide gene which can kill carcinoma cells by converting a non-poisonous pro-drug 5-flucytosine (5-FC) into a poisonous cytotoxic 5-fluorouracil (5-FU). We combined super-paramagnetic iron oxide (SPIO) nanoparticles labeled EPCs with CD gene to treat grafted liver carcinomas and tracked them with 7.0 T Magnetic resonance imaging (MRI). Results showed that the therapeutic EPCs loaded with CD plus 5-Fc provided stronger carcinoma growth suppression compared with treatment using CD alone. The CD/5-Fc significantly inhibited the growth of endothelial cells and induced carcinoma cells apoptosis. These results indicate that EPCs transfected with anti-carcinoma genes can be used in carcinoma therapy as a novel therapeutic modality.

  19. Does adenosine deaminase activity play a role in the early diagnosis of ectopic pregnancy?

    PubMed

    Turkmen, G G; Karçaaltıncaba, D; Isık, H; Fidancı, V; Kaayalp, D; Tımur, H; Batıoglu, S

    2016-01-01

    Early diagnosis of ectopic pregnancy (EP) is important due to life-threatening consequences in the first trimester of pregnancy. In this study we aimed to investigate the role of adenosine deaminase (ADA) activity in the prediction of EP. Forty-one patients with unruptured ectopic pregnancy comprised the case group and forty-two first trimester pregnant women with shown foetal heart beating in ultrasound comprised the control group. The mean ADA level in EP (10.9 ± 3.0 IU/L) was higher than that in control group (9.2 ± 3.6 IU/L) (p = 0.018). Receiver operating characteristics or ROC curve identified ADA value of 10.95 IU/L as optimal threshold for the prediction of EP with 56% sensitivity and 67% specificity. High ADA levels are valuable in the early diagnosis of EP. However more comprehensive studies are required.

  20. Expression of activation-induced cytidine deaminase decreases throughout the life.

    PubMed

    Radu, D L; Kodera, T; Bona, C

    2003-01-01

    Activation-induced cytidine deaminase (AID) is an RNA editing enzyme, which contributes to generation of new functional genes from a restricted number of genes of plant and animal genome. This enzyme was involved in the process of somatic mutation and class switching in vertebrate. Since the rate of somatic mutations is variable throughout ontogeny, we have studied the transcription of AID in 3 to 24 month-old Balb/c mice. Our results demonstrate a significant decrease of the transcription of the AID gene with aging. The decreased AID activity is not related to variation of phenotypic and functional properties of B cells throughout the life. This observation can explain the low rate of somatic mutation in aged animals.

  1. Induction of homologous recombination between sequence repeats by the activation induced cytidine deaminase (AID) protein.

    PubMed

    Buerstedde, Jean-Marie; Lowndes, Noel; Schatz, David G

    2014-07-08

    The activation induced cytidine deaminase (AID) protein is known to initiate somatic hypermutation, gene conversion or switch recombination by cytidine deamination within the immunoglobulin loci. Using chromosomally integrated fluorescence reporter transgenes, we demonstrate a new recombinogenic activity of AID leading to intra- and intergenic deletions via homologous recombination of sequence repeats. Repeat recombination occurs at high frequencies even when the homologous sequences are hundreds of bases away from the positions of AID-mediated cytidine deamination, suggesting DNA end resection before strand invasion. Analysis of recombinants between homeologous repeats yielded evidence for heteroduplex formation and preferential migration of the Holliday junctions to the boundaries of sequence homology. These findings broaden the target and off-target mutagenic potential of AID and establish a novel system to study induced homologous recombination in vertebrate cells.DOI: http://dx.doi.org/10.7554/eLife.03110.001.

  2. Cloning of L-amino acid deaminase gene from Proteus vulgaris.

    PubMed

    Takahashi, E; Ito, K; Yoshimoto, T

    1999-12-01

    The L-amino acid degrading enzyme gene from Proteus vulgaris was cloned and the nucleotide sequence of the enzyme gene was clarified. An open reading frame of 1,413 bp starting at an ATG methionine codon was found, which encodes a protein of 471 amino acid residues, the calculated molecular weight of which is 51,518. The amino acid sequence of P. vulgaris was 58.6% identical with the L-amino acid deaminase of P. mirabilis. A significantly conserved sequence was found around the FAD-binding sequence of flavo-proteins. The partially purified wild and recombinant enzymes had the same substrate specificity for L-amino acids to form the respective keto-acids, however not for D-amino acids.

  3. Non-infectious lung disease in patients with adenosine deaminase deficient severe combined immunodeficiency.

    PubMed

    Booth, C; Algar, V E; Xu-Bayford, J; Fairbanks, L; Owens, C; Gaspar, H B

    2012-06-01

    Adenosine deaminase deficiency is a disorder of purine metabolism manifesting severe combined immunodeficiency (ADA-SCID) and systemic abnormalities. Increased levels of the substrate deoxyadenosine triphosphate (dATP) lead to immunodeficiency and are associated in a murine model with pulmonary insufficiency. We compared a cohort of patients with ADA-SCID and X-linked SCID and found that despite similar radiological and respiratory findings, positive microbiology is significantly less frequent in ADA-SCID patients (p < 0.0005), suggesting a metabolic pathogenesis for the lung disease. Clinicians should be aware of this possibility and correct metabolic abnormalities either through enzyme replacement or haematopoietic stem cell transplant, in addition to treating infectious complications.

  4. Correlation between tumor histology, steroid receptor status, and adenosine deaminase complexing protein immunoreactivity in ovarian cancer.

    PubMed

    Rao, B R; Slotman, B J; Geldof, A A; Dinjens, W N

    1990-01-01

    Adenosine deaminase complexing protein (ADCP) immunoreactivity was investigated in 40 ovarian tumors and correlated with clinicopathologic parameters, including tumor steroid receptor content. Ten (29%) of 34 common epithelial ovarian carcinomas showed ADCP reactivity. Reactivity for ADCP was seen more frequently in mucinous (100%; p less than 0.001), well-differentiated (73%; p less than 0.001) and Stage I (56%; p less than 0.05) ovarian carcinomas. Furthermore, tumors that contained high levels of androgen receptors and tumors that did not contain estrogen receptors were more frequently ADCP positive (p less than 0.05). However, after stratifying for histologic grade, no correlation between ADCP reactivity and receptor status was found. Determination of ADCP reactivity appears to be of limited value in ovarian cancer.

  5. Traffic flow characteristics in a mixed traffic system consisting of ACC vehicles and manual vehicles: A hybrid modelling approach

    NASA Astrophysics Data System (ADS)

    Yuan, Yao-Ming; Jiang, Rui; Hu, Mao-Bin; Wu, Qing-Song; Wang, Ruili

    2009-06-01

    In this paper, we have investigated traffic flow characteristics in a traffic system consisting of a mixture of adaptive cruise control (ACC) vehicles and manual-controlled (manual) vehicles, by using a hybrid modelling approach. In the hybrid approach, (i) the manual vehicles are described by a cellular automaton (CA) model, which can reproduce different traffic states (i.e., free flow, synchronised flow, and jam) as well as probabilistic traffic breakdown phenomena; (ii) the ACC vehicles are simulated by using a car-following model, which removes artificial velocity fluctuations due to intrinsic randomisation in the CA model. We have studied the traffic breakdown probability from free flow to congested flow, the phase transition probability from synchronised flow to jam in the mixed traffic system. The results are compared with that, where both ACC vehicles and manual vehicles are simulated by CA models. The qualitative and quantitative differences are indicated.

  6. Coexistence of Two Forms of LTP in ACC Provides a Synaptic Mechanism for the Interactions between Anxiety and Chronic Pain

    PubMed Central

    Koga, Kohei; Descalzi, Giannina; Chen, Tao; Ko, Hyoung-Gon; Lu, Jinshan; Li, Shermaine; Son, Junehee; Kim, TaeHyun; Kwak, Chuljung; Huganir, Richard L.; Zhao, Ming-gao; Kaang, Bong-Kiun; Collingridge, Graham L.; Zhuo, Min

    2015-01-01

    SUMMARY Chronic pain can lead to anxiety and anxiety can enhance the sensation of pain. Unfortunately, little is known about the synaptic mechanisms that mediate these re-enforcing interactions. Here we characterized two forms of long-term potentiation (LTP) in the anterior cingulate cortex (ACC); a presynaptic form (pre-LTP) that requires kainate receptors and a postsynaptic form (post-LTP) that requires N-methyl-D-aspartate receptors. Pre-LTP also involves adenylyl cyclase and protein kinase A and is expressed via a mechanism involving hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. Interestingly, chronic pain and anxiety both result in selective occlusion of pre-LTP. Significantly, microinjection of the HCN blocker ZD7288 into the ACC in vivo produces both anxiolytic and analgesic effects. Our results provide a mechanism by which two forms of LTP in the ACC may converge to mediate the interaction between anxiety and chronic pain. PMID:25556835

  7. Dispersed sites of HIV Vif-dependent polyubiquitination in the DNA deaminase APOBEC3F

    PubMed Central

    Albin, John S.; Anderson, John S.; Johnson, Jeffrey R.; Harjes, Elena; Matsuo, Hiroshi; Krogan, Nevan J.; Harris, Reuben S.

    2013-01-01

    APOBEC3F and APOBEC3G are DNA cytosine deaminases that potently restrict Human Immunodeficiency Virus-type 1 replication when the virus is deprived of its accessory protein Vif. Vif counteracts these restriction factors by recruiting APOBEC3F and APOBEC3G to an E3 ubiquitin ligase complex that mediates their polyubiquitination and proteasomal degradation. While previous efforts have identified single amino acid residues in APOBEC3 proteins required for Vif recognition, less is known about the downstream ubiquitin acceptor sites that are targeted. One prior report identified a cluster of polyubiquitinated residues in APOBEC3G and proposed an antiparallel model of APOBEC3G interaction with the Vif-E3 ubiquitin ligase complex wherein Vif binding at one terminus of APOBEC3G orients the opposite terminus for polyubiquitination [Iwatani Y, et al. (2009) PNAS 106(46):19539–19544]. To test the generalizability of this model, we carried out a complete mutagenesis of the lysine residues in APOBEC3F and used a complementary, unbiased proteomic approach to identify ubiquitin acceptor sites targeted by Vif. Our data indicate that internal lysines are the dominant ubiquitin acceptor sites in both APOBEC3F and APOBEC3G. In contrast with the proposed antiparallel model, however, we find that the Vif-dependent polyubiquitination of APOBEC3F and APOBEC3G can occur at multiple acceptor sites dispersed along predicted lysine-enriched surfaces of both the N- and C-terminal deaminase domains. These data suggest an alternative model for binding of APOBEC3 proteins to the Vif-E3 ubiquitin ligase complex and diminish enthusiasm for the amenability of APOBEC3 ubiquitin acceptor sites to therapeutic intervention. PMID:23318957

  8. Nucleic acid determinants for selective deamination of DNA over RNA by activation-induced deaminase.

    PubMed

    Nabel, Christopher S; Lee, Jae W; Wang, Laura C; Kohli, Rahul M

    2013-08-27

    Activation-induced deaminase (AID), a member of the larger AID/APOBEC family, is the key catalyst in initiating antibody somatic hypermutation and class-switch recombination. The DNA deamination model accounting for AID's functional role posits that AID deaminates genomic deoxycytosine bases within the immunoglobulin locus, activating downstream repair pathways that result in antibody maturation. Although this model is well supported, the molecular basis for AID's selectivity for DNA over RNA remains an open and pressing question, reflecting a broader need to elucidate how AID/APOBEC enzymes engage their substrates. To address these questions, we have synthesized a series of chimeric nucleic acid substrates and characterized their reactivity with AID. These chimeric substrates feature targeted variations at the 2'-position of nucleotide sugars, allowing us to interrogate the steric and conformational basis for nucleic acid selectivity. We demonstrate that modifications to the target nucleotide can significantly alter AID's reactivity. Strikingly, within a substrate that is otherwise DNA, a single RNA-like 2'-hydroxyl substitution at the target cytosine is sufficient to compromise deamination. Alternatively, modifications that favor a DNA-like conformation (or sugar pucker) are compatible with deamination. AID's closely related homolog APOBEC1 is similarly sensitive to RNA-like substitutions at the target cytosine. Inversely, with unreactive 2'-fluoro-RNA substrates, AID's deaminase activity was rescued by introducing a trinucleotide DNA patch spanning the target cytosine and two nucleotides upstream. These data suggest a role for nucleotide sugar pucker in explaining the molecular basis for AID's DNA selectivity and, more generally, suggest how other nucleic acid-modifying enzymes may distinguish DNA from RNA.

  9. Characterization of a novel resistance-related deoxycytidine deaminase from Brassica oleracea var. capitata.

    PubMed

    Shibu, Marthandam Asokan; Yang, Hsueh-Hui; Lo, Chaur-Tsuen; Lin, Hong-Shin; Liu, Shu-Ying; Peng, Kou-Cheng

    2014-02-26

    Brassica oleracea deoxycytidine deaminase (BoDCD), a deoxycytidine deaminase (DCD, EC 3.5.4.14) enzyme, is known to play an important role in the Trichoderma harzianum ETS 323 mediated resistance mechanism in young leaves of B. oleracea var. capitata during Rhizoctonia solani infection. BoDCD potentially neutralizes cytotoxic products of host lipoxygenase activity, and thereby BoDCD restricts the hypersensitivity-related programmed cell death induced in plants during the initial stages of infection. To determine the biochemical characteristics and to partially elucidate the designated functional properties of BoDCD, the enzyme was cloned into an Escherichia coli expression system, and its potential to neutralize the toxic analogues of 2'-deoxycytidine (dC) was examined. BoDCD transformants of E. coli cells were found to be resistant to 2'-deoxycytidine analogues at all of the concentrations tested. The BoDCD enzyme was also overexpressed as a histidine-tagged protein and purified using nickel chelating affinity chromatography. The molecular weight of BoDCD was determined to be 20.8 kDa as visualized by SDS-PAGE. The substrate specificity and other kinetic properties show that BoDCD is more active in neutralizing cytotoxic cytosine β-d-arabinofuranoside than in deaminating 2'-deoxycytinde to 2'-deoxyuridine in nucleic acids or in metabolizing cytidine to uridine. The optimal temperature and pH of the enzyme were 27 °C and 7.5. The Km and Vmax values of BoDCD were, respectively, 91.3 μM and 1.475 mM for its natural substrate 2'-deoxycytidine and 63 μM and 2.072 mM for cytosine β-d-arabinofuranoside. The phenomenon of neutralization of cytotoxic dC analogues by BoDCD is discussed in detail on the basis of enzyme biochemical properties.

  10. Targeted endostatin-cytosine deaminase fusion gene therapy plus 5-fluorocytosine suppresses ovarian tumor growth.

    PubMed

    Sher, Y-P; Chang, C-M; Juo, C-G; Chen, C-T; Hsu, J L; Lin, C-Y; Han, Z; Shiah, S-G; Hung, M-C

    2013-02-28

    There are currently no effective therapies for cancer patients with advanced ovarian cancer, therefore developing an efficient and safe strategy is urgent. To ensure cancer-specific targeting, efficient delivery, and efficacy, we developed an ovarian cancer-specific construct (Survivin-VISA-hEndoyCD) composed of the cancer specific promoter survivin in a transgene amplification vector (VISA; VP16-GAL4-WPRE integrated systemic amplifier) to express a secreted human endostatin-yeast cytosine deaminase fusion protein (hEndoyCD) for advanced ovarian cancer treatment. hEndoyCD contains an endostatin domain that has tumor-targeting ability for anti-angiogenesis and a cytosine deaminase domain that converts the prodrug 5-fluorocytosine (5-FC) into the chemotherapeutic drug, 5-fluorouracil. Survivin-VISA-hEndoyCD was found to be highly specific, selectively express secreted hEndoyCD from ovarian cancer cells, and induce cancer-cell killing in vitro and in vivo in the presence of 5-FC without affecting normal cells. In addition, Survivin-VISA-hEndoyCD plus 5-FC showed strong synergistic effects in combination with cisplatin in ovarian cancer cell lines. Intraperitoneal (i.p.) treatment with Survivin-VISA-hEndoyCD coupled with liposome attenuated tumor growth and prolonged survival in mice bearing advanced ovarian tumors. Importantly, there was virtually no severe toxicity when hEndoyCD is expressed by Survivin-VISA plus 5-FC compared with CMV plus 5-FC. Thus, the current study demonstrates an effective cancer-targeted gene therapy that is worthy of development in clinical trials for treating advanced ovarian cancer.

  11. Acute intermittent porphyria: expression of mutant and wild-type porphobilinogen deaminase in COS-1 cells.

    PubMed Central

    Mustajoki, S.; Laine, M.; Lahtela, M.; Mustajoki, P.; Peltonen, L.; Kauppinen, R.

    2000-01-01

    BACKGROUND: Acute intermittent porphyria (AIP) is an autosomal dominant disorder that results from the partial deficiency of porphobilinogen deaminase (PBGD) in the heme biosynthetic pathway. Patients with AIP can experience acute attacks consisting of abdominal pain and various neuropsychiatric symptoms. Although molecular biological studies on the porphobilinogen deaminase (PBGD) gene have revealed several mutations responsible for AIP, the properties of mutant PBGD in eukaryotic expression systems have not been studied previously. MATERIALS AND METHODS: Seven mutations were analyzed using transient expression of the mutated polypeptides in COS-1 cells. The properties of mutated polypeptides were studied by enzyme activity measurement, Western blot analysis, pulse-chase experiments, and immunofluorescence staining. RESULTS: Of the mutants studied, R26C, R167W, R173W, R173Q, and R225X resulted in a decreased enzyme activity (0-5%), but R225G and 1073delA (elongated protein) displayed a significant residual activity of 16% and 50%, respectively. In Western blot analysis, the polyclonal PBGD antibody detected all mutant polypeptides except R225X, which was predicted to result in a truncated protein. In the pulse-chase experiment, the mutant polypeptides were as stable as the wild-type enzyme. In the immunofluorescence staining both wild-type and mutant polypeptides were diffusely dispersed in the cytoplasm and, thus, no accumulation of mutated proteins in the cellular compartments could be observed. CONCLUSIONS: The results confirm the causality of mutations for the half normal enzyme activity measured in the patients' erythrocytes. In contrast to the decreased enzyme activity, the majority of the mutations produced a detectable polypeptide, and the stability and the intracellular processing of the mutated polypeptides were both comparable to that of the wild-type PBGD and independent of the cross-reacting immunological material (CRIM) class. PMID:11055586

  12. Acceleration of a Particle-in-Cell Code for Space Plasma Simulations with OpenACC

    NASA Astrophysics Data System (ADS)

    Peng, Ivy Bo; Markidis, Stefano; Vaivads, Andris; Vencels, Juris; Deca, Jan; Lapenta, Giovanni; Hart, Alistair; Laure, Erwin

    2015-04-01

    We simulate space plasmas with the Particle-in-cell (PIC) method that uses computational particles to mimic electrons and protons in solar wind and in Earth magnetosphere. The magnetic and electric fields are computed by solving the Maxwell's equations on a computational grid. In each PIC simulation step, there are four major phases: interpolation of fields to particles, updating the location and velocity of each particle, interpolation of particles to grids and solving the Maxwell's equations on the grid. We use the iPIC3D code, which was implemented in C++, using both MPI and OpenMP, for our case study. By November 2014, heterogeneous systems using hardware accelerators such as Graphics Processing Unit (GPUs) and the Many Integrated Core (MIC) coprocessors for high performance computing continue growth in the top 500 most powerful supercomputers world wide. Scientific applications for numerical simulations need to adapt to using accelerators to achieve portability and scalability in the coming exascale systems. In our work, we conduct a case study of using OpenACC to offload the computation intensive parts: particle mover and interpolation of particles to grids, in a massively parallel Particle-in-Cell simulation code, iPIC3D, to multi-GPU systems. We use MPI for inter-node communication for halo exchange and communicating particles. We identify the most promising parts suitable for GPUs accelerator by profiling using CrayPAT. We implemented manual deep copy to address the challenges of porting C++ classes to GPU. We document the necessary changes in the exiting algorithms to adapt for GPU computation. We present the challenges and findings as well as our methodology for porting a Particle-in-Cell code to multi-GPU systems using OpenACC. In this work, we will present the challenges, findings and our methodology of porting a Particle-in-Cell code for space applications as follows: We profile the iPIC3D code by Cray Performance Analysis Tool (CrayPAT) and identify

  13. Identification and characterization of an Apis cerana cerana Delta class glutathione S-transferase gene (AccGSTD) in response to thermal stress.

    PubMed

    Yan, Huiru; Jia, Haihong; Wang, Xiuling; Gao, Hongru; Guo, Xingqi; Xu, Baohua

    2013-02-01

    Glutathione S-transferases (GSTs) are members of a multifunctional enzyme super family that plays a pivotal role in both insecticide resistance and protection against oxidative stress. In this study, we identified a single-copy gene, AccGSTD, as being a Delta class GST in the Chinese honey bee (Apis cerana cerana). A predicted antioxidant response element, CREB, was found in the 1,492-bp 5'-flanking region, suggesting that AccGSTD may be involved in oxidative stress response pathways. Real-time PCR and immunolocalization studies demonstrated that AccGSTD exhibited both developmental- and tissue-specific expression patterns. During development, AccGSTD transcript was increased in adults. The AccGSTD expression level was the highest in the honey bee brain. Thermal stress experiments demonstrated that AccGSTD could be significantly upregulated by temperature changes in a time-dependent manner. It is hypothesized that high expression levels might be due to the increased levels of oxidative stress caused by the temperature challenges. Additionally, functional assays of the recombinant AccGSTD protein revealed that AccGSTD has the capability to protect DNA from oxidative damage. Taken together, these data suggest that AccGSTD may be responsible for antioxidant defense in adult honey bees.

  14. Identification and characterization of an Apis cerana cerana Delta class glutathione S-transferase gene ( AccGSTD) in response to thermal stress

    NASA Astrophysics Data System (ADS)

    Yan, Huiru; Jia, Haihong; Wang, Xiuling; Gao, Hongru; Guo, Xingqi; Xu, Baohua

    2013-02-01

    Glutathione S-transferases (GSTs) are members of a multifunctional enzyme super family that plays a pivotal role in both insecticide resistance and protection against oxidative stress. In this study, we identified a single-copy gene, AccGSTD, as being a Delta class GST in the Chinese honey bee ( Apis cerana cerana). A predicted antioxidant response element, CREB, was found in the 1,492-bp 5'-flanking region, suggesting that AccGSTD may be involved in oxidative stress response pathways. Real-time PCR and immunolocalization studies demonstrated that AccGSTD exhibited both developmental- and tissue-specific expression patterns. During development, AccGSTD transcript was increased in adults. The AccGSTD expression level was the highest in the honey bee brain. Thermal stress experiments demonstrated that AccGSTD could be significantly upregulated by temperature changes in a time-dependent manner. It is hypothesized that high expression levels might be due to the increased levels of oxidative stress caused by the temperature challenges. Additionally, functional assays of the recombinant AccGSTD protein revealed that AccGSTD has the capability to protect DNA from oxidative damage. Taken together, these data suggest that AccGSTD may be responsible for antioxidant defense in adult honey bees.

  15. The ADA*2 allele of the adenosine deaminase gene (20q13.11) and recurrent spontaneous abortions: an age-dependent association

    PubMed Central

    Nunes, Daniela Prudente Teixeira; Spegiorin, Lígia Cosentino Junqueira Franco; de Mattos, Cinara Cássia Brandão; Oliani, Antonio Helio; Vaz-Oliani, Denise Cristina Mós; de Mattos, Luiz Carlos

    2011-01-01

    OBJECTIVE: Adenosine deaminase acts on adenosine and deoxyadenosine metabolism and modulates the immune response. The adenosine deaminase G22A polymorphism (20q.11.33) influences the level of adenosine deaminase enzyme expression, which seems to play a key role in maintaining pregnancy. The adenosine deaminase 2 phenotype has been associated with a protective effect against recurrent spontaneous abortions in European Caucasian women. The aim of this study was to investigate whether the G22A polymorphism of the adenosine deaminase gene is associated with recurrent spontaneous abortions in Brazilian women. METHODS: A total of 311 women were recruited to form two groups: G1, with a history of recurrent spontaneous abortions (N = 129), and G2, without a history of abortions (N = 182). Genomic DNA was extracted from peripheral blood with a commercial kit and PCR-RFLP analysis was used to identify the G22A genetic polymorphism. Fisher's exact test and odds ratio values were used to compare the proportions of adenosine deaminase genotypes and alleles between women with and without a history of recurrent spontaneous abortion (p<0.05). The differences between mean values for categorical data were calculated using unpaired t tests. The Hardy-Weinberg equilibrium was assessed with a chi-square test. RESULTS: Statistically significant differences were identified for the frequencies of adenosine deaminase genotypes and alleles between the G1 and G2 groups when adjusted for maternal age. CONCLUSIONS: The results suggest that the adenosine deaminase *2 allele is associated with a low risk for recurrent spontaneous abortions, but this association is dependent on older age. PMID:22086524

  16. N-terminal amino acid sequences of D-serine deaminases of wild-type and operator-constitutive strains of Escherichia coli K-12.

    PubMed Central

    Heincz, M C; McFall, E

    1975-01-01

    The N-terminal amino acid sequences of the D-serine deaminases from strains of Escherichia coli K-12 that harbor wild-type and high-level constitutive catabolite-insensitive operator-initiator regions are identical: Met-Ser-GluNH2-Ser-Gly-Arg-His-Cys. This result indicates that the operator-initiator region is probably distinct from the D-serine deaminase structural gene. Images PMID:1099073

  17. N-terminal and C-terminal cytosine deaminase domain of APOBEC3G inhibit hepatitis B virus replication

    PubMed Central

    Lei, Yan-Chang; Tian, Yong-Jun; Ding, Hong-Hui; Wang, Bao-Ju; Yang, Yan; Hao, You-Hua; Zhao, Xi-Ping; Lu, Meng-Ji; Gong, Fei-Li; Yang, Dong-Liang

    2006-01-01

    AIM: To investigate the effect of human apolipoprotein B mRNA-editing enzyme catalytic-polypeptide 3G (APOBEC3G) and its N-terminal or C-terminal cytosine deaminase domain-mediated antiviral activity against hepatitis B virus (HBV) in vitro and in vivo. METHODS: The mammalian hepatoma cells HepG2 and HuH7 were cotransfected with APOBEC3G and its N-terminal or C-terminal cytosine deaminase domain expression vector and 1.3-fold-overlength HBV DNA as well as the linear monomeric HBV of genotype B and C. For in vivo study, an HBV vector-based mouse model was used in which APOBEC3G and its N-terminal or C-terminal cytosine deaminase domain expression vectors were co-delivered with 1.3-fold-overlength HBV DNA via high-volume tail vein injection. Levels of hepatitis B virus surface antigen (HBsAg) and hepatitis B virus e antigen (HBeAg) in the media of the transfected cells and in the sera of mice were determined by ELISA. The expression of hepatitis B virus core antigen (HBcAg) in the transfected cells was determined by Western blot analysis. Core-associated HBV DNA was examined by Southern blot analysis. Levels of HBV DNA in the sera of mice as well as HBV core-associated RNA in the liver of mice were determined by quantitative PCR and quantitative RT-PCR analysis, respectively. RESULTS: Human APOBEC3G exerted an anti-HBV activity in a dose-dependent manner in HepG2 cells, and comparable suppressive effects were observed on genotype B and C as that of genotype A. Interestingly, the N-terminal or C-terminal cytosine deaminase domain alone could also inhibit HBV replication in HepG2 cells as well as Huh7 cells. Consistent with in vitro results, the levels of HBsAg in the sera of mice were dramatically decreased, with more than 50 times decrease in the levels of serum HBV DNA and core-associated RNA in the liver of mice treated with APOBEC3G and its N-terminal or C-terminal cytosine deaminase domain as compared to the controls. CONCLUSION: Our findings provide probably the

  18. Passive monitoring of VOC in air using ACC. Final report, revised edition

    SciTech Connect

    Gesser, H.D.

    1997-12-31

    The report describes a project that developed a method of using activated carbon cloth (ACC) as a sampler for measuring volatile organic compounds (VOCs) in air. Strips of carbon cloth mounted in slide holders were tested as diffusive samplers exposed to known concentrations of standard chemicals in test chambers. Adsorbed chemicals were extracted with solvents and analyzed, and comparison was made between the results obtained with the cloths and with passive samplers. Preliminary tests were carried out to determine the effects on carbon cloth performance of relative humidity, methods of extraction, materials for storing the cloth strips, and the type of cloth. Appendices include tables of experimental data and a report on a novel method of solventless extraction based on a combination of solid phase micro-extraction and purge/trap methods.

  19. Overview of the arthritis Cost Consequence Evaluation System (ACCES): a pharmacoeconomic model for celecoxib.

    PubMed

    Pettitt, D; Goldstein, J L; McGuire, A; Schwartz, J S; Burke, T; Maniadakis, N

    2000-12-01

    Pharmacoeconomic analyses have become useful and essential tools for health care decision makers who increasingly require such analyses prior to placing a drug on a national, regional or hospital formulary. Previous health economic models of non-steroidal anti-inflammatory drugs (NSAIDs) have been restricted to evaluating a narrow range of agents within specific health care delivery systems using medical information derived from homogeneous clinical trial data. This paper summarizes the Arthritis Cost Consequence Evaluation System (ACCES)--a pharmacoeconomic model that has been developed to predict and evaluate the costs and consequences associated with the use of celecoxib in patients with arthritis, compared with other NSAIDs and NSAIDs plus gastroprotective agents. The advantage of this model is that it can be customized to reflect local practice patterns, resource utilization and costs, as well as provide context-specific health economic information to a variety of providers and/or decision makers.

  20. Sequencing of allotetraploid cotton (Gossypium hirsutum L. acc. TM-1) provides a resource for fiber improvement.

    PubMed

    Zhang, Tianzhen; Hu, Yan; Jiang, Wenkai; Fang, Lei; Guan, Xueying; Chen, Jiedan; Zhang, Jinbo; Saski, Christopher A; Scheffler, Brian E; Stelly, David M; Hulse-Kemp, Amanda M; Wan, Qun; Liu, Bingliang; Liu, Chunxiao; Wang, Sen; Pan, Mengqiao; Wang, Yangkun; Wang, Dawei; Ye, Wenxue; Chang, Lijing; Zhang, Wenpan; Song, Qingxin; Kirkbride, Ryan C; Chen, Xiaoya; Dennis, Elizabeth; Llewellyn, Danny J; Peterson, Daniel G; Thaxton, Peggy; Jones, Don C; Wang, Qiong; Xu, Xiaoyang; Zhang, Hua; Wu, Huaitong; Zhou, Lei; Mei, Gaofu; Chen, Shuqi; Tian, Yue; Xiang, Dan; Li, Xinghe; Ding, Jian; Zuo, Qiyang; Tao, Linna; Liu, Yunchao; Li, Ji; Lin, Yu; Hui, Yuanyuan; Cao, Zhisheng; Cai, Caiping; Zhu, Xiefei; Jiang, Zhi; Zhou, Baoliang; Guo, Wangzhen; Li, Ruiqiang; Chen, Z Jeffrey

    2015-05-01

    Upland cotton is a model for polyploid crop domestication and transgenic improvement. Here we sequenced the allotetraploid Gossypium hirsutum L. acc. TM-1 genome by integrating whole-genome shotgun reads, bacterial artificial chromosome (BAC)-end sequences and genotype-by-sequencing genetic maps. We assembled and annotated 32,032 A-subgenome genes and 34,402 D-subgenome genes. Structural rearrangements, gene loss, disrupted genes and sequence divergence were more common in the A subgenome than in the D subgenome, suggesting asymmetric evolution. However, no genome-wide expression dominance was found between the subgenomes. Genomic signatures of selection and domestication are associated with positively selected genes (PSGs) for fiber improvement in the A subgenome and for stress tolerance in the D subgenome. This draft genome sequence provides a resource for engineering superior cotton lines.

  1. Reward sensitivity modulates brain activity in the prefrontal cortex, ACC and striatum during task switching.

    PubMed

    Fuentes-Claramonte, Paola; Ávila, César; Rodríguez-Pujadas, Aina; Ventura-Campos, Noelia; Bustamante, Juan C; Costumero, Víctor; Rosell-Negre, Patricia; Barrós-Loscertales, Alfonso

    2015-01-01

    Current perspectives on cognitive control acknowledge that individual differences in motivational dispositions may modulate cognitive processes in the absence of reward contingencies. This work aimed to study the relationship between individual differences in Behavioral Activation System (BAS) sensitivity and the neural underpinnings involved in processing a switching cue in a task-switching paradigm. BAS sensitivity was hypothesized to modulate brain activity in frontal regions, ACC and the striatum. Twenty-eight healthy participants underwent fMRI while performing a switching task, which elicited activity in fronto-striatal regions during the processing of the switch cue. BAS sensitivity was negatively associated with activity in the lateral prefrontal cortex, anterior cingulate cortex and the ventral striatum. Combined with previous results, our data indicate that BAS sensitivity modulates the neurocognitive processes involved in task switching in a complex manner depending on task demands. Therefore, individual differences in motivational dispositions may influence cognitive processing in the absence of reward contingencies.

  2. Reward Sensitivity Modulates Brain Activity in the Prefrontal Cortex, ACC and Striatum during Task Switching

    PubMed Central

    Fuentes-Claramonte, Paola; Ávila, César; Rodríguez-Pujadas, Aina; Ventura-Campos, Noelia; Bustamante, Juan C.; Costumero, Víctor; Rosell-Negre, Patricia; Barrós-Loscertales, Alfonso

    2015-01-01

    Current perspectives on cognitive control acknowledge that individual differences in motivational dispositions may modulate cognitive processes in the absence of reward contingencies. This work aimed to study the relationship between individual differences in Behavioral Activation System (BAS) sensitivity and the neural underpinnings involved in processing a switching cue in a task-switching paradigm. BAS sensitivity was hypothesized to modulate brain activity in frontal regions, ACC and the striatum. Twenty-eight healthy participants underwent fMRI while performing a switching task, which elicited activity in fronto-striatal regions during the processing of the switch cue. BAS sensitivity was negatively associated with activity in the lateral prefrontal cortex, anterior cingulate cortex and the ventral striatum. Combined with previous results, our data indicate that BAS sensitivity modulates the neurocognitive processes involved in task switching in a complex manner depending on task demands. Therefore, individual differences in motivational dispositions may influence cognitive processing in the absence of reward contingencies. PMID:25875640

  3. Crystal Structure of Staphylococcus aureus tRNA Adenosine Deaminase TadA in Complex with RNA

    SciTech Connect

    Losey,H.; Ruthenburg, A.; Verdine, G.

    2006-01-01

    Bacterial tRNA adenosine deaminases (TadAs) catalyze the hydrolytic deamination of adenosine to inosine at the wobble position of tRNA(Arg2), a process that enables this single tRNA to recognize three different arginine codons in mRNA. In addition, inosine is also introduced at the wobble position of multiple eukaryotic tRNAs. The genes encoding these deaminases are essential in bacteria and yeast, demonstrating the importance of their biological activity. Here we report the crystallization and structure determination to 2.0 A of Staphylococcus aureus TadA bound to the anticodon stem-loop of tRNA(Arg2) bearing nebularine, a non-hydrolyzable adenosine analog, at the wobble position. The cocrystal structure reveals the basis for both sequence and structure specificity in the interactions of TadA with RNA, and it additionally provides insight into the active site architecture that promotes efficient hydrolytic deamination.

  4. Protein preparation and preliminary X-ray crystallographic analysis of a putative glucosamine 6-phosphate deaminase from Streptococcus mutants

    SciTech Connect

    Hu, Guan-Jing; Li, Lan-Fen; Li, Dan; Liu, Cong; Wei, Shi-Cheng; Liang, Yu-He Su, Xiao-Dong

    2007-09-01

    A glucosamine 6-phosphate deaminase homologue from S. mutans was expressed, purified and crystallized. Diffraction data have been collected to 2.4 Å resolution. The SMU.636 protein from Streptococcus mutans is a putative glucosamine 6-phosphate deaminase with 233 residues. The smu.636 gene was PCR-amplified from S. mutans genomic DNA and cloned into the expression vector pET-28a(+). The resultant His-tagged fusion protein was expressed in Escherichia coli and purified to homogeneity in two steps. Crystals of the fusion protein were obtained by the hanging-drop vapour-diffusion method. The crystals diffracted to 2.4 Å resolution and belong to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 53.83, b = 82.13, c = 134.70 Å.

  5. Improved Performance of an Air Cooled Condenser (ACC) Using SPX Wind Guide Technology at Coal-Based Thermoelectric Power Plants

    SciTech Connect

    Ken Mortensen

    2010-12-31

    This project added a new airflow enhancement technology to an existing ACC cooling process at a selected coal power plant. Airflow parameters and efficiency improvement for the main plant cooling process using the applied technology were determined and compared with the capabilities of existing systems. The project required significant planning and pre-test execution in order to reach the required Air Cooled Condenser system configuration for evaluation. A host Power Plant ACC system had to be identified, agreement finalized, and addition of the SPX ACC Wind Guide Technology completed on that site. Design of the modification, along with procurement, fabrication, instrumentation, and installation of the new airflow enhancement technology were executed. Baseline and post-modification cooling system data was collected and evaluated. The improvement of ACC thermal performance after SPX wind guide installation was clear. Testing of the improvement indicates there is a 5% improvement in heat transfer coefficient in high wind conditions and 1% improvement at low wind speed. The benefit increased with increasing wind speed. This project was completed on schedule and within budget.

  6. 24 CFR 884.105 - Maximum total ACC commitment and project account (private-owner/PHA projects).

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 24 Housing and Urban Development 4 2010-04-01 2010-04-01 false Maximum total ACC commitment and project account (private-owner/PHA projects). 884.105 Section 884.105 Housing and Urban Development Regulations Relating to Housing and Urban Development (Continued) OFFICE OF THE ASSISTANT SECRETARY...

  7. The human ACC2 CT-domain C-terminus is required for full functionality and has a novel twist

    PubMed Central

    Madauss, Kevin P.; Burkhart, William A.; Consler, Thomas G.; Cowan, David J.; Gottschalk, William K.; Miller, Aaron B.; Short, Steven A.; Tran, Thuy B.; Williams, Shawn P.

    2009-01-01

    Inhibition of acetyl-CoA carboxylase (ACC) may prevent lipid-induced insulin resistance and type 2 diabetes, making the enzyme an attractive pharmaceutical target. Although the enzyme is highly conserved amongst animals, only the yeast enzyme structure is available for rational drug design. The use of biophysical assays has permitted the identification of a specific C-terminal truncation of the 826-residue human ACC2 carboxyl transferase (CT) domain that is both functionally competent to bind inhibitors and crystallizes in their presence. This C-terminal truncation led to the determination of the human ACC2 CT domain–CP-640186 complex crystal structure, which revealed distinctions from the yeast-enzyme complex. The human ACC2 CT-domain C-terminus is com­prised of three intertwined α-helices that extend outwards from the enzyme on the opposite side to the ligand-binding site. Differences in the observed inhibitor conformation between the yeast and human structures are caused by differing residues in the binding pocket. PMID:19390150

  8. Draft Genome Sequence of Edwardsiellapiscicida Strain ACC35.1 Isolated from Diseased Turbot (Scophthalmus maximus) in Europe.

    PubMed

    Buján, Noemí; Toranzo, Alicia E; Magariños, Beatriz

    2017-02-16

    Edwardsiella piscicida is a bacterial fish pathogen with a high degree of virulence. The strain ACC35.1 was isolated from diseased turbot in Europe. The draft genome sequence comprises 3.84 Mb with a G+C content of 59.8% and >3,450 protein-coding genes.

  9. Tuning the light in senior care: Evaluating a trial LED lighting system at the ACC Care Center in Sacramento, CA

    SciTech Connect

    Davis, Robert G.; Wilkerson, Andrea M.

    2016-08-31

    This report summarizes the results from a trial installation of light-emitting diode (LED) lighting systems in several spaces within the ACC Care Center in Sacramento, CA. The Sacramento Municipal Utility District (SMUD) coordinated the project and invited the U.S. Department of Energy (DOE) to document the performance of the LED lighting systems as part of a GATEWAY evaluation. DOE tasked the Pacific Northwest National Laboratory (PNNL) to conduct the investigation. SMUD and ACC staff coordinated and completed the design and installation of the LED systems, while PNNL and SMUD staff evaluated the photometric performance of the systems. ACC staff also track behavioral and health measures of the residents; some of those results are reported here, although PNNL staff were not directly involved in collecting or interpreting those data. The trial installation took place in a double resident room and a single resident room, and the corridor that connects those (and other) rooms to the central nurse station. Other spaces in the trial included the nurse station, a common room called the family room located near the nurse station, and the ACC administrator’s private office.

  10. Implementation and efficiency analysis of parallel computation using OpenACC: a case study using flow field simulations

    NASA Astrophysics Data System (ADS)

    Zhang, Shanghong; Yuan, Rui; Wu, Yu; Yi, Yujun

    2016-01-01

    The Open Accelerator (OpenACC) application programming interface is a relatively new parallel computing standard. In this paper, particle-based flow field simulations are examined as a case study of OpenACC parallel computation. The parallel conversion process of the OpenACC standard is explained, and further, the performance of the flow field parallel model is analysed using different directive configurations and grid schemes. With careful implementation and optimisation of the data transportation in the parallel algorithm, a speedup factor of 18.26× is possible. In contrast, a speedup factor of just 11.77× was achieved with the conventional Open Multi-Processing (OpenMP) parallel mode on a 20-kernel computer. These results demonstrate that optimised feature settings greatly influence the degree of speedup, and models involving larger numbers of calculations exhibit greater efficiency and higher speedup factors. In addition, the OpenACC parallel mode is found to have good portability, making it easy to implement parallel computation from the original serial model.

  11. The human ACC2 CT-domain C-terminus is required for full functionality and has a novel twist

    SciTech Connect

    Madauss, Kevin P.; Burkhart, William A.; Consler, Thomas G.; Cowan, David J.; Gottschalk, William K.; Miller, Aaron B; Short, Steven A.; Tran, Thuy B.; Williams, Shawn P.

    2009-06-15

    Inhibition of acetyl-CoA carboxylase (ACC) may prevent lipid-induced insulin resistance and type 2 diabetes, making the enzyme an attractive pharmaceutical target. Although the enzyme is highly conserved amongst animals, only the yeast enzyme structure is available for rational drug design. The use of biophysical assays has permitted the identification of a specific C-terminal truncation of the 826-residue human ACC2 carboxyl transferase (CT) domain that is both functionally competent to bind inhibitors and crystallizes in their presence. This C-terminal truncation led to the determination of the human ACC2 CT domain-CP-640186 complex crystal structure, which revealed distinctions from the yeast-enzyme complex. The human ACC2 CT-domain C-terminus is comprised of three intertwined -helices that extend outwards from the enzyme on the opposite side to the ligand-binding site. Differences in the observed inhibitor conformation between the yeast and human structures are caused by differing residues in the binding pocket.

  12. Draft Genome Sequence of Edwardsiella piscicida Strain ACC35.1 Isolated from Diseased Turbot (Scophthalmus maximus) in Europe

    PubMed Central

    Toranzo, Alicia E.; Magariños, Beatriz

    2017-01-01

    ABSTRACT   Edwardsiella piscicida is a bacterial fish pathogen with a high degree of virulence. The strain ACC35.1 was isolated from diseased turbot in Europe. The draft genome sequence comprises 3.84 Mb with a G+C content of 59.8% and >3,450 protein-coding genes. PMID:28209828

  13. NATO Network Enabled Capability (NNEC) challenges: Why NATO Air Command and Control System (ACCS) might be a good case?

    DTIC Science & Technology

    2011-06-01

    Picture ( RAP ), Air Tasking Order (ATO), Air Coordination order (ACO), etc. 2. Non Functional: providing all other functionalities, e.g. IA/security...with their respective characteristics in the project management behavior and environmental parameters. It should be noted that SOA implementation... influence its transformation. The roles and perspectives of the main ACCS transformation actors are described in this section. Several stakeholders

  14. Expression of recombinant AccMRJP1 protein from royal jelly of Chinese honeybee in Pichia pastoris and its proliferation activity in an insect cell line

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Main royal jelly protein 1 (MRJP1) is the most abundant member of the main royal jelly protein (MRJP) family among honeybees. Mature MRJP1 cDNA of the Chinese honeybee (Apis cerana cerana MRJP1, or AccMRJP1) was expressed in Pichia pastoris. SDS-PAGE showed that recombinant AccMRJP1 was identical in...

  15. Isolation of a Saccharomyces cerevisiae mutant strain deficient in deoxycytidylate deaminase activity and partial characterization of the enzyme.

    PubMed Central

    McIntosh, E M; Haynes, R H

    1984-01-01

    Deoxycytidylate deaminase activity in Saccharomyces cerevisiae has been partially characterized. The yeast enzyme was found to exhibit properties similar to those of dCMP deaminases isolated from higher eucaryotes. A mutant strain completely deficient in dCMP deaminase activity was isolated by selection for resistance to 5-fluoro-2'-deoxycytidylate followed by screening for cross sensitivity to 5-fluoro-2'-deoxyuridylate, a potent inhibitor of the yeast thymidylate synthetase. We have designated this new allele dcd1 . A strain exhibiting an auxotrophic requirement for dUMP was isolated after mutagenesis of a dcd1 tup7 haploid. Genetic analysis revealed that this auxotrophic phenotype resulted from a combination of the dcd1 allele and a second, unlinked, nuclear mutation that we designated dmp1 . This allele, which by itself conveys no readily discernible phenotype, presumably impairs efficient synthesis of dUMP from UDP. The auxotrophic requirement of dcd1 dmp1 tup7 strains also can be satisfied by exogenous dTMP but not deoxyuridine. PMID:6373725

  16. Active RNAP pre-initiation sites are highly mutated by cytidine deaminases in yeast, with AID targeting small RNA genes

    PubMed Central

    Taylor, Benjamin JM; Wu, Yee Ling; Rada, Cristina

    2014-01-01

    Cytidine deaminases are single stranded DNA mutators diversifying antibodies and restricting viral infection. Improper access to the genome leads to translocations and mutations in B cells and contributes to the mutation landscape in cancer, such as kataegis. It remains unclear how deaminases access double stranded genomes and whether off-target mutations favor certain loci, although transcription and opportunistic access during DNA repair are thought to play a role. In yeast, AID and the catalytic domain of APOBEC3G preferentially mutate transcriptionally active genes within narrow regions, 110 base pairs in width, fixed at RNA polymerase initiation sites. Unlike APOBEC3G, AID shows enhanced mutational preference for small RNA genes (tRNAs, snoRNAs and snRNAs) suggesting a putative role for RNA in its recruitment. We uncover the high affinity of the deaminases for the single stranded DNA exposed by initiating RNA polymerases (a DNA configuration reproduced at stalled polymerases) without a requirement for specific cofactors. DOI: http://dx.doi.org/10.7554/eLife.03553.001 PMID:25237741

  17. Molecular cloning of cDNA for double-stranded RNA adenosine deaminase, a candidate enzyme for nuclear RNA editing.

    PubMed Central

    Kim, U; Wang, Y; Sanford, T; Zeng, Y; Nishikura, K

    1994-01-01

    We have cloned human cDNA encoding double-stranded RNA adenosine deaminase (DRADA). DRADA is a ubiquitous nuclear enzyme that converts multiple adenosines to inosines in double-helical RNA substrates without apparent sequence specificity. The A --> I conversion activity of the protein encoded by the cloned cDNA was confirmed by recombinant expression in insect cells. Use of the cloned DNA as a molecular probe documented sequence conservation across mammals and detected a single transcript of 7 kb in RNA of all human tissues analyzed. The deduced primary structure of human DRADA revealed a bipartite nuclear localization signal, three repeats of a double-stranded RNA binding motif, and the presence of sequences conserved in the catalytic center of other deaminases, including a cytidine deaminase involved in the RNA editing of apolipoprotein B. These structural properties are consistent with the enzymatic signature of DRADA, and strengthen the hypothesis that DRADA carries out the RNA editing of transcripts encoding glutamate-gated ion channels in brain. Images PMID:7972084

  18. Auxin and ethylene induce flavonol accumulation through distinct transcriptional networks.

    PubMed

    Lewis, Daniel R; Ramirez, Melissa V; Miller, Nathan D; Vallabhaneni, Prashanthi; Ray, W Keith; Helm, Richard F; Winkel, Brenda S J; Muday, Gloria K

    2011-05-01

    Auxin and ethylene are key regulators of plant growth and development, and thus the transcriptional networks that mediate responses to these hormones have been the subject of intense research. This study dissected the hormonal cross talk regulating the synthesis of flavonols and examined their impact on root growth and development. We analyzed the effects of auxin and an ethylene precursor on roots of wild-type and hormone-insensitive Arabidopsis (Arabidopsis thaliana) mutants at the transcript, protein, and metabolite levels at high spatial and temporal resolution. Indole-3-acetic acid (IAA) and 1-aminocyclopropane-1-carboxylic acid (ACC) differentially increased flavonol pathway transcripts and flavonol accumulation, altering the relative abundance of quercetin and kaempferol. The IAA, but not ACC, response is lost in the transport inhibitor response1 (tir1) auxin receptor mutant, while ACC responses, but not IAA responses, are lost in ethylene insensitive2 (ein2) and ethylene resistant1 (etr1) ethylene signaling mutants. A kinetic analysis identified increases in transcripts encoding the transcriptional regulators MYB12, Transparent Testa Glabra1, and Production of Anthocyanin Pigment after hormone treatments, which preceded increases in transcripts encoding flavonoid biosynthetic enzymes. In addition, myb12 mutants were insensitive to the effects of auxin and ethylene on flavonol metabolism. The equivalent phenotypes for transparent testa4 (tt4), which makes no flavonols, and tt7, which makes kaempferol but not quercetin, showed that quercetin derivatives are the inhibitors of basipetal root auxin transport, gravitropism, and elongation growth. Collectively, these experiments demonstrate that auxin and ethylene regulate flavonol biosynthesis through distinct signaling networks involving TIR1 and EIN2/ETR1, respectively, both of which converge on MYB12. This study also provides new evidence that quercetin is the flavonol that modulates basipetal auxin transport.

  19. Calcium ion dependency of ethylene production in segments of primary roots of Zea mays

    NASA Technical Reports Server (NTRS)

    Hasenstein, K. H.; Evans, M. L.

    1986-01-01

    We investigated the effect of Ca2+ on ethylene production in 2-cm long apical segments from primary roots of corn (Zea mays L., B73 x Missouri 17) seedlings. The seedlings were raised under different conditions of Ca2+ availability. Low-Ca and high-Ca seedlings were raised by soaking the grains and watering the seedlings with distilled water or 10 mM CaCl2, respectively. Segments from high-Ca roots produced more than twice as much ethylene as segments from low-Ca roots. Indoleacetic acid (IAA; 1 micromole) enhanced ethylene production in segments from both low-Ca and high-Ca roots but auxin-induced promotion of ethylene production was consistently higher in segments from high-Ca roots. Addition of 1-aminocyclopropane-1-carboxylic acid (ACC) to root segments from low-Ca seedlings doubled total ethylene production and the rate of production remained fairly constant during a 24 h period of monitoring. In segments from high-Ca seedlings ACC also increased total ethylene production but most of the ethylene was produced within the first 6 h. The data suggest that Ca2+ enhances the conversion of ACC to ethylene. The terminal 2 mm of the root tip were found to be especially important to ethylene biosynthesis by apical segments and, experiments using 45Ca2+ as tracer indicated that the apical 2 mm of the root is the region of strongest Ca2+ accumulation. Other cations such as Mn2+, Mg2+, and K+ could largely substitute for Ca2+. The significance of these findings is discussed with respect to recent evidence for gravity-induced Ca2+ redistribution and its relationship to the establishment of asymmetric growth during gravitropic curvature.

  20. Identification and characterization of cDNAs encoding ethylene biosynthetic enzymes from Pelargonium x hortorum cv Snow Mass leaves.

    PubMed Central

    Wang, T W; Arteca, R N

    1995-01-01

    Two Pelargonium 1-aminocyclopropane-1-carboxylate (ACC) synthase cDNAs (GAC-1 and GAC-2) were identified and characterized. GAC-1 is 1934 bp long with a 1446-bp open reading frame encoding a 54.1-kD polypeptide. GAC-2 is a 1170-bp-long ACC synthase polymerase chain reaction fragment encoding 390 amino acids. Expression of GAC-1 and GAC-2 together with a previously identified ACC oxidase (GEFE-1) was examined in different Pelargonium plant parts, and leaves were subjected to osmotic stress (sorbitol), metal ion stress (CuCl2), auxin (2,4-dichlorophenoxyacetic acid [2,4-D]), and ethylene. GAC-1 expression was not detectable in any of the plant parts tested, whereas high levels of GAC-2 were expressed in the leaf bud, young leaf, young floret, fully open floret, and senescing floret. GAC-2 was expressed to a lesser degree in fully expanded leaves or roots and was undetectable in old leaves and floret buds. GEFE-1 was detectable at all leaf ages tested, in young and fully open florets, and in the roots; however, the highest degree of expression was in the senescing florets. GAC-1 was induced by sorbitol. Both GAC-1 and GAC-2 were only slightly affected by CuCl2 and induced indirectly by 2,4-D. GEFE-1 was highly induced by sorbitol, CuCl2, and 2,4-D. GAC-1, GAC-2, and GEFE-1 were unaffected by ethylene treatment. These results suggest that GAC-1 is only induced by stress and that GAC-2 may be developmentally regulated, whereas GEFE-1 is influenced by both stress and development. PMID:7480351

  1. Effects of abscisic acid on ethylene biosynthesis and perception in Hibiscus rosa-sinensis L. flower development.

    PubMed

    Trivellini, Alice; Ferrante, Antonio; Vernieri, Paolo; Serra, Giovanni

    2011-11-01

    The effect of the complex relationship between ethylene and abscisic acid (ABA) on flower development and senescence in Hibiscus rosa-sinensis L. was investigated. Ethylene biosynthetic (HrsACS and HrsACO) and receptor (HrsETR and HrsERS) genes were isolated and their expression evaluated in three different floral tissues (petals, style-stigma plus stamens, and ovaries) of detached buds and open flowers. This was achieved through treatment with 0.1 mM 1-aminocyclopropane-1-carboxylic acid (ACC) solution, 500 nl l(-1) methylcyclopropene (1-MCP), and 0.1 mM ABA solution. Treatment with ACC and 1-MCP confirmed that flower senescence in hibiscus is ethylene dependent, and treatment with exogenous ABA suggested that ABA may play a role in this process. The 1-MCP impeded petal in-rolling and decreased ABA content in detached open flowers after 9 h. This was preceded by an earlier and sequential increase in ABA content in 1-MCP-treated petals and style-stigma plus stamens between 1 h and 6 h. ACC treatment markedly accelerated flower senescence and increased ethylene production after 6 h and 9 h, particularly in style-stigma plus stamens. Ethylene evolution was positively correlated in these floral tissues with the induction of the gene expression of ethylene biosynthetic and receptor genes. Finally, ABA negatively affected the ethylene biosynthetic pathway and tissue sensitivity in all flower tissues. Transcript abundance of HrsACS, HrsACO, HrsETR, and HrsERS was reduced by exogenous ABA treatment. This research underlines the regulatory effect of ABA on the ethylene biosynthetic and perception machinery at a physiological and molecular level when inhibitors or promoters of senescence are exogenously applied.

  2. Coupling of Physiological and Proteomic Analysis to Understand the Ethylene- and Chilling-Induced Kiwifruit Ripening Syndrome

    PubMed Central

    Minas, Ioannis S.; Tanou, Georgia; Karagiannis, Evangelos; Belghazi, Maya; Molassiotis, Athanassios

    2016-01-01

    Kiwifruit [Actinidia deliciosa (A. Chev.) C.F. Liang et A.R. Ferguson, cv. “Hayward”] is classified as climacteric fruit and the initiation of endogenous ethylene production following harvest is induced by exogenous ethylene or chilling exposure. To understand the biological basis of this “dilemma,” kiwifruit ripening responses were characterized at 20°C following treatments with exogenous ethylene (100 μL L−1, 20°C, 24 h) or/and chilling temperature (0°C, 10 days). All treatments elicited kiwifruit ripening and induced softening and endogenous ethylene biosynthesis, as determined by 1-aminocyclopropane-1-carboxylic acid (ACC) content and ACC synthase (ACS) and ACC oxidase (ACO) enzyme activities after 10 days of ripening at 20°C. Comparative proteomic analysis using two-dimensional gel electrophoresis (2DE-PAGE) and nanoscale liquid chromatography coupled to tandem mass spectrometry (nanoLC-MS/MS) revealed 81 kiwifruit proteins associated with ripening. Thirty-one kiwifruit proteins were identified as commonly regulated by the three treatments accompanied by dynamic changes of 10 proteins specific to exogenous ethylene, 2 to chilling treatment, and 12 to their combination. Ethylene and/or chilling-responsive proteins were mainly involved in disease/defense, energy, protein destination/storage, and cell structure/cell wall. Interactions between the identified proteins were demonstrated by bioinformatics analysis, allowing a more complete insight into biological pathways and molecular functions affected by ripening. The present approach provides a quantitative basis for understanding the ethylene- and chilling-induced kiwifruit ripening and climacteric fruit ripening in general. PMID:26913040

  3. Hormonal changes in relation to biomass partitioning and shoot growth impairment in salinized tomato (Solanum lycopersicum L.) plants.

    PubMed

    Albacete, Alfonso; Ghanem, Michel Edmond; Martínez-Andújar, Cristina; Acosta, Manuel; Sánchez-Bravo, José; Martínez, Vicente; Lutts, Stanley; Dodd, Ian C; Pérez-Alfocea, Francisco

    2008-01-01

    Following exposure to salinity, the root/shoot ratio is increased (an important adaptive response) due to the rapid inhibition of shoot growth (which limits plant productivity) while root growth is maintained. Both processes may be regulated by changes in plant hormone concentrations. Tomato plants (Solanum lycopersicum L. cv Moneymaker) were cultivated hydroponically for 3 weeks under high salinity (100 mM NaCl) and five major plant hormones (abscisic acid, ABA; the cytokinins zeatin, Z, and zeatin-riboside, ZR; the auxin indole-3-acetic acid, IAA; and the ethylene precursor 1-aminocyclopropane-1-carboxylic acid, ACC) were determined weekly in roots, xylem sap, and leaves. Salinity reduced shoot biomass by 50-60% and photosynthetic area by 20-25% both by decreasing leaf expansion and delaying leaf appearance, while root growth was less affected, thus increasing the root/shoot ratio. ABA and ACC concentrations strongly increased in roots, xylem sap, and leaves after 1 d (ABA) and 15 d (ACC) of salinization. By contrast, cytokinins and IAA were differentially affected in roots and shoots. Salinity dramatically decreased the Z+ZR content of the plant, and induced the conversion of ZR into Z, especially in the roots, which accounted for the relative increase of cytokinins in the roots compared to the leaf. IAA concentration was also strongly decreased in the leaves while it accumulated in the roots. Decreased cytokinin content and its transport from the root to the shoot were probably induced by the basipetal transport of auxin from the shoot to the root. The auxin/cytokinin ratio in the leaves and roots may explain both the salinity-induced decrease in shoot vigour (leaf growth and leaf number) and the shift in biomass allocation to the roots, in agreement with changes in the activity of the sink-related enzyme cell wall invertase.

  4. Hormonal changes in relation to biomass partitioning and shoot growth impairment in salinized tomato (Solanum lycopersicum L.) plants

    PubMed Central

    Albacete, Alfonso; Ghanem, Michel Edmond; Martínez-Andújar, Cristina; Acosta, Manuel; Sánchez-Bravo, José; Martínez, Vicente; Lutts, Stanley; Dodd, Ian C.; Pérez-Alfocea, Francisco

    2008-01-01

    Following exposure to salinity, the root/shoot ratio is increased (an important adaptive response) due to the rapid inhibition of shoot growth (which limits plant productivity) while root growth is maintained. Both processes may be regulated by changes in plant hormone concentrations. Tomato plants (Solanum lycopersicum L. cv Moneymaker) were cultivated hydroponically for 3 weeks under high salinity (100 mM NaCl) and five major plant hormones (abscisic acid, ABA; the cytokinins zeatin, Z, and zeatin-riboside, ZR; the auxin indole-3-acetic acid, IAA; and the ethylene precursor 1-aminocyclopropane-1-carboxylic acid, ACC) were determined weekly in roots, xylem sap, and leaves. Salinity reduced shoot biomass by 50–60% and photosynthetic area by 20–25% both by decreasing leaf expansion and delaying leaf appearance, while root growth was less affected, thus increasing the root/shoot ratio. ABA and ACC concentrations strongly increased in roots, xylem sap, and leaves after 1 d (ABA) and 15 d (ACC) of salinization. By contrast, cytokinins and IAA were differentially affected in roots and shoots. Salinity dramatically decreased the Z+ZR content of the plant, and induced the conversion of ZR into Z, especially in the roots, which accounted for the relative increase of cytokinins in the roots compared to the leaf. IAA concentration was also strongly decreased in the leaves while it accumulated in the roots. Decreased cytokinin content and its transport from the root to the shoot were probably induced by the basipetal transport of auxin from the shoot to the root. The auxin/cytokinin ratio in the leaves and roots may explain both the salinity-induced decrease in shoot vigour (leaf growth and leaf number) and the shift in biomass allocation to the roots, in agreement with changes in the activity of the sink-related enzyme cell wall invertase. PMID:19036841

  5. Endogenous ethylene does not regulate opening of unstressed Iris flowers but strongly inhibits it in water-stressed flowers.

    PubMed

    Çelikel, Fisun G; van Doorn, Wouter G

    2012-09-15

    The floral buds of Iris flowers (Iris x hollandica) are enclosed by two sheath leaves. Flower opening depends on lifting the flower up to a position whereby the tepals can move laterally. This upward movement is carried out by elongation of the subtending pedicel and ovary. In the pedicels and ovaries of unstressed control flowers, the concentration of ACC (1-aminocyclopropane-1-carboxylic acid) and the rate of ethylene production increased during d 0-1 of flower opening, and then decreased. Exposure to ≥200 nL L(-1) ethylene for 24 h at 20°C inhibited elongation of the pedicel+ovary, and inhibited flower opening. However, pulsing of unstressed flowers with solutions containing inhibitors of ethylene synthesis (AOA, AVG), or an inhibitor of ethylene action (STS), did not affect pedicel+ovary elongation or flower opening. When the flowers were dehydrated for 2 d at 20°C and 60% RH, they did not open when subsequently placed in water, and showed inhibited elongation in the pedicel+ovary. This dehydration treatment resulted in elevated pedicel+ovary ACC levels and in increased ethylene production. Treatment with STS prevented the increase in ACC levels and ethylene production, overcame the effect of dehydration on elongation of the pedicel+ovary, and resulted in full flower opening. It is concluded that flower opening in unstressed Iris flowers is not regulated by endogenous ethylene. An increase in endogenous ethylene above normal levels during stress, by contrast, strongly inhibited flower opening, due to its inhibitory effect on elongation of the pedicel+ovary.

  6. Exogenous ethylene influences flower opening of cut roses (Rosa hybrida) by regulating the genes encoding ethylene biosynthesis enzymes.

    PubMed

    Ma, Nan; Cai, Lei; Lu, Wangjin; Tan, Hui; Gao, Junping

    2005-10-01

    The purpose of this paper is to investigate the differential responses of flower opening to ethylene in two cut rose cultivars, 'Samantha', whose opening process is promoted, and 'Kardinal', whose opening process is inhibited by ethylene. Ethylene production and 1-aminocyclopropane-1-carboxylate (ACC) synthase and oxidase activities were determined first. After ethylene treatment, ethylene production, ACC synthase (ACS) and ACC oxidase (ACO) activities in petals increased and peaked at the earlier stage (stage 3) in 'Samantha', and they were much more dramatically enhanced and peaked at the later stage (stage 4) in 'Kardinal' than control during vasing. cDNA fragments of three Rh-ACSs and one Rh-ACO genes were cloned and designated as Rh-ACS1, Rh-ACS2, Rh-ACS3 and Rh-ACO1 respectively. Northern blotting analysis revealed that, among three genes of ACS, ethylene-in- duced expression patterns of Rh-ACS3 gene corresponded to ACS activity and ethylene production in both cultivars. A more dramatic accumulation of Rh-ACS3 mRNA was induced by ethylene in 'Kardinal' than that of 'Samantha'. As an ethylene action inhibitor, STS at concentration of 0.2 mmol/L generally inhibited the expression of Rh-ACSs and Rh-ACO in both cultivars, although it induced the expression of Rh-ACS3 transiently in 'Kardinal'. Our results suggests that 'Kardinal' is more sensitive to ethylene than 'Samantha'; and the changes of Rh-ACS3 expression caused by ethylene might be related to the acceleration of flower opening in 'Samantha' and the inhibition in 'Kardinal'. Additional results indicated that three Rh-ACSs genes were differentially associated with flower opening and senescence as well as wounding

  7. Arabidopsis Hexokinase-Like1 and Hexokinase1 Form a Critical Node in Mediating Plant Glucose and Ethylene Responses

    SciTech Connect

    Karve, Abhijit A; Xioxia, Xia; Moore, Brandon

    2012-01-01

    Arabidopsis (Arabidopsis thaliana) hexokinase-like1 (HKL1) lacks Glc phosphorylation activity and has been shown to act as a negative regulator of plant growth. Interestingly, the protein has a largely conserved Glc binding domain and protein overexpression was shown previously to promote seedling tolerance to exogenous 6% (w/v) Glc. Since these phenotypes occur independently of cellular Glc signaling activities, we have tested whether HKL1 might promote crosstalk between the normal antagonists Glc and ethylene. We show that repression by 1-aminocyclopropane-1-carboxylic acid (ACC) of the Glc-dependent developmental arrest of wild-type Arabidopsis seedlings requires the HKL1 protein. We also describe an unusual root hair phenotype associated with growth on high Glc media that occurs prominently in HKL1 overexpression lines and in gin2-1, a null mutant of hexokinase1 (HXK1). Seedlings of these lines produce bulbous root hairs with an enlarged base, after transfer from agar plates with normal media to plates with 6% Glc. Seedling transfer to plates with 2% Glc plus ACC mimics the high Glc affect in the HKL1 overexpression line, but not in gin2-1. A similar ACC-stimulated, bulbous root hair phenotype also was observed in wild-type seedlings transferred to plates with 9% Glc. From transcript expression analyses, we found that HKL1 and HXK1 have differential roles in Glc-dependent repression of some ethylene biosynthesis genes. Since we show by co-immunoprecipitation assays that HKL1 and HXK1 can interact, these two proteins likely form a critical node in Glc signaling that mediates overlapping, but also distinct cellular responses to Glc and ethylene treatments.

  8. Effects of abscisic acid on ethylene biosynthesis and perception in Hibiscus rosa-sinensis L. flower development

    PubMed Central

    Trivellini, Alice; Ferrante, Antonio; Vernieri, Paolo; Serra, Giovanni

    2011-01-01

    The effect of the complex relationship between ethylene and abscisic acid (ABA) on flower development and senescence in Hibiscus rosa-sinensis L. was investigated. Ethylene biosynthetic (HrsACS and HrsACO) and receptor (HrsETR and HrsERS) genes were isolated and their expression evaluated in three different floral tissues (petals, style–stigma plus stamens, and ovaries) of detached buds and open flowers. This was achieved through treatment with 0.1 mM 1-aminocyclopropane-1-carboxylic acid (ACC) solution, 500 nl l−1 methylcyclopropene (1-MCP), and 0.1 mM ABA solution. Treatment with ACC and 1-MCP confirmed that flower senescence in hibiscus is ethylene dependent, and treatment with exogenous ABA suggested that ABA may play a role in this process. The 1-MCP impeded petal in-rolling and decreased ABA content in detached open flowers after 9 h. This was preceded by an earlier and sequential increase in ABA content in 1-MCP-treated petals and style–stigma plus stamens between 1 h and 6 h. ACC treatment markedly accelerated flower senescence and increased ethylene production after 6 h and 9 h, particularly in style–stigma plus stamens. Ethylene evolution was positively correlated in these floral tissues with the induction of the gene expression of ethylene biosynthetic and receptor genes. Finally, ABA negatively affected the ethylene biosynthetic pathway and tissue sensitivity in all flower tissues. Transcript abundance of HrsACS, HrsACO, HrsETR, and HrsERS was reduced by exogenous ABA treatment. This research underlines the regulatory effect of ABA on the ethylene biosynthetic and perception machinery at a physiological and molecular level when inhibitors or promoters of senescence are exogenously applied. PMID:21841180

  9. Methyl jasmonate-induced ethylene production is responsible for conifer phloem defense responses and reprogramming of stem cambial zone for traumatic resin duct formation.

    PubMed

    Hudgins, J W; Franceschi, Vincent R

    2004-08-01

    Conifer stem pest resistance includes constitutive defenses that discourage invasion and inducible defenses, including phenolic and terpenoid resin synthesis. Recently, methyl jasmonate (MJ) was shown to induce conifer resin and phenolic defenses; however, it is not known if MJ is the direct effector or if there is a downstream signal. Exogenous applications of MJ, methyl salicylate, and ethylene were used to assess inducible defense signaling mechanisms in conifer stems. MJ and ethylene but not methyl salicylate caused enhanced phenolic synthesis in polyphenolic parenchyma cells, early sclereid lignification, and reprogramming of the cambial zone to form traumatic resin ducts in Pseudotsuga menziesii and Sequoiadendron giganteum. Similar responses in internodes above and below treated internodes indicate transport of a signal giving a systemic response. Studies focusing on P. menziesii showed MJ induced ethylene production earlier and 77-fold higher than wounding. Ethylene production was also induced in internodes above the MJ-treated internode. Pretreatment of P. menziesii stems with the ethylene response inhibitor 1-methylcyclopropene inhibited MJ and wound responses. Wounding increased 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase protein, but MJ treatment produced a higher and more rapid ACC oxidase increase. ACC oxidase was most abundant in ray parenchyma cells, followed by cambial zone cells and resin duct epithelia. The data show these MJ-induced defense responses are mediated by ethylene. The cambial zone xylem mother cells are reprogrammed to differentiate into resin-secreting epithelial cells by an MJ-induced ethylene burst, whereas polyphenolic parenchyma cells are activated to increase polyphenol production. The results also indicate a central role of ray parenchyma in ethylene-induced defense.

  10. Methyl Jasmonate-Induced Ethylene Production Is Responsible for Conifer Phloem Defense Responses and Reprogramming of Stem Cambial Zone for Traumatic Resin Duct Formation

    PubMed Central

    Hudgins, J.W.; Franceschi, Vincent R.

    2004-01-01

    Conifer stem pest resistance includes constitutive defenses that discourage invasion and inducible defenses, including phenolic and terpenoid resin synthesis. Recently, methyl jasmonate (MJ) was shown to induce conifer resin and phenolic defenses; however, it is not known if MJ is the direct effector or if there is a downstream signal. Exogenous applications of MJ, methyl salicylate, and ethylene were used to assess inducible defense signaling mechanisms in conifer stems. MJ and ethylene but not methyl salicylate caused enhanced phenolic synthesis in polyphenolic parenchyma cells, early sclereid lignification, and reprogramming of the cambial zone to form traumatic resin ducts in Pseudotsuga menziesii and Sequoiadendron giganteum. Similar responses in internodes above and below treated internodes indicate transport of a signal giving a systemic response. Studies focusing on P. menziesii showed MJ induced ethylene production earlier and 77-fold higher than wounding. Ethylene production was also induced in internodes above the MJ-treated internode. Pretreatment of P. menziesii stems with the ethylene response inhibitor 1-methylcyclopropene inhibited MJ and wound responses. Wounding increased 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase protein, but MJ treatment produced a higher and more rapid ACC oxidase increase. ACC oxidase was most abundant in ray parenchyma cells, followed by cambial zone cells and resin duct epithelia. The data show these MJ-induced defense responses are mediated by ethylene. The cambial zone xylem mother cells are reprogrammed to differentiate into resin-secreting epithelial cells by an MJ-induced ethylene burst, whereas polyphenolic parenchyma cells are activated to increase polyphenol production. The results also indicate a central role of ray parenchyma in ethylene-induced defense. PMID:15299142

  11. Shedding light on ethylene metabolism in higher plants

    PubMed Central

    Rodrigues, Maria A.; Bianchetti, Ricardo E.; Freschi, Luciano

    2014-01-01

    Ethylene metabolism in higher plants is regulated by a wide array of endogenous and environmental factors. During most physiological processes, ethylene levels are mainly determined by a strict control of the rate-limiting biosynthetic steps responsible for the production of 1-aminocyclopropane-1-carboxylic acid (ACC) and its subsequent conversion to ethylene. Responsible for these reactions, the key enzymes ACC synthase and ACC oxidase are encoded by multigene families formed by members that can be differentially regulated at the transcription and post-translational levels by specific developmental and environmental signals. Among the wide variety of environmental cues controlling plant ethylene production, light quality, duration, and intensity have consistently been demonstrated to influence the metabolism of this plant hormone in diverse plant tissues, organs, and species. Although still not completely elucidated, the mechanisms underlying the interaction between light signal transduction and ethylene evolution appears to involve a complex network that includes central transcription factors connecting multiple signaling pathways, which can be reciprocally modulated by ethylene itself, other phytohormones, and specific light wavelengths. Accumulating evidence has indicated particular photoreceptors as essential mediators in light-induced signaling cascades affecting ethylene levels. Therefore, this review specifically focuses on discussing the current knowledge of the potential molecular mechanisms implicated in the light-induced responses affecting ethylene metabolism during the regulation of developmental and metabolic plant responses. Besides presenting the state of the art in this research field, some overlooked mechanisms and future directions to elucidate the exact nature of the light–ethylene interplay in higher plants will also be compiled and discussed. PMID:25520728

  12. Effects of wintertime fasting and seasonal adaptation on AMPK and ACC in hypothalamus, adipose tissue and liver of the raccoon dog (Nyctereutes procyonoides).

    PubMed

    Kinnunen, Sanni; Mänttäri, Satu; Herzig, Karl-Heinz; Nieminen, Petteri; Mustonen, Anne-Mari; Saarela, Seppo

    2016-02-01

    The raccoon dog (Nyctereutes procyonoides) is a canid with autumnal fattening and passive wintering strategy. We examined the effects of wintertime fasting and seasonality on AMP-activated protein kinase (AMPK), a regulator of metabolism, and its target, acetyl-CoA carboxylase (ACC) on the species. Twelve farmed raccoon dogs (eleven females/one male) were divided into two groups: half were fasted for ten weeks in December-March (winter fasted) and the others were fed ad libitum (winter fed). A third group (autumn fed, eight females) was fed ad libitum and sampled in December. Total AMPK, ACC and their phosphorylated forms (pAMPK, pACC) were measured from hypothalamus, liver, intra-abdominal (iWAT) and subcutaneous white adipose tissues (sWAT). The fasted animals lost 32% and the fed 20% of their body mass. Hypothalamic AMPK expression was lower and pACC levels higher in the winter groups compared to the autumn fed group. Liver pAMPK was lower in the winter fasted group, with consistently decreased ACC and pACC. AMPK and pAMPK were down-regulated in sWAT and iWAT of both winter groups, with a parallel decline in pACC in sWAT. The responses of AMPK and ACC to fasting were dissimilar to the effects observed previously in non-seasonal mammals and hibernators. Differences between the winter fed and autumn fed groups indicate that the functions of AMPK and ACC could be regulated in a season-dependent manner. Furthermore, the distinctive effects of prolonged fasting and seasonal adaptation on AMPK-ACC pathway could contribute to the wintering strategy of the raccoon dog.

  13. Improving derivatization efficiency of BMAA utilizing AccQ-Tag in a complex cyanobacterial matrix.

    PubMed

    Eriksson, Johan; Jonasson, Sara; Papaefthimiou, Dimitra; Rasmussen, Ulla; Bergman, Birgitta

    2009-01-01

    Two different assays have been developed and used in order to investigate the optimal conditions for derivatization and detection of acid beta-N-methyl-amino-L-alanine (BMAA) in a cyanobacterial sample. BMAA was extracted from cyanobacterial cultures both from the cytosolic ("free") fraction and in the precipitated ("protein") fraction using a newly developed extraction scheme and the sample matrix was standardized according to protein concentration to ensure the highest possible derivative yield. A rapid and sensitive HPLC method for fluorescence detection of the non-protein amino acid BMAA in cyanobacteria, utilizing the Waters AccQ-Tag chemistry and Chromolith Performance RP-18e columns was developed. Using this new method and utilizing a different buffer system and column than that recommended by Waters, we decreased the time between injections by 75%. The limit of quantification was determined to be 12 nmol and limit of detection as 120 fmol. The linear range was in the range of 8.5 nmol-84 pmol. Accuracy and precision were well within FDA guidelines for bioanalysis.

  14. Anatomically shaped cranial collimation (ACC) for lateral cephalometric radiography: a technical report.

    PubMed

    Hoogeveen, R C; van der Stelt, P F; Berkhout, W E R

    2014-01-01

    Lateral cephalograms in orthodontic practice display an area cranial of the base of the skull that is not required for diagnostic evaluation. Attempts have been made to reduce the radiation dose to the patient using collimators combining the shielding of the areas above the base of the skull and below the mandible. These so-called "wedge-shaped" collimators have not become standard equipment in orthodontic offices, possibly because these collimators were not designed for today's combination panoramic-cephalometric imaging systems. It also may be that the anatomical variability of the area below the mandible makes this area unsuitable for standardized collimation. In addition, a wedge-shaped collimator shields the cervical vertebrae; therefore, assessment of skeletal maturation, which is based on the stage of development of the cervical vertebrae, cannot be performed. In this report, we describe our investigations into constructing a collimator to be attached to the cephalostat and shield the cranial area of the skull, while allowing the visualization of diagnostically relevant structures and markedly reducing the size of the irradiated area. The shape of the area shielded by this "anatomically shaped cranial collimator" (ACC) was based on mean measurements of cephalometric landmarks of 100 orthodontic patients. It appeared that this collimator reduced the area of irradiation by almost one-third without interfering with the imaging system or affecting the quality of the image. Further research is needed to validate the clinical efficacy of the collimator.

  15. A portable platform for accelerated PIC codes and its application to GPUs using OpenACC

    NASA Astrophysics Data System (ADS)

    Hariri, F.; Tran, T. M.; Jocksch, A.; Lanti, E.; Progsch, J.; Messmer, P.; Brunner, S.; Gheller, C.; Villard, L.

    2016-10-01

    We present a portable platform, called PIC_ENGINE, for accelerating Particle-In-Cell (PIC) codes on heterogeneous many-core architectures such as Graphic Processing Units (GPUs). The aim of this development is efficient simulations on future exascale systems by allowing different parallelization strategies depending on the application problem and the specific architecture. To this end, this platform contains the basic steps of the PIC algorithm and has been designed as a test bed for different algorithmic options and data structures. Among the architectures that this engine can explore, particular attention is given here to systems equipped with GPUs. The study demonstrates that our portable PIC implementation based on the OpenACC programming model can achieve performance closely matching theoretical predictions. Using the Cray XC30 system, Piz Daint, at the Swiss National Supercomputing Centre (CSCS), we show that PIC_ENGINE running on an NVIDIA Kepler K20X GPU can outperform the one on an Intel Sandy bridge 8-core CPU by a factor of 3.4.

  16. Adenosine deaminase activity level as a tool for diagnosing tuberculous pleural effusion.

    PubMed

    Khow-Ean, Nathapol; Booraphun, Suchart; Aekphachaisawat, Noppadol; Sawanyawisuth, Kittisak

    2013-07-04

    The yield for using a pleural fluid culture to diagnose tuberculous pleural effusion (TPE) is low. Adenosine deaminase activity (ADA) has been shown to have good diagnostic value for TPE. The ADA cutoff point for the diagnosis of TPE is unclear. We attempted to determine the ADA level cutoff point for diagnosing of TPE in Thailand, where tuberculosis is endemic. We reviewed the medical records of patients with newly diagnosed pleural effusion aged >15 years who had a pleural fluid ADAlevel and who underwent a pleural biopsy. The study period was from March 1, 2010 to January 31, 2011. The diagnoses of TPE and malignant pleural effusion (MPE) were based on pathological findings. The diagnostic cutoff level for using ADA to diagnose TPE was determined. Forty-eight patients met study criteria. Of those, 18 patients (37.5%) were diagnosed with TPE. The mean ADA level was significantly higher among patients in the TPE group than in the MPE group (38.2 vs 14.8 U/l, p < 0.001). The cutoff level of 17.5 U/l gave sensitivity, specificity, positive likelihood ratio, and negative likelihood ratio of 88.9%, 73.3%, 3.33, and 0.15, respectively. An ADA level >17.5 U/l had good diagnostic values among TPE patients in our study.

  17. Activation-induced cytidine deaminase (AID) is localized to subnuclear domains enriched in splicing factors

    SciTech Connect

    Hu, Yi Ericsson, Ida Doseth, Berit Liabakk, Nina B. Krokan, Hans E. Kavli, Bodil

    2014-03-10

    Activation-induced cytidine deaminase (AID) is the mutator enzyme in adaptive immunity. AID initiates the antibody diversification processes in activated B cells by deaminating cytosine to uracil in immunoglobulin genes. To some extent other genes are also targeted, which may lead to genome instability and B cell malignancy. Thus, it is crucial to understand its targeting and regulation mechanisms. AID is regulated at several levels including subcellular compartmentalization. However, the complex nuclear distribution and trafficking of AID has not been studied in detail previously. In this work, we examined the subnuclear localization of AID and its interaction partner CTNNBL1 and found that they associate with spliceosome-associated structures including Cajal bodies and nuclear speckles. Moreover, protein kinase A (PKA), which activates AID by phosphorylation at Ser38, is present together with AID in nuclear speckles. Importantly, we demonstrate that AID physically associates with the major spliceosome subunits (small nuclear ribonucleoproteins, snRNPs), as well as other essential splicing components, in addition to the transcription machinery. Based on our findings and the literature, we suggest a transcription-coupled splicing-associated model for AID targeting and activation. - Highlights: • AID and its interaction partner CTNNBL1 localize to Cajal bodies and nuclear speckles. • AID associates with its activating kinase PKA in nuclear speckles. • AID is linked to the splicing machinery in switching B-cells. • Our findings suggest a transcription-coupled splicing associated mechanism for AID targeting and activation.

  18. miR-181b negatively regulates activation-induced cytidine deaminase in B cells.

    PubMed

    de Yébenes, Virginia G; Belver, Laura; Pisano, David G; González, Susana; Villasante, Aranzazu; Croce, Carlo; He, Lin; Ramiro, Almudena R

    2008-09-29

    Activated B cells reshape their primary antibody repertoire after antigen encounter by two molecular mechanisms: somatic hypermutation (SHM) and class switch recombination (CSR). SHM and CSR are initiated by activation-induced cytidine deaminase (AID) through the deamination of cytosine residues on the immunoglobulin loci, which leads to the generation of DNA mutations or double-strand break intermediates. As a bystander effect, endogenous AID levels can also promote the generation of chromosome translocations, suggesting that the fine tuning of AID expression may be critical to restrict B cell lymphomagenesis. To determine whether microRNAs (miRNAs) play a role in the regulation of AID expression, we performed a functional screening of an miRNA library and identified miRNAs that regulate CSR. One such miRNA, miR-181b, impairs CSR when expressed in activated B cells, and results in the down-regulation of AID mRNA and protein levels. We found that the AID 3' untranslated region contains multiple putative binding sequences for miR-181b and that these sequences can be directly targeted by miR-181b. Overall, our results provide evidence for a new regulatory mechanism that restricts AID activity and can therefore be relevant to prevent B cell malignant transformation.

  19. Heme-Biosynthetic Porphobilinogen Deaminase Protects Aspergillus nidulans from Nitrosative Stress

    PubMed Central