bluuebunny/crossref_metadata_2025 huggingface dataset:
Efficient infrared electroabsorption with 1 V applied voltage swing using intersubband transitions
Authors: Holmström P. , Jänes P. , Ekenberg U. , Thylén L.
Published: November 2008

We have demonstrated efficient intersubband electroabsorption in InGaAs/InAlGaAs/InAlAs step quantum wells grown by metal-organic vapor-phase epitaxy. An absorption modulation of 6 dB (Δα=2300 cm−1) at λ∼5.7 μm due to Stark shift of the intersubband resonance was achieved at a low applied voltage swing of ±0.5 V in a multipass waveguide structure. The interface intermixing was estimated by comparing experimental and theoretical Stark shifts. It is predicted that the present material in a strongly confining surface plasmon waveguide can yield an electroabsorption modulator with a peak-to-peak voltage of Vpp=0.9 V and modulation speed of f3dB≈130 GHz.
Simulation of a plasmonic tip-terminated scanning nanowire waveguide for molecular imaging
Authors: Malcolm Nathan P. , Heltzel Alex J. , Sokolov Konstantin V. , Shi Li , Howell John R.
Published: November 2008

Finite difference time domain simulation reveals plasmon coupling and local field enhancement at the gap between the gold nanoparticle (NP) tip of a ZnO nanowire (NW) waveguide and a gold-coated substrate or a gold NP probe. The region of field enhancement is about three times smaller than the 100 nm diameter of the gold NP tip, making the NW waveguide grown on a transparent microcantilever well-suited for near field imaging of single molecules immobilized on a gold substrate or gold NP-labeled cell membranes with superior spatial resolution and signal to noise ratio.
Spray-deposited poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) top electrode for organic solar cells
Authors: Lim Yee-Fun , Lee Sungsoo , Herman David J. , Lloyd Matthew T. , Anthony John E. , Malliaras George G.
Published: November 2008

Spray deposition is emerging as an attractive low-cost and high throughput method for organic solar cell (OSC) fabrication. In this letter, we demonstrate that a highly conductive formulation of poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) can be spray deposited to form the top electrode for an OSC. An inverted solar cell fabricated in this way with a blend of poly(3-hexylthiophene) and [6,6]-phenyl-C61 butyric acid methyl ester as the active layer achieved a power conversion efficiency of 2.0% under AM 1.5 100 mW/cm2 illumination.
Low power density multihole cathode very-high-frequency plasma for mixed phase Si:H thin films
Authors: Jariwala C. , Chainani A. , Eguchi R. , Matsunami M. , Shin S. , Bhatt S. , Dalal V. , John P. I.
Published: November 2008

A low power density very-high-frequency (VHF) (55 MHz) H2 plasma in a capacitively coupled multihole-cathode (MHC) geometry is studied using Langmuir probe measurements. Radial profiles show a higher ion density (Ni) and lower electron temperature (Te) compared to a MHC 13.56 MHz H2 plasma. The Ni dependence on power indicates an Ohmic plasma, while Te is essentially constant. The MHC-VHF plasma is used to investigate mixed phase microcrystalline+amorphous (μc+a-) Si:H thin films at a substrate temperature of 60 °C. High-resolution photoemission suggests two types of Si, with concentrations in agreement with atomic force microscopy images showing ∼510±40 nm crystallites embedded in a-Si:H matrix. The results show that the low power density MHC-VHF plasma is a high-Ni Ohmic collisional plasma, suitable for low temperature deposition of μc+a-Si:H thin films.
Invisibility cloak without singularity
Authors: Jiang Wei Xiang , Cui Tie Jun , Yang Xin Mi , Cheng Qiang , Liu Ruopeng , Smith David R.
Published: November 2008

An elliptical invisible cloak is proposed using a coordinate transformation in the elliptical-cylindrical coordinate system, which crushes the cloaked object to a line segment instead of a point. The elliptical cloak is reduced to a nearly circular cloak if the elliptical focus becomes very small. The advantage of the proposed invisibility cloak is that none of the parameters is singular and the changing range of all parameters is relatively small.
Polarized emission from high quality microcavity based on active organic layered domains
Authors: Stelitano S. , De Luca G. , Savasta S. , Patané S.
Published: November 2008

We demonstrate a giant polarization splitting of the emission of a high quality monolithic microcavity with an embedded ultrathin organic tetrakis(4-methoxyphenyl)porphyrin layer. The usual employed mechanism, based on the mismatch between the center of the mirror stopband and the wavelength of the microcavity, accounts only for a small fraction of the observed splitting. Optical and atomic force microscopy measurements allowed us to attribute it to local crystalline molecular order. This opens the way for the design and optimization of highly polarized compact optical sources.
High electric tunability of relaxor ferroelectric Langmuir–Blodgett terpolymer films
Authors: Wang J. L. , Meng X. J. , Yuan S. Z. , Yang J. , Sun J. L. , Xu H. S. , Chu J. H.
Published: November 2008

Poly(vinylidene fluoride-trifluoroethylene-chlorofloroethylene) terpolymer films have been fabricated on Al-coated polyimide substrates using the Langmuir–Blodgett (LB) technology. The electrical properties of the films have been investigated from 150 to 350 K. The temperature of the dielectric maximum and the measured frequency obeys the Vogel–Fulcher [J. Am. Ceram. Soc. 8, 339 (1925); Phys. Z. 22, 645 (1921)] law. The electric tunabilities of the films at room temperature are 42.6% and 80% under 50 and 240 MV/m, respectively. The reasons for the high tunability are discussed based on the special microstructure of the LB films.
Benefits of winter legume cover crops require early sowing
Authors: Gselman A. , Kramberger B.
Published: November 2008

Winter cover crops are beneficial, especially legumes that can supply nitrogen (N) to the next crop. The purpose of this study, involving separate experiments carried out at 2 different locations in north-eastern Slovenia, was to determine the most appropriate sowing time (early, early autumn SD1; late, mid autumn SD2; very late, late autumn SD3) for winter legumes (Trifolium subterraneum L., T. incarnatum L., T. pratense L., and Vicia villosa Roth) for the optimal yield of beneficial dry matter and soil N cycling. The control treatment used Lolium multiflorum Lam. For legume cover crops in SD1, from 915.0 (T. subterraneum) to 2495.0 (V. villosa) kg herbage dry matter yield (HDMY)/ha, 52.3 (T. pratense) to 148.4 (T. incarnatum) kg accumulated N (AN)/ha, and 14.5 (T. pratense) to 114.5 (T. incarnatum) kg symbiotically fixed N (Nsymb)/ha was obtained to the end of autumn. Until the spring ploughing-in, which was before maize sowing, legume cover crops in SD1 yielded 1065.0 (T. subterraneum) to 4440.0 (T. incarnatum) kg HDMY/ha, 74.9 (T. subterraneum) to 193.0 (V. villosa) kg AN/ha, and 4.7 (T. subterraneum) to 179.0 (V. villosa) kg Nsymb/ha. All parameters in SD2 were significantly lower than in SD1, whereas the SD3 sowing was not suitable for the legumes. The benefits of legume winter cover crops with regard to symbiotic N fixation were achieved only by early sowing; however, the amount of soil mineral N in late autumn and in early spring was decreased under L. multiflorum more than under the legumes.
Tomato irrigation scheduling improved by using percent canopy cover and crop developmental stage
Authors: Cetin Oner , Uygan Demet , Boyaci Hasan
Published: November 2008

The aim of this study was to evaluate whether it is possible to conserve water and improve yield using canopy cover in order to adjust the amount of water needed for drip-irrigated tomatoes. The experiments were carried out using field-grown tomatoes (Lycopercion esculentum cv. Dual Large, F1) in Central Anatolia, Turkey from 2003 to 2005. The experimental design used split-plots in randomised blocks with 3 replications. The main plots in this study were: I1, constant Class A pan evaporation (Ep) (i.e. K = 1.0) throughout the irrigation season; I2, varying Ep proportion according to crop development stage (i.e. planting time to flowering stage 0.50; flowering stage to first harvesting 1.0; and first harvest to last harvest 0.75. In subplots, wetted area percentages or canopy cover were used. In K1, the wetted area percentage measured 90% of the experimental site. In K2, the wetted area percentage varied depending on the canopy cover. In K3, the percentage of wetted area was set to a value of 65% throughout the irrigation season. The maximum yield of 134.8 t/ha was obtained with the I2K2 treatment, with a seasonal irrigation requirement of 515 mm. A tomato yield of 122.8 t/ha was obtained under conditions of 552 mm seasonal irrigation water applied using the I1K2 treatment. Although 37 mm less water was used in the I2K2 treatment, the yield obtained from that treatment was higher by 12.0 t/ha than the I1K2 treatment. In addition, the yield with all other treatments was lower than the I2K2 treatment. Similarly, the maximum irrigation water-use efficiency (IWUE) of 26.2 kg/m3 was obtained with the I2K2 treatment. Thus, use of different coefficients of Ep varying with crop growth stages and percentage of canopy cover to estimate irrigation water requirement (i.e. irrigation scheduling) maximised the yield of tomatoes and improved the IWUE.
Strategies for developing a core collection of bladder clover (Trifolium spumosum L.) using ecological and agro-morphological data
Authors: Ghamkhar K. , Snowball R. , Wintle B. J. , Brown A. H. D.
Published: November 2008

Core collections are a way to improve the functioning of germplasm collections through systematic evaluation, exploitation, and improved management of the whole collection. The published sizes of cores have ranged from 10 to 30% of the collection. The first step in developing a core collection is usually stratification of the whole collection. We adopted three approaches in the process of core designation. First, the existing ecological data and newly acquired agro-morphological (evaluation) data were analysed to determine the optimal method for capturing the ecological and morphological diversity of accessions. Second, different selection strategies were compared to identify the most powerful analysis with the greatest likelihood of covering maximum diversity. Third, the correlation between the two datasets was investigated. The annual pasture legume, bladder clover (Trifolium spumosum L.), was used for this pilot study because 80% of the existing 398 accessions have near-complete ecological data. The study found that a two-step selection process using ecological data and stratified proportional strategy followed by combined datasets and maximising strategy best represents the whole collection of T. spumosum in the core.
Cell Wall Architecture of the Elongating Maize Coleoptile
Authors: Carpita Nicholas C. , Defernez Marianne , Findlay Kim , Wells Brian , Shoue Douglas A. , Catchpole Gareth , Wilson Reginald H. , McCann Maureen C.
Published: November 2008

The primary walls of grasses are composed of cellulose microfibrils, glucuronoarabinoxylans (GAXs), and mixed-linkage β-glucans, together with smaller amounts of xyloglucans, glucomannans, pectins, and a network of polyphenolic substances. Chemical imaging by Fourier transform infrared microspectroscopy revealed large differences in the distributions of many chemical species between different tissues of the maize (Zea mays) coleoptile. This was confirmed by chemical analyses of isolated outer epidermal tissues compared with mesophyll-enriched preparations. Glucomannans and esterified uronic acids were more abundant in the epidermis, whereas β-glucans were more abundant in the mesophyll cells. The localization of β-glucan was confirmed by immunocytochemistry in the electron microscope and quantitative biochemical assays. We used field emission scanning electron microscopy, infrared microspectroscopy, and biochemical characterization of sequentially extracted polymers to further characterize the cell wall architecture of the epidermis. Oxidation of the phenolic network followed by dilute NaOH extraction widened the pores of the wall substantially and permitted observation by scanning electron microscopy of up to six distinct microfibrillar lamellae. Sequential chemical extraction of specific polysaccharides together with enzymic digestion of β-glucans allowed us to distinguish two distinct domains in the grass primary wall. First, a β-glucan-enriched domain, coextensive with GAXs of low degrees of arabinosyl substitution and glucomannans, is tightly associated around microfibrils. Second, a GAX that is more highly substituted with arabinosyl residues and additional glucomannan provides an interstitial domain that interconnects the β-glucan-coated microfibrils. Implications for current models that attempt to explain the biochemical and biophysical mechanism of wall loosening during cell growth are discussed.
Expression of an Antimicrobial Peptide via the Chloroplast Genome to Control Phytopathogenic Bacteria and Fungi
Authors: DeGray Gerald , Rajasekaran Kanniah , Smith Franzine , Sanford John , Daniell Henry
Published: November 2008

The antimicrobial peptide MSI-99, an analog of magainin 2, was expressed via the chloroplast genome to obtain high levels of expression in transgenic tobacco (Nicotiana tabacum var. Petit Havana) plants. Polymerase chain reaction products and Southern blots confirmed integration of MSI-99 into the chloroplast genome and achievement of homoplasmy, whereas northern blots confirmed transcription. Contrary to previous predictions, accumulation of MSI-99 in transgenic chloroplasts did not affect normal growth and development of the transgenic plants. This may be due to differences in the lipid composition of plastid membranes compared with the membranes of susceptible target microbes. In vitro assays with protein extracts from T1 and T2 plants confirmed that MSI-99 was expressed at high levels to provide 88% (T1) and 96% (T2) inhibition of growth against Pseudomonas syringae pv tabaci, a major plant pathogen. When germinated in the absence of spectinomycin selection, leaf extracts from T2 generation plants showed 96% inhibition of growth against P. syringae pv tabaci. In addition, leaf extracts from transgenic plants (T1) inhibited the growth of pregerminated spores of three fungal species,Aspergillus flavus, Fusarium moniliforme, and Verticillium dahliae, by more than 95% compared with non-transformed control plant extracts. In planta assays with the bacterial pathogen P. syringae pv tabaciresulted in areas of necrosis around the point of inoculation in control leaves, whereas transformed leaves showed no signs of necrosis, demonstrating high-dose release of the peptide at the site of infection by chloroplast lysis. In planta assays with the fungal pathogen,Colletotrichum destructivum, showed necrotic anthracnose lesions in non-transformed control leaves, whereas transformed leaves showed no lesions. Genetically engineering crop plants for disease resistance via the chloroplast genome instead of the nuclear genome is desirable to achieve high levels of expression and to prevent pollen-mediated escape of transgenes.
Enhanced Levels of the Aroma and Flavor CompoundS-Linalool by Metabolic Engineering of the Terpenoid Pathway in Tomato Fruits
Authors: Lewinsohn Efraim , Schalechet Fernond , Wilkinson Jack , Matsui Kenji , Tadmor Yaakov , Nam Kyoung-Hee , Amar Orit , Lastochkin Elena , Larkov Olga , Ravid Uzi , Hiatt William , Gepstein Shimon , Pichersky Eran
Published: November 2008

The aromas of fruits, vegetables, and flowers are mixtures of volatile metabolites, often present in parts per billion levels or less. We show here that tomato (Lycopersicon esculentumMill.) plants transgenic for a heterologous Clarkia breweri S-linalool synthase (LIS) gene, under the control of the tomato late-ripening-specific E8promoter, synthesize and accumulate S-linalool and 8-hydroxylinalool in ripening fruits. Apart from the difference in volatiles, no other phenotypic alterations were noted, including the levels of other terpenoids such as γ- and α-tocopherols, lycopene, β-carotene, and lutein. Our studies indicate that it is possible to enhance the levels of monoterpenes in ripening fruits by metabolic engineering.
A Novel Protein Elicitor (PaNie) from Pythium aphanidermatum Induces Multiple Defense Responses in Carrot, Arabidopsis, and Tobacco
Authors: Veit Stefan , Wörle Jörg Manfred , Nürnberger Thorsten , Koch Wolfgang , Seitz Hanns Ulrich
Published: November 2008

A novel protein elicitor (PaNie234) fromPythium aphanidermatum (Edson) Fitzp. was purified, microsequenced, and the corresponding cDNA was cloned. The deduced amino acid sequence contains a putative eukaryotic secretion signal with a proteinase cleavage site. The heterologously expressed elicitor protein without the secretion signal of 21 amino acids (PaNie213) triggered programmed cell death and de novo formation of 4-hydroxybenzoic acid in cultured cells of carrot (Daucus carota). Programmed cell death was determined using the tetrazolium assay and DNA laddering. Infiltration of PaNie213 into the intercellular space of leaves of Arabidopsis (Columbia-0, wild type) resulted in necroses and deposition of callose on the cell walls of spongy parenchyma cells surrounding the necrotic mesophyll cells. Necroses were also formed in tobacco (Nicotiana tabacum cv Wisconsin W38, wild type) and tomato (Lycopersicon esculentum Mill.) but not in maize (Zea mays), oat (Avena sativa), andTradescantia zebrina (Bosse), indicating that monocotyledonous plants are unable to perceive the signal. The reactions observed after treatment with the purified PaNie213 were identical to responses measured after treatment with a crude elicitor preparation from the culture medium ofP. aphanidermatum, described previously. The availability of the pure protein offers the possibility to isolate the corresponding receptor and its connection to downstream signaling-inducing defense reactions.
The Arabidopsis Protein SHI Represses Gibberellin Responses in Arabidopsis and Barley
Authors: Fridborg Ingela , Kuusk Sandra , Robertson Masumi , Sundberg Eva
Published: November 2008

The current model of gibberellin (GA) signal transduction is based on a derepressible system and a number of candidate negative regulators have been identified in Arabidopsis. We previously have reported the identification of the Arabidopsis gene SHORT INTERNODES (SHI) that causes suppression of GA responses when constitutively activated. In this paper, we show by using reporter gene analysis that the SHI gene is expressed in young organs, e.g. shoot apices and root tips. The model predicts a suppressor of GA responses to be active in these tissues to prevent premature growth or development. To study the effect of SHI on GA signaling, we used a functional assay that measures effects of signaling components on a well-defined GA response; the up-regulation of α-amylase in barley (Hordeum vulgare) aleurones in response to GA treatment. We found that SHI was able to specifically block the activity of a high-isoelectric point α-amylase promoter following GA3 treatment, which further supports that SHI is a suppressor of GA responses. We have identified two putative loss-of-function insertion alleles of SHI and lines homozygous for either of the new alleles show no phenotypic deviations from wild type. Because SHI belongs to a gene family consisting of nine members, we suggest that SHI and theSHI-related genes are functionally redundant. We also show that a functional ERECTA allele is able to partly suppress the dwarfing effect of the shi gain-of-function mutation, suggesting that the erecta mutation harbored by the Landsberg erecta ecotype is an enhancer of theshi dwarf phenotype.
Different Mechanisms of Four Aluminum (Al)-Resistant Transgenes for Al Toxicity in Arabidopsis
Authors: Ezaki Bunichi , Katsuhara Maki , Kawamura Masako , Matsumoto Hideaki
Published: November 2008

We have characterized the mechanism of action of four transgenes (AtBCB [Arabidopsis blue copper-binding protein], parB [tobacco {Nicotiana tabacum} glutathione S-transferase],NtPox [tobacco peroxidase], and NtGDI1[tobacco GDP dissociation inhibitor]) that independently Al resistance on transgenic Arabidopsis. All four transgenic lines showed lower deposition of callose after Al treatment than the Landsberg erecta ecotype of Arabidopsis, confirming that the four genes function to ameliorate Al toxicity. Influx and efflux experiments of Al ions suggested that theAtBCB gene may suppress Al absorption, whereas expression of the NtGDI1 gene promotes a release of Al in the root tip region of Arabidopsis. The total enzyme activities of glutathione S-transferases or peroxidases in transgenic lines carrying either the parB or NtPoxgenes were significantly higher than in the Landsbergerecta ecotype of Arabidopsis, and these enzyme activities were maintained at higher levels during Al stress. Furthermore, lipid peroxidation caused by Al stress was repressed in these two transgenic lines, suggesting that overexpression of these two genes diminishes oxidative damage caused by Al stress. Al-treated roots of transgenic plants were also stained by 4′,6-diamino-2-phenylindole to monitor cell death caused by Al toxicity. The result suggested that cell death is repressed in the NtPox line. Analysis of F1 hybrids between the four transgenic lines suggests that more resistant transgenic plants can be constructed by combinations of these four genes.
External Ca2+ Is Essential for Chloroplast Movement Induced by Mechanical Stimulation But Not by Light Stimulation,
Authors: Sato Yoshikatsu , Wada Masamitsu , Kadota Akeo
Published: November 2008

In the fern Adiantum capillus-veneris, chloroplast movement is induced by mechanical stimulation as well as by light stimulation. Directional movement of both types depends on an actin-based motile system. To investigate the physiological relationship between mechanical and light signaling in the regulation of chloroplast movement, we examined the mechano-response of chloroplasts whose motility had been already restricted after photo-relocation. Chloroplast mechano-avoidance movement was induced under all of the photo-relocation conditions tested, indicating that mechano-specific signals generated by mechanical stimulation dominate over the light signals and reactivate the motility of chloroplasts. When the effects of external Ca2+ on the induction of mechano- and light responses were examined, strikingly different requirements of external Ca2+ were found for each. In medium without Ca2+, the mechano-response was suppressed but no effects were observed on photo-response. Mechano-relocation movement of chloroplasts was inhibited by 100 μmlanthanum (La3+), a plasma membrane calcium channel blocker, and by 10 μm gadolinium (Gd3+), a stretch-activated channel blocker. However, the same concentrations of these drugs did not affect the photo-relocation movement at all. These results suggest that the influx of external Ca2+ is crucial for the early signaling step of chloroplast mechano-relocation but not for that of photo-relocation. This is the first report showing the separation of signaling pathways in mechano- and photo-relocation of chloroplasts.
Diversity of Arabidopsis Genes Encoding Precursors for Phytosulfokine, a Peptide Growth Factor
Authors: Yang Heping , Matsubayashi Yoshikatsu , Nakamura Kenzo , Sakagami Youji
Published: November 2008

Phytosulfokine-α (PSK-α), a unique plant peptide growth factor, was originally isolated from conditioned medium of asparagus (Asparagus officinalis) mesophyll cell cultures. PSK-α has several biological activities including promoting plant cell proliferation. Four genes that encode precursors of PSK-α have been identified from Arabidopsis. Analysis of cDNAs for two of these,AtPSK2 and AtPSK3, shows that both of these genes consist of two exons and one intron. The predicted precursors have N-terminal signal peptides and only a single PSK-α sequence located close to their carboxyl termini. Both precursors contain dibasic processing sites flanking PSK, analogous to animal and yeast prohormones. Although the PSK domain including the sequence of PSK-α and three amino acids preceding it are perfectly conserved, the precursors bear very limited similarity among Arabidopsis and rice (Oryza sativa), suggesting a new level of diversity among polypeptides that are processed into the same signaling molecule in plants, a scenario not found in animals and yeast. Unnatural [serine-4]PSK-β was found to be secreted by transgenic Arabidopsis cells expressing a mutant of either AtPSK2 orAtPSK3 cDNAs, suggesting that both AtPSK2and AtPSK3 encode PSK-α precursors.AtPSK2 and AtPSK3 were expressed demonstrably not only in cultured cells but also in intact plants, suggesting that PSK-α may be essential for plant cell proliferation in vivo as well as in vitro. Overexpression of either precursor gene allowed the transgenic calli to grow twice as large as the controls. However, the transgenic cells expressing either antisense cDNA did not dramatically decrease mitogenic activity, suggesting that these two genes may act redundantly.
Photoperiod and Temperature Interactions Regulate Low-Temperature-Induced Gene Expression in Barley
Authors: Fowler D. Brian , Breton Ghislaı̀n , Limin Allen E. , Mahfoozi Siroos , Sarhan Fathey
Published: November 2008

Vernalization and photoperiod (PP) responses are developmental mechanisms that allow plants to synchronize their growth and reproductive cycles with the seasonal weather changes. Vernalization requirement has been shown to influence the length of time that low-temperature (LT)-induced genes are up-regulated when cereal species are exposed to acclimating temperatures. The objective of the present study was to determine whether expression of LT-inducedWcs and Wcor gene families is also developmentally regulated by PP response. The LT-tolerant, highly short-day (SD)-sensitive barley (Hordeum vulgare L. cv Dicktoo) was subjected to 8-h SD and 20-h long-day PPs at cold-acclimating temperatures over a period of 70 d. A delay in transition from the vegetative to the reproductive stage under SD resulted in an increased level and longer retention of LT tolerance. Similar WCS and WCOR protein homologs were expressed, but levels of expression were much higher in plants acclimated under SD, indicating that the poor LT tolerance of long-day plants was the result of an inability to maintain LT-induced genes in an up-regulated state. These observations indicate that the PP and vernalization genes influence the expression of LT-induced genes in cereals through separate pathways that eventually converge to activate genes controlling plant development. In both instances, the delay in the transition from the vegetative to the reproductive stage produces increased LT tolerance that is sustained for a longer period of time, indicating that the developmental genes determine the duration of expression of LT-induced structural genes.
Identification and Analysis of Arabidopsis Expressed Sequence Tags Characteristic of Non-Coding RNAs
Authors: MacIntosh Gustavo C. , Wilkerson Curtis , Green Pamela J.
Published: November 2008

Sequencing of the Arabidopsis genome has led to the identification of thousands of new putative genes based on the predicted proteins they encode. Genes encoding tRNAs, ribosomal RNAs, and small nucleolar RNAs have also been annotated; however, a potentially important class of genes has largely escaped previous annotation efforts. These genes correspond to RNAs that lack significant open reading frames and encode RNA as their final product. Accumulating evidence indicates that such “non-coding RNAs” (ncRNAs) can play critical roles in a wide range of cellular processes, including chromosomal silencing, transcriptional regulation, developmental control, and responses to stress. Approximately 15 putative Arabidopsis ncRNAs have been reported in the literature or have been annotated. Although several have homologs in other plant species, all appear to be plant specific, with the exception of signal recognition particle RNA. Conversely, none of the ncRNAs reported from yeast or animal systems have homologs in Arabidopsis or other plants. To identify additional genes that are likely to encode ncRNAs, we used computational tools to filter protein-coding genes from genes corresponding to 20,000 expressed sequence tag clones. Using this strategy, we identified 19 clones with characteristics of ncRNAs, nine putative peptide-coding RNAs with open reading frames smaller than 100 amino acids, and 11 that could not be differentiated between the two categories. Again, none of these clones had homologs outside the plant kingdom, suggesting that most Arabidopsis ncRNAs are likely plant specific. These data indicate that ncRNAs represent a significant and underdeveloped aspect of Arabidopsis genomics that deserves further study.
Molecular Cloning, Functional Characterization, and Subcellular Localization of Soybean Nodule Dihydrolipoamide Reductase,
Authors: Moran Jose F. , Sun Zhaohui , Sarath Gautam , Arredondo-Peter Raúl , James Euan K. , Becana Manuel , Klucas Robert V.
Published: November 2008

Nodule ferric leghemoglobin reductase (FLbR) and leaf dihydrolipoamide reductase (DLDH) belong to the same family of pyridine nucleotide-disulfide oxidoreductases. We report here the cloning, expression, and characterization of a second protein with FLbR activity, FLbR-2, from soybean (Glycine max) nodules. The cDNA is 1,779 bp in length and codes for a precursor protein comprising a 30-residue mitochondrial transit peptide and a 470-residue mature protein of 50 kD. The derived protein has considerable homology with soybean nodule FLbR-1 (93% identity) and pea (Pisum sativum) leaf mitochondria DLDH (89% identity). The cDNA encoding the mature protein was overexpressed in Escherichia coli. The recombinant enzyme showed K  mand k  cat values for ferric leghemoglobin that were very similar to those of DLDH. The transcripts of FLbR-2 were more abundant in stems and roots than in nodules and leaves. Immunoblots of nodule fractions revealed that an antibody raised against pea leaf DLDH cross-reacted with recombinant FLbR-2, native FLbR-2 of soybean nodule mitochondria, DLDH from bacteroids, and an unknown protein of approximately 70 kD localized in the nodule cytosol. Immunogold labeling was also observed in the mitochondria, cytosol, and bacteroids of soybean nodules. The similar biochemical, kinetic, and immunological properties, as well as the high amino acid sequence identity and mitochondrial localization, draw us to conclude that FLbR-2 is soybean DLDH.
Analysis of the Raffinose Family Oligosaccharide Pathway in Pea Seeds with Contrasting Carbohydrate Composition
Authors: Peterbauer Thomas , Lahuta Leslaw B. , Blöchl Andreas , Mucha Jan , Jones David A. , Hedley Cliff L. , Gòrecki Richard J. , Richter Andreas
Published: November 2008

Raffinose family oligosaccharides (RFOs) are synthesized by a set of galactosyltransferases, which sequentially add galactose units from galactinol to sucrose. The accumulation of RFOs was studied in maturing seeds of two pea (Pisum sativum) lines with contrasting RFO composition. Seeds of the line SD1 accumulated stachyose as the predominant RFO, whereas verbascose, the next higher homolog of stachyose, was almost absent. In seeds of the line RRRbRb, a high level of verbascose was accumulated alongside with stachyose. The increase in verbascose in developing RRRbRb seeds was associated with galactinol-dependent verbascose synthase activity. In addition, a galactinol-independent enzyme activity was detected, which catalyzed transfer of a galactose residue from one stachyose molecule to another. The two enzyme activities synthesizing verbascose showed an optimum at pH 7.0. Both activities were almost undetectable inSD1. Maximum activity of stachyose synthase was about 4-fold higher in RRRbRb compared with SD1, whereas the activities of galactinol synthase and raffinose synthase were only about 1.5-fold higher in RRRbRb. The levels of galactinol synthase and stachyose synthase activity were reflected by steady-state levels of corresponding mRNAs. We suggest that the accumulation of verbascose in RRRbRb was controlled by a coordinated up-regulation of the last steps of verbascose biosynthesis.
Endogenous Nod-Factor-Like Signal Molecules Promote Early Somatic Embryo Development in Norway Spruce
Authors: Dyachok Julia V. , Wiweger Malgorzata , Kenne Lennart , von Arnold Sara
Published: November 2008

Embryogenic cultures of Norway spruce (Picea abies) are composed of pro-embryogenic masses (PEMs) and somatic embryos of various developmental stages. Auxin is important for PEM formation and proliferation. In this report we show that depletion of auxin blocks PEM development and causes large-scale cell death. Extracts of the media conditioned by embryogenic cultures stimulate development of PEM aggregates in auxin-deficient cultures. Partial characterization of the conditioning factor shows that it is a lipophilic, low-molecular-weight molecule, which is sensitive to chitinase and contains GlcNAc residues. On the basis of this information, we propose that the factor is a lipophilic chitin oligosaccharide (LCO). The amount of LCO correlates to the developmental stages of PEMs and embryos, with the highest level in the media conditioned by developmentally blocked cultures. LCO is not present in nonembryogenic cultures. Cell death, induced by withdrawal of auxin, is suppressed by extra supply of endogenous LCO or Nod factor from Rhizobium sp. NGR234. The effect can be mimicked by a chitotetraose or chitinase from Streptomyces griseus. Taken together, our data suggest that endogenous LCO acts as a signal molecule stimulating PEM and early embryo development in Norway spruce.
Gravity-Stimulated Changes in Auxin and Invertase Gene Expression in Maize Pulvinal Cells
Authors: Long Joanne C. , Zhao Wei , Rashotte Aaron M. , Muday Gloria K. , Huber Steven C.
Published: November 2008

Maize (Zea mays) stem gravitropism involves differential elongation of cells within a highly specialized region, the stem internodal pulvinus. In the present study, we investigated factors that control gravitropic responses in this system. In the graviresponding pulvinus, hexose sugars (d-Glc andd-Fru) accumulated asymmetrically across the pulvinus. This correlated well with an asymmetric increase in acid invertase activity across the pulvinus. Northern analyses revealed asymmetric induction of one maize acid invertase gene, Ivr2, consistent with transcriptional regulation by gravistimulation. Several lines of evidence indicated that auxin redistribution, as a result of polar auxin transport, is necessary for gravity-stimulatedIvr2 transcript accumulation and differential cell elongation across the maize pulvinus. First, the auxin transport inhibitor, N-1-naphthylphthalamic acid, inhibited gravistimulated curvature and Ivr2 transcript accumulation. Second, a transient gradient of free indole-3-acetic acid (IAA) across the pulvinus was apparent shortly after initiation of gravistimulation. This temporarily free IAA gradient appears to be important for differential cell elongation and Ivr2transcript accumulation. This is based on the observation thatN-1-naphthylphthalamic acid will not inhibit gravitropic responses when applied to pulvinus tissue after the free IAA gradient peak has occurred. Third, IAA alone can stimulate Ivr2transcript accumulation in non-gravistimulated pulvini. The gravity- and IAA-stimulated increase in Ivr2 transcripts was sensitive to the protein synthesis inhibitor, cycloheximide. Based on these results, a two-phase model describing possible relationships between gravitropic curvature, IAA redistribution, andIvr2 expression is presented.
Flavone Glucoside Uptake into Barley Mesophyll and Arabidopsis Cell Culture Vacuoles. Energization Occurs by H+-Antiport and ATP-Binding Cassette-Type Mechanisms
Authors: Frangne Nathalie , Eggmann Thomas , Koblischke Carsten , Weissenböck Gottfried , Martinoia Enrico , Klein Markus
Published: November 2008

In many cases, secondary plant products accumulate in the large central vacuole of plant cells. However, the mechanisms involved in the transport of secondary compounds are only poorly understood. Here, we demonstrate that the transport mechanisms for the major barley (Hordeum vulgare) flavonoid saponarin (apigenin 6-C-glucosyl-7-O-glucoside) are different in various plant species: Uptake into barley vacuoles occurs via a proton antiport and is competitively inhibited by isovitexin (apigenin 6-C-glucoside), suggesting that both flavone glucosides are recognized by the same transporter. In contrast, the transport into vacuoles from Arabidopsis, which does not synthesize flavone glucosides, displays typical characteristics of ATP-binding cassette transporters. Transport of saponarin into vacuoles of both the species is saturable with a K  m of 50 to 100 μm. Furthermore, the uptake of saponarin into vacuoles from a barley mutant exhibiting a strongly reduced flavone glucoside biosynthesis is drastically decreased when compared with the parent variety. Thus, the barley vacuolar flavone glucoside/H+antiporter could be modulated by the availability of the substrate. We propose that different vacuolar transporters may be responsible for the sequestration of species-specific/endogenous and nonspecific/xenobiotic secondary compounds in planta.
Characterization of a Family of Arabidopsis Genes Related to Xyloglucan Fucosyltransferase1
Authors: Sarria Rodrigo , Wagner Tanya A. , O'Neill Malcolm A. , Faik Ahmed , Wilkerson Curtis G. , Keegstra Kenneth , Raikhel Natasha V.
Published: November 2008

To understand primary cell wall assembly in Arabidopsis, we have focused on identifying and characterizing enzymes involved in xyloglucan biosynthesis. Nine genes (AtFUT2–10) were identified that share between 47% and 62% amino acid similarity with the xyloglucan-specific fucosyltransferase AtFUT1. Reverse transcriptase-PCR analysis indicates that all these genes are expressed. Bioinformatic analysis predicts that these family members are fucosyltransferases, and we first hypothesized that some may also be involved in xyloglucan biosynthesis. AtFUT3,AtFUT4, and AtFUT5 were expressed in tobacco (Nicotiana tabacum L. cv BY2) suspension culture cells, and the resulting proteins did not transfer fucose (Fuc) from GDP-Fuc to tamarind xyloglucan. AtFUT3, AtFUT4, and AtFUT5 were overexpressed in Arabidopsis plants. Leaves of plants overexpressing AtFUT4 or AtFUT5 contained more Fuc than wild-type plants. Stems of plants overexpressing AtFUT4 or AtFUT5 contained more xylose, less arabinose, and less galactose than wild-type plants. We suggest that the AtFUT family is likely to include fucosyltransferases important for the synthesis of wall carbohydrates. A targeted analysis of isolated cell wall matrix components from plants altered in expression of these proteins will help determine their specificity and biological function.
Identification, Purification, and Characterization of a Thermally Stable Lipase from Rice Bran. A New Member of the (Phospho) Lipase Family
Authors: Bhardwaj Kanchan , Raju Aruna , Rajasekharan Ram
Published: November 2008

A thermally stable lipase (EC 3.1.1.3.) was first identified in rice (Oryza sativa) bran, and the enzyme was purified to homogeneity using octyl-Sepharose chromatography. The enzyme was purified to 7.6-fold with the final specific activity of 0.38 μmol min−1 mg−1 at 80°C using [9,10-3H]triolein as a substrate. The purified enzyme was found to be a glycoprotein of 9.4 kD. Enzyme showed a maximum activity at 80°C and at pH 11.0. The protein was biologically active and retained most of its secondary structure even at 90°C as judged by the enzymatic assays and far-ultraviolet circular dichroism spectroscopy, respectively. Differential scanning calorimetric studies indicated that the transition temperature was 76°C and enthalpy 1.3 × 105 Calorie mol−1 at this temperature. The purified lipase also exhibited phospholipase A2 activity. Colocalization of both the hydrolytic activities in reverse-phase high-performance liquid chromatography and isoelectric focusing showed that the dual activity was associated with a single protein. Further, a direct interaction between both the substrates and the purified protein was demonstrated by photoaffinity labeling, using chemically synthesized analogs of triolein and phosphatidylcholine (PC). Apparent K  m for triolein (6.71 mm) was higher than that for PC (1.02 mm). The enzyme preferentially hydrolyzed thesn-2 position of PC, whereas it apparently exhibited no positional specificity toward triacylglycerol. Diisopropyl fluorophosphate inhibited both lipase and phospholipase activities of the purified enzyme. This enzyme is a new member from plants in the family of lipases capable of hydrolyzing phospholipids.
Efficient Down-Regulation of the Major Vegetative Storage Protein Genes in Transgenic Soybean Does Not Compromise Plant Productivity
Authors: Staswick Paul E. , Zhang Zhanyuan , Clemente Thomas E. , Specht James E.
Published: November 2008

Soybean (Glycine max L. Merr.) contains two related and abundant proteins, VSPα and VSPβ, that have been called vegetative storage proteins (VSP) based on their pattern of accumulation, degradation, tissue localization, and other characteristics. To determine whether these proteins play a critical role in sequestering N and other nutrients during early plant development, a VspA antisense gene construct was used to create transgenic plants in which VSP expression was suppressed in leaves, flowers, and seed pods. Total VSP was reduced at least 50-fold due to a 100-fold reduction in VSPα and a 10-fold reduction in VSPβ. Transgenic lines were grown in replicated yield trials in the field in Nebraska during the summer of 1999 and seed harvested from the lines was analyzed for yield, protein, oil, and amino acid composition. No significant difference (α = 0.05) was found between down-regulated lines and controls for any of the traits tested. Young leaves of antisense plants grown in the greenhouse contained around 3% less soluble leaf protein than controls at the time of flowering. However, total leaf N did not vary. Withdrawing N from plants during seed fill did not alter final seed protein content of antisense lines compared with controls. These results indicate that the VSPs play little if any direct role in overall plant productivity under typical growth conditions. The lack of VSPs in antisense plants might be partially compensated for by increases in other proteins and/or non-protein N. The results also suggest that the VSPs could be genetically engineered or replaced without deleterious effects.
Oxygen Deficiency Responsive Gene Expression inChlamydomonas reinhardtii through a Copper-Sensing Signal Transduction Pathway
Authors: Quinn Jeanette M. , Eriksson Mats , Moseley Jeffrey L. , Merchant Sabeeha
Published: November 2008

Chlamydomonas reinhardtii activatesCpx1, Cyc6, and Crd1, encoding, respectively, coproporphyrinogen oxidase, cytochromec  6, and a novel di-iron enzyme when transferred to oxygen-deficient growth conditions. This response is physiologically relevant because C. reinhardtiiexperiences these growth conditions routinely, and furthermore, one of the target genes, Crd1, is functionally required for normal growth under oxygen-depleted conditions. The same genes are activated also in response to copper-deficiency through copper-response elements that function as target sites for a transcriptional activator. The core of the copper-response element, GTAC, is required also for the hypoxic response, as is a trans-acting locus, CRR1. Mercuric ions, which antagonize the copper-deficiency response, also antagonize the oxygen-deficiency response of these target genes. Taken together, these observations suggest that the oxygen- and copper-deficiency responses share signal transduction components. Nevertheless, whereas the copper-response element is sufficient for the nutritional copper response, the oxygen-deficiency response requires, in addition, a second cis-element, indicating that the response to oxygen depletion is not identical to the nutritional copper response. The distinction between the two responses is also supported by comparative analysis of the response of the target genes,Cyc6, Cpx1, and Crd1, to copper versus oxygen deficiency. A Crr1-independent pathway forHyd1 expression in oxygen-depleted C. reinhardtii demonstrates the existence of multiple oxygen/redox-responsive circuits in this model organism.
Antisense RNA-Mediated Suppression of Benzophenanthridine Alkaloid Biosynthesis in Transgenic Cell Cultures of California Poppy
Authors: Park Sang-Un , Yu Min , Facchini Peter J.
Published: November 2008

California poppy (Eschscholzia californicaCham.) cell cultures produce several benzophenanthridine alkaloids, such as sanguinarine, chelirubine, and macarpine, with potent pharmacological activity. Antisense constructs of genes encoding two enzymes involved in benzophenanthridine alkaloid biosynthesis, the berberine bridge enzyme (BBE) and N-methylcoclaurine 3′-hydroxylase (CYP80B1), were introduced separately into California poppy cell cultures. Transformed cell lines expressing antisenseBBE or antisense CYP80B1 constructs and displaying low levels of BBE or CYP80B1 mRNAs, respectively, showed reduced accumulation of benzophenanthridine alkaloids compared with control cultures transformed with a β-glucuronidase gene. Pathway intermediates were not detected in any of the transformed cell lines. The suppression of benzophenanthridine alkaloid biosynthesis using BBE or CYP80B1 antisense RNA constructs also reduced the growth rate of the cultures. Two-dimensional 1H-nuclear magnetic resonance and in vivo 15N-nuclear magnetic resonance spectroscopy showed no difference in the abundance of carbohydrate metabolites in the various transgenic cell lines. However, transformed cells with reduced benzophenanthridine alkaloid levels contained larger cellular pools of several amino acids including alanine, leucine, phenylalanine, threonine, and valine compared with controls. The relative abundance of tyrosine, from which benzophenanthridine alkaloids are derived, was less than 2-fold higher in antisense-suppressed cells relative to controls. These results show that alterations in the metabolic flux through benzophenanthridine alkaloid biosynthesis can affect the regulation of amino acid pools. These data provide new insight into the metabolic engineering of benzophenanthridine alkaloid pathways.
Electrophysiological Analysis of Cloned Cyclic Nucleotide-Gated Ion Channels
Authors: Leng Qiang , Mercier Richard W. , Hua Bao-Guang , Fromm Hillel , Berkowitz Gerald A.
Published: November 2008

Electrophysiological studies were conducted on the cloned plant cyclic nucleotide-gated ion channels AtCNGC2 and AtCNGC1 from Arabidopsis, and NtCBP4 from tobacco (Nicotiana tobacum). The nucleotide coding sequences for these proteins were expressed in Xenopus laevis oocytes or HEK 293 cells. Channel characteristics were evaluated using voltage clamp analysis of currents in the presence of cAMP. AtCNGC2 was demonstrated to conduct K+ and other monovalent cations, but exclude Na+; this conductivity profile is unique for any ion channel not possessing the amino acid sequence found in the selectivity filter of K+-selective ion channels. Application of cAMP evoked currents in membrane patches of oocytes injected with AtCNGC2 cRNA. Direct activation of the channel by cyclic nucleotide, demonstrated by application of cyclic nucleotide to patches of membranes expressing such channels, is a hallmark characteristic of this ion channel family. Voltage clamp studies (two-electrode configuration) demonstrated that AtCNGC1 and NtCBP4 are also cyclic nucleotide-gated channels. Addition of a lipophilic analog of cAMP to the perfusion bath of oocytes injected with NtCBP4 and AtCNGC1 cRNAs induced inward rectified, noninactivating K+currents.