bluuebunny/crossref_metadata_2025_split huggingface dataset:
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

Abstract
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

AbstractThe 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

Abstract
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

Abstract
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

Abstract
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

Abstract
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

Abstract
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

Abstract
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

Abstract
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

Abstract
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

AbstractRaffinose 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

AbstractEmbryogenic 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

AbstractMaize (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.