bluuebunny/crossref_metadata_2025 huggingface dataset:
Analysis of Sec22p in Endoplasmic Reticulum/Golgi Transport Reveals Cellular Redundancy in SNARE Protein Function
Authors: Liu Yiting , Barlowe Charles
Published: September 2002

Membrane-bound soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins form heteromeric complexes that are required for intracellular membrane fusion and are proposed to encode compartmental specificity. In yeast, the R-SNARE protein Sec22p acts in transport between the endoplasmic reticulum (ER) and Golgi compartments but is not essential for cell growth. Other SNARE proteins that function in association with Sec22p (i.e., Sed5p, Bos1p, and Bet1p) are essential, leading us to question how transport through the early secretory pathway is sustained in the absence of Sec22p. In wild-type strains, we show that Sec22p is directly required for fusion of ER-derived vesicles with Golgi acceptor membranes. Insec22Δ strains, Ykt6p, a related R-SNARE protein that operates in later stages of the secretory pathway, is up-regulated and functionally substitutes for Sec22p. In vivo combination of thesec22Δ mutation with a conditionalykt6-1 allele results in lethality, consistent with a redundant mechanism. Our data indicate that the requirements for specific SNARE proteins in intracellular membrane fusion are less stringent than appreciated and suggest that combinatorial mechanisms using both upstream-targeting elements and SNARE proteins are required to maintain an essential level of compartmental organization.
Characterization of a Novel Lipoxygenase-Independent Senescence Mechanism in Alstroemeria peruviana Floral Tissue
Authors: Leverentz Michael K. , Wagstaff Carol , Rogers Hilary J. , Stead Anthony D. , Chanasut Usawadee , Silkowski Helena , Thomas Brian , Weichert Heiko , Feussner Ivo , Griffiths Gareth
Published: September 2002

The role of lipoxygenase (lox) in senescence ofAlstroemeria peruviana flowers was investigated using a combination of in vitro assays and chemical profiling of the lipid oxidation products generated. Phospholipids and galactolipids were extensively degraded during senescence in both sepals and petals and the ratio of saturated/unsaturated fatty acids increased. Lox protein levels and enzymatic activity declined markedly after flower opening. Stereochemical analysis of lox products showed that 13-lox was the major activity present in both floral tissues and high levels of 13-keto fatty acids were also synthesized. Lipid hydroperoxides accumulated in sepals, but not in petals, and sepals also had a higher chlorophyll to carotenoid ratio that favors photooxidation of lipids. Loss of membrane semipermeability was coincident for both tissue types and was chronologically separated from lox activity that had declined by over 80% at the onset of electrolyte leakage. Thus, loss of membrane function was not related to lox activity or accumulation of lipid hydroperoxides per se and differs in these respects from other ethylene-insensitive floral tissues representing a novel pattern of flower senescence.
Differential Expression of a Metallothionein Gene during the Presymbiotic versus the Symbiotic Phase of an Arbuscular Mycorrhizal Fungus
Authors: Lanfranco Luisa , Bolchi Angelo , Ros Emanuele Cesale , Ottonello Simone , Bonfante Paola
Published: September 2002

A full-length cDNA encoding a metallothionein (MT)-like polypeptide, designated GmarMT1, was identified in an expressed sequence tag collection from germinated spores of the arbuscular mycorrhizal fungus Gigaspora margarita(BEG34). The GmarMT1 gene is composed of two exons separated by an 81-bp intron. It codes for a 65-amino acid polypeptide comprising a plant type 1 MT-like N-terminal domain and a C-terminal domain that is most closely related to an as-yet-uncharacterized fungal MT. As revealed by heterologous complementation assays in yeast,GmarMT1 encodes a functional polypeptide capable of conferring increased tolerance against Cd and Cu. TheGmarMT1 RNA is expressed in both presymbiotic spores and symbiotic mycelia, even in the absence of metal exposure, but is significantly less abundant in the latter stage. An opposite pattern was observed upon Cu exposure, which up-regulatedGmarMT1 expression in symbiotic mycelia but not in germinated spores. Together, these data provide the first evidence, to our knowledge, for the occurrence in an arbuscular mycorrhizal fungus of a structurally novel MT that is modulated in a metal and life cycle stage-dependent manner and may afford protection against heavy metals (and other types of stress) to both partners of the endomycorrhizal symbiosis.
Loss of Nuclear Gene Expression during the Phytochrome A-Mediated Far-Red Block of Greening Response
Authors: McCormac Alex C. , Terry Matthew J.
Published: September 2002

We have examined the expression of the HEMA1 gene, which encodes the key chlorophyll synthesis enzyme glutamyl-tRNA reductase, during the phytochrome A-mediated far-red light (FR) block of greening response in Arabidopsis. Our results demonstrate that the FR block of greening comprises two separate responses: a white light (WL) intensity-independent response that requires 3 d of FR and is associated with a loss of expression of the nuclear genesHEMA1 and Lhcb following the transfer to WL (transcriptionally coupled response) and a WL intensity-dependent response that is induced by 1 d of FR and is transcriptionally uncoupled. Both responses required phytochrome A. The transcriptionally uncoupled response correlated with a deregulation of tetrapyrrole synthesis and potential photooxidative damage and was inhibited by cytokinin. The transcriptionally coupled FR response was additive with the loss of expression following Norflurazon-induced photobleaching and was absent in the presence of sucrose or after lower fluence rate (1 μmol m−2 s−1) FR treatments. Both pathways leading to the loss of nuclear gene expression were inhibited by overexpression of NADPH:protochlorophyllide oxidoreductase, indicating a role for plastid signaling in the FR-mediated pathway. The significance of identifying a distinct phytochrome A-mediated plastid signaling pathway is discussed.
Plasmalemma Abscisic Acid Perception Leads to RAB18Expression via Phospholipase D Activation in Arabidopsis Suspension Cells
Authors: Hallouin Matthieu , Ghelis Thanos , Brault Mathias , Bardat Françoise , Cornel Daniel , Miginiac Emile , Rona Jean-Pierre , Sotta Bruno , Jeannette Emmanuelle
Published: September 2002

Abscisic acid (ABA) plays a key role in the control of stomatal aperture by regulating ion channel activities and water exchanges across the plasma membrane of guard cells. Changes in cytoplasmic calcium content and activation of anion and outward-rectifying K+ channels are among the earliest cellular responses to ABA in guard cells. In Arabidopsis suspension cells, we have demonstrated that outer plasmalemma perception of ABA triggered similar early events. Furthermore, a Ca2+influx and the activation of anion channels are part of the ABA-signaling pathway leading to the specific expression ofRAB18. Here, we determine whether phospholipases are involved in ABA-induced RAB18 expression. Phospholipase C is not implicated in this ABA pathway. Using a transphosphatidylation reaction, we show that ABA plasmalemma perception results in a transient stimulation of phospholipase D (PLD) activity, which is necessary for RAB18 expression. Further experiments showed that PLD activation was unlikely to be regulated by heterotrimeric G proteins. We also observed that ABA-dependent stimulation of PLD was necessary for the activation of plasma anion current. However, when ABA activation of plasma anion channels was inhibited, the ABA-dependent activation of PLD was unchanged. Thus, we conclude that in Arabidopsis suspension cells, ABA stimulation of PLD acts upstream from anion channels in the transduction pathway leading to RAB18 expression.
Sterol C-24 Methyltransferase Type 1 Controls the Flux of Carbon into Sterol Biosynthesis in Tobacco Seed
Authors: Holmberg Niklas , Harker Mark , Gibbard Carl L. , Wallace Andrew D. , Clayton John C. , Rawlins Sally , Hellyer Amanda , Safford Richard
Published: September 2002

The first committed step in the conversion of cycloartenol into Δ5 C24-alkyl sterols in plants is catalyzed by anS-adenosyl-methionine-dependent sterol-C24-methyltransferase type 1 (SMT1). We report the consequences of overexpressing SMT1 in tobacco (Nicotiana tabacum), under control of either the constitutive carnation etched ring virus promoter or the seed-specific Brassica napusacyl-carrier protein promoter, on sterol biosynthesis in seed tissue. Overexpression of SMT1 with either promoter increased the amount of total sterols in seed tissue by up to 44%. The sterol composition was also perturbed with levels of sitosterol increased by up to 50% and levels of isofucosterol and campesterol increased by up to 80%, whereas levels of cycloartenol and cholesterol were decreased by up to 53% and 34%, respectively. Concomitant with the enhanced SMT1 activity was an increase in endogenous 3-hydroxy-3-methylglutaryl coenzyme A reductase activity, from which one can speculate that reduced levels of cycloartenol feed back to up-regulate 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and thereby control the carbon flux into sterol biosynthesis. This potential regulatory role of SMT1 in seed sterol biosynthesis is discussed.
Probing in Vivo Metabolism by Stable Isotope Labeling of Storage Lipids and Proteins in Developing Brassica napusEmbryos
Authors: Schwender Jörg , Ohlrogge John B.
Published: September 2002

Developing embryos of Brassica napusaccumulate both triacylglycerols and proteins as major storage reserves. To evaluate metabolic fluxes during embryo development, we have established conditions for stable isotope labeling of cultured embryos under steady-state conditions. Sucrose supplied via the endosperm is considered to be the main carbon and energy source for seed metabolism. However, in addition to 220 to 270 mmcarbohydrates (sucrose, glucose, and fructose), analysis of endosperm liquid revealed up to 70 mm amino acids as well as 6 to 15 mm malic acid. Therefore, a labeling approach with multiple carbon sources is a precondition to quantitatively reflect fluxes of central carbon metabolism in developing embryos. Mid-cotyledon stageB. napus embryos were dissected from plants and cultured for 15 d on a complex liquid medium containing13C-labeled carbohydrates. The 13C enrichment of fatty acids and amino acids (after hydrolysis of the seed proteins) was determined by gas chromatography/mass spectrometry. Analysis of 13C isotope isomers of labeled fatty acids and plastid-derived amino acids indicated that direct glycolysis provides at least 90% of precursors of plastid acetyl-coenzyme A (CoA). Unlabeled amino acids, when added to the growth medium, did not reduce incorporation of 13C label into plastid-formed fatty acids, but substantially diluted 13C label in seed protein. Approximately 30% of carbon in seed protein was derived from exogenous amino acids and as a consequence, the use of amino acids as a carbon source may have significant influence on the total carbon and energy balance in seed metabolism. 13C label in the terminal acetate units of C20 and C22 fatty acids that derive from cytosolic acetyl-CoA was also significantly diluted by unlabeled amino acids. We conclude that cytosolic acetyl-CoA has a more complex biogenetic origin than plastidic acetyl-CoA. Malic acid in the growth medium did not dilute 13C label incorporation into fatty acids or proteins and can be ruled out as a source of carbon for the major storage components of B. napusembryos.
Two New Loci, PLEIADE and HYADE, Implicate Organ-Specific Regulation of Cytokinesis in Arabidopsis
Authors: Müller Sabine , Fuchs Esther , Ovecka Miroslav , Wysocka-Diller Joanna , Benfey Philip N. , Hauser Marie-Theres
Published: September 2002

In screens for regulators of root morphogenesis in Arabidopsis we isolated six new recessive mutants with irregular cell expansion. Complementation analyses placed the mutations in two loci, PLEIADE (PLE) andHYADE (HYA). Phenotypic analyses revealed multinucleated cells, cell wall stubs, and synchronized cell divisions in incompletely separated cells that are all characteristics of defective cytokinesis. These defects were pronounced in roots and undetectable in aerial organs. In addition, fertility and germination were not affected by the mutations. Thus, the alleles that we have isolated of PLE and HYA suggest that the genes may encode organ-specific components needed primarily during root development. Analysis of microtubule arrays during cell cycle inple and hya roots indicates that the presence of several synchronized nuclei influences the position of preprophase band, mitotic spindles, and phragmoplasts. The enhanced and synergistic phenotype ofPLE/ple.hya/hyaseedlings and double mutants point to a role of PLE andHYA in the same process. These mutants provide tools to elucidate the regulation of nuclear cytoskeletal interactions during cell division and cytokinesis.
A Strobilurin Fungicide Enhances the Resistance of Tobacco against Tobacco Mosaic Virus and Pseudomonas syringae pvtabaci
Authors: Herms Stefan , Seehaus Kai , Koehle Harald , Conrath Uwe
Published: September 2002

The strobilurin class of fungicides comprises a variety of synthetic plant-protecting compounds with broad-spectrum antifungal activity. In the present study, we demonstrate that a strobilurin fungicide, F 500 (Pyraclostrobin), enhances the resistance of tobacco (Nicotiana tabacum cv Xanthi nc) against infection by either tobacco mosaic virus (TMV) or the wildfire pathogenPseudomonas syringae pv tabaci. F 500 was also active at enhancing TMV resistance in NahGtransgenic tobacco plants unable to accumulate significant amounts of the endogenous inducer of enhanced disease resistance, salicylic acid (SA). This finding suggests that F 500 enhances TMV resistance in tobacco either by acting downstream of SA in the SA signaling mechanism or by functioning independently of SA. The latter assumption is the more likely because in infiltrated leaves, F 500 did not cause the accumulation of SA-inducible pathogenesis-related (PR)-1 proteins that often are used as conventional molecular markers for SA-induced disease resistance. However, accumulation of PR-1 proteins and the associated activation of the PR-1 genes were elicited upon TMV infection of tobacco leaves and both these responses were induced more rapidly in F 500-pretreated plants than in the water-pretreated controls. Taken together, our results suggest that F 500, in addition to exerting direct antifungal activity, may also protect plants by priming them for potentiated activation of subsequently pathogen-induced cellular defense responses.
An Endoplasmic Reticulum-Bound Ca2+/Mn2+Pump, ECA1, Supports Plant Growth and Confers Tolerance to Mn2+ Stress
Authors: Wu Zhongyi , Liang Feng , Hong Bimei , Young Jeff C. , Sussman Michael R. , Harper Jeffrey F. , Sze Heven
Published: September 2002

Plants can grow in soils containing highly variable amounts of mineral nutrients, like Ca2+ and Mn2+, though the mechanisms of adaptation are poorly understood. Here, we report the first genetic study to determine in vivo functions of a Ca2+ pump in plants. Homozygous mutants of Arabidopsis harboring a T-DNA disruption in ECA1 showed a 4-fold reduction in endoplasmic reticulum-type calcium pump activity. Surprisingly, the phenotype of mutant plants was indistinguishable from wild type when grown on standard nutrient medium containing 1.5 mmCa2+ and 50 μm Mn2+. However, mutants grew poorly on medium with low Ca2+ (0.2 mm) or high Mn2+ (0.5 mm). On high Mn2+, the mutants failed to elongate their root hairs, suggesting impairment in tip growth processes. Expression of the wild-type gene (CAMV35S::ECA1) reversed these conditional phenotypes. The activity of ECA1 was examined by expression in a yeast (Saccharomyces cerevisiae) mutant, K616, which harbors a deletion of its endogenous calcium pumps. In vitro assays demonstrated that Ca2+, Mn2+, and Zn2+stimulated formation of a phosphoenzyme intermediate, consistent with the translocation of these ions by the pump. ECA1 provided increased tolerance of yeast mutant to toxic levels of Mn2+ (1 mm) and Zn2+(3 mm), consistent with removal of these ions from the cytoplasm. These results show that despite the potential redundancy of multiple Ca2+ pumps and Ca2+/H+ antiporters in Arabidopsis, pumping of Ca2+ and Mn2+ by ECA1 into the endoplasmic reticulum is required to support plant growth under conditions of Ca2+ deficiency or Mn2+ toxicity.
Mitochondrial-Driven Bicarbonate Transport Supports Photosynthesis in a Marine Microalga
Authors: Huertas I. Emma , Colman Brian , Espie George S.
Published: September 2002

The CO2-concentrating mechanism (CCM) of the marine eustigmatophycean microalga Nannochloropsis gaditana consists of an active HCO3  −transport system and an internal carbonic anhydrase to facilitate accumulation and conversion of HCO3  − to CO2 for photosynthetic fixation. Aqueous inlet mass spectrometry revealed that a portion of the CO2 generated within the cells leaked to the medium, resulting in a significant rise in the extracellular CO2 concentration to a level above its chemical equilibrium that was diagnostic for active HCO3  − transport. The transient rise in extracellular CO2 occurred in the light and the dark and was resolved from concurrent respiratory CO2efflux using H13CO3  − stable isotope techniques. H13CO3  −pump-13CO2 leak activity of the CCM was unaffected by 10 μm3(3,4-dichlorophenyl)-1,1-dimethylurea, an inhibitor of chloroplast linear electron transport, although photosynthetic O2evolution was reduced by 90%. However, low concentrations of cyanide, azide, and rotenone along with anoxia significantly reduced or abolished 13CO2 efflux in the dark and light. These results indicate that H13CO3  − transport was supported by mitochondrial energy production in contrast to other algae and cyanobacteria in which it is supported by photosynthetic electron transport. This is the first report of a direct role for mitochondria in the energization and functioning of the CCM in a photosynthetic organism.
Effect of Regulated Overexpression of the MADS Domain Factor AGL15 on Flower Senescence and Fruit Maturation
Authors: Fang Su-Chiung , Fernandez Donna E.
Published: September 2002

We have examined the effect of regulated overexpression of AGL15, a member of the MADS domain family of regulatory factors, on reproductive tissues. Using molecular and physiological markers, we show that constitutive overexpression of AGL15 in Arabidopsis leads to delay and down-regulation of senescence programs in perianth organs and developing fruits and alters the process of seed desiccation. Through genetic crosses, we show that the rate of water loss in the maturing seeds is dictated by the genetic composition and physiological state of the maternal tissue, rather than the embryo. To define the developmental time and/or place when senescence programs are most affected by elevated AGL15 levels, we expressed AGL15 under the control of various promoters. Expression during senescence or in abscission zone cells did not produce delays in floral organ senescence or abscission. Using a glucocorticoid-inducible expression system, we show that an increase in AGL15 levels around the time of flower opening is necessary to delay senescence and increase floral organ longevity.
Early Embryo Development in Fucus distichusIs Auxin Sensitive
Authors: Basu Swati , Sun Haiguo , Brian Leigh , Quatrano Ralph L. , Muday Gloria K.
Published: September 2002

Auxin and polar auxin transport have been implicated in controlling embryo development in land plants. The goal of these studies was to determine if auxin and auxin transport are also important during the earliest stages of development in embryos of the brown alga Fucus distichus. Indole-3-acetic acid (IAA) was identified in F. distichus embryos and mature tissues by gas chromatography-mass spectroscopy. F. distichus embryos accumulate [3H]IAA and an inhibitor of IAA efflux, naphthylphthalamic acid (NPA), elevates IAA accumulation, suggesting the presence of an auxin efflux protein complex similar to that found in land plants. F. distichus embryos normally develop with a single unbranched rhizoid, but growth on IAA leads to formation of multiple rhizoids and growth on NPA leads to formation of embryos with branched rhizoids, at concentrations that are active in auxin accumulation assays. The effects of IAA and NPA are complete before 6 h after fertilization (AF), which is before rhizoid germination and cell division. The maximal effects of IAA and NPA are between 3.5 and 5 h AF and 4 and 5.5 h AF, respectively. Although, the location of the planes of cell division was significantly altered in NPA- and IAA-treated embryos, these abnormal divisions occurred after abnormal rhizoid initiation and branching was observed. The results of this study suggest that auxin acts in the formation of apical basal patterns inF. distichus embryo development.
Developmentally Regulated Dual-Specificity Kinase from Peanut That Is Induced by Abiotic Stresses
Authors: Rudrabhatla Parvathi , Rajasekharan Ram
Published: September 2002

Tyrosine (Tyr) phosphorylation represents an important biochemical mechanism to regulate many cellular processes. No Tyr kinase has been cloned so far in plants. Dual-specificity kinases are reported in plants and the function of these kinases remains unknown. A 1.7-kb cDNA that encodes serine/threonine/Tyr (STY) kinase was isolated by screening peanut (Arachis hypogaea) expression library using the anti-phospho-Tyr antibody. The histidine-tagged recombinant kinase histidine-6-STY predominantly autophosphorylated on Tyr and phosphorylated the histone primarily on threonine. Genomic DNA gel-blot analysis revealed that STY kinase is a member of a small multigene family. The transcript of STY kinase is accumulated in the mid-maturation stage of seed development, suggesting a role in the signaling of storage of seed reserves. The STY kinase mRNA expression, as well as kinase activity, markedly increased in response to cold and salt treatments; however, no change in the protein level was observed, suggesting a posttranslational activation mechanism. The activation of the STY kinase is detected after 12 to 48 h of cold and salt treatments, which indicates that the kinase may not participate in the initial response to abiotic stresses, but may play a possible role in the adaptive process to adverse conditions. The transcript levels and kinase activity were unaltered with abscisic acid treatment, suggesting an abscisic acid-independent cold and salt signaling pathway. Here, we report the first identification of a non-MAP kinase cascade dual-specificity kinase involved in abiotic stress and seed development.
Resistance of Cultivated Tomato to Cell Content-Feeding Herbivores Is Regulated by the Octadecanoid-Signaling Pathway
Authors: Li Chuanyou , Williams Mark M. , Loh Ying-Tsu , Lee Gyu In , Howe Gregg A.
Published: September 2002

The octadecanoid signaling pathway has been shown to play an important role in plant defense against various chewing insects and some pathogenic fungi. Here, we examined the interaction of a cell-content feeding arachnid herbivore, the two-spotted spider mite (Tetranychus urticae Koch), with cultivated tomato (Lycopersicon esculentum) and an isogenic mutant line (defenseless-1 [def-1]) that is deficient in the biosynthesis of the octadecanoid pathway-derived signal, jasmonic acid (JA). Spider mite feeding and fecundity ondef-1 plants was significantly greater than on wild-type plants. Decreased resistance of def-1 plants was correlated with reduced JA accumulation and expression of defensive proteinase inhibitor (PI) genes, which were induced in mite-damaged wild-type leaves. Treatment of def-1 plants with methyl-JA restored resistance to spider mite feeding and reduced the fecundity of female mites. Plants expressing a35S::prosystemin transgene that constitutively activates the octadecanoid pathway in a Def-1-dependent manner were highly resistant to attack by spider mites and western flower thrips (Frankliniella occidentalis), another cell-content feeder of economic importance. These findings indicate that activation of the octadecanoid signaling pathway promotes resistance of tomato to a broad spectrum of herbivores. The techniques of amplified fragment length polymorphism (AFLP) and bulk segregant analysis were used to map the Def-1 gene to a region on the long arm of chromosome 3 that is genetically separable from the map position of known JA biosynthetic genes. Tight linkage ofDef-1 to a T-DNA insertion harboring the maize (Zea mays) Dissociation transposable element suggests a strategy for directed transposon tagging of the gene.
Patterns of Expression and Normalized Levels of the Five Arabidopsis Phytochromes
Authors: Sharrock Robert A. , Clack Ted
Published: September 2002

Using monoclonal antibodies specific for each apoprotein and full-length purified apoprotein standards, the levels of the five Arabidopsis phytochromes and their patterns of expression in seedlings and mature plants and under different light conditions have been characterized. Phytochrome levels are normalized to the DNA content of the various tissue extracts to approximate normalization to the number of cells in the tissue. One phytochrome, phytochrome A, is highly light labile. The other four phytochromes are much more light stable, although among these, phytochromes B and C are reduced 4- to 5-fold in red- or white-light-grown seedlings compared with dark-grown seedlings. The total amount of extractable phytochrome is 23-fold lower in light-grown than dark-grown tissues, and the percent ratios of the five phytochromes, A:B:C:D:E, are measured as 85:10:2:1.5:1.5 in etiolated seedlings and 5:40:15:15:25 in seedlings grown in continuous white light. The four light-stable phytochromes are present at nearly unchanging levels throughout the course of development of mature rosette and reproductive-stage plants and are present in leaves, stems, roots, and flowers. Phytochrome protein expression patterns over the course of seed germination and under diurnal and circadian light cycles are also characterized. Little cycling in response to photoperiod is observed, and this very low amplitude cycling of some phytochrome proteins is out of phase with previously reported cycling ofPHY mRNA levels. These studies indicate that, with the exception of phytochrome A, the family of phytochrome photoreceptors in Arabidopsis constitutes a quite stable and very broadly distributed array of sensory molecules.
A Role for the DOF Transcription Factor BPBF in the Regulation of Gibberellin-Responsive Genes in Barley Aleurone
Authors: Mena Montaña , Cejudo Francisco Javier , Isabel-Lamoneda Ines , Carbonero Pilar
Published: September 2002

Functional analyses of a number of hydrolase gene promoters, induced by gibberellin (GA) in aleurone cells following germination, have identified a GA-responsive complex as a tripartite element containing a pyrimidine box motif 5′-CCTTTT-3′. We describe here that BPBF, a barley (Hordeum vulgare) transcription factor of the DOF (DNA-Binding with One Finger) class, previously shown to be an activator of reserve protein encoding genes during development, also has a role in the control of hydrolase genes following seed germination. Northern-blot, reverse transcriptase-polymerase chain reaction, and in situ hybridization analyses evidenced that the transcripts of the BPBF-encoding gene (Pbf), besides being present during endosperm development, are also expressed in aleurone cells of germinated seeds where they are induced by GA, an effect counteracted by abscisic acid. Electrophoretic mobility shift assays have shown that the BPBF protein binds specifically to the pyrimidine box motif in vitro within the different sequence contexts that naturally occur in the promoters of genes encoding a cathepsin B-like protease (Al21) and a low-isoelectric point α-amylase (Amy2/32b), both induced in the aleurone layers in response to GA. In transient expression experiments, BPBF repressed transcription of theAl21 promoter in GA-treated barley aleurone layers and reverted the GAMYB-mediated activation of this protease promoter.
The Bifunctional LKR/SDH Locus of Plants Also Encodes a Highly Active Monofunctional Lysine-Ketoglutarate Reductase Using a Polyadenylation Signal Located within an Intron,
Authors: Tang Guiliang , Zhu Xiaohong , Gakiere Bertrand , Levanony Hanna , Kahana Anat , Galili Gad
Published: September 2002

Both plants and animals catabolize lysine (Lys) via two consecutive enzymes, Lys-ketoglutarate reductase (LKR) and saccharopine dehydrogenase (SDH), which are linked on a single polypeptide encoded by a single LKR/SDHgene. We have previously shown that the ArabidopsisLKR/SDH gene also encodes a monofunctional SDH that is transcribed from an internal promoter. In the present report, we have identified two cDNAs derived from cotton (Gossypium hirsutum) boll abscission zone that encode a novel enzymatic form of Lys catabolism, i.e. a catabolic monofunctional LKR. The monofunctional LKR mRNA is also encoded by theLKR/SDH gene, using two weak polyadenylation sites located within an intron. In situ mRNA hybridization and quantitative reverse transcriptase-polymerase chain reaction analyses also suggest that the cotton monofunctional LKR is relatively abundantly expressed in parenchyma cells of the abscission zone. DNA sequence analysis of theLKR/SDH genes of Arabidopsis, maize (Zea mays), and tomato (Lycopersicon esculentum) suggests that these genes can also encode a monofunctional LKR mRNA by a similar mechanism. To test whether the LKR/SDH and monofunctional LKR enzymes possess different biochemical properties, we used recombinant Arabidopsis LKR/SDH and monofunctional LKR enzymes expressed in yeast (Saccharomyces cerevisiae) cells. The K  m of the monofunctional LKR to Lys was nearly 10-fold lower than its counterpart that is linked to SDH. Taken together, our results suggest that theLKR/SDH locus of plants is a super-composite locus that can encode three related but distinct enzymes of Lys catabolism. These three enzymes apparently operate in concert to finely regulate Lys catabolism during plant development.
Engineering Herbicide Metabolism in Tobacco and Arabidopsis with CYP76B1, a Cytochrome P450 Enzyme from Jerusalem Artichoke
Authors: Didierjean Luc , Gondet Laurence , Perkins Roberta , Lau Sze-Mei Cindy , Schaller Hubert , O'Keefe Daniel P. , Werck-Reichhart Danièle
Published: September 2002

The Jerusalem artichoke (Helianthus tuberosus) xenobiotic inducible cytochrome P450, CYP76B1, catalyzes rapid oxidative dealkylation of various phenylurea herbicides to yield nonphytotoxic metabolites. We have found that increased herbicide metabolism and tolerance can be achieved by ectopic constitutive expression of CYP76B1 in tobacco (Nicotiana tabacum) and Arabidopsis. Transformation with CYP76B1 conferred on tobacco and Arabidopsis a 20-fold increase in tolerance to linuron, a compound detoxified by a single dealkylation, and a 10-fold increase in tolerance to isoproturon or chlortoluron, which need successive catalytic steps for detoxification. Two constructs for expression of translational fusions of CYP76B1 with P450 reductase were prepared to test if they would yield even greater herbicide tolerance. Plants expressing these constructs had lower herbicide tolerance than CYP76B1 alone, which is apparently a consequence of reduced stability of the fusion proteins. In all cases, increased herbicide tolerance results from more extensive metabolism, as demonstrated with exogenously fed phenylurea. Beside increased herbicide tolerance, expression of CYP76B1 has no other visible phenotype in the transgenic plants. Our data indicate that CYP76B1 can function as a selectable marker for plant transformation, allowing efficient selection in vitro and in soil-grown plants. Plants expressing CYP76B1 may also be a potential tool for phytoremediation of contaminated sites.
Molecular Identification of Cytosolic, Patatin-Related Phospholipases A from Arabidopsis with Potential Functions in Plant Signal Transduction
Authors: Holk André , Rietz Steffen , Zahn Marc , Quader Hartmut , Scherer Günther F.E.
Published: September 2002

Rapid activation of phospholipase A (PLA) by auxin or plant-pathogen interaction suggests a function in signal transduction for this enzyme, but the molecular identification of a cytosolic PLA carrying out this function remains open. We isolated four cDNA sequences from Arabidopsis (ecotype Columbia), AtPLA I,AtPLA IIA, AtPLA IVA, andAtPLA IVC, which are members of the patatin-related PLA gene family in plants and which are homologous to the animal Ca2+-independent PLA2 gene family. Expression was measured by reverse transcriptase-polymerase chain reaction, andAtPLA I transcripts were found preferentially in shoots,AtPLA IIA and AtPLA IVA in roots, andAtPLA IVC in flowers. Transient expression of the four PLA-green fluorescent protein fusion proteins in tobacco (Nicotiana tabacum) leaves showed they were located in the cytosol and not in the vacuoles. Surprisingly,AtPLA::green fluorescent protein was also localized to chloroplasts. The enzymatic activity of the purified recombinant AtPLA IVA toward phosphatidylcholine was dependent on Ca2+, saturated at 0.5 mm, and had a pH optimum of about 7.0. It had both PLA1 and PLA2 specificity. The enzyme showed in vitro highest sensitivity toward the PLA2 inhibitors palmitoyltrifluoromethyl ketone (PACOCF3,K  i approximately 30 nm), arachidonyltrifluoromethyl ketone (AACOCF3,K  i approximately 25 μm), and tetrahydro-3-(1-naphtalenyl)-2H-pyran-2-one (K  i approximately 200 nm) and was also sensitive to other previously used inhibitors 5,8,11,14-eicosatetraynoic acid (K  iapproximately 3 μm) and nordihydroguajaretic acid (K  i approximately 15 μm). The influence of these PLA2 inhibitors on elongation in etiolated Arabidopsis seedlings was tested, and tetrahydro-3-(1-naphtalenyl)-2H-pyran-2-one and 5,8,11,14-eicosatetraynoic acid inhibited hypocotyl elongation maximally at concentrations close to theirK  i in vitro.
Movement of Potato Spindle Tuber Viroid Reveals Regulatory Points of Phloem-Mediated RNA Traffic
Authors: Zhu Yali , Qi Yijun , Xun Yan , Owens Robert , Ding Biao
Published: September 2002

Increasing evidence indicates that the phloem mediates traffic of selective RNAs within a plant. How an RNA enters, moves in, and exits the phloem is poorly understood. Potato spindle tuber viroid (PSTVd) is a pathogenic RNA that does not encode proteins and is not encapsidated, and yet it replicates autonomously and traffics systemically within an infected plant. The viroid RNA genome must interact directly with cellular factors to accomplish these functions and is, therefore, an excellent probe to study mechanisms that regulate RNA traffic. Our analyses of PSTVd traffic in Nicotiana benthamianayielded evidence that PSTVd movement within sieve tubes does not simply follow mass flow from source to sink organs. Rather, this RNA is transported into selective sink organs. Furthermore, two PSTVd mutants can enter the phloem to spread systemically but cannot exit the phloem in systemic leaves of tobacco (Nicotiana tabacum). A viroid most likely has evolved structural motifs that mimic endogenous plant RNA motifs so that they are recognized by cellular factors for traffic. Thus, analysis of PSTVd traffic functions may provide insights about endogenous mechanisms that control phloem entry, transport, and exit of RNAs.
F-Actin-Dependent Endocytosis of Cell Wall Pectins in Meristematic Root Cells. Insights from Brefeldin A-Induced Compartments
Authors: Baluška František , Hlavacka Andrej , Šamaj Jozef , Palme Klaus , Robinson David G. , Matoh Toru , McCurdy David W. , Menzel Diedrik , Volkmann Dieter
Published: September 2002

Brefeldin A (BFA) inhibits exocytosis but allows endocytosis, making it a valuable agent to identify molecules that recycle at cell peripheries. In plants, formation of large intracellular compartments in response to BFA treatment is a unique feature of some, but not all, cells. Here, we have analyzed assembly and distribution of BFA compartments in development- and tissue-specific contexts of growing maize (Zea mays) root apices. Surprisingly, these unique compartments formed only in meristematic cells of the root body. On the other hand, BFA compartments were absent from secretory cells of root cap periphery, metaxylem cells, and most elongating cells, all of which are active in exocytosis. We report that cell wall pectin epitopes counting rhamnogalacturonan II dimers cross-linked by borate diol diester, partially esterified (up to 40%) homogalacturonan pectins, and (1→4)-β-d-galactan side chains of rhamnogalacturonan I were internalized into BFA compartments. In contrast, Golgi-derived secretory (esterified up to 80%) homogalacturonan pectins localized to the cytoplasm in control cells and did not accumulate within characteristic BFA compartments. Latrunculin B-mediated depolymerization of F-actin inhibited internalization and accumulation of cell wall pectins within intracellular BFA compartments. Importantly, cold treatment and protoplasting prevented internalization of wall pectins into root cells upon BFA treatment. These observations suggest that cell wall pectins of meristematic maize root cells undergo rapid endocytosis in an F-actin-dependent manner.
Regulated Expression of Arabidopsis Phosphate Transporters
Authors: Karthikeyan Athikkattuvalasu S. , Varadarajan Deepa K. , Mukatira Uthappa T. , D'Urzo Matilde Paino , Damsz Barbara , Raghothama Kashchandra G.
Published: September 2002

Phosphorus deficiency is one of the major abiotic stresses affecting plant growth. Plants respond to the persistent deficiency of phosphate (Pi) by coordinating the expression of genes involved in alleviation of the stress. The high-affinity Pi transporters are among the major molecular determinants that are activated during Pi stress. In this study, using three reporter genes (green fluorescent protein, luciferase, and β-glucuronidase) regulated by two Pi transporter promoters, we have carried out an extensive analysis of transcriptional and spatial regulation of gene expression. Activation of the genes was rapid, repressible, and specific in response to changes in Pi availability. The phytohormones auxin and cytokinin suppressed the expression of the reporter gene driven by the AtPT1promoter, and that of the native gene, suggesting that hormones may be involved in regulation of some component(s) of Pi starvation response pathway. These studies also provide molecular evidence for a potential role of high-affinity Pi transporters in mobilizing Pi into reproductive organs. The results suggest that members of the Pi transporter family may have similar but nonredundant functions in plants.
Blurring services and resources: Boston College’s implementation of MetaLib and SFX
Authors: Gerrity Bob , Lyman Theresa , Tallent Ed
Published: September 2002

Over the past year, the Boston College libraries have been engaged in making available to our users two new resources, MetaLib and SFX. MetaLib is portal/gateway software that allows for simultaneous searching of several databases, as well as some options to customize the interface. SFX technology provides context‐sensitive reference linking from citations to extended services. In this article we review the pedagogical, technical, and service reasons for making these resources available. We discuss how we believe these technologies respond to current student use of the library, and how we approached the installation process. We also review the challenges of the installation (both technical and service) and future possibilities.
Toxic processes challenged at the Strategos “Innovation Academy”
Authors: Merlyn Paul , Välikangas Liisa
Published: September 2002

In a session of the Strategos Innovation Academy, participants considered how a number of core management processes – for example, strategic planning, capital budgeting, performance assessment and product and process development – inhibit innovation. Working in groups, the participants identified problems with existing practices and then suggested a number of ways to make the process less toxic to innovation. Today’s strategic‐planning processes rarely emphasize radical innovation – the new business concepts and operational models that are necessary to keep corporations at the head of the pack – either implicitly or explicitly. Another failure that participants identified is the linkage between strategy planning and the annual budgetary cycle. To improve strategic planning, participants made a number of other suggestions, many of which derive from the toxicities and failures of the existing strategic‐planning process. Companies should first ensure that their business definition and associated mission statement are broad. Narrow definitions are likely to reduce a company’s identity to its current business model, thereby impeding the possibility of renewal. Companies should also explicitly include innovation in the strategic‐planning process. A chief innovation officer – a new senior‐level appointee in the company – can help ensure that innovation remains central to the strategic‐planning process. Greater scrutiny of strategic plans can also help. For example, CEOs can reject strategic plans that do not include a substantial amount of innovation. The introduction of new metrics for innovation would help formalize this commitment to innovation. Participants also recommended that companies find ways to dissociate the strategic‐planning process from an annual schedule. Instead, the process needs to become continuous. To this end, some participants advocated renaming the process strategic evolution instead of strategic planning.
Theoretical and Experimental Dipole Moments of Purines
Authors: Párkányi Cyril , Boniface Christian , Aaron Jean-Jacques , Bulaceanu-MacNair Mihaela , Dakkouri Marwan
Published: September 2002

As a follow-up on our previous study of a series of purines (purine, 6-chloropurine, purine-6-thiol, hypoxanthine, theobromine, theophylline, caffeine, and uric acid), we have investigated six additional biologically important purines (adenine, guanine, isoguanine, thioguanine, xanthine, and kinetin). Their ground-state dipole moments were measured in dioxane at 293 K. The first excited singlet-state dipole moments were obtained using the solvatochromic shift equations (McRae, Suppan, Bakhshiev, and Kawski-Chamma-Viallet). The theoretical dipole moments were calculated as a combination of the π-moment (PPP method) and the σ-moment (vector sum of the σ-bond and σ-group moments). The same approach was used to obtain their first excited singlet-state dipole moments (excited state π-moment; σ-moment assumed to be the same as in the ground state).Ab initioHF 6-31G**calculations were also used to obtain ground-state dipole moments for all the fourteen purines under study. In addition, a DFT/B3PW91/6311++(2df,2p) calculation has been carried out for purine for comparison. The different sets of theoretical dipole moments were compared with the respective experimental values. There is an approximately equally good agreement among the experimental dipole moments and the PPP + σ dipole moments (±6.9%) and theab initiodipole moments (±7.4%). The effect of structure on the dipole moments is discussed.
Determination of Nitroaromatic Photodecomposition Products of Tecnazene Using Liquid Chromatography with Amperometric Detection
Authors: Kubáň Pavel , Flowers Hugh
Published: September 2002

Isocratic liquid chromatographic methods with mobile phases consisting of 20 mmol l-1 sulfuric acid and 60% acetonitrile (for 9 quickly eluted nitroaromatic compounds) or 75% acetonitrile (for 2 strongly retained nitrobenzenes) at a flow rate of 1.2 ml min-1 using a DC amperometric detection with a glassy carbon working electrode at -0.5 V vs Ag/AgCl reference electrode have been developed for the determination of 11 nitroaromatic compounds. The overall repeatability of the retention times was better than 0.5% and the repeatability of the peak areas ranged from 1.2 to 8.3% (n = 7) based on the manual injection of 300 μl of standard solution. Limits of detection (LODs for 3.S/N) of 25-250 nmol l-1 were obtained. The developed methods have been applied to the determination of nitroaromatic decomposition products in the UV-irradiated samples of a tecnazene (1,2,4,5-tetrachloro-3-nitrobenzene) solution. Due to the high selectivity of the DC amperometric detection in reductive mode, no interferences occurred in the analyses of UV-irradiated tecnazene solutions although high concentrations of other compounds (e.g., phenols, chlorophenols, inorganic anions, and carboxylic acids) were present.
Synthesis and Potentiometric Titrations of Some New 4-(Benzylideneamino)-4,5-dihydro-1H-1,2,4-triazol-5-one Derivatives in Non-Aqueous Media
Authors: Bahçeci Şule , Yüksek Haydar , Ocak Zafer , Azakli Ibrahim , Alkan Muzaffer , Özdemir Mustafa
Published: September 2002

The synthesis of 3-alkyl(aryl)-4-[4-(dimethylamino)benzylideneamino]-4,5-dihydro-1H-1,2,4-triazol-5-ones (3) from the reactions of 3-alkyl(aryl)-4-amino-4,5-dihydro-1H-1,2,4-triazol-5-ones (2) with 4-(dimethylamino)benzaldehyde is described. The newly synthesized 3 type compounds were titrated potentiometrically with tetrabutylammonium hydroxide in three non-aqueous solvents, including acetonitrile, isopropyl alcohol and N,N-dimethylformamide. The half-neutralization potential values and the corresponding pKa values were determined for all cases. Thus, the effects of solvents and molecular structure upon acidity were investigated. In addition, N-acetyl derivatives of 3 type compounds were also obtained. The new synthesized compounds in the study were fully characterized.
What distinguishes unintentional injuries from injuries due to intimate partner violence: a study in Greek ambulatory care settings
Authors: Petridou E , Browne A , Lichter E , Dedoukou X , Alexe D , Dessypris N
Published: September 2002

Objectives:Intimate partner violence (IPV) is an important sociocultural and public health problem. This study aims to assess sociodemographic and injury characteristics of IPV victims among adults in a traditional southern European population.

Setting:Accident and emergency departments of three sentinel hospitals in Greece participating in the Emergency Department Injury Surveillance System (EDISS).

Methods:Data on sociodemographic variables, as well as event and injury characteristics were retrieved from the EDISS database during the three year period 1996–98. Out of a total of 27 319 injured women aged 19 years or more, 312 (1.1%) were reported as IPV related and were compared with 26 466 women with unintentional injuries. Among the 35 174 men with injuries 39 (0.1%) were reported as IPV related and were compared with 34 049 men with unintentional injuries. The data were analyzed through simple cross tabulations and multiple logistic regression. Positive predicted values for selected injury characteristics were also calculated.

Results:IPV is more common in rural than in urban areas of Greece. Women are 10 times more frequently IPV victims but men are also IPV victims; younger women and older men are disproportionately affected by IPV. The relative frequency of the phenomenon increases during the late evening and night hours. Certain types of injuries, notably multiple facial injuries, and presentation of the injured person on his/her own at the emergency department or combinations of predictive characteristics are strongly indicative of IPV.

Conclusions:Injuries due to IPV are not uncommon in Greece, not withstanding the traditional structure of the society and the tendency of under-reporting. Certain injury characteristics have high positive predictive values and could be used in screening protocols aiming at the correct identification of the underlying external cause in injuries that may be caused by IPV.