bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tissue‐specific oxylipin signature of tomato flowers: allene oxide cyclase is highly expressed in distinct flower organs and vascular bundles
Authors: Hause Bettina , Stenzel Irene , Miersch Otto , Maucher Helmut , Kramell Robert , Ziegler Jöörg , Wasternack Claus 
Published: March 2003

SummaryA crucial step in the biosynthesis of jasmonic acid (JA) is the formation of its correct stereoisomeric precursor,cis(+)12‐oxophytodienoic acid (OPDA). This step is catalysed by allene oxide cyclase (AOC), which has been recently cloned from tomato . In stems, young leaves and young flowers, AOC mRNA accumulates to a low level , contrasting with a high accumulation in flower buds, flower stalks and roots. The high levels of AOC mRNA and AOC protein in distinct flower organs correlate with high AOC activity, and with elevated levels of JA, OPDA and JA isoleucine conjugate. These compounds accumulate in flowers to levels of about 20 nmol g−1fresh weight, which is two orders of magnitude higher than in leaves. In pistils, the level of OPDA is much higher than that of JA, whereas in flower stalks, the level of JA exceeds that of OPDA. In other flower tissues, the ratios among JA, OPDA and JA isoleucine conjugate differ remarkably, suggesting a tissue‐specific oxylipin signature. Immunocytochemical analysis revealed the specific occurrence of the AOC protein in ovules, the transmission tissue of the style and in vascular bundles of receptacles, flower stalks, stems, petioles and roots. Based on the tissue‐specific AOC expression and formation of JA, OPDA and JA amino acid conjugates, a possible role for these compounds in flower development is discussed in terms of their effect on sink–source relationships and plant defence reactions. Furthermore, the AOC expression in vascular bundles might play a role in the systemin‐mediated wound response of tomato.
Alternative splicing of a novel diacylglycerol kinase in tomato leads to a calmodulin‐binding isoform
Authors: Snedden Wayne A. , Blumwald Eduardo 
Published: March 2003

SummaryCalmodulin is a regulatory protein activated during Ca2+ signalling. We have isolated a cDNA, designated LeCBDGK (Lycopersicon esculentum calmodulin‐binding diacylglycerol kinase) encoding a novel calmodulin‐binding protein with sequence similarity to diacylglycerol kinases from animals. Diacylglycerol kinases convert diacylglycerol to phosphatidic acid. We delineated the calmodulin‐binding domain to approximately 25 residues near the C‐terminus of LeCBDGK. We have also isolated a second diacylglycerol kinase cDNA, designated LeDGK1, identical to LeCBDGK, except that it lacks the calmodulin‐binding domain. Both recombinant LeCBDGK and LeDGK1 were catalytically active in vitro. Anti‐DGK antiserum detected two immunoreactive proteins associated with microsomal and plasma membrane fractions from cell suspensions. The higher molecular weight immunoreactive protein was also present in soluble extracts and bound to calmodulin–agarose in the presence of calcium, demonstrating that native LeCBDGK is a calmodulin‐binding protein. In the presence of calcium, LeCBDGK associated with membrane cell fractions in vitro, but calmodulin antagonists disrupted this association, suggesting a possible role of calcium in the recruitment of LeCBDGK from soluble to membrane cell fractions. Native LeCBDGK and calmodulin co‐immunoprecipitated from tomato soluble cell extracts, suggesting their interaction in vivo. The same gene encodes both LeCBDGK and LeDGK1 and the calmodulin‐binding domain of LeCBDGK is encoded by a separate exon. Thus, alternative transcript splicing leads to calmodulin‐binding and non‐binding forms of diacylglycerol kinases in tomato. Possible roles of LeCBDGK and LeDGK1 in calcium and lipid signalling are discussed.
AtFim1 is an actin filament crosslinking protein from Arabidopsis thaliana
Authors: Kovar David R. , Staiger Christopher J. , Weaver Eric A. , McCurdy David W. 
Published: March 2003

Summary ATFIM1 is a widely expressed gene in Arabidopsis thaliana that encodes a putative actin filament‐crosslinking protein, AtFim1, belonging to the fimbrin/plastin class of actin‐binding proteins. In this report we have used bacterially expressed AtFim1 and actin isolated from Zea mays pollen to demonstrate that AtFim1 functions as an actin filament‐crosslinking protein. AtFim1 binds pollen actin filaments (F‐actin) in a calcium‐independent manner, with an average dissociation constant (Kd) of 0.55 ± 0.21 µM and with a stoichiometry at saturation of 1 : 4 (mol AtFim1 : mol actin monomer). AtFim1 also crosslinks pollen F‐actin by a calcium‐independent mechanism, in contrast to crosslinking of plant actin by human T‐plastin, a known calcium‐sensitive actin‐crosslinking protein. When micro‐injected at high concentration into living Tradescantia virginiana stamen hair cells, AtFim1 caused cessation of both cytoplasmic streaming and transvacuolar strand dynamics within 2–4 min. Using the ‘nuclear displacement assay’ as a measure of the integrity of the actin cytoskeleton in living stamen hair cells, we demonstrated that AtFim1 protects actin filaments in these cells from Z. mays profilin (ZmPRO5)‐induced depolymerization, in a dose‐dependent manner. The apparent ability of AtFim1 to protect actin filaments in vivo from profilin‐mediated depolymerization was confirmed by in vitro sedimentation assays. Our results indicate that AtFim1 is a calcium‐independent, actin filament‐crosslinking protein that interacts with the actin cytoskeleton in living plant cells.
Plastid‐expressed 5‐enolpyruvylshikimate‐3‐phosphate synthase genes provide high level glyphosate tolerance in tobacco
Authors: Ye Guang‐Ning , Hajdukiewicz Peter T.J. , Broyles Debra , Rodriguez Damian , Xu Charles W. , Nehra Narender , Staub Jeffrey M. 
Published: March 2003

SummaryPlastid transformation (transplastomic) technology has several potential advantages for biotechnological applications including the use of unmodified prokaryotic genes for engineering, potential high‐level gene expression and gene containment due to maternal inheritance in most crop plants. However, the efficacy of a plastid‐encoded trait may change depending on plastid number and tissue type. We report a feasibility study in tobacco plastids to achieve high‐level herbicide resistance in both vegetative tissues and reproductive organs. We chose to test glyphosate resistance via over‐expression in plastids of tolerant forms of 5‐enolpyruvylshikimate‐3‐phosphate synthase (EPSPS). Immunological, enzymatic and whole‐plant assays were used to prove the efficacy of three different prokaryotic (Achromobacter, Agrobacterium and Bacillus) EPSPS genes. Using the Agrobacterium strain CP4 EPSPS as a model we identified translational control sequences that direct a 10 000‐fold range of protein accumulation (to >10% total soluble protein in leaves). Plastid‐expressed EPSPS could provide very high levels of glyphosate resistance, although levels of resistance in vegetative and reproductive tissues differed depending on EPSPS accumulation levels, and correlated to the plastid abundance in these tissues. Paradoxically, higher levels of plastid‐expressed EPSPS protein accumulation were apparently required for efficacy than from a similar nuclear‐encoded gene. Nevertheless, the demonstration of high‐level glyphosate tolerance in vegetative and reproductive organs using transplastomic technology provides a necessary step for transfer of this technology to other crop species.
Negative autoregulation of the Arabidopsis homeobox gene ATHB‐2
Authors: Ohgishi Maki , Oka Atsuhiro , Morelli Giorgio , Ruberti Ida , Aoyama Takashi 
Published: March 2003

SummaryThe Arabidopsis homeobox gene ATHB‐2 is tightly regulated by light signals, and is thought to direct morphological changes during shade avoidance responses. To understand how ATHB‐2 mediates light signals in plant morphogenesis, we investigated its transcriptional network. We constructed a gene encoding a chimeric transcription factor (HD‐Zip‐2‐V‐G) that is expected to activate target genes of ATHB‐2 in a glucocorticoid‐dependent manner. In transgenic Arabidopsis plants expressing HD‐Zip‐2‐V‐G, glucocorticoid treatment activates the ATHB‐2 gene itself, independent of de novo protein synthesis. An in vitro DNase I‐footprinting experiment showed that recombinant ATHB‐2 protein specifically bound to an ATHB‐2 promoter region. These complementary results indicate that ATHB‐2 recognizes its own promoter. Consistent with the fact that ATHB‐2 itself has been shown to act as a repressor, expression of the endogenous ATHB‐2 gene was repressed in transgenic plants overexpressing an ATHB‐2 transgene. Moreover, target‐gene analysis using the HD‐Zip‐2‐V‐G suggested that ATHB‐2 recognizes other HD‐Zip II subfamily genes. We conclude that ATHB‐2 has a negative autoregulatory loop and may be involved in a complicated transcriptional network involving paralogous genes, as is the case with animal homeobox genes.
Analysis of the function of two circadian‐regulated CONSTANS‐LIKE genes
Authors: Ledger Susan , Strayer Carl , Ashton Fern , Kay Steve A. , Putterill Jo 
Published: March 2003

SummaryThe Arabidopsis genes CONSTANS‐LIKE 1 (COL1) and CONSTANS‐LIKE 2 (COL2) are predicted to encode zinc finger proteins with ∼67% amino acid identity to the protein encoded by the flowering‐time gene CONSTANS (CO). We show that the circadian clock regulates expression of COL1 and COL2 with a peak in transcript levels around dawn. We analyzed transgenic plants misexpressing COL1, COL2 and CO. Unlike CO, altered expression of COL1 and COL2 in transgenic plants had little effect on flowering time. However, analysis of circadian phenotypes in the transgenic plants showed that over‐expression of COL1 can shorten the period of two distinct circadian rhythms. Experiments with the highest COL1 over‐expressing line indicate that its circadian defects are fluence rate‐dependent, suggesting an effect on a light input pathway(s).
The male sterile G cytoplasm of wild beet displays modified mitochondrial respiratory complexes
Authors: Ducos Eric , Touzet Pascal , Boutry Marc 
Published: March 2003

SummaryCytoplasmic male sterility (CMS) in higher plants has been mainly studied in cultivated species. In most cases, pollen abortion is linked to the presence of an additional mitochondrial polypeptide leading to organelle dysfunction in reproductive tissues. In wild beet, both CMS and hermaphrodite plants coexist in natural populations. The G cytoplasm is widely distributed along the Western European coast, and previous genetic studies have demonstrated that this cytoplasm confers male sterility in beet. In the present study, we have identified two mutations of G mitochondrial genes, each of which results in the production of a respiratory chain complex subunit with an altered molecular weight; the NAD9 subunit has a C‐terminal extension while the COX2 subunit has a truncated C‐terminus. NADH dehydrogenase activity was unchanged in leaves, but cytochrome c oxidase activity was reduced by 50%. Moreover, Western blot analyses revealed that alternative oxidase was more abundant in male sterile G plants than in a fertile control (Nv), suggesting that this alternative pathway might compensate for the cytochrome c oxidase deficiency. Implications of respiratory chain changes and a putative link with CMS are discussed.
The 5′ untranslated region of the At‐P5R gene is involved in both transcriptional and post‐transcriptional regulation
Authors: Hua Xue Jun , Van De Cotte Brigitte , Van Montagu Marc , Verbruggen Nathalie 
Published: March 2003

SummaryThe steady‐state level of transcripts coding for the pyrroline‐5‐carboxylate reductase of Arabidopsis (At‐P5R) increased under salt and heat stress, mainly because of an enhanced mRNA stability. However, the At‐P5R protein level was not induced, and its translation was inhibited at initiation stage and probably also at later stages. Replacement of the 5′ untranslated region (5′UTR) and β‐glucuronidase (gus) fusion analysis revealed that the first 92 bp region of the At‐P5R 5′UTR was sufficient to mediate transcript stabilization and translation inhibition during salt and heat stresses. Furthermore, the first 92 bp region of the At‐P5R 5′UTR was also involved in transcription efficiency in a promoter‐dependent manner. The results demonstrated that the stress regulation of At‐P5R is complex and involves the 5′UTR which acts at three levels, partly in opposing directions.
The role of calcium in ABA‐induced gene expression and stomatal movements
Authors: Webb Alex A. R. , Larman Mark G. , Montgomery Lucy T. , Taylor Jane E. , Hetherington Alistair M. 
Published: March 2003

SummaryThere is much interest in the transduction pathways by which abscisic acid (ABA) regulates stomatal movements (ABA‐turgor signalling) and by which this phytohormone regulates the pattern of gene expression in plant cells (ABA‐nuclear signalling). A number of second messengers have been identified in both the ABA‐turgor and ABA‐nuclear signalling pathways. A major challenge is to understand the architecture of ABA‐signalling pathways and to determine how the ABA signal is coupled to the appropriate response. We have investigated whether separate Ca2+‐dependent and ‐independent ABA‐signalling pathways are present in guard cells. Our data suggest that increases in [Ca2+]i are a common component of the guard cell ABA‐turgor and ABA‐nuclear signalling pathways. The effects of Ca2+ antagonists on ABA‐induced stomatal closure and the ABA‐responsive CDeT6‐19 gene promoter suggest that Ca2+ is involved in both ABA‐turgor signalling and ABA‐nuclear signalling in guard cells. However, the sensitivity of these pathways to alterations in the external calcium concentration differ, suggesting that the ABA‐nuclear and ABA‐turgor signalling pathways are not completely convergent. Our data suggest that whilst Ca2+‐independent signalling elements are present in the guard cell, they do not form a completely separate Ca2+‐independent ABA‐signalling pathway.
Identification of a cis‐regulatory element for L1 layer‐specific gene expression, which is targeted by an L1‐specific homeodomain protein
Authors: Abe Mitsutomo , Takahashi Taku , Komeda Yoshibumi 
Published: March 2003

SummaryThe Arabidopsis thaliana PROTODERMAL FACTOR1 (PDF1) gene encoding a putative extracellular proline‐rich protein is exclusively expressed in the L1 layer of shoot apices and the protoderm of organ primordia. In order to identify essential cis‐regulatory sequences required for the L1 layer‐specific expression, a series of 5′ deletions of the PDF1 promoter were fused to the β‐glucronidase (GUS) gene and introduced into Arabidopsis plants. Our analysis revealed that the minimum region necessary to confer L1‐specific expression of PDF1 is confined within a 260‐bp fragment upstream of the transcription start site. We identified an 8‐bp motif in this region that is conserved between promoter regions of all the L1‐specific genes so far cloned, and we designated it the L1 box. Electrophoretic mobility shift assays demonstrated that the L1‐specific homeodomain protein ATML1 can bind to the L1 box sequence in vitro. The GUS expression in transgenic plants disappeared when a mutation that abolishes binding of ATML1 was introduced into the PDF1 l1 box sequence of the construct. These results suggest that the L1 box plays a crucial role in the regulation of PDF1 expression in L1 cells and that ATML1 could cooperate to drive L1‐specific expression.
Identification of a novel pathogen‐responsive element in the promoter of the tobacco gene HSR203J, a molecular marker of the hypersensitive response
Authors: Pontier Dominique , Balagué Claudine , Bezombes‐Marion Isabelle , Tronchet Maurice , Deslandes Laurent , Roby Dominique 
Published: March 2003

SummaryThe tobacco gene, HSR203J, which is specifically activated during the early steps of incompatible plant/pathogen interactions has been shown to be a molecular marker of the hypersensitive response (HR). It constitutes an ideal model for the identification of HR‐responsive cis‐regulatory elements. As a first step in the promoter dissection, deletion mutants of the 5′ flanking sequence of HSR203J fused to the GUS reporter gene were analyzed. Then, the construction and study of chimeric constructs containing HSR203J promoter fragments fused to a minimal promoter enabled us to identify a 28‐bp regulatory element located between −106 and −79 upstream of the transcription initiation site. This element has been shown to be necessary and sufficient for transcriptional activation in response to pathogen. It contains a 10‐bp palindrome followed by its unperfect repeat. The mutagenesis of these two sequence elements led to the identification of a 12‐bp motif termed HSRE (HSR203 responsive element) responsible for the marked induction of the HSR203J gene during the HR. Since this DNA region did not show any homology with known regulatory sequences, this 12 bp motif corresponds to a novel cis‐regulatory element.
An Arabidopsis inositol phospholipid kinase strongly expressed in procambial cells: Synthesis of PtdIns(4,5)P2 and PtdIns(3,4,5)P3 in insect cells by 5‐phosphorylation of precursors
Authors: Elge Stephan , Brearley Charles , Xia Hui‐Jun , Kehr Julia , Xue Hong‐Wei , Mueller‐Roeber Bernd 
Published: March 2003

SummaryWe have cloned a phosphatidylinositol‐4‐phosphate 5‐kinase (PIP5K) cDNA (AtP5K1) from Arabidopsis thaliana. By the application of cell permeabilization and short‐term nonequilibrium labelling we show that expression of AtP5K1 in Baculovirus‐infected insect (Spodoptera frugiperda) cells directs synthesis of PtdIns(4,5)P2 and PtdIns(3,4,5)P3. The same phosphoinositides were produced by isolated whole‐cell membrane fractions of AtP5K1‐expressing insect cells. Their synthesis was not affected by adding defined precursor lipids, that is PtdIns(3)P, PtdIns(4)P, PtdIns(3,4)P2, or PtdIns(4,5)P2, in excess, indicating that substrates for the plant enzyme were not limiting in vivo. Enzymatic dissection of lipid headgroups revealed that AtP5K1‐directed synthesis of PtdIns(4,5)P2 and PtdIns(3,4,5)P3 proceeds via 5‐phosphory lation of precursors. Analysis of promoter‐reporter gene (β‐glucuronidase) fusions in transgenic plants revealed that expression of the AtP5K1 gene is strongest in vascular tissues of leaves, flowers, and roots, namely in cells of the lateral meristem, that is the procambium. Single‐cell sampling of sap from flower stem meristem tissue and neighbouring phloem cells, when coupled to reverse transcriptase – polymerase chain reaction, confirmed preferential expression of AtP5K1 in procambial tissue. We hypothesize that AtP5K1, like animal and yeast PIP5K, may be involved in the control of cell proliferation.
Knock‐out of the plastid ribosomal protein L11 in Arabidopsis: effects on mRNA translation and photosynthesis
Authors: Pesaresi Paolo , Varotto Claudio , Meurer Jörg , Jahns Peter , Salamini Francesco , Leister Dario 
Published: March 2003

SummaryThe prpl11–1 mutant of Arabidopsis thaliana was identified among a collection of T‐DNA tagged lines on the basis of a decrease in the effective quantum yield of photosystem II. The mutation responsible was localized to Prpl11, a single‐copy nuclear gene that encodes PRPL11, a component of the large subunit of the plastid ribosome. The amino acid sequence of Arabidopsis PRPL11 is very similar to those of L11 proteins from spinach and prokaryotes. In the prpl11–1 mutant, photosensitivity and chlorophyll fluorescence parameters are significantly altered owing to changes in the levels of thylakoid protein complexes and stromal proteins. The abundance of most plastome transcripts examined, such as those of genes coding for the photosystem II core complex and RbcL, is not decreased. Plastid ribosomal RNA accumulates in wild‐type amounts, and the assembly of plastid polysomes on the transcripts of the rbcL, psbA and psbE genes remains mainly unchanged in mutant plants, indicating that lack of PRPL11 affects neither the abundance of plastid ribosomes nor their assembly into polysomes. However, in vivo translation assays demonstrate that the rate of translation of the large subunit of Rubisco (RbcL) is significantly reduced in prpl11–1 plastids. Our data suggest a major role for PRPL11 in plastid ribosome activity per se, consistent with its location near the GTPase‐binding centre of the chloroplast 50S ribosomal subunit. Additional effects of the mutation, including the pale green colour of the leaves and a drastic reduction in growth rate under greenhouse conditions, are compatible with reduced levels of protein synthesis in plastids.
Expression of Clarkia S‐linalool synthase in transgenic petunia plants results in the accumulation of S‐linalyl‐β‐d‐glucopyranoside
Authors: Lücker Joost , Bouwmeester Harro J. , Schwab Wilfried , Blaas Jan , Van Der Plas Linus H. W. , Verhoeven Harrie A. 
Published: March 2003

Summary Petunia hybrida W115 was transformed with a Clarkia breweri S‐linalool synthase cDNA (lis). Lis was expressed in all tissues analysed, and linalool was detected in leaves, sepals, corolla, stem and ovary, but not in nectaries, roots, pollen and style. However, the S‐linalool produced by the plant in the various tissues is not present as free linalool, but was efficiently converted to non‐volatile S‐linalyl‐β‐d‐glucopyranoside by the action of endogenous glucosyltransferase. The results presented demonstrate that monoterpene production can be altered by genetic modification, and that the compounds produced can be converted by endogenous enzymatic activity.
A nuclear gene encoding mitochondrial Δ1‐pyrroline‐5‐carboxylate dehydrogenase and its potential role in protection from proline toxicity
Authors: Deuschle Karen , Funck Dietmar , Hellmann Hanjo , Däschner Klaus , Binder Stefan , Frommer Wolf B. 
Published: March 2003

SummaryΔ1‐pyrroline‐5‐carboxylate (P5C), an intermediate in biosynthesis and degradation of proline (Pro), is assumed to play a role in cell death in plants and animals. Toxicity of external Pro and P5C supply to Arabidopsis suggested that P5C dehydrogenase (P5CDH; EC 1.2.1.12) plays a crucial role in this process by degrading the toxic Pro catabolism intermediate P5C. Also in a Δput2 yeast mutant, lacking P5CDH, Pro led to growth inhibition and formation of reactive oxygen species (ROS). Complementation of the Δput2 mutant allowed identification of the Arabidopsis P5CDH gene. AtP5CDH is a single‐copy gene and the encoded protein was localized to the mitochondria. High homology of AtP5CDH to LuFIS1, an mRNA up‐regulated during susceptible pathogen attack in flax, suggested a role for P5CDH in inhibition of hypersensitive reactions. An Arabidopsis mutant (cpr5) displaying a constitutive pathogen response was found to be hypersensitive to external Pro. In agreement with a role in prevention of cell death, AtP5CDH was expressed at a basal level in all tissues analysed. The highest expression was found in flowers that are known to contain the highest Pro levels under normal conditions. External supply of Pro induced AtP5CDH expression, but much more slowly than Pro dehydrogenase (AtProDH) expression. Uncoupled induction of the AtProDH and AtP5CDH genes further supports the hypothesis that P5C levels have to be tightly controlled. These results indicate that, in addition to the well‐studied functions of Pro, for example in osmoregulation, the Pro metabolism intermediate P5C also serves as a regulator of cellular stress responses.