bluuebunny/crossref_metadata_2025_split huggingface dataset:
Isolation of a gene encoding Arabidopsis membrane‐associated acyl‐CoA binding protein and immunolocalization of its gene product
Authors: Chye Mee‐Len , Huang Bing‐Quan , Zee Sze Yong 
Published: March 2003

SummaryUntil recently, only cytosolic acyl‐CoA binding proteins (ACBPs) have been characterized. The isolation of an Arabidopsis thaliana cDNA encoding a novel membrane‐associated ACBP that accumulates in developing seeds, designated ACBP1, has provided evidence for the existence of membrane‐associated forms of ACBPs (
Chye 1998, Plant Mol. Biol. 38, 827–838). We now report on the isolation of its corresponding gene from an A. thaliana Columbia genomic library using the ACBP1 cDNA as a hybridization probe. Nucleotide sequence analysis of Arabidopsis ACBP1 showed that its promoter lacks a TATA box, resembling the promoters of rat, Drosophila and human genes encoding cytosolic ACBP and suggesting that it is a housekeeping gene. We show by Western blot analysis that ACBP1 expression in developing seeds coincides with lipid deposition and that homologues of membrane‐associated ACBP1 exist in other plants. Using light microscopy, we show that ACBP1 is strongly expressed in the embryo at the cotyledons, hypocotyl, procambium of the axis and in most peripheral cells of the cotyledons and hypocotyl. Immunogold labelling localized ACBP1 to vesicles, to the plasma membrane especially at epidermal cells of heart, torpedo and cotyledonary stage embryos, and to the cell wall of the outer integument cells at the seed coat. Our results suggest that ACBP1 is involved in intermembrane lipid transport from the ER via vesicles to the plasma membrane where it could maintain a membrane‐associated acyl pool; its immunolocalization to the cell wall of outer integument cells at the seed coat suggests a role in cuticle and cutin formation.
A G‐box motif (GCCACGTGCC) tetramer confers high‐level constitutive expression in dicot and monocot plants
Authors: Ishige Fumiharu , Takaichi Miyuki , Foster Randy , Chua Nam‐Hai , Oeda Kenji 
Published: March 2003

SummaryGUS reporter expression from 11 basal promoters (CaMV –90) with G‐box cores (CACGTG) was analysed to evaluate the regulatory roles of G‐box flanking sequences. While most G‐box motifs exhibited some tissue preference of gene expression, a distinct tissue‐specific expression was not apparent. However, one of 11 G‐box sequences, the G‐box 10 (GCCACGTGCC) tetramer, conferred a high‐level constitutive expression in seed, root, leaf, axillary bud, almost all parts of flower buds and pollen of transgenic tobacco plants. Furthermore, the G‐box 10 tetramer promoter exhibited high‐level expression in transgenic dicot carrot and monocot rice. This is apparently the first report of a G‐box motif conferring a high‐level constitutive expression in a non‐tissue‐specific manner.
The Nicotiana tabacum plasma membrane aquaporin NtAQP1 is mercury‐insensitive and permeable for glycerol
Authors: Biela Alexander , Grote Karsten , Otto Beate , Hoth Stefan , Hedrich Rainer , Kaldenhoff Ralf 
Published: March 2003

Summary A new aquaporin from Nicotiana tabacum (cv. Samsun) was characterized. It shares sequence homology to the Arabidopsis thaliana PIP1 protein family. By two‐phase partitioning and immunoblot analysis, plasma membrane localization could be demonstrated. The corresponding mRNA is highly abundant in roots and flowers, while it is rarely expressed in leaves and stems. Functional expression in Xenopus oocytes revealed that NtAQP1 can mediate glycerol transport in addition to water flow. However, NtAQP1 is impermeable for Na

+

, K

+

and Cl

–

ions. The water permeability and selectivity could not be modulated by addition of mercurials or the activity of cAMP‐dependent protein kinases.
Regulation of phytochrome B signaling by phytochrome A and FHY1 in Arabidopsis thaliana
Authors: Cerdán Pablo D. , Yanovsky Marcelo J. , Reymundo F. Carolina , Nagatani Akira , Staneloni Roberto J. , Whitelam Garry C. , Casal Jorge J. 
Published: March 2003

Summary Phytochrome A (phyA) and phytochrome B (phyB) share the control of many processes but little is known about mutual signaling regulation. Here, we report on the interactions between phyA and phyB in the control of the activity of an Lhcb1*2 gene fused to a reporter, hypocotyl growth and cotyledon unfolding in etiolated Arabidopsis thaliana. The very‐low fluence responses (VLFR) induced by pulsed far‐red light and the high‐irradiance responses (HIR) observed under continuous far‐red light were absent in the phyA and phyA phyB mutants, normal in the phyB mutant, and reduced in the fhy1 mutant that is defective in phyA signaling. VLFR were also impaired in Columbia compared to Landsberg erecta. The low‐fluence responses (LFR) induced by red‐light pulses and reversed by subsequent far‐red light pulses were small in the wild type, absent in phyB and phyA phyB mutants but strong in the phyA and fhy1 mutants. This indicates a negative effect of phyA and FHY1 on phyB‐mediated responses. However, a pre‐treatment with continuous far‐red light enhanced the LFR induced by a subsequent red‐light pulse. This enhancement was absent in phyA
, phyB
, or phyA phyB and partial in fhy1
. The levels of phyB were not affected by the phyA or fhy1 mutations or by far‐red light pre‐treatments. We conclude that phyA acting in the VLFR mode (i.e. under light pulses) is antagonistic to phyB signaling whereas phyA acting in the HIR mode (i.e. under continuous far‐red light) operates synergistically with phyB signaling, and that both types of interaction require FHY1.
Stable chloroplast transformation in potato: use of green fluorescent protein as a plastid marker
Authors: Sidorov Vladimir A. , Kasten Daniel , Pang Sheng‐Zhi , Hajdukiewicz Peter T. J. , Staub Jeffrey M. , Nehra Narender S. 
Published: March 2003

SummaryWe describe here the development of a reproducible plastid transformation system for potato and regeneration of plants with uniformly transformed plastids. Two distinct tobacco‐specific plastid vectors, pZS197 (Prrn/aadA/TpsbA) and pMON30125 (Prrn/GFP/Trps16::PpsbA/aadA/TpsbA), designed for integration into the large single copy and inverted repeat regions of the plastid genome, respectively, were bombarded into leaf explants of potato line FL1607. A total of three transgenic lines were selected out of 46 plates bombarded with pZS197 and three transgenic lines out of 104 plates were obtained with pMON30125. Development of a high frequency leaf‐based regenera‐ tion system, a stringent selection scheme and optimization of biolistic transformation protocol were critical for recovery of plastid transformants. Plastid‐expressed green fluorescent protein was used as a visual marker for identification of plastid transformants at the early stage of selection and shoot regeneration. The establishment of a plastid transformation system in potato, which has several advantages over routinely used nuclear transformation, offers new possibilities for genetic improvement of this crop.
PR‐1 protein inhibits the differentiation of rust infection hyphae in leaves of acquired resistant broad bean
Authors: Rauscher Martina , Ádám Attila L. , Wirtz Sabine , Guggenheim Richard , Mendgen Kurt , Deising Holger B. 
Published: March 2003

SummaryTreatment of broad bean leaves with salicylic acid (SA) or 2,6‐dichloro‐isonicotinic acid (DCINA) induces resistance against the rust fungus Uromyces fabae resulting in reduced rust pustule density. Light‐microscopy studies showed that in induced resistant plants the rust fungus is inhibited immediately after penetration through the stomatal pore. The differentia‐ tion of infection structures growing within the intercellular space of the leaf, i.e. infection hyphae and haustorial mother cells, is inhibited. Furthermore, low‐temperature scanning electron microscopy studies of freeze fractures revealed protrusions at the tips of infection hyphae growing in induced resistant broad bean leaves. Treatment of in vitro‐differentiating rust infection structures with intercellular fluids (IFs) from induced resistant plants confirmed that the fungus is sensitive towards an apoplastic anti‐fungal activity only after having formed appressoria. Other legume rusts such as U. vignae and U. appendiculatus were likewise inhibited in the presence of IF from SA‐treated broad bean leaves. Heterologous antibodies were used to study changes in the extracellular pathogenesis‐related (PR) protein pattern after resistance induction. Western blots indicated that chitinases and β‐1,3‐glucanases were present in both induced and control plants. In contrast, PR‐1 proteins were newly synthesized in response to SA or DCINA application. The major induced PR‐1 protein was purified and exhibited strong differentiation‐inhibiting activity towards U. fabae infection structures. We conclude that the inhibition of rust infection hyphae in acquired resistant broad bean plants is mainly due to the anti‐fungal activity of this induced basic PR‐1 protein.
Long‐chain acyl‐CoA oxidases of Arabidopsis
Authors: Hooks Mark A. , Kellas Fiona , Graham Ian A. 
Published: March 2003

SummaryFull‐length cDNAs coding for two distinct acyl‐CoA oxidases were isolated by screening an Arabidopsis cDNA library. The genes for the two acyl‐CoA oxidases have been termed AtACX1 and AtACX2. AtACX1 encodes a peptide of 664 amino acids possessing a molecular mass of 74.3 kDa. AtACX2 encodes a peptide of 691 amino acids in length with a molecular mass of 77.5 kDa. Peroxisomal targeting signals were identified in the primary sequences. AtACX1 has a putative PTS1, whereas AtACX2 has a characteristic PTS2. Expression of AtACX1 and AtACX2 in Escherichia coli gave active enzymes for enzymatic and biochemical analysis. AtACX1 was active with both medium‐and long‐chain saturated fatty acyl‐CoAs and showed maximal activity with C14‐CoA. Activity with mono‐unsaturated acyl‐CoAs was slightly higher than with the corresponding saturated acyl‐CoA. AtACX2 was active with long‐chain acyl‐CoAs and showed maximal activity with C18‐CoA. AtACX2 activity with mono‐unsaturated acyl‐CoAs was approximately twice as high as with the corresponding saturated acyl‐CoA. Both enzymes have an apparent Km of approximately 5 μm with the preferred substrate. Northern analysis was conducted to determine the expression patterns of AtACX1 and AtACX2 during germination and in various tissues of a mature plant. The two genes showed generally similar expression profiles and steady‐state mRNA levels in seedlings and mature tissues, but subtle differences were observed. Enzymatic analyses of plant extracts revealed that AtACX1 and AtACX2 are members of a family that includes acyl‐CoA oxidases specific for shorter‐chain acyl‐CoAs. Through expression of antisense constructs of the individual genes, we were able to decrease long‐chain oxidase activity only in antisense AtACX1 plants. Seedlings with long‐chain oxidase activity reduced down to 30% of wild‐type levels germinated and established normally; however, reduced root growth appeared to be a general feature of antisense AtACX1 plants.
Seed‐specific overexpression of phytoene synthase: increase in carotenoids and other metabolic effects
Authors: Shewmaker Christine K. , Sheehy Julie A. , Daley Maureen , Colburn Susan , Ke Dang Yang 
Published: March 2003

SummaryA bacterial phytoene synthase (crtB) gene was overexpressed in a seed‐specific manner and the protein product targeted to the plastid in Brassica napus (canola). The resultant embryos from these transgenic plants were visibly orange and the mature seed contained up to a 50‐fold increase in carotenoids. The predominant carotenoids accumulating in the seeds of the transgenic plants were alpha and beta‐carotene. Other precursors such as phytoene were also detected. Lutein, the predominant carotenoid in control seeds, was not substantially increased in the transgenics. The total amount of carotenoids in these seeds is now equivalent to or greater than those seen in the mesocarp of oil palm. Other metabolites in the isoprenoid pathway were examined in these seeds. Sterol levels remained essentially the same, while tocopherol levels decreased significantly as compared to non‐transgenic controls. Chlorophyll levels were also reduced in developing transgenic seed. Additionally, the fatty acyl composition was altered with the transgenic seeds having a relatively higher percentage of the 18 : 1 (oleic acid) component and a decreased percentage of the 18 : 2 (linoleic acid) and 18 : 3 (linolenic acid) components. This dramatic increase in flux through the carotenoid pathway and the other metabolic effects are discussed.
Transcript profiling in rice (Oryza sativa L.) seedlings using serial analysis of gene expression (SAGE)
Authors: Matsumura Hideo , Nirasawa Shizuko , Terauchi Ryohei 
Published: March 2003

SummarySerial analysis of gene expression (SAGE) was applied for profiling expressed genes in rice seedlings. In the SAGE method, a 9–11 bp fragment (tag) represents each transcript, and frequency of a tag in the sample directly reflects the abundance of the respective mRNA. We studied 10 122 tags derived from 5921 expressed genes in rice (Oryza sativa L.) seedlings, among which only 1367 genes (23.1%) matched the rice cDNA or EST sequences in the DNA database. SAGE showed that most of the highly expressed genes in rice seedlings belong to the category of housekeeping genes (genes encoding ribosomal proteins or proteins responsible for metabolism and cell structure). Unexpectedly, the most highly expressed gene in rice seedlings was a metallothionein (MT) gene, and together with three other messages for MT, it accounts for 2.7% of total gene expression. To our knowledge, this is the first quantitative study of global gene expression in a higher plant. We further applied the SAGE technique to identify differentially expressed genes between anaerobically treated and untreated rice seedlings. Additionally, we show that a longer cDNA fragment can be easily recovered by PCR using the SAGE tag sequence as a primer, thereby facilitating the analysis of unknown genes identified by tag sequence in SAGE. In combination with micro‐array analysis, SAGE should serve as a highly efficient tool for the identification and isolation of differentially expressed genes in plants.
Diurnal regulation of Hsp70s in leaf tissue
Authors: Li Qin‐Bao , Haskell Dale , Zhang Chun , Sung Dong‐Yul , Guy Charles 
Published: March 2003

SummarySteady‐state mRNA levels for three Hsp70s were found to be regulated by a distinctive light/dark mechanism in spinach leaves. Messenger RNAs for the chloroplast stromal and two cytosolic forms displayed a diurnal expression pattern under isothermal conditions that appeared to be independent of circadian control. While protein blot data showed relatively constant Hsp70 protein levels, the higher Hsp70 mRNA levels in the light paralleled the diurnal cycle of total cell protein synthesis. Fractionation studies showed that the major cytosolic Hsp70 cognate group was associated with polysomes. Therefore, the variation of Hsp70 mRNAs is consistent with the diurnal metabolic activity of plant photosynthetic cells in which the demand of protein biogenesis for chaperone function and tissue temperature are highest during the day.
Matrix polysaccharide precursors in Arabidopsis cell walls are synthesized by alternate pathways with organ‐specific expression patterns
Authors: Seitz Beate , Klos Christoph , Wurm Marita , Tenhaken Raimund 
Published: March 2003

SummaryThe expression pattern of the single‐copy gene UDP‐glucose dehydrogenase (Ugd) was analysed in transgenic Arabidopsis plants by promoter::GUS and GFP fusions, Western blots, activity assays and histochemical activity staining. The enzyme oxidizes UDP‐glucose to UDP‐glucuronic acid and thus directs carbohydrates irreversibly into a cell wall‐specific pool of nucleotide sugars. UDP‐glucuronic acid is the central intermediate in the interconversion pathway to other nucleotide sugars, including the UDP‐derivatives of arabinose, xylose, apiose and galacturonic acid which account for half the biomass of a typical Arabidopsis leaf cell wall. These activated sugars are needed as substrates for the biosynthesis of matrix polysaccharide polymers. In plants up to 5 days old the Ugd gene is strongly expressed in young roots, but very little in hypocotyls. Older plants show a more uniform expression pattern with a preference for the vascular system. A complex expression pattern was observed in flowers with high activity in the stamen, stigma and nectaries. Meristems in the leaf axil of rosette and inflorescence leaves exhibit a high level of activity of the Ugd gene. Although many of the growing tissues show high activity levels of the Ugd gene, others such as the hypocotyl and the cotyledons of young seedlings do not. Instead these tissues efficiently incorporate 3H‐inositol into their cell walls. This indicates the biosynthesis of UDP‐glucuronic acid through an alternative pathway via the oxidation of inositol to glucuronic acid and subsequent activation to the nucleotide sugar. The data strongly suggest two alternative pathways for matrix polysaccharide precursors with spatial and developmental regulation.
Stringent control of transgene expression in Arabidopsis thaliana using the Top10 promoter system
Authors: Love John , Scott Amie C. , Thompson William F. 
Published: March 2003

SummaryWe show that the tightly regulated tetracycline‐sensitive Top10 promoter system (Weinmann et al. Plant J. 1994, 5, 559–569) is functional in Arabidopsis thaliana. A pure breeding A. thaliana line (JL‐tTA/8) was generated which expressed a chimeric fusion of the tetracycline repressor and the activation domain of Herpes simplex virus (tTA), from a single transgenic locus. Plants from this line were crossed with transgenics carrying the ER‐targeted green fluorescent protein coding sequence (mGFP5) under control of the Top10 promoter sequence. Progeny from this cross displayed ER‐targeted GFP fluorescence throughout the plant, indicating that the tTA–Top10 promoter interaction was functional in A. thaliana. GFP expression was repressed by 100 ng ml−1 tetracycline, an order of magnitude lower than the concentration used previously to repress expression in Nicotiana tabacum. Moreover, the level of GFP expression was controlled by varying the concentration of tetracycline in the medium, allowing a titred regulation of transgenic activity that was previously unavailable in A. thaliana. The kinetics of GFP activity were determined following de‐repression of the Top10::mGFP5 transgene, with a visible ER‐targeted GFP signal appearing from 24 to 48 h after de‐repression.
Cell‐to‐cell movement of TMV RNA is temperature‐dependent and corresponds to the association of movement protein with microtubules
Authors: Boyko Vitaly , Ferralli Jacqueline , Heinlein Manfred 
Published: March 2003

SummaryThe movement protein (MP) of tobacco mosaic virus (TMV) is essential for spread of the viral RNA genome from cell to cell. During infection, the MP associates with microtubules, and it has been proposed that the cytoskeleton transports the viral ribonucleoprotein complex from ER sites of synthesis to plasmodesmata through which infection spreads into adjacent cells. However, microtubule association of MP was observed in cells undergoing late infection rather than in cells undergoing early infection at the leading edge of expanding infection sites where virus RNA cell‐to‐cell spread occurs. Therefore, alternative roles for microtubules in virus infection have been proposed, including a role in MP degradation. To further investigate the role of microtubules in virus pathogenesis, we tested the efficiency of cell‐to‐cell spread of infection and microtubule association of the MP in response to changes in temperature. We show that the subcellular distribution of MP is temperature‐dependent and that a higher efficiency of intercellular transport of virus RNA at elevated temperatures corresponds to an increased association of MP with microtubules early in infection.
rgf1, a mutation reducing grain filling in maize through effects on basal endosperm and pedicel development†
Authors: Maitz Monika , Santandrea Geraldina , Zhang Zhiyong , Lal Shailesh , Hannah L. Curtis , Salamini Francesco , Thompson Richard D. 
Published: March 2003

SummaryThe maize cob presents an excellent opportunity to screen visually for mutations affecting assimilate partitioning in the developing kernel. We have identified a defective kernel mutant termed rgf1, reduced grain filling, with a final grain weight 30% of the wild type. In contrast with most defective endosperm mutants, rgf1 shows gene dosage‐dependent expression in the endosperm. rgf1 kernels possess a small endosperm incompletely filling the papery pericarp, but embryo development is unaffected and the seeds are viable. The mutation conditions defective pedicel development and greatly reduces expression of endosperm transfer layer‐specific markers. rgf1 exhibits striking morphological similarities to the mn1 mutant, but maps to a locus approximately 4 c
m away from mn1 on chromosome 2 of maize. Despite reduced starch accumulation in the mutant, no obvious lesion in starch biosynthesis has been detected. Free sugar levels are unaltered in rgf1 endosperm. Rates of sugar uptake, measured over short (8 h) periods in cultured kernels, are increased in rgf1 compared to the wild type. rgf1 and wild‐type kernels, excised at 5 DAP and cultured in vitro also develop differently in response to variations in sugar regime: glucose concentrations above 1% arrest placentochalazal development of rgf1 kernels, but have no effect on cultured wild‐type kernels. These findings suggest that either uptake or perception of sugar(s) in endosperm cells at 5–10 DAP determines the rgf1 kernel phenotype.
Comparative physical mapping of segments of the genome of Brassica oleracea var. alboglabra that are homoeologous to sequenced regions of chromosomes 4 and 5 of Arabidopsis thaliana
Authors: O'neill Carmel M. , Bancroft Ian 
Published: March 2003

SummaryDue to their relatedness to Arabidopsis thaliana (Arabidopsis), the cultivated Brassica species represent the first group of crops with which to evaluate comparative genomics approaches to understanding biological processes and manipulating traits. We have constructed a high‐quality binary BAC library (JBo) from genomic DNA of Brassica oleracea var. alboglabra, in order to underpin such investigations. Using the Arabidopsis genome sequence and clones from the JBo library, we have analysed aspects of gene conservation and microsynteny between a 222 kb region of the genome of Arabidopsis and homoeologous segments of the genome of B. oleracea. All 19 predicted genes tested were found to hybridize to clones in the JBo library, indicating a high level of gene conservation. Further analyses and physical mapping with the BAC clones identified allowed us to construct clone contig maps and analyse in detail the gene content and organization in the set of paralogous segments identified in the genome of B. oleracea. Extensive divergence of gene content was observed, both between the B. oleracea paralogous segments and between them and their homoeologous segment within the genome of Arabidopsis. However, the genes present show highly conserved collinearity with their orthologues in the genome of Arabidopsis. We have identified one example of a Brassica gene in a non‐collinear position and one rearrangement. Some of the genes not present in the discernible homoeologous regions appear to be located elsewhere in the B. oleracea genome. The implications of our findings for comparative map‐based cloning of genes from crop species are discussed.