bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular comparison of monocot and dicot U1 and U2 snRNAs
Authors: Musci Michael A. , Egeland Daniel B. , Schuler Mary A. 
Published: March 2003

SummaryTo elucidate differences between the pre‐mRNA splicing components in monocots and dicots, we have cloned and characterized several U1 and U2 snRNA sequence variants expressed in wheat seedling nuclei. Primer extension sequencing on wheat and pea snRNA populations has demonstrated that two 5′‐ terminal nucleotides found in most other U1 snRNAs are missing/modified in many plant U1 snRNAs. Comparison of the wheat U1 and U2 snRNA variants with their counterparts expressed in pea nuclei has defined regions of structural divergence between monocot and dicot U1 and U2 snRNAs. The U1 and U2 snRNA sequences involved in RNA:RNA interaction with pre‐mRNAs are absolutely conserved. Significant differences occur between wheat and pea U1 snRNAs in stem I and II structures implicated in the binding of U1‐specific proteins suggesting that the monocot and dicot U1‐specific snRNP proteins differ in their binding specificities. Stem III structures, which are required in mammalian systems for splicing complex formation but not for U1‐specific protein binding, differ more extensively than stems I, II, or IV. In U2 snRNAs, the sequence differences between these two species are primarily localized in stem III and in stem IV which has been implicated in snRNP protein binding. These differences suggest that monocot and dicot U1 and U2 snRNPs represent distinct entities that may have monocot‐ and dicot‐specific snRNP protein variants associated with each snRNA.
PCR‐amplified microsatellites as markers in plant genetics
Authors: Morgante M. , Olivieri A.M. 
Published: March 2003

SummaryIn order to assess the feasibility of using microsatellites as markers in plant genetics, a survey of published DNA sequence data for presence, abundance and ubiquity in higher plants of all types of dinucleotide and trinucleotide repeats with a minimum number of 10 and 7 units, respectively, was conducted. This search revealed that such microsatellites are frequent and widely distributed; they were uncovered in 34 species, with a frequency of one every 50 kb. AT repeats were by far the most frequently observed class of dinucleotide microsatellites, whereas AC/TG repeats, which are common in animals, were observed only once. TAT repeats prevailed among trinucleotides. Polymerase chain reaction amplification of (AT)n and (TAT)n micro‐satellites in soybean (Glycine max (L.) Merr.) revealed that they are highly polymorphic, as a consequence of length variation, somatically stable and inherited in a co‐dominant Mendelian manner. The abundance and amount of information derived from such markers, together with the ease by which they can be identified, make them ideal markers for plant genetic linkage and physical mapping, population studies and varietal identification.
An Arabidopsis thaliana RFLP mapping set to localize mutations to chromosomal regions
Authors: Fabri Christoph O. , Schäffner Anton R. 
Published: March 2003

SummaryMapping of newly identified Arabidopsis thaliana mutants is an important step towards their molecular characterization and the attempt to saturate the genome by known mutations. The classical genetic analysis using phenotypic tester lines is well‐established, but laborious, time‐consuming and potentially ambiguous. An alternative molecular strategy was developed that is based on RFLPs. Subcloned DNA markers that detect only segregating RFLP bands distinguishing A. thaliana ecotype Landsberg from Columbia or Enkheim after EcoRI restriction digestion compose an Arabidopsis RFLP mapping set (ARMS). Up to 13 markers uniformly cover the five A. thaliana chromosomes and can be scored in only two successive Southern experiments on a single blot without mutual interference of the signals. Thus, this system allows a simple, reliable, rapid and especially inexpensive mapping of any monogenic mutant locus to the A. thaliana chromosomes. Several loci can be analysed in one experiment if the respective blots are hybridized together. This paper demonstrates the mapping of two recessive mutants affecting the development of A. thaliana leaves which had been generated in the Columbia and Enkheim ecotype by analysing less than 20 F2 individuals. Further markers to refine or verify the result on the same blot can be chosen out of 14 additional probes detecting single segregating EcoRI polymorphic bands as well.
Correlation between the activities of five ribosome‐inactivating proteins in depurination of tobacco ribosomes and inhibition of tobacco mosaic virus infection
Authors: Taylor Sally , Massiah Andrea , Lomonossoff George , Roberts Lynne M. , Lord J. Michael , Hartley Martin 
Published: March 2003

SummaryThe rRNA depurination activities of five ribosome‐inactivating proteins (RIPs) were compared in vitro using yeast and tobacco leaf ribosomes as substrates. All of the RIPs (pokeweed antiviral protein (PAP), dianthin 32, tritin, barley RIP and ricin A‐chain) were active on yeast ribosomes. PAP and dianthin 32 were highly active and ricin A‐chain weakly active on tobacco ribosomes, whereas tritin and barley RIP were inactive. PAP and dianthin 32 were highly effective in inhibiting the formation of local lesions caused by tobacco mosaic virus (TMV) on tobacco leaves, whereas tritin, barley RIP and ricin A‐chain were ineffective. The apparent anomaly between the in vitro rRNA depurination activity, but lack of antiviral activity of ricin A‐chain was further investigated by assaying for rRNA depurination in situ following the topical application of the RIP to leaves. No activity was detected, a finding consistent with the apparent lack of antiviral activity of this RIP. Thus, it is concluded that there is a positive correlation between RIP‐catalysed depurination of tobacco ribosomes and antiviral activity which gives strong support to the hypothesis that the antiviral activity of RIPs works through ribosome inactivation.
Spatial patterns of expression of flavonoid/isoflavonoid pathway genes during interactions between roots of Medicago truncatula and the mycorrhizal fungus Glomus versiforme
Authors: Harrison Maria J. , Dixon Richard. A. 
Published: March 2003

SummaryA combination of Northern blot analysis and in situ hybridization was used to measure and localize changes in the levels of transcripts encoding five enzymes of phenylpropanoid/flavonoid/isoflavonoid metabolism in Medicago truncatula roots following colonization with the mycorrhizal fungus Glomus versiforme. In colonized roots, elevated levels of phenylalanine ammonia‐lyase (PAL) and chalcone synthase (CHS) transcripts were detected specifically in the cortical cells containing arbuscules. This localization was discrete, and elevated levels of transcripts were not observed in adjacent, non‐colonized cells. In contrast, isoflavone reductase (IFR) transcripts, which were detected at relatively high levels in the cortical cells of non‐colonized roots, were almost undetectable in the colonized cortical cells containing arbuscules and were also lowered in adjacent cells. The expression of chalcone isomerase (CHI) and isoliquiritigenin 2′‐O‐methyltransferase (ChalOMT) was unaffected by colonization, and the tissue‐specific patterns of gene expression were the same as in non‐colonized roots. It is concluded that the establishment of the mycorrhizal interaction results in cell type‐specific differential expression of genes of phenylpropanoid/flavonoid/isoflavonoid biosynthesis, which may be causally related to arbuscule development.
The developmental expression of the asparagus intracellular PR protein (AoPR1) gene correlates with sites of phenylpropanoid biosynthesis
Authors: Warner Simon A.J. , Gill Aideen , Draper John 
Published: March 2003

SummaryPrevious reports have described the induction, by either wounding or attempted pathogen invasion, of an Asparagus officinalis intracellular pathogenesis‐related (AoPR1) promoter—GUS gene fusion in transgenic tobacco. Here we describe the unexpected developmental expression pattern of the AoPR1—GUS gene which correlates well, temporally and spatially, with the developmental expression observed for GUS fusions with promoters derived from genes coding for enzymes in the ‘core phenylpropanoid pathway’. Analysis of endogenous AoPR1 gene expression in asparagus and both AoPR1—GUS and AoPR1—luciferase gene fusions in transgenic tobacco suggests that the AoPR1 promoter directs similar cell‐specific transcription patterns in both asparagus and transgenic tobacco. The AoPR1 promoter contains sequence motifs similar to those implicated as important in the regulation of phenylpropanoid pathway genes and another ‘intracellular’ PR gene. Treatment with salicylic acid enhances AoPR1 promoter gene activity both in tobacco and in asparagus.
Functional reconstitution of the solubilized Arabidopsis thaliana STP1 monosaccharide‐H+ symporter in lipid vesicles and purification of the histidine tagged protein from transgenic Saccharomyces cerevisiae
Authors: Stolz Jürgen , Stadler Ruth , Opekarová Miroslava , Sauer Norbert 
Published: March 2003

SummaryComplete DNA sequences encoding the Arabidopsis thaliana STP1 monosaccharide/H+ symporter or a histidine‐tagged STP1‐His6 protein were expressed in baker's yeast Saccharomyces cerevisiae. Both wild‐type STP1 and the recombinant his‐tagged protein were located in the plasma membranes of transformed yeast cells. The C‐terminal modification caused no loss of transport activity compared with the wild‐type protein. Anti‐STP1‐antibodies were used to confirm the identity of the protein in yeast and to compare the apparent molecular weights of STP1 proteins in membrane extracts from yeast or Arabidopsis thaliana. Purified yeast plasma membranes were fused with proteoliposomes consisting of Escherichia coli lipids and beef heart cytochrome‐c oxidase. Addition of ascorbate/TMPD/cytochrome‐c to these fused vesicles caused an immediate formation of membrane potential (inside negative; monitored with [3H]tetraphenylphosphonium cations) and a simultaneous, uncoupler‐sensitive influx of d‐glucose into the energized vesicles. STP1‐His6 protein is functionally active after solubilization with octyl‐β‐d‐glucoside, which was shown by insertion of the protein into proteoliposomes by detergent dilution and determination of the resulting transport capacity. Detergent extracts from either total membranes or plasma membranes of transgenic yeast cells were used for one‐step purification of the STP1‐His6 protein on Ni2+‐NTA columns. The identity of the purified protein was checked by immunoblotting and N‐terminal sequencing.
Analysis of phytochrome‐ and ABA‐deficient mutants suggests that ABA degradation is controlled by light in Nicotiana plumbaginifolia
Authors: Kraepiel Yvan , Rousselin Philippe , Sotta Bruno , Kerhoas Lucien , Einhorn Jacques , Caboche Michel , Miginiac Emile 
Published: March 2003

SummaryThe phytochrome chromophore‐deficient mutant, pew1, of Nicotiana plumbaginifolia exhibited decreased germination and slower dehydration of detached leaves during water stress as compared with the wild‐type. These physiological processes are controlled by abscisic acid (ABA) and we examined, therefore, whether phytochrome plays a specific role in the regulation of ABA metabolism using the pew1 mutant. The ABA contents of mature seeds and young leaves were analysed and in both cases mutant material was found to contain higher amounts of ABA as compared with the wild‐type. This indicates that the phytochrome activation can lead to a decrease of the ABA level in the wild‐type plant. The role of phytochromes was investigated in greater detail using the ABA‐deficient mutant aba1 of N. plumbaginifolia exhibiting an early and synchronous germination. This mutant accumulates at very high levels a metabolite derived from a precursor (ABA‐aldehyde) in the ABA biosynthetic pathway. The first biochemical characterization of this molecule, which corresponds to the glucose‐conjugated ABA‐alcohol (ABA‐AG) is described. A pew1‐aba1 double mutant exhibiting both an etiolated growth and early germination was also obtained. The comparable accumulation of ABA‐AG in the pew1‐aba1 double mutant as compared with the aba1 mutant allowed the proposition that, in a wild‐type plant, the phytochrome‐mediated light signal enhances ABA degradation rather than inhibits its biosynthesis.
Starch and fatty acid synthesis in plastids from developing embryos of oilseed rape (Brassica napus L.)
Authors: Kang Fan , Rawsthorne Stephen 
Published: March 2003

SummaryThe aim of this work was to investigate the capacity for synthesis of starch and fatty acids from exogenous metabolites by plastids from developing embryos of oilseed rape (Brassica napus L.). A method was developed for the rapid isolation from developing embryos of intact plastids with low contamination by cytosolic enzymes. The plastids contain a complete glycolytic pathway, NADP‐glucose‐6‐phosphate dehydrogenase, NADP‐6‐phosphogluconate dehydrogenase, fructose‐1,6‐bisphosphatase, NADP‐malic enzyme, the pyruvate dehydrogenase complex (PDC), and acetyl‐CoA carboxylase. Organelle fractionation studies showed that 67% of the total cellular PDC activity was in the plastids. The isolated plastids were fed with 14C‐labelled carbon precursors and the incorporation of 14C into starch and fatty acids was determined. 14C from glucose‐6‐phosphate (G‐6‐P), fructose, glucose, fructose‐6‐phosphate and dihydroxyacetone phosphate (DHAP) was incorporated into starch in an intactness‐ and ATP‐dependent manner. The rate of starch synthesis was highest from G‐6‐P, although fructose gave rates which were 70% of those from G‐6‐P. Glucose‐1‐phosphate was not utilized by intact plastids for starch synthesis. The plastids utilized pyruvate, G‐6‐P, DHAP, malate and acetate as substrates for fatty acid synthesis. Of these substrates, pyruvate and G‐6‐P supported the highest rates of synthesis. These studies show that several cytosolic metabolites may contribute to starch and/or fatty acid synthesis in the developing embryos of oilseed rape.
The axr4 auxin‐resistant mutants of Arabidopsis thaliana define a gene important for root gravitropism and lateral root initiation
Authors: Hobbie Lawrence , Estelle Mark 
Published: March 2003

SummaryTo understand the molecular mechanism of auxin action, mutants of Arabidopsis thaliana with altered responses to auxin have been identified and characterized. Here the isolation of two auxin‐resistant mutants that define a new locus involved in auxin response, named AXR4, is reported. The axr4 mutations are recessive and map near the ch1 mutation on chromosome 1. Mutant plants are specifically resistant to auxin and defective in root gravitropism. Double mutants between axr4 and the recessive auxin‐resistant mutants axr1‐3 and aux1‐7 were characterized to ascertain possible genetic interactions between the mutations. The roots of the axr4 axr1‐3 double mutant plants are less sensitive to auxin, respond more slowly to gravity, and form fewer lateral roots than either parental single mutant. These results suggest that the two mutations have additive or even synergistic effects. The AXR1 and AXR4 gene products may therefore act in separate pathways of auxin response or perhaps perform partially redundant functions in a single pathway. The axr4 aux1‐7 double mutant has the same sensitivity to auxin as the aux1‐7 mutant but forms far fewer lateral roots than either parental single mutant. The aux1‐7 mutation thus appears to be epistatic to axr4 with respect to auxin‐resistant root elongation, whereas in lateral root formation, the effects of the two mutations are additive. The complexity of the genetic interactions indicated by these results may reflect differences in the mechanism of auxin action during root elongation and the formation of lateral roots. The AXR4 gene product, along with those of the AXR1 and AUX1 genes, is important for normal auxin sensitivity, gravitropic response in roots and lateral root formation.
A constituent of the chloroplast import complex represents a new type of GTP‐binding protein
Authors: Seedorf Matthias , Waegemann Karin , Soll Jürgen 
Published: March 2003

SummaryThe 34 kDa polypeptide of the outer envelope membranes from pea chloroplasts (OEP 34) is a major constituent of this membrane. OEP 34 is detected on polyacrylamide gels under non‐reducing condition in association with OEP 75, the putative protein translocation pore. An antiserum against OEP 34 is able to co‐immunoprecipitate the precursor of Rubisco small subunit from a partially purified import complex of chloroplast outer envelope membranes. A full‐length cDNA clone coding for pea OEP 34 has been isolated. Analysis of the deduced amino acid sequence revealed typical and conserved sequence motifs found in GTP‐binding proteins, making it a new and unique member of this superfamily. OEP 34 behaves as an integral constituent of the outer chloroplast envelope, which is anchored by its C‐terminus into the membrane, while the majority of the protein projects into the cytoplasm. OEP 34 does not possess a cleavable N‐terminal transit sequence but it is targeted to the chloroplasts and integrated into the outer membranes by internal sequence information which seems to be present in the C‐terminal membrane anchor region. Productive integration of OEP 34 into the outer envelope requires, in contrast to other OEPs, protease‐sensitive chloroplast surface components and is stimulated by ATR. The GTP binding specificity of OEP 34 is demonstrated by photo‐affinity labelling in the presence of [α‐32P]GTP. Overexpressed and purified OEP 34 possesses endogenous GTPase activity. These results indicate a possible regulatory function of OEP 34 in protein translocation into chloroplasts.
Altered lignin composition in transgenic tobacco expressing O‐methyltransferase sequences in sense and antisense orientation
Authors: Atanassova Rossitza , Favet Noëlle , Martz Françoise , Chabbert Brigitte , Tollier Marie‐Thérèse , Monties Bernard , Fritig Bernard , Legrand Michel 
Published: March 2003

SummaryThe monomeric composition of tobacco lignin has been modified by genetic engineering. Sense or antisense expression of sequences encoding O‐methyltransferase (OMT), a lignin biosynthetic enzyme, was shown to modulate enzyme activity. Ten constructs harboring the entire or a partial OMT cDNA were used. Populations of 20 trensgenic plants per construct were analyzed for OMT activity and compared with untransformed controls. As expected, expression of only the full‐length sense construct led to an increase in OMT activity. An important reduction of activity was found in a variable number of plantlets from all other transgenic populations but the inhibition was sustained through the adult stage only in plants transformed with the complete cDNA. T‐DNA genes were shown to be stably integrated into the tobacco genome and to be transmitted to the progeny. By using gene‐specific probes, OMT inhibition in stems was correlated to a parallel disappearance of OMT transcripts originating from both the resident gene and the transgene. In contrast, transgene transcripts were detected in leaf tissues where the resident gene is poorly expressed, thus indicating that relative expression of the two OMT genes controls transcript turnover. In stems of inhibited plants, a marked decrease of syringyl units and the appearance of 5‐hydroxy guaiacyl units were demonstrated. These two structural features are also characteristic of natural mutants of maize with an improved digestibility compared with wild lines. These data demonstrate the feasibility and the potential benefits of lignin manipulation.
Low threshold levels of ultraviolet‐B in a background of photosynthetically active radiation trigger rapid degradation of the D2 protein of photosystem‐II
Authors: Jansen Marcel A.K. , Gaba Victor , Greenberg Bruce M. , Mattoo Autar K. , Edelman Marvin 
Published: March 2003

SummaryThe photosystem II reaction centre has at its core a heterodimer made up of two proteins, D1 and D2. The D1 protein is known to be rapidly degraded by photosyn‐thetically active radiation while the D2 protein is relatively stable. This paper reports that when the aquatic higher plant, Spirodela was exposed to ultraviolet‐B radiation, D2 degradation accelerated markedly and half life times approached those of the D1 protein. Moreover, in the presence of an environmentally relevant background of photosynthetically active radiation, low fluxes of ultraviolet‐B (but not ultraviolet‐A) radiation synergistically stimulated degradation of the D2 protein within functional reaction centres. Thus, above a critical threshold, ultraviolet‐B specifically targets the D1–D2 heterodimer for accelerated degradation.
Isolation of early genes expressed in reproductive organs of the dioecious white campion (Silene latifolia) by subtraction cloning using an asexual mutant
Authors: Barbacar Nicolae , Hinnisdaels Stefan , Farbos Isabelle , Monéger Françoise , Lardon André , Delichère Catherine , Mouras Armand , Negrutiu Ioan 
Published: March 2003

SummaryThe dioecious white campion (Silene latifolia) has been chosen as a working model for sexual development. In this species, sexual dimorphism is achieved through two distinct developmental blocks: inhibition of carpel development in male flowers, and early arrest of anther differentiation in female flowers. The combined advantages of the dioecious system and the availability of a sexual mutant lacking both male and female reproductive organs have been exploited in a molecular subtraction approach using male and asexual flower buds. This resulted in the cloning of 22 cDNA clones expressed in stamens at distinct stages of development. Fourteen of these clones corresponded to genes whose expression was detected in pre‐meiotic stamens, a stage of development for which very little information is presently available. Furthermore, the absence of similarities with database sequences for ten clones suggests that they represent novel genes. Functional analysis of each clone will enable their positioning within the reproductive organ developmental pathway(s). In parallel, these clones are being exploited as developmental markers of early differentiation within the flower.
Cell‐to‐cell movement of potato spindle tuber viroid
Authors: Ding Biao , Kwon Myoung‐Ok , Hammond Rosemarie , Owens Robert 
Published: March 2003

SummaryViroids are non‐translatable, autonomously replicating circular RNAs that infect only plants. An important component of the viroid infection process is cell‐to‐cell movement; however, there is virtually no information available about the pathways and mechanisms of this process. In this study, potato spindle tuber viroid (PSTVd) has been used as a model system to investigate the mechanism of viroid cell‐to‐cell transport. Infectious RNA transcripts were produced from PSTVd cDNA clones in vitro, labeled with the nucleotide‐specific fluorescent dye TOTO‐1 iodide, and used for micro‐injection. When injected into symplasmically isolated guard cells of mature tomato and tobacco leaves, PSTVd remained in the injected cells; in contrast, PSTVd injected into symplasmically connected mesophyll cells moved rapidly from cell to cell. A 1400 nt RNA containing only vector sequences was unable to move out of the injected mesophyll cells, but when PSTVd was fused to this transcript, the fusion RNA moved from cell to cell. At the DNA level, PSTVd cDNA also appears able to mediate cell‐to‐cell movement of plasmid DNA. These data indicate that (i) PSTVd moves from cell to cell via plasmodesmata, and (ii) this movement may be mediated by a specific sequence or structural motif.