bluuebunny/crossref_metadata_2025_split huggingface dataset:
Regulation of expression of a cDNA from barley roots encoding a high affinity sulphate transporter
Authors: Smith Frank W. , Hawkesford Malcolm J. , Ealing Paul M. , Clarkson David T. , Vanden Berg Peter J. , Belcher Ann R. , Warrilow Andrew G.S. 
Published: March 2003

SummaryA cDNA encoding a high‐affinity sulphate transporter has been isolated from barley by complementation of a yeast mutant. The cDNA, designated HVST1, encodes a polypeptide of 660 amino acids (Mr = 72 550), which is predicted to have 12 membrane‐spanning domains and has extensive sequence homology with other identified eukaryotic sulphate transporters. The Km for sulphate was 6.9 µM when the HVST1 cDNA was expressed in a yeast mutant deficient in the gene encoding for the yeast SUL1 sulphate transporter. The strong pH‐dependency of sulphate uptake when HVST1 was expressed heterologously in yeast suggests that the HVST1 polypeptide is a proton/sulphate co‐transporter. The gene encoding HVST1 is expressed specifically in root tissues and the abundance of the mRNA is strongly influenced by sulphur nutrition. During sulphur‐starvation of barley, the abundance of mRNA corresponding to HVST1, and the capacity of the roots to take up sulphate, both increase. Upon re‐supply of sulphate, the abundance of the mRNA corresponding to HVST1, and the capacity of the roots to take up sulphate, decrease rapidly, concomitant with rises in tissue sulphate, cysteine and glutathione contents. Addition of the cysteine precursor, O‐acetylserine, to plants grown with adequate sulphur supply, leads to increases in sulphate transporter mRNA, sulphate uptake rates and tissue contents of glutathione and cysteine. It is suggested, that whilst sulphate, cysteine and glutathione may be candidates for negative metabolic regulators of sulphate transporter gene expression, this regulation may be overridden by O‐acetylserine acting as a positive regulator.
Transposon sequences drive tissue‐specific expression of the maize regulatory gene R‐s
Authors: May Bruce P. , Dellaporta Stephen L. 
Published: March 2003

SummaryThe R family of genes in maize encodes transcription factors that activate anthocyanin synthesis. The R‐r complex at the r locus contains two components, of which one, the aleurone‐pigmenting R‐s subcomplex, is an unusual promoter flanked by two head‐to‐head, nearly identical coding regions. The central region of the promoter, designated σ, contains inverted terminal repeats and subterminal repeats, both of which are features of the CACTA transposon family. Transcription from the R‐s promoter was strongest in aleurone but transcripts had a surprisingly variable initiation site and splicing pattern. Microprojectile bombardment indicated that the sequences introduced by transposition act as an enhancer of transcription in aleurone but not in coleoptile or mesocotyl tissue. Moreover, the region containing the subterminal repeats was able to activate expression when fused to a minimal promoter, suggesting that a factor that binds to the repeats may be responsible. Regions downstream of σ encode a leader containing three ORFs, two of which are variably removed by splicing. The results presented here show that transposons can mediate the rearrangement of genes and confer new splicing and expression patterns. In the case of the r complex, this has resulted in a new phenotype, the pigmentation of aleurone cells.
Transgenic analysis of tomato endo‐β‐1,4‐glucanase gene function. Role of cel1 in floral abscission
Authors: Lashbrook Coralie C. , Giovannoni James J. , Hall Bradford D. , Fischer Robert L. , Bennett Alan B. 
Published: March 2003

SummaryEndo‐β‐1,4‐glucanase (EGase) expression in ripening tomato fruit and abscising tomato flowers reflects the overlapping accumulation of structurally divergent gene transcripts, including EGase1 (Cel1) and EGase2 (Cel2). Transgenic plants in which cel1 expression was specifically inhibited were evaluated in order to assess the potential role(s) played by Cel1 during fruit ripening and floral abscission. The constitutive expression of an antisense cel1 gene driven by the 35S promoter of cauliflower mosaic virus resulted in the reduction of Cel1 mRNA to trace levels in both ripening fruit and abscising abscission zones of transgenic tomato plants. Transgenic fruit in which cel1 expression was reduced to less than 0.1% of wild‐type levels exhibited normal growth and softening behaviour. Transgenic floral abscission zones expressing 5–7% of wild‐type levels of Cel1 mRNA exhibited an incidence of abscission reduced by up to one third. The incomplete loss of abscission ability by transgenic flowers may indicate that the contribution of Cel1 activity is insufficient to account for all floral abscission and attests to the presence in tomato abscission zones of additional cell wall hydrolase activities, including Cel2.
Two bZIP proteins from Antirrhinum flowers preferentially bind a hybrid C‐box/G‐box motif and help to define a new sub‐family of bZIP transcription factors
Authors: Martínez‐García Jaime F. , Moyano Enriqueta , Alcocer Marcos J. C. , Martin Cathie 
Published: March 2003

SummaryTwo genes encoding bZIP proteins are expressed in flowers of Antirrhinum majus, predominantly in vascular tissues, carpels and anthers. The sequences of cDNA clones encoding these proteins show them to belong to a distinct sub‐family of bZIP proteins which also includes LIP19 from rice and MLIPI5 and OBF1 from maize. The sub‐family is characterized by the inclusion of very small proteins consisting of essentially a basic domain and a long leucine zipper. Members also have a conserved upstream open reading frame (uORF) in their 5′ leader sequences, implying a common mode of post‐transcriptional control. In vitro, the Antirrhinum bZIP proteins preferentially bind to a novel hybrid C‐box/G‐box motif which is found in the promoters of some plant histone genes and of some nuclear‐encoded genes with plastidial protein products. Expression of the bZIP proteins in transgenic tobacco under control of the CaMV 35S promoter supports the view that they can regulate expression of genes which contain the preferred target motif within their regulatory sequences, although both enhancement and repression of transcript levels of target genes were observed, indicating that the bZIP proteins probably interact with other factors to modulate transcription in different ways, as has been observed for the small MAF family of bZIP proteins in vertebrates.
Low temperature signal transduction during cold acclimation: protein phosphatase 2A as an early target for cold‐inactivation
Authors: Monroy Antonio F. , Sangwan Veena , Dhindsa Rajinder S. 
Published: March 2003

SummaryThe authors have previously shown that cold acclimation and cold acclimation‐specific (cas) gene expression in alfalfa require cold‐triggered calcium influx and phosphorylation of specific pre‐existing proteins. In this study, the authors used the expression of cas15 gene as an end‐point marker to examine the role of protein phosphorylation in low temperature signal transduction in alfalfa cells. Whereas the protein kinase inhibitor staurosporine prevented the cold induction of cas15, the protein phosphatase inhibitor okadaic acid induced the cas15 at 25°C. Upon exposure of cells to cold, total cellular protein phosphatase activity rapidly declined by 30% but all of this decline could be attributed to an almost complete inhibition of protein phosphatase 2A (PP2A). This cold‐inactivation of PP2A was found to be mediated by calcium influx and could be reproduced at 25°C by treating the cells with the calcium ionophore A23187 or with the calcium channel agonist Bay K8644. On exposure of cells to cold, the transcript and protein levels of the PP2A catalytic subunit (PP2Ac) did not decline, but the binding of anti‐PP2Ac antibody to native PP2Ac increased, indicating a demasking of the PP2Ac epitope. The possible mechanisms by which cold might inhibit PP2A are discussed.
Molecular cloning and functional analysis of the Arabidopsis thaliana DNA ligase I homologue
Authors: Taylor Richard M. , Hamer Marcus J. , Rosamond John , Bray Clifford M. 
Published: March 2003

SummaryA cDNA encoding the DNA ligase I homologue has been isolated from Arabidopsis thaliana using a degenerate PCR approach. The ORF of this cDNA encodes an amino acid sequence of 790 residues, representing a protein with a theoretical molecular mass of 87.8 kDa and an isoelectric point (pI) of 8.20. Alignment of the A. thaliana DNA ligase protein sequence with the sequence of DNA ligases from human (Homo sapiens), murine (Mus musculus), clawed toad (Xenopus laevis) and the yeasts Schizosaccharomyces pombe and Saccharomyces cerevisiae showed good sequence homology (42–45% identity, 61–66% similarity), particularly around the active site. Sequence data indicate that the Arabidopsis DNA ligase is the homologue of the animal DNA ligase I species. Functional analysis of the cDNA clone demonstrated its ability to complement the conditional lethal phenotype of an S. cerevisiae cdc9 mutant defective in DNA ligase activity, confirming that the cloned sequence encodes an active DNA ligase. The level of the DNA ligase transcript was not increased in A. thaliana seedlings in response to DNA damage induced by a period of enhanced UV‐B irradiation. However, the cellular level of the DNA ligase mRNA transcript does correlate with the replicative state of plant cells.
Targeting and modification of prokaryotic cell–cell junctions by tobacco mosaic virus cell‐to‐cell movement protein
Authors: Heinlein Manfred , Wood Malcolm R. , Thiel Teresa , Beachy Roger N. 
Published: March 2003

SummaryThe movement protein (MP) of tobacco mosaic virus (TMV) facilitates the cell‐to‐cell spread of infection by altering the structure and function of plasmodesmata, the intercellular communication channels in plants. Because the protein was shown to interfere with intercellular communication when expressed in the cyanobacterium Anabaena sp. strain PCC 7120, whether the ability of the protein to target and to modify intercellular communication channels in plants is conserved in this prokaryote was investigated. It was found that the MP localizes to the cell junctions and induces the formation of filamentous structures that traverse the septa. It is proposed that the protein interacts with host components that are similar between plants and Anabaena and that may be evolutionarily related. The observations in Anabaena suggest that the MP modifies plasmodesmata by forming a filamentous aggregate within the pore.
Homology modelling and molecular dynamics aided analysis of ligand complexes demonstrates functional properties of lipid‐transfer proteins encoded by the barley low‐temperature‐inducible gene family, blt4
Authors: Keresztessy Zsolt , Hughes Monica A. 
Published: March 2003

SummaryThe homology modelling technique was used to predict the tertiary structures of three members of the low‐temperature‐inducible barley vegetative shoot epidermal lipid‐transfer protein (LTP) family, BLT4, on the basis of the X‐ray crystallographically determined three‐dimensional structure of a maize seedling LTP. Differences between the maize LTP and the BLT4 family include amino acid substitutions around the entrance and inside the predicted hydrophobic binding tunnels of these proteins. Because of the deletion of the loop region corresponding to Val60–Gly62 of the maize LTP from all three BLT4 LTPs, their internal hydrophobic tunnels are longer. Molecular dynamics modelling shows that BLT4.9 can accommodate hexadecanoic acid in its binding tunnel in similar conformation to the maize LTP. However, modelled cis,cis‐9,12‐octadecandienoic acid had a more favourable interaction with the BLT4.9 LTP than with the maize protein. Di‐cis,cis‐9,12‐octadecandienoyl phosphatidylglycerol and di‐cis,cis‐9,12‐octadecandienoyl phosphatidylcholine were modelled in the BLT4.9 structure with the fatty acyl group at position 1 embedded in the binding tunnel and the group at position 2 located on the solvent accessible surface of the protein. The results of the modelling suggest that the phospholipid headgroup can form hydrogen and salt bridges with polar and charged residues outside the binding tunnel and the exposed hydrocarbon chain interacts with hydrophobic amino acids on the surface. These results are consistent with the proposal that BLT4 LTPs have a lipid‐transfer function associated with frost acclimation in barley.
Brown‐midrib maize (bm1) – a mutation affecting the cinnamyl alcohol dehydrogenase gene
Authors: Halpin Claire , Holt Karen , Chojecki Jan , Oliver Duncan , Chabbert Brigitte , Monties Bernard , Edwards Keith , Barakate Abdellah , Foxon Geoffrey A. 
Published: March 2003

Summary Brown‐midrib (bm) mutants of maize have modified lignin of reddish‐brown colour. Although four independent bm loci are known, only one of the mutant genes has been previously identified. We report here that maize bm1, one of the less characterised mutants, shows severely reduced CAD activity in lignified tissues, resulting in the production of a modified lignin. Both the total lignin content and the structure of the polymer are altered by the mutation. We further describe the isolation and characterisation of the maize CAD cDNA and mapping of the CAD gene. CAD maps very closely to the known location of bm1 and co‐segregates with the bm1 locus in two independent recombinant inbred populations. These data strongly support the premise that maize bm1 directly affects expression of the CAD gene.
Molecular characterisation of a novel plant homeobox gene expressed in the maturing xylem zone ofPopulus tremula× tremuloides
Authors: Hertzberg Magnus , Of Olsson Ol 
Published: March 2003

SummaryIn trees, wood is produced from the vascular cambium. However, little is known about how differentiation of the cambial derivatives is controlled at the molecular level. Hybrid aspen,Populus tremula × tremuloides, is a fast‐growing, wood‐forming plant from which an efficient transformation and regeneration protocol has been developed. We use this species as a model to study xylem cell differentiation. In a cross‐section of the stem, cells in the cambial region at different stages of development can be seen in a radial distribution. We dissected out the cambial region and constructed a cDNA‐library from this tissue. The library was screened with a degenerated primer mix complementary to the third helix region of the homeobox. A number of cDNAs hybridising to the probe were isolated, purified, cloned and sequenced. Characterisation of the expression pattern of two such homeobox cDNAs revealed that one of the cDNAs, denotedPttHB1, was specifically expressed in the xylem maturation zone, indicating a role in the regulation of secondary wall formation, while the other, denotedPttHB2was expressed in differentiating xylem and phloem cells. Structurally, the two hybrid aspen homeobox genes are homologous to each other, but no close relationship could be identified between these genes and any other homeobox sequence described thus far, in plants or other eukaryotes. We therefore conclude that thePttHB1andPttHB2genes belong to a novel hitherto undescribed sequence class of homeobox genes. We designate this as PALE.
cDNA cloning, gene expression and subcellular localization of anthocyanin 5‐aromatic acyltransferase fromGentiana triflora
Authors: Fujiwara Hiroyuki , Tanaka Yoshikazu , Yonekura‐Sakakibara Keiko , Fukuchi‐Mizutani Masako , Nakao Masahiro , Fukui Yuko , Yamaguchi Masaatsu , Ashikari Toshihiko , Kusumi Takaaki 
Published: March 2003

Summary Acylation of anthocyanins with hydroxycinnamic acid derivatives is one of the most important and less understood modification reactions during anthocyanin biosynthesis. Anthocyanin aromatic acyltransferase catalyses the transfer of hydroxycinnamic acid moieties from their CoA esters to the glycosyl groups of anthocyanins. A full‐length cDNA encoding the anthocyanin 5‐aromatic acyltransferase (5AT) (
EC 2.3.1.153
) that acylates the glucose bound at the 5‐position of anthocyanidin 3,5‐diglucoside was isolated from petals of

Gentiana triflora

on the basis of the amino acid sequence of the purified enzyme. The isolated full‐length cDNA had an open reading frame of 469 amino acids and the calculated molecular weight was 52 736. The deduced amino acid sequence contains consensus motifs that are conserved among the putative acyl CoA‐mediated acyltransferases, and this indicates that 5AT is a member of a proposed superfamily of multifunctional acyltransferases (

St‐Pierre

et al

. (1998

)

Plant J.

14, 703–713). The cDNA was expressed in

Escherichia coli

and yeast, and confirmed to encode 5AT. The enzymatic characteristics of the recombinant 5AT were consistent with those of the native gentian 5AT. Immunoblot analysis using specific antibodies to 5AT showed that the 5AT protein is present in petals, but not in sepals, stems or leaves of

G. triflora

. RNA blot analysis showed that the

5AT

gene is expressed only in petals and that its expression is temporally regulated during flower development coordinately with other anthocyanin biosynthetic genes. Immunohistochemical analysis demonstrated that the 5AT protein is specifically expressed in the outer epidermal cells of gentian petals and that it is localized mainly in the cytosol.
Transgenic plants changed in carbon allocation pattern display a shift in diurnal growth pattern
Authors: Kehr Julia , Hustiak Frauke , Walz Christina , Willmitzer Lothar , Fisahn Joachim 
Published: March 2003

Summary Photosynthesis, partitioning of carbohydrates and growth have to be highly orchestrated to enable an efficient performance of plants. To study the diurnal relationships between carbon distribution and growth, we analysed transgenic potato plants with altered carbon allocation patterns. To modify carbohydrate supply of growing sinks, we used plants that accumulated starch as a consequence of inhibition in triose‐phosphate export from chloroplasts and plants that were genetically inhibited in starch production. Carbon assimilation was analysed by gas exchange and single cell analysis of source leaves. Export was determined by microanalysis of phloem exudates and internodal growth rates were measured by displacement transducers. Gas exchange measurements showed similar assimilation rates in the wild‐type and transgenic plants during the light period. Sugar analysis of phloem exudates and epidermal cells revealed a severe shift of sucrose concentrations in the individual plant lines. Moreover, epidermal cells turned out to be a potential storage site for carbohydrates in potato. Finally, we could demonstrate that changing the diurnal rhythm of carbon allocation results in a change in the diurnal growth pattern.
Transient expression of green fluorescent protein in various plastid types following microprojectile bombardment
Authors: Hibberd Julian M. , Linley Philip J. , Khan Muhammad S. , Gray John C. 
Published: March 2003

Summary The green fluorescent protein gene (

gfp

) is a widely used reporter in both animals and plants. Fusions between the plastid

rrn

promoter or the

Escherichia coli trc

promoter and the

gfp

coding region have been delivered to chloroplasts using gold or tungsten microprojectiles, and fluorescence from GFP was visible in individual tobacco chloroplasts and in the abnormally large chloroplasts of the

arc

6 mutant of

Arabidopsis thaliana

2–4 days after bombardment. The fusion of the

gfp

coding region to the bacterial

trc

promoter demonstrated that a bacterial promoter is active in chloroplasts

in vivo

. GFP was also detectable in amyloplasts of potato tubers and in chromoplasts of marigold petals, carrot roots and pepper fruits 4 days after bombardment. This demonstrates that GFP can be used as a reporter for transient gene expression in chloroplasts and in non‐photosynthetic plastids in a range of higher plants.
Floral dip: a simplified method forAgrobacterium‐mediated transformation ofArabidopsis thaliana
Authors: Clough Steven J. , Bent Andrew F. 
Published: March 2003

Summary The

Agrobacterium

vacuum infiltration method has made it possible to transform

Arabidopsis thaliana

without plant tissue culture or regeneration. In the present study, this method was evaluated and a substantially modified transformation method was developed. The labor‐intensive vacuum infiltration process was eliminated in favor of simple dipping of developing floral tissues into a solution containing

Agrobacterium tumefaciens

, 5% sucrose and 500 microliters per litre of surfactant Silwet L‐77. Sucrose and surfactant were critical to the success of the floral dip method. Plants inoculated when numerous immature floral buds and few siliques were present produced transformed progeny at the highest rate. Plant tissue culture media, the hormone benzylamino purine and pH adjustment were unnecessary, and

Agrobacterium

could be applied to plants at a range of cell densities. Repeated application of

Agrobacterium

improved transformation rates and overall yield of transformants approximately twofold. Covering plants for 1 day to retain humidity after inoculation also raised transformation rates twofold. Multiple ecotypes were transformable by this method. The modified method should facilitate high‐throughput transformation of

Arabidopsis

for efforts such as T‐DNA gene tagging, positional cloning, or attempts at targeted gene replacement.
Target genes and regulatory domains of the GAMYB transcriptional activator in cereal aleurone
Authors: Gubler Frank , Raventos Dora , Keys Margaret , Watts Robyn , Mundy John , Jacobsen John V. 
Published: March 2003

SummaryGAMYB is an MYB transcription factor which is expressed in cereal aleurone cells in response to gibberellin (GA). HvGAMYB binds to the TAACAAA box of a high‐pI α‐amylase gene promoter and transcriptionally activates its expression. In this study, we examined the role of HvGAMYB in activating expression of other GA‐regulated genes encoding hydrolytic enzymes. In transient expression experiments, HvGAMYB transactivated expression of reporter genes fused to a low‐pI α‐amylase gene promoter, an EII(1–3,1–4)‐β‐glucanase gene promoter and a cathepsin B‐like protease promoter. HvGAMYB DNA binding specificity was determined using a PCR‐based random site selection using HvGAMYB fusion protein isolated fromE. coli. The deduced consensus closely resembled gibberellin response elements in α‐amylase promoters. Functional analysis of HvGAMYB by transient expression of C terminal HvGAMYB deletions in barley aleurone cells identified two transcriptional activation domains (TADs) which function in transcriptional regulation of both high‐ and low‐pI α‐amylase promoters. The same TADs were identified using a heterologous yeast expression system. Together, these results indicate that HvGAMYB has two TADs. These domains are C‐terminal to its DNA‐binding domain.