bluuebunny/crossref_metadata_2025_split huggingface dataset:
Direct repeats of T‐DNA integrated in tobacco chromosome: characterization of junction regions
Authors: Krizkova Lucie , Hrouda Milan 
Published: March 2003

Summary Plant transformation via

Agrobacterium

frequently results in formation of multiple copy T‐DNA arrays at one target site of the chromosome. The T‐DNA copies are arranged in repeats, direct or inverted around one of the T‐DNA borders. A Ti plasmid‐derived transformation vector has been constructed enabling direct selection of transformants carrying at least two linked copies of T‐DNA in the same orientation. The selection is based on expression of a promoterless neomycin phosphotransferase gene on one T‐DNA copy from a promoter located on the other T‐DNA copy. After co‐cultivation of tobacco protoplasts with

Agrobacterium

, as many as 30% of regenerated transformed plants carried directly repeated T‐DNA copies. The junction regions between two T‐DNAs were amplified and 13 amplified fragments were cloned and sequenced. The involvement of T‐DNA left and right border sequences in direct repeat junctions was determined. In some junctions, additional filler DNA was detected. The length of filler DNA varied from a few up to almost 300 bp. The longer filler DNAs from two clones were found to be T‐DNA fragments in direct or reverse orientation. We discuss the recently suggested models for T‐DNA integration and propose that the formation of direct repeats in genomes does not necessarily result from ligation of intermediates (i.e. T‐strands), but more likely from the co‐integration of several intermediates into one target site.
Treatment ofVicia fabaroot tip cells with specific inhibitors to cyclin‐dependent kinases leads to abnormal spindle formation
Authors: Binarová Pavla , Doležel Jaroslav , Draber Pavel , Heberle‐Bors Erwin , Strnad Miroslav , Bögre László 
Published: March 2003

Summary Many events during cell division are triggered by an evolutionary conserved regulator, the cyclin‐dependent kinase (Cdk). Here we used two novel drugs, the purine analogues bohemine and roscovitine, to study the role of Cdks in cell cycle progression and microtubule organisation in

Vicia faba

root tip cells. Both drugs inhibited the activity of immunopurified

Vicia faba

and alfalfa Cdc2‐kinase. The transcript levels of an A‐ and B‐type cyclin, as well as of the

cdc2

genes, declined in treated root tips, while the mRNA level of a D‐type cyclin gene was not affected. An observed transient arrest at the G1/S and G2/M regulatory points indicated that inhibition of the Cdc2‐kinase had an effect on both transitions. In contrast to the regular bipolar spindle in untreated cell, in drug‐treated metaphase cells abnormally short and dense kinetochore microtubule fibres were observed. These microtubules were randomly arranged in the vicinity of the kinetochores and connected the chromosomes. Thus, the chromosomes were not aligned on the metaphase plate but were arranged in a circle, with kinetochores pointing inwards and chromosome arms pointing outwards. γ‐Tubulin, which plays a role in microtubule nucleation, also localised to the centre of the monopolar spindle. The observed abnormalities in mitosis, after inhibition of Cdc2‐kinase by specific drugs, suggest a role for this enzyme in regulating some of the steps leading to a bipolar spindle structure.
KCS1encodes a fatty acid elongase 3‐ketoacyl‐CoA synthase affecting wax biosynthesis inArabidopsis thaliana
Authors: Todd James , Post‐Beittenmiller Dusty , Jaworski Jan G. 
Published: March 2003

Summary An Arabidopsis fatty acid elongase gene,

KCS1

, with a high degree of sequence identity to

FAE1

, encodes a 3‐ketoacyl‐CoA synthase which is involved in very long chain fatty acid synthesis in vegetative tissues, and which also plays a role in wax biosynthesis. Sequence analysis of

KCS1

predicted that this synthase was anchored to a membrane by two adjacent N‐terminal, membrane‐spanning domains. Analysis of a T‐DNA tagged

kcs1–1

mutant demonstrated the involvement of the

KCS1

in wax biosynthesis. Phenotypic changes in the

kcs1–1

mutant included thinner stems and less resistance to low humidity stress at a young age. Complete loss of

KCS1

expression resulted in decreases of up to 80% in the levels of C26 to C30 wax alcohols and aldehydes, but much smaller effects were observed on the major wax components, i.e. the C29 alkanes and C29 ketones on leaves, stems and siliques. In no case did the loss of

KCS1

expression result in complete loss of any individual wax component or significantly decrease the total wax load. This indicated that there was redundancy in the elongase KCS activities involved in wax synthesis. Furthermore, since alcohol, aldehyde, alkane and ketone levels were affected to varying degrees, involvement of the

KCS1

synthase in both the decarbonylation and acyl‐reduction wax synthesis pathways was demonstrated.
A partially assembled complex I in NAD4‐deficient mitochondria of maize
Authors: Karpova Olga V. , Newton Kathleen J. 
Published: March 2003

Summary The proton‐translocating NADH:ubiquinone oxidoreductase (respiratory complex I) consists of at least 32 subunits in higher plants, nine of which are mitochondrially encoded (NAD 1–7, NAD4L, NAD9). Complex I (CI) has been analyzed from a mitochondrial mutant of maize, NCS2, that carries a deletion for the 3′ end of the
nad4
gene. Mitochondria from highly defective, near‐homoplasmic mutant plants have only trace amounts of the normal complex I. Instead, a reduced amount of a smaller complex, which also exhibits NADH dehydrogenase activity, is detected on ‘blue‐native’ polyacrylamide gels. Subunits of 76 kDa, 40 kDa and 55 kDa, as well as NAD7 and NAD9, have been identified in the subcomplex by their cross‐reactivity with heterologous antisera. The corresponding subunits in
Neurospora
are localized in a ‘peripheral arm’ of CI, which is known to assemble independently of a ‘membrane arm’. The maize NCS2 CI subcomplex is loosely bound to the membrane and is missing several subunits that could be membrane components. Thus, the mutant CI subcomplex may consist of a peripheral arm. A reduction in the steady‐state levels of NAD7 and NAD9 in NCS2 mitochondria occurs despite normal rates of biosynthesis and there is a concomitant decrease of the nuclear encoded 76 kDa subunit. The reduction in CI‐associated NADH dehydrogenase activity in the
nad4
‐deficient NCS2 mutant mitochondria is not associated with a compensatory increase in the activities or amounts of the putative ‘exogenous’ NAD(P)H dehydrogenases that are found in plant mitochondria.
Cell wall alterations and localized accumulation of feruloyl‐3′‐methoxytyramine in onion epidermis at sites of attempted penetration by Botrytis allii are associated with actin polarisation, peroxidase activity and suppression of flavonoid biosynthesis
Authors: McLusky Sarah R. , Bennett Mark H. , Beale Michael H. , Lewis Mervyn J. , Gaskin Paul , Mansfield John W. 
Published: March 2003

Summary Granular deposits of reaction material (RM) were formed in onion epidermal cells at sites of attempted penetration by
Botrytis allii
. Both RM and the adjacent cell wall fluoresced blue under UV excitation. The blue autofluorescence was caused primarily by the accumulation of feruloyl‐3′‐methoxytyramine (FMT) and feruloyltyramine (FT) within challenged tissues. Additional phenolics increasing at infection sites were identified as coumaroyl glucose, coumaroyltyramine (CT) and 2‐hydroxy‐2‐(4‐hydroxyphenyl) ethylferulate. The major autofluorescent components of RM, the hydroxycinnamic acid amides FMT and FT, were bound by ether linkage onto the cell wall as well as being present in methanol soluble granules. Formation of RM was associated with early increases in peroxidase activity detected by histochemistry at reaction sites and striking polarisation of actin microfilaments. Quantitative analysis of quercetin and cyanidin glucosides revealed that the localized synthesis and deposition of feruloyltyramine derivatives was associated with suppression of flavonoid and anthocyanin accumulation in a zone of cells around those accumulating RM. No antifungal activity was detected in FMT, FT or CT, nevertheless it is proposed that the phenolics have a key role in resistance by preventing fungal degradation of the cell wall.
Cell wall antibodies without immunization: generation and use of de‐esterified homogalacturonan block‐specific antibodies from a naive phage display library
Authors: Willats William G. T. , Gilmartin Philip M. , Mikkelsen Jørn Dalgaard , Knox J. Paul 
Published: March 2003

SummaryHomogalacturonan (HG) is a multi‐functional pectic polysaccharide of primary cell walls involved in calcium cross‐linking and gel formation, and the regulation of ionic status and porosity of the cell wall matrix, and is a source of oligosaccharins functioning in development and defence. Phage display monoclonal antibodies with specificity for de‐esterified stretches (‘blocks’) of pectic HG have been isolated from a naive phage display library without the need for immunization of animals or conjugation of an oligosaccharide to protein. These antibodies, designated PAM1 and PAM2, bind specifically to de‐esterified and un‐substituted HG. Assays with a series of pectins de‐esterified by the action of plant or fungal pectin methyl esterases indicated that the antibodies were specific to de‐esterified blocks resulting from the blockwise action of plant pectin methyl esterases. Analysis of antibody binding to a series of oligogalacturonides indicated that optimal binding required in the region of 30 de‐esterified GalA residues. The recognition of such a large epitope by these antibodies allows the HG block architecture of primary cell walls to be identified and localized for the first time. Furthermore, we have demonstrated that monoclonal antibodies with high specificity and avidity to cell wall epitopes can be generated using a ‘single pot’ phage display approach.
The GRAS gene family in Arabidopsis: sequence characterization and basic expression analysis of the SCARECROW‐LIKE genes
Authors: Pysh Leonard D. , Wysocka‐Diller Joanna W. , Camilleri Christine , Bouchez David , Benfey Philip N. 
Published: March 2003

SummaryMutations at the SCARECROW (SCR) locus in Arabidopsis thaliana result in defective radial patterning in the root and shoot. The SCR gene product contains sequences which suggest that it is a transcription factor. A number of Arabidopsis Expressed Sequence Tags (ESTs) have been identified that encode gene products bearing remarkable similarity to SCR throughout their carboxyl‐termini, indicating that SCR is the prototype of a novel gene family. These ESTs have been designated SCARECROW‐LIKE (SCL). The gene products of the GIBBERELLIN‐INSENSITIVE (GAI) and the REPRESSOR of ga1–3 (RGA) loci show high structural and sequence similarity to SCR and the SCLs. Sequence analysis of the products of the GRAS (GAI, RGA, SCR) gene family indicates that they share a variable amino‐terminus and a highly conserved carboxyl‐terminus that contains five recognizable motifs. The SCLs have distinct patterns of expression, but all of those analyzed show expression in the root. One of them, SCL3, has a tissue‐specific pattern of expression in the root similar to SCR. The importance of the GRAS gene family in plant biology has been established by the functional analyses of SCR, GAI and RGA.
Two MAR DNA‐binding proteins of the pea nuclear matrix identify a new class of DNA‐binding proteins
Authors: Hatton Diane , Gray John C. 
Published: March 2003

SummaryFour MAR‐binding proteins of 60, 65, 70 and 72 kDa have been detected by South‐Western blotting and isolated from pea nuclear matrices. Two cDNAs encoding the 60 and 65 kDa proteins (MARBP‐1 and MARBP‐2) were isolated from a pea leaf cDNA library by screening with a PCR product obtained using degenerate primers based on an amino acid sequence from the 60 kDa protein. The proteins of 560 and 550 amino acids are 86% identical and contain several KKD/E repeats near the C‐terminus. Escherichia coli‐expressed MARBP‐1 specifically binds A/T‐rich MAR DNA. The interaction of MARBP‐1/MARBP‐2 with MAR DNA involves novel DNA‐binding motifs. The MARBP‐1 and MARBP‐2 genes are expressed in a range of pea tissues and are encoded by genes at different loci. MARBP‐1 and MARBP‐2 are homologous to yeast nucleolar proteins Nop56p and Nop58p, which are involved in ribosome biogenesis, and to similar highly conserved proteins in other eukaryotes and in archaebacteria. MARBP‐1 and MARBP‐2 may have multifunctional roles in chromatin organisation and ribosome biogenesis.
Oxygen deprivation stimulates Ca2+‐mediated phosphorylation of mRNA cap‐binding protein eIF4E in maize roots
Authors: Manjunath Sivalinganna , Williams Alan J. , Bailey‐Serres Julia 
Published: March 2003

SummaryFlooding of maize seedlings causes O2 deprivation that leads to a global reduction in protein synthesis and selective translation of cytoplasmic mRNAs. Since selective translation in animal cells can involve the cap‐binding protein eIF4E, we characterized the distinct mRNA cap‐binding proteins eIF4E and eIFiso4E of maize. These proteins have 45% deduced amino acid sequence identity and are highly conserved at residues of eIF4E that function in intermolecular interactions in animals. Maize eIF4E is a phosphoprotein. O2 deprivation resulted in a decrease in the isoelectric point of eIF4E, consistent with additional phosphorylation. Modification of eIF4E was mimicked by treatment with caffeine under aerobic conditions and blocked by treatment with ruthenium red under O2 deprivation, implicating Ca2+ as a second messenger in eIF4E modification. In contrast, no isoelectric variants of eIFiso4E were detected. The possible role of cytosolic Ca2+ and pH in regulation of mRNA cap‐binding protein activity under O2 deprivation is discussed.
Emission of nitrous oxide (N2O) from transgenic tobacco expressing antisense NiR mRNA
Authors: Goshima Naoki , Mukai Toshihiro , Suemori Mamiko , Takahashi Misa , Caboche Michel , Morikawa Hiromichi 
Published: March 2003

SummaryThe emission of N2 and N2O from intact transgenic tobacco (clone 271) expressing antisense nitrite reductase (NiR) mRNA, and wild‐type plants grown aseptically, on NO3–, NO2– or NH4+ ‐containing medium was investigated. 15N contents of gas sampled from gas‐sealed pots, in which the plants were grown on 15N‐containing medium, were analyzed by gas chromato‐ graphy and mass spectrometry (GC–MS). No emission of N2 was detected in either of the gas samples from plant clone 271 or the wild‐type grown on NO3–‐containing medium. N2O emission from clone 271 grown on NO3–‐containing medium was detected, but not from the wild‐type plants. The N2O emission rate of clone 271 was 106 ng N2O mg–1 incorporated N week–1 and the N2O emission was inhibited by tungstate (a nitrate reductase inhibitor). No emission of N2O was found from clone 271 or wild‐type plants grown on medium containing NH4+. Emission of N2O also was detected from clone 271 grown on NO2–‐containing medium and its emission rate increased with increasing NO2– levels in plants. We speculate that NO3– is reduced to NO2– and that a part of NO2– is metabolized to N2O in clone 271.
The distinctive roles of five different ARC genes in the chloroplast division process in Arabidopsis
Authors: Marrison Joanne L. , Rutherford Stephen M. , Robertson Elizabeth J. , Lister Clare , Dean Caroline , Leech Rachel M. 
Published: March 2003

Summary ARC (accumulation and replication of chloroplasts) genes control different aspects of the chloroplast division process in higher plants. In order to establish the hierarchy of the ARC genes in the chloroplast division process and to provide evidence for their specific roles, double mutants were constructed between arc11, arc6, arc5, arc3 and arc1 in all combinations and phenotypically analysed. arc11 is a new nuclear recessive mutant with 29 chloroplasts compared with 120 in wild type. All the phenotypes of the double mutants are unambiguous. ARC1 down‐regulates proplastid division but is on a separate pathway from ARC3, ARC5, ARC6 and ARC11. ARC6 initiates both proplastid and chloroplast division. ARC3 controls the rate of chloroplast expansion and ARC11 the central positioning of the final division plane in chloroplast division. ARC5 facilitates separation of the two daughter chloroplasts. ARC5 maps to chromosome 3 and ARC11 and ARC6 map approximately 60 cM apart on chromosome 5.
AtAMT1 gene expression and NH4+ uptake in roots of Arabidopsis thaliana: evidence for regulation by root glutamine levels
Authors: Rawat Suman R. , Silim Salim N. , Kronzucker Herbert J. , Siddiqi M. Yaeesh , Glass Anthony D. M. 
Published: March 2003

SummaryThe mechanisms involved in regulating high‐affinity ammonium (NH4+) uptake and the expression of the AtAMT1 gene encoding a putative high‐affinity NH4+ transporter were investigated in the roots of Arabidopsis thaliana. Under conditions of steady‐state nitrogen (N) supply, transcript levels of the AtAMT1 gene and Vmax values for high‐affinity 13NH4+ influx were inversely correlated with levels of N provision. Following re‐supply of NH4NO3 to N‐starved plants, AtAMT1 mRNA levels and 13NH4+ influx declined rapidly but remained high when the conversion of NH4+ to glutamine (Gln) was blocked with methionine sulfoximine (MSX). This result demonstrates that end products of NH4+ assimilation, rather than NH4+ itself, are responsible for regulating AtAMT1 gene expression. Consistent with this hypothesis, AtAMT1 gene expression and NH4+ influx were suppressed by provision of Gln alone, or together with NH4NO3 plus MSX. Furthermore, AtAMT1 transcript levels and 13NH4+ influx were negatively correlated with root Gln concentrations, following re‐supply of N to N‐starved plants. In addition to this level of control, the data suggest that high cytoplasmic [NH4+] may inhibit NH4+ influx.
Uncoupling secretion and tip growth in lily pollen tubes: evidence for the role of calcium in exocytosis
Authors: Roy Stéphane J. , Holdaway‐Clarke Terena L. , Hackett Grant R. , Kunkel Joseph G. , Lord Elizabeth M. , Hepler Peter K. 
Published: March 2003

SummaryCytoplasmic calcium concentration ([Ca2+]i) and extracellular calcium (Ca2+o) influx has been studied in pollen tubes of Lilium longliflorum in which the processes of cell elongation and exocytosis have been uncoupled by use of Yariv phenylglycoside ((β‐D‐Glc)3). Growing pollen tubes were pressure injected with the ratio dye fura‐2 dextran and imaged after application of (β‐D‐Glc)3, which binds arabinogalactan proteins (AGPs). Application of (β‐D‐Glc)3 inhibited growth but not secretion. Ratiometric imaging of [Ca2+]i revealed an initial spread in the locus of the apical [Ca2+]i gradient and substantial elevations in basal [Ca2+]i followed by the establishment of new regions of elevated [Ca2+]i on the flanks of the tip region. Areas of elevated [Ca2+]i corresponded to sites of pronounced exocytosis, as evidenced by the formation of wall ingrowths adjacent to the plasma membrane. Ca2+o influx at the tip of (β‐D‐Glc)3‐treated pollen tubes was not significantly different to that of control tubes. Taken together these data indicate that regions of elevated [Ca2+]i, probably resulting from Ca2+o influx across the plasma membrane, stimulate exocytosis in pollen tubes independent of cell elongation.
Rhizobium Nod factors induce increases in intracellular free calcium and extracellular calcium influxes in bean root hairs
Authors: Cárdenas Luis , Feijó José A. , Kunkel Joseph G. , Sánchez Federico , Holdaway‐Clarke Terena , Hepler Peter K. , Quinto Carmen 
Published: March 2003

SummaryApplication of Nod factors to growing, responsive root hairs of the bean Phaseolus vulgaris induces marked changes in both the intracellular cytosolic free calcium (Ca2+) and in the influx of extracellular [Ca2+]. The intracellular [Ca2+], which has been measured by ratiometric imaging in cells microinjected with fura‐2‐dextran (70 kDa), elevates within 5 min from approximately 400 n
m to 1500 n
m in localised zones in the root hair apex. Of particular note is the observation that the elevated regions of [Ca2+] appear to shift position during short time intervals. Increases in and fluctuations of the intracellular [Ca2+] are also observed in the perinuclear region after 10–15 min treatment with Nod factors. The extracellular Ca2+ flux, detected with the non‐invasive, calcium specific vibrating electrode, is inwardly directed and also increases quickly in response to Nod factors from 13 pmol cm–2 s–1 to 28 pmol cm–2 s–1. Chitin‐oligomers, which are structurally similar but biologically inactive when compared to the active Nod factors, fail to elicit changes in either intracellular or extracellular Ca2+. The similar timing and location of the intracellular elevations and the increased extracellular influx provide support for the idea that Ca2+ participates in secretion and cell wall remodelling, which occur in anticipation of root hair deformation and curling.
CYP78A5 encodes a cytochrome P450 that marks the shoot apical meristem boundary in Arabidopsis
Authors: Zondlo Susan C. , Irish Vivian F. 
Published: March 2003

SummaryThe normal development of shoot structures depends on controlling the growth, proliferation and differentiation of cells derived from the shoot apical meristem. We have identified the CYP78A5 gene encoding a putative cytochrome P450 monooxygenase that is the first member of the CYP78 family from Arabidopsis. This gene is strongly expressed in the peripheral regions of the vegetative and reproductive shoot apical meristems, defining a boundary between the central meristematic zone and the developing organ primordia. In addition, CYP78A5 shows a dynamic pattern of expression during floral development. Overexpression of CYP78A5 affects multiple cell types, causing twisting and kinking of the stem and defects in floral development. To define the relationship of CYP78A5 to genes controlling meristem function, we examined CYP78A5 expression in plants mutant for SHOOT MERISTEMLESS, ZWILLE and ARGONAUTE, and have found that CYP78A5 expression is altered in these mutant backgrounds. We propose that CYP78A5 has a role in regulating directional growth in the peripheral region of the shoot apical meristem in response to cues established by genes regulating meristem function.
The AtSUC1 sucrose carrier may represent the osmotic driving force for anther dehiscence and pollen tube growth in Arabidopsis
Authors: Stadler Ruth , Truernit Elisabeth , Gahrtz Manfred , Sauer Norbert 
Published: March 2003

SummaryThe Arabidopsis AtSUC1 protein has previously been characterized as a plasma membrane H+‐sucrose symporter. This paper describes the sites of AtSUC1 gene expression and AtSUC1 protein localization and assigns specific functions to this sucrose transporter in anther development and pollen tube growth. RNase protection assays revealed AtSUC1 expression exclusively in floral tissue, which was confirmed by analyses of AtSUC1 promoter‐β‐glucuronidase (GUS) plants. In situ hybridizations identified AtSUC1 expression in anther connective tissue, in funiculi and in fully developed pollen grains. Indirect immuno‐fluorescence analyses with anti‐AtSUC1 antiserum confirmed AtSUC1 protein localization in the connective tissue and funiculi. In mature pollen grains, however, despite high AtSUC1 mRNA levels no AtSUC1 protein was found. Only after pollination of stylar papillae was AtSUC1 protein detected inside the pollen and later inside the growing pollen tubes, suggesting a translation of pre‐existing AtSUC1 mRNA after pollination. Pollen germination analyses underlined the important role of sucrose for pollen tube growth. The data presented suggest a role of AtSUC1 in the controlled dehiscence of Arabidopsis anthers. It is postulated that an important function of AtSUC1 is the cell‐specific modulation of water potentials.