bluuebunny/crossref_metadata_2025 huggingface dataset:
Serial detection of Epstein–Barr virus DNA in sera and peripheral blood leukocyte samples of pediatric renal allograft recipients with persistent mononucleosis‐like symptoms defines patients at risk to develop post‐transplant lymphoproliferative disease
Authors: Campe Hartmut , Jaeger Gundula , Abou‐Ajram Claudia , Nitschko Hans , Griebel Martin , Montoya Carmen , Klare Bernd , Koszinowski Ulrich
Published: March 2003

Abstract:  We tested blood samples of 25 pediatric renal transplant recipients for Epstein–Barr virus (EBV) DNA load by quantitative polymerase chain reaction (PCR). Eleven of these transplant recipients showed clinical persistent mononucleosis‐like symptoms years after transplantation (Tx). A quantitation of EBV DNA by PCR in peripheral blood lymphocyte (PBL) and serum samples revealed variable EBV DNA titers. The majority of EBV PCR results in samples of the 14 asymptomatic transplant recipients was repeatedly below detection limit. In contrast, patients with mononucleosis‐like symptoms showed persistent EBV genome titers over a period of 6 months, ranging from 75 to 18 750 copies/10 000 PBL and from 680 to 335 000 copies/mL serum, respectively. One child suffering from this mononucleosis‐like condition developed an EBV‐associated Burkitt‐like lymphoma 29 months after Tx. Whereas clinical and histological investigations did not indicate a post‐transplant lymphoproliferative disorder (PTLD) until tumor detection, EBV titers in PBL and serum had been high for at least 8 months. We propose that pediatric transplant recipients who show both, recurrent mononucleosis‐like symptoms and a sustained high EBV genome load, are at increased risk for severe EBV‐related post‐transplant complications.
Sunflower seedlings subjected to increasing water deficit stress: oxidative stress and defence mechanisms
Authors: Sgherri Cristina Luisa Maria , Navari‐Izzo Flavia
Published: March 2003

Sunflower seedlings (Helianthus annuus cv. Licia Stella) reached a mild, moderate and severe level of water deficit stress after 5, 8 and 11 days of soil water depletion, respectively. At a moderate level of water deficit stress, an elastic adjustment occurred. At the same time, in response to a minor osmotic potential (Ψπ) and an intermediate rate of water potential (Ψw) decrease, glutathione level increased and enzyme activities related to the ascorbate/glutathione cycle were induced. At severe water deficit stress, the efficiency of this defence mechanism fell, oxidative processes intensified and soluble protein content decreased further. The results of the present experiments suggest that during water deficit stress the water status of the plants plays a key role in the activation of defence mechanisms.
Auxin increases elastic wall‐properties in rye coleoptiles: implications for the mechanism of wall loosening
Authors: Edelmann Hans G. , Köhler Kurt
Published: March 2003

Auxin‐induced changes of wall‐rheological properties during different growth rates of rye coleoptile segments (Secale cereale L.) were investigated. In addition, changes of osmotic concentration and turgor pressure were measured. Decrease of turgor and of osmotic concentration followed a synchronous time course. Auxin‐incubated segments exhibited a faster decrease and eventually lower values of both parameters. Creep test extensibility measurements demonstrate that apparent plastic as well as elastic extensibility of distilled‐water‐incubated segments strongly decreased during 24 h. In auxin‐incubated segments apparent plastic as well as elastic extensibilities were strongly increased, even in the absence of growth due to insufficient turgor pressure. The increasing effect of auxin on elastic wall properties is also reflected by an increase in relative reversible length (part of segment length by which segments shrink after freezing/thawing as referred to total length) and a complementary decrease of relative irreversible length (remaining length after turgor elimination as referred to turgid length); again the effects were independent of growth rate and turgor pressure. Cellulose synthesis inhibition of approx. 80% by dichlorobenzonitrile (DCB) had no significant effect either on growth or on wall‐rheological properties. Independent of whether the changed rheological wall behaviour of auxin‐incubated segments is causally related to the mechanism of auxin‐induced wall loosening, it indicates changes of wall polymer properties and/or interactions which are conserved when no actual length increase occurs due to insufficient turgor pressure. The results suggest that IAA‐induced wall loosening may be primarily mediated by cell wall changes other than cleavage of covalent, load‐bearing bonds as hypothesized in various wall loosening models.
Identification of a phenylalanine ammonia‐Iyase inactivating factor in harvested head lettuce (Lactuca sativa)
Authors: Ritenour Mark A. , Saltveit Mikal E.
Published: March 2003

Exposing head lettuce (Lactuca sativa L., crisphead or Iceberg type) leaf tissue to hormonal levels of ethylene (10 μl l−1) at 5°C promotes the de novo synthesis of phenylalanine ammonia‐lyase (PAL, EC 4.3.1.5) and an increase in its activity. It also promotes the appearance of the postharvest physiological disorder called russet spotting (RS). Discontinuing ethylene exposure after 4 days resulted in a rapid decline in PAL activity which was delayed by treating excised midrib leaf tissue with actinornycin D or cycloheximide at 5°C. Only cycloheximide delayed the loss of PAL activity in tissue that was transferred from 5 to 15°C. Activity of PAL from Rhodolorula glutinis was. slowly lost during incubation in buffer alone, but there was a logarithmic decline in its activity over time when it was incubated with aliquots of the resuspended 10000 g pellet from homogenized, lettuce tissue affected with RS. The in vitro loss in PAL activity was 9–fold higher in extracts from lettuce showing RS symptoms than from control lettuce, boiled samples or the buffer control. The PAL‐inactivating factor isolated from lettuce affected with RS had a pH optimum around 8.0. It appears that the rapid loss in PAL activity after the discontinuation of exposure to ethylene is dependent on the de novo synthesis of a PAL‐inactivating factor.
Induction of cell division‐related genes in quiescent (G0) sugar beet cells
Authors: Fowler Mark R. , Kirby Melissa J. , Scott Nigel W. , Slater Adrian , Elliott Malcolm C.
Published: March 2003

Sugar beet cells maintained in the stationary phase of the batch culture cycle for 2 or more days have been shown to exhibit many of the characteristics of quiescent (G0) cells. When such cells were subcultured into fresh medium they progressed through a period of DNA synthesis to a highly synchronised first division, 6 days after subculture. The onset of DNA synthesis and cell division were each delayed by 2 days relative to the timing of the events when the cells were subcultured immediately before entry into the stationary phase. The regulation of gene expression during this extended transition from the G0 phase back to the cell division cycle was investigated.

The cell division cycle‐related genes Bvcdc2, Betvu;CycA2, Arath;CycB1;1 histone H4 and Bvcrk1 (a novel cdc2‐like gene) showed widely differing patterns of expression. Bvcdc2 transcripts were present at low levels in quiescent cells whereas crk1, cyclin and histone transcripts were not detectable. Expression of both Bvcrk1 and Betvu; CycA2 was induced within 1 h after subculture into fresh medium, whereas histone H4 gene expression was not detectable for 24 h and showed a marked increase between 24 and 48 h. B‐type cyclin transcripts were not detectable until more than 48 h after subculture.

The addition of either sucrose or MS macronutrients to quiescent sugar beet cells was not sufficient to re‐initiate cell division but both medium components were able to stimulate the expression of the two ‘early’ genes (Betvu;CycA2 and Bvcrk1) within 6 h. Furthermore, although the sugar beet cells were habituated, i.e. they were routinely grown without added plant growth regulators, treatment of quiescent cells with IAA and kinetin also induced expression of Betvu;CycA2 and Bvcrk1 without subsequent cell division.
Phenotypic alterations and component analysis of seed yield in transgenic Brassica napus plants expressing the tzs gene
Authors: Roeckel P. , Oancia T. , Drevet J. R.
Published: March 2003

Cytokinins play an important role in plant development. We investigated the possibility that the nopaline Ti plasmid gene (tzs) from Agrobacterium tumefaciens could encode a protein able to participate in plant cytokinin production and lead to alterations in plant phenotype as a result of the expression of endogenous tzs. tzs was placed under the control of a heat‐inducible promoter from the Zea mays hsp70 gene. The expression of this fused gene was examined in transgenic Brassica napus plants. The tzs gene, which encodes the enzyme dimethylallyl transferase, was used as a cytokinin biosynthetic gene. The expression of the tzs gene was monitored by RNA hybridization and analysis of cytokinin content. Overproduction of cytokinin was observed even when the plants had not been heat‐shocked, and the plants displayed a reduced root system, increased height and branching, and delayed flowering. In addition, a significant increase in seed yield was observed in the transgenic plants, accounted for by increased number of seeds per silique and seed weight. The results suggest that increased levels of cytokinins, through the expression of tzs, are correlated with growth rather than with differentiation processes.
Inhibition of glucose transport by phenylglyoxal: Both dissipation of ΔpH and essential arginyl residues are involved
Authors: Guy Micha , Volokita Micha
Published: March 2003

The effects of the arginine modifying reagent phenylglyoxal (PGO) on solute transport was studied in two cellular systems: protoplasts isolated from the mesophyll of Vicia faba L. and XD cell suspension culture of Nicotiana tabacum L. cv. Xanthi. The solutes in the case of the protoplasts were the non‐metabolizable glucose analog 3‐O‐methyl‐D‐glucose (MeG), and a non‐metabolizable amino acid analog α‐aminoisobutyric acid (AIB), whereas the solutes for the cell suspension were AIB and nitrate. Solute transport in both systems was rapidly inhibited by PGO. Exposure of the protoplasts to light enhanced the initial rate of MeG uptake. PGO rapidly inhibited MeG uptake in both the light and the dark, the half‐time for inactivation being less than 3 min. Flux analysis of double‐labeled MeG showed that initial MeG uptake was mediated mainly by the plasma membrane transport system and that it was inhibited by PGO. Maximal inhibition of initial MeG uptake rate was observed at PGO concentrations of 1 mM and above. PGO treatment altered rapidly the equilibrium distribution of the ΔpH probe dimethyloxazolidine (DMO) in both cellular systems, indicating dissipation of ΔpH between cell and medium. In the protoplasts, PGO inhibited both DMO and MeG uptake at pH 5.5; however, at pH 7.0, where ΔpH is minimal, only MeG uptake was inhibited. Our results suggest that PGO has two effects on glucose uptake: an indirect effect through ΔpH dissipation and a direct effect through interaction with essential arginyl residues in the glucose transporter.
Water relations and osmotic adjustment in Apium graveolens during long‐term NaCl stress and subsequent relief
Authors: Pardossi Alberto , Malorgio Fernando , Oriolo Domenico , Gucci Riccardo , Serra Giovanni , Tognoni Franco
Published: March 2003

To investigate how the regulation of plant water status contributes to salt tolerance in celery (Apium graveolens L. cv. Istar) we determined growth, leaf gas exchange and water relations in hydroponically‐grown plants supplied with nutrient solutions containing 5 (control), 50, 100 and 300 mM NaCl for 4 weeks, and supplied subsequently with the control solution to relieve salt stress. At salinity levels of 50 and 100 mM NaCl there was a moderate, albeit significant, reduction in biomass production. A drastic decrease in both dry weight and leaf area occurred at 300 mM NaCl, but the plants remained viable and showed appreciable photosynthesis and growth. Leaf water potential, turgor pressure, stomatal conductance and plant hydraulic conductance (Lp) decreased during salt exposure. After the first week of stress there was a progressive increase in leaf turgor and Lp, which was associated with substantial recovery of stomatal conductance and photosynthesis in 50 and 100 mM NaCl‐treated plants. The analysis of the pressure‐volume isotherms showed that osmotic adjustment had occurred under all NaCl treatments. Leaf osmotic adjustment in salt‐stressed plants was primarily accomplished by accumulation of Na+ and Cl−. Upon relief of stress, salinized plants rehydrated and regained turgor rapidly, and growth and photosynthesis resumed almost fully. These results indicate that maintenance of a favourable water status contributes substantially to the salt resistance of celery.
Postharvest changes in endogenous cytokinins and cytokinin efficacy in potato tubers in relation to bud endodormancy
Authors: Suttle Jeffrey C.
Published: March 2003

Using an indirect enzyme‐linked immunosorbent assay (ELISA), the effects of postharvest storage duration and temperature on endogenous cytokinins in potato (Solanum tuberosum L. cv. Russet Burbank) tuber apical bud tissues in relation to endodormancy status were determined. Following fractionation by HPLC, a total of eight cytokinins were detected and these were: zeatin riboside‐5′‐monophosphate (ZRMP), zeatin‐O‐glucoside (ZOG), zeatin (Z), zeatin riboside (ZR), isopentenyl adenosine‐5′‐monophosphate (IPMP), isopentenyl adenine‐9‐glucoside (IP‐9‐G), isopentenyl adenine (IP) and isopentenyl adenosine (IPA). Regardless of postharvest storage temperature or endodormancy status, IP‐9‐G was the most abundant cytokinin detected while ZRMP and ZOG were the least abundant ones. In tubers preincubated at a growth‐permissive temperature (20°C) prior to extraction, the loss of endodormancy was preceded by significant increases in the endogenous levels of Z, ZR, IPMP and IP‐9‐G. When stored continuously at a growth‐inhibiting temperature (3°C), significant increases in ZR, IP‐9‐G and IP + IPA were observed. The total content of cytokinins increased by over 7‐fold during postharvest storage and this increase was a result of de novo biosynthesis. Dose‐response studies using IPA and ZR demonstrated a time‐dependent increase in apparent cytokinin sensitivity during postharvest storage. With the exception of IP‐9‐G, injection of any of these endogenous cytokinins resulted in the rapid and complete termination of tuber endodormancy. The significance of these results with respect to endodormancy regulation and the possible mechanisms controlling cytokinin levels in potato tubers are discussed.
Elevated pyruvate,orthophosphate dikinase (PPDK) activity alters carbon metabolism in C3 transgenic potatoes with a C4 maize PPDK gene
Authors: Ishimaru Ken , Ohkawa Yasunobu , Ishige Teruo , Tobias Dennis J. , Ohsugi Ryu
Published: March 2003

Changes in carbon metabolism and δ13C value of transgenic potato plants with a maize pyruvate,orthophosphate dikinase (PPDK; EC 2.7.9.1) gene are reported. PPDK catalyzes the formation of phosphoenolpyruvate (PEP), the initial acceptor of CO2 in the C4 photosynthetic pathway. PPDK activities in the leases of transgenic potatoes were up to 5.4‐fold higher than those of control potato plants (wild‐type and treated control plants). In the transgenic potato plants, PPDK activity in leaves was negatively correlated with pyruvate content (r2= 0.81), and was positively correlated with malate content (r2= 0.88). A significant increase in the δ13C value was observed in the transgenic potato plants, suggesting a certain contribution of PEP carboxylase as the initial acceptor of atmospheric CO2. These data suggest that elevated PPDK activity may alter carbon metabolism and lead to a partial operation of C4‐type carbon metabolism. However, since parameters associated with CO2 gas exchange were not affected, the altered carbon metabolism had only a small effect on the total photosynthetic characteristics of the transgenic plants.
The association of actin and tubulin with plasma membranes: Characterization using inside‐out vesicles formed by Brij 58
Authors: Sonesson Anders , Widell Susanne
Published: March 2003

Most processes of eukaryotic cells depend on the cortical cytoskeleton (CS), a protein filament structure associated to the plasma membrane (PM). With animal cells, much information has been collected on the mechanisms behind CS‐PM interactions, but for plant cells the CS‐PM links are poorly characterized. To allow investigations on these links, isolated PM from cauliflower were here treated with Brij 58, a detergent that causes the PM vesicles to turn inside‐out (cytoplasmic side‐out), thereby exposing the CS components. When actin and tubulin co‐pelleted with inside‐out PM were separated using sucrose gradient centrifugation, actin and tubulin were recovered with PM‐marker activities, supporting intact links between these CS proteins and the Brij‐treated PM. Inside‐out PM was also treated with different media to learn more about the CS‐PM interaction. Extensive dialysis against a low ionic strength medium released actin but not tubulin from these PM, while dialysis against 0.7 M NaCl had no effect. Neither 50 mM DTT, 10 mM CaCl2 nor 2 M NaCl had any effect on the release of either actin or tubulin from PM, but actin was completely released with 6 M urea or 0.6 M KI. Tubulin was also released by urea but not by KI. Incubation of PM in sodium carbonate at increasing pH led to a total release of actin at pH 10, of α‐tubulin at pH 11 and of β‐tubulin at pH 11.4. In many respects, these characteristics agree with reported findings using e.g., fluorescence microscopy with protoplast ghosts, suggesting that inside‐out vesicles obtained with Brij 58 can be used in investigations aimed at understanding the role of the cortical CS in regulating PM‐bound components.
Individual members of the light‐harvesting complex II chlorophyll a/b‐binding protein gene family in pea (Pisum sativum) show differential responses to ultraviolet‐B radiation
Authors: A.‐H.‐Mackerness Soheila , Liu Liansen , Thomas Brian , Thompson William F. , Jordan Brian R. , White Michael J.
Published: March 2003

Light‐harvesting complex II chlorophyll a/b‐binding protein (Lhcb) mRNA levels are differentially affected by ultraviolet‐B radiation (UV‐B, 280‐320 nm) at different stages of development of pea (Pisum sativum L. cv. Feltham first) seedlings. Addition of UV‐B radiation to the light periods of diurnal cycles of white light resulted in reduction of total Lhcb mRNAs in green leaves but a transient increase in etiolated buds. The aims of this study were to determine the stage during de‐etiolation at which supplementary UV‐B began to inhibit Lhcb gene expression, and to determine whether differential regulation of individual Lhcb genes could explain the differential response to supplementary UV‐B at different developmental stages. All seven Lhcb mRNAs were shown to increase in etiolated buds transferred to a diurnal cycle with supplementary UV‐B during the light periods, but were greatly reduced in green leaves given the same treatment. Therefore, the different responses of total Lhcb mRNA levels to UV‐B radiation in green leaves and etiolated buds are not primarily due to the expression of different members of the Lhcb gene family at different developmental stages. However, the Lhcb genes could be divided into two groups based on their sensitivities to UV‐B. Transcripts from the three genes, Lhcb1*2, Lhcb1*3 and Lhcb1*5, which were undetectable in dark‐grown etiolated buds, exhibited stronger responses to supplementary UV‐B in green leaves than the four genes, Lhcb1*1, Lhcb1*4, Lhcb2*1 and Lhcb3*1, which showed low levels of initial transcript accumulation in dark‐grown etiolated buds. The effect of UV‐B on Lhcb mRNA levels were, however, correlated with chlorophyll content, suggesting that the developmental stage of chloroplasts may be important in determining the responses of the Lhcb genes to supplementary UV‐B radiation in pea seedlings.
Growth and physiology of aspen supplied with different fertilizer addition rates
Authors: Coleman Mark D. , Dickson Richard E. , Isebrands J. G.
Published: March 2003

Variable internal plant nutrient content may confound plant response to environmental stress. Plant nutrient content may be controlled with relative addition rate techniques in solution culture. However, because raising large numbers of plants in flowing solution culture is difficult, we investigated the feasibility of raising plants in soil mix using relative fertilizer additions. Aspen (Populus tremuloides Michx.) clones (216, 259 and 271) planted in pots containing a peat, sand and vermiculite (2:1:1, v/v/v) soil mix were grown with exponentially increasing fertilizer concentrations and harvested periodically to assess growth. Addition rate treatments ranged from 0.01 to 0.05 day−1. The lag phase of growth, in which plants adjusted to the fertilizer regime, lasted 40 days after which plants entered the experimental period characterized by constant relative growth rates equivalent to applied fertilizer addition rates. Total plant nutrient concentration was (1) unique for each addition rate, (2) linearly related to addition rate and growth rate, and (3) tended to increase at the highest, and decrease at the lowest addition rates. Regardless, the plants appeared to have attained steady‐state conditions. Allocation of carbon to roots increased with lower addition rate treatments and was not dependent upon ontogeny. There were no treatment differences in growth response among aspen clones. Yet there were treatment differences in leaf chlorophyll and photosynthesis within the clones. For the 0.05 day−1 addition rate treatment, chlorophyll, leaf N concentration and photosynthetic rate were strongly correlated with one another, were at a maximum in recently mature leaves, and rapidly declined with leaf age. The rate of decline in these leaf characteristics was slowest in clone 271, consistent with the leaf longevity stress response reported elsewhere. Plant responses from these relative fertilizer addition trials in soil mix agree closely with those run in hydroponics, indicating that steady‐state nutrition can be achieved with a technically simple experimental assemblage.
Possible involvement of phospholipase A2 in light signal transduction of guard cells of Commelina communis
Authors: Suh SuJeong , Park Joung‐gun , Lee Youngsook
Published: March 2003

Polyunsaturated fatty acids induce stomatal opening (Y. Lee, H. Lee, R. C. Crain, A. Lee and S. J. Korn. 1994. Cell Signal. 6: 181–186), but it is not known whether they function as second messengers in guard cells exposed to signals that open stomata. To test the hypothesis that phospholipase A2 (PLA2), which produces fatty acids and lysophospholipids, is involved in light signal transduction in guard cells, we treated epidermal peels of Commelina communis L. with PLA2 inhibitors and followed the changes in stomatal apertures in response to light. Stomatal opening by white, blue, or red light was inhibited by 2–3 different PLA2 inhibitors in concentration ranges that have been reported to inhibit PLA2 activity. However, the PLA2 inhibitors could not block stomatal opening induced by a polyunsaturated fatty acid. These results suggest that PLA2 functions as a signal transducer for both blue and red light in guard cells.
Polyamine conjugate levels and ethylene biosynthesis: inverse relationship with vegetative bud formation in tobacco thin layers
Authors: Scaramagli S. , Biondi S. , Capitani F. , Gerola P. , Altamura M. M. , Torrigiani P.
Published: March 2003

The effects of two inhibitors of polyamine (spermidine and spermine) biosynthesis, cyclohexylamine (CHA; 5 and 10 mM) and methylglyoxal(bis‐guanylhydrazone) (MGBG; 0.1, 0.5 and 1 mM), on the organogenic response in vegetative bud‐forming tobacco (Nicotiana tabacum L. cv. Samsun) thin layer explants were evaluated micro‐ and macroscopically at different times during culture. The final number of buds formed and the percentage of organogenic explants was significantly reduced by both inhibitors, but much more so by MGBG than CHA. This inhibitory effect was already evident in MGBG‐treated explants on day 5, in terms of the number of meristemoids per explant. On the contrary, in the presence of CHA, the number of meristemoids on day 5 was higher than that in the controls. Between days 9 and 13, meristemoid formation slowed down considerably in inhibitor‐treated explants compared with controls. On day 13, the number of bud primordia was similar in control and CHA‐treated explants, but significantly lower in MGBG‐treated explants. This inhibitor also induced peculiar cytohistological events, such as a reduced formation of oval‐shaped cell aggregates on the explant surface and more frequent cases of nucleolar extrusion, while CHA led to the appearance of hypertrophic epidermal cells; callus formation at the basal end of the explant and xylogenesis were also affected by the inhibitors. Ethylene biosynthesis, measured as [

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C]methionine incorporation, was stimulated 2‐ (day 2) to 3‐fold (15 h) by 0.5 mM MGBG, whereas CHA (10 mM) had little effect and aminoethoxyvinylglycine (AVG; 0.1 μM), an ethylene synthesis inhibitor, was strongly inhibitory. In control explants, the incorporation of labelled methionine into ethylene and spermidine followed an inverse trend up to day 8. In these explants, free putrescine increased 32‐fold and spermidine increased about 10‐fold between days 0 and 8. Trichloroacetic acid (TCA)‐soluble conjugated putrescine also accumulated dramatically during culture. While CHA provoked a decline in spermidine levels, MGBG caused an unexpected increase in free spermidine and spermine titres; however, its most conspicuous effect was on the further enhancement of putrescine conjugate accumulation, while CHA and AVG had the opposite effect. Results are discussed in view of establishing a putative link between MGBG‐enhanced ethylene synthesis, increased conjugate titres and inhibition of meristemoid formation.
Molecular determinants of the Arabidopsis AKT1 K+ channel ionic selectivity investigated by expression in yeast of randomly mutated channels
Authors: Ros R. , Lemaillet G. , Fonrouge A.G. , Daram P. , Enjuto M. , Salmon J.M. , Thibaud J.B. , Sentenac H.
Published: March 2003

The Arabidopsis thaliana K+ channel AKT1 was expressed in a yeast strain defective for K+ uptake at low K+ concentrations (<3 mM). Besides restoring K+ transport in this strain, AKT1 expression increased its tolerance to salt (NaCl or LiCl), whatever the external K+ concentration used (50 μM, 5 mM, or 50 mM). We took advantage of the latter phenomenon for screening a library of channels randomly mutated in the region that shares homologies with the pore forming domain (the so‐called P domain) of animal K+ channels (Shaker family). Cassette mutagenesis was performed using a degenerate oligonucleotide that was designed to ensure, theoretically, a single mutation per P cassette. The mean number of amino acid exchanges per cassette turned out to be 1.4. Mutant channels that conferred on the transformed cells a reduction in salt tolerance (increased Na+ content, decreased K+ content, and lower growth rate, as compared to control cells expressing the wild‐type channel) were selected. By co‐expressing them with the wild‐type AKT1 cDNA, it was shown that the mutated polypeptides were expressed, stable and correctly targeted to the cell membrane where they formed channels with altered properties. Analysis of the mutation distribution in these channels suggests that the AKT1 P domain has a structure similar to that of animal Shaker channels (a strongly constrained central region lining the tunnel that includes the highly conserved consensus motif TXXTXGYGD, and flanking regions forming the outer mouth of the pore), with an additional selectivity filter located upstream from the tunnel and formed by residues present in the N‐terminal flanking region.
Lateral domain diversity in membranes of callus and root cells of potato as revealed by EPR spectroscopy
Authors: Svetek Jelka , Kirn Borut , Vilhar Barbara , Schara Milan
Published: March 2003

Lateral heterogeneity in terms of co‐existing domains with a distinct molecular organization is an area of increasing interest in membrane biology. The structural and dynamic aspects of the in‐plane domain organization of lipids are becoming well documented, especially for model membrane systems. Potato (Solanum tuberosum L. cv. Desirée) callus cells and roots of plantlets from stem node culture were doped with a spin‐labeled analog of the methyl ester of palmitic acid bearing the paramagnetic nitroxide group at position C—5 of the acyl chain, which serves as a monitor of membrane fluidity of the region close to the polar phospholipid head groups of the bilayer. Model reconstruction of the line‐shapes of the experimental spectra revealed the co‐existence of two types of membrane domains with different ordering and dynamics of lipids in the membranes of both callus and root cells. With changes in temperature, relatively small differences were detected in either type of domain in the lipid ordering of the bilayer as characterized by order parameter S. However, the relative population of domains in the bilayer exhibited stronger temperature dependence. Typically, the relative proportion of disordered domains with less molecular order (smaller S) was larger in the membranes of callus cells compared to those of root cells, indicating higher fluidity throughout the measured temperature range (5–35°C). The Arrhenius activation energies for rearrangement of lipid molecules within the bilayer were found to be higher for root tissue membranes, indicating the ability of root cells to oppose actively any drastic changes of membrane structuring under temperature stress. The distinctions in organization of lateral domains between the callus and root cell membranes may be correlated with differences in growth rate and metabolic activity between these two types of tissue.
Purification and characterization of a safener‐induced glutathione S‐transferase from wheat (Triticum aestivum)
Authors: Pascal Sophie , Scalla René
Published: March 2003

The genome of cultivated wheat is hexaploid, and in consequence a large number of glutathione S‐transferase (GSTs, EC 2.5.1.18) isozymes is expected in that organism. Wheat GST subunits were first analyzed by reverse‐phase high performance liquid chromatography (RP‐HPLC). In root and shoot tissues, subunits 4, 8, and 9 were constitutively expressed whereas subunits 2, 3, and 5 were inducible by the herbicide safener naphthalic anhydride (NA). Significant differences were observed, however, between the distributions of these six major subunits in roots and shoots. A major GST isozyme was purified from the shoots of plants treated by NA. A combination of ammonium sulphate precipitation, hydrophobic interaction chromatography (HIC) and affinity chromatography resulted in purification with an apparent yield of 4.6% and a 48‐fold increase in specific activity toward 1‐chloro‐2,4‐dinitrobenzene (CDNB). Analysis by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) showed a single band at 24.5 kDa. Molecular mass estimated by nondenaturing PAGE was 49.5 kDa. These results suggest that the enzyme exists as a dimer. A pI of 5.2 was determined by native isoelectric focusing (IEF). Analysis by 2‐D electrophoresis showed a single spot, with a pI of 5.8–5.9. However, further analysis by RP‐HPLC revealed that the two subunits were different. They were characterized and identified by electrospray ionization mass spectrometry (ESI‐MS) as subunits 2 and 3, molecular masses 24 924±3 and 24 958±5 Da, respectively. Therefore, GST(2–3) is apparently a heterodimer consisting of subunits 2 and 3. Apparent KM values were 424 μM for CDNB and 228 μM for glutathione (GSH). GST(2–3) metabolized the herbicide fluorodifen, and a KM of 22 μM was determined for the herbicide.
Stomatal closure is induced rather by prevailing xylem abscisic acid than by accumulated amount of xylem‐derived abscisic acid
Authors: Jia Wensuo , Zhang Jianhua
Published: March 2003

We have studied the stomatal response in relation to the xylem‐derived abscisic acid (ABA) accumulation in sunflower leaves. When ABA was introduced into detached leaves of the sunflower through xylem flux, stomatal conductance was regulated, water flux was changed as a result and at the same time the xylem‐derived ABA was metabolised in the leaves. We computed the xylem‐derived ABA accumulation in the leaves as a function of time by taking into account the variation of ABA flux into the leaves (the product of water flux and ABA concentration) and a continuing ABA metabolism. We found that ABA accumulation was rapid during an initial lag phase, much slowed down during the decreasing phase of stomatal conductance, but still substantial when stomatal conductance reached a new stable state. The results show a poor link between the kinetics of ABA‐induced stomatal closure and the xylem‐derived ABA accumulation. Xylem‐derived ABA was metabolised rapidly in the leaves. Tetcyclacis, as an inhibitor, substantially inhibited this process. Two hours after ABA was fed into a leaf, about 70% of the fed ABA was metabolised, but when tetcyclacis was added into the feeding solution, less than 30% of ABA was metabolised, even after 24 h of incubation. The inhibition of ABA metabolism by tetcyclacis did not lead to more stomatal closure, which was still concentration‐dependent. Since the accumulation of xylem‐derived ABA was enhanced substantially by the presence of tetcyclacis, these results strongly indicate that stomata mainly respond to the prevailing ABA concentration in the xylem stream, rather than to the accumulated amount of xylem‐derived ABA in the leaves.
Effects of chronic ozone and elevated atmospheric CO2 concentrations on ribulose‐1,5‐bisphosphate in soybean (Glycine max)
Authors: Reid Chantal D. , Fiscus Edwin L. , Burkey Kent O.
Published: March 2003

Ribulose‐1,5‐bisphosphate (RuBP) pool size was determined at regular intervals during the growing season to understand the effects of tropospheric ozone concentrations, elevated atmospheric carbon dioxide concentrations and their interactions on the photosynthetic limitation by RuBP regeneration. Soybean (Glycine max [L.] Merr. cv. Essex) was grown from seed to maturity in open‐top field chambers in charcoal‐filtered air (CF) either without (22 nmol O3 mol−1) or with added O3 (83 nmol mol−1) at ambient (AA, 369 μmol CO2 mol−1) or elevated CO2 (710 μmol mol−1). The RuBP pool size generally declined with plant age in all treatments when expressed on a unit leaf area and in all treatments but CF‐AA when expressed per unit ribulose‐1,5‐bisphosphate carboxylase/oxygenase (Rubisco; EC 4.1.1.39) binding site. Although O3 in ambient CO2 generally reduced the RuBP pool per unit leaf area, it did not change the RuBP pool per unit Rubisco binding site. Elevated CO2, in CF or O3‐fumigated air, generally had no significant effect on RuBP pool size, thus mitigating the negative O3 effect. The RuBP pools were below 2 mol mol−1 binding site in all treatments for most of the season, indicating limiting RuBP regeneration capacity. These low RuBP pools resulted in increased RuBP regeneration via faster RuBP turnover, but only in CF air and during vegetative and flowering stages at elevated CO2. Also, the low RuBP pool sizes did not always reflect RuBP consumption rates or the RuBP regeneration limitation relative to potential carboxylation (%RuBP). Rather, %RuBP increased linearly with decrease in the RuBP pool turnover time. These data suggest that amelioration of damage from O3 by elevated atmospheric CO2 to the RuBP regeneration may be in response to changes in the Rubisco carboxylation.
Arabinogalactan proteins in embryogenic and non‐embryogenic callus cultures of Euphorbia pulcherrima
Authors: Saare‐Surminski Katja , Preil Walter , Knox J. Paul , Lieberei Reinhard
Published: March 2003

The arabinogalactan protein (AGP) fractions of embryogenic and non‐embryogenic callus lines of Euphorbia pulcherrima Willd. ex. Klotzsch were analysed over a cultivation period of 9 weeks using the β‐glucosyl Yariv reagent and an anti‐AGP antibody (LM2). The amount of AGPs detected with the Yariv reagent increased in embryogenic cultures during the development of somatic embryos. The embryogenic and non‐embryogenic callus contained different sets of AGPs characterized with the Yariv reagent and the LM2 monoclonal antibody. AGPs recognized by LM2 are localized primarily in the protodermal cells of globular somatic embryos. The development of somatic embryos of E. pulcherrima appears to be associated with the presence of particular AGPs.
Priming bitter gourd seeds with selenium solution enhances germinability and antioxidative responses under sub‐optimal temperature
Authors: Chen Chang Chi , Sung Jih Min
Published: March 2003

Priming offers an effective means for counteracting sub‐optimal temperature induced oxidative injury and raising seed performance in many crop species. In Taiwan, bitter gourd is frequently seeded when ambient temperatures are below the optimum. The objectives of the present study were to evaluate the effect of priming on germinability and antioxidative capacity in two bitter gourd cultivars germinated under 20 and 25°C conditions. Sodium selenite was also added to the priming solution. Priming was achieved by moistening the seeds with 1, 2, 5, or 10 mg l−1 sodium selenite solutions on germination paper at 25°C for 48 h and then air‐drying them to their original moisture levels. A sub‐optimal temperature of 20°C decreased seed performance by enhancing peroxidative injury. The sub‐optimal temperature germinability was partially restored by priming the seeds with selenium solutions up to the 2 mg l−1 level. The improved seed performances were linked to the priming‐enhanced free radical and peroxide‐scavenging activities linked to the ascorbate‐glutathione cycle. However, glutathione peroxidase was the only enzyme showing positive response with increasing selenium level up to 10 mg l−1, suggesting that this enzyme is selenium‐inducible. The changes in germination and antioxidative activities in relation to selenium priming and sub‐optimal temperature were similar for the two cultivars, despite differences in their germination performance and related antioxidative traits.
Light‐controlled growth of the maize seedling mesocotyl: Mechanical cell‐wall changes in the elongation zone and related changes in lignification
Authors: Schopfer Peter , Lapierre Catherine , Nolte Titus
Published: March 2003

Cell extension in the mesocotyl elongation zone (MEZ) of maize (Zea mays L.) seedlings is inhibited by light. The growth inhibition by blue light in the MEZ was reversible upon transfer to darkness. This experimental system was used for investigating the modification of mechanical cell‐wall properties and the role of cell‐wall lignification in cell elongation. The occurrence of lignin in the cortex and vascular bundle tissues of the MEZ was demonstrated by the isolation of diagnostic monomers released after thioacidolysis of the cell walls. Concomitantly with the inhibition of growth, blue light induces an increase in cell‐wall stiffness (tensile modulus) as well as an increase in extractable lignin in the outer MEZ tissues (cortex+epidermis). Both effects are reversed when growth is resumed in the MEZ in darkness after a period of growth inhibition induced by 3 h light. In the vascular bundle light produces no comparable change in lignin content. Appearance and disappearance of phenylpropanoid material in MEZ cell walls in the light, or in darkness following a brief light treatment, respectively, can be visualized under the fluorescence microscope by characteristic changes in autofluorescence of tissue sections upon excitation with UV radiation. It is concluded from these results that light‐induced lignification of primary walls is involved in cell‐wall stiffening and thus inhibition of elongation growth in the MEZ of maize seedlings. Resumption of growth upon redarkening may be initiated by wall loosening in the uppermost MEZ region which displaces the lignified cell walls towards the lower mesocotyl region.
N‐Bromosuccinimide modification of tryptophan 241 at the C‐terminus of the manganese stabilizing protein of plant photosystem II influences its structure and function
Authors: Yu Yong , Li Rong , Xu Chunhe , Ruan Kangcheng , Shen Yunkang , Govindjee
Published: March 2003

Life on earth depends upon the ability of oxygenic photosynthesis to oxidize water to molecular oxygen. This process is catalyzed by water–plastoquinone oxido‐reductase complex. In addition to the photosystem II (PSII) reaction core, it includes a manganese stabilizing protein (MSP) that plays an important regulatory role in the process in plants and algae. Tryptophan 241, located at the carboxyl‐terminus of the MSP, is its sole tryptophan. Modification of MSP by N‐bromosuccinimide (NBS) was carried out to explore the role of Trp241 in maintaining its structure and function. Data and arguments are presented to show that it is Trp241, not other tyrosines in MSP, that is involved in the modification and changes observed in this study. Further, the pH‐dependence of the modification and the comparison of features of fluorescence spectra of MSP suggested that Trp241 is buried in the hydrophobic interior of the protein. Hydropathy analysis revealed that Trp241 is located in the middle of the hydrophobic region at the C‐terminus of MSP. Circular dichroism spectroscopy showed that NBS modification of Trp241 dramatically modified the protein structure. The affinity of MSP to PSII decreased greatly after the modification of Trp241, and no oxygen‐evolving activity was recovered after its reconstitution. This study provides a novel demonstration that Trp241 at the C‐terminus hydrophobic region of the MSP is critical for maintaining appropriate structure and function of MSP.
Increased glutamine synthetase activity and changes in amino acid pools in leaves treated with 5‐aminoimidazole‐4‐carboxiamide ribonucleoside (AICAR)
Authors: Man Hui‐Min , Kaiser Werner M.
Published: March 2003

Feeding 5‐aminoimidazole‐4‐carboxiamide ribonucleoside (AICAR) through the petiole of detached young barley leaves rapidly increased activities of NADH‐nitrate reductase (NR) and glutamine synthetase (GS) in leaf extracts and at least partly prevented the usual slow decrease of these enzyme activities during prolonged illumination. Further, AICAR caused drastic changes in amino acid levels: glutamine and serine levels were increased whereas glutamate and glycine were decreased, probably indicating a higher GS activity and more rapid conversion of glycine into serine. The latter may be responsible for the higher ammonium contents found in AICAR treated leaves. We tentatively suggest that GS (located in the chloroplast) and glycine decarboxylase (located in the mitochondria) are regulated in a manner similar to NR. This is discussed in the light of recent reports that 14‐3‐3 isoforms exist in chloroplasts and that GS binds to 14‐3‐3s in vitro.
Seasonal fluctuations in the concentration of UV‐absorbing compounds in the leaves of some Mediterranean plants under field conditions
Authors: Liakoura V. , Manetas Y. , Karabourniotis G.
Published: March 2003

Leaves of 14 representative Mediterranean plant species were collected on a monthly basis and assayed for UV‐absorbing compounds concentration, either on an area or a dry mass basis, from 1995 to 1997. Strong seasonal fluctuations were observed in eight species (all evergreens, two phrygana, one deciduous, one summer perennial and one winter perennial). Two different patterns of changing concentrations of UV‐absorbing compounds were observed. In the first, concentration of these compounds was higher in young developing leaves and concentration declined during maturation, whereas in other plants, the opposite trend was observed. These differences could be attributed to the particular leaf surface morphology of each plant. The observed seasonal fluctuations of UV‐absorbing compounds seem to be more correlated to developmental processes, than to seasonal fluctuations of the naturally occurring UV‐B radiation. Most of the winter perennials did not show strong fluctuations during the period of development. The concentration of these compounds varied not only on a seasonal basis among the examined plants, but between different life forms as well: during winter, examination of the leaves of 13 species showed that evergreen sclerophylls and phrygana had at least two‐fold higher concentration of UV‐B‐absorbing compounds on a leaf area basis than winter perennials. In addition, during the same season and irrespective of life form and species, the absorbance at 300 nm per unit of mature leaf area followed an asymptotic exponential decrease when specific leaf area increased. The UV‐B radiation screening capacity of the leaves of these plants is discussed in relation to each adaptive strategy.
Temporal and tissue localization of a cowpea (Vigna unguiculata) cystatin
Authors: Flores Victor M. Q. , Louro Ricardo P. , Xavier‐Filho José , Barratt Daniel H. P. , Shewry Peter R. , Fernandes Kátia V. S.
Published: March 2003

A cowpea (Vigna unguiculata L. Walpers cv. Pitiúba) cystatin was analysed to determine its localization during development and germination of cowpea seeds, using western blotting with a specific antiserum. The pattern of immunoreactive proteins changed during development, with the major reactive bands present in dried seeds being mobilized after a 62‐h period of imbibition. Immunohistochemical analysis revealed that cowpea cystatin is distributed in both embryonic axes and cotyledons with the highest level being present in the outer cell walls of the adaxial surface of the cotyledons.
Two distinct mechanisms regulate the transcription of photosystem II genes in Synechocystis sp. PCC 6803
Authors: Herranen Mirkka , Aro Eva‐Mari , Tyystjärvi Taina
Published: March 2003

Expression and regulation of psb genes, encoding various subunits of photosystem II (PSII), were studied in the cyanobacterium Synechocystis sp. PCC 6803. Transcription of the psbA and psbD genes, encoding the PSII reaction centre proteins D1 and D2, was rapidly activated upon onset of illumination and the transcription rates were enhanced at high irradiance. Gel retardation analysis demonstrated dark‐enhanced binding of proteins to the upstream region of the psbA2 gene, pointing to a repressor‐protein‐based transcriptional regulation mechanism. Transcription of all the other psb genes also required light, but unlike the psbA and psbD genes, these psb genes did not respond specifically to high‐light. Moreover, the transcription of these psb genes was activated slowly at onset of illumination, and was strictly dependent on de novo protein synthesis. We suggest that these psb genes are up‐regulated in the light via transcriptional activator proteins, and the slow activation may be related to production of new PSII centres during growth. Apart from the two distinct mechanisms for transcriptional regulation, all psb genes shared a common regulation mechanism at the level of transcript stability, mediated by the redox poise of intersystem electron carrier(s).