bluuebunny/crossref_metadata_2025 huggingface dataset:
The molecular characterization of a set of cDNAs differentially expressed during Douglas‐fir germination and early seedling development
Authors: Tranbarger Timothy J. , Misra Santosh
Published: May 2006

We constructed a cDNA library with mRNA pooled from 4‐ to 6‐day‐old Douglas‐fir (Pseudotsuga menziesii [Mirb.] Franco) seedlings. To identify genes with increased expression during germination and early seedling development, the library was screened differentially with cDNA probes synthesized using mRNA isolated from mature seeds and 6‐day‐old seedlings. Partial DNA sequence analysis and predicted amino acid sequences revealed cDNA clones with homology to several plant proteins including: a chaperonin 60β (cpn60β), a low molecular weight heat shock protein (LMW HSP), a luminal binding protein (BiP), a type II chlorophyll a/b‐binding protein (CAB) and a cysteine protease. In mature seeds, transcripts corresponding to each cDNA clone were detected and their levels increased after 2 days of exposure to the germination conditions. The clone that shares identity with the cysteine protease (DF3‐3) and DF6‐7 had peak transcript levels during the germination phase, whereas the cpn60β (DF3), a LMW HSP (DF4‐5), BiP (DF5‐1) and CAB (DF6‐3) showed peaks during the post‐germination phase. All the transcripts were present at lower levels in 10‐week‐old needles except for CAB. As a comparison, an additional clone which was present at similar levels in both mature seeds and in 6‐day‐old seedlings was isolated and found to be homologous to a NADPH‐cytochrome P450 reductase (CPR) (EC 1.6.2.4). The expression of CPR changed little during germination and early seedling development with lower transcript levels in 10‐week‐old needles. The significance of the individual genes, their expression, and possible roles during germination is discussed.
Purification and characterization of a 22‐kDa protein in chloroplasts from green spores of the fern Osmunda japonica
Authors: Inoue Hiroshi , Takano Atuo , Asano Yasuhisa , Katoh Yasuo
Published: May 2006

Changes in the polypeptide composition of chloroplasts were investigated during germination of green spores of the fern Osmunda japonica. The polypeptide composition of chloroplasts was appreciably changed during a germination time course of 48 h. Levels of five polypeptides with apparent molecular masses of 47, 44, 42, 22 and 18.5 kDa in the soluble fraction of chloroplasts and three polypeptides with molecular masses of 24, 22 and 15 kDa in the thylakoid membranes decreased during germination. In contrast, no decrease of chloroplast polypeptides was observed in the spores incubated with cycloheximide for 48 h. A new 22‐kDa protein was isolated from thylakoid membranes of spores and the amino‐terminal sequence of the purified protein was determined. High levels of alanine and glycine were found in the basic protein (pl > 10.3). This protein, with a native molecular mass of 80 kDa, was characterized by a subunit band observed at a molecular mass of 22 kDa on SDS‐PAGE and by the disappearance of the band during spore germination. Protease activity against the 22‐kDa protein was observed in an extract prepared from chloroplasts of quiescent spores. A hypothetical cytosolic proteinaceous factor is implicated in the regulation of protein degradation in chloroplasts.
Hyphal contribution to water uptake in mycorrhizal plants as affected by the fungal species and water status
Authors: Ruiz‐Lozano J. M. , Azcón R.
Published: May 2006

Vesicular‐arbuscular mycorrhizae may increase resistance of plants to drought by a number of mechanisms, such as increased root hydraulic conductivity, stomatal regulation, hyphal water uptake and osmotic adjustment. However, a substantial contribution of vesicular‐arbuscular mycorrhizal (VAM) hyphae to water uptake has not been demonstrated unequivocally. The objective of this investigation was to examine the contribution of hyphae from two VAM fungi to water uptake and transport by the host plant. Lettuce (Lactuca sativa L.) plants were grown in a container divided by a screen into two compartments. One was occupied by roots, the other only by VAM hyphae, which the screen permitted to pass. Roots were colonized by the VAM fungi Glomus deserticola or Glomus fasciculatum, or were left uninoculated but P‐supplemented. Water was supplied to the hyphal compartment at a distance of 10 cm from the screen (root). CO2 exchange rate, water‐use efficiency, transpiration, stomatal conductance and photosynthetic phosphorus‐use efficiency of VAM or P‐amended control plants were evaluated at three levels of water application in the hyphal compartment. Results indicate that much of the water was taken up by the hyphae in VAM plants. VAM plants, which had access to the hyphal compartment, had higher water and nutrient contents. G. deserticola functioned efficiently under water limitation and mycelium from G. fasciculatum‐colonized plants was very sensitive to water in the medium. This discrepancy in VAM behaviour reflects the various abilities of each fungus according to soil water levels. Different abilities of specific mycelia were also expressed in terms of nutritional and leaf gas‐exchange parameters. G. fasciculatum caused a significant increase in net photosynthesis and rate of water use efficiency compared to G. deserticola and P‐fertilized plants. In contrast, the G. deserticola treatment was the most efficient affecting N, P and K nutrition, leaf conductance and transpiration. Since no differences in the intra‐ and extra‐radical hyphal extension of the two endophytes were found, the results demonstrate that mycorrhizal hyphae can take up water and that there are considerable variations in both the behaviour of these two VAM fungi and in the mechanisms involved in their effects on plant water relations.
Quantitative analysis of the control of physiological phenomena by plant hormones
Authors: Weyers Jonathan D. B. , Paterson Neil W. , A'Brook Richard , Peng Zhi‐Yong
Published: May 2006

We highlight the need to combine information about hormone concentrations and tissue sensitivity when assessing control of physiological phenomena by plant hormones. A quantitative approach to this problem is suggested and the method illustrated using data for stomatal closure induced by abscisic acid. The technique requires accurate estimates of hormone concentration at the putative site of action and information about the kinetics of responses in the form of dose‐response curves obtained under relevant conditions. Extending this approach, hormonal control is seen as relative, in that it must be considered against a background of other controlling influences. Possible means of evaluating this ‘control potential’ from dose‐response curves are discussed. Finally, we draw together the above in an analytical framework for assessing hormonal control. This involves (1) matching observed and predicted responses; (2) studying effects of manipulating hormone concentration or sensitivity; and (3) assessing the relative control potential of the compound of interest.
Phase reversal in Sequoia sempervirens in relation to mtDNA
Authors: Huang Li‐Chun , Lin Lih‐Yuan , Chen Chung‐Mong , Chen Li‐Jeng , Huang Bau‐Lian , Murashige Toshio
Published: May 2006

Restriction fragment analysis revealed differences in mitochondrial DNA (mtDNA) between adult and juvenile coastal redwood, Sequoia sempervirens (D. Don) Endl. Thus, for example, juvenile shoots contained 4.0‐ and 3.6‐kb BamHI], restricted fragments that were absent in adult material. Rejuvenation by 5 repetitive graftings onto juvenile rootstocks resulted in the same 4.0‐ and 3.6‐kb fragments as were present in the juvenile material. The similarities in mtDNA fragments of juvenile and rejuvenated. shoots, as well as their distinction from adult shoots, were confirmed by double digestion, Southern hybridization and DNA sequencing.
Properties of proton and sugar transport at the tonoplast of tomato (Lycopersicon esculentum) fruit
Authors: Milner I. D. , Ho L. C. , Hall J. L.
Published: May 2006

Tonoplast vesicles were isolated from tomato (Lycopersicon esculentum Mill.) fruit pericarp and purified on a discontinuous sucrose gradient. ATPase activity was inhibited by nitrate and bafilomycin A1 but was insensitive to vanadate and azide. PPase hydrolytic activity was inhibited by NaF but was insensitive to nitrate, bafilomycin A1 vanadate and azide. Kimetic studies of PPase activity gave an apparent Km, for PP3 of 18 μM. Identical distributions of bafilomycin‐ and NO3‐sensitive ATPase activities within continuous sucrose density gradients, confirmed that bafilomycin‐sensitive ATPase activity is a suitable marker for the tonoplast. By comparing the distribution of bafilomycin‐sensitive ATPase activity with that of PPase activity, it was possible to locate the PPase enzyme exclusively at the tonoplast. The apparent density of the tonoplast did not change during fruit development. Measurements of tonoplast PPase and ATPase activities during fruit development over a 35‐day period revealed an 80% reduction in PPase specific activity and a small decrease in ATPase specific activity. ATP‐ and PP1‐dependent ΔpH generation was measured by the quenching of quinacrine fluorescence in tonoplast vesicles prepared on a discontinuous Dextran gradient. No H+ efflux was detected on the addition of sucrose to energized vesicles. Therefore a H+/sucrose antiport may not be the mechanism of sucrose uptake at the tomato fruit tonoplast. Similar results were obtained with glucose, fructose and sorbitol. The lack of ATP (or PP1) stimulation of [14C]‐sucrose uptake also suggested that an antiport was not involved. Initial uptake rates of radiolabelled glucose and fructose were almost double that for sucrose. The inhibition of hexose uptake by p‐chloromercuribenzene sulphonate (PCMBS) implicated the involvement of a carrier. Therefore storage of hexose in the tomato fruit vacuole and maintenance of a downhill sucrose concentration gradient into sink cells is likely to be regulated by the activity of sucrose metabolizing enzymes, rather than by energy‐requiring uptake mechanisms at the tonoplast.
Sugar degradation during autoclaving: Effects of duration and solution volume on breakdown of glucose
Authors: Wang Xiao‐Jia , Hsiao Ke‐Cheng
Published: May 2006

As the autoclaving of a sugar together with other nutrient components enhances its degradation with the associated formation of toxic products, it is advisable to autoclave it separately from other medium components. In such cases, and to prepare growth media of consistent quality, it may also be necessary to adjust the duration of autoclaving according to the volume of the sugar solution. The results of an attempt to fine‐adjust the autoclaving procedure are presented.

Glucose, used as model sugar in this investigation, was autoclaved for 20,32 or 44 min in volumes of 0.25. 1.2 and 5 1. The rise in temperature of the solutions was monitored by thermocouples. Glucose degradation was estimated by the rate of cyanide‐initiated oxygen consumption. The rates were found to be inversely proportional to the volumes in a semilogarithmic plot. The T‐t (temperature‐time) value, the area under the solution temperature vs time curve, was found to be an independent variable from which the extent of glucose degradation could be deduced. For a chosen level of thermodegradation of glucose, the duration of autoclaving for solutions of different volumes could be determined graphically from a T‐t value vs time plot.
Re‐examination of the localization of Mg‐chelatase within the chloroplast
Authors: Walker Caroline J. , Weinstein Jon D.
Published: May 2006

In a plastid‐free assay, Mg‐chelatase from pea (Pisum sativum L. cv. Spring) and cucumber (Cucumis sativus L. cv. Sumter) chloroplasts is inhibited to equal extents by the mercurial reagents. p‐chloromercuribenzoate (PCMB) and p‐chloromercuribenzene sulfonate (PCMBS). However, in intact chloroplasts PCMB inhibits Mg‐chelatase fourfold more strongly than does PCMBS. Since PCMBS cannot penetrate membranes as readily as PCMB, Mg‐chelatase may be localized interior to the inner chloroplast envelope. When Mg‐chelatase is assayed with photosynthetically generated ATP, the presence of an external ATP trap does not inhibit activity, suggesting that the enzyme is not located in the interenvelope space. None of the components of Mg‐chelatase are integral membrane proteins: Mg‐chelatase activity is readily solubilized by washing the total chloroplast membranes in buffers of low MgCl2 content. This precludes localization by purifying individual thylakoid and envelope membranes which requires low MgCl2 concentrations.
Enriched rhizosphere CO2 concentrations can ameliorate the influence of salinity on hydroponically grown tomato plants
Authors: Cramer M. D. , Lips S. H.
Published: May 2006

Our previous work indicated that salinity caused a shift in the predominant site of nitrate reduction and assimilation from the shoot to the root in tomato plants. In the present work we tested whether an enhanced supply of dissolved inorganic carbon (DIC, CO2+ HCO3) to the root solution could increase anaplerotic provision of carbon compounds for the increased nitrogen assimilation in the root of salinity‐stressed Lycopersicon esculentum (L.) Mill. cv. F144. The seedlings were grown in hydroponic culture with 0 or 100mM NaCl and aeration of the root solution with either ambient or CO2‐enriched air (5000 μmol mol−1). The salinity‐treated plants accumulated more dry weight and higher total N when the roots were supplied with CO2‐enriched aeration than when aerated with ambient air. Plants grown with salinity and enriched DIC also had higher rates of NO−3 uptake and translocated more NO−3 and reduced N in the xylem sap than did equivalent plants grown with ambient DIC. Incorporation of DIC was measured by supplying a 1 ‐h pulse of H14CO−3 to the roots followed by extraction with 80% ethanol. Enriched DIC increased root incorporation of DIC 10‐fold in both salinized and non‐salinized plants. In salinity‐stressed plants, the products of dissolved inorganic 14C were preferentially diverted into amino acid synthesis to a greater extent than in non‐salinized plants in which label was accumulated in organic acids. It was concluded that enriched DIC can increase the supply of N and anaplerotic carbon for amino acid synthesis in roots of salinized plants. Thus enriched DIC could relieve the limitation of carbon supply for ammonium assimilation and thus ameliorate the influence of salinity on NO−3 uptake and assimilation as well as on plant growth.
Alterations in photosynthesis and pigment distributions in pea leaves following UV‐B exposure
Authors: Day T. A. , Vogelmann T. C.
Published: May 2006

We compared photosynthetic and UV‐B‐absorbing pigment concentrations, gas‐exchange rates and photosystem II (PSII) electron transport rates in leaves of pea (Pisum sativum mutant Argenteum) grown without UV‐B or under an enhanced UV‐B treatment (18 kJ m−2 biologically effective daily dose) in a greenhouse. We also compared the distribution of chlorophyll by depth within leaves of each treatment by using image analysis of chlorophyll autofluorescence. Ultraviolet‐B treatment elicited putative protective responses such as an 80% increase in UV‐B‐absorbing compound concentrations (leaf‐area basis), and a slight increase in mesophyll thickness (178 in controls compared to 191 μm in UV‐B‐treated leaves). However, photosynthetic rates of UV‐B‐treated leaves were only 80% of those of controls. This was paralleled by reductions in leaf conductance to water vapor (50% of controls) and intercellular CO2 concentrations, suggesting that stomatal limitations were at least partly responsible for lower photosynthetic rates under the UV‐B treatment. Total chlorophyll concentrations (leaf‐area basis) in UV‐B‐treated leaves were only 70% of controls, and there was a shift in the relative distribution of chlorophyll with depth in UV‐B‐treated leaves. In control leaves chlorophyll concentrations were highest near the adaxial surface of the upper palisade, dropped with depth and then increased slightly in the bottom of the spongy mesophyll nearest the abaxial surface. In contrast, in UV‐B‐treated leaves chlorophyll concentrations were lowest at the adaxial surface of the upper palisade and increased with depth through the leaf. The most notable treatment difference in chlorophyll concentrations was in the upper palisade near the adaxial surface of leaves, where we estimate that chlorophyll concentrations in each 1‐μm‐thick paradermal layer were about 50% lower in UV‐B‐treated leaves than in controls. We found reduced electron transport capacity in UV‐B‐treated leaves, based on lower maximum fluorescence (Fm), variable to maximum fluorescence ratios (F,/Fm) and quantum yield of PSII electron transport (Y). However, the above were assessed from fluorometer measurements on the adaxial leaf surface and may reflect the markedly lower chlorophyll concentrations in the upper palisade of UV‐B‐treated leaves.
Lipid peroxidation and peroxide‐scavenging enzymes associated with accelerated aging and hydration of watermelon seeds differing in ploidy
Authors: Chiu K. Y. , Wang C. S. , Sung J. M.
Published: May 2006

Hydration offers an effective means for raising seed performance in many crop species. The objective of this study was to evaluate the effect of vermiculite hydration on germinability and several physiological activities related to vigor in artificially aged watermelon seeds differing in ploidy. Aging was achieved by incubating the seeds at 45°C and 79% relative humidity for 6 days, then the seeds were air‐dried to their original moisture level (4.7%). Hydration was achieved by mixing the untreated and aged seeds with moist vermiculite No. 3 at 25°C for 24 h. The partially hydrated seeds were air‐dried at 25°C for 36 h to 4.7% moisture level. Significant differences existed between unaged and aged seeds, with lower germination percentage and slower germination speed in the latter. Aging also increased lipid peroxidation and reduced the activity of peroxide‐scavenging enzymes. The germinability of aged watermelon seed was restored partially by vermiculite hydration. The activities of protein synthesis and peroxide‐scavenging enzymes in axis and cotyledon portions of the seeds were also increased by hydration treatment. The changes in germination and related physiological responses in relation to aging and hydration are similar in seeds differing in ploidy, despite differences in their germination performance, seed leakage, extent of lipid peroxidation and activities of peroxide‐scavenging enzymes.
Effect of temperature on biomass allocation in tomato (Lycopersicon esculentum)
Authors: Heuvelink E.
Published: May 2006

Temperature may influence dry matter partitioning between fruits and vegetative plant parts either directly or indirectly through its influence on development, flower and/or fruit abortion. The objective of the present work was to investigate whether there is any direct effect of temperature on dry matter partitioning between fruits and vegetative plant parts in tomato. A greenhouse experiment was conducted, with alternating 3‐week periods of high (23°C) and low (18°C) temperature setpoint. Dry matter partitioning during these 3‐week periods was determined from destructive plant harvests at two levels of fruit pruning (3 and 7 fruits per truss). Indirect temperature effects on dry matter partitioning were excluded by fruit pruning.

On average, the fraction of dry matter distributed to the fruits during a 12‐week period, starting with the flowering of the fifth truss (28 days after planting), was 0.53 (3 fruits per truss) and 0.70 (7 fruits per truss). These ratios were also calculated for every 3‐week period separately and did not depend on the average temperature (18–24°C) during that period.

It is concluded that dry matter distribution in tomato is not significantly affected by temperature directly, which means that the temperature effect (18–24°C) on the generative sink strength is not much different from the temperature effect on the vegetative sink strength.
Effect of apex excision and replacement by 1‐naphthylacetic acid on cytokinin concentration and apical dominance in pea plants
Authors: Li C.‐J. , Guevara E. , Herrera J. , Bangerth F.
Published: May 2006

As known from literature lateral buds from pea (Pisum sativum) plants are released from apical dominance when repeatedly treated with exogenous cytokinins. Little is known, however, about the endogenous role of cytokinins in this process and whether they interact with basipolar transported IAA, generally regarded as the main signal controlling apical dominance. This paper presents evidence that such an interaction exists.

The excision of the apex of pea plants resulted in the release of inhibited lateral buds from apical dominance (AD). This could be entirely prevented by applying 1‐naphthylacetic acid (NAA) to the cut end of the shoot. Removal of the apex also resulted in a rapid and rather large increase in the endogenous concentrations of zeatin riboside (ZR), isopentenyladenosine (iAdo) and an as yet unidentified polar zeatin derivative in the node and internode below the point of decapitation. This accumulation of ZR and iAdo, was strongly reduced by the application of NAA. The observed increase in cytokinin concentration preceded the elongation of the lateral buds, suggesting that endogenous cytokinins play a significant role in the release of lateral buds from AD. However, the effect of NAA on the concentration of cytokinins clearly demonstrated the dominant role of the polar basipetally transported auxin in AD. The results suggest a mutual interaction between the basipolar IAA transport system and cytokinins obviously produced in the roots and transported via the xylem into the stem of the pea plants.
Dynamic changes in leaf bulk water relations during stomatal oscillations in mangrove species. Continuous analysis using a dewpoint hygrometer
Authors: Herppich Werner B. , von Willert Dieter J.
Published: May 2006

Plants of the mangrove species Pelliciera rhizophoreae and Avicennia germinans, exhibit pronounced oscillations in stomatal aperture under certain climatic conditions. During these oscillations, changes in transpirational water loss were closely followed by those in leaf water potential (ψ1) as indicated by continuous monitoring with an in situ dewpoint hygrometer. With this instrument, it was possible to measure dynamic changes in ψ1 for several days under constant conditions. Subsequently, the leaf was detached from the shoot and a pressure‐volume (PV) curve was established by repeatedly weighing the leaf, still attached to the hygrometer during short interruptions of the water potential recordings. The pressure‐volume relationship was then used to derive other water relations parameters from these water potential data. Thus, the procedure described herein allows a continuous analysis of the relevant components of bulk leaf water relations. Oscillations in water potential were also measured with single leaves using a pressure chamber. Water relations data obtained with these two different methods were in good agreement. In addition, osmotic potentials derived from the PV‐analysis were well within the range of those determined cryoscopically using extracted cell sap.
Purification and characterization of an algal (Dunaliella tertiolecta) protein cross‐reacting with an anti‐Ga‐common antibody
Authors: Takahashi Koji , Yuasa Takashi , Muto Shoshi
Published: May 2006

A 26‐kDa and a 36‐kDa protein that cross‐reacted with anti‐Ga‐common and anti‐Gβ antibodies, respectively, were detected in Dunaliella cells. The 26‐kDa protein was solubilized from a crude membrane fraction with deoxycholate and purified to homogeneity by DE52 and hydroxylapatite chromatography and DEAE‐5PW high performance liquid chromatography (HPLC). The hydroxylapatite‐purified preparation had GTPγS binding and GTPase activities, but the homogeneous 26‐kDa protein had none. The sequence of the 28 N‐terminal amino acids of the 26‐kDa protein had no homology to any GTP binding protein thus far reported.
Rheological analysis of viscoelastic cell wall changes in maize coleoptiles as affected by auxin and osmotic stress
Authors: Hohl Michael , Schopfer Peter
Published: May 2006

The effects of auxin and osmotic stress on elongation growth of maize (Zea mays L.) coleoptile segments are accompanied by characteristic changes in the extensibility of the growth‐limiting cell walls. At full turgor auxin causes growth by an increase in wall extensibility (wall looseining). Growth can be stopped by an osmotically produced step‐down in turgor of 0.45 MPa. Under these conditions auxin causes the accumulation of a potential for future wall extension which is released after restoration of full turgor. Turgor reduction causes a reversible decrease in wall extensibility (wall stiffening) both in the presence and absence of auxin. These changes in vivo are correlated with corresponding changes in the rheological properties of the cell walls in vitro which can be traced back to specific modifications in the shape of the hysteretic stress‐strain relationship. The longitudinally load‐bearing walls of the coleoptile demonstrate almost perfect viscoelasticity as documented by a nearly closed hysteresis loop. Auxin‐mediated wall loosening causes an increase of loop width and thus affects primarily the amount of hysteresis in the isolated wall. In contrast, turgor reduction by osmotic stress reduces loop length and thus affects primarily the amount of viscoelastic wall extensibility. Pretreatment of segments with anoxia and H2O2 modify the hysteresis loop in agreement with the conclusion that the wall‐stiffening reaction visualized under osmotic stress in vivo is an O2‐dependent process in which O2 can be substituted by H2O2. Cycloheximide specifically inhibits auxin‐mediated wall loosening without affecting wall stiffening, and this is mirrored in specific changes of the hysteresis loop. Corroborating a previous in vivo study (Hohl et al. 1995, Physiol. Plant. 94: 491–498) these results show that cell wall stiffening in vivo can also be demonstrated by Theological measurements with the isolated cell wall and that this process can be separated from cell wall loosening by specific changes in the shape of the hysteresis loop.
Solute leakage, lipid peroxidation, and protein degradation during the senecence of Iris tepals
Authors: Celikel Fisun G. , van Doorn Wouter G.
Published: May 2006

A colour change and inrolling of the tepal edges are the first symptoms of senescence of Iris flowers (Iris x hollandica Tub., cv. Blue Magic). Tepals showed an increase in leakage of both ions and anthocyanins, prior to the visible senescence symptoms. Increased leakage occurred irrespective of the time at which the tepals were severed and placed in water, indicating that the senescence process is inherent in the tepal cells. Net loss of proteins in the tepal edges started after flower opening, and after two more days, when the first symptoms of senescence were observed, the protein level was only 20% of that at harvest. Cycloheximide delayed senescence and resulted in a lower rate of protein loss. Phenylmethylsulfony fluoride (PMSF), a protease inhibitor, had a similar effect on protein levels but did not affect the time to visible senescence, and also several other protease inhibitors did not affect the time to senescence.

During senescence the rate of respiration of the tepals remained unchanged and their rate of ethylene production decreaased. The rate of ethane production, an indicator of lipid peroxidation, was very low and remained unaltered. Antioxidants (lascorbic acid, benzoic acid, butylated hydroxytoluene, diphenylamine, propyl gallate, propyl‐p‐hydroxybenzoate and sodium benzoate) had no effect on the time to tepal senescence. It is concluded that tepal wilting is due to transfer of solutes from the symplast to the apoplast. Although net protein degradation occurs early during the senescence process, its inhibition is not correlated with a delay in the time to senescence. Furthermore, the results do not support the hypothesis that the increase in solute leakage is due to (free radical‐mediated) peroxidation of membrane lipids. The present results are in contrast with the ethylene‐regulated petal senescence of carnation, which is accompanied by lipid peroxidation.
Expression of chloroplastic and cytosolic glutamine synthetase in barley leaves after cold‐sensitive blocking of β‐carotene synthesis by amitrole or mutation
Authors: Zito Francesca , Gough Simon , Marcato Caterina , Rascio Nicoletta , Casadoro Giorgio
Published: May 2006

3‐Amino‐ 1,2,4‐triazole (amitrole) provided to germinating barley at 20°C in the light led to bleached seedling leaves and photodynamic destruction of chloroplast structure, whereas normal greening and chloroplast ultrastructure was obtained when the seedlings developed in the presence of amitrole in the light at 30°C. Mass spectrometric analysis of the extractable herbicide demonstrated the same content of amitrole in leaves developed at 20 and 30°C. A very similar temperature‐sensitive syndrome is characteristic for the nuclear gene mutant ligrina‐o34 in barley. Amitrole and the mutation were shown to inhibit the cyclization of lycopene, leading to severe deficiencies in β‐carotene and its xanthophyll derivative lutein. Besides accumulation of lycopene, also its precursors phytoene, phytofluene and ξ‐carotene accumulated. Inhibition of carotenoid biosynthesis by amitrole and the mutation at 20°C in the light led to a strong reduction of both transcript and protein levels for chloroplastic glutamine synthetase (GS2) while transcript amount and protein of the cytosolic isoenzyme (GS1) were unaffected. At 30°C increased levels of mRNA for the chloroplastic isoform GS2 were observed in wild type, mutant and amitrole‐treated seedlings, but protein levels remained unchanged. Turnover rates of the GS2 protein were the same at 20 and 30°C. This extensive translational control of chloroplastic GS2 synthesis was also observed in a heat shock experiment, which revealed transiently increased mRNA levels for chloroplastic GS2 but unchanged protein levels.

Permissive synthesis of β‐carotene and chloroplastic glutamine synthetase (GS2) at 30°C in the presence of amitrole or the tigrina‐o34 mutation might be due to two alternative pathways of ionone ring formation using either lycopene or neurosporene as substrates for cyclization.
Characterization of the blue light‐induced extracellular alkalinization associated with the monovalent anion uptake byMonorapidium braunii.Competition between NO3and Cl−
Authors: Witt Federico G. , Aparicio Pedro J.
Published: May 2006

The blue light‐elicited monovalent anion‐dependent alkalinization of the medium ofMonoraphidium braunü(Legnerová, 202–7d) was characterized for the NO‐3and Cl‐ uptake. The maximal H+uptake rates for these two anions have a similar optimum pH around 8.5, and quite similar Ksvalues for high (38 üMfor Cl‐ and 35üMfor NO‐3) and low (320 üMfor Cl‐ and 335 üMfor NO‐3) affinities. The steady H+uptake associated with the uptake and reduction of NO‐3showed a Ksof 125 üM. which in this alga corresponds to the NO‐3reductase (EC 1.6.6.2) Kmfor NO‐3. The only and striking difference found in the uptake properties of these anions was the delay time between the switching on of the blue light and the start of the alkalinization, which increased from 10 to 90 s as the initial pH decreased from 8.5 to 6.5 in the presence of NO‐3, whereas for Cl‐ uptake this delay time (10s) did not vary in relation to the initial pH. When the NO‐3concentration in the medium was low (100 üM), the presence of relatively high concentrations of Cl‐ (3 üM), on the one hand, greatly stimulated the maximal alkalinization rates but, on the other, Cl‐ severely reduced the steady NO‐3‐dependent rate of alkalinization. The data indicate that Cl‐ inhibits competitively NO‐3uptake with a Kiof 750 üM. Moreover, high concentrations of NO‐3(above 5 üM) reduced its own maximal, but not the steady, uptake rates. The above results allow us to propose that most of the components of the individual NO‐3and Cl‐ transport systems are under identical light control and, as the competition data suggest, that these two anions may be taken up by the same transport system.
Involvement of NAD(P)+‐glutamate dehydrogenase isoenzymes in carbon and nitrogen metabolism in Chlamydomonas reinhardtü
Authors: Moyano E. , Cárdenas J. , Muñoz‐Blanco J.
Published: May 2006

The involvement of three NAD(P)+‐L‐glutamate dehydrogenase (GDH; EC 1.4.1.3) isoenzymes, named GDH1, GDH2 and GDH3, in the carbon and nitrogen metabolism of the green alga Chlamydomonas reinhardtü 6145c has been investigated under different environmental and stress conditions. GDH1 activity decreased, but GDH2 and GDH3 activities increased with the age of cultures. When the extracellular ammonium concentration was high, only GDH1 activity increased with growth whereas GDH2 and GDH3 remained unchanged. In the presence of L‐methionine‐D,L‐sulfoximine (MSX), an inhibitor of L‐glutamine synthetase (GS), a significant increase of GDH1 and a slight decrease in GDH3 activity was observed, whereas GDH2 did not change. A significant increase in the intracellular 2‐oxoglutarate was also found upon addition of azaserine, an inhibitor of L‐glutamate synthase (GOGAT) activity. However, no significant changes in GDH isoenzyme activities were observed after addition of azaserine or azaserine plus MSX, except an induction of GDH3 in the latter case. Moreover, in the presence of ethoxyzolamide (ETZ), an inhibitor of carbonic anhydrase activity, an induction of total GDH activity, mainly due to an increase in GDH1 and to a minor extent in GDH2, was observed in cells under low CO2 (0.03%). In the dark, cells showed an increase in GDH1 activity, but when acetate was present GDH1 activity was repressed. All these results taken together suggest a relationship between GDH1 and nitrogen assimilation, whereas GDH2 and GDH3 seem to be involved in the production of 2‐oxoglutarate to fuel the tricarboxylic acid (TCA) cycle.
Purification and properties of glutamine synthetase from Douglas fir roots
Authors: Bedell Jean‐Philippe , Chalot Michel , Brun Annick , Botton Bernard
Published: May 2006

Glutamine synthetase (GS. EC 6.3.1.2) was purified to apparent electrophoretic homogeneity from roots of Pseudotsuga menziesii (Mirb) Franco by a three‐step procedure involving diethylaminoethyl (DEAE)‐Trisacryl chromatography, affinity chromatography on Matrex Gel Red A. and preparative polyacrylamide gel electrophoresis. The enzyme was purified 40‐fold with a 16% recovery. The native enzyme had a molecular mass of 460 ± 5 kDa as estimated by gel filtration, interpolation of the Ferguson plots and non‐denaturing gradient‐PAGE. It was composed of two different subunits of 54 and 64 kDa. Affinity constants for glutamate (Glu), glutamine (Gln), ATP and ADP were 2.6, 10.5, 0.5 and 0.083 mM. respectively. The enzyme exhibited a negative cooperativity for ammonium (Hill number of 0.7) with two Km values which were 11 and 75 μM in the presence of ammonium concentrations lower and higher than 1.3 mM, respectively. Glycine and ADP appeared as potential inhibitors of the GS activity. The optimum pH values were 7.2 and 7.6 for the transferase and the biosynthetic assays, respectively. The enzyme lost 30% of its activity within 25 days of storage at 4°C. The optimum temperatures of activity were 40°C and 45°C for the transferase and bio‐synthetic activities, respectively.
Somatic embryogenesis of Cyclamen persicum in liquid medium
Authors: Kreuger Marc , Postma Erik , Brouwer Yvon , van Holst Gerrit‐Jan
Published: May 2006

A method is described for the production of somatic embryos of Cyclamen persicum Mill. in liquid medium. Five steps are involved; initiation of embryogenic cell lines, proliferation of pro‐embryogenic masses (PEMs) on auxin‐containing medium, development of somatic embryos on hormone‐free medium with high osmolarity, germination and subsequent plantlet formation. Cell lines were initiated by culturing the explant, the seedling tuber, directly in liquid medium. Three parameters were important for obtaining embryogenic cell lines; explant density, hormone concentrations and subculture regime. The rate of uptake of the hormones 2,4‐dichlorophenoxyacetic acid (2,4‐D) and kinetin influenced the formation of PEMs. Highly embryogenic cell lines were obtained only when PEMs had formed within 5–7 weeks. PEMs were proliferated for at least 24 months and could be isolated from each subculture for the production of somatic embryos. A high sucrose content (175 mM) in the development medium without hormones ensured efficient embryo development from PEMs. A subsequent subculture in low sucrose concentration (58 mM) induced the formation of a tuber, thus promoting germination. Arabinogalactan‐proteins (AGPs) from carrot seeds and AGPs bound by the monoclonal antibody ZUM 18 increased the number of PEMs in a culture, showing that the activity of AGPs is not species specific.
Stomatal and photosynthetic responses to shade in sorghum, soybean and eastern gamagrass
Authors: Fay Philip A. , Knapp Alan K.
Published: May 2006

We studied photosynthetic and stomatal responses of grain sorghum (Sorghum bicolor [L.] Moench cv. Pioneer 8500), soybean (Glycine max L. cv. Flyer) and eastern gamagrass (Tripsacum dactyloides L.) during experimental sun and shade periods simulating summer cloud cover. Leaf gas exchange measurements of field plants showed that short‐term (5 min) shading of leaves to 300–400 μmol m−2 s−1 photosynthetic photon flux density reduced photosynthesis, leaf temperature, stomatal conductance, transpiration and water use efficiency and increased intercellular CO2 partial pressure. In all species, photosynthetic recovery was delayed when leaves were reilluminated, apparently by stomatal closure. The strongest stomatal response was in soybean. Photosynthetic recovery was studied further with soybeans grown indoors (maximum photosynthetic photon flux density 1 200 μmol m−2 s−1). Plants grown indoors had responses to shade similar to those of field plants, except for brief nonstomatal limitation immediately after reillumination. These responses indicated the importance of the light environment during leaf development on assimilation responses to variable light, and suggested different limitations on carbon assimilation in different parts of the soybean canopy. Photosynthetic oxygen evolution recovered immediately upon reillumination, indicating that the light reactions did not limit soybean photosynthetic recovery. While shade periods caused stomatal closure and reduced carbon gain and water loss in all species, the consequences for carbon gain/water loss were greatest in soybean. The occurrence of stomatal closure in all three species may arise from their shared phenologies and herbaceous growth forms.
Gloeobacter violaceus— investigation of an unusual photosynthetic apparatus. Absence of the long wavelength emission of photosystem I in 77 K fluorescence spectra
Authors: Koenig Friederike , Schmidt Martin
Published: May 2006

The deeply purple cyanobacterium Gloeobacter violaceus is subject of this investigation. It does not contain thylakoids, and the photosynthetic apparatus is located in the only membrane of the cell, the plasma membrane. Upon excitation with blue light, the 77 K fluorescence emission spectra of neither intact cells (excited with 427 nm) nor of the isolated plasma membrane (excited with 430 nm), show the expected long wavelength photosystem I emission characteristic for low energy chlorophylls. Maximal fluorescence emission was observed at 688 nm. independent on the excitation wavelength, 427 (430) nm blue light, exciting mainly chlorophyll, or 550 nm green light, exciting mainly phycoerythin. The ratio of P700 to chlorophyll was 175. O2‐evolution was 160 μmol mg‐1 chlorophyll h‐1 in saturating white light; the compensation point was reached at 6 μmol m2 s‐1 in cultures grown at 25 μmol m2 s‐1. Dark O2 uptake was 50 μmol mg‐1 chlorophyll h‐1. During adaptation to increasing white light intensities Gloeobacter reduces the amount of phycocyanin and chlorophyll per cell and strongly increases the concentration of carotenoids relative to chlorophyll. The carotenoid concentration per cell increases with increasing light intensity. Apparently, part of the carotenoids is not located in the plasma membrane.
Partial purification and properties of a 36‐kDa 1‐aminocyclopropane‐1‐carboxylate N‐malonyltransferase from mung bean
Authors: Benichou Mohamed , Martinez‐Reina Gracia , Romojaro Felix , Pech Jean‐Claude , Latché Alain
Published: May 2006

A 36‐kDa 1‐aminocyclopropane‐1‐carboxylate (ACC) N‐malonyltransferase, which converts the ethylene precursor ACC into the conjugated derivative malonyl‐ACC (MACC), has been isolated from etiolated mung bean (Vigna radiata) hypocotyls, and partially purified in a four‐step procedure. The enzyme is stimulated about 7‐fold by 100 mM K+ salts or 0.5 mM Co2+ salts, and is inhibited competitively by D‐phenylalanine (Ki= 1.3 mM) and non competitively by CoASH (0.3 mM). Beside malonyl‐CoA, it is capable to use succinyl‐CoA as an acyl donor. The 36‐kDa enzyme described here exhibits a lower optimum temperature (40°C) and a 7‐ or 3‐fold lower apparent Km for ACC (68 μM) and malonyl‐CoA (74 μM), respectively, when compared with its 55 kDa isoform already isolated from the same plant material. This data support the idea that several isoforms of ACC N‐malonyltransferase exist in plants. These isoforms may play a differential role in regulating the availability of ACC, and consequently the rate of ethylene production, as well as detoxifying cells from D‐amino acids.
Water stress‐induced alterations in the proline metabolism of drought‐susceptible and ‐tolerant cassava (Manihot esculenta) cultivars
Authors: Sundaresan S. , Sudhakaran P. R.
Published: May 2006

The free proline levels and activities of ornithine aminotransferase (EC 2.6.1.13) and proline oxidase (EC 1.5.2.2), two of the enzymes involved in proline metabolism were studied during the induction of water stress in a drought susceptible (M‐4) and a drought tolerant (S‐1315) cultivar of cassava (Manihot esculenta Crantz). Water stress induced by polyethylene glycol (MW 6000, osmotic potential — 1.65 MPa) caused a ca 25‐fold increase in proline in young excised leaves of the susceptible cultivar (M‐4) while the increase was about 9‐fold in the tolerant cultivar (S‐1315). The activity of ornithine aminotransferase (OAT), a key enzyme involved in the biosynthesis of proline, was found to increase 3‐fold in water stressed leaves of M‐4 and about 2‐fold in those of S‐1315. The activity of proline oxidase, which is involved in the degradation of proline to pyrroline‐5‐carboxylate, was reduced by 50% in M‐4 and nearly 25% in S‐1315 on water stress. Comparison of the kinetic properties of OAT showed that the enzyme from water‐stressed leaves is more stable to heat inactivation compared to that of control. These results indicate that during water stress there are alterations in the metabolism of proline in cassava, and the extent of alteration varies between drought‐susceptible and ‐tolerant cultivars.
Phosphatidylcholine biosynthesis and cell growth in Apium graveolens
Authors: Parkin Edward T. , Goad L. John , Rolph Carole E.
Published: May 2006

Xenobiotics directed against sterol biosynthesis have proved to be useful tools in the determination of which sterol molecules are necessary for successful plant cell growth. However, the exact mode of action by which sterols are able to trigger cell growth remains to be elucidated. Previous studies using the triazole paclobutrazol, demonstrated that in Apium graveolens (cv. New Dwarf White) suspension cultures, sterol and phosphatidylcholine biosynthesis are co‐ordinately regulated (C. E. Rolph and L. J. Goad 1991, Physiol. Plant. 83: 605–610). The studies presented herein, were designed to investigate the possible role of phosphatidylcholine in the stimulation of plant cell growth.

Sterol biosynthesis, and hence cell growth, was inhibited by the use of the azole xenobiotic miconazole. Treatment of the cultures with miconazole lead to compositional changes in the free sterol content of the cells. For example, 30 μM miconazole treatment led to a reduction in the stigmasterol/sitosterol ratios from 1.53 to 1.24. In contrast, the phospholipid content of the cells remained relatively unchanged with phosphatidylcholine accounting for approximately 25% of the total phospholipids present in both control and miconazole‐treated cells. The cytostatic effect of miconazole could be partially counteracted by supplementation of the growth medium with the phytosterol stigmasterol and/or the unsaturated fatty acids oleate and linoleate.

The activity of CTP:cholinephosphate cytidylyltransferase (EC 2.7.7.15), a rate‐limiting enzyme in phosphatidylcholine biosynthesis, was significantly reduced in cells whose growth had been arrested by miconazole treatment. In miconazole‐treated cultures whose growth had been partially restored by supplementation with either specific sterols or unesterified fatty acids, the activity of this key enzyme was increased. In the case of stigmasterol, oleate and linoleate supplementation, the microsomal activity was found to be similar to that exhibited by control cultures.

From these studies, it may be concluded that certain phytosterols and unsaturated fatty acids play key roles with respect to phosphatidylcholine biosynthesis and that phosphatidylcholine biosynthesis via the CDP‐base pathway is an important pre‐ and/or co‐requisite for successful culture growth.
Characterization and partial purification of indole‐3‐butyric acid synthetase from maize (Zea mays)
Authors: Ludwig‐Müller Jutta , Hilgenberg Willy
Published: May 2006

In previous work it has been shown that the route from indoleacetic acid (IAA) to indolebutyric acid (IBA) is likely to be a two‐step process with an unknown intermediate designated ‘product X′. Our objective was to characterize and purify enzyme activities that are involved in these reactions. Indole‐3‐butyric acid synthetase was isolated and characterized from light‐grown maize seedlings (Zea mays L.), which were able to synthesize IBA from indole‐3‐acetic acid (IAA) with ATP and acetyl‐CoA as cofactors. The enzyme activity is most likely located on the membranes of the endoplasmic reticulum, as shown by means of aqueous two‐phase partitioning and sucrose density gradient centrifugation, with subsequent marker enzyme analysis. It was possible to solubilize the enzyme from the membranes with a detergent (CHAPS) and high concentrations of NaCl. The molecular mass of solubilized IBA synthetase was ca 31 kDa and its isoelectric point was at pH 4.8. The enzyme forming the reaction intermediate had a molecular mass of only 20 kDa and it seemed to be located on different membranes. Inhibition experiments with reducing agents and sulfhydryl reagents indicated that no sulfhydryl groups or disulfide bridges were present in the active centre of IBA synthetase. KCN inhibited the enzyme activity completely, and sodium azide by about 50%. Substrate analogs. such as 1‐IAA, 2,4‐dichlorophenoxyacetic acid, phenylacetic acid, and naphthaleneacetic acid, inhibited IBA formation to a high extent. Experiments with tunicamycin gave evidence that the enzyme is not a glycoprotein. These findings were confirmed by affinity chromatography with Concanavalin A. where the enzyme did not bind to the matrix. Further purification of the IBA synthetase on an ATP‐affinity column resulted in a more than 1 000‐fold purification compared to the microsomal membranes. IBA synthetase activity was also present in other plant families. Our results present further evidence that IBA is synthesized by a two‐step mechanism involving two different enzyme activities.