Plant Cell and Environment

Seventy one cultivars of sweet sorghum (Sorghum bicolor L.) were screened for aluminium (Al) tolerance by measuring relative root growth (RRG). Two contrasting cultivars, ROMA (Al-tolerant) and POTCHETSTRM (Al-sensitive), were selected to study shorter-term responses to Al stress. POTCHETSTRM had higher callose synthase activity, lower β-1,3-glucanase activity and more callose deposition in the root apices during Al treatment compared with ROMA. We monitored the expression of twelve genes involved in callose synthesis and degradation and found that one of these, SbGlu1 (Sb03g045630.1), which encodes a β-1,3-glucanase enzyme, best explained the contrasting deposition of callose in ROMA and POTCHETSTRM during Al treatment. Full length cDNAs of SbGlu1 was prepared from ROMA and POTCHETSTRM and expressed in Arabidopsis thaliana using the constitutive CaMV 35S promoter. Independent transgenic lines displayed significantly greater Al tolerance than wild-type plants and vector-only controls. This phenotype was associated with greater total β-1,3-glucanase activity, less Al accumulation and reduced callose deposition in the roots. These results suggest that callose production is not just an early indicator of Al stress in plants but likely to be part of the toxicity pathway that leads to the inhibition of root growth.

Plants sense light and gravity to orient their direction of growth. One common component in the early events of both phototropic and gravitropic signal transduction is activation of phospholipase C (PLC), which leads to an increase in inositol 1,4,5-triphosphate (InsP(3)) levels. The InsP(3) signal is terminated by hydrolysis of InsP(3) through inositolpolyphosphate-5-phosphatases (InsP 5-ptases). Arabidopsis plants expressing a heterologous InsP 5-ptase have low basal InsP(3) levels and exhibit reduced gravitropic and phototropic bending. Downstream effects of InsP(3)-mediated signalling are not understood. We used comparative transcript profiling to characterize gene expression changes in gravity- or light-stimulated Arabidopsis root apices that were manipulated in their InsP(3) metabolism either through inhibition of PLC activity or expression of InsP 5-ptase. We identified InsP(3)-dependent and InsP(3)-independent co-regulated gene sets in response to gravity or light stimulation. Inhibition of PLC activity in wild-type plants caused similar changes in transcript abundance in response to gravitropic and phototropic stimulation as in the transgenic lines. Therefore, we conclude that changes in gene expression in response to gravitropic and phototropic stimulation are mediated by two signal transduction pathways that vary in their dependence on changes in InsP(3).

This study was conducted to unravel a mechanism for the gravitropic curvature response in oat (Avena sativa) shoot pulvini. For this purpose, we examined the downward movement of starch-filled chloroplast gravisensors, differential changes in inositol 1,4,5-trisphosphate (IP(3)) levels, transport of indole-3-acetic acid (IAA) and gravitropic curvature. Upon gravistimulation, the ratio for IAA levels in lower halves versus those in upper halves (L/U) increased from 1.0 at 0 h and reached a maximum value of 1.45 at 8 h. When shoots were grown in the dark for 10 d, to deplete starch in the chloroplast, the gravity-induced L/U of IAA was reduced to 1.0. N-naphthylphthalamic acid (NPA) and 2,3,5-triiodobenzoic acid (TIBA), both auxin transport inhibitors, significantly reduced the amount of gravitropic curvature and gravity-induced lateral IAA transport, but did not reduce the gravity-induced late change in the L/U ratio of IP(3) levels. U73122, a specific phospholipase C (PLC) inhibitor, decreased gravity-induced curvature. Because U73122 reduced the ratio of L/U of IAA imposed by gravistimulation, it is clear that IAA transport is correlated with changes in IP(3) levels upon gravistimulation. These results indicate that gravistimulation-induced differential lateral IAA transport may result from the onset of graviperception in the chloroplast gravisensors coupled with gravity-induced asymmetric changes in IP(3) levels in oat shoot pulvini.

The activity of the photosynthetic carbon-fixing enzyme, ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), is partially inhibited by arsenite in the millimolar concentration range. However, micromolar arsenite can fully inhibit Rubisco in the presence of a potentiating monothiol such as cysteine, cysteamine, 2-mercaptoethanol or N-acetylcysteine, but not glutathione. Arsenite reacts specifically with the vicinal Cys172-Cys192 from the large subunit of Rubisco and with the monothiol to establish a ternary complex which is suggested to be a trithioarsenical. The stability of the complex is strongly dependent on the nature of the monothiol. Enzyme activity is fully recovered through the disassembly of the complex after eliminating arsenite and/or the thiol from the medium. The synergic combination of arsenite and a monothiol acts also in vivo stopping carbon dioxide fixation in illuminated cultures of Chlamydomonas reinhardtii. Again, this effect may be reverted by washing the cells. However, in vivo inhibition does not result from the blocking of Rubisco since mutant strains carrying Rubiscos with Cys172 and/or Cys192 substitutions (which are insensitive to arsenite in vitro) are also arrested. This suggests the existence of a specific sensor controlling carbon fixation that is even more sensitive than Rubisco to the arsenite-thiol synergism.

Cyclic electron flow around photosystem I (CEF1) is thought to augment chloroplast ATP production to meet metabolic needs. Very little is known about the induction and regulation of CEF1. We investigated the effects on CEF1 of antisense suppression of the Calvin-Benson enzymes glyceraldehyde-3-phosphate dehydrogenase (gapR), and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) small subunit (SSU), in tobacco (Nicotiana tabacum cv. Wisconsin 38). The gapR, but not ssuR, mutants showed substantial increases in CEF1, demonstrating that specific intermediates, rather than slowing of assimilation, induce CEF1. Both types of mutant showed increases in steady-state transthylakoid proton motive force (pmf) and subsequent activation of the photoprotective q(E) response. With gapR, the increased pmf was caused both by up-regulation of CEF1 and down-regulation of the ATP synthase. In ssuR, the increased pmf was attributed entirely to a decrease in ATP synthase activity, as previously seen in wild-type plants when CO₂ levels were decreased. Comparison of major stromal metabolites in gapR, ssuR and hcef1, a mutant with decreased fructose 1,6-bisphosphatase activity, showed that neither the ATP/ADP ratio, nor major Calvin-Benson cycle intermediates can directly account for the activation of CEF1, suggesting that chloroplast redox status or reactive oxygen species regulate CEF1.

Chloroplast protrusions (CPs) are often observed under environmental stresses, but their role has not been elucidated. The formation of CPs was observed in the leaf of rice plants treated with 75 mm NaCl for 14 d. Some CPs were almost separated from the main chloroplast body. In some CPs, inner membrane structures and crystalline inclusions were included. Similar structures surrounded by double membranes were observed in the cytoplasm and vacuole. Ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) was detected in CPs and the similar structures in the cytoplasm and vacuole. These results suggest that CP is one of the pathways of Rubisco exclusion from chloroplasts into the cytoplasm under salinity, and the exclusions could be transported to vacuole for their degradation.

Although ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) was discovered nearly 60 years ago, the associated chemical mechanism of the reaction is still incompletely understood. The catalytic cycle consists of four major steps: ribulose-1,5-bisphosphate binding, enolization, CO(2) or O(2) addition and hydration and cleavage of the intermediate. The use of individual rate constants for these elemental steps yields mathematical expressions for usual kinetic constants (k(cat) , K(m) ), CO(2) vs. O(2) specificity (S(c/o) ) as well as other chemical parameters such as the (12) C/(13) C isotope effect. That said, most of them are not simple and thus the interpretation of experimental, observed values of k(cat) , K(m) and S(c/o) may be more complicated than expected. That is, Rubisco effective catalysis depends on several kinetic parameters which are influenced by both the biological origin and the cellular medium (that can in turn vary with environmental conditions). In this brief review, we present the basic model of Rubisco kinetics and describe how subtle biochemical changes (which may have occurred along Evolution) can easily modify Rubisco catalysis.

D-ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyses the first step in photosynthetic carbon assimilation and represents the largest sink for nitrogen in plants. Improvement of its kinetic properties or the efficiency with which it is used in planta would benefit photosynthesis, nitrogen and water use efficiency, and yield. This paper presents a new non-radioactive microplate-based assay, which determines the product [3-phosphoglycerate (3-PGA)] in an enzymic cycle between glycerol-3-phosphate dehydrogenase and glycerol-3-phosphate oxidase. High sensitivity permits use of highly diluted extracts, and a short reaction time to avoid problems due to fall-off. Throughput was several hundreds of samples per person per day. Sensitivity and convenience compared favourably with radioisotopic assays, which were previously used to assay Rubisco. Its use is illustrated in three applications. (1) Maximal and initial activities and the K(m) for ribulose-1,5-bisphosphate were determined in raw extracts of leaves from several species. Similar values were obtained from those in the literature. (2) Diurnal changes were compared in rosettes of wild-type (WT) Arabidopsis and the starchless pgm mutant. Despite these dramatic differences in carbon metabolism, Rubisco activity and activation were similar in both genotypes. (3) A preliminary association mapping study was performed with 118 Arabidopsis accessions, using 183 markers that probably cover approximately 3-8% of the total genome. At a P-value < 0.005, two, two and no quantitative trait loci (QTL) were found for Rubisco maximal activity, initial activity and activation state, respectively. Inspection of the genomic regions that span these markers revealed these QTL involved genes not previously implicated in the regulation of Rubisco expression or activity.

During photosynthesis, triose-phosphates (trioseP) exported from the chloroplast to the cytosol are converted to sucrose via cytosolic fructose-1,6-bisphosphatase (cFBPase). Expression analysis in rice suggests that OscFBP1 plays a major role in the cytosolic conversion of trioseP to sucrose in leaves during the day. The isolated OscFBP1 mutants exhibited markedly decreased photosynthetic rates and severe growth retardation with reduced chlorophyll content, which results in plant death. Analysis of primary carbon metabolites revealed both significantly reduced levels of sucrose, glucose, fructose and starch in leaves of these mutants, and a high accumulation of sucrose to starch in leaves of rice plants. In the oscfbp1 mutants, products of glycolysis and the TCA cycle were significantly increased. A partitioning experiment of (14)C-labelled photoassimilates revealed altered carbon distributions including a slight increase in the insoluble fraction representing transitory starch, a significant decrease in the neutral fraction corresponding to soluble sugars and a high accumulation of phosphorylated intermediates and carboxylic acid fractions in the oscfbp1 mutants. These results indicate that the impaired synthesis of sucrose in rice cannot be sufficiently compensated for by the transitory starch-mediated pathways that have been found to facilitate plant growth in the equivalent Arabidopsis mutants.

A full-length FBPase cDNA has been isolated from Fragaria x ananassa (strawberry) corresponding to a novel putative chloroplastic FBPase but lacking the regulatory redox domain, a characteristic of the plastidial isoenzyme (cpFBPaseI). Another outstanding feature of this novel isoform, called cpFBPaseII, is the absence of the canonical active site. Enzymatic assays with cpFBPaseII evidenced clear Mg(2+)-dependent FBPase activity and a K(m) for fructose-1,6-bisphosphate (FBP) of 1.3 mM. Immunolocalization experiments and chloroplast isolation confirmed that the new isoenzyme is located in the stroma. Nevertheless, unlike cpFBPaseI, which is redox activated, cpFBPaseII did not increase its activity in the presence of either DTT or thioredoxin f (TRX f) and is resistant to H(2)O(2) inactivation. Additionally, the novel isoform was able to complement the growth deficiency of the yeast FBP1 deletion fed with a non-fermentable carbon source. Furthermore, orthologues are restricted to land plants, suggesting that cpFBPaseII is a novel and an intriguing chloroplastic FBPase that emerged late in the evolution of photosynthetic organisms, possibly because of a pressing need of land plants.

The role of cysteines 449 (Cys449) and 459 (Cys459) from the large subunit (LS) of ribulose 1-5-bisphosphate carboxylase/oxygenase (Rubisco) in the reduction-oxidation (redox) regulation of the enzyme was assessed by site-directed mutagenesis of these residues and chloroplast transformation of Chlamydomonas reinhardtii. In vitro studies indicated that mutations C449S, C459S or C449S/ C459S do not affect the activity and proteolytic susceptibility of the enzyme in the reduced state. However, when oxidized, the mutant enzymes differed from the wild type (WT), showing an increased resistance to inactivation and, in the case of the double mutant (DM), an altered structural conformation as reflected by the kinetics of proteolysis with subtilisin. The response of the DM strain to saline stress revealed that the absence of Cys449 and Cys459 intensifies Rubisco degradation and the covalent disulfide and non-disulfide polymerization of the enzyme in vivo. Saline stress also induced Rubisco translocation to a membrane (M) fraction that contained only covalently polymerized enzyme. Rubisco mobilization to this M fraction was enhanced also in the DM strain. Altogether, these results indicate that Cys449 and Cys459 participate in the modulation of the conformational changes promoted by oxidative modifications retarding processes related to the catabolism of the enzyme in vivo.

Photosynthesis depends on the diffusion of gaseous CO(2) inside the leaf spaces from the stomatal entry point to the mesophyll cell walls. Although most research considers only the vertical diffusion from stomata on upper and/or leaf lower surfaces, some of the gas will diffuse in the lateral (paradermal) direction. The importance of lateral CO(2) diffusion is reviewed, and the anatomical characteristics of leaves, including the variation of air space volume between species and conditions are discussed. The contribution of the air space conductance to the limitation of photosynthesis by the overall CO(2) diffusion pathway is usually ignored. However, the need to consider three-dimensional diffusion at the small scale of a few stomata is emphasized because stomata are discrete, and separated by 20-300 microm. At the large scale of 100s of micrometres, there may be barriers to CO(2) caused by the vascular tissue, particularly if there are bundle sheath extensions. The possible extent and controls on CO(2) lateral and vertical diffusion in different species and conditions are illustrated using chlorophyll a fluorescence imaging techniques. It is clear that there is a range of effective lateral permeabilities depending on the particular vascular patterns and cell arrangements, and that species cannot be simply divided into homobaric and heterobaric anatomies. Lateral diffusion in more permeable leaves can be sufficient to affect measurements of leaf gas exchange, particularly when fluxes are low, although its contribution to leaf photosynthesis in natural conditions needs clarification.

Leaf area expansion is affected by environmental conditions because of differences in cell number and/or cell size. Increases in the DNA content (ploidy) of a cell by endoreduplication are related to its size. The aim of this work was to determine how cell ploidy interacts with the regulation of cell size and with leaf area expansion. The approach used was to grow Arabidopsis thaliana plants performing increased or decreased rounds of endoreduplication under shading and water deficit. The shading and water deficit treatments reduced final leaf area and cell number; however, cell area was increased and decreased, respectively. These differences in cell size were unrelated to alterations of the endocycle, which was reduced by these treatments. The genetic modification of the extent of endoreduplication altered leaf growth responses to shading and water deficit. An increase in the extent of endoreduplication in a leaf rendered it more sensitive to the shade treatment but less sensitive to water deficit conditions. The link between the control of whole organ and individual cell expansion under different environmental conditions was demonstrated by the correlation between the plasticity of cell size and the changes in the duration of leaf expansion.

While there is currently intense effort to examine the (13)C signal of CO(2) evolved in the dark, less is known on the isotope composition of day-respired CO(2). This lack of knowledge stems from technical difficulties to measure the pure respiratory isotopic signal: day respiration is mixed up with photorespiration, and there is no obvious way to separate photosynthetic fractionation (pure c(i)/c(a) effect) from respiratory effect (production of CO(2) with a different delta(13)C value from that of net-fixed CO(2)) at the ecosystem level. Here, we took advantage of new simple equations, and applied them to sunflower canopies grown under low and high [CO(2)]. We show that whole mesocosm-respired CO(2) is slightly (13)C depleted in the light at the mesocosm level (by 0.2-0.8 per thousand), while it is slightly (13)C enriched in darkness (by 1.5-3.2 per thousand). The turnover of the respiratory carbon pool after labelling appears similar in the light and in the dark, and accordingly, a hierarchical clustering analysis shows a close correlation between the (13)C abundance in day- and night-evolved CO(2). We conclude that the carbon source for respiration is similar in the dark and in the light, but the metabolic pathways associated with CO(2) production may change, thereby explaining the different (12)C/(13)C respiratory fractionations in the light and in the dark.

In leaves, although it is accepted that CO(2) evolved by dark respiration after illumination is naturally (13) C-enriched compared to organic matter or substrate sucrose, much uncertainty remains on whether day respiration produces (13) C-depleted or (13) C-enriched CO(2). Here, we applied equations described previously for mesocosm CO(2) exchange to investigate the carbon isotope composition of CO(2) respired by autotrophic and heterotrophic tissues of Pelargonium × hortorum leaves, taking advantage of leaf variegation. Day-respired CO(2) was slightly (13) C-depleted compared to organic matter both under 21% O(2) and 2% O(2). Furthermore, most, if not all CO(2) molecules evolved in the light came from carbon atoms that had been fixed previously before the experiments, in both variegated and green leaves. We conclude that the usual definition of day respiratory fractionation, that assumes carbon fixed by current net photosynthesis is the respiratory substrate, is not valid in Pelargonium leaves under our conditions. In variegated leaves, total organic matter was slightly (13) C-depleted in white areas and so were most primary metabolites. This small isotopic difference between white and green areas probably came from the small contribution of photosynthetic CO(2) refixation and the specific nitrogen metabolism in white leaf areas.

Combined δ13C and δ18O analyses of leaf material were used to infer changes in photosynthetic capacity (Amax) and stomatal conductance (gl) in Fagus sylvatica and Picea abies trees growing under natural and controlled conditions. Correlation between gl and δ18O in leaf cellulose (δ18Ocel) allowed us to apply a semi-quantitative model to infer gl from δ18Ocel and also interpret variation in δ13C as reflecting variation in Amax. Extraction of leaf cellulose was necessary, because δ18O from leaf organic matter (δ18OLOM) and δ18Ocel was not reliably correlated. In juvenile trees, the model predicted elevated carbon dioxide (CO2) to reduce Amax in both species, whereas ozone (O3) only affected beech by reducing CO2 uptake via lowered gl. In adult trees, Amax declined with decreasing light level as gl was unchanged. O3 did not significantly affect isotopic signatures in leaves of adult trees, reflecting the higher O3 susceptibility of juvenile trees under controlled conditions. The isotopic analysis compared favourably to the performance of leaf gas exchange, underlining that the semi-quantitative model approach provides a robust way to gather time-integrated information on photosynthetic performance of trees under multi-faced ecological scenarios, in particular when information needed for quantitative modelling is only scarcely available.

We characterized differences in carbon isotopic content (delta(13)C) and sugar concentrations in phloem exudates from Eucalyptus globulus (Labill) plantations across a rainfall gradient in south-western Australia. Phloem sap delta(13)C and sugar concentrations varied with season and annual rainfall. Annual bole growth was negatively related to phloem sap delta(13)C during summer, suggesting a water limitation, yet was positively related in winter. We conclude that when water is abundant, variations in carboxylation rates become significant to overall growth. Concentrations of sucrose in phloem sap varied across sites by up to 600 mm, and raffinose by 300 mm. These compounds play significant roles in maintaining osmotic balance and facilitating carbon movement into the phloem, and their relative abundances contribute strongly to overall delta(13)C of phloem sap. Taken together, the delta(13)C and concentrations of specific sugars in phloem sap provide significant insights to functions supporting growth at the tree, site and landscape scale.

(13) C discrimination between atmosphere and bulk leaf matter (Δ(13) C(lb) ) is frequently used as a proxy for transpiration efficiency (TE). Nevertheless, its relevance is challenged due to: (1) potential deviations from the theoretical discrimination model, and (2) complex time integration and upscaling from leaf to whole plant. Six hybrid genotypes of Populus deltoides×nigra genotypes were grown in climate chambers and tested for whole-plant TE (i.e. accumulated biomass/water transpired). Net CO(2) assimilation rates (A) and stomatal conductance (g(s) ) were recorded in parallel to: (1) (13) C in leaf bulk material (δ(13) C(lb) ) and in soluble sugars (δ(13) C(ss) ) and (2) (18) O in leaf water and bulk leaf material. Genotypic means of δ(13) C(lb) and δ(13) C(ss) were tightly correlated. Discrimination between atmosphere and soluble sugars was correlated with daily intrinsic TE at leaf level (daily mean A/g(s) ), and with whole-plant TE. Finally, g(s) was positively correlated to (18) O enrichment of bulk matter or water of leaves at individual level, but not at genotype level. We conclude that Δ(13) C(lb) captures efficiently the genetic variability of whole-plant TE in poplar. Nevertheless, scaling from leaf level to whole-plant TE requires to take into account water losses and respiration independent of photosynthesis, which remain poorly documented.

Isotopic labelling experiments were conducted to assess relationships among (13)C of recently assimilated carbon (deltaC(A)), foliage respiration (deltaC(F)), soluble carbohydrate (deltaC(SC)), leaf waxes (deltaC(LW)) and bulk organic matter (deltaC(OM)). Slash pine, sweetgum and maize were grown under (13)C depleted CO(2) to label biomass and then placed under ambient conditions to monitor the loss of label. In pine and sweetgum, deltaC(F) of labelled plants (approximately -44 and -35 per thousand, respectively) rapidly approached control values but remained depleted by approximately 4-6 per thousand after 3-4 months. For these tree species, no or minimal label was lost from deltaC(SC), deltaC(LW) and deltaC(OM) during the observation periods. deltaC(F) and deltaC(SC) of labelled maize plants rapidly changed and were indistinguishable from controls after 1 month, while deltaC(LW) and deltaC(OM) more slowly approached control values and remained depleted by 2-6 per thousand. Changes in deltaC(F) in slash pine and sweetgum fit a two-pool exponential model, with the fast turnover metabolic pool (approximately 3-4 d half-life) constituting only 1-2% of the total. In maize, change in deltaC(F) fits a single pool model with a half-life of 6.4 d. The (13)C of foliage respiration and biochemical pools reflect temporally integrated values of deltaC(A), with change in isotopic composition dampened by the size of metabolic carbon reserves and turnover rates.

Little is known about the dynamics of concentrations and carbon isotope ratios of individual carbohydrates in leaves in response to climatic and physiological factors. Improved knowledge of the isotopic ratio in sugars will enhance our understanding of the tree ring isotope ratio and will help to decipher environmental conditions in retrospect more reliably. Carbohydrate samples from larch (Larix gmelinii) needles of two sites in the continuous permafrost zone of Siberia with differing growth conditions were analyzed with the Compound-Specific Isotope Analysis (CSIA). We compared concentrations and carbon isotope values (δ(13) C) of sucrose, fructose, glucose and pinitol combined with phenological data. The results for the variability of the needle carbohydrates show high dynamics with distinct seasonal characteristics between and within the studied years with a clear link to the climatic conditions, particularly vapor pressure deficit. Compound-specific differences in δ(13) C values as a response to climate were detected. The δ(13) C of pinitol, which contributes up to 50% of total soluble carbohydrates, was almost invariant during the whole growing season. Our study provides the first in depth characterization of compound-specific needle carbohydrate isotope variability, identifies involved mechanisms and shows the potential of such results for linking tree physiological responses to different climatic conditions. This article is protected by copyright. All rights reserved.

Water-use efficiency and stable isotope composition were studied in three tropical tree species. Seedlings of Tectona grandis, Swietenia macrophylla and Platymiscium pinnatum were grown at either high or low water supply, and with or without added fertilizer. These three species previously exhibited low, intermediate and high whole-plant water-use efficiency (TE) when grown at high water supply in unfertilized soil. Responses of TE to water and nutrient availability varied among species. The TE was calculated as experiment-long dry matter production divided by cumulative water use. Species-specific offsets were observed in relationships between TE and whole-plant (13)C discrimination (Delta(13)C(p)). These offsets could be attributed to a breakdown in the relationship between Delta(13)C(p) and the ratio of intercellular to ambient CO(2) partial pressures (c(i)/c(a)) in P. pinnatum, and to variation among species in the leaf-to-air vapour pressure difference (v). Thus, a plot of v.TE against c(i)/c(a) showed a general relationship among species. Relationships between delta(18)O of stem dry matter and stomatal conductance ranged from strongly negative for S. macrophylla to no relationship for T. grandis. Results suggest inter-specific variation among tropical tree species in relationships between stable isotope ratios (delta(13)C and delta(18)O) and the gas exchange processes thought to affect them.

In many tree species, physiological adaptations to drought include the accumulation of osmotically active substances and/or the presence of particular compatible solutes, among them cyclitols. Recently, the cyclitol quercitol was identified in species of Eucalyptus , a diverse genus whose speciation is probably driven by adaptation to water availability. We subjected seedlings of 13 Eucalyptus species from different ecosystems (‘mesic’ and ‘xeric’) and different sub‐generic taxonomic groups to 10 weeks of water deficit (WD) treatment. Pre‐dawn water potentials (ψ pdwn ) and relative water content (RWC) were determined in shoots, and total osmolality, soluble low‐molecular‐weight carbohydrates and cyclitols were measured in leaves and roots. Responses to water deficit followed two distinct patterns: Eucalyptus species from ‘mesic’ environments adjusted concentrations of sucrose (through increased levels of sucrose and decreases in RWC) in response to water deficit, whereas ‘xeric’ species increased concentrations of quercitol (through reductions in RWC). In root tissues, only species from xeric environments contained high levels of quercitol and mannitol, increasing under WD conditions. We suggest that the former (mesic) strategy may be beneficial to respond to short‐lasting drought conditions, because sucrose is easily metabolized, whereas the latter (xeric) strategy may relate to an effective acclimation to longer‐lasting drought. These physiological response groups are also related to taxonomic groups within the genus.

A deeper understanding of the drought response and genetic improvement of the cultivated crops for better tolerance requires attention because of the complexity of the drought response syndrome and the loss of genetic diversity during domestication. We initially screened about 200 wild emmer wheat genotypes and then focused on 26 of these lines, which led to the selection of two genotypes with contrasting responses to water deficiency. Six subtractive cDNA libraries were constructed, and over 13 000 expressed sequence tags (ESTs) were sequenced using leaf and root tissues of wild emmer wheat genotypes TR39477 (tolerant) and TTD-22 (sensitive), and modern wheat variety Kiziltan drought stressed for 7 d. Clustering and assembly of ESTs resulted in 2376 unique sequences (1159 without hypothetical proteins and no hits), 75% of which were represented only once. At this level of EST sampling, each tissue shared a very low percentage of transcripts (13-26%). The data obtained indicated that the genotypes shared common elements of drought stress as well as distinctly differential expression patterns that might be illustrative of their contrasting ability to tolerate water deficiencies. The new EST data generated here provide a highly diverse and rich source for gene discovery in wheat and other grasses.

We report diurnal variations in (13)C discrimination ((13)Delta) of Picea sitchensis (Bong.) Carr. branches measured in the field using a branch chamber technique. The observations were compared to predicted (13)Delta based on concurrent measurements of branch gas exchange. Observed (13)Delta values were described well by the classical model of (13)Delta including isotope effects during photorespiration, day respiration and CO(2) transfer through a series of resistances to the sites of carboxylation. A simplified linear of model (13)Delta did not capture the observed diurnal variability. At dawn and dusk, we measured very high (13)Delta values that were not predicted by either of the said models. Exploring the sensitivity of (13)Delta to possible respiratory isotope effects, we conclude that isotopic disequilibria between the gross fluxes of photosynthesis and day respiration can explain the high observed (13)Delta values during net photosynthetic gas exchange. Based on the classical model, a revised formulation incorporating an isotopically distinct substrate for day respiration was able to account well for the high observed dawn and dusk (13)Delta values.