Showing papers in "Plant Cell and Environment in 2006"

Can improvement in photosynthesis increase crop yields

Abstract: The yield potential ( Y p ) of a grain crop is the seed mass per unit ground area obtained under optimum growing conditions without weeds, pests and diseases. It is determined by the product of the available light energy and by the genetically determined properties: efficiency of light capture ( e i ), the efficiency of conversion of the intercepted light into biomass ( e c ) and the proportion of biomass partitioned into grain ( h ). Plant breeding brings h and e i close to their theoretical maxima, leaving e c , primarily determined by photosynthesis, as the only remaining major prospect for improving Y p . Leaf photosynthetic rate, however, is poorly correlated with yield when different genotypes of a crop species are compared. This led to the viewpoint that improvement of leaf photosynthesis has little value for improving Y p . By contrast, the many recent experiments that compare the growth of a genotype in current and future projected elevated [CO 2 ] environments show that increase in leaf photosynthesis is closely associated with similar increases in yield. Are there opportunities to achieve similar increases by genetic manipulation? Six potential routes of increasing e c by improving photosynthetic efficiency were explored, ranging from altered canopy architecture to improved regeneration of the acceptor molecule for CO 2 . Collectively, these changes could improve e c and, therefore, Y p by c . 50%. Because some changes could be achieved by transgenic technology, the time of the development of commercial cultivars could be considerably less than by conventional breeding and potentially, within 10‐15 years.

The hydraulic limitation hypothesis revisited

Abstract: We proposed the hydraulic limitation hypothesis (HLH) as a mechanism to explain universal patterns in tree height, and tree and stand biomass growth: height growth slows down as trees grow taller, maximum height is lower for trees of the same species on resource-poor sites and annual wood production declines after canopy closure for even-aged forests. Our review of 51 studies that measured one or more of the components necessary for testing the hypothesis showed that taller trees differ physiologically from shorter, younger trees. Stomatal conductance to water vapour ( g s ), photosynthesis ( A ) and leaf-specific hydraulic conductance ( K L ) are often, but not always, lower in taller trees. Additionally, leaf mass per area is often greater in taller trees, and leaf area:sapwood area ratio changes with tree height. We conclude that hydraulic limitation of gas exchange with increasing tree size is common, but not universal. Where hydraulic limitations to A do occur, no evidence supports the original expectation that hydraulic limitation of carbon assimilation is sufficient to explain observed declines in wood production. Any limit to height or height growth does not appear to be related to the so-called age-related decline in wood production of forests after canopy closure. Future work on this problem should explicitly link leaf or canopy gas exchange with tree and stand growth, and consider a more fundamental assumption: whether tree biomass growth is limited by carbon availability.

Cadmium effect on oxidative metabolism of pea (Pisum sativum L.) roots. Imaging of reactive oxygen species and nitric oxide accumulation in vivo

Abstract: Growth of pea (Pisum sativum L.) plants with 50 microM CdCl2 for 15 d produced a reduction in the number and length of lateral roots, and changes in structure of the principal roots affecting the xylem vessels. Cadmium induced a reduction in glutathione (GSH) and ascorbate (ASC) contents, and catalase (CAT), GSH reductase (GR) and guaiacol peroxidase (GPX) activities. CuZn-superoxide dismutase (SOD) activity was also diminished by the Cd treatment, although Mn-SOD was slightly increased. CAT and CuZn-SOD were down-regulated at transcriptional level, while Mn-SOD, Fe-SOD and GR were up-regulated. Analysis of reactive oxygen species (ROS) and nitric oxide (NO) levels by fluorescence and confocal laser microscopy (CLM) showed an over-accumulation of O2*- and H2O2, and a reduction in the NO content in lateral and principal roots. ROS overproduction was dependent on changes in intracellular Ca+2 content, and peroxidases and NADPH oxidases were involved. Cadmium also produced an increase in salicylic acid (SA), jasmonic acid (JA) and ethylene (ET) contents. The rise of ET and ROS, and the NO decrease are in accordance with senescence processes induced by Cd, and the increase of JA and SA could regulate the cellular response to cope with damages imposed by cadmium.

Metabolic signalling in defence and stress: the central roles of soluble redox couples

Abstract: Plant growth and development are driven by electron transfer reactions. Modifications of redox components are both monitored and induced by cells, and are integral to responses to environmental change. Key redox compounds in the soluble phase of the cell are NAD, NADP, glutathione and ascorbate--all of which interact strongly with reactive oxygen. This review takes an integrated view of the NAD(P)-glutathione-ascorbate network. These compounds are considered not as one-dimensional 'reductants' or 'antioxidants' but as redox couples that can act together to condition cellular redox tone or that can act independently to transmit specific information that tunes signalling pathways. Emphasis is placed on recent developments highlighting the complexity of redox-dependent defence reactions, and the importance of interactions between the reduction state of soluble redox couples and their concentration in mediating dynamic signalling in response to stress. Signalling roles are assessed within the context of interactions with reactive oxygen, phytohormones and calcium, and the biochemical reactions through which redox couples could be sensed are discussed.

Drought stress response in wheat: physiological and molecular analysis of resistant and sensitive genotypes

Abstract: Water deficit is a severe environmental stress and the major constraint on plant productivity with an evident effect on plant growth. The aim of this work was to study Triticum and Aegilops seedlings differing in their response to drought stress at the physiological and molecular levels. The identification of resistant and sensitive genotypes was firstly based on the relative water content (RWC) measurement. Further characterization of genotypes contrasting in their response to water stress was performed at the physiological level by determination of RWC, water loss rate (WLR) and free proline content after different hours of dehydration. Modification in the expression level of five dehydrin (DHN) genes was also analysed by reverse transcription-polymerase chain reaction (RT-PCR). Five cDNAs coding for different DHNs were identified and characterized. These genes are not expressed in the well-watered plants, but only in the stressed plants. Four of these cDNAs are related to novel DHN sequences. The results obtained clearly indicate a relation between the expression of these genes and tissue water content. In particular, in the resistant genotypes the expression of DHN genes is initiated even though tissue hydration levels are still high, indicating also in wheat the involvement of these proteins in water retention.

Carotenoid accumulation and function in seeds and non-green tissues.

Abstract: Carotenoids are plant pigments that function as antioxidants, hormone precursors, colourants and essential components of the photosynthetic apparatus. Carotenoids accumulate in nearly all types of plastids, not just the chloroplast, and are thus found in most plant organs and tissues, albeit at trace levels in some tissues. In this review we summarise the current knowledge of the carotenoid content of non-green plastids and discuss what is known about the regulation of their biosynthesis in roots, fruits, flowers, tubers and seeds. The emphasis is on food crops as carotenoids are essential components of human diets, primarily as some are precursors of vitamin A. The low carotenoid content of many staple foods, such as cereals, can exacerbate dietary deficiencies. The World Health Organisation has estimated that more than 100 million children are vitamin A-deficient and up to 500,000 of these children become blind each year. Many of these children die within 12 months of going blind. Thus, understanding the regulation of carotenoid accumulation in food crops, especially tubers and cereals, should facilitate improvements to nutritional value with potentially significant health benefits.

Induction of systemic resistance in tomato by N-acyl-L-homoserine lactone-producing rhizosphere bacteria.

Abstract: N-acyl-L-homoserine lactone (AHL) signal molecules are utilized by Gram-negative bacteria to monitor their population density (quorum sensing) and to regulate gene expression in a density-dependent manner. We show that Serratia liquefaciens MG1 and Pseudomonas putida IsoF colonize tomato roots, produce AHL in the rhizosphere and increase systemic resistance of tomato plants against the fungal leaf pathogen, Alternaria alternata. The AHL-negative mutant S. liquefaciens MG44 was less effective in reducing symptoms and A. alternata growth as compared to the wild type. Salicylic acid (SA) levels were increased in leaves when AHL-producing bacteria colonized the rhizosphere. No effects were observed when isogenic AHL-negative mutant derivatives were used in these experiments. Furthermore, macroarray and Northern blot analysis revealed that AHL molecules systemically induce SA- and ethylene-dependent defence genes (i.e. PR1a, 26 kDa acidic and 30 kDa basic chitinase). Together, these data support the view that AHL molecules play a role in the biocontrol activity of rhizobacteria through the induction of systemic resistance to pathogens.

Silicon deposition in the root reduces sodium uptake in rice (Oryza sativa L.) seedlings by reducing bypass flow.

Abstract: Sodium chloride reduces the growth of rice seedlings, which accumulate excessive concentrations of sodium and chloride ions in their leaves. In this paper, we describe how silicon decreases transpirational bypass flow and ion concentrations in the xylem sap in rice (Oryza sativa L.) seedlings growing under NaCl stress. Salt (50 mM NaCl) reduced the growth of shoots and roots: adding silicate (3 mM) to the saline culture solution improved the growth of the shoots, but not roots. The improvement of shoot growth in the presence of silicate was correlated with reduced sodium concentration in the shoot. The net transport rate of Na from the root to shoot (expressed per unit of root mass) was also decreased by added silicate. There was, however, no effect of silicate on the net transport of potassium. Furthermore, in salt-stressed plants, silicate did not decrease the transpiration, and even increased it in seedlings pre-treated with silicate for 7 d prior to salt treatment, indicating that the reduction of sodium uptake by silicate was not simply through a reduction in volume flow from root to shoot. Experiments using trisodium-8-hydroxy-1,3,6-pyrenetrisulphonic acid (PTS), an apoplastic tracer, showed that silicate dramatically decreased transpirational bypass flow in rice (from about 4.2 to 0.8%), while the apparent sodium concentration in the xylem, which was estimated indirectly from the flux data, decreased from 6.2 to 2.8 mM. Direct measurements of the concentration of sodium in xylem sap sampled using Philaenus spumarius confirmed that the apparent reduction was not a consequence of sodium recycling. X-ray microanalysis showed that silicon was deposited in the outer part of the root and in the endodermis, being more obvious in the latter than in the former. The results suggest that silicon deposition in the exodermis and endodermis reduced sodium uptake in rice (Oryza sativa L.) seedlings under NaCl stress through a reduction in apoplastic transport across the root.

On the induction of volatile organic compound emissions by plants as consequence of wounding or fluctuations of light and temperature

Abstract: Among the volatile organic compounds (VOCs) emitted by plants, some are characteristic of stress conditions, but their biosynthesis and the metabolic and environmental control over the emission are still unclear. We performed experiments to clarify whether (1) the emission following wounding can occur at distance from the wounding site, from VOC pools subjected to metabolic signals; and (2) the emission of biogenic VOCs generated by membrane damage (e.g. consequent to wounding or ozone exposure) can also be induced by exposure to high light and high temperature, recurrent in nature. In Phragmites australis, leaf cutting caused large and rapid bursts of acetaldehyde both at the cutting site and on parts of the cut leaf distant from the cutting site. This emission was preceded by a transient stomatal opening and did not occur in conditions preventing stomatal opening. This suggests the presence of a large pool of leaf acetaldehyde whose release is under stomatal control. VOCs other than isoprene, particularly acetaldehyde and (E)-2-hexenal, one of the C-6 compounds formed by the denaturation of membrane lipids, were released by leaves exposed to high temperature and high light. The high-temperature treatment (45 degrees C) also caused a rapid stimulation and then a decay of isoprene emission in Phragmites leaves. Isoprene recovered to the original emission level after suspending the high-temperature treatment, suggesting a temporary deficit of photosynthetically formed substrate under high temperature. Emission of C-6 compounds was slowly induced by high temperature, and remained high, indicating that membrane denaturation occurs also after suspending the high-temperature treatment. Conversely, the emission of C-6 compounds was limited to the high-light episode in Phragmites. This suggests that a membrane denaturation may also occur in conditions that do not damage other important plant processes such as the photochemistry of photosynthesis of photoinhibition-insensitive plants. In the photoinhibition-sensitive Arabidopsis thaliana mutant NPQ1, a large but transient emission of (E)-2-hexenal was also observed a few minutes after the high-light treatment, indicating extensive damage to the membranes. However, (E)-2-hexenal emission was not observed in Arabidopsis plants fumigated with isoprene during the high-light treatment. This confirms that isoprene can effectively protect cellular membranes from denaturation. Our study indicates that large, though often transient, VOC emissions by plants occur in nature. In particular, we demonstrate that VOCs can be released by much larger tissues than those wounded and that even fluctuations of light and temperature regularly observed in nature can induce their emissions. This knowledge adds information that is useful for the parameterization of the emissions and for the estimate of biogenic VOC load in the atmosphere.

Managing sulphur metabolism in plants.

Abstract: Resolution and analysis of genes encoding components of the pathways of primary sulphur assimilation have provided the potential to elucidate how sulphur is managed by plants. Individual roles for members of gene families and regulatory mechanisms operating at gene, cellular and whole plant levels have been recognized. Sulphur is taken up and transported around the plant principally as sulphate, catalysed for the most part by a single gene family of highly regulated transporters. Additional regulation occurs in the pathway of reduction of sulphate to sulphide and its incorporation into cysteine, which occurs principally within the plastid. Cellular and whole-plant regulation of uptake, and the assimilatory pathway attempt to balance supply with demand for growth and include mechanisms for re-mobilization and redistribution of sulphur. Furthermore, optimization of sulphur assimilation requires coordination with carbon and nitrogen pathways, and multiple processes have been proposed to contribute to this balance. Present studies on cis and trans elements are focusing on transcriptional regulation, but this regulation still needs to be linked to apparent metabolite sensing. Whilst the components of the assimilatory pathways have been resolved after many years of controversy, uncertainties remain concerning roles of individual genes in gene families, their sub-cellular localization and their significance in balancing sulphur flux to sulphur demand of the plant for growth under variable environmental conditions.

Metabolic consequences of susceptibility and resistance (race‐specific and broad‐spectrum) in barley leaves challenged with powdery mildew

Abstract: In a compatible interaction biotrophic fungi often lower the yield of their hosts by reducing photosynthesis and altering the fluxes of carbon within the infected leaf. In contrast, comparatively little is known about the metabolic consequences of activating resistance responses. In this study we investigated the hypothesis that the activation of both race-specific (Mla12) and broad-spectrum (mlo) resistance pathways in barley leaves infected with Blumeria graminis represents a cost to the plant in terms of carbon production and utilization. We have shown, using quantitative imaging of chlorophyll fluorescence, that during a susceptible interaction, photosynthesis was progressively reduced both in cells directly below fungal colonies and in adjacent cells when compared with uninoculated leaves. The lower rate of photosynthesis was associated with an increase in invertase activity, an accumulation of hexoses and a down-regulation of photosynthetic gene expression. During both Mla12- and mlo-mediated resistance, photosynthesis was also reduced, most severely inhibited in cells directly associated with attempted penetration of the fungus but also in surrounding cells. These cells displayed intense autofluorescence under ultraviolet illumination indicative of the accumulation of phenolic compounds and/or callose deposition. The depression in photosynthesis was not due only to cell death but also to an alteration in source-sink relations and carbon utilization. Apoplastic (cell wall-bound) invertase activity increased more rapidly and to a much greater extent than in infected susceptible leaves and was accompanied by an accumulation of hexoses that was localized to areas of the leaf actively exhibiting resistance responses. The accumulation of hexoses was accompanied by a down-regulation in the expression of Rubisco (rbcS) and chlorophyll a/b binding protein (cab) genes (although to a lesser extent than in a compatible interaction) and with an up-regulation in the expression of the pathogenesis-related protein 1 (PR-1). These results are consistent with a role for invertase in the generation of hexoses, which may supply energy for defence reactions and/or act as signals inducing defence gene expression.

Comparative transcriptome analysis of toxic metal responses in Arabidopsis thaliana and the Cd2+-hypertolerant facultative metallophyte Arabidopsis halleri

Abstract: Toxic effects of both essential and non-essential heavy metals are well documented in plants. Very little is known, however, about their modes of toxicity, about tolerance mechanisms and the signalling cascades involved in mediating transcriptional responses to toxic metal excess. We analysed transcriptome changes upon Cd2+ and Cu2+ exposure in roots of Arabidopsis thaliana and the Cd(2+)-hypertolerant metallophyte Arabidopsis halleri. Particularly, three categories of genes were identified with the help of this comparative approach: (1) common responses, which might indicate stable and functionally relevant changes conserved across plant species; (2) metallophyte-specific responses as well as transcripts differentially regulated between the two species, representing candidate genes for Cd2+ hypertolerance; and (3) those specifically responsive to Cd2+ and therefore indicative of toxicity mechanisms or potentially involved in signalling cascades. Our data define, for instance, Arabidopsis core responses to Cd2+ and Cu2+. In addition, they suggest that Cd2+ exposure very rapidly results in apparent Zn deficiency, and they show the existence of highly specific Cd2+ responses and distinct signalling cascades. Array results were independently confirmed by real-time quantitative PCR, thereby further validating cross-species transcriptome analysis with oligonucleotide microarrays.

Over‐expression of different aldehyde dehydrogenase genes in Arabidopsis thaliana confers tolerance to abiotic stress and protects plants against lipid peroxidation and oxidative stress

Abstract: Aldehyde dehydrogenases (ALDHs) play a major role in the detoxification processes of aldehydes generated in plants when exposed to abiotic stress. In previous studies, we have shown that the Arabidopsis thaliana ALDH3I1 gene is transcriptionally activated by abiotic stress, and over-expression of the ALDH3I1 gene confers stress tolerance in transgenic plants. The A. thaliana genome contains 14 ALDH genes expressed in different sub-cellular compartments and are presumably involved in different reactions. The purpose of this study was to compare the potential of a cytoplasmic and a chloroplastic stress-inducible ALDH in conferring stress tolerance under different conditions. We demonstrated that constitutive or stress-inducible expression of both the chloroplastic ALDH3I1 and the cytoplasmic ALDH7B4 confers tolerance to osmotic and oxidative stress. Stress tolerance in transgenic plants is accompanied by a reduction of H2O2 and malondialdehyde (MDA) derived from cellular lipid peroxidation. Involvement of ALDHs in stress tolerance was corroborated by the analysis of ALDH3I1 and ALDH7B4 T-DNA knockout (KO) mutants. Both mutant lines exhibited higher sensitivity to dehydration and salt than wild-type (WT) plants. The results indicate that ALDH3I1 and ALDH7B4 not only function as aldehyde-detoxifying enzymes, but also as efficient reactive oxygen species (ROS) scavengers and lipid peroxidation-inhibiting enzymes. The potential of ALDHs to interfere with H2O2 was also shown for recombinant bacterial proteins.

Cadmium affects tobacco cells by a series of three waves of reactive oxygen species that contribute to cytotoxicity.

Abstract: Cadmium is suspected to exert its toxic action on cells through oxidative damage. However, the transition metal is unable to directly generate reactive oxygen species (ROS) via redox reactions with molecular oxygen in a biological environment. Here, we show that bright yellow-2 (BY-2) tobacco cells exposed to millimolar concentrations of CdCl(2) developed cell death within 2-3 h. The death process was preceded by two successive waves of ROS differing in their nature and subcellular localization. Firstly, these consisted in the transient NADPH oxidase-dependent accumulation of H(2)O(2) followed by the accumulation of O(2) (-*) in mitochondria. A third wave of ROS consisting in fatty acid hydroperoxide accumulation was concomitant with cell death. Accumulation of H(2)O(2) was preceded by an increase in cytosolic free calcium concentration originating from internal pools that was essential to activate the NADPH oxidase. The cell line gp3, impaired in NADPH oxidase activity, and that was unable to accumulate H(2)O(2) in response to Cd(2+), was nevertheless poisoned by the metal. Therefore, this first wave of ROS was not sufficient to trigger all the cadmium-dependent deleterious effects. However, we show that the accumulation of O(2) (-*) of mitochondrial origin and membrane peroxidation are key players in Cd(2+)-induced cell death.

MRI of long-distance water transport: a comparison of the phloem and xylem flow characteristics and dynamics in poplar, castor bean, tomato and tobacco

Abstract: We used dedicated magnetic resonance imaging (MRI) equipment and methods to study phloem and xylem transport in large potted plants. Quantitative flow profiles were obtained on a per-pixel basis, giving parameter maps of velocity, flow-conducting area and volume flow (flux). The diurnal xylem and phloem flow dynamics in poplar, castor bean, tomato and tobacco were compared. In poplar, clear diurnal differences in phloem flow profile were found, but phloem flux remained constant. In tomato, only small diurnal differences in flow profile were observed. In castor bean and tobacco, phloem flow remained unchanged. In all plants, xylem flow profiles showed large diurnal variation. Decreases in xylem flux were accompanied by a decrease in velocity and flow-conducting area. The diurnal changes in flow-conducting area of phloem and xylem could not be explained by pressure-dependent elastic changes in conduit diameter. The phloem to xylem flux ratio reflects what fraction of xylem water is used for phloem transport (Munch's counterflow). This ratio was large at night for poplar (0.19), castor bean (0.37) and tobacco (0.55), but low in tomato (0.04). The differences in phloem flow velocity between the four species, as well as within a diurnal cycle, were remarkably small (0.25-0.40 mm s(-1)). We hypothesize that upper and lower bounds for phloem flow velocity may exist: when phloem flow velocity is too high, parietal organelles may be stripped away from sieve tube walls; when sap flow is too slow or is highly variable, phloem-borne signalling could become unpredictable.

Senescence-specific regulation of catalases in Arabidopsis thaliana (L.) Heynh

Abstract: Oxygen free radicals are thought to play an essential role in senescence, especially those derived from peroxisomes. Therefore, the activities of different isoforms of the peroxisomal hydrogen peroxide (H2O2)-scavenging enzyme catalase (CAT) were analysed during senescence of Arabidopsis. CAT2 activity decreased with bolting time parallel with cytosolic ascorbate peroxidase 1 (APX1) activity before loss of chlorophyll could be measured. At the same time point, the H2O2 content increased. Subsequently, the stress-inducible CAT3 isoform was activated and APX1 activity was recovered, accompanied by a decline of the H2O2 content. In very late stages, low activities of the seed-specific CAT1 became detectable in leaves, but H2O2 increased again. Further analyses of CAT expression by promoter: beta-glucuronidase (GUS) fusions in transgenic plants revealed a vasculature-specific CAT3 expression, whereas CAT2 expression turned out to be specific for photosynthetic active tissues. CAT2 expression is down-regulated during leaf senescence, while CAT3 expression is induced with age and corresponds to an accumulation of H2O2 in the vascular bundles. CAT2 down-regulation on the transcriptional level appears as the initial step in creating the H2O2 peak during bolting time, while the decrease in APX1 activity might only be a secondary and amplifying effect.

Evidence that differential gene expression between the halophyte, Thellungiella halophila, and Arabidopsis thaliana is responsible for higher levels of the compatible osmolyte proline and tight control of Na+ uptake in T. halophila.

Abstract: Salt-sensitive glycophytes and salt-tolerant halophytes employ common mechanisms to cope with salinity, and it is hypothesized that differences in salt tolerance arise because of changes in the regulation of a basic set of salt tolerance genes. We explored the expression of genes involved in two key salt tolerance mechanisms in Arabidopsis thaliana and the halophytic A. thaliana relative model system (ARMS), Thellungiella halophila. Salt overly sensitive 1 (SOS1) is a plasma membrane Na + /H + antiporter that retrieves and loads Na + ions from and into the xylem. Shoot SOS1 transcript was more strongly induced by salt in T. halophila while root SOS1 was constitutively higher in unstressed T. halophila. This is consistent with a lower salt-induced rise in T. halophila xylem sap Na + concentration than in A. thaliana. Thellungiella halophila contained higher unstressed levels of the compatible osmolyte proline than A. thaliana, while under salt stress, T. halophila accumulated more proline mainly in shoots. Expression of the A. thaliana ortholog of proline dehydrogenase (PDH), involved in proline catabolism, was undetectable in T. halophila shoots. The PDH enzyme activity was lower and T. halophila seedlings were hypersensitive to exogenous proline, indicating repression of proline catabolism in T. halophila. Our results suggest that differential gene expression between glycophytes and halophytes contributes to the salt tolerance of halophytes.

Spatial coordination of aluminium uptake, production of reactive oxygen species, callose production and wall rigidification in maize roots.

Abstract: Aluminium (Al) toxicity associated with acid soils represents one of the biggest limitations to crop production worldwide. Although Al specifically inhibits the elongation of root cells, the exact mechanism by which this growth reduction occurs remains controversial. The aim of this study was to investigate the spatial and temporal dynamics of Al migration into roots of maize (Zea mays L.) and the production of the stress response compound callose. Using the Al-specific fluorescent probe morin, we demonstrate the gradual penetration of AI into roots. Al readily accumulates in the root's epidermal and outer cortical cell layers but does not readily penetrate into the inner cortex. After prolonged exposure times (12-24 h), Al had entered all areas of the root apex. The spatial and temporal accumulation of Al within the root is similarly matched by the production of the cell wall polymer callose, which is also highly localized to the epidermis and outer cortical region. Exposure to Al induced the rapid production of reactive oxygen species and induced a significant rigidification of the cell wall. Our results suggest that Al-induced root inhibition in maize occurs by rigidification of the epidermal layers.

Conifer defence against insects: microarray gene expression profiling of Sitka spruce (Picea sitchensis) induced by mechanical wounding or feeding by spruce budworms (Choristoneura occidentalis) or white pine weevils (Pissodes strobi ) reveals large-scale changes of the host transcriptome

Abstract: Conifers are resistant to attack from a large number of potential herbivores or pathogens. Previous molecular and biochemical characterization of selected conifer defence systems support a model of multigenic, constitutive and induced defences that act on invading insects via physical, chemical, biochemical or ecological (multitrophic) mecha- nisms. However, the genomic foundation of the complex defence and resistance mechanisms of conifers is largely unknown. As part of a genomics strategy to characterize inducible defences and possible resistance mechanisms of conifers against insect herbivory, we developed a cDNA microarray building upon a new spruce ( Picea spp.) expressed sequence tag resource. This first-generation spruce cDNA microarray contains 9720 cDNA elements representing c. 5500 unique genes. We used this array to monitor gene expression in Sitka spruce ( Picea sitchensis ) bark in response to herbivory by white pine weevils ( Pis- sodes strobi , Curculionidae) or wounding, and in young shoot tips in response to western spruce budworm ( Chori- stoneura occidentalis , Lepidopterae) feeding. Weevils are stem-boring insects that feed on phloem, while budworms are foliage feeding larvae that consume needles and young shoot tips. Both insect species and wounding treatment caused substantial changes of the host plant transcriptome detected in each case by differential gene expression of several thousand array elements at 1 or 2 d after the onset of treatment. Overall, there was considerable overlap among differentially expressed gene sets from these three stress treatments. Functional classification of the induced transcripts revealed genes with roles in general plant defence, octadecanoid and ethylene signalling, transport, secondary metabolism, and transcriptional regulation. Sev- eral genes involved in primary metabolic processes such as photosynthesis were down-regulated upon insect feeding or wounding, fitting with the concept of dynamic resource allocation in plant defence. Refined expression analysis using gene-specific primers and real-time PCR for selected transcripts was in agreement with microarray results for most genes tested. This study provides the first large-scale survey of insect-induced defence transcripts in a gymno- sperm and provides a platform for functional investigation of plant-insect interactions in spruce. Induction of spruce genes of octadecanoid and ethylene signalling, terpenoid biosynthesis, and phenolic secondary metabolism are dis- cussed in more detail.

Sugar and ABA response pathways and the control of gene expression

Abstract: Sugars are essential to plant growth and metabolism, both as energy source and as structural components. Sugar production and use are in part controlled at the level of gene expression by the sugars themselves. Responses to sugar are closely integrated with response pathways that indicate environmental conditions such as light and water availability. High sugar levels inhibit seedling development, repress photosynthetic gene expression and induce genes of storage metabolism such as those of starch biosynthesis. Genetic approaches have demonstrated the importance of abscisic acid (ABA) and the transcriptional regulator ABA-insensitive4 (ABI4) in sugar response pathways. Recent analysis of both photosynthetic and starch biosynthetic gene promoters suggest a direct role for ABI4 in their control. The increased understanding of the regulatory promoter elements controlling gene expression, in response to sugar and ABA, allows transcriptional networks to be understood at a molecular level.

Internal conductance does not scale with photosynthetic capacity: implications for carbon isotope discrimination and the economics of water and nitrogen use in photosynthesis

Abstract: Central paradigms of ecophysiology are that there are recognizable and even explicit and predictable patterns among species, genera, and life forms in the economics of water and nitrogen use in photosynthesis and in carbon isotope discrimination (delta). However most previous examinations have implicitly assumed an infinite internal conductance (gi) and/or that internal conductance scales with the biochemical capacity for photosynthesis. Examination of published data for 54 species and a detailed examination for three well-characterized species--Eucalyptus globulus, Pseudotsuga menziesii and Phaseolus vulgaris--show these assumptions to be incorrect. The reduction in concentration of CO2 between the substomatal cavity (Ci) and the site of carbon fixation (Cc) varies greatly among species. Photosynthesis does not scale perfectly with gi and there is a general trend for plants with low gi to have a larger draw-down from Ci to Cc, further confounding efforts to scale photosynthesis and other attributes with gi. Variation in the gi-photosynthesis relationship contributes to variation in photosynthetic 'use' efficiency of N (PNUE) and water (WUE). Delta is an information-rich signal, but for many species only about two-thirds of this information relates to A/gs with the remaining one-third related to A/gi. Using data for three well-studied species we demonstrate that at common WUE, delta may vary by up to 3 per thousand. This is as large or larger than is commonly reported in many interspecific comparisons of delta, and adds to previous warnings about simplistic interpretations of WUE based on delta. A priority for future research should be elucidation of relationships between gi and gs and how these vary in response to environmental conditions (e.g. soil water, leaf-to-air vapour pressure deficit, temperature) and among species.

The CBF1-dependent low temperature signalling pathway, regulon and increase in freeze tolerance are conserved in Populus spp

Abstract: The meristematic tissues of temperate woody perennials must acclimate to freezing temperatures to survive the winter and resume growth the following year. To determine whether the C-repeat binding factor (CBF) family of transcription factors contributing to this process in annual herbaceous species also functions in woody perennials, we investigated the changes in phenotype and transcript profile of transgenic Populus constitutively expressing CBF1 from Arabidopsis (AtCBF1). Ectopic expression of AtCBF1 was sufficient to significantly increase the freezing tolerance of non-acclimated leaves and stems relative to wild-type plants. cDNA microarray experiments identified genes up-regulated by ectopic AtCBF1 expression in Populus, demonstrated a strong conservation of the CBF regulon between Populus and Arabidopsis and identified differences between leaf and stem regulons. We studied the induction kinetics and tissue specificity of four CBF paralogues identified from the Populus balsamifera subsp. trichocarpa genome sequence (PtCBFs). All four PtCBFs are cold-inducible in leaves, but only PtCBF1 and PtCBF3 show significant induction in stems. Our results suggest that the central role played by the CBF family of transcriptional activators in cold acclimation of Arabidopsis has been maintained in Populus. However, the differential expression of the PtCBFs and differing clusters of CBF-responsive genes in annual (leaf) and perennial (stem) tissues suggest that the perennial-driven evolution of winter dormancy may have given rise to specific roles for these 'master-switches' in the different annual and perennial tissues of woody species.

Identification of QTL controlling root growth response to phosphate starvation in Arabidopsis thaliana

Abstract: One of the responses of plants to low sources of external phosphorus (P) is to modify root architecture. In Arabidopsis thaliana plantlets grown on low P, the primary root length (PRL) is reduced whereas lateral root growth is promoted. By using the Bay-0 x Shahdara recombinant inbred line (RIL) population, we have mapped three quantitative trait loci (QTL) involved in the root growth response to low P. The Shahdara alleles at these three QTL promote the response of the primary root to low P (i.e., root length reduction). One of these QTL, LPR1, located in a 2.8 Mb region at the top of chromosome 1, explains 52% of the variance of the PRL. We also detected a single QTL associated with primary root cell elongation in response to low P which colocalizes with LPR1. LPR1 does not seem to be involved in other typical P-starvation responses such as growth and density of root hairs, excretion of acid phosphatases, anthocyanin accumulation or the transcriptional induction of the P transporter Phtl;4. LPR1 might highlight new aspects of root growth that are revealed specifically under low P conditions.

Mechanical reinforcement of tracheids compromises the hydraulic efficiency of conifer xylem

Abstract: Wood structure and function of juvenile wood from 18 conifer species from four conifer families (Araucariaceae, Cupressaceae, Pinaceae and Podocarpaceae) were examined for a trade-off between wood reinforcement and hydraulic efficiency. Wood density and tracheid ‘thicknessto-span’ ratio were used as anatomical proxies for mechanical properties. The thickness:span represented the ratio of tracheid double wall thickness to lumen diameter. Hydraulic resistivity (R) of tracheids on a cross-sectional area basis (R CA ) increased over 50-fold with increasing density and thickness:span, implying a strength versus efficiency conflict. The conflict arose because density and thickness:span were increased by narrowing tracheid diameter rather than by thickening walls, which may be developmentally difficult. In the Pinaceae and Cupressaceae species, density and thickness:span correlated strongly with protection from drought-induced embolism, suggesting that mechanical strength was required in part to withstand tracheid collapse by negative sap pressure. These species showed a corresponding trade-off between increasing R CA and embolism protection. In contrast, species of Podocarpaceae and Araucariaceae were overbuilt for their embolism protection and were hydraulically inefficient, having greater density, thickness:span and R CA , none of which were correlated with vulnerability to embolism.

The dehydrogenase‐mediated recycling of NADPH is a key antioxidant system against salt‐induced oxidative stress in olive plants

Abstract: NADPH is an important molecule in the redox balance of the cell. In this paper, using olive tissue cultures as a model of the function of the NADPH-generating dehydrogenases in the mechanism of oxidative stress induced by severe salinity conditions was studied. When olive (Olea europaea) plants were grown with 200 mM NaCl, a 40% reduction in leaf fresh weight was produced. The content of non-enzymatic antioxidants such as ascorbate and glutathione was diminished between 20% to 39%, whereas the H2O2 content was increased threefold. In contrast, the analysis of the activity and protein contents of the main antioxidative enzymes showed a significant increase of catalase, superoxide dismutase and glutathione reductase. Overall, these changes strongly suggests that NaCl induces oxidative stress in olive plants. On the other hand, while the content of glucose-6-phosphate was increased almost eightfold in leaves of plants grown under salt stress, the content of NAD(P)H (reduced and oxided forms) did not show significant variations. Under salt stress conditions, the activity and protein contents of the main NADPH-recycling enzymes, glucose-6-phosphate dehydrogenase (G6PDH), isocitrate dehydrogenase (ICDH), malic enzyme (ME) and ferrodoxin-NADP reductase (FNR) showed an enhancement of 30-50%. In leaves of olive plants grown with 200 mM NaCl, analysis of G6PDH by immunocytochemistry and confocal laser scanning microscopy showed a general increase of this protein in epidermis, palisade and spongy mesophyll cells. These results indicate that in olive plants, salinity causes reactive oxygen species (ROS)-mediated oxidative stress, and plants respond to this situation by inducing different antioxidative enzymes, especially the NADPH-producing dehydrogenases in order to recycle NADPH necessary for the protection against oxidative damages. These NADP-dehydrogenases appear to be key antioxidative enzymes in olive plants under salt stress conditions.

Evidence from Amazonian forests is consistent with isohydric control of leaf water potential

Abstract: Climate modelling studies predict that the rain forests of the Eastern Amazon basin are likely to experience reduc- tions in rainfall of up to 50% over the next 50-100 years. Efforts to predict the effects of changing climate, especially drought stress, on forest gas exchange are currently limited by uncertainty about the mechanism that controls stomatal closure in response to low soil moisture. At a through-fall exclusion experiment in Eastern Amazonia where water was experimentally excluded from the soil, we tested the hypothesis that plants are isohydric, that is, when water is scarce, the stomata act to prevent leaf water potential from dropping below a critical threshold level. We made diurnal measurements of leaf water potential ( Ψ l ), stomatal con- ductance ( g s ), sap flow and stem water potential ( Ψ stem ) in the wet and dry seasons. We compared the data with the predictions of the soil-plant-atmosphere (SPA) model, which embeds the isohydric hypothesis within its stomatal conductance algorithm. The model inputs for meteorology, leaf area index (LAI), soil water potential and soil-to-leaf hydraulic resistance ( R ) were altered between seasons in accordance with measured values. No optimization param- eters were used to adjust the model. This 'mechanistic' model of stomatal function was able to explain the individ- ual tree-level seasonal changes in water relations ( r 2 = 0.85, 0.90 and 0.58 for Ψ l , sap flow and g s , respectively). The model indicated that the measured increase in R was the dominant cause of restricted water use during the dry sea- son, resulting in a modelled restriction of sap flow four times greater than that caused by reduced soil water poten- tial. Higher resistance during the dry season resulted from an increase in below-ground resistance (including root and soil-to-root resistance) to water flow.

Both abscisic acid (ABA)‐dependent and ABA‐independent pathways govern the induction of NCED3, AAO3 and ABA1 in response to salt stress

Abstract: The responsiveness of plants to osmotic stress is critically mediated by the increase in abscisic acid (ABA) levels. Osmotic stress induces the biosynthesis of ABA, whose increased levels subsequently exert a positive feedback on its own biosynthetic pathway. As only qualitative or semi-quantitative analyses were performed to test the inducibility of ABA biosynthetic genes in Arabidopsis thaliana, we used quantitative reverse transcriptase-polymerase chain reaction to re-examine the induction of the ABA1, ABA2, ABA3, NCED3 and AAO3 genes by NaCl and ABA. Quantitative gene expression data obtained from wild-type plants and severely ABA-deficient mutants support the prevailing notion that the 9-cis-epoxycarotenoid cleavage reaction is the key regulatory step in NaCl-induced ABA biosynthesis. Interestingly, strong induction by NaCl of NCED3 was still observed in severe ABA-deficient mutants, pointing to an ABA-independent induction pathway for NCED3 that is NaCl-dependent. Therefore, in the absence of the ABA-mediated positive feedback on ABA biosynthesis, the ABA-independent pathway makes a major contribution to the induction of key ABA biosynthetic genes, such as NCED3, AAO3 and ABA1. In addition, and in contrast to some previous reports, our data do not support the limited ability of ABA to induce NCED3 expression. Under our experimental conditions, the induction of NCED3 by ABA, either in wild-type plants or ABA-deficient mutants, was predominant over that of other ABA biosynthetic genes. Natural variability was found in the induction by NaCl and ABA of NCED3 and ABA1 expression in different Arabidopsis accessions, although NCED3 expression was clearly predominant.

13CO2 pulse-labelling of photoassimilates reveals carbon allocation within and between tree rings

Abstract: Post-photosynthetic fractionation processes during translocation, storage and remobilization of photoassimilate are closely related to intra-annual δ13C of tree rings, and understanding how these processes affect tree-ring δ13C is therefore indispensable for improving the quality of climate reconstruction. Our first objective was to study the relationship between translocation path and phloem grain. We pulse-labelled a branch of Larix gmelinii (Rupr.) Rupr. and later analysed the δ13C distribution in the stem. A 13C spiral translocation path closely related to the spiral grain was observed. Our second objective was to study the use of remobilized storage material for earlywood formation in spring, which is a suspected cause of the autocorrelation (correlation of ring parameters to the climate in the previous year) observed in (isotope) dendroclimatology. We pulse-labelled whole trees to study how spring, summer and autumn photoassimilate is later used for both earlywood and latewood formation. Analysis of intra-annual δ13C of the tree rings formed after the labelling revealed that earlywood contained photoassimilate from the previous summer and autumn as well as from the current spring. Latewood was mainly composed of photoassimilate from the current year’s summer/autumn, although it also relied on stored material in some cases. These results emphasize the need for separating earlywood and latewood for climate reconstruction work with narrow boreal tree rings.

Estimating stomatal conductance with thermal imagery

Abstract: Most thermal methods for the study of drought responses in plant leaves are based on the calculation of 'stress indices'. This paper proposes and compares three main extensions of these for the direct estimation of absolute values of stomatal conductance to water vapour (g s ) using infrared thermography (IRT). All methods use the measured leaf temperature and two environmental variables (air temperature and boundary layer resistance) as input. Additional variables required, depending on the method, are the temperatures of wet and dry reference surfaces, net radiation and relative humidity. The methods were compared using measured g s data from a vineyard in Southern Portugal. The errors in thermal estimates of conductance were of the same order as the measurement errors using a porometer. Observed variability was also compared with theoretical estimates of errors in estimated g, determined on the basis of the errors in the input variables (leaf temperature, boundary layer resistance, net radiation) and the partial derivatives of the energy balance equations used for the g, calculations. The full energy balance approach requires accurate estimates of net radiation absorbed, which may not be readily available in field conditions, so alternatives using reference surfaces are shown to have advantages. A new approach using a dry reference leaf is particularly robust and recommended for those studies where the specific advantages of thermal imagery, including its noncontact nature and its ability to sample large numbers of leaves, are most apparent. Although the results suggest that estimates of the absolute magnitude of gs are somewhat subjective, depending on the skill of the experimenter at selecting evenly exposed leaves, relative treatment differences in conductance are sensitively detected by different experimenters.

Photosynthetic capacity is related to the cellular and subcellular partitioning of Na+, K+ and Cl- in salt-affected barley and durum wheat.

Abstract: The capacity of plants to tolerate high levels of salinity depends on the ability to exclude salt from the shoot, or to tolerate high concentrations of salt in the leaf (tissue tolerance) It is widely held that a major component of tissue tolerance is the capacity to compartmentalize salt into safe storage places such as vacuoles This mechanism would avoid toxic effects of salt on photosynthesis and other key metabolic processes To test this, the relationship between photosynthetic capacity and the cellular and subcellular distribution of Na + , K + and Cl - was studied in salt-sensitive durum wheat (cv Wollaroi) and salt-tolerant barley (cv Franklin) seedlings grown in a range of salinity treatments Photosynthetic capacity parameters (V cmax , J max ) of salt-stressed Wollaroi decreased at a lower leaf Na + concentration than in Franklin Vacuolar concentrations of Na + , K + and Cl - in mesophyll and epidermal cells were measured using cryo-scanning electron microscopy (SEM) X-ray microanalysis In both species, the vacuolar Na + concentration was similar in mesophyll and epidermal cells, whereas K + was at higher concentrations in the mesophyll, and Cl - higher in the epidermis The calculated cytoplasmic Na + concentration increased to higher concentrations with increasing bulk leaf Na + concentration in Wollaroi compared to Franklin Vacuolar K + concentration was lower in the epidermal cells of Franklin than Wollaroi, resulting in higher cytoplasmic K + concentrations and a higher K + : Na + ratio This study indicated that the maintenance of photosynthetic capacity (and the resulting greater salt tolerance) at higher leaf Na + levels of barley compared to durum wheat was associated with the maintenance of higher K + , lower Na + and the resulting higher K + :Na + in the cytoplasm of mesophyll cells of barley