Showing papers in "Journal of Experimental Botany in 2002"

Cellular mechanisms for heavy metal detoxification and tolerance

Abstract: Heavy metals such as Cu and Zn are essential for normal plant growth, although elevated concentrations of both essential and non-essential metals can result in growth inhibition and toxicity symptoms. Plants possess a range of potential cellular mechanisms that may be involved in the detoxification of heavy metals and thus tolerance to metal stress. These include roles for the following: for mycorrhiza and for binding to cell wall and extracellular exudates; for reduced uptake or efflux pumping of metals at the plasma membrane; for chelation of metals in the cytosol by peptides such as phytochelatins; for the repair of stress-damaged proteins; and for the compartmentation of metals in the vacuole by tonoplast-located transporters. This review provides a broad overview of the evidence for an involvement of each mechanism in heavy metal detoxification and tolerance.

Role of superoxide dismutases (SODs) in controlling oxidative stress in plants

Abstract: Reactive O2 species (ROS) are produced in both unstressed and stressed cells. Plants have welldeveloped defence systems against ROS, involving both limiting the formation of ROS as well as instituting its removal. Under unstressed conditions, the formation and removal of O2 are in balance. However, the defence system, when presented with increased ROS formation under stress conditions, can be overwhelmed. Within a cell, the superoxide dismutases (SODs) constitute the first line of defence against ROS. Specialization of function among the SODs may be due to a combination of the influence of subcellular location of the enzyme and upstream sequences in the genomic sequence. The commonality of elements in the upstream sequences of Fe, Mn and CuuZn SODs suggests a relatively recent origin for those regulatory regions. The differences in the upstream regions of the three FeSOD genes suggest differing regulatory control which is borne out in the research literature. The finding that the upstream sequences of Mn and peroxisomal CuuZn SODs have three common elements suggests a common regulatory pathway. The tools are available to dissect further the molecular basis for antioxidant defence responses in plant cells. SODs are clearly among the most important of those defences, when coupled with the necessary downstream events for full detoxification of ROS.

Plant responses to abiotic stresses: heavy metal‐induced oxidative stress and protection by mycorrhization

Abstract: The aim of this review is to assess the mode of action and role of antioxidants as protection from heavy metal stress in roots, mycorrhizal fungi and mycorrhizae. Based on their chemical and physical properties three different molecular mechanisms of heavy metal toxicity can be distinguished: (a) production of reactive oxygen species by autoxidation and Fenton reaction; this reaction is typical for transition metals such as iron or copper, (b) blocking of essential functional groups in biomolecules, this reaction has mainly been reported for non-redox-reactive heavy metals such as cadmium and mercury, (c) displacement of essential metal ions from biomolecules; the latter reaction occurs with different kinds of heavy metals. Transition metals cause oxidative injury in plant tissue, but a literature survey did not provide evidence that this stress could be alleviated by increased levels of antioxidative systems. The reason may be that transition metals initiate hydroxyl radical production, which can not be controlled by antioxidants. Exposure of plants to non-redox reactive metals also resulted in oxidative stress as indicated by lipid peroxidation, H(2)O(2) accumulation, and an oxidative burst. Cadmium and some other metals caused a transient depletion of GSH and an inhibition of antioxidative enzymes, especially of glutathione reductase. Assessment of antioxidative capacities by metabolic modelling suggested that the reported diminution of antioxidants was sufficient to cause H(2)O(2) accumulation. The depletion of GSH is apparently a critical step in cadmium sensitivity since plants with improved capacities for GSH synthesis displayed higher Cd tolerance. Available data suggest that cadmium, when not detoxified rapidly enough, may trigger, via the disturbance of the redox control of the cell, a sequence of reactions leading to growth inhibition, stimulation of secondary metabolism, lignification, and finally cell death. This view is in contrast to the idea that cadmium results in unspecific necrosis. Plants in certain mycorrhizal associations are less sensitive to cadmium stress than non-mycorrhizal plants. Data about antioxidative systems in mycorrhizal fungi in pure culture and in symbiosis are scarce. The present results indicate that mycorrhization stimulated the phenolic defence system in the Paxillus-Pinus mycorrhizal symbiosis. Cadmium-induced changes in mycorrhizal roots were absent or smaller than those in non-mycorrhizal roots. These observations suggest that although changes in rhizospheric conditions were perceived by the root part of the symbiosis, the typical Cd-induced stress responses of phenolics were buffered. It is not known whether mycorrhization protected roots from Cd-induced injury by preventing access of cadmium to sensitive extra- or intracellular sites, or by excreted or intrinsic metal-chelators, or by other defence systems. It is possible that mycorrhizal fungi provide protection via GSH since higher concentrations of this thiol were found in pure cultures of the fungi than in bare roots. The development of stress-tolerant plant-mycorrhizal associations may be a promising new strategy for phytoremediation and soil amelioration measures.

Cereal seed storage proteins: structures, properties and role in grain utilization

Abstract: Storage proteins account for about 50% of the total protein in mature cereal grains and have important impacts on their nutritional quality for humans and livestock and on their functional properties in food processing. Current knowledge of the structures and properties of the prolamin and globulin storage proteins of cereals and their mechanisms of synthesis, trafficking and deposition in the developing grain is briefly reviewed here. The role of the gluten proteins of wheat in determining the quality of the grain for breadmaking and how their amount and composition can be manipulated leading to changes in dough mixing properties is also discussed.

Regulation and function of ascorbate peroxidase isoenzymes

Abstract: Even under optimal conditions, many metabolic processes, including the chloroplastic, mitochondrial, and plasma membrane-linked electron transport systems of higher plants, produce active oxygen species (AOS). Furthermore, the imposition of biotic and abiotic stress conditions can give rise to excess concentrations of AOS, resulting in oxidative damage at the cellular level. Therefore, antioxidants and antioxidant enzymes function to interrupt the cascades of uncontrolled oxidation in each organelle. Ascorbate peroxidase (APX) exists as isoenzymes and plays an important role in the metabolism of H(2)O(2) in higher plants. APX is also found in eukaryotic algae. The characterization of APX isoenzymes and the sequence analysis of their clones have led to a number of investigations that have yielded interesting and novel information on these enzymes. Interestingly, APX isoenzymes of chloroplasts in higher plants are encoded by only one gene, and their mRNAs are generated by alternative splicing of the gene's two 3'-terminal exons. Manipulation of the expression of the enzymes involved in the AOS-scavenging systems by gene-transfer technology has provided a powerful tool for increasing the present understanding of the potential of the defence network against oxidative damage caused by environmental stresses. Transgenic plants expressing E. coli catalase to chloroplasts with increased tolerance to oxidative stress indicate that AOS-scavenging enzymes, especially chloroplastic APX isoenzymes are sensitive under oxidative stress conditions. It is clear that a high level of endogenous ascorbate is essential effectively to maintain the antioxidant system that protects plants from oxidative damage due to biotic and abiotic stresses.

Hydrogen peroxide and nitric oxide as signalling molecules in plants

Abstract: It is now clear that hydrogen peroxide (H(2)O(2)) and nitric oxide (NO) function as signalling molecules in plants. A wide range of abiotic and biotic stresses results in H(2)O(2) generation, from a variety of sources. H(2)O(2) is removed from cells via a number of antioxidant mechanisms, both enzymatic and non-enzymatic. Both biotic and abiotic stresses can induce NO synthesis, but the biosynthetic origins of NO in plants have not yet been resolved. Cellular responses to H(2)O(2) and NO are complex, with considerable cross-talk between responses to several stimuli. In this review the potential roles of H(2)O(2) and NO during various stresses and the signalling pathways they activate are discussed. Key signalling components that might provide targets for enhancing crop production are also identified.

Signal transduction during oxidative stress

Abstract: As an unfortunate consequence of aerobic life, active oxygen species (AOS) are formed by partial reduction of molecular oxygen. Plants possess a complex battery of enzymatic and non-enzymatic antioxidants that can protect cells from oxidative damage by scavenging AOS. It is becoming evident that AOS, which are generated during pathogen attack and abiotic stress situations, are recognized by plants as a signal for triggering defence responses. An overview of the literature is presented on the signalling role of AOS in plant defence responses, cell death, and development. Special attention is given to AOS and redox-regulated gene expression and the role of kinases and phosphatases in redox signal transduction.

Ethylene biosynthesis and action in tomato: a model for climacteric fruit ripening

Abstract: Elucidating the mechanisms involved in ripening of climacteric fruit and the role that ethylene plays in the process are key to understanding fruit production and quality. In this review, which is based largely on research in tomato, particular attention is paid to the role of specific isoforms of ACC synthase and ACC oxidase in controlling ethylene synthesis during the initiation and subsequent autocatalytic phase of ethylene production during ripening. Recent information on the structure and role of six different putative ethylene receptors in tomato is discussed, including evidence supporting the receptor inhibition model for ripening, possible differences in histidine kinase activity between receptors, and the importance of receptor LeETR4 in ripening. A number of ethylene-regulated ripening-related genes are discussed, including those involved in ethylene synthesis, fruit texture, and aroma volatile production, as well as experiments designed to elucidate the ethylene signalling pathway from receptor through intermediate components similar to those found in Arabidopsis, leading to transcription factors predicted to control the expression of ethylene-regulated genes.

Interactions between biosynthesis, compartmentation and transport in the control of glutathione homeostasis and signalling

Abstract: Glutathione has numerous roles in cellular defence and in sulphur metabolism. These functions depend or impact on the concentration and/or redox state of leaf glutathione pools. Effective function requires homeostatic control of concentration and redox state, with departures from homeostasis acting as signals that trigger adaptive responses. Intercellular and intracellular glutathione pools are linked by transport across membranes. It is shown that glutathione can cross the chloroplast envelope at rates similar to the speed of biosynthesis. Control of glutathione concentration and redox state is therefore due to a complex interplay between biosynthesis, utilization, degradation, oxidation/reduction, and transport. All these factors must be considered in order to evaluate the significance of glutathione as a signalling component during development, abiotic stress, or pathogen attack.

Water stress‐induced abscisic acid accumulation triggers the increased generation of reactive oxygen species and up‐regulates the activities of antioxidant enzymes in maize leaves

Abstract: The interrelationship among water-stress-induced abscisic acid (ABA) accumulation, the generation of reactive oxygen species (ROS), and the activities of several antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR) was investigated in leaves of detached maize (Zea mays L.) plants exposed to -0.7 MPa water stress induced by polyethylene glycol (PEG 6000). Time-course analyses of ABA content, the production of ROS, and the activities of antioxidant enzymes in water-stressed leaves showed that a significant increase in the content of ABA preceded that of ROS, which was followed by a marked increase in the activities of these antioxidant enzymes. Pretreatment with an ABA biosynthesis inhibitor, tungstate, significantly suppressed the accumulation of ABA, and also reduced the increased generation of ROS and the up-regulation of these antioxidant enzymes in water-stressed leaves. A mild oxidative stress induced by paraquat, which generates O - 2 and then H 2 O 2 , resulted In a significant enhancement in the activities of antioxidant enzymes in non-water-stressed leaves. Pretreatment with some ROS scavengers, such as Tiron and dimethylthiourea (DMTU), and an inhibitor of NAD(P)H oxidase, diphenyleneiodonium (DPI), almost completely arrested the increase In ROS and the activities of these antioxidant enzymes induced by water stress or ABA treatment. These data suggest that water stress-Induced ABA accumulation triggers the increased generation of ROS, which, In turn, leads to the up-regulation of the antioxidant defence system.

Regulation of nitric oxide (NO) production by plant nitrate reductase in vivo and in vitro.

Abstract: NO (nitric oxide) production from sunflower plants (Helianthus annuus L.), detached spinach leaves (Spinacia oleracea L.), desalted spinach leaf extracts or commercial maize (Zea mays L.) leaf nitrate reductase (NR, EC 1.6.6.1) was continuously followed as NO emission into the gas phase by chemiluminescence detection, and its response to post-translational NR modulation was examined in vitro and in vivo. NR (purified or in crude extracts) in vitro produced NO at saturating NADH and nitrite concentrations at about 1% of its nitrate reduction capacity. The K(m) for nitrite was relatively high (100 microM) compared to nitrite concentrations in illuminated leaves (10 microM). NO production was competitively inhibited by physiological nitrate concentrations (K(i)=50 microM). Importantly, inactivation of NR in crude extracts by protein phosphorylation with MgATP in the presence of a protein phosphatase inhibitor also inhibited NO production. Nitrate-fertilized plants or leaves emitted NO into purified air. The NO emission was lower in the dark than in the light, but was generally only a small fraction of the total NR activity in the tissue (about 0.01-0.1%). In order to check for a modulation of NO production in vivo, NR was artificially activated by treatments such as anoxia, feeding uncouplers or AICAR (a cell permeant 5'-AMP analogue). Under all these conditions, leaves were accumulating nitrite to concentrations exceeding those in normal illuminated leaves up to 100-fold, and NO production was drastically increased especially in the dark. NO production by leaf extracts or intact leaves was unaffected by nitric oxide synthase inhibitors. It is concluded that in non-elicited leaves NO is produced in variable quantities by NR depending on the total NR activity, the NR activation state and the cytosolic nitrite and nitrate concentration.

N uptake and distribution in crops: an agronomical and ecophysiological perspective

Abstract: The rate of N uptake of crops is highly variable during crop development and between years and sites. However, under ample soil N availability, crop N accumulation is highly related to crop growth rate and to biomass accumulation. Critical N concentration has been defined as the minimum N concentration which allows maximum growth rate. Critical N concentration declines during crop growth. The relationship between critical N concentration and biomass accumulation over the growth period of a crop is broadly similar within major C(3) and C(4) cultivated species. Therefore, the critical N concentration concept is widely used in agronomy as the basis of the diagnosis of crop N status, and allows discrimination between situations of sub-optimal and supra-optimal N supply. The relationship between N and biomass accumulation in crops, relies on the interregulation of multiple crop physiological processes. Among these processes, N uptake, crop C assimilation and thus growth rate, and C and N allocation between organs and between plants, play a particular role. Under sub-optimal N supply, N uptake of the crop depends on soil mineral N availability and distribution, and on root distribution. Under ample N supply, N uptake largely depends on growth rate via internal plant regulation. Carbon assimilation of the crop is related to crop N through the distribution of N between mature leaves with consequences for leaf and canopy photosynthesis. However, although less commonly emphasized, carbon assimilation of the crop also depends on crop N through leaf area development. Therefore, crop growth rate fundamentally relies on the balance of N allocation between growing and mature leaves. Nitrogen uptake and distribution also depends on C allocation between organs and N composition of these organs. Within shoots, allocation of C to stems generally increases in relation to C allocation to the leaves over the crop growth period. Allocation of C and N between shoots and roots also changes to a large extent in relation to soil N and/or crop N. These alterations in C and N allocation between plant organs have implications, together with soil availability and carbon assimilation, on N uptake and distribution in crops. Therefore, N uptake and distribution in plants and crops involves many aspects of growth and development. Regulation of nitrogen assimilation needs to be considered in the context of these interregulatory processes.

The apoplastic oxidative burst in response to biotic stress in plants: a three‐component system

Abstract: The oxidative burst, the generation of reactive oxygen species (ROS) in response to microbial pathogen attack, is a ubiquitous early part of the resistance mechanisms of plant cells. It has also become apparent from the study of a number of plant-pathogen interactions and those modelled by elicitor treatment of cultured cells that there may be more than one mechanism operating. However, one mechanism may be dominant in any given species. NADPH oxidases have been implicated in a number of systems and have been cloned and characterized. However, the enzyme system which is the major source of ROS in French bean (Phaseolus vulgaris) cells treated with a cell wall elicitor from Colletotrichum lindemuthianum, appears to be dependent on an exocellular peroxidase. The second component, the extracellular alkalinization, occurs as a result of the Ca(2+) and proton influxes and the K(+) efflux common to most elicitation systems as one of the earliest responses. The third component, the actual reductant/substrate, has remained elusive. The low molecular weight compound composition of apoplastic fluid was compared before and after elicitation. The substrate only becomes available some min after elicitation and can be extracted, so that by comparing the profiles by LC-MS it has been possible to identify possible substrates. The mechanism has proved to be complex and may involve a number of low molecular weight components. Stimulation of H(2)O(2) production was observed with saturated fatty acids such as palmitate and stearate without concomitant oxylipin production. This biochemical evidence is supported by immunolocalization studies on papillae forming at bacterial infection sites that show the peroxidase isoform present at sites of H(2)O(2) production revealed by cerium chloride staining together with the cross-linked wall proteins and callose and callose synthase. The peroxidase has been cloned and expressed in Pichia pastoris and has been shown to catalyse the oxidation reaction with the same kinetics as the purified enzyme. Furthermore, Arabidopsis plants transformed heterologously using the French bean peroxidase in antisense orientation have proved to be highly susceptible to bacterial and fungal pathogens. Thus it is possible that Arabidopsis is another species with the potential to mount an apoplastic oxidative burst and these transformed plant lines may be useful to identify the peroxidase that is responsible.

Nitrogen metabolism and remobilization during senescence.

Abstract: Senescence is a highly organized and well-regulated process. As much as 75% of total cellular nitrogen may be located in mesophyll chloroplasts of C(3)-plants. Proteolysis of chloroplast proteins begins in an early phase of senescence and the liberated amino acids can be exported to growing parts of the plant (e.g. maturing fruits). Rubisco and other stromal enzymes can be degraded in isolated chloroplasts, implying the involvement of plastidial peptide hydrolases. Whether or not ATP is required and if stromal proteins are modified (e.g. by reactive oxygen species) prior to their degradation are questions still under debate. Several proteins, in particular cysteine proteases, have been demonstrated to be specifically expressed during senescence. Their contribution to the general degradation of chloroplast proteins is unclear. The accumulation in intact cells of peptide fragments and inhibitor studies suggest that multiple degradation pathways may exist for stromal proteins and that vacuolar endopeptidases might also be involved under certain conditions. The breakdown of chlorophyll-binding proteins associated with the thylakoid membrane is less well investigated. The degradation of these proteins requires the simultaneous catabolism of chlorophylls. The breakdown of chlorophylls has been elucidated during the last decade. Interestingly, nitrogen present in chlorophyll is not exported from senescencing leaves, but remains within the cells in the form of linear tetrapyrrolic catabolites that accumulate in the vacuole. The degradation pathways for chlorophylls and chloroplast proteins are partially interconnected.

The role of glutamine synthetase and glutamate dehydrogenase in nitrogen assimilation and possibilities for improvement in the nitrogen utilization of crops.

Abstract: This short review outlines the central role of glutamine synthetase (GS) in plant nitrogen metabolism and discusses some possibilities for crop improvement. GS functions as the major assimilatory enzyme for ammonia produced from N fixation, and nitrate or ammonia nutrition. It also reassimilates ammonia released as a result of photorespiration and the breakdown of proteins and nitrogen transport compounds. GS is distributed in different subcellular locations (chloroplast and cytoplasm) and in different tissues and organs. This distribution probably changes as a function of the development of the tissue, for example, GS1 appears to play a key role in leaf senescence. The enzyme is the product of multiple genes with complex promoters that ensure the expression of the genes in an organ- and tissue-specific manner and in response to a number of environmental variables affecting the nutritional status of the cell. GS activity is also regulated post-translationally in a manner that involves 14-3-3 proteins and phosphorylation. GS and plant nitrogen metabolism is best viewed as a complex matrix continually changing during the development cycle of plants. Along with GS, a number of other enzymes play key roles in maintaining the balance of carbon and nitrogen. It is proposed that one of these is glutamate dehydrogenase (GDH). There is considerable evidence for a GDH shunt to return the carbon in amino acids back into reactions of carbon metabolism and the tri-carboxylic acid cycle. Results with transgenic plants containing transferred GS genes suggest that there may be ways in which it is possible to improve the efficiency with which crop plants use nitrogen. Marker-assisted breeding may also bring about such improvements.

Carbon and nitrogen assimilation in relation to yield: mechanisms are the key to understanding production systems

Abstract: Improved understanding of crop production systems in relation to N-supply has come from a knowledge of basic plant biochemistry and physiology. Gene expression leads to protein synthesis and the formation of metabolic systems; the ensuing metabolism determines the capacity for growth, development and yield production. This constitutes the genetic potential. These processes set the requirements for the supply of resources. The interactions between carbon dioxide (CO(2)) and nitrate () assimilation and their dynamics are of key importance for crop production. In particular, an adequate supply of, its assimilation to amino acids (for which photosynthesized carbon compounds are required) and their availability for protein synthesis, are essential for metabolism. An adequate supply of stimulates leaf growth and photosynthesis, the former via cell growth and division, the latter by larger contents of components of the light reactions, and those of CO(2) assimilation and related processes. If the supply of resources exceeds the demand set by the genetic potential then production is maximal, but if it is less then potential is not reached; matching resources to potential is the aim of agriculture. However, the connection between metabolism and yield is poorly quantified. Biochemical characteristics and simulation models must be better used and combined to improve fertilizer-N application, efficiency of N-use, and yields. Increasing N-uptake at inadequate N-supply by increasing rooting volume and density is feasible, increasing affinity is less so. It would increase biomass and N/C ratio. With adequate N, at full genetic potential, more C-assimilation per unit N would increase biomass, but energy would be limiting at full canopy. Increasing C-assimilation per unit N would increase biomass but decrease N/C at both large and small N-supply. Increasing production of all biochemical components would increase biomass and demand for N, and maintain N/C ratio. Changing C- or N-assimilation requires modifications to many processes to effect improvements in the whole system; genetic engineering/molecular biological alterations to single steps in the central metabolism are unlikely to achieve this, because targets are unclear, and also because of the complex interactions between processes and environment. Achievement of the long-term objectives of improving crop N-use and yield with fewer inputs and less pollution, by agronomy, breeding or genetic engineering, requires a better understanding of the whole system, from genes via metabolism to yield.

Steps towards an integrated view of nitrogen metabolism

Abstract: This article discusses how nitrate assimilation is integrated with nitrate uptake, with ammonium assimilation and amino acid synthesis, with pH regulation, and with the sugar supply in tobacco leaves. During the first part of the light period, nitrate assimilation exceeds nitrate uptake by 2-fold and ammonium assimilation by 50%, leading to rapid depletion of nitrate and accumulation of ammonium, glutamine, glycine and serine. NIA, NII and PPC expression show a shared maximum early in the diurnal cycle to direct carbon towards malate synthesis for pH regulation. Later in the diurnal cycle an orchestrated increase of GLN2, PKc, CS, and ICDH-1 expression re-establishes a balance between nitrate assimilation and ammonium metabolism. Nitrate uptake continues throughout the night, replenishing the leaf nitrate pool. These diurnal changes are attenuated or abolished in mutants with low NIA activity, and modified in wild-type plants growing on different nitrogen sources or elevated [CO(2)]. Comparison across genotypes and conditions reveals that NIA transcript levels are always closely related to the balance between nitrate influx and assimilation, but are unrelated to changes of glutamine or 2-oxoglutarate. In a systematic search for other downstream regulators, a wide range of downstream metabolites was fed to detached leaves and glutamate, cysteine, asparagine, and malate identified as candidates. Low sugars totally inhibit nitrate assimilation, overriding signals from nitrogen metabolism. Moderate changes act post-transcriptionally, and larger changes lead to a collapse of the NIA transcript. Low sugars also lead to a collapse of minor amino acids and a dramatic decrease of phenylpropanoids and nicotine. Consequently, wild-type plants growing in unfavourable light regimes and antisense RBCS transformants are simultaneously carbon- and nitrogen-limited.

Molecular and physiological approaches to maize improvement for drought tolerance

Abstract: Average maize yields have increased steadily over the years in the USA and yet the variations in harvestable yield have also markedly increased. Much of the increase in yield variability can be attributed to (1) varying environmental stress conditions; (2) improved nitrogen inputs and better weed control; and (3) continuing sensitivity of different maize lines to the variation in input supply, especially rainfall. Drought stress alone can account for a significant percentage of average yield losses. Yet despite variable environments, new commercially available maize hybrids continue to be produced each year with ever-increasing harvestable yield. Since many factors contribute to high plant performance under water deficits, efforts are being made to elucidate the nature of water-stress tolerance in an attempt to improve maize hybrids further. Such factors include better partitioning of biomass to the developing ear resulting in faster spikelet growth and improved reproductive success. An emphasis on faster spikelet growth rate may result in a reduction in the number of spikelets formed on the ear that facilitates overall seed set by reducing water and carbon constraints per spikelet. To understand the molecular mechanisms for drought tolerance in improved maize lines better, a variety of genomic tools are being used. Newer molecular markers and comprehensive gene expression profiling methods provide opportunities to direct the continued breeding of genotypes that provide stable grain yield under widely varied environmental conditions.

Reactive oxygen species, antioxidant systems and nitric oxide in peroxisomes

Abstract: Peroxisomes are subcellular organelles with an essentially oxidative type of metabolism. Like chloroplasts and mitochondria, plant peroxisomes also produce superoxide radicals (O .- 2 ) and there are, at least, two sites of superoxide generation: one in the organelle matrix, the generating system being xanthine oxidase, and another site in the peroxisomal membranes dependent on NAD(P)H. In peroxisomal membranes, three integral polypeptides (PMPs) with molecular masses of 18, 29 and 32 kDa have been shown to generate O .- 2 radicals. Besides catalase, several antioxidative systems have been demonstrated in plant peroxisomes, including different superoxide dismutases, the ascorbate-glutathione cycle, and three NADP-dependent dehydrogenases. A CuZn-SOD and two Mn-SODs have been purified and characterized from different types of peroxisomes. The four enzymes of the ascorbate-glutathione cycle (ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase, and glutathione reductase) as well as the antioxidants glutathione and ascorbate have been found in plant peroxisomes. The recycling of NADPH from NADP + can be carried out in peroxisomes by three dehydrogenases: glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and isocitrate dehydrogenase. In the last decade, different experimental evidence has suggested the existence of cellular functions for peroxisomes related to reactive oxygen species (ROS), but the recent demonstration of the presence of nitric oxide synthase (NOS) in plant peroxisomes implies that these organelles could also have a function in plant cells as a source of signal molecules like nitric oxide (NO), superoxide radicals, hydrogen peroxide, and possibly S-nitrosoglutathione (GSNO).

ABA, ethylene and the control of shoot and root growth under water stress

Abstract: The question of whether abscisic acid (ABA) acts as an inhibitor or promoter of shoot growth in plants growing in drying soil is examined, drawing on current understanding of the role of ABA in root growth maintenance. Particular consideration is given to studies of endogenous ABA deficiency, which have shown that an important role of ABA is to limit ethylene production, and that this interaction is involved in the effects of ABA status on shoot and root growth.

Use of infrared thermography for monitoring stomatal closure in the field: application to grapevine

Abstract: This paper reviews and discusses strategies for the use of thermal imaging for studies of stomatal con ductance in the field and compares techniques for image collection and analysis. Measurements were taken under a range of environmental conditions and on sunlit and shaded canopies to illustrate the vari ability of temperatures and derived stress indices. A simple procedure is presented for correcting for cali bration drift within the images from the low-cost thermal imager used (Snapshot 225, Infrared Solutions, Inc.). The use of wet and dry reference surfaces as thresholds to eliminate the inclusion of non-leaf material in the analysis of canopy tempera ture is discussed. An index that is proportional to stomatal conductance was compared with stomatal measurements with a porometer. The advantages and disadvantages of a possible new approach to the use of thermal imagery for the detection of stomatal closure in grapevine canopies, based on an analysis of the temperature of shaded leaves, rather than sunlit leaves, are discussed. Evidence is presented that the temperature of reference surfaces exposed within the canopy can be affected by the canopy water status.

Imaging of photo-oxidative stress responses in leaves

Abstract: High resolution digital imaging was used to identify sites of photo-oxidative stress responses in Arabidopsis leaves non-invasively, and to demonstrate the potential of using a suite of imaging techniques for the study of oxidative metabolism in planta. Tissue-specific photoinhibition of photosynthesis in individual chloroplasts in leaves was imaged by chlorophyll fluorescence microscopy. Singlet oxygen production was assessed by imaging the quenching of the fluorescence of dansyl-2,2,5,5-tetramethyl-2,5-dihydro-1H-pyrrole (DanePy) that results from its reaction with singlet oxygen. Superoxide and hydrogen peroxide accumulation were visualized by the reduction of nitroblue tetrazolium (NBT) to formazan deposits and by polymerization with 3,3'-diaminobenzidine (DAB), respectively. Stress-induced expression of a gene involved with antioxidant metabolism was imaged from the bioluminescence from leaves of an Arabidopsis APX2-LUC transformant, which co-expresses an ascorbate peroxidase (APX2) with firefly luciferase. Singlet oxygen and superoxide production were found to be primarily located in mesophyll tissues whereas hydrogen peroxide accumulation and APX2 gene expression were primarily localized in the vascular tissues.

The regulation of nitrate and ammonium transport systems in plants

Abstract: Inorganic nitrogen concentrations in soil solutions vary across several orders of magnitude among different soils and as a result of seasonal changes In order to respond to this heterogeneity, plants have evolved mechanisms to regulate and influx In addition, efflux analysis using (13)N has revealed that there is a co-ordinated regulation of all component fluxes within the root, including biochemical fluxes Physiological studies have demonstrated the presence of two high-affinity transporter systems (HATS) for and one HATS for in roots of higher plants By contrast, in Arabidopsis thaliana there exist seven members of the NRT2 family encoding putative HATS for and five members of the AMT1 family encoding putative HATS for The induction of high-affinity transport and Nrt21 and Nrt22 expression occur in response to the provision of, while down-regulation of these genes appear to be due to the effects of glutamine High-affinity transport and AMT11 expression also appear to be subject to down-regulation by glutamine In addition, there is evidence that accumulated and may act post-transcriptionally on transporter function The present challenge is to resolve the functions of all of these genes In Aspergillus nidulans and Chlamydomonas reinhardtii there are but two high-affinity transporters and these appear to have undergone kinetic differentiation that permits a greater efficiency of absorption over the wide range of concentration normally found in nature Such kinetic differentiation may also have occurred among higher plant transporters The characterization of transporter function in higher plants is currently being inferred from patterns of gene expression in roots and shoots, as well as through studies of heterologous expression systems and knockout mutants

Localized versus systemic effect of arbuscular mycorrhizal fungi on defence responses to Phytophthora infection in tomato plants

Abstract: Development of biological control for plant diseases is accepted as a durable and environmentally friendly alternative for agrochemicals. Arbuscular mycorrhizal fungi (AMF), which form symbiotic associations with root systems of most agricultural, horticultural and hardwood crop species, have been suggested as widespread potential bioprotective agents. In the present study the ability of two AMF (Glomus mosseae and Glomus intraradices) to induce local or systemic resistance to Phytophthora parasitica in tomato roots have been compared using a split root experimental system. Glomus mosseae was effective in reducing disease symptoms produced by P. parasitica infection, and evidence points to a combination of local and systemic mechanisms being responsible for this bioprotector effect. The biochemical analysis of different plant defencerelated enzymes showed a local induction of mycorrhiza-related new isoforms of the hydrolytic enzymes chitinase, chitosanase and b-1,3-glucanase, as well as superoxide dismutase, an enzyme which is involved in cell protection against oxidative stress. Systemic alterations of the activity of some of the constitutive isoforms were also observed in non-mycorrhizal roots of mycorrhizal plants. Studies on the lytic activity against Phytophthora cell wall of root protein extracts also corroborated a systemic effect of mycorrhizal symbiosis on tomato resistance to Phytophthora.

The role of phytochelatins in constitutive and adaptive heavy metal tolerances in hyperaccumulator and non‐hyperaccumulator metallophytes

Abstract: Using the g-glutamylcysteine synthetase inhibitor, L-buthionine-[S,R]-sulphoximine (BSO), the role for phytochelatins (PCs) was evaluated in Cu, Cd, Zn, As, Ni, and Co tolerance in non-metallicolous and metallicolous, hypertolerant populations of Silene vulgaris (Moench) Garcke, Thlaspi caerulescens J.&C. Presl., Holcus lanatus L., and Agrostis castellana Boiss. et Reuter. Based on plant-internal PCthiol to metal molar ratios, the metals’ tendency to induce PC accumulation decreased in the order As/Cd/Cu > Zn > Ni/Co, and was consistently higher in non-metallicolous plants than in hypertolerant ones, except for the case of As. The sensitivities to Cu, Zn, Ni, and Co were consistently unaffected by BSO treatment, both in non-metallicolous and hypertolerant plants, suggesting that PC-based sequestration is not essential for constitutive tolerance or hypertolerance to these metals. Cd sensitivity was considerably increased by BSO, though exclusively in plants lacking Cd hypertolerance, suggesting that adaptive cadmium hypertolerance is not dependent on PC-mediated sequestration. BSO dramatically increased As sensitivity, both in non-adapted and As-hypertolerant plants, showing that PC-based sequestration is essential for both normal constitutive tolerance and adaptive hypertolerance to this metalloid. The primary function of PC synthase in plants and algae remains elusive.

Linking drought‐resistance mechanisms to drought avoidance in upland rice using a QTL approach: progress and new opportunities to integrate stomatal and mesophyll responses

Abstract: The advent of saturated molecular maps promised rapid progress towards the improvement of crops for genetically complex traits like drought resistance via analysis of quantitative trait loci (QTL). Progress with the identification of QTLs for drought resistance-related traits in rice is summarized here with the emphasis on a mapping population of a cross between drought-resistant varieties Azucena and Bala. Data which have used root morphological traits and indicators of drought avoidance in field-grown plants are reviewed, highlighting problems and uncertainties with the QTL approach. The contribution of root-growth QTLs to drought avoidance appears small in the experiments so far conducted, and the limitations of screening methodologies and the involvement of shoot-related mechanisms of drought resistance are studied. When compared to Azucena, Bala has been observed to have highly sensitive stomata, does not roll its leaves readily, has a greater ability to adjust osmotically, slows growth more rapidly when droughted and has a lower water-use efficiency. It is also a semi-dwarf variety and hence has a different canopy structure. There is a need to clarify the contribution of the shoot to drought resistance from the level of the biochemistry of photosynthesis through stomatal behaviour and leaf anatomy to canopy architecture. Recent advances in studying the physical and biochemical processes related to water use and drought stress offer the opportunity to advance a more holistic understanding of drought resistance. These include the potential use of infrared thermal imaging to study energy balance, integrated and online stable isotope analysis to dissect processes involved in carbon dioxide fixation and water evaporation, and leaf fluorescence to monitor photosynthesis and photochemical quenching. Justification and a strategy for this integrated approach is described, which has relevance to the study of drought resistance in most crops.

Regulation of carotenoid formation during tomato fruit ripening and development

Abstract: Carotenoid biosynthesis and its regulation during tomato fruit development and ripening is a complex process that occurs alongside the differentiation of chloroplasts into chromoplasts and changes to the organoleptic properties of the fruit. Unusually for plants, the ripe tomato fruit accumulates large amounts of lycopene, as the pattern of gene expression found in green fruit changes during fruit ripening. Although the control of gene expression is thought to be the main regulatory mechanism for these alterations in carotenoids, post-transcriptional regulation has also been reported, including feedback inhibition. The use of genetic manipulation of carotenogenesis in tomato has been used primarily for biotechnological reasons, but it has also facilitated investigations into these regulatory mechanisms, as well as into the effects of such perturbations on other isoprenoids such as gibberellins, tocopherols and sterols.

Characteristics of cadmium uptake in two contrasting ecotypes of the hyperaccumulator Thlaspi caerulescens

Abstract: Uptake of Cd and Zn by intact seedlings of two contrasting ecotypes of the hyperaccumulator Thlaspi caerulescens was characterized using radioactive tracers. Uptake of Cd and Zn at 2 degrees C was assumed to represent mainly apoplastic binding in the roots, whereas the difference in uptake between 22 degrees C and 2 degrees C represented metabolically dependent influx. There was no significant difference between the two ecotypes in the apoplastic binding of Cd or Zn. Metabolically dependent uptake of Cd was 4.5-fold higher in the high Cd-accumulating ecotype, Ganges, than in the low Cd-accumulating ecotype, Prayon. By contrast, there was only a 1.5-fold difference in the Zn uptake between the two ecotypes. For the Ganges ecotype, Cd uptake could be described by Michaelis-Menten kinetics with a V(max) of 143 nmol g(-1) root FW h(-1) and a K(m) of 0.45 microM. Uptake of Cd by the Ganges ecotype was not inhibited by La, Zn, Cu, Co, Mn, Ni or Fe(II), and neither by increasing the Ca concentration. By contrast, addition of La, Zn or Mn, or increasing the Ca concentration in the uptake solution decreased Cd uptake by Prayon. Uptake of Ca was larger in Prayon than in Ganges. The results suggest that Cd uptake by the low Cd-accumulating ecotype (Prayon) may be mediated partly via Ca channels or transporters for Zn and Mn. By contrast, there may exist a highly selective Cd transport system in the root cell membranes of the high Cd-accumulating ecotype (Ganges) of T. caerulescens.

Regulation of the plant defence response in arbuscular mycorrhizal symbiosis.

Abstract: The response of plants to arbuscular mycorrhizal fungi involves a temporal and spatial activation of different defence mechanisms. The activation and regulation of these defences have been proposed to play a role in the maintenance of the mutualistic status of the association, however, how these defences affect the functioning and development of arbuscular mycorrhiza remains unclear. A number of regulatory mechanisms of plant defence response have been described during the establishment of the arbuscular mycorrhizal symbiosis, including elicitor degradation, modulation of second messenger concentration, nutritional and hormonal plant defence regulation, and activation of regulatory symbiotic gene expression. The functional characterization of these regulatory mechanisms on arbuscular mycorrhiza, including cross-talk between them, will be the aim and objective of future work on this topic.

Pectate lyases, cell wall degradation and fruit softening.

Abstract: This is a brief review of what is known about the role of pectate lyases in plants. The mode of action and three-dimensional structure of microbial pectate lyases is discussed first and then the limited information on the plant proteins is presented. Pectate lyase-like genes have been isolated from a wide range of plant tissues including germinating seeds, pollen, cell cultures, and ripening fruits. The abundance of ESTs for these genes in tomato and the presence of pectate lyase-like transcripts in many other fruits may indicate that these enzymes have a more important role in ripening than previously suspected.