How the Ethylene Biosynthesis Pathway of Semi-Halophytes Is Modified with Prolonged Salinity Stress Occurrence?

The mechanism of ethylene (ET)-regulated salinity stress response remains largely unexplained, especially for semi-halophytes and halophytes. Here, we present the results of the multifaceted analysis of the model semi-halophyte Mesembryanthemum crystallinum L. (common ice plant) ET biosynthesis pathway key components' response to prolonged (14 days) salinity stress. Transcriptomic analysis revealed that the expression of 3280 ice plant genes was altered during 14-day long salinity (0.4 M NaCl) stress. A thorough analysis of differentially expressed genes (DEGs) showed that the expression of genes involved in ET biosynthesis and perception (ET receptors), the abscisic acid (ABA) catabolic process, and photosynthetic apparatus was significantly modified with prolonged stressor presence. To some point this result was supported with the expression analysis of the transcript amount (qPCR) of key ET biosynthesis pathway genes, namely ACS6 (1-aminocyclopropane-1-carboxylate synthase) and ACO1 (1-aminocyclopropane-1-carboxylate oxidase) orthologs. However, the pronounced circadian rhythm observed in the expression of both genes in unaffected (control) plants was distorted and an evident downregulation of both orthologs' was induced with prolonged salinity stress. The UPLC-MS analysis of the ET biosynthesis pathway rate-limiting semi-product, namely of 1-aminocyclopropane-1-carboxylic acid (ACC) content, confirmed the results assessed with molecular tools. The circadian rhythm of the ACC production of NaCl-treated semi-halophytes remained largely unaffected by the prolonged salinity stress episode. We speculate that the obtained results represent an image of the steady state established over the past 14 days, while during the first hours of the salinity stress response, the view could be completely different.

Plasticity of Plantago lanceolata L. in Adaptation to Extreme Environmental Conditions.

This study aimed at characterizing some adaptive changes in Plantago lanceolata L. exposed to harsh conditions of a desert-like environment generating physiological stress of limited water availability and exposure to strong light. It was clearly shown that the plants were capable of adapting their root system and vascular tissues to enable efficient vegetative performance. Soil analyses, as well as nitrogen isotope discrimination data show that P. lanceolata leaves in a desert-like environment had better access to nitrogen (nitrite/nitrate) and were able to fix it efficiently, as compared to the plants growing in the surrounding forest. The arbuscular mycorrhiza was also shown to be well-developed, and this was accompanied by higher bacterial frequency in the root zone, which might further stimulate plant growth. A closer look at the nitrogen content and leaf veins with a higher number of vessels and a greater vessel diameter made it possible to define the changes developed by the plants populating sandy habitats as compared with the vegetation sites located in the nearby forest. A determination of the photosynthesis parameters indicates that the photochemical apparatus in P. lanceolata inhabiting the desert areas adapted slightly to the desert-like environment and the time of day, with some changes of the reaction center (RC) size (photosystem II, PSII), while the plants' photochemical activity was at a similar level. No differences between the two groups of plants were observed in the dissipation of light energy. The exposure of plants to harsh conditions of a desert-like environment increased the water use efficiency (WUE) value in parallel with possible stimulation of the ß-carboxylation pathway.

Cadmium-Tolerant Rhizospheric Bacteria of the C3/CAM Intermediate Semi-Halophytic Common Ice Plant (Mesembryanthemum crystallinum L.) Grown in Contaminated Soils.

The common ice plant, Mesembryanthemum crystallinum L., has recently been found as a good candidate for phytoremediation of heavy-metal polluted soils. This semi-halophyte is a C3/CAM (Crassulacean acid metabolism) intermediate plant capable of tolerating extreme levels of cadmium in the soil. The aim of the work was to obtain and characterize novel, Cd-tolerant microbial strains that populate the root zone of M. crystallinum performing different types of photosynthetic metabolism and growing in Cd-contaminated substrates. The plants exhibiting either C3 or CAM photosynthesis were treated for 8 days with different CdCl2 doses to obtain final Cd concentrations ranging from 0.82 to 818 mg⋅kg-1 of soil d.w. The CAM phase was induced by highly saline conditions. After treatment, eighteen bacterial and three yeast strains were isolated from the rhizosphere and, after preliminary Cd-resistance in vitro test, five bacterial strains were selected and identified with a molecular proteomics technique. Two strains of the species Providencia rettgeri (W6 and W7) were obtained from the C3 phase and three (one Paenibacillus glucanolyticus S7 and two Rhodococcus erythropolis strains: S4 and S10) from the CAM performing plants. The isolates were further tested for Cd-resistance (treatment with either 1 mM or 10 mM CdCl2) and salinity tolerance (0.5 M NaCl) in model liquid cultures (incubation for 14 days). Providencia rettgeri W7 culture remained fully viable at 1 mM Cd, whereas Rh. erythropolis S4 and S10 together with P. glucanolyticus S7 were found to be resistant to 10 mM Cd in the presence of 0.5 M NaCl. It is suggested that the high tolerance of the common ice plant toward cadmium may result from the synergic action of the plant together with the Cd/salt-resistant strains occurring within rhizospheral microbiota. Moreover, the isolated bacteria appear as promising robust microorganisms for biotechnological applications in bio- and phytoremediation projects.

ROS-Scavengers, Osmoprotectants and Violaxanthin De-Epoxidation in Salt-Stressed Arabidopsis thaliana with Different Tocopherol Composition.

To determine the role of α- and γ-tocopherol (TC), this study compared the response to salt stress (200 mM NaCl) in wild type (WT) Arabidopsis thaliana (L.) Heynh. And its two mutants: (1) totally TC-deficient vte1; (2) vte4 accumulating γ-TC instead of α-TC; and (3) tmt transgenic line overaccumulating α-TC. Raman spectra revealed that salt-exposed α-TC accumulating plants were more flexible in regulating chlorophyll, carotenoid and polysaccharide levels than TC deficient mutants, while the plants overaccumulating γ-TC had the lowest levels of these biocompounds. Tocopherol composition and NaCl concentration affected xanthophyll cycle by changing the rate of violaxanthin de-epoxidation and zeaxanthin formation. NaCl treated plants with altered TC composition accumulated less oligosaccharides than WT plants. α-TC deficient plants increased their oligosaccharide levels and reduced maltose amount, while excessive accumulation of α-TC corresponded with enhanced amounts of maltose. Salt-stressed TC-deficient mutants and tmt transgenic line exhibited greater proline levels than WT plants, lower chlorogenic acid levels, and lower activity of catalase and peroxidases. α-TC accumulating plants produced more methylated proline- and glycine- betaines, and showed greater activity of superoxide dismutase than γ-TC deficient plants. Under salt stress, α-TC demonstrated a stronger regulatory effect on carbon- and nitrogen-related metabolites reorganization and modulation of antioxidant patterns than γ-TC. This suggested different links of α- and γ-TCs with various metabolic pathways via various functions and metabolic loops.

Exogenous Methyl Jasmonate Improves Cold Tolerance with Parallel Induction of Two Cold-Regulated (COR) Genes Expression in Triticum aestivum L.

Methyl jasmonate (MJ) is an important plant growth regulator that plays a key role in tolerance to biotic and abiotic stresses. In this research, the effects of exogenous MJ on cold tolerance, photosynthesis, activity and gene expression of antioxidant enzymes, proline accumulation, and expression of cold-regulated (COR) genes in wheat seedlings under low temperature (4 °C) were investigated. Exogenous MJ treatment (1 µM) promoted wheat cold tolerance before and during cold exposure. Low temperature significantly decreased photosynthetic parameters, whereas MJ application led to their partial recovery under cold exposure. Hydrogen peroxide (H2O2) and malondialdehyde (MDA) levels increased in response to low temperature, and this was counteracted by MJ application. Exogenous MJ significantly enhanced the activities of antioxidant enzymes and upregulated the expression of MnSOD and CAT during cold exposure. MJ application also led to enhanced proline content before 4 °C exposure, whereas the P5CS gene expression was upregulated by MJ's presence at both normal (22 °C) and low (4 °C) temperatures. It was also shown that MJ tended to upregulate the expression of the COR genes WCS19 and WCS120 genes. We conclude that exogenous MJ can alleviate the negative effect of cold stress thus increasing wheat cold tolerance.

At the Edges of Photosynthetic Metabolic Plasticity-On the Rapidity and Extent of Changes Accompanying Salinity Stress-Induced CAM Photosynthesis Withdrawal.

The common ice plant (Mesembryanthemum crystallinum L.) is a facultative crassulacean acid metabolism (CAM) plant, and its ability to recover from stress-induced CAM has been confirmed. We analysed the photosynthetic metabolism of this plant during the 72-h response period following salinity stress removal from three perspectives. In plants under salinity stress (CAM) we found a decline of the quantum efficiencies of PSII (Y(II)) and PSI (Y(I)) by 17% and 15%, respectively, and an increase in nonphotochemical quenching (NPQ) by almost 25% in comparison to untreated control. However, 48 h after salinity stress removal, the PSII and PSI efficiencies, specifically Y(II) and Y(I), elevated nonphotochemical quenching (NPQ) and donor side limitation of PSI (YND), were restored to the level observed in control (C3 plants). Swelling of the thylakoid membranes, as well as changes in starch grain quantity and size, have been found to be components of the salinity stress response in CAM plants. Salinity stress induced an over 3-fold increase in average starch area and over 50% decline of average seed number in comparison to untreated control. However, in plants withdrawn from salinity stress, during the first 24 h of recovery, we observed chloroplast ultrastructures closely resembling those found in intact (control) ice plants. Rapid changes in photosystem functionality and chloroplast ultrastructure were accompanied by the induction of the expression (within 24 h) of structural genes related to the PSI and PSII reaction centres, including PSAA, PSAB, PSBA (D1), PSBD (D2) and cp43. Our findings describe one of the most flexible photosynthetic metabolic pathways among facultative CAM plants and reveal the extent of the plasticity of the photosynthetic metabolism and related structures in the common ice plant.

Cooling effect of fungal stromata in the Dactylis-Epichloë-Botanophila symbiosis.

The stromata of Epichloë fungi are structures covering part of the stem of grasses. Under the fungal layer, still green tissues of the plant survive, although the development of the new leaves is inhibited. Stromata are the places where conidia and ascospores develop. Also, here Botanophila flies dine on mycelium, lay the eggs, defecate, and the larvae develop. The interaction of the three symbionts was analyzed concerning the organisms' adaptation to understand the differences in physiology and ecology of this microenvironment that support stable symbiosis spreading presently in Europe since the beginning of the XXI century. For analysis of the infrared radiation emitted by stromata, a high-resolution infrared camera FLIR E50 was used. The visualization of stromata temperature profiles was shown in the form of pseudo-colored (false) infrared images. The 13C discrimination was used to characterize photosynthesis of the plant tissue enclosed within the stromata. The stromata had a substantially lower temperature than the green plant tissues. The difference reached ~5.6°C during midday hours, whereas it was smaller in the evening, reaching only ~3.6°C. The mycelium of Epichloë cultivated on agar showed about 2°C lower temperature in comparison to the surrounding. The plant tissues enclosed within the stroma were photosynthetically active, although this activity was of phosphoenolpyruvate carboxylase (PEPC) type and less involved in heat dissipation during the day. The stromata, built by fungal hyphae, on which fungal reproductive structures develop, form a cool shelter. This shelter provides a place for the larvae of Botanophila flies.

Tocopherols mutual balance is a key player for maintaining Arabidopsis thaliana growth under salt stress.

Enhanced channeling carbon through pathways: shikimate/chorismate, benzenoid-phenylopropanoid or 2-C-methyl-D-erythritol 4-phosphate (MEP) provides a multitude of secondary metabolites and cell wall components and allows plants response to environmental stresses. Through the biosynthetic pathways, different secondary metabolites, like tocopherols (TCs), are bind to mutual dependencies and metabolic loops, that are not yet fully understood. We compared, in parallel, the influence of α- and γ-TCs on metabolites involved in osmoprotective/antioxidative response, and physico-chemical modification of plasma membrane and cell wall. We studied Arabidopsis thaliana Columbia ecotype (WT), mutant vte1 deficient in α- and γ-TCs, mutant vte4 over-accumulating γ-TC instead of α-TC, and transgenic line tmt over-accumulating α-TC; exposed to NaCl. The results indicate that salt stress activates ß-carboxylation processes in WT plants and in plants with altered TCs accumulation. In α-TC-deficient plants, NaCl causes ACC decrease, but does not change SA, whose concentration remains higher than in α-TC accumulating plants. α/γ-TCs contents influence carbohydrates, poliamines, phenolic (caffeic, ferrulic, cinnamic) acids accumulation patterns. Salinity results in increased detection of the LM5 galactan and LM19 homogalacturonan epitopes in α-TC accumulating plants, and the LM6 arabinan and MAC207 AGP epitopes in α-TC deficient mutants. Parallel, plants with altered TCs composition show decreased both the cell turgor and elastic modulus determined at the individual cell level. α-TC deficient plants reveal lower values of cell turgor and elastic modulus, but higher cell hydraulic conductivity than α-TC accumulating plants. Under salt stress, α-TC shows stronger regulatory effect than γ-TC through the impact on chloroplastic biosynthetic pathways and ROS/osmotic-modulating compounds.

The Common Ice Plant (Mesembryanthemum crystallinum L.)-Phytoremediation Potential for Cadmium and Chromate-Contaminated Soils.

The common ice plant (Mesembryanthemum crystallinum L.) is a widely studied model due to its tolerance to numerous biotic and abiotic stresses. In this study, carried out in model pots, the plants were treated with variant doses of Cd(II) and Cr(VI) and proved resistant to extreme levels of these heavy metals. Initial toxicity symptoms were observed upon final concentrations of 818 mg Cd kg-1 soil d.w., and 1699 mg Cr kg-1 applied as potassium chromate. Biometric analyses revealed that none of the Cr(VI) doses affected dry weight of the plant organs thus maintaining the shoot-to-root ratio. The Cd and Cr hypertolerance strategies were divergent and resulted in different accumulation patterns. For the case of Cd(II), an excluder-like mechanism was developed to prevent the plant from toxicity. For chromate, high accumulation potential together with Cr(VI) root-to-shoot translocation at sublethal concentrations was revealed (up to 6152 mg Cr kg-1 shoot at 4248 mg Cr kg-1 soil). It is concluded that M. crystallinum reveals considerable phytoremediation capabilities due to unique growth potential in contaminated substrates and is suitable for bioreclamation of degraded soils. The plant is especially applicable for efficient phytoextraction of chromate-contamination, whereas for Cd-affected areas it may have a phytostabilizing effect.

Antioxidative action of polyamines in protection of phospholipid membranes exposed to ozone stress.

In the present work, Langmuir monolayers were used to study the interaction of putrescine (a cationic antioxidant) with anionic charged membranes (1,2-dioleoyl-sn-glycerol-3-phosphate) under oxidative stress caused by the presence of ozone in the water phase. Calcium ions and acidic environment were used to compare the electrostatic and antioxidant effects of putrescine with those of an inorganic cation. It has been shown that the main role of putrescine in protecting systems against oxidation is its rapid reaction with ROS. The initial rate of ROS neutralization rose as the concentration of putrescine increased. No such reaction was observed for calcium ions. The consequence of putrescine's ozone removal was lesser lipid destruction that depended on the pH conditions.

Expression of Genes Involved in Heavy Metal Trafficking in Plants Exposed to Salinity Stress and Elevated Cd Concentrations.

Many areas intended for crop production suffer from the concomitant occurrence of heavy metal pollution and elevated salinity; therefore, halophytes seem to represent a promising perspective for the bioremediation of contaminated soils. In this study, the influence of Cd treatment (0.01-10.0 mM) and salinity stress (0.4 M NaCl) on the expression of genes involved in heavy metal uptake (irt2-iron-regulated protein 2, zip4-zinc-induced protein 4), vacuolar sequestration (abcc2-ATP-binding cassette 2, cax4-cation exchanger 2 pcs1-phytochelatin synthase 1) and translocation into aerial organs (hma4-heavy metal ATPase 4) were analyzed in a soil-grown semi-halophyte Mesembryanthemum crystallinum. The upregulation of irt2 expression induced by salinity was additionally enhanced by Cd treatment. Such changes were not observed for zip4. Stressor-induced alterations in abcc2, cax4, hma4 and pcs1 expression were most pronounced in the root tissue, and the expression of cax4, hma4 and pcs1 was upregulated in response to salinity and Cd. However, the cumulative effect of both stressors, similar to the one described for irt2, was observed only in the case of pcs1. The importance of salt stress in the irt2 expression regulation mechanism is proposed. To the best of our knowledge, this study is the first to report the combined effect of salinity and heavy metal stress on genes involved in heavy metal trafficking.

The response of a model C3/CAM intermediate semi-halophyte Mesembryanthemum crystallinum L. to elevated cadmium concentrations.

Many areas exhibiting increased concentrations of soluble salts are simultaneously polluted with heavy metals (HM), and halophytes with extended tolerance to heavy metal toxicity seem to represent a promising tool for their phytoremediation. In this study, the response of the soil-grown C3-CAM (Crassulacean acid metabolism) intermediate halophyte Mesembryanthemum crystallinum (common ice plant) to increased concentrations of Cd (0.01-1 mM) was investigated. None of the tested Cd treatments affected growth parameters or tissue water content of either C3 or CAM-performing plants. Chlorophyll a fluorescence confirmed high tolerance of the photosynthetic apparatus of both metabolic states towards Cd. Plants performing both photosynthesis types accumulated significant Cd amounts only under the highest (1 mM) treatment, and the metal was primarily deposited in the roots, which are features typical of an excluding strategy. Upon the application of 1 mM Cd solution CAM-performing plants, due to the NaCl pre-treatment applied for CAM induction, were exposed to significantly higher amounts of bioavailable Cd in comparison with those of C3-performing plants. As a result, roots of CAM plants accumulated over 4-fold higher Cd amounts when compared with C3 plants. In our opinion, enhanced Cd-accumulating potential observed in CAM-performing plants was the effect of osmotic stress episode and resulting modifications e.g. in the detoxifying capacity of the antioxidative system. Increased antioxidative potential of NaCl pre-treated plants was pronounced with significantly higher activity of CuZnSOD (copper-zinc superoxide dismutase), not achievable in C3 plants subjected to high Cd concentrations. Moreover, the applied Cd doses induced SOD activity in a compartment-dependent manner only in C3 plants. We confirmed that none of the applied Cd concentrations initiated the metabolic shift from C3 to CAM.

Spatial referencing of chlorophyll fluorescence images for quantitative assessment of infection propagation in leaves demonstrated on the ice plant: Botrytis cinerea pathosystem.

BACKGROUND: Chlorophyll fluorescence analysis is one of the non-invasive techniques widely used to detect and quantify the stress-induced changes in the photosynthetic apparatus. Quantitative information is obtained as a series of images and the specific fluorescence parameters are evaluated inside the regions of interest outlined separately on each leaf image. As the performance of photosynthesis is highly heterogeneous over a leaf surface, the areas of interest selected for generating numeric data are crucial for a reliable analysis. The differences in intact leaf physio-morphological characters and in the structural effects of stress between leaves increase the risk of artefacts. RESULTS: The authors propose a new enhanced method for precise assessment of stress-induced spatiotemporal changes in chlorophyll a fluorescence exemplified in the leaves of common ice plants infected with a fungal pathogen. The chl a fluorescence leaf image series obtained with Imaging-PAM fluorometer are aligned both by affine and nonlinear spline transforms based on the set of control points defined interactively. The successive readings were taken on the same leaf and this image sequence registration allows to capture quantitative changes of fluorescence parameters in time and along selected directions on the leaf surface. The time series fluorescence images of attached leaf, aligned according to the proposed method, provide a specific disease signature for an individual leaf. The results for C3 and Crassulacean Acid Metabolism (CAM) plants have been compared with respect to the type of photosynthetic metabolism and the image alignment accuracy has also been discussed. CONCLUSIONS: The image alignment applied to the series of fluorescence images allows to evaluate the dynamics of biotic stress propagation in individual plant leaves with better accuracy than previous methods. An important use of this method is the ability to map the fluorescence signal horizontally in one leaf during disease development and to accurately compare the results between leaves which differ in morphology or in the structural effects of stress. This approach in analysing chlorophyll fluorescence changes can be used to receive spatial and temporal information over a sample area in leaves infected by different pathogenic fungi and bacteria.

How to define obligatory anaerobiosis? An evolutionary view on the antioxidant response system and the early stages of the evolution of life on Earth.

One of the former definitions of "obligate anaerobiosis" was based on three main criteria: 1) it occurs in organisms, so-called obligate anaerobes, which live in environments without oxygen (O2), 2) O2-dependent (aerobic) respiration, and 3) antioxidant enzymes are absent in obligate anaerobes. In contrast, aerobes need O2 in order to grow and develop properly. Obligate (or strict) anaerobes belong to prokaryotic microorganisms from two domains, Bacteria and Archaea. A closer look at anaerobiosis covers a wide range of microorganisms that permanently or in a time-dependent manner tolerate different concentrations of O2 in their habitats. On this basis they can be classified as obligate/facultative anaerobes, microaerophiles and nanaerobes. Paradoxically, O2 tolerance in strict anaerobes is usually, as in aerobes, associated with the activity of the antioxidant response system, which involves different antioxidant enzymes responsible for removing excess reactive oxygen species (ROS). In our opinion, the traditional definition of "obligate anaerobiosis" loses its original sense. Strict anaerobiosis should only be restricted to the occurrence of O2-independent pathways involved in energy generation. For that reason, a term better than "obligate anaerobes" would be O2/ROS tolerant anaerobes, where the role of the O2/ROS detoxification system is separated from O2-independent metabolic pathways that supply energy. Ubiquitous key antioxidant enzymes like superoxide dismutase (SOD) and superoxide reductase (SOR) in contemporary obligate anaerobes might suggest that their origin is ancient, maybe even the beginning of the evolution of life on Earth. It cannot be ruled out that c. 3.5 Gyr ago, local microquantities of O2/ROS played a role in the evolution of the last universal common ancestor (LUCA) of all modern organisms. On the basis of data in the literature, the hypothesis that LUCA could be an O2/ROS tolerant anaerobe is discussed together with the question of the abiotic sources of O2/ROS and/or the early evolution of cyanobacteria that perform oxygenic photosynthesis.

Foliar application of selenium for protection against the first stages of mycotoxin infection of crop plant leaves.

BACKGROUND: The aim of this study was to investigate whether the application of selenium (Se) ions directly to the leaf surface can protect plants against infection by the fungal toxin zearalenone (ZEA). The experiments were performed for the most common and agronomically important crops such as wheat, oat, and barley (both tolerant and sensitive varieties) because mycotoxin accumulation in plants is the cause of many diseases in animals and people. RESULTS: ZEA at a concentration of 10 µmol L-1 either alone or in combination with Se (5 µmol L-1 Na2 SeO4 ) was applied to the second leaf of seedlings. Visualization of leaf temperature profiles by infrared thermography demonstrated a decrease in temperature at the location of ZEA infection that was more noticeable in sensitive genotypes. The presence of Se significantly suppressed changes at the site of ZEA application in all tested plants, especially the tolerant genotypes. Microscopic observations confirmed that foliar administration of ZEA resulted in its penetration to deeper localized cells and that damage induced by ZEA (mainly to chloroplasts) decreased after Se application. Analyses of antioxidant enzymes demonstrated the involvement of Se in antioxidation mechanisms, in particular by activating SOD and CAT under ZEA-induced stress conditions. CONCLUSION: The foliar application of Se to seedling leaves may be a non-invasive method of protecting crops against the first steps of ZEA infection. © 2018 Society of Chemical Industry.

Withdrawal from functional Crassulacean acid metabolism (CAM) is accompanied by changes in both gene expression and activity of antioxidative enzymes.

In Mesembryanthemum crystallinum, crassulacean acid metabolism (CAM) is seemingly reversible, but unequivocal evidence for functional CAM withdrawal has yet to be shown. In this study, we confirmed the rapid downregulation of PEPC1 expression just 1 h after the removal of NaCl from the plant growth media. At the same time, the Δ malate values in desalted plants rapidly (1 d) re-established to values typical for C3 plants. This phenomenon allowed us to confirm functional CAM withdrawal in the desalted plants. Desalting altered the expression of the genes of the main antioxidative enzymes and/or the activity of their respective proteins; for catalase (CAT), both gene expression and protein activity were restored to levels observed in C3 plants in response to desalting, while for cooper-zinc superoxide dismutase (CuZnSOD) and ascorbate peroxidase (APX), only protein activity was restored. Therefore, we conclude that during the C3→CAM transition the CAM-specific antioxidative enzyme activity profile constitutes a transient and fully reversible response to abiotic stress.

Seasonal variations of leaf chlorophyll-protein complexes in the wintergreen herbaceous plant Ajuga reptans L.

The chlorophyll and carotenoid content, and the spectra of low-temperature fluorescence of the leaves, chloroplasts and isolated pigment-protein complexes in the perennial herbaceous wintergreen plant Ajuga reptans L. (bugle) in different seasons of the year were studied. During winter, these plants downregulate photosynthesis and the PSA is reorganised, including the loss of chlorophyll, possible reductions in the number of functional reaction centres of PSII, and changes in aggregation of the thylakoid protein complexes. We also observed a restructuring of the PSI-PSII megacomplex and the PSII-light-harvesting complex II supercomplex in leaves covered by snow. After snowmelt, the monomeric form of the chl a/b pigment-protein complex associated with PSII (LHCII) and the free pigments were also detected. We expect that snow cover provides favourable conditions for keeping photosynthetic machinery ready for photosynthesis in spring just after snowmelt. During winter, the role of the zeaxanthin-dependent protective mechanism, which is responsible for the dissipation of excess absorbed light energy, is likely to increase.

Photosynthetic activity of vascular bundles in Plantago media leaves.

Photosynthetic processes in the leaf lamina and midribs of Plantago media were investigated using plants grown in high light (HL) or low light (LL) conditions. The fluorescence parameters, which indicate photochemical/photosynthetic activity, were different in HL and LL grown plants, but no major differences between lamina and midribs were found. An OJIP test (chlorophyll a fluorescence transient induction) of LL grown plants, indicative of the chloroplast electron transport chain, also showed both tissues to be similar. In HL plants, a partial blockage of electron flow between QA (the primary plastoquinone electron acceptor of PSII) and QB (the secondary plastoquinone acceptor of PSII) was found, and this was less visible in midribs. The effective dissipation of quantum energy per reaction center (DI0/RC) was similar in both tissues of HL grown plants, while in the midribs of LL leaves, this process seemed to be less effective. Measurements of 13C discrimination showed that the midrib tissues of LL and HL leaves effectively used ß-carboxylation products to accumulate their biomass. Thus, the well protected activity of electron transport in midribs with their limited capacity to fix CO2 from the air may indicate the involvement of this tissue in ß-carboxylation, transport or signaling. Carbon accumulated in roots showed a lower 13C discrimination value (more negative) than the values observed in lamina. This could indicate that roots are supplied with assimilates mostly during the light phase of the day cycle with intensive C3 photosynthesis.

Effects of exogenously applied hydrogen peroxide on antioxidant and osmoprotectant profiles and the C3-CAM shift in the halophyte Mesembryanthemum crystallinum L.

Exogenously applied H2O2 (50, 100 and 200mM) to Mesembryanthemum crystallinum root medium induced a transition from C3 to Crassulacean Acid Metabolism (CAM), as evaluated by diurnal malate (Δmal) fluctuations. A very high concentration of H2O2 (400mM) reduced Δmal below the value measured in control plants. An increase of malate content during the night in 400mM H2O2-treated plants might suggest that malate decarboxylation is crucial for CAM functioning. We conclude that malate plays a dual role: i) a protective and signaling function before CAM expression, and ii) a storage form of CO2 in plants performing CAM. A slight stimulation of photosystem II (PSII) photochemistry and net photosynthesis observed during the C3-CAM shift indicated that neither photoinhibition nor reduction of the photosynthetic rate were prerequisites for CAM. Moreover, CAM induction corresponded to a decrease of catalase activity. In CAM-performing plants, α-tocopherol, polyamines (putrescine and spermidine) and proline showed daily alterations and the content of α-tocopherol and polyamines was lower at the end of the day. In contrast, the proline concentration correlated with the applied H2O2 concentration and was higher at the end of the day in treated plants. The dynamic changes of antioxidant and osmolyte levels suggest their active role in preventing oxidative damage, stress acclimation mechanisms and involvement in metabolic regulation and/or signal transduction cascades.

Photosynthesis-related characteristics of the midrib and the interveinal lamina in leaves of the C3-CAM intermediate plant Mesembryanthemum crystallinum.

BACKGROUND AND AIMS: Leaf veins are usually encircled by specialized bundle sheath cells. In C4 plants, they play an important role in CO2 assimilation, and the photosynthetic activity is compartmentalized between the mesophyll and the bundle sheath. In C3 and CAM (Crassulacean acid metabolism) plants, the photosynthetic activity is generally attributed to the leaf mesophyll cells, and the vascular parenchymal cells are rarely considered for their role in photosynthesis. Recent studies demonstrate that enzymes required for C4 photosynthesis are also active in the veins of C3 plants, and their vascular system contains photosynthetically competent parenchyma cells. However, our understanding of photosynthesis in veins of C3 and CAM plants still remains insufficient. Here spatial analysis of photosynthesis-related properties were applied to the midrib and the interveinal lamina cells in leaves of Mesembryanthemum crystallinum, a C3-CAM intermediate plant. METHODS: The midrib anatomy as well as chloroplast structure and chlorophyll fluorescence, diurnal gas exchange profiles, the immunoblot patterns of PEPC (phosphoenolpyruvate carboxylase) and RubisCO (ribulose-1,5-bisphosphate carboxylase/oxygenase), H2O2 localization and antioxidant enzyme activities were compared in the midrib and in the interveinal mesophyll cells in leaves of C3 and CAM plants. KEY RESULTS: Leaf midribs were structurally competent to perform photosynthesis in C3 and CAM plants. The midrib chloroplasts resembled those in the bundle sheath cells of C4 plants and were characterized by limited photosynthetic activity. CONCLUSIONS: The metabolic roles of midrib chloroplasts differ in C3 and CAM plants. It is suggested that in leaves of C3 plants the midrib chloroplasts could be involved in the supply of CO2 for carboxylation, and in CAM plants they could provide malate to different metabolic processes and mediate H2O2 signalling.