Light Quality and Intensity Modulate Cold Acclimation in Arabidopsis.

Plant survival in temperate zones requires efficient cold acclimation, which is strongly affected by light and temperature signal crosstalk, which converge in modulation of hormonal responses. Cold under low light conditions affected Arabidopsis responses predominantly in apices, possibly because energy supplies were too limited for requirements of these meristematic tissues, despite a relatively high steady-state quantum yield. Comparing cold responses at optimal light intensity and low light, we found activation of similar defence mechanisms-apart from CBF1-3 and CRF3-4 pathways, also transient stimulation of cytokinin type-A response regulators, accompanied by fast transient increase of trans-zeatin in roots. Upregulated expression of components of strigolactone (and karrikin) signalling pathway indicated involvement of these phytohormones in cold responses. Impaired response of phyA, phyB, cry1 and cry2 mutants reflected participation of these photoreceptors in acquiring freezing tolerance (especially cryptochrome CRY1 at optimal light intensity and phytochrome PHYA at low light). Efficient cold acclimation at optimal light was associated with upregulation of trans-zeatin in leaves and roots, while at low light, cytokinin (except cis-zeatin) content remained diminished. Cold stresses induced elevation of jasmonic acid and salicylic acid (in roots). Low light at optimal conditions resulted in strong suppression of cytokinins, jasmonic and salicylic acid.

Plant Abiotic Stress Proteomics: The Major Factors Determining Alterations in Cellular Proteome.

HIGHLIGHTS: Major environmental and genetic factors determining stress-related protein abundance are discussed.Major aspects of protein biological function including protein isoforms and PTMs, cellular localization and protein interactions are discussed.Functional diversity of protein isoforms and PTMs is discussed. Abiotic stresses reveal profound impacts on plant proteomes including alterations in protein relative abundance, cellular localization, post-transcriptional and post-translational modifications (PTMs), protein interactions with other protein partners, and, finally, protein biological functions. The main aim of the present review is to discuss the major factors determining stress-related protein accumulation and their final biological functions. A dynamics of stress response including stress acclimation to altered ambient conditions and recovery after the stress treatment is discussed. The results of proteomic studies aimed at a comparison of stress response in plant genotypes differing in stress adaptability reveal constitutively enhanced levels of several stress-related proteins (protective proteins, chaperones, ROS scavenging- and detoxification-related enzymes) in the tolerant genotypes with respect to the susceptible ones. Tolerant genotypes can efficiently adjust energy metabolism to enhanced needs during stress acclimation. Stress tolerance vs. stress susceptibility are relative terms which can reflect different stress-coping strategies depending on the given stress treatment. The role of differential protein isoforms and PTMs with respect to their biological functions in different physiological constraints (cellular compartments and interacting partners) is discussed. The importance of protein functional studies following high-throughput proteome analyses is presented in a broader context of plant biology. In summary, the manuscript tries to provide an overview of the major factors which have to be considered when interpreting data from proteomic studies on stress-treated plants.

Proteomic Response of Hordeum vulgare cv. Tadmor and Hordeum marinum to Salinity Stress: Similarities and Differences between a Glycophyte and a Halophyte.

Response to a high salinity treatment of 300 mM NaCl was studied in a cultivated barley Hordeum vulgare Syrian cultivar Tadmor and in a halophytic wild barley H. marinum. Differential salinity tolerance of H. marinum and H. vulgare is underlied by qualitative and quantitative differences in proteins involved in a variety of biological processes. The major aim was to identify proteins underlying differential salinity tolerance between the two barley species. Analyses of plant water content, osmotic potential and accumulation of proline and dehydrin proteins under high salinity revealed a relatively higher water saturation deficit in H. marinum than in H. vulgare while H. vulgare had lower osmotic potential corresponding with high levels of proline and dehydrins. Analysis of proteins soluble upon boiling isolated from control and salt-treated crown tissues revealed similarities as well as differences between H. marinum and H. vulgare. The similar salinity responses of both barley species lie in enhanced levels of stress-protective proteins such as defense-related proteins from late-embryogenesis abundant family, several chaperones from heat shock protein family, and others such as GrpE. However, there have also been found significant differences between H. marinum and H. vulgare salinity response indicating an active stress acclimation in H. marinum while stress damage in H. vulgare. An active acclimation to high salinity in H. marinum is underlined by enhanced levels of several stress-responsive transcription factors from basic leucine zipper and nascent polypeptide-associated complex families. In salt-treated H. marinum, enhanced levels of proteins involved in energy metabolism such as glycolysis, ATP metabolism, and photosynthesis-related proteins indicate an active acclimation to enhanced energy requirements during an establishment of novel plant homeostasis. In contrast, changes at proteome level in salt-treated H. vulgare indicate plant tissue damage as revealed by enhanced levels of proteins involved in proteasome-dependent protein degradation and proteins related to apoptosis. The results of proteomic analysis clearly indicate differential responses to high salinity and provide more profound insight into biological mechanisms underlying salinity response between two barley species with contrasting salinity tolerance.

Global Scale Transcriptional Profiling of Two Contrasting Barley Genotypes Exposed to Moderate Drought Conditions: Contribution of Leaves and Crowns to Water Shortage Coping Strategies.

Drought is a serious threat for sustainable agriculture. Barley represents a species well adapted to environmental stresses including drought. To elucidate the adaptive mechanism of barley on transcriptional level we evaluated transcriptomic changes of two contrasting barley cultivars upon drought using the microarray technique on the level of leaves and crowns. Using bioinformatic tools, differentially expressed genes in treated vs. non-treated plants were identified. Both genotypes revealed tissue dehydration under drought conditions as shown at water saturation deficit and osmotic potential data; however, dehydration was more severe in Amulet than in drought-resistant Tadmor under the same ambient conditions. Performed analysis showed that Amulet enhanced expression of genes related to active plant growth and development, while Tadmor regarding the stimulated genes revealed conservative, water saving strategy. Common reactions of both genotypes and tissues included an induction of genes encoding several stress-responsive signaling proteins, transcription factors as well as effector genes encoding proteins directly involved in stress acclimation. In leaf, tolerant cultivar effectively stimulated mainly the expression of genes encoding proteins and enzymes involved in protein folding, sulfur metabolism, ROS detoxification or lipid biosynthesis and transport. The crown specific reaction of tolerant cultivar was an enhanced expression of genes encoding proteins and enzymes involved in cell wall lignification, ABRE-dependent abscisic acid (ABA) signaling, nucleosome remodeling, along with genes for numerous jasmonate induced proteins.

Quantitative analysis of proteome extracted from barley crowns grown under different drought conditions.

Barley cultivar Amulet was used to study the quantitative proteome changes through different drought conditions utilizing two-dimensional difference gel electrophoresis (2D-DIGE). Plants were cultivated for 10 days under different drought conditions. To obtain control and differentially drought-treated plants, the soil water content was kept at 65, 35, and 30% of soil water capacity (SWC), respectively. Osmotic potential, water saturation deficit, (13)C discrimination, and dehydrin accumulation were monitored during sampling of the crowns for proteome analysis. Analysis of the 2D-DIGE gels revealed 105 differentially abundant spots; most were differentially abundant between the controls and drought-treated plants, and 25 spots displayed changes between both drought conditions. Seventy-six protein spots were successfully identified by tandem mass spectrometry. The most frequent functional categories of the identified proteins can be put into the groups of: stress-associated proteins, amino acid metabolism, carbohydrate metabolism, as well as DNA and RNA regulation and processing. Their possible role in the response of barley to drought stress is discussed. Our study has shown that under drought conditions barley cv. Amulet decreased its growth and developmental rates, displayed a shift from aerobic to anaerobic metabolism, and exhibited increased levels of several protective proteins. Comparison of the two drought treatments revealed plant acclimation to milder drought (35% SWC); but plant damage under more severe drought treatment (30% SWC). The results obtained revealed that cv. Amulet is sensitive to drought stress. Additionally, four spots revealing a continuous and significant increase with decreasing SWC (UDP-glucose 6-dehydrogenase, glutathione peroxidase, and two non-identified) could be good candidates for testing of their protein phenotyping capacity together with proteins that were significantly distinguished in both drought treatments.

Wheat and barley dehydrins under cold, drought, and salinity - what can LEA-II proteins tell us about plant stress response?

Dehydrins as a group of late embryogenesis abundant II proteins represent important dehydration-inducible proteins whose accumulation is induced by developmental processes (embryo maturation) as well as by several abiotic stress factors (low temperatures, drought, salinity). In the review, an overview of studies aimed at investigation of dehydrin accumulation patterns at transcript and protein levels as well as their possible functions in common wheat (Triticum aestivum), durum wheat (T. durum), and barley (Hordeum vulgare) plants exposed to various abiotic stress factors (cold, frost, drought, salinity) is provided. Possible roles of dehydrin proteins in an acquisition and maintenance of an enhanced frost tolerance are analyzed in the context of plant developmental processes (vernalization). Quantitative and qualitative differences as well as post-translational modifications in accumulated dehydrin proteins between barley cultivars revealing differential tolerance to drought and salinity are also discussed. Current knowledge on dehydrin role in wheat and barley response to major dehydrative stresses is summarized and the major challenges in dehydrin research are outlined.

Proteomics of stress responses in wheat and barley-search for potential protein markers of stress tolerance.

Wheat (Triticum aestivum; T. durum) and barley (Hordeum vulgare) agricultural production is severely limited by various abiotic and biotic stress factors. Proteins are directly involved in plant stress response so it is important to study proteome changes under various stress conditions. Generally, both abiotic and biotic stress factors induce profound alterations in protein network covering signaling, energy metabolism (glycolysis, Krebs cycle, ATP biosynthesis, photosynthesis), storage proteins, protein metabolism, several other biosynthetic pathways (e.g., S-adenosylmethionine metabolism, lignin metabolism), transport proteins, proteins involved in protein folding and chaperone activities, other protective proteins (LEA, PR proteins), ROS scavenging enzymes as well as proteins affecting regulation of plant growth and development. Proteins which have been reported to reveal significant differences in their relative abundance or posttranslational modifications between wheat, barley or related species genotypes under stress conditions are listed and their potential role in underlying the differential stress response is discussed. In conclusion, potential future roles of the results of proteomic studies in practical applications such as breeding for an enhanced stress tolerance and the possibilities to test and use protein markers in the breeding are suggested.

Effect of heat stress on polyamine metabolism in proline-over-producing tobacco plants.

The effect of heat stress on the accumulation of proline and on the level of polyamines (PAs) in tobacco plants was investigated. Responses to heat stress were compared in the upper and lower leaves and roots of tobacco plants that constitutively over-express a modified gene for the proline biosynthetic enzyme Δ1-pyrroline-5-carboxylate synthetase (P5CSF129A) and in the corresponding wild-type. In the initial phases of heat stress (after 2h at 40°C), the accumulation of proline increased in the wild type but slightly decreased in the transformants. The response to heat stress in proline-over-producing tobacco plants involved a transient increase in the levels of free and conjugated putrescine (Put) and in the levels of free spermidine (Spd), norspermidine (N-Spd) and spermine (Spm) after a 2-h lag phase, which correlated with stimulation of the activity of the corresponding biosynthetic enzymes. Diamine oxidase (DAO) activity increased in both plant genotypes, most significantly in the leaves of WT plants. Polyamine oxidase (PAO) activity increased in the roots of WT plants and decreased in the leaves and roots of the transformants. After 6h of heat stress, proline accumulation was observed in the transformants, especially in the lower leaves; much more modest increase was observed in the WT plants. A decrease in the levels of free and conjugated Put coincided with down-regulation of the activity of ornithine decarboxylase and marked stimulation of DAO activity in the leaves and roots of the transformants. PAO activity increased in the roots of the transformants but decreased in the leaves. Conversely, in WT tobacco subjected to 6h of heat stress, slight increases in free and conjugated PA levels were observed and the activity of DAO only increased in the roots; PAO activity did not change from the value observed during the initial phase of heat stress. 6 Hours' heat stress had no effect on the level of malondialdehyde (MDA; a product of lipid peroxidation), in the upper leaves of either genotype. After a recovery period (2h at 25°C), most of the studied parameters exhibited values comparable to those observed in untreated plants. The coordination of the proline and polyamine biosynthetic pathways during heat stress conditions is discussed.

Analysis of proteome and frost tolerance in chromosome 5A and 5B reciprocal substitution lines between two winter wheats during long-term cold acclimation.

Dynamics of cold tolerance and crown proteome composition has been analysed in a set of two winter wheat cultivars Mironovskaya 808 and Bezostaya 1 and four reciprocal substitution lines with interchanged chromosomes 5A and 5B during a long-term cold-acclimation (CA) treatment. Proteome analysis has revealed 298 differently abundant spots during experiment. Most of them (260) were changed due to CA process and only 52 spots displayed differences between genotypes. Two hundred and seven protein spots were successfully identified by tandem mass spectrometry. Comparison of samples before and after vernalization fulfillment by a combination of ANOVA and Student' T-test displayed ten differentially abundant protein spots (e.g. chopper chaperones). However, differences in the accumulation of these spots did not reflect differences in vernalization requirement of genotypes. Therefore, our results indicate that vernalization process has not influenced total proteome of CA wheat crowns. A few protein spots (14 spots; e.g. malate dehydrogenase) revealed differential accumulation levels between the individual genotypes or their groups possessing chromosome 5A or 5B from Mironovskaya 808 versus Bezostaya 1. The study has shown the effect of chromosome 5A on physiological traits and also proteome in winter wheat. Putative candidate protein markers for cold tolerance in wheat are discussed.

Comparison of hormonal responses to heat, drought and combined stress in tobacco plants with elevated proline content.

In order to test the possibility of improving tolerance to heat and drought (alone and in combination) by elevation of the osmoprotectant proline (Pro) content, stress responses were compared in tobacco plants constitutively over-expressing a gene for the Pro biosynthetic enzyme Δ(2)-pyrroline-5-carboxylate synthetase (P5CSF129A; EC 2.7.2.11/1.2.1.41) and in the corresponding wild-type. Significantly enhanced Pro production in the transformant coincided with a more negative leaf osmotic potential (both at control conditions and following stress) and enhanced production of protective xanthophyll cycle pigments. Heat stress (40 °C) caused a transient increase in the level of bioactive cytokinins, predominantly N(6)-(2-isopentenyl)adenosine, accompanied by down-regulation of the activity of the main cytokinin degrading enzyme cytokinin oxidase/dehydrogenase (EC 1.4.3.18/1.5.99.12). No significant differences were found between the tested genotypes. In parallel, a transient decrease of abscisic acid was observed. Following drought stress, bioactive cytokinin levels decreased in the whole plants, remaining relatively higher in preferentially protected upper leaves and in roots. Cytokinin suppression was less pronounced in Pro transformants. Exposure to heat stress (40 °C for 2h) at the end of 10-d drought period strongly enhanced the severity of the water stress, as indicated by a drop in leaf water potential. The activity of cytokinin oxidase/dehydrogenase was strongly stimulated in upper leaves and roots of stressed plants, coinciding with strong down-regulation of bioactive cytokinins in whole plants. Combined heat and drought stress resulted in a minor decrease in abscisic acid, but only in non-wilty upper leaves. Both stresses as well as their combination were associated with elevation of free auxin, indolylacetic acid, in lower leaves and/or in roots. Auxin increase was dependent on the stress strength. After rehydration, a marked elevation of bioactive cytokinins in leaves was observed. A greater increase in cytokinin content in Pro transformants indicated a mild elevation of their stress tolerance.