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In nature, plants are exposed to a range of climatic conditions. Those negatively impacting plant growth and survival are called abiotic stresses. Although abiotic stresses have been extensively studied separately, little is known about their interactions. Here, we investigate the impact of long-term mild metal exposure on the cold acclimation of Salix viminalis roots using physiological, transcriptomic, and proteomic approaches. We found that, while metal exposure significantly affected plant morphology and physiology, it did not impede cold acclimation. Cold acclimation alone increased glutathione content and glutathione reductase activity. It also resulted in the increase in transcripts and proteins belonging to the heat-shock proteins and related to the energy metabolism. Exposure to metals decreased antioxidant capacity but increased catalase and superoxide dismutase activity. It also resulted in the overexpression of transcripts and proteins related to metal homeostasis, protein folding, and the antioxidant machinery. The simultaneous exposure to both stressors resulted in effects that were not the simple addition of the effects of both stressors taken separately. At the antioxidant level, the response to both stressors was like the response to metals alone. While this should have led to a reduction of frost tolerance, this was not observed. The impact of the simultaneous exposure to metals and cold acclimation on the transcriptome was unique, while at the proteomic level the cold acclimation component seemed to be dominant. Some genes and proteins displayed positive interaction patterns. These genes and proteins were related to the mitigation and reparation of oxidative damage, sugar catabolism, and the production of lignans, trehalose, and raffinose. Interestingly, none of these genes and proteins belonged to the traditional ROS homeostasis system. These results highlight the importance of the under-studied role of lignans and the ROS damage repair and removal system in plants simultaneously exposed to multiple stressors. [ABSTRACT FROM AUTHOR]

Abstract: The response of two willow clones (Salix fragilis (Sf) and Salix aurita (Sa)) to the presence of metals (Zn, Cu, Cd, Ni) was studied. Rooted cuttings were planted in control and contaminated soil. After 100days, different parameters (biomass, chlorophyll fluorescence (Fv/Fm), pigment and sugar concentrations, electrolyte leakage and proteome-level changes) were analyzed. The growth of Sa was not influenced by metals whereas Sf produced significantly less biomass when exposed to the pollutants. Furthermore, although Sa did not show a growth reduction in the presence of metals, the overall view of the physiological results among others the changes in the accumulation of sugars and pigments indicated that metals had a more severe impact on this clone. The response at the proteome level confirmed these observations. The growth reduction and the proteomic changes in Sf indicate that this clone adjusts its metabolism to maintain cellular homeostasis. Sa on the contrary maintains growth but the physiological and proteomics data suggests that this can only be done at the cost of cellular deregulation. Therefore high biomass is not linked with a good tolerance strategy. In a long-term study the survival of Sa might be compromised making it a poorer candidate for phytoremediation efforts. Biological significance: In the last centuries human activity has resulted in the dispersal of heavy metals with potential phytotoxic effects over large areas. The increased knowledge of the responses of Salix-species, a group of trees with potential as biomass producer but also as phytoremediation agent, when growing on metal-polluted substrate provided by this study has the potential to help in the improved selection of clones with more or less potential for these aims. Contrary to most studies the trees in the current study were exposed to a mixture of metals, thereby facing a closer resemblance to the situation on soils polluted by human activity. Whereas many papers focused on the two main phenotypic characteristics (biomass and accumulation), fewer papers studied proteomic and physiological parameters which allow to have a global view of the tolerance of probable willow candidates for phytoremediation purposes. Our data demonstrates that higher biomass production in presence of metals is not necessarily linked with higher tolerance whereas growth reduction might indicate longer long-term tolerance. In the long term and in the purpose of a future use in phytoremediation, the survival of this high producer clone could be compromised. [Copyright &y& Elsevier]

Abstract: The key to successful cryopreservation lies in the induction of tolerance towards dehydration/desiccation and freezing. The accumulation of osmo-active compounds, which can be induced by drought and cold stress, is therefore important. In the present study, three-week old shoots from in vitro plantlets of the cultivated potato Solanum tuberosum and its frost-resistant relative Solanum commersonii were submitted to osmotic stress (by using sucrose) and chilling (6°C). After 14days of exposure, shoot tips were sampled in order to gain an insight into changes of the proteome and soluble sugars. Also, the effect of these treatments on growth performance behaviour and on the success of cryopreservation was evaluated. Identified proteins that changed in abundance due to stress were associated with stress response. Additionally, carbohydrate analyses in both species, after exposure to chilling, also indicated species-related differences; this observation could point towards a better-adapted physiological state of the donor plants of S. commersonii prior to the cryoprocedure and therefore a better recovery of the meristems. Biological significance: To our knowledge, this is the first study in which cryopreservation experiments are combined with the observation of the responses to abiotic stress exposure involving the potato species S. commersonii and S. tuberosum. These two species are known to have a different cold-acclimation behaviour, which seems to be closely related to their tolerance towards cryopreservation. Furthermore, common and differential responses to abiotic stresses were observed in the two species indicating that some pathways could be crucial not only in the plant's response to stress but also in tolerance towards cryopreservation. [Copyright &y& Elsevier]

Better knowledge on responses to dehydration stress could help to improve the existing cryopreservation protocols for potato, since plant tissues processed for cryopreservation are often submitted to similar in vitro stress conditions. Cryopreservation (the best method of conservation for vegetatively propagated plants) of potato still needs to be standardized to make it available and to conserve the wide diversity of this crop. In the present work, the response to osmotic stress and chilling temperature was investigated in two potato species, Solanum tuberosum and its relative, frost-tolerant S. commersonii. After 14 days of exposure, different growth parameters, such as shoot length and number of leaves, were measured. Furthermore, differentially abundant proteins were identified after performing 2-fluorescence difference gel electrophoresis (2-DIGE) experiments, and soluble carbohydrates were analyzed by High Performance Anion Exchange Chromatography with Pulsed Amperometric Detection (HPAEC-PAD). The results show different responses in both species depending on the stress treatment. Focusing on the differences in growth parameters during the treatments, Solanum commersonii seems to be more affected than S. tuberosum cv. Désirée. At the molecular level, there are some differences and similarities between the two potato species studied that are dependent on the type of stressor. [ABSTRACT FROM AUTHOR]

Abstract: The forest ecosystem is of particular importance from an economic and ecological perspective. However, the stress physiology of trees, perennial and woody plants, is far from being fully understood. For that purpose, poplar plants were exposed to drought; the plants exhibited commonly reported drought stress traits in the different plant tissues. Leafy rooted cuttings of poplar were investigated through a proteomic approach in order to compare the water constraint response of two plant tissues, namely leaf and cambium. Sampling was realized during the drought period at 2 time points with increased drought intensity and 7days after rewatering. Our data show that there is a difference in the moment of response to the water constraint between the two tissues, cambium being affected later than leaves. In leaves, drought induced a decrease in rubisco content, and an increase in the abundance of light harvesting complex proteins as well as changes in membrane-related proteins. In the cambial tissue, the salient proteome pattern change was the decrease of multiple proteins identified as bark storage proteins. After rewatering, almost all changes in cambial proteome disappeared whereas a significant number of leaf proteins appeared to be differentially regulated only during the recovery from drought. [Copyright &y& Elsevier]

Salix viminalis is a fast growing willow species with potential as a plant used for biomass feedstock or for phytoremediation. However, few reference genes (RGs) for quantitative real-time polymerase chain reaction (qPCR) are available in S. viminalis, thereby limiting gene expression studies. Here, we investigated the expression stability of 14 candidate reference genes (RGs) across various organs exposed to five abiotic stresses (cold, heat, drought, salt, and poly-metals). Four RGs ranking algorithms, namely geNormPLUS, BestKeeper, NormFinder, and GrayNorm were applied to analyze the qPCR data and the outputs were merged into consensus lists with RankAggreg, a rank aggregation algorithm. In addition, the optimal RG combinations were determined with geNormPLUS and GrayNorm. The genes that were the most stable in the roots were TIP41 and CDC2. In the leaves, TIP41 was the most stable, followed by EF1b and ARI8, depending on the condition tested. Conversely, GAPDH and β-TUB, two genes commonly used for qPCR data normalization were the least stable across all organs. Nevertheless, both geNormPLUS and GrayNorm recommended the use of a combination of genes rather than a single one. These results are valuable for research of transcriptomic responses in different S. viminalis organs. [ABSTRACT FROM AUTHOR]