Root proteomics reveals cucumber 24-epibrassinolide responses under Ca(NO<Subscript>3</Subscript>)<Subscript>2</Subscript> stress

Key message

The application of exogenous 24-epibrassinolide promotes Brassinosteroids intracellular signalling in cucumber, which leads to differentially expressed proteins that participate in different life process to relieve Ca(NO ₃ ) ₂ damage.

Abstract

NO₃ ^? and Ca²⁺ are the main anion and cation of soil secondary salinization during greenhouse cultivation. Brassinosteroids (BRs), steroidal phytohormones, regulate various important physiological and developmental processes and are used against abiotic stress. A two-dimensional electrophoresis gel coupled with MALDI-TOF/TOF MS was performed to investigate the effects of exogenous 24-epibrassinolide (EBL) on proteomic changes in cucumber seedling roots under Ca(NO₃)₂ stress. A total of 80 differentially accumulated protein spots in response to stress and/or exogenous EBL were identified and grouped into different categories of biological processes according to Gene Ontology. Under Ca(NO₃)₂ stress, proteins related to nitrogen metabolism and lignin biosynthesis were induced, while those related to cytoskeleton organization and cell-wall neutral sugar metabolism were inhibited. However, the accumulation of abundant proteins involved in protein modification and degradation, defence mechanisms against antioxidation and detoxification and lignin biosynthesis by exogenous EBL might play important roles in salt tolerance. Real-time quantitative PCR was performed to investigate BR signalling. BR signalling was induced intracellularly under Ca(NO₃)₂ stress. Exogenous EBL can alleviate the root indices, effectively reduce the Ca²⁺ content and increase the K⁺ content in cucumber roots under Ca(NO₃)₂ stress. This study revealed the differentially expressed proteins and BR signalling-associated mRNAs induced by EBL in cucumber seedling roots under Ca(NO₃)₂ stress, providing a better understanding of EBL-induced salt resistance in cucumber seedlings. The mechanism for alleviation provides valuable insight into improving Ca(NO₃)₂ stress tolerance of other horticultural plants.

     

High-mass-resolution MALDI mass spectrometry imaging reveals detailed spatial distribution of metabolites and lipids in roots of barley seedlings in response to salinity stress

Introduction

Mass spectrometry imaging (MSI) is a technology that enables the visualization of the spatial distribution of hundreds to thousands of metabolites in the same tissue section simultaneously. Roots are below-ground plant organs that anchor plants to the soil, take up water and nutrients, and sense and respond to external stresses. Physiological responses to salinity are multifaceted and have predominantly been studied using whole plant tissues that cannot resolve plant salinity responses spatially.

Objectives

This study aimed to use a comprehensive approach to study the spatial distribution and profiles of metabolites, and to quantify the changes in the elemental content in young developing barley seminal roots before and after salinity stress.

Methods

Here, we used a combination of liquid chromatography–mass spectrometry (LC–MS), inductively coupled plasma mass spectrometry (ICP–MS), and matrix-assisted laser desorption/ionization (MALDI–MSI) platforms to profile and analyze the spatial distribution of ions, metabolites and lipids across three anatomically different barley root zones before and after a short-term salinity stress (150 mM NaCl).

Results

We localized, visualized and discriminated compounds in fine detail along longitudinal root sections and compared ion, metabolite, and lipid composition before and after salt stress. Large changes in the phosphatidylcholine (PC) profiles were observed as a response to salt stress with PC 34:n showing an overall reduction in salt treated roots. ICP–MS analysis quantified changes in the elemental content of roots with increases of Na⁺ and decreases of K⁺ content.

Conclusion

Our results established the suitability of combining three mass spectrometry platforms to analyze and map ionic and metabolic responses to salinity stress in plant roots and to elucidate tolerance mechanisms in response to abiotic stress, such as salinity stress.

     

Involvement of nitric oxide-mediated alternative pathway in tolerance of wheat to drought stress by optimizing photosynthesis

Key message

NO-mediated alternative pathway plays an important role in protecting wheat seedlings against drought stress through dissipating excessive reducing equivalents generated by photosynthesis.

Abstract

Alternative pathway (AP) has been proven to be involved in responses to various stresses. However, the mechanisms of AP in defense response to drought stress are still lacking. The aims of this work are to investigate the role of AP in drought tolerance and how AP is induced under drought stress using two wheat cultivars with different drought tolerance. Our results showed that Longchun22 cultivar is more tolerant to drought than 98SN146 cultivar. Seedlings exposed to drought led to a significant increase in AP, and it increased more in Longchun22. Furthermore, chlorophyll fluorescence parameters (Fv/Fm, ΦPSII, qP) decreased significantly in drought-treated seedlings, especially in 98SN146, indicating that photoinhibition occurred under drought stress. Pretreatment with SHAM, the malate–oxaloacetate shuttle activity and photosynthetic efficiency were further inhibited in drought-treated seedlings, resulting in more serious oxidative damage as indicated by higher levels of malondialdehyde and hydrogen peroxide. Moreover, NO modulated AP under drought stress by increasing AOX1a expression and pyruvate content. Taken together, these results indicate that NO-mediated AP is involved in optimizing photosynthesis under drought stress by avoiding the over-reduction of photosynthetic electron transport chain, thus reducing reactive oxygen species production and oxidative damage in wheat leaves.

     

Association mapping of drought tolerance-related traits in barley to complement a traditional biparental QTL mapping study

Key message

Association mapping of drought-related traits in barley was used to increase the density of existing QTL maps without recreating mapping populations.

Abstract

We used 109 spring barley genotypes exhibiting high or low drought tolerance to elucidate the associations between diversity array technology sequencing (DArTseq) and single nucleotide polymorphism (SNP) markers and various physiological parameters related to plant responses to drought conditions. The study was performed in controlled conditions (growth chambers), drought tolerance was phenotyped in the four-leaf seedlings. We identified 58 associations including 34 new markers (i.e., 16 DArTseq and 18 SNP markers). The results for three markers were consistent with the data obtained in an earlier traditional biparental QTL mapping study. The regions neighboring markers on linkage group 2H contained the highest number of significant marker–trait associations. Five markers related to the photosynthetic activity of photosystem II were detected on chromosome 4H. The lowest number of associations were observed for the sequences neighboring DArT markers on linkage group 6H. A chromosome 3H region related to water use efficiency and net photosynthesis rate in both biparental QTL, and association study, may be particularly valuable, as these parameters correspond to the ability of plants to remain highly productive under water deficit stress. Our findings confirm that association mapping can increase the density of existing QTL maps without recreating mapping populations.

     

Effects of heterogeneous salinity on growth,water uptake,and tissue ion concentrations of alfalfa

Aims

Soil salinity varies greatly in the plant rhizosphere. The effect of nonuniform salinity on the growth and physiology response of alfalfa plants was determined to improve understanding of salt stress tolerance mechanisms of alfalfa.

Methods

Plant growth, predawn leaf water potential, water uptake, and tissue ionic content were studied in alfalfa plants grown hydroponically for 9 days using a split-root system, with uniform salinity or horizontally nonuniform salinity treatments (0/S, 75/S, and 150/S corresponding to 0, 75, and 150 mM NaCl on the low salt side, respectively).

Results

Compared with uniform high salinity, 0/S and 75/S treatments significantly increased the alfalfa shoot dry mass and stem extension rate. Compensatory water uptake by low salt roots of 0/S and 75/S treatments was observed. However, decreased leaf Na⁺ concentration, increased leaf K⁺/Na⁺, and compensatory growth of roots on the low salt side were observed only following the 0/S treatment.

Conclusions

Nonuniform salinity dose not enhance plant growth once a threshold NaCl concentration in low salinity growth medium has been reached. Compensation of water uptake from the low-salt root zone and regulation of K⁺/Na⁺ homeostasis in low salt root play more important role than regulation of leaf ions in enhancing alfalfa growth under nonuniform salinity.

     

MdSOS2L1 phosphorylates MdVHA-B1 to modulate malate accumulation in response to salinity in apple

Key message

Salt-induced phosphorylation of MdVHA-B1 protein was mediated by MdSOS2L1 protein kinase, and thereby increasing malate content in apple.

Abstract

Salinity is an important environmental factor that influences malate accumulation in apple. However, the molecular mechanism by which salinity regulates this process is poorly understood. In this work, we found that MdSOS2L1, a novel AtSOS2-LIKE protein kinase, interacts with V-ATPase subunit MdVHA-B1. Furthermore, MdSOS2L1 directly phosphorylates MdVHA-B1 at Ser³⁹⁶ site to modulate malate accumulation in response to salt stress. Meanwhile, a series of transgenic analyses in apple calli showed that the MdSOS2L1–MdVHAB1 pathway was involved in the regulation of malate accumulation. Finally, a viral vector-based transformation approach demonstrated that the MdSOS2L1–MdVHAB1 pathway also modulated malate accumulation in apple fruits with or without salt stress. Collectively, our findings provide a new insight into the mechanism by which MdSOS2L1 phosphorylates MdVHA-B1 to modulate malate accumulation in response to salinity in apple.

     

Ectopic expression of phloem motor protein pea forisome PsSEO-F1 enhances salinity stress tolerance in tobacco

Key message

PsSEOF-1 binds to calcium and its expression is upregulated by salinity treatment. PsSEOF - 1 -overexpressing transgenic tobacco showed enhanced salinity stress tolerance by maintaining cellular ion homeostasis and modulating ROS-scavenging pathway.

Abstract

Calcium (Ca²⁺) plays important role in growth, development and stress tolerance in plants. Cellular Ca²⁺ homeostasis is achieved by the collective action of channels, pumps, antiporters and by Ca²⁺ chelators present in the cell like calcium-binding proteins. Forisomes are ATP-independent mechanically active motor proteins known to function in wound sealing of injured sieve elements of phloem tissue. The Ca²⁺-binding activity of forisome and its role in abiotic stress signaling were largely unknown. Here we report the Ca²⁺-binding activity of pea forisome (PsSEO-F1) and its novel function in promoting salinity tolerance in transgenic tobacco. Native PsSEO-F1 promoter positively responded in salinity stress as confirmed using GUS reporter. Overexpression of PsSEO-F1 tobacco plants confers salinity tolerance by alleviating ionic toxicity and increased ROS scavenging activity which probably results in reduced membrane damage and improved yield under salinity stress. Evaluation of several physiological indices shows an increase in relative water content, electrolyte leakage, proline accumulation and chlorophyll content in transgenic lines as compared with null-segregant control. Expression of several genes involved in cellular homeostasis is perturbed by PsSEO-F1 overexpression. These findings suggest that PsSEO-F1 provides salinity tolerance through cellular Ca²⁺ homeostasis which in turn modulates ROS machinery providing indirect link between Ca²⁺ and ROS signaling under salinity-induced perturbation. PsSEO-F1 most likely functions in salinity stress tolerance by improving antioxidant machinery and mitigating ion toxicity in transgenic lines. This finding should make an important contribution in our better understanding of the significance of calcium signaling in phloem tissue leading to salinity stress tolerance.

     

Association analysis of salt tolerance in cowpea (<Emphasis Type="Italic">Vigna unguiculata</Emphasis> (L.) Walp) at germination and seedling stages

Key message

This is the first report on association analysis of salt tolerance and identification of SNP markers associated with salt tolerance in cowpea.

Abstract

Cowpea (Vigna unguiculata (L.) Walp) is one of the most important cultivated legumes in Africa. The worldwide annual production in cowpea dry seed is 5.4 million metric tons. However, cowpea is unfavorably affected by salinity stress at germination and seedling stages, which is exacerbated by the effects of climate change. The lack of knowledge on the genetic underlying salt tolerance in cowpea limits the establishment of a breeding strategy for developing salt-tolerant cowpea cultivars. The objectives of this study were to conduct association mapping for salt tolerance at germination and seedling stages and to identify SNP markers associated with salt tolerance in cowpea. We analyzed the salt tolerance index of 116 and 155 cowpea accessions at germination and seedling stages, respectively. A total of 1049 SNPs postulated from genotyping-by-sequencing were used for association analysis. Population structure was inferred using Structure 2.3.4; K optimal was determined using Structure Harvester. TASSEL 5, GAPIT, and FarmCPU involving three models such as single marker regression, general linear model, and mixed linear model were used for the association study. Substantial variation in salt tolerance index for germination rate, plant height reduction, fresh and dry shoot biomass reduction, foliar leaf injury, and inhibition of the first trifoliate leaf was observed. The cowpea accessions were structured into two subpopulations. Three SNPs, Scaffold87490_622, Scaffold87490_630, and C35017374_128 were highly associated with salt tolerance at germination stage. Seven SNPs, Scaffold93827_270, Scaffold68489_600, Scaffold87490_633, Scaffold87490_640, Scaffold82042_3387, C35069468_1916, and Scaffold93942_1089 were found to be associated with salt tolerance at seedling stage. The SNP markers were consistent across the three models and could be used as a tool to select salt-tolerant lines for breeding improved cowpea tolerance to salinity.

     

Metabolic profiling of <Emphasis Type="Italic">Daphnia magna</Emphasis> exposed to environmental stressors by GC–MS and chemometric tools

Introduction

Climate change is a major concern for the scientific community, demanding novel information about the effects of environmental stressors on living organisms. Metabolic profiling is required for achieving the most extensive possible range of compounds and their concentration changes on stressed conditions.

Objectives

Individuals of the crustacean species Daphnia magna were exposed to three different abiotic factors linked to global climate change: high salinity, high temperature levels and hypoxia. Advanced chemometric tools were used to characterize the metabolites affected by the exposure.

Method

An exploratory analysis of gas chromatography-mass spectrometry (GC–MS) data was performed to discriminate between control and exposed daphnid samples. Due to the complexity of these GC–MS data sets, a comprehensive untargeted analysis of the full scan data was performed using multivariate curve resolution-alternating least squares (MCR-ALS) method. This approach enabled to resolve most of the metabolite signals from interference peaks caused by derivatization reactions. Metabolites with significant changes in their peak areas were tentatively identified and the involved metabolic pathways explored.

Results

D. magna metabolic biomarkers are proposed for the considered physical factors. Metabolites related with energy metabolic pathways including some amino acids, carbohydrates, organic acids and nucleosides were identified as potential biomarkers of the investigated treatments.

Conclusions

The proposed untargeted GC–MS metabolomics strategy and multivariate data analysis tools were useful to investigate D. magna metabolome under environmental stressed conditions.

     

Effect of ectomycorrhizal composition on survival and growth of <Emphasis Type="Italic">Pinus thunbergii</Emphasis> seedlings varying in resistance to the pine wilt nematode

Key message

Ectomycorrhizal composition and associated fungi affect the intra-specific ability of resistant black pines for physiological adaptation.

Abstract

Since Japanese black pine (Pinus thunbergii Parl.) forests have been widely devastated by pine wilt disease, several kinds of resistant black pines have been developed. Although all of the resistant black pines are the same species, these resistant trees show different physiological characteristics. We investigated the survival rates and growth rates, as well as ectomycorrhizal composition and associated fungi, on four kinds of Japanese black pine seedlings (three pine wilt-resistant and one non-resistant), and elucidated the factors affecting the various physiological characteristics. We found that the abundance of ectomycorrhizal types differed even though seedlings were grown sympatrically in the same areas for about 2 years. The seedlings that had plentiful white ectomycorrhizae showed the highest survival and growth rates regardless of the variety of black pine. Sequence similarities of the white ectomycorrhizae in the rDNA ITS region were best matched with members of Astraeus sp., Atheliaceae, Boletaceae and Thelephoraceae. Our findings indicate that intra-specific physiological adaptation might be affected by ectomycorrhizal composition or by the specific ectomycorrhizal species.

     

Extraction of natural products from bark of <Emphasis Type="Italic">Betula pendula</Emphasis> using ionic liquids

Introduction

Aqueous–methanol mixtures have successfully been applied to extract a broad range of metabolites from plant tissue. However, a certain amount of material remains insoluble.

Objectives

To enlarge the metabolic compendium, two ionic liquids were selected to extract the methanol insoluble part of trunk from Betula pendula.

Methods

The extracted compounds were analyzed by LC/MS and GC/MS.

Results

The results show that 1-butyl-3-methylimidazolium acetate (IL-Ac) predominantly resulted in fatty acids, whereas 1-ethyl-3-methylimidazolium tosylate (IL-Tos) mostly yielded phenolic structures. Interestingly, bark yielded more ionic liquid soluble metabolites compared to interior wood.

Conclusion

From this one can conclude that the application of ionic liquids may expand the metabolic snapshot.

     

An aquaporin gene from halophyte <Emphasis Type="Italic">Sesuvium portulacastrum</Emphasis>, <Emphasis Type="Italic">SpAQP1</Emphasis>, increases salt tolerance in transgenic tobacco

Key message

SpAQP1 was strongly induced by salt in an ABA-independent way, promoted seed germination and root growth in transgenic tobaccos and increased salt tolerance by increasing the activities of antioxidative enzymes.

Abstract

Aquaporin (AQP) plays crucial roles in the responses of plant to abiotic stresses such as drought, salt and cold. Compared to glycophytes, halophytes often have excellent salt and drought tolerances. To uncover the molecular mechanism of halophyte Sesuvium portulacastrum tolerance to salt, in this study, an AQP gene, SpAQP1, from S. portulacastrum was isolated and characterized. The amino acid sequence of SpAQP1 shared high homology with that of plant plasma membrane intrinsic proteins (PIPs) and contained the distinct molecular features of PIPs. In the phylogenic tree, SpAQP1 was evidently classified as the PIP2 subfamily. SpAQP1 is expressed in roots, stems and leaves, and was significantly induced by NaCl treatment and inhibited by abscisic acid (ABA) treatment. When heterologously expressed in yeast and tobacco, SpAQP1 enhanced the salt tolerance of yeast strains and tobacco plants and promoted seed germination and root growth under salt stress in transgenic plants. The activity of antioxidative enzymes including superoxide dismutase, peroxidase and catalase was increased in transgenic plants overexpressing SpAQP1. Taken together, our studies suggested that SpAQP1 functioned in the responses of S. portulacastrum to salt stress and could increase salt tolerance by enhancing the antioxidative activity of plants.