Molecular Mechanisms Underlying Salt Tolerance Research Articles

Transcriptome Analysis of the Regulatory Mechanisms of Holly (Ilex dabieshanensis) under Salt Stress Conditions.

The holly Ilex dabieshanensis K. Yao & M. B. Deng, a tree endemic to the Dabieshan Mountains region in China, is a commonly used landscaping plant. Like other crops, its growth is affected by salt stress. The molecular mechanism underlying salt tolerance in holly is still unclear. In this study, we used NaCl treatment and RNA sequencing (RNA-seq) at different times to identify the salt stress response genes of holly. A total of 4775 differentially expressed genes (DEGs) were identified. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of the DEGs obtained at different salt treatment times (3, 6, 9, 12, and 24 h), as compared to control (ck, 0 h), showed that plant hormone signal transduction and carotenoid biosynthesis were highly enriched. The mechanism by which holly responds to salt stress involves many plant hormones, among which the accumulation of abscisic acid (ABA) and its signal transduction may play an important role. In addition, ion homeostasis, osmotic metabolism, accumulation of antioxidant enzymes and nonenzymatic antioxidant compounds, and transcription factors jointly regulate the physiological balance in holly, providing important guarantees for its growth and development under conditions of salt stress. These results lay the foundation for studying the molecular mechanisms of salt tolerance in holly and for the selection of salt-tolerant varieties.

Weighted gene co-expression network analysis reveals hub genes regulating response to salt stress in peanut

Peanut (Arachis hypogaea L.) is an important oilseed crop worldwide. However, soil salinization becomes one of the main limiting factors of peanut production. Therefore, developing salt-tolerant varieties and understanding the molecular mechanisms of salt tolerance is important to protect peanut yield in saline areas. In this study, we selected four peanut varieties with contrasting response to salt challenges with T1 and T2 being tolerance and S1 and S2 being susceptible. High-throughput RNA sequencing resulted in more than 314.63 Gb of clean data from 48 samples. We identified 12,057 new genes, 7,971of which have functional annotations. KEGG pathway enrichment analysis of uniquely expressed genes in salt-tolerant peanut revealed that upregulated genes in the root are involved in the MAPK signaling pathway, fatty acid degradation, glycolysis/gluconeogenesis, and upregulated genes in the shoot were involved in plant hormone signal transduction and the MAPK signaling pathway. Na+ content, K+ content, K+/ Na+, and dry mass were measured in root and shoot tissues, and two gene co-expression networks were constructed based on weighted gene co-expression network analysis (WGCNA) in root and shoot. In this study, four key modules that are highly related to peanut salt tolerance in root and shoot were identified, plant hormone signal transduction, phenylpropanoid biosynthesis, starch and sucrose metabolism, flavonoid biosynthesis, carbon metabolism were identified as the key biological processes and metabolic pathways for improving peanut salt tolerance. The hub genes include genes encoding ion transport (such as HAK8, CNGCs, NHX, NCL1) protein, aquaporin protein, CIPK11 (CBL-interacting serine/threonine-protein kinase 11), LEA5 (late embryogenesis abundant protein), POD3 (peroxidase 3), transcription factor, and MAPKKK3. There were some new salt-tolerant genes identified in peanut, including cytochrome P450, vinorine synthase, sugar transport protein 13, NPF 4.5, IAA14, zinc finger CCCH domain-containing protein 62, beta-amylase, fatty acyl-CoA reductase 3, MLO-like protein 6, G-type lectin S-receptor-like serine/threonine-protein kinase, and kinesin-like protein KIN-7B. The identification of key modules, biological pathways, and hub genes in this study enhances our understanding of the molecular mechanisms underlying salt tolerance in peanuts. This knowledge lays a theoretical foundation for improving and innovating salt-tolerant peanut germplasm.

Identification of core genes involved in the response of Apocynum venetum to salt stress based on transcriptome sequencing and WGCNA.

Apocynum venetum L. belongs to the Apocynaceae family and is a plant that is highly resistant to stress. It is important in the fields of ecology, feeding, industry and medicine. The molecular mechanism underlying salt tolerance has not been elucidated. In this study, RNA-seq based transcriptome sequencing of A. venetum leaves after 0, 2, 6, 12, 24 and 48 h of treatment with 300 mM NaCl was performed. We conducted a comprehensive analysis of the transcriptome expression profiles of A. venetum under salt stress using the WGCNA method and identified red, black, and brown as the core modules regulating the salt tolerance of A. venetum. A co-expression regulatory network was constructed to identify the core genes in the module according to the correlations between genes. The genes TRINITY_DN102_c0_g1 (serine carboxypeptidase), TRINITY_DN3073_c0_g1 (SOS signaling pathway) and TRINITY_DN6732_c0_g1 (heat shock transcription factor) in the red module were determined to be the core genes. Two core genes in the black module, TRINITY_DN9926_c0_g1 and TRINITY_DN7962_c0_g1, are pioneer candidate salt tolerance-associated genes in A. venetum. The genes in the brown module were mainly enriched in two pathways, namely photosynthesis and osmotic balance. Among them, the TRINITY_DN6321_c0_g2 and TRINITY_DN244_c0_g1 genes encode aquaporin, which is helpful for maintaining the cell water balance and plays a protective role in defending A. venetum under abiotic stress. Our findings contribute to the identification of core genes involved in the response of A. venetum to salt stress.

Unraveling wheat’s response to salt stress during early growth stages through transcriptomic analysis and co-expression network profiling

BackgroundSoil salinization is one of the vital factors threatening the world’s food security. To reveal the biological mechanism of response to salt stress in wheat, this study was conducted to resolve the transcription level difference to salt stress between CM6005 (salt-tolerant) and KN9204 (salt-sensitive) at the germination and seedling stage.ResultsTo investigate the molecular mechanism underlying salt tolerance in wheat, we conducted comprehensive transcriptome analyses at the seedling and germination stages. Two wheat cultivars, CM6005 (salt-tolerant) and KN9204 (salt-sensitive) were subjected to salt treatment, resulting in a total of 24 transcriptomes. Through expression-network analysis, we identified 17 modules, 16 and 13 of which highly correlate with salt tolerance-related phenotypes in the germination and seedling stages, respectively. Moreover, we identified candidate Hub genes associated with specific modules and explored their regulatory relationships using co-expression data. Enrichment analysis revealed specific enrichment of gibberellin-related terms and pathways in CM6005, highlighting the potential importance of gibberellin regulation in enhancing salt tolerance. In contrast, KN9204 exhibited specific enrichment in glutathione-related terms and activities, suggesting the involvement of glutathione-mediated antioxidant mechanisms in conferring resistance to salt stress. Additionally, glucose transport was found to be a fundamental mechanism for salt tolerance during wheat seedling and germination stages, indicating its potential universality in wheat. Wheat plants improve their resilience and productivity by utilizing adaptive mechanisms like adjusting osmotic balance, bolstering antioxidant defenses, accumulating compatible solutes, altering root morphology, and regulating hormones, enabling them to better withstand extended periods of salt stress.ConclusionThrough utilizing transcriptome-level analysis employing WGCNA, we have revealed a potential regulatory mechanism that governs the response to salt stress and recovery in wheat cultivars. Furthermore, we have identified key candidate central genes that play a crucial role in this mechanism. These central genes are likely to be vital components within the gene expression network associated with salt tolerance. The findings of this study strongly support the molecular breeding of salt-tolerant wheat, particularly by utilizing the genetic advancements based on CM6005 and KN9204.

Combined transcriptomic and metabolomic analysis of alginate oligosaccharides alleviating salt stress in rice seedlings

BackgroundSalt stress is one of the key factors limiting rice production. Alginate oligosaccharides (AOS) enhance plant stress resistance. However, the molecular mechanism underlying salt tolerance in rice induced by AOS remains unclear. FL478, which is a salt-tolerant indica recombinant inbred line and IR29, a salt-sensitive rice cultivar, were used to comprehensively analyze the effects of AOS sprayed on leaves in terms of transcriptomic and metabolite profiles of rice seedlings under salt stress.ResultsIn this experiment, exogenous application of AOS increased SOD, CAT and APX activities, as well as GSH and ASA levels to reduce the damage to leaf membrane, increased rice stem diameter, the number of root tips, aboveground and subterranean biomass, and improved rice salt tolerance. Comparative transcriptomic analyses showed that the regulation of AOS combined with salt treatment induced the differential expression of 305 and 1030 genes in FL478 and IR29. The expressed genes enriched in KEGG pathway analysis were associated with antioxidant levels, photosynthesis, cell wall synthesis, and signal transduction. The genes associated with light-trapping proteins and RLCK receptor cytoplasmic kinases, including CBA, LHCB, and Lhcp genes, were fregulated in response to salt stress. Treatment with AOS combined with salt induced the differential expression of 22 and 50 metabolites in FL478 and IR29. These metabolites were mainly related to the metabolism of amino and nucleotide sugars, tryptophan, histidine, and β -alanine. The abundance of metabolites associated with antioxidant activity, such as 6-hydroxymelatonin, wedelolactone and L-histidine increased significantly. Combined transcriptomic and metabolomic analyses revealed that dehydroascorbic acid in the glutathione and ascorbic acid cycles plays a vital role in salt tolerance mediated by AOS.ConclusionAOS activate signal transduction, regulate photosynthesis, cell wall formation, and multiple antioxidant pathways in response to salt stress. This study provides a molecular basis for the alleviation of salt stress-induced damage by AOS in rice.

The ZmbHLH32-ZmIAA9-ZmARF1 module regulates salt tolerance in maize

The growth and productivity of maize (Zea mays), along with other crop plants, can be significantly hindered by salt stress. Nevertheless, the precise molecular mechanism underlying salt tolerance in maize has yet to be fully elucidated. Hence, it was attempted to identify ZmIAA9, a member of the maize Aux/IAA gene family, as a positive regulator of salt tolerance in maize, which was accompanied by the increased ROS detoxification and elevated transcript abundances of ROS scavenging genes. Molecular and biochemical assays have provided compelling evidence that ZmbHLH32, a transcription factor belonging to the bHLH family, was capable of binding directly to the promoter region of ZmIAA9, thereby activating its expression. This interaction between ZmbHLH32 and ZmIAA9 could be critical for the regulation of salt tolerance in maize. As expected, overexpression of ZmbHLH32 led to the enhanced salt tolerance. In contrast, decreased salt tolerance was attained after application of knockout mutants of ZmbHLH32. Furthermore, ZmARF1, which could act as a downstream of ZmIAA9, was found to physically interact with ZmIAA9 and repress the expression levels of ROS scavenging genes. Thus, our work uncovers a novel mechanism of ZmbHLH32-ZmIAA9-ZmARF1 module-mediated salt tolerance in maize, which can be exploited for breeding salt-tolerant maize varieties.

Transcriptomic Analysis and Salt-Tolerance Gene Mining during Rice Germination.

Salt stress is an important environmental factor affecting crop growth and development. One of the important ways to improve the salt tolerance of rice is to identify new salt-tolerance genes, reveal possible mechanisms, and apply them to the creation of new germplasm and the breeding of new varieties. In this study, the salt-sensitive japonica variety Tong 35 (T35) and salt-tolerant japonica variety Ji Nongda 709 (JND709) were used. Salt stress treatment with a 150 mmol/L NaCl solution (the control group was tested without salt stress treatment simultaneously) was continued until the test material was collected after the rice germination period. Twelve cDNA libraries were constructed, and 5 comparator groups were established for transcriptome sequencing. On average, 9.57G of raw sequencing data were generated per sample, with alignment to the reference genome above 96.88% and alignment to guanine-cytosine (GC) content above 53.86%. A total of 16,829 differentially expressed genes were present in the five comparison groups, of which 2390 genes were specifically expressed in T35 (category 1), 3306 genes were specifically expressed in JND709 (category 2), and 1708 genes were differentially expressed in both breeds (category 3). Differentially expressed genes were subjected to gene ontology (GO), functional enrichment analysis, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, which revealed that these genes belonged to three main classes: molecular function, cellular components, and biological processes. KEGG pathway analysis showed that the significantly enriched pathways for these differentially expressed genes included phenylpropane biosynthesis, phytohormone signaling, and the interaction of plants with pathogens. In this study, we provided a reference for studying the molecular mechanism underlying salt tolerance during germination.

Comparative transcriptome analysis provides novel insights into molecular response of salt-tolerant and sensitive polyembryonic mango genotypes to salinity stress at seedling stage.

Increased soil salinity in the recent years has adversely affected the productivity of mango globally. Extending the cultivation of mango in salt affected regions warrants the use of salinity tolerant/resistant rootstocks. However, the lack of sufficient genomic and transcriptomic information impedes comprehensive research at the molecular level. We employed RNA sequencing-based transcriptome analysis to gain insight into molecular response to salt stress by using two polyembryonic mango genotypes with contrasting response to salt stress viz., salt tolerant Turpentine and salt susceptible Mylepelian. RNA sequencing by Novaseq6000 resulted in a total of 2795088, 17535948, 7813704 and 5544894 clean reads in Mylepelian treated (MT), Mylepelian control (MC), Turpentine treated (TT) and Turpentine control (TC) respectively. In total, 7169 unigenes annotated against all the five public databases, including NR, NT, PFAM, KOG, Swissport, KEGG and GO. Further, maximum number of differentially expressed genes were found between MT and MC (2106) followed by MT vs TT (1158) and TT and TC (587). The differentially expressed genes under different treatment levels included transcription factors (bZIP, NAC, bHLH), genes involved in Calcium-dependent protein kinases (CDPKs), ABA biosynthesis, Photosynthesis etc. Expression of few of these genes was experimentally validated through quantitative real-time PCR (qRT-PCR) and contrasting expression pattern of Auxin Response Factor 2 (ARF2), Late Embryogenesis Abundant (LEA) and CDPK genes were observed between Turpentine and Mylepelian. The results of this study will be useful in understanding the molecular mechanism underlying salt tolerance in mango which can serve as valuable baseline information to generate new targets in mango breeding for salt tolerance.

Halophyte Nitraria billardieri CIPK25 promotes photosynthesis in Arabidopsis under salt stress.

The calcineurin B-like (CBL)-interacting protein kinases (CIPKs), a type of plant-specific genes in the calcium signaling pathway, function in response to adverse environments. However, few halophyte derived CIPKs have been studied for their role in plant physiological and developmental adaptation during abiotic stresses, which inhibits the potential application of these genes to improve environmental adaptability of glycophytes. In this study, we constructed Nitraria billardieri CIPK25 overexpressing Arabidopsis and analyzed the seedling development under salt treatment. Our results show that Arabidopsis with NbCIPK25 expression exhibits more vigorous growth than wild type plants under salt condition. To gain insight into the molecular mechanisms underlying salt tolerance, we profiled the transcriptome of WT and transgenic plants via RNA-seq. GO and KEGG analyses revealed that upregulated genes in NbCIPK25 overexpressing seedlings under salt stress are enriched in photosynthesis related terms; Calvin-cycle genes including glyceraldehyde-3-phosphate dehydrogenases (GAPDHs) are significantly upregulated in transgenic plants, which is consistent with a decreased level of NADPH (GAPDH substrate) and increased level of NADP+. Accordingly, NbCIPK25 overexpressing plants exhibited more efficient photosynthesis; soluble sugar and proteins, as photosynthesis products, showed a higher accumulation in transgenic plants. These results provide molecular insight into how NbCIPK25 promotes the expression of genes involved in photosynthesis, thereby maintaining plant growth under salt stress. Our finding supports the potential application of halophyte-derived NbCIPK25 in genetic modification for better salt adaptation.

Genome-Wide Analysis of CqCrRLK1L and CqRALF Gene Families in Chenopodium quinoa and Their Roles in Salt Stress Response.

Chenopodium quinoa is a halophyte with exceptional nutritional qualities, and therefore it is potentially an ideal crop to grow in saline soils, not only addressing the problem of land salinization, but also providing nutrient food for the health of humans. Currently, the molecular mechanisms underlying salt tolerance in quinoa are still largely unknown. In Arabidopsis thaliana, Catharanthus roseus receptor-like kinase (CrRLK1Ls) FERONIA (FER) and its ligands rapid alkalinization factors (RALFs) have been reported that participate in the regulation of salt tolerance. Here, we performed a genome-wide analysis and identified 26 CqCrRLK1L and 18 CqRALF family genes in quinoa genome. Transcriptomic profiling of the leaf, root, stamen, and pistil tissues of quinoa reveals that different CqCrRLK1L and CqRALF genes exhibit tissue-specific expression patterns, which is consistent with that observed in other plant species. RNA-seq data show that three CqCrRLK1L genes are highly up-regulated after salt treatment, suggesting that some CqCrRLK1L family genes are transcriptionally responsive to salt stress in quinoa. Biochemical study indicates that CqRALF15, a paralog of Arabidopsis RALF22, is physically associated with CrRLK1L proteins CqFER and AtFER. CqRALF15 and AtRALF22 are functionally conserved in inducing the internalization of AtFER and in triggering root growth inhibition in both quinoa and Arabidopsis. Moreover, overexpression of CqRALF15 in Arabidopsis results in enhanced leaf bleaching under salt stress, indicating that CqRALF15 is involved in salt stress response. Together, our study characterizes CqCrRLK1L and CqRALF family genes in quinoa at genomic, transcriptional, and protein levels, and provides evidence to support their roles in salt stress response.

Root Na+ Content Negatively Correlated to Salt Tolerance Determines the Salt Tolerance of Brassica napus L. Inbred Seedlings.

Soil salinization is a major environmental stressor that reduces the growth and yield of crops. Maintaining the balance of ions under salinity is vital for plant salt tolerance; however, little is known about the correlation between the salt tolerance of crops and the ion contents of their roots and shoots. Here, we investigated the poorly understood salt-tolerance mechanisms, particularly regarding ion contents (particularly Na+), in Brassica napus subsp. napus L., an agriculturally important species. Twenty B. napus inbred lines were randomly chosen from five salt-tolerance categories and treated with increasing concentrations of NaCl (0–200 mmol) for this work. We found that the root Na+ content is the most correlated limiting factor for the salt tolerance of B. napus; the higher the salt tolerance, the lower the root Na+ content. Correspondingly, the Ca2+/Na+ and K+/Na+ ratios of the roots were highly correlated with B. napus salt tolerance, indicating that the selective absorption ability of these ions by the roots and their translocation to the shoots play a pivotal role in this trait. These data provide a foundation for the further study of the molecular mechanisms underlying salt tolerance and for breeding salt-tolerant B. napus cultivars.

Integrated Physiological, Transcriptomic, and Metabolomic Analyses Revealed Molecular Mechanism for Salt Resistance in Soybean Roots.

Salinity stress is a threat to yield in many crops, including soybean (Glycine max L.). In this study, three soybean cultivars (JD19, LH3, and LD2) with different salt resistance were used to analyze salt tolerance mechanisms using physiology, transcriptomic, metabolomic, and bioinformatic methods. Physiological studies showed that salt-tolerant cultivars JD19 and LH3 had less root growth inhibition, higher antioxidant enzyme activities, lower ROS accumulation, and lower Na+ and Cl- contents than salt-susceptible cultivar LD2 under 100 mM NaCl treatment. Comparative transcriptome analysis showed that compared with LD2, salt stress increased the expression of antioxidant metabolism, stress response metabolism, glycine, serine and threonine metabolism, auxin response protein, transcription, and translation-related genes in JD19 and LH3. The comparison of metabolite profiles indicated that amino acid metabolism and the TCA cycle were important metabolic pathways of soybean in response to salt stress. In the further validation analysis of the above two pathways, it was found that compared with LD2, JD19, and LH3 had higher nitrogen absorption and assimilation rate, more amino acid accumulation, and faster TCA cycle activity under salt stress, which helped them better adapt to salt stress. Taken together, this study provides valuable information for better understanding the molecular mechanism underlying salt tolerance of soybean and also proposes new ideas and methods for cultivating stress-tolerant soybean.

Comparative transcriptome analysis reveals the molecular mechanism of salt tolerance in Apocynum venetum

Apocynum venetum is a traditional Chinese medicinal herb with tolerance to various abiotic stresses, especially, salinity. However, only a few studies have investigated the salt-tolerant mechanism of this non-halophyte under salt stress at phenotypic and physiological levels. To explore the molecular mechanism of salinity tolerance in A. venetum, the global transcriptome profiles of seedling leaves under different salt-stress durations, using 200 mM NaCl, were analyzed. De novo assembly of approximately 715 million high-quality reads and approximately 105.61 Gb sequence data was performed. In total, 2822 differentially expressed genes (DEGs) were identified. DEGs were significantly enriched in flavonoid metabolism-related pathways such as “flavonoid biosynthesis” and “phenylpropanoid biosynthesis”. Most of these DEGs were downregulated under salt stress. However, genes encoding the non-selective cation channels and antioxidants were upregulated under salt stress, whereas most cell wall-related DEGs were downregulated. Consequently, the concentration of flavonoids decreased, whereas that of Na+ increased with exposure time. Thus, we hypothesized that the accumulation of Na+ in the leaves, which resulted in reduced flavonoid concentration under salt stress, directly led to a decrease in the salt tolerance of A. venetum. This was verified by overexpressing four flavonoid synthesis pathway genes in Arabidopsis. The transgenic plants showed higher salt tolerance than the wild-type plants due to the accumulation of total flavonoids. These physiological and transcriptome analyses of A. venetum revealed major molecular underpinnings contributing to the responses of A. venetum to salt stress, thereby improving our understanding of the molecular mechanisms underlying salt tolerance in A. venetum and plants in general. The findings serve as a basis for functional studies on and engineering strategies for plant salinity tolerance.

Salt-tolerance of Phaseolus vulgaris L. is a function of the potentiation extent of antioxidant enzymes and the expression profiles of polyamine encoding genes

Selection and improvement of crop salt tolerance constitutes an urgent need for increasing agricultural and food production to feed the growing human population. Two bean (Phaseolus vulgaris L.) genotypes, namely Tema and Djadida varying in their tolerance to NaCl, were selected to investigate the determinant physiological and molecular mechanisms underlying salt tolerance. At the physiological level, salinity resulted in a significant decrease in growth, water status, stomatal conductance and photosynthesis for both genotypes. The potassium content was higher in the salt-tolerant genotype Tema under normal and saline conditions. Both genotypes had a significant increase in proline, glucose, fructose and sucrose concentrations, but glucose and fructose were differentially higher in the salt-tolerant genotype. While putrescine and spermidine concentrations decreased, spermine concentration significantly enhanced under salt stress along with antioxidant enzymes activities namely catalase, glutathione reductase and ascorbate peroxidase. The increase was higher in the salt-tolerant genotype. The expression levels of genes encoding enzymes involved in spermine biosynthesis revealed an upregulation of spermine synthase (SPMS) and a downregulation of polyamine oxidase (PAO4). Overall, the difference between genotypes is more quantitative than qualitative; the salt-tolerant genotype Tema displayed a better physiological and molecular response under salinity. By measuring these molecules in salt-stressed plants, several genotypes could be screened for breeding programmes leading to improve salt tolerance in beans.

Comparative transcriptome and metabolome profiling reveal molecular mechanisms underlying OsDRAP1-mediated salt tolerance in rice

Integration of transcriptomics and metabolomics data can provide detailed information for better understanding the molecular mechanisms underlying salt tolerance in rice. In the present study, we report a comprehensive analysis of the transcriptome and metabolome of rice overexpressing the OsDRAP1 gene, which encodes an ERF transcription factor and was previously identified to be conferring drought tolerance. Phenotypic analysis showed that OsDRAP1 overexpression (OE) improved salt tolerance by increasing the survival rate under salt stress. OsDRAP1 affected the physiological indices such as superoxide dismutase (SOD), catalase (CAT) and malondialdehyde (MDA) to enhance redox homeostasis and membrane stability in response to salt stress. Higher basal expression of OsDRAP1 resulted in differential expression of genes that potentially function in intrinsic salt tolerance. A core set of genes with distinct functions in transcriptional regulation, organelle gene expression and ion transport were substantially up-regulated in the OE line in response to salt stress, implying their important role in OsDRAP1-mediated salt tolerance. Correspondingly, metabolome profiling detected a number of differentially metabolites in the OE line relative to the wild type under salt stress. These metabolites, including amino acids (proline, valine), organic acids (glyceric acid, phosphoenolpyruvic acid and ascorbic acid) and many secondary metabolites, accumulated to higher levels in the OE line, demonstrating their role in salt tolerance. Integration of transcriptome and metabolome analysis highlights the crucial role of amino acids and carbohydrate metabolism pathways in OsDRAP1-mediated salt tolerance.

Transcriptional profiling of two contrasting genotypes uncovers molecular mechanisms underlying salt tolerance in alfalfa

Alfalfa is an important forage crop that is moderately tolerant to salinity; however, little is known about its salt-tolerance mechanisms. We studied root and leaf transcriptomes of a salt-tolerant (G03) and a salt-sensitive (G09) genotype, irrigated with waters of low and high salinities. RNA sequencing led to 1.73 billion high-quality reads that were assembled into 418,480 unigenes; 35% of which were assigned to 57 Gene Ontology annotations. The unigenes were assigned to pathway databases for understanding high-level functions. The comparison of two genotypes suggested that the low salt tolerance index for transpiration rate and stomatal conductance of G03 compared to G09 may be due to its reduced salt uptake under salinity. The differences in shoot biomass between the salt-tolerant and salt-sensitive lines were explained by their differential expressions of genes regulating shoot number. Differentially expressed genes involved in hormone-, calcium-, and redox-signaling, showed treatment- and genotype-specific differences and led to the identification of various candidate genes involved in salinity stress, which can be investigated further to improve salinity tolerance in alfalfa. Validation of RNA-seq results using qRT-PCR displayed a high level of consistency between the two experiments. This study provides valuable insight into the molecular mechanisms regulating salt tolerance in alfalfa.

Physiological and Proteomic Analysis of Brassica napus in Response to Salt Stress

Salinity is a major abiotic stress that adversely affects plant growth and development. Canola (Brassica napus L.) is an important oilseed crop in the world, and its yield decreases drastically with increasing salinity. To date, little is known about the molecular mechanisms underlying its salt stress response and tolerance. This study combines physiological assays with comparative proteomics to understand how B. napus plants respond to salt stress. The changes in relative water content, electrical conductance, stomata conductance, intercellular CO2 concentration, transpiration rate, photosynthesis rate, water usage efficiency, respiration rate, chlorophyll fluorescence, antioxidant enzyme activities, soluble sugar, proline and betaine in B. napus plants under different NaCl concentrations were analyzed. Proteomic profiles of B. napus plants under 100, 200 and 400 mM NaCl treatment at 7 day and 14 day were acquired using iTRAQ LC-MS/MS based quantitative proteomics. A total of 2316 proteins were identified in B. napus leaves, of which 614 proteins showed differential expression under salt stress. These proteins were mainly involved in 10 processes, of which proteins in stress and defense, metabolism and photosynthesis pathways ranked the top three. Subcellular localization analysis showed that most proteins were located in chloroplast, cytoplasm, mitochondria and nucleus. A total of 138 differentially expressed proteins were predicted to interact with each other. These results have provided a comprehensive view of the physiological and molecular processes taken place in B. napus leaves under salt stress, and revealed the molecular mechanisms underlying salt tolerance of B. napus plants.

Comparative Transcriptome, Metabolome, and Ionome Analysis of Two Contrasting Common Bean Genotypes in Saline Conditions.

Soil salinity is a major abiotic stress factor that limits agricultural productivity worldwide, and this problem is expected to grow in the future. Common bean is an important protein source in developing countries however highly susceptible to salt stress. To understand the underlying mechanism of salt stress responses, transcriptomics, metabolomics, and ion content analysis were performed on both salt-tolerant and susceptible common bean genotypes in saline conditions. Transcriptomics has demonstrated increased photosynthesis in saline conditions for tolerant genotype while the susceptible genotype acted in contrast. Transcriptome also displayed active carbon and amino-acid metabolism for the tolerant genotype. Analysis of metabolites with GC-MS demonstrated the boosted carbohydrate metabolism in the tolerant genotype with increased sugar content as well as better amino-acid metabolism. Accumulation of lysine, valine, and isoleucine in the roots of the susceptible genotype suggested a halted stress response. According to ion content comparison, the tolerant genotype managed to block accumulation of Na+ in the leaves while accumulating significantly less Na+ in the roots compared to susceptible genotype. K+ levels increased in the leaves of both genotype and the roots of the susceptible one but dropped in the roots of the tolerant genotype. Additionally, Zn+2 and Mn+2 levels were dropped in the tolerant roots, while Mo+2 levels were significantly higher in all tissues in both control and saline conditions for tolerant genotype. The results of the presented study have demonstrated the differences in contrasting genotypes and thus provide valuable information on the pivotal molecular mechanisms underlying salt tolerance.

Low Mannitol Concentrations in Arabidopsis thaliana Expressing Ectocarpus Genes Improve Salt Tolerance.

Mannitol is abundant in a wide range of organisms, playing important roles in biotic and abiotic stress responses. Nonetheless, mannitol is not produced by a vast majority of plants, including many important crop plants. Mannitol-producing transgenic plants displayed improved tolerance to salt stresses though mannitol production was rather low, in the µM range, compared to mM range found in plants that innately produce mannitol. Little is known about the molecular mechanisms underlying salt tolerance triggered by low concentrations of mannitol. Reported here is the production of mannitol in Arabidopsis thaliana, by expressing two mannitol biosynthesis genes from the brown alga Ectocarpus sp. strain Ec32. To date, no brown algal genes have been successfully expressed in land plants. Expression of mannitol-1-phosphate dehydrogenase and mannitol-1-phosphatase genes was associated with the production of 42.3–52.7 nmol g−1 fresh weight of mannitol, which was sufficient to impart salinity and temperature stress tolerance. Transcriptomics revealed significant differences in the expression of numerous genes, in standard and salinity stress conditions, including genes involved in K+ homeostasis, ROS signaling, plant development, photosynthesis, ABA signaling and secondary metabolism. These results suggest that the improved tolerance to salinity stress observed in transgenic plants producing mannitol in µM range is achieved by the activation of a significant number of genes, many of which are involved in priming and modulating the expression of genes involved in a variety of functions including hormone signaling, osmotic and oxidative stress, and ion homeostasis.

Differentially expressed genes related to oxidoreductase activity and glutathione metabolism underlying the adaptation of Phragmites australis from the salt marsh in the Yellow River Delta, China.

The common reed (Phragmites australis) is a dominant species in the coastal wetlands of the Chinese Yellow River Delta, where it tolerates a wide range of salinity. Recent environmental changes have led to the increase of soil salinity in this region, which has degraded much of the local vegetation. Clones of common reeds from the tidal marsh may have adapted to local high salinity habitat through selection on genes and metabolic pathways conferring salt tolerance. This study aims to reveal molecular mechanisms underlying salt tolerance in the tidal reed by comparing them to the salt-sensitive freshwater reed under salt stress. We employed comparative transcriptomics to reveal the differentially expressed genes (DEGs) between these two types of common reeds under different salinity conditions. The results showed that only three co-expressed genes were up-regulated and one co-expressed gene was down-regulated between the two reed types. On the other hand, 1,371 DEGs were exclusively up-regulated and 285 DEGs were exclusively down-regulated in the tidal reed compared to the control, while 115 DEGs were exclusively up-regulated and 118 DEGs were exclusively down-regulated in the freshwater reed compared to the control. From the pattern of enrichment of transcripts involved in salinity response, the tidal reed was more active and efficient in scavenging reactive oxygen species (ROS) than the freshwater reed, with the tidal reed showing significantly higher gene expression in oxidoreductase activity. Furthermore, when the reeds were exposed to salt stress, transcripts encoding glutathione metabolism were up-regulated in the tidal reed but not in the freshwater reed. DEGs related to encoding glutathione reductase (GR), glucose-6-phosphate 1-dehydrogenase (G6PDH), 6-phosphogluconate dehydrogenase (6PD), glutathione S-transferase (GST) and L-ascorbate peroxidase (LAP) were revealed as especially highly differentially regulated and therefore represented candidate genes that could be cloned into plants to improve salt tolerance. Overall, more genes were up-regulated in the tidal reed than in the freshwater reed from the Yellow River Delta when under salt stress. The tidal reed efficiently resisted salt stress by up-regulating genes encoding for oxidoreductase activity and glutathione metabolism. We suggest that this type of common reed could be extremely useful in the ecological restoration of degraded, high salinity coastal wetlands in priority.