The sugar beet gene encoding the sodium/proton exchanger 1 (BvNHX1) is regulated by a MYB transcription factor

Sodium/proton exchangers (NHX) are key players in the plant response to salinity and have a central role in establishing ion homeostasis. NHXs can be localized in the tonoplast or plasma membranes, where they exchange sodium ions for protons, resulting in sodium ions being removed from the cytosol into the vacuole or extracellular space. The expression of most plant NHX genes is modulated by exposure of the organisms to salt stress or water stress. We explored the regulation of the vacuolar NHX1 gene from the salt-tolerant sugar beet plant (BvNHX1) using Arabidopsis plants transformed with an array of constructs of BvHNX1::GUS, and the expression patterns were characterized using histological and quantitative assays. The 5′ UTR of BvNHX1, including its intron, does not modulate the activity of the promoter. Serial deletions show that a 337 bp promoter fragment sufficed for driving activity that indistinguishable from that of the full-length (2,464 bp) promoter. Mutating four putative cis-acting elements within the 337 bp promoter fragment revealed that MYB transcription factor(s) are involved in the activation of the expression of BvNHX1 upon exposure to salt and water stresses. Gel mobility shift assay confirmed that the WT but not the mutated MYB binding site is bound by nuclear protein extracted from salt-stressed Betavulgaris leaves

Arabidopsis Fatty Acid Desaturase FAD2 Is Required for Salt Tolerance during Seed Germination and Early Seedling Growth

Fatty acid desaturases play important role in plant responses to abiotic stresses. However, their exact function in plant resistance to salt stress is unknown. In this work, we provide the evidence that FAD2, an endoplasmic reticulum localized ω-6 desaturase, is required for salt tolerance in Arabidopsis. Using vacuolar and plasma membrane vesicles prepared from the leaves of wild-type (Col-0) and the loss-of-function Arabidopsis mutant, fad2, which lacks the functional FAD2, we examined the fatty acid composition and Na+-dependent H+ movements of the isolated vesicles. We observed that, when compared to Col-0, the level of vacuolar and plasma membrane polyunsaturation was lower, and the Na+/H+ exchange activity was reduced in vacuolar and plasma membrane vesicles isolated from fad2 mutant. Consistent with the reduced Na+/H+ exchange activity, fad2 accumulated more Na+ in the cytoplasm of root cells, and was more sensitive to salt stress during seed germination and early seedling growth, as indicated by CoroNa-Green staining, net Na+ efflux and salt tolerance analyses. Our results suggest that FAD2 mediated high-level vacuolar and plasma membrane fatty acid desaturation is essential for the proper function of membrane attached Na+/H+ exchangers, and thereby to maintain a low cytosolic Na+ concentration for salt tolerance during seed germination and early seedling growth in Arabidopsis

Isolation and Characterization of Maize PMP3 Genes Involved in Salt Stress Tolerance

Plasma membrane protein 3 (PMP3), a class of small hydrophobic polypeptides with high sequence similarity, is responsible for salt, drought, cold, and abscisic acid. These small hydrophobic ploypeptides play important roles in maintenance of ion homeostasis. In this study, eight ZmPMP3 genes were cloned from maize and responsive to salt, drought, cold and abscisic acid. The eight ZmPMP3s were membrane proteins and their sequences in trans-membrane regions were highly conserved. Phylogenetic analysis showed that they were categorized into three groups. All members of group II were responsive to ABA. Functional complementation showed that with the exception of ZmPMP3-6, all were capable of maintaining membrane potential, which in turn allows for regulation of intracellular ion homeostasis. This process was independent of the presence of Ca2+. Lastly, over-expression of ZmPMP3-1 enhanced growth of transgenic Arabidopsis under salt condition. Through expression analysis of deduced downstream genes in transgenic plants, expression levels of three ion transporter genes and four important antioxidant genes in ROS scavenging system were increased significantly in transgenic plants during salt stress. This tolerance was likely achieved through diminishing oxidative stress due to the possibility of ZmPMP3-1's involvement in regulation of ion homeostasis, and suggests that the modulation of these conserved small hydrophobic polypeptides could be an effective way to improve salt tolerance in plants

Natural Variation in Arabidopsis thaliana Revealed a Genetic Network Controlling Germination Under Salt Stress

Plant responses to environmental stresses are polygenic and complex traits. In this study quantitative genetics using natural variation in Arabidopsis thaliana was used to investigate the genetic architecture of plant responses to salt stress. Eighty seven A. thaliana accessions were screened and showed a large variation for root development and seed germination under 125 and 200 mM NaCl, respectively. Twenty two quantitative trait loci for these traits have been detected by phenotyping two recombinants inbred line populations, Sha x Col and Sha x Ler. Four QTLs controlling germination under salt were detected in the Sha x Col population. Interestingly, only one allelic combination at these four QTLs inhibits germination under salt stress, implying strong epistatic interactions between them. In this interacting context, we confirmed the effect of one QTL by phenotyping selected heterozygous inbred families. We also showed that this QTL is involved in the control of germination under other stress conditions such as KCl, mannitol, cold, glucose and ABA. Our data highlights the presence of a genetic network which consists of four interacting QTLs and controls germination under limiting environmental conditions

Glutathione Transferase from Trichoderma virens Enhances Cadmium Tolerance without Enhancing Its Accumulation in Transgenic Nicotiana tabacum

BACKGROUND: Cadmium (Cd) is a major heavy metal pollutant which is highly toxic to plants and animals. Vast agricultural areas worldwide are contaminated with Cd. Plants take up Cd and through the food chain it reaches humans and causes toxicity. It is ideal to develop plants tolerant to Cd, without enhanced accumulation in the edible parts for human consumption. Glutathione transferases (GST) are a family of multifunctional enzymes known to have important roles in combating oxidative stresses induced by various heavy metals including Cd. Some GSTs are also known to function as glutathione peroxidases. Overexpression/heterologous expression of GSTs is expected to result in plants tolerant to heavy metals such as Cd. RESULTS: Here, we report cloning of a glutathione transferase gene from Trichoderma virens, a biocontrol fungus and introducing it into Nicotiana tabacum plants by Agrobacterium-mediated gene transfer. Transgenic nature of the plants was confirmed by Southern blot hybridization and expression by reverse transcription PCR. Transgene (TvGST) showed single gene Mendelian inheritance. When transgenic plants expressing TvGST gene were exposed to different concentrations of Cd, they were found to be more tolerant compared to wild type plants, with transgenic plants showing lower levels of lipid peroxidation. Levels of different antioxidant enzymes such as glutathione transferase, superoxide dismutase, ascorbate peroxidase, guiacol peroxidase and catalase showed enhanced levels in transgenic plants expressing TvGST compared to control plants, when exposed to Cd. Cadmium accumulation in the plant biomass in transgenic plants were similar or lower than wild-type plants. CONCLUSION: The results of the present study suggest that transgenic tobacco plants expressing a Trichoderma virens GST are more tolerant to Cd, without enhancing its accumulation in the plant biomass. It should be possible to extend the present results to crop plants for developing Cd tolerance and in limiting Cd availability in the food chain

A cotton miRNA is involved in regulation of plant response to salt stress

The present study functionally identified a new microRNA (microRNA ovual line 5, miRNVL5) with its target gene GhCHR from cotton (Gossypium hirsutum). The sequence of miRNVL5 precursor is 104 nt long, with a well developed secondary structure. GhCHR contains two DC1 and three PHD Cys/His-rich domains, suggesting that GhCHR encodes a zinc-finger domain-containing transcription factor. miRNVL5 and GhCHR express at various developmental stages of cotton. Under salt stress (50–400 mM NaCl), miRNVL5 expression was repressed, with concomitant high expression of GhCHR in cotton seedlings. Ectopic expression of GhCHR in Arabidopsis conferred salt stress tolerance by reducing Na+ accumulation in plants and improving primary root growth and biomass. Interestingly, Arabidopsis constitutively expressing miRNVL5 showed hypersensitivity to salt stress. A GhCHR orthorlous gene At2g44380 from Arabidopsis that can be cleaved by miRNVL5 was identified by degradome sequencing, but no confidential miRNVL5 homologs in Arabidopsis have been identified. Microarray analysis of miRNVL5 transgenic Arabidopsis showed six downstream genes (CBF1, CBF2, CBF3, ERF4, AT3G22920, and AT3G49200), which were induced by salt stress in wild-type but repressed in miRNVL5-expressing Arabidopsis. These results indicate that miRNVL5 is involved in regulation of plant response to salt stress

Enhancement of stress tolerance in transgenic tobacco plants constitutively expressing AtIpk2β, an inositol polyphosphate 6-/3-kinase from Arabidopsis thaliana

Inositol phosphates (IPs) and their turnover products have been implicated to play important roles in stress signaling in eukaryotic cells. In higher plants genes encoding inositol polyphosphate kinases have been identified previously, but their physiological functions have not been fully resolved. Here we expressed Arabidopsis inositol polyphosphate 6-/3-kinase (AtIpk2β) in two heterologous systems, i.e. the yeast Saccharomycescerevisiae and in tobacco (Nicotiana tabacum), and tested the effect on abiotic stress tolerance. Expression of AtIpk2β rescued the salt-, osmotic- and temperature-sensitive growth defects of a yeast mutant strain (arg82Δ) that lacks inositol polyphosphate multikinase activity encoded by the ARG82/IPK2 gene. Transgenic tobacco plants constitutively expressing AtIpk2β under the control of the Cauliflower Mosaic Virus 35S promoter were generated and found to exhibit improved tolerance to diverse abiotic stresses when compared to wild type plants. Expression patterns of various stress responsive genes were enhanced, and the activities of anti-oxidative enzymes were elevated in transgenic plants, suggesting a possible involvement of AtIpk2β in plant stress responses

Genetic Variation of HvCBF Genes and Their Association with Salinity Tolerance in Tibetan Annual Wild Barley

The evaluation of both the genetic variation and the identification of salinity tolerant accessions of Tibetan annual wild barley (hereafter referred to as Tibetan barley) (Hordeum vulgare L. ssp. Spontaneum and H. vulgare L. ssp. agriocrithum) are essential for discovering and exploiting novel alleles involved in salinity tolerance. In this study, we examined tissue dry biomass and the Na+ and K+ contents of 188 Tibetan barley accessions in response to salt stress. We investigated the genetic variation of transcription factors HvCBF1, HvCBF3 and HvCBF4 within these accessions, conducting association analysis between these three genes and the respective genotypic salt tolerance. Salt stress significantly reduced shoot and root dry weight by 27.6% to 73.1% in the Tibetan barley lines. HvCBF1, HvCBF3 and HvCBF4 showed diverse sequence variation in amplicon as evident by the identification of single nucleotide polymorphisms (SNPs) and 3, 8 and 13 haplotypes, respectively. Furthermore, the decay of Linkage disequilibrium (LD) of chromosome 5 was 8.9 cM (r2<0.1). Marker bpb-4891 and haplotype 13 (Ps 610) of the HvCBF4 gene were significantly (P<0.05) and highly significantly (P<0.001) associated with salt tolerance. However, HvCBF1 and HvCBF3 genes were not associated with salinity tolerance. The accessions from haplotype 13 of the HvCBF4 gene showed high salinity tolerance, maintaining significantly lower Na+/K+ ratios and higher dry weight. It is thus proposed that these Tibetan barley accessions could be of value for enhancing salinity tolerance in cultivated barley