Dehydrin Protein Research Articles

Overexpression of soybean GmDHN9 gene enhances drought resistance of transgenic Arabidopsis

ABSTRACT Soybean is one of the important oil crops and a major source of protein and lipids. Drought can cause severe soybean yields. Dehydrin protein (DHN) is a subfamily of LEA proteins that play an important role in plant responses to abiotic stresses. In this study, the soybean GmDHN9 gene was cloned and induced under a variety of abiotic stresses. Results showed that the GmDHN9 gene response was more pronounced under drought induction. Subcellular localization results indicated that the protein was localized in the cytoplasm. The role of transgenic Arabidopsis plants in drought stress response was further studied. Under drought stress, the germination rate, root length, chlorophyll, proline, relative water content, and antioxidant enzyme content of transgenic Arabidopsis thaliana transgenic genes were higher than those of wild-type plants, and transgenic plants contained less O2 −, H2O2 and MDA contents. In short, the GmDHN9 gene can regulate the homeostasis of ROS and enhance the drought resistance of plants.

Heterologous overexpression of Tawzy1-2 gene encoding an SK3 dehydrin enhances multiple abiotic stress tolerance in Escherichia coli and Nicotiania benthamiana.

The nuclear localized TaWZY1-2 helps plants resist abiotic stress by preserving the cell's ability to remove reactive oxygen species and decrease lipid oxidation under such conditions. In light of the unpredictable environmental conditions in which food crops grow, precise strategies must be developed by crops to effectively cope with abiotic stress and minimize damage over their lifespan. A key component in this endeavor is the group II of late embryogenesis abundant (LEA) proteins, known as dehydrins, which play crucial roles in enhancing the tolerance of plants to abiotic stress. Tawzy1-2 is a dehydrin-encoding gene which is constitutively expressed in various tissues of wheat. However, the biological function of TaWZY1-2 is not yet fully understood. In this study, TaWZY1-2 was isolated and identified in the wheat genome, and its functional role in conferring tolerance to abiotic stresses was detected in both prokaryotic and eukaryotic cells. Results showed that TaWZY1-2 is a nuclear localized hydrophilic protein that accumulates in response to multiple stresses. Escherichia coli cells expressing TaWZY1-2 showed enhanced tolerance to multiple stress conditions. Overexpression of TaWZY1-2 in Nicotiania benthamiana improved growth, germination and survival rate of the transgenic plants exposed to four kinds of abiotic stress conditions. Our results show that Tawzy1-2 transgenic plants exhibit improved capability in clearing reactive oxygen species and reducing lipid degradation, thereby enhancing their resistance to abiotic stress. This demonstrates a significant role of TaWZY1-2 in mitigating abiotic stress-induced damage. Consequently, these findings not only establish a basis for future investigation into the functional mechanism of TaWZY1-2 but also contribute to the expansion of functional diversity within the dehydrin protein family. Moreover, they identify potential candidate genes for crop optimization.

The positive impact of the NtTAS14-like1 gene on osmotic stress response in Nicotiana tabacum.

NtTAS14-like1 enhances osmotic tolerance through coordinately activating the expression of osmotic- and ABA-related genes. Osmotic stress is one of the most important limiting factors for tobacco (Nicotiana tabacum) growth and development. Dehydrin proteins are widely involved in plant adaptation to osmotic stress, but few of these proteins have been functionally characterized in tobacco. Here, to identify genes required for osmotic stress response in tobacco, an encoding dehydrin protein gene NtTAS14-like1 was isolated based on RNA sequence data. The expression of NtTAS14-like1 was obviously induced by mannitol and abscisic acid (ABA) treatments. Knock down of NtTAS14-like1 expression reduced osmotic tolerance, while overexpression of NtTAS14-like1 conferred tolerance to osmotic stress in transgenic tobacco plants, as determined by physiological analysis of the relative electrolyte leakage and malonaldehyde accumulation. Further expression analysis by quantitative real-time PCR indicated that NtTAS14-like1 participates in osmotic stress response possibly through coordinately activating osmotic- and ABA-related genes expression, such as late embryogenesis abundant (NtLEA5), early responsive to dehydration 10C (NtERD10C), calcium-dependent protein kinase 2 (NtCDPK2), ABA-responsive element-binding protein (NtAREB), ABA-responsive element-binding factor 1 (NtABF1), dehydration-responsive element-binding genes (NtDREB2A), xanthoxin dehydrogenase/reductase (NtABA2), ABA-aldehyde oxidase 3 (NtAAO3), 9-cis-epoxycarotenoid dioxygenase (NtNCED3). Together, this study will facilitate to improve our understandings of molecular and functional properties of plant TAS14 proteins and to improve genetic evidence on the involvement of the NtTAS14-like1 in osmotic stress response of tobacco.

Overexpression of ZmDHN15 Enhances Cold Tolerance in Yeast and Arabidopsis.

Maize (Zea mays L.) originates from the subtropical region and is a warm-loving crop affected by low-temperature stress. Dehydrin (DHN) protein, a member of the Group 2 LEA (late embryogenesis abundant proteins) family, plays an important role in plant abiotic stress. In this study, five maize DHN genes were screened based on the previous transcriptome sequencing data in our laboratory, and we performed sequence analysis and promoter analysis on these five DHN genes. The results showed that the promoter region has many cis-acting elements related to cold stress. The significantly upregulated ZmDHN15 gene has been further screened by expression pattern analysis. The subcellular localization results show that ZmDHN15 fusion protein is localized in the cytoplasm. To verify the role of ZmDHN15 in cold stress, we overexpressed ZmDHN15 in yeast and Arabidopsis. We found that the expression of ZmDHN15 can significantly improve the cold resistance of yeast. Under cold stress, ZmDHN15-overexpressing Arabidopsis showed lower MDA content, lower relative electrolyte leakage, and less ROS (reactive oxygen species) when compared to wild-type plants, as well as higher seed germination rate, seedling survival rate, and chlorophyll content. Furthermore, analysis of the expression patterns of ROS-associated marker genes and cold-response-related genes indicated that ZmDHN15 genes play an important role in the expression of these genes. In conclusion, the overexpression of the ZmDHN15 gene can effectively improve the tolerance to cold stress in yeast and Arabidopsis. This study is important for maize germplasm innovation and the genetic improvement of crops.

Transcriptomic analysis reveals the mechanism of low/high temperature resistance in an outstanding diet alga Nannochloropsis oceanica

Low/high temperature stress is one of the key abiotic factors restricting the implementation of microalgal large-scale production. Metabolism, involved in abiotic stress resistances, have been reported in plants, but the related research in microalgae is far behind. This study analyzed the transcriptional changes in Nannochloropsis oceanica under low and high temperature stresses by RNA-seq technology. Results indicated that the total differentially expressed gene (DEG) amount under high temperature stress was much more than low temperature stress. GO enrichment analysis suggested that DEGs under low temperature were mainly concentrated in protein modification and phosphorus metabolic process, while transmembrane transport, lipid localization and transport, and cellular homeostasis were the most enriched terms under high temperature. Moreover, “membrane” was the most enriched term in both low and high temperature groups. The gene expressions of 37 stress resistance-related enzymes/proteins (172 genes) were analyzed concretely in N. oceanica for the first time. Results revealed that ornithine decarboxylase (ODC) was significantly down-regulated, while choline dehydrogenase (CHDH), late embryogenesis abundant (LEA), dehydrin (DHN), chitinase (CHI), calmodulin (CaM), and lipid transfer protein (LTP) were extremely up-regulated under low temperature. Under high temperature, more genes with drastic changes were identified. Superoxide dismutase (SOD), ascorbate peroxidase (APX), glutathione synthetase (GSS), and glyoxalase II (Gly II) exhibited down-regulated significantly, while glutathione S-transferase (GST), CHDH, L-asparaginase (L-ASP), LEA, DHN, early response to dehydration protein (ERD), heat shock protein (HSP), and CaM were dramatically up-regulated. No significant changes were detected in glutathione peroxidase (GPX), tocopherol cyclase (TC), gamma-glutamylcysteine synthetase (γ-GCS), pyrroline‐5–carboxylate reductase (P5CR), asparagine synthetase (ASN ), succinate-semialdehyde dehydrogenase (SSADH), pyruvate decarboxylase (PDC), and S-adenosylmethionine synthetase (SAMS) under low or high temperature. Furthermore, 166 transcription factor (TF) genes related to plant stress resistance were characterized and categorized into 20 TF families. MYB was the most abundant and active one, revealing that the MYB family played a critical role of temperature resistance in N. oceanica. These new findings could benefit the understanding of the temperature resistance mechanism in Nannochloropsis. The significantly changed genes might be ideal candidates for breeding elite Nannochloropsis strains with stronger resistance to temperature stress using genetic engineering methods.

SsDHN, a dehydrin protein from Suaeda salsa, enhances salt stress tolerance in transgenic tobacco plants

SsDHN, a dehydrin protein from Suaeda salsa, enhances salt stress tolerance in transgenic tobacco plants

SDS induced structural transition and cryoprotective function of plant IDP, dehydrin and its mutants

Dehydrin is an IDP that belongs to the Late Embryogenesis Abundant protein family which has been shown to have important role in abiotic stress tolerance. Here, we investigated the DHN1 (dehydrin) protein from Zea mays (167 aa). The role of conserved segment and its folding in presence of membrane is vital for understanding the functional mechanism of DHN1. To investigate the structural change and cryoprotection mechanism of DHN1, three mutants: DHN1 CW1 (Trp122–Cys62), DHN1 CW2 (Trp132–Cys62) and DHN1 W3 (Trp3) were generated using site directed mutagenesis.

AnDHN, a Dehydrin Protein From Ammopiptanthus nanus, Mitigates the Negative Effects of Drought Stress in Plants.

Dehydrins (DHNs) play crucial roles in a broad spectrum of abiotic stresses in model plants. However, the evolutionary role of DHNs has not been explored, and the function of DHN proteins is largely unknown in Ammopiptanthus nanus (A. nanus), an ancient and endangered legume species from the deserts of northwestern China. In this study, we isolated a drought-response gene (c195333_g1_i1) from a drought-induced RNA-seq library of A. nanus. Evolutionary bioinformatics showed that c195333_g1_i1 is an ortholog of Arabidopsis DHN, and we renamed it AnDHN. Moreover, DHN proteins may define a class of proteins that are evolutionarily conserved in all angiosperms that have experienced a contraction during the evolution of legumes. Arabidopsis plants overexpressing AnDHN exhibited morpho-physiological changes, such as an increased germination rate, higher relative water content (RWC), higher proline (PRO) content, increased peroxidase (POD) and catalase (CAT) activities, lower contents of malondialdehyde (MDA), H2O2 and O2–, and longer root length. Our results showed that the transgenic lines had improved drought resistance with deep root system architecture, excellent water retention, increased osmotic adjustment, and enhanced reactive oxygen species (ROS) scavenging. Furthermore, the transgenic lines also had enhanced salt and cold tolerance. Our findings demonstrate that AnDHN may be a good candidate gene for improving abiotic stress tolerance in crops.Key Message: Using transcriptome analysis in Ammopiptanthus nanus, we isolated a drought-responsive gene, AnDHN, that plays a key role in enhancing abiotic stress tolerance in plants, with strong functional diversification in legumes.

Molecular characterization and expression studiesof Eucalyptus globulus stress-responsive gene DHN-10

The Eucalyptus globulus is one of the important forest tree species which reveals enhanced tolerance to multiple abiotic stresses. The enhanced tolerance of E. globulus against low temperature is linked to the presence of enhanced accumulation of lysine-rich dehydrin proteins. The accumulation of dehydrin protein of Mr 10 kDa in response to multiple abiotic stresses prevents cells from dehydration. It is encoded by a novel dehydrin-10 (DHN-10) gene. In the present study, we have used the DHN-10 gene of E. globulus of Pakistani origin, which was cloned in a bacterial expression vector pET30(a) and sequenced. We have found the 315 bp long coding sequence of this gene which has been enrolled as Eucalyptus GZJ-2018 DHN-10 gene under GenBank accession number MG948256.1. The in silico studies have identified several differences of this gene from the earlier enrolled DHN-10 genes: the studied gene possessed higher amphipathic character because of the presence of five additional electrically charged amino acids (two positively charged histidine, one negatively charged glutamate, and two negatively charged aspartate residues) and one extra lysine residue. The studied DHN-10 gene has been successfully expressed in the BL21-DE3 expression strain of Escherichia coli and 10 kDa protein has been detected on the nitrocellulose membrane. Our study is the first report of the sub-cloning of the DHN-10 gene and its expression outside the Eucalyptus cell.

Quantitative analysis of differential dehydrin regulation in pine and spruce seedlings under water deficit

Dehydrins are well-known components of plant responses to different stresses that cause dehydration, including drought, freezing, salinity, etc. In conifers, the dehydrin gene family is very large, implying that the members of this family have important physiological functions in conifer stress tolerance. However, dehydrin gene expression displays a wide range of responses to stress, from thousand-fold increased expression to decreased expression, and it is generally unknown how regulatory systems are connected at the mRNA and protein levels. Therefore, we studied these aspects of dehydrin regulation in Scots pine (Pinus sylvestris L.) and Norway spruce (Picea abies (L.) H. Karst) seedlings under polyethylene glycol 6000-induced osmotic stress ranging from relatively low (culture medium water potential of −0.15 MPa) to very high (−1.0 MPa) intensities. In pine, the major dehydrin protein was Dhn1 in both the roots and needles, and in spruce, two isoforms of the Dhn4 protein were the major dehydrins; both of these proteins are AESK-type dehydrins. The genes encoding these major proteins were highly expressed even under control conditions; surprisingly, we also observed several highly expressed dehydrin genes that were not abundantly translated. Under osmotic stress, the most prominent expression changes were observed for the dehydrin genes with low basal expression levels, whereas highly expressed genes generally demonstrated rather modest changes in expression. We report proposed constitutive physiological functions of the AESK-type dehydrins in Pinaceae plants.

Functional characterization of VviDHN2 and VviDHN4 dehydrin isoforms from Vitis vinifera (L.): An in silico and in vitro approach

Dehydrins, a family of hydrophilic and intrinsically disordered proteins, are a subgroup of late embryogenesis abundant proteins that perform different protective roles in plants. Although the transition from a disordered to an ordered state has been associated with dehydrin function or interactions with specific partner molecules, the question of how the primary and secondary dehydrin protein structure is related to specific functions or target molecule preferences remains unresolved. This work addresses the in silico sequencing analysis and in vitro functional characterization of two dehydrin isoforms, VviDHN2 and VviDHN4, from Vitis vinifera. Conformational changes suggest potential interactions with a broad range of molecules and could point to more than one function. The in silico analysis showed differences in conserved segments, specific amino acid binding sequences, heterogeneity of structural properties and predicted sites accessible for various post-translational modifications between the sequence of both dehydrins. Moreover, in vitro functional analysis revealed that although they both showed slight antifungal activity, only VviDHN4 acts as a molecular shield that protects proteins from freezing and dehydration. VviDHN4 also demonstrated high potential as a chaperone and reactive oxygen species scavenger, in addition to presenting antifreeze activity, all of which confirms its multifunctional nature. Our findings highlight the significant role of Y-segments and the differential and specific amino acid composition of less conserved segments that are rich in polar/charged residues between S- and K-segments, coupled with post-translational modifications, in modulating and switching dehydrin biological function.

Isolation and characterization of Dehydrin promoter region from sugarcane (Saccharum officinarum L.)

The development of molecular biology techniques nowadays has enabled to engineer drought tolerant sugarcane by genetic engineering to accelerate the breeding program. Dehydrin (DHN) is known to have an important role in plant response and adaptation to abiotic stresses (drought, high salinity, cold, heat, etc.). While plant tissues are subjected to drought stress (dehydration), DHN protein is accumulated to high content throughout all vegetative or generative tissues. The research aimed to isolate and characterize the DHN promoter from sugarcane that can be used as transformation material in generating drought tolerant sugarcane. Specific primers for DHN promoter amplification were designed and DHN promoter region was successfully isolated by PCR cloning method. Two putative promoter sequences were identified namely Pr-1DHNSo and Pr-2DHNSo. In silicoanalyses were carried out and cis-regulatory elements motifs that play a role in adaptation on abiotic stress as well as biotic stress including ABRE, MBS, CGTCA-motif, TGACG-motif, GARE-motif, P-box TCA-element and Box-W1 were identified. The promoter Pr-1DHNSo was then cloned into pBI121 expression vector by Overlap Extention PCR (OE-PCR) for further characterization. Functional test of the promoter construct pBI- Pr-1DHNSo was conducted through Agrobacterium transformation into sugarcane calli. GUS assay and PCR analysis showed that the DHN promoter was transformed and expressed in the sugarcane calli.

Functional Analysis of <i>Tadhn1312</i> by Virus-Induced Gene Silencing (VIGS) in Common Wheat (<i>Triticum aestivum</i> L.)

Virus-induced gene silencing (VIGS) is an important tool for gene function analysis in plants. In the present study, the function identification of a candidate gene named Tadhn1312 encoding Dehydrin protein obtained from the high-throughput transcriptome was carried out by VIGS. Leaves appeared the chlorotic phenotype and the transcript level of TaPDS decreased rapidly at 7 days after the inoculation by BSMV: TaPDS, which indicated that the virus had successfully infected wheat leaves and the BSMV system was efficient. The chlorophyll content of leaves with BSMV: TaPDS and BSMV: Tadhn1312 inoculation decreased at 7 days, and reached significant level compared with the control. After the inoculation by BSMV: Tadhn1312, the transcript level of Tadhn1312 was rapidly decreased at 7 days, and reached the minimum value at 21 days, indicated that Tadhn1312 had been silenced. The spike differentiation procession of wheat plants inoculated by BSMV: Tadhn1312 was late than that inoculated by BSMV: 00. The result showed that the silencing of Tadhn1312 prolonged the spike differentiation process, illustrating that Tadhn1312 gene was involved in the spike differentiation process of wheat directly or indirectly.

Dehydration Sensitivity at the Early Seedling Establishment Stages of the European Beech (Fagus sylvatica L.)

Shortage of water is a limiting factor for the growth and development of plants, particularly at early developmental stages. We focused on the European beech (Fagus sylvatica L.), which produces seeds and further seedlings in large intervals of up to ten years. To explore the beech seedling establishment process, six stages referring to embryo expansion were studied to determine sensitivity to dehydration. The characterization of the response of elongating embryonic axes and cotyledons included a viability test before and after dehydration and measurement of the amounts of electrolyte leakage, concentration, and arrangement of storage materials, changes in chaperone proteins related to water deficit, and accumulation of hydrogen peroxide and superoxide anion radicals. Elongating embryonic axes and cotyledons differed in water content, dehydration rates, membrane permeability before and after dehydration, protein, and lipid decomposition pattern, and amount of 44-kDa dehydrin and 22-kDa small heat shock protein (sHSP). Protruding embryonic axes were more sensitive to dehydration than cotyledons, although dehydration caused transient reinduction of three dehydrin-like proteins and sHSP synthesis, which accompany desiccation tolerance. Extended deterioration, including overproduction of hydrogen peroxide and depletion of superoxide anion radicals, was reported in dehydrated embryonic axes longer than 10 mm characterized by highly elevated cellular leakage. The apical part elongating embryonic axes consisting of the radicles was the most sensitive part of the seed to dehydration, and the root apical meristem area was the first to become inviable. The effects of severe dehydration involving ROS imbalance and reduced viability in beech seedlings with embryonic axes longer than 10 mm might help to explain the difficulties in beech seedling establishment observed in drought-affected environments. The conversion of environmental drought into climate-originated oxidative stress affecting beech seedling performance is discussed in this report.

Diverse responsiveness of dehydrin genes to abscisic acid and water stress treatments in cucumber F1 cultivar hybrids

ABSTRACTExpression of putative dehydrin (DHN) genes of cucumber (Cucumis sativus L.) was investigated under water deficit stress and after external abscisic acid (ABA) treatment. Three typically open field cultivated along with four greenhouse grown F1 cultivar hybrids were used in the experiments. ‘K’ segment specific Western blot was applied to characterise DHN proteins expressed in water deprivation while transcription of selected DHN genes was studied by RT-PCR and RT-qPCR. Water-deficit induced differential accumulation of DHN polypeptides in heat stable protein fractions of leaf samples. Greenhouse cultivated hybrids exhibited more pronounced induction of low molecular weight DHN proteins under this condition. The basal transcript level of the YnSKn type DHN2 gene in the absence of treatments was lower than that of the SKn type DHN3. DHN2 was highly inducible by water stress and ABA application. Transcription of DHN3 was moderately increased by desiccation in most hybrids, while it was only slightly responsive to ABA in all genotypes. F1 hybrids of the same cultivation type did not form distinct groups according to their transcriptional responses to either treatments. Comparison of the expression patterns at the transcript and protein levels pointed to a possibility of translational and/or post-translational regulation of DHN protein abundance.

Transcriptome Analysis and Differential Expression in Tall Fescue Harboring Different Endophyte Strains in Response to Water Deficit

Two tall fescue [Lolium arundinaceum (Schreb.) Darbysh. = Schedonorus arundinaceus (Schreb.) Dumort. = Festuca arundinacea var. arundinacea Schreb.] plant genotypes with an Epichloë coenophiala (Morgan-Jones & W. Gams) C.W. Bacon & Schardl common toxic endophyte (CTE), one with a nontoxic strain (NTE19) and one with another Epichloë species (FaTG-4) were evaluated and compared with their respective endophyte-free clones for responses to water-deficit stress in the greenhouse. One of the plant genotypes (P27) showed a positive effect of its CTE strain on tiller production after stress and resumed watering. In transcriptome analysis of the pseudostems (leaf sheath whorls), differentially expressed genes (DEGs) were defined as having at least twofold expression difference and false discovery rate (FDR) < 0.05 in comparisons of water treatment (stressed or watered), endophyte presence or absence, or both. Stress affected 38% of the plant transcripts including those for the expected stress-response pathways. The DEGs affected by endophyte in stressed plants were unique to individual plant genotypes. In unstressed plants, endophyte presence tended to reduce expression of genes putatively for defense against fungi, but in unstressed P27 endophyte presence there was enhanced expression of dehydrin and heat shock protein genes. Our results indicated subtle and variable effects of endophytes on tall fescue gene expression; where the endophyte confers protection, its effects on plant gene expression may help prime the plant for stress resistance.

Relationship Between Dehydrin Accumulation and Winter Survival in Winter Wheat and Barley Grown in the Field.

Low temperatures represent a crucial environmental factor determining winter survival (WS) of barley and wheat winter-type varieties. In laboratory experiments, low temperatures induce an active plant acclimation response, which is associated with an enhanced accumulation of several stress-inducible proteins including dehydrins. Here, dehydrin accumulations in sampled wheat (WCS120 protein family, or WCS120 and WDHN13 transcripts) and barley (DHN5 protein) varieties grown in two locations for two winters were compared with the variety WS evaluated by a provocation wooden-box test. A high correlation between dehydrin transcripts or protein relative accumulation and variety WS score was found only in samples taken prior vernalization fulfillment, when high tolerant varieties accumulated dehydrins earlier and to higher level than less tolerant varieties, and the plants have not yet been vernalized. After vernalization fulfillment, the correlation was weak, and the apical development indicated that plants reached double ridge (DR) in barley or stayed before DR in wheat. Dehydrin proteins and transcripts can be thus used as reliable markers of wheat or barley variety winter hardiness in the field conditions; however, only at the beginning of winter, when the plants have not yet finished vernalization. In wheat, a higher correlation was obtained for the total amount of dehydrins than for the individual dehydrin proteins.HIGHLIGHTS-More tolerant winter-type wheat and barley plants reveal higher threshold induction temperatures for dehydrin accumulation in comparison to less tolerant varieties. Thus, more tolerant winter cereals have higher dehydrin levels than the less tolerant ones upon the same ambient temperature in November samplings.-A significant correlation between dehydrin transcript/protein accumulation and winter survival was found in both winter wheat and winter barley plants in the field conditions, but only prior to vernalization fulfillment.

Characterization of Dehydrin protein, CdDHN4-L and CdDHN4-S, and their differential protective roles against abiotic stress in vitro

BackgroundDehydrins play positive roles in regulating plant abiotic stress responses. The objective of this study was to characterize two dehydrin genes, CdDHN4-L and CdDHN4-S, generated by alternative splicing of CdDHN4 in bermudagrass.ResultsOverexpression of CdDHN4-L with φ-segment and CdDHN4-S lacking of φ-segment in Arabidopsis significantly increased tolerance against abiotic stresses. The growth phenotype of Arabidopsis exposed to NaCl at 100 mM was better in plants overexpressing CdDHN4-L than those overexpressing CdDHN4-S, as well as better in E.coli cells overexpressing CdDHN4-L than those overexpressing CdDHN4-S in 300 and 400 mM NaCl, and under extreme temperature conditions at − 20 °C and 50 °C. The CdDHN4-L had higher disordered characterization on structures than CdDHN4-S at temperatures from 10 to 90 °C. The recovery activities of lactic dehydrogenase (LDH) and alcohol dehydrogenase (ADH) in presence of CdDHN4-L and CdDHN4-S were higher than that of LDH and ADH alone under freeze-thaw damage and heat. Protein-binding and bimolecular fluorescence complementation showed that both proteins could bind to proteins with positive isoelectric point via electrostatic forces.ConclusionsThese results indicate that CdDHN4-L has higher protective ability against abiotic stresses due to its higher flexible unfolded structure and thermostability in comparison with CdDHN4-S. These provided direct evidence of the function of the φ-segment in dehydrins for protecting plants against abiotic stress and to show the electrostatic interaction between dehydrins and client proteins.

PicW2 from Picea wilsonii: preparation, purification, crystallization and X-ray diffraction analysis.

Low temperature is a major limiting factor for plant growth and development. Dehydrin proteins are generally induced in response to low-temperature stress. In previous research, a full-length dehydrin gene, PicW2, was isolated from Picea wilsonii and its expression was associated with hardiness to cold. In order to gain insight into the mechanism of low-temperature tolerance by studying its three-dimensional crystal structure, prokaryotically expressed PicW2 dehydrin protein was purified using chitosan-affinity chromatography and gel filtration, and crystallized using the vapour-diffusion method. The crystal grew in a condition consisting of 0.1 M HEPES pH 8.0, 25%(w/v) PEG 3350 using 4 mg ml-1 protein solution at 289 K. X-ray diffraction data were collected from a crystal at 100 K to 2.82 Å resolution. The crystal belonged to space group C121, with unit-cell parameters a = 121.55, b = 33.26, c = 73.39 Å, α = γ = 90.00, β= 109.01°. The asymmetric unit contained one molecule of the protein, with a corresponding Matthews coefficient of 2.87 Å3 Da-1 and a solvent content of 57.20%. Owing to a lack of structures of homologous dehydrin proteins, molecular-replacement trials failed. Data collection for selenium derivatization of PicW2 and crystal structure determination is currently in progress.

Genome-wide identification and molecular characterization of abiotic stress related gene families in chickpea and other plants

Various abiotic stresses such as high salinity and drought affect various aspects of plant physiology and metabolism and as a result, severely diminish crop productivity in some area. In chickpea, three genomes including both ICC4985 (desi type) and CDC frontier (kabuli type) from Cicer arietinum as well as the wild species PI489777 ( Cicer reticulatum ) have been sequenced, which provided a basis to further explore the mechanism on abiotic stress tolerance. In this study, we genome-widely identified 6 abiotic stress related gene families in these three chickpea genotypes and compared these families with those from other plants. These 6 gene families include high-affinity K + transporter (HKT), K + uptake permease (KUP), cation proton antiporters (CPA), vacuolar H(+)-translocating inorganic pyrophosphatase (VHP), dehydrin (DHN) and stress responsive protein (SRP) families. Our data showed that three chickpea and other plant genomes encoded different sizes of family members and exhibited obvious difference in the family expansion and evolutionary selection. Both segmental and tandem duplication significantly contributed to the CPA family expansion. Although the CPA family showed the strong purifying selection within a species, positive selection was detected among different species of chickpea. Expression analysis showed that most of these family members were expressed in multiple tissues and no tissue-preferred genes were detected. Furthermore, up to 17.9-100% of these family members were up- or down-regulated by either high salinity or drought stress, suggesting their important roles in abiotic stress signaling pathways.