Cold Stress Conditions Research Articles

PKS5 Confers Cold Tolerance by Controlling Stomatal Movement and Regulating Cold-Responsive Genes in Arabidopsis.

Cold stress limits plant growth and development; however, the precise mechanisms underpinning plant acclimation to cold stress remain largely unknown. In this study, the Ser/Thr protein kinase SOS2-LIKE PROTEIN KINASE5 (PKS5) was shown to play a positive role in plant responses to cold stress. A PKS5 loss-of-function mutant (pks5-1) exhibited elevated sensitivity to cold stress, as well as a lower survival rate and increased ion leakage. Conversely, PKS5 gain-of-function mutants (pks5-3, pks5-4) were more tolerant to cold stress and exhibited higher survival rates and decreased ion leakage. Stomatal aperture analysis revealed that stomatal closure was slower during the first 25 min after cold exposure in pks5-1 compared to wild-type, whereas pks5-3 and pks5-4 displayed accelerated stomatal closure over the same time period. Further stomatal aperture analysis under an abscisic acid (ABA) treatment showed slower closure in pks5-1 and more rapid closure in pks5-3 and pks5-4. Finally, expression levels of cold-responsive genes were regulated by PKS5 under cold stress conditions, while cold stress and ABA treatment can regulate PKS5 expression. Taken together, these results suggest that PKS5 plays a positive role in short-term plant acclimation to cold stress by regulating stomatal aperture, possibly via ABA pathways, and in long-term acclimation by regulating cold-responsive genes.

Influence of Seed Soaking and Foliar Application Using Ozonated Water on Two Sweet Pepper Hybrids under Cold Stress

The harmful impacts of ozone (O3) on plant development and productivity have been excessively studied. Furthermore, the positive influences of its low concentrations still need to be explored further. The present study was performed to assess the impact of low concentrations of O3 on two sweet pepper hybrids under cold stress. The ozonated water was utilized for seed soaking or foliar application at concentrations of 0, 10, 20, 30, and 40 ppm. Seed soaking using ozonated water for 1 h was compared to soaking in distilled water as a control. Moreover, exogenously ozonated water was sprayed thrice at three-day intervals compared with untreated control. The differences between the applied methods (seed soaking and foliar application using ozonated water) were not statistically detected in most of the evaluated parameters. On the other hand, the evaluated hybrids displayed significant differences in the studied parameters, with the superiority of the Lirica evident in most germination and seedling growth parameters. Both applied methods significantly improved germination and seedling growth parameters. In particular, the concentration of 40 ppm displayed the highest enhancement of the germination index, coefficient velocity, and seedling quality. In addition, it promoted the seedling maintenance of high relative water content (RWC), chlorophyll, proline, and ascorbate peroxidase activity under cold stress conditions. Moreover, it protected the cell wall from damage by decreasing membrane permeability (MP). Generally, the best results were obtained from 40 ppm followed by 30 ppm of O3 as seed soaking or foliar spray. The results pointed out the possible use of O3 in a low concentration to protect the plants from cold stress during germination and early plant growth.

Characterization of invertase inhibitors (InvInhs) in tea plant, and their potential roles in participating in growth, development and cold response

Invertase inhibitor (InvInh) represses acid invertase (AI) activity though interacting with AI at post-translation level, which plays important roles in vegetative and reproductive processes and stress responses of plant. Our previous studies revealed that CsAI genes were involved in regulating cold response of tea plant, while the post-translation regulation mechanism of AI and the roles of InvInh in tea plant have not been reported yet. Herein, we identified 4 CsInvInhs, including 2 tandem repeat genes (CsInvInh1/2) and 2 segmental duplication genes (CsInvInh3/4), from three sequenced tea plant genomes, and further purified CsInvInh1/2 proteins. Bioinformatics analysis results showed that both of CsInvInh1/2 were located on chromosome (Chr) 5, while CsInvInh3/4 were respectively located on Chr9 and Chr12, which was consistent with the results of inter-species collinearity analysis. Under various abiotic stress conditions, the transcription levels of CsInvInhs were induced with different extent at different treatment time points. In addition, all of CsInvInhs differentially expressed among different tea plant cultivars under drought stress condition. Moreover, CsInvInh1/2 shared similar expression patterns in different tea plant tissues, and under cold stress or cold acclimation conditions, suggesting that CsInvInh1/2 may have functional redundancy in tea plant. Correlation analysis results showed that the changes of CWIN and VIN activities were not significantly correlated with the expression patterns of 5 CsAI genes, while negatively correlated with the expression patterns of 3 CsInvInhs in different tea plant tissues, and under cold stress or cold acclimation conditions, indicating that CsInvInhs participate in growth, development and cold response of tea plant mainly through inhibiting AI activity. These results will provide theoretical foundation for the cold-resistant molecular mechanism of tea plant, and also provide new genetic resources and approaches for cold-resistant molecular breeding of tea plant.

Genome-wide identification and expression analysis of glutathione S-transferase gene family to reveal their role in cold stress response in cucumber.

The plant glutathione S-transferases (GSTs) are versatile proteins encoded by several genes and play vital roles in responding to various physiological processes. Members of plant GSTs have been identified in several species, but few studies on cucumber (Cucumis sativus L.) have been reported. In this study, we identified 46 GST genes, which were divided into 11 classes. Chromosomal location and genome mapping revealed that cucumber GSTs (CsGSTs) were unevenly distributed in seven chromosomes, and the syntenic regions differed in each chromosome. The conserved motifs and gene structure of CsGSTs were analyzed using MEME and GSDS 2.0 online tools, respectively. Transcriptome and RT-qPCR analysis revealed that most CsGST members responded to cold stress. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses for differentially expressed CsGSTs under cold stress revealed that these genes responded to cold stress probably through “glutathione metabolism.” Finally, we screened seven candidates that may be involved in cold stress using Venn analysis, and their promoters were analyzed using PlantCARE and New PLACE tools to predict the factors regulating these genes. Antioxidant enzyme activities were increased under cold stress conditions, which conferred tolerance against cold stress. Our study illustrates the characteristics and functions of CsGST genes, especially in responding to cold stress in cucumber.

Robustness of the honeybee neuro-muscular octopaminergic system in the face of cold stress.

In recent decades, our planet has undergone dramatic environmental changes resulting in the loss of numerous species. This contrasts with species that can adapt quickly to rapidly changing ambient conditions, which require physiological plasticity and must occur rapidly. The Western honeybee (Apis mellifera) apparently meets this challenge with remarkable success, as this species is adapted to numerous climates, resulting in an almost worldwide distribution. Here, coordinated individual thermoregulatory activities ensure survival at the colony level and thus the transmission of genetic material. Recently, we showed that shivering thermogenesis, which is critical for honeybee thermoregulation, depends on octopamine signaling. In this study, we tested the hypothesis that the thoracic neuro-muscular octopaminergic system strives for a steady-state equilibrium under cold stress to maintain endogenous thermogenesis. We can show that this applies for both, octopamine provision by flight muscle innervating neurons and octopamine receptor expression in the flight muscles. Additionally, we discovered alternative splicing for AmOARβ2. At least the expression of one isoform is needed to survive cold stress conditions. We assume that the thoracic neuro-muscular octopaminergic system is finely tuned in order to contribute decisively to survival in a changing environment.

Association between SSR Markers and Phenologic Plus Agronomic Traits in Barley (Hordeum valgare L.) Under Cold Stress Conditions

Association between SSR Markers and Phenologic Plus Agronomic Traits in Barley (Hordeum valgare L.) Under Cold Stress Conditions

Global identification of quantitative trait loci and candidate genes for cold stress and chilling acclimation in rice through GWAS and RNA-seq

Associated analysis of GWAS with RNA-seq had detected candidate genes responsible for cold stress and chilling acclimation in rice. Haplotypes of two candidate genes and geographic distribution were analyzed. To explore new candidate genes and genetic resources for cold tolerance improvement in rice, genome-wide association study (GWAS) mapping experiments with 351 rice core germplasms was performed for three traits (survival rate, shoot length and chlorophyll content) under three temperature conditions (normal temperature, cold stress and chilling acclimation), yielding a total of 134 QTLs, of which 54, 59 and 21 QTLs were responsible for normal temperature, cold stress and chilling acclimation conditions, respectively. Integrated analysis of significant SNPs in 134 QTLs further identified 116 QTLs for three temperature treatments, 53, 43 and 18 QTLs responsible for normal temperature, cold stress and chilling acclimation, respectively, and 2 QTLs were responsible for both cold stress and chilling acclimation. Matching differentially expressed genes from RNA-seq to 43 and 18 QTLs for cold stress and chilling acclimation, we identified 69 and 44 trait-associated candidate genes, respectively, to be classified into six and five groups, particularly involved in metabolisms, reactive oxygen species scavenging and hormone signaling. Interestingly, two candidate genes LOC_Os01g04814, encoding a vacuolar protein sorting-associating protein 4B, and LOC_Os01g48440, encoding glycosyltransferase family 43 protein, showed the highest expression levels under chilling acclimation. Haplotype analysis revealed that both genes had a distinctive differentiation with subpopulation. Haplotypes of both genes with more japonica accessions have higher latitude distribution and higher chilling tolerance than the chilling sensitive indica accessions. These findings reveal the new insight into the molecular mechanism and candidate genes for cold stress and chilling acclimation in rice.

Effect of a Radical Mutation in Plastidic Starch Phosphorylase PHO1a on Potato Growth and Cold Stress Response

The plant response to stresses includes changes in starch metabolism regulated by a complex catalytic network, in which plastidic starch phosphorylase PHO1a is one of the key players. In this study, we used the CRISPR-Cas9 system to edit the PHO1a gene in four potato (Solanum tuberosum L.) cultivars, which resulted in the introduction of a radical mutation, G261V, into the PHO1a functional domain. The mutants had altered morphology and differed from wild-type plants in starch content in the roots and leaves. Exposure to cold stress revealed the differential response of parental and transgenic plants in terms of starch content and the expression of genes coding for β-amylases, amylase inhibitors, and stress-responsive MADS-domain transcription factors. These results suggest that the G261V mutation causes changes in the functional activity of PHO1a, which in turn affect the coordinated operation of starch catabolism enzymes both under normal and cold stress conditions, possibly through differential expression of MADS-domain transcription factors. Our results highlight a critical regulatory role of PHO1a in starch metabolism, root and shoot development, and stress response in potatoes.

Identification of AP2/ERF transcription factors in Tetrastigma hemsleyanum revealed the specific roles of ERF46 under cold stress.

Tetrastigma hemsleyanum (T. hemsleyanum) is a traditional medicinal plant that is widely used in China. Cultivated T. hemsleyanum usually encounters cold stress, limiting its growth and quality at key developmental stages. APETALA2 (AP2)/ethylene-responsive factor (ERF) transcription factors (TFs) comprise one of the largest gene superfamilies in plants and are widely involved in biotic and abiotic stresses. To reveal the roles of AP2/ERF TFs during T. hemsleyanum development, 70 AP2/ERF TFs were identified in T. hemsleyanum. Among them, 18 and 2 TFs were classified into the AP2 and RAV families, respectively. The other 50 TFs belonged to the ERF family and were further divided into the ERF and (dehydration reaction element binding factor) DREB subfamilies. The ERF subfamily contained 46 TFs, while the DREB subfamily contained 4 TFs. Phylogenetic analysis indicated that AP2/ERF TFs could be classified into five groups, in which 10 conserved motifs were confirmed. Several motifs were group- or subgroup-specific, implying that they were significant for the functions of the AP2/ERF TFs of these clades. In addition, 70 AP2/ERF TFs from the five groups were used for an expression pattern analysis under three low-temperature levels, namely, –4, 0, and 4°C. The majority of these AP2/ERF TFs exhibited a positive response to cold stress conditions. Specifically, ThERF5, ThERF31, ThERF46, and ThERF55 demonstrated a more sensitive response to cold stress. Moreover, AP2/ERF TFs exhibited specific expression patterns under cold stress. Transient overexpression and RNA interference indicated that ThERF46 has a specific tolerance to cold stress. These new insights provide the basis for further studies on the roles of AP2/ERF TFs in cold stress tolerance in T. hemsleyanum.

A low concentration of exogenous salicylic acid enhances cold tolerance in Hami melons (Cucumis melo var. saccharinus) by modulating salicylic acid-response CmGST genes

Long-term cold storage of Hami melons (Cucumis melo var. saccharinus) induces severe chilling injuries, downgrading their storage quality. Three different concentrations (1, 3, and 9 mM) of exogenous salicylic acid (SA) were applied under cold stress conditions (0.5 ± 0.5 °C) to prolong the shelf-life of cold-sensitive ‘Golden Empress-308′ (GE-melon) Hami melons. Afterward, the phenotypic index, bioinformatics of CmGSTs, expression patterns of CmGSTs, endogenous SA content, molecular docking, and molecular dynamics simulation were investigated. The phenotypic index indicated that low concentrations (1 and 3 mM) of SA ameliorated chilling injuries, whereas a high concentration of SA (9 mM) treatment adversely affected the GE-melons. The RT-qPCR analysis determined that SA treatment induced five out of seven CmGST-containing SA-response cis-elements. The content of endogenous SA was elevated by exogenous SA. Interestingly, both high and low levels of endogenous SA promoted chilling injuries in GE-melons under cold stress conditions. Furthermore, molecular docking and molecular dynamics simulation signaled that SA bound to the target CmGSTs in a stable conformation. In conclusion, administration of a low concentration of SA enhances cold tolerance in Hami melons by modulating the SA-responsive CmGSTs under cold stress conditions.

Physiological and production performance of maize and soybean in crop-livestock-forest integration system under hydrogel rates and nitrogen and potassium topdressing

The purpose of this research was to determine the effects of copper nanoparticles (CuNP) supplementation on growth performance, intestinal morphology, intraepithelial lymphocytes (IELs), goblet cell (GC) count, stress hormones and oxidant/antioxidant balance of broilers exposed to cold stress. One day old broiler chickens (n=400) were allocated into five different groups and were raised under standard conditions until the 21st day. The five groups were designated as thermo-neutral (TN reared in 26°C), cold stress (CS raised under 13°C±2°C for 8 hours/day from d 22- d 35), CS5 (CS+ 5mg/kg diet CuNP), CS10 (CS+10mg/kg diet CuNP) and CS15 (CS+15mg/kg diet CuNP). On 35th day, 16 birds per group were selected randomly for blood collection and were then humanely sacrificed for tissue sample collection. CS group had a reduced (P<0.05) body weight (BW), villus surface area (VSA), villus height to crypt depth ratio (VH: CD) but had a higher (P<0.05) feed conversion ratio (FCR), goblet cell count, serum corticosterone and malondialdehyde (MDA) levels than the TN group. Inclusion of CuNP in diet linearly increased (P<0.05) BW, VH, VW, CD, VSA and GC count in jejunum and ileum. Conversely, MDA and corticosterone hormone levels and FCR, decreased linearly (P<0.05) with CuNP supplementation. In conclusion, during cold stress conditions, CuNP is a valuable feed additive in broilers with positive effects on growth performance and intestinal morphology.

Effects of Copper nanoparticle supplementation on growth, intestinal morphology and antioxidant status in broilers exposed to cyclic cold stress

The purpose of this research was to determine the effects of copper nanoparticles (CuNP) supplementation on growth performance, intestinalmorphology, intraepithelial lymphocytes (IELs), goblet cell (GC) count, stress hormones and oxidant/antioxidant balance of broilers exposedto cold stress. One day old broiler chickens (n=400) were allocated into five different groups and were raised under standard conditionsuntil the 21st day. The five groups were designated as thermo-neutral (TN reared in 26°C), cold stress (CS raised under 13°C±2°C for8 hours/day from d 22- d 35), CS5 (CS+ 5mg/kg diet CuNP), CS10 (CS+10mg/kg diet CuNP) and CS15 (CS+15mg/kg diet CuNP). On35th day, 16 birds per group were selected randomly for blood collection and were then humanely sacrificed for tissue sample collection.CS group had a reduced (P<0.05) body weight (BW), villus surface area (VSA), villus height to crypt depth ratio (VH: CD) but had ahigher (P<0.05) feed conversion ratio (FCR), goblet cell count, serum corticosterone and malondialdehyde (MDA) levels than the TNgroup. Inclusion of CuNP in diet linearly increased (P<0.05) BW, VH, VW, CD, VSA and GC count in jejunum and ileum. Conversely,MDA and corticosterone hormone levels and FCR, decreased linearly (P<0.05) with CuNP supplementation. In conclusion, during coldstress conditions, CuNP is a valuable feed additive in broilers with positive effects on growth performance and intestinal morphology

Comparative bioinformatics analysis and abiotic stress responses of expansin proteins in Cucurbitaceae members: watermelon and melon.

Watermelon and melon are members of the Cucurbitaceae family including economically significant crops in the world. The expansin protein family, which is one of the members of the cell wall, breaks down the non-covalent bonds between cell wall polysaccharides, causing pressure-dependent cell expansion. Comparative bioinformatics and molecular characterization analysis of the expansin protein family were carried out in the watermelon (Citrullus lanatus) and melon (Cucumis melo) plants in the study. Gene expression levels of expansin family members were analyzed in leaf and root tissues of watermelon and melon under ABA, drought, heat, cold, and salt stress conditions by quantitative real-time PCR analysis. After comprehensive searches, 40 expansin proteins (22 ClaEXPA, 14 ClaEXPLA, and 4 ClaEXPB) in watermelon and 43 expansin proteins (19 CmEXPA, 15 CmEXPLA, 3 CmEXPB, and 6 CmEXPLB) in melon were identified. The greatest orthologous genes were identified with soybean expansin genes for watermelon and melon. However, the latest divergence time between orthologous genes was determined with poplar expansin genes for watermelon and melon expansin genes. ClaEXPA-04, ClaEXPA-09, ClaEXPB-01, ClaEXPB-03, and ClaEXPLA-13 genes in watermelon and CmEXPA-12, CmEXPA-10, and CmEXPLA-01 genes in melon can be involved in tissue development and abiotic stress response of the plant. The current study combining bioinformatics and experimental analysis can provide a detailed characterization of the expansin superfamily which has roles in growth and reaction to the stress of the plant. The study ensures detailed data for future studies examining gene functions including the roles in plant growth and stress conditions.

Influence of dietary supplemental guanidinoacetic acid on performance, haematological parameters, carcass characteristics and enzyme activities in male broilers with cold-induced ascites

ABSTRACT This study aimed to evaluate the effects of dietary guanidinoacetic acid (GAA) on performance, enzyme activity and biochemical variables of broilers with cold-induced ascites. A total of 640 day-old male broiler chicks (Cobb 500) were randomly assigned to four treatments (with 8 replicates of 20 birds) including a control and diets supplemented with 0.6, 1.2 and 1.8 g of GAA per kg of feed. On day 14, the temperature was reduced to 17°C during the daytime and 14°C at night. Dietary inclusion of 0.6 g/kg GAA decreased BWG in compared to those fed other levels in the grower period (P < 0.05). The ratio of right to total ventricle, mortality and relative weights of liver were lower in birds fed the diet with 1.2 g/kg GAA compared to bids fed other levels (P < 0.05). The plasma activity of creatine kinase (CK) increased in the birds fed with 1.2 and 1.8 g/kg GAA compared to chicks fed the control diet. It can be concluded that dietary supplemental 1.2 g/kg GAA would be a beneficial way to decrease the mortality rate of broiler reared under cold stress conditions. In addition, the blood levels of HDL-C increased in broilers fed the 1.2 g/kg GAA diet.

Transcriptome and Metabonomics Combined Analysis Revealed the Defense Mechanism Involved in Hydrogen-Rich Water-Regulated Cold Stress Response of Tetrastigma hemsleyanum.

The poor resistance to cold stress conditions has become the bottleneck problem in Tetrastigma hemsleyanum (T. hemsleyanum) planting industry. Exogenous hydrogen (H2) plays an important role in improving stress resistance in plants. However, the key factors and regulatory network of plants in response to hydrogen-rich water (HRW) treatment under environmental stress are not clear. Here, we conducted integrative analyses of metabolome and transcriptome profiles to reveal the defense mechanism involved in the HRW-regulated cold stress response of T. hemsleyanum. The application of 75% HRW could alleviate stress damage by decreasing stomatal apparatus density and significantly increasing photosynthetic efficiency and mitigating physiological indexes of resistance, such as Pn, Cond, MDA, SOD, etc., which were changed by cold stress conditions. A total of 7,883 DEGs and 439 DEMs were identified. DEGs were the most relevant to phenylpropanoid, isoflavonoid, monoterpenoid, and flavonoid biosynthesis pathways. Using gene co-expression analysis (WGCNA), we identified one gene module that showed a strong correlation between total antioxidant capacity and transpiration rate. Trend analysis indicated that the phenylpropanoid biosynthesis pathway played a major role in the transcription and metabolism process of HRW treatment under cold stress. Based on the integrated analysis of genes and metabolites, the results showed cold stress upregulated the expression of PAL, CHS, COMT, CCR, AtBG1, etc., resulting in the accumulation of coniferyl alcohol and eriodictyol contents in T. hemsleyanum under cold stress, but the 75% HRW treatment could attenuate the enhancement. The study not only identified the main strategy of HRW protection against cold stress but also provided candidate genes for flavonoid biosynthesis, so as to better improve cold tolerance through molecular breeding techniques.

A 3-D virtual human model for simulating heat and cold stress.

In this study, we extended our previously developed anatomically detailed three-dimensional (3-D) thermoregulatory virtual human model for predicting heat stress to allow for predictions of heat and cold stress in one unified model. Starting with the modified Pennes bioheat transfer equation to estimate the spatiotemporal temperature distribution within the body as the underlying modeling structure, we developed a new formulation to characterize the spatial variation of blood temperature between body elements and within the limbs. We also implemented the means to represent heat generated from shivering and skin blood flow that apply to air exposure and water immersion. Then, we performed simulations and validated the model predictions with experimental data from nine studies, representing a wide range of heat- and cold-stress conditions in air and water and physical activities. We observed excellent agreement between model predictions and measured data, with average root mean squared errors of 0.2°C for core temperature, 0.9°C for mean skin temperature, and 27 W for heat from shivering. We found that a spatially varying blood temperature profile within the limbs was crucial to accurately predict core body temperature changes during very cold exposures. Our 3-D thermoregulatory virtual human model consistently predicted the body's thermal state accurately for each of the simulated hot and cold environmental conditions and exertional heat stress. As such, it serves as a reliable tool to assess whole body, localized tissue, and, potentially, organ-specific injury risks, helping develop injury prevention and mitigation strategies in a systematic and expeditious manner.NEW & NOTEWORTHY This work provides a new, unified modeling framework to accurately predict the human body's thermal response to both heat and cold stress caused by environmental conditions and exertional physical activity in one mathematical model. We show that this 3-D anatomically detailed model accurately predicts the spatiotemporal temperature distribution in the body under extreme conditions for exposures to air and water and could be used to help design medical interventions and countermeasures to prevent injuries.

The Performances of Some Tomato Pure Lines under Cold Stress in the Vegetative and Generative Stage

Especially, in tomatoes occur due to low temperature stress serious yield and quality decreases in greenhouse conditions. For successful tomato cultivation under the cold stress, cultivars performances are extremely important both vegetative and reproductive growth stage. In this study, 20 tomato pure lines and 3 commercial cultivars (Cigdem F1, Anit F1 and Bestona F1) and also Solanum hirsutum (LA 1777) known as tolerant genotypes were evaluated at vegetative and reproductive stage. The studies were conducted under both the cold stress in growth chamber and the optimal temperature condition (control) in the greenhouse. They were evaluated by measuring malondialdehyde (MDA), electrolyte leakage (EL) and dry matter yield (DM) at vegetative stage. The results showed that EL rate and MDA content increased while DM decreased under the cold stress when compared with leaves of plants grown at optimal temperature. In reproductive stage, pollen viability and pollen germination were evaluated under both cold stress and control conditions for all genotypes. All the sensitive genotypes exhibited low pollen viability and pollen germination. Consequently, three pure lines were identified with low-temperature tolerant in vegetative and reproductive growth stage.

The Kandelia obovata transcription factor KoWRKY40 enhances cold tolerance in transgenic Arabidopsis

BackgroundWRKY transcription factors play key roles in plant development processes and stress response. Kandelia obovata is the most cold-resistant species of mangrove plants, which are the important contributors to coastal marine environment. However, there is little known about the WRKY genes in K. obovata.ResultsIn this study, a WRKY transcription factor gene, named KoWRKY40, was identified from mangrove plant K. obovata. The full-length cDNA of KoWRKY40 gene was 1420 nucleotide bases, which encoded 318 amino acids. The KoWRKY40 protein contained a typical WRKY domain and a C2H2 zinc-finger motif, which were common signatures to group II of WRKY family. The three-dimensional (3D) model of KoWRKY40 was formed by one α-helix and five β-strands. Evolutionary analysis revealed that KoWRKY40 has the closest homology with a WRKY protein from another mangrove plant Bruguiera gymnorhiza. The KoWRKY40 protein was verified to be exclusively located in nucleus of tobacco epidermis cells. Gene expression analysis demonstrated that KoWRKY40 was induced highly in the roots and leaves, but lowly in stems in K. obovata under cold stress. Overexpression of KoWRKY40 in Arabidopsis significantly enhanced the fresh weight, root length, and lateral root number of the transgenic lines under cold stress. KoWRKY40 transgenic Arabidopsis exhibited higher proline content, SOD, POD, and CAT activities, and lower MDA content, and H2O2 content than wild-type Arabidopsis under cold stress condition. Cold stress affected the expression of genes related to proline biosynthesis, antioxidant system, and the ICE-CBF-COR signaling pathway, including AtP5CS1, AtPRODH1, AtMnSOD, AtPOD, AtCAT1, AtCBF1, AtCBF2, AtICE1, AtCOR47 in KoWRKY40 transgenic Arabidopsis plants.ConclusionThese results demonstrated that KoWRKY40 conferred cold tolerance in transgenic Arabidopsis by regulating plant growth, osmotic balance, the antioxidant system, and ICE-CBF-COR signaling pathway. The study indicates that KoWRKY40 is an important regulator involved in the cold stress response in plants.

MsSAMS, a cold stress-responsive gene, provides resistance to environmental stress in T2-generation transgenic plants

The SAMS (S-adenosylmethionine synthetase) gene is known to play an important role in the mechanism of cold resistance, as overexpression of this gene results in phenotypic changes in T1-generation transgenic plants. Accordingly, this study was conducted to test the expression of the MsSAMS gene in T2-generation transgenic plants and to investigate the resistance of these plants and the function of the transgene in response to various environmental stresses. For the morphological analysis of T2-generation transgenic plants overexpressing the MsSAMS gene, observations using scanning electron microscopy (SEM) were performed. T2-generation transgenic plants were obtained by planting a total of 5 lines, and their characteristics were tested by comparisons with those of the control. SEM revealed that the thickest leaves were produced by the T6 transgenic line-161.24 ± 8.05µm. The number of stomata ranged from 20.00 ± 2.65 to 34.00 ± 1.00 in the T2-generation transgenic plants, but the control had more stomata. Resistance to various factors, such as low temperature, drought, and oxidative stress, in the T2-generation transgenic plants was also confirmed. Under cold-stress conditions, the T6 transgenic line presented the lowest value (22.73%) of ion leakage, and under drought-stress conditions, compared with the control, the transgenic lines presented lower ion leakage after being treated with various concentrations of mannitol. Even under oxidative-stress conditions, the T2-generation transgenic plants presented ion leakage levels that were 32.91 ± 4.24 to 48.33 ± 3.54% lower than those of the control after treatment with various concentrations of methyl viologen. Regarding SAMS enzyme activity, as the duration of cold treatment increased, the activity in the transgenic plants tended to decrease and then increase. During 48h of cold treatment, the control showed a decrease in SAM content, while the T2-generation transgenic plants presented an increase in SAM content, from 13.58 ± 1.04 to 22.75 ± 1.95mg protein/g FW. The results suggest that the MsSAMS gene may be important to the mechanisms of resistance to oxidative and drought stresses in addition to its previously known association with cold resistance. Based on these results, it was suggested that the MsSAMS gene, whose expression is induced by cold stress, can serve as a marker of various responses to environmental stresses, because resistance to cold damage and various environmental stresses are stably inherited in the T2 generation.

Variations in the Adaptive Response of &lt;i&gt;Burkholderia pseudomallei&lt;/i&gt; to Cold Stress

The aim of the study was to analyze differential gene expression in Burkholderia pseudomallei strains with different survival rates under cold stress conditions.Materials and methods. Three strains of B. pseudomallei (sequence types ST 46, ST 70, and ST 85) were used as model strains. The RNA was isolated using the membrane columns method and stabilized through dehydration. The cDNA was sequenced on the Illumina MiSeq platform. Gene functions were classified using the KEGG PATHWAY database.Results and discussion. Based on the analysis of transcriptomes of B. pseudomallei strains after prolonged exposure to cold stress, the molecular mechanisms of B. pseudomallei adaptation to low temperatures have been described for the first time ever. It was shown that adaptation of B. pseudomallei to cold stress is associated with regulatory processes leading to a significant decrease in the total transcriptional activity. Two strategies of adaptation to low temperatures have been found: 1) modulation of regulatory processes leading to suppression of gene expression of the main metabolic pathways to the minimum level that ensures cell viability and activation of the minimum required set of stress response genes, and 2) less noticeable suppression of general metabolism in combination with activation of expression of an extended range of genes for cold and heat shock, general, osmotic, and universal stresses. Both mechanisms provide the causative agent of melioidosis with survival under conditions of prolonged cold stress at low positive temperatures. The first strategy showed greater efficiency at negative temperatures. The transition of B. pseudomallei to a viable but uncultivated state occurs in the long term (at least 2 years). While with the second strategy, this happens within 2 months. Assessment of the potential and molecular mechanisms of adaptation of this bacterium to cold stress is necessary to understand the degree of risk in case of a possible introduction of B. pseudomallei into regions with a temperate climate and to develop effective measures to ensure the biosafety of the environment.