Molecular Mechanisms Of Cold Tolerance Research Articles

Establishment and characterization of a gill cell line from Takifugu obscurus and transcriptome analysis of its gene expression profiles upon low temperature

Takifugu obscurus is a farmed fish of great economic importance in China. The rapid development of T. obscurus aquaculture industry has been accompanied by disease and low-temperature stress, resulting in huge economic losses. Cell lines are used extensively in teleost physiology and pathology as the most cost-effective platform for in vitro research. A novel gill cell line of T. obscurus (named TOG) was first successfully established, and passed through 52 generations. The optimal conditions for TOG growth were 20 % FBS concentration and 24 °C, TOG could be grown in both hypotonic (150 mOsmol-kg-1) and hypertonic (600 mOsmol-kg-1) environments. TOG was determined to be derived from T. obscurus by sequencing the mitochondrial COI gene. Karyotype analysis revealed that the chromosome number of TOG was 44 (2n = 44). Transfection experiment showed that TOG was able to express foreign genes. Furthermore, several immune-related genes were significantly up-regulated in TOG after LPS and poly (I:C) stimulation, including tlr3, isg15, il1β and il10. Additionally, transcriptome analysis of TOG under low-temperature stress (24 °C, 18 °C, 12 °C, 10 °C and 8 °C) found that differentially expressed genes (DEGs) were significantly clustered in several immunological and energy metabolic pathways, and cold stress could disrupt the immune barrier and reduce immunity by downregulating the immune-related pathways. Additionally, weighted gene co-expression network analysis (WGCNA) revealed that bule module and turquoise module, which were closely correlated with low temperature and the degree of fish damage, were both predominantly found in PPAR, NOD-like receptor and Toll-like receptor signaling pathway. Hub genes were identified in these two modules, including mre11, clpb, dhx15, ddx18 and utp15. TOG cell line will become an effective experimental platform for genetic and immunological research, and our results would help us gain a deeper insight into the molecular mechanism of cold tolerance in teleost.

Comparative Transcriptome Analysis Reveals Different Responses in Three Developmental Stages of Mythimna loreyi to Cold Stress.

The loreyi leafworm Mythimna loreyi (Lepidoptera: Noctuidae) is a serious pest of agriculture that causes particular damage to Gramineae crops in Asia, Europe, Australia, Africa, and the Middle East. Low temperature is one of the important environmental factors that limits the survival, distribution, colonization, and abundance of M. loreyi. However, the metabolic synthesis pathways of cold-tolerant substances in M. loreyi and the key genes involved in the regulation under cold stress remain largely unknown. In this study, we sequenced the transcriptomes of three developmental stages (larvae, pupae, and adults) of M. loreyi to discover the molecular mechanisms of their responses to cold stress. In total, sequencing generated 120.64 GB of clean data from 18 samples, of which 19,459 genes and 1740 differentially expressed genes (DEGs) were identified. The enrichment analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that many DEGs were mainly enriched in pathways associated with energy metabolism and hormone metabolism. Among these, genes encoding multiple metabolic enzymes, cuticle proteins (CPs), and heat shock proteins (HSPs) were differentially expressed. These results indicate that there are significant differences among the three developmental stages of M. loreyi exposed to cold stress and provide a basis for further studying the molecular mechanisms of cold tolerance in insects.

The Molecular Mechanism of Cold-Stress Tolerance: Cold Responsive Genes and Their Mechanisms in Rice (Oryza sativa L.).

Rice (Oryza sativa L.) production is highly susceptible to temperature fluctuations, which can significantly reduce plant growth and development at different developmental stages, resulting in a dramatic loss of grain yield. Over the past century, substantial efforts have been undertaken to investigate the physiological, biochemical, and molecular mechanisms of cold stress tolerance in rice. This review aims to provide a comprehensive overview of the recent developments and trends in this field. We summarized the previous advancements and methodologies used for identifying cold-responsive genes and the molecular mechanisms of cold tolerance in rice. Integration of new technologies has significantly improved studies in this era, facilitating the identification of essential genes, QTLs, and molecular modules in rice. These findings have accelerated the molecular breeding of cold-resistant rice varieties. In addition, functional genomics, including the investigation of natural variations in alleles and artificially developed mutants, is emerging as an exciting new approach to investigating cold tolerance. Looking ahead, it is imperative for scientists to evaluate the collective impacts of these novel genes to develop rice cultivars resilient to global climate change.

TaMAPK3 phosphorylates TaCBF and TaICE and plays a negative role in wheat freezing tolerance

Freezing temperature during overwintering often kills plants; plants have thus, developed a defense mechanism called ‘cold acclimation’, in which a number of genes are involved in increasing cell protection and gene expression. Mitogen-activated protein kinase (MAPK) controls proteins' activities by phosphorylation and is involved in numerous metabolic pathways. In this study, we identified the protein interaction between TaMAPK3 and the proteins in the cold response pathway, ICE41, ICE87, and CBFIVd-D9. The subcellular localization and bimolecular fluorescence complement (BiFC) assays revealed that these proteins interact in the nucleus or in the plasma membrane. Furthermore, MAPK3-mediated phosphorylation of ICE41, ICE87, and CBFIVd-D9 was verified using an in vitro phosphorylation assay. TaMAPK3-overexpressing transgenic Brachypodium showed a lower survival rate upon freezing stress and lower proline content during cold acclimation, compared to wild-type plants. Furthermore, cold response gene expression analysis revealed that the expression of these genes was suppressed in the transgenic lines under cold treatment. It was further elucidated that MAPK3 mediates the degradation of ICE and CBF proteins, which implies the negative impact of MAPK3 on the freezing tolerance of plants. This study will help to elucidate the molecular mechanisms of cold tolerance and the activity of MAPK3 in wheat.

Full-length transcriptome sequencing provides insights into alternative splicing under cold stress in peanut.

Peanut (Arachis hypogaea L.), also called groundnut is an important oil and cash crop grown widely in the world. The annual global production of groundnuts has increased to approximately 50 million tons, which provides a rich source of vegetable oils and proteins for humans. Low temperature (non-freezing) is one of the major factors restricting peanut growth, yield, and geographic distribution. Since the complexity of cold-resistance trait, the molecular mechanism of cold tolerance and related gene networks were largely unknown in peanut. In this study, comparative transcriptomic analysis of two peanut cultivars (SLH vs. ZH12) with differential cold tolerance under low temperature (10°C) was performed using Oxford Nanopore Technology (ONT) platform. As a result, we identified 8,949 novel gene loci and 95,291 new/novel isoforms compared with the reference database. More differentially expressed genes (DEGs) were discovered in cold-sensitive cultivar (ZH12) than cold-tolerant cultivar (SLH), while more alternative splicing events were found in SLH compared to ZH12. Gene Ontology (GO) analyses of the common DEGs showed that the "response to stress", "chloroplast part", and "transcription factor activity" were the most enriched GO terms, indicating that photosynthesis process and transcription factors play crucial roles in cold stress response in peanut. We also detected a total of 708 differential alternative splicing genes (DASGs) under cold stress compared to normal condition. Intron retention (IR) and exon skipping (ES) were the most prevalent alternative splicing (AS) events. In total, 4,993 transcription factors and 292 splicing factors were detected, many of them had differential expression levels and/or underwent AS events in response to cold stress. Overexpression of two candidate genes (encoding trehalose-6-phosphatephosphatases, AhTPPs) in yeast improves cold tolerance. This study not only provides valuable resources for the study of cold resistance in peanut but also lay a foundation for genetic modification of cold regulators to enhance stress tolerance in crops.

Integration of transcriptome and metabolome reveals molecular mechanisms responsive to cold stress in gynogenetic mrigal carp (Cirrhinus mrigala)

Cold stress is a major environmental stimulus affecting the physiological and metabolic activities of farmed fish. In our previous study, we obtained gynogenetic mrigal carp (Cirrhinus mrigala, GMC) with significantly improved cold tolerance compared with the wild individuals. To gain a comprehensive and unbiased understanding of the molecular mechanism of cold tolerance in GMC, transcriptomes and metabolomics of liver tissues were comparatively analysed. After cold treatment at 14 °C for 48 h, histology analysis revealed that the hepatocyte nucleus was shifted, blurred and disintegrated; the terminus of the gill filament was enlarged; and the number of mucus cells and their secretions were increased. A total of 4844 differentially expressed genes (DEGs), including 2420 upregulated and 2424 downregulated genes, were identified. The upregulated KEGG pathways were mainly involved in oxidative phosphorylation, the proteasome and the citrate cycle, while the downregulated pathways were involved in staphylococcus aureus infection, systemic lupus erythematosus and leishmaniasis. A total of 160 differential metabolites (DMs) were identified, including 39 increased and 121 reduced metabolites. The KEGG pathway analysis revealed significant enrichment in retinol metabolism and folate biosynthesis. KEGG function analysis showed that the DEGs and DMs were both highly enriched in purine metabolism, neuroactive ligand–receptor interaction and galactose metabolism. Correlation analysis between DEGs and DMs revealed that multiple pathways were active and strongly related to immunity and disease, metabolism and growth under cold stress. This study can facilitate our understanding of the molecular mechanisms of fish response to cold stress and provide insight into the application of gynogenesis in farmed fish.

Differential Regulations of Antioxidant Metabolism and Cold-Responsive Genes in Three Bermudagrass Genotypes under Chilling and Freezing Stress.

As a typical warm-season grass, bermudagrass growth and turf quality begin to decrease when the environmental temperature drops below 20 °C. The current study investigated the differential responses of three bermudagrass genotypes to chilling stress (8/4 °C) for 15 days and then freezing stress (2/-2 °C) for 2 days. The three genotypes exhibited significant variation in chilling and freezing tolerance, and Chuannong-3, common bermudagrass 001, and Tifdwarf were ranked as cold-tolerant, -intermediate, and -sensitive genotypes based on evaluations of chlorophyll content, the photochemical efficiency of photosystem II, oxidative damage, and cell membrane stability, respectively. Chuannong-3 achieved better tolerance through enhancing the antioxidant defense system to stabilize cell membrane and reactive oxygen species homeostasis after being subjected to chilling and freezing stresses. Chuannong-3 also downregulated the ethylene signaling pathway by improving CdCTR1 expression and suppressing the transcript levels of CdEIN3-1 and CdEIN3-2; however, it upregulated the hydrogen sulfide signaling pathway via an increase in CdISCS expression under cold stress. In addition, the molecular basis of cold tolerance could be associated with the mediation of key genes in the heat shock pathway (CdHSFA-2b, CdHSBP-1, CdHSP22, and CdHSP40) and the CdOSMOTIN in Chuannong-3 because the accumulation of stress-defensive proteins, including heat shock proteins and osmotin, plays a positive role in osmoprotection, osmotic adjustment, or the repair of denatured proteins as molecular chaperones under cold stress. The current findings give an insight into the physiological and molecular mechanisms of cold tolerance in the new cultivar Chuannong-3, which provides valuable information for turfgrass breeders and practitioners.

The genome-wide characterization and associated cold-tolerance function of the superoxide dismutase in the cold response of the tick Haemaphysalis longicornis

Accumulating evidence suggests that superoxide dismutase (SOD) is the first line of antioxidant defense in organisms and plays an important role in scavenging reactive oxygen species produced during environmental stress. However, limited information is available regarding the response of SOD genes to cold stress in ticks. Therefore, in the present study, SOD genes were cloned and identified from the genome of Haemaphysalis longicornis, and the function of SOD during the cold response was further explored. Seven SOD genes were characterized: HlCCS1, HlCCS2, HlMSD, HlCSD1, HlCSD2, HlCSD3, and HlCSD4. Bioinformatics analysis showed that HlCCS1 and HlCCS2 are copper chaperones of SODs. HlCSD1–HlCSD4 belong to the Cu/Zn SOD, whereas HlMSD belongs to the Mn SOD gene family. Fluorescence quantitative PCR showed that the expression of HlCCS2, HlMSD, and HlCSD1–3 was upregulated, whereas HlCCS1 and HlCSD4 were downregulated during the cold response of H. longicornis. Western blotting confirmed changes in the relative expression of HlCSD3 and HlMSD in H. longicornis after cold treatment. Mortality of H. longicornis increased significantly after dsRNA injection of HlCCS2, HlMSD, HlCSD1, and HlCSD3. The above results show that SODs have different regulatory functions during the cold response in H. longicornis, and there might be an interaction between treatment temperature and duration. Furthermore, the results lay a foundation for subsequent research on the molecular mechanism of cold tolerance in H. longicornis and shed light on the population distribution and diffusion limit of ticks.

Genome-wide analysis of the WRKY genes and their important roles during cold stress in white clover.

White clover (Trifolium repens L) is a high-quality forage grass with a high protein content, but it is vulnerable to cold stress, which can negatively affect its growth and development. WRKY transcription factor is a family of plant transcription factors found mainly in higher plants and plays an important role in plant growth, development, and stress response. Although WRKY transcription factors have been studied extensively in other plants, it has been less studied in white clover. In the present research, we have performed a genome-wide analysis of the WRKY gene family of white clover, in total, there were 145 members of WRKY transcription factors identified in white clover. The characterization of the TrWRKY genes was detailed, including conserved motif analysis, phylogenetic analysis, and gene duplication analysis, which have provided a better understanding of the structure and evolution of the TrWRKY genes in white clover. Meanwhile, the genetic regulation network (GRN) containing TrWRKY genes was reconstructed, and Gene Ontology (GO) annotation analysis of these function genes showed they contributed to regulation of transcription process, response to wounding, and phosphorylay signal transduction system, all of which were important processes in response to abiotic stress. To determine the TrWRKY genes function under cold stress, the RNA-seq dataset was analyzed; most of TrWRKY genes were highly upregulated in response to cold stress, particularly in the early stages of cold stress. These results were validated by qRT-PCR experiment, implying they are involved in various gene regulation pathways in response to cold stress. The results of this study provide insights that will be useful for further functional analyses of TrWRKY genes in response to biotic or abiotic stresses in white clover. These findings are likely to be useful for further research on the functions of TrWRKY genes and their role in response to cold stress, which is important to understand the molecular mechanism of cold tolerance in white clover and improve its cold tolerance.

Genome-wide analysis of the CDPK gene family and their important roles response to cold stress in white clover

ABSTRACT Calcium-dependent protein kinases (CDPKs) are an important class of calcium-sensitive response proteins that play an important regulatory role in response to abiotic stresses. To date, little is known about the CDPK genes in white clover. White clover is a high-quality forage grass with high protein content, but it is susceptible to cold stress. Therefore, we performed a genome-wide analysis of the CDPK gene family in white clover and identified 50 members of the CDPK genes. Phylogenetic analysis using CDPKs from the model plant Arabidopsis divided the TrCDPK genes into four groups based on their sequence similarities. Motif analysis showed that TrCDPKs within the same group had similar motif compositions. Gene duplication analysis revealed the evolution and expansion of TrCDPK genes in white clover. Meanwhile, a genetic regulatory network (GRN) containing TrCDPK genes was reconstructed, and gene ontology (GO) annotation analysis of these functional genes showed that they contribute to signal transduction, cellular response to stimuli, and biological regulation, all of which are important processes in response to abiotic stresses. To determine the function of TrCDPK genes, we analyzed the RNA-seq dataset and found that most TrCDPK genes were highly up-regulated under cold stress, particularly in the early stages of cold stress. These results were validated by qRT-PCR experiments, implying that TrCDPK genes are involved in various gene regulatory pathways in response to cold stress. Our study may help to further investigate the function of TrCDPK genes and their role in response to cold stress, which is important for understanding the molecular mechanisms of cold tolerance in white clover and improving its cold tolerance.

Transcriptome and proteome depth analysis indicate ABA, MAPK cascade and Ca2+ signaling co-regulate cold tolerance in Rhododendron chrysanthum Pall.

Cold stress is a global common problem that significantly limits plant development and geographical distribution. Plants respond to low temperature stress by evolving interrelated regulatory pathways to respond and adapt to their environment in a timely manner. Rhodoendron chrysanthum Pall. (R. chrysanthum) is a perennial evergreen dwarf shrub used for adornment and medicine that thrives in the Changbai Mountains at high elevations and subfreezing conditions. In this study, a comprehensive investigation of cold tolerance (4°C, 12h) in R. chrysanthum leaves under cold using physiological combined with transcriptomic and proteomic approaches. There were 12,261 differentially expressed genes (DEGs) and 360 differentially expressed proteins (DEPs) in the low temperature (LT) and normal treatment (Control). Integrated transcriptomic and proteomic analyses showed that MAPK cascade, ABA biosynthesis and signaling, plant-pathogen interaction, linoleic acid metabolism and glycerophospholipid metabolism were significantly enriched in response to cold stress of R. chrysanthum leaves. We analyzed the involvement of ABA biosynthesis and signaling, MAPK cascade, and Ca2+ signaling, that may jointly respond to stomatal closure, chlorophyll degradation, and ROS homeostasis under low temperature stress. These results propose an integrated regulatory network of ABA, MAPK cascade and Ca2+ signaling comodulating the cold stress in R. chrysanthum, which will provide some insights to elucidate the molecular mechanisms of cold tolerance in plants.

Tetratricopeptide repeat protein SlREC2 positively regulates cold tolerance in tomato.

Cold stress is a key environmental constraint that dramatically affects the growth, productivity, and quality of tomato (Solanum lycopersicum); however, the underlying molecular mechanisms of cold tolerance remain poorly understood. In this study, we identified REDUCED CHLOROPLAST COVERAGE 2 (SlREC2) encoding a tetratricopeptide repeat protein that positively regulates tomato cold tolerance. Disruption of SlREC2 largely reduced abscisic acid (ABA) levels, photoprotection, and the expression of C-REPEAT BINDING FACTOR (CBF)-pathway genes in tomato plants under cold stress. ABA deficiency in the notabilis (not) mutant, which carries a mutation in 9-CIS-EPOXYCAROTENOID DIOXYGENASE 1 (SlNCED1), strongly inhibited the cold tolerance of SlREC2-silenced plants and empty vector control plants and resulted in a similar phenotype. In addition, foliar application of ABA rescued the cold tolerance of SlREC2-silenced plants, which confirms that SlNCED1-mediated ABA accumulation is required for SlREC2-regulated cold tolerance. Strikingly, SlREC2 physically interacted with β-RING CAROTENE HYDROXYLASE 1b (SlBCH1b), a key regulatory enzyme in the xanthophyll cycle. Disruption of SlBCH1b severely impaired photoprotection, ABA accumulation, and CBF-pathway gene expression in tomato plants under cold stress. Taken together, this study reveals that SlREC2 interacts with SlBCH1b to enhance cold tolerance in tomato via integration of SlNCED1-mediated ABA accumulation, photoprotection, and the CBF-pathway, thus providing further genetic knowledge for breeding cold-resistant tomato varieties.

Transcriptomics Profiling of Acer pseudosieboldianum Molecular Mechanism against Freezing Stress

Low temperature is an important environmental factor that affects the growth and development of trees and leads to the introduction of failure in the genetic improvement of trees. Acer pseudosieboldianum is a tree species that is well-known for its bright red autumn leaf color. These trees are widely used in landscaping in northeast China. However, due to their poor cold resistance, introduced A. pseudosieboldianum trees suffer severe freezing injury in many introduced environments. To elucidate the physiological indicators and molecular mechanisms associated with freezing damage, we analyzed the physiological indicators and transcriptome of A. pseudosieboldianum, using kits and RNA-Seq technology. The mechanism of A. pseudosieboldianum in response to freezing stress is an important scientific question. In this study, we used the shoots of four-year-old A. pseudosieboldianum twig seedlings, and the physiological index and the transcriptome of A. pseudosieboldianum under low temperature stress were investigated. The results showed that more than 20,000 genes were detected in A. pseudosieboldianum under low temperature (4 °C) and freezing temperatures (-10 °C, -20 °C, -30 °C, and -40 °C). There were 2505, 6021, 5125, and 3191 differential genes (DEGs) between -10 °C, -20°C, -30°C, -40 °C, and CK (4 °C), respectively. Among these differential genes, 48 genes are involved in the MAPK pathway and 533 genes are involved in the glucose metabolism pathway. In addition, the important transcription factors (MYB, AP2/ERF, and WRKY) involved in freezing stress were activated under different degrees of freezing stress. A total of 10 sets of physiological indicators of A. pseudosieboldianum were examined, including the activities of five enzymes and the accumulation of five hormones. All of the physiological indicators except SOD and GSH-Px reached their maximum values at -30 °C. The enzyme activity of SOD was highest at -10 °C, and that of GSH-Px was highest at -20 °C. Our study is the first to provide a more comprehensive understanding of the differential genes (DEGs) involved in A. pseudosieboldianum under freezing stress at different temperatures at the transcriptome level. These results may help to clarify the molecular mechanism of cold tolerance of A. pseudosieboldianum and provide new insights and candidate genes for the genetic improvement of the freezing tolerance of A. pseudosieboldianum.

Metabolic profiling and gene expression analyses provide insights into cold adaptation of an Antarctic moss Pohlia nutans.

Antarctica is the coldest, driest, and most windy continent on earth. The major terrestrial vegetation consists of cryptogams (mosses and lichens) and two vascular plant species. However, the molecular mechanism of cold tolerance and relevant regulatory networks were largely unknown in these Antarctic plants. Here, we investigated the global alterations in metabolites and regulatory pathways of an Antarctic moss (Pohlia nutans) under cold stress using an integrated multi-omics approach. We found that proline content and several antioxidant enzyme activities were significantly increased in P. nutans under cold stress, but the contents of chlorophyll and total flavonoids were markedly decreased. A total of 559 metabolites were detected using ultra high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). We observed 39 and 71 differentially changed metabolites (DCMs) after 24 h and 60 h cold stress, indicating that several major pathways were differentially activated for producing fatty acids, alkaloids, flavonoids, terpenoids, and phenolic acids. In addition, the quantitative transcriptome sequencing was conducted to uncover the global transcriptional profiles of P. nutans under cold stress. The representative differentially expressed genes (DEGs) were identified and summarized to the function including Ca2+ signaling, ABA signaling, jasmonate signaling, fatty acids biosynthesis, flavonoid biosynthesis, and other biological processes. The integrated dataset analyses of metabolome and transcriptome revealed that jasmonate signaling, auxin signaling, very-long-chain fatty acids and flavonoid biosynthesis pathways might contribute to P. nutans acclimating to cold stress. Overall, these observations provide insight into Antarctic moss adaptations to polar habitats and the impact of global climate change on Antarctic plants.

Quantitative Trait Loci Mapping Analysis for Cold Tolerance Under Cold Stress and Brassinosteroid-Combined Cold Treatment at Germination and Bud Burst Stages in Rice.

Low temperature is one of the major abiotic stresses limiting seed germination and early seedling growth in rice. Brassinosteroid (BR) application can improve cold tolerance in rice. However, the regulatory relationship between cold tolerance and BR in rice remains undefined. Here, we constructed a population of 140 backcross recombinant inbred lines (BRILs) derived from a cross between a wild rice (Dongxiang wild rice, DXWR) and a super rice (SN265). The low-temperature germination rate (LTG), survival rate (SR), plant height (PH), and first leaf length (FLL) were used as indices for assessing cold tolerance under cold stress and BR-combined cold treatment at seed germination and bud burst stages. A high-resolution SNP genetic map, covering 1,145 bin markers with a distance of 3188.33 cM onto 12 chromosomes, was constructed using the GBS technique. A total of 73 QTLs were detected, of which 49 QTLs were identified under cold stress and 24 QTLs under BR-combined cold treatment. Among these, intervals of 30 QTLs were pairwise coincident under cold stress and BR-combined cold treatment, as well as different traits including SR and FLL, and PH and FLL, respectively. A total of 14 candidate genes related to cold tolerance or the BR signaling pathway, such as CBF/DREB (LOC_Os08g43200), bHLH (LOC_Os07g08440 and LOC_Os07g08440), WRKY (LOC_Os06g30860), MYB (LOC_Os01g62410 and LOC_Os05g51160), and BRI1-associated receptor kinase 1 precursor (LOC_Os06g16300), were located. Among these, the transcript levels of 10 candidate genes were identified under cold stress and BR-combined cold treatment by qRT-PCR. These findings provided an important basis for further mining the genes related to cold tolerance or the BR signaling pathway and understanding the molecular mechanisms of cold tolerance in rice.

The genetic basis of cold tolerance in cucumber (Cucumis sativus L.)-the latest developments and perspectives.

Cold stress is one of the main causes of yield losses in plant production in temperate climate areas. Cold stress slows down and even stops plant growth and development and causes injuries that may result in the plant's death. Cucumber (Cucumis sativus L.), an economically important vegetable, is sensitive to low temperatures, thus improving cold tolerance in cucumber would benefit cucumber producers, particularly those farming in temperate climates and higher altitude areas. So far, single cucumber accessions showing different degrees of cold tolerance have been identified, and genetic studies have revealed biparentally and maternally inherited genetic factors responsible for chilling tolerance. Paternally transmitted chilling tolerance has also been suggested. Quantitative trait loci (QTL) associated with seed germination ability at low temperature and seedling recovery from chilling have been described. Several transgenic attempts have been made to improve cold tolerance in cucumber. Despite numerous studies, the molecular mechanisms of cold tolerance in cucumber have still not been sufficiently elucidated. In this review, we summarise the results of research focused on understanding the genetic basis of cold tolerance in cucumber and their implications for cucumber breeding.

Evaluation of cold tolerance and gene expression patterns associated with low-temperature stress in giant freshwater prawn Macrobrachium rosenbergii

The giant freshwater prawn (GFP) Macrobrachium rosenbergii is a commercially important cultured species that originates from tropical and subtropical regions and has poor cold tolerance. This study investigated the species’ cold tolerance and regulation patterns of the related response genes, using GFP juveniles subjected to extreme low-temperature treatment. The water temperature was decreased from 26 °C at a rate of approximately 1 °C every 4 h until all prawns died. The temperatures resulting in lowest LT50 (lethal temperature at which 50% prawn die) and lowest lethal temperature were approximately 12.5 °C and 11.5 °C, respectively. Phenotypic traits and hardness of the exoskeleton generally had little effect on the cold tolerance of the prawns; however, two body-weight groups showed a highly significant difference (P < 0.01) in cold-tolerance ability, with smaller prawns (≤2 g) more susceptible to low temperatures than larger ones. Furthermore, the cold-tolerant group (last 10% of mortalities) had better cold tolerance than the cold-sensitive group (first 10% of mortalities). Additionally, comparative transcriptomic analysis was performed for the cold-tolerant and cold-sensitive groups under low-temperature stress, and for the control group maintained at ~26 °C. Through the identification of differentially expressed genes (DEGs) and RT-qPCR validation experiments, several key candidate genes were screened out, including acetyl-CoA carboxylase and laccase1-like protein, which may exert considerable influence on the response of GFP to low-temperature stress. More DEGs were found in hepatopancreas than in muscle, and several key pathways were significantly enriched in hepatopancreas, including folate biosynthesis, starch and sucrose metabolism, and pancreatic secretion, indicating the hepatopancreas plays a critical role in cold tolerance. Within the pathway of folate biosynthesis, the key upregulated gene is Alkaline phosphatase, suggesting that this gene is critical in response to low temperatures. The present results contribute to understanding the molecular mechanism of cold tolerance in this species.

Gene-coexpression network analysis identifies specific modules and hub genes related to cold stress in rice

BackgroundWhen plants are subjected to cold stress, they undergo a series of molecular and physiological changes to protect themselves from injury. Indica cultivars can usually withstand only mild cold stress in a relatively short period. Hormone-mediated defence response plays an important role in cold stress. Weighted gene co-expression network analysis (WGCNA) is a very useful tool for studying the correlation between genes, identifying modules with high phenotype correlation, and identifying Hub genes in different modules. Many studies have elucidated the molecular mechanisms of cold tolerance in different plants, but little information about the recovery process after cold stress is available.ResultsTo understand the molecular mechanism of cold tolerance in rice, we performed comprehensive transcriptome analyses during cold treatment and recovery stage in two cultivars of near-isogenic lines (9311 and DC907). Twelve transcriptomes in two rice cultivars were determined. A total of 2509 new genes were predicted by fragment splicing and assembly, and 7506 differentially expressed genes were identified by pairwise comparison. A total of 26 modules were obtained by expression-network analysis, 12 of which were highly correlated with cold stress or recovery treatment. We further identified candidate Hub genes associated with specific modules and analysed their regulatory relationships based on coexpression data. Results showed that various plant-hormone regulatory genes acted together to protect plants from physiological damage under short-term low-temperature stress. We speculated that this may be common in rice. Under long-term cold stress, rice improved the tolerance to low-temperature stress by promoting autophagy, sugar synthesis, and metabolism.ConclusionThrough WGCNA analysis at the transcriptome level, we provided a potential regulatory mechanism for the cold stress and recovery of rice cultivars and identified candidate central genes. Our findings provided an important reference for the future cultivation of rice strains with good tolerance.

Transcriptome analysis and differential gene expression profiling of wucai (Brassica campestris L.) in response to cold stress

BackgroundWucai suffers from low temperature during the growth period, resulting in a decline in yield and poor quality. But the molecular mechanisms of cold tolerance in wucai are still unclear.ResultsAccording to the phenotypes and physiological indexes, we screened out the cold-tolerant genotype “W18” (named CT) and cold-sensitive genotype “Sw-1” (named CS) in six wucai genotypes. We performed transcriptomic analysis using seedling leaves after 24 h of cold treatment. A total of 3536 and 3887 differentially expressed genes (DEGs) were identified between the low temperature (LT) and control (NT) comparative transcriptome in CT and CS, respectively, with 1690 DEGs specific to CT. The gene ontology (GO) analysis showed that the response to cadmium ion (GO:0,046,686), response to jasmonic acid (GO:0,009,753), and response to wounding (GO:0,009,611) were enriched in CT (LT vs NT). The DEGs were enriched in starch and sucrose metabolism and glutathione metabolism in both groups, and α-linolenic acid metabolism was enriched only in CT (LT vs NT). DEGs in these processes, including glutathione S-transferases (GSTs), 13S lipoxygenase (LOX), and jasmonate ZIM-domain (JAZ), as well as transcription factors (TFs), such as the ethylene-responsive transcription factor 53 (ERF53), basic helix-loop-helix 92 (bHLH92), WRKY53, and WRKY54.We hypothesize that these genes play important roles in the response to cold stress in this species.ConclusionsOur data for wucai is consistent with previous studies that suggest starch and sucrose metabolism increased the content of osmotic substances, and the glutathione metabolism pathway enhance the active oxygen scavenging. These two pathways may participated in response to cold stress. In addition, the activation of α-linolenic acid metabolism may promote the synthesis of methyl jasmonate (MeJA), which might also play a role in the cold tolerance of wucai.

Integrated Transcriptomics and Metabolomics Analysis Reveal Key Metabolism Pathways Contributing to Cold Tolerance in Peanut.

Low temperature (non-freezing) is one of the major limiting factors in peanut (Arachis hypogaea L.) growth, yield, and geographic distribution. Due to the complexity of cold-resistance trait in peanut, the molecular mechanism of cold tolerance and related gene networks were largely unknown. In this study, metabolomic analysis of two peanut cultivars subjected to chilling stress obtained a set of cold-responsive metabolites, including several carbohydrates and polyamines. These substances showed a higher accumulation pattern in cold-tolerant variety SLH than cold-susceptible variety ZH12 under cold stress, indicating their importance in protecting peanut from chilling injuries. In addition, 3,620 cold tolerance genes (CTGs) were identified by transcriptome sequencing, and the CTGs were most significantly enriched in the “phenylpropanoid biosynthesis” pathway. Two vital modules and several novel hub genes were obtained by weighted gene co-expression network analysis (WGCNA). Several key genes involved in soluble sugar, polyamine, and G-lignin biosynthetic pathways were substantially higher and/or responded more quickly in SLH (cold tolerant) than ZH12 (cold susceptible) under low temperature, suggesting they might be crucial contributors during the adaptation of peanut to low temperature. These findings will not only provide valuable resources for study of cold resistance in peanut but also lay a foundation for genetic modification of cold regulators to enhance stress tolerance in crops.