Cold-regulated Genes Research Articles

Cycling DOF factor mediated seasonal regulation of sexual reproduction and cold response is not conserved in Physcomitrium patens

Cycling DOF factor mediated seasonal regulation of sexual reproduction and cold response is not conserved in <i>Physcomitrium patens</i>

EjCBF3 conferred cold-resistance through the enhancement of antioxidase activity in loquat (Eriobotrya japonica Lindl.)

Low-temperature is a serious threat to production of plants, and cold damage of loquat (Eriobotrya japonica Lindl.) reduces fruit yield tremendously. C-Repeat Binding Factors (CBFs) are known to work as core regulators for cold adaption in plants. However, loquat EjCBFs and their mechanism for cold tolerance have not been identified. In this study, 3 EjCBFs were discovered from loquat using genome-wide searching strategy, which were candidates responding to low-temperature stress supported by analysis of conserved domains and cis-elements in promoters. The greatest up-regulation was shown on EjCBF3 both in loquat leaves and young fruits subjected to low-temperature treatment. EjCBF3 was located in the cell nucleus of Nicotiana benthamiana (N. benthamiana), and its transcriptional activation activity was verified in yeast. Transient over-expression of EjCBF3 alleviated chilling injury through the increase of antioxidase peroxidase (POD) and superoxide dismutase (SOD) activity in N. benthamiana, and the removal of H2O2 with the application of scavenger attenuated cold damage in loquat leaves. Higher survival rate and lower electrolyte leakage were found in Arabidopsis thaliana (A. thaliana) treated with low-temperature stress with the over-expression (OE) of EjCBF3. Furthermore, the induction of cold-regulated genes (CORs) including COR15A, COR47, COR78, and KIN1 were greater in OE plants than that of wild type (WT) suffering cold stress. Results provide new insights into the cold tolerance of loquat and a novel cold-resistant gene, EjCBF3, for loquat breeding.

You can have your cake and eat it too: Ectopic expression of COLD-REGULATED genes reshapes the salicylic acid-mediated growth-defense tradeoff.

You can have your cake and eat it too: Ectopic expression of COLD-REGULATED genes reshapes the salicylic acid-mediated growth-defense tradeoff.

Altering cold-regulated gene expression decouples the salicylic acid-growth trade-off in Arabidopsis.

In Arabidopsis (Arabidopsis thaliana), overproduction of salicylic acid (SA) increases disease resistance and abiotic stress tolerance but penalizes growth. This growth-defense trade-off has hindered the adoption of SA-based disease management strategies in agriculture. However, investigation of how SA inhibits plant growth has been challenging because many SA-hyperaccumulating Arabidopsis mutants have developmental defects due to the pleiotropic effects of the underlying genes. Here, we heterologously expressed a bacterial SA synthase gene in Arabidopsis and observed that elevated SA levels decreased plant growth and reduced the expression of cold-regulated (COR) genes in a dose-dependent manner. Growth suppression was exacerbated at below-ambient temperatures. Severing the SA-responsiveness of individual COR genes was sufficient to overcome the growth inhibition caused by elevated SA at ambient and below-ambient temperatures while preserving disease- and abiotic-stress-related benefits. Our results show the potential of decoupling SA-mediated growth and defense trade-offs for improving crop productivity.

Autophagy is suppressed by low temperatures and is dispensable for cold acclimation in Arabidopsis.

Plants have evolved various mechanisms to adapt to the ever-changing external environment. Autophagy is one such mechanism and has been suggested to play a key role in responding to and adapting to abiotic stresses in plants. However, the role of autophagy in adaptation to cold and freezing stresses remains to be characterized in detail. Here, we investigated the role of autophagy in the low-temperature response of Arabidopsis using atg mutants. Both the atg5-1 and atg10-1 mutants exhibited normal freezing tolerance, regardless of cold acclimation. A comparison of fresh weights indicated that the difference in growth between the wild-type and atg plants under cold conditions was rather small compared with that under normal conditions. Analysis of COLD-REGULATED gene expression showed no significant differences between the atg mutants and wild type. Treatment with 3-methyladenine, an inhibitor of autophagy, did not impair the induction of COR15Apro::LUC expression upon exposure to low temperature. Evaluation of autophagic activity using transgenic plants expressing RBCS-mRFP demonstrated that autophagy was rarely induced by cold exposure, even in the dark. Taken together, these data suggest that autophagy is suppressed by low temperatures and is dispensable for cold acclimation and freezing tolerance in Arabidopsis.

Cold stress induces rapid gene-specific changes in the levels of H3K4me3 and H3K27me3 in Arabidopsis thaliana.

When exposed to low temperatures, plants undergo a drastic reprogramming of their transcriptome in order to adapt to their new environmental conditions, which primes them for potential freezing temperatures. While the involvement of transcription factors in this process, termed cold acclimation, has been deeply investigated, the potential contribution of chromatin regulation remains largely unclear. A large proportion of cold-inducible genes carries the repressive mark histone 3 lysine 27 trimethylation (H3K27me3), which has been hypothesized as maintaining them in a silenced state in the absence of stress, but which would need to be removed or counteracted upon stress perception. However, the fate of H3K27me3 during cold exposure has not been studied genome-wide. In this study, we offer an epigenome profiling of H3K27me3 and its antagonistic active mark H3K4me3 during short-term cold exposure. Both chromatin marks undergo rapid redistribution upon cold exposure, however, the gene sets undergoing H3K4me3 or H3K27me3 differential methylation are distinct, refuting the simplistic idea that gene activation relies on a switch from an H3K27me3 repressed chromatin to an active form enriched in H3K4me3. Coupling the ChIP-seq experiments with transcriptome profiling reveals that differential histone methylation only weakly correlates with changes in expression. Interestingly, only a subset of cold-regulated genes lose H3K27me3 during their induction, indicating that H3K27me3 is not an obstacle to transcriptional activation. In the H3K27me3 methyltransferase curly leaf (clf) mutant, many cold regulated genes display reduced H3K27me3 levels but their transcriptional activity is not altered prior or during a cold exposure, suggesting that H3K27me3 may serve a more intricate role in the cold response than simply repressing the cold-inducible genes in naïve conditions.

HbBIN2 Functions in Plant Cold Stress Resistance through Modulation of HbICE1 Transcriptional Activity and ROS Homeostasis in Hevea brasiliensis.

Cold stress poses significant limitations on the growth, latex yield, and ecological distribution of rubber trees (Hevea brasiliensis). The GSK3-like kinase plays a significant role in helping plants adapt to different biotic and abiotic stresses. However, the functions of GSK3-like kinase BR-INSENSITIVE 2 (BIN2) in Hevea brasiliensis remain elusive. Here, we identified HbBIN2s of Hevea brasiliensis and deciphered their roles in cold stress resistance. The transcript levels of HbBIN2s are upregulated by cold stress. In addition, HbBIN2s are present in both the nucleus and cytoplasm and have the ability to interact with the INDUCER OF CBF EXPRESSION1(HbICE1) transcription factor, a central component in cold signaling. HbBIN2 overexpression in Arabidopsis displays decreased tolerance to chilling stress with a lower survival rate and proline content but a higher level of electrolyte leakage (EL) and malondialdehyde (MDA) than wild type under cold stress. Meanwhile, HbBIN2 transgenic Arabidopsis treated with cold stress exhibits a significant increase in the accumulation of reactive oxygen species (ROS) and a decrease in the activity of antioxidant enzymes. Further investigation reveals that HbBIN2 inhibits the transcriptional activity of HbICE1, thereby attenuating the expression of C-REPEAT BINDING FACTOR (HbCBF1). Consistent with this, overexpression of HbBIN2 represses the expression of CBF pathway cold-regulated genes under cold stress. In conclusion, our findings indicate that HbBIN2 functions as a suppressor of cold stress resistance by modulating HbICE1 transcriptional activity and ROS homeostasis.

Genome-wide characterization of tobacco B-BOX gene family identified two close members play contrast roles under cold stress

Cold stress influences the growth and geographical distribution of plants. Tobacco (Nicotiana tabacum L.), a model in plant biology studies and a world-widely cultivated economic plant from tropical America, exhibits extreme sensitivity to low temperatures. The conserved B-BOX (BBX) transcription factors play vital roles in plant photomorphogenesis and flowering regulation, and their functions in abiotic stresses, including cold stress, have been uncovered recently. However, few studies on tobacco BBX genes have been conducted to date. In this study, 49 non-redundant BBX genes have been identified in the reference tobacco genome. Gene structure, phylogenetic relationship, prediction of cis-regulatory elements in promoters, tissue-specific expression patterns, and response to cold stress were conducted on these BBX gene members. The expression of NtBBX9 and NtBBX11, two close members in the amplified group II, was up-regulated under cold treatments. Overexpression of NtBBX11 embedded the cold tolerance of transgenic plants, while the NtBBX9 overexpressors were susceptible to cold. The NtBBX9 functions as a negative regulator of tobacco response to cold stress, which was confirmed by the phenotype of NtBBX9 RNA-silenced lines. Furthermore, we demonstrated that NtBBX9 and NtBBX11 regulated the cold stress response by alternating the expression of critical cold-regulated genes such as CBFs, LEA14, and LTI65 and reactive oxygen species (ROS) production. The combination of NtBBX9 and NtBBX11 could facilitate tobacco’s rapid response to and recovery from transient cold stress and regulate plant cold-responsive growth in tropical regions.

Coping with the cold: unveiling cryoprotectants, molecular signaling pathways, and strategies for cold stress resilience.

Low temperature stress significantly threatens crop productivity and economic sustainability. Plants counter this by deploying advanced molecular mechanisms to perceive and respond to cold stress. Transmembrane proteins initiate these responses, triggering a series of events involving secondary messengers such as calcium ions (Ca2+), reactive oxygen species (ROS), and inositol phosphates. Of these, calcium signaling is paramount, activating downstream phosphorylation cascades and the transcription of cold-responsive genes, including cold-regulated (COR) genes. This review focuses on how plants manage freeze-induced damage through dual strategies: cold tolerance and cold avoidance. Tolerance mechanisms involve acclimatization to decreasing temperatures, fostering gradual accumulation of cold resistance. In contrast, avoidance mechanisms rely on cryoprotectant molecules like potassium ions (K+), proline, glycerol, and antifreeze proteins (AFPs). Cryoprotectants modulate intracellular solute concentration, lower the freezing point, inhibit ice formation, and preserve plasma membrane fluidity. Additionally, these molecules demonstrate antioxidant activity, scavenging ROS, preventing protein denaturation, and subsequently mitigating cellular damage. By forming extensive hydrogen bonds with water molecules, cryoprotectants also limit intercellular water movement, minimizing extracellular ice crystal formation, and cell dehydration. The deployment of cryoprotectants is a key adaptive strategy that bolsters plant resilience to cold stress and promotes survival in freezing environments. However, the specific physiological and molecular mechanisms underlying these protective effects remain insufficiently understood. Therefore, this review underscores the need for further research to elucidate these mechanisms and assess their potential impact on crop productivity and sustainability, contributing to the progressive discourse in plant biology and environmental science.

Natural variation in OsSEC13 HOMOLOG 1 modulates redox homeostasis to confer cold tolerance in rice.

Rice (Oryza sativa L.) is a cold-sensitive species that often faces cold stress, which adversely affects yield productivity and quality. However, the genetic basis for low-temperature adaptation in rice remains unclear. Here, we demonstrate that two functional polymorphisms in Oryza sativa SEC13 Homolog 1 (OsSEH1), encoding a WD40-repeat nucleoporin, between the two subspecies Oryza sativa japonica and Oryza sativa indica rice may have facilitated cold adaptation in japonica rice. We show that OsSEH1 of the japonica variety expressed in OsSEH1MSD plants (transgenic line overexpressing the OsSEH1 allele from Mangshuidao [MSD], cold-tolerant landrace) has a higher affinity for metallothionein 2b (OsMT2b) than that of OsSEH1 of indica. This high affinity of OsSEH1MSD for OsMT2b results in inhibition of OsMT2b degradation, with decreased accumulation of reactive oxygen species and increased cold tolerance. Transcriptome analysis indicates that OsSEH1 positively regulates the expression of the genes encoding dehydration-responsive element-binding transcription factors, i.e., OsDREB1 genes, and induces the expression of multiple cold-regulated genes to enhance cold tolerance. Our findings highlight a breeding resource for improving cold tolerance in rice.

Chemical screening approach using single leaves identifies compounds that affect cold signaling in Arabidopsis.

The identification of chemical compounds that affect intracellular processes has greatly contributed to our understanding of plant growth and development. In most cases, these compounds have been identified in germinated seedlings. However, chemical screening using mature plants would benefit and advance our understanding of environmental responses. In this study, we developed a high-throughput screening method using single leaves of mature plants to identify small molecules that affect cold-regulated gene expression. A single excised leaf of Arabidopsis (Arabidopsis thaliana) grown in submerged cultures responded to low temperatures in terms of COLD-REGULATED (COR) gene expression. We used transgenic Arabidopsis harboring a COLD-REGULATED 15A (COR15A) promoter::luciferase (COR15Apro::LUC) construct to screen natural compounds that affect the cold induction of COR15Apro::LUC. This approach allowed us to identify derivatives of 1,4-naphthoquinone as specific inhibitors of COR gene expression. Moreover, 1,4-naphthoquinones appeared to inhibit the rapid induction of upstream C-REPEAT BINDING FACTOR (CBF) transcription factors upon exposure to low temperature, suggesting that 1,4-naphthoquinones alter upstream signaling processes. Our study offers a chemical screening scheme for identifying compounds that affect environmental responses in mature plants. This type of analysis is likely to reveal an unprecedented link between certain compounds and plant environmental responses.

COLD REGULATED GENE 27 and 28 Antagonize the Transcriptional Activity of the RVE8/LNK1/LNK2 Circadian Complex.

Many molecular and physiological processes in plants occur at a specific time of day. These daily rhythms are coordinated in part by the circadian clock, a timekeeper that uses daylength and temperature to maintain rhythms of approximately 24 hours in various clock-regulated phenotypes. The circadian MYB-like transcription factor REVEILLE 8 (RVE8) interacts with its transcriptional coactivators NIGHT LIGHT INDUCIBLE AND CLOCK REGULATED 1 (LNK1) and LNK2 to promote the expression of evening-phased clock genes and cold tolerance factors. While genetic approaches have commonly been used to discover connections within the clock and between clock elements and other pathways, here we used affinity purification coupled with mass spectrometry to identify time-of-day-specific protein interactors of the RVE8-LNK1/LNK2 complex in Arabidopsis (Arabidopsis thaliana). Among the interactors of RVE8/LNK1/LNK2 were COLD REGULATED GENE 27 (COR27) and COR28, which coprecipitated in an evening-specific manner. In addition to COR27 and COR28, we found an enrichment of temperature-related interactors that led us to establish a previously uncharacterized role for LNK1 and LNK2 in temperature entrainment of the clock. We established that RVE8, LNK1, and either COR27 or COR28 form a tripartite complex in yeast (Saccharomyces cerevisiae) and that the effect of this interaction in planta serves to antagonize transcriptional activation of RVE8 target genes, potentially through mediating RVE8 protein degradation in the evening. Together, these results illustrate how a proteomic approach can be used to identify time-of-day-specific protein interactions. Discovery of the RVE8-LNK-COR protein complex indicates a previously unknown regulatory mechanism for circadian and temperature signaling pathways.

High-throughput miRNA sequencing and identification of a novel ICE1-targeting miRNA in response to low temperature stress in Eucalyptus camaldulensis.

MicroRNAs (miRNAs) play a crucial role in the growth, development, morphogenesis, signal transduction, and stress response in plants. The ICE (Inducer of CBF expression)-CBF (C-repeat binding factor)-COR (Cold-regulated gene) regulatory cascade is an important signalling pathway in plant response to low temperature stress, and it remains unknown whether this pathway is regulated by miRNAs. In this study, high-throughput sequencing was employed for predicting and identifying the miRNAs that were likely to target the ICE-CBF-COR pathway in Eucalyptus camaldulensis. A novel ICE1-targeting miRNA, eca-novel-miR-259-5p (nov-miR259), was further analysed. A total of 392 conserved miRNAs and 97 novel miRNAs were predicted, including 80 differentially expressed miRNAs. Of these, 30 miRNAs were predicted to be associated with the ICE-CBF-COR pathway. The full-length of mature nov-miR259 was 22 bp and its precursor gene was 60 bp in length, with a typical hairpin structure. The RNA ligase-mediated 5' amplification of cDNA ends (5'-RLM-RACE) and Agrobacterium-mediated tobacco transient expression assays demonstrated that nov-miR259 could cleave EcaICE1 in vivo. Moreover, qRT-PCR and Pearson's correlation analysis further revealed that the expression levels of nov-miR259 were almost significantly negativelycorrelated with those of its target gene, EcaICE1, and the other genes in the ICE-CBF-COR pathway. We first identified the nov-miR259 as a novel ICE1-targeting miRNA, and the nov-miR259-ICE1 module may be involved in regulating the cold stress response in E.camaldulensis.

Cross‐disciplinary insights into the mechanisms of plant cold hardiness: From molecules to ecosystems

Cross‐disciplinary insights into the mechanisms of plant cold hardiness: From molecules to ecosystems

Progress and Prospects of the Molecular Basis of Soybean Cold Tolerance.

Cold stress is a major factor influencing the geographical distribution of soybean growth and causes immense losses in productivity. Understanding the molecular mechanisms that the soybean has undergone to survive cold temperatures will have immense value in improving soybean cold tolerance. This review focuses on the molecular mechanisms involved in soybean response to cold. We summarized the recent studies on soybean cold-tolerant quantitative trait loci (QTLs), transcription factors, associated cold-regulated (COR) genes, and the regulatory pathways in response to cold stress. Cold-tolerant QTLs were found to be overlapped with the genomic region of maturity loci of E1, E3, E4, pubescence color locus of T, stem growth habit gene locus of Dt1, and leaf shape locus of Ln, indicating that pleiotropic loci may control multiple traits, including cold tolerance. The C-repeat responsive element binding factors (CBFs) are evolutionarily conserved across species. The expression of most GmDREB1s was upregulated by cold stress and overexpression of GmDREB1B;1 in soybean protoplast, and transgenic Arabidopsis plants can increase the expression of genes with the DRE core motif in their promoter regions under cold stress. Other soybean cold-responsive regulators, such as GmMYBJ1, GmNEK1, GmZF1, GmbZIP, GmTCF1a, SCOF-1 and so on, enhance cold tolerance by regulating the expression of COR genes in transgenic Arabidopsis. CBF-dependent and CBF-independent pathways are cross-talking and work together to activate cold stress gene expression. Even though it requires further dissection for precise understanding, the function of soybean cold-responsive transcription factors and associated COR genes studied in Arabidopsis shed light on the molecular mechanism of cold responses in soybeans and other crops. Furthermore, the findings may also provide practical applications for breeding cold-tolerant soybean varieties in high-latitude and high-altitude regions.

Integrated Metabolomic and Transcriptomic Analysis Reveals That Amino Acid Biosynthesis May Determine Differences in Cold-Tolerant and Cold-Sensitive Tea Cultivars.

Cold stress is one of the major abiotic stresses limiting tea production. The planting of cold-resistant tea cultivars is one of the most effective measures to prevent chilling injury. However, the differences in cold resistance between tea cultivars remain unclear. In the present study, we perform a transcriptomic and metabolomic profiling of Camellia sinensis var. "Shuchazao" (cold-tolerant, SCZ) and C. sinensis var. assamica "Yinghong 9" (cold-sensitive, YH9) during cold acclimation and analyze the correlation between gene expression and metabolite biosynthesis. Our results show that there were 51 differentially accumulated metabolites only up-regulated in SCZ in cold-acclimation (CA) and de-acclimation (DA) stages, of which amino acids accounted for 18%. The accumulation of L-arginine and lysine in SCZ in the CA stage was higher than that in YH9. A comparative transcriptomic analysis showed an enrichment of the amino acid biosynthesis pathway in SCZ in the CA stage, especially "arginine biosynthesis" pathways. In combining transcriptomic and metabolomic analyses, it was found that genes and metabolites associated with amino acid biosynthesis were significantly enriched in the CA stage of SCZ compared to CA stage of YH9. Under cold stress, arginine may improve the cold resistance of tea plants by activating the polyamine synthesis pathway and CBF (C-repeat-binding factor)-COR (cold-regulated genes) regulation pathway. Our results show that amino acid biosynthesis may play a positive regulatory role in the cold resistance of tea plants and assist in understanding the cold resistance mechanism differences among tea varieties.

The involvement of Antifreeze protein maxi-like and Cold-shock domain-containing protein genes in cold-induced larval diapause and cold-shock treatment of khapra beetle

The khapra beetle, Trogoderma granarium (Everts) (Coleoptera: Dermestideae), is a cosmopolitan and one of the most destructive pests of various stored grains and grain products. This pest is categorized as a quarantine threat in many countries. The larvae undergo diapause that lasts for several years depending on different stress conditions such as temperature, insecticidal applications, starvation, humidity, crowding, and fecal pellets. When conditions are favorable for the development, diapause terminates and the larva continues its development. In the present study, two cold-regulated genes Antifreeze protein maxi-like (TgAFP) and Cold-shock domain-containing protein E1 (TgCSDP) were identified in the cDNA library of T. granarium. Nucleotide sequences of two unigene cDNAs encoding TgAFP and TgCSDP were verified using RT-PCR and RACE-PCR amplifications. Transcriptional regulations of TgAFP and TgCSDP were examined in cold-induced diapause larvae, cold-shocked larvae, and at different developmental stages. Gene expression pattern of TgAFP revealed the highest mRNA levels during pre-diapause (25 °C) followed by the larvae exposed to 5 °C during the diapause phase. This can be attributable to the protective role of TgAFP against temperature fall. Significant upregulation of TgCSDP at 15 °C might indicate its probable chaperone role in toleration for cold-induced diapause. TgAFP level was downregulated after cold-shock treatment (CST), while it was slightly upregulated by recovery, indicating that it might confer tolerance for the recovery period (RP). The highest TgCSDP expression after the CST might suggest its involvement in response to acute CST. Low abundance of TgAFP expression in each developmental stage might suggest that 33 °C temperature does not induce the synthesis of TgAFP. Significant amount of TgCSDP levels in adults might indicate its putative role in development. These findings suggest that TgAFP and TgCSDP genes might be crucial for cold survival where the T. granarium undergoes its facultative diapause.

Biochemical and Transcriptional Responses in Cold-Acclimated and Non-Acclimated Contrasting Camelina Biotypes under Freezing Stress.

Cold-acclimated and non-acclimated contrasting Camelina (Camelina sativa L.) biotypes were investigated for changes in stress-associated biomarkers, including antioxidant enzyme activity, lipid peroxidation, protein, and proline content. In addition, a well-known freezing tolerance pathway participant known as C-repeat/DRE-binding factors (CBFs), an inducer of CBF expression (ICE1), and a cold-regulated (COR6.6) genes of the ICE-CBF-COR pathway were studied at the transcriptional level on the doubled-haploid (DH) lines. Freezing stress had significant effects on all studied parameters. The cold-acclimated DH34 (a freezing-tolerant line) showed an overall better performance under freezing stress than non-acclimated plants. The non-cold-acclimated DH08 (a frost-sensitive line) showed the highest electrolyte leakage after freezing stress. The highest activity of antioxidant enzymes (glutathione peroxidase, superoxide dismutase, and catalase) was also detected in non-acclimated plants, whereas the cold-acclimated plants showed lower enzyme activities upon stress treatment. Cold acclimation had a significantly positive effect on the total protein and proline content of stressed plants. The qRT-PCR analysis revealed significant differences in the expression and cold-inducibility of CsCBF1-3, CsICE1, and CsCOR6.6 genes among the samples of different treatments. The highest expression of all CBF genes was recorded in the non-acclimated frost-tolerant biotype after freezing stress. Interestingly a significantly higher expression of COR6.6 was detected in cold-acclimated samples of both frost-sensitive and -tolerant biotypes after freezing stress. The presented results provide more insights into freezing tolerance mechanisms in the Camelina plant from both a biochemical point of view and the expression of the associated genes.

StLTO1, a lumen thiol oxidoreductase in Solanum tuberosum L., enhances the cold resistance of potato plants

Cold stress reduces plant photosynthesis and increases the accumulation of reactive oxygen species (ROS) in plants, thereby dramatically affecting plant growth, crop productivity and quality. Here, we report that lumen thiol oxidoreductase 1 (StLTO1), a vitamin K epoxide reductase (VKOR)-like protein in the thylakoid membrane of Solanum tuberosum L., enhances the cold tolerance of potato plants. Under normal conditions, overexpression of StLTO1 in plants promoted plant growth. In addition, potato plants overexpressing StLTO1 displayed enhanced photosynthetic capacity and increased capacity for scavenging ROS compared to StLTO1 knockdown and wild-type potato plants under cold conditions. Overexpression of StLTO1 in potato plants also improved cold-regulated (COR) gene expression after cold stress. Our results suggest that StLTO1 acts as a positive regulator of cold resistance in potato plants.

CsCBF5 depletion impairs cold tolerance in tea plants

CBFs play important roles in tea plant cold tolerance. In our study, 16 tea varieties were used to investigate the relationship between the expression level of CsCBFs and cold tolerance in field experiments. A strong and positive correlation was found between cold stress-regulated CsCBF1, CsCBF3 and CsCBF5 expression levels (R2 > 0.8) in tea mesophyll cells and cold tolerance in 16 tea varieties. A previous study reported that CsCBF1 and CsCBF3 were important components associated with cold tolerance in tea plants; thus, the function of CsCBF5 in the CsCBF family was targeted. Our previous study reported that CsCBF5 was localized in the nucleus and exhibited transcriptional activity. In the current study, MDA content in leaves was significantly increased in CsCBF5-silenced leaves, which exhibited poor cold tolerance, compared with WT plants under cold stress. In contrast, increased germination rates and antioxidant enzyme activities under cold conditions compared with WT plants. Furthermore, CsCBF5 overexpression in Arabidopsis promoted the expression levels of the cold-regulated genes AtCOR15a, AtCOR78, AtERD4 and AtRD29B; however, the expression levels of downstream genes, including CsCOR47, CsCOR413, CsERD4 and CsRD29B, were significantly reduced in CsCBF5-silenced tea leaves. Taken together, our results indicated that CsCBF5 could function as a positive regulator in the cold stress response.