C-repeat Binding Factor Research Articles

C-repeat binding factor (CBF) identification and expression pattern to abiotic stress response in Casuarina equisetifolia and overexpression of CeCBF4 increase stress tolerance in Arabidopsis

Casuarina equisetifolia stands out as a highly valuable tree species crucial for industries and ecological restoration projects in tropical and subtropical regions, owing to its exceptional salt resistance. However, its growth is hampered in cold and drought-prone areas. Understanding the genetic basis for abiotic stress tolerance is imperative for enhancing stress-resistant breeding efforts of C. equisetifolia. However, the CBF genes are renowned for their pivotal roles in cold response and broad resilience to other adverse environments, such as drought, and high salinity. In this study, five CeCBFs were identified from the genome of C. equisetifolia. CeCBF1 and CeCBF3 belong to Clade I group, which diverged early from DREBIII members, while CeCBF2, CeCBF4, and CeCBF5 are classified into Clade II, a group innovated from Clade I. These genes lack introns in their sequences. The proteins encoded by CeCBFs feature classic AP2 domains and CBF signature sequences, localize in the nucleus and exhibit transactivation activity. Additionally, stress-related, abscisic acid, and jasmonic acid responsive elements are present on the CeCBFs promoters. Expression analysis revealed that CeCBFs were prodominantly induced by cold, drought, and salinity treatments, except for CeCBF1, which showed repression specifically under low temperature. Moreover, CeCBF1, CeCBF4, and CeCBF5 were inhibited by abscisic acid, while CeCBF1 showed reduced transcript levels in response to methyl jasmonate treatment. Furthermore, overexpression of CeCBF4 enhanced tolerance to low temperature, depriving of water, and salt by up-regulating stress-responsive genes expression and promoting proline accumulation, albeit with a trade-off in growth and productivity. Hence, CeCBFs emerge as potential regulators contributing significantly to stress tolerance mechanisms in C. equisetifolia.

Understanding cold stress response mechanisms in plants: an overview

Low-temperature stress significantly impacts plant growth, development, yield, and geographical distribution. However, during the long-term process of evolution, plants have evolved complicated mechanisms to resist low-temperature stress. The cold tolerance trait is regulated by multiple pathways, such as the Ca2+ signaling cascade, mitogen-activated protein kinase (MAPK) cascade, inducer of CBF expression 1 (ICE1)-C-repeat binding factor (CBF)-cold-reulated gene (COR) transcriptional cascade, reactive oxygen species (ROS) homeostasis regulation, and plant hormone signaling. However, the specific responses of these pathways to cold stress and their interactions are not fully understood. This review summarizes the response mechanisms of plants to cold stress from four aspects, including cold signal perception and transduction, ICE1-CBF-COR transcription cascade regulation, ROS homeostasis regulation and plant hormone signal regulation. It also elucidates the mechanism of cold stress perception and Ca2+ signal transduction in plants, and proposes the important roles of transcription factors (TFs), post-translational modifications (PTMs), light signals, circadian clock factors, and interaction proteins in the ICE1-CBF-COR transcription cascade. Additionally, we analyze the importance of ROS homeostasis and plant hormone signaling pathways in plant cold stress response, and explore the cross interconnections among the ICE1-CBF-COR cascade, ROS homeostasis, and plant hormone signaling. This comprehensive review enhances our understanding of the mechanism of plant cold tolerance and provides a molecular basis for genetic strategies to improve plant cold tolerance.

The nuclear exosome subunit HEN2 acts independently of the core exosome to assist transcription in Arabidopsis.

Regulation of gene expression is at the frontier of plant responses to various external stimuli including stress. RNA polymerase-based transcription and post-transcriptional degradation of RNA play vital roles in this regulation. Here, we show that HUA ENHANCER 2 (HEN2), a co-factor of the nuclear exosome complex, influences RNAPII transcription elongation in Arabidopsis (Arabidopsis thaliana) under cold conditions. Our results demonstrate that a hen2 mutant is cold sensitive and undergoes substantial transcriptional changes compared to wild type when exposed to cold conditions. We found an accumulation of 5' fragments from a subset of genes (including C-repeat binding factors 1-3 [CBF1-3]) that do not carry over to their 3' ends. In fact, hen2 mutants have lower levels of full-length mRNA for a subset of genes. This distinct 5'-end accumulation and 3'-end depletion was not observed in other NEXT complex members or core exosome mutants, highlighting HEN2's distinctive role. We further used RNAPII-associated nascent RNA to confirm the transcriptional phenotype is a result of lower active transcription specifically at the 3'-end of these genes in a hen2 mutant. Taken together, our data point to the unique role of HEN2 in maintaining RNAPII transcription dynamics especially highlighted under cold stress.

An AP2/ERF transcription factor confers chilling tolerance in rice.

Cold stress, a prominent adverse environmental factor, severely hinders rice growth and productivity. Unraveling the complex mechanisms governing chilling tolerance in rice is crucial for molecular breeding of cold-tolerant varieties. Here, we identify an APETALA2/Ethylene Responsive Factor (AP2/ERF) transcription factor, OsERF52, as a positive modulator in response to low temperatures. OsERF52 directly regulates the expression of C-Repeat Binding Factor (CBF) genes in rice. In addition, Osmotic Stress/ABA-Activated Protein Kinase 9-mediated phosphorylation of OsERF52 at S261 enhances its stability and interaction with Ideal Plant Architecture 1 and OsbHLH002/OsICE1. This collaborative activation leads to the expression of OsCBFs, thereby initiating the chilling response in rice. Notably, plants with base-edited OsERF52S261D-3HA exhibit enhanced chilling resistance without yield penalty. Our findings unveil the mechanism orchestrated by a regulatory framework involving a protein kinase and transcription factors from diverse families, offering potential genetic resources for developing chilling-tolerant rice varieties.

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.

HOS15-mediated turnover of PRR7 enhances freezing tolerance.

Arabidopsis PSEUDORESPONSE REGULATOR7 (PRR7) is a core component of the circadian oscillator which also plays a crucial role in freezing tolerance. PRR7 undergoes proteasome-dependent degradation to discretely phase maximal expression in early evening. While its repressive activity on downstream genes is integral to cold regulation, the mechanism of the conditional regulation of the PRR7 abundance is unknown. We used mutant analysis, protein interaction and ubiquitylation assays to establish that the ubiquitin ligase adaptor, HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENE 15 (HOS15), controls the protein accumulation pattern of PRR7 through direct protein-protein interactions at low temperatures. Freezing tolerance and electrolyte leakage assays show that PRR7 enhances cold temperature sensitivity, supported by ChIP-qPCR at C-REPEAT BINDING FACTOR1 (CBF1) and COLD-REGULATED 15A (COR15A) promoters where PRR7 levels were higher in hos15 mutants. HOS15 mediates PRR7 turnover through enhanced ubiquitylation at low temperature in the dark. Under the same conditions, increased PRR7 association with the promoters of CBFs and COR15A in hos15 correlates with decreased CBF1 and COR15A transcription and enhanced freezing sensitivity. We propose a novel mechanism whereby HOS15-mediated degradation of PRR7 provides an intersection between the circadian system and other cold acclimation pathways that lead to increased freezing tolerance.

CaMYB80 enhances the cold tolerance of pepper by directly targeting CaPOA1.

Cold temperatures negatively impact crop yield and quality, posing significant limitations to the advancement of the vegetable industry. MYB transcription factors are pivotal in enhancing plant resilience against various abiotic stresses, including low-temperature stress. Pepper (Capsicum annuum L.) is a nutrient-rich vegetable crop sensitive to low temperatures. This study aimed to determine the function of CaMYB80 in the cold stress response of pepper through virus-induced silencing. The study also conducted heterologous expression of CaMYB80 in Arabidopsis and tomato plants. The results showed that CaMYB80 could respond to low-temperature stress in pepper. CaMYB80 was localized in the nucleus and cytoplasm and exhibited transcriptional activation ability. Moreover, CaMYB80 silencing decreased cold tolerance in pepper, while its heterologous overexpression increased cold tolerance in Arabidopsis and tomato. Further analysis showed that CaMYB80 interacted with CaPOA1 (peroxidase N1-like). Similarly, the expression of CaPOA1 also responded to low-temperature stress. Overexpression of CaPOA1 enhanced freezing tolerance in Arabidopsis, while its silencing reduced cold stress tolerance in pepper. Furthermore, overexpression of CaMYB80 in Arabidopsis and tomato could increase the activity of peroxidases and the expression levels of genes in the ICE-CBF-COR (inducer of CBF expression, C-repeat binding factor, cold-responsive) regulatory network. In conclusion, our research results indicate that CaMYB80 enhances pepper cold tolerance by interacting with CaPOA1 to increase peroxidase activity and influence the expression of ICE-CBF-COR related genes.

Comparing time-series transcriptomes between chilling-resistant and -susceptible rice reveals potential transcription factors responding to chilling stress.

Low temperature is one of the most important environmental factors that inhibits rice growth and grain yield. Transcription factors (TFs) play crucial roles in chilling acclimation by regulating gene expression. However, transcriptional dynamics and key regulators responding to low temperature remain largely unclear in rice. In this study, a transcriptome-based comparative analysis was performed to explore genome-wide gene expression profiles between a chilling-resistant cultivar DC90 and a chilling-susceptible cultivar 9311 at a series of time points under low temperature treatment and recovery condition. A total of 3,590 differentially expressed genes (DEGs) between two cultivars were determined and divided into 12 co-expression modules. Meanwhile, several biological processes participating in the chilling response such as abscisic acid (ABA) responses, water deprivation, protein metabolic processes, and transcription regulator activities were revealed. Through weighted gene co-expression network analysis (WGCNA), 15 hub TFs involved in chilling conditions were identified. Further, we used the gene regulatory network (GRN) to evaluate the top 50 TFs, which might have potential roles responding to chilling stress. Finally, five TFs, including a C-repeat binding factor (OsCBF3), a zinc finger-homeodomain protein (OsZHD8), a tandem zinc finger protein (OsTZF1), carbon starved anther (CSA), and indeterminate gametophyte1 (OsIG1) were identified as crucial candidates responsible for chilling resistance in rice. This study deepens our understanding in the gene regulation networks of chilling stress in rice and offers potential gene resources for breeding climate-resilient crops.

HOS15-mediated turnover of PRR7 enhances freezing tolerance.

Arabidopsis PSEUDO RESPONSE REGULATOR7 (PRR7) is a core component of the circadian oscillator which also plays a crucial role in freezing tolerance. PRR7 undergoes proteasome-dependent degradation to discretely phase maximal expression in early evening. While its transcriptional repressive activity on downstream genes is integral to cold regulation, the mechanism of the conditional regulation of the PRR7 protein activity is unknown. We used double mutant analysis, protein interaction and ubiquitylation assays to establish that the ubiquitin ligase adaptor, HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENE 15 (HOS15), controls the protein accumulation pattern of PRR7 through direct protein-protein interactions. Freezing tolerance and electrolyte leakage assays show that PRR7 enhances cold temperature sensitivity, supported by ChIP-qPCR at C-REPEAT BINDING FACTOR (CBF) and COLD REGULATED 15A (COR15A) promoters where PRR7 levels were higher in hos15 mutants. We establish that HOS15 mediates PRR7 protein turnover through enhanced ubiquitylation at low temperature in the dark. Under the same conditions, increased PRR7 association with the promoter regions of CBFs and COR15A in hos15 correlates with decreased CBF1 and COR15A transcription and enhanced freezing sensitivity. We propose a novel mechanism whereby HOS15-mediated regulation of PRR7 provides an intersection between the circadian system and other cold acclimation pathways leading to freezing tolerance through upregulation of CBF1 and COR15A.

The NAC056 transcription factor confers freezing tolerance by positively regulating expression of CBFs and NIA1 in Arabidopsis

Freezing stress can seriously affect plant growth and development, but the mechanisms of these effects and plant responses to freezing stress require further exploration. Here, we identified a NAM, ATAF1/2, and CUC2 (NAC)-family transcription factor (TF), NAC056, that can promote freezing tolerance in Arabidopsis. NAC056 mRNA levels are strongly induced by freezing stress in roots, and the nac056 mutant exhibits compromised freezing tolerance. NAC056 acts positively in response to freezing by directly promoting key C-repeat-binding factor (CBF) pathway genes. Interestingly, we found that CBF1 regulates nitrate assimilation by regulating the nitrate reductase gene NIA1 in plants; therefore, NAC056–CBF1–NIA1 form a regulatory module for the assimilation of nitrate and the growth of roots under freezing stress. In addition, 35S::NAC056 transgenic plants show enhanced freezing tolerance, which is partially reversed in the cbfs triple mutant. Thus, NAC056 confers freezing tolerance through the CBF pathway, mediating plant responses to balance growth and freezing stress tolerance.

Freezing treatment under light conditions leads to a dramatic enhancement of freezing tolerance in cold-acclimated Arabidopsis.

Overwintering plants survive subzero temperatures by cold acclimation (CA), wherein they acquire freezing tolerance through short-term exposure to low temperatures above 0°C. The freezing tolerance of CA plants increases when they are subsequently exposed to mild subzero temperatures, a phenomenon known as second-phase cold hardening (2PH). Here, we explored the molecular mechanism and physiological conditions of 2PH. The results show that, compared with supercooling, a freezing treatment during 2PH after CA enhanced the freezing tolerance of Arabidopsis. This required CA as a pretreatment, and was designated as second-phase freezing acclimation (2PFA). Light increased the effect of 2PFA to enhance freezing tolerance after CA. C-repeat binding factor and cold-regulatedgenes were downregulated by light during the 2PFA treatment, a different transcription profile from that during CA. The freezing tolerance of 2PFA plants was decreased by the presence of the photosynthetic electron transfer inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylureaduring the 2PFA treatment. Compared with wild-type plants, phototropin1,2 and phyb mutants showed lower freezing tolerance after 2PFA treatment. These results show that exposure to freezing after CA increases freezing tolerance as a secondary process, and that freezing under light conditions further increases freezing tolerance via pathways involving photoreceptors and photosynthetic electron transfer.

Target enrichment sequencing coupled with GWAS identifies MdPRX10 as a candidate gene in the control of budbreak in apple.

The timing of floral budbreak in apple has a significant effect on fruit production and quality. Budbreak occurs as a result of a complex molecular mechanism that relies on accurate integration of external environmental cues, principally temperature. In the pursuit of understanding this mechanism, especially with respect to aiding adaptation to climate change, a QTL at the top of linkage group (LG) 9 has been identified by many studies on budbreak, but the genes underlying it remain elusive. Here, together with a dessert apple core collection of 239 cultivars, we used a targeted capture sequencing approach to increase SNP resolution in apple orthologues of known or suspected A. thaliana flowering time-related genes, as well as approximately 200 genes within the LG9 QTL interval. This increased the 275 223 SNP Axiom® Apple 480K array dataset by an additional 40 857 markers. Robust GWAS analyses identified MdPRX10, a peroxidase superfamily gene, as a strong candidate that demonstrated a dormancy-related expression pattern and down-regulation in response to chilling. In-silico analyses also predicted the residue change resulting from the SNP allele associated with late budbreak could alter protein conformation and likely function. Late budbreak cultivars homozygous for this SNP allele also showed significantly up-regulated expression of C-REPEAT BINDING FACTOR (CBF) genes, which are involved in cold tolerance and perception, compared to reference cultivars, such as Gala. Taken together, these results indicate a role for MdPRX10 in budbreak, potentially via redox-mediated signaling and CBF gene regulation. Moving forward, this provides a focus for developing our understanding of the effects of temperature on flowering time and how redox processes may influence integration of external cues in dormancy pathways.

Genome-Wide Analysis of C-Repeat Binding Factor Gene Family in Capsicum baccatum and Functional Exploration in Low-Temperature Response.

Capsicum baccatum is a close relative of edible chili peppers (Capsicum annuum) with high economic value. The CBF gene family plays an important role in plant stress resistance physiology. We detected a total of five CBF genes in the C. baccatum genome-wide sequencing data. These genes were scattered irregularly across four chromosomes. The genes were categorized into three groupings according to their evolutionary relationships, with genes in the same category showing comparable principles for motif composition. The 2000 bp upstream of CbCBF contains many resistance-responsive elements, hormone-responsive elements, and transcription factor binding sites. These findings emphasize the crucial functions of these genes in responding to challenging conditions and physiological regulation. Analysis of tissue-specific expression revealed that CbCBF3 exhibited the greatest level of expression among all tissues. Under conditions of low-temperature stress, all CbCBF genes exhibited different levels of responsiveness, with CbCBF3 showing a considerable up-regulation after 0.25 h of cold stress, indicating a high sensitivity to low-temperature response. The importance of the CbCBF3 gene in the cold response of C. baccatum was confirmed by the use of virus-induced gene silencing (VIGS) technology, as well as the prediction of its protein interaction network. To summarize, this study conducts a thorough bioinformatics investigation of the CbCBF gene family, showcases the practicality of employing VIGS technology in C. baccatum, and confirms the significance of the CbCBF3 gene in response to low temperatures. These findings provide significant references for future research on the adaptation of C. baccatum to low temperatures.

The cold-responsive C-repeat binding factors in Betula platyphylla Suk. positively regulate cold tolerance

Cold stress is one of the most destructive abiotic stresses limiting plant growth and development. CBF (C-repeat binding factor) transcription factors and their roles in cold response have been identified in Arabidopsis as well as several other plant species. However, the biological functions and related molecular mechanisms of CBFs in birch (Betula platyphylla Suk.) remain undetermined. In this study, five cold-responsive BpCBF genes, BpCBF1, BpCBF2, BpCBF7, BpCBF10 and BpCBF12 were cloned. Via protoplast transformation, BpCBF7 was found to be localized in nucleus. The result of yeast one hybrid assay validated the binding of BpCBF7 to the CRT/DRE (C-repeat/dehydration responsive element) elements in the promoter of BpERF1.1 gene. By overexpressing and repressing BpCBFs in birch plants, it was proven that BpCBFs play positive roles in the cold tolerance. At the metabolic level, BpCBFs OE lines had lower ROS accumulation, as well as higher activities of antioxidant enzymes (SOD, POD and CAT) and higher accumulation of protective substances (soluble sugar, soluble protein and proline). Via yeast one hybrid and co-transformation of effector and reporter vectors assay, it was proven that BpCBF7 can regulate the expression of BpERF5 and BpZAT10 genes by directly binding to their promoters. An interacting protein of BpCBF7, BpWRKY17, was identified by yeast two hybrid library sequencing and the interaction was validated with in vivo methods. These results indicates that BpCBFs can increase the cold tolerance of birch plants, partly by gene regulation and protein interaction. This study provides a reference for the research on CBF transcription factors and genetic improvement of forest trees upon abiotic stresses.

Physiological and transcriptomic analyses reveal mechanisms of exogenous strigolactones to regulate cold tolerance in litchi fruit

Strigolactones (SLs) are a class of carotenoid-derived phytohormones that play diverse biological functions in plants. In the current study, we conducted physiological and transcriptomic analysis to explore the effects of two different concentrations (0.1 and 1.0 μM) of GR24, a synthetic analogue of SLs, on chilling injury (CI) and related mechanisms in litchi fruit during refrigeration at 4 °C. The results showed that 0.1 μM GR24 (GR24–0.1) dipping treatment retarded the advancement of browning and red loss in litchi pericarp during refrigeration. GR24–0.1 treatment ameliorated oxidative stress and restrained membrane deterioration, as manifested by lowered reactive oxygen species (ROS) production, relative electrolyte leakage (REL) and malondialdehyde (MDA) content. GR24–0.1 treatment inhibited the enhancement of polyphenol oxidase (PPO) and peroxidase (POD) activities, which was concomitant with diminished contents of total phenolics, flavonoids and anthocyanins. Transcriptomic analysis revealed that GR24–0.1 treatment up-regulated the expression of genes associated with abscisic acid (ABA) biosynthesis and signal transduction, C-repeat binding factor (CBF) signaling pathway, anthocyanin synthesis and transport, antioxidant system, oxidative protein repair system as well as energy metabolism, contributing to the enhanced levels of some metabolites in these biochemical pathways. In contrast to GR24–0.1 treatment, reverse transcriptional and physiological effects in parallel with aggravated CI severity occurred in GR24–1.0 treated fruit, indicating a dosage-dependent regulatory effect of GR24 on litchi CI. The present findings provide insights into the potential mechanisms by which SLs regulate cold tolerance in postharvest litchi fruit.

The MdCBF1/2-MdTST1/2 module regulates sugar accumulation in response to low temperature in apple.

Soluble sugar content is a key component in controlling fruit flavor, and its accumulation in fruit is largely determined by sugar metabolism and transportation. When the diurnal temperature range is greater, the fleshy fruits accumulated more soluble sugars and become more sweeter. However, the molecular mechanism underlying this response remains largely unknown. In this study, we verified that low-temperature treatment promoted soluble sugar accumulation in apple fruit and found that this was due to the upregulation of the Tonoplast Sugar Transporter genes MdTST1/2. A combined strategy using assay for transposase-accessible chromatin (ATAC) sequencing and gene expression and cis-acting elements analyses, we identified two C-repeat Binding Factors, MdCBF1 and MdCBF2, that were induced by low temperature and that might be upstream transcription factors of MdTST1/2. Further studies established that MdCBF1/2 could bind to the promoters of MdTST1/2 and activate their expression. Overexpression of MdCBF1 or MdCBF2 in apple calli and fruit significantly upregulated MdTST1/2 expression and increased the concentrations of glucose, fructose, and sucrose. Suppression of MdTST1 and/or MdTST2 in an MdCBF1/2-overexpression background abolished the positive effect of MdCBF1/2 on sugar accumulation. In addition, simultaneous silencing of MdCBF1/2 downregulated MdTST1/2 expression and apple fruits failed to accumulate more sugars under low-temperature conditions, indicating that MdCBF1/2-mediated sugar accumulation was dependent on MdTST1/2 expression. Hence, we concluded that the MdCBF1/2-MdTST1/2 module is crucial for sugar accumulation in apples in response to low temperatures. Our findings provide mechanistic components coordinating the relationship between low temperature and sugar accumulation as well as new avenues to improve fruit quality.

Экспрессия транскрипционных факторов CBF (C-REPEAT BINDING FACTOR) у проростков ржи посевной (Secale cereale L.) при холодовом стрессе

Winter rye is one of the most important food crops in the world. Due to its high adaptability, winter hardiness and ability to yield on low-fertility soils, winter rye helps stabilize the gross grain harvest. For the vast majority of crop plants, the main stress-causing environmental factor is temperature. This is especially true for its lower values in cultivation areas. The response to low temperatures in plants is formed at the cellular level. Genes and transcription factors responsible for cold resistance are activated, which ensure plant survival. One of the most important mechanisms capable of activating the adaptive response of rye to cold is the CBF (C-repeat Binding Factor) family of transcription factors. The aim of this work was to identify the expression level of some genes of the ScCBF family (ScCBF1, ScCBF4, ScCBF14, ScCBF18) in seedlings of sowing rye (Secale cereale L.) under cold stress. S. cereale seedlings were divided into four groups based on the duration of cold stress (1, 6, 12, 24 h). One control group was not exposed to cold stress. RNA of seedlings from different cold stress groups was analyzed by real-time PCR. The study showed that the expression levels of ScCBF1, ScCBF4, ScCBF14, and ScCBF18 genes changed to varying degrees in response to low positive temperature. The highest level of normalized expression of ScCBF1 and ScCBF14 genes was observed under 6-hour cold stress. In the case of ScCBF4 gene, the highest level of its normalized expression was observed under 24-hour cold stress. The level of normalized expression of ScCBF18 gene reached its peak under 12-hour stress.

Frost survival and gene expression in timothy (Phleum pratense L.) cultivars as affected by age and selection in diverse field environments.

The sustainable production of perennial grasses in Northern Norway is at risk due to the ongoing climate change. The predicted increase in temperatures and variable weather patterns are further expected to create challenges for winter survival of timothy (Phleum pratense L.). Knowledge about the molecular mechanisms underlying freezing tolerance is crucial for developing robust cultivars. The current study is aimed at identifying genes involved in freezing stress response of timothy and studying gene expression differentiation due to field selection in contrasting environments using RNAseq. Four timothy cultivars were field tested for three years in Tromsø and Vesterålen, in Northern Norway. The surviving material from the field tests, along with plants raised from the original seed lots, were subjected to freezing tests. LT50 values varied across cultivars and materials. Many genes coding for transcription factors and proteins known to play an important role in freezing tolerance, like dehydrins, c-repeat binding factors, and late embryogenesis abundant proteins were upregulated with decreasing temperatures. Moreover, genes associated with glycolysis/gluconeogenesis, TCA cycle, glutathione metabolism, proteasome pathways and genes encoding autophagy-related proteins, plasma membrane-associated proteins, sugar and amino acid transporters had elevated expression in field survivors compared to plants raised from the original material. The lower freezing stress tolerance of field survivors despite the elevated expression of several stress-responsive genes might be due to a combination of selection in the field and the age effect. Furthermore, differences in freezing stress response between northern and southern adapted cultivars and surviving material from two field trial locations are discussed.

Melatonin as a key regulator in seed germination under abiotic stress.

Seed germination (SG) is the first stage in a plant's life and has an immense importance in sustaining crop production. Abiotic stresses reduce SG by increasing the deterioration of seed quality, and reducing germination potential, and seed vigor. Thus, to achieve a sustainable level of crop yield, it is important to improve SG under abiotic stress conditions. Melatonin (MEL) is an important biomolecule that interplays in developmental processes and regulates many adaptive responses in plants, especially under abiotic stresses. Thus, this review specifically summarizes and discusses the mechanistic basis of MEL-mediated SG under abiotic stresses. MEL regulates SG by regulating some stress-specific responses and some common responses. For instance, MEL induced stress specific responses include the regulation of ionic homeostasis, and hydrolysis of storage proteins under salinity stress, regulation of C-repeat binding factors signaling under cold stress, starch metabolism under high temperature and heavy metal stress, and activation of aquaporins and accumulation of osmolytes under drought stress. On other hand, MEL mediated regulation of gibberellinsbiosynthesis and abscisic acidcatabolism, redox homeostasis, and Ca2+ signaling are amongst the common responses. Nonetheless factors such as endogenous MEL contents, plant species, and growth conditions also influence above-mentioned responses. In conclusion, MEL regulates SG under abiotic stress conditions by interacting with different physiological mechanisms.

SVALKA-POLYCOMB REPRESSIVE COMPLEX2 module controls C-REPEAT BINDING FACTOR3 induction during cold acclimation.

C-REPEAT BINDING FACTORS (CBFs) are highly conserved plant transcription factors that promote cold tolerance. In Arabidopsis (Arabidopsis thaliana), three CBFs (CBF1 to CBF3) play a critical role in cold acclimation, and the expression of their corresponding genes is rapidly and transiently induced during this adaptive response. Cold induction of CBFs has been extensively studied and shown to be tightly controlled, yet the molecular mechanisms that restrict the expression of each CBF after their induction during cold acclimation are poorly understood. Here, we present genetic and molecular evidence that the decline in the induction of CBF3 during cold acclimation is epigenetically regulated through the Polycomb Repressive Complex (PRC) 2. We show that this complex promotes the deposition of the repressive mark H3K27me3 at the coding region of CBF3, silencing its expression. Our results indicate that the cold-inducible long noncoding RNA SVALKA is essential for this regulation by recruiting PRC2 to CBF3. These findings unveil a SVALKA-PRC2 regulatory module that ensures the precise timing of CBF3 induction during cold acclimation and the correct development of this adaptive response.