Cold Stress Tolerance In Plants Research Articles

Cold Tolerance of ScCBL6 Is Associated with Tonoplast Transporters and Photosynthesis in Arabidopsis.

Plants that are adapted to harsh environments offer enormous opportunity to understand stress responses in ecological systems. Stipa capillacea is widely distributed in the frigid and arid region of the Tibetan Plateau, but its signal transduction system under cold stress has not been characterized. In this study, we isolated a cDNA encoding the signal transduction protein, ScCBL6, from S. capillacea, and evaluated its role in cold tolerance by ectopically expressing it in Arabidopsis. Full-length ScCBL6 encode 227 amino acids, and are clustered with CBL6 in Stipa purpurea and Oryza sativa in a phylogenetic analysis. Compared with tolerance in wild-type (WT) plants, ScCBL6-overexpressing plants (ScCBL6-OXP) were more tolerant to cold stress but not to drought stress, as confirmed by their high photosynthetic capacity (Fv/Fm) and survival rate under cold stress. We further compared their cold-responsive transcriptome profiles by RNA sequencing. In total, 3931 genes were differentially expressed by the introduction of ScCBL6. These gene products were involved in multiple processes such as the immune system, lipid catabolism, and secondary metabolism. A KEGG pathway analysis revealed that they were mainly enriched in plant hormone signal transduction and biomacromolecule metabolism. Proteins encoded by differentially expressed genes were predicted to be localized in chloroplasts, mitochondria, and vacuoles, suggesting that ScCBL6 exerts a wide range of functions. Based on its tonoplast subcellular location combined with integrated transcriptome and physiological analyses of ScCBL6-OXP, we inferred that ScCBL6 improves plant cold stress tolerance in Arabidopsis via the regulation of photosynthesis, redox status, and tonoplast metabolite transporters.

Cold adaptation strategies in plants-An emerging role of epigenetics and antifreeze proteins to engineer cold resilient plants.

Cold stress adversely affects plant growth, development, and yield. Also, the spatial and geographical distribution of plant species is influenced by low temperatures. Cold stress includes chilling and/or freezing temperatures, which trigger entirely different plant responses. Freezing tolerance is acquired via the cold acclimation process, which involves prior exposure to non-lethal low temperatures followed by profound alterations in cell membrane rigidity, transcriptome, compatible solutes, pigments and cold-responsive proteins such as antifreeze proteins. Moreover, epigenetic mechanisms such as DNA methylation, histone modifications, chromatin dynamics and small non-coding RNAs play a crucial role in cold stress adaptation. Here, we provide a recent update on cold-induced signaling and regulatory mechanisms. Emphasis is given to the role of epigenetic mechanisms and antifreeze proteins in imparting cold stress tolerance in plants. Lastly, we discuss genetic manipulation strategies to improve cold tolerance and develop cold-resistant plants.

Mechanistic Insights Into Trehalose-Mediated Cold Stress Tolerance in Rapeseed (Brassica napus L.) Seedlings.

Cold stress (CS) severely affects several physiological, biochemical, and molecular mechanisms and limits the growth and production of rapeseed (Brassica napus L.). Trehalose (Tre) acts as a growth modulator, which is extensively used to improve the tolerance to multiple plant stresses. Further, Tre also serves as an external force in inducing plant signaling molecules, regulating the expression of stress-responsive genes, and enhancing the CS tolerance in plants. Nevertheless, the importance of exogenous Tre in improving the CS tolerance in rapeseed is still unclear. Therefore, the current study was designed to get mechanistic insights into Tre-mediated CS tolerance in rapeseed seedlings. To explore the Tre role, we designed four treatments [control (CK), CK + 20 mM L–1 Tre, Cold, and Cold + 20 mM L–1 Tre] and three CS conditions (4, 0, and −4°C). The results showed that Tre treatments significantly mitigated the adverse effects of CS on the seedlings and increased the survival rate of Tre-treated seedlings under CS conditions. The exogenous Tre dramatically increased the contents of osmoprotectants, including the soluble sugar (SS), soluble protein (SP), and proline (Pro), and the activities of antioxidant enzymes, such as catalase (CAT), peroxidase (POD), superoxide dismutase (SOD), and ascorbate peroxidase (APX) were also increased under CS conditions. Additionally, Tre decreased the malondialdehyde (MDA) contents to protect the rapeseed seedlings. Moreover, Tre also remarkably augmented the expression levels of antioxidant genes (CAT12, POD34, and FSD7), CS-responsive marker genes (CBF1, CBF2, CBF4, COR6.6, COR15, COR25, COL1, and KIN1), and Tre-biosynthesis genes (TPS4, TPS8, and TPS9). Briefly, exogenous Tre not only regulates the antioxidant and osmotic balance, but it also significantly participates in Tre metabolism and signaling network to improve the CS tolerance in rapeseed. Thus, Tre-induced supervisory connections between physiological or/and biochemical attributes provide information to dissect the mechanisms of Tre-mediated CS tolerance.

Role of the Rice BAHD Acyltransferase Gene OsAt10 in Plant Cold Stress Tolerance

Role of the Rice BAHD Acyltransferase Gene OsAt10 in Plant Cold Stress Tolerance

Cold plasma seed treatment improves chilling resistance of tomato plants through hydrogen peroxide and abscisic acid signaling pathway

How to develop a simple and economic approach to improve plant cold stress tolerance is an important scientific problem. With the hope that we explored the effect and metabolism of cold plasma (CP) seed treatment on the chilling tolerance in tomato plants. 75 W CP seed treatment showed the best mitigative effect on cold-induced injury of tomato seedlings, as evidenced by the higher maximum photochemical efficiency of PSII (Fv/Fm), lower ion leakage and chilling injury index. Moreover, the results showed that CP-induced chilling tolerance was related to the hydrogen peroxide (H2O2) mediated by respiratory burst oxidase homologue 1 (RBOH1), which was proved by the decrease low temperature tolerance observed in RBOH1 silence or chemical scavenging of H2O2 seedlings. Furthermore, RBOH1-mediated H2O2 acted as the downstream signaling of CP treatment to enhance the levels of abscisic acid (ABA) by increasing the transcript of 9-cis-epoxycarotenoid dioxygenase 1 (NCED1). Mutation of NCED1 completely abolished CP-induced cold resistance. Genetic evidence showed that H2O2 and ABA were positive regulators of cold stress tolerance. Thus, CP-induced H2O2 and ABA cascade signal up-regulated the regulatory genes (ICE1 and CBF1) of cold acclimation, which increased the osmotic adjustment substances (proline and soluble sugar) accumulation and antioxidant enzymes (SOD, APX and CAT) activities. Our results indicate that H2O2 and ABA signals are involved in conferring cold stress tolerance induced by CP seed treatment in tomato plants.

Study on the mechanism of exogenous serotonin improving cold tolerance of rapeseed (Brassica napus L.) seedlings

Cold stress disturbs numerous physiological and biochemical processes, restricting plant growth and productivity. In the current study, the protective role of exogenous serotonin in alleviating cold stress was investigated in rapeseed (Brassica napus L.) seedlings. The rapeseed seedlings were pretreated with different concentrations (0, 0.01, 0.02, 0.03, and 0.04 g L− 1) of serotonin solution and then were exposed to cold stress (4, 2, 0, and − 2 °C). The results indicated that the pretreatment of serotonin significantly increased the survival rate. Mainly, 0.03 g L− 1 of serotonin increased the survival rate by 75% compared to control conditions. The physiological and biochemical indexes and the expression of cold tolerance-related genes were analyzed in the seedlings pretreated with 0.03 g L− 1 of serotonin. The contents of proline (PRO), soluble sugar (SS), and soluble protein (SP); and the activities of antioxidant defense such as catalase (CAT), peroxidase (POD), and superoxide dismutase (SOD) were significantly increased by exogenous serotonin under cold stress. Supplemented serotonin significantly increased the expression of SOD, COR6.6, COR15, and CBFs genes under cold stress.Overall, our results indicate that the optimal concentration (0.03 g L− 1) of exogenous serotonin maintained the osmotic potential balance in cells under cold stress by increasing the content of osmotic regulatory substances (SS, SP, and PRO), improving the scavenging ability of reactive oxygen species (ROS), increasing the antioxidant enzyme activities (CAT, POD, and SOD) and the transcriptional level of cold stress-related genes, helping rapeseed seedlings to cope with the cold stress. Therefore, serotonin-induced regulatory interactions between physiological and biochemical processes and the elevated expression of stress-associated genes may be a beneficial technique for cold stress tolerance in plants.

Physiological and Molecular Mechanism Involved in Cold Stress Tolerance in Plants.

Previous studies have reported that low temperature (LT) constrains plant growth and restricts productivity in temperate regions. However, the underlying mechanisms are complex and not well understood. Over the past ten years, research on the process of adaptation and tolerance of plants during cold stress has been carried out. In molecular terms, researchers prioritize research into the field of the ICE-CBF-COR signaling pathway which is believed to be the important key to the cold acclimation process. Inducer of CBF Expression (ICE) is a pioneer of cold acclimation and plays a central role in C-repeat binding (CBF) cold induction. CBFs activate the expression of COR genes via binding to cis-elements in the promoter of COR genes. An ICE-CBF-COR signaling pathway activates the appropriate expression of downstream genes, which encodes osmoregulation substances. In this review, we summarize the recent progress of cold stress tolerance in plants from molecular and physiological perspectives and other factors, such as hormones, light, and circadian clock. Understanding the process of cold stress tolerance and the genes involved in the signaling network for cold stress is essential for improving plants, especially crops.

Role of the Arabidopsis calcineurin B-like protein-interacting protein kinase CIPK21 in plant cold stress tolerance

The Arabidopsis calcineurin B-like protein-interacting protein kinase CIPK21 gene (AtCIPK21) plays important roles in cell metabolism, plant development, and abiotic stress responses. However, the function of the AtCIPK21 gene in cold stress tolerance in plant cells is not fully understood. In this study, we use cell cultures of three plant species including rice (Oryza sativa L.), cotton (Gossypium hirsutum L.), and white pine (Pinus strobus L.), as well as Agrobacterium tumefaciens strain GV3101 harboring pBI-AtCIPK21 to generate transgenic cell lines. After confirmation of integration of the AtCIPK21 gene into the genome by polymerase chain reaction (PCR), southern blotting, and northern blotting analyses, the rice, cotton, and pine AtCIPK21 transgenic cell lines were used to examine cold stress tolerance. The experimental results demonstrated that overexpression of the AtCIPK21 gene enhanced cold stress tolerance of transgenic cells by increasing cell viability and cell growth rate, decreasing lipid peroxidation and ion leakage, increasing the content of polyamines, as well as elevating the activity of antioxidative enzymes. In rice cells, AtCIPK21 increases expression of Ca2+-dependent protein kinase (CPK) genes and mitogen-activated protein kinase (MAPK) genes under cold stress. These results indicated that overexpression of the AtCIPK21 gene in plant cells improved cold stress tolerance by elevating polyamines content, antioxidative enzyme activity, and expression of CPK genes and MAPK genes. Overexpression of the AtCIPK21 gene may be a valuable approach for engineering plant cold stress tolerance.

Sesquiterpene glucosylation mediated by glucosyltransferase UGT91Q2 is involved in the modulation of cold stress tolerance in tea plants.

Plants produce and emit terpenes, including sesquiterpenes, during growth and development, which serve different functions in plants. The sesquiterpene nerolidol has health-promoting properties and adds a floral scent to plants. However, the glycosylation mechanism of nerolidol and its biological roles in plants remained unknown. Sesquiterpene UDP-glucosyltransferases were selected by using metabolites-genes correlation analysis, and its roles in response to cold stress were studied. We discovered the first plant UGT (UGT91Q2) in tea plant, whose expression is strongly induced by cold stress and which specifically catalyzes the glucosylation of nerolidol. The accumulation of nerolidol glucoside was consistent with the expression level of UGT91Q2 in response to cold stress, as well as in different tea cultivars. The reactive oxygen species (ROS) scavenging capacity of nerolidol glucoside was significantly higher than that of free nerolidol. Down-regulation of UGT91Q2 resulted in reduced accumulation of nerolidol glucoside, ROS scavenging capacity and tea plant cold tolerance. Tea plants absorbed airborne nerolidol and converted it to its glucoside, subsequently enhancing tea plant cold stress tolerance. Nerolidol plays a role in response to cold stress as well as in triggering plant-plant communication in response to cold stress. Our findings reveal previously unidentified roles of volatiles in response to abiotic stress in plants.

H2O2 and NO are involved in trehalose-regulated oxidative stress tolerance in cold-stressed tomato plants

Cold adversely affects tomato (Solanum lycopersicum) growth and yield during winter and early spring in northern China. Trehalose (Tre) is a growth regulator that is widely used to enhance plant stress tolerance. However, there are few reports on the mechanistic role of Tre in regulating tomato cold tolerance or the interactions between hydrogen peroxide (H2O2) and nitric oxide (NO) in Tre-induced cold stress tolerance in plants. Here, we explored the role of Tre in tomato cold tolerance and evaluated whether H2O2 and NO were involved in this regulation. Tre pretreatment significantly upregulated superoxide dismutase (Cu/Zn SOD) and glutathione reductase (GR1) transcription and increased catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR) activities under normal temperatures. Under cold stress, Tre pretreatment dramatically elevated Cu/Zn SOD and CAT1 transcription and increased SOD and CAT activities. Tre pretreatment reduced membrane lipid peroxidation and alleviated plant growth inhibition caused by cold stress conditions. Under normal temperatures, Tre pretreatment upregulated the expression of respiratory burst oxidase homolog1 (RBOH1), nitrate reductase (NR), and nitric oxide synthase (NOS) and elevated H2O2 and NO levels. Treating tomato leaves with H2O2 and NO scavengers [dimethylthiourea (DMTU) and 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO), respectively] significantly diminished Tre-mediated alleviation of membrane lipid peroxidation. The effect of H2O2 on reducing cold stress-induced damage in tomato was essentially abolished by cPTIO treatment. These results indicated that exogenous Tre enhanced cold stress tolerance in tomato by regulating the antioxidant defense system. Tre may act as an exogenous inducer of H2O2 and NO that regulate antioxidase gene expression and activities to promote plant responses to cold stress, enhance cold stress tolerance, and reduce membrane lipid peroxidation and cellular injury. NO may act downstream of H2O2 to mediate Tre-regulated tomato oxidative stress tolerance under cold stress.

RNA Regulation in Plant Cold Stress Response.

In addition to plants, all organisms react to environmental stimuli via the perception of signals and subsequently respond through alterations of gene expression. However, genes/mRNAs are usually not the functional unit themselves, and instead, resultant protein products with individual functions result in various acquired phenotypes. In order to fully characterize the adaptive responses of plants to environmental stimuli, it is essential to determine the level of proteins, in addition to the regulation of mRNA expression. This regulatory step, which is referred to as "mRNA posttranscriptional regulation," occurs subsequent to mRNA transcription and prior to translation. Although these RNA regulatory mechanisms have been well-studied in many organisms, including plants, it is not fully understood how plants respond to environmental stimuli, such as cold stress, via these RNA regulations.A recent study described several RNA regulatory factors in relation to environmental stress responses, including plant cold stress tolerance. In this chapter, the functions of RNA regulatory factors and comprehensive analyses related to the RNA regulations involved in cold stress response are summarized, such as mRNA maturation, including capping, splicing, polyadenylation of mRNA, and the quality control system of mRNA; mRNA degradation, including the decapping step; and mRNA stabilization. In addition, the putative roles of messenger ribonucleoprotein (mRNP) granules, such as processing bodies (PBs) and stress granules (SGs), which are cytoplasmic particles, are described in relation to RNA regulations under stress conditions. These RNA regulatory systems are important for adjusting or fine-tuning and determining the final levels of mRNAs and proteins in order to adapt or respond to environmental stresses. Collectively, these new areas of study revealed that plants possess precise novel regulatory mechanisms which specifically function in the response to cold stress.

RNA Chaperone Function of a Universal Stress Protein in Arabidopsis Confers Enhanced Cold Stress Tolerance in Plants.

The physiological function of Arabidopsis thaliana universal stress protein (AtUSP) in plant has remained unclear. Thus, we report here the functional role of the Arabidopsis universal stress protein, AtUSP (At3g53990). To determine how AtUSP affects physiological responses towards cold stress, AtUSP overexpression (AtUSP OE) and T-DNA insertion knock-out (atusp, SALK_146059) mutant lines were used. The results indicated that AtUSP OE enhanced plant tolerance to cold stress, whereas atusp did not. AtUSP is localized in the nucleus and cytoplasm, and cold stress significantly affects RNA metabolism such as by misfolding and secondary structure changes of RNA. Therefore, we investigated the relationship of AtUSP with RNA metabolism. We found that AtUSP can bind nucleic acids, including single- and double-stranded DNA and luciferase mRNA. AtUSP also displayed strong nucleic acid-melting activity. We expressed AtUSP in RL211 Escherichia coli, which contains a hairpin-loop RNA structure upstream of chloramphenicol acetyltransferase (CAT), and observed that AtUSP exhibited anti-termination activity that enabled CAT gene expression. AtUSP expression in the cold-sensitive Escherichia coli (E. coli) mutant BX04 complemented the cold sensitivity of the mutant cells. As these properties are typical characteristics of RNA chaperones, we conclude that AtUSP functions as a RNA chaperone under cold-shock conditions. Thus, the enhanced tolerance of AtUSP OE lines to cold stress is mediated by the RNA chaperone function of AtUSP.

An Overview of Signaling Regulons During Cold Stress Tolerance in Plants.

Plants, being sessile organisms, constantly withstand environmental fluctuations, including low-temperature, also referred as cold stress. Whereas cold poses serious challenges at both physiological and developmental levels to plants growing in tropical or sub-tropical regions, plants from temperate climatic regions can withstand chilling or freezing temperatures. Several cold inducible genes have already been isolated and used in transgenic approach to generate cold tolerant plants. The conventional breeding methods and marker assisted selection have helped in developing plant with improved cold tolerance, however, the development of freezing tolerant plants through cold acclimation remains an unaccomplished task. Therefore, it is essential to have a clear understanding of how low temperature sensing strategies and corresponding signal transduction act during cold acclimation process. Herein, we synthesize the available information on the molecular mechanisms underlying cold sensing and signaling with an aim that the summarized literature will help develop efficient strategies to obtain cold tolerant plants.

Beneficial Soil Bacterium Pseudomonas frederiksbergensis OS261 Augments Salt Tolerance and Promotes Red Pepper Plant Growth.

Soil salinity, being a part of natural ecosystems, is an increasing problem in agricultural soils throughout the world. Pseudomonas frederiksbergensis OS261 has already been proved to be an effective bio-inoculant for enhancing cold stress tolerance in plants, however, its effect on salt stress tolerance is unknown. The main aim of the present study was to elucidate P. frederiksbergensis OS261 mediated salt stress tolerance in red pepper. The plants were exposed to a salt stress using NaCl at the concentrations of 50, 100, and 150 mM after 12 days of transplantation, while plant growth and enzyme activity were estimated 50 days after sowing. The height in P. frederiksbergensis OS261 inoculated plants was significantly increased by 19.05, 34.35, 57.25, and 61.07% compared to un-inoculated controls at 0, 50, 100, and 150 mM of NaCl concentrations, respectively, under greenhouse conditions. The dry biomass of the plants increased by 31.97, 37.47, 62.67, and 67.84% under 0, 50, 100, and 150 mM of NaCl concentrations, respectively. A high emission of ethylene was observed in un-inoculated red pepper plants under salinity stress. P. frederiksbergensis OS261 inoculation significantly reduced ethylene emission by 20.03, 18.01, and 20.07% at 50, 100, and 150 mM of NaCl concentrations, respectively. Furthermore, the activity of antioxidant enzymes (ascorbate peroxidase, superoxide dismutase, and catalase) also varied in the inoculated red pepper plants. Salt stress resistance in the bacterized plants was evident from the improved antioxidant activity in leaf tissues and the decreased hydrogen ion concentration. Thus, we conclude that P. frederiksbergensis OS261 possesses stress mitigating property which can enhance plant growth under high soil salinity by reducing the emission of ethylene and regulating antioxidant enzymes.

Breeding approaches and genomics technologies to increase crop yield under low-temperature stress.

Improved knowledge about plant cold stress tolerance offered by modern omics technologies will greatly inform future crop improvement strategies that aim to breed cultivars yielding substantially high under low-temperature conditions. Alarmingly rising temperature extremities present a substantial impediment to the projected target of 70% more food production by 2050. Low-temperature (LT) stress severely constrains crop production worldwide, thereby demanding an urgent yet sustainable solution. Considerable research progress has been achieved on this front. Here, we review the crucial cellular and metabolic alterations in plants that follow LT stress along with the signal transduction and the regulatory network describing the plant cold tolerance. The significance of plant genetic resources to expand the genetic base of breeding programmes with regard to cold tolerance is highlighted. Also, the genetic architecture of cold tolerance trait as elucidated by conventional QTL mapping and genome-wide association mapping is described. Further, global expression profiling techniques including RNA-Seq along with diverse omics platforms are briefly discussed to better understand the underlying mechanism and prioritize the candidate gene (s) for downstream applications. These latest additions to breeders' toolbox hold immense potential to support plant breeding schemes that seek development of LT-tolerant cultivars. High-yielding cultivars endowed with greater cold tolerance are urgently required to sustain the crop yield under conditions severely challenged by low-temperature.

Plastid ribosomal protein S5 is involved in photosynthesis, plant development, and cold stress tolerance in Arabidopsis.

Plastid ribosomal proteins are essential components of protein synthesis machinery and have diverse roles in plant growth and development. Mutations in plastid ribosomal proteins lead to a range of developmental phenotypes in plants. However, how they regulate these processes is not fully understood, and the functions of some individual plastid ribosomal proteins remain unknown. To identify genes responsible for chloroplast development, we isolated and characterized a mutant that exhibited pale yellow inner leaves with a reduced growth rate in Arabidopsis. The mutant (rps5) contained a missense mutation of plastid ribosomal protein S5 (RPS5), which caused a dramatically reduced abundance of chloroplast 16S rRNA and seriously impaired 16S rRNA processing to affect ribosome function and plastid translation. Comparative proteomic analysis revealed that the rps5 mutation suppressed the expression of a large number of core components involved in photosystems I and II as well as many plastid ribosomal proteins. Unexpectedly, a number of proteins associated with cold stress responses were greatly decreased in rps5, and overexpression of the plastid RPS5 improved plant cold stress tolerance. Our results indicate that RPS5 is an important constituent of the plastid 30S subunit and affects proteins involved in photosynthesis and cold stress responses to mediate plant growth and development.

Efficient purification of cryoprotective dehydrin protein from the radish (Raphanus sativus) taproot

Dehydrin is a protein that is related to cold stress tolerance in plants. Because dehydrin shows potent cryoprotective activity, it has the potential to be used in food storage applications. In this paper, we presented an efficient purification method for native dehydrin from radishes (Raphanus sativus). Immunoblot analysis using an anti-Arabidopsis KS-type dehydrin antibody revealed that the related dehydrin accumulates in the radish taproot. The radish dehydrin that accumulated in the vascular tissues of the taproot was purified through two simple chromatographic steps: immobilized metal affinity chromatography followed by anion exchange chromatography. The yield was higher than yields previously reported on a fresh weight basis. The cryoprotective activity for malate dehydrogenase shown by purified dehydrin was more potent than that shown by small molecule cryoprotectants. This suggests that the radish is an appropriate source for the production of native dehydrin.

Plastid signals induce ALTERNATIVE OXIDASE expression to enhance the cold stress tolerance in Arabidopsis thaliana

The external environment affects the metabolic activities of plants, and plastids play an important role in sensing the environmental situation and executing adaptive responses to environmental stresses. Previous studies have indicated that some plastid-signaling deficient mutants have impaired basal thermo-tolerance, drought tolerance, and high-light tolerance. In this study, we investigated the role of plastid signals in alleviating cold stress damage in Arabidopsis (Arabidopsis thaliana). Thermographic analysis revealed that wide-type (Col-0) plants maintained a higher temperature at about 22 °C, whereas the leaf temperature of plastid-signaling deficient mutants was about 18 °C upon 4 °C cold stress. The relative water content of the mutants significantly decreased after cold treatment, whereas H2O2 content, electrolyte leakage, and malonaldehyde content increased compared with Col-0. The cyanide-resistant respiration rates of the mutants were lower than those of wild-type plants. Quantitative real-time PCR analysis revealed that AOX1a (encoding alternative oxidase, a key enzyme for cyanide-resistant respiration) transcripts in the mutants were much lower than wild-type after cold stress. Moreover, investigations with aox1a mutant further indicated that plastid signals enhance plant cold stress tolerance mainly through the induction of AOX1a.

Molecular characterization of cold stress-related transcription factors, CaEREBP-C1, -C2, -C3, and CaWRKY1A from Capsicum annuum L.

EREBP (ethylene responsive element binding protein) and WRKY (wizz-like transcription factor) are known to play roles in plant tolerance to abiotic stresses, such as low temperatures. Previously, we used cDNA microarrays and Northern blot analysis to determine the mechanisms responsible for those underlying defenses. These analyses led to the identification of CaEREBP-C1, -C2, -C3 and CaWRKY1A genes that encode the ethylene responsive element binding protein and wizz-like transcription factors, respectively, from hot pepper (Capsicum annuum). In that study, we demonstrated that the CaEREBP-C1, -C2, -C3 and CaWRKY1A genes were strongly induced by cold stress. Here, we used Tiplasmid and Agrobecterium-mediated transformation to engineer CaEREBP-C1, -C2, -C3 and CaWRKY1A under control of the CaMV 35S promoter for constitutive expression in transgenic plants. The resultant transgenic plants exhibited significantly increased tolerance to low temperatures. In addition, none of the CaEREBP-C1, -C2, -C3 and CaWRKY1A transgenic plants showed visible phenotypic alteration when compared to wild type plants. Taken together, these results suggest that CaEREBP-C1, -C2, -C3 and CaWRKY1A play important biological roles in conferring plant cold stress tolerance.

683 PB 221 IDENTIFICATION OF MOLECULAR MARKERS LINKED TO LOW-TEMPERATURE RESPONSES

Identification of the genes involved in freezing tolerance in oilseed Erussica could lead to genetic improvement of winter survival of this crop and other species, as well as provide greater understanding of the basis of cold stress tolerance in plants. We developed a genetic linkage map for B. rapa using restriction fragment length polymorphisms (RFLPs) and identified molecular markers which are linked to genes controlling vernalization requirement and freezing tolerance. We mapped the location of a group of cold-regulated (`cor') genes from Arabidopsis thaliana in this population and determined their association with freezing tolerance and vernalization requirement. We developed genetically fixed, recombinant inbred lines of B. rapa to assay the physiological processes involved in these cold responses. Specifically, we measured the differences in lipid composition of the plasma membranes of acclimated and nonacclimated plants of a subset of this population. We will determine if the genes involved in the physiological responses to cold temperature are also associated with the acquisition of freezing tolerance.