Cold-regulated Genes Research Articles

Comparative analysis of the variable 3′ UTR and gene expression of the KIN and KIN-homologous LEA genes in Capsella bursa-pastoris

As the crucial members of the cold-regulated (COR) gene family, KIN genes are involved in diverse abiotic stress responses in plants. In the present study, KIN genes from the widespread plant Capsella bursa-pastoris were identified and analyzed to better understand the powerful adaptation of this species. Two KIN genes were cloned and sequenced by 3' RACE. As some COR genes are homologous to LEA genes, three KIN-homologous LEA genes were also identified. We deduced the amino acid sequences of the five proteins to estimate their phylogenetic relationships, and grouped them into three subfamilies (CI, CII, and CIII). Variable 3' UTRs were found in CI, CII, and CIII genes. Using qPCR, we evaluated the transcriptional levels of the five genes in different organs and embryonic stages. Two CI genes were exclusively expressed in early embryos and flowers. The CII and CIII genes showed obvious up-regulation in young leaves after heat stress, cold stress, and ABA treatment. Two of the CI genes, however, rarely responded to those stresses in young leaves. In contrast, all five genes showed differential responses in flowers when C. bursa-pastoris plants were sprayed with ABA. Furthermore, the expression of these genes in C. bursa-pastoris was compared to that of the corresponding Arabidopsis genes, and similar gene expression profiles were found in both species. Our findings suggest that these five genes play different roles in development and the responses to abiotic stresses in C. bursa-pastoris. Key message We characterized two KIN and three KIN-homologous LEA genes, and analyzed their variable 3'UTR and organ-specific, embryo-developmental, stress-induced gene expression in Capsella bursa-pastoris.

Modulation of gene expression in cold-induced sweetening resistant potato species Solanum berthaultii exposed to low temperature

Cold-induced sweetening (CIS) is a crucial factor influencing the processing quality of potato tubers. To better understand the molecular events of potato CIS and different CIS-sensitivity among various potato species, a suppression subtractive hybridization library and cDNA microarray gene filters were developed. A total of 188 genes were found to be differentially expressed (DE) in Solanum berthaultii (ber) upon cold stimulation. These functional genes were mostly related to cell rescue, defense and virulence, metabolism, energy and protein fate, included in various processes of plant defense against abiotic stresses. Four expression patterns of these DE genes were profiled by qRT-PCR using the cold-stored tubers of both CIS-resistant (ber) and CIS-sensitive (E-potato 3, a variety of S. tuberosum) potatoes. The expression pattern and abundance of many DE genes encoding proteins involved in metabolism were different in these two potato tubers, especially genes associated with amylolysis, sucrose decomposition and glycolysis pathways, indicating distinct regulatory mechanisms between ber and E3 in response to cold stress, which may be crucial for potato CIS. Further investigation of these cold-regulated genes will deepen our understanding of the regulatory mechanisms of potato CIS and direct approaches for the genetic improvement of potato processing quality.

Vitis CBF1 and Vitis CBF4 differ in their effect on Arabidopsis abiotic stress tolerance, development and gene expression

Plants growing in temperate regions encode several C-repeat binding factor/dehydration responsive element binding factors (CBF/DREB1) and the question is whether these transcription factors have different functions. In this study, Arabidopsis transformed with grape CBF1 (VrCBF1) or grape CBF4 (VrCBF4) were characterized. Electrolyte leakage assays showed that the freezing tolerance of transgenic lines was correlated with the level of VrCBF expression irrespective of the type of CBF, while drought tolerance was most increased by VrCBF1. VrCBF overexpression coincided with an increase in the expression of the cold-regulated genes AtCOR15a, AtRD29A, AtCOR6.6 and AtCOR47. In addition, the development of grape CBF overexpressing plants was seen to be altered and resulted in dwarf plants which flowered later and had thicker rosette leaves with a higher stomatal density. Analysis of gene expression showed that these morphological changes may be because of an increase in the expression of AtRGL3 in VrCBF4 lines or AtGA2ox7 in VrCBF1 lines, and AtFLC in both. In addition, the results show for the first time that CBFs can positively affect the expression of AtICE1/SCREAM1, the gene that is known to induce AtCBF3 expression. The difference in gene induction by VrCBF1 compared with VrCBF4 suggests that these CBFs have different regulons.

TheArabidopsisNAC Transcription Factor VNI2 Integrates Abscisic Acid Signals into Leaf Senescence via theCOR/RDGenes

Leaf aging is a highly regulated developmental process, which is also influenced profoundly by diverse environmental conditions. Accumulating evidence in recent years supports that plant responsiveness to abiotic stress is intimately related with leaf longevity. However, molecular mechanisms underlying the signaling crosstalks and regulatory schemes are yet unknown. In this work, we demonstrate that an abscisic acid (ABA)-responsive NAC transcription factor VND-INTERACTING2 (VNI2) integrates ABA-mediated abiotic stress signals into leaf aging by regulating a subset of COLD-REGULATED (COR) and RESPONSIVE TO DEHYDRATION (RD) genes. The VNI2 gene was induced by high salinity in an ABA-dependent manner. In addition, spatial and temporal expression patterns of the VNI2 gene are correlated with leaf aging and senescence. Accordingly, leaf aging was delayed in transgenic plants overexpressing the VNI2 gene but significantly accelerated in a VNI2-deficient mutant. The VNI2 transcription factor regulates the COR and RD genes by binding directly to their promoters. Notably, transgenic plants overexpressing the COR or RD genes exhibited prolonged leaf longevity. These observations indicate that the VNI2 transcription factor serves as a molecular link that integrates plant responses to environmental stresses into modulation of leaf longevity.

ICE1 Ser403 is necessary for protein stabilization and regulation of cold signaling and tolerance

ICE1, a MYC-type transcription factor, has an important role in the induction of CBF3/DREB1A for regulation of cold signaling and tolerance. Here we reveal that serine 403 of ICE1 is involved in regulating the transactivation and stability of the ICE1 protein. Substitution of serine 403 by alanine enhanced the transactivational activity of ICE1 in Arabidopsis protoplasts. Over-expression of ICE1(S403A) conferred more freezing tolerance than ICE1(WT) in Arabidopsis, and the expression of cold-regulated genes such as CBF3/DREB1A, COR47 and KIN1 was enhanced in plants over-expressing ICE1(S403A). Furthermore, the ICE1(S403A) protein level was not changed after cold treatment, whereas the ICE1(WT) protein level was reduced. Interestingly, polyubiquitylation of the ICE1(S403A) protein in vivo was apparently blocked. These results demonstrate that serine 403 of ICE1 has roles in both transactivation and cold-induced degradation of ICE1 via the ubiquitin/26S proteasome pathway, suggesting that serine 403 is a key residue for the attenuation of cold-stress responses by HOS1-mediated degradation of ICE1.

A comparison of the low temperature transcriptomes and CBF regulons of three plant species that differ in freezing tolerance: Solanum commersonii, Solanum tuberosum, and Arabidopsis thaliana

Solanum commersonii and Solanum tuberosum are closely related plant species that differ in their abilities to cold acclimate; whereas S. commersonii increases in freezing tolerance in response to low temperature, S. tuberosum does not. In Arabidopsis thaliana, cold-regulated genes have been shown to contribute to freezing tolerance, including those that comprise the CBF regulon, genes that are controlled by the CBF transcription factors. The low temperature transcriptomes and CBF regulons of S. commersonii and S. tuberosum were therefore compared to determine whether there might be differences that contribute to their differences in ability to cold acclimate. The results indicated that both plants alter gene expression in response to low temperature to similar degrees with similar kinetics and that both plants have CBF regulons composed of hundreds of genes. However, there were considerable differences in the sets of genes that comprised the low temperature transcriptomes and CBF regulons of the two species. Thus differences in cold regulatory programmes may contribute to the differences in freezing tolerance of these two species. However, 53 groups of putative orthologous genes that are cold-regulated in S. commersonii, S. tuberosum, and A. thaliana were identified. Given that the evolutionary distance between the two Solanum species and A. thaliana is 112–156 million years, it seems likely that these conserved cold-regulated genes—many of which encode transcription factors and proteins of unknown function—have fundamental roles in plant growth and development at low temperature.

Identification of Cold-Temperature-Regulated Genes in Flavobacterium psychrophilum

Flavobacterium psychrophilum is the etiological agent of bacterial coldwater disease (BCWD) and rainbow trout fry syndrome (RTFS). It causes disease primarily in fresh water-reared salmonids, but other fish species can also be affected. A diverse array of clinical conditions is associated with BCWD, including tail rot (peduncle disease), necrotic myositis, and cephalic osteochondritis. Degradation of connective and muscular tissues by extracellular proteases is common to all of these presentations. There are no effective vaccines to prevent BCWD or RTFS, and antibiotics are often used to prevent and control disease. To identify virulence factors that might permit development of an efficacious vaccine, cDNA suppression subtractive hybridization (SSH) was used to identify cold-regulated genes in a virulent strain of F. psychrophilum. Genes predicted to encode a two-component system sensor histidine kinase (LytS), an ATP-dependent RNA helicase, a multidrug ABC transporter permease/ATPase, an outer membrane protein/protective antigen OMA87, an M43 cytophagalysin zinc-dependent metalloprotease, a hypothetical protein, and four housekeeping genes were upregulated at 8°C versus the level of expression at 20°C. Because no F. psychrophilum gene was known to be suitable as an internal standard in reverse transcription-quantitative real-time PCR (RT-qPCR) experiments, the expression stability of nine commonly used reference genes was evaluated at 8°C and 20°C. Expression of the 16S rRNA was equivalent at both temperatures, and this gene was used in RT-qPCR experiments to verify the SSH findings. With the exception of the ATCC 49513 strain, similar patterns of gene expression were obtained with 11 other representative strains of F. psychrophilum.

A novel cold-regulated gene, COR25, of Brassica napus is involved in plant response and tolerance to cold stress

Cold stress, which causes dehydration damage to the plant cell, is one of the most common abiotic stresses that adversely affect plant growth and crop productivity. To improve its cold-tolerance, plants often enhance expression of some cold-related genes. In this study, a cold-regulated gene encoding 25 KDa of protein was isolated from Brassica napus cDNA library using a macroarray analysis, and is consequently designated as BnCOR25. RT-PCR analysis demonstrated that BnCOR25 was expressed at high levels in hypocotyls, cotyledons, stems, and flowers, but its mRNA was found at low levels in roots and leaves. Northern blot analysis revealed that BnCOR25 transcripts were significantly induced by cold and osmotic stress treatment. The data also showed that BnCOR25 gene expression is mediated by ABA-dependent pathway. Overexpression of BnCOR25 in yeast (Schizosaccharomyces pombe) significantly enhanced the cell survival probability under cold stress, and overexpression of BnCOR25 in Arabidopsis enhances plant tolerance to cold stress. These results suggested that the BnCOR25 gene may play an important role in conferring freezing/cold tolerance in plants.

Nitric oxide participates in cold‐responsive phosphosphingolipid formation and gene expression in Arabidopsis thaliana

Chilling triggers rapid molecular responses that permit the maintenance of plant cell homeostasis and plant adaptation. Recent data showed that nitric oxide (NO) is involved in plant acclimation and tolerance to cold. The participation of NO in the early transduction of the cold signal in Arabidopsis thaliana was investigated. The production of NO after a short exposure to cold was assessed using the NO-sensitive fluorescent probe 4, 5-diamino fluoresceine diacetate and chemiluminescence. Pharmacological and genetic approaches were used to analyze NO sources and NO-mediated changes in cold-regulated gene expression, phosphatidic acid (PtdOH) synthesis and sphingolipid phosphorylation. NO production was detected after 1-4h of chilling. It was impaired in the nia1nia2 nitrate reductase mutant. Moreover, NO accumulation was not observed in H7 plants overexpressing the A. thaliana nonsymbiotic hemoglobin Arabidopsis haemoglobin 1 (AHb1). Cold-regulated gene expression was affected in nia1nia2 and H7 plants. The synthesis of PtdOH upon chilling was not modified by NO depletion. By contrast, the formation of phytosphingosine phosphate and ceramide phosphate, two phosphorylated sphingolipids that are transiently synthesized upon chilling, was negatively regulated by NO. Taken together, these data suggest a new function for NO as an intermediate in gene regulation and lipid-based signaling during cold transduction.

Systemic low temperature signaling in Arabidopsis

When leaves are exposed to low temperature, sugars accumulate and transcription factors in the C-repeat binding factor (CBF) family are expressed, which, together with CBF-independent pathways, are known to contribute to the cold acclimation process and an increase in freezing tolerance. What is not known, however, is whether expression of these cold-regulated genes can be induced systemically in response to a localized cold treatment. To address this, pre-existing, mature leaves of warm-grown Arabidopsis thaliana were exposed to a localized cold treatment (near 10 °C) whilst conjoined newly developing leaves continued only to experience warmer temperatures. In initial experiments on wild-type A. thaliana (Col-0) using real-time reverse transcription--PCR (RT-PCR) we observed that some genes--including CBF genes, certain downstream cold-responsive (COR) targets and CBF-independent transcription factors--respond to a direct 9 °C treatment of whole plants. In subsequent experiments, we found that the treatment of expanded leaves with temperatures near 10 °C can induce cold-associated genes in conjoined warm-maintained tissues. CBF1 showed a particularly strong systemic response, although CBF-independent transcription factors also responded. Moreover, the localized cold treatment of A. thaliana (C24) plants with a luciferase reporter fused to the promoter region of KIN2 indicated that in warm-maintained leaves, KIN2 might respond to a systemic signal from remote, directly cold-treated leaves. Collectively, our study provides strong evidence that the processes involved in cold acclimation are partially mediated by a signal that acts systemically. This has the potential to act as an early-warning system to enable developing leaves to cope better with the cold environment in which they are growing.

Histone dynamics and roles of histone acetyltransferases during cold-induced gene regulation in Arabidopsis

In Arabidopsis, CBF transcription factors bind to and activate certain cold-regulated (COR) gene promoters during cold acclimation. Consistent with the prevailing model that histone acetylation and nucleosomal depletion correspond with transcriptionally active genes, we now report that H3 acetylation increases and nucleosome occupancy decreases at COR gene promoters upon cold acclimation. Overexpression of CBF1 resulted in a constitutive increase in H3 acetylation and decrease in nucleosome occupancy, consistent with the constitutive activation of COR gene expression. Overexpression of a truncated form of CBF2 lacking its transcriptional activation domain resulted in a cold-stimulated increase in H3 acetylation, but no change in nucleosomal occupancy or COR gene expression, indicating that histone acetylation is congruent with but not sufficient for cold-activation of COR gene expression. Plants homozygous for T-DNA disruption alleles of GCN5 (encoding a histone acetyltransferase) or ADA2b (a GCN5-interacting protein) show diminished expression of COR genes during cold acclimation. Contrary to expectations, H3 acetylation at COR gene promoters was stimulated upon cold acclimation in ada2b and gcn5 plants as in wild type plants, but the decrease in nucleosome occupancy was diminished. Thus, GCN5 is not the HAT responsible for histone acetylation at COR gene promoters during cold acclimation. Several other HAT mutant plants were also tested; although some do affect COR gene expression, none affected histone acetylation. Therefore, H3 acetylation at the COR gene promoters is not solely dependent on any of the HATs tested.

Regulation of Freezing Tolerance and Flowering in Temperate Cereals: The VRN-1 Connection

In winter wheat (Triticum spp.) and barley (Hordeum vulgare) varieties, long exposures to nonfreezing cold temperatures accelerate flowering time (vernalization) and improve freezing tolerance (cold acclimation). However, when plants initiate their reproductive development, freezing tolerance decreases, suggesting a connection between the two processes. To better understand this connection, we used two diploid wheat (Triticum monococcum) mutants, maintained vegetative phase (mvp), that carry deletions encompassing VRN-1, the major vernalization gene in temperate cereals. Homozygous mvp/mvp plants never flower, whereas plants carrying at least one functional VRN-1 copy (Mvp/-) exhibit normal flowering and high transcript levels of VRN-1 under long days. The Mvp/- plants showed reduced freezing tolerance and reduced transcript levels of several cold-induced C-REPEAT BINDING FACTOR transcription factors and COLD REGULATED genes (COR) relative to the mvp/mvp plants. Diploid wheat accessions with mutations in the VRN-1 promoter, resulting in high transcript levels under both long and short days, showed a significant down-regulation of COR14b under long days but not under short days. Taken together, these studies suggest that VRN-1 is required for the initiation of the regulatory cascade that down-regulates the cold acclimation pathway but that additional genes regulated by long days are required for the down-regulation of the COR genes. In addition, our results show that allelic variation in VRN-1 is sufficient to determine differences in freezing tolerance, suggesting that quantitative trait loci for freezing tolerance previously mapped on this chromosome region are likely a pleiotropic effect of VRN-1 rather than the effect of a separate closely linked locus (FROST RESISTANCE-1), as proposed in early freezing tolerance studies.

Overexpression of SlCZFP1, a Novel TFIIIA-type Zinc Finger Protein from Tomato, Confers Enhanced Cold Tolerance in Transgenic Arabidopsis and Rice

The C2H2-type (also called TFIIIA-type) zinc finger protein family is a major family of eukaryotic transcription factors. C2H2-type zinc finger proteins in plants play important roles in regulating various developmental processes or responses to abiotic stresses. We isolated a complementary deoxyribonucleic acid (cDNA) clone, designated Solanumlycopersicumcold zinc finger protein 1 (SlCZFP1), from tomato by analyzing the previous microarray data of the low-temperature transcriptome of tomato. SlCZFP1 encodes a novel TFIIIA-type zinc finger protein, which contains two typical zinc finger motifs, CX2–4 CX3FX3QALGGHX3–5H, and a potential nuclear localization signal (NLS). The SlCZFP1–GFP fusion protein was localized to the nucleus in a transient expression assay. Expression of SlCZFP1 was strongly induced by cold stress, dehydration, and salt treatment, but not by abscisic acid (ABA). Overexpression of SlCZFP1 in transgenic Arabidopsis and rice induced constitutive expression of cold-regulated (COR) or cold-responsive genes and conferred enhanced tolerance to freezing or cold treatments for non-acclimate transgenic plants, compared with wild-type plants. However, there was no obvious enhancement observed in drought and salt tolerance. Our data suggest that SlCZFP1 plays an important role in plant responses to cold stress by regulating cold-responsive gene expression and that SlCZEP1 might be a useful gene for improving cold tolerance in crop plants.

Using Specialized cDNA Microarrays to Analyze Arabidopsis Gene Expression Under Cold Stress

Cold acclimation enables plants to withstand low but non-freezing temperatures. Biochemical and physiological changes include a reduction in tissue water content and altered composition of membrane lipids. These responses are correlated with fluctuations in the expression of cold-induced genes such as LTI (low-temperature-induced), KIN (cold-inducible), RD (responsive to desiccation), and ERD (early dehydration-inducible). We performed time-course experiments with specialized cDNA microarrays comprising 712 cDNAs selected by SAGE and SSH methods. Expression dynamics were monitored in the leaves of Arabidopsis. Profiles of nine samples from plants chilled for various time periods revealed 264 cold-inducible genes and 33 repressed genes, for which expression was altered (up or down) by at least twofold. These included not only several previously reported cold-regulated genes, e.g., rd, lea, and CBF3, but also candidate genes such as those for alcohol dehydrogenase and transport inhibitor response 1. All genes were grouped according to their expression patterns; most tended to shift their expression at 3 ∼ 8 h after cold treatment. Two cold-associated transcriptional activators, CBF2 and CBF3, did not have parallel patterns of expression, although both were induced within 15 min. Our results suggest different roles for CBF2 and CBF3 in the signal-transduction pathway for cold acclimation. We believe that, compared with standard differential screening, our microarray analysis is a more useful technique for the selection of new candidate genes responsible for cold acclimation.

A novel cold-regulated gene from Camellia sinensis, CsCOR1, enhances salt- and dehydration-tolerance in tobacco

In present research, the full-length cDNA and the genomic sequence of a novel cold-regulated gene, CsCOR1, were isolated from Camellia sinensis L. The deduced protein CsCOR1 contains a hydrophobic N-terminus as a signal peptide and a hydrophilic C-terminal domain that is rich in glycine, arginine and proline. Two internal repetitive tridecapeptide fragments (HSVTAGRGGYNRG) exist in the middle of the C-terminal domain and the two nucleotide sequences encoding them are identical. CsCOR1 was localized in the cell walls of transgenic-tobaccos via CsCOR1:: GFP fusion approach. The expression of CsCOR1 in tea leaves was enhanced dramatically by both cold- and dehydration-stress. And overexpression of CsCOR1 in transgenic-tobaccos improved obviously the tolerance to salinity and dehydration.

Application of functional genomics and proteomics to plant cryopreservation.

Plant cryobiology has primarily emerged from the classical fields of cryobiology and plant stress physiology. Cryopreservation tools are now available to geneticists for germplasm preservation and the field itself is advancing significantly through the use of molecular techniques. Long-term preservation of vegetatively propagated tissues can minimize the risks of long-term maintenance under tissue culture or field conditions. Cells can be successfully cryopreserved when the adverse affects of ice crystal formation are mitigated by the removal of water or procedures to limit ice formation and crystal growth. The addition of cryoprotectant solutions to hydrated cells may improve the survival of microdissected shoot tips or embryonic axes. Recent discoveries in the genetic pathways leading to cold acclimation and freezing tolerance suggest the involvement of key cold-regulated genes in the acquisition of cold tolerance in plant tissues. Model systems of banana and Arabidopsis have revealed the involvement of genes and proteins in the glycolytic and other metabolic pathways, particularly processes involved in dehydration tolerance, osmoprotection, and membrane transport. Furthermore, successful recovery appears to be dependent upon the presence of antioxidant protection from reactive oxygen species. Characterization of specific genes and proteins will lead to significant advances in plant cryobiology research.

Depletion of the Membrane-Associated Acyl-Coenzyme A-Binding Protein ACBP1 Enhances the Ability of Cold Acclimation in Arabidopsis

In Arabidopsis (Arabidopsis thaliana), a family of six genes encodes acyl-coenzyme A-binding proteins (ACBPs). A member of this family, ACBP1, contains an amino-terminal transmembrane domain that targets it to the plasma membrane and the endoplasmic reticulum. To investigate ACBP1 function, ACBP1-overexpressing transgenic Arabidopsis plants were characterized using lipid analysis. ACBP1 overexpressors showed reduction in several species of diunsaturated phosphatidylcholine (PC), prompting us to investigate if they were altered in response to freezing stress. ACBP1 overexpressors demonstrated increased freezing sensitivity accompanied by a decrease in PC and an increase in phosphatidic acid (PA), while acbp1 mutant plants showed enhanced freezing tolerance associated with PC accumulation and PA reduction. We also showed binding of a recombinant eukaryotic ACBP (ACBP1) to PA, indicative of the possibility of enhanced PA interaction in ACBP1 overexpressors. Since phospholipase Dalpha1 (PLDalpha1) is a major enzyme promoting the hydrolysis of PC to PA, PLDalpha1 expression was examined and was observed to be higher in ACBP1 overexpressors than in acbp1 mutant plants. In contrast, the expression of PLDdelta, which plays a positive role in freezing tolerance, declined in the ACBP1 overexpressors but increased in acbp1 mutant plants. Given that ACBP1 is localized to the endoplasmic reticulum and plasma membrane, it may regulate the expression of PLDalpha1 and PLDdelta by maintaining a membrane-associated PA pool through its ability to bind PA. Moreover, both genotypes showed no alterations in proline and soluble sugar content or in cold-regulated (COR6.6 and COR47) gene expression, suggesting that the ACBP1-mediated response is PLD associated and is independent of osmolyte accumulation.

What happens in plant molecular responses to cold stress?

Low temperature is one of the major abiotic stresses limiting the productivity and the geographical distribution of many important crops. Many plants increase in freezing tolerance in response to low temperatures. This phenomenon needs a vast reprogramming of gene expression which results in the adjusted metabolic-structural alterations. However, the efficient adjustments are dependent on proper cold signal transduction. The first stage is cold stress signal perception which is carried out by different pathways. Transcriptional cascades are next players which operate through ABA-dependent and ABA-independent pathways to induce cold-regulated (COR) gene expression and the result is increasing in the levels of hundreds of metabolites, which some of them are known to have protective effects against the damaging effects of cold stress and some like soluble sugars, reactive oxygen species and photosynthetic metabolites are thought to act as signaling molecules and regulate special COR genes. The different aspects of these events are discussed in detail below.

How do bacteria sense and respond to low temperature?

Rigidification of the membrane appears to be the primary signal perceived by a bacterium when exposed to low temperature. The perception and transduction of the signal then occurs through a two-component signal transduction pathway consisting of a membrane-associated sensor and a cytoplasmic response regulator and as a consequence a set of cold-regulated genes are activated. In addition, changes in DNA topology due to change in temperature may also trigger cold-responsive mechanisms. Inducible proteins thus accumulated repair the damage caused by cold stress. For example, the fluidity of the rigidified membrane is restored by altering the levels of saturated and unsaturated fatty acids, by altering the fatty acid chain length, by changing the proportion of cis to trans fatty acids and by changing the proportion of anteiso to iso fatty acids. Bacteria could also achieve membrane fluidity changes by altering the protein content of the membrane and by altering the levels of the type of carotenoids synthesized. Changes in RNA secondary structure, changes in translation and alteration in protein conformation could also act as temperature sensors. This review highlights the various strategies by which bacteria senses low temperature signal and as to how it responds to the change.

Functional Identification of a C-repeat Binding Factor Transcriptional Activator from Blueberry Associated with Cold Acclimation and Freezing Tolerance

Highbush blueberry (Vaccinium corymbosum) is susceptible to winter freezing injury and frost damage in the spring. As part of an ongoing project to understand the process of cold acclimation, we isolated a C-repeat binding factor (CBF) transcriptional activator gene-coding region from the highbush blueberry cultivar Bluecrop. Expression of the highbush blueberry CBF gene was compared in floral buds of the cold-tolerant northern highbush cultivar Bluecrop and the more cold-sensitive southern rabbiteye (V. virgatum) blueberry cultivar Tifblue. Relative gene expression was higher in ‘Bluecrop’ than in ‘Tifblue’. Expression in both cultivars was highest at the earliest time point in the fall (coincident with the first stage of cold acclimation), declined during the later fall and winter, and, in ‘Bluecrop’, increased again as buds deacclimated, when temperatures tend to fluctuate. To confirm the putative identity of the gene as a member of the CBF gene family, and to determine if expression in a heterologous system could enhance freezing tolerance, the blueberry gene coding sequence was overexpressed in transgenic Arabidopsis thaliana under the control of the cauliflower mosaic virus 35S promoter. Transgenic plants expressing the putative blueberry CBF gene exhibited induced expression of the A. thaliana cold-regulated (COR) genes COR78 and COR6.6, under non-inducing conditions (i.e., 23 °C); however, expression of two other COR genes was unaffected. Transgenic plants also exhibited enhanced freezing tolerance under non-acclimated conditions, but not to the level of acclimated control plants. Thus, the expression pattern in floral buds and the ability of the isolated gene to turn on a subset of COR genes and increase freezing tolerance in a heterologous system suggest it is a functional member of the CBF gene family in blueberry.