Temperature-responsive Genes Research Articles

SUCROSE NONFERMENTING1-RELATED PROTEIN KINASE2.6, an ortholog of OPEN STOMATA1, is a negative regulator of strawberry fruit development and ripening.

Whereas the regulatory mechanisms that direct fruit ripening have been studied extensively, little is known about the signaling mechanisms underlying this process, especially for nonclimacteric fruits. In this study, we demonstrated that a SUCROSE NONFERMENTING1-RELATED PROTEIN KINASE2, designated as FaSnRK2.6, is a negative regulator of fruit development and ripening in the nonclimacteric fruit strawberry (Fragaria × ananassa) and can also mediate temperature-modulated strawberry fruit ripening. FaSnRK2.6 was identified as an ortholog of OPEN STOMATA1. Levels of FaSnRK2.6 transcript rapidly decreased during strawberry fruit development and ripening. FaSnRK2.6 was found to be capable of physically interacting with strawberry ABSCISIC ACID INSENSITIVE1, a negative regulator in strawberry fruit ripening. RNA interference-induced silencing of FaSnRK2.6 significantly promoted fruit ripening. By contrast, overexpression of FaSnRK2.6 arrested fruit ripening. Strawberry fruit ripening is highly sensitive to temperature, with high temperatures promoting ripening and low temperatures delaying it. As the temperature increased, the level of FaSnRK2.6 expression declined. Furthermore, manipulating the level of FaSnRK2.6 expression altered the expression of a variety of temperature-responsive genes. Taken together, this study demonstrates that FaSnRK2.6 is a negative regulator of strawberry fruit development and ripening and, furthermore, that FaSnRK2.6 mediates temperature-modulated strawberry fruit ripening.

A transcriptomic analysis of Chrysanthemum nankingense provides insights into the basis of low temperature tolerance.

BackgroundA major constraint affecting the quality and productivity of chrysanthemum is the unusual period of low temperature occurring during early spring, late autumn, and winter. Yet, there has been no systematic investigation on the genes underlying the response to low temperature in chrysanthemum. Herein, we used RNA-Seq platform to characterize the transcriptomic response to low temperature by comparing different transcriptome of Chrysanthemum nankingense plants and subjecting them to a period of sub-zero temperature, with or without a prior low temperature acclimation.ResultsSix separate RNA-Seq libraries were generated from the RNA samples of leaves and stems from six different temperature treatments, including one cold acclimation (CA), two freezing treatments without prior CA, two freezing treatments with prior CA and the control. At least seven million clean reads were obtained from each library. Over 77% of the reads could be mapped to sets of C. nankingense unigenes established previously. The differentially transcribed genes (DTGs) were identified as low temperature sensing and signalling genes, transcription factors, functional proteins associated with the abiotic response, and low temperature-responsive genes involved in post-transcriptional regulation. The differential transcription of 15 DTGs was validated using quantitative RT-PCR.ConclusionsThe large number of DTGs identified in this study, confirmed the complexity of the regulatory machinery involved in the processes of low temperature acclimation and low temperature/freezing tolerance.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-844) contains supplementary material, which is available to authorized users.

Physiological and transcriptional responses of anaerobic chemostat cultures of Saccharomyces cerevisiae subjected to diurnal temperature cycles.

Diurnal temperature cycling is an intrinsic characteristic of many exposed microbial ecosystems. However, its influence on yeast physiology and the yeast transcriptome has not been studied in detail. In this study, 24-h sinusoidal temperature cycles, oscillating between 12°C and 30°C, were imposed on anaerobic, glucose-limited chemostat cultures of Saccharomyces cerevisiae. After three diurnal temperature cycles (DTC), concentrations of glucose and extracellular metabolites as well as CO2 production rates showed regular, reproducible circadian rhythms. DTC also led to waves of transcriptional activation and repression, which involved one-sixth of the yeast genome. A substantial fraction of these DTC-responsive genes appeared to respond primarily to changes in the glucose concentration. Elimination of known glucose-responsive genes revealed an overrepresentation of previously identified temperature-responsive genes as well as genes involved in the cell cycle and de novo purine biosynthesis. In-depth analysis demonstrated that DTC led to a partial synchronization of the cell cycle of the yeast populations in chemostat cultures, which was lost upon release from DTC. Comparison of DTC results with data from steady-state cultures showed that the 24-h DTC was sufficiently slow to allow S. cerevisiae chemostat cultures to acclimate their transcriptome and physiology at the DTC temperature maximum and to approach acclimation at the DTC temperature minimum. Furthermore, this comparison and literature data on growth rate-dependent cell cycle phase distribution indicated that cell cycle synchronization was most likely an effect of imposed fluctuations of the relative growth rate (μ/μmax) rather than a direct effect of temperature.

A change in temperature modulates defence to yellow (stripe) rust in wheat line UC1041 independently of resistance gene Yr36.

BackgroundRust diseases are of major importance in wheat production worldwide. With the constant evolution of new rust strains and their adaptation to higher temperatures, consistent and durable disease resistance is a key challenge. Environmental conditions affect resistance gene performance, but the basis for this is poorly understood.ResultsHere we show that a change in day temperature affects wheat resistance to Puccinia striiformis f. sp tritici (Pst), the causal agent of yellow (or stripe) rust. Using adult plants of near-isogenic lines UC1041 +/- Yr36, there was no significant difference between Pst percentage uredia coverage in plants grown at day temperatures of 18°C or 25°C in adult UC1041 + Yr36 plants. However, when plants were transferred to the lower day temperature at the time of Pst inoculation, infection increased up to two fold. Interestingly, this response was independent of Yr36, which has previously been reported as a temperature-responsive resistance gene as Pst development in adult UC1041 -Yr36 plants was similarly affected by the plants experiencing a temperature reduction. In addition, UC1041 -Yr36 plants grown at the lower temperature then transferred to the higher temperature were effectively resistant and a temperature change in either direction was shown to affect Pst development up to 8 days prior to inoculation. Results for seedlings were similar, but more variable compared to adult plants. Enhanced resistance to Pst was observed in seedlings of UC1041 and the cultivar Shamrock when transferred to the higher temperature. Resistance was not affected in seedlings of cultivar Solstice by a temperature change in either direction.ConclusionsYr36 is effective at 18°C, refining the lower range of temperature at which resistance against Pst is conferred compared to previous studies. Results reveal previously uncharacterised defence temperature sensitivity in the UC1041 background which is caused by a change in temperature and independently of Yr36. This novel phenotype is present in some cultivars but absent in others, suggesting that Pst defence may be more stable in some cultivars than others when plants are exposed to varying temperatures.

Time-based comparative transcriptomics in engineered xylose-utilizing Saccharomyces cerevisiae identifies temperature-responsive genes during ethanol production

Agricultural residues comprising lignocellulosic materials are excellent sources of pentose sugar, which can be converted to ethanol as fuel. Ethanol production via consolidated bioprocessing requires a suitable microorganism to withstand the harsh fermentation environment of high temperature, high ethanol concentration, and exposure to inhibitors. We genetically enhanced an industrial Saccharomyces cerevisiae strain, sun049, enabling it to uptake xylose as the sole carbon source at high fermentation temperature. This strain was able to produce 13.9 g/l ethanol from 50 g/l xylose at 38 °C. To better understand the xylose consumption ability during long-term, high-temperature conditions, we compared by transcriptomics two fermentation conditions: high temperature (38 °C) and control temperature (30 °C) during the first 12 h of fermentation. This is the first long-term, time-based transcriptomics approach, and it allowed us to discover the role of heat-responsive genes when xylose is the sole carbon source. The results suggest that genes related to amino acid, cell wall, and ribosomal protein synthesis are down-regulated under heat stress. To allow cell stability and continuous xylose uptake in order to produce ethanol, hexose transporter HXT5, heat shock proteins, ubiquitin proteins, and proteolysis were all induced at high temperature. We also speculate that the strong relationship between high temperature and increased xylitol accumulation represents the cell's mechanism to protect itself from heat degradation.

A Temperature-Responsive Network Links Cell Shape and Virulence Traits in a Primary Fungal Pathogen

Author SummaryMicrobial pathogens of humans display the ability to thrive at host temperature. So-called “thermally dimorphic” fungal pathogens, which include Histoplasma capsulatum, are a class of soil fungi that upon being inhaled into the human lung, undergo dramatic changes in cell shape and virulence gene expression in response to host temperature. The ability of these pathogens to cause disease is exquisitely coupled to temperature response. Here we elucidate the regulatory network that governs the ability of H. capsulatum to switch from a filamentous form in the soil environment to a pathogenic yeast form at body temperature. The circuit is driven by three transcription regulators (Ryp1, Ryp2, and Ryp3) that control yeast-phase growth. We show that these factors, which include two highly conserved proteins of the Velvet family of unknown function, bind to specific regulatory DNA elements and directly regulate expression of virulence genes. We identify and characterize Ryp4, a fourth regulator of this pathway, and define DNA motifs that recruit these transcription factors to their temperature-responsive target genes. Our results provide a molecular understanding of how changes in cell shape are linked to expression of virulence genes in thermally dimorphic fungi.

Transcriptomic analysis of acclimation to temperature and light stress in Saccharina latissima (Phaeophyceae).

Kelps, brown algae of the order Laminariales, dominate rocky shores and form huge kelp beds which provide habitat and nurseries for various marine organisms. Whereas the basic physiological and ecophysiological characteristics of kelps are well studied, the molecular processes underlying acclimation to different light and temperature conditions are still poorly understood. Therefore we investigated the molecular mechanisms underlying the physiological acclimation to light and temperature stress. Sporophytes of S. latissima were exposed to combinations of light intensities and temperatures, and microarray hybridizations were performed to determine changes in gene expression patterns. This first large-scale transcriptomic study of a kelp species shows that S. latissima responds to temperature and light stress with a multitude of transcriptional changes: up to 32% of genes showed an altered expression after the exposure experiments. High temperature had stronger effects on gene expression in S. latissima than low temperature, reflected by the higher number of temperature-responsive genes. We gained insights into underlying molecular processes of acclimation, which includes adjustment of the primary metabolism as well as induction of several ROS scavengers and a sophisticated regulation of Hsps. We show that S. latissima, as a cold adapted species, must make stronger efforts for acclimating to high than to low temperatures. The strongest response was caused by the combination of high temperatures with high light intensities, which proved most harmful for the alga.

Transgenic tobacco plants over expressing cold regulated protein CbCOR15b from Capsella bursa-pastoris exhibit enhanced cold tolerance

Low temperature is among the most significant abiotic stresses, restricting the habitats of sessile plants and reducing crop productivity. Cold regulated (COR) genes are low temperature-responsive genes expressing under regulation of a specific signal transduction pathway, which is designated C-repeat-binding-factor (CBF) signaling pathway. In the present article, cold bioassay showed that the transcript level of cold regulated gene CbCOR15b from shepherd's purse (Capsella bursa-pastoris) was obviously elevated under cold treatments. Reverse transcription-PCR (RT-PCR) and GUS report system revealed that unlike AtCOR15b, CbCOR15b expressed not only in leaves but also in stems and maturation zone of roots. When transgenic tobacco plants ectopically expressing CbCOR15b were exposed to chilling and freezing temperatures, they displayed more cold tolerance compared to control plants. According to the electrolyte leakage, the relative water content, the glucose content and the phenotype observation, CbCOR15b transformants suffered less damage under cold stress. Further investigation of the subcellular localization of CbCOR15b by transient expression of fusion protein CbCOR15b–GFP revealed that it was localized exclusively in the chloroplasts of tobacco mesophyll cells and in the cytoplasm of onion epidermal cells. It can be concluded that CbCOR15b which located in the chloroplasts and in the cytoplasm of cells without chloroplasts was involved in cold response of C. bursa-pastoris and conferred enhanced cold tolerance in transgenic tobacco plants.

Thermogenetic tools to monitor temperature-dependent gene expression in bacteria

Free-living bacteria constantly monitor their ambient temperature. Drastic deviations elicit immediate protective responses known as cold shock or heat shock response. Many mammalian pathogens use temperature surveillance systems to recognize the successful invasion of a host by its body temperature, usually 37°C. Translation of temperature-responsive genes can be modulated by RNA thermometers (RNATs). RNATs form complex structures primarily in the 5'-untranslated region of their transcripts. Most RNATs block the ribosome binding site at low temperatures. Translation is induced at increasing temperature by melting of the RNA structure. The analysis of such temperature-dependent RNA elements calls for adequate test systems that function in the appropriate temperature range. Here, we summarize previously established reporter gene systems based on the classical β-galactosidase LacZ, the heat-stable β-galactosidase BgaB and the green fluorescent protein GFP. We validate these systems by testing known RNATs and describe the construction and application of an optimized bgaB system. Finally, two novel RNA thermometer candidates from Escherichia coli and Salmonella will be presented.

Characterization and expression of high temperature stress responsive genes in bread wheat (Triticum aestivum L.)

To elucidate the effects of high temperatures, wheat plants (Triticum aestivum cv. CPAN 1676) were given heat shocks at 37°C and 42°C for two hours, and responsive genes were identified through PCR-Select Subtraction technology. Four subtractive cDNA libraries, including three forward and one reverse subtraction, were constructed from three different developmental stages. A total of 5500 ESTs were generated and 3516 high quality ESTs were submitted to Genbank. More than one third of the ESTs generated fall in unknown/no hit categories upon a homology search through BLAST analysis. A large number of high temperature responsive genes have been identified and characterized. Reverse subtraction analysis in developing grains showed extensive transcriptional changes upon heat stress as revealed by comparative analysis with forward subtraction. Differential expression was confirmed by cDNA macroarray and by northern/RT-PCR analysis. Expression analysis of wheat plants subjected to high temperature stress, after one and four days of recovery, showed fast recovery in seedling tissues. However, recovery was small in the developing seed tissue after two hours of heat stress. Ten selected genes were analysed in further detail by quantitative real-time PCR in an array of 35 different wheat tissues representing major developmental stages as well as different abiotic stresses. Tissue specificity was examined along with cross talk with other abiotic stresses and putative signalling molecules. The results obtained contribute towards understanding the regulation of genes at different developmental stages in wheat crucial to withstanding and recovery from heat stress.

Bioinformatic prediction of the AP2/ERF family genes in Eucalyptus grandis: focus on the CBF family

Background Due to their very high economic importance, Eucalyptus tree species are among the most planted hardwoods in the world with over 20 million hectares. However, as long-lived evergreen species, this genus is particularly exposed to cold. Frost tolerance varies among species and is inversely correlated to productivity. The AP2/ERF gene family includes developmentally and physiologically important transcription factors characterized by the presence of the AP2/ERF DNA-binding domain. AP2 proteins contain two AP2-like domains and RAV family proteins contain one AP2 domain and one B3 domain. ERF family proteins exhibit only one AP2 domain and are further divided into the DREB subfamily and the ERF subfamily [1]. The CBF/DREB1 protein differ from the other DREB proteins by the presence of “signature sequences” (PKK/RPAGRxKFxETRHP and DSAWR) flanking the DNA-binding AP2 domain [2]. The DREB factors recognize the C-repeat or dehydration response element (DRE) in the promoters of low temperature and/or water deficit responsive genes and would play a crucial role in response to abiotic stresses. CBF/DREB1 are the key regulators of the cold-responsive (COR) genes. So far CBF transcription factors have been mainly characterized in model plants such as Arabidopsis, but lately they were identified in several tree species including Eucalyptus [2]. The Eucalyptus cold tolerance was greatly improved in our hands when two genes from the four CBF members isolated from a tolerant species E. gunnii were individually constitutively overexpressed in the frost sensitive E. urophylla x E. grandis hybrid [3]. In the present study E. grandis AP2/ERF family genes were identified based on the presence of putative encoding AP2-domain(s) and were studied with regard to the model herbaceous Arabidopsis as well as the main sequenced woody plants. Within this family, a part of the study focused on the CBF/DREB1 subfamily which was compared to the four genes already characterized in E. gunnii[2].

Identification and expression analysis of CjLTI, a novel low temperature responsive gene from Caragana jubata

Using rapid amplification of cDNA ends, a full length cDNA (CjLTI) was cloned from apical buds of Caragana jubata, a plant species that grows under extreme cold. The cDNA obtained was 573 bp long consisting of an open reading frame of 351 bp encoding 116 amino acids. Homology analysis did not exhibit significant similarity with any sequence at NCBI database, therefore it was deduced as a novel gene. Secondary structure analysis suggested that the deduced CjLTI contained 25.86% α-helices, 4.31% β-turns, 6.90% extended strands, and 62.93% random coils. The hydropathy profile suggested CjLTI to be a hydrophobic protein having characteristic features of signal peptides at N-terminus. The gene exhibited down-regulation at 5 min of exposure to low temperature (LT, 4 ± 3 °C) followed by a strong up-regulation after 15 min and onwards. Methyl jasmonate (MJ) lead to up-regulation of CjLTI starting at 5 min onwards. The gene exhibited up- and down-regulation of expression pattern in response to abscisic acid (ABA) and salicylic acid (SA). Mild drought stress slightly up-regulated gene expression and at severe drought (up to 115% reduction in leaf water potential) slight down-regulation of gene expression was observed. These results suggested CjLTI to be a LT responsive gene wherein MJ, ABA and SA pathways might be involved in regulating the gene expression.

Comparative physiology: a “crystal ball” for predicting consequences of global change

Comparative physiology offers powerful approaches for developing causal, mechanistic explanations of shifts in biogeographic patterning occurring in concert with global change. These analyses can identify the cellular loci and intensities of stress-induced perturbation and generate predictions about ecosystem alterations in a changing world. Congeneric species adapted to different abiotic conditions offer excellent study systems for these purposes. Several findings have emerged from such comparative studies: 1) In aquatic and terrestrial habitats, the most heat-tolerant ectotherms may be most threatened by further increases in temperature, due to proximity of these species' thermal optima and tolerance limits to current maximal ambient temperatures and limited capacities for acclimatization to higher temperatures. 2) Cardiac function is a "weak link" in acute thermal tolerance. 3) Stress-induced changes in gene expression comprise a graded response involving genes linked to damage repair, lysis of irreversibly damaged molecules, and downregulation of cell proliferation. Transcriptomic and proteomic analyses provide "biomarkers" for diagnosing degrees of stress. 4) Different abiotic stresses may have synergistic or opposing effects on gene expression, a complexity needing consideration when developing integrated pictures of effects of global change. 5) Adaptation of proteins can result from one to a few amino acid substitutions, which can occur at many sites in a protein, a discovery with implications for rates of adaptive evolution. 6) Greater thermal tolerance of invasive species may favor their replacement of natives. 7) Losses of protein-coding genes and temperature-responsive gene regulatory abilities in stenothermal ectotherms of the Southern Ocean may lead to broad extinctions.

Ambient temperature response establishes ELF3 as a required component of the core Arabidopsis circadian clock.

Circadian clocks synchronize internal processes with environmental cycles to ensure optimal timing of biological events on daily and seasonal time scales. External light and temperature cues set the core molecular oscillator to local conditions. In Arabidopsis, EARLY FLOWERING 3 (ELF3) is thought to act as an evening-specific repressor of light signals to the clock, thus serving a zeitnehmer function. Circadian rhythms were examined in completely dark-grown, or etiolated, null elf3-1 seedlings, with the clock entrained by thermocycles, to evaluate whether the elf3 mutant phenotype was light-dependent. Circadian rhythms were absent from etiolated elf3-1 seedlings after exposure to temperature cycles, and this mutant failed to exhibit classic indicators of entrainment by temperature cues, consistent with global clock dysfunction or strong perturbation of temperature signaling in this background. Warm temperature pulses failed to elicit acute induction of temperature-responsive genes in elf3-1. In fact, warm temperature-responsive genes remained in a constitutively "ON" state because of clock dysfunction and, therefore, were insensitive to temperature signals in the normal time of day-specific manner. These results show ELF3 is broadly required for circadian clock function regardless of light conditions, where ELF3 activity is needed by the core oscillator to allow progression from day to night during either light or temperature entrainment. Furthermore, robust circadian rhythms appear to be a prerequisite for etiolated seedlings to respond correctly to temperature signals.

Comparative characterization of a temperature responsive gene (lactate dehydrogenase-B, ldh-b) in two congeneric tropical fish, Lates calcarifer and Lates niloticus.

The characterization of candidate loci is a critical step in obtaining insight into adaptation and acclimation of organisms. In this study of two non-model tropical (to sub-tropical) congeneric perciformes (Lates calcarifer and Lates niloticus) we characterized both coding and non-coding regions of lactate dehydrogenase-B (ldh-b), a locus which exhibits temperature-adaptive differences among temperate and sub-tropical populations of the North American killifish Fundulus heteroclitus. Ldh-b was 5,004 and 3,527 bp in length in L. calcarifer and L. niloticus, respectively, with coding regions comprising 1,005 bp in both species. A high level of sequence homology existed between species for both coding and non-coding regions of ldh-b (> 97% homology), corresponding to a 98.5% amino acid sequence homology. All six known functional sites within the encoded protein sequence (LDH-B) were conserved between the two Lates species. Ten simple sequence repeat (SSR) motifs (mono-, di-, tri- and tetranucleotide) and thirty putative microRNA elements (miRNAs) were identified within introns 1, 2, 5 and 6 of both Lates species. Five single nucleotide polymorphisms (SNPs) were also identified within miRNA containing intron regions. Such SNPs are implicated in several complex human conditions and/or diseases (as demonstrated by extensive genome-wide association studies). This novel characterization serves as a platform to further examine how non-model species may respond to changes in their native temperatures, which are expected to increase by up to 6 degrees C over the next century.

Differential SAGE analysis in Arabidopsis uncovers increased transcriptome complexity in response to low temperature

BackgroundAbiotic stress, including low temperature, limits the productivity and geographical distribution of plants, which has led to significant interest in understanding the complex processes that allow plants to adapt to such stresses. The wide range of physiological, biochemical and molecular changes that occur in plants exposed to low temperature require a robust global approach to studying the response. We have employed Serial Analysis of Gene Expression (SAGE) to uncover changes in the transcriptome of Arabidopsis thaliana over a time course of low temperature stress.ResultsFive SAGE libraries were generated from A. thaliana leaf tissue collected at time points ranging from 30 minutes to one week of low temperature treatment (4°C). Over 240,000 high quality SAGE tags, corresponding to 16,629 annotated genes, provided a comprehensive survey of changes in the transcriptome in response to low temperature, from perception of the stress to acquisition of freezing tolerance. Interpretation of these data was facilitated by representing the SAGE data by gene identifier, allowing more robust statistical analysis, cross-platform comparisons and the identification of genes sharing common expression profiles. Simultaneous statistical calculations across all five libraries identified 920 low temperature responsive genes, only 24% of which overlapped with previous global expression analysis performed using microarrays, although similar functional categories were affected. Clustering of the differentially regulated genes facilitated the identification of novel loci correlated with the development of freezing tolerance. Analysis of their promoter sequences revealed subsets of genes that were independent of CBF and ABA regulation and could provide a mechanism for elucidating complementary signalling pathways. The SAGE data emphasised the complexity of the plant response, with alternate pre-mRNA processing events increasing at low temperatures and antisense transcription being repressed.ConclusionAlternate transcript processing appears to play an important role in enhancing the plasticity of the stress induced transcriptome. Novel genes and cis-acting sequences have been identified as compelling targets to allow manipulation of the plant's ability to protect against low temperature stress. The analyses performed provide a contextual framework for the interpretation of quantitative sequence tag based transcriptome analysis which will prevail with the application of next generation sequencing technology.

Temperature‐induced gene expression associated with different thermal reaction norms for growth rate

Although nearly all organisms are subject to fluctuating temperature regimes in their natural habitat, little is known about the genetics underlying the response to thermal conditions, and even less about the genetic differences that cause individual variation in thermal response. Here, we aim to elucidate possible pathways involved in temperature-induced phenotypic plasticity of growth rate. Our model organism is the collembolan Orchesella cincta that occurs in a wide variety of habitats and is known to be adapted to local thermal conditions. Because sequence information is lacking in O. cincta, we constructed cDNA libraries enriched for temperature-responsive genes using suppression subtractive hybridization. We compared gene expression of O. cincta with steep thermal reaction norms (high plasticity) to those with flat thermal reaction norms (low plasticity) for juvenile growth after exposure to a temperature switch composed of a cooling or a warming treatment. Using suppression subtractive hybridization, we found differential expression of ten nuclear genes, including several genes involved in energy metabolism, such as pantothenate kinase and carbonic anhydrase. In addition, seven mitochondrial genes were found in the cloned subtracted library, but further analysis showed this was caused by allelic variation in mitochondrial genes in our founder population, and that a specific haplotype was associated with high thermal responsiveness. Future work will focus on candidate genes from pathways such as the oxidative phosphorylation and biosynthesis of coenzyme A which are possibly involved in thermal responsiveness of juvenile growth rate.

Expression patterns of low temperature responsive genes in a dominant ABA‐less‐sensitive mutant line of common wheat

Abscisic acid (ABA) plays important role in mediating stress responses and in acquiring desiccation tolerance and dormancy of plant seeds. To study roles of ABA in cold acclimation and freezing tolerance in wheat, expression profiles of Cor/Lea and their putative transcription factor (TF) genes were analysed using a dominant mutation line of common wheat EH47‐1 lacking seed dormancy. The mutant line was less sensitive to exogenous ABA than the original line as judged by the magnitude of ABA inhibition of seedling growth. Expression analysis of Cor/Lea and TF genes however, showed that more transcripts were present in ABA‐treated seedlings of the mutant line. In developing caryopses, the same tendency was observed. The mutant line showed no changes in the cold acclimation ability, but it showed a higher level of freezing tolerance than the original line without cold acclimation. No significant differences were observed in the expression profiles of Cor/Lea and TF genes during cold acclimation between the two lines. Our results imply the presence of an unknown ABA‐dependent cold responsive pathway, which enhances the basal level of freezing tolerance by a dominant mutation in EH47‐1.

Transcriptional Analysis of Long-Term Adaptation ofYersinia enterocoliticato Low-Temperature Growth

To analyze the transcriptional response of Yersinia enterocolitica cells to prolonged growth at low temperature, a collection of luxCDABE transposon mutants was cultivated in parallel at optimal (30 degrees C) and suboptimal (10 degrees C) temperatures and screened for enhanced promoter activities during growth until entering stationary phase. Among 5,700 Y. enterocolitica mutants, 42 transcriptional units were identified with strongly enhanced or reduced promoter activity at 10 degrees C compared to 30 degrees C, and changes in their transcriptional levels over time were measured. Green fluorescent protein fusions to 10 promoter regions confirmed the data. The temporal order of induction of the temperature-responsive genes of Y. enterocolitica was deduced, starting with the expression of cold shock genes cspA and cspB and the elevated transcription of a glutamate-aspartate symporter. Subsequently, cold-adapted cells drastically up-regulated genes encoding environmental sensors and regulators, such as UhpABC, ArcA, and methyl-accepting chemotaxis protein I (MCPI). Among the most prominent cold-responsive elements that were transcriptionally induced during growth in early and middle exponential phase are the insecticidal toxin genes tcaA and tcaB, as well as genes involved in flagellar synthesis and chemotaxis. The expression pattern of the late-exponential- to early-stationary-growth phase is dominated by factors involved in biodegradative metabolism, namely, a histidine ammonia lyase, three enzymes responsible for uptake and utilization of glycogen, the urease complex, and a subtilisin-like protease. Double-knockout mutants and complementation studies demonstrate inhibitory effects of MCPI and UhpC on the expression of a putative hemolysin transporter. The data partially delineate the spectrum of gene expression of Y. enterocolitica at environmental temperatures, providing evidence that an as-yet-unknown insect phase is part of the life cycle of this human pathogen.

Temperature-dependent growth rates and gene expression patterns of various medaka Oryzias latipes cell lines derived from different populations

Medaka Oryzias latipes has several geographically and genetically distinct populations. We examined temperature acclimation response in various medaka cell lines derived from different populations. Measurement of cell growth at various temperatures suggested that 15 degrees Celsius was the permissive growth temperature in all cell lines from the Northern Japanese and East Korean populations, but not in those from the Southern Japanese population and medaka-related species Oryzias celebensis, which inhabits a tropical zone. RT-PCR for 102 temperature-responsive genes, previously reported in other species, revealed that the accumulated mRNA level of a gene encoding HSP47 was lower at 25 degrees Celsius than at 33 degrees Celsius, and vice versa for 12 genes including IkappaBalpha and Rab-1c, in OLHNI-1 cell line from the Northern Japanese population. Further analysis by real-time PCR demonstrated that the accumulated mRNA levels of IkappaBalpha and Rab-1c in OLHNI-1 and OLSOK-e7 cell lines from the East Korean population were increased when the culture temperature was shifted from 33 to 15 degrees Celsius, but not in OLHdrR-e3 cell line from the Southern Japanese population. Since IkappaBalpha and Rab-1c are related to the NFkappaB cascade and endoplasmic reticulum-to-Golgi transport, respectively, it is inferred that immune responses and intracellular transport are possibly critical to temperature adaptation for medaka.