Unknown,Transcriptomics,Genomics,Proteomics

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Cardiac transcriptomic shifts during first 24h of thermal acclimation in the porcelain crab, Petrolisthes cinctipes

ABSTRACT: Intertidal zone organisms can experience transient freezing temperatures during winter low tides, but their extreme cold tolerance mechanisms are not known. Petrolisthes cinctipes is a temperate mid-high intertidal zone crab species that can experience wintertime habitat temperatures below the freezing point of seawater. We examined how cold tolerance changed during the initial phase of thermal acclimation to cold and warm temperatures, as well as the persistence of cold tolerance during long-term thermal acclimation. Thermal acclimation for as little as 6 hours at 8˚C enhanced crab tolerance during a 1h exposure to -2°C relative to crabs acclimated to 18˚C. Potential mechanisms for this enhanced tolerance were elucidated using cDNA microarrays to probe for differences in gene expression in cardiac tissue of warm and cold acclimated crabs during the first day of thermal acclimation. No changes in gene expression were detected until 12h of thermal acclimation. Genes strongly upregulated in warm acclimated crabs represented immune response and extracellular / intercellular processes, suggesting that warm acclimated crabs had a generalized stress response and may have been remodelling tissues or altering intercellular processes. Genes strongly upregulated in cold acclimated crabs included many that are involved in glucose production suggesting that cold acclimation involves increasing intracellular glucose as a cryoprotectant. Structural cytoskeletal proteins were also strongly represented among the genes upregulated in only cold acclimated crabs. There were no consistent changes in composition or the level of unsaturation of membrane phospholipid fatty acids with cold acclimation, which suggests that neither short- nor long-term changes in cold tolerance are mediated by changes in membrane fatty acid composition. Overall, our study demonstrates that initial changes in cold tolerance are likely not regulated by transcriptomic responses, but that gene expression-related changes in homeostasis begin within 12 hours – the length of a tidal cycle. all array data and raw images archived at the Porcelain Crab Array Database (http://array.sfsu.edu) n=264 specimens were divided into warm (18°C, n=96), cold (8°C, n=96), and control (13°C, n=72) acclimation groups. Crabs were sampled from the 13°C group at 0 h (the start of the experiment) and 24 h, the termination of the experiment. Crabs were sampled from the warm and cold acclimation groups at 6, 12, 18, and 24 hours following the start of thermal acclimation. At each time point, heart tissue from n=16 crabs from each group was dissected, flash frozen and stored at −80°C. A pooled total aRNA sample was prepared for each group by mixing equal quantities of total RNA from n=5 individuals in each group in order to have the same amount of biological diversity within each pooled RNA sample. For microarray hybridizations we used n=25 slides in an incomplete loop design where each sample was hybridized n=5 times, 2-3 times labelled with each Cy dye

ORGANISM(S): Petrolisthes cinctipes

SUBMITTER: Jonathon Stillman

PROVIDER: E-GEOD-35307 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Intertidal zone organisms can experience transient freezing temperatures during winter low tides, but their extreme cold tolerance mechanisms are not known. Petrolisthes cinctipes is a temperate mid-high intertidal zone crab species that can experience wintertime habitat temperatures below the freezing point of seawater. We examined how cold tolerance changed during the initial phase of thermal acclimation to cold and warm temperatures, as well as the persistence of cold tolerance during long-term thermal acclimation. Thermal acclimation for as little as 6 hours at 8˚C enhanced crab tolerance during a 1h exposure to -2°C relative to crabs acclimated to 18˚C. Potential mechanisms for this enhanced tolerance were elucidated using cDNA microarrays to probe for differences in gene expression in cardiac tissue of warm and cold acclimated crabs during the first day of thermal acclimation. No changes in gene expression were detected until 12h of thermal acclimation. Genes strongly upregulated in warm acclimated crabs represented immune response and extracellular / intercellular processes, suggesting that warm acclimated crabs had a generalized stress response and may have been remodelling tissues or altering intercellular processes. Genes strongly upregulated in cold acclimated crabs included many that are involved in glucose production suggesting that cold acclimation involves increasing intracellular glucose as a cryoprotectant. Structural cytoskeletal proteins were also strongly represented among the genes upregulated in only cold acclimated crabs. There were no consistent changes in composition or the level of unsaturation of membrane phospholipid fatty acids with cold acclimation, which suggests that neither short- nor long-term changes in cold tolerance are mediated by changes in membrane fatty acid composition. Overall, our study demonstrates that initial changes in cold tolerance are likely not regulated by transcriptomic responses, but that gene expression-related changes in homeostasis begin within 12 hours – the length of a tidal cycle. all array data and raw images archived at the Porcelain Crab Array Database (http://array.sfsu.edu)

Project description:During low temperature exposure, temperate plant species increase their freezing tolerance in a process termed cold acclimation. During deacclimation in response to warm temperatures cold acclimated plants lose freezing tolerance and resume growth and development. While considerable effort has been directed toward understanding the molecular and metabolic basis of cold acclimation, much less information is available about the regulation of deacclimation. Here, we report metabolic (GC-MS) and transcriptional (microarrays, qRT-PCR) responses underlying deacclimation during the first 24 h after a shift of cold acclimated Arabidopsis thaliana (Columbia-0) plants to warm temperature. The data revealed a faster response of the transcriptome than of the metabolome and provided evidence for tightly regulated temporal responses at both levels. Metabolically deacclimation is associated with decreasing contents of sugars, amino acids and glycolytic and TCA cycle intermediates, indicating an increased need for carbon sources and respiratory energy production associated with growth resumption under warm temperature conditions. Deacclimation also involves extensive down-regulation of protein synthesis and changes in the metabolism of lipids and cell wall components. Altered hormonal regulation appears particularly important during deacclimation, with changes in the expression of genes related to auxin, gibberellin, brassinosteroid, jasmonate and ethylene metabolisms. Several transcription factor families controlling fundamental aspects of plant development are significantly regulated during deacclimation, emphasizing that loss of freezing tolerance and growth resumption are interrelated processes that are transcriptionally highly interrelated. Expression patterns of some clock oscillator components during deacclimation resembled those under warm conditions, indicating at least partial re-activation of the circadian clock. This study provide the first comprehensive analysis of the regulation of deacclimation in cold acclimated plants. The data indicate cascades of rapidly regulated genes and metabolites that underly the developmental switch resulting in reduced freezing tolerance and the resumption of growth. They constitute a reference dataset of genes, metabolites and pathways that are crucial during the first rapid phase of deacclimation and will be useful for the further analysis of this important but under-researched plant process. We used whole genome microarrays to monitor changes in gene expression in the Arabidopsis thaliana accession Columbia-0 during the first 24 h after a shift of cold acclimated plants to warm temperature.

Project description:During cold acclimation plants increase their freezing tolerance in response to low non-freezing temperatures. This is accompanied by many physiological, biochemical and molecular changes that have been extensively investigated. In addition, many cold acclimated plants become more freezing tolerant during exposure to mild, non-damaging sub-zero temperatures. There is hardly any information available about the molecular basis of this adaptation. However, Arabidopsis thaliana is among the species that acclimate to sub-zero temperatures. This makes it possible to use the molecular and genetic tools available in this species to identify components of sub-zero signal transduction and acclimation. Here, we have used microarrays and a qRT-PCR primer platform covering 1880 genes encoding transcription factors to monitor changes in gene expression in the accessions Columbia-0, Rschew and Tenela during the first three days of sub-zero acclimation at -3°C. The results indicate that gene expression during sub-zero acclimation follows a tighly controlled time-course. Especially AP2/EREBP and WRKY transcription factors may be important regulators of sub-zero acclimation, although the CBF signal transduction pathway seems to be less important during sub-zero than during cold acclimation. Globally, we estimate that approximately 5% of all Arabidopsis genes are regulated during sub-zero acclimation. Particularly photosynthesis-related genes were down-regulated and genes belonging to the functional classes of cell wall biosynthesis, hormone metabolism and RNA regulation of transcription were up-regulated. Collectively, these data provide the first global analysis of gene expression during sub-zero acclimation and allow the identification of candidate genes for forward and reverse genetic studies into the molecular mechanisms of sub-zero acclimation. We used whole genome microarrays to monitor changes in gene expression in the Arabidopsis thaliana accessions Columbia-0, Rschew and Tenela during three days of acclimation to sub-zero temperature at -3°C after cold acclimation Plants from Arabidopsis thaliana accessions Columbia-0, Rschew and Tenela were cold acclimated at 4°C for two weeks. Detached leaves were then sub-zero acclimated at -3°C for 8 h, 1 d or 3 d at -3°C. Leaves of cold acclimated plants and sub-zero acclimated leaves were collected for RNA extraction and hybridization on Affymetrix ATH1 microarrays in order to explore temporal transcriptome changes during sub-zero acclimation. For each sample total RNA was isolated from a pool of three leaves from three different plants. The experiment was performed in three idenpendent biological replicates.

Project description:Proteomic response of adult Drosophila melanogaster acclimated at three contrasted thermal conditions (11, 25 and 31 degree Celsius) investigated by 2D-DIGE experiment. Thermal acclimation drastically alters thermotolerance of ectotherms, but the mechanisms determining this plastic response are not fully understood. The present study investigates the proteomic response (2D-DIGE) of adult Drosophila melanogaster acclimated at 11, 25 or 31 °C. As expected 11 °C-acclimation improved cold tolerance and 31 °C-acclimation improved heat tolerance. We hypothesized that the marked organismal responses to acclimation could be detected at the proteomic level assuming that changes in the abundance of specific proteins are linked to the physiological changes underlying the phenotypic response. The 31 °C-acclimated flies displayed a particular divergent proteomic profile where molecular chaperones made up a large number of the proteins that were modulated during heat acclimation. Many other proteins showed significant modulation during acclimation including proteins involved in iron ion and cell redox homeostasis, carbohydrate and energy metabolism, chromatin remodeling and translation, and contractile machinery. Interestingly the changes in protein abundance were often unrelated to transcriptional activity of the genes coding for the proteins, except for the most strongly expressed proteins (e.g. Hsp70). The 11 °C-acclimation evoked weak proteomic response despite the marked effect on the organismal phenotype. Thus the acquired cold tolerance observed here may involve regulatory process such as posttranslational regulation rather than de novo protein synthesis. Bioinformatics and data processing: The proteinScape 2.1 software (BrukerDaltonik GmbH) was used to submit MS/MS data to the following database: NCBI restricted to Drosophila (June 2011, 223,543 sequences) using the Mascot search engine (Mascot server v2.2, http://www.matrixscience.com). Parameters were set as follows: trypsin as enzyme with one allowed miscleavage, carbamidomethylation of cysteins as fixed modification and methionine oxidation as variable modifications. The mass tolerance for parent and fragment ions was set to 0.5 Da. Peptide identifications were accepted if the individual ion Mascot scores were above the identity threshold (the ion score is −10*log(P), where P is the probability that the observed match is a random event, P < 0.05). In case of ambiguous assignments (one compound fit to more than one peptide), peptide were accepted based on the peptide score, meaning that the peptide sequence with the highest score is accepted. The compilation of identified peptides to proteins was performed with the ProteinExtractor algorithm (Thiele et al., 2008, 2010), so that every protein reported was identified by at least one peptide with significant ion Mascot score (above the identity threshold).

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Cardiac transcriptomic shifts during first 24h of thermal acclimation in the porcelain crab, Petrolisthes cinctipes

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