Heat Stress Response Mechanisms Research Articles

Analysis of Genomic Alternative Splicing Patterns in Rat under Heat Stress Based on RNA-Seq Data.

Heat stress is one of the most severe challenges faced in livestock production in summer. Alternative splicing as an important post-transcriptional regulation is rarely studied in heat-stressed animals. Here, we performed and analyzed RNA-sequencing assays on the liver of Sprague-Dawley rats in control (22 °C, n = 5) and heat stress (4 °C for 120 min, H120; n = 5) groups, resulting in the identification of 636 differentially expressed genes. Identification analysis of the alternative splicing events revealed that heat stress-induced alternative splicing events increased by 20.18%. Compared with other types of alternative splicing events, the alternative start increased the most (43.40%) after heat stress. Twenty-eight genes were differentially alternatively spliced (DAS) between the control and H120 groups, among which Acly, Hnrnpd and mir3064 were also differentially expressed. For DAS genes, Srebf1, Shc1, Srsf5 and Ensa were associated with insulin, while Cast, Srebf1, Tmem33, Tor1aip2, Slc39a7 and Sqstm1 were enriched in the composition of the endoplasmic reticulum. In summary, our study conducts a comprehensive profile of alternative splicing in heat-stressed rats, indicating that alternative splicing is one of the molecular mechanisms of heat stress response in mammals and providing reference data for research on heat tolerance in mammalian livestock.

Investigating the biochemical effects of heat stress and sample quenching approach on the metabolic profiling of abalone (Haliotis iris).

Ocean temperatures have been consistently increasing due to climate change, and the frequency of heatwave events on shellfish quality is a growing concern worldwide. Typically, shellfish growing areas are in remote or difficult to access locations which makes in-field sampling and sample preservation of shellfish heat stress difficult. As such, there is a need to investigate in-field sampling approaches that facilitate the study of heat stress in shellfish. This study aims to apply a gas chromatography-mass spectrometry (GC-MS) based metabolomics approach to examine molecular mechanisms of heat stress responses in shellfish using abalone as a model, and compare the effects of different quenching protocols on abalone metabolic profiles. Twenty adult Haliotis iris abalone were exposed to two temperatures (14°C and 24°C) for 24h. Then, haemolymph and muscle tissues of each animal were sampled and quenched with 4 different protocols (liquid nitrogen, dry ice, cold methanol solution and normal ice) which were analyzed via GC-MS for central carbon metabolites. The effects of different quenching protocols were only observed in muscle tissues in which the cold methanol solution and normal ice caused some changes in the observed metabolic profiles, compared to dry ice and liquid nitrogen. Abalone muscle tissues were less affected by thermal stress than haemolymph. There were 10 and 46 compounds significantly influenced by thermal stress in muscle and haemolymph, respectively. The changes of these metabolite signatures indicate oxidative damage, disturbance of amino acid and fatty acid metabolism, and a shift from aerobic metabolism to anaerobic pathways. The study provided insights into the heat response of abalone, which could be useful for understanding the effects of marine heatwaves and summer mortality events on abalone. Dry ice appeared to be a suitable protocol, and safer in-field alternative to liquid nitrogen, for quenching of abalone tissues.

Transcriptomic Response under Heat Stress in Chickens Revealed the Regulation of Genes and Alteration of Metabolism to Maintain Homeostasis.

Simple SummaryWith the increased global temperature, the threat from climate change has already affected the livestock industry. Exposure to heat stress is a major factor responsible for impacts on the overall livestock production, which ultimately results in economic losses. With no exception, poultry is among the most vulnerable livestock to environmental stress. Hence, a comprehensive study is required to understand the molecular mechanisms and to improve the breeding program to overcome economic losses. Therefore, we investigated growth related phenotypes and performed transcriptome analysis to understand the heat stress response in chickens. Animal experiments were designed with two groups, which were kept at 21 and 33 °C for 2 weeks as the control and treatment groups. The transcriptome analysis used blood samples from each chicken. In this study, we identified a total of 245 differentially expressed genes (DEGs) with important roles in various biological processes, such as cell protection, energy conversion in the mitochondria, and protein quality control. The results indicate that the heat stress environment regulates genes and alter the metabolism to adjust for the heat environment in chickens. These findings could be useful to help understand the heat stress response in poultry.Chicken is important livestock that serves as a vital food source which remain largely affected by heat stress. Therefore, we performed the transcriptome analysis to help understand the mechanisms of heat stress response in chickens. In the animal experiments, we grouped them into a normal and severe at 21 and 33 °C, with identified physiologic parameters for 2-weeks. Subsequently, RNA-seq analysis was performed to identify DEGs with a false discovery rate < 0.05 and a fold change ≥ 1.5. In the physiological parameters, we observed average daily gain was declined, rectal temperature and respiration rate was increased in severe group. Among total 245 DEGs, 230 and 15 genes were upregulated and downregulated, respectively. In upregulated DEGs, HSPs, MYLK2, and BDKRB1 genes were identified as key genes in heat stress. The KEGG pathway analysis showed involvement in the ATP metabolic process, MAPK signaling pathway and calcium signaling pathway with related protein processing and synthesis. In conclusion, with induced heat stress, such changes in physiologic parameters alter the neuroendocrine system, and we observed that the heat stress environment regulates such Heat shock protein genes to protect the cells and proteins from an altered metabolism. These findings provide a more comprehensive understanding of the heat stress response in poultry.

ITRAQ-based quantitative proteomic analysis of heat stress-induced mechanisms in pepper seedlings.

BackgroundAs one of the most important vegetable crops, pepper has rich nutritional value and high economic value. Increasing heat stress due to the global warming has a negative impact on the growth and yield of pepper.MethodsTo understand the heat stress response mechanism of pepper, an iTRAQ-based quantitative proteomic analysis was employed to identify possible heat-responsive proteins and metabolic pathways in 17CL30 and 05S180 pepper seedlings under heat stress.ResultIn the present study, we investigated the changes of phenotype, physiology, and proteome in heat-tolerant (17CL30) and heat-sensitive (05S180) pepper cultivars in response to heat stress. Phenotypic and physiological changes showed that 17CL30 had a stronger ability to resist heat stress compared with 05S180. In proteomic analysis, a total of 3,874 proteins were identified, and 1,591 proteins were considered to participate in the process of heat stress response. According to bioinformatic analysis of heat-responsive proteins, the heat tolerance of 17CL30 might be related to a higher ROS scavenging, photosynthesis, signal transduction, carbohydrate metabolism, and stress defense, compared with 05S180.

Heat stress response mechanisms in pollen development.

SummaryBeing rooted in place, plants are faced with the challenge of responding to unfavourable local conditions. One such condition, heat stress, contributes massively to crop losses globally. Heatwaves are predicted to increase, and it is of vital importance to generate crops that are tolerant to not only heat stress but also to several other abiotic stresses (e.g. drought stress, salinity stress) to ensure that global food security is protected. A better understanding of the molecular mechanisms that underlie the temperature stress response in pollen will be a significant step towards developing effective breeding strategies for high and stable production in crop plants. While most studies have focused on the vegetative phase of plant growth to understand heat stress tolerance, it is the reproductive phase that requires more attention as it is more sensitive to elevated temperatures. Every phase of reproductive development is affected by environmental challenges, including pollen and ovule development, pollen tube growth, male–female cross‐talk, fertilization, and embryo development. In this review we summarize how pollen is affected by heat stress and the molecular mechanisms employed during the stress period, as revealed by classical and ‐omics experiments.

Regulation of DNA (de)Methylation Positively Impacts Seed Germination during Seed Development under Heat Stress.

Seed development needs the coordination of multiple molecular mechanisms to promote correct tissue development, seed filling, and the acquisition of germination capacity, desiccation tolerance, longevity, and dormancy. Heat stress can negatively impact these processes and upon the increase of global mean temperatures, global food security is threatened. Here, we explored the impact of heat stress on seed physiology, morphology, gene expression, and methylation on three stages of seed development. Notably, Arabidopsis Col-0 plants under heat stress presented a decrease in germination capacity as well as a decrease in longevity. We observed that upon mild stress, gene expression and DNA methylation were moderately affected. Nevertheless, upon severe heat stress during seed development, gene expression was intensively modified, promoting heat stress response mechanisms including the activation of the ABA pathway. By analyzing candidate epigenetic markers using the mutants’ physiological assays, we observed that the lack of DNA demethylation by the ROS1 gene impaired seed germination by affecting germination-related gene expression. On the other hand, we also observed that upon severe stress, a large proportion of differentially methylated regions (DMRs) were located in the promoters and gene sequences of germination-related genes. To conclude, our results indicate that DNA (de)methylation could be a key regulatory process to ensure proper seed germination of seeds produced under heat stress.

Differential requirement of MED14/17 recruitment for activation of heat inducible genes.

The mechanism of heat stress response in plants has been studied, focusing on the function of transcription factors (TFs). Generally, TFs recruit coactivators, such as Mediator, are needed to assemble the transcriptional machinery. However, despite the close relationship with TFs, how coactivators are involved in transcriptional regulation under heat stress conditions is largely unclear. We found a severe thermosensitive phenotype of Arabidopsis mutants of MED14 and MED17. Transcriptomic analysis revealed that a quarter of the heat stress (HS)-inducible genes were commonly downregulated in these mutants. Furthermore, chromatin immunoprecipitation assay showed that the recruitment of Mediator by HsfA1s, the master regulators of heat stress response, is an important step for the expression of HS-inducible genes. There was a differential requirement of Mediator among genes; TF genes have a high requirement whereas heat shock proteins (HSPs) have a low requirement. Furthermore, artificial activation of HsfA1d mimicking perturbation of protein homeostasis induced HSP gene expression without MED14 recruitment but not TF gene expression. Considering the essential role of MED14 in Mediator function, other coactivators may play major roles in HSP activation depending on the cellular conditions. Our findings highlight the importance of differential recruitment of Mediator for the precise control of HS responses in plants.

Transcriptional Basis for Differential Thermosensitivity of Seedlings of Various Tomato Genotypes.

Transcriptional reprograming after the exposure of plants to elevated temperatures is a hallmark of stress response which is required for the manifestation of thermotolerance. Central transcription factors regulate the stress survival and recovery mechanisms and many of the core responses controlled by these factors are well described. In turn, pathways and specific genes contributing to variations in the thermotolerance capacity even among closely related plant genotypes are not well defined. A seedling-based assay was developed to directly compare the growth and transcriptome response to heat stress in four tomato genotypes with contrasting thermotolerance. The conserved and the genotype-specific alterations of mRNA abundance in response to heat stress were monitored after exposure to three different temperatures. The transcripts of the majority of genes behave similarly in all genotypes, including the majority of heat stress transcription factors and heat shock proteins, but also genes involved in photosynthesis and mitochondrial ATP production. In turn, genes involved in hormone and RNA-based regulation, such as auxin- and ethylene-related genes, or transcription factors like HsfA6b, show a differential regulation that associates with the thermotolerance pattern. Our results provide an inventory of genes likely involved in core and genotype-dependent heat stress response mechanisms with putative role in thermotolerance in tomato seedlings.

Effects of chronic heat stress on mRNA and miRNA expressions in dairy cows

Heat stress has a negative impact on dairy cow productivity. In order to reveal the mechanisms of heat-stress response, the mRNA and miRNA expression profiles of five cows under chronic heat-stress and thermoneutral conditions were assayed in blood by high-throughput sequencing technology. A total of 540 mRNAs and 9 miRNAs were expressed differently under heat-stress and thermoneutral conditions. Functional analyses revealed that MAPK signaling pathway, cellular senescence, circadian entrainment, aldosterone synthesis and secretion, and pathways in cancer were enriched for differently expressed mRNAs; meanwhile cGMP-PKG signaling pathway, thermogenesis, and protein digestion and absorption were enriched for differently expressed miRNAs. In addition, GADD45G, TGFB2, and GNG11 may play roles in the heat stress, and bta-miR-423-5p might be one of the regulators of heat-stress response in cows as potential mediators of chronic heat-stress response. In conclusion, the present study described the mRNA and miRNA expression patterns in blood extracted from cows during the transition from heat-stress to thermoneutral conditions. The results provide new data that could help in identifying mechanisms that mediate cows' response to chronic heat stress.

Proteomic analysis of Aspergillus niger 3.316 under heat stress.

β‐Glucosidase production by Aspergillus niger is accompanied by an inevitable temperature increase in the industrial fermentation environment. Hence, the synthetic process of β‐glucosidase is negatively affected. However, our understanding of the heat stress response (HSR) mechanism in A. niger is still incomplete. The current study explored the intracellular proteome profile of A. niger 3.316 in group T (50°C stress) and group C (30°C control) using two proteomic approaches (isobaric tags for relative and absolute quantitation [iTRAQ] and label‐free) and examined the expression of four proteins using a parallel reaction monitoring (PRM) approach. Based on the result of the iTRAQ proteomic analysis, 1,025 proteins were differentially expressed in group T compared to group C. Using the label‐free approach, we only focused on 77 proteins with significant changes in their protein expression levels. In addition, we performed bioinformatics analysis on all these proteins and obtained detailed gene ontology (GO) enrichment and Kyoto encyclopedia of genes and genomes (KEGG) pathway results. Under heat stress conditions, the relative expression levels of proteins with protection and repair functions were upregulated in A. niger 3.316. These proteins were involved in metabolic pathways, oxidative phosphorylation, porphyrin and chlorophyll metabolism, pyruvate metabolism, and the citrate cycle (TCA cycle). The insights obtained from the presented proteomics and bioinformatics analyses can be used to further explore the HSR mechanism of A. niger.

YODA-HSP90 Module Regulates Phosphorylation-Dependent Inactivation of SPEECHLESS to Control Stomatal Development under Acute Heat Stress in Arabidopsis

Stomatal ontogenesis, patterning, and function are hallmarks of environmental plant adaptation, especially to conditions limiting plant growth, such as elevated temperatures and reduced water availability. The specification and distribution of a stomatal cell lineage and its terminal differentiation into guard cells require a master regulatory protein phosphorylation cascade involving the YODA mitogen-activated protein kinase kinase kinase. YODA signaling results in the activation of MITOGEN-ACTIVATED PROTEIN KINASEs (MPK3 and MPK6), which regulate transcription factors, including SPEECHLESS (SPCH). Here, we report that acute heat stress affects the phosphorylation and deactivation of SPCH and modulates stomatal density. By using complementary molecular, genetic, biochemical, and cell biology approaches, we provide solid evidence that HEAT SHOCK PROTEINS 90 (HSP90s) play a crucial role in transducing heat-stress response through the YODA cascade. Genetic studies revealed that YODA and HSP90.1 are epistatic, and they likely function linearly in the same developmental pathway regulating stomata formation. HSP90s interact with YODA, affect its cellular polarization, and modulate the phosphorylation of downstream targets, such as MPK6 and SPCH, under both normal and heat-stress conditions. Thus, HSP90-mediated specification and differentiation of the stomatal cell lineage couples stomatal development to environmental cues, providing an adaptive heat stress response mechanism in plants.

Cardiac and Skeletal Muscle Transcriptome Response to Heat Stress in Kenyan Chicken Ecotypes Adapted to Low and High Altitudes Reveal Differences in Thermal Tolerance and Stress Response.

Heat stress (HS) negatively affects chicken performance. Agricultural expansion will happen in regions that experience high ambient temperatures, where fast-growing commercial chickens are vulnerable. Indigenous chickens of such regions, due to generations of exposure to environmental challenges, might have higher thermal tolerance. In this study, two indigenous chicken ecotypes, from the hot and humid Mombasa (lowland) and the colder Naivasha (highland) regions, were used to investigate the effects of acute (5 h, 35°C) and chronic (3 days of 35°C for 8 h/day) HS on the cardiac and skeletal muscle, through RNA sequencing. The rectal temperature gain and the number of differentially expressed genes (DEGs) [False Discovery Rate (FDR) < 0.05] were two times higher in the acute stage than in the chronic stage in both ecotypes, suggesting that cyclic exposure to HS can lead to adaptation. A tissue- and stage-specific difference in response to HS was observed, with peroxisome proliferator-activated-receptor (PPAR) signaling and mitogen-activate protein kinase (MAPK) signaling pathways, enriched in heart and skeletal muscle, respectively, and the p53 pathway enriched only in the acute stage in both tissues. The acute and chronic stage DEGs were integrated by a region-specific gene coexpression network (GCN), and genes with the highest number of connections (hub genes) were identified. The hub genes in the lowland network were CCNB2, Crb2, CHST9, SESN1, and NR4A3, while COMMD4, TTC32, H1F0, ACYP1, and RPS28 were the hub genes in the highland network. Pathway analysis of genes in the GCN showed that p53 and PPAR signaling pathways were enriched in both low and highland networks, while MAPK signaling and protein processing in endoplasmic reticulum were enriched only in the gene network of highland chickens. This shows that to dissipate the accumulated heat, to reduce heat induced apoptosis, and to promote DNA damage repair, the ecotypes activated or suppressed different genes, indicating the differences in thermal tolerance and HS response mechanisms between the ecotypes. This study provides information on the HS response of chickens, adapted to two different agro climatic environments, extending our understanding of the mechanisms of HS response and the effect of adaptation in counteracting HS.

Targeting Hsp20 Using the Novel Small Non-coding RNA DnrH Regulates Heat Tolerance in Deinococcus radiodurans.

Small non-coding RNAs (ncRNAs) are a class of regulatory molecules, which remain understudied in bacteria. In the extremophilic bacterium Deinococcus radiodurans, although hundreds of ncRNAs have been identified, few have been characterized in detail. In this study, we report the identification and characterization of a novel heat-inducible ncRNA named DnrH. Heat tolerance analysis showed that deleting DnrH significantly inhibited viability in response to high temperature conditions. Comparative phenotypic and qRT-PCR analyses of a DnrH mutant (ΔDnrH) and wild-type (WT) D. radiodurans suggested that DnrH is potentially involved in regulating the expression of the heat shock-related gene Hsp20. Microscale thermophoresis and genetic complementation showed that a 28-nucleotide (nt) sequence in the stem-loop structure of DnrH (143–170 nt) pairs with its counterpart in the coding region of Hsp20 mRNA (91–117 nt) via a 22 nt region. In vivo, mutation of the 22-nt region in the D. radiodurans genome led to a reduction in heat tolerance similar to that observed in the DnrH-mutant. Our results show that DnrH positively influences heat tolerance by increasing the transcription of Hsp20 mRNA, demonstrating, for the first time, a ncRNA that directly controls the expression of a heat stress-resistance gene. This work provides new insight into the heat stress response mechanism of D. radiodurans as well as other extremophiles that express similar Hsp20 proteins.

Transcriptomic insights into the heat stress response of Dunaliella bardawil

The halophilic green alga Dunaliella bardawil has been used for commercial production of natural β-carotene by large-scale outdoor cultivation, which often suffers from heat stress especially at noon in hot summers. In this study, the effects of heat stress on cell growth, pigment contents, and activities of antioxidant system in D. bardawil were studied, and RNA-seq experiment was conducted to analyze the transcriptional response to heat stress (42 °C for 2 h) in D. bardawil. High temperature (42 °C) for short time treatment (≤3 h) did not severely affect the cell growth and pigment accumulation of D. bardawil. Multiple genes encoding heat shock proteins for protein folding and antioxidant enzymes against toxic reactive oxygen species were substantially up-regulated significantly under heat stress. D. bardawil cells tended to shift from aerobic to glycolytic metabolism for energy production to increase survival chances under heat stress. Furthermore, the enrichment of ascorbate-glutathione cycle, up-regulation of genes responsible for chloroplast membranes, and changes in lipid characteristics like carbon chain length and unsaturation degree could play a vital role in achieving thermotolerance of D. bardawil. Taken together, this study improved our understanding of the molecular mechanisms of heat stress responses in D. bardawil.

Variations in physiological response and expression profiles of proline metabolism-related genes and heat shock transcription factor genes in petunia subjected to heat stress

High temperature is a critical constraint factor limiting plant growth and productivity of crops throughout the world. Petunia (Petunia hybrida) is a well-known ornamental plant which is widely grown in gardens. However, petunia does not tolerate high temperatures well and seedling production and landscape uses are seriously inhibited in the summer. In this study, a total of ten physiological indexes were selected to analyze the physiological responses of petunia ‘W115’ under heat stress. Partial least squares-discriminant analysis showed that the physiological responses of petunia ‘W115’ to heat stress could be classified into three critical time points (0.5 h, 3 h, and 72 h after stress), and that proline concentration was the most important core physiological index to reflect the heat-stress response of petunia seedlings. In order to elucidate the molecular regulatory mechanism of heat-stress response in petunia ‘W115’, the expression patterns of eight proline metabolism pathway genes and of 12 heat shock transcription factors (Hsfs) were studied by real-time quantitative PCR under heat stress. Expression analysis indicated that the expression trend of the proline synthetase gene (PhP5CR) was most closely correlated with the change in proline concentration, and the expression levels of most Hsfs were markedly up-regulated, especially PhHsf19 and PhHsf21, which were highly expressed during the entire heat-stress period, suggesting that these genes could play important roles in response to heat stress in petunia ‘W115’. We hope this research could deepen our understanding of the heat-stress response mechanism of petunia ‘W115’ seedlings and identify candidate genes for molecular breeding for heat tolerance in the future.

Physicochemical changes in liver and Hsc70 expression in pikeperch Sander lucioperca under heat stress

The pikeperch Sander lucioperca is an economically important freshwater species that is currently threatened by higher summer temperatures caused by global warming. To clarify the physiological state of pikeperch reared under relatively high temperatures and to acquire valuable biomarkers to monitor heat stress in this species, 100 fish were subjected to five different temperature treatments, ranging from 23 °C (control) to 36 °C. The physiological and biochemical indexes of liver and blood were determined, and heat-shock cognate 70 kDa protein (Hsc70) mRNA expression profiles were analyzed. The results showed that the activities of superoxide dismutase, catalase, and glutathione peroxidase in heat-stressed pikeperch first increased and then decreased, exhibiting peaks at 34 °C, 28 °C, and 28 °C, respectively. The level of thiobarbituric acid-reactive substances (TBARS) in all experimental groups was significantly higher than that of the control. The numbers of red blood cells, the packed-cell volume, and the contents of hemoglobin were significantly higher in the 34 °C and 36 °C treatment groups. Under heat stress, the albumin, cholesterol, and triglycerides contents decreased with increasing temperatures. Real-time fluorescence-based quantitative RT-PCR showed that Hsc70 mRNA levels increased in all eight of the tested tissues under heat stress. Expression reached maximum levels at 34 °C in the muscle, heart and gill tissues, and at 36 °C in the other five tissues. These results demonstrate that several physiological and biochemical phenotypes, such as oxidative stress, antioxidant enzymes and molecular chaperones, could be important biomarkers of heat stress in pikeperch, and are potentially valuable to uncover the mechanisms of heat-stress responses in fish.

Identification of Potential Genes Responsible for Thermotolerance in Wheat under High Temperature Stress.

Wheat, a major worldwide staple food crop, is relatively sensitive to a changing environment, including high temperature. The comprehensive mechanism of heat stress response at the molecular level and exploitation of candidate tolerant genes are far from enough. Using transcriptome data, we analyzed the gene expression profiles of wheat under heat stress. A total of 1705 and 17 commonly differential expressed genes (DEGs) were identified in wheat grain and flag leaf, respectively, through transcriptome analysis. Gene Ontology (GO) and pathway enrichment were also applied to illustrate the functions and metabolic pathways of DEGs involved in thermotolerance of wheat grain and flag leaf. Furthermore, our data suggest that there may be a more complex molecular mechanism or tighter regulatory network in flag leaf than in grain under heat stress over time, as less commonly DEGs, more discrete expression profiles of genes (principle component analysis) and less similar pathway response were observed in flag leaf. In addition, we found that transcriptional regulation of zeatin, brassinosteroid and flavonoid biosynthesis pathways may play an important role in wheat’s heat tolerance. The expression changes of some genes were validated using quantitative real-time polymerase chain reaction and three potential genes involved in the flavonoid biosynthesis process were identified.

Transcriptomic responses to heat stress in rainbow trout Oncorhynchus mykiss head kidney

Rainbow trout (Oncorhynchus mykiss) are widely cultured throughout the word for commercial aquaculture. However, as a cold-water species, rainbow trout are highly susceptible to heat stress, which may cause pathological signs or diseases by alleviating the immune roles and then lead to mass mortality. Understanding the molecular mechanisms that occur in the rainbow trout in response to heat stress will be useful to decrease heat stress-related morbidity and mortality in trout aquaculture. In the present study, we conducted transcriptome analysis of head kidney tissue in rainbow trout under heat-stress (24 °C) and control (18 °C) conditions, to identify heat stress-induced genes and pathways. More than 281 million clean reads were generated from six head kidney libraries. Using an adjusted P-value of P < 0.05 as the threshold, a total of 443 differentially expressed genes (DEGs) were identified, including members of the HSP90, HSP70, HSP60, and HSP40 family and several cofactors or cochaperones. The RNA-seq results were confirmed by RT-qPCR. Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis of DEGs were performed. Many genes involved in maintaining homeostasis or adapting to stress and stimuli were highly induced in response to high temperature. The most significantly enriched pathway was “Protein processing in endoplasmic reticulum (ER)”, a quality control system that ensures correct protein folding or degradation of misfolded polypeptides by ER-associated degradation. Other signaling pathways involved in regulation of immune system and post-transcriptional regulation of spliceosome were also critical for thermal adaptation. These findings improve our understanding of the molecular mechanisms of heat stress responses and are useful to develop strategies for the improvement of rainbow trout survival rate during summer high-temperature period.

Physiological and transcriptomic analysis revealed the involvement of crucial factors in heat stress response of Rhododendron hainanense

Molecular regulatory mechanism of heat stress response (HSR) in Ericaceae remains unknown. Here, we sought to identify HSR mechanisms in Rhododendron hainanense, a Ericaceae species, through a combination of physiological and transcriptomic studies. The levels of MDA, H2O2, Pro, SOD, CAT and APX in leaves of R. hainanense were analyzed to characterize a dramatic difference in varied temperature treatment. Also, three sequencing libraries, including one control and two heat stress (HS)-treated samples, were constructed for comparative transcriptomic analysis. By Illumina sequencing and Trinity strategy, 350 million clean reads (average length = 149 bp) was assembled into 183,486 unigenes. According to analysis of differential expression genes (DEGs), a total of 2658 DEGs were obtained. Moreover, a complex interaction network of 982 DEGs was established, of which master portions were comprised of 109 transcription factors (TFs). Importantly, integrated differential expression profiling, qRT-PCR and functional analysis, several TFs of R. hainanense (ABR1, IAA26, OBF1, LUX, SCL3, DIV, NAC29, NAC72 and TCP3) and their potential regulations for the crosstalk between hormonal signal and HSR were identified. These findings will contribute to our understanding of the regulatory mechanisms of HSR in R. hainanense, breeding cultivars with improved thermotolerance.

Antioxidant response to heat stress in seagrasses. A gene expression study

Seawater warming associated to the ongoing climate change threatens functioning and survival of keystone coastal benthic species such as seagrasses. Under elevated temperatures, the production of reactive oxygen species (ROS) is increased and plants must activate their antioxidant defense mechanisms to protect themselves from oxidative damage. Here we explore from a molecular perspective the ability of Mediterranean seagrasses to activate heat stress response mechanisms, with particular focus on antioxidants. The level of expression of targeted genes was analyzed in shallow and deep plants of the species Posidonia oceanica and in shallow plants of Cymodocea nodosa along an acute heat exposure of several days and after recovery. The overall gene expression response of P. oceanica was more intense and complete than in C. nodosa and reflected a higher oxidative stress level during the experimental heat exposure. The strong activation of genes with chaperone activity (heat shock proteins and a luminal binding protein) just in P. oceanica plants, suggested the higher sensitivity of the species to increased temperatures. In spite of the interspecific differences, genes from the superoxide dismutase (SOD) family seem to play a pivotal role in the thermal stress response of Mediterranean seagrasses as previously reported for other marine plant species. Shallow and deep P. oceanica ecotypes showed a different timing of response to heat. Shallow plants early responded to heat and after a few days relaxed their response which suggests a successful early metabolic adjustment. The response of deep plants was delayed and their recovery incomplete evidencing a lower resilience to heat in respect to shallow ecotypes. Moreover, shallow ecotypes showed some degree of pre-adaptation to heat as most analyzed genes showed higher constitutive expression levels than in deep ecotypes. The recurrent exposure of shallow plants to elevated summer temperatures has likely endowed them with a higher basal level of antioxidant defense and a faster responsiveness to warming than deep plants. Our findings match with previous physiological studies and supported the idea that warming will differently impact Mediterranean seagrass meadows depending on the species as well as on the depth (i.e. thermal regimen) at which the meadow grows. The increase in the incidence of summer heat waves could therefore produce a significant change in the distribution and composition of Mediterranean seagrass meadows with considerable consequences for the functioning of the whole ecosystem and for the socio-economic services that these ecosystems offer to the riverine populations.