Larvae of Drosophila melanogaster exhibit transcriptional activation of immune response pathways and antimicrobial peptides during recovery from supercooling stress
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ABSTRACT: The biochemical and molecular mechanisms of insect cold acclimation prior to cold stress have been relatively well explored. The mechanisms linked to recovery and repair after cold stress received much less attention.Here we focus on recovery from cold stress in the larvae of vinegar fly (Drosophila melanogaster) that were exposed to two physiologically distinct cold stress situations; supercooling (S, survival > 90%) and freezing (F, survival < 10%), both at -5°C.The transcriptomic responses to cold stress were analyzed.
Project description:Stress acclimation is an effective mechanism that plants acquired for adaption to dynamic environmental conditions. After undergoing cold acclimation, plants become more tolerant to cold stress. In order to understand the mechanism of cold acclimation, we performed a systematic, comprehensive study of cold response and acclimation in Cassava (Manihot esculenta), a staple crop and major food source in the tropical regions of the world. We profiled mRNA genes and small-RNA species, using next generation sequencing, and performed an integrative analysis of the transcriptome and microRNAome of Cassava across the normal condition, a moderate cold stress at 14M-BM-0C, a harsh stress at 4M-BM-0C after cold acclimation at 14M-BM-0C, and a cold shock from 24M-BM-0C to 4M-BM-0C. Two results from the analysis were striking. First, the moderate stress and cold shock, despite a difference of 10M-BM-0C between the two, triggered comparable degrees of perturbation to the transcriptome; in contrary, further harsh stress after cold acclimation resulted in a much smaller degree of transcriptome variation. Second and more importantly, about two thirds of the up- or down-regulated genes after moderate stress reversed their expression to down- or up-regulation, respectively, under harsh stress after cold acclimation, resulting in a genome-wide rewiring of regulatory networks. MicroRNAs, which are key post-transcriptional gene regulators, were major players in this massive rewiring of genetic circuitry. Further, a function enrichment analysis of the perturbed genes revealed that cold acclimation helped the plant to develop immunity to further harsh stress by exclusively inducing genes with functions of nutrient reservoir; in contrast, many genes with functions of viral reproduction were induced by cold shock. Our study revealed, for the first time, the molecular basis of stress acclimation in plants, and shed lights on the role of microRNA gene regulation in cold response and acclimation in Euphorbia. Three organs/tissues (folded leaf, fully expanded leaf and roots) of Cassava cultivar SC124 harvested at 6h, 24h and 5d for three cold treatments of CA, CCA and CS, for gene expression profiling at the stages of initial response, secondary response, and functional adaption to cold stresses. Total RNA of each sample was isolated individually, and then pooled with an equal amount from each sample into one for profiling. As a result, four mRNA libraries and four small-RNA libraries, corresponding to the conditions of CA, CCA, CS and NC, were constructed.
Project description:Rice in tropical and sub-tropical areas is often subjected to cold stress at the seedling stage resulting in poor growth and yield loss. In general, japonica rice has stronger cold tolerance (CT) than indica rice. However, several favorite alleles for CT exist in indica rice and can be used to enhance CT under japonica background. Genome-wide gene expression profiling is the efficient way to decipher the molecular genetic mechanism of CT enhancement and provide valuable information for the CT improvement in rice molecular breeding. In this study, the transcriptome of a CT introgressed line (IL) K354 and its recurrent parent C418 under cold stress were comparatively analyzed to explore the possible CT enhancement mechanism of K354.Totally 3184 differentially expressed genes (DEGs) including 195 transcription factors were identified in both lines under cold stress, about half of them were commonly regulated, which were involved in the major cold responsive pathways including OsDREB1s regulon. The K354-specific cold-induced DEGs were mainly related to stimulus response, cellular cell wall organization, and microtubule-based movement process compared with commonly cold-induced DEGs by GO analysis. Moreover, 296 constitutive DEGs with significantly different transcription level between C418 and K354 were detected under both control and cold stress conditions. K354-specific cold-regulated and constitutive DEGs jointly account for the CT improvement of K354. Pathway analysis unraveled up-regulation of starch and sucrose metabolism in both genotypes and presumably weaker defense response to stress in K354 than C418 under cold stress. Candidate genes prediction based on previous putative CT genetic networks revealed genotype-dependent CT enhancement mechanism in CT IL K354 vs recurrent parent C418, including Sir2, OsFAD7, OsWAK112d, and PCD related genes, etc.We propose a hypothesis of the CT enhancement mechanism in rice based on the results in present study. Firstly, a number of cold-regulated genes are able to express constitutively at high level or absent under control condition for standing by the arrival of cold stress. Next, under cold stress slower perception of stress signal from the more cold-tolerant membrane can postpone the activation of defensive system which may have possible negative effects on rice growth. Then, PCD will be launched to sacrifice a few cells for maintaining the basal growth of most cells. Finally, the protective responses on multiple aspects of cold damage will be postponed because of delayed several cold-defensive pathways (i.e. OsDREB1C regulon). It might explain why the recovery capacity of K354 from cold stress to control condition is stronger than C418. The CT enhancement mechanism can be regarded as the possible way to improve CT of japonica rice using indica germplasm in rice breeding program. In this study, the specific gene expression patterns of two genotypes (C418 and K354) at 2h, 6h, 12h, 24h and 48h during cold stress treatment (4 C) and control were characterized by using the Affymetrix rice microarray platform.
Project description:Control of mRNA half-life is a powerful strategy to adjust individual mRNA levels to various stress conditions, because the mRNA degradation rate controls not only the steady-state mRNA level but also the transition speed of mRNA levels. Here, we analyzed mRNA half-life changes in response to cold stress in Arabidopsis cells using genome-wide analysis, in which mRNA half-life measurements and transcriptome analysis were combined. Half-lives of average transcripts were determined to be elongated under cold conditions. Taking this general shift into account, we identified more than a thousand transcripts that were classified as relatively stabilized or relatively destabilized. The relatively stabilized class was predominantly observed in functional categories that included various regulators involved in transcriptional, post-transcriptional and post-translational processes. On the other hand, the relatively destabilized class was enriched in categories related to stress and hormonal response proteins, supporting the idea that rapid decay of mRNA is advanta- geous for swift responses to stress. In addition, pentatricopeptide repeat, cyclin-like F-box and Myb transcription factor protein families were significantly over-represented in the relatively destabilized class. The global analysis presented here demonstrates not only the importance of mRNA turn-over control in the cold stress response but also several structural characteristics that might be important in the control of mRNA stability. To demonstrate the importance of mRNA stability control in cold stress response, we investigated global changes in mRNA half-lives in response to cold treatment by micaroarray using Arabidopsis suspension cell cultures (T87 cells). Control cells were collected prior to transcriptional inhibitor (cordycepin) treatment (0 h), and 1 and 3 h after the start of cordycepin treatment. For cold-treated cells, 6 h samples were also used for microarray analyses. The experiments were performed with triplicate sets for each time point.
Project description:In the field, insects suffer multiple cold exposures during winter. When exposed to repeated low temperatures, Drosophila melanogaster females showed an increase in survival, but a reduction in reproduction. In this study, the microarrays were used to analyze the gene expression of female D. melanogaster after multiple, single sustained (or single prolonged) and single short cold treatments, which exposed the flies at 0 M-0C for repeated 2 h, single 10 h and single 2 h respectively. Candidate genes that were involved in 6 h recovery from different types of cold exposures were identified. After repeated cold exposures, candidates particularly included genes involved in muscle protein and muscle activity. Stress-related genes, Turandot A, Turandot C, and Turandot M were up-regulated in response to multiple cold exposures, and improve the cold survival in female D. melanogaster. This work also suggested a strong relationship between cold exposure and the immune system. I suggest that in fruit flies, chilling injuries after cold exposure may induce immune responses and contribute to recovery from cold.
Project description:Lysine acetylation in proteins is a dynamic and reversible Post-translational modification and plays an important role in diverse cellular processes, but limited knowledge is available for acetylation modifications in pepper (Capsicum annuum L.) resistance of cold stress. In this study, the proteome and acetylome of two peppers with different cold resistance under different cold stress and recovery treatment were investigated. In total, 6213 proteins groups and 4574 lysine acetylation sites of 2261 protein groups were identified. Cold stress and recovery treatment results in 3008 differentially expressed proteins (DEPs) and 768 differentially expressed acetylated proteins (DEAPs). Further analysis found a total 1988 proteins were both identifed in the proteome and acetylome data and elucidated the functional differences between the up-regulated and down-regulated proteins in these co-identified proteins through GO enrichment. Twenty acetylation motifs were defined on 3934 unique lysine acetylation sites and motifs *KS*, *KY*, and *KH* occupied the highest proportion of all the identified peptides. Subcellular distribution predictions showed that acetylated proteins in pepper leaves distributed predominantly in chloroplast, cytoplasm and nuclear. KEGG analysis showed 397 identified acetylated proteins were involved in 93 different metabolic pathways. Then the dynamic changes of acetylated proteins in photosynthesis and carbon fixation in photosynthetic organisms pathways under cold stress were further analyzed and many key acetylated proteins regulating cold resistance of pepper were found in these two metabolic pathways. This study is the first to identify the acetylome in pepper, expands greatly the catalog of lysine acetylation substrates and sites in Solanaceae crops and provide a new insight for the post-translational modification study.
Project description:In this experiment, Drosophila melanogaster larvae were fed with Proline and Arginine compounds and showed a better tolerance to freezing compared to control larvae. Using a microarray approach, we investigated the alteration of transcriptome profiles in larvae fed with Proline or Arginine versus control larvae.
Project description:We combined heritability analysis of larval development rate with a global expression analysis of this phenotype to investigate genotype by environment interactions across three ecologically relevant temperatures in the Glanville fritillary butterfly (Melitaea cinxia). We focused upon the development of final instar caterpillars which is greatly affected by temperature, and during this stage the caterpillars build up most of the resources for adult life. Second generation, lab reared larvae, initially collected from the M-CM-^Eland metapopulation, were reared in standard lab condition until 6th larval instar. At the beginning of the final (7th) instar stage the larvae were separated into of three temperature conditions: Cold treatment (temperature profile: 8M-BM-0C 18:00-9:59, 14M-BM-0C 10:00-11:59, 20M-BM-0C 12:00-15:59 and 14M-BM-0C 16:00-17:59) Standard treatment (temperature profile: 15M-BM-0C 17:00-6:59, 18M-BM-0C 7:00-8:59, 22M-BM-0C 9:00-10:59 and 26M-BM-0C 11:00-16:59) Hot treatment (temperature profile: 8M-BM-0C 20:00-7:59, 15M-BM-0C 8:00-9:59, 35M-BM-0C 10:00-17:59 and 15M-BM-0C 18:00-19:59) The temperature profiles mimic the diurnal thermal variation of the natural habitat (M-CM-^Eland islands) of samples. Cold treatment mimics a cool and cloudy summer, Standard represents an average temperature profile in the M-CM-^Eland islands and Hot treatment mimics an exceptionally hot and sunny summer, with cold night-time temperatures. The experiment contained several larval families (full-sib) of which three were selected for gene expression analysis. Samples were snap-frozen in liquid nitrogen during mid-development (after 6, 5 and 4 days, for Cold, Standard and Hot respectively). Additional larvae from the same treatments were assayed for survival and growth. Gene expression was analyzed using a mixed model approach to identify genes with potential heritable expression variation (variation among families), genes with plastic expression responses (treatment induced changes) and genes with treatment dependent expression that varies among families (family by treatment interactions). Full-sib larvae from three families (N170, N74 and O171) were exposed to three temperature treatments (Cold, Standard and Hot) during final (7th) instar stage. A total of 35 samples were used to analyze family, treatment and family by treatment interactions in gene expression, using a mixed model approach. This included 3 biological replicates in Cold and Hot treatments from each family and 5-6 biological replicates in Standard (5 in O171) per family. Techinal replicates from each family and additional techical replicates in family O171 (including dye swap replicates) were used to assess robustness of the findings.
Project description:Gut and rest of body tissue from fifth instar larvae which fed for three days a diet containing different doses of gossypol. Transcriptional profiling comparing gut and rest of body samples for three gossypol concentrations (0%, 0.016%-hormetic dose & 0.16%-detrimental dose). Hormesis is a biphasic biological response characterized by the stimulatory effect at relatively low amounts of chemical compounds otherwise known to be detrimental at higher concentrations. A hormetic response in larval growth rates has been observed in cotton-feeding insects in response to increasing concentrations of gossypol, a toxic metabolite found in the pigment glands of some Malvaceae plants. We investigated the developmental effect of gossypol in the cotton bollworm, Helicoverpa armigera, an important heliothine pest species, by exposing larvae to different doses of this metabolite in their diet. In addition, we sought to determine the underlying transcriptional responses to different gossypol doses. Larval weight gain, pupal weight and larval development time were measured in feeding experiments and a hormetic response was seen for the first two characters. On the basis of net larval weight gain responses to gossypol, three treatments, that is 0%, 0.016% and 0.16% gossypol , were selected for transcript profiling in the gut and the rest of the body in a two-color double reference design microarray experiment. Two-color double reference design. Reference sample was the 0% (CT) gossypol condition (either Gut or Rest of Body) or one of two experimental conditions {0.016(T5) and 0.16%(T7)}. Biological replicates: 4 (10 individuals per replicate). 32 samples total.
Project description:Small noncoding RNA (sncRNA), including microRNAs (miRNAs) and endogenous small-interfering RNAs (endo-siRNAs) are key gene regulators in eukaryotes, playing critical roles in plant development and stress tolerance. Trans-acting siRNAs (ta-siRNAs), which are secondary siRNAs triggered by miRNAs, and siRNAs from natural antisense transcripts (nat-siRNAs) are two well-studied classes of endo-siRNAs. In order to understand sncRNAsM-bM-^@M-^Y roles in plant cold response and stress acclimation, we studied miRNAs and endo-siRNAs in Cassava (Manihot esculenta), a major source of food for the world populations in tropical regions. Combining Next-Generation sequencing and computational and experimental analyses, we profiled and characterized sncRNA species and mRNA genes from the plants that experienced severe and moderate cold stresses, that underwent further severe cold stress after cold acclimation at moderate stress, and that grew under the normal condition. We also included Castor bean (Ricinus communis) to understand conservation of sncRNAs. In addition to known miRNAs, we identified dozens of novel miRNAs as well as ta-siRNA-yielding and nat-siRNA-yielding loci in Cassava and Castor bean, respectively. Among the expressed sncRNAs, many sncRNAs were differentially expressed under cold stresses. Our study provided the results on gene regulation by sncRNAs in cold acclimation of Euphorbiaceous plants and the role of sncRNA-mediated pathways affected by cold stress and stress acclimation in Cassava. Examination of small RNA populations in Cassava cultivar SC124 under the normal condition (NC), gradual cold acclimation (CA), cold shock (CS) and stress acclimation Cold stress after cold acclimation (CCA).
Project description:Abiotic stress is a major environmental factor that limits cotton growth and yield, moreover, this problem has become more and more serious recently and multiple stresses often occur simultaneously due to the global climate change and environmental pollution. We used microarrays to analyze the crosstalk of responsive genes to multiple abiotic stresses including ABA, cold, drought, salinity and alkalinity in cotton. Cotton seedlings with different abiotic stress treatment were selected at 14-day after germination for RNA extraction and hybridization on Affymetrix microarrays. We sought to identify genes involved in diverse stresses including abscisic acid (A), cold (C), drought (M), salinity (N) and alkalinity (P) by comparative microarray analysis (3 biological replicates for each abiotic stress treatment).