Embryonic incubation temperature has a long-term effect on the microRNA transcriptome of adult zebrafish
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ABSTRACT: Zebrafish eggs were collected at 28 °C, and split into three temperature groups (24 °C, 28 °C, 32 °C) for incubation. When larvae reaching first feeding, the incubation temperature of larvae from 24 °C and 32 °C was gadually changed to 28 °C, and all fish were kept at 28 °C for later development until adulthood. At 100 d post-fertilization, seven replicates from each group were intraperitoneally (i.p.) injected with 2 μl of 50 mg/ml lipopolysaccharide (LPS) from Pseudomonas aeruginosa 10, while another seven replicates were i.p. injected with 2 μl phosphate-buffered saline as control. At 12 h post-injection, the spleen was dissected, and total RNA was isolated. Five LPS-treated replicates and five controls were used for building small RNA libraries and sequencing. A total of 112 novel miRNAs were identified using miRDeep2. By comparing fish from different embryonic incubation tmeperatures, 32 miRNAs (29 up-/3 down-regulated) were identified differentially expressed (DEmiRs, adjusted p-value < 0.05, |fold change| > 2.0, DESeq2) in fish from embryonic incubation temperature of 32 °C compared to fish kept at constant 28 °C, 3 DEmiRs (2 up-/1 down-regulated) were identified in response to LPS in fish kept at 28 °C. A total of 9,116 target genes were predicted using miRanda (v3.3a), including 8,224 genes targeted by 32 DE miRNAs and 892 genes targeted by 3 DE miRNAs.
Project description:Zebrafish eggs were collected at 28 °C, and split into three embryonic incubation temperature groups (24 °C, 28 °C, 32 °C). When larvae reaching first feeding, the incubation temperature of larvae from 24 °C and 32 °C was gradually changed to 28 °C, and all fish were kept at 28 °C for later development until adulthood. At 100 d post-fertilization, 7 replicates from each temperature group were intraperitoneally (i.p.) injected with 2 μl of 50 mg/ml lipopolysaccharide (LPS) from Pseudomonas aeruginosa 10, while another 7 replicates were i.p. injected with 2 μl phosphate-buffered saline as control. At 12 h post-injection, the spleen was dissected, and total RNA was extracted. Five LPS-treated replicates and five controls were used for building RNA libraries and sequencing. The differential gene expression analysis were performed between fish from different embryonic incubation temperatures (24 °C vs 28 °C; 32 °C vs 28 °C), and between LPS treatment and control within each temperature group (24 °C LPS vs control; 28 °C LPS vs control; 32 °C LPS vs control). Totally, 251 differentially expressed genes (DEGs, 71 up-/180 down-regulated) were identified in fish from 24 °C embryonic incubation temperature compared to fish kept at constant 28 °C (DESeq2, adjusted p-value < 0.05, |fold change| > 1.5); and 660 DEGs (385 up-/275 down-regulated) were identified in fish from 32 °C embryonic incubation temperature compared to fish kept at 28 °C. By comparing LPS-treated fish to control, 567 DEGs (271 up-/296 down-regulated) were identified in fish from embryonic incubation temperature of 24 °C, 140 DEGs (80 up-/60 down-regulated) were identified in fish kept at constant 28 °C; and 49 DEGs (11 up-/38 down-regulated) were identified in fish from the 32 °C embryonic incubation temperature group.
Project description:We report the effect of incubation temperature on embryos and larvae of Solea senegalensis subjected to two different incubation temperatures (15 °C or 21 °C). We found that at some stages, a higher incubation temperature was associated with the expression of miRNAs positively related with growth
Project description:Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) is an inflammatory process of the lungs characterized by increased permeability of the alveolar-capillary membrane with subsequent interstitial/alveolar edema and diffuse alveolar damage. ALI/ARDS can be the results of either direct or indirect lung injury, with pneumonia being the most common direct pulmonary insult and sepsis the most common extra-pulmonary cause. In this study, we employed the murine lipopolysaccharide (LPS)-induced direct and indirect lung injury model to explore the pathogenic mechanisms of pulmonary and extra-pulmonary ARDS, using an unbiased, discovery and quantitative proteomic approach. A total of 1,017 proteins were both identified and quantified in bronchoalveolar lavage fluid (BALF) from control, intratracheal LPS (I.T. LPS, 0.1 mg/kg) and intraperitoneal LPS (I.P. LPS, 5 mg/kg) treated mice. The two LPS groups shared 13 up-regulated and 22 down-regulated proteins compared to the control group. Among them, molecules related to bronchial and type II alveolar epithelial cell functions including cell adhesion molecule 1 and surfactant protein B were reduced, whereas lactotransferrin and resistin like alpha involved in lung innate immunity were upregulated in both LPS groups. Proteomic profiling also identified significant differences in BALF proteins between I.T. and I.P. LPS groups. Ingenuity pathway analysis revealed that acute-phase response signaling was activated by both I.T. and I.P. LPS, however, the magnitude of activation is much greater in I.T. LPS group compared to I.P. LPS group. Intriguingly, two canonical signaling pathways, liver X receptor/retinoid X receptor activation and the production of nitric oxide and reactive oxygen species in macrophages, were activated by I.T. LPS but suppressed by I.P. LPS. In addition, CXCL15 (also known as lungkine) was also up-regulated by I.T LPS but down-regulated by I.P. LPS. In conclusion, our quantitative discovery-based proteomic approach identified commonalities as well as significant differences in BALF protein expression profiles in LPS-induced direct and indirect lung injury, and importantly, LPS-induced indirect lung injury results in suppression of select components of lung innate immunity, which could contribute to the so-called “immunoparalysis” in sepsis patients.
Project description:The cold stress is considered the bottleneck problem limiting the development of red tilapia industry. Fish (initial weight: 72.71±1.32 g) was divided into the cold stressed group (C) and the control normal water temperature group (N). The water temperature was reduced by 2℃ per day from 20℃ to 8℃ in cold group, and the control group kept the water temperature at 20℃ during the experiment. At the end of the experiment, the gill(G)、liver(L) and skin(S) tissues from the fish of cold(C) group and control normal water temperature (N) group were taken. The DIA proteome analysis of 18 samples was completed, and 6341 trusted proteins containing quantitative information were obtained. Our aim is to understand the protein expression profiles of cold stress in red tilapia.
Project description:LPS (10 μg/50 μl PBS; E. coli Serotype 055:B5, Sigma-Aldrich) was administered onto the nares. Baseline levels of genes in mice treated with vehicle (Hanks Balanced Salt solution (HBSS) and PBS), Intranasal S100s (10 μg/50 μl HBSS) were given 2 h before LPS; control mice received equal volumes of PBS and HBSS. Mice were sacrificed 4 h post LPS inhalation.
Project description:Here we analyzed the proteomic response of the important Gram-negative fish pathogen A. salmonicida to iron-limitation, the antibiotic florfenicol and an elevated incubation temperature. Proteins of different subcellular fractions (cytosol, inner membrane, outer membrane, extracellular and outer membrane vesicles) were enriched and analyzed.
Project description:Two experiments were preformed, one for evaluating the effect of E2 exposure in males and a second experiment for comparing gene expression in the liver of non-vitellogenic females with that of vitellogenic females. The experiments designs were as follows:<br>1) Four months old zebrafish were divided into three groups consisting of 8 fish in each group: i) males (N=8) were exposed to E2 (Sigma-Aldrich, Israel) by immersion for 48 h (group E). The concentration used was 5 ?g/L (18 nM), as 3-4 ng/ml was determined to be the E2 natural concentration in the plasma of adult vitellogenic female ZF (Heiden et al 2006); ii) untreated males(N=8); iii) untreated vitellogenic females(N=8). Four replicate samples were prepared for each group and each replicate consisted of a pooled sample from livers of two fish. <br>2) In order to reveal the differences between vitellogenic and non-vitellogenic females, a second experiment was designed. One month old zebrafish were divided into two groups consisting of 32 fish in each group. Due to the small size of the liver, pooled samples from eight fish were prepared for each of the four replicates in the expression studies. The groups consisted of: i) fish that were kept under a light/dark cycle of 14/10 h for 5 weeks (N=32). with ovaries in vitellogenic stage; ii) fish that were kept under a light/dark cycle of 6/18 h for the same time period (N=32) with non-vitellogenic ovaries.
Project description:Low temperatures may cause severe growth inhibition and mortality in fish. In order to understand the mechanism of cold tolerance, a transgenic zebrafish Tg (smyd1:m3ck) model was established to study the effect of energy homeostasis during cold stress. The muscle-specific promoter Smyd1 was used to express the carp muscle form III of creatine kinase (M3-CK), which maintained enzymatic activity at a relatively low temperature, in zebrafish skeletal muscle. In situ hybridization showed that M3-CK was expressed strongly in the skeletal muscle. When exposed to 13°C, Tg (smyd1:m3ck) fish maintained their swimming behavior, while the wild-type could not. Energy measurements showed that the concentration of ATP increased in Tg (smyd1:m3ck) versus wild-type fish at 28°C. After 2 h at 13°C, ATP concentrations were 2.16-fold higher in Tg (smyd1:m3ck) than in wild-type (P < 0.05). At 13°C, the ATP concentration in Tg (smyd1:m3ck) fish and wild-type fish was 63.3% and 20.0%, respectively, of that in wild-type fish at 28°C. Microarray analysis revealed differential expression of 1249 transcripts in Tg (smyd1:m3ck) versus wild-type fish under cold stress. Biological processes that were significantly overrepresented in this group included circadian rhythm, energy metabolism, lipid transport, and metabolism. These results are clues to understanding the mechanisms underlying temperature acclimation in fish.
Project description:In the present study, the eggs of Chinese pond turtles (Mauremys reevesii) were incubated at three temperatures (26℃, 29℃ and 32℃). During the thermosensitive period (TSP) of incubation, the adrenal-kidney-gonad (AKG) complexes were sampled, and the transcriptome differences of the AKG samples were investigated using next-generation sequencing, then candidate genes associated with temperature-dependent sex determination of Reeves' Turtle were identified.
Project description:Streptococcus agalactiae is one of the most important pathogens associated with outbreaks of streptococcis in Nile tilapia farms around the world. High water temperature (above 27°C) have been described as factor predisposing for disease in fish. On the other hand, at low temperature (below 25°C) fish mortalities are no usually observed in farms. The temperature variation can modulate the expression of genes and proteins involved with metabolism, adaptation and bacterial pathogenicity, increasing or decreasing the host susceptibility to infection. The aim of this study was to evaluate the transcriptome and proteome of fish-pathogenic S. agalactiae strain (SA53) submitted to in vitro growth under different temperatures using microarray and label-free shotgun LC-HDMSE approach, and to compare the expression trends of proteins shared among GBS strains from different hosts (SA53 and NEM316). Biological triplicates of isolates were cultured in BHIT broth at 22°C or 32°C for RNA and protein isolation and submitted to transcriptomic and proteomic analysis. Total of 1730 transcripts were identified in SA53, being 107 genes differentially expressed among the temperature evaluated. A higher number of genes related with metabolism were detected as up-regulated proteins at 32°C, mainly PTS system and ABC transport system. In proteome analysis, 1046 proteins were identified in SA53 strain, being 81 proteins differentially regulated at 22 and 32ºC. Proteins involved in Defense mechanisms (V), Lipid transport and metabolism (I), and Nucleotide transport and metabolism (F) were up-regulated at 32ºC. A higher number of interactions was observed in the category F. The induction of genes/proteins involved in virulence were detected in both temperatures evaluated. A low correlation between transcriptome and proteome datasets was observed. And there is a distinct adaptation between fish and human GBS strains at the proteome level. Our study showed that the transcriptome and proteome of fish-adapted GBS strain are modulated by temperature, especially regulating the differential expression of genes/proteins involved with metabolism, adaptation and virulence, and revealing a host specificity at proteome regulation for human and fish hosts