- Results: Larvae of Songpu mirror carp and Barbless carp that were pretreated at 18 °C for 24 h significantly. - However, Songpu mirror carp exhibited stronger abilities of cold tolerance and acclimation than Barbless carp. - Transcriptomic profiles of Songpu mirror carp and Barbless carp larvae at 28 °C and 18 °C were compared during cold acclimation through RNA-seq. - Differentially expressed genes that are closely associated with the differences in cold acclimation between two carp species were identified through. - component assembly involved in morphogenesis, secondary alcohol metabolism and drug transport were the most up-regulated biological processes during cold acclimation of Songpu mirror carp. - Conversely, positive regulation of macroautophagy, intracellular protein transport, and organonitrogen compound catabolism were the most down- regulated biological processes during cold acclimation of Barbless carp. - Previous studies have reported that Songpu mirror carp (Cyprinus carpio.) have a strong ability of cold tolerance [37, 38] and Barb- less carp (Cyprinus pellegrini) that specifically live in Xingyun Lake of the Yunnan-Guizhou Plateau in China [39] are sensitive to cold temperature [36]. - Cold tolerance and acclimation of Songpu mirror carp and barbless carp. - Songpu mirror carp and Barbless carp are two closely re- lated species whose larvae rearing at 28 °C exhibited similar developmental phenotypes at 15 dpf (Fig. - Larvae of Songpu mirror carp and Barbless carp at 9 dpf. - We further compared the difference of Songpu mir- ror carp and Barbless carp in cold acclimation. - 1d) (Additional file 1), indicating that both Songpu mirror carp and Barbless carp possess the ability of cold acclimation and Songpu mirror carp has a stronger ability of cold acclimation than Barbless carp.. - A total of twelve cDNA libraries for Songpu mirror carp and Barbless carp were constructed and subjected to high-throughput sequencing, followed by extensive bio- informatics analysis (Fig. - 2b), indicating that Songpu mirror carp and Barbless carp are two closely related species of the carp family that possess different abilities of cold acclimation.. - 1 The difference of Songpu mirror carp and Barbless carp in cold tolerance and cold acclimation. - a Lateral views of Songpu mirror carp (upper) and Barbless carp (below) at 15 dpf. - c Survival rates of Songpu mirror carp and Barbless carp at 12 dpf in AS groups. - d Survival rates of Songpu mirror carp and Barbless carp at 12 dpf in CA groups. - Identification of differentially expressed genes for cold tolerance and acclimation between Songpu mirror carp and barbless carp. - After obtaining counts for annotated genes in two con- trol groups (Ctrl) and cold acclimation groups (CA) of Songpu mirror carp and Barbless carp, we performed a comparative analysis of differentially expressed genes be- tween two of Ctrl and/or CA groups (Fig. - Differen- tially expressed genes in group I represent the developmental and physiological differences between Songpu mirror carp and Barbless carp growing at nor- mal temperature. - II represent the differences between Songpu mirror carp and Barbless carp during cold acclimation. - Differentially expressed genes in group III represent cold-induced and -inhibited genes in Songpu mirror carp. - Next, we performed analysis of differentially genes with Venn’s diagrams to identify potential genes that are specifically required for cold tolerance of Songpu mirror carp and for cold acclimation of Songpu mirror carp and Barbless carp. - 3b and c, differentially expressed genes in group I contain genes that are re- sponsible for the difference of Songpu mirror carp and Barbless carp in cold tolerance to lethal stress (Fig. - Songpu mirror carp Barbless carp. - III (a, b, c, a’, b’, and c’) contain candidate genes that are responsible for the strong ability of cold acclima- tion in Songpu mirror carp. - Some of differentially expressed genes in groups III and IV (d, e, d’ and e’) are involved in cold acclimation of both Songpu mir- ror carp and Barbless carp. - Some of differentially expressed genes in group IV (f, g, f’ and g’) are likely required for cold acclimation of Barbless carp. - These sets of differentially expressed genes were further used for enrichment analysis to explore biological processes and signaling pathways for cold acclimation.. - GO enrichment analysis to identify biological processes for the difference in cold acclimation between Songpu mirror carp and barbless carp. - Differentially expressed genes that are potentially associ- ated with cold acclimation of Songpu mirror carp and Barbless carp were analyzed with GO enrichment and displayed in Additional file 6 and representatives of GO term belonged to the biological process obtained through REVIGO tool were displayed in Additional file 7.. - Candidate genes up-regulated in groups III and IV (d and e) during cold acclimation of both Songpu mirror carp and Barbless carp were mainly enriched in bio- logical processes including lipid homeostasis, cellular biosynthesis, lipid transport, response to endogenous stimulus and other processes (Fig. - 3 Identify genes involved in cold acclimation of Songpu mirror carp and Barbless carp. - Genes involved in cold acclimation of Songpu mirror carp and Barbless carp were identified through Venn ’ s diagrams analysis of up-regulated genes (B) and down-regulated genes (C). - encode specific factors for the formation of cold acclimation in Songpu mirror carp, and those in group IV (f, g, f ’ and g. - encode specific factors for the formation of cold acclimation in Barbless carp. - Common factors required for the formation of cold acclimation of two carp species are encoded by differentially expressed genes that are overlapped in group III and IV (d, e, d ’ and e. - during cold acclimation of both Songpu mirror carp and Barbless carp were overrepresented in biological pro- cesses such as protein catabolism, regulation of RNA splicing, striated muscle tissue development and primary metabolism (Fig. - 5 Heat maps from KEGG enrichment analysis of genes associated with cold acclimation in Songpu mirror carp and Barbless carp. - were enriched in signaling pathways that are potentially required for the formation of cold acclimation in Songpu mirror carp. - were enriched in signaling pathways that are likely important for the formation of cold acclimation in Barbless carp. - were enriched in signaling pathways that are potentially required for the formation of cold acclimation in two carp species. - Candidate genes down- regulated in groups III and IV (f’ and g’) during cold acclimation of Barbless carp were overrepresented in biological processes including positive regulation of macroautophagy, organonitrogen compound catabolism, intracellular protein transport, protein deubiquitination and other processes (Fig. - KEGG enrichment analysis to identify signaling pathways for cold acclimation difference between Songpu mirror carp and barbless carp. - Differentially expressed genes that are potentially associ- ated with cold acclimation of Songpu mirror carp and Barbless carp were analyzed with the KEGG pathway en- richment. - and c’) during the formation of strong cold acclimation in Songpu mirror carp were mainly overrepresented in signaling pathways such as protein processing in endo- plasmic reticulum, antigen processing and presentation and amino sugar and nucleotide sugar metabolism (Fig. - Candidate genes up-regulated in groups III and IV (d and e) during cold acclimation of both Songpu mirror carp and Barbless carp were enriched in signaling path- ways including cholesterol metabolism, PPAR signaling pathway and spliceosome (Fig. - and e’) during cold acclimation of both Songpu mirror carp and Barbless carp were mainly overrepresented in. - Candidate genes up-regulated in groups III and IV (f and g) during cold acclimation of Barbless carp were mainly enriched in signaling pathways such as insulin signaling pathway, PPAR signaling pathways, adipocyto- kine signaling pathway and ribosome biogenesis in eukaryotes (Fig. - Candidate genes down-regulated in groups III and IV (f’ and g’) during cold acclimation of Barbless carp were overrepre- sented in signaling pathways of thermogenesis, cellular senescence and oxidative phosphorylation (Fig. - FoxO signaling pathway played a key role in the difference of cold acclimation between Songpu mirror carp and barbless carp. - Regulation of autophagy was down-regulated by cold-inhibited BNIP3 (Bcl-2/adenovirus E1B 19-kDa interacting pro- tein) expression in groups III and IV (d’ and e’ in gray) of both Songpu mirror carp and Barbless carp (Fig. - During the cold acclimation of Barbless carp, the AMPK. - Next, inhibitors of FOXO (AS1842856), AMPK (Dorsomorphin 2HCl) and JNK (SP600125) and activator of Akt (SC79) were used to further verify the function of FoxO signaling in the formation of cold acclimation of Songpu mirror carp following a strategy in Fig. - Moreover, the survival rate of Songpu mirror carp was significantly reduced after activation of Akt signaling by SC79. - 6 Signaling pathways associated with cold acclimation of two carp species activated in the FoxO signaling pathway. - Taken together, these data suggest that FoxO signaling played a crucial role in cold acclimation of two carp spe- cies and distinctive signaling pathways upstream of FOXO activation contributed to the difference of cold acclimation in Songpu mirror carp and Barbless carp.. - Songpu mirror carp and Barbless carp are two closely related species of the carp family since they can hybrid to generate the off- spring [39]. - identified many differentially expressed genes in Songpu mirror carp and Barbless carp during acute cold stress and acclimation. - Moreover, FoxO-related signaling pathways appear to play crucial roles in the formation of strong cold toler- ance and acclimation of Songpu mirror carp.. - In addition to Songpu mirror carp and Barbless carp, or- ganisms including zebrafish [33, 34], Caenorhabditis ele- gans [43, 44] and plants [45] possess a cold acclimation ability to allow their survival in extremely low temperature. - Thus, the relatively low level of cholesterol content is likely associated with the strong cold acclimation of Songpu mirror carp.. - Consistent with one of our previous studies during cold acclimation of zebrafish [33], we found the gene of hmgb3 (high mobility group protein B3) and cirbpb (cold-inducible RNA-binding protein B) was up- regulated by 2.7- and 2.8- folds in Songpu mirror carp, and by 3.1- and 3.5-folds in Barbless carp during cold acclimation (Additional file 5). - Another two genes (cry2 and nr1d2) that are related to circadian clocks were also induced during cold acclimation of Songpu mirror carp and Barbless carp. - characterization of cold acclimation responses in differ- ent fish species.. - 7 Verification of FoxO-related signaling pathways involved in cold acclimation of Songpu mirror carp. - Larvae of Songpu mirror carp were pre-treated at 18 °C with or without inhibitors or activator for 24 h and then moved to severe low. - b Cold acclimation of Songpu mirror carp was significantly affected by inhibition or activation of key signaling factors acting upstream of FoxO signaling. - Larvae of Songpu mirror carp were treated with inhibitors (AS1842856 for FOXO at 5 μ M, dorsomorphin 2HCl for AMPK at 10 μ M or SP600125 for JNK at 20 μ M) or an activator (SC79 for Akt at 4 μ M) for 24 h. - d A working model for the roles of FoxO-related signaling pathways in cold acclimation of carps. - with the cold acclimation of both Songpu mirror carp and Barbless carp.. - In this study, the representative of enriched GO terms in groups III and IV (d and e) include lipid homeostasis (GO:0055088), lipid transport (GO:0006869), and lipid metabolism (GO:0006629) (Figure S3), indicating that the changes in lipid homeostasis plays an important role in the cold acclimation of both Songpu mirror carp and Barbless carp.. - We also found some biological processes that are spe- cifically enriched during the cold acclimation of Songpu mirror carp. - These findings suggest that the formation of a strong cold acclimation in Songpu mirror carp is closely associated with changes in mul- tiple tissues.. - It is likely that the limitation of these biological processes is mainly contributed to the weak ability of cold acclimation in Barbless carp.. - Signal pathways that were significantly activated dur- ing the cold acclimation of both Songpu mirror carp and Barbless carp include insulin signaling pathway, choles- terol metabolism, PPAR signaling pathway, and spliceo- some (Fig. - transport and lipid metabolism were highly enriched during the cold acclimation of both Songpu mirror carp and Barbless carp (Fig. - Thermogenesis is activated during the cold acclimation of mice however, thermogenesis is inhibited in Barbless carp but not in the Songpu mirror carp during the cold acclimation (Fig. - Further investigations are need to examine the contribution of PPAR signaling pathway and thermogenesis to a strong cold acclimation of Songpu mirror carp.. - Therefore, it is likely that FoxO-related signaling pathways play a key role in cold acclimation of Songpu mirror carp. - In this study, Songpu mirror carp was found to have stronger abilities of cold tolerance and acclimation than Barbless carp. - High-throughput RNA-seq analysis have identified many of differentially expressed genes, biological processes and signaling pathways between Songpu mirror carp and Barbless carp during cold accli- mation. - Moreover, FoxO- related signaling pathways appear to play crucial roles in the formation of strong cold tolerance and acclimation of Songpu mirror carp. - Fertilized eggs of Songpu mirror carp and Barbless carp were obtained from Heilongjiang River Fishery Research Institute of Chinese Academy of Fishery Sciences. - To compare differences in cold tolerance and acclima- tion between Songpu mirror carp and Barbless carp,. - Larvae at 8 dpf of Songpu mirror carp and Barbless carp were kept in dark throughout the experiment to avoid the influence of light exposure on gene expres- sion [34]. - Twenty larvae at 10 dpf of Songpu mirror carp and Barbless carp growing at 28 °C were sampled in triplets and served as controls (Ctrl). - Twenty larvae at 9 dpf of Songpu mirror carp and Barbless carp grow- ing at 28 °C were subjected to cold acclimation at 18 °C for 24 h, sampled in triplets at 10 dpf and served as cold acclimation groups (CA). - Larvae at 9 dpf of Songpu mirror carp were treated with or without inhibitors or activators of FoxO signaling pathway during cold acclimation at 18 °C for 24 h, col- lected and lysed in cell lysis buffer for Western and IP (Beyotime, P0013J) containing 1% protease inhibitors (Protease Inhibitor Cocktail, Bimake) and 1% phosphat- ase inhibitors (Phosphatase Inhibitor Cocktail, Bimake).. - Survival rates of Songpu mirror carp and Barbless carp under cold stress.. - CA-cold acclimation. - CA: cold acclimation. - CA: cold acclimation;. - Cold acclimation alters the connective tissue content of the zebrafish (Danio rerio) heart:. - Thyroid hormone regulates cardiac performance during cold acclimation in zebrafish (Danio rerio). - Thyroid hormone regulates muscle function during cold acclimation in zebrafish (Danio rerio). - Transcriptomic characterization of cold acclimation in larval zebrafish. - Inheritance of growth traits in Songpu mirror carp ( Cyprinus carpio L.) cultured in Northeast China. - Effects of dietary protein and temperature on growth and flesh quality of Songpu Mirror carp
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