College of Life Sciences, College of Marine Science and Engineering, Nanjing Normal University, Nanjing 210023, China; Co-Innovation Center for Marine Bio-Industry Technology of Jiangsu Province, Lianyungang, Jiangsu 222005, China.
College of Life Sciences, College of Marine Science and Engineering, Nanjing Normal University, Nanjing 210023, China; Co-Innovation Center for Marine Bio-Industry Technology of Jiangsu Province, Lianyungang, Jiangsu 222005, China.
J Proteomics. 2019 Jun 15;201:27-36. doi: 10.1016/j.jprot.2019.04.004. Epub 2019 Apr 5.
Low temperatures profoundly influence the physiological and behavioural processes of ectotherms, especially teleosts, which have made them the subjects of strong interest over time. However, the characteristics of fish cold-tolerance at the protein level remain unclear. Therefore, to shed further light on the molecular mechanisms of low temperature adaptation in fish, we conducted quantitative proteomics on the T. fasciatus liver using iTRAQ. Comparing the proteomic profiles of the T. fasciatus liver at 12 °C and 26 °C, a total of 3741 proteins were identified, and 160 were differentially abundant proteins (DAPs). Among the DAPs, the most significant changes were noted in proteins involved in oxidative stress (nine proteins), mitochondrial enzymes (eleven proteins) and signal transduction (thirteen proteins). The KEGG enrichment analysis indicated significant enhancement of D-arginine and D-ornithine metabolism, MAPK signalling, Wnt signalling and Gap junction pathway. Subsequently, three significantly up-regulated proteins (CIRB, HSP90 and GST) and two significantly down-regulated proteins (FLNB and A2ML1) were validated with parallel reaction monitoring (PRM) assays. Furthermore, the changes in abundance of proteins that are involved in oxidative stress, mitochondrial enzymes and signal transduction were validated at the transcriptional level with qPCR. These verification results show that the experimental data of iTRAQ are reliable. Our results not only deepen the understanding of the mechanisms underlying low-temperature tolerance in fish, but they also may contribute to the enhancement of cold tolerance during its breeding process. SIGNIFICANCE OF THE STUDY: The study focused on a comparative quantitative proteomics analysis of the T. fasciatus liver in response to low temperatures using iTRAQ, which has not yet been reported in the literatures. The results showed that the effect of low temperature on T. fasciatus is significant, including a detoxification of metabolic by-products and oxidative stress, an activation of the mitochondrial enzyme to strengthen energy metabolism, and a negative effect on signal transduction, which result in dysfunction or suboptimal performance. These low-temperature-related changes in the liver proteome of T. fasciatus can facilitate the understanding of the low temperature-related response that takes place in similar conditions in the liver and may contribute to the breeding of cold-resistant strains.
低温深刻影响着变温动物（尤其是硬骨鱼类）的生理和行为过程，这使得它们成为研究热点。然而，鱼类在蛋白质水平上的耐寒特性仍不清楚。因此，为了进一步阐明鱼类适应低温的分子机制，我们使用 iTRAQ 对 T. fasciatus 肝脏进行了定量蛋白质组学研究。比较 T. fasciatus 肝脏在 12°C 和 26°C 时的蛋白质组图谱，共鉴定出 3741 种蛋白质，其中 160 种为差异丰度蛋白（DAPs）。在这些 DAPs 中，参与氧化应激（9 种蛋白质）、线粒体酶（11 种蛋白质）和信号转导（13 种蛋白质）的蛋白质变化最为显著。KEGG 富集分析表明，D-精氨酸和 D-鸟氨酸代谢、MAPK 信号通路、Wnt 信号通路和间隙连接途径显著增强。随后，用平行反应监测（PRM）实验验证了 3 个显著上调的蛋白（CIRB、HSP90 和 GST）和 2 个显著下调的蛋白（FLNB 和 A2ML1）。此外，用 qPCR 验证了与氧化应激、线粒体酶和信号转导相关的蛋白质丰度的变化在转录水平上的变化。这些验证结果表明，iTRAQ 的实验数据是可靠的。我们的研究结果不仅加深了对鱼类耐低温机制的理解，而且可能有助于在其繁殖过程中提高耐寒性。研究意义：本研究采用 iTRAQ 对 T. fasciatus 肝脏在低温下的蛋白质组进行了比较定量蛋白质组学分析，这在文献中尚未报道。结果表明，低温对 T. fasciatus 的影响显著，包括代谢产物解毒和氧化应激的减轻、线粒体酶的激活以增强能量代谢、以及对信号转导的负面影响，导致其功能障碍或表现不佳。这些 T. fasciatus 肝脏蛋白质组与低温相关的变化有助于理解在类似条件下肝脏发生的与低温相关的反应，并可能有助于耐寒品系的选育。
