Temperature affects almost all aspects of the fish life. To cope with low temperature, fish have evolved the ability of cold acclimation for survival. However, intracellular signaling events underlying cold acclimation in fish remain largely unknown. Here, the formation of cold acclimation in zebrafish embryonic fibroblasts (ZF4) is monitored and the phosphorylation events during the process are investigated through a large-scale quantitative phosphoproteomic approach. In total, 11 474 phosphorylation sites are identified on 4066 proteins and quantified 5772 phosphosites on 2519 proteins. Serine, threonine, and tyrosine (Ser/Thr/Tyr) phosphorylation accounted for 85.5%, 13.3%, and 1.2% of total phosphosites, respectively. Among all phosphosites, 702 phosphosites on 510 proteins show differential regulation during cold acclimation of ZF4 cells. These phosphosites are divided into six clusters according to their dynamic changes during cold exposure. Kinase-substrate prediction reveals that mitogen-activated protein kinase (MAPK) among the kinase groups is predominantly responsible for phosphorylation of these phosphosites. The differentially regulated phosphoproteins are functionally associated with various cellular processes such as regulation of actin cytoskeleton and MAPK signaling pathway. These data enrich the database of protein phosphorylation sites in zebrafish and provide key clues for the elucidation of intracellular signaling networks during cold acclimation of fish.

中文翻译：

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斑马鱼胚胎成纤维细胞在冷驯化过程中的动态磷酸化蛋白质组学分析。

温度几乎影响鱼类生活的各个方面。为了应对低温，鱼类已经进化出冷驯化的生存能力。然而，在鱼类中冷适应的细胞内信号转导事件仍然是未知的。在这里，斑马鱼胚胎成纤维细胞（ZF4）的冷驯化的形成受到监控，并通过大规模定量磷酸化蛋白质组学方法研究了该过程中的磷酸化事件。总共在4066个蛋白质上鉴定了11 474个磷酸化位点，在2519个蛋白质上鉴定了5772个磷酸化位点。丝氨酸，苏氨酸和酪氨酸（Ser / Thr / Tyr）磷酸化分别占总磷酸酯位的85.5％，13.3％和1.2％。在所有磷酸酯中，510蛋白上的702个磷酸酯在ZF4细胞冷驯化过程中显示出不同的调节作用。根据它们在冷暴露期间的动态变化，将这些磷酸酯分为六个簇。激酶底物的预测表明，激酶组中的促分裂原活化蛋白激酶（MAPK）主要负责这些磷酸位的磷酸化。差异调节的磷蛋白在功能上与各种细胞过程相关，例如肌动蛋白细胞骨架和MAPK信号通路的调节。这些数据丰富了斑马鱼中蛋白质磷酸化位点的数据库，并为阐明鱼类冷适应过程中的细胞内信号网络提供了重要线索。激酶底物的预测表明，激酶组中的促分裂原活化蛋白激酶（MAPK）主要负责这些磷酸位的磷酸化。差异调节的磷蛋白在功能上与各种细胞过程相关，例如肌动蛋白细胞骨架和MAPK信号通路的调节。这些数据丰富了斑马鱼中蛋白质磷酸化位点的数据库，并为阐明鱼类冷适应过程中的细胞内信号网络提供了重要线索。激酶底物的预测表明，激酶组中的促分裂原活化蛋白激酶（MAPK）主要负责这些磷酸位的磷酸化。差异调节的磷蛋白在功能上与各种细胞过程相关，例如肌动蛋白细胞骨架和MAPK信号通路的调节。这些数据丰富了斑马鱼中蛋白质磷酸化位点的数据库，并为阐明鱼类冷适应过程中的细胞内信号网络提供了重要线索。