Global Phosphoproteomic Analysis Reveals Significant Metabolic Reprogramming in the Termination of Liver Regeneration in Mice
Global Phosphoproteomic Analysis Reveals Significant Metabolic Reprogramming in the Termination of Liver Regeneration in Mice
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全球磷酸化蛋白质组学分析揭示小鼠肝脏再生终止过程中的显着代谢重编程
DOI：
10.1021/acs.jproteome.0c00028
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发表时间：
2020
影响因子：
4.4
通讯作者：
Fang Lei
中科院分区：
文献类型：
--
作者：
Zhang Jingzi;Tang Neng;Zhao Yinjuan;Zhao Ruoyu;Fu Xiao;Zhao D;an;Zhao Yue;Huang Lan;Li Chaojun;Qin Yudong;Xue Bin;Fang Lei
磷酸化在调节各种生物过程中至关重要。然而，全面的磷酸化蛋白质组学分析在肝再生（LR）的终止仍然缺失。在这里，我们使用串联质量标签（TMT）标记加上磷酸肽富集和二维（2D）液相色谱-质谱（LC-MS）/MS分析，以建立一个全球磷酸蛋白质组学地图在小鼠肝脏部分肝切除术（PH）后第5天。共鉴定了3443种蛋白质的9731个磷酸化位点，并对2980种蛋白质的7802个磷酸化位点进行了定量。识别的磷酸化位点的基序分析揭示了一系列不同的共有序列，表明多个激酶家族，包括ERK/MAPK，PKA/PKC，CaMK-II，CKII和CDK可能参与LR的终止。功能聚类分析显示，磷酸化位点异常的蛋白质主要参与代谢途径、DNA复制和紧密连接。更重要的是，肝脏中PP 2Ac α的缺失显著改变了整体磷酸化谱，表明其在调节LR终止中的关键作用。最后，通过免疫共沉淀和Western blot验证了几个差异磷酸化位点。总之，我们的数据揭示了小鼠LR终止的第一个全面的磷酸化蛋白质组学图谱，这极大地扩展了我们对这一过程复杂调控的认识，并为使用肝切除治疗肝癌提供了新的方向。
Phosphorylation is crucial in regulating various biological processes. However, comprehensive phosphoproteomic profiling in the termination of liver regeneration (LR) is still missing. Here, we used Tandem Mass Tag (TMT) labeling coupled with phosphopeptide enrichment and two-dimensional (2D) liquid chromatography–mass spectrometry (LC–MS)/MS analysis to establish a global phosphoproteomic map in the liver of mice at day 5 after partial hepatectomy (PH). Altogether, 9731 phosphosites from 3443 proteins were identified and 7802 phosphosites from 2980 proteins were quantified. Motif analysis of the identified phosphosites revealed a diverse array of consensus sequences, suggesting that multiple kinase families including ERK/MAPK, PKA/PKC, CaMK-II, CKII, and CDK may be involved in the termination of LR. Functional clustering analysis of proteins with dysregulated phosphosites showed that they mainly participate in metabolic pathways, DNA replication, and tight junction. More importantly, the deletion of PP2Acα in the liver remarkably changes the overall phosphorylation profile, indicating its critical role in regulating the termination of LR. Finally, several differentially phosphorylated sites were validated by co-immunoprecipitation and Western blot. Taken together, our data unravel the first comprehensive phosphoproteomic map in the termination of LR in mice, which greatly expands our knowledge in the complicated regulation of this process and provides new directions for the treatment of liver cancer using liver resection.