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利用 iTRAQ 联合 LC-MS/MS 技术对感染猪德尔塔冠状病毒的猪小肠上皮细胞进行定量蛋白质组学分析。

Quantitative Proteomic Analysis of Porcine Intestinal Epithelial Cells Infected with Porcine Deltacoronavirus Using iTRAQ-Coupled LC-MS/MS.

机构信息

Key Laboratory of Animal Epidemiology of Ministry of Agriculture and Rural Affairs, College of Veterinary Medicine, China Agricultural University, No. 2 Yuanmingyuan West Road, Haidian District, Beijing 100193, PR China.

出版信息

J Proteome Res. 2020 Nov 6;19(11):4470-4485. doi: 10.1021/acs.jproteome.0c00592. Epub 2020 Oct 12.

Abstract

Porcine deltacoronavirus (PDCoV) is an emergent enteropathogenic coronavirus associated with swine diarrhea. Porcine small intestinal epithelial cells (IPEC) are the primary target cells of PDCoV infection in vivo. Here, isobaric tags for relative and absolute quantification (iTRAQ) labeling coupled to liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to quantitatively identify differentially expressed proteins (DEPs) in PDCoV-infected IPEC-J2 cells. A total of 78 DEPs, including 23 upregulated and 55 downregulated proteins, were identified at 24 h postinfection. The data are available via ProteomeXchange with identifier PXD019975. To ensure reliability of the proteomics data, two randomly selected DEPs, the downregulated anaphase-promoting complex subunit 7 (ANAPC7) and upregulated interferon-induced protein with tetratricopeptide repeats 1 (IFIT1), were verified by real-time PCR and Western blot, and the results of which indicate that the proteomics data were reliable and valid. Bioinformatics analyses, including GO, COG, KEGG, and STRING, further demonstrated that a majority of the DEPs are involved in numerous crucial biological processes and signaling pathways, such as immune system, digestive system, signal transduction, RIG-I-like receptor, mTOR, PI3K-AKT, autophagy, and cell cycle signaling pathways. Altogether, this is the first study on proteomes of PDCoV-infected host cells, which shall provide valuable clues for further investigation of PDCoV pathogenesis.

摘要

猪德尔塔冠状病毒(PDCoV)是一种新兴的肠致病性冠状病毒,与猪腹泻有关。猪小肠上皮细胞(IPEC)是 PDCoV 感染体内的主要靶细胞。在这里,采用相对和绝对定量标记(iTRAQ)联合液相色谱-串联质谱(LC-MS/MS)技术定量鉴定 PDCoV 感染的 IPEC-J2 细胞中的差异表达蛋白(DEPs)。在感染后 24 小时,共鉴定出 78 个 DEPs,包括 23 个上调和 55 个下调蛋白。该数据可通过 ProteomeXchange 以标识符 PXD019975 获得。为确保蛋白质组学数据的可靠性,随机选择了两个 DEPs,下调的有丝分裂促进复合物亚基 7(ANAPC7)和上调的干扰素诱导蛋白具有四肽重复 1(IFIT1),通过实时 PCR 和 Western blot 进行了验证,结果表明蛋白质组学数据是可靠和有效的。生物信息学分析,包括 GO、COG、KEGG 和 STRING,进一步表明大多数 DEPs 参与了许多关键的生物过程和信号通路,如免疫系统、消化系统、信号转导、RIG-I 样受体、mTOR、PI3K-AKT、自噬和细胞周期信号通路。总之,这是首次对 PDCoV 感染宿主细胞的蛋白质组进行研究,为进一步研究 PDCoV 的发病机制提供了有价值的线索。

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