基于TMT技术对低氧与常氧条件下牦牛PASMCs的蛋白组学分析

doi:10.11843/j.issn.0366-6964.2022.07.015

摘要/Abstract

摘要： 旨在利用串联质谱标签(tandem mass tag,TMT)技术对牦牛肺动脉平滑肌细胞(pulmonary arterial smooth muscle cells,PASMCs)在低氧和常氧不同培养条件下的蛋白组学进行差异表达蛋白(differentially expressed proteins,DEPs)分析，进而了解高原牦牛PASMCs的DEPs。本试验采用贴壁培养法培养PASMCs，分别在低氧与常氧条件下培养72 h，然后收集细胞，提取两组细胞的总蛋白，利用TMT标记蛋白定量技术筛选DEPs进行分析，并进行GO功能注释和KEGG通路分析。在比较低氧和常氧下PASMCs的2组蛋白样品中共获得6 859个蛋白，以FC (fold change)≥1.3或≤0.78，且P<0.05条件进行筛选，获得DEPs共531个，其中上调DEPs有186个，下调DEPs有345个。富集分析结果表明，DEPs主要富集在中枢碳代谢、糖酵解与糖代谢合成、HIF-1信号通路、次生代谢产物的生物合成等低氧适应相关通路中。在这些通路中发现其中有7个重要的DEPs (PGK、HK、LDH、PGAM、pfkA、IL6ST、PDK1)。亚细胞定位中质膜(28.9%)所占比例最高。结构域分析中主要以表皮生长因子样结构域为主。结果表明，DEPs分别富集在代谢、酶活性、低氧适应性等相关类别与通路中，并发现7个重要的DEPs (PGK、HK、LDH、PGAM、pfkA、IL6ST、PDK1)可能与缺氧环境的适应有关。这些结果为进一步研究牦牛PASMCs在低氧中的适应性提供了理论支持。

关键词: 牦牛, PASMCs, 低氧与常氧, TMT技术

Abstract: The purpose of this study was to analyze the differential expression proteins(DEPs) of yak pulmonary arterial smooth muscle cells (PASMCs) under different culture conditions of hypoxia and normoxia by using TMT technology, so as to understand DEPs for PASMCs of plateau yak. In this study, PASMCs were cultured by sticking culture method and cultured for 72 h under hypoxia and normoxia conditions, respectively. Then cells were collected and the total protein of cells was extracted. DEPs were screened for analysis using TMT labeled protein quantitative technology, and GO functional annotation and KEGG pathway analysis were performed. A total of 6 859 proteins were obtained by comparing the samples in 2 groups of PASMCs under hypoxia and normoxia. With the standard of FC(fold change) ≥ 1.3 or ≤ 0.78 and P<0.05, a total of 531 DEPs were obtained, among them, 186 DEPs were up-regulated and 345 DEPs were down-regulated. The results of enrichment analysis showed DEPs were mainly enriched in the hypoxia adaptation related pathways such as central carbon metabolism, glycolysis and glucose metabolism synthesis, HIF-1 signaling pathway, and biosynthesis of secondary metabolites. Seven important DEPs (PGK, HK, LDH, PGAM, pfkA, IL6ST and PDK1) were found in these pathways. Subcellular localization of plasma membrane (28.9%) accounted for the highest proportion. Epidermal growth factor-like domain was the main domain in domain analysis. The results indicated that DEPs were enriched in metabolism, enzyme activity, hypoxia adaptation and other related categories and pathways. Seven important DEPs (PGK, HK, LDH, PGAM, pfkA, IL6ST and PDK1) may be related to the adaptation to hypoxia environment. These results provide the theoretical support for further study on adaptation of yak PASMCs to hypoxia.

Key words: yak, PASMCs, hypoxia and normoxia, TMT technology

中图分类号:

S823.85

张兰, 姚一凡, 覃安琪, 李睿, 张伊阳, 陈树吾, 许瑾, 乔自林, 杨琨. 基于TMT技术对低氧与常氧条件下牦牛PASMCs的蛋白组学分析[J]. 畜牧兽医学报, 2022, 53(7): 2182-2193.

ZHANG Lan, YAO Yifan, QIN Anqi, LI Rui, ZHANG Yiyang, CHEN Shuwu, XU Jin, QIAO Zilin, YANG Kun. Proteomic Analysis for Yaks PASMCs under Hypoxia and Normoxia Conditions Based on TMT Technique[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(7): 2182-2193.

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