Quantification of tunicamycin-induced protein expression and N-glycosylation changes in yeast.
Quantification of tunicamycin-induced protein expression and N-glycosylation changes in yeast.
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DOI：
10.1039/c6an00144k
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发表时间：
2016-06
期刊：
影响因子：
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通讯作者：
Haopeng Xiao;Johanna M. Smeekens;Ronghu Wu
中科院分区：
文献类型：
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作者：
Haopeng Xiao;Johanna M. Smeekens;Ronghu Wu
衣霉素是一种有效的蛋白质N-糖基化抑制剂，经常被用来操纵细胞中的蛋白质糖基化。然而，衣霉素治疗引起的蛋白质表达和糖基化变化仍不清楚。我们以酵母为模型系统，在蛋白质组和N-糖蛋白组水平上系统地研究了衣霉素对细胞的响应。利用现代基于质谱学的蛋白质组学，我们量化了4259个蛋白质，几乎覆盖了整个酵母蛋白质组。衣霉素处理3h后，超过5%的蛋白质至少下调2倍，其中与几种糖代谢和糖酵解相关途径相关的蛋白质高度丰富。此外，由于蛋白质N-糖基化的抑制影响了蛋白质的折叠和运输，所以典型的未折叠蛋白质反应通路中的几个蛋白质被上调。我们还全面量化了衣霉素处理的细胞中蛋白质糖基化的变化，超过三分之一的量化独特糖肽(465个多肽中的168个)下调。根据基因本体论聚类，含有下调糖肽的蛋白质与糖基化、糖蛋白代谢过程、碳水化合物过程和细胞壁组织相关。目前的结果提供了第一个在蛋白质组和糖蛋白组水平上对衣霉素的细胞反应的整体看法。
Tunicamycin is a potent protein N-glycosylation inhibitor that has frequently been used to manipulate protein glycosylation in cells. However, protein expression and glycosylation changes as a result of tunicamycin treatment are still unclear. Using yeast as a model system, we systematically investigated the cellular response to tunicamycin at the proteome and N-glycoproteome levels. By utilizing modern mass spectrometry-based proteomics, we quantified 4259 proteins, which nearly covers the entire yeast proteome. After the three-hour tunicamycin treatment, more than 5% of proteins were down-regulated by at least 2 fold, among which proteins related to several glycan metabolism and glycolysis-related pathways were highly enriched. Furthermore, several proteins in the canonical unfolded protein response pathway were up-regulated because the inhibition of protein N-glycosylation impacts protein folding and trafficking. We also comprehensively quantified protein glycosylation changes in tunicamycin-treated cells, and more than one third of quantified unique glycopeptides (168 of 465 peptides) were down-regulated. Proteins containing down-regulated glycopeptides were related to glycosylation, glycoprotein metabolic processes, carbohydrate processes, and cell wall organization according to gene ontology clustering. The current results provide the first global view of the cellular response to tunicamycin at the proteome and glycoproteome levels.