基于iTRAQ技术分析不同浓度的雷公藤甲素作用肺癌细胞后的差异蛋白表达

Identification of differential expression proteins in lung cancer cells treated with triptolide by iTRAQ

目的:分析雷公藤甲素(TP)作用人肺癌细胞(A549)后差异表达的蛋白质,以探讨TP抗肺癌的作用机制。方法:以12.5,50和200 ng·mL^(-1)3个浓度梯度的TP作用A549细胞后,提取细胞总蛋白,采用同位素标记的同位素相对和绝对定量(iTRAQ)技术结合纳升液相色谱-串联质谱(Nano LC-MS/MS)的定量蛋白质组学方法,分析和鉴定差异表达的蛋白质,运用生物信息学分析差异表达的蛋白质功能。结果:质谱共鉴定到4 977个蛋白(Local FDR<5%)。其中,在3个浓度均有差异表达蛋白质81个,呈下调趋势的40个,呈上调趋势的41个。通过基因富集分析发现,这些差异蛋白参与了细胞程序性死亡、代谢、蛋白合成等生物学过程。与蛋白合成相关的RNA转运信号通路和剪接体信号通路被显著抑制。结论:TP的广谱的抗肺癌作用机制,可能是通过抑制RNA转运和剪接相关的信号通路来实现的。

Objective： To analyze the differentially expressed proteins in human lung cancer cells（ A549）treated with triptolide（ TP）,and explore the anti-tumor mechanism of TP in proteomics level. Methods： A549 cells were treated with different concentrations of TP（ 12. 5,50 and 200 ng·mL-1） and the total proteins were extracted. After equal amount of protein digested by trypsin,the i TRAQ-labeled proteins were separated by nano liquid chromatography,and differentially expressed proteins were detected and identified by nano-MS/MS. The functions of proteins were analyzed by bioinformatics. Results： A total of 4 977 proteins（ local FDR 5%） were identified by mass spectrometry. Among them,there were 81 differentially expressed proteins in three concentrations,with 41 proteins significantly up-regulated and 40 down regulated. Through gene enrichment analysis,these differential proteins are involved in biological processes such as programmed cell death,metabolism,protein synthesis and so on. The RNA transport and splicesome signaling pathway associated with protein synthesis were significantly suppressed. Conclusion： The broad spectrum of anti-cancer mechanisms of TP may be associated with inhibiting RNA transport and splicesome signaling pathways.