Triple SILAC exosomal quantitative proteomic analysis

Exosomes are 30-100 nm sized membrane vesicles released by cells into extracellular space and mediate the intercellular communication via transfer of proteins and RNAs. To better understand the function of these microvesicles in lung carcinogenesis, we employed a Triple SILAC quantitative proteomic strategy to examine the differential protein abundance between exosomes derived from an immortalized normal bronchial epithelial cell line and two non-small cell lung cancer (NSCLC) cell lines harboring distinct activating mutations in the cell signaling molecules Kirsten rat sarcoma viral oncogene homolog (KRAS) or epidermal growth factor receptor (EGFR). We were able to quantify 727 exosomal proteins derived from the three cell lines. Proteins associated with signal transduction are enriched in NSCLC exosomes which are functionally active in regulating cell proliferation. The study is the first to investigate protein abundance differences in exosomes derived from NSCLC cells and their normal counterparts, and reveals the role of exosomes in NSCLC cancer progression, which may have clinical implications in biomarker development for patients with NSCLC.

外泌体(Exosomes)是一类直径30至100纳米的膜囊泡，由细胞释放至细胞外间隙，通过转运蛋白质与RNA介导细胞间通讯。为深入解析这类微囊泡在肺癌发生发展中的功能，我们采用三重稳定同位素标记氨基酸细胞培养（SILAC）定量蛋白质组学策略，对比分析了永生化正常支气管上皮细胞系与两种分别携带细胞信号分子Kirsten大鼠肉瘤病毒癌基因同源物(KRAS)或表皮生长因子受体(EGFR)功能性激活突变的非小细胞肺癌(NSCLC)细胞系所来源外泌体的蛋白质丰度差异。我们成功对这三种细胞系来源的727种外泌体蛋白质完成了定量分析。与信号转导相关的蛋白质在NSCLC来源的外泌体中显著富集，且这类外泌体在调控细胞增殖方面具有功能活性。本研究首次针对NSCLC细胞系及其正常同源细胞来源的外泌体蛋白质丰度差异展开研究，揭示了外泌体在NSCLC进展中的关键作用，其研究成果或可为NSCLC患者的生物标志物开发提供潜在临床应用价值。