Screening for EphB Signaling Effectors Using SILAC with a Linear Ion Trap-Orbitrap Mass Spectrometer

Erythropoietin-producing hepatocellular carcinoma (Eph) receptors play important roles in development, neural plasticity, and cancer. We used an Orbitrap mass spectrometer and stable isotope labeling by amino acids in cell culture (SILAC) to identify and quantify 204 proteins with significantly changed abundance in antiphosphotyrosine immunoprecipitates after ephrinB1-Fc stimulation. More than half of all known effectors downstream of EphB receptors were identified in this study, as well as numerous novel candidates for EphB signaling.

促红细胞生成素产生肝细胞癌（Erythropoietin-producing hepatocellular carcinoma, Eph）受体在个体发育、神经可塑性及肿瘤发生过程中发挥关键调控作用。本研究采用轨道阱质谱仪（Orbitrap mass spectrometer）与细胞培养氨基酸稳定同位素标记（Stable Isotope Labeling by Amino Acids in Cell Culture, SILAC）技术，对经ephrinB1-Fc刺激后的抗磷酸酪氨酸免疫沉淀物中丰度发生显著变化的204种蛋白质完成了鉴定与定量分析。本研究共鉴定出EphB受体下游半数以上已知效应蛋白，同时还发现了诸多参与EphB信号通路的新型候选蛋白。