Receptor tyrosine kinases (RTK) bind growth factors and are critical for cell proliferation and differentiation. Their dysregulation leads to a loss of growth control, often resulting in cancer. Epidermal growth factor receptor (EGFR) is the prototypic RTK and can bind several ligands exhibiting distinct mitogenic potentials. Whereas the phosphorylation on individual EGFR sites and their roles for downstream signaling have been extensively studied, less is known about ligand-specific ubiquitination events on EGFR, which are crucial for signal attenuation and termination. We used a proteomics-based workflow for absolute quantitation combined with mathematical modeling to unveil potentially decisive ubiquitination events on EGFR from the first 30 seconds to 15 minutes of stimulation. Four ligands were used for stimulation: epidermal growth factor (EGF), heparin-binding-EGF like growth factor, transforming growth factor-α and epiregulin. Whereas only little differences in the order of individual ubiquitination sites were observed, the overall amount of modified receptor differed depending on the used ligand, indicating that absolute magnitude of EGFR ubiquitination, and not distinctly regulated ubiquitination sites, is a major determinant for signal attenuation and the subsequent cellular outcomes.

受体酪氨酸激酶 (RTK) 结合生长因子，对细胞增殖和分化至关重要。它们的失调会导致生长失控，通常会导致癌症。表皮生长因子受体 (EGFR) 是原型 RTK，可以结合几种具有不同促有丝分裂潜力的配体。尽管已经广泛研究了单个 EGFR 位点的磷酸化及其对下游信号传导的作用，但对 EGFR 上的配体特异性泛素化事件知之甚少，这对于信号衰减和终止至关重要。我们使用基于蛋白质组学的工作流程进行绝对定量，并结合数学建模来揭示从刺激的前 30 秒到 15 分钟对 EGFR 具有潜在决定性的泛素化事件。四种配体用于刺激：表皮生长因子 (EGF)、肝素结合-EGF 样生长因子、转化生长因子-α 和上皮调节蛋白。虽然观察到单个泛素化位点的顺序差异很小，但修饰受体的总量因使用的配体而异，表明 EGFR 泛素化的绝对量级，而不是明显调节的泛素化位点，是信号衰减和随后的细胞结果。