Stable and EGF-Induced Temporal Interactome Profiling of CBL and CBLB Highlights Their Signaling Complex Diversity
Stable and EGF-Induced Temporal Interactome Profiling of CBL and CBLB Highlights Their Signaling Complex Diversity
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DOI：
10.1021/acs.jproteome.1c00284
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发表时间：
2021-06-17
影响因子：
4.4
通讯作者：
Tian, Ruijun
中科院分区：
文献类型：
--
作者：
Liu, Jie;Yang, Lijun;Tian, Ruijun
表皮生长因子受体（EGFR）信号调节细胞增殖、迁移和存活。EGFR的异常激活构成各种癌症的主要原因。CBL蛋白介导的受体泛素化和降解起负调控作用，控制信号传导的强度和持续时间。通过构建可诱导表达FLAG标记的CBL或CBLB的稳定细胞系，我们通过亲和纯化随后质谱（AP-MS）方法分别鉴定了102和82个CBL和CBLB的稳定相互作用蛋白。在6个不同时间点的时间分辨分析结合时间相互作用组的功能注释，提供了对EGFR信号传导激活后信号蛋白的动态组装的见解。CBL和CBLB相互作用组的比较表明，它们的功能是冗余的，但也是互补的。重要的是，我们通过生化测定验证了EPS 15 L1和ITSN 2的稳定关联以及TNK 2与CBL和CBLB的时间关联。总的来说，这些结果为CBL和CBLB相互作用组提供了有用的资源，并突出了它们在EGFR信号传导网络中的突出和多样化的作用。
The epidermal growth factor receptor (EGFR) signal modulates cell proliferation, migration, and survival. Aberrant activation of EGFR constitutes the major cause of various cancers. Receptor ubiquitination and degradation mediated by CBL proteins play negative regulatory roles and control the intensity and duration of the signaling. With the construction of stable cell lines inducibly expressing FLAG-tagged CBL or CBLB, we identified 102 and 82 stable interacting proteins of CBL and CBLB, respectively, through the affinity purification followed by mass spectrometry (AP-MS) approach. Time-resolved profiling at six different time points combined with functional annotations of the temporal interactomes provides insights into the dynamic assembly of signal proteins upon EGFR signaling activation. Comparison between the interactomes of CBL and CBLB indicates their redundant but also complementary functions. Importantly, we validated the stable association of EPS15L1 and ITSN2 and temporal association of TNK2 to both CBL and CBLB through biochemical assays. Collectively, these results offer a useful resource for CBL and CBLB interactomes and highlight their prominent and diverse roles in the EGFR signaling network.