SEL-10/Fbw7-dependent negative feedback regulation of LIN-45/Braf signaling in C. elegans via a conserved phosphodegron
SEL-10/Fbw7-dependent negative feedback regulation of LIN-45/Braf signaling in C. elegans via a conserved phosphodegron
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DOI：
10.1101/gad.203703.112
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发表时间：
2012-11-15
影响因子：
10.5
通讯作者：
Greenwald, Iva
中科院分区：
文献类型：
--
作者：
de la Cova, Claire;Greenwald, Iva
在秀丽隐杆线虫中命名为SEL-10的保守的E3泛素连接酶组分和在哺乳动物中命名为Fbw 7的保守的E3泛素连接酶组分通过称为Cdc 4磷酸降解决定子（CPD）的高亲和力结合位点靶向泛素介导的降解底物。由于Fbw 7的许多已知底物都是癌蛋白，因此新底物的鉴定可以提供对癌症生物学以及蛋白质组调控方面的见解。在这里，我们评估了进化上保守的CPD的存在是否是一个可行的补充，以蛋白质组学为基础的方法，以确定新的潜在底物。对于功能评估，我们专注于LIN-45，它是外阴诱导的信号转导途径的一个组成部分，也是人类Braf的直系同源物，Braf是许多癌症中Ras的效应子。我们的分析表明，LIN-45作为SEL-10的真正底物，CPD突变或sel-10缺失导致体内活性和蛋白质稳定性增加。此外，在外阴诱导过程中，下游激酶MPK-1/ERK也是负反馈回路中LIN-45蛋白降解所需的，导致ERK高度活性的LIN-45降解。由于CPD共有序列在人Braf中是保守的，我们提出Fbw 7也可以在某些细胞环境中调节Braf稳定性。我们讨论了我们的研究结果的影响，在C外阴发育。elegans，对人Braf的潜在适用性，以及基于CPD的人Fbw 7底物预测方法的价值。
The conserved E3 ubiquitin ligase component named SEL-10 in Caenorhabditis elegans and Fbw7 in mammals targets substrates for ubiquitin-mediated degradation through a high-affinity binding site called a Cdc4 phosphodegron (CPD). As many known substrates of Fbw7 are oncoproteins, the identification of new substrates may offer insight into cancer biology as well as aspects of proteome regulation. Here, we evaluated whether the presence of an evolutionarily conserved CPD would be a feasible complement to proteomics-based approaches for identifying new potential substrates. For functional assessments, we focused on LIN-45, a component of the signal transduction pathway underlying vulval induction and the ortholog of human Braf, an effector of Ras in numerous cancers. Our analysis demonstrates that LIN-45 behaves as a bona fide substrate of SEL-10, with mutation of the CPD or loss of sel-10 resulting in increased activity and protein stability in vivo. Furthermore, during vulval induction, the downstream kinase MPK-1/ERK is also required for LIN-45 protein degradation in a negative feedback loop, resulting in degradation of LIN-45 where ERK is highly active. As the CPD consensus sequence is conserved in human Braf, we propose that Fbw7 may also regulate Braf stability in some cell contexts. We discuss the implications of our findings for vulval development in C. elegans, the potential applicability to human Braf, and the value of a CPD-based predictive approach for human Fbw7 substrates.