Division of Basic Sciences, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue N, Seattle, WA 98109, USA.
J Cell Sci. 2014 Feb 1;127(Pt 3):509-20. doi: 10.1242/jcs.127829. Epub 2013 Nov 27.
Phosphorylation-dependent protein ubiquitylation and degradation provides an irreversible mechanism to terminate protein kinase signaling. Here, we report that mammary epithelial cells require cullin-5-RING-E3-ubiquitin-ligase complexes (Cul5-CRLs) to prevent transformation by a Src-Cas signaling pathway. Removal of Cul5 stimulates growth-factor-independent growth and migration, membrane dynamics and colony dysmorphogenesis, which are all dependent on the endogenous tyrosine kinase Src. Src is activated in Cul5-deficient cells, but Src activation alone is not sufficient to cause transformation. We found that Cul5 and Src together stimulate degradation of the Src substrate p130Cas (Crk-associated substrate). Phosphorylation stimulates Cas binding to the Cul5-CRL adaptor protein SOCS6 and consequent proteasome-dependent degradation. Cas is necessary for the transformation of Cul5-deficient cells. Either knockdown of SOCS6 or use of a degradation-resistant Cas mutant stimulates membrane ruffling, but not other aspects of transformation. Our results show that endogenous Cul5 suppresses epithelial cell transformation by several pathways, including inhibition of Src-Cas-induced ruffling through SOCS6.
磷酸化依赖性蛋白泛素化和降解提供了一种终止蛋白激酶信号的不可逆机制。在这里，我们报告乳腺上皮细胞需要 Cullin-5-RING-E3-泛素连接酶复合物（Cul5-CRL）来防止Src-Cas 信号通路的转化。Cul5 的缺失会刺激生长因子非依赖性生长和迁移、膜动力学和集落形态异常发生，所有这些都依赖于内源性酪氨酸激酶Src。Cul5 缺陷细胞中Src 被激活，但Src 的单独激活不足以导致转化。我们发现Cul5 和 Src 一起刺激 Src 底物 p130Cas（Crk 相关底物）的降解。磷酸化刺激 Cas 与 Cul5-CRL 衔接蛋白 SOCS6 的结合，并随后进行蛋白酶体依赖性降解。Cas 是 Cul5 缺陷细胞转化所必需的。SOCS6 的敲低或使用降解抗性 Cas 突变体均能刺激细胞膜皱襞，但不能刺激其他转化方面。我们的结果表明，内源性 Cul5 通过多种途径抑制上皮细胞转化，包括通过 SOCS6 抑制 Src-Cas 诱导的皱襞。
