CbI-directed monoubiquitination of CIN85 is involved in regulation of ligand-induced degradation of EGF receptors

摘要

Addition of ubiquitin or ubiquitin chains to target proteins leads to their mono- or polyubiquitination, respectively. Whereas poly-ubiquitination targets proteins for degradation, monoubiquitination is thought to regulate receptor internalization and endosomal sorting. CbI proteins are major ubiquitin ligases that promote ligand-dependent polyubiquitination and degradation of receptor tyrosine kinases. They also recruit CIN85-endophilin in the complex with activated receptors, thus controlling receptor endocytosis. Here we show that the adaptor protein CIN85 and its homologue CMS are monoubiquitinated by CbI/CbI-b after epidermal growth factor (EGF) stimulation. Monoubiquitination of CIN85 required direct interactions between CIN85 and CbI, the intact RING finger domain of Cbl and a ubiquitin acceptor site present in the carboxyl terminus of CIN85. CbI-b and monoubiquitinated CIN85 are found in the complex with polyubiquitinated EGF receptors during prolonged EGF stimulation and are degraded together in the lyso-some. Dominant interfering forms of CIN85, which have been shown previously to delay EGF receptor degradation, were also impaired in their monoubiquitination. Thus, our data demonstrate that CbI/CbI-b can mediate polyubiquitination of cargo as well as monoubiquitination of CIN85 to control endosomal sorting and degradation of receptor tyrosine kinases.