c-Cbl directs EGF receptors into an endocytic pathway that involves the ubiquitin-interacting motif of Eps15.

c-Cbl associates with the activated EGF receptor before endocytosis. We here reveal that the capacity of c-Cbl to promote receptor internalization depends on its ubiquitin ligase activity, which functionally connects the EGF receptor to Eps15, a mediator of clathrin-coated pit formation. EGF-induced phosphorylation of Eps15, as well as recruitment of Eps15 to the plasma membrane and its co-localization with the EGF receptor in endosomes required the ubiquitin ligase activity of c-Cbl. This suggested that ubiquitin provides a direct or indirect link between the receptor and Eps15. Indeed, EGF-induced redistribution of Eps15 to the plasma membrane and endosomes depended on its ubiquitin-interacting motif. Upon over-expression, the ubiquitin-interacting motif abrogated the capacity of c-Cbl to promote EGF receptor endocytosis and only allowed receptor internalization via a route that lacked Eps15. Our findings disclose a novel function for the c-Cbl ubiquitin ligase and identify ubiquitin as a module that directs the EGF receptor into an endocytic pathway involving Eps15.