The epsilon hinge-ear region regulates membrane localization of the AP-4 complex

Adaptor protein (AP) complexes are key factors for the spatial and temporal regulation of intracellular trafficking events. Four complexes (AP-1, -2, -3, -4) are known, among which AP-4 is only poorly characterized. Recent work suggests a role for AP-4 in the intracellular trafficking of the β-amyloid precursor protein and molecular genetics showed that the loss of functional AP-4 is associated with congenital neuronal disorders of severe cognitive dysfunction. To unravel the molecular mechanisms controlling AP-4 functions, we established the intracellular expression of recombinant AP-4 complex. This approach combined with the analysis of mutant complexes allowed us to discover that the epsilon adaptin hinge-ear region has a function in membrane recruitment of AP-4. We further show that this process is phosphorylation dependent and involves PP2A-like protein phosphatases and a staurosporine-sensitive kinase. Deletion of the residues 839-871 in the carboxy-terminal region of the hinge of epsilon adaptin abrogated the membrane/cytosol recycling of AP-4. As targets of phosphorylation, we identified three serine residues: S847, S868 and S871. We conclude that the terminal hinge region and the appendage of the AP-4 epsilon subunit are involved in membrane association in a process that is controlled by phosphorylation and dephosphorylation events.