Systematic evaluation of candidate ligands regulating ectodomain shedding of amyloid precursor protein

Despite intense interest in the proteolysis of the β-Amyloid Precursor Protein (APP) in Alzheimer's disease, how the normal processing of this type I receptor-like glycoprotein is physiologically regulated remains ill-defined. In recent years, several candidate protein ligands for APP, including F-spondin, Reelin, β1 Integrin, Contactins, Lingo-1, and Pancortin, have been reported. However, a cognate ligand for APP that regulates its processing by α- or β-secretase has yet to be widely confirmed in multiple laboratories. Here, we developed new assays in an effort to confirm a role for one or more of these candidate ligands in regulating APP ectodomain shedding in a biologically relevant context. A comprehensive quantification of APPsα and APPsβ, the immediate products of secretase processing, in both non-neuronal cell lines and primary neuronal cultures expressing endogenous APP yielded no evidence that any of these published candidate ligands stimulate ectodomain shedding. Rather, Reelin, Lingo-1, and Pancortin-1 emerged as the most consistent ligands for significantly inhibiting ectodomain shedding. These findings led us to conduct further detailed analyses of the interactions of Reelin and Lingo-1 with APP.