The role of retromer in sorla-dependent APP transport and processing

Abstract

The retromer complex has been implicated in the intracellular transport of the amyloid precursor protein (APP). Retromer-deficiency leads to aberrant APP processing and the level of retromer proteins are altered in Alzheimer's disease (AD). SorLA is a neuronal sorting receptor genetically linked to late-onset AD with important functions in neuronal sorting of APP. The current study describes the interaction between sorLA and retromer and its role in APP transport and processing. Protein interactions were determined by fluorescence life-time imaging microscopy (FLIM) and co-immunoprecipitation analysis. APP processing products were determined by WB and ELISA. The subcellular localizations of sorLA and APP were characterized by confocal microscopy and electron microscopy analysis. Knockdown of expression of the VPS26 retromer subunit was undertaken by siRNA and shRNA technologies. We find that sorLA and retromer co-localize in neurons within Golgi and endosomal compartments, and that knockdown of retromer leads to a mislocalization of sorLA. Mutations of key amino acids responsible for retromer binding in the cytoplasmic domain of sorLA results in a redistribution of the receptors that is indistinguishable from that observed in cells following knockdown of VPS26. In retromer expressing cells, wild-type sorLA binds APP in the trans-Golgi network (TGN), diminishing processing of APP by both the amyloidogeninc and non-amyloidogenic pathway. The sorLA mutant that does not associate with retromer binds APP, as demonstrated by FLIM studies, but within distinct cellular compartments as compared to wildtype sorLA. Interestingly, the mutant receptor exhibits no protection against amyloid production. We have mapped the binding site for retromer in sorLA and provide evidence for a regulatory role of this receptor in retromer-dependent APP sorting and processing. We propose a model whereby retromer functions in the retrieval of sorLA from endosomes to the TGN.