Abstract
Accumulation of extracellular amyloid beta peptide (Abeta), generated from amyloid precursor protein (APP) processing by beta- and gamma-secretases, is toxic to neurons and is central to the pathogenesis of Alzheimer disease. Production of Abeta from APP is greatly affected by the subcellular localization and trafficking of APP. Here we have identified a novel intracellular adaptor protein, sorting nexin 17 (SNX17), that binds specifically to the APP cytoplasmic domain via the YXNPXY motif that has been shown previously to bind several cell surface adaptors, including Fe65 and X11. Overexpression of a dominant-negative mutant of SNX17 and RNA interference knockdown of endogenous SNX17 expression both reduced steady-state levels of APP with a concomitant increase in Abeta production. RNA interference knockdown of SNX17 also decreased APP half-life, which led to the decreased steady-state levels of APP. Immunofluorescence staining confirmed a colocalization of SNX17 and APP in the early endosomes. We also showed that a cell surface adaptor protein, Dab2, binds to the same YXNPXY motif and regulates APP endocytosis at the cell surface. Our results thus provide strong evidence that both cell surface and intracellular adaptor proteins regulate APP endocytic trafficking and processing to Abeta. The identification of SNX17 as a novel APP intracellular adaptor protein highly expressed in neurons should facilitate the understanding of the relationship between APP intracellular trafficking and processing to Abeta.
Highlights
Sorting nexin 17 (SNX17) is a member of the sorting nexin family characterized by the presence of a phox (PX) domain [12], which serves to localize SNX17 to intracellular vesicles by interacting with specific phosphatidylinositol phosphates [13, 14]
Because SNX17 binds to lipoprotein receptor-related protein 1 (LRP1) through an analogous NPXY motif [17], we tested the ability of SNX17 to bind the amyloid precursor protein (APP) cytoplasmic domain
To analyze whether the YXNPXY motif within the APP tail is responsible for SNX17 binding, we mutated each residue within this motif to alanine, and their binding to SNX17 was examined by GST pulldown assay (Fig. 1A)
Summary
Sorting nexin 17 (SNX17) is a member of the sorting nexin family characterized by the presence of a phox (PX) domain [12], which serves to localize SNX17 to intracellular vesicles by interacting with specific phosphatidylinositol phosphates [13, 14]. Several cell surface adaptor proteins, including Fe65, X11, and Dab1, bind to the NPXY motif within the APP cytoplasmic domain and regulate its trafficking and processing to A (6 – 8). GST or the GST-fused APP cytoplasmic domain was incubated with lysates from U87 cells stably expressing Myc-tagged SNX17, and binding was examined by Western blotting using anti-Myc antibody.
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