Clustering, processing and internalization of the amyloid precursor protein (APP) at the plasma membrane

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Alzheimer´s disease (AD) is a neurodegenerative disorder for which the deposition of amyloid plaques in the brain is a hallmark. Plaques contain Abeta peptides generated from the amyloid precursor protein (APP) by two consecutive cleavage steps. APP mutants were informative in elucidating putative disease mechanisms, suggesting that the rates of Abeta production or its propensity to aggregates play a role. 6 mutations are localized to the N-terminal region of Abeta, from which 5 are pathogenic and one is protective.We recently found that the first 5 amino acids of the N-terminal Abeta region mediate clustering of the APP protein in the plasma membrane and that clustering is required for entering the endocytic pathway (Schreiber et al., Biophysical Journal, 2012), via which production of the neurotoxic Abeta peptide is initiated. Preliminary data presented in the proposal show that the protective APP mutant, which carries a point mutation within the clustering region we previously described, is less clustered and less efficiently sorted to endocytic structures.These findings underline the importance of the N-terminal Abeta segment in regulating the dynamics of plasmalemmal APP. In addition, they suggest that the protective effect of the mutant is explained by less clustering and diminished endocytic uptake. Longer residency times in the plasma membrane would increase processing by alpha-secretases, generating non-toxic p3 peptides. Finally, as the clustering region is extracellular, a therapeutic strategy becomes plausible based on interfering with APP clustering from the extracellular site.Based on these data, first we aim for investigating the molecular mechanisms of plasmalemmal APP clustering mediated by the N-terminal segment of Abeta (2.3.1). We will clarify whether (i) self-oligomerization is involved (2.3.1.1), (ii) homophilic interactions are sufficient or other co-factors are required (2.3.1.2) and (iii) relevant residues can be better confined (2.3.1.3). To obtain proof-of-principle whether the above therapeutic strategy is feasible, we will screen for an aptamer binding to the clustering region with the intention of using it for abrogating clustering from the extracellular side (2.3.1.4).Second, as the rates of APP cleavage and internalization determine how much Abeta or p3 are produced, we will study their kinetics selectively at the plasma membranes. We will investigate whether changed lateral mobility and/or altered propensities of the mutants to aggregate are related to their processing in the plasma membrane (2.3.2.1) and/or internalization (2.3.2.3). Moreover, we will analyze the subcellular co-distribution of APP and secretases on the level of nano-domains (2.3.2.2).The project will provide novel insights in APP research, as APP clustering and trafficking, the understanding of how mutants exert protective/pathological effects and the exploration of a novel therapeutic strategy against AD.