Alzheimer's disease is characterized by the presence of both beta- amyloid plaques and neurofibrillary tangles in cortex. Increasing evidence favors the deposition of amyloid beta-protein (Abeta) in plaques as an early and possibley primary event in the pathogenesis of Alzheimer's disease, a process that may be related to altered expression or processing of the amyloid precursor protein (APP). Recent studies have further implicated the longer Abeta species, specifically Abeta peptides of 42 amino acids long (Abeta42) as potentially critical for amyloid deposition and fibril formation. The pathways that lead to the generation of Abeta and Abeta42 have not been clearly defined. The foundation that guides this ongoing project is that processing of APP in the endocytic pathway is important to Abeta production. Accordingly, we have formulated two working hypotheses to direct our continuing research efforts: 1) the APP internalization pathway is the primary route for Abeta production and subsequent release into the medium, and 2) familial Alzheimer's disease mutations alter APP trafficking and, in turn, Abeta production. Three Specific Aims are proposed for the next granting period to test the two working hypotheses. The first Specific Aim examines the role of endocytic processing in the production and release of Abeta42. The second Specific Aim will analyze the relationship between presenilin-1 mutations, APP trafficking and Abeta42 production. In the third Specific Aim, the mechanism by which the APP codon 717 mutations increases Abeta42 production will be explored with regards to the relationship between internalization and gamma-secretase APP cleavage. This investigation of the APP trafficking pathways in a cell culture system will examine fundamental processes that are critical for Abeta (Abeta42) production. Results from these studies may provide important insight into the pathogenesis of Alzheimer's disease.