PROJECT SUMMARY/ABSTRACT Living longer does not necessarily mean living better. As the global demographic shifts towards increased lifespan, so does the incidence and prevalence of aging-related diseases. Most common among them, Alzheimer's disease (AD) has no effective treatments for either prevention or recovery. This is primarily due to insufficient understanding of its pathophysiology. Increasing evidence connects AD to the primary cilium, a historically overlooked organelle that serves as the neuron's antenna. My analysis of differentially expressed genes (DEGs) in postmortem human AD brains has revealed a high prevalence of DEGs intricately linked to primary cilia. Notably, the gene ADCY3, which encodes for primary cilia-specific adenylate cyclase 3 (AC3), is significantly downregulated in human AD brains. Primary cilia are dependent on AC3, and loss of functional AC3 is associated with cognitive decline and reduced brain mass. One hallmark of AD is aberrantly processed amyloid precursor protein (APP), and it is notable that APP normally localizes near AC3 in the primary cilia. My preliminary studies further show that AC3 becomes insoluble at early stages of AD and localizes with amyloid beta (Ab) peptides outside of the neuron. This suggests a role for APP in normal maintenance of cilia structure, and the potential for interaction between AC3 and various Ab fragments derived from APP that could promote abnormal extracellular amyloid aggregation. My goals are (1) to characterize primary cilia pathology in AD patients and a preclinical mouse AD model, (2) to establish whether the accumulation APP within primary cilia can deteriorate cilial length, and (3) to test whether AC3-Ab interaction promotes accumulation of aberrant amyloid aggregates in AD. To this end, I propose three Specific Aims that combine in vivo and in vitro approaches. In Aim 1, I will characterize AD-related impairments to primary cilia structure using immunohistochemistry and electron microscopy in the brains of both human and mouse AD. In Aim 2, I will engineer cells in which the putative cilia-localization sequences within APP are abolished to determine whether the presence of APP within cilia is required for normal cilia structure. In Aim 3, I will define how AC3 impacts aberrant Ab fragment accumulation by utilizing an in vitro model of Ab aggregation into fibrils through coincubation with AC3 and the various Ab peptides found in normal and AD brain. Through these aims, I will establish the role that deterioration of primary cilia plays in AD. By providing key insights into the aggregation properties and kinetics of AC3 in amyloid plaque formation, I will also clarify how AC3 interacts with Ab peptides. My work could identify new therapeutic targets for patients with AD. I will additionally receive rigorous training in biostatistics, aggregation kinetics, and ultrastructural analysis of primary cilia and amyloid fragments. This will help prepare me for my desired caree...