PROJECT SUMMARY/ABSTRACT The long-term goal of this project is to understand the age-related, neuron-microglia-astrocyte-specific proteome dynamics and interaction in the brain and their deregulation in the pathogenesis of Alzheimer?s disease (AD). Our ongoing large-scale profiling of transcriptome and proteome in bulk AD brain tissue have revealed that the transcriptome is often not an accurate indicator of protein abundance, due to posttranscriptional events, such as protein turnover. Genetic studies have revealed over 20 AD-related genes, many of which are associated with specific cell types. We hypothesize that dysfunction of neuron-microglia-astrocyte interaction contributes to AD pathogenesis, and propose to use the latest cutting edge technologies to profile cell type specific proteome, in situ transcriptome, and proteome turnover in the three cell populations from multiple common AD-related mouse models, to decipher molecular signaling networks in disease development, and to validate targets in human AD cases, mouse models, and human organoid models. Three specific aims are: (i) to disentangle dynamic neuron- microglia-astrocyte-specific proteome and interaction in AD mice by BONCAT-mediated single cell type proteomics and spatial transcriptomics, (ii) to measure cell type-specific proteome-wide protein turnover in AD by BONCAT pulse-chase analysis, and (iii) to study deregulation of proteome dynamics in human AD cases, mouse models and brain organoids. Successful outcome of this project will identify novel, cell type specific molecular components and pathways regulating cell-cell interactions during AD pathogenesis, which may provide new therapeutic strategies for effective treatment of this devastating disorder.