Comprehensive and Cell Type-Specific Proteogenomic Profiling of Aberrant and Mis-Processed Proteins in Alzheimer's Disease and Tauopathy Models - Project Summary In Alzheimer's disease (AD), the most common form of dementia, aberrant and mis-processed proteins accumulate throughout the brain. AD-associated aberrant and mis-processed proteins may result from a variety of mechanisms, including protein-altering DNA sequence differences, dysregulated mRNA splicing, post- translational modifications, proteolytic cleavages, and protein misfolding. Although these mechanisms are known to affect a handful of proteins in AD, we have not been able to comprehensively characterize their impact on the proteome because of technological and methodological limitations. Thus, new approaches that can systematically profile aberrant and mis-processed proteins in AD are needed to improve our understanding of the disease's causes and identify actionable therapeutic targets. AD-associated protein dysfunctions also differentially affect distinct brain regions and cell types. To understand the factors responsible for this heterogeneity, we need approaches with sufficient scale to profile multiple brain regions and sufficient resolution to profile individual cells. Here, we pair state-of-the-art proteomic methods that we developed with proteogenomics, an approach that integrates genomic, transcriptomic, and proteomic data. The resulting proteogenomic pipeline will allow us to globally profile disease-associated proteins with amino acid substitutions, alternative protein isoforms, post-translationally modified proteins, and proteolytic cleavage products. In Aim 1, we apply our proteogenomic pipeline to multiple brain regions in a large AD cohort and multiple tauopathy mouse models. By profiling brain regions selectively vulnerable versus resilient to neurodegeneration in AD, the full spectrum of AD pathological stages, multiple disease subtypes, and multiple tauopathy mouse models using mass spectrometry methods that provide high proteome and individual protein sequence coverage, we will substantially expand our understanding of protein dysfunctions in AD. In Aim 2, we pair proteogenomics with the pioneering single-cell mass spectrometric proteomic methods we developed. We will profile individual neurons and non-neuronal cells extracted from human AD and tauopathy mouse brain tissues to provide needed insights into the cell type-specific factors that cause protein dysfunctions in AD. Across both aims, we apply rigorous experimental methods for functional validation and analytical approaches for causal inference to mechanistically characterize AD-associated aberrant and mis-processed proteins. Collectively, our aims will improve our understanding of AD's causes, identify new AD therapeutic targets, and provide new tools for studying protein dysfunctions in AD.
