In vivo single-cell analysis of Alzheimer's disease-associated regulatory elements

PROJECT SUMMARY Treatment options for Alzheimer’s disease (AD) have been elusive, in large part because the molecular and cellular mechanisms underlying AD pathogenesis remain unclear. Recent studies have identified various cis-regulatory elements (CREs) associated with AD, including 75 genomic regions identified by genome-wide association studies (GWAS) and thousands of genomic regions with altered chromatin architecture in postmortem brains from individuals with AD. These CREs are thought to influence AD pathogenesis by affecting gene regulation in a cell type-specific and age-dependent manner. Therefore, it is crucial to study how AD-associated CREs orchestrate gene expression at single-cell resolution. To achieve this, we propose a scalable platform, single-cell Massively Parallel Reporter Assays (scMPRA), to investigate how AD-associated CREs regulate gene expression across different cell types. The scMPRA platform combines high-throughput sequencing and barcoding technology with single-cell RNA-sequencing to simultaneously assess the regulatory effects of thousands of CREs in a cell type-specific manner. We will apply scMPRA to in vivo mouse brains from two well-established AD models, which recapitulate both shared and distinct pathological aspects of AD, along with age-matched littermate controls. This will allow us to examine the regulatory effects of CREs within an intact physiological context, accounting for natural cellular interactions and aging processes, thereby offering new insights into the genetic architecture of AD. Upon completion, we aim to establish a novel genomic toolbox for identifying cell type-specific CREs, pinpoint cell types critical for aging and AD pathogenesis, and understand gene-environment interactions in the context of AD.