Defining gene-by-environment interactions using multiplex single-cell genomics

PROJECT SUMMARY The proposed research program aims to develop systems for the multiplex assessment of the effect of genetic perturbation on the response of cells to exogenous and endogenous exposures at the resolution of individual cells . Specifically, we will develop a flexible and scalable in vitro platform for high-throughput single-cell gene- by-environment interaction screens that couples CRISPR-based single-cell pooled genetic screens to a ?nuclear hashing?-based sample multiplexing strategy and a single-cell genomic readout. This platform will allow for the simultaneous molecular description of 1,000s-10,000s of unique conditions (i.e. gene and exposure combinations) at the resolution of individual cell types and cell states. In addition, we propose to develop a set of tools based on heterologous viral receptor expression for the in situ and cell-type specific delivery of CRISPR- based genetic perturbations in a manner compatible with single-cell RNA-seq. We highlight the versatility and generalizability of our approach across three applications of our technologies that aim to define gene-by- environment interactions in neurobiological disease. We propose to dissect the genetic interactions between mutations associated with Alzheimer?s disease and the transcriptional response of neural cell types to oxidative stress and peptide aggregates; to characterize the transcriptional effects associated with cell-type specific modulation of genetic modifier activity in response to mutations in genes that cause epilepsy disorder and; to determine how genes that are commonly mutated in brain cancer alter the response of murine tumors to therapies in vivo. Our goal is to arrive at clinically actionable molecular descriptions of how individual genes contribute to the response of individuals to exposure.