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Title: “Identification of CAR-Responsive Active Enhancers at DNase-I Hypersensitive Sites by Massively Parallel Reporter Assay in TCPOBOP-Exposed Mouse Liver”

Committee: David J. Waxman, PhD – Biology, BME (Advisor, Chair) Ahmad S. Khalil, PhD – BME John Ngo, PhD – BME Juan Fuxman Bass, PhD – Biology

Abstract: Functional noncoding transcripts can participate in biological processes through a role in gene regulation and epigenetic regulation affecting chromatin structure and accessibility. However, compared with nearly complete genome sequencing, gene annotation and activity profiling across the entire genome is still lagging, especially within noncoding regions. Lack of annotation on regulatory elements hinders the understanding of response pathways involved. This proposal capitalizes on massively paralleled reporter assays (MPRAs), multi-omics sequencing, and computational biology to systematically dissect mechanisms underlying the disruptive actions of environmental chemicals upon the activation of nuclear receptor/transcription factor CAR (Nr1i3) in mouse liver. Here, the proposed aims will study the transcription activity of open chromatin regions by using an in vivo genome-wide study (STARR-seq) and characterize those active enhancers in terms of their epigenetic features, transcription factor binding activity, responsiveness to environmental chemicals and enhancer-promoter interaction toward their gene targets. Furthermore, a novel single-cell reporter assay (sc-STARR-seq) will facilitate the understanding of how active enhancers contribute to the heterogeneity of metabolic capability. Lastly, the functional association between active enhancers and target genes will be verified through epigenetic perturbations. The work proposed will shed light on the mechanisms whereby foreign chemicals dysregulate gene expression linked to dysfunctional metabolism and pathophysiological diseases.