All ETDs from UAB

Advisory Committee Chair

Jeremy J Day

Advisory Committee Members

Matthew S Alexander

Eric D Roberson

Summer B Thyme

Linda S Overstreet-Wadiche

Document Type

Dissertation

Date of Award

2020

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Genomic enhancers are cis-regulatory elements in the DNA that exhibit spatial and temporal control over intricate gene expression programs shaping fate and function of heterogeneous cell populations in the developing and adult brain. In addition to their role in cellular development and maintenance, enhancer activity drives transcriptional changes in response to stimulation. Active enhancers are subject to bidirectional transcription that yields non-coding enhancer RNAs (eR-NAs). However, the functional role of eRNAs in transcriptional regulation remains controversial. Therefore, we used primary neuronal cultures to investigate en-hancers and eRNAs in the rat genome. We identified and verified enhancer-gene pairs, characterized their response to different types of neuronal stimulation, and demonstrated that Fos eRNAs serve as critical transcriptional regulators of the immediate early gene Fos. Moreover, we provide evidence that the underlying regulatory mechanisms are mediated by eRNA-protein interactions between Fos eRNAs and acetyltransferase CBP. Furthermore, we developed a novel Cre-dependent CRISPR activation (CRISPRa) tool for cell type-specific activation of endogenous genomic loci, and optimized an intron-containing, double-floxed dCas9-VPR construct. This new strategy alleviated leaky target gene induction compared to a traditional Cre-dependent CRISPRa construct. Overall, the work described in this dissertation indicates that eRNAs regulate gene expression facili-tating transcriptional adaptations to neuronal activity, and introduces a novel tool to facilitate future studies of distinct genes and regulation mechanisms.

Share

COinS