Molecular Mechanisms and Functions of Estrogen Receptor Enhancers in Hormone-Dependent Gene Expression

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2017-11-21

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Abstract

Transcription is a fundamental regulatory mechanism of biological processes in a range of physiological and pathological conditions. Transcription enhancers are DNA regulatory elements that regulate the expression of the target genes by accommodating transcription factor (TF) binding through sequence specificity. Estrogen receptor alpha (ERα) belongs to ligand-dependent nuclear receptor superfamily. Upon activation by estrogenic ligands, ERα binds to specific sites on chromatin, and assembles and activates enhancer complexes, which in turn lead to the transcription of target genes. Various molecular events have been associated with enhancer function, including coregulator recruitment, induction of enhancer-enriched histone modifications, nucleosome remodeling, enhancer-promoter chromatin interactions, and transcription activation at the enhancer, as well as the target gene promoter. However, we lack a clear understanding of the order of events, the specific roles of each coregulator and enhancer-enriched chromatin features, and the functional relationships among them. Using ERα in estrogen (E2)-regulated gene transcription as a model in combination with molecular and cellular biology, as well as genomic and computational approaches, my dissertation herein describes a series of studies elucidating the molecular mechanisms and functions of these evens that lead to ERα enhancer activation. Collectively, it demonstrates that (1) ERα enhancer assembly and activation is a dynamic process, (2) the temporally-defined recruitment and activation of key coregulators are required for successful activation of ERα enhancers, and (3) enhancer transcripts (eRNA) mark active enhancers. Lastly, I delineate the development of a new technology, single-cell Global Run-on Sequencing (scGRO-seq), to uncover the link between enhancer activity and target gene transcription at the single-cell level. Single-cell imaging and sequencing technologies have demonstrated the heterogeneous nature of gene expression and enhancer activity in a wide range of biological systems, including clonally-expanded populations of cultured cells. However, our understanding on the molecular basis of heterogeneous gene expression is limited because of a lack of technologies that allow us to simultaneously examine enhancer activity and target gene transcription at the single-cell level. scGRO-seq will overcome this problem by capturing active transcription at the enhancers, which is an indicative of enhancer activity, and at the target gene in the same cells.

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Adaptor Proteins, Signal Transducing, Enhancer Elements, Genetic, Estrogen Receptor alpha, Estrogens, RNA, Untranslated

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