Date of Award

2015

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Pharmacology

First Advisor

Gerd A. Blobel

Abstract

Bromodomain and Extra-Terminal motif proteins (BETs) associate with acetylated histones and transcription factors. While pharmacologic inhibition of this ubiquitous protein family is an emerging therapeutic approach for neoplastic and inflammatory disease, the mechanisms through which BETs act remain largely uncharacterized. Here we explore the role of BETs in the physiologically relevant context of erythropoiesis driven by the transcription factor GATA1. First, we characterize functions of the BET family as a whole using a pharmacologic approach. We find that BETs are broadly required for GATA1-mediated transcriptional activation, but that repression is largely BET-independent. BETs support activation by facilitating both GATA1 occupancy and transcription downstream of its binding. Second, we test the specific roles of BETs BRD2, BRD3, and BRD4 in GATA1-activated transcription. BRD2 and BRD4 are required for efficient transcriptional activation by GATA1. Despite co-localizing with the great majority of GATA1 binding sites, we find that BRD3 is not required for GATA1-mediated transcriptional activation. However, exogenous BRD3 efficiently compensates for BRD2 loss, suggesting that BRD2 and BRD3 function redundantly. Third, we tested the role of BETs in mitotic bookmarking. We present evidence that mitotic binding by BRD4, the BET most strongly implicated in preserving transcriptional state during mitosis, is extensively remodeled during this phase of the cell cycle. Additionally, disruption of mitotic BET occupancy has no measurable impact on post-mitotic gene reactivation, calling into question the role of BETs in mitotic bookmarking. These results elucidate new factors critical for GATA1-mediated erythropoiesis. In addition to furthering understanding of the mechanisms underlying BET function, these findings have important consequences for the rational development of BET inhibitors.

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