TOPOLOGICAL REGULATION OF ENHANCER-PROMOTER COMMUNICATION

Abstract

The activation of gene expression by transcriptional enhancers remains a central mystery in molecular biology. One prevailing view has been that enhancers loop directly to the promoters of genes they regulate in a targeted fashion. However, recent studies have suggested that enhancer-promoter interactions occur with the context of topologically associating domains, opening up the possibility for potential enhancer-promoter and enhancer-enhancer crosstalk within these domains. Further, it is still unknown at what distances (both genomic and physical) enhancers can interact with their target promoters to elicit transcription.

In this dissertation, I employ quantitative live-imaging methods to investigate enhancer function in living Drosophila embryos. In Chapter 1, I introduce what is known about transcriptional regulation, the cis-regulatory elements that mediate it, and the current models for enhancer function in this process. In Chapter 2, I describe our strategy to visualize the process of transvection, whereby enhancers located on one homolog activate transcription on the other homolog. Interestingly, a shared enhancer was found to produce coordinated transcription from linked reporter genes in trans. In Chapter 3, I describe work done to explore transcriptional regulation at the endogenous fushi tarazu (ftz) and even-skipped (eve) loci. Evidence for enhancer-enhancer communication was found upon deletion of the eve stripe 1 enhancer. In Chapter 4, I investigate the distances that exist between genes co-regulated by a single enhancer during transcription. Using the transvection assay, large distances of at least 100-200nm were found to separate an enhancer from its target promoter.

Finally, in Chapter 5, I argue that these observations are all consistent with the occurrence of transcription “hubs”, which trap the transcriptional machineries mediating gene expression. These “hubs” therefore allow for action at a distance from transcriptional activators binding and acting at enhancers to the recruitment and release of Pol II at promoters. Together, these findings and resultant models bring novel insights to our understanding transcriptional regulation as well as highlight future directions for research.