Enhancers are critical genetic elements that regulate transcription from promoters, shaping the spatial and temporal patterns of gene expression during development. Despite their importance, the mechanisms that allow enhancers to act across large genomic distances are not fully understood. Three-dimensional genome organization, including dynamic chromatin looping by cohesin, has been proposed to enable enhancer-promoter communication, but its precise role remains challenging to dissect.
This dissertation investigates the contribution of cohesin loop extrusion to long-range transcriptional regulation. Leveraging recent advances in genome engineering and inducible degradation systems, I acutely disrupt loop extrusion without arresting the cell cycle in mouse embryonic stem cells. Transcriptomic profiling reveals that not all loci with distal enhancers depend equally on loop extrusion. Using comparative genome editing, I demonstrate that enhancer-promoter communication across as little as 20 kilobases can require cohesin. However, promoter-proximal regulatory elements can support long-range, cohesin-independent enhancer action, either upon disabling extrusion or across strong insulators.
To assess whether loop extrusion is more broadly required during development, I extend these studies to neural progenitors and differentiation models. Genes dysregulated upon cohesin inhibition differ across cell types, likely reflecting distinct enhancer landscapes, yet transcriptional responses to differentiation cues remain largely intact.
Together, these findings provide mechanistic insight into how genome architecture and regulatory context shape enhancer function, with broad implications for development and disease.
