Investigating factors contributing to enhancer-promoter interaction

Thesis

Enhancer-promoter interactions are drivers of tissue-specific gene expression and there are several mechanisms which contribute to their establishment and maintenance. These include accessibility and recruitment of transcription factors and co-factors to enhancer and promoter elements within the context of chromatin; compatibility of enhancer-promoter pairs; and the extent to which interactions are constrained by the 3D structure of the genome. Enhancer promoter interactions usually occur within topologically associated domains (TADs); large regions (200-2000kb) of interacting chromatin delimited by binding sites for the insulating factor CTCF. Whilst we know that all these mechanisms play a role, it remains unclear how each contributes. This thesis aims to address less well understood mechanisms of how enhancer-promoter communication is fine tuned in the context of the murine ⍺-globin locus. I report on a functional assay which highlights how actively transcribing genes can themselves act as insulator elements, similar in strength to a known strong CTCF site (HS-38). This assay also revealed the unexpected importance of gene orientation relative to their enhancers, such that maximum enhancer-induced activation is achieved when the genes are in their native orientation, whilst in the reverse orientation transcription is impeded. Finally, I also find enhancer-promoter communication, in the context of the ⍺-globin locus, is partially independent from CTCF sites throughout the locus; removal of most sites individually or in combination has no effect on alpha globin expression. However, when all 9 CTCF sites across the locus are deleted, ⍺-globin transcription is reduced. These results highlight the relationship between transcription and cohesin-mediated contacts and overall, the nuances of enhancer-promoter interactions which together achieve the optimised level of gene expression.

Authors: Cornell, LJ

• DOI: 10.5287/ora-zrjkxy77m
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English