Investigating the mechanisms and elements responsible for nuclear compartmentalisation during gene regulation. (360G-Wellcome-109110_Z_15_A)

The genome is organised into a highly ordered and complex structure. It is separated into distinct regulatory compartments which have a functional impact on transcriptional regulation. CTCF has been found to act as a boundary, with the ability to block enhancer-promoter interactions and insulate regulatory territories. The process that creates and regulates this genome organisation is not well understood, therefore, the main aim of the proposed research is to investigate the role of CTCF, enhancer and promoter elements in generating and defining regulatory domains. To achieve this, we will initially expand a series of models mutating the CTCF sites within the well characterised mouse alpha-globin locus, and explore genome-wide, polymorphic CTCF binding sites between inbred mice strains. The effects on local chromatin and transcription will be assessed using a combination of standard genomic assays used within the Hughes group, and the chromosome conformation capture technique NG Capture-C developed by them. To complement this, we will use genetic engineering to find the minimal requirements for compartment formation using the well-characterised functional elements from the mouse alpha-globin locus out of their native chromosomal context, to define the basic rules underlying their formation.