Elucidating the function and requirement of insulator protein CTCF in transcriptional regulation in the mouse. (360G-Wellcome-099684_Z_12_A)

Regions of open chromatin referred to as DNaseI hypersensitive sites co-localise in mammals with putative cis-regulatory regions that include promoters, enhancers and CTCF binding sites. Of these, the function of CTCF binding sites is the least well understood, although roles in chromosome folding, partitioning and transcriptional regulation have been identified. Our aim is to identify the function of CTCF binding sites in the regulation ofgene transcription and the establishment of the local chromatin architecture. To achieve this, we will explore two methods to study the effects of differential CTCF binding on gene regulation; the disruption of CTCF binding by TALEN-mediated mutagenesis and the study of differential CTCF binding events between inbred mouse strains. The effects on local chromatin and transcription will be assessed by chromosome conformation capture techniques, chromatin immunoprecipitation and expression analysis of local genes. In support of these strategies, we will characterise the binding of several knownCTCF binding partners to sub-classify CTCF binding sites. Thus, we aim to use a combination of functional studies and genome-wide approaches to elucidate the role of CTCF binding sites in gene transcription and chromatin organisation.