Principal Supervisor: Professor Ferenc Mueller, Professor of Develpomental Genetics, Institute of Cancer and Genomic Sciences
Co-supervisor: Dr. Andrew Beggs, Reader, Institute of Cancer and Genomic Sciences
PhD project title: Mechanism and sequence determinants of regulatory (Enhancer – promoter) interactions
University of Registration: University of Birmingham
Mutations in non-coding functional elements of genes (such as cis-regulatory elements) cause misregulation of genes and lead to both congenital and multifactorial diseases including cancers. However, finding such mutations is much more problematic than in case of protein coding sequences, as we have little knowledge about the structure and function of non-coding functional elements of genes. Firstly, there is still no complete picture of how enhancers interact with and regulate their target genes. Secondly, the regulatory targets of enhancers and the mechanism and determinants of targeting the promoters they interact with are to a large extent unknown, which is a limiting factor in predicting the targets and limits functional validation. Understanding of enhancer-promoter interaction specificity is prerequisite to improving our chances of identifying the targets of predicted cis-regulatory mutations. The zebrafish, which produces a high number of transparent, extrauterine, rapidly developing embryos with easy transgenesis and high resolution in vivo imaging, is an ideal in vivo model for genetic /functional analysis of cis-regulatory elements and their interactions (Gehrig et al., 2009; Andersson et al., 2014; Pasquali et al., 2014; Haberle et al., 2014). In this project we shall utilise the apparent evolutionary conservation of higher order chromatin topology between fish and human and study the mechanisms and DNA sequence codes of enhancer-promoter interaction specificity in order to allow us to predict with much greater confidence the target genes of large number of distal regulatory elements and apply the uncovered principles on human genome, thereby improve the identification of disease relevant sequence variants and mutations, which fall in such regulatory elements and to aid in predicting and validating their target genes. We shall generate and utilise existing epigenomic profiles of enhancers including enhancer associated chromatin features (e.g. open chromatin, histone posttranslational modifications, enhancer transcription) and associate them with candidate target promoters, and manipulate predicted sequence determinants of enhancer promoter interaction using targeted genome editing of zebrafish. We shall use in vivo transgenic reporter essays for monitoring transcription and its aberrations upon targeted manipulation of candidate cis regulatory elements. The design of the zebrafish work will be informed by findings from whole genome sequencing of human disease patients and will focus on addressing the causes of misregulation of disease causing/associated genes mutated in their regulatory elements that are conserved between human and fish.
- Andersson, R., Gebhard, C., Miguel -Escalada, I., et al., Müller, F., FANTOM Consortium, Forrest, F., Carninci, P., Rehli, M., Sandelin A., Systematic in-vivo characterization of active enhancers across the human body. Nature, 507(7493):455-61.
- Haberle, V., Li, N., Hadzhiev, Y., et al., Carninci, P., Müller, F.*, Lenhard B. Two independent transcription initiation codes overlap on vertebrate core promoters. Nature, 507(7492):381-5.*co-corresponding author
- Pasquali, L., Rodríguez-Seguí, S., Miguel-Escalada, et al., Gomez Skarmeta, JL., Müller, F., McCarthy, M, and Ferrer J. (2014) Deconstructing the epigenomic and cis-regulatory code of human pancreatic islet-cells. Nat. Genet, 46(2):136-43.
- Gehrig J, Reischl M, Kalmár E, Ferg M, Hadzhiev Y, Zaucker A, Song C, Schindler S, Liebel U. and Müller, F. Automated high-throughput mapping of promoter-enhancer interactions in zebrafish embryos (2009). Nat. Methods, 6(12):911-6.
BBSRC Strategic Research Priority: Molecules, Cells and Systems
Techniques that will be undertaken during the project:
The project will provide training in genomics and embryo manipulation tools (NGS sequencing and computational analysis of epigenome features, targeted cas9 mediated genome editing and zebrafish transgenesis, in vivo imaging).
Contact: Professor Ferenc Mueller, Professor of Develpomental Genetics, Institute of Cancer and Genomic Sciences