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Title: Exploring genetic susceptibility : using a combined systems biology, in vitro and ex vivo approach to understand the pathology of ulcerative colitis
Author: Brooks, Johanne
ISNI:       0000 0004 7227 6583
Awarding Body: University of East Anglia
Current Institution: University of East Anglia
Date of Award: 2018
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The overall aim of this PhD is to use a multidisciplinary approach to determine the function of Ulcerative Colitis (UC) associated SNPs, to help understand the role of SNPs in the pathogenesis of UC in general and in a patient specific context. UC is a chronic, relapsing inflammatory disease of the large bowel for which the aetiology is thought to be a trifecta of 1) dysregulation of the immune system in response to 2) an environmental trigger in a 3) genetically susceptible host. Genetic susceptibility or susceptibility loci for UC have been identified by Genome Wide Associations Scanning and subsequent fine mapping and deep sequencing. This work intended to further characterise these susceptibility loci at a global level and a patient specific level using both a systems biology approach and experimental validation of the in-silico work. Using publicly available datasets non exonic UC associated SNPs were functionally annotated to regulatory regions within the genome. Exonic SNPs were also analysed looking at impacts in protein linear motifs and splice enhancement motifs. Bioinformatics was used to identify interacting proteins and create a UC-interactome network. This suggested that UC was a disease of fine regulators as opposed to a disease of specific target proteins. Analysis of the UC-interactome identified the focal adhesion complex (FAC) that is involved in regulating wound healing as major component of the network. One member of the FAC, Leupaxin (LPXN), was identified as a potential target for validation. Using CRISPR-Cas9 technology, LPXN overexpressing cell lines and knock out cell lines were created. Wound healing assays and cytokine analysis identified that overexpression of LPXN impaired wound healing and reduced the secretion of MCP-1. In addition, using genotyped colonic biopsies from UC patients and control patients in a polarised in vitro organ culture (pIVOC) system we show that the LPXN risk allele may impact on cytokine production. Finally, UKIBD genetics consortium data was used to access a pilot dataset of 58 patients’ SNP profiles from Immunochip data who were patients at the Norfolk and Norwich University Hospital to create patient-specific UC-interactomes. Analysis of these footprints identified convergent interacting proteins affected by multiple SNPs and novel pathogenic pathways.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available