NOD2 is an intracellular innate immune receptor expressed in dendritic cells and gastrointestinal epithelial cells. Polymorphisms in the NOD2 gene convey a strong predisposition to Crohn’s disease (CD), a form of inflammatory bowel disease. Understanding the function of NOD2, and in what way it is aberrant in the presence of NOD2 polymorphisms, would confer a valuable paradigm for understanding Crohn’s pathogenesis. CD is thought to arise both from defects in the gut mucosal barrier and from a dysregulated Th17 immune response to commensal gut flora. Aberrant expression of IL-23 is present in both human CD and in murine models of colitis. Wild-type NOD2 contributes to NFκB activation and pro-inflammatory cytokine production on recognition of its cognate ligand, a function that is lost in CD-associated mutations. How the predominantly loss-of-function CD-NOD2 contributes to the pro-inflammatory response present in Crohn’s is not yet understood. In this thesis a set of experiments is described that aim to shed light on the function of NOD2, firstly through identification of negative regulators of immune activation that are dependent on NOD2 for their expression. This work identifies the microRNAs that are expressed following NOD2 triggering in human dendritic cells. Specifically, up-regulation of the miR-29 family was found to be dependent on wild-type NOD2 function. A number of novel miR-29 targets and their functional consequences are presented, including the cytokine subunits IL-12p40 and IL-23p19, directly linking NOD2 polymorphisms and aberrant IL-23 expression. Secondly, a project aiming to identify components of the NOD2 signalling complex (or signalosome) is described. To this end I employed a model system that involved tagging NOD2, and stable expression in a human cell line. These clones were validated for expression and function before an immunoprecipitation protocol was optimised. Mass spectrometry analysis of these samples identified the known NOD2-interacting protein Erbin.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:560929 |
| Date | January 2013 |
| Creators | Brain, Andrew Oliver Seaward |
| Contributors | Simmons, Alison |
| Publisher | University of Oxford |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://ora.ox.ac.uk/objects/uuid:d04c56dd-1325-4ff3-8810-4991d2fd61b0 |
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