Thesis presented July 02, 2009
Abstract: The deregulation of NF-κB signalling pathways, involved in cell survival and inflammation, leads to chronic inflammation and cancers. The aim of this thesis was to identify negative regulators of the conserved NF-κB pathways, Toll and Imd, in
Drosophila melanogaster. The stability or activity of several compounds of NF-κB pathways is regulated by ubiquitination. Consequently, the ubiquitine specific proteases (USPs) constitute a new area to look for regulators of these pathways. To do this, I constructed a collection of interfering RNA able to inactivate the 21 USPs of drosophila in S2 cells. The screening of this collection identified three negative regulators of the Imd pathway, one of them having also an effect on the Toll pathway. The target specificity of USPs could explain the small number of candidates. Among these candidates, dUSP36, a homologue of human USP36, was previously detected in the lab in a genetic screening. Team studies, in which I participated, show its
in vivo effect on the adaptative protein Imd through its catalytic activity. In order to characterise the two other candidates I performed transgenesis experiments in drosophila. These studies show that the two USPs are able to prevent activation of the Imd pathway in case of infection and that they are required to maintain the inactivated state of the Imd pathway in absence of infection. I also started to characterise the catalytic activity of both candidates
in vitro. My work's novelty was to limit the screening to only one gene family, which allowed the detection of new regulating genes which had not been revealed in previous screenings performed on a large part or the entire genome.
Keywords: NF-κB, ubiquitin protease, signal transduction, innate immunity, stress response
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