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Identifying the intracellular survival strategies of chlamydiae

Clissold, Hayley (2017) Identifying the intracellular survival strategies of chlamydiae. PhD thesis, University of York.

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Chlamydia is the most commonly diagnosed sexually transmitted infection in the UK. The etiological agent, Chlamydia trachomatis (Ct), is an obligate intracellular bacterium that resides within an intracellular niche, termed the inclusion, following entry into host cells. Chlamydiae avoid lysosomal destruction by diverging away from the normal endocytic trafficking pathway. Ct expresses a type-III secretion system (T3SS) that enables the translocation of Ct effector proteins into the host cytoplasm and these proteins are essential for virulence. Relatively little is understood regarding the pathogenesis of chlamydiae and the mechanisms they use to manipulate host defences because, until recently, they have been intractable to genetic manipulation. In this thesis, we describe both a targeted and random screening approach for the identification of Ct T3SS effector proteins in the yeast Saccharomyces cerevisiae. We firstly use an in silico prediction program to identify Ct proteins that are likely to be secreted by the bacterium’s T3SS. The predicted proteins were then screened for their ability to disrupt membrane trafficking using an established pathogen effector protein screening in yeast (PEPSY) screening method. In parallel, we also generated a Ct genomic library in order to PEPSY screen the entire chlamydial genome for proteins involved in disrupting membrane trafficking. We identified two chlamydial deubiquitinases (DUB), ChlaDUB1 and ChlaDUB2, which disrupt intracellular membrane trafficking in a yeast model system. These chlamydial DUBs were expressed in mammalian cells and their effects on endosomal compartments, EGFR internalisation and degradation, IκBα levels and global ubiquitin levels were examined. Our findings suggest that ChlaDUB1 and ChlaDUB2 are likely to demonstrate broad substrate specificity towards host substrates and this research paves the way for future research to investigate the role of chlamydial DUBs in the manipulation of host membrane trafficking during infection.

Item Type: Thesis (PhD)
Academic Units: The University of York > Biology (York)
Depositing User: Miss Hayley Clissold
Date Deposited: 11 Jun 2018 09:41
Last Modified: 11 Jun 2018 09:41
URI: http://etheses.whiterose.ac.uk/id/eprint/20403

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