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Exploring bacterial binding to receptors of the CEACAM family as a basis for translational approaches

Exploring bacterial binding to receptors of the CEACAM family as a basis for translational approaches

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ROTH, Alexandra, 2013. Exploring bacterial binding to receptors of the CEACAM family as a basis for translational approaches [Dissertation]. Konstanz: University of Konstanz

@phdthesis{Roth2013Explo-24324, title={Exploring bacterial binding to receptors of the CEACAM family as a basis for translational approaches}, year={2013}, author={Roth, Alexandra}, address={Konstanz}, school={Universität Konstanz} }

2013 Roth, Alexandra deposit-license 2013-08-26T06:58:21Z eng Roth, Alexandra Exploring bacterial binding to receptors of the CEACAM family as a basis for translational approaches Carcinoembryonic antigen-related cell adhesion molecules (CEACAMs), expressed on human epithelia, are engaged by several Gram-negative human-restricted pathogens, including Neisseria gonorrhoeae. In detail,N. gonorrhoeae, the causative agent of gonorrhea, expresses several colony opacity (Opa) proteins that interact with the amino-terminal domain of epithelial CEACAMs (CEACAM1, CEA and CEACAM6). Accordingly, the interaction of gonococcal Opa proteins and CEACAMs enables N. gonorrhoeae to colonize human mucosal surfaces. In contrast to the benefit N. gonorrhoeae gains by binding to epithelial CEACAMs, binding of Opa proteins to the granulocyte receptor CEACAM3, results in bacterial phagocytosis and destruction. In the first part of the study we summarized the CEACAM3-mediated internalization and killing of bacteria and compared the CEACAM3-signaling pathways to the well-studied Fcγ receptors (FcγRs) and Dectin-1.<br />CEACAM3-mediated elimination of gonococci, might contribute to the fact that most gonococcal infections are local. Though, in rare cases, gonococci enter the bloodstream and cause systemic or disseminating gonococcal infections (DGI). Here we analyzed the CEACAM3-binding capacity of the DGI strain VP1. Accordingly, we identified the whole repertoire of VP1 opa genes and constitutively expressed them in Escherichia coli. Strikingly, several VP1 Opa proteins interacted with epithelial CEACAMs, but not a single VP1 Opa protein bound to the phagocyte receptor CEACAM3. Thus, E. coli expressing VP1 Opa proteins were not phagocytosed and eliminated by human granulocytes. Furthermore, the analysis of Opa variants from four additional clinical DGI isolates again demonstrated a lack of CEACAM3-binding. In conclusion, our results suggest that CEACAM3-avoidance of N. gonorrhoeae could be one factor that contributes to DGI.<br /><br />A major challenge for the treatment of gonorrhea is the emergence of multi-drug resistant gonococcal strains. In line with this, the investigation of additional treatment options is essential. Here, we tested a novel passive immunization approach which is based on the opsonin-dependent, FcγR-mediated phagocytosis of CEACAM-binding gonococci by human granulocytes. Precisely, we opsonized CEA-binding N. gonorrhoeae strains, including a multi-drug resistant strain, with soluble CEA-Fc fusion proteins. Importantly, we demonstrated that CEA-Fc opsonized gonococci are taken up into FcγR-expressing human cells as well as into human granulocytes. Phagocytosis of CEA-Fc opsonized gonococci by granulocytes resulted in an oxidative burst response and bacterial destruction. Thus, CEA-Fc fusion proteins could be a starting point for the development of novel therapeutics against antibiotic-resistant gonococci.<br /><br />CEACAM orthologues are also expressed in other mammals, but no bacterial ligands have been identified for non-human CEACAMs. Another aim of this study was to search for CEACAM-binding bacteria in rhesus monkey. For that purpose, we generated a soluble construct of the macaque orthologue of human CEA and developed a cultivation-independent screen to enrich a pool of macaque stool samples for macaque CEA-binding bacteria. By 16S rRNA gene pyrosequencing we identified several bacteria, belonging to the genus Prevotella, to be potential macaque CEA-binding bacteria. Biochemical approaches confirmed the interaction of Prevotella and macaque CEA. Unexpectedly, Prevotella also bound to several human CEACAMs, but not to CEACAMs from more distantly related mammals. Our observations indicate that CEACAM-engagement is species-specific and might contribute to the specific microbiota of different mammalian hosts.<br /><br />Altogether, the findings obtained in this study allow new insights in gonococcal pathogenicity and provide a basis for novel translational research approaches.

Dateiabrufe seit 01.10.2014 (Informationen über die Zugriffsstatistik)

Doktorarbeit Alexandra Roth.pdf 358

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