Development of bacterial autolysis strains to detect innate immune responses triggered by production of local mediators active on mucosal surfaces
Final Report Abstract
The type VI secretion system (T6SS) is a syringe-like nanomachine that is present in ~25% of the Gram-negative bacteria sequenced to date. It functions similar to a contractile phage tail in reverse orientation to translocate effector molecules from one prokaryotic cell into another bacterial cell or eukaryotic cells to either kill them or alter their cell biological processes. Vibrio cholerae, the causative agent of the diarrheal disease cholera, uses its T6SS to target commensal organisms in the gut. Injection of toxic T6SS effectors into sensitive ‘prey’ cells presumably leads to the release of their contents by lysis. This lysate is likely rich in molecules that display pathogen-associated molecular patterns (PAMPs) such as fragments of lipopolysaccharide (LPS) and peptidoglycan (PG). The initial sensing of PAMPs by eukaryotic cells is mediated by pattern-recognition receptors (PRRs) that initiate signal transduction cascades leading to the release of inflammatory cytokines. In vivo T6SS activity has been linked to host immune responses and an enhanced virulence gene expression by V. cholerae which suggests that killing of prey cells is sensed by the host and induces changes in the gut microenvironment that in turn modulates V. cholerae virulence properties. To address the role of cell lysis modulating host mucosal responses, I developed two orthogonal strategies to create ‘biological signal amplifiers’ that generate various PAMPs to induce transcriptional changes in the mucosal milieu. In one system I adapted the CRISPR-interference (CRISPRi) system for the knockdown of V. cholerae genes encoding essential proteins. As a second approach I used murein hydrolases as a life-death switch for the controlled lysis of bacteria. These approaches uncovered unexpected host responses that stimulated the ability of V. cholerae to cause disease. Thus, our method offers a unique new way to probe the response of the mucosal surface to categories of PAMP release.
Publications
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Poster flash talk and poster presentation at the Microbial Adhesion and Signal Transduction Gordon Research Conference (July 2023, US) Title: Programming bacterial lysis in vivo: Application in stimulating innate immune responses on mucosal surfaces
Mareike Lembke
