Characterization of a novel Salmonella regulator of IL-10 production and its role in pathogenesis

Duke University, Durham NC

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Abstract

? DESCRIPTION (provided by applicant): Salmonella enterica has been observed to manipulate the host pro-inflammatory immune response to optimize its own growth conditions. However, how S. enterica impacts and manipulates anti-inflammatory responses is poorly understood. Characterizing the mechanisms by which pathogens can trigger anti- inflammatory cytokines could have profound consequences for developing treatments for both infectious diseases and autoimmune disorders that are regulated by this anti-inflammatory cytokine. Thus, my long-term goal is to understand the mechanisms of pathogen manipulation of anti-inflammatory pathways and the resulting implications on infection and autoimmunity. The objective of this application is to characterize how S. enterica serovar Typhimurium induces production of the anti-inflammatory cytokine interleukin-10 (IL-10) and the role this plays on the progression and severity of infection. In pursuit of this objective, naturally occurring variation n the ability of S. enterica serovars to induce IL-10 revealed a genetic region present only in IL-10 inducing serovars. The hypothesis is that this region contains a gene encoding a Salmonella secreted effector that manipulates a host signaling pathway to produce IL-10. Deletion of this gene renders Salmonella unable to induce IL-10. Indicative of likely in vivo relevance, the deletion induces less IL-10 in mouse spleens, as measured by both qRT-PCR and IL-10-GFP reporter mice. Aim 1 of this proposal will determine how Salmonella induces IL-10 production. The hypothesis is that this unique protein is sufficient to induce IL-10 and interacts with host proteins to effect this response. This will be tested through 1) determining if expression of the gene in host cells is sufficient to induce IL-10 without accompanying S. enterica infection; 2) identification via RNAseq of host pathways altered in cells infected with wildtype vs. a mutant unable to induce IL-10; and 3) characterization of physical interactions with host proteins. Aim 2 will identify the cell types that are producing IL-10 in vivo and characterize the effects of the I-10-inducing factor and the induced IL-10 on infection progression and outcome. The hypothesis is that S. Typhimurium lacking induction of this anti-inflammatory pathway will show a defect in virulence. This will be tested by utilizing mouse models to examine bacterial loads, cytokine responses, and survival. IL-10-GFP reporter mice will be used to identify which cell types are producing IL-10 by flow cytometry with cell type-specific markers. Elucidating how intracellular S. enterica induces the IL-10 anti-inflammatory pathway will lead to a better understanding of how pathogens can modulate inflammation and how this can contribute to the effectiveness of clearance, chronic infection, and autoimmunity. In addition, the initiation of this IL-10 productio pathway may represent a new target for autoimmune therapies.