Intestinal granuloma formation during Yersinia pseudotuberculosis infection

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Project Summary/Abstract In various infectious and non-infectious contexts, chronic immune stimulation induces the formation of granulomas: aggregations of recruited immune cells that are thought to encapsulate pathogens and prevent their dissemination. Despite being a prominent feature of numerous infections, key gaps in our knowledge are the mechanisms of granuloma formation and the functional role of these structures in controlling infectious disease. Yersinia are bacterial pathogens that block immune cell function and induce granuloma formation in lymphoid tissues. Yersinia pseudotuberculosis (Yptb) causes self-limiting gastroenteritis and lymphadenitis in immunocompetent hosts following fecal-oral transmission. Yptb subverts the immune response through the injection of Yersinia outer proteins (Yops) into nearby immune cells through a needle-like type III secretion system, blocking functions such as phagocytosis and pro-inflammatory gene expression. While granulomas are well-described during Yersinia infection of lymphatic tissue, relatively little is known about early intestinal infection. In this proposal, we describe, for the first time, a murine model of granuloma formation in the intestinal mucosa during acute Yptb infection. Importantly, live bacteria are abundant within granulomas but are largely absent from surrounding non-granuloma intestinal tissue, suggesting that granulomas play a previously uncharacterized role at the intestinal mucosa, an immunological barrier that bottlenecks Yersinia dissemination. Interestingly, Yptb lacking Yops does not induce intestinal granuloma formation, suggesting that these structures form in response to features of bacterial virulence. Further, intestinal granulomas are highly enriched in neutrophils and inflammatory monocytes. Strikingly, monocyte-deficient Ccr2- /- mice show defects in restriction of bacterial dissemination, succumbing to acute infection. Similarly, mice deficient in tumor necrosis factor (TNF) signaling, a cytokine that enhances phagocyte microbicidal function, exhibit defects in bacterial restriction. I therefore hypothesize that blockade of immune cell function by Yop effector proteins induces the formation of intestinal granulomas, which protect the host through bacterial restriction mediated by monocyte-derived TNF. In this proposal, I will investigate the formation and function of intestinal granulomas from both the bacterial and host sides. First, I will uncover how Yersinia virulence factors induce intestinal granuloma formation by testing a panel of Yop mutant strains, in addition to using an injection reporter strain (Aim 1). Second, I will dissect immune cell functions that are necessary for restriction of Yersinia by intestinal granulomas through complementary mechanistic studies using chimeric animals and genetic ablation systems (Aim 2). This work will mechanistically define a previously unappreciated facet of the host immune response to Yersinia infection. Findings from this novel model of intestinal granuloma formation will provide insight into future work on poorly-studied granulomatous disorders and underscore potential therapeutic targets for the treatment of chronic disease.