Clostridioides difficile Toxin B Activates Group 3 Innate Lymphocytes

Group 3 innate lymphocytes (ILC3s) are rare immune cells localized in mucosal tissues, especially the gastrointestinal (GI) tract. Despite their rarity, they are a major source of the cytokine interleukin-22 (IL-22), which protects the GI epithelium during inflammation and infection. Although ILC3s have been demonstrated to be important for defense against Clostridioides difficile infection, the exact mechanisms through which they sense productive infection and become activated to produce IL-22 remain poorly understood. In this study, we identified a novel mechanism of ILC3 activation after exposure to C. difficile. Toxin B (TcdB) from C. difficile directly induced production of IL-22 in ILC3s, and this induction was dependent on the glucosyltransferase activity of the toxin, which inhibits small GTPases. Pharmacological inhibition of the small GTPase Cdc42 also enhanced IL-22 production in ILC3s, indicating that Cdc42 is a negative regulator of ILC3 activation. Further gene expression analysis revealed that treatment with TcdB modulated the expression of several inflammation-related genes in ILC3s. These findings demonstrate that C. difficile toxin-mediated inhibition of Cdc42 leads to the activation of ILC3s, providing evidence for how these cells are recruited into the immune response against the pathobiont.