983 Corynebacterium diphtheriae causes keratinocyte-intrinsic ribotoxic stress and NLRP1 inflammasome activation in a model of cutaneous diphtheria

Human NLRP1 is a multifunctional sensor predominantly expressed in the skin and airway epithelium. Recent work from our lab and others has demonstrated that human NLRP1 can sense viral infections, activating a pro-inflammatory form of cell death. In our latest unpublished work, we report that human NLRP1 can also sense bacterial infections. Mechanistically NLRP1 functions as a component of the ribotoxic stress response (RSR) and is indirectly, activated by bacterial exotoxins that target human ribosome elongation factors EEF1 and EEF2. As such, NLRP1 can sense exotoxins including Diphtheria Toxin (DT) from Corynebacterium diphtheriae, exotoxin A from Pseudomonas aeruginosa and sidI from Legionella pneumophila leading to pro-inflammatory cell death. We show that in primary epithelial cells, toxigenic Corynebacterium diphtheriae filtrate triggers ribotoxic stress signaling and NLRP1-dependent pyroptosis, in a manner that requires ZAKα and p38 kinase activity in the host. Furthermore, using 3D human skin cultures, Corynebacterium diphtheriae infection severely disrupts skin barrier function and induces IL-1 driven inflammation. Pharmacologic inhibition of p38 and ZAKα kinase activity suppresses keratinocyte pyroptosis and rescues skin barrier integrity of Corynebacterium diphtheriae-treated 3D skin. In summary, we report that multiple bacterial exotoxins that target distinct ribosome elongation factors activate the NLRP1 inflammasome in primary epithelial cells. Using cutaneous diphtheria as a model, we demonstrate that one of these toxins, DT, disrupts epidermal barrier integrity via ribotoxic stress and pyroptosis. Pharmacologic inhibition of RSR prevents DT-driven pyroptosis and as a result alleviates C. diphtheria-induced skin damage and inflammation. These findings implicate a broader role of RSR and the NLRP1 inflammasome in innate immunity against bacterial diseases and might explain certain aspects of diphtheria pathogenesis, such as the fatal systemic dissemination of DT following skin barrier disruption.