NOX1 drives inflammatory bowel disease through oxidant-dependent stabilization of IκBζ

NADPH oxidase 1 (NOX1), predominantly expressed in colonic epithelial cells, regulates mucosal immunity and homeostasis through the production of reactive oxygen species (ROS). However, its dysregulation may contribute to inflammatory bowel disease (IBD). IκBζ, an atypical IκB family member, amplifies inflammation by inducing the expression of several pro-inflammatory genes. Here, we investigated whether NOX1 could regulate IκBζ stabilization thus sustaining pro-inflammatory gene expression in IBD. NOX1 and IκBζ expression were upregulated in colonic tissues and organoids from IBD patients, and this expression pattern was reproduced in healthy organoids stimulated with key inflammatory mediators associated with IBD, namely flagellin, TNFα and IL-17. Silencing NOX1 across multiple models including T84 colonic epithelial cells, human intestinal organoids, and mice, inhibited IκBζ induction in response to inflammatory mediators, demonstrating a critical role of NOX1 in regulating IκBζ expression. Furthermore, ROS directly oxidized IκBζ in colonic epithelial cells under inflammatory conditions, preventing its proteasomal degradation and promoting its stabilization. Mutation of cysteine 638, a solvent-exposed residue of IκBζ, abolished protein stabilization, suggesting that oxidation occurs at this site. Transcriptomic analysis of organoids under inflammatory conditions after silencing NOX1 or IκBζ, revealed that the NOX1-IκBζ axis controls a network of inflammatory genes, including CCL2, identified as a hub gene. In IBD patients, elevated CCL2 secretion correlated with IκBζ oxidation, reflecting increased IκBζ stabilization. Together, these findings uncover a redox-sensitive NOX1-ROS-IκBζ axis that sustains intestinal inflammation through IκBζ stabilization and pro-inflammatory gene induction, and identify this axis as a potential therapeutic target in IBD.