Basal IFN-λ2/3 expression mediates tight junction formation in human epithelial cells

Type-III interferons (or IFNλs) play important roles in antiviral defense and intestinal epithelial barrier integrity. While interferon expression has been primarily studied in response to pathogens, basal interferon expression also occurs in pathogen-free environments. However, the mechanisms regulating basal IFN-λ expression and their functions have not yet been elucidated. Here, we show that basal IFN-λ2/3 expression is linked to the development of an intact cellular epithelium characterized by formation of tight junctions and establishment of barrier function. Our findings indicate that basal IFN-λ2/3 expression depends on cGAS-STING-mediated mitochondrial DNA detection, while it is inhibited by the Hippo mechanotransduction pathway at low cellular densities. Cells lacking basal IFN-λ2/3 expression fail to develop proper tight junctions and establish normal barrier function. Mechanistically, IFN-λ2/3 suppresses Claudin-2 expression, thereby promoting barrier formation as cells become confluent. These results demonstrate a previously unknown function of basal IFNλ expression in regulating epithelial cell junction formation and highlight their importance not only during pathogen challenges but also in maintaining epithelial cell function under steady-state conditions.