Macrophage USP9X attenuates colitis by restricting oncostatin M production via K27-linked deubiquitination of STAT1.

Dysregulated posttranslational modifications drive inflammatory bowel disease (IBD) pathogenesis, yet the specific roles of deubiquitinases in intestinal macrophages remain poorly characterized. Here, we identify ubiquitin-specific peptidase 9X (USP9X) as a critical negative regulator of intestinal inflammation. USP9X expression is reduced in UC patients and inversely correlated with disease severity. Myeloid-specific deletion of USP9X exacerbates colitis in both DSS- and TNBS-induced models, whereas its overexpression ameliorates disease. Mechanistically, USP9X interacts with signal transducer and activator of transcription 1 (STAT1) and removes K27-linked polyubiquitin chains from lysine 544. The loss of USP9X enhances STAT1 nuclear translocation and transcriptional activity, resulting in the upregulation of oncostatin M (OSM) expression and exacerbating inflammation. Mutation of STAT1 at K544 attenuated both basal and USP9X knockdown-induced responses. Notably, neutralizing OSM abrogates the exacerbated colitis phenotype in USP9X-deficient mice. These findings define a USP9X-STAT1-OSM axis that connects ubiquitin editing to transcriptional regulation and represents a potential therapeutic target for IBD. USP9X suppresses STAT1 transcriptional activation by editing K27-linked ubiquitination at lysine 544 In wild-type conditions, USP9X removes K27-linked polyubiquitin chains from STAT1 at lysine 544, limiting its nuclear retention and transcriptional activity. In macrophages, loss of USP9X leads to enhanced K27-linked ubiquitination of STAT1, enhanced STAT1 nuclear translocation, and increased STAT1 occupancy at the Osm promoter, resulting in elevated OSM expression. AAV9-mediated USP9X delivery ameliorates intestinal inflammation by restoring USP9X expression. USP9X, ubiquitin-specific peptidase 9X; STAT1, signal transducer and activator of transcription 1; OSM, Oncostatin M. (Created with Biorender).