Pathogenic OTUD3 mutations predispose to ulcerative colitis due to barrier dysfunction.

Background and aims: The contribution of common genetic polymorphisms to ulcerative colitis (UC) pathogenesis is modest, however families with severe colitis may harbor rare variants with large effect sizes that highlight unrecognized pathways.

Methods: A multigenerational family with UC necessitating colectomy was identified. Whole exome sequencing of this kindred was performed, implicating a rare variant in OTUD3. Constitutive knock-out and intestinal specific Otud3 deficient and heterozygous mice were generated. OTUD3 expression in human colonic biopsies and intestinal organoids was assessed using qRT-PCR and immunofluorescence. Prevalence of rare, damaging variants were compared in distinct patient cohorts. Plasmids containing OTUD3 missense variants were introduced into cell lines where OTUD3 was disrupted to determine their effects on cellular response to cytokine stimulation.

Results: Constitutive disruption or heterozygosity of Otud3 in mice, or intestinal-specific deletion, resulted in impaired barrier integrity, tight-junction dysregulation, increased ER stress, and penetration of luminal bacteria deep into the colonic crypts that preceded a spontaneous progressive colitis. Analysis of distinct UC cohorts demonstrated enrichment of rare, damaging variants in OTUD3. Introduction of OTUD3 variants in intestinal cell lines phenocopied the epithelial immune dysregulation observed in knockout mice. Finally, OTUD3 mRNA and epithelial protein expression were decreased in the quiescent colonic epithelial tissue of genotype-unselected individuals with UC compared to matched non-UC controls.

Conclusions: Our results demonstrate that OTUD3 is required for colonic epithelial barrier function, and plays a role in the pathogenesis of UC.