RUNX2 promotes epigenetic WNT signaling in inflamed intestinal epithelial cells

Cabrera-Silva, R. I.; Wilson, Z. S.; Miranda, J.; Fan, S.; Dame, M. K.; Bishu, S.; Spence, J. R.; Brazil, J.; Colacino, J.; Nusrat, A.; Parkos, C. A.

doi:10.1101/2025.11.17.688458

bioRxiv · 10.1101/2025.11.17.688458

RUNX2 promotes epigenetic WNT signaling in inflamed intestinal epithelial cells

Cabrera-Silva, R. I.·Wilson, Z. S.·Miranda, J.·Fan, S.·Dame, M. K.·Bishu, S.·Spence, J. R.·Brazil, J.·Colacino, J.·Nusrat, A.·Parkos, C. A.

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Abstract

Ulcerative colitis (UC) is characterized by chronic mucosal inflammation, recurrent epithelial injury, and impaired colonic mucosal wound healing. While WNT/{beta}-catenin dysregulation has been reported in UC, the mechanisms of such abnormalities remain unclear. To investigate epithelial intrinsic alterations associated with UC, we performed single-nucleus RNA-seq (snRNA-seq) and ATAC-seq (snATAC-seq) multiomics on human primary colonic epithelial cells (colonoids) from healthy donors and patients with inactive or active UC. Colonoids were cultured in a 3D matrix recapitulating crypt base cells or grown as 2D monolayers in differentiation medium to recapitulate luminal epithelial cells. Colonoids from active UC had a unique cell population with elevated CTNNB1 and reduced APC expression. Chromatin profiling identified enrichment of RUNX2 motifs in this UC-associated cell population. Active UC colonoids exhibited reduced OLFM4 expression in 3D and the differentiation marker VIL1 in 2D, suggesting impaired self-renewal and maturation. RUNX2 inhibition using CADD522 reduced {beta}-catenin levels in 3D colonoids and restored VIL1 expression and junctional {beta}-catenin localization in 2D cultures. These findings reveal an intrinsic defect in epithelial renewal in UC, driven in part by RUNX2-dependent WNT dysregulation. Our study identifies RUNX2 as a transcriptional regulator of epithelial stem cell function and WNT signaling in the inflamed human colon. Graphical Abstract summarySingle-nucleus RNA and ATAC sequencing of UC patient-derived colonoids reveals a RUNX2-associated WNT signature in active inflammation. Elevated {beta}-catenin and reduced OLFM4 and VIL1 expression indicate impaired self-renewal and differentiation. Pharmacologic inhibition of RUNX2 restores epithelial maturation, identifying RUNX2 as a key regulator of epithelial dysfunction in UC. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=134 SRC="FIGDIR/small/688458v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@14c0093org.highwire.dtl.DTLVardef@b9b6a3org.highwire.dtl.DTLVardef@858ee2org.highwire.dtl.DTLVardef@67b98c_HPS_FORMAT_FIGEXP M_FIG C_FIG

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Cabrera-Silva, R. I., Wilson, Z. S., Miranda, J., Fan, S., Dame, M. K., Bishu, S., Spence, J. R., Brazil, J., Colacino, J., Nusrat, A., Parkos, C. A.. 2025-11-17. RUNX2 promotes epigenetic WNT signaling in inflamed intestinal epithelial cells. https://doi.org/10.1101/2025.11.17.688458

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