Wnt/β-catenin signaling is a primary pathway for stem cell maintenance during tissue renewal and a frequent target for mutations in cancer. Impaired Wnt receptor endocytosis due to loss of the ubiquitin ligase RNF43 gives rise to Wnt-hypersensitive tumors that are susceptible to anti-Wnt-based therapy. Contrary to this paradigm, we identify a class of RNF43 truncating cancer mutations that induce β-catenin-mediated transcription, despite exhibiting retained Wnt receptor downregulation. These mutations interfere with a ubiquitin-independent suppressor role of the RNF43 cytosolic tail that involves Casein kinase 1 (CK1) binding and phosphorylation. Mechanistically, truncated RNF43 variants trap CK1 at the plasma membrane, thereby preventing β-catenin turnover and propelling ligand-independent target gene transcription. Gene editing of human colon stem cells shows that RNF43 truncations cooperate with p53 loss to drive a niche-independent program for self-renewal and proliferation. Moreover, these RNF43 variants confer decreased sensitivity to anti-Wnt-based therapy. Our data demonstrate the relevance of studying patient-derived mutations for understanding disease mechanisms and improved applications of precision medicine.

Central European Institute of Technology Masaryk University Brno Czech Republic

Department of Cell Biology and Oncode Institute Center for Molecular Medicine University Medical Center Utrecht Utrecht The Netherlands

Department of Experimental Biology Faculty of Science Masaryk University Brno Czech Republic

Department of Molecular Biology and Oncode Institute Faculty of Science Radboud Institute for Molecular Life Sciences Radboud University Nijmegen Nijmegen The Netherlands

Hubrecht Organoid Technology Utrecht The Netherlands

Institute of Molecular Biotechnology of the Austrian Academy of Sciences Vienna Austria

Laboratory for Experimental Oncology and Radiobiology and Oncode Institute Center for Experimental and Molecular Medicine Amsterdam UMC Cancer Center Amsterdam University of Amsterdam Amsterdam The Netherlands

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Targeting Oncogenic Pathways in the Era of Personalized Oncology: A Systemic Analysis Reveals Highly Mutated Signaling Pathways in Cancer Patients and Potential Therapeutic Targets

RNF43 inhibits WNT5A-driven signaling and suppresses melanoma invasion and resistance to the targeted therapy

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