UBE2K-FBXO11 complex activates Wnt signaling by stabilizing β-catenin through lysine 63-linked polyubiquitination

Abstract Β-catenin is regulated by phosphorylation and lysine (K)48-linked ubiquitination, which target it for proteasomal degradation. While the degradation machinery of β-catenin, including the destruction complex, has been extensively studied, regulators that stabilize β-catenin remain largely unexplored. A Wnt pathway double death trap (DDT) reporter system was established, followed by a genome-wide CRISPR screen that identified UBE2K and FBXO11 as novel positive regulators of Wnt signaling. Mechanistically, FBXO11 interacts with UBE2K to catalyze K63-linked polyubiquitination of β-catenin at residues K292 and K345, thereby enhancing its protein stability. Additionally, targeting the UBE2K-FBXO11 regulatory axis demonstrates a potent inhibitory effect on the growth of cancer cells driven by CTNNB1 mutations. This study reveals the UBE2K-FBXO11 complex as a previously unrecognized E2-E3 ubiquitination module that positively regulates β-catenin stability, thereby expanding the understanding of ubiquitin-mediated regulation of β-catenin beyond its canonical role in proteasomal degradation.