Dishevelled (DVL) is a key scaffolding protein and a branching point in Wnt signaling pathways. Here, we present conclusive evidence that DVL regulates the centrosomal cycle. We demonstrate that DVL dishevelled and axin (DIX) domain, but not DIX domain-mediated multimerization, is essential for DVL's centrosomal localization. DVL accumulates during the cell cycle and associates with NIMA-related kinase 2 (NEK2), which is able to phosphorylate DVL at a multitude of residues, as detected by a set of novel phospho-specific antibodies. This creates interfaces for efficient binding to CDK5 regulatory subunitassociated protein 2 (CDK5RAP2) and centrosomal Nek2-associated protein 1 (C-NAP1), two proteins of the centrosomal linker. Displacement of DVL from the centrosome and its release into the cytoplasm on NEK2 phosphorylation is coupled to the removal of linker proteins, an event necessary for centrosomal separation and proper formation of the mitotic spindle. Lack of DVL prevents NEK2-controlled dissolution of loose centrosomal linker and subsequent centrosomal separation. Increased DVL levels, in contrast, sequester centrosomal NEK2 and mimic monopolar spindle defects induced by a dominant negative version of this kinase. Our study thus uncovers molecular crosstalk between centrosome and Wnt signaling.Wnt signaling | centrosome | Dishevelled | NEK2 | linker proteins

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.

Our findings expand current knowledge of TTBK2 substrates and provide insight into extent of TTBK2-mediated phosphorylations in the context of basal body proteins and PC initiation. We further show that phosphorylation of CEP164 N-term by TTBK2 regulates ciliogenesis.