Splice variants of CK1α and CK1α-like: Comparative analysis of subcellular localization, kinase activity, and function in the Wnt signaling pathway

• Published in: The Journal of biological chemistry 2024-07, Vol.300 (7), p.107407, Article 107407
• Main Authors: Gybeľ, Tomáš, Čada, Štěpán, Klementová, Darja, Schwalm, Martin P., Berger, Benedict-Tilman, Šebesta, Marek, Knapp, Stefan, Bryja, Vítězslav
• Format: Article
• Language: English
• Subjects: Alternative Splicing; Axin; beta Catenin - genetics; beta Catenin - metabolism; casein kinase 1 alpha (CK1α); casein kinase 1 alpha-like (CK1α-like); Casein Kinase Ialpha - genetics; Casein Kinase Ialpha - metabolism; gene knockout; Glycogen Synthase Kinase 3 - genetics; Glycogen Synthase Kinase 3 - metabolism; Glycogen Synthase Kinase 3 beta - genetics; Glycogen Synthase Kinase 3 beta - metabolism; HEK293 Cells; Humans; inhibitor; NanoBRET; Phosphorylation; Wnt pathway; Wnt Signaling Pathway; Wnt3A Protein - genetics; Wnt3A Protein - metabolism; β-catenin
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1016/j.jbc.2024.107407

Members of the casein kinase 1 (CK1) family are important regulators of multiple signaling pathways. CK1α is a well-known negative regulator of the Wnt/β-catenin pathway, which promotes the degradation of β-catenin via its phosphorylation of Ser45. In contrast, the closest paralog of CK1α, CK1α-like, is a poorly characterized kinase of unknown function. In this study, we show that the deletion of CK1α, but not CK1α-like, resulted in a strong activation of the Wnt/β-catenin pathway. Wnt-3a treatment further enhanced the activation, which suggests there are at least two modes, a CK1α-dependent and Wnt-dependent, of β-catenin regulation. Rescue experiments showed that only two out of ten naturally occurring splice CK1α/α-like variants were able to rescue the augmented Wnt/β-catenin signaling caused by CK1α deficiency in cells. Importantly, the ability to phosphorylate β-catenin on Ser45 in the in vitro kinase assay was required but not sufficient for such rescue. Our compound CK1α and GSK3α/β KO models suggest that the additional nonredundant function of CK1α in the Wnt pathway beyond Ser45-β-catenin phosphorylation includes Axin phosphorylation. Finally, we established NanoBRET assays for the three most common CK1α splice variants as well as CK1α-like. Target engagement data revealed comparable potency of known CK1α inhibitors for all CK1α variants but not for CK1α-like. In summary, our work brings important novel insights into the biology of CK1α, including evidence for the lack of redundancy with other CK1 kinases in the negative regulation of the Wnt/β-catenin pathway at the level of β-catenin and Axin.