FAM83F regulates canonical Wnt signalling through an interaction with CK1α

Abstract

The function of the FAM83F protein, like the functions of many members of the FAM83 family, is poorly understood. Here we show that injection of Fam83f mRNA into Xenopus embryos causes axis duplication, a phenotype indicative of enhanced Wnt signalling.Consistent with this, overexpression of FAM83F activates Wnt signalling, whilst ablation of FAM83F from human colorectal cancer (CRC) cells attenuates it. We demonstrate that FAM83F is farnesylated and interacts and co-localises with CK1α at the plasma membrane.This interaction with CK1α is essential for FAM83F to activate Wnt signalling, and FAM83F mutants that do not interact with CK1α fail to induce axis duplication in Xenopus embryos and to activate Wnt signalling in cells. FAM83F acts upstream of the β-catenin destruction complex, because the attenuation of Wnt signalling caused by loss of FAM83F can be rescued by GSK-3 inhibition. Introduction of a farnesyl-deficient mutant mis-localises the FAM83F-CK1α complex to the nucleus and significantly attenuates Wnt signalling, indicating that FAM83F exerts its effects on Wnt signalling at the plasma membrane.

IMiDs induce FAM83F degradation via an interaction with CK1α to attenuate Wnt signalling

“…Endogenous FAM83F is predominately located at the plasma membrane as observed by immunofluorescence of HCT116 cells in which a GFP tag was knocked in homozygously at the N-terminus of the FAM83F gene ( GFP/GFP FAM83F cells) ( Figs 2A , S3A , and S4A ) ( Dunbar et al, 2020 ). Upon treatment of HCT116 GFP/GFP FAM83F cells with pomalidomide, the GFP signal was lost from the plasma membrane ( Fig 2A ).…”

“…When DLD-1 wild-type cells were treated with pomalidomide, a reduction in the levels of both FAM83F and CK1α protein was observed in the membrane fraction, whereas CK1α protein levels in the cytoplasmic and nuclear fractions did not change relative to untreated DLD-1 wild-type cells ( Fig 2B ). FAM83F interacts with CK1α and is responsible for delivering it to the plasma membrane ( Dunbar et al, 2020 ). Interestingly, the pomalidomide-induced reduction of CK1α protein from the plasma membrane in DLD-1 wild-type cells was comparable with the stable reduction of membranous CK1α observed in FAM83F-knockout (FAM83F −/− ) DLD-1 cells, generated using CRISPR/Cas9 genome editing, in the absence of pomalidomide treatment ( Figs 2B , S3A , and S4B ).…”

“…Several conserved residues within the DUF1669 domains of FAM83 proteins have been identified as critical mediators of the FAM83-CK1 interaction ( Fulcher et al, 2018 ). For FAM83F, mutation of two phenylalanine residues at positions 284 and 288 to alanine would be predicted to abolish association with CK1α ( Dunbar et al, 2020 ). As expected, FAM83F co-precipitated with CK1α in CK1α immunoprecipitates (IPs) from HCT116 wild-type cells but not from FAM83F-knockout (FAM83F −/− ) cells generated using CRISPR/Cas9 ( Figs 3A , S3A , and S4B ).…”

“…We have recently established that FAM83F mediates canonical Wnt signalling through association with CK1α ( Dunbar et al, 2020 ). Ablation of FAM83F significantly inhibits Wnt signalling in multiple cell lines.…”

“…All FAM83 proteins interact with CK1α, whereas FAM83A, B, E, and H also interact with CK1δ and ε ( Fulcher et al, 2018 ). Whereas the FAM83 family remains largely uncharacterised, roles for specific FAM83–CK1α complexes have been established in mitosis ( Fulcher et al, 2019 ; Fulcher & Sapkota, 2020 ) and canonical Wnt signalling ( Bozatzi et al, 2018 ; Wu et al, 2019 ; Dunbar et al, 2020 ). Given the reports of IMiD-induced degradation of CK1α, we sought to establish the effect of IMiDs on the stability of FAM83 proteins and different FAM83–CK1α complexes.…”