CEP78 is a centrosomal protein implicated in ciliogenesis and ciliary length control, and mutations in the CEP78 gene cause retinal cone-rod dystrophy associated with hearing loss. However, the mechanism by which CEP78 affects cilia formation is unknown. Based on a recently discovered disease-causing CEP78 p.L150S mutation, we identified the disease-relevant interactome of CEP78. We confirmed that CEP78 interacts with the EDD1 -DYRK2-DDB1VPRBP E3 ubiquitin lig- ase complex, which is involved in CP110 ubiquitination and degradation, and identified a novel interaction between CEP78 and CEP350 that is weakened by the CEP78L150S mutation. We show that CEP350 promotes centrosomal recruitment and stability of CEP78, which in turn leads to cen- trosomal recruitment of EDD 1. Consistently, cells lacking CEP78 display significantly increased cellular and centrosomal levels of CP110, and depletion of CP110 in CEP78-deficient cells restored ciliation frequency to normal. We propose that CEP78 functions downstream of CEP350 to pro- mote ciliogenesis by negatively regulating CP110 levels via an EDD 1-dependent mechanism.
Bibliografisk noteFunding Information:
We thank Søren Lek Johansen and Maria Schrøder Holm for expert technical assistance, Benedicte Schultz Kappel and Frida Roikjer Rasmussen for help with plasmid generation, and Nynne Chris-tensen, Center for Advanced Bioimaging (CAB) Denmark, for help with 3D-SIM acquisition. Dr. Kay Schink kindly provided plasmids for lentivirus particle generation and assisted in 3D-SIM acquisition at OUH Radiumhospitalet. We are grateful to Drs. Peter K. Jackson, Tomoharu Kanie, Francesc Garcia-Gonzalo, Laurence Pelletier, Brian David Dynlacht, Anne-Marie Tassin, Maria Gavilan, Rosa M. Rios, Éric A. Cohen and Peter ten Dijke for reagents, and Dr. Michel Bornens for sharing original EM microcraphs. This study was supported grants from Independent Research Fund Denmark (#8020‐00162B to PF and LBP, and #8021-00425A to JSA), from the Carlsberg Foundation (#CF18‐0294 to LBP) and from the Swiss National Science Foundation (#176097 to CR).
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