Proteomic analysis identifies the E3 ubiquitin ligase Pdzrn3 as a regulatory target of Wnt5a-Ror signaling

Wnt5a-Ror signaling is a conserved pathway that regulates morphogenetic processes during vertebrate development, but its downstream signaling events remain poorly understood. By conducting a large-scale proteomic screen in mouse embryonic fibroblasts, we identified the E3 ubiquitin ligase Pdzrn3 as a new regulatory target that is degraded upon pathway activation in a β-catenin-independent, ubiquitin-proteasome system-dependent manner. We developed a flow cytometry-based reporter to monitor Pdzrn3 abundance and delineated a signaling cascade involving Frizzled, Dishevelled, Casein kinase 1, and Glycogen synthase kinase 3 that regulates Pdzrn3 stability. Genetic epistasis analysis suggests that Pdzrn3 degradation occurs downstream of Dishevelled but independently of Kif26b, a previously identified Wnt5a-Ror-Dishevelled signaling target. Further, we discovered that Pdzrn3 degradation requires Wnt5a-dependent phosphorylation of three residues within its C-terminal LNX3H domain, which is conserved in other homologs and likely functions as a Wnt5a-responsive domain. Collectively, this work establishes a new Wnt5a-Ror signaling cascade involving Pdzrn3 phosphorylation and degradation.