Abstract B12: March E3 ubiquitin ligase is required for head formation by mediating Dishevelled degradation during Xenopus development

Wnt signaling controls numerous biological events including vertebrate development, adult tissue homeostasis and organogenesis. In addition, dysregulation of Wnt itself or its downstream signaling components causes human birth defects and cancer. Dishevelled (Dvl) is a crucial scaffolding protein that exerts activation of Wnt signaling pathway. Recently, several Dvl-interacting proteins have been suggested to mediate the stability of Dvl. However, the molecular mechanisms by which Dvl undergoes degradation remain to be elucidated. Here we identify March, a RING domain-containing ubiquitin ligase, targeting Dvl protein and show that it regulates canonical Wnt signaling in Xenopus . During Xenopus embryogenesis, March transcripts were ubiquitously expressed. We intriguingly found that March depletion by microinjection of morpholino oligonucleotide caused defects in the anterior head formation, including loss of the forebrain, cement gland and eye structures. We also found that these defective phenotypes were caused by the Wnt antagonistic function of March in the head organizer region. Knockdown of March suppressed expression of head organizer genes Gsc, Chd and Otx2 at the dorsal marginal region of the gastrula embryo. Wnt reporter assay, western blot analysis measuring phosphorylation of LRP6 and activation of β-catenin, RT-PCR and axis duplication assay in Xenopus revealed that March inhibits canonical Wnt signaling pathway. Furthermore, we identified that March protein associates with Dvl and mediates its poly-ubiquitination for lysosomal degradation in Xenopus embryos. Taken together, these results suggest that March retains cytosolic pools of Dvl protein and leads subsequent limitation of Wnt signaling for vertebrate head development. Citation Format: Hyeyoon Lee, Seong-Moon Cheong, Jin-Kwan Han. March E3 ubiquitin ligase is required for head formation by mediating Dishevelled degradation during Xenopus development. [abstract]. In: Proceedings of the AACR Special Conference: Developmental Biology and Cancer; Nov 30-Dec 3, 2015; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(4_Suppl):Abstract nr B12.