EXPLORATION OF UBE3A IN WNT SIGNALING AND NEURAL CREST DEVELOPMENT

Abstract

UBE3A is an E3 ubiquitin ligase that was first discovered due to its association with HPV’s E6 oncoprotein in inhibiting apoptosis through the degradation of p53 in cancerous states. Aside from UBE3A’s implication in cancer, it has also been determined to be the causative gene of Angelman Syndrome (AS), a rare neurodevelopmental and genetic disorder. Additionally, it has been documented that patients with AS have hypopigmentation and craniofacial defects, which are classic symptoms of neurocristopathies and are caused by a dysregulation of neural crest (NC) development. While many signaling pathways are required for proper development, one of the most imperative is canonical Wnt signaling. Wnt signaling begins to play a role in development as soon as fertilization occurs, with cortical rotation, and remains integral as NC induction, migration, and differentiation occur. UBE3A has been shown to activate Wnt signaling, and neurocristopathies often arise from a loss of Wnt signaling during development. Therefore, we hypothesize that loss of UBE3A leads to neural crest defects via Wnt signaling. Here, we show that Ube3a is localized to the NC in Xenopus tropicalis during development, and additionally discovered that Ube3a is a highly abundant maternally deposited protein, which prevented us from knocking it down or out. Moving to an in vitro model of the HEK293STF cell line, we confirmed that UBE3A activates Wnt signaling in a dose-dependent manner. Collaborating with clinicians, have identified a novel cystine to tryptophan mutation (C843W) in UBE3A mutation in an AS patient, and we found that the C843 residue is localized in the active site and likely required for proper E3 ligase activity. We have determined that this mutation makes the protein less stable than its wild-type counterpart and unable to induce Wnt signaling. Future efforts will be focused on a new knock-in mouse line harboring the patient-derived mutation and germ-line knockout required for developmental studies in X. tropicalis.