title: c-Abl phosphorylates E6AP and regulates its E3 ubiquitin ligase activity.
abstract: In human papillomavirus (HPV)-infected cells, the p53 tumor suppressor is tightly regulated by the HPV-E6-E6AP complex, which promotes it for proteasomal degradation. We previously demonstrated that c-Abl tyrosine kinase protects p53 from HPV-E6-E6AP complex-mediated ubiquitination and degradation under stress conditions. However, the underlying mechanism was not defined. In this study, we explored the possibility that c-Abl targets E6AP and thereby protects p53. We demonstrated that c-Abl interacts with and phosphorylates E6AP. We determined that the E3 ligase activity of E6AP is impaired in response to phosphorylation by c-Abl. We mapped the phosphorylation site to tyrosine 636 within the HECT catalytic domain of E6AP, and using substitution mutants, we showed that this residue dictates the E3 ligase activity of E6AP, in a substrate-specific manner. On the basis of the crystal structure of the HECT domain of E6AP, we propose a model in which tyrosine 636 plays a regulatory role in the oligomerization of E6AP, which is a process implicated in its E3 ubiquitin ligase activity. Our results suggest that c-Abl protects p53 from HPV-E6-E6AP complex-mediated degradation by phosphorylating E6AP and impairing its E3 ligase activity, thus providing a molecular explanation for the stress-induced protection of p53 in HPV-infected cells.
authors.affiliation: Research Division, The Peter MacCallum Cancer Centre, St. Andrew's Place, East Melbourne 3002, Victoria, Australia.
authors.forename: Ai-Leen
authors.initials: AL
authors.lastname: Chan
authors.forename: Tamar
authors.initials: T
authors.lastname: Grossman
authors.forename: Valentina
authors.initials: V
authors.lastname: Zuckerman
authors.forename: Dafne
authors.initials: D
authors.lastname: Campigli Di Giammartino
authors.forename: Ofra
authors.initials: O
authors.lastname: Moshel
authors.forename: Martin
authors.initials: M
authors.lastname: Scheffner
authors.forename: Brendon
authors.initials: B
authors.lastname: Monahan
authors.forename: Pat
authors.initials: P
authors.lastname: Pilling
authors.forename: Yong-Hui
authors.initials: YH
authors.lastname: Jiang
authors.forename: Sue
authors.initials: S
authors.lastname: Haupt
authors.forename: Ora
authors.initials: O
authors.lastname: Schueler-Furman
authors.forename: Ygal
authors.initials: Y
authors.lastname: Haupt
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D020134:Catalytic Domain; D002460:Cell Line; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D016315:Proto-Oncogene Proteins c-abl / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: C471639:UBE3A protein, human; D044767:Ubiquitin-Protein Ligases; D016315:Proto-Oncogene Proteins c-abl