title: Resistance to UV-induced apoptosis by β-HPV5 E6 involves targeting of activated BAK for proteolysis by recruitment of the HERC1 ubiquitin ligase.
abstract: UV exposure is the main etiological agent in the development of non-melanoma skin cancer (NMSC), but mounting evidence suggests a co-factorial role for β-genus HPV types early in tumor initiation or progression. UV damage initiates an apoptotic response, driven at the mitochondrial level by BCL-2 family proteins, that eliminates damaged cells that may accumulate deleterious mutations and acquire tumorigenic properties. BAK is a pro-apoptotic BCL-2 protein that functions ultimately to form pores that permeabilize the mitochondrial outer membrane, thereby committing a cell to death, a process involving changes in BAK phosphorylation and conformation. The E6 protein of β-type HPV5 signals BAK for proteasomal degradation, a function that confers protection from UV-induced apoptosis. We find that HPV5 E6 does not constitutively target BAK for proteolysis, but targets the latter stages of BAK activation, following changes in phosphorylation and conformation. A mutational analysis identified the lysine residue on BAK required for proteolysis, and a functional siRNA screen identified the HECT domain E3 ubiquitin ligase HERC1 as being required for E6-mediated BAK degradation. We show that HERC1 interacts with BAK in E6-expressing cells that have been damaged by UV, and provide evidence that the interaction of HERC1 with BAK requires access to a hydrophobic surface on BAK that binds BH3 domains of BCL-2 proteins. We also show that HERC1 contains a putative BH3 domain that can bind to BAK. These findings reveal a specific and unique mechanism used by the HPV5 E6 protein to target BAK.
authors.affiliation: Department of Oncology, Weatherall Institute of Molecular Medicine,University of Oxford, OX3 9DS, United Kingdom.
authors.forename: Amy
authors.initials: A
authors.lastname: Holloway
authors.forename: Mark
authors.initials: M
authors.lastname: Simmonds
authors.forename: Abul
authors.initials: A
authors.lastname: Azad
authors.forename: Joanna L
authors.initials: JL
authors.lastname: Fox
authors.forename: Alan
authors.initials: A
authors.lastname: Storey
mesh_terms: D017209:Apoptosis / Q000528:radiation effects*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D005434:Flow Cytometry; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism*; D045325:HCT116 Cells; D006801:Humans; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000528:radiation effects; D011485:Protein Binding; D059748:Proteolysis / Q000528:radiation effects; D034622:RNA Interference; D044767:Ubiquitin-Protein Ligases; D014466:Ultraviolet Rays; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: C487708:E6 protein, human papillomavirus type 5; D020662:Guanine Nucleotide Exchange Factors; D009856:Oncogene Proteins, Viral; D050998:bcl-2 Homologous Antagonist-Killer Protein; C102169:HERC1 protein, human; D044767:Ubiquitin-Protein Ligases; D008239:Lysine
keywords: BAK; HERC1; HPV; apoptosis