Insights into the Novel Adaptor-Independent Apoptosis Mediated by Human Papillomavirus E2 Protein

Abstract

Early protein E2 of human papillomaviruses (HPV), that are associated with cervical and anogenital cancers, regulates viral DNA replication and transactivation of essential viral oncogenes. Apart from these functions, E2 protein from high risk virus types such as HPV-16 and −18 triggers apoptosis in the host cell. Although the exact mechanism is unclear, recent literature suggests that in HPV-18 E2, the N-terminal transactivation domain directly interacts with procaspase-8, a component of Death Inducing Signaling Complex (DISC) in the extrinsic cell death pathway. This interaction bypasses the requirement of upstream adaptor proteins which are essentially required for DISC formation, thereby representing a novel adaptor-independent caspase activation pathway. In this work, we dissected the binding interface of E2-procasapse-8 interaction using an interdisciplinary approach employing techniques such as in silico, mutational, biochemical and biophysical analyses. In vitro pull-down and co-expression studies show that E2 specifically interacts with procaspase-8 death effector domain (DED) B. We further delineated the minimal binding region in DED B using different deletion constructs. Based upon docking analyses, site directed mutagenesis of E2 was carried out and critical residues involved in this protein-protein interaction were identified. Our results provide a molecular basis of this novel E2-procaspase-8 interaction and help in providing a model for E2-induced apoptosis in high risk HPV types. This information may be utilized in future studies to design E2 analogs so as to modulate procaspase-8 activation and hence promote apoptosis.