Post-Translational Control of IL-1β via the Human Papillomavirus Type 16 E6 Oncoprotein: A Novel Mechanism of Innate Immune Escape Mediated by the E3-Ubiquitin Ligase E6-AP and p53

Abstract

Infections with high-risk human papillomaviruses (HPVs) are causally involved in the development of anogenital cancer. HPVs apparently evade the innate immune response of their host cells by dysregulating immunomodulatory factors such as cytokines and chemokines, thereby creating a microenvironment that favors malignancy. One central key player in the immune surveillance interactome is interleukin-1 beta (IL-1β) which not only mediates inflammation, but also links innate and adaptive immunity. Because of its pleiotropic physiological effects, IL-1β production is tightly controlled on transcriptional, post-translational and secretory levels. Here, we describe a novel mechanism how the high-risk HPV16 E6 oncoprotein abrogates IL-1β processing and secretion in a NALP3 inflammasome-independent manner. We analyzed IL-1β regulation in immortalized keratinocytes that harbor the HPV16 E6 and/or E7 oncogenes as well as HPV-positive cervical tumor cells. While in primary and in E7-immortalized human keratinocytes the secretion of IL-1β was highly inducible upon inflammasome activation, E6-positive cells did not respond. Western blot analyses revealed a strong reduction of basal intracellular levels of pro-IL-1β that was independent of dysregulation of the NALP3 inflammasome, autophagy or lysosomal activity. Instead, we demonstrate that pro-IL-1β is degraded in a proteasome-dependent manner in E6-positive cells which is mediated via the ubiquitin ligase E6-AP and p53. Conversely, in E6- and E6/E7-immortalized cells pro-IL-1β levels were restored by siRNA knock-down of E6-AP and simultaneous recovery of functional p53. In the context of HPV-induced carcinogenesis, these data suggest a novel post-translational mechanism of pro-IL-1β regulation which ultimately inhibits the secretion of IL-1β in virus-infected keratinocytes. The clinical relevance of our results was further confirmed in HPV-positive tissue samples, where a gradual decrease of IL-1β towards cervical cancer could be discerned. Hence, attenuation of IL-1β by the HPV16 E6 oncoprotein in immortalized cells is apparently a crucial step in viral immune evasion and initiation of malignancy.

Lipopolysaccharide Primes the NALP3 Inflammasome by Inhibiting Its Ubiquitination and Degradation Mediated by the SCFFBXL2 E3 Ligase

“…However, recent studies suggest that assembly of the NALP3 inflammasome to activate caspase 1 is regulated mainly through nontranscriptional processes (14,15). Furthermore, the final products of the activated NALP3 inflammasome, IL-1␤ and IL-18, are possibly regulated at posttranslational levels, suggesting concerted actions of modulators through this mode of regulatory control (16,17).…”

Regulation of inflammasomes by ubiquitination

“…HPV16 harboring the E6 oncogene encodes the ubiquitin ligase E6-AP, which increases the ubiquitination and proteasomal degradation of pro-IL-1β. 54 This exemplifies a possible therapeutic strategy to reduce IL-1β signaling by targeting ubiquitin.…”

Inflammasomes and its importance in viral infections

“…Additionally, papilloma virus has been shown to possess a post-translational mechanism to regulate production of IL-1b; however, this process is regulated independently of NLRP3 [130]. Notably, this phenomenon is promoted by the E6 oncoprotein that activates the proteasome pathway and culminates in degradation of pro-IL-1b [131]. It is relevant to mention that many viral proteins can translocate into the nucleus of infected cells, probably to regulate the expression of genes involved in immunological processes Inflammasome complex activation occurs through a two-step process, which is initiated by the stimulation of inflammasome components, leading to the production of pro-IL-1b and pro-IL-18 through NF-jB activation, followed by a final signal that triggers inflammasome complex activation resulting in caspase-1 catalysis.…”