Papillomavirus Infection Requires γ Secretase

Abstract

The mechanism by which papillomaviruses breach cellular membranes to deliver their genomic cargo to the nucleus is poorly understood. Here, we show that infection by a broad range of papillomavirus types requires the intramembrane protease ␥ secretase. The ␥-secretase inhibitor (S, The necessary causal association of persistent infection by an "oncogenic" type of human papillomavirus (HPV) with cervical cancer is firmly established (52,53). HPV is the most prevalent sexually transmitted infection, and although the majority of patients clear their infection, HPV is directly responsible for 5% of all cancer deaths worldwide (30). HPV is also associated with multiple other anogenital cancers and oropharyngeal cancers.SThe life cycle of HPV is closely linked to epithelial differentiation within stratified squamous epithelia (16). Initial infection occurs within the undifferentiated proliferative basal cell layer in which only the viral early proteins are expressed, whereas production of the late proteins and, thus, progeny virus is restricted to the terminally differentiated suprabasal compartment (53). The exquisite dependence of virion production upon epithelial differentiation and lack of a rapid phenotype in culture can be circumvented by ectopic expression of the capsid proteins L1 and L2 in cells maintaining viral genome or reporter constructs as episomes, resulting in "quasivirions" or "pseudovirions," respectively, whose infectivity can be readily and rapidly quantified in vitro or in vivo (6,11,35,41).The completion of the entire papillomavirus life cycle is species specific. However, studies with bovine papillomavirus (BPV) in horses and hamsters, HPV pseudovirions in mouse challenge models, and quasivirions in rabbits suggest that virion internalization and delivery of the encapsidated DNA to the nucleus are promiscuous and that tropism is determined at a later stage of the life cycle (11,27,29,39).Although significant progress has been made in understanding the HPV life cycle and virion structure, many of the molecular events of virus internalization and infection are poorly defined (43). Both the L1 (major) and L2 (minor) capsid proteins provide essential functions during infection (41) (8). L1 is sufficient to form empty capsids, termed virus-like particles (VLPs) (25), which bind to basement membrane and to the cell surface and which also form the basis of the licensed HPV vaccines (10). Glycosaminoglycans (GAGs), most notably heparan sulfate (HS), play a critical role in virion binding and infection, both in vitro and in the murine vaginal challenge model, although differences between HPV types and target cells in vitro have been described (14,19,20), for example, between HPV16 and HPV31 (4,34,42). Once bound to the basement membrane, the virions undergo a conformation change resulting in the surface display of the amino terminus of L2 and its cleavage by a proprotein convertase (PC), furin and/or PC5/PC6, and the transfer of virions to the cell surface (24). The uptake of the virions is apparently ...

Genome-wide siRNA screen identifies the retromer as a cellular entry factor for human papillomavirus

“…However, some of the hits-furin, the γ-secretase complex, and the vacuolar ATPase complex (which causes endosome acidification)-were reported by others to be required for HPV infection or are consistent with our prior understanding of this process (17)(18)(19)(20). The identification of these genes validated our screen design and suggested that our findings were relevant to HPV16 infection.…”

“…HPV is then thought to be transferred to an as-yetunidentified cell-surface receptor, followed by endocytosis and intracellular trafficking (8)(9)(10)(11)(12)(13)(14)(15). Cyclophilin B and the proteases furin and γ-secretase play essential but not clearly understood roles during HPV entry (16)(17)(18)(19). After HPV is internalized, capsid disassembly is initiated in the endosome by acidification (11,15,20).…”

Multiple Heparan Sulfate Binding Site Engagements Are Required for the Infectious Entry of Human Papillomavirus Type 16

“…Before we investigated the defect of these mutant PsVs in more detail, we tested whether the mutant viruses follow the same internalization pathway as wt HPV16. To test this, HeLa cells were infected with wt and mutant PsVs in the presence of well-established inhibitors of HPV16 infection targeting CyPs, ␥-secretase, tyrosine kinases, and endosomal acidification and trafficking (38,46,(48)(49)(50)(51)(52)(53)). All mutant PsVs tested displayed the same inhibition profile as wt HPV16 (Fig.…”

A Transmembrane Domain and GxxxG Motifs within L2 Are Essential for Papillomavirus Infection

“…A55 is predicted to be the most deleterious mutation, disturbing all the TM domain GxxxG motifs. A60 preserves the 52 GxxxG 56 motif but disrupts all downstream motifs, and A64 disrupts only the C-terminal 61 GxxxG 65 and 63 GxxxG 67 motifs. The A55 mutant was completely noninfectious, the A60 mutant had drastically reduced infectivity, and the A64 mutant had no reduction in infectivity (Fig.…”

“…Lastly, the intramembrane protease complex, ␥-secretase, has been shown to be absolutely critical for HPV infection (61). Biochemical inhibition of ␥-secretase potently blocks HPV16 infection, again by preventing the endosomal translocation of vDNA.…”