Neutralization sites of human papillomavirus-6 relate to virus attachment and entry phase in viral infection

Abstract

Human papillomavirus type 6 (HPV6) is the major etiologic agent of genital warts and recurrent respiratory papillomatosis. Although the commercial HPV vaccines cover HPV6, the neutralization sites and mode for HPV6 are poorly understood. Here, we identify the HPV6 neutralization sites and discriminate the inhibition of virus attachment and entry by three potent neutralizing antibodies (nAbs), 5D3, 17D5, and 15F7. Mutagenesis assays showed that these nAbs predominantly target surface loops BC, DE, and FG of HPV6 L1. Cryo-EM structures of the HPV6 pseudovirus (PsV) and its immune complexes revealed three distinct binding modalities – full-occupation-bound to capsid, top-center-bound-, and top-rim-bound to pentamers – and illustrated a structural atlas for three classes of antibody-bound footprints that are located at center-distal ring, center, and center-proximal ring of pentamer surface for 5D3, 17D5, and 15F7, respectively. Two modes of neutralization were identified: mAb 5D3 and 17D5 block HPV PsV from attaching to the extracellular matrix (ECM) and the cell surface, whereas 15F7 allows PsV attachment but prohibits PsV from entering the cell. These findings highlight three neutralization sites of HPV6 L1 and outline two antibody-mediated neutralization mechanisms against HPV6, which will be relevant for HPV virology and antiviral inhibitor design. HighlightsMajor neutralization sites of HPV6 were mapped on the pseudovirus cryo-EM structuremAb 15F7 binds HPV6 capsid with a novel top-rim binding modality and confers a post-attachment neutralizationmAb 17D5 binds capsid in top-centre manner but unexpectedly prevents virus from attachment to cell surface

Structural Basis for the Shared Neutralization Mechanism of Three Classes of Human Papillomavirus Type 58 Antibodies with Disparate Modes of Binding

“…8A and D). A similar binding mode was also previously reported for HPV11 and in our previous structural study of HPV58 and HPV6 (12,17,36).…”

“…Comprehensive neutralization epitopes are vital for an in-depth understanding of HPV infection mechanisms and for virology research. Generally, the five surface loops (BC, DE, EF, FG, and HI) of the HPV L1 protein compose the major antigenic epitopes of HPV, and this knowledge has been widely confirmed in the literature, for example, for HPV16 (38,40,41), HPV31 (42), HPV33 (43), and HPV6 (16,17). In addition, the C-terminal arm of L1 serves as a neutralizing epitope, and this has been validated with loopswapping analysis and high-resolution cryo-EM structure determination (11).…”

“…U4) have been generated and are extensively used in HPV vaccine research and related serological assays (13,14). Biochemical and structural analyses have mapped the neutralizing binding sites for HPV6, -11, -16, -31, -33, -52, -58, and -59 to the hypervariable surface loops BC, DE, EF, FG, and HI of the L1 major capsid protein, as well as the C-terminal arm (8,11,12,(15)(16)(17). For HPV58, although several antibody panels have been developed (18), detailed information on the neutralization sites remain unknown.…”

Structural characterization of a neutralizing mAb H16.001, a potent candidate for a common potency assay for various HPV16 VLPs

“…To date, there are several reported cryo-EM structures of immune complex containing HPV PsV, wherein the highest resolution was only at 4.7 Å and could not afford a confident model building at near atomic level 16 , 19 – 21 . We circumvented the resolution bottleneck over 4 Å resolution for HPV capsid immune complex through new emerging subparticle approach.…”

Cryo EM Analysis Reveals Inherent Flexibility of Authentic Murine Papillomavirus Capsids

“…The complete T = 7d icosahedral papillomavirus capsid has previously been visualized to modest resolution by cryogenic electron microscopy (cryo EM) with icosahedral symmetry imposed during the reconstruction [ 4 , 8 , 9 , 10 , 11 , 12 , 13 , 14 ]. No structural studies of papillomavirus capsids with 60-fold icosahedral symmetry imposed have exceeded 4.1–4.4 Å resolution [ 13 , 14 , 15 , 16 ]. The structure of HPV16 QV published by Guan et al at 4.3 Å showed there is likely inherent flexibility of the papillomavirus capsid as evidenced by the variability in capsid resolution, and differing capsid diameters [ 13 ].…”