title: Maturation of the human papillomavirus 16 capsid.
abstract: Papillomaviruses are a family of nonenveloped DNA viruses that infect the skin or mucosa of their vertebrate hosts. The viral life cycle is closely tied to the differentiation of infected keratinocytes. Papillomavirus virions are released into the environment through a process known as desquamation, in which keratinocytes lose structural integrity prior to being shed from the surface of the skin. During this process, virions are exposed to an increasingly oxidative environment, leading to their stabilization through the formation of disulfide cross-links between neighboring molecules of the major capsid protein, L1. We used time-lapse cryo-electron microscopy and image analysis to study the maturation of HPV16 capsids assembled in mammalian cells and exposed to an oxidizing environment after cell lysis. Initially, the virion is a loosely connected procapsid that, under in vitro conditions, condenses over several hours into the more familiar 60-nm-diameter papillomavirus capsid. In this process, the procapsid shrinks by ~5% in diameter, its pentameric capsomers change in structure (most markedly in the axial region), and the interaction surfaces between adjacent capsomers are consolidated. A C175S mutant that cannot achieve normal inter-L1 disulfide cross-links shows maturation-related shrinkage but does not achieve the fully condensed 60-nm form. Pseudoatomic modeling based on a 9-Å resolution reconstruction of fully mature capsids revealed C-terminal disulfide-stabilized "suspended bridges" that form intercapsomeric cross-links. The data suggest a model in which procapsids exist in a range of dynamic intermediates that can be locked into increasingly mature configurations by disulfide cross-linking, possibly through a Brownian ratchet mechanism. Importance: Human papillomaviruses (HPVs) cause nearly all cases of cervical cancer, a major fraction of cancers of the penis, vagina/vulva, anus, and tonsils, and genital and nongenital warts. HPV types associated with a high risk of cancer, such as HPV16, are generally transmitted via sexual contact. The nonenveloped virion of HPVs shows a high degree of stability, allowing the virus to persist in an infectious form in environmental fomites. In this study, we used cryo-electron microscopy to elucidate the structure of the HPV16 capsid at different stages of maturation. The fully mature capsid adopts a rigid, highly regular structure stabilized by intermolecular disulfide bonds. The availability of a pseudoatomic model of the fully mature HPV16 virion should help guide understanding of antibody responses elicited by HPV capsid-based vaccines.
authors.affiliation: Laboratory of Structural Biology, National Institute for Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health (NIH), Bethesda, Maryland, USA.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Cardone
authors.affiliation: Laboratory of Cellular Oncology, National Cancer Institute, NIH, Bethesda, Maryland, USA.
authors.forename: Adam L
authors.initials: AL
authors.lastname: Moyer
authors.affiliation: Laboratory of Structural Biology, National Institute for Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health (NIH), Bethesda, Maryland, USA.
authors.forename: Naiqian
authors.initials: N
authors.lastname: Cheng
authors.affiliation: Laboratory of Cellular Oncology, National Cancer Institute, NIH, Bethesda, Maryland, USA.
authors.forename: Cynthia D
authors.initials: CD
authors.lastname: Thompson
authors.affiliation: Yale University, New Haven, Connecticut, USA.
authors.forename: Israel
authors.initials: I
authors.lastname: Dvoretzky
authors.affiliation: Laboratory of Cellular Oncology, National Cancer Institute, NIH, Bethesda, Maryland, USA.
authors.forename: Douglas R
authors.initials: DR
authors.lastname: Lowy
authors.affiliation: Laboratory of Cellular Oncology, National Cancer Institute, NIH, Bethesda, Maryland, USA.
authors.forename: John T
authors.initials: JT
authors.lastname: Schiller
authors.affiliation: Laboratory of Structural Biology, National Institute for Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health (NIH), Bethesda, Maryland, USA.
authors.forename: Alasdair C
authors.initials: AC
authors.lastname: Steven
authors.affiliation: Laboratory of Cellular Oncology, National Cancer Institute, NIH, Bethesda, Maryland, USA.
authors.forename: Christopher B
authors.initials: CB
authors.lastname: Buck
authors.affiliation: Imaging Sciences Laboratory, Center for Information Technology, NIH, Bethesda, Maryland, USA trusb@mail.nih.gov.
authors.forename: Benes L
authors.initials: BL
authors.lastname: Trus
mesh_terms: D036022:Capsid Proteins / Q000648:ultrastructure*; D002460:Cell Line; D020285:Cryoelectron Microscopy; D052162:Human papillomavirus 16 / Q000254:growth & development* / Q000648:ultrastructure*; D006801:Humans; D017433:Protein Structure, Secondary; D014771:Virion / Q000648:ultrastructure
chemical_list: D036022:Capsid Proteins
references.citation: Cell. 1998 Jul 10;94(1):55-60
references.pmid: 9674427
references.citation: Adv Exp Med Biol. 2012;726:403-22
references.pmid: 22297524
references.citation: J Comput Chem. 2005 Dec;26(16):1781-802
references.pmid: 16222654
references.citation: Curr Protoc Cell Biol. 2007 Dec;Chapter 26:Unit 26.1
references.pmid: 18228512
references.citation: EMBO J. 2002 Sep 16;21(18):4754-62
references.pmid: 12234916
references.citation: Curr Opin Struct Biol. 2005 Apr;15(2):227-36
references.pmid: 15837183
references.citation: EMBO J. 2005 Apr 6;24(7):1352-63
references.pmid: 15775971
references.citation: J Pharm Sci. 2005 Jul;94(7):1538-51
references.pmid: 15929070
references.citation: Nat Struct Biol. 2003 May;10(5):334-41
references.pmid: 12704429
references.citation: J Virol. 1997 Aug;71(8):6247-52
references.pmid: 9223527
references.citation: Proc Natl Acad Sci U S A. 2010 Apr 6;107(14):6298-303
references.pmid: 20308582
references.citation: Int J Cancer. 2012 Nov 1;131(9):1969-82
references.pmid: 22623137
references.citation: J Virol. 2005 Apr;79(7):3938-48
references.pmid: 15767396
references.citation: J Virol. 2007 Aug;81(16):8784-92
references.pmid: 17553881
references.citation: Clin Dermatol. 1997 May-Jun;15(3):309-19
references.pmid: 9255438
references.citation: J Mol Graph. 1996 Feb;14(1):33-8, 27-8
references.pmid: 8744570
references.citation: J Virol. 1998 Mar;72(3):2160-7
references.pmid: 9499072
references.citation: J Comput Chem. 2004 Oct;25(13):1605-12
references.pmid: 15264254
references.citation: J Virol. 1993 Jan;67(1):315-22
references.pmid: 8380079
references.citation: Mol Cell. 2000 Mar;5(3):557-67
references.pmid: 10882140
references.citation: Gynecol Oncol. 2010 Jun;118(1 Suppl):S12-7
references.pmid: 20494219
references.citation: J Clin Virol. 2005 Mar;32 Suppl 1:S7-15
references.pmid: 15753007
references.citation: J Natl Cancer Inst Monogr. 2003;(31):14-9
references.pmid: 12807940
references.citation: J Virol. 2005 Mar;79(5):2839-46
references.pmid: 15709003
references.citation: J Virol. 2003 Nov;77(21):11625-32
references.pmid: 14557648
references.citation: Nat Struct Biol. 1997 May;4(5):413-20
references.pmid: 9145113
references.citation: J Virol. 1998 Jul;72(7):6186-9
references.pmid: 9621087
references.citation: J Mol Biol. 1996 Jun 7;259(2):249-63
references.pmid: 8656427
references.citation: Electrophoresis. 2001 Jun;22(10):2058-65
references.pmid: 11465506
references.citation: J Struct Biol. 2008 Mar;161(3):232-42
references.pmid: 17869539
references.citation: J Virol. 2008 May;82(9):4638-46
references.pmid: 18305047
references.citation: Science. 2001 Apr 27;292(5517):744-8
references.pmid: 11326105
references.citation: Curr Protoc Protein Sci. 2001 May;Chapter 8:Unit8.3
references.pmid: 18429205
references.citation: J Virol. 2008 Jun;82(11):5190-7
references.pmid: 18367526
references.citation: J Struct Biol. 1996 Jan-Feb;116(1):120-30
references.pmid: 8742733
references.citation: J Mol Biol. 2001 Mar 16;307(1):173-82
references.pmid: 11243812
references.citation: Structure. 2008 May;16(5):673-83
references.pmid: 18462672
references.citation: J Gen Virol. 1995 Sep;76 ( Pt 9):2407-12
references.pmid: 7561785
references.citation: J Struct Biol. 1993 Nov-Dec;111(3):222-33
references.pmid: 8003383
references.citation: J Virol. 2004 Jan;78(2):751-7
references.pmid: 14694107
references.citation: Methods Mol Med. 2005;119:445-62
references.pmid: 16350417