Binding of herpes simplex virus (HSV) glycoprotein D (gD) to a cell surface receptor is required to trigger membrane fusion during entry into host cells. Nectin-1 is a cell adhesion molecule and the main HSV receptor in neurons and epithelial cells. We report the structure of gD bound to nectin-1 determined by x-ray crystallography to 4.0 ？ resolution. The structure reveals that the nectin-1 binding site on gD differs from the binding site of the HVEM receptor. A surface on the first Ig-domain of nectin-1, which mediates homophilic interactions of Ig-like cell adhesion molecules, buries an area composed by residues from both the gD N- and C-terminal extensions. Phenylalanine 129, at the tip of the loop connecting β-strands F and G of nectin-1, protrudes into a groove on gD, which is otherwise occupied by C-terminal residues in the unliganded gD and by N-terminal residues in the gD/HVEM complex. Notably, mutation of Phe129 to alanine prevents nectin-1 binding to gD and HSV entry. Together these data are consistent with previous studies showing that gD disrupts the normal nectin-1 homophilic interactions. Furthermore, the structure of the complex supports a model in which gD-receptor binding triggers HSV entry through receptor-mediated displacement of the gD C-terminal region.
References
[1]
Heldwein EE, Krummenacher C (2008) Entry of herpesviruses into mammalian cells. Cell Mol Life Sci 65: 1653–1668.
[2]
Connolly SA, Jackson JO, Jardetzky TS, Longnecker R (2011) Fusing structure and function: a structural view of the herpesvirus entry machinery. Nat Rev Microbiol 9: 369–381.
[3]
Eisenberg RJ, Heldwein EE, Cohen GH, Krummenacher C (2011) Recent progress in understanding herpes simplex virus entry: relationship of structure to function. In: Weller SK, editor. Alphaherpesviruses: Molecular and cellular Biology. Norwich, UK: Horizon Scientific Press. pp. 131–152.
[4]
Atanasiu D, Saw WT, Cohen GH, Eisenberg RJ (2010) Cascade of events governing cell-cell fusion induced by herpes simplex virus glycoproteins gD, gH/gL, and gB. J Virol 84: 12292–12299.
[5]
Chowdary TK, Cairns TM, Atanasiu D, Cohen GH, Eisenberg RJ, et al. (2010) Crystal structure of the conserved herpesvirus fusion regulator complex gH-gL. Nat Struct Mol Biol 17: 882–888.
[6]
Milne RS, Nicola AV, Whitbeck JC, Eisenberg RJ, Cohen GH (2005) Glycoprotein D receptor-dependent, low-pH-independent endocytic entry of herpes simplex virus type 1. J Virol 79: 6655–6663.
[7]
Nicola AV, Hou J, Major EO, Straus SE (2005) Herpes simplex virus type 1 enters human epidermal keratinocytes, but not neurons, via a pH-dependent endocytic pathway. J Virol 79: 7609–7616.
[8]
Nicola AV, Straus SE (2004) Cellular and viral requirements for rapid endocytic entry of herpes simplex virus. J Virol 78: 7508–7517.
[9]
Montgomery RI, Warner MS, Lum BJ, Spear PG (1996) Herpes simplex virus-1 entry into cells mediated by a novel member of the TNF/NGF receptor family. Cell 87: 427–436.
[10]
Tiwari V, Clement C, Scanlan PM, Kowlessur D, Yue BY, et al. (2005) A role for herpesvirus entry mediator as the receptor for herpes simplex virus 1 entry into primary human trabecular meshwork cells. J Virol 79: 13173–13179.
[11]
Akhtar J, Tiwari V, Oh MJ, Kovacs M, Jani A, et al. (2008) HVEM and Nectin-1 are the Major Mediators of Herpes Simplex Virus 1 (HSV-1) Entry into Human Conjunctival Epithelium. Invest Ophthalmol Vis Sci 49: 4026–4035.
[12]
Krummenacher C, Baribaud F, Ponce De Leon M, Baribaud I, Whitbeck JC, et al. (2004) Comparative usage of herpesvirus entry mediator A and nectin-1 by laboratory strains and clinical isolates of herpes simplex virus. Virology 322: 286–299.
[13]
Simpson SA, Manchak MD, Hager EJ, Krummenacher C, Whitbeck JC, et al. (2005) Nectin-1/HveC Mediates herpes simplex virus type 1 entry into primary human sensory neurons and fibroblasts. J Neurovirol 11: 208–218.
[14]
Galen B, Cheshenko N, Tuyama A, Ramratnam B, Herold BC (2006) Access to nectin favors herpes simplex virus infection at the apical surface of polarized human epithelial cells. J Virol 80: 12209–12218.
[15]
Huber MT, Wisner TW, Hegde NR, Goldsmith KA, Rauch DA, et al. (2001) Herpes Simplex Virus with Highly Reduced gD Levels Can Efficiently Enter and Spread between Human Keratinocytes. J Virol 75: 10309–10318.
[16]
Tiwari V, Clement C, Xu D, Valyi-Nagy T, Yue BY, et al. (2006) Role for 3-o-sulfated heparan sulfate as the receptor for herpes simplex virus type 1 entry into primary human corneal fibroblasts. J Virol Methods 80: 8970–8980.
[17]
Delboy MG, Patterson JL, Hollander AM, Nicola AV (2006) Nectin-2-mediated entry of a syncytial strain of herpes simplex virus via pH-independent fusion with the plasma membrane of Chinese hamster ovary cells. Virol J 3: 105.
[18]
Stiles KM, Milne RS, Cohen GH, Eisenberg RJ, Krummenacher C (2008) The herpes simplex virus receptor nectin-1 is down-regulated after trans-interaction with glycoprotein D. Virology 373: 98–111.
[19]
Stiles KM, Krummenacher C (2010) Glycoprotein D actively induces rapid internalization of two nectin-1 isoforms during herpes simplex virus entry. Virology 399: 109–119.
[20]
Stiles KM, Whitbeck JC, Lou H, Cohen GH, Eisenberg RJ, et al. (2010) Herpes simplex virus glycoprotein D interferes with binding of herpesvirus entry mediator to its ligands through downregulation and direct competition. J Virol 84: 11646–11660.
[21]
Carfi A, Willis SH, Whitbeck JC, Krummenacher C, Cohen GH, et al. (2001) Herpes simplex virus glycoprotein D bound to the human receptor HveA. Mol Cell 8: 169–179.
[22]
Krummenacher C, Supekar VM, Whitbeck JC, Lazear E, Connolly SA, et al. (2005) Structure of unliganded HSV gD reveals a mechanism for receptor-mediated activation of virus entry. EMBO J 24: 4144–4153.
[23]
Connolly SA, Landsburg DJ, Carfi A, Wiley DC, Cohen GH, et al. (2003) Structure-based mutagenesis of herpes simplex virus glycoprotein D defines three critical regions at the gD/HveA interface. J Virol 77: 8127–8140.
[24]
Yoon M, Zago A, Shukla D, Spear PG (2003) Mutations in the N termini of herpes simplex virus type 1 and 2 gDs alter functional interactions with the entry/fusion receptors HVEM, Nectin-2, and 3-O-sulfated heparan sulfate but not with Nectin-1. J Virol 77: 9221–9231.
[25]
Krummenacher C, Nicola AV, Whitbeck JC, Lou H, Hou W, et al. (1998) Herpes simplex virus glycoprotein D can bind to poliovirus receptor-related protein 1 or herpesvirus entry mediator, two structurally unrelated mediators of virus entry. J Virol 72: 7064–7074.
[26]
Narita H, Yamamoto Y, Suzuki M, Miyazaki N, Yoshida A, et al. (2011) Crystal structure of the cis-dimer of nectin-1: implications for the architecture of cell-cell junctions. J Biol Chem 286: 12659–12669.
[27]
Cocchi F, Lopez M, Menotti L, Aoubala M, Dubreuil P, et al. (1998) The V domain of herpesvirus Ig-like receptor (HIgR) contains a major functional region in herpes simplex virus-1 entry into cells and interacts physically with the viral glycoprotein D. Proc Natl Acad Sci U S A 95: 15700–15705.
[28]
Krummenacher C, Rux AH, Whitbeck JC, Ponce de Leon M, Lou H, et al. (1999) The first immunoglobulin-like domain of HveC is sufficient to bind herpes simplex virus gD with full affinity while the third domain is involved in oligomerization of HveC. J Virol 73: 8127–8137.
[29]
Cocchi F, Lopez M, Dubreuil P, Campadelli Fiume G, Menotti L (2001) Chimeric nectin1-poliovirus receptor molecules identify a nectin1 region functional in herpes simplex virus entry. J Virol 75: 7987–7994.
[30]
Krummenacher C, Baribaud I, Ponce de Leon M, Whitbeck JC, Lou H, et al. (2000) Localization of a binding site for herpes simplex virus glycoprotein D on the herpesvirus entry mediator C by using anti-receptor monoclonal antibodies. J Virol 74: 10863–10872.
[31]
Takai Y, Miyoshi J, Ikeda W, Ogita H (2008) Nectins and nectin-like molecules: roles in contact inhibition of cell movement and proliferation. Nat Rev Mol Cell Biol 9: 603–615.
[32]
Krummenacher C, Baribaud I, Sanzo JF, Cohen GH, Eisenberg RJ (2002) Effects of herpes simplex virus on structure and function of nectin- 1/HveC. J Virol 76: 2424–2433.
[33]
Sakisaka T, Taniguchi T, Nakanishi H, Takahashi K, Miyahara M, et al. (2001) Requirement of interaction of nectin-1alpha/HveC with afadin for efficient cell-cell spread of herpes simplex virus type 1. J Virol 75: 4734–4743.
Skubak P, Murshudov GN, Pannu NS (2004) Direct incorporation of experimental phase information in model refinement. Acta Crystallogr D 60: 2196–2201.
[36]
Spear PG, Manoj S, Yoon M, Jogger CR, Zago A, et al. (2006) Different receptors binding to distinct interfaces on herpes simplex virus gD can trigger events leading to cell fusion and viral entry. Virology 344: 17–24.
[37]
Geraghty RJ, Fridberg A, Krummenacher C, Cohen GH, Eisenberg RJ, et al. (2001) Use of chimeric nectin-1(HveC)-related receptors to demonstrate that ability to bind alphaherpesvirus gD is not necessarily sufficient for viral entry. Virology 285: 366–375.
[38]
Zhang P, Mueller S, Morais MC, Bator CM, Bowman VD, et al. (2008) Crystal structure of CD155 and electron microscopic studies of its complexes with polioviruses. Proc Natl Acad Sci U S A 105: 18284–18289.
[39]
Kwong PD, Wyatt R, Robinson J, Sweet RW, Sodroski J, et al. (1998) Structure of an HIV gp120 envelope glycoprotein in complex with the CD4 receptor and a neutralizing human antibody. Nature 393: 648–659.
[40]
Bewley MC, Springer K, Zhang YB, Freimuth P, Flanagan JM (1999) Structural analysis of the mechanism of adenovirus binding to its human cellular receptor, CAR. Science 286: 1579–1583.
[41]
Kirchner E, Guglielmi KM, Strauss HM, Dermody TS, Stehle T (2008) Structure of reovirus sigma1 in complex with its receptor junctional adhesion molecule-A. PLoS Pathog 4: e1000235.
[42]
Menotti L, Cerretani A, Hengel H, Campadelli-Fiume G (2008) Construction of a fully retargeted herpes simplex virus 1 recombinant capable of entering cells solely via human epidermal growth factor receptor 2. J Virol 82: 10153–10161.
[43]
Martinez WM, Spear PG (2002) Amino acid substitutions in the V domain of nectin-1 (HveC) that impair entry activity for herpes simplex virus types 1 and 2 but not for Pseudorabies virus or bovine herpesvirus 1. J Virol 76: 7255–7262.
[44]
Uchida H, Shah WA, Ozuer A, Frampton AR Jr, Goins WF, et al. (2009) Generation of herpesvirus entry mediator (HVEM)-restricted herpes simplex virus type 1 mutant viruses: resistance of HVEM-expressing cells and identification of mutations that rescue nectin-1 recognition. J Virol 83: 2951–2961.
[45]
Whitbeck JC, Muggeridge MI, Rux A, Hou W, Krummenacher C, et al. (1999) The major neutralizing antigenic site on herpes simplex virus glycoprotein D overlaps a receptor-binding domain. J Virol 73: 9879–9890.
[46]
Connolly SA, Landsburg DJ, Carfi A, Whitbeck JC, Zuo Y, et al. (2005) Potential nectin-1 binding site on herpes simplex virus glycoprotein D. J Virol 79: 1282–1295.
[47]
Struyf F, Martinez WM, Spear PG (2002) Mutations in the N-terminal domains of nectin-1 and nectin-2 reveal differences in requirements for entry of various alphaherpesviruses and for nectin-nectin interactions. J Virol 76: 12940–12950.
[48]
Dong X, Xu F, Gong Y, Gao J, Lin P, et al. (2006) Crystal structure of the V domain of human Nectin-like molecule-1/Syncam3/Tsll1/Igsf4b, a neural tissue-specific immunoglobulin-like cell-cell adhesion molecule. J Biol Chem 281: 10610–10617.
[49]
van Raaij MJ, Chouin E, van der Zandt H, Bergelson JM, Cusack S (2000) Dimeric structure of the coxsackievirus and adenovirus receptor D1 domain at 1.7 ？ resolution. Structure 8: 1147–1155.
[50]
Hashiguchi T, Ose T, Kubota M, Maita N, Kamishikiryo J, et al. (2011) Structure of the measles virus hemagglutinin bound to its cellular receptor SLAM. Nat Struct Mol Biol 18: 135–141.
[51]
Connolly SA, Landsburg DJ, Carfi A, Wiley DC, Eisenberg RJ, et al. (2002) Structure-based analysis of the herpes simplex virus glycoprotein D binding site present on herpesvirus entry mediator HveA(HVEM). J Virol 76: 10894–10904.
[52]
Lazear E, Carfi A, Whitbeck JC, Cairns TM, Krummenacher C, et al. (2008) Engineered disulfide bonds in herpes simplex virus type 1 gD separate receptor binding from fusion initiation and viral entry. J Virol 82: 700–709.
[53]
Manoj S, Jogger CR, Myscofski D, Yoon M, Spear PG (2004) Mutations in herpes simplex virus glycoprotein D that prevent cell entry via nectins and alter cell tropism. Proc Natl Acad Sci U S A 101: 12414–12421.
[54]
Yoon M, Spear PG (2002) Disruption of adherens junctions liberates nectin-1 to serve as receptor for herpes simplex virus and pseudorabies virus entry. J Virol 76: 7203–7208.
[55]
Bergelson JM, Cunningham JA, Droguett G, Kurt-Jones EA, Krithivas A, et al. (1997) Isolation of a common receptor for Coxsackie B viruses and adenoviruses 2 and 5. Science 275: 1320–1323.
[56]
Mendelsohn CL, Wimmer E, Racaniello VR (1989) Cellular receptor for poliovirus: molecular cloning, nucleotide sequence, and expression of a new member of the immunoglobulin superfamily. Cell 56: 855–865.
[57]
Barton ES, Forrest JC, Connolly JL, Chappell JD, Liu Y, et al. (2001) Junction adhesion molecule is a receptor for reovirus. Cell 104: 441–451.
[58]
Walters RW, Freimuth P, Moninger TO, Ganske I, Zabner J, et al. (2002) Adenovirus fiber disrupts CAR-mediated intercellular adhesion allowing virus escape. Cell 110: 789–799.
[59]
Menotti L, Cocchi F, Campadelli-Fiume G (2002) Critical residues in the CC′ ridge of the human nectin1 receptor V domain enable herpes simplex virus entry into the cell and act synergistically with the downstream region. Virology 301: 6–12.
[60]
Geraghty RJ, Krummenacher C, Eisenberg RJ, Cohen GH, Spear PG (1998) Entry of alphaherpesviruses mediated by poliovirus receptor related protein 1 and poliovirus receptor. Science 280: 1618–1620.
[61]
Willis SH, Rux AH, Peng C, Whitbeck JC, Nicola AV, et al. (1998) Examination of the kinetics of herpes simplex virus glycoprotein D binding to the herpesvirus entry mediator, using surface plasmon resonance. J Virol 72: 5937–5947.
[62]
Gianni T, Amasio M, Campadelli-Fiume G (2009) Herpes simplex virus gD forms distinct complexes with fusion executors gB and gH/gL in part through the C-terminal profusion domain. J Biol Chem 284: 17370–17382.
[63]
Perez-Romero P, Perez A, Capul A, Montgomery R, Fuller AO (2005) Herpes simplex virus entry mediator associates in infected cells in a complex with viral proteins gD and at least gH. J Virol 79: 4540–4544.
[64]
Cocchi F, Fusco D, Menotti L, Gianni T, Eisenberg RJ, et al. (2004) The soluble ectodomain of herpes simplex virus gD contains a membrane-proximal pro-fusion domain and suffices to mediate virus entry. Proc Natl Acad Sci U S A 101: 7445–7450.
[65]
Zhou G, Ye GJ, Debinski W, Roizman B (2002) Engineered herpes simplex virus 1 is dependent on IL13Ralpha 2 receptor for cell entry and independent of glycoprotein D receptor interaction. Proc Natl Acad Sci U S A 99: 15124–15129.
[66]
Zago A, Jogger CR, Spear PG (2004) Use of herpes simplex virus and pseudorabies virus chimeric glycoprotein D molecules to identify regions critical for membrane fusion. Proc Natl Acad Sci U S A 101: 17498–17503.
[67]
Zhou G, Avitabile E, Campadelli-Fiume G, Roizman B (2003) The domains of glycoprotein D required to block apoptosis induced by herpes simplex virus 1 are largely distinct from those involved in cell-cell fusion and binding to nectin1. J Virol 77: 3759–3767.
[68]
Kamiyama H, Zhou G, Roizman B (2006) Herpes simplex virus 1 recombinant virions exhibiting the amino terminal fragment of urokinase-type plasminogen activator can enter cells via the cognate receptor. Gene Ther 13: 621–629.
[69]
Menotti L, Cerretani A, Campadelli-Fiume G (2006) A herpes simplex virus recombinant that exhibits a single-chain antibody to HER2/neu enters cells through the mammary tumor receptor, independently of the gD receptors. J Virol 80: 5531–5539.
[70]
Menotti L, Nicoletti G, Gatta V, Croci S, Landuzzi L, et al. (2009) Inhibition of human tumor growth in mice by an oncolytic herpes simplex virus designed to target solely HER-2-positive cells. Proc Natl Acad Sci U S A 106: 9039–9044.
[71]
Zhou G, Roizman B (2007) Separation of receptor-binding and profusogenic domains of glycoprotein D of herpes simplex virus 1 into distinct interacting proteins. Proc Natl Acad Sci U S A 104: 4142–4146.
[72]
Otwinowski Z, Minor W (1997) Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol 276: 307–326.
[73]
CCP4 (1994) The CCP4 suite: programs for protein crystallography. Acta Crystallogr D 50: 760–763.
[74]
Yeates TO (1997) Detecting and overcoming crystal twinning. Methods Enzymol 276: 344–358.
[75]
Adams PD, Afonine PV, Bunkoczi G, Chen VB, Davis IW, et al. (2010) PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr D 66: 213–221.
[76]
Krummenacher C, Baribaud I, Eisenberg RJ, Cohen GH (2003) Cellular localization of nectin-1 and glycoprotein D during herpes simplex virus infection. J Virol 77: 8985–8999.
