title: 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.
abstract: Multiple cell surface molecules (herpesvirus entry mediator [HVEM], nectin-1, nectin-2, and 3-O-sulfated heparan sulfate) can serve as entry receptors for herpes simplex virus type 1 (HSV-1) or HSV-2 and also as receptors for virus-induced cell fusion. Viral glycoprotein D (gD) is the ligand for these receptors. A previous study showed that HVEM makes contact with HSV-1 gD at regions within amino acids 7 to 15 and 24 to 32 at the N terminus of gD. In the present study, amino acid substitutions and deletions were introduced into the N termini of HSV-1 and HSV-2 gDs to determine the effects on interactions with all of the known human and mouse entry/fusion receptors, including mouse HVEM, for which data on HSV entry or cell fusion were not previously reported. A cell fusion assay was used to assess functional activity of the gD mutants with each entry/fusion receptor. Soluble gD:Fc hybrids carrying each mutation were tested for the ability to bind to cells expressing the entry/fusion receptors. We found that deletions overlapping either or both of the HVEM contact regions, in either HSV-1 or HSV-2 gD, severely reduced cell fusion and binding activity with all of the human and mouse receptors except nectin-1. Amino acid substitutions described previously for HSV-1 (L25P, Q27P, and Q27R) were individually introduced into HSV-2 gD and, for both serotypes, were found to be without effect on cell fusion and the binding activity for nectin-1. Each of these three substitutions in HSV-1 gD enhanced fusion with cells expressing human nectin-2 (ordinarily low for wild-type HSV-1 gD), but the same substitutions in HSV-2 gD were without effect on the already high level of cell fusion observed with the wild-type protein. The Q27P or Q27R substitution in either HSV-1 and HSV-2 gD, but not the L25P substitution, significantly reduced cell fusion and binding activity for both human and mouse HVEM. Each of the three substitutions in HSV-1 gD, as well as the deletions mentioned above, reduced fusion with cells bearing 3-O-sulfated heparan sulfate. Thus, the N terminus of HSV-1 or HSV-2 gD is not necessary for functional interactions with nectin-1 but is necessary for all of the other receptors tested here. The sequence of the N terminus determines whether nectin-2 or 3-O-sulfated heparan sulfate, as well as HVEM, can serve as entry/fusion receptors.
authors.affiliation: Department of Microbiology-Immunology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois 60611, USA.
authors.forename: Miri
authors.initials: M
authors.lastname: Yoon
authors.forename: Anna
authors.initials: A
authors.lastname: Zago
authors.forename: Deepak
authors.initials: D
authors.lastname: Shukla
authors.forename: Patricia G
authors.initials: PG
authors.lastname: Spear
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D015815:Cell Adhesion Molecules / Q000502:physiology; D006224:Cricetinae; D004274:DNA, Recombinant / Q000235:genetics; D006497:Heparan Sulfate / Q000502:physiology; D018259:Herpesvirus 1, Human / Q000235:genetics* / Q000502:physiology*; D018258:Herpesvirus 2, Human / Q000235:genetics* / Q000502:physiology*; D006801:Humans; D008561:Membrane Fusion / Q000502:physiology; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D000075983:Nectins; D018124:Receptors, Tumor Necrosis Factor / Q000502:physiology; D053240:Receptors, Tumor Necrosis Factor, Member 14; D011991:Receptors, Virus / Q000502:physiology; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000502:physiology*
chemical_list: D015815:Cell Adhesion Molecules; D004274:DNA, Recombinant; D006497:Heparan Sulfate; D000075983:Nectins; D018124:Receptors, Tumor Necrosis Factor; D053240:Receptors, Tumor Necrosis Factor, Member 14; D011991:Receptors, Virus; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; C000619010:NECTIN1 protein, human; C000619015:Nectin1 protein, mouse; C506028:TNFRSF14 protein, human; C506029:Tnfrsf14 protein, mouse; C039447:glycoprotein D, Human herpesvirus 1; C040570:glycoprotein D-herpes simplex virus type 2