title: Receptor binding, fusion inhibition, and induction of cross-reactive neutralizing antibodies by a soluble G glycoprotein of Hendra virus.
abstract: Hendra virus (HeV) and Nipah virus (NiV) are closely related emerging viruses comprising the Henipavirus genus of the Paramyxovirinae, which are distinguished by their ability to cause fatal disease in both animal and human hosts. These viruses infect cells by a pH-independent membrane fusion event mediated by their attachment (G) and fusion (F) glycoproteins. Previously, we reported on HeV- and NiV-mediated fusion activities and detailed their host-cell tropism characteristics. These studies also suggested that a common cell surface receptor, which could be destroyed by protease, was utilized by both viruses. To further characterize the G glycoprotein and its unknown receptor, soluble forms of HeV G (sG) were constructed by replacing its cytoplasmic tail and transmembrane domains with an immunoglobulin kappa leader sequence coupled to either an S-peptide tag (sG(S-tag)) or myc-epitope tag (sG(myc-tag)) to facilitate purification and detection. Expression of sG was verified in cell lysates and culture supernatants by specific affinity precipitation. Analysis of sG by size exclusion chromatography and sucrose gradient centrifugation demonstrated tetrameric, dimeric, and monomeric species, with the majority of the sG released as a disulfide-linked dimer. Immunofluorescence staining revealed that sG specifically bound to HeV and NiV infection-permissive cells but not to a nonpermissive HeLa cell line clone, suggesting that it binds to virus receptor on host cells. Preincubation of host cells with sG resulted in dose-dependent inhibition of both HeV and NiV cell fusion as well as infection by live virus. Taken together, these data indicate that sG retains important native structural features, and we further demonstrate that administration of sG to rabbits can elicit a potent cross-reactive neutralizing antibody response against infectious HeV and NiV. This HeV sG glycoprotein will be exceedingly useful for structural studies, receptor identification strategies, and vaccine development goals for these important emerging viral agents.
authors.affiliation: Department of Microbiology and Immunology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Road, Bethesda, MD 20814-4799, USA.
authors.forename: Katharine N
authors.initials: KN
authors.lastname: Bossart
authors.forename: Gary
authors.initials: G
authors.lastname: Crameri
authors.forename: Antony S
authors.initials: AS
authors.lastname: Dimitrov
authors.forename: Bruce A
authors.initials: BA
authors.lastname: Mungall
authors.forename: Yan-Ru
authors.initials: YR
authors.lastname: Feng
authors.forename: Jared R
authors.initials: JR
authors.lastname: Patch
authors.forename: Anil
authors.initials: A
authors.lastname: Choudhary
authors.forename: Lin-Fa
authors.initials: LF
authors.lastname: Wang
authors.forename: Bryan T
authors.initials: BT
authors.lastname: Eaton
authors.forename: Christopher C
authors.initials: CC
authors.lastname: Broder
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral / Q000096:biosynthesis; D001483:Base Sequence; D002460:Cell Line; D002522:Chlorocebus aethiops; D003429:Cross Reactions; D004279:DNA, Viral / Q000235:genetics; D006367:HeLa Cells; D045404:Hendra Virus / Q000235:genetics / Q000276:immunology / Q000472:pathogenicity* / Q000502:physiology*; D006801:Humans; D008561:Membrane Fusion / Q000502:physiology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009500:Neutralization Tests; D045405:Nipah Virus / Q000235:genetics / Q000276:immunology / Q000472:pathogenicity / Q000502:physiology; D011991:Receptors, Virus / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012995:Solubility; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000502:physiology*
chemical_list: D000914:Antibodies, Viral; D004279:DNA, Viral; D011991:Receptors, Virus; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C116204:attachment protein G