title: Substitution of specific cysteine residues in the E1 glycoprotein of classical swine fever virus strain Brescia affects formation of E1-E2 heterodimers and alters virulence in swine.
abstract: E1, along with E(rns) and E2, is one of the three envelope glycoproteins of classical swine fever virus (CSFV). E1 and E2 are anchored to the virus envelope at their carboxyl termini, and E(rns) loosely associates with the viral envelope. In infected cells, E2 forms homodimers and heterodimers with E1 mediated by disulfide bridges between cysteine residues. The E1 protein of CSFV strain Brescia contains six cysteine residues at positions 5, 20, 24, 94, 123, and 171. The role of these residues in the formation of E1-E2 heterodimers and their effect on CSFV viability in vitro and in vivo remain unclear. Here we observed that recombinant viruses harboring individual cysteine-to-serine substitutions within the E1 envelope protein still have formation of E1-E2 heterodimers which are functional in terms of allowing efficient virus progeny yields in infected primary swine cells. Additionally, these single cysteine mutant viruses were virulent in infected swine. However, a double mutant harboring Cys24Ser and Cys94Ser substitutions within the E1 protein altered formation of E1-E2 heterodimers in infected cells. This recombinant virus, E1ΔCys24/94v, showed delayed growth kinetics in primary swine macrophage cultures and was attenuated in swine. Furthermore, despite the observed diminished growth in vitro, infection with E1ΔCys24/94v protected swine from challenge with virulent CSFV strain Brescia at 3 and 28 days postinfection.
authors.affiliation: Plum Island Animal Disease Center, USDA/ARS/NAA, P.O. Box 848, Greenport, NY 11944-0848, USA.
authors.forename: I
authors.initials: I
authors.lastname: Fernández-Sainz
authors.forename: L G
authors.initials: LG
authors.lastname: Holinka
authors.forename: D
authors.initials: D
authors.lastname: Gladue
authors.forename: V
authors.initials: V
authors.lastname: O'Donnell
authors.forename: Z
authors.initials: Z
authors.lastname: Lu
authors.forename: B K
authors.initials: BK
authors.lastname: Gavrilov
authors.forename: G R
authors.initials: GR
authors.lastname: Risatti
authors.forename: M V
authors.initials: MV
authors.lastname: Borca
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003545:Cysteine / Q000235:genetics*; D017931:DNA Primers; D053118:Influenza A Virus, H1N1 Subtype / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017386:Sequence Homology, Amino Acid; D013552:Swine; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*
chemical_list: D017931:DNA Primers; D014764:Viral Proteins; D003545:Cysteine