title: Recombinant Sendai viruses expressing fusion proteins with two furin cleavage sites mimic the syncytial and receptor-independent infection properties of respiratory syncytial virus.
abstract: Cell entry by paramyxoviruses requires fusion between viral and cellular membranes. Paramyxovirus infection also gives rise to the formation of multinuclear, fused cells (syncytia). Both types of fusion are mediated by the viral fusion (F) protein, which requires proteolytic processing at a basic cleavage site in order to be active for fusion. In common with most paramyxoviruses, fusion mediated by Sendai virus F protein (F(SeV)) requires coexpression of the homologous attachment (hemagglutinin-neuraminidase [HN]) protein, which binds to cell surface sialic acid receptors. In contrast, respiratory syncytial virus fusion protein (F(RSV)) is capable of fusing membranes in the absence of the viral attachment (G) protein. Moreover, F(RSV) is unique among paramyxovirus fusion proteins since F(RSV) possesses two multibasic cleavage sites, which are separated by an intervening region of 27 amino acids. We have previously shown that insertion of both F(RSV) cleavage sites in F(SeV) decreases dependency on the HN attachment protein for syncytium formation in transfected cells. We now describe recombinant Sendai viruses (rSeV) that express mutant F proteins containing one or both F(RSV) cleavage sites. All cleavage-site mutant viruses displayed reduced thermostability, with double-cleavage-site mutants exhibiting a hyperfusogenic phenotype in infected cells. Furthermore, insertion of both F(RSV) cleavage sites in F(SeV) reduced dependency on the interaction of HN with sialic acid for infection, thus mimicking the unique ability of RSV to fuse and infect cells in the absence of a separate attachment protein.
authors.affiliation: Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, 28220 Madrid, Spain. joannarawling@yahoo.co.uk
authors.forename: Joanna
authors.initials: J
authors.lastname: Rawling
authors.forename: Olga
authors.initials: O
authors.lastname: Cano
authors.forename: Dominique
authors.initials: D
authors.lastname: Garcin
authors.forename: Daniel
authors.initials: D
authors.lastname: Kolakofsky
authors.forename: José A
authors.initials: JA
authors.lastname: Melero
mesh_terms: D002459:Cell Fusion; D045683:Furin / Q000378:metabolism*; D055550:Protein Stability; D012136:Respiratory Syncytial Viruses / Q000235:genetics / Q000502:physiology*; D029082:Sendai virus / Q000235:genetics / Q000502:physiology*; D013696:Temperature; D014760:Viral Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D053586:Virus Internalization
chemical_list: D014760:Viral Fusion Proteins; D045683:Furin