title: A charged second-site mutation in the fusion peptide rescues replication of a mutant avian sarcoma and leukosis virus lacking critical cysteine residues flanking the internal fusion domain.
abstract: The entry process of the avian sarcoma and leukosis virus (ASLV) family of retroviruses requires first a specific interaction between the viral surface (SU) glycoproteins and a receptor on the cell surface at a neutral pH, triggering conformational changes in the viral SU and transmembrane (TM) glycoproteins, followed by exposure to low pH to complete fusion. The ASLV TM glycoprotein has been proposed to adopt a structure similar to that of the Ebola virus GP2 protein: each contains an internal fusion peptide flanked by cysteine residues predicted to be in a disulfide bond. In a previous study, we concluded that the cysteines flanking the internal fusion peptide in ASLV TM are critical for efficient function of the ASLV viral glycoproteins in mediating entry. In this study, replication-competent ASLV mutant subgroup A [ASLV(A)] variants with these cysteine residues mutated were constructed and genetically selected for improved replication capacity in chicken fibroblasts. Viruses with single cysteine-to-serine mutations reverted to the wild-type sequence. However, viruses with both C9S and C45S (C9,45S) mutations retained both mutations and acquired a second-site mutation that significantly improved the infectivity of the genetically selected virus population. A charged-amino-acid second-site substitution in the TM internal fusion peptide at position 30 is preferred to rescue the C9,45S mutant ASLV(A). ASLV(A) envelope glycoproteins that contain the C9,45S and G30R mutations bind the Tva receptor at wild-type levels and have improved abilities to trigger conformational changes and to form stable TM oligomers compared to those of the C9,45S mutant glycoprotein.
authors.affiliation: Department of Molecular Medicine, Mayo Clinic, 200 First Street, SW, Rochester, MN 55905, USA.
authors.forename: Deborah C
authors.initials: DC
authors.lastname: Melder
authors.forename: Xueqian
authors.initials: X
authors.lastname: Yin
authors.forename: Sue E
authors.initials: SE
authors.lastname: Delos
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Federspiel
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D030161:Avian Proteins / Q000378:metabolism; D001358:Avian Sarcoma Viruses / Q000235:genetics / Q000502:physiology*; D002478:Cells, Cultured; D002645:Chickens; D003545:Cysteine / Q000235:genetics; D005347:Fibroblasts / Q000821:virology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense; D011485:Protein Binding; D011991:Receptors, Virus / Q000378:metabolism; D013489:Suppression, Genetic; D014760:Viral Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D053586:Virus Internalization
chemical_list: D030161:Avian Proteins; D011991:Receptors, Virus; C088745:Tva receptor; D014760:Viral Fusion Proteins; D003545:Cysteine