title: Roles in cell-to-cell fusion of two conserved hydrophobic regions in the murine coronavirus spike protein.
abstract: The spike (S) protein of coronavirus mouse hepatitis virus (MHV), mediates attachment and fusion during viral entry and cell-to-cell fusion later in infection. By analogy with other viral proteins that induce cell fusion the MHV S protein would be expected to have a hydrophobic stretch of amino acids that serves as a fusion peptide. Sequence analysis suggests that the S protein falls within the group of fusion proteins having internal rather than N-terminal fusion peptides. Based on the features of known viral fusion peptides, we identified two regions (PEP1 and PEP2) of MHV-A59 S2 as possible fusion peptides. Site-directed mutagenesis and an in viro cell-to-cell fusion assay were used to evaluate the roles of PEP1 and PEP2, as well as a third previously identified putative fusion domain (PEP3) in membrane fusion. Substitution of bulky hydrophobic residues with charged residues within PEP1 affects the fusion activity of the S protein without affecting processing and surface expression. Similar substitutions within PEP2 result in a fusion-negative phenotype; however, these mutant S proteins also exhibit defects in protein processing and surface expression which likely explain the loss of the ability to induce fusion. Thus PEP1 remains a candidate fusion peptide, while PEP2 may play a significant role in the overall structure or oligomerization of the S protein. PEP3 is an unlikely putative fusion peptide since it is not conserved among coronaviruses and nonconservative amino acid substitutions in PEP3 have minimal effects on cell-to-cell fusion.
authors.affiliation: Department of Microbiology, University of Pennsylvania School of Medicine, Philadelphia 19104-6076, USA.
authors.forename: Z
authors.initials: Z
authors.lastname: Luo
authors.forename: S R
authors.initials: SR
authors.lastname: Weiss
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D002459:Cell Fusion / Q000502:physiology*; D002460:Cell Line; D017124:Conserved Sequence; D017934:Coronavirus / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology; D006224:Cricetinae; D017931:DNA Primers / Q000235:genetics; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D014760:Viral Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*
chemical_list: D017931:DNA Primers; D008562:Membrane Glycoproteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus