title: The paramyxovirus fusion protein C-terminal region: mutagenesis indicates an indivisible protein unit.
abstract: Paramyxoviruses enter host cells by fusing the viral envelope with a host cell membrane. Fusion is mediated by the viral fusion (F) protein, and it undergoes large irreversible conformational changes to cause membrane merger. The C terminus of PIV5 F contains a membrane-proximal 7-residue external region (MPER), followed by the transmembrane (TM) domain and a 20-residue cytoplasmic tail. To study the sequence requirements of the F protein C terminus for fusion, we constructed chimeras containing the ectodomain of parainfluenza virus 5 F (PIV5 F) and either the MPER, the TM domain, or the cytoplasmic tail of the F proteins of the paramyxoviruses measles virus, mumps virus, Newcastle disease virus, human parainfluenza virus 3, and Nipah virus. The chimeras were expressed, and their ability to cause cell fusion was analyzed. The chimeric proteins were variably expressed at the cell surface. We found that chimeras containing the ectodomain of PIV5 F with the C terminus of other paramyxoviruses were unable to cause cell fusion. Fusion could be restored by decreasing the activation energy of refolding through introduction of a destabilizing mutation (S443P). Replacing individual regions, singly or doubly, in the chimeras with native PIV5 F sequences restored fusion to various degrees, but it did not have an additive effect in restoring activity. Thus, the F protein C terminus may be a specific structure that only functions with its cognate ectodomain. Alanine scanning mutagenesis of MPER indicates that it has a regulatory role in fusion since both hyperfusogenic and hypofusogenic mutations were found.
authors.affiliation: Department of Molecular Biosciences, Northwestern University, Evanston, Illinois, USA.
authors.forename: Aarohi
authors.initials: A
authors.lastname: Zokarkar
authors.forename: Robert A
authors.initials: RA
authors.lastname: Lamb
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D002459:Cell Fusion; D002460:Cell Line; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D018184:Paramyxoviridae Infections / Q000662:veterinary / Q000821:virology; D018098:Paramyxovirinae / Q000737:chemistry* / Q000235:genetics* / Q000502:physiology; D019213:Rubulavirus / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D016415:Sequence Alignment; D014760:Viral Fusion Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism / Q000502:physiology
chemical_list: D014760:Viral Fusion Proteins