title: Effect of cysteine mutations in the extracellular domain of CM2 on the influenza C virus replication.
abstract: CM2 is the second membrane protein of influenza C virus and possesses three conserved cysteines at residue 1, 6 and 20 in its extracellular domain, all of which are involved in the formation of disulfide-linked oligomers of the molecule. In the present study, to examine the effect of CM2 oligomerization on virus replication, we generated a mutant recombinant virus, rC1620A, in which all three cysteines on CM2 were substituted to alanines. The rC1620A virus was more attenuated than the recombinant wild-type (rWT) virus in cultured cells. The CM2 protein synthesized in rC1620A-infected cells could not apparently be detected as a tetramer and was transported to the cell surface less efficiently than was authentic CM2. The amount of CM2 protein incorporated into the rC1620A virions was comparable to that into the rWT virions, although the main CM2 species in the rC1620A virions was in the form of a dimer. Analyses of one-step grown virions and virus-infected cells could not provide evidence for any difference in growth between rC1620A and rWT. On the other hand, the amount of genome present in VLPs possessing the mutant CM2 (C1620A-VLPs) was approximately 31% of that in VLPs possessing wild-type CM2 (WT-VLPs). The incoming genome from VLPs was less efficiently transported to the nucleus in the C1620A-VLP-infected cells than in WT-VLP-infected cells, leading to reduced reporter gene expression in the C1620A-VLP-infected cells. Taken together, these findings demonstrate that CM2 oligomerization affects the packaging and uncoating processes. Thus, we concluded that disulfide-linked CM2 oligomers facilitate virus growth by affecting the replication processes.
authors.affiliation: Department of Microbiology, Kanazawa Medical University School of Medicine Uchinada, Ishikawa, Japan. ymuraki@kanazawa-med.ac.jp
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Muraki
authors.forename: Takako
authors.initials: T
authors.lastname: Okuwa
authors.forename: Toshiki
authors.initials: T
authors.lastname: Himeda
authors.forename: Seiji
authors.initials: S
authors.lastname: Hongo
authors.forename: Yoshiro
authors.initials: Y
authors.lastname: Ohara
mesh_terms: D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry*; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D006801:Humans; D009982:Gammainfluenzavirus / Q000502:physiology*; D009154:Mutation; D054730:Protein Interaction Domains and Motifs / Q000235:genetics*; D055503:Protein Multimerization; D014763:Viral Matrix Proteins / Q000737:chemistry / Q000235:genetics*; D014764:Viral Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D019065:Virus Assembly; D014779:Virus Replication / Q000235:genetics*; D057646:Virus Uncoating
chemical_list: C090877:CM2 protein, Influenza C virus; D014763:Viral Matrix Proteins; D014764:Viral Proteins; D003545:Cysteine