title: Importance of arginine 20 of the swine vesicular disease virus 2A protease for activity and virulence.
abstract: A major virulence determinant of swine vesicular disease virus (SVDV), an Enterovirus that causes an acute vesicular disease, has been mapped to residue 20 of the 2A protease. The SVDV 2A protease cleaves the 1D-2A junction in the viral polyprotein, induces cleavage of translation initiation factor eIF4GI, and stimulates the activity of enterovirus internal ribosome entry sites (IRESs). The 2A protease from an attenuated strain of SVDV (Ile at residue 20) is significantly defective at inducing cleavage of eIF4GI and the activation of IRES-dependent translation compared to the 2A protease from a pathogenic strain (J1/73, Arg at residue 20), but the two proteases have similar 1D-2A cleavage activities (Y. Sakoda, N. Ross-Smith, T. Inoue, and G. J. Belsham, J. Virol. 75:10643-10650, 2001). Residue 20 has now been modified to every possible amino acid, and the activities of each mutant 2A protease has been analyzed. Selected mutants were reconstructed into full-length SVDV cDNA, and viruses were rescued. The rate of virus growth in cultured swine kidney cells reflected the efficiency of 2A protease activity. In experimentally infected pigs, all four of the mutant viruses tested displayed much-reduced virulence compared to the J1/73 virus but a significant, albeit reduced, level of viral replication and excretion was detected. Direct sequencing of cDNA derived from samples taken early and late in infection indicated that a gradual selection-reversion to a more efficient protease occurred. The data indicated that extensive sequence change and selection may introduce a severe bottleneck in virus replication, leading to a decreased viral load and reduced or no clinical disease.
authors.affiliation: Department of Exotic Disease, National Institute of Animal Health, Kodaira, Tokyo, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Inoue
authors.forename: Soren
authors.initials: S
authors.lastname: Alexandersen
authors.forename: Angela T
authors.initials: AT
authors.lastname: Clark
authors.forename: Ciara
authors.initials: C
authors.lastname: Murphy
authors.forename: Melvyn
authors.initials: M
authors.lastname: Quan
authors.forename: Scott M
authors.initials: SM
authors.lastname: Reid
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Sakoda
authors.forename: Helen L
authors.initials: HL
authors.lastname: Johns
authors.forename: Graham J
authors.initials: GJ
authors.lastname: Belsham
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine; D001483:Base Sequence; D003546:Cysteine Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029822:Enterovirus B, Human / Q000235:genetics / Q000472:pathogenicity*; D004769:Enterovirus Infections / Q000503:physiopathology / Q000662:veterinary* / Q000821:virology; D015967:Gene Expression Regulation, Viral; D008969:Molecular Sequence Data; D009154:Mutation; D017422:Sequence Analysis, DNA; D013555:Swine Vesicular Disease / Q000503:physiopathology* / Q000821:virology; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014774:Virulence
chemical_list: D014764:Viral Proteins; D001120:Arginine; D003546:Cysteine Endopeptidases; C057400:picornain 2A, Picornavirus