TY - JOUR
T1 - Mutational analysis of the sendai virus V protein
T2 - Importance of the conserved residues for Zn binding, virus pathogenesis, and efficient RNA editing
AU - Fukuhara, Noriko
AU - Huang, Cheng
AU - Kiyotani, Katsuhiro
AU - Yoshida, Tetsuya
AU - Sakaguchi, Takemasa
N1 - Funding Information:
We thank Atsushi Kato and Yoshiyuki Nagai (National Institute of Infectious Diseases, Japan) for providing the SeV recovery system from cDNA. We also thank the Research Center for Molecular Medicine and the Laboratory for Development of Genetic Diagnosis and Gene Ther- apy, Hiroshima University School of Medicine, fo r allowing us to use their facilities. This study was supported in part by a grant-in-aid from the Japan Society for the Promotion of Science.
PY - 2002
Y1 - 2002
N2 - The V protein of Sendai virus (SeV) is nonessential for virus replication in cell culture but indispensable for viral pathogenicity in mice. At the C terminus of the V protein, there are amino acid residues conserved among the members of the Paramyxovinae subfamily that are clustered in three regions: region I, just downstream of the RNA editing site; and regions II and III, cysteine-rich zinc-finger-like regions. In the present study, we introduced mutations into the conserved amino acids and generated nine mutant viruses. All of the viruses had impaired virus replication in mouse lungs and attenuated virulence in mice. Furthermore, the C-terminal polypeptides fused with glutathione-S-transferase with a mutation in region I, II, or III all had impaired Zn binding in a 65Zn-binding assay in solution. These results demonstrate that the conserved amino acids are important for V protein function, probably via protein conformation dependent on Zn binding. One mutant, SeV V-H318N, had inefficient RNA editing, indicating that the nucleotide that is a part of the codon encoding histidine at position 318 is conserved for the RNA editing machinery. In addition, to determine the function of the C-terminal extension of the V protein, which is not translated in recent virulent field isolates, a translational stop codon was introduced to generate the corresponding short V protein. The mutant virus showed similar virus propagation and pathogenicity, indicating that C-terminal extension of the V protein is not relevant to virus pathogenesis.
AB - The V protein of Sendai virus (SeV) is nonessential for virus replication in cell culture but indispensable for viral pathogenicity in mice. At the C terminus of the V protein, there are amino acid residues conserved among the members of the Paramyxovinae subfamily that are clustered in three regions: region I, just downstream of the RNA editing site; and regions II and III, cysteine-rich zinc-finger-like regions. In the present study, we introduced mutations into the conserved amino acids and generated nine mutant viruses. All of the viruses had impaired virus replication in mouse lungs and attenuated virulence in mice. Furthermore, the C-terminal polypeptides fused with glutathione-S-transferase with a mutation in region I, II, or III all had impaired Zn binding in a 65Zn-binding assay in solution. These results demonstrate that the conserved amino acids are important for V protein function, probably via protein conformation dependent on Zn binding. One mutant, SeV V-H318N, had inefficient RNA editing, indicating that the nucleotide that is a part of the codon encoding histidine at position 318 is conserved for the RNA editing machinery. In addition, to determine the function of the C-terminal extension of the V protein, which is not translated in recent virulent field isolates, a translational stop codon was introduced to generate the corresponding short V protein. The mutant virus showed similar virus propagation and pathogenicity, indicating that C-terminal extension of the V protein is not relevant to virus pathogenesis.
KW - Conserved amino acid
KW - Paramyxovirus
KW - Pathogenicity
KW - Reverse genetics
KW - Sendai virus
KW - V protein
KW - Zn binding
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U2 - 10.1006/viro.2002.1516
DO - 10.1006/viro.2002.1516
M3 - Article
C2 - 12202220
AN - SCOPUS:0036432812
SN - 0042-6822
VL - 299
SP - 172
EP - 181
JO - Virology
JF - Virology
IS - 2
ER -