Involvement of the zinc-binding capacity of Sendai virus V protein in viral pathogenesis.
The V protein of Sendai virus (SeV) is nonessential to virus replication in cell culture but indispensable to viral pathogenicity in mice. The highly conserved cysteine-rich zinc finger-like domain in its carboxyl terminus is believed to be responsible for this viral pathogenicity. In the present study, we showed that the cysteine-rich domain of the SeV V protein could actually bind zinc by using glutathione-S-transferase fusion proteins. When the seven conserved cysteine residues at positions 337, 341, 353, 355, 358, 362, and 365 were replaced individually, the zinc-binding capacities of the mutant proteins were greatly impaired, ranging from 22 to 68% of that of the wild type. We then recovered two mutant SeVs from cDNA, which have V-C(341)S and V-C(365)R mutations and represent maximal and minimal zinc-binding capacities among the corresponding mutant fusion proteins, respectively. The mutant viruses showed viral protein synthesis and growth patterns similar to those of wild-type SeV in cultured cells. However, the mutant viruses were strongly attenuated in mice in a way similar to that of SeV V(DeltaC), which has a truncated V protein lacking the cysteine-rich domain, by exhibiting earlier viral clearance from the mouse lung and less virulence to mice. We therefore conclude that the zinc-binding capacity of the V protein is involved in viral pathogenesis.
Huang
C
C
Department of Bacteriology, Hiroshima University School of Medicine, Hiroshima 734-8551, Japan.
Kiyotani
K
K
Fujii
Y
Y
Fukuhara
N
N
Kato
A
A
Nagai
Y
Y
Yoshida
T
T
Sakaguchi
T
T
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D003545:Cysteine / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D018108:Respirovirus / Q000235:genetics / Q000472:pathogenicity*; D014764:Viral Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication / Q000502:physiology; D015032:Zinc / Q000378:metabolism*; D016335:Zinc Fingers
D011993:Recombinant Fusion Proteins; C111739:V protein, Sendai virus; D014764:Viral Proteins; D005982:Glutathione Transferase; D015032:Zinc; D003545:Cysteine