Alteration of Sendai virus morphogenesis and nucleocapsid incorporation due to mutation of cysteine residues of the matrix protein.
The matrix (M) protein of Sendai virus (SeV) has five cysteine residues, at positions 83, 106, 158, 251, and 295. To determine the roles of the cysteine residues in viral assembly, we generated mutant M cDNA possessing a substitution to serine at one of the cysteine residues or at all of the cysteine residues. Some mutant M proteins were unstable when expressed in cultured cells, suggesting that cysteine residues affect protein stability, probably by disrupting the proper conformation. In an attempt to generate virus from cDNA, SeV M-C(83)S, SeV M-C(106)S, and SeV M-C(295)S were successfully recovered from cDNA, while recombinant SeVs possessing other mutations were not. SeV M-C(83)S and SeV M-C(106)S had smaller virus particles than did the wild-type SeV, whereas SeV M-C(295)S had larger and heterogeneously sized particles. Furthermore, SeV M-C(106)S had a significant amount of empty particles lacking nucleocapsids. These results indicate that a single-point mutation at a cysteine residue of the M protein affects virus morphology and nucleocapsid incorporation, showing direct involvement of the M protein in SeV assembly. Cysteine-dependent conformation of the M protein was not due to disulfide bond formation, since the cysteines were shown to be free throughout the viral life cycle.
Sakaguchi
Takemasa
T
Department of Bacteriology, Hiroshima University School of Medicine, Hiroshima 734-8551, Japan. tsaka@hiroshima-u.ac.jp
Uchiyama
Tsuneo
T
Huang
Cheng
C
Fukuhara
Noriko
N
Kiyotani
Katsuhiro
K
Nagai
Yoshiyuki
Y
Yoshida
Tetsuya
T
D000818:Animals; D002460:Cell Line; D002499:Centrifugation, Density Gradient; D003545:Cysteine / Q000235:genetics* / Q000378:metabolism; D008854:Microscopy, Electron; D009024:Morphogenesis; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; D011487:Protein Conformation; D010253:Respirovirus Infections / Q000821:virology; D029082:Sendai virus / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism*; D014758:Viral Core Proteins / Q000378:metabolism*; D014763:Viral Matrix Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014771:Virion / Q000378:metabolism / Q000648:ultrastructure; D019065:Virus Assembly
C097020:M protein, Sendai virus; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; D014758:Viral Core Proteins; D014763:Viral Matrix Proteins; D003545:Cysteine