1999 Fiscal Year Final Research Report Summary
A structure-functional analysis of the paramyxovirus M protein by using a virus recovery system from cDNA
Project/Area Number |
10670286
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Research Category |
Grant-in-Aid for Scientific Research (C)
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Allocation Type | Single-year Grants |
Section | 一般 |
Research Field |
Virology
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Research Institution | Hiroshima University |
Principal Investigator |
SAKAGUCHI Takemasa Hiroshima University Faculty of Medicine, Associate Professor, 医学部, 助教授 (70196070)
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Project Period (FY) |
1998 – 1999
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Keywords | paramyxovirus / M protein / virus recovery / cysteine / morphogenesis / reverse genetics |
Research Abstract |
The matrix (M) protein of Sendai virus (SeV) has five cysteine residues, at positions 83, 106, 158, 251 and 295. The fast migration of the M protein in SDS-PAGE in a non-reducing condition suggests that it forms a specific structure. The structure is considered to depend on cysteine residues, since mutations of the cysteine residues affect migration in SDS-PAGE. To determine the cysteine-dependent structure of the M protein in viral replication, we tried to recover virus from mutant genomic cDNA possessing a substitution to serine at one of the cysteine residues or at all of the cysteine residues. SeV M-CィイD283ィエD2S, SeV M-CィイD2106ィエD2S, and SeV M-CィイD2295ィエD2S were successfully recovered from cDNA, while recombinant SeVs possessing M-CィイD2158ィエD2S, M-CィイD2251ィエD2S, and M-C(-) mutations were not, suggesting the importance of the cysteine residues at positions 158 and/or 251 in virus replication. SeV M-CィイD283ィエD2S and SeV M-CィイD2106ィエD2S had smaller virus particles than did the wild-type SeV, whereas SeV M-CィイD2295ィエD2S had larger and heterogeneous particles. Furthermore, SeV M-CィイD2106ィエD2S had a significant amount of empty particles lacking the viral genome. These results indicate that a single-point mutation at the cysteine residues of the M protein affects virus morphology and genome incorporation. SeV M-CィイD283ィエD2S and SeV M-CィイD2106ィエD2S exhibited a lower virus growth in cultured cells and mouse lungs, leading to a lower pathogenicity to mice compared with the wild-type virus, while the SeV M-CィイD2295ィエD2S mutant grew as efficiently as the wild-type virus. The infection of cultured cells with SeV M-CィイD283ィエD2S and SeV M-CィイD2106ィエD2S suggested that virus assembly and/or budding steps might be abrogated.
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