Generation of recombinant influenza A virus without M2 ion-channel protein by introduction of a point mutation at the 5' end of the viral intron.
The aim of this study was to inhibit influenza virus M2 protein expression by mutating the splicing signal of the M gene. Mutations were introduced into the GU dinucleotide sequence at the 5'-proximal splicing site of the M gene (corresponding to nt 52-53 of M cRNA). Transfected cells expressing mutated M viral ribonucleoproteins failed to generate M2 mRNA. Interestingly, recombinant viruses with mutations at the dinucleotide sequence were viable, albeit attenuated, in cell culture. These recombinants failed to express M2 mRNA and M2 protein. These observations demonstrated that the GU invariant dinucleotide sequence at the 5'-proximal splicing site of M gene is essential for M2 mRNA synthesis. These results also indicated that the M2 ion-channel protein is critical, but not essential, for virus replication in cell culture. This approach may provide a new way of producing attenuated influenza A virus.
Cheung
T K W
TKW
Department of Microbiology, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong SAR.
Guan
Y
Y
Department of Microbiology, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong SAR.
Ng
S S F
SSF
Department of Microbiology, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong SAR.
Chen
H
H
Department of Microbiology, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong SAR.
Wong
C H K
CHK
Department of Microbiology, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong SAR.
Peiris
J S M
JSM
Department of Microbiology, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong SAR.
Poon
L L M
LLM
Department of Microbiology, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong SAR.
D002478:Cells, Cultured; D006801:Humans; D009980:Influenza A virus / Q000737:chemistry* / Q000235:genetics* / Q000254:growth & development; D007438:Introns; D017354:Point Mutation; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D012367:RNA, Viral / Q000032:analysis / Q000096:biosynthesis; D014763:Viral Matrix Proteins / Q000096:biosynthesis* / Q000235:genetics*; D000086082:Viroporin Proteins
D012333:RNA, Messenger; D012367:RNA, Viral; D014763:Viral Matrix Proteins; C488937:M2 protein, Influenza A virus; D000086082:Viroporin Proteins