Production of infectious human respiratory syncytial virus from cloned cDNA confirms an essential role for the transcription elongation factor from the 5' proximal open reading frame of the M2 mRNA in gene expression and provides a capability for vaccine development.
Infectious human respiratory syncytial virus (RSV) was produced by the intracellular coexpression of five plasmid-borne cDNAs. One cDNA encoded a complete positive-sense version of the RSV genome (corresponding to the replicative intermediate RNA or antigenome), and each of the other four encoded a separate RSV protein, namely, the major nucleocapsid N protein, the nucleocapsid P phosphoprotein, the major polymerase L protein, or the protein from the 5' proximal open reading frame of the M2 mRNA [M2(ORF1)]. RSV was not produced if any of the five plasmids was omitted. The requirement for the M2(ORF1) protein is consistent with its recent identification as a transcription elongation factor and confirms its importance for RSV gene expression. It should thus be possible to introduce defined changes into infectious RSV. This should be useful for basic studies of RSV molecular biology and pathogenesis; in addition, there are immediate applications to the development of live attenuated vaccine strains bearing predetermined defined attenuating mutations.
Collins
P L
PL
Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892-0720, USA.
Hill
M G
MG
Camargo
E
E
Grosfeld
H
H
Chanock
R M
RM
Murphy
B R
BR
D001483:Base Sequence; D003001:Cloning, Molecular; D016373:DNA, Antisense; D018076:DNA, Complementary / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D015707:HN Protein; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D012333:RNA, Messenger / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D018113:Respiratory Syncytial Virus, Human / Q000235:genetics* / Q000254:growth & development* / Q000276:immunology; D014158:Transcription, Genetic; D014162:Transfection; D014614:Vaccines, Synthetic / Q000235:genetics; D014759:Viral Envelope Proteins; D010948:Viral Plaque Assay; D014764:Viral Proteins / Q000378:metabolism*; D014765:Viral Vaccines / Q000235:genetics
D016373:DNA, Antisense; D018076:DNA, Complementary; D015707:HN Protein; D012333:RNA, Messenger; D012367:RNA, Viral; D014614:Vaccines, Synthetic; D014759:Viral Envelope Proteins; D014764:Viral Proteins; D014765:Viral Vaccines; C116204:attachment protein G