An infectious clone of human parainfluenza virus type 3.
A full-length clone of the human parainfluenza virus type 3 (HPIV-3) genome (called pHPIV-3) was constructed, and recombinant, infectious HPIV-3 was generated by transfecting pHPIV-3 and support plasmids encoding the HPIV-3 NP, P, and L proteins into HeLa cells infected with a vaccinia virus recombinant which expresses T7 RNA polymerase. T7 RNA polymerase promoters on the transfected plasmids direct the synthesis of transcripts encoding the NP, P, and L proteins and a full-length, positive-sense copy of the HPIV-3 genome. Generation of virus was dependent on transfection of pHPIV-3 and the HPIV-3 P- and L-encoding plasmids. However, a plasmid encoding the NP protein was not required since NP was expressed from pHPIV-3. Recovered virus was neutralized by anti-HPIV-3 antisera and shown to contain specific base substitutions characteristic of pHPIV-3. Recombination was shown to occur during recovery, as viruses with two distinct genotypes and phenotypes were isolated. The ability to produce infectious HPIV-3 engineered to contain specific alterations within the HPIV-3 genes and cis-acting elements expedites the study of all aspects of the virus replication cycle. Additionally, analysis of mutations may lead to the identification of attenuating genotypes, a key step in the development of a live virus vaccine.
Hoffman
M A
MA
Department of Molecular Biology, Research Institute, The Cleveland Clinic Foundation, Ohio 44195, USA.
Banerjee
A K
AK
D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D006367:HeLa Cells; D006801:Humans; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; D010224:Parainfluenza Virus 3, Human / Q000235:genetics*; D010957:Plasmids; D012093:Replicon; D014758:Viral Core Proteins / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics; D014779:Virus Replication
D018076:DNA, Complementary; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; D014758:Viral Core Proteins; D014764:Viral Proteins