Influenza virus naked RNA can be expressed upon transfection into cells co-expressing the three subunits of the polymerase and the nucleoprotein from simian virus 40 recombinant viruses.
The functionality of the influenza virus polymerase subunits and the nucleoprotein expressed from simian virus 40 (SV40) recombinants has been tested by their ability to direct the in vivo expression of influenza virus-like RNAs. These RNAs, which contained either the chloramphenicol acetyltransferase (CAT) or haemagglutinin (HA) genes, were synthesized and reconstituted in vitro into viral ribonucleoproteins with a polymerase/nucleoprotein mixture purified from influenza virus-infected cells. Only the coinfection with SV40 recombinant viruses expressing the three polymerase subunits and the nucleoprotein allowed the expression of the transfecting CAT or HA RNAs, confirming that this set of viral genes is the minimal requirement for viral gene expression. Unexpectedly, transfection of the corresponding naked RNAs into SV40 recombinant-infected cells was as effective in directing the synthesis of CAT or HA proteins as the standard reconstituted ribonucleoprotein transfection. These results may be important for the genetic analysis of trans-acting factors involved in influenza virus transcription and replication and may open the way to rescuing influenza viruses in the absence of a helper virus.
Centro Nacional de Biotecnología CSIC, Universidad Autónoma, Cantoblanco, Madrid, Spain.
S
S
de la Luna
J
J
Martín
A
A
Portela
J
J
Ortín
D000818:Animals; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D002522:Chlorocebus aethiops; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000502:physiology*; D015870:Gene Expression; D005814:Genes, Viral / Q000502:physiology; D006389:Hemagglutinins, Viral / Q000235:genetics; D009698:Nucleoproteins / Q000235:genetics / Q000502:physiology*; D009975:Orthomyxoviridae / Q000235:genetics*; D012367:RNA, Viral / Q000096:biosynthesis*; D011993:Recombinant Fusion Proteins; D013539:Simian virus 40; D014158:Transcription, Genetic; D014162:Transfection / Q000235:genetics*; D014779:Virus Replication
D006389:Hemagglutinins, Viral; D009698:Nucleoproteins; D012367:RNA, Viral; D011993:Recombinant Fusion Proteins; D015500:Chloramphenicol O-Acetyltransferase; D012321:DNA-Directed RNA Polymerases