Highly stable expression of a foreign gene from rabies virus vectors.
A reverse genetics approach was applied to generate a chimeric nonsegmented negative strand RNA virus, rabies virus (RV) of the Rhabdoviridae family, that expresses a foreign protein. DNA constructs containing the entire open reading frame of the bacterial chloramphenicol acetyltransferase (CAT) gene and an upstream RV cistron border sequence were inserted either into the nontranslated pseudogene region of a full-length cDNA copy of the RV genome or exchanged with the pseudogene region. After intracellular T7 RNA polymerase-driven expression of full-length antigenome RNA transcripts and RV nucleoprotein, phosphoprotein and polymerase from transfected plasmids, RVs transcribing novel monocistronic mRNAs and expressing CAT at high levels, were recovered. The chimeric viruses possessed the growth characteristics of standard RV and were genetically stable upon serial cell culture passages. CAT activity was still observed in cell cultures infected with viruses passaged for more than 25 times. Based on the unprecedented stability of the chimeric RNA genomes, which is most likely due to the structure of the rhabdoviral ribonucleoprotein complex, we predict the successful future use of recombinant rhabdovirus vectors for displaying foreign antigens or delivering therapeutic genes.
Mebatsion
T
T
Department of Clinical Virology, Federal Research Centre for Virus Diseases of Animals, Tübingen, Germany.
Schnell
M J
MJ
Cox
J H
JH
Finke
S
S
Conzelmann
K K
KK
D000818:Animals; D002460:Cell Line; D002678:Chimera; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D018076:DNA, Complementary; D017353:Gene Deletion; D005814:Genes, Viral; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors; D016679:Genome, Viral; D051379:Mice; D008815:Mice, Inbred Strains; D011544:Pseudogenes; D011820:Rabies virus; D011994:Recombinant Proteins / Q000096:biosynthesis*; D013698:Templates, Genetic; D014158:Transcription, Genetic; D014162:Transfection / Q000379:methods*; D015678:Viral Structural Proteins / Q000235:genetics
D018076:DNA, Complementary; D011994:Recombinant Proteins; D015678:Viral Structural Proteins; D015500:Chloramphenicol O-Acetyltransferase