Efficient production of human gamma interferon in tobacco protoplasts by genetically engineered brome mosaic virus RNAs.
We succeeded in producing human gamma interferon (IFN-gamma) in tobacco protoplasts in quantity using genetically engineered brome mosaic virus (BMV strain ATCC66). This strain of BMV produces two types of coat protein, a full-length coat protein (20K) and a truncated coat protein (19K) which are translated from the first and second initiation codons, respectively. We replaced the truncated coat protein gene with the IFN-gamma gene and synthesized BMV-IFN-gamma chimera RNAs using an in vitro transcription system. The BMV-IFN-gamma chimera RNAs were used to inoculate tobacco protoplasts together with BMV RNA 1 and RNA 2 and produced IFN-gamma to a level of 5 to 10% of total extracted proteins per infected protoplast after 24 h of incubation. The efficient production of IFN-gamma was attributed to the high translation activity of the BMV-IFN-gamma chimera RNA. We demonstrate that 24 nucleotides coding for the N-terminal amino acids of the full-length coat protein were probably involved in the high translation activity of the BMV-IFN-gamma chimera RNA.
Mori
M
M
Laboratory of Plant Pathology, Faculty of Agriculture, Kyoto University, Japan.
Zhang
G H
GH
Kaido
M
M
Okuno
T
T
Furusawa
I
I
D001483:Base Sequence; D002478:Cells, Cultured; D005818:Genetic Engineering / Q000379:methods; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis* / Q000494:pharmacology; D008969:Molecular Sequence Data; D009029:Mosaic Viruses / Q000235:genetics* / Q000502:physiology; D009838:Oligodeoxyribonucleotides; D010947:Plants, Toxic; D014176:Protein Biosynthesis; D011523:Protoplasts; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins; D012845:Sindbis Virus / Q000187:drug effects / Q000472:pathogenicity / Q000502:physiology; D014026:Nicotiana / Q000378:metabolism*; D014779:Virus Replication
D009838:Oligodeoxyribonucleotides; D012367:RNA, Viral; D011994:Recombinant Proteins; D007371:Interferon-gamma