Rapid, high-level expression of biologically active alpha-trichosanthin in transfected plants by an RNA viral vector.
alpha-Trichosanthin, a eukaryotic ribosome-inactivating protein from Trichosanthes kirilowii, inhibits the replication of the human immunodeficiency virus (HIV) in vitro. The alpha-trichosanthin gene was placed under the transcriptional control of a tobamovirus subgenomic promoter in a plant RNA viral vector. Two weeks after inoculation, transfected Nicotiana benthamiana plants accumulated alpha-trichosanthin to levels of at least 2% of total soluble protein. The recombinant alpha-trichosanthin was purified and its structural and biological properties were analyzed. The 23-amino acid signal peptide was recognized by N. benthamiana and the processed enzyme caused a concentration-dependent inhibition of protein synthesis in vitro. The high level of heterologous gene expression observed in these studies is due to the unique features of the RNA viral-based transfection system.
Kumagai
M H
MH
Biosource Genetics Corp., Vacaville, CA 95688.
Turpen
T H
TH
Weinzettl
N
N
della-Cioppa
G
G
Turpen
A M
AM
Donson
J
J
Hilf
M E
ME
Grantham
G L
GL
Dawson
W O
WO
Chow
T P
TP
D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000378:metabolism*; D001483:Base Sequence; D004305:Dose-Response Relationship, Drug; D005822:Genetic Vectors / Q000235:genetics; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000378:metabolism; D011500:Protein Synthesis Inhibitors / Q000378:metabolism*; D014027:Tobacco Mosaic Virus / Q000235:genetics; D014162:Transfection; D015978:Trichosanthin / Q000096:biosynthesis* / Q000235:genetics
D000998:Antiviral Agents; D021382:Protein Sorting Signals; D011500:Protein Synthesis Inhibitors; D015978:Trichosanthin