Expression of the rabies virus glycoprotein in transgenic tomatoes.
We have engineered tomato plants (Lycopersicon esculentum Mill var. UC82b) to express a gene for the glycoprotein (G-protein), which coats the outer surface of the rabies virus. The recombinant constructs contained the G-protein gene from the ERA strain of rabies virus, including the signal peptide, under the control of the 35S promoter of cauliflower mosaic virus. Plants were transformed by Agrobacterium tumefaciens-mediated transformation of cotyledons and tissue culture on selective media. PCR confirmed the presence of the G-protein gene in plants surviving selection. Northern blot analysis indicated that RNA of the appropriate molecular weight was produced in both leaves and fruit of the transgenic plants. The recombinant G-protein was immunoprecipitated and detected by Western blot from leaves and fruit using different antisera. The G-protein expressed in tomato appeared as two distinct bands with apparent molecular mass of 62 and 60 kDa as compared to the 66 kDa observed for G-protein from virus grown in BHK cells. Electron microscopy of leaf tissue using immunogold-labeling and antisera specific for rabies G-protein showed localization of the G-protein to the Golgi bodies, vesicles, plasmalemma and cell walls of vascular parenchyma cells. In light of our previous demonstration that orally administered rabies G-protein from the same ERA strain elicits protective immunity in animals, these transgenic plants should provide a valuable tool for the development of edible oral vaccines.
McGarvey
P B
PB
Thomas Jefferson University, Department of Microbiology and Immunology, Philadelphia, PA 19107, USA. pmcgarve@lac.jci.tju.edu
Hammond
J
J
Dienelt
M M
MM
Hooper
D C
DC
Fu
Z F
ZF
Dietzschold
B
B
Koprowski
H
H
Michaels
F H
FH
D016960:Agrobacterium tumefaciens / Q000235:genetics; D000956:Antigens, Viral; D015152:Blotting, Northern; D015153:Blotting, Western; D017796:Caulimovirus / Q000235:genetics; D015870:Gene Expression; D006023:Glycoproteins / Q000235:genetics*; D007163:Immunosorbent Techniques; D018551:Solanum lycopersicum / Q000737:chemistry / Q000235:genetics*; D008854:Microscopy, Electron; D018515:Plant Leaves / Q000737:chemistry; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D015202:Protein Engineering; D012313:RNA / Q000032:analysis; D011994:Recombinant Proteins; D014162:Transfection; D014759:Viral Envelope Proteins / Q000235:genetics*
D000956:Antigens, Viral; D006023:Glycoproteins; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; C064951:glycoprotein G, Rabies virus; D012313:RNA