Epitope presentation system based on cucumber mosaic virus coat protein expressed from a potato virus X-based vector.
The Cucumber mosaic virus Ixora isolate (CMV) coat protein gene (CP) was placed under the transcriptional control of the duplicated subgenomic CP promoter of a Potato virus X (PVX)-based vector. In vitro RNA transcripts were inoculated onto Nicotiana benthamiana plants and recombinant CMV capsid proteins were identified on Western blots probed with CMV antibodies 5-7 days post-inoculation. PVX-produced CMV CP subunits were capable of assembling into virus-like particles (VLPs), which were visualized by electron microscopy. We further used the PVX/CMVCP system for transient expression of recombinant CMV CP constructs containing different neutralizing epitopes of Newcastle disease virus (NDV) engineered into the internal betaH-betaI (motif 5) loop. Both crude plant extracts and purified VLPs were immunoreactive with CMV antibodies as well as with epitope-specific antibodies to NDV, thus confirming the surface display of the engineered NDV epitope. Our study demonstrates the potential of PVX/CMVCP as an expression tool and as a presentation system for promising epitopes.
USDA-ARS Molecular Plant Pathology Laboratory, Beltsville, Maryland 20705, USA.
A
A
Natilla
R W
RW
Hammond
L G
LG
Nemchinov
D015153:Blotting, Western; D036022:Capsid Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D017799:Cucumovirus / Q000235:genetics / Q000276:immunology*; D000939:Epitopes / Q000096:biosynthesis* / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D016253:Microscopy, Immunoelectron; D008958:Models, Molecular; D009522:Newcastle disease virus / Q000235:genetics / Q000276:immunology; D017863:Potexvirus / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D011995:Recombination, Genetic; D014026:Nicotiana / Q000737:chemistry / Q000235:genetics; D022701:Virosomes / Q000302:isolation & purification / Q000648:ultrastructure
D036022:Capsid Proteins; D000939:Epitopes; D012367:RNA, Viral; D011994:Recombinant Proteins; D022701:Virosomes