Plasma membrane display of anti-viral single chain Fv fragments confers resistance to tobacco mosaic virus

We tested the hypothesis that membrane-anchored anti-viral antibodies can c onfer viral resistance to transgenic plants. A heterologous expression syst em was developed for plasma membrane targeting of anti-viral antibodies usi ng mammalian transmembrane domains. A tobacco mosaic virus (TMV) neutralizi ng single-chain Fv antibody fragment (scFv24) was targeted to the endoplasm ic reticulum and integrated into the plasma membrane of tobacco cells, usin g mammalian signal peptides and membrane receptor transmembrane domains. Th e human platelet-derived growth factor receptor (PDGFR) transmembrane domai n or the T-cell receptor beta-domain (TcR beta) transmembrane domain was fu sed to the C-terminus of TMV-specific scFv24 to target expression of scFv24 as an extracellularly facing plasma membrane protein. Western blot and ELI SA analyses were carried out to confirm functional expression of the recomb inant fusion proteins scFv24-PDGFR and scFv24-TcR beta in transgenic tobacc o suspension cultures and transgenic plants. Immunofluorescence and electro n microscopy showed that the TcR beta transmembrane domain targeted scFv24 to the tobacco plasma membrane. Bioassays of viral infection showed that tr ansgenic tobacco plants expressing scFv24-TcR beta were resistant to TMV in fection. These results demonstrated that membrane anchored anti-viral antib ody fragments are functional, can be targeted to the plasma membrane in pla nta and are a novel approach for engineering disease-resistant crops.