Homology-dependent resistance to tomato ringspot nepovirus in plants transformed with the VPg–protease coding region1

The coding region for the VPg (genome-linked viral protein) and protease (Pro) of a raspberry isolate (Rasp-2) of tomato ringspot virus (ToRSV) was cloned into a plant expression vector and transformed into Nicotiana benthamiana using Agrobacterium tumefaciens. Transgenic lines were also produced that expressed two mutant derivatives of the VPg–Pro coding region (i.e., containing point mutations in the putative catalytic triad and substrate-binding pocket of the protease). A large proportion of the R1 transgenic lines displayed extreme resistance to ToRSV (raspberry isolate Rasp-1, 99% homology to the transgene at the nucleotide level), especially when transformed with the wild-type VPg–Pro coding region. Several transgenic lines displaying extreme resistance to ToRSV were further tested at the R2 generation. These R2 lines were resistant to Rasp-1 isolate but were susceptible to GYV isolate (79% homology to the transgene). These transgenic lines were also resistant to a potato virus X (PVX) hybrid containing the VPg–Pro coding region of ToRSV (Rasp-2 isolate) but were susceptible to the wild-type PVX. These results imply a role for homology-dependent silencing as a mechanism for resistance in these lines.