REPLICASE-MEDIATED RESISTANCE TO ALFALFA MOSAIC-VIRUS

Tobacco plants transformed with the P1 and P2 replicase genes of alfal fa mosaic virus (AIMV) have been shown to produce functional replicase proteins, permitting their infection with AIMV inocula lacking the ge nome segments encoding P1 and P2, respectively. To see whether express ion of a mutant P2 protein would interfere with the assembly of a func tional replicase complex, tobacco plants were transformed with modifie d P2 genes. When plants were transformed with a P2 gene encoding an N- terminally truncated protein which mimicked the tobacco mosaic virus 5 4K protein, no resistance was observed with 10 independent lines of tr ansformants. Similarly, when the GDD motif in the full-length P2 prote in was changed into VDD, no resistance was observed in 14 transgenic l ines. However, when the GDD motif was changed into GGD (5 lines), GVD (15 lines), or DDD (13 lines), 20 to 30% of the transgenic lines showe d a high level of resistance to AIMV infection. This resistance was ef fective to inoculum concentrations of 10 to 25 mu g/ml of virus and 10 0 mu g/ml of viral RNA, causing severe necrosis of control plants. For all transgenic lines, the expression of the transgenes was analyzed a t the RNA level. With the GGD, GVD, and DDD mutants, resistance was ge nerally observed in plants with a relatively high expression level. Th is indicates that the resistance is due to the mutant replicase rather than to an RNA-mediated cosuppression phenomenon. (C) 1995 Academic P ress, Inc.