MUTATIONAL ANALYSIS OF THE TOBACCO VEIN MOTTLING VIRUS GENOME

We have used a cDNA clone of the potyvirus, tobacco vein mottling viru s, to construct 19 mutants bearing 12-nt insertions in the viral genom e. These mutants display a variety of phenotypes in inoculated tobacco plants or protoplasts. All mutants with insertions in P3, CI, 6K, Nla , or Nib failed to produce detectable amounts of progeny viral RNA in protoplasts or plants which suggests that they all may be directly inv olved in replication. Mutants (one in P1 and one in HCpro) presumably affected in polyprotein processing also did not replicate in plants or protoplasts. Seven mutants, with insertions in the 5' noncoding regio n, P1, HCpro, or CP regions of the genome, were able to infect protopl asts. Three of the 7 mutants (1 in the 5' noncoding region and 2 in HC pro) were able to infect protoplasts but not plants. The remaining 4 m utants replicated in protoplasts and were able to cause systemic infec tion in plants. The mutation in the CP had no effect on virus accumula tion or symptom development in inoculated plants, whereas the other 3 (1 in PI and a in HCpro) induced cyclical patterns of symptom expressi on. These symptoms ranged from very mild to wild-type-like as new leav es emerged and, as the plants continued to grow, this pattern was repe ated. These results support the assignment of roles in replication to five coding regions in the genome and demonstrate that sequence altera tions in many parts of other regions of the viral genome may have pron ounced effects on replication and the expression of disease symptoms. (C) 1994 Academic Press, Inc.