Abstract:
The tobacco etch potyvirus (TEV) polyprotein is proteolytically processed by three viral proteinases (NIa,
HC-Pro, and P1). While the NIa and HC-Pro proteinases each provide multiple functions essential for viral
infectivity, the role of the P1 proteinase beyond its autoproteolytic activity is understood poorly. To determine
if P1 is necessary for genome amplification and/or virus movement from cell to cell, a mutant lacking the entire
P1 coding region (ΔP1 mutant) was produced with a modified TEV strain (TEV-GUS) expressing β-glucuronidase
(GUS) as a reporter, and its replication and movement phenotypes were assayed in tobacco protoplasts
and plants. The ΔP1 mutant accumulated in protoplasts to approximately 2 to 3% the level of parental
TEV-GUS, indicating that the P1 protein may contribute to but is not strictly required for viral RNA
amplification. The ΔP1 mutant was capable of cell-to-cell and systemic (leaf-to-leaf) movement in plants but
at reduced rates compared with parental virus. This is in contrast to the S256A mutant, which encodes a
processing-defective P1 proteinase and which was nonviable in plants. Both ΔP1 and S256A mutants were
complemented by P1 proteinase expressed in a transgenic host. In transgenic protoplasts, genome amplification
of the ΔP1 mutant relative to parental virus was stimulated five- to sixfold. In transgenic plants, the level
of accumulation of the DP1 mutant was stimulated, although the rate of cell-to-cell movement was the same as
in nontransgenic plants. Also, the S256A mutant was capable of replication and systemic infection in P1-
expressing transgenic plants. These data suggest that, in addition to providing essential processing activity, the
P1 proteinase functions in trans to stimulate genome amplification.
