Virus recovery is induced in Brome mosaic virus p2 transgenic plants showing synchronous complementation and RNA-2-specific silencing.

Nicotiana benthamiana plants expressing Brome mosaic virus (BMV) p2 protein complemented replication of RNAs1 + 3 but, surprisingly, supported little or no replication of RNA-2. Despite this, the p2 transgenic plants were able to support systemic migration of RNAs-1 and -3. Kinetic analyses showed identical degradation rates for RNAs-2 and -3, greatly detracting from the concept of an induction of an RNA-2-specific degradation system. Deletion analysis identified a 200-nucleotide sequence that may contribute to silencing in a context-specific manner. When R1 progeny of a severely silencing p2 transgenic line were tested for virus resistance, three different classes of reactions were observed. In class 1 and class 3 plants, the virus moved systemically and showed various extents of RNA-2 silencing. However, in class 2 plants, there was a stochastic onset of post-transcriptional silencing in the systemic leaves that was reminiscent of virus recovery. Plants showing recovery tended to have a greater number of transgene loci than did those exhibiting component-specific silencing. The induction of silencing did not appear to be dependent solely on the combined steady state levels of the transgene and viral RNA. Some plants transformed with a p2 frameshift construct showed a complete silencing phenotype, but none showed RNA-2-specific silencing. While the relationship between the two types of silencing remains unclear, we speculate that our observations reflect early events in the induction of virus recovery.