Tobacco mosaic virus defective RNAs expressing C-terminal methyltransferase domain sequences are severely impaired in long-distance movement in Nicotiana benthamiana.

Tobamovirus replicase proteins, which function in replication and gene expression, are also implicated in viral cell-to-cell and long-distance movement. The role(s) of Tobacco mosaic virus (TMV) 126-/183-kDa replicase protein in the complex movement process are not understood due to lack of systems that can separate the multiple steps involved. We previously developed a bipartite TMV-defective RNA (dRNA) system to dissect the role of the N-terminal methyltransferase (MT) domain in accumulation and cell-to-cell movement of dRNAs [Knapp, E., Danyluk, G.M., Achor, D., Lewandowski, D.J., 2005. A bipartite Tobacco mosaic virus-defective RNA (dRNA) system to study the role of the N-terminal methyltransferase domain in cell-to-cell movement of dRNAs. Virology 341, 47-58]. In the current study we analyzed long-distance movement of dRNAs in the presence of helper virus in Nicotiana benthamiana. dRNAs expressing approximately 50% of the MT domain (DeltaHinc151) moved long-distances in more than half of the plants. dRNAs expressing approximately 90% of the MT domain sequences (DeltaCla151) predominantly failed to accumulate in upper leaves. The helper virus moved systemically when inoculated alone or with a dRNA. In inoculated leaves, more DeltaHinc151-induced infection foci spread adjacent to class V veins compared to those of DeltaCla151. Consequently, DeltaHinc151 infected more class V veins than DeltaCla151. DeltaCla151 was only detected in bundle sheath cells, whereas DeltaHinc151 could accumulate in bundle sheath and phloem parenchyma cells of class V veins. However, the latter accumulation pattern did not always result in systemic accumulation of DeltaHinc151, suggesting that factors in addition to those affecting cell-to-cell movement played a role in long-distance movement.