Expression of functional replication protein from tomato golden mosaic virus in transgenic tobacco plants

The A component of the bipartite genome of the geminivirus tomato golden mosaic virus (TGMV) encodes the viral protein (AL1) that is required for viral DNA replication. We have constructed transgenic Nicotiana benthamiana plants in which the AL1 open reading frame is transcribed under the control of the cauliflower mosaic virus 35S promoter. The transgenic plants, which were phenotypically normal, produced a single transcript from the 35S-AL1 construct and a 40-kDa protein that cross-reacted with a polyclonal antiserum raised against AL1 protein overproduced in Escherichia coli. Six of nine transgenic lines complemented a TGMV A variant with a mutation in AL1 when coinoculated with the B component of the TGMV genome. Single- and double-stranded forms of the B component were synthesized in leaf discs from a complementing, transgenic line in the absence of TGMV A. These results establish that the transgenic plants express functional AL1 protein and show that this viral protein is not only required, but sufficient, for single- and double-stranded replication of TGMV DNA in the presence of host proteins. These results also show that the AL1 protein is not by itself a determinant of disease or pathogenesis.