Analysis of multiple copies of geminiviral DNA in the genome of four closely related Nicotiana species suggest a unique integration event

Previously, we discovered multiple direct repeats of geminivirus-related DNA (GRD) sequences clustered at a single chromosomal position in Nicotiana tabacum (tobacco). Here we show that, in addition to tobacco, multiple copies of these elements occur in the genomes of three related Nicotiana species, all in the section Tomentosae: N. tomentosiformis, N. tomentosa and N. Kawakamii, but not in 9 other more distantly related Nicotiana species, nor in various other solanaceous and non-solanaecous plants. DNA sequence analysis of 18 GRD copies reveal 4 distinct, but highly related, sub-families: GRD5, GRD3 and GRD53 in tobacco, GRD5 in N. tomentosiformis and N. kawakamii; and GRD2 in N. tomentosa. In addition to novel sequences, all elements share significant but varying lengths of DNA sequence similarity with the geminiviral replication origin plus the adjacent rep gene. There is extended sequence similarity to REP protein at the deduced amino acid sequence level, including motifs associated with other rolling circle replication proteins. Our data suggest that all GRD elements descend from a unique geminiviral integration event, most likely in a common ancestor of these Tomentosae species.