Summary
The geminivirus tomato golden mosaic virus (TGMV) replicates in nuclei and expresses genes from high copy number DNA episomes. The authors used TGMV as a vector to determine whether episomal DNA can cause silencing of homologous, chromosomal genes. Two markers were used to asses silencing: (1) the sulfur allele (su) of magnesium chelatase, an enzyme required for chlorophyll formation; and (2) the firefly luciferase gene (luc). Various portions of both marker genes were inserted into TGMV in place of the coat protein openβreading frame and the constructs were introduced into intact plants using particle bombardment. When TGMV vectors carrying fragments of su (TGMV::su) were introduced into leaves of wild type Nicotiana benthamiana, circular, yellow spots with an area of several hundred cells formed after 3β5 days. Systemic movement of TGMV::su subsequently produced varigated leaf and stem tissue. Fragments that caused silencing included a 786 bp 5' fragment of the 1392 bp su cDNA in sense and antiβsense orientation, and a 403 bp 3' fragment. TGMV::suβinduced silencing was propogated through tissue culture, along with the viral episome, but was not retained through meiosis. Systemic downregulation of a constitutively expresse luciferase transgene in plants was achieved following infection with TGMV vectors carrying a 623 bp portion of luc in sense or antiβsense orientation. These results establish that homologous DNA sequences localized in nuclear episomes can modulate the expression of active chromosomal genes.