THE INFLUENCE OF SMALL CHANGES IN TRANSGENE TRANSCRIPTION ON HOMOLOGY-DEPENDENT VIRUS-RESISTANCE AND GENE SILENCING

A tissue culture/regeneration scheme was employed to introduce epigene tic variation into tobacco lines that either do (T4) or do not (T19) e xhibit post-transcriptional silencing of beta-glucuronidase (GUS) tran sgenes. Homology-dependent resistance to a potato virus X derivative c arrying GUS (PVX-GUS), an indicator of post-transcriptional gene silen cing, was used as a screen for epigenetic changes. Regenerant families derived from the high-expressing line T19 and nontransformed tobacco remained completely susceptible to PVX-GUS, like the progenitors. Twen ty out of twenty T4 regenerant families were also susceptible to PVX-G US and were therefore very distinct from the low-expressing, PVX-GUS-r esistant T4 progenitor line. This change to PVX-GUS susceptibility was not associated with DNA structural changes, but was associated with s light increases in GUS mRNA accumulation, increased 35S promoter methy lation and correspondingly decreased transcription of GUS, These resul ts suggest that homology-dependent virus resistance and gene silencing are very sensitive to small changes in transgene transcription.