Expression of active enzymes from an inducible tomato-mosaic-virus-based vector in cultured transgenic tobacco BY-2 cells

Abstract

Previously, we reported on an inducible viral vector system for foreign protein production in cultured plant cells, in which the transcription of the recombinant viral vector RNA encoding a foreign protein is controlled by an estradiol-inducible promoter. In this study, we used the inducible virus vector system to test the efficiency of a modified tomato mosaic virus (ToMV) encoding the movement protein. The virus inducibly produced a foreign jellyfish green fluorescent protein encoded in the virus in transgenic tobacco BY-2 suspension-cultured cells as efficiently as modified ToMV without the movement protein. We then produced transgenic BY-2 cell lines ER–ToMV–MP–DHFR and ER–ToMV–MP–GUS, which encoded modified ToMV with Escherichia coli dihydrofolate reductase (DHFR) and β-glucuronidase (GUS), respectively. After estradiol was added to the cell culture, DHFR and GUS activity was detected in the ER–ToMV–MP–DHFR and ER–ToMV–MP–GUS cells, respectively. In contrast, no DHFR or GUS activity was detected in untreated transgenic lines. Three days after induction, DHFR accumulation accounted for up to 15% of the total soluble proteins extracted from the cells, indicating that an inducible viral vector is an effective option for efficiently producing active enzymes in cultured plant cells.