COPPER-CONTROLLABLE GENE-EXPRESSION SYSTEM FOR WHOLE PLANTS

We describe a system for gene expression in plants based on the regulation mechanism of the yeast metallothionein (MT) gene. The system consists of two elements: (i) the yeast ace1 (activating copper-MT expression) gene encoding a transcription factor under control of the cauliflower mosaic virus (CaMV) 35S RNA promoter, and (ii) a gene of interest under control of a chimeric promoter consisting of the 90-base-pair domain A of the CaMV 35S RNA promoter linked to the ACE1 transcription factor-binding site. At elevated copper ion concentrations, the ACE1 protein changes conformation, binds to, and activates transcription from the chimeric promoter. To test the functioning of the system in plants, a construct containing the beta-glucuronidase (GUS) reporter gene under control of the chimeric promoter was prepared, and transgenic tobacco plants were produced. It was shown that GUS activity in the leaves of transgenic plants increased up to 50-fold, either after addition of 50 muM CuSO4 to the nutrient solution or after application of 0.5 muM CuSO4 to the plants in a foliar spray. This GUS expression was repressed after the removal of copper ions. The results show that the activity of the described chimeric promoter directly depends on copper ion concentration and that this system can be used in experiments that demand precise timing of expression.