Chemical-inducible, ecdysone receptor-based gene expression system for plants.

Abstract

We have developed an inducible gene expression system with potential for field application using the ecdysone receptor (EcR) from the spruce budworm and the non-steroidal EcR agonist, methoxyfenozide. Chimeric transcription activators were constructed with EcR ligand binding domain, GAL4 and LexA DNA binding domains, and VP16 activation domain. In the presence of methoxyfenozide, the transcription activators induced expression of the luciferase reporter gene cloned downstream of a promoter containing GAL4A- or LexA-response element and a minimal 35S promoter. Low basal and high induced luciferase expression was optimized by cloning the activator and the reporter genes in different tandem orientations. Many transgenic Arabidopsis and tobacco plants were obtained with little or no basal expression in the absence of methoxyfenozide and inducible expression that was several fold higher than that observed with the constitutive 35S promoter. Moreover, gene expression was controlled over a wide range of methoxyfenozide concentration. Our results demonstrate that the inducible gene expression system based on the spruce budworm EcR ligand binding domain with methoxyfenozide as a ligand is very effective in regulating transgenes in plants. It is suitable for field applications because methoxyfenozide is commercially available and has an exceptional health and environmental safety profile.