Tetracycline repressor, tetR, rather than the tetR-mammalian cell transcription factor fusion derivatives, regulates inducible gene expression in mammalian cells.

This article describes the first (to our knowledge) tetracycline-inducible regulatory system that demonstrates that the tetracycline repressor (tetR) alone, rather than tetR-mammalian cell transcription factor fusion derivatives, can function as a potent trans-modulator to regulate gene expression in mammalian cells. With proper positioning of tetracycline operators downstream of the TATA element and of human epidermal growth factor (hEGF) as a reporter, we show that gene expression from the tetracycline operator-bearing hCMV major immediate-early enhancer-promoter (pcmvtetO) can be regulated by tetR over three orders of magnitude in response to tetracycline when (1) the reporter was cotransfected with tetR-expressing plasmid in transient expression assays, and (2) the reporter unit was stably integrated into the chromosome of a tetR-expressing cell line. This level of tetR-mediated inducible gene regulation is significantly higher than that of other repression-based mammalian cell transcription switch systems. In an in vivo porcine wound model, close to 60-fold tetR-mediated regulatory effects were detected and it was reversed when tetracycline was administered. Collectively, this study provides a direct implementation of this tetracycline-inducible regulatory switch for controlling gene expression in vitro, in vivo, and in gene therapy.