Ecdysone Receptor-based Singular Gene Switches for Regulated Transgene Expression in Cells and Adult Rodent Tissues

• Published in: Molecular Therapy: Nucleic Acids
• Main Authors: Seoghyun Lee, Kyung-Cheol Sohn, Dae-Kyoung Choi, Minho Won, Kyeong Ah Park, Sung-Kyu Ju, Kidong Kang, Young-Ki Bae, Gang Min Hur, Hyunju Ro
• Format: Article
• Language: English
• Published: Elsevier 2016-01-01
• Subjects: adenovirus; controlled gene expression; ecdysone receptor (EcR)-based gene regulatory system; tetracycline; tebufenozide
• Online Access: http://www.sciencedirect.com/science/article/pii/S2162253117300926

Controlled gene expression is an indispensable technique in biomedical research. Here, we report a convenient, straightforward, and reliable way to induce expression of a gene of interest with negligible background expression compared to the most widely used tetracycline (Tet)-regulated system. Exploiting a Drosophila ecdysone receptor (EcR)-based gene regulatory system, we generated nonviral and adenoviral singular vectors designated as pEUI(+) and pENTR-EUI, respectively, which contain all the required elements to guarantee regulated transgene expression (GAL4-miniVP16-EcR, termed GvEcR hereafter, and 10 tandem repeats of an upstream activation sequence promoter followed by a multiple cloning site). Through the transient and stable transfection of mammalian cell lines with reporter genes, we validated that tebufenozide, an ecdysone agonist, reversibly induced gene expression, in a dose- and time-dependent manner, with negligible background expression. In addition, we created an adenovirus derived from the pENTR-EUI vector that readily infected not only cultured cells but also rodent tissues and was sensitive to tebufenozide treatment for regulated transgene expression. These results suggest that EcR-based singular gene regulatory switches would be convenient tools for the induction of gene expression in cells and tissues in a tightly controlled fashion.