An Improved Tet-On System for Gene Expression in Neurons Delivered by a Single Lentiviral Vector

作者：Tian, Xuebi; Wang, Gongming; Xu, Ying; Wang, Ping; Chen, Shasha; Yang, Hui; Gao, Feng; Xu, Aijun; Cao, Fei; Jin, Xiaogao; Manyande, Anne; Tian, Yuke^*

来源：Human Gene Therapy, 2009, 20(2): 113-123.

DOI：10.1089/hum.2008.018

• TETRACYCLINE-RESPONSIVE PROMOTERS
• REGULATED TRANSGENE EXPRESSION
• IN-VIVO
• MAMMALIAN-CELLS
• ENOLASE PROMOTER
• VIRAL EVOLUTION
• TIGHT CONTROL
• BRAIN
• EFFICIENT
• TRANSDUCTION

In most cases, the successful application of gene therapy requires the development of vectors that can provide regulated control of therapeutic gene expression. We have reconstituted the Tet-On (tetracycline-regulated transgene expression) two-component system in a single lentiviral vector with insertion of a chicken chromatin insulator (cHS4) element between the two expression cassettes. Optimization of this vector included an improved reverse tetracycline-dependent trans-activator (rtTA) sequence developed through HIV viral evolution, and an rtTA-dependent, Tet-responsive element containing modifications of the tetO sequence (TRE-tight1) to improve leaky basal transcription. Transduction of HeLa cells with these lentiviral vectors resulted in a high level of rtTA expression in the presence of doxycycline. In neuronal cells, rtTA expression driven by a neuron-specific enolase (NSE) promoter was more efficient than gene expression from a murine cytomegalovirus promoter. Transgene expression from the NSE promoter also provided tightly regulated gene expression in neurons in vivo.

• 出版日期2009-2
• 单位山东省立医院; 中国医科大学; 华中科技大学