One-step selection of artificial transcription factors using an in vivo screening system

Gene expression is regulated in large part at the level of transcription under the control of sequence-specific transcriptional regulatory proteins. Therefore, the abil-ity to affect gene expression at will using sequence-specific artificial transcription factors would provide researchers with a powerful tool for biotechnology re-search and drug discovery. Previously, we isolated 56 novel sequence-specific DNA-binding domains from the human genome by in vivo selection. We hypothesized that these domains might be more useful for regulating gene expression in higher eukaryotic cells than those selected in vitro using phage display. However, an un-predictable factor, termed the “context effect”, is asso-ciated with the construction of novel zinc finger tran-scription factors--- DNA-binding proteins that bind specifically to 9-base pair target sequences. In this study, we directly selected active artificial zinc finger proteins from a zinc finger protein library. Direct in vivo selec-tion of constituents of a zinc finger protein library may be an efficient method for isolating multi-finger DNA binding proteins while avoiding the context effect