PROMOTER-SPECIFIC ACTIVATION OF GENE-EXPRESSION DIRECTED BY BACTERIOPHAGE-SELECTED ZINC FINGERS

It has been shown that sequence-specific DNA-binding domains containin g zinc fingers can be selected from libraries displayed on filamentous bacteriophage. The affinity and specificity of these peptides are wel l characterised in vitro, but few data are available to demonstrate sp ecific DNA binding and discrimination between closely related DNA sequ ences in vivo. Transient transactivation assays were performed in mamm alian cells, using expression plasmids which produce different amounts of a model transcription factor containing a phage-selected zinc fing er DNA-binding domain, and reporter plasmids which carry systematic va riations of the promoter sequence. When the intracellular concentratio n of the transcription factor was appropriate, activation of gene expr ession was absolutely dependent on a promoter having the same DNA sequ ence as that originally used to select the zinc finger domain by phage display. However, excessive intracellular concentrations of the trans cription factor resulted in some less-specific DNA binding, leading to gene activation from similar promoters containing a maximum of two ba se changes. Thus, provided delivery is carefully controlled, highly sp ecific control of gene expression in vivo can be achieved using artifi cial transcription factors containing phage-selected zinc finger DNA-b inding domains. (C) 1997 Academic Press Limited.