Y. Choo et al., PROMOTER-SPECIFIC ACTIVATION OF GENE-EXPRESSION DIRECTED BY BACTERIOPHAGE-SELECTED ZINC FINGERS, Journal of Molecular Biology, 273(3), 1997, pp. 525-532
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.