Synthetic zinc finger transcription factor action at an endogenous chromosomal site - Activation of the human erythropoietin gene

We have targeted the activation of an endogenous chromosomal locus includin g the human erythropoietin gene using synthetic transcription factors. Thes e transcription factors are targeted to particular DNA sequences in the 5'- flanking region of the erythropoietin gene through engineering of a zinc fi nger DNA binding domain. The DNA binding domain is linked to a VP16 transcr iptional activation domain. We find that these synthetic transcription fact ors invariably activate transiently transfected templates in which sequence s within the 5' flank of the erythropoietin gene are fused to a luciferase reporter. The efficiency of activation under these circumstances at a defin ed site is dependent on DNA binding affinity. In contrast, only a subset of these same zinc finger proteins is able to activate the endogenous chromos omal locus. The activity of these proteins is influenced by their capacity to gain access to their recognition elements within the chromatin infrastru cture, Zinc finger transcription factors will provide a powerful tool to pr obe the determinants of chromatin accessibility and remodeling within endog enous chromosomal loci.