Chromatin structure and transcription factor activity collaborate to set the transcription level of a gene. Our understanding of the relative contributions of each of these factors at a specific gene is limited. We studied the effects of an altered chromatin environment on the activity of the estrogen-responsive pS2 promoter. We created stable cell lines with the pS2 promoter situated in an alternative chromatin site in addition to it being in its native site. Both promoters were estrogen-responsive for estrogen receptor alpha (ERalpha) recruitment, but transcription was inducible only at the native site. At the recombinant site, transcription was high and constitutive. Higher histone H3 and H4 acetylation (acH3 and acH4), as well as trimethylated lysine 4 on histone H3 levels, was observed at the recombinant site compared to the native site in vehicle treated cells. Inhibition of histone deacetylases (HDACs) resulted in increased acH4, but only modest increases in acH3, ERalpha binding and basal transcription at the native pS2 site. Inhibiting HDACs had no effect on transcription from the recombinant site. These data suggest that highly active chromatin is not only permissive for transcription, but can override the requirement for the transcription factor at an inducible promoter.