We have investigated chromatin organization over the MMTV LTR, a promoter regulated by steroid hormones. The studies were performed on cell lines containing BPV-based episomal constructs. Nucleosome positioning was determined by localization of sites sensitive to the enzyme micrococcal nuclease, or to the chemical MPE-Fe(II). Experiments with both reagents indicate that nucleosomes are specifically positioned in MMTV LTR chromatin. In the absence of hormone a regular cutting pattern is obtained, with cleavage sites at +136, -60, -250, -444, -651, -826 and -1019 relative to the Cap site. In the presence of hormone the cutting pattern is unchanged, except for a region between -60 and -250 that becomes hypersensitive to MPE-Fe(II). This region contains the DNA sequences to which steroid receptor complexes bind during transcriptional activation. Our results indicate that this region is associated in chromatin from uninduced cells with a macromolecular complex (probably a nucleosome core), and this complex is displaced (or modified) upon binding of activated receptor.