Transcriptional activation of gene expression by glucocorticoid hormones is mediated by the interaction of hormone-receptor complexes with specific DNA sequences called glucocorticoid responsive elements (GREs) (refs 1-3, see ref. 4 for review). Deletion of this sequence abolishes glucocorticoid induction of transcription. According to a current model, activation of the cytoplasmic receptor protein by hormone binding leads to its increased affinity for and translocation to the nucleus. However, recent reports that the oestradiol and progesterone receptors are localized in the nucleus in the absence of steroid led us to examine whether the free receptor interacts in vivo with its DNA binding site in the absence of hormone binding. We used the genomic footprinting technique to show that changes in in vivo protein-DNA interactions within the GREs of the tyrosine aminotransferase gene (TAT) can be detected only after hormone treatment in hepatoma cells. Such changes are not detected in fibroblast cells, in which the TAT gene is not expressed. Many of the changes in dimethylsulphate reactivity observed in the living cell are also found in vitro using cloned DNA and a partially purified glucocorticoid receptor.