Six mitochondrial genome sequences, showing strong similarity to the glucocorticoid responsive element consensus sequence (GRE), four localized within the cytochrome c oxidase (COX) subunit I and III genes (GREs I–IV) and two within the D-loop region (GREs a and b) have been examined as binding sites of glucocorticoid receptor (GR) from rat liver cytosol. Purified GR from rat liver cytosol binds with high specificity to all potential mitochondrial GREs, as shown by filter retention and gel shift assays. Specific binding of protein(s), present in a mitochondrial extract from dexamethasone-induced mice, to all six putative mitochondrial GREs was also documented by the same methodology. Both purified GR and protein(s) from mitochondrial extract give the same band in the gel retardation assay. Using monospecific anti-glucocorticoid receptor polyclonal antibody (EP), a supershift of the gel retarded protein-DNA band was obtained. These results demonstrate that the mitochondrial genome sequences examined have characteristics of GREs, since they show the capacity to specifically bind the respective receptor protein. These findings support the hypothesis that the mitochondrial genome is a primary site of action of steroid and thyroid hormones (Sekeris C.E.: The mitochondrial genome: a possible primary site of action of steroid hormones, In vivo 4 (1990) 317–320).