Steroid receptor coactivator-1 (SRC-1) family members interact with steroid receptors, including estrogen receptor alpha (ERalpha) and progesterone receptor (PR), to enhance ligand-dependent transcription. However, the expression of ERalpha and SRC-1 was found to be segregated in distinct subsets of cells within the epithelium of the estrogen-responsive rat mammary gland. This finding was in contrast to the finding for the stroma, where significant numbers of cells coexpressed ERalpha and SRC-1. Treatment of animals with estrogen induced PR expression in the ERalpha-expressing mammary epithelial cells in the absence of detectable SRC-1 and did not affect the segregated pattern of SRC-1 and ERalpha expression. PR was neither expressed nor induced by estrogen treatment in stroma, despite the coexpression of ERalpha and SRC-1. These results suggest that SRC-1 is not necessary for ERalpha-mediated induction of PR in mammary epithelial cells and is also not sufficient for PR induction in stromal cells expressing both ERalpha and SRC-1. Furthermore, the expression of SRC-1 in a subpopulation of mammary epithelial cells distinct from those expressing ERalpha or PR raises the possibility that SRC-1 has cell type-specific functions other than simply to act as coactivator for ERalpha or PR in the mammary epithelium.