Previous biochemical studies have shown that c-fos mRNA expression is rapidly and dramatically induced by estrogen in the immature rat uterus. In this work we have used in situ hybridization and immunocytochemistry to analyze the cellular pattern of proto-oncogene induction following administration of estradiol. The c-fos transcript and protein levels were low in control animals. Three hours after hormone treatment, the luminal and glandular epithelium, stroma, and myometrium all showed a clear increase in mRNA levels of the proto-oncogene as judged by in situ hybridization; this increase was most pronounced in the epithelial cells. Immunocytochemistry also demonstrated Fos expression in all cell layers with a higher fraction of cells staining in the epithelial and stromal layers than in the myometrium. Estradiol thus increased expression of c-fos in all major cell types of the immature uterus, although each tissue layer exhibited a specific pattern and degree of fos transcript and protein expression. This proto-oncogene is thus a marker for estrogen action in all uterine cell types at this stage of development. Since estrogen stimulates proliferation in all cell layers of the immature uterus, the cellular pattern of fos expression also correlates with the proliferative response to the hormone.