The conversion of androgens to estrogens is catalyzed by the aromatase cytochrome P-450 (P-450(AROM)) in a veriety of tissues and cell types in vertebrates. The manner in which aromatase activity is regulated appears to be quite different, even between different tissues of a single species. In the current study, we have determined the sequence of the 5' end of the aromatase mRNA in a rat Leydig tumor cell line, R2C, and in three rat tissue [adult ovary, perinatal amygdala (AmY), and medial basal hypothalamus-preoptic area (MBH-POA)], each of which shows different patterns of aromatase expression. S(1) nuclease protection and primer-extension analyses establish that the site of transcription initiation of the aromatase mRNA present in rat ovarh and the rat Leydig tumor cell line R2C is located approximately 97 nucleotides upstream from the initiator methionine. By contrast, although aromatase mRNA was detected in S(1) nuclease protection experiments using a probe derived from the aromatase open-reading frame, transcripts initiating at this site were absent from RNA samples prepared from perinatal rat AMY and MBH-POA tissue. S(1) mapping and sequencing of the 5' end of AMY and MBH-POA aromatase cDNAs indicate that the aromatase mRNA transcripts present in these rat neural tissues from perinatal animals contain a distinctive 5' terminus and are derived from a different promoter.