We isolated and characterized the human oxytocin receptor gene. Southern blots indicated that the human genome has a single copy of the gene. Chromosomal localization by fluorescence in situ hybridization also showed that the gene was a single copy, assigned to 3p26.2 of the human chromosome. The gene spans approximately 17 kilobases and contains 3 introns and 4 exons. Exons 1 and 2 correspond to the 5'-non-coding region, followed by exons 3 and 4 encoding the amino acids of the receptor. Intron 3, which is the largest at 12 kilobases, separates the coding region immediately after the putative sixth transmembrane-spanning domain. The transcription start sites, demonstrated by primer extension analysis, lie 618 and 621 base pairs upstream of the methionine initiation codon. Near these putative transcription start sites, we found a TATA-like motif and a potential SP-1 binding site at about 30 and 65 base pairs, respectively. We also found other known binding sites of transcription regulating factors, such as AP-1, AP-2, GATA-1, Myb, nucleofactor-interleukin 6 binding consensus sequence, and an acute phase reactant-responsive element. No estrogen-responsive element was observed except three half-palindromic estrogen-responsive element motifs. Our findings of the oxytocin receptor gene structure should help to elucidate the mechanism by which the gene expression is induced drastically at parturition in the uterus and how the gene is regulated in other organs such as the mammary gland or central nervous system.