In humans, transcription of the gonadotropin-releasing hormone (GnRH) gene can be initiated at two transcription start sites to produce different GnRH mRNAs. The upstream transcription start site is used only in reproductive tissues and tumors. To determine if a similar pattern of GnRH gene expression exists in non-human primates, we cloned GnRH cDNA from rhesus monkey hypothalamic RNA using reverse transcriptase-polymerase chain reaction (RT-PCR) and the 5' flanking region of the monkey GnRH gene by PCR. A 96% similarity between monkey and human GnRH cDNA was found with 94% similarity in the upstream promoter region. An upstream transcriptional start site, was identified in cynomolgus monkey testicular mRNA, 504 base pairs upstream from the hypothalamic site, which was different from that identified in the human GnRH gene. Various cynomolgus monkey reproductive tissues were found to utilize this upstream transcriptional start site. In contrast, no evidence was found for the use of upstream transcriptional start sites in rat testis or placenta, suggesting that the reproductive tissue specificity of the upstream transcription start site may be a primate specific feature.