Isolation and characterization of the rat prolactin-releasing peptide gene: Multiple TATA boxes in the promoter region

The prolactin-releasing peptide (PrRP) gene is a novel bioactive peptide ex pressed in very restricted regions in the brain. To explore the molecular m echanism of PrRP gene expression, we cloned and characterized the gene and its promoter region. The gene spans approximately 2.4 kb and contains three exons and two introns. 3'RACE analysis showed that a polyadenylation signa l 103 bp downstream from the stop codon was functional. Primer extension an alysis indicated three transcriptional start sites (TSSs) 92, 199, and 325 bp upstream from the translational start site. Interestingly, in addition t o the putative binding sites for SP1-1, AP-2, and Oct-2A, three characteris tic TATA boxes were identified close to these TSSs. Transient transfection study using a series of deletion mutants revealed that the middle TATA box is important for the promoter activity. Furthermore, the cloned 1.6 kh prom oter region was active only in neuron- and pituitary-derived cell lines, an d the promoter region -1600 similar to -800 bp worked as a negative regulat ory element. We demonstrated for the first time, the genomic organization a nd promoter function of the PrRP gene, and this knowledge will facilitate e lucidation of transcriptional control of the PrRP gene. (C) 2001 Academic P ress.