To understand the regulation of the insulin-like growth factor binding protein-5 (IGFBP-5) gene expression, we have cloned the IGFBP-5 gene from rat genomic libraries and determined its genomic organization as well as the DNA sequence at the 5' flanking region of the gene. The rat IGFBP-5 gene spans at least 17 kilobases (kb) of the genome and contains 4 exons interrupted by 3 introns of approximately 10, 0.6 and 0.7 kb, respectively. Southern blot analysis of the rat chromosome DNA revealed a single copy gene for IGFBP-5 in the haploid genome. Primer extension experiments demonstrated a single transcriptional start site located at 772 nucleotides 5' of the ATG translational start codon. In addition to a TATA box and a CAAT box, multiple putative cis-regulatory elements, including an AP-1, an AP-2 and a binding site for progesterone receptor are present in the promoter region. This finding suggests that the IGFBP-5 gene is one of those that are expressed in a tissue-specific manner.