We have studied the activity for the insulin-like growth factor binding protein-1 (IGFBP-1) gene promoter in human endometrial stromal cells by transient transfection. The promoter activity derived from p3.6CAT or p3.6Luc (3400 bp IGFBP-1 promoter 5' to the reporter gene chloramphenicol acetyltransferase or luciferase) was minimal in unstimulated cells. A time study over 13 days of culture showed that the promoter activity increased exponentially to > 10(4) fold in cells treated with medroxyprogesterone acetate (MPA) and relaxin (RLX). Induction of the IGFBP-1 gene promoter activity by hormones was similar to the secretion pattern of IGFBP-1 in endometrial stromal cells. MPA alone caused a moderate induction, 3-40-fold increase over the control. Deletion analysis showed that two regions in the IGFBP-1 gene promoter were responsible for the activation of the IGFBP-1 gene. The basal promoter region, termed bp1-A (+68 bp to -1.205 kb), contains multiple sections of regulatory sequence including a cis-element CCAAT (-72 bp). A DNase I protection assay in the bp-1A region revealed four distinct binding regions, one of which contained the CCAAT box region. Another promoter region, termed bp1-B (-2.6 to -3.4 kb), mediated 95% of the total promoter activity in endometrial stromal cells. The bp1-B region also contains multiple regulatory sequences. Mutation and DNase I protection assay suggest that Sp1-like binding site at -2.63 kb was a regulatory site responsible for the activation of IGFBP-1 gene promoter.