The promoter of the FSH receptor (R) gene has been cloned from several species. Although some of its regulatory elements have been identified, its function still remains poorly characterized. Using transient transfections of luciferase reporter constructs, driven by various fragments of the murine (m) FSHR promoter, we identified a cell-specific promoter region. This domain is located in the distal part of the mFSHR promoter, -1,110 to -1,548 bp upstream of the translation initiation site, and it contains two steroidogenic factor 1 (SF-1) like binding sites (SLBS). The cellular levels of SF-1 mRNA and protein closely correlated in various steroidogenic cell lines with activity of the transfected mFSHR promoter/luciferase reporter construct carrying the distal activator domain. A dose-dependent increase in FSHR promoter activity was shown in nonsteroidogenic HEK 293 cells transiently transfected with SF-1 cDNA. SF-1 was found to bind to a nonconsensus 5'-CAAGGACT-3' SLBS-3 motif in the distal part of the promoter; formation of the SF-1/SLBS-3 complex could be reversed by addition of SF-1 antibody. Mutation in the SLBS-3 domain abolished the SF-1/SLBS-3 complex in gel-shift assays and led to a significant loss of SF-1-mediated mFSHR promoter activity. The second SLBS appeared to have minor role in SF-1-regulated mFSHR expression. In conclusion, we have identified a regulatory domain in the mFSHR promoter participating in the cell-specific regulation of FSHR expression. We demonstrated for the first time that the mFSHR promoter possesses functional SF-1 binding sites and thus belongs to the group of SF-1-regulated genes. These findings provide further evidence for the key role of SF-1 in the regulation of genes involved in gonadal differentiation and endocrine functions.