The promoter of murine follicle-stimulating hormone receptor: functional characterization and regulation by transcription factor steroidogenic factor 1

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.     

Activation of luteinizing hormone beta gene by gonadotropin-releasing hormone requires the synergy of early growth response-1 and steroidogenic factor-1.

We have previously shown that early growth response (Egr) 1-deficient mice exhibit female infertility, reflecting a luteinizing hormone (LH) beta deficiency. Egr-1 activates the LHbeta gene in vitro through synergy with steroidogenic factor-1 (SF-1), a protein required for gonadotrope function. To test if this synergy is essential for gonadotropin-releasing hormone (GnRH) stimulation of LHbeta, we examined the activity of the LHbeta promoter in the gonadotrope cell line LbetaT2. GnRH markedly stimulated the LHbeta promoter (15-fold). Mutation of either Egr-1 or SF-1 elements within the LHbeta promoter attenuated this stimulation, whereas mutation of both promoter elements abrogated GnRH induction of the LHbeta promoter. Furthermore, GnRH stimulated Egr-1 but not SF-1 expression in LbetaT2 cells. Importantly, overexpression of Egr-1 alone was sufficient to enhance LHbeta expression. Although other Egr proteins are expressed in LbetaT2 cells and are capable of interacting with SF-1, GnRH stimulation of Egr-1 was the most robust. We also found that the nuclear receptor DAX-1, a repressor of SF-1 activity, reduced Egr-1-SF-1 synergy and diminished GnRH stimulation of the LHbeta promoter. We conclude that the synergy between Egr-1 and SF-1 is essential for GnRH stimulation of the LHbeta gene and plays a central role in the dynamic regulation of LHbeta expression.