Sp1, steroidogenic factor 1 (SF-1), and early growth response protein 1 (Egr-1) binding sites form a tripartite gonadotropin-releasing hormone response element in the rat luteinizing hormone-β gene promoter: An integral role for SF-1

Recently, several cis-regulatory elements that play roles in LH beta gene expression, and their cognate DNA-binding transcription factors, have been identified. These factors include Sp1, steroidogenic factor-1 (SF-1), and early growth response protein 1 (Egr-1). Using the GH, pituitary cell line (which lacks SF-1) as a model, we demonstrate that expression of SF-1 or Egr-1 increases rat LH beta gene promoter activity but has little effect on the fold response to GnRH. However, expression of both SF-1 and Egr-1 synergistically enhances LH beta gene promoter activity and prevents further stimulation of activity by GnRH. Mutations in the Spl binding sites of the rat LH beta gene promoter decrease GnRH responsiveness, whereas mutations in the SF-1 and/or Egr-1 binding sites alone have little effect on the GnRH response. Combinatorial mutations in both the Sp1 and Egr-1 binding elements result in almost complete loss of the GnRH response. In contrast, in GH, cells cotransfected with SF-1, mutations in the Sp1, SF-1, or Egr-1 binding elements independently decrease GnRH responsiveness. In L beta T2 cells, a gonadotrope-derived cell line that expresses SF-1 endogenously, mutations in either the Sp1 or Egr-1 binding elements decrease GnRH responsiveness. These data suggest that the Sp1, SF-1, and Egr-1 binding sites form a tripartite GnRH response element in the rat LH beta gene promoter. Changes in the spacing between the upstream Sp1 binding sites and the downstream SF-1/Egr-1 binding elements reduce the response to GnRH. SF-1, while having little direct effect on GnRH responsiveness, has a critical role in integrating the effects of Spl and Egr-1.