PARENT SESSION
Signal Transduction

IDENTIFICATION OF SMAD BINDING ELEMENT WITHIN THE RAT FOLLICLE STIMULATING HORMONE -SUBUNIT .

Suszko, Magdalena¹, Woodruff, Teresa¹, ¹

ABSTRACT- Follicle stimulating hormone (FSH) is an essential component of the mammalian reproductive axis. It is a member of structurally related pituitary glycoproteins consisting of two subunits, a common -subunit and a unique -subunit. The subunit constitutes possible target for regulation of FSH by a variety of positive and negative stimuli. Synthesis and release of FSH from cells of the anterior pituitary is highly regulated by activin, a member of the TGF-superfamily of ligands. Most of the TGF ligands signal through a common pathway that involves cytoplasmic and nuclear mediators, Smads. Specific transcriptional elements of FSH regulation by activin, however, are still largely unknown. Elucidation of molecular mechanisms underlying activin action on FSH is necessary for understanding of normal reproductive function. We used the pituitary-derived cell line, LT2, as an in vitro model of FSH regulation to identify and characterize activin responsive genetic elements. The cells were transiently transfected with rat FSH promoter fused to the luciferase reporter gene. Activin treatment resulted in the up-regulation of the luciferase expression, while co-treatment with follistatin prevented this induction. Based on the transfection experiments of different deletion constructs a minimal activin responsive FSH promoter region was identified and further characterized by electrophoretic mobility shift assay. Competitive binding and antibody shift experiments identified a region of 29 nucleotides that contains Smad binding element. Further studies are being performed to narrow down the specific Smad binding element and to identify other factors contained within the nuclear transcription complex.

KEY WORDS: Activin, Follicle Stimulating Hormone , Smad