Thyroid stimulating hormone (TSH) is the pituitary glycoprotein hormone responsible for the synthesis and secretion of thyroid hormone from the thyroid gland. As such, it plays a crucial role in the maintenance of normal thyroid function, and is a necessary component in clinical treatment regimes for diseases such as thyroid cancer and thyroid disfunction. The long term goals of the laboratory are to understand the molecular mechanisms dictating the induction of thyrotrope differentiation in the anterior pituitary gland. Using targeted tumorigenesis in transgenic mice, we have created a novel thyrotrope cell line, termed 5.5alphaT-1, which expresses differentiated markers of the thyrotrope lineage. This cell line is a unique model system in which the molecular mechanism of tissue-specific gene regulation can be dissected. The objectives of this project are to: 1) Identify the cis-acting DNA elements required for thyrotrope-specific expression of the alpha- and beta-subunit genes of TSH. These studies will include the identification of thyrotrope- specific regulatory elements in the 5' flanking regions of the TSH subunit genes using transient transfection assays. Subsequent DNAse I protection assays will define distinct DNA-binding elements bound by thyrotrope nuclear factors. 2) Examine the hormonal regulation of TSH subunit genes by Thyrotropin Releasing Hormone (TRH). The 5.5alphaT-1 cells will be treated with TRH to characterize their response to the appropriate regulatory hormone and signal transduction cascades. Hormone-responsive elements will then be analyzed as in the first specific aim. This analysis will provide the necessary preliminary information for further characterization of the protein factors and signal transduction pathways involved in specifying the differentiated phenotype of thyrotrope cells in a future Independent Investigator grant application.
