The human glycoprotein hormones participate in the control of many aspects of reproduction, development and metabolic stability. Chorionic gonadotropin (CG) is responsible for the maintenance of pregnancy and luteinizing hormone (LH) and follicle stimulating hormone (FSH) regulate gonadal function. Thyroid stimulating hormone (TSH) stimulates the production of thyroid hormones which are involved in development and maintenance of metabolic stability. Each hormone is comprised of two subunits, the common alpha subunit and a unique beta subunit. The alpha subunit is encoded by a single gene which is expressed as a component of all four hormones. Thus, it is produced by the gonadotropes and thyrotropes of the anterior pituitary and by the trophoblasts of the placental. In addition, the alpha-subunit gene is produced ectopically by a variety of human tumors. Since each of these hormones is independently regulated, the alpha-subunit gene must contain regulatory sequences which direct its expression in several specific tissues during different times in development, and facilitate its differential hormonal responsiveness. Using gene transfer experiments, we have demonstrated that the alpha-subunit gene 5' flanking region carries sequences involved in placental- specific and cAMP-responsive expression. These interdependent control elements act as regulated and tissue-specific enhancers. In addition, DNA-binding factors which associate specifically with these elements have been identified. In the proposed studies, the control of alpha-subunit gene transcription will be investigated using integrated molecular and biochemical approaches. The specific DNA-binding proteins which associate with the alpha- subunit gene will be characterized and additional factors will be identified. These will be compared to known transcription factors and their presence in various cell types assessed. Selected factors will be purified and biochemically characterized. In addition, in vitro transcription systems derived from placental tissues or complemented with placental fractions will be used to reconstitute the specific and regulated transcription of the alpha- subunit gene. These will be fractionated to identify factors important in the transcription of this gene and will also be used to investigate the roles of the DNA-binding factors. The juxtaposition of DNA-binding studies, protein purification and reconstitution of activity using in vitro systems will allow us to gain precise and detailed information concerning the nature of the factors which control alpha-subunit gene transcription and transcription in general.
