Dr. Gurr wants to determine, in molecular terms, how the biosynthesis of the alpha and beta subunits of thyrotropin (TSH) is regulated by thyroid hormone (T3). He previously demonstrated that T3 inhibits the transcription of alpha and beta TSH genes, and the inhibition of TSH beta gene transcription was more marked than that of alpha. He now wants to identify cis-acting DNA sequence elements and trans-acting factors which modulate the response of alpha and beta TSH to T3 and which determine tissue-specific TSH alpha and beta gene activity. The binding sites for the T3 receptor on the alpha and beta genes will be delineated by DNAase footprinting and methylation protection, using T3 receptor protein which has been synthesized in vitro from T3 receptor cDNA. Other specific trans-acting factors interacting with these genes will be identified in nuclear extracts from thyrotropic tumors. Genomic footprinting will be used to assess the relevance of the in vitro binding data to the situation in intact chromatin. Cis-acting sequence elements and trans-acting factors which are involved in determining tissue specific alpha and beta TSH gene expression will be identified using in vitro transcription, with nuclear extracts from thyrotropic tumors and tissues in which the alpha and beta TSH genes are not expressed, e.g., liver and kidney. These experiments are important since they will demonstrate the mechanism by which hormones are able to regulate gene expression within specific tissues. While many investigators are working on this general problem, Dr. Gurr is studying the negative regulation of a hormone which is composed of 2 subunits, one common to other pituitary hormones and one that is unique, and is an example of feedback inhibition of a substance which regulates its production.
