STRUCTURAL AND FUNCTIONAL-CHARACTERIZATION OF THE GENOMIC LOCUS ENCODING THE MURINE BETA-2 THYROID-HORMONE RECEPTOR

beta 1 and beta 2 are functional thyroid hormone receptors (TRs) that are generated from the same genomic locus by splicing of a different a mino terminus onto a common carboxyl region containing the DNA and hor mone binding domains. TR beta 1 is widely expressed whereas TR beta 2 is found primarily in the pituitary gland although low levels of expre ssion have been described in other tissues. To gain insight into the m echanisms governing expression of this complex transcriptional unit, w e cloned mouse genomic fragments containing the common carboxyl termin us as well as the unique TR beta 2 amino-terminal sequence that was lo cated at least 25 kilobases upstream. The DNA and ligand binding exons are identical in size and location of their boundaries to those of th e human TR beta 1 gene. To determine whether the region 5' of the TR b eta 2 amino terminus represented the promoter region, we examined it f or sites of transcriptional initiation and for its ability to function as a promoter in TR beta 2-expressing thyrotrope cells. Multiple tran scriptional start sites extending over 400 base pairs (bp) were identi fied with those more proximal showing inhibition by TB Transcription w as not detected more than 400 bp upstream from the putative AUG codon, although initiation downstream of this AUG was demonstrated indicatin g alternative AUG usage. A fragment containing 500 bp of the TR beta 2 5'-region exhibited preferential promoter activity when transfected i nto thyrotrope cells that express endogenous TR beta 2. Deletion studi es demonstrated that removal of consensus binding sites for the transc ription factor Pit-1 resulted in loss of this cell specificity. We the refore conclude that the promoter region responsible for expression of the TR beta 2 isoform in pituitary thyrotropes is distinct from that described for TR beta 1 and is located many kilobases upstream from th eir common exons.