DELINEATION OF 3 DIFFERENT THYROID HORMONE-RESPONSE ELEMENTS IN PROMOTER OF RAT SARCOPLASMIC-RETICULUM CA2-ATPASE GENE - DEMONSTRATION THATRETINOID-X RECEPTOR BINDS 5' TO THYROID-HORMONE RECEPTOR IN RESPONSE ELEMENT-1()

Thyroid hormone (3,5,3'-triiodothyronine) positively regulates transcr iption of the sarcoplasmic reticulum Ca(2+)ATPase gene in rat heart, a nd sequences within 559 nucleotides upstream from the transcription st art site confer thyroid hormone responsiveness upon a reporter gene. I n the present study, three thyroid hormone-response elements (TREs) ar e identified between nucleotides -485 and -190. Each TRE is active in transient transfection assays and specifically binds 3,5,3'-triiodothy ronine receptors (TRs) alpha 1 and beta 1 alone and in combination wit h retinoid X receptors (RXRs) alpha and beta. TRE 1 is a direct repeat of two half-sites separated by four nucleotides; TREs 2 and 3 are inv erted palindromes of two half-sites separated by four and six nucleoti des, respectively. Methylation interference analysis of TRE 1 showed b inding of a TR alpha 1 monomer to the 3' half-site, whereas the hetero dimer contacts both half-sites. Subsequent studies employed TR beta an d RXR alpha mutants in which their P-boxes were replaced with the P-bo x of the glucocorticoid receptor. Bandshifts of wild type and mutant p roteins with either wild type TRE 1 or a mutant version, in which the 5' half-site was converted to a glucocorticoid response element half-s ite, demonstrated preferential binding of RXR to the 5' half-site and of TR to the 3' half-site of TRE 1.