IDENTIFICATION OF A FUNCTIONAL THYROID-HORMONE RESPONSE ELEMENT IN THE UPSTREAM FLANKING REGION OF THE HUMAN NA, K-ATPASE BETA-1 GENE

The human Na,K-ATPase beta1 subunit gene promoter activity is stimulated by thyroid hormone (T3) in the human intestinal Caco-2 cells. To identify potential cis-acting transcriptional regulatory elements involved in this process, chimeric plasmids containing varying lengths of the 5' flanking region of the human beta1 Na,K-ATPase gene linked to the firefly luciferase reporter gene were introduced into Caco-2 cells by transient transfection. Analysis of T3-regulated luciferase activity of cells carrying these plasmids, and subsequent use of site-directed mutagenesis revealed that a region from - 459 to - 438 (relative to the transcriptional start site) is required for the induction of the beta1 Na,K-ATPase gene by T3. An oligonucleotide containing this sequence from - 465 to - 433 confers T3 responsiveness to a heterologous promoter. Gel mobility shift assays showed specific binding of nuclear proteins of Caco-2 cells to this region and immunoreactive T3 receptor was identified in one of these complexes. These data demonstrate that there is a cis-acting thyroid hormone responsive element in the 5' flanking region of the human beta1 Na,K-ATPase gene and induction of transcription of this gene by T3 involves specific binding of the thyroid hormone receptor to the TRE located at position - 459 to - 438.