The N-terminal domain of thyroid hormone receptor-alpha is required for its biological activities.
Thyroid hormone (T3) receptors (T3Rs) are ligand-modulated transcription factors that belong to the nuclear receptor superfamily. Whereas the well-conserved DNA-binding domain and the relatively well-conserved ligand-binding domain in T3Rs have been characterized in detail, limited information is available on the contribution of the variable N terminus to the transcriptional properties of T3Rs. To gain greater insight into the function of the N terminus, we generated a deletion mutant of T3Ralpha, T3Ralpha-deltaN1, that lacks amino acids 7-45 and assessed the effect of this deletion on all known transcriptional activities of T3Ralpha. Despite the fact that T3Ralpha-deltaN1 was expressed and bound T3 with an affinity similar to that of wildtype T3Ralpha, all of its common transcriptional activities were lost. That is, T3Ralpha-deltaN1 did not activate transcription from a positive or negative T3 response element, and it could not interfere with AP-1 transcriptional activity. Surprisingly, T3Ralpha-deltaN1 lost its ability to bind DNA, which can account for its deficiencies as a transcriptional activator. In contrast, the ability of T3Ralpha-deltaN1 to interact with putative coactivators or corepressors was not significantly altered from that of wildtype T3Ralpha. However, overall folding of T3Ralpha-deltaN1 was altered, as indicated by differential sensitivity to limited protease digestion. These data document that the N terminus of T3Ralpha, albeit relatively short and representing a variable and unconserved region when compared with other nuclear receptors, has a critical role in proper folding of the DNA-binding domain and is required for the biological activities of full-length T3Ralpha.
Thuestad
G
G
Department of Pharmacy, University of Oslo, Norway.
Kraus
I
I
Apriletti
J
J
Saatcioglu
F
F
D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D010450:Endopeptidases / Q000378:metabolism; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D016296:Mutagenesis; D011485:Protein Binding; D011988:Receptors, Thyroid Hormone / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism; D017384:Sequence Deletion; D015534:Trans-Activators / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection
D004268:DNA-Binding Proteins; D011988:Receptors, Thyroid Hormone; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D015534:Trans-Activators; D004247:DNA; D008156:Luciferases; D015500:Chloramphenicol O-Acetyltransferase; D010450:Endopeptidases