Recombinant thyroid hormone receptor and retinoid X receptor stimulate ligand-dependent transcription in vitro

Insong J. Lee, Paul H. Driggers, Jeffrey A. Medin, Vera M. Nikodem, Keiko Ozato

Research output: Contribution to journalArticle

Abstract

The thyroid hormone and retinoid X receptors form a heterodimer with each other and mediate thyroid hormone (T3)-dependent transcription. Retinoid X receptor, in addition, forms a homodimer and mediates 9-cis-retinoic acid- dependent transcription. Here, recombinant thyroid hormone receptor and recombinant retinoid X receptor β expressed from baculovirus vectors have been studied for ligand-mediated activation of transcription in vitro. We show that the two recombinant receptors, most likely as a heterodimer, cooperatively enhance transcription in vitro from a template containing functional T3 responsive elements. The enhancement was specific for the T3 responsive element and was greatest when T3 was added to the reaction (≃14- fold increase). Albeit to a lesser degree, the two receptors also directed transcription in the absence of T3. Template competition experiments suggest that the two receptors enhance formation of the preinitiation complex and that activation by T3 occurs when the ligand binds the receptor prior to (or during), but not after, the formation of the preinitiation complex. Although 9-cis-retinoic acid had no effect on the T3-dependent transcription, this ligand activated transcription in vitro directed by recombinant retinoic X receptor β, most likely as a homodimer. This activation was observed when using nuclear extracts from embryonal carcinoma cells as a source of basal transcription factors, but not those from B lymphocytes. These results demonstrate that transcriptional activation mediated by T3 and 9-cis-retinoic acid can be reconstituted in vitro with the respective recombinant receptors.

Original languageEnglish (US)
Pages (from-to)1647-1651
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume91
Issue number5
DOIs
StatePublished - Mar 1 1994

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