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
StatePublished - Mar 1 1994
Externally publishedYes

Fingerprint

Retinoid X Receptors
Thyroid Hormone Receptors
Ligands
Transcriptional Activation
CD3 Antigens
Embryonal Carcinoma Stem Cells
Baculoviridae
Triiodothyronine
Thyroid Hormones
B-Lymphocytes
Transcription Factors
In Vitro Techniques
alitretinoin

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Recombinant thyroid hormone receptor and retinoid X receptor stimulate ligand-dependent transcription in vitro. / Lee, Insong J.; Driggers, Paul H.; Medin, Jeffrey A.; Nikodem, Vera M.; Ozato, Keiko.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 91, No. 5, 01.03.1994, p. 1647-1651.

Research output: Contribution to journalArticle

Lee, Insong J. ; Driggers, Paul H. ; Medin, Jeffrey A. ; Nikodem, Vera M. ; Ozato, Keiko. / Recombinant thyroid hormone receptor and retinoid X receptor stimulate ligand-dependent transcription in vitro. In: Proceedings of the National Academy of Sciences of the United States of America. 1994 ; Vol. 91, No. 5. pp. 1647-1651.
@article{b1ef4de83de94026a442df327bc99c4a,
title = "Recombinant thyroid hormone receptor and retinoid X receptor stimulate ligand-dependent transcription in vitro",
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.",
author = "Lee, {Insong J.} and Driggers, {Paul H.} and Medin, {Jeffrey A.} and Nikodem, {Vera M.} and Keiko Ozato",
year = "1994",
month = "3",
day = "1",
language = "English (US)",
volume = "91",
pages = "1647--1651",
journal = "Proceedings of the National Academy of Sciences of the United States of America",
issn = "0027-8424",
number = "5",

}

TY - JOUR

T1 - Recombinant thyroid hormone receptor and retinoid X receptor stimulate ligand-dependent transcription in vitro

AU - Lee, Insong J.

AU - Driggers, Paul H.

AU - Medin, Jeffrey A.

AU - Nikodem, Vera M.

AU - Ozato, Keiko

PY - 1994/3/1

Y1 - 1994/3/1

N2 - 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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=0028194869&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0028194869&partnerID=8YFLogxK

M3 - Article

C2 - 8127860

AN - SCOPUS:0028194869

VL - 91

SP - 1647

EP - 1651

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

SN - 0027-8424

IS - 5

ER -