Multiple mechanisms of chicken ovalbumin upstream promoter transcription factor-dependent repression of transactivation by the vitamin D, thyroid hormone, and retinoic acid receptors.

A.J Cooney,X Leng,S.Y Tsai,B.W O'Malley,M.J Tsai

doi:10.1016/s0021-9258(18)53592-4

Abstract

The chicken ovalbumin upstream promoter transcription factor (COUP-TF) is a member of the steroid/thyroid hormone receptor superfamily about which little is known of its functional role in the cell. However, it is able to repress hormonal induction of target genes by vitamin D3 receptor (VDR), thyroid hormone receptor (TR), and retinoic acid receptor (RAR). We have shown previously that COUP-TF can bind a wide variety of A/GGGTCA repeats. This promiscuous recognition of response elements correlates with the ability of COUP-TF I to repress other receptors that bind to A/GGGTCA repeats with different spacings between the half-sites. Here we show that repression of transactivation by these receptors is a general phenomenon for the COUP-TF subfamily, as inhibition is also observed with COUP-TF II. This repression is also dose-dependent on COUP-TF. Inhibition of VDR, TR, and RAR activities also occurs through natural physiological response elements found in the osteocalcin, myosin heavy chain, and beta RAR promoters, respectively. In search of the mechanisms of repression by COUP-TF we show that it does not involve the formation of detectable functionally inactive heterodimers between COUP-TF and VDR, TR, and RAR. Instead, we show that the mechanism of repression could occur at three different levels: (a) active silencing of transcription and dual competition for; (b) occupancy of DNA binding sites; and (c), heterodimer formation with retinoid X receptor, the coregulator of VDR, TR, and RAR. The silencing activity was localized to the putative ligand binding domain of COUP-TF. We postulate that COUP-TF may play a master role in regulating transactivation by VDR, TR, and RAR.

Highlights

Of gene expression is governed bythe combinationof response little is known of its functional role in the cell
Except for retinoid Xreceptor (RXR),’ no ligands for search of the mechanisms of repression by COUP-TF these transcription factorshave yet been identified, and they we show that it doesnotinvolve theformation of have been grouped as orphan members of the steroid/
We show that themechanism of repression could occur at three different levels: (a)active silencing of transcription and dualcompetition for; ( b )occupancy of DNA bindthyroid hormone receptor superfamily

Summary

To whom correspondence should be sent

Dept. of Cell Biology, Baylor Collegeof Medicine, 1Baylor Plaza, Houston,TX 77030. Tel.: 713-798-6253;Fax: 713-790-1275. This rule was extended to cover the direct repeat spacing of one nucleotide when it was shownthat RXRspecifically activates transcription in a hormone-dependent manner through such an element [17,18,19] This simplification of the response element discrimination by this group of receptors was recently complicatedwhen the genes encodingthecoregulators or auxiliary factors of VDR, TR, and RAR were cloned and identified as RXRs [20,21,22,23,24,25]. The resin was washed twice each with suspension buffer and transcription buffer (20 mM Hepes, levels: by competition for DNA binding site occupancy; the pH 7.9, 20% glycerol, 0.1 M KC1,0.2 mM EDTA, 5 mM MgCl,, 2 mM formation of nonfunctional heterodimers between COUP-TF dithiothreitol). A30-pl aliquot of 2 X SDS loadingbuffer

EXPERIMENTAL PROCEDURES

RESULTS

Competition for the Binding Site

DISCUSSION