Estradiol repression of tumor necrosis factor-α transcription requires estrogen receptor activation function-2 and is enhanced by coactivators

Estradiol repression of tumor necrosis factor-α transcription requires estrogen receptor activation function-2 and is enhanced by coactivators

^*Department of Obstetrics, Gynecology and Reproductive Sciences, Center for Reproductive Sciences, and ^†Metabolic Research Unit, University of California, San Francisco, 513 Parnassus Avenue, San Francisco, CA 94143; ^‡Department of Pharmaceutical Sciences, University of Brasília, Campus Universitário/Asa Norte, Brasília, DF, 70910–900, Brazil; and ^§Department of Medical Nutrition, Karolinska Institute, NOVUM HS, S-141 86 Huddinge, Sweden

Communicated by Keith R. Yamamoto, University of California, San Francisco, CA, and approved October 19, 1999 (received for review April 12, 1999)

Abstract

The tumor necrosis factor-α (TNF-α) promoter was used to explore the molecular mechanisms of estradiol (E₂)-dependent repression of gene transcription. E₂ inhibited basal activity and abolished TNF-α activation of the TNF-α promoter. The E₂-inhibitory element was mapped to the −125 to −82 region of the TNF-α promoter, known as the TNF-responsive element (TNF-RE). An AP-1-like site in the TNF-RE is essential for repression activity. Estrogen receptor (ER) β is more potent than ERα at repressing the −1044 TNF-α promoter and the TNF-RE upstream of the herpes simplex virus thymidine kinase promoter, but weaker at activating transcription through an estrogen response element. The activation function-2 (AF-2) surface in the ligand-binding domain is required for repression, because anti-estrogens and AF-2 mutations impair repression. The requirement of the AF-2 surface for repression is probably due to its capacity to recruit p160 coactivators or related coregulators, because overexpressing the coactivator glucocorticoid receptor interacting protein-1 enhances repression, whereas a glucocorticoid receptor interacting protein-1 mutant unable to interact with the AF-2 surface is ineffective. Furthermore, receptor interacting protein 140 prevents repression by ERβ, probably by interacting with the AF-2 surface and blocking the binding of endogenous coactivators. These studies demonstrate that E₂-mediated repression requires the AF-2 surface and the participation of coactivators or other coregulatory proteins.

Footnotes

↵ ¶ To whom reprint requests should be addressed. E-mail: leitman{at}itsa.ucsf.edu.
Abbreviations:

AF,

activation function;

DBD,

DNA-binding domain;

E2,

estradiol;

ER,

estrogen receptor;

ERE,

estrogen response element;

GRIP,

glucocorticoid receptor-interacting protein;

LBD,

ligand-binding domain;

Luc,

luciferase;

NR,

nuclear receptor;

RIP140,

receptor interacting protein 140;

TNF-α,

tumor necrosis factor-α;

TNF-RE,

TNF-responsive element;

TK,

thymidine kinase;

ICI,

Imperial Chemical Industries