Characterization of a HeLa Cell Factor Which Negatively Regulates Transcriptional Activation in Vitro by Transcriptional Enhancer Factor-1 (TEF-1)

doi:10.1074/jbc.270.8.3631

Characterization of a HeLa Cell Factor Which Negatively Regulates Transcriptional Activation in Vitro by Transcriptional Enhancer Factor-1 (TEF-1) (*)

From the Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP, Collège de France, B.P. 163-67404 Illkirch Cedex, France

^¶ To whom correspondence should be addressed. Tel.: 33-88-65-34-40 (or 45); Fax: 33-88-65-32-01.

Abstract

A novel negatively acting factor has been identified and partially purified from HeLa and BJA-B cell extracts by chromatographic fractionation. Addition of this factor to HeLa cell extracts or to a reconstituted HeLa cell transcription system repressed transcriptional activation by a chimeric activator, GAL-TEF-1, containing the activation function of transcriptional enhancer factor-1 (TEF-1). In contrast, this factor did not repress transactivation by the chimeric GAL-VP16 activator. Repression of transactivation by GAL-TEF-1 could be alleviated by the addition of immunopurified HeLa cell TFIID, but not by increased quantities of GAL-TEF-1. These observations suggest that this negatively acting factor represses transactivation by interfering with the function of, or competing for, the TATA-binding protein-associated coactivators which mediate the activity TEF-1.

Footnotes

↵^§ Supported by fellowships from the Association pour la Recherche contre le Cancer and the CNRS. Present address: Biotechnology Research, Ciba Geigy Corp., P. O. Box 12257, Research Triangle Park, NC 27709-2257.
↵* This work was supported in part by grants from the CNRS, the INSERM, the Centre Hospitalier Universitaire Régionale, the Ministère de la Recherche et de la Technologie, the Fondation pour la Recherche Médicale, and the Association pour la Recherche Contre le Cancer. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

TBP

TATA-binding protein

TAF

TBP-associated factor

TEF-1

transcriptional enhancer factor-1

PC

phosphocellulose

ER

estrogen receptor

WCE

whole cell extract

HPLC

high performance liquid chromatography

IP

immunoprecipitation(s)

AdMLP

adenovirus major late promoter.
↵²S. Chaudhary, L. Tora, and I. Davidson, unpublished data.
- Received October 19, 1994.
- Revision received December 15, 1994.