TEF-1 Transrepression in BeWo Cells Is Mediated through Interactions with the TATA-binding Protein, TBP

doi:10.1074/jbc.271.16.9510

TEF-1 Transrepression in BeWo Cells Is Mediated through Interactions with the TATA-binding Protein, TBP (*)

From the (1)Endocrine Research Unit, Departments of Medicine and
(2)Biochemistry/Molecular Biology, Mayo Clinic, Rochester, Minnesota 55905

^§To whom correspondence should be addressed. Tel.: 507-255-6554; Fax: 507-255-4828.

Abstract

Transcription enhancer factor-1 (TEF-1) has been implicated in transactivating a placental enhancer (CSEn) that regulates human chorionic somatomammotropin (hCS) gene activity. We demonstrated that TEF-1 represses hCS promoter activity in choriocarcinoma (BeWo) cells (Jiang, S. W., and Eberhardt, N. L.(1995) J. Biol. Chem. 270, 13609-13915), suggesting that TEF-1 interacts with basal transcription factors. Here we demonstrate that hTEF-1 overexpression inhibits minimal hCS promoters containing TATA and/or initiator elements, Rous sarcoma virus and thymidine kinase promoters in BeWo cells. Cotransfection of TEF-1 antisense oligonucleotides alleviated exogenous TEF-1-mediated repression and increased basal hCS promoter activity, indicating that endogenous TEF-1 exerts repressor activity. GST-TEF-1 fusion peptides fixed to glutathione-Sepharose beads retained in vitro-generated human TATA-binding protein, hTBP. The TEF-1 proline-rich domain was essential for TBP binding, but polypeptides also containing the zinc finger domain bound TBP with higher apparent affinity. TBP supershifted hTEF-GT-IIC DNA complexes, but TEF-1 inhibited in vitro binding of TBP to the TATA motif. Coexpression of TBP and TEF-1 in BeWo cells alleviated TEF-1-mediated transrepression, indicating that the TBP-TEF-1 interaction is functional in vivo. The data indicate that TEF-1 transrepression is mediated by direct interactions with TBP, possibly by inhibiting preinitiation complex formation.

Footnotes

↵* This work was supported by National Institutes of Health Grant DK41206 (to N. L. E.). 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:

TEF-1

transcription enhancer factor-1

hCS

human chorionic somatomammotropin

CSEn

chorionic somatomammotropin gene enhancer

TBP

TATA-binding protein

GST

glutathione S-transferase

ANOVA

analysis of variance

TK

thymidine kinase

PAGE

polyacrylamide gel electrophoresis

TRβ

thyroid hormone receptor β

RSV

Rous sarcoma virus

AdML

adenovirus major late.
- Received September 26, 1995.
- Revision received January 25, 1996.