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J. Biol. Chem., Vol. 280, Issue 7, 6222-6230, February 18, 2005

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Efficient Binding of NC2·TATA-binding Protein to DNA in the Absence of TATA^*

Siv Gilfillan, Gertraud Stelzer, Elisa Piaia, Markus G. Hofmann, and Michael Meisterernst

From the Gene Expression, Institute of Molecular Immunology, GSF-National Research Center for Environment and Health, Marchionini-Strasse 25, D-81377 Munich, Germany

Negative cofactor 2 (NC2) forms a stable complex with TATA-binding protein (TBP) on promoters. This prevents the assembly of transcription factor (TF) IIA and TFIIB and leads to repression of RNA polymerase II transcription. Here we have revisited the interactions of NC2·TBP with DNA. We show that NC2·TBP complexes exhibit a significantly reduced preference for TATA box sequences compared with TBP and TBP·TFIIA complexes. In chromatin immunoprecipitations, NC2 is found on a variety of human TATA-containing and TATA-less promoters. Substantial amounts of NC2 are present in a complex with TBP in bulk chromatin. A complex of NC2·TBP displays a K_D for DNA of 2 x 10^-9 M for a 35-bp major late promoter oligonucleotide. While preferentially recognizing promoter-bound TBP, NC2 also accelerates TBP binding to promoters and stabilizes TBP·DNA complexes. Our data suggest that NC2 controls TBP binding and maintenance on DNA that is largely independent of a canonical TATA sequence.

Received for publication, June 7, 2004 , and in revised form, November 29, 2004.

^* This work was supported by grants from the Deutsche Forschungsgemeinschaft, the Training and Mobility of Researchers program of the European Commission (HPRN-CT-2002-00261), and the Bundesministerium fur Bildung und Forschung proteomics platform technology program (031U101F and 0313030A) to M. M. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

To whom correspondence should be addressed. Tel.: 49-89-7099-202; Fax: 49-89-7099-537; E-mail: Meisterernst{at}gsf.de.

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