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J. Biol. Chem., Vol. 280, Issue 2, 1199-1208, January 14, 2005

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A Novel Repressive E2F6 Complex Containing the Polycomb Group Protein, EPC1, That Interacts with EZH2 in a Proliferation-specific Manner^*

Claire Attwooll, Sergio Oddi, Peter Cartwright, Elena Prosperini, Karl Agger, Peter Steensgaard, Christian Wagener¶, Claude Sardet||, M. Cristina Moroni, and Kristian Helin**

From the European Institute of Oncology, Department of Experimental Oncology, Via Ripamonti 435, Milan, 20141, Italy, the ^||Institut de Génétique Moléculaire, UMR5535 1919 route de Mende, 34293 Montpellier, France, and the ^**Biotech Research & Innovation Centre, Fruebjergvej 3, 2100 Copenhagen, Denmark

The transcriptional repressor E2F6 has been identified as a component of two distinct polycomb group protein (PcG)-containing complexes, suggesting a mechanism for the recruitment of repressive complexes to target sequences in DNA. Whereas one complex is involved in the repression of classic E2F target genes in G₀, a role for E2F6 within the cell cycle has yet to be defined. We searched for novel E2F6-binding proteins using a yeast two-hybrid screen and identified the PcG protein, EPC1. We showed that, both in vitro and in vivo, E2F6, DP1, and EPC1 form a stable core complex with repressive activity. Furthermore, we identified the proliferation-specific PcG, EZH2, as an EPC1-interacting protein. Using affinity purification, we showed that E2F6, DP1, EPC1, EZH2, and Sin3B co-elute, suggesting the identification of a novel E2F6 complex that exists in vivo in both normal and transformed human cell lines. EZH2 is required for cellular proliferation and consistent with this, EZH2 elutes with the E2F6-EPC1 complex only in proliferating cells. Thus we have identified a novel E2F6-PcG complex (E2F6-EPC1) that interacts with EZH2 and may regulate genes required for cell cycle progression.

Received for publication, November 4, 2004

^* This work was supported by grants from AIRC, The Italian Health Ministry, the Danish Research Ministry, the Danish Medical Research Council, and La Ligue Nationale contre le Cancer. 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.

Supported by Marie Curie fellowships from the European Union.

^¶ Present address: Capsulution Nanoscience AG, Volmerstrasse 7b, D-12489 Berlin, Germany.

To whom correspondence should be addressed. Tel.: 45-3917-9666; Fax: 45-3917-9669; E-mail: kristian.helin{at}bric.dk.

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