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J. Biol. Chem., Vol. 281, Issue 21, 14719-14728, May 26, 2006

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Active Transcription of the Human FASL/CD95L/TNFSF6 Promoter Region in T Lymphocytes Involves Chromatin Remodeling

ROLE OF DNA METHYLATION AND PROTEIN ACETYLATION SUGGEST DISTINCT MECHANISMS OF TRANSCRIPTIONAL REPRESSION^*

Rémy Castellano¹, Bérengère Vire¹, Marjorie Pion, Vincent Quivy^¶2, Daniel Olive, Ivan Hirsch, Carine Van Lint^¶3, and Yves Collette⁴

From the INSERM UMR599, Centre de Recherche en Cancérologie de Marseille, Université de la Méditerranée, 13009 Marseille, France, INSERM U372, Unite de Pathogenie des Infections a Lentivirus, Parc Scientifique et Technologique de Luminy, 13009 Marseille, France, and ^¶Université Libre de Bruxelles, Institut de Biologie et de Médecine Moléculaires, Laboratoire de Virologie Moléculaire, 6041 Gosselies, Belgium

Fas ligand (FasL/CD95L/TNFSF6), a member of the tumor necrosis factor family, initiates apoptosis in lymphoid and nonlymphoid tissues by binding to its receptor Fas (CD95/TNFRSF6). Although the transcriptional control of TNFSF6 gene expression is subjected to intense study, the role of its chromatin organization and accessibility to the transcriptional machinery is not known. Here, we determined the chromatin organization of TNFSF6 gene 5' regulatory regions. Using the indirect end-labeling technique, a unique region named HSS1 and encompassing nucleotides –189 to +185 according to the transcriptional start site, was identified throughout a 20-kilobase nucleosomal DNA domain surrounding the promoter. The HSS1 region displayed hypersensitivity to in vivo DNase I digestion in TNFSF6-expressing cells only, including upon T cell activation. Hypersensitivity to micrococcal nuclease digestion and to specific restriction enzyme digestion suggested the precise positioning of two nucleosomes across the transcription start site and minimal promoter region, likely interfering with TNFSF6 active transcription in T lymphocytes. Indeed, HSS1 hypersensitivity to nuclease digestion strictly correlated with TNFSF6 transcription, including in primary and leukemia T cells. HSS1 chromatin remodeling preceded detectable TNFSF6 mRNA accumulation and was blocked by cycloheximide that also prevented TNFSF6 transcription. However, DNA methylation levels of the TNFSF6 HSS1 region did not correlate with transcriptional activation. Induction of global protein acetylation by treatment with histone deacetylase inhibitors was not accompanied by HSS1 chromatin remodeling and/or TNFSF6 transcription. We conclude that chromatin remodeling is a primary event in the activation of TNFSF6 expression in primary and leukemia T cells and that mechanisms independent of protein deacetylation and of DNA methylation of the TNFSF6 promoter region are involved in the repression of TNFSF6 gene expression.

Received for publication, March 14, 2006

^* This work was supported by grants from the INSERM and the Université Libre de Bruxelles ARC Program 04/09-309 and by grants from the Association Nationale de Recherche contre le Syndrome d'Immunodéficience acquise (SIDA), Fonds National de la Recherche Scientifique (Belgium), Télévie-program, the International Brachet Stiftung, Commissariat Général aux Relations Internationnales/INSERM cooperation, région Wallonne-Commission Européenne Fonds Européen de Développement Régional, and Theyskens-Mineur Foundation. 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.

¹ These authors equally contributed to this work and are fellows from Association Nationale de Recherche contre le Syndrome d'Immunodéficience acquise (SIDA).

² Aspirant of the Fonds National de la Recherche.

³ Maître de Recherches of the Fonds National de la Recherche.

⁴ To whom correspondence should be addressed: UMR599 INSERM, Centre de Recherche en Cancérologie de Marseille, 27 Boulevard Lei Roure, 13009 Marseille, France. Tel.: 33-491-75-84-13; Fax: 33-491-26-03-64; E-mail: collette{at}marseille.inserm.fr.