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J. Biol. Chem., Vol. 279, Issue 11, 9725-9732, March 12, 2004

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Repression by TTK69 of GAGA-mediated Activation Occurs in the Absence of TTK69 Binding to DNA and Solely Requires the Contribution of the POZ/BTB Domain of TTK69^*

Sara Pagans, David Piñeyro¶, Ana Kosoy||, Jordi Bernués, and Fernando Azorín**

From the Department de Biologia Molecular i Cellular; Institut de Biologia Molecular de Barcelona, Consejo Superior de Investigaciones Científicas, Jordi Girona Salgado, 18-26, 08034 Barcelona, Spain

tramtrack 69 (TTK69) is known to repress GAGA-mediated activation of the eve promoter in S2 cells. Here, we show that repression by TTK69 occurs in the absence of bona fide TTK69-binding sites on the template, indicating that it does not require the binding of TTK69 to DNA. Consistent with this interpretation, the POZ/BTB domain of TTK69, which does not bind DNA, is sufficient for repression. Moreover, a fusion protein in which the POZ/BTB domain of GAGA is replaced by that of TTK69 is not capable of activating the eve promoter but efficiently represses GAGA-dependent activation. Repression involves GAGA-TTK69 interaction because TTK69 is not capable of repressing basal transcription. Most probably, GAGA-TTK69 interaction occurs at the promoter because GAGA·TTK69 complexes are fully competent in binding DNA in vitro. Our results also show that repression by TTK69 of GAGA-dependent activation of the eve promoter is not mediated by any of the co-repressors known to interact with TTK69 (dMi2 or C-terminal binding protein) or by trichostatin A-sensitive histone deacetylases. Altogether, these observations strongly suggest that the binding of TTK69 prevents the interaction of GAGA with the transcription machinery and, therefore, compromises its activation potential.

Received for publication, December 3, 2003

^* This work was financed by grants from the Ministero de Ciencia y Tecnologie (BMC2000-878 and BMC2000-898) and the Commissió Interdepartamental de Recerca i Innovacio Tecnologica (CIRIT) (2001SGR-00344). This work was carried out within the framework of the Centre de Referència en Biotecnologia de la Generalitat de Catalunya. 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.

Present address: Gladstone Institute of Virology and Immunology, University of California, San Francisco, CA 94141.

Supported by a doctoral fellowship from CIRIT.

^¶ Supported by a Formación Profesora de Universitario doctoral fellowship from Ministero de Educación y Ciencia.

^|| Present address: Derald H. Ruttenberg Cancer Center, 1425 Madison Ave., New York, NY 10029.

^** To whom correspondence should be addressed: Dept. de Biologia Molecular i Cellular, Institut de Biologia Molecular de Barcelona, Consejo Superior de Investigaciones Científicas, Jordi Girona Salgado, 18-26, 08034 Barcelona, Spain. Tel.: 3493-4006137; Fax: 3493-2045904; E-mail: fambmc{at}cid.csic.es.

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