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Structure and VP16 binding of the Mediator Med25 activator interaction domain

Abstract

Eukaryotic transcription is regulated by interactions between gene-specific activators and the coactivator complex Mediator. Here we report the NMR structure of the Mediator subunit Med25 (also called Arc92) activator interaction domain (ACID) and analyze the structural and functional interaction of ACID with the archetypical acidic transcription activator VP16. Unlike other known activator targets, ACID forms a seven-stranded β-barrel framed by three helices. The VP16 subdomains H1 and H2 bind to opposite faces of ACID and cooperate during promoter-dependent activated transcription in a in vitro system. The activator-binding ACID faces are functionally required and conserved among higher eukaryotes. Comparison with published activator structures reveals that the VP16 activation domain uses distinct interaction modes to adapt to unrelated target surfaces and folds that evolved for activator binding.

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Figure 1: Solution structure of Med25 ACID.
Figure 2: VP16–ACID interaction.
Figure 3: VP16-binding interface of ACID.
Figure 4: Functional ACID-VP16 interaction.
Figure 5: Model of an activated transcription initiation complex.
Figure 6: Comparison of ACID with known activator–target complexes.

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Acknowledgements

We thank D. Kostrewa and K. Leike for help, and T. Fröhlich for help with MALDI and ESI experiments. M. Seizl was supported by the Boehringer Ingelheim Fonds and Elitenetzwerk Bayern. A.M. was supported by a Ph.D. fellowship (SFRH/BD/22323/2005) from the Portuguese Foundation for Science and Technology (FCT). M. Sattler acknowledges support by the Deutsche Forschungsgemeinschaft, the European Commission (3D Repertoire LSHG-CT-2005-512028) and the Bavarian NMR Center. P.C. was supported by the Deutsche Forschungsgemeinschaft, the Sonderforschungsbereich SFB646, the SFB Transregio 5, the Jung-Stiftung and the Fonds der chemischen Industrie.

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E.V., A.M. and B.S., NMR data acquisition and analysis; E.V., M. Seizl, L.W., L.L. and S.B., sample preparation and functional assays; K.B. and M.M., mammalian transcription assays; M.M., M. Sattler and P.C., project design and supervision; E.V., M. Sattler and P.C., manuscript preparation.

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Correspondence to Michael Sattler or Patrick Cramer.

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Vojnic, E., Mourão, A., Seizl, M. et al. Structure and VP16 binding of the Mediator Med25 activator interaction domain. Nat Struct Mol Biol 18, 404–409 (2011). https://doi.org/10.1038/nsmb.1997

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