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Published February 23, 2016 | Supplemental Material + Published
Journal Article Open

Structure of the Brd4 ET domain bound to a C-terminal motif from γ-retroviral integrases reveals a conserved mechanism of interaction

Abstract

The bromodomain and extraterminal domain (BET) protein family are promising therapeutic targets for a range of diseases linked to transcriptional activation, cancer, viral latency, and viral integration. Tandem bromodomains selectively tether BET proteins to chromatin by engaging cognate acetylated histone marks, and the extraterminal (ET) domain is the focal point for recruiting a range of cellular and viral proteins. BET proteins guide γ-retroviral integration to transcription start sites and enhancers through bimodal interaction with chromatin and the γ-retroviral integrase (IN). We report the NMR-derived solution structure of the Brd4 ET domain bound to a conserved peptide sequence from the C terminus of murine leukemia virus (MLV) IN. The complex reveals a protein–protein interaction governed by the binding-coupled folding of disordered regions in both interacting partners to form a well-structured intermolecular three-stranded β sheet. In addition, we show that a peptide comprising the ET binding motif (EBM) of MLV IN can disrupt the cognate interaction of Brd4 with NSD3, and that substitutions of Brd4 ET residues essential for binding MLV IN also impair interaction of Brd4 with a number of cellular partners involved in transcriptional regulation and chromatin remodeling. This suggests that γ-retroviruses have evolved the EBM to mimic a cognate interaction motif to achieve effective integration in host chromatin. Collectively, our findings identify key structural features of the ET domain of Brd4 that allow for interactions with both cellular and viral proteins.

Additional Information

© 2016 National Academy of Sciences. Published online before print February 8, 2016. We thank Matthew Plumb, Nikoloz Shkriabai, Annie Moradian, Roxana Eggleston-Rangel, and Michael Sweredoski for help with bioinformatics analysis and proteomics experiments and Punit Uphadyaya, Dehua Pei, Michael Poirier, Aparna Unnikrishnan, Claire Chandler, Chris French, and Geoff Rosenfeld for encouragement and discussions. This work was supported in part by NIH Grant AI062520 (to M.K.) and NIH Grant AI124463 (to M.P.F.). The Proteome Exploration Laboratory (S.H.) is supported by the Gordon and Betty Moore Foundation through Grant GBMF775 and the Beckman Institute. Author contributions: B.L.C., R.C.L., C.Y., M.K., and M.P.F. designed research; B.L.C., R.C.L., C.Y., S.H., and M.P.F. performed research; B.L.C., R.C.L., C.Y., S.H., M.K., and M.P.F. analyzed data; and B.L.C., R.C.L., C.Y., S.H., M.K., and M.P.F. wrote the paper. The authors declare no conflict of interest. This article is a PNAS Direct Submission. Data deposition: NMR, atomic coordinates, chemical shifts, and restraints have been deposited in the Protein Data Bank, www.pdb.org (PDB ID code 2N3K), and BioMagResBank, www.bmrb.wisc.edu (BMRB ID code 25649). This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1516813113/-/DCSupplemental.

Attached Files

Published - PNAS-2016-Crowe-2086-91.pdf

Supplemental Material - pnas.1516813113.sd01.xlsx

Supplemental Material - pnas.201516813SI.pdf

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August 22, 2023
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