Identification of transcription complexes that contain the double bromodomain protein Brd2 and chromatin remodeling machines

Boston University, Boston, Massachusetts, United States
Journal of Proteome Research (Impact Factor: 5). 04/2006; 5(3):502-11. DOI: 10.1021/pr050430u
Source: PubMed

ABSTRACT We use affinity purification of the double bromodomain protein Brd2 to isolate a multicomponent nuclear complex from cultured cells, and apply mass spectrometry/proteomics methods to identify the participants. We then confirm by immunoblot several transcription co-activators and co-repressors, proteins of the Swi/Snf chromatin remodeling complex, which regulate transcription control of cyclin A. This multiprotein complex is likely to contribute to cell cycle control and play a role in proliferation and cancer.

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Available from: Mark E Mccomb, Aug 01, 2015
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    • "Brd2 likely co-activates MLL target genes in 11q23 mixed lineage leukemias (Guo et al., 2000). Through its bromodomains and carboxyl-terminal domain for association with E2F-containing protein complexes, Brd2 provides a scaffold on chromatin (Denis, 2001b) that recruits histone acetyltransferase and chromatin remodeling activities (Denis et al., 2006) to the cyclin A promoter (Sinha et al., 2005), thereby coupling histone acetylation to transcription (LeRoy et al., 2008). B cellrestricted constitutive expression in mice of Brd2 inappropriately transactivates the cyclin A gene in pre-malignant B cells (Greenwald et al., 2004) to cause a malignancy that is highly similar to human DLBCL (Lenburg et al., 2007). "
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    ABSTRACT: Double bromodomain proteins bind to acetylated lysines in histones, bringing associated histone modification and nucleosome remodeling activity to chromatin. The ability of bromodomain regulators to alter chromatin status and control gene expression has long been appreciated to be important in the development of certain human cancers. However, bromodomain proteins have now been found also to be critical, non-redundant players in diverse, non-malignant phenotypes, directing transcriptional programs that control adipogenesis, energy metabolism and inflammation. The fact that such different processes are functionally linked by the same molecular machinery suggests a common epigenetic basis to understand and interpret the origins of several important co-morbidities, such as asthma or cancer that occurs in obesity, and complex inflammatory diseases like cardiovascular disease, systemic lupus erythematosus, rheumatoid arthritis and insulin resistance that may be built on a common pro-inflammatory foundation.
    Discovery medicine 12/2010; 10(55):489-99. · 3.50 Impact Factor
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    • "It was further shown that Brd2 and Brd3 associate with hyperacetylated chromatin along the entire lengths of transcribed genes and that Brd2-and Brd3-associated chromatin is enriched in H4K5, H4K12, and H3K14 acetylation, confirming earlier experiments using FRET analysis (Kanno et al., 2004). Brd2 has been reported to form complexes with proteins such as E2F, Snf2β, Baf155, HDAC11, CAF1b, NAP1L3, some components of TAFIID, as well as histones (Denis et al., 2006). Brd2 has also been shown to interact with the major latency-associated nuclear antigen (LANA) of Kaposi's sarcoma herpesvirus/ human herpesvirus-8 (KSHV/HHV-8) and, similar to what was seen for Brd4 in the papillomavirus system (You et al., 2004), to tether the viral genome to mitotic chromosomes (An et al., 2005; Mattsson et al., 2002; Platt et al., 1999; Viejo-Borbolla et al., 2005). "
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    ABSTRACT: The BET subfamily of bromodomain-containing genes is characterized by the presence of two bromodomains and a unique ET domain at their carboxyl termini. Here, we show that the founding member of this subfamily, Brd2, is an essential gene by generating a mutant mouse line lacking Brd2 function. Homozygous Brd2 mutants are embryonic lethal, with most Brd2(-/-) embryos dying by embryonic day 11.5. Before death, the homozygous embryos were notably smaller and exhibited abnormalities in the neural tube where the gene is highly expressed. Brd2-deficient embryonic fibroblast cells were observed to proliferate more slowly than controls. Experiments to explore whether placental insufficiency could be a cause of the embryonic lethality showed that injecting diploid mutant embryonic stem cells into tetraploid wild-type blastocysts did not rescue the lethality; that is Brd2-deficient embryos could not be rescued by wild-type extraembryonic tissues. Furthermore, there were enhanced levels of cell death in Brd2-deficient embryos.
    Developmental Dynamics 04/2009; 238(4):908-17. DOI:10.1002/dvdy.21911 · 2.67 Impact Factor
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    • "Histone acetylation is a key regulator of transcription; however, it is still a matter of debate as to whether the regulatory features are influenced by modification at specific lysine sites or it is the cumulative electrostatic consequences of random lysine acetylation. Mounting evidence indicates that large multiprotein chromatin remodeling complexes regulate transcription by localizing to certain chromatin sites through the interaction of bromodomain (BrD) 1 proteins with acetylated nucleosomes [1] [2] [3] [4] [5] [6] [7]. The BrD is a protein motif, approximately 100 amino acids in length, primarily associated with targeting subunits of chromatin remodeling complexes [8]. "
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    ABSTRACT: An acetyl-histone peptide library was used to determine the thermodynamic parameters that define acetylation-dependent bromodomain-histone interactions. Bromodomains interact with histones by binding acetylated lysines. The bromodomain used in this study, BrD3, is derived from the polybromo-1 protein, which is a subunit of the PBAF chromatin remodeling complex. Steady-state fluorescence anisotropy was used to examine the variations in specificity and affinity that drive molecular recognition. Temperature and salt concentration dependence studies demonstrate that the hydrophobic effect is the primary driving force, consistent with lysine acetylation being required for binding. An electrostatic effect was observed in only two complexes where the acetyl-lysine was adjacent to an arginine. The large change in heat capacity determined for the specific complex suggests that the dehydrated BrD3-histone interface forms a tightly bound, high-affinity complex with the target site. These explorations into the thermodynamic driving forces that confer acetylation site-dependent BrD3-histone interactions improve our understanding of how individual bromodomains work in isolation. Furthermore, this work will permit the development of hypotheses regarding how the native Pb1, and the broader class of bromodomain proteins, directs multisubunit chromatin remodeling complexes to specific acetyl-nucleosome sites in vivo.
    Analytical Biochemistry 04/2008; 374(2):304-12. DOI:10.1016/j.ab.2007.12.008 · 2.22 Impact Factor
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