The Hbo1-Brd1/Brpf2 complex is responsible for global acetylation of H3K14 and required for fetal liver erythropoiesis

Department of Cellular and Molecular Medicine, Graduate School of Medicine, Graduate School of Pharmaceutical, Sciences, Chiba University, Chiba, Japan.
Blood (Impact Factor: 10.45). 07/2011; 118(9):2443-53. DOI: 10.1182/blood-2011-01-331892
Source: PubMed


The histone acetyltransferases (HATs) of the MYST family include TIP60, HBO1, MOZ/MORF, and MOF and function in multisubunit protein complexes. Bromodomain-containing protein 1 (BRD1), also known as BRPF2, has been considered a subunit of the MOZ/MORF H3 HAT complex based on analogy with BRPF1 and BRPF3. However, its physiologic function remains obscure. Here we show that BRD1 forms a novel HAT complex with HBO1 and regulates erythropoiesis. Brd1-deficient embryos showed severe anemia because of impaired fetal liver erythropoiesis. Biochemical analyses revealed that BRD1 bridges HBO1 and its activator protein, ING4. Genome-wide mapping in erythroblasts demonstrated that BRD1 and HBO1 largely colocalize in the genome and target key developmental regulator genes. Of note, levels of global acetylation of histone H3 at lysine 14 (H3K14) were profoundly decreased in Brd1-deficient erythroblasts and depletion of Hbo1 similarly affected H3K14 acetylation. Impaired erythropoiesis in the absence of Brd1 accompanied reduced expression of key erythroid regulator genes, including Gata1, and was partially restored by forced expression of Gata1. Our findings suggest that the Hbo1-Brd1 complex is the major H3K14 HAT required for transcriptional activation of erythroid developmental regulator genes.

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    • "When BRD1 [39] was first evaluated for FRAP under the same conditions as those in other assays, GFP-tagged BRD1 appeared to be localised exclusively to distinct nuclear speckles, without any diffuse distribution across the nucleus, consistent with reported immunohistological staining results [40]. Attempted FRAP assays with these cells produced almost no recovery after photobleaching (data not shown), indicating that BRD1 is almost entirely immobilised in these speckles within the time frame of the FRAP experiments. "
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    ABSTRACT: Background Acetylation of lysine residues in histone tails plays an important role in the regulation of gene transcription. Bromdomains are the readers of acetylated histone marks, and, consequently, bromodomain-containing proteins have a variety of chromatin-related functions. Moreover, they are increasingly being recognised as important mediators of a wide range of diseases. The first potent and selective bromodomain inhibitors are beginning to be described, but the diverse or unknown functions of bromodomain-containing proteins present challenges to systematically demonstrating cellular efficacy and selectivity for these inhibitors. Here we assess the viability of fluorescence recovery after photobleaching (FRAP) assays as a target agnostic method for the direct visualisation of an on-target effect of bromodomain inhibitors in living cells. Results Mutation of a conserved asparagine crucial for binding to acetylated lysines in the bromodomains of BRD3, BRD4 and TRIM24 all resulted in reduction of FRAP recovery times, indicating loss of or significantly reduced binding to acetylated chromatin, as did the addition of known inhibitors. Significant differences between wild type and bromodomain mutants for ATAD2, BAZ2A, BRD1, BRD7, GCN5L2, SMARCA2 and ZMYND11 required the addition of the histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA) to amplify the binding contribution of the bromodomain. Under these conditions, known inhibitors decreased FRAP recovery times back to mutant control levels. Mutation of the bromodomain did not alter FRAP recovery times for full-length CREBBP, even in the presence of SAHA, indicating that other domains are primarily responsible for anchoring CREBBP to chromatin. However, FRAP assays with multimerised CREBBP bromodomains resulted in a good assay to assess the efficacy of bromodomain inhibitors to this target. The bromodomain and extraterminal protein inhibitor PFI-1 was inactive against other bromodomain targets, demonstrating the specificity of the method. Conclusions Viable FRAP assays were established for 11 representative bromodomain-containing proteins that broadly cover the bromodomain phylogenetic tree. Addition of SAHA can overcome weak binding to chromatin, and the use of tandem bromodomain constructs can eliminate masking effects of other chromatin binding domains. Together, these results demonstrate that FRAP assays offer a potentially pan-bromodomain method for generating cell-based assays, allowing the testing of compounds with respect to cell permeability, on-target efficacy and selectivity.
    Epigenetics & Chromatin 07/2014; 7(1). DOI:10.1186/1756-8935-7-14 · 5.33 Impact Factor
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    • "MommeD42 was identified as an enhancer of variegation and carries a nonsense mutation in exon 11 of Bromodomain containing 1 (Brd1; Figure 3d and Table 1). Brd1 has recently been reported to form a complex with the histone acetyltransferase HBO1 and is required for the transcriptional activation of the erythroid-specific regulator genes in fetal liver [45]. "
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    Genome biology 09/2013; 14(9):R96. DOI:10.1186/gb-2013-14-9-r96 · 10.81 Impact Factor
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    • "On the other hand, PZPM was also conserved across its homological components in yeast-NuA3-like HAT complexes (Fig. S1 and Qin et al., 2011) and we suggest that these PZPMs played a similar role in their own complexes, like the one in BRPF2. Recently , BRPF2 was identified to preferentially form a novel HAT complex with HBO1 (human acetylase binding to ORC1) and ING4 (inhibitor of growth protein 4) and regulate erythropoiesis and the BRPF2-containing HAT complex was probably an important H3K14 HAT required for transcriptional activation of erythroid developmental regulator genes, which further emphasized the physiologic significance of this complex (Mishima et al., 2011). However the DNA binding affinity is limited and unspecific, it is possible that the atypical PHD finger of BRPF2 possesses additional undiscovered biological function which is to be revealed by further work. "
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    ABSTRACT: Plant homeodomain (PHD) finger is found to be a versatile reader that functions in recruiting transcription factors and chromatin modification complexes. Bromodomain- and PHD finger-containing (BRPF) proteins are identified as scaffold component in a couple of histone acetyltransferase (HATs) complexes but the biological function of PHD fingers, composing the motif called PZPM (PHD/Zn-knuckle/PHD Motif), in BRPF proteins is far from being well understood. Here we report the three-dimensional solution structure of the second PHD finger of PZPM in human BRPF2. According to the structure, BRPF2 PHD2 possesses a two-strand β sheet which is different from any other PHD fingers. Functionally, this PHD finger can potentially bind DNA non-specifically with an evolutionarily conserved and positively charged surface. We provide the structural and interaction information of this atypical PHD finger and categorize this BRPF2 PHD2 into a new subset of PHD finger. Moreover our work also shed light on the functional aspect of the PZPM.
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