BET domain co-regulators in obesity, inflammation and cancer

Abstract

The bromodomain is a highly conserved motif of 110 amino acids that is bundled into four anti-parallel α-helices and found in proteins that interact with chromatin, such as transcription factors, histone acetylases and nucleosome remodelling complexes. Bromodomain proteins are chromatin 'readers'; they recruit chromatin-regulating enzymes, including 'writers' and 'erasers' of histone modification, to target promoters and to regulate gene expression. Conventional wisdom held that complexes involved in chromatin dynamics are not 'druggable' targets. However, small molecules that inhibit bromodomain and extraterminal (BET) proteins have been described. We examine these developments and discuss the implications for small molecule epigenetic targeting of chromatin networks in cancer.

Figure 1. Structure and relationships among bromodomain-containing proteins

a. The anti-parallel α-helices of the bromodomain bundle are shown in association with the small-molecule inhibitor I-BET and a histone H4 lysine peptide acetylated at position 12 (REF. 97). The BC and ZA loops form the binding pocket for the ε-acetyl-lysine groups of nucleosomal histones in the structure, which the Zhou group first described in detail. b. Relatedness among bromodomain families, as defined by selectivity for JQ1, is measured by differential scanning fluorimetry. The BET proteins BRD2, BRD3 and BRD4 are shown to be closely related, with respect to both the first and second bromodomains, as well as the first bromodomain of BRDT. The second bromodomain of BRDT was not tested (shown in grey). Part a is reproduced, with permission, from REF. © (2010) Macmillan Publishers Ltd. All rights reserved. Part b reproduced, with permission, from REF. © (2010) Macmillan Publishers Ltd. All rights reserved.

Figure 2. Motif alignment of double bromodomain-containing proteins

The dual, tandem bromodomains (BDs) are mutually related and always positioned at the amino terminus, where anchoring to nucleosomal histones is encoded, and the carboxy-terminal end of each polypeptide is available for interaction with chromatin-modifying factors, transcription factors, histone-modification enzymes and other proteins. This recruitment takes place either through poorly understood protein–protein interaction ET domains or through SEED domains rich in acidic, phosphorylatable amino acids that resemble the C-terminal domain of RNA polymerase II. The human, fruitfly and yeast proteins contain putative or verified nuclear localization signals (shown in blue) or ATP binding, kinase catalytic sites (shown by grey triangles). Additional features of the Drosophila melanogaster protein include large insertions (represented by triangles) and stretches of polyglutamine (poly Q), a motif that is frequently associated with transcriptional activation. The C-terminal domain of the long isoform of BRD4 is unstructured and does not contain well-established protein–protein interaction or transcriptional-activation motifs, but is nevertheless partly responsible for functional differences between this isoform of BRD4 and other, shorter BET proteins. The chromosome on which each gene is located is identified.

Figure 3. Small-molecule inhibitors of BET proteins

Recently reported chemical structures and measurements of inhibition constants (IC₅₀) or dissociation constants (K_d) for JQ1 (REF. 79), I-BET (REF. 97), I-BET151 (REF. 92) and other structures that incorporate acetyl-lysine bioisoteres are shown. The parts of each molecule that displace the ε-acetyl-lysine group of the histone are circled in red.

Figure 4. Model for BET protein co-repression of PPARγ-responsive genes

Transcriptional co-repression of specific loci is an active process that requires the recruitment of repressor complexes. In the case of peroxisome proliferator-activated receptor-γ (PPARγ), co-repression is enabled through BRD2 association with RXR, which is known to heterodimerize with PPARγ. Removal of BRD2 by genetic ablation promotes the transcription of adipogenic networks, analogous to thiazolidinedione drug treatment. Small-molecule inhibitors of BET proteins would be expected to produce a similar result.

Figure 5. BET proteins co-regulate transcriptional networks of transcriptional activation and repression

Several functional networks are co-regulated by BET protein interactions. Some interactions involve transcriptional co-repression, such as insulin transcription, peroxisome proliferator-activated receptor-γ (PPARγ)-controlled adipogenic differentiation in adipose tissue^, and GATA1-controlled haematopoietic differentiation^,. Other interactions involve transcriptional co-activation, such as the activation of genes that promote cell cycle progression controlled by MYC^,,, and E2F proteins^,,,,; nuclear factor-κB (NF-κB)-controlled synthesis of pro-inflammatory cytokines^,,,; and cellular genes regulated by P-TEFb. The transcription and replication of latent viruses seem to exploit BET protein capacity for either co-repression or co-activation, depending on the demands of the virus, through P-TEFb^,,,,, human papilloma virus (HPV) E2 protein^– or Kaposi’s sarcoma-associated herpesvirus (KSHV) LANA1 protein^,–. Recent data also implicate BRD4 in the maintenance of higher order chromatin structure. EBV, Epstein–Barr virus.