Bromodomains in living cells participate in deciphering the histone code Alejandra Loyola and Genevieve Almouzni UMR 218 CNRS Institut Curie, Section de Recherche, 26 rue d’Ulm, 75248 Paris Cedex 05, France The bromodomain, a module of , 110 amino acids, is found in several chromatin-associated proteins, includ- ing histone acetyltransferases and chromatin-remodel- ing factors, and can bind to acetylated lysines. Such post-translational modifications occur mainly in the N-terminal tail of the histone proteins and, in combi- nation with other modifications, are thought to partici- pate in defining a histone code. Recent findings provide a model for how bromodomain-containing proteins participate in the recognition of acetylated histones. Post-translational modifications are important in regulat- ing protein – protein interactions. In the case of histones, defined patterns of modifications, including acetylation, phosphorylation and methylation, can function as a recognition code for the recruitment of different factors [1–3]. This is known as the ‘histone code’ hypothesis (Box 1). However, for this to occur, specific protein domains that recognize these modifications are required. Support for this hypothesis comes, in part, from structural studies revealing that bromodomains are modules that can bind to acetylated lysine [4–8]. This article will illustrate how recent studies in vivo [9] provide further insights that support this hypothesis, focussing on the recognition of acetylated histones. Bromodomains: families and protein structure Bromodomains are found in several chromatin-associated proteins. Their name comes from the Drosophila protein ‘Brahma’, which was the first bromodomain-containing protein to be identified, and the ATPase subunit of the Brahma chromatin-remodeling complex. The proteins can be divided into three families: (i) histone acetyltrans- ferases (HATs), including Gcn5, p300 cAMP-response- element-binding (CREB)-binding protein-associated factor (PCAF) and TATA-binding-protein-associated factor (TAF) II 250 – a subunit of the transcription factor TF II D; (ii) ATP-dependent chromatin-remodeling complexes, including Brahma, Swi2, Snf2 and Brg1; and (iii) the less-characterized BET (bromodomain and ET domain) family, which is a novel class of transcriptional regulators that includes Bdf1, Bdf2, Bdr4 and Brd2. The bromodomain is composed of four left-handed helix bundles (a Z , a A , a B , and a C ), with a long loop (ZA) connecting helices Z and A oriented against the small loop (BC) connecting helices B and C (Figure 1). These loops are organized to form an accessible hydrophobic pocket in which the interaction with the acetylated lysine residue occurs (for review, see Ref. [10]). Bromodomain function Interactions between bromodomains and acetylated his- tones are thought to mediate several effects, including transcriptional activation, memory of the transcription- ally activated chromosomal regions and anti-silencing. The role of the interactions between bromodomains and acetylated histone in transcriptional activation is exem- plified by the viral activation of the gene interferon b. This results in a unique pattern of histone modifications, particularly acetylation, at the promoter of the gene that can be used as a signal to induce the recruitment of the transcriptional machinery [11]. The sequential recruit- ment of two bromodomain-containing proteins is then observed: first, Brg1, which is a component of the SWI – SNF complex and, second, the transcription factor TF II D through TAF II 250. Their recruitment is mediated by interactions with acetylated (Ac) Lys8 on histone H4 (AcK8 H4), and acetylated Lys9 and acetylated Lys14 on histone H3 (AcK9 and AcK14 H3), respectively [11]. In addition, memory of activated states is illustrated in vitro by the interaction of the chromatin-remodeling complex SWI–SNF and the HAT complex SAGA (Spt – Ada – Gcn5 – acetyltransferase) with the promoters, in the absence of activators. These associations occur through the bromodomain-containing proteins Swi2 or Snf2 and Gcn5, respectively, after the promoter has been acetylated by activator-recruited SAGA [12]. Interestingly, only Swi2 or Snf2, and Gcn5 mediate the association, independent of the presence of several other bromo- domain-containing proteins in both complexes. These data highlight the specificity and selectivity within bromodomains. An anti-silencing role has been assigned to the BET family bromodomain-containing protein Bdf1, through its association with acetylated histone H4 [13]. Chromatin immunoprecipitation assays show that Bdf1 can bind to Box 1. The histone code Histones are post-translationally modified (e.g. by acetylation, phosphorylation, methylation and ubiquitination) on specific resi- dues that are located mostly in their N-terminal tail [18]. The histone code hypothesis proposes that defined patterns of modifications, possibly acting in combination, can be recognized by specific factors. In turn, this recognition can induce the recruitment of particular effectors, thereby translating the histone-modification pattern into a particular chromatin state. Corresponding author: Genevieve Almouzni (genevieve.almouzni@curie.fr). Available online 11 May 2004 Update TRENDS in Cell Biology Vol.14 No.6 June 2004 279 www.sciencedirect.com