CHD4 Is a Peripheral Component of the Nucleosome Remodeling and Deacetylase Complex

Abstract

Chromatin remodeling enzymes act to dynamically regulate gene accessibility. In many cases, these enzymes function as large multicomponent complexes that in general comprise a central ATP-dependent Snf2 family helicase that is decorated with a variable number of regulatory subunits. The nucleosome remodeling and deacetylase (NuRD) complex, which is essential for normal development in higher organisms, is one such macromolecular machine. The NuRD complex comprises ϳ10 subunits, including the histone deacetylases 1 and 2 (HDAC1 and HDAC2), and is defined by the presence of a CHD family remodeling enzyme, most commonly CHD4 (chromodomain helicase DNA-binding protein 4). The existing paradigm holds that CHD4 acts as the central hub upon which the complex is built. We show here that this paradigm does not, in fact, hold and that CHD4 is a peripheral component of the NuRD complex. A complex lacking CHD4 that has HDAC activity can exist as a stable species. The addition of recombinant CHD4 to this nucleosome deacetylase complex reconstitutes a NuRD complex with nucleosome remodeling activity. These data contribute to our understanding of the architecture of the NuRD complex.Nucleosomes effectively act as a roadblock to all aspects of genome biology. ATP-dependent chromatin remodeling enzymes solve this problem by using ATP-derived energy to alter the positions, occupancy and composition of nucleosomes. All remodelers possess a highly related ATPase motor domain from the helicase family and are classified into four subfamilies (INO80, ISWI, SWR1, and CHD) based on sequence similarity (1). Each subfamily is represented in nearly all eukaryotes, suggesting that they catalyze different remodeling events. For example, ISWI proteins reposition (or slide) nucleosomes to create regularly spaced arrays; this periodic organization is a key characteristic of DNA at the start of genes (2). SWR1 and INO80 enzymes have opposing roles in histone variant dynamics; the former incorporates these histone variants (e.g. H2A.Z), whereas the latter removes them. These variants set up specific chromatin structures that modulate transcription and replication, although the roles of many variants are still under debate (3). Fundamentally, these remodeling enzymes all alter the accessibility of DNA to other DNA-binding factors and thereby broadly underpin genome biology.Remodelers frequently act in the context of large multisubunit complexes, and in general, the "mixing and matching" of complex composition can generate complexes with varying activities; the human ISWI protein Snf2h for instance has been identified in six distinct complexes (4). Likewise, the accessory subunits can also modulate remodeler activity. For example, the paralogous methyl-CpG-binding domain proteins 2 and 3 (MBD2 and MBD3) subunits of the nucleosome remodeling and deacetylase (NuRD) 6 complex are mutually exclusive (5); MBD2 recognizes 5-methylcytosine-modified DNA, whereas MBD3 instead binds to 5-hydroxymethylated DNA (6, 7). Unsurprisingly, it has been observed tha...

Refinement of the subunit interaction network within the nucleosome remodelling and deacetylase (NuRD) complex

“…In contrast, however, MALLS and iBAQ data on D. melanogaster NuRD argue for a 2:1 HDAC:MBD ratio and a sub-stoichiometric amount of GATAD2 [41]. A sub-stoichiometric GATAD2 is unlikely; we have previously shown, using blue-native-PAGE (BN-PAGE) and two-dimensional BN-SDS-PAGE that GATAD2 is associated with the NuRD complex in all its forms (including the CHD-less NuDe complex) [32]. This apparent discrepancy might be attributable to biological differences between the mammalian and fly NuRD.…”

“…4A). In contrast, NuRD subunits could be readily detected in HEK293 cell lysates and also in a pulldown from HEK293 cells using FOG1(1-45) (which selectively enriches the NuRD complex from cell extracts [31][32][33]) as bait.…”

The Nucleosome Remodeling and Deacetylase complex has an asymmetric, dynamic, and modular architecture

“…We purified native mammalian NuRD complex from murine erythroleukemia cells using our previously established protocol (Figure 1a, (44)). This strategy also yields a complex lacking the CHD subunit, which we have previously termed the Nucleosome Deacetylase (NuDe) complex.…”

“…Samples of NuDe and NuRD were isolated by FOG1 affinity pulldown from cultured mouse erythroleukemia (MEL) cells as described in (44).…”

The Nucleosome Remodelling and Deacetylation complex suppresses transcriptional noise during lineage commitment

“…Nucleosome Remodelling and Deacetylation is comprised of two enzymatically and biochemically distinct subcomplexes: a chromatin remodelling and a deacetylase subcomplex. The chromatin remodelling subcomplex contains a nucleosome remodelling ATPase protein (Chd3/4/5) along with one of the zinc finger proteins Gatad2a/b and the Doc1/Cdk2ap1 protein, while the deacetylase subcomplex contains class I histone deacetylase proteins Hdac1/2, the histone chaperones Rbbp4/7, the Metastasis Tumour Antigen family of proteins, Mta1, Mta2 and Mta3 and, in pluripotent cells, the zinc finger proteins Sall1/4 (Lauberth & Rauchman, ; Allen et al , ; Kloet et al , ; Bode et al , ; Low et al , ; Miller et al , ; Spruijt et al , ; Zhang et al , ). These two subcomplexes are bridged by Mbd2/3, creating intact NuRD (Fig A).…”