Cross‐linking mass spectrometry reveals the structural topology of peripheral NuRD subunits relative to the core complex

Spruijt, Cornelia G.; Gräwe, Cathrin; Kleinendorst, Simone C.; Baltissen, Marijke P; Vermeulen, Michiel

doi:10.1111/febs.15650

Cited by 18 publications

(21 citation statements)

References 38 publications

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“…4A-4C). Both copies of MBD3 MBD localize close to the MTA1 BAH domain, which is consistent with the location observed for MBD2 MBD in an independent cryo-EM map of a 2:2:1 MTA1:HDAC1:MBD2 complex (Millard et al, 2020) and an independent set of crosslinks (Spruijt et al, 2021) (Fig. 4A-4E).…”

Section: Resultssupporting

confidence: 86%

“…The quality of the models was assessed by the fit to input data (Fig. S5-S7), as well as data not used in modeling, such as an independent, published crosslinking dataset (Spruijt et al, 2021), cryo-EM maps (Millard et al, 2020), published biochemical data (Desai et al, 2015; Millard et al, 2020; Pflum et al, 2001; Zhang et al, 1999), human cancer-associated mutations (COSMIC) (Table S1) (Forbes et al, 2006), and predictions from AlphaFold (Jumper et al, 2021). The robustness of the models was also assessed by jack-knifing.…”

Section: Resultsmentioning

confidence: 99%

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Molecular architecture of nucleosome remodeling and deacetylase sub-complexes by integrative structure determination

Arvindekar

Jackman

Low

et al. 2021

Preprint

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The Nucleosome Remodeling and Deacetylase (NuRD) complex is a chromatin-modifying assembly that regulates gene expression and DNA damage repair. Despite its importance, limited structural information is available on the complex and a detailed understanding of its mechanism is lacking. We investigated the molecular architecture of three NuRD sub-complexes: MTA1-HDAC1-RBBP4 (MHR), MTA1N-HDAC1-MBD3GATAD2CC (MHM), and MTA1-HDAC1-RBBP4-MBD3-GATAD2 (NuDe) using Bayesian integrative structure determination with IMP (Integrative Modeling Platform), drawing on information from SEC-MALLS, DIA-MS, XLMS, negative stain EM, X-ray crystallography, NMR spectroscopy, secondary structure and homology predictions. The structures were corroborated by independent cryo-EM maps, biochemical assays, and known cancer-associated mutations. Our integrative structure of the 2:2:2 MHM complex shows asymmetric binding of MBD3, whereas our structure of the NuDe complex shows MBD3 localized precisely to a single position distant from the MTA1 dimerization interface. Our models suggest a possible mechanism by which asymmetry is introduced in NuRD, and indicate three previously unrecognized subunit interfaces in NuDe: HDAC1C-MTA1BAH, MTA1BAH-MBD3, and HDAC160-100-MBD3. We observed that a significant number of cancer-associated mutations mapped to protein-protein interfaces in NuDe. Our approach also allows us to localize regions of unknown structure, such as HDAC1C and MBD3IDR, thereby resulting in the most complete structural characterization of these NuRD sub-complexes so far.

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Section: Resultssupporting

confidence: 86%

Section: Resultsmentioning

confidence: 99%

Molecular architecture of nucleosome remodeling and deacetylase sub-complexes by integrative structure determination

Arvindekar

Jackman

Low

et al. 2021

Preprint

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“…Alternatively, MBD2 interactions with methylated DNA could impede the complete assembly of the NuRD complex, resulting in a limited pool of functional complexes. Based on recent crosslinking mass spectrometry and/or structural data and modelling of the complete NuRD complex, MBD proteins have been shown to bridge the HDAC and CHD modules (10)(11)(12). However, if this role, which involves multiple protein-protein interactions with the MBD domain, is compatible with binding to DNA, remains to be fully elucidated.…”

Section: Discussionmentioning

confidence: 99%

“…Additionally, the methyl-CpG binding protein family members MBD2 or MBD3 are essential but mutually exclusive NuRD complex members, therefore assembling distinct MBD2-NuRD or MBD3-NuRD complexes (4,9). Recent structural and biochemical data support the notion that the MBD2 and MBD3 proteins function as a link between the MTA:HDAC:RBBP core and the peripheral GATAD2:CHD:CDK2AP remodeling module (10)(11)(12). Absence of MBD2 or MBD3 therefore disrupts NuRD complex functionality.…”

Section: Introductionmentioning

confidence: 97%

Dissecting the roles of MBD2 isoforms in regulating NuRD complex function during cellular differentiation

Schmolka

Bhaskaran

Karemaker

et al. 2021

Preprint

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The Nucleosome Remodelling and Deacetylation (NuRD) complex is a crucial regulator of cellular differentiation. Two members of the Methyl-CpG-binding domain (MBD) protein family, MBD2 and MBD3, are known to be integral, but mutually exclusive subunits of the NuRD com-plex. Several MBD2 and MBD3 isoforms are present in mammalian cells, resulting in distinct MBD-NuRD complexes. If these different complexes serve distinct biochemical and/or functional activities during differentiation is not completely understood. Based on the essential role of MBD3 in lineage commitment, we systematically investigated a diverse set of MBD3 and MBD2 vari-ants for their potential to rescue the differentiation block observed in mouse embryonic stem cells (ESCs) lacking MBD3. Our study reveals that while MBD3 is indeed crucial for ESC differentiation to neuronal cells, this function is independent of its MBD domain or binding to methylated DNA. While MBD3 isoforms are highly redundant, we identify that MBD2 isoforms vary in their potential to fully rescue the absence of MBD3 during lineage commitment. Full-length MBD2a only partially rescues the differentiation block; MBD2b, which lacks the N-terminal GR-rich repeat, fully rescues the differentiation block in MBD3 KO ES cells, and cells expressing the testis-specific isoform MBD2t that lacks the coiled-coil domain required for NuRD interac-tions are not able to generate any differentiated cells. In case of MBD2a, we further show that removing the m-CpG DNA binding capacity or the GR-rich repeat renders the protein fully redundant to MBD3, highlighting the requirements for these domains in diversifying NuRD complex function. In sum, our results highlight a partial redundancy of MBD2 and MBD3 during cellular differentiation and point to specific functions of distinct MBD2 isoforms and specific domains within the NuRD complex.

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“…B) Normalised protein expression of HDAC1and HDAC2, the normalised iBAQ value is reported in ProteomicsDB[37].Supplemental Table 2. Number of unique peptides detected for each subunit of the NCOR complex and their representation in the Contaminant Repository for Affinity Purification MassSpectrometry Data (CRAPome)[43].…”

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confidence: 99%

Unique Protein Interaction Networks Define The Chromatin Remodeling Module of The NuRD Complex

Tabar

Giardina

Feng

et al. 2021

Preprint

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The combination of four proteins and their paralogues including MBD2/3, GATAD2A/B, CDK2AP1, and CHD3/4/5, which we refer to as the MGCC module, form the chromatin remodeling module of the Nucleosome Remodeling and Deacetylase (NuRD) complex, a gene repressor complex. Specific paralogues of the MGCC subunits such as MBD2 and CHD4 are amongst the key repressors of adult-stage fetal globin and provide important targets for molecular therapies in beta (β)-thalassemia. However, mechanisms by which the MGCC module acquires paralogue-specific function and specificity have not been addressed to date. Understanding the protein-protein interaction (PPI) network of the MGCC subunits is essential in defining underlying mechanisms and developing treatment strategies. Therefore, using pulldown followed by mass spectrometry analysis (PD-MS) we report a proteome-wide interaction network of the MGCC module in a paralogue-specific manner. Our data also demonstrate that the disordered C-terminal region of CHD3/4/5 is a gateway to incorporate remodeling activity into both the ChAHP (CHD4, ADNP, HP1γ) and NuRD complexes in a mutually exclusive manner. We define a short aggregation prone region (APR) within the C-terminal segment of GATAD2B that is essential for the interaction of CHD4 and CDK2AP1 with the NuRD complex. Finally, we also report an association of CDK2AP1 with the Nuclear Receptor Co-Repressor (NCOR) complex. Overall, this study provides insight into the possible mechanisms through which the MGCC module can achieve specificity and diverse biological functions.

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Cross‐linking mass spectrometry reveals the structural topology of peripheral NuRD subunits relative to the core complex

Cited by 18 publications

References 38 publications

Molecular architecture of nucleosome remodeling and deacetylase sub-complexes by integrative structure determination

Molecular architecture of nucleosome remodeling and deacetylase sub-complexes by integrative structure determination

Dissecting the roles of MBD2 isoforms in regulating NuRD complex function during cellular differentiation

Unique Protein Interaction Networks Define The Chromatin Remodeling Module of The NuRD Complex

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