The nucleosome remodeling and deacetylase (NuRD) complex is a widely conserved transcriptional co-regulator that harbors both nucleosome remodeling and histone deacetylase activities. Although the subunit composition of the complex is fairly well characterized, its exact stoichiometry, arrangement of subunits within the complex, and dynamics of these subunit interactions are largely unknown. Given its importance throughout the animal kingdom, knowledge of NuRD structure and function will provide key insight into the mechanisms of eukaryotic gene regulation.
The work presented here, as part of a program of research, aims to elucidate the stoichiometry and interaction map of a subcomplex of the NuRD complex that comprises HDAC1, MTA2 and MBD3 (MMH). The MMH subcomplex was expressed in HEK293 cells, affinity purified and fractionated using sucrose gradient centrifugation. The subunit stoichiometry was then quantified via data-independent acquisition mass spectrometry. Our results reveal a 2:2:2 stoichiometry of the MBD3-MTA2-HDAC1 subcomplex. The architecture of the MMH complex was also mapped using covalent crosslinking combined with mass spectrometry (XL-MS). We obtained 56 high-confidence inter-protein cross-linked peptides and 42 intra-protein peptides that will allow us to understand the way in which the subunits are arranged. These data will help to build a better understanding of the structure of the NuRD complex.