MARCS: Modification Atlas for Regulation by Chromatin States

Goal of this project was the identification of chromatin interacting proteins whose binding is differentially regulated by various combinatorial chromatin modifications found in different chromatin states such as promoters, enhancers and heterochromatin. To achieve this, recombinant modified nucleosomes representing different chromatin states were assembled from canonical histones, H2A.Z and modified ligated H3/H4 histone proteins and biotinylated nucleosomal DNAs. Assembled nucleosomes were immobilized on streptavidin-coated beads via biotinylated di-nucleosomal 601 DNA and used for nucleosome affinity purifications to identify proteins regulated by chromatin modifications from SILAC-labelled HeLaS3 nuclear extracts (Arg10 and Lys8). SILAC affinity purifications were carried out in "forward" (heavy extract on modified nucleosome and light extract on unmodified nucleosome) and "reverse" (light extract on modified nucleosome and heavy extract on unmodified nucleosome) label-swap experiments and protein abundances were quantified by MaxQuant. Initial trial experiments with biotinylated mono-, di- and tetra- 601 nucleosomes are also deposited along with the data for the di-nucleosome experiments. See also: Bartke et al., 2010. Nucleosome-interacting proteins regulated by DNA and histone methylation. Cell 143, 470–484; doi:10.1016/j.cell.2010.10.012. The H4K20me2 samples from this experiment were previously deposited with identifier PXD009281. These were published in: Nakamura et al., 2019. H4K20me0 recognition by BRCA1-BARD1 directs homologous recombination to sister chromatids. Nature Cell Biology 21, 311–318; doi: 10.1038/s41556-019-0282-9.