Elucidating Combinatorial Chromatin States by Co-ImmunoPrecipitation (comb-ChIP)

Chromatin ImmunoPrecipitation followed by massively parallel sequencing (ChIP-Seq) has been instrumental to our current view of chromatin structure and function, and identifies correlating histone marks, which together demarcate biologically-relevant domains. However, as with most genome-wide assays, ChIP-seq is an ensemble measurement that reports on the average occupancy of individual modifications in a population of cells. Consequently, our understanding of the combinatorial nature of chromatin states relies almost exclusively on spatial correlations. Here, we report the development of a novel protocol, called indexed Combinatorial ChIP (comb-ChIP), which has the power to determine the genome-wide co-occurrence of histone marks at single nucleosome resolution. We show that at regions of overlapping ChIP signals, certain combinations of marks (H3K36me3 and H3K79me3) tend to co-occur on the same nucleosome, while other combinations (H3K4me3 and H3K36me3) do not, reflecting differences in the underlying chromatin pathways. We further use comb-ChIP to detect changes in histone mark co-occurrence upon genetic perturbation, illuminating new aspects of the Set2-RPD3S pathway. Overall, comb-ChIP promises to greatly improve our understanding of the structural and functional complexity of chromatin.