Figure 1. H3K27M correlates with H3K27ac and is excluded from PRC2 targets.
(A) Representative UCSC genome browser views of genomic regions enriched after chromatin immunoprecipitation (ChIP) for histones, histone post-translational modifications and proteins listed on the left of each track in SF8628. Input served as background control. (B) Heat map analysis of the most highly H3.3K27M enriched genomic loci (first 5000 in descending order top to bottom) in SF8628. The center of each vertical lane represents the summit of the peak while the left and right borders represent −5 kb and +5 kb, respectively. All the other antigens occupancy at those regions are plotted accordingly. Input served as background control. Scale bar represents relative intensities. (C) Metaplot showing average signal accumulation (RPM, reads per million) of the top 5000 regions bound by H3.3K27M in SF8628. All the other antigens average occupancy at those regions are plotted accordingly. Each plot is centered on the summit of the average occupancy and extended 5 kb upstream and downstream (−5kb and +5kb, respectively). Input served as background control. (D) Western blot analysis of immunoprecipitated (IP) histone H3K27ac from mononucleosomes containing C-terminal FLAG-tagged wild type H3.3 or mutant H3.3K27M purified from 293T cells (image representative of at least 2 independent experiments. Uncropped images in Supplementary Figure 18). (E) Same as (A). (F) As described in (B) showing 5000 most enriched regions bound by H3K27me3.