L3MBTL2 and E2F6 contribute to chromatin binding of PRC1.6.

<p>(A) Western blot analysis of L3MBTL2, MGA, E2F6, PCGF6 and RING2 in L3MBTL2<i>ko</i>, E2F6<i>ko</i> and PCGF6<i>ko</i> cells. Re-probing for Tubulin (TUB) controlled loading of extracts. The Tubulin blots are related to the MGA blots. Uncropped versions of the MGA blots are shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1007193#pgen.1007193.s004" target="_blank">S4 Fig</a>. (B) Representative genome browser screenshots of a 0.5 Mb region of chromosome 19 showing reduced binding of PRC1.6 components to several promoters in L3MBTL2<i>ko</i> and in E2F6<i>ko</i> cells but not in PCGF6<i>ko</i> cells. (C) Heat map views of the distribution of MGA, L3MBTL2 and E2F6 peaks in wild type cells (n = 8342) and in MGA<i>ko</i>, L3MBTL2<i>ko</i>, E2F6<i>ko</i> and PCGF6<i>ko</i> cells at +/- 2 kb regions centred over the MGA peaks. (D) Scatter plots comparing the signal intensity of MGA, L3MBTL2 and E2F6 peaks in wild type cells with the signal intensity of corresponding peaks in L3MBTL2<i>ko</i> (left panels), E2F6<i>ko</i> (middle panels) or PCGF6<i>ko</i> cells (right panels). Normalized ChIP-seq read counts in MGA ChIP-seq peak regions of wild type cells were plotted against the normalized read counts in corresponding peak regions of L3MBTL2<i>ko</i>, E2F6<i>ko</i> or PCGF6<i>ko</i> cells. (E) Left panel, scatter plot showing the correlation between reduced MGA binding and reduced L3MBTL2 binding in E2F6<i>ko</i> cells. Right panel, scatter plot showing the correlation between reduced MGA binding and reduced E2F6 binding in L3MBTL<i>ko</i> cells. The top 500 ranked MGA binding sites were used to calculate the fold change of normalized ChIP-seq read counts in L3MBTL2<i>ko</i> and E2F6<i>ko</i> cells relative to wild type cells.