NSL proteins preferably associate with the promoters of constitutively active genes.

<p>(A) Metagene profiles of histone modifications reveal higher ratios of active chromatin marks H3K4me2/3, H4K16ac and H3K9ac for active genes bound by the NSL complex compared to active NSL-non-bound and inactive genes. On the contrary, the repressive mark H3K27me2 is not enriched on gene promoters bound by the NSL complex. Active genes were defined according to the expression data from <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen.1002736-Cherbas1" target="_blank">[28]</a> (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#s3" target="_blank">Materials and Methods</a>). The expression levels of NSL-bound and NSL-non-bound active genes are similar (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen.1002736.s002" target="_blank">Figure S2A</a>). The log₂ ratios ( = log₂FC (ChIP/input)) of the histone modifications were obtained from modENCODE, extracted for 200 bp bins, and normalized to H4 Chip-chip signals. (B) The chromatin color model contains <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen.1002736-Filion1" target="_blank">[29]</a> two states of euchromatin: “yellow” and “red”. NSL-bound TSSs are predominantly associated with “yellow”, but not “red” chromatin. NSL-non-bound genes display chromatin color ratios that resemble the pattern seen by Filion et al. for tissue-specific genes. (“Green” and “blue” correspond to classical and PcG heterochromatin, respectively, while “black” denotes regions of repressive chromatin). (C) For a different model of chromatin states devised by Kharchenko et al., similar results as in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen-1002736-g002" target="_blank">Figure 2B</a> were obtained: The pie chart depicts that 93% of all peaks of NSL1, MCRS2, NSL3 and MBD-R2 that localize within +/−200 bp of the nearest TSS associate with regions of chromatin state 1. This is defined as the state of actively transcribed TSSs <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002736#pgen.1002736-Kharchenko1" target="_blank">[30]</a>. Complementary, as shown in the bar chart, NSL-bound TSSs of expressed genes are significantly enriched in chromatin state 1 and depleted of chromatin state 9 (p-values<2.2e-16; binomial test) while NSL-non-bound genes are more equally distributed between states of active TSSs (1) and elongation marks (states 2, 3, 4). (D) Heatmaps of ChIP-seq signals (log₂FCs) demonstrate the strong enrichment of NSL binding around the TSSs of constitutively transcribed genes. In contrast to the Pol II signal that is present in both constitutive and regulatory (not constitutive) active genes, the NSL proteins are predominantly found around the TSSs of constitutively transcribed genes. As indicated on the left-hand side, genes were sorted according to their genomic location. The proteins' binding intensities can be directly compared between the different panels. The inlay (right) illustrates the findings of the heatmap with a focus on genes that are bound by all NSLs concomitantly: 85.5% of NSL-bound promoters are constitutively expressed (light blue area). Active (not constitutive) and inactive genes are represented by dark blue and white areas, respectively.