Genome-Wide H3K9m2 Profiling Figure 5. H3K9m2 methylation and expression levels of genes. (a) Histogram of H3K9m2 methylation of genes present in euchromatic arms (red line) or present in pericentromeric regions (blue bars). X-axis corresponds to average Z-score of each gene and Y-axis is the percent of the genes falling within each Z-score interval. (b) Expression level of all genes located in pericentromeric regions (green), those that are H3K9m2 methylated (red) or are unmethylated (blue). doi:10.1371/journal.pone.0003156.g005 arms (Figure 7b). We found that a significant fraction of repeats found in pericentromeric heterochromatin showed a peak of very high H3K9m2 levels, which was higher than the levels found in H3K9 methylated repeats in the arms. These results are consistent with the general trend of very high levels of H3K9m2 in pericentromeric regions, and suggest that many types of repeated DNAs become highly H3K9 methylated in these regions. Abundant evidence shows that small interfering RNA (siRNAs) can recruit DNA methylation and gene silencing [50]. We carried out an analysis of H3K9m2 within clusters of endogenous siRNAs defined by massively parallel signature sequencing (MPSS) [51] by determining how many of those clusters fall within H3K9m2 regions. As expected, the majority of siRNA clusters present in pericentromeric regions were highly associated with H3K9m2 (90% or higher overlap), independent of whether the siRNA cluster was defined as dense, moderate or sparse (Figure 8). However, in the euchromatic arms of the chromosomes, sparse clusters behaved differently from the dense/moderate clusters. In particular, while the percent of H3K9m2 dense clusters stayed relatively high (84%), the percent of H3K9m2 methylated sparse clusters decreased to 37%. These results indicate that dense siRNA clusters are strongly associated with H3K9m2, independent of their position in the genome, while sparse siRNA clusters are not as efficient at recruiting H3K9m2 in euchromatic arms. These findings also once again PLoS ONE | www.plosone.org 7 September 2008 | Volume 3 | Issue 9 | e3156