Nucleosome organization in the Drosophila genome

Abstract

Comparative genomics of nucleosome positions provides a powerful means for understanding how the organization of chromatin and the transcription machinery co-evolve. Here we produce a high-resolution reference map of H2A.Z and bulk nucleosome locations across the genome of the fly Drosophila melanogaster and compare it to that from the yeast Saccharomyces cerevisiae. Like Saccharomyces, Drosophila nucleosomes are organized around active transcription start sites in a canonical -1, nucleosome-free region, +1 arrangement. However, Drosophila does not incorporate H2A.Z into the -1 nucleosome and does not bury its transcriptional start site in the +1 nucleosome. At thousands of genes, RNA polymerase II engages the +1 nucleosome and pauses. How the transcription initiation machinery contends with the +1 nucleosome seems to be fundamentally different across major eukaryotic lines.

Figure 1. H2A.Z nucleosomal organization around the 5 ′ end of Drosophila genes

a, Browser shot of an arbitrary locus (nbs-defl). Bar graph represents the number of “W” or “C” (top and bottom traces, respectively) strand reads mapped to each coordinate. b, Composite distribution of H2A.Z nucleosomes relative to the TSS. Nearby genes were either included (gray) or eliminated (black) from the analysis, and normalized accordingly. The equivalent Saccharomyces profile is shown in green. c, Correlation of the number of H2A.Z nucleosomal sequencing reads (per gene) to mRNA levels in 8–12 hr embryos.

Figure 2. Organization of conserved DNA motifs around TSSs (left) and nucleosomes (right)

a, Distribution of representative conserved DNA motifs from four distribution classes. b, Distribution of all motifs plotted with Treeview. Bin counts from all motifs in Fig. S12 were converted to fold deviations from the regional average (−/+ 1 kb), converted to a log₂ scale, and plotted. Red/black/green denotes above, near, or below average deviations, respectively.

Figure 3. Positioning properties of Drosophila nucleosomes and DNA

a, Composite distribution of WW and SS dinucleotides (as indicated) along the 147 bp axis of nucleosomal DNA (p-value = 0). The equivalent yeast profile is shown in light shading in the background. b, Average correlation of all Drosophila promoter regions to nucleosome positioning sequence (NPS) patterns, comparing an AA/TT and a CC/GG pattern. The distribution of H2A.Z nucleosomes from Fig. 1b is shown as a gray backdrop. The span of the +1 nucleosome is indicated by the horizontal bar.

Figure 4. H2A.Z nucleosomal organization around the 3′ end of Drosophila genes

a, Composite distribution of nucleosomes relative to ORF end points. Also shown is the distribution of transcript termination sites (polyA sites) in red. Nearby genes were either included (gray) or eliminated (black) from the analysis. b, Average correlation of all Drosophila gene terminal regions to AA/TT or CC/GG NPS patterns. The nucleosome profile from panel a is shown as a gray backdrop.

Figure 5. Distribution of Pol II and Pol II-engaged nucleosomes around the 5′ end of genes

a, Genome-wide location of “Paused” Pol II relative to the TSS. ChIP-chip genomic profiling of Pol II was conducted on Drosophila embryos. The black filled plot shows the distribution of Pol II at 1,956 “Paused” Pol II genes (see Methods). The yellow trace shows the distribution of permanganate-reactive thymines (an indicator of pausing) in 50 genes that undergo pausing. The distribution of Pol II at these 50 genes is similar to the bulk profile (Fig. S18a). b, H2A.Z nucleosomal distribution at 1,956 genes that contained “Paused” Pol II. The “Not paused” class represents those where Pol II is either absent or not paused. c, Distribution of Pol II-bound nucleosomes. Pol II ChIP was performed on bulk MNase-digested mono-nucleosomal DNA and hybridized to genome-wide tiling arrays. The three traces include distributions at 1,956 Pol II “Paused” genes, and control distributions either at 788 genes that have “No H2A.Z” within 1 kb of the TSS, or at 8,736 genes of the “Not paused” class.