Fig. 1

Concurrent chemical cleavages of H3 histones eliminate the background noise in the chemical mapping experiments. a Integrative Genomics Viewer browser snapshot illustrating nucleosome maps obtained by different techniques. Previous chemical maps, using mutated histones H4S47C (tracks 1–5), are affected by increased background of non-specific cleavages produced by free phenanthroline. Tracks 1 and 2 show the nucleosome occupancy inferred from H4S47C cleavages reported by [23] by symmetrically extending each cleavage position to a footprint of 101 bp (to emphasize the linker positions) between neighboring nucleosomes. Tracks 3–5 show similar data from [26]. The background from the promoter of YBR084W is strongly reduced when we consider only concurrent chemical cleavages, both produced by H3Q85C (tracks 6–8). Tracks 6–8 show the occupancy produced by symmetrically extending the ~ 50-bp fragment centers to a footprint of 101 bp. The absence of histones from the promoter is confirmed by two alternative chromatin immunoprecipitation sequencing (ChIP-seq) methods (tracks 9 and 10) (data from [51, 60]). To make the occupancy profiles comparable, the MNase-ChIP-seq nucleosome map was generated by symmetrically extending the position of the dyad to a footprint of 101 bp. For sonication-ChIP-seq profiles, the occupancy was computed by stacking all the fragments with length between 50 and 200 bp, as in this case the centers of the fragments do not necessarily represent nucleosome dyads. b Average dyad distributions obtained from our H3Q85C cleavage experiment show reduced background noise compared to the map obtained from the H4S47C cleavage experiment. c Length distribution of DNA fragments before and after gel filtration. d Left-right V-plot display of chemical cleavages, where the X-axis shows the position of the center of all 16 aligned centromeres and the Y-axis shows the length of each fragment; red pixels represent the left fragment ends and blue pixels represent the right ends. The two edges of the ~ 80-bp centromere DNA element II (CDEII) are indicated by dotted lines. For H4S47C, the red and blue vertical features between the dotted lines imply that cleavages occur at two distinct positions within CDEII over the population. However, for H3Q85C, no cleavages are seen within the centromeric region, indicating that centromeric nucleosomes do not contain H3 histones. e Cleavage density (left) and average occupancy (right) plots of the data shown in d, comparing H4S47C and H3Q85C cleavage data. Mock control data were obtained by phenanthroline treatment and cleavage reactions performed using wild-type cells as described [26]. f Cleavage density plots comparing H4S47C, H3Q85C, and Mock control cleavage data over aligned transfer RNA (tRNA) genes. g The preferred rotational positions are more evident in the H3S47C cleavage data, even when comparing the nucleosome positions called from the H4S47C cleavage experiment. All dyad distributions were normalized such that the average dyad density equals 1 for every chromosome