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. 2016 Nov 23;5:e20148. doi: 10.7554/eLife.20148

Figure 4. Attenuation of replication coupled nucleosome disruption at the MBT.

(A) Predicted nucleosome occupancy is plotted for the selected time points (top) for the 800 bp region flanking a set of early embryonic promoters, centered over the TSS. These timepoints correspond to the initial phases of DNA replication in each cell cycle. Nucleosome profiles for the complete timecourse are provided in Figure 4—figure supplement 1. Promoters are ordered on the y-axis by the average −1 nucleosome position over the entire time course, and the relative nucleosome occupancy is represented by the colorbar at left. The log10 average occupancy signal for each time point is plotted below each heatmap (red). Maximum nucleosome signal at the TSS over the entire timecourse is indicated by the grey line. (B) Mean-normalized relative NFR sizes for time (red) and N:C ratio (blue) dependent loci are plotted for haploid embryos for comparison with diploid embryos (green). Data are plotted as a function of N:C ratio (x-axis). NFRs for time-dependent loci demonstrate minimal closing during metaphase (N:C = 0.25, p<0.01, asterisk) and early S-phase (N:C = 0.5, p<0.01, asterisk) compared with N:C-ratio-dependent loci and N:C ratio matched diploid loci. p-Values indicate frequencies of observing differences between randomly selected loci greater than or equal to that observed for the time- and N:C-ratio-dependent groups (one-tailed permutation test for n = 1×104 trials). Error bars show the 95% confidence interval for differences between the plotted median values. (C) Mean-normalized relative NFR sizes for Zelda-associated (red) and GAF-associated (blue) promoters was plotted for haploid embryos. Plotting and significance testing is as described for panel B. Zelda-associated promoters demonstrate increased NFR stability during early S-phase during both N:C = 0.25 and N:C = 0.5 (asterisks, p<0.01, one-tailed permutation test for n = 1×104 trials). Error bars show the 95% confidence interval for differences between the plotted median values.

DOI: http://dx.doi.org/10.7554/eLife.20148.034

Figure 4.

Figure 4—figure supplement 1. Nucleosome positioning over promoters.

Figure 4—figure supplement 1.

(A) Predicted nucleosome occupancy is plotted for all time points (top) for the 800 bp region flanking a set of early embryonic promoters, centered over the TSS. Promoters are ordered on the y-axis by the average −1 nucleosome position over the entire time course, and the relative nucleosome occupancy is represented by the colorbar at left. The log10 average occupancy signal for each time point is plotted below each heatmap (red). Maximum nucleosome signal at the TSS over the entire timecourse is indicated by the grey line. (B) The average NFR size for each timepoint was calculated and plotted. (C) Average nucleosome occupancies were normalized to the mean occupancies of the flanking 200 bp of each 800 bp region and plotted. Top plot shows average occupancy data for all time points in grey, and the per-cell-cycle average as indicated by the legend. Bottom plot shows average occupancy data for all time points according to the color bar at right.
DOI: http://dx.doi.org/10.7554/eLife.20148.035

Figure 4—figure supplement 2. Changes in accessibility at nucleosome-free regions over the cell cycle.

Figure 4—figure supplement 2.

To calculate changes in chromatin accessibility at NFRs over the cell cycle, the maximum extent of each NFR was calculated from the predicted extreme positions of −1 and +1 nucleosomes as determined by NucleoATAC. For each timepoint, the maximum ATAC-seq count value corresponding to open and accessible chromatin was calculated for each NFR (n = 2470) and plotted. Chromatin accessibility at NFRs doubles on average over the course of each cell cycle (Left Panel). The boxplots show the calculated log2 fold change between the first and last timepoint in each indicated cell cycle for each NFR. A greater negative log2 fold change in accessbility is observed between metaphase 11 and early interphase 12 than between metaphase 12 and interphase 13 (Middle Panel). The dotted grey line indicates no difference in fold change between compared timepoints. To estimate the latency in recovery of accessibility to prior levels of chromatin accessibility, data for each NFR was subjected to spline interpolation and the timepoint when accessibility in the following cell cycle is equal to or greater than accessibility for the preceding metaphase was calculated. The right panel shows the kernel density estimate for the distribution of latency times for recovery during NC12 and NC13 (black and red traces, respectively). The distribution of latency estimates is biphasic for both cell cycles, with NC12 showing nearly equivalent distribution between early (3 min) and late (9 min) recovering regions. A larger proportion of NFRs recover by 3 min during NC13. Median values for raw, non-interpolated data (inset) are +9 min for NC12, and +6 min for NC13.
DOI: http://dx.doi.org/10.7554/eLife.20148.036