FIGURE 2.

Assay for transposase-accessible chromatin using sequencing (ATAC-seq) analysis on fluorescence-activated cell sorting (FACS)-based sorted tdTomato^Low (Tom^Low) and tdTomato^High (Tom^High) populations. a) Timeline of tamoxifen treatment of Sftpc^CreERT2/+;tdTomato^flox/flox mice (n=3). ATAC-seq was carried out on FACS-based sorted Tom^Low and Tom^High subpopulations. d: day. b, c) Coverage heat maps of Tom^Low and Tom^High, displaying genome-wide regions of differential open chromatin peaks in Tom^Low versus Tom^High. Tom^Low chromatin is less open and transcriptionally less active compared to Tom^High. Ctrl-low shows the peaks related to open chromatin regions in Tom^Low. Ctrl-high shows the peaks related to open chromatin regions in Tom^High. Common shows peaks detected in both Tom^Low and Tom^High subpopulations. d) ATAC-seq analysis of peaks based on the cut-offs shows 3605 upregulated in Tom^High (false discovery rate (FDR)<0.05, log₂(FC)>0.585, base mean>20), 3512 upregulated in Tom^Low (FDR<0.05, log₂(FC)>0.585, base mean>20) and 3878 non-regulated (base mean>20, FDR>0.5, log₂(FC) between −0.15 and 0.15), which means 32% and 32.2% of the genome is differently accessible in Tom^Low and Tom^High, respectively. e) ATAC-seq histogram of average read coverage at the Rosa26 locus shows distinct ATAC-seq peaks at the promoter and denser chromatin in Tom^Low compared to Tom^High in this locus. Representative peaks of Tom^Low and Tom^High are the average of three independent samples. f) Quantification of peaks at the Rosa26 locus. g) ATAC-seq profile at Sftpc locus shows distinct ATAC-seq peaks at the promoter and denser chromatin in Tom^Low compared to Tom^High. Representative peaks of Tom^Low and Tom^High are averages of three independent samples. h) Quantification of peak regions of Sftpc locus. The ATAC-seq data have been normalised for sequencing depth and the scale on the y-axis was chosen for optimal visualisation of peaks. i) ATAC-Heat-pVAI Z-score, Pearson, average heat map based on the ATAC-seq data of top cell cycle-related genes differentially regulated in Tom^Low and Tom^High. FDR: the significance of results by Benjamini–Hochberg correction of multiple tests (n=3).