Fig. 5: mtDNA depletion delays cell-cycle progression and relaxed replication.
a, UMAP clustering based on RNA-seq before regression of cell-cycle heterogeneity. Cells are colored by cell-cycle phase. b, Proportion of cells in each perturbation group assigned to each cell-cycle stage. Two-sided chi-squared tests with multiple testing correction (Benjamini–Hochberg) were used to test for significant differences. c, Cell-cycle phase duration times in FUCCI-expressing WT and TFAM-KO HeLa cells. Phase duration was calculated across one full cycle (G1 and S/G2) for 20 cells per condition. Comparisons were conducted using two-tailed unpaired t-tests. **P < 0.01 and ****P < 0.0001. Data are presented as the mean ± s.d. d, Cell-cycle phase duration in ΔH2.1 mtDNA deletion cybrid clones stably expressing PIP–FUCCI, cultured in high-glucose or low-glucose medium and imaged every 20 min over a period of 72 h. Phase duration was calculated across one full cycle (G1, S and G2/M) for 20 cells (early, first 24 h; late, after 24 h). Data are presented as the mean ± s.d. One-way ANOVAs were performed per cell-cycle phase for each clone with Tukey’s post hoc test. ****P < 0.0001. All significant pairwise comparisons are shown. N/A, no observed cells progressed to this phase during the experiment. e, mtDNA coverage across Seurat cell-cycle phases. A one-way ANOVA with Tukey’s post hoc test was used to test for significant differences. ****P < 0.0001. f, mtDNA coverage across cell-cycle pseudotime with cells ranked on the basis of cell cycle pseudotime (tricycle π) score. The fit line is a smoothed locally estimated scatterplot smoothing regression (span = 0.2). Shaded regions around the fit line indicate the 95% CI. g, mtDNA CN in WT PIP–FUCCI-HeLa cells flow-sorted by cell-cycle phase (sorting strategy in Extended Data Fig. 9e). Cell numbers per group are indicated on the graph. Data are presented as the mean ± s.d. A one-way ANOVA with Tukey’s post hoc test was used to test for significant differences. ***P < 0.001.
