Extended Data Fig. 10: Oncogene-induced polyploidization.
From: Multigenerational cell tracking of DNA replication and heritable DNA damage
(a) Western blot analysis of 53BP1-PCNA U-2 OS cells overexpressing HRAS. (b) Western blot analysis of 53BP1-PCNA U-2 OS cells overexpressing Cyclin E1. (c) DNA fibre analysis of 53BP1-PCNA U-2 OS cells overexpressing either HRAS or Cyclin E1. Statistical analysis by one-way ANOVA followed by Tukey’s post hoc test. (d) Analysis of micronuclei induction in 53BP1-PCNA U-2 OS cells overexpressing either HRAS or Cyclin E1 compared to empty vector transfected control cells. Mean ± SD are depicted. Statistical analysis was performed with one-way ANOVA followed by Tukey’s post hoc test. (e) Single cell lineage from empty vector control 53BP1-PCNA U-2 OS cells. PCNA foci, 53BP1 foci and area of the nucleus over 70 h of live imaging are depicted. Sequential staining intensities from individual cells of the lineage and mean intensities from the corresponding reference population are depicted for the markers pRB, γH2AX, p21 and p53. The total DAPI intensity of the nucleus from the corresponding reference population is depicted as well as from the single cells at the end of live imaging. (f) Single cell lineage from 53BP1-PCNA U-2 OS cells overexpressing HRAS. Live+QIBC data is shown as in (e). (g) Single cell lineage from 53BP1-PCNA U-2 OS cells overexpressing Cyclin E1. Live+QIBC data is shown as in (e). (h) Single cell lineage from 53BP1-PCNA cells overexpressing Cyclin E1. Live+QIBC data is shown as in (e). Representative images of the polyploid cells are included as well as their position within the DAPI scatter plot of the G1 and G2 populations. Scale bars, 10 µm. Box plot limits indicate 25th percentile (Q1) and 75th percentile (Q3); boxes represent interquartile range (IQR, Q3-Q1) with medians (solid lines). Whiskers define lower and upper adjacent value; dots show outliers greater than Q3 + 1.5xIQR. For gel source data, see Supplementary Fig. 1.
