Fig. 3: Effect of Notch1 activation on cell death and tumourigenesis.

a Predicted effect of candidate genes, as indicated by their sense fraction of insertions based on the direction of the CAG promoter and the transcriptional direction of the inserted gene. Genes with a strong bias towards sense insertions are expected to be activated and might serve as oncogenic drivers. Conversely, those biased towards antisense insertions are predicted to be inactivated or yield truncated products and serve as tumour suppressors. The dashed line in the middle indicates the equal ratio of sense and antisense insertions. b Structure of Notch1 and transposon insertion sites within the Notch1 gene. Blue arrows indicate that the promoter in the transposon is in the same orientation as the host gene, and red arrows indicate different directions. Insertion frequencies are indicated by numbers. c Expression analysis of all exons of the Notch1 gene based on RNA-seq of Notch1-driven SB tumours. d Activated Notch1 protein expression analysis based on western blotting compared with non-Notch1-driven tumours. e Western blot analysis of Notch1-driven tumours (MK1370-3R) and non-Notch1-driven tumours (MK1097-5R) after shRNA knockdown. f, g Tumour volume measured at day 30 of MK1370-3RMT (f) and MK1097-5RMT (g) after shRNA-mediated knockdown of endogenous Notch1 or pan-Notch1; n = 4 biologically independent animals. The t-test was used to determine the significance of the difference between the different sets of data. h Kaplan–Meier curve showing the mammary tumours-free rate for SB mice with Notch1-driven tumours (n = 93) and non-Notch1-driven tumours (n = 152), as well as BrW (n = 62) and BrM (n = 56) control mice. Notch1-driven tumours tended to show earlier onset compared with non-Notch1-driven tumours (p < 0.0001) by the log-rank test. i Percentage of different types of ES colonies with wild-type Brca1 and knockout after treatment with 4-HT to induce Brca1 knockout in p53 mutant, ICN1-overexpressing and parental ES cell lines. Cells were inoculated with 4-HT for 3 days (to delete Brca1), followed by disassociation and replating on feeder cells with regular medium. Single ES colonies were selected 7 days later for genotyping of their Brca1 status. Data are presented as mean values ± s.d.