File S1 - Suppression of Akt-mTOR Pathway-A Novel Component of Oncogene Induced DNA Damage Response Barrier in Breast Tumorigenesis

Supporting information figures and tables. Figure S1, Cyclin E levels in MCF10A parental (P) or cyclin E over expressing stable cell clones {MCF10A CyE (B10) and MCF10A CyE (C3) (a) measured at RNA level by QPCR using primers that span cyclin E and Flag sequence thus show the levels of transfected flag tagged cyclin E in breast cells. (b) measured by immunofluorescence (IF) by using cyclin E antibody that picked up both the endogenous and transfected cyclin E levels. Figure S2, Cyclin E over expression does not cause cell death through apoptosis. (a) Bar diagram showing the % of cells undergoing early apoptosis, late apoptosis and necrosis in cyclin E over expressing cell clones {MCF10A CyE (B10) and MCF10A CyE (C3) as compared to MCF10A parental (P). 100,000 cells were plated in 6-well plate and next day cells were stained with Annexin V-FITC and PI and were sorted by flow cytometry. The values shown are mean + standard error obtained by repeating the cell cycle assay 3 times. Figure S3, Transient over expression of Cyclin E suppresses AKT-mTOR pathway in MCF10A parental (P) cells. (a) levels of indicated proteins were measured by western blotting. Table S1, P values for pair wise comparisons of the three groups. Table S2, The heterogeneity (e) and 95% CI of the biomarkers. (PDF)