Skip to main content
. 2016 Aug 24;7:12595. doi: 10.1038/ncomms12595

Figure 1. Genetic targeting of PML hampers breast cancer initiation potential.

Figure 1

(a) PML levels (representative western blot out of four independent experiments) upon PML silencing with two shRNAs (sh) in MDA-MB-231 cells. (b) Percentage of ALDH1+ cells upon PML silencing with two shRNAs in MDA-MB-231 cells (n=4). (c) Representative flow cytometry analysis out of three independent experiments of the ALDH1+ population in shC or shPML-transduced MDA-MB-231 cells (FITC: fluorescein-isothiocyanate, SSC-A: side-scatter). (d) Effect of PML silencing on primary (OSI) and secondary (OSII) OS formation in MDA-MB-231 cells (n=5 for OSII in shPML cells and n=6 for shC and OSI in shPML cells). (e,f) PML levels (representative western blot out of three independent experiments) (e) and OS formation (n=3) (f) upon PML inducible silencing (shPML#4) with the indicated doses of doxycycline in MDA-MB-231 cells. (g) Limiting dilution experiment after xenotransplantation. Nude mice were inoculated with 500,000 or 50,000 MDA-MB-231 cells (n=12 injections per experimental condition). Tumour-initiating cell number was calculated using the ELDA platform. A log-fraction plot of the limiting dilution model fitted to the data is presented. The slope of the line is the log-active cell fraction (solid lines: mean; dotted lines: 95% confidence interval; circles: values obtained in each cell dilution). (h) PML levels (representative western blot out of four independent experiments) upon PML silencing in the PDX44-derived cell line. (i) OSI formation upon PML silencing in PDX44 cells (n=3). (j) Limiting dilution experiment after xenotransplantation. Nude mice were inoculated either with 100,000 or 10,000 PDX44 cells (n=20 injections per experimental condition). Tumour-initiating cell number was calculated using the ELDA platform as in g. Error bars represent s.e.m., P value (*P<0.05; **P<0.01; ***P<0.001 compared with shC or as indicated). Statistics test: one-tail unpaired t-test (b,d,i), analysis of variance (f) and χ2-test (g,j). dox, doxycycline; OS, oncospheres; shC, Scramble shRNA; sh2, sh4 and sh5, shRNA against PML.