Fig. 6: BRCA1 silencing reduces luminal properties in a PLK1-dependent manner.
From: Pathogenic BRCA1 variants disrupt PLK1-regulation of mitotic spindle orientation
a Colony structures derived from 100 MCF10A RFP-TUBA1B cells that were transduced either with non-hairpin shRNA (shNHP, hereafter control-transduced) or with BRCA1-targeting shRNA (shBRCA1, hereafter BRCA1-silenced) and classified at day 5 by the density of nuclei in the colony, as outlined in Supplementary Figure S14 (mean ± SD from four wells across duplicate experiments). ***P = 0.0005; two-tailed unpaired t-test. b Day 5 colony structures derived from 100 BRCA1-silenced (shBRCA1) or 100 control-transduced (shNHP) MCF10A RFP-TUBA1B cells grown in the presence of DMSO or BI2536 (mean ± SD from four wells across duplicate experiments). ****P = 1.09E−04, ns P = 0.5425; one-way ANOVA. c Colony formation assays for control-transduced (shNHP) or BRCA1-silenced (shBRCA1) MCF10A RFP-TUBA1B cells imaged every 15 min for 36 h. 2–4 cell clusters were chosen for imaging at 36 h after seeding. Motion tracking through colony formation is indicated. Scale bar = 50 μm. d Cell number expansion (black y-axis, LHS) and cell distances measured by Delaunay triangulation (red y-axis, RHS) plotted through the colony formation process for control-transduced (shNHP) MCF10A RFP-TUBA1B cells. Panels d–g plot: mean ± SD band plotted except for division angle mean ± SEM plotted, n = 3 wells imaged per experiment, triplicate experiments. e Cell contacts per cell (black y-axis, LHS) and cell division angles relative to the cell’s long axis (red y-axis, RHS) plotted through the colony formation process for control-transduced (shNHP) MCF10A RFP-TUBA1B cells. f Cell number expansion (black y-axis, LHS) and cell distances measured by delaunay triangulation (blue y-axis, RHS) plotted through the colony formation process for BRCA1-silenced (shBRCA1) MCF10A RFP-TUBA1B cells. g Cell contacts per cell (black y-axis, LHS) and cell division angles relative to the cell’s long axis (blue y-axis, RHS) plotted through the colony formation process for BRCA1-silenced (shBRCA1) MCF10A RFP-TUBA1B cells.
