Fig. 1: ATR-dependent Pol I inhibition and nucleolar segregation.
From: Treacle controls the nucleolar response to rDNA breaks via TOPBP1 recruitment and ATR activation
a Time-lapse microscopy of U2OS cells expressing low levels of GFP-tagged Treacle, after transfection with I-Ppo1. b Timeline of the morphological changes of the nucleoli after I-Ppo1 transfection. c Morphological changes of the nucleoli after I-Ppo1 transfection in ATMi (KU-55933) and ATRi (VE-821) treated cells. Displayed are maximum intensity projections of confocal z-stacks. NPM stands for nucleophosmin d Quantification of Treacle foci number per nuceolus, nucleolar volume and nucleolar sphericity by segmentation of 3D reconstituted z-stacks. DMSO/0 min n = 262, DMSO/60 min n = 239, DMSO/120 min n = 264, ATMi/0 min n = 174, ATMi/60 min n = 166, ATMi/120 min n = 179, ATRi/0 min n = 208, ATRi/60 min n = 218, ATRi/120 min n = 201 independent cells. Bars represent means, dotted lines within the violin plots represent median and quartiles. e ATM and ATR recruitment to sites of nucleolar DSBs in response to I-Ppo1 transfection. Displayed are maximum intensity projections of confocal z-stacks. NPM stands for nucleophosmin. f Quantification of nucleolar EU incorporation after I-Ppo1 transfection in I-Ppo1 H98A/DMSO (n = 215), DMSO (n = 205), ATMi (KU-55933, n = 206) and ATRi (VE-821, n = 219) treated cells. g Quantification of nucleolar EU incorporation in siCtrl I-Ppo1 H98A and WT (n = 203) and siATR I-Ppo1 H98A and WT (n = 210) treated cells. h Quantification of nucleolar EU incorporation after I-Ppo1 H98A (n = 229) and I-Ppo1 WT transfection in control (DMSO, n = 231), ATRi (VE-821, n = 237), CHK1i/CHK2i (AZD7762, n = 246) and CHK1i (GDC-0575, n = 255) treated cells. f–h Boxes represent the 25–75 percentile range with median and whiskers represent the 5–95 percentile range. Data points outside of this range are shown individually. All scalebars = 10 µm. Source data are provided as a Source Data file.
