Fig. 3: APH-induced chromosome gaps and breaks and CFS expression do not depend on MiDAS.

A Timeline of experiments where high-dose APH (2 μM) and EdU were added at 1 h, 2 h, or 3 h prior to chromosome harvest to inhibit MiDAS and for visualizing MiDAS foci, respectively. B Comparison of EdU focus yield between untreated and high-dose APH-treated HCT116 and GM12878 cells. Gray bars are the mean of two independent biological replicates (orange and blue sample points). Number of HCT116 metaphase cells scored: 1 hr EdU 104 + 92; 2 hr EdU, 88 + 103; 3 hr EdU, 110 + 96; 1 hr EdU+2uMAPH, 109 + 100; 2 hr EdU+2uMAPH, 63 + 89; 3 hr EdU+2uM APH, 78 + 92. Number of GM12878 metaphase cells: 1 hr EdU, 59 + 102; 2 hr EdU, 59 + 88; 3 hr EdU, 41 + 92; 1 hr EdU+2uMAPH, 21 + 104; 2 hr EdU+2uMAPH, 29 + 83; 3 hr EdU+2uM APH, 21 + 77. C, D Total gaps and breaks (left) and CFS expression (right) in HCT116 and GM12878 cells, respectively, with and without high-dose APH to suppress MiDAS. Chromosome were analyzed in 25 metaphases from each of two biological replicates for each cell line. Gray bars, mean. E FRA3B and FRA116D CFS gaps/breaks (arrows) in a representative HCT116 cell treated with high-dose APH for 2 h from the set of experiments described in C. Scale bar (black line), 5 µm. F Additional examples of FRA3B and FRA16D CFS gaps/breaks (arrows) in HCT116 and GM12878 cells treated with high-dose APH for 2 h or 1 h, respectively, from the set of experiments described in (C) and (D). Scale bar (black line), 5 µm. Source data are provided as a Source Data file.