Fig. 6
Replication stress induces mitotic telomere deprotection. a Experimental time course for (b, c). b Example images of cytocentrifuged chromosome spreads from HT1080 6TG cells treated with DMSO or APH and stained with DAPI (blue), γ-H2AX immunofluorescence (red) and telomere FISH (green). c Quantitation of mitotic telomeric and genomic DDR foci in APH or HU treated HT1080 6TG cells (three biological replicates scoring n ≥ 81 chromosome spreads per condition are compiled in a Tukey box plot, Mann–Whitney test). d Western blot of whole cell extracts from HT1080 6TG parental and HT1080 6TG mCherry-TRF1OE cells. e Representative images of cytocentrifuged chromosomes from HT1080 6TG mCherry-TRF1OE cells treated with DMSO or APH, stained with DAPI (blue) and γ-H2AX IF (green). f Quantitation of mitotic telomeric DDR foci in HT1080 6TG parental and HT1080 6TG mCherry-TRF1OE cells (two biological replicates scoring n ≥ 48 chromosome spreads per condition are compiled in a Tukey box plot, Mann–Whitney test). g Mitotic duration of HT1080 6TG parental and HT1080 6TG mCherry-TRF1OE cells with DMSO or APH treatment (three biological replicates scoring n ≥ 57 mitoses per condition compiled in a Tukey box plot, Mann–Whitney test). h Outcome of the mitotic events in (g) (mean ± s.e.m., n = 3 biological replicates, Fisher’s Exact Test). i Representative images of the mitotic DDR on cytocentrifuged chromosome as shown in (b) for HT1080 6TG cells treated with APH ± reversine, Hesperadin or KU-55933. j Quantitation of mitotic telomere and genomic DDR foci for the conditions shown in (i) (three biological replicates scoring n ≥ 37 chromosome spreads per condition are compiled in a Tukey box plot, Mann–Whitney test). k Telomere PNA FISH signal intensity in arbitrary units (A.U.) from individual telomeres in APH treated HT1080 6TG cells, sorted by γ-H2AX status (three biological replicates scoring n ≥ 1529 telomeres per condition compiled into a Tukey box plot, Mann–Whitney test). For all panels, ns = not significant, **p < 0.01. Scale bars represent 10 µm. Source data are provided as a Source Data file
