Figure 1.

Mitotic telomere deprotection is suppressed by WRN helicase independently of its catalytic activities. (a) Immunoblot of WRN in IMR-90 E6E7 hTERT cells transduced with shWRN or shScramble for 5 days. GAPDH serves as a loading control. (b) Representative images of meta-TIF assay from WRN knockdown cells after treatment with 100 ng/ml colcemid. The images show DAPI (blue), γ-H2AX (red), and telomere FISH (green). Scale bar, 10 µm. (c) Quantification of telomeric signals colocalized with γ-H2AX foci per chromosome spread in indicated conditions. Violin plots illustrate the distribution of all data and averages from three independent experiments (n = 15/experiment for 2 h colcemid; n = 30/experiment for 24 h colcemid; mean ± s.e.m.; Kruskal–Wallis followed by Dunn’s test). (d) Immunoblot of WRN in IMR-90 E6E7 hTERT cells expressing exogenous WRN^RES or Vector. Cells were transduced with shScramble or shWRN for 5 days before analysis. GAPDH serves as a loading control. (e) Quantification of telomeric signals colocalized with γ-H2AX foci in indicated conditions as shown in c (n = 15/experiment for 2 h colcemid; n = 30/experiment for 24 h colcemid; mean ± s.e.m.; Kruskal–Wallis followed by Dunn’s test). (f) Immunoblot of WRN in cells expressing WRN^RES and indicated WRN mutants. Transduced cells were harvested on day 10 post-infection. GAPDH serves as a loading control. (g) Quantification of telomeric signals colocalized with γ-H2AX foci in indicated conditions as shown in c (n = 30/experiment; mean ± s.e.m.; Kruskal–Wallis followed by Dunn's test). Unprocessed blot images are shown in Supplementary Information files.