Fig. 2: Impairment of ESCRT-III function affects micronuclei integrity.

a HeLa cells were transfected with the indicated sRNAi for 48 h and the percentages of cells with at least one micronucleus were quantified for each treatment (minimum 250 cells per treatment, per repeat). Results were analyzed using a one-way ANOVA with Dunnett’s post hoc test. b HeLa cells were transfected with the indicated sRNAi for either 24, 48, or 72 h. The number of micronucleated cells were quantified for each one of the treatments (minimum 200 cells were scored for each treatment, per repeat). Results were analyzed using a two-way ANOVA with Dunnett’s post hoc test. c The average number of micronuclei observed per micronucleated cell were quantified (minimum 150 cells per treatment, per repeat). Results were analyzed using a one-way ANOVA with Dunnett’s post hoc test. d Micronuclei in HeLa cells transfected for 48 h with the indicated sRNAi and scored for presence of CREST (minimum 200 micronuclei per treatment, per repeat). Results were analyzed using a one-way ANOVA with Dunnett’s post hoc test. e HeLa cells were transfected with the indicated sRNAi for 48 h and scored for the status of Lamin B, as defined in supplementary Figure 2A (minimum 100 micronuclei per treatment, per repeat). Results were analyzed using a one-way ANOVA with Dunnett’s post hoc test. Statistics shown for the “absent” category. f HeLa cells were transfected with the indicated sRNAi for 48 h and scored for the status of mab414, as defined in supplementary Figure 2B (minimum 100 micronuclei per treatment, per repeat). Results were analyzed using a one-way ANOVA with Dunnett’s post hoc test. Statistics shown for the ‘absent’ category. g Micronuclei in HeLa cells transfected for 48 h with the indicated sRNAi and scored for presence of PDI within micronuclear chromatin (PDI-invaded; minimum 100 micronuclei per treatment, per repeat). Results were analyzed using a one-way ANOVA with Dunnett’s post hoc test. h Micronuclei in HeLa cells transfected for 48 h with the indicated sRNAi and scored for presence of PARP1 (minimum 100 micronuclei per treatment, per repeat). Results were analyzed using a one-way ANOVA with Dunnett’s post hoc test. Averages and SEM are shown (N = 3, unless stated otherwise). i HeLa cells treated with control or CHMP7 sRNAi, then transfected with GFP-NLS post-seeding and imaged 48 h post sRNAi depletion. Examples of micronuclei with GFP-NLS absent (top panel) or present (bottom panel) (arrowheads). Scale bar 10 μm. j Quantification of GFP-NLS retention in micronuclei treated as in g as judged visually by the loss of nuclear signal (25 micronuclei scored per treatment, per repeat). Averages of N = 2 biological repeats are shown