Fig. 3: Decreased PML-NB number correlates with SG persistence in HeLa cells.
From: Loss of PML nuclear bodies in familial amyotrophic lateral sclerosis-frontotemporal dementia
A, B G3BP2-GFP HeLa Kyoto cells were lipofected with a non-targeting control siRNA (siRNA control) or with siRNA specific for PML or Ubc9, respectively. 72 h post-transfection, cells were either left untreated or treated with sodium arsenite (500 μM) for 45 min. Where indicated cells were allowed to recover in drug-free medium for 4 h (rec. 4 h). Cells were fixed (A, B), subjected to immunostaining with a PML-specific antibody (A). Nucleic acid was labeled using DAPI (A, B). PML-NBs were automatically segmented and quantified with ScanR (A). The number of PML-NBs analyzed is reported: siRNA control (1026, control; 1161, arsenite); siRNA PML (1042, control; 1260, arsenite); siRNA Ubc9 (1098, control; 1164, arsenite). One-way ANOVA, Post-hoc Bonferroni-Holm test. Panel B shows the percentage of SG-positive cells and representative images. n = 3, +/− SEM; P = n.s. (non-significant). One-way ANOVA, Post-hoc Bonferroni-Holm test. C Immunoblotting showing PML (left panel) and Ubc9 (right panel) siRNA efficacy in mammalian cells. Ponceau staining shows equal loading. A, B Scale bar is 10 μm.
