Fig. 4. Effect of transcription blockage on hOGG1 relocalisation to nuclear speckles. (A) Images corresponding to single optical sections from the middle of cells were thresholded and binarised and the percentage of round speckles calculated in NI and UVA-irradiated cells. Representative images are presented. (B) RNA synthesis was visualised by incorporation of BrU for 60 minutes in NI and UVA irradiated cells. BrU incorporation was detected with an antibody directed against BrdU (red). Arrows indicate nucleoli in the NI cells. (C) Cells were treated for 2 hours with the transcription inhibitors actinomycin-D (2 µg/ml) or -amanitin (50 µg/ml) and immunostained with SC35 antibodies (red). hOGG1-GFP was excluded from speckles (see insets). (D) Actinomycin-D-treated cells were stained with PI-RNase (red). (E) hOGG1-GFP-expressing cells were UVC-irradiated and stained with the antibody against SC35 (red). Insets show the exclusion of hOGG1-GFP from nuclear speckles. Bar, 4 µm (B,C,E); 2 µm (D).