Fig. 7

NAT10-independent recruitment of ac4C RNA to UVA-damaged chromatin. A Nuclear distribution of NAT10 (red) and ac4C RNA (green) in physiological conditions. MCF7 cells were studied instead of MEFs as antibodies are only available against the human epitope. B Local laser microirradiation showed that NAT10 acetyltransferase (red) does not recruit to DNA lesions (positive on ac4C RNA; green) induced in MCF7 cells. Scale bars are 5 µm. C Western blots showed no changes in NAT10 levels in cells exposed to UVA and UVC light. An effect of UV- as well as γ-radiation, was confirmed by an increased γH2AX level. D shows western blot results on the level of NAT10 and γH2AX in NAT10 (wt) and NAT10 (dn) cells. Western blot data were normalized to the level of α-tubulin, and proteins were loaded following the identical total protein levels. E A representative anti-ac4C dot blot (#A18806, Abclonal) was performed to study total, long, and small RNA in NAT10 (wt) and NAT10 (dn) cells. F Quantification of dot blot results from E is shown in panel F. Asterisks (*) indicate a statistically increased level of ac4C in RNA. G The level of ac4C RNA (green) and NAT10 (red) in non-irradiated control and UVA- or UVC-irradiated whole populations of a NAT10 (wt) and b NAT10 (dn) HeLa cells. H Box plot graphs display the absolute intensity of ac4C RNA in the nucleoplasm (nucleus), ***p ≤ 0.001 (ANOVA One-Way test). I Box plot graphs show the total intensity of fluorescently stained ac4C RNA in nucleoli, ***p ≤ 0.001, **p ≤ 0.01. J Box plot graphs depict the ratio of the fluorescent intensity of ac4C RNA occupying nucleoli and the nucleoplasm (whole nucleus), ***p < 0.001, **p ≤ 0.01. K The level of ac4C RNA (green) and NAT10 (red) in microirradiated a NAT10 (wt) and b NAT10 (dn) HeLa cells