Fig. 5: FTO-PARP1 interaction, but not FTO demethylase activity, is required for PARP1 clustering in cells.

a A schematic diagram of the domain structure of FTO depicting binding sites for Fe²⁺ and PARP1. b Recombinant Flag-PARP1 proteins have reduced binding to His-FTO-Δ28 (28 amino acid deletion at the N-terminal) compared to full-length (FL) His-FTO. c Quantification of PARP1 binding. Data were normalized to the His-FTO-FL group (N = 7) and analyzed using a two-tailed paired t-test. d FTO KO HeLa cells expressing myc-FTO (WT, Δ28 or DD) were irradiated with 100 J UVC and recovered for 4 min. PLA signals (gray) between endogenous PARP1 and myc-FTO were present within nuclei (marked by dashed lines that outline the DAPI stain). Transfected cells were identified by the presence of myc staining (green). Scale bar, 20 μm. e Quantification of the number of PLA puncta per nucleus. (WT-Ctrl, N = 55 cells; WT-UV, N = 51 cells; Δ28-Ctrl, N = 48 cells; Δ28-UV = 29 cells; DD-Ctrl = 43 cells; DD-UV = 35 cells). Data were analyzed using one-way ANOVA with Tukey’s multiple comparison test. f Representative Airyscan2 confocal images of FTO KO HeLa cells overexpressing myc-FTO (WT, Δ28 or DD) that were irradiated with 100 J UVC or not, and immunostained for myc (green), PARP1 (magenta) and DAPI (blue). Scale bars, 10 μm and 1 μm (inset). g Quantification of the number of PARP1 clusters per nucleus area (KO-Ctrl, N = 80 clusters; KO-UV, N = 64 clusters; WT-Ctrl, N = 64 clusters; WT-UV, N = 39 clusters; Δ28-Ctrl, N = 52 clusters; Δ28-UV = 36 clusters; DD-Ctrl = 59 clusters; DD-UV = 43 clusters). Data were analyzed using one-way ANOVA with Tukey’s multiple comparison test. All data are represented as mean ± SEM. Source data are provided as a Source Data file.