Fig. 5. DNA damage abolishes the ciliary SUMO protease SENP1 and enhances FBF1 SUMOylation.
a Immunofluorescent images for SENP1 staining in RCTE cells. Basal bodies were labeled with HYLS1. Scale bar, 10 μm. b Structured illumination microscopic images stained for FBF1 and SENP1 at ciliary base in IMR-90 cells. Scale bar, 1 μm. c Co-immunoprecipitation of SENP1 and FBF1 in 293T cells. d Endogenous SENP1 or FBF1 immunoprecipitates with FBF1 or SENP1 in RCTE cells. e Immunofluorescent images in RCTE cells at day 7 without or with irradiation treatment using antibodies against SENP1 and FBF1. Basal bodies were labeled with HYLS1. Scale bar, 10 μm. f Endogenous FBF1 immunoprecipitates with SENP1 in RCTE cells with or without irradiation. g SENP1 diminished FBF1 SUMOylation in a co-immunoprecipitation assay in 293T cells. h Immunofluorescent images for FBF1, PML and YFP in WT or ARL3−/− RCTE cells co-transfected with CFP-FRB-CEP170c and YFP-FKBP-SENP1 plasmids at day 7 after irradiation. n=30 cells. Scale bar, 10 μm. i Proposed model: DNA-damage stress strongly downregulates ARLs, enables FBF1-UBC9 interaction, and results in FBF1 SUMOylation. SUMOylated FBF1 translocates from the ciliary base to PML-NBs, which is required for stress-induced PML-NB upregulation and senescence initiation. Three experiments were repeated independently with similar results (a–h). One-way ANOVA followed by Bonferroni multiple-comparison analysis was employed for h. Source data are provided as a Source Data file.