Fig. 8: Nucleolar segregation of broken repeats requires UFD1L in human cells.
a Retention of rDNA locus (NPM1) in human cells upon DNA damage treated with siRNA against UFD1L. Human RPE cells stably expressing FKBP12-HAI-PpoI were transfected with siRNAs against UFD1L (siUFD1La and siUFD1Lb), or treated with the ATM kinase inhibitor KU55933 (ATMi). Representative images of immunofluorescence staining after rDNA damage, using the indicated antibodies, are shown. Scale bar, 10 µm. b Quantification of human RPE cells transfected with siRNA against a control (siControl; n = 4), RNF4 (siRNF4; n = 3), siUFD1L (siUFD1La and siUFD1Lb; n = 4), combination thereof (n = 3), or ATMi treated (n = 4), showing the percentage of cells with nucleolar retention of γH2AX foci. Data are mean ± SEM of n biologically independent experiments. Statistical analysis was performed using ANOVA, and letters denote significant differences with a Tukey’s post hoc test at P ≤ 0.05. c Immunoblot of RPE cells from Fig. 7a after rDNA damage, showing knock-down efficiency of UFD1L and HAI-PpoI induction. H3 served as loading control. d Model for rDNA release upon DNA damage: Under normal conditions, the rDNA repeats are kept inside the nucleolus by Cdc14-mediated dephosphorylation of Nur1, which maintains the interaction of the rDNA tethering complex. When a repeat unit is damaged, SUMOylation of CLIP-cohibin disrupts the complex through Cdc48-Npl4-Ufd1, allowing rDNA relocation outside the nucleolus. Nur1 is also phosphorylated, further supporting the broken repeat relocation. Source data are provided as a Source Data file.
