EXD2’s Nuclease Activity Is Required to Suppress Replication Fork Collapse
(A) Quantification of the frequency of 53BP1 foci in HeLa WT and EXD2−/− S/G2 cells and representative images. Cyclin A (green) acts as a marker for S/G2 cells, DAPI acts as a nuclear stain (mean ± SEM, n = 3 independent experiments, Mann-Whitney). Scale bar, 10 μm.
(B) Quantification of the frequency of chromosomal aberrations from mitotic spreads from HeLa WT and EXD2−/− cells (mean ± SEM, n = 75 metaphase spreads pooled from 3 independent experiments, t test).
(C) Representative images of metaphase spreads from B). Arrows indicate chromatid breaks. Scale bar, 6.5 μm.
(D) Boxplot of CldU tract length ratios of associated sister forks from HeLa WT, EXD2−/−, and EXD2−/− cells complemented with either Flag-EXD2 WT or Flag-EXD2 nuclease dead (ND) mutant protein (5–95 percentile, n ≥ 60 sister fork pairs pooled from 3 independent experiments, Mann-Whitney).