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. 2019 Aug 8;75(3):605–619.e6. doi: 10.1016/j.molcel.2019.05.026

Figure 1.

Figure 1

EXD2 Is Recruited to Stressed Replication Forks

(A) Western blot of iPOND samples. Thymidine chase analysis illustrates that EXD2 specifically associates with the replisome. PCNA acts as a control.

(B) Schematic of the proximity ligation assay (PLA) employed to detect colocalization of target proteins with nascent DNA.

(C) Percentage of cells with MRE11/biotin PLA foci (mean ± SEM, n = 3 independent experiments, t test). Right: representative images of PLA foci (red), DAPI acts as a nuclear counterstain. Scale bar, 10 μm.

(D) Percentage of cells with GFP/biotin PLA foci (mean ± SEM, n = 3 independent experiments, t test) in U2OS control cells and U2OS cells expressing GFP-EXD2. Right: representative images of PLA foci (red), DAPI acts as a nuclear counterstain. Scale bar, 10 μm.

(E) Laser microirradiation induces rapid redistribution of GFP-EXD2 to damaged chromatin; representative images showing GFP-EXD2 accumulation at laser-generated DNA lesions. GFP-CtIP was used as a positive control. Scale bar, 10 μm.

(F) Quantification of GFP-EXD2 (left panel) and GFP-CtIP (right panel) recruitment kinetics (intensity versus time) to laser-generated DNA lesions (mean ± SE, n ≥ 10 cells from 2 independent experiments).