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. 2015 Nov 5;60(3):362–373. doi: 10.1016/j.molcel.2015.09.019

Figure 2.

Figure 2

USP4 Functions in DNA-End Resection

(A) USP4 depletion caused HR defects [direct-repeat (DR)-GFP reporter assays]. Quantifications were normalized to control siRNA-treated cells and set to 100% (mean ± SEM; n = 4).

(B) USP4 depletion sensitized U2OS cells to camptothecin (CPT) (mean ± SEM; n = 3 and CtIP siRNA-treatment was the positive control).

(C) USP4 depletion reduced RPA2 Ser-4/Ser-8 phosphorylation (S4S8p) after IR (10 Gy).

(D) USP4 depletion reduced RPA2 S4S8p after camptothecin (1 μM, 1 hr) treatment. Intensities were quantified with Odysey CLx (LI-COR) and Image Studio 4.x software and RPA2 S4S8p was normalized to RPA2. Quantifications were normalized to the camptothecin-treated siControl and set to 100% (mean ± SEM; n = 3).

(E) USP4 siRNA treatment, followed by camptothecin (1 μM, 1 hr) treatment, of U2OS cells reduced resection (BrdU intensities). Quantifications were normalized to the camptothecin-treated siControl (CtIP depletion was the positive control; mean ± SEM; n = 3).

(F) GFP-USP4 WT(L)-complemented U2OS cells restored resection defects (BrdU intensities) observed upon USP4 depletion. Quantifications were normalized to camptothecin and Control siRNA-treated GFP cells (mean ± SEM; n = 3; ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; n.s., not significant). See also Figure S2.