Figure 1.
UBC13 Contributes to DSB Repair by Recruiting BRCA1 and RAP80 onto DSB Sites during the G1 Phase
(A) Average number of etoposide-induced FK2 foci in MCF-7 cells synchronized during the G1 phase by serum starvation (24 h). We examined wild-type MCF-7 cells treated with shRNA targeting UBC13 (shUBC13) and non-targeting shRNA (shControl). Synchronized cells were treated with etoposide (10 μM) for 30 min, washed, and incubated with etoposide-free media. Error bars show the standard deviation (SD) from three independent experiments. At least 50 G1-phase (cyclin A-negative) cells per experiment were counted. Single asterisk indicates p = 1.1 × 10−3, calculated by Student's t test. Representative images and box plots of FK2 foci are shown in Figures S1A and S1D, respectively.
(B) DSB-repair kinetics of G1-phase MCF-7 cells after pulse exposure (0–0.5 h) to etoposide (10 μM). Average number of γH2AX foci was counted at 0.5 and 8.5 h after addition of etoposide. Data are as shown in (A). Single, double, and triple asterisks indicate p = 1.2 × 10−3, p = 5.6 × 10−4, and p = 3.2 × 10−5, respectively, calculated by Student's t test. The box plots of γH2AX foci are shown in Figure S1F.
(C and D) Average number of etoposide-induced BRCA1 (C) and RAP80 (D) foci per cell. The experimental procedure and data are as shown in (A). Asterisks indicate p = 9.3 × 10−4 in (C) and p = 8.0 × 10−3 in (D). Representative images and box plots of BRCA1 and RAP80 foci are shown in Figures S1I and S1J, respectively.