Figure 3.

SPOC1 dose-dependent modulation of DDR after γ-IR. SPOC1 expression in CV1 cell lines was determined by immunoblotting cell extracts with the indicated antibodies (Supplementary Information). (A) SPOC1 protein levels 72 h post-siRNA transfection; (B) Endogenous and FLAG-SPOC1 expression in three stably transformed CV1 cell lines (clones #14, #17 and #23) pre- or 24 h post-Dox treatment. Signals were quantified by densitometry. (C–F) IRIF visualized by indirect immunofluorescence of CV1 cells (unpublished observations) were counted by automated imaging at various times after γ-irradiation. (C, E) γH2AX and 53BP1 foci in CV1 cells 72 h after siRNA-mediated reduction of SPOC1 (siSPOC1) compared to control cells (siCtr). (D, F) γH2AX and 53BP1 IRIF in CV1 clone #14 before and after Dox-induced (24 h) expression of FLAG-SPOC1. The IRIF of 700–1000 cells were counted in each of three independent radiation experiments and at different time points, and are represented as the mean/cell ± SEM. *P ≤ 0.05, **P ≤ 0.01, ***P≤ 0.001.