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. Author manuscript; available in PMC: 2016 Oct 14.
Published in final edited form as: Oncogene. 2016 Apr 4;35(41):5362–5376. doi: 10.1038/onc.2016.77

Figure 6. 53BP1 depletion rescues growth arrest in VDR-depleted cells.

Figure 6

(A) MCF-7 cells carrying an HR reporter construct (DR-GFP) were lentivirally transduced with shRNA targeting VDR or shRNA control (shscr). We monitored the percentage of GFP-positive cells resulting from HR of I-SceI-induced DSBs by FACS in 4 independent experiments. Note how depletion of VDR (immunoblotting) results in decreased HR efficiency. (B) BJ+hTert fibroblasts were depleted of 53BP1 via lentiviral transduction with specific shRNA (sh53BP1) prior to depletion of VDR (shVDR). Hairpins shscr and shLuc were used as control for the depletions. After dual selection, cells were processed for immunoblotting to monitor the levels of BRCA1, VDR, and 53BP1. Vinculin was used as loading control. (C) Percentage of cells positive for SA-β-Gal activity was calculated 6 days after the second lentiviral transduction with shRNAs. Graph shows average±sem of 2 biological repeats. (D) Proliferation of the different cell lines generated was evaluated by counting cells days 2–11 after dual selection, as described in methods. The graph shows average±sem of 4 biological repeats. Note how depletion of 53BP1 prevents growth arrest in VDR-depleted cells (Day 11, *p=0.001).