Figure 5.

PRMT1 protects breast cancer cells from IR-induced apoptosis. (A) Apoptosis determination in MCF7 cells subjected to the indicated treatments using an ELISA (left panel). At the zero time point, the cells were treated with AA and AdOx. On day 1, the cells were irradiated (10 Gy), and apoptosis was assayed on day 4. When the cells were not irradiated, apoptosis analyses were performed on the fourth day. In the right panel, MCF7-Luc2 cells were subjected to indicated siRNA transfections. The number of cells remaining at three days post-IR was quantified by bioluminescence imaging, while apoptosis was determined by ELISA (histogram; *P < 0.05). (B) Depletion of PRMT1 sensitizes 4T1 cells to IR in vitro and in vivo. The images in the left panel show the effect of IR on apoptosis in the indicated 4T1 cells. The scale bar (100 μm) refers to both panels. 4T1-control-transfected and 4T1-PRMT1-KO cells were subcutaneously injected near the mammary fat pads of athymic nu/nu mice (arrows represent the initial injection site). Primary tumors were recorded by using bioluminescence imaging (n = 5 per group) and compare nonirradiated (w/o) vs. IR-treated mice (middle panel). Right panel shows the tumor sizes at 40 days after the tumor cells were injected (scale bar, 1 cm). The growth of 4T1-PRMT1-KO tumors in IR-treated mice was significantly reduced (P < 0.005) compared with the other groups. (C) Overexpression of PRMT1 protects MCF7 cells from IR-induced apoptosis. Western blots showing PRMT1 expression (left panel). The images show the effect of IR on apoptosis in MCF7 cells using Hoechst 33342. Analyses of apoptosis were performed at 72 h after IR. The scale bar (100 μm) refers to both panels. (D) Nuclear focus formation of γH2AX in MCF7 cells was observed following double-stranded DNA damage after the indicated treatments. Scale bar represents 8 μm. Blots in this figure were cropped from different gels. Full blots are shown in the Supplementary Information, Fig. S8.