Fig. 3

ArgS regulates antioxidative proteins to decrease cell recovery and migration. A Immunoblot analysis (left panel) of the antioxidant proteins HO-1, CISD2, SOD1, and GSR in BT-549 cells treated with ArgS or TN (2 μg/ml) for 24 h. One representative immunoblot (n = 3) is shown. The relative protein level (right panel) was calculated by normalizing the protein signal intensity in the + Arg group, which was set to 1 after normalization with Lamin A/C. B Analysis of ROS levels in BT-549 (top panel) and MDA-MB-231 (bottom panel) cells treated with a combination of ArgS and HO-1 activators or inhibitors (CoPPIX or ZnPPIX) or HO-1 knockdown/overexpression. BT-549 cells were transfected with siHMOX1 (30 nM) or treated with ZnPPIX (5 μM), and MDA-MB-231 cells were stably overexpressing HO-1 or treated with COPPIX (12.5 μM). ROS levels were monitored by DCF-DA oxidation after 24 h of ArgS treatment. C Analysis of cell recovery in BT-549 (left panel) and MDA-MB-231 (right panel) cells treated with a combination of ArgS and HO-1 activators or inhibitors (CoPPIX or ZnPPIX) or HO-1 knockdown/overexpression, as described in B. The cell recovery was determined as described previously in Fig. 2D. D Analysis of cell migration in MDA-MB-231 cells stably overexpressing HO-1 or treated with COPPIX (12.5 μM) after 24 h of ArgS treatment, as described previously in Fig. 2E; n = 3. Data are shown as mean ± s.e.m.; *p < 0.05; **p < 0.01; ***p < 0.001; Two-Way ANOVA followed by Dunett’s (A) or Tukey's (B-D) multiple comparison tests