Figure 6

Effect of HSD10-modification on CypD and how it may influence cancer cell growth and death. A. EV and HSD10 ov whole cell lysates were analyzed for CypD protein expression using immunoblotting. β-actin was used as the loading control, and CypD was normalized to actin (n = 4). B. Control shRNA and HSD10 shRNA whole cell lysates were analyzed for CypD protein expression using immunoblotting. β-actin was used as the loading control, and CypD was normalized to actin (n = 4). C. EV and HSD10 ov whole cell lysates were analyzed for HSD10-CypD complexes using co-immunoprecipitation. β-actin was used as the loading control for the input. The immunoblots demonstrate an increased HSD10-CypD interaction in PC-12 cells overexpressing HSD10 compared to EV cells. D. Confocal microscopy demonstrating immunofluorescence staining of HSD10 alone (red), CypD alone (green), and these two antigens co-localized (yellow) in EV and HSD10 ov cells. E. Immunofluorescence staining of HSD10 alone (red), mitochondrial marker SODII alone (green), and these two antigens co-localized (yellow) in HSD10 ov cells. F. Immunofluorescence staining of CypD alone (green), mitochondrial marker Hsp60 alone (red), and these two antigens co-localized (yellow) in HSD10 ov cells. Scale bar in F: 20 μm. G-H. Quantification of HSD10 and CypD fluorescence densities (depicted in D) displayed as fold increase (n = 4). Data presented as mean ± SE. *P < 0.01 versus transfected control group. I. Hypothetical mechanism of action for HSD10-mediated cancer cell growth. Top panel, cells overexpressing HSD10 bind to CypD and sequester it in the mitochondrial matrix, thereby avoiding cell death induction; this resistance allows for continued cancer cell proliferation. Bottom panel, cells under-expressing HSD10 cannot bind all of the available CypD; thus unbound CypD translocates to the IM where it induces cell death via MPTP opening.