FIG. 4.

SIRT3 expression is required for survival of HeLa cells after genotoxic stress. (A) Representative Western blot showing endogenous expression of SIRT3 in control cells and cells subjected to SIRT3 knockout. Note reduced levels of both forms of SIRT3 after siRNA-mediated knockdown. (B) Western blot showing expression of SIRT3 in control and stable HeLa cells. (C) Western blot showing reduced levels of both forms of SIRT3 after siRNA-mediated knockdown in stable cells. (D) HeLa cells (control and endogenous SIRT3 knockdown cells) were treated with MNNG (500 μM) for 10 min and then washed and kept in complete medium for 6 h. Cell death was determined by Hoechst and PI staining followed by FACS analysis. (E) SIRT3 stable cells expressing wt SIRT3 or mt SIRT3 were examined for MNNG-mediated cell death. (F) SIRT3 stable cells were subjected to SIRT3 knockdown by siRNA and then tested for MNNG-mediated cell death. SIRT3 antibody PAB-11098 was used in all these Western blot assays. Values are means ± standard errors of three experiments.