Skip to main content
. 2013 Apr;33(7):1285–1302. doi: 10.1128/MCB.06798-11

Fig 8.

Fig 8

SCO2-mediated apoptosis is ASK-1 dependent. (A) Two ASK-1 knockout cell lines were prepared from HCT p53+/+ and HCT p53−/− cells. The cell lines were validated for the presence of ASK-1 and p53 proteins by Western blotting. The results show that ASK-1 protein was missing from the HCT p53+/+ ASK-1−/− and HCT p53−/− ASK-1−/− cell lines (n = 4). (B) SCO2-induced apoptosis was observed in normoxic and hypoxic HCT p53+/+ ASK-1−/− and HCT p53−/− ASK-1−/− cell lines. Both tamoxifen and SCO2 induced significant apoptosis in normoxic cancer cells. However, SCO2-induced apoptosis was absent in ASK-1−/− cells (bar 3). Tamoxifen was not able to induce a high apoptotic fraction in the hypoxic cancer cells (bar 5). SCO2 induced high-level apoptosis in hypoxic cancer cells with the ASK-1+/+ genotype. SCO2-induced apoptosis was absent in ASK-1−/− cells (bar 6) (n = 7; error bars represent standard deviations determined by analysis of variance; an asterisk represents a significant difference in apoptosis between ASK-1+/+ and ASK-1−/− cells [P < 0.02]). (C) The role of ASK-1 in SCO2-induced tumors was studied by using tumor regression in SCO2-transfected HCT p53+/+ ASK-1−/− and HCT p53−/− ASK-1−/− tumor xenografts. Results show that SCO2 induced apoptosis in ASK-1+/+ tumors (graphs 1 and 2) but did not induce tumor regression in ASK-1−/− knockdown tumor xenografts (graphs 3 and 4). Untreated tumors and tumors where ROS was quenched by NAC served as controls (error bars represent standard deviations determined by analysis of variance; n = 5). (D) The role of ASK-1 in SCO2-mediated apoptosis in tumor xenografts was also observed in ASK-1 knockout tumor xenografts by using a FLIVO in vivo apoptosis kit. Results show significant decreases in the apoptotic fraction in ASK-1−/− tumor xenografts (bars 3 and 4) (a red asterisk represents a significant difference in SCO2-induced apoptosis between the HCT p53+/+ and HCT p53+/+ ASK-1−/− tumors [P < 0.04]; a blue asterisk represents a significant difference in SCO2-induced apoptosis between the HCT p53−/− and HCT p53−/− ASK-1−/− tumors [P < 0.043]; error bars represent standard deviations determined by analysis of variance; n = 7).