Research Paper Volume 13, Issue 5 pp 7300—7313

Maf1 suppression of ATF5-dependent mitochondrial unfolded protein response contributes to rapamycin-induced radio-sensitivity in lung cancer cell line A549

Rapamycin suppresses IR-induced UPRmtthrough Maf1 in A549 cells. (A, B) A549 cells treated with IR and rapamycin were examined for UPRmt marker genes (mHSP70 and HSP60) expression by RT-qPCR. Experiments were performed for 3 biological repeats and data were normalized to non-treated control. (C, D) HSP70 and HSP60 protein levels were analyzed by Western blot. Representative results are shown in (C) and quantification of 3 biological repeats in (D). Data were normalized to non-treated control. (E, F) Maf1 knockdown activated UPRmt in an ATF5-dependent manner. A549 cells were knocked down for Maf1, ATF5 or both, then treated with rapamycin and IR. mHSP70 and HSP60 protein levels were analyzed by Western blot. Representative results are shown in (E). Experiment were performed for 3 times. Data were normalized to non-transfected control. For all bar graph, the error bars stand for Standard Deviation (SD) of the mean. Statistical significance was evaluated by 2-tailed, paired student’s t-test (ns, not significant, *, P

Figure 3. Rapamycin suppresses IR-induced UPRmtthrough Maf1 in A549 cells. (A, B) A549 cells treated with IR and rapamycin were examined for UPRmt marker genes (mHSP70 and HSP60) expression by RT-qPCR. Experiments were performed for 3 biological repeats and data were normalized to non-treated control. (C, D) HSP70 and HSP60 protein levels were analyzed by Western blot. Representative results are shown in (C) and quantification of 3 biological repeats in (D). Data were normalized to non-treated control. (E, F) Maf1 knockdown activated UPRmt in an ATF5-dependent manner. A549 cells were knocked down for Maf1, ATF5 or both, then treated with rapamycin and IR. mHSP70 and HSP60 protein levels were analyzed by Western blot. Representative results are shown in (E). Experiment were performed for 3 times. Data were normalized to non-transfected control. For all bar graph, the error bars stand for Standard Deviation (SD) of the mean. Statistical significance was evaluated by 2-tailed, paired student’s t-test (ns, not significant, *, P<0.05, **, P<0.01, ***, P<0.001).