Persistent ER Stress Induces the Spliced Leader RNA Silencing Pathway (SLS), Leading to Programmed Cell Death in Trypanosoma brucei
Figure 9
ΔΨm depolarization and ROS formation during SLS.
A. Measurements of mitochondrial outer membrane potential. Cells were harvested and loaded with 150 nM Tetramethyl rhodamine methyl-ester (TMRM) in serum free medium. The samples were incubated in the dark for 15 min at 27°C and then analyzed by FACS. i. Cells uninduced (control) and silenced cell [72 hours (SRα) or 48 hours (SEC63, SEC61)] (+Tet) ii. Cells treated with 4 mM DTT for 0 to 5 hours. The data represent the percentage of cells lacking mitochondria membrane potential obtained from three independent experiments and the S.D. is indicated. B. Measurements of reactive oxygen species formation. Cells were harvested and incubated at 27°C for 30 min with 10 µM DCFH-DA. The cells were washed once with PBS and analyzed by FACS. i. Uninduced cells (control) and silenced cells [72 hours (SRα) or 48 hours (SEC63, SEC61)] (+Tet). ii. Cells treated with 4 mM DTT for 0 to 5 hours. The data represent the average of three independent experiments and the S.D is indicated.