Fig. 4

MitoVES affects mitochondrial complexes. (A) NeuTL spheres were treated with 2 μM MitoVE for and 4 h, before they were harvested, permeablised with saponin and evaluated for respiration at the presence of substrates specific for CI and CII using the protocal indicated in more detail in Materials and Methods. The abbreviations in the top left line graph are: L, leak; CI, complex I; CII, complex II; ETS, electron transfer system (uncoupled resiraiton); CII’, uncoupled respiration via CII; ROX, residual respiration; PMG, pyruvate, malate and glutamate; cyt c, cytochrome c; succ, succinate; F, FCCP; rot, rotenone; ama, antimycin A. (B) The respiration via CI and CII, and the uncoupled respiration via CI (CI’) and CII (CII’) as derived from results shown in panel A is documented in control cells and cells exposed to 10 μM MitoVES for 2 and 4 h. (C) The mitochondrial fraction, prepared from control NeuTL cells or cells exposed to 10 μM MitoVES for 2 and 4 h, was lysed in the presence of digitonin and subjected to native blue gel electrophoresis as detailed in Materials and Methods. Specific subunits of individual complexes were detected using the antibodies as shown. HSP60 was used as a loading control. The symbol ‘*’ in panels indicates statistically significant differences (p < 0.05) for the respiration after cells were exposed to MitoVES