Figure 1

Impaired mitochondrial respiration in PINK1 −/− fibroblasts. A. Representative oxygraphs of PINK1−/− (−/−) and wild-type (+/+) MEFs energized with glucose (10 mM). The arrow indicates the time MEFs are added to the chamber. B. Bars show oxygen consumption, which represents the endogenous respiratory activity in PINK1−/− and +/+ MEFs. C Representative oxygraphs of PINK1−/− and +/+ MEFs energized with 10 mM glutamate/malate (complex I substrate), 10 mM succinate (complex II substrate) or 1 mM TMPD/1 mM ascorbate (complex IV substrate) in the presence of ADP (1 mM). Arrows indicate the time of the addition of either the substrates or oligomycin (2 μM). D. Bars show State 3 respiratory activity for complex I, II and IV in PINK1−/− and +/+ MEFs permeabilized with digitonin. E. Enzymatic activities of complexes I, II, III and IV of the mitochondrial electron transport system, as measured by spectrophotometric assays and after normalization to citrate synthase activity. F. Graph represents succinate-cytochrome C oxidase (complex II + III) activity. G. Upper panel: Representative western blot showing levels of cytochrome C in the mitochondrial fraction of PINK1−/− and +/+ MEFs. Hsp75 is used as a control for the total amount of mitochondrial proteins loaded in each well. Lower panel: The bar graph shows relative quantification of the level of Cytochrome C using Hsp75 as loading control. All data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01.