Figure 6.

Endogenous parkin regulates the apoptotic release of cytochrome c from mitochondria. (A) The ectopic expression of parkin in SH-SY5Y cells substantially attenuates the BH3 peptide induce cytochrome c release from isolated mitochondria, with a concomitant decrease in the retained cytochrome c (B), as determined by western blot. Equal loading of mitochondrial proteins was confirmed by western blot analysis of VDAC. (C) Whole cell lysates from naïve SH-SY5Y cells and those stably transduced with a lentivirus encoding an shRNA directed against the expression of endogenous parkin were probed by western blot for both parkin and actin levels. (D) Mitochondria from SH-SY5Y cell lines with and without shRNA against parkin expression were incubated with DMSO vehicle, 1 µm of the purified BH3 domain peptides of Bid, Bim and PUMA, alamethicin (40 µg/ml), or 1% NP-40. Released and retained cytochrome c, and mitochondrial VDAC levels were analyzed by western blot. (D) is representative of at least three independent experiments.