Figure 1. Mitochondrial localization of... | Rockefeller University Press

$Figure 1. Mitochondrial localization of Parkin is etiologically important. (A) HeLa cells expressing HA-Parkin were treated with CCCP or DMSO (control) and then immunostained with the indicated antibodies. (B) HeLa cells stably expressing HA-Parkin or intact SH-SY5Y cells were treated with CCCP or DMSO and subjected to fractionation experiments. I, C, and M indicate input, cytosol-rich supernatant, and mitochondria-rich membrane pellet, respectively. (C) Schematic diagram of disease-relevant mutants of Parkin used in this study. IBR, in between RING; Ubl, ubiquitin like. (D) Polarized mitochondria stained with MitoTracker red (red arrowheads) were not labeled by Parkin. In contrast, damaged mitochondria marked by Parkin (green arrowheads) were not stained with MitoTracker red. (E) HeLa cells expressing HA-Parkin with various pathogenic mutations were treated with CCCP, followed by immunocytochemistry. (A, D, and E) Higher magnification views of the boxed areas are shown in the insets. (F) Parkin colocalization with mitochondria was analyzed in >100 cells per mutation. Example figures indicative of robust colocalization (counted as 1), weak colocalization (counted as 0.5), and no colocalization (counted as 0) are shown. Error bars represent the mean ± SD values of at least three experiments. Bars: (A, E, and F) 10 µm; (D) 30 µm.$

Figure 1.

Mitochondrial localization of Parkin is etiologically important. (A) HeLa cells expressing HA-Parkin were treated with CCCP or DMSO (control) and then immunostained with the indicated antibodies. (B) HeLa cells stably expressing HA-Parkin or intact SH-SY5Y cells were treated with CCCP or DMSO and subjected to fractionation experiments. I, C, and M indicate input, cytosol-rich supernatant, and mitochondria-rich membrane pellet, respectively. (C) Schematic diagram of disease-relevant mutants of Parkin used in this study. IBR, in between RING; Ubl, ubiquitin like. (D) Polarized mitochondria stained with MitoTracker red (red arrowheads) were not labeled by Parkin. In contrast, damaged mitochondria marked by Parkin (green arrowheads) were not stained with MitoTracker red. (E) HeLa cells expressing HA-Parkin with various pathogenic mutations were treated with CCCP, followed by immunocytochemistry. (A, D, and E) Higher magnification views of the boxed areas are shown in the insets. (F) Parkin colocalization with mitochondria was analyzed in >100 cells per mutation. Example figures indicative of robust colocalization (counted as 1), weak colocalization (counted as 0.5), and no colocalization (counted as 0) are shown. Error bars represent the mean ± SD values of at least three experiments. Bars: (A, E, and F) 10 µm; (D) 30 µm.

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