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Fig. 7 | Molecular Neurodegeneration

Fig. 7

From: Hexokinases link DJ-1 to the PINK1/parkin pathway

Fig. 7

AKT signaling promotes PINK1/parkin pathway activation. a Mitochondrial membrane potential after overnight incubation with MK2206 in HeLa cells with and without the protonophore CCCP. b-d AKT was inhibited in YFP-parkin expressing HeLa cells using MK2206 (10 μM) for 24 h prior to a 1-h treatment with 10 μM CCCP and then YFP-parkin was immunoprecipitated. Levels of immunoprecipitated HK2, VDAC1, and parkin proteins are shown in (b) and quantified in (c) and (d) (n = 7 replicates done over the course of 4 separate experiments, one sample t-test comparing MK2206/DMSO versus 1). e-g HeLa cells stably expressing YFP-parkin and MitodsRed2 were treated with MK2206 (10 μM) prior to 1-h treatment with CCCP (5 μM) and cells were analyzed using high-content imaging. Photomicrographs of Hoechst (nuclei), YFP-parkin, MitodsRed2, and pUbiquitin S65 are shown in (e). Quantification of the overlap between YFP-parkin and MitodsRed2 is shown in (f), while the intensity of pUbiquitin S65 in mitochondria is displayed in (g) (n = 400 cells/well, 5 wells per condition). Two-way ANOVA with Bonferroni’s multiple comparison test was used to compare groups. Scale bar: 50 μm. h HeLa cells stably expressing YFP-parkin and MitodsRed2 were treated with MK2206 prior to 2-h treatment with valinomycin (10 μM). i Quantification of YFP-parkin overlap with MitodsRed2 from (h) (n = 400 cells/well, 11–12 wells per condition). Comparison and scale bar as shown in (e-g)

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