PINK1 Is Selectively Stabilized on Impaired Mitochondria to Activate Parkin
Figure 2
PINK1 accumulates following inhibition of voltage-sensitive cleavage.
(A) HeLa cells stably expressing YFP-Parkin were treated with DMSO for 3.5 h, 2 µM CCCP for 3.5 h, or CCCP for 3 h followed by washout of CCCP for 0.5 h in the absence of serum. 50 µM MG132 and/or 100 µM cyclohexamide were added for the last 1 h of treatment. Whole-cell lysates (WCL) run on SDS gels and immunoblotted for endogenous PINK1 and tubulin. (B) Model depicting the two-step processing of PINK1. (C) Quantitative RT-PCR was used to measure relative PINK1 mRNA expression in HeLa cells treated with DMSO or CCCP for 1 h. The graph represents the results from four independent experiments. As a positive control, relative PINK1 mRNA levels were also measured in HeLa cells following exogenous expression of PINK1. PINK1 mRNA expression levels were normalized to the housekeeping gene β-actin. WT, wild type.