Abstract
In this study, we develop a simple assay to identify mitophagy inducers on the basis of the use of fluorescently tagged mitochondria that undergo a colour change on lysosomal delivery. Using this assay, we identify iron chelators as a family of compounds that generate a strong mitophagy response. Iron chelation-induced mitophagy requires that cells undergo glycolysis, but does not require PINK1 stabilization or Parkin activation, and occurs in primary human fibroblasts as well as those isolated from a Parkinson's patient with Parkin mutations. Thus, we have identified and characterized a mitophagy pathway, the induction of which could prove beneficial as a potential therapy for several neurodegenerative diseases in which mitochondrial clearance is advantageous.
Highlights
Mitochondrial damage is associated with many human diseases and maintenance of a viable pool of mitochondria is considered fundamental to cell function [1]
The assay relies on differences in pKa of green fluorescent protein (GFP) and mCherry and is adapted from a method developed for general autophagy [10]
The assay consists of cells expressing a tandem mCherry–GFP tag attached to the outer mitochondrial membrane localization signal of the protein FIS1
Summary
Mitochondrial damage is associated with many human diseases and maintenance of a viable pool of mitochondria is considered fundamental to cell function [1]. Received 4 June 2013; revised 30 September 2013; accepted 1 October 2013; published online 1 November 2013 observations that the protein kinase PINK1 and E3 ubiquitin ligase Parkin, both mutated in early onset forms of Parkinson’s disease, act to induce mitophagy on mitochondrial membrane depolarization [7]. This implies, at least in patients harbouring mutated PINK1 or Parkin, impaired removal of mitochondria might contribute to neuronal death. We screened compounds known to disrupt mitochondrial function or induce autophagy and identified iron chelation as a strong PINK1/Parkin-independent activator of mitophagy
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