Semi-automated quantitation of mitophagy in cells and tissues

Lambert Montava-Garriga, Francois Singh, Graeme Ball, Ian G. Ganley (Lead / Corresponding author)

Research output: Contribution to journalArticlepeer-review

3 Citations (Scopus)
137 Downloads (Pure)

Abstract

Mitophagy is a natural phenomenon and entails the lysosomal degradation of mitochondria by the autophagy pathway. In recent years, the development of fluorescent pH-sensitive mitochondrial reporters has greatly facilitated the monitoring of mitophagy by distinguishing between cytosolic mitochondria or those delivered to acidic lysosomes. We recently published the mito-QC reporter, which consists of a mitochondrial outer membrane-localised tandem mCherry-GFP tag. This allows the quantification of mitophagy via the increase in red-only mCherry signal that arises when the GFP signal is quenched upon mitochondrial delivery to lysosomes. Here we develop a macro for FIJI, the mito-QC Counter, and describe its use to allow reliable and consistent semi-automated quantification of mitophagy. In this methods article we describe step-by-step how to detect and quantify mitophagy and show that mitophagy levels can be reliably calculated in different cell lines and under distinct stimuli. Finally, we show that the mito-QC Counter can be used to quantify mitophagy in tissues of mito-QC transgenic mice. We demonstrate that mitophagy levels in skeletal muscle correlates with glycolytic activity. Our present data show that the mito-QC Counter macro for FIJI enables the robust quantification of mitophagy both in vitro and in vivo.
Original languageEnglish
Article number111196
Number of pages12
JournalMechanisms of Ageing and Development
Volume185
Early online date13 Dec 2019
DOIs
Publication statusPublished - Jan 2020

Keywords

  • mitophagy
  • mitochondria
  • autophagy
  • FIJI
  • mito-QC
  • mitolysosome
  • Mitochondria
  • Mitolysosome
  • Mitophagy
  • Autophagy

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