Description
Basal mitophagy is widespread in Drosophila but minimally affected by loss of Pink1 or parkin
Juliette J. Lee1, Alvaro Sanchez-Martinez1, Aitor Martinez Zarate2,3, Cristiane Benincá1, Ugo Mayor2, Michael J. Clague3, Alexander J. Whitworth1, 4
1MRC Mitochondrial Biology Unit, University of Cambridge, Cambridge Biomedical Campus, Hills Road, Cambridge CB2 0XY, United Kingdom; 2Department of Biochemistry and Molecular Biology UPV/EHU, University of the Basque Country, 48940, Leioa - Bizkaia, Spain; Department of Molecular and Cellular Physiology, Institute of Translational medicine, University of Liverpool, Crown St., Liverpool, L69 3BX, United Kingdom
Mitochondria are essential organelles that perform many critical metabolic functions but are also a major source of damaging reactive oxygen species (ROS) and harbour pro-apoptotic factors. Multiple homeostatic processes operate to maintain mitochondrial integrity, however, terminally damaged organelles are degraded through the process of targeted mitochondrial autophagy (mitophagy) to prevent potentially catastrophic consequences. Such homeostatic mechanisms are particularly important for post-mitotic, energetically demanding tissues such as nerves and muscles. There is increasing evidence that failure of this mechanism is linked to normal ageing and some neurodegenerative disorders. Interestingly, two proteins linked to Parkinson's Disease (PD), Parkin, a cytosolic ubiquitin ligase, and PINK1, a mitochondrially targeted kinase, have been shown to play key roles in this mitophagy. However, little is known about their impact on basal mitophagy in vivo.
We generated transgenic Drosophila expressing fluorescent mitophagy reporters, the mt-Keima and the mito-QC, to evaluate the impact of Pink1/parkin mutations on basal mitophagy under physiological conditions. The mt-Keima construct is a mitochondrial matrix-targeted pH-sensitive variant of GFP, while the mito-QC encodes a tandem GFP-mCherry fusion protein targeted to the outer mitochondrial membrane. Both systems exploit the pH-sensitive properties of mKeima and GFP to enable differential labelling of mitochondria in the acidic microenvironment of the lysosome as a proxy end-point readout of mitophagy.
We find that mitophagy is readily detectable and abundant in many tissues including Parkinson's Disease relevant dopaminergic neurons. However, we did not detect mitolysosomes in flight muscle. Surprisingly, in Pink1 or parkin null flies we did not observe any substantial impact on basal mitophagy. As these flies exhibit locomotor defects and dopaminergic neuron loss, our findings raise questions about current assumptions of the pathogenic mechanism associated with the PINK1/Parkin pathway. Our findings provide evidence that PINK1 and Parkin are not essential for bulk basal mitophagy in Drosophila. They also emphasize that mechanisms underpinning basal mitophagy remain largely obscure.