Katayama, H. et al. Cell 181, 1176–1187.e16 (2020).
Dysfunctional mitochondria are a hallmark of many diseases, but methods to study mitophagy, the process by which lysosomes remove these mitochondria, are limited. Katayama et al. have developed an improved indicator for mitophagy. They first engineered a cyan fluorescent protein called TOLLES (Tolerance of Lysosomal Environments) that is resistant to both acidic environments and proteases present in the lysosome. They then fused TOLLES to the yellow fluorescent protein YPet. This construct loses its yellow fluorescence in the lysosomal environment, as YPet is both acid- and protease-sensitive. The retained cyan fluorescence can be observed in fixed and living cells as a readout for autophagy. This probe, which the researchers call signal-retaining autophagy indicator (SRAI), was then specifically targeted to generate the mitophagy probe mito-SRAI. The researchers used mito-SRAI in a screen for compounds that induce mitophagy and to study mitophagy in neurons in a mouse model of Parkinson’s disease.