The Many Faces of Mitochondrial Autophagy: Making Sense of Contrasting Observations in Recent Research
Figure 3
Biochemical probes for monitoring mitophagy in yeast. The localisation of probes within the different compartments of the mitochondrion is shown. A–G are fluorescence-based probes, while H is an enzymatic approach. (A) OM45-GFP is expressed from a chromosomal location in fusion with the endogenous OM protein OM45. GFP is exposed to the cytosol. (B) mt-Rosella II is an nonoligomerising biosensor comprising a red FP and pH-sensitive GFP expressed as a fusion to ATP3 from a genomic location (Lucarelli, May, Devenish and Prescott; unpublished). (C–H) Plasmid-derived combinations of FPs are targeted to the matrix space using different targeting sequences (TS) as follows: (C) isocitrate dehydrogenase, (D–E) F0 ATP synthase subunit c and (F–G) citrate synthase. (H) mito-Pho8 is a an acid phosphatase that is only active at vacuolar pH. When targeted to the matrix by a COXIV TS, the enzymatic activity provides a measure of mitophagy in strains disrupted for the endogenous PHO8 and PHO13 genes. Alternative targeting sequences allow targeting of probes to different compartments. OM = mitochondrial outer membrane, IMS = intermembrane space, IM = mitochondrial inner membrane, TS = targeting sequence.
