Fig. 4: Activation of the MitoSR is specific to mitochondrial damage in neurons and targets negative regulators of autophagy.

A–D Concerted degradation of MTMR5, MTMR2, and Rubicon is mediated by diverse triggers of mitophagy. A Representative western blot from WT cortical neuronal lysates treated with vehicle (EtOH, DMSO), Ant A, Oligo A, or CCCP for 2 hrs (concentration of drugs as indicated in the image). B MTMR5 levels normalized to total protein upon treatment of WT neurons with vehicle, Ant A, Oligo A, or CCCP for 2 hrs. C Ratio of intact MTMR2 normalized to total (intact + degraded) upon treatment of WT neurons with vehicle, Ant A, Oligo A, or CCCP for 2 hrs. D Rubicon band intensity normalized to total protein upon treatment of WT cortical neurons with vehicle, Ant A, Oligo A, or CCCP for 2 hrs (A–D: N = 4 experiments, one-way ANOVA with Dunnett’s multiple comparison test). E–H Lysosomal damage does not trigger the degradation of the negative regulators of autophagy. E Representative western blot from lysates of WT murine embryonic cortical neurons treated with vehicle (EtOH) or 1 mM LLOMe for 2 hrs. F Fold change in MTMR5 levels upon treatment of WT neurons with LLOMe as compared to with vehicle. G Fold change in MTMR2 levels upon treatment of WT neurons with LLOMe as compared to with vehicle. H Fold change in Rubicon levels upon treatment of WT neurons with LLOMe as compared to with vehicle (E–H: N = 3 experiments, Mann–Whitney test). I–M Damage to the endoplasmic reticulum does not trigger the degradation of the negative regulators of autophagy. I Representative western blot from lysates of WT murine embryonic cortical neurons treated with vehicle (DMSO) or 50 nM Tunicamycin (TM) for 3 hrs. J Fold change in ATF4 levels upon treatment of WT neurons with TM as compared to with vehicle. K Fold change in MTMR5 levels upon treatment of WT neurons with TM as compared to with vehicle. L Fold change in MTMR2 levels upon treatment of WT neurons with TM as compared to with vehicle. M Fold change in Rubicon levels upon treatment of WT neurons with TM as compared to with vehicle (I–M: N = 4 experiments, Mann–Whitney test). N–Q MitoSR is not triggered by mitochondrial damage in astrocytes. N Representative western blot from lysates of WT murine astrocytes treated with vehicle (EtOH, DMSO) or 5 μM Ant A, 10 μM Oligo A for 6 hrs. O Fold change in MFN-2 levels upon treatment of WT astrocytes with 5 μM Ant A, 10 μM Oligo A as compared to with vehicle. P Fold change in MTMR2 levels upon treatment of WT astrocytes with 5 μM Ant A, 10 μM Oligo A as compared to with vehicle. Q Fold change in Rubicon levels upon treatment of WT astrocytes with 5 μM Ant A, 10 μM Oligo A as compared to with vehicle (N–Q: N = 4 experiments, Mann–Whitney test). Error bars indicate SEM. Source data are provided as a Source Data file.