The 18-kDa TSPO (translocator protein) localizes on the outer mitochondrial membrane (OMM) and participates in cholesterol transport. Here, we report that TSPO inhibits mitochondrial autophagy downstream of the PINK1-PARK2 pathway, preventing essential ubiquitination of proteins. TSPO abolishes mitochondrial relocation of SQSTM1/p62 (sequestosome 1), and consequently that of the autophagic marker LC3 (microtubule-associated protein 1 light chain 3), thus leading to an accumulation of dysfunctional mitochondria, altering the appearance of the network. Independent of cholesterol regulation, the modulation of mitophagy by TSPO is instead dependent on VDAC1 (voltagedependent anion channel 1), to which TSPO binds, reducing mitochondrial coupling and promoting an overproduction of reactive oxygen species (ROS) that counteracts PARK2-mediated ubiquitination of proteins. These data identify TSPO as a novel element in the regulation of mitochondrial quality control by autophagy, and demonstrate the importance for cell homeostasis of its expression ratio with VDAC1.

SummaryMitophagy is central to mitochondrial and cellular homeostasis and operates via the PINK1/Parkin pathway targeting mitochondria devoid of membrane potential (ΔΨm) to autophagosomes. Although mitophagy is recognized as a fundamental cellular process, selective pharmacologic modulators of mitophagy are almost nonexistent. We developed a compound that increases the expression and signaling of the autophagic adaptor molecule P62/SQSTM1 and forces mitochondria into autophagy. The compound, P62-mediated mitophagy inducer (PMI), activates mitophagy without recruiting Parkin or collapsing ΔΨm and retains activity in cells devoid of a fully functional PINK1/Parkin pathway. PMI drives mitochondria to a process of quality control without compromising the bio-energetic competence of the whole network while exposing just those organelles to be recycled. Thus, PMI circumvents the toxicity and some of the nonspecific effects associated with the abrupt dissipation of ΔΨm by ionophores routinely used to induce mitophagy and represents a prototype pharmacological tool to investigate the molecular mechanisms of mitophagy.

Mitochondria drive cellular adaptation to stress by retro-communicating with the nucleus. This process is known as mitochondrial retrograde response (MRR) and is induced by mitochondrial dysfunction. MRR results in the nuclear stabilization of prosurvival transcription factors such as the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB). Here, we demonstrate that MRR is facilitated by contact sites between mitochondria and the nucleus. The translocator protein (TSPO) by preventing the mitophagy-mediated segregation o mitochonria is required for this interaction. The complex formed by TSPO with the protein kinase A (PKA), via the A-kinase anchoring protein acyl-CoA binding domain containing 3 (ACBD3), established the tethering. The latter allows for cholesterol redistribution of cholesterol in the nucleus to sustain the prosurvival response by blocking NF-κB deacetylation. This work proposes a previously unidentified paradigm in MRR: the formation of contact sites between mitochondria and nucleus to aid communication.