Molecular Regulation of Mitochondrial Biogenesis by Mitophagy Mediated Inhibition of PARIS

Mitochondrial volume and quality are tightly regulated by two key mechanisms: removal of dysfunctional/damaged mitochondria by selective autophagy (mitophagy) and mitochondrial biogenesis. Disruption of either one or both processes results in the accumulation of dysfunctional mitochondria and increased oxidative stress. Emerging evidence suggest that mitophagy and mitochondrial biogenesis are tightly coordinated, working in concert to maintain mitochondrial homeostasis and quality control. Previously, we reported that tumor necrosis factor α (TNFα), a proinflammatory cytokine produced during inflammatory airway disease, promotes mitochondrial biogenesis by enhancing peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) signaling, the nodal regulator of mitochondrial biogenesis, thereby increasing mitochondrial volume density in human airway smooth muscle (hASM) cells. Additional evidence indicates that TNFα induces mitochondrial depolarization, which is known to trigger a mitophagy pathway involving PTEN-induced putative kinase 1 (PINK1) and Parkin, an E3 ubiquitin ligase. Mitochondrial depolarization facilitates the mitochondrial accumulation of PINK1 and its activation by autophosphorylation at ser228 (pPINK1 s228 ). pPINK1 s228 promotes the recruitment and phosphorylation of Parkin at ser65 (pParkin s65 ), which ubiquitylates several mitochondrial associated proteins promoting the removal of damaged mitochondria. The decrease in the number of functional mitochondria by mitophagy can be offset by increasing mitochondrial biogenesis through the upregulation of PGC-1α signaling. Therefore, we hypothesize that PARIS (ZNF746), a transcriptional repressor of PGC-1α, is a target of PINK1/Parkin-mediated ubiquitination and degradation in hASM cells. In the present study, primary hASM cells were isolated from bronchiolar tissue of 6 subjects with no history of chronic pulmonary disease or smoking and separated into the following treatment groups: 1) FCCP (1 µM, 6 h; positive control), a proton ionophore that induces mitochondrial depolarization; 2) TNFα (20 ng/mL, 6 h); and 3) untreated control. We observed that FCCP and TNFα-induced mitochondrial depolarization triggered mitochondrial translocation of PINK1 and increased phosphorylation of pPINK1 s228 . Additionally, phosphorylation of pParkin s65 and ubiquitination of several mitochondrial associated proteins was increased. Immunoprecipitation of ubiquitinated proteins revealed increased ubiquitination of PARIS, while total expression of PARIS was decreased. Mitochondria and lysosomes were labelled with MitoTracker green (200 nM) and LysoTracker red (100 nM), respectively, and imaged in 3D (Z optical slice of 0.5 μm) to demonstrate the coefficient for colocalization of mitochondria with lysosomes. We observed increased formation of mitophagosomes, characteristic of mitophagy. Our findings demonstrate that TNFα-induces both mitophagy and mitochondrial biogenesis via the activation of PINK1/Parkin-mediated mitophagy pathway. Such coordination of mitophagy and mitochondrial biogenesis has significant ramifications for the maintenance of mitochondrial homeostasis, quality control and cellular energy metabolism. Supported by NIH grant HL157984 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.