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Synaptic damage is a crucial pathological process in traumatic brain injury. However, the mechanisms driving this process remain poorly understood. In this report, we demonstrate that the accumulation of damaged mitochondria, resulting from impaired mitphagy, plays a significant role in causing synaptic damage. Moreover, copper induced downregulation of BNIP3 is a key player in regulating mitophagy. DMSA alleviates synaptic damage and mitochondrial dysfunction by promoting urinary excretion of copper. Mechanistically, we find that copper downregulate BNIP3 by increasing the nuclear translocation of NFKB, which is triggered by TRIM25-mediated ubiquitination-dependent degradation of NFKBIA. Our study underscores the importance of copper accumulation in the regulation of BNIP3-mediated mitophagy and suggests that therapeutic targeting of the copper-TRIM25-NFKB-BNIP3 axis holds promise to attenuate synaptic damage after traumatic brain injury. [ABSTRACT FROM AUTHOR]

Skeletal muscle plays a crucial role in generating force to facilitate movement. Skeletal muscle is a heterogenous tissue composed of diverse fibers with distinct contractile and metabolic profiles. The intricate classification of skeletal muscle fibers exists on a continuum ranging from type I (slow-twitch, oxidative) to type II (fast-twitch, glycolytic). The heterogenous distribution and characteristics of fibers within and between skeletal muscles profoundly influences cellular signaling; however, this has not been broadly discussed as it relates to macroautophagy/autophagy. The growing interest in skeletal muscle autophagy research underscores the necessity of comprehending the interplay between autophagic responses among skeletal muscles and fibers with different contractile properties, metabolic profiles, and other related signaling processes. We recommend approaching the interpretation of autophagy findings with careful consideration for two key reasons: 1) the distinct behaviors and responses of different skeletal muscles or fibers to various perturbations, and 2) the potential impact of alterations in skeletal muscle fiber type or metabolic profile on observed autophagic outcomes. This review provides an overview of the autophagic profile and response in skeletal muscles/fibers of different types and metabolic profiles. Further, this review discusses autophagic findings in various conditions and diseases that may differentially affect skeletal muscle. Finally, we provide key points of consideration to better enable researchers to fine-tune the design and interpretation of skeletal muscle autophagy experiments. Abbreviation: AKT1: AKT serine/threonine kinase 1; AMPK: AMP-activated protein kinase; ATG: autophagy related; ATG4: autophagy related 4 cysteine peptidase; ATG5: autophagy related 5; ATG7: autophagy related 7; ATG12: autophagy related 12; BECN1: beclin 1; BNIP3: BCL2 interacting protein 3; CKD: chronic kidney disease; COPD: chronic obstructive pulmonary disease; CS: citrate synthase; DIA: diaphragm; EDL: extensor digitorum longus; FOXO3/FOXO3A: forkhead box O3; GAS; gastrocnemius; GP: gastrocnemius-plantaris complex; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAPK: mitogen-activated protein kinase; MYH: myosin heavy chain; PINK1: PTEN induced kinase 1; PLANT: plantaris; PRKN: parkin RBR E3 ubiquitin protein ligase; QUAD: quadriceps; RA: rectus abdominis; RG: red gastrocnemius; RQ: red quadriceps; SOL: soleus; SQSTM1: sequestosome 1; TA: tibialis anterior; WG: white gastrocnemius; WQ: white quadriceps; WVL: white vastus lateralis; VL: vastus lateralis; ULK1: unc-51 like autophagy activating kinase 1. [ABSTRACT FROM AUTHOR]

Mitochondria undergo fission and fusion, and their coordinated balance is crucial for maintaining mitochondrial homeostasis. In yeast, the dynamin-related protein Dnm1 is a mitochondrial fission factor acting from outside the mitochondria. We recently reported the mitochondrial intermembrane space protein Atg44/mitofissin/Mdi1/Mco8 as a novel fission factor, but the relationship between Atg44 and Dnm1 remains elusive. Here, we show that Atg44 is required to complete Dnm1-mediated mitochondrial fission under homeostatic conditions. Atg44-deficient cells often exhibit enlarged mitochondria with accumulated Dnm1 and rosary-like mitochondria with Dnm1 foci at constriction sites. These mitochondrial constriction sites retain the continuity of both the outer and inner membranes within an extremely confined space, indicating that Dnm1 is unable to complete mitochondrial fission without Atg44. Moreover, accumulated Atg44 proteins are observed at mitochondrial constriction sites. These findings suggest that Atg44 and Dnm1 cooperatively execute mitochondrial fission from inside and outside the mitochondria, respectively. Abbreviation: ATG: autophagy related; CLEM: correlative light and electron microscopy; EM: electron microscopy; ER: endoplasmic reticulum; ERMES: endoplasmic reticulum-mitochondria encounter structure; GA: glutaraldehyde; GFP: green fluorescent protein; GTP: guanosine triphosphate: IMM: inner mitochondrial membrane; IMS: intermembrane space; OMM: outer mitochondrial membrane; PB: phosphate buffer; PBS: phosphate-buffered saline; PFA: paraformaldehyde; RFP: red fluorescent protein; WT: wild type. [ABSTRACT FROM AUTHOR]

Regressing the accelerated degradation of skeletal muscle protein is a significant goal for cancer cachexia management. Here, we show that genetic deletion of Pgam5 ameliorates skeletal muscle atrophy in various tumor-bearing mice. pgam5 ablation represses excessive myoblast mitophagy and effectively suppresses mitochondria meltdown and muscle wastage. Next, we define BNIP3 as a mitophagy receptor constitutively associating with PGAM5. bnip3 deletion restricts body weight loss and enhances the gastrocnemius mass index in the age- and tumor size-matched experiments. The NH2-terminal region of PGAM5 binds to the PEST motif-containing region of BNIP3 to dampen the ubiquitination and degradation of BNIP3 to maintain continuous mitophagy. Finally, we identify S100A9 as a pro-cachectic chemokine via activating AGER/RAGE. AGER deficiency or S100A9 inhibition restrains skeletal muscle loss by weakening the interaction between PGAM5 and BNIP3. In conclusion, the AGER-PGAM5-BNIP3 axis is a novel but common pathway in cancer-associated muscle wasting that can be targetable. Abbreviation: AGER/RAGE: advanced glycation end-product specific receptor; BA1: bafilomycin A1; BNIP3: BCL2 interacting protein 3; BNIP3L: BCL2 interacting protein 3 like; Ckm-Cre: creatinine kinase, muscle-specific Cre; CM: conditioned medium; CON/CTRL: control; CRC: colorectal cancer; FUNDC1: FUN14 domain containing 1; MAP1LC3A/LC3A: microtubule associated protein 1 light chain 3 alpha; PGAM5: PGAM family member 5, mitochondrial serine/threonine protein phosphatase; S100A9: S100 calcium binding protein A9; SQSTM1/p62: sequestosome 1; TOMM20: translocase of outer mitochondrial membrane 20; TIMM23: translocase of inner mitochondrial membrane 23; TSKO: tissue-specific knockout; VDAC1: voltage dependent anion channel 1. [ABSTRACT FROM AUTHOR]

Epidemiology has shown a strong relationship between fine particulate matter (PM) exposure and cardiovascular disease. However, it remains unknown whether PM aggravates myocardial ischemia-reperfusion (I/R) injury, and the related mechanisms are unclear. Our previous study has shown that adipose stem cell-derived exosomes (ADSC-Exos) contain high levels of Mir221 and Mir222. The present study investigated the effects of PM exposure on I/R-induced cardiac injury through mitophagy and apoptosis, as well as the potential role of Mir221 and Mir222 in ADSC-Exos. Wild-type, mir221- and mir222-knockout (KO), and Mir221- and Mir222-overexpressing transgenic (TG) mice were intratracheally injected with PM (10 mg/kg). After 24 h, mice underwent left coronary artery ligation for 30 min, followed by 3 h of reperfusion (I/R). H9c2 cardiomyocytes were cultured under 1% O2 for 6 h, then reoxygenated for 12 h (hypoxia-reoxygenation [H/R]). PM aggravated I/R (or H/R) cardiac injury by increasing ROS levels and causing mitochondrial dysfunction, which increased the expression of mitochondrial fission-related proteins (DNM1L/Drp1 and MFF) and mitophagy-related proteins (BNIP3 and MAP1LC3B/LC3B) in vivo and in vitro. Treatment with ADSC-Exos or Mir221- and Mir222-mimics significantly reduced PM+I/R-induced cardiac injury. Importantly, ADSC-Exos contain Mir221 and Mir222, which directly targets BNIP3, MAP1LC3B/LC3B, and BBC3/PUMA, decreasing their expression and ultimately reducing cardiomyocyte mitophagy and apoptosis. The present data showed that ADSC-Exos treatment regulated mitophagy and apoptosis through the Mir221 and Mir222-BNIP3-MAP1LC3B-BBC3/PUMA pathway and significantly reduced the cardiac damage caused by PM+I/R. The present study revealed the novel therapeutic potential of ADSC-Exos in alleviating PM-induced exacerbation of myocardial I/R injury.Abbreviation: ADSC-Exos: adipose-derived stem cell exosomes; AL: autolysosome; ATP: adenosine triphosphate; BBC3/PUMA: BCL2 binding component 3; BNIP3: BCL2/adenovirus E1B interacting protein 3; CASP3: caspase 3; CASP9: caspase 9; CDKN1B/p27: cyclin dependent kinase inhibitor 1B; CVD: cardiovascular disease; DCFH-DA: 2‘,7’-dichlorodihydrofluorescein diacetate; DHE: dihydroethidium; DNM1L/Drp1: dynamin 1-like; EF: ejection fraction; FS: fractional shortening; H/R: hypoxia-reoxygenation; I/R: ischemia-reperfusion; LDH: lactate dehydrogenase; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; MFF: mitochondrial fission factor; miRNA: microRNA; NAC: N-acetylcysteine; OCR: oxygen consumption rate; PIK3C3/Vps34: phosphatidylinositol 3-kinase catalytic subunit type 3; PM: particulate matter; PRKAA1/AMPK: protein kinase AMP-activated catalytic subunit alpha 1; ROS: reactive oxygen species; SQSTM1/p62: sequestosome 1; TEM: transmission electron microscopy; TRP53/p53: transformation related protein 53; TUNEL: terminal deoxynucleotidyl transferase dUTP nick end labeling. [ABSTRACT FROM AUTHOR]

Macroautophagy/autophagy is a constitutively active catabolic lysosomal degradation pathway, often found dysregulated in human diseases. It is often considered to act in a cytoprotective manner and is commonly upregulated in cells undergoing stress. Its initiation is regulated at the protein level and does not require de novo protein synthesis. Historically, autophagy has been regarded as nonselective; however, it is now clear that different stimuli can lead to the selective degradation of cellular components via selective autophagy receptors (SARs). Due to its selective nature and the existence of multiple degradation pathways potentially acting in concert, monitoring of autophagy flux, i.e. selective autophagy-dependent protein degradation, should address this complexity. Here, we introduce a targeted proteomics approach monitoring abundance changes of 37 autophagy-related proteins covering process-relevant proteins such as the initiation complex and the Atg8-family protein lipidation machinery, as well as most known SARs. We show that proteins involved in autophagosome biogenesis are upregulated and spared from degradation under autophagy-inducing conditions in contrast to SARs, in a cell-line dependent manner. Classical bulk stimuli such as nutrient starvation mainly induce degradation of ubiquitin-dependent soluble SARs and not of ubiquitin-independent, membrane-bound SARs. In contrast, treatment with the iron chelator deferiprone leads to the degradation of ubiquitin-dependent and -independent SARs linked to mitophagy and reticulophagy/ER-phagy. Our approach is automatable and supports large-scale screening assays paving the way to (pre)clinical applications and monitoring of specific autophagy flux.Abbreviation: AMBRA1: autophagy and beclin 1 regulator 1; ATG: autophagy related; BafA1: bafilomycin A1; BNIP1: BCL2 interacting protein 1; BNIP3: BCL2 interacting protein 3; BNIP3L/NIX: BCL2 interacting protein 3-like; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CCPG1: cell cycle progression 1; CV: coefficients of variations; CCCP: carbonyl cyanide m-chlorophenyl hydrazone; DFP: deferiprone; ER: endoplasmic reticulum; FKBP8: FKBP prolyl isomerase 8; GABARAPL: GABA type A receptor associated protein like; LC: liquid chromatography; LOD: limit of detection; LOQ: limit of quantification; MAP1LC3: microtubule associated protein 1 light chain 3; MS: mass spectrometry; NCOA4: nuclear receptor coactivator 4; NBR1: NBR1 autophagy cargo receptor; NUFIP1: nuclear FMR1 interacting protein 1; OPTN: optineurin; PHB2: prohibitin 2; PNPLA2/ATGL: patatin like phospholipase domain containing 2; POI: protein of interest; PTM: posttranslational modification; PRM: parallel reaction monitoring; RB1CC1/FIP200: RB1 inducible coiled-coil 1; RETREG1/FAM134B: reticulophagy regulator 1; RPS6KB1: ribosomal protein S6 kinase B1; RTN3: reticulon 3; SARs: selective autophagy receptors; SQSTM1/p62: sequestosome 1; STBD1: starch binding domain 1; TAX1BP1: Tax1 binding protein 1; TFEB: transcription factor EB; TNIP1: TNFAIP3 interacting protein 1; TOLLIP: toll interacting protein; ULK1: unc-51 like autophagy activating kinase 1; WBP2: WW domain binding protein 2; WDFY3/Alfy: WD repeat and FYVE domain containing 3; WIPI2: WD repeat domain, phosphoinositide interacting 2. [ABSTRACT FROM AUTHOR]

Uveal melanoma (UM) is an aggressive intraocular malignancy derived from melanocytes in the uvea tract of the eye. Up to 50% of patients with UM develop distant metastases which is usually fatal within one year; preventing metastases is therefore essential. Metabolic reprogramming plays a critical role in UM progression and metastasis. However, the metabolic phenotype of UM cells in the hypoxic tumor is not well understood. Here, we report that hypoxia-induced BNIP3 reprograms tumor cell metabolism, promoting their survival and metastasis. In response to hypoxia, BNIP3-mediated mitophagy alleviates mitochondrial dysfunction and enhances mitochondrial oxidative phosphorylation (OXPHOS) while simultaneously reducing mitochondrial reactive oxygen species (mtROS) production. This, in turn, impairs HIF1A/HIF-1α protein stability and inhibits glycolysis. Inhibition of mitophagy significantly suppresses BNIP3-induced UM progression and metastasis in vitro and in vivo. Collectively, these observations demonstrate a novel mechanism whereby BNIP3 promotes UM metabolic reprogramming and malignant progression by mediating hypoxia-induced mitophagy and suggest that BNIP3 could be an important therapeutic target to prevent metastasis in patients with UM.Abbreviations: AOD: average optical density; BNIP3: BCL2/adenovirus E1B interacting protein 3; CQ: chloroquine; CoCl2: cobalt chloride; GEPIA: Gene Expression Profiling Interactive Analysis; HIF1A: hypoxia inducible factor 1, alpha subunit; IHC: immunohistochemistry; mtROS: mitochondrial reactive oxygen species; NAC: N-acetylcysteine; OCR: oxygen consumption rate; OXPHOS: oxidative phosphorylation; ROS: reactive oxygen species; TCGA: The Cancer Genome Atlas; UM: uveal melanoma. [ABSTRACT FROM AUTHOR]

The selective removal of dysfunctional mitochondria, a process termed mitophagy, is critical for cellular health and impairments have been linked to aging, Parkinson disease, and other neurodegenerative conditions. A central mitophagy pathway is orchestrated by the ubiquitin (Ub) kinase PINK1 together with the E3 Ub ligase PRKN/Parkin. The decoration of damaged mitochondrial domains with phosphorylated Ub (p-S65-Ub) mediates their elimination though the autophagy system. As such p-S65-Ub has emerged as a highly specific and quantitative marker of mitochondrial damage with significant disease relevance. Existing p-S65-Ub antibodies have been successfully employed as research tools in a range of applications including western blot, immunocytochemistry, immunohistochemistry, and enzyme-linked immunosorbent assay. However, physiological levels of p-S65-Ub in the absence of exogenous stress are very low, therefore difficult to detect and require reliable and ultrasensitive methods. Here we generated and characterized a collection of novel recombinant, rabbit monoclonal p-S65-Ub antibodies with high specificity and affinity in certain applications that allow the field to better understand the molecular mechanisms and disease relevance of PINK1-PRKN signaling. These antibodies may also serve as novel diagnostic or prognostic tools to monitor mitochondrial damage in various clinical and pathological specimens. Abbreviations: AD: Alzheimer disease; CCCP: carbonyl cyanide 3-chlorophenylhydrazone; ELISA: enzyme-linked immunosorbent assay; HEK293E cell: human embryonic kidney E cell; ICC: immunocytochemistry; IHC: immunohistochemistry: KO: knockout; LoB: limit of blank; LoD: limit of detection; LoQ: limit of quantification; MEF: mouse embryonic fibroblast; MSD: Meso Scale Discovery; n.s.: non-significant; nonTg: non-transgenic; PBMC: peripheral blood mononuclear cell; PD: Parkinson disease; p-S65-PRKN: phosphorylated PRKN at serine 65; p-S65-Ub: phosphorylated Ub at serine 65; Ub: ubiquitin; WT: wild-type. [ABSTRACT FROM AUTHOR]

Dominant variants in WFS1 (wolframin ER transmembrane glycoprotein), the gene coding for a mitochondria-associated endoplasmic reticulum (ER) membrane (MAM) resident protein, have been associated with Wolfram-like syndrome (WLS). In vitro and in vivo, WFS1 loss results in reduced ER to mitochondria calcium (Ca2+) transfer, mitochondrial dysfunction, and enhanced macroautophagy/autophagy and mitophagy. However, in the WLS pathological context, whether the mutant protein triggers the same cellular processes is unknown. Here, we show that in human fibroblasts and murine neuronal cultures the WLS protein WFS1E864K leads to decreases in mitochondria bioenergetics and Ca2+ uptake, deregulation of the mitochondrial quality system mechanisms, and alteration of the autophagic flux. Moreover, in the Wfs1E864K mouse, these alterations are concomitant with a decrease of MAM number. These findings reveal pathophysiological similarities between WS and WLS, highlighting the importance of WFS1 for MAM's integrity and functionality. It may open new treatment perspectives for patients with WLS. Abbreviations: BafA1: bafilomycin A1; ER: endoplasmic reticulum; HSPA9/GRP75: heat shock protein family A (Hsp70) member 9; ITPR/IP3R: inositol 1,4,5-trisphosphate receptor; MAM: mitochondria-associated endoplasmic reticulum membrane; MCU: mitochondrial calcium uniporter; MFN2: mitofusin 2; OCR: oxygen consumption rate; ROS: reactive oxygen species; ROT/AA: rotenone+antimycin A; VDAC1: voltage dependent anion channel 1; WLS: Wolfram-like syndrome; WS: Wolfram syndrome; WT: wild-type [ABSTRACT FROM AUTHOR]

During tumor expansion, breast cancer (BC) cells often experience reactive oxygen species accumulation and mitochondrial damage because of glucose shortage. However, the mechanism by which BC cells deal with the glucose-shortage-induced oxidative stress remains unclear. Here, we showed that MANF (mesencephalic astrocyte derived neurotrophic factor)-mediated mitophagy facilitates BC cell survival under glucose-starvation conditions. MANF-mediated mitophagy also promotes fatty acid oxidation in glucose-starved BC cells. Moreover, during glucose starvation, SENP1-mediated de-SUMOylation of MANF increases cytoplasmic MANF expression through the inhibition of MANF’s nuclear translocation and hence renders mitochondrial distribution of MANF. MANF mediates mitophagy by binding to PRKN (parkin RBR E3 ubiquitin protein ligase), a key mitophagy regulator, in the mitochondria. Under conditions of glucose starvation, protein oxidation inhibits PRKN activity; nevertheless, the CXXC motif of MANF alleviates protein oxidation in RING II-domain of PRKN and restores its E3 ligase activity. Furthermore, MANF-PRKN interactions are essential for BC tumor growth and metastasis. High MANF expression predicts poor outcomes in patients with BC. Our results highlight the prosurvival role of MANF-mediated mitophagy in BC cells during glucose starvation, suggesting MANF as a potential therapeutic target.Abbreviation: 2DG, 2-deoxy-D-glucose; 5TG, 5-thio-D-glucose; ACSL4/FACL4, acyl-CoA synthetase long chain family member 4; Baf A1, bafilomycin A1; BRCA, breast cancer; CHX, cycloheximide; DMF, distant metastasis-free; DMFS, distant metastasis-free survival; ECM, extracellular matrix; ER, endoplasmic reticulum; ERS, endoplasmic reticulum stress; F-1,6-BP, fructose-1,6-bisphosphate; FAO, fatty acid oxidation; GSH, reduced glutathione; GSVA, gene set variation analysis; HCC, hepatocellular carcinoma; ICC, intrahepatic cholangiocarcinoma; IF, immunofluorescence; MANF, mesencephalic astrocyte derived neurotrophic factor; Mdivi-1, mitochondrial division inhibitor 1; MFI, mean fluorescence intensity; NAC, N-acetyl-L-cysteine; OCR, oxygen-consumption rate; OS, overall survival; PMI, SQSTM1/p62-mediated mitophagy inducer; PPP, pentose phosphate pathway; PRKN, parkin RBR E3 ubiquitin protein ligase; RBR, RING in between RING; RFS, relapse-free survival; ROS, reactive oxygen species; SAPLIPs, saposin-like proteins; TCGA, The Cancer Genome Atlas; TNBC, triple-negative breast cancer; WT, wild type. [ABSTRACT FROM AUTHOR]

Severe fever with thrombocytopenia syndrome is an emerging viral hemorrhagic fever caused by a tick-borne bunyavirus, severe fever with thrombocytopenia syndrome virus (SFTSV), with a high case fatality. We previously found that SFTSV nucleoprotein (NP) induces macroautophagy/autophagy to facilitate virus replication. However, the role of NP in antagonizing host innate immunity remains unclear. Mitophagy, a selected form of autophagy, eliminates damaged mitochondria to maintain mitochondrial homeostasis. Here, we demonstrate that SFTSV NP triggers mitophagy to degrade MAVS (mitochondrial antiviral signaling protein), thereby blocking MAVS-mediated antiviral signaling to escape the host immune response. Mechanistically, SFTSV NP translocates to mitochondria by interacting with TUFM (Tu translation elongation factor, mitochondrial), and mediates mitochondrial sequestration into phagophores through interacting with LC3, thus inducing mitophagy. Notably, the N-terminal LC3-interacting region (LIR) motif of NP is essential for mitophagy induction. Collectively, our results demonstrated that SFTSV NP serves as a novel virulence factor, inducing TUFM-mediated mitophagy to degrade MAVS and evade the host immune response.Abbreviation: 3-MA: 3-methyladenine; ACTB: actin beta; co-IP: co-immunoprecipitation; CQ: chloroquine; DAPI: 4ʹ,6-diamidino-2-phenylindole, dihydrochloride; DMSO: dimethyl sulfoxide; FCCP: carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone; GFP: green fluorescent protein; HTNV: Hantan virus; IAV: influenza A virus; IFN: interferon; LAMP1: lysosomal associated membraneprotein 1; LIR: LC3-interacting region; MAP1LC3B/LC3B: microtubule associatedprotein 1 light chain 3 beta; MAVS: mitochondrial antiviral signaling protein; Mdivi-1: mitochondrial division inhibitor 1; MOI: multiplicity of infection; MT-CO2/COXII: mitochondrially encoded cytochrome C oxidase II; NP: nucleoprotein; NSs: nonstructural proteins; poly(I:C): polyinosinic:polycytidylic acid; RIGI: RNA sensor RIG-I; RLR: RIGI-like receptor; SFTSV: severe fever withthrombocytopenia syndrome virus; TCID50: 50% tissue culture infectiousdose; TIMM23: translocase of inner mitochondrial membrane 23; TOMM20:translocase of outer mitochondrial membrane 20; TUFM: Tu translation elongationfactor, mitochondrial. [ABSTRACT FROM AUTHOR]

Aging is a gradual and irreversible physiological process that significantly increases the risks of developing a variety of pathologies, including neurodegenerative, cardiovascular, metabolic, musculoskeletal, and immune system diseases. Mitochondria are the energy-producing organelles, and their proper functioning is crucial for overall cellular health. Over time, mitochondrial function declines causing an increased release of harmful reactive oxygen species (ROS) and DNA, which leads to oxidative stress, inflammation and cellular damage, common features associated with various age-related pathologies. The impairment of mitophagy, the selective removal of damaged or dysfunctional mitochondria by autophagy, is relevant to the development and progression of age-related diseases. The molecular mechanisms that regulates mitophagy levels in aging remain largely uncharacterized. AMBRA1 is an intrinsically disordered scaffold protein with a unique property of regulating the activity of both proliferation and autophagy core machineries. While the role of AMBRA1 during embryonic development and neoplastic transformation has been extensively investigated, its functions in post-mitotic cells of adult tissues have been limited due to the embryonic lethality caused by AMBRA1 deficiency. Recently, a key role of AMBRA1 in selectively regulating mitophagy in post-mitotic cells has emerged. Here we summarize and discuss these results with the aim of providing a comprehensive view of the mitochondrial roles of AMBRA1, and how defective activity of AMBRA1 has been functionally linked to mitophagy alterations observed in age-related degenerative disorders, including muscular dystrophy/sarcopenia, Parkinson diseases, Alzheimer diseases and age-related macular degeneration.Abbreviations: AD: Alzheimer disease; AMD: age‐related macular degeneration; AMBRA1: autophagy and beclin 1 regulator 1; APOE4: apolipoprotein E4; ATAD3A: ATPase family AAA domain containing 3A; ATG: autophagy related; BCL2: BCL2 apoptosis regulator; BH3: BCL2-homology-3; BNIP3L/NIX: BCL2 interacting protein 3 like; CDK: cyclin dependent kinase; CHUK/IKKα: component of inhibitor of nuclear factor kappa B kinase complex; CRL2: CUL2-RING ubiquitin ligase; DDB1: damage specific DNA binding protein 1; ER: endoplasmic reticulum; FOXO: forkhead box O; FUNDC1: FUN14 domain containing 1; GBA/β-glucocerebrosidase: glucosylceramidase beta; HUWE1: HECT, UBA and WWE domain containing E3 ubiquitin protein ligase 1; IDR: intrinsically disordered region; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAVS: mitochondrial antiviral signaling protein; MCL1: MCL1 apoptosis regulator, BCL2 family member; MFN2: mitofusin 2; MTOR: mechanistic target of rapamycin kinase; MSA: multiple system atrophy; MYC: MYC proto-oncogene, bHLH transcription factor; NUMA1: nuclear mitotic apparatus protein 1; OMM; mitochondria outer membrane; PD: Parkinson disease; PHB2: prohibitin 2; PINK1: PTEN induced kinase 1; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PTK2/FAK: protein tyrosine kinase 2; ROS: reactive oxygen species; RPE: retinal pigment epithelium; SAD: sporadic AD; SOCS3: suppressor of cytokine signaling 3; SRC, SRC proto-oncogene, non-receptor tyrosine kinase; STAT3: signal transducer and activator of transcription 3; STING1: stimulator of interferon response cGAMP interactor 1; SQSTM1/p62: sequestosome 1; TBK1: TANK binding kinase 1; TGFB/TGFβ: transforming growth factor beta; TOMM: translocase of outer mitochondrial membrane; TRAF6: TNF receptor associated factor 6; TRIM32: tripartite motif containing 32; ULK1: unc-51 like autophagy activating kinase 1. [ABSTRACT FROM AUTHOR]

Disruption of mitochondrial function is observed in multiple drug-induced liver injuries (DILIs), a significant global health threat. However, how the mitochondrial dysfunction occurs and whether maintain mitochondrial homeostasis is beneficial for DILIs remains unclear. Here, we show that defective mitophagy by OPTN (optineurin) ablation causes disrupted mitochondrial homeostasis and aggravates hepatocytes necrosis in DILIs, while OPTN overexpression protects against DILI depending on its mitophagic function. Notably, mass spectrometry analysis identifies a new mitochondrial substrate, GCDH (glutaryl-CoA dehydrogenase), which can be selectively recruited by OPTN for mitophagic degradation, and a new cofactor, VCP (valosin containing protein) that interacts with OPTN to stabilize BECN1 during phagophore assembly, thus boosting OPTN-mediated mitophagy initiation to clear damaged mitochondria and preserve mitochondrial homeostasis in DILIs. Then, the accumulation of OPTN in different DILIs is further validated with a protective effect, and pyridoxine is screened and established to alleviate DILIs by inducing OPTN-mediated mitophagy. Collectively, our findings uncover a dual role of OPTN in mitophagy initiation and implicate the preservation of mitochondrial homeostasis via inducing OPTN-mediated mitophagy as a potential therapeutic approach for DILIs.Abbreviation: AILI: acetaminophen-induced liver injury; ALS: amyotrophic lateral sclerosis; APAP: acetaminophen; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CHX: cycloheximide; Co-IP: co-immunoprecipitation; DILI: drug-induced liver injury; FL: full length; GCDH: glutaryl-CoA dehydrogenase; GOT1/AST: glutamic-oxaloacetic transaminase 1; GO: gene ontology; GSEA: gene set enrichment analysis; GPT/ALT: glutamic – pyruvic transaminase; INH: isoniazid; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MMP: mitochondrial membrane potential; MST: microscale thermophoresis; MT-CO2/COX-II: mitochondrially encoded cytochrome c oxidase II; OPTN: optineurin; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; TIMM23: translocase of inner mitochondrial membrane 23; TOMM20: translocase of outer mitochondrial membrane 20; TSN: toosendanin; VCP: valosin containing protein, WIPI2: WD repeat domain, phosphoinositide interacting 2. [ABSTRACT FROM AUTHOR]

Defective mitophagy is consistently found in postmortem brain and iPSC-derived neurons from Alzheimer disease (AD) patients. However, there is a lack of extensive examination of mitophagy status in serum or cerebrospinal fluid (CSF), and the clinical potential of mitophagy biomarkers has not been tested. We quantified biomarkers of mitophagy/autophagy and lysosomal degradation (PINK1, BNIP3L and TFEB) in CSF and serum from 246 individuals, covering mild cognitive impairment due to AD (MCI-AD, n = 100), dementia due to AD (AD-dementia, n = 100), and cognitively unimpaired individuals (CU, n = 46), recruited from the Czech Brain Aging Study. Cognitive function and brain atrophy were also assessed. Our data show that serum and CSF PINK1 and serum BNIP3L were higher, and serum TFEB was lower in individuals with AD than in corresponding CU individuals. Additionally, the magnitude of mitophagy impairment correlated with the severity of clinical indicators in AD patients. Specifically, levels of PINK1 positively correlated with phosphorylated (p)-MAPT/tau (181), total (t)-MAPT/tau, NEFL (neurofilament light chain), and NRGN (neurogranin) levels in CSF and negatively with memory, executive function, and language domain. Serum TFEB levels negatively correlated with NEFL and positively with executive function and language. This study reveals mitophagy impairment reflected in biofluid biomarkers of individuals with AD and associated with more advanced AD pathology. Abbreviation: Aβ: amyloid beta; AD: Alzheimer disease; AVs: autophagic vacuoles; BNIP3L: BCL2 interacting protein 3 like; CU: cognitively unimpaired; CSF: cerebrospinal fluid; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCI: mild cognitive impairment; NRGN: neurogranin; NEFL: neurofilament light chain; p-MAPT/tau: phosphorylated microtubule associated protein tau; PINK1: PTEN induced kinase 1; t-MAPT/tau: total microtubule associated protein tau; TFEB: transcription factor EB; TMT: Trail Making Test. [ABSTRACT FROM AUTHOR]

Sarcopenia is a major contributor to disability in older adults, and thus, it is key to elucidate the mechanisms underlying its development. Increasing evidence suggests that impaired macroautophagy/autophagy contributes to the development of sarcopenia. However, the mechanisms leading to reduced autophagy during aging remain largely unexplored, and whether autophagy activation protects from sarcopenia has not been fully addressed. Here we show that the autophagy regulator TP53INP2/TRP53INP2 is decreased during aging in mouse and human skeletal muscle. Importantly, chronic activation of autophagy by muscle-specific overexpression of TRP53INP2 prevents sarcopenia and the decline of muscle function in mice. Acute re-expression of TRP53INP2 in aged mice also improves muscle atrophy, enhances mitophagy, and reduces ROS production. In humans, high levels of TP53INP2 in muscle are associated with increased muscle strength and healthy aging. Our findings highlight the relevance of an active muscle autophagy in the maintenance of muscle mass and prevention of sarcopenia. Abbreviation: ATG7: autophagy related 7; BMI: body mass index; EIF4EBP1: eukaryotic translation initiation factor 4E binding protein 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; ROS: reactive oxygen species; TP53INP2: tumor protein p53 inducible nuclear protein 2; WT: wild type [ABSTRACT FROM AUTHOR]

Drug-tolerant persister (DTP) cancer cells drive residual tumor and relapse. However, the mechanisms underlying DTP state development are largely unexplored. In a recent study, we determined that PINK1-mediated mitophagy favors DTP generation in the context of MAPK inhibition therapy. DTP cells that persist in the presence of a MAPK inhibitor exhibit mitochondriadependent metabolism. During DTP state development, MYC depletion alleviates the transcriptional repression of PINK1, resulting in PINK1 upregulation and mitophagy activation. PINK1-mediated mitophagy is essential for mitochondrial homeostasis in DTP cells. Either knockdown of PINK1 or inhibition of mitophagy eradicates DTP cells and achieves complete responses to MAPK inhibition therapy. This study reveals a novel role of mitophagy as a protective mechanism for DTP development.

Spautin-1 is a well-known macroautophagy/autophagy inhibitor via suppressing the deubiquitinases USP10 and USP13 and promoting the degradation of the PIK3C3/VPS34-BECN1 complex, while its effect on selective autophagy remains poorly understood. Mitophagy is a selective form of autophagy for removal of damaged and superfluous mitochondria via the autophagy-lysosome pathway. Here, we report a surprising discovery that, while spautin-1 remains as an effective autophagy inhibitor, it promotes PINK1-PRKN-dependent mitophagy induced by mitochondrial damage agents. Mechanistically, spautin-1 facilitates the stabilization and activation of the full-length PINK1 at the outer mitochondrial membrane (OMM) via binding to components of the TOMM complex (TOMM70 and TOMM20), leading to the disruption of the mitochondrial import of PINK1 and prevention of PARL-mediated PINK1 cleavage. Moreover, spautin-1 induces neuronal mitophagy in Caenorhabditis elegans (C. elegans) in a PINK-1-PDR-1-dependent manner. Functionally, spautin-1 is capable of improving associative learning capability in an Alzheimer disease (AD) C. elegans model. In summary, we report a novel function of spautin-1 in promoting mitophagy via the PINK1-PRKN pathway. As deficiency of mitophagy is closely implicated in the pathogenesis of neurodegenerative disorders, the pro-mitophagy function of spautin-1 might suggest its therapeutic potential in neurodegenerative disorders such as AD.Abbreviations: AD, Alzheimer disease; ATG, autophagy related; BafA1, bafilomycin A1; CALCOCO2/NDP52, calcium binding and coiled-coil domain 2; CCCP, carbonyl cyanide m-chlorophenyl hydrazone; COX4/COX IV, cytochrome c oxidase subunit 4; EBSS, Earle’s balanced salt; ECAR, extracellular acidification rate; GFP, green fluorescent protein; IA, isoamyl alcohol; IMM, inner mitochondrial membrane; MAP1LC3/LC3, microtubule associated protein 1 light chain 3; MMP, mitochondrial membrane potential; mtDNA, mitochondrial DNA; nDNA, nuclear DNA; O/A, oligomycin-antimycin; OCR, oxygen consumption rate; OMM, outer mitochondrial membrane; OPTN, optineurin; PARL, presenilin associated rhomboid like; PINK1, PTEN induced kinase 1; PRKN, parkin RBR E3 ubiquitin protein ligase; p-Ser65-Ub, phosphorylation of Ub at Ser65; TIMM23, translocase of inner mitochondrial membrane 23; TOMM, translocase of outer mitochondrial membrane; USP10, ubiquitin specific peptidase 10; USP13, ubiquitin specific peptidase 13; VAL, valinomycin; YFP, yellow fluorescent protein. [ABSTRACT FROM AUTHOR]

Osteopenia and osteoporosis are among the most common metabolic bone diseases and represent major public health problems, with sufferers having an increased fracture risk. Diabetes is one of the most common diseases contributing to osteopenia and osteoporosis. However, the mechanisms underlying diabetes-induced osteopenia and osteoporosis remain unclear. Bone reconstruction, including bone formation and absorption, is a dynamic process. Large-conductance Ca2+-activated K+ channels (BK channels) regulate the function of bone marrow-derived mesenchymal stem cells, osteoblasts, and osteoclasts. Our previous studies revealed the relationship between BK channels and the function of osteoblasts via various pathways under physiological conditions. In this study, we reported a decrease in the expression of BK channels in mice with diabetes-induced osteopenia. BK deficiency enhanced mitochondrial Ca2+ and activated classical PINK1 (PTEN induced putative kinase 1)-PRKN/Parkin (parkin RBR E3 ubiquitin protein ligase)-dependent mitophagy, whereas the upregulation of BK channels inhibited mitophagy in osteoblasts. Moreover, SLC25A5/ANT2 (solute carrier family 25 (mitochondrial carrier, adenine nucleotide translocator), member 5), a critical inner mitochondrial membrane protein participating in PINK1-PRKN-dependent mitophagy, was also regulated by BK channels. Overall, these data identified a novel role of BK channels in regulating mitophagy in osteoblasts, which might be a potential target for diabetes-induced bone diseases.Abbreviations: AGE, advanced glycation end products; Baf A1, bafilomycin A1; BK channels, big-conductance Ca2+-activated K+ channels; BMSCs, bone marrow-derived mesenchymal stem cells; BSA, bovine serum albumin; FBG, fasting blood glucose; IMM, inner mitochondrial membrane; ITPR1, inositol 1,4,5-trisphosphate receptor 1; MAM, mitochondria-associated ER membrane; OMM, outer mitochondrial membrane; PINK1, PTEN induced putative kinase 1; PPID/CyP-D, peptidylprolyl isomerase D (cyclophilin D); PRKN/PARK2, parkin RBR E3 ubiquitin protein ligase; ROS, reactive oxygen species; SLC25A5/ANT2, solute carrier family 25 (mitochondrial carrier, adenine nucleotide translocator), member 5; STZ, streptozotocin. [ABSTRACT FROM AUTHOR]

Mitophagy involves the selective elimination of defective mitochondria during chemotherapeutic stress to maintain mitochondrial homeostasis and sustain cancer growth. Here, we showed that CLU (clusterin) is localized to mitochondria to induce mitophagy controlling mitochondrial damage in oral cancer cells. Moreover, overexpression and knockdown of CLU establish its mitophagy-specific role, where CLU acts as an adaptor protein that coordinately interacts with BAX and LC3 recruiting autophagic machinery around damaged mitochondria in response to cisplatin treatment. Interestingly, CLU triggers class III phosphatidylinositol 3-kinase (PtdIns3K) activity around damaged mitochondria, and inhibition of mitophagic flux causes the accumulation of excessive mitophagosomes resulting in reactive oxygen species (ROS)-dependent apoptosis during cisplatin treatment in oral cancer cells. In parallel, we determined that PPARGC1A/PGC1α (PPARG coactivator 1 alpha) activates mitochondrial biogenesis during CLU-induced mitophagy to maintain the mitochondrial pool. Intriguingly, PPARGC1A inhibition through small interfering RNA (siPPARGC1A) and pharmacological inhibitor (SR-18292) treatment counteracts CLU-dependent cytoprotection leading to mitophagy-associated cell death. Furthermore, co-treatment of SR-18292 with cisplatin synergistically suppresses tumor growth in oral cancer xenograft models. In conclusion, CLU and PPARGC1A are essential for sustained cancer cell growth by activating mitophagy and mitochondrial biogenesis, respectively, and their inhibition could provide better therapeutic benefits against oral cancer. [ABSTRACT FROM AUTHOR]

CARM1 (coactivator associated arginine methyltransferase 1) has recently emerged as a powerful regulator of skeletal muscle biology. However, the molecular mechanisms by which the methyltransferase remodels muscle remain to be fully understood. In this study, carm1 skeletal muscle-specific knockout (mKO) mice exhibited lower muscle mass with dysregulated macroautophagic/autophagic and atrophic signaling, including depressed AMP-activated protein kinase (AMPK) site-specific phosphorylation of ULK1 (unc-51 like autophagy activating kinase 1; Ser555) and FOXO3 (forkhead box O3; Ser588), as well as MTOR (mechanistic target of rapamycin kinase)-induced inhibition of ULK1 (Ser757), along with AKT/protein kinase B site-specific suppression of FOXO1 (Ser256) and FOXO3 (Ser253). In addition to lower mitophagy and autophagy flux in skeletal muscle, carm1 mKO led to increased mitochondrial PRKN/parkin accumulation, which suggests that CARM1 is required for basal mitochondrial turnover and autophagic clearance. carm1 deletion also elicited PPARGC1A (PPARG coactivator 1 alpha) activity and a slower, more oxidative muscle phenotype. As such, these carm1 mKO-evoked adaptations disrupted mitophagy and autophagy induction during food deprivation and collectively served to mitigate fasting-induced muscle atrophy. Furthermore, at the threshold of muscle atrophy during food deprivation experiments in humans, skeletal muscle CARM1 activity decreased similarly to our observations in mice, and was accompanied by site-specific activation of ULK1 (Ser757), highlighting the translational impact of the methyltransferase in human skeletal muscle. Taken together, our results indicate that CARM1 governs mitophagic, autophagic, and atrophic processes fundamental to the maintenance and remodeling of muscle mass. Targeting the enzyme may provide new therapeutic approaches for mitigating skeletal muscle atrophy. Abbreviation: ADMA: asymmetric dimethylarginine; AKT/protein kinase B: AKT serine/threonine kinase; AMPK: AMP-activated protein kinase; ATG: autophagy related; BECN1: beclin 1; BNIP3: BCL2 interacting protein 3; CARM1: coactivator associated arginine methyltransferase 1; Col: colchicine; CSA: cross-sectional area; CTNS: cystinosin, lysosomal cystine transporter; EDL: extensor digitorum longus; FBXO32/MAFbx: F-box protein 32; FOXO: forkhead box O; GAST: gastrocnemius; H2O2: hydrogen peroxide; IMF: intermyofibrillar; LAMP1: lysosomal associated membrane protein 1; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; mKO: skeletal muscle-specific knockout; MMA: monomethylarginine; MTOR: mechanistic target of rapamycin kinase; MYH: myosin heavy chain; NFE2L2/NRF2: NFE2 like bZIP transcription factor 2; OXPHOS: oxidative phosphorylation; PABPC1/PABP1: poly(A) binding protein cytoplasmic 1; PPARGC1A/PGC-1α: PPARG coactivator 1 alpha; PRKN/parkin: parkin RBR E3 ubiquitin protein ligase; PRMT: protein arginine methyltransferase; Sal: saline; SDMA: symmetric dimethylarginine; SIRT1: sirtuin 1; SKP2: S-phase kinase associated protein 2; SMARCC1/BAF155: SWI/SNF related, matrix associated, actin dependent regulator of chromatin subfamily c member 1; SOL: soleus; SQSTM1/p62: sequestosome 1; SS: subsarcolemmal; TA: tibialis anterior; TFAM: transcription factor A, mitochondrial; TFEB: transcription factor EB; TOMM20: translocase of outer mitochondrial membrane 20; TRIM63/MuRF1: tripartite motif containing 63; ULK1: unc-51 like autophagy activating kinase 1; VPS11: VPS11 core subunit of CORVET and HOPS complexes; WT: wild-type. [ABSTRACT FROM AUTHOR]

Senecavirus A (SVA) is a newly emerging picornavirus associated with swine vesicular lesions and neonatal mortality, threatening the global pig industry. Despite sustained efforts, the molecular mechanisms of SVA pathogenesis have not yet been fully elucidated. Here, we demonstrate for the first time that SVA infection can induce complete mitophagy in host cells, which depends on SVA replication. Mitophagy has been subsequently proven to promote SVA replication in host cells. Genome-wide screening of SVA proteins involved in inducing mitophagy showed that although VP2, VP3, 2C, and 3A proteins can independently induce mitophagy, only the 2C protein mediates mitophagy through direct interaction with TUFM (Tu translation elongation factor, mitochondrial). The glutamic acids at positions 196 and 211 of TUFM were shown to be two key sites for its interaction with 2C protein. Moreover, TUFM was discovered to interact directly with BECN1 and indirectly with the ATG12–ATG5 conjugate. Further experiments revealed that TUFM needs to undergo ubiquitination modification before being recognized by the macroautophagy/autophagy receptor protein SQSTM1/p62, and E3 ubiquitin ligase RNF185 catalyzes K27-linked polyubiquitination of TUFM through the interaction between RNF185's transmembrane domain 1 and TUFM to initiate SVA-induced mitophagy. The ubiquitinated TUFM is recognized and bound by SQSTM1, which in turn interacts with MAP1LC3/LC3, thereby linking the 2C-anchored mitochondria to the phagophore for sequestration into mitophagosomes, which ultimately fuse with lysosomes to achieve complete mitophagy. Overall, our results elucidated the molecular mechanism by which SVA induces mitophagy to promote self-replication and provide new insights into SVA pathogenesis.Abbreviations: aa: amino acid; Baf A1: bafilomycin A1; BHK-21: baby hamster kidney-21; CCCP: carbonyl cyanide m-chlorophenyl hydrazone; co-IP: co-immunoprecipitation; CQ: chloroquine; DAPI: 4',6-diamidino-2'-phenylindole; DMSO: dimethyl sulfoxide; EGFP: enhanced green fluorescent protein; ER: endoplasmic reticulum; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFP: green fluorescent protein; GST: glutathione S-transferase; HA: hemagglutinin; hpi: hours post-infection; hpt: hours post-transfection; IPTG: isopropyl β-D-1-thiogalactopyranoside; mAb: monoclonal antibody; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAVS: mitochondrial antiviral signaling protein; Mdivi-1: mitochondrial division inhibitor-1; MOI: multiplicity of infection; mRFP: monomeric red fluorescent protein; MS: mass spectrometry; ORF: open reading frame; PBS: phosphate-buffered saline; SD: standard deviation; SQSTM1/p62: sequestosome 1; ST: swine testis; SVA: Senecavirus A; TCID50: 50% tissue culture infectious dose; TIMM23: translocase of inner mitochondrial membrane 23; TM: transmembrane; TOMM20: translocase of outer mitochondrial membrane 20; TUFM: Tu translation elongation factor, mitochondrial; Ub: ubiquitin; UV: ultraviolet; VDAC1: voltage dependent anion channel 1; WT: wild-type; μg: microgram; μm: micrometer; μM: micromole. [ABSTRACT FROM AUTHOR]

Mitophagy is the process of selective autophagy that removes superfluous and dysfunctional mitochondria. Mitophagy was first characterized in mammalian cells and is now recognized to follow several pathways including basal forms in specific organs. Mitophagy pathways are regulated by multiple, often interconnected factors. The present review aims to streamline this complexity and evaluate common elements that may define the evolutionary origin of mitophagy. Key issues surrounding mitophagy signaling at the mitochondrial surface may fundamentally derive from mitochondrial membrane dynamics. Elements of such membrane dynamics likely originated during the endosymbiosis of the alphaproteobacterial ancestor of our mitochondria but underwent an evolutionary leap forward in basal metazoa that determined the currently known variations in mitophagy signaling. Abbreviations: AGPAT, 1-acylglycerol-3-phosphate O-acyltransferase; ATG, autophagy related; BCL2L13, BCL2 like 13; BNIP3, BCL2 interacting protein 3; BNIP3L, BCL2 interacting protein 3 like; CALCOCO, calcium binding and coiled-coil domain; CL, cardiolipin; ER, endoplasmic reticulum; ERMES, ER-mitochondria encounter structure; FBXL4, F-box and leucine rich repeat protein 4; FUNDC1, FUN14 domain containing 1; GABARAPL1, GABA type A receptor associated protein like 1; HIF, hypoxia inducible factor; IMM, inner mitochondrial membrane; LBPA/BMP, lysobisphosphatidic acid; LIR, LC3-interacting region; LPA, lysophosphatidic acid; MAM, mitochondria-associated membranes; MAP1LC3/LC3, microtubule associated protein 1 light chain 3; MCL, monolysocardiolipin; ML, maximum likelihood; NBR1, NBR1 autophagy cargo receptor; OMM, outer mitochondrial membrane; PA, phosphatidic acid; PACS2, phosphofurin acidic cluster sorting protein 2; PC/PLC, phosphatidylcholine; PE, phosphatidylethanolamine; PHB2, prohibitin 2; PINK1, PTEN induced kinase 1; PtdIns, phosphatidylinositol; SAR, Stramenopiles, Apicomplexa and Rhizaria; TAX1BP1, Tax1 binding protein 1; ULK1, unc-51 like autophagy activating kinase 1; VDAC/porin, voltage dependent anion channel. [ABSTRACT FROM AUTHOR]

The ubiquitin kinase-ligase pair PINK1-PRKN recognizes and transiently labels damaged mitochondria with ubiquitin phosphorylated at Ser65 (p-S65-Ub) to mediate their selective degradation (mitophagy). Complete loss of PINK1 or PRKN function unequivocally leads to early-onset Parkinson disease, but it is debated whether impairments in mitophagy contribute to disease later in life. While the pathway has been extensively studied in cell culture upon acute and massive mitochondrial stress, basal levels of activation under endogenous conditions and especially in vivo in the brain remain undetermined. Using rodent samples, patient-derived cells, and isogenic neurons, we here identified age-dependent, brain region-, and cell type-specific effects and determined expression levels and extent of basal and maximal activation of PINK1 and PRKN. Our work highlights the importance of defining critical risk and therapeutically relevant levels of PINK1-PRKN signaling which will further improve diagnosis and prognosis and will lead to better stratification of patients for future clinical trials. [ABSTRACT FROM AUTHOR]

Renal fibrosis is a typical pathological change in chronic kidney disease (CKD). Epithelial-mesenchymal transition (EMT) is the predominant stage. Activation of macroautophagy/autophagy plays a crucial role in the process of EMT. Lycopene (LYC) is a highly antioxidant carotenoid with pharmacological effects such as anti-inflammation, anti-apoptosis and mediation of autophagy. In this study, we demonstrated the specific mechanism of LYC in activating mitophagy and improving renal fibrosis. The enrichment analysis results of GO and KEGG showed that LYC had high enrichment values with autophagy. In this study, we showed that LYC alleviated aristolochic acid I (AAI)-induced intracellular expression of PINK1, TGFB/TGF-β, p-SMAD2, p-SMAD3, and PRKN/Parkin, recruited expression of MAP1LC3/LC3-II and SQSTM1/p62, decreased mitochondrial membrane potential (MMP), and ameliorated renal fibrosis in mice. When we simultaneously intervened NRK52E cells using bafilomycin A1 (Baf-A1), AAI, and LYC, intracellular MAP1LC3-II and SQSTM1 expression was significantly increased. A similar result was seen in renal tissue and cells when treated in vitro and in vivo with CQ, AAI, and LYC, and the inhibitory effect of LYC on the AAI-activated SMAD2-SMAD3 signaling pathway was attenuated. Molecular docking simulation experiments showed that LYC stably bound to the AKT active site. After intervention of cells with AAI and GSK-690693, the expression of PINK1, PRKN, MAP1LC3-II, BECN1, p-SMAD2 and p-SMAD3 was increased, and the expression of SQSTM1 was decreased. However, SC79 inhibited autophagy and reversed the inhibitory effect of LYC on EMT. The results showed that LYC could inhibit the AKT signaling pathway to activate mitophagy and reduce renal fibrosis. Abbreviation: AA: aristolochic acid; ACTA2/α-SMA: actin alpha 2, smooth muscle, aorta; ACTB: actin beta; AKT/protein kinase B: thymoma viral proto-oncogene; BAF-A1: bafilomycin A1; BECN1: beclin 1, autophagy related; CCN2/CTGF: cellular communication network factor 2; CDH1/E-Cadherin: cadherin 1; CKD: chronic kidney disease; COL1: collagen, type I; COL3: collagen, type III; CQ: chloroquine; ECM: extracellular matrix; EMT: epithelial-mesenchymal transition; FN1: fibronectin 1; LYC: lycopene; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MMP: mitochondrial membrane potential; MTOR: mechanistic target of rapamycin kinase ; PI3K: phosphoinositide 3-kinase; PINK1: PTEN induced putative kinase 1; PRKN/Parkin: parkin RBR E3 ubiquitin protein ligase; PPI: protein-protein interaction; SMAD2: SMAD family member 2; SMAD3: SMAD family member 3; SQSTM1/p62: sequestosome 1; TGFB/TGFβ: transforming growth factor, beta; VIM: vimentin [ABSTRACT FROM AUTHOR]

Mitophagy, the selective autophagic clearance of damaged mitochondria, is considered vital for maintaining mitochondrial quality and cellular homeostasis; however, its molecular mechanisms, particularly under basal conditions, and its role in cellular physiology remain poorly characterized. We recently demonstrated that basal mitophagy is a key feature of primary human cells and is downregulated by immortalization, suggesting its dependence on the primary cell state. Mechanistically, we demonstrated that the PINK1-PRKN-SQSTM1 pathway regulates basal mitophagy, with SQSTM1 sensing superoxide-enriched mitochondria through its redox-sensitive cysteine residues, which mediate SQSTM1 oligomerization and mitophagy activation. We developed STOCK1N–57534, a small molecule that targets and promotes this SQSTM1 activation mechanism. Treatment with STOCK1N–57534 reactivates mitophagy downregulated in senescent and naturally aged donor-derived primary cells, improving cellular senescence(−like) phenotypes. Our findings highlight that basal mitophagy is protective against cellular senescence and aging, positioning its pharmacological reactivation as a promising anti-aging strategy.Abbreviation: IR: ionizing radiation; ROS: reactive oxygen species; SARs: selective autophagy receptors. [ABSTRACT FROM AUTHOR]

Mitochondria are crucial organelles in maintaining cellular homeostasis. They are involved in processes such as energy production, metabolism of lipids and glucose, and cell death regulation. Mitochondrial dysfunction can lead to various health issues such as aging, cancer, neurodegenerative diseases, and chronic liver diseases. While mitophagy is the main process for getting rid of excess or damaged mitochondria, there are additional mechanisms for preserving mitochondrial quality. One such alternative mechanism we have discovered is a hybrid organelle called mitochondrial-lysosome-related-organelle (MLRO), which functions independently of the typical autophagy process. More recently, another type of vesicle called vesicle derived from the inner mitochondrial membrane (VDIM) has been identified to break down the inner mitochondrial membrane without involving the standard autophagy pathway. In this article, we will delve into the similarities and differences between MLRO and VDIM, including their structure, regulation, and relevance to human diseases. [ABSTRACT FROM AUTHOR]

Outer membrane vesicles (OMVs) are nanometer-sized membrane blebs secreted by all Gram-negative bacteria to facilitate bacterial communication and modulate the external environment, including in the context of host-microbe interactions. Neisseria gonorrhoeae releases OMVs during interactions with epithelial cells; however, beneficial functional activities for these OMVs have not yet been demonstrated. Our recent study shows that gonococcal OMVs are endocytosed by epithelial cells and subsequently induce mitophagy through a dual PorB-dependent mechanism. PorB is the major gonococcal outer membrane porin protein, which is able to translocate to mitochondria and dissipate the mitochondrial membrane potential, leading to the initiation of a conventional mitophagy mechanism that is dependent on PINK1 and the receptor proteins OPTN or CALCOCO2/NDP52. A second SQSTM1/p62-dependent mitophagy pathway results from direct K63-linked polyubiquitination of PorB lysine residue 171 by the E3 ubiquitin ligase RNF213. Induction of mitophagy favors intracellular gonococcal survival, because it reduces the release of bactericidal mitochondrial reactive oxygen species. These findings highlight a sophisticated bimodal PorB-dependent mechanism by which gonococcal OMVs modulate the intracellular environment to enhance survival in this hostile niche. [ABSTRACT FROM AUTHOR]

Mutations in the PINK1 kinase cause Parkinson disease (PD) through physiological processes that are not yet fully elucidated. PINK1 kinase accumulates selectively on damaged mitochondria, where it recruits the E3 ubiquitin ligase PRKN/Parkin to mediate mitophagy. Upon mitochondrial import failure, PINK1 accumulates in association with the translocase of outer mitochondrial membrane (TOMM). However, the molecular basis of this PINK1 accumulation on the TOMM complex remain elusive. We recently demonstrated that TIMM23 (translocase of the inner mitochondrial membrane 23) is a component of the PINK1-supercomplex formed in response to mitochondrial stress. We also uncovered that PINK1 is required for the formation of this supercomplex and highlighted the biochemical regulation and significance of this supercomplex; expanding our understanding of mitochondrial quality control and PD pathogenesis. [ABSTRACT FROM AUTHOR]

The serine/threonine kinase, PINK1, and the E3 ubiquitin ligase, PRKN/Parkin facilitate LC3-dependent autophagosomal encasement and lysosomal clearance of dysfunctional mitochondria, and defects in this pathway contribute to the pathogenesis of numerous cardiometabolic and neurological diseases. Although dynamic actin remodeling has recently been shown to play an important role in governing spatiotemporal control of mitophagy, the mechanisms remain unclear. We recently found that the RhoGAP, ARHGAP26/GRAF1 is a PRKN-binding protein that is rapidly recruited to damaged mitochondria where upon phosphorylation by PINK1 it serves to coordinate phagophore capture by regulating mitochondrial-associated actin remodeling and by facilitating PRKN-LC3 interactions. Because ARHGAP26 phosphorylation on PINK1-dependent sites is dysregulated in human heart failure and ARHGAP26 depletion in mouse hearts blunts mitochondrial clearance and attenuates compensatory metabolic adaptations to stress, this enzyme may be a tractable target to treat the many diseases associated with mitochondrial dysfunction. [ABSTRACT FROM AUTHOR]

Intervertebral disc degeneration (IDD) is the most critical pathological factor in the development of low back pain. The maintenance of nucleus pulposus (NP) cell and intervertebral disc integrity benefits largely from well-controlled mitochondrial quality, surveilled by mitochondrial dynamics (fission and fusion) and mitophagy, but the outcome is cellular context-dependent that remain to be clarified. Our studies revealed that the loss of NLRX1 is correlated with NP cell senescence and IDD progression, which involve disordered mitochondrial quality. Further using animal and in vitro tissue and cell models, we demonstrated that NLRX1 could facilitate mitochondrial quality by coupling mitochondrial dynamic factors (p-DNM1L, L-OPA1:S-OPA1, OMA1) and mitophagy activity. Conversely, mitochondrial collapse occurred in NLRX1-defective NP cells and switched on the compensatory PINK1-PRKN pathway that led to excessive mitophagy and aggressive NP cell senescence. Mechanistically, NLRX1 was originally shown to interact with zinc transporter SLC39A7 and modulate mitochondrial Zn2+ trafficking via the formation of an NLRX1-SLC39A7 complex on the mitochondrial membrane of NP cells, subsequently orchestrating mitochondrial dynamics and mitophagy. The restoration of NLRX1 function by gene overexpression or pharmacological agonist (NX-13) treatment showed great potential for regulating mitochondrial fission with synchronous fusion and mitophagy, thus sustaining mitochondrial homeostasis, ameliorating NP cell senescence and rejuvenating intervertebral discs. Collectively, our findings highlight a working model whereby the NLRX1-SLC39A7 complex coupled mitochondrial dynamics and mitophagy activity to surveil and target damaged mitochondria for degradation, which determines the beneficial function of the mitochondrial surveillance system and ultimately rejuvenates intervertebral discs. Abbreviations: 3-MA: 3-methyladenine; Baf-A1: bafilomycin A1; CDKN1A/p21: cyclin dependent kinase inhibitor 1A; CDKN2A/p16: cyclin dependent kinase inhibitor 2A; DNM1L/DRP1: dynamin 1 like; EdU: 5-Ethynyl-2'-deoxyuridine; HE: hematoxylin-eosin; IDD: intervertebral disc degeneration; IL1B/IL-1β: interleukin 1 beta; IL6: interleukin 6; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MKI67/Ki67: marker of proliferation Ki-67; LBP: low back pain; MMP: mitochondrial membrane potential; MFN1: mitofusin 1; MFN2: mitofusin 2; MFF: mitochondrial fission factor; NP: nucleus pulposus; NLRX1: NLR family member X1; OMA1: OMA1 zinc metallopeptidase; OPA1: OPA1 mitochondrial dynamin like GTPase; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; ROS: reactive oxidative species; SASP: senescence-associated secretory phenotype; SA-GLB1/β-gal: senescence-associated galactosidase beta 1; SO: safranin o; TBHP: tert-butyl hydroperoxide; TP53/p53: tumor protein p53; SLC39A7/ZIP7: solute carrier family 39 member 7; TOMM20: translocase of outer mitochondrial membrane 20; TIMM23: translocase of inner mitochondrial membrane 23. [ABSTRACT FROM AUTHOR]

Sorafenib is the most widely used first-line drug for the treatment of the advanced hepatocellular carcinoma (HCC). Unfortunately, sorafenib resistance often limits its therapeutic efficacy. To evaluate the efficacy of artesunate against sorafenib-resistant HCC and to investigate its underlying pharmacological mechanisms, a "sorafenib resistance related gene-ART candidate target" interaction network was constructed, and a signaling axis consisting with artesunate candidate target AFAP1L2 and sorafenib target SRC, and the downstream FUNDC1-dependent mitophagy was identified as a major contributor to the sorafenib resistance and a potential way of artesunate to mitigate resistance. Notably, our clinical data demonstrated that AFAP1L2 expression in HCC tissues was markedly higher than that in adjacent non-cancerous liver tissues (P < 0.05), and high AFAP1L2 expression was also significantly associated with an unfavorable overall survival of HCC patients (P < 0.05). Experimentally, AFAP1L2 was overexpressed in sorafenib resistant cells, leading to the activation of downstream SRC-FUNDC1 signaling axis, further blocking the FUNDC1 recruitment of LC3B to mitochondria and inhibiting the activation of mitophagy, based on both in vitro and in vivo systems. Moreover, artesunate significantly enhanced the inhibitory effects of sorafenib on resistant cells and tumors by inducing excessive mitophagy. Mechanically, artesunate reduced the expression of AFAP1L2 protein, suppressed the phosphorylation levels of SRC and FUNDC1 proteins, promoted the FUNDC1 recruitment of massive LC3B to mitochondria, and further overactivated the mitophagy and subsequent cell apoptosis of sorafenib resistant cells. In conclusion, artesunate may be a promising strategy to mitigate sorafenib resistance in HCC via exacerbating AFAP1L2-SRC-FUNDC1 axis-dependent mitophagy. Abbreviations: AFAP1L2, actin filament associated protein 1 like 2; ANOVA, analysis of variance; ANXA5, annexin V; ART: artesunate; CETSA, cellular thermal shift assay; CI: combination index; CO-IP: co-immunoprecipitation; CQ: chloroquine; CT, computed tomography; [18F]-FDG, fluoro-2-D-deoxyglucose F18; FUNDC1: FUN14 domain containing 1; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; HCC, hepatocellular carcinoma; H&E Staining: hematoxylin – eosin staining; HepG2R, sorafenib resistant HepG2; IF, immunofluorescence; IHC, immunohistochemistry; LAMP1: lysosomal associated membrane protein 1; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; miR, microRNA; mRNA: messenger RNA; OE, overexpression; OS, overall survival; PET, positron emission tomography; qRT-PCR: quantitative real-time PCR; sh, short hairpin; shNC: negative control shRNA; shAFAP1L2: short hairpin AFAP1L2; SORA, sorafenib; SPR, surface plasmon resonance; SRC, SRC proto-oncogene, non-receptor tyrosine kinase; SUV, standardized uptake value; TEM, transmission electron microscopy; TOMM20: translocase of outer mitochondrial membrane 20. [ABSTRACT FROM AUTHOR]

Cerebral ischemia induces massive mitochondrial damage, leading to neuronal death. The elimination of damaged mitochondria via mitophagy is critical for neuroprotection. Here we show that the level of PA2G4/EBP1 (proliferation-associated 2G4) was notably increased early during transient middle cerebral artery occlusion and prevented neuronal death by eliciting cerebral ischemia-reperfusion (IR)-induced mitophagy. Neuron-specific knockout of Pa2g4 increased infarct volume and aggravated neuron loss with impaired mitophagy and was rescued by introduction of adeno-associated virus serotype 2 expressing PA2G4/EBP1. We determined that PA2G4/EBP1 is ubiquitinated on lysine 376 by PRKN/PARKIN on the damaged mitochondria and interacts with receptor protein SQSTM1/p62 for mitophagy induction. Thus, our study suggests that PA2G4/EBP1 ubiquitination following cerebral IR-injury promotes mitophagy induction, which may be implicated in neuroprotection. Abbreviations: AAV: adeno-associated virus; ACTB: actin beta; BNIP3L/NIX: BCL2 interacting protein 3 like; CA1: Cornu Ammonis 1; CASP3: caspase 3; CCCP: carbonyl cyanide m-chlorophenyl hydrazone; DMSO: dimethyl sulfoxide; PA2G4/EBP1: proliferation-associated 2G4; FUNDC1: FUN14 domain containing 1; IB: immunoblotting; ICC: immunocytochemistry; IHC: immunohistochemistry; IP: immunoprecipitation; MCAO: middle cerebral artery occlusion; MEF: mouse embryonic fibroblast; OGD: oxygen-glucose deprivation; PRKN/PARKIN: parkin RBR E3 ubiquitin protein ligase; PINK1: PTEN induced kinase 1; RBFOX3/NeuN: RNA binding fox-1 homolog 3; SQSTM1/p62: sequestosome 1; TIMM23: translocase of inner mitochondrial membrane 23; TOMM20: translocase of outer mitochondrial membrane 20; TUBB: tubulin beta class I; WT: wild-type [ABSTRACT FROM AUTHOR]

Mitophagy is a critical mitochondrial quality control process that selectively removes dysfunctional or excess mitochondria through the autophagy-lysosome system. The process is tightly controlled to ensure cellular and physiological homeostasis. Insufficient mitophagy can result in failure to remove damaged mitochondria and consequent cellular degeneration, but it is equally important to appropriately restrain mitophagy to prevent excessive mitochondrial depletion. Here, we discuss our recent discovery that the SKP1-CUL1-F-box (SCF)-FBXL4 (F-box and leucine-rich repeat protein 4) E3 ubiquitin ligase localizes to the mitochondrial outer membrane, where it constitutively mediates the ubiquitination and degradation of BNIP3L/NIX and BNIP3 mitophagy receptors to suppress mitophagy. The post-translational regulation of BNIP3L and BNIP3 is disrupted in mitochondrial DNA depletion syndrome 13 (MTDPS13), a multi-systemic disorder caused by mutations in the FBXL4 gene and characterized by elevated mitophagy and mitochondrial DNA/mtDNA depletion in patient fibroblasts. Our results demonstrate that mitophagy is not solely stimulated in response to specific conditions but is instead also actively suppressed through the continuous degradation of BNIP3L and BNIP3 mediated by the SCF-FBXL4 ubiquitin ligase. Thus, cellular conditions or signaling events that prevent the FBXL4-mediated turnover of BNIP3L and BNIP3 on specific mitochondria are expected to facilitate their selective removal. [ABSTRACT FROM AUTHOR]

Parkinson disease remains a debilitating neurodegenerative disorder, despite the discovery of multiple causative genes that account for familial forms. Prominent among these are PRKN/Parkin and PINK1, whose protein products participate in mitochondrial turnover, or mitophagy. But our poor understanding of the basic biological mechanisms driven by those genes in neurons limits our ability to target them therapeutically. Here, we summarize our recent findings enabled by a new platform to track individual mitochondria in neurons. Our analysis delineates the steps of PINK1- and PRKN-dependent mitochondrial turnover, including the unexplored fates of mitochondria after fusion with lysosomes. These studies reveal unexpected mechanisms of mitochondrial quality control, which may contribute to the reliance of neurons on PINK1 under conditions of stress.

We have employed artificial intelligence to streamline the small molecule drug screening pipeline and identified the cholesterol-reducing compound probucol in the process. Probucol augmented mitophagy and prevented loss of dopaminergic neurons in flies and zebrafish challenged with mitochondrial toxins. Further dissection of the mechanism of action led to the identification of ABCA1, the target of probucol, as a mitophagy modulator. Probucol treatment regulates lipid droplet dynamics during mitophagy and ABCA1 is required for these effects. Here we will summarize the combination of in silico and cell-based screening that led us to identify and characterize probucol as a compound that enhances mitophagy and include thoughts about future directions for the topics explored in our study. Abbreviations: ABCA1: ATP binding cassette transporter protein 1; ATP: Adenosine tri-phosphate; CCCP: carbonyl cyanide m-chlorophenylhydrazone; DsRed: Discosoma red; FDA: Food and drug administration; GFP: Green fluorescent protein; LAMP: lysosome-associated membrane glycoproteins; LD: Lipid droplet; PD: Parkinson's disease; PINK: PTEN-induced kinase [ABSTRACT FROM AUTHOR]

Mitophagy, the process of removing damaged mitochondria to promote cell survival, plays a crucial role in cellular functionality. However, excessive, or uncontrolled mitophagy can lead to reduced mitochondrial content that burdens the remaining organelles, triggering mitophagy-mediated cell death. FBXL4 mutations, which affect the substrate-binding adaptor of the CUL1 (cullin 1)-RING ubiquitin ligase complex (CRL1), have been linked to mitochondrial DNA depletion syndrome type 13 (MTDPS13) characterized by reduced mtDNA content and impaired energy production in affected organs. However, the mechanism behind FBXL4 mutation-driven MTDPS13 remain poorly understood. In a recent study, we demonstrate that the CRL1-FBXL4 complex promotes the degradation of BNIP3 and BNIP3L, two key mitophagy cargo receptors. Deficiency of FBXL4 results in a strong accumulation of BNIP3 and BNIP3L proteins and triggers high levels of BNIP3- and BNIP3L-dependent mitophagy. Patient-derived FBXL4 mutations do not affect its interaction with BNIP3 and BNIP3L but impair the assembly of an active CRL1-FBXL4 complex. Furthermore, excessive mitophagy is observed in knockin mice carrying a patient-derived FBXL4 mutation, and in cortical neurons generated from human patient induced pluripotent stem cells (hiPSCs). These findings support the model that the CRL1-FBXL4 complex tightly restricts basal mitophagy, and its dysregulation leads to severe symptoms of MTDPS13. [ABSTRACT FROM AUTHOR]

Despite growing evidence that has declared the importance of circRNAs in neurodegenerative diseases, the clinical significance of circRNAs in dopaminergic (DA) neuronal degeneration in the pathogenesis of Parkinson disease (PD) remains unclear. Here, we performed rRNA-depleted RNA sequencing and detected more than 10,000 circRNAs in the plasma samples of PD patients. In consideration of ROC and the correlation between Hohen-Yahr stage (H-Y stage) and Unified Parkinson Disease Rating Scale-motor score (UPDRS) of 40 PD patients, circEPS15 was selected for further research. Low expression of circEPS15 was found in PD patients and there was a negative positive correlation between the circEPS15 level and severity of PD motor symptoms, while overexpression of circEPS15 protected DA neurons against neurotoxin-induced PD-like neurodegeneration in vitro and in vivo. Mechanistically, circEPS15 acted as a MIR24-3p sponge to promote the stable expression of target gene PINK1, thus enhancing PINK1-PRKN-dependent mitophagy to eliminate damaged mitochondria and maintain mitochondrial homeostasis. Thus, circEPS15 rescued DA neuronal degeneration through the MIR24-3p-PINK1 axis-mediated improvement of mitochondrial function. This study reveals that circEPS15 exerts a critical role in participating in PD pathogenesis, and may give us an insight into the novel avenue to develop potential biomarkers and therapeutic targets for PD. Abbreviations: AAV: adeno-associated virus; DA: dopaminergic; FISH: fluorescence in situ hybridizations; HPLC: high-performance liquid chromatography; H-Y stage: Hohen-Yahr stage; LDH: lactate dehydrogenase; MMP: mitochondrial membrane potential; MPTP/p: 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/probenecid; NC: negative control; PD: Parkinson disease; PINK1: PTEN induced kinase 1; PBS: phosphate-buffered saline; ROS: reactive oxygen species; SNpc: substantia nigra pars compacta; TEM: transmission electron microscopy; UPDRS: Unified Parkinson's Disease Rating Scale-motor score [ABSTRACT FROM AUTHOR]

Mitophagy, which selectively eliminates the dysfunctional and excess mitochondria by autophagy, is crucial for cellular homeostasis under stresses such as hypoxia. Dysregulation of mitophagy has been increasingly linked to many disorders including neurodegenerative disease and cancer. Triple-negative breast cancer (TNBC), a highly aggressive breast cancer subtype, is reported to be characterized by hypoxia. However, the role of mitophagy in hypoxic TNBC as well as the underlying molecular mechanism is largely unexplored. Here, we identified GPCPD1 (glycerophosphocholine phosphodiesterase 1), a key enzyme in choline metabolism, as an essential mediator in hypoxia-induced mitophagy. Under the hypoxic condition, we found that GPCPD1 was depalmitoylated by LYPLA1, which facilitated the relocating of GPCPD1 to the outer mitochondrial membrane (OMM). Mitochondria-localized GPCPD1 could bind to VDAC1, the substrate for PRKN/PARKIN-dependent ubiquitination, thus interfering with the oligomerization of VDAC1. The increased monomer of VDAC1 provided more anchor sites to recruit PRKN-mediated polyubiquitination, which consequently triggered mitophagy. In addition, we found that GPCPD1-mediated mitophagy exerted a promotive effect on tumor growth and metastasis in TNBC both in vitro and in vivo. We further determined that GPCPD1 could serve as an independent prognostic indicator in TNBC. In conclusion, our study provides important insights into a mechanistic understanding of hypoxia-induced mitophagy and elucidates that GPCPD1 could act as a potential target for the future development of novel therapy for TNBC patients. Abbreviations: ACTB: actin beta; 5-aza: 5-azacytidine; BNIP3: BCL2 interacting protein 3; BNIP3L: BCL2 interacting protein 3 like; CCCP: carbonyl cyanide m-chlorophenyl hydrazone; ChIP: chromatin immunoprecipitation; co-IP: co-immunoprecipitation; CQ: chloroquine; CsA: cyclosporine; DOX: doxorubicin; FIS1: fission, mitochondrial 1; FUNDC1: FUN14 domain containing 1; GPCPD1: glycerophosphocholine phosphodiesterase 1; HAM: hydroxylamine; HIF1A: hypoxia inducible factor 1 subunit alpha; HRE: hypoxia response element; IF: immunofluorescence; LB: lysis buffer; LC3B/MAP1LC3B: microtubule associated protein 1 light chain 3 beta; LC-MS: liquid chromatography-mass spectrometry; LYPLA1: lysophospholipase 1; LYPLA2: lysophospholipase 2; MDA231: MDA-MB-231; MDA468: MDA-MB-468; MFN1: mitofusin 1; MFN2: mitofusin 2; MKI67: marker of proliferation Ki-67; OCR: oxygen consumption rate; OMM: outer mitochondrial membrane; OS: overall survival; PalmB: palmostatin B; PBS: phosphate-buffered saline; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; SDS: sodium dodecyl sulfate; TOMM20: translocase of outer mitochondrial membrane 20; TNBC: triple-negative breast cancer; VBIT-4: VDAC inhibitor; VDAC1: voltage dependent anion channel 1; WT: wild type. [ABSTRACT FROM AUTHOR]

Mitophagy regulates cancer stem cell (CSC) populations affecting tumorigenicity and malignancy in various cancer types. Here, we report that cisplatin treatment led to the activation of higher mitophagy through regulating CLU (clusterin) levels in oral CSCs. Moreover, both the gain-of-function and loss-of-function of CLU indicated its mitophagy-specific role in clearing damaged mitochondria. CLU also regulates mitochondrial fission by activating the Ser/Thr kinase AKT, which triggered phosphorylation of DNM1L/Drp1 at the serine 616 residue initiating mitochondrial fission. More importantly, we also demonstrated that CLU-mediated mitophagy positively regulates oral CSCs through mitophagic degradation of MSX2 (msh homeobox 2), preventing its nuclear translocation from suppressing SOX2 activity and subsequent inhibition of cancer stemness and self-renewal ability. However, CLU knockdown disturbed mitochondrial metabolism generating excessive mitochondrial superoxide, which improves the sensitivity to cisplatin in oral CSCs. Notably, our results showed that CLU-mediated cytoprotection relies on SOX2 expression. SOX2 inhibition through genetic (shSOX2) and pharmacological (KRX-0401) strategies reverses CLU-mediated cytoprotection, sensitizing oral CSCs toward cisplatin-mediated cell death. [ABSTRACT FROM AUTHOR]

Although macroautophagy/autophagy deficiency causes degenerative diseases, the deletion of essential autophagy genes in adipocytes paradoxically reduces body weight. Brown adipose tissue (BAT) plays an important role in body weight regulation and metabolic control. However, the key cellular mechanisms that maintain BAT function remain poorly understood. in this study, we showed that global or brown adipocyte-specific deletion of pink1, a Parkinson disease-related gene involved in selective mitochondrial autophagy (mitophagy), induced BAT dysfunction, and obesity-prone type in mice. Defective mitochondrial function is among the upstream signals that activate the NLRP3 inflammasome. NLRP3 was induced in brown adipocyte precursors (BAPs) from pink1 knockout (KO) mice. Unexpectedly, NLRP3 induction did not induce canonical inflammasome activity. Instead, NLRP3 induction led to the differentiation of pink1 KO BAPs into white-like adipocytes by increasing the expression of white adipocyte-specific genes and repressing the expression of brown adipocyte-specific genes. nlrp3 deletion in pink1 knockout mice reversed BAT dysfunction. Conversely, adipose tissue-specific atg7 KO mice showed significantly lower expression of Nlrp3 in their BAT. Overall, our data suggest that the role of mitophagy is different from general autophagy in regulating adipose tissue and whole-body energy metabolism. Our results uncovered a new mitochondria-NLRP3 pathway that induces BAT dysfunction. The ability of the nlrp3 knockouts to rescue BAT dysfunction suggests the transcriptional function of NLRP3 as an unexpected, but a quite specific therapeutic target for obesity-related metabolic diseases.Abbreviations: ACTB: actin, beta; BAPs: brown adipocyte precursors; BAT: brown adipose tissue; BMDMs: bone marrow-derived macrophages; CASP1: caspase 1; CEBPA: CCAAT/enhancer binding protein (C/EBP), alpha; ChIP: chromatin immunoprecipitation; EE: energy expenditure; HFD: high-fat diet; IL1B: interleukin 1 beta; ITT: insulin tolerance test; KO: knockout; LPS: lipopolysaccharide; NLRP3: NLR family, pyrin domain containing 3; PINK1: PTEN induced putative kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; RD: regular diet; ROS: reactive oxygen species; RT: room temperature; UCP1: uncoupling protein 1 (mitochondrial, proton carrier); WT: wild-type.

Substantial evidence indicates that a decline in mitochondrial health contributes to the development of Parkinson disease. Accordingly, therapeutic stimulation of mitophagy, the autophagic turnover of dysfunctional mitochondria, is a promising approach to treat Parkinson disease. An attractive target in such a setting is PINK1, a protein kinase that initiates the mitophagy cascade. Previous reports suggest that PINK1 kinase activity can be enhanced by kinetin triphosphate (KTP), an enlarged ATP analog that acts as an alternate phosphate donor for PINK1 during phosphorylation. However, the mechanism of how KTP could exert such an effect on PINK1 was unclear. In a recent study, we demonstrate that contrary to previous thinking, KTP cannot be used by PINK1. Nucleotide-bound PINK1 structures indicate that KTP would clash with the back of PINK1’s ATP binding pocket, and enlarging this pocket by mutagenesis is required to enable PINK1 to use KTP. Strikingly, mutation shifts PINK1’s nucleotide preference from ATP to KTP. Similar results could be demonstrated in cells with kinetin, a membrane-permeable precursor of KTP. These results overturn the previously accepted mechanism of how kinetin enhances mitophagy and indicate that kinetin and its derivatives instead function through a currently unidentified mechanism. [ABSTRACT FROM AUTHOR]

Mitophagy is a selective form of autophagy that targets dysfunctional or superfluous mitochondria for degradation. During mitophagy, specific selective autophagy receptors (SARs) mark a portion of mitochondria to recruit the autophagy-related (Atg) machinery and nucleate a phagophore. The phagophore expands and surrounds the mitochondrial cargo, forming an autophagosome. Fission plays a crucial role in separating the targeted portion of mitochondria from the main body to sequester it within the autophagosome. Our recent study, utilizing fission and budding yeasts as model systems, has identified Atg44 as a mitochondrial fission factor that generates mitochondrial fragments suitable for phagophore engulfment. Atg44 resides in the mitochondrial intermembrane space (IMS) and interacts with lipid membranes, with the capacity of mediating membrane fragility and fission. Based on our findings, we propose the term mitofissin to refer to Atg44 and its homologous proteins, which might participate in diverse cellular processes requiring membrane remodeling across various species. Atg: autophagy related; IMM: inner mitochondrial membrane; IMS: intermembrane space; PAS: phagophore assembly site; SAR: selective autophagy receptor [ABSTRACT FROM AUTHOR]

Autophagy plays a crucial role in tumor initiation and progression. However, targeting autophagy in cancer has proven challenging due to genetic or epigenetic factors that may affect the efficacy of autophagy inhibition. Therefore, identifying biomarkers is crucial for selecting patients who are likely to benefit from this treatment modality. We show that dysregulation of mitochondrial translation caused by CBFB (core-binding factor subunit beta) deficiency can sensitize the tumors to autophagy inhibition. CBFB and its binding partner HNRNPK (heterogeneous nuclear ribonucleoprotein K) interact with mRNAs encoded by the mitochondrial genome (mt-mRNAs) and maintain their translation. Specifically, CBFB enhances the binding of TUFM (Tu translation elongation factor, mitochondrial), an elongation factor for mitochondrial translation, to mt-mRNAs. CBFB deficiency, which often occurs in estrogen receptor-positive breast tumors, results in elevated autophagy and mitophagy that promote cancer cell survival. Consequently, these cells are hypersensitive to autophagy inhibition, creating a targetable vulnerability. Studies using in vivo models have shown that inhibiting autophagy selectively eliminates breast tumor cells with mitochondrial translation defects resulting from CBFB deficiency. Our results suggest that autophagy inhibition may be an effective treatment option for breast tumors carrying CBFB alterations. AMPK, AMP-activated protein kinase; BioID, biotinylation identification; CBFB, core-binding factor subunit beta; HCQ, hydroxychloroquine; HNRNPK, heterogeneous nuclear ribonucleoprotein K; PDX, patient-derived xenograft; PIK3CA, phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha; TUFM, Tu translation elongation factor, mitochondrial; ETC, electron transport chain [ABSTRACT FROM AUTHOR]

Mitophagy is a form of autophagy that plays a key role in maintaining the homeostasis of functional mitochondria in the cell. Viruses have evolved various strategies to manipulate mitophagy to escape host immune responses and promote virus replication. In this study, the nucleoprotein (NP) of H1N1 virus (PR8 strain) was identified as a regulator of mitophagy. We revealed that NP-mediated mitophagy leads to the degradation of the mitochondria-anchored protein MAVS, thereby blocking MAVS-mediated antiviral signaling and promoting virus replication. The NP-mediated mitophagy is dependent on the interaction of NP with MAVS and the cargo receptor TOLLIP. Moreover, Y313 of NP is a key residue for the MAVS-NP interaction and NP-mediated mitophagy. The NPY313F mutation significantly attenuates the virus-induced mitophagy and the virus replication in vitro and in vivo. Taken together, our findings uncover a novel mechanism by which the NP of influenza virus induces mitophagy to attenuate innate immunity. Abbreviations: ACTB: actin beta; ATG7: autophagy related 7; ATG12: autophagy related 12; CCCP: carbonyl cyanide 3-chlorophenyl hydrazone; co-IP: co-immunoprecipitation; COX4/COXIV: cytochrome c oxidase subunit 4; DAPI: 4ʹ,6-diamidino-2-phenylindole, dihydrochloride; EID50: 50% egg infective dose; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFP: green fluorescent protein; HEK: human embryonic kidney; hpi: hours post-infection; IAV: influenza A virus; IFN: interferon; IP: immunoprecipitation; LAMP1: lysosomal associated membrane protein 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAVS: mitochondrial antiviral signaling protein; Mdivi-1: mitochondrial division inhibitor 1; MLD50: 50% mouse lethal dose; MOI: multiplicity of infection; NBR1: NBR1 autophagy cargo receptor; NP: nucleoprotein; PB1: basic polymerase 1; RFP: red fluorescent protein; RIGI: RNA sensor RIG-I; RIGI-N: RIGI-CARD; SeV: Sendai virus; SQSTM1/p62: sequestosome 1; TIMM23: translocase of inner mitochondrial membrane 23; TOLLIP: toll interacting protein; TOMM20: translocase of outer mitochondrial membrane 20; TUBA: tubulin alpha; Vec: empty vector; vRNP: viral ribonucleoprotein. [ABSTRACT FROM AUTHOR]

Dysfunction of the endosomal sorting complex required for transport (ESCRT) has been linked to frontotemporal dementia (FTD) due in part to the accumulation of unsealed autophagosomes. However, the mechanisms of ESCRT-mediated membrane closure events on phagophores remain largely unknown. In this study, we found that partial knockdown of non-muscle MYH10/myosin IIB/zip rescues neurodegeneration in both Drosophila and human iPSC-derived cortical neurons expressing FTD-associated mutant CHMP2B, a subunit of ESCRT-III. We also found that MYH10 binds and recruits several autophagy receptor proteins during autophagosome formation induced by mutant CHMP2B or nutrient starvation. Moreover, MYH10 interacted with ESCRT-III to regulate phagophore closure by recruiting ESCRT-III to damaged mitochondria during PRKN/parkin-mediated mitophagy. Evidently, MYH10 is involved in the initiation of induced but not basal autophagy and also links ESCRT-III to mitophagosome sealing, revealing novel roles of MYH10 in the autophagy pathway and in ESCRT-related FTD pathogenesis. Abbreviations: ALS: amyotrophic lateral sclerosis; AP: autophagosome; Atg: autophagy-related; ESCRT: endosomal sorting complex required for transport; FTD: frontotemporal dementia [ABSTRACT FROM AUTHOR]

The contribution of mitochondria to the metabolic function of hypoxic NP cells has been overlooked. We have shown that NP cells contain networked mitochondria and that mitochondrial translocation of BNIP3 mediates hypoxia-induced mitophagy. However, whether BNIP3 also plays a role in governing mitochondrial function and metabolism in hypoxic NP cells is not known. BNIP3 knockdown altered mitochondrial morphology, and number, and increased mitophagy. Interestingly, BNIP3 deficiency in NP cells reduced glycolytic capacity reflected by lower production of lactate/H+ and lower ATP production rate. Widely targeted metabolic profiling and flux analysis using 1-2-13C-glucose showed that the BNIP3 loss resulted in redirection of glycolytic flux into pentose phosphate and hexosamine biosynthesis as well as pyruvate resulting in increased TCA flux. An overall reduction in one-carbon metabolism was noted suggesting reduced biosynthesis. U13C-glutamine flux analysis showed preservation of glutamine utilization to maintain TCA intermediates. The transcriptomic analysis of the BNIP3-deficient cells showed dysregulation of cellular functions including membrane and cytoskeletal integrity, ECM-growth factor signaling, and protein quality control with an overall increase in themes related to angiogenesis and innate immune response. Importantly, we observed strong thematic similarities with the transcriptome of a subset of human degenerative samples. Last, we noted increased autophagic flux, decreased disc height index and aberrant COL10A1/collagen X expression, signs of early disc degeneration in young adult bnip3 knockout mice. These results suggested that in addition to mitophagy regulation, BNIP3 plays a role in maintaining mitochondrial function and metabolism, and dysregulation of mitochondrial homeostasis could promote disc degeneration. Abbreviations: ECAR extracellular acidification rate; HIF hypoxia inducible factor; MFA metabolic flux analysis; NP nucleus pulposus; OCR oxygen consumption rate; ShBnip3 short-hairpin Bnip3 [ABSTRACT FROM AUTHOR]

The ubiquitin (Ub) kinase-ligase pair PINK1-PRKN mediates the degradation of damaged mitochondria by macroautophagy/autophagy (mitophagy). PINK1 surveils mitochondria and upon stress accumulates on the mitochondrial surface where it phosphorylates serine 65 of Ub to activate PRKN and to drive mitochondrial turnover. While loss of either PINK1 or PRKN is genetically linked to Parkinson disease (PD) and activating the pathway seems to have great therapeutic potential, there is no formal proof that stimulation of mitophagy is always beneficial. Here we used biochemical and cell biological methods to study single nucleotide variants in the activation loop of PINK1 to modulate the enzymatic function of this kinase. Structural modeling and in vitro kinase assays were used to investigate the molecular mechanism of the PINK1 variants. In contrast to the PD-linked PINK1G411S mutation that diminishes Ub kinase activity, we found that the PINK1G411A variant significantly boosted Ub phosphorylation beyond levels of PINK1 wild type. This resulted in augmented PRKN activation, mitophagy rates and increased viability after mitochondrial stress in midbrain-derived, gene-edited neurons. Mechanistically, the G411A variant stabilizes the kinase fold of PINK1 and transforms Ub to adopt the preferred, C-terminally retracted conformation for improved substrate turnover. In summary, we identify a critical role of residue 411 for substrate receptivity that may now be exploited for drug discovery to increase the enzymatic function of PINK1. The genetic substitution of Gly411 to Ala increases mitophagy and may be useful to confirm neuroprotection in vivo and might serve as a critical positive control during therapeutic development. Abbreviations: ATP: adenosine triphosphate; CCCP: carbonyl cyanide m-chlorophenyl hydrazone; Ub-CR: ubiquitin with C-terminally retracted tail; CTD: C-terminal domain (of PINK1); ELISA: enzyme-linked immunosorbent assay; HCI: high-content imaging; IB: immunoblot; IF: immunofluorescence; NPC: neuronal precursor cells; MDS: molecular dynamics simulation; PD: Parkinson disease; p-S65-Ub: ubiquitin phosphorylated at Ser65; RMSF: root mean scare fluctuation; TOMM: translocase of outer mitochondrial membrane; TVLN: ubiquitin with T66V and L67N mutation, mimics Ub-CR; Ub: ubiquitin; WT: wild-type. [ABSTRACT FROM AUTHOR]

Mitophagy is a form of autophagy that selectively removes damaged mitochondria and attenuates mitochondrial-dependent apoptosis during viral infection, but how arboviruses balance mitophagy and apoptosis to facilitate persistent viral infection in insect vectors without causing evident fitness cost remains elusive. Here, we identified mitochondrial VDAC1 (voltage-dependent anion channel 1) that could be hijacked by nonstructural protein Pns11 of rice gall dwarf virus (RGDV), a plant nonenveloped double-stranded RNA virus, to synergistically activate pro-viral extensive mitophagy and limited apoptosis in leafhopper vectors. The direct target of fibrillar structures constructed by Pns11 with VDAC1 induced mitochondrial degeneration. Moreover, the degenerated mitochondria were recruited into Pns11-induced phagophores to initiate mitophagy via interaction of VDAC1 with Pns11 and an autophagy protein, ATG8. Such mitophagy mediated by Pns11 and VDAC1 required the classical PRKN/Parkin-PINK1 pathway. VDAC1 regulates apoptosis by controlling the release of apoptotic signaling molecules through its pore, while the anti-apoptotic protein GSN (gelsolin) could bind to VDAC1 pore. We demonstrated that the interaction of Pns11 with VDAC1 and gelsolin decreased VDAC1 expression but increased GSN expression, which prevented the extensive apoptotic response in virus-infected regions. Meanwhile, virus-induced mitophagy also effectively prevented extensive apoptotic response to decrease apoptosis-caused insect fitness cost. The subsequent fusion of virus-loaded mitophagosomes with lysosomes is prevented, and thus such mitophagosomes are exploited for persistent spread of virions within insect bodies. Our results reveal a new strategy for arboviruses to balance and exploit mitophagy and apoptosis, resulting in an optimal intracellular environment for persistent viral propagation in insect vectors. Abbreviations: ATG: autophagy related; BNIP3: BCL2 interacting protein 3; CYCS/CytC: cytochrome c, somatic; dsGSN: double-stranded RNAs targeting GSN/gelsolin; dsGFP: double-stranded RNAs targeting green fluorescent protein; dsPRKN: double-stranded RNAs targeting PRKN; dsPns11: double-stranded RNAs targeting Pns11; dsRNA: double-stranded RNA; EC: epithelia cell; GST: glutathione S-transferase; LAMP1: lysosomal associated membrane protein 1; Mito: mitochondrion; Mmg: middle midgut; MP, mitophagosome; PG, phagophore. padp: post-first access to diseased plants; PINK1: PTEN induced kinase 1; RGDV: rice gall dwarf virus; SQSTM1: sequestosome 1; TOMM20: translocase of outer mitochondrial membrane 20; TUNEL: terminal deoxynucleotidyl transferase dUTP nick end labeling; VDAC1: voltage dependent anion channel 1. [ABSTRACT FROM AUTHOR]

Mitophagy is a key intracellular process that selectively removes damaged mitochondria to prevent their accumulation that can cause neuronal degeneration. During mitophagy, PINK1 (PTEN induced kinase 1), a serine/threonine kinase, works with PRKN/parkin, an E3 ubiquitin ligase, to target damaged mitochondria to the lysosome for degradation. Mutations in the PINK1 and PRKN genes cause early-onset Parkinson disease that is also associated with mitochondrial dysfunction. There are a large number of reports indicating the critical role of PINK1 in mitophagy. However, most of these findings were obtained from in vitro experiments with exogenous PINK1 expression and acute damage of mitochondria by toxins. Recent studies using novel animal models suggest that PINK1-PRKN can also function independent of mitochondria. In this review, we highlight the major differences between in vitro and in vivo models for investigating PINK1 and discuss the potential mechanisms underlying these differences with the aim of understanding how PINK1 functions under different circumstances.Abbreviations: AAV: adeno-associated viruses;AD: Alzheimer disease; CCCP: carbonyl cyanidem-chlorophenyl hydrazone; HD: Huntington disease; MPTP: 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine; MTS: mitochondrial targeting sequence; PD: Parkinson diseases; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; ROS: reactive oxygen species; UIM, ubiquitin interacting motif. [ABSTRACT FROM AUTHOR]

Macroautophagy/autophagy or mitophagy plays crucial roles in the maintenance of pancreatic β-cell function. PPP3/calcineurin can modulate the activity of TFEB, a master regulator of lysosomal biogenesis and autophagy gene expression, through dephosphorylation. We studied whether PPP3/calcineurin inhibitors can affect the mitophagy of pancreatic β-cells and pancreatic β-cell function employing FK506, an immunosuppressive drug against graft rejection. FK506 suppressed rotenone- or oligomycin+antimycin-A-induced mitophagy measured by Mito-Keima localization in acidic lysosomes or RFP-LC3 puncta colocalized with TOMM20 in INS-1 insulinoma cells. FK506 diminished nuclear translocation of TFEB after treatment with rotenone or oligomycin+antimycin A. Forced TFEB nuclear translocation by a constitutively active TFEB mutant transfection restored impaired mitophagy by FK506, suggesting the role of decreased TFEB nuclear translocation in FK506-mediated mitophagy impairment. Probably due to reduced mitophagy, recovery of mitochondrial potential or quenching of mitochondrial ROS after removal of rotenone or oligomycin+antimycin A was delayed by FK506. Mitochondrial oxygen consumption was reduced by FK506, indicating reduced mitochondrial function by FK506. Likely due to mitochondrial dysfunction, insulin release from INS-1 cells was reduced by FK506 in vitro. FK506 treatment also reduced insulin release and impaired glucose tolerance in vivo, which was associated with decreased mitophagy and mitochondrial COX activity in pancreatic islets. FK506-induced mitochondrial dysfunction and glucose intolerance were ameliorated by an autophagy enhancer activating TFEB. These results suggest that diminished mitophagy and consequent mitochondrial dysfunction of pancreatic β-cells contribute to FK506-induced β-cell dysfunction or glucose intolerance, and autophagy enhancement could be a therapeutic modality against post-transplantation diabetes mellitus caused by PPP3/calcineurin inhibitors. [ABSTRACT FROM AUTHOR]