Adenine nucleotide translocator (ANT) is a mitochondrial protein involved in the exchange of ADP and ATP across the mitochondrial inner membrane. It plays a crucial role in cellular energy metabolism by facilitating the transport of ATP synthesized within the mitochondria to the cytoplasm. The isoform ANT1 predominately expresses in cardiac and skeletal muscles. Mutations or dysregulation in ANT1 have been implicated in various mitochondrial disorders and neuromuscular diseases. We aimed to examine whether ANT1 deletion may affect mitochondrial redox state in our established ANT1-deficient mice. Hearts and quadriceps resected from age-matched wild type (WT) and ANT1-deficient mice were snap-frozen in liquid nitrogen. The Chance redox scanner was utilized to perform 3D optical redox imaging. Each sample underwent scanning across 3-5 sections. Global averaging analysis showed no significant differences in the redox indices (NADH, flavin adenine dinucleotide containing-flavoproteins Fp, and the redox ratio Fp/(NADH+Fp) between WT and ANT1-deficient groups. However, quadriceps had higher Fp than hearts in both groups (p = 0.0004 and 0.01, respectively). Furthermore, the quadriceps were also more oxidized (a higher redox ratio) than hearts in WT group (p = 0.004). NADH levels were similar in all cases. Our data suggest that under non-stressful physical condition, the ANT1-deficient muscle cells were in the same mitochondrial state as WT ones and that the significant difference in the mitochondrial redox state between quadriceps and hearts found in WT might be diminished in ANT1-deficient ones. Redox imaging of muscles under physical stress can be conducted in future.

Background: The dysregulation of extraocular muscles (EOMs) in the strabismus may be partly due to modification in the mitochondrial DNA (mtDNA). Currently, little is known about changes occurring in mtDNA of EOMs in patients with strabismus, therefore the aim of our study was to analyze if there are any changes occurring in the mitochondrial DNA of extraocular muscles in children that underwent strabismus surgery in our clinic. Methods: MtDNA was isolated from the tissue material using the Qiagen kit. Assessment of mtDNA mutations was performed by next-generation sequencing (NGS) using the Illumina MiSeq protocol. Results: The examination revealed the presence of atrophic changes in muscle fibers. NGS evaluation revealed a dominant genetic mutation in the ANT1 gene in 12 of the 15 patients examined. Conclusions: The presented results constitute the beginning of research on changes in mtDNA occurring in the muscles of children with strabismus surgery. Further studies are necessary in the context of resolving the transcriptomic differences between strabismic and non-strabismic EOMs. Better understanding of the molecular genetics of strabismus will lead to improved knowledge of the disease mechanisms and ultimately to a more effective treatment.

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Hepatic encephalopathy (HE) implies high morbidity and mortality. The assessment of covert HE (CHE) [i.e. minimal HE (MHE) plus grade 1 HE] is often neglected in Taiwan. Therefore, the aim was to investigate the potential of the animal naming test (ANT1 and simplified ANT1 (S-ANT1)) for assessing CHE in Chinese-speaking regions, specifically Taiwan.
A prospective cohort study was conducted, comprising 65 cirrhotic patients and 29 healthy controls (relatives of the patients). Patients were followed up every three months and censored after two years or until death. Hospitalization for overt HE (OHE) and mortality were considered. All subjects underwent ANT1, psychometric HE score (PHES), and mini-mental state examination (MMSE). The patients underwent an electroencephalogram (EEG) to detect slowing indicative of MHE. Cut-off values for ANT1 and S-ANT1 were assessed by ROC analysis and Youden\'s index, considering CHE as a reference. The prognostic values for OHE and OHE-free survival were assessed.
Preliminary analysis confirmed that PHES ≤-4 is a good discriminant point for abnormal results. CHE was found in 29 patients: 9 had MHE (PHES ≤ -4 or altered EEG) and 20 had grade 1 HE. ANT1 and S-ANT1 were found to have diagnostic values for CHE: AUC = 0.807, 0.786; cut off: 18 and 19, respectively. ANT1 and S-ANT1 were found to have prognostic value for OHE, number of hospitalization episodes for OHE, and OHE recurrence-free survival.
ANT1 shows promise as a tool for CHE detection, quantification, and follow-up in Taiwan and other Chinese-speaking regions.Key messagesThe animal naming test (ANT1) is a simple and valid semantic fluency test that can be easily performed in outpatient or bedside settings in one minute and can also be used as a tool for covert hepatic encephalopathy (CHE) detection, quantification, and follow-up in Taiwan, other Chinese-speaking regions, and many other countries.The diagnostic value of ANT1 and S-ANT1 for CHE were found to be significant, with area under the receiver operating characteristic curve (AUROC) values of 0.807 and 0.786 respectively, and cut-off scores of 18 and 19.ANT1 and S-ANT1 have prognostic value for the first breakthrough of overt hepatic encephalopathy (OHE), number of hospitalization episodes for OHE, and OHE recurrence-free survival, independent of the MELD score.

Mitochondrial biogenesis requires the import of >1,000 mitochondrial preproteins from the cytosol. Most studies on mitochondrial protein import are focused on the core import machinery. Whether and how the biophysical properties of substrate preproteins affect overall import efficiency is underexplored. Here, we show that protein traffic into mitochondria can be disrupted by amino acid substitutions in a single substrate preprotein. Pathogenic missense mutations in ADP/ATP translocase 1 (ANT1), and its yeast homolog ADP/ATP carrier 2 (Aac2), cause the protein to accumulate along the protein import pathway, thereby obstructing general protein translocation into mitochondria. This impairs mitochondrial respiration, cytosolic proteostasis, and cell viability independent of ANT1\'s nucleotide transport activity. The mutations act synergistically, as double mutant Aac2/ANT1 causes severe clogging primarily at the translocase of the outer membrane (TOM) complex. This confers extreme toxicity in yeast. In mice, expression of a super-clogger ANT1 variant led to neurodegeneration and an age-dependent dominant myopathy that phenocopy ANT1-induced human disease, suggesting clogging as a mechanism of disease. More broadly, this work implies the existence of uncharacterized amino acid requirements for mitochondrial carrier proteins to avoid clogging and subsequent disease.
Inside our cells, compartments known as mitochondria generate the chemical energy required for life processes to unfold. Most of the proteins found within mitochondria are manufactured in another part of the cell (known as the cytosol) and then imported with the help of specialist machinery. For example, the TOM and TIM22 channels provide a route for the proteins to cross the two membrane barriers that separate the cytosol from the inside of a mitochondrion. ANT1 is a protein that is found inside mitochondria in humans, where it acts as a transport system for the cell’s energy currency. Specific mutations in the gene encoding ANT1 have been linked to degenerative conditions that affect the muscles and the brain. However, it remains unclear how these mutations cause disease. To address this question, Coyne et al. recreated some of the mutations in the gene encoding the yeast equivalent of ANT1 (known as Aac2). Experiments in yeast cells carrying these mutations showed that the Aac2 protein accumulated in the TOM and TIM22 channels, creating a ‘clog’ that prevented other essential proteins from reaching the mitochondria. As a result, the yeast cells died. Mutant forms of the human ANT1 protein also clogged up the TOM and TIM22 channels of human cells in a similar way. Further experiments focused on mice genetically engineered to produce a “super-clogger” version of the mouse equivalent of ANT1. The animals soon developed muscle and neurological conditions similar to those observed in human diseases associated with ANT1. The findings of Coyne et al. suggest that certain genetic mutations in the gene encoding the ANT1 protein cause disease by blocking the transport of other proteins to the mitochondria, rather than by directly affecting ANT1’s nucleotide trnsport role in the cell. This redefines our understanding of diseases associated with mitochondrial proteins, potentially altering how treatments for these conditions are designed.

Non-essential proteins for viral replication affect host cell metabolism, while the function of the UL43 protein of herpes simplex virus 1 (HSV-1) is not clear. Herein, we performed a comprehensive microarray analysis of HUVEC cells infected with HSV-1 and its UL43-deficient mutant and found significant variation in genes associated with cellular energy metabolic pathways. The localization of UL43 protein in host cells and how it affects cellular energy metabolism pathways were further investigated. Internalization analysis showed that the UL43 protein could be endocytosis-mediated by YPLF motif (aa144-147) and localized to mitochondria. At the same time, more ATP was produced by coupling with mitochondrial small G protein ARF-like 2 (ARL2) GTPase, which triggered the phosphorylation of ANT1 (SLC25A4) to affect the opening degree of mitochondrial permeability transition pore (mPTP), and significantly promoted the aerobic oxidation and oxidative phosphorylation of glucose. Our study shows that UL43 mediates the improvement of host cell metabolism after HSV-1 infection. Additionally, UL43 protein could be a valuable ATP-stimulating factor for mammalian cells.

Protein glycosylation plays a crucial role in central nervous system, and abnormal glycosylation has major implications for human diseases. This study aims to evaluate an etiological implication of the variation in glycosylation for Parkinson\'s disease (PD), a neurodegenerative disorder. Based on a PD mouse model constructed by the intraperitoneal injection with 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine, glycosylation variation was accessed using biotinylated lectin of dolichos biflorus agglutinin (DBA) specific for the exposed N-acetylgalactosamine linked to glycoprotein. Consequently, a glycoprotein with a significantly reduced N-acetylgalactosamination was identified as ADP/ATP translocase 1 (ANT1) by lectin affinity chromatography coupled with MALDI-TOF MS/MS (mass spectrometry), and confirmed by the analysis of dual co-immunofluorescence and Western blot. A tissue-specific distribution of de-N-acetylgalactosaminated ANT1 was found to be correlated with high risk of PD. At cellular level, an obvious co-aggregation between ANT1 and DBA was only found in the MPP+-induced PD-like cell model using dual co-immunofluorescence. Thus, we found that ANT1 was a potential glycoprotein with terminal N-acetylgalactosamine moiety, and the variation of glycosylation in ANT1 was associated with PD. This investigation provides an innovative insight in protein glycosylation with PD pathogenesis.

Exposure to benzo (a)pyrene (BaP) has been confirmed to interfere with embryo implantation. As the primary organ of progesterone synthesis during early pregnancy, the ovarian corpus luteum (CL) is essential for embryo implantation and pregnancy maintenance. We previously demonstrated that BaP impaired luteal function, but the molecular mechanism remains unclear. In CL cells, mitochondria are the main sites of progesterone synthesis. Mitophagy, a particular type of autophagy, regulates mitochondrial quality by degrading damaged mitochondria and ensuring the homeostasis of cell physiology. Therefore, the present study investigated the effects and the potential molecular mechanisms of BaP on ovarian mitophagy during early pregnancy. We found that BaP and its metabolite, BPDE, inhibited autophagy and PINK1/Parkin-mediated mitophagy in the pregnant ovaries and luteinized granulosa cell, KGN. Notably, adenine nucleotide translocator 1 (ANT1), a crucial mediator of PINK1-dependent mitophagy, was suppressed by BaP and BPDE both in vivo and in vitro. The inhibition of ANT1 leads to the decrease in the PINK1 bound to the outer membrane of mitochondria and consequently reduces recruitment of Parkin to the mitochondria, which is required for the subsequent clearance of mitochondria. Meanwhile, exposure to BPDE also damaged mitochondrial function, causing the reduction in mitochondrial potential and ATP production. Overexpression of ANT1 in KGN cells partially relieved the inhibition of mitophagy caused by BPDE, restored mitochondrial function and expression of hormone synthesis-associated genes. Collectively, our study firstly clarified that BaP and BPDE suppress mitophagy of CL cells via the ANT1-PINK1-Parkin pathway, which provides a new insight to explore the detailed mechanism of the BaP-induced ovarian toxicity.

Adenine nucleotide translocase-1 (ANT1) is an ADP/ATP transporter protein located in the inner mitochondrial membrane. ANT1 is involved not only in the processes of ADP/ATP exchange but also in the composition of the mitochondrial membrane permeability transition pore (mPTP); and the function of ANT1 is closely related to its own conformational changes. Notably, various viral proteins can interact directly with ANT1 to influence mitochondrial membrane potential by regulating the opening of mPTP, thereby affecting tumor cell fate. The Epstein-Barr virus (EBV) encodes the key tumorigenic protein, latent membrane protein 1 (LMP1), which plays a pivotal role in promoting therapeutic resistance in related tumors. In our study, we identified a novel mechanism for EBV-LMP1-induced alteration of ANT1 conformation in cisplatin resistance in nasopharyngeal carcinoma. Here, we found that EBV-LMP1 localizes to the inner mitochondrial membrane and inhibits the opening of mPTP by binding to ANT1, thereby favoring tumor cell survival and drug resistance. The ANT1 conformational inhibitor carboxyatractyloside (CATR) in combination with cisplatin improved the chemosensitivity of EBV-LMP1-positive cells. This finding confirms that ANT1 is a novel therapeutic target for overcoming cisplatin resistance in the future.

BACKGROUND: ADP/ATP translocase 1 (ANT1) is involved in the exchange of cytosolic ADP and mitochondrial ATP, and its defection plays an important role in mitochondrial pathogenesis. To reveal an etiological implication of ANT1 for Parkinson\'s disease (PD), a neurodegenerative disorder, a mouse model treated with 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine and neuroblastoma cell model induced by 1-methyl-4-pehny1-pyridine were utilized in this study.
RESULTS: The tissue-specific abundance in ANT1 in mouse brains was accessed using the analysis of Western blot and immunohistochemistry. Down-regulated soluble ANT1 was found to be correlated with PD, and ANT1 was associated with PD pathogenesis via forming protein aggregates with α-synuclein. This finding was confirmed at cellular level using neuroblastoma cell models. ANT1 supplement in neuronal cells revealed the protective roles of ANT1 against cytotoxicity caused by MPP+. Protein interaction assay, coupled with the analysis of LC-MS/MS, silver-stained SDS-PAGE and Western blot against anti-ANT1 antibody respectively, illustrated the interaction of ANT1 with α-synuclein using the expressed α-synuclein as a bite. Additionally, a significant increasing ROSs was detected in the MPP+-treated cells.
CONCLUSIONS: This study indicated that ANT1 was a potentially causative factor of PD, and led to neuropathogenic injury via promoting the formation of protein aggregates with α-synuclein. This investigation potentially promotes an innovative understanding of ANT1 on the etiology of PD and provides valuable information on developing potential drug targets in PD treatment or reliable biomarkers in PD prognostication.