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Objective: To evaluate the effects of uncoupling protein (UCP) 1, UCP2 and UCP3 gene variants on body composition and metabolic changes in response to chronic overfeeding and the recovery after the period of overfeeding. Subjects and design: Twenty-four normal weight men (21±2 y), who constituted 12 pairs of identical twins, ate a 4.2 MJ/day energy surplus, 6 days a week, during a period of 100 days. The subjects were asked to return to the laboratory for testing at 4 months and for a final examination 5 y after completion of the overfeeding protocol. Methods: Resting metabolic rate (RMR) measurements were performed before and after overfeeding. A 4.2 MJ test meal was consumed, after which calorimetric measurements were continued for 240 min. Total body fat was assessed by hydrodensitometry and total subcutaneous fat by the sum of eight skinfolds. Polymorphisms were typed by PCR and PCR-RFLP-techniques. Thyroid stimulating hormone (TSH) concentrations after a thyrotropin releasing hormone (TRH) injection were measured by radioimmunoassay (RIA). Results: The changes in body weight and adiposity were not different between UCP1 Bcl I, UCP2 alanine to valine (A55V), UCP2 insertion/deletion (I/D) or UCP3 Rsa I genotypes. However, the recovery from overfeeding was worse among G-allele carriers of the UCP1 Bcl I, I allele non-carriers of the UCP2 I/D, AV heterozygote subjects of the UCP2 A55V and CC subjects of the UCP3 Rsa I polymorphisms. RMR was lower both before (P=0.01) and after (P=0.001) overfeeding in subjects with the CC genotype of the UCP3 Rsa I polymorphism. Moreover, after overfeeding, the UCP2 A55V heterozygote and UCP3 Rsa I CC homozygote subjects had significantly higher respiratory quotient (RQ) values at rest (P

Exercise powerfully increases energy metabolism and substrate flux in tissues, a process reliant on dramatic changes in mitochondrial energetics. Liver mitochondria play a multi-factorial role during exercise to fuel hepatic glucose output. We previously showed acute exercise activates hepatic mitophagy, a pathway to recycle low-functioning/damaged mitochondria, however little is known how individual bouts of exercise alters the hepatic mitochondrial proteome. Here we leveraged proteomics to examine changes in isolated hepatic mitochondria both immediately after and 2 hours post an acute, 1 hour bout of treadmill exercise in female mice. Further, we utilized leupeptin, a lysosomal inhibitor, to capture and measure exercise-induced changes in mitochondrial proteins that would have been unmeasured due to their targeting for lysosomal degradation. Proteomic analysis of enriched hepatic mitochondria identified 3241 total proteins. Functional enrichment analysis revealed robust enrichment for proteins critical to the mitochondria including metabolic pathways, tricarboxylic acid cycle, and electron transport system. Compared to the sedentary condition, exercise elevated processes regulating lipid localization, Il-5 signaling, and protein phosphorylation in isolated mitochondria. t-SNE analysis identified 4 unique expressional clusters driven by time-dependent changes in protein expression. Isolation of proteins significantly altered with exercise from each cluster revealed influences of leupeptin and exercise both independently and cooperatively modulating mitochondrial protein expressional profiles. Overall, we provide evidence that acute exercise rapidly modulates changes in the proteins/pathways of isolated hepatic mitochondria that include fatty acid metabolism/storage, post-translational protein modification, inflammation, and oxidative stress. In conclusion, the hepatic mitochondrial proteome undergoes extensive remodeling with a bout of exercise.

Oxaliplatin (OXA) is a usual chemotherapeutic agent applied in the colorectal cancer (CRC) clinical treatment. Interferon-alpha inducible protein 6 (IFI6) has been proved to promote proliferation and suppress apoptosis in several tumor cells, while the impacts of IFI6 on OXA resistance in CRC still need exploration. HCT116 and SW620 cells were used as the parental to obtain OXA-resistant cells. The influence of IFI6 on OXA sensitivity, cell proliferation and apoptosis were evaluated by overexpression or knockdown IFI6 in cells. In this work, we found that the level of IFI6 was significantly enhanced in HCT116/OXA and SW620/OXA cells as compared to the parental cells. Overexpression of IFI6 decreased the sensitivity of HCT116 and SW620 cells to OXA. However, knockdown of IFI6 enhanced the sensitivity of HCT116/OXA and SW620/OXA cells to OXA. And upregulated IFI6 promoted the proliferation and repressed apoptosis in HCT116 cells, while suppressed IFI6 markedly reduced proliferation and increased apoptosis in HCT116/OXA cells. Additionally, IFI6 suppressed the phosphorylation level of p38, and silenced IFI6 enhanced it. The addition of the p38 kinase inhibitor, SB203580, alleviated the decreased cell proliferation and increased apoptosis in HCT116/OXA cells. Suppressed IFI6 enhanced the reactive oxygen species (ROS) level in HCT116/OXA cells, and blockade of ROS with N-acetyl-L-cysteine (NAC) decreased the enhancement level of ROS and the phosphorylation level of the p38, which was induced by IFI6 down-regulation. We, therefore, implied that suppressed IFI6 reverses OXA-resistance of CRC cells via promoting the ROS-induced p38 mitogen-activated protein kinase (MAPK) signaling pathway.

The hypoxia-inducible factor 1 (HIF-1) cascade is an ancient and strongly evolutionarily conserved signaling pathway that is involved in the hypoxic responses of most metazoans. Despite immense advances in the understanding of the HIF-1-mediated regulation of hypoxic responses in mammals, the contribution of the hif-1 cascade in the hypoxic adaptation of nonmodel invertebrates remains unclear. In this study, we used the oriental river prawn Macrobrachium nipponense for investigating the roles of hif-1-regulated mitophagy in crustacean testes under hypoxic conditions. We identified that the Bcl-2/adenovirus E1B 19-kDa interacting protein (bnip3) functions as a regulator of mitophagy in M. nipponense and demonstrated that hif-1α activates bnip3 by binding to the bnip3 promoter. Hif-1α knockdown suppressed the expression of multiple mitophagy-related genes, and prawns with hif-1α knockdown exhibited higher mortality under hypoxic conditions. We observed that the levels of BNIP3 were induced under hypoxic conditions and detected that bnip3 knockdown inhibited the mitochondrial translocation of dynamin-related protein 1 (drp1), which is associated with mitochondrial fission. Notably, bnip3 knockdown inhibited hypoxia-induced mitophagy and aggravated the deleterious effects of hypoxia-induced reactive oxygen species (ROS) production and apoptosis. The experimental studies demonstrated that hypoxia induced mitochondrial fission in M. nipponense via drp1. Altogether, the study elucidated the mechanism underlying hif-1/bnip3-mediated mitochondrial fission and mitophagy and demonstrated that this pathway protects crustaceans against ROS production and apoptosis induced by acute hypoxia.

Brown adipose tissue (BAT) is rich in mitochondria containing uncoupling protein 1 (UCP1), and dissipates energy through thermogenesis. However, even though BAT mass and its UCP1 content increase in rodents chronically fed a high-fat sucrose-enriched (HFS) diet, marked expansion of adiposity still occurs in these animals, suggesting insufficient BAT-mediated HFS diet-induced thermogenesis. Thus, the objective of this study was to investigate the metabolic and molecular mechanisms that regulate BAT thermogenesis in HFS-induced obesity. To accomplish this, rats were fed either a standard chow or HFS diet for 8 weeks. Subsequently, glucose and fatty acid metabolism and the molecular mechanisms underlying these processes were assessed in freshly isolated primary BAT adipocytes. Despite increasing BAT mass and its UCP1 content, the HFS diet reduced uncoupled glucose and palmitate oxidation in BAT adipocytes. It also markedly diminished tyrosine hydroxylase content and lipolysis in these cells. Conversely, glucose uptake, lactate production, glycerol incorporation into lipids, palmitate incorporation into triacylglycerol (TAG), phosphoenolpyruvate carboxykinase and glycerol kinase levels, and lipoprotein lipase and cluster of differentiation 36 gene expression were increased. In summary, a HFS diet enhanced glyceroneogenesis and shifted BAT metabolism toward TAG synthesis by impairing UCP1-mediated substrate oxidation and by enhancing fatty acid esterification in intact brown adipocytes. These adaptive metabolic responses to chronic HFS feeding attenuated BAT thermogenic capacity and favoured the development of obesity. KEY POINTS: Despite increasing brown adipose tissue (BAT) mass and levels of thermogenic proteins such as peroxisome proliferator-activated receptor γ coactivator 1α, carnitine palmitoyltransferase 1B and uncoupling protein 1 (UCP1), an obesogenic high-fat sucrose-enriched (HFS) diet attenuated uncoupled glucose and fatty acid oxidation in brown adipocytes. Brown adipocytes diverted glycerol and fatty acids toward triacylglycerol (TAG) synthesis by elevating the cellular machinery that promotes fatty acid uptake along with phosphoenolpyruvate carboxykinase and glycerol kinase levels. The HFS diet increased glucose uptake that supported lactate production and provided substrate for glyceroneogenesis and TAG synthesis in brown adipocytes. Impaired UCP-1-mediated thermogenic capacity and enhanced TAG storage in BAT adipocytes were consistent with reduced adipose triglyceride lipase and tyrosine hydroxylase levels in HFS diet-fed animals.

Necroptosis is the major cause of death in alveolar epithelial cells (AECs) during acute lung injury (ALI). Here, we report a previously unrecognized mechanism for necroptosis. We found an accumulation of mitochondrial citrate (citratemt) in lipopolysaccharide (LPS)-treated AECs because of the downregulation of Idh3α and citrate carrier (CIC, also known as Slc25a1). shRNA- or inhibitor–mediated inhibition of Idh3α and Slc25a1 induced citratemt accumulation and necroptosis in vitro. Mice with AEC-specific Idh3α and Slc25a1 deficiency exhibited exacerbated lung injury and AEC necroptosis. Interestingly, the overexpression of Idh3α and Slc25a1 decreased citratemt levels and rescued AECs from necroptosis. Mechanistically, citratemt accumulation induced mitochondrial fission and excessive mitophagy in AECs. Furthermore, citratemt directly interacted with FUN14 domain-containing protein 1 (FUNDC1) and promoted the interaction of FUNDC1 with dynamin-related protein 1 (DRP1), leading to excessive mitophagy-mediated necroptosis and thereby initiating and promoting ALI. Importantly, necroptosis induced by citratemt accumulation was inhibited in FUNDC1-knockout AECs. We show that citratemt accumulation is a novel target for protection against ALI involving necroptosis.

Twinkle is a mitochondrial replicative helicase which can self-load onto and unwind mitochondrial DNA. Nearly 60 mutations on Twinkle have been linked to human mitochondrial diseases. Using cryo-electron microscopy (cryo-EM) and high-speed atomic force microscopy (HS-AFM), we obtained the atomic-resolution structure of a vertebrate Twinkle homolog with DNA and captured in real-time how Twinkle is self-loaded onto DNA. Our data highlight the important role of the non-catalytic N-terminal domain of Twinkle. The N-terminal domain directly contacts the C-terminal helicase domain, and the contact interface is a hotspot for disease-related mutations. Mutations at the interface destabilize Twinkle hexamer and reduce helicase activity. With HS-AFM, we observed that a highly dynamic Twinkle domain, which is likely to be the N-terminal domain, can protrude ∼5 nm to transiently capture nearby DNA and initialize Twinkle loading onto DNA. Moreover, structural analysis and subunit doping experiments suggest that Twinkle hydrolyzes ATP stochastically, which is distinct from related helicases from bacteriophages.

Mitochondria are involved in various cellular processes, such as energy production, inflammatory responses and cell death. Mitochondrial dysfunction is associated with many age-related diseases, including neurological disorders and heart failure. Mitochondrial quality is strictly maintained by mitochondrial dynamics linked to an adequate supply of phospholipids and other substances from the endoplasmic reticulum (ER). The outer mitochondrial membrane-localized E3 ubiquitin ligase MITOL/MARCHF5 is responsible for mitochondrial quality control through the regulation of mitochondrial dynamics, formation of mitochondria-ER contacts and mitophagy. MITOL deficiency has been shown to impair mitochondrial function, cause an excessive inflammatory response and increase vulnerability to stress, resulting in the exacerbation of the disease. In this study, we overview the ubiquitin-mediated regulation of mitochondrial function by MITOL and the relationship between MITOL and diseases.

Glioblastoma (GBM) is the most malignant tumor in human brain. High invasiveness of this tumor is the main reason causing treatment failure and recurrence. Previous study has found that LACTB is a novel tumor suppressor in breast cancer. Moreover, the function of LACTB in other tumors and mechanisms involving LACTB were also reported. However, the role and relevant mechanisms of LACTB in GBM invasion remains to be revealed. Our aim is to investigate the role LACTB in GBM migration and invasion. We found that LACTB was downregulated in gliomas compared to normal brain tissues. Overexpression of LACTB suppressed migration and invasion of LN229 and U87 cell lines. Mechanistically, LACTB overexpression downregulated the mesenchymal markers. Moreover, LACTB overexpression downregulated the expression of RHOC and inhibited RHOC/Cofilin signaling pathway. The study suggests that LACTB suppresses migration and invasion of GBM cell lines via downregulating RHOC/Cofilin signaling pathway. These findings suggest that LACTB may be a potential treatment target of GBM.

The mhg1 (NCU02695/ada-23) gene encodes the mitochondrial high-mobility group box (HMG-box or HMGB) protein in Neurospora crassa. The mhg1 KO strain (mhg1

Mitochondrial alternative oxidase is an important protein involved in maintaining cellular metabolic and energy balance, especially under stress conditions. AOX genes knockout is aimed at revealing the functions of AOX genes. Under unfavourable conditions, AOX-suppressed plants (mainly based on Arabidopsis AOX1a-knockout lines) usually experience strong oxidative stress. However, a compensation effect, which consists of the absence of AOX1a leading to an increase in defence response mechanisms, concomitant with a decrease in ROS content, has also been demonstrated. This review briefly describes the possible mechanisms underlying the compensation effect upon the suppression of AOX1a. Information about mitochondrial retrograde regulation of AOX is given. The importance of ROS and mitochondrial membrane potential in triggering the signal transmission from mitochondria in the absence of AOX or disturbance of mitochondrial electron transport chain functions is indicated. The few available data on the response of the cell to the absence of AOX at the level of changes in the hormonal balance and the reactions of chloroplasts are presented. The decrease in the relative amount of reduced ascorbate at stable ROS levels as a result of compensation in AOX1a-suppressed plants is proposed as a sign of stress development. Obtaining direct evidence on the mechanisms and signalling pathways involved in AOX modulation in the genome should facilitate a deeper understanding of the role of AOX in the integration of cellular signalling pathways.

Mitochondrial [2Fe-2S] cluster biosynthesis is driven by the coordinated activities of the Iron-Sulfur Cluster (ISC) pathway protein machinery. Within the ISC machinery, the protein that provides a structural scaffold on which [2Fe-2S] clusters are assembled is the ISCU protein in humans; this protein is referred to as the "Scaffold" protein. Truncation of the C-terminal portion of ISCU causes the fatal disease "ISCU Myopathy", which exhibits phenotypes of reduced Fe-S cluster assembly in cells. In this report, the yeast ISCU ortholog "Isu1" has been characterized to gain a better understanding of the role of the scaffold protein in relation to [2Fe-2S] assembly and ISCU Myopathy. Here we explored the biophysical characteristics of the C-terminal region of Isu1, the segment of the protein that is truncated on the human ortholog during the disease ISCU Myopathy. We characterized the role of this region in relation to iron binding, protein stability, assembly of the ISC multiprotein complex required to accomplish Fe-S cluster assembly, and finally on overall cell viability. We determined the Isu1 C-terminus is essential for the completion of the Fe-S cluster assembly but serves a function independent of protein iron binding.

Transmembrane protein 160 (TMEM160) was recently reported to be localized to the mitochondrial inner membrane, but mitochondrial function was noted to be unaffected by loss of TMEM160. In contrast to these previously published findings, we report here that the absence of TMEM160 influences intracellular responses. After confirming that TMEM160 is localized in the inner mitochondrial membrane, we knocked down TMEM160 in human cultured cells and analyzed the changes in cellular responses. TMEM160 depletion led to an upregulation of the mitochondrial chaperone HSPD1, suggesting that depletion induced the mitochondrial unfolded protein response (UPR

Leaf senescence is a highly complex and coordinated process that involves ordered and continuous changes in the external environment and endogenous signaling pathways. Alternative oxidase (AOX), a key enzyme in the respiratory pathway, is involved in the regulation of plant senescence. Salicylic acid (SA) and reactive oxygen species (ROS), two significant inducers of plant senescence, also play an important role in plant senescence, but the relationship between AOX, ROS and SA is unclear. In this study, an RNA interference and overexpression transgenic strain of AOX was constructed using tomato as experimental material. It was found that AOX was able to reduce the accumulation of ROS during dark-induced plant senescence, and that AOX suppressed the synthetic gene SALICYLIC ACID INDUCTION DEFICIENT 2 (SID2) in the SA signaling pathway and salicylate receptor gene NONEXPRESSER OF PR GENES 1 (NPR1), decreasing endogenous SA content and blunting SA perception in plant leaf cells, thereby inhibiting SA-induced leaf senescence. Also, spraying the plants with ROS and ROS scavengers revealed that the synthesis of endogenous SA was affected and that AOX eliminated excess ROS, which in turn affected the SA content and delayed plant senescence. Based on these results, we propose that AOX mediates SA synthesis by balancing ROS content and plays a key role in leaf senescence.

Background: Upregulation of mitochondrial E3 ubiquitin ligase 1 (Mul1) contributes to brain injury in ischemic stroke due to disturbance of mitochondrial dynamics, and bioinformatics analysis predicts that Mul1 is a potential target of Dipsacoside B. Objective: The aim of the study was to explore whether Dipsacoside B can exert a beneficial effect on brain injury in the ischemic stroke rat via targeting Mul1. Methods: The SD rat brains or PC12 cells were subjected to 2 h-ischemia or 8 h-hypoxia plus 24 h-reperfusion or 24 h-reoxygenation to establish the ischemic stroke rat model in vivo or in vitro, which were treated with Dipsacoside B at different dosages. The brain or PC12 cell injury, relevant protein levels and mitochondrial functions were measured by methods of biochemistry, flow cytometry or Western blot. Results: The neurological dysfunction and brain injury (such as infarction and apoptosis) observed in the ischemic stroke rats were accompanied by increases in Mul1 and dynamin-related protein 1 (Drp1) levels along with decreases in mitofusin 2 (Mfn2) level and ATP production. These effects were attenuated by Dipsacoside B. Consistently, cell injury (necroptosis and apoptosis) occurred in the PC12 cells exposed to hypoxia concomitant with the upregulation of Mul1 and Drp1 along with downregulation of Mfn2 and mitochondrial functions (such as increases in reactive oxygen species production and mitochondrial fission and decreases in mitochondrial membrane potential and ATP production).These phenomena were reversed in the presence of Dipsacoside B. Conclusion: Dipsacoside B can protect the rat brain against ischemic injury via inhibition of Mul1 due to the improvement of mitochondrial function.

Purpose of Review This study is aimed at reviewing the recent progress in Drp1 inhibition as a novel approach for reducing doxorubicin-induced cardiotoxicity and for improving cancer treatment. Recent Findings Anthracyclines (e.g. doxorubicin) are one of the most common and effective chemotherapeutic agents to treat a variety of cancers. However, the clinical usage of doxorubicin has been hampered by its severe cardiotoxic side effects leading to heart failure. Mitochondrial dysfunction is one of the major aetiologies of doxorubicin-induced cardiotoxicity. The morphology of mitochondria is highly dynamic, governed by two opposing processes known as fusion and fission, collectively known as mitochondrial dynamics. An imbalance in mitochondrial dynamics is often reported in tumourigenesis which can lead to adaptive and acquired resistance to chemotherapy. Drp1 is a key mitochondrial fission regulator, and emerging evidence has demonstrated that Drp1-mediated mitochondrial fission is upregulated in both cancer cells to their survival advantage and injured heart tissue in the setting of doxorubicin-induced cardiotoxicity. Summary Effective treatment to prevent and mitigate doxorubicin-induced cardiotoxicity is currently not available. Recent advances in cardio-oncology have highlighted that Drp1 inhibition holds great potential as a targeted mitochondrial therapy for doxorubicin-induced cardiotoxicity.

Small ubiquitin-like modifier (SUMO) plays a key regulatory role in cardiovascular diseases, such as cardiac hypertrophy, hypertension, atherosclerosis, and cardiac ischemia-reperfusion injury. As a multifunctional posttranslational modification molecule in eukaryotic cells, SUMOylation is essentially associated with the regulation of mitochondrial dynamics, especially mitophagy, which is involved in the progression and development of cardiovascular diseases. SUMOylation targeting mitochondrial-associated proteins is admittedly considered to regulate mitophagy activation and mitochondrial functions and dynamics, including mitochondrial fusion and fission. SUMOylation triggers mitochondrial fusion to promote mitochondrial dysfunction by modifying Fis1, OPA1, MFN1/2, and DRP1. The interaction between SUMO and DRP1 induces SUMOylation and inhibits lysosomal degradation of DRP1, which is further involved in the regulation of mitochondrial fission. Both SUMOylation and deSUMOylation contribute to the initiation and activation of mitophagy by regulating the conjugation of MFN1/2 SERCA2a, HIF1α, and PINK1. SUMOylation mediated by the SUMO molecule has attracted much attention due to its dual roles in the development of cardiovascular diseases. In this review, we systemically summarize the current understanding underlying the expression, regulation, and structure of SUMO molecules; explore the biochemical functions of SUMOylation in the initiation and activation of mitophagy; discuss the biological roles and mechanisms of SUMOylation in cardiovascular diseases; and further provide a wider explanation of SUMOylation and deSUMOylation research to provide a possible therapeutic strategy for cardiovascular diseases. Considering the precise functions and exact mechanisms of SUMOylation in mitochondrial dysfunction and mitophagy will provide evidence for future experimental research and may serve as an effective approach in the development of novel therapeutic strategies for cardiovascular diseases. Regulation and effect of SUMOylation in cardiovascular diseases via mitophagy. SUMOylation is involved in multiple cardiovascular diseases, including cardiac hypertrophy, hypertension, atherosclerosis, and cardiac ischemia-reperfusion injury. Since it is expressed in multiple cells associated with cardiovascular disease, SUMOylation can be regulated by numerous ligases, including the SENP family proteins PIAS1, PIASy/4, UBC9, and MAPL. SUMOylation regulates the activation and degradation of PINK1, SERCA2a, PPARγ, ERK5, and DRP1 to mediate mitochondrial dynamics, especially mitophagy activation. Mitophagy activation regulated by SUMOylation further promotes or inhibits ventricular diastolic dysfunction, perfusion injury, ventricular remodelling and ventricular noncompaction, which contribute to the development of cardiovascular diseases.

Cardiotoxicity is the major side effect of anthracyclines (doxorubicin, daunorubicin, epirubicin, and idarubicin), though being the most commonly used chemotherapy drugs and the mainstay of therapy in solid and hematological neoplasms. Advances in the field of cardio-oncology have expanded our understanding of the molecular mechanisms underlying anthracycline-induced cardiotoxicity (AIC). AIC has a complex pathogenesis that includes a variety of aspects such as oxidative stress, autophagy, and inflammation. Emerging evidence has strongly suggested that the loss of mitochondrial quality control (MQC) plays an important role in the progression of AIC. Mitochondria are vital organelles in the cardiomyocytes that serve as the key regulators of reactive oxygen species (ROS) production, energy metabolism, cell death, and calcium buffering. However, as mitochondria are susceptible to damage, the MQC system, including mitochondrial dynamics (fusion/fission), mitophagy, mitochondrial biogenesis, and mitochondrial protein quality control, appears to be crucial in maintaining mitochondrial homeostasis. In this review, we summarize current evidence on the role of MQC in the pathogenesis of AIC and highlight the therapeutic potential of restoring the cardiomyocyte MQC system in the prevention and intervention of AIC.

Mitochondrial protein homeostasis in cardiomyocyte injury determines not only the normal operation of mitochondrial function but also the fate of mitochondria in cardiomyocytes. Studies of mitochondrial protein homeostasis have become an integral part of cardiovascular disease research. Modulation of the mitochondrial unfolded protein response (UPRmt), a protective factor for cardiomyocyte mitochondria, may in the future become an important treatment strategy for myocardial protection in cardiovascular disease. However, because of insufficient understanding of the UPRmt and inadequate elucidation of relevant mechanisms, few therapeutic drugs targeting the UPRmt have been developed. The UPRmt maintains a series of chaperone proteins and proteases and is activated when misfolded proteins accumulate in the mitochondria. Mitochondrial injury leads to metabolic dysfunction in cardiomyocytes. This paper reviews the relationship of the UPRmt and mitochondrial quality monitoring with cardiomyocyte protection. This review mainly introduces the regulatory mechanisms of the UPRmt elucidated in recent years and the relationship between the UPRmt and mitophagy, mitochondrial fusion/fission, mitochondrial biosynthesis, and mitochondrial energy metabolism homeostasis in order to generate new ideas for the study of the mitochondrial protein homeostasis mechanisms as well as to provide a reference for the targeted drug treatment of imbalances in mitochondrial protein homeostasis following cardiomyocyte injury.

The small cyprinid genus Prolabeops Schultz, 1941 is restricted to the Nyong and Sanaga River systems in Cameroon. In the past, the genus had been suggested to be either a member of the Labeoninae, Torinae or the Smiliogastrinae mainly on the basis of morphological similarities, and it is nowadays considered as incertae sedis within the Cypriniformes. This study provides the first attempt to reveal the phylogenetic position of Prolabeops using molecular data. For this purpose, the authors sequenced a large fraction of the mitochondrial genome (c. 13,600 bp), including all mitochondrial protein coding genes, of two Prolabeops melanhypopterus specimens and an additional four Enteromius specimens. The large-scale phylogenetic analysis was based on an alignment including all mitochondrial protein coding genes of 902 specimens representing c. 899 cypriniform species. Prolabeops was clearly recovered within the African Smiliogastrinae, forming a weakly supported clade together with Enteromius jae, Enteromius hulstaerti and Barboides gracilis. The study data underline the urgent need of a thorough taxonomic revision of the small African barbs collectively placed in the genus Enteromius.

Background: Experimental evidence suggests a key role of SIRT1 (silent information regulator 1) in age- and metabolic-related vascular dysfunction. Whether these effects hold true in the human microvasculature is unknown. We aimed to investigate the SIRT1 role in very early stages of age- and obesity-related microvascular dysfunction in humans. Methods: Ninety-five subjects undergoing elective laparoscopic surgery were recruited and stratified based on their body mass index status (above or below 30 kg/m 2 ) and age (above or below 40 years) in 4 groups: Young Nonobese, Young Obese, Old Nonobese, and Old Obese. We measured small resistance arteries’ endothelial function by pressurized micromyography before and after incubation with a SIRT1 agonist (SRT1720) and a mitochondria reactive oxygen species (mtROS) scavenger (MitoTEMPO). We assessed vascular levels of mtROS and nitric oxide availability by confocal microscopy and vascular gene expression of SIRT1 and mitochondrial proteins by qPCR. Chromatin immunoprecipitation assay was employed to investigate SIRT1-dependent epigenetic regulation of mitochondrial proteins. Results: Compared with Young Nonobese, obese and older patients showed lower vascular expression of SIRT1 and antioxidant proteins (FOXO3 [forkhead box protein O3] and SOD2) and higher expression of pro-oxidant and aging mitochondria proteins p66 Shc and Arginase II. Old Obese, Young Obese and Old Nonobese groups endothelial dysfunction was rescued by SRT1720. The restoration was comparable to the one obtained with mitoTEMPO. These effects were explained by SIRT1-dependent chromatin changes leading to reduced p66 Shc expression and upregulation of proteins involved in mitochondria respiratory chain. Conclusions: SIRT1 is a novel central modulator of the earliest microvascular damage induced by age and obesity. Through a complex epigenetic control mainly involving p66 Shc and Arginase II, it influences mtROS levels, NO availability, and the expression of proteins of the mitochondria respiratory chain. Therapeutic modulation of SIRT1 restores obesity- and age-related endothelial dysfunction. Early targeting of SIRT1 might represent a crucial strategy to prevent age- and obesity-related microvascular dysfunction.

Although stress-induced mitochondrial hyperfusion (SIMH) exerts a protective role in aiding cell survival, in the absence of mitochondrial fission, SIMH drives oxidative stress-related induction of apoptosis. In this study, our data showed that MTP18, a mitochondrial fission-promoting protein expression, was increased in oral cancer. We have screened and identified S28, a novel inhibitor of MTP18, which was found to induce SIMH and subsequently trigger apoptosis. Interestingly, it inhibited MTP18-mediated mitochondrial fission, as shown by a decrease in p-Drp1 along with increased Mfn1 expression in oral cancer cells. Moreover, S28 induced autophagy but not mitophagy due to the trouble in engulfment of hypoperfused mitochondria. Interestingly, S28-mediated SIMH resulted in the loss of mitochondrial membrane potential, leading to the consequent generation of mitochondrial superoxide to induce intrinsic apoptosis. Mechanistically, S28-induced mitochondrial superoxide caused lysosomal membrane permeabilization (LMP), resulting in decreased lysosomal pH, which impaired autophagosome-lysosome fusion. In this setting, it showed that overexpression of MTP18 resulted in mitochondrial fission leading to mitophagy and inhibition of superoxide-mediated LMP and apoptosis. Further, S28, in combination with FDA-approved anticancer drugs, exhibited higher apoptotic activity and decreased cell viability, suggesting the MTP18 inhibition combined with the anticancer drug could have greater efficacy against cancer.

The mitochondrial replisome replicates the 16.6 kb mitochondria DNA (mtDNA). The proper functioning of this multicomponent protein complex is vital for the integrity of the mitochondrial genome. One of the critical protein components of the mitochondrial replisome is the Twinkle helicase, a member of the Superfamily 4 (SF4) helicases. Decades of research has uncovered common themes among SF4 helicases including self-assembly, ATP-dependent translocation, and formation of protein-protein complexes. Some of the molecular details of these processes are still unknown for the mitochondria SF4 helicase, Twinkle. Here, we describe a protocol for expression, purification, and single-particle cryo-electron microscopy of the Twinkle helicase clinical variant, W315L, which resulted in the first high-resolution structure of Twinkle helicase. The methods described here serve as an adaptable protocol to support future high-resolution studies of Twinkle helicase or other SF4 helicases.

Introduction Thoracic perivascular adipose tissue (tPVAT) has a phenotype resembling brown AT. Dysfunctional tPVAT appears to be linked to vascular dysfunction. Methods We evaluated uncoupling protein 1 (UCP1) expression by Western blot, oxidative stress by measuring lipid peroxidation, the antioxidant capacity by HPLC and spectrophotometry, and mitochondrial respiration by high-resolution respirometry (HRR) in tPVAT, compared to inguinal white AT (iWAT), obtained from non-diabetic (NDM) and streptozocin-induced diabetic (STZ-DM) mice. Mitochondrial respiration was assessed by HRR using protocol 1: complex I and II oxidative phosphorylation (OXPHOS) and protocol 2: fatty acid oxidation (FAO) OXPHOS. OXPHOS capacity in tPVAT was also evaluated after UCP1 inhibition by guanosine 5'-diphosphate (GDP). Results UCP1 expression was higher in tPVAT when compared with iWAT in both NDM and STZ-DM mice. The malondialdehyde concentration was elevated in tPVAT from STZ-DM compared to NDM mice. Glutathione peroxidase and reductase activities, as well as reduced glutathione levels, were not different between tPVAT from NDM and STZ-DM mice but were lower compared to iWAT of STZ-DM mice. OXPHOS capacity of tPVAT was significantly decreased after UCP1 inhibition by GDP in protocol 1. While there were no differences in the OXPHOS capacity between NDM and STZ-DM mice in protocol 1, it was increased in STZ-DM compared to NDM mice in protocol 2. Moreover, complex II- and FAO-linked respiration were elevated in STZ-DM mice under UCP1 inhibition. Conclusions Pharmacological therapies could be targeted to modulate UCP1 activity with a significant impact in the uncoupling of mitochondrial bioenergetics in tPVAT.

During adaptive radiation, mitochondria have co-evolved with their hosts, leading to gain or loss of subunits and assembly factors of respiratory complexes. Plant mitochondrial complex I harbors ∼40 nuclear- and 9 mitochondrial-encoded subunits, and is formed by stepwise assembly during which different intermediates are integrated via various assembly factors. In mammals, the mitochondrial complex I intermediate assembly (MCIA) complex is required for building the membrane arm module. However, plants have lost almost all of the MCIA complex components, giving rise to the hypothesis that plants follow an ancestral pathway to assemble the membrane arm subunits. Here, we characterize a maize crumpled seed mutant, crk1, and reveal by map-based cloning that CRK1 encodes an ortholog of human complex I assembly factor 1, zNDUFAF1, the only evolutionarily conserved MCIA subunit in plants. zNDUFAF1 is localized in the mitochondria and accumulates in two intermediate complexes that contain complex I membrane arm subunits. Disruption of zNDUFAF1 results in severe defects in complex I assembly and activity, a cellular bioenergetic shift to aerobic glycolysis, and mitochondrial vacuolation. Moreover, we found that zNDUFAF1, the putative mitochondrial import inner membrane translocase ZmTIM17-1, and the isovaleryl-coenzyme A dehydrogenase ZmIVD1 interact each other, and could be co-precipitated from the mitochondria and co-migrate in the same assembly intermediates. Knockout of either ZmTIM17-1 or ZmIVD1 could lead to the significantly reduced complex I stability and activity as well as defective seeds. These results suggest that zNDUFAF1, ZmTIM17-1 and ZmIVD1 probably form an MCIA-like complex that is essential for the biogenesis of mitochondrial complex I and seed development in maize. Our findings also imply that plants and mammals recruit MCIA subunits independently for mitochondrial complex I assembly, highlighting the importance of parallel evolution in mitochondria adaptation to their hosts.

Background: Colorectal cancer (CRC) is a common malignancy of digestive tract. Pinocembrin (PINO) has been discovered to have proapoptotic effect on CRC. This study aimed to elucidate how other biological behaviors of CRC cells were affected under PINO treatment. Materials & Methods: The effect of PINO on HT29 and HCT116 cells were detected through treatment of different concentrations of PINO. The role of LACTB in PINO treatment was investigated by transfection of siRNA-LACTB. Cell counting kit-8 assay, wound healing assay, and Transwell assay were conducted to evaluate the proliferation, migration, and invasiveness of CRC cells, respectively. Western blot or quantitative reverse transcription-polymerase chain reaction was carried out to measure the expressions of LACTB, matrix metalloproteinase (MMP)-2, E-cadherin, and N-cadherin. Results: Gradient PINO inhibited the viability, migration, invasiveness, and expressions of MMP-2 and N-cadherin in CRC cells, while promoted E-cadherin and LACTB expressions. Silencing LACTB promoted the viability, migration, invasiveness, and expressions of MMP-2 and N-cadherin in CRC cells and inhibited E-cadherin expression. PINO counteracted the effect of silenced LACTB, and yet silencing LACTB partially abolished the effect of PINO on CRC cells. Conclusion: PINO inhibited the proliferation, migration, invasiveness, and epithelial-to-mesenchymal transition of CRC cells by regulating LACTB.

Splicing of plant mitochondrial introns is facilitated by numerous nucleus-encoded protein factors. Although some splicing factors have been identified in plants, the mechanism underlying mitochondrial intron splicing remains largely unclear. In this study, we identified a small P-type pentatricopeptide repeat (PPR) protein containing merely four PPR repeats, small PPR protein 2 (SPR2), which is required for the splicing of more than half of the introns in maize (Zea mays) mitochondria. Null mutations of Spr2 severely impair the splicing of 15 out of the 22 mitochondrial Group II introns, resulting in substantially decreased mature transcripts, which abolished the assembly and activity of mitochondrial complex I. Consequently, embryogenesis and endosperm development were arrested in the spr2 mutants. Yeast two-hybrid, luciferase complementation imaging, bimolecular fluorescence complementation, and semi-in vivo pull-down analyses indicated that SPR2 interacts with small MutS-related domain protein PPR-SMR1, both of which are required for the splicing of 13 introns. In addition, SPR2 and/or PPR-SMR1 interact with other splicing factors, including PPR proteins EMPTY PERICARP16, PPR14, and chloroplast RNA splicing and ribosome maturation (CRM) protein Zm-mCSF1, which participate in the splicing of specific intron(s) of the 13 introns. These results prompt us to propose that SPR2/PPR-SMR1 serves as the core component of a splicing complex and possibly exerts the splicing function through a dynamic interaction with specific substrate recognizing PPR proteins in mitochondria.

The BolA gene family member (BOLA1–3) plays an important role in regulating normal and pathological biological processes including liver tumorigenesis. However, their expression patterns as prognostic factors in hepatocellular carcinoma (HCC) patients have not to be elucidated. We examined the transcriptional expressions and survival data of BolA family member in patients with HCC from online databases including ONCOMINE, TCGA, UALCAN, Gene Expression Profiling Interactive Analysis (GEPIA), Kaplan-Meier plotter, SurvExpress, cBioPortal, and Exobase. Network molecular interaction views of BolA family members and their neighborhoods were constructed by the IntAct web server. In our research, we had found that the expression levels of BolA /2/3 mRNA were higher in HCC tissue than in normal liver tissues from TGCA databases. Moreover, the BolA family gene expression level is significantly associated with distinct tumor pathological grade, TMN stage, and overall survival (OS). The BolA family can be considered as prognostic risk biomarkers of HCC. A small number of BolA gene-mutated samples were detected in the HCC tissue. IntAct analysis revealed that BolA1/2/3 was closely associated with the GLRX3 expression in HCC, which is implicated in the regulation of the cellular iron homeostasis and tumor growth. Furthermore, prognostic values of altered BolAs and their neighbor GLRX3 gene in HCC patients were validated by SurvExpress analysis. In conclusion, the membrane BolA family identified in this study provides very useful information for the mechanism of hepatic tumorigenesis.

Obesity is linked to reproductive disorders. Novel neuropeptide phoenixin demonstrated many therapeutic actions. In this study, we aim to evaluate phoenixin’s potential effect in obesity-induced infertility through modulating mitochondrial dynamics. Ninety adult female rats were divided to 4 groups: (I), fed with normal pellet diet; (II), given phoenixin; (III), fed with high-fat diet. Rats that developed obesity and infertility were divided to 2 groups: (III-A), received no further treatment; (III-B), given phoenixin. Our results showed that phoenixin treatment in obese infertile rats significantly decreased serum levels of insulin and testosterone and ovarian levels of dynamin-related protein1(Drp1),reactive oxygen species ROS, TNF-α, MDA, and caspase-3. Phoenixin treatment also significantly increased serum estrogen progesterone, LH, and FSH together with ovarian levels of GnRH receptor (GnRHR), mitofusin2(Mfn2), mitochondrial transmembrane potential (ΔΨm), and electron transport chain (ETC) complex-I significantly when compared with obese group. Ovarian histopathological changes were similarly improved by phoenixin. Our data demonstrate phoenixin’s role in improving obesity-induced infertility. Graphical abstract

Tax1 banding protein 1 (Tax1bp1) was originally identified as an NF-κB regulatory protein that participated in inflammatory, antiviral and innate immune processes. Tax1bp1 also functions as an autophagy receptor that plays a role in autophagy. Our previous study shows that Tax1bp1 protects against cardiomyopathy in STZ-induced diabetic mice. In this study we investigated the role of Tax1bp1 in heart failure. Pressure overload-induced heart failure model was established in mice by aortic banding (AB) surgery, and angiotensin II (Ang II)-induced heart failure model was established by infusion of Ang II through osmotic minipump for 4 weeks. We showed that the expression levels of Tax1bp1 in the heart were markedly increased 2 and 4 weeks after AB surgery. Knockdown of Tax1bp1 in mouse hearts significantly ameliorated both AB- and Ang II infusion-induced heart failure parameters. On the contrary, AB-induced heart failure was aggravated in cardiac-specific Tax1bp1 transgenic mice. Similar results were observed in neonatal rat cardiomyocytes (NRCMs) under Ang II insult. We demonstrated that the pro-heart failure effect of Tax1bp1 resulted from its interaction with the E3 ligase ITCH to promote the transcription factor P73 ubiquitination and degradation, causing enhanced BCL2 interacting protein 3 (BNIP3)-mediated cardiomyocyte apoptosis. Knockdown ITCH or BNIP3 in NRCMs significantly reduced Ang II-induced apoptosis in vitro. Similarly, BNIP3 knockdown attenuated heart failure in cardiac-specific Tax1bp1 transgenic mice. In the left ventricles of heart failure patients, Tax1bp1 expression level was significantly increased; Tax1bp1 gene expression was negatively correlated with left ventricular ejection fraction in heart failure patients. Collectively, the Tax1bp1 increase in heart failure enhances ITCH-P73-BNIP3-mediated cardiomyocyte apoptosis and induced cardiac injury. Tax1bp1 may serve as a potent therapeutic target for the treatment of heart failure.• Cardiac Tax1bp1 transgene mice were more vulnerable to cardiac dysfunction under stress.• Cardiac Tax1bp1 transgene mice were more vulnerable to cardiac dysfunction under stress.• Knockout of Tax1bp1 in mouse hearts ameliorated heart failure induced by pressure overload.• Tax1bp1 interacts with the E3 ligase Itch to promote P73 ubiquitination and degradation, causing enhanced BNIP3-mediated apoptosis.• Tax1bp1 may become a target of new therapeutic methods for treating heart failure.

Ca2+ signaling is central to plant development and acclimation. While Ca2+-responsive proteins have been investigated intensely in plants, only a few Ca2+-permeable channels have been identified, and our understanding of how intracellular Ca2+ fluxes is facilitated remains limited. Arabidopsis thaliana homologs of the mammalian channel-forming mitochondrial calcium uniporter (MCU) protein showed Ca2+ transport activity in vitro. Yet, the evolutionary complexity of MCU proteins, as well as reports about alternative systems and unperturbed mitochondrial Ca2+ uptake in knockout lines of MCU genes, leave critical questions about the in vivo functions of the MCU protein family in plants unanswered. Here, we demonstrate that MCU proteins mediate mitochondrial Ca2+ transport in planta and that this mechanism is the major route for fast Ca2+ uptake. Guided by the subcellular localization, expression, and conservation of MCU proteins, we generated an mcu triple knockout line. Using Ca2+ imaging in living root tips and the stimulation of Ca2+ transients of different amplitudes, we demonstrated that mitochondrial Ca2+ uptake became limiting in the triple mutant. The drastic cell physiological phenotype of impaired subcellular Ca2+ transport coincided with deregulated jasmonic acid-related signaling and thigmomorphogenesis. Our findings establish MCUs as a major mitochondrial Ca2+ entry route in planta and link mitochondrial Ca2+ transport with phytohormone signaling. Monitoring of subcellular Ca2+ dynamics in living Arabidopsis roots reveals that MCU proteins provide the dominant mitochondrial Ca2+ uptake mechanism in vivo.

Friedreich Ataxia (FA) is a rare and often fatal autosomal recessive disease in which a mitochondrial protein, frataxin (FXN), is severely reduced in all tissues. With loss of FXN, mitochondrial metabolism is severely disrupted. Multiple therapeutic approaches are in development, but a key limitation is the lack of biomarkers reflecting the activity of FXN in a timely fashion. We predicted this dysregulated metabolism would present a unique metabolite profile in blood of FA patients versus Controls (Con). Plasma from 10 FA and 11 age and sex matched Con subjects was analyzed by targeted mass spectrometry and untargeted NMR. This combined approach yielded quantitative measurements for 540 metabolites and found 59 unique metabolites (55 from MS and 4 from NMR) that were significantly different between cohorts. Correlation-based network analysis revealed several clusters of pathway related metabolites including a cluster associated with one‑carbon (1C) metabolism composed of formate, sarcosine, hypoxanthine, and homocysteine. Receiver operator characteristics analyses demonstrated an excellent ability to discriminate between Con and FA with AUC values0.95. These results are the first reported metabolomic analyses of human patients with FA. The metabolic perturbations, especially those related to 1C metabolism, may serve as a valuable biomarker panel of disease progression and response to therapy. The identification of dysregulated 1C metabolism may also inform the search for new therapeutic targets related to this pathway.

Chemotherapy induces a cachectic-like phenotype, accompanied by skeletal muscle wasting, weakness and mitochondrial dysfunction. Peroxisome proliferator-activated receptor-gamma coactivator-1 alpha (PGC1α), a regulator of mitochondrial biogenesis, is often reduced in cachectic skeletal muscle. Overexpression of PGC1α has yielded mixed beneficial results in cancer cachexia, yet investigations using such approach in a chemotherapy setting are limited. Utilizing transgenic mice, we assessed whether overexpression of PGC1α could combat the skeletal muscle consequences of cisplatin.Young (2 month) and old (18 month) wild-type (WT) and PGC1α transgenic male and female mice (Tg) were injected with cisplatin (C; 2.5 mg/kg) for 2 weeks, while control animals received saline (n = 5-9/group). Animals were assessed for muscle mass and force, motor unit connectivity, and expression of mitochondrial proteins.Young WT + C mice displayed reduced gastrocnemius mass (male: -16%, P 0.0001; female: -11%, P 0.001), muscle force (-6%, P 0.05, both sexes), and motor unit number estimation (MUNE; male: -53%, P 0.01; female: -51%, P 0.01). Old WT + C male and female mice exhibited gastrocnemius wasting (male: -22%, P 0.05; female: -27%, P 0.05), muscle weakness (male: -20%, P 0.0001; female: -17%, P 0.01), and loss of MUNE (male: -82%, P 0.01; female: -62%, P 0.05), suggesting exacerbated cachexia compared with younger animals. Overexpression of PGC1α had mild protective effects on muscle mass in young Tg + C male only (gastrocnemius: +10%, P 0.05); however, force and MUNE were unchanged in both young Tg + C male and female, suggesting preservation of neuromuscular function. In older male, protective effects associated with PGC1α overexpression were heighted with Tg + C demonstrating preserved muscle mass (gastrocnemius: +34%, P 0.001), muscle force (+13%, P 0.01), and MUNE (+3-fold, P 0.05). Similarly, old female Tg + C did not exhibit muscle wasting or reductions in MUNE, and had preserved muscle force (+11%, P 0.05) compared with female WT + C. Follow-up molecular analysis demonstrated that aged WT animals were more susceptible to cisplatin-induced loss of mitochondrial proteins, including PGC1α, OPA1, cytochrome-C, and Cox IV.In our study, the negative effects of cisplatin were heighted in aged animals, whereas overexpression of PGC1α was sufficient to combat the neuromuscular dysfunction caused by cisplatin, especially in older animals. Hence, our observations indicate that aged animals may be more susceptible to develop chemotherapy side toxicities and that mitochondria-targeted strategies may serve as a tool to prevent chemotherapy-induced muscle wasting and weakness.

Background. Liver hepatocellular carcinoma (LIHC) is the predominant type of liver cancer, and its treatment still faces great challenges presently. Mitochondrial inner membrane protein MPV17 is reported to be involved in multiple biological activities of cancers. Here, we seek to investigate the specific role and functions of MPV17 in LIHC progression. Methods. Firstly, MPV17 expressions in various tumors and corresponding normal samples and LIHC groups with various clinical features were analyzed, respectively. Next, the relationship between MPV17 expression and LIHC survival was analyzed and verified by AUC curves. Besides, differentially expressed genes (DEGs) for LIHC were screened from TCGA and then analyzed by GO and KEGG. Then, MPV17 was analyzed by prognostic model, Cox analysis, predictive nomogram, pathway correlation, and immunoassay. Finally, the functions of MPV17 were determined by CCK-8 and Tranwell assays. Results. In most tumors, MPV17 expression was higher than that in the normal group, and it was related to LIHC clinical features. In the LIHC survival analysis, highly expressed MPV17 was associated with a poor prognosis. Besides, 314 upregulated and 193 downregulated DEGs are mainly involved in the TNF signaling pathway and tyrosine metabolism. Through prognostic model, Cox analysis, and predictive nomogram, MPV17 had the prognostic value for LIHC. Gene-pathway correlation analysis showed that MPV17 had the strongest correlation with the G2M_checkpoint pathway. In an immunoassay, MPV17 had a strong correlation with many immune cells. Functional assays showed that MPV17 reduction in LIHC cells could inhibit cell invasion, migration, and proliferation. Conclusion. MPV17, as a tumor promoter, could be a new biomarker for LIHC diagnosis and prognosis and probably shed new light on the exploration of LIHC therapies.

Targeting the cancer cell mitochondrion is a promising approach for developing novel anticancer agents. The experimental anticancer agent N,N'-bis(3,5-dichlorophenyl)urea (SR4) induces apoptotic cell death in several cancer cell lines by uncoupling mitochondrial oxidative phosphorylation (OxPhos) using a protein-free mechanism. However, the precise mechanism by which SR4 depolarizes mitochondria is unclear because SR4 lacks an acidic functional group typically found in protein-independent uncouplers. Recently, it was shown that structurally related thioureas can facilitate proton transport across lipid bilayers by a fatty acid-activated mechanism, in which the fatty acid acts as the site of protonation/deprotonation and the thiourea acts as an anion transporter that shuttles deprotonated fatty acids across the phospholipid bilayer to enable proton leak. In this paper, we show that SR4-mediated proton transport is enhanced by the presence of free fatty acids in the lipid bilayer, indicating that SR4 uncouples mitochondria through the fatty acid-activated mechanism. This mechanistic insight was used to develop a library of substituted bisaryl ureas for structure-activity relationship studies and subsequent cell testing. It was found that lipophilic electron-withdrawing groups on bisaryl ureas enhanced electrogenic proton transport via the fatty acid-activated mechanism and had the capacity to depolarize mitochondria and reduce the viability of MDA-MB-231 breast cancer cells. The most active compound in the series reduced cell viability with greater potency than SR4 and was more effective at inhibiting adenosine triphosphate production.

Plant cells have two endosymbiotic organelles, chloroplasts, and mitochondria. These organelles perform specific functions that depend on organelle-specific proteins. The majority of chloroplast and mitochondrial proteins are specifically imported by the transit peptide and presequence, respectively. However, a significant number of proteins are also dually targeted to these two organelles. Currently, it is not fully understood how proteins are dually targeted to both chloroplasts and mitochondria. In this study, the mechanism underlying mitochondrial targeting of dual targeting AtSufE1 in Arabidopsis was elucidated. The N-terminal fragment containing 80 residues of AtSufE1 (AtSufE1N80) was sufficient to confer dual targeting of reporter protein, AtSufE1N80:GFP, in protoplasts. Two sequence motifs, two arginine residues at 15

Mitochondrial energetic adaptations encompass a plethora of conserved processes that maintain cell and organismal fitness and survival in the changing environment by adjusting the respiratory capacity of mitochondria. These mitochondrial responses are governed by general principles of regulatory biology exemplified by changes in gene expression, protein translation, protein complex formation, transmembrane transport, enzymatic activities and metabolite levels. These changes can promote mitochondrial biogenesis and membrane dynamics that in turn support mitochondrial respiration. The main regulatory components of mitochondrial energetic adaptation include: the transcription coactivator peroxisome proliferator-activated receptor-γ (PPARγ) coactivator 1α (PGC1α) and associated transcription factors; mTOR and endoplasmic reticulum stress signalling; TOM70-dependent mitochondrial protein import; the cristae remodelling factors, including mitochondrial contact site and cristae organizing system (MICOS) and OPA1; lipid remodelling; and the assembly and metabolite-dependent regulation of respiratory complexes. These adaptive molecular and structural mechanisms increase respiration to maintain basic processes specific to cell types and tissues. Failure to execute these regulatory responses causes cell damage and inflammation or senescence, compromising cell survival and the ability to adapt to energetically demanding conditions. Thus, mitochondrial adaptive cellular processes are important for physiological responses, including to nutrient availability, temperature and physical activity, and their failure leads to diseases associated with mitochondrial dysfunction such as metabolic and age-associated diseases and cancer.

Individuals can exhibit differences in metabolism caused by the interplay of genetic background, nutritional input, microbiota, and other environmental factors(1–4). It is difficult to connect differences in metabolism to genomic variation and derive underlying molecular mechanisms in humans due to differences in diet and lifestyle, among others. Here, we introduce the nematode Caenorhabditis elegans as a model to study inter-individual variation in metabolism. By comparing three wild strains and the commonly used N2 laboratory strain, we find differences in the abundances of both known and previously undescribed metabolites. Novel metabolites include conjugates between 3-hydroxypropionate (3HP) and several amino acids (3HP-AAs) that are much higher in abundance in one of the wild strains. 3HP is an intermediate in the propionate shunt pathway, which is activated when flux through the canonical, vitamin B12-dependent propionate breakdown pathway is perturbed(5). We demonstrate that increased 3HP-AA accumulation is caused by genetic variation in HPHD-1, for which 3HP is a substrate. Our results suggest that 3HP-AA production represents a ‘shunt-within-a-shunt’ pathway to accommodate a reduction-of-function allele in hphd-1. This study provides a first step to developing metabolic network models that capture individual-specific differences of metabolism and more closely represent the diversity found in entire species.

Coiled-coil-helix-coiled-coil-helix domain containing 2 (CHCHD2) is a mitochondrial protein that plays important roles in cristae structure, oxidative phosphorylation and apoptosis. Multiple mutations in CHCHD2 have been associated with Lewy body disorders (LBDs), such as Parkinson’s disease (PD) and dementia with Lewy bodies, with the CHCHD2-T61I mutation being the most widely studied. However, at present, only CHCHD2 knockout or CHCHD2/CHCHD10 double knockout mouse models have been investigated. They do not recapitulate the pathology seen in patients with CHCHD2 mutations. We generated the first transgenic mouse model expressing the human PD-linked CHCHD2-T61I mutation driven by the mPrP promoter. We show that CHCHD2-T61I Tg mice exhibit perinuclear mitochondrial aggregates, neuroinflammation, and have impaired long-term synaptic plasticity associated with synaptic dysfunction. Dopaminergic neurodegeneration, a hallmark of PD, is also observed along with α-synuclein pathology. Significant motor dysfunction is seen with no changes in learning and memory at 1 year of age. A minor proportion of the CHCHD2-T61I Tg mice (~10%) show a severe motor phenotype consistent with human Pisa Syndrome, an atypical PD phenotype. Unbiased proteomics analysis reveals surprising increases in many insoluble proteins predominantly originating from mitochondria and perturbing multiple canonical biological pathways as assessed by ingenuity pathway analysis, including neurodegenerative disease-associated proteins such as tau, cofilin, SOD1 and DJ-1. Overall, CHCHD2-T61I Tg mice exhibit pathological and motor changes associated with LBDs, indicating that this model successfully captures phenotypes seen in human LBD patients with CHCHD2 mutations and demonstrates changes in neurodegenerative disease-associated proteins, which delineates relevant pathological pathways for further investigation.

The reproductive consequences of global warming representing heat stress (HS) have been widely received more attention in the last decades. HS induced significant influence on the male reproductive cell, especially sperm functionally. Reduction in the sperm function induced by HS leads to failure of fertility potential. The main effects of HS on sperm are reducing sperm motility, increased abnormalities and changes in the fluidity of the membrane as well as cell morphology. Moreover, the destruction of mitochondrial function could be the result of adverse influences of HS. The protein contents and enzymes of mitochondria were lowered after the exposure of sperm to HS. Some natural antioxidants were used for improving sperm mitochondrial function under HS conditions. In this review, it was highlighted the potential influences of HS on sperm function through reduction in ATP Synthesis yield, mitochondrial activity, mitochondrial protein contents and mitochondrial enzymes, which involves the interference of mitochondrial remodelling in sperm of animals.

Mitochondrial temperature is produced by various metabolic processes inside the mitochondria, particularly oxidative phosphorylation. It was recently reported that mitochondria could normally operate at high temperatures that can reach 50℃. The aim of this review is to identify mitochondrial temperature differences between normal cells and cancer cells. Herein, we discussed the different types of mitochondrial thermosensors and their advantages and disadvantages. We reviewed the studies assessing the mitochondrial temperature in cancer cells and normal cells. We shed the light on the factors involved in maintaining the mitochondrial temperature of normal cells compared to cancer cells.

Brown adipose tissue (BAT) mitochondria generate heat via uncoupled respiration due to excessive proton leak through uncoupling proteins (UCPs). We previously found hyperthermia in a newborn mouse model of fragile X syndrome and excessive leak in Fmr1 KO forebrain mitochondria caused by CoQ deficiency. The inefficient thermogenic nature of Fmr1 mutant forebrain mitochondria was reminiscent of BAT metabolic features. Thus, we aimed to characterize BAT mitochondrial function in these hyperthermic mice using a top-down approach. Although there was no change in steady-state levels of UCP1 expression between strains, BAT weighed significantly less in Fmr1 mutants compared with controls. Fmr1 KO BAT mitochondria demonstrated impaired substrate oxidation, lower mitochondrial membrane potentials and rates of respiration, and CoQ deficiency. The CoQ analog decylubiquinone normalized CoQ-dependent electron flux and unmasked excessive proton leak. Unlike mutant forebrain, where such deficiency resulted in pathological proton leak, CoQ deficiency within BAT mitochondria resulted largely in abnormal substrate oxidation. This suggests that CoQ is important in BAT for uncoupled respiration to produce heat during development. Although our data provide further evidence of a link between fragile X mental retardation protein (FMRP) and CoQ biosynthesis, the results highlight the importance of CoQ in developing tissues and suggest tissue-specific differences from CoQ deficiency. Because BAT mitochondria are primarily responsible for regulating core body temperature, the defects we describe in Fmr1 KOs could manifest as an adaptive downregulated response to hyperthermia or could result from FMRP deficiency directly.

Mitochondria play an important role in both normal heart function and disease etiology. We report analysis of common genetic variations contributing to mitochondrial and heart functions using an integrative proteomics approach in a panel of inbred mouse strains called the Hybrid Mouse Diversity Panel (HMDP). We performed a whole heart proteome study in the HMDP (72 strains, n=2-3 mice) and retrieved 848 mitochondrial proteins (quantified in ≥50 strains). High-resolution association mapping on their relative abundance levels revealed three trans-acting genetic loci on chromosomes (chr) 7, 13 and 17 that regulate distinct classes of mitochondrial proteins as well as cardiac hypertrophy. DAVID enrichment analyses of genes regulated by each of the loci revealed that the chr13 locus was highly enriched for complex-I proteins (24 proteins, P=2.2E-61), the chr17 locus for mitochondrial ribonucleoprotein complex (17 proteins, P=3.1E-25) and the chr7 locus for ubiquinone biosynthesis (3 proteins, P=6.9E-05). Follow-up high resolution regional mapping identified NDUFS4, LRPPRC and COQ7 as the candidate genes for chr13, chr17 and chr7 loci, respectively, and both experimental and statistical analyses supported their causal roles. Furthermore, a large cohort of Diversity Outbred mice was used to corroborate Lrpprc gene as a driver of mitochondrial DNA (mtDNA)-encoded gene regulation, and to show that the chr17 locus is specific to heart. Variations in all three loci were associated with heart mass in at least one of two independent heart stress models, namely, isoproterenol-induced heart failure and diet-induced obesity. These findings suggest that common variations in certain mitochondrial proteins can act in trans to influence tissue-specific mitochondrial functions and contribute to heart hypertrophy, elucidating mechanisms that may underlie genetic susceptibility to heart failure in human populations.

Mitochondria have their own DNA (mtDNA), which encodes essential respiratory subunits. Under live imaging, mitochondrial nucleoids, composed of several copies of mtDNA and DNA-binding proteins, such as mitochondrial transcription factor A (TFAM), actively move inside mitochondria and change the morphology, in concert with mitochondrial membrane fission. Here we found the mitochondrial inner membrane-anchored AAA-ATPase protein ATAD3A mediates the nucleoid dynamics. Its ATPase domain exposed to the matrix binds directly to TFAM and mediates nucleoid trafficking along mitochondria by ATP hydrolysis. Nucleoid trafficking also required ATAD3A oligomerization via an interaction between the coiled-coil domains in intermembrane space. In ATAD3A deficiency, impaired nucleoid trafficking repressed the clustered and enlarged nucleoids observed in mitochondrial fission-deficient cells resulted in dispersed distribution of small nucleoids observed throughout the mitochondrial network, and this enhanced respiratory complex formation. Thus, mitochondrial fission and nucleoid trafficking cooperatively determine the size, number, and distribution of nucleoids in mitochondrial network, which should modulate respiratory complex formation.

Mitochondria are involved in a variety of critical cellular functions, and their impairment drives cell injury. The mitochondrial ribosome (mitoribosome) is responsible for the protein synthesis of mitochondrial DNA-encoded genes. These proteins are involved in oxidative phosphorylation, respiration, and ATP production required in the cell. Mitoribosome components originate from both mitochondrial and nuclear genomes. Their dysfunction can be caused by impaired mitochondrial protein synthesis or mitoribosome misassembly, leading to a decline in mitochondrial translation. This decrease can trigger mitochondrial ribosomal stress and contribute to pulmonary cell injury, death, and diseases. This review focuses on the contribution of the impaired mitoribosome structural components and function to respiratory disease pathophysiology. We present recent findings in the fields of lung cancer, chronic obstructive pulmonary disease, interstitial lung disease, and asthma. We also include reports on the mitoribosome dysfunction in pulmonary hypertension, high-altitude pulmonary edema, and bacterial and viral infections. Studies of the mitoribosome alterations in respiratory diseases can lead to novel therapeutic targets.

Recently, we found that myoglobin (Mb) localizes in both the cytosol and mitochondrial intermembrane space in rodent skeletal muscle. Most proteins of the intermembrane space pass through the outer mitochondrial membrane via the translocase of the outer membrane (TOM) complex. However, whether the TOM complex imports Mb remains unknown. The purpose of this study was to investigate the involvement of the TOM complex in Mb import into the mitochondria. A proteinase K protection assay of mitochondria from C2C12 myotubes confirmed that Mb integrated into the mitochondria. An immunoprecipitation assay verified the interaction of Mb and TOM complex receptors (Tom20, Tom70) in isolated mitochondria. The assay showed a clear interaction of Mb with Tom20 and Tom70. A knockdown experiment using siRNA for TOM complex receptors (Tom20, Tom70) and TOM complex channel (Tom40) did not alter the amount of Mb expression in the mitochondrial fraction. These results suggested that Mb does not necessarily require the TOM complex for mitochondrial import of Mb. Although the physiological role of Mb interactions with TOM complex receptors remains unclear, further studies are needed to clarify how Mb enters the mitochondria independently of the TOM complex.

Redox homeostasis is elemental for the normal physiology of all cell types. Cells use multiple mechanisms to tightly regulate the redox balance. The onset and progression of many metabolic and aging-associated diseases occur due to the dysregulation of redox homeostasis. Thus, it is critical to identify and therapeutically target mechanisms that precipitate abnormalities in redox balance. Reactive oxygen species (ROS) produced within the immune cells regulate homeostasis, hyperimmune and hypoimmune cell responsiveness, apoptosis, immune response to pathogens, and tumor immunity. Immune cells have both cytosolic and organelle-specific redox regulatory systems to maintain appropriate levels of ROS. Nicotinamide nucleotide transhydrogenase (NNT) is an essential mitochondrial redox regulatory protein. Dysregulation of NNT function prevents immune cells from mounting an adequate immune response to pathogens, promotes a chronic inflammatory state associated with aging and metabolic diseases, and initiates conditions related to a dysregulated immune system such as autoimmunity. Although many studies have reported on NNT in different cell types, including cancer cells, relatively few studies have explored NNT in immune cells. This review provides an overview of NNT and focuses on the current knowledge of NNT in the immune cells.

Acute myelogenous leukemia (AML) is a very common hematopoietic malignancy. Hematopoietic stem cell transplantation can improve the therapeutic effect of AML, but the 5-year survival rate is very low. CD123 imbalance, abnormal gene expression, and epigenetics play an important role in the pathogenesis of AML. This research was to explore the differential expression of CD123-related long non-coding RNA (lncRNA) in AML bone marrow mononuclear cells and provide a theoretical basis for targeted therapy of AML. High-throughput sequencing was performed to screen differentially expressed lncRNA in bone marrow mononuclear immunophenotypes of CD123+ and CD123- from patients with primary AML, and real-time quantitative PCR was adopted for screening and validation. There were 933 differentially expressed lncRNAs in the CD123+ group and the CD123- group, 407 lncRNAs were up-regulated and 463 lncRNAs were down-regulated in the CD123+ group. 14 lncRNAs with more than 2 times of difference were screened for identification, and it was found that compared with CD123- group, there was no substantial difference in the expression of JHDM1D-AS1, LINC01355, CASC15, FAM13A-AS1, HSPC324, LOC339803, LINC00877, and MAG12-AS3 in CD123+ group (P0.05). The expressions of LOC101929698, BaALC-AS2, BOLA3-AS1, and FBX19-AS1 were considerably up-regulated (P0.05), while the expressions of LOC100132249 and LINC02085 were considerably down-regulated (P0.05). In summary, differentially expressed lncRNAs in bone marrow samples of CD123+ and CD123- group of newly diagnosed AML patients may be involved in the process of AML and seriously affect the prognosis of patients.

Mitophagy, a mitochondria-specific form of autophagy, removes dysfunctional mitochondria and is hence an essential process contributing to mitochondrial quality control. PTEN-induced kinase 1 (PINK1) and the E3 ubiquitin ligase Parkin are critical molecules involved in stress-induced mitophagy, but the intracellular signaling mechanisms by which this pathway is regulated are unclear. We tested the hypothesis that signaling through RhoA, a small GTPase, induces mitophagy via modulation of the PINK1/Parkin pathway as a protective mechanism against ischemic stress. We demonstrate that expression of constitutively active RhoA as well as sphingosine-1-phosphate induced activation of endogenous RhoA in cardiomyocytes result in an accumulation of PINK1 at mitochondria. This is accompanied by translocation of Parkin to mitochondria and ubiquitination of mitochondrial proteins leading to recognition of mitochondria by autophagosomes and their lysosomal degradation. Expression of RhoA in cardiomyocytes confers protection against ischemia, and this cardioprotection is attenuated by siRNA-mediated PINK1 knockdown. In vivo myocardial infarction elicits increases in mitochondrial PINK1, Parkin, and ubiquitinated mitochondrial proteins. AAV9-mediated RhoA expression potentiates these responses and a concurrent decrease in infarct size is observed. Interestingly, induction of mitochondrial PINK1 accumulation in response to RhoA signaling is neither mediated through its transcriptional upregulation nor dependent on depolarization of the mitochondrial membrane, the canonical mechanism for PINK1 accumulation. Instead, our results reveal that RhoA signaling inhibits PINK1 cleavage, thereby stabilizing PINK1 protein at mitochondria. We further show that active RhoA localizes at mitochondria and interacts with PINK1, and that the mitochondrial localization of RhoA is regulated by its downstream effector protein kinase D. These findings demonstrate that RhoA activation engages a unique mechanism to regulate PINK1 accumulation, induce mitophagy and protect against ischemic stress, and implicates regulation of RhoA signaling as a potential strategy to enhance mitophagy and confer protection under stress conditions.

Gene expression in plant mitochondria is predominantly governed at the post-transcriptional level and relies mostly on nuclear-encoded proteins. However, the protein factors involved and the underlying molecular mechanisms are still not well understood. Here, we report on the function of the MITOCHONDRIAL STABILITY FACTOR 3 (MTSF3) protein, previously named EMBRYO DEFECTIVE 2794 (EMB2794), and show that it is essential for accumulation of the mitochondrial NADH dehydrogenase subunit 2 (nad2) transcript in Arabidopsis (Arabidopsis thaliana) but not for splicing of nad2 intron 2 as previously proposed. The MTSF3 gene encodes a pentatricopeptide repeat protein that localizes in the mitochondrion. An MTSF3 null mutation induces embryonic lethality, but viable mtsf3 mutant plants can be generated through partial complementation with the developmentally regulated ABSCISIC ACID INSENSITIVE3 promoter. Genetic analyses revealed growth retardation in rescued mtsf3 plants owing to the specific destabilization of mature nad2 mRNA and a nad2 precursor transcript bearing exons 3 to 5. Biochemical data demonstrate that MTSF3 protein specifically binds to the 3′ terminus of nad2. Destabilization of nad2 mRNA induces a substantial decrease in complex I assembly and activity and overexpression of the alternative respiratory pathway. Our results support a role for MTSF3 protein in protecting two nad2 transcripts from degradation by mitochondrial exoribonucleases by binding to their 3′ extremities.