Oxidative Phosphorylation Complexes Research Articles

Oxidative phosphorylation decline and mitochondrial dynamics disequilibrium are involved in chicken large white follicle atresia

In domestic hens, the atresia of large white follicles (LWFs) directly affects the number of follicles that enter the hierarchical development and ovulation. Figuring out factors responsible for LWFs atresia is helpful to improve egg production of hens. At the LWF stage, yellow yolk begins to be deposited into the follicles via receptor mediated endocytosis, which requires large amounts of ATP. Mitochondrial oxidative phosphorylation (OXPHOS) is the primary source of ATP for follicular development. However, it is not clear whether the OXPHOS is changed along LWFs atresia. In this study, firstly, differences in morphological appearance, histology, cell proliferation, apoptosis, OXPHOS and mitochondrial dynamics between LWFs and atretic large white follicles (ALWFs) in hens at the peak laying stage (35W) were determined to elucidate whether OXPHOS changes in ALWFs. Then, these differences of LWFs between the peak laying hens (35W-LWFs) and the late laying hens (70W-LWFs) were detected to confirm whether OXPHOS changes during LWFs atresia. The results showed that ALWFs exhibited a wrinkled surface with several hemorrhage spots, and numerous intercellular vacuoles, as well as severe nuclear pyknosis. Compared to LWFs, a higher cell apoptosis rate and a lower proliferation rate were observed in ALWFs. In ALWFs, OXPHOS declined as manifested by reductions in ATP levels, ATP synthetase abundance, NAD+, NADH and NAD+/NADH ratio, and mRNA levels of genes associated with OXPHOS complexes I-V. Meanwhile, mitochondrial dynamics disequilibrium was detected in ALWFs as the expression levels of proteins and genes related to mitochondrial fusion (MFN1, MFN2, and OPA1) decreased, while the expression levels of proteins and genes related to mitochondrial fission (DRP1 and FIS1) increased. Further, compared to 35W-LWFs, 70W-LWFs showed a histology resembling to ALWFs, manifested as a slightly loosen structure of granulosa layers, and a lower cell proliferation rate. Moreover, both lower OXPHOS and impaired mitochondrial dynamics were detected in 70W-LWFs. In conclusion, our results indicated that OXPHOS decline and mitochondrial dynamics disequilibrium are involved in LWFs atresia in laying hens.

Adipose Tissue-Resident Sphingomonas Paucimobilis Suppresses Adaptive Thermogenesis by Reducing 15-HETE Production and Inhibiting AMPK Pathway.

Obesity represents a low-grade chronic inflammation status, which is associated with compromised adaptive thermogenesis. However, the mechanisms underlying the defective activation of thermogenesis in chronic inflammation remain unclear. Here, a chronic inflammatory model is first estabolished by injecting mice with low-dose lipopolysaccharide (LPS) before cold exposure, and then it is verified that LPS treatment can decrease the core body temperature of mice and alter the microbial distribution in epididymal white adipose tissue (eWAT). An adipose tissue-resident bacterium Sphingomonas paucimobilis is identified as a potential inhibitor on the activation of brown fat and browning of inguinal WAT, resulting in defective adaptive thermogenesis. Mechanically, LPS and S. paucimobilis inhibit the production and release of 15-HETE by suppressing its main metabolic enzyme 12 lipoxygenase (12-LOX) and 15- Hydroxyeicosatetraenoic acid (15-HETE) rescues the impaired thermogenesis. Interestingly, 15-HETE directly binds to AMP-activated protein kinase α (AMPKα) and elevates the phosphorylation of AMPK, leading to the activation of uncoupling protein 1 (UCP1) and mitochondrial oxidative phosphorylation (OXPHOS) complexes. Further analysis with human obesity subjects reveals that individuals with high body mass index displayed lower 15-HETE levels. Taken together, this work improves the understanding of how chronic inflammation impairs adaptive thermogenesis and provides novel targets for alleviating obesity.

A novel m.5906G > a variant in MT-CO1 causes MELAS/Leigh overlap syndrome.

The MELAS/Leigh overlap syndrome manifests with a blend of clinical and radiographic traits from both MELAS and LS. However, the association of MELAS/Leigh overlap syndrome with MT-CO1 gene variants has not been previously reported. In this study, we report a patient diagnosed with MELAS/Leigh overlap syndrome harboring the m.5906G > A variant in MT-CO1, with biochemical evidence supporting the pathogenicity of the variant. The variant m.5906G > A that led to a synonymous variant in the start codon of MT-CO1 was filtered as the candidate disease-causing variant of the patient. Patient-derived fibroblasts were used to generate a series of monoclonal cells carrying different m.5906G > A variant loads for further functional assays. The oxygen consumption rate, ATP production, mitochondrial membrane potential and lactate assay indicated an impairment of cellular bioenergetics due to the m.5906G > A variant. Blue native PAGE analysis revealed that the m.5906G > A variant caused a deficiency in the content of mitochondrial oxidative phosphorylation complexes. Furthermore, molecular biology assays performed for the pathogenesis, mtDNA copy number, mtDNA-encoded subunits, and recovery capacity of mtDNA were all deficient due to the m.5906G > A variant, which might be caused by mtDNA replication deficiency. Overall, our findings demonstrated the pathogenicity of m.5906G > A variant and proposed a potential pathogenic mechanism, thereby expanding the genetic spectrum of MELAS/Leigh overlap syndrome.

Rapid evolution of mitochondrion-related genes in haplodiploid arthropods

BackgroundMitochondrial genes and nuclear genes cooperate closely to maintain the functions of mitochondria, especially in the oxidative phosphorylation (OXPHOS) pathway. However, mitochondrial genes among arthropod lineages have dramatic evolutionary rate differences. Haplodiploid arthropods often show fast-evolving mitochondrial genes. One hypothesis predicts that the small effective population size of haplodiploid species could enhance the effect of genetic drift leading to higher substitution rates in mitochondrial and nuclear genes. Alternatively, positive selection or compensatory changes in nuclear OXPHOS genes could lead to the fast-evolving mitochondrial genes. However, due to the limited number of arthropod genomes, the rates of evolution for nuclear genes in haplodiploid species, besides hymenopterans, are largely unknown. To test these hypotheses, we used data from 76 arthropod genomes, including 5 independently evolved haplodiploid lineages, to estimate the evolutionary rates and patterns of gene family turnover of mitochondrial and nuclear genes.ResultsWe show that five haplodiploid lineages tested here have fast-evolving mitochondrial genes and fast-evolving nuclear genes related to mitochondrial functions, while nuclear genes not related to mitochondrion showed no significant evolutionary rate differences. Among hymenopterans, bees and ants show faster rates of molecular evolution in mitochondrial genes and mitochondrion-related nuclear genes than sawflies and wasps. With genome data, we also find gene family expansions and contractions in mitochondrion-related genes of bees and ants.ConclusionsOur results reject the small population size hypothesis in haplodiploid species. A combination of positive selection and compensatory changes could lead to the observed patterns in haplodiploid species. The elevated evolutionary rates in OXPHOS complex 2 genes of bees and ants suggest a unique evolutionary history of social hymenopterans.

Ndufs4 knockout mice with isolated complex I deficiency engage a futile adaptive brain response

Paediatric Leigh syndrome (LS) is an early-onset and fatal neurodegenerative disorder lacking treatment options. LS is frequently caused by mutations in the NDUFS4 gene, encoding an accessory subunit of mitochondrial complex I (CI), the first complex of the oxidative phosphorylation (OXPHOS) system. Whole-body Ndufs4 knockout (KO) mice (WB-KO mice) are widely used to study isolated CI deficiency, LS pathology and interventions. These animals develop a brain-specific phenotype via an incompletely understood pathomechanism. Here we performed a quantitative analysis of the sub-brain proteome in six-weeks old WB-KO mice vs. wildtype mice. Brain regions comprised of a brain slice (BrSl), cerebellum (CB), cerebral cortex (CC), hippocampus (HC), inferior colliculus (IC), and superior colliculus (SC). Proteome analysis demonstrated similarities between CC/HC, and between IC/SC, whereas BrSl and CB differed from these two groups and each other. All brain regions displayed greatly reduced levels of two CI structural subunits (NDUFS4, NDUFA12) and an increased level of the CI assembly factor NDUFAF2. The level of CI-Q module subunits was significantly more reduced in IC/SC than in BrSl/CB/CC/HC, whereas other OXPHOS complex levels were not reduced. Gene ontology and pathway analysis demonstrated specific and common proteome changes between brain regions.Across brain regions, upregulation of cold-shock-associated proteins, mitochondrial fatty acid (FA) oxidation and synthesis (mtFAS) were the most prominent. FA-related pathways were predominantly upregulated in CB and HC. Based upon these results, we argue that stimulation of these pathways is futile and pro-pathological and discuss alternative strategies for therapeutic intervention in LS. SignificanceThe Ndufs4 knockout mouse model is currently the most relevant and most widely used animal model to study the brain-linked pathophysiology of human Leigh Syndrome (LS) and intervention strategies. We demonstrate that the Ndufs4 knockout brain engages futile and pro-pathological responses. These responses explain both negative and positive outcomes of intervention studies in Leigh Syndrome mice and patients, thereby guiding novel intervention opportunities.

Enhanced Anticancer Activity of 7MeERT over Ertredin: A Comparative Study on Cancer Cell Proliferation and NDUFA12 Binding.

We have previously identified Ertredin (3-(2-amino-5-bromophenyl) quinoxalin-2(1H)-one) as a compound that suppresses 3D spheroid formation and tumorigenesis in NIH3T3 cells induced by Epidermal Growth Factor Receptor variant III (EGFRvIII) transduction. One of its targets has been shown to be NDUFA12 (NADH Dehydrogenase (Ubiquinone) 1 Alpha Subcomplex Subunit 12), a component protein of oxidative phosphorylation complex I. In this report, we compared the growth inhibitory activity of Ertredin with its methylated analogue 7MeERT (3-(2-amino-5-bromophenyl)-7-methylquinoxalin-2(1H)-one) on human cancer cells. 7MeERT induced the inhibition of the proliferation of various cancer cells similarly to Ertredin and showed higher activity in glioblastoma cells, A431 cells overexpressing EGFR (wild type), and multiple myeloma cells. Molecular docking analysis and a Cellular Thermal Shift Assay (CETSA) suggested that 7MeERT binds to NDUFA12 similarly to Ertredin. The binding of 7MeERT and Ertredin to NDUFA12 in glioblastoma was further supported by the inhibition of the oxygen consumption rate. These results suggest that 7MeERT also binds to NDUFA12, inhibits oxidative phosphorylation, and has a higher anti-cancer cell growth inhibitory activity than Ertredin.

Biochemical and neurophysiological effects of deficiency of the mitochondrial import protein TIMM50.

TIMM50, an essential TIM23 complex subunit, is suggested to facilitate the import of ∼60% of the mitochondrial proteome. In this study, we characterized a TIMM50 disease causing mutation in human fibroblasts and noted significant decreases in TIM23 core protein levels (TIMM50, TIMM17A/B, and TIMM23). Strikingly, TIMM50 deficiency had no impact on the steady state levels of most of its putative substrates, suggesting that even low levels of a functional TIM23 complex are sufficient to maintain the majority of TIM23 complex-dependent mitochondrial proteome. As TIMM50 mutations have been linked to severe neurological phenotypes, we aimed to characterize TIMM50 defects in manipulated mammalian neurons. TIMM50 knockdown in mouse neurons had a minor effect on the steady state level of most of the mitochondrial proteome, supporting the results observed in patient fibroblasts. Amongst the few affected TIM23 substrates, a decrease in the steady state level of components of the intricate oxidative phosphorylation and mitochondrial ribosome complexes was evident. This led to declined respiration rates in fibroblasts and neurons, reduced cellular ATP levels and defective mitochondrial trafficking in neuronal processes, possibly contributing to the developmental defects observed in patients with TIMM50 disease. Finally, increased electrical activity was observed in TIMM50 deficient mice neuronal cells, which correlated with reduced levels of KCNJ10 and KCNA2 plasma membrane potassium channels, likely underlying the patients' epileptic phenotype.

Vgll2 as an integrative regulator of mitochondrial function and contractility specific to skeletal muscle.

During skeletal muscle adaptation to physiological or pathophysiological signals, contractile apparatus and mitochondrial function are coordinated to alter muscle fiber type. Although recent studies have identified various factors involved in modifying contractile proteins and mitochondrial function, the molecular mechanisms coordinating contractile and metabolic functions during muscle fiber transition are not fully understood. Using a gene-deficient mouse approach, our previous studies uncovered that vestigial-like family member 2 (Vgll2), a skeletal muscle-specific transcription cofactor activated by exercise, is essential for fast-to-slow adaptation of skeletal muscle. The current study provides evidence that Vgll2 plays a role in increasing muscle mitochondrial mass and oxidative capacity. Transgenic Vgll2 overexpression in mice altered muscle fiber composition toward the slow type and enhanced exercise endurance, which contradicted the outcomes observed with Vgll2 deficiency. Vgll2 expression was positively correlated with the expression of genes related to mitochondrial function in skeletal muscle, mitochondrial DNA content, and protein abundance of oxidative phosphorylation complexes. Additionally, Vgll2 overexpression significantly increased the maximal respiration of isolated muscle fibers and enhanced the suppressive effects of endurance training on weight gain. Notably, no additional alteration in expression of myosin heavy chain genes was observed after exercise, suggesting that Vgll2 plays a direct role in regulating mitochondrial function, independent of its effect on contractile components. The observed increase in exercise endurance and metabolic efficiency may be attributed to the acute upregulation of genes promoting fatty acid utilization as a direct consequence of Vgll2 activation facilitated by endurance exercise. Thus, the current study establishes that Vgll2 is an integrative regulator of mitochondrial function and contractility in skeletal muscle.

605. MITOCHONDRIAL METABOLISM IN UPPER GI ADENOCARCINOMA DETERMINES PATIENT SURVIVAL

Abstract Background Mitochondrial oxidative phosphorylation (OXPHOS) is responsible for meeting the vast majority of cellular energy demands. Ever since the description of aerobic glycolysis by Warburg in 1927, it has been recognised that cancers have dysfunctional metabolism. Clinically, this Warburg effect is the basis of FDG-PET imaging, increased FDG uptake represents increased tumour glycolysis and translates to worse upper GI adenocarcinoma outcomes. The cause of OXPHOS disruption in upper GI adenocarcinoma is poorly understood, its impact on potential therapeutic strategies is yet to be explored. This is particularly pertinent in upper GI adenocarcinomas as despite recent improvements, 5-year survival remains alarmingly low. Method Oesophageal and gastric adenocarcinoma samples collected in the UK from 175 patients (89 oesophageal adenocarcinoma, 86 gastric adenocarcinoma, 131 male, 44 female, median age=65, IQR=12) who underwent radical resection with curative intent between 1995 and 2009. Quantitative immunofluorescence were used to determine expressions of OXPHOS Complex I and IV at a single cell level. Patients were divided into high vs. low expressors of Complex I and IV using K-means clustering to allow for subsequent subgroup analysis. Result Survival analysis demonstrated significantly worse outcomes for low expressors as compared to high expressors of Complex I at 5 years post-op (5-year survival probability low expressors=0.440, 95%CI=0.358-0.539, high expressors=0.629, 95%CI=0.515-0.769, P-value=0.048). Similarly comparison of Complex IV expression also demonstrates disadvantageous survival for Complex IV low expressors (5-year survival probability low expressors=0.432, 95%CI=0.346-0.538, high expressors=0.603, 95%CI=0.499-0.729, P-value=0.042). The low expressors of Complex I and IV encompassed significantly more Stage III disease as per AJCC 7th edition. The low vs. high expression Complex I and IV groups are similarly matched for baseline patient characteristics. Conclusion These results demonstrate reduced capacity to carry out OXPHOS in oesophageal and gastric adenocarcinoma is characteristic of more advanced cancer and results in worse patient outcomes. Further work is needed to determine if OXPHOS status impacts on neo-adjuvant therapy response and how to exploit OXPHOS deficiency therapeutically in oesophageal and gastric adenocarcinomas. The limitations of this preliminary work include small sample size and tissue derived from historic patients over a long period of time, during which treatment and outcomes for upper GI cancer have significantly changed.

Akt1 deficiency does not affect fiber type composition or mitochondrial protein expression in skeletal muscle of male mice.

Insulin-like growth factor-1-induced activation of ATP citrate lyase (ACLY) improves muscle mitochondrial function through an Akt-dependent mechanism. In this study, we examined whether Akt1 deficiency alters skeletal muscle fiber type and mitochondrial function by regulating ACLY-dependent signaling in male Akt1 knockout (KO) mice (12-16 weeks old). Akt1 KO mice exhibited decreased body weight and muscle wet weight, with reduced cross-sectional areas of slow- and fast-type muscle fibers. Loss of Akt1 did not affect the phosphorylation status of ACLY in skeletal muscle. The skeletal muscle fiber type and expression of mitochondrial oxidative phosphorylation complex proteins were unchanged in Akt1 KO mice compared with the wild-type control. These observations indicate that Akt1 is important for the regulation of skeletal muscle fiber size, whereas the regulation of muscle fiber type and muscle mitochondrial content occurs independently of Akt1 activity.

Airway Epithelium-derived CXCL14 Promotes Eosinophil Accumulation in Allergic Airway Inflammation.

C-X-C motif chemokine ligand 14 (CXCL14) is expressed in the airway epithelial cells of patients with asthma. However, the mechanisms of CXCL14 secretion and its effects on asthma pathogenesis remain unclear. Here, we investigated the role of CXCL14 in allergic airway inflammation and its effects on eosinophil infiltration. Our findings showed that Alternaria alternata, a major environmental allergen, stimulated CXCL14 secretion from airway epithelial cells via reactive oxygen species (ROS) generated in mitochondrial oxidative phosphorylation (OXPHOS) complexes, especially in OXPHOS complex II. In vivo, in a mouse model of allergic airway inflammation, intranasal administration of anti-CXCL14 antibody suppressed eosinophil and dendritic cell infiltration into the airways and goblet cell hyperplasia. In vitro, in human eosinophil-like cells, CXCL14 promoted cell migration through C-X-C chemokine receptor type 4 (CXCR4) binding. Eosinophil CXCR4 expression was upregulated by Alternaria stimulation via ROS production. These findings suggest that the crosstalk between Alternaria-stimulated airway epithelial CXCL14 secretion and eosinophil CXCR4 upregulation plays an important role in eosinophil infiltration into the lungs during allergic airway inflammation. In summary, this study demonstrates that CXCL14 could be a therapeutic target for allergic airway inflammation.

Effects of Atorvastatin and Simvastatin on the Bioenergetic Function of Isolated Rat Brain Mitochondria.

Little is known about the effects of statins, which are cholesterol-lowering drugs, on the bioenergetic functions of mitochondria in the brain. This study aimed to elucidate the direct effects of atorvastatin and simvastatin on the bioenergetics of isolated rat brain mitochondria by measuring the statin-induced changes in respiratory chain activity, ATP synthesis efficiency, and the production of reactive oxygen species (ROS). Our results in isolated brain mitochondria are the first to demonstrate that atorvastatin and simvastatin dose-dependently significantly inhibit the activity of the mitochondrial respiratory chain, resulting in a decreased respiratory rate, a decreased membrane potential, and increased ROS formation. Moreover, the tested statins reduced mitochondrial coupling parameters, the ADP/O ratio, the respiratory control ratio, and thus, the oxidative phosphorylation efficiency in brain mitochondria. Among the oxidative phosphorylation complexes, statin-induced mitochondrial impairment concerned complex I, complex III, and ATP synthase activity. The calcium-containing atorvastatin had a significantly more substantial effect on isolated brain mitochondria than simvastatin. The higher inhibitory effect of atorvastatin was dependent on calcium ions, which may lead to the disruption of calcium homeostasis in mitochondria. These findings suggest that while statins are effective in their primary role as cholesterol-lowering agents, their use may impair mitochondrial function, which may have consequences for brain health, particularly when mitochondrial energy efficiency is critical.

Misregulation of mitochondrial 6mA promotes the propagation of mutant mtDNA and causes aging in C.elegans.

In virtually all eukaryotes, the mitochondrial DNA (mtDNA) encodes proteins necessary for oxidative phosphorylation (OXPHOS) and RNAs required for their synthesis. The mechanisms of regulation of mtDNA copy number and expression are not completely understood but crucially ensure the correct stoichiometric assembly of OXPHOS complexes from nuclear- and mtDNA-encoded subunits. Here, we detect adenosine N6-methylation (6mA) on the mtDNA of diverse animal and plant species. This modification is regulated in C.elegans by the DNA methyltransferase DAMT-1 and demethylase ALKB-1. Misregulation of mtDNA 6mA through targeted modulation of these activities inappropriately alters mtDNA copy number and transcript levels, impairing OXPHOS function, elevating oxidative stress, and shortening lifespan. Compounding these defects, mtDNA 6mA hypomethylation promotes the cross-generational propagation of a deleterious mtDNA. Together, these results reveal that mtDNA 6mA is highly conserved among eukaryotes and regulates lifespan by influencing mtDNA copy number, expression, and heritable mutation levels invivo.

KARS Mutations Impair Brain Myelination by Inducing Oligodendrocyte Deficiency: One Potential Mechanism and Improvement by Melatonin.

It is very crucial to investigate key molecules that are involved in myelination to gain an understanding of brain development and injury. We have reported for the first time that pathogenic variants p.R477H and p.P505S in KARS, which encodes lysyl-tRNA synthetase (LysRS), cause leukoencephalopathy with progressive cognitive impairment in humans. The role and action mechanisms of KARS in brain myelination during development are unknown. Here, we first generated Kars knock-in mouse models throughthe CRISPR-Cas9 system. Kars knock-in mice displayed significant cognitive deficits. These mice also showed significantly reduced myelin density and content, as well as significantly decreased myelin thickness during development. In addition, Kars mutations significantly induced oligodendrocyte differentiation arrest and reduction in the brain white matter of mice. Mechanically, oligodendrocytes' significantly imbalanced expression of differentiation regulators and increased capase-3-mediated apoptosis were observed in the brain white matter of Kars knock-in mice. Furthermore, Kars mutations significantly reduced the aminoacylation and steady-state level of mitochondrial tRNALys and decreased the protein expression of subunits of oxidative phosphorylation complexes in the brain white matter. Kars knock-in mice showed decreased activity of complex IV and significantly reduced ATP production and increased reactive oxygen species in the brain white matter. Significantly increased percentages of abnormal mitochondria and mitochondrion area were observed in the oligodendrocytes of Kars knock-in mouse brain. Finally, melatonin (a mitochondrion protectant) significantly attenuated mitochondrion and oligodendrocyte deficiency in the brain white matter of KarsR504H/P532S mice. The mice treated with melatonin also showed significantly restored myelination and cognitive function. Our study first establishes Kars knock-in mammal models of leukoencephalopathy and cognitive impairment and indicates important roles of KARS in the regulation of mitochondria, oligodendrocyte differentiation and survival, and myelination during brain development and application prospects of melatonin in KARS (or even aaRS)-related diseases.

Alcohol- and Low-Iron Induced Changes in Antioxidant and Energy Metabolism Associated with Protein Lys Acetylation.

Understanding the role of iron in ethanol-derived hepatic stress could help elucidate the efficacy of dietary or clinical interventions designed to minimize liver damage from chronic alcohol consumption. We hypothesized that normal levels of iron are involved in ethanol-derived liver damage and reduced dietary iron intake would lower the damage caused by ethanol. We used a pair-fed mouse model utilizing basal Lieber-DeCarli liquid diets for 22 weeks to test this hypothesis. In our mouse model, chronic ethanol exposure led to mild hepatic stress possibly characteristic of early-stage alcoholic liver disease, seen as increases in liver-to-body weight ratios. Dietary iron restriction caused a slight decrease in non-heme iron and ferritin (FeRL) expression while it increased transferrin receptor 1 (TfR1) expression without changing ferroportin 1 (FPN1) expression. It also elevated protein lysine acetylation to a more significant level than in ethanol-fed mice under normal dietary iron conditions. Interestingly, iron restriction led to an additional reduction in nicotinamide adenine dinucleotide (NAD+) and NADH levels. Consistent with this observation, the major mitochondrial NAD+-dependent deacetylase, NAD-dependent deacetylase sirtuin-3 (SIRT3), expression was significantly reduced causing increased protein lysine acetylation in ethanol-fed mice at normal and low-iron conditions. In addition, the detection of superoxide dismutase 1 and 2 levels (SOD1 and SOD2) and oxidative phosphorylation (OXPHOS) complex activities allowed us to evaluate the changes in antioxidant and energy metabolism regulated by ethanol consumption at normal and low-iron conditions. We observed that the ethanol-fed mice had mild liver damage associated with reduced energy and antioxidant metabolism. On the other hand, iron restriction may exacerbate certain activities of ethanol further, such as increased protein lysine acetylation and reduced antioxidant metabolism. This metabolic change may prove a barrier to the effectiveness of dietary reduction of iron intake as a preventative measure in chronic alcohol consumption.

The Uncoupling Effect of 17β-Estradiol Underlies the Resilience of Female-Derived Mitochondria to Damage after Experimental TBI.

Current literature finds females have improved outcomes over their male counterparts after severe traumatic brain injury (TBI), while the opposite seems to be true for mild TBI. This begs the question as to what may be driving these sex differences after TBI. Estrogen is thought to be neuroprotective in certain diseases, and its actions have been shown to influence mitochondrial function. Mitochondrial impairment is a major hallmark of TBI, and interestingly, this dysfunction has been shown to be more severe in males than females after brain injury. This suggests estrogen could be playing a role in promoting "mitoprotection" following TBI. Despite the existence of estrogen receptors in mitochondria, few studies have examined the direct role of estrogen on mitochondrial function, and no studies have explored this after TBI. We hypothesized ex vivo treatment of isolated mitochondria with 17β-estradiol (E2) would improve mitochondrial function after experimental TBI in mice. Total mitochondria from the ipsilateral (injured) and contralateral (control) cortices of male and female mice were isolated 24 h post-controlled severe cortical impact (CCI) and treated with vehicle, 2 nM E2, or 20 nM E2 immediately before measuring reactive oxygen species (ROS) production, bioenergetics, electron transport chain complex (ETC) activities, and β-oxidation of palmitoyl carnitine. Protein expression of oxidative phosphorylation (OXPHOS) complexes was also measured in these mitochondrial samples to determine whether this influenced functional outcomes with respect to sex or injury. While mitochondrial ROS production was affected by CCI in both sexes, there were other sex-specific patterns of mitochondrial injury 24 h following severe CCI. For instance, mitochondria from males were more susceptible to CCI-induced injury with respect to bioenergetics and ETC complex activities, whereas mitochondria from females showed only Complex II impairment and reduced β-oxidation after injury. Neither concentration of E2 influenced ETC complex activities themselves, but 20 nM E2 appeared to uncouple mitochondria isolated from the contralateral cortex in both sexes, as well as the injured ipsilateral cortex of females. These studies highlight the significance of measuring mitochondrial dysfunction in both sexes after TBI and also shed light on another potential neuroprotective mechanism in which E2 may attenuate mitochondrial dysfunction after TBI in vivo.

A common variant in the iron regulatory gene (Hfe) alters the metabolic and transcriptional landscape in brain regions vulnerable to neurodegeneration.

The role of iron dyshomeostasis in neurodegenerative disease has implicated the involvement of genes that regulate brain iron. The homeostatic iron regulatory gene (HFE) has been at the forefront of these studies given the role of the H63D variant (H67D in mice) in increasing brain iron load. Despite iron's role in oxidative stress production, H67D mice have shown robust protection against neurotoxins and improved recovery from intracerebral hemorrhage. Previous data support the notion that H67D mice adapt to the increased brain iron concentrations and hence develop a neuroprotective environment. This adaptation is particularly evident in the lumbar spinal cord (LSC) and ventral midbrain (VM), both relevant to neurodegeneration. We studied C57BL6/129 mice with homozygous H67D compared to WT HFE. Immunohistochemistry was used to analyze dopaminergic (in the VM) and motor (in the LSC) neuron population maturation in the first 3 months. Immunoblotting was used to measure protein carbonyl content and the expression of oxidative phosphorylation complexes. Seahorse assay was used to analyze metabolism of mitochondria isolated from the LSC and VM. Finally, a Nanostring transcriptomic analysis of genes relevant to neurodegeneration within these regions was performed. Compared to WT mice, we found no difference in the viability of motor neurons in the LSC, but the dopaminergic neurons in H67D mice experienced significant decline before 3 months of age. Both regions in H67D mice had alterations in oxidative phosphorylation complex expression indicative of stress adaptation. Mitochondria from both regions of H67D mice demonstrated metabolic differences compared to WT. Transcriptional differences in these regions of H67D mice were related to cell structure and adhesion as well as cell signaling. Overall, we found that the LSC and VM undergo significant and distinct metabolic and transcriptional changes in adaptation to iron-related stress induced by the H67D HFE gene variant.

Joint and Individual Mitochondrial DNA Variation and Cognitive Outcomes in Black and White Older Adults.

Mitochondrial dysfunction manifests in neurodegenerative diseases and other age-associated disorders. In this study, we examined variation in inherited mitochondrial DNA (mtDNA) sequences in Black and White participants from 2 large aging studies to identify variants related to cognitive function. Participants included self-reported Black and White adults aged ≥70 years in the Lifestyle Interventions and Independence for Elders (LIFE; N = 1319) and Health Aging and Body Composition (Health ABC; N = 788) studies. Cognitive function was measured by the Digit-Symbol Substitution Test (DSST), and the Modified Mini-Mental State Examination (3MSE) at baseline and over follow-up in LIFE (3.6 years) and Health ABC (10 years). We examined the joint effects of multiple variants across 16 functional mitochondrial regions with cognitive function using a sequence kernel association test. Based on these results, we prioritized meta-analysis of common variants in Black and White participants using mixed effects models. A Bonferroni-adjusted p value of <.05 was considered statistically significant. Joint variation in subunits ND1, ND2, and ND5 of Complex I, 12S RNA, and hypervariable region (HVR) were significantly associated with DSST and 3MSE at baseline. In meta-analyses among Black participants, variant m.4216T>C, ND1 was associated with a faster decline in 3MSE, and variant m.462C>T in the HVR was associated with a slower decline in DSST. Variant m.5460G>C, ND2 was associated with slower and m.182C>T in the HVR was associated with faster decline in 3MSE in White participants. Among Black and White adults, oxidative phosphorylation Complex I variants were associated with cognitive function.

A roadmap for ribosome assembly in human mitochondria.

Mitochondria contain dedicated ribosomes (mitoribosomes), which synthesize the mitochondrial-encoded core components of the oxidative phosphorylation complexes. The RNA and protein components of mitoribosomes are encoded on two different genomes (mitochondrial and nuclear) and are assembled into functional complexes with the help of dedicated factors inside the organelle. Defects in mitoribosome biogenesis are associated with severe human diseases, yet the molecular pathway of mitoribosome assembly remains poorly understood. Here, we applied a multidisciplinary approach combining biochemical isolation and analysis of native mitoribosomal assembly complexes with quantitative mass spectrometry and mathematical modeling to reconstitute the entire assembly pathway of the human mitoribosome. We show that, in contrast to its bacterial and cytosolic counterparts, human mitoribosome biogenesis involves the formation of ribosomal protein-only modules, which then assemble on the appropriate ribosomal RNA moiety in a coordinated fashion. The presence of excess protein-only modules primed for assembly rationalizes how mitochondria cope with the challenge of forming a protein-rich ribonucleoprotein complex of dual genetic origin. This study provides a comprehensive roadmap of mitoribosome biogenesis, from very early to late maturation steps, and highlights the evolutionary divergence from its bacterial ancestor.

Mitochondrion-to-nucleus communication mediated by RNA export: a survey of potential mechanisms and players across eukaryotes.

The nucleus interacts with the other organelles to perform essential functions of the eukaryotic cell. Mitochondria have their own genome and communicate back to the nucleus in what is known as mitochondrial retrograde response. Information is transferred to the nucleus in many ways, leading to wide-ranging changes in nuclear gene expression and culminating with changes in metabolic, regulatory or stress-related pathways. RNAs are emerging molecules involved in this signalling. RNAs encode precise information and are involved in highly target-specific signalling, through a wide range of processes known as RNA interference. RNA-mediated mitochondrial retrograde response requires these molecules to exit the mitochondrion, a process that is still mostly unknown. We suggest that the proteins/complexes translocases of the inner membrane, polynucleotide phosphorylase, mitochondrial permeability transition pore, and the subunits of oxidative phosphorylation complexes may be responsible for RNA export.