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  • Calcium Overload Decreases Net Free Radical Emission in Cardiac Mitochondria
    Mitochondrion (IF 3.449) Pub Date : 2020-01-23
    Quynh V. Duong; Adrianna Hoffman; Katie Zhong; Maria J. Dessinger; Yizhu Zhang; Jason N. Bazil

    Elevated calcium and reactive oxygen species (ROS) are responsible for the bulk of cell death occurring in a variety of clinical settings that include acute coronary events, cerebrovascular accidents, and acute kidney injury. It is commonly believed that calcium and ROS participate in a viscous cycle during these events. However, the precise feedback mechanisms responsible are unknown. In this study, we quantitatively demonstrate that, on the contrary, calcium does not stimulate free radical production but suppresses it. Isolated mitochondria from guinea pig hearts were energized with a variety of substrates and exposed to calcium concentrations designed to induce moderate calcium overload conditions associated with ischemia/reperfusion injury but do not elicit the well-known mitochondrial permeability transition phenomenon. Metabolic function and free radical emission were simultaneously quantified using high-resolution respirometry and fluorimetry. In parallel, membrane potential, high amplitude swelling, and calcium dynamics were also quantified. Our results reveal that calcium overload does not lead to excessive ROS emission but does decrease ADP stimulated respiration rates for NADH-dependent pathways. Moreover, we developed an empirical model of mitochondrial free radical homeostasis to identify the processes that are different for each substrate and calcium condition. In summary, we show that in healthy guinea pig mitochondria, calcium uptake and free radical generation do not contribute to a viscous cycle and that the relationship between net free radical production and oxygen concentration is hyperbolic. Altogether, these results lay out an important foundation necessary to quantitatively determine the role of calcium in IR injury and ROS production.

  • Redox Homeostasis, Oxidative Stress and Mitophagy
    Mitochondrion (IF 3.449) Pub Date : 2020-01-20
    Carla Garza-Lombó; Aglaia Pappa; Mihalis I. Panayiotidis; Rodrigo Franco

    Autophagy is a ubiquitous homeostatic mechanism for the degradation or turnover of cellular components. Degradation of mitochondria via autophagy (mitophagy) is involved in a number of physiological processes including cellular homeostasis, differentiation and aging. Upon stress or injury, mitophagy prevents the accumulation of damaged mitochondria and the increased steady state levels of reactive oxygen species leading to oxidative stress and cell death. A number of human diseases, particularly neurodegenerative disorders, have been linked to the dysregulation of mitophagy. In this mini-review, we aimed to review the molecular mechanisms involved in the regulation of mitophagy and their relationship with redox signaling and oxidative stress.

  • MMPdb and MitoPredictor: tools for facilitating comparative analysis of animal mitochondrial proteomes
    Mitochondrion (IF 3.449) Pub Date : 2020-01-20
    Viraj Muthye; Gaurav Kandoi; Dennis Lavrov

    Comparative analysis of animal mitochondrial proteomes faces two challenges: the scattering of data on experimentally-characterized animal mitochondrial proteomes across several databases, and the lack of data on mitochondrial proteomes from the majority of metazoan lineages. In this study, we developed two resources to address these challenges: 1] the Metazoan Mitochondrial Proteome Database (MMPdb), which consolidates data on experimentally-characterized mitochondrial proteomes of vertebrate and invertebrate model organisms, and 2] MitoPredictor, a novel machine-learning tool for prediction of mitochondrial proteins in animals. MMPdb allows comparative analysis of animal mitochondrial proteomes by integrating results from orthology analysis, prediction of mitochondrial targeting signals, protein domain analysis, and Gene Ontology analysis. Additionally, for mammalian mitochondrial proteins, MMPdb includes experimental evidence of localization from MitoMiner and the Human Protein Atlas. MMPdb is publicly available at https://mmpdb.eeob.iastate.edu/. MitoPredictor is a Random Forest classifier which uses orthology, mitochondrial targeting signal prediction and protein domain content to predict mitochondrial proteins in animals.

  • Detection of mitochondrial DNA variants at low level heteroplasmy in pediatric CNS and extra-CNS solid tumors with three different enrichment methods
    Mitochondrion (IF 3.449) Pub Date : 2020-01-20
    Kristiyana Kaneva; Daria Merkurjev; Dejerianne Ostrow; Alex Ryutov; Petr Triska; Kevin Stachelek; David Cobrinik; Jaclyn A. Biegel; Xiaowu Gai

    The mitochondrial genome is small, 16.5kb, and yet complex to study due to an abundance of mitochondria in any given cell or tissue. Mitochondrial DNA (mtDNA) mutations have been previously described in cancer, many of which were detected at low heteroplasmy. In this study we enriched the mitochondrial genome in primary pediatric tumors for detection of mtDNA variants. We completed mtDNA enrichment using REPLI-g, Agilent SureSelect, and long-range polymerase chain reaction (LRPCR) followed by next generation sequencing (NGS) on Illumina platforms. Primary tumor and germline genomic DNA from a variety of pediatric central nervous system (CNS) and extra-CNS solid tumors were analyzed by the three different methods. Although all three methods performed equally well for detecting variants at high heteroplasmy or homoplasmy, only LRPCR and SureSelect-based enrichment methods provided consistent results for variants that were present at less than five percent heteroplasmy. We then applied both LRPCR and SureSelect to three successive samples from a patient with multiply-recurrent gliofibroma and detected a low-level novel mutation as well as a change in heteroplasmy levels of a synonymous variant that was correlated with progression of disease. Implication: This study demonstrates that LRPCR and SureSelect enrichment, but not REPLI-g, followed by NGS are accurate methods for studying the mtDNA variations at low heteroplasmy, which may be applied to studying mtDNA mutations in cancer.

  • Relative Telomere Length and Mitochondrial DNA Copy Number Variation with Age: Association with Plasma Folate and Vitamin B12
    Mitochondrion (IF 3.449) Pub Date : 2020-01-11
    Guruvaiah Praveen; Tattari Shalini; Mudili Sivaprasad; G. Bhanuprakash Reddy

    Telomere attrition and mitochondrial DNA variations are implicated in the biological aging process and genomic stability can be influenced by nutritional factors. This study aims to analyze the relative telomere length (rTL) and mitochondrial DNA copy number (mtCN) in aged individuals and their association with plasma folate and vitamin B12 levels. This community-based cross-sectional study involves 428 subjects (<60 years: 242 & ≥60 years: 186). Quantitative real-time PCR was used to measure rTL and mtCN variation, and radioimmunoassay to measure plasma folate and vitamin B12 levels. The subjects in the ≥60 years age group have significantly shorter telomeres and lower mtCN compared to the <60 years age group. A significant positive correlation was observed between the rTL and mtCN, and both of them were positively associated with plasma folate and vitamin B12 levels. In the ≥60 age group; folate and vitamin B12 positively correlated with rTL and vitamin B12 with mtCN. The study revealed a decline of rTL and mtCN with age in the Indian population and their association suggests that they may co-regulate each other with age. In conclusion, folate and vitamin B12 may delay aging by preventing the reduction in rTL length and mtCN.

  • Revealing mitogenome-wide DNA methylation and RNA editing of three Ascomycotina fungi using SMRT sequencing
    Mitochondrion (IF 3.449) Pub Date : 2020-01-07
    Chaoxia Wang; Jianhua Feng; Yujiao Chen; Dongmei Li; Li Liu; Yuqian Wu; Shujun Zhang; Simiao Du; Yaozhou Zhang

    Beauveria bassiana, Cordyceps militaris and Ophiocordyceps sinensis (Ascomycotina) are traditional Chinese medicines. Here, mitogenomes of these three Ascomycotina fungi were sequenced and de-novo assembled using single-molecule real-time sequencing. The results showed that their complete mitogenomes were 31,258, 31,854 and 157,584 bp, respectively, with sequencing depth approximately 278,760×, 326,283× and 69,385×. Types of repeat sequences were mainly (AA)n, (AAT)n, (TA)n and (TATT)n. DNA methylation motifs were revealed in DNA modifications of these three fungi. We discovered new models of RNA editing through analysis of transcriptomes from B. bassiana and C. militaris. These data lay a solid foundation for further genetic and biological studies about these three fungi, especially for elucidating the mitogenome evolution and exploring the regulatory mechanism of adapting environment.

  • LONP1 de novo dominant mutation causes mitochondrial encephalopathy with loss of LONP1 chaperone activity and excessive LONP1 proteolytic activity
    Mitochondrion (IF 3.449) Pub Date : 2020-01-07
    Arnaud Besse; Daniel Brezavar; Jennifer Hanson; Austin Larson; Penelope E Bonnen

    LONP1 is an ATP-dependent protease and chaperone that plays multiple vital roles in mitochondria. LONP1 is essential for mitochondrial homeostasis due to its role in maintenance of the mitochondrial genome and its central role in regulating mitochondrial processes such as oxidative phosphorylation, mitophagy, and heme biosynthesis. Bi-allelic LONP1 mutations have been reported to cause a constellation of clinical presentations. We report a patient heterozygous for a de novo mutation in LONP1: c.901C>T,p.R301W presenting as a neonate with seizures, encephalopathy, pachygyria and microcephaly. Assays of respiratory chain activity in muscle showed complex II-III function at 8% of control. Functional studies in patient fibroblasts showed a signature of dysfunction that included significant decreases in known proteolytic targets of LONP1 (TFAM, PINK1, phospho-PDH E1α) as well as loss of mitochondrial ribosome subunits MRPL44 and MRPL11 with concomitant decreased activity and level of protein subunits of oxidative phosphorylation complexes I and IV. These results indicate excessive LONP1 proteolytic activity and a loss of LONP1 chaperone activity. Further, we demonstrate that the LONP1 N-terminal domain is involved in hexamer stability of LONP1 and that the ability to make conformational changes is necessary for LONP1 to regulate proper functioning of both its proteolytic and chaperone activities.

  • 更新日期:2019-12-25
  • Perspective: Cell Danger Response Biology—The New Science that Connects Environmental Health with Mitochondria and the Rising Tide of Chronic Illness
    Mitochondrion (IF 3.449) Pub Date : 2019-12-23
    Robert K. Naviaux

    This paper is written for non-specialists in mitochondrial biology to provide access to an important area of science that has broad implications for all people. The cell danger response (CDR) is a universal response to environmental threat or injury. Once triggered, healing cannot be completed until the choreographed stages of the CDR are returned to an updated state of readiness. Although the CDR is a cellular response, it has the power to change human thought and behavior, child development, physical fitness and resilience, fertility, and the susceptibility of entire populations to disease. Mitochondria regulate the CDR by monitoring and responding to the physical, chemical, and microbial conditions within and around the cell. In this way, mitochondria connect cellular health to environmental health. Over 7,000 chemicals are now made or imported to the US for industrial, agricultural, and personal care use in amounts ranging from 25,000 to over 1 million pounds each year, and plastic waste now exceeds 83 billion pounds/year. This chemical load creates a rising tide of manmade pollutants in the oceans, air, water, and food chain. Fewer than 5% of these chemicals have been tested for developmental toxicity. In the 1980s, 5-10% of children lived with a chronic illness. As of 2018, 40% of children, 50% of teens, 60% of adults under age 65, and 90% of adults over 65 live with chronic illness. Several studies now report the presence of dozens to hundreds of manmade chemicals and pollutants in placenta, umbilical cord blood, and newborn blood spots. New methods in metabolomics and exposomics allow scientists to measure thousands of chemicals in blood, air, water, soil, and the food chain. Systematic measurements of environmental chemicals can now be correlated with annual and regional patterns of childhood illness. These data can be used to prepare a prioritized list of molecules for congressional action, ranked according to their impact on human health. “When a deep injury is done to us, we never heal until we forgive.” --Nelson Mandela (1918-2013)

  • OPA1 regulates respiratory supercomplexes assembly: the role of mitochondrial swelling
    Mitochondrion (IF 3.449) Pub Date : 2019-12-20
    Sehwan Jang; Sabzali Javadov

    Optic atrophy type 1 protein (OPA1), a dynamin-related GTPase, that, in addition to mitochondrial fusion, plays an important role in maintaining the structural organization and integrity of the inner mitochondrial membrane (IMM). OPA1 exists in two forms: IMM-bound long-OPA1 (L-OPA1) and soluble short-OPA1 (S-OPA1), a product of L-OPA1 proteolytic cleavage localized in the intermembrane space. In addition to OPA1, the structural and functional integrity of IMM can be regulated by changes in the matrix volume due to the opening/closure of permeability transition pores (PTP). Herein, we investigated the crosstalk between the PTP and OPA1 to clarify whether PTP opening is involved in OPA1-mediated regulation of respiratory chain supercomplexes (RCS) assembly using cardiac mitochondria and cell line. We found that: 1) Proteolytic cleavage of L-OPA1 is stimulated by PTP-induced mitochondrial swelling, 2) OPA1 knockdown reduces PTP-induced mitochondrial swelling but enhances ROS production, 3) OPA1 deficiency impairs the RCS assembly associated with diminished ETC activity and oxidative phosphorylation, 4) OPA1 has no physical interaction with phospholipid scramblase 3 although OPA1 downregulation increases expression of the scramblase. Thus, this study demonstrates that L-OPA1 cleavage depends on the PTP-induced mitochondrial swelling suggesting a regulatory role of the PTP-OPA1 axis in RCS assembly and mitochondrial bioenergetics.

  • Positive regulation of human PINK1 and Parkin gene expression by Nuclear respiratory factor 1
    Mitochondrion (IF 3.449) Pub Date : 2019-12-18
    Yapeng Lu; Wangwang Ding; Bo Wang; Lu Wang; Huiwen Kan; Xueting Wang; Dan Wang; Li Zhu

    Previous studies have demonstrated that two Parkinson's disease-associated genes PINK1 and Parkin play a key role in mitochondrial quality control. But until now, the transcriptional regulation of these two genes under normal physiological conditions are not well understood. In this study, the transcriptional regulation of PINK1 and Parkin genes by nuclear respiratory factor 1 (NRF-1) and its effect on PINK1/ Parkin-mediated mitophagy were studied. The NRF-1 binding sites in the promoter regions of human PINK1 and Parkin genes were analyzed by JASPER software and were confirmed by chromatin immunoprecipitation (ChIP) assay. The transcriptional activities and the expressions of PINK1 and Parkin genes were positively regulated by NRF-1 in HEK293T cells and in SH-SY5Y cells. Furthermore, NRF-1 over-expression (OE) up-regulated the protein level of full-length PINK1 in CCCP-treated cells, indicating the enhanced PINK1/Parkin-mediated mitophagy. When NRF-1 expression was transient or stable knockdown, the CCCP-induced mitophagy was alleviated as characterized by the reduced protein level of full-length PINK1, the declined ratio of LC3 II to LC3 I, and the decreased ratio of Mt -keima fluorescence intensity excited at 552 nm to that excited at 488 nm. In conclusion, NRF-1 has a positive regulatory effect on the transcription of PINK1 and Parkin genes, and involves in mitochondrial quality control through regulating PINK1/Parkin-mediated mitophagy.

  • Genomic profiling of Mitochondrial DNA reveals novel complex gene mutations in familial type 2 diabetes mellitus individuals from Mizo ethnic population, Northeast India
    Mitochondrion (IF 3.449) Pub Date : 2019-12-17
    Freda Lalrohlui; John Zohmingthanga; Vanlal hruaii; Nachimuthu Senthil Kumar

    The variants reported for mitochondrial DNA (mtDNA) and type 2 diabetes (T2D) may not be accountable for the disease in certain other populations and the risk depends upon numerous factors which may include genetics, environment as well as ethnicity. This leads to a challenge in identifying, exploring and comparing the variants between diabetic cases and healthy controls in a remote unexplored tribal population. To study the possible contribution of mtDNA variants, we sequenced the entire mitochondrial genomes and the frequencies of mtSNPs, their association with familial T2D and the potential impact of non-synonymous substitutions on protein functions were determined. The mtSNP 8584 G>A (ATP6: A20T) was detected in 14.28% of the diabetic patients and none in the control groups. The mitochondrial ND3 variant 10398A>G was found to be significantly associated with the risk of T2D (OR=9.489, 95% CI=1.161-77.54, P value=0.036). A novel Frame-shift substitution ND5: 81_81ins A at position 12417 was observed in 53.57% of diabetic individuals. Majority of the variants lie in tRNA-Phe in the non-protein coding region of mtDNA for both diabetic cases and common cases. We concluded that mutations in the coding (synonymous or non-synonymous) and noncoding regions of the mitochondria might have contribution towards the development of T2D. Our study is the first to report the distinct mitochondrial variants which may be attributed to the susceptibility as well as development of type 2 diabetes in an ethnic tribe from northeast India.

  • Decreased membrane cholesterol in liver mitochondria of the point mutation mouse model of juvenile Niemann-Pick C1, Npc1nmf164
    Mitochondrion (IF 3.449) Pub Date : 2019-12-17
    Robert P Erickson; Siddhesh Aras; Neeraja Purandare; Maik Hüttemann; Jenney Liu; Jessica Dragotto; Maria Teresa Fiorenza; Lawrence I. Grossman

    It has long been known that there is decreased mitochondrial function in several tissues of Niemann-Pick C1 model mice and cultured cells. These defects contribute to the accumulation of Reactive Oxygen Species (ROS) and tissue damage. It is also well established that there is increased unesterified cholesterol, stored in late endosomes/lysosomes, in many tissues in mutant humans, mouse models, and mutant cultured cells. Using a mouse model with an NPC1 point mutation that is more typical of the most common form of the disease, and highly purified liver mitochondria, we find markedly decreased mitochondrial membrane cholesterol. This is compared to previous reports of increased mitochondrial membrane cholesterol. We also find that, although in wild-type or heterozygous mitochondria cytochrome c oxidase (COX) activity decreases with age as expected, surprisingly, COX activity in homozygous mutant mice improves with age. COX activity is less than half of wild-type amounts in young mutant mice but later reaches wild-type levels while total liver cholesterol is decreasing. Mutant mice also contain a decreased number of mitochondria that are morphologically abnormal. We suggest that the decreased mitochondrial membrane cholesterol is causative for the mitochondrial energy defects. In addition, we find that the mitochondrial stress regulator protein MNRR1 can stimulate NPC1 synthesis and is deficient in mutant mouse livers. Furthermore, the age curve of MNRR1 deficiency paralleled levels of total cholesterol. The role of such altered mitochondria in initiating the abnormal autophagy and neuroinflammation found in NPC1 mouse models is discussed.

  • Mitochondrial mRNA Fragments are Circularized in a Human HEK Cell Line
    Mitochondrion (IF 3.449) Pub Date : 2019-12-09
    Landon G. Mance, Ishaat Mawla, Steven M. Shell, A. Bruce Cahoon

    The relatively recent focus on the widespread occurrence and abundance of circular RNAs (circRNA) in the human cell nucleus has sparked an intensive interest in their existence and possible roles in cell gene expression and physiology. The presence of circRNAs in mammalian mitochondria, however, has been under-explored. Mitochondrial mRNAs differ from those produced from nuclear genes because they lack introns and are transcribed as poly-cistronic transcripts that are endonucleolytically cleaved, leaving transcripts with very small 5’ and 3’ UTRs. Circular RNAs have been identified in the semi-autonomous organelles of single-celled organisms and plants but their purpose has not been clearly demonstrated. The goal of our project was to test the hypothesis, processed mRNAs are circularized in vertebrate mitochondria as a necessary RNA processing step prior to translation. Mitochondrial mRNAs were isolated from the human cell line HEK293 and evidence of circularization sought by treating RNA with RNAse-R and then amplifying putative 3’-5’ junction sites. Sequence results demonstrated the occurrence of mRNA circularization within each coding region of the mitochondrial genome. However, in most cases the circRNAs carried coding regions that had been truncated, suggesting they were not translatable. Quantification of the circularized versions of the mRNAs revealed they comprise a small portion (∼10%) of the total mRNA. These findings demonstrate that mRNA circularization occurs in mammalian mitochondria but it does not appear to play a role in making translatable mRNAs.

  • Mitochondrial Function and Brain Metabolic Score (BMS) in Ischemic Stroke: Evaluation of “Neuroprotectants” Safety and Efficacy
    Mitochondrion (IF 3.449) Pub Date : 2019-11-29
    Avraham Mayevsky, Hofit Kutai-Asis, Michael Tolmasov

    The initial and significant event developed in ischemic stroke is the sudden decrease in blood flow and oxygen supply to brain tissue, leading to dysfunction of the mitochondria. Many attempts were and are being made to develop new drugs and treatments that will save the ischemic brain, but the efficacy is not optimal and in many patients, irreversible damage to the brain will persist. We review a unique approach to evaluate mitochondrial function and microcirculatory hemodynamic in real time in vivo. Three out of four monitored physiological parameters are integrated into a new Brain Metabolic Score (BMS) calculated in real time and is correlated to Brain Oxygen Balance. The technology was adapted to various experimental as well as clinical situations for monitoring the brain in real time. The developed protocols could be used in testing the efficacy and safety of new drugs in experimental animals. Few models of brain monitoring during partial or complete ischemia were developed and used in naive animals or under brain activation protocols. It was found that mitochondrial function/dysfunction is the major and dominant parameter affecting the calculated Brain Metabolic Score. Using our monitoring system and protocols will provide direct information regarding the ability of the tested brain to provide enough oxygen consumed by the mitochondria in the “resting” or in the “activated” brain in vivo and in real-time. Preliminary studies, indicated that testing the efficacy and safety of new neuroprotectant drugs provided significant results to the R&D studies of ischemic stroke related to mitochondrial function.

  • Non-invasive versus ex vivo measurement of mitochondrial function in an endotoxemia model in rat: toward monitoring of mitochondrial therapy
    Mitochondrion (IF 3.449) Pub Date : 2019-11-23
    Mark A. Wefers Bettink, Floor A. Harms, Nathalie Dollee, Patricia A.C. Specht, Nicolaas J.H. Raat, G.C. Schoonderwoerd, Egbert G. Mik

    Mitochondrial function has been predominantly measured ex vivo. Due to isolation and preservation procedures ex vivo measurements might misrepresent in vivo mitochondrial conditions. Direct measurement of in vivo mitochondrial oxygen tension (mitoPO2) and oxygen disappearance rate (ODR) with the protoporphyrin IX‐triplet state lifetime technique (PpIX-TSLT) might increase our understanding of mitochondrial dysfunction in the pathophysiology of acute disease. LPS administration decreased mitochondrial respiration (ODR) in vivo but did not alter mitochondrial function as assessed with ex vivo techniques (high resolution respirometry and specific complex determinations). PpIX-TSLT measures in vivo mitoPO2 and ODR and can be applied non-invasively at the skin.

  • Cerebrospinal fluid ATP as a potential biomarker in patients with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke like episodes (MELAS)
    Mitochondrion (IF 3.449) Pub Date : 2019-11-19
    Takamasa Nukui, Atsushi Matsui, Hideki Niimi, Mamoru Yamamoto, Noriyuki Mastuda, Jin-Lan Piao, Kyo Noguchi, Isao Kitajima, Yuji Nakastuji

    Mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) is caused by defective oxidative phosphorylation in the cerebral parenchyma, cerebral blood vessels, and leptomeningeal tissue. Although increased serum and cerebrospinal fluid (CSF) lactate level has been used as a diagnostic biomarker in patients with MELAS, no biomarkers reflecting disease activity exist. Since we have developed a highly sensitive ATP assay system using luciferase luminous reaction, we examined CSF ATP in patients with MELAS and found that it negatively correlates with disease activity and that it reflects the efficacy of the treatment. CSF ATP might be a novel disease monitoring marker for MELAS.

  • Cell Stress Management by the Mitochondrial LonP1 Protease-nsights into Mitigating Developmental, Oncogenic, Cardiac Stress
    Mitochondrion (IF 3.449) Pub Date : 2019-11-19
    Sundararajan Venkatesh, Carolyn K. Suzuki

    Mitochondrial LonP1 is an essential stress response protease that mediates mitochondrial proteostasis, metabolism and bioenergetics. Homozygous and compound heterozygous variants in the LONP1 gene encoding the LonP1 protease have recently been shown to cause a diverse spectrum of human pathologies, ranging from classical mitochondrial disease phenotypes, in addition to profound neurologic impairment and multi-organ dysfunctions, some of which are uncommon to mitochondrial disorders. In this review, we focus primarily on human LonP1 and discuss findings, which demonstrate its multidimensional roles in maintaining mitochondrial proteostasis and adapting cells to metabolic flux and stress during normal physiology and disease processes. We also discuss emerging roles of LonP1 in responding to developmental, oncogenic and cardiac stress.

  • Hydrogen Sulfide Attenuates Hyperhomocysteinemia-Induced Mitochondrial Dysfunctions in Brain
    Mitochondrion (IF 3.449) Pub Date : 2019-11-18
    Mohit Kumar, Rajat Sandhir

    Hyperhomocysteinemia (HHcy) has been implicated in development of neurodegenerative conditions and mild cognitive disorders. Mitochondrial dysfunctions are the major mechanisms involved in homocysteine (Hcy)-induced neurotoxicity. Although, hydrogen sulfide has been reported as potent antioxidant, its effects on Hcy-induced mitochondrial dysfunctions have not been studied. Therefore, the present study has been designed to evaluate the protective effect of NaHS on Hcy-induced mitochondrial dysfunctions in brain. NaHS supplementation decreased reactive oxygen species and nitrite levels in the cortex and hippocampus of animals with HHcy. NaHS supplementation increased the activity of mitochondrial electron transport chain components; NADH dehydrogenase, cytochrome c oxidase and F0-F1 ATPase in the cortex and hippocampus of HHcy animals along with in-gel activity of complex I – complex V in the mitochondria isolated from the cortex and hippocampus of HHcy animals. Moreover, NaHS supplementation also increased the mitochondrial complex I, II and IV mediated oxygen consumption rate in Hcy treated mitochondria. NaHS administration prevented the Hcy-induced mitochondrial damage as suggested by the deceased mitochondrial swelling in the cortex and hippocampus of HHcy animals. NaHS supplementation decreased the activity of lactate dehydrogenase isozymes (1-5) in the brain regions of HHcy animals. The expression of protein kinase c δ was also decreased in the brain regions of HHcy animals following NaHS supplementation. This was accompanied by reduced activity of caspase-3 indicating anti-apoptotic effect of H2S. Taken together, the findings suggest that H2S supplementation ameliorates Hcy-induced oxidative stress and mitochondrial dysfunctions suggesting it be a novel therapeutic to treat HHcy associated neurological disorders.

  • Analysis of secondary mtDNA mutations in families with Leber’s hereditary optic neuropathy: four novel variants and their association with clinical presentation
    Mitochondrion (IF 3.449) Pub Date : 2019-11-16
    Jasna Jancic, Branislav Rovcanin, Vesna Djuric, Ana Pepic, Janko Samardzic, Blazo Nikolic, Ivana Novakovic, Vladimir S. Kostic

    Leber’s hereditary optic neuropathy (LHON) is a mitochondrial disease characterized by subacute optic atrophy which results in severe visual impairment. The penetrance, clinical expression and disease onset are variable, and frequently associated with other extraocular symptoms. The disease phenotype remains to be an intriguing question which is dependent upon primary as well as secondary mtDNA mutations. In this study we analyzed the whole mtDNA sequence in six LHON families from Serbian population. The mtDNA sequencing was performed by Sanger’s method and various bioinformatic tools were used for analysis of detected mutations. LHON patients carry all three (m.3460G>A, m.11778G>A and m.14484T>C) primary mutations, together with numerous secondary mtDNA mutations. Four novel mutations (m.4516G>A, m.8779C>T, m.13138G>A and m.15986insG) in four different families were discovered. The m.8779C>T and m.13138G>A mutations could have a potential influence on LHON symptoms, but the issue of effect of secondary mtDNA mutations in LHON patients needs to be better clarified in future studies.

  • Exploring the links between lipid geometry and mitochondrial fission: Emerging concepts
    Mitochondrion (IF 3.449) Pub Date : 2019-07-25
    Ashutosh Agrawal, Rajesh Ramachandran

    Mitochondria, the double membrane-walled powerhouses of the eukaryotic cell, are also the seats of synthesis of two critical yet prevalent nonbilayer-prone phospholipids, namely phosphatidylethanolamine (PE) and cardiolipin (CL). Besides their established biochemical roles in the regulation of partner protein function, PE and CL are also key protagonists in the biophysics of mitochondrial membrane remodeling and dynamics. In this review, we address lipid geometry and behavior at the single-molecule level as well as their intimate coupling to whole organelle morphology and remodeling during the concerted events of mitochondrial fission. We present evidence from recent experimental measurements ably supported and validated by computational modeling studies to support our notion that conical lipids play a catalytic as well as a structural role in mitochondrial fission.

  • Physics-based oligomeric models of the yeast mitofusin Fzo1 at the molecular scale in the context of membrane docking
    Mitochondrion (IF 3.449) Pub Date : 2019-07-12
    Astrid Brandner, Dario De Vecchis, Marc Baaden, Mickael M. Cohen, Antoine Taly

    Tethering and homotypic fusion of mitochondrial outer membranes is mediated by large GTPases of the dynamin-related proteins family called the mitofusins. The yeast mitofusin Fzo1 forms high molecular weight complexes and its assembly during membrane fusion likely involves the formation of high order complexes. Consistent with this possibility, mitofusins form oligomers in both cis (on the same lipid bilayer) and trans to mediate membrane attachment and fusion. Here, we utilize our recent Fzo1 model to investigate and discuss the formation of cis and trans mitofusin oligomers. We have built three distinct cis-assembly Fzo1 models that gave rise to three distinct trans-oligomeric models of mitofusin constructs. Each model involves two main components of mitofusin oligomerization: the GTPase and the trunk domains. The oligomeric models proposed in this study were further assessed for stability and dynamics in a membrane environment using a coarse-grained molecular dynamics (MD) simulation approach. A narrow opening ‘head-to-head’ cis-oligomerization (via the GTPase domain) followed by the antiparallel ‘back-to-back’ trans-associations (via the trunk domain) appears to be in agreement with all of the available experimental data. More broadly, this study opens new possibilities to start exploring cis and trans conformations for Fzo1 and mitofusins in general.

  • Dynamic of mitochondrial network, cristae, and mitochondrial nucleoids in pancreatic β-cells
    Mitochondrion (IF 3.449) Pub Date : 2019-06-25
    Petr Ježek, Andrea Dlasková

    Type 2 diabetes progression stems from dysfunction of β-cells, besides the peripheral insulin resistance. Mitochondria as glucose sensor and regulation center are impaired at various stages of this progression. Their biogenesis and functional impairment is reflected by altered morphology of the mitochondrial network and ultramorphology of cristae and mitochondrial DNA loci, termed nucleoids. Aspects of all above changes are reviewed here together with a brief introduction to proteins involved in mitochondrial network dynamics, cristae shaping, and mtDNA nucleoid structure and maintenance. Most frequently, pathology is reflected by the fragmentation of network, cristae inflation or absence and declining number of nucleoids.

  • Mitochondrial dynamics and their potential as a therapeutic target
    Mitochondrion (IF 3.449) Pub Date : 2019-06-19
    B.N. Whitley, E.A. Engelhart, S. Hoppins

    Mitochondrial dynamics shape the mitochondrial network and contribute to mitochondrial function and quality control. Mitochondrial fusion and division are integrated into diverse cellular functions and respond to changes in cell physiology. Imbalanced mitochondrial dynamics are associated with a range of diseases that are broadly characterized by impaired mitochondrial function and increased cell death. In various disease models, modulating mitochondrial fusion and division with either small molecules or genetic approaches has improved function. Although additional mechanistic understanding of mitochondrial fusion and division will be critical to inform further therapeutic approaches, mitochondrial dynamics represent a powerful therapeutic target in a wide range of human diseases.

  • Linking mitochondrial dynamics, cristae remodeling and supercomplex formation: How mitochondrial structure can regulate bioenergetics
    Mitochondrion (IF 3.449) Pub Date : 2019-06-15
    Nicole Baker, Jeel Patel, Mireille Khacho

    The dynamic and fluid nature of mitochondria allows for modifications in mitochondrial shape, connectivity and cristae architecture. The precise balance of mitochondrial dynamics is among the most critical features in the control of mitochondrial function. In the past few years, mitochondrial shape has emerged as a key regulatory factor in the determination of the bioenergetic capacity of cells. This is mostly due to the recent discoveries linking changes in cristae organization with supercomplex assembly of the electron transport chain (ETC), also defined as the formation of respirosomes. Here we will review the most current advances demonstrating the impact of mitochondrial dynamics and cristae shape on oxidative metabolism, respiratory efficiency, and redox state. Furthermore, we will discuss the implications of mitochondrial dynamics and supercomplex assembly under physiological and pathological conditions.

  • Mitochondrial interaction with the endosomal compartment in endocytosis and mitochondrial transfer
    Mitochondrion (IF 3.449) Pub Date : 2019-05-14
    Kiran Todkar, Lilia Chikhi, Marc Germain

    Mitochondria are essential organelles required for cellular processes ranging from energy production to cellular differentiation. To perform these functions, mitochondria physically and functionally interact with other organelles such as the endoplasmic reticulum (ER) and endosomes. While the role of ER-mitochondria contact sites is well established, the interaction between mitochondria and endosomes has only recently been reported. These interactions are involved in lipid and ion transfer and potentially play a crucial role in mitochondria quality control and the release of mitochondrial components within extracellular vesicles. Here, we will discuss the current view of mitochondria-endosome interaction, both physically and functionally.

  • Proteolytic regulation of mitochondrial dynamics
    Mitochondrion (IF 3.449) Pub Date : 2019-04-25
    Jonathan V. Dietz, Iryna Bohovych, Martonio Ponte Viana, Oleh Khalimonchuk

    Spatiotemporal changes in the abundance, shape, and cellular localization of the mitochondrial network, also known as mitochondrial dynamics, are now widely recognized to play a key role in mitochondrial and cellular physiology as well as disease states. This process involves coordinated remodeling of the outer and inner mitochondrial membranes by conserved dynamin-like guanosine triphosphatases and their partner molecules in response to various physiological and stress stimuli. Although the core machineries that mediate fusion and partitioning of the mitochondrial network have been extensively characterized, many aspects of their function and regulation are incompletely understood and only beginning to emerge. In the present review we briefly summarize current knowledge about how the key mitochondrial dynamics-mediating factors are regulated via selective proteolysis by mitochondrial and cellular proteolytic machineries.

  • Mitochondrial complex IV is lost in neurons in the cuprizone mouse model
    Mitochondrion (IF 3.449) Pub Date : 2019-10-31
    Kristin N. Varhaug, Torbjørn Kråkenes, Maria N. Alme, Christian A. Vedeler, Laurence A. Bindoff
  • Mitochondrial Dynamics in Signaling and Disease.
    Mitochondrion (IF 3.449) Pub Date : 2019-07-30
    Timothy Shutt

  • Corrigendum to "Topical Coenzyme Q10 demonstrates mitochondrial-mediated neuroprotection in a rodent model of ocular hypertension" [Mitochondrion 36 (2017) 114-123].
    Mitochondrion (IF 3.449) Pub Date : 2019-05-21
    B M Davis,K Tian,M Pahlitzsch,J Brenton,N Ravindran,G Butt,G Malaguarnera,E M Normando,L Guo,M F Cordeiro

  • The process of RNA editing in plant mitochondria.
    Mitochondrion (IF 3.449) Pub Date : 2008-03-08
    Mizuki Takenaka,Daniil Verbitskiy,Johannes A van der Merwe,Anja Zehrmann,Axel Brennicke

    RNA editing changes more than 400 cytidines to uridines in the mRNAs of mitochondria in flowering plants. In other plants such as ferns and mosses, RNA editing reactions changing C to U and U to C are observed at almost equal frequencies. Development of transfection systems with isolated mitochondria and of in vitro systems with extracts from mitochondria has considerably improved our understanding of the recognition of specific editing sites in the last few years. These assays have also yielded information about the biochemical parameters, but the enzymes involved have not yet been identified. Here we summarize our present understanding of the process of RNA editing in flowering plant mitochondria.

  • Linear plasmids in plant mitochondria: peaceful coexistences or malicious invasions?
    Mitochondrion (IF 3.449) Pub Date : 2008-03-08
    Hirokazu Handa

    Plant mitochondria contain small extrachromosomal DNAs in addition to a large and complex main mitochondrial genome. These molecules can be regarded as extrachromosomal replicons or plasmids, of which there are two forms, circular and linear. Linear mitochondrial plasmids are present in many fungi and in some plants, but they seem to be absent from most animal cells. They usually have a common structural feature, called an invertron, that is characterized by the presence of terminal inverted repeats and proteins covalently attached to their 5 termini. Linear mitochondrial plasmids possess one to six ORFs that can encode unknown proteins but often code for the DNA and RNA polymerases. Although the functions of most linear plasmids in plant mitochondria are unknown, some plasmids may be associated with mitochondrial genome rearrangements and may have phenotypic effects due to their integration into mitochondrial genome. The Brassica 11.6-kb plasmid, one of the linear mitochondrial plasmids in plants, shows a non-maternal inheritance, in contrast to mitochondrial genomes. The origin of these plasmids is still a mystery, but indirect evidence indicates the possibility of horizontal transfer from fungal mitochondria. In this review, the main features of these unique DNAs present in plant mitochondria are described.

  • Differential action of iodine on mitochondria from human tumoral- and extra-tumoral tissue in inducing the release of apoptogenic proteins.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Geeta Upadhyay,Rajesh Singh,Ramesh Sharma,Anil K Balapure,Madan M Godbole

    Iodide is actively concentrated in the thyroid gland for thyroid hormone biosynthesis. Excess iodine has been observed to induce apoptosis in thyrocytes and mammary cells. The mechanism of iodine induced apoptosis is poorly understood. Among various cell organelles, mitochondria is known to provide conducive environment for the organification of iodine, i.e. iodination of different proteins. Mitochondria also play a central role in execution of apoptosis. To study the role of mitochondria in iodine induced apoptosis, we investigated the direct interaction of iodine and human breast mitochondria vis-a-vis its role in the initiation of apoptosis in vitro. We observed that mitochondria isolated from the tumor (TT) and extra-tumoral tissue (ET) of human breast display significant uptake of iodine. Mitochondrial proteins were observed to be predominantly iodinated in ET but not in TT mitochondria. Treatment with iodine showed an increase in mitochondrial permeability transition of TT and decrease in ET. Iodine induced released factor(s) other than cytochrome c from tumor mitochondria initiate(s) apoptosis in vitro, while those from ET mitochondria were non-apoptogenic in nature. To our knowledge, this is first report demonstrating that iodine acts differentially on mitochondria of tumor and extratumoral origin to release apoptogenic proteins from TT and has a protective effect on ET.

  • Complex II inactivation is lethal in the nematode Caenorhabditis elegans.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Harumi Ichimiya,R Giselle Huet,Phil Hartman,Hisako Amino,Kiyoshi Kita,Naoaki Ishii

    RNA-mediated interference (RNAi) was employed to systematically inactivate the four subunits of complex II in the mitochondrial electron transport chain. Embryonic lethality was the predominant result of inactivating three subunits (ceSDHB, ceSDHC, and ceSDHD) when using the soaking method to inactivate RNA. The feeding method was employed to deliver dsRNA from the fourth subunit (ceSDHA) to wild-type, mev-1 (mutated in ceSDHC of complex II), and gas-1 animals (mutated in a complex I gene). Survival was reduced only in the mev-1 genetic background, and in an oxygen-dependent fashion. Collectively, these data provide further evidence that compromised complex II integrity can result in sensitivity to oxidative stress.

  • Effect of the first window of ischemic preconditioning on mitochondrial dysfunction following global cerebral ischemia.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Miguel A Pérez-Pinzón,Abdul Basit,Kunjan R Dave,Raul Busto,Clemence Veauvy,Isabel Saul,Myron D Ginsberg,Thomas J Sick

    Rats may develop sustained tolerance against lethal cerebral ischemia after exposure to a sublethal ischemic insult (ischemic preconditioning (IPC)). Two windows for the induction of tolerance by IPC have been proposed, one that occurs within 1h following IPC, and the other one that occurs 1-3 days after IPC. An important difference between these two windows is that in contrast to the second window, neuroprotection against lethal ischemia is transient in the first window. We tested the hypothesis that rapid IPC would reduce or prevent ischemia-induced changes in mitochondrial function. IPC and ischemia were produced by bilateral carotid occlusions and systemic hypotension (50 mmHg) for 2 and 10 min, respectively. The non-synaptosomal mitochondria were harvested 30 min following the 10 min 'test' ischemia. Mitochondrial rate of respiration decreased by 10% when the substrates were pyruvate and malate, and 29% when the substrates were ascorbic acid and N,N,N',N'-tetramethyl-p-phenylenediamine ( P< 0.01). The activities of complex I-III decreased in ischemic group by 16, 23 (P < 0.05) and 24%, respectively. IPC was unable to prevent decreases in the rate of respiration and activities of different complexes. These data suggest that rapidly induced IPC is unable to protect the integrity of mitochondrial oxidative phosphorylation following cerebral ischemia, perhaps explaining why IPC only provides transitory protection in the 'first window'.

  • Mechanism of leflunomide-induced proliferation of mitochondria in mammalian cells.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Jan H Spodnik,Michal Wozniak,Dorota Budzko,Masa-Aki Teranishi,Mariusz Karbowski,Yuji Nishizawa,Jiro Usukura,Takashi Wakabayashi

    Leflunomide (LFM) is an inhibitor of mitochondrial enzyme dihydroorotate dehydrogenase (DHODH) that catalyzes the conversion of dihydroorotate to orotate coupled with the generation of reactive oxygen species (ROS) from mitochondria. We demonstrate here that LFM causes an unrestrained proliferation of mitochondria both in human osteosarcoma cell line 143B cells and rat liver derived RL-34 cells. Increases in the total mass of mitochondria per cell in LFM-treated cells were evidenced by the application of Green FM or 10-n-nonyl acridine orange to flow cytometry, an enhanced replication of mtDNA and electron microscopy. Externally added uridine improved the disturbance in cell cycle progression in LFM-treated cells, but failed to suppress such unrestrained mitochondrial proliferation. On the contrary, lapacol and 5-fluoroorotate, inhibitors of DHODH besides LFM, suppressed the biogenesis of mitochondria during the cell cycle progression. LFM, but not lapacol or 5-fluoroorotate, caused increases of the intracellular level of acetylated alpha-tubulin. These data suggest that the inhibition of DHODH may not be at least primarily related to the LFM-induced abnormal proliferation of mitochondria, and support our recent published observation that changes in the physicochemical properties of microtubules may be in someway concerned with the biogenesis of mitochondria.

  • Inheritance of mitochondrial disorders.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Patrick F Chinnery

    Over the last decade there have been major advances in our understanding of the genetic basis of mitochondrial disease, enabling genetic counseling for patients with autosomal dominant and autosomal recessive disorders. Genetic counseling for patients with mitochondrial DNA (mtDNA) mutations is less well established. Approximately one-third of adults with a mtDNA disorder are sporadic cases, usually due to a single deletion of mtDNA. About two-thirds of adults with mtDNA disease harbor a maternally transmitted point mutation. The recurrence risks are well documented for homoplasmic mtDNA mutations causing Leber hereditary optic neuropathy, but the situation is less clear for families with heteroplasmic mtDNA disorders. Two large studies have shown that for some heteroplasmic point mutations there appears to be a relationship between the percentage level of mutant mtDNA in a mother's blood and her risk of having clinically affected offspring. The situation is less clear for other point mutations, some of which may cause sporadic disease. Recent evidence has cast light on the general principles behind the transmission of heteroplasmic mtDNA point mutations, which may be important for genetic counseling in the future.

  • Mitochondrial neurogastrointestinal encephalomyopathy and thymidine metabolism: results and hypotheses.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Ramon Marti,Antonella Spinazzola,Ichizo Nishino,Antonio L Andreu,Ali Naini,Saba Tadesse,Juan A Oliver,Michio Hirano

    Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is an autosomal recessive disease with mitochondrial DNA (mtDNA) alterations and is caused by mutations in the nuclear gene encoding thymidine phosphorylase (TP). The cardinal clinical manifestations are ptosis, ophthalmoparesis, gastrointestinal dysmotility, cachexia, peripheral neuropathy, and leukoencephalopathy. Skeletal muscle shows mitochondrial abnormalities, including ragged-red fibers and cytochrome c oxidase deficiency, together with mtDNA depletion, multiple deletions or both. In MNGIE patients, TP mutations cause a loss-of-function of the cytosolic enzyme, TP. As a direct consequence of the TP defect, thymidine metabolism is altered. High blood levels of this nucleoside are likely to lead to mtDNA defects even in cells that do not express TP, such as skeletal muscle. We hypothesize that high concentrations of thymidine affect dNTP (deoxyribonucleoside triphosphate) metabolism in mitochondria more than in cytosol or nuclei, because mitochondrial dNTPs depend mainly on the thymidine salvage pathway, whereas nuclear dNTPs depend mostly on de novo pathway. The imbalance in the mitochondrial dNTP homeostasis affects mtDNA replication, leading to mitochondrial dysfunction.

  • Wobble modification defect suppresses translational activity of tRNAs with MERRF and MELAS mutations.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Takehiro Yasukawa,Tsutomu Suzuki,Shigeo Ohta,Kimitsuna Watanabe

    By purifying mutant mitochondrial tRNAs, we were able to ascertain that post-transcriptional modification at the anticodon wobble uridine is absent in tRNA(Lys) with the 8344 MERRF mutation and in tRNA(Leu(UUR)) with either the 3243 or 3271 MELAS mutation. Both the MERRF and MELAS mutant tRNAs substantially lost their translational ability, the extent of the loss in each mutant corresponding to the reduction in actual mitochondrial translational activity. Lack of the wobble modification deprived mutant tRNA(Lys) of interaction with the cognate codons. These features indicate that the modification defect plays a primary role in the molecular pathophysiology of these mitochondrial diseases.

  • New perspectives on the assembly process of mitochondrial respiratory chain complex cytochrome c oxidase.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Cristina Ugalde,Marieke J H Coenen,Murtada H Farhoud,Stefanie Gilinsky,Werner J H Koopman,Lambert P van den Heuvel,Jan A M Smeitink,Leo G J Nijtmans

    The assembly of cytochrome c oxidase (COX) is a complicated process and requires a number of assembly factors to put all the necessary subunits in the correct position. Defects in COX assembly lead in particular to serious neuromuscular disorders. We demonstrated that COX-deficient patients can be associated with different assembly patterns. To obtain more insight in the biogenesis of COX in a living cell, we used yeast as a model organism to design a way to pulse label holo-COX with green fluorescent protein (GFP). Using blue native electrophoresis, we showed that the GFP-tagged subunit is incorporated into fully assembled COX and this GFP tagged complex still has enzymatic activity. This allows us to correlate the GFP fluorescence signal detected in vivo by microscopy with the synthesis, turnover and assembly of COX.

  • The human complex I NDUFS4 subunit: from gene structure to function and pathology.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    S M S Budde,L P W J van den Heuvel,J A M Smeitink

    Complex I is the first and largest enzyme of the oxidative phosphorylation system. It consists of at least 43 subunits. Recent studies have shown that the NDUFS4 subunit of complex I contributes to the activation of the complex through cAMP dependent phosphorylation of a conserved site (RVS) located at the C-terminal region of this protein. This report focuses on the NDUFS4 subunit. Summarized is the current knowledge of this subunit, from gene structure to function and pathology.

  • Targeting of tRNA into yeast and human mitochondria: the role of anticodon nucleotides.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    O Kolesnikova,N Entelis,H Kazakova,I Brandina,R P Martin,I Tarassov

    In vivo, yeast mitochondria import a single cytoplasmic tRNA, tRNA(CUU)Lys, while human mitochondria do not import any cytoplasmic tRNA. We have previously demonstrated that both yeast and human isolated mitochondria can specifically internalize tRNA(CUU)Lys, several of its mutant versions and some mutant versions of yeast cytosolic tRNA(UUU)Lys (not imported in vivo). Aminoacylation of tRNA(CUU)Lys by the cytoplasmic lysyl-tRNA synthetase was a prerequisite for its import. Here we are studying the influence of one-base replacements in the anticodon of tRNAs(Lys) on their aminoacylation, on binding to the precursor of the mitochondrial lysyl-tRNA synthetase (carrier protein directing the import), and on the efficiency of import into isolated yeast and human mitochondria. We show that the base U35 is the main identity element for the yeast cytoplasmic lysyl-tRNA synthetase. The single replacement that abolished import was C34G, while all the others only modulated the import efficiency. The need of aminoacylation for import and for interaction with the carrier protein was shown only for a subset of mutant versions, while the others could be recognized and internalized without aminoacylation or in misacylated forms.

  • Frataxin deficiency and mitochondrial dysfunction.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Massimo Pandolfo

    Friedreich ataxia (FA) is an inherited recessive disorder characterized by progressive neurological disability and heart abnormalities. The Friedreich ataxia gene (FRDA) encodes a small mitochondrial protein, frataxin, which is produced in insufficient amounts in the disease as a consequence of a GAA triplet repeat expansion in the first intron of the gene. Frataxin deficiency leads to excessive free radical production, dysfunction of Fe-S center containing enzymes (in particular respiratory complexes I, II and III, and aconitase), and progressive iron accumulation in mitochondria. Frataxin may be a mitochondrial iron-binding protein that prevents this metal from participating in Fenton chemistry to generate toxic hydroxyl radicals. We investigated whether frataxin deficiency may in addition interfere with signaling pathways. First, we showed that exposure of FA fibroblasts to iron fails to produce the normally observed increase in expression of the stress defense protein manganese superoxide dismutase. This impaired induction involves a nuclear factor-kappaB-independent pathway that does not require free radical signaling intermediates. We also examined the role of frataxin in neuronal differentiation by using stably transfected clones of P19 embryonic carcinoma cells with antisense or sense frataxin constructs. We found that during retinoic acid-induced neurogenesis frataxin deficiency enhances apoptosis and reduces the number of terminally differentiated neuronal-like cells. The addition of the antioxidant N-acetyl-cysteine only rescues cells non-committed to the neuronal lineage, indicating that frataxin deficiency impairs differentiation mechanisms and survival responses through different mechanisms. Both studies suggest that some abnormalities in frataxin-deficient cells are related to free radical independent signaling pathways.

  • Biogenesis of iron-sulfur proteins in eukaryotes: components, mechanism and pathology.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Jana Gerber,Roland Lill

    Iron-sulfur (Fe-S) clusters are ubiquitous co-factors of proteins that play an important role in metabolism, electron-transfer and regulation of gene expression. In eukaryotes mitochondria are the primary site of Fe-S cluster biogenesis. The organelles contain some ten proteins of the so-called iron-sulfur cluster (ISC) assembly machinery that is well-conserved in bacteria and eukaryotes. The ISC assembly machinery is responsible for biogenesis of Fe-S proteins within mitochondria. In addition, this machinery is involved in the maturation of extra-mitochondrial Fe-S proteins by cooperating with mitochondrial proteins with an exclusive function in this process. This review summarizes recent developments in our understanding of the biogenesis of cellular Fe-S proteins in eukaryotes. Particular emphasis is given to disorders in Fe-S protein biogenesis causing human disease.

  • A novel defense system of mitochondria in mice and human subjects for preventing expression of mitochondrial dysfunction by pathogenic mutant mtDNAs.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Kazuto Nakada,Tomoko Ono,Jun-Ichi Hayashi

    Recently, we generated mtDNA-based disease mice (mito mice) by introduction of respiration-deficient mitochondria possessing pathogenic mutant mtDNA with a 4696 bp deletion (deltamtDNA4696) from somatic cells into mouse zygotes. Mito mice and cytochrome c oxidase (COX) electronmicrographs, that could identify the respiration enzyme activity at individual mitochondrial levels, enabled precise investigation of the pathogenesis of deltamtDNA4696. All the observations represented unambiguous evidence for the presence of extensive and continuous exchange of genetic contents between mitochondria. Thus, the inter-mitochondrial interaction could correspond to a very unique and effective defense system of the highly oxidative organelles for preventing mice and human subjects from expressing mitochondrial dysfunction caused by mtDNA lesions, which have been continuously created by oxidative stresses during aging. Here, we would like to propose a new hypothesis on mitochondrial biogenesis, 'the interaction theory of mammalian mitochondria': mitochondria exchange genetic contents, and thus lose individuality and function as a single dynamic cellular unit.

  • Expression of bacterial endonucleases in Saccharomyces cerevisiae mitochondria.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Sarah L Donahue,Uma Lakshmipathy,Colin Campbell

    Expression vectors were created in which the 5' end of the Saccharomyces cerevisiae CDC9 gene, which encodes a mitochondrial targeting peptide, was cloned in-frame with the coding regions of the EcoR I, Hind III, and Pst I endonuclease genes. Expression of the EcoR I and Hind III fusion proteins inhibited growth of yeast on glycerol-containing media and resulted in the nearly quantitative restriction digestion of their mitochondrial DNA. In contrast, expression of Pst I, which does not recognize any sites within yeast mitochondrial DNA, had no effect on growth in glycerol-containing media, and did not affect the integrity of the mitochondrial genome.

  • Inherited Mendelian defects of nuclear-mitochondrial communication affecting the stability of mitochondrial DNA.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Anna Limongelli,Valeria Tiranti

    The presence of mtDNA abnormalities inherited as Mendelian traits indicates the existence of mutations in nuclear genes affecting the integrity of the mitochondrial genome. Two groups of nucleus-driven abnormalities have been described: qualitative alterations of mtDNA, i.e. multiple large-scale deletions of mtDNA, and quantitative decrease of the mtDNA copy number, i.e. tissue-specific depletion of mtDNA. Autosomal dominant or recessive (adPEO), progressive ophthalmoplegia and autosomal-recessive mitochondrial neurogastrointestinal encephalomyopathy (MNGIE), are three neurodegenerative disorders associated with the coexistence of wild-type mtDNA with several deletion-containing mtDNA species. Heterozygous mutations of the genes encoding the muscle-heart isoform of the adenosine diphosphate/adenosine triphosphate mitochondrial translocator (ANT1), the main subunit of polymerase gamma (POLG1), and of the putative mtDNA helicase (Twinkle) have been found in adPEO families linked to three different loci, on chromosomes 4q34-35, 10q24, and 15q25, respectively. Mutations in the gene encoding thymidine phosphorylase have been identified in several MNGIE patients. Severe, tissue-specific depletion of mtDNA is the molecular hallmark of rapidly progressive hepatopathies or myopathies of infancy and childhood. Two genes, deoxyguanosine kinase and thymidine kinase type 2, both involved in the mitochondrion-specific salvage pathways of deoxynucleotide pools, have been associated with depletion syndromes in selected families.

  • Role of mitochondrial DNA in human aging.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Giuseppe Attardi

  • The thankless task of playing genetics with mammalian mitochondrial DNA: a 30-year review.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    M Pilar Bayona-Bafaluy,Patricio Fernández-Silva,José A Enríquez

    The advances obtained through the genetic tools available in yeast for studying the oxidative phosphorylation (OXPHOS) biogenesis and in particular the role of the mtDNA encoded genes, strongly contrast with the very limited benefits that similar approaches have generated for the study of mammalian mtDNA. Here we review the use of the genetic manipulation in mammalian mtDNA, its difficulty and the main types of mutants accumulated in the past 30 years and the information derived from them. We also point out the need for a substantial improvement in this field in order to obtain new tools for functional genetic studies and for the generation of animal models of mtDNA-linked diseases.

  • In vitro evidence of inhibition of mitochondrial protease processing by HIV-1 protease inhibitors in yeast: a possible contribution to lipodystrophy syndrome.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Abhijit Mukhopadhyay,Baoxian Wei,Steven J Zullo,Lauren V Wood,Henry Weiner

    Highly active antiretroviral therapy has been associated with the emergence of lipodystrophy syndromes that have clinical features commonly seen in patients with mitochondrial dysfunction. The effect of therapeutic protease inhibitors (PIs) on mitochondrial function is unknown. Mitochondrial matrix space proteins possess an amino-terminal leader peptide that is removed by the mitochondrial processing protease (MPP). Lack of cleavage could result in non- or dysfunctional mitochondrial proteins. The effects of different PIs on protease processing using pure MPP or yeast mitochondria, recognized models for mammalian counterparts, were examined in vitro. Multiple PIs were found to inhibit MPP, evidenced by accumulation of immature pALDH and decreased levels of processed ALDH. Both indinavir and amprenavir at 5.0 mg/ml resulted in significant inhibition of MPP. Although inhibition of MPP was also observed with ritonavir and saquinavir, the inhibition was difficult to quantify due to background inhibition of MPP by DMSO that was required to solubilize the drugs for the in vitro studies. Indinavir was also shown to inhibit MPP within yeast mitochondria. Lack of processing may impair mitochondrial function and contribute to the observed mitochondrial dysfunctions in patients receiving HAART and implicated in antiretroviral-associated lipodystrophy.

  • The complete mitochondrial DNA sequence of the cestode Echinococcus multilocularis (Cyclophyllidea: Taeniidae).
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Minoru Nakao,Noriko Yokoyama,Yasuhito Sako,Masahito Fukunaga,Akira Ito

    The 13,738 bp mitochondrial DNA from the cestode Echinococcus multilocularis has been sequenced. It contains two major noncoding regions and 36 genes (12 for proteins involved in oxidative phosphorylation, two for rRNAs and 22 for tRNAs) but a gene for ATPase subunit 8 is missing. All genes are transcribed in the same direction. Putative secondary structures of tRNAs indicate that most of them are conventional clover leaves but the dihydrouridine arm is unpaired in tRNA(Ser(AGN)), tRNA(Ser(UCN)), tRNA(Arg) and tRNA(Cys). The base composition at the wobble positions of fourfold degenerate codon families is highly biased toward U and against C.

  • 4-Hydroxytamoxifen is a potent inhibitor of the mitochondrial permeability transition.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Carla M P Cardoso,Leonor M Almeida,José B A Custódio

    The effects of 4-hydroxytamoxifen (OHTAM), the major active metabolite of the antiestrogen tamoxifen used in the breast cancer therapy, were studied on the mitochondrial permeability transition (MPT) and bioenergetic functions of mitochondria to evaluate the mechanisms underlying the cell death and toxic effects. The MPT was induced in vitro by incubating rat liver mitochondria with 1 mM inorganic phosphate plus Ca2+ and with tert-butyl hydroperoxide. OHTAM provides protection against the Ca2+-induced mitochondrial swelling, depolarization of the mitochondrial membrane potential (deltapsi), loss of electrophoretic Ca2+ uptake capacity and uncoupling of respiration, similarly to cyclosporine A. The concentrations of OHTAM used do not significantly affect deltapsi, respiratory control and adenosine diphosphate/oxygen ratios and induce repolarization and Ca2+ re-uptake, suggesting that such inhibitory effects of OHTAM were due to the prevention of the MPT induction and not due to the inhibition of the mitochondrial Ca2+ uniporter. Since the MPT induction has been linked to an oxidized shift in the mitochondrial redox state and/or increase in the generation of reactive oxygen species, the MPT prevention by OHTAM may be related to its high antioxidant capacity.

  • A Russian family of Slavic origin carrying mitochondrial DNA with a 9-bp deletion in region V and a long C-stretch in D-loop.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Vassilina A Sokolova,Vadim B Vasilyev,Thomas Delefosse,Catherine Hänni,Dominique Rochet,Catherine Godinot

    A 9-bp deletion first described in the mitochondrial DNA (mtDNA) for East Asian, Polynesian or Indian American populations of the B haplogroup is now discovered in Slavs. The Russian family carrying that deletion belongs to a new branch of the T haplogroup as deduced from D-loop sequence and haplogroup-specific restriction fragment length polymorphism analysis. One family member had a Kearns-Sayre syndrome with a 5.5 kb mtDNA deletion. This family also presented a long C-stretch in the D-loop. Whether or not the formation of the 5.5 kb deletion might be related to the 9-bp deletion or to the long C-stretch in the D-loop is discussed.

  • High-throughput assessment of mitochondrial membrane potential in situ using fluorescence resonance energy transfer.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    James A Dykens,Beth Fleck,Soumitra Ghosh,Michelle Lewis,Gonul Velicelebi,Manus W Ward

    Mitochondrial dysfunction causes dozens of debilitating diseases, and is implicated in the etiology of type 2 diabetes, Parkinson's, and Alzheimer's diseases, among others. However, development of mitochondrially targeted therapeutic agents has been impeded by the lack of high-throughput screening techniques that are capable of distinguishing in intact cells the mitochondrial membrane potential (deltapsi(m)) from the plasma membrane potential, (deltapsi(p)). We report here a fluorescence resonance energy transfer (FRET) assay that specifically monitors deltapsi(m) that is not confounded by background signal arising from potentiometric dye responding to deltapsi(p). The technique relies on energy transfer between nonyl acridine orange (NAO), which stains diphosphatidyl glycerol (cardiolipin) that is indigenous to the inner mitochondrial membrane, and tetramethylrhodamine methyl ester (TMR), a potentiometric dye that is sequestered by mitochondria as a Nernstian function of deltapsi(m) and concentration. FRET occurs only when both dyes co-localize to the mitochondria, and results in quenching of NAO emission by TMR in proportion to deltapsi(m). Validation studies using compounds with well-characterized mitochondrial effects, including oligomycin, CCCP+, bongkrekic acid, cyclosporin A, nigericin, ADP, and ruthenium red, demonstrate that the FRET-based deltapsi(m) assay responds in accord with the known pharmacology. Validation studies assessing the suitability of the technique for high-throughput compound screening indicate that the assay provides a sensitive and robust assessment not only of mitochondrial integrity in situ, but also, when used in conjunction with agents such as cyclosporin A, an indicator of permeability transition.

  • t-Loops in yeast mitochondria.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Lubomir Tomaska,Alexander M Makhov,Jack D Griffith,Jozef Nosek

    Mitochondria of several yeast species contain a linear DNA genome possessing specific terminal DNA structures dubbed mitochondrial telomeres. Several tandemly repeated units and a 5' single-stranded extension characterize mitochondrial telomeres in Candida parapsilosis, Pichia philodendra and Candida salmanticensis. Resemblance of this type of mitochondrial telomeres to typical nuclear telomeres suggests that they might form t-loop structures. Therefore we adopted a protocol for stabilization of potential t-loops in the mtDNA of C. parapsilosis and observed several loops at the ends of the mtDNA. A potential role of t-loops in protection of the ends of mtDNA and/or in mitochondrial telomere dynamics is discussed.

  • Flow cytometry in the study of mitochondrial respiratory chain disorders.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Karen Setterfield,Andrew J Williams,Jennifer Donald,David R Thorburn,Denise M Kirby,Ian Trounce,John Christodoulou

    We have developed a flow cytometric assay to measure the oxidative capacity of cultured lymphoblasts as a possible screening test for patients suspected of having a defect of the mitochondrial respiratory chain. Cells were incubated overnight in serum free media, followed by incubation with dihydroethidium with and without rotenone, and then analysed using flow cytometry to measure fluorescence. Inhibition with rotenone gave an increase in fluorescence compared to uninhibited cells. The change in fluorescence was significantly lower in four of the six patient cell lines, with a correlation between the activity of complex I and change in fluorescence. This method may be applicable to cell lines with defects in other complexes of the respiratory chain.

  • Catalase-dependent measurement of H2O2 in intact mitochondria.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Xia Li,James M May

    Mitochondria generate potentially damaging amounts of superoxide and H2O2 during oxidative metabolism. Although many assays are available to measure mitochondrial H2O2 generation, most detect H2O2 that has escaped the organelle. To measure H2O2 within mitochondria that contain catalase, we have developed an assay based on the ability of H2O2 to inhibit catalase in the presence of 3-amino-1,2,4-triazole. The assay is simple to perform, does not require expensive instrumentation, and is specific for H2O2. Results from this assay show that H2O2 generation in rat heart mitochondria reflects the activity of the electron transport chain. Further, liver mitochondria prepared from selenium-deficient rats have increased succinate-stimulated rates of H2O2 generation. This indicates that mitochondrial selenoenzymes are important for H2O2 removal. It also demonstrates the utility of this assay in measuring H2O2 release from mitochondria that do not contain catalase. The assay should be useful for study of both superoxide-dependent H2O2 generation in situ, and the role of endogenous mitochondrial catalase in H2O2 removal.

  • Cell cycle dependent morphology changes and associated mitochondrial DNA redistribution in mitochondria of human cell lines.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Daciana H Margineantu,W Gregory Cox,Linda Sundell,Steven W Sherwood,Joseph M Beechem,Roderick A Capaldi

    Mitochondria of osteosarcoma cells (143B) in culture have variable morphologies, classified according to the shape and size of the organelle as reticular, fragmented or intermediate. Synchronization and release from G0 has shown that the morphology of mitochondria oscillates between the reticular and fragmented state in a cell cycle dependent manner. Cells in G1 have reticular mitochondria while those in S phase have fragmented mitochondria. By using a novel method of fluorescence in situ hybridization, the morphology of mitochondria was correlated with mitochondrial DNA distribution. MtDNA molecules were seen in clusters of two to four along mitochondrial filaments. In the fully fragmented state, each mitochondrion contained at least one cluster. We discuss the importance of fission and fusion events in regulating the morphology of mitochondria, segregation of mtDNA and maintenance of the organelle's functional unity.

  • Role of intramitochondrial nitric oxide in rat heart and kidney during hypertension.
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Leopoldo Aguilera-Aguirre,Juan Carlos González-Hernández,Victoriano Pérez-Vázquez,Joel Ramírez,Mónica Clemente-Guerrero,Rafael Villalobos-Molina,Alfredo Saavedra-Molina

    Nitric oxide (NO) is an important reactive molecule in many organisms. A mitochondrial nitric oxide synthase has been described; however, the role of NO in this organelle is not yet fully clear. We tested the effect of intramitochondrial NO on various functions from spontaneously hypertensive rats (SHR) and their normotensive genetic control, Wistar-Kyoto (WKY) rats. While the stimulation of intramitochondrial NOS increased calcium- and phosphate-induced permeability transition pore opening, its inhibition partially prevented it, without affecting membrane potential. Matrix free calcium and the pH decreased with NOS inhibition. Basal [NO] was lower in SHR than in WKY. Our data suggest that intramitochondrial NO plays an important role in mitochondrial regulation during hypertension.

  • Mitochondrial dysfunction and antiretroviral nucleoside analog toxicities: what is the evidence?
    Mitochondrion (IF 3.449) Pub Date : 2005-08-27
    Tamir Dagan,Craig Sable,June Bray,Mariana Gerschenson

    Mitochondrial dysfunction has been associated with long-term toxicities of human immunodeficiency virus (HIV) therapy, particularly with the nucleoside analog reverse transcriptase inhibitors (NRTIs). Lactic acidosis, hepatic steatosis, myopathies, cardiomyopathies, neuropathies, and lipodystrophy are frequently attributed to mitochondrial toxicity. Since mitochondrial toxicity could pose a major threat to the long-term success of HIV therapy, the scientific evidence underlying an association between mitochondrial toxicity and antiretroviral therapies, must be carefully examined. There is some data to support the association between NRTIs and mitochondria dysfunction. In this review, we examine human, animal, and in vitro data implicating mitochondrial dysfunction as the causal mechanism of NRTI-associated toxicity in HIV-infected patients.

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上海纽约大学William Glover