An evolutionarily diverged mitochondrial protein controls biofilm growth and virulence in Candida albicans.

A forward genetic screening approach identified orf19.2500 as a gene controlling Candida albicans biofilm dispersal and biofilm detachment. Three-dimensional (3D) protein modeling and bioinformatics revealed that orf19.2500 is a conserved mitochondrial protein, structurally similar to, but functionally diverged from, the squalene/phytoene synthases family. The C. albicans orf19.2500 is distinguished by 3 evolutionarily acquired stretches of amino acid inserts, absent from all other eukaryotes except a small number of ascomycete fungi. Biochemical assays showed that orf19.2500 is required for the assembly and activity of the NADH ubiquinone oxidoreductase Complex I (CI) of the respiratory electron transport chain (ETC) and was thereby named NDU1. NDU1 is essential for respiration and growth on alternative carbon sources, important for immune evasion, required for virulence in a mouse model of hematogenously disseminated candidiasis, and for potentiating resistance to antifungal drugs. Our study is the first report on a protein that sets the Candida-like fungi phylogenetically apart from all other eukaryotes, based solely on evolutionary "gain" of new amino acid inserts that are also the functional hub of the protein.

Mitochondrial fusion and fission in cell life and death.

Mitochondria are dynamic organelles that constantly fuse and divide. These processes (collectively termed mitochondrial dynamics) are important for mitochondrial inheritance and for the maintenance of mitochondrial functions. The core components of the evolutionarily conserved fusion and fission machineries have now been identified, and mechanistic studies have revealed the first secrets of the complex processes that govern fusion and fission of a double membrane-bound organelle. Mitochondrial dynamics was recently recognized as an important constituent of cellular quality control. Defects have detrimental consequences on bioenergetic supply and contribute to the pathogenesis of neurodegenerative diseases. These findings open exciting new directions to explore mitochondrial biology.

Hidden cases of tRNA gene duplication and remolding in mitochondrial genomes of amphipods.

The evolution of tRNA genes in mitochondrial (mt) genomes is a complex process that includes duplications, degenerations, and transpositions, as well as a specific process of identity change through mutations in the anticodon (tRNA gene remolding or tRNA gene recruitment). Using amphipod-specific tRNA models for annotation, we show that tRNA duplications are more common in the mt genomes of amphipods than what was revealed by previous annotations. Seventeen cases of tRNA gene duplications were detected in the mt genomes of amphipods, and ten of them were tRNA genes that underwent remolding. The additional tRNA gene findings were verified using phylogenetic analysis and genetic distance analysis. The majority of remolded tRNA genes (seven out of ten cases) were found in the mt genomes of endemic amphipod species from Lake Baikal. All additional mt tRNA genes arose independently in the Baikalian amphipods, indicating the unusual plasticity of tRNA gene evolution in these species assemblages. The possible reasons for the unusual abundance of additional tRNA genes in the mt genomes of Baikalian amphipods are discussed. The amphipod-specific tRNA models developed for MiTFi refine existing predictions of tRNA genes in amphipods and reveal additional cases of duplicated tRNA genes overlooked by using less specific Metazoa-wide models. The application of these models for mt tRNA gene prediction will be useful for the correct annotation of mt genomes of amphipods and probably other crustaceans.

Macrophages Promote Oxidative Metabolism To Drive Nitric Oxide Generation in Response to Trypanosoma cruzi.

Trypanosoma cruzi is the causative agent of chronic chagasic cardiomyopathy. Why macrophages (mφs), the early responders to infection, fail to achieve parasite clearance is not known. Mouse (RAW 264.7) and human (THP-1 and primary) mφs were infected for 3 h and 18 h with T. cruzi TcI isolates, SylvioX10/4 (SYL, virulent) and TCC (nonpathogenic), which represent mφ stimulation and infection states, respectively. Mφs incubated with lipopolysaccharide and gamma interferon (LPS/IFN-γ) and with interleukin-4 (IL-4) were used as controls. We monitored the cytokine profile (using enzyme-linked immunosorbent assay [ELISA]), reactive oxygen species (ROS; fluorescent probes), nitric oxide (·NO; Griess assay), and metabolic state using a custom-designed mitoxosome array and Seahorse XF24 Analyzer. LPS/IFN-γ treatment of mφs elicited a potent increase in production of tumor necrosis alpha (TNF-α) at 3 h and of ROS and ·NO by 18 h. Upon SYL infection, murine mφs elicited an inflammatory cytokine profile (TNF-α ≫ TGF-ß + IL-10) and low levels of ·NO and ROS production. LPS/IFN-γ treatment resulted in the inhibition of oxidative metabolism at the gene expression and functional levels and a switch to the glycolytic pathway in mφs, while IL-4-treated mφs utilized oxidative metabolism to meet energy demands. SYL infection resulted in an intermediate functional metabolic state with increased mitoxosome gene expression and glycolysis, and IFN-γ addition shut down the oxidative metabolism in SYL-infected mφs. Further, TCC- and SYL-stimulated mφs exhibited similar levels of cell proliferation and production of TNF-α and ROS, while TCC-stimulated mφs exhibited up to 2-fold-higher levels of oxidative metabolism and ·NO production than SYL-infected mφs. Inhibiting ATP-coupled O2 consumption suppressed the ·NO generation in SYL-infected mφs. Mitochondrial oxygen consumption constitutes a mechanism for stimulating ·NO production in mφs during T. cruzi infection. Enhancing the oxidative metabolism provides an opportunity for increased ·NO production and pathogen clearance by mφs to limit disease progression.

Mitonuclear Ecology.

Eukaryotes were born of a chimeric union between two prokaryotes--the progenitors of the mitochondrial and nuclear genomes. Early in eukaryote evolution, most mitochondrial genes were lost or transferred to the nucleus, but a core set of genes that code exclusively for products associated with the electron transport system remained in the mitochondrion. The products of these mitochondrial genes work in intimate association with the products of nuclear genes to enable oxidative phosphorylation and core energy production. The need for coadaptation, the challenge of cotransmission, and the possibility of genomic conflict between mitochondrial and nuclear genes have profound consequences for the ecology and evolution of eukaryotic life. An emerging interdisciplinary field that I call "mitonuclear ecology" is reassessing core concepts in evolutionary ecology including sexual reproduction, two sexes, sexual selection, adaptation, and speciation in light of the interactions of mitochondrial and nuclear genomes.

NRIP1/RIP140 siRNA-mediated attenuation counteracts mitochondrial dysfunction in Down syndrome.

Mitochondrial dysfunction, which is consistently observed in Down syndrome (DS) cells and tissues, might contribute to the severity of the DS phenotype. Our recent studies on DS fetal hearts and fibroblasts have suggested that one of the possible causes of mitochondrial dysfunction is the downregulation of peroxisome proliferator-activated receptor gamma, coactivator 1 alpha (PGC-1α or PPARGC1A)--a key modulator of mitochondrial function--and of several nuclear-encoded mitochondrial genes (NEMGs). Re-analysis of publicly available expression data related to manipulation of chromosome 21 (Hsa21) genes suggested the nuclear receptor interacting protein 1 (NRIP1 or RIP140) as a good candidate Hsa21 gene for NEMG downregulation. Indeed, NRIP1 is known to affect oxidative metabolism and mitochondrial biogenesis by negatively controlling mitochondrial pathways regulated by PGC-1α. To establish whether NRIP1 overexpression in DS downregulates both PGC-1α and NEMGs, thereby causing mitochondrial dysfunction, we used siRNAs to decrease NRIP1 expression in trisomic human fetal fibroblasts. Levels of PGC-1α and NEMGs were increased and mitochondrial function was restored, as shown by reactive oxygen species decrease, adenosine 5'-triphosphate (ATP) production and mitochondrial activity increase. These findings indicate that the Hsa21 gene NRIP1 contributes to the mitochondrial dysfunction observed in DS. Furthermore, they suggest that the NRIP1-PGC-1α axe might represent a potential therapeutic target for restoring altered mitochondrial function in DS.

ABCC6 localizes to the mitochondria-associated membrane.

RATIONALE: Mutations of the orphan transporter ABCC6 (ATP-binding cassette, subfamily C, member 6) cause the connective tissue disorder pseudoxanthoma elasticum. ABCC6 was thought to be located on the plasma membrane of liver and kidney cells. OBJECTIVE: Mouse systems genetics and bioinformatics suggested that ABCC6 deficiency affects mitochondrial gene expression. We therefore tested whether ABCC6 associates with mitochondria. METHODS AND RESULTS: We found ABCC6 in crude mitochondrial fractions and subsequently pinpointed its localization to the purified mitochondria-associated membrane fraction. Cell-surface biotinylation in hepatocytes confirmed that ABCC6 is intracellular. Abcc6-knockout mice demonstrated mitochondrial abnormalities and decreased respiration reserve capacity. CONCLUSIONS: Our finding that ABCC6 localizes to the mitochondria-associated membrane has implications for its mechanism of action in normal and diseased states.

ERRgamma directs and maintains the transition to oxidative metabolism in the postnatal heart.

At birth, the heart undergoes a critical metabolic switch from a predominant dependence on carbohydrates during fetal life to a greater dependence on postnatal oxidative metabolism. This remains the principle metabolic state throughout life, although pathologic conditions such as heart failure and cardiac hypertrophy reactivate components of the fetal genetic program to increase carbohydrate utilization. Disruption of the ERRgamma gene (Esrrg), which is expressed at high levels in the fetal and postnatal mouse heart, blocks this switch, resulting in lactatemia, electrocardiographic abnormalities, and death during the first week of life. Genomic ChIP-on-chip and expression analysis identifies ERRgamma as both a direct and an indirect regulator of a nuclear-encoded mitochondrial genetic network that coordinates the postnatal metabolic transition. These findings reveal an unexpected and essential molecular genetic component of the oxidative metabolic gene program in the heart and highlight ERRgamma in the study of cardiac hypertrophy and failure.

Maternally inherited susceptibility to cancer.

Tumor microenvironment promotes mtDNA mutations. A number of these mutations will affect cell metabolism and increase cell survival. These mutations are positively selected and contribute to other tumor features, such as extracellular matrix remodeling and angiogenic processes, thus favoring metastases. Like somatic mutations, although with less marked effects, some mtDNA population polymorphisms will affect OXPHOS function, cell metabolism, and homeostasis. Thus, they could behave as inherited susceptibility factors for cancer. However, in addition to epidemiological evidence, other more direct clues are required. The cybrid approach can help to clarify this issue.

Genetic insights into OXPHOS defect and its role in cancer.

Warburg proposed that cancer originates from irreversible injury to mitochondrial oxidative phosphorylation (mtOXPHOS), which leads to an increase rate of aerobic glycolysis in most cancers. However, despite several decades of research related to Warburg effect, very little is known about the underlying genetic cause(s) of mtOXPHOS impairment in cancers. Proteins that participate in mtOXPHOS are encoded by both mitochondrial DNA (mtDNA) as well as nuclear DNA. This review describes mutations in mtDNA and reduced mtDNA copy number, which contribute to OXPHOS defects in cancer cells. Maternally inherited mtDNA renders susceptibility to cancer, and mutation in the nuclear encoded genes causes defects in mtOXPHOS system. Mitochondria damage checkpoint (mitocheckpoint) induces epigenomic changes in the nucleus, which can reverse injury to OXPHOS. However, irreversible injury to OXPHOS can lead to persistent mitochondrial dysfunction inducing genetic instability in the nuclear genome. Together, we propose that "mitocheckpoint" led epigenomic and genomic changes must play a key role in reversible and irreversible injury to OXPHOS described by Warburg. These epigenetic and genetic changes underlie the Warburg phenotype, which contributes to the development of cancer.

Mitochondrial transcription and its regulation in mammalian cells.

Human mitochondria contain multiple copies of a small double-stranded DNA genome that encode 13 components of the electron-transport chain and RNA components that are needed for mitochondrial translation. The mitochondrial genome is transcribed by a specialized machinery that includes a monomeric RNA polymerase, the mitochondrial transcription factor A and one of the two mitochondrial transcription factor B paralogues, TFB1M or TFB2M. Today, the components of the basal transcription machinery in mammalian mitochondria are known and their mechanisms of action are gradually being established. In addition, regulatory factors govern transcription levels both at the stage of initiation and termination, but the detailed biochemical understanding of these processes is largely missing.

Role of mitochondrial genetic interactions in determining adaptation to high altitude human population.

Physiological and haplogroup studies performed to understand high-altitude adaptation in humans are limited to individual genes and polymorphic sites. Due to stochastic evolutionary forces, the frequency of a polymorphism is affected by changes in the frequency of a near-by polymorphism on the same DNA sample making them connected in terms of evolution. Here, first, we provide a method to model these mitochondrial polymorphisms as "co-mutation networks" for three high-altitude populations, Tibetan, Ethiopian and Andean. Then, by transforming these co-mutation networks into weighted and undirected gene-gene interaction (GGI) networks, we were able to identify functionally enriched genetic interactions of CYB and CO3 genes in Tibetan and Andean populations, while NADH dehydrogenase genes in the Ethiopian population playing a significant role in high altitude adaptation. These co-mutation based genetic networks provide insights into the role of different set of genes in high-altitude adaptation in human sub-populations.

Molecular phylogeny of Japanese Catocala moths based on nucleotide sequences of the mitochondrial ND5 gene.

Phylogenetic relationships of 31 Japanese Catocala species were analyzed based on the partial nucleotide sequences of the mitochondrial NADH dehydrogenase subunit 5 (ND5) gene (762 bp). When several non-Catocala Noctuidae moths were designated as the outgroup, these Catocala species formed a monophyletic group. However, divergences between these Catocala species were very deep, and no close phylogenetic relationships were recognized among them except for that between the two recently separated species, C. xarippe and C. fulminea. The remote relationships implied for several pairs of species suggest that the color of the hindwings is a changeable characteristic, and does not reflect phylogenetic lineage. Continental specimens were analyzed in 20 of 31 Catocala species, and all of them showed a close relationship with their Japanese counterpart. However, the closeness of the nucleotide sequences between the Japanese and continental individuals of the same species varied from species to species, indicating that isolation between the Japanese and continental populations of these species occurred at many different times. The two analyzed species endemic to North America showed a close relationship with their morphologically inferred Japanese counterparts, indicating that the geographic separation and following speciation between these Eurasian and American species occurred much more recently compared with the speciation events among the Catocala species now found in Japan.

Mitochondrial deafness alleles confer misreading of the genetic code.

Despite the fact that important genetic diseases are caused by mutant mitochondrial ribosomes, the molecular mechanisms by which such ribosomes result in a clinical phenotype remain largely unknown. The absence of experimental models for mitochondrial diseases has also prevented the rational search for therapeutic interventions. Here, we report on the construction of bacterial hybrid ribosomes that contain various versions of the mitochondrial decoding region of ribosomal RNA. We show that the pathogenic mutations A1555G and C1494T decrease the accuracy of translation and render the ribosomal decoding site hypersusceptible to aminoglycoside antibiotics. This finding suggests misreading of the genetic code as an important molecular mechanism in disease pathogenesis.

New Insights Into Mitochondrial Dysfunction at Disease Susceptibility Loci in the Development of Type 2 Diabetes.

This study investigated the potential genetic mechanisms which underlie adipose tissue mitochondrial dysfunction in Type 2 diabetes (T2D), by systematically identifying nuclear-encoded mitochondrial genes (NEMGs) among the genes regulated by T2D-associated genetic loci. The target genes of these 'disease loci' were identified by mapping genetic loci associated with both disease and gene expression levels (expression quantitative trait loci, eQTL) using high resolution genetic maps, with independent estimates co-locating to within a small genetic distance. These co-locating signals were defined as T2D-eQTL and the target genes as T2D cis-genes. In total, 763 cis-genes were associated with T2D-eQTL, of which 50 were NEMGs. Independent gene expression datasets for T2D and insulin resistant cases and controls confirmed that the cis-genes and cis-NEMGs were enriched for differential expression in cases, providing independent validation that genetic maps can identify informative functional genes. Two additional results were consistent with a potential role of T2D-eQTL in regulating the 50 identified cis-NEMGs in the context of T2D risk: (1) the 50 cis-NEMGs showed greater differential expression compared to other NEMGs and (2) other NEMGs showed a trend towards significantly decreased expression if their expression levels correlated more highly with the subset of 50 cis-NEMGs. These 50 cis-NEMGs, which are differentially expressed and associated with mapped T2D disease loci, encode proteins acting within key mitochondrial pathways, including some of current therapeutic interest such as the metabolism of branched-chain amino acids, GABA and biotin.

Functional annotation of heart enriched mitochondrial genes GBAS and CHCHD10 through guilt by association.

Despite the mitochondria ubiquitous nature many of their components display divergences in their expression profile across different tissues. Using the bioinformatics-approach of guilt by association (GBA) we exploited these variations to predict the function of two so far poorly annotated genes: Coiled-coil-helix-coiled-coil-helix domain containing 10 (CHCHD10) and glioblastoma amplified sequence (GBAS). We predicted both genes to be involved in oxidative phosphorylation. Through in vitro experiments using gene-knockdown we could indeed confirm this and furthermore we asserted CHCHD10 to play a role in complex IV activity.

The mitochondrial gene orfH79 plays a critical role in impairing both male gametophyte development and root growth in CMS-Honglian rice.

BACKGROUND: Cytoplasmic male sterility (CMS) has often been associated with abnormal mitochondrial open reading frames. The mitochondrial gene orfH79 is a candidate gene for causing the CMS trait in CMS-Honglian (CMS-HL) rice. However, whether the orfH79 expression can actually induce CMS in rice remains unclear. RESULTS: Western blot analysis revealed that the ORFH79 protein is mainly present in mitochondria of CMS-HL rice and is absent in the fertile line. To investigate the function of ORFH79 protein in mitochondria, this gene was fused to a mitochondrial transit peptide sequence and used to transform wild type rice, where its expression induced the gametophytic male sterile phenotype. In addition, excessive accumulation of reactive oxygen species (ROS) in the microspore, a reduced ATP/ADP ratio, decreased mitochondrial membrane potential and a lower respiration rate in the transgenic plants were found to be similar to those in CMS-HL rice. Moreover, retarded growth of primary and lateral roots accompanied by abnormal accumulation of ROS in the root tip was observed in both transgenic rice and CMS-HL rice (YTA). CONCLUSION: These results suggest that the expression of orfH79 in mitochondria impairs mitochondrial function, which affects the development of both male gametophytes and the roots of CMS-HL rice.

Analysis of genetic diversity in cytoplasmic male sterility, and association of mitochondrial genes with petaloid-type cytoplasmic male sterility in tuber mustard (Brassica juncea var. tumida Tsen et Lee).

In our previous study, we bred a stable cytoplasmic male sterility (CMS) line of tuber mustard by using distant hybridization and subsequent backcrosses. In this CMS plants, all floral organs are normal except the anthers, which are transformed into petals or tubular structures. Recently, 2 mitochondrial genes-atpA and orf220-that are distinctively present in the CMS line of tuber mustard were cloned and partially characterized. In our study of genetic diversity analysis of CMS, 7 species of Brassica and Raphanus crops, which included 5 CMS lines and their respective maintainer lines, were used to compare the constitution of protein-coding genes in the mitochondrial genomes. In 4 of the 43 mitochondrial genes, namely, atpA, orf220, orf256, and orf305/orf324, polymorphisms were detected among the tuber mustard CMS line and its maintainer line. The results of a cluster analysis indicate that petaloid CMS phenotype of tuber mustard is a novel CMS type and is nearer to the nap CMS in Brassica napus at the phylogenetic level. The results of individual amplifications of these genes indicate the presence of 4 sequence-characterized amplified region (SCAR) markers, which enable rapid and reliable identification of this CMS. Expressions of the orf220 and orf256 genes were detected only in the CMS line, while expression of the orf305 gene was detected in the maintainer line. The different expression patterns of different mitochondrial-specific marker genes indicate that the quantity of mitochondrial proteins is differentially regulated during organ/tissue development in tuber mustard. The results of this study suggest that the above mentioned 4 mitochondrial genes are associated with the petaloid CMS phenotype in tuber mustard.

Evolutionary genomics analysis of human nucleus-encoded mitochondrial genes: implications for the roles of energy production and metabolic pathways in the pathogenesis and pathophysiology of demyelinating diseases.

The myelin sheath is a plasma membrane extension that lines nerve fibers to protect, support and insulate neurons. The myelination of axons in vertebrates enables fast, saltatory impulse propagation, and this process relies on organelles, especially on mitochondria to supply energy. Approximately 99% of mitochondrial proteins are encoded in the nucleus. Since mitochondria play a central role in the energy production and metabolic pathways, which are essential for myelinogenesis, studying these nucleus-encoded genes (nMGs) may provide new insights into the roles of energy metabolism in demyelinating diseases. In this work, a multiomics-based approach was employed to 1) construct a 1,740 human nMG subset with mitochondrial localization evidence obtained from the Integrated Mitochondrial Protein Index (IMPI) and MitoCarta databases, 2) conduct an evolutionary genomics analysis across mouse, rat, monkey, chimp, and human models, 3) examine dysmyelination phenotype-related genes (nMG subset genes with oligodendrocyte- ​and myelin-related ​phenotypes, OMP-nMGs) in MGI mouse lines and human patients, 4) determine the expression discrepancy of OMP-nMGs in brain tissues of cuprizone-treated mice, multiple sclerosis patients, and normal controls, and 5) conduct literature data mining to explore OMP-nMG-associated disease impacts. By contrasting OMP-nMGs with other genes, OMP-nMGs were found to be more ubiquitously expressed (59.1% vs. 16.1%), disease-associated (67.3% vs. 20.2%), and evolutionarily conserved within the human populations. Our multiomics-based analysis identified 110 OMP-nMGs implicated in energy production and lipid and glycan biosynthesis in the pathogenesis and pathophysiology of demyelinating disorders. Future targeted characterization of OMP-nMGs in abnormal myelination conditions may allow the discovery of novel nMG mediated mechanisms underlying myelinogenesis and related diseases.

Alterations in Mitochondrial Dynamic-related Genes in the Peripheral Blood of Alzheimer's Disease Patients.

BACKGROUND: Mitochondrial dysfunction is a pathological feature that manifests early in the brains of patients with Alzheimer's Disease (AD). The disruption of mitochondrial dynamics contributes to mitochondrial morphological and functional impairments. Our previous study demonstrated that the expression of genes involved in amyloid beta generation was altered in the peripheral blood of AD patients. OBJECTIVE: The aim of this study was to further investigate the relative levels of mitochondrial genes involved in mitochondrial dynamics, including mitochondrial fission and fusion, and mitophagy in peripheral blood samples from patients with AD compared to healthy controls. METHODS: The mRNA levels were analyzed by real-time polymerase chain reaction. Gene expression profiles were assessed in relation to cognitive performance. RESULTS: Significant changes were observed in the mRNA expression levels of fission-related genes; Fission1 (FIS1) levels in AD subjects were significantly higher than those in healthy controls, whereas Dynamin- related protein 1 (DRP1) expression was significantly lower in AD subjects. The levels of the mitophagy-related genes, PTEN-induced kinase 1 (PINK1) and microtubule-associated protein 1 light chain 3 (LC3), were significantly increased in AD subjects and elderly controls compared to healthy young controls. The mRNA levels of Parkin (PARK2) were significantly decreased in AD. Correlations were found between the expression levels of FIS1, DRP1 and PARK2 and cognitive performance scores. CONCLUSION: Alterations in mitochondrial dynamics in the blood may reflect impairments in mitochondrial functions in the central and peripheral tissues of AD patients. Mitochondrial fission, together with mitophagy gene profiles, might be potential considerations for the future development of blood-based biomarkers for AD.