Science.gov

Sample records for mitochondrial protease gene

  1. StAR enhances transcription of genes encoding the mitochondrial proteases involved in its own degradation.

    PubMed

    Bahat, Assaf; Perlberg, Shira; Melamed-Book, Naomi; Lauria, Ines; Langer, Thomas; Orly, Joseph

    2014-02-01

    Steroidogenic acute regulatory protein (StAR) is essential for steroid hormone synthesis in the adrenal cortex and the gonads. StAR activity facilitates the supply of cholesterol substrate into the inner mitochondrial membranes where conversion of the sterol to a steroid is catalyzed. Mitochondrial import terminates the cholesterol mobilization activity of StAR and leads to mounting accumulation of StAR in the mitochondrial matrix. Our studies suggest that to prevent mitochondrial impairment, StAR proteolysis is executed by at least 2 mitochondrial proteases, ie, the matrix LON protease and the inner membrane complexes of the metalloproteases AFG3L2 and AFG3L2:SPG7/paraplegin. Gonadotropin administration to prepubertal rats stimulated ovarian follicular development associated with increased expression of the mitochondrial protein quality control system. In addition, enrichment of LON and AFG3L2 is evident in StAR-expressing ovarian cells examined by confocal microscopy. Furthermore, reporter studies of the protease promoters examined in the heterologous cell model suggest that StAR expression stimulates up to a 3.5-fold increase in the protease gene transcription. Such effects are StAR-specific, are independent of StAR activity, and failed to occur upon expression of StAR mutants that do not enter the matrix. Taken together, the results of this study suggest the presence of a novel regulatory loop, whereby acute accumulation of an apparent nuisance protein in the matrix provokes a mitochondria to nucleus signaling that, in turn, activates selected transcription of genes encoding the enrichment of mitochondrial proteases relevant for enhanced clearance of StAR. PMID:24422629

  2. Biochemical and functional analysis of the YME1 gene product, an ATP and zinc-dependent mitochondrial protease from S. cerevisiae.

    PubMed Central

    Weber, E R; Hanekamp, T; Thorsness, P E

    1996-01-01

    Inactivation of YME1 in yeast causes several distinct phenotypes: an increased rate of DNA escape from mitochondria, temperature-sensitive growth on nonfermentable carbon sources, extremely slow growth when mitochondrial DNA is completely absent from the cell, and altered morphology of the mitochondrial compartment. The protein encoded by YME1, Yme1p, contains two highly conserved sequence elements, one implicated in the binding and hydrolysis of ATP, and the second characteristic of active site residues found in neutral, zinc-dependent proteases. Both the putative ATPase and zinc-dependent protease elements are necessary for the function of Yme1p as genes having mutations in critical residues of either of these motifs are unable to suppress any of the phenotypes exhibited by yme1 deletion strains. Yme1p co-fractionates with proteins associated with the mitochondrial inner membrane, is tightly associated with this membrane, and is oriented with the bulk of the protein facing the matrix. Unassembled subunit II of cytochrome oxidase is stabilized in yme1 yeast strains. The data support a model in which Yme1p is an ATP and zinc-dependent protease associated with the matrix side of the inner mitochondrial membrane. Subunit II of cytochrome oxidase, when not assembled into a higher order complex, is a likely substrate of Yme1p. Images PMID:8688560

  3. Mitochondrial Proteases as Emerging Pharmacological Targets.

    PubMed

    Gibellini, Lara; De Biasi, Sara; Nasi, Milena; Iannone, Anna; Cossarizza, Andrea; Pinti, Marcello

    2016-01-01

    The preservation of mitochondrial function and integrity is critical for cell viability. Under stress conditions, unfolded, misfolded or damaged proteins accumulate in a certain compartment of the organelle, interfering with oxidative phosphorylation and normal mitochondrial functions. In stress conditions, several mechanisms, including mitochondrial unfolded protease response (UPRmt), fusion and fission, and mitophagy are engaged to restore normal proteostasis of the organelle. Mitochondrial proteases are a family of more than 20 enzymes that not only are involved in the UPRmt, but actively participate at multiple levels in the stress-response system. Alterations in their expression levels, or mutations that determine loss or gain of function of these proteases deeply impair mitochondrial functionality and can be associated with the onset of inherited diseases, with the development of neurodegenerative disorders and with the process of carcinogenesis. In this review, we focus our attention on six of them, namely CLPP, HTRA2 and LONP1, by analysing the current knowledge about their functions, their involvement in the pathogenesis of human diseases, and the compounds currently available for inhibiting their functions. PMID:26831646

  4. Mitochondrial cereblon functions as a Lon-type protease

    PubMed Central

    Kataoka, Kosuke; Nakamura, China; Asahi, Toru; Sawamura, Naoya

    2016-01-01

    Lon protease plays a major role in the protein quality control system in mammalian cell mitochondria. It is present in the mitochondrial matrix, and degrades oxidized and misfolded proteins, thereby protecting the cell from various extracellular stresses, including oxidative stress. The intellectual disability-associated and thalidomide-binding protein cereblon (CRBN) contains a large, highly conserved Lon domain. However, whether CRBN has Lon protease-like function remains unknown. Here, we determined if CRBN has a protective function against oxidative stress, similar to Lon protease. We report that CRBN partially distributes in mitochondria, suggesting it has a mitochondrial function. To specify the mitochondrial role of CRBN, we mitochondrially expressed CRBN in human neuroblastoma SH-SY5Y cells. The resulting stable SH-SY5Y cell line showed no apparent effect on the mitochondrial functions of fusion, fission, and membrane potential. However, mitochondrially expressed CRBN exhibited protease activity, and was induced by oxidative stress. In addition, stably expressed cells exhibited suppressed neuronal cell death induced by hydrogen peroxide. These results suggest that CRBN functions specifically as a Lon-type protease in mitochondria. PMID:27417535

  5. SUMO-specific Protease 1 Regulates Mitochondrial Biogenesis through PGC-1α*

    PubMed Central

    Cai, Rong; Yu, Tingting; Huang, Chao; Xia, Xuefeng; Liu, Xiaobing; Gu, Jianmin; Xue, Song; Yeh, Edward T.H.; Cheng, Jinke

    2012-01-01

    Peroxisome proliferator-activated receptor γ (PPARγ) coactivator 1α (PGC-1α) is a master regulator of mitochondrial biogenesis in response to changes in the cellular environment, physiological or pathological status of mammals. PGC-1α is known to be modified by SUMO (Small Ubiquitin-like Modifier). However, it is not known whether SUMOylation could affect the function of PGC-1α in mitochondrial biogenesis and that how PGC-1α SUMOylation is regulated. In this study, we have identified the role of Sentrin/SUMO-specific protease 1 (SENP1) as a specific SUMO protease to regulate SUMOylation status of PGC-1α. More importantly, we have also found that SENP1 promotes PGC-1α transcription activity, which is essential for the expression of mitochondrial genes and subsequently mitochondrial biogenesis. Thus, we reveal that the SUMOylation of PGC-1α controlled by SENP1 plays an important role in mitochondrial biogenesis and function. PMID:23152500

  6. Mitochondrial RNA granules: Compartmentalizing mitochondrial gene expression.

    PubMed

    Jourdain, Alexis A; Boehm, Erik; Maundrell, Kinsey; Martinou, Jean-Claude

    2016-03-14

    In mitochondria, DNA replication, gene expression, and RNA degradation machineries coexist within a common nondelimited space, raising the question of how functional compartmentalization of gene expression is achieved. Here, we discuss the recently characterized "mitochondrial RNA granules," mitochondrial subdomains with an emerging role in the regulation of gene expression. PMID:26953349

  7. Lon protease: A key enzyme controlling mitochondrial bioenergetics in cancer

    PubMed Central

    Quirós, Pedro M; Bárcena, Clea; López-Otín, Carlos

    2014-01-01

    We have recently explored the in vivo functional and oncologic relevance of Lon protease (LONP1), an enzyme involved in mitochondrial quality control. We found that LONP1 is an essential protein for life and that it also performs a critical function in tumorigenesis by regulating the bioenergetics of cancer cells. PMID:27308364

  8. Mitochondrial AAA proteases--towards a molecular understanding of membrane-bound proteolytic machines.

    PubMed

    Gerdes, Florian; Tatsuta, Takashi; Langer, Thomas

    2012-01-01

    Mitochondrial AAA proteases play an important role in the maintenance of mitochondrial proteostasis. They regulate and promote biogenesis of mitochondrial proteins by acting as processing enzymes and ensuring the selective turnover of misfolded proteins. Impairment of AAA proteases causes pleiotropic defects in various organisms including neurodegeneration in humans. AAA proteases comprise ring-like hexameric complexes in the mitochondrial inner membrane and are functionally conserved from yeast to man, but variations are evident in the subunit composition of orthologous enzymes. Recent structural and biochemical studies revealed how AAA proteases degrade their substrates in an ATP dependent manner. Intersubunit coordination of the ATP hydrolysis leads to an ordered ATP hydrolysis within the AAA ring, which ensures efficient substrate dislocation from the membrane and translocation to the proteolytic chamber. In this review, we summarize recent findings on the molecular mechanisms underlying the versatile functions of mitochondrial AAA proteases and their relevance to those of the other AAA+ machines. PMID:22001671

  9. Biological Roles of the Podospora anserina Mitochondrial Lon Protease and the Importance of Its N-Domain

    PubMed Central

    Adam, Céline; Picard, Marguerite; Déquard-Chablat, Michelle; Sellem, Carole H.; Denmat, Sylvie Hermann-Le; Contamine, Véronique

    2012-01-01

    Mitochondria have their own ATP-dependent proteases that maintain the functional state of the organelle. All multicellular eukaryotes, including filamentous fungi, possess the same set of mitochondrial proteases, unlike in unicellular yeasts, where ClpXP, one of the two matricial proteases, is absent. Despite the presence of ClpXP in the filamentous fungus Podospora anserina, deletion of the gene encoding the other matricial protease, PaLon1, leads to lethality at high and low temperatures, indicating that PaLON1 plays a main role in protein quality control. Under normal physiological conditions, the PaLon1 deletion is viable but decreases life span. PaLon1 deletion also leads to defects in two steps during development, ascospore germination and sexual reproduction, which suggests that PaLON1 ensures important regulatory functions during fungal development. Mitochondrial Lon proteases are composed of a central ATPase domain flanked by a large non-catalytic N-domain and a C-terminal protease domain. We found that three mutations in the N-domain of PaLON1 affected fungal life cycle, PaLON1 protein expression and mitochondrial proteolytic activity, which reveals the functional importance of the N-domain of the mitochondrial Lon protease. All PaLon1 mutations affected the C-terminal part of the N-domain. Considering that the C-terminal part is predicted to have an α helical arrangement in which the number, length and position of the helices are conserved with the solved structure of its bacterial homologs, we propose that this all-helical structure participates in Lon substrate interaction. PMID:22693589

  10. Two mitochondrial matrix proteases act sequentially in the processing of mammalian matrix enzymes.

    PubMed

    Kalousek, F; Hendrick, J P; Rosenberg, L E

    1988-10-01

    The imported precursors of the mammalian matrix enzymes malate dehydrogenase [(S)-malate:NAD+ oxidoreductase, EC 1.1.1.37] and ornithine transcarbamylase (carbamoyl-phosphate:L-ornithine carbamoyltransferase, EC 2.1.3.3) are cleaved to their mature subunits in two steps, each catalyzed by matrix-localized processing proteases. The number and properties of these proteases are the subjects of this report. We have identified and characterized two distinct protease activities in a crude matrix fraction from rat liver: processing protease I, which cleaves these precursors to the corresponding intermediate form; and processing protease II, which cleaves the intermediate forms to mature subunits. Protease I is insensitive to chelation by EDTA and to inactivation with N-ethylmaleimide; protease II is inhibited by 5 mM EDTA and is inactivated by treatment with N-ethylmaleimide. We have prepared from mitochondrial matrix an 800-fold-enriched protease I fraction free of protease II activity by using the following steps: ion exchange, hydroxyapatite, molecular sieving, and hydrophobic chromatography. Using similar procedures, we also have prepared an approximately 2000-fold-enriched protease II fraction, which has a trace amount of contaminating protease I. This enriched protease II fraction has little or no cleavage activity toward mitochondrial precursors but rapidly and efficiently converts intermediate forms to mature size. Finally, we show that protease I alone is sufficient to cleave the precursor of a third nuclear-encoded mitochondrial protein subunit--the beta subunit of propionyl-CoA carboxylase [propanoyl-CoA:carbon dioxide ligase (ADP-forming), EC 6.4.1.3]--to its mature size. PMID:3050998

  11. CODAS Syndrome Is Associated with Mutations of LONP1, Encoding Mitochondrial AAA+ Lon Protease

    PubMed Central

    Strauss, Kevin A.; Jinks, Robert N.; Puffenberger, Erik G.; Venkatesh, Sundararajan; Singh, Kamalendra; Cheng, Iteen; Mikita, Natalie; Thilagavathi, Jayapalraja; Lee, Jae; Sarafianos, Stefan; Benkert, Abigail; Koehler, Alanna; Zhu, Anni; Trovillion, Victoria; McGlincy, Madeleine; Morlet, Thierry; Deardorff, Matthew; Innes, A. Micheil; Prasad, Chitra; Chudley, Albert E.; Lee, Irene Nga Wing; Suzuki, Carolyn K.

    2015-01-01

    CODAS syndrome is a multi-system developmental disorder characterized by cerebral, ocular, dental, auricular, and skeletal anomalies. Using whole-exome and Sanger sequencing, we identified four LONP1 mutations inherited as homozygous or compound-heterozygous combinations among ten individuals with CODAS syndrome. The individuals come from three different ancestral backgrounds (Amish-Swiss from United States, n = 8; Mennonite-German from Canada, n = 1; mixed European from Canada, n = 1). LONP1 encodes Lon protease, a homohexameric enzyme that mediates protein quality control, respiratory-complex assembly, gene expression, and stress responses in mitochondria. All four pathogenic amino acid substitutions cluster within the AAA+ domain at residues near the ATP-binding pocket. In biochemical assays, pathogenic Lon proteins show substrate-specific defects in ATP-dependent proteolysis. When expressed recombinantly in cells, all altered Lon proteins localize to mitochondria. The Old Order Amish Lon variant (LONP1 c.2161C>G[p.Arg721Gly]) homo-oligomerizes poorly in vitro. Lymphoblastoid cell lines generated from affected children have (1) swollen mitochondria with electron-dense inclusions and abnormal inner-membrane morphology; (2) aggregated MT-CO2, the mtDNA-encoded subunit II of cytochrome c oxidase; and (3) reduced spare respiratory capacity, leading to impaired mitochondrial proteostasis and function. CODAS syndrome is a distinct, autosomal-recessive, developmental disorder associated with dysfunction of the mitochondrial Lon protease. PMID:25574826

  12. The human LON protease binds to mitochondrial promoters in a single-stranded, site-specific, strand-specific manner.

    PubMed

    Fu, G K; Markovitz, D M

    1998-02-17

    LON proteases, which are ATP-dependent and exhibit ATPase activity, are found in bacteria, yeast, and humans. In Escherichia coli, LON is known to regulate gene expression by targeting specific regulatory proteins for degradation. The yeast and human LON proteins are encoded in the nucleus but localize to the mitochondrial matrix. In yeast, LON has been shown to be essential for the maintenance of the integrity of the mitochondrial genome. E. coli Lon has long been known to bind DNA, but we have only recently demonstrated that it binds preferentially to a specific TG-rich double-stranded sequence. We now show that human LON recognizes a very similar site in both the light and heavy chain promoters of the mitochondrial genome, in a region which is involved in regulating both DNA replication and transcription. Unlike E. coli Lon, however, human LON specifically binds to the TG-rich element only when it is presented in the context of a single DNA strand. These findings suggest that the human LON protease might regulate mitochondrial DNA replication and/or gene expression using site-specific, single-stranded DNA binding to target the degradation of regulatory proteins binding to adjacent sites in mitochondrial promoters. PMID:9485316

  13. Silencing of mitochondrial Lon protease deeply impairs mitochondrial proteome and function in colon cancer cells.

    PubMed

    Gibellini, Lara; Pinti, Marcello; Boraldi, Federica; Giorgio, Valentina; Bernardi, Paolo; Bartolomeo, Regina; Nasi, Milena; De Biasi, Sara; Missiroli, Sonia; Carnevale, Gianluca; Losi, Lorena; Tesei, Anna; Pinton, Paolo; Quaglino, Daniela; Cossarizza, Andrea

    2014-12-01

    Lon is a nuclear-encoded, mitochondrial protease that assists protein folding, degrades oxidized/damaged proteins, and participates in maintaining mtDNA levels. Here we show that Lon is up-regulated in several human cancers and that its silencing in RKO colon cancer cells causes profound alterations of mitochondrial proteome and function, and cell death. We silenced Lon in RKO cells by constitutive or inducible expression of Lon shRNA. Lon-silenced cells displayed altered levels of 39 mitochondrial proteins (26% related to stress response, 14.8% to ribosome assembly, 12.7% to oxidative phosphorylation, 8.5% to Krebs cycle, 6.3% to β-oxidation, and 14.7% to crista integrity, ketone body catabolism, and mtDNA maintenance), low levels of mtDNA transcripts, and reduced levels of oxidative phosphorylation complexes (with >90% reduction of complex I). Oxygen consumption rate decreased 7.5-fold in basal conditions, and ATP synthesis dropped from 0.25 ± 0.04 to 0.03 ± 0.001 nmol/mg proteins, in the presence of 2-deoxy-d-glucose. Hydrogen peroxide and mitochondrial superoxide anion levels increased by 3- and 1.3-fold, respectively. Mitochondria appeared fragmented, heterogeneous in size and shape, with dilated cristae, vacuoles, and electrondense inclusions. The triterpenoid 2-cyano-3,12-dioxooleana-1,9,-dien-28-oic acid, a Lon inhibitor, partially mimics Lon silencing. In summary, Lon is essential for maintaining mitochondrial shape and function, and for survival of RKO cells. PMID:25154874

  14. Emerging role of Lon protease as a master regulator of mitochondrial functions.

    PubMed

    Pinti, Marcello; Gibellini, Lara; Nasi, Milena; De Biasi, Sara; Bortolotti, Carlo Augusto; Iannone, Anna; Cossarizza, Andrea

    2016-08-01

    Lon protease is a nuclear-encoded, mitochondrial ATP-dependent protease highly conserved throughout the evolution, crucial for the maintenance of mitochondrial homeostasis. Lon acts as a chaperone of misfolded proteins, and is necessary for maintaining mitochondrial DNA. The impairment of these functions has a deep impact on mitochondrial functionality and morphology. An altered expression of Lon leads to a profound reprogramming of cell metabolism, with a switch from respiration to glycolysis, which is often observed in cancer cells. Mutations of Lon, which likely impair its chaperone properties, are at the basis of a genetic inherited disease named of the cerebral, ocular, dental, auricular, skeletal (CODAS) syndrome. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi. PMID:27033304

  15. Reversal of mitochondrial defects with CSB-dependent serine protease inhibitors in patient cells of the progeroid Cockayne syndrome

    PubMed Central

    Chatre, Laurent; Biard, Denis S. F.; Sarasin, Alain; Ricchetti, Miria

    2015-01-01

    UV-sensitive syndrome (UVSS) and Cockayne syndrome (CS) are human disorders caused by CSA or CSB gene mutations; both conditions cause defective transcription-coupled repair and photosensitivity. Patients with CS also display neurological and developmental abnormalities and dramatic premature aging, and their cells are hypersensitive to oxidative stress. We report CSA/CSB-dependent depletion of the mitochondrial DNA polymerase-γ catalytic subunit (POLG1), due to HTRA3 serine protease accumulation in CS, but not in UVsS or control fibroblasts. Inhibition of serine proteases restored physiological POLG1 levels in either CS fibroblasts and in CSB-silenced cells. Moreover, patient-derived CS cells displayed greater nitroso-redox imbalance than UVSS cells. Scavengers of reactive oxygen species and peroxynitrite normalized HTRA3 and POLG1 levels in CS cells, and notably, increased mitochondrial oxidative phosphorylation, which was altered in CS cells. These data reveal critical deregulation of proteases potentially linked to progeroid phenotypes in CS, and our results suggest rescue strategies as a therapeutic option. PMID:26038566

  16. Detergent alkaline proteases: enzymatic properties, genes, and crystal structures.

    PubMed

    Saeki, Katsuhisa; Ozaki, Katsuya; Kobayashi, Tohru; Ito, Susumu

    2007-06-01

    Subtilisin-like serine proteases from bacilli have been used in various industrial fields worldwide, particularly in the production of laundry and automatic dishwashing detergents. They belong to family A of the subtilase superfamily, which is composed of three clans, namely, true subtilisins, high-alkaline proteases, and intracellular proteases. We succeeded in the large-scale production of a high-alkaline protease (M-protease) from alkaliphilic Bacillus clausii KSM-K16, and the enzyme has been introduced into compact heavy-duty laundry detergents. We have also succeeded in the industrial-scale production of a new alkaline protease, KP-43, which was originally resistant to chemical oxidants and to surfactants, produced by alkaliphilic Bacillus sp. strain KSM-KP43 and have incorporated it into laundry detergents. KP-43 and related proteases form a new clan, oxidatively stable proteases, in subtilase family A. In this review, we describe the enzymatic properties, gene sequences, and crystal structures of M-protease, KP-43, and related enzymes. PMID:17630120

  17. Disorders of phospholipid metabolism: an emerging class of mitochondrial disease due to defects in nuclear genes

    PubMed Central

    Lu, Ya-Wen; Claypool, Steven M.

    2015-01-01

    The human nuclear and mitochondrial genomes co-exist within each cell. While the mitochondrial genome encodes for a limited number of proteins, transfer RNAs, and ribosomal RNAs, the vast majority of mitochondrial proteins are encoded in the nuclear genome. Of the multitude of mitochondrial disorders known to date, only a fifth are maternally inherited. The recent characterization of the mitochondrial proteome therefore serves as an important step toward delineating the nosology of a large spectrum of phenotypically heterogeneous diseases. Following the identification of the first nuclear gene defect to underlie a mitochondrial disorder, a plenitude of genetic variants that provoke mitochondrial pathophysiology have been molecularly elucidated and classified into six categories that impact: (1) oxidative phosphorylation (subunits and assembly factors); (2) mitochondrial DNA maintenance and expression; (3) mitochondrial protein import and assembly; (4) mitochondrial quality control (chaperones and proteases); (5) iron–sulfur cluster homeostasis; and (6) mitochondrial dynamics (fission and fusion). Here, we propose that an additional class of genetic variant be included in the classification schema to acknowledge the role of genetic defects in phospholipid biosynthesis, remodeling, and metabolism in mitochondrial pathophysiology. This seventh class includes a small but notable group of nuclear-encoded proteins whose dysfunction impacts normal mitochondrial phospholipid metabolism. The resulting human disorders present with a diverse array of pathologic consequences that reflect the variety of functions that phospholipids have in mitochondria and highlight the important role of proper membrane homeostasis in mitochondrial biology. PMID:25691889

  18. Disorders of phospholipid metabolism: an emerging class of mitochondrial disease due to defects in nuclear genes.

    PubMed

    Lu, Ya-Wen; Claypool, Steven M

    2015-01-01

    The human nuclear and mitochondrial genomes co-exist within each cell. While the mitochondrial genome encodes for a limited number of proteins, transfer RNAs, and ribosomal RNAs, the vast majority of mitochondrial proteins are encoded in the nuclear genome. Of the multitude of mitochondrial disorders known to date, only a fifth are maternally inherited. The recent characterization of the mitochondrial proteome therefore serves as an important step toward delineating the nosology of a large spectrum of phenotypically heterogeneous diseases. Following the identification of the first nuclear gene defect to underlie a mitochondrial disorder, a plenitude of genetic variants that provoke mitochondrial pathophysiology have been molecularly elucidated and classified into six categories that impact: (1) oxidative phosphorylation (subunits and assembly factors); (2) mitochondrial DNA maintenance and expression; (3) mitochondrial protein import and assembly; (4) mitochondrial quality control (chaperones and proteases); (5) iron-sulfur cluster homeostasis; and (6) mitochondrial dynamics (fission and fusion). Here, we propose that an additional class of genetic variant be included in the classification schema to acknowledge the role of genetic defects in phospholipid biosynthesis, remodeling, and metabolism in mitochondrial pathophysiology. This seventh class includes a small but notable group of nuclear-encoded proteins whose dysfunction impacts normal mitochondrial phospholipid metabolism. The resulting human disorders present with a diverse array of pathologic consequences that reflect the variety of functions that phospholipids have in mitochondria and highlight the important role of proper membrane homeostasis in mitochondrial biology. PMID:25691889

  19. The SUMO protease SENP5 is required to maintain mitochondrial morphology and function.

    PubMed

    Zunino, Rodolfo; Schauss, Astrid; Rippstein, Peter; Andrade-Navarro, Miguel; McBride, Heidi M

    2007-04-01

    Mitochondria are dynamic organelles that undergo regulated fission and fusion events that are essential to maintain metabolic stability. We previously demonstrated that the mitochondrial fission GTPase DRP1 is a substrate for SUMOylation. To further understand how SUMOylation impacts mitochondrial function, we searched for a SUMO protease that may affect mitochondrial dynamics. We demonstrate that the cytosolic pool of SENP5 catalyzes the cleavage of SUMO1 from a number of mitochondrial substrates. Overexpression of SENP5 rescues SUMO1-induced mitochondrial fragmentation that is partly due to the downregulation of DRP1. By contrast, silencing of SENP5 results in a fragmented and altered morphology. DRP1 was stably mono-SUMOylated in these cells, suggesting that SUMOylation leads to increased DRP1 mediated fission. In addition, the reduction of SENP5 levels resulted in a significant increase in the production of free radicals. Reformation of the mitochondrial tubules by expressing the dominant interfering DRP1 or by RNA silencing of endogenous DRP1 protein rescued both the morphological aberrations and the increased production of ROS induced by downregulation of SENP5. These data demonstrate the importance of SENP5 as a new regulator of SUMO1 proteolysis from mitochondrial targets, impacting mitochondrial morphology and metabolism. PMID:17341580

  20. Regulation of Skeletal Muscle Oxidative Capacity and Insulin Signaling by the Mitochondrial Rhomboid Protease PARL

    PubMed Central

    Civitarese, Anthony E.; MacLean, Paul S.; Carling, Stacy; Kerr-Bayles, Lyndal; McMillan, Ryan P.; Pierce, Anson; Becker, Thomas C.; Moro, Cedric; Finlayson, Jean; Lefort, Natalie; Newgard, Christopher B.; Mandarino, Lawrence; Cefalu, William; Walder, Ken; Collier, Greg R.; Hulver, Matthew W.; Smith, Steven R.; Ravussin, Eric

    2010-01-01

    SUMMARY Type 2 diabetes Mellitus (T2DM) and aging are characterized by insulin resistance, lower mitochondrial density and function and increased production of reactive oxygen species (ROS). In lower organisms continuous remodeling critically maintains the function and life cycle of mitochondria, in part by the protease pcp1 (PARL ortholog). We therefore examined whether variation in PARL protein content is associated with mitochondrial abnormalities and insulin resistance. Relative to healthy, young individuals (23±1y), PARL mRNA and mitochondrial mass were both reduced in elderly subjects (64.4±1.2 y; 51% and 44% respectively) and in subjects with T2DM (51.8±3 y; 31% and 41% respectively; all p<0.05). Muscle knock-down of PARL in mice resulted in lower mitochondrial content (−31±3%, p<0.05), lower OPA1 and PGC1α protein levels and impaired insulin signaling. Furthermore, mitochondrial cristae were malformed and resulted in elevated in vivo oxidative stress. Adenoviral suppression of PARL protein in healthy myotubes lowered mitochondrial mass (−33±8%), insulin stimulated glycogen synthesis (−33±9%) and increased ROS production (2-fold) (all p<0.05). We propose that lower PARL expression may contribute to the mitochondrial abnormalities seen in aging and T2DM. PMID:20444421

  1. Cloning and nucleotide sequence of the Vibrio cholerae hemagglutinin/protease (HA/protease) gene and construction of an HA/protease-negative strain.

    PubMed Central

    Häse, C C; Finkelstein, R A

    1991-01-01

    The structural gene hap for the extracellular hemagglutinin/protease (HA/protease) of Vibrio cholerae was cloned and sequenced. The cloned DNA fragment contained a 1,827-bp open reading frame potentially encoding a 609-amino-acid polypeptide. The deduced protein contains a putative signal sequence followed by a large propeptide. The extracellular HA/protease consists of 414 amino acids with a computed molecular weight of 46,700. In the absence of protease inhibitors, this is processed to the 32-kDa form which is usually isolated. The deduced amino acid sequence of the mature HA/protease showed 61.5% identity with the Pseudomonas aeruginosa elastase. The cloned hap gene was inactivated and introduced into the chromosome of V. cholerae by recombination to construct the HA/protease-negative strain HAP-1. The cloned fragment containing the hap gene was then shown to complement the mutant strain. Images PMID:2045361

  2. Mature DIABLO/Smac Is Produced by the IMP Protease Complex on the Mitochondrial Inner Membrane

    PubMed Central

    Burri, Lena; Strahm, Yvan; Hawkins, Christine J.; Gentle, Ian E.; Puryer, Michelle A.; Verhagen, Anne; Callus, Bernard; Vaux, David; Lithgow, Trevor

    2005-01-01

    DIABLO/Smac is a mitochondrial protein that can promote apoptosis by promoting the release and activation of caspases. To do so, DIABLO/Smac must first be processed by a mitochondrial protease and then released into the cytosol, and we show this in an intact cellular system. We propose that the precursor form of DIABLO/Smac enters the mitochondria through a stop-transfer pathway and is processed to its active form by the inner membrane peptidase (IMP) complex. Catalytic subunits of the mammalian IMP complex were identified based on sequence conservation and functional complementation, and the novel sequence motif RX5P in Imp1 and NX5S in Imp2 distinguish the two catalytic subunits. DIABLO/Smac is one of only a few specific proteins identified as substrates for the IMP complex in the mitochondrial intermembrane space. PMID:15814844

  3. Mitochondrial DNA Damage and its Consequences for Mitochondrial Gene Expression

    PubMed Central

    Cline, Susan D.

    2012-01-01

    How mitochondria process DNA damage and whether a change in the steady-state level of mitochondrial DNA damage (mtDNA) contributes to mitochondrial dysfunction are questions that fuel burgeoning areas of research into aging and disease pathogenesis. Over the past decade, researchers have identified and measured various forms of endogenous and environmental mtDNA damage and have elucidated mtDNA repair pathways. Interestingly, mitochondria do not appear to contain the full range of DNA repair mechanisms that operate in the nucleus, although mtDNA contains types of damage that are targets of each nuclear DNA repair pathway. The reduced repair capacity may, in part, explain the high mutation frequency of the mitochondrial chromosome. Since mtDNA replication is dependent on transcription, mtDNA damage may alter mitochondrial gene expression at three levels: by causing DNA polymerase γ nucleotide incorporation errors leading to mutations, by interfering with the priming of mtDNA replication by the mitochondrial RNA polymerase, or by inducing transcriptional mutagenesis or premature transcript termination. This review summarizes our current knowledge of mtDNA damage, its repair, and its effects on mtDNA integrity and gene expression. PMID:22728831

  4. Identification of potential mitochondrial CLPXP protease interactors and substrates suggests its central role in energy metabolism

    PubMed Central

    Fischer, Fabian; Langer, Julian D.; Osiewacz, Heinz D.

    2015-01-01

    Maintenance of mitochondria is achieved by several mechanisms, including the regulation of mitochondrial proteostasis. The matrix protease CLPXP, involved in protein quality control, has been implicated in ageing and disease. However, particularly due to the lack of knowledge of CLPXP’s substrate spectrum, only little is known about the pathways and mechanisms controlled by this protease. Here we report the first comprehensive identification of potential mitochondrial CLPXP in vivo interaction partners and substrates using a combination of tandem affinity purification and differential proteomics. This analysis reveals that CLPXP in the fungal ageing model Podospora anserina is mainly associated with metabolic pathways in mitochondria, e.g. components of the pyruvate dehydrogenase complex and the tricarboxylic acid cycle as well as subunits of electron transport chain complex I. These data suggest a possible function of mitochondrial CLPXP in the control and/or maintenance of energy metabolism. Since bioenergetic alterations are a common feature of neurodegenerative diseases, cancer, and ageing, our data comprise an important resource for specific studies addressing the role of CLPXP in these adverse processes. PMID:26679294

  5. Evolution of mitochondrial gene order in Annelida.

    PubMed

    Weigert, Anne; Golombek, Anja; Gerth, Michael; Schwarz, Francine; Struck, Torsten H; Bleidorn, Christoph

    2016-01-01

    Annelida is a highly diverse animal group with over 21,000 described species. As part of Lophotrochozoa, the vast majority of annelids are currently classified into two groups: Errantia and Sedentaria, together forming Pleistoannelida. Besides these taxa, Sipuncula, Amphinomidae, Chaetopteridae, Oweniidae and Magelonidae can be found branching at the base of the tree. Comparisons of mitochondrial genomes have been used to investigate phylogenetic relationship within animal taxa. Complete annelid mitochondrial genomes are available for some Sedentaria and Errantia and in most cases exhibit a highly conserved gene order. Only two complete genomes have been published from the basal branching lineages and these are restricted to Sipuncula. We describe the first complete mitochondrial genome sequences for all other basal branching annelid families: Owenia fusiformis (Oweniidae), Magelona mirabilis (Magelonidae), Eurythoe complanata (Amphinomidae), Chaetopterus variopedatus and Phyllochaetopterus sp. (Chaetopteridae). The mitochondrial gene order of all these taxa is substantially different from the pattern found in Pleistoannelida. Additionally, we report the first mitochondrial genomes in Annelida that encode genes on both strands. Our findings demonstrate that the supposedly highly conserved mitochondrial gene order suggested for Annelida is restricted to Pleistoannelida, representing the ground pattern of this group. All investigated basal branching annelid taxa show a completely different arrangement of genes than observed in Pleistoannelida. The gene order of protein coding and ribosomal genes in Magelona mirabilis differs only in two transposition events from a putative lophotrochozoan ground pattern and might be the closest to an ancestral annelid pattern. The mitochondrial genomes of Myzostomida show the conserved pattern of Pleistoannelida, thereby supporting their inclusion in this taxon. PMID:26299879

  6. Role of mitochondrial processing peptidase and AAA proteases in processing of the yeast acetohydroxyacid synthase precursor.

    PubMed

    Dasari, Suvarna; Kölling, Ralf

    2016-07-01

    We studied presequence processing of the mitochondrial-matrix targeted acetohydroxyacid synthase (Ilv2). C-terminal 3HA-tagging altered the cleavage pattern from a single step to sequential two-step cleavage, giving rise to two Ilv2-3HA forms (A and B). Both cleavage events were dependent on the mitochondrial processing peptidase (MPP). We present evidence for the involvement of three AAA ATPases, m- and i-AAA proteases, and Mcx1, in Ilv2-3HA processing. Both, precursor to A-form and A-form to B-form cleavage were strongly affected in a ∆yme1 mutant. These defects could be suppressed by overexpression of MPP, suggesting that MPP activity is limiting in the ∆yme1 mutant. Our data suggest that for some substrates AAA ATPases could play an active role in the translocation of matrix-targeted proteins. PMID:27398316

  7. The mitochondrial intramembrane protease PARL cleaves human Pink1 to regulate Pink1 trafficking.

    PubMed

    Meissner, Cathrin; Lorenz, Holger; Weihofen, Andreas; Selkoe, Dennis J; Lemberg, Marius K

    2011-06-01

    Intramembrane proteolysis is a conserved mechanism that regulates a variety of cellular processes ranging from transcription control to signaling. In mitochondria, the inner membrane rhomboid protease PARL has been implicated in the control of life span and apoptosis by a so far uncharacterized mechanism. Here, we show that PARL cleaves human Pink1, which is implicated in Parkinson's disease, within its conserved membrane anchor. Mature Pink1 is then free to be released into the cytosol or the mitochondrial intermembrane space. Upon depolarization of the mitochondrial membrane potential, the canonical import of Pink1 and PARL-catalyzed processing is blocked, leading to accumulation of the Pink1 precursor. As targeting of this precursor to the outer mitochondrial membrane has been shown to trigger mitophagy, we suggest that the PARL-catalyzed removal of the Pink1 signal sequence in the canonical import pathway acts as a cellular checkpoint for mitochondrial integrity. Furthermore, we show that two Parkinson's disease-causing mutations decrease the processing of Pink1 by PARL, with attendant implications for pathogenesis. PMID:21426348

  8. Analysis of the immunoglobulin A protease gene of Streptococcus sanguis.

    PubMed Central

    Gilbert, J V; Plaut, A G; Wright, A

    1991-01-01

    The amino acid sequence T-P-P-T-P-S-P-S is tandemly duplicated in the heavy chain of human immunoglobulin A1 (IgA1), the major antibody in secretions. The bacterial pathogen Streptococcus sanguis, a precursor to dental caries and a cause of bacterial endocarditis, yields IgA protease that cleaves only the Pro-Thr peptide bond in the left duplication, while the type 2 IgA proteases of the genital pathogen Neisseria gonorrhoeae and the respiratory pathogen Haemophilus influenzae cleave only the P-T bond in the right half. We have sequenced the entire S. sanguis iga gene cloned into Escherichia coli. A segment consisting of 20 amino acids tandemly repeated 10 times, of unknown function, occurs near the amino-terminal end of the enzyme encoded in E. coli. Identification of a predicted zinc-binding region in the S. sanguis enzyme and the demonstration that mutations in this region result in production of a catalytically inactive protein support the idea that the enzyme is a metalloprotease. The N. gonorrhoeae and H. influenzae enzymes were earlier shown to be serine-type proteases, while the Bacteroides melaninogenicus IgA protease was shown to be a cysteine-type enzyme. The streptococcal IgA protease amino acid sequence has no significant homology with either of the two previously determined IgA protease sequences, that of type 2 N. gonorrhoeae and type 1 H. influenzae. The differences in both structure and mechanism among these functionally analogous enzymes underscore their role in the infectious process and offer some prospect of therapeutic intervention. Images PMID:1987065

  9. New progress in snake mitochondrial gene rearrangement.

    PubMed

    Chen, Nian; Zhao, Shujin

    2009-08-01

    To further understand the evolution of snake mitochondrial genomes, the complete mitochondrial DNA (mtDNA) sequences were determined for representative species from two snake families: the Many-banded krait, the Banded krait, the Chinese cobra, the King cobra, the Hundred-pace viper, the Short-tailed mamushi, and the Chain viper. Thirteen protein-coding genes, 22-23 tRNA genes, 2 rRNA genes, and 2 control regions were identified in these mtDNAs. Duplication of the control region and translocation of the tRNAPro gene were two notable features of the snake mtDNAs. These results from the gene rearrangement comparisons confirm the correctness of traditional classification schemes and validate the utility of comparing complete mtDNA sequences for snake phylogeny reconstruction. PMID:19479623

  10. Mitochondrial Lon protease at the crossroads of oxidative stress, ageing and cancer.

    PubMed

    Pinti, Marcello; Gibellini, Lara; Liu, Yongzhang; Xu, Shan; Lu, Bin; Cossarizza, Andrea

    2015-12-01

    Lon protease is a nuclear DNA-encoded mitochondrial enzyme highly conserved throughout evolution, involved in the degradation of damaged and oxidized proteins of the mitochondrial matrix, in the correct folding of proteins imported in mitochondria, and in the maintenance of mitochondrial DNA. Lon expression is induced by various stimuli, including hypoxia and reactive oxygen species, and provides protection against cell stress. Lon down-regulation is associated with ageing and with cell senescence, while up-regulation is observed in tumour cells, and is correlated with a more aggressive phenotype of cancer. Lon up-regulation contributes to metabolic reprogramming observed in cancer, favours the switch from a respiratory to a glycolytic metabolism, helping cancer cell survival in the tumour microenvironment, and contributes to epithelial to mesenchymal transition. Silencing of Lon, or pharmacological inhibition of its activity, causes cell death in various cancer cells. Thus, Lon can be included in the growing class of proteins that are not responsible for oncogenic transformation, but that are essential for survival and proliferation of cancer cells, and that can be considered as a new target for development of anticancer drugs. PMID:26363553

  11. Higher plant mitochondrial DNA: Genomes, genes, mutants, transcription, translation

    SciTech Connect

    Not Available

    1986-01-01

    This volume contains brief summaries of 63 presentations given at the International Workshop on Higher Plant Mitochondrial DNA. The presentations are organized into topical discussions addressing plant genomes, mitochondrial genes, cytoplasmic male sterility, transcription, translation, plasmids and tissue culture. (DT)

  12. Proteolytic processing of Atg32 by the mitochondrial i-AAA protease Yme1 regulates mitophagy.

    PubMed

    Wang, Ke; Jin, Meiyan; Liu, Xu; Klionsky, Daniel J

    2013-11-01

    Mitophagy, the autophagic removal of mitochondria, occurs through a highly selective mechanism. In the yeast Saccharomyces cerevisiae, the mitochondrial outer membrane protein Atg32 confers selectivity for mitochondria sequestration as a cargo by the autophagic machinery through its interaction with Atg11, a scaffold protein for selective types of autophagy. The activity of mitophagy in vivo must be tightly regulated considering that mitochondria are essential organelles that produce most of the cellular energy, but also generate reactive oxygen species that can be harmful to cell physiology. We found that Atg32 was proteolytically processed at its C terminus upon mitophagy induction. Adding an epitope tag to the C terminus of Atg32 interfered with its processing and caused a mitophagy defect, suggesting the processing is required for efficient mitophagy. Furthermore, we determined that the mitochondrial i-AAA protease Yme1 mediated Atg32 processing and was required for mitophagy. Finally, we found that the interaction between Atg32 and Atg11 was significantly weakened in yme1∆ cells. We propose that the processing of Atg32 by Yme1 acts as an important regulatory mechanism of cellular mitophagy activity. PMID:24025448

  13. New insights into the evolution of subtilisin-like serine protease genes in Pezizomycotina

    PubMed Central

    2010-01-01

    Background Subtilisin-like serine proteases play an important role in pathogenic fungi during the penetration and colonization of their hosts. In this study, we perform an evolutionary analysis of the subtilisin-like serine protease genes of subphylum Pezizomycotina to find if there are similar pathogenic mechanisms among the pathogenic fungi with different life styles, which utilize subtilisin-like serine proteases as virulence factors. Within Pezizomycotina, nematode-trapping fungi are unique because they capture soil nematodes using specialized trapping devices. Increasing evidence suggests subtilisin-like serine proteases from nematode-trapping fungi are involved in the penetration and digestion of nematode cuticles. Here we also conduct positive selection analysis on the subtilisin-like serine protease genes from nematode-trapping fungi. Results Phylogenetic analysis of 189 subtilisin-like serine protease genes from Pezizomycotina suggests five strongly-supported monophyletic clades. The subtilisin-like serine protease genes previously identified or presumed as endocellular proteases were clustered into one clade and diverged the earliest in the phylogeny. In addition, the cuticle-degrading protease genes from entomopathogenic and nematode-parasitic fungi were clustered together, indicating that they might have overlapping pathogenic mechanisms against insects and nematodes. Our experimental bioassays supported this conclusion. Interestingly, although they both function as cuticle-degrading proteases, the subtilisin-like serine protease genes from nematode-trapping fungi and nematode-parasitic fungi were not grouped together in the phylogenetic tree. Our evolutionary analysis revealed evidence for positive selection on the subtilisin-like serine protease genes of the nematode-trapping fungi. Conclusions Our study provides new insights into the evolution of subtilisin-like serine protease genes in Pezizomycotina. Pezizomycotina subtilisins most likely evolved

  14. Distinct types of protease systems are involved in homeostasis regulation of mitochondrial morphology via balanced fusion and fission.

    PubMed

    Saita, Shotaro; Ishihara, Takaya; Maeda, Maki; Iemura, Shun-Ichiro; Natsume, Tohru; Mihara, Katsuyoshi; Ishihara, Naotada

    2016-05-01

    Mitochondrial morphology is dynamically regulated by fusion and fission. Several GTPase proteins control fusion and fission, and posttranslational modifications of these proteins are important for the regulation. However, it has not been clarified how the fusion and fission is balanced. Here, we report the molecular mechanism to regulate mitochondrial morphology in mammalian cells. Ablation of the mitochondrial fission, by repression of Drp1 or Mff, or by over-expression of MiD49 or MiD51, results in a reduction in the fusion GTPase mitofusins (Mfn1 and Mfn2) in outer membrane and long form of OPA1 (L-OPA1) in inner membrane. RNAi- or CRISPR-induced ablation of Drp1 in HeLa cells enhanced the degradation of Mfns via the ubiquitin-proteasome system (UPS). We further found that UPS-related protein BAT3/BAG6, here we identified as Mfn2-interacting protein, was implicated in the turnover of Mfns in the absence of mitochondrial fission. Ablation of the mitochondrial fission also enhanced the proteolytic cleavage of L-OPA1 to soluble S-OPA1, and the OPA1 processing was reversed by inhibition of the inner membrane protease OMA1 independent on the mitochondrial membrane potential. Our findings showed that the distinct degradation systems of the mitochondrial fusion proteins in different locations are enhanced in response to the mitochondrial morphology. PMID:26935475

  15. Protochlamydia Induces Apoptosis of Human HEp-2 Cells through Mitochondrial Dysfunction Mediated by Chlamydial Protease-Like Activity Factor

    PubMed Central

    Matsuo, Junji; Nakamura, Shinji; Ito, Atsushi; Yamazaki, Tomohiro; Ishida, Kasumi; Hayashi, Yasuhiro; Yoshida, Mitsutaka; Takahashi, Kaori; Sekizuka, Tsuyoshi; Takeuchi, Fumihiko; Kuroda, Makoto; Nagai, Hiroki; Hayashida, Kyoko; Sugimoto, Chihiro; Yamaguchi, Hiroyuki

    2013-01-01

    Obligate amoebal endosymbiotic bacterium Protochlamydia with ancestral pathogenic chlamydial features evolved to survive within protist hosts, such as Acanthamoba, 0.7–1.4 billion years ago, but not within vertebrates including humans. This observation raises the possibility that interactions between Protochlamydia and human cells may result in a novel cytopathic effect, leading to new insights into host-parasite relationships. Previously, we reported that Protochlamydia induces apoptosis of the immortalized human cell line, HEp-2. In this study, we attempted to elucidate the molecular mechanism underlying this apoptosis. We first confirmed that, upon stimulation with the bacteria, poly (ADP-ribose) polymerase (PARP) was cleaved at an early stage in HEp-2 cells, which was dependent on the amount of bacteria. A pan-caspase inhibitor and both caspase-3 and -9 inhibitors similarly inhibited the apoptosis of HEp-2 cells. A decrease of the mitochondrial membrane potential was also confirmed. Furthermore, lactacystin, an inhibitor of chlamydial protease-like activity factor (CPAF), blocked the apoptosis. Cytochalasin D also inhibited the apoptosis, which was dependent on the drug concentration, indicating that bacterial entry into cells was required to induce apoptosis. Interestingly, Yersinia type III inhibitors (ME0052, ME0053, and ME0054) did not have any effect on the apoptosis. We also confirmed that the Protochlamydia used in this study possessed a homologue of the cpaf gene and that two critical residues, histidine-101 and serine-499 of C. trachomatis CPAF in the active center, were conserved. Thus, our results indicate that after entry, Protochlamydia-secreted CPAF induces mitochondrial dysfunction with a decrease of the membrane potential, followed by caspase-9, caspase-3 and PARP cleavages for apoptosis. More interestingly, because C. trachomatis infection can block the apoptosis, our finding implies unique features of CPAF between pathogenic and primitive

  16. Inhibition of the Mitochondrial Protease ClpP as a Therapeutic Strategy for Human Acute Myeloid Leukemia.

    PubMed

    Cole, Alicia; Wang, Zezhou; Coyaud, Etienne; Voisin, Veronique; Gronda, Marcela; Jitkova, Yulia; Mattson, Rachel; Hurren, Rose; Babovic, Sonja; Maclean, Neil; Restall, Ian; Wang, Xiaoming; Jeyaraju, Danny V; Sukhai, Mahadeo A; Prabha, Swayam; Bashir, Shaheena; Ramakrishnan, Ashwin; Leung, Elisa; Qia, Yi Hua; Zhang, Nianxian; Combes, Kevin R; Ketela, Troy; Lin, Fengshu; Houry, Walid A; Aman, Ahmed; Al-Awar, Rima; Zheng, Wei; Wienholds, Erno; Xu, Chang Jiang; Dick, John; Wang, Jean C Y; Moffat, Jason; Minden, Mark D; Eaves, Connie J; Bader, Gary D; Hao, Zhenyue; Kornblau, Steven M; Raught, Brian; Schimmer, Aaron D

    2015-06-01

    From an shRNA screen, we identified ClpP as a member of the mitochondrial proteome whose knockdown reduced the viability of K562 leukemic cells. Expression of this mitochondrial protease that has structural similarity to the cytoplasmic proteosome is increased in leukemic cells from approximately half of all patients with AML. Genetic or chemical inhibition of ClpP killed cells from both human AML cell lines and primary samples in which the cells showed elevated ClpP expression but did not affect their normal counterparts. Importantly, Clpp knockout mice were viable with normal hematopoiesis. Mechanistically, we found that ClpP interacts with mitochondrial respiratory chain proteins and metabolic enzymes, and knockdown of ClpP in leukemic cells inhibited oxidative phosphorylation and mitochondrial metabolism. PMID:26058080

  17. Inhibition of the mitochondrial protease, ClpP, as a therapeutic strategy for human acute myeloid leuekmia

    PubMed Central

    Cole, Alicia; Wang, Zezhou; Coyaud, Etienne; Voisin, Veronique; Gronda, Marcela; Jitkova, Yulia; Mattson, Rachel; Hurren, Rose; Babovic, Sonja; Maclean, Neil; Restall, Ian; Wang, Xiaoming; Jeyaraju, Danny V.; Sukhai, Mahadeo A.; Prabha, Swayam; Bashir, Shaheena; Ramakrishnan, Ashwin; Leung, Elisa; Qia, Yi Hua; Zhang, Nianxian; Combes, Kevin R.; Ketela, Troy; Lin, Fengshu; Houry, Walid A.; Aman, Ahmed; Al-awar, Rima; Zheng, Wei; Wienholds, Erno; Xu, Chang Jiang; Dick, John; Wang, Jean C.Y.; Moffat, Jason; Minden, Mark D.; Eaves, Connie J.; Bader, Gary D.; Hao, Zhenyue; Kornblau, Steven M.; Raught, Brian; Schimmer, Aaron D.

    2015-01-01

    Summary From an shRNA screen, we identified ClpP as a member of the mitochondrial proteome whose knockdown reduced the viability of K562 leukemic cells. Expression of this mitochondrial protease that has structural similarity to the cytoplasmic proteosome is increased in the leukemic cells from approximately half of patients with AML. Genetic or chemical inhibition of ClpP killed cells from both human AML cell lines and primary samples in which the cells showed elevated ClpP expression, but did not affect their normal counterparts. Importantly, Clpp knockout mice were viable with normal hematopoiesis. Mechanistically, we found ClpP interacts with mitochondrial respiratory chain proteins and metabolic enzymes, and knockdown of ClpP in leukemic cells inhibited oxidative phosphorylation and mitochondrial metabolism. PMID:26058080

  18. Echinochrome A Increases Mitochondrial Mass and Function by Modulating Mitochondrial Biogenesis Regulatory Genes

    PubMed Central

    Jeong, Seung Hun; Kim, Hyoung Kyu; Song, In-Sung; Noh, Su Jin; Marquez, Jubert; Ko, Kyung Soo; Rhee, Byoung Doo; Kim, Nari; Mishchenko, Natalia P.; Fedoreyev, Sergey A.; Stonik, Valentin A.; Han, Jin

    2014-01-01

    Echinochrome A (Ech A) is a natural pigment from sea urchins that has been reported to have antioxidant properties and a cardio protective effect against ischemia reperfusion injury. In this study, we ascertained whether Ech A enhances the mitochondrial biogenesis and oxidative phosphorylation in rat cardio myoblast H9c2 cells. To study the effects of Ech A on mitochondrial biogenesis, we measured mitochondrial mass, level of oxidative phosphorylation, and mitochondrial biogenesis regulatory gene expression. Ech A treatment did not induce cytotoxicity. However, Ech A treatment enhanced oxygen consumption rate and mitochondrial ATP level. Likewise, Ech A treatment increased mitochondrial contents in H9c2 cells. Furthermore, Ech A treatment up-regulated biogenesis of regulatory transcription genes, including proliferator-activated receptor gamma co-activator (PGC)-1α, estrogen-related receptor (ERR)-α, peroxisome proliferator-activator receptor (PPAR)-γ, and nuclear respiratory factor (NRF)-1 and such mitochondrial transcription regulatory genes as mitochondrial transcriptional factor A (TFAM), mitochondrial transcription factor B2 (TFB2M), mitochondrial DNA direct polymerase (POLMRT), single strand binding protein (SSBP) and Tu translation elongation factor (TUFM). In conclusion, these data suggest that Ech A is a potentiated marine drug which enhances mitochondrial biogenesis. PMID:25196935

  19. The mitochondrial protease AtFTSH4 safeguards Arabidopsis shoot apical meristem function.

    PubMed

    Dolzblasz, Alicja; Smakowska, Elwira; Gola, Edyta M; Sokołowska, Katarzyna; Kicia, Marta; Janska, Hanna

    2016-01-01

    The shoot apical meristem (SAM) ensures continuous plant growth and organogenesis. In LD 30 °C, plants lacking AtFTSH4, an ATP-dependent mitochondrial protease that counteracts accumulation of internal oxidative stress, exhibit a puzzling phenotype of premature SAM termination. We aimed to elucidate the underlying cellular and molecular processes that link AtFTSH4 with SAM arrest. We studied AtFTSH4 expression, internal oxidative stress accumulation, and SAM morphology. Directly in the SAM we analysed H2O2 accumulation, mitochondria behaviour, and identity of stem cells using WUS/CLV3 expression. AtFTSH4 was expressed in proliferating tissues, particularly during the reproductive phase. In the mutant, SAM, in which internal oxidative stress accumulates predominantly at 30 °C, lost its meristematic fate. This process was progressive and stage-specific. Premature meristem termination was associated with an expansion in SAM area, where mitochondria lost their functionality. All these effects destabilised the identity of the stem cells. SAM termination in ftsh4 mutants is caused both by internal oxidative stress accumulation with time/age and by the tissue-specific role of AtFTSH4 around the flowering transition. Maintaining mitochondria functionality within the SAM, dependent on AtFTSH4, is vital to preserving stem cell activity throughout development. PMID:27321362

  20. The mitochondrial protease AtFTSH4 safeguards Arabidopsis shoot apical meristem function

    PubMed Central

    Dolzblasz, Alicja; Smakowska, Elwira; Gola, Edyta M.; Sokołowska, Katarzyna; Kicia, Marta; Janska, Hanna

    2016-01-01

    The shoot apical meristem (SAM) ensures continuous plant growth and organogenesis. In LD 30 °C, plants lacking AtFTSH4, an ATP-dependent mitochondrial protease that counteracts accumulation of internal oxidative stress, exhibit a puzzling phenotype of premature SAM termination. We aimed to elucidate the underlying cellular and molecular processes that link AtFTSH4 with SAM arrest. We studied AtFTSH4 expression, internal oxidative stress accumulation, and SAM morphology. Directly in the SAM we analysed H2O2 accumulation, mitochondria behaviour, and identity of stem cells using WUS/CLV3 expression. AtFTSH4 was expressed in proliferating tissues, particularly during the reproductive phase. In the mutant, SAM, in which internal oxidative stress accumulates predominantly at 30 °C, lost its meristematic fate. This process was progressive and stage-specific. Premature meristem termination was associated with an expansion in SAM area, where mitochondria lost their functionality. All these effects destabilised the identity of the stem cells. SAM termination in ftsh4 mutants is caused both by internal oxidative stress accumulation with time/age and by the tissue-specific role of AtFTSH4 around the flowering transition. Maintaining mitochondria functionality within the SAM, dependent on AtFTSH4, is vital to preserving stem cell activity throughout development. PMID:27321362

  1. Mitochondrial complex 1 gene analysis in keratoconus

    PubMed Central

    Pathak, Dhananjay; Nayak, Bhagabat; Singh, Manvendra; Sharma, Namrata; Tandon, Radhika; Sinha, Rajesh; Titiyal, Jeewan S.

    2011-01-01

    Purpose Keratoconus is characterized by the thinning of corneal stroma, resulting in reduced vision. The exact etiology of keratoconus (KC) is still unknown. The involvement of oxidative stress (OS) in this disease has been reported. However, the exact mechanism of OS in keratoconus is still unknown. Thus we planned this study to screen mitochondrial complex I genes for sequence changes in keratoconus patients and controls, as mitochondrial complex I is the chief source of reactive oxygen species (ROS) production. Methods A total of 20 keratoconus cases and 20 healthy controls without any ocular disorder were enrolled in this study. Mitochondrial complex I genes (ND1, 2, 3, 4, 4L, 5, and 6) were amplified in all patients and controls using 12 pairs of primers by PCR. After sequencing, DNA sequences were analyzed against the mitochondrial reference sequence NC_012920. Haplogroup frequency based Principle Component Analysis (PCA) was constructed to determine whether the gene pool of keratoconus patients is closer to major populations in India. Results DNA sequencing revealed a total 84 nucleotide variations in patients and 29 in controls. Of 84 nucleotide changes, 18 variations were non-synonymous and two novel frame-shift mutations were detected in cases. Non-synonymous mtDNA sequence variations may account for increased ROS and decreased ATP production. This ultimately leads to OS; which is a known cause for variety of corneal abnormalities. Haplotype analysis showed that most of the patients were clustered under the haplogroups: T, C4a2a, R2’TJ, M21’Q1a, M12’G2a2a, M8’CZ and M7a2a, which are present as negligible frequency in normal Indian population, whereas only few patients were found to be a part of the other haplogroups like U7 (Indo-European), R2 and R31, whose origin is contentious. Conclusions Mt complex I sequence variations are the main cause of elevated ROS production which leads oxidative stress. This oxidative stress then starts a cascade of

  2. Mutations in nuclear genes alter post-transcriptional regulation of mitochondrial genes.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nuclear gene products are required for the expression of mitochondrial genes and elaboration of functional mitochondrial protein complexes. To better understand the roles of these nuclear genes, we exploited the mitochondrial encoded S-type of cytoplasmic male sterility (CMS-S) and developed a nove...

  3. The Kunitz-protease inhibitor domain in amyloid precursor protein reduces cellular mitochondrial enzymes expression and function.

    PubMed

    Chua, Li-Min; Lim, Mei-Li; Wong, Boon-Seng

    2013-08-01

    Mitochondrial dysfunction is a prominent feature of Alzheimer's disease (AD) and this can be contributed by aberrant metabolic enzyme function. But, the mechanism causing this enzymatic impairment is unclear. Amyloid precursor protein (APP) is known to be alternatively spliced to produce three major isoforms in the brain (APP695, APP751, APP770). Both APP770 and APP751 contain the Kunitz Protease Inhibitory (KPI) domain, but the former also contain an extra OX-2 domain. APP695 on the other hand, lacks both domains. In AD, up-regulation of the KPI-containing APP isoforms has been reported. But the functional contribution of this elevation is unclear. In the present study, we have expressed and compared the effect of the non-KPI containing APP695 and the KPI-containing APP751 on mitochondrial function. We found that the KPI-containing APP751 significantly decreased the expression of three major mitochondrial metabolic enzymes; citrate synthase, succinate dehydrogenase and cytochrome c oxidase (COX IV). This reduction lowers the NAD(+)/NADH ratio, COX IV activity and mitochondrial membrane potential. Overall, this study demonstrated that up-regulation of the KPI-containing APP isoforms is likely to contribute to the impairment of metabolic enzymes and mitochondrial function in AD. PMID:23872114

  4. Loss of the m-AAA protease subunit AFG₃L₂ causes mitochondrial transport defects and tau hyperphosphorylation.

    PubMed

    Kondadi, Arun Kumar; Wang, Shuaiyu; Montagner, Sara; Kladt, Nikolay; Korwitz, Anne; Martinelli, Paola; Herholz, David; Baker, Michael J; Schauss, Astrid C; Langer, Thomas; Rugarli, Elena I

    2014-05-01

    The m-AAA protease subunit AFG₃L₂ is involved in degradation and processing of substrates in the inner mitochondrial membrane. Mutations in AFG₃L₂ are associated with spinocerebellar ataxia SCA28 in humans and impair axonal development and neuronal survival in mice. The loss of AFG₃L₂ causes fragmentation of the mitochondrial network. However, the pathogenic mechanism of neurodegeneration in the absence of AFG₃L₂ is still unclear. Here, we show that depletion of AFG₃L₂ leads to a specific defect of anterograde transport of mitochondria in murine cortical neurons. We observe similar transport deficiencies upon loss of AFG₃L₂ in OMA1-deficient neurons, indicating that they are not caused by OMA1-mediated degradation of the dynamin-like GTPase OPA1 and inhibition of mitochondrial fusion. Treatment of neurons with antioxidants, such as N-acetylcysteine or vitamin E, or decreasing tau levels in axons restored mitochondrial transport in AFG₃L₂-depleted neurons. Consistently, tau hyperphosphorylation and activation of ERK kinases are detected in mouse neurons postnatally deleted for Afg3l2. We propose that reactive oxygen species signaling leads to cytoskeletal modifications that impair mitochondrial transport in neurons lacking AFG₃L₂. PMID:24681487

  5. Validation of Mitochondrial Gene Delivery in Liver and Skeletal Muscle via Hydrodynamic Injection Using an Artificial Mitochondrial Reporter DNA Vector.

    PubMed

    Yasuzaki, Yukari; Yamada, Yuma; Ishikawa, Takuya; Harashima, Hideyoshi

    2015-12-01

    For successful mitochondrial transgene expression, two independent processes, i.e., developing a mitochondrial gene delivery system and construction of DNA vector to achieve mitochondrial gene expression, are required. To date, very few studies dealing with mitochondrial gene delivery have been reported and, in most cases, transgene expression was not validated, because the construction of a reporter DNA vector for mitochondrial gene expression is the bottleneck. In this study, mitochondrial transgene expression by the in vivo mitochondrial gene delivery of an artificial mitochondrial reporter DNA vector via hydrodynamic injection is demonstrated. In the procedure, a large volume of naked plasmid DNA (pDNA) is rapidly injected. We designed and constructed pHSP-mtLuc (CGG) as a mitochondrial reporter DNA vector that possesses a mitochondrial heavy strand promoter (HSP) and an artificial mitochondrial genome with the reporter NanoLuc (Nluc) luciferase gene that records adjustments to the mitochondrial codon system. We delivered the pDNA into mouse liver mitochondria by hydrodynamic injection, and detected exogenous mRNA in the liver using reverse transcription PCR analysis. The hydrodynamic injection of pHSP-mtLuc (CGG) resulted in the expression of the Nluc luciferase protein in liver and skeletal muscle. Our mitochondrial transgene expression reporter system would contribute to mitochondrial gene therapy and further studies directed at mitochondrial molecular biology. PMID:26567847

  6. Oligodendroglial differentiation induces mitochondrial genes and inhibition of mitochondrial function represses oligodendroglial differentiation

    PubMed Central

    Schoenfeld, Robert; Wong, Alice; Silva, Jillian; Li, Ming; Itoh, Aki; Horiuchi, Makoto; Itoh, Takayuki; Pleasure, David; Cortopassi, Gino

    2011-01-01

    Demyelination occurs in multiple inherited mitochondrial diseases. We studied which genes were induced as a consequence of differentiation in rodent and human oligodendroglia. Cholesterol, myelin and mitochondrial genes were significantly increased with oligodendroglial differentiation. Mitochondrial DNA content per cell and acetyl CoA-related transcripts increased significantly; thus, the large buildup of cholesterol necessary for myelination appears to require mitochondrial production of acetyl-CoA. Oligodendroglia were treated with low doses of the mitochondrial inhibitor rotenone to test the dependence of differentiation on mitochondrial function. Undifferentiated cells were resistant to rotenone, whereas differentiating cells were much more sensitive. Very low doses of rotenone that did not affect viability or ATP synthesis still inhibited differentiation, as measured by reduced levels of the myelin transcripts 2′,3′-Cyclic Nucleotide-3′-Phosphodiesterase and Myelin Basic Protein. Thus, mitochondrial transcripts and mtDNA are amplified during oligodendroglial differentiation, and differentiating oligodendroglia are especially sensitive to mitochondrial inhibition, suggesting mechanisms for demyelination observed in mitochondrial disease. PMID:20005986

  7. Codon usage trend in mitochondrial CYB gene.

    PubMed

    Uddin, Arif; Chakraborty, Supriyo

    2016-07-15

    Here we reported the pattern of codon usage and the factors which influenced the codon usage pattern in mitochondrial cytochrome B (MT-CYB) gene among pisces, aves and mammals. The F1 axis of correspondence analysis showed highly significant positive correlation with nucleobases A3, C and C3 and significant negative correlation with T and T3 while F2 of correspondence analysis showed significant positive correlation with C and C3 and significant negative correlation with A and A3. From the neutrality plot, it was evident that the GC12 was influenced by mutation pressure and natural selection with a ratio of 0.10/0.90=0.11 in pisces, 0.024/0.976=0.0245 in aves and in mammals 0.215/0.785=0.273, which indicated that the role of natural selection was more than mutation pressure on structuring the bases at the first and second codon positions. Natural selection played the major role; but compositional constraint and mutation pressure also played a significant role in codon usage pattern. Analysis of codon usage pattern has contributed to the better understanding of the mechanism of distribution of codons and the evolution of MT-CYB gene. PMID:27063508

  8. Chymotrypsin protease inhibitor gene family in rice: Genomic organization and evidence for the presence of a bidirectional promoter shared between two chymotrypsin protease inhibitor genes.

    PubMed

    Singh, Amanjot; Sahi, Chandan; Grover, Anil

    2009-01-01

    Protease inhibitors play important roles in stress and developmental responses of plants. Rice genome contains 17 putative members in chymotrypsin protease inhibitor (ranging in size from 7.21 to 11.9 kDa) gene family with different predicted localization sites. Full-length cDNA encoding for a putative subtilisin-chymotrypsin protease inhibitor (OCPI2) was obtained from Pusa basmati 1 (indica) rice seedlings. 620 bp-long OCPI2 cDNA contained 219 bp-long ORF, coding for 72 amino acid-long 7.7 kDa subtilisin-chymotrypsin protease inhibitor (CPI) cytoplasmic protein. Expression analysis by semi-quantitative RT-PCR analysis showed that OCPI2 transcript is induced by varied stresses including salt, ABA, low temperature and mechanical injury in both root and shoot tissues of the seedlings. Transgenic rice plants produced with OCPI2 promoter-gus reporter gene showed that this promoter directs high salt- and ABA-regulated expression of the GUS gene. Another CPI gene (OCPI1) upstream to OCPI2 (with 1126 bp distance between the transcription initiation sites of the two genes; transcription in the reverse orientation) was noted in genome sequence of rice genome. A vector that had GFP and GUS reporter genes in opposite orientations driven by 1881 bp intergenic sequence between the OCPI2 and OCPI1 (encompassing the region between the translation initiation sites of the two genes) was constructed and shot in onion epidermal cells by particle bombardment. Expression of both GFP and GUS from the same epidermal cell showed that this sequence represents a bidirectional promoter. Examples illustrating gene pairs showing co-expression of two divergent neighboring genes sharing a bidirectional promoter have recently been extensively worked out in yeast and human systems. We provide an example of a gene pair constituted of two homologous genes showing co-expression governed by a bidirectional promoter in rice. PMID:18952157

  9. The dsbB gene product is required for protease production by Burkholderia cepacia.

    PubMed Central

    Abe, M; Nakazawa, T

    1996-01-01

    Burkholderia cepacia KF1, isolated from a pneumonia patient, produces a 37-kDa extracellular metalloprotease. A protease-deficient and lipase-proficient mutant, KFT1007, was complemented by a clone having an open reading frame coding for a 170-amino-acid polypeptide which showed significant homology to Escherichia coli DsbB. KFT1007, a presumed dsbB mutant, also failed to show motility, and both protease secretion and motility were restored by the introduction of the cloned dsbB gene of B. cepacia. The mutant KFT1007 excreted a 43-kDa polypeptide that is immunologically related to the 37-kDa mature protease. These results suggested that the dsbB mutant secretes a premature and catalytically inactive form of protease and that disulfide formation is required for the production of extracellular protease by B. cepacia. PMID:8926116

  10. Gene therapy for the treatment of mitochondrial DNA disorders.

    PubMed

    Taylor, Robert W

    2005-02-01

    Despite recent epidemiological studies confirming that mitochondrial respiratory chain disorders due to mutations in either the mitochondrial or nuclear genome are amongst the most common inherited human diseases, realistic therapeutic strategies for these patients remain limited. The disappointing response to various vitamins, cofactors and electron acceptors that have been administered to patients in an attempt to bypass the underlying respiratory chain defect, coupled with the complexities of human mitochondrial genetics, means that novel and innovative means are required to offer realistic treatments. Several 'gene therapy' strategies have therefore been proposed to treat patients with pathogenic mitochondrial DNA mutations, and although these are not without their own inherent problems, several exciting approaches promise much in the near future. This review will provide a basic background to mitochondrial genetics and mitochondrial DNA disorders before introducing the various strategies being tested in vitro at present, in cell culture and animal models, and, in the example of therapeutic exercise, in patients themselves. PMID:15757380

  11. Leishmania aethiopica: identification and characterization of cathepsin L-like cysteine protease genes.

    PubMed

    Kuru, Teklu; Jirata, Dagim; Genetu, Abebe; Barr, Stephen; Mengistu, Yohannes; Aseffa, Abraham; Gedamu, Lashitew

    2007-03-01

    There is limited information on the biology and pathogenesis of Leishmania aethiopica, causative agent of cutaneous leishmaniasis (CL) in Ethiopia. In this study we have identified and characterized two cathepsin L-like cysteine protease genes, Laecpa and Laecpb, from L. aethiopica. The predicted amino acid sequence of Laecpa and Laecpb is more than 75% identical with homologous cathepsin L-like cysteine protease genes of other Leishmania species and less than 50% identical with human cathepsin L. Laecpa is expressed predominantly in the stationary, and to a lower level, during the amastigote stage while Laecpb is specifically expressed in the stationary stage of L. aethiopica development. Phylogenetic analysis showed that the two genes are grouped into separate clades which are the result of gene duplication. The isolation of these genes will be useful in developing Leishmania species specific diagnostics for molecular epidemiological studies and serves as a first step to study the role of cysteine proteases in L. aethiopica pathogenesis. PMID:17083936

  12. MOLECULAR IDENTIFICATION OF CYSTEINE AND TRYPSIN PROTEASE, EFFECT OF DIFFERENT HOSTS ON PROTEASE EXPRESSION, AND RNAI MEDIATED SILENCING OF CYSTEINE PROTEASE GENE IN THE SUNN PEST.

    PubMed

    Amiri, Azam; Bandani, Ali Reza; Alizadeh, Houshang

    2016-04-01

    Sunn pest, Eurygaster integriceps, is a serious pest of cereals in the wide area of the globe from Near and Middle East to East and South Europe and North Africa. This study described for the first time, identification of E. integriceps trypsin serine protease and cathepsin-L cysteine, transcripts involved in digestion, which might serve as targets for pest control management. A total of 478 and 500 base pair long putative trypsin and cysteine gene sequences were characterized and named Tryp and Cys, respectively. In addition, the tissue-specific relative gene expression levels of these genes as well as gluten hydrolase (Gl) were determined under different host kernels feeding conditions. Result showed that mRNA expression of Cys, Tryp, and Gl was significantly affected after feeding on various host plant species. Transcript levels of these genes were most abundant in the wheat-fed E. integriceps larvae compared to other hosts. The Cys transcript was detected exclusively in the gut, whereas the Gl and Tryp transcripts were detectable in both salivary glands and gut. Also possibility of Sunn pest gene silencing was studied by topical application of cysteine double-stranded RNA (dsRNA). The results indicated that topically applied dsRNA on fifth nymphal stage can penetrate the cuticle of the insect and induce RNA interference. The Cys gene mRNA transcript in the gut was reduced to 83.8% 2 days posttreatment. Also, it was found that dsRNA of Cys gene affected fifth nymphal stage development suggesting the involvement of this protease in the insect growth, development, and molting. PMID:26609789

  13. Purification and characterization of cloned alkaline protease gene of Geobacillus stearothermophilus.

    PubMed

    Iqbal, Irfana; Aftab, Muhammad Nauman; Afzal, Mohammed; Ur-Rehman, Asad; Aftab, Saima; Zafar, Asma; Ud-Din, Zia; Khuharo, Ateeque Rahman; Iqbal, Jawad; Ul-Haq, Ikram

    2015-02-01

    Thermostable alkaline serine protease gene of Geobacillus stearothermophilus B-1172 was cloned and expressed in Escherichia coli BL21 (DE3) using pET-22b(+), as an expression vector. The growth conditions were optimized for maximal production of the protease using variable fermentation parameters, i.e., pH, temperature, and addition of an inducer. Protease, thus produced, was purified by ammonium sulfate precipitation followed by ion exchange chromatography with 13.7-fold purification, with specific activity of 97.5 U mg(-1) , and a recovery of 23.6%. Molecular weight of the purified protease, 39 kDa, was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme was stable at 90 °C at pH 9. The enzyme activity was steady in the presence of EDTA indicating that the protease was not a metalloprotease. No significant change in the activity of protease after addition of various metal ions further strengthened this fact. However, an addition of 1% Triton X-100 or SDS surfactants constrained the enzyme specific activity to 34 and 19%, respectively. Among organic solvents, an addition of 1-butanol (20%) augmented the enzyme activity by 29% of the original activity. With casein as a substrate, the enzyme activity under optimized conditions was found to be 73.8 U mg(-1) . The effect of protease expression on the host cells growth was also studied and found to negatively affect E. coli cells to certain extent. Catalytic domains of serine proteases from eight important thermostable organisms were analyzed through WebLogo and found to be conserved in all serine protease sequences suggesting that protease of G. stearothermophilus could be beneficially used as a biocontrol agent and in many industries including detergent industry. PMID:25224381

  14. Mitochondrial Cyclic AMP Response Element-binding Protein (CREB) Mediates Mitochondrial Gene Expression and Neuronal Survival*S

    PubMed Central

    Lee, Junghee; Kim, Chun-Hyung; Simon, David K.; Aminova, Lyaylya R.; Andreyev, Alexander Y.; Kushnareva, Yulia E.; Murphy, Anne N.; Lonze, Bonnie E.; Kim, Kwang-Soo; Ginty, David D.; Ferrante, Robert J.; Ryu, Hoon; Ratan, Rajiv R.

    2008-01-01

    Cyclic AMP response element-binding protein (CREB) is a widely expressed transcription factor whose role in neuronal protection is now well established. Here we report that CREB is present in the mitochondrial matrix of neurons and that it binds directly to cyclic AMP response elements (CREs) found within the mitochondrial genome. Disruption of CREB activity in the mitochondria decreases the expression of a subset of mitochondrial genes, including the ND5 subunit of complex I, down-regulates complex I-dependent mitochondrial respiration, and increases susceptibility to 3-nitropropionic acid, a mitochondrial toxin that induces a clinical and pathological phenotype similar to Huntington disease. These results demonstrate that regulation of mitochondrial gene expression by mitochondrial CREB, in part, underlies the protective effects of CREB and raise the possibility that decreased mitochondrial CREB activity contributes to the mitochondrial dysfunction and neuronal loss associated with neurodegenerative disorders. PMID:16207717

  15. Gemini surfactants mediate efficient mitochondrial gene delivery and expression.

    PubMed

    Cardoso, Ana M; Morais, Catarina M; Cruz, A Rita; Cardoso, Ana L; Silva, Sandra G; do Vale, M Luísa; Marques, Eduardo F; Pedroso de Lima, Maria C; Jurado, Amália S

    2015-03-01

    Gene delivery targeting mitochondria has the potential to transform the therapeutic landscape of mitochondrial genetic diseases. Taking advantage of the nonuniversal genetic code used by mitochondria, a plasmid DNA construct able to be specifically expressed in these organelles was designed by including a codon, which codes for an amino acid only if read by the mitochondrial ribosomes. In the present work, gemini surfactants were shown to successfully deliver plasmid DNA to mitochondria. Gemini surfactant-based DNA complexes were taken up by cells through a variety of routes, including endocytic pathways, and showed propensity for inducing membrane destabilization under acidic conditions, thus facilitating cytoplasmic release of DNA. Furthermore, the complexes interacted extensively with lipid membrane models mimicking the composition of the mitochondrial membrane, which predicts a favored interaction of the complexes with mitochondria in the intracellular environment. This work unravels new possibilities for gene therapy toward mitochondrial diseases. PMID:25634573

  16. Evolutionary relationship of nuclear genes encoding mitochondrial proteins across grasses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Comparative genome studies were done across taxa to provide a basic understanding of genome evolution regarding nuclear genes encoding for mitochondrial proteins and their conservation in grass species. Two different mitochondria-related gene sets, one from rice and another from Arabidopsis, were us...

  17. The effect of environmental conditions on expression of Bacteroides fragilis and Bacteroides thetaiotaomicron C10 protease genes

    PubMed Central

    2012-01-01

    Background Bacteroides fragilis and Bacteroides thetaiotaomicron are members of the normal human intestinal microbiota. However, both organisms are capable of causing opportunistic infections, during which the environmental conditions to which the bacteria are exposed change dramatically. To further explore their potential for contributing to infection, we have characterized the expression in B. thetaiotaomicron of four homologues of the gene encoding the C10 cysteine protease SpeB, a potent extracellular virulence factor produced by Streptococcus pyogenes. Results We identified a paralogous set of genes (btp genes) in the B. thetaiotaomicron genome, that were related to C10 protease genes we recently identified in B. fragilis. Similar to C10 proteases found in B. fragilis, three of the B. thetaiotaomicron homologues were transcriptionally coupled to genes encoding small proteins that are similar in structural architecture to Staphostatins, protease inhibitors associated with Staphopains in Staphylococcus aureus. The expression of genes for these C10 proteases in both B. fragilis and B. thetaiotaomicron was found to be regulated by environmental stimuli, in particular by exposure to oxygen, which may be important for their contribution to the development of opportunistic infections. Conclusions Genes encoding C10 proteases are increasingly identified in operons which also contain genes encoding proteins homologous to protease inhibitors. The Bacteroides C10 protease gene expression levels are responsive to different environmental stimuli suggesting they may have distinct roles in the bacterial-host interaction. PMID:22943521

  18. Mitochondrial and Metabolic Gene Expression in the Aged Rat Heart.

    PubMed

    Barton, Gregory P; Sepe, Joseph J; McKiernan, Susan H; Aiken, Judd M; Diffee, Gary M

    2016-01-01

    Aging is associated with a decline in cardiac function. Exercise intervention has been suggested as a way to improve this decrement. Age-related decline in cardiac function is associated with decreases in fatty acid oxidation, mitochondrial function, and AMP-activated protein kinase (AMPK) activity. The molecular mechanisms involved with age-related changes in mitochondrial function and substrate metabolism are poorly understood. We determined gene expression differences in hearts of Young (6 mo), Old (33 mo), and old exercise trained (Old + EXE) (34 mo) FBN rats, using Qiagen PCR arrays for Glucose, Fatty acid, and Mitochondrial metabolism. Old rats demonstrated decreased (p < 0.05) expression for key genes in fatty acid oxidation, mitochondrial function, and AMPK signaling. There were no differences in the expression of genes involved in glucose metabolism with age. These gene expression changes occurred prior to altered protein translation as we found no differences in the protein content of peroxisome proliferator activated receptor gamma, coactivators 1 alpha (PGC-1α), peroxisome proliferator activated receptor alpha (PPARα), and AMPKα2 between young and old hearts. Four months of exercise training did not attenuate the decline in the gene expression in aged hearts. Despite this lack of change in gene expression, exercise-trained rats demonstrated increased exercise capacity compared to their sedentary counterparts. Taken together, our results show that differential expression of genes associated with fatty acid metabolism, AMPK signaling and mitochondrial function decrease in the aging heart which may play a role in age-related declines in fatty acid oxidation, AMPK activity, and mitochondrial function in the heart. PMID:27601998

  19. Mitochondrial and Metabolic Gene Expression in the Aged Rat Heart

    PubMed Central

    Barton, Gregory P.; Sepe, Joseph J.; McKiernan, Susan H.; Aiken, Judd M.; Diffee, Gary M.

    2016-01-01

    Aging is associated with a decline in cardiac function. Exercise intervention has been suggested as a way to improve this decrement. Age-related decline in cardiac function is associated with decreases in fatty acid oxidation, mitochondrial function, and AMP-activated protein kinase (AMPK) activity. The molecular mechanisms involved with age-related changes in mitochondrial function and substrate metabolism are poorly understood. We determined gene expression differences in hearts of Young (6 mo), Old (33 mo), and old exercise trained (Old + EXE) (34 mo) FBN rats, using Qiagen PCR arrays for Glucose, Fatty acid, and Mitochondrial metabolism. Old rats demonstrated decreased (p < 0.05) expression for key genes in fatty acid oxidation, mitochondrial function, and AMPK signaling. There were no differences in the expression of genes involved in glucose metabolism with age. These gene expression changes occurred prior to altered protein translation as we found no differences in the protein content of peroxisome proliferator activated receptor gamma, coactivators 1 alpha (PGC-1α), peroxisome proliferator activated receptor alpha (PPARα), and AMPKα2 between young and old hearts. Four months of exercise training did not attenuate the decline in the gene expression in aged hearts. Despite this lack of change in gene expression, exercise-trained rats demonstrated increased exercise capacity compared to their sedentary counterparts. Taken together, our results show that differential expression of genes associated with fatty acid metabolism, AMPK signaling and mitochondrial function decrease in the aging heart which may play a role in age-related declines in fatty acid oxidation, AMPK activity, and mitochondrial function in the heart. PMID:27601998

  20. MALT1 Protease Activity Controls the Expression of Inflammatory Genes in Keratinocytes upon Zymosan Stimulation.

    PubMed

    Schmitt, Anja; Grondona, Paula; Maier, Tabea; Brändle, Marc; Schönfeld, Caroline; Jäger, Günter; Kosnopfel, Corinna; Eberle, Franziska C; Schittek, Birgit; Schulze-Osthoff, Klaus; Yazdi, Amir S; Hailfinger, Stephan

    2016-04-01

    The protease activity of the paracaspase mucosa-associated lymphoid tissue lymphoma translocation gene 1 (MALT1) plays an important role in antigen receptor-mediated lymphocyte activation by controlling the activity of the transcription factor nuclear factor-κB and is thus essential for the expression of inflammatory target genes. MALT1 is not only present in cells of the hematopoietic lineage, but is ubiquitously expressed. Here we report that stimulation with zymosan or Staphylococcus aureus induced MALT1 protease activity in human primary keratinocytes. Inhibition of the Src family of kinases or novel protein kinase C isoforms as well as silencing of CARMA2 or BCL10 interfered with activation of MALT1 protease. Silencing or inhibition of MALT1 protease strongly decreased the expression of important inflammatory genes such as TNFα, IL-17C, CXCL8 and HBD-2. MALT1-inhibited cells were unable to mount an antimicrobial response upon zymosan stimulation or phorbolester/ionomycin treatment, demonstrating a central role of MALT1 protease activity in keratinocyte immunity and suggesting MALT1 as a potential target in inflammatory skin diseases. PMID:26767426

  1. Limitations of allotopic expression of mitochondrial genes in mammalian cells.

    PubMed Central

    Oca-Cossio, Jose; Kenyon, Lesley; Hao, Huiling; Moraes, Carlos T

    2003-01-01

    The possibility of expressing mitochondrial DNA-coded genes in the nuclear-cytoplasmic compartment provides an attractive system for genetic treatment of mitochondrial disorders associated with mitochondrial DNA mutations. In theory, by recoding mitochondrial genes to adapt them to the universal genetic code and by adding a DNA sequence coding for a mitochondrial-targeting sequence, one could achieve correct localization of the gene product. Such transfer has occurred in nature, and certain species of algae and plants express a number of polypeptides that are commonly coded by mtDNA in the nuclear-cytoplasmic compartment. In the present study, allotopic expression of three different mtDNA-coded polypeptides (ATPase8, apocytochrome b, and ND4) into COS-7 and HeLa cells was analyzed. Among these, only ATPase8 was correctly expressed and localized to mitochondria. The full-length, as well as truncated forms, of apocytochrome b and ND4 decorated the periphery of mitochondria, but also aggregated in fiber-like structures containing tubulin and in some cases also vimentin. The addition of a hydrophilic tail (EGFP) to the C terminus of these polypeptides did not change their localization. Overexpression of molecular chaperones also did not have a significant effect in preventing aggregations. Allotopic expression of apocytochrome b and ND4 induced a loss of mitochondrial membrane potential in transfected cells, which can lead to cell death. Our observations suggest that only a subset of mitochondrial genes can be replaced allotopically. Analyses of the hydrophobic patterns of different polypeptides suggest that hydrophobicity of the N-terminal segment is the main determinant for the importability of peptides into mammalian mitochondria. PMID:14573482

  2. Cysteine protease gene expression and proteolytic activity during senescence of Alstroemeria petals.

    PubMed

    Wagstaff, Carol; Leverentz, Michael K; Griffiths, Gareth; Thomas, Brian; Chanasut, Usawadee; Stead, Anthony D; Rogers, Hilary J

    2002-02-01

    The functional life of the flower is terminated by senescence and/or abscission. Multiple processes contribute to produce the visible signs of petal wilting and inrolling that typify senescence, but one of the most important is that of protein degradation and remobilization. This is mediated in many species through protein ubiquitination and the action of specific protease enzymes. This paper reports the changes in protein and protease activity during development and senescence of Alstroemeria flowers, a Liliaceous species that shows very little sensitivity to ethylene during senescence and which shows perianth abscission 8-10 d after flower opening. Partial cDNAs of ubiquitin (ALSUQ1) and a putative cysteine protease (ALSCYP1) were cloned from Alstroemeria using degenerate PCR primers and the expression pattern of these genes was determined semi-quantitatively by RT-PCR. While the levels of ALSUQ1 only fluctuated slightly during floral development and senescence, there was a dramatic increase in the expression of ALSCYP1 indicating that this gene may encode an important enzyme for the proteolytic process in this species. Three papain class cysteine protease enzymes showing different patterns of activity during flower development were identified on zymograms, one of which showed a similar expression pattern to the cysteine protease cDNA. PMID:11807127

  3. Production, characterization, gene cloning, and nematocidal activity of the extracellular protease from Stenotrophomonas maltophilia N4.

    PubMed

    Jankiewicz, Urszula; Larkowska, Ewa; Swiontek Brzezinska, Maria

    2016-06-01

    A rhizosphere strain of the bacterium Stenotrophomonas maltophilia N4 secretes the serine protease PN4, whose molecular mass is approximately 42 kDa. The optimal temperature for the enzyme activity of the 11-fold purified protein was 50°C and the optimal pH was 10.5. The activity of the enzyme was strongly inhibited by specific serine protease inhibitors, which allowed for its classification as an alkaline serine protease family. Ca(2+) ions stimulated the activity of the protease PN4, while Mg(2+) ions stabilized its activity, and Zn(2+) and Cd(2+) ions strongly inhibited its activity. The enzyme has broad substrate specificity. For example, it is able to hydrolyse casein, keratin, albumin, haemoglobin, and gelatin, as well as the insoluble modified substrates azure keratin and azocoll. The gene that encodes the 1740 bp precursor form of the enzyme (accession number: LC031815) was cloned. We then deduced that its amino acid sequence includes the region of the conserved domain of the S8 family of peptidases as well as the catalytic triad Asp/His/Ser. The bacterial culture fluid as well as the purified protease PN4 demonstrated biocidal activity with regard to the nematodes Caenorhabditis elegans and Panagrellus spp. PMID:26896861

  4. Polymorphisms in mitochondrial genes and prostate cancer risk

    PubMed Central

    Wang, Liang; McDonnell, Shannon K.; Hebbring, Scott J.; Cunningham, Julie M.; Sauver, Jennifer St; Cerhan, James R.; Isaya, Grazia; Schaid, Daniel J.; Thibodeau, Stephen N.

    2009-01-01

    The mitochondrion, conventionally thought to be an organelle specific to energy metabolism, is in fact multi-functional and implicated in many diseases, including cancer. To evaluate whether mitochondria-related genes are associated with increased risk for prostate cancer, we genotyped 24 single nucleotide polymorphisms (SNPs) within the mitochondrial genome (mtSNPs) and 376 tagSNPs localized to 78 nuclear-encoded mitochondrial genes. The tagSNPs were selected to achieve ≥80% coverage based on linkage disequilibrium. We compared allele and haplotype frequencies in ~1000 prostate cancer cases with ~500 population controls. An association with prostate cancer was not detected for any of the mtSNPs individually or for 10 mitochondrial common haplotypes when evaluated using a global score statistic. For the nuclear-encoded genes, none of the tagSNPs were significantly associated with prostate cancer after adjusting for multiple testing. Nonetheless, we evaluated unadjusted p-values by comparing our results with those from the CGEMS phase I data set. Seven tagSNPs had unadjusted p-values ≤ 0.05 in both our data and in CGEMS (two SNPs were identical and five were in strong linkage disequilibrium with CGEMS SNPs). These seven SNPs (rs17184211, rs4147684, rs4233367, rs2070902, rs3829037, rs7830235, and rs1203213) are located in genes MTRR, NDUFA9, NDUFS2, NDUFB9 and COX7A2, respectively. Five of the seven SNPs were further included in the CGEMS phase II study, however, none of the findings for these were replicated. Overall, these results suggest that polymorphisms in the mitochondrial genome and those in the nuclear encoded mitochondrial genes evaluated are not substantial risk factors for prostate cancer. PMID:19064571

  5. Protease Omi cleaving Hax-1 protein contributes to OGD/R-induced mitochondrial damage in neuroblastoma N2a cells and cerebral injury in MCAO mice

    PubMed Central

    Wu, Jia-yuan; Li, Mei; Cao, Li-juan; Sun, Mei-ling; Chen, Dong; Ren, Hai-gang; Xia, Qin; Tao, Zhou-teng; Qin, Zheng-hong; Hu, Qing-song; Wang, Guang-hui

    2015-01-01

    Aim: In the penumbra after focal cerebral ischemia, an increase of protease Omi is linked to a decrease of Hs1-associated protein X-1 (Hax-1), a protein belonging to the Bcl-2 family. In this study we investigated the mechanisms underlying the regulation of Hax-1 by protease Omi in cerebral ischemia/reperfusion (I/R) injury. Methods: Mouse neuroblastoma N2a cells were subjected to oxygen-glucose deprivation and reoxygenation (OGD/R); cell viability was assessed with MTT assay. Mice underwent 2-h middle cerebral artery occlusion (MCAO) and reperfusion, and the infarct volume was determined with TTC staining. The expression of Omi and Hax-1 was detected using immunoblot and immunofluorescence assays. The mitochondrial membrane potential was measured using TMRM staining. Results: In the brains of MCAO mice, the protein level of Omi was significantly increased, while the protein level of Hax-1 was decreased. Similar changes were observed in OGD/R-treated N2a cells, but the mRNA level of Hax-1 was not changed. Furthermore, in OGD/R-treated N2a cells, knockdown of Omi significantly increased Hax-1 protein level. Immunofluorescence assay showed that Omi and Hax-1 were co-localized in mitochondria of N2a cells. OGD/R caused marked mitochondrial damage and apoptosis in N2a cells, while inhibition of Omi protease activity with UCF-101 (10 μmol/L) or overexpression of Hax-1 could restore the mitochondrial membrane potential and attenuate cell apoptosis. Moreover, pretreatment of MCAO mice with UCF-101 (7.15 mg/kg, ip) could restore Hax-1 expression, inhibit caspase activation, and significantly reduce the infarct volume. Conclusion: Protease Omi impairs mitochondrial function by cleaving Hax-1, which induces apoptosis in OGD/R-treated N2a cells and causes I/R injury in MCAO mice. PMID:26299953

  6. Alternative splicing, a new target to block cellular gene expression by poliovirus 2A protease

    SciTech Connect

    Alvarez, Enrique; Castello, Alfredo; Carrasco, Luis; Izquierdo, Jose M.

    2011-10-14

    Highlights: {yields} Novel role for poliovirus 2A protease as splicing modulator. {yields} Poliovirus 2A protease inhibits the alternative splicing of pre-mRNAs. {yields} Poliovirus 2A protease blocks the second catalytic step of splicing. -- Abstract: Viruses have developed multiple strategies to interfere with the gene expression of host cells at different stages to ensure their own survival. Here we report a new role for poliovirus 2A{sup pro} modulating the alternative splicing of pre-mRNAs. Expression of 2A{sup pro} potently inhibits splicing of reporter genes in HeLa cells. Low amounts of 2A{sup pro} abrogate Fas exon 6 skipping, whereas higher levels of protease fully abolish Fas and FGFR2 splicing. In vitro splicing of MINX mRNA using nuclear extracts is also strongly inhibited by 2A{sup pro}, leading to accumulation of the first exon and the lariat product containing the unspliced second exon. These findings reveal that the mechanism of action of 2A{sup pro} on splicing is to selectively block the second catalytic step.

  7. Serine and cysteine protease-like genes in the genome of a gall midge and their interactions with host plant genotypes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    For plant-feeding insects, digestive proteases are targets for engineering protease inhibitors for pest control. In this study, we identified 105 putative serine- and cysteine-protease genes from Hessian fly genome. Among the genes, 31 encode putative trypsins, 18 encode putative chymotrypsins, se...

  8. Functional diversification of a protease inhibitor gene in the genus Drosophila and its molecular basis.

    PubMed

    Börner, Stefan; Ragg, Hermann

    2008-05-31

    The mutually exclusive use of alternative reactive site loop (RSL) cassettes due to alternative splicing of serpin (serine protease inhibitor) gene transcripts is a widespread strategy to create target-selective protease inhibitors in the animal kingdom. Since molecular basis and evolution of serpin RSL cassette exon amplification and diversification are unexplored, the exon-intron organization of the serpin gene spn4 from 12 species of the genus Drosophila was studied. The analysis of the gene structures shows that both number and target enzyme specificities of Spn4 RSL cassettes are highly variable in fruit flies and includes inhibitor variants with novel antiproteolytic activities in some species, indicating that RSL diversity is the result of adaptive evolution. Comparative genomics suggests that interallelic gene conversion and/or recombination events contribute to RSL cassette exon amplification. Due to an intron that is located at the most suitable position within the RSL region, multiple inhibitors can be formed in an economic manner that are both efficient and target-selective, allowing fruit flies to control an astonishing variety of proteases with different cleavage chemistry and evolutionary ancestry. PMID:18395367

  9. Evidence of a bigenomic regulation of mitochondrial gene expression by thyroid hormone during rat brain development

    SciTech Connect

    Sinha, Rohit Anthony; Pathak, Amrita; Mohan, Vishwa; Babu, Satish; Pal, Amit; Khare, Drirh; Godbole, Madan M.

    2010-07-02

    Hypothyroidism during early mammalian brain development is associated with decreased expression of various mitochondrial encoded genes along with evidence for mitochondrial dysfunction. However, in-spite of the similarities between neurological disorders caused by perinatal hypothyroidism and those caused by various genetic mitochondrial defects we still do not know as to how thyroid hormone (TH) regulates mitochondrial transcription during development and whether this regulation by TH is nuclear mediated or through mitochondrial TH receptors? We here in rat cerebellum show that hypothyroidism causes reduction in expression of nuclear encoded genes controlling mitochondrial biogenesis like PGC-1{alpha}, NRF-1{alpha} and Tfam. Also, we for the first time demonstrate a mitochondrial localization of thyroid hormone receptor (mTR) isoform in developing brain capable of binding a TH response element (DR2) present in D-loop region of mitochondrial DNA. These results thus indicate an integrated nuclear-mitochondrial cross talk in regulation of mitochondrial transcription by TH during brain development.

  10. Mitochondrial Lon protease controls ROS-dependent apoptosis in cardiomyocyte under hypoxia.

    PubMed

    Kuo, Chan-Yen; Chiu, Yi-Chieh; Lee, Alan Yueh-Luen; Hwang, Tsong-Long

    2015-07-01

    Apoptosis of cardiomyocytes, under ischemic conditions, has been identified as an essential process in the progression of heart failure. Under hypoxic conditions, mitochondria can become a threat to the cell because of their capacity to generate reactive oxygen species (ROS). As ROS appear to have a critical role in heart failure, there has been considerable interest in identifying the candidate proteins involved and in developing strategies to reduce oxidative stress. Lon protease (Lon) is a multifunctional protein that mediates protein quality control and stress response in mitochondria. However, comprehensive and detailed studies, on the role of Lon in hypoxia-induced cardiomyocyte apoptosis, have yet to be carried out. In the present study, we demonstrated that hypoxia induced ROS-dependent cardiomyocyte apoptosis. Lon was upregulated in hypoxia-induced cardiomyocytes. Lon downregulation attenuated hypoxia-induced cardiomyocyte apoptosis through a reduction of ROS level. Moreover, overexpression of Lon stimulated ROS production and induced apoptosis under normoxic conditions in cardiomyocytes. Our results identify Lon as a novel regulator of cardiomyocyte fate and offers exciting new insights into the therapeutic potential of hypoxia-induced cardiomyocyte apoptosis. PMID:25922169

  11. PLANT-PIs: a database for plant protease inhibitors and their genes

    PubMed Central

    De Leo, F.; Volpicella, M.; Licciulli, F.; Liuni, S.; Gallerani, R.; Ceci, L. R.

    2002-01-01

    PLANT-PIs is a database developed to facilitate retrieval of information on plant protease inhibitors (PIs) and related genes. For each PI, links to sequence databases are reported together with a summary of the functional properties of the molecule (and its mutants) as deduced from literature. PLANT-PIs contains information for 351 plant PIs, plus several isoinhibitors. The database is accessible at http://bighost.area.ba.cnr.it/PLANT-PIs. PMID:11752333

  12. Genomic modulation of mitochondrial respiratory genes in the hypertrophied heart reflects adaptive changes in mitochondrial and contractile function

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We hypothesized the coordinate induction of mitochondrial regulatory genes in the hypertrophied right ventricle to sustain mitochondrial respiratory capacity and contractile function in response to increased load. Wistar rats were exposed to hypobaric hypoxia (11% O(2)) or normoxia for 2 wk. Cardiac...

  13. Molecular mechanisms of extensive mitochondrial gene rearrangementin plethodontid salamanders

    SciTech Connect

    Mueller, Rachel Lockridge; Boore, Jeffrey L.

    2005-06-01

    Extensive gene rearrangement is reported in the mitochondrial genomes of lungless salamanders (Plethodontidae). In each genome with a novel gene order, there is evidence that the rearrangement was mediated by duplication of part of the mitochondrial genome, including the presence of both pseudogenes and additional, presumably functional, copies of duplicated genes. All rearrangement-mediating duplications include either the origin of light strand replication and the nearby tRNA genes or the regions flanking the origin of heavy strand replication. The latter regions comprise nad6, trnE, cob, trnT, an intergenic spacer between trnT and trnP and, in some genomes, trnP, the control region, trnF, rrnS, trnV, rrnL, trnL1, and nad1. In some cases, two copies of duplicated genes, presumptive regulatory regions, and/or sequences with no assignable function have been retained in the genome following the initial duplication; in other genomes, only one of the duplicated copies has been retained. Both tandem and non-tandem duplications are present in these genomes, suggesting different duplication mechanisms. In some of these mtDNAs, up to 25 percent of the total length is composed of tandem duplications of non-coding sequence that includes putative regulatory regions and/or pseudogenes of tRNAs and protein-coding genes along with otherwise unassignable sequences. These data indicate that imprecise initiation and termination of replication, slipped-strand mispairing, and intra-molecular recombination may all have played a role in generating repeats during the evolutionary history of plethodontid mitochondrial genomes.

  14. Human mast cell tryptase: Multiple cDNAs and genes reveal a multigene serine protease family

    SciTech Connect

    Vanderslice, P.; Ballinger, S.M., Tam, E.K.; Goldstein, S.M.; Craik, C.S.; Caughey, G.H. )

    1990-05-01

    Three different cDNAs and a gene encoding human skin mast cell tryptase have been cloned and sequenced in their entirety. The deduced amino acid sequences reveal a 30-amino acid prepropeptide followed by a 245-amino acid catalytic domain. The C-terminal undecapeptide of the human preprosequence is identical in dog tryptase and appears to be part of a prosequence unique among serine proteases. The differences among the three human tryptase catalytic domains include the loss of a consensus N-glycosylation site in one cDNA, which may explain some of the heterogeneity in size and susceptibility to deglycosylation seen in tryptase preparations. All three tryptase cDNAs are distinct from a recently reported cDNA obtained from a human lung mast cell library. A skin tryptase cDNA was used to isolate a human tryptase gene, the exons of which match one of the skin-derived cDNAs. The organization of the {approx}1.8-kilobase-pair tryptase gene is unique and is not closely related to that of any other mast cell or leukocyte serine protease. The 5{prime} regulatory regions of the gene share features with those of other serine proteases, including mast cell chymase, but are unusual in being separated from the protein-coding sequence by an intron. High-stringency hybridization of a human genomic DNA blot with a fragment of the tryptase gene confirms the presence of multiple tryptase genes. These findings provide genetic evidence that human mast cell tryptases are the products of a multigene family.

  15. The impact of mitochondrial DNA and nuclear genes related to mitochondrial functioning on the risk of Parkinson's disease.

    PubMed

    Gaweda-Walerych, Katarzyna; Zekanowski, Cezary

    2013-12-01

    Mitochondrial dysfunction and oxidative stress are the major factors implicated in Parkinson's disease (PD) pathogenesis. The maintenance of healthy mitochondria is a very complex process coordinated bi-genomically. Here, we review association studies on mitochondrial haplogroups and subhaplogroups, discussing the underlying molecular mechanisms. We also focus on variation in the nuclear genes (NDUFV2, PGC-1alpha, HSPA9, LRPPRC, MTIF3, POLG1, and TFAM encoding NADH dehydrogenase (ubiquinone) flavoprotein 2, peroxisome proliferator-activated receptor gamma coactivator 1-alpha, mortalin, leucine-rich pentatricopeptide repeat containing protein, translation initiation factor 3, mitochondrial DNA polymerase gamma, and mitochondrial transcription factor A, respectively) primarily linked to regulation of mitochondrial functioning that recently have been associated with PD risk. Possible interactions between mitochondrial and nuclear genetic variants and related proteins are discussed. PMID:24532986

  16. The Impact of Mitochondrial DNA and Nuclear Genes Related to Mitochondrial Functioning on the Risk of Parkinson’s Disease

    PubMed Central

    Gaweda-Walerych, Katarzyna; Zekanowski, Cezary

    2013-01-01

    Mitochondrial dysfunction and oxidative stress are the major factors implicated in Parkinson’s disease (PD) pathogenesis. The maintenance of healthy mitochondria is a very complex process coordinated bi-genomically. Here, we review association studies on mitochondrial haplogroups and subhaplogroups, discussing the underlying molecular mechanisms. We also focus on variation in the nuclear genes (NDUFV2, PGC-1alpha, HSPA9, LRPPRC, MTIF3, POLG1, and TFAM encoding NADH dehydrogenase (ubiquinone) flavoprotein 2, peroxisome proliferator-activated receptor gamma coactivator 1-alpha, mortalin, leucine-rich pentatricopeptide repeat containing protein, translation initiation factor 3, mitochondrial DNA polymerase gamma, and mitochondrial transcription factor A, respectively) primarily linked to regulation of mitochondrial functioning that recently have been associated with PD risk. Possible interactions between mitochondrial and nuclear genetic variants and related proteins are discussed. PMID:24532986

  17. Genes or culture: are mitochondrial genes associated with tool use in bottlenose dolphins (Tursiops sp.)?

    PubMed

    Bacher, K; Allen, S; Lindholm, A K; Bejder, L; Krützen, M

    2010-09-01

    Some bottlenose dolphins use marine sponges as foraging tools ('sponging'), which appears to be socially transmitted from mothers mainly to their female offspring. Yet, explanations alternative to social transmission have been proposed. Firstly, the propensity to engage in sponging might be due to differences in diving ability caused by variation of mitochondrial genes coding for proteins of the respiratory chain. Secondly, the cultural technique of sponging may have selected for changes in these same genes (or other autosomal ones) among its possessors. We tested whether sponging can be predicted by mitochondrial coding genes and whether these genes are under selection. In 29 spongers and 54 non-spongers from two study sites, the non-coding haplotype at the HVRI locus was a significant predictor of sponging, whereas the coding mitochondrial genes were not. There was no evidence of selection in the investigated genes. Our study shows that mitochondrial gene variation is unlikely to be a viable alternative to cultural transmission as a primary driver of tool use in dolphins. PMID:20582623

  18. Characterization and gene cloning of a novel serine protease with nematicidal activity from Trichoderma pseudokoningii SMF2.

    PubMed

    Chen, Lei-Lei; Liu, Li-Jun; Shi, Mei; Song, Xiao-Yan; Zheng, Chang-Ying; Chen, Xiu-Lan; Zhang, Yu-Zhong

    2009-10-01

    Trichoderma pseudokoningii SMF2 is a biocontrol fungus with inhibitory ability against phytopathogenic fungi. Here, a crude extract of strain SMF2 in a solid ferment exhibited strong nematicidal activity against Meloidogyne incognita, and a novel serine protease SprT with nematicidal activity was purified from the crude extract. Protease SprT has a molecular mass of 31 kDa, a pH optimum of 8.5, and a temperature optimum of 60-65 degrees C. It had good thermostability, and was stable in an alkaline environment. SprT could degrade bovine serum albumin, lysozyme, and gelatin, and its activity was enhanced by many metal ions. The cuticles of nematodes treated by protease SprT obviously crimpled. Purified protease SprT could kill juveniles of M. incognita and inhibit egg hatch, suggesting that it is involved in the nematicidal process of T. pseudokoningii SMF2. The full-length cDNA gene-encoding protease SprT was cloned by rapid amplification of cDNA ends. Sequence analysis showed that SprT is a monodomain subtilase containing 284 amino acid residues. It had higher identities and a closer relation to the nematicidal serine proteases (59-69%) from nematode parasitic fungi than to the serine proteases (<50%) from Trichoderma. Protease SprT represents the first well-characterized subtilase with nematicidal activity from Trichoderma. PMID:19702879

  19. Decrypting the Mitochondrial Gene Pool of Modern Panamanians

    PubMed Central

    Angerhofer, Norman; Ekins, Jayne E.; Olivieri, Anna; Woodward, Scott R.; Pascale, Juan Miguel; Cooke, Richard; Motta, Jorge; Achilli, Alessandro

    2012-01-01

    The Isthmus of Panama–the narrow neck of land connecting the northern and southern American landmasses–was an obligatory corridor for the Paleo-Indians as they moved into South America. Archaeological evidence suggests an unbroken link between modern natives and their Paleo-Indian ancestors in some areas of Panama, even if the surviving indigenous groups account for only 12.3% of the total population. To evaluate if modern Panamanians have retained a larger fraction of the native pre-Columbian gene pool in their maternally-inherited mitochondrial genome, DNA samples and historical records were collected from more than 1500 volunteer participants living in the nine provinces and four indigenous territories of the Republic. Due to recent gene-flow, we detected ∼14% African mitochondrial lineages, confirming the demographic impact of the Atlantic slave trade and subsequent African immigration into Panama from Caribbean islands, and a small European (∼2%) component, indicating only a minor influence of colonialism on the maternal side. The majority (∼83%) of Panamanian mtDNAs clustered into native pan-American lineages, mostly represented by haplogroup A2 (51%). These findings reveal an overwhelming native maternal legacy in today's Panama, which is in contrast with the overall concept of personal identity shared by many Panamanians. Moreover, the A2 sub-clades A2ad and A2af (with the previously named 6 bp Huetar deletion), when analyzed at the maximum level of resolution (26 entire mitochondrial genomes), confirm the major role of the Pacific coastal path in the peopling of North, Central and South America, and testify to the antiquity of native mitochondrial genomes in Panama. PMID:22675545

  20. Decrypting the mitochondrial gene pool of modern Panamanians.

    PubMed

    Perego, Ugo A; Lancioni, Hovirag; Tribaldos, Maribel; Angerhofer, Norman; Ekins, Jayne E; Olivieri, Anna; Woodward, Scott R; Pascale, Juan Miguel; Cooke, Richard; Motta, Jorge; Achilli, Alessandro

    2012-01-01

    The Isthmus of Panama--the narrow neck of land connecting the northern and southern American landmasses--was an obligatory corridor for the Paleo-Indians as they moved into South America. Archaeological evidence suggests an unbroken link between modern natives and their Paleo-Indian ancestors in some areas of Panama, even if the surviving indigenous groups account for only 12.3% of the total population. To evaluate if modern Panamanians have retained a larger fraction of the native pre-Columbian gene pool in their maternally-inherited mitochondrial genome, DNA samples and historical records were collected from more than 1500 volunteer participants living in the nine provinces and four indigenous territories of the Republic. Due to recent gene-flow, we detected ~14% African mitochondrial lineages, confirming the demographic impact of the Atlantic slave trade and subsequent African immigration into Panama from Caribbean islands, and a small European (~2%) component, indicating only a minor influence of colonialism on the maternal side. The majority (~83%) of Panamanian mtDNAs clustered into native pan-American lineages, mostly represented by haplogroup A2 (51%). These findings reveal an overwhelming native maternal legacy in today's Panama, which is in contrast with the overall concept of personal identity shared by many Panamanians. Moreover, the A2 sub-clades A2ad and A2af (with the previously named 6 bp Huetar deletion), when analyzed at the maximum level of resolution (26 entire mitochondrial genomes), confirm the major role of the Pacific coastal path in the peopling of North, Central and South America, and testify to the antiquity of native mitochondrial genomes in Panama. PMID:22675545

  1. Suppression of cytoplasmic male sterility by nuclear genes alters expression of a novel mitochondrial gene region.

    PubMed Central

    Singh, M; Brown, G G

    1991-01-01

    To identify regions of the mitochondrial genome that potentially could specify the "Polima" (pol) cytoplasmic male sterility (CMS) of Brassica napus, transcripts of 14 mitochondrial genes from nap (male fertile), pol (male sterile), and nuclear fertility-restored pol cytoplasm plants were analyzed. Transcriptional differences among these plants were detected only with the ATPase subunit 6 (atp6) gene. Structural analysis of the atp6 gene regions of pol and nap mitochondrial DNAs showed that rearrangements in the pol mitochondrial genome occurring upstream of atp6 have generated a chimeric 224-codon open reading frame, designated orf224, that is cotranscribed with atp6. In CMS plants, most transcripts of this region are dicistronic, comprising both orf224 and atp6 sequences. Nuclear restorer genes at either of two distinct loci appear to specifically alter this transcript pattern such that monocistronic atp6 transcripts predominate. The differences in expression of this region appear to result, in part, from differential processing of a tRNA-like element comprising a tRNA pseudogene present immediately upstream of atp6 in both the sterile and fertile mitochondrial DNAs. Possible mechanisms by which expression of the orf224/atp6 locus and the Polima CMS trait may be specifically related are considered. PMID:1840901

  2. Review: Progress in the Researches on Insect Mitochondrial Genome and Analysis of Gene Order

    NASA Astrophysics Data System (ADS)

    Hu, Li; Jianyu, Gao; Haiyu, Liu; Wanzhi, Cai

    2009-04-01

    Insect mitochondrial genome is a double-stranded circular genomes which range from 14,503 bp to 19,571 bp in size. Nearly all the sequenced insect mitochondrial genomes encode 37 genes: two for rRNAs, 13 for proteins and 22 for tRNAs. This review compares and summarizes the features of complete mitochondrial genomes from 175 sequenced species of insects in 22 orders. The genomic organization, contents, gene order, and rearrangements of gene order are analyzed.

  3. Force generation and protease gene expression in organotypic co-cultures of fibroblasts and keratinocytes.

    PubMed

    Wall, Ivan B; Bhadal, Navneet; Broad, Simon; Whawell, Simon A; Mudera, Vivek; Lewis, Mark P

    2009-12-01

    Fibroblast-epithelium interactions are crucial for successful tissue engineering of skin and oral mucosal equivalents. In this study, we assessed early force generation in organotypic fibroblast-epithelium co-cultures, using normal human keratinocytes (NHK) and HPV16-transformed (UP) cells. During the initial 2 h period, organotypic co-cultures containing both epithelial cell types produced significantly more force than fibroblasts alone (p < 0.05). After 2 h, the epithelial contribution became diminished and did not significantly contribute to intrinsic force generation by fibroblasts, and no differences were observed when using UP vs. NHK. We then measured protease gene expression at the end of the experimental period. Distinct differences were evident in protease expression both between NHK-human skin fibroblast (HSF) vs. UP-HSF co-cultures and compared to fibroblasts alone. We conclude that whilst the very early contractile response of fibroblasts is enhanced by the overlying epithelium, this becomes diminished as the fibroblast response becomes predominant and it does contribute to tissue remodelling via regulation of protease expression. PMID:19701934

  4. Biogenesis of mitochondria: the mitochondrial gene (aap1) coding for mitochondrial ATPase subunit 8 in Saccharomyces cerevisiae.

    PubMed Central

    Macreadie, I G; Novitski, C E; Maxwell, R J; John, U; Ooi, B G; McMullen, G L; Lukins, H B; Linnane, A W; Nagley, P

    1983-01-01

    A mitochondrial gene (denoted aap1) in Saccharomyces cerevisiae has been characterized by nucleotide sequence analysis of a region of mtDNA between the oxi3 and oli2 genes. The reading frame of the aap1 gene specifies a hydrophobic polypeptide containing 48 amino acids. The functional nature of this reading frame was established by sequence analysis of a series of mit- mutants and revertants. Evidence is presented that the aap1 gene codes for a mitochondrially synthesized polypeptide associated with the mitochondrial ATPase complex. This polypeptide (denoted subunit 8) is a proteolipid whose size has been previously assumed to be 10 kilodaltons based on its mobility on SDS-polyacrylamide gels, but the sequence of the aap1 gene predicts a molecular weight of 5,815 for this protein. PMID:6223276

  5. Mitochondrial gene expression, antioxidant responses, and histopathology after cadmium exposure.

    PubMed

    Al Kaddissi, Simone; Legeay, Alexia; Elia, Antonia Concetta; Gonzalez, Patrice; Floriani, Magali; Cavalie, Isabelle; Massabuau, Jean-Charles; Gilbin, Rodolphe; Simon, Olivier

    2014-08-01

    The present study investigates cadmium effects on the transcription of mitochondrial genes of Procambarus clarkii after acute (0.05, 0.5, and 5 mg Cd/L; 4-10 days) and chronic exposures (10 μg Cd/L; 30-60 days). Transcriptional responses of cox1, atp6, and 12S using quantitative real-time RT-PCR were assessed in gills and hepatopancreas. Additionally, the expression levels of genes involved in detoxification and/or oxidative stress responses [mt, sod(Mn)] and enzymatic activities of antioxidants (SOD, CAT, GPX, and GST) were analyzed. The histopathological effects in hepatopancreas of crayfish were evaluated by light microscopy. Relationships between endpoints at different levels of biological organization and Cd bioaccumulation were also examined. Cd induced high levels of bioaccumulation, which was followed by mitochondrial dysfunction and histological alterations in both experiments. Moreover, perturbations in the defence mechanisms against oxidative stress tended to increase with time. Results also showed that molecular responses can vary depending on the intensity and duration of the chemical stress applied to the organisms and that the study of mt gene expression levels seemed to be the best tool to assess Cd intoxication. PMID:23065898

  6. The Spn4 gene from Drosophila melanogaster is a multipurpose defence tool directed against proteases from three different peptidase families

    PubMed Central

    Brüning, Mareke; Lummer, Martina; Bentele, Caterina; Smolenaars, Marcel M. W.; Rodenburg, Kees W.; Ragg, Hermann

    2006-01-01

    By alternative use of four RSL (reactive site loop) coding exon cassettes, the serpin (serine protease inhibitor) gene Spn4 from Drosophila melanogaster was proposed to enable the synthesis of multiple protease inhibitor isoforms, one of which has been shown to be a potent inhibitor of human furin. Here, we have investigated the inhibitory spectrum of all Spn4 RSL variants. The analyses indicate that the Spn4 gene encodes inhibitors that may inhibit serine proteases of the subtilase family (S8), the chymotrypsin family (S1), and the papain-like cysteine protease family (C1), most of them at high rates. Thus a cohort of different protease inhibitors is generated simply by grafting enzyme-adapted RSL sequences on to a single serpin scaffold, even though the target proteases contain different types and/or a varying order of catalytic residues and are descendents of different phylogenetic lineages. Since all of the Spn4 RSL isoforms are produced as intracellular residents and additionally as variants destined for export or associated with the secretory pathway, the Spn4 gene represents a versatile defence tool kit that may provide multiple antiproteolytic functions. PMID:16989645

  7. IgA Protease Activity in Haemophilus parasuis in the Absence of a Recognizable IgA Protease Gene

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background. Haemophilus parasuis, the bacterium responsible for Glasser’s disease, is a pathogen of significant concern in modern high-health swine production systems. Little is known regarding the molecular mechanisms of H. parasuis infection. In some Pasteurellaceae species, IgA proteases aid in d...

  8. Effects of dietary fatty acids on mitochondrial phospholipid compositions, oxidative status and mitochondrial gene expression of zebrafish at different ages.

    PubMed

    Betancor, M B; Almaida-Pagán, P F; Hernández, A; Tocher, D R

    2015-10-01

    Mitochondrial decay is generally associated with impairment in the organelle bioenergetics function and increased oxidative stress, and it appears that deterioration of mitochondrial inner membrane phospholipids (PL) and accumulation of mitochondrial DNA (mtDNA) mutations are among the main mechanisms involved in this process. In the present study, mitochondrial membrane PL compositions, oxidative status (TBARS content and SOD activity) and mtDNA gene expression of muscle and liver were analyzed in zebrafish fed two diets with lipid supplied either by rapeseed oil (RO) or a blend 60:40 of RO and DHA500 TG oil (DHA). Two feeding trials were performed using zebrafish from the same population of two ages (8 and 21 months). Dietary FA composition affected fish growth in 8-month-old animals, which could be related to an increase in stress promoted by diet composition. Lipid peroxidation was considerably higher in mitochondria of 8-month-old zebrafish fed the DHA diet than in animals fed the RO diet. This could indicate higher oxidative damage to mitochondrial lipids, very likely due to increased incorporation of DHA in PL of mitochondrial membranes. Lipids would be among the first molecules affected by mitochondrial reactive oxygen species, and lipid peroxidation could propagate oxidative reactions that would damage other molecules, including mtDNA. Mitochondrial lipid peroxidation and gene expression of 21-month-old fish showed lower responsiveness to diet composition than those of younger fish. Differences found in the effect of diet composition on mitochondrial lipids between the two age groups could be indicating age-related changes in the ability to maintain structural homeostasis of mitochondrial membranes. PMID:26156499

  9. The mitochondrial ATP-dependent Lon protease: a novel target in lymphoma death mediated by the synthetic triterpenoid CDDO and its derivatives

    PubMed Central

    Venkatesh, Sundararajan; Li, Min; Lee, Jae; Lu, Bin; Hilchey, Shannon P.; Morse, Kimberly M.; Metcalfe, Hollie M.; Skalska, Jolanta; Andreeff, Michael; Brookes, Paul S.

    2012-01-01

    Synthetic triterpenoids are multitarget compounds exhibiting promise as preventative and therapeutic agents for cancer. Their proposed mechanism of action is by forming Michael adducts with reactive nucleophilic groups on target proteins. Our previous work demonstrates that the 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO) and its derivatives promote B-lymphoid cell apoptosis through a mitochondria-mediated pathway linked to mitochondrial protein aggregation. As one function of the Lon protease is to eliminate abnormal mitochondrial proteins, we hypothesized that CDDO-induced protein aggregation and lymphoma apoptosis occur by inactivating this enzyme. Here, we show that CDDO and its derivatives directly and selectively inhibit Lon. CDDO blocks Lon-mediated proteolysis in biochemical and cellular assays, but does not inhibit the 20S proteasome. Furthermore, a biotinylated-CDDO conjugate modifies mitochondrial Lon. A striking common phenotype of CDDO-treated lymphoma cells and Lon-knockdown cells is the accumulation of electron-dense aggregates within mitochondria. We also show that Lon protein levels are substantially elevated in malignant lymphoma cells, compared with resting or activated B cells. Finally, we demonstrate that Lon knockdown leads to lymphoma cell death. Together, these findings suggest that Lon inhibition plays a contributory role in CDDO-induced lymphoma cell death, and support the concept that mitochondrial Lon is a novel anticancer drug target. PMID:22323447

  10. Progressive mitochondrial myopathy, deafness, and sporadic seizures associated with a novel mutation in the mitochondrial tRNASer(AGY) gene.

    PubMed

    Cardaioli, Elena; Malfatti, Edoardo; Da Pozzo, Paola; Gallus, Gian Nicola; Carluccio, Maria Alessandra; Rufa, Alessandra; Volpi, Nila; Dotti, Maria Teresa; Federico, Antonio

    2011-04-15

    We sequenced the mitochondrial genome from a patient with progressive mitochondrial myopathy associated with deafness, sporadic seizures, and histological and biochemical features of mitochondrial respiratory chain dysfunction. Direct sequencing showed a heteroplasmic mutation at nucleotide 12262 in the tRNASer(AGY) gene. RFLP analysis confirmed that 63% of muscle mtDNA harboured the mutation, while it was absent in all the other tissues. The mutation is predicted to influence the functional behaviour of the aminoacyl acceptor stem of the tRNA. Several point mutations on mitochondrial tRNA genes have been reported in patients affected by encephalomyopathies, but between them only four were reported for tRNASer(AGY). PMID:21257182

  11. Genome-wide identification, evolutionary and expression analysis of the aspartic protease gene superfamily in grape

    PubMed Central

    2013-01-01

    Background Aspartic proteases (APs) are a large family of proteolytic enzymes found in almost all organisms. In plants, they are involved in many biological processes, such as senescence, stress responses, programmed cell death, and reproduction. Prior to the present study, no grape AP gene(s) had been reported, and their research on woody species was very limited. Results In this study, a total of 50 AP genes (VvAP) were identified in the grape genome, among which 30 contained the complete ASP domain. Synteny analysis within grape indicated that segmental and tandem duplication events contributed to the expansion of the grape AP family. Additional analysis between grape and Arabidopsis demonstrated that several grape AP genes were found in the corresponding syntenic blocks of Arabidopsis, suggesting that these genes arose before the divergence of grape and Arabidopsis. Phylogenetic relationships of the 30 VvAPs with the complete ASP domain and their Arabidopsis orthologs, as well as their gene and protein features were analyzed and their cellular localization was predicted. Moreover, expression profiles of VvAP genes in six different tissues were determined, and their transcript abundance under various stresses and hormone treatments were measured. Twenty-seven VvAP genes were expressed in at least one of the six tissues examined; nineteen VvAPs responded to at least one abiotic stress, 12 VvAPs responded to powdery mildew infection, and most of the VvAPs responded to SA and ABA treatments. Furthermore, integrated synteny and phylogenetic analysis identified orthologous AP genes between grape and Arabidopsis, providing a unique starting point for investigating the function of grape AP genes. Conclusions The genome-wide identification, evolutionary and expression analyses of grape AP genes provide a framework for future analysis of AP genes in defining their roles during stress response. Integrated synteny and phylogenetic analyses provide novel insight into the

  12. Construction of a yeast strain devoid of mitochondrial introns and its use to screen nuclear genes involved in mitochondrial splicing.

    PubMed Central

    Séraphin, B; Boulet, A; Simon, M; Faye, G

    1987-01-01

    We have constructed a respiring yeast strain devoid of mitochondrial introns to screen nuclear pet- mutants for those that play a direct role in mitochondrial intron excision. Intron-less mitochondria are introduced by cytoduction into pet- strains that have been made rho0; cytoductants therefrom recover respiratory competency if the original pet- mutation is required only for mitochondrial splicing. By this means, we have identified 11 complementation groups of such genes. Their total number may be estimated as about 18. Images PMID:3309947

  13. Mitochondrial gene order change in Schistosoma (Platyhelminthes: Digenea: Schistosomatidae).

    PubMed

    Webster, Bonnie L; Littlewood, D Timothy J

    2012-01-01

    In the flatworm genus Schistosoma, species of which include parasites of biomedical and veterinary importance, mitochondrial gene order is radically different in some species. A PCR-based survey of 19 schistosomatid spp. established which of 14 Schistosoma spp. have the ancestral (plesiomorphic) or derived gene order condition. A phylogeny for Schistosoma was estimated and used to infer the origin of the gene order change which is present in all members of a clade containing Schistosoma incognitum and members of the traditionally recognised Schistosoma indicum, Schistosoma mansoni and Schistosomahaematobium spp. groups. Schistosoma turkestanicum, with the plesiomorphic gene order state, is sister to this clade. Common interval analysis suggests change in gene order, from ancestral to derived, consisted of two sequential transposition events: (a) nad1_nad3 to nad3_nad1 and (b) [atp6,nad2]_[nad3,-nad1,cox1,rrnL,rrnS,cox2,nad6] to [nad3,nad1,cox1,rrnL,rrnS,cox2,nad6]_[atp6,nad2], where gene order offragments within square brackets remain unchanged. Gene order change is rare in parasitic flatworms and is a robust synapomorphy for schistosome spp. that exhibit it. The schistosomatid phylogeny casts some doubt on the origin of Schistosoma (Asian or African), highlights the propensity for species to hosts witch amongst mammalian (definitive) hosts, and indicates the likely importance of snail (intermediate)hosts in determining and defining patterns of schistosome radiation and continental invasion. Mitogenomic sampling of Schistosoma dattai and Schistosoma harinasutai to determine gene order, and within key species, especially S. turkestanicum and S. incognitum, to determine ancestral ranges, may help discover the geographic origins of gene order change in the genus. Samples of S. incognitum from India and Thailand suggest this taxon may include cryptic species. PMID:23362512

  14. Characterization of the Treponema denticola prtP gene encoding a prolyl-phenylalanine-specific protease (dentilisin).

    PubMed Central

    Ishihara, K; Miura, T; Kuramitsu, H K; Okuda, K

    1996-01-01

    A chymotrypsin-like protease from Treponema denticola ATCC 35405 was purified by chromatographic techniques. The purified enzyme consisted of three polypeptides (38, 43, and 72 kDa). The protease exhibited specificity for peptide bonds containing phenylalanine and proline at the P1 and P2 positions, respectively, and was classified as a serine protease on the basis of inhibition studies. Naturally occurring protease inhibitors such as alpha1-antitrypsin and alpha1-antichymotrypsin had no effect on enzymatic activity. The enzyme degraded fibronectin, alpha1-antitrypsin, and gelatin while weakly degrading the immunoglobulin G heavy chain and type IV collagen. N-terminal amino acid sequences were determined for the 43- and 72-kDa proteins. On the basis of these sequences, the genes coding for the 43- and 72-kDa proteins were isolated and sequenced. The open reading frame which codes for the 72-kDa protein was designated prtP. This gene consists of 2,169 bp and codes for a protein with an Mr of 77,471. The protein appeared to be composed of a signal peptide region followed by a prosequence and the mature protein domain. The deduced amino acid sequence exhibited similarity with that of the Bacillus subtilis serine protease subtilisin. The deduced properties of the sequence suggest that the 72-kDa protein is a chymotrypsin-like protease. However, the nature and function of the 43-kDa protein have not yet been determined. PMID:8945563

  15. Detection of gene-anchored amplification polymorphism (GAAP) in the vicinity of plant mitochondrial genes.

    PubMed

    Loridon, K; Saumitou-Laprade, P

    2002-05-01

    A simple, semi-automatable method was established for assessing polymorphism in plant mitochondrial genome. A set of 41 mitochondrial markers based on the published Arabidopsis thaliana sequence was developed in Brassicaceae using a gene-anchored amplification polymorphism (GAAP) strategy. PCR primers were selected based on conserved coding regions of mitochondrial genes and used to amplify the corresponding 5' and/or 3' non-coding flanking regions in order to maximise sequence variability between haplotypes. The variations in fragment size were analysed on a LiCor DNA sequencer, but the methodology is compatible with various sequencing systems using denaturing polyacrylamide gels. One advantage of the method is that GAAP products can be directly sequenced (without any cloning steps) through labelled M13 consensus sequences. Mitochondrial GAAP loci gave clear and simple patterns (one or two bands) that were easy to score and highly reproducible. Nearly all mitochondrial loci examined in A. thaliana were conserved within the Brassicaceae family, and half of the primers generated products when DNA from a distant species, Beta vulgaris (Chenopodiaceae), was used as template. The GAAP markers revealed low levels of polymorphism within species but exhibited a high level of polymorphism among genera and families. Our results showed some discrepancies with respect to the published mtDNA sequence of A. thaliana. PMID:12073035

  16. MAP-1 and IAP-1, two novel AAA proteases with catalytic sites on opposite membrane surfaces in mitochondrial inner membrane of Neurospora crassa.

    PubMed

    Klanner, C; Prokisch, H; Langer, T

    2001-09-01

    Eukaryotic AAA proteases form a conserved family of membrane-embedded ATP-dependent proteases but have been analyzed functionally only in the yeast Saccharomyces cerevisiae. Here, we have identified two novel members of this protein family in the filamentous fungus Neurospora crassa, which were termed MAP-1 and IAP-1. Both proteins are localized to the inner membrane of mitochondria. They are part of two similar-sized high molecular mass complexes, but expose their catalytic sites to opposite membrane surfaces, namely, the intermembrane and the matrix space. Disruption of iap-1 by repeat-induced point mutation caused a slow growth phenotype at high temperature and stabilization of a misfolded inner membrane protein against degradation. IAP-1 could partially substitute for functions of its yeast homolog Yme1, demonstrating functional conservation. However, respiratory growth at 37 degrees C was not restored. Our results identify two components of the quality control system of the mitochondrial inner membrane in N. crassa and suggest that AAA proteases with catalytic sites exposed to opposite membrane surfaces are present in mitochondria of all eukaryotic cells. PMID:11553723

  17. Identification of two new keratinolytic proteases from a Bacillus pumilus strain using protein analysis and gene sequencing.

    PubMed

    Fellahi, Soltana; Chibani, Abdelwaheb; Feuk-Lagerstedt, Elisabeth; Taherzadeh, Mohammad J

    2016-12-01

    The Bacillus strain (CCUG 66887) has a high capacity to excrete keratinase with the ability to degrade both alpha- and beta keratin. In this study we aimed to show the characteristics of the keratinolytic protease and to identify its gene by using liquid chromatography-electrospray ionization tandem mass spectrometry methods (nanoHPLC-ESI-MS/MS) followed by Mascot data base search. The results showed that the enzyme in fact consists of two different keratinases, both with a molecular mass of 38 kDa. Further, DNA sequencing generated the open reading frame (ORF) of one of the genes (Ker1), and de novo genome sequencing identified the ORF of the second gene (Ker2). The two keratinase genes contain 1153 base pairs each and have a gene similarity of 67 %. In addition, the Bacillus strain was classified as Bacillus pumilus and its genes were annotated in the GeneBank at NCBI (accession: CP011109.1). Amino acid sequences alignment with known B. pumilus proteases indicated that the two keratinases of B. pumilus strain C4 are subtilisin-like serine proteases belonging to the Protease S8 family. Taken together, these result suggest the two keratinases as promising candidates for enzymatic processing of keratinous wastes in waste refinery. PMID:27363997

  18. Expression and characterization of Drosophila signal peptide peptidase-like (sppL), a gene that encodes an intramembrane protease.

    PubMed

    Casso, David J; Liu, Songmei; Biehs, Brian; Kornberg, Thomas B

    2012-01-01

    Intramembrane proteases of the Signal Peptide Peptidase (SPP) family play important roles in developmental, metabolic and signaling pathways. Although vertebrates have one SPP and four SPP-like (SPPL) genes, we found that insect genomes encode one Spp and one SppL. Characterization of the Drosophila sppL gene revealed that the predicted SppL protein is a highly conserved structural homolog of the vertebrate SPPL3 proteases, with a predicted nine-transmembrane topology, an active site containing aspartyl residues within a transmembrane region, and a carboxy-terminal PAL domain. SppL protein localized to both the Golgi and ER. Whereas spp is an essential gene that is required during early larval stages and whereas spp loss-of-function reduced the unfolded protein response (UPR), sppL loss of function had no apparent phenotype. This was unexpected given that genetic knockdown phenotypes in other organisms suggested significant roles for Spp-related proteases. PMID:22439002

  19. Regulation of nuclear genes encoding mitochondrial proteins in Saccharomyces cerevisiae.

    PubMed Central

    Brown, T A; Evangelista, C; Trumpower, B L

    1995-01-01

    Selection for mutants which release glucose repression of the CYB2 gene was used to identify genes which regulate repression of mitochondrial biogenesis. We have identified two of these as the previously described GRR1/CAT80 and ROX3 genes. Mutations in these genes not only release glucose repression of CYB2 but also generally release respiration of the mutants from glucose repression. In addition, both mutants are partially defective in CYB2 expression when grown on nonfermentable carbon sources, indicating a positive regulatory role as well. ROX3 was cloned by complementation of a glucose-inducible flocculating phenotype of an amber mutant and has been mapped as a new leftmost marker on chromosome 2. The ROX3 mutant has only a modest defect in glucose repression of GAL1 but is substantially compromised in galactose induction of GAL1 expression. This mutant also has increased SUC2 expression on nonrepressing carbon sources. We have also characterized the regulation of CYB2 in strains carrying null mutation in two other glucose repression genes, HXK2 and SSN6, and show that HXK2 is a negative regulator of CYB2, whereas SSN6 appears to be a positive effector of CYB2 expression. PMID:7592476

  20. High Variability of Mitochondrial Gene Order among Fungi

    PubMed Central

    Aguileta, Gabriela; de Vienne, Damien M.; Ross, Oliver N.; Hood, Michael E.; Giraud, Tatiana; Petit, Elsa; Gabaldón, Toni

    2014-01-01

    From their origin as an early alpha proteobacterial endosymbiont to their current state as cellular organelles, large-scale genomic reorganization has taken place in the mitochondria of all main eukaryotic lineages. So far, most studies have focused on plant and animal mitochondrial (mt) genomes (mtDNA), but fungi provide new opportunities to study highly differentiated mtDNAs. Here, we analyzed 38 complete fungal mt genomes to investigate the evolution of mtDNA gene order among fungi. In particular, we looked for evidence of nonhomologous intrachromosomal recombination and investigated the dynamics of gene rearrangements. We investigated the effect that introns, intronic open reading frames (ORFs), and repeats may have on gene order. Additionally, we asked whether the distribution of transfer RNAs (tRNAs) evolves independently to that of mt protein-coding genes. We found that fungal mt genomes display remarkable variation between and within the major fungal phyla in terms of gene order, genome size, composition of intergenic regions, and presence of repeats, introns, and associated ORFs. Our results support previous evidence for the presence of mt recombination in all fungal phyla, a process conspicuously lacking in most Metazoa. Overall, the patterns of rearrangements may be explained by the combined influences of recombination (i.e., most likely nonhomologous and intrachromosomal), accumulated repeats, especially at intergenic regions, and to a lesser extent, mobile element dynamics. PMID:24504088

  1. Gene Duplication and Adaptive Evolution of Digestive Proteases in Drosophila arizonae Female Reproductive Tracts

    PubMed Central

    Kelleher, Erin S; Swanson, Willie J; Markow, Therese A

    2007-01-01

    It frequently has been postulated that intersexual coevolution between the male ejaculate and the female reproductive tract is a driving force in the rapid evolution of reproductive proteins. The dearth of research on female tracts, however, presents a major obstacle to empirical tests of this hypothesis. Here, we employ a comparative EST approach to identify 241 candidate female reproductive proteins in Drosophila arizonae, a repleta group species in which physiological ejaculate–female coevolution has been documented. Thirty-one of these proteins exhibit elevated amino acid substitution rates, making them candidates for molecular coevolution with the male ejaculate. Strikingly, we also discovered 12 unique digestive proteases whose expression is specific to the D. arizonae lower female reproductive tract. These enzymes belong to classes most commonly found in the gastrointestinal tracts of a diverse array of organisms. We show that these proteases are associated with recent, lineage-specific gene duplications in the Drosophila repleta species group, and exhibit strong signatures of positive selection. Observation of adaptive evolution in several female reproductive tract proteins indicates they are active players in the evolution of reproductive tract interactions. Additionally, pervasive gene duplication, adaptive evolution, and rapid acquisition of a novel digestive function by the female reproductive tract points to a novel coevolutionary mechanism of ejaculate–female interaction. PMID:17784792

  2. Identification and mapping of tRNA genes on the Helianthus annuus mitochondrial genome.

    PubMed

    Ceci, L R; Veronico, P; Gallerani, R

    1996-01-01

    The physical map for seventeen tRNA genes on the mitochondrial genome of the dicotyledonous plant Helianthus annuus has been established. Eleven are genuine mitochondrial genes, while the other six show a high degree of similarity with the chloroplast counterparts. The genes, with the exception of the genuine trnS(GCT) and of the chloroplast-like trnV and trnP, are expressed. The comparison of the organization of some tRNA genes in the H. annuus mitochondrial genome with that of similar genes detectable in other plants reveals that their association is common to several dicotyledons. PMID:8722570

  3. The complete mitochondrial genome of Pseudocellus pearsei (Chelicerata: Ricinulei) and a comparison of mitochondrial gene rearrangements in Arachnida

    PubMed Central

    Fahrein, Kathrin; Talarico, Giovanni; Braband, Anke; Podsiadlowski, Lars

    2007-01-01

    Background Mitochondrial genomes are widely utilized for phylogenetic and population genetic analyses among animals. In addition to sequence data the mitochondrial gene order and RNA secondary structure data are used in phylogenetic analyses. Arachnid phylogeny is still highly debated and there is a lack of sufficient sequence data for many taxa. Ricinulei (hooded tickspiders) are a morphologically distinct clade of arachnids with uncertain phylogenetic affinities. Results The first complete mitochondrial DNA genome of a member of the Ricinulei, Pseudocellus pearsei (Arachnida: Ricinulei) was sequenced using a PCR-based approach. The mitochondrial genome is a typical circular duplex DNA molecule with a size of 15,099 bp, showing the complete set of genes usually present in bilaterian mitochondrial genomes. Five tRNA genes (trnW, trnY, trnN, trnL(CUN), trnV) show different relative positions compared to other Chelicerata (e.g. Limulus polyphemus, Ixodes spp.). We propose that two events led to this derived gene order: (1) a tandem duplication followed by random deletion and (2) an independent translocation of trnN. Most of the inferred tRNA secondary structures show the common cloverleaf pattern except tRNA-Glu where the TψC-arm is missing. In phylogenetic analyses (maximum likelihood, maximum parsimony, Bayesian inference) using concatenated amino acid and nucleotide sequences of protein-coding genes the basal relationships of arachnid orders remain unresolved. Conclusion Phylogenetic analyses (ML, MP, BI) of arachnid mitochondrial genomes fail to resolve interordinal relationships of Arachnida and remain in a preliminary stage because there is still a lack of mitogenomic data from important taxa such as Opiliones and Pseudoscorpiones. Gene order varies considerably within Arachnida – only eight out of 23 species have retained the putative arthropod ground pattern. Some gene order changes are valuable characters in phylogenetic analysis of intraordinal

  4. Critical COPD respiratory illness is linked to increased transcriptomic activity of neutrophil proteases genes

    PubMed Central

    2012-01-01

    essential role of neutrophil proteases in COPD patients with critical respiratory illness. Measurement and modulation of the expression of these genes could present an option for clinical monitoring and treatment of severe COPD exacerbations. PMID:22852767

  5. Genetic Variants in Nuclear-Encoded Mitochondrial Genes Influence AIDS Progression

    PubMed Central

    Hendrickson, Sher L.; Lautenberger, James A.; Chinn, Leslie Wei; Malasky, Michael; Sezgin, Efe; Kingsley, Lawrence A.; Goedert, James J.; Kirk, Gregory D.; Gomperts, Edward D.; Buchbinder, Susan P.; Troyer, Jennifer L.; O'Brien, Stephen J.

    2010-01-01

    Background The human mitochondrial genome includes only 13 coding genes while nuclear-encoded genes account for 99% of proteins responsible for mitochondrial morphology, redox regulation, and energetics. Mitochondrial pathogenesis occurs in HIV patients and genetically, mitochondrial DNA haplogroups with presumed functional differences have been associated with differential AIDS progression. Methodology/Principal Findings Here we explore whether single nucleotide polymorphisms (SNPs) within 904 of the estimated 1,500 genes that specify nuclear-encoded mitochondrial proteins (NEMPs) influence AIDS progression among HIV-1 infected patients. We examined NEMPs for association with the rate of AIDS progression using genotypes generated by an Affymetrix 6.0 genotyping array of 1,455 European American patients from five US AIDS cohorts. Successfully genotyped SNPs gave 50% or better haplotype coverage for 679 of known NEMP genes. With a Bonferroni adjustment for the number of genes and tests examined, multiple SNPs within two NEMP genes showed significant association with AIDS progression: acyl-CoA synthetase medium-chain family member 4 (ACSM4) on chromosome 12 and peroxisomal D3,D2-enoyl-CoA isomerase (PECI) on chromosome 6. Conclusions Our previous studies on mitochondrial DNA showed that European haplogroups with presumed functional differences were associated with AIDS progression and HAART mediated adverse events. The modest influences of nuclear-encoded mitochondrial genes found in the current study add support to the idea that mitochondrial function plays a role in AIDS pathogenesis. PMID:20877624

  6. Physella acuta: atypical mitochondrial gene order among panpulmonates (Gastropoda)

    PubMed Central

    Nolan, Journey R.; Bergthorsson, Ulfar; Adema, Coen M.

    2014-01-01

    Mitochondrial (mt) sequences are frequently used for phylogenetic reconstruction and for identification of species of molluscs. This study expands the phylogenetic range of Hygrophila (Panpulmonata) for which such sequence data are available by characterizing the full mt genome of the invasive freshwater snail Physella acuta (Physidae). The mt genome sequences of two P. acuta isolates from Stubblefield Lake, New Mexico, USA, differed in length (14,490 vs 14,314 bp) and showed 11.49% sequence divergence, whereas ITS1 and ITS2 sequences from the nuclear genome differed by 1.75%. The mt gene order of P. acuta (cox1, P, nad6, nad5, nad1, D, F, cox2, Y, W, nad4L, C, Q, atp6, R, E, rrnS, M, T, cox3, I, nad2, K, V, rrnL, L1, A, cytb, G, H, L2, atp8, N, nad2, S1, S2, nad4) differs considerably from the relatively conserved gene order within Panpulmonata. Phylogenetic trees show that the 13 protein-encoding mt gene sequences (equivalent codons) of P. acuta group according to gastropod phylogeny, yet branch lengths and dN/dS ratios for P. acuta indicate elevated amino acid substitutions relative to other gastropods. This study indicates that mt sequences of P. acuta are phylogenetically informative despite a considerable intraspecific divergence and the atypical gene order in its mt genome. PMID:25368439

  7. Using in silico techniques: Isolation and characterization of an insect cuticle-degrading-protease gene from Beauveria bassiana.

    PubMed

    Khan, Sehroon; Nadir, Sadia; Wang, Xuewen; Khan, Afsar; Xu, Jianchu; Li, Meng; Tao, Lihong; Khan, Siraj; Karunarathna, Samantha C

    2016-08-01

    Cuticle-degrading-proteases (CDPs) secreted by Beauveria spp. are pivotal biocontrol substances, possessing commercial potential for developing bio-pesticides. Therefore, a thoughtful and contemplative understanding and assessment of the structural and functional features of these proteases would markedly assist the development of biogenic pesticides. Computational molecular biology is a new facile alternative approach to the tedious experimental molecular biology; therefore, by using bioinformatics tools, we isolated and characterized an insect CDP gene from Beauveria bassiana 70 s.l. genomic DNA. The CDP gene (1240 bp with GeneBank accession no. KT804651.1) consisted of three introns and four CDS exons, and shared 74-100% sequence identity to the reference CDP genes. Its phylogenetic tree results showed a unique evolution pattern, and the predicted amino acid peptide (PAAP) consisted of 344 amino acid residues with pI, molecular weight, instability index, grand average hydropathicity value and aliphatic index of 7.2, 35.4 kDa, 24.45, -0.149, and 76.63, respectively. The gene possessed 74-89% amino acid sequence similarity to the 12 reference strains. Three motifs (Peptidase_S8 subtilase family) were detected in the PAAP, and the computed 3D structure possessed 79.09% structural identity to alkaline serine proteases. The PAAP had four (three serine proteases and one Pyridoxal-dependent decarboxylase) conserved domains, a disulfide bridge, two calcium binding sites, MY domain, and three predicted active sites in the serine family domains. These results will set the groundwork for further exploitation of proteases and understanding the mechanism of disease caused by cuticle-degrading-serine-proteases from entomopathogenic fungi. PMID:27287496

  8. Cloning, characterization, expression analysis and inhibition studies of a novel gene encoding Bowman-Birk type protease inhibitor from rice bean

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper presents the first study describing the isolation, cloning and characterization of a full length gene encoding Bowman-Birk protease inhibitor (RbTI) from rice bean (Vigna umbellata). A full-length protease inhibitor gene with complete open reading frame of 327bp encoding 109 amino acids w...

  9. Regulation of Bacterial Gene Expression by Protease-Alleviated Spatial Sequestration (PASS).

    PubMed

    Pitner, Ragan A; Scarpelli, Andrew H; Leonard, Joshua N

    2015-09-18

    In natural microbial systems, conditional spatial sequestration of transcription factors enables cells to respond rapidly to changes in their environment or intracellular state by releasing presynthesized regulatory proteins. Although such a mechanism may be useful for engineering synthetic biology technologies ranging from cell-based biosensors to biosynthetic platforms, to date it remains unknown how or whether such conditional spatial sequestration may be engineered. In particular, based upon seemingly contradictory reports in the literature, it is not clear whether subcellular spatial localization of a transcription factor within the cytoplasm is sufficient to preclude regulation of cognate promoters on plasmid-borne or chromosomal loci. Here, we describe a modular, orthogonal platform for investigating and implementing this mechanism using protease-alleviated spatial sequestration (PASS). In this system, expression of an exogenous protease mediates the proteolytic release of engineered transcriptional regulators from the inner face of the Escherichia coli cytoplasmic membrane. We demonstrate that PASS mediates robust, conditional regulation of either transcriptional repression, via tetR, or transcriptional activation, by the λ phage CI protein. This work provides new insights into a biologically important facet of microbial gene expression and establishes a new strategy for engineering conditional transcriptional regulation for the microbial synthetic biology toolbox. PMID:25822588

  10. Cloning and expression analysis of cysteine protease gene (MwCP) in Agropyron mongolicum Keng.

    PubMed

    Ao, T G B Y; Lang, M L; Li, Y Q; Zhao, Y; Wang, L C; Yang, X J

    2016-01-01

    In this study, a cysteine protease gene (MwCP) from Agropyron mongolicum Keng was isolated using RACE. Sequence analysis indicated that MwCP was 1473 bp, and it contained a 1134-bp open reading frame, which encoded 377 amino acids with a 24-amino acid N-terminal signal peptide. The results indicated that the MwCP protein was a new member of the papain C1A family, and it was predicted to be an extracellular, secretory stable hydrophilic protein. The secondary structure of MwCP was mainly composed of α-helices and random coils, and the space structure primarily contained α-helices, β-sheets, and β-turns. Homology analyses showed the 98% homology between MwCP amino acids and a cysteine protease found in Triticum aestivum (GenBank accession No. AAW21813.1). Analysis of mRNA using semi-quantitative RT-PCR indicated that during a 48-h drought stress period, MwCP was expressed during the 4th hour, and the expression level peaked during the 6th hour before declining to the original level. The results revealed that MwCP was involved in drought-resistant physiological processes of A. mongolicum. Moreover, the MwCP expression levels were highest in leaves, intermediate in roots, and lowest in stems. PMID:26909915

  11. Protease inhibitor 15, a candidate gene for abdominal aortic internal elastic lamina ruptures in the rat

    PubMed Central

    Falak, Samreen; Schafer, Sebastian; Baud, Amelie; Hummel, Oliver; Schulz, Herbert; Gauguier, Dominique; Osborne-Pellegrin, Mary

    2014-01-01

    The inbred Brown Norway (BN) rat develops spontaneous ruptures of the internal elastic lamina (RIEL) of the abdominal aorta (AA) and iliac arteries. Prior studies with crosses of the BN/Orl RJ (susceptible) and LOU/M (resistant) showed the presence of a significant QTL on chromosome 5 and the production of congenic rats proved the involvement of this locus. In this study, we further dissected the above-mentioned QTL by creating a new panel of LOU.BN(chr5) congenic and subcongenic lines and reduced the locus to 5.2 Mb. Then we studied 1,002 heterogeneous stock (HS) rats, whose phenotyping revealed a low prevalence and high variability for RIEL. High-resolution mapping in the HS panel detected the major locus on chromosome 5 (log P > 35) and refined it to 1.4 Mb. Subsequently, RNA-seq analysis on AA of BN, congenics, and LOU revealed expression differences for only protease inhibitor 15 (Pi15) gene and a putative long intergenic noncoding RNA (lincRNA) within the linkage region. The high abundance of lincRNA with respect to reduced Pi15 expression, in conjunction with exertion of longitudinal strain, may be related to RIEL, indicating the potential importance of proteases in biological processes related to defective aortic internal elastic lamina structure. Similar mechanisms may be involved in aneurysm initiation in the human AA. PMID:24790086

  12. Identification and Transcriptional Control of the Genes Encoding the Caulobacter crescentus ClpXP Protease

    PubMed Central

    Østerås, Magne; Stotz, Agathe; Nuoffer, Stefanie Schmid; Jenal, Urs

    1999-01-01

    The region of the Caulobacter crescentus chromosome harboring the genes for the ClpXP protease was isolated and characterized. Comparison of the deduced amino acid sequences of the C. crescentus ClpP and ClpX proteins with those of their homologues from several gram-positive and gram-negative bacteria revealed stronger conservation for the ATPase regulatory subunit (ClpX) than for the peptidase subunit (ClpP). The C. crescentus clpX gene was shown by complementation analysis to be functional in Escherichia coli. However, clpX from E. coli was not able to substitute for the essential nature of the clpX gene in C. crescentus. The clpP and clpX genes are separated on the C. crescentus chromosome by an open reading frame pointing in the opposite direction from the clp genes, and transcription of clpP and clpX was found to be uncoupled. clpP is transcribed as a monocistronic unit with a promoter (PP1) located immediately upstream of the 5′ end of the gene and a terminator structure following its 3′ end. PP1 is under heat shock control and is induced upon entry of the cells into the stationary phase. At least three promoters for clpX (PX1, PX2, and PX3) were mapped in the clpP-clpX intergenic region. In contrast to PP1, the clpX promoters were found to be downregulated after heat shock but were also subject to growth phase control. In addition, the clpP and clpX promoters showed different activity patterns during the cell cycle. Together, these results demonstrate that the genes coding for the peptidase and the regulatory subunits of the ClpXP protease are under independent transcriptional control in C. crescentus. Determination of the numbers of ClpP and ClpX molecules per cell suggested that ClpX is the limiting component compared with ClpP. PMID:10322004

  13. Isolation of cDNA from Jacaratia mexicana encoding a mexicain-like cysteine protease gene.

    PubMed

    Ramos-Martínez, Erick M; Herrera-Ramírez, Alejandra C; Badillo-Corona, Jesús Agustín; Garibay-Orijel, Claudio; González-Rábade, Nuria; Oliver-Salvador, María Del Carmen

    2012-07-01

    Cysteine proteases (CPs) from the C1 family, which are similar to papain, can be found in animals and plants, as well as some viruses and prokaryotes. These enzymes have diverse physiological functions and are thus very attractive for science and industry. Jacaratia mexicana, a member of the Caricaceae plant family, contains several CPs, the principal being mexicain, found to favorably compete against papain for many industrial applications due to its high stability and specific activity. In this study, leaves of J. mexicana were used to isolate a CP-coding gene, similar to those that code for mexicain and chymomexicain. By using rapid amplification of cDNA ends (RACE) as well as oligonucleotide design from papain-like conserved amino acids (aa), a sequence of 1404 bp consisting of a 5' terminal untranslated region (UTR) of 153 bp, a 3' terminal UTR of 131 bp, with a polyadenylation (poly(A)) signal sequence and a poly(A) tail, and an open reading frame (ORF) of 1046 bp, was obtained by overlapping three partial sequences. Two full-length cDNA sequences that encode for mexicain-like proteases were cloned from mRNA (JmCP4 and JmCP5). JmCP4 is predicted to have an ORF of 1044 bp, which codifies for polypeptides that have a 26 aa signal peptide region, a 108 aa propeptide region and a mature enzyme of 214 aa. A 969 bp fragment (JmCP5) encodes for a partial sequence of a CP gene, without the signal peptide region but with a full-length propeptide region. The sequence analysis showed that this protease presented a high similarity to other plant CPs from J. mexicana, Vasconcellea cundinamarcensis, Vasconcellea stipulata, and Carica papaya, among others, mainly at the conserved catalytic site. Obtaining the sequence of this CP gene from J. mexicana provides an alternative for production in a standard system and could be an initial step towards the commercialization of this enzyme. PMID:22543019

  14. Repeated, recent and diverse transfers of a mitochondrial gene to the nucleus in flowering plants.

    PubMed

    Adams, K L; Daley, D O; Qiu, Y L; Whelan, J; Palmer, J D

    2000-11-16

    A central component of the endosymbiotic theory for the bacterial origin of the mitochondrion is that many of its genes were transferred to the nucleus. Most of this transfer occurred early in mitochondrial evolution; functional transfer of mitochondrial genes has ceased in animals. Although mitochondrial gene transfer continues to occur in plants, no comprehensive study of the frequency and timing of transfers during plant evolution has been conducted. Here we report frequent loss (26 times) and transfer to the nucleus of the mitochondrial gene rps10 among 277 diverse angiosperms. Characterization of nuclear rps10 genes from 16 out of 26 loss lineages implies that many independent, RNA-mediated rps10 transfers occurred during recent angiosperm evolution; each of the genes may represent a separate functional gene transfer. Thus, rps10 has been transferred to the nucleus at a surprisingly high rate during angiosperm evolution. The structures of several nuclear rps10 genes reveal diverse mechanisms by which transferred genes become activated, including parasitism of pre-existing nuclear genes for mitochondrial or cytoplasmic proteins, and activation without gain of a mitochondrial targeting sequence. PMID:11099041

  15. A novel mitochondrial ATP8 gene mutation in a patient with apical hypertrophic cardiomyopathy and neuropathy.

    PubMed

    Jonckheere, An I; Hogeveen, Marije; Nijtmans, Leo; van den Brand, Mariel; Janssen, Antoon; Diepstra, Heleen; van den Brandt, Frans; van den Heuvel, Bert; Hol, Frans; Hofste, Tom; Kapusta, Livia; Dillmann, U; Shamdeen, M; Smeitink, J; Smeitink, J; Rodenburg, Richard

    2009-01-01

    To identify the biochemical and molecular genetic defect in a 16-year-old patient presenting with apical hypertrophic cardiomyopathy and neuropathy suspected for a mitochondrial disorder.Measurement of the mitochondrial energy-generating system (MEGS) capacity in muscle and enzyme analysis in muscle and fibroblasts were performed. Relevant parts of the mitochondrial DNA were analysed by sequencing.A homoplasmic nonsense mutation m.8529G→A (p.Trp55X) was found in the mitochondrial ATP8 gene in the patient's fibroblasts and muscle tissue. Reduced complex V activity was measured in the patient's fibroblasts and muscle tissue, and was confirmed in cybrid clones containing patient-derived mitochondrial DNAWe describe the first pathogenic mutation in the mitochondrial ATP8 gene, resulting in an improper assembly and reduced activity of the complex V holoenzyme. PMID:21686774

  16. The spectrum of low molecular weight alpha-amylase/protease inhibitor genes expressed in the US bread wheat Butte 86

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The complement of genes encoding alpha-amylase/protease inhibitors expressed in Triticum aestivum cv. Butte 86 was characterized by transcript and proteomic analysis. Coding sequences for 18 distinct proteins were identified among a collection of expressed sequence tags (ESTs) from Butte 86 developi...

  17. Inventory of the Human Mitochondrial Gene Expression Machinery with Links to Disease

    PubMed Central

    Shutt, Timothy E.; Shadel, Gerald S.

    2010-01-01

    Mammalian mitochondrial DNA encodes thirty-seven essential genes required for ATP production via oxidative phosphorylation, instability or misregulation of which is associated with human diseases and aging. Other than the mtDNA-encoded RNA species (thirteen mRNAs, 12S and 16S rRNAs, and twenty-two tRNAs), the many remaining factors needed for mitochondrial gene expression (i.e. transcription, RNA processing/modification and translation), including a dedicated set of mitochondrial ribosomal proteins, are products of nuclear genes that are imported into the mitochondrial matrix. Herein, we inventory the human mitochondrial gene expression machinery, and while doing so highlight specific associations of these regulatory factors with human disease. Major new breakthroughs have been made recently in this burgeoning area that set the stage for exciting future studies on the key outstanding issue of how mitochondrial gene expression is regulated differentially in vivo. This should promote a greater understanding of why mtDNA mutations and dysfunction cause the complex and tissue-specific pathology characteristic of mitochondrial disease states and how mitochondrial dysfunction contributes to more common human pathology and aging. PMID:20544879

  18. Glucose repression of yeast mitochondrial transcription: kinetics of derepression and role of nuclear genes.

    PubMed Central

    Ulery, T L; Jang, S H; Jaehning, J A

    1994-01-01

    Yeast mitochondrial transcript and gene product abundance has been observed to increase upon release from glucose repression, but the mechanism of regulation of this process has not been determined. We report a kinetic analysis of this phenomenon, which demonstrates that the abundance of all classes of mitochondrial RNA changes slowly relative to changes observed for glucose-repressed nuclear genes. Several cell doublings are required to achieve the 2- to 20-fold-higher steady-state levels observed after a shift to a nonrepressing carbon source. Although we observed that in some yeast strains the mitochondrial DNA copy number also increases upon derepression, this does not seem to play the major role in increased RNA abundance. Instead we found that three- to sevenfold increases in RNA synthesis rates, measured by in vivo pulse-labelling experiments, do correlate with increased transcript abundance. We found that mutations in the SNF1 and REG1 genes, which are known to affect the expression of many nuclear genes subject to glucose repression, affect derepression of mitochondrial transcript abundance. These genes do not appear to regulate mitochondrial transcript levels via regulation of the nuclear genes RPO41 and MTF1, which encode the subunits of the mitochondrial RNA polymerase. We conclude that a nuclear gene-controlled factor(s) in addition to the two RNA polymerase subunits must be involved in glucose repression of mitochondrial transcript abundance. Images PMID:8289797

  19. Multiple losses and transfers to the nucleus of two mitochondrial succinate dehydrogenase genes during angiosperm evolution.

    PubMed Central

    Adams, K L; Rosenblueth, M; Qiu, Y L; Palmer, J D

    2001-01-01

    Unlike in animals, the functional transfer of mitochondrial genes to the nucleus is an ongoing process in plants. All but one of the previously reported transfers in angiosperms involve ribosomal protein genes. Here we report frequent transfer of two respiratory genes, sdh3 and sdh4 (encoding subunits 3 and 4 of succinate dehydrogenase), and we also show that these genes are present and expressed in the mitochondria of diverse angiosperms. Southern hybridization surveys reveal that sdh3 and sdh4 have been lost from the mitochondrion about 40 and 19 times, respectively, among the 280 angiosperm genera examined. Transferred, functional copies of sdh3 and sdh4 were characterized from the nucleus in four and three angiosperm families, respectively. The mitochondrial targeting presequences of two sdh3 genes are derived from preexisting genes for anciently transferred mitochondrial proteins. On the basis of the unique presequences of the nuclear genes and the recent mitochondrial gene losses, we infer that each of the seven nuclear sdh3 and sdh4 genes was derived from a separate transfer to the nucleus. These results strengthen the hypothesis that angiosperms are experiencing a recent evolutionary surge of mitochondrial gene transfer to the nucleus and reveal that this surge includes certain respiratory genes in addition to ribosomal protein genes. PMID:11454775

  20. Dietary fatty acids affect mitochondrial phospholipid compositions and mitochondrial gene expression of rainbow trout liver at different ages.

    PubMed

    Almaida-Pagán, P F; De Santis, C; Rubio-Mejía, O L; Tocher, D R

    2015-01-01

    Mitochondria are among the first responders to various stressors that challenge the homeostasis of cells and organisms. Mitochondrial decay is generally associated with impairment in the organelle bioenergetics function and increased oxidative stress, and it appears that deterioration of mitochondrial inner membrane phospholipids (PL), particularly cardiolipin (CL), and accumulation of mitochondrial DNA (mtDNA) mutations are among the main mechanisms involved in this process. In the present study, liver mitochondrial membrane PL compositions, lipid peroxidation, and mtDNA gene expression were analyzed in rainbow trout fed three diets with the same base formulation but with lipid supplied either by fish oil (FO), rapeseed oil (RO), or high DHA oil (DHA) during 6 weeks. Specifically, two feeding trials were performed using fish from the same population of two ages (1 and 3 years), and PL class compositions of liver mitochondria, fatty acid composition of individual PL classes, TBARS content, and mtDNA expression were determined. Dietary fatty acid composition strongly affected mitochondrial membrane composition from trout liver but observed changes did not fully reflect the diet, particularly when it contained high DHA. The changes were PL specific, CL being particularly resistant to changes in DHA. Some significant differences observed in expression of mtDNA with diet may suggest long-term dietary effects in mitochondrial gene expression which could affect electron transport chain function. All the changes were influenced by fish age, which could be related to the different growth rates observed between 1- and 3-year-old trout but that could also indicate age-related changes in the ability to maintain structural homeostasis of mitochondrial membranes. PMID:25398637

  1. Whole Cell Formaldehyde Cross-Linking Simplifies Purification of Mitochondrial Nucleoids and Associated Proteins Involved in Mitochondrial Gene Expression

    PubMed Central

    Rajala, Nina; Hensen, Fenna; Wessels, Hans J. C. T.; Ives, Daniel; Gloerich, Jolein; Spelbrink, Johannes N.

    2015-01-01

    Mitochondrial DNA/protein complexes (nucleoids) appear as discrete entities inside the mitochondrial network when observed by live-cell imaging and immunofluorescence. This somewhat trivial observation in recent years has spurred research towards isolation of these complexes and the identification of nucleoid-associated proteins. Here we show that whole cell formaldehyde crosslinking combined with affinity purification and tandem mass-spectrometry provides a simple and reproducible method to identify potential nucleoid associated proteins. The method avoids spurious mitochondrial isolation and subsequent multifarious nucleoid enrichment protocols and can be implemented to allow for label-free quantification (LFQ) by mass-spectrometry. Using expression of a Flag-tagged Twinkle helicase and appropriate controls we show that this method identifies many previously identified nucleoid associated proteins. Using LFQ to compare HEK293 cells with and without mtDNA, but both expressing Twinkle-FLAG, identifies many proteins that are reduced or absent in the absence of mtDNA. This set not only includes established mtDNA maintenance proteins but also many proteins involved in mitochondrial RNA metabolism and translation and therefore represents what can be considered an mtDNA gene expression proteome. Our data provides a very valuable resource for both basic mitochondrial researchers as well as clinical geneticists working to identify novel disease genes on the basis of exome sequence data. PMID:25695250

  2. Transcriptional activation of the human cytotoxic serine protease gene CSP-B in T lymphocytes.

    PubMed Central

    Hanson, R D; Ley, T J

    1990-01-01

    The cytotoxic serine protease B (CSP-B) gene is activated during cytotoxic T-lymphocyte maturation. In this report, we demonstrate that the PEER T-cell line (bearing gamma/delta T-cell receptors) accumulates CSP-B mRNA following exposure to 12-O-tetradecanoylphorbol-13-acetate (TPA) and N6-2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (bt2cAMP) because of transcriptional activation of the CSP-B gene. TPA and bt2cAMP act synergistically to induce CSP-B expression, since neither agent alone causes activation of CSP-B transcription or mRNA accumulation. Chromatin upstream from the CSP-B gene is resistant to DNase I digestion in untreated PEER cells, but becomes sensitive following TPA-bt2cAMP treatment. Upon activation of PEER cells, a DNase I-hypersensitive site forms upstream from the CSP-B gene within a region that is highly conserved in the mouse. Transient transfection of CSP-B promoter constructs identified two regulatory regions in the CSP-B 5'-flanking sequence, located at positions -609 to -202 and positions -202 to -80. The region from -615 to -63 is sufficient to activate a heterologous promoter in activated PEER cells, but activation is orientation specific, suggesting that this region behaves as an upstream promoter element rather than a classical enhancer. Consensus AP-1, AP-2, and cAMP response elements are found upstream from the CSP-B gene (as are several T-cell-specific consensus elements), but the roles of these elements in CSP-B gene activation have yet to be determined. Images PMID:2233710

  3. Evolution of the mitochondrial genome in snakes: Gene rearrangements and phylogenetic relationships

    PubMed Central

    Yan, Jie; Li, Hongdan; Zhou, Kaiya

    2008-01-01

    Background Snakes as a major reptile group display a variety of morphological characteristics pertaining to their diverse behaviours. Despite abundant analyses of morphological characters, molecular studies using mitochondrial and nuclear genes are limited. As a result, the phylogeny of snakes remains controversial. Previous studies on mitochondrial genomes of snakes have demonstrated duplication of the control region and translocation of trnL to be two notable features of the alethinophidian (all serpents except blindsnakes and threadsnakes) mtDNAs. Our purpose is to further investigate the gene organizations, evolution of the snake mitochondrial genome, and phylogenetic relationships among several major snake families. Results The mitochondrial genomes were sequenced for four taxa representing four different families, and each had a different gene arrangement. Comparative analyses with other snake mitochondrial genomes allowed us to summarize six types of mitochondrial gene arrangement in snakes. Phylogenetic reconstruction with commonly used methods of phylogenetic inference (BI, ML, MP, NJ) arrived at a similar topology, which was used to reconstruct the evolution of mitochondrial gene arrangements in snakes. Conclusion The phylogenetic relationships among the major families of snakes are in accordance with the mitochondrial genomes in terms of gene arrangements. The gene arrangement in Ramphotyphlops braminus mtDNA is inferred to be ancestral for snakes. After the divergence of the early Ramphotyphlops lineage, three types of rearrangements occurred. These changes involve translocations within the IQM tRNA gene cluster and the duplication of the CR. All phylogenetic methods support the placement of Enhydris plumbea outside of the (Colubridae + Elapidae) cluster, providing mitochondrial genomic evidence for the familial rank of Homalopsidae. PMID:19038056

  4. Potential efficacy of mitochondrial genes for animal DNA barcoding: a case study using eutherian mammals

    PubMed Central

    2011-01-01

    Background A well-informed choice of genetic locus is central to the efficacy of DNA barcoding. Current DNA barcoding in animals involves the use of the 5' half of the mitochondrial cytochrome oxidase 1 gene (CO1) to diagnose and delimit species. However, there is no compelling a priori reason for the exclusive focus on this region, and it has been shown that it performs poorly for certain animal groups. To explore alternative mitochondrial barcoding regions, we compared the efficacy of the universal CO1 barcoding region with the other mitochondrial protein-coding genes in eutherian mammals. Four criteria were used for this comparison: the number of recovered species, sequence variability within and between species, resolution to taxonomic levels above that of species, and the degree of mutational saturation. Results Based on 1,179 mitochondrial genomes of eutherians, we found that the universal CO1 barcoding region is a good representative of mitochondrial genes as a whole because the high species-recovery rate (> 90%) was similar to that of other mitochondrial genes, and there were no significant differences in intra- or interspecific variability among genes. However, an overlap between intra- and interspecific variability was still problematic for all mitochondrial genes. Our results also demonstrated that any choice of mitochondrial gene for DNA barcoding failed to offer significant resolution at higher taxonomic levels. Conclusions We suggest that the CO1 barcoding region, the universal DNA barcode, is preferred among the mitochondrial protein-coding genes as a molecular diagnostic at least for eutherian species identification. Nevertheless, DNA barcoding with this marker may still be problematic for certain eutherian taxa and our approach can be used to test potential barcoding loci for such groups. PMID:21276253

  5. Extensive mitochondrial gene rearrangement in a genus of plant parasitic nematodes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The nematodes Globodera pallida and G. rostochiensis are two of the only animals known to have multipartite mitochondrial genomes. In such genomes, mitochondrial genes are distributed on multiple circles. The entire sequence of a nematode (Radopholus similis) that belongs to the same superfamily (...

  6. The mitochondrial genome of the onychophoran Opisthopatus cinctipes (Peripatopsidae) reflects the ancestral mitochondrial gene arrangement of Panarthropoda and Ecdysozoa.

    PubMed

    Braband, Anke; Cameron, Stephen L; Podsiadlowski, Lars; Daniels, Savel R; Mayer, Georg

    2010-10-01

    The ancestral genome composition in Onychophora (velvet worms) is unknown since only a single species of Peripatidae has been studied thus far, which shows a highly derived gene order with numerous translocated genes. Due to this lack of information from Onychophora, it is difficult to infer the ancestral mitochondrial gene arrangement patterns for Panarthropoda and Ecdysozoa. Hence, we analyzed the complete mitochondrial genome of the onychophoran Opisthopatus cinctipes, a representative of Peripatopsidae. Our data show that O. cinctipes possesses a highly conserved gene order, similar to that found in various arthropods. By comparing our results to those from different outgroups, we reconstruct the ancestral gene arrangement in Panarthropoda and Ecdysozoa. Our phylogenetic analysis of protein-coding gene sequences from 60 protostome species (including outgroups) provides some support for the sister group relationship of Onychophora and Arthropoda, which was not recovered by using a single species of Peripatidae, Epiperipatus biolleyi, in a previous study. A comparison of the strand-specific bias between onychophorans, arthropods, and a priapulid suggests that the peripatid E. biolleyi is less suitable for phylogenetic analyses of Ecdysozoa using mitochondrial genomic data than the peripatopsid O. cinctipes. PMID:20493270

  7. Mutant alcohol dehydrogenase (ADH III) presequences that affect both in vitro mitochondrial import and in vitro processing by the matrix protease.

    PubMed Central

    Mooney, D T; Pilgrim, D B; Young, E T

    1990-01-01

    Point mutations in the presequence of the mitochondrial alcohol dehydrogerase isoenzyme (ADH III) have been shown to affect either the import of the precursor protein into yeast mitochondria in vivo or its processing within the organelle. In the present work, the behavior of these mutants during in vitro import into isolated mitochondria was investigated. All point mutants tested were imported with a slower initial rate than that of the wild-type precursor. This defect was corrected when the precursors were treated with urea prior to import. Once imported, the extent of processing to the mature form of mutant precursors varied greatly and correlated well with the defects observed in vivo. This result was not affected by prior urea treatment. When matrix extracts enriched for the processing protease were used, this defect was shown to be due to failure of the protease to efficiently recognize or cleave the presequence, rather than to a lack of access to the precursor. The rate of import of two ADH III precursors bearing internal deletions in the leader sequence was similar to those of the point mutants, whereas a deletion leading to the removal of the 15 amino-terminal amino acids was poorly imported. The mature amino terminus of wild-type ADH III was determined to be Gln-25. Mutant m01 (Ser-26 to Phe), which reduced the efficiency of cleavage in vitro by 80%, was cleaved at the correct site. Images PMID:2188098

  8. Cloning, expression, and sequencing of a protease gene (tpr) from Porphyromonas gingivalis W83 in Escherichia coli.

    PubMed Central

    Bourgeau, G; Lapointe, H; Péloquin, P; Mayrand, D

    1992-01-01

    Porphyromonas gingivalis is a highly proteolytic organism which metabolizes small peptides and amino acids. Indirect evidence suggests that the proteases produced by this microorganism constitute an important virulence factor. In this study, a gene bank of P. gingivalis W83 DNA was constructed by cloning 0.5- to 20-kb HindIII-cut DNA fragments into Escherichia coli DH5 alpha by using the plasmid vector pUC19. A clone expressing a protease from P. gingivalis was isolated on LB agar containing 1% skim milk. The clone contained a 3.0-kb insert that coded for a protease with an apparent molecular mass of 64 kDa. Sequencing part of the 3.0-kb DNA fragment revealed an open reading frame encoding a protein of 482 amino acids with a molecular mass of 62.5 kDa. Putative promoter and termination elements flanking the open reading frame were identified. The activity expressed in E. coli was extensively characterized by using various substrates and protease inhibitors, and the results suggest that it is possibly a thiol protease. Images PMID:1322368

  9. Identification and Partial Characterization of Extracellular Aspartic Protease Genes from Metschnikowia pulcherrima IWBT Y1123 and Candida apicola IWBT Y1384

    PubMed Central

    Reid, Vernita J.; Theron, Louwrens W.; du Toit, Maret

    2012-01-01

    The extracellular acid proteases of non-Saccharomyces wine yeasts may fulfill a number of roles in winemaking, which include increasing the available nitrogen sources for the growth of fermentative microbes, affecting the aroma profile of the wine, and potentially reducing protein haze formation. These proteases, however, remain poorly characterized, especially at genetic level. In this study, two extracellular aspartic protease-encoding genes were identified and sequenced, from two yeast species of enological origin: one gene from Metschnikowia pulcherrima IWBT Y1123, named MpAPr1, and the other gene from Candida apicola IWBT Y1384, named CaAPr1. In silico analysis of these two genes revealed a number of features peculiar to aspartic protease genes, and both the MpAPr1 and CaAPr1 putative proteins showed homology to proteases of yeast genera. Heterologous expression of MpAPr1 in Saccharomyces cerevisiae YHUM272 confirmed that it encodes an aspartic protease. MpAPr1 production, which was shown to be constitutive, and secretion were confirmed in the presence of bovine serum albumin (BSA), casein, and grape juice proteins. The MpAPr1 gene was found to be present in 12 other M. pulcherrima strains; however, plate assays revealed that the intensity of protease activity was strain dependent and unrelated to the gene sequence. PMID:22820332

  10. Mammalian mitochondrial ribosomal small subunit (MRPS) genes: A putative role in human disease.

    PubMed

    Gopisetty, Gopal; Thangarajan, Rajkumar

    2016-09-01

    Mitochondria are prominently understood as power houses producing ATP the primary energy currency of the cell. However, mitochondria are also known to play an important role in apoptosis and autophagy, and mitochondrial dysregulation can lead to pathological outcomes. Mitochondria are known to contain 1500 proteins of which only 13 are coded by mitochondrial DNA and the rest are coded by nuclear genes. Protein synthesis in mitochondria involves mitochondrial ribosomes which are 55-60S particles and are composed of small 28S and large 39S subunits. A feature of mammalian mitoribosome which differentiate it from bacterial ribosomes is the increased protein content. The human mitochondrial ribosomal protein (MRP) gene family comprises of 30 genes which code for mitochondrial ribosomal small subunit and 50 genes for the large subunit. The present review focuses on the mitochondrial ribosomal small subunit genes (MRPS), presents an overview of the literature and data gleaned from publicly available gene and protein expression databases. The survey revealed aberrations in MRPS gene expression patterns in varied human diseases indicating a putative role in their etiology. PMID:27170550

  11. Independent replication of mitochondrial genes supports the transcriptional program in developing fiber cells of cotton (Gossypium hirsutum L.).

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The mitochondrial genomes of flowering plants exist both as a "master circle" chromosome and as numerous subgenomic sublimons that are generated by intramolecular recombination. Differential stability or replication of these sublimons allows individual mitochondrial gene copy numbers to vary indepe...

  12. Use of protease sensitivity to probe the conformations of newly synthesised mutant forms of mitochondrial aspartate aminotransferase.

    PubMed

    Azzariti, A; Giannattasio, S; Doonan, S; Merafina, R S; Marra, E; Quagliariello, E

    1995-10-24

    Sensitivity to digestion with pronase has been used to show that the precursor form of mitochondrial aspartate aminotransferase, the form lacking the N-terminal presequence, that with a deletion of the first 9 residues and mutants of the mature enzyme in which residue Cys-166 is mutated to alanine or serine, all retain unfolded conformations after synthesis in a reticulocyte lysate. In the presence of lysed mitochondria the various forms of mitochondrial aspartate aminotransferase retained their susceptibilities to pronase in a way that mirrored the efficiencies with which they are imported into intact mitochondria. The results are interpreted as showing that the presequence of mitochondrial aspartate aminotransferase is not uniquely required for interaction with cytosolic factors required to maintain the newly synthesised protein in a form competent for interacting with, and being imported into, mitochondria. PMID:7488044

  13. Crystal structure of the caseinolytic protease gene regulator, a transcriptional activator in actinomycetes.

    PubMed

    Russo, Santina; Schweitzer, Jens-Eric; Polen, Tino; Bott, Michael; Pohl, Ehmke

    2009-02-20

    Human pathogens of the genera Corynebacterium and Mycobacterium possess the transcriptional activator ClgR (clp gene regulator) which in Corynebacterium glutamicum has been shown to regulate the expression of the ClpCP protease genes. ClgR specifically binds to pseudo-palindromic operator regions upstream of clpC and clpP1P2. Here, we present the first crystal structure of a ClgR protein from C. glutamicum. The structure was determined from two different crystal forms to resolutions of 1.75 and 2.05 A, respectively. ClgR folds into a five-helix bundle with a helix-turn-helix motif typical for DNA-binding proteins. Upon dimerization the two DNA-recognition helices are arranged opposite to each other at the protein surface in a distance of approximately 30 A, which suggests that they bind into two adjacent major grooves of B-DNA in an anti-parallel manner. A binding pocket is situated at a strategic position in the dimer interface and could possess a regulatory role altering the positions of the DNA-binding helices. PMID:19019826

  14. Gene identification and molecular characterization of solvent stable protease from a moderately haloalkaliphilic bacterium, Geomicrobium sp. EMB2.

    PubMed

    Karan, Ram; Singh, Raj Kumar Mohan; Kapoor, Sanjay; Khare, S K

    2011-02-01

    Cloning and characterization of the gene encoding a solvent-tolerant protease from the haloalkaliphilic bacterium Geomicrobium sp. EMB2 are described. Primers designed based on the N-terminal amino acid sequence of the purified EMB2 protease helped in the amplification of a 1,505-bp open reading frame that had a coding potential of a 42.7-kDa polypeptide. The deduced EMB2 protein contained a 35.4-kDa mature protein of 311 residues, with a high proportion of acidic amino acid residues. Phylogenetic analysis placed the EMB2 gene close to a known serine protease from Bacillus clausii KSM-K16. Primary sequence analysis indicated a hydrophobic inclination of the protein; and the 3D structure modeling elucidated a relatively higher percentage of small (glycine, alanine, and valine) and borderline (serine and threonine) hydrophobic residues on its surface. The structure analysis also highlighted enrichment of acidic residues at the cost of basic residues. The study indicated that solvent and salt stabilities in Geomicrobium sp. protease may be accorded to different structural features; that is, the presence of a number of small hydrophobic amino acid residues on the surface and a higher content of acidic amino acid residues, respectively. PMID:21364294

  15. The gene expression landscape of thermogenic skunk cabbage suggests critical roles for mitochondrial and vacuolar metabolic pathways in the regulation of thermogenesis.

    PubMed

    Ito-Inaba, Yasuko; Hida, Yamato; Matsumura, Hideo; Masuko, Hiromi; Yazu, Fumiko; Terauchi, Ryohei; Watanabe, Masao; Inaba, Takehito

    2012-03-01

    Floral thermogenesis has been described in several plant species. Because of the lack of comprehensive gene expression profiles in thermogenic plants, the molecular mechanisms by which floral thermogenesis is regulated remain to be established. We examined the gene expression landscape of skunk cabbage (Symplocarpus renifolius) during thermogenic and post-thermogenic stages and identified expressed sequence tags from different developmental stages of the inflorescences using super serial analysis of gene expression (SuperSAGE). In-depth analysis suggested that cellular respiration and mitochondrial functions are significantly enhanced during the thermogenic stage. In contrast, genes involved in stress responses and protein degradation were significantly up-regulated during post-thermogenic stages. Quantitative comparisons indicated that the expression levels of genes involved in cellular respiration were higher in thermogenic spadices than in Arabidopsis inflorescences. Thermogenesis-associated genes seemed to be expressed abundantly in the peripheral tissues of the spadix. Our results suggest that cellular respiration and mitochondrial metabolism play key roles in heat production during floral thermogenesis. On the other hand, vacuolar cysteine protease and other degradative enzymes seem to accelerate senescence and terminate thermogenesis in the post-thermogenic stage. PMID:21955303

  16. Compilation and classification of higher plant mitochondrial tRNA genes.

    PubMed Central

    Veronico, P; Gallerani, R; Ceci, L R

    1996-01-01

    This compilation reports the tRNA genes detected on higher plant mitochondrial genomes subdivided into the widely accepted categories of 'genuine' and 'chloroplast-like' genes. Moreover, it includes a list of pseudo or truncated genes divided in the same way. PMID:8710486

  17. Compilation and classification of higher plant mitochondrial tRNA genes.

    PubMed

    Veronico, P; Gallerani, R; Ceci, L R

    1996-06-15

    This compilation reports the tRNA genes detected on higher plant mitochondrial genomes subdivided into the widely accepted categories of 'genuine' and 'chloroplast-like' genes. Moreover, it includes a list of pseudo or truncated genes divided in the same way. PMID:8710486

  18. Expression profiling of Drosophila mitochondrial genes via deep mRNA sequencing

    PubMed Central

    Torres, Tatiana Teixeira; Dolezal, Marlies; Schlötterer, Christian; Ottenwälder, Birgit

    2009-01-01

    Mitochondria play an essential role in several cellular processes. Nevertheless, very little is known about patterns of gene expression of genes encoded by the mitochondrial DNA (mtDNA). In this study, we used next-generation sequencing (NGS) for transcription profiling of genes encoded in the mitochondrial genome of Drosophila melanogaster and D. pseudoobscura. The analysis of males and females in both species indicated that the expression pattern was conserved between the two species, but differed significantly between both sexes. Interestingly, mRNA levels were not only different among genes encoded by separate transcription units, but also showed significant differences among genes located in the same transcription unit. Hence, mRNA abundance of genes encoded by mtDNA seems to be heavily modulated by post-transcriptional regulation. Finally, we also identified several transcripts with a noncanonical structure, suggesting that processing of mitochondrial transcripts may be more complex than previously assumed. PMID:19843606

  19. Isolation of the human PC6 gene encoding the putative host protease for HIV-1 gp160 processing in CD4+ T lymphocytes.

    PubMed Central

    Miranda, L; Wolf, J; Pichuantes, S; Duke, R; Franzusoff, A

    1996-01-01

    Production of infectious HIV-1 virions is dependent on the processing of envelope glycoprotein gp160 by a host cell protease. The protease in human CD4+ T lymphocytes has not been unequivocally identified, yet members of the family of mammalian subtilisin-like protein convertases (SPCs), which are soluble or membrane-bound proteases of the secretory pathway, best fulfill the criteria. These proteases are required for proprotein maturation and cleave at paired basic amino acid motifs in numerous cellular and viral glycoprotein precursors, both in vivo and in vitro. To identify the gp160 processing protease, we have used reverse transcription-PCR and Northern blot analyses to ascertain the spectrum of SPC proteases in human CD4+ T cells. We have cloned novel members of the SPC family, known as the human PC6 genes. Two isoforms of the hPC6 protease are expressed in human T cells, hPC6A and the larger hPC6B. The patterns of SPC gene expression in human T cells has been compared with the furin-defective LoVo cell line, both of which are competent in the production of infectious HIV virions. This comparison led to the conclusion that the hPC6 gene products are the most likely candidates for the host cell protease responsible for HIV-1 gp160 processing in human CD4+ T cells. Images Fig. 1 Fig. 3 PMID:8755538

  20. Gene characterization of two digestive serine proteases in orange blossom wheat midge (Sitodiplosis mosellana)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Two full length cDNA sequences, encoding digestive serine proteases (designated as SmPROT-1 and SmPROT-2), were recovered from the midgut of the wheat midge, Sitodiplosis mosellana in an ongoing EST project. The deduced amino acid sequences shared homology with digestive serine proteases from insect...

  1. Tripartite mitochondrial genome of spinach: physical structure, mitochondrial gene mapping, and locations of transposed chloroplast DNA sequences.

    PubMed Central

    Stern, D B; Palmer, J D

    1986-01-01

    A complete physical map of the spinach mitochondrial genome has been established. The entire sequence content of 327 kilobase pairs (kb) is postulated to occur as a single circular molecule. Two directly repeated elements of approximately 6 kb, located on this "master chromosome", are proposed to participate in an intragenomic recombination event that reversibly generates two "subgenomic" circles of 93 kb and 234 kb. The positions of protein and ribosomal RNA-encoding genes, determined by heterologous filter hybridizations, are scattered throughout the genome, with duplicate 26S rRNA genes located partially or entirely within the 6 kb repeat elements. Filter hybridizations between spinach mitochondrial DNA and cloned segments of spinach chloroplast DNA reveal at least twelve dispersed regions of inter-organellar sequence homology. Images PMID:3016660

  2. Comparative characterization of the iga gene encoding IgA1 protease in Neisseria meningitidis, Neisseria gonorrhoeae and Haemophilus influenzae.

    PubMed

    Lomholt, H; Poulsen, K; Kilian, M

    1995-02-01

    Cloning and sequencing of the IgA1 protease gene (iga) from Neisseria meningitidis strain HF13 showed an overall structure equivalent to iga genes from Neisseria gonorrhoeae and Haemophilus influenzae, although no region corresponding to the gonococcal alpha-peptide was evident. An additional 18 N. meningitidis and 3 H. influenzae iga genes were amplified by the polymerase chain reaction technique and sequenced corresponding approximately to the N-terminal half of the mature enzyme. Comparative analyses of a total of 29 iga genes showed that pathogenic Neisseria have iga genes with a significantly lower degree of heterogeneity than H. influenzae iga genes. Recombinational events indicated by mosaic-like structures corresponding to those found among N. gonorrhoeae protease genes were detected among N. meningitidis iga genes. One region showed characteristic differences in sequence and length which correlated with each of the different cleavage specificities. Meningococci were extremely conserved in this region with no evidence of recombination between isolates of different cleavage specificities. Sequences further downstream showed no obvious relationship with enzyme cleavage type. This region consisted of conserved areas interspersed with highly variable areas. Amino acid sequence homologies in the variable regions of meningococci reflected the antigenic types defined by using polyclonal neutralizing antibodies. PMID:7783620

  3. Sessile snails, dynamic genomes: gene rearrangements within the mitochondrial genome of a family of caenogastropod molluscs

    PubMed Central

    2010-01-01

    Background Widespread sampling of vertebrates, which comprise the majority of published animal mitochondrial genomes, has led to the view that mitochondrial gene rearrangements are relatively rare, and that gene orders are typically stable across major taxonomic groups. In contrast, more limited sampling within the Phylum Mollusca has revealed an unusually high number of gene order arrangements. Here we provide evidence that the lability of the molluscan mitochondrial genome extends to the family level by describing extensive gene order changes that have occurred within the Vermetidae, a family of sessile marine gastropods that radiated from a basal caenogastropod stock during the Cenozoic Era. Results Major mitochondrial gene rearrangements have occurred within this family at a scale unexpected for such an evolutionarily young group and unprecedented for any caenogastropod examined to date. We determined the complete mitochondrial genomes of four species (Dendropoma maximum, D. gregarium, Eualetes tulipa, and Thylacodes squamigerus) and the partial mitochondrial genomes of two others (Vermetus erectus and Thylaeodus sp.). Each of the six vermetid gastropods assayed possessed a unique gene order. In addition to the typical mitochondrial genome complement of 37 genes, additional tRNA genes were evident in D. gregarium (trnK) and Thylacodes squamigerus (trnV, trnLUUR). Three pseudogenes and additional tRNAs found within the genome of Thylacodes squamigerus provide evidence of a past duplication event in this taxon. Likewise, high sequence similarities between isoaccepting leucine tRNAs in Thylacodes, Eualetes, and Thylaeodus suggest that tRNA remolding has been rife within this family. While vermetids exhibit gene arrangements diagnostic of this family, they also share arrangements with littorinimorph caenogastropods, with which they have been linked based on sperm morphology and primary sequence-based phylogenies. Conclusions We have uncovered major changes in gene

  4. N-acetylcysteine inhibits the upregulation of mitochondrial biogenesis genes in livers from rats fed ethanol chronically

    PubMed Central

    Caro, Andres A.; Bell, Matthew; Ejiofor, Shannon; Zurcher, Grant; Petersen, Dennis R.; Ronis, Martin J. J.

    2014-01-01

    Background Chronic ethanol administration to experimental animals induces hepatic oxidative stress and upregulates mitochondrial biogenesis. The mechanisms by which chronic ethanol upregulates mitochondrial biogenesis have not been fully explored. In this work, we hypothesized that oxidative stress is a factor that triggers mitochondrial biogenesis after chronic ethanol feeding. If our hypothesis is correct, co-administration of antioxidants should prevent upregulation of mitochondrial biogenesis genes. Methods Rats were fed an ethanol-containing diet intragastrically by total enteral nutrition for 150 days, in the absence or presence of the antioxidant N-acetylcysteine (NAC) at 1.7 g/kg/day; control rats were administered isocaloric diets where carbohydrates substituted for ethanol calories. Results Ethanol administration significantly increased hepatic oxidative stress, evidenced as decreased liver total glutathione and GSH/GSSG ratio. These effects were inhibited by co-administration of ethanol and NAC. Chronic ethanol increased the expression of mitochondrial biogenesis genes including peroxisome proliferator activated receptor gamma-coactivator-1 alpha and mitochondrial transcription factor A, and mitochondrial DNA; co-administration of ethanol and NAC prevented these effects. Chronic ethanol administration was associated with decreased mitochondrial mass, inactivation and depletion of mitochondrial complex I and complex IV, and increased hepatic mitochondrial oxidative damage, effects that were not prevented by NAC. Conclusions These results suggest that oxidative stress caused by chronic ethanol triggered the upregulation of mitochondrial biogenesis genes in rat liver, because an antioxidant such as NAC prevented both effects. Because NAC did not prevent liver mitochondrial oxidative damage, extra-mitochondrial effects of reactive oxygen species may regulate mitochondrial biogenesis. In spite of the induction of hepatic mitochondrial biogenesis genes by

  5. A novel aspartic acid protease gene from pineapple fruit (Ananas comosus): cloning, characterization and relation to postharvest chilling stress resistance.

    PubMed

    Raimbault, Astrid-Kim; Zuily-Fodil, Yasmine; Soler, Alain; Cruz de Carvalho, Maria H

    2013-11-15

    A full-length cDNA encoding a putative aspartic acid protease (AcAP1) was isolated for the first time from the flesh of pineapple (Ananas comosus) fruit. The deduced sequence of AcAP1 showed all the common features of a typical plant aspartic protease phytepsin precursor. Analysis of AcAP1 gene expression under postharvest chilling treatment in two pineapple varieties differing in their resistance to blackheart development revealed opposite trends. The resistant variety showed an up-regulation of AcAP1 precursor gene expression whereas the susceptible showed a down-regulation in response to postharvest chilling treatment. The same trend was observed regarding specific AP enzyme activity in both varieties. Taken together our results support the involvement of AcAP1 in postharvest chilling stress resistance in pineapple fruits. PMID:23838125

  6. Isolation and gene expression analysis of a papain-type cysteine protease in thermogenic skunk cabbage (Symplocarpus renifolius).

    PubMed

    Ito-Inaba, Yasuko; Masuko, Hiromi; Watanabe, Masao; Inaba, Takehito

    2012-01-01

    Skunk cabbage (Symplocarpus renifolius) spadices contain abundant transcripts for cysteine protease (CP). From thermogenic spadices, we isolated SrCPA, a highly expressed CP gene that encoded a papain-type CP. SrCPA is structurally similar to other plant CPs, including the senescence-associated CPs found in aroids. The expression of SrCPA increased during floral development, and was observed in all floral tissues except for the stamens. PMID:23047088

  7. Molecular cloning and functional analysis of duck ubiquitin-specific protease 18 (USP18) gene.

    PubMed

    Qian, Wei; Wei, Xiaoqin; Zhou, Hongbo; Jin, Meilin

    2016-09-01

    In mammals, ubiquitin-specific protease 18 (USP18) is an interferon (IFN)-inducible gene and is a negative regulator of Toll-like receptor-mediated nuclear factor kappa B (NF-κB) activation. The role of USP18 in ducks (duUSP18) remains poorly understood. In the present study, we cloned and characterized the full-length coding sequence of duUSP18 from duck embryo fibroblasts (DEFs). In healthy ducks, duUSP18 transcripts were broadly expressed in different tissues, with higher expression levels in the spleen, lung and kidney. Quantitative real-time PCR (qRT-PCR) analysis revealed that duUSP18 could be induced by treatment with Poly(I:C) or LPS. Overexpression of duUSP18 inhibited NF-κB and IFN-β expression. Furthermore, deletion mutant analysis revealed that the duUSP18 region between aa 75 and 304 was essential for inhibiting NF-κB. In addition, overexpression of duUSP18 also suppressed the secretion of NF-κB-dependent proinflammatory cytokines. Taken together, these results suggest that duUSP18 regulates duck innate immune responses. PMID:27133094

  8. PhAP protease from Pseudoalteromonas haloplanktis TAC125: Gene cloning, recombinant production in E. coli and enzyme characterization

    NASA Astrophysics Data System (ADS)

    de Pascale, D.; Giuliani, M.; De Santi, C.; Bergamasco, N.; Amoresano, A.; Carpentieri, A.; Parrilli, E.; Tutino, M. L.

    2010-08-01

    Cold-adapted proteases have been found to be the dominant activity throughout the cold marine environment, indicating their importance in bacterial acquisition of nitrogen-rich complex organic compounds. However, few extracellular proteases from marine organisms have been characterized so far, and the mechanisms that enable their activity in situ are still largely unknown. Aside from their ecological importance and use as model enzyme for structure/function investigations, cold-active proteolytic enzymes offer great potential for biotechnological applications. Our studies on cold adapted proteases were performed on exo-enzyme produced by the Antarctic marine bacterium Pseudoalteromonas haloplanktis TAC125. By applying a proteomic approach, we identified several proteolytic activities from its culture supernatant. PhAP protease was selected for further investigations. The encoding gene was cloned and the protein was recombinantly produced in E. coli cells. The homogeneous product was biochemically characterised and it turned out that the enzyme is a Zn-dependent aminopeptidase, with an activity dependence from assay temperature typical of psychrophilic enzymes.

  9. Mitochondrial Homeostasis Molecules: Regulation by a Trio of Recessive Parkinson's Disease Genes

    PubMed Central

    Han, Ji-Young; Kim, Ji-Soo

    2014-01-01

    Mitochondria are small organelles that produce the majority of cellular energy as ATP. Mitochondrial dysfunction has been implicated in the pathogenesis of Parkinson's disease (PD), and rare familial forms of PD provide valuable insight into the pathogenic mechanism underlying mitochondrial impairment, even though the majority of PD cases are sporadic. The regulation of mitochondria is crucial for the maintenance of energy-demanding neuronal functions in the brain. Mitochondrial biogenesis and mitophagic degradation are the major regulatory pathways that preserve optimal mitochondrial content, structure and function. In this mini-review, we provide an overview of the mitochondrial quality control mechanisms, emphasizing regulatory molecules in mitophagy and biogenesis that specifically interact with the protein products of three major recessive familial PD genes, PINK1, Parkin and DJ-1. PMID:25548534

  10. The mitochondrial genome of the stramenopile alga Chrysodidymus synuroideus. Complete sequence, gene content and genome organization

    PubMed Central

    Chesnick, Joby M.; Goff, Megan; Graham, James; Ocampo, Christopher; Lang, B. Franz; Seif, Elias; Burger, Gertraud

    2000-01-01

    This is the first report of a complete mitochondrial genome sequence from a photosynthetic member of the stramenopiles, the chrysophyte alga Chrysodidymus synuroideus. The circular-mapping mitochondrial DNA (mtDNA) of 34 119 bp contains 58 densely packed genes (all without introns) and five unique open reading frames (ORFs). Protein genes code for components of respiratory chain complexes, ATP synthase and the mitoribosome, as well as one product of unknown function, encoded in many other protist mtDNAs (YMF16). In addition to small and large subunit ribosomal RNAs, 23 tRNAs are mtDNA-encoded, permitting translation of all codons present in protein-coding genes except ACN (Thr) and CGN (Arg). The missing tRNAs are assumed to be imported from the cytosol. Comparison of the C.synuroideus mtDNA with that of other stramenopiles allowed us to draw conclusions about mitochondrial genome organization, expression and evolution. First, we provide evidence that mitochondrial ORFs code for highly derived, unrecognizable versions of ribosomal or respiratory genes otherwise ‘missing’ in a particular mtDNA. Secondly, the observed constraints in mitochondrial genome rearrangements suggest operon-based, co-ordinated expression of genes functioning in common biological processes. Finally, stramenopile mtDNAs reveal an unexpectedly low variability in genome size and gene complement, testifying to substantial differences in the tempo of mtDNA evolution between major eukaryotic lineages. PMID:10871400

  11. Global variability in gene expression and alternative splicing is modulated by mitochondrial content.

    PubMed

    Guantes, Raul; Rastrojo, Alberto; Neves, Ricardo; Lima, Ana; Aguado, Begoña; Iborra, Francisco J

    2015-05-01

    Noise in gene expression is a main determinant of phenotypic variability. Increasing experimental evidence suggests that genome-wide cellular constraints largely contribute to the heterogeneity observed in gene products. It is still unclear, however, which global factors affect gene expression noise and to what extent. Since eukaryotic gene expression is an energy demanding process, differences in the energy budget of each cell could determine gene expression differences. Here, we quantify the contribution of mitochondrial variability (a natural source of ATP variation) to global variability in gene expression. We find that changes in mitochondrial content can account for ∼50% of the variability observed in protein levels. This is the combined result of the effect of mitochondria dosage on transcription and translation apparatus content and activities. Moreover, we find that mitochondrial levels have a large impact on alternative splicing, thus modulating both the abundance and type of mRNAs. A simple mathematical model in which mitochondrial content simultaneously affects transcription rate and splicing site choice can explain the alternative splicing data. The results of this study show that mitochondrial content (and/or probably function) influences mRNA abundance, translation, and alternative splicing, which ultimately affects cellular phenotype. PMID:25800673

  12. Brief Report: High Frequency of Biochemical Markers for Mitochondrial Dysfunction in Autism: No Association with the Mitochondrial Aspartate/Glutamate Carrier "SLC25A12" Gene

    ERIC Educational Resources Information Center

    Correia, Catarina; Coutinho, Ana M.; Diogo, Luisa; Grazina, Manuela; Marques, Carla; Miguel, Teresa; Ataide, Assuncao; Almeida, Joana; Borges, Luis; Oliveira, Catarina; Oliveira, Guiomar; Vicente, Astrid M.

    2006-01-01

    In the present study we confirm the previously reported high frequency of biochemical markers of mitochondrial dysfunction, namely hyperlactacidemia and increased lactate/pyruvate ratio, in a significant fraction of 210 autistic patients. We further examine the involvement of the mitochondrial aspartate/glutamate carrier gene ("SLC25A12") in…

  13. Gene organization and characterization of the complete mitochondrial genome of Hainan black goat (Capra hircus).

    PubMed

    Hu, Jiangtao; Zhao, Wei; Niu, Lili; Wang, Linjie; Li, Li; Zhang, Hongping; Zhong, Tao

    2016-05-01

    The complete mitochondrial genome sequence of Hainan black goat was determined for the first time by the PCR-based method. The total length of the mitogenome was 16,641 bp, including 33.54% A, 26.04% C, 27.31% T, 13.11% G. The genome structure contained 22 tRNA genes, 2 rRNA genes, 13 protein-coding genes and 1 control region (D-loop region). These results have extended more detail information of mitochondrial genome, thus being useful for further study on the genetic divergence and phylogenetic resolution of global goats. PMID:25211090

  14. A nuclear genetic lesion affecting Saccharomyces cerevisiae mitochondrial translation is complemented by a homologous Bacillus gene.

    PubMed Central

    Kim, S I; Stange-Thomann, N; Martins, O; Hong, K W; Söll, D; Fox, T D

    1997-01-01

    A novel Bacillus gene was isolated and characterized. It encodes a homolog of Saccharomyces cerevisiae Pet112p, a protein that has no characterized relative and is dispensable for cell viability but required for mitochondrial translation. Expression of the Bacillus protein in yeast, modified to ensure mitochondrial targeting, partially complemented the phenotype of the pet112-1 mutation, demonstrating a high degree of evolutionary conservation for this as yet unidentified component of translation. PMID:9287027

  15. The complete mitochondrial genome sequence and gene organization of Tridentiger trigonocephalus (Gobiidae: Gobionellinae) with phylogenetic consideration.

    PubMed

    Wei, Hongqing; Ma, Hongyu; Ma, Chunyan; Zhang, Fengying; Wang, Wei; Chen, Wei; Ma, Lingbo

    2016-09-01

    The complete mitochondrial genome plays an important role in studies of genome-level characteristics and phylogenetic relationships. Here we determined the complete mitogenome sequence of Tridentiger trigonocephalus (Perciformes, Gobiidae), and discovered its phylogenetic relationship. This circular genome was 16 662 bp in length, and consisted of 37 typical genes, including 13 protein-coding genes, 22 tRNA genes, and two rRNA genes. The gene order of T. trigonocephalus mitochondrial genome was identical to those observed in most other vertebrates. Of 37 genes, 28 were encoded by heavy strand, while the others were encoded by light strand. The phylogenetic tree constructed by 13 concatenated protein-coding genes showed that T. trigonocephalus was closest to T. bifasciatus, and then to T. barbatus among the 20 species within suborder Gobioidei. This work should facilitate the studies on population genetic diversity, and molecular evolution in Gobioidei fishes. PMID:26370266

  16. Molecular cloning, sequencing analysis, and chromosomal localization of the human protease inhibitor 4 (Kallistatin) gene (P14)

    SciTech Connect

    Chai, K.X.; Chao, J.; Chao, L.; Ward, D.C.

    1994-09-15

    The gene encoding human protease inhibitor 4 (kallistatin; gene symbol PI4), a novel serine proteinase inhibitor (serpin), has been isolated and completely sequenced. The kallistatin gene is 9618 bp in length and contains five exons and four introns. The structure and organization of the kallistatin gene are similar to those of the genes encoding {alpha}{sub 1}-antichymotrypsin. The kallistatin gene is also similar to the genes encoding rat and mouse kallikrein-binding proteins. The first exon of the kallistatin gene is a noncoding 89-bp fragment, as determined by primer extension. The fifth exon, which contains 308 bp of noncoding sequence, encodes the reactive center of kallistatin. In the 5`-flanking region of the kallistatin gene, 1125 bp have been sequenced and a consensus promoter segment with potential transcription regulatory sites, including CAAT and TATA boxes, an AP-2 binding site, a GC-rich region, a cAMP response element, and an AP-1 binding site, has been identified within this region. The kallistatin gene was localized by in situ hybridization to human chromosome 14q31-132.1, close to the serpin genes encoding {alpha}{sub 1}-antichymotrypsin, protein C inhibitor, {alpha}{sub 1}-antitrypsin, and corticosteroid-binding globulin. In a genomic DNA Southern blot, kallistatin-related genes were identified in monkey, mouse, rat, bovine, dog, cat, and a ground mole. The patterns of hybridization revealed clues of human serpin evolution. 34 refs., 6 figs.

  17. Uniparental Inheritance of Mitochondrial Genes in Yeast: Dependence on Input Bias of Mitochondrial DNA and Preliminary Investigations of the Mechanism

    PubMed Central

    Birky, C. William; Demko, Catherine A.; Perlman, Philip S.; Strausberg, Robert

    1978-01-01

    In Saccharomyces cerevisiae, previous studies on the inheritance of mitochondrial genes controlling antibiotic resistance have shown that some crosses produce a substantial number of uniparental zygotes , which transmit to their diploid progeny mitochondrial alleles from only one parent. In this paper, we show that uniparental zygotes are formed especially when one parent (majority parent) contributes substantially more mitochondrial DNA molecules to the zygote than does the other (minority) parent. Cellular contents of mitochondrial DNA (mtDNA) are increased in these experiments by treatment with cycloheximide, alpha-factor, or the uvsρ5 nuclear mutation. In such a biased cross, some zygotes are uniparental for mitochondrial alleles from the majority parent, and the frequency of such zygotes increases with increasing bias. In two- and three-factor crosses, the cap1, ery1, and oli1 loci behave coordinately, rather than independently; minority markers tend to be transmitted or lost as a unit, suggesting that the uniparental mechanism acts on entire mtDNA molecules rather than on individual loci. This rules out the possibility that uniparental inheritance can be explained by the conversion of minority markers to the majority alleles during recombination. Exceptions to the coordinate behavior of different loci can be explained by marker rescue via recombination. Uniparental inheritance is largely independent of the position of buds on the zygote. We conclude that it is due to the failure of minority markers to replicate in some zygotes, possibly involving the rapid enzymatic destruction of such markers. We have considered two general classes of mechanisms: (1) random selection of molecules for replication, as for example by competition for replicating sites on a membrane; and (2) differential marking of mtDNA molecules in the two parents, possibly by modification enzymes, followed by a mechanism that "counts" molecules and replicates only the majority type. These

  18. The PEP4 gene encodes an aspartyl protease implicated in the posttranslational regulation of Saccharomyces cerevisiae vacuolar hydrolases.

    PubMed Central

    Woolford, C A; Daniels, L B; Park, F J; Jones, E W; Van Arsdell, J N; Innis, M A

    1986-01-01

    pep4 mutants of Saccharomyces cerevisiae accumulate inactive precursors of vacuolar hydrolases. The PEP4 gene was isolated from a genomic DNA library by complementation of the pep4-3 mutation. Deletion analysis localized the complementing activity to a 1.5-kilobase pair EcoRI-XhoI restriction enzyme fragment. This fragment was used to identify an 1,800-nucleotide mRNA capable of directing the synthesis of a 44,000-dalton polypeptide. Southern blot analysis of yeast genomic DNA showed that the PEP4 gene is unique; however, several related sequences exist in yeasts. Tetrad analysis and mitotic recombination experiments localized the PEP4 gene proximal to GAL4 on chromosome XVI. Analysis of the DNA sequence indicated that PEP4 encodes a polypeptide with extensive homology to the aspartyl protease family. A comparison of the PEP4 predicted amino acid sequence with the yeast protease A protein sequence revealed that the two genes are, in fact, identical (see also Ammerer et al., Mol. Cell. Biol. 6:2490-2499, 1986). Based on our observations, we propose a model whereby inactive precursor molecules produced from the PEP4 gene self-activate within the yeast vacuole and subsequently activate other vacuolar hydrolases. Images PMID:3537721

  19. Yeast PPR proteins, watchdogs of mitochondrial gene expression

    PubMed Central

    Herbert, Christopher J; Golik, Pawel; Bonnefoy, Nathalie

    2013-01-01

    PPR proteins are a family of ubiquitous RNA-binding factors, found in all the Eukaryotic lineages, and are particularly numerous in higher plants. According to recent bioinformatic analyses, yeast genomes encode from 10 (in S. pombe) to 15 (in S. cerevisiae) PPR proteins. All of these proteins are mitochondrial and very often interact with the mitochondrial membrane. Apart from the general factors, RNA polymerase and RNase P, most yeast PPR proteins are involved in the stability and/or translation of mitochondrially encoded RNAs. At present, some information concerning the target RNA(s) of most of these proteins is available, the next challenge will be to refine our understanding of the function of the proteins and to resolve the yeast PPR-RNA-binding code, which might differ significantly from the plant PPR code. PMID:24184848

  20. Gene clusters for ribosomal proteins in the mitochondrial genome of a liverwort, Marchantia polymorpha.

    PubMed Central

    Takemura, M; Oda, K; Yamato, K; Ohta, E; Nakamura, Y; Nozato, N; Akashi, K; Ohyama, K

    1992-01-01

    We detected 16 genes for ribosomal proteins in the complete sequence of the mitochondrial DNA from a liverwort, Marchantia polymorpha. The genes formed two major clusters, rps12-rps7 and rps10-rpl2-rps19-rps3-rpl16-rpl5- rps14-rps8- rpl6-rps13-rps11-rps1, very similar in organization to Escherichia coli ribosomal protein operons (str and S10-spc-alpha operons, respectively). In contrast, rps2 and rps4 genes were located separately in the liverwort mitochondrial genome (the latter was part of the alpha operon in E. coli). Furthermore, several ribosomal proteins encoded by the liverwort mitochondrial genome differed substantially in size from their counterparts in E. coli and liverwort chloroplast. PMID:1620617

  1. Gene organization and complete sequence of the mitochondrial genome of Linwu mallard.

    PubMed

    Tian, Ke-Xiong; Liu, Li-Li; Yu, Qi-Fang; He, Shao-Ping; He, Jian-Hua

    2016-01-01

    Linwu mallard is an excellent native breeds from Hunan province in China. This is the first study to determine the complete mitochondrial genome sequence of L. mallard using PCR-based amplification and Sanger sequencing. The characteristic of the entire mitochondrial genome was analyzed in detail, with the base composition of 29.19% A, 22.19% T, 32.83% C, 15.79% G in the L. mallard (16,605 bp in length). It contained 2 ribosomal RNA genes, 13 protein-coding genes, 22 transfer RNA genes and a major non-coding control region (D-loop region). The complete mitochondrial genome sequence of L. mallard will be useful for the phylogenetics of poultry, and be available as basic data for the genetics and breeding. PMID:24938102

  2. The mitochondrial genome of Anopheles quadrimaculatus species A: complete nucleotide sequence and gene organization.

    PubMed

    Mitchell, S E; Cockburn, A F; Seawright, J A

    1993-12-01

    The complete sequence (15,455 bp) of the mitochondrial DNA of the mosquito Anopheles quadrimaculatus species A is reported. This genome is compact and very A+T rich (77.4% A+T). It contains genes for 2 ribosomal RNAs (rRNAs), 22 transfer RNAs (tRNAs), and 13 subunits of the mitochondrial inner membrane respiratory complexes. The gene arrangement is the same as in Drosophila yakuba, except that the positions of two contiguous tRNAs are reversed and a third tRNA is transcribed from the complementary strand. Protein-coding genes, rRNAs, and most tRNAs were similar to D. yakuba. Two tRNAs had nonstandard secondary structures comparable with those of nematode mitochondrial tRNAs. The very small putative control region (625 bp) contains no sequence motifs similar to those used in vertebrates and other insects for initiation of transcription and replication. PMID:8112570

  3. Mitochondrial content is central to nuclear gene expression: Profound implications for human health.

    PubMed

    Muir, Rebecca; Diot, Alan; Poulton, Joanna

    2016-02-01

    We review a recent paper in Genome Research by Guantes et al. showing that nuclear gene expression is influenced by the bioenergetic status of the mitochondria. The amount of energy that mitochondria make available for gene expression varies considerably. It depends on: the energetic demands of the tissue; the mitochondrial DNA (mtDNA) mutant load; the number of mitochondria; stressors present in the cell. Hence, when failing mitochondria place the cell in energy crisis there are major effects on gene expression affecting the risk of degenerative diseases, cancer and ageing. In 2015 the UK parliament approved a change in the regulation of IVF techniques, allowing "Mitochondrial replacement therapy" to become a reproductive choice for women at risk of transmitting mitochondrial disease to their children. This is the first time that this technique will be available. Therefore understanding the interaction between mitochondria and the nucleus has never been more important. PMID:26725055

  4. Mitochondrial content is central to nuclear gene expression: Profound implications for human health

    PubMed Central

    Muir, Rebecca; Diot, Alan

    2016-01-01

    We review a recent paper in Genome Research by Guantes et al. showing that nuclear gene expression is influenced by the bioenergetic status of the mitochondria. The amount of energy that mitochondria make available for gene expression varies considerably. It depends on: the energetic demands of the tissue; the mitochondrial DNA (mtDNA) mutant load; the number of mitochondria; stressors present in the cell. Hence, when failing mitochondria place the cell in energy crisis there are major effects on gene expression affecting the risk of degenerative diseases, cancer and ageing. In 2015 the UK parliament approved a change in the regulation of IVF techniques, allowing “Mitochondrial replacement therapy” to become a reproductive choice for women at risk of transmitting mitochondrial disease to their children. This is the first time that this technique will be available. Therefore understanding the interaction between mitochondria and the nucleus has never been more important. PMID:26725055

  5. Complete sequence and gene organization of the mitochondrial genome of Asio flammeus (Strigiformes, strigidae).

    PubMed

    Zhang, Yanan; Song, Tao; Pan, Tao; Sun, Xiaonan; Sun, Zhonglou; Qian, Lifu; Zhang, Baowei

    2016-07-01

    The complete sequence of the mitochondrial genome was determined for Asio flammeus, which is distributed widely in geography. The length of the complete mitochondrial genome was 18,966 bp, containing 2 rRNA genes, 22 tRNA genes, 13 protein-coding genes (PCGs), and 1 non-coding region (D-loop). All the genes were distributed on the H-strand, except for the ND6 subunit gene and eight tRNA genes which were encoded on the L-strand. The D-loop of A. flammeus contained many tandem repeats of varying lengths and repeat numbers. The molecular-based phylogeny showed that our species acted as the sister group to A. capensis and the supported Asio was the monophyletic group. PMID:25980662

  6. Insertion near the mitochondrial tyrosine tRNA gene in patients with mitochondrial diseases

    SciTech Connect

    Goto, Y.; Nonaka, I.; Horai, S.

    1994-09-01

    The 3243 mutation commonly found in patients with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) has been occasionally detected in patients with chronic progressive external opthalmoplegia (CPEO). To elucidate the molecular mechanism underlying this phenomenon, an extensive mitochondrial (mt) DNA study was performed on such a patient (3243-CPEO). The newly discovered insertion was located in the noncoding region between cytrochrome c oxidase subunit 1 and tyrosine tRNA. The insertion was not found in 58 or 22 CPEO patients with or without mtDNA large-scale deletion but in another 3243-CPEO patient. In addition, the insertion was present in 1 of 116 normal Japanese, who had no 3243 mutation, and in 3 of 68 3243-MELAS patients. These results raise the possibility that the phenotypic expression of the 3243 mutation could be modulated or arranged by additional mtDNA mutations.

  7. Improved systematic tRNA gene annotation allows new insights into the evolution of mitochondrial tRNA structures and into the mechanisms of mitochondrial genome rearrangements

    PubMed Central

    Jühling, Frank; Pütz, Joern; Bernt, Matthias; Donath, Alexander; Middendorf, Martin; Florentz, Catherine; Stadler, Peter F.

    2012-01-01

    Transfer RNAs (tRNAs) are present in all types of cells as well as in organelles. tRNAs of animal mitochondria show a low level of primary sequence conservation and exhibit ‘bizarre’ secondary structures, lacking complete domains of the common cloverleaf. Such sequences are hard to detect and hence frequently missed in computational analyses and mitochondrial genome annotation. Here, we introduce an automatic annotation procedure for mitochondrial tRNA genes in Metazoa based on sequence and structural information in manually curated covariance models. The method, applied to re-annotate 1876 available metazoan mitochondrial RefSeq genomes, allows to distinguish between remaining functional genes and degrading ‘pseudogenes’, even at early stages of divergence. The subsequent analysis of a comprehensive set of mitochondrial tRNA genes gives new insights into the evolution of structures of mitochondrial tRNA sequences as well as into the mechanisms of genome rearrangements. We find frequent losses of tRNA genes concentrated in basal Metazoa, frequent independent losses of individual parts of tRNA genes, particularly in Arthropoda, and wide-spread conserved overlaps of tRNAs in opposite reading direction. Direct evidence for several recent Tandem Duplication-Random Loss events is gained, demonstrating that this mechanism has an impact on the appearance of new mitochondrial gene orders. PMID:22139921

  8. Secretory leukocyte protease inhibitor gene deletion alters bleomycin-induced lung injury, but not development of pulmonary fibrosis.

    PubMed

    Habgood, Anthony N; Tatler, Amanda L; Porte, Joanne; Wahl, Sharon M; Laurent, Geoffrey J; John, Alison E; Johnson, Simon R; Jenkins, Gisli

    2016-06-01

    Idiopathic pulmonary fibrosis is a progressive, fatal disease with limited treatment options. Protease-mediated transforming growth factor-β (TGF-β) activation has been proposed as a pathogenic mechanism of lung fibrosis. Protease activity in the lung is tightly regulated by protease inhibitors, particularly secretory leukocyte protease inhibitor (SLPI). The bleomycin model of lung fibrosis was used to determine the effect of increased protease activity in the lungs of Slpi(-/-) mice following injury. Slpi(-/-), and wild-type, mice received oropharyngeal administration of bleomycin (30 IU) and the development of pulmonary fibrosis was assessed. Pro and active forms of matrix metalloproteinase (MMP)-2 and MMP-9 were measured. Lung fibrosis was determined by collagen subtype-specific gene expression, hydroxyproline concentration, and histological assessment. Alveolar TGF-β activation was measured using bronchoalveolar lavage cell pSmad2 levels and global TGF-β activity was assessed by pSmad2 immunohistochemistry. The active-MMP-9 to pro-MMP-9 ratio was significantly increased in Slpi(-/-) animals compared with wild-type animals, demonstrating enhanced metalloproteinase activity. Wild-type animals showed an increase in TGF-β activation following bleomycin, with a progressive and sustained increase in collagen type I, alpha 1 (Col1α1), III, alpha 1(Col3α1), IV, alpha 1(Col4α1) mRNA expression, and a significant increase in total lung collagen 28 days post bleomycin. In contrast Slpi(-/-) mice showed no significant increase of alveolar TGF-β activity following bleomycin, above their already elevated levels, although global TGF-β activity did increase. Slpi(-/-) mice had impaired collagen gene expression but animals demonstrated minimal reduction in lung fibrosis compared with wild-type animals. These data suggest that enhanced proteolysis does not further enhance TGF-β activation, and inhibits sustained Col1α1, Col3α1, and Col4α1 gene expression

  9. Biased introgression of mitochondrial and nuclear genes: a comparison of diploid and haplodiploid systems.

    PubMed

    Patten, Manus M; Carioscia, Sara A; Linnen, Catherine R

    2015-10-01

    Hybridization between recently diverged species, even if infrequent, can lead to the introgression of genes from one species into another. The rates of mitochondrial and nuclear introgression often differ, with some taxa showing biases for mitochondrial introgression and others for nuclear introgression. Several hypotheses exist to explain such biases, including adaptive introgression, sex differences in dispersal rates, sex-specific prezygotic isolation and sex-specific fitness of hybrids (e.g. Haldane's rule). We derive a simple population genetic model that permits an analysis of sex-specific demographic and fitness parameters and measures the relative rates of mitochondrial and nuclear introgression between hybridizing pairs. We do this separately for diploid and haplodiploid species. For diploid taxa, we recover results consistent with previous hypotheses: an excess of one sex among the hybridizing migrants or sex-specific prezygotic isolation causes a bias for one type of marker or the other; when Haldane's rule is obeyed, we find a mitochondrial bias in XY systems and a nuclear bias in ZW systems. For haplodiploid taxa, the model reveals that owing to their unique transmission genetics, they are seemingly assured of strong mitochondrial biases in introgression rates, unlike diploid taxa, where the relative fitness of male and female hybrids can tip the bias in either direction. This heretofore overlooked aspect of hybridization in haplodiploids provides what is perhaps the most likely explanation for differential introgression of mitochondrial and nuclear markers and raises concerns about the use of mitochondrial DNA barcodes for species delimitation in these taxa. PMID:26173469

  10. Recent stable insertion of mitochondrial DNA into an Arabidopsis polyubiquitin gene by nonhomologous recombination.

    PubMed

    Sun, C W; Callis, J

    1993-01-01

    Sequence analysis of a newly identified polyubiquitin gene (UBQ13) from the Columbia ecotype of Arabidopsis thaliana revealed that the gene contained a 3.9-kb insertion in the coding region. All subclones of the 3.9-kb insert hybridized to isolated mitochondrial DNA. The insert was found to consist of at least two, possibly three, distinct DNA segments from the mitochondrial genome. A 590-bp region of the insert is nearly identical to the Arabidopsis mitochondrial nad1 gene. UBQ13 restriction fragments in total cellular DNA from ecotypes Ler, No-0, Be-0, WS, and RLD were identified and, with the exception of Be-0, their sizes were equivalent to that predicted from the corresponding ecotype Columbia UBQ13 restriction fragment without the mitochondrial insert. Isolation by polymerase chain reaction and sequence determination of UBQ13 sequences from the other ecotypes showed that all lacked the mitochondrial insert. All ecotypes examined, except Columbia, contain intact open reading frames in the region of the insert, including four ubiquitin codons which Columbia lacks. This indicates that the mitochondrial DNA in UBQ13 in ecotype Columbia is the result of an integration event that occurred after speciation of Arabidopsis rather than a deletion event that occurred in all ecotypes except Columbia. This stable movement of mitochondrial DNA to the nucleus is so recent that there are few nucleotide changes subsequent to the transfer event. This allows for precise analysis of the sequences involved and elucidation of the possible mechanism. The presence of intron sequences in the transferred nucleic acid indicates that DNA was the transfer intermediate. The lack of sequence identity between the integrating sequence and the target site, represented by the other Arabidopsis ecotypes, suggests that integration occurred via nonhomologus recombination. This nuclear/organellar gene transfer event is strikingly similar to the experimentally accessible process of nuclear

  11. The natural killer cell serine protease gene Lmet1 maps to mouse chromosome 10

    SciTech Connect

    Thia, K.Y.T.; Smyth, M.J.; Jenkins, N.A.; Gilbert, D.J.; Copeland, N.G.

    1995-01-01

    Cytotoxic lymphocytes play a key role in immune responses against viruses and tumors. Lymphocyte-mediated cytolysis by both cytotoxic T lymphocytes (CTL) and natural killer (NK) cells is often associated with the formation of membrane lesions on target cells caused by exocytosis of cytoplasmic granule serine proteases and a pore-forming protein, perforin. A variety of granzymes have been found to reside within the cytoplasmic granules of cytotoxic lymphocytes, but unlike perforin, isolated serine proteases are not intrinsically lytic. However, a role for serine proteases in cellular cytotoxicity has been supported by the ability of protease inhibitors to completely abrogate lymphocyte cytotoxicity, and the demonstration that serine proteases can initiate DNA fragmentation in target cells transfected or pretreated with a sublytic concentration of perforin. Granzymes cloned in human, mouse, and rat encode four granzyme activities and all are expressed in either T cells, their thymic precursors, and/or NK cells. In particular, a rat granzyme that cleaves after methionine residues, but not phenylalanine residues and its human equivalent, human Met-ase 1, are unique granzymes with restricted expression in CD3-NK cells. 24 refs., 2 figs.

  12. Deregulation of genes related to iron and mitochondrial metabolism in refractory anemia with ring sideroblasts.

    PubMed

    del Rey, Mónica; Benito, Rocío; Fontanillo, Celia; Campos-Laborie, Francisco J; Janusz, Kamila; Velasco-Hernández, Talía; Abáigar, María; Hernández, María; Cuello, Rebeca; Borrego, Daniel; Martín-Zanca, Dionisio; De Las Rivas, Javier; Mills, Ken I; Hernández-Rivas, Jesús M

    2015-01-01

    The presence of SF3B1 gene mutations is a hallmark of refractory anemia with ring sideroblasts (RARS). However, the mechanisms responsible for iron accumulation that characterize the Myelodysplastic Syndrome with ring sideroblasts (MDS-RS) are not completely understood. In order to gain insight in the molecular basis of MDS-RS, an integrative study of the expression and mutational status of genes related to iron and mitochondrial metabolism was carried out. A total of 231 low-risk MDS patients and 81 controls were studied. Gene expression analysis revealed that iron metabolism and mitochondrial function had the highest number of genes deregulated in RARS patients compared to controls and the refractory cytopenias with unilineage dysplasia (RCUD). Thus mitochondrial transporters SLC25 (SLC25A37 and SLC25A38) and ALAD genes were over-expressed in RARS. Moreover, significant differences were observed between patients with SF3B1 mutations and patients without the mutations. The deregulation of genes involved in iron and mitochondrial metabolism provides new insights in our knowledge of MDS-RS. New variants that could be involved in the pathogenesis of these diseases have been identified. PMID:25955609

  13. Deregulation of Genes Related to Iron and Mitochondrial Metabolism in Refractory Anemia with Ring Sideroblasts

    PubMed Central

    del Rey, Mónica; Benito, Rocío; Fontanillo, Celia; Campos-Laborie, Francisco J.; Janusz, Kamila; Velasco-Hernández, Talía; Abáigar, María; Hernández, María; Cuello, Rebeca; Borrego, Daniel; Martín-Zanca, Dionisio; De Las Rivas, Javier; Mills, Ken I.; Hernández-Rivas, Jesús M.

    2015-01-01

    The presence of SF3B1 gene mutations is a hallmark of refractory anemia with ring sideroblasts (RARS). However, the mechanisms responsible for iron accumulation that characterize the Myelodysplastic Syndrome with ring sideroblasts (MDS-RS) are not completely understood. In order to gain insight in the molecular basis of MDS-RS, an integrative study of the expression and mutational status of genes related to iron and mitochondrial metabolism was carried out. A total of 231 low-risk MDS patients and 81 controls were studied. Gene expression analysis revealed that iron metabolism and mitochondrial function had the highest number of genes deregulated in RARS patients compared to controls and the refractory cytopenias with unilineage dysplasia (RCUD). Thus mitochondrial transporters SLC25 (SLC25A37 and SLC25A38) and ALAD genes were over-expressed in RARS. Moreover, significant differences were observed between patients with SF3B1 mutations and patients without the mutations. The deregulation of genes involved in iron and mitochondrial metabolism provides new insights in our knowledge of MDS-RS. New variants that could be involved in the pathogenesis of these diseases have been identified. PMID:25955609

  14. Effects of dietary soybean stachyose and phytic acid on gene expressions of serine proteases in Japanese flounder ( Paralichthys olivaceus)

    NASA Astrophysics Data System (ADS)

    Mi, Haifeng; Mai, Kangsen; Zhang, Wenbing; Wu, Chenglong; Cai, Yinghua

    2011-09-01

    Soybean stachyose (SBS) and phytic acid (PA) are anti-nutritional factors (ANF) which have deleterious effects on the growth and digestibility in fish. The present research studied the effects of dietary SBS and PA on the expression of three serine protease genes in the liver of Japanese flounder ( Paralichthys olivaceus). These genes are trypsinogen 1 (poTRY), elastase 1 (poEL) and chymotrypsinogen 1 (poCTRY). Eight artificial diets with graded levels of supplemented ANFs were formulated to 4 levels of SBS (0.00, 0.40, 0.80 and 1.50%), 4 levels of PA (0.00, 0.20, 0.40 and 0.80), respectively. Japanese flounder (initial weight 2.45 g ± 0.01 g) were fed with these diets for 10 weeks with three replications per treatment. At the end of 10 weeks, supplementation of 0.40% of dietary SBS or PA significantly increased the gene expression of poTRY and poCTRY ( P<0.05). The same level of dietary SBS significantly decreased the gene expression of poEL. In comparison with the control group (ANF-free), dietary PA (0.2% and 0.8%) significantly decreased the gene expression of poTRY, poCTRY and poEL ( P<0.05). However, excessive supplement of dietary SBS (1.5%) has no significant effects on these gene expressions ( P>0.05). These results suggested that dietary SBS and dietary PA could directly affect the serine protease genes at the transcriptional level in Japanese flounder, and these genes' expression was more sensitive to dietary PA than to SBS under the current experimental conditions.

  15. Mitochondrial Gene Expression Is Responsive to Starvation Stress and Developmental Transition in Trypanosoma cruzi

    PubMed Central

    Shaw, Aubie K.; Kalem, Murat C.

    2016-01-01

    ABSTRACT Trypanosoma cruzi parasites causing Chagas disease are passed between mammals by the triatomine bug vector. Within the insect, T. cruzi epimastigote-stage cells replicate and progress through the increasingly nutrient-restricted digestive tract, differentiating into infectious, nonreplicative metacyclic trypomastigotes. Thus, we evaluated how nutrient perturbations or metacyclogenesis affects mitochondrial gene expression in different insect life cycle stages. We compared mitochondrial RNA abundances in cultures containing fed, replicating epimastigotes, differentiating cultures containing both starved epimastigotes and metacyclic trypomastigotes and epimastigote starvation cultures. We observed increases in mitochondrial rRNAs and some mRNAs in differentiating cultures. These increases predominated only for the edited CYb mRNA in cultures enriched for metacyclic trypomastigotes. For the other transcripts, abundance increases were linked to starvation and were strongest in culture fractions with a high population of starved epimastigotes. We show that loss of both glucose and amino acids results in rapid increases in RNA abundances that are quickly reduced when these nutrients are returned. Furthermore, the individual RNAs exhibit distinct temporal abundance patterns, suggestive of multiple mechanisms regulating individual transcript abundance. Finally, increases in mitochondrial respiratory complex subunit mRNA abundances were not matched by increases in abundances of nucleus-encoded subunit mRNAs, nor were there statistically significant increases in protein levels of three nucleus-encoded subunits tested. These results show that, similarly to that in T. brucei, the mitochondrial genome in T. cruzi has the potential to alter gene expression in response to environmental or developmental stimuli but for an as-yet-unknown purpose. IMPORTANCE Chagas disease is caused by insect-transmitted Trypanosoma cruzi. Halting T. cruzi’s life cycle in one of its

  16. Mitochondrial neurogastrointestinal encephalomyopathy: novel pathogenic mutations in thymidine phosphorylase gene in two Italian brothers.

    PubMed

    Libernini, Laura; Lupis, Chiara; Mastrangelo, Mario; Carrozzo, Rosalba; Santorelli, Filippo Maria; Inghilleri, Maurizio; Leuzzi, Vincenzo

    2012-08-01

    Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE, MIM 603041) is an autosomal recessive multisystem disorder occurring due to mutations in a nuclear gene coding for the enzyme thymidine phosphorylase (TYMP). Clinical features of MNGIE include gastrointestinal dysmotility, cachexia, ptosis or ophthalmoparesis, peripheral neuropathy, diffuse leukoencephalopathy, and signs of mitochondrial dysfunction in tissues. We report the clinical and molecular findings in two brothers in whom novel TYMP gene mutations (c.215-13_215delinsGCGTGA; c.1159 + 2T > A) were associated with different clinical presentations and outcomes. PMID:22618301

  17. A new point mutation in the ND1 mitochondrial gene identified in a type II diabetic patient

    SciTech Connect

    Kalinin, V.N.; Schmidt, W.; Olek, K.

    1995-08-01

    A novel mutation in a mitochondrial gene was identified in a patient with type II diabetes mellitus. G-to-A transition was localized at the nt3316 position of gene ND1 and resulted in alanine threonine replacement at position 4 of mitochondrial NAD-H-dehydrogenase. 6 refs., 2 figs.

  18. The plant mitochondrial mat-r gene/nad1 gene complex

    SciTech Connect

    Wolstenhome, D.R.

    1996-12-31

    We have completed sequencing segments of the maize mitochondrial (mt) DNA that contains all five of the exons (A-E) of the gene (nad1) for subunit I of the respiratory chain NADH dehydrogenase. Analysis of these sequences indicates that exons B and C are joined by a continuous group II intron, but the remaining exons are associated with partial group II introns and are encoded at widely separated locations in the maize mtDNA molecule. We have shown that mature transcripts of the maize nad1 gene contain 23 edited nucleotides, and that transcripts of maize and soybean mat-r genes contain 15 and 14 edits, respectively. The majority of edits in nad1 transcripts result in amino acid replacements that increase similarity between the maize NAD1 protein and NAD1 proteins of other plant species and of animal species. We found that the intron between exons b and c is not edited. From data obtained using PCR and sequencing we have shown that transcripts containing all possible exon combinations exist in maize mitochondria.

  19. Palindromic Genes in the Linear Mitochondrial Genome of the Nonphotosynthetic Green Alga Polytomella magna

    PubMed Central

    Smith, David Roy; Hua, Jimeng; Archibald, John M.; Lee, Robert W.

    2013-01-01

    Organelle DNA is no stranger to palindromic repeats. But never has a mitochondrial or plastid genome been described in which every coding region is part of a distinct palindromic unit. While sequencing the mitochondrial DNA of the nonphotosynthetic green alga Polytomella magna, we uncovered precisely this type of genic arrangement. The P. magna mitochondrial genome is linear and made up entirely of palindromes, each containing 1–7 unique coding regions. Consequently, every gene in the genome is duplicated and in an inverted orientation relative to its partner. And when these palindromic genes are folded into putative stem-loops, their predicted translational start sites are often positioned in the apex of the loop. Gel electrophoresis results support the linear, 28-kb monomeric conformation of the P. magna mitochondrial genome. Analyses of other Polytomella taxa suggest that palindromic mitochondrial genes were present in the ancestor of the Polytomella lineage and lost or retained to various degrees in extant species. The possible origins and consequences of this bizarre genomic architecture are discussed. PMID:23940100

  20. Palindromic genes in the linear mitochondrial genome of the nonphotosynthetic green alga Polytomella magna.

    PubMed

    Smith, David Roy; Hua, Jimeng; Archibald, John M; Lee, Robert W

    2013-01-01

    Organelle DNA is no stranger to palindromic repeats. But never has a mitochondrial or plastid genome been described in which every coding region is part of a distinct palindromic unit. While sequencing the mitochondrial DNA of the nonphotosynthetic green alga Polytomella magna, we uncovered precisely this type of genic arrangement. The P. magna mitochondrial genome is linear and made up entirely of palindromes, each containing 1-7 unique coding regions. Consequently, every gene in the genome is duplicated and in an inverted orientation relative to its partner. And when these palindromic genes are folded into putative stem-loops, their predicted translational start sites are often positioned in the apex of the loop. Gel electrophoresis results support the linear, 28-kb monomeric conformation of the P. magna mitochondrial genome. Analyses of other Polytomella taxa suggest that palindromic mitochondrial genes were present in the ancestor of the Polytomella lineage and lost or retained to various degrees in extant species. The possible origins and consequences of this bizarre genomic architecture are discussed. PMID:23940100

  1. Profiling Gene Expression Induced by Protease-Activated Receptor 2 (PAR2) Activation in Human Kidney Cells

    PubMed Central

    Suen, Jacky Y.; Gardiner, Brooke; Grimmond, Sean; Fairlie, David P.

    2010-01-01

    Protease-Activated Receptor-2 (PAR2) has been implicated through genetic knockout mice with cytokine regulation and arthritis development. Many studies have associated PAR2 with inflammatory conditions (arthritis, airways inflammation, IBD) and key events in tumor progression (angiogenesis, metastasis), but they have relied heavily on the use of single agonists to identify physiological roles for PAR2. However such probes are now known not to be highly selective for PAR2, and thus precisely what PAR2 does and what mechanisms of downstream regulation are truly affected remain obscure. Effects of PAR2 activation on gene expression in Human Embryonic Kidney cells (HEK293), a commonly studied cell line in PAR2 research, were investigated here by comparing 19,000 human genes for intersecting up- or down-regulation by both trypsin (an endogenous protease that activates PAR2) and a PAR2 activating hexapeptide (2f-LIGRLO-NH2). Among 2,500 human genes regulated similarly by both agonists, there were clear associations between PAR2 activation and cellular metabolism (1,000 genes), the cell cycle, the MAPK pathway, HDAC and sirtuin enzymes, inflammatory cytokines, and anti-complement function. PAR-2 activation up-regulated four genes more than 5 fold (DUSP6, WWOX, AREG, SERPINB2) and down-regulated another six genes more than 3 fold (TXNIP, RARG, ITGB4, CTSD, MSC and TM4SF15). Both PAR2 and PAR1 activation resulted in up-regulated expression of several genes (CD44, FOSL1, TNFRSF12A, RAB3A, COPEB, CORO1C, THBS1, SDC4) known to be important in cancer. This is the first widespread profiling of specific activation of PAR2 and provides a valuable platform for better understanding key mechanistic roles of PAR2 in human physiology. Results clearly support the development of both antagonists and agonists of human PAR2 as potential disease modifying therapeutic agents. PMID:21072196

  2. Gene Arrangement Convergence, Diverse Intron Content, and Genetic Code Modifications in Mitochondrial Genomes of Sphaeropleales (Chlorophyta)

    PubMed Central

    Fučíková, Karolina; Lewis, Paul O.; González-Halphen, Diego; Lewis, Louise A.

    2014-01-01

    The majority of our knowledge about mitochondrial genomes of Viridiplantae comes from land plants, but much less is known about their green algal relatives. In the green algal order Sphaeropleales (Chlorophyta), only one representative mitochondrial genome is currently available—that of Acutodesmus obliquus. Our study adds nine completely sequenced and three partially sequenced mitochondrial genomes spanning the phylogenetic diversity of Sphaeropleales. We show not only a size range of 25–53 kb and variation in intron content (0–11) and gene order but also conservation of 13 core respiratory genes and fragmented ribosomal RNA genes. We also report an unusual case of gene arrangement convergence in Neochloris aquatica, where the two rns fragments were secondarily placed in close proximity. Finally, we report the unprecedented usage of UCG as stop codon in Pseudomuriella schumacherensis. In addition, phylogenetic analyses of the mitochondrial protein-coding genes yield a fully resolved, well-supported phylogeny, showing promise for addressing systematic challenges in green algae. PMID:25106621

  3. Species boundaries of Gulf of Mexico vestimentiferans (Polychaeta, Siboglinidae) inferred from mitochondrial genes

    NASA Astrophysics Data System (ADS)

    Pia Miglietta, Maria; Hourdez, Stephane; Cowart, Dominique A.; Schaeffer, Stephen W.; Fisher, Charles

    2010-11-01

    At least six morphospecies of vestimentiferan tubeworms are associated with cold seeps in the Gulf of Mexico (GOM). The physiology and ecology of the two best-studied species from depths above 1000 m in the upper Louisiana slope (Lamellibrachia luymesi and Seepiophila jonesi) are relatively well understood. The biology of one rare species from the upper slope (escarpiid sp. nov.) and three morphospecies found at greater depths in the GOM (Lamellibrachia sp. 1, L. sp. 2, and Escarpia laminata) are not as well understood. Here we address species distributions and boundaries of cold-seep tubeworms using phylogenetic hypotheses based on two mitochondrial genes. Fragments of the mitochondrial large ribosomal subunit rDNA (16S) and cytochrome oxidase subunit I (COI) genes were sequenced for 167 vestimentiferans collected from the GOM and analyzed in the context of other seep vestimentiferans for which sequence data were available. The analysis supported five monophyletic clades of vestimentiferans in the GOM. Intra-clade variation in both genes was very low, and there was no apparent correlation between the within-clade diversity and collection depth or location. Two of the morphospecies of Lamellibrachia from different depths in the GOM could not be distinguished by either mitochondrial gene. Similarly, E. laminata could not be distinguished from other described species of Escarpia from either the west coast of Africa or the eastern Pacific using COI. We suggest that the mitochondrial COI and 16S genes have little utility as barcoding markers for seep vestimentiferan tubeworms.

  4. Potential impact of human mitochondrial replacement on global policy regarding germline gene modification.

    PubMed

    Ishii, Tetsuya

    2014-08-01

    Previous discussions regarding human germline gene modification led to a global consensus that no germline should undergo genetic modification. However, the UK Human Fertilisation and Embryology Authority, having conducted at the UK Government's request a scientific review and a wide public consultation, provided advice to the Government on the pros and cons of Parliament's lifting a ban on altering mitochondrial DNA content of human oocytes and embryos, so as to permit the prevention of maternal transmission of mitochondrial diseases. In this commentary, relevant ethical and biomedical issues are examined and requirements for proceeding with this novel procedure are suggested. Additionally, potentially significant impacts of the UK legalization on global policy concerning germline gene modification are discussed in the context of recent advances in genome-editing technology. It is concluded that international harmonization is needed, as well as further ethical and practical consideration, prior to the legalization of human mitochondrial replacement. PMID:24832374

  5. Genome-wide identification, evolutuionary and expression analysis of aspartic proteases gene superfamily in grape

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspartic proteases (APs) are a large family of proteolytic enzymes in vertebrates, plants, yeast, nematodes, parasites, fungi, and viruses. In plants, they are involved in many biological processes, such as plant senescence, stress response, programmed cell death, and reproduction. Prior to the pr...

  6. New genes and pathomechanisms in mitochondrial disorders unraveled by NGS technologies.

    PubMed

    Legati, Andrea; Reyes, Aurelio; Nasca, Alessia; Invernizzi, Federica; Lamantea, Eleonora; Tiranti, Valeria; Garavaglia, Barbara; Lamperti, Costanza; Ardissone, Anna; Moroni, Isabella; Robinson, Alan; Ghezzi, Daniele; Zeviani, Massimo

    2016-08-01

    Next Generation Sequencing (NGS) technologies are revolutionizing the diagnostic screening for rare disease entities, including primary mitochondrial disorders, particularly those caused by nuclear gene defects. NGS approaches are able to identify the causative gene defects in small families and even single individuals, unsuitable for investigation by traditional linkage analysis. These technologies are contributing to fill the gap between mitochondrial disease cases defined on the basis of clinical, neuroimaging and biochemical readouts, which still outnumber by approximately 50% the cases for which a molecular-genetic diagnosis is attained. We have been using a combined, two-step strategy, based on targeted genes panel as a first NGS screening, followed by whole exome sequencing (WES) in still unsolved cases, to analyze a large cohort of subjects, that failed to show mutations in mtDNA and in ad hoc sets of specific nuclear genes, sequenced by the Sanger's method. Not only this approach has allowed us to reach molecular diagnosis in a significant fraction (20%) of these difficult cases, but it has also revealed unexpected and conceptually new findings. These include the possibility of marked variable penetrance of recessive mutations, the identification of large-scale DNA rearrangements, which explain spuriously heterozygous cases, and the association of mutations in known genes with unusual, previously unreported clinical phenotypes. Importantly, WES on selected cases has unraveled the presence of pathogenic mutations in genes encoding non-mitochondrial proteins (e.g. the transcription factor E4F1), an observation that further expands the intricate genetics of mitochondrial disease and suggests a new area of investigation in mitochondrial medicine. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi. PMID:26968897

  7. Host mitochondrial association evolved in the human parasite Toxoplasma gondii via neofunctionalization of a gene duplicate

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In Toxoplasma gondii, an intracellular parasite of humans and other warm-blooded animals, the ability to associate with host mitochondria (HMA) is driven by a locally expanded gene family that encodes multiple mitochondrial association factor 1 (MAF1) proteins. The importance of copy number in the e...

  8. Complete Sequence and Gene Organization of the Mitochondrial Genome of the Land Snail Albinaria Coerulea

    PubMed Central

    Hatzoglou, E.; Rodakis, G. C.; Lecanidou, R.

    1995-01-01

    The complete sequence (14,130 bp) of the mitochondrial DNA (mtDNA) of the land snail Albinaria coerulea was determined. It contains 13 protein, two rRNA and 22 tRNA genes. Twenty-four of these genes are encoded by one and 13 genes by the other strand. The gene arrangement shares almost no similarities with that of two other molluscs for which the complete gene content and arrangement are known, the bivalve Mytilus edulis and the chiton Katharina tunicata; the protein and rRNA gene order is similar to that of another terrestrial gastropod, Cepaea nemoralis. Unusual features include the following: (1) the absence of lengthy noncoding regions (there are only 141 intergenic nucleotides interspersed at different gene borders, the longest intergenic sequence being 42 nucleotides), (2) the presence of several overlapping genes (mostly tRNAs), (3) the presence of tRNA-like structures and other stem and loop structures within genes. An RNA editing system acting on tRNAs must necessarily be invoked for posttranscriptional extension of the overlapping tRNAs. Due to these features, and also because of the small size of its genes (e.g., it contains the smallest rRNA genes among the known coelomates), it is one of the most compact mitochondrial genomes known to date. PMID:7498775

  9. Molecular systematics of the genus Sigmodon: results from mitochondrial and nuclear gene sequences

    PubMed Central

    Henson, Dallas D.; Bradley, Robert D.

    2010-01-01

    Phylogenetic relationships within the genus Sigmodon Say and Ord, 1825 were examined using sequence data from multiple gene regions, including exon 1 of the nuclear-encoded interphotoreceptor retinoid binding protein, intron 7 of the nuclear beta-fibrinogen gene, and the mitochondrial cytochrome b gene from 27 individuals representing 11 species of Sigmodon. Nuclear genes were analyzed independently, combined with each other, and combined with the mitochondrial data. Topologies were constructed using parsimony and Bayesian methods, with nodal support provided by bootstrap and posterior probability values. All analyses recovered four independent clades (I–IV), each representing unique species groups: hispidus, fulviventer, peruanus, and alstoni. The analyses from the combined data also provided support for relationships previously proposed within those species groups. PMID:20407590

  10. Increased Incidence of Mitochondrial Cytochrome C Oxidase 1 Gene Mutations in Patients with Primary Ovarian Insufficiency

    PubMed Central

    Zhen, Xiumei; Wu, Bailin; Wang, Jian; Lu, Cuiling; Gao, Huafang; Qiao, Jie

    2015-01-01

    Primary ovarian insufficiency (POI), also known as premature ovarian failure (POF), is defined as more than six months of cessation of menses before the age of 40 years, with two serum follicle stimulating hormone (FSH) levels (at least 1 month apart) falling in the menopause range. The cause of POI remains undetermined in the majority of cases, although some studies have reported increased levels of reactive oxygen species (ROS) in idiopathic POF. The role of mitochondrial DNA in the pathogenesis of POI has not been studied extensively. This aim of this study was to uncover underlying mitochondrial genetic defects in patients with POI. The entire region of the mitochondrial genome was amplified in subjects with idiopathic POI (n=63) and age-matched healthy female controls (n=63) using nine pair sets of primers, followed by screening of the mitochondrial genome using an Illumina MiSeq. We identified a total of 96 non-synonymous mitochondrial variations in POI patients and 93 non-synonymous variations in control subjects. Of these, 21 (9 in POI and 12 in control) non-synonymous variations had not been reported previously. Eight mitochondrial cytochrome coxidase 1 (MT-CO1) missense variants were identified in POI patients, whereas only four missense mutations were observed in controls. A high incidence of MT-CO1 missense variants were identified in POI patients compared with controls, and the difference between the groups was statistically significant (13/63 vs. 5/63, p=0.042). Our results show that patients with primary ovarian insufficiency exhibit an increased incidence of mitochondrial cytochrome c oxidase 1 gene mutations, suggesting that MT-CO1 gene mutation may be causal in POI. PMID:26225554

  11. Mitochondrial impairment increases FL-PINK1 levels by calcium-dependent gene expression☆

    PubMed Central

    Gómez-Sánchez, Rubén; Gegg, Matthew E.; Bravo-San Pedro, José M.; Niso-Santano, Mireia; Alvarez-Erviti, Lydia; Pizarro-Estrella, Elisa; Gutiérrez-Martín, Yolanda; Alvarez-Barrientos, Alberto; Fuentes, José M.; González-Polo, Rosa Ana; Schapira, Anthony H.V.

    2014-01-01

    Mutations of the PTEN-induced kinase 1 (PINK1) gene are a cause of autosomal recessive Parkinson's disease (PD). This gene encodes a mitochondrial serine/threonine kinase, which is partly localized to mitochondria, and has been shown to play a role in protecting neuronal cells from oxidative stress and cell death, perhaps related to its role in mitochondrial dynamics and mitophagy. In this study, we report that increased mitochondrial PINK1 levels observed in human neuroblastoma SH-SY5Y cells after carbonyl cyanide m-chlorophelyhydrazone (CCCP) treatment were due to de novo protein synthesis, and not just increased stabilization of full length PINK1 (FL-PINK1). PINK1 mRNA levels were significantly increased by 4-fold after 24 h. FL-PINK1 protein levels at this time point were significantly higher than vehicle-treated, or cells treated with CCCP for 3 h, despite mitochondrial content being decreased by 29%. We have also shown that CCCP dissipated the mitochondrial membrane potential (Δψm) and induced entry of extracellular calcium through L/N-type calcium channels. The calcium chelating agent BAPTA-AM impaired the CCCP-induced PINK1 mRNA and protein expression. Furthermore, CCCP treatment activated the transcription factor c-Fos in a calcium-dependent manner. These data indicate that PINK1 expression is significantly increased upon CCCP-induced mitophagy in a calcium-dependent manner. This increase in expression continues after peak Parkin mitochondrial translocation, suggesting a role for PINK1 in mitophagy that is downstream of ubiquitination of mitochondrial substrates. This sensitivity to intracellular calcium levels supports the hypothesis that PINK1 may also play a role in cellular calcium homeostasis and neuroprotection. PMID:24184327

  12. Complete mitochondrial genome of Tubulipora flabellaris (Bryozoa: Stenolaemata): the first representative from the class Stenolaemata with unique gene order.

    PubMed

    Sun, Ming'an; Shen, Xin; Liu, Huilian; Liu, Xixing; Wu, Zhigang; Liu, Bin

    2011-09-01

    Mitochondrial genomes play a significant role in the reconstruction of phylogenetic relationships within metazoans. There are still many controversies concerning the phylogenetic position of the phylum Bryozoa. In this research, we have finished the complete mitochondrial genome of one bryozoan (Tubulipora flabellaris), which is the first representative from the class Stenolaemata. The complete mitochondrial genome of T. flabellaris is 13,763bp in length and contains 36 genes, which lacks the atp8 gene in contrast to the typical metazoan mitochondrial genomes. Gene arrangement comparisons indicate that the mitochondrial genome of T. flabellaris has unique gene order when compared with other metazoans. The four known bryozoans complete mitochondrial genomes also have very different gene arrangements, indicates that bryozoan mitochondrial genomes have experienced drastic rearrangements. To investigate the phylogenetic relationship of Bryozoa, phylogenetic analyses based on amino acid sequences of 11 protein coding genes (excluding atp6 and atp8) from 26 metazoan complete mitochondrial genomes were made utilizing Maximum Likelihood (ML) and Bayesian methods, respectively. The results indicate the monopoly of Lophotrochozoa and a close relationship between Chaetognatha and Bryozoa. However, more evidences are needed to clarify the relationship between two groups. Lophophorate appeared to be polyphyletic according to our analyses. Meanwhile, neither analysis supports close relationship between Branchiopod and Phoronida. Four bryozoans form a clade and the relationship among them is T. flabellaris+(F. hispida+(B. neritina+W. subtorquata)), which is in coincidence with traditional classification system. PMID:21867967

  13. FOXO3a regulates reactive oxygen metabolism by inhibiting mitochondrial gene expression.

    PubMed

    Ferber, E C; Peck, B; Delpuech, O; Bell, G P; East, P; Schulze, A

    2012-06-01

    Forkhead transcription factors of the O class (FOXOs) are important targets of the phosphatidylinositol 3-kinase/Akt pathway, and are key regulators of the cell cycle, apoptosis and response to oxidative stress. FOXOs have been shown to have tumour suppressor function and are important for stem cell maintenance. We have performed a detailed analysis of the transcriptional programme induced in response to Forkhead-box protein O3a (FOXO3a) activation. We observed that FOXO3a activation results in the repression of a large number of nuclear-encoded genes with mitochondrial function. Repression of these genes was mediated by FOXO3a-dependent inhibition of c-Myc. FOXO3a activation also caused a reduction in mitochondrial DNA copy number, expression of mitochondrial proteins, respiratory complexes and mitochondrial respiratory activity. FOXO3a has been previously implicated in the detoxification of reactive oxygen species (ROS) through induction of manganese-containing superoxide dismutase (SOD2). We observed that reduction in ROS levels following FOXO3a activation was independent of SOD2, but required c-Myc inhibition. Hypoxia increases ROS production from the mitochondria, which is required for stabilisation of the hypoxia-inducible factor-1α (HIF-1α). FOXO3a activation blocked the hypoxia-dependent increase in ROS and prevented HIF-1α stabilisation. Our data suggest that FOXO factors regulate mitochondrial activity through inhibition of c-Myc function and alter the hypoxia response. PMID:22139133

  14. FOXO3a regulates reactive oxygen metabolism by inhibiting mitochondrial gene expression

    PubMed Central

    Ferber, E C; Peck, B; Delpuech, O; Bell, G P; East, P; Schulze, A

    2012-01-01

    Forkhead transcription factors of the O class (FOXOs) are important targets of the phosphatidylinositol 3-kinase/Akt pathway, and are key regulators of the cell cycle, apoptosis and response to oxidative stress. FOXOs have been shown to have tumour suppressor function and are important for stem cell maintenance. We have performed a detailed analysis of the transcriptional programme induced in response to Forkhead-box protein O3a (FOXO3a) activation. We observed that FOXO3a activation results in the repression of a large number of nuclear-encoded genes with mitochondrial function. Repression of these genes was mediated by FOXO3a-dependent inhibition of c-Myc. FOXO3a activation also caused a reduction in mitochondrial DNA copy number, expression of mitochondrial proteins, respiratory complexes and mitochondrial respiratory activity. FOXO3a has been previously implicated in the detoxification of reactive oxygen species (ROS) through induction of manganese-containing superoxide dismutase (SOD2). We observed that reduction in ROS levels following FOXO3a activation was independent of SOD2, but required c-Myc inhibition. Hypoxia increases ROS production from the mitochondria, which is required for stabilisation of the hypoxia-inducible factor-1α (HIF-1α). FOXO3a activation blocked the hypoxia-dependent increase in ROS and prevented HIF-1α stabilisation. Our data suggest that FOXO factors regulate mitochondrial activity through inhibition of c-Myc function and alter the hypoxia response. PMID:22139133

  15. Thyroid hormone-regulated brain mitochondrial genes revealed by differential cDNA cloning.

    PubMed Central

    Vega-Núñez, E; Menéndez-Hurtado, A; Garesse, R; Santos, A; Perez-Castillo, A

    1995-01-01

    Thyroid hormone (T3) plays a critical role in the development of the central nervous system and its deficiency during the early neonatal period results in severe brain damage. However the mechanisms involved and the genes specifically regulated by T3 during brain development are largely unknown. By using a subtractive hybridization technique we have isolated a number of cDNAs that represented mitochondrial genes (12S and 16S rRNAs and cytochrome c oxidase subunit III). The steady state level of all three RNAs was reduced in hypothyroid animals during the postnatal period and T3 administration restored control levels. During fetal life the level of 16S rRNA was decreased in the brain of hypothyroid animals, suggesting a prenatal effect of thyroid hormone on brain development. Since T3 does not affect the amount of mitochondrial DNA, the results suggest that the effect of T3 is at transcriptional and/or postranscriptional level. In addition, the transcript levels for two nuclear-encoded mitochondrial cytochrome c oxidase subunits: subunits IV and VIc were also decreased in the brains of hypothyroid animals. Hypothyroidism-induced changes in mitochondrial RNAs were followed by a concomitant 40% decrease in cytochrome c oxidase activity. This study shows that T3 is an important regulator of mitochondrial function in the neonatal brain and, more importantly, provides a molecular basis for the specific action of this hormone in the developing brain. Images PMID:7635984

  16. Molecular cloning of a thermostable neutral protease gene from Bacillus stearothermophilus in a vector plasmid and its expression in Bacillus stearothermophilus and Bacillus subtilis.

    PubMed Central

    Fujii, M; Takagi, M; Imanaka, T; Aiba, S

    1983-01-01

    The structural gene for a thermostable protease from Bacillus stearothermophilus was cloned in plasmid pTB90. It is expressed in both B. stearothermophilus and Bacillus subtilis. B. stearothermophilus carrying the recombinant plasmid produced about 15-fold more protease (310 U/mg of cell dry weight) than did the wild-type strain of B. stearothermophilus. Some properties of the proteases that have been purified from the transformants of B. stearothermophilus and B. subtilis were examined. No significant difference was observed among the enzyme properties studied here despite the difference in host cells. We found that the protease, neutral in pH characteristics and with a molecular weight of 36,000, retained about 80% of its activity even after treatment of 65 degrees C for 30 min. Images PMID:6302083

  17. Identification, Characterization and Down-Regulation of Cysteine Protease Genes in Tobacco for Use in Recombinant Protein Production

    PubMed Central

    Duwadi, Kishor; Chen, Ling; Menassa, Rima; Dhaubhadel, Sangeeta

    2015-01-01

    Plants are an attractive host system for pharmaceutical protein production. Many therapeutic proteins have been produced and scaled up in plants at a low cost compared to the conventional microbial and animal-based systems. The main technical challenge during this process is to produce sufficient levels of recombinant proteins in plants. Low yield is generally caused by proteolytic degradation during expression and downstream processing of recombinant proteins. The yield of human therapeutic interleukin (IL)-10 produced in transgenic tobacco leaves was found to be below the critical level, and may be due to degradation by tobacco proteases. Here, we identified a total of 60 putative cysteine protease genes (CysP) in tobacco. Based on their predicted expression in leaf tissue, 10 candidate CysPs (CysP1-CysP10) were selected for further characterization. The effect of CysP gene silencing on IL-10 accumulation was examined in tobacco. It was found that the recombinant protein yield in tobacco could be increased by silencing CysP6. Transient expression of CysP6 silencing construct also showed an increase in IL-10 accumulation in comparison to the control. Moreover, CysP6 localizes to the endoplasmic reticulum (ER), suggesting that ER may be the site of IL-10 degradation. Overall results suggest that CysP6 is important in determining the yield of recombinant IL-10 in tobacco leaves. PMID:26148064

  18. Characterization of a juvenile hormone-regulated chymotrypsin-like serine protease gene in Aedes aegypti mosquito

    PubMed Central

    Bian, Guowu; Raikhel, Alexander S; Zhu, Jinsong

    2008-01-01

    After female mosquitoes ingest blood from vertebrate hosts, exopeptidases and endopeptidases are required for digesting blood proteins in the midgut into amino acids, which female mosquitoes use to build yolk proteins. These proteases are not always present in the midgut, and their diverse expression patterns suggest that production of these enzymes is highly regulated in order to meet specific physiological demands at various stages. Here we report identification of a serine-type protease, JHA15, in the yellow fever mosquito Aedes aegypti. This protein shares high sequence homology with chymotrypsins, and indeed exhibits specific chymotrypsin enzymatic activity. The JHA15 gene is expressed primarily in the midgut of adult female mosquitoes. Our results indicate that its transcription is activated by juvenile hormone in the newly emerged female adults. Although its mRNA profile is similar to that of the early trypsin gene, we found that JHA15 proteins were readily detected in the midgut epithelium cells of both non-blood-fed and blood-fed mosquitoes. Analysis of polysomal RNA further substantiated that synthesis of JHA15 occurs before and shortly after blood feeding. Knocking down expression of JHA15 resulted in no evident phenotypic changes, implying that functional redundancy exists among those proteolytic enzymes. PMID:18207080

  19. Effects of TCDD on the Expression of Nuclear Encoded Mitochondrial Genes

    PubMed Central

    Forgacs, Agnes L.; Burgoon, Lyle D.; Lynn, Scott G.; LaPres, John J.; Zacharewski, Timothy

    2014-01-01

    Generation of mitochondrial reactive oxygen species (ROS) can be perturbed following exposure to environmental chemicals such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Reports indicate that the aryl hydrocarbon receptor (AhR) mediates TCDD-induced sustained hepatic oxidative stress by decreasing hepatic ATP levels and through hyperpolarization of the inner mitochondrial membrane. To further elucidate the effects of TCDD on the mitochondria, high-throughput quantitative real-time PCR (HTP-QRTPCR) was used to evaluate the expression of 90 genes encoding mitochondrial proteins involved in electron transport, oxidative phosphorylation, uncoupling, and associated chaperones. HTP-QRTPCR analysis of time course (30 μg/kg TCDD at 2, 4, 8, 12, 18, 24, 72, and 168 hrs) liver samples obtained from orally gavaged immature, ovariectomized C57BL/6 mice identified 54 differentially expressed genes (|fold change|>1.5 and P-value <0.1). Of these, 8 exhibited a dose response (0.03 to 300 μg/kg TCDD) at 4, 24 or 72 hrs. Dose responsive genes encoded proteins associated with electron transport chain (ETC) complex I (NADH dehydrogenase), III (cytochrome c reductase), IV (cytochrome c oxidase), and V (ATP synthase) and could be generally categorized as having proton gradient, ATP synthesis, and chaperone activities. In contrast, transcript levels of ETC complex II, succinate dehydrogenase, remained unchanged. Putative dioxin response elements were computationally found in the promoter regions of the 8 dose-responsive genes. This high-throughput approach suggests that TCDD alters the expression of genes associated with mitochondrial function which may contribute to TCDD-elicited mitochondrial toxicity. PMID:20399798

  20. Effects of TCDD on the expression of nuclear encoded mitochondrial genes

    SciTech Connect

    Forgacs, Agnes L.; Burgoon, Lyle D.; Lynn, Scott G.; LaPres, John J.; Zacharewski, Timothy

    2010-07-15

    Generation of mitochondrial reactive oxygen species (ROS) can be perturbed following exposure to environmental chemicals such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Reports indicate that the aryl hydrocarbon receptor (AhR) mediates TCDD-induced sustained hepatic oxidative stress by decreasing hepatic ATP levels and through hyperpolarization of the inner mitochondrial membrane. To further elucidate the effects of TCDD on the mitochondria, high-throughput quantitative real-time PCR (HTP-QRTPCR) was used to evaluate the expression of 90 nuclear genes encoding mitochondrial proteins involved in electron transport, oxidative phosphorylation, uncoupling, and associated chaperones. HTP-QRTPCR analysis of time course (30 {mu}g/kg TCDD at 2, 4, 8, 12, 18, 24, 72, and 168 h) liver samples obtained from orally gavaged immature, ovariectomized C57BL/6 mice identified 54 differentially expressed genes (|fold change| > 1.5 and P-value < 0.1). Of these, 8 exhibited a sigmoidal or exponential dose-response profile (0.03 to 300 {mu}g/kg TCDD) at 4, 24 or 72 h. Dose-responsive genes encoded proteins associated with electron transport chain (ETC) complexes I (NADH dehydrogenase), III (cytochrome c reductase), IV (cytochrome c oxidase), and V (ATP synthase) and could be generally categorized as having proton gradient, ATP synthesis, and chaperone activities. In contrast, transcript levels of ETC complex II, succinate dehydrogenase, remained unchanged. Putative dioxin response elements were computationally found in the promoter regions of all 8 dose-responsive genes. This high-throughput approach suggests that TCDD alters the expression of genes associated with mitochondrial function which may contribute to TCDD-elicited mitochondrial toxicity.

  1. Complete mitochondrial genome sequence and gene organization of Chinese indigenous chickens with phylogenetic considerations.

    PubMed

    Zhao, F P; Fan, H Y; Li, G H; Zhang, B K

    2016-01-01

    In this study, we sequenced the complete mitochondrial DNA of Chinese indigenous Jinhu Black-bone and Rugao chickens. The two chicken mitochondrial genomes were deposited in GenBank under accession Nos. KP742951 and KR347464, respectively. The complete mitochondrial genomes of Jinhu Black-bone and Rugao chickens were sequenced and found to span 16,785 and 16,786 bp, respectively, and consisted of 22 tRNA genes, two rRNA genes (12S rRNA and 16S rRNA), 13 protein-coding genes, and one control region (D-loop). The majority of genes were positioned on the H-strand, and the ND6 and eight tRNA genes were found to be encoded on the L-strand. The mitogenomes showed a similar gene order to that of the published Gallus gallus genome, as neither included a control region. The overall base composition of the genome of the two chickens was A = 30.22/30.28%, G = 13.57/13.49%, T = 23.74/23.76%, and C = 32.48/32.48%. Nucleotide skewness of the coding strands of the two chicken genomes (AT-skew = 0.12, GC-skew = -0.41) was biased towards T and G. Phylogenetic analysis revealed 29 subspecies, and the molecular genetic relationship among the 29 subspecies was identical to that of traditional taxonomy. PMID:27421002

  2. Complete mitochondrial genome DNA sequence for two ophiuroids and a holothuroid: the utility of protein gene sequence and gene maps in the analyses of deep deuterostome phylogeny.

    PubMed

    Scouras, Andrea; Beckenbach, Karen; Arndt, Allan; Smith, Michael J

    2004-04-01

    The complete mitochondrial genome sequences have been determined for the holothuroid Cucumaria miniata and two ophiuroid species Ophiopholis aculeata and Ophiura lütkeni. In addition, the nucleotide sequence of the mitochondrial protein-coding genes for the asteroid Pisaster ochraceus has been completed. Maximum-likelihood and LogDet distance analyses of concatenated protein-coding sequences produced a series of trees that did not conclusively support generally accepted models of echinoderm phylogeny. The ophiuroid data consistently demonstrated accelerated nucleotide divergence rates and lack of stationarity. This confounds the phylogenetic analyses. Molecular investigations using individual protein-coding gene alignments demonstrated that the cytochrome b gene exhibits the least deviation in rate and stationarity and generated some trees consistent with proposed echinoderm phylogenies. Phylogenies based on echinoderm mitochondrial gene rearrangements also proved problematic because of extensive variation in gene order between and within classes. A comparison of the two distinctive ophiuroid mitochondrial gene orders supports the hypothesis that O. lütkeni has a more derived mitochondrial gene order versus O. aculeata. The variation in the echinoderm mitochondrial gene maps reinforces the limitations of the application of mitochondrial gene rearrangements as a global phylogenetic tool. PMID:15019608

  3. Cloning and analysis of a Trichinella pseudospiralis muscle larva secreted serine protease gene

    PubMed Central

    Cwiklinski, Krystyna; Meskill, Diana; Robinson, Mark W.; Pozio, E.; Appleton, Judith A.; Connolly, Bernadette

    2009-01-01

    Nematode parasites of the genus Trichinella are intracellular and distinct life cycle stages invade intestinal epithelial and skeletal muscle cells. Within the genus, Trichinella spiralis and Trichinella pseudospiralis exhibit species-specific differences with respect to host-parasite complex formation and host immune modulation. Parasite excretory-secretory (ES) proteins play important roles at the host-parasite interface and are thought to underpin these differences in biology. Serine proteases are among the most abundant group of T. spiralis ES proteins and multiple isoforms of the muscle larvae-specific TspSP-1 serine protease have been identified. Recently, a similar protein (TppSP-1) in T. pseudospiralis muscle larvae was identified. Here we report the cloning and characterisation of the full-length transcript of TppSP-1 and present comparative data between TspSP-1 and TppSP-1. PMID:19054614

  4. Cloning and analysis of a Trichinella pseudospiralis muscle larva secreted serine protease gene.

    PubMed

    Cwiklinski, Krystyna; Meskill, Diana; Robinson, Mark W; Pozio, Eduardo; Appleton, Judith A; Connolly, Bernadette

    2009-02-23

    Nematode parasites of the genus Trichinella are intracellular and distinct life cycle stages invade intestinal epithelial and skeletal muscle cells. Within the genus, Trichinella spiralis and Trichinella pseudospiralis exhibit species-specific differences with respect to host-parasite complex formation and host immune modulation. Parasite excretory-secretory (ES) proteins play important roles at the host-parasite interface and are thought to underpin these differences in biology. Serine proteases are among the most abundant group of T. spiralis ES proteins and multiple isoforms of the muscle larvae-specific TspSP-1 serine protease have been identified. Recently, a similar protein (TppSP-1) in T. pseudospiralis muscle larvae was identified. Here we report the cloning and characterisation of the full-length transcript of TppSP-1 and present comparative data between TspSP-1 and TppSP-1. PMID:19054614

  5. Nutrition Therapy for Mitochondrial Neurogastrointestinal Encephalopathy with Homozygous Mutation of the TYMP Gene.

    PubMed

    Wang, Jing; Chen, Wei; Wang, Fang; Wu, Dong; Qian, Jiaming; Kang, Junren; Li, Hailong; Ma, Enling

    2015-04-01

    Mitochondrial neurogastrointestinal encephalopathy (MNGIE) is characterized by significant gastrointestinal dysmotility. Early and long-term nutritional therapy is highly recommended. We report a case of MNGIE in a patient who was undergoing long-term nutrition therapy. The patient was diagnosed with a serious symptom of fatty liver and hyperlipidemia complications, along with homozygous mutation of the thymidine phosphorylase (TYMP) gene (c.217G > A). To our knowledge, this is the first report of such a case. Herein, we describe preventive measures for the aforementioned complications and mitochondrial disease-specific nutritional therapy. PMID:25954734

  6. Nutrition Therapy for Mitochondrial Neurogastrointestinal Encephalopathy with Homozygous Mutation of the TYMP Gene

    PubMed Central

    Wang, Jing; Wang, Fang; Wu, Dong; Qian, Jiaming; Kang, Junren; Li, Hailong; Ma, Enling

    2015-01-01

    Mitochondrial neurogastrointestinal encephalopathy (MNGIE) is characterized by significant gastrointestinal dysmotility. Early and long-term nutritional therapy is highly recommended. We report a case of MNGIE in a patient who was undergoing long-term nutrition therapy. The patient was diagnosed with a serious symptom of fatty liver and hyperlipidemia complications, along with homozygous mutation of the thymidine phosphorylase (TYMP) gene (c.217G > A). To our knowledge, this is the first report of such a case. Herein, we describe preventive measures for the aforementioned complications and mitochondrial disease-specific nutritional therapy. PMID:25954734

  7. 5-HT2 Receptor Regulation of Mitochondrial Genes: Unexpected Pharmacological Effects of Agonists and Antagonists.

    PubMed

    Harmon, Jennifer L; Wills, Lauren P; McOmish, Caitlin E; Demireva, Elena Y; Gingrich, Jay A; Beeson, Craig C; Schnellmann, Rick G

    2016-04-01

    In acute organ injuries, mitochondria are often dysfunctional, and recent research has revealed that recovery of mitochondrial and renal functions is accelerated by induction of mitochondrial biogenesis (MB). We previously reported that the nonselective 5-HT2 receptor agonist DOI [1-(4-iodo-2,5-dimethoxyphenyl)propan-2-amine] induced MB in renal proximal tubular cells (RPTCs). The goal of this study was to determine the role of 5-HT2 receptors in the regulation of mitochondrial genes and oxidative metabolism in the kidney. The 5-HT2C receptor agonist CP-809,101 [2-[(3-chlorophenyl)methoxy]-6-(1-piperazinyl)pyrazine] and antagonist SB-242,084 [6-chloro-2,3-dihydro-5-methyl-N-[6-[(2-methyl-3-pyridinyl)oxy]-3-pyridinyl]-1H-indole-1-carboxyamide dihydrochloride] were used to examine the induction of renal mitochondrial genes and oxidative metabolism in RPTCs and in mouse kidneys in the presence and absence of the 5-HT2C receptor. Unexpectedly, both CP-809,101 and SB-242,084 increased RPTC respiration and peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) mRNA expression in RPTCs at 1-10 nM. In addition, CP-809,101 and SB-242,084 increased mRNA expression of PGC-1α and the mitochondrial proteins NADH dehydrogenase subunit 1 and NADH dehydrogenase (ubiquinone) β subcomplex 8 in mice. These compounds increased mitochondrial genes in RPTCs in which the 5-HT2C receptor was downregulated with small interfering RNA and in the renal cortex of mice lacking the 5-HT2C receptor. By contrast, the ability of these compounds to increase PGC-1α mRNA and respiration was blocked in RPTCs treated with 5-HT2A receptor small interfering RNA or the 5-HT2A receptor antagonist eplivanserin. In addition, the 5-HT2A receptor agonist NBOH-2C-CN [4-[2-[[(2-hydroxyphenyl)methyl]amino]ethyl]-2,5-dimethoxybenzonitrile] increased RPTC respiration at 1-100 nM. These results suggest that agonism of the 5-HT2A receptor induces MB and that the classic 5-HT2C receptor agonist CP

  8. Multisystem disorder associated with a missense mutation in the mitochondrial cytochrome b gene.

    PubMed

    Wibrand, F; Ravn, K; Schwartz, M; Rosenberg, T; Horn, N; Vissing, J

    2001-10-01

    Mitochondrial cytochrome b mutations have been reported to have a homogenous phenotype of pure exercise intolerance. We describe a novel mutation in the cytochrome b gene of mitochondrial DNA (A15579G) associated with a selective decrease of muscle complex III activity in a patient who, besides severe exercise intolerance, also has multisystem manifestations (deafness, mental retardation, retinitis pigmentosa, cataract, growth retardation, epilepsy). The point mutation is heteroplasmic in muscle (88%) and leukocytes (15%), and changes a highly conserved tyrosine to cysteine at amino acid position 278. PMID:11601507

  9. Thymidine phosphorylase gene mutations in patients with mitochondrial neurogastrointestinal encephalomyopathy syndrome.

    PubMed

    Slama, A; Lacroix, C; Plante-Bordeneuve, V; Lombès, A; Conti, M; Reimund, J M; Auxenfants, E; Crenn, P; Laforêt, P; Joannard, A; Seguy, D; Pillant, H; Joly, P; Haut, S; Messing, B; Said, G; Legrand, A; Guiochon-Mantel, A

    2005-04-01

    The mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) syndrome is characterized by the association of gastrointestinal and neurological symptoms. It is a rare autosomal recessive mitochondrial disorder with multiple mitochondrial DNA deletions and/or depletion. It is caused by thymidine phosphorylase (TP) gene mutations resulting in a complete abolition of TP activity. We tested 31 unrelated patients presenting either with a complete MNGIE syndrome (8 patients), a severe intestinal pseudo-obstruction (10 patients), and multiple deletions and/or depletion of mitochondrial DNA (13 patients). All the tested patients presenting with a complete MNGIE had increased thymidine levels in plasma and urine, and no TP activity. The group with pseudo-obstruction syndrome had normal or partial reduction of TP activity. We found pathogenic mutations on TP gene only in the MNGIE syndrome group: all the MNGIE patients were compound heterozygous or homozygous for mutations in the TP gene. Eight of these mutations are yet unreported, confirming the lack of genotype/phenotype correlation in this syndrome. Enzymatic activity and thymidine level are thus rapid diagnosis tests to detect MNGIE affected patients prior to genetic testing for patients with gastrointestinal symptoms. PMID:15781193

  10. Identification and mapping of trnI, trnE and trnfM genes in the sunflower mitochondrial genome.

    PubMed

    Ceci, L R; Veronico, P; Siculella, L; Gallerani, R

    1995-01-01

    Three sunflower mitochondrial HindIII restriction fragments containing the tRNA genes trnI, trnE and trnfM have been sequenced. The genes are present in single copy on the whole genome and are transcribed. Hybridization experiments and sequence analysis of the HindIII fragments allowed the precise mapping and orientation of each gene on the sunflower mitochondrial genome. PMID:7579587

  11. Different control mechanisms regulate glucoamylase and protease gene transcription in Aspergillus oryzae in solid-state and submerged fermentation.

    PubMed

    te Biesebeke, R; van Biezen, N; de Vos, W M; van den Hondel, C A M J J; Punt, P J

    2005-04-01

    Solid-state fermentation (SSF) with Aspergillus oryzae results in high levels of secreted protein. However, control mechanisms of gene expression in SSF have been only poorly studied. In this study we show that both glucoamylase (glaB) and protease (alpA, nptB) genes are highly expressed during surface cultivation on wheat-based solid medium, and even higher during cultivation on wheat kernels. In wheat-based liquid medium, low levels of gene expression are observed. Typical SSF cultivation conditions, such as low water activity and the formation of aerial hyphae, did not contribute to the high-level gene expression on wheat-based solid medium. Analysis of wheat-based solid and liquid cultivations showed differences in carbon and nitrogen utilisation and external pH. The results presented show that the difference in regulation of transcription of the alpA and nptB genes in wheat-based liquid and solid medium could be pH dependent, involving a pH-dependent transcription regulator. The results obtained suggest that the difference in regulation of transcription of the glaB gene in wheat-based liquid and solid medium is caused by a difference in carbohydrate degradation and consumption under the different culture conditions. PMID:15800731

  12. Characteristic features of the nucleotide sequences of yeast mitochondrial ribosomal protein genes as analyzed by computer program GeneMark.

    PubMed

    Isono, K; McIninch, J D; Borodovsky, M

    1994-01-01

    The nucleotide sequence data for yeast mitochondrial ribosomal protein (MRP) genes were analyzed by the computer program GeneMark which predicts the presence of likely genes in sequence data by calculating statistical biases in the appearance of consecutive nucleotides. The program uses a set of standard sequence data for this calculation. We used this program for the analysis of yeast nucleotide sequence data containing MRP genes, hoping to obtain information as to whether they share features in common that are different from other yeast genes. Sequence data sets for ordinary yeast genes and for 27 known MRP genes were used. The MRP genes were nicely predicted as likely genes regardless of the data sets used, whereas other yeast genes were predicted to be likely genes only when the data set for ordinary yeast genes was used. The assembled sequence data for chromosomes II, III, VIII and XI as well as the segmented data for chromosome V were analyzed in a similar manner. In addition to the known MRP genes, eleven ORF's were predicted to be likely MRP genes. Thus, the method seems very powerful in analyzing genes of heterologous origins. PMID:7719921

  13. Discovery of the rpl10 Gene in Diverse Plant Mitochondrial Genomes and Its Probable Replacement by the Nuclear Gene for Chloroplast RPL10 in Two Lineages of Angiosperms

    PubMed Central

    Kubo, Nakao; Arimura, Shin-ichi

    2010-01-01

    Mitochondrial genomes of plants are much larger than those of mammals and often contain conserved open reading frames (ORFs) of unknown function. Here, we show that one of these conserved ORFs is actually the gene for ribosomal protein L10 (rpl10) in plant. No rpl10 gene has heretofore been reported in any mitochondrial genome other than the exceptionally gene-rich genome of the protist Reclinomonas americana. Conserved ORFs corresponding to rpl10 are present in a wide diversity of land plant and green algal mitochondrial genomes. The mitochondrial rpl10 genes are transcribed in all nine land plants examined, with five seed plant genes subject to RNA editing. In addition, mitochondrial-rpl10-like cDNAs were identified in EST libraries from numerous land plants. In three lineages of angiosperms, rpl10 is either lost from the mitochondrial genome or a pseudogene. In two of them (Brassicaceae and monocots), no nuclear copy of mitochondrial rpl10 is identifiably present, and instead a second copy of nuclear-encoded chloroplast rpl10 is present. Transient assays using green fluorescent protein indicate that this duplicate gene is dual targeted to mitochondria and chloroplasts. We infer that mitochondrial rpl10 has been functionally replaced by duplicated chloroplast counterparts in Brassicaceae and monocots. PMID:19934175

  14. [Complete sequence and gene organization of the Tibetan chicken mitochondrial genome].

    PubMed

    Tong, Xiao-Mei; Liang, Yu; Wang, Wei; Xu, Shu-Qing; Zheng, Xiao-Guang; Wang, Jian; Yu, Jun

    2006-07-01

    Using PCR amplification, sequencing and assembling, we obtained the complete mitochondrial genome of Tibetan chicken. The complete mitochondrial genome was 16 783 bp in length. It contained 37 genes (13 protein coding genes, 2 rRNA, 22 tRNA) and a control region. The deduced restriction map revealed a unique pattern of Dra I restriction in Tibetan chicken. Phylogenetic trees based on the D-loop locus and the 13 protein coding genes by Neighbor-joining and Maximum Parsimony analysis indicated that the red junglefowl was the direct ancestor of Tibetan chicken and Tibetan chicken was closest to white leghorn and white plymouth rock, although the evolution of Tibetan chicken appeared to be relatively independent from them. A possible explanation is that the ancestor of Tibetan chicken lived in a relatively isolated environment after entering into the high altitude area and developed unique genetic characters. PMID:16825161

  15. Towards germline gene therapy of inherited mitochondrial diseases

    PubMed Central

    Tachibana, Masahito; Amato, Paula; Sparman, Michelle; Woodward, Joy; Sanchis, Dario Melguizo; Ma, Hong; Gutierrez, Nuria Marti; Tippner-Hedges, Rebecca; Kang, Eunju; Lee, Hyo-Sang; Ramsey, Cathy; Masterson, Keith; Battaglia, David; Lee, David; Wu, Diana; Jensen, Jeffrey; Patton, Phillip; Gokhale, Sumita; Stouffer, Richard; Mitalipov, Shoukhrat

    2012-01-01

    Mutations in mitochondrial DNA (mtDNA) are associated with serious human diseases and inherited from mother's eggs. Here we investigated the feasibility of mtDNA replacement in human oocytes by spindle transfer (ST). Of 106 human oocytes donated for research, 65 were subjected to reciprocal ST and 33 served as controls. Fertilization rate in ST oocytes (73%) was similar to controls (75%). However, a significant portion of ST zygotes (52%) displayed abnormal fertilization as determined by irregular number of pronuclei. Among normally fertilized ST zygotes, blastocyst development (62%) and embryonic stem cell (ESC) isolation (38%) rates were comparable to controls. All ESC lines derived from ST zygotes displayed normal euploid karyotypes and contained exclusively donor mtDNA. The mtDNA can be efficiently replaced in human oocytes. Although some ST oocytes displayed abnormal fertilization, remaining embryos were capable of developing to blastocysts and producing ESCs similar to controls. PMID:23103867

  16. Strikingly Bacteria-Like and Gene-Rich Mitochondrial Genomes throughout Jakobid Protists

    PubMed Central

    Burger, Gertraud; Gray, Michael W.; Forget, Lise; Lang, B. Franz

    2013-01-01

    The most bacteria-like mitochondrial genome known is that of the jakobid flagellate Reclinomonas americana NZ. This genome also encodes the largest known gene set among mitochondrial DNAs (mtDNAs), including the RNA subunit of RNase P (transfer RNA processing), a reduced form of transfer–messenger RNA (translational control), and a four-subunit bacteria-like RNA polymerase, which in other eukaryotes is substituted by a nucleus-encoded, single-subunit, phage-like enzyme. Further, protein-coding genes are preceded by potential Shine–Dalgarno translation initiation motifs. Whether similarly ancestral mitochondrial characters also exist in relatives of R. americana NZ is unknown. Here, we report a comparative analysis of nine mtDNAs from five distant jakobid genera: Andalucia, Histiona, Jakoba, Reclinomonas, and Seculamonas. We find that Andalucia godoyi has an even larger mtDNA gene complement than R. americana NZ. The extra genes are rpl35 (a large subunit mitoribosomal protein) and cox15 (involved in cytochrome oxidase assembly), which are nucleus encoded throughout other eukaryotes. Andalucia cox15 is strikingly similar to its homolog in the free-living α-proteobacterium Tistrella mobilis. Similarly, a long, highly conserved gene cluster in jakobid mtDNAs, which is a clear vestige of prokaryotic operons, displays a gene order more closely resembling that in free-living α-proteobacteria than in Rickettsiales species. Although jakobid mtDNAs, overall, are characterized by bacteria-like features, they also display a few remarkably divergent characters, such as 3′-tRNA editing in Seculamonas ecuadoriensis and genome linearization in Jakoba libera. Phylogenetic analysis with mtDNA-encoded proteins strongly supports monophyly of jakobids with Andalucia as the deepest divergence. However, it remains unclear which α-proteobacterial group is the closest mitochondrial relative. PMID:23335123

  17. Whole mitochondrial genome analysis in two families with dilated mitochondrial cardiomyopathy: detection of mutations in MT-ND2 and MT-TL1 genes.

    PubMed

    Alila, Olfa Fersi; Rebai, Emna Mkaouar; Tabebi, Mouna; Tej, Amel; Chamkha, Imen; Tlili, Abdelaziz; Bouguila, Jihene; Tilouche, Samia; Soyah, Nejla; Boughamoura, Lamia; Fakhfakh, Faiza

    2016-07-01

    Pathogenic mitochondrial DNA (mtDNA) mutations leading to mitochondrial dysfunction can cause cardiomyopathy and heart failure. These mutations were described in the mt-tRNA genes and in the mitochondrial protein-coding genes. The aim of this study was to identify the genetic defect in two patients belonging to two families with cardiac dysfunction associated to a wide spectrum of clinical phenotypes. The sequencing analysis of the whole mitochondrial DNA in the two patients and their parents revealed the presence of known polymorphisms associated to cardiomyopathy and two pathogenic mutations in DNA extracted from blood leucocytes: the heteroplasmic m.3243A > G mutation in the MT-TL1 gene in patient A; and the homoplasmic m.5182C > T mutation in the ND2 gene in patient B. Secondary structure analysis of the ND2 protein further supported the deleterious role of the m.5182C > T mutation, as it was found to be involved an extended imbalance in its hydrophobicity and affect its function. In addition, the mitochondrial variants identified in patients A and B classify both of them in the same haplogroup H2a2a1. PMID:26258512

  18. Identification of the gene encoding the mitochondrial elongation factor G in mammals.

    PubMed Central

    Barker, C; Makris, A; Patriotis, C; Bear, S E; Tsichlis, P N

    1993-01-01

    Protein synthesis in cytosolic and rough endoplasmic reticulum associated ribosomes is directed by factors, many of which have been well characterized. Although these factors have been the subject of intense study, most of the corresponding factors regulating protein synthesis in the mitochondrial ribosomes remain unknown. In this report we present the cloning and initial characterization of the gene encoding the rat mitochondrial elongation factor-G (rEF-Gmt). The rat gene encoding EF-Gmt (rMef-g) maps to rat chromosome 2 and it is expressed in all tissues with highest levels in liver, thymus and brain. Its DNA sequence predicts a 752 amino acid protein exhibiting 72% homology to the yeast Saccharomyces cerevisiae mitochondrial elongation factor-G (YMEF-G), 62% and 61% homology to the Thermus thermophilus and E. coli elongation factor-G (EF-G) respectively and 52% homology to the rat elongation factor-2 (EF-2). The deduced amino acid sequence of EF-G contains characteristic motifs shared by all GTP binding proteins. Therefore, similarly to other elongation factors, the enzymatic function of EF-Gmt is predicted to depend on GTP binding and hydrolysis. EF-Gmt differs from its cytoplasmic homolog, EF-2, in that it contains an aspartic acid residue at amino acid position 621 which corresponds to the EF-2 histidine residue at position 715. Since this histidine residue, following posttranslational modification into diphthamide, appears to be the sole cellular target of diphtheria toxin and Pseudomonas aeruginosa endotoxin A, we conclude that EF-Gmt will not be inactivated by these toxins. The severe effects of these toxins on protein elongation in tissues expressing EF-Gmt suggest that EF-Gmt and EF-2 exhibit nonoverlapping functions. The cloning and characterization of the mammalian mitochondrial elongation factor G will permit us to address its role in the regulation of normal mitochondrial function and in disease states attributed to mitochondrial dysfunction. Images

  19. Mitochondrial genomes of Clymenella torquata (Maldanidae) and Riftia pachyptila (Siboglinidae): evidence for conserved gene order in annelida.

    PubMed

    Jennings, Robert M; Halanych, Kenneth M

    2005-02-01

    Mitochondrial genomes are useful tools for inferring evolutionary history. However, many taxa are poorly represented by available data. Thus, to further understand the phylogenetic potential of complete mitochondrial genome sequence data in Annelida (segmented worms), we examined the complete mitochondrial sequence for Clymenella torquata (Maldanidae) and an estimated 80% of the sequence of Riftia pachyptila (Siboglinidae). These genomes have remarkably similar gene orders to previously published annelid genomes, suggesting that gene order is conserved across annelids. This result is interesting, given the high variation seen in the closely related Mollusca and Brachiopoda. Phylogenetic analyses of DNA sequence, amino acid sequence, and gene order all support the recent hypothesis that Sipuncula and Annelida are closely related. Our findings suggest that gene order data is of limited utility in annelids but that sequence data holds promise. Additionally, these genomes show AT bias (approximately 66%) and codon usage biases but have a typical gene complement for bilaterian mitochondrial genomes. PMID:15483328

  20. Screen for mitochondrial DNA copy number maintenance genes reveals essential role for ATP synthase

    PubMed Central

    Fukuoh, Atsushi; Cannino, Giuseppe; Gerards, Mike; Buckley, Suzanne; Kazancioglu, Selena; Scialo, Filippo; Lihavainen, Eero; Ribeiro, Andre; Dufour, Eric; Jacobs, Howard T

    2014-01-01

    The machinery of mitochondrial DNA (mtDNA) maintenance is only partially characterized and is of wide interest due to its involvement in disease. To identify novel components of this machinery, plus other cellular pathways required for mtDNA viability, we implemented a genome-wide RNAi screen in Drosophila S2 cells, assaying for loss of fluorescence of mtDNA nucleoids stained with the DNA-intercalating agent PicoGreen. In addition to previously characterized components of the mtDNA replication and transcription machineries, positives included many proteins of the cytosolic proteasome and ribosome (but not the mitoribosome), three proteins involved in vesicle transport, some other factors involved in mitochondrial biogenesis or nuclear gene expression, > 30 mainly uncharacterized proteins and most subunits of ATP synthase (but no other OXPHOS complex). ATP synthase knockdown precipitated a burst of mitochondrial ROS production, followed by copy number depletion involving increased mitochondrial turnover, not dependent on the canonical autophagy machinery. Our findings will inform future studies of the apparatus and regulation of mtDNA maintenance, and the role of mitochondrial bioenergetics and signaling in modulating mtDNA copy number. PMID:24952591

  1. The Mitochondrial Genome of Soybean Reveals Complex Genome Structures and Gene Evolution at Intercellular and Phylogenetic Levels

    PubMed Central

    Chang, Shengxin; Wang, Yankun; Lu, Jiangjie; Gai, Junyi; Li, Jijie; Chu, Pu; Guan, Rongzhan; Zhao, Tuanjie

    2013-01-01

    Determining mitochondrial genomes is important for elucidating vital activities of seed plants. Mitochondrial genomes are specific to each plant species because of their variable size, complex structures and patterns of gene losses and gains during evolution. This complexity has made research on the soybean mitochondrial genome difficult compared with its nuclear and chloroplast genomes. The present study helps to solve a 30-year mystery regarding the most complex mitochondrial genome structure, showing that pairwise rearrangements among the many large repeats may produce an enriched molecular pool of 760 circles in seed plants. The soybean mitochondrial genome harbors 58 genes of known function in addition to 52 predicted open reading frames of unknown function. The genome contains sequences of multiple identifiable origins, including 6.8 kb and 7.1 kb DNA fragments that have been transferred from the nuclear and chloroplast genomes, respectively, and some horizontal DNA transfers. The soybean mitochondrial genome has lost 16 genes, including nine protein-coding genes and seven tRNA genes; however, it has acquired five chloroplast-derived genes during evolution. Four tRNA genes, common among the three genomes, are derived from the chloroplast. Sizeable DNA transfers to the nucleus, with pericentromeric regions as hotspots, are observed, including DNA transfers of 125.0 kb and 151.6 kb identified unambiguously from the soybean mitochondrial and chloroplast genomes, respectively. The soybean nuclear genome has acquired five genes from its mitochondrial genome. These results provide biological insights into the mitochondrial genome of seed plants, and are especially helpful for deciphering vital activities in soybean. PMID:23431381

  2. A One-Megabase Physical Map Provides Insights on Gene Organization in the Enormous Mitochondrial Genome of Cucumber

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cucumber has one of the largest mitochondrial genomes known among all eukaryotes, due in part to the accumulation of short repetitive-DNA motifs. Recombination among these repetitive DNAs produces rearrangements affecting organization and expression of mitochondrial genes. In order to more efficie...

  3. Molecular characterization of a gene encoding extracellular serine protease isolated from a subtilisin inhibitor-deficient mutant of Streptomyces albogriseolus S-3253.

    PubMed Central

    Taguchi, S; Odaka, A; Watanabe, Y; Momose, H

    1995-01-01

    An extracellular serine protease produced by a mutant, M1, derived from Streptomyces albogriseolus S-3253 that no longer produces a protease inhibitor (Streptomyces subtilisin inhibitor [SSI]) was isolated. A 20-kDa protein was purified by its affinity for SSI and designated SAM-P20. The amino acid sequence of the amino-terminal region of SAM-P20 revealed high homology with the sequences of Streptomyces griseus proteases A and B, and the gene sequence confirmed the relationships. The sequence also revealed a putative amino acid signal sequence for SAM-P20 that apparently functioned to allow secretion of SAM-P20 from Escherichia coli carrying the recombinant gene. SAM-P20 produced by E. coli cells was shown to be sensitive to SSI inhibition. PMID:7887600

  4. Rearrangement of mitochondrial tRNA genes in flat bugs (Hemiptera: Aradidae)

    PubMed Central

    Song, Fan; Li, Hu; Shao, Renfu; Shi, Aimin; Bai, Xiaoshuan; Zheng, Xiaorong; Heiss, Ernst; Cai, Wanzhi

    2016-01-01

    The typical insect mitochondrial (mt) genome organization, which contains a single chromosome with 37 genes, was found in the infraorder Pentatomomorpha (suborder Heteroptera). The arrangement of mt genes in these true bugs is usually the same as the ancestral mt gene arrangement of insects. Rearrangement of transfer RNA (tRNA) genes, however, has been found in two subfamilies of flat bugs (Mezirinae and Calisiinae, family Aradidae). In this study, we sequenced the complete mt genomes of four species from three other subfamilies (Aradinae, Carventinae and Aneurinae). We found tRNA gene rearrangement in all of these four species. All of the rearranged tRNA genes are located between the mitochondrial control region and cox1, indicating this region as a hotspot for gene rearrangement in flat bugs; the rearrangement is likely caused by events of tandem duplication and random deletion of genes. Furthermore, our phylogenetic and dating analyses indicated that the swap of positions between trnQ and trnI occurred ~162 million years ago (MYA) in the most recent common ancestor of the five subfamilies of flat bugs investigated to date, whereas the swap of positions between trnC and trnW occurred later in the lineage leading to Calisiinae, and the translocation of trnC and trnY occurred later than 134 MYA in the lineage leading to Aradinae. PMID:27180804

  5. Rearrangement of mitochondrial tRNA genes in flat bugs (Hemiptera: Aradidae).

    PubMed

    Song, Fan; Li, Hu; Shao, Renfu; Shi, Aimin; Bai, Xiaoshuan; Zheng, Xiaorong; Heiss, Ernst; Cai, Wanzhi

    2016-01-01

    The typical insect mitochondrial (mt) genome organization, which contains a single chromosome with 37 genes, was found in the infraorder Pentatomomorpha (suborder Heteroptera). The arrangement of mt genes in these true bugs is usually the same as the ancestral mt gene arrangement of insects. Rearrangement of transfer RNA (tRNA) genes, however, has been found in two subfamilies of flat bugs (Mezirinae and Calisiinae, family Aradidae). In this study, we sequenced the complete mt genomes of four species from three other subfamilies (Aradinae, Carventinae and Aneurinae). We found tRNA gene rearrangement in all of these four species. All of the rearranged tRNA genes are located between the mitochondrial control region and cox1, indicating this region as a hotspot for gene rearrangement in flat bugs; the rearrangement is likely caused by events of tandem duplication and random deletion of genes. Furthermore, our phylogenetic and dating analyses indicated that the swap of positions between trnQ and trnI occurred ~162 million years ago (MYA) in the most recent common ancestor of the five subfamilies of flat bugs investigated to date, whereas the swap of positions between trnC and trnW occurred later in the lineage leading to Calisiinae, and the translocation of trnC and trnY occurred later than 134 MYA in the lineage leading to Aradinae. PMID:27180804

  6. Probable presence of an ubiquitous cryptic mitochondrial gene on the antisense strand of the cytochrome oxidase I gene

    PubMed Central

    2011-01-01

    Background Mitochondria mediate most of the energy production that occurs in the majority of eukaryotic organisms. These subcellular organelles contain a genome that differs from the nuclear genome and is referred to as mitochondrial DNA (mtDNA). Despite a disparity in gene content, all mtDNAs encode at least two components of the mitochondrial electron transport chain, including cytochrome c oxidase I (Cox1). Presentation of the hypothesis A positionally conserved ORF has been found on the complementary strand of the cox1 genes of both eukaryotic mitochondria (protist, plant, fungal and animal) and alpha-proteobacteria. This putative gene has been named gau for gene antisense ubiquitous in mtDNAs. The length of the deduced protein is approximately 100 amino acids. In vertebrates, several stop codons have been found in the mt gau region, and potentially functional gau regions have been found in nuclear genomes. However, a recent bioinformatics study showed that several hypothetical overlapping mt genes could be predicted, including gau; this involves the possible import of the cytosolic AGR tRNA into the mitochondria and/or the expression of mt antisense tRNAs with anticodons recognizing AGR codons according to an alternative genetic code that is induced by the presence of suppressor tRNAs. Despite an evolutionary distance of at least 1.5 to 2.0 billion years, the deduced Gau proteins share some conserved amino acid signatures and structure, which suggests a possible conserved function. Moreover, BLAST analysis identified rare, sense-oriented ESTs with poly(A) tails that include the entire gau region. Immunohistochemical analyses using an anti-Gau monoclonal antibody revealed strict co-localization of Gau proteins and a mitochondrial marker. Testing the hypothesis This hypothesis could be tested by purifying the gau gene product and determining its sequence. Cell biological experiments are needed to determine the physiological role of this protein. Implications of

  7. The complete mitochondrial genome of the fire coral-inhabiting barnacle Megabalanus ajax (Sessilia: Balanidae): gene rearrangements and atypical gene content.

    PubMed

    Shen, Xin; Chu, Ka Hou; Chan, Benny Kwok Kan; Tsang, Ling Ming

    2016-01-01

    The complete mitochondrial genome of Megabalanus ajax Darwin, 1854 (Sessilia: Balanidae) is reported. Compared to typical gene content of metazoan mitochondrial genomes, duplication of one tRNA gene (trnL2) and absence of another tRNA gene (trnS1) are identified in M. ajax mitochondrial genome. There is a replacement of one tRNA (trnS1) by another tRNA (trnL2) in M. ajax mitochondrial genome compared to Megabalanus volcano mitochondrial genome. Inversion of a six-gene block (trnP-nd4L-nd4-trnH-nd5-trnF) is found between M. ajax/M. volcano and Tetraclita japonica mitochondrial genomes. With reference to the pancrustacean mitochondrial ground pattern, there is an inversion of a large gene block from the light strand to heavy strand in the two Megabalanus mitochondrial genomes, including three PCGs and two tRNAs (nd4L-nd4-trnH-nd5-trnF). Furthermore, four tRNAs (trnA, trnE, trnQ and trnC) exhibit translocation, while translocation and inversion occur in three tRNAs (trnP, trnY and trnK). PMID:25050875

  8. Mitochondrially-targeted expression of a cytoplasmic male sterility-associated orf220 gene causes male sterility in Brassica juncea

    PubMed Central

    2010-01-01

    Background The novel chimeric open reading frame (orf) resulting from the rearrangement of a mitochondrial genome is generally thought to be a causal factor in the occurrence of cytoplasmic male sterility (CMS). Both positive and negative correlations have been found between CMS-associated orfs and the occurrence of CMS when CMS-associated orfs were expressed and targeted at mitochondria. Some orfs cause male sterility or semi-sterility, while some do not. Little is currently known about how mitochondrial factor regulates the expression of the nuclear genes involved in male sterility. The purpose of this study was to investigate the biological function of a candidate CMS-associated orf220 gene, newly isolated from cytoplasmic male-sterile stem mustard, and show how mitochondrial retrograde regulated nuclear gene expression is related to male sterility. Results It was shown that the ORF220 protein can be guided to the mitochondria using the mitochondrial-targeting sequence of the β subunit of F1-ATPase (atp2-1). Transgenic stem mustard plants expressed the chimeric gene containing the orf220 gene and a mitochondrial-targeting sequence of the β subunit of F1-ATPase (atp2-1). Transgenic plants were male-sterile, most being unable to produce pollen while some could only produce non-vigorous pollen. The transgenic stem mustard plants also showed aberrant floral development identical to that observed in the CMS stem mustard phenotype. Results obtained from oligooarray analysis showed that some genes related to mitochondrial energy metabolism were down-regulated, indicating a weakening of mitochondrial function in transgenic stem mustard. Some genes related to pollen development were shown to be down-regulated in transgenic stem mustard and the expression of some transcription factor genes was also altered. Conclusion The work presented furthers our understanding of how the mitochondrially-targeted expression of CMS-associated orf220 gene causes male sterility through

  9. Identification and characterization of alkaline protease producing Bacillus firmus species EMBS023 by 16S rRNA gene sequencing.

    PubMed

    Wishard, Rohan; wishard, Rohan; Jaiswal, Mahak; Parveda, Maheshwari; Amareshwari, P; Bhadoriya, Sneha Singh; Rathore, Pragya; Yadav, Mukesh; Nayarisseri, Anuraj; Nair, Achuthsankar S

    2014-12-01

    Probiotic microorganisms are those which exert a positive exect on the growth of the host, when administered as a dietary mixture in an adequate amount. They form the best alternative to the use of antibiotics for controlling enteric diseases in poultry farm animals, especially in the light of the gruesome problems of development of antibiotic resistance in enteric pathogens and the contamination of poultry products with antibiotics. 16S rDNA sequencing which has gained wide popularity amongst microbiologists for the molecular characterization and identification of newly discovered isolates provides accurate identification of isolates down to the level of sub-species (strain). It's most important advantage over the traditional biochemical characterization methods are that it can provide an accurate identification of strains with atypical phenotypic characters as well. The following work is an application of 16S rRNA gene sequencing approach to identify a novel, alkaline protease producing bacteria, from poultry farm waste. The sample was collected from a local poultry farm in the Guntur district, Andhra Pradesh, India. Subsequently the sample was serially diluted and the aliquots were incubated for a suitable time period following which the suspected colony was subjected to 16S rDNA sequencing. The results showed the isolate to be a novel, high alkaline protease producing bacteria, which was named Bacillus firmus isolate EMBS023, after characterization the sequence of isolate was deposited in GenBank with accession number JN990980. PMID:25118655

  10. Phenotypic characterization of virological failure following lopinavir/ritonavir monotherapy using full-length gag–protease genes

    PubMed Central

    Sutherland, Katherine A.; Mbisa, Jean L.; Ghosn, Jade; Chaix, Marie-Laure; Cohen-Codar, Isabelle; Hue, Stephane; Delfraissy, Jean-Francois; Delaugerre, Constance; Gupta, Ravindra K.

    2014-01-01

    Objectives Major protease mutations are rarely observed following first-line failure with PIs and interpretation of genotyping results in this context may be difficult. We performed extensive phenotyping of viruses from five patients failing lopinavir/ritonavir monotherapy in the MONARK study without major PI mutations by standard genotyping. Methods Phenotypic susceptibility testing and viral infectivity assessments were performed using a single-cycle assay and fold changes (FC) relative to a lopinavir-susceptible reference strain were calculated. Results >10-fold reduced baseline susceptibility to lopinavir occurred in two of five patients and >5-fold in another two. Four of five patients exhibited phylogenetic evidence of a limited viral evolution between baseline and failure, with amino acid changes at drug resistance-associated positions in one: T81A emerged in Gag with M36I in the protease gene, correlating with a reduction in lopinavir susceptibility from FC 7 (95% CI 6–8.35) to FC 13 (95% CI 8.11–17.8). Reductions in darunavir susceptibility (>5 FC) occurred in three individuals. Discussion This study suggests both baseline reduced susceptibility and evolution of resistance could be contributing factors to PI failure, despite the absence of classical PI resistance mutations by standard testing methods. Use of phenotyping also reveals lower darunavir susceptibility, warranting further study as this agent is commonly used following lopinavir failure. PMID:25096075

  11. The Yeast Gene, MDM20, Is Necessary for Mitochondrial Inheritance and Organization of the Actin Cytoskeleton

    PubMed Central

    Hermann, Greg J.; King, Edward J.; Shaw, Janet M.

    1997-01-01

    In Saccharomyces cerevisiae, the growing bud inherits a portion of the mitochondrial network from the mother cell soon after it emerges. Although this polarized transport of mitochondria is thought to require functions of the cytoskeleton, there are conflicting reports concerning the nature of the cytoskeletal element involved. Here we report the isolation of a yeast mutant, mdm20, in which both mitochondrial inheritance and actin cables (bundles of actin filaments) are disrupted. The MDM20 gene encodes a 93-kD polypeptide with no homology to other characterized proteins. Extra copies of TPM1, a gene encoding the actin filament–binding protein tropomyosin, suppress mitochondrial inheritance defects and partially restore actin cables in mdm20Δ cells. Synthetic lethality is also observed between mdm20 and tpm1 mutant strains. Overexpression of a second yeast tropomyosin, Tpm2p, rescues mutant phenotypes in the mdm20 strain to a lesser extent. Together, these results provide compelling evidence that mitochondrial inheritance in yeast is an actin-mediated process. MDM20 and TPM1 also exhibit the same pattern of genetic interactions; mutations in MDM20 are synthetically lethal with mutations in BEM2 and MYO2 but not SAC6. Although MDM20 and TPM1 are both required for the formation and/or stabilization of actin cables, mutations in these genes disrupt mitochondrial inheritance and nuclear segregation to different extents. Thus, Mdm20p and Tpm1p may act in vivo to establish molecular and functional heterogeneity of the actin cytoskeleton. PMID:9105043

  12. Clinical and Neuroimaging Features in Two Children with Mutations in the Mitochondrial ND5 Gene.

    PubMed

    Sonam, Kothari; Bindu, P S; Taly, Arun B; Govindaraju, Chikkanna; Gayathri, Narayanappa; Arvinda, Hanumanthapura R; Nagappa, Madhu; Sinha, Sanjib; Khan, Nahid Akthar; Govindaraj, Periyasamy; Thangaraj, Kumarasamy

    2015-08-01

    Mutations in the mitochondrial-encoded nicotinamide adenine dinucleotide dehydrogenase 5 gene (MT-ND5) has been implicated as an important genetic cause of childhood mitochondrial encephalomyopathies. This study reports the clinical and magnetic resonance imaging findings in two pediatric patients with mutations in the ND5 gene of mitochondrial DNA. The 8-month-old boy with m.13513 G>A mutation presented with infantile basal ganglia stroke syndrome secondary to mineralizing angiopathy. The 7-year-old girl with the m.13514A>G mutation had episodic regression, progressive ataxia, optic atrophy, and hyperactivity. Magnetic resonance imaging of the brain showed bilateral symmetrical signal intensity changes in the thalamus, tectal plate, and inferior olivary nucleus, which subsided on follow-up image. Both the patients had a stable course. Familiarity with the various phenotypic and magnetic resonance imaging findings and the clinical course in childhood mitochondrial encephalomyopathies may help the physician in targeted metabolic-genetic testing and prognostication. PMID:25974876

  13. A Mutation in the Mitochondrial Fission Gene Dnm1l Leads to Cardiomyopathy

    PubMed Central

    Ashrafian, Houman; Docherty, Louise; Leo, Vincenzo; Towlson, Christopher; Neilan, Monica; Steeples, Violetta; Lygate, Craig A.; Hough, Tertius; Townsend, Stuart; Williams, Debbie; Wells, Sara; Norris, Dominic; Glyn-Jones, Sarah; Land, John; Barbaric, Ivana; Lalanne, Zuzanne; Denny, Paul; Szumska, Dorota; Bhattacharya, Shoumo; Griffin, Julian L.; Hargreaves, Iain; Fernandez-Fuentes, Narcis; Cheeseman, Michael; Watkins, Hugh; Dear, T. Neil

    2010-01-01

    Mutations in a number of genes have been linked to inherited dilated cardiomyopathy (DCM). However, such mutations account for only a small proportion of the clinical cases emphasising the need for alternative discovery approaches to uncovering novel pathogenic mutations in hitherto unidentified pathways. Accordingly, as part of a large-scale N-ethyl-N-nitrosourea mutagenesis screen, we identified a mouse mutant, Python, which develops DCM. We demonstrate that the Python phenotype is attributable to a dominant fully penetrant mutation in the dynamin-1-like (Dnm1l) gene, which has been shown to be critical for mitochondrial fission. The C452F mutation is in a highly conserved region of the M domain of Dnm1l that alters protein interactions in a yeast two-hybrid system, suggesting that the mutation might alter intramolecular interactions within the Dnm1l monomer. Heterozygous Python fibroblasts exhibit abnormal mitochondria and peroxisomes. Homozygosity for the mutation results in the death of embryos midway though gestation. Heterozygous Python hearts show reduced levels of mitochondria enzyme complexes and suffer from cardiac ATP depletion. The resulting energy deficiency may contribute to cardiomyopathy. This is the first demonstration that a defect in a gene involved in mitochondrial remodelling can result in cardiomyopathy, showing that the function of this gene is needed for the maintenance of normal cellular function in a relatively tissue-specific manner. This disease model attests to the importance of mitochondrial remodelling in the heart; similar defects might underlie human heart muscle disease. PMID:20585624

  14. Clock-genes and mitochondrial respiratory activity: Evidence of a reciprocal interplay.

    PubMed

    Scrima, Rosella; Cela, Olga; Merla, Giuseppe; Augello, Bartolomeo; Rubino, Rosa; Quarato, Giovanni; Fugetto, Sabino; Menga, Marta; Fuhr, Luise; Relógio, Angela; Piccoli, Claudia; Mazzoccoli, Gianluigi; Capitanio, Nazzareno

    2016-08-01

    In the past few years mounting evidences have highlighted the tight correlation between circadian rhythms and metabolism. Although at the organismal level the central timekeeper is constituted by the hypothalamic suprachiasmatic nuclei practically all the peripheral tissues are equipped with autonomous oscillators made up by common molecular clockworks represented by circuits of gene expression that are organized in interconnected positive and negative feed-back loops. In this study we exploited a well-established in vitro synchronization model to investigate specifically the linkage between clock gene expression and the mitochondrial oxidative phosphorylation (OxPhos). Here we show that synchronized cells exhibit an autonomous ultradian mitochondrial respiratory activity which is abrogated by silencing the master clock gene ARNTL/BMAL1. Surprisingly, pharmacological inhibition of the mitochondrial OxPhos system resulted in dramatic deregulation of the rhythmic clock-gene expression and a similar result was attained with mtDNA depleted cells (Rho0). Our findings provide a novel level of complexity in the interlocked feedback loop controlling the interplay between cellular bioenergetics and the molecular clockwork. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi. PMID:27060253

  15. Partial kinetoplast-mitochondrial gene organization and expression in the respiratory deficient plant trypanosomatid Phytomonas serpens.

    PubMed

    Maslov, D A; Nawathean, P; Scheel, J

    1999-04-30

    In plant-dwelling trypanosomatids from the genus Phytomonas, mitochondrial functions, such as cytochrome mediated respiration, ATP production and Krebs cycle, are missing, and cell energetics is based on the glycolysis. Using Blue Native/Tricine-SDS two-dimensional gel electrophoretic analysis, we observed that mitochondrial respiratory Complexes III (cytochrome bc1) and IV (cytochrome c oxidase) were absent in Phytomonas serpens; however, Complex V (ATPase) was present. A deletion of the genes for cytochrome c oxidase subunit III (COIII) and apocytochrome b (Cyb) was identified within the 6234 bp sequenced region of the 31 kb maxicircle kinetoplast DNA. Genes, found in this region, include 12S and 9S ribosomal RNAs, subunits 7, 8 and 9 of NADH dehydrogenase (ND7, ND8 and ND9) and subunit 6 of ATPase (A6 or MURF4), as well as the genes (MURF1, MURF5 and G3) with unknown function. Most genes are actively transcribed and some mRNAs are edited. Fully edited mRNAs for A6 and G3 were abundant, while edited ND7 transcripts were rare, and only partially edited and pre-edited transcripts for ND8 were detected. The data show that the mitochondrial genome of P. serpens is functional, although its functions may be limited to expressing the ATPase and, possibly, NADH dehydrogenase complexes. PMID:10340485

  16. Maintenance and Integrity of the Mitochondrial Genome: a Plethora of Nuclear Genes in the Budding Yeast

    PubMed Central

    Contamine, Véronique; Picard, Marguerite

    2000-01-01

    Instability of the mitochondrial genome (mtDNA) is a general problem from yeasts to humans. However, its genetic control is not well documented except in the yeast Saccharomyces cerevisiae. From the discovery, 50 years ago, of the petite mutants by Ephrussi and his coworkers, it has been shown that more than 100 nuclear genes directly or indirectly influence the fate of the rho+ mtDNA. It is not surprising that mutations in genes involved in mtDNA metabolism (replication, repair, and recombination) can cause a complete loss of mtDNA (rho0 petites) and/or lead to truncated forms (rho−) of this genome. However, most loss-of-function mutations which increase yeast mtDNA instability act indirectly: they lie in genes controlling functions as diverse as mitochondrial translation, ATP synthase, iron homeostasis, fatty acid metabolism, mitochondrial morphology, and so on. In a few cases it has been shown that gene overexpression increases the levels of petite mutants. Mutations in other genes are lethal in the absence of a functional mtDNA and thus convert this petite-positive yeast into a petite-negative form: petite cells cannot be recovered in these genetic contexts. Most of the data are explained if one assumes that the maintenance of the rho+ genome depends on a centromere-like structure dispensable for the maintenance of rho− mtDNA and/or the function of mitochondrially encoded ATP synthase subunits, especially ATP6. In fact, the real challenge for the next 50 years will be to assemble the pieces of this puzzle by using yeast and to use complementary models, especially in strict aerobes. PMID:10839818

  17. Multiple Origins of Eukaryotic cox15 Suggest Horizontal Gene Transfer from Bacteria to Jakobid Mitochondrial DNA.

    PubMed

    He, Ding; Fu, Cheng-Jie; Baldauf, Sandra L

    2016-01-01

    The most gene-rich and bacterial-like mitochondrial genomes known are those of Jakobida (Excavata). Of these, the most extreme example to date is the Andalucia godoyi mitochondrial DNA (mtDNA), including a cox15 gene encoding the respiratory enzyme heme A synthase (HAS), which is nuclear-encoded in nearly all other mitochondriate eukaryotes. Thus cox15 in eukaryotes appears to be a classic example of mitochondrion-to-nucleus (endosymbiotic) gene transfer, with A. godoyi uniquely retaining the ancestral state. However, our analyses reveal two highly distinct HAS types (encoded by cox15-1 and cox15-2 genes) and identify A. godoyi mitochondrial cox15-encoded HAS as type-1 and all other eukaryotic cox15-encoded HAS as type-2. Molecular phylogeny places the two HAS types in widely separated clades with eukaryotic type-2 HAS clustering with the bulk of α-proteobacteria (>670 sequences), whereas A. godoyi type-1 HAS clusters with an eclectic set of bacteria and archaea including two α-proteobacteria missing from the type-2 clade. This wide phylogenetic separation of the two HAS types is reinforced by unique features of their predicted protein structures. Meanwhile, RNA-sequencing and genomic analyses fail to detect either cox15 type in the nuclear genome of any jakobid including A. godoyi. This suggests that not only is cox15-1 a relatively recent acquisition unique to the Andalucia lineage but also the jakobid last common ancestor probably lacked both cox15 types. These results indicate that uptake of foreign genes by mtDNA is more taxonomically widespread than previously thought. They also caution against the assumption that all α-proteobacterial-like features of eukaryotes are ancient remnants of endosymbiosis. PMID:26412445

  18. Mitochondrial DNA mutation-elicited oxidative stress, oxidative damage, and altered gene expression in cultured cells of patients with MERRF syndrome.

    PubMed

    Wu, Shi-Bei; Ma, Yi-Shing; Wu, Yu-Ting; Chen, Yin-Chiu; Wei, Yau-Huei

    2010-06-01

    Myoclonic epilepsy and ragged-red fibers (MERRF) syndrome is a rare disorder characterized by myoclonus, muscle weakness, cerebellar ataxia, heart conduction block, and dementia. It has been documented that 80-90% of the patients with MERRF syndrome are caused by the A8344G mutation in the tRNA(Lys) gene of mitochondrial DNA (mtDNA). We and other investigators have reported that the mtDNA mutation results in not only inefficient generation of adenosine triphosphate but also increased production of reactive oxygen species (ROS) in cultured cells harboring A8344G mutation of mtDNA. In addition, we found an imbalance in the gene expression of antioxidant enzymes in the skin fibroblasts of MERRF patients. The mRNA, protein, and enzyme activity levels of manganese-superoxide dismutase were increased, but those of Cu,Zn-SOD, catalase, and glutathione peroxidase did not show significant changes. Recently, we showed that the excess ROS could damage voltage-dependent anion channel, prohibitin, Lon protease, and aconitase in the MERRF cells. Moreover, there was a dramatic increase in the gene expression and activity of matrix metalloproteinase 1, which may contribute to the cytoskeleton remodeling involved in the weakness and atrophy of muscle commonly seen in MERRF patients. Taken together, we suggest that mtDNA mutation-elicited oxidative stress, oxidative damage, and altered gene expression are involved in the pathogenesis and progression of MERRF syndrome. PMID:20411357

  19. Mitochondrial-related gene expression changes are sensitive to agonal-pH state: implications for brain disorders

    PubMed Central

    Vawter, MP; Tomita, H; Meng, F; Bolstad, B; Li, J; Evans, S; Choudary, P; Atz, M; Shao, L; Neal, C; Walsh, DM; Burmeister, M; Speed, T; Myers, R; Jones, EG; Watson, SJ; Akil, H; Bunney, WE

    2010-01-01

    Mitochondrial defects in gene expression have been implicated in the pathophysiology of bipolar disorder and schizophrenia. We have now contrasted control brains with low pH versus high pH and showed that 28% of genes in mitochondrial-related pathways meet criteria for differential expression. A majority of genes in the mitochondrial, chaperone and proteasome pathways of nuclear DNA-encoded gene expression were decreased with decreased brain pH, whereas a majority of genes in the apoptotic and reactive oxygen stress pathways showed an increased gene expression with a decreased brain pH. There was a significant increase in mitochondrial DNA copy number and mitochondrial DNA gene expression with increased agonal duration. To minimize effects of agonal-pH state on mood disorder comparisons, two classic approaches were used, removing all subjects with low pH and agonal factors from analysis, or grouping low and high pH as a separate variable. Three groups of potential candidate genes emerged that may be mood disorder related: (a) genes that showed no sensitivity to pH but were differentially expressed in bipolar disorder or major depressive disorder; (b) genes that were altered by agonal-pH in one direction but altered in mood disorder in the opposite direction to agonal-pH and (c) genes with agonal-pH sensitivity that displayed the same direction of changes in mood disorder. Genes from these categories such as NR4A1 and HSPA2 were confirmed with Q-PCR. The interpretation of postmortem brain studies involving broad mitochondrial gene expression and related pathway alterations must be monitored against the strong effect of agonal-pH state. Genes with the least sensitivity to agonal-pH could present a starting point for candidate gene search in neuropsychiatric disorders. PMID:16636682

  20. Mitochondrial-related gene expression changes are sensitive to agonal-pH state: implications for brain disorders.

    PubMed

    Vawter, M P; Tomita, H; Meng, F; Bolstad, B; Li, J; Evans, S; Choudary, P; Atz, M; Shao, L; Neal, C; Walsh, D M; Burmeister, M; Speed, T; Myers, R; Jones, E G; Watson, S J; Akil, H; Bunney, W E

    2006-07-01

    Mitochondrial defects in gene expression have been implicated in the pathophysiology of bipolar disorder and schizophrenia. We have now contrasted control brains with low pH versus high pH and showed that 28% of genes in mitochondrial-related pathways meet criteria for differential expression. A majority of genes in the mitochondrial, chaperone and proteasome pathways of nuclear DNA-encoded gene expression were decreased with decreased brain pH, whereas a majority of genes in the apoptotic and reactive oxygen stress pathways showed an increased gene expression with a decreased brain pH. There was a significant increase in mitochondrial DNA copy number and mitochondrial DNA gene expression with increased agonal duration. To minimize effects of agonal-pH state on mood disorder comparisons, two classic approaches were used, removing all subjects with low pH and agonal factors from analysis, or grouping low and high pH as a separate variable. Three groups of potential candidate genes emerged that may be mood disorder related: (a) genes that showed no sensitivity to pH but were differentially expressed in bipolar disorder or major depressive disorder; (b) genes that were altered by agonal-pH in one direction but altered in mood disorder in the opposite direction to agonal-pH and (c) genes with agonal-pH sensitivity that displayed the same direction of changes in mood disorder. Genes from these categories such as NR4A1 and HSPA2 were confirmed with Q-PCR. The interpretation of postmortem brain studies involving broad mitochondrial gene expression and related pathway alterations must be monitored against the strong effect of agonal-pH state. Genes with the least sensitivity to agonal-pH could present a starting point for candidate gene search in neuropsychiatric disorders. PMID:16636682

  1. Host Generated siRNAs Attenuate Expression of Serine Protease Gene in Myzus persicae

    PubMed Central

    Bhatia, Varnika; Bhattacharya, Ramcharan; Uniyal, Prem L.; Singh, Rajendra; Niranjan, Rampal S.

    2012-01-01

    Background Sap sucking hemipteran aphids damage diverse crop species. Although delivery of ds-RNA or siRNA through microinjection/feeding has been demonstrated, the efficacy of host-mediated delivery of aphid-specific dsRNA in developing aphid resistance has been far from being elucidated. Methodology/Principal Findings Transgenic Arabidopsis expressing ds-RNA of Myzus persicae serine protease (MySP) was developed that triggered the generation of corresponding siRNAs amenable for delivery to the feeding aphids. M. persicae when fed on the transgenic plants for different time intervals under controlled growth conditions resulted in a significant attenuation of the expression of MySP and a commensurate decline in gut protease activity. Although the survivability of these aphids was not affected, there was a noticeable decline in their fecundity resulting in a significant reduction in parthenogenetic population. Conclusions/Significance The study highlighted the feasibility of developing host based RNAi-mediated resistance against hemipteran pest aphids. PMID:23071558

  2. Tomato transgenic plants expressing hairpin construct of a nematode protease gene conferred enhanced resistance to root-knot nematodes

    PubMed Central

    Dutta, Tushar K.; Papolu, Pradeep K.; Banakar, Prakash; Choudhary, Divya; Sirohi, Anil; Rao, Uma

    2015-01-01

    Root-knot nematodes (Meloidogyne incognita) cause substantial yield losses in vegetables worldwide, and are difficult to manage. Continuous withdrawal of environmentally-harmful nematicides from the global market warrants the need for novel nematode management strategies. Utility of host-delivered RNAi has been demonstrated in several plants (Arabidopsis, tobacco, and soybean) that exhibited resistance against root-knot and cyst nematodes. Herein, a M. incognita-specific protease gene, cathepsin L cysteine proteinase (Mi-cpl-1), was targeted to generate tomato transgenic lines to evaluate the genetically modified nematode resistance. In vitro knockdown of Mi-cpl-1 gene led to the reduced attraction and penetration of M. incognita in tomato, suggesting the involvement of Mi-cpl-1 in nematode parasitism. Transgenic expression of the RNAi construct of Mi-cpl-1 gene resulted in 60–80% reduction in infection and multiplication of M. incognita in tomato. Evidence for in vitro and in vivo silencing of Mi-cpl-1 was confirmed by expression analysis using quantitative PCR. Our study demonstrates that Mi-cpl-1 plays crucial role during plant-nematode interaction and plant-mediated downregulation of this gene elicits detrimental effect on M. incognita development, reinforcing the potential of RNAi technology for management of phytonematodes in crop plants. PMID:25883594

  3. Resolution of the African hominoid trichotomy by use of a mitochondrial gene sequence

    SciTech Connect

    Ruvolo, M.; Disotell, T.R.; Allard, M.W. ); Brown, W.M. ); Honeycutt, R.L. )

    1991-02-15

    Mitochondrial DNA sequences encoding the cytochrome oxidase subunit II gene have been determined for five primate species, siamang (Hylobates syndactylus), lowland gorilla (Gorilla gorilla), pygmy chimpanzee (Pan paniscus), crab-eating macaque (Macaca fascicularis), and green monkey (Cercopithecus aethiops), and compared with published sequences of other primate and nonprimate species. Comparisons of cytochrome oxidase subunit II gene sequences provide clear-cut evidence from the mitochondrial genome for the separation of the African ape trichotomy into two evolutionary lineages, one leading to gorillas and the other to humans and chimpanzees. Several different tree-building methods support this same phylogenetic tree topology. The comparisons also yield trees in which a substantial length separates the divergence point of gorillas from that of humans and chimpanzees, suggesting that the lineage most immediately ancestral to humans and chimpanzees may have been in existence for a relatively long time.

  4. Multiple Conserved Heteroplasmic Sites in tRNA Genes in the Mitochondrial Genomes of Terrestrial Isopods (Oniscidea).

    PubMed

    Chandler, Christopher H; Badawi, Myriam; Moumen, Bouziane; Grève, Pierre; Cordaux, Richard

    2015-07-01

    Mitochondrial genome structure and organization are relatively conserved among metazoans. However, in many isopods, especially the terrestrial isopods (Oniscidea), the mitochondrial genome consists of both ∼14-kb linear monomers and ∼28-kb circular dimers. This unusual organization is associated with an ancient and conserved constitutive heteroplasmic site. This heteroplasmy affects the anticodon of a tRNA gene, allowing this single locus to function as a "dual" tRNA gene for two different amino acids. Here, we further explore the evolution of these unusual mitochondrial genomes by assembling complete mitochondrial sequences for two additional Oniscidean species, Trachelipus rathkei and Cylisticus convexus. Strikingly, we find evidence of two additional heteroplasmic sites that also alter tRNA anticodons, creating additional dual tRNA genes, and that are conserved across both species. These results suggest that the unique linear/circular organization of isopods' mitochondrial genomes may facilitate the evolution of stable mitochondrial heteroplasmies, and, conversely, once such heteroplasmies have evolved, they constrain the multimeric structure of the mitochondrial genome in these species. Finally, we outline some possible future research directions to identify the factors influencing mitochondrial genome evolution in this group. PMID:25911226

  5. Multiple Conserved Heteroplasmic Sites in tRNA Genes in the Mitochondrial Genomes of Terrestrial Isopods (Oniscidea)

    PubMed Central

    Chandler, Christopher H.; Badawi, Myriam; Moumen, Bouziane; Grève, Pierre; Cordaux, Richard

    2015-01-01

    Mitochondrial genome structure and organization are relatively conserved among metazoans. However, in many isopods, especially the terrestrial isopods (Oniscidea), the mitochondrial genome consists of both ∼14-kb linear monomers and ∼28-kb circular dimers. This unusual organization is associated with an ancient and conserved constitutive heteroplasmic site. This heteroplasmy affects the anticodon of a tRNA gene, allowing this single locus to function as a “dual” tRNA gene for two different amino acids. Here, we further explore the evolution of these unusual mitochondrial genomes by assembling complete mitochondrial sequences for two additional Oniscidean species, Trachelipus rathkei and Cylisticus convexus. Strikingly, we find evidence of two additional heteroplasmic sites that also alter tRNA anticodons, creating additional dual tRNA genes, and that are conserved across both species. These results suggest that the unique linear/circular organization of isopods’ mitochondrial genomes may facilitate the evolution of stable mitochondrial heteroplasmies, and, conversely, once such heteroplasmies have evolved, they constrain the multimeric structure of the mitochondrial genome in these species. Finally, we outline some possible future research directions to identify the factors influencing mitochondrial genome evolution in this group. PMID:25911226

  6. Genome-wide identification and immune response analysis of serine protease inhibitor genes in the silkworm, Bombyx mori.

    PubMed

    Zhao, Ping; Dong, Zhaoming; Duan, Jun; Wang, Genhong; Wang, Lingyan; Li, Youshan; Xiang, Zhonghuai; Xia, Qingyou

    2012-01-01

    In most insect species, a variety of serine protease inhibitors (SPIs) have been found in multiple tissues, including integument, gonad, salivary gland, and hemolymph, and are required for preventing unwanted proteolysis. These SPIs belong to different families and have distinct inhibitory mechanisms. Herein, we predicted and characterized potential SPI genes based on the genome sequences of silkworm, Bombyx mori. As a result, a total of eighty SPI genes were identified in B. mori. These SPI genes contain 10 kinds of SPI domains, including serpin, Kunitz_BPTI, Kazal, TIL, amfpi, Bowman-Birk, Antistasin, WAP, Pacifastin, and alpha-macroglobulin. Sixty-three SPIs contain single SPI domain while the others have at least two inhibitor units. Some SPIs also contain non-inhibitor domains for protein-protein interactions, including EGF, ADAM_spacer, spondin_N, reeler, TSP_1 and other modules. Microarray analysis showed that fourteen SPI genes from lineage-specific TIL family and Group F of serpin family had enriched expression in the silk gland. The roles of SPIs in resisting pathogens were investigated in silkworms when they were infected by four pathogens. Microarray and qRT-PCR experiments revealed obvious up-regulation of 8, 4, 3 and 3 SPI genes after infection with Escherichia coli, Bacillus bombysepticus, Beauveria bassiana or B. mori nuclear polyhedrosis virus (BmNPV), respectively. On the contrary, 4, 11, 7 and 9 SPI genes were down-regulated after infection with E. coli, B. bombysepticus, B. bassiana or BmNPV, respectively. These results suggested that these SPI genes may be involved in resistance to pathogenic microorganisms. These findings may provide valuable information for further clarifying the roles of SPIs in the development, immune defence, and efficient synthesis of silk gland protein. PMID:22348050

  7. MitoNuc: a database of nuclear genes coding for mitochondrial proteins. Update 2002.

    PubMed

    Attimonelli, Marcella; Catalano, Domenico; Gissi, Carmela; Grillo, Giorgio; Licciulli, Flavio; Liuni, Sabino; Santamaria, Monica; Pesole, Graziano; Saccone, Cecilia

    2002-01-01

    Mitochondria, besides their central role in energy metabolism, have recently been found to be involved in a number of basic processes of cell life and to contribute to the pathogenesis of many degenerative diseases. All functions of mitochondria depend on the interaction of nuclear and organelle genomes. Mitochondrial genomes have been extensively sequenced and analysed and data have been collected in several specialised databases. In order to collect information on nuclear coded mitochondrial proteins we developed MitoNuc, a database containing detailed information on sequenced nuclear genes coding for mitochondrial proteins in Metazoa. The MitoNuc database can be retrieved through SRS and is available via the web site http://bighost.area.ba.cnr.it/mitochondriome where other mitochondrial databases developed by our group, the complete list of the sequenced mitochondrial genomes, links to other mitochondrial sites and related information, are available. The MitoAln database, related to MitoNuc in the previous release, reporting the multiple alignments of the relevant homologous protein coding regions, is no longer supported in the present release. In order to keep the links among entries in MitoNuc from homologous proteins, a new field in the database has been defined: the cluster identifier, an alpha numeric code used to identify each cluster of homologous proteins. A comment field derived from the corresponding SWISS-PROT entry has been introduced; this reports clinical data related to dysfunction of the protein. The logic scheme of MitoNuc database has been implemented in the ORACLE DBMS. This will allow the end-users to retrieve data through a friendly interface that will be soon implemented. PMID:11752284

  8. MitoNuc: a database of nuclear genes coding for mitochondrial proteins. Update 2002

    PubMed Central

    Attimonelli, Marcella; Catalano, Domenico; Gissi, Carmela; Grillo, Giorgio; Licciulli, Flavio; Liuni, Sabino; Santamaria, Monica; Pesole, Graziano; Saccone, Cecilia

    2002-01-01

    Mitochondria, besides their central role in energy metabolism, have recently been found to be involved in a number of basic processes of cell life and to contribute to the pathogenesis of many degenerative diseases. All functions of mitochondria depend on the interaction of nuclear and organelle genomes. Mitochondrial genomes have been extensively sequenced and analysed and data have been collected in several specialised databases. In order to collect information on nuclear coded mitochondrial proteins we developed MitoNuc, a database containing detailed information on sequenced nuclear genes coding for mitochondrial proteins in Metazoa. The MitoNuc database can be retrieved through SRS and is available via the web site http://bighost.area.ba.cnr.it/mitochondriome where other mitochondrial databases developed by our group, the complete list of the sequenced mitochondrial genomes, links to other mitochondrial sites and related information, are available. The MitoAln database, related to MitoNuc in the previous release, reporting the multiple alignments of the relevant homologous protein coding regions, is no longer supported in the present release. In order to keep the links among entries in MitoNuc from homologous proteins, a new field in the database has been defined: the cluster identifier, an alpha numeric code used to identify each cluster of homologous proteins. A comment field derived from the corresponding SWISS-PROT entry has been introduced; this reports clinical data related to dysfunction of the protein. The logic scheme of MitoNuc database has been implemented in the ORACLE DBMS. This will allow the end-users to retrieve data through a friendly interface that will be soon implemented. PMID:11752284

  9. Timing major conflict between mitochondrial and nuclear genes in species relationships of Polygonia butterflies (Nymphalidae: Nymphalini)

    PubMed Central

    Wahlberg, Niklas; Weingartner, Elisabet; Warren, Andrew D; Nylin, Sören

    2009-01-01

    Background Major conflict between mitochondrial and nuclear genes in estimating species relationships is an increasingly common finding in animals. Usually this is attributed to incomplete lineage sorting, but recently the possibility has been raised that hybridization is important in generating such phylogenetic patterns. Just how widespread ancient and/or recent hybridization is in animals and how it affects estimates of species relationships is still not well-known. Results We investigate the species relationships and their evolutionary history over time in the genus Polygonia using DNA sequences from two mitochondrial gene regions (COI and ND1, total 1931 bp) and four nuclear gene regions (EF-1α, wingless, GAPDH and RpS5, total 2948 bp). We found clear, strongly supported conflict between mitochondrial and nuclear DNA sequences in estimating species relationships in the genus Polygonia. Nodes at which there was no conflict tended to have diverged at the same time when analyzed separately, while nodes at which conflict was present diverged at different times. We find that two species create most of the conflict, and attribute the conflict found in Polygonia satyrus to ancient hybridization and conflict found in Polygonia oreas to recent or ongoing hybridization. In both examples, the nuclear gene regions tended to give the phylogenetic relationships of the species supported by morphology and biology. Conclusion Studies inferring species-level relationships using molecular data should never be based on a single locus. Here we show that the phylogenetic hypothesis generated using mitochondrial DNA gives a very different interpretation of the evolutionary history of Polygonia species compared to that generated from nuclear DNA. We show that possible cases of hybridization in Polygonia are not limited to sister species, but may be inferred further back in time. Furthermore, we provide more evidence that Haldane's effect might not be as strong a process in

  10. A FastA based compilation of higher plant mitochondrial tRNA genes.

    PubMed Central

    Sagliano, A; Volpicella, M; Gallerani, R; Ceci, L R

    1998-01-01

    A new version of the compilation of higher plant mitochondrial tRNA genes (http://www.ebi.ac.uk/service ) has been obtained by means of the FastA program for similarity searching in nucleotide sequence Databases. This approach improves the previous collection, which was based on literature data analysis. The current compilation contains 158 sequences with an increase of 43 units. In this paper, some interesting features of the new entries are briefly presented. PMID:9399821