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Mitochondrial DNA (mtDNA) mutations are maternally inherited and are associated with a broad range of debilitating and fatal diseases (1). Reproductive technologies designed to uncouple the inheritance of mtDNA from nuclear DNA may enable affected women to have a genetically related child with a greatly reduced risk of mtDNA disease. Here we report the first preclinical studies on pronuclear transplantation (PNT). Surprisingly, techniques used in proof-of-concept studies involving abnormally fertilized human zygotes (2) were not well tolerated by normally fertilized zygotes. We have therefore developed an alternative approach based on transplanting pronuclei shortly after completion of meiosis rather than shortly before the first mitotic division. This promotes efficient development to the blastocyst stage with no detectable effect on aneuploidy or gene expression. After optimization, mtDNA carryover was reduced to, Predicting the risk of serious disease in children of women who carry mtDNA mutations is complicated by a number of factors. Mutations in mtDNA can be either homoplasmic (all copies [...]

Mitochondrial diseases are commonly caused by mutated mitochondrial DNA (mtDNA), which in most cases coexists with wild-type mtDNA, resulting in mtDNA heteroplasmy. We have engineered transcription activator-like effector nucleases (TALENs) [...]

8-Oxoguanine (8-oxoG), a common DNA lesion caused by reactive oxygen species, is associated with carcinogenesis and neurodegeneration. Although the mechanism by which 8-oxoG causes carcinogenesis is well understood, the mechanism by which it causes neurodegeneration is unknown. Here, we report that neurode-generation is triggered by MUTYH-mediated excision repair of 8-oxoG-paired adenine. Mutant mice lacking 8-oxo-2'-deoxyguanosine triphosphate-depleting (8-oxo-dGTP-depleting) MTH1 and/or 8-oxoG-excising, OGG1 exhibited severe striatal neurodegeneration, whereas mutant mice lacking MUTYH or OGG1/MUTYH were resistant to neurodegeneration under conditions of oxidative stress. These results indicate that OGG1 and MTH1 are protective, [...]

Mitochondrial respiratory function is frequently impaired in human Cancers (1-4). However, the mechanisms by which mitochondrial dysfunction contributes to tumour progression remain elusive. Here we show in Drosophila imaginal epithelium that defects in mitochondrial function potently induce tumour progression of surrounding tissue in conjunction with oncogenic Ras. Our data show that Ras activation and mitochondrial dysfunction cooperatively stimulate production of reactive oxygen species, which causes activation of c-Jun amino (N)-terminal kinase (JNK) signalling. JNK cooperates with oncogenic Ras to inactivate the Hippo pathway, leading to upregulation of its targets Unpaired (an interleukin-6 homologue) and Wingless (a Wnt homologue). Mitochondrial dysfunction in Ras-activated cells further cooperates with Ras signalling in neighbouring cells with normal mitochondrial function, causing benign tumours to exhibit metastatic behaviour. Our findings provide a mechanistic basis for interclonal tumour progression driven by mitochondrial dysfunction and oncogenic Ras., Cell-cell interactions between oncogenic cells and surrounding normal cells in the tumour microenvironment play crucial roles in cancer progression (5). To study the genetic basis of tumour progression through cell-cell [...]

Abstract As part of the life sciences series, this article describes the role of deoxyribonucleic acid and ribonucleic acid, genes and chromosomes. The processes of mitosis and meiosis are discussed [...]

BACKGROUND: The incidence of asbestos-induced human cancers is increasing worldwide, and considerable evidence suggests that reactive oxygen species (ROS) are important mediators of these diseases. Our previous studies suggested that [...]

ABSTRACT Biological and ecological research is seriously handicapped because of difficulties experienced in the reliable detection and quantification of bivalve larvae. This is a critical issue in the case of [...]

Spermatozoon of Atlantic halibut (Hippoglossus hippoglossus (L., 1758)) is uniflagellated, lacks an acrosome, and is differentiated into a head, midpiece, and flagellum. There are two to five mitochondria in the midpiece, as well as proximal and distal centrioles. The flagellum consisted of 9 + 2 microtubules surrounded by plasma membrane, which is extended at the proximal part of the flagellum owing to the presence of vacuoles. After sperm activation in seawater, sperm motility and velocity decreased from 98.4% ± 3.4% and 170.3 ± 8.9 µm x [s.sup.-1] at 15 s after sperm activation to 4.8% ± 4.7% and 9.2 ± 8.9 µm x [s.sup.-1] at 120 s after sperm activation, respectively. ATP content (nmol x [L.sup.-1] ATP per [10.sup.8] spermatozoa) significantly decreased at 60 s after sperm activation (5.9 ± 1.5) compared with at 0 and 30 s after sperm activation (14.9 ± 1.5 and 14.5 ± 1.5, respectively). Beating waves propagated along the full length of the flagellum after sperm activation, whereas waves were restricted to the proximal section during the latter motility period. Wave amplitude significantly decreased at 45 s after sperm activation, but wavelength did not differ. The present study showed associations among sperm morphology, ATP content, flagellar wave parameters, and sperm velocity, which could be used in comparative spermatology. Le spermatozoide du fletan atlantique (Hippoglossus hippoglossus (L., 1758)) est uniflagelle, sans acrosome et differencie en tete, corps median et flagelle. Il y a deux a cinq mitochondries dans le corps median, ainsi que des centrioles proximaux et distaux. Le flagelle consiste en 9 + 2 microtubules entoures d'une membrane plasmatique qui se prolonge dans la partie proximale du flagelle a cause de la presence de vacuoles. Apres l'activation des spermatozoides dans l'eau de mer, la motilite et la vitesse des spermatozoides diminuent respectivement de 98,4 % ± 3,4 % et de 170,3 ± 8,9 µm x [s.sub.-1] a 15 s apres l'activation a 4,8 % ± 4,7 % et 9,2 ± 8,9 µm x [s.sup.-1] a 120 s apres l'activation. Le contenu en ATP (nmol x [L.sub.-1] ATP par 108 spermatozoides) diminue significativement 60 s apres l'activation (5,9 ± 1,5) par comparaison a 0 et 30 s apres l'activation (respectivement 14,9 ± 1,5 et 14,5 ± 1,5). Les ondulations de battement se propagent sur toute la longueur du flagelle apres l'activation, mais sont restreintes a la partie proximale durant la periode de motilite; subsequente. L'amplitude des ondulations diminue significativement a 45 s apres l'activation, mais leur longueur d'onde ne change pas. Notre etude met en lumiere des associations entre la morphologie des spermatozoides, leur contenu en ATP, les variables ondulatoires des flagelles et la vitesse des spermatozoides qui pourraient servir en spermatologie comparee. [Traduit par la Redaction], Introduction Morphology, motility, and seminal plasma composition are considered major elements in the study of comparative fish spermatology (Jamieson 1991; Ishijima et al. 1998; Alavi et al. 2008a). Fish spermatozoa [...]

Myocardial infarction is a leading cause of mortality worldwide. Here we report that modulation of microRNA-499 (miR-499) levels affects apoptosis and the severity of myocardial infarction and cardiac dysfunction induced by ischemia-reperfusion. We found that both the α- and β-isoforms of the calcineurin catalytic subunit are direct targets of miR-499 and that miR-499 inhibits cardiomyocyte apoptosis through its suppression of calcineurin-mediated dephosphorylation of dynamin-related protein-1 (Drp1), thereby decreasing Drp1 accumulation in mitochondria and Drp1-mediated activation of the mitochondrial fission program. We also found that p53 transcriptionally downregulates miR-499 expression. Our data reveal a role for miR-499 in regulating the mitochondrial fission machinery and we suggest that modulation of miR-499 levels may provide a therapeutic approach for treating myocardial infarction., MicroRNAs are about 22 nucleotides long and act as negative regulators of gene expression by inhibiting mRNA translation or promoting mRNA degradation (1). A single miRNA can modulate complex physiological [...]

Although the proteins BAX and BAK are required for initiation of apoptosis at the mitochondria, how BAX and BAK are activated remains unsettled. We provide in viva evidence demonstrating an essential role of the proteins BID, BIM, and PUMA in activating BAX and BAK. Bid, Bim, and Puma triple-knockout mice showed the same developmental defects that are associated with deficiency of Bax and Bak, including persistent interdigital webs and imperforate vaginas. Genetic deletion of Bid, Bim, and Puma prevented the homo-oligomerization of BAX and BAK, and thereby cytochrome c--mediated activation of caspases in response to diverse death signals in neurons and T lymphocytes, despite the presence of other BH3-only molecules. Thus, many forms of apoptosis require direct activation of BAX and BAK at the mitochondria by a member of the BID, BIM, or PUMA ramify of proteins. 10.1126/science.1190217

At present, there are few means to track symptomatic stages of CNS aging. Thus, although metabolic changes are implicated in mtDNA mutation-driven aging, the manifestations remain unclear. Here, we used normally aging and prematurely aging mtDNA mutator mice to establish a molecular link between mitochondrial dysfunction and abnormal metabolism in the aging process. Using proton magnetic resonance spectroscopy and HPLC, we found that brain lactate levels were increased twofold in both normally and prematurely aging mice during aging. To correlate the striking increase in lactate with tissue pathology, we investigated the respiratory chain enzymes and detected mitochondrial failure in key brain areas from both normally and prematurely aging mice. We used in situ hybridization to show that increased brain lactate levels were caused by a shift in transcriptional activities of the lactate dehydrogenases to promote pyruvate to lactate conversion. Separation of the five tetrameric lactate dehydrogenase (LDH) isoenzymes revealed an increase of those dominated by the Ldh-A product and a decrease of those rich in the Ldh-B product, which, in turn, increases pyruvate to lactate conversion. Spectrophotometric assays measuring LDH activity from the pyruvate and lactate sides of the reaction showed a higher pyruvate [right arrow] lactate activity in the brain. We argue for the use of lactate proton magnetic resonance spectroscopy as a noninvasive strategy for monitoring this hallmark of the aging process. The mtDNA mutator mouse allows us to conclude that the increased LDH-A/LDH-B ratio causes high brain lactate levels, which, in turn, are predictive of aging phenotypes. mtDNA mutator mouse | proton magnetic resonance spectroscopy | in situ hybridization | COX/SDH enzyme histochemistry | HPLC doi/ 10.1073/pnas.1008189107

Human tyrosyl-DNA phosphodiesterase (TDP1) hydrolyzes the phosphodiester bond at a DNA 3'-end linked to a tyrosyl moiety and has been implicated in the repair of topoisomerase I (Top1)-DNA covalent complexes. TDP1 can also hydrolyze other 3'-end DNA alterations including 3'-phosphoglycolate and 3'-abasic sites, and exhibits 3'-nucleosidase activity indicating it may function as a general 3'-end-processing DNA repair enzyme. Here, using laser confocal microscopy, subcellular fractionation and biochemical analyses we demonstrate that a fraction of the TDP1 encoded by the nuclear TDP1 gene localizes to mitochondria. We also show that mitochondrial base excision repair depends on TDP1 activity and provide evidence that TDP1 is required for efficient repair of oxidative damage in mitochondrial DNA. Together, our findings provide evidence for TDP1 as a novel mitochondrial enzyme. DNA repair | ligase III | oxidative DNA damage | topoisomerase I | mitochondrial BER doi/ 10.1073/pnas.1009814107

Disrupted-in-schizophrenia 1 (DISC1) has emerged as a schizophrenia-susceptibility gene affecting various neuronal functions. In this study, we characterized Mitofilin, a mitochondrial inner membrane protein, as a mediator of the mitochondrial function of DISC1. A fraction of DISC1 was localized to the inside of mitochondria and directly interacts with Mitofilin. A reduction in DISC1 function induced mitochondrial dysfunction, evidenced by decreased mitochondrial NADH dehydrogenase activities, reduced cellular ATP contents, and perturbed mitochondrial [Ca.sup.2+] dynamics. In addition, deficiencies in DISC1 and Mitofilin induced a reduction in mitochondrial monoamine oxidase-A activity. The mitochondrial dysfunctions evoked by the deficiency of DISC1 were partially pheno-copied by an overexpression of truncated DISC1 that is associated with schizophrenia in human. DISC1 deficiencies induced the ubiquitination of Mitofilin, suggesting that DISC1 is critical for the stability of Mitofilin. Finally, the mitochondrial dysfunction induced by DISC1 deficiency was partially reversed by coexpression of Mitofilin, confirming a functional link between DISC1 and Mitofilin for the normal mitochondrial function. According to these results, we propose that DISC1 plays essential roles for mitochondrial function in collaboration with a mitochondrial interacting partner, Mitofilin. IMMT | mitochondrial dysfunctions | hyperdopaminergia | calcium buffering | psychiatric disorders doi/ 10.1073/pnas.1004361107

The fishing community of Bering Island, located in the Russian Commander Islands off the Kamchatka Peninsula, was originally founded by a small number of Russian soldiers and merchants, along with Aleuts forcibly relocated from the western region of the Aleutian archipelago. The purpose of this study is to characterize the genetic variation of Bering Island inhabitants for autosomal, mitochondrial, and Y-chromosome DNA and classic genetic markers and to investigate the genetic impact of the 19th-century founding and subsequent demographic events on this heterogeneous community. Our results show a loss of diversity among maternal lineages in the Bering Aleut population, with fixation of mtDNA haplogroup D, as revealed by median-joining network analysis and mismatch differences. Conversely, paternal haplotypes exhibit an increase in diversity and the presence of a substantial number of non-Native lineages. Admixture results, based on autosomal STR data, indicate that parental contributions to the mixed Aleut population of Bering are approximately 60% Aleut and 40% Russian. Classic genetic markers show affinities between the Bering Island Aleuts and the other historically founded Aleut communities of St. Paul and St. George in the Pribilof Islands, Alaska. This study demonstrates that the opposing evolutionary forces of genetic drift and gene flow acted on the maternal and paternal lineages, respectively, to shape the genetic structure of the present-day inhabitants of Bering Island. KEY WORDS: ALEUTS, BERING ISLAND, GENETIC DRIFT, ADMIXTURE, Y-CHROMOSOME HAPLOGROUPS, MTDNA HAPLOGROUPS, AUTOSOMAL STRS, CLASSIC GENETIC MARKERS., To date, a number of studies have used classic genetic and/or molecular markers to investigate the genetic and evolutionary consequences of geographic isolation on small populations, often reporting dramatic decreases [...]

We summarize the results of a field and laboratory research program (1999-2006) in the Aleutian Islands on the origins of the inhabitants of the archipelago and the genetic structure of these populations. The Aleuts show closest genetic affinity to the contemporary Siberian Eskimos and Chukchi of Chukotka and differ significantly from the populations of Kamchatka (the terminus of the archipelago) and Alaskan Eskimos. Our findings support the hypothesis that the ancestors of the Aleuts crossed Beringia and expanded westerly into the islands approximately 9,000 years ago. The Monmonier algorithm indicates genetic discontinuity between contemporary Kamchatkan populations and western Aleut populations, suggesting that island hopping from Kamchatka into the western Aleutian Islands was highly unlikely. The primary determinant of the distribution of genes throughout the archipelago is geography. The most intimate relationship exists between the genetics (based on mtDNA sequences and intermatch/mismatch distances) and geographic distances (measured in kilometers). However, the Y-chromosome haplogroup frequencies are not significantly correlated with the geography of the Aleutian Islands. The underlying patterns of precontact genetic structure based on Y-chromosome markers of the Aleut populations is obscured because of the gene flow from Russian male colonizers and Scandinavian and English fishermen. We consider alternative theories about the peopling of the Americas from Siberia. In addition, we attempt a synthesis between archaeological and genetic data for the Aleutian Islands. KEY WORDS: MITOCHONDRIAL DNA SEQUENCES, Y-CHROMOSOME MARKERS, GENETIC DISCONTINUITY, GENETIC STRUCTURE, ALEUTIAN ARCHIPELAGO, SIBERIAN ESKIMOS, ALASKAN ESKIMOS, CHUKCHI, KAMCHATKA, ALEUTS., Until the mid-1980s and the development of molecular methods, the reconstruction of human origins was based primarily on archaeological, morphological, linguistic, and genetic data. The genetic characterizations of populations were [...]

The Aleutian Islands were colonized, perhaps several times, from the Alaskan mainland. Earlier work documented transitions in the relative frequencies of mtDNA haplogroups over time, but little is known about potential source populations for prehistoric Aleut migrants. As part of a pilot investigation, we sequenced the mtDNA first hypervariable region (HVRI) in samples from two archaeological sites on the Alaska Peninsula (the Hot Springs site near Port Moller, Alaska; and samples from a cluster of sites in the Brooks River area near Katmai National Park and Preserve) and one site from Prince William Sound (Mink Island). The sequences revealed not only the mtDNA haplogroups typically found in both ancient and modern Aleut populations (A2 and D2) but also haplogroups B2 and D1 in the Brooks River samples and haplogroup D3 in one Mink Islander. These preliminary results suggest greater mtDNA diversity in prehistoric populations than previously observed and facilitate reconstruction of migration scenarios from the peninsula into the Aleutian archipelago in the past. KEY WORDS: ALEUTIAN ISLANDS, ADNA, HVRI, MTDNA HAPLOGROUPS, ALEUTIAN PREHISTORY., Hrdlicka's (1945) inference that the modern population of the Aleutian Islands derived from a migration of people from mainland Alaska through the Alaska Peninsula who replaced the original inhabitants of [...]

The eastern Aleutian prehistoric archaeological sequence is key for understanding population movements, cultural exchanges, and adaptations to environmental changes over a wide area of the north Pacific and Bering Sea during the Holocene. An important question is, Can the settlement history of the eastern Aleutians be understood as a single continuous tradition lasting some 9,000 years, or were there major population and cultural influxes along with periods of widespread population abandonment? We review the available archaeological evidence with reference to recent mtDNA and nucleic DNA studies of prehistoric and contemporary Arctic and Subarctic populations and conclude that the evidence points to an overall cultural continuity with notable incursions and excursions of people and cultural elements into and out of the eastern Aleutians. KEY WORDS: EASTERN ALEUTIANS, PREHISTORIC ARCHAEOLOGY, EASTERN ALEUTIAN PREHISTORY, ARCTIC SMALL TOOL TRADITION, HOG ISLAND, AMAKNAK ISLAND, MARGARET BAY, UMNAK ISLAND, UNALASKA ISLAND., The eastern Aleutian region begins on the Alaska Peninsula at Port Moller and extends about 750 km westward through the Fox Island group to the Islands of the Four Mountains [...]

The human mitochondrial transcription machinery generates the primers required for initiation of leading-strand DNA replication. According to one model, the 3' end of the primer is defined by transcription termination at conserved sequence block II (CSB II) in the mitochondrial DNA control region. We here demonstrate that this site-specific termination event is caused by G-quadruplex structures formed in nascent RNA upon transcription of CSB II. We also demonstrate that a poly-dT stretch downstream of CSB II has a modest stimulatory effect on the termination efficiency. The mechanism is reminiscent of Rho-independent transcription termination in prokaryotes, with the exception that a G-quadruplex structure replaces the hairpin loop formed in bacterial mRNA during transcription of terminator sequences. mitochondrion | RNA polymerase www.pnas.org/cgi/doi/10.1073/pnas.1006026107

The integral peroxisomal membrane proteins PEX10, PEX2, and PEX12 contain a zinc RING finger close to the C terminus. Loss of function of these peroxins causes embryo lethality at the heart stage in Arabidopsis. Preventing the coordination of [Zn.sup.2+] ions by amino acid substitutions in PEX10, PEX2, and PEX12 and overexpressing the resulting conditional sublethal mutations in WT uncovered additional functions of PEX10. Plants overexpressing [DELTA]Zn-mutant PEX10 display deformed peroxisomal shapes causing diminished contact with chloroplasts and possibly with mitochondria. These changes correlated with impaired metabolite transfer and, at high C[O.sub.2], recoverable defective photorespiration plus dwarfish phenotype. The N-terminal PEX10 domain is critical for peroxisome biogenesis and plant development. A point mutation in the highly conserved TLGEEY motif results in vermiform peroxisome shape without impairing organelle contact. Addition of an N-terminal T7 tag to WT PEX0 resulted in partially recoverable reduced growth and defective inflorescences persisting under high C[O.sub.2]. In contrast, plants overexpressing PEX2-[DELTA]Zn-T7 grow like WT in normal atmosphere, contain normal-shaped peroxisomes, but display impaired peroxisomal matrix protein import. PEX12-[DELTA]Zn-T7 mutants exhibit unimpaired import of matrix protein and normal-shaped peroxisomes when grown in normal atmosphere. During seed germination, glyoxysomes form a reticulum around the lipid bodies for mobilization of storage oil. The formation of this glyoxysomal reticulum seemed to be impaired in PEX10-[DELTA]Zn but not in PEX2-[DELTA]Zn-T7 or PEX12-[DELTA]Zn-T7 plants. Both cytosolic PEX10 domains seem essential for peroxisome structure but differ in metabolic function, suggesting a role for this plant peroxin in addition to the import of matrix protein via ubiquitination of PEX5. Thr Leu Gly Glu Glu Tyr motif in peroxin PEX10 | glyoxysome | metabolomics doi/ 10.1073/pnas.1009174107

The core human mitochondrial transcription apparatus is currently regarded as an obligate three-component system comprising the bacteriophage T7-related mitochondrial RNA polymerase, the rRNA methyltransferase-related transcription factor, h-mtTFB2, and the high mobility group box transcription/DNA-packaging factor, h-mtTFA/TFAM. Using a faithful recombinant human mitochondrial transcription system from Escherichia coli, we demonstrate that specific initiation from the mtDNA promoters, LSP and HSP1, only requires mitochondrial RNA polymerase and h-mtTFB2 in vitro. When h-mtTFA is added to these basal components, LSP exhibits a much lower threshold for activation and a larger amplitude response than HSP1. In addition, when LSP and HSP1 are together on the same transcription template, h-mtTFA-independent transcription from HSP1 and h-mtTFA-dependent transcription from both promoters is enhanced and a higher concentration of h-mtTFA is required to stimulate HSP1. Promoter competition experiments revealed that, in addition to LSP competing transcription components away from HSP1, additional cis-acting signals are involved in these aspects of promoter regulation. Based on these results, we speculate that the human mitochondrial transcription system may have evolved to differentially regulate transcription initiation and transcription-primed mtDNA replication in response to the amount of h-mtTFA associated with nucleoids, which could begin to explain the heterogeneity of nucleoid structure and activity in vivo. Furthermore, this study sheds new light on the evolution of mitochondrial transcription components by showing that the human system is a regulated two-component system in vitro, and thus more akin to that of budding yeast than thought previously. h-mtTFA/TFAM | mtDNA | nucleoid | POLRMT | h-mtTFB2/TFB2M doi/10.1073/pnas.0910581107

We examined the population genetics and phylogenetics of Falcipennis canadensis isleibi, a subspecies of Spruce Grouse from the Alexander Archipelago of southeast Alaska, which was recently given subspecies status on the basis of subtle differences in plumage coloration and its limited distribution on several islands. The taxonomic status of E c. isleibi is particularly consequential, both because little is known about its evolutionary, demographic, and conservation status and because island endemics often face high extinction risks. Samples were collected from central Alaska, British Columbia, and Prince of Wales (POW) and Zarembo (ZAM) islands in the Alexander Archipelago and identified to subspecies using established morphological traits. We sequenced the cytochrome-c oxidase I (COI) subunit of the mitochondrial genome (n = 62) and genotyped each individual at six nuclear microsatellite loci (n = 65). Individuals from POW and ZAM shared a unique mitochondrial haplotype not observed in other populations of other subspecies (F. c.franklinii and F. c. canadensis), whereas haplotypes were shared by individuals identified as franklinii of canadensis. Microsatellite loci revealed significant divergence among all subspecies populations ([[bar.F].sub.ST] = 0.352) as well as divergence between POW and ZAM populations of F. c. isleibi. These data corroborate the morphological classification of F. c. isleibi as a separate subspecies. Spruce Grouse are not managed as a single species by the state of Alaska, but instead as an aggregate with other forest grouse species. Our results indicate that populations of F. c. isleibi warrant special management attention to maintain this distinct evolutionary lineage. Received 16 July 2009, accepted 7 March 2010. Key words: Alexander Archipelago, Falcipennis canadensis, microsatellites, mtDNA, population structure, Spruce Grouse. Examinamos la genetica de poblaciones y la filogeograffa de Falcipennis canadensis isleibi, una subespecie del archipielago Alexander del sudeste de Alaska, a la cual se le confirio recientemente el estatus de subespecie con base en diferencias sutiles en la coloracion del plumaje y en su distribucion restringida en varias islas. El estatus taxonomico de F. c. isleibi es particularmente importante tanto porque se sabe poco sobre su estado evolutivo, demografico y de conservacion, como porque los endemicos insulares estan generalmente sujetos a un alto riesgo de extincion. Recolectamos muestras en Alaska central, British Columbia y en las islas Prince of Wales (POW) y Zarembo (ZAM) en el archipielago de Alexander, e identificamos las muestras hasta el nivel de subespecie utilizando caracteres morfologicos establecidos. Secuenciamos la subunidad I de la citocromo-c oxidasa (COI) del genoma mitocondrial (n = 62) y genotipificamos cada individuo con base en seis loci nucleares microsatelitales (n = 65). Los individuos de POW y ZAM compartieron un unico haplotipo mitocondrial que no fue observado en otras poblaciones de otras subespecies (F. c. franklinii y F. c. canadensis), mientras que individuos identificados comofranklinii o canadensis compartieron haplotipos. Los loci microsatelitales revelaron una divergencia significativa entre todas las poblaciones de subespecies ([[barra.F].sub.ST] = 0.352) y tambien divergencia entre las poblaciones de E c. isleibi de POW y ZAM. Estos datos corroboran la clasificacion morfologica de F. c. isleibi como una subespecie. Falcipennis canadensis no es manejada como una unica especie por el estado de Alaska, sino como un agregado junto con otras especies de bosque del mismo genero. Nuestros resultados sugieren que las poblaciones de F. c. isleibi requieren de una atencion especial en el manejo para mantener ese linaje evolutivo distinto. DOI: 10.1525/auk.2010.09086

Examination of contact zones between closely related species is important for understanding speciation, because the interactions in such zones may change the evolutionary direction of one or both taxa. To expose the composition and dynamics of a contact zone between two dove species, Streptopelia vinacea and S. capicola, we used mitochondrial DNA, amplified fragment-length polymorphism (AFLP) markers, and morphological and color measurements. We combined a field study on contact-zone and parental-species individuals with an examination of [F.sub.1] hybrids reared in captivity. We found that the contact zone is a narrow hybrid zone characterized by a high frequency of hybrids, a lack of clear parental species forms, a high incidence of S. capicola mtDNA, and an AFLP marker distribution more similar to that of S. vinacea. In morphology and color, field hybrids were more similar to S. vinacea and significantly different from S. capicola. [F.sub.1] hybrids were more similar to S. capicola in color. The laboratory data showed that both types of mixed matings produced viable [F.sub.1] offspring. Taken together, the results indicate the existence of a unimodal hybrid zone with asymmetric introgression from S. capicola into S. vinacea. The origin of this hybrid zone is most likely a combination of geographic and behavioral factors and suggests a balance between dispersal and selection. Received 30 July 2009, accepted 28 February 2010. Key words: AFLP, doves, genetics, hybrid zone, morphology, mtDNA, Streptopelia, vocalization. Examinar las zonas de contacto entre especies cercanamente emparentadas es importante para entender la especiacion porque las interacciones en dichas zonas pueden cambiar la trayectoria de uno o los dos taxones. Para dar a conocer la composicion y dinamica de una zona de contacto entre dos especies de palomas, Streptopelia vinacea y S. capicola, empleamos ADN mitocondrial, marcadores de polimorfismos amplificados de fragmentos de longitud variable (AFLP) y medidas de morfologia y color. Combinamos un estudio de campo con individuos de la zona de contacto y de las especies parentales con un examen de hibridos de la [F.sub.1] criados en cautiverio. Encontramos que la zona de contacto es una zona de hibridacion estrecha caracterizada por una alta frecuencia de hibridos, una ausencia de formas claramente parentales, una alta incidencia de ADNmt de S. capicola y una distribucion de marcadores AFLP mas similar a la de S. vinacea. En morfologia y color, los hibridos encontrados en el campo fueron mas similares a S. vinacea y significativamente diferentes de S. capicola. Los hibridos de la [F.sub.1] fueron mas similares en color a S. capicola. Los datos de laboratorio mostraron que ambos tipos de apareamientos mixtos produjeron crias [F.sub.1] viables. En conjunto, los resultados indican la existencia de una zona de hibridacion unimodal con introgresion asimetrica de S. capicola hacia S. vinacea. El origen de esta zona de hibridacion probablemente se relaciona con una combinacion de factores geograficos y de comportamiento, y sugiere un balance entre la dispersion y la seleccion. DOI: 10.1525/auk.2010.10002

Mitochondrial genomes with deleterious mutations can replicate in cells along with wild-type genomes in a state of heteroplasmy, and are a cause of severe inherited syndromes, such as mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke (MELAS), neuropathy, ataxia, retinitis pigmentosa-maternally inherited Leigh syndrome (NARP-MILS), and Leber's hereditary optic neuropathy (LHON). The cytosolic E3 ligase, Parkin, commonly mutated in recessive familial parkinsonism, translocates to depolarized mitochondria and induces their autophagic elimination, suggesting that Parkin may signal the selective removal of defective mitochondria within the cell. We report that long-term overexpression of Parkin can eliminate mitochondria with deleterious COXI mutations in heteroplasmic cybrid cells, thereby enriching cells for wild-type mtDNA and restoring cytochrome c oxidase activity. After relieving cybrid cells of Parkin overexpression, a more favorable wild-type to mutant mitochondrial genome ratio is stably maintained. These data support the model that Parkin functions in a mitochondrial quality control pathway. Additionally, they suggest that transiently increasing levels of Parkin expression might ameliorate certain mitochondrial diseases. autophagy | neurodegeneration | Parkinson disease | PINK1 | mitochondria doi/ 10.1073/pnas.0914569107

An insertion polymorphism of the angiotensin-I converting enzyme gene (ACE) is common in humans and the higher expressing allele is associated with an increased risk of diabetic complications. The ACE polymorphism does not significantly affect blood pressure or angiotensin II levels, suggesting that the kallikrein-kinin system partly mediates the effects of the polymorphism. We have therefore explored the influence of lack of both bradykinin receptors (B1R and B2R) on diabetic nephropathy, neuropathy, and osteopathy in male mice heterozygous for the Akita diabetogenic mutation in the insulin 2 gene (Ins2): We find that all of the detrimental phenotypes observed in Akita diabetes are enhanced by lack of both B1R and B2R, including urinary albumin excretion, glomerulosclerosis, glomerular basement membrane thickening, mitochondrial DNA deletions, reduction of nerve conduction velocities and of heat sensation, and bone mineral loss. Absence of the bradykinin receptors also enhances the diabetes-associated increases in plasma thiobarbituric acid-reactive substances, mitochondrial DNA deletions, and renal expression of fibrogenic genes, including transforming growth factor beta1, connective tissue growth factor, and endothelin-1. Thus, lack of B1R and B2R exacerbates diabetic complications. The enhanced renal injury in diabetic mice caused by lack of B1R and B2R may be mediated by a combination of increases in oxidative stress, mitochondrial DNA damage and over expression of fibrogenic genes. diabetes mellitus complications | kinins doi/ 10.1073/pnas.1005144107

The TIM22 protein import pathway mediates the import of membrane proteins into the mitochondrial inner membrane and consists of two intermembrane space chaperone complexes, the Tim9-Tim 10 and Tim8-Tim13 complexes. To facilitate mechanistic studies, we developed a chemical-genetic approach to identify small molecule agonists that caused lethality to a tim10-1 yeast mutant at the permissive temperature. One molecule, MitoBIoCK-1, attenuated the import of the carrier proteins including the ADP/ATP and phosphate carriers, but not proteins that used the TIM23 or the Mia40/ Erv1 translocation pathways. MitoBloCK-1 impeded binding of the Tim9-Tim10 complex to the substrate during an early stage of translocation, when the substrate was crossing the outer membrane. As a probe to determine the substrate specificity of the small Tim proteins, MitoBloCK-1 impaired the import of Tim22 and Tafazzin, but not Tim23, indicating that the Tim9-Tim10 complex mediates the import of a subset of inner membrane proteins. MitoBloCK-1 also inhibited growth of mammalian cells and import of the ADP/ATP carrier, but not TIM23 substrates, confirming that MitoBloCK-1 can be used to understand mammalian mitochondrial import and dysfunction linked to inherited human disease. Our approach of screening chemical libraries for compounds causing synthetic genetic lethality to identify inhibitors of mitochondrial protein translocation in yeast validates the generation of new probes to facilitate mechanistic studies in yeast and mammalian mitochondria. chemical biology | chemical genetics www.pnas.org/cgi/doi/10.1073/pnas.0914387107

[alpha]E-catenin binds the cell--cell adhesion complex of [alpha]E-cadherin and [beta]-catenin ([beta]-cat) regulates filamentous actin (F-actin) dynamics. In vitro, binding of [alpha]E-catenin to the E-cadherin-[beta]-cat complex lowers [alpha]E-catenin affinity for F-actin, and [alpha]E-catenin alone can bind F-actin and inhibit Arp2/3 complex--mediated actin polymerization. In cells, to test whether [alpha]E-catenin regulates actin dynamics independently of the cadherin complex, the cytosolic [alpha]E-catenin pool was sequestered to mitochondria without affecting overall levels of [alpha]E-catenin or the cadherin--catenin complex. Sequestering cytosolic [alpha]E-catenin to mitochondria alters lamellipodia architecture and increases membrane dynamics and cell migration without affecting cell--cell adhesion. In contrast, sequestration of cytosolic [alpha]E-catenin to the plasma membrane reduces membrane dynamics. These results demonstrate that the cytosolic pool of [alpha]E-catenin regulates actin dynamics independently of cell--cell adhesion. doi/10.1083/jcb.200910041

Parkinson's disease (PD) is a prevalent neurodegenerative disorder. Recent identification of genes linked to familial forms of PD such as Parkin and PINK1 (PTEN-induced putative kinase 1) has revealed that ubiquitylation and mitochondrial integrity are key factors in disease pathogenesis. However, the exact mechanism underlying the functional interplay between Parkin-catalyzed ubiquitylation and PINK1-regulated mitochondrial quality control remains an enigma. In this study, we show that PINK1 is rapidly and constitutively degraded under steady-state conditions in a mitochondrial membrane potential--dependent manner and that a loss in mitochondrial membrane potential stabilizes PINK1 mitochondrial accumulation. Furthermore, PINK1 recruits Parkin from the cytoplasm to mitochondria with low membrane potential to initiate the autophagic degradation of damaged mitochondria. Interestingly, the ubiquitin ligase activity of Parkin is repressed in the cytoplasm under steady-state conditions; however, PINK1-dependent mitochondrial localization liberates the latent enzymatic activity of Parkin. Some pathogenic mutations of PINK1 and Parkin interfere with the aforementioned events, suggesting an etiological importance. These results provide crucial insight into the pathogenic mechanisms of PD. doi/10.1083/jcb.200910140

Riboswitches are natural RNA sensors that regulate gene expression in response to ligand binding. Riboswitches have been identified in prokaryotes and eukaryotes but are unknown in organelles (mitochondria and plastids). Here we have tested the possibility to engineer riboswitches for plastids (chloroplasts), a genetic system that largely relies on translational control of gene expression. To this end, we have used bacterial riboswitches and modified them in silico to meet the requirements of translational regulation in plastids. These engineered switches were then tested for functionality in vivo by stable transformation of the tobacco chloroplast genome. We report the identification of a synthetic riboswitch that functions as an efficient translational regulator of gene expression in plastids in response to its exogenously applied ligand theophylline. This riboswitch provides a novel tool for plastid genome engineering that facilitates the tightly regulated inducible expression of chloroplast genes and transgenes and thus has wide applications in functional genomics and biotechnology. chloroplast | plastid transformation | translational regulation doi/10.1073/pnas.0914423107

The Quebec population is one of the rare populations of its size for which genealogical information is available for an uninterrupted period of almost four centuries. This allows for in-depth studies on the formation and evolution of a young founder population. Using data from two major population registers, in this study we focus on the maternal and paternal lineages (i.e., strictly female or male genealogical lines) that can be traced back within the Quebec genealogies. Through the analysis of these lineages it is possible to characterize the founders who transmitted to the contemporary population their mitochondrial (for females) and Y-chromosome (for males) DNA. The basic material consists of 2,221 ascending genealogies of subjects who married in the Quebec population between 1945 and 1965. On average, more than nine generations of ancestors were identified among the lineages. Analyses of maternal and paternal lineages show that the number of paternal founders is higher and their origins and genetic contributions are more variable than that of maternal founders, leading to a larger effective population size and greater diversity of Y chromosomes than of mtDNA. This is explained for the most part by differential migratory patterns among male and female founders of the Quebec population. Comparisons of sex-specific genetic contributions with total genetic contribution showed a strong correlation between the two values, with some discrepancies related to sex ratio differences among the founders' first descendants. KEY WORDS: MATERNAL LINEAGES, PATERNAL LINEAGES, GENEALOGIES, FOUNDERS, GENETIC CONTRIBUTION, KINSHIP, QUEBEC POPULATION, CANADA., Paternal lines of descent correspond to the patronymic lineage (transmission of surnames) and to the transmission line of Y chromosomes, whereas female lineages correspond to the transmission line of mitochondrial [...]

Mitochondrial DNA from 14 archaeological samples at the Ural State University in Yekaterinburg, Russia, was extracted to test the feasibility of ancient DNA work on their collection. These samples come from a number of sites that fall into two groupings. Seven samples are from three sites, dating to the 8th-12th century AD, that belong to a northern group of what are thought to be Ugrians, who lived along the Ural Mountains in northwestern Siberia. The remaining seven samples are from two sites that belong to a southern group representing the Sargat culture, dating between roughly the 5th century BC and the 5th century AD, from southwestern Siberia near the Ural Mountains and the present-day Kazakhstan border. The samples are derived from several burial types, including kurgan burials. They also represent a number of different skeletal elements and a range of observed preservation. The northern sites repeatedly failed to amplify after multiple extraction and amplification attempts, but the samples from the southern sites were successfully extracted and amplified. The sequences obtained from the southern sites support the hypothesis that the Sargat culture was a potential zone of intermixture between native Ugrian and/or Siberian populations and steppe peoples from the south, possibly early Iranian or Indo-Iranian, which has been previously suggested by archaeological analysis. KEY WORDS: D-LOOP, RUSSIA, SIBERIA, ADNA, MTDNA, UGRIAN POPULATIONS, SARGAT CULTURE, KURGAN BURIALS., Human habitation of western Siberia dates back to at least the late Paleolithic. The region has long formed an identifiable cultural area marked by similar material culture, including such aspects [...]

Regulations of proliferation and quiescence in response to nutritional cues are important for medicine and basic biology. The fission yeast Schizosaccharomyces pombe serves as a model, owing to the shift of proliferating cells to the metabolically active quiescence (designate GO phase hereafter) by responding to low nitrogen source. S. pombe GO phase cells keep alive for months without growth and division. Nitrogen replenishment reinstates vegetative proliferation phase (designate VEG). Some 40 genes required for GO maintenance were identified, but many more remain to be identified. We here show, using mutants, that the proteasome is required for maintaining GO quiescence. Functional outcomes of proteasome in GO and VEG phases appear to be distinct. Upon proteasome dysfunction, a number of antioxidant proteins and compounds responsive to ROS (reactive oxygen species) are produced. In addition, autophagy-mediated destruction of mitochondria occurs, which suppresses the loss of viability by eliminating ROS-generating mitochondria. These defensive responses are found in GO but not in VEG, suggesting that the main function of proteasome in GO phase homeostasis is to minimize ROS. Proteasome and autophagy are thus collaborative to support the lifespan of S. pombe GO phase. cell cycle | cellular quiescence | proteolysis | mitochondria doi/10.1073/pnas.0911055107

The kangaroo rat Dipodomys merriami occurs widely over the Baja California peninsula, inhabiting all the arid and sandy lowlands. Its range encompasses diverse climatic, edaphic, and vegetation zones, including 2 islands. The 11 taxa recognized in the merriami complex in this region almost 50 years ago remain valid to the present. The 2 island forms originally were described as different species, and have been treated in different ways by subsequent authors. In this investigation we used a genetic analysis of 2 mitochondrial genes, cytochrome b (Cytb) and cytochrome c oxidase subunit III (COIII), to study patterns of genetic structuring in this species throughout the peninsula. We supplemented existing morphometric and pelage color data with new data, and integrated these data sets into a phylogenetic analysis. Finally, we explored how our results compared with the existing taxonomic arrangement of species and subspecies. Our phylogenetic analyses of molecular data used the concatenation of 1,140 base pairs (bp) of the Cytb gene and 690 bp of the COIII gene; cladograms were generated using maximum-parsimony, maximum-likelihood, and Bayesian inference procedures. A hierarchy of nested clades was produced, the highest level of which revealed 2 reciprocally monophyletic clades separated by 20 mutational steps. There is a Southern Clade extending north to the Vizcaino Desert in the west and San Jose Island in the east, and a Northern Clade that includes all the populations farther to the north. At a shallower level, the Southern Clade contains 4 subclades, including the populations of San Jose and Margarita islands, whereas the Northern Clade is composed of 3 subclades. Adding the morphometric and pelage coloration data sets to the analyses resulted in support for the 11 previously recognized taxa arranged in a single species, Dipodomys merriami, but with improved understanding of how the subspecies relate to each other. As is increasingly appreciated, our investigation supports a comprehensive approach involving multiple data sets that are sensitive to a wide temporal range of evolutionary history for phylogenetic reconstruction. Key words: kangaroo rat, mitochondrial DNA, multiple data sets, nested clade phylogeographic analysis, phylogenetics, subspecies, taxonomy

In vertebrates, thyroid hormones (THs, thyroxine, and triiodothyronine) are critical cell signaling molecules. THs regulate and coordinate physiology within and between cells, tissues, and whole organisms, in addition to controlling embryonic growth and development, via dose-dependent regulatory effects on essential genes. While invertebrates and plants do not have thyroid glands, many utilize THs for development, while others store iodine as TH derivatives or TH precursor molecules (iodotyrosines)--or produce similar hormones that act in analogous ways. Such common developmental roles for iodotyrosines across kingdoms suggest that a common endocrine signaling mechanism may account for coordinated evolutionary change in all multi-cellular organisms. Here, I expand my earlier hypothesis for the role of THs in vertebrate evolution by proposing a critical evolutionary role for iodine, the essential ingredient in all iodotyrosines and THs. Iodine is known to be crucial for life in many unicellular organisms (including evolutionarily ancient cyanobacteria), in part, because it acts as a powerful antioxidant. I propose that during the last 3-4 billion years, the ease with which various iodine species become volatile, react with simple organic compounds, and catalyze biochemical reactions explains why iodine became an essential constituent of life and the Earth's atmosphere--and a potential marker for the origins of life. From an initial role as membrane antioxidant and biochemical catalyst, spontaneous coupling of iodine with tyrosine appears to have created a versatile, highly reactive and mobile molecule, which over time became integrated into the machinery of energy production, gene function, and DNA replication in mitochondria. Iodotyrosines later coupled together to form THs, the ubiquitous cell-signaling molecules used by all vertebrates. Thus, due to their evolutionary history, THs, and their derivative and precursors molecules not only became essential for communicating within and between cells, tissues and organs, and for coordinating development and whole-body physiology in vertebrates, but they can also be shared between organisms from different kingdoms.

The Yakuts are a Turkic-speaking population from northeastern Siberia who are believed to have originated from ancient Turkic populations in South Siberia, based on archaeological and ethno-historical evidence. In order to better understand Yakut origins, we modeled 25 demographic scenarios and tested by coalescent simulation whether any are consistent with the patterns of mtDNA diversity observed in present-day Yakuts. The models consist of either two simulated demes that represent Yakuts and a South Siberian ancestral population, or three demes that also include a regional Northeast Siberian population that served as a source of local gene flow into the Yakut deme. The model that produced the best fit to the observed data defined a founder group with an effective female population size of only 150 individuals that migrated northwards approximately 1,000 years BP and who experienced significant admixture with neighboring populations in Northeastern Siberia. These simulation results indicate a pronounced founder effect that was primarily kin-structured and reconcile reported discrepancies between Yakut mtDNA and Y chromosome diversity levels. KEY WORDS Yakut origins; mitochondrial DNA; coalescent simulation

Mitochondrial DNA is thought to be especially prone to oxidative damage by reactive oxygen species generated through electron transport during cellular respiration. This damage is mitigated primarily by the base excision repair (BER) pathway, one of the few DNA repair pathways with confirmed activity on mitochondrial DNA. Through genetic epistasis analysis of the yeast Saccharomyces cevisiae, we examined the genetic interaction between each of the BER proteins previously shown to localize to the mitochondria. In addition, we describe a series of genetic interactions between BER components and the MutS homolog MSHI, a respiration-essential gene. We show that, in addition to their variable effects on mitochondrial function, mutant mshl alleles conferring partial function interact genetically at different points in mitochondrial BER. In addition to this separation of function, we also round that the role of Mshlp in BER is unlikely to be involved in the avoidance of large-scale deletions and rearrangements. DOI: 10.1534/genetics.109.103796

Mitochondria are critical for cellular bioenergetics, and they mediate apoptosis within cells. We used whole body peroxisome proliferator-activated receptor-[gamma] coactivator-1[alpha] (PGC-1[alpha]) knockout (KO) animals to investigate its role on organelle function, apoptotic signaling, and cytochrome-c oxidase activity, an indicator of mitochondrial content, in muscle and other tissues (brain, liver, and pancreas). Lack of PGC-1[alpha] reduced mitochondrial content in all muscles (17-44%; P < 0.05) but had no effect in brain, liver, and pancreas. However, the tissue expression of proteins involved in mitochondrial DNA maintenance [transcription factor A (Tfam)], import (Tim23), and remodeling [mitofusin 2 (Mfn2) and dynamin-related protein 1 (Drp1)] did not parallel the decrease in mitochondrial content in PGC-1[alpha] KO animals. These proteins remained unchanged or were upregulated (P < 0.05) in the highly oxidative heart, indicating a change in mitochondrial composition. A change in muscle organelle composition was also evident from the alterations in subsarcolemmal and intermyofibrillar mitochondrial respiration, which was impaired in the absence of PGC-1[alpha]. However, endurance-trained KO animals did not exhibit reduced mitochondrial respiration. Mitochondrial reactive oxygen species (ROS) production was not affected by the lack of PGC-1[alpha], but subsarcolemmal mitochondria from PGC-1[alpha] KO animals released a greater amount of cytochrome c than in WT animals following exogenous ROS treatment. Our results indicate that the lack of PGC-1[alpha] results in 1) a muscle type-specific suppression of mitochondrial content that depends on basal oxidative capacity, 2) an alteration in mitochondrial composition, 3) impaired mitochondrial respiratory function that can be improved by training, and 4) a greater basal protein release from subsarcolemmal mitochondria, indicating an enhanced mitochondrial apoptotic susceptibility. endurance training; exercise; mitochondrial biogenesis; reactive oxygen species

Signal transducer and activator of transcription 3 (STAT3) is a latent cytoplasmic transcription factor responsive to cytokine signaling and tyrosine kinase oncoproteins by nuclear translocation when it is tyrosine-phosphorylated. We report that malignant transformation by activated Ras is impaired without STAT3, in spite of the inability of Ras to drive STAT3 tyrosine phosphorylation or nuclear translocation. Moreover, STAT3 mutants that cannot be tyrosine-phosphorylated, that are retained in the cytoplasm, or that cannot bind DNA nonetheless supported Ras-mediated transformation. Unexpectedly, STAT3 was detected within mitochondria, and exclusive targeting of STAT3 to mitochondria without nuclear accumulation facilitated Ras transformation. Mitochondrial STAT3 sustained altered glycolytic and oxidative phosphorylation activities characteristic of cancer cells. Thus, in addition to its nuclear transcriptional rote, STAT3 regulates a rnetabotic function in mitochondria, supporting Ras-dependent malignant transformation.

Nuclear copies of mitochondrial DNA fragments (numts) have been reported in a number of mammalian taxa. The inclusion of these numt pseudogenes within mitochondrial data sets can unknowingly compromise the integrity of phylogenetic, systematic, and biogeographic studies. Here, we evaluate 6 unique cytochrome-b numt sequences isolated in the southern red-backed vole (Myodes [formerly Clethrionomys] gapperi). Each sequence contains features characteristic of numt pseudogenes including premature stop codons, insertions--deletions (indels), flame-shift substitutions, and transposable element insertions. Mitochondrial and nuclear sequences are evolving at different rates. Molecular dating indicates that this group of mitochondrial transfers originated ~4.6 million years ago, which predates the presumptive origin of the genus. Phylogenetic analyses of mitochondrial and nuclear sequences suggest that the numts are the result of at least 1 nuclear insertion followed by subsequent duplication events, but pairwise sequence comparisons do not support this assertion. This conflict is likely the result of incomplete numt sampling from the Myodes nuclear genome. Genome sequencing efforts will ultimately provide the data needed to fully characterize numt demography in different lineages, and we predict that numts will have phylogenetic utility as informative synapomorphies in closely related species. Key words: Arvicolinae, Clethrionomys, cytochrome b, mitochondrial transfer, nuclear integration

The first large-scale fine characterization of Tunisian H lineages clarifies that the post-Last glacial maximum expansion originating in Iberia hot only led to the resettlement of Europe but also of North Africa. We found that 46% of 81 Tunisian H lineages subscreened for 1,580 bp in mtDNA coding region were affiliated with H1 and H3 subhaplogroups, which are known to have originated in Iberia. Although no signs of local expansion were detected, which would allow a clear dating of their introduction, the younger and less diverse Tunisian H1 and H3 lineages indicate Iberia as the radiating centre. Major contributions from historical migrations to this Iberian genetic imprint in Tunisia were ruled out by the mtDNA gene pool similarity between Berber/Arab/cosmopolitan samples and some 'Andalusian' communities, settled by the descendents of the 'Moors' who once lived in Iberia for 10 centuries (between 8th and 17th centuries), before being expelled to Tunisia. KEY WORDS mtDNA; haplogroup H; Tunisia; Andalusians

Molecules of mitochondrial DNA (mtDNA) are packed into nucleic acid-protein complexes termed mitochondrial nucleoids (mt-nucleoids). In this study, we analysed mt-nucleoids of the yeast Candida parapsilosis, which harbours a linear form of the mitochondrial genome. To identify conserved as well as specific features of mt-nucleoids in this species, we employed two strategies for analysis of their components. First, we investigated the protein composition of mt-nucleoids isolated from C. parapsilosis mitochondria, determined N-terminal amino acid sequences of 14 proteins associated with the mt-nucleoids and identified corresponding genes. Next, we complemented the list of mt-nucleoid components with additional candidates identified in the complete genome sequence of C. parapsilosis as homologues of Saccharomyces cerevisiae mt-nucleoid proteins. Our approach revealed several known mt-nucleoid proteins as well as additional components that expand the repertoire of proteins associated with these cytological structures. In particular, we identified and purified the protein Gcf1, which is abundant in the mt-nucleoids and exhibits structural features in common with the mtDNA packaging protein Abf2 from S. cerevisiae. We demonstrate that Gcf1 p co-localizes with mtDNA, has DNA-binding activity in vitro, and is able to stabilize mtDNA in the S. cerevisiae [DELTA]abf2 mutant, all of which points to a role in the maintenance of the C. parapsilosis mitochondrial genome. Importantly, in contrast to Abf2p, in silico analysis of Gcf1p predicted the presence of a coiled-coil domain and a single high-mobility group (HMG) box, suggesting that it represents a novel type of mitochondrial HMG protein.

Cucumber (Cucumis sativus) has one of the largest mitochondrial genomes known among all eukaryotes, due in part to the accumulation of short 20 to 60 by repetitive DNA motifs. Recombination among these repetitive DNAs produces rearrangements affecting organization and expression of mitochondrial genes. To more efficiently identify rearrangements in the cucumber mitochondrial DNA, we built two nonoverlapping 800 and 220 kb BAC contigs and assigned major mitochondrial genes to these BACs. Polymorphism carried on the largest BAC contig was used to confirm paternal transmission. Mitochondrial genes were distributed across BACs and physically distant, although occasional clustering was observed. Introns in the nad1, nad4, and nad7 genes were larger than those reported in other plants, due in part to accumulation of short repetitive DNAs and indicating that increased intron sizes contributed to mitochondrial genome expansion in cucumber. Mitochondrial genes atp6 and atp9 are physically close to each other and cotranscribed. These physical contigs will be useful for eventual sequencing of the cucumber mitochondrial DNA, which can be exploited to more efficiently screen for unique rearrangements affecting mitochondrial gene expression. Key words: Cucumis sativus, bacterial artificial chromosome, FPC, organellar genome. Le concombre (Cucumis sativus) possede fun des plus grands genomms mitochondriaux chez les eucaryotes en raison, en partie, dune accumulation de courts motifs repetitifs d'ADN de 20 a 60 pb. La recombinaison entre ces sequences repetees produit des rearrangements qui affectent l'organisation et (expression des genes mitochondriaux. Afin d'identifier plus efficacement les rearrangements au sein de l'ADN mitochondrial du concombre, les auteurs ont assemble deux contigs totalisant 800 et 220 kb de clones BAC sans chevauchements et ont assigne les principaux genes mitochondriaux a ces BAC. Le polymorphisme present sur le plus grand contig BAC a ete employe pour en confirmer la transmission paternelle. Les genes mitochondriaux etaient distrbbues sur les BAC et separes dune grande distance physique, bien que quelques groupements aient ete observes. Les Introns dens les genes nad1, nad4 et nad7 etaient plus grands que leurs homologues chez d'autres plantes en raison, en partie, de l'accumulation de courts motifs repetes d'ADN, ce qui indique que des Introns de taille accrue ont contribue a l'expansion du genome mitochondrial chez le concombre. Les genes mitochondriaux atp6 et atp9 sont physiquement proches l'un de l'autre et co-transcrits. Ces contigs physiques seront utiles en vue du sequencage de l'ADN mitochondrial du concombre qu'il serait possible d'exploiter pour identifier efficacement des rearrangements uniques affectent (expression de genes mitochondriaux. Mots-cles : Cucumis sativus, chromosome bacterien artifcial, FBC, genome d'organite. [Traduit par la Redaction], Introduction Plant mitochondrial genomes are larger than those of most other eukaryotes, even though the numbers of mitochondrially encoded genes are similar (Unseld et al. 1997; Kubo et al. 2000; [...]

An increasing number of cancer types have been found to be linked to specific mutations in the mitochondrial DNA, which result in specific structural changes of the respiratory enzyme complexes. In this study, we have investigated the effect of 2 such mutations identified in colon cancer patients, leading to the amino acid substitutions Ser458Pro and Gly125Asp in subunit I of cytochrome c oxidase (complex IV) [Greaves et al. (2006) Proc Natl Acad Sci USA 103:714-719]. We introduced these mutations in Rhodobacter sphaeroides, which carries an oxidase that serves as a model of the mitochondrial counterpart. The lack of expression of the former variant indicates that the amino acid substitution results in severely altered overall structure of the enzyme. The latter mutation (Gly171Asp in the bacterial oxidase) resulted in a structurally intact enzyme, but with reduced activity (approximately 30%), mainly due to slowed reduction of the redox site heme a. Furthermore, even though the Gly171Asp CytcO pumps protons, an intrinsic proton leak was identified, which would lead to a decreased overall energy-conversion efficiency of the respiratory chain, and would also perturb transport processes such as protein, ion, and metabolite trafficking. Furthermore, the specific leak may act to alter the balance between the electrical and chemical components of the proton electrochemical gradient. cytochrome [aa.sub.3] | electrochemical | electron transfer | pump | respiratory oxidase

Nerve, muscle, and inflammatory cells involved in gastrointestinal (GI) function have high-energy requirements and are affected in mitochondrial disorders. Familial aggregation of irritable bowel syndrome (IBS) frequently involves mothers and their children. Since mitochondrial DNA (mtDNA) is maternally inherited, mtDNA single nucleotide polymorphisms (SNPs) could confer risk to the development of IBS. The mtDNA SNPs, 16519C>T and 3010G>A, are associated with migraine and childhood cyclic vomiting syndrome. Our hypothesis is that these mtDNA SNPs are associated with functional GI disorders (FGIDs) and GI functions. The mt genome was first tested for the 7028C polymorphism (defining haplogroup H) in 699 patients or controls, and those with 7028C were further genotyped at 16519 and 3010. Phenotypes were based on symptoms (validated questionnaires and criteria) and GI physiology using validated motor and sensory studies. Constipation-predominant IBS and alternating constipation and diarrhea IBS are less prevalent in individuals with the 7028C mtDNA polymorphism than in individuals with 7028T. Conversely, 7028C is associated with higher maximum tolerated volume (lower satiation) compared with 7028T. Among those with 7028C, nonspecific abdominal pain (chronic abdominal pain or dyspepsia) was significantly associated with 3010A compared with 3010G (odds ratio 3.3, P = 0.02), and slower gastric emptying was statistically associated with 3010G. There were no significant associations of mtDNA genotypes tested and stomach volumes, small bowel or colonic transit, rectal compliance, and motor or sensory functions. Thus variation in mtDNA may be associated with satiation, gastric emptying, and possibly pain; further studies of mtDNA in appetite regulation and larger numbers of patients with FGIDs are warranted. 16519T; 3010A; 7028C; irritable bowel syndrome; dyspepsia; haplogroup H; genotype; constipation; nonspecific abdominal pain; gastric emptying; accommodation; satiation; somatic symptoms

Mitochondrial (mt)DNA mutations contribute to various disease states characterized by low ATP production. In contrast, thyroid hormone [3,3',5-triiodothyronine ([T.sub.3])] induces mitochondrial biogenesis and enhances ATP generation within cells. To evaluate the role of T3-mediated mitochondrial biogenesis in patients with mtDNA mutations, three fibroblast cell lines with mtDNA mutations were evaluated, including two patients with Leigh's syndrome and one with hypertrophic cardiomyopathy. Compared with control cells, patient fibroblasts displayed similar levels of mitochondrial mass, peroxisome proliferator-activated receptor-[gamma] coactivator-1[alpha] (PGC1[alpha]), mitochondrial transcription factor A (Tram), and uncoupling protein 2 (UCP2) protein expression. However, patient cells exhibited a 1.6-fold elevation in ROS production, a 1.7-fold elevation in cytoplasmic Ca2+ levels, a 1.2-fold elevation in mitochondrial membrane potential, and 30% less complex V activity compared with control cells. Patient cells also displayed 20-25% reductions in both cytochrome c oxidase (COX) activity and MnSOD protein levels compared with control cells. After [T.sub.3] treatment of patient cells, ROS production was decreased by 40%, cytoplasmic [Ca.sup.2+] was reduced by 20%, COX activity was increased by 1.3-fold, and ATP levels were elevated by 1.6-fold, despite the absence of a change in mitochondrial mass. There were no significant alterations in the protein expression of PGC-1[alpha], Tfam, or UCP2 in either [T.sub.3]-treated patient or control cells. However, [T.sub.3] restored the mitochondrial membrane potential, complex V activity, and levels of MnSOD to normal values in patient cells and elevated MnSOD levels by 21% in control cells. These results suggest that [T.sub.3] acts to reduce cellular oxidative stress, which may help attenuate ROSmediated damage, along with improving mitochondrial function and energy status in cells with mtDNA defects. reactive oxygen species; mitochondrial biogenesis; cytoplasmic calcium; mitochondrial disease; 3,3',5-triiodothyronine; mitochondrial DNA

We investigated the genetic population structure of the Black-throated Blue Warbler (Dendroica caerulescens), a Nearctic-Neotropic migrant passerine that breeds in cool mixed deciduous--coniferous forests in eastern North America. A cline in plumage color in breeding populations in the central Appalachian Mountains suggests either a contact zone between two formerly allopatric populations or the presence of a strong contemporary selection gradient. Analysis of 333 base pairs of the mitochondrial control region from 287 individuals sampled from 14 populations revealed relatively high haplotype diversity, low nucleotide diversity, and limited but significant phylogeographic structure across the breeding range (analysis of molecular variance [AMOVA], variation among populations = 2.7%; P < 0.01) and between northern and southern population groups (AMOVA, variation among groups = 2.9%; P < 0.01). Genetic differentiation among populations did not conform to an isolation-by-distance model. Nucleotide diversity was generally highest in the central Appalachians and lower in geographically peripheral populations. Populations from the northwestern periphery of the breeding range in Michigan had the lowest haplotype diversity and were genetically distinct from populations in the southern Appalachians. The star-shaped haplotype network, extensive sharing of common haplotypes among populations, and the haphazard distribution of rare haplotypes are most likely attributable to the combined effects of postglacial expansion from a single refugium (12,000-84,000 years ago) and long-distance dispersal events. The existence of a cline in plumage color, in the face of inferred recent gene flow, suggests that a strong selection gradient is operating, perhaps related to the migratory divide postulated from stableisotope data. Key words: Appalachian Mountains, Black-throated Blue Warbler, Dendroica caerulescens, genetic structure, glaciation, mitochondrial DNA, plumage color, stable isotopes.

The Northern Talysh from Azerbaijan and the Southern Talysh from Iran self-identify as one ethnic group and speak a Northwestern Iranian language. However, the Northern and Southern Talysh dialects are so different that they may actually be separate languages. Does this linguistic differentiation reflect internal change due to isolation, or could contact-induced change have played a role? We analyzed mtDNA HVI sequences, 11 Y-chromosome bi-allelic markers, and 9 YSTR loci in Northern and Southern Talysh and compared them with their neighboring groups. The mtDNA data show a close relatedness of both groups with each other and with neighboring groups, whereas the Northern Talysh Y-chromosome variation differs from that of neighboring groups, probably as a result of genetic drift. This genetic drift most likely reflects a founder event in the male gene pool of Northern Talysh: either fewer males than females migrated to Azerbaijan, or there was a higher degree of relatedness among the male migrants. Since we find no evidence of substantial genetic contact between either Northern or Southern Talysh and neighboring groups, we conclude that internal change, rather than contact-induced change, most likely explains the linguistic differentiation between Northern and Southern Talysh. Am J Phys Anthropol 138:82-89, 2009. KEY WORDS Northern Talysh; Southern Talysh; Y-chromosome; mtDNA