WorldWideScience

Sample records for positive-strand dna phenotype

  1. DNA repair phenotype and dietary antioxidant supplementation

    DEFF Research Database (Denmark)

    Guarnieri, Serena; Loft, Steffen; Riso, Patrizia

    2008-01-01

    Phytochemicals may protect cellular DNA by direct antioxidant effect or modulation of the DNA repair activity. We investigated the repair activity towards oxidised DNA in human mononuclear blood cells (MNBC) in two placebo-controlled antioxidant intervention studies as follows: (1) well-nourished......Phytochemicals may protect cellular DNA by direct antioxidant effect or modulation of the DNA repair activity. We investigated the repair activity towards oxidised DNA in human mononuclear blood cells (MNBC) in two placebo-controlled antioxidant intervention studies as follows: (1) well......-nourished subjects who ingested 600 g fruits and vegetables, or tablets containing the equivalent amount of vitamins and minerals, for 24 d; (2) poorly nourished male smokers who ingested 500 mg vitamin C/d as slow- or plain-release formulations together with 182 mg vitamin E/d for 4 weeks. The mean baseline levels...

  2. Epigenetic reversion of breast carcinoma phenotype is accompaniedby DNA sequestration

    Energy Technology Data Exchange (ETDEWEB)

    Sandal, Tone; Valyi-Nagy, Klara; Spencer, Virginia A.; Folberg,Robert; Bissell, Mina J.; Maniotis, Andrew J.

    2006-07-19

    The importance of microenvironment and context in regulation of tissue-specific genes is finally well established. DNA exposure to, or sequestration from, nucleases can be used to detect differences in higher order chromatin structure in intact cells without disturbing cellular or tissue architecture. To investigate the relationship between chromatin organization and tumor phenotype, we utilized an established 3-D assay where normal and malignant human breast cells can be easily distinguished by the morphology of the structures they make (acinus-like vs tumor-like, respectively). We show that these phenotypes can be distinguished also by sensitivity to AluI digestion where the malignant cells are resistant to digestion relative to non-malignant cells. Reversion of the T4-2 breast cancer cells by either cAMP analogs, or a phospatidylinositol 3-kinase (P13K) inhibitor not only reverted the phenotype, but also the chromatin sensitivity to AluI. By using different cAMP-analogs, we show that the cAMP-induced phenotypic reversion, polarization, and shift in DNA organization act through a cAMP-dependent-protein-kinase A-coupled signaling pathway. Importantly, inhibitory antibody to fibronectin also reverted the malignant phenotype, polarized the acini, and changed chromatin sequestration. These experiments show not only that modifying the tumor microenvironment can alter the organization of tumor cells but also that architecture of the tissues and the global chromatin organization are coupled and yet highly plastic.

  3. Nuclear DNA but not mtDNA controls tumor phenotypes in mouse cells

    International Nuclear Information System (INIS)

    Akimoto, Miho; Niikura, Mamoru; Ichikawa, Masami; Yonekawa, Hiromichi; Nakada, Kazuto; Honma, Yoshio; Hayashi, Jun-Ichi

    2005-01-01

    Recent studies showed high frequencies of homoplasmic mtDNA mutations in various human tumor types, suggesting that the mutated mtDNA haplotypes somehow contribute to expression of tumor phenotypes. We directly addressed this issue by isolating mouse mtDNA-less (ρ 0 ) cells for complete mtDNA replacement between normal cells and their carcinogen-induced transformants, and examined the effect of the mtDNA replacement on expression of tumorigenicity, a phenotype forming tumors in nude mice. The results showed that genome chimera cells carrying nuclear DNA from tumor cells and mtDNA from normal cells expressed tumorigenicity, whereas those carrying nuclear DNA from normal cells and mtDNA from tumor cells did not. These observations provided direct evidence that nuclear DNA, but not mtDNA, is responsible for carcinogen-induced malignant transformation, although it remains possible that mtDNA mutations and resultant respiration defects may influence the degree of malignancy, such as invasive or metastatic properties

  4. Use of Cellular Decapping Activators by Positive-Strand RNA Viruses

    Directory of Open Access Journals (Sweden)

    Jennifer Jungfleisch

    2016-12-01

    Full Text Available Positive-strand RNA viruses have evolved multiple strategies to not only circumvent the hostile decay machinery but to trick it into being a priceless collaborator supporting viral RNA translation and replication. In this review, we describe the versatile interaction of positive-strand RNA viruses and the 5′-3′ mRNA decay machinery with a focus on the viral subversion of decapping activators. This highly conserved viral trickery is exemplified with the plant Brome mosaic virus, the animal Flock house virus and the human hepatitis C virus.

  5. DNA Phenotyping: The prediction of human pigmentation traits from genetic data

    NARCIS (Netherlands)

    S. Walsh (Susan)

    2013-01-01

    textabstractPhenotyping is the ability to assign characteristics to an organism based on certain measurable parameters. In the case of DNA phenotyping, it is limited to the sole use of DNA to determine a phenotype such as an externally visible characteristic. In a forensic setting, it encompasses

  6. Forensic DNA phenotyping: Developing a model privacy impact assessment.

    Science.gov (United States)

    Scudder, Nathan; McNevin, Dennis; Kelty, Sally F; Walsh, Simon J; Robertson, James

    2018-05-01

    Forensic scientists around the world are adopting new technology platforms capable of efficiently analysing a larger proportion of the human genome. Undertaking this analysis could provide significant operational benefits, particularly in giving investigators more information about the donor of genetic material, a particularly useful investigative lead. Such information could include predicting externally visible characteristics such as eye and hair colour, as well as biogeographical ancestry. This article looks at the adoption of this new technology from a privacy perspective, using this to inform and critique the application of a Privacy Impact Assessment to this emerging technology. Noting the benefits and limitations, the article develops a number of themes that would influence a model Privacy Impact Assessment as a contextual framework for forensic laboratories and law enforcement agencies considering implementing forensic DNA phenotyping for operational use. Copyright © 2018 Elsevier B.V. All rights reserved.

  7. Forensic DNA Phenotyping: Predicting human appearance from crime scene material for investigative purposes.

    Science.gov (United States)

    Kayser, Manfred

    2015-09-01

    Forensic DNA Phenotyping refers to the prediction of appearance traits of unknown sample donors, or unknown deceased (missing) persons, directly from biological materials found at the scene. "Biological witness" outcomes of Forensic DNA Phenotyping can provide investigative leads to trace unknown persons, who are unidentifiable with current comparative DNA profiling. This intelligence application of DNA marks a substantially different forensic use of genetic material rather than that of current DNA profiling presented in the courtroom. Currently, group-specific pigmentation traits are already predictable from DNA with reasonably high accuracies, while several other externally visible characteristics are under genetic investigation. Until individual-specific appearance becomes accurately predictable from DNA, conventional DNA profiling needs to be performed subsequent to appearance DNA prediction. Notably, and where Forensic DNA Phenotyping shows great promise, this is on a (much) smaller group of potential suspects, who match the appearance characteristics DNA-predicted from the crime scene stain or from the deceased person's remains. Provided sufficient funding being made available, future research to better understand the genetic basis of human appearance will expectedly lead to a substantially more detailed description of an unknown person's appearance from DNA, delivering increased value for police investigations in criminal and missing person cases involving unknowns. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  8. Morphological analysis and DNA methylation in Conyza bonariensis L. cronquist (Asteraceae phenotypes

    Directory of Open Access Journals (Sweden)

    Juliana Maria de Paula

    2017-08-01

    Full Text Available ABSTRACT The species Conyza bonariensis (L. cause losses in agriculture due to their invasive capacity and resistance to herbicides like glyphosate. The species of this genus exhibit phenotypic plasticity, which complicates their identification and characterization. Thus, experiments were performed with 2 extreme C. bonariensis phenotypes (called broad leaf and narrow leaf in greenhouse conditions and in the laboratory, in order to verify if the morphological differences among these phenotypes are a genetic character or result from environmental effects. In addition to the comparative morphological analysis, assessment of DNA methylation profile was performed to detect the occurrence, or not, of differences in the epigenetic level. The morphological characteristics evaluated were length, width, shape, margin and leaves indument; plant height and stem indument; the number of capitula, flowers and seeds. The Methylation Sensitive Amplified Polymorphism technique was used to investigate the methylation levels. The morphological differences of phenotypes supposed to be C. bonariensis are probably genetic in origin and not the result of environmental effects, since, after 6 crop cycles in a greenhouse under the same environmental conditions, these phenotypes remained with the same morphological characteristics and seed production in relation to the original phenotypes found in the collection site. The different phenotypes did not show differences corresponding to DNA methylation patterns that could indicate an epigenetic effect as the cause of the differences between the 2 phenotypes. The results of morphological analysis and methylation probably indicate that maybe they are individuals of populations from different taxa not registered yet in the literature.

  9. Ectopic ERK Expression Induces Phenotypic Conversion of C10 Cells and Alters DNA Methyltransferase Expression

    Energy Technology Data Exchange (ETDEWEB)

    Sontag, Ryan L.; Weber, Thomas J.

    2012-05-04

    In some model systems constitutive extracellular signal regulated kinase (ERK) activation is sufficient to promote an oncogenic phenotype. Here we investigate whether constitutive ERK expression influences phenotypic conversion in murine C10 type II alveolar epithelial cells. C10 cells were stably transduced with an ERK1-green fluorescent protein (ERK1-GFP) chimera or empty vector and ectopic ERK expression was associated with the acquisition of soft agar focus-forming potential in late passage, but not early passage cells. Late passage ERK1-GFP cells exhibited a significant increase in the expression of DNA methyl transferases (DNMT1 and 3b) and a marked increase in sensitivity to 5-azacytidine (5-azaC)-mediated toxicity, relative to early passage ERK1-GFP cells and vector controls. The expression of xeroderma pigmentosum complementation group A (XPA) and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) were significantly increased in late passage cells, suggesting enhanced DNA damage recognition and repair activity which we interpret as a reflection of genomic instability. Phospho-ERK levels were dramatically decreased in late passage ERK1-GFP cells, relative to early passage and vector controls, and phospho-ERK levels were restored by treatment with sodium orthovanadate, indicating a role for phosphatase activity in this response. Collectively these observations suggest that ectopic ERK expression promotes phenotypic conversion of C10 cells that is associated with latent effects on epigenetic programming and phosphatase activities.

  10. Genotype-Phenotype Correlation of Maternally Inherited Disorders due to Mutations in Mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    Peterus Thajeb

    2006-09-01

    Full Text Available Mitochondrial disorders are heterogeneous systemic ailments that are most often caused by maternal inheritance of a variety of mutations of the mitochondrial (mt DNA. Paternal inheritance and somatic mutation are rare. The disorders are well recognized not only for the genotypic heterogeneity, but also the phenotypic variation among the affected members of a single family. The genotype-phenotype correlation of the diversity of the syndromic and non-syndromic features of mitochondrial disorders are discussed. Some aspects of the molecular mechanisms of this heterogeneity, and the histopathologic findings are highlighted.

  11. Failure to induce a DNA repair gene, RAD54, in Saccharomyces cerevisiae does not affect DNA repair or recombination phenotypes

    International Nuclear Information System (INIS)

    Cole, G.M.; Mortimer, R.K.

    1989-01-01

    The Saccharomyces cerevisiae RAD54 gene is transcriptionally regulated by a broad spectrum of DNA-damaging agents. Induction of RAD54 by DNA-damaging agents is under positive control. Sequences responsible for DNA damage induction (the DRS element) lie within a 29-base-pair region from -99 to -70 from the most proximal transcription start site. This inducible promoter element is functionally separable from a poly(dA-dT) region immediately downstream which is required for constitutive expression. Deletions which eliminate induction of RAD54 transcription by DNA damage but do not affect constitutive expression have no effect on growth or survival of noninducible strains relative to wild-type strains in the presence of DNA-damaging agents. The DRS element is also not required for homothallic mating type switching, transcriptional induction of RAD54 during meiosis, meiotic recombination, or spontaneous or X-ray-induced mitotic recombination. We find no phenotype for a lack of induction of RAD54 message via the damage-inducible DRS, which raises significant questions about the physiology of DNA damage induction in S. cerevisiae

  12. Characterization of DNA repair phenotypes of Xeroderma pigmentosum cell lines by a paralleled in vitro test

    International Nuclear Information System (INIS)

    Raffin, A.L.

    2009-06-01

    DNA is constantly damaged modifying the genetic information for which it encodes. Several cellular mechanisms as the Base Excision Repair (BER) and the Nucleotide Excision Repair (NER) allow recovering the right DNA sequence. The Xeroderma pigmentosum is a disease characterised by a deficiency in the NER pathway. The aim of this study was to propose an efficient and fast test for the diagnosis of this disease as an alternative to the currently available UDS test. DNA repair activities of XP cell lines were quantified using in vitro miniaturized and paralleled tests in order to establish DNA repair phenotypes of XPA and XPC deficient cells. The main advantage of the tests used in this study is the simultaneous measurement of excision or excision synthesis (ES) of several lesions by only one cellular extract. We showed on one hand that the relative ES of the different lesions depend strongly on the protein concentration of the nuclear extract tested. Working at high protein concentration allowed discriminating the XP phenotype versus the control one, whereas it was impossible under a certain concentration's threshold. On the other hand, while the UVB irradiation of control cells stimulated their repair activities, this effect was not observed in XP cells. This study brings new information on the XPA and XPC protein roles during BER and NER and underlines the complexity of the regulations of DNA repair processes. (author)

  13. DNA methylation profiling reveals the presence of population-specific signatures correlating with phenotypic characteristics.

    Science.gov (United States)

    Giri, Anil K; Bharadwaj, Soham; Banerjee, Priyanka; Chakraborty, Shraddha; Parekatt, Vaisak; Rajashekar, Donaka; Tomar, Abhishek; Ravindran, Aarthi; Basu, Analabha; Tandon, Nikhil; Bharadwaj, Dwaipayan

    2017-06-01

    Phenotypic characteristics are known to vary substantially among different ethnicities around the globe. These variations are mediated by number of stochastic events and cannot be attributed to genetic architecture alone. DNA methylation is a well-established mechanism that sculpts our epigenome influencing phenotypic variation including disease manifestation. Since DNA methylation is an important determinant for health issues of a population, it demands a thorough investigation of the natural differences in genome wide DNA methylation patterns across different ethnic groups. This study is based on comparative analyses of methylome from five different ethnicities with major focus on Indian subjects. The current study uses hierarchical clustering approaches, principal component analysis and locus specific differential methylation analysis on Illumina 450K methylation data to compare methylome of different ethnic subjects. Our data indicates that the variations in DNA methylation patterns of Indians are less among themselves compared to other global population. It empirically correlated with dietary, cultural and demographical divergences across different ethnic groups. Our work further suggests that Indians included in this study, despite their genetic similarity with the Caucasian population, are in close proximity with Japanese in terms of their methylation signatures.

  14. dNTP pool levels modulate mutator phenotypes of error-prone DNA polymerase ε variants.

    Science.gov (United States)

    Williams, Lindsey N; Marjavaara, Lisette; Knowels, Gary M; Schultz, Eric M; Fox, Edward J; Chabes, Andrei; Herr, Alan J

    2015-05-12

    Mutator phenotypes create genetic diversity that fuels tumor evolution. DNA polymerase (Pol) ε mediates leading strand DNA replication. Proofreading defects in this enzyme drive a number of human malignancies. Here, using budding yeast, we show that mutator variants of Pol ε depend on damage uninducible (Dun)1, an S-phase checkpoint kinase that maintains dNTP levels during a normal cell cycle and up-regulates dNTP synthesis upon checkpoint activation. Deletion of DUN1 (dun1Δ) suppresses the mutator phenotype of pol2-4 (encoding Pol ε proofreading deficiency) and is synthetically lethal with pol2-M644G (encoding altered Pol ε base selectivity). Although pol2-4 cells cycle normally, pol2-M644G cells progress slowly through S-phase. The pol2-M644G cells tolerate deletions of mediator of the replication checkpoint (MRC) 1 (mrc1Δ) and radiation sensitive (Rad) 9 (rad9Δ), which encode mediators of checkpoint responses to replication stress and DNA damage, respectively. The pol2-M644G mutator phenotype is partially suppressed by mrc1Δ but not rad9Δ; neither deletion suppresses the pol2-4 mutator phenotype. Thus, checkpoint activation augments the Dun1 effect on replication fidelity but is not required for it. Deletions of genes encoding key Dun1 targets that negatively regulate dNTP synthesis, suppress the dun1Δ pol2-M644G synthetic lethality and restore the mutator phenotype of pol2-4 in dun1Δ cells. DUN1 pol2-M644G cells have constitutively high dNTP levels, consistent with checkpoint activation. In contrast, pol2-4 and POL2 cells have similar dNTP levels, which decline in the absence of Dun1 and rise in the absence of the negative regulators of dNTP synthesis. Thus, dNTP pool levels correlate with Pol ε mutator severity, suggesting that treatments targeting dNTP pools could modulate mutator phenotypes for therapy.

  15. DNA fragmentation and nuclear phenotype in tendons exposed to low-intensity infrared laser

    Science.gov (United States)

    de Paoli, Flavia; Ramos Cerqueira, Larissa; Martins Ramos, Mayara; Campos, Vera M.; Ferreira-Machado, Samara C.; Geller, Mauro; de Souza da Fonseca, Adenilson

    2015-03-01

    Clinical protocols are recommended in device guidelines outlined for treating many diseases on empirical basis. However, effects of low-intensity infrared lasers at fluences used in clinical protocols on DNA are controversial. Excitation of endogenous chromophores in tissues and free radicals generation could be described as a consequence of laser used. DNA lesions induced by free radicals cause changes in DNA structure, chromatin organization, ploidy degrees and cell death. In this work, we investigated whether low-intensity infrared laser therapy could alter the fibroblasts nuclei characteristics and induce DNA fragmentation. Tendons of Wistar rats were exposed to low-intensity infrared laser (830 nm), at different fluences (1, 5 and 10 J/cm2), in continuous wave (power output of 10mW, power density of 79.6 mW/cm2). Different frequencies were analyzed for the higher fluence (10 J/cm2), at pulsed emission mode (2.5, 250 and 2500 Hz), with the laser source at surface of skin. Geometric, densitometric and textural parameters obtained for Feulgen-stained nuclei by image analysis were used to define nuclear phenotypes. Significant differences were observed on the nuclear phenotype of tendons after exposure to laser, as well as, high cell death percentages was observed for all fluences and frequencies analyzed here, exception 1 J/cm2 fluence. Our results indicate that low-intensity infrared laser can alter geometric, densitometric and textural parameters in tendon fibroblasts nuclei. Laser can also induce DNA fragmentation, chromatin lost and consequently cell death, using fluences, frequencies and emission modes took out from clinical protocols.

  16. To What Extent Does DNA Methylation Affect Phenotypic Variation in Cattle?

    Directory of Open Access Journals (Sweden)

    Stephanie McKAY

    2015-07-01

    Full Text Available DNA methylation is an environmentally influenced epigenetic modification that regulates gene transcription and has the potential to influence variation in economically important phenotypes in agricultural species. We have utilized a novel approach to evaluate the relationship between genetic and epigenetic variation and downstream phenotypes. To begin with, we have integrated RNA-Seq and methyl binding domain sequencing (MBD-Seq data in order to determine the extent to which DNA methylation affects phenotypic variation in economically important traits of cattle. MBD-Seq is a technique that involves the sample enrichment of methylated genomic regions followed by their next-generation sequencing. This study utilized Illumina next generation sequencing technology to perform both RNA-Seq and MBD-Seq. NextGENe software (SoftGenetics, State College, PA was employed for quality trimming and aligning the sequence reads to the UMD3.1 bovine reference genome, generating counts of matched reads and methylated peak identification. Subsequently, we identified and quantified genome-wide methylated regions and characterized the extent of differential methylation and differential expression between two groups of animals with extreme phenotypes. The program edgeR from the R software package (version 3.0.1 was employed for identifying differentially methylated regions and regions of differential expression. Finally, Partial Correlation with Information Theory (PCIT was performed to identify transcripts and methylation events that exhibit differential hubbing. A differential hub is defined as a gene network hub that is more highly connected in one treatment group than the other. This analysis produced every possible pair-wise interaction that subsequently enabled us to look at network interactions of how methylation affects expression. (co-expression, co-methylation, methylation x expression. Genomic regions of interest derived from this analysis were then aligned

  17. Sequence homology at the breakpoint and clinical phenotype of mitochondrial DNA deletion syndromes.

    Science.gov (United States)

    Sadikovic, Bekim; Wang, Jing; El-Hattab, Ayman W; Landsverk, Megan; Douglas, Ganka; Brundage, Ellen K; Craigen, William J; Schmitt, Eric S; Wong, Lee-Jun C

    2010-12-20

    Mitochondrial DNA (mtDNA) deletions are a common cause of mitochondrial disorders. Large mtDNA deletions can lead to a broad spectrum of clinical features with different age of onset, ranging from mild mitochondrial myopathies (MM), progressive external ophthalmoplegia (PEO), and Kearns-Sayre syndrome (KSS), to severe Pearson syndrome. The aim of this study is to investigate the molecular signatures surrounding the deletion breakpoints and their association with the clinical phenotype and age at onset. MtDNA deletions in 67 patients were characterized using array comparative genomic hybridization (aCGH) followed by PCR-sequencing of the deletion junctions. Sequence homology including both perfect and imperfect short repeats flanking the deletion regions were analyzed and correlated with clinical features and patients' age group. In all age groups, there was a significant increase in sequence homology flanking the deletion compared to mtDNA background. The youngest patient group (deletion distribution in size and locations, with a significantly lower sequence homology flanking the deletion, and the highest percentage of deletion mutant heteroplasmy. The older age groups showed rather discrete pattern of deletions with 44% of all patients over 6 years old carrying the most common 5 kb mtDNA deletion, which was found mostly in muscle specimens (22/41). Only 15% (3/20) of the young patients (deletion, which is usually present in blood rather than muscle. This group of patients predominantly (16 out of 17) exhibit multisystem disorder and/or Pearson syndrome, while older patients had predominantly neuromuscular manifestations including KSS, PEO, and MM. In conclusion, sequence homology at the deletion flanking regions is a consistent feature of mtDNA deletions. Decreased levels of sequence homology and increased levels of deletion mutant heteroplasmy appear to correlate with earlier onset and more severe disease with multisystem involvement.

  18. Mutations in BALB mitochondrial DNA induce CCL20 up-regulation promoting tumorigenic phenotypes

    Energy Technology Data Exchange (ETDEWEB)

    Sligh, James [Department of Medicine—Dermatology Division, University of Arizona, Tucson, AZ 857 24 (United States); University of Arizona Cancer Center, Tucson, AZ 85724 (United States); Janda, Jaroslav [University of Arizona Cancer Center, Tucson, AZ 85724 (United States); Jandova, Jana, E-mail: jjandova@email.arizona.edu [Department of Medicine—Dermatology Division, University of Arizona, Tucson, AZ 857 24 (United States); University of Arizona Cancer Center, Tucson, AZ 85724 (United States)

    2014-11-15

    Highlights: • Alterations in mitochondrial DNA are commonly found in various human cancers. • Mutations in BALB mitochondrial DNA induce up-regulation of chemokine CCL20. • Increased growth and motility of mtBALB cells is associated with CCL20 levels. • mtDNA changes in BALB induce in vivo tumor growth through CCL20 up-regulation. • Mutations in mitochondrial DNA play important roles in keratinocyte neoplasia. - Abstract: mtDNA mutations are common in human cancers and are thought to contribute to the process of neoplasia. We examined the role of mtDNA mutations in skin cancer by generating fibroblast cybrids harboring a mutation in the gene encoding the mitochondrial tRNA for arginine. This somatic mutation (9821insA) was previously reported in UV-induced hyperkeratotic skin tumors in hairless mice and confers specific tumorigenic phenotypes to mutant cybrids. Microarray analysis revealed and RT-PCR along with Western blot analysis confirmed the up-regulation of CCL20 and its receptor CCR6 in mtBALB haplotype containing the mt-Tr 9821insA allele compared to wild type mtB6 haplotype. Based on reported role of CCL20 in cancer progression we examined whether the hyper-proliferation and enhanced motility of mtBALB haplotype would be associated with CCL20 levels. Treatment of both genotypes with recombinant CCL20 (rmCCL20) resulted in enhanced growth and motility of mtB6 cybrids. Furthermore, the acquired somatic alteration increased the in vivo tumor growth of mtBALB cybrids through the up-regulation of CCL20 since neutralizing antibody significantly decreased in vivo tumor growth of these cells; and tumors from anti-CCL20 treated mice injected with mtBALB cybrids showed significantly decreased CCL20 levels. When rmCCL20 or mtBALB cybrids were used as chemotactic stimuli, mtB6 cybrids showed increased motility while anti-CCL20 antibody decreased the migration and in vivo tumor growth of mtBALB cybrids. Moreover, the inhibitors of MAPK signaling and NF

  19. Phenotypic and mtDNA variation in Philippine Kappaphycus cottonii (Gigartinales, Rhodophyta).

    Science.gov (United States)

    Dumilag, Richard V; Gallardo, William George M; Garcia, Christian Philip C; You, YeaEun; Chaves, Alyssa Keren G; Agahan, Lance

    2017-11-09

    Members of the carrageenan-producing seaweeds of the genus Kappapphycus have a complicated taxonomic history particularly with regard to species identification. Many taxonomic challenges in this group have been currently addressed with the use of mtDNA sequences. The phylogenetic status and genetic diversity of one of the lesser known species, Kappaphycus cottonii, have repeatedly come into question. This study explored the genetic variation in Philippine K. cottonii using the mtDNA COI-5P gene and cox2-3 spacer sequences. The six phenotypic forms in K. cottonii did not correspond to the observed genetic variability; hinting at the greater involvement of environmental factors in determining changes to the morphology of this alga. Our results revealed that the Philippine K. cottonii has the richest number of haplotypes that have been detected, so far, for any Kappaphycus species. Our inferred phylogenetic trees suggested two lineages: a lineage, which exclusively includes K. cottonii and another lineage comprising the four known Kappaphycus species: K. alvarezii, K. inermis, K. malesianus, and K. striatus. The dichotomy supports the apparent synamorphy for each of these lineages (the strictly terete thalli, lack of protuberances, and the presence of a hyphal central core in the latter group, while the opposite of these morphologies in K. cottonii). These findings shed new light on understanding the evolutionary history of the genus. Assessing the breadth of the phenotypic and genetic variation in K. cottonii has implications for the conservation and management of the overall Kappaphycus genetic resources, especially in the Philippines.

  20. Accumulation of pathogenic ΔmtDNA induced deafness but not diabetic phenotypes in mito-mice

    International Nuclear Information System (INIS)

    Nakada, Kazuto; Sato, Akitsugu; Sone, Hideyuki; Kasahara, Atsuko; Ikeda, Katsuhisa; Kagawa, Yasuo; Yonekawa, Hiromichi; Hayashi, Jun-Ichi

    2004-01-01

    Mito-mice carrying various proportions of deletion mutant mtDNA (ΔmtDNA) were generated by introduction of the ΔmtDNA from cultured cells into fertilized eggs of C57BL/6J (B6) strain mice. Great advantages of mito-mice are that they share exactly the same nuclear-genome background, and that their genetic variations are restricted to proportions of pathogenic ΔmtDNA. Since accumulation of ΔmtDNA to more than 75% induced respiration defects, the disease phenotypes observed exclusively in mito-mice carrying more than 75% ΔmtDNA should be due to accumulated ΔmtDNA. In this study, we focused on the expressions of hearing loss and diabetic phenotypes, since these common age-associated abnormalities have sometimes been reported to be inherited maternally and to be associated with pathogenic mutant mtDNAs. The results showed that accumulation of exogenously introduced ΔmtDNA was responsible for hearing loss, but not for expression of diabetic phenotypes in mito-mice

  1. Phenotypic Analysis of ATM Protein Kinase in DNA Double-Strand Break Formation and Repair.

    Science.gov (United States)

    Mian, Elisabeth; Wiesmüller, Lisa

    2017-01-01

    Ataxia telangiectasia mutated (ATM) encodes a serine/threonine protein kinase, which is involved in various regulatory processes in mammalian cells. Its best-known role is apical activation of the DNA damage response following generation of DNA double-strand breaks (DSBs). When DSBs appear, sensor and mediator proteins are recruited, activating transducers such as ATM, which in turn relay a widespread signal to a multitude of downstream effectors. ATM mutation causes Ataxia telangiectasia (AT), whereby the disease phenotype shows differing characteristics depending on the underlying ATM mutation. However, all phenotypes share progressive neurodegeneration and marked predisposition to malignancies at the organismal level and sensitivity to ionizing radiation and chromosome aberrations at the cellular level. Expression and localization of the ATM protein can be determined via western blotting and immunofluorescence microscopy; however, detection of subtle alterations such as resulting from amino acid exchanges rather than truncating mutations requires functional testing. Previous studies on the role of ATM in DSB repair, which connects with radiosensitivity and chromosomal stability, gave at first sight contradictory results. To systematically explore the effects of clinically relevant ATM mutations on DSB repair, we engaged a series of lymphoblastoid cell lines (LCLs) derived from AT patients and controls. To examine DSB repair both in a quantitative and qualitative manners, we used an EGFP-based assay comprising different substrates for distinct DSB repair mechanisms. In this way, we demonstrated that particular signaling defects caused by individual ATM mutations led to specific DSB repair phenotypes. To explore the impact of ATM on carcinogenic chromosomal aberrations, we monitored chromosomal breakage at a breakpoint cluster region hotspot within the MLL gene that has been associated with therapy-related leukemia. PCR-based MLL-breakage analysis of HeLa cells

  2. Approaching ethical, legal and social issues of emerging forensic DNA phenotyping (FDP) technologies comprehensively: Reply to ‘Forensic DNA phenotyping: Predicting human appearance from crime scene material for investigative purposes’ by Manfred Kayser

    NARCIS (Netherlands)

    Toom, V.; Wienroth, M.; M'charek, A.; Prainsack, B.; Williams, R.; Duster, T.; Heinemann, T.; Kruse, C.; Machado, H.; Murphy, E.

    2016-01-01

    In a recent special issue of the journal on new trends in forensic genetics, Manfred Kayser contributed a review of developments, opportunities and challenges of forensic DNA phenotyping (FDP). In his article he argues that FDP technologies - such as determining eye, hair and skin color - should be

  3. Unique DNA methylome profiles in CpG island methylator phenotype colon cancers

    Science.gov (United States)

    Xu, Yaomin; Hu, Bo; Choi, Ae-Jin; Gopalan, Banu; Lee, Byron H.; Kalady, Matthew F.; Church, James M.; Ting, Angela H.

    2012-01-01

    A subset of colorectal cancers was postulated to have the CpG island methylator phenotype (CIMP), a higher propensity for CpG island DNA methylation. The validity of CIMP, its molecular basis, and its prognostic value remain highly controversial. Using MBD-isolated genome sequencing, we mapped and compared genome-wide DNA methylation profiles of normal, non-CIMP, and CIMP colon specimens. Multidimensional scaling analysis revealed that each specimen could be clearly classified as normal, non-CIMP, and CIMP, thus signifying that these three groups have distinctly different global methylation patterns. We discovered 3780 sites in various genomic contexts that were hypermethylated in both non-CIMP and CIMP colon cancers when compared with normal colon. An additional 2026 sites were found to be hypermethylated in CIMP tumors only; and importantly, 80% of these sites were located in CpG islands. These data demonstrate on a genome-wide level that the additional hypermethylation seen in CIMP tumors occurs almost exclusively at CpG islands and support definitively that these tumors were appropriately named. When these sites were examined more closely, we found that 25% were adjacent to sites that were also hypermethylated in non-CIMP tumors. Thus, CIMP is also characterized by more extensive methylation of sites that are already prone to be hypermethylated in colon cancer. These observations indicate that CIMP tumors have specific defects in controlling both DNA methylation seeding and spreading and serve as an important first step in delineating molecular mechanisms that control these processes. PMID:21990380

  4. eCOMPAGT integrates mtDNA: import, validation and export of mitochondrial DNA profiles for population genetics, tumour dynamics and genotype-phenotype association studies

    Directory of Open Access Journals (Sweden)

    Specht Günther

    2010-03-01

    Full Text Available Abstract Background Mitochondrial DNA (mtDNA is widely being used for population genetics, forensic DNA fingerprinting and clinical disease association studies. The recent past has uncovered severe problems with mtDNA genotyping, not only due to the genotyping method itself, but mainly to the post-lab transcription, storage and report of mtDNA genotypes. Description eCOMPAGT, a system to store, administer and connect phenotype data to all kinds of genotype data is now enhanced by the possibility of storing mtDNA profiles and allowing their validation, linking to phenotypes and export as numerous formats. mtDNA profiles can be imported from different sequence evaluation programs, compared between evaluations and their haplogroup affiliations stored. Furthermore, eCOMPAGT has been improved in its sophisticated transparency (support of MySQL and Oracle, security aspects (by using database technology and the option to import, manage and store genotypes derived from various genotyping methods (SNPlex, TaqMan, and STRs. It is a software solution designed for project management, laboratory work and the evaluation process all-in-one. Conclusions The extended mtDNA version of eCOMPAGT was designed to enable error-free post-laboratory data handling of human mtDNA profiles. This software is suited for small to medium-sized human genetic, forensic and clinical genetic laboratories. The direct support of MySQL and the improved database security options render eCOMPAGT a powerful tool to build an automated workflow architecture for several genotyping methods. eCOMPAGT is freely available at http://dbis-informatik.uibk.ac.at/ecompagt.

  5. eCOMPAGT integrates mtDNA: import, validation and export of mitochondrial DNA profiles for population genetics, tumour dynamics and genotype-phenotype association studies.

    Science.gov (United States)

    Weissensteiner, Hansi; Schönherr, Sebastian; Specht, Günther; Kronenberg, Florian; Brandstätter, Anita

    2010-03-09

    Mitochondrial DNA (mtDNA) is widely being used for population genetics, forensic DNA fingerprinting and clinical disease association studies. The recent past has uncovered severe problems with mtDNA genotyping, not only due to the genotyping method itself, but mainly to the post-lab transcription, storage and report of mtDNA genotypes. eCOMPAGT, a system to store, administer and connect phenotype data to all kinds of genotype data is now enhanced by the possibility of storing mtDNA profiles and allowing their validation, linking to phenotypes and export as numerous formats. mtDNA profiles can be imported from different sequence evaluation programs, compared between evaluations and their haplogroup affiliations stored. Furthermore, eCOMPAGT has been improved in its sophisticated transparency (support of MySQL and Oracle), security aspects (by using database technology) and the option to import, manage and store genotypes derived from various genotyping methods (SNPlex, TaqMan, and STRs). It is a software solution designed for project management, laboratory work and the evaluation process all-in-one. The extended mtDNA version of eCOMPAGT was designed to enable error-free post-laboratory data handling of human mtDNA profiles. This software is suited for small to medium-sized human genetic, forensic and clinical genetic laboratories. The direct support of MySQL and the improved database security options render eCOMPAGT a powerful tool to build an automated workflow architecture for several genotyping methods. eCOMPAGT is freely available at http://dbis-informatik.uibk.ac.at/ecompagt.

  6. DNA variation in the phenotypically-diverse brown alga Saccharina japonica

    Directory of Open Access Journals (Sweden)

    Balakirev Evgeniy S

    2012-07-01

    Full Text Available Abstract Background Saccharina japonica (Areschoug Lane, Mayes, Druehl et Saunders is an economically important and highly morphologically variable brown alga inhabiting the northwest Pacific marine waters. On the basis of nuclear (ITS, plastid (rbcLS and mitochondrial (COI DNA sequence data, we have analyzed the genetic composition of typical Saccharina japonica (TYP and its two common morphological varieties, known as the “longipes” (LON and “shallow-water” (SHA forms seeking to clarify their taxonomical status and to evaluate the possibility of cryptic species within S. japonica. Results The data show that the TYP and LON forms are very similar genetically in spite of drastic differences in morphology, life history traits, and ecological preferences. Both, however, are genetically quite different from the SHA form. The two Saccharina lineages are distinguished by 109 fixed single nucleotide differences as well as by seven fixed length polymorphisms (based on a 4,286 bp concatenated dataset that includes three gene regions. The GenBank database reveals a close affinity of the TYP and LON forms to S. japonica and the SHA form to S. cichorioides. The three gene markers used in the present work have different sensitivity for the algal species identification. COI gene was the most discriminant gene marker. However, we have detected instances of interspecific COI recombination reflecting putative historical hybridization events between distantly related algal lineages. The recombinant sequences show highly contrasted level of divergence in the 5’- and 3’- regions of the gene, leading to significantly different tree topologies depending on the gene segment (5’- or 3’- used for tree reconstruction. Consequently, the 5’-COI “barcoding” region (~ 650 bp can be misleading for identification purposes, at least in the case of algal species that might have experienced historical hybridization events. Conclusion Taking into account

  7. Murine Lupus Susceptibility Locus Sle2 Activates DNA-Reactive B Cells through Two Sub-Loci with Distinct Phenotypes

    Science.gov (United States)

    Zeumer, Leilani; Sang, Allison; Niu, Haitao; Morel, Laurence

    2010-01-01

    The NZM2410-derived Sle2 lupus susceptibility locus induces an abnormal B cell differentiation which most prominently leads to the expansion of autoreactive B1a cells. We have mapped the expansion of B1a cells to three Sle2 sub-loci, Sle2a, Sle2b, and Sle2c. Sle2 also enhances the breach of B cell tolerance to nuclear antigens in the 56R anti-DNA immunoglobulin transgenic (Tg) model. This study used the Sle2 sub-congenic strains to map the activation of 56R Tg B cells. Sle2c strongly sustained the breach of tolerance and the activation of anti-DNA B cells. The production of Tg-encoded anti-DNA antibodies was more modest in Sle2a expressing mice, but Sle2a was responsible for the recruitment for Tg B cells to the marginal zone, a phenotype that has been found for 56R Tg B cells in mice expressing the whole Sle2 interval. In addition, Sle2a promoted the production of endogenously encoded anti-DNA antibodies. Overall, this study showed that at least two Sle2 genes are involved in the activation of anti-DNA B cells, and excluded more than two-thirds of the Sle2 interval from contributing to this phenotype. This constitutes an important step toward the identification of novel genes that play a critical role in B cell tolerance. PMID:21270826

  8. Development of a porcine skeletal muscle cDNA microarray: analysis of differential transcript expression in phenotypically distinct muscles

    Directory of Open Access Journals (Sweden)

    Stear Michael

    2003-03-01

    Full Text Available Abstract Background Microarray profiling has the potential to illuminate the molecular processes that govern the phenotypic characteristics of porcine skeletal muscles, such as hypertrophy or atrophy, and the expression of specific fibre types. This information is not only important for understanding basic muscle biology but also provides underpinning knowledge for enhancing the efficiency of livestock production. Results We report on the de novo development of a composite skeletal muscle cDNA microarray, comprising 5500 clones from two developmentally distinct cDNA libraries (longissimus dorsi of a 50-day porcine foetus and the gastrocnemius of a 3-day-old pig. Clones selected for the microarray assembly were of low to moderate abundance, as indicated by colony hybridisation. We profiled the differential expression of genes between the psoas (red muscle and the longissimus dorsi (white muscle, by co-hybridisation of Cy3 and Cy5 labelled cDNA derived from these two muscles. Results from seven microarray slides (replicates correctly identified genes that were expected to be differentially expressed, as well as a number of novel candidate regulatory genes. Quantitative real-time RT-PCR on selected genes was used to confirm the results from the microarray. Conclusion We have developed a porcine skeletal muscle cDNA microarray and have identified a number of candidate genes that could be involved in muscle phenotype determination, including several members of the casein kinase 2 signalling pathway.

  9. Leaf margin phenotype-specific restriction-site-associated DNA-derived markers for pineapple (Ananas comosus L.).

    Science.gov (United States)

    Urasaki, Naoya; Goeku, Satoko; Kaneshima, Risa; Takamine, Tomonori; Tarora, Kazuhiko; Takeuchi, Makoto; Moromizato, Chie; Yonamine, Kaname; Hosaka, Fumiko; Terakami, Shingo; Matsumura, Hideo; Yamamoto, Toshiya; Shoda, Moriyuki

    2015-06-01

    To explore genome-wide DNA polymorphisms and identify DNA markers for leaf margin phenotypes, a restriction-site-associated DNA sequencing analysis was employed to analyze three bulked DNAs of F1 progeny from a cross between a 'piping-leaf-type' cultivar, 'Yugafu', and a 'spiny-tip-leaf-type' variety, 'Yonekura'. The parents were both Ananas comosus var. comosus. From the analysis, piping-leaf and spiny-tip-leaf gene-specific restriction-site-associated DNA sequencing tags were obtained and designated as PLSTs and STLSTs, respectively. The five PLSTs and two STSLTs were successfully converted to cleaved amplified polymorphic sequence (CAPS) or simple sequence repeat (SSR) markers using the sequence differences between alleles. Based on the genotyping of the F1 with two SSR and three CAPS markers, the five PLST markers were mapped in the vicinity of the P locus, with the closest marker, PLST1_SSR, being located 1.5 cM from the P locus. The two CAPS markers from STLST1 and STLST3 perfectly assessed the 'spiny-leaf type' as homozygotes of the recessive s allele of the S gene. The recombination value between the S locus and STLST loci was 2.4, and STLSTs were located 2.2 cM from the S locus. SSR and CAPS markers are applicable to marker-assisted selection of leaf margin phenotypes in pineapple breeding.

  10. A Phenotype-Driven Approach to Generate Mouse Models with Pathogenic mtDNA Mutations Causing Mitochondrial Disease

    Directory of Open Access Journals (Sweden)

    Johanna H.K. Kauppila

    2016-09-01

    Full Text Available Mutations of mtDNA are an important cause of human disease, but few animal models exist. Because mammalian mitochondria cannot be transfected, the development of mice with pathogenic mtDNA mutations has been challenging, and the main strategy has therefore been to introduce mutations found in cell lines into mouse embryos. Here, we describe a phenotype-driven strategy that is based on detecting clonal expansion of pathogenic mtDNA mutations in colonic crypts of founder mice derived from heterozygous mtDNA mutator mice. As proof of concept, we report the generation of a mouse line transmitting a heteroplasmic pathogenic mutation in the alanine tRNA gene of mtDNA displaying typical characteristics of classic mitochondrial disease. In summary, we describe a straightforward and technically simple strategy based on mouse breeding and histology to generate animal models of mtDNA-mutation disease, which will be of great importance for studies of disease pathophysiology and preclinical treatment trials.

  11. MitoLSDB: a comprehensive resource to study genotype to phenotype correlations in human mitochondrial DNA variations.

    Directory of Open Access Journals (Sweden)

    Shamnamole K

    Full Text Available Human mitochondrial DNA (mtDNA encodes a set of 37 genes which are essential structural and functional components of the electron transport chain. Variations in these genes have been implicated in a broad spectrum of diseases and are extensively reported in literature and various databases. In this study, we describe MitoLSDB, an integrated platform to catalogue disease association studies on mtDNA (http://mitolsdb.igib.res.in. The main goal of MitoLSDB is to provide a central platform for direct submissions of novel variants that can be curated by the Mitochondrial Research Community. MitoLSDB provides access to standardized and annotated data from literature and databases encompassing information from 5231 individuals, 675 populations and 27 phenotypes. This platform is developed using the Leiden Open (source Variation Database (LOVD software. MitoLSDB houses information on all 37 genes in each population amounting to 132397 variants, 5147 unique variants. For each variant its genomic location as per the Revised Cambridge Reference Sequence, codon and amino acid change for variations in protein-coding regions, frequency, disease/phenotype, population, reference and remarks are also listed. MitoLSDB curators have also reported errors documented in literature which includes 94 phantom mutations, 10 NUMTs, six documentation errors and one artefactual recombination. MitoLSDB is the largest repository of mtDNA variants systematically standardized and presented using the LOVD platform. We believe that this is a good starting resource to curate mtDNA variants and will facilitate direct submissions enhancing data coverage, annotation in context of pathogenesis and quality control by ensuring non-redundancy in reporting novel disease associated variants.

  12. Forensic DNA phenotyping in criminal investigations and criminal courts: assessing and mitigating the dilemmas inherent in the science.

    Science.gov (United States)

    MacLean, Charles E; Lamparello, Adam

    2014-01-01

    Forensic DNA Phenotyping ("FDP"), estimating the externally visible characteristics ("EVCs") of the source of human DNA left at a crime scene, is evolving from science fiction toward science fact. FDP can already identify a source's gender with 100% accuracy, and likely hair color, iris color, adult height, and a number of other EVCs with accuracy rates approaching 70%. Patent applications have been filed for approaches to generating 3D likenesses of DNA sources based on the DNA alone. Nonetheless, criminal investigators, particularly in the United States, have been reticent to apply FDP in their casework. The reticence is likely related to a number of perceived and real dilemmas associated with FDP: is FDP racial profiling, should we test unknown and unseen physical conditions, does testing for behavioral characteristics impermissibly violate the source's privacy, ought testing be permitted for samples from known sources or DNA databases, and should FDP be limited to use in investigations only or is FDP appropriate for use in a criminal court. As this article explains, although those dilemmas are substantive, they are not insurmountable, and can be quite easily managed with appropriate regulation and protocols. As FDP continues to develop, there will be less need for criminal investigators to shy away from FDP. Cold cases, missing persons, and victims in crimes without other evidence will one day soon all be well served by FDP.

  13. Clinical phenotype associations with various types of anti-dsDNA antibodies in patients with recent onset of rheumatic symptoms. Results from a multicentre observational study

    DEFF Research Database (Denmark)

    Compagno, Michele; Rekvig, Ole P; Bengtsson, Anders A

    2014-01-01

    UNLABELLED: Despite anti-dsDNA antibodies constitute a wide range of specificities, they are considered as the hallmark for systemic lupus erythematosus (SLE). OBJECTIVE: To identify clinical phenotypes associated with anti-dsDNA antibodies, independently of any clinical diagnoses. METHODS...

  14. Parallel Genetic and Phenotypic Evolution of DNA Superhelicity in Experimental Populations of Escherichia coli

    DEFF Research Database (Denmark)

    Crozat, Estelle; Winkworth, Cynthia; Gaffé, Joël

    2010-01-01

    , indicate that changes in DNA superhelicity have been important in the evolution of these populations. Surprisingly, however, most of the evolved alleles we tested had either no detectable or slightly deleterious effects on fitness, despite these signatures of positive selection.......DNA supercoiling is the master function that interconnects chromosome structure and global gene transcription. This function has recently been shown to be under strong selection in Escherichia coli. During the evolution of 12 initially identical populations propagated in a defined environment...

  15. Picornaviruses and nuclear functions: targeting a cellular compartment distinct from the replication site of a positive-strand RNA virus

    Directory of Open Access Journals (Sweden)

    Dylan eFlather

    2015-06-01

    Full Text Available The compartmentalization of DNA replication and gene transcription in the nucleus and protein production in the cytoplasm is a defining feature of eukaryotic cells. The nucleus functions to maintain the integrity of the nuclear genome of the cell and to control gene expression based on intracellular and environmental signals received through the cytoplasm. The spatial separation of the major processes that lead to the expression of protein-coding genes establishes the necessity of a transport network to allow biomolecules to translocate between these two regions of the cell. The nucleocytoplasmic transport network is therefore essential for regulating normal cellular functioning. The Picornaviridae virus family is one of many viral families that disrupt the nucleocytoplasmic trafficking of cells to promote viral replication. Picornaviruses contain positive-sense, single-stranded RNA genomes and replicate in the cytoplasm of infected cells. As a result of the limited coding capacity of these viruses, cellular proteins are required by these intracellular parasites for both translation and genomic RNA replication. Being of messenger RNA polarity, a picornavirus genome can immediately be translated upon entering the cell cytoplasm. However, the replication of viral RNA requires the activity of RNA-binding proteins, many of which function in host gene expression, and are consequently localized to the nucleus. As a result, picornaviruses disrupt nucleocytoplasmic trafficking to exploit protein functions normally localized to a different cellular compartment from which they translate their genome to facilitate efficient replication. Furthermore, picornavirus proteins are also known to enter the nucleus of infected cells to limit host-cell transcription and down-regulate innate antiviral responses. The interactions of picornavirus proteins and host-cell nuclei are extensive, required for a productive infection, and are the focus of this review.

  16. Isolation and characterization of Solenopsis invicta virus 3, a new positive-strand RNA virus infecting the red imported fire ant, Solenopsis invicta

    International Nuclear Information System (INIS)

    Valles, Steven M.; Hashimoto, Yoshifumi

    2009-01-01

    We report the discovery of a new virus from the red imported fire ant, Solenopsis invicta. Solenopsis invicta virus 3 (SINV-3) represents the third virus discovered from this ant species using the metagenomics approach. The single (positive)-strand RNA, monopartite, bicistronic genome of SINV-3 was sequenced in entirety (GenBank accession number (FJ528584)), comprised of 10,386 nucleotides, and polyadenylated at the 3' terminus. This genome size was confirmed by Northern analysis. The genome revealed 2 large open reading frames (ORFs) in the sense orientation with an untranslated region (UTR) at each end and between the two ORFs. The 5' proximal ORF (ORF 1) encoded a predicted protein of 299.1 kDa (2580 amino acids). The 3' proximal ORF (ORF 2) encoded a predicted protein of 73.2 kDa (651 amino acids). RNA-dependent RNA polymerase (RdRp), helicase, and protease domains were recognized in ORF 1. SDS-PAGE separation of purified SINV-3 particles yielded 2 bands (ostensibly capsid proteins) with a combined molecular mass of 77.3 kDa which was similar to the mass predicted by ORF 2 (73.2 kDa). Phylogenetic analysis of the conserved amino acid sequences containing domains I to VIII of the RdRp from dicistroviruses, iflaviruses, plant small RNA viruses, picornaviruses, and 4 unassigned positive-strand RNA viruses revealed a trichotomous phenogram with SINV-3 and Kelp fly virus comprising a unique cluster. Electron microscopic examination of negatively stained samples of SINV-3 revealed isometric particles with apparent projections and a diameter of 27.3 ± 1.3 nm. SINV-3 was successfully transmitted to uninfected workers by feeding. The minus (replicative) strand of SINV-3 was detected in worker ants indicating replication of the virus. The possibility of using SINV-3 as a microbial control agent for fire ants is discussed.

  17. Long-term effects of inducible mutagenic DNA repair on relative fitness and phenotypic diversification in Pseudomonas cichorii 302959.

    Science.gov (United States)

    Weigand, Michael R; Sundin, George W

    2009-01-01

    Mutagenic DNA repair (MDR) employs low-fidelity DNA polymerases capable of replicating past DNA lesions resulting from exposure to high-energy ultraviolet radiation (UVR). MDR confers UVR tolerance and activation initiates a transient mutator phenotype that may provide opportunities for adaptation. To investigate the potential role of MDR in adaptation, we have propagated parallel lineages of the highly mutable epiphytic plant pathogen Pseudomonas cichorii 302959 with daily UVR activation (UVR lineages) for approximately 500 generations. Here we examine those lineages through the measurement of relative fitness and observation of distinct colony morphotypes that emerged. Isolates and population samples from UVR lineages displayed gains in fitness relative to the ancestor despite increased rates of inducible mutation to rifampicin resistance. Regular activation of MDR resulted in the maintenance of genetic diversity within UVR lineages, including the reproducible diversification and coexistence of "round" and "fuzzy" colony morphotypes. These results suggest that inducible mutability may present a reasonable strategy for adaptive evolution in stressful environments by contributing to gains in relative fitness and diversification.

  18. A Distinct DNA Methylation Shift in a Subset of Glioma CpG Island Methylator Phenotypes during Tumor Recurrence

    Directory of Open Access Journals (Sweden)

    Camila Ferreira de Souza

    2018-04-01

    Full Text Available Summary: Glioma diagnosis is based on histomorphology and grading; however, such classification does not have predictive clinical outcome after glioblastomas have developed. To date, no bona fide biomarkers that significantly translate into a survival benefit to glioblastoma patients have been identified. We previously reported that the IDH mutant G-CIMP-high subtype would be a predecessor to the G-CIMP-low subtype. Here, we performed a comprehensive DNA methylation longitudinal analysis of diffuse gliomas from 77 patients (200 tumors to enlighten the epigenome-based malignant transformation of initially lower-grade gliomas. Intra-subtype heterogeneity among G-CIMP-high primary tumors allowed us to identify predictive biomarkers for assessing the risk of malignant recurrence at early stages of disease. G-CIMP-low recurrence appeared in 9.5% of all gliomas, and these resembled IDH-wild-type primary glioblastoma. G-CIMP-low recurrence can be characterized by distinct epigenetic changes at candidate functional tissue enhancers with AP-1/SOX binding elements, mesenchymal stem cell-like epigenomic phenotype, and genomic instability. Molecular abnormalities of longitudinal G-CIMP offer possibilities to defy glioblastoma progression. : IDH-mutant lower-grade glioma glioblastoma often progresses to a more aggressive phenotype upon recurrence. de Souza et al. examines the intra-subtype heterogeneity of initial G-CIMP-high and use this information to identify predictive biomarkers for assessing the risk of recurrence and malignant transformation. Keywords: longitudinal gliomas, DNA methylation, IDH mutation, G-CIMP-high, intra-subtype heterogeneity, malignant transformation and recurrence, G-CIMP-low, stem cell-like glioblastoma, predictive biomarkers

  19. Insights into hominin phenotypic and dietary evolution from ancient DNA sequence data.

    Science.gov (United States)

    Perry, George H; Kistler, Logan; Kelaita, Mary A; Sams, Aaron J

    2015-02-01

    Nuclear genome sequence data from Neandertals, Denisovans, and archaic anatomically modern humans can be used to complement our understanding of hominin evolutionary biology and ecology through i) direct inference of archaic hominin phenotypes, ii) indirect inference of those phenotypes by identifying the effects of previously-introgressed alleles still present among modern humans, or iii) determining the evolutionary timing of relevant hominin-specific genetic changes. Here we review and reanalyze published Neandertal and Denisovan genome sequence data to illustrate an example of the third approach. Specifically, we infer the timing of five human gene presence/absence changes that may be related to particular hominin-specific dietary changes and discuss these results in the context of our broader reconstructions of hominin evolutionary ecology. We show that pseudogenizing (gene loss) mutations in the TAS2R62 and TAS2R64 bitter taste receptor genes and the MYH16 masticatory myosin gene occurred after the hominin-chimpanzee divergence but before the divergence of the human and Neandertal/Denisovan lineages. The absence of a functional MYH16 protein may explain our relatively reduced jaw muscles; this gene loss may have followed the adoption of cooking behavior. In contrast, salivary amylase gene (AMY1) duplications were not observed in the Neandertal and Denisovan genomes, suggesting a relatively recent origin for the AMY1 copy number gains that are observed in modern humans. Thus, if earlier hominins were consuming large quantities of starch-rich underground storage organs, as previously hypothesized, then they were likely doing so without the digestive benefits of increased salivary amylase production. Our most surprising result was the observation of a heterozygous mutation in the first codon of the TAS2R38 bitter taste receptor gene in the Neandertal individual, which likely would have resulted in a non-functional protein and inter-individual PTC

  20. Quantitative DNA methylation analysis improves epigenotype-phenotype correlations in Beckwith-Wiedemann syndrome

    Science.gov (United States)

    Calvello, Mariarosaria; Tabano, Silvia; Colapietro, Patrizia; Maitz, Silvia; Pansa, Alessandra; Augello, Claudia; Lalatta, Faustina; Gentilin, Barbara; Spreafico, Filippo; Calzari, Luciano; Perotti, Daniela; Larizza, Lidia; Russo, Silvia; Selicorni, Angelo; Sirchia, Silvia M; Miozzo, Monica

    2013-01-01

    Beckwith-Wiedemann syndrome (BWS) is a rare disorder characterized by overgrowth and predisposition to embryonal tumors. BWS is caused by various epigenetic and/or genetic alterations that dysregulate the imprinted genes on chromosome region 11p15.5. Molecular analysis is required to reinforce the clinical diagnosis of BWS and to identify BWS patients with cancer susceptibility. This is particularly crucial prenatally because most signs of BWS cannot be recognized in utero. We established a reliable molecular assay by pyrosequencing to quantitatively evaluate the methylation profiles of ICR1 and ICR2. We explored epigenotype-phenotype correlations in 19 patients that fulfilled the clinical diagnostic criteria for BWS, 22 patients with suspected BWS, and three fetuses with omphalocele. Abnormal methylation was observed in one prenatal case and 19 postnatal cases, including seven suspected BWS. Seven cases showed ICR1 hypermethylation, five cases showed ICR2 hypomethylation, and eight cases showed abnormal methylation of ICR1 and ICR2 indicating paternal uniparental disomy (UPD). More cases of ICR1 alterations and UPD were found than expected. This is likely due to the sensitivity of this approach, which can detect slight deviations in methylation from normal levels. There was a significant correlation (p < 0.001) between the percentage of ICR1 methylation and BWS features: severe hypermethylation (range: 75–86%) was associated with macroglossia, macrosomia, and visceromegaly, whereas mild hypermethylation (range: 55–59%) was associated with umbilical hernia and diastasis recti. Evaluation of ICR1 and ICR2 methylation by pyrosequencing in BWS can improve epigenotype-phenotype correlations, detection of methylation alterations in suspected cases, and identification of UPD. PMID:23917791

  1. The methylation of nuclear and mitochondrial DNA in ageing phenotypes and longevity.

    Science.gov (United States)

    Bacalini, Maria Giulia; D'Aquila, Patrizia; Marasco, Elena; Nardini, Christine; Montesanto, Alberto; Franceschi, Claudio; Passarino, Giuseppe; Garagnani, Paolo; Bellizzi, Dina

    2017-07-01

    An increasing body of data is progressively indicating that the comprehension of the epigenetic landscape, actively integrated with the genetic elements, is crucial to delineate the molecular basis of the inter-individual complexity of ageing process. Indeed, it has emerged that DNA methylation changes occur during ageing, consisting mainly in a progressive process of genome demethylation, in a hypermethylation of gene-specific CpG dinucleotides, as well as in an inter-individual divergence of the epigenome due to stochastic events and environmental exposures throughout life, namely as epigenetic drift. Additionally, it has also come to light an implication of the mitochondrial genome in the regulation of the intracellular epigenetic landscape, as demonstrated by the being itself object of epigenetic modifications. An overview of DNA methylation changes occurring during ageing process at both nuclear and mitochondrial level will be described in this review, also taking into account the recent and promising data available on the 5-hydroxymethylcytosine. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Enrichment of Phosphatidylethanolamine in Viral Replication Compartments via Co-opting the Endosomal Rab5 Small GTPase by a Positive-Strand RNA Virus.

    Directory of Open Access Journals (Sweden)

    Kai Xu

    2016-10-01

    Full Text Available Positive-strand RNA viruses build extensive membranous replication compartments to support replication and protect the virus from antiviral responses by the host. These viruses require host factors and various lipids to form viral replication complexes (VRCs. The VRCs built by Tomato bushy stunt virus (TBSV are enriched with phosphatidylethanolamine (PE through a previously unknown pathway. To unravel the mechanism of PE enrichment within the TBSV replication compartment, in this paper, the authors demonstrate that TBSV co-opts the guanosine triphosphate (GTP-bound active form of the endosomal Rab5 small GTPase via direct interaction with the viral replication protein. Deletion of Rab5 orthologs in a yeast model host or expression of dominant negative mutants of plant Rab5 greatly decreases TBSV replication and prevents the redistribution of PE to the sites of viral replication. We also show that enrichment of PE in the viral replication compartment is assisted by actin filaments. Interestingly, the closely related Carnation Italian ringspot virus, which replicates on the boundary membrane of mitochondria, uses a similar strategy to the peroxisomal TBSV to hijack the Rab5-positive endosomes into the viral replication compartments. Altogether, usurping the GTP-Rab5-positive endosomes allows TBSV to build a PE-enriched viral replication compartment, which is needed to support peak-level replication. Thus, the Rab family of small GTPases includes critical host factors assisting VRC assembly and genesis of the viral replication compartment.

  3. Identification of cis-acting elements on positive-strand subgenomic mRNA required for the synthesis of negative-strand counterpart in bovine coronavirus.

    Science.gov (United States)

    Yeh, Po-Yuan; Wu, Hung-Yi

    2014-07-30

    It has been demonstrated that, in addition to genomic RNA, sgmRNA is able to serve as a template for the synthesis of the negative-strand [(-)-strand] complement. However, the cis-acting elements on the positive-strand [(+)-strand] sgmRNA required for (-)-strand sgmRNA synthesis have not yet been systematically identified. In this study, we employed real-time quantitative reverse transcription polymerase chain reaction to analyze the cis-acting elements on bovine coronavirus (BCoV) sgmRNA 7 required for the synthesis of its (-)-strand counterpart by deletion mutagenesis. The major findings are as follows. (1) Deletion of the 5'-terminal leader sequence on sgmRNA 7 decreased the synthesis of the (-)-strand sgmRNA complement. (2) Deletions of the 3' untranslated region (UTR) bulged stem-loop showed no effect on (-)-strand sgmRNA synthesis; however, deletion of the 3' UTR pseudoknot decreased the yield of (-)-strand sgmRNA. (3) Nucleotides positioned from -15 to -34 of the sgmRNA 7 3'-terminal region are required for efficient (-)-strand sgmRNA synthesis. (4) Nucleotide species at the 3'-most position (-1) of sgmRNA 7 is correlated to the efficiency of (-)-strand sgmRNA synthesis. These results together suggest, in principle, that the 5'- and 3'-terminal sequences on sgmRNA 7 harbor cis-acting elements are critical for efficient (-)-strand sgmRNA synthesis in BCoV.

  4. Relationship between commercially available DNA analysis and phenotypic observations on beef quality and tenderness.

    Science.gov (United States)

    Magolski, J D; Buchanan, D S; Maddock-Carlin, K R; Anderson, V L; Newman, D J; Berg, E P

    2013-11-01

    Warner-Bratzler shear force values from 560 mixed breed heifers and steers were used to determine estimates of genetic selection. Cattle were marketed from 2008 to 2011, and included five feedlot based research projects at the North Dakota State University-Carrington Research Extension Center. Samples were collected for IGENITY® analysis providing information that included selection indices and estimated breeding values for carcass traits. DNA-based test results were compared with actual carcass measurements. Marbling accounted for over 10% of the variation in WBSF while hot carcass weight was the second most influential carcass trait accounting for 4% (Pfeeding a diet that meets or exceeds recommended nutrients for growth are the most important factors influencing beef tenderness and acceptability. Copyright © 2013 Elsevier Ltd. All rights reserved.

  5. The usefulness of DNA sequencing after extraction by Whatman FTA filter matrix technology and phenotypic tests for differentiation of Candida albicans and Candida dubliniensis.

    Science.gov (United States)

    Kiraz, Nuri; Oz, Yasemin; Aslan, Huseyin; Muslumanoglu, Hamza

    2014-02-01

    Since C. dubliniensis is similar to C. albicans phenotypically, it can be misidentified as C. albicans. We aimed to investigate the prevalence of C. dubliniensis among isolates previously identified as C. albicans in our stocks and to compare the phenotypic methods and DNA sequencing of D1/D2 region on the ribosomal large subunit (rLSU) gene. A total of 850 isolates included in this study. Phenotypic identification was performed based on germ tube formation, chlamydospore production, colony colors on chromogenic agar, inability of growth at 45 °C and growth on hypertonic Sabouraud dextrose agar. Eighty isolates compatible with C. dubliniensis by at least one phenotypic test were included in the sequence analysis. Nested PCR amplification of D1/D2 region of the rLSU gene was performed after the fungal DNA extraction by Whatman FTA filter paper technology. The sequencing analysis of PCR products carried out by an automated capillary gel electrophoresis device. The rate of C. dubliniensis was 2.35 % (n = 20) among isolates previously described as C. albicans. Consequently, none of the phenotypic tests provided satisfactory performance alone in our study, and molecular methods required special equipment and high cost. Thus, at least two phenotypic methods can be used for identification of C. dubliniensis, and molecular methods can be used for confirmation.

  6. Identification of Cis-Acting Elements on Positive-Strand Subgenomic mRNA Required for the Synthesis of Negative-Strand Counterpart in Bovine Coronavirus

    Directory of Open Access Journals (Sweden)

    Po-Yuan Yeh

    2014-07-01

    Full Text Available It has been demonstrated that, in addition to genomic RNA, sgmRNA is able to serve as a template for the synthesis of the negative-strand [(−-strand] complement. However, the cis-acting elements on the positive-strand [(+-strand] sgmRNA required for (−-strand sgmRNA synthesis have not yet been systematically identified. In this study, we employed real-time quantitative reverse transcription polymerase chain reaction to analyze the cis-acting elements on bovine coronavirus (BCoV sgmRNA 7 required for the synthesis of its (−-strand counterpart by deletion mutagenesis. The major findings are as follows. (1 Deletion of the 5'-terminal leader sequence on sgmRNA 7 decreased the synthesis of the (−-strand sgmRNA complement. (2 Deletions of the 3' untranslated region (UTR bulged stem-loop showed no effect on (−-strand sgmRNA synthesis; however, deletion of the 3' UTR pseudoknot decreased the yield of (−-strand sgmRNA. (3 Nucleotides positioned from −15 to −34 of the sgmRNA 7 3'-terminal region are required for efficient (−-strand sgmRNA synthesis. (4 Nucleotide species at the 3'-most position (−1 of sgmRNA 7 is correlated to the efficiency of (−-strand sgmRNA synthesis. These results together suggest, in principle, that the 5'- and 3'-terminal sequences on sgmRNA 7 harbor cis-acting elements are critical for efficient (−-strand sgmRNA synthesis in BCoV.

  7. The 3'-terminal 55 nucleotides of bovine coronavirus defective interfering RNA harbor cis-acting elements required for both negative- and positive-strand RNA synthesis.

    Directory of Open Access Journals (Sweden)

    Wei-Yu Liao

    Full Text Available The synthesis of the negative-strand [(--strand] complement of the ∼30 kilobase, positive-strand [(+-strand] coronaviral genome is a necessary early step for genome replication. The identification of cis-acting elements required for (--strand RNA synthesis in coronaviruses, however, has been hampered due to insufficiencies in the techniques used to detect the (--strand RNA species. Here, we employed a method of head-to-tail ligation and real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR to detect and quantitate the synthesis of bovine coronavirus (BCoV defective interfering (DI RNA (- strands. Furthermore, using the aforementioned techniques along with Northern blot assay, we specifically defined the cis-acting RNA elements within the 3'-terminal 55 nucleotides (nts which function in the synthesis of (-- or (+-strand BCoV DI RNA. The major findings are as follows: (i nts from -5 to -39 within the 3'-terminal 55 nts are the cis-acting elements responsible for (--strand BCoV DI RNA synthesis, (ii nts from -3 to -34 within the 3'-terminal 55 nts are cis-acting elements required for (+-strand BCoV DI RNA synthesis, and (iii the nucleotide species at the 3'-most position (-1 is important, but not critical, for both (-- and (+-strand BCoV DI RNA synthesis. These results demonstrate that the 3'-terminal 55 nts in BCoV DI RNA harbor cis-acting RNA elements required for both (-- and (+-strand DI RNA synthesis and extend our knowledge on the mechanisms of coronavirus replication. The method of head-to-tail ligation and qRT-PCR employed in the study may also be applied to identify other cis-acting elements required for (--strand RNA synthesis in coronaviruses.

  8. Resistance to DNA Damaging Agents Produced Invasive Phenotype of Rat Glioma Cells—Characterization of a New in Vivo Model

    Directory of Open Access Journals (Sweden)

    Sonja Stojković

    2016-06-01

    Full Text Available Chemoresistance and invasion properties are severe limitations to efficient glioma therapy. Therefore, development of glioma in vivo models that more accurately resemble the situation observed in patients emerges. Previously, we established RC6 rat glioma cell line resistant to DNA damaging agents including antiglioma approved therapies such as 3-bis(2-chloroethyl-1-nitrosourea (BCNU and temozolomide (TMZ. Herein, we evaluated the invasiveness of RC6 cells in vitro and in a new orthotopic animal model. For comparison, we used C6 cells from which RC6 cells originated. Differences in cell growth properties were assessed by real-time cell analyzer. Cells’ invasive potential in vitro was studied in fluorescently labeled gelatin and by formation of multicellular spheroids in hydrogel. For animal studies, fluorescently labeled cells were inoculated into adult male Wistar rat brains. Consecutive coronal and sagittal brain sections were analyzed 10 and 25 days post-inoculation, while rats’ behavior was recorded during three days in the open field test starting from 25th day post-inoculation. We demonstrated that development of chemoresistance induced invasive phenotype of RC6 cells with significant behavioral impediments implying usefulness of orthotopic RC6 glioma allograft in preclinical studies for the examination of new approaches to counteract both chemoresistance and invasion of glioma cells.

  9. A Distinct DNA Methylation Shift in a Subset of Glioma CpG Island Methylator Phenotypes during Tumor Recurrence.

    Science.gov (United States)

    de Souza, Camila Ferreira; Sabedot, Thais S; Malta, Tathiane M; Stetson, Lindsay; Morozova, Olena; Sokolov, Artem; Laird, Peter W; Wiznerowicz, Maciej; Iavarone, Antonio; Snyder, James; deCarvalho, Ana; Sanborn, Zachary; McDonald, Kerrie L; Friedman, William A; Tirapelli, Daniela; Poisson, Laila; Mikkelsen, Tom; Carlotti, Carlos G; Kalkanis, Steven; Zenklusen, Jean; Salama, Sofie R; Barnholtz-Sloan, Jill S; Noushmehr, Houtan

    2018-04-10

    Glioma diagnosis is based on histomorphology and grading; however, such classification does not have predictive clinical outcome after glioblastomas have developed. To date, no bona fide biomarkers that significantly translate into a survival benefit to glioblastoma patients have been identified. We previously reported that the IDH mutant G-CIMP-high subtype would be a predecessor to the G-CIMP-low subtype. Here, we performed a comprehensive DNA methylation longitudinal analysis of diffuse gliomas from 77 patients (200 tumors) to enlighten the epigenome-based malignant transformation of initially lower-grade gliomas. Intra-subtype heterogeneity among G-CIMP-high primary tumors allowed us to identify predictive biomarkers for assessing the risk of malignant recurrence at early stages of disease. G-CIMP-low recurrence appeared in 9.5% of all gliomas, and these resembled IDH-wild-type primary glioblastoma. G-CIMP-low recurrence can be characterized by distinct epigenetic changes at candidate functional tissue enhancers with AP-1/SOX binding elements, mesenchymal stem cell-like epigenomic phenotype, and genomic instability. Molecular abnormalities of longitudinal G-CIMP offer possibilities to defy glioblastoma progression. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

  10. Limited phylogeographic signal in sex-linked and autosomal loci despite geographically, ecologically, and phenotypically concordant structure of mtDNA variation in the Holarctic avian genus Eremophila.

    Directory of Open Access Journals (Sweden)

    Sergei V Drovetski

    Full Text Available Phylogeographic studies of Holarctic birds are challenging because they involve vast geographic scale, complex glacial history, extensive phenotypic variation, and heterogeneous taxonomic treatment across countries, all of which require large sample sizes. Knowledge about the quality of phylogeographic information provided by different loci is crucial for study design. We use sequences of one mtDNA gene, one sex-linked intron, and one autosomal intron to elucidate large scale phylogeographic patterns in the Holarctic lark genus Eremophila. The mtDNA ND2 gene identified six geographically, ecologically, and phenotypically concordant clades in the Palearctic that diverged in the Early-Middle Pleistocene and suggested paraphyly of the horned lark (E. alpestris with respect to the Temminck's lark (E. bilopha. In the Nearctic, ND2 identified five subclades which diverged in the Late Pleistocene. They overlapped geographically and were not concordant phenotypically or ecologically. Nuclear alleles provided little information on geographic structuring of genetic variation in horned larks beyond supporting the monophyly of Eremophila and paraphyly of the horned lark. Multilocus species trees based on two nuclear or all three loci provided poor support for haplogroups identified by mtDNA. The node ages calculated using mtDNA were consistent with the available paleontological data, whereas individual nuclear loci and multilocus species trees appeared to underestimate node ages. We argue that mtDNA is capable of discovering independent evolutionary units within avian taxa and can provide a reasonable phylogeographic hypothesis when geographic scale, geologic history, and phenotypic variation in the study system are too complex for proposing reasonable a priori hypotheses required for multilocus methods. Finally, we suggest splitting the currently recognized horned lark into five Palearctic and one Nearctic species.

  11. A positive-strand RNA virus uses alternative protein-protein interactions within a viral protease/cofactor complex to switch between RNA replication and virion morphogenesis.

    Science.gov (United States)

    Dubrau, Danilo; Tortorici, M Alejandra; Rey, Félix A; Tautz, Norbert

    2017-02-01

    The viruses of the family Flaviviridae possess a positive-strand RNA genome and express a single polyprotein which is processed into functional proteins. Initially, the nonstructural (NS) proteins, which are not part of the virions, form complexes capable of genome replication. Later on, the NS proteins also play a critical role in virion formation. The molecular basis to understand how the same proteins form different complexes required in both processes is so far unknown. For pestiviruses, uncleaved NS2-3 is essential for virion morphogenesis while NS3 is required for RNA replication but is not functional in viral assembly. Recently, we identified two gain of function mutations, located in the C-terminal region of NS2 and in the serine protease domain of NS3 (NS3 residue 132), which allow NS2 and NS3 to substitute for uncleaved NS2-3 in particle assembly. We report here the crystal structure of pestivirus NS3-4A showing that the NS3 residue 132 maps to a surface patch interacting with the C-terminal region of NS4A (NS4A-kink region) suggesting a critical role of this contact in virion morphogenesis. We show that destabilization of this interaction, either by alanine exchanges at this NS3/4A-kink interface, led to a gain of function of the NS3/4A complex in particle formation. In contrast, RNA replication and thus replicase assembly requires a stable association between NS3 and the NS4A-kink region. Thus, we propose that two variants of NS3/4A complexes exist in pestivirus infected cells each representing a basic building block required for either RNA replication or virion morphogenesis. This could be further corroborated by trans-complementation studies with a replication-defective NS3/4A double mutant that was still functional in viral assembly. Our observations illustrate the presence of alternative overlapping surfaces providing different contacts between the same proteins, allowing the switch from RNA replication to virion formation.

  12. A positive-strand RNA virus uses alternative protein-protein interactions within a viral protease/cofactor complex to switch between RNA replication and virion morphogenesis

    Science.gov (United States)

    Rey, Félix A.

    2017-01-01

    The viruses of the family Flaviviridae possess a positive-strand RNA genome and express a single polyprotein which is processed into functional proteins. Initially, the nonstructural (NS) proteins, which are not part of the virions, form complexes capable of genome replication. Later on, the NS proteins also play a critical role in virion formation. The molecular basis to understand how the same proteins form different complexes required in both processes is so far unknown. For pestiviruses, uncleaved NS2-3 is essential for virion morphogenesis while NS3 is required for RNA replication but is not functional in viral assembly. Recently, we identified two gain of function mutations, located in the C-terminal region of NS2 and in the serine protease domain of NS3 (NS3 residue 132), which allow NS2 and NS3 to substitute for uncleaved NS2-3 in particle assembly. We report here the crystal structure of pestivirus NS3-4A showing that the NS3 residue 132 maps to a surface patch interacting with the C-terminal region of NS4A (NS4A-kink region) suggesting a critical role of this contact in virion morphogenesis. We show that destabilization of this interaction, either by alanine exchanges at this NS3/4A-kink interface, led to a gain of function of the NS3/4A complex in particle formation. In contrast, RNA replication and thus replicase assembly requires a stable association between NS3 and the NS4A-kink region. Thus, we propose that two variants of NS3/4A complexes exist in pestivirus infected cells each representing a basic building block required for either RNA replication or virion morphogenesis. This could be further corroborated by trans-complementation studies with a replication-defective NS3/4A double mutant that was still functional in viral assembly. Our observations illustrate the presence of alternative overlapping surfaces providing different contacts between the same proteins, allowing the switch from RNA replication to virion formation. PMID:28151973

  13. A positive-strand RNA virus uses alternative protein-protein interactions within a viral protease/cofactor complex to switch between RNA replication and virion morphogenesis.

    Directory of Open Access Journals (Sweden)

    Danilo Dubrau

    2017-02-01

    Full Text Available The viruses of the family Flaviviridae possess a positive-strand RNA genome and express a single polyprotein which is processed into functional proteins. Initially, the nonstructural (NS proteins, which are not part of the virions, form complexes capable of genome replication. Later on, the NS proteins also play a critical role in virion formation. The molecular basis to understand how the same proteins form different complexes required in both processes is so far unknown. For pestiviruses, uncleaved NS2-3 is essential for virion morphogenesis while NS3 is required for RNA replication but is not functional in viral assembly. Recently, we identified two gain of function mutations, located in the C-terminal region of NS2 and in the serine protease domain of NS3 (NS3 residue 132, which allow NS2 and NS3 to substitute for uncleaved NS2-3 in particle assembly. We report here the crystal structure of pestivirus NS3-4A showing that the NS3 residue 132 maps to a surface patch interacting with the C-terminal region of NS4A (NS4A-kink region suggesting a critical role of this contact in virion morphogenesis. We show that destabilization of this interaction, either by alanine exchanges at this NS3/4A-kink interface, led to a gain of function of the NS3/4A complex in particle formation. In contrast, RNA replication and thus replicase assembly requires a stable association between NS3 and the NS4A-kink region. Thus, we propose that two variants of NS3/4A complexes exist in pestivirus infected cells each representing a basic building block required for either RNA replication or virion morphogenesis. This could be further corroborated by trans-complementation studies with a replication-defective NS3/4A double mutant that was still functional in viral assembly. Our observations illustrate the presence of alternative overlapping surfaces providing different contacts between the same proteins, allowing the switch from RNA replication to virion formation.

  14. Somatic mitochondrial DNA mutations in cancer escape purifying selection and high pathogenicity mutations lead to the oncocytic phenotype: pathogenicity analysis of reported somatic mtDNA mutations in tumors

    International Nuclear Information System (INIS)

    Pereira, Luísa; Soares, Pedro; Máximo, Valdemar; Samuels, David C

    2012-01-01

    The presence of somatic mitochondrial DNA (mtDNA) mutations in cancer cells has been interpreted in controversial ways, ranging from random neutral accumulation of mutations, to positive selection for high pathogenicity, or conversely to purifying selection against high pathogenicity variants as occurs at the population level. Here we evaluated the predicted pathogenicity of somatic mtDNA mutations described in cancer and compare these to the distribution of variations observed in the global human population and all possible protein variations that could occur in human mtDNA. We focus on oncocytic tumors, which are clearly associated with mitochondrial dysfunction. The protein variant pathogenicity was predicted using two computational methods, MutPred and SNPs&GO. The pathogenicity score of the somatic mtDNA variants were significantly higher in oncocytic tumors compared to non-oncocytic tumors. Variations in subunits of Complex I of the electron transfer chain were significantly more common in tumors with the oncocytic phenotype, while variations in Complex V subunits were significantly more common in non-oncocytic tumors. Our results show that the somatic mtDNA mutations reported over all tumors are indistinguishable from a random selection from the set of all possible amino acid variations, and have therefore escaped the effects of purifying selection that act strongly at the population level. We show that the pathogenicity of somatic mtDNA mutations is a determining factor for the oncocytic phenotype. The opposite associations of the Complex I and Complex V variants with the oncocytic and non-oncocytic tumors implies that low mitochondrial membrane potential may play an important role in determining the oncocytic phenotype

  15. Flow cytometric characterization of phenotype, DNA indices and p53 gene expression in 55 cases of acute leukemia.

    Science.gov (United States)

    Powari, Manish; Varma, Neelam; Varma, Subhash; Marwaha, Ram Kumar; Sandhu, Harpreet; Ganguly, Nirmal Kumar

    2002-06-01

    To characterize the phenotype of acute leukemia cases using flow cytometry, to detect mixed lineage cases and to use DNA index determination, including S-phase fraction (SPF) and p53 detection, to find if there was any correlation of SPF and p53 expression with outcome. Fifty-five cases of acute leukemia were enrolled in this study. A complete hemogram and routine bone marrow examination, including cytochemistry, was done. Mycloperoxidase-negative cases were evaluated on a flow cytometer using monoclonal antibodies. DNA indices were determined by flow cytometry in all cases, and p53 was detected immunohistochemically using the alkaline phosphatase/antialkaline phosphatase technique. Acute myeloblastic leukemia (AML) was diagnosed in 32 cases; acute lymphoblastic leukemia (ALL) was diagnosed in 18 (14 B lineage and 4 T line age). Four cases showed mixed lineage leukemia, and undifferentiated acute leukemia was diagnosed in one case. The mean/range of SPF for these groups were 3.76/0.33-6.91, 6.25/0.15-21.4, 2.89/0.35-10.64, 2.60/0.72-6.94 and 7.34, respectively. Aneuploidy was detected in two cases of B-lineage ALL and tetraploidy in a case of AML-M7, while all others were diploid p53. Was detected in 6 of 55 cases (10.90%). Follow-up was available for 24 patients. Five patients relapsed, and four had B-cell type ALL and were diploid and expressed no p53 gene. SPF% did not show any correlation with outcome. These data suggest that within acute leukemia subtypes, there is a wide variation in SPF. SPF does not seem to correlate with outcome. Immunophenotyping is essential to determine the lineage in myeloperoxidase-negative cases. It is perhaps the only way to diagnose mixed lineage leukemia and aberrant expression of markers presently. The p53 gene was detected less frequently. However, more studies are required from different centers with longer follow-up to evaluate prognostic significance.

  16. Correlation of pathologic features with CpG island methylator phenotype (CIMP) by quantitative DNA methylation analysis in colorectal carcinoma.

    Science.gov (United States)

    Ogino, Shuji; Odze, Robert D; Kawasaki, Takako; Brahmandam, Mohan; Kirkner, Gregory J; Laird, Peter W; Loda, Massimo; Fuchs, Charles S

    2006-09-01

    Extensive gene promoter methylation in colorectal carcinoma has been termed the CpG island methylator phenotype (CIMP). Previous studies on CIMP used primarily methylation-specific polymerase chain reaction (PCR), which, unfortunately, may detect low levels of methylation that has little or no biological significance. Utilizing quantitative real-time PCR (MethyLight), we measured DNA methylation in a panel of 5 CIMP-specific gene promoters (CACNA1G, CDKN2A (p16), CRABP1, MLH1, and NEUROG1) in 459 colorectal carcinomas obtained from 2 large prospective cohort studies. CIMP was defined as tumors that showed methylation in >or=4/5 promoters. CIMP was significantly associated with the presence of mucinous or signet ring cell morphology, marked Crohn's-like lymphoid reaction, tumor infiltrating lymphocytes, marked peritumoral lymphocytic reaction, tumor necrosis, tumor cell sheeting, and poor differentiation. All these features have previously been associated with microsatellite instability (MSI). Therefore, we divided the 459 colorectal carcinomas into 6 subtypes, namely, MSI-high (MSI-H)/CIMP, MSI-H/non-CIMP, MSI-low (MSI-L)/CIMP, MSI-L/non-CIMP, microsatellite stable/CIMP, and micro satellite sstable/non-CIMP. Compared with MSI-H/non-CIMP, MSI-H/CIMP was associated with marked tumor infiltrating lymphocytes, tumor necrosis, sheeting, and poor differentiation (all PCIMP, MSI-L/CIMP was associated with tumors that had CIMP. Both MSI and CIMP appear to play a role in the pathogenesis of specific morphologic patterns of colorectal carcinoma.

  17. Transfection with extracellularly UV-damaged DNA induces human and rat cells to express a mutator phenotype towards parvovirus H-1

    International Nuclear Information System (INIS)

    Dinsart, C.; Cornelis, J.J.; Klein, B.; van der Eb, A.J.; Rommelaere, J.

    1984-01-01

    Human and rat cells transfected with UV-irradiated linear double-stranded DNA from calf thymus displayed a mutator activity. This phenotype was identified by growing a lytic thermosensitive single-stranded DNA virus (parvovirus H-1) in those cells and determining viral reversion frequencies. Likewise, exogenous UV-irradiated closed circular DNAs, either double-stranded (simian virus 40) or single-stranded (phi X174), enhanced the ability of recipient cells to mutate parvovirus H-1. The magnitude of mutator activity expression increased along with the number of UV lesions present in the inoculated DNA up to a saturation level. Unirradiated DNA displayed little inducing capacity, irrespective of whether it was single or double stranded. Deprivation of a functional replication origin did not impede UV-irradiated simian virus 40 DNA from providing rat and human cells with a mutator function. Our data suggest that in mammalian cells a trans-acting mutagenic signal might be generated from UV-irradiated DNA without the necessity for damaged DNA to replicate

  18. Decreased DNA Methylation in the Shati/Nat8l Promoter in Both Patients with Schizophrenia and a Methamphetamine-Induced Murine Model of Schizophrenia-Like Phenotype.

    Directory of Open Access Journals (Sweden)

    Kyosuke Uno

    Full Text Available The number of patients with schizophrenia has increased over the past decade. Previously, many studies have been performed to establish its diagnostic criteria, prophylactic methods, and effective therapies. In this study, we analyzed whether the ratios of DNA methylation in CpG islands of the Shati/Nat8l is decreased in model mice of schizophrenia-like phenotype using genomic DNA collected from brain regions and peripheral blood, since the mouse model of schizophrenia-like phenotype, mice treated repeatedly with methamphetamine showed increase of Shati/Nat8l mRNA expression in our previous experiment. The ratios of Shati/Nat8l CpG island methylation were significantly decreased in both the nucleus accumbens and the peripheral blood of model mice compared with those of control mice. We also investigated Shati/Nat8l methylation in the blood of patients with schizophrenia. We found that Shati/Nat8l CpG island methylation ratios were lower in the patients with schizophrenia than in the healthy controls, which is consistent with our findings in the mice model. To our knowledge, this is the first study to show similar alterations in methylation status of a particular genomic DNA site in both the brain and peripheral blood of mice. Furthermore, the same phenomenon was observed in corresponding human genomic sequences of the DNA extracted from the peripheral blood of patients with schizophrenia. Based on our findings, DNA methylation profiles of the CpG island of Shati/Nat8l might be a diagnostic biomarker of schizophrenia.

  19. Hypoxia-induced DNA hypermethylation in human pulmonary fibroblasts is associated with Thy-1 promoter methylation and the development of a pro-fibrotic phenotype

    Directory of Open Access Journals (Sweden)

    Robinson Claire M

    2012-08-01

    Full Text Available Abstract Background Pulmonary fibrosis is a debilitating and lethal disease with no effective treatment options. Understanding the pathological processes at play will direct the application of novel therapeutic avenues. Hypoxia has been implicated in the pathogenesis of pulmonary fibrosis yet the precise mechanism by which it contributes to disease progression remains to be fully elucidated. It has been shown that chronic hypoxia can alter DNA methylation patterns in tumour-derived cell lines. This epigenetic alteration can induce changes in cellular phenotype with promoter methylation being associated with gene silencing. Of particular relevance to idiopathic pulmonary fibrosis (IPF is the observation that Thy-1 promoter methylation is associated with a myofibroblast phenotype where loss of Thy-1 occurs alongside increased alpha smooth muscle actin (α-SMA expression. The initial aim of this study was to determine whether hypoxia regulates DNA methylation in normal human lung fibroblasts (CCD19Lu. As it has been reported that hypoxia suppresses Thy-1 expression during lung development we also studied the effect of hypoxia on Thy-1 promoter methylation and gene expression. Methods CCD19Lu were grown for up to 8 days in hypoxia and assessed for global changes in DNA methylation using flow cytometry. Real-time PCR was used to quantify expression of Thy-1, α-SMA, collagen I and III. Genomic DNA was bisulphite treated and methylation specific PCR (MSPCR was used to examine the methylation status of the Thy-1 promoter. Results Significant global hypermethylation was detected in hypoxic fibroblasts relative to normoxic controls and was accompanied by increased expression of myofibroblast markers. Thy-1 mRNA expression was suppressed in hypoxic cells, which was restored with the demethylating agent 5-aza-2′-deoxycytidine. MSPCR revealed that Thy-1 became methylated following fibroblast exposure to 1% O2. Conclusion These data suggest that global and

  20. Genome-wide DNA methylation alterations of Alternanthera philoxeroides in natural and manipulated habitats: implications for epigenetic regulation of rapid responses to environmental fluctuation and phenotypic variation.

    Science.gov (United States)

    Gao, Lexuan; Geng, Yupeng; Li, Bo; Chen, Jiakuan; Yang, Ji

    2010-11-01

    Alternanthera philoxeroides (alligator weed) is an invasive weed that can colonize both aquatic and terrestrial habitats. Individuals growing in different habitats exhibit extensive phenotypic variation but little genetic differentiation in its introduced range. The mechanisms underpinning the wide range of phenotypic variation and rapid adaptation to novel and changing environments remain uncharacterized. In this study, we examined the epigenetic variation and its correlation with phenotypic variation in plants exposed to natural and manipulated environmental variability. Genome-wide methylation profiling using methylation-sensitive amplified fragment length polymorphism (MSAP) revealed considerable DNA methylation polymorphisms within and between natural populations. Plants of different source populations not only underwent significant morphological changes in common garden environments, but also underwent a genome-wide epigenetic reprogramming in response to different treatments. Methylation alterations associated with response to different water availability were detected in 78.2% (169/216) of common garden induced polymorphic sites, demonstrating the environmental sensitivity and flexibility of the epigenetic regulatory system. These data provide evidence of the correlation between epigenetic reprogramming and the reversible phenotypic response of alligator weed to particular environmental factors. © 2010 Blackwell Publishing Ltd.

  1. Complementation of the UV-sensitive phenotype of a xeroderma pigmentosum human cell line by transfection with a cDNA clone library

    International Nuclear Information System (INIS)

    Teitz, T.; Naiman, T.; Avissar, S.S.; Bar, S.; Okayama, H.; Canaani, D.

    1987-01-01

    In previous work, a xeroderma pigmentosum cell line belonging to complementation group C was established by transformation with origin-defective simian virus 40. We now report the complementation of the UV sensitivity of this cell line by gene transfer. A human cDNA clone library constructed in a mammalian expression vector, and itself incorporated in a lambda phage vector, was introduced into the cells as a calcium phosphate precipitate. Following selection to G418 resistance, provided by the neo gene of the vector, transformants were selected for UV resistance. Twenty-one cell clones were obtained with UV-resistance levels typical of normal human fibroblasts. All transformants contained vector DNA sequences in their nuclei. Upon further propagation in the absence of selection for G418 resistance, about half of the primary transformants remained UV-resistant. Secondary transformants were generated by transfection with a partial digest of total chromosomal DNA from one of these stable transformants. This resulted in 15 G418-resistant clones, 2 of which exhibited a UV-resistant phenotype. The other primary clones lost UV resistance rapidly when subcultured in the absence of G418. Importantly, several retained UV resistance under G418 selection pressure. The acquisition of UV resistance by secondary transformants derived by transfection of DNA from a stable primary transformant, and the linkage between G418 and UV resistances in the unstable primary transformants, strongly suggests that the transformants acquired UV resistance through DNA-mediated gene transfer and not by reversion

  2. Dietary restriction delays the secretion of senescence associated secretory phenotype by reducing DNA damage response in the process of renal aging.

    Science.gov (United States)

    Wang, Wenjuan; Cai, Guangyan; Chen, Xiangmei

    2017-09-13

    Dietary restriction (DR) has multiple and essential effects in protecting against DNA damage in model organisms. Persistent DNA damage plays a central role in the process of aging. Senescence-associated secretory phenotype (SASP), as a product of cellular aging, can accelerate the process of cellular senescence as a feedback. In this study, we directly observed whether a DR of 30% for 6months in aged rats could retard SASP by delaying the progression of DNA damage and also found the specific mechanism. The results revealed that a 30% DR could significantly improve renal pathology and some metabolic characteristics. The biomarkers and products of DNA damage were decreased in the process of renal aging on a 30% DR. A series of SASP, notably cytokine, chemokine, and growth factor, were obviously reduced by DR during renal aging. The phosphorylation levels of NF-κB and IκBα in aged kidneys of DR group were markedly reduced. These findings suggest that a 30% DR for 6months can delay renal aging and reduce the accumulation of SASP by retarding the progression of DNA damage and decreasing the transcription activity of NF-κB, thus providing a target to delay renal aging. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Efficient Translation of Epstein-Barr Virus (EBV) DNA Polymerase Contributes to the Enhanced Lytic Replication Phenotype of M81 EBV.

    Science.gov (United States)

    Church, Trenton Mel; Verma, Dinesh; Thompson, Jacob; Swaminathan, Sankar

    2018-03-15

    Epstein-Barr virus (EBV) is linked to the development of both lymphoid and epithelial malignancies worldwide. The M81 strain of EBV, isolated from a Chinese patient with nasopharyngeal carcinoma (NPC), demonstrates spontaneous lytic replication and high-titer virus production in comparison to the prototype B95-8 EBV strain. Genetic comparisons of M81 and B95-8 EBVs were previously been performed in order to determine if the hyperlytic property of M81 is associated with sequence differences in essential lytic genes. EBV SM is an RNA-binding protein expressed during early lytic replication that is essential for virus production. We compared the functions of M81 SM and B95-8 SM and demonstrate that polymorphisms in SM do not contribute to the lytic phenotype of M81 EBV. However, the expression level of the EBV DNA polymerase protein was much higher in M81- than in B95-8-infected cells. The relative deficiency in the expression of B95-8 DNA polymerase was related to the B95-8 genome deletion, which truncates the BALF5 3' untranslated region (UTR). Similarly, the insertion of bacmid DNA into the widely used recombinant B95-8 bacmid creates an inefficient BALF5 3' UTR. We further showed that the while SM is required for and facilitates the efficient expression of both M81 and B95-8 mRNAs regardless of the 3' UTR, the BALF5 3' UTR sequence is important for BALF5 protein translation. These data indicate that the enhanced lytic replication and virus production of M81 compared to those of B95-8 are partly due to the robust translation of EBV DNA polymerase required for viral DNA replication due to a more efficient BALF5 3' UTR in M81. IMPORTANCE Epstein-Barr virus (EBV) infects more than 90% of the human population, but the incidence of EBV-associated tumors varies greatly in different parts of the world. Thus, understanding the connection between genetic polymorphisms from patient isolates of EBV, gene expression phenotypes, and disease is important and may help in

  4. Randomly detected genetically modified (GM maize (Zea mays L. near a transport route revealed a fragile 45S rDNA phenotype.

    Directory of Open Access Journals (Sweden)

    Nomar Espinosa Waminal

    Full Text Available Monitoring of genetically modified (GM crops has been emphasized to prevent their potential effects on the environment and human health. Monitoring of the inadvertent dispersal of transgenic maize in several fields and transport routes in Korea was carried out by qualitative multiplex PCR, and molecular analyses were conducted to identify the events of the collected GM maize. Cytogenetic investigations through fluorescence in situ hybridization (FISH of the GM maize were performed to check for possible changes in the 45S rDNA cluster because this cluster was reported to be sensitive to replication and transcription stress. Three GM maize kernels were collected from a transport route near Incheon port, Korea, and each was found to contain NK603, stacked MON863 x NK603, and stacked NK603 x MON810 inserts, respectively. Cytogenetic analysis of the GM maize containing the stacked NK603 x MON810 insert revealed two normal compact 5S rDNA signals, but the 45S rDNA showed a fragile phenotype, demonstrating a "beads-on-a-string" fragmentation pattern, which seems to be a consequence of genetic modification. Implications of the 45S rDNA cluster fragility in GM maize are also discussed.

  5. Smad mediated regulation of inhibitor of DNA binding 2 and its role in phenotypic maintenance of human renal proximal tubule epithelial cells.

    Directory of Open Access Journals (Sweden)

    Mangalakumar Veerasamy

    Full Text Available The basic-Helix-Loop-Helix family (bHLH of transcriptional factors plays a major role in regulating cellular proliferation, differentiation and phenotype maintenance. The downregulation of one of the members of bHLH family protein, inhibitor of DNA binding 2 (Id2 has been shown to induce de-differentiation of epithelial cells. Opposing regulators of epithelial/mesenchymal phenotype in renal proximal tubule epithelial cells (PTEC, TGFβ1 and BMP7 also have counter-regulatory effects in models of renal fibrosis. We investigated the regulation of Id2 by these growth factors in human PTECs and its implication in the expression of markers of epithelial versus myofibroblastic phenotype. Cellular Id2 levels were reduced by TGFβ1 treatment; this was prevented by co-incubation with BMP7. BMP7 alone increased cellular levels of Id2. TGFβ1 and BMP7 regulated Id2 through Smad2/3 and Smad1/5 dependent mechanisms respectively. TGFβ1 mediated Id2 suppression was essential for α-SMA induction in PTECs. Although Id2 over-expression prevented α-SMA induction, it did not prevent E-cadherin loss under the influence of TGFβ1. This suggests that the loss of gate keeper function of E-cadherin alone may not necessarily result in complete EMT and further transcriptional re-programming is essential to attain mesenchymal phenotype. Although BMP7 abolished TGFβ1 mediated α-SMA expression by restoring Id2 levels, the loss of Id2 was not sufficient to induce α-SMA expression even in the context of reduced E-cadherin expression. Hence, a reduction in Id2 is critical for TGFβ1-induced α-SMA expression in this model of human PTECs but is not sufficient in it self to induce α-SMA even in the context of reduced E-cadherin.

  6. Mitochondrial DNA polymerase editing mutation, PolgD257A, reduces the diabetic phenotype of Akita male mice by suppressing appetite.

    Science.gov (United States)

    Fox, Raymond; Kim, Hyung-Suk; Reddick, Robert L; Kujoth, Gregory C; Prolla, Tomas A; Tsutsumi, Shuichi; Wada, Youichiro; Smithies, Oliver; Maeda, Nobuyo

    2011-05-24

    Diabetes and the development of its complications have been associated with mitochondrial DNA (mtDNA) dysfunction, but causal relationships remain undetermined. With the objective of testing whether increased mtDNA mutations exacerbate the diabetic phenotype, we have compared mice heterozygous for the Akita diabetogenic mutation (Akita) with mice homozygous for the D257A mutation in mitochondrial DNA polymerase gamma (Polg) or with mice having both mutations (Polg-Akita). The Polg-D257A protein is defective in proofreading and increases mtDNA mutations. At 3 mo of age, the Polg-Akita and Akita male mice were equally hyperglycemic. Unexpectedly, as the Polg-Akita males aged to 9 mo, their diabetic symptoms decreased. Thus, their hyperglycemia, hyperphagia and urine output declined significantly. The decrease in their food intake was accompanied by increased plasma leptin and decreased plasma ghrelin, while hypothalamic expression of the orexic gene, neuropeptide Y, was lower and expression of the anorexic gene, proopiomelanocortin, was higher. Testis function progressively worsened with age in the double mutants, and plasma testosterone levels in 9-mo-old Polg-Akita males were significantly reduced compared with Akita males. The hyperglycemia and hyperphagia returned in aged Polg-Akita males after testosterone administration. Hyperglycemia-associated distal tubular damage in the kidney also returned, and Polg-D257A-associated proximal tubular damage was enhanced. The mild diabetes of female Akita mice was not affected by the Polg-D257A mutation. We conclude that reduced diabetic symptoms of aging Polg-Akita males results from appetite suppression triggered by decreased testosterone associated with damage to the Leydig cells of the testis.

  7. Integrative DNA methylation and gene expression analysis to assess the universality of the CpG island methylator phenotype.

    Science.gov (United States)

    Moarii, Matahi; Reyal, Fabien; Vert, Jean-Philippe

    2015-10-13

    The CpG island methylator phenotype (CIMP) was first characterized in colorectal cancer but since has been extensively studied in several other tumor types such as breast, bladder, lung, and gastric. CIMP is of clinical importance as it has been reported to be associated with prognosis or response to treatment. However, the identification of a universal molecular basis to define CIMP across tumors has remained elusive. We perform a genome-wide methylation analysis of over 2000 tumor samples from 5 cancer sites to assess the existence of a CIMP with common molecular basis across cancers. We then show that the CIMP phenotype is associated with specific gene expression variations. However, we do not find a common genetic signature in all tissues associated with CIMP. Our results suggest the existence of a universal epigenetic and transcriptomic signature that defines the CIMP across several tumor types but does not indicate the existence of a common genetic signature of CIMP.

  8. The mitochondrial DNA T16189C polymorphism and HIV-associated cardiomyopathy: a genotype-phenotype association study

    Directory of Open Access Journals (Sweden)

    Poulton Joanna

    2009-04-01

    Full Text Available Abstract Background The mitochondrial DNA (mtDNA T16189C polymorphism, with a homopolymeric C-tract of 10–12 cytosines, is a putative genetic risk factor for idiopathic dilated cardiomyopathy in the African and British populations. We hypothesized that this variant may predispose to dilated cardiomyopathy in people who are infected with the human immunodeficiency virus (HIV. Methods A case-control study of 30 HIV-positive cases with dilated cardiomyopathy and 37 HIV-positive controls without dilated cardiomyopathy was conducted. The study was confined to persons of black African ancestry to minimize confounding of results by population admixture. HIV-positive patients with an echocardiographically confirmed diagnosis of dilated cardiomyopathy and HIV-positive controls with echocardiographically normal hearts were studied. Patients with secondary causes of cardiomyopathy (such as hypertension, diabetes, pregnancy, alcoholism, valvular heart disease, and opportunistic infection were excluded from the study. DNA samples were sequenced for the mtDNA T16189C polymorphism with a homopolymeric C-tract in the forward and reverse directions on an ABI3100 sequencer. Results The cases and controls were well matched for age (median 35 years versus 34 years, P = 0.93, gender (males 60% vs 53%, P = 0.54, and stage of HIV disease (mean CD4 T cell count 260.7/μL vs. 176/μL, P = 0.21. The mtDNA T16189C variant with a homopolymeric C-tract was detected at a frequency of 26.7% (8/30 in the HIV-associated cardiomyopathy cases and 13.5% (5/37 in the HIV-positive controls. There was no significant difference between cases and controls (Odds Ratio 2.33, 95% Confidence Interval 0.67–8.06, p = 0.11. Conclusion The mtDNA T16189C variant with a homopolymeric C-tract is not associated with dilated cardiomyopathy in black African people infected with HIV.

  9. CpG island methylator phenotype (CIMP) of colorectal cancer is best characterised by quantitative DNA methylation analysis and prospective cohort studies.

    Science.gov (United States)

    Ogino, S; Cantor, M; Kawasaki, T; Brahmandam, M; Kirkner, G J; Weisenberger, D J; Campan, M; Laird, P W; Loda, M; Fuchs, C S

    2006-07-01

    The concept of CpG island methylator phenotype (CIMP) is not universally accepted. Even if specific clinicopathological features have been associated with CIMP, investigators often failed to demonstrate a bimodal distribution of the number of methylated markers, which would suggest CIMP as a distinct subtype of colorectal cancer. Previous studies primarily used methylation specific polymerase chain reaction which might detect biologically insignificant low levels of methylation. To demonstrate a distinct genetic profile of CIMP colorectal cancer using quantitative DNA methylation analysis that can distinguish high from low levels of DNA methylation. We developed quantitative real time polymerase chain reaction (MethyLight) assays and measured DNA methylation (percentage of methylated reference) of five carefully selected loci (promoters of CACNA1G, CDKN2A (p16), CRABP1, MLH1, and NEUROG1) in 460 colorectal cancers from large prospective cohorts. There was a clear bimodal distribution of 80 microsatellite instability-high (MSI-H) tumours according to the number of methylated promoters, with no tumours showing 3/5 methylated loci. Thus we defined CIMP as having >or=4/5 methylated loci, and 17% (78) of the 460 tumours were classified as CIMP. CIMP was significantly associated with female sex, MSI, BRAF mutations, and wild-type KRAS. Both CIMP MSI-H tumours and CIMP microsatellite stable (MSS) tumours showed much higher frequencies of BRAF mutations (63% and 54%) than non-CIMP counterparts (non-CIMP MSI-H (0%, pCIMP MSS tumours (6.6%, pCIMP is best characterised by quantitative DNA methylation analysis. CIMP is a distinct epigenotype of colorectal cancer and may be less frequent than previously reported.

  10. A KAS2 cDNA complements the phenotypes of the Arabidopsis fab1 mutant that differs in a single residue bordering the substrate binding pocket

    DEFF Research Database (Denmark)

    Carlsson, A.S.; LaBrie, S.T.; Kinney, A.J.

    2002-01-01

    The fab1 mutant of Arabidopsis is partially deficient in activity of ß-ketoacyl-[acyl carrier protein] synthase II (KAS II). This defect results in increased levels of 16 : 0 fatty acid and is associated with damage and death of the mutants at low temperature. Transformation of fab1 plants with a c......DNA from Brassica napus encoding a KAS II enzyme resulted in complementation of both mutant phenotypes. The dual complementation by expression of the single gene proves that low-temperature damage is a consequence of altered membrane unsaturation. The fab1 mutation is a single nucleotide change...... chain to bend. For functional analysis the equivalent Leu207Phe mutation was introduced into the fabB gene encoding the E. coli KAS I enzyme. Compared to wild-type, the Leu207Phe protein showed a 10-fold decrease in binding affinity for the fatty acid substrate, exhibited a modified behavior during size...

  11. Natural variation in life history and aging phenotypes is associated with mitochondrial DNA deletion frequency in Caenorhabditis briggsae

    Directory of Open Access Journals (Sweden)

    Smith Samson W

    2011-01-01

    Full Text Available Abstract Background Mutations that impair mitochondrial functioning are associated with a variety of metabolic and age-related disorders. A barrier to rigorous tests of the role of mitochondrial dysfunction in aging processes has been the lack of model systems with relevant, naturally occurring mitochondrial genetic variation. Toward the goal of developing such a model system, we studied natural variation in life history, metabolic, and aging phenotypes as it relates to levels of a naturally-occurring heteroplasmic mitochondrial ND5 deletion recently discovered to segregate among wild populations of the soil nematode, Caenorhabditis briggsae. The normal product of ND5 is a central component of the mitochondrial electron transport chain and integral to cellular energy metabolism. Results We quantified significant variation among C. briggsae isolates for all phenotypes measured, only some of which was statistically associated with isolate-specific ND5 deletion frequency. We found that fecundity-related traits and pharyngeal pumping rate were strongly inversely related to ND5 deletion level and that C. briggsae isolates with high ND5 deletion levels experienced a tradeoff between early fecundity and lifespan. Conversely, oxidative stress resistance was only weakly associated with ND5 deletion level while ATP content was unrelated to deletion level. Finally, mean levels of reactive oxygen species measured in vivo showed a significant non-linear relationship with ND5 deletion level, a pattern that may be driven by among-isolate variation in antioxidant or other compensatory mechanisms. Conclusions Our findings suggest that the ND5 deletion may adversely affect fitness and mitochondrial functioning while promoting aging in natural populations, and help to further establish this species as a useful model for explicit tests of hypotheses in aging biology and mitochondrial genetics.

  12. Phenotyping of VIGS-mediated gene silencing in rice using a vector derived from a DNA virus.

    Science.gov (United States)

    Kant, Ravi; Dasgupta, Indranil

    2017-07-01

    Target genes in rice can be optimally silenced if inserted in antisense or hairpin orientation in the RTBV-derived VIGS vector and plants grown at 28 °C and 80% humidity after inoculation. Virus induced gene silencing (VIGS) is a method used to transiently silence genes in dicot as well as monocot plants. For the important monocot species rice, the Rice tungro bacilliform virus (RTBV)-derived VIGS system (RTBV-VIGS), which uses agroinoculation to initiate silencing, has not been standardized for optimal use. Here, using RTBV-VIGS, three sets of conditions were tested to achieve optimal silencing of the rice marker gene phytoene desaturase (pds). The effect of orientation of the insert in the RTBV-VIGS plasmid (sense, antisense and hairpin) on the silencing of the target gene was then evaluated using rice magnesium chelatase subunit H (chlH). Finally, the rice Xa21 gene, conferring resistance against bacterial leaf blight disease (BLB) was silenced using RTBV-VIGS system. In each case, real-time PCR-based assessment indicated approximately 40-80% fall in the accumulation levels of the transcripts of pds, chlH and Xa21. In the case of pds, the appearance of white streaks in the emerging leaves, and for chlH, chlorophyll levels and F v /F m ratio were assessed as phenotypes for silencing. For Xa21, the resistance levels to BLB were assessed by measuring the lesion length and the percent diseased areas of leaves, following challenge inoculation with Xanthomonas oryzae. In each case, the RTBV-MVIGS system gave rise to a discernible phenotype indicating the silencing of the respective target gene using condition III (temperature 28 °C, humidity 80% and 1 mM MES and 20 µM acetosyringone in secondary agrobacterium culture), which revealed the robustness of this gene silencing system for rice.

  13. HDAC4 and HDAC6 sustain DNA double strand break repair and stem-like phenotype by promoting radioresistance in glioblastoma cells.

    Science.gov (United States)

    Marampon, Francesco; Megiorni, Francesca; Camero, Simona; Crescioli, Clara; McDowell, Heather P; Sferra, Roberta; Vetuschi, Antonella; Pompili, Simona; Ventura, Luca; De Felice, Francesca; Tombolini, Vincenzo; Dominici, Carlo; Maggio, Roberto; Festuccia, Claudio; Gravina, Giovanni Luca

    2017-07-01

    The role of histone deacetylase (HDAC) 4 and 6 in glioblastoma (GBM) radioresistance was investigated. We found that tumor samples from 31 GBM patients, who underwent temozolomide and radiotherapy combined treatment, showed HDAC4 and HDAC6 expression in 93.5% and 96.7% of cases, respectively. Retrospective clinical data analysis demonstrated that high-intensity HDAC4 and/or HDAC6 immunostaining was predictive of poor clinical outcome. In vitro experiments revealed that short hairpin RNA-mediated silencing of HDAC4 or HDAC6 radiosensitized U87MG and U251MG GBM cell lines by promoting DNA double-strand break (DSBs) accumulation and by affecting DSBs repair molecular machinery. We found that HDAC6 knock-down predisposes to radiation therapy-induced U251MG apoptosis- and U87MG autophagy-mediated cell death. HDAC4 silencing promoted radiation therapy-induced senescence, independently by the cellular context. Finally, we showed that p53 WT expression contributed to the radiotherapy lethal effects and that HDAC4 or HDAC6 sustained GBM stem-like radioresistant phenotype. Altogether, these observations suggest that HDAC4 and HDAC6 are guardians of irradiation-induced DNA damages and stemness, thus promoting radioresistance, and may represent potential prognostic markers and therapeutic targets in GBM. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Mechanistically Distinct Pathways of Divergent Regulatory DNA Creation Contribute to Evolution of Human-Specific Genomic Regulatory Networks Driving Phenotypic Divergence of Homo sapiens.

    Science.gov (United States)

    Glinsky, Gennadi V

    2016-09-19

    Thousands of candidate human-specific regulatory sequences (HSRS) have been identified, supporting the hypothesis that unique to human phenotypes result from human-specific alterations of genomic regulatory networks. Collectively, a compendium of multiple diverse families of HSRS that are functionally and structurally divergent from Great Apes could be defined as the backbone of human-specific genomic regulatory networks. Here, the conservation patterns analysis of 18,364 candidate HSRS was carried out requiring that 100% of bases must remap during the alignments of human, chimpanzee, and bonobo sequences. A total of 5,535 candidate HSRS were identified that are: (i) highly conserved in Great Apes; (ii) evolved by the exaptation of highly conserved ancestral DNA; (iii) defined by either the acceleration of mutation rates on the human lineage or the functional divergence from non-human primates. The exaptation of highly conserved ancestral DNA pathway seems mechanistically distinct from the evolution of regulatory DNA segments driven by the species-specific expansion of transposable elements. Genome-wide proximity placement analysis of HSRS revealed that a small fraction of topologically associating domains (TADs) contain more than half of HSRS from four distinct families. TADs that are enriched for HSRS and termed rapidly evolving in humans TADs (revTADs) comprise 0.8-10.3% of 3,127 TADs in the hESC genome. RevTADs manifest distinct correlation patterns between placements of human accelerated regions, human-specific transcription factor-binding sites, and recombination rates. There is a significant enrichment within revTAD boundaries of hESC-enhancers, primate-specific CTCF-binding sites, human-specific RNAPII-binding sites, hCONDELs, and H3K4me3 peaks with human-specific enrichment at TSS in prefrontal cortex neurons (P sapiens is driven by the evolution of human-specific genomic regulatory networks via at least two mechanistically distinct pathways of creation of

  15. Tumor cell heterogeneity in Small Cell Lung Cancer (SCLC: phenotypical and functional differences associated with Epithelial-Mesenchymal Transition (EMT and DNA methylation changes.

    Directory of Open Access Journals (Sweden)

    Alexander Krohn

    Full Text Available Small Cell Lung Cancer (SCLC is a specific subtype of lung cancer presenting as highly metastatic disease with extremely poor prognosis. Despite responding initially well to chemo- or radiotherapy, SCLC almost invariably relapses and develops resistance to chemotherapy. This is suspected to be related to tumor cell subpopulations with different characteristics resembling stem cells. Epithelial-Mesenchymal Transition (EMT is known to play a key role in metastatic processes and in developing drug resistance. This is also true for NSCLC, but there is very little information on EMT processes in SCLC so far. SCLC, in contrast to NSCLC cell lines, grow mainly in floating cell clusters and a minor part as adherent cells. We compared these morphologically different subpopulations of SCLC cell lines for EMT and epigenetic features, detecting significant differences in the adherent subpopulations with high levels of mesenchymal markers such as Vimentin and Fibronectin and very low levels of epithelial markers like E-cadherin and Zona Occludens 1. In addition, expression of EMT-related transcription factors such as Snail/Snai1, Slug/Snai2, and Zeb1, DNA methylation patterns of the EMT hallmark genes, functional responses like migration, invasion, matrix metalloproteases secretion, and resistance to chemotherapeutic drug treatment all differed significantly between the sublines. This phenotypic variability might reflect tumor cell heterogeneity and EMT during metastasis in vivo, accompanied by the development of refractory disease in relapse. We propose that epigenetic regulation plays a key role during phenotypical and functional changes in tumor cells and might therefore provide new treatment options for SCLC patients.

  16. Cattle phenotypes can disguise their maternal ancestry.

    Science.gov (United States)

    Srirattana, Kanokwan; McCosker, Kieren; Schatz, Tim; St John, Justin C

    2017-06-26

    Cattle are bred for, amongst other factors, specific traits, including parasite resistance and adaptation to climate. However, the influence and inheritance of mitochondrial DNA (mtDNA) are not usually considered in breeding programmes. In this study, we analysed the mtDNA profiles of cattle from Victoria (VIC), southern Australia, which is a temperate climate, and the Northern Territory (NT), the northern part of Australia, which has a tropical climate, to determine if the mtDNA profiles of these cattle are indicative of breed and phenotype, and whether these profiles are appropriate for their environments. A phylogenetic tree of the full mtDNA sequences of different breeds of cattle, which were obtained from the NCBI database, showed that the mtDNA profiles of cattle do not always reflect their phenotype as some cattle with Bos taurus phenotypes had Bos indicus mtDNA, whilst some cattle with Bos indicus phenotypes had Bos taurus mtDNA. Using D-loop sequencing, we were able to contrast the phenotypes and mtDNA profiles from different species of cattle from the 2 distinct cattle breeding regions of Australia. We found that 67 of the 121 cattle with Bos indicus phenotypes from NT (55.4%) had Bos taurus mtDNA. In VIC, 92 of the 225 cattle with Bos taurus phenotypes (40.9%) possessed Bos indicus mtDNA. When focusing on oocytes from cattle with the Bos taurus phenotype in VIC, their respective oocytes with Bos indicus mtDNA had significantly lower levels of mtDNA copy number compared with oocytes possessing Bos taurus mtDNA (P cattle with a Bos taurus phenotype. The phenotype of cattle is not always related to their mtDNA profiles. MtDNA profiles should be considered for breeding programmes as they also influence phenotypic traits and reproductive capacity in terms of oocyte quality.

  17. Partial sleep deprivation activates the DNA damage response (DDR) and the senescence-associated secretory phenotype (SASP) in aged adult humans.

    Science.gov (United States)

    Carroll, Judith E; Cole, Steven W; Seeman, Teresa E; Breen, Elizabeth C; Witarama, Tuff; Arevalo, Jesusa M G; Ma, Jeffrey; Irwin, Michael R

    2016-01-01

    Age-related disease risk has been linked to short sleep duration and sleep disturbances; however, the specific molecular pathways linking sleep loss with diseases of aging are poorly defined. Key cellular events seen with aging, which are thought to contribute to disease, may be particularly sensitive to sleep loss. We tested whether one night of partial sleep deprivation (PSD) would increase leukocyte gene expression indicative of DNA damage responses (DDR), the senescence-associated secretory phenotype (SASP), and senescence indicator p16(INK4a) in older adult humans, who are at increased risk for cellular senescence. Community-dwelling older adults aged 61-86years (n=29; 48% male) underwent an experimental partial sleep deprivation (PSD) protocol over 4 nights, including adaptation, an uninterrupted night of sleep, partial sleep deprivation (sleep restricted 3-7AM), and a subsequent full night of sleep. Blood samples were obtained each morning to assess peripheral blood mononuclear cell (PBMC) gene expression using Illumina HT-12 arrays. Analyses of microarray results revealed that SASP (psleep deprivation activates PBMC gene expression patterns consistent with biological aging in this older adult sample. PSD enhanced the SASP and increased the accumulation of damage that initiates cell cycle arrest and promotes cellular senescence. These findings causally link sleep deprivation to the molecular processes associated with biological aging. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Lithium Chloride Dependent Glycogen Synthase Kinase 3 Inactivation Links Oxidative DNA Damage, Hypertrophy and Senescence in Human Articular Chondrocytes and Reproduces Chondrocyte Phenotype of Obese Osteoarthritis Patients.

    Directory of Open Access Journals (Sweden)

    Serena Guidotti

    Full Text Available Recent evidence suggests that GSK3 activity is chondroprotective in osteoarthritis (OA, but at the same time, its inactivation has been proposed as an anti-inflammatory therapeutic option. Here we evaluated the extent of GSK3β inactivation in vivo in OA knee cartilage and the molecular events downstream GSK3β inactivation in vitro to assess their contribution to cell senescence and hypertrophy.In vivo level of phosphorylated GSK3β was analyzed in cartilage and oxidative damage was assessed by 8-oxo-deoxyguanosine staining. The in vitro effects of GSK3β inactivation (using either LiCl or SB216763 were evaluated on proliferating primary human chondrocytes by combined confocal microscopy analysis of Mitotracker staining and reactive oxygen species (ROS production (2',7'-dichlorofluorescin diacetate staining. Downstream effects on DNA damage and senescence were investigated by western blot (γH2AX, GADD45β and p21, flow cytometric analysis of cell cycle and light scattering properties, quantitative assessment of senescence associated β galactosidase activity, and PAS staining.In vivo chondrocytes from obese OA patients showed higher levels of phosphorylated GSK3β, oxidative damage and expression of GADD45β and p21, in comparison with chondrocytes of nonobese OA patients. LiCl mediated GSK3β inactivation in vitro resulted in increased mitochondrial ROS production, responsible for reduced cell proliferation, S phase transient arrest, and increase in cell senescence, size and granularity. Collectively, western blot data supported the occurrence of a DNA damage response leading to cellular senescence with increase in γH2AX, GADD45β and p21. Moreover, LiCl boosted 8-oxo-dG staining, expression of IKKα and MMP-10.In articular chondrocytes, GSK3β activity is required for the maintenance of proliferative potential and phenotype. Conversely, GSK3β inactivation, although preserving chondrocyte survival, results in functional impairment via

  19. Lithium Chloride Dependent Glycogen Synthase Kinase 3 Inactivation Links Oxidative DNA Damage, Hypertrophy and Senescence in Human Articular Chondrocytes and Reproduces Chondrocyte Phenotype of Obese Osteoarthritis Patients.

    Science.gov (United States)

    Guidotti, Serena; Minguzzi, Manuela; Platano, Daniela; Cattini, Luca; Trisolino, Giovanni; Mariani, Erminia; Borzì, Rosa Maria

    2015-01-01

    Recent evidence suggests that GSK3 activity is chondroprotective in osteoarthritis (OA), but at the same time, its inactivation has been proposed as an anti-inflammatory therapeutic option. Here we evaluated the extent of GSK3β inactivation in vivo in OA knee cartilage and the molecular events downstream GSK3β inactivation in vitro to assess their contribution to cell senescence and hypertrophy. In vivo level of phosphorylated GSK3β was analyzed in cartilage and oxidative damage was assessed by 8-oxo-deoxyguanosine staining. The in vitro effects of GSK3β inactivation (using either LiCl or SB216763) were evaluated on proliferating primary human chondrocytes by combined confocal microscopy analysis of Mitotracker staining and reactive oxygen species (ROS) production (2',7'-dichlorofluorescin diacetate staining). Downstream effects on DNA damage and senescence were investigated by western blot (γH2AX, GADD45β and p21), flow cytometric analysis of cell cycle and light scattering properties, quantitative assessment of senescence associated β galactosidase activity, and PAS staining. In vivo chondrocytes from obese OA patients showed higher levels of phosphorylated GSK3β, oxidative damage and expression of GADD45β and p21, in comparison with chondrocytes of nonobese OA patients. LiCl mediated GSK3β inactivation in vitro resulted in increased mitochondrial ROS production, responsible for reduced cell proliferation, S phase transient arrest, and increase in cell senescence, size and granularity. Collectively, western blot data supported the occurrence of a DNA damage response leading to cellular senescence with increase in γH2AX, GADD45β and p21. Moreover, LiCl boosted 8-oxo-dG staining, expression of IKKα and MMP-10. In articular chondrocytes, GSK3β activity is required for the maintenance of proliferative potential and phenotype. Conversely, GSK3β inactivation, although preserving chondrocyte survival, results in functional impairment via induction of

  20. Characterization of DNA repair phenotypes of Xeroderma pigmentosum cell lines by a paralleled in vitro test; Phenotypage de la reparation de l'ADN de lignees Xeroderma pigmentosum, par un test in vitro multiparametrique

    Energy Technology Data Exchange (ETDEWEB)

    Raffin, A.L.

    2009-06-15

    DNA is constantly damaged modifying the genetic information for which it encodes. Several cellular mechanisms as the Base Excision Repair (BER) and the Nucleotide Excision Repair (NER) allow recovering the right DNA sequence. The Xeroderma pigmentosum is a disease characterised by a deficiency in the NER pathway. The aim of this study was to propose an efficient and fast test for the diagnosis of this disease as an alternative to the currently available UDS test. DNA repair activities of XP cell lines were quantified using in vitro miniaturized and paralleled tests in order to establish DNA repair phenotypes of XPA and XPC deficient cells. The main advantage of the tests used in this study is the simultaneous measurement of excision or excision synthesis (ES) of several lesions by only one cellular extract. We showed on one hand that the relative ES of the different lesions depend strongly on the protein concentration of the nuclear extract tested. Working at high protein concentration allowed discriminating the XP phenotype versus the control one, whereas it was impossible under a certain concentration's threshold. On the other hand, while the UVB irradiation of control cells stimulated their repair activities, this effect was not observed in XP cells. This study brings new information on the XPA and XPC protein roles during BER and NER and underlines the complexity of the regulations of DNA repair processes. (author)

  1. Prognostication of patients with clear cell renal cell carcinomas based on quantification of DNA methylation levels of CpG island methylator phenotype marker genes.

    Science.gov (United States)

    Tian, Ying; Arai, Eri; Gotoh, Masahiro; Komiyama, Motokiyo; Fujimoto, Hiroyuki; Kanai, Yae

    2014-10-20

    The CpG island methylator phenotype (CIMP) of clear cell renal cell carcinomas (ccRCCs) is characterized by accumulation of DNA methylation at CpG islands and poorer patient outcome. The aim of this study was to establish criteria for prognostication of patients with ccRCCs using the ccRCC-specific CIMP marker genes. DNA methylation levels at 299 CpG sites in the 14 CIMP marker genes were evaluated quantitatively in tissue specimens of 88 CIMP-negative and 14 CIMP-positive ccRCCs in a learning cohort using the MassARRAY system. An additional 100 ccRCCs were also analyzed as a validation cohort. Receiver operating characteristic curve analysis showed that area under the curve values for the 23 CpG units including the 32 CpG sites in the 7 CIMP-marker genes, i.e. FAM150A, ZNF540, ZNF671, ZNF154, PRAC, TRH and SLC13A5, for discrimination of CIMP-positive from CIMP-negative ccRCCs were larger than 0.95. Criteria combining the 23 CpG units discriminated CIMP-positive from CIMP-negative ccRCCs with 100% sensitivity and specificity in the learning cohort. Cancer-free and overall survival rates of patients with CIMP-positive ccRCCs diagnosed using the criteria combining the 23 CpG units in a validation cohort were significantly lower than those of patients with CIMP-negative ccRCCs (P = 1.41 × 10-5 and 2.43 × 10-13, respectively). Patients with CIMP-positive ccRCCs in the validation cohort had a higher likelihood of disease-related death (hazard ratio, 75.8; 95% confidence interval, 7.81 to 735; P = 1.89 × 10-4) than those with CIMP-negative ccRCCs. The established criteria are able to reproducibly diagnose CIMP-positive ccRCCs and may be useful for personalized medicine for patients with ccRCCs.

  2. Comparison of the accuracy of two conventional phenotypic methods and two MALDI-TOF MS systems with that of DNA sequencing analysis for correctly identifying clinically encountered yeasts.

    Science.gov (United States)

    Chao, Qiao-Ting; Lee, Tai-Fen; Teng, Shih-Hua; Peng, Li-Yun; Chen, Ping-Hung; Teng, Lee-Jene; Hsueh, Po-Ren

    2014-01-01

    We assessed the accuracy of species-level identification of two commercially available matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) systems (Bruker Biotyper and Vitek MS) and two conventional phenotypic methods (Phoenix 100 YBC and Vitek 2 Yeast ID) with that of rDNA gene sequencing analysis among 200 clinical isolates of commonly encountered yeasts. The correct identification rates of the 200 yeast isolates to species or complex (Candida parapsilosis complex, C. guilliermondii complex and C. rugosa complex) levels by the Bruker Biotyper, Vitek MS (using in vitro devices [IVD] database), Phoenix 100 YBC and Vitek 2 Yeast ID (Sabouraud's dextrose agar) systems were 92.5%, 79.5%, 89%, and 74%, respectively. An additional 72 isolates of C. parapsilosis complex and 18 from the above 200 isolates (30 in each of C. parapsilosis, C. metapsilosis, and C. orthopsilosis) were also evaluated separately. Bruker Biotyper system could accurately identify all C. parapsilosis complex to species level. Using Vitek 2 MS (IVD) system, all C. parapsilosis but none of C. metapsilosis, or C. orthopsilosis could be accurately identified. Among the 89 yeasts misidentified by the Vitek 2 MS (IVD) system, 39 (43.8%), including 27 C. orthopsilosis isolates, could be correctly identified Using the Vitek MS Plus SARAMIS database for research use only. This resulted in an increase in the rate of correct identification of all yeast isolates (87.5%) by Vitek 2 MS. The two species in C. guilliermondii complex (C. guilliermondii and C. fermentati) isolates were correctly identified by cluster analysis of spectra generated by the Bruker Biotyper system. Based on the results obtained in the current study, MALDI-TOF MS systems present a promising alternative for the routine identification of yeast species, including clinically commonly and rarely encountered yeast species and several species belonging to C. parapsilosis complex, C. guilliermondii complex

  3. Comparison of the accuracy of two conventional phenotypic methods and two MALDI-TOF MS systems with that of DNA sequencing analysis for correctly identifying clinically encountered yeasts.

    Directory of Open Access Journals (Sweden)

    Qiao-Ting Chao

    Full Text Available We assessed the accuracy of species-level identification of two commercially available matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS systems (Bruker Biotyper and Vitek MS and two conventional phenotypic methods (Phoenix 100 YBC and Vitek 2 Yeast ID with that of rDNA gene sequencing analysis among 200 clinical isolates of commonly encountered yeasts. The correct identification rates of the 200 yeast isolates to species or complex (Candida parapsilosis complex, C. guilliermondii complex and C. rugosa complex levels by the Bruker Biotyper, Vitek MS (using in vitro devices [IVD] database, Phoenix 100 YBC and Vitek 2 Yeast ID (Sabouraud's dextrose agar systems were 92.5%, 79.5%, 89%, and 74%, respectively. An additional 72 isolates of C. parapsilosis complex and 18 from the above 200 isolates (30 in each of C. parapsilosis, C. metapsilosis, and C. orthopsilosis were also evaluated separately. Bruker Biotyper system could accurately identify all C. parapsilosis complex to species level. Using Vitek 2 MS (IVD system, all C. parapsilosis but none of C. metapsilosis, or C. orthopsilosis could be accurately identified. Among the 89 yeasts misidentified by the Vitek 2 MS (IVD system, 39 (43.8%, including 27 C. orthopsilosis isolates, could be correctly identified Using the Vitek MS Plus SARAMIS database for research use only. This resulted in an increase in the rate of correct identification of all yeast isolates (87.5% by Vitek 2 MS. The two species in C. guilliermondii complex (C. guilliermondii and C. fermentati isolates were correctly identified by cluster analysis of spectra generated by the Bruker Biotyper system. Based on the results obtained in the current study, MALDI-TOF MS systems present a promising alternative for the routine identification of yeast species, including clinically commonly and rarely encountered yeast species and several species belonging to C. parapsilosis complex, C. guilliermondii

  4. Novel de novo variant in EBF3 is likely to impact DNA binding in a patient with a neurodevelopmental disorder and expanded phenotypes: patient report, in silico functional assessment, and review of published cases.

    Science.gov (United States)

    Blackburn, Patrick R; Barnett, Sarah S; Zimmermann, Michael T; Cousin, Margot A; Kaiwar, Charu; Pinto E Vairo, Filippo; Niu, Zhiyv; Ferber, Matthew J; Urrutia, Raul A; Selcen, Duygu; Klee, Eric W; Pichurin, Pavel N

    2017-05-01

    Pathogenic variants in EBF3 were recently described in three back-to-back publications in association with a novel neurodevelopmental disorder characterized by intellectual disability, speech delay, ataxia, and facial dysmorphisms. In this report, we describe an additional patient carrying a de novo missense variant in EBF3 (c.487C>T, p.(Arg163Trp)) that falls within a conserved residue in the zinc knuckle motif of the DNA binding domain. Without a solved structure of the DNA binding domain, we generated a homology-based atomic model and performed molecular dynamics simulations for EBF3, which predicted decreased DNA affinity for p.(Arg163Trp) compared with wild-type protein and control variants. These data are in agreement with previous experimental studies of EBF1 showing the paralogous residue is essential for DNA binding. The conservation and experimental evidence existing for EBF1 and in silico modeling and dynamics simulations to validate comparable behavior of multiple variants in EBF3 demonstrates strong support for the pathogenicity of p.(Arg163Trp). We show that our patient presents with phenotypes consistent with previously reported patients harboring EBF3 variants and expands the phenotypic spectrum of this newly identified disorder with the additional feature of a bicornuate uterus.

  5. HIV-1 Env DNA vaccine plus protein boost delivered by EP expands B- and T-cell responses and neutralizing phenotype in vivo.

    Directory of Open Access Journals (Sweden)

    Kar Muthumani

    Full Text Available An effective HIV vaccine will most likely require the induction of strong T-cell responses, broadly neutralizing antibodies (bNAbs, and the elicitation of antibody-dependent cellular cytotoxicity (ADCC. Previously, we demonstrated the induction of strong HIV/SIV cellular immune responses in macaques and humans using synthetic consensus DNA immunogens delivered via adaptive electroporation (EP. However, the ability of this improved DNA approach to prime for relevant antibody responses has not been previously studied. Here, we investigate the immunogenicity of consensus DNA constructs encoding gp140 sequences from HIV-1 subtypes A, B, C and D in a DNA prime-protein boost vaccine regimen. Mice and guinea pigs were primed with single- and multi-clade DNA via EP and boosted with recombinant gp120 protein. Sera were analyzed for gp120 binding and induction of neutralizing antibody activity. Immunization with recombinant Env protein alone induced low-titer binding antibodies with limited neutralization breath. In contrast, the synthetic DNA prime-protein boost protocol induced significantly higher antibody binding titers. Furthermore, sera from DNA prime-protein boost groups were able to neutralize a broader range of viruses in a panel of tier 1 clade B viruses as well as multiple tier 1 clade A and clade C viruses. Further investigation of synthetic DNA prime plus adaptive EP plus protein boost appears warranted.

  6. Aging and photo-aging DNA repair phenotype of skin cells-Evidence toward an effect of chronic sun-exposure

    Energy Technology Data Exchange (ETDEWEB)

    Prunier, Chloe; Masson-Genteuil, Gwenaeelle [Laboratoire Lesions des Acides Nucleiques, CEA, DSM, INAC, SCIB, UMR-E CEA/UJF-Grenoble 1, 17 Rue des Martyrs, F-38054 Grenoble Cedex 9 (France); Ugolin, Nicolas [Laboratoire de Cancerologie Experimentale, CEA, DSV, IRCM, SREIT, BP6, Fontenay-aux-Roses Cedex F-92265 (France); Sarrazy, Fanny [Laboratoire Lesions des Acides Nucleiques, CEA, DSM, INAC, SCIB, UMR-E CEA/UJF-Grenoble 1, 17 Rue des Martyrs, F-38054 Grenoble Cedex 9 (France); Sauvaigo, Sylvie, E-mail: sylvie.sauvaigo@cea.fr [Laboratoire Lesions des Acides Nucleiques, CEA, DSM, INAC, SCIB, UMR-E CEA/UJF-Grenoble 1, 17 Rue des Martyrs, F-38054 Grenoble Cedex 9 (France)

    2012-08-01

    Several studies have demonstrated the deleterious effect of aging on the capacity of cells to repair their DNA. However, current existing assays aimed at measuring DNA repair address only a specific repair step dedicated to the correction of a specific DNA lesion type. Consequently they provide no information regarding the repair pathways that handle other types of lesions. In addition to aging, consequences of photo-exposure on these repair processes remain elusive. In this study we evaluated the consequence of aging and of chronic and/or acute photo-exposure on DNA repair in human skin fibroblasts using a multiplexed approach, which provided detailed information on several repair pathways at the same time. The resulting data were analyzed with adapted statistics/bioinformatics tools. We showed that, irrespective of the repair pathway considered, excision/synthesis was less efficient in non-exposed cells from elderly compared to cells from young adults and that photo-exposure disrupted this very clear pattern. Moreover, it was evidenced that chronic sun-exposure induced changes in DNA repair properties. Finally, the identification of a specific signature at the level of the NER pathway in cells repeatedly exposed to sun revealed a cumulative effect of UVB exposure and chronic sun irradiation. The uses of bioinformatics tools in this study was essential to fully take advantage of the large sum of data obtained with our multiplexed DNA repair assay and unravel the effects of environmental exposure on DNA repair pathways.

  7. Differential radiosensitivity phenotypes of DNA-PKcs mutations affecting NHEJ and HRR systems following irradiation with gamma-rays or very low fluences of alpha particles.

    Science.gov (United States)

    Lin, Yu-Fen; Nagasawa, Hatsumi; Little, John B; Kato, Takamitsu A; Shih, Hung-Ying; Xie, Xian-Jin; Wilson, Paul F; Brogan, John R; Kurimasa, Akihiro; Chen, David J; Bedford, Joel S; Chen, Benjamin P C

    2014-01-01

    We have examined cell-cycle dependence of chromosomal aberration induction and cell killing after high or low dose-rate γ irradiation in cells bearing DNA-PKcs mutations in the S2056 cluster, the T2609 cluster, or the kinase domain. We also compared sister chromatid exchanges (SCE) production by very low fluences of α-particles in DNA-PKcs mutant cells, and in homologous recombination repair (HRR) mutant cells including Rad51C, Rad51D, and Fancg/xrcc9. Generally, chromosomal aberrations and cell killing by γ-rays were similarly affected by mutations in DNA-PKcs, and these mutant cells were more sensitive in G1 than in S/G2 phase. In G1-irradiated DNA-PKcs mutant cells, both chromosome- and chromatid-type breaks and exchanges were in excess than wild-type cells. For cells irradiated in late S/G2 phase, mutant cells showed very high yields of chromatid breaks compared to wild-type cells. Few exchanges were seen in DNA-PKcs-null, Ku80-null, or DNA-PKcs kinase dead mutants, but exchanges in excess were detected in the S2506 or T2609 cluster mutants. SCE induction by very low doses of α-particles is resulted from bystander effects in cells not traversed by α-particles. SCE seen in wild-type cells was completely abolished in Rad51C- or Rad51D-deficient cells, but near normal in Fancg/xrcc9 cells. In marked contrast, very high levels of SCEs were observed in DNA-PKcs-null, DNA-PKcs kinase-dead and Ku80-null mutants. SCE induction was also abolished in T2609 cluster mutant cells, but was only slightly reduced in the S2056 cluster mutant cells. Since both non-homologous end-joining (NHEJ) and HRR systems utilize initial DNA lesions as a substrate, these results suggest the possibility of a competitive interference phenomenon operating between NHEJ and at least the Rad51C/D components of HRR; the level of interaction between damaged DNA and a particular DNA-PK component may determine the level of interaction of such DNA with a relevant HRR component.

  8. Whole Genome DNA Methylation Analysis of Obstructive Sleep Apnea: IL1R2, NPR2, AR, SP140 Methylation and Clinical Phenotype.

    Science.gov (United States)

    Chen, Yung-Che; Chen, Ting-Wen; Su, Mao-Chang; Chen, Chung-Jen; Chen, Kuang-Den; Liou, Chia-Wei; Tang, Petrus; Wang, Ting-Ya; Chang, Jen-Chieh; Wang, Chin-Chou; Lin, Hsin-Ching; Chin, Chien-Hung; Huang, Kuo-Tung; Lin, Meng-Chih; Hsiao, Chang-Chun

    2016-04-01

    We hypothesized that DNA methylation patterns may contribute to disease severity or the development of hypertension and excessive daytime sleepiness (EDS) in patients with obstructive sleep apnea (OSA). Illumina's (San Diego, CA, USA) DNA methylation 27-K assay was used to identify differentially methylated loci (DML). DNA methylation levels were validated by pyrosequencing. A discovery cohort of 15 patients with OSA and 6 healthy subjects, and a validation cohort of 72 patients with sleep disordered breathing (SDB). Microarray analysis identified 636 DMLs in patients with OSA versus healthy subjects, and 327 DMLs in patients with OSA and hypertension versus those without hypertension. In the validation cohort, no significant difference in DNA methylation levels of six selected genes was found between the primary snoring subjects and OSA patients (primary outcome). However, a secondary outcome analysis showed that interleukin-1 receptor 2 (IL1R2) promoter methylation (-114 cytosine followed by guanine dinucleotide sequence [CpG] site) was decreased and IL1R2 protein levels were increased in the patients with SDB with an oxygen desaturation index > 30. Androgen receptor (AR) promoter methylation (-531 CpG site) and AR protein levels were both increased in the patients with SDB with an oxygen desaturation index > 30. Natriuretic peptide receptor 2 (NPR2) promoter methylation (-608/-618 CpG sites) were decreased, whereas levels of both NPR2 and serum C type natriuretic peptide protein were increased in the SDB patients with EDS. Speckled protein 140 (SP140) promoter methylation (-194 CpG site) was increased, and SP140 protein levels were decreased in the patients with SDB and EDS. IL1R2 hypomethylation and AR hypermethylation may constitute an important determinant of disease severity, whereas NPR2 hypomethylation and SP140 hypermethylation may provide a biomarker for vulnerability to EDS in OSA. A commentary on this article appears in this issue on page 723. © 2016

  9. Dried Blood Spots, an Affordable Tool to Collect, Ship, and Sequence gDNA from Patients with an X-Linked Agammaglobulinemia Phenotype Residing in a Developing Country

    Directory of Open Access Journals (Sweden)

    Gesmar R. S. Segundo

    2018-02-01

    Full Text Available BackgroundNew sequencing techniques have revolutionized the identification of the molecular basis of primary immunodeficiency disorders (PID not only by establishing a gene-based diagnosis but also by facilitating defect-specific treatment strategies, improving quality of life and survival, and allowing factual genetic counseling. Because these techniques are generally not available for physicians and their patients residing in developing countries, collaboration with overseas laboratories has been explored as a possible, albeit cumbersome, strategy. To reduce the cost of time and temperature-sensitive shipping, we selected Guthrie cards, developed for newborn screening, to collect dried blood spots (DBS, as a source of DNA that can be shipped by regular mail at minimal cost.MethodBlood was collected and blotted onto the filter paper of Guthrie cards by completely filling three circles. We enrolled 20 male patients with presumptive X-linked agammaglobulinemia (XLA cared for at the Vietnam National Children’s Hospital, their mothers, and several sisters for carrier analysis. DBS were stored at room temperature until ready to be shipped together, using an appropriately sized envelope, to a CLIA-certified laboratory in the US for sequencing. The protocol for Sanger sequencing was modified to account for the reduced quantity of gDNA extracted from DBS.ResultHigh-quality gDNA could be extracted from every specimen. Bruton tyrosine kinase (BTK mutations were identified in 17 of 20 patients studied, confirming the diagnosis of XLA in 85% of the study cohort. Type and location of the mutations were similar to those reported in previous reviews. The mean age when XLA was suspected clinically was 4.6 years, similar to that reported by Western countries. Two of 15 mothers, each with an affected boy, had a normal BTK sequence, suggesting gonadal mosaicism.ConclusionDBS collected on Guthrie cards can be shipped inexpensively by airmail across continents

  10. Phenotypic variability among strains of Pasteurella multocida ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-05-02

    May 2, 2008 ... Available online at http://www.academicjournals.org/AJB. ISSN 1684–5315 ... extended phenotypic characterization methods supported by DNA ... septicaemia African (Obudu) strain (E:2) which are currently employed as ...

  11. Meta-analysis of archived DNA microarrays identifies genes regulated by hypoxia and involved in a metastatic phenotype in cancer cells

    International Nuclear Information System (INIS)

    Pierre, Michael; DeHertogh, Benoît; Gaigneaux, Anthoula; DeMeulder, Bertrand; Berger, Fabrice; Bareke, Eric; Michiels, Carine; Depiereux, Eric

    2010-01-01

    Metastasis is a major cancer-related cause of death. Recent studies have described metastasis pathways. However, the exact contribution of each pathway remains unclear. Another key feature of a tumor is the presence of hypoxic areas caused by a lack of oxygen at the center of the tumor. Hypoxia leads to the expression of pro-metastatic genes as well as the repression of anti-metastatic genes. As many Affymetrix datasets about metastasis and hypoxia are publicly available and not fully exploited, this study proposes to re-analyze these datasets to extract new information about the metastatic phenotype induced by hypoxia in different cancer cell lines. Affymetrix datasets about metastasis and/or hypoxia were downloaded from GEO and ArrayExpress. AffyProbeMiner and GCRMA packages were used for pre-processing and the Window Welch t test was used for processing. Three approaches of meta-analysis were eventually used for the selection of genes of interest. Three complementary approaches were used, that eventually selected 183 genes of interest. Out of these 183 genes, 99, among which the well known JUNB, FOS and TP63, have already been described in the literature to be involved in cancer. Moreover, 39 genes of those, such as SERPINE1 and MMP7, are known to regulate metastasis. Twenty-one genes including VEGFA and ID2 have also been described to be involved in the response to hypoxia. Lastly, DAVID classified those 183 genes in 24 different pathways, among which 8 are directly related to cancer while 5 others are related to proliferation and cell motility. A negative control composed of 183 random genes failed to provide such results. Interestingly, 6 pathways retrieved by DAVID with the 183 genes of interest concern pathogen recognition and phagocytosis. The proposed methodology was able to find genes actually known to be involved in cancer, metastasis and hypoxia and, thus, we propose that the other genes selected based on the same methodology are of prime interest in

  12. A structured simple form for ordering genetic tests is needed to ensure coupling of clinical detail (phenotype) with DNA variants (genotype) to ensure utility in publication and databases.

    Science.gov (United States)

    Cotton, R G H; Auerbach, A D; Brown, A F; Carrera, P; Christodoulou, J; Claustres, M; Compton, J; Cox, D W; De Baere, E; den Dunnen, J T; Greenblatt, M; Fujiwara, M; Hilbert, P; Jani, A; Lehvaslaiho, H; Nebert, D W; Verma, I; Vihinen, M

    2007-10-01

    Researchers and clinicians ideally need instant access to all the variation in their gene/locus of interest to efficiently conduct their research and genetic healthcare to the highest standards. Currently much key data resides in the laboratory books or patient records around the world, as there are many impediments to submitting this data. It would be ideal therefore if a semiautomated pathway was available, with a minimum of effort, to make the deidentified data publicly available for others to use. The Human Variome Project (HVP) meeting listed 96 recommendations to work toward this situation. This article is planned to initiate a strategy to enhance the collection of phenotype and genotype data from the clinician/diagnostic laboratory nexus. Thus, the aim is to develop universally applicable forms that people can use when investigating patients for each inherited disease, to assist in satisfying many of the recommendations of the HVP Meeting [Cotton et al., 2007]. We call for comment and collaboration in this article. Copyright 2007 Wiley-Liss, Inc.

  13. The Rhizobium etli rpoN locus: DNA sequence analysis and phenotypical characterization of rpoN, ptsN, and ptsA mutants.

    Science.gov (United States)

    Michiels, J; Van Soom, T; D'hooghe, I; Dombrecht, B; Benhassine, T; de Wilde, P; Vanderleyden, J

    1998-04-01

    The rpoN region of Rhizobium etli was isolated by using the Bradyrhizobium japonicum rpoN1 gene as a probe. Nucleotide sequence analysis of a 5,600-bp DNA fragment of this region revealed the presence of four complete open reading frames (ORFs), ORF258, rpoN, ORF191, and ptsN, coding for proteins of 258, 520, 191, and 154 amino acids, respectively. The gene product of ORF258 is homologous to members of the ATP-binding cassette-type permeases. ORF191 and ptsN are homologous to conserved ORFs found downstream from rpoN genes in other bacterial species. Unlike in most other microorganisms, rpoN and ORF191 are separated by approximately 1.6 kb. The R. etli rpoN gene was shown to control in free-living conditions the production of melanin, the activation of nifH, and the metabolism of C4-dicarboxylic acids and several nitrogen sources (ammonium, nitrate, alanine, and serine). Expression of the rpoN gene was negatively autoregulated and occurred independently of the nitrogen source. Inactivation of the ptsN gene resulted in a decrease of melanin synthesis and nifH expression. In a search for additional genes controlling the synthesis of melanin, an R. etli mutant carrying a Tn5 insertion in ptsA, a gene homologous to the Escherichia coli gene coding for enzyme I of the phosphoenolpyruvate:sugar phosphotransferase system, was obtained. The R. etli ptsA mutant also displayed reduced expression of nifH. The ptsN and ptsA mutants also displayed increased sensitivity to the toxic effects of malate and succinate. Growth of both mutants was inhibited by these C4-dicarboxylates at 20 mM at pH 7.0, while wild-type cells grow normally under these conditions. The effect of malate occurred independently of the nitrogen source used. Growth inhibition was decreased by lowering the pH of the growth medium. These results suggest that ptsN and ptsA are part of the same regulatory cascade, the inactivation of which renders the cells sensitive to toxic effects of elevated concentrations of

  14. Global organization of a positive-strand RNA virus genome.

    Directory of Open Access Journals (Sweden)

    Baodong Wu

    Full Text Available The genomes of plus-strand RNA viruses contain many regulatory sequences and structures that direct different viral processes. The traditional view of these RNA elements are as local structures present in non-coding regions. However, this view is changing due to the discovery of regulatory elements in coding regions and functional long-range intra-genomic base pairing interactions. The ∼4.8 kb long RNA genome of the tombusvirus tomato bushy stunt virus (TBSV contains these types of structural features, including six different functional long-distance interactions. We hypothesized that to achieve these multiple interactions this viral genome must utilize a large-scale organizational strategy and, accordingly, we sought to assess the global conformation of the entire TBSV genome. Atomic force micrographs of the genome indicated a mostly condensed structure composed of interconnected protrusions extending from a central hub. This configuration was consistent with the genomic secondary structure model generated using high-throughput selective 2'-hydroxyl acylation analysed by primer extension (i.e. SHAPE, which predicted different sized RNA domains originating from a central region. Known RNA elements were identified in both domain and inter-domain regions, and novel structural features were predicted and functionally confirmed. Interestingly, only two of the six long-range interactions known to form were present in the structural model. However, for those interactions that did not form, complementary partner sequences were positioned relatively close to each other in the structure, suggesting that the secondary structure level of viral genome structure could provide a basic scaffold for the formation of different long-range interactions. The higher-order structural model for the TBSV RNA genome provides a snapshot of the complex framework that allows multiple functional components to operate in concert within a confined context.

  15. Phenotypic characterization and 16S rDNA identification of culturable non-obligate halophilic bacterial communities from a hypersaline lake, La Sal del Rey, in extreme South Texas (USA).

    Science.gov (United States)

    Phillips, Kristen; Zaidan, Frederic; Elizondo, Omar R; Lowe, Kristine L

    2012-02-02

    La Sal del Rey ("the King's Salt") is one of several naturally-occurring salt lakes in Hidalgo County, Texas and is part of the Lower Rio Grande Valley National Wildlife Refuge. The research objective was to isolate and characterize halophilic microorganisms from La Sal del Rey. Water samples were collected from the lake and a small creek that feeds into the lake. Soil samples were collected from land adjacent to the water sample locations. Sample salinity was determined using a refractometer. Samples were diluted and cultured on a synthetic saline medium to grow halophilic bacteria. The density of halophiles was estimated by viable plate counts. A collection of isolates was selected, gram-stained, tested for catalase, and characterized using API 20E® test strips. Isolates were putatively identified by sequencing the 16S rDNA. Carbon source utilization by the microbial community from each sample site was examined using EcoPlate™ assays and the carbon utilization total activity of the community was determined. Results showed that salinity ranged from 4 parts per thousand (ppt) at the lake water source to 420 ppt in water samples taken just along the lake shore. The density of halophilic bacteria in water samples ranged from 1.2 × 102 - 5.2 × 103 colony forming units per ml (cfu ml-1) whereas the density in soil samples ranged from 4.0 × 105 - 2.5 × 106 colony forming units per gram (cfu g-1). In general, as salinity increased the density of the bacterial community decreased. Microbial communities from water and soil samples were able to utilize 12 - 31 carbon substrates. The greatest number of substrates utilized was by water-borne communities compared to soil-based communities, especially at lower salinities. The majority of bacteria isolated were gram-negative, catalase-positive, rods. Biochemical profiles constructed from API 20E® test strips showed that bacterial isolates from low-salinity water samples (4 ppt) showed the greatest phenotypic diversity

  16. Quality Control Test for Sequence-Phenotype Assignments

    Science.gov (United States)

    Ortiz, Maria Teresa Lara; Rosario, Pablo Benjamín Leon; Luna-Nevarez, Pablo; Gamez, Alba Savin; Martínez-del Campo, Ana; Del Rio, Gabriel

    2015-01-01

    Relating a gene mutation to a phenotype is a common task in different disciplines such as protein biochemistry. In this endeavour, it is common to find false relationships arising from mutations introduced by cells that may be depurated using a phenotypic assay; yet, such phenotypic assays may introduce additional false relationships arising from experimental errors. Here we introduce the use of high-throughput DNA sequencers and statistical analysis aimed to identify incorrect DNA sequence-phenotype assignments and observed that 10–20% of these false assignments are expected in large screenings aimed to identify critical residues for protein function. We further show that this level of incorrect DNA sequence-phenotype assignments may significantly alter our understanding about the structure-function relationship of proteins. We have made available an implementation of our method at http://bis.ifc.unam.mx/en/software/chispas. PMID:25700273

  17. Ancient DNA

    DEFF Research Database (Denmark)

    Willerslev, Eske; Cooper, Alan

    2004-01-01

    ancient DNA, palaeontology, palaeoecology, archaeology, population genetics, DNA damage and repair......ancient DNA, palaeontology, palaeoecology, archaeology, population genetics, DNA damage and repair...

  18. Genetic variants influencing phenotypic variance heterogeneity.

    Science.gov (United States)

    Ek, Weronica E; Rask-Andersen, Mathias; Karlsson, Torgny; Enroth, Stefan; Gyllensten, Ulf; Johansson, Åsa

    2018-03-01

    Most genetic studies identify genetic variants associated with disease risk or with the mean value of a quantitative trait. More rarely, genetic variants associated with variance heterogeneity are considered. In this study, we have identified such variance single-nucleotide polymorphisms (vSNPs) and examined if these represent biological gene × gene or gene × environment interactions or statistical artifacts caused by multiple linked genetic variants influencing the same phenotype. We have performed a genome-wide study, to identify vSNPs associated with variance heterogeneity in DNA methylation levels. Genotype data from over 10 million single-nucleotide polymorphisms (SNPs), and DNA methylation levels at over 430 000 CpG sites, were analyzed in 729 individuals. We identified vSNPs for 7195 CpG sites (P mean DNA methylation levels. We further showed that variance heterogeneity between genotypes mainly represents additional, often rare, SNPs in linkage disequilibrium (LD) with the respective vSNP and for some vSNPs, multiple low frequency variants co-segregating with one of the vSNP alleles. Therefore, our results suggest that variance heterogeneity of DNA methylation mainly represents phenotypic effects by multiple SNPs, rather than biological interactions. Such effects may also be important for interpreting variance heterogeneity of more complex clinical phenotypes.

  19. Asthma phenotypes in childhood.

    Science.gov (United States)

    Reddy, Monica B; Covar, Ronina A

    2016-04-01

    This review describes the literature over the past 18 months that evaluated childhood asthma phenotypes, highlighting the key aspects of these studies, and comparing these studies to previous ones in this area. Recent studies on asthma phenotypes have identified new phenotypes on the basis of statistical analyses (using cluster analysis and latent class analysis methodology) and have evaluated the outcomes and associated risk factors of previously established early childhood asthma phenotypes that are based on asthma onset and patterns of wheezing illness. There have also been investigations focusing on immunologic, physiologic, and genetic correlates of various phenotypes, as well as identification of subphenotypes of severe childhood asthma. Childhood asthma remains a heterogeneous condition, and investigations into these various presentations, risk factors, and outcomes are important since they can offer therapeutic and prognostic relevance. Further investigation into the immunopathology and genetic basis underlying childhood phenotypes is important so therapy can be tailored accordingly.

  20. Clinical phenotypes of asthma

    NARCIS (Netherlands)

    Bel, Elisabeth H.

    2004-01-01

    PURPOSE OF REVIEW: Asthma is a phenotypically heterogeneous disorder and, over the years, many different clinical subtypes of asthma have been described. A precise definition of asthma phenotypes is now becoming more and more important, not only for a better understanding of pathophysiologic

  1. Molecular and phenotypic characterization of anamorphic fungi

    OpenAIRE

    Madrid Lorca, Hugo

    2011-01-01

    Anamorphic fungi (those reproducing asexually) are a big part of kingdom Fungi. Most of them occur as saprobes in nature, but numerous species are pathogenic to plants and animals including man. With the aim of contributing to the knowledge of the diversity and distribution of anamorphic fungi, we performed a phenotypic and molecular characterization of environmental and clinical isolates of these fungi. Based on a polyphasic taxonomy approach which included morphology, physiology and DNA seq...

  2. Sequence analysis and transcript identification within 1.5 MB of DNA deleted together with the NDP and MAO genes in atypical Norrie disease patients presenting with a profound phenotype.

    Science.gov (United States)

    Suárez-Merino, B; Bye, J; McDowall, J; Ross, M; Craig, I W

    2001-06-01

    Mutations at the Norrie disease gene locus, NDP, manifest in a broad range of defects. These range from a relatively mild, late-onset, exudative vitreoretinopathy to congenital blindness and sensorineural deafness combined, in some cases, with mental retardation. In addition, extensive deletions involving the NDP locus, located at Xp11.3, the adjacent monoamine oxidadase genes MAOA and MAOB, and additional material, result in a more severe pattern of symptoms. The phenotypes include all or some of the following; mental retardation, involuntary movements, hypertensive crises and hypogonadism. We extended an existing YAC contig to embrace the boundaries of three of the largest deletions and converted this into four PAC contigs. Computer analysis and experimental data have resulted in the identification of several putative loci, including a phosphatase inhibitor 2-like gene (dJ154.1) and a 250-bp sequence which resembles a homeobox domain (dA113.3), 1.2 Mb and 400 kb respectively from the MAO/NDP cluster. The pattern of expression of dJ154.1 suggests that it may represent an important factor contributing to the complex phenotypes of these deletion patients. Hum Mutat 17:523, 2001. Copyright 2001 Wiley-Liss, Inc.

  3. HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways

    Science.gov (United States)

    Guo, Fang; Zhao, Qiong; Cheng, Junjun; Qi, Yonghe; Su, Qing; Wei, Lai; Li, Wenhui; Chang, Jinhong

    2017-01-01

    Hepatitis B virus (HBV) core protein assembles viral pre-genomic (pg) RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs) and sulfamoylbenzamides (SBAs), have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or “empty” capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc) DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B. PMID:28945802

  4. HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways.

    Science.gov (United States)

    Guo, Fang; Zhao, Qiong; Sheraz, Muhammad; Cheng, Junjun; Qi, Yonghe; Su, Qing; Cuconati, Andrea; Wei, Lai; Du, Yanming; Li, Wenhui; Chang, Jinhong; Guo, Ju-Tao

    2017-09-01

    Hepatitis B virus (HBV) core protein assembles viral pre-genomic (pg) RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs) and sulfamoylbenzamides (SBAs), have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or "empty" capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc) DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B.

  5. HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways.

    Directory of Open Access Journals (Sweden)

    Fang Guo

    2017-09-01

    Full Text Available Hepatitis B virus (HBV core protein assembles viral pre-genomic (pg RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs and sulfamoylbenzamides (SBAs, have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or "empty" capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B.

  6. COPD: Definition and Phenotypes

    DEFF Research Database (Denmark)

    Vestbo, J.

    2014-01-01

    particles or gases. Exacerbations and comorbidities contribute to the overall severity in individual patients. The evolution of this definition and the diagnostic criteria currently in use are discussed. COPD is increasingly divided in subgroups or phenotypes based on specific features and association...

  7. Phenotypic Resistance to Antibiotics

    Directory of Open Access Journals (Sweden)

    Jose L. Martinez

    2013-04-01

    Full Text Available The development of antibiotic resistance is usually associated with genetic changes, either to the acquisition of resistance genes, or to mutations in elements relevant for the activity of the antibiotic. However, in some situations resistance can be achieved without any genetic alteration; this is called phenotypic resistance. Non-inherited resistance is associated to specific processes such as growth in biofilms, a stationary growth phase or persistence. These situations might occur during infection but they are not usually considered in classical susceptibility tests at the clinical microbiology laboratories. Recent work has also shown that the susceptibility to antibiotics is highly dependent on the bacterial metabolism and that global metabolic regulators can modulate this phenotype. This modulation includes situations in which bacteria can be more resistant or more susceptible to antibiotics. Understanding these processes will thus help in establishing novel therapeutic approaches based on the actual susceptibility shown by bacteria during infection, which might differ from that determined in the laboratory. In this review, we discuss different examples of phenotypic resistance and the mechanisms that regulate the crosstalk between bacterial metabolism and the susceptibility to antibiotics. Finally, information on strategies currently under development for diminishing the phenotypic resistance to antibiotics of bacterial pathogens is presented.

  8. Adjusting phenotypes by noise control.

    Directory of Open Access Journals (Sweden)

    Kyung H Kim

    2012-01-01

    Full Text Available Genetically identical cells can show phenotypic variability. This is often caused by stochastic events that originate from randomness in biochemical processes involving in gene expression and other extrinsic cellular processes. From an engineering perspective, there have been efforts focused on theory and experiments to control noise levels by perturbing and replacing gene network components. However, systematic methods for noise control are lacking mainly due to the intractable mathematical structure of noise propagation through reaction networks. Here, we provide a numerical analysis method by quantifying the parametric sensitivity of noise characteristics at the level of the linear noise approximation. Our analysis is readily applicable to various types of noise control and to different types of system; for example, we can orthogonally control the mean and noise levels and can control system dynamics such as noisy oscillations. As an illustration we applied our method to HIV and yeast gene expression systems and metabolic networks. The oscillatory signal control was applied to p53 oscillations from DNA damage. Furthermore, we showed that the efficiency of orthogonal control can be enhanced by applying extrinsic noise and feedback. Our noise control analysis can be applied to any stochastic model belonging to continuous time Markovian systems such as biological and chemical reaction systems, and even computer and social networks. We anticipate the proposed analysis to be a useful tool for designing and controlling synthetic gene networks.

  9. Phenotype ontologies and cross-species analysis for translational research.

    Directory of Open Access Journals (Sweden)

    Peter N Robinson

    2014-04-01

    Full Text Available The use of model organisms as tools for the investigation of human genetic variation has significantly and rapidly advanced our understanding of the aetiologies underlying hereditary traits. However, while equivalences in the DNA sequence of two species may be readily inferred through evolutionary models, the identification of equivalence in the phenotypic consequences resulting from comparable genetic variation is far from straightforward, limiting the value of the modelling paradigm. In this review, we provide an overview of the emerging statistical and computational approaches to objectively identify phenotypic equivalence between human and model organisms with examples from the vertebrate models, mouse and zebrafish. Firstly, we discuss enrichment approaches, which deem the most frequent phenotype among the orthologues of a set of genes associated with a common human phenotype as the orthologous phenotype, or phenolog, in the model species. Secondly, we introduce and discuss computational reasoning approaches to identify phenotypic equivalences made possible through the development of intra- and interspecies ontologies. Finally, we consider the particular challenges involved in modelling neuropsychiatric disorders, which illustrate many of the remaining difficulties in developing comprehensive and unequivocal interspecies phenotype mappings.

  10. Fanconi anemia and DNA repair.

    Science.gov (United States)

    Grompe, M; D'Andrea, A

    2001-10-01

    Fanconi anemia (FA) is an autosomal recessive disorder caused by defects in at least eight distinct genes FANCA, B, C, D1, D2, E, F and G. The clinical phenotype of all FA complementation groups is similar and is characterized by progressive bone marrow failure, cancer proneness and typical birth defects. The principal cellular phenotype is hypersensitivity to DNA damage, particularly interstrand DNA crosslinks. The FA proteins constitute a multiprotein pathway whose precise biochemical function(s) remain unknown. Five of the FA proteins (FANCA, C, E, F and G) interact in a nuclear complex upstream of FANCD2. FANCB and FANCD1 have not yet been cloned, but it is likely that FANCB is part of the nuclear complex and that FANCD1 acts downstream of FANCD2. The FA nuclear complex regulates the mono-ubiquitination of FANCD2 in response to DNA damage, resulting in targeting of this protein into nuclear foci. These foci also contain BRCA1 and other DNA damage response proteins. In male meiosis, FANCD2 also co-localizes with BRCA1 at synaptonemal complexes. Together, these data suggest that the FA pathway functions primarily as a DNA damage response system, although its exact role (direct involvement in DNA repair versus indirect, facilitating role) has not yet been defined.

  11. Modeling DNA

    Science.gov (United States)

    Robertson, Carol

    2016-01-01

    Deoxyribonucleic acid (DNA) is life's most amazing molecule. It carries the genetic instructions that almost every organism needs to develop and reproduce. In the human genome alone, there are some three billion DNA base pairs. The most difficult part of teaching DNA structure, however, may be getting students to visualize something as small as a…

  12. Clinical strains of acinetobacter classified by DNA-DNA hybridization

    International Nuclear Information System (INIS)

    Tjernberg, I.; Ursing, J.

    1989-01-01

    A collection of Acinetobacter strains consisting of 168 consecutive clinical strains and 30 type and reference strains was studied by DNA-DNA hybridization and a few phenotypic tests. The field strains could be allotted to 13 DNA groups. By means of reference strains ten of these could be identified with groups described by Bouvet and Grimont (1986), while three groups were new; they were given the numbers 13-15. The type strain of A. radioresistens- recently described by Nishimura et al. (1988) - was shown to be a member of DNA group 12, which comprised 31 clinical isolates. Of the 19 strains of A. junii, eight showed hemolytic acitivity on sheep and human blood agar and an additional four strains on human blood agar only. Strains of this species have previously been regarded as non-hemolytic. Reciprocal DNA pairing data for the reference strains of the DNA gropus were treated by UPGMA clustering. The reference strains for A. calcoaceticus, A. baumannii and DNA groups 3 and 13 formed a cluster with about 70% relatedness within the cluster. Other DNA groups joined at levels below 60%. (author)

  13. Clinical strains of acinetobacter classified by DNA-DNA hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Tjernberg, I; Ursing, J [Department of Medical Microbiology, University of Lund, Malmoe General Hospital, Malmoe (Sweden)

    1989-01-01

    A collection of Acinetobacter strains consisting of 168 consecutive clinical strains and 30 type and reference strains was studied by DNA-DNA hybridization and a few phenotypic tests. The field strains could be allotted to 13 DNA groups. By means of reference strains ten of these could be identified with groups described by Bouvet and Grimont (1986), while three groups were new; they were given the numbers 13-15. The type strain of A. radioresistens- recently described by Nishimura et al. (1988) - was shown to be a member of DNA group 12, which comprised 31 clinical isolates. Of the 19 strains of A. junii, eight showed hemolytic acitivity on sheep and human blood agar and an additional four strains on human blood agar only. Strains of this species have previously been regarded as non-hemolytic. Reciprocal DNA pairing data for the reference strains of the DNA gropus were treated by UPGMA clustering. The reference strains for A. calcoaceticus, A. baumannii and DNA groups 3 and 13 formed a cluster with about 70% relatedness within the cluster. Other DNA groups joined at levels below 60%. (author).

  14. Targeting phenotypically tolerant Mycobacterium tuberculosis

    Science.gov (United States)

    Gold, Ben; Nathan, Carl

    2016-01-01

    While the immune system is credited with averting tuberculosis in billions of individuals exposed to Mycobacterium tuberculosis, the immune system is also culpable for tempering the ability of antibiotics to deliver swift and durable cure of disease. In individuals afflicted with tuberculosis, host immunity produces diverse microenvironmental niches that support suboptimal growth, or complete growth arrest, of M. tuberculosis. The physiological state of nonreplication in bacteria is associated with phenotypic drug tolerance. Many of these host microenvironments, when modeled in vitro by carbon starvation, complete nutrient starvation, stationary phase, acidic pH, reactive nitrogen intermediates, hypoxia, biofilms, and withholding streptomycin from the streptomycin-addicted strain SS18b, render M. tuberculosis profoundly tolerant to many of the antibiotics that are given to tuberculosis patients in a clinical setting. Targeting nonreplicating persisters is anticipated to reduce the duration of antibiotic treatment and rate of post-treatment relapse. Some promising drugs to treat tuberculosis, such as rifampicin and bedaquiline, only kill nonreplicating M. tuberculosis in vitro at concentrations far greater than their minimal inhibitory concentrations against replicating bacilli. There is an urgent demand to identify which of the currently used antibiotics, and which of the molecules in academic and corporate screening collections, have potent bactericidal action on nonreplicating M. tuberculosis. With this goal, we review methods of high throughput screening to target nonreplicating M. tuberculosis and methods to progress candidate molecules. A classification based on structures and putative targets of molecules that have been reported to kill nonreplicating M. tuberculosis revealed a rich diversity in pharmacophores. However, few of these compounds were tested under conditions that would exclude the impact of adsorbed compound acting during the recovery phase of

  15. DNA repair deficiency in neurodegeneration

    DEFF Research Database (Denmark)

    Jeppesen, Dennis Kjølhede; Bohr, Vilhelm A; Stevnsner, Tinna V.

    2011-01-01

    Deficiency in repair of nuclear and mitochondrial DNA damage has been linked to several neurodegenerative disorders. Many recent experimental results indicate that the post-mitotic neurons are particularly prone to accumulation of unrepaired DNA lesions potentially leading to progressive...... neurodegeneration. Nucleotide excision repair is the cellular pathway responsible for removing helix-distorting DNA damage and deficiency in such repair is found in a number of diseases with neurodegenerative phenotypes, including Xeroderma Pigmentosum and Cockayne syndrome. The main pathway for repairing oxidative...... base lesions is base excision repair, and such repair is crucial for neurons given their high rates of oxygen metabolism. Mismatch repair corrects base mispairs generated during replication and evidence indicates that oxidative DNA damage can cause this pathway to expand trinucleotide repeats, thereby...

  16. From metabolome to phenotype

    DEFF Research Database (Denmark)

    Khakimov, Bekzod; Rasmussen, Morten Arendt; Kannangara, Rubini Maya

    2017-01-01

    for ideal vegetable protein production and for augmented β-glucan production. Seeds from three barley lines (Bomi, lys3.a and lys5.f) were sampled eight times during grain filling and analysed for metabolites using gas chromatography-mass spectrometry (GC-MS). The lys3.a mutation disrupts a regulator gene...... their successful application to link genetic and environmental factors with the seed phenotype of unique and agro-economically important barley models for optimal vegetable protein and dietary fibre production......., causing an increase in proteins rich in the essential amino acid lysine, while lys5.f carries a mutation in an ADP-glucose transporter gene leading to a significant increase in production of mixed-linkage β-glucan at the expense of α-glucan. Unique metabolic patterns associated with the tricarboxylic acid...

  17. Mechanistic phenotypes: an aggregative phenotyping strategy to identify disease mechanisms using GWAS data.

    Directory of Open Access Journals (Sweden)

    Jonathan D Mosley

    Full Text Available A single mutation can alter cellular and global homeostatic mechanisms and give rise to multiple clinical diseases. We hypothesized that these disease mechanisms could be identified using low minor allele frequency (MAF<0.1 non-synonymous SNPs (nsSNPs associated with "mechanistic phenotypes", comprised of collections of related diagnoses. We studied two mechanistic phenotypes: (1 thrombosis, evaluated in a population of 1,655 African Americans; and (2 four groupings of cancer diagnoses, evaluated in 3,009 white European Americans. We tested associations between nsSNPs represented on GWAS platforms and mechanistic phenotypes ascertained from electronic medical records (EMRs, and sought enrichment in functional ontologies across the top-ranked associations. We used a two-step analytic approach whereby nsSNPs were first sorted by the strength of their association with a phenotype. We tested associations using two reverse genetic models and standard additive and recessive models. In the second step, we employed a hypothesis-free ontological enrichment analysis using the sorted nsSNPs to identify functional mechanisms underlying the diagnoses comprising the mechanistic phenotypes. The thrombosis phenotype was solely associated with ontologies related to blood coagulation (Fisher's p = 0.0001, FDR p = 0.03, driven by the F5, P2RY12 and F2RL2 genes. For the cancer phenotypes, the reverse genetics models were enriched in DNA repair functions (p = 2×10-5, FDR p = 0.03 (POLG/FANCI, SLX4/FANCP, XRCC1, BRCA1, FANCA, CHD1L while the additive model showed enrichment related to chromatid segregation (p = 4×10-6, FDR p = 0.005 (KIF25, PINX1. We were able to replicate nsSNP associations for POLG/FANCI, BRCA1, FANCA and CHD1L in independent data sets. Mechanism-oriented phenotyping using collections of EMR-derived diagnoses can elucidate fundamental disease mechanisms.

  18. Deep Learning for Plant Phenotyping

    OpenAIRE

    Mori, Matteo

    2016-01-01

    Plant Phenotyping is an emerging science which provides us the knowledge to better understand plants. Indeed, the study of the link between genetic background and environment in which plants develop can help us to determine cures for plants’ sicknesses and new ways to improve yields using limited resources. In this regard, one of the main aspects of Plant Phenotyping that were studied in the past, was Root Phenotyping, which is based on the study of the root architectures. In particular, toda...

  19. Epidemiological markers in Neisseria meningitidis: an estimate of the performance of genotyping vs phenotyping

    DEFF Research Database (Denmark)

    Weis, N; Lind, I

    1998-01-01

    In order to estimate the performance of genotypic vs phenotypic characterization of Neisseria meningitidis, 2 methods, DNA fingerprinting and multilocus enzyme electrophoresis (MEE), were assessed as regards applicability, reproducibility and discriminating capacity. 50 serogroup B and 52 serogro......, and as applied in the study MEE was superior to DNA fingerprinting. Clusters of invasive strains were reliably identified by phenotyping alone, whereas determination of identity of carrier strains and an invasive strain required genotyping.......In order to estimate the performance of genotypic vs phenotypic characterization of Neisseria meningitidis, 2 methods, DNA fingerprinting and multilocus enzyme electrophoresis (MEE), were assessed as regards applicability, reproducibility and discriminating capacity. 50 serogroup B and 52 serogroup...... C Neisseria meningitidis strains from 96 patients with meningococcal disease and 22 serogroup C strains from healthy carriers were investigated. Both methods were 100% applicable to meningococcal strains and results of DNA fingerprinting as well as of MEE were reproducible. The number of types...

  20. DNA Camouflage

    Science.gov (United States)

    2016-01-08

    1 DNA Camouflage Supplementary Information Bijan Zakeri1,2*, Timothy K. Lu1,2*, Peter A. Carr2,3* 1Department of Electrical Engineering and...ll.mit.edu). Distribution A: Public Release   2 Supplementary Figure 1 DNA camouflage with the 2-state device. (a) In the presence of Cre, DSD-2[α...10 1 + Cre 1 500 1,000 length (bp) chromatogram alignment template − Cre   4 Supplementary Figure 3 DNA camouflage with a switchable

  1. Disentangling hexaploid genetics : towards DNA-informed breeding for postharvest performance in chrysanthemum

    NARCIS (Netherlands)

    Geest, van Geert

    2017-01-01

    DNA-informed selection can strongly improve the process of plant breeding. It requires the detection of DNA polymorphisms, calculation of genetic linkage, access to reliable phenotypes and methods to detect genetic loci associated with phenotypic traits of interest. Cultivated chrysanthemum is an

  2. Plant Phenotype Characterization System

    Energy Technology Data Exchange (ETDEWEB)

    Daniel W McDonald; Ronald B Michaels

    2005-09-09

    This report is the final scientific report for the DOE Inventions and Innovations Project: Plant Phenotype Characterization System, DE-FG36-04GO14334. The period of performance was September 30, 2004 through July 15, 2005. The project objective is to demonstrate the viability of a new scientific instrument concept for the study of plant root systems. The root systems of plants are thought to be important in plant yield and thus important to DOE goals in renewable energy sources. The scientific study and understanding of plant root systems is hampered by the difficulty in observing root activity and the inadequacy of existing root study instrumentation options. We have demonstrated a high throughput, non-invasive, high resolution technique for visualizing plant root systems in-situ. Our approach is based upon low-energy x-ray radiography and the use of containers and substrates (artificial soil) which are virtually transparent to x-rays. The system allows us to germinate and grow plant specimens in our containers and substrates and to generate x-ray images of the developing root system over time. The same plant can be imaged at different times in its development. The system can be used for root studies in plant physiology, plant morphology, plant breeding, plant functional genomics and plant genotype screening.

  3. Sex hormone binding globulin phenotypes

    DEFF Research Database (Denmark)

    Cornelisse, M M; Bennett, Patrick; Christiansen, M

    1994-01-01

    Human sex hormone binding globulin (SHBG) is encoded by a normal and a variant allele. The resulting SHBG phenotypes (the homozygous normal SHBG, the heterozygous SHBG and the homozygous variant SHBG phenotype) can be distinguished by their electrophoretic patterns. We developed a novel detection....... This method of detection was used to determine the distribution of SHBG phenotypes in healthy controls of both sexes and in five different pathological conditions characterized by changes in the SHBG level or endocrine disturbances (malignant and benign ovarian neoplasms, hirsutism, liver cirrhosis...... on the experimental values. Differences in SHBG phenotypes do not appear to have any clinical significance and no sex difference was found in the SHBG phenotype distribution....

  4. Phenotype determination guides swift genotyping of a DFNA2/KCNQ4 family with a hot spot mutation (W276S).

    NARCIS (Netherlands)

    Topsakal, V.; Pennings, R.J.E.; Brinke, H. te; Hamel, B.C.J.; Huygen, P.L.M.; Kremer, J.M.J.; Cremers, C.W.R.J.

    2005-01-01

    OBJECTIVE: Genotype a family trait with autosomal dominant nonsyndromic sensorineural hearing impairment guided only by the phenotype. STUDY DESIGN: Family study. SETTING: Tertiary referral center. PATIENTS: Fifteen family members. METHODS: In the first phase, sequence analysis was performed on DNA

  5. DNA glue

    DEFF Research Database (Denmark)

    Filichev, Vyacheslav V; Astakhova, Irina V.; Malakhov, Andrei D.

    2008-01-01

    Significant alterations in thermal stability of parallel DNA triplexes and antiparallel duplexes were observed upon changing the attachment of ethynylpyrenes from para to ortho in the structure of phenylmethylglycerol inserted as a bulge into DNA (TINA). Insertions of two ortho-TINAs as a pseudo...

  6. Hyperstretching DNA

    NARCIS (Netherlands)

    Schakenraad, Koen; Biebricher, Andreas S.; Sebregts, Maarten; Ten Bensel, Brian; Peterman, Erwin J.G.; Wuite, Gijs J L; Heller, Iddo; Storm, Cornelis; Van Der Schoot, Paul

    2017-01-01

    The three-dimensional structure of DNA is highly susceptible to changes by mechanical and biochemical cues in vivo and in vitro. In particular, large increases in base pair spacing compared to regular B-DNA are effected by mechanical (over)stretching and by intercalation of compounds that are widely

  7. Involvement of the yeast DNA polymerase delta in DNA repair in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Giot, L. [State University of New York at Stony Brook, Stony Brook, NY. (United States); Chanet, R.; Simon, M.; Facca, C.; Faye, G.

    1997-08-15

    The POL3 encoded catalytic subunit of DNA polymerase delta possesses a highly conserved C-terminal cysteine-rich domain in Saccharomyces cerevisiae. Mutations in some of its cysteine codons display a lethal phenotype, which demonstrates an essential function of this domain. The thermosensitive mutant pol3-13, in which a serine replaces a cysteine of this domain, exhibits a range of defects in DNA repair, such as hypersensitivity to different DNA-damaging agents and deficiency for induced mutagenesis and for recombination. These phenotypes are observed at 24 degrees, a temperature at which DNA replication is almost normal; this differentiates the functions of POL3 in DNA repair and DNA replication. Since spontaneous mutagenesis and spontaneous recombination are efficient in pol3-13, we propose that POL3 plays an important role in DNA repair after irradiation, particularly in the error-prone and recombinational pathways. Extragenic suppressors of pol3-13 are allelic to sdp5-1, previously identified as an extragenic suppressor of pol3-11. SDP5, which is identical to HYS2, encodes a protein homologous to the p50 subunit of bovine and human DNA polymerase delta. SDP5 is most probably the p55 subunit of Pol delta of S. cerevisiae and seems to be associated with the catalytic subunit for both DNA replication and DNA repair. (author)

  8. Same MSH2 Gene Mutation But Variable Phenotypes in 2 Families With Lynch Syndrome: Two Case Reports and Review of Genotype-Phenotype Correlation.

    Science.gov (United States)

    Liccardo, Raffaella; De Rosa, Marina; Duraturo, Francesca

    2018-01-01

    Lynch syndrome is an autosomal dominant syndrome that can be subdivided into Lynch syndrome I, or site-specific colonic cancer, and Lynch syndrome II, or extracolonic cancers, particularly carcinomas of the stomach, endometrium, biliary and pancreatic systems, and urinary tract. Lynch syndrome is associated with point mutations and large rearrangements in DNA MisMatch Repair ( MMR ) genes. This syndrome shows a variable phenotypic expression in people who carry pathogenetic mutations. So far, a correlation in genotype-phenotype has not been definitely established. In this study, we describe 2 Lynch syndrome cases presenting with the same genotype but different phenotypes and discuss possible reasons for this.

  9. Phenotypic plasticity, costs of phenotypes, and costs of plasticity

    DEFF Research Database (Denmark)

    Callahan, Hilary S; Maughan, Heather; Steiner, Uli

    2008-01-01

    Why are some traits constitutive and others inducible? The term costs often appears in work addressing this issue but may be ambiguously defined. This review distinguishes two conceptually distinct types of costs: phenotypic costs and plasticity costs. Phenotypic costs are assessed from patterns...... of covariation, typically between a focal trait and a separate trait relevant to fitness. Plasticity costs, separable from phenotypic costs, are gauged by comparing the fitness of genotypes with equivalent phenotypes within two environments but differing in plasticity and fitness. Subtleties associated with both...... types of costs are illustrated by a body of work addressing predator-induced plasticity. Such subtleties, and potential interplay between the two types of costs, have also been addressed, often in studies involving genetic model organisms. In some instances, investigators have pinpointed the mechanistic...

  10. Deep Phenotyping: Deep Learning For Temporal Phenotype/Genotype Classification

    OpenAIRE

    Najafi, Mohammad; Namin, Sarah; Esmaeilzadeh, Mohammad; Brown, Tim; Borevitz, Justin

    2017-01-01

    High resolution and high throughput, genotype to phenotype studies in plants are underway to accelerate breeding of climate ready crops. Complex developmental phenotypes are observed by imaging a variety of accessions in different environment conditions, however extracting the genetically heritable traits is challenging. In the recent years, deep learning techniques and in particular Convolutional Neural Networks (CNNs), Recurrent Neural Networks (RNNs) and Long-Short Term Memories (LSTMs), h...

  11. Escherichia coli DnaE Polymerase Couples Pyrophosphatase Activity to DNA Replication.

    Directory of Open Access Journals (Sweden)

    Fabio Lapenta

    Full Text Available DNA Polymerases generate pyrophosphate every time they catalyze a step of DNA elongation. This elongation reaction is generally believed as thermodynamically favoured by the hydrolysis of pyrophosphate, catalyzed by inorganic pyrophosphatases. However, the specific action of inorganic pyrophosphatases coupled to DNA replication in vivo was never demonstrated. Here we show that the Polymerase-Histidinol-Phosphatase (PHP domain of Escherichia coli DNA Polymerase III α subunit features pyrophosphatase activity. We also show that this activity is inhibited by fluoride, as commonly observed for inorganic pyrophosphatases, and we identified 3 amino acids of the PHP active site. Remarkably, E. coli cells expressing variants of these catalytic residues of α subunit feature aberrant phenotypes, poor viability, and are subject to high mutation frequencies. Our findings indicate that DNA Polymerases can couple DNA elongation and pyrophosphate hydrolysis, providing a mechanism for the control of DNA extension rate, and suggest a promising target for novel antibiotics.

  12. DNA probes

    International Nuclear Information System (INIS)

    Castelino, J.

    1992-01-01

    The creation of DNA probes for detection of specific nucleotide segments differs from ligand detection in that it is a chemical rather than an immunological reaction. Complementary DNA or RNA is used in place of the antibody and is labelled with 32 P. So far, DNA probes have been successfully employed in the diagnosis of inherited disorders, infectious diseases, and for identification of human oncogenes. The latest approach to the diagnosis of communicable and parasitic infections is based on the use of deoxyribonucleic acid (DNA) probes. The genetic information of all cells is encoded by DNA and DNA probe approach to identification of pathogens is unique because the focus of the method is the nucleic acid content of the organism rather than the products that the nucleic acid encodes. Since every properly classified species has some unique nucleotide sequences that distinguish it from every other species, each organism's genetic composition is in essence a finger print that can be used for its identification. In addition to this specificity, DNA probes offer other advantages in that pathogens may be identified directly in clinical specimens

  13. DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Castelino, J

    1993-12-31

    The creation of DNA probes for detection of specific nucleotide segments differs from ligand detection in that it is a chemical rather than an immunological reaction. Complementary DNA or RNA is used in place of the antibody and is labelled with {sup 32}P. So far, DNA probes have been successfully employed in the diagnosis of inherited disorders, infectious diseases, and for identification of human oncogenes. The latest approach to the diagnosis of communicable and parasitic infections is based on the use of deoxyribonucleic acid (DNA) probes. The genetic information of all cells is encoded by DNA and DNA probe approach to identification of pathogens is unique because the focus of the method is the nucleic acid content of the organism rather than the products that the nucleic acid encodes. Since every properly classified species has some unique nucleotide sequences that distinguish it from every other species, each organism`s genetic composition is in essence a finger print that can be used for its identification. In addition to this specificity, DNA probes offer other advantages in that pathogens may be identified directly in clinical specimens 10 figs, 2 tabs

  14. PHENOTYPIC CORRELATIONS AND BODY WEIGHTS ...

    African Journals Online (AJOL)

    Dr Osondu

    Ethiopian Journal of Environmental Studies and Management Vol. 4 No.3 2011. PHENOTYPIC ... because of its high meat quality and acceptance by her populace. The meat is ... commands high price in the restaurants and markets than other ...

  15. DNA Damage and Pulmonary Hypertension

    Science.gov (United States)

    Ranchoux, Benoît; Meloche, Jolyane; Paulin, Roxane; Boucherat, Olivier; Provencher, Steeve; Bonnet, Sébastien

    2016-01-01

    Pulmonary hypertension (PH) is defined by a mean pulmonary arterial pressure over 25 mmHg at rest and is diagnosed by right heart catheterization. Among the different groups of PH, pulmonary arterial hypertension (PAH) is characterized by a progressive obstruction of distal pulmonary arteries, related to endothelial cell dysfunction and vascular cell proliferation, which leads to an increased pulmonary vascular resistance, right ventricular hypertrophy, and right heart failure. Although the primary trigger of PAH remains unknown, oxidative stress and inflammation have been shown to play a key role in the development and progression of vascular remodeling. These factors are known to increase DNA damage that might favor the emergence of the proliferative and apoptosis-resistant phenotype observed in PAH vascular cells. High levels of DNA damage were reported to occur in PAH lungs and remodeled arteries as well as in animal models of PH. Moreover, recent studies have demonstrated that impaired DNA-response mechanisms may lead to an increased mutagen sensitivity in PAH patients. Finally, PAH was linked with decreased breast cancer 1 protein (BRCA1) and DNA topoisomerase 2-binding protein 1 (TopBP1) expression, both involved in maintaining genome integrity. This review aims to provide an overview of recent evidence of DNA damage and DNA repair deficiency and their implication in PAH pathogenesis. PMID:27338373

  16. DNA methylation

    DEFF Research Database (Denmark)

    Williams, Kristine; Christensen, Jesper; Helin, Kristian

    2012-01-01

    DNA methylation is involved in key cellular processes, including X-chromosome inactivation, imprinting and transcriptional silencing of specific genes and repetitive elements. DNA methylation patterns are frequently perturbed in human diseases such as imprinting disorders and cancer. The recent...... discovery that the three members of the TET protein family can convert 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC) has provided a potential mechanism leading to DNA demethylation. Moreover, the demonstration that TET2 is frequently mutated in haematopoietic tumours suggests that the TET...... proteins are important regulators of cellular identity. Here, we review the current knowledge regarding the function of the TET proteins, and discuss various mechanisms by which they contribute to transcriptional control. We propose that the TET proteins have an important role in regulating DNA methylation...

  17. DNA data

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Raw DNA chromatogram data produced by the ABI 373, 377, 3130 and 3730 automated sequencing machines in ABI format. These are from fish (primarily Sebastes spp.,...

  18. DNA nanotechnology

    Science.gov (United States)

    Seeman, Nadrian C.; Sleiman, Hanadi F.

    2018-01-01

    DNA is the molecule that stores and transmits genetic information in biological systems. The field of DNA nanotechnology takes this molecule out of its biological context and uses its information to assemble structural motifs and then to connect them together. This field has had a remarkable impact on nanoscience and nanotechnology, and has been revolutionary in our ability to control molecular self-assembly. In this Review, we summarize the approaches used to assemble DNA nanostructures and examine their emerging applications in areas such as biophysics, diagnostics, nanoparticle and protein assembly, biomolecule structure determination, drug delivery and synthetic biology. The introduction of orthogonal interactions into DNA nanostructures is discussed, and finally, a perspective on the future directions of this field is presented.

  19. Comparing transformation methods for DNA microarray data

    NARCIS (Netherlands)

    Thygesen, Helene H.; Zwinderman, Aeilko H.

    2004-01-01

    Background: When DNA microarray data are used for gene clustering, genotype/phenotype correlation studies, or tissue classification the signal intensities are usually transformed and normalized in several steps in order to improve comparability and signal/noise ratio. These steps may include

  20. DNA expressions - A formal notation for DNA

    NARCIS (Netherlands)

    Vliet, Rudy van

    2015-01-01

    We describe a formal notation for DNA molecules that may contain nicks and gaps. The resulting DNA expressions denote formal DNA molecules. Different DNA expressions may denote the same molecule. Such DNA expressions are called equivalent. We examine which DNA expressions are minimal, which

  1. What Is Mitochondrial DNA?

    Science.gov (United States)

    ... DNA What is mitochondrial DNA? What is mitochondrial DNA? Although most DNA is packaged in chromosomes within ... proteins. For more information about mitochondria and mitochondrial DNA: Molecular Expressions, a web site from the Florida ...

  2. Establishing the UK DNA Bank for motor neuron disease (MND).

    Science.gov (United States)

    Smith, Lucy; Cupid, B C; Dickie, B G M; Al-Chalabi, A; Morrison, K E; Shaw, C E; Shaw, P J

    2015-07-14

    In 2003 the Motor Neurone Disease (MND) Association, together with The Wellcome Trust, funded the creation of a national DNA Bank specific for MND. It was anticipated that the DNA Bank would constitute an important resource to researchers worldwide and significantly increase activity in MND genetic research. The DNA Bank houses over 3000 high quality DNA samples, all of which were donated by people living with MND, family members and non-related controls, accompanied by clinical phenotype data about the patients. Today the primary focus of the UK MND DNA Bank still remains to identify causative and disease modifying factors for this devastating disease.

  3. Ins and Outs of Multipartite Positive-Strand RNA Plant Viruses: Packaging versus Systemic Spread

    Directory of Open Access Journals (Sweden)

    Mattia Dall’Ara

    2016-08-01

    Full Text Available Viruses possessing a non-segmented genome require a specific recognition of their nucleic acid to ensure its protection in a capsid. A similar feature exists for viruses having a segmented genome, usually consisting of viral genomic segments joined together into one viral entity. While this appears as a rule for animal viruses, the majority of segmented plant viruses package their genomic segments individually. To ensure a productive infection, all viral particles and thereby all segments have to be present in the same cell. Progression of the virus within the plant requires as well a concerted genome preservation to avoid loss of function. In this review, we will discuss the “life aspects” of chosen phytoviruses and argue for the existence of RNA-RNA interactions that drive the preservation of viral genome integrity while the virus progresses in the plant.

  4. Positive-Strand RNA Viruses Infecting the Red Imported Fire Ant, Solenopsis invicta

    Directory of Open Access Journals (Sweden)

    Steven M. Valles

    2012-01-01

    Full Text Available The imported fire ants, Solenopsis invicta and S. richteri were introduced into the USA between 1918 and 1945. Since that time, they have expanded their USA range to include some 138 million hectares. Their introduction has had significant economic consequences with costs associated with damage and control efforts estimated at 6 billion dollars annually in the USA. The general consensus of entomologists and myrmecologists is that permanent, sustainable control of these ants in the USA will likely depend on self-sustaining biological control agents. A metagenomics approach successfully resulted in discovery of three viruses infecting S. invicta. Solenopsis invicta virus 1 (SINV-1, SINV-2, and SINV-3 are all positive, single-stranded RNA viruses and represent the first viral discoveries in any ant species. Molecular characterization, host relationships, and potential development and use of SINV-1, SINV-2, and SINV-3 as biopesticides are discussed.

  5. Repair of DNA damage in Deinococcus radiodurans

    International Nuclear Information System (INIS)

    Evans, D.M.

    1984-01-01

    The repair of DNA lesions in Deinococcus radiodurans was examined with particular reference to DNA excision repair of ultraviolet light (UV) induced pyrimidine dimers. The characteristics of excision repair via UV endonucleases α and β in vivo varied with respect to (a) the substrate range of the enzymes, (b) the rate of repair of DNA damage (c) the requirement for a protein synthesised in response to DNA damage to attenuate exonuclease action at repairing regions. UV endonuclease α is postulated to incise DNA in a different manner from UV endonuclease β thus defining the method of subsequent repair. Several DNA damage specific endonuclease activities independent of α and β are described. Mutations of the uvsA, uvsF and uvsG genes resulted in an increase in single-strand breaks in response to DNA damage producing uncontrolled DNA degradation. Evidence is presented that these genes have a role in limiting the access of UV endonuclease β to DNA lesions. uvsF and uvsG are also shown to be linked to the mtoA gene. Mutation of uvsH and reo-1 produces further distinct phenotypes which are discussed. An overall model of excision repair of DNA damage in Deinococcus radiodurans is presented. (author)

  6. Role of oxidative DNA damage in genome instability and cancer

    International Nuclear Information System (INIS)

    Bignami, M.; Kunkel, T.

    2009-01-01

    Inactivation of mismatch repair (MMR) is associated with a dramatic genomic instability that is observed experimentally as a mutator phenotype and micro satellite instability (MSI). It has been implicit that the massive genetic instability in MMR defective cells simply reflects the accumulation of spontaneous DNA polymerase errors during DNA replication. We recently identified oxidation damage, a common threat to DNA integrity to which purines are very susceptible, as an important cofactor in this genetic instability

  7. Childhood asthma-predictive phenotype.

    Science.gov (United States)

    Guilbert, Theresa W; Mauger, David T; Lemanske, Robert F

    2014-01-01

    Wheezing is a fairly common symptom in early childhood, but only some of these toddlers will experience continued wheezing symptoms in later childhood. The definition of the asthma-predictive phenotype is in children with frequent, recurrent wheezing in early life who have risk factors associated with the continuation of asthma symptoms in later life. Several asthma-predictive phenotypes were developed retrospectively based on large, longitudinal cohort studies; however, it can be difficult to differentiate these phenotypes clinically as the expression of symptoms, and risk factors can change with time. Genetic, environmental, developmental, and host factors and their interactions may contribute to the development, severity, and persistence of the asthma phenotype over time. Key characteristics that distinguish the childhood asthma-predictive phenotype include the following: male sex; a history of wheezing, with lower respiratory tract infections; history of parental asthma; history of atopic dermatitis; eosinophilia; early sensitization to food or aeroallergens; or lower lung function in early life. Copyright © 2014 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  8. Mutagenic DNA repair in enterobacteria

    International Nuclear Information System (INIS)

    Sedgwick, S.G.; Chao Ho; Woodgate, R.

    1991-01-01

    Sixteen species of enterobacteria have been screened for mutagenic DNA repair activity. In Escherichia coli, mutagenic DNA repair is encoded by the umuDC operon. Synthesis of UmuD and UmuC proteins is induced as part of the SOS response to DNA damage, and after induction, the UmuD protein undergoes an autocatalytic cleavage to produce the carboxy-terminal UmuD' fragment needed for induced mutagenesis. The presence of a similar system in other species was examined by using a combined approach of inducible-mutagenesis assays, cross-reactivity to E. coli UmuD and UmuD' antibodies to test for induction and cleavage of UmuD-like proteins, and hybridization with E. coli and Salmonella typhimurium u mu DNA probes to map umu-like genes. The results indicate a more widespread distribution of mutagenic DNA repair in other species than was previously thought. They also show that umu loci can be more complex in other species than in E. coli. Differences in UV-induced mutability of more than 200-fold were seen between different species of enteric bacteria and even between multiple natural isolates of E. coli, and yet some of the species which display a poorly mutable phenotype still have umu-like genes and proteins. It is suggested that umuDC genes can be curtailed in their mutagenic activities but that they may still participate in some other, unknown process which provides the continued stimulus for their retention

  9. Epigenetic changes of DNA repair genes in cancer.

    Science.gov (United States)

    Lahtz, Christoph; Pfeifer, Gerd P

    2011-02-01

    'Every Hour Hurts, The Last One Kills'. That is an old saying about getting old. Every day, thousands of DNA damaging events take place in each cell of our body, but efficient DNA repair systems have evolved to prevent that. However, our DNA repair system and that of most other organisms are not as perfect as that of Deinococcus radiodurans, for example, which is able to repair massive amounts of DNA damage at one time. In many instances, accumulation of DNA damage has been linked to cancer, and genetic deficiencies in specific DNA repair genes are associated with tumor-prone phenotypes. In addition to mutations, which can be either inherited or somatically acquired, epigenetic silencing of DNA repair genes may promote tumorigenesis. This review will summarize current knowledge of the epigenetic inactivation of different DNA repair components in human cancer.

  10. Trabectedin – the DNA minor groove binder

    Directory of Open Access Journals (Sweden)

    G. A. Belitsky

    2015-01-01

    Full Text Available Trabectedin (ET-743, Yondelis is an alkaloid that was originally isolated from the Caribbean Sea squirt, Ecteinascidia turbinata and is now produced synthetically. Its chemical structure consists in three fused tetrahydroisoquinoline rings. Two of them, A and B, binds covalently to guanine residues in the minor groove of the DNA double helix to bend the molecule toward the major groove and the third ring C protrudes from the DNA duplex, apparently allowing interactions with several nuclear proteins. Binding to the minor groove of DNA, trabectedin trigger a cascade of events that interfere with several transcription factors, DNA binding proteins, and DNA repair pathways in particular nucleotide excision repair. It acts both as a DNA-alkylating drug and topoisomerase poison. Trabectedin-DNA adduct traps the nucleotide excision repair proteins repairing the DNA damage in transcribing genes and induces DNA strand breaks. Cells deficient in homologous recombination pathway which repairs these double-strand breaks show increased sensitivity to trabectedin. The most sensitive of them were myxoid liposarcomas. Trabectedin is also effective in chemotherapy-experienced patients with advanced, recurrent liposarcoma or leiomyosarcoma as well as in women with ovarian cancer and breast cancer with BRCAness phenotype. Besides of tumor cells Trabectedin inhibits inflammatory cells by affecting directly monocytes and tumorassociated macrophages and indirectly by inhibiting production of inflammatory mediators, the cytokines and chemokines. It inhibits also the MDR-1 gene, which is responsible for the resistance of cancer cells to chemotherapeutic agents and strikes tumor angiogenesis.

  11. DNA Vaccines

    Indian Academy of Sciences (India)

    diseases. Keywords. DNA vaccine, immune response, antibodies, infectious diseases. GENERAL .... tein vaccines require expensive virus/protein purification tech- niques as ... sphere continue to remain major health hazards in developing nations. ... significance since it can be produced at a very low cost and can be stored ...

  12. DNA Investigations.

    Science.gov (United States)

    Mayo, Ellen S.; Bertino, Anthony J.

    1991-01-01

    Presents a simulation activity that allow students to work through the exercise of DNA profiling and to grapple with some analytical and ethical questions involving a couple arranging with a surrogate mother to have a baby. Can be used to teach the principles of restriction enzyme digestion, gel electrophoresis, and probe hybridization. (MDH)

  13. Multiple sporadic colorectal cancers display a unique methylation phenotype.

    Directory of Open Access Journals (Sweden)

    Victoria Gonzalo

    Full Text Available Epigenetics are thought to play a major role in the carcinogenesis of multiple sporadic colorectal cancers (CRC. Previous studies have suggested concordant DNA hypermethylation between tumor pairs. However, only a few methylation markers have been analyzed. This study was aimed at describing the epigenetic signature of multiple CRC using a genome-scale DNA methylation profiling. We analyzed 12 patients with synchronous CRC and 29 age-, sex-, and tumor location-paired patients with solitary tumors from the EPICOLON II cohort. DNA methylation profiling was performed using the Illumina Infinium HM27 DNA methylation assay. The most significant results were validated by Methylight. Tumors samples were also analyzed for the CpG Island Methylator Phenotype (CIMP; KRAS and BRAF mutations and mismatch repair deficiency status. Functional annotation clustering was performed. We identified 102 CpG sites that showed significant DNA hypermethylation in multiple tumors with respect to the solitary counterparts (difference in β value ≥0.1. Methylight assays validated the results for 4 selected genes (p = 0.0002. Eight out of 12(66.6% multiple tumors were classified as CIMP-high, as compared to 5 out of 29(17.2% solitary tumors (p = 0.004. Interestingly, 76 out of the 102 (74.5% hypermethylated CpG sites found in multiple tumors were also seen in CIMP-high tumors. Functional analysis of hypermethylated genes found in multiple tumors showed enrichment of genes involved in different tumorigenic functions. In conclusion, multiple CRC are associated with a distinct methylation phenotype, with a close association between tumor multiplicity and CIMP-high. Our results may be important to unravel the underlying mechanism of tumor multiplicity.

  14. Phenotypic spectrum of GABRA1

    DEFF Research Database (Denmark)

    Johannesen, Katrine; Marini, Carla; Pfeffer, Siona

    2016-01-01

    OBJECTIVE: To delineate phenotypic heterogeneity, we describe the clinical features of a cohort of patients with GABRA1 gene mutations. METHODS: Patients with GABRA1 mutations were ascertained through an international collaboration. Clinical, EEG, and genetic data were collected. Functional analy...

  15. Leaf segmentation in plant phenotyping

    NARCIS (Netherlands)

    Scharr, Hanno; Minervini, Massimo; French, Andrew P.; Klukas, Christian; Kramer, David M.; Liu, Xiaoming; Luengo, Imanol; Pape, Jean Michel; Polder, Gerrit; Vukadinovic, Danijela; Yin, Xi; Tsaftaris, Sotirios A.

    2016-01-01

    Image-based plant phenotyping is a growing application area of computer vision in agriculture. A key task is the segmentation of all individual leaves in images. Here we focus on the most common rosette model plants, Arabidopsis and young tobacco. Although leaves do share appearance and shape

  16. Delineating SPTAN1 associated phenotypes

    DEFF Research Database (Denmark)

    Syrbe, Steffen; Harms, Frederike L; Parrini, Elena

    2017-01-01

    De novo in-frame deletions and duplications in the SPTAN1 gene, encoding the non-erythrocyte αII spectrin, have been associated with severe West syndrome with hypomyelination and pontocerebellar atrophy. We aimed at comprehensively delineating the phenotypic spectrum associated with SPTAN1 mutati...

  17. Cloning of Salmonella typhimurium DNA encoding mutagenic DNA repair

    International Nuclear Information System (INIS)

    Thomas, S.M.; Sedgwick, S.G.

    1989-01-01

    Mutagenic DNA repair in Escherichia coli is encoded by the umuDC operon. Salmonella typhimurium DNA which has homology with E. coli umuC and is able to complement E. coli umuC122::Tn5 and umuC36 mutations has been cloned. Complementation of umuD44 mutants and hybridization with E. coli umuD also occurred, but these activities were much weaker than with umuC. Restriction enzyme mapping indicated that the composition of the cloned fragment is different from the E. coli umuDC operon. Therefore, a umu-like function of S. typhimurium has been found; the phenotype of this function is weaker than that of its E. coli counterpart, which is consistent with the weak mutagenic response of S. typhimurium to UV compared with the response in E. coli

  18. DNA damage preceding dopamine neuron degeneration in A53T human α-synuclein transgenic mice

    International Nuclear Information System (INIS)

    Wang, Degui; Yu, Tianyu; Liu, Yongqiang; Yan, Jun; Guo, Yingli; Jing, Yuhong; Yang, Xuguang; Song, Yanfeng; Tian, Yingxia

    2016-01-01

    Defective DNA repair has been linked with age-associated neurodegenerative disorders. Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by genetic and environmental factors. Whether damages to nuclear DNA contribute to neurodegeneration of PD still remain obscure. in this study we aim to explore whether nuclear DNA damage induce dopamine neuron degeneration in A53T human α-Synuclein over expressed mouse model. We investigated the effects of X-ray irradiation on A53T-α-Syn MEFs and A53T-α-Syn transgene mice. Our results indicate that A53T-α-Syn MEFs show a prolonged DNA damage repair process and senescense phenotype. DNA damage preceded onset of motor phenotype in A53T-α-Syn transgenic mice and decrease the number of nigrostriatal dopaminergic neurons. Neurons of A53T-α-Syn transgenic mice are more fragile to DNA damages. - Highlights: • This study explore contribution of DNA damage to neurodegeneration in Parkinson's disease mice. • A53T-α-Syn MEF cells show a prolonged DNA damage repair process and senescense phenotype. • DNA damage preceded onset of motor phenotype in A53T-α-Syn transgenic mice. • DNA damage decrease the number of nigrostriatal dopaminergic neurons. • Neurons of A53T-α-Syn transgenic mice are more fragile to DNA damages.

  19. Interoperability between phenotype and anatomy ontologies.

    Science.gov (United States)

    Hoehndorf, Robert; Oellrich, Anika; Rebholz-Schuhmann, Dietrich

    2010-12-15

    Phenotypic information is important for the analysis of the molecular mechanisms underlying disease. A formal ontological representation of phenotypic information can help to identify, interpret and infer phenotypic traits based on experimental findings. The methods that are currently used to represent data and information about phenotypes fail to make the semantics of the phenotypic trait explicit and do not interoperate with ontologies of anatomy and other domains. Therefore, valuable resources for the analysis of phenotype studies remain unconnected and inaccessible to automated analysis and reasoning. We provide a framework to formalize phenotypic descriptions and make their semantics explicit. Based on this formalization, we provide the means to integrate phenotypic descriptions with ontologies of other domains, in particular anatomy and physiology. We demonstrate how our framework leads to the capability to represent disease phenotypes, perform powerful queries that were not possible before and infer additional knowledge. http://bioonto.de/pmwiki.php/Main/PheneOntology.

  20. DNA repair

    International Nuclear Information System (INIS)

    Setlow, R.

    1978-01-01

    Some topics discussed are as follows: difficulty in extrapolating data from E. coli to mammalian systems; mutations caused by UV-induced changes in DNA; mutants deficient in excision repair; other postreplication mechanisms; kinds of excision repair systems; detection of repair by biochemical or biophysical means; human mutants deficient in repair; mutagenic effects of UV on XP cells; and detection of UV-repair defects among XP individuals

  1. DNA-PKcs is critical for telomere capping

    Energy Technology Data Exchange (ETDEWEB)

    Gilley, David; Tanaka, Hiromi; Hande, M. Prakash; Kurimasa,Akihiro; Li, Gloria C.; Chen, David J.

    2001-04-10

    The DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is critical for DNA repair via the non-homologous end joining (NHEJ) pathway. Previously, it was reported that bone marrow cells and spontaneously transformed fibroblasts from SCID (severe combined immunodeficiency) mice have defects in telomere maintenance. The genetically defective SCID mouse arose spontaneously from its parental strain CB17. One known genomic alteration in SCID mice is a truncation of the extreme carboxyl-terminus of DNA-PKcs, but other as yet unidentified alterations may also exist. We have used a defined system, the DNA-PKcs knockout mouse, to investigate specifically the role DNA-PKcs specifically plays in telomere maintenance. We report that primary mouse embryonic fibroblasts (MEFs) and primary cultured kidney cells from 6-8 month old DNA-PKcs deficient mice accumulate a large number of telomere fusions, yet still retain wildtype telomere length. Thus, the phenotype of this defect separates the two-telomere related phenotypes, capping and length maintenance. DNA-PKcs deficient MEFs also exhibit elevated levels of chromosome fragments and breaks, which correlate with increased telomere fusions. Based on the high levels of telomere fusions observed in DNA-PKcs deficient cells, we conclude that DNA-PKcs plays an important capping role at the mammalian telomere.

  2. CpG Island Methylator Phenotype in Primary Gastric Carcinoma

    OpenAIRE

    TOJO Masayuki:筆頭著者; KONISHI Kazuo; YANO Yuichiro; KATAGIRI Atsushi; NOZAWA Hisako; KUBOTA Yutaro; MURAMOTO Takashi; KONDA Kenichi; SHINMURA Kensuke; TAKIMOTO Masafumi; IMAWARI Michio; YOSHIDA Hitoshi

    2013-01-01

    Gastric cancers (GC) with methylation of multiple CpG islands have a CpG island methylator phenotype (CIMP) and they can have different biological features. The aim of this study was to investigate the DNA methylation status of GCs and its association with their clinicopathological features. We evaluated the methylation status of four genes (MINT1, MINT2, MINT25 and MINT31) in 105 primary GCs using bisulfite-pyrosequencing analysis. We classified tumors as CIMP-high (CIMP-H), CIMP-low (CIMP-L...

  3. Radiation-induced senescence-like phenotype in proliferating and plateau-phase vascular endothelial cells

    International Nuclear Information System (INIS)

    Igarashi, Kaori; Sakimoto, Ippei; Kataoka, Keiko; Ohta, Keisuke; Miura, Masahiko

    2007-01-01

    The effects of ionizing radiation (IR) on tumor angiogenesis still remain largely unknown. In this study, we found that IR (8 Gy) induces a high-frequency (80-90%) senescence-like phenotype in vascular endothelial cells (ECs) undergoing exponential growth. This finding allowed us to characterize the IR-induced senescence-like (IRSL) phenotype by examining the gene expression profiles and in vitro angiogenic activities of these ECs. The expression levels of genes associated with cell cycle progression and DNA replication were remarkably reduced in the IRSL ECs. Additionally, the in vitro invasion and migration activities of these cells through Matrigel were significantly suppressed. We also found that confluent ECs exhibited a high-frequency IRSL phenotype when they were replated immediately after irradiation, whereas incubation in plateau-phase conditions reduced the induction of this phenotype and enhanced colony formation. The kinetics of DNA double-strand break repair, which showed a faster time course in confluent ECs than in growing ECs, may contribute to the protective mechanism associated with the IRSL phenotype. These results imply that the IRSL phenotype may be important for determining the angiogenic activity of ECs following irradiation. The present study should contribute to the understanding of the effects of IR on tumor angiogenesis

  4. The CpG island methylator phenotype (CIMP) in colorectal cancer.

    Science.gov (United States)

    Nazemalhosseini Mojarad, Ehsan; Kuppen, Peter Jk; Aghdaei, Hamid Asadzadeh; Zali, Mohammad Reza

    2013-01-01

    It is clear that colorectal cancer (CRC) develops through multiple genetic and epigenetic pathways. These pathways may be determined on the basis of three molecular features: (i) mutations in DNA mismatch repair genes, leading to a DNA microsatellite instability (MSI) phenotype, (ii) mutations in APC and other genes that activate Wnt pathway, characterized by chromosomal instability (CIN) phenotype, and (iii) global genome hypermethylation, resulting in switch off of tumor suppressor genes, indicated as CpG island methylator phenotype (CIMP). Each of these pathways is characterized by specific pathological features, mechanisms of carcinogenesis and process of tumor development. The molecular aspects of these pathways have been used clinically in the diagnosis, screening and management of patients with colorectal cancer. In this review we especially describe various aspects of CIMP, one of the important and rather recently discovered pathways that lead to colorectal cancer.

  5. Automated phenotyping of permanent crops

    Science.gov (United States)

    McPeek, K. Thomas; Steddom, Karl; Zamudio, Joseph; Pant, Paras; Mullenbach, Tyler

    2017-05-01

    AGERpoint is defining a new technology space for the growers' industry by introducing novel applications for sensor technology and data analysis to growers of permanent crops. Serving data to a state-of-the-art analytics engine from a cutting edge sensor platform, a new paradigm in precision agriculture is being developed that allows growers to understand the unique needs of each tree, bush or vine in their operation. Autonomous aerial and terrestrial vehicles equipped with multiple varieties of remote sensing technologies give AGERpoint the ability to measure key morphological and spectral features of permanent crops. This work demonstrates how such phenotypic measurements combined with machine learning algorithms can be used to determine the variety of crops (e.g., almond and pecan trees). This phenotypic and varietal information represents the first step in enabling growers with the ability to tailor their management practices to individual plants and maximize their economic productivity.

  6. Phenotypic deconstruction of gene circuitry.

    Science.gov (United States)

    Lomnitz, Jason G; Savageau, Michael A

    2013-06-01

    It remains a challenge to obtain a global perspective on the behavioral repertoire of complex nonlinear gene circuits. In this paper, we describe a method for deconstructing complex systems into nonlinear sub-systems, based on mathematically defined phenotypes, which are then represented within a system design space that allows the repertoire of qualitatively distinct phenotypes of the complex system to be identified, enumerated, and analyzed. This method efficiently characterizes large regions of system design space and quickly generates alternative hypotheses for experimental testing. We describe the motivation and strategy in general terms, illustrate its use with a detailed example involving a two-gene circuit with a rich repertoire of dynamic behavior, and discuss experimental means of navigating the system design space.

  7. Phenotypic deconstruction of gene circuitry

    Science.gov (United States)

    Lomnitz, Jason G.; Savageau, Michael A.

    2013-06-01

    It remains a challenge to obtain a global perspective on the behavioral repertoire of complex nonlinear gene circuits. In this paper, we describe a method for deconstructing complex systems into nonlinear sub-systems, based on mathematically defined phenotypes, which are then represented within a system design space that allows the repertoire of qualitatively distinct phenotypes of the complex system to be identified, enumerated, and analyzed. This method efficiently characterizes large regions of system design space and quickly generates alternative hypotheses for experimental testing. We describe the motivation and strategy in general terms, illustrate its use with a detailed example involving a two-gene circuit with a rich repertoire of dynamic behavior, and discuss experimental means of navigating the system design space.

  8. Wine Expertise Predicts Taste Phenotype.

    Science.gov (United States)

    Hayes, John E; Pickering, Gary J

    2012-03-01

    Taste phenotypes have long been studied in relation to alcohol intake, dependence, and family history, with contradictory findings. However, on balance - with appropriate caveats about populations tested, outcomes measured and psychophysical methods used - an association between variation in taste responsiveness and some alcohol behaviors is supported. Recent work suggests super-tasting (operationalized via propylthiouracil (PROP) bitterness) not only associates with heightened response but also with more acute discrimination between stimuli. Here, we explore relationships between food and beverage adventurousness and taste phenotype. A convenience sample of wine drinkers (n=330) were recruited in Ontario and phenotyped for PROP bitterness via filter paper disk. They also filled out a short questionnaire regarding willingness to try new foods, alcoholic beverages and wines as well as level of wine involvement, which was used to classify them as a wine expert (n=110) or wine consumer (n=220). In univariate logisitic models, food adventurousness predicted trying new wines and beverages but not expertise. Likewise, wine expertise predicted willingness to try new wines and beverages but not foods. In separate multivariate logistic models, willingness to try new wines and beverages was predicted by expertise and food adventurousness but not PROP. However, mean PROP bitterness was higher among wine experts than wine consumers, and the conditional distribution functions differed between experts and consumers. In contrast, PROP means and distributions did not differ with food adventurousness. These data suggest individuals may self-select for specific professions based on sensory ability (i.e., an active gene-environment correlation) but phenotype does not explain willingness to try new stimuli.

  9. From plant genomes to phenotypes

    OpenAIRE

    Bolger, Marie; Gundlach, Heidrun; Scholz, Uwe; Mayer, Klaus; Usadel, Björn; Schwacke, Rainer; Schmutzer, Thomas; Chen, Jinbo; Arend, Daniel; Oppermann, Markus; Weise, Stephan; Lange, Matthias; Fiorani, Fabio; Spannagl, Manuel

    2017-01-01

    Recent advances in sequencing technologies have greatly accelerated the rate of plant genome and applied breeding research. Despite this advancing trend, plant genomes continue to present numerous difficulties to the standard tools and pipelines not only for genome assembly but also gene annotation and downstream analysis.Here we give a perspective on tools, resources and services necessary to assemble and analyze plant genomes and link them to plant phenotypes.

  10. Phenotype-Driven Plasma Biobanking Strategies and Methods

    Directory of Open Access Journals (Sweden)

    Erica A. Bowton

    2015-05-01

    Full Text Available Biobank development and integration with clinical data from electronic medical record (EMR databases have enabled recent strides in genomic research and personalized medicine. BioVU, Vanderbilt’s DNA biorepository linked to de-identified clinical EMRs, has proven fruitful in its capacity to extensively appeal to numerous areas of biomedical and clinical research, supporting the discovery of genotype-phenotype interactions. Expanding on experiences in BioVU creation and development, we have recently embarked on a parallel effort to collect plasma in addition to DNA from blood specimens leftover after routine clinical testing at Vanderbilt. This initiative offers expanded utility of BioVU by combining proteomic and metabolomic approaches with genomics and/or clinical outcomes, widening the breadth for potential research and subsequent future impact on clinical care. Here, we describe the considerations and components involved in implementing a plasma biobank program from a feasibility assessment through pilot sample collection.

  11. Phenotypic covariance at species' borders.

    Science.gov (United States)

    Caley, M Julian; Cripps, Edward; Game, Edward T

    2013-05-28

    Understanding the evolution of species limits is important in ecology, evolution, and conservation biology. Despite its likely importance in the evolution of these limits, little is known about phenotypic covariance in geographically marginal populations, and the degree to which it constrains, or facilitates, responses to selection. We investigated phenotypic covariance in morphological traits at species' borders by comparing phenotypic covariance matrices (P), including the degree of shared structure, the distribution of strengths of pair-wise correlations between traits, the degree of morphological integration of traits, and the ranks of matricies, between central and marginal populations of three species-pairs of coral reef fishes. Greater structural differences in P were observed between populations close to range margins and conspecific populations toward range centres, than between pairs of conspecific populations that were both more centrally located within their ranges. Approximately 80% of all pair-wise trait correlations within populations were greater in the north, but these differences were unrelated to the position of the sampled population with respect to the geographic range of the species. Neither the degree of morphological integration, nor ranks of P, indicated greater evolutionary constraint at range edges. Characteristics of P observed here provide no support for constraint contributing to the formation of these species' borders, but may instead reflect structural change in P caused by selection or drift, and their potential to evolve in the future.

  12. Adaptive evolution of molecular phenotypes

    International Nuclear Information System (INIS)

    Held, Torsten; Nourmohammad, Armita; Lässig, Michael

    2014-01-01

    Molecular phenotypes link genomic information with organismic functions, fitness, and evolution. Quantitative traits are complex phenotypes that depend on multiple genomic loci. In this paper, we study the adaptive evolution of a quantitative trait under time-dependent selection, which arises from environmental changes or through fitness interactions with other co-evolving phenotypes. We analyze a model of trait evolution under mutations and genetic drift in a single-peak fitness seascape. The fitness peak performs a constrained random walk in the trait amplitude, which determines the time-dependent trait optimum in a given population. We derive analytical expressions for the distribution of the time-dependent trait divergence between populations and of the trait diversity within populations. Based on this solution, we develop a method to infer adaptive evolution of quantitative traits. Specifically, we show that the ratio of the average trait divergence and the diversity is a universal function of evolutionary time, which predicts the stabilizing strength and the driving rate of the fitness seascape. From an information-theoretic point of view, this function measures the macro-evolutionary entropy in a population ensemble, which determines the predictability of the evolutionary process. Our solution also quantifies two key characteristics of adapting populations: the cumulative fitness flux, which measures the total amount of adaptation, and the adaptive load, which is the fitness cost due to a population's lag behind the fitness peak. (paper)

  13. Epigenome-associated phenotypic acclimatization to ocean acidification in a reef-building coral

    KAUST Repository

    Liew, Yi Jin; Zoccola, Didier; Li, Yong; Tambutté , Eric; Venn, Alexander A.; Michell, Craig; Cui, Guoxin; Deutekom, Eva S.; Kaandorp, Jaap A.; Voolstra, Christian R.; Forê t, Sylvain; Allemand, Denis; Tambutté , Sylvie; Aranda, Manuel

    2017-01-01

    Over the last century, the anthropogenic production of CO2 has led to warmer (+0.74 C) and more acidic (-0.1 pH) oceans, resulting in increasingly frequent and severe mass bleaching events worldwide that precipitate global coral reef decline. To mitigate this decline, proposals to augment the stress tolerance of corals through genetic and non-genetic means have been gaining traction. Work on model systems has shown that environmentally induced alterations in DNA methylation can lead to phenotypic acclimatization. While DNA methylation has been observed in corals, its potential role in phenotypic plasticity has not yet been described. Here, we show that, similar to findings in mice, DNA methylation significantly reduces spurious transcription in the Red Sea coral Stylophora pistillata, suggesting the evolutionary conservation of this essential mechanism in corals. Furthermore, we find that DNA methylation also reduces transcriptional noise by fine-tuning the expression of highly expressed genes. Analysis of DNA methylation patterns of corals subjected to long-term pH stress showed widespread changes in pathways regulating cell cycle and body size. Correspondingly, we found significant increases in cell and polyp sizes that resulted in more porous skeletons, supporting the maintenance of linear extension rates under conditions of reduced calcification. These findings suggest an epigenetic component in phenotypic acclimatization, providing corals with an additional mechanism to cope with climate change.

  14. Epigenome-associated phenotypic acclimatization to ocean acidification in a reef-building coral

    KAUST Repository

    Liew, Yi Jin

    2017-09-14

    Over the last century, the anthropogenic production of CO2 has led to warmer (+0.74 C) and more acidic (-0.1 pH) oceans, resulting in increasingly frequent and severe mass bleaching events worldwide that precipitate global coral reef decline. To mitigate this decline, proposals to augment the stress tolerance of corals through genetic and non-genetic means have been gaining traction. Work on model systems has shown that environmentally induced alterations in DNA methylation can lead to phenotypic acclimatization. While DNA methylation has been observed in corals, its potential role in phenotypic plasticity has not yet been described. Here, we show that, similar to findings in mice, DNA methylation significantly reduces spurious transcription in the Red Sea coral Stylophora pistillata, suggesting the evolutionary conservation of this essential mechanism in corals. Furthermore, we find that DNA methylation also reduces transcriptional noise by fine-tuning the expression of highly expressed genes. Analysis of DNA methylation patterns of corals subjected to long-term pH stress showed widespread changes in pathways regulating cell cycle and body size. Correspondingly, we found significant increases in cell and polyp sizes that resulted in more porous skeletons, supporting the maintenance of linear extension rates under conditions of reduced calcification. These findings suggest an epigenetic component in phenotypic acclimatization, providing corals with an additional mechanism to cope with climate change.

  15. DNA repair

    International Nuclear Information System (INIS)

    Van Zeeland, A.A.

    1984-01-01

    In this chapter a series of DNA repair pathways are discussed which are available to the cell to cope with the problem of DNA damaged by chemical or physical agents. In the case of microorganisms our knowledge about the precise mechanism of each DNA repair pathway and the regulation of it has been improved considerably when mutants deficient in these repair mechanisms became available. In the case of mammalian cells in culture, until recently there were very little repair deficient mutants available, because in almost all mammalian cells in culture at least the diploid number of chromosomes is present. Therefore the frequency of repair deficient mutants in such populations is very low. Nevertheless because replica plating techniques are improving some mutants from Chinese hamsters ovary cells and L5178Y mouse lymphoma cells are now available. In the case of human cells, cultures obtained from patients with certain genetic diseases are available. A number of cells appear to be sensitive to some chemical or physical mutagens. These include cells from patients suffering from xeroderma pigmentosum, Ataxia telangiectasia, Fanconi's anemia, Cockayne's syndrome. However, only in the case of xeroderma pigmentosum cells, has the sensitivity to ultraviolet light been clearly correlated with a deficiency in excision repair of pyrimidine dimers. Furthermore the work with strains obtained from biopsies from man is difficult because these cells generally have low cloning efficiencies and also have a limited lifespan in vitro. It is therefore very important that more repair deficient mutants will become available from established cell lines from human or animal origin

  16. Epidemiological markers in Neisseria meningitidis: an estimate of the performance of genotyping vs phenotyping

    DEFF Research Database (Denmark)

    Weis, N; Lind, I

    1998-01-01

    In order to estimate the performance of genotypic vs phenotypic characterization of Neisseria meningitidis, 2 methods, DNA fingerprinting and multilocus enzyme electrophoresis (MEE), were assessed as regards applicability, reproducibility and discriminating capacity. 50 serogroup B and 52 serogroup...... C Neisseria meningitidis strains from 96 patients with meningococcal disease and 22 serogroup C strains from healthy carriers were investigated. Both methods were 100% applicable to meningococcal strains and results of DNA fingerprinting as well as of MEE were reproducible. The number of types...... defined by DNA fingerprinting and MEE as compared to that defined by phenotypic characteristics (serogroup, serotype, serosubtype and sulphonamide resistance) was as follows: for serogroup B strains from patients, 11 and 12 vs 8; for serogroup C strains from patients, 10 and 15 vs 8; and for serogroup C...

  17. Translation of Genotype to Phenotype by a Hierarchy of Cell Subsystems.

    Science.gov (United States)

    Yu, Michael Ku; Kramer, Michael; Dutkowski, Janusz; Srivas, Rohith; Licon, Katherine; Kreisberg, Jason; Ng, Cherie T; Krogan, Nevan; Sharan, Roded; Ideker, Trey

    2016-02-24

    Accurately translating genotype to phenotype requires accounting for the functional impact of genetic variation at many biological scales. Here we present a strategy for genotype-phenotype reasoning based on existing knowledge of cellular subsystems. These subsystems and their hierarchical organization are defined by the Gene Ontology or a complementary ontology inferred directly from previously published datasets. Guided by the ontology's hierarchical structure, we organize genotype data into an "ontotype," that is, a hierarchy of perturbations representing the effects of genetic variation at multiple cellular scales. The ontotype is then interpreted using logical rules generated by machine learning to predict phenotype. This approach substantially outperforms previous, non-hierarchical methods for translating yeast genotype to cell growth phenotype, and it accurately predicts the growth outcomes of two new screens of 2,503 double gene knockouts impacting DNA repair or nuclear lumen. Ontotypes also generalize to larger knockout combinations, setting the stage for interpreting the complex genetics of disease.

  18. Induced pluripotent stem cells with a pathological mitochondrial DNA deletion

    Science.gov (United States)

    Cherry, Anne B. C.; Gagne, Katelyn E.; McLoughlin, Erin M.; Baccei, Anna; Gorman, Bryan; Hartung, Odelya; Miller, Justine D.; Zhang, Jin; Zon, Rebecca L.; Ince, Tan A.; Neufeld, Ellis J.; Lerou, Paul H.; Fleming, Mark D.; Daley, George Q.; Agarwal, Suneet

    2013-01-01

    In congenital mitochondrial DNA (mtDNA) disorders, a mixture of normal and mutated mtDNA (termed heteroplasmy) exists at varying levels in different tissues, which determines the severity and phenotypic expression of disease. Pearson marrow pancreas syndrome (PS) is a congenital bone marrow failure disorder caused by heteroplasmic deletions in mtDNA. The cause of the hematopoietic failure in PS is unknown, and adequate cellular and animal models are lacking. Induced pluripotent stem (iPS) cells are particularly amenable for studying mtDNA disorders, as cytoplasmic genetic material is retained during direct reprogramming. Here we derive and characterize iPS cells from a patient with PS. Taking advantage of the tendency for heteroplasmy to change with cell passage, we isolated isogenic PS-iPS cells without detectable levels of deleted mtDNA. We found that PS-iPS cells carrying a high burden of deleted mtDNA displayed differences in growth, mitochondrial function, and hematopoietic phenotype when differentiated in vitro, compared to isogenic iPS cells without deleted mtDNA. Our results demonstrate that reprogramming somatic cells from patients with mtDNA disorders can yield pluripotent stem cells with varying burdens of heteroplasmy that might be useful in the study and treatment of mitochondrial diseases. PMID:23400930

  19. Knowledge-based analysis of phenotypes

    KAUST Repository

    Hoendorf, Robert

    2016-01-27

    Phenotypes are the observable characteristics of an organism, and they are widely recorded in biology and medicine. To facilitate data integration, ontologies that formally describe phenotypes are being developed in several domains. I will describe a formal framework to describe phenotypes. A formalized theory of phenotypes is not only useful for domain analysis, but can also be applied to assist in the diagnosis of rare genetic diseases, and I will show how our results on the ontology of phenotypes is now applied in biomedical research.

  20. Peruvian Maca (Lepidium peruvianum): (I) Phytochemical and Genetic Differences in Three Maca Phenotypes.

    Science.gov (United States)

    Meissner, Henry O; Mscisz, Alina; Mrozikiewicz, Mieczyslaw; Baraniak, Marek; Mielcarek, Sebastian; Kedzia, Bogdan; Piatkowska, Ewa; Jólkowska, Justyna; Pisulewski, Pawel

    2015-09-01

    Glucosinolates were previously reported as physiologically-important constituents present in Peruvian Maca (Lepidium peruvianum Chacon) and linked to various therapeutic functions of differently-colored Peruvian Maca hypocotyls. In two separate Trials, three colours of Maca hypocotyls "Black", "Red" and "Yellow" (termed "Maca phenotypes"), were selected from mixed crops of Peruvian Maca for laboratory studies as fresh and after being dried. Individual Maca phenotypes were cultivated in the highlands of the Peruvian Andes at 4,200m a.s.l. (Junin and Ninacaca). Glucosinolate levels, chromatographic HPLC profiles and DNA variability in the investigated Maca phenotypes are presented. Genotypic profiles were determined by the ISSR-PCR and RAPD techniques. Compared to the Black and Red phenotypes, the Yellow phenotype contained much lower Glucosinolate levels measured against Glucotropaeolin and m-methoxy-glucotropaeolin standards, and exhibited different RAPD and ISSR-PCR reactions. The Red Maca phenotype showed the highest concentrations of Glucosinolates as compared to the Black and Yellow Maca. It appears that the traditional system used by natives of the Peruvian Andean highlands in preparing Maca as a vegetable dish (boiling dried Maca after soaking in water), to supplement their daily meals, is as effective as laboratory methods - for extracting Glucosinolates, which are considered to be one of the key bioactive constituents responsible for therapeutic functions of Peruvian Maca phenotypes. It is reasonable to assume that the HPLC and DNA techniques combined, or separately, may assist in determining ID and "Fingerprints" identifying individual Peruvian Maca phenotypes, hence confirming the authenticity of marketable Maca products. The above assumptions warrant further laboratory testing.

  1. NIH Mouse Metabolic Phenotyping Centers: the power of centralized phenotyping.

    Science.gov (United States)

    Laughlin, Maren R; Lloyd, K C Kent; Cline, Gary W; Wasserman, David H

    2012-10-01

    The Mouse Metabolic Phenotyping Centers (MMPCs) were founded in 2001 by the National Institutes of Health (NIH) to advance biomedical research by providing the scientific community with standardized, high-quality phenotyping services for mouse models of diabetes, obesity, and their complications. The intent is to allow researchers to take optimum advantage of the many new mouse models produced in labs and in high-throughput public efforts. The six MMPCs are located at universities around the country and perform complex metabolic tests in intact mice and hormone and analyte assays in tissues on a fee-for-service basis. Testing is subsidized by the NIH in order to reduce the barriers for mouse researchers. Although data derived from these tests belong to the researcher submitting mice or tissues, these data are archived after publication in a public database run by the MMPC Coordinating and Bioinformatics Unit. It is hoped that data from experiments performed in many mouse models of metabolic diseases, using standard protocols, will be useful in understanding the nature of these complex disorders. The current areas of expertise include energy balance and body composition, insulin action and secretion, whole-body and tissue carbohydrate and lipid metabolism, cardiovascular and renal function, and metabolic pathway kinetics. In addition to providing services, the MMPC staff provides expertise and advice to researchers, and works to develop and refine test protocols to best meet the community's needs in light of current scientific developments. Test technology is disseminated by publications and through annual courses.

  2. The Human Phenotype Ontology in 2017

    International Nuclear Information System (INIS)

    Köhler, Sebastian; Vasilevsky, Nicole A.; Engelstad, Mark; Foster, Erin; McMurry, Julie

    2016-01-01

    Deep phenotyping has been defined as the precise and comprehensive analysis of phenotypic abnormalities in which the individual components of the phenotype are observed and described. The three components of the Human PhenotypeOntology (HPO; www.human-phenotype-ontology.org) project are the phenotype vocabulary, disease-phenotype annotations and the algorithms that operate on these. These components are being used for computational deep phenotyping and precision medicine as well as integration of clinical data into translational research. The HPO is being increasingly adopted as a standard for phenotypic abnormalities by diverse groups such as international rare disease organizations, registries, clinical labs, biomedical resources, and clinical software tools and will thereby contribute toward nascent efforts at global data exchange for identifying disease etiologies. This update article reviews the progress of the HPO project since the debut Nucleic Acids Research database article in 2014, including specific areas of expansion such as common (complex) disease, new algorithms for phenotype driven genomic discovery and diagnostics, integration of cross-species mapping efforts with the Mammalian Phenotype Ontology, an improved quality control pipeline, and the addition of patient-friendly terminology.

  3. Involvement of specialized DNA polymerases Pol II, Pol IV and DnaE2 in DNA replication in the absence of Pol I in Pseudomonas putida

    International Nuclear Information System (INIS)

    Sidorenko, Julia; Jatsenko, Tatjana; Saumaa, Signe; Teras, Riho; Tark-Dame, Mariliis; Horak, Rita; Kivisaar, Maia

    2011-01-01

    The majority of bacteria possess a different set of specialized DNA polymerases than those identified in the most common model organism Escherichia coli. Here, we have studied the ability of specialized DNA polymerases to substitute Pol I in DNA replication in Pseudomonas putida. Our results revealed that P. putida Pol I-deficient cells have severe growth defects in LB medium, which is accompanied by filamentous cell morphology. However, growth of Pol I-deficient bacteria on solid rich medium can be restored by reduction of reactive oxygen species in cells. Also, mutants with improved growth emerge rapidly. Similarly to the initial Pol I-deficient P. putida, its adapted derivatives express a moderate mutator phenotype, which indicates that DNA replication carried out in the absence of Pol I is erroneous both in the original Pol I-deficient bacteria and the adapted derivatives. Analysis of the spectra of spontaneous Rif r mutations in P. putida strains lacking different DNA polymerases revealed that the presence of specialized DNA polymerases Pol II and Pol IV influences the frequency of certain base substitutions in Pol I-proficient and Pol I-deficient backgrounds in opposite ways. Involvement of another specialized DNA polymerase DnaE2 in DNA replication in Pol I-deficient bacteria is stimulated by UV irradiation of bacteria, implying that DnaE2-provided translesion synthesis partially substitutes the absence of Pol I in cells containing heavily damaged DNA.

  4. Emergent properties of gene evolution: Species as attractors in phenotypic space

    Science.gov (United States)

    Reuveni, Eli; Giuliani, Alessandro

    2012-02-01

    The question how the observed discrete character of the phenotype emerges from a continuous genetic distance metrics is the core argument of two contrasted evolutionary theories: punctuated equilibrium (stable evolution scattered with saltations in the phenotype) and phyletic gradualism (smooth and linear evolution of the phenotype). Identifying phenotypic saltation on the molecular levels is critical to support the first model of evolution. We have used DNA sequences of ∼1300 genes from 6 isolated populations of the budding yeast Saccharomyces cerevisiae. We demonstrate that while the equivalent measure of the genetic distance show a continuum between lineage distance with no evidence of discrete states, the phenotypic space illustrates only two (discrete) possible states that can be associated with a saltation of the species phenotype. The fact that such saltation spans large fraction of the genome and follows by continuous genetic distance is a proof of the concept that the genotype-phenotype relation is not univocal and may have severe implication when looking for disease related genes and mutations. We used this finding with analogy to attractor-like dynamics and show that punctuated equilibrium could be explained in the framework of non-linear dynamics systems.

  5. Genotypic and phenotypic diversity of Alicyclobacillus acidocaldarius isolates.

    Science.gov (United States)

    Félix-Valenzuela, L; Guardiola-Avila, I; Burgara-Estrella, A; Ibarra-Zavala, M; Mata-Haro, V

    2015-10-01

    The fruit juice industry recognizes Alicyclobacillus as a major quality control target micro-organism. In this study, we analysed 19 bacterial isolates to identify Alicyclobacillus species by polymerase chain reaction (PCR) and sequencing analyses. Phenotypic and genomic diversity among isolates were investigated by API 50CHB system and ERIC-PCR (enterobacterial repetitive intergenic consensus-PCR) respectively. All bacterial isolates were identified as Alicyclobacillus acidocaldarius, and almost all showed identical DNA sequences according to their 16S rRNA (rDNA) gene partial sequences. Only few carbohydrates were fermented by A. acidocaldarius isolates, and there was little variability in the biochemical profile. Genotypic fingerprinting of the A. acidocaldarius isolates showed high diversity, and clusters by ERIC-PCR were distinct to those obtained from the 16S rRNA gene phylogenetic tree. There was no correlation between phenotypic and genotypic variability in the A. acidocaldarius isolates analysed in this study. Detection of Alicyclobacillus strains is imperative in fruit concentrates and juices due to the production of guaiacol. Identification of the genera originates rejection of the product by processing industry. However, not all the Alicyclobacillus species are deteriorative and hence the importance to differentiate among them. In this study, partial 16S ribosomal RNA sequence alignment allowed the differentiation of species. In addition, ERIC-PCR was introduced for the genotypic characterization of Alicyclobacillus, as an alternative for differentiation among isolates from the same species. © 2015 The Society for Applied Microbiology.

  6. Using systems and structure biology tools to dissect cellular phenotypes.

    Science.gov (United States)

    Floratos, Aris; Honig, Barry; Pe'er, Dana; Califano, Andrea

    2012-01-01

    The Center for the Multiscale Analysis of Genetic Networks (MAGNet, http://magnet.c2b2.columbia.edu) was established in 2005, with the mission of providing the biomedical research community with Structural and Systems Biology algorithms and software tools for the dissection of molecular interactions and for the interaction-based elucidation of cellular phenotypes. Over the last 7 years, MAGNet investigators have developed many novel analysis methodologies, which have led to important biological discoveries, including understanding the role of the DNA shape in protein-DNA binding specificity and the discovery of genes causally related to the presentation of malignant phenotypes, including lymphoma, glioma, and melanoma. Software tools implementing these methodologies have been broadly adopted by the research community and are made freely available through geWorkbench, the Center's integrated analysis platform. Additionally, MAGNet has been instrumental in organizing and developing key conferences and meetings focused on the emerging field of systems biology and regulatory genomics, with special focus on cancer-related research.

  7. Phenotypic variability in Meesmann's dystrophy

    DEFF Research Database (Denmark)

    Ehlers, Niels; Hjortdal, Jesper; Nielsen, Kim

    2008-01-01

    symptoms often include blurred vision and ocular irritation. Typical cases may be entirely free of complaints. Intermittent pain episodes, such as occur in recurrent erosion syndrome, are not the rule. Genetic sequencing indicated a familial relationship with the originally described Meesmann family......'s dystrophy occurs worldwide. The largest family described is the original German one, now supplemented with a Danish branch. Despite the presence of an identical genetic defect, the clinical phenotype varies. This suggests that non-KRT12-related mechanisms are responsible for the variation....

  8. The thrifty phenotype hypothesis revisited

    DEFF Research Database (Denmark)

    Vaag, A A; Grunnet, L G; Arora, G P

    2012-01-01

    Twenty years ago, Hales and Barker along with their co-workers published some of their pioneering papers proposing the 'thrifty phenotype hypothesis' in Diabetologia (4;35:595-601 and 3;36:62-67). Their postulate that fetal programming could represent an important player in the origin of type 2...... of the underlying molecular mechanisms. Type 2 diabetes is a multiple-organ disease, and developmental programming, with its idea of organ plasticity, is a plausible hypothesis for a common basis for the widespread organ dysfunctions in type 2 diabetes and the metabolic syndrome. Only two among the 45 known type 2...

  9. DNA repair mechanism in radioresistant bacteria

    International Nuclear Information System (INIS)

    Kitayama, Shigeru

    1992-01-01

    Many radiation resistant bacteria have been isolated from various sources which are not in high background field. Since Deinococcus radiodurans had been isolated first in 1956, studies on the mechanism for radioresistance were carried out mostly using this bacterium. DNA in this bacterium isn't protected against injury induced by not only ionizing radiation but also ultraviolet light. Therefore, DNA damages induced by various treatments are efficiently and accurately repaired in this cells. Damages in base and/or sugar in DNA are removed by endonucleases which, if not all, are synthesized during postirradiation incubation. Following the endonucleolytic cleavage the strand scissions in DNA are seemed to be rejoined by a process common for the repair of strand scissions induced by such as ionizing radiations. Induce protein(s) is also involved in this rejoining process of strand scissions. DNA repair genes were classified into three phenotypic groups. (1)Genes which are responsible for the endonucleolytic activities. (2) Genes involved in the rejoining of DNA strand scissions. (3) Genes which participate in genetic recombination and repair. Three genes belong to (1) and (2) were cloned onto approximately 1 kbp DNA fragments which base sequences have been determined. (author)

  10. DNA repair mechanism in radioresistant bacteria

    International Nuclear Information System (INIS)

    Kitayama, Shigeru

    1992-01-01

    Many radiation resistant bacteria have been isolated from various sources which are not in high background field. Since Deinococcus radiodurans had been isolated first in 1956, the studies on the mechanism of radioresistance were mostly carried out using this bacterium. DNA in this bacterium isn't protected against injury induced by not only ionizing radiation but also ultraviolet light. Therefore, DNA damages induced by various treatments are efficiently and accurately repaired in this cells. Damages in base and/or sugar in DNA are removed by endonucleases which, if not all, are synthesized during postirradiation incubation. Following the endonucleolytic cleavage the strand scissions in DNA are seemed to be rejoined by a process common for the repair of strand scissions induced by such as ionizing radiations. Induce protein(s) is also involved in this rejoining process of strand scissions. DNA repair genes were classified into three phenotypic groups. (1) Genes which are responsible for the endonucleolytic activities. (2) Genes involved in the rejoining of DNA strand scissions. (3) Genes which participate in genetic recombination and repair. Three genes belong to (1) and (2) were cloned onto approximately 1 kbp DNA fragments which base sequences have been determined. (author)

  11. A newly identified DNA ligase of Saccharomyces cerevisiae involved in RAD52-independent repair of DNA double-strand breaks

    Science.gov (United States)

    Schär, Primo; Herrmann, Gernot; Daly, Graham; Lindahl, Tomas

    1997-01-01

    Eukaryotic DNA ligases are ATP-dependent DNA strand-joining enzymes that participate in DNA replication, repair, and recombination. Whereas mammalian cells contain several different DNA ligases, encoded by at least three distinct genes, only one DNA ligase has been detected previously in either budding yeast or fission yeast. Here, we describe a newly identified nonessential Saccharomyces cerevisiae gene that encodes a DNA ligase distinct from the CDC9 gene product. This DNA ligase shares significant amino acid sequence homology with human DNA ligase IV; accordingly, we designate the yeast gene LIG4. Recombinant LIG4 protein forms a covalent enzyme-AMP complex and can join a DNA single-strand break in a DNA/RNA hybrid duplex, the preferred substrate in vitro. Disruption of the LIG4 gene causes only marginally increased cellular sensitivity to several DNA damaging agents, and does not further sensitize cdc9 or rad52 mutant cells. In contrast, lig4 mutant cells have a 1000-fold reduced capacity for correct recircularization of linearized plasmids by illegitimate end-joining after transformation. Moreover, homozygous lig4 mutant diploids sporulate less efficiently than isogenic wild-type cells, and show retarded progression through meiotic prophase I. Spore viability is normal, but lig4 mutants appear to produce a higher proportion of tetrads with only three viable spores. The mutant phenotypes are consistent with functions of LIG4 in an illegitimate DNA end-joining pathway and ensuring efficient meiosis. PMID:9271115

  12. CpG island methylator phenotype (CIMP) in cancer: causes and implications.

    Science.gov (United States)

    Teodoridis, Jens M; Hardie, Catriona; Brown, Robert

    2008-09-18

    Strong evidence exists for a subgroup of tumours, from a variety of tissue types, exhibiting concordant tumour specific DNA methylation: the "CpG island methylator phenotype" (CIMP). Occurrence of CIMP is associated with a range of genetic and environmental factors, although the molecular causes are not well-understood. Both increased expression and aberrant targeting of DNA methyltransferases (DNMTs) could contribute to the occurrence of CIMP. One under-explored area is the possibility that DNA damage may induce or select for CIMP during carcinogenesis or treatment of tumours with chemotherapy. DNA damaging agents can induce DNA damage at guanine rich regions throughout the genome, including CpG islands. This DNA damage can result in stalled DNA synthesis, which will lead to localised increased DNMT1 concentration and therefore potentially increased DNA methylation at these sites. Chemotherapy can select for cells which have increased tolerance to DNA damage due to increased lesion bypass, in some cases by mechanisms which involve inactivation of genes by CpG island methylation. CIMP has been associated with worse patient prognosis, probably due to increased epigenetic plasticity. Therefore, further clinical testing of the diagnostic and prognostic value of the current CIMP markers, as well as increasing our understanding of the molecular causes underlying CIMP are required.

  13. The Composition and Metabolic Phenotype of Neisseria gonorrhoeae Biofilms

    Directory of Open Access Journals (Sweden)

    Michael A Apicella

    2011-04-01

    Full Text Available N. gonorrhoeae has been shown to form biofilms during cervical infection. Thus, biofilm formation may play an important role in the infection of women. The ability of N. gonorrhoeae to form membrane blebs is crucial to biofilm formation. Blebs contain DNA and outer membrane structures, which have been shown to be major constituents of the biofilm matrix. The organism expresses a DNA thermonuclease that is involved in remodeling of the biofilm matrix. Comparison of the transcriptional profiles of gonococcal biofilms and planktonic runoff indicate that genes involved in anaerobic metabolism and oxidative stress tolerance are more highly expressed in biofilm. The expression of aniA, ccp, and norB, which encode nitrite reductase, cytochrome c peroxidase, and nitric oxide reductase respectively, is required for mature biofilm formation over glass and human cervical cells. In addition, anaerobic respiration occurs in the substratum of gonococcal biofilms and disruption of the norB gene required for anaerobic respiration, results in a severe biofilm attenuation phenotype. It has been demonstrated that accumulation of nitric oxide (NO contributes to the phenotype of a norB mutant and can retard biofilm formation. However, NO can also enhance biofilm formation, and this is largely dependent on the concentration and donation rate or steady state kinetics of NO. The majority of the genes involved in gonococcal oxidative stress tolerance are also required for normal biofilm formation, as mutations in the following genes result in attenuated biofilm formation over cervical cells and/or glass: oxyR, gor, prx, mntABC, trxB, and estD. Overall, biofilm formation appears to be an adaptation for coping with the environmental stresses present in the female genitourinary tract. Therefore, this review will discuss the studies, which describe the composition and metabolic phenotype of gonococcal biofilms.

  14. ACE phenotyping in Gaucher disease.

    Science.gov (United States)

    Danilov, Sergei M; Tikhomirova, Victoria E; Metzger, Roman; Naperova, Irina A; Bukina, Tatiana M; Goker-Alpan, Ozlem; Tayebi, Nahid; Gayfullin, Nurshat M; Schwartz, David E; Samokhodskaya, Larisa M; Kost, Olga A; Sidransky, Ellen

    2018-04-01

    Gaucher disease is characterized by the activation of splenic and hepatic macrophages, accompanied by dramatically increased levels of angiotensin-converting enzyme (ACE). To evaluate the source of the elevated blood ACE, we performed complete ACE phenotyping using blood, spleen and liver samples from patients with Gaucher disease and controls. ACE phenotyping included 1) immunohistochemical staining for ACE; 2) measuring ACE activity with two substrates (HHL and ZPHL); 3) calculating the ratio of the rates of substrate hydrolysis (ZPHL/HHL ratio); 4) assessing the conformational fingerprint of ACE by evaluating the pattern of binding of monoclonal antibodies to 16 different ACE epitopes. We show that in patients with Gaucher disease, the dramatically increased levels of ACE originate from activated splenic and/or hepatic macrophages (Gaucher cells), and that both its conformational fingerprint and kinetic characteristics (ZPHL/HHL ratio) differ from controls and from patients with sarcoid granulomas. Furthermore, normal spleen was found to produce high levels of endogenous ACE inhibitors and a novel, tightly-bound 10-30 kDa ACE effector which is deficient in Gaucher spleen. The conformation of ACE is tissue-specific. In Gaucher disease, ACE produced by activated splenic macrophages differs from that in hepatic macrophages, as well as from macrophages and dendritic cells in sarcoid granulomas. The observed differences are likely due to altered ACE glycosylation or sialylation in these diseased organs. The conformational differences in ACE may serve as a specific biomarker for Gaucher disease. Copyright © 2018 Elsevier Inc. All rights reserved.

  15. Global DNA hypermethylation-associated cancer chemotherapy resistance and its reversion with the demethylating agent hydralazine

    Directory of Open Access Journals (Sweden)

    Benitez-Bribiesca Luis

    2006-08-01

    Full Text Available Abstract Background The development of resistance to cytotoxic chemotherapy continues to be a major obstacle for successful anticancer therapy. It has been shown that cells exposed to toxic concentrations of commonly used cancer chemotherapy agents develop DNA hypermetylation. Hence, demethylating agents could play a role in overcoming drug resistance. Methods MCF-7 cells were rendered adriamycin-resistant by weekly treatment with adriamycin. Wild-type and the resulting MCF-7/Adr cells were analyzed for global DNA methylation. DNA methyltransferase activity and DNA methyltransferase (dnmt gene expression were also determined. MCF-7/Adr cells were then subjected to antisense targeting of dnmt1, -3a, and -b genes and to treatment with the DNA methylation inhibitor hydralazine to investigate whether DNA demethylation restores sensitivity to adriamycin. Results MCF-7/Adr cells exhibited the multi-drug resistant phenotype as demonstrated by adriamycin resistance, mdr1 gene over-expression, decreased intracellular accumulation of adriamycin, and cross-resistance to paclitaxel. The mdr phenotype was accompanied by global DNA hypermetylation, over-expression of dnmt genes, and increased DNA methyltransferase activity as compared with wild-type MCF-7 cells. DNA demethylation through antisense targeting of dnmts or hydralazine restored adriamycin sensitivity of MCF-7/Adr cells to a greater extent than verapamil, a known inhibitor of mdr protein, suggesting that DNA demethylation interferes with the epigenetic reprogramming that participates in the drug-resistant phenotype. Conclusion We provide evidence that DNA hypermethylation is at least partly responsible for development of the multidrug-resistant phenotype in the MCF-7/Adr model and that hydralazine, a known DNA demethylating agent, can revert the resistant phenotype.

  16. Regenerant arabidopsis lineages display a distinct genome-wide spectrum of mutations conferring variant phenotypes

    KAUST Repository

    Jiang, Caifu

    2011-07-28

    Multicellular organisms can be regenerated from totipotent differentiated somatic cell or nuclear founders [1-3]. Organisms regenerated from clonally related isogenic founders might a priori have been expected to be phenotypically invariant. However, clonal regenerant animals display variant phenotypes caused by defective epigenetic reprogramming of gene expression [2], and clonal regenerant plants exhibit poorly understood heritable phenotypic ("somaclonal") variation [4-7]. Here we show that somaclonal variation in regenerant Arabidopsis lineages is associated with genome-wide elevation in DNA sequence mutation rate. We also show that regenerant mutations comprise a distinctive molecular spectrum of base substitutions, insertions, and deletions that probably results from decreased DNA repair fidelity. Finally, we show that while regenerant base substitutions are a likely major genetic cause of the somaclonal variation of regenerant Arabidopsis lineages, transposon movement is unlikely to contribute substantially to that variation. We conclude that the phenotypic variation of regenerant plants, unlike that of regenerant animals, is substantially due to DNA sequence mutation. 2011 Elsevier Ltd. All rights reserved.

  17. Transgenerational effects of stress exposure on offspring phenotypes in apomictic dandelion.

    Directory of Open Access Journals (Sweden)

    Koen J F Verhoeven

    Full Text Available Heritable epigenetic modulation of gene expression is a candidate mechanism to explain parental environmental effects on offspring phenotypes, but current evidence for environment-induced epigenetic changes that persist in offspring generations is scarce. In apomictic dandelions, exposure to various stresses was previously shown to heritably alter DNA methylation patterns. In this study we explore whether these induced changes are accompanied by heritable effects on offspring phenotypes. We observed effects of parental jasmonic acid treatment on offspring specific leaf area and on offspring interaction with a generalist herbivore; and of parental nutrient stress on offspring root-shoot biomass ratio, tissue P-content and leaf morphology. Some of the effects appeared to enhance offspring ability to cope with the same stresses that their parents experienced. Effects differed between apomictic genotypes and were not always consistently observed between different experiments, especially in the case of parental nutrient stress. While this context-dependency of the effects remains to be further clarified, the total set of results provides evidence for the existence of transgenerational effects in apomictic dandelions. Zebularine treatment affected the within-generation response to nutrient stress, pointing at a role of DNA methylation in phenotypic plasticity to nutrient environments. This study shows that stress exposure in apomictic dandelions can cause transgenerational phenotypic effects, in addition to previously demonstrated transgenerational DNA methylation effects.

  18. Transgenerational effects of stress exposure on offspring phenotypes in apomictic dandelion.

    Science.gov (United States)

    Verhoeven, Koen J F; van Gurp, Thomas P

    2012-01-01

    Heritable epigenetic modulation of gene expression is a candidate mechanism to explain parental environmental effects on offspring phenotypes, but current evidence for environment-induced epigenetic changes that persist in offspring generations is scarce. In apomictic dandelions, exposure to various stresses was previously shown to heritably alter DNA methylation patterns. In this study we explore whether these induced changes are accompanied by heritable effects on offspring phenotypes. We observed effects of parental jasmonic acid treatment on offspring specific leaf area and on offspring interaction with a generalist herbivore; and of parental nutrient stress on offspring root-shoot biomass ratio, tissue P-content and leaf morphology. Some of the effects appeared to enhance offspring ability to cope with the same stresses that their parents experienced. Effects differed between apomictic genotypes and were not always consistently observed between different experiments, especially in the case of parental nutrient stress. While this context-dependency of the effects remains to be further clarified, the total set of results provides evidence for the existence of transgenerational effects in apomictic dandelions. Zebularine treatment affected the within-generation response to nutrient stress, pointing at a role of DNA methylation in phenotypic plasticity to nutrient environments. This study shows that stress exposure in apomictic dandelions can cause transgenerational phenotypic effects, in addition to previously demonstrated transgenerational DNA methylation effects.

  19. Regenerant arabidopsis lineages display a distinct genome-wide spectrum of mutations conferring variant phenotypes

    KAUST Repository

    Jiang, Caifu; Mithani, Aziz; Gan, Xiangchao; Belfield, Eric J.; Klingler, John  P.; Zhu, Jian-Kang; Ragoussis, Jiannis; Mott, Richard; Harberd, Nicholas  P.

    2011-01-01

    Multicellular organisms can be regenerated from totipotent differentiated somatic cell or nuclear founders [1-3]. Organisms regenerated from clonally related isogenic founders might a priori have been expected to be phenotypically invariant. However, clonal regenerant animals display variant phenotypes caused by defective epigenetic reprogramming of gene expression [2], and clonal regenerant plants exhibit poorly understood heritable phenotypic ("somaclonal") variation [4-7]. Here we show that somaclonal variation in regenerant Arabidopsis lineages is associated with genome-wide elevation in DNA sequence mutation rate. We also show that regenerant mutations comprise a distinctive molecular spectrum of base substitutions, insertions, and deletions that probably results from decreased DNA repair fidelity. Finally, we show that while regenerant base substitutions are a likely major genetic cause of the somaclonal variation of regenerant Arabidopsis lineages, transposon movement is unlikely to contribute substantially to that variation. We conclude that the phenotypic variation of regenerant plants, unlike that of regenerant animals, is substantially due to DNA sequence mutation. 2011 Elsevier Ltd. All rights reserved.

  20. Transcription factors as readers and effectors of DNA methylation.

    Science.gov (United States)

    Zhu, Heng; Wang, Guohua; Qian, Jiang

    2016-08-01

    Recent technological advances have made it possible to decode DNA methylomes at single-base-pair resolution under various physiological conditions. Many aberrant or differentially methylated sites have been discovered, but the mechanisms by which changes in DNA methylation lead to observed phenotypes, such as cancer, remain elusive. The classical view of methylation-mediated protein-DNA interactions is that only proteins with a methyl-CpG binding domain (MBD) can interact with methylated DNA. However, evidence is emerging to suggest that transcription factors lacking a MBD can also interact with methylated DNA. The identification of these proteins and the elucidation of their characteristics and the biological consequences of methylation-dependent transcription factor-DNA interactions are important stepping stones towards a mechanistic understanding of methylation-mediated biological processes, which have crucial implications for human development and disease.

  1. Viral interference with DNA repair by targeting of the single-stranded DNA binding protein RPA.

    Science.gov (United States)

    Banerjee, Pubali; DeJesus, Rowena; Gjoerup, Ole; Schaffhausen, Brian S

    2013-10-01

    Correct repair of damaged DNA is critical for genomic integrity. Deficiencies in DNA repair are linked with human cancer. Here we report a novel mechanism by which a virus manipulates DNA damage responses. Infection with murine polyomavirus sensitizes cells to DNA damage by UV and etoposide. Polyomavirus large T antigen (LT) alone is sufficient to sensitize cells 100 fold to UV and other kinds of DNA damage. This results in activated stress responses and apoptosis. Genetic analysis shows that LT sensitizes via the binding of its origin-binding domain (OBD) to the single-stranded DNA binding protein replication protein A (RPA). Overexpression of RPA protects cells expressing OBD from damage, and knockdown of RPA mimics the LT phenotype. LT prevents recruitment of RPA to nuclear foci after DNA damage. This leads to failure to recruit repair proteins such as Rad51 or Rad9, explaining why LT prevents repair of double strand DNA breaks by homologous recombination. A targeted intervention directed at RPA based on this viral mechanism could be useful in circumventing the resistance of cancer cells to therapy.

  2. Breast tumor copy number aberration phenotypes and genomic instability

    International Nuclear Information System (INIS)

    Fridlyand, Jane; Jain, Ajay N; McLennan, Jane; Ziegler, John; Chin, Koei; Devries, Sandy; Feiler, Heidi; Gray, Joe W; Waldman, Frederic; Pinkel, Daniel; Albertson, Donna G; Snijders, Antoine M; Ylstra, Bauke; Li, Hua; Olshen, Adam; Segraves, Richard; Dairkee, Shanaz; Tokuyasu, Taku; Ljung, Britt Marie

    2006-01-01

    Genomic DNA copy number aberrations are frequent in solid tumors, although the underlying causes of chromosomal instability in tumors remain obscure. Genes likely to have genomic instability phenotypes when mutated (e.g. those involved in mitosis, replication, repair, and telomeres) are rarely mutated in chromosomally unstable sporadic tumors, even though such mutations are associated with some heritable cancer prone syndromes. We applied array comparative genomic hybridization (CGH) to the analysis of breast tumors. The variation in the levels of genomic instability amongst tumors prompted us to investigate whether alterations in processes/genes involved in maintenance and/or manipulation of the genome were associated with particular types of genomic instability. We discriminated three breast tumor subtypes based on genomic DNA copy number alterations. The subtypes varied with respect to level of genomic instability. We find that shorter telomeres and altered telomere related gene expression are associated with amplification, implicating telomere attrition as a promoter of this type of aberration in breast cancer. On the other hand, the numbers of chromosomal alterations, particularly low level changes, are associated with altered expression of genes in other functional classes (mitosis, cell cycle, DNA replication and repair). Further, although loss of function instability phenotypes have been demonstrated for many of the genes in model systems, we observed enhanced expression of most genes in tumors, indicating that over expression, rather than deficiency underlies instability. Many of the genes associated with higher frequency of copy number aberrations are direct targets of E2F, supporting the hypothesis that deregulation of the Rb pathway is a major contributor to chromosomal instability in breast tumors. These observations are consistent with failure to find mutations in sporadic tumors in genes that have roles in maintenance or manipulation of the genome

  3. DNA Repair Systems

    Indian Academy of Sciences (India)

    DNA molecule which makes it ideal for storage and propagation of genetic information. ... of these errors are broadly referred to as DNA repair. DNA can ... changes occur in the human genome per day. ..... nails, frequent physical and mental.

  4. Synthesis of DNA

    Science.gov (United States)

    Mariella, Jr., Raymond P.

    2008-11-18

    A method of synthesizing a desired double-stranded DNA of a predetermined length and of a predetermined sequence. Preselected sequence segments that will complete the desired double-stranded DNA are determined. Preselected segment sequences of DNA that will be used to complete the desired double-stranded DNA are provided. The preselected segment sequences of DNA are assembled to produce the desired double-stranded DNA.

  5. DNA watermarks: A proof of concept

    Directory of Open Access Journals (Sweden)

    Barnekow Angelika

    2008-04-01

    Full Text Available Abstract Background DNA-based watermarks are helpful tools to identify the unauthorized use of genetically modified organisms (GMOs protected by patents. In silico analyses showed that in coding regions synonymous codons can be used to insert encrypted information into the genome of living organisms by using the DNA-Crypt algorithm. Results We integrated an authenticating watermark in the Vam7 sequence. For our investigations we used a mutant Saccharomyces cerevisiae strain, called CG783, which has an amber mutation within the Vam7 sequence. The CG783 cells are unable to sporulate and in addition display an abnormal vacuolar morphology. Transformation of CG783 with pRS314 Vam7 leads to a phenotype very similar to the wildtype yeast strain CG781. The integrated watermark did not influence the function of Vam7 and the resulting phenotype of the CG783 cells transformed with pRS314 Vam7-TB shows no significant differences compared to the CG783 cells transformed with pRS314 Vam7. Conclusion From our experiments we conclude that the DNA watermarks produced by DNA-Crypt do not influence the translation from mRNA into protein. By analyzing the vacuolar morphology, growth rate and ability to sporulate we confirmed that the resulting Vam7 protein was functionally active.

  6. The Mitochondrial DNA (mtDNA)-Associated Protein SWIB5 Influences mtDNA Architecture and Homologous Recombination

    KAUST Repository

    Blomme, Jonas

    2017-04-19

    In addition to the nucleus, mitochondria and chloroplasts in plant cells also contain genomes. Efficient DNA repair pathways are crucial in these organelles to fix damage resulting from endogenous and exogenous factors. Plant organellar genomes are complex compared with their animal counterparts, and although several plant-specific mediators of organelle DNA repair have been reported, many regulators remain to be identified. Here, we show that a mitochondrial SWI/SNF (nucleosome remodeling) complex B protein, SWIB5, is capable of associating with mitochondrial DNA (mtDNA) in Arabidopsis thaliana. Gainand loss-of-function mutants provided evidence for a role of SWIB5 in influencing mtDNA architecture and homologous recombination at specific intermediate-sized repeats both under normal and genotoxic conditions. SWIB5 interacts with other mitochondrial SWIB proteins. Gene expression and mutant phenotypic analysis of SWIB5 and SWIB family members suggests a link between organellar genome maintenance and cell proliferation. Taken together, our work presents a protein family that influences mtDNA architecture and homologous recombination in plants and suggests a link between organelle functioning and plant development.

  7. Refined Phenotyping of Modic Changes

    Science.gov (United States)

    Määttä, Juhani H.; Karppinen, Jaro; Paananen, Markus; Bow, Cora; Luk, Keith D.K.; Cheung, Kenneth M.C.; Samartzis, Dino

    2016-01-01

    Abstract Low back pain (LBP) is the world's most disabling condition. Modic changes (MC) are vertebral bone marrow changes adjacent to the endplates as noted on magnetic resonance imaging. The associations of specific MC types and patterns with prolonged, severe LBP and disability remain speculative. This study assessed the relationship of prolonged, severe LBP and back-related disability, with the presence and morphology of lumbar MC in a large cross-sectional population-based study of Southern Chinese. We addressed the topographical and morphological dimensions of MC along with other magnetic resonance imaging phenotypes (eg, disc degeneration and displacement) on the basis of axial T1 and sagittal T2-weighted imaging of L1-S1. Prolonged severe LBP was defined as LBP lasting ≥30 days during the past year, and a visual analog scale severest pain intensity of at least 6/10. An Oswestry Disability Index score of 15% was regarded as significant disability. We also assessed subject demographics, occupation, and lifestyle factors. In total, 1142 subjects (63% females, mean age 53 years) were assessed. Of these, 282 (24.7%) had MC (7.1% type I, 17.6% type II). MC subjects were older (P = 0.003), had more frequent disc displacements (P disability. The strength of the associations increased with the number of MC. This large-scale study is the first to definitively note MC types and specific morphologies to be independently associated with prolonged severe LBP and back-related disability. This proposed refined MC phenotype may have direct implications in clinical decision-making as to the development and management of LBP. Understanding of these imaging biomarkers can lead to new preventative and personalized therapeutics related to LBP. PMID:27258491

  8. Utility of 16S rDNA Sequencing for Identification of Rare Pathogenic Bacteria.

    Science.gov (United States)

    Loong, Shih Keng; Khor, Chee Sieng; Jafar, Faizatul Lela; AbuBakar, Sazaly

    2016-11-01

    Phenotypic identification systems are established methods for laboratory identification of bacteria causing human infections. Here, the utility of phenotypic identification systems was compared against 16S rDNA identification method on clinical isolates obtained during a 5-year study period, with special emphasis on isolates that gave unsatisfactory identification. One hundred and eighty-seven clinical bacteria isolates were tested with commercial phenotypic identification systems and 16S rDNA sequencing. Isolate identities determined using phenotypic identification systems and 16S rDNA sequencing were compared for similarity at genus and species level, with 16S rDNA sequencing as the reference method. Phenotypic identification systems identified ~46% (86/187) of the isolates with identity similar to that identified using 16S rDNA sequencing. Approximately 39% (73/187) and ~15% (28/187) of the isolates showed different genus identity and could not be identified using the phenotypic identification systems, respectively. Both methods succeeded in determining the species identities of 55 isolates; however, only ~69% (38/55) of the isolates matched at species level. 16S rDNA sequencing could not determine the species of ~20% (37/187) of the isolates. The 16S rDNA sequencing is a useful method over the phenotypic identification systems for the identification of rare and difficult to identify bacteria species. The 16S rDNA sequencing method, however, does have limitation for species-level identification of some bacteria highlighting the need for better bacterial pathogen identification tools. © 2016 Wiley Periodicals, Inc.

  9. Mitochondrial DNA mutations in mutator mice confer respiration defects and B-cell lymphoma development.

    Directory of Open Access Journals (Sweden)

    Takayuki Mito

    Full Text Available Mitochondrial DNA (mtDNA mutator mice are proposed to express premature aging phenotypes including kyphosis and hair loss (alopecia due to their carrying a nuclear-encoded mtDNA polymerase with a defective proofreading function, which causes accelerated accumulation of random mutations in mtDNA, resulting in expression of respiration defects. On the contrary, transmitochondrial mito-miceΔ carrying mtDNA with a large-scale deletion mutation (ΔmtDNA also express respiration defects, but not express premature aging phenotypes. Here, we resolved this discrepancy by generating mtDNA mutator mice sharing the same C57BL/6J (B6J nuclear background with that of mito-miceΔ. Expression patterns of premature aging phenotypes are very close, when we compared between homozygous mtDNA mutator mice carrying a B6J nuclear background and selected mito-miceΔ only carrying predominant amounts of ΔmtDNA, in their expression of significant respiration defects, kyphosis, and a short lifespan, but not the alopecia. Therefore, the apparent discrepancy in the presence and absence of premature aging phenotypes in mtDNA mutator mice and mito-miceΔ, respectively, is partly the result of differences in the nuclear background of mtDNA mutator mice and of the broad range of ΔmtDNA proportions of mito-miceΔ used in previous studies. We also provided direct evidence that mtDNA abnormalities in homozygous mtDNA mutator mice are responsible for respiration defects by demonstrating the co-transfer of mtDNA and respiration defects from mtDNA mutator mice into mtDNA-less (ρ(0 mouse cells. Moreover, heterozygous mtDNA mutator mice had a normal lifespan, but frequently developed B-cell lymphoma, suggesting that the mtDNA abnormalities in heterozygous mutator mice are not sufficient to induce a short lifespan and aging phenotypes, but are able to contribute to the B-cell lymphoma development during their prolonged lifespan.

  10. The phenotypic variance gradient - a novel concept.

    Science.gov (United States)

    Pertoldi, Cino; Bundgaard, Jørgen; Loeschcke, Volker; Barker, James Stuart Flinton

    2014-11-01

    Evolutionary ecologists commonly use reaction norms, which show the range of phenotypes produced by a set of genotypes exposed to different environments, to quantify the degree of phenotypic variance and the magnitude of plasticity of morphometric and life-history traits. Significant differences among the values of the slopes of the reaction norms are interpreted as significant differences in phenotypic plasticity, whereas significant differences among phenotypic variances (variance or coefficient of variation) are interpreted as differences in the degree of developmental instability or canalization. We highlight some potential problems with this approach to quantifying phenotypic variance and suggest a novel and more informative way to plot reaction norms: namely "a plot of log (variance) on the y-axis versus log (mean) on the x-axis, with a reference line added". This approach gives an immediate impression of how the degree of phenotypic variance varies across an environmental gradient, taking into account the consequences of the scaling effect of the variance with the mean. The evolutionary implications of the variation in the degree of phenotypic variance, which we call a "phenotypic variance gradient", are discussed together with its potential interactions with variation in the degree of phenotypic plasticity and canalization.

  11. Stress-induced DNA methylation changes and their heritability in asexual dandelions

    NARCIS (Netherlands)

    Verhoeven, K.J.F.; Jansen, J.J.; Van Dijk, P.J.; Biere, A.

    2010-01-01

    DNA methylation can cause heritable phenotypic modifications in the absence of changes in DNA sequence. Environmental stresses can trigger methylation changes and this may have evolutionary consequences, even in the absence of sequence variation. However, it remains largely unknown to what extent

  12. Stress-induced DNA methylation changes and their heritability in asexual dandelions

    NARCIS (Netherlands)

    Verhoeven, K.J.F.; Jansen, J.J.; Dijk, P.J.; Biere, A.

    2010-01-01

    DNA methylation can cause heritable phenotypic modifications in the absence of changes in DNA sequence. Environmental stresses can trigger methylation changes and this may have evolutionary consequences, even in the absence of sequence variation. However, it remains largely unknown to what extent

  13. DNA based methods used for characterization and detection of food ...

    African Journals Online (AJOL)

    Detection of food borne pathogen is of outmost importance in the food industries and related agencies. For the last few decades conventional methods were used to detect food borne pathogens based on phenotypic characters. At the advent of complementary base pairing and amplification of DNA, the diagnosis of food ...

  14. Virulence properties and random amplification of polymorphic DNA ...

    African Journals Online (AJOL)

    Genotypic and phenotypic characterization as well as studies on the virulence factors of Candida albicans isolates obtained from oral cavity of patients was carried out using random amplified polymorphic DNA (RAPD) fingerprinting and epithelial cells adherence assay, respectively. RAPD patterns revealed the presence of ...

  15. Exome sequencing for gene discovery in lethal fetal disorders--harnessing the value of extreme phenotypes.

    Science.gov (United States)

    Filges, Isabel; Friedman, Jan M

    2015-10-01

    Massively parallel sequencing has revolutionized our understanding of Mendelian disorders, and many novel genes have been discovered to cause disease phenotypes when mutant. At the same time, next-generation sequencing approaches have enabled non-invasive prenatal testing of free fetal DNA in maternal blood. However, little attention has been paid to using whole exome and genome sequencing strategies for gene identification in fetal disorders that are lethal in utero, because they can appear to be sporadic and Mendelian inheritance may be missed. We present challenges and advantages of applying next-generation sequencing approaches to gene discovery in fetal malformation phenotypes and review recent successful discovery approaches. We discuss the implication and significance of recessive inheritance and cross-species phenotyping in fetal lethal conditions. Whole exome sequencing can be used in individual families with undiagnosed lethal congenital anomaly syndromes to discover causal mutations, provided that prior to data analysis, the fetal phenotype can be correlated to a particular developmental pathway in embryogenesis. Cross-species phenotyping allows providing further evidence for causality of discovered variants in genes involved in those extremely rare phenotypes and will increase our knowledge about normal and abnormal human developmental processes. Ultimately, families will benefit from the option of early prenatal diagnosis. © 2014 John Wiley & Sons, Ltd.

  16. DNA damage preceding dopamine neuron degeneration in A53T human α-synuclein transgenic mice.

    Science.gov (United States)

    Wang, Degui; Yu, Tianyu; Liu, Yongqiang; Yan, Jun; Guo, Yingli; Jing, Yuhong; Yang, Xuguang; Song, Yanfeng; Tian, Yingxia

    2016-12-02

    Defective DNA repair has been linked with age-associated neurodegenerative disorders. Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by genetic and environmental factors. Whether damages to nuclear DNA contribute to neurodegeneration of PD still remain obscure. in this study we aim to explore whether nuclear DNA damage induce dopamine neuron degeneration in A53T human α-Synuclein over expressed mouse model. We investigated the effects of X-ray irradiation on A53T-α-Syn MEFs and A53T-α-Syn transgene mice. Our results indicate that A53T-α-Syn MEFs show a prolonged DNA damage repair process and senescense phenotype. DNA damage preceded onset of motor phenotype in A53T-α-Syn transgenic mice and decrease the number of nigrostriatal dopaminergic neurons. Neurons of A53T-α-Syn transgenic mice are more fragile to DNA damages. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. The Human Phenotype Ontology project: linking molecular biology and disease through phenotype data

    NARCIS (Netherlands)

    Kohler, S.; Doelken, S.C.; Mungall, C.J.; Bauer, S.; Firth, H.V.; Bailleul-Forestier, I.; Black, G.C.M.; Brown, D.L.; Brudno, M.; Campbell, J.; FitzPatrick, D.R.; Eppig, J.T.; Jackson, A.P.; Freson, K.; Girdea, M.; Helbig, I.; Hurst, J.A.; Jahn, J.; Jackson, L.G.; Kelly, A.M.; Ledbetter, D.H.; Mansour, S.; Martin, C.L.; Moss, C.; Mumford, A.; Ouwehand, W.H.; Park, S.M.; Riggs, E.R.; Scott, R.H.; Sisodiya, S.; Vooren, S. van der; Wapner, R.J.; Wilkie, A.O.; Wright, C.F.; Silfhout, A.T. van; Leeuw, N. de; Vries, B. de; Washingthon, N.L.; Smith, C.L.; Westerfield, M.; Schofield, P.; Ruef, B.J.; Gkoutos, G.V.; Haendel, M.; Smedley, D.; Lewis, S.E.; Robinson, P.N.

    2014-01-01

    The Human Phenotype Ontology (HPO) project, available at http://www.human-phenotype-ontology.org, provides a structured, comprehensive and well-defined set of 10,088 classes (terms) describing human phenotypic abnormalities and 13,326 subclass relations between the HPO classes. In addition we have

  18. The yeast Saccharomyces cerevisiae DNA polymerase IV: possible involvement in double strand break DNA repair.

    Science.gov (United States)

    Leem, S H; Ropp, P A; Sugino, A

    1994-08-11

    We identified and purified a new DNA polymerase (DNA polymerase IV), which is similar to mammalian DNA polymerase beta, from Saccharomyces cerevisiae and suggested that it is encoded by YCR14C (POLX) on chromosome III. Here, we provided a direct evidence that the purified DNA polymerase IV is indeed encoded by POLX. Strains harboring a pol4 deletion mutation exhibit neither mitotic growth defect nor a meiosis defect, suggesting that DNA polymerase IV participates in nonessential functions in DNA metabolism. The deletion strains did not exhibit UV-sensitivity. However, they did show weak sensitivity to MMS-treatment and exhibited a hyper-recombination phenotype when intragenic recombination was measured during meiosis. Furthermore, MAT alpha pol4 delta segregants had a higher frequency of illegitimate mating with a MAT alpha tester strain than that of wild-type cells. These results suggest that DNA polymerase IV participates in a double-strand break repair pathway. A 3.2kb of the POL4 transcript was weakly expressed in mitotically growing cells. During meiosis, a 2.2 kb POL4 transcript was greatly induced, while the 3.2 kb transcript stayed at constant levels. This induction was delayed in a swi4 delta strain during meiosis, while no effect was observed in a swi6 delta strain.

  19. Daddy issues: paternal effects on phenotype.

    Science.gov (United States)

    Rando, Oliver J

    2012-11-09

    The once popular and then heretical idea that ancestral environment can affect the phenotype of future generations is coming back into vogue due to advances in the field of epigenetic inheritance. How paternal environmental conditions influence the phenotype of progeny is now a tractable question, and researchers are exploring potential mechanisms underlying such effects. Copyright © 2012 Elsevier Inc. All rights reserved.

  20. Phenotypes of organ involvement in sarcoidosis

    NARCIS (Netherlands)

    Schupp, Jonas Christian; Freitag-Wolf, Sandra; Bargagli, Elena; Mihailović-Vučinić, Violeta; Rottoli, Paola; Grubanovic, Aleksandar; Müller, Annegret; Jochens, Arne; Tittmann, Lukas; Schnerch, Jasmin; Olivieri, Carmela; Fischer, Annegret; Jovanovic, Dragana; Filipovic, Snežana; Videnovic-Ivanovic, Jelica; Bresser, Paul; Jonkers, René; O'Reilly, Kate; Ho, Ling-Pei; Gaede, Karoline I.; Zabel, Peter; Dubaniewicz, Anna; Marshall, Ben; Kieszko, Robert; Milanowski, Janusz; Günther, Andreas; Weihrich, Anette; Petrek, Martin; Kolek, Vitezslav; Keane, Michael P.; O'Beirne, Sarah; Donnelly, Seamas; Haraldsdottir, Sigridur Olina; Jorundsdottir, Kristin B.; Costabel, Ulrich; Bonella, Francesco; Wallaert, Benoît; Grah, Christian; Peroš-Golubičić, Tatjana; Luisetti, Mauritio; Kadija, Zamir; Pabst, Stefan; Grohé, Christian; Strausz, János; Vašáková, Martina; Sterclova, Martina; Millar, Ann; Homolka, Jiří; Slováková, Alena; Kendrick, Yvonne; Crawshaw, Anjali; Wuyts, Wim; Spencer, Lisa; Pfeifer, Michael; Valeyre, Dominique; Poletti, Venerino; Wirtz, Hubertus; Prasse, Antje; Schreiber, Stefan; Krawczak, Michael; Müller-Quernheim, Joachim

    2018-01-01

    Sarcoidosis is a highly variable, systemic granulomatous disease of hitherto unknown aetiology. The GenPhenReSa (Genotype-Phenotype Relationship in Sarcoidosis) project represents a European multicentre study to investigate the influence of genotype on disease phenotypes in sarcoidosis. The baseline

  1. Emerging semantics to link phenotype and environment

    Directory of Open Access Journals (Sweden)

    Anne E. Thessen

    2015-12-01

    Full Text Available Understanding the interplay between environmental conditions and phenotypes is a fundamental goal of biology. Unfortunately, data that include observations on phenotype and environment are highly heterogeneous and thus difficult to find and integrate. One approach that is likely to improve the status quo involves the use of ontologies to standardize and link data about phenotypes and environments. Specifying and linking data through ontologies will allow researchers to increase the scope and flexibility of large-scale analyses aided by modern computing methods. Investments in this area would advance diverse fields such as ecology, phylogenetics, and conservation biology. While several biological ontologies are well-developed, using them to link phenotypes and environments is rare because of gaps in ontological coverage and limits to interoperability among ontologies and disciplines. In this manuscript, we present (1 use cases from diverse disciplines to illustrate questions that could be answered more efficiently using a robust linkage between phenotypes and environments, (2 two proof-of-concept analyses that show the value of linking phenotypes to environments in fishes and amphibians, and (3 two proposed example data models for linking phenotypes and environments using the extensible observation ontology (OBOE and the Biological Collections Ontology (BCO; these provide a starting point for the development of a data model linking phenotypes and environments.

  2. Haptoglobin Phenotypes and Hypertension in Indigenous Zambians ...

    African Journals Online (AJOL)

    Haptoglobin Phenotypes and Hypertension in Indigenous Zambians at the University Teaching Hospital, Lusaka, Zambia. MM Phiri, T Kaile, FM Goma. Abstract. Objectives: The aim of the study was to investigate the association between presence of haptoglobin phenotypes and hypertension in indigenous Zambian patients ...

  3. Knowledge-based analysis of phenotypes

    KAUST Repository

    Hoendorf, Robert

    2016-01-01

    a formal framework to describe phenotypes. A formalized theory of phenotypes is not only useful for domain analysis, but can also be applied to assist in the diagnosis of rare genetic diseases, and I will show how our results on the ontology

  4. The Neuroanatomy of the Autistic Phenotype

    Science.gov (United States)

    Fahim, Cherine; Meguid, Nagwa A.; Nashaat, Neveen H.; Yoon, Uicheul; Mancini-Marie, Adham; Evans, Alan C.

    2012-01-01

    The autism phenotype is associated with an excess of brain volume due in part to decreased pruning during development. Here we aimed at assessing brain volume early in development to further elucidate previous findings in autism and determine whether this pattern is restricted to idiopathic autism or shared within the autistic phenotype (fragile X…

  5. Cooperation of DNA-PKcs and WRN helicase in the maintenance of telomeric D-loops

    DEFF Research Database (Denmark)

    Kusumoto-Matsuo, Rika; Opresko, Patricia L; Ramsden, Dale

    2010-01-01

    Werner syndrome is an inherited human progeriod syndrome caused by mutations in the gene encoding the Werner Syndrome protein, WRN. It has both 3'-5' DNA helicase and exonuclease activities, and is suggested to have roles in many aspects of DNA metabolism, including DNA repair and telomere...... D-loop model substrate. In addition, the length of telomeric G-tails decreases in DNA-PKcs knockdown cells, and this phenotype is reversed by overexpression of WRN helicase. These results suggest that WRN and DNA-PKcs may cooperatively prevent G-tail shortening in vivo....

  6. Evolving phenotypic networks in silico.

    Science.gov (United States)

    François, Paul

    2014-11-01

    Evolved gene networks are constrained by natural selection. Their structures and functions are consequently far from being random, as exemplified by the multiple instances of parallel/convergent evolution. One can thus ask if features of actual gene networks can be recovered from evolutionary first principles. I review a method for in silico evolution of small models of gene networks aiming at performing predefined biological functions. I summarize the current implementation of the algorithm, insisting on the construction of a proper "fitness" function. I illustrate the approach on three examples: biochemical adaptation, ligand discrimination and vertebrate segmentation (somitogenesis). While the structure of the evolved networks is variable, dynamics of our evolved networks are usually constrained and present many similar features to actual gene networks, including properties that were not explicitly selected for. In silico evolution can thus be used to predict biological behaviours without a detailed knowledge of the mapping between genotype and phenotype. Copyright © 2014 The Author. Published by Elsevier Ltd.. All rights reserved.

  7. Federated Tensor Factorization for Computational Phenotyping

    Science.gov (United States)

    Kim, Yejin; Sun, Jimeng; Yu, Hwanjo; Jiang, Xiaoqian

    2017-01-01

    Tensor factorization models offer an effective approach to convert massive electronic health records into meaningful clinical concepts (phenotypes) for data analysis. These models need a large amount of diverse samples to avoid population bias. An open challenge is how to derive phenotypes jointly across multiple hospitals, in which direct patient-level data sharing is not possible (e.g., due to institutional policies). In this paper, we developed a novel solution to enable federated tensor factorization for computational phenotyping without sharing patient-level data. We developed secure data harmonization and federated computation procedures based on alternating direction method of multipliers (ADMM). Using this method, the multiple hospitals iteratively update tensors and transfer secure summarized information to a central server, and the server aggregates the information to generate phenotypes. We demonstrated with real medical datasets that our method resembles the centralized training model (based on combined datasets) in terms of accuracy and phenotypes discovery while respecting privacy. PMID:29071165

  8. Nordic research infrastructures for plant phenotyping

    Directory of Open Access Journals (Sweden)

    Kristiina Himanen

    2018-03-01

    Full Text Available Plant phenomics refers to the systematic study of plant phenotypes. Together with closely monitored, controlled climates, it provides an essential component for the integrated analysis of genotype-phenotype-environment interactions. Currently, several plant growth and phenotyping facilities are under establishment globally, and numerous facilities are already in use. Alongside the development of the research infrastructures, several national and international networks have been established to support shared use of the new methodology. In this review, an overview is given of the Nordic plant phenotyping and climate control facilities. Since many areas of phenomics such as sensor-based phenotyping, image analysis and data standards are still developing, promotion of educational and networking activities is especially important. These facilities and networks will be instrumental in tackling plant breeding and plant protection challenges. They will also provide possibilities to study wild species and their ecological interactions under changing Nordic climate conditions.

  9. Mutagenesis and repair of DNA

    International Nuclear Information System (INIS)

    Janion, C.; Grzesiuk, E.; Fabisiewicz, A.; Tudek, B.; Ciesla, J.; Graziewicz, M.; Wojcik, A.; Speina, E.

    1998-01-01

    Full text. The discovery that the mfd gene codes for a transcription-coupling repair factor (TRCF) prompted us to re-investigate the MFD (mutation frequency decline) phenomenon in E.coli K-12 strain when mutations were induced by ultraviolet light, halogen light or MMS-treatment. These studies revealed that: (i) the process of MFD involves the proofreading activity of DNA pol III and the mismatch repair system, as well as, TRCF and the UvrABC-excinuclease (ii) a semi-rich plate test may be replaced by a rich liquid medium, (iii) the T-T pyrimidine dimers are the lesions excised with the highest activity, and (iv) overproduction of UmuD(D'C) proteins leads to a great increase in mutant frequency in irradiated and MMS-treated cells. The role of mismatch repair (MR) in MMS-induced mutagenesis is obscured by the fact that the spectra of mutational specificity are different in bacteria proficient and deficient in MR. It has been found that transposons Tn10 (and Tn5) when inserted into chromosomal DNA of E. coli influence the phenotype lowering the survival and frequency of mutations induced by UV or halogen light irradiation. This is connected with a deficiency of UmuD(D') and UmuC proteins. Transformation of bacteria with plasmids bearing the umuD(D')C genes, suppresses the effects of the transposon insertion, a phenomenon which has not been described before. Single-stranded DNA of M13mp18 phage was oxidized in vitro by a hydroxyl radical generating system including hypoxanthine/xanthine oxidase/Fe3+/EDTA, and it was found that Fapy-Ade, Fapy-Gua, 8-oxyAde and thymine glycol were the main products formed. Replication of the oxidized template by T7 phage DNA polymerase, Klenow fragment of polymerase I, or polymerase beta from bovine thymus has revealed that oxidized pyrimidines are stronger blockers than oxidized purines for T7 phage and Klenow fragment polymerases and the blocking potency depends on the neighboring bases and on the type of polymerase. Studies of

  10. DNA methyltransferase 1 mutations and mitochondrial pathology: is mtDNA methylated?

    Directory of Open Access Journals (Sweden)

    Alessandra eMaresca

    2015-03-01

    Full Text Available Autosomal dominant cerebellar ataxia, deafness and narcolepsy (ADCA-DN and Hereditary sensory neuropathy with dementia and hearing loss (HSN1E are two rare, overlapping neurodegenerative syndromes that have been recently linked to allelic dominant pathogenic mutations in the DNMT1 gene, coding for DNA (cytosine-5-methyltransferase 1. DNMT1 is the enzyme responsible for maintaining the nuclear genome methylation patterns during the DNA replication and repair, thus regulating gene expression. The mutations responsible for ADCA-DN and HSN1E affect the replication foci targeting sequence domain, which regulates DNMT1 binding to chromatin. DNMT1 dysfunction is anticipated to lead to a global alteration of the DNA methylation pattern with predictable downstream consequences on gene expression. Interestingly, ADCA-DN and HSN1E phenotypes share some clinical features typical of mitochondrial diseases, such as optic atrophy, peripheral neuropathy and deafness, and some biochemical evidence of mitochondrial dysfunction. The recent discovery of a mitochondrial isoform of DNMT1 and its proposed role in methylating mitochondrial DNA (mtDNA suggests that DNMT1 mutations may directly affect mtDNA and mitochondrial physiology. On the basis of this latter finding the link between DNMT1 abnormal activity and mitochondrial dysfunction in ADCA-DN and HSN1E appears intuitive, however mtDNA methylation remains highly debated. In the last years several groups demonstrated the presence of 5-methylcytosine in mtDNA by different approaches, but, on the other end, the opposite evidence that mtDNA is not methylated has also been published. Since over 1500 mitochondrial proteins are encoded by the nuclear genome, the altered methylation of these genes may well have a critical role in leading to the mitochondrial impairment observed in ADCA-DN and HSN1E. Thus, many open questions still remain unanswered, such as why mtDNA should be methylated, and how this process is

  11. DNA methylation patterns in cord blood DNA and body size in childhood.

    Directory of Open Access Journals (Sweden)

    Caroline L Relton

    Full Text Available Epigenetic markings acquired in early life may have phenotypic consequences later in development through their role in transcriptional regulation with relevance to the developmental origins of diseases including obesity. The goal of this study was to investigate whether DNA methylation levels at birth are associated with body size later in childhood.A study design involving two birth cohorts was used to conduct transcription profiling followed by DNA methylation analysis in peripheral blood. Gene expression analysis was undertaken in 24 individuals whose biological samples and clinical data were collected at a mean ± standard deviation (SD age of 12.35 (0.95 years, the upper and lower tertiles of body mass index (BMI were compared with a mean (SD BMI difference of 9.86 (2.37 kg/m(2. This generated a panel of differentially expressed genes for DNA methylation analysis which was then undertaken in cord blood DNA in 178 individuals with body composition data prospectively collected at a mean (SD age of 9.83 (0.23 years. Twenty-nine differentially expressed genes (>1.2-fold and p<10(-4 were analysed to determine DNA methylation levels at 1-3 sites per gene. Five genes were unmethylated and DNA methylation in the remaining 24 genes was analysed using linear regression with bootstrapping. Methylation in 9 of the 24 (37.5% genes studied was associated with at least one index of body composition (BMI, fat mass, lean mass, height at age 9 years, although only one of these associations remained after correction for multiple testing (ALPL with height, p(Corrected = 0.017.DNA methylation patterns in cord blood show some association with altered gene expression, body size and composition in childhood. The observed relationship is correlative and despite suggestion of a mechanistic epigenetic link between in utero life and later phenotype, further investigation is required to establish causality.

  12. C-terminal phenylalanine of bacteriophage T7 single-stranded DNA-binding protein is essential for strand displacement synthesis by T7 DNA polymerase at a nick in DNA.

    Science.gov (United States)

    Ghosh, Sharmistha; Marintcheva, Boriana; Takahashi, Masateru; Richardson, Charles C

    2009-10-30

    Single-stranded DNA-binding protein (gp2.5), encoded by gene 2.5 of bacteriophage T7, plays an essential role in DNA replication. Not only does it remove impediments of secondary structure in the DNA, it also modulates the activities of the other replication proteins. The acidic C-terminal tail of gp2.5, bearing a C-terminal phenylalanine, physically and functionally interacts with the helicase and DNA polymerase. Deletion of the phenylalanine or substitution with a nonaromatic amino acid gives rise to a dominant lethal phenotype, and the altered gp2.5 has reduced affinity for T7 DNA polymerase. Suppressors of the dominant lethal phenotype have led to the identification of mutations in gene 5 that encodes the T7 DNA polymerase. The altered residues in the polymerase are solvent-exposed and lie in regions that are adjacent to the bound DNA. gp2.5 lacking the C-terminal phenylalanine has a lower affinity for gp5-thioredoxin relative to the wild-type gp2.5, and this affinity is partially restored by the suppressor mutations in DNA polymerase. gp2.5 enables T7 DNA polymerase to catalyze strand displacement DNA synthesis at a nick in DNA. The resulting 5'-single-stranded DNA tail provides a loading site for T7 DNA helicase. gp2.5 lacking the C-terminal phenylalanine does not support this event with wild-type DNA polymerase but does to a limited extent with T7 DNA polymerase harboring the suppressor mutations.

  13. C-terminal Phenylalanine of Bacteriophage T7 Single-stranded DNA-binding Protein Is Essential for Strand Displacement Synthesis by T7 DNA Polymerase at a Nick in DNA*

    Science.gov (United States)

    Ghosh, Sharmistha; Marintcheva, Boriana; Takahashi, Masateru; Richardson, Charles C.

    2009-01-01

    Single-stranded DNA-binding protein (gp2.5), encoded by gene 2.5 of bacteriophage T7, plays an essential role in DNA replication. Not only does it remove impediments of secondary structure in the DNA, it also modulates the activities of the other replication proteins. The acidic C-terminal tail of gp2.5, bearing a C-terminal phenylalanine, physically and functionally interacts with the helicase and DNA polymerase. Deletion of the phenylalanine or substitution with a nonaromatic amino acid gives rise to a dominant lethal phenotype, and the altered gp2.5 has reduced affinity for T7 DNA polymerase. Suppressors of the dominant lethal phenotype have led to the identification of mutations in gene 5 that encodes the T7 DNA polymerase. The altered residues in the polymerase are solvent-exposed and lie in regions that are adjacent to the bound DNA. gp2.5 lacking the C-terminal phenylalanine has a lower affinity for gp5-thioredoxin relative to the wild-type gp2.5, and this affinity is partially restored by the suppressor mutations in DNA polymerase. gp2.5 enables T7 DNA polymerase to catalyze strand displacement DNA synthesis at a nick in DNA. The resulting 5′-single-stranded DNA tail provides a loading site for T7 DNA helicase. gp2.5 lacking the C-terminal phenylalanine does not support this event with wild-type DNA polymerase but does to a limited extent with T7 DNA polymerase harboring the suppressor mutations. PMID:19726688

  14. The Nature of Stable Insomnia Phenotypes

    Science.gov (United States)

    Pillai, Vivek; Roth, Thomas; Drake, Christopher L.

    2015-01-01

    Study Objectives: We examined the 1-y stability of four insomnia symptom profiles: sleep onset insomnia; sleep maintenance insomnia; combined onset and maintenance insomnia; and neither criterion (i.e., insomnia cases that do not meet quantitative thresholds for onset or maintenance problems). Insomnia cases that exhibited the same symptom profile over a 1-y period were considered to be phenotypes, and were compared in terms of clinical and demographic characteristics. Design: Longitudinal. Setting: Urban, community-based. Participants: Nine hundred fifty-four adults with Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition based current insomnia (46.6 ± 12.6 y; 69.4% female). Interventions: None. Measurements and results: At baseline, participants were divided into four symptom profile groups based on quantitative criteria. Follow-up assessment 1 y later revealed that approximately 60% of participants retained the same symptom profile, and were hence judged to be phenotypes. Stability varied significantly by phenotype, such that sleep onset insomnia (SOI) was the least stable (42%), whereas combined insomnia (CI) was the most stable (69%). Baseline symptom groups (cross-sectionally defined) differed significantly across various clinical indices, including daytime impairment, depression, and anxiety. Importantly, however, a comparison of stable phenotypes (longitudinally defined) did not reveal any differences in impairment or comorbid psychopathology. Another interesting finding was that whereas all other insomnia phenotypes showed evidence of an elevated wake drive both at night and during the day, the “neither criterion” phenotype did not; this latter phenotype exhibited significantly higher daytime sleepiness despite subthreshold onset and maintenance difficulties. Conclusions: By adopting a stringent, stability-based definition, this study offers timely and important data on the longitudinal trajectory of specific insomnia phenotypes. With

  15. DNA testing in hereditary neuropathies.

    LENUS (Irish Health Repository)

    Murphy, Sinéad M

    2013-01-01

    The inherited neuropathies are a clinically and genetically heterogeneous group of disorders in which there have been rapid advances in the last two decades. Molecular genetic testing is now an integral part of the evaluation of patients with inherited neuropathies. In this chapter we describe the genes responsible for the primary inherited neuropathies. We briefly discuss the clinical phenotype of each of the known inherited neuropathy subgroups, describe algorithms for molecular genetic testing of affected patients and discuss genetic counseling. The basic principles of careful phenotyping, documenting an accurate family history, and testing the available genes in an appropriate manner should identify the vast majority of individuals with CMT1 and many of those with CMT2. In this chapter we also describe the current methods of genetic testing. As advances are made in molecular genetic technologies and improvements are made in bioinformatics, it is likely that the current time-consuming methods of DNA sequencing will give way to quicker and more efficient high-throughput methods, which are briefly discussed here.

  16. Comparison of duplex PCR and phenotypic analysis in differentiating Candida dubliniensis from Candida albicans from oral samples.

    Science.gov (United States)

    Sampath, Asanga; Weerasekera, Manjula; Dilhari, Ayomi; Gunasekara, Chinthika; Bulugahapitiya, Uditha; Fernando, Neluka; Samaranayake, Lakshman

    2017-12-01

    Candida dubliniensis shares a wide range of phenotypic characteristics with Candida albicans including a common trait called germ tube positivity. Hence, laboratory differentiation of these two species is cumbersome. Duplex PCR analyses for C. albicans and C. dubliniensis was performed directly on DNA extracted from a total of 122 germ tube positive isolates derived from 100 concentrated oral rinse samples from a random cohort of diabetics attending a clinic in Sri Lanka. These results were confirmed by DNA sequencing of internal transcribed spacer (ITS) region of rDNA of the yeasts. Performance efficacy of duplex PCR was then compared with phenotypic identification using a standard battery of phenotypic tests. Of the 122 germ tube positive isolates three were identified by duplex PCR as C. dubliniensis and the remainder as C. albicans. On the contrary, when the standard phenotypic tests, sugar assimilation and chlamydospore formation, were used to differentiate the two species 13 germ tube positive isolates were erroneously identified as C. dubliniensis. Duplex PCR was found to be rapid, sensitive and more specific than phenotypic identification methods in discriminating C. dubliniensis from C. albicans. This is also the first report on the oral carriage of C. dubliniensis in a Sri Lankan population.

  17. The Phenotype/Genotype Correlation of Lactase Persistence among Omani Adults

    Directory of Open Access Journals (Sweden)

    Abdulrahim Al-Abri

    2013-09-01

    Full Text Available Objective: To examine the correlation of lactase persistence phenotype with genotype in Omani adults.Methods: Lactase persistence phenotype was tested by hydrogen breath test in 52 Omani Adults using the Micro H2 analyzer. Results were checked against genotyping using direct DNA sequencing.Results: Forty one individuals with C/C-13910 and T/T-13915 genotypes had positive breath tests (≥20 ppm; while eight of nine individuals with T/C-13910 or T/G-13915 genotypes had negative breath tests (<20 ppm and two subjects were non-hydrogen producers. The agreement between phenotype and genotype using Kappa value was very good (0.93.Conclusion: Genotyping both T/C-13910 and T/G-13915 alleles can be used to assist diagnosis and predict lactose intolerance in the Omani population.

  18. The Role of ATRX in the Alternative Lengthening of Telomeres (ALT Phenotype

    Directory of Open Access Journals (Sweden)

    João P. Amorim

    2016-09-01

    Full Text Available Telomeres are responsible for protecting chromosome ends in order to prevent the loss of coding DNA. Their maintenance is required for achieving immortality by neoplastic cells and can occur by upregulation of the telomerase enzyme or through a homologous recombination-associated process, the alternative lengthening of telomeres (ALT. The precise mechanisms that govern the activation of ALT or telomerase in tumor cells are not fully understood, although cellular origin may favor one of the other mechanisms that have been found thus far in mutual exclusivity. Specific mutational events influence ALT activation and maintenance: a unifying frequent feature of tumors that acquire this phenotype are the recurrent mutations of the Alpha Thalassemia/Mental Retardation Syndrome X-Linked (ATRX or Death-Domain Associated Protein (DAXX genes. This review summarizes the established criteria about this phenotype: its prevalence, theoretical molecular mechanisms and relation with ATRX, DAXX and other proteins (directly or indirectly interacting and resulting in the ALT phenotype.

  19. The phenotype of FancB-mutant mouse embryonic stem cells

    OpenAIRE

    Kim, Tae Moon; Ko, Jun Ho; Choi, Yong Jun; Hu, Lingchuan; Hasty, Paul

    2011-01-01

    Fanconi anemia (FA) is a rare autosomal recessive disease characterized by bone marrow failure, developmental defects and cancer. There are multiple FA genes that enable the repair of interstrand crosslinks (ICLs) in coordination with a variety of other DNA repair pathways in a way that is poorly understood. Here we present the phenotype of mouse embryonic stem (ES) cells mutated for FancB. We found FancB-mutant cells exhibited reduced cellular proliferation, hypersensitivity to the crosslink...

  20. Mitochondrial Haplogroups Define Two Phenotypes of Osteoarthritis

    Science.gov (United States)

    Fernández-Moreno, Mercedes; Soto-Hermida, Angel; Oreiro, Natividad; Pértega, Sonia; Fenández-López, Carlos; Rego-Pérez, Ignacio; Blanco, Francisco J.

    2012-01-01

    Objective: To assess a mitochondrion-related phenotype in patients with osteoarthritis (OA). Methods: Serum levels of the following OA-related biomarkers: matrix metalloproteinase-1 (MMP-1); MMP-3; MMP-13; myeloperoxidase (MPO); a peptide of the alpha-helical region of type II collagen, Coll2-1, and its nitrated form Coll2-1NO2; a C-terminal neoepitope generated by the collagenase-mediated cleavage of collagen type II triple helix, C2C; the C-propeptide of collagen type II, CPII; hyaluronic acid (HA); human cartilage glycoprotein 39, YKL-40; cartilage oligomeric matrix protein; and cathepsin K were analyzed in 48 OA patients and 52 healthy controls carrying the haplogroups H and J. Logistic regression models and receiver operating characteristic (ROC) curves were performed to predict the onset of OA. Results: MMP-13 was the only biomarker significantly increased in OA patients compared to healthy controls in both haplogroups H and J. The collagen type II biomarkers, Coll2-1, Coll2-1NO2, the Coll2-1NO2/Coll2-1 ratio, C2C, CPII, and the C2C:CPII ratio were significantly increased in OA patients carrying haplogroup H compared to OA carriers of the haplogroup J. Two logistic regression models for diagnosis were constructed and adjusted for age, gender, and body mass index. For haplogroup H, the biomarkers significantly associated with OA were MMP-13 and Coll2-1; the area under the curve (AUC) of the ROC curve for this model was 0.952 (95% CI = 0.892–1.012). For haplogroup J, the only biomarker significantly associated with OA was MMP-13; the AUC for this model was 0.895 (95% CI = 0.801–0.989). Conclusion: The mitochondrial DNA haplogroups are potential complementary candidates for biomarkers of OA; their genotyping in conjunction with the assessment of classical protein molecular markers is recommended. PMID:22593743

  1. Semantic Disease Gene Embeddings (SmuDGE): phenotype-based disease gene prioritization without phenotypes

    KAUST Repository

    AlShahrani, Mona; Hoehndorf, Robert

    2018-01-01

    In the past years, several methods have been developed to incorporate information about phenotypes into computational disease gene prioritization methods. These methods commonly compute the similarity between a disease's (or patient's) phenotypes and a database of gene-to-phenotype associations to find the phenotypically most similar match. A key limitation of these methods is their reliance on knowledge about phenotypes associated with particular genes which is highly incomplete in humans as well as in many model organisms such as the mouse. Results: We developed SmuDGE, a method that uses feature learning to generate vector-based representations of phenotypes associated with an entity. SmuDGE can be used as a trainable semantic similarity measure to compare two sets of phenotypes (such as between a disease and gene, or a disease and patient). More importantly, SmuDGE can generate phenotype representations for entities that are only indirectly associated with phenotypes through an interaction network; for this purpose, SmuDGE exploits background knowledge in interaction networks comprising of multiple types of interactions. We demonstrate that SmuDGE can match or outperform semantic similarity in phenotype-based disease gene prioritization, and furthermore significantly extends the coverage of phenotype-based methods to all genes in a connected interaction network.

  2. Semantic Disease Gene Embeddings (SmuDGE): phenotype-based disease gene prioritization without phenotypes

    KAUST Repository

    Alshahrani, Mona

    2018-04-30

    In the past years, several methods have been developed to incorporate information about phenotypes into computational disease gene prioritization methods. These methods commonly compute the similarity between a disease\\'s (or patient\\'s) phenotypes and a database of gene-to-phenotype associations to find the phenotypically most similar match. A key limitation of these methods is their reliance on knowledge about phenotypes associated with particular genes which is highly incomplete in humans as well as in many model organisms such as the mouse. Results: We developed SmuDGE, a method that uses feature learning to generate vector-based representations of phenotypes associated with an entity. SmuDGE can be used as a trainable semantic similarity measure to compare two sets of phenotypes (such as between a disease and gene, or a disease and patient). More importantly, SmuDGE can generate phenotype representations for entities that are only indirectly associated with phenotypes through an interaction network; for this purpose, SmuDGE exploits background knowledge in interaction networks comprising of multiple types of interactions. We demonstrate that SmuDGE can match or outperform semantic similarity in phenotype-based disease gene prioritization, and furthermore significantly extends the coverage of phenotype-based methods to all genes in a connected interaction network.

  3. Human cytomegalovirus uracil DNA glycosylase associates with ppUL44 and accelerates the accumulation of viral DNA

    Directory of Open Access Journals (Sweden)

    Dixon Melissa

    2005-07-01

    Full Text Available Abstract Background Human cytomegalovirus UL114 encodes a uracil-DNA glycosylase homolog that is highly conserved in all characterized herpesviruses that infect mammals. Previous studies demonstrated that the deletion of this nonessential gene delays significantly the onset of viral DNA synthesis and results in a prolonged replication cycle. The gene product, pUL114, also appears to be important in late phase DNA synthesis presumably by introducing single stranded breaks. Results A series of experiments was performed to formally assign the observed phenotype to pUL114 and to characterize the function of the protein in viral replication. A cell line expressing pUL114 complemented the observed phenotype of a UL114 deletion virus in trans, confirming that the observed defects were the result of a deficiency in this gene product. Stocks of recombinant viruses without elevated levels of uracil were produced in the complementing cells; however they retained the phenotype of poor growth in normal fibroblasts suggesting that poor replication was unrelated to uracil content of input genomes. Recombinant viruses expressing epitope tagged versions of this gene demonstrated that pUL114 was expressed at early times and that it localized to viral replication compartments. This protein also coprecipitated with the DNA polymerase processivity factor, ppUL44 suggesting that these proteins associate in infected cells. This apparent interaction did not appear to require other viral proteins since ppUL44 could recruit pUL114 to the nucleus in uninfected cells. An analysis of DNA replication kinetics revealed that the initial rate of DNA synthesis and the accumulation of progeny viral genomes were significantly reduced compared to the parent virus. Conclusion These data suggest that pUL114 associates with ppUL44 and that it functions as part of the viral DNA replication complex to increase the efficiency of both early and late phase viral DNA synthesis.

  4. Newly identified CHO ERCC3/XPB mutations and phenotype characterization

    Science.gov (United States)

    Rybanská, Ivana; Gurský, Ján; Fašková, Miriam; Salazar, Edmund P.; Kimlíčková-Polakovičová, Erika; Kleibl, Karol; Thompson, Larry H.; Piršel, Miroslav

    2010-01-01

    Nucleotide excision repair (NER) is a complex multistage process involving many interacting gene products to repair a wide range of DNA lesions. Genetic defects in NER cause human hereditary diseases including xeroderma pigmentosum (XP), Cockayne syndrome (CS), trichothiodystrophy and a combined XP/CS overlapping symptom. One key gene product associated with all these disorders is the excision repair cross-complementing 3/xeroderma pigmentosum B (ERCC3/XPB) DNA helicase, a subunit of the transcription factor IIH complex. ERCC3 is involved in initiation of basal transcription and global genome repair as well as in transcription-coupled repair (TCR). The hamster ERCC3 gene shows high degree of homology with the human ERCC3/XPB gene. We identified new mutations in the Chinese hamster ovary cell ERCC3 gene and characterized the role of hamster ERCC3 protein in DNA repair of ultraviolet (UV)-induced and oxidative DNA damage. All but one newly described mutations are located in the protein C-terminal region around the last intron–exon boundary. Due to protein truncations or frameshifts, they lack amino acid Ser751, phosphorylation of which prevents the 5′ incision of the UV-induced lesion during NER. Thus, despite the various locations of the mutations, their phenotypes are similar. All ercc3 mutants are extremely sensitive to UV-C light and lack recovery of RNA synthesis (RRS), confirming a defect in TCR of UV-induced damage. Their limited global genome NER capacity averages ∼8%. We detected modest sensitivity of ercc3 mutants to the photosensitizer Ro19-8022, which primarily introduces 8-oxoguanine lesions into DNA. Ro19-8022-induced damage interfered with RRS, and some of the ercc3 mutants had delayed kinetics. All ercc3 mutants showed efficient base excision repair (BER). Thus, the positions of the mutations have no effect on the sensitivity to, and repair of, Ro19-8022-induced DNA damage, suggesting that the ERCC3 protein is not involved in BER. PMID:19942596

  5. Methylator phenotype of malignant germ cell tumours in children identifies strong candidates for chemotherapy resistance.

    Science.gov (United States)

    Jeyapalan, J N; Noor, D A Mohamed; Lee, S-H; Tan, C L; Appleby, V A; Kilday, J P; Palmer, R D; Schwalbe, E C; Clifford, S C; Walker, D A; Murray, M J; Coleman, N; Nicholson, J C; Scotting, P J

    2011-08-09

    Yolk sac tumours (YSTs) and germinomas are the two major pure histological subtypes of germ cell tumours. To date, the role of DNA methylation in the aetiology of this class of tumour has only been analysed in adult testicular forms and with respect to only a few genes. A bank of paediatric tumours was analysed for global methylation of LINE-1 repeat elements and global methylation of regulatory elements using GoldenGate methylation arrays. Both germinomas and YSTs exhibited significant global hypomethylation of LINE-1 elements. However, in germinomas, methylation of gene regulatory regions differed little from control samples, whereas YSTs exhibited increased methylation at a large proportion of the loci tested, showing a 'methylator' phenotype, including silencing of genes associated with Caspase-8-dependent apoptosis. Furthermore, we found that the methylator phenotype of YSTs was coincident with higher levels of expression of the DNA methyltransferase, DNA (cytosine-5)-methyltransferase 3B, suggesting a mechanism underlying the phenotype. Epigenetic silencing of a large number of potential tumour suppressor genes in YSTs might explain why they exhibit a more aggressive natural history than germinomas and silencing of genes associated with Caspase-8-dependent cell death might explain the relative resistance of YSTs to conventional therapy.

  6. Phenotypic spectrum associated with mutations of the mitochondrial polymerase gamma gene.

    Science.gov (United States)

    Horvath, Rita; Hudson, Gavin; Ferrari, Gianfrancesco; Fütterer, Nancy; Ahola, Sofia; Lamantea, Eleonora; Prokisch, Holger; Lochmüller, Hanns; McFarland, Robert; Ramesh, V; Klopstock, Thomas; Freisinger, Peter; Salvi, Fabrizio; Mayr, Johannes A; Santer, Rene; Tesarova, Marketa; Zeman, Jiri; Udd, Bjarne; Taylor, Robert W; Turnbull, Douglass; Hanna, Michael; Fialho, Doreen; Suomalainen, Anu; Zeviani, Massimo; Chinnery, Patrick F

    2006-07-01

    Mutations in the gene coding for the catalytic subunit of the mitochondrial DNA (mtDNA) polymerase gamma (POLG1) have recently been described in patients with diverse clinical presentations, revealing a complex relationship between genotype and phenotype in patients and their families. POLG1 was sequenced in patients from different European diagnostic and research centres to define the phenotypic spectrum and advance understanding of the recurrence risks. Mutations were identified in 38 cases, with the majority being sporadic compound heterozygotes. Eighty-nine DNA sequence changes were identified, including 2 predicted to alter a splice site, 1 predicted to cause a premature stop codon and 13 predicted to cause novel amino acid substitutions. The majority of children had a mutation in the linker region, often 1399G-->A (A467T), and a mutation affecting the polymerase domain. Others had mutations throughout the gene, and 11 had 3 or more substitutions. The clinical presentation ranged from the neonatal period to late adult life, with an overlapping phenotypic spectrum from severe encephalopathy and liver failure to late-onset external ophthalmoplegia, ataxia, myopathy and isolated muscle pain or epilepsy. There was a strong gender bias in children, with evidence of an environmental interaction with sodium valproate. POLG1 mutations cause an overlapping clinical spectrum of disease with both dominant and recessive modes of inheritance. 1399G-->A (A467T) is common in children, but complete POLG1 sequencing is required to identify multiple mutations that can have complex implications for genetic counselling.

  7. Integrating phenotype ontologies with PhenomeNET

    KAUST Repository

    Rodriguez-Garcia, Miguel Angel

    2017-12-19

    Background Integration and analysis of phenotype data from humans and model organisms is a key challenge in building our understanding of normal biology and pathophysiology. However, the range of phenotypes and anatomical details being captured in clinical and model organism databases presents complex problems when attempting to match classes across species and across phenotypes as diverse as behaviour and neoplasia. We have previously developed PhenomeNET, a system for disease gene prioritization that includes as one of its components an ontology designed to integrate phenotype ontologies. While not applicable to matching arbitrary ontologies, PhenomeNET can be used to identify related phenotypes in different species, including human, mouse, zebrafish, nematode worm, fruit fly, and yeast. Results Here, we apply the PhenomeNET to identify related classes from two phenotype and two disease ontologies using automated reasoning. We demonstrate that we can identify a large number of mappings, some of which require automated reasoning and cannot easily be identified through lexical approaches alone. Combining automated reasoning with lexical matching further improves results in aligning ontologies. Conclusions PhenomeNET can be used to align and integrate phenotype ontologies. The results can be utilized for biomedical analyses in which phenomena observed in model organisms are used to identify causative genes and mutations underlying human disease.

  8. Redefining Aging in HIV Infection Using Phenotypes.

    Science.gov (United States)

    Stoff, David M; Goodkin, Karl; Jeste, Dilip; Marquine, Maria

    2017-10-01

    This article critically reviews the utility of "phenotypes" as behavioral descriptors in aging/HIV research that inform biological underpinnings and treatment development. We adopt a phenotypic redefinition of aging conceptualized within a broader context of HIV infection and of aging. Phenotypes are defined as dimensions of behavior, closely related to fundamental mechanisms, and, thus, may be more informative than chronological age. Primary emphasis in this review is given to comorbid aging and cognitive aging, though other phenotypes (i.e., disability, frailty, accelerated aging, successful aging) are also discussed in relation to comorbid aging and cognitive aging. The main findings that emerged from this review are as follows: (1) the phenotypes, comorbid aging and cognitive aging, are distinct from each other, yet overlapping; (2) associative relationships are the rule in HIV for comorbid and cognitive aging phenotypes; and (3) HIV behavioral interventions for both comorbid aging and cognitive aging have been limited. Three paths for research progress are identified for phenotype-defined aging/HIV research (i.e., clinical and behavioral specification, biological mechanisms, intervention targets), and some important research questions are suggested within each of these research paths.

  9. Evolution of molecular phenotypes under stabilizing selection

    International Nuclear Information System (INIS)

    Nourmohammad, Armita; Schiffels, Stephan; Lässig, Michael

    2013-01-01

    Molecular phenotypes are important links between genomic information and organismic functions, fitness, and evolution. Complex phenotypes, which are also called quantitative traits, often depend on multiple genomic loci. Their evolution builds on genome evolution in a complicated way, which involves selection, genetic drift, mutations and recombination. Here we develop a coarse-grained evolutionary statistics for phenotypes, which decouples from details of the underlying genotypes. We derive approximate evolution equations for the distribution of phenotype values within and across populations. This dynamics covers evolutionary processes at high and low recombination rates, that is, it applies to sexual and asexual populations. In a fitness landscape with a single optimal phenotype value, the phenotypic diversity within populations and the divergence between populations reach evolutionary equilibria, which describe stabilizing selection. We compute the equilibrium distributions of both quantities analytically and we show that the ratio of mean divergence and diversity depends on the strength of selection in a universal way: it is largely independent of the phenotype’s genomic encoding and of the recombination rate. This establishes a new method for the inference of selection on molecular phenotypes beyond the genome level. We discuss the implications of our findings for the predictability of evolutionary processes. (paper)

  10. Replicative DNA polymerase mutations in cancer☆

    Science.gov (United States)

    Heitzer, Ellen; Tomlinson, Ian

    2014-01-01

    Three DNA polymerases — Pol α, Pol δ and Pol ɛ — are essential for DNA replication. After initiation of DNA synthesis by Pol α, Pol δ or Pol ɛ take over on the lagging and leading strand respectively. Pol δ and Pol ɛ perform the bulk of replication with very high fidelity, which is ensured by Watson–Crick base pairing and 3′exonuclease (proofreading) activity. Yeast models have shown that mutations in the exonuclease domain of Pol δ and Pol ɛ homologues can cause a mutator phenotype. Recently, we identified germline exonuclease domain mutations (EDMs) in human POLD1 and POLE that predispose to ‘polymerase proofreading associated polyposis’ (PPAP), a disease characterised by multiple colorectal adenomas and carcinoma, with high penetrance and dominant inheritance. Moreover, somatic EDMs in POLE have also been found in sporadic colorectal and endometrial cancers. Tumors with EDMs are microsatellite stable and show an ‘ultramutator’ phenotype, with a dramatic increase in base substitutions. PMID:24583393

  11. Replicative DNA polymerase mutations in cancer.

    Science.gov (United States)

    Heitzer, Ellen; Tomlinson, Ian

    2014-02-01

    Three DNA polymerases - Pol α, Pol δ and Pol ɛ - are essential for DNA replication. After initiation of DNA synthesis by Pol α, Pol δ or Pol ɛ take over on the lagging and leading strand respectively. Pol δ and Pol ɛ perform the bulk of replication with very high fidelity, which is ensured by Watson-Crick base pairing and 3'exonuclease (proofreading) activity. Yeast models have shown that mutations in the exonuclease domain of Pol δ and Pol ɛ homologues can cause a mutator phenotype. Recently, we identified germline exonuclease domain mutations (EDMs) in human POLD1 and POLE that predispose to 'polymerase proofreading associated polyposis' (PPAP), a disease characterised by multiple colorectal adenomas and carcinoma, with high penetrance and dominant inheritance. Moreover, somatic EDMs in POLE have also been found in sporadic colorectal and endometrial cancers. Tumors with EDMs are microsatellite stable and show an 'ultramutator' phenotype, with a dramatic increase in base substitutions. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  12. Genetic and phenotypic diversity of Rhizobium isolates from Southern Ecuador

    Directory of Open Access Journals (Sweden)

    Roldán Torres-Gutiérrez

    Full Text Available ABSTRACT Rhizobium-legume symbioses play relevant roles in agriculture but have not been well studied in Ecuador. The aim of this study was to characterize the genetic and phenotypic diversity of Rhizobium isolates associated with Phaseolus vulgaris from southern Ecuador. Morpho-cultural characterization, biochemical tests and physiological analyses were conducted to authenticate and determine the diversity of bacteria Rhizobium-like isolates. The genetic diversity of the isolates was determined by molecular techniques, which consisted of bacteria DNA extraction and amplification and sequencing of the 16S rRNA gene. The nodulation parameters and nitrogen fixation for P. vulgaris under greenhouse conditions were also assessed to determine the phenotypic diversity among isolates. Furthermore, bacteria indole-acetic-acid production was evaluated by the colorimetric method. Morpho-cultural and biochemical characteristic assessments demonstrated that Rhizobium-like bacteria was associated with the P. vulgaris nodules. The diversity among the isolates, as determined by physiological analyses, revealed the potential of several isolates to grow at different pH values, salinity conditions and temperatures. Partial sequencing of the 16S rRNA gene identified the Rhizobium genus in every sampling site. From a total of 20 aligned sequences, nine species of Rhizobium were identified. Nodule formation and biomass, as well as nitrogen fixation, showed an increase in plant phenotypic parameters, which could be influenced by IAA production, especially for the strains R. mesoamericanum NAM1 and R. leguminosarum bv. viciae COL6. These results demonstrated the efficiency of native symbiotic diazotrophic strains inoculants for legume production. This work can serve as the basis for additional studies of native Rhizobium strains and to help spread the use of biofertilizers in Ecuadorian fields.

  13. The nature of stable insomnia phenotypes.

    Science.gov (United States)

    Pillai, Vivek; Roth, Thomas; Drake, Christopher L

    2015-01-01

    We examined the 1-y stability of four insomnia symptom profiles: sleep onset insomnia; sleep maintenance insomnia; combined onset and maintenance insomnia; and neither criterion (i.e., insomnia cases that do not meet quantitative thresholds for onset or maintenance problems). Insomnia cases that exhibited the same symptom profile over a 1-y period were considered to be phenotypes, and were compared in terms of clinical and demographic characteristics. Longitudinal. Urban, community-based. Nine hundred fifty-four adults with Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition based current insomnia (46.6 ± 12.6 y; 69.4% female). None. At baseline, participants were divided into four symptom profile groups based on quantitative criteria. Follow-up assessment 1 y later revealed that approximately 60% of participants retained the same symptom profile, and were hence judged to be phenotypes. Stability varied significantly by phenotype, such that sleep onset insomnia (SOI) was the least stable (42%), whereas combined insomnia (CI) was the most stable (69%). Baseline symptom groups (cross-sectionally defined) differed significantly across various clinical indices, including daytime impairment, depression, and anxiety. Importantly, however, a comparison of stable phenotypes (longitudinally defined) did not reveal any differences in impairment or comorbid psychopathology. Another interesting finding was that whereas all other insomnia phenotypes showed evidence of an elevated wake drive both at night and during the day, the 'neither criterion' phenotype did not; this latter phenotype exhibited significantly higher daytime sleepiness despite subthreshold onset and maintenance difficulties. By adopting a stringent, stability-based definition, this study offers timely and important data on the longitudinal trajectory of specific insomnia phenotypes. With the exception of daytime sleepiness, few clinical differences are apparent across stable phenotypes.

  14. Assessing the fidelity of ancient DNA sequences amplified from nuclear genes

    DEFF Research Database (Denmark)

    Binladen, Jonas; Wiuf, Carsten Henrik; Gilbert, M. Thomas P.

    2006-01-01

    To date, the field of ancient DNA has relied almost exclusively on mitochondrial DNA (mtDNA) sequences. However, a number of recent studies have reported the successful recovery of ancient nuclear DNA (nuDNA) sequences, thereby allowing the characterization of genetic loci directly involved...... in phenotypic traits of extinct taxa. It is well documented that postmortem damage in ancient mtDNA can lead to the generation of artifactual sequences. However, as yet no one has thoroughly investigated the damage spectrum in ancient nuDNA. By comparing clone sequences from 23 fossil specimens, recovered from...... adenine), respectively. Type 2 transitions are by far the most dominant and increase relative to those of type 1 with damage load. The results suggest that the deamination of cytosine (and 5-methyl cytosine) to uracil (and thymine) is the main cause of miscoding lesions in both ancient mtDNA and nu...

  15. Phenotyping animal models of diabetic neuropathy

    DEFF Research Database (Denmark)

    Biessels, G J; Bril, V; Calcutt, N A

    2014-01-01

    NIDDK, JDRF, and the Diabetic Neuropathy Study Group of EASD sponsored a meeting to explore the current status of animal models of diabetic peripheral neuropathy. The goal of the workshop was to develop a set of consensus criteria for the phenotyping of rodent models of diabetic neuropathy...... with a discussion on the merits and limitations of a unified approach to phenotyping rodent models of diabetic neuropathy and a consensus formed on the definition of the minimum criteria required for establishing the presence of the disease. A neuropathy phenotype in rodents was defined as the presence...

  16. Evidence that DNA polymerase δ contributes to initiating leading strand DNA replication in Saccharomyces cerevisiae.

    Science.gov (United States)

    Garbacz, Marta A; Lujan, Scott A; Burkholder, Adam B; Cox, Phillip B; Wu, Qiuqin; Zhou, Zhi-Xiong; Haber, James E; Kunkel, Thomas A

    2018-02-27

    To investigate nuclear DNA replication enzymology in vivo, we have studied Saccharomyces cerevisiae strains containing a pol2-16 mutation that inactivates the catalytic activities of DNA polymerase ε (Pol ε). Although pol2-16 mutants survive, they present very tiny spore colonies, increased doubling time, larger than normal cells, aberrant nuclei, and rapid acquisition of suppressor mutations. These phenotypes reveal a severe growth defect that is distinct from that of strains that lack only Pol ε proofreading (pol2-4), consistent with the idea that Pol ε is the major leading-strand polymerase used for unstressed DNA replication. Ribonucleotides are incorporated into the pol2-16 genome in patterns consistent with leading-strand replication by Pol δ when Pol ε is absent. More importantly, ribonucleotide distributions at replication origins suggest that in strains encoding all three replicases, Pol δ contributes to initiation of leading-strand replication. We describe two possible models.

  17. DNA preservation in silk.

    Science.gov (United States)

    Liu, Yawen; Zheng, Zhaozhu; Gong, He; Liu, Meng; Guo, Shaozhe; Li, Gang; Wang, Xiaoqin; Kaplan, David L

    2017-06-27

    The structure of DNA is susceptible to alterations at high temperature and on changing pH, irradiation and exposure to DNase. Options to protect and preserve DNA during storage are important for applications in genetic diagnosis, identity authentication, drug development and bioresearch. In the present study, the stability of total DNA purified from human dermal fibroblast cells, as well as that of plasmid DNA, was studied in silk protein materials. The DNA/silk mixtures were stabilized on filter paper (silk/DNA + filter) or filter paper pre-coated with silk and treated with methanol (silk/DNA + PT-filter) as a route to practical utility. After air-drying and water extraction, 50-70% of the DNA and silk could be retrieved and showed a single band on electrophoretic gels. 6% silk/DNA + PT-filter samples provided improved stability in comparison with 3% silk/DNA + filter samples and DNA + filter samples for DNA preservation, with ∼40% of the band intensity remaining at 37 °C after 40 days and ∼10% after exposure to UV light for 10 hours. Quantitative analysis using the PicoGreen assay confirmed the results. The use of Tris/borate/EDTA (TBE) buffer enhanced the preservation and/or extraction of the DNA. The DNA extracted after storage maintained integrity and function based on serving as a functional template for PCR amplification of the gene for zinc finger protein 750 (ZNF750) and for transgene expression of red fluorescence protein (dsRed) in HEK293 cells. The high molecular weight and high content of a crystalline beta-sheet structure formed on the coated surfaces likely accounted for the preservation effects observed for the silk/DNA + PT-filter samples. Although similar preservation effects were also obtained for lyophilized silk/DNA samples, the rapid and simple processing available with the silk-DNA-filter membrane system makes it appealing for future applications.

  18. Force induced DNA melting

    International Nuclear Information System (INIS)

    Santosh, Mogurampelly; Maiti, Prabal K

    2009-01-01

    When pulled along the axis, double-strand DNA undergoes a large conformational change and elongates by roughly twice its initial contour length at a pulling force of about 70 pN. The transition to this highly overstretched form of DNA is very cooperative. Applying a force perpendicular to the DNA axis (unzipping), double-strand DNA can also be separated into two single-stranded DNA, this being a fundamental process in DNA replication. We study the DNA overstretching and unzipping transition using fully atomistic molecular dynamics (MD) simulations and argue that the conformational changes of double-strand DNA associated with either of the above mentioned processes can be viewed as force induced DNA melting. As the force at one end of the DNA is increased the DNA starts melting abruptly/smoothly above a critical force depending on the pulling direction. The critical force f m , at which DNA melts completely decreases as the temperature of the system is increased. The melting force in the case of unzipping is smaller compared to the melting force when the DNA is pulled along the helical axis. In the case of melting through unzipping, the double-strand separation has jumps which correspond to the different energy minima arising due to sequence of different base pairs. The fraction of Watson-Crick base pair hydrogen bond breaking as a function of force does not show smooth and continuous behavior and consists of plateaus followed by sharp jumps.

  19. DNA damage and autophagy

    International Nuclear Information System (INIS)

    Rodriguez-Rocha, Humberto; Garcia-Garcia, Aracely; Panayiotidis, Mihalis I.; Franco, Rodrigo

    2011-01-01

    Both exogenous and endogenous agents are a threat to DNA integrity. Exogenous environmental agents such as ultraviolet (UV) and ionizing radiation, genotoxic chemicals and endogenous byproducts of metabolism including reactive oxygen species can cause alterations in DNA structure (DNA damage). Unrepaired DNA damage has been linked to a variety of human disorders including cancer and neurodegenerative disease. Thus, efficient mechanisms to detect DNA lesions, signal their presence and promote their repair have been evolved in cells. If DNA is effectively repaired, DNA damage response is inactivated and normal cell functioning resumes. In contrast, when DNA lesions cannot be removed, chronic DNA damage triggers specific cell responses such as cell death and senescence. Recently, DNA damage has been shown to induce autophagy, a cellular catabolic process that maintains a balance between synthesis, degradation, and recycling of cellular components. But the exact mechanisms by which DNA damage triggers autophagy are unclear. More importantly, the role of autophagy in the DNA damage response and cellular fate is unknown. In this review we analyze evidence that supports a role for autophagy as an integral part of the DNA damage response.

  20. DNA Open states and DNA hydratation

    International Nuclear Information System (INIS)

    Lema-Larre, B. de; Martin-Landrove, M

    1995-01-01

    It is a very well-known fact that an protonic exchange exists among natural DNA filaments and synthetic polynucleotides with the solvent (1--2). The existence of DNA open states, that is to say states for which the interior of the DNA molecule is exposed to the external environment, it has been demonstrated by means of proton-deuterium exchange (3). This work has carried out experiments measuring the dispersion of the traverse relaxation rate (4), as a pulsation rate function in a Carr-Purcell-Meiboom-Gill (CPMG) pulses sequence rate, to determine changes in the moist layer of the DNA molecule. The experiments were carried out under different experimental conditions in order to vary the probability that open states occurs, such as temperature or the exposure to electromagnetic fields. Some theoretical models were supposed to adjust the experimental results including those related to DNA non linear dynamic [es

  1. Phenotypic profiles of Armenian grape cultivars

    Directory of Open Access Journals (Sweden)

    Aroutiounian Rouben

    2015-01-01

    Full Text Available The conservation and sustainable use of grapevine biodiversity in Armenia is particularly important due to the large number of traditional local varieties. Being partially different from European grapevine gene pool, the material of Armenian local cultivars significantly contributes to the understanding of the genetic variation and is valuable source for target selection. During last years many Armenian grapevine cultivars have been already described and their genotypes determined, but some local varieties and wild accessions remain unidentified and their phenotypic characteristics overlooked. The comprehensive analysis of phenotypes is essential for research, including genetic association studies, cultivar evaluation and selection. The goal of our research was the phenotyping on the base of reproductive, carpological and analytical characteristics of 80 Armenian aboriginal and new grape cultivars. Description of phenotypic profiles is important step towards identification and conservation of genetic resources of Armenian grapes. In future, these data can be applied for breeding of improved grape varieties targeted to fresh consumption and wine production.

  2. Phenotypic diversity and phylogenetic relationship between the ...

    African Journals Online (AJOL)

    Phenotypic diversity and phylogenetic relationship between the Bakosi/Baweri and other pig breeds ( Sus scrofa Domesticus ) in the humid forest with monomodal rainfall agro-ecological zone of Cameroon.

  3. Mining skeletal phenotype descriptions from scientific literature.

    Directory of Open Access Journals (Sweden)

    Tudor Groza

    Full Text Available Phenotype descriptions are important for our understanding of genetics, as they enable the computation and analysis of a varied range of issues related to the genetic and developmental bases of correlated characters. The literature contains a wealth of such phenotype descriptions, usually reported as free-text entries, similar to typical clinical summaries. In this paper, we focus on creating and making available an annotated corpus of skeletal phenotype descriptions. In addition, we present and evaluate a hybrid Machine Learning approach for mining phenotype descriptions from free text. Our hybrid approach uses an ensemble of four classifiers and experiments with several aggregation techniques. The best scoring technique achieves an F-1 score of 71.52%, which is close to the state-of-the-art in other domains, where training data exists in abundance. Finally, we discuss the influence of the features chosen for the model on the overall performance of the method.

  4. Integrating phenotype ontologies with PhenomeNET

    KAUST Repository

    Rodriguez-Garcia, Miguel Angel; Gkoutos, Georgios V.; Schofield, Paul N.; Hoehndorf, Robert

    2017-01-01

    in clinical and model organism databases presents complex problems when attempting to match classes across species and across phenotypes as diverse as behaviour and neoplasia. We have previously developed PhenomeNET, a system for disease gene prioritization

  5. Wiedemann-Rautenstrauch syndrome: A phenotype analysis

    NARCIS (Netherlands)

    Paolacci, Stefano; Bertola, Debora; Franco, José; Mohammed, Shehla; Tartaglia, Marco; Wollnik, Bernd; Hennekam, Raoul C.

    2017-01-01

    Wiedemann-Rautenstrauch syndrome (WRS) is a neonatal progeroid disorder characterized by growth retardation, lipodystrophy, a distinctive face, and dental anomalies. Patients reported to date demonstrate a remarkable variability in phenotype, which hampers diagnostics. We performed a literature

  6. REVIEW ARTICLE One gene, many phenotypes

    African Journals Online (AJOL)

    salah

    Phenotype descriptions are valuable information right at the interface of medi- cine and biology. ... the interaction of alleles at different loci. Modifier genes. 5. ... the amount of normal protein is called ..... Institute, using computer simulations,.

  7. The spatial patterns of directional phenotypic selection

    KAUST Repository

    Siepielski, Adam M.; Gotanda, Kiyoko M.; Morrissey, Michael B.; Diamond, Sarah E.; DiBattista, Joseph; Carlson, Stephanie Marie

    2013-01-01

    the spatial patterns of selection, namely the extent of variation among populations in the strength and direction of selection. Here, we analyse a data set of spatially replicated studies of directional phenotypic selection from natural populations. The data

  8. A Mutator Phenotype Promoting the Emergence of Spontaneous Oxidative Stress-Resistant Mutants in Campylobacter jejuni.

    Science.gov (United States)

    Dai, Lei; Sahin, Orhan; Tang, Yizhi; Zhang, Qijing

    2017-12-15

    Campylobacter jejuni is a leading cause of foodborne illnesses worldwide. As a microaerophilic organism, C. jejuni must be able to defend against oxidative stress encountered both in the host and in the environment. How Campylobacter utilizes a mutation-based mechanism for adaptation to oxidative stress is still unknown. Here we present a previously undescribed phenotypic and genetic mechanism that promotes the emergence of oxidative stress-resistant mutants. Specifically, we showed that a naturally occurring mutator phenotype, resulting from a loss of function mutation in the DNA repair enzyme MutY, increased oxidative stress resistance (OX R ) in C. jejuni We further demonstrated that MutY malfunction did not directly contribute to the OX R phenotype but increased the spontaneous mutation rate in the peroxide regulator gene perR , which functions as a repressor for multiple genes involved in oxidative stress resistance. Mutations in PerR resulted in loss of its DNA binding function and derepression of PerR-controlled oxidative stress defense genes, thereby conferring an OX R phenotype and facilitating Campylobacter survival under oxidative stress. These findings reveal a new mechanism that promotes the emergence of spontaneous OX R mutants in bacterial organisms. IMPORTANCE Although a mutator phenotype has been shown to promote antibiotic resistance in many bacterial species, little is known about its contribution to the emergence of OX R mutants. This work describes the link between a mutator phenotype and the enhanced emergence of OX R mutants as well as its underlying mechanism involving DNA repair and mutations in PerR. Since DNA repair systems and PerR are well conserved in many bacterial species, especially in Gram positives, the same mechanism may operate in multiple bacterial species. Additionally, we developed a novel method that allows for rapid quantification of spontaneous OX R mutants in a bacterial population. This method represents a technical

  9. Atypical disease phenotypes in pediatric ulcerative colitis

    DEFF Research Database (Denmark)

    Levine, Arie; de Bie, Charlotte I; Turner, Dan

    2013-01-01

    Definitive diagnosis of pediatric ulcerative colitis (UC) may be particularly challenging since isolated colitis with overlapping features is common in pediatric Crohn's disease (CD), while atypical phenotypes of UC are not uncommon. The Paris classification allows more accurate phenotyping...... of atypical inflammatory bowel disease (IBD) patients. Our aim was to identify the prevalence of atypical disease patterns in new-onset pediatric UC using the Paris classification....

  10. A Regulatory RNA Inducing Transgenerationally Inherited Phenotypes

    DEFF Research Database (Denmark)

    Jensen, Lea Møller

    . The variation in Arabidopsis enables different regulatory networks and mechanisms to shape the phenotypic characteristics. The thesis describes the identification of regulatory RNA encoded by an enzyme encoding gene. The RNA regulates by inducing transgenerationally inherited phenotypes. The function of the RNA...... is dependent on the genetic background illustrating that polymorphisms are found in either interactors or target genes of the RNA. Furthermore, the RNA provides a mechanistic link between accumulation of glucosinolate and onset of flowering....

  11. Volatility of Mutator Phenotypes at Single Cell Resolution.

    Directory of Open Access Journals (Sweden)

    Scott R Kennedy

    2015-04-01

    Full Text Available Mutator phenotypes accelerate the evolutionary process of neoplastic transformation. Historically, the measurement of mutation rates has relied on scoring the occurrence of rare mutations in target genes in large populations of cells. Averaging mutation rates over large cell populations assumes that new mutations arise at a constant rate during each cell division. If the mutation rate is not constant, an expanding mutator population may contain subclones with widely divergent rates of evolution. Here, we report mutation rate measurements of individual cell divisions of mutator yeast deficient in DNA polymerase ε proofreading and base-base mismatch repair. Our data are best fit by a model in which cells can assume one of two distinct mutator states, with mutation rates that differ by an order of magnitude. In error-prone cell divisions, mutations occurred on the same chromosome more frequently than expected by chance, often in DNA with similar predicted replication timing, consistent with a spatiotemporal dimension to the hypermutator state. Mapping of mutations onto predicted replicons revealed that mutations were enriched in the first half of the replicon as well as near termination zones. Taken together, our findings show that individual genome replication events exhibit an unexpected volatility that may deepen our understanding of the evolution of mutator-driven malignancies.

  12. Immunoassay of DNA damage

    International Nuclear Information System (INIS)

    Gasparro, F.P.; Santella, R.M.

    1988-01-01

    The direct photomodification of DNA by ultraviolet light or the photo-induced addition of exogenous compounds to DNA components results in alterations of DNA structure ranging from subtle to profound. There are two consequences of these conformational changes. First, cells in which the DNA has been damaged are capable of executing repair steps. Second, the DNA which is usually of very low immunogenicity now becomes highly antigenic. This latter property has allowed the production of a series of monoclonal antibodies that recognize photo-induced DNA damage. Monoclonal antibodies have been generated that recognize the 4',5'-monoadduct and the crosslink of 8-methoxypsoralen in DNA. In addition, another antibody has been prepared which recognizes the furan-side monoadduct of 6,4,4'-trimethylangelicin in DNA. These monoclonal antibodies have been characterized as to sensitivity and specificity using non-competitive and competitive enzyme-linked-immunosorbent assays (ELISA). (author)

  13. Immunoassay of DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Gasparro, F P; Santella, R M

    1988-09-01

    The direct photomodification of DNA by ultraviolet light or the photo-induced addition of exogenous compounds to DNA components results in alterations of DNA structure ranging from subtle to profound. There are two consequences of these conformational changes. First, cells in which the DNA has been damaged are capable of executing repair steps. Second, the DNA which is usually of very low immunogenicity now becomes highly antigenic. This latter property has allowed the production of a series of monoclonal antibodies that recognize photo-induced DNA damage. Monoclonal antibodies have been generated that recognize the 4',5'-monoadduct and the crosslink of 8-methoxypsoralen in DNA. In addition, another antibody has been prepared which recognizes the furan-side monoadduct of 6,4,4'-trimethylangelicin in DNA. These monoclonal antibodies have been characterized as to sensitivity and specificity using non-competitive and competitive enzyme-linked-immunosorbent assays (ELISA).

  14. DNA computing models

    CERN Document Server

    Ignatova, Zoya; Zimmermann, Karl-Heinz

    2008-01-01

    In this excellent text, the reader is given a comprehensive introduction to the field of DNA computing. The book emphasizes computational methods to tackle central problems of DNA computing, such as controlling living cells, building patterns, and generating nanomachines.

  15. DNA tagged microparticles

    Science.gov (United States)

    Farquar, George Roy; Leif, Roald N; Wheeler, Elizabeth

    2015-05-05

    A simulant that includes a carrier and DNA encapsulated in the carrier. Also a method of making a simulant including the steps of providing a carrier and encapsulating DNA in the carrier to produce the simulant.

  16. Modeling DNA Replication.

    Science.gov (United States)

    Bennett, Joan

    1998-01-01

    Recommends the use of a model of DNA made out of Velcro to help students visualize the steps of DNA replication. Includes a materials list, construction directions, and details of the demonstration using the model parts. (DDR)

  17. Phenex: ontological annotation of phenotypic diversity.

    Directory of Open Access Journals (Sweden)

    James P Balhoff

    2010-05-01

    Full Text Available Phenotypic differences among species have long been systematically itemized and described by biologists in the process of investigating phylogenetic relationships and trait evolution. Traditionally, these descriptions have been expressed in natural language within the context of individual journal publications or monographs. As such, this rich store of phenotype data has been largely unavailable for statistical and computational comparisons across studies or integration with other biological knowledge.Here we describe Phenex, a platform-independent desktop application designed to facilitate efficient and consistent annotation of phenotypic similarities and differences using Entity-Quality syntax, drawing on terms from community ontologies for anatomical entities, phenotypic qualities, and taxonomic names. Phenex can be configured to load only those ontologies pertinent to a taxonomic group of interest. The graphical user interface was optimized for evolutionary biologists accustomed to working with lists of taxa, characters, character states, and character-by-taxon matrices.Annotation of phenotypic data using ontologies and globally unique taxonomic identifiers will allow biologists to integrate phenotypic data from different organisms and studies, leveraging decades of work in systematics and comparative morphology.

  18. Phenex: ontological annotation of phenotypic diversity.

    Science.gov (United States)

    Balhoff, James P; Dahdul, Wasila M; Kothari, Cartik R; Lapp, Hilmar; Lundberg, John G; Mabee, Paula; Midford, Peter E; Westerfield, Monte; Vision, Todd J

    2010-05-05

    Phenotypic differences among species have long been systematically itemized and described by biologists in the process of investigating phylogenetic relationships and trait evolution. Traditionally, these descriptions have been expressed in natural language within the context of individual journal publications or monographs. As such, this rich store of phenotype data has been largely unavailable for statistical and computational comparisons across studies or integration with other biological knowledge. Here we describe Phenex, a platform-independent desktop application designed to facilitate efficient and consistent annotation of phenotypic similarities and differences using Entity-Quality syntax, drawing on terms from community ontologies for anatomical entities, phenotypic qualities, and taxonomic names. Phenex can be configured to load only those ontologies pertinent to a taxonomic group of interest. The graphical user interface was optimized for evolutionary biologists accustomed to working with lists of taxa, characters, character states, and character-by-taxon matrices. Annotation of phenotypic data using ontologies and globally unique taxonomic identifiers will allow biologists to integrate phenotypic data from different organisms and studies, leveraging decades of work in systematics and comparative morphology.

  19. Dissecting molecular stress networks: identifying nodes of divergence between life-history phenotypes.

    Science.gov (United States)

    Schwartz, Tonia S; Bronikowski, Anne M

    2013-02-01

    The complex molecular network that underlies physiological stress response is comprised of nodes (proteins, metabolites, mRNAs, etc.) whose connections span cells, tissues and organs. Variable nodes are points in the network upon which natural selection may act. Thus, identifying variable nodes will reveal how this molecular stress network may evolve among populations in different habitats and how it might impact life-history evolution. Here, we use physiological and genetic assays to test whether laboratory-born juveniles from natural populations of garter snakes (Thamnophis elegans), which have diverged in their life-history phenotypes, vary concomitantly at candidate nodes of the stress response network, (i) under unstressed conditions and (ii) in response to an induced stress. We found that two common measures of stress (plasma corticosterone and liver gene expression of heat shock proteins) increased under stress in both life-history phenotypes. In contrast, the phenotypes diverged at four nodes both under unstressed conditions and in response to stress: circulating levels of reactive oxygen species (superoxide, H(2)O(2)); liver gene expression of GPX1 and erythrocyte DNA damage. Additionally, allele frequencies for SOD2 diverge from neutral markers, suggesting diversifying selection on SOD2 alleles. This study supports the hypothesis that these life-history phenotypes have diverged at the molecular level in how they respond to stress, particularly in nodes regulating oxidative stress. Furthermore, the differences between the life-history phenotypes were more pronounced in females. We discuss the responses to stress in the context of the associated life-history phenotype and the evolutionary pressures thought to be responsible for divergence between the phenotypes. © 2012 Blackwell Publishing Ltd.

  20. Epigenetic telomere protection by Drosophila DNA damage response pathways.

    Science.gov (United States)

    Oikemus, Sarah R; Queiroz-Machado, Joana; Lai, KuanJu; McGinnis, Nadine; Sunkel, Claudio; Brodsky, Michael H

    2006-05-01

    Analysis of terminal deletion chromosomes indicates that a sequence-independent mechanism regulates protection of Drosophila telomeres. Mutations in Drosophila DNA damage response genes such as atm/tefu, mre11, or rad50 disrupt telomere protection and localization of the telomere-associated proteins HP1 and HOAP, suggesting that recognition of chromosome ends contributes to telomere protection. However, the partial telomere protection phenotype of these mutations limits the ability to test if they act in the epigenetic telomere protection mechanism. We examined the roles of the Drosophila atm and atr-atrip DNA damage response pathways and the nbs homolog in DNA damage responses and telomere protection. As in other organisms, the atm and atr-atrip pathways act in parallel to promote telomere protection. Cells lacking both pathways exhibit severe defects in telomere protection and fail to localize the protection protein HOAP to telomeres. Drosophila nbs is required for both atm- and atr-dependent DNA damage responses and acts in these pathways during DNA repair. The telomere fusion phenotype of nbs is consistent with defects in each of these activities. Cells defective in both the atm and atr pathways were used to examine if DNA damage response pathways regulate telomere protection without affecting telomere specific sequences. In these cells, chromosome fusion sites retain telomere-specific sequences, demonstrating that loss of these sequences is not responsible for loss of protection. Furthermore, terminally deleted chromosomes also fuse in these cells, directly implicating DNA damage response pathways in the epigenetic protection of telomeres. We propose that recognition of chromosome ends and recruitment of HP1 and HOAP by DNA damage response proteins is essential for the epigenetic protection of Drosophila telomeres. Given the conserved roles of DNA damage response proteins in telomere function, related mechanisms may act at the telomeres of other organisms.

  1. DNA: Structure and function

    DEFF Research Database (Denmark)

    Sinden, Richard R.; E. Pearson, Christopher; N. Potaman, Vladimir

    1998-01-01

    This chapter discusses the structure and function of DNA. DNA occupies a critical role in cells, because it is the source of all intrinsic genetic information. Chemically, DNA is a very stable molecule, a characteristic important for a macromolecule that may have to persist in an intact form...

  2. Replicating animal mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    Emily A. McKinney

    2013-01-01

    Full Text Available The field of mitochondrial DNA (mtDNA replication has been experiencing incredible progress in recent years, and yet little is certain about the mechanism(s used by animal cells to replicate this plasmid-like genome. The long-standing strand-displacement model of mammalian mtDNA replication (for which single-stranded DNA intermediates are a hallmark has been intensively challenged by a new set of data, which suggests that replication proceeds via coupled leading-and lagging-strand synthesis (resembling bacterial genome replication and/or via long stretches of RNA intermediates laid on the mtDNA lagging-strand (the so called RITOLS. The set of proteins required for mtDNA replication is small and includes the catalytic and accessory subunits of DNA polymerase y, the mtDNA helicase Twinkle, the mitochondrial single-stranded DNA-binding protein, and the mitochondrial RNA polymerase (which most likely functions as the mtDNA primase. Mutations in the genes coding for the first three proteins are associated with human diseases and premature aging, justifying the research interest in the genetic, biochemical and structural properties of the mtDNA replication machinery. Here we summarize these properties and discuss the current models of mtDNA replication in animal cells.

  3. Sensitive periods in epigenetics: bringing us closer to complex behavioral phenotypes.

    Science.gov (United States)

    Nagy, Corina; Turecki, Gustavo

    2012-08-01

    Genetic studies have attempted to elucidate causal mechanisms for the development of complex disease, but genome-wide associations have been largely unsuccessful in establishing these links. As an alternative link between genes and disease, recent efforts have focused on mechanisms that alter the function of genes without altering the underlying DNA sequence. Known as epigenetic mechanisms, these include DNA methylation, chromatin conformational changes through histone modifications, ncRNAs and, most recently, 5-hydroxymethylcytosine. Although DNA methylation is involved in normal development, aging and gene regulation, altered methylation patterns have been associated with disease. It is generally believed that early life constitutes a period during which there is increased sensitivity to the regulatory effects of epigenetic mechanisms. The purpose of this review is to outline the contribution of epigenetic mechanisms to genomic function, particularly in the development of complex behavioral phenotypes, focusing on the sensitive periods.

  4. Correlation of iris biometrics and DNA

    DEFF Research Database (Denmark)

    Harder, Stine; Clemmensen, Line Katrine Harder; Dahl, Anders Bjorholm

    2013-01-01

    The presented work concerns prediction of complex human phenotypes from genotypes. We were interested in correlating iris color and texture with DNA. Our data consist of 212 eye images along with DNA: 32 single-nucleotide polymorphisms (SNPs). We used two types of biometrics to describe the eye...... images: One for iris color and one for iris texture. Both biometrics were high dimensional and a sparse principle component analysis (SPCA) reduced the dimensions and resulted in a representation of data with good interpretability. The correlations between the sparse principal components (SPCs......) and the 32 SNPs were found using a canonical correlation analysis (CCA). The result was a single significant canonical correlation (CC) for both biometrics. Each CC comprised two correlated canonical variables, consisting of a linear combination of SPCs and a linear combination of SNPs, respectively...

  5. Targeting DNA Methyltranferases in Urological Tumors

    Directory of Open Access Journals (Sweden)

    Ângela Marques-Magalhães

    2018-04-01

    Full Text Available Urological cancers are a heterogeneous group of malignancies accounting for a considerable proportion of cancer-related morbidity and mortality worldwide. Aberrant epigenetic traits, especially altered DNA methylation patterns constitute a hallmark of these tumors. Nonetheless, these alterations are reversible, and several efforts have been carried out to design and test several epigenetic compounds that might reprogram tumor cell phenotype back to a normal state. Indeed, several DNMT inhibitors are currently under evaluation for therapeutic efficacy in clinical trials. This review highlights the critical role of DNA methylation in urological cancers and summarizes the available data on pre-clinical assays and clinical trials with DNMT inhibitors in bladder, kidney, prostate, and testicular germ cell cancers.

  6. Mitochondrial DNA Copy Number in Sleep Duration Discordant Monozygotic Twins.

    Science.gov (United States)

    Wrede, Joanna E; Mengel-From, Jonas; Buchwald, Dedra; Vitiello, Michael V; Bamshad, Michael; Noonan, Carolyn; Christiansen, Lene; Christensen, Kaare; Watson, Nathaniel F

    2015-10-01

    Mitochondrial DNA (mtDNA) copy number is an important component of mitochondrial function and varies with age, disease, and environmental factors. We aimed to determine whether mtDNA copy number varies with habitual differences in sleep duration within pairs of monozygotic twins. Academic clinical research center. 15 sleep duration discordant monozygotic twin pairs (30 twins, 80% female; mean age 42.1 years [SD 15.0]). Sleep duration was phenotyped with wrist actigraphy. Each twin pair included a "normal" (7-9 h/24) and "short" (sleeping twin. Fasting peripheral blood leukocyte DNA was assessed for mtDNA copy number via the n-fold difference between qPCR measured mtDNA and nuclear DNA creating an mtDNA measure without absolute units. We used generalized estimating equation linear regression models accounting for the correlated data structure to assess within-pair effects of sleep duration on mtDNA copy number. Mean within-pair sleep duration difference per 24 hours was 94.3 minutes (SD 62.6 min). We found reduced sleep duration (β = 0.06; 95% CI 0.004, 0.12; P sleep efficiency (β = 0.51; 95% CI 0.06, 0.95; P sleep duration was associated with a decrease in mtDNA copy number of 0.06. Likewise, a 1% decrease in actigraphy-defined sleep efficiency was associated with a decrease in mtDNA copy number of 0.51. Reduced sleep duration and sleep efficiency were associated with reduced mitochondrial DNA copy number in sleep duration discordant monozygotic twins offering a potential mechanism whereby short sleep impairs health and longevity through mitochondrial stress. © 2015 Associated Professional Sleep Societies, LLC.

  7. Genotypes and clinical phenotypes in children with cytochrome-c oxidase deficiency.

    Science.gov (United States)

    Darin, N; Moslemi, A-R; Lebon, S; Rustin, P; Holme, E; Oldfors, A; Tulinius, M

    2003-12-01

    Cytochrome c oxidase (COX) deficiency has been associated with a wide spectrum of clinical features and may be caused by mutations in different genes of both the mitochondrial and the nuclear DNA. In an attempt to correlate the clinical phenotype with the genotype in 16 childhood cases, mtDNA was analysed for deletion, depletion, and mutations in the three genes encoding COX subunits and the 22 tRNA genes. Furthermore, nuclear DNA was analysed for mutations in the SURF1, SCO2, COX10, and COX17 genes and cases with mtDNA depletion were analysed for mutations in the TK2 gene. SURF1-mutations were identified in three out of four cases with Leigh syndrome while a mutation in the mitochondrial tRNA (trp) gene was identified in the fourth. One case with mtDNA depletion had mutations in the TK2 gene. In two cases with leukoencephalopathy, one case with encephalopathy, five cases with fatal infantile myopathy and cardiomyopathy, two cases with benign infantile myopathy, and one case with mtDNA depletion, no mutations were identified. We conclude that COX deficiency in childhood should be suspected in a wide range of clinical settings and although an increasing number of genetic defects have been identified, the underlying mutations remain unclear in the majority of the cases.

  8. Identity of the xerophilic species Aspergillus penicillioides: Integrated analysis of the genotypic and phenotypic characters.

    Science.gov (United States)

    Tamura, Miki; Kawasaki, Hiroko; Sugiyama, Junta

    1999-02-01

    We examined the identity of Aspergillus penicillioides, the typical xerophilic and strictly anamorphic species, using an integrated analysis of the genotypic and phenotypic characters. Our experimental methods on two genotypic characters, i.e., DNA base composition using the HPLC method and DNA relatedness using the nitrocellulose filter hybridization technique between A. flavus, A. oryzae, and their close relations revealed a good agreement with the values by buoyant density (for DNA base composition) and spectrophotometric determination (for DNA relatedness) reported by Kurtzman et al. in 1986. On the basis of these comparisons, we examined DNA base composition and DNA relatedness of six selected strains of A. penicillioides, including IFO 8155 (originally described as A. vitricola), one strain of A. restrictus, and the respective strains from Eurotium amstelodami, E. repens, and E. rubrum. As a result, five strains within A. penicillioides, including the neotype strain NRRL 4548, had G+C contents of 46 to 49 mol%, whereas IFO 8155 had 50 mol%. A. restrictus had 52 mol%, and three Eurotium species ranged from 46 to 49 mol%. The DNA relatedness between A. penicillioides (five strains), except for IFO 8155, exhibited values greater than 70%, but the DNA complementarity between four strains and IFO 8155 in A. penicillioides revealed values of less than 40%. DNA relatedness values between three species of Eurotium were 65 to 72%. We determined 18S, 5.8S, and ITS rDNA sequences as other genotypic characters from A. penicillioides (six strains), A. restrictus, and related teleomorphic species of Eurotium. In three phylogenetic trees inferred from these sequences, five strains of A. penicillioides, including the neotype strain, were closely related to each other, whereas IFO 8155 was distantly related and grouped with other xerophilic species. Our results have suggested that A. penicillioides typified by NRRL 4548 and A. penicillioides IFO 8155 (ex holotype of A

  9. Fast phylogenetic DNA barcoding

    DEFF Research Database (Denmark)

    Terkelsen, Kasper Munch; Boomsma, Wouter Krogh; Willerslev, Eske

    2008-01-01

    We present a heuristic approach to the DNA assignment problem based on phylogenetic inferences using constrained neighbour joining and non-parametric bootstrapping. We show that this method performs as well as the more computationally intensive full Bayesian approach in an analysis of 500 insect...... DNA sequences obtained from GenBank. We also analyse a previously published dataset of environmental DNA sequences from soil from New Zealand and Siberia, and use these data to illustrate the fact that statistical approaches to the DNA assignment problem allow for more appropriate criteria...... for determining the taxonomic level at which a particular DNA sequence can be assigned....

  10. Radiation and DNA

    Energy Technology Data Exchange (ETDEWEB)

    Riabchenko, N I

    1979-01-01

    Consideration is given to the effects of ionizing radiation on the structure of DNA. Physical and chemical methods of determining radiation damage to the primary (polynucleotide chain and nitrogenous base) and secondary (helical) structure of DNA are discussed, and the effects of ionizing radiation on deoxyribonucleoprotein complexes are considered. The radiolysis of DNA in vitro and in bacterial and mammalian cells is examined and cellular mechanisms for the repair of radiation-damaged DNA are considered, taking into account single-strand and double-strand breaks, gamma-radiation damage and deoxyribonucleoprotein-membrane complex damage. Postradiation DNA degradation in bacteria and lymphatic cells is also discussed.

  11. DNA-Mediated Electrochemistry

    Science.gov (United States)

    Gorodetsky, Alon A.; Buzzeo, Marisa C.

    2009-01-01

    The base pair stack of DNA has been demonstrated as a medium for long range charge transport chemistry both in solution and at DNA-modified surfaces. This chemistry is exquisitely sensitive to structural perturbations in the base pair stack as occur with lesions, single base mismatches, and protein binding. We have exploited this sensitivity for the development of reliable electrochemical assays based on DNA charge transport at self-assembled DNA monolayers. Here we discuss the characteristic features, applications, and advantages of DNA-mediated electrochemistry. PMID:18980370

  12. DNA methyltransferase expressions in Japanese rice fish (Oryzias latipes) embryogenesis is developmentally regulated and modulated by ethanol and 5-azacytidine

    Science.gov (United States)

    We aimed to investigate the impact of the epigenome in inducting fetal alcohol spectrum disorder (FASD) phenotypes in Japanese rice fish embryogenesis. One of the significant events in epigenome is DNA methylation which is catalyzed by DNA methyl transferase (DNMT) enzymes. We analyzed DNMT enzyme m...

  13. Modulation of DNA methylation machineries in japanese rice fish (Oryzias latipes) embryogenesis by ethanol and 5-azacytidine

    Science.gov (United States)

    As a sequel of our investigations on the impact of epigenome in inducing fetal alcohol spectrum disorder (FASD) phenotypes in Japanese rice fish, we investigated on several DNA methylation machinery genes including DNA methyl transferase 3ba (dnmt3ba) and methyl binding proteins (MBPs), namely, mbdl...

  14. Mitochondrial DNA mutation load in a family with the m.8344A>G point mutation and lipomas

    DEFF Research Database (Denmark)

    Jeppesen, Tina Dysgaard; Al-Hashimi, Noor; Duno, Morten

    2017-01-01

    Studies have shown that difference in mtDNA mutation load among tissues is a result of postnatal modification. We present five family members with the m.8344A>G with variable phenotypes but uniform intrapersonal distribution of mutation load, indicating that there is no postnatal modification of mt......DNA mutation load in this genotype....

  15. Age is associated with asthma phenotypes.

    Science.gov (United States)

    Ponte, Eduardo V; Lima, Aline; Almeida, Paula C A; de Jesus, Juliana P V; Lima, Valmar B; Scichilone, Nicola; Souza-Machado, Adelmir; Cruz, Álvaro A

    2017-11-01

    The relationship between age and asthma phenotypes is important as population is ageing, asthma is becoming common in older ages and recently developed treatments for asthma are guided by phenotypes. The aim of this study is to evaluate whether age is associated with specific asthma phenotypes. This is a cross-sectional study. We included subjects with asthma of varied degrees of severity. Subjects underwent spirometry, skin prick test to aeroallergens, answered the Asthma Control Questionnaire and had blood samples collected. We performed binary logistic regression analysis to evaluate whether age is associated with asthma phenotypes. We enrolled 868 subjects. In comparison with subjects ≤ 40 years, older subjects had high odds of irreversible airway obstruction (from 41 to 64 years, OR: 1.83 (95% CI: 1.32-2.54); ≥65 years, OR: 3.45 (2.12-5.60)) and severe asthma phenotypes (from 41 to 64 years, OR: 3.23 (2.26-4.62); ≥65 years, OR: 4.55 (2.39-8.67)). Older subjects had low odds of atopic (from 41 to 64 years, OR: 0.56 (0.39-0.79); ≥65 years, OR: 0.47 (0.27-0.84)) and eosinophilic phenotypes (from 41 to 64 years, OR: 0.63 (0.46-0.84); ≥65 years, OR: 0.39 (0.24-0.64)). Older subjects with asthma have low odds of atopic and eosinophilic phenotypes, whereas they present high odds of irreversible airway obstruction and severe asthma. © 2017 Asian Pacific Society of Respirology.

  16. DAF-16/FOXO and EGL-27/GATA promote developmental growth in response to persistent somatic DNA damage.

    Science.gov (United States)

    Mueller, Michael M; Castells-Roca, Laia; Babu, Vipin; Ermolaeva, Maria A; Müller, Roman-Ulrich; Frommolt, Peter; Williams, Ashley B; Greiss, Sebastian; Schneider, Jennifer I; Benzing, Thomas; Schermer, Bernhard; Schumacher, Björn

    2014-12-01

    Genome maintenance defects cause complex disease phenotypes characterized by developmental failure, cancer susceptibility and premature ageing. It remains poorly understood how DNA damage responses function during organismal development and maintain tissue functionality when DNA damage accumulates with ageing. Here we show that the FOXO transcription factor DAF-16 is activated in response to DNA damage during development, whereas the DNA damage responsiveness of DAF-16 declines with ageing. We find that in contrast to its established role in mediating starvation arrest, DAF-16 alleviates DNA-damage-induced developmental arrest and even in the absence of DNA repair promotes developmental growth and enhances somatic tissue functionality. We demonstrate that the GATA transcription factor EGL-27 co-regulates DAF-16 target genes in response to DNA damage and together with DAF-16 promotes developmental growth. We propose that EGL-27/GATA activity specifies DAF-16-mediated DNA damage responses to enable developmental progression and to prolong tissue functioning when DNA damage persists.

  17. DNA fragmentation in spermatozoa

    DEFF Research Database (Denmark)

    Rex, A S; Aagaard, J.; Fedder, J

    2017-01-01

    Sperm DNA Fragmentation has been extensively studied for more than a decade. In the 1940s the uniqueness of the spermatozoa protein complex which stabilizes the DNA was discovered. In the fifties and sixties, the association between unstable chromatin structure and subfertility was investigated....... In the seventies, the impact of induced DNA damage was investigated. In the 1980s the concept of sperm DNA fragmentation as related to infertility was introduced as well as the first DNA fragmentation test: the Sperm Chromatin Structure Assay (SCSA). The terminal deoxynucleotidyl transferase nick end labelling...... (TUNEL) test followed by others was introduced in the nineties. The association between DNA fragmentation in spermatozoa and pregnancy loss has been extensively investigated spurring the need for a therapeutic tool for these patients. This gave rise to an increased interest in the aetiology of DNA damage...

  18. Biophysics of DNA

    CERN Document Server

    Vologodskii, Alexander

    2015-01-01

    Surveying the last sixty years of research, this book describes the physical properties of DNA in the context of its biological functioning. It is designed to enable both students and researchers of molecular biology, biochemistry and physics to better understand the biophysics of DNA, addressing key questions and facilitating further research. The chapters integrate theoretical and experimental approaches, emphasising throughout the importance of a quantitative knowledge of physical properties in building and analysing models of DNA functioning. For example, the book shows how the relationship between DNA mechanical properties and the sequence specificity of DNA-protein binding can be analyzed quantitatively by using our current knowledge of the physical and structural properties of DNA. Theoretical models and experimental methods in the field are critically considered to enable the reader to engage effectively with the current scientific literature on the physical properties of DNA.

  19. Epigenetics in Alzheimer's Disease: Perspective of DNA Methylation.

    Science.gov (United States)

    Qazi, Talal Jamil; Quan, Zhenzhen; Mir, Asif; Qing, Hong

    2018-02-01

    Research over the years has shown that causes of Alzheimer's disease are not well understood, but over the past years, the involvement of epigenetic mechanisms in the developing memory formation either under pathological or physiological conditions has become clear. The term epigenetics represents the heredity of changes in phenotype that are independent of altered DNA sequences. Different studies validated that cytosine methylation of genomic DNA decreases with age in different tissues of mammals, and therefore, the role of epigenetic factors in developing neurological disorders in aging has been under focus. In this review, we summarized and reviewed the involvement of different epigenetic mechanisms especially the DNA methylation in Alzheimer's disease (AD), late-onset Alzheimer's disease (LOAD), familial Alzheimer's disease (FAD), and autosomal dominant Alzheimer's disease (ADAD). Down to the minutest of details, we tried to discuss the methylation patterns like mitochondrial DNA methylation and ribosomal DNA (rDNA) methylation. Additionally, we mentioned some therapeutic approaches related to epigenetics, which could provide a potential cure for AD. Moreover, we reviewed some recent studies that validate DNA methylation as a potential biomarker and its role in AD. We hope that this review will provide new insights into the understanding of AD pathogenesis from the epigenetic perspective especially from the perspective of DNA methylation.

  20. Aging of hematopoietic stem cells: DNA damage and mutations?

    Science.gov (United States)

    Moehrle, Bettina M; Geiger, Hartmut

    2016-10-01

    Aging in the hematopoietic system and the stem cell niche contributes to aging-associated phenotypes of hematopoietic stem cells (HSCs), including leukemia and aging-associated immune remodeling. Among others, the DNA damage theory of aging of HSCs is well established, based on the detection of a significantly larger amount of γH2AX foci and a higher tail moment in the comet assay, both initially thought to be associated with DNA damage in aged HSCs compared with young cells, and bone marrow failure in animals devoid of DNA repair factors. Novel data on the increase in and nature of DNA mutations in the hematopoietic system with age, the quality of the DNA damage response in aged HSCs, and the nature of γH2AX foci question a direct link between DNA damage and the DNA damage response and aging of HSCs, and rather favor changes in epigenetics, splicing-factors or three-dimensional architecture of the cell as major cell intrinsic factors of HSCs aging. Aging of HSCs is also driven by a strong contribution of aging of the niche. This review discusses the DNA damage theory of HSC aging in the light of these novel mechanisms of aging of HSCs. Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

  1. DNA repair is indispensable for survival after acute inflammation

    Science.gov (United States)

    Calvo, Jennifer A.; Meira, Lisiane B.; Lee, Chun-Yue I.; Moroski-Erkul, Catherine A.; Abolhassani, Nona; Taghizadeh, Koli; Eichinger, Lindsey W.; Muthupalani, Sureshkumar; Nordstrand, Line M.; Klungland, Arne; Samson, Leona D.

    2012-01-01

    More than 15% of cancer deaths worldwide are associated with underlying infections or inflammatory conditions, therefore understanding how inflammation contributes to cancer etiology is important for both cancer prevention and treatment. Inflamed tissues are known to harbor elevated etheno-base (ε-base) DNA lesions induced by the lipid peroxidation that is stimulated by reactive oxygen and nitrogen species (RONS) released from activated neutrophils and macrophages. Inflammation contributes to carcinogenesis in part via RONS-induced cytotoxic and mutagenic DNA lesions, including ε-base lesions. The mouse alkyl adenine DNA glycosylase (AAG, also known as MPG) recognizes such base lesions, thus protecting against inflammation-associated colon cancer. Two other DNA repair enzymes are known to repair ε-base lesions, namely ALKBH2 and ALKBH3; thus, we sought to determine whether these DNA dioxygenase enzymes could protect against chronic inflammation-mediated colon carcinogenesis. Using established chemically induced colitis and colon cancer models in mice, we show here that ALKBH2 and ALKBH3 provide cancer protection similar to that of the DNA glycosylase AAG. Moreover, Alkbh2 and Alkbh3 each display apparent epistasis with Aag. Surprisingly, deficiency in all 3 DNA repair enzymes confers a massively synergistic phenotype, such that animals lacking all 3 DNA repair enzymes cannot survive even a single bout of chemically induced colitis. PMID:22684101

  2. Ancient mtDNA genetic variants modulate mtDNA transcription and replication.

    Directory of Open Access Journals (Sweden)

    Sarit Suissa

    2009-05-01

    Full Text Available Although the functional consequences of mitochondrial DNA (mtDNA genetic backgrounds (haplotypes, haplogroups have been demonstrated by both disease association studies and cell culture experiments, it is not clear which of the mutations within the haplogroup carry functional implications and which are "evolutionary silent hitchhikers". We set forth to study the functionality of haplogroup-defining mutations within the mtDNA transcription/replication regulatory region by in vitro transcription, hypothesizing that haplogroup-defining mutations occurring within regulatory motifs of mtDNA could affect these processes. We thus screened >2500 complete human mtDNAs representing all major populations worldwide for natural variation in experimentally established protein binding sites and regulatory regions comprising a total of 241 bp in each mtDNA. Our screen revealed 77/241 sites showing point mutations that could be divided into non-fixed (57/77, 74% and haplogroup/sub-haplogroup-defining changes (i.e., population fixed changes, 20/77, 26%. The variant defining Caucasian haplogroup J (C295T increased the binding of TFAM (Electro Mobility Shift Assay and the capacity of in vitro L-strand transcription, especially of a shorter transcript that maps immediately upstream of conserved sequence block 1 (CSB1, a region associated with RNA priming of mtDNA replication. Consistent with this finding, cybrids (i.e., cells sharing the same nuclear genetic background but differing in their mtDNA backgrounds harboring haplogroup J mtDNA had a >2 fold increase in mtDNA copy number, as compared to cybrids containing haplogroup H, with no apparent differences in steady state levels of mtDNA-encoded transcripts. Hence, a haplogroup J regulatory region mutation affects mtDNA replication or stability, which may partially account for the phenotypic impact of this haplogroup. Our analysis thus demonstrates, for the first time, the functional impact of particular mtDNA

  3. International Symposium on Positive Strand RNA Viruses (2nd) Held in Vienna, Austria on June 26-30, 1989. Abstracts

    Science.gov (United States)

    1989-07-01

    DEAE dextran-treated chicken embryo Iosdr specific probe hybridised in -olont bluis to a fiibroblasts (CEF). VEE antigens were demonstrated in 1,1...are protein dimers but show differences reflects the destiny of the protein. The with respect to molecular organization as effect of sorting sequences...exchange and virus directly from virusinfected chick embryo homogena- had lost their capacity to bind the tes, which is the source of virus used for

  4. Ultraviolet light-resistant primary transfectants of xeroderma pigmentosum cells are also DNA repair-proficient

    International Nuclear Information System (INIS)

    Stark, M.; Naiman, T.; Canaani, D.

    1989-01-01

    In a previous work, an immortal xeroderma pigmentosum cell line belonging to complementation group C was complemented to a UV-resistant phenotype by transfection with a human cDNA clone library. We now report that the primary transformants selected for UV-resistance also acquired normal levels of DNA repair. This was assessed both by measurement of UV-induced [ 3 H]thymidine incorporation and by equilibrium sedimentation analysis of repair-DNA synthesis. Therefore, the transduced DNA element which confers normal UV-resistance also corrects the excision repair defect of the xeroderma pigmentosum group C cell line

  5. DNA Methylation: An Epigenetic Risk Factor in Preterm Birth

    Science.gov (United States)

    Menon, Ramkumar; Conneely, Karen N.; Smith, Alicia K.

    2012-01-01

    Spontaneous preterm birth (PTB; birth prior to 37 weeks of gestation) is a complex phenotype with multiple risk factors that complicate our understanding of its etiology. A number of recent studies have supported the hypothesis that epigenetic modifications such as DNA methylation induced by pregnancy-related risk factors may influence the risk of PTB or result in changes that predispose a neonate to adult-onset diseases. The critical role of timing of gene expression in the etiology of PTB makes it a highly relevant disorder in which to examine the potential role of epigenetic changes. Because changes in DNA methylation patterns can result in long-term consequences, it is of critical interest to identify the epigenetic patterns associated with adverse pregnancy outcomes. This review examines the potential role of DNA methylation as a risk factor for PTB and discusses several issues and limitations that should be considered when planning DNA methylation studies. PMID:22228737

  6. Overeating phenotypes in overweight and obese children.

    Science.gov (United States)

    Boutelle, Kerri N; Peterson, Carol B; Crosby, Ross D; Rydell, Sarah A; Zucker, Nancy; Harnack, Lisa

    2014-05-01

    The purpose of this study was to identify overeating phenotypes and their correlates in overweight and obese children. One hundred and seventeen treatment-seeking overweight and obese 8-12year-old children and their parents completed the study. Children completed an eating in the absence of hunger (EAH) paradigm, the Eating Disorder Examination interview, and measurements of height and weight. Parents and children completed questionnaires that evaluated satiety responsiveness, food responsiveness, negative affect eating, external eating and eating in the absence of hunger. Latent profile analysis was used to identify heterogeneity in overeating phenotypes in the child participants. Latent classes were then compared on measures of demographics, obesity status and nutritional intake. Three latent classes of overweight and obese children were identified: High Satiety Responsive, High Food Responsive, and Moderate Satiety and Food Responsive. Results indicated that the High Food Responsive group had higher BMI and BMI-Z scores compared to the High Satiety Responsive group. No differences were found among classes in demographics or nutritional intake. This study identified three overeating phenotypes, supporting the heterogeneity of eating patterns associated with overweight and obesity in treatment-seeking children. These finding suggest that these phenotypes can potentially be used to identify high risk groups, inform prevention and intervention targets, and develop specific treatments for these behavioral phenotypes. Copyright © 2014. Published by Elsevier Ltd.

  7. Overproduction of single-stranded-DNA-binding protein specifically inhibits recombination of UV-irradiated bacteriophage DNA in Escherichia coli

    International Nuclear Information System (INIS)

    Moreau, P.L.

    1988-01-01

    Overproduction of single-stranded DNA (ssDNA)-binding protein (SSB) in uvr Escherichia coli mutants results in a wide range of altered phenotypes. (i) Cell survival after UV irradiation is decreased; (ii) expression of the recA-lexA regulon is slightly reduced after UV irradiation, whereas it is increased without irradiation; and (iii) recombination of UV-damaged lambda DNA is inhibited, whereas recombination of nonirradiated DNA is unaffected. These results are consistent with the idea that in UV-damaged bacteria, SSB is first required to allow the formation of short complexes of RecA protein and ssDNA that mediate cleavage of the LexA protein. However, in a second stage, SSB should be displaced from ssDNA to permit the production of longer RecA-ssDNA nucleoprotein filaments that are required for strand pairing and, hence, recombinational repair. Since bacteria overproducing SSB appear identical in physiological respects to recF mutant bacteria, it is suggested that the RecF protein (alone or with other proteins of the RecF pathway) may help RecA protein to release SSB from ssDNA

  8. Introgression of mitochondrial DNA among Myodes voles: consequences for energetics?

    Directory of Open Access Journals (Sweden)

    Boratyński Zbyszek

    2011-12-01

    Full Text Available Abstract Background Introgression of mitochondrial DNA (mtDNA is among the most frequently described cases of reticulate evolution. The tendency of mtDNA to cross interspecific barriers is somewhat counter-intuitive considering the key function of enzymes that it encodes in the oxidative-phosphorylation process, which could give rise to hybrid dysfunction. How mtDNA reticulation affects the evolution of metabolic functions is, however, uncertain. Here we investigated how morpho-physiological traits vary in natural populations of a common rodent (the bank vole, Myodes glareolus and whether this variation could be associated with mtDNA introgression. First, we confirmed that M. glareolus harbour mtDNA introgressed from M. rutilus by analyzing mtDNA (cytochrome b, 954 bp and nuclear DNA (four markers; 2333 bp in total sequence variation and reconstructing loci phylogenies among six natural populations in Finland. We then studied geographic variation in body size and basal metabolic rate (BMR among the populations of M. glareolus and tested its relationship with mtDNA type. Results Myodes glareolus and its arctic neighbour, M. rutilus, are reciprocally monophyletic at the analyzed nuclear DNA loci. In contrast, the two northernmost populations of M. glareolus have a fixed mitotype that is shared with M. rutilus, likely due to introgressive hybridization. The analyses of phenotypic traits revealed that the body mass and whole-body, but not mass corrected, BMR are significantly reduced in M. glareolus females from northern Finland that also have the introgressed mitotype. Restricting the analysis to the single population where the mitotypes coexist, the association of mtDNA type with whole-body BMR remained but those with mass corrected BMR and body mass did not. Mitochondrial sequence variation in the introgressed haplotypes is compatible with demographic growth of the populations, but may also be a result of positive selection. Conclusion Our

  9. Reversion of mtDNA depletion in a patient with TK2 deficiency.

    Science.gov (United States)

    Vilà, M R; Segovia-Silvestre, T; Gámez, J; Marina, A; Naini, A B; Meseguer, A; Lombès, A; Bonilla, E; DiMauro, S; Hirano, M; Andreu, A L

    2003-04-08

    Mutations in the thymidine kinase 2 (TK2) gene cause a myopathic form of the mitochondrial DNA depletion syndrome (MDS). Here, the authors report the unusual clinical, biochemical, and molecular findings in a 14-year-old patient in whom pathogenic mutations were identified in the TK2 gene. This report extends the phenotypic expression of primary TK2 deficiency and suggests that factors other than TK2 may modify expression of the clinical phenotype in patients with MDS syndrome.

  10. Genome-Wide Prediction of DNA Methylation Using DNA Composition and Sequence Complexity in Human.

    Science.gov (United States)

    Wu, Chengchao; Yao, Shixin; Li, Xinghao; Chen, Chujia; Hu, Xuehai

    2017-02-16

    DNA methylation plays a significant role in transcriptional regulation by repressing activity. Change of the DNA methylation level is an important factor affecting the expression of target genes and downstream phenotypes. Because current experimental technologies can only assay a small proportion of CpG sites in the human genome, it is urgent to develop reliable computational models for predicting genome-wide DNA methylation. Here, we proposed a novel algorithm that accurately extracted sequence complexity features (seven features) and developed a support-vector-machine-based prediction model with integration of the reported DNA composition features (trinucleotide frequency and GC content, 65 features) by utilizing the methylation profiles of embryonic stem cells in human. The prediction results from 22 human chromosomes with size-varied windows showed that the 600-bp window achieved the best average accuracy of 94.7%. Moreover, comparisons with two existing methods further showed the superiority of our model, and cross-species predictions on mouse data also demonstrated that our model has certain generalization ability. Finally, a statistical test of the experimental data and the predicted data on functional regions annotated by ChromHMM found that six out of 10 regions were consistent, which implies reliable prediction of unassayed CpG sites. Accordingly, we believe that our novel model will be useful and reliable in predicting DNA methylation.

  11. Step-wise and punctuated genome evolution drive phenotype changes of tumor cells

    International Nuclear Information System (INIS)

    Stepanenko, Aleksei; Andreieva, Svitlana; Korets, Kateryna; Mykytenko, Dmytro; Huleyuk, Nataliya; Vassetzky, Yegor; Kavsan, Vadym

    2015-01-01

    Highlights: • There are the step-wise continuous and punctuated phases of cancer genome evolution. • The system stresses during the different phases may lead to very different responses. • Stable transfection of an empty vector can result in genome and phenotype changes. • Functions of a (trans)gene can be opposite/versatile in cells with different genomes. • Contextually, temozolomide can both promote and suppress tumor cell aggressiveness. - Abstract: The pattern of genome evolution can be divided into two phases: the step-wise continuous phase (step-wise clonal evolution, stable dominant clonal chromosome aberrations (CCAs), and low frequency of non-CCAs, NCCAs) and punctuated phase (marked by elevated NCCAs and transitional CCAs). Depending on the phase, system stresses (the diverse CIN promoting factors) may lead to the very different phenotype responses. To address the contribution of chromosome instability (CIN) to phenotype changes of tumor cells, we characterized CCAs/NCCAs of HeLa and HEK293 cells, and their derivatives after genotoxic stresses (a stable plasmid transfection, ectopic expression of cancer-associated CHI3L1 gene or treatment with temozolomide) by conventional cytogenetics, copy number alterations (CNAs) by array comparative genome hybridization, and phenotype changes by cell viability and soft agar assays. Transfection of either the empty vector pcDNA3.1 or pcDNA3.1-CHI3L1 into 293 cells initiated the punctuated genome changes. In contrast, HeLa-CHI3L1 cells demonstrated the step-wise genome changes. Increased CIN correlated with lower viability of 293-pcDNA3.1 cells but higher colony formation efficiency (CFE). Artificial CHI3L1 production in 293-CHI3L1 cells increased viability and further contributed to CFE. The opposite growth characteristics of 293-CHI3L1 and HeLa-CHI3L1 cells were revealed. The effect and function of a (trans)gene can be opposite and versatile in cells with different genetic network, which is defined by

  12. Step-wise and punctuated genome evolution drive phenotype changes of tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Stepanenko, Aleksei, E-mail: a.a.stepanenko@gmail.com [Department of Biosynthesis of Nucleic Acids, Institute of Molecular Biology and Genetics, National Academy of Sciences of Ukraine, Kyiv 03680 (Ukraine); Andreieva, Svitlana; Korets, Kateryna; Mykytenko, Dmytro [Department of Biosynthesis of Nucleic Acids, Institute of Molecular Biology and Genetics, National Academy of Sciences of Ukraine, Kyiv 03680 (Ukraine); Huleyuk, Nataliya [Institute of Hereditary Pathology, National Academy of Medical Sciences of Ukraine, Lviv 79008 (Ukraine); Vassetzky, Yegor [CNRS UMR8126, Université Paris-Sud 11, Institut de Cancérologie Gustave Roussy, Villejuif 94805 (France); Kavsan, Vadym [Department of Biosynthesis of Nucleic Acids, Institute of Molecular Biology and Genetics, National Academy of Sciences of Ukraine, Kyiv 03680 (Ukraine)

    2015-01-15

    Highlights: • There are the step-wise continuous and punctuated phases of cancer genome evolution. • The system stresses during the different phases may lead to very different responses. • Stable transfection of an empty vector can result in genome and phenotype changes. • Functions of a (trans)gene can be opposite/versatile in cells with different genomes. • Contextually, temozolomide can both promote and suppress tumor cell aggressiveness. - Abstract: The pattern of genome evolution can be divided into two phases: the step-wise continuous phase (step-wise clonal evolution, stable dominant clonal chromosome aberrations (CCAs), and low frequency of non-CCAs, NCCAs) and punctuated phase (marked by elevated NCCAs and transitional CCAs). Depending on the phase, system stresses (the diverse CIN promoting factors) may lead to the very different phenotype responses. To address the contribution of chromosome instability (CIN) to phenotype changes of tumor cells, we characterized CCAs/NCCAs of HeLa and HEK293 cells, and their derivatives after genotoxic stresses (a stable plasmid transfection, ectopic expression of cancer-associated CHI3L1 gene or treatment with temozolomide) by conventional cytogenetics, copy number alterations (CNAs) by array comparative genome hybridization, and phenotype changes by cell viability and soft agar assays. Transfection of either the empty vector pcDNA3.1 or pcDNA3.1-CHI3L1 into 293 cells initiated the punctuated genome changes. In contrast, HeLa-CHI3L1 cells demonstrated the step-wise genome changes. Increased CIN correlated with lower viability of 293-pcDNA3.1 cells but higher colony formation efficiency (CFE). Artificial CHI3L1 production in 293-CHI3L1 cells increased viability and further contributed to CFE. The opposite growth characteristics of 293-CHI3L1 and HeLa-CHI3L1 cells were revealed. The effect and function of a (trans)gene can be opposite and versatile in cells with different genetic network, which is defined by

  13. Constraints on the evolution of phenotypic plasticity

    DEFF Research Database (Denmark)

    Murren, Courtney J; Auld, Josh R.; Callahan, Hilary S

    2015-01-01

    Phenotypic plasticity is ubiquitous and generally regarded as a key mechanism for enabling organisms to survive in the face of environmental change. Because no organism is infinitely or ideally plastic, theory suggests that there must be limits (for example, the lack of ability to produce...... an optimal trait) to the evolution of phenotypic plasticity, or that plasticity may have inherent significant costs. Yet numerous experimental studies have not detected widespread costs. Explicitly differentiating plasticity costs from phenotype costs, we re-evaluate fundamental questions of the limits...... to the evolution of plasticity and of generalists vs specialists. We advocate for the view that relaxed selection and variable selection intensities are likely more important constraints to the evolution of plasticity than the costs of plasticity. Some forms of plasticity, such as learning, may be inherently...

  14. Phenotype Development in Adolescents With Tourette Syndrome

    DEFF Research Database (Denmark)

    Groth, Camilla; Debes, Nanette Mol; Skov, Liselotte

    2017-01-01

    Tourette syndrome (TS) is a neurodevelopmental disorder characterized by frequent comorbidities and a wide spectrum of phenotype presentations. This study aimed to describe the development of phenotypes in TS and tic-related impairment in a large longitudinal study of 226 children and adolescents...... followed up after 6 years. The participants were clinically examined to assess tic severity and impairment, obsessive compulsive disorder (OCD), and attention-deficit/hyperactivity disorder (ADHD). The development in phenotypes changed toward less comorbidity with 40% TS-only (no OCD or ADHD) (TS without...... OCD or ADHD) at baseline and 55% at follow-up.Tic-related impairment was expected to improve with an age-related tic decline, but surprisingly the impairment score did not reflect the tic decline. Sex, vocal and motor tics, and OCD and ADHD severity were highly significantly correlated...

  15. Delineating the GRIN1 phenotypic spectrum

    DEFF Research Database (Denmark)

    Lemke, Johannes R; Geider, Kirsten; Helbig, Katherine L

    2016-01-01

    consequences of GRIN1 mutations were investigated in Xenopus laevis oocytes. RESULTS: We identified heterozygous de novo GRIN1 mutations in 14 individuals and reviewed the phenotypes of all 9 previously reported patients. These 23 individuals presented with a distinct phenotype of profound developmental delay......, severe intellectual disability with absent speech, muscular hypotonia, hyperkinetic movement disorder, oculogyric crises, cortical blindness, generalized cerebral atrophy, and epilepsy. Mutations cluster within transmembrane segments and result in loss of channel function of varying severity...... impairment as well as oculomotor and movement disorders being discriminating phenotypic features. Loss of NMDA receptor function appears to be the underlying disease mechanism. The identification of both heterozygous and homozygous mutations blurs the borders of dominant and recessive inheritance of GRIN1...

  16. Semi-supervised Learning for Phenotyping Tasks.

    Science.gov (United States)

    Dligach, Dmitriy; Miller, Timothy; Savova, Guergana K

    2015-01-01

    Supervised learning is the dominant approach to automatic electronic health records-based phenotyping, but it is expensive due to the cost of manual chart review. Semi-supervised learning takes advantage of both scarce labeled and plentiful unlabeled data. In this work, we study a family of semi-supervised learning algorithms based on Expectation Maximization (EM) in the context of several phenotyping tasks. We first experiment with the basic EM algorithm. When the modeling assumptions are violated, basic EM leads to inaccurate parameter estimation. Augmented EM attenuates this shortcoming by introducing a weighting factor that downweights the unlabeled data. Cross-validation does not always lead to the best setting of the weighting factor and other heuristic methods may be preferred. We show that accurate phenotyping models can be trained with only a few hundred labeled (and a large number of unlabeled) examples, potentially providing substantial savings in the amount of the required manual chart review.

  17. High-resolution analysis of cytosine methylation in ancient DNA.

    Directory of Open Access Journals (Sweden)

    Bastien Llamas

    Full Text Available Epigenetic changes to gene expression can result in heritable phenotypic characteristics that are not encoded in the DNA itself, but rather by biochemical modifications to the DNA or associated chromatin proteins. Interposed between genes and environment, these epigenetic modifications can be influenced by environmental factors to affect phenotype for multiple generations. This raises the possibility that epigenetic states provide a substrate for natural selection, with the potential to participate in the rapid adaptation of species to changes in environment. Any direct test of this hypothesis would require the ability to measure epigenetic states over evolutionary timescales. Here we describe the first single-base resolution of cytosine methylation patterns in an ancient mammalian genome, by bisulphite allelic sequencing of loci from late Pleistocene Bison priscus remains. Retrotransposons and the differentially methylated regions of imprinted loci displayed methylation patterns identical to those derived from fresh bovine tissue, indicating that methylation patterns are preserved in the ancient DNA. Our findings establish the biochemical stability of methylated cytosines over extensive time frames, and provide the first direct evidence that cytosine methylation patterns are retained in DNA from ancient specimens. The ability to resolve cytosine methylation in ancient DNA provides a powerful means to study the role of epigenetics in evolution.

  18. Relationship between endophenotype and phenotype in ADHD

    Directory of Open Access Journals (Sweden)

    Buitelaar Jan K

    2008-01-01

    Full Text Available Abstract Background It has been hypothesized that genetic and environmental factors relate to psychiatric disorders through the effect of intermediating, vulnerability traits called endophenotypes. The study had a threefold aim: to examine the predictive validity of an endophenotypic construct for the ADHD diagnosis, to test whether the magnitude of group differences at the endophenotypic and phenotypic level is comparable, and to investigate whether four factors (gender, age, IQ, rater bias have an effect (moderation or mediation on the relation between endophenotype and phenotype. Methods Ten neurocognitive tasks were administered to 143 children with ADHD, 68 non-affected siblings, and 120 control children (first-borns and 132 children with ADHD, 78 non-affected siblings, and 113 controls (second-borns (5 – 19 years. The task measures have been investigated previously for their endophenotypic viability and were combined to one component which was labeled 'the endophenotypic construct': one measure representative of endophenotypic functioning across several domains of functioning. Results The endophenotypic construct classified children with moderate accuracy (about 50% for each of the three groups. Non-affected children differed as much from controls at the endophenotypic as at the phenotypic level, but affected children displayed a more severe phenotype than endophenotype. Although a potentially moderating effect (age and several mediating effects (gender, age, IQ were found affecting the relation between endophenotypic construct and phenotype, none of the effects studied could account for the finding that affected children had a more severe phenotype than endophenotype. Conclusion Endophenotypic functioning is moderately predictive of the ADHD diagnosis, though findings suggest substantial overlap exists between endophenotypic functioning in the groups of affected children, non-affected siblings, and controls. Results suggest other

  19. Phenotypic Approaches to Drought in Cassava: Review

    Directory of Open Access Journals (Sweden)

    Emmanuel eOkogbenin

    2013-05-01

    Full Text Available Cassava is an important crop in Africa, Asia, Latin America and the Caribbean. Cassava can be produced adequately in drought conditions making it the ideal food security crop in marginal environments. Although cassava can tolerate drought stress, it can be genetically improved to enhance productivity in such environments. Drought adaptation studies in over three decades in cassava have identified relevant mechanisms which have been explored in conventional breeding. Drought is a quantitative trait and its multigenic nature makes it very challenging to effectively manipulate and combine genes in breeding for rapid genetic gain and selection process. Cassava has a long growth cycle of 12 - 18 months which invariably contributes to a long breeding scheme for the crop. Modern breeding using advances in genomics and improved genotyping, is facilitating the dissection and genetic analysis of complex traits including drought tolerance, thus helping to better elucidate and understand the genetic basis of such traits. A beneficial goal of new innovative breeding strategies is to shorten the breeding cycle using minimized, efficient or fast phenotyping protocols. While high throughput genotyping have been achieved, this is rarely the case for phenotyping for drought adaptation. Some of the storage root phenotyping in cassava are often done very late in the evaluation cycle making selection process very slow. This paper highlights some modified traits suitable for early-growth phase phenotyping that may be used to reduce drought phenotyping cycle in cassava. Such modified traits can significantly complement the high throughput genotyping procedures to fast track breeding of improved drought tolerant varieties. The need for metabolite profiling, improved phenomics to take advantage of next generation sequencing technologies and high throughput phenotyping are basic steps for future direction to improve genetic gain and maximize speed for drought tolerance

  20. Beyond DNA repair: DNA-PK function in cancer

    OpenAIRE

    Goodwin, Jonathan F.; Knudsen, Karen E.

    2014-01-01

    The DNA-dependent protein kinase (DNA-PK) is a pivotal component of the DNA repair machinery that governs the response to DNA damage, serving to maintain genome integrity. However, the DNA-PK kinase component was initially isolated with transcriptional complexes, and recent findings have illuminated the impact of DNA-PK-mediated transcriptional regulation on tumor progression and therapeutic response. DNA-PK expression has also been correlated with poor outcome in selected tumor types, furthe...

  1. Global DNA Methylation in the Chestnut Blight Fungus Cryphonectria parasitica and Genome-Wide Changes in DNA Methylation Accompanied with Sectorization

    Directory of Open Access Journals (Sweden)

    Kum-Kang So

    2018-02-01

    Full Text Available Mutation in CpBck1, an ortholog of the cell wall integrity mitogen-activated protein kinase kinase kinase (MAPKKK of Saccharomyces cerevisiae, in the chestnut blight fungus Cryphonectria parasitica resulted in a sporadic sectorization as culture proceeded. The progeny from the sectored area maintained the characteristics of the sector, showing a massive morphogenetic change, including robust mycelial growth without differentiation. Epigenetic changes were investigated as the genetic mechanism underlying this sectorization. Quantification of DNA methylation and whole-genome bisulfite sequencing revealed genome-wide DNA methylation of the wild-type at each nucleotide level and changes in DNA methylation of the sectored progeny. Compared to the wild-type, the sectored progeny exhibited marked genome-wide DNA hypomethylation but increased methylation sites. Expression analysis of two DNA methyltransferases, including two representative types of DNA methyltransferase (DNMTase, demonstrated that both were significantly down-regulated in the sectored progeny. However, functional analysis using mutant phenotypes of corresponding DNMTases demonstrated that a mutant of CpDmt1, an ortholog of RID of Neurospora crassa, resulted in the sectored phenotype but the CpDmt2 mutant did not, suggesting that the genetic basis of fungal sectorization is more complex. The present study revealed that a mutation in a signaling pathway component resulted in sectorization accompanied with changes in genome-wide DNA methylation, which suggests that this signal transduction pathway is important for epigenetic control of sectorization via regulation of genes involved in DNA methylation.

  2. Using Dendritic Heat Maps to Simultaneously Display Genotype Divergence with Phenotype Divergence.

    Science.gov (United States)

    Kellom, Matthew; Raymond, Jason

    2016-01-01

    The advancement of techniques to visualize and analyze large-scale sequencing datasets is an area of active research and is rooted in traditional techniques such as heat maps and dendrograms. We introduce dendritic heat maps that display heat map results over aligned DNA sequence clusters for a range of clustering cutoffs. Dendritic heat maps aid in visualizing the effects of group differences on clustering hierarchy and relative abundance of sampled sequences. Here, we artificially generate two separate datasets with simplified mutation and population growth procedures with GC content group separation to use as example phenotypes. In this work, we use the term phenotype to represent any feature by which groups can be separated. These sequences were clustered in a fractional identity range of 0.75 to 1.0 using agglomerative minimum-, maximum-, and average-linkage algorithms, as well as a divisive centroid-based algorithm. We demonstrate that dendritic heat maps give freedom to scrutinize specific clustering levels across a range of cutoffs, track changes in phenotype inequity across multiple levels of sequence clustering specificity, and easily visualize how deeply rooted changes in phenotype inequity are in a dataset. As genotypes diverge in sample populations, clusters are shown to break apart into smaller clusters at higher identity cutoff levels, similar to a dendrogram. Phenotype divergence, which is shown as a heat map of relative abundance bin response, may or may not follow genotype divergences. This joined view highlights the relationship between genotype and phenotype divergence for treatment groups. We discuss the minimum-, maximum-, average-, and centroid-linkage algorithm approaches to building dendritic heat maps and make a case for the divisive "top-down" centroid-based clustering methodology as being the best option visualize the effects of changing factors on clustering hierarchy and relative abundance.

  3. Using Dendritic Heat Maps to Simultaneously Display Genotype Divergence with Phenotype Divergence.

    Directory of Open Access Journals (Sweden)

    Matthew Kellom

    Full Text Available The advancement of techniques to visualize and analyze large-scale sequencing datasets is an area of active research and is rooted in traditional techniques such as heat maps and dendrograms. We introduce dendritic heat maps that display heat map results over aligned DNA sequence clusters for a range of clustering cutoffs. Dendritic heat maps aid in visualizing the effects of group differences on clustering hierarchy and relative abundance of sampled sequences. Here, we artificially generate two separate datasets with simplified mutation and population growth procedures with GC content group separation to use as example phenotypes. In this work, we use the term phenotype to represent any feature by which groups can be separated. These sequences were clustered in a fractional identity range of 0.75 to 1.0 using agglomerative minimum-, maximum-, and average-linkage algorithms, as well as a divisive centroid-based algorithm. We demonstrate that dendritic heat maps give freedom to scrutinize specific clustering levels across a range of cutoffs, track changes in phenotype inequity across multiple levels of sequence clustering specificity, and easily visualize how deeply rooted changes in phenotype inequity are in a dataset. As genotypes diverge in sample populations, clusters are shown to break apart into smaller clusters at higher identity cutoff levels, similar to a dendrogram. Phenotype divergence, which is shown as a heat map of relative abundance bin response, may or may not follow genotype divergences. This joined view highlights the relationship between genotype and phenotype divergence for treatment groups. We discuss the minimum-, maximum-, average-, and centroid-linkage algorithm approaches to building dendritic heat maps and make a case for the divisive "top-down" centroid-based clustering methodology as being the best option visualize the effects of changing factors on clustering hierarchy and relative abundance.

  4. DNA topology and transcription

    Science.gov (United States)

    Kouzine, Fedor; Levens, David; Baranello, Laura

    2014-01-01

    Chromatin is a complex assembly that compacts DNA inside the nucleus while providing the necessary level of accessibility to regulatory factors conscripted by cellular signaling systems. In this superstructure, DNA is the subject of mechanical forces applied by variety of molecular motors. Rather than being a rigid stick, DNA possesses dynamic structural variability that could be harnessed during critical steps of genome functioning. The strong relationship between DNA structure and key genomic processes necessitates the study of physical constrains acting on the double helix. Here we provide insight into the source, dynamics, and biology of DNA topological domains in the eukaryotic cells and summarize their possible involvement in gene transcription. We emphasize recent studies that might inspire and impact future experiments on the involvement of DNA topology in cellular functions. PMID:24755522

  5. Eukaryotic DNA Replication Fork.

    Science.gov (United States)

    Burgers, Peter M J; Kunkel, Thomas A

    2017-06-20

    This review focuses on the biogenesis and composition of the eukaryotic DNA replication fork, with an emphasis on the enzymes that synthesize DNA and repair discontinuities on the lagging strand of the replication fork. Physical and genetic methodologies aimed at understanding these processes are discussed. The preponderance of evidence supports a model in which DNA polymerase ε (Pol ε) carries out the bulk of leading strand DNA synthesis at an undisturbed replication fork. DNA polymerases α and δ carry out the initiation of Okazaki fragment synthesis and its elongation and maturation, respectively. This review also discusses alternative proposals, including cellular processes during which alternative forks may be utilized, and new biochemical studies with purified proteins that are aimed at reconstituting leading and lagging strand DNA synthesis separately and as an integrated replication fork.

  6. Biological significance of facilitated diffusion in protein-DNA interactions. Applications to T4 endonuclease V-initiated DNA repair

    International Nuclear Information System (INIS)

    Dowd, D.R.; Lloyd, R.S.

    1990-01-01

    Facilitated diffusion along nontarget DNA is employed by numerous DNA-interactive proteins to locate specific targets. Until now, the biological significance of DNA scanning has remained elusive. T4 endonuclease V is a DNA repair enzyme which scans nontarget DNA and processively incises DNA at the site of pyrimidine dimers which are produced by exposure to ultraviolet (UV) light. In this study we tested the hypothesis that there exists a direct correlation between the degree of processivity of wild type and mutant endonuclease V molecules and the degree of enhanced UV resistance which is conferred to repair-deficient Eshcerichia coli. This was accomplished by first creating a series of endonuclease V mutants whose in vitro catalytic activities were shown to be very similar to that of the wild type enzyme. However, when the mechanisms by which these enzymes search nontarget DNA for its substrate were analyzed in vitro and in vivo, the mutants displayed varying degrees of nontarget DNA scanning ranging from being nearly as processive as wild type to randomly incising dimers within the DNA population. The ability of these altered endonuclease V molecules to enhance UV survival in DNA repair-deficient E. coli then was assessed. The degree of enhanced UV survival was directly correlated with the level of facilitated diffusion. This is the first conclusive evidence directly relating a reduction of in vivo facilitated diffusion with a change in an observed phenotype. These results support the assertion that the mechanisms which DNA-interactive proteins employ in locating their target sites are of biological significance

  7. Mgm101p is a novel component of the mitochondrial nucleoid that binds DNA and is required for the repair of oxidatively damaged mitochondrial DNA

    International Nuclear Information System (INIS)

    Meeusen, S.; Tieu, Q.; Wong, E.; Weiss, E.; Schieltz, D.; Yates, J.R.; Nunnari, J.

    1999-01-01

    Maintenance of mitochondrial DNA (mtDNA) during cell division is required for progeny to be respiratory competent. Maintenance involves the replication, repair, assembly, segregation, and partitioning of the mitochondrial nucleoid. MGM101 has been identified as a gene essential for mtDNA maintenance in S. cerevisiae, but its role is unknown. Using liquid chromatography coupled with tandem mass spectrometry, we identified Mgm101p as a component of highly enriched nucleoids, suggesting that it plays a nucleoid-specific role in maintenance. Subcellular fractionation, indirect immunofluorescence and GFP tagging show that Mgm101p is exclusively associated with the mitochondrial nucleoid structure in cells. Furthermore, DNA affinity chromatography of nucleoid extracts indicates that Mgm101p binds to DNA, suggesting that its nucleoid localization is in part due to this activity. Phenotypic analysis of cells containing a temperature sensitive mgm101 allele suggests that Mgm101p is not involved in mtDNA packaging, segregation, partitioning or required for ongoing mtDNA replication. We examined Mgm101p's role in mtDNA repair. As compared with wild-type cells, mgm101 cells were more sensitive to mtDNA damage induced by UV irradiation and were hypersensitive to mtDNA damage induced by gamma rays and H2O2 treatment. Thus, we propose that Mgm101p performs an essential function in the repair of oxidatively damaged mtDNA that is required for the maintenance of the mitochondrial genome. (author)

  8. DNA-based machines.

    Science.gov (United States)

    Wang, Fuan; Willner, Bilha; Willner, Itamar

    2014-01-01

    The base sequence in nucleic acids encodes substantial structural and functional information into the biopolymer. This encoded information provides the basis for the tailoring and assembly of DNA machines. A DNA machine is defined as a molecular device that exhibits the following fundamental features. (1) It performs a fuel-driven mechanical process that mimics macroscopic machines. (2) The mechanical process requires an energy input, "fuel." (3) The mechanical operation is accompanied by an energy consumption process that leads to "waste products." (4) The cyclic operation of the DNA devices, involves the use of "fuel" and "anti-fuel" ingredients. A variety of DNA-based machines are described, including the construction of "tweezers," "walkers," "robots," "cranes," "transporters," "springs," "gears," and interlocked cyclic DNA structures acting as reconfigurable catenanes, rotaxanes, and rotors. Different "fuels", such as nucleic acid strands, pH (H⁺/OH⁻), metal ions, and light, are used to trigger the mechanical functions of the DNA devices. The operation of the devices in solution and on surfaces is described, and a variety of optical, electrical, and photoelectrochemical methods to follow the operations of the DNA machines are presented. We further address the possible applications of DNA machines and the future perspectives of molecular DNA devices. These include the application of DNA machines as functional structures for the construction of logic gates and computing, for the programmed organization of metallic nanoparticle structures and the control of plasmonic properties, and for controlling chemical transformations by DNA machines. We further discuss the future applications of DNA machines for intracellular sensing, controlling intracellular metabolic pathways, and the use of the functional nanostructures for drug delivery and medical applications.

  9. DNA repair and cancer

    International Nuclear Information System (INIS)

    Rathore, Shakuntla; Joshi, Pankaj Kumar; Gaur, Sudha

    2012-01-01

    DNA repair refers to a collection of processes by which a cell identifies and corrects damage to the DNA molecule that encode it's genome. In human cells, both normal metabolic activities and environmental factors such as UV light and radiation can cause DNA damage, resulting in as many one million individual molecular lesions per day. Many of these lesions cause structural damage to the DNA molecule and can alter or eliminate the cell's ability to transcribe the gene that the affected DNA encodes. Other lesions include potentially harmful mutation in cell's genome which affect the survival of it's daughter cells after it undergoes mitosis. As a consequence, the DNA repair process is constantly active as it responds to damage in the DNA structure. Inherited mutation that affect DNA repair genes are strongly associated with high cancer risks in humans. Hereditary non polyposis colorectal cancer (HNPCC) is strongly associated with specific mutation in the DNA mismatch repair pathway. BRCA1, BRCA2 two famous mutation conferring a hugely increased risk of breast cancer on carrier, are both associated with a large number of DNA repair pathway, especially NHEJ and homologous recombination. Cancer therapy procedures such as chemotherapy and radiotherapy work by overwhelming the capacity of the cell to repair DNA damage, resulting in cell death. Cells that are most rapidly dividing most typically cancer cells are preferentially affected. The side effect is that other non-cancerous but rapidly dividing cells such as stem cells in the bone marrow are also affected. Modern cancer treatment attempt to localize the DNA damage to cells and tissue only associated with cancer, either by physical means (concentrating the therapeutic agent in the region of the tumor) or by biochemical means (exploiting a feature unique to cancer cells in the body). (author)

  10. DNA hypomethylation at ALOX12 is associated with persistent wheezing in childhood.

    Science.gov (United States)

    Morales, Eva; Bustamante, Mariona; Vilahur, Nadia; Escaramis, Georgia; Montfort, Magda; de Cid, Rafael; Garcia-Esteban, Raquel; Torrent, Maties; Estivill, Xavier; Grimalt, Joan O; Sunyer, Jordi

    2012-05-01

    Epigenetic changes may play a role in the occurrence of asthma-related phenotypes. To identify epigenetic marks in terms of DNA methylation of asthma-related phenotypes in childhood, and to assess the effect of prenatal exposures and genetic variation on these epigenetic marks. Data came from two cohorts embedded in the Infancia y Medio Ambiente (INMA) PROJECT: Menorca (n = 122) and Sabadell (n = 236). Wheezing phenotypes were defined at age 4-6 years. Cytosine-guanine (CpG) dinucleotide site DNA methylation differences associated with wheezing phenotypes were screened in children of the Menorca study using the Illumina GoldenGate Panel I. Findings were validated and replicated using pyrosequencing. Information on maternal smoking and folate supplement use was obtained through questionnaires. Dichlorodiphenyldichloroethylene was measured in cord blood or maternal serum. Genotypes were extracted from genome-wide data. Screening identified lower DNA methylation at a CpG site in the arachidonate 12-lipoxygenase (ALOX12) gene in children having persistent wheezing compared with those never wheezed (P = 0.003). DNA hypomethylation at ALOX12 loci was associated with higher risk of persistent wheezing in the Menorca study (odds ratio per 1% methylation decrease, 1.13; 95% confidence interval, 0.99-1.29; P = 0.077) and in the Sabadell study (odds ratio, 1.16; 95% confidence interval, 1.03-1.37; P = 0.017). Higher levels of prenatal dichlorodiphenyldichloroethylene were associated with DNA hypomethylation of ALOX12 in the Menorca study (P = 0.033), but not in the Sabadell study (P = 0.377). ALOX12 DNA methylation was strongly determined by underlying genetic polymorphisms. DNA methylation of ALOX12 may be an epigenetic biomarker for the risk of asthma-related phenotypes.

  11. Nonisotopic DNA probe techniques

    National Research Council Canada - National Science Library

    Kricka, Larry J

    1992-01-01

    The objective of this book is to bring together descriptions of the principal nonisotopic methods for DNA hybridization assays, together with experimental details of the methods, including labelling...

  12. DNA replication and cancer

    DEFF Research Database (Denmark)

    Boyer, Anne-Sophie; Walter, David; Sørensen, Claus Storgaard

    2016-01-01

    A dividing cell has to duplicate its DNA precisely once during the cell cycle to preserve genome integrity avoiding the accumulation of genetic aberrations that promote diseases such as cancer. A large number of endogenous impacts can challenge DNA replication and cells harbor a battery of pathways...... causing DNA replication stress and genome instability. Further, we describe cellular and systemic responses to these insults with a focus on DNA replication restart pathways. Finally, we discuss the therapeutic potential of exploiting intrinsic replicative stress in cancer cells for targeted therapy....

  13. Forensic DNA testing.

    Science.gov (United States)

    Butler, John M

    2011-12-01

    Forensic DNA testing has a number of applications, including parentage testing, identifying human remains from natural or man-made disasters or terrorist attacks, and solving crimes. This article provides background information followed by an overview of the process of forensic DNA testing, including sample collection, DNA extraction, PCR amplification, short tandem repeat (STR) allele separation and sizing, typing and profile interpretation, statistical analysis, and quality assurance. The article concludes with discussions of possible problems with the data and other forensic DNA testing techniques.

  14. Structure of DNA toroids and electrostatic attraction of DNA duplexes

    International Nuclear Information System (INIS)

    Cherstvy, A G

    2005-01-01

    DNA-DNA electrostatic attraction is considered as the driving force for the formation of DNA toroids in the presence of DNA condensing cations. This attraction comes from the DNA helical charge distribution and favours hexagonal toroidal cross-sections. The latter is in agreement with recent cryo-electron microscopy studies on DNA condensed with cobalt hexammine. We treat the DNA-DNA interactions within the modern theory of electrostatic interaction between helical macromolecules. The size and thickness of the toroids is calculated within a simple model; other models of stability of DNA toroids are discussed and compared

  15. A mild mutator phenotype arises in a mouse model for malignancies associated with neurofibromatosis type 1

    International Nuclear Information System (INIS)

    Garza, Rene; Hudson, Robert A.; McMahan, C. Alex; Walter, Christi A.; Vogel, Kristine S.

    2007-01-01

    Defects in genes that control DNA repair, proliferation, and apoptosis can increase genomic instability, and thus promote malignant progression. Although most tumors that arise in humans with neurofibromatosis type 1 (NF1) are benign, these individuals are at increased risk for malignant peripheral nerve sheath tumors (MPNST). To characterize additional mutations required for the development of MPNST from benign plexiform neurofibromas, we generated a mouse model for these tumors by combining targeted null mutations in Nf1 and p53, in cis. CisNf1+/-; p53+/- mice spontaneously develop PNST, and these tumors exhibit loss-of-heterozygosity at both the Nf1 and p53 loci. Because p53 has well-characterized roles in the DNA damage response, DNA repair, and apoptosis, and because DNA repair genes have been proposed to act as modifiers in NF1, we used the cisNf1+/-; p53+/- mice to determine whether a mutator phenotype arises in NF1-associated malignancies. To quantitate spontaneous mutant frequencies (MF), we crossed the Big Blue mouse, which harbors a lacI transgene, to the cisNf1+/-; p53+/- mice, and isolated genomic DNA from both tumor and normal tissues in compound heterozygotes and wild-type siblings. Many of the PNST exhibited increased mutant frequencies (MF = 4.70) when compared to normal peripheral nerve and brain (MF = 2.09); mutations occurred throughout the entire lacI gene, and included base substitutions, insertions, and deletions. Moreover, the brains, spleens, and livers of these cisNf1+/-; p53+/- animals exhibited increased mutant frequencies when compared to tissues from wild-type littermates. We conclude that a mild mutator phenotype arises in the tumors and tissues of cisNf1+/-; p53+/- mice, and propose that genomic instability influences NF1 tumor progression and disease severity

  16. Onco-proteogenomics: Multi-omics level data integration for accurate phenotype prediction.

    Science.gov (United States)

    Dimitrakopoulos, Lampros; Prassas, Ioannis; Diamandis, Eleftherios P; Charames, George S

    2017-09-01

    The overall goal of translational oncology is to identify molecular alterations indicative of cancer or of responsiveness to specific therapeutic regimens. While next-generation sequencing has played a pioneering role in this quest, the latest advances in proteomic technologies promise to provide a holistic approach to the further elucidation of tumor biology. Genetic information may be written in DNA and flow from DNA to RNA to protein, according to the central dogma of molecular biology, but the observed phenotype is dictated predominantly by the DNA protein coding region-derived proteotype. Proteomics holds the potential to bridge the gap between genotype and phenotype, because the powerful analytical tool of mass spectrometry has reached a point of maturity to serve this purpose effectively. This integration of "omics" data has given birth to the novel field of onco-proteogenomics, which has much to offer to precision medicine and personalized patient management. Here, we review briefly how each "omics" technology has individually contributed to cancer research, discuss technological and computational advances that have contributed to the realization of onco-proteogenomics, and summarize current and future translational applications.

  17. Absence of Wip1 partially rescues Atm deficiency phenotypes in mice

    Science.gov (United States)

    Darlington, Yolanda; Nguyen, Thuy-Ai; Moon, Sung-Hwan; Herron, Alan; Rao, Pulivarthi; Zhu, Chengming; Lu, Xiongbin; Donehower, Lawrence A.

    2011-01-01

    Wildtype p53-Induced Phosphatase 1 (WIP1) is a serine/threonine phosphatase that dephosphorylates proteins in the ataxia telangiectasia mutated (ATM)-initiated DNA damage response pathway. WIP1 may play a homeostatic role in ATM signaling by returning the cell to a normal pre-stress state following completion of DNA repair. To better understand the effects of WIP1 on ATM signaling, we crossed Atm-deficient mice to Wip1-deficient mice and characterized phenotypes of the double knockout progeny. We hypothesized that the absence of Wip1 might rescue Atm deficiency phenotypes. Atm null mice, like ATM-deficient humans with the inherited syndrome ataxia telangiectasia, exhibit radiation sensitivity, fertility defects, and are T-cell lymphoma prone. Most double knockout mice were largely protected from lymphoma development and had a greatly extended lifespan compared to Atm null mice. Double knockout mice had increased p53 and H2AX phosphorylation and p21 expression compared to their Atm null counterparts, indicating enhanced p53 and DNA damage responses. Additionally, double knockout splenocytes displayed reduced chromosomal instability compared to Atm null mice. Finally, doubly null mice were partially rescued from infertility defects observed in Atm null mice. These results indicate that inhibition of WIP1 may represent a useful strategy for cancer treatment in general and A-T patients in particular. PMID:21765465

  18. Characterization of the Pseudomonas aeruginosa recA gene: the Les- phenotype

    International Nuclear Information System (INIS)

    Kokjohn, T.A.; Miller, R.V.

    1988-01-01

    The Les- phenotype (lysogeny establishment deficient) is a pleiotropic effect of the lesB908 mutation of Pseudomonas aeruginosa PAO. lesB908-containing strains are also (i) deficient in general recombination, (ii) sensitive to UV irradiation, and (iii) deficient in UV-stimulated induction of prophages. The P. aeruginosa recA-containing plasmid pKML3001 complemented each of these pleiotropic characteristics of the lesB908 mutation, supporting the hypothesis that lesB908 is an allele of the P. aeruginosa recA gene. The phenotypic effects of the lesB908 mutation may be best explained by the hypothesis that the lesB908 gene product is altered in such a way that it has lost synaptase activity but possesses intrinsic protease activity in the absence of DNA damage. The Les- phenotype is a result of the rapid destruction of newly synthesized phage repressor, resulting in lytic growth of the infecting virus. This hypothesis is consistent with the observations that increasing the number of copies of the phage repressor gene by increasing the multiplicity of infection (i.e., average number of phage genomes per cell) or by introducing the cloned phage repressor gene into a lesB908 mutant will also suppress the Les- phenotype in a phage-specific fashion

  19. Distribution of genotype network sizes in sequence-to-structure genotype-phenotype maps.

    Science.gov (United States)

    Manrubia, Susanna; Cuesta, José A

    2017-04-01

    An essential quantity to ensure evolvability of populations is the navigability of the genotype space. Navigability, understood as the ease with which alternative phenotypes are reached, relies on the existence of sufficiently large and mutually attainable genotype networks. The size of genotype networks (e.g. the number of RNA sequences folding into a particular secondary structure or the number of DNA sequences coding for the same protein structure) is astronomically large in all functional molecules investigated: an exhaustive experimental or computational study of all RNA folds or all protein structures becomes impossible even for moderately long sequences. Here, we analytically derive the distribution of genotype network sizes for a hierarchy of models which successively incorporate features of increasingly realistic sequence-to-structure genotype-phenotype maps. The main feature of these models relies on the characterization of each phenotype through a prototypical sequence whose sites admit a variable fraction of letters of the alphabet. Our models interpolate between two limit distributions: a power-law distribution, when the ordering of sites in the prototypical sequence is strongly constrained, and a lognormal distribution, as suggested for RNA, when different orderings of the same set of sites yield different phenotypes. Our main result is the qualitative and quantitative identification of those features of sequence-to-structure maps that lead to different distributions of genotype network sizes. © 2017 The Author(s).

  20. Genotype-phenotype variations in five Spanish families with Norrie disease or X-linked FEVR.

    Science.gov (United States)

    Riveiro-Alvarez, Rosa; Trujillo-Tiebas, Maria José; Gimenez-Pardo, Ascension; Garcia-Hoyos, Maria; Cantalapiedra, Diego; Lorda-Sanchez, Isabel; Rodriguez de Alba, Marta; Ramos, Carmen; Ayuso, Carmen

    2005-09-02

    Norrie disease (OMIM 310600) is a rare X-linked disorder characterized by congenital blindness in males. Approximately 40 to 50% of the cases develop deafness and mental retardation. X-linked familial exudative vitreoretinopathy (XL-FEVR) is a hereditary ocular disorder characterized by a failure of peripheral retinal vascularization. Both X-linked disorders are due to mutations in the NDP gene, which encodes a 133 amino acid protein called Norrin, but autosomal recessive (AR) and autosomal dominant (AD) forms of FEVR have also been described. In this study, we report the molecular findings and the related phenotype in five Spanish families affected with Norrie disease or XL-FEVR due to mutations of the NDP gene. The study was conducted in 45 subjects from five Spanish families. These families were clinically diagnosed with Norrie disease or similar conditions. The three exons of the NDP gene were analyzed by automatic DNA sequencing. Haplotype analyses were also performed. Two new nonsense mutations, apart from other mutations previously described in the NDP gene, were found in those patients affected with ND or X-linked FEVR. An important genotype-phenotype variation was found in relation to the different mutations of the NDP gene. In fact, the same mutation may be responsible for different phenotypes. We speculate that there might be other molecular factors that interact in the retina with Norrin, which contribute to the resultant phenotypes.

  1. Phenotypic heterogeneity associated with a novel mutation (Gly112Glu) in the Norrie disease protein.

    Science.gov (United States)

    Allen, R C; Russell, S R; Streb, L M; Alsheikheh, A; Stone, E M

    2006-02-01

    To determine the molecular pathology and clinical severity of two pedigrees with a history of early retinal detachment and peripheral retinal vascular abnormalities. Longitudinal cohort study. A longitudinal clinical study and DNA analysis was performed on 49 family members of two pedigrees. Nine individuals were found to be hemizygous for a mutation at codon 112 (Gly112Glu) of the Norrie disease protein (NDP) in one pedigree. Significant phenotypic heterogeneity was found. The proband presented with a unilateral subtotal retinal detachment at the age of 3 years, and subsequently developed a slowly progressive tractional retinal detachment involving the macula in the contralateral eye at the age of 4 years. One individual had only mild peripheral retinal pigmentary changes with normal vision at the age of 79 years. The remaining seven individuals had varying degrees of peripheral retinal vascular abnormalities and anterior segment findings. Seven affected members of a second pedigree affected by a previously reported mutation, Arg74Cys, also demonstrated wide ocular phenotypic variation. A novel mutation (Gly112Glu), which represents the most carboxy located, NDP mutation reported, results in significant phenotypic heterogeneity. These data support the contention that the spectrum of ocular disease severity associated with these NDP mutations is broad. Use of terms that characterize this entity by phenotypic appearance, such as familial exudative vitreoretinopathy, do not adequately communicate the potential spectrum of severity of this disorder to affected or carrier family members.

  2. Phenotypic characterisation and molecular polymorphism of ...

    African Journals Online (AJOL)

    The study of the phenotypic characterisation and molecular polymorphism of local chicken populations was carried out in Benin on 326 chickens of the Forest ecological area and 316 of the Savannah ecological area, all were 7 months old at least. The collection of blood for the molecular typing was achieved on 121 ...

  3. Evidence for a Broad Autism Phenotype

    NARCIS (Netherlands)

    K. de Groot (Kristel); J.W. van Strien (Jan)

    2017-01-01

    textabstractThe broad autism phenotype implies the existence of a continuum ranging from individuals displaying almost no autistic traits to severely impaired diagnosed individuals. Recent studies have linked this variation in autistic traits to several domains of functioning. However, studies

  4. phenotype correlation of methylene tetrahydrofolate reductase ...

    African Journals Online (AJOL)

    Rabah M. Shawky

    2014-06-21

    Jun 21, 2014 ... children with autism and to correlate them with different phenotypes. Subjects and ... of impairments in communication, reciprocal social interac- tions, and ... isolation was obtained from peripheral blood samples using the spin ... IQ, while ten of them (50%) had mild mental retardation and six patients (30%) ...

  5. Phenotype Presentation of Hypophosphatemic Rickets in Adults

    DEFF Research Database (Denmark)

    Beck-Nielsen, Signe S; Brusgaard, Klaus; Rasmussen, Lars M

    2010-01-01

    Hypophosphatemic rickets (HR) is a group of rare disorders caused by excessive renal phosphate wasting. The purpose of this cross-sectional study of 38 HR patients was to characterize the phenotype of adult HR patients. Moreover, skeletal and endodontic severity scores were defined to assess poss...

  6. Phenotypic and molecular characterization of Salmonella serotypes ...

    African Journals Online (AJOL)

    The presence of Salmonella and human pathogens in unpasteurized milk remains a public health hazard. The study reported the phenotypic and molecular characterization of Salmonella serotypes in cow raw milk, cheese and traditional yoghurt marketed for man's consumption in Nigeria. Isolation of Salmonella was done ...

  7. (RR) soybean cultivars estimated by phenotypic characteristics

    African Journals Online (AJOL)

    SAM

    2014-06-25

    Jun 25, 2014 ... phenotypic characteristics and microsatellite molecular markers (SSR). ... discriminatory analysis, principal components, coordinate and cluster analysis .... were employed with 10.000 simulations to attribute significance values to ...... association analysis of protein and oil content in food-grade soybeans ...

  8. Colorectal Cancer "Methylator Phenotype": Fact or Artifact?

    Directory of Open Access Journals (Sweden)

    Charles Anacleto

    2005-04-01

    Full Text Available It has been proposed that human colorectal tumors can be classified into two groups: one in which methylation is rare, and another with methylation of several loci associated with a "CpG island methylated phenotype (CIMP," characterized by preferential proximal location in the colon, but otherwise poorly defined. There is considerable overlap between this putative methylator phenotype and the well-known mutator phenotype associated with microsatellite instability (MSI. We have examined hypermethylation of the promoter region of five genes (DAPK, MGMT, hMLH1, p16INK4a, and p14ARF in 106 primary colorectal cancers. A graph depicting the frequency of methylated loci in the series of tumors showed a continuous, monotonically decreasing distribution quite different from the previously claimed discontinuity. We observed a significant association between the presence of three or more methylated loci and the proximal location of the tumors. However, if we remove from analysis the tumors with hMLH1 methylation or those with MSI, the significance vanishes, suggesting that the association between multiple methylations and proximal location was indirect due to the correlation with MSI. Thus, our data do not support the independent existence of the so-called methylator phenotype and suggest that it rather may represent a statistical artifact caused by confounding of associations.

  9. DNA methylation mediates genetic variation for adaptive transgenerational plasticity.

    Science.gov (United States)

    Herman, Jacob J; Sultan, Sonia E

    2016-09-14

    Environmental stresses experienced by individual parents can influence offspring phenotypes in ways that enhance survival under similar conditions. Although such adaptive transgenerational plasticity is well documented, its transmission mechanisms are generally unknown. One possible mechanism is environmentally induced DNA methylation changes. We tested this hypothesis in the annual plant Polygonum persicaria, a species known to express adaptive transgenerational plasticity in response to parental drought stress. Replicate plants of 12 genetic lines (sampled from natural populations) were grown in dry versus moist soil. Their offspring were exposed to the demethylating agent zebularine or to control conditions during germination and then grown in dry soil. Under control germination conditions, the offspring of drought-stressed parents grew longer root systems and attained greater biomass compared with offspring of well-watered parents of the same genetic lines. Demethylation removed these adaptive developmental effects of parental drought, but did not significantly alter phenotypic expression in offspring of well-watered parents. The effect of demethylation on the expression of the parental drought effect varied among genetic lines. Differential seed provisioning did not contribute to the effect of parental drought on offspring phenotypes. These results demonstrate that DNA methylation can mediate adaptive, genotype-specific effects of parental stress on offspring phenotypes. © 2016 The Author(s).

  10. Peruvian Maca (Lepidium peruvianum) - III: The Effects of Cultivation Altitude on Phytochemical and Genetic Differences in the Four Prime Maca Phenotypes.

    Science.gov (United States)

    Meissner, Henry O; Mscisz, Alina; Baraniak, Marek; Piatkowska, Ewa; Pisulewski, Pawel; Mrozikiewicz, Mieczyslaw; Bobkiewicz-Kozlowska, Teresa

    2017-06-01

    In two trials, dietary and Glucosinolates' characteristics in four Maca phenotypes have been examined with an extension into the determination of DNA sequences. Hypocotyls of the four prime phenotypes of Peruvian Maca - Lepidium peruvianum Chacon, labelled as "Yellow", "Black", "Red" and "Purple" were separated from mixed Maca crops cultivated in four geographically-distant locations in the Peruvian Andes at altitudes between 2,800m and 4,300 m a.s.l. It was found that at higher altitudes where Red and Purple Maca phenotypes were grown, the significantly higher ( P Maca physiological activity, were observed with the Purple phenotype showing the highest Glucosinolates' content at 4,300m a.s.l., followed by the Red-coloured hypocotyls. Black Maca showed a reversal, but also a significant ( P 0.05) and has consistently the lowest Glucosinolates content. Thus, it is reasonable to assume that the altitude at which Red, Purple and Black phenotypes of L. peruvianum are grown, may be responsible for the variation in physiologic functionalities, leading to different than expected specific therapeutic and health benefits induced by Maca phenotypes grown at diverse altitudes. Although promising, insufficiently precise differences in DNA sequences failed to distinguish, without any reasonable doubt, four Maca phenotypes cultivated either in the same or geographically-distant locations, and harvested at different altitudes a.s.l. Further research on DNA sequences is needed, with more primers and larger number of Maca phenotypes, considering biosynthesis of secondary metabolites and adaptation pathways induced by harsh environment at altitudes where Maca is cultivated.

  11. Peruvian Maca (Lepidium peruvianum) – III: The Effects of Cultivation Altitude on Phytochemical and Genetic Differences in the Four Prime Maca Phenotypes

    Science.gov (United States)

    Meissner, Henry O.; Mscisz, Alina; Baraniak, Marek; Piatkowska, Ewa; Pisulewski, Pawel; Mrozikiewicz, Mieczyslaw; Bobkiewicz-Kozlowska, Teresa

    2017-01-01

    In two trials, dietary and Glucosinolates’ characteristics in four Maca phenotypes have been examined with an extension into the determination of DNA sequences. Hypocotyls of the four prime phenotypes of Peruvian Maca - Lepidium peruvianum Chacon, labelled as “Yellow”, “Black”, “Red” and “Purple” were separated from mixed Maca crops cultivated in four geographically-distant locations in the Peruvian Andes at altitudes between 2,800m and 4,300 m a.s.l. It was found that at higher altitudes where Red and Purple Maca phenotypes were grown, the significantly higher (PMaca physiological activity, were observed with the Purple phenotype showing the highest Glucosinolates’ content at 4,300m a.s.l., followed by the Red-coloured hypocotyls. Black Maca showed a reversal, but also a significant (P0.05) and has consistently the lowest Glucosinolates content. Thus, it is reasonable to assume that the altitude at which Red, Purple and Black phenotypes of L. peruvianum are grown, may be responsible for the variation in physiologic functionalities, leading to different than expected specific therapeutic and health benefits induced by Maca phenotypes grown at diverse altitudes. Although promising, insufficiently precise differences in DNA sequences failed to distinguish, without any reasonable doubt, four Maca phenotypes cultivated either in the same or geographically-distant locations, and harvested at different altitudes a.s.l. Further research on DNA sequences is needed, with more primers and larger number of Maca phenotypes, considering biosynthesis of secondary metabolites and adaptation pathways induced by harsh environment at altitudes where Maca is cultivated. PMID:28824342

  12. Worm Phenotype Ontology: Integrating phenotype data within and beyond the C. elegans community

    Directory of Open Access Journals (Sweden)

    Yook Karen

    2011-01-01

    Full Text Available Abstract Background Caenorhabditis elegans gene-based phenotype information dates back to the 1970's, beginning with Sydney Brenner and the characterization of behavioral and morphological mutant alleles via classical genetics in order to understand nervous system function. Since then C. elegans has become an important genetic model system for the study of basic biological and biomedical principles, largely through the use of phenotype analysis. Because of the growth of C. elegans as a genetically tractable model organism and the development of large-scale analyses, there has been a significant increase of phenotype data that needs to be managed and made accessible to the research community. To do so, a standardized vocabulary is necessary to integrate phenotype data from diverse sources, permit integration with other data types and render the data in a computable form. Results We describe a hierarchically structured, controlled vocabulary of terms that can be used to standardize phenotype descriptions in C. elegans, namely the Worm Phenotype Ontology (WPO. The WPO is currently comprised of 1,880 phenotype terms, 74% of which have been used in the annotation of phenotypes associated with greater than 18,000 C. elegans genes. The scope of the WPO is not exclusively limited to C. elegans biology, rather it is devised to also incorporate phenotypes observed in related nematode species. We have enriched the value of the WPO by integrating it with other ontologies, thereby increasing the accessibility of worm phenotypes to non-nematode biologists. We are actively developing the WPO to continue to fulfill the evolving needs of the scientific community and hope to engage researchers in this crucial endeavor. Conclusions We provide a phenotype ontology (WPO that will help to facilitate data retrieval, and cross-species comparisons within the nematode community. In the larger scientific community, the WPO will permit data integration, and

  13. Diagnosis of becker muscular dystrophy: Results of Re-analysis of DNA samples.

    Science.gov (United States)

    Straathof, Chiara S M; Van Heusden, Dave; Ippel, Pieternella F; Post, Jan G; Voermans, Nicol C; De Visser, Marianne; Brusse, Esther; Van Den Bergen, Janneke C; Van Der Kooi, Anneke J; Verschuuren, Jan J G M; Ginjaar, Hendrika B

    2016-01-01

    The phenotype of Becker muscular dystrophy (BMD) is highly variable, and the disease may be underdiagnosed. We searched for new mutations in the DMD gene in a cohort of previously undiagnosed patients who had been referred in the period 1985-1995. All requests for DNA analysis of the DMD gene in probands with suspected BMD were re-evaluated. If the phenotype was compatible with BMD, and no deletions or duplications were detected, DNA samples were screened for small mutations. In 79 of 185 referrals, no mutation was found. Analysis could be performed on 31 DNA samples. Seven different mutations, including 3 novel ones, were found. Long-term clinical follow-up is described. Refining DNA analysis in previously undiagnosed cases can identify mutations in the DMD gene and provide genetic diagnosis of BMD. A delayed diagnosis can still be valuable for the proband or the relatives of BMD patients. © 2015 Wiley Periodicals, Inc.

  14. DNA methylation-based variation between human populations.

    Science.gov (United States)

    Kader, Farzeen; Ghai, Meenu

    2017-02-01

    Several studies have proved that DNA methylation affects regulation of gene expression and development. Epigenome-wide studies have reported variation in methylation patterns between populations, including Caucasians, non-Caucasians (Blacks), Hispanics, Arabs, and numerous populations of the African continent. Not only has DNA methylation differences shown to impact externally visible characteristics, but is also a potential biomarker for underlying racial health disparities between human populations. Ethnicity-related methylation differences set their mark during early embryonic development. Genetic variations, such as single-nucleotide polymorphisms and environmental factors, such as age, dietary folate, socioeconomic status, and smoking, impacts DNA methylation levels, which reciprocally impacts expression of phenotypes. Studies show that it is necessary to address these external influences when attempting to differentiate between populations since the relative impacts of these factors on the human methylome remain uncertain. The present review summarises several reported attempts to establish the contribution of differential DNA methylation to natural human variation, and shows that DNA methylation could represent new opportunities for risk stratification and prevention of several diseases amongst populations world-wide. Variation of methylation patterns between human populations is an exciting prospect which inspires further valuable research to apply the concept in routine medical and forensic casework. However, trans-generational inheritance needs to be quantified to decipher the proportion of variation contributed by DNA methylation. The future holds thorough evaluation of the epigenome to understand quantification, heritability, and the effect of DNA methylation on phenotypes. In addition, methylation profiling of the same ethnic groups across geographical locations will shed light on conserved methylation differences in populations.

  15. The single-strand DNA binding activity of human PC4 preventsmutagenesis and killing by oxidative DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Jen-Yeu; Sarker, Altaf Hossain; Cooper, Priscilla K.; Volkert, Michael R.

    2004-02-01

    Human positive cofactor 4 (PC4) is a transcriptional coactivator with a highly conserved single-strand DNA (ssDNA) binding domain of unknown function. We identified PC4 as a suppressor of the oxidative mutator phenotype of the Escherichia coli fpg mutY mutant and demonstrate that this suppression requires its ssDNA binding activity. Yeast mutants lacking their PC4 ortholog Sub1 are sensitive to hydrogen peroxide and exhibit spontaneous and peroxide induced hypermutability. PC4 expression suppresses the peroxide sensitivity of the yeast sub l{Delta} mutant, suggesting that the human protein has a similar function. A role for yeast and human proteins in DNA repair is suggested by the demonstration that Sub1 acts in a peroxide-resistance pathway involving Rad2 and by the physical interaction of PC4 with the human Rad2 homolog XPG. We show XPG recruits PC4 to a bubble-containing DNA substrate with resulting displacement of XPG and formation of a PC4-DNA complex. We discuss the possible requirement for PC4 in either global or transcription-coupled repair of oxidative DNA damage to mediate the release of XPG bound to its substrate.

  16. Extended DNA Tile Actuators

    DEFF Research Database (Denmark)

    Kristiansen, Martin; Kryger, Mille; Zhang, Zhao

    2012-01-01

    A dynamic linear DNA tile actuator is expanded to three new structures of higher complexity. The original DNA actuator was constructed from a central roller strand which hybridizes with two piston strands by forming two half-crossover junctions. A linear expansion of the actuator is obtained...

  17. Dna fingerprinting - review paper

    OpenAIRE

    Blundell, Renald

    2006-01-01

    Before the Polymerase Chain Reaction (PCR) was established, DNA fingerprinting technology has relied for years on Restriction Fragment Length Polymorphism (RFLP) and Variable Number of Tandom Repeats (VNTR) analysis, a very efficient technique but quite laborious and not suitable for high throughput mapping. Since its, development, PCR has provided a new and powerful tool for DNA fingerprinting.

  18. DNA Repair Systems

    Indian Academy of Sciences (India)

    Thanks to the pioneering research work of Lindahl, Sancar, Modrich and their colleagues, we now have an holistic awareness of how DNA damage occurs and how the damage is rectified in bacteria as well as in higher organisms including human beings. A comprehensive understanding of DNA repair has proven crucial ...

  19. DNA repair genes

    International Nuclear Information System (INIS)

    Morimyo, Mitsuoki

    1995-01-01

    Fission yeast S. pombe is assumed to be a good model for cloning of human DNA repair genes, because human gene is normally expressed in S. pombe and has a very similar protein sequence to yeast protein. We have tried to elucidate the DNA repair mechanisms of S. pombe as a model system for those of mammals. (J.P.N.)

  20. DNA-cell conjugates

    Science.gov (United States)

    Hsiao, Shih-Chia; Francis, Matthew B.; Bertozzi, Carolyn; Mathies, Richard; Chandra, Ravi; Douglas, Erik; Twite, Amy; Toriello, Nicholas; Onoe, Hiroaki

    2018-05-15

    The present invention provides conjugates of DNA and cells by linking the DNA to a native functional group on the cell surface. The cells can be without cell walls or can have cell walls. The modified cells can be linked to a substrate surface and used in assay or bioreactors.

  1. Characterization of muntjac DNA

    International Nuclear Information System (INIS)

    Davis, R.C.

    1981-01-01

    Sister chromatid exchange (SCE) in muntjac chromosomes is generally proportional to the chromosomal DNA content, but the SCE frequency is reduced in the heterochromatic neck region of the X chromosome. The physical properties of muntjac DNA and the kinetics of repair of UV damage in muntjac heterochromatin and euchromatin were examined and compared with the distribution of sister chromatid exchange

  2. DNA-cell conjugates

    Science.gov (United States)

    Hsiao, Shih-Chia; Francis, Matthew B.; Bertozzi, Carolyn; Mathies, Richard; Chandra, Ravi; Douglas, Erik; Twite, Amy; Toriello, Nicholas; Onoe, Hiroaki

    2016-05-03

    The present invention provides conjugates of DNA and cells by linking the DNA to a native functional group on the cell surface. The cells can be without cell walls or can have cell walls. The modified cells can be linked to a substrate surface and used in assay or bioreactors.

  3. Characterization of muntjac DNA

    Energy Technology Data Exchange (ETDEWEB)

    Davis, R.C.

    1981-05-27

    Sister chromatid exchange (SCE) in muntjac chromosomes is generally proportional to the chromosomal DNA content, but the SCE frequency is reduced in the heterochromatic neck region of the X chromosome. The physical properties of muntjac DNA and the kinetics of repair of UV damage in muntjac heterochromatin and euchromatin were examined and compared with the distribution of sister chromatid exchange.

  4. The influence of DNA sequence on epigenome-induced pathologies

    Directory of Open Access Journals (Sweden)

    Meagher Richard B

    2012-07-01

    Full Text Available Abstract Clear cause-and-effect relationships are commonly established between genotype and the inherited risk of acquiring human and plant diseases and aberrant phenotypes. By contrast, few such cause-and-effect relationships are established linking a chromatin structure (that is, the epitype with the transgenerational risk of acquiring a disease or abnormal phenotype. It is not entirely clear how epitypes are inherited from parent to offspring as populations evolve, even though epigenetics is proposed to be fundamental to evolution and the likelihood of acquiring many diseases. This article explores the hypothesis that, for transgenerationally inherited chromatin structures, “genotype predisposes epitype”, and that epitype functions as a modifier of gene expression within the classical central dogma of molecular biology. Evidence for the causal contribution of genotype to inherited epitypes and epigenetic risk comes primarily from two different kinds of studies discussed herein. The first and direct method of research proceeds by the examination of the transgenerational inheritance of epitype and the penetrance of phenotype among genetically related individuals. The second approach identifies epitypes that are duplicated (as DNA sequences are duplicated and evolutionarily conserved among repeated patterns in the DNA sequence. The body of this article summarizes particularly robust examples of these studies from humans, mice, Arabidopsis, and other organisms. The bulk of the data from both areas of research support the hypothesis that genotypes predispose the likelihood of displaying various epitypes, but for only a few classes of epitype. This analysis suggests that renewed efforts are needed in identifying polymorphic DNA sequences that determine variable nucleosome positioning and DNA methylation as the primary cause of inherited epigenome-induced pathologies. By contrast, there is very little evidence that DNA sequence directly

  5. DNA alkylation damage as a sensor of nitrosative stress in Mycobacterium tuberculosis

    OpenAIRE

    Durbach, S I; Springer, B; Machowski, E E; North, R J; Papavinasasundaram, K G; Colston, M J; Böttger, E C; Mizrahi, V

    2003-01-01

    One of the cellular consequences of nitrosative stress is alkylation damage to DNA. To assess whether nitrosative stress is registered on the genome of Mycobacterium tuberculosis, mutants lacking an alkylation damage repair and reversal operon were constructed. Although hypersensitive to the genotoxic effects of N-methyl-N′-nitro-N-nitrosoguanidine in vitro, the mutants displayed no phenotype in vivo, suggesting that permeation of nitrosative stress to the level of cytotoxic DNA damage is res...

  6. Uncommon nucleotide excision repair phenotypes revealed by targeted high-throughput sequencing.

    Science.gov (United States)

    Calmels, Nadège; Greff, Géraldine; Obringer, Cathy; Kempf, Nadine; Gasnier, Claire; Tarabeux, Julien; Miguet, Marguerite; Baujat, Geneviève; Bessis, Didier; Bretones, Patricia; Cavau, Anne; Digeon, Béatrice; Doco-Fenzy, Martine; Doray, Bérénice; Feillet, François; Gardeazabal, Jesus; Gener, Blanca; Julia, Sophie; Llano-Rivas, Isabel; Mazur, Artur; Michot, Caroline; Renaldo-Robin, Florence; Rossi, Massimiliano; Sabouraud, Pascal; Keren, Boris; Depienne, Christel; Muller, Jean; Mandel, Jean-Louis; Laugel, Vincent

    2016-03-22

    Deficient nucleotide excision repair (NER) activity causes a variety of autosomal recessive diseases including xeroderma pigmentosum (XP) a disorder which pre-disposes to skin cancer, and the severe multisystem condition known as Cockayne syndrome (CS). In view of the clinical overlap between NER-related disorders, as well as the existence of multiple phenotypes and the numerous genes involved, we developed a new diagnostic approach based on the enrichment of 16 NER-related genes by multiplex amplification coupled with next-generation sequencing (NGS). Our test cohort consisted of 11 DNA samples, all with known mutations and/or non pathogenic SNPs in two of the tested genes. We then used the same technique to analyse samples from a prospective cohort of 40 patients. Multiplex amplification and sequencing were performed using AmpliSeq protocol on the Ion Torrent PGM (Life Technologies). We identified causative mutations in 17 out of the 40 patients (43%). Four patients showed biallelic mutations in the ERCC6(CSB) gene, five in the ERCC8(CSA) gene: most of them had classical CS features but some had very mild and incomplete phenotypes. A small cohort of 4 unrelated classic XP patients from the Basque country (Northern Spain) revealed a common splicing mutation in POLH (XP-variant), demonstrating a new founder effect in this population. Interestingly, our results also found ERCC2(XPD), ERCC3(XPB) or ERCC5(XPG) mutations in two cases of UV-sensitive syndrome and in two cases with mixed XP/CS phenotypes. Our study confirms that NGS is an efficient technique for the analysis of NER-related disorders on a molecular level. It is particularly useful for phenotypes with combined features or unusually mild symptoms. Targeted NGS used in conjunction with DNA repair functional tests and precise clinical evaluation permits rapid and cost-effective diagnosis in patients with NER-defects.

  7. U-251 revisited: genetic drift and phenotypic consequences of long-term cultures of glioblastoma cells

    International Nuclear Information System (INIS)

    Torsvik, Anja; Stieber, Daniel; Enger, Per Øyvind; Golebiewska, Anna; Molven, Anders; Svendsen, Agnete; Westermark, Bengt; Niclou, Simone P; Olsen, Thale Kristin; Chekenya Enger, Martha; Bjerkvig, Rolf

    2014-01-01

    It is well known that in vitro subculture represents a selection pressure on cell lines, and over time this may result in a genetic drift in the cancer cells. In addition, long-term cultures harbor the risk of cross-contamination with other cell lines. The consequences may have major impact on experimental results obtained in various laboratories, where the cell lines no longer reflect the original tumors that they are supposed to represent. Much neglected in the scientific community is a close monitoring of cell cultures by regular phenotypic and genetic characterization. In this report, we present a thorough characterization of the commonly used glioblastoma (GBM) model U-251, which in numerous publications has been wrongly identified as U-373, due to an earlier cross-contamination. In this work, the original U-251 and three subclones of U-251, commonly referred to as U-251 or U-373, were analyzed with regard to their DNA profile, morphology, phenotypic expression, and growth pattern. By array comparative genomic hybridization (aCGH), we show that only the original low-passaged U-251 cells, established in the 1960s, maintain a DNA copy number resembling a typical GBM profile, whereas all long-term subclones lost the typical GBM profile. Also the long-term passaged subclones displayed variations in phenotypic marker expression and showed an increased growth rate in vitro and a more aggressive growth in vivo. Taken together, the variations in genotype and phenotype as well as differences in growth characteristics may explain different results reported in various laboratories related to the U-251 cell line

  8. Comparison of DNA quantification methodology used in the DNA extraction protocol for the UK Biobank cohort.

    Science.gov (United States)

    Welsh, Samantha; Peakman, Tim; Sheard, Simon; Almond, Rachael

    2017-01-05

    UK Biobank is a large prospective cohort study in the UK established by the Medical Research Council (MRC) and the Wellcome Trust to enable approved researchers to investigate the role of genetic factors, environmental exposures and lifestyle in the causes of major diseases of late and middle age. A wide range of phenotypic data has been collected at recruitment and has recently been enhanced by the UK Biobank Genotyping Project. All UK Biobank participants (500,000) have been genotyped on either the UK Biobank Axiom® Array or the Affymetrix UK BiLEVE Axiom® Array and the workflow for preparing samples for genotyping is described. The genetic data is hoped to provide further insight into the genetics of disease. All data, including the genetic data, is available for access to approved researchers. Data for two methods of DNA quantification (ultraviolet-visible spectroscopy [UV/Vis]) measured on the Trinean DropSense™ 96 and PicoGreen®) were compared by two laboratories (UK Biobank and Affymetrix). The sample processing workflow established at UK Biobank, for genotyping on the custom Affymetrix Axiom® array, resulted in high quality DNA (average DNA concentration 38.13 ng/μL, average 260/280 absorbance 1.91). The DNA generated high quality genotype data (average call rate 99.48% and pass rate 99.45%). The DNA concentration measured on the Trinean DropSense™ 96 at UK Biobank correlated well with DNA concentration measured by PicoGreen® at Affymetrix (r = 0.85). The UK Biobank Genotyping Project demonstrated that the high throughput DNA extraction protocol described generates high quality DNA suitable for genotyping on the Affymetrix Axiom array. The correlation between DNA concentration derived from UV/Vis and PicoGreen® quantification methods suggests, in large-scale genetic studies involving two laboratories, it may be possible to remove the DNA quantification step in one laboratory without affecting downstream analyses. This would result in

  9. Phenotypic equilibrium as probabilistic convergence in multi-phenotype cell population dynamics.

    Directory of Open Access Journals (Sweden)

    Da-Quan Jiang

    Full Text Available We consider the cell population dynamics with n different phenotypes. Both the Markovian branching process model (stochastic model and the ordinary differential equation (ODE system model (deterministic model are presented, and exploited to investigate the dynamics of the phenotypic proportions. We will prove that in both models, these proportions will tend to constants regardless of initial population states ("phenotypic equilibrium" under weak conditions, which explains the experimental phenomenon in Gupta et al.'s paper. We also prove that Gupta et al.'s explanation is the ODE model under a special assumption. As an application, we will give sufficient and necessary conditions under which the proportion of one phenotype tends to 0 (die out or 1 (dominate. We also extend our results to non-Markovian cases.

  10. Invasion strategies in clonal aquatic plants: Are phenotypic differences caused by phenotypic plasticity or local adaptation?

    DEFF Research Database (Denmark)

    Riis, Tenna; Lambertini, Carla; Olesen, Birgit

    2010-01-01

    conditions and plant morphological characteristics. Conclusions: The results indicate that at the current stage of spread into New Zealand, the primary adaptive strategy of these three invasive macrophytes is phenotypic plasticity. However, while limited, the possibility that genetic diversity between......Background and Aims: The successful spread of invasive plants in new environments is often linked to multiple introductions and a diverse gene pool that facilitates local adaptation to variable environmental conditions. For clonal plants, however, phenotypic plasticity may be equally important....... Methods: Field populations with a large phenotypic variety were sampled in a range of lakes and streams with different chemical and physical properties. The phenotypic plasticity of the species before and after cultivation was studied in a common garden growth experiment, and the genetic diversity...

  11. A further case of a Prader-Willi syndrome phenotype in a patient with Angelman syndrome molecular defect

    Directory of Open Access Journals (Sweden)

    De Molfetta Greice Andreotti

    2002-01-01

    Full Text Available Angelman syndrome (AS and Prader-Willi syndrome (PWS are distinct human neurogenetic disorders; however, a clinical overlap between AS and PWS has been identified. We report on a further case of a patient showing the PWS phenotype with the AS molecular defect. Despite the PWS phenotype, the DNA methylation analysis of SNRPN revealed an AS pattern. Cytogenetic and FISH analysis showed normal chromosomes 15 and microsatellite analysis showed heterozygous loci inside and outside the 15q11-13 region. The presence of these atypical cases could be more frequent than previously expected and we reinforce that the DNA methylation analysis is important for the correct diagnosis of severe mental deficiency, congenital hypotonia and obesity.

  12. Whose DNA is this?

    DEFF Research Database (Denmark)

    Taroni, Franco; Biedermann, Alex; Vuille, Joëlle

    2013-01-01

    This communication seeks to draw the attention of researchers and practitioners dealing with forensic DNA profiling analyses to the following question: is a scientist's report, offering support to a hypothesis according to which a particular individual is the source of DNA detected during...... evoked during the international conference "The hidden side of DNA profiles. Artifacts, errors and uncertain evidence" held in Rome (April 27th to 28th, 2012). Indeed, despite the fact that this conference brought together some of the world's leading forensic DNA specialists, it appeared clearly...... talk considerably different languages. It thus is fundamental to address this issue of communication about results of forensic DNA analyses, and open a dialogue with practicing non-scientists at large who need to make meaningful use of scientific results to approach and help solve judicial cases...

  13. DNA repair protocols

    DEFF Research Database (Denmark)

    Bjergbæk, Lotte

    In its 3rd edition, this Methods in Molecular Biology(TM) book covers the eukaryotic response to genomic insult including advanced protocols and standard techniques in the field of DNA repair. Offers expert guidance for DNA repair, recombination, and replication. Current knowledge of the mechanisms...... that regulate DNA repair has grown significantly over the past years with technology advances such as RNA interference, advanced proteomics and microscopy as well as high throughput screens. The third edition of DNA Repair Protocols covers various aspects of the eukaryotic response to genomic insult including...... recent advanced protocols as well as standard techniques used in the field of DNA repair. Both mammalian and non-mammalian model organisms are covered in the book, and many of the techniques can be applied with only minor modifications to other systems than the one described. Written in the highly...

  14. Racemic DNA crystallography.

    Science.gov (United States)

    Mandal, Pradeep K; Collie, Gavin W; Kauffmann, Brice; Huc, Ivan

    2014-12-22

    Racemates increase the chances of crystallization by allowing molecular contacts to be formed in a greater number of ways. With the advent of protein synthesis, the production of protein racemates and racemic-protein crystallography are now possible. Curiously, racemic DNA crystallography had not been investigated despite the commercial availability of L- and D-deoxyribo-oligonucleotides. Here, we report a study into racemic DNA crystallography showing the strong propensity of racemic DNA mixtures to form racemic crystals. We describe racemic crystal structures of various DNA sequences and folded conformations, including duplexes, quadruplexes, and a four-way junction, showing that the advantages of racemic crystallography should extend to DNA. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Decomposing phenotype descriptions for the human skeletal phenome.

    Science.gov (United States)

    Groza, Tudor; Hunter, Jane; Zankl, Andreas

    2013-01-01

    Over the course of the last few years there has been a significant amount of research performed on ontology-based formalization of phenotype descriptions. The intrinsic value and knowledge captured within such descriptions can only be expressed by taking advantage of their inner structure that implicitly combines qualities and anatomical entities. We present a meta-model (the Phenotype Fragment Ontology) and a processing pipeline that enable together the automatic decomposition and conceptualization of phenotype descriptions for the human skeletal phenome. We use this approach to showcase the usefulness of the generic concept of phenotype decomposition by performing an experimental study on all skeletal phenotype concepts defined in the Human Phenotype Ontology.

  16. Occupational trichloroethylene hypersensitivity syndrome: human herpesvirus 6 reactivation and rash phenotypes.

    Science.gov (United States)

    Kamijima, Michihiro; Wang, Hailan; Yamanoshita, Osamu; Ito, Yuki; Xia, Lihua; Yanagiba, Yukie; Chen, Cishan; Okamura, Ai; Huang, Zhenlie; Qiu, Xinxiang; Song, Xiangrong; Cai, Tingfeng; Liu, Lili; Ge, Yichen; Deng, Yingyu; Naito, Hisao; Yoshikawa, Tetsushi; Tohyama, Mikiko; Li, Laiyu; Huang, Hanlin; Nakajima, Tamie

    2013-12-01

    Trichloroethylene (TCE) is an industrial solvent which can cause severe generalized dermatitis, i.e., occupational TCE hypersensitivity syndrome. Reactivation of latent human herpesvirus 6 (HHV6) can occur in such patients, which has made TCE known as a causative chemical of drug-induced hypersensitivity syndrome (DIHS). This study aimed to clarify HHV6 status, cytokine profiles and their association with rash phenotypes in patients with TCE hypersensitivity syndrome. HHV6 DNA copy numbers, anti-HHV6 antibody titers, and cytokines were measured in blood prospectively sampled 5-7 times from 28 hospitalized patients with the disease. The patients (19 had exfoliative dermatitis (ED) and 9 had non-ED type rash) generally met the diagnostic criteria for DIHS. Viral reactivation defined as increases in either HHV6 DNA (≥100 genomic copies/10(6) peripheral blood mononuclear cells) or antibody titers was identified in 24 (89%) patients. HHV6 DNA, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-5, IL-6 and IL-10 concentrations were remarkably higher in the patients than in the healthy workers (p<0.01). Positive correlations between HHV6 DNA, TNF-α, IFN-γ, IL-6 and IL-10 were significant (p<0.05) except for that between HHV6 DNA and IFN-γ. An increase in HHV6 DNA was positively associated with an increase in TNF-α on admission (p<0.01). HHV6 DNA, the antibody titers, TNF-α and IL-10 concentrations were significantly higher in ED than in the non-ED type (p<0.05). Reactivated HHV6 and the increased cytokines could be biomarkers of TCE hypersensitivity syndrome. The higher-level reactivation and stronger humoral responses were associated with ED-type rash. Copyright © 2013 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

  17. Comparative analysis of Edwardsiella isolates from fish in the eastern United States identifies two distinct genetic taxa amongst organisms phenotypically classified as E. tarda

    Science.gov (United States)

    Griffin, Matt J.; Quiniou, Sylvie M.; Cody, Theresa; Tabuchi, Maki; Ware, Cynthia; Cipriano, Rocco C.; Mauel, Michael J.; Soto, Esteban

    2013-01-01

    Edwardsiella tarda, a Gram-negative member of the family Enterobacteriaceae, has been implicated in significant losses in aquaculture facilities worldwide. Here, we assessed the intra-specific variability of E. tarda isolates from 4 different fish species in the eastern United States. Repetitive sequence mediated PCR (rep-PCR) using 4 different primer sets (ERIC I & II, ERIC II, BOX, and GTG5) and multi-locus sequence analysis of 16S SSU rDNA, groEl, gyrA, gyrB, pho, pgi, pgm, and rpoA gene fragments identified two distinct genotypes of E. tarda (DNA group I; DNA group II). Isolates that fell into DNA group II demonstrated more similarity to E. ictaluri than DNA group I, which contained the reference E. tarda strain (ATCC #15947). Conventional PCR analysis using published E. tarda-specific primer sets yielded variable results, with several primer sets producing no observable amplification of target DNA from some isolates. Fluorometric determination of G + C content demonstrated 56.4% G + C content for DNA group I, 60.2% for DNA group II, and 58.4% for E. ictaluri. Surprisingly, these isolates were indistinguishable using conventional biochemical techniques, with all isolates demonstrating phenotypic characteristics consistent with E. tarda. Analysis using two commercial test kits identified multiple phenotypes, although no single metabolic characteristic could reliably discriminate between genetic groups. Additionally, anti-microbial susceptibility and fatty acid profiles did not demonstrate remarkable differences between groups. The significant genetic variation (<90% similarity at gyrA, gyrB, pho, phi and pgm; <40% similarity by rep-PCR) between these groups suggests organisms from DNA group II may represent an unrecognized, genetically distinct taxa of Edwardsiella that is phenotypically indistinguishable from E. tarda.

  18. Predictive genomics: a cancer hallmark network framework for predicting tumor clinical phenotypes using genome sequencing data.

    Science.gov (United States)

    Wang, Edwin; Zaman, Naif; Mcgee, Shauna; Milanese, Jean-Sébastien; Masoudi-Nejad, Ali; O'Connor-McCourt, Maureen

    2015-02-01

    Tumor genome sequencing leads to documenting thousands of DNA mutations and other genomic alterations. At present, these data cannot be analyzed adequately to aid in the understanding of tumorigenesis and its evolution. Moreover, we have little insight into how to use these data to predict clinical phenotypes and tumor progression to better design patient treatment. To meet these challenges, we discuss a cancer hallmark network framework for modeling genome sequencing data to predict cancer clonal evolution and associated clinical phenotypes. The framework includes: (1) cancer hallmarks that can be represented by a few molecular/signaling networks. 'Network operational signatures' which represent gene regulatory logics/strengths enable to quantify state transitions and measures of hallmark traits. Thus, sets of genomic alterations which are associated with network operational signatures could be linked to the state/measure of hallmark traits. The network operational signature transforms genotypic data (i.e., genomic alterations) to regulatory phenotypic profiles (i.e., regulatory logics/strengths), to cellular phenotypic profiles (i.e., hallmark traits) which lead to clinical phenotypic profiles (i.e., a collection of hallmark traits). Furthermore, the framework considers regulatory logics of the hallmark networks under tumor evolutionary dynamics and therefore also includes: (2) a self-promoting positive feedback loop that is dominated by a genomic instability network and a cell survival/proliferation network is the main driver of tumor clonal evolution. Surrounding tumor stroma and its host immune systems shape the evolutionary paths; (3) cell motility initiating metastasis is a byproduct of the above self-promoting loop activity during tumorigenesis; (4) an emerging hallmark network which triggers genome duplication dominates a feed-forward loop which in turn could act as a rate-limiting step for tumor formation; (5) mutations and other genomic alterations have

  19. Genetic variants and early cigarette smoking and nicotine dependence phenotypes in adolescents.

    Directory of Open Access Journals (Sweden)

    Jennifer O'Loughlin

    Full Text Available While the heritability of cigarette smoking and nicotine dependence (ND is well-documented, the contribution of specific genetic variants to specific phenotypes has not been closely examined. The objectives of this study were to test the associations between 321 tagging single-nucleotide polymorphisms (SNPs that capture common genetic variation in 24 genes, and early smoking and ND phenotypes in novice adolescent smokers, and to assess if genetic predictors differ across these phenotypes.In a prospective study of 1294 adolescents aged 12-13 years recruited from ten Montreal-area secondary schools, 544 participants who had smoked at least once during the 7-8 year follow-up provided DNA. 321 single-nucleotide polymorphisms (SNPs in 24 candidate genes were tested for an association with number of cigarettes smoked in the past 3 months, and with five ND phenotypes (a modified version of the Fagerstrom Tolerance Questionnaire, the ICD-10 and three clusters of ND symptoms representing withdrawal symptoms, use of nicotine for self-medication, and a general ND/craving symptom indicator.The pattern of SNP-gene associations differed across phenotypes. Sixteen SNPs in seven genes (ANKK1, CHRNA7, DDC, DRD2, COMT, OPRM1, SLC6A3 (also known as DAT1 were associated with at least one phenotype with a p-value <0.01 using linear mixed models. After permutation and FDR adjustment, none of the associations remained statistically significant, although the p-values for the association between rs557748 in OPRM1 and the ND/craving and self-medication phenotypes were both 0.076.Because the genetic predictors differ, specific cigarette smoking and ND phenotypes should be distinguished in genetic studies in adolescents. Fifteen of the 16 top-ranked SNPs identified in this study were from loci involved in dopaminergic pathways (ANKK1/DRD2, DDC, COMT, OPRM1, and SLC6A3.Dopaminergic pathways may be salient during early smoking and the development of ND.

  20. Helicobacter pylori ATCC 43629/NCTC 11639 Outer Membrane Vesicles (OMVs) from Biofilm and Planktonic Phase Associated with Extracellular DNA (eDNA)

    Science.gov (United States)

    Grande, Rossella; Di Marcantonio, Maria C.; Robuffo, Iole; Pompilio, Arianna; Celia, Christian; Di Marzio, Luisa; Paolino, Donatella; Codagnone, Marilina; Muraro, Raffaella; Stoodley, Paul; Hall-Stoodley, Luanne; Mincione, Gabriella

    2015-01-01

    Helicobacter pylori persistence is associated with its capacity to develop biofilms as a response to changing environmental conditions and stress. Extracellular DNA (eDNA) is a component of H. pylori biofilm matrix but the lack of DNase I activity supports the hypothesis that eDNA might be protected by other extracellular polymeric substances (EPS) and/or Outer Membrane Vesicles (OMVs), which bleb from the bacteria surface during growth. The aim of the present study was to both identify the eDNA presence on OMVs segregated from H. pylori ATCC 43629/NCTC 11639 biofilm (bOMVs) and its planktonic phase (pOMVs) and to characterize the physical-chemical properties of the OMVs. The presence of eDNA in bOMVs and pOMVs was initially carried out using DNase I-gold complex labeling and Transmission Electron Microscope analysis (TEM). bOMVs and pOMVs were further isolated and physical-chemical characterization carried out using dynamic light scattering (DLS) analysis. eDNA associated with OMVs was detected and quantified using a PicoGreen spectrophotometer assay, while its extraction was performed with a DNA Kit. TEM images showed that eDNA was mainly associated with the OMV membrane surfaces; while PicoGreen staining showed a four-fold increase of dsDNA in bOMVs compared with pOMVs. The eDNA extracted from OMVs was visualized using gel electrophoresis. DLS analysis indicated that both planktonic and biofilm H. pylori phenotypes generated vesicles, with a broad distribution of sizes on the nanometer scale. The DLS aggregation assay suggested that eDNA may play a role in the aggregation of OMVs, in the biofilm phenotype. Moreover, the eDNA associated with vesicle membrane may impede DNase I activity on H. pylori biofilms. These results suggest that OMVs derived from the H. pylori biofilm phenotype may play a structural role by preventing eDNA degradation by nucleases, providing a bridging function between eDNA strands on OMV surfaces and promoting aggregation. PMID:26733944

  1. Myopathic mtDNA Depletion Syndrome Due to Mutation in TK2 Gene.

    Science.gov (United States)

    Martín-Hernández, Elena; García-Silva, María Teresa; Quijada-Fraile, Pilar; Rodríguez-García, María Elena; Rivera, Henry; Hernández-Laín, Aurelio; Coca-Robinot, David; Fernández-Toral, Joaquín; Arenas, Joaquín; Martín, Miguel A; Martínez-Azorín, Francisco

    2017-01-01

    Whole-exome sequencing was used to identify the disease gene(s) in a Spanish girl with failure to thrive, muscle weakness, mild facial weakness, elevated creatine kinase, deficiency of mitochondrial complex III and depletion of mtDNA. With whole-exome sequencing data, it was possible to get the whole mtDNA sequencing and discard any pathogenic variant in this genome. The analysis of whole exome uncovered a homozygous pathogenic mutation in thymidine kinase 2 gene ( TK2; NM_004614.4:c.323 C>T, p.T108M). TK2 mutations have been identified mainly in patients with the myopathic form of mtDNA depletion syndromes. This patient presents an atypical TK2-related myopathic form of mtDNA depletion syndromes, because despite having a very low content of mtDNA (TK2 gene in mtDNA depletion syndromes and expanded the phenotypic spectrum.

  2. Physiological differences and changes in global DNA methylation levels in Agave angustifolia Haw. albino variant somaclones during the micropropagation process.

    Science.gov (United States)

    Duarte-Aké, Fátima; Castillo-Castro, Eduardo; Pool, Felipe Barredo; Espadas, Francisco; Santamaría, Jorge M; Robert, Manuel L; De-la-Peña, Clelia

    2016-12-01

    Global DNA methylation changes caused by in vitro conditions are associated with the subculturing and phenotypic variation in Agave angustifolia Haw. While the relationship between the development of albinism and in vitro culture is well documented, the role of epigenetic processes in this development leaves some important questions unanswered. During the micropropagation of Agave angustifolia Haw., we found three different phenotypes, green (G), variegated (V) and albino (A). To understand the physiological and epigenetic differences among the somaclones, we analyzed several morphophysiological parameters and changes in the DNA methylation patterns in the three phenotypes during their in vitro development. We found that under in vitro conditions, the V plantlets maintained their CAM photosynthetic capacity, while the A variant showed no pigments and lost its CAM photosynthetic ability. Epigenetic analysis revealed that global DNA methylation increased in the G phenotype during the first two subcultures. However, after that time, DNA methylation levels declined. This hypomethylation correlated with the appearance of V shoots in the G plantlets. A similar correlation occurred in the V phenotype, where an increase of 2 % in the global DNA methylation levels was correlated with the generation of A shoots in the V plantlets. This suggests that an "epigenetic stress memory" during in vitro conditions causes a chromatin shift that favors the generation of variegated and albino shoots.

  3. Advanced phenotyping and phenotype data analysis for the plant growth and development study

    Directory of Open Access Journals (Sweden)

    Md. Matiur eRahaman

    2015-08-01

    Full Text Available Due to increase in the consumption of food, feed, fuel and to ensure global food security for rapidly growing human population, there is need to breed high yielding crops that can adapt to future climate. To solve these global issues, novel approaches are required to provide quantitative phenotypes to elucidate the genetic basis of agriculturally import traits and to screen germplasm with super performance in function under resource-limited environment. At present, plant phenomics has offered and integrated suite technologies for understanding the complete set of phenotypes of plants, towards the progression of the full characteristics of plants with whole sequenced genomes. In this aspect, high-throughput phenotyping platforms have been developed that enables to capture extensive and intensive phenotype data from non-destructive imaging over time. These developments advance our view on plant growth and performance with responses to the changing climate and environment. In this paper, we present a brief review on currently developed high-throughput plant phenotyping infrastructures based on imaging techniques and corresponding principles for phenotype data analysis.

  4. ABO blood group phenotype frequency estimation using molecular phenotyping in rhesus and cynomolgus macaques.

    Science.gov (United States)

    Kanthaswamy, S; Ng, J; Oldt, R F; Valdivia, L; Houghton, P; Smith, D G

    2017-11-01

    A much larger sample (N = 2369) was used to evaluate a previously reported distribution of the A, AB and B blood group phenotypes in rhesus and cynomolgus macaques from six different regional populations. These samples, acquired from 15 different breeding and research facilities in the United States, were analyzed using a real-time quantitative polymerase chain reaction (qPCR) assay that targets single nucleotide polymorphisms (SNPs) responsible for the macaque A, B and AB phenotypes. The frequency distributions of blood group phenotypes of the two species differ significantly from each other and significant regional differentiation within the geographic ranges of each species was also observed. The B blood group phenotype was prevalent in rhesus macaques, especially those from India, while the frequencies of the A, B and AB phenotypes varied significantly among cynomolgus macaques from different geographic regions. The Mauritian cynomolgus macaques, despite having originated in Indonesia, showed significant (P ≪ .01) divergence from the Indonesian animals at the ABO blood group locus. Most Mauritian animals belonged to the B blood group while the Indonesian animals were mostly A. The close similarity in blood group frequency distributions between the Chinese rhesus and Indochinese cynomolgus macaques demonstrates that the introgression between these two species extends beyond the zone of intergradation in Indochina. This study underscores the importance of ABO blood group phenotyping of the domestic supply of macaques and their biospecimens. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  5. Elucidating the genotype–phenotype map by automatic enumeration and analysis of the phenotypic repertoire

    Science.gov (United States)

    Lomnitz, Jason G; Savageau, Michael A

    2015-01-01

    Background: The gap between genotype and phenotype is filled by complex biochemical systems most of which are poorly understood. Because these systems are complex, it is widely appreciated that quantitative understanding can only be achieved with the aid of mathematical models. However, formulating models and measuring or estimating their numerous rate constants and binding constants is daunting. Here we present a strategy for automating difficult aspects of the process. Methods: The strategy, based on a system design space methodology, is applied to a class of 16 designs for a synthetic gene oscillator that includes seven designs previously formulated on the basis of experimentally measured and estimated parameters. Results: Our strategy provides four important innovations by automating: (1) enumeration of the repertoire of qualitatively distinct phenotypes for a system; (2) generation of parameter values for any particular phenotype; (3) simultaneous realization of parameter values for several phenotypes to aid visualization of transitions from one phenotype to another, in critical cases from functional to dysfunctional; and (4) identification of ensembles of phenotypes whose expression can be phased to achieve a specific sequence of functions for rationally engineering synthetic constructs. Our strategy, applied to the 16 designs, reproduced previous results and identified two additional designs capable of sustained oscillations that were previously missed. Conclusions: Starting with a system’s relatively fixed aspects, its architectural features, our method enables automated analysis of nonlinear biochemical systems from a global perspective, without first specifying parameter values. The examples presented demonstrate the efficiency and power of this automated strategy. PMID:26998346

  6. Elucidating the genotype-phenotype map by automatic enumeration and analysis of the phenotypic repertoire.

    Science.gov (United States)

    Lomnitz, Jason G; Savageau, Michael A

    The gap between genotype and phenotype is filled by complex biochemical systems most of which are poorly understood. Because these systems are complex, it is widely appreciated that quantitative understanding can only be achieved with the aid of mathematical models. However, formulating models and measuring or estimating their numerous rate constants and binding constants is daunting. Here we present a strategy for automating difficult aspects of the process. The strategy, based on a system design space methodology, is applied to a class of 16 designs for a synthetic gene oscillator that includes seven designs previously formulated on the basis of experimentally measured and estimated parameters. Our strategy provides four important innovations by automating: (1) enumeration of the repertoire of qualitatively distinct phenotypes for a system; (2) generation of parameter values for any particular phenotype; (3) simultaneous realization of parameter values for several phenotypes to aid visualization of transitions from one phenotype to another, in critical cases from functional to dysfunctional; and (4) identification of ensembles of phenotypes whose expression can be phased to achieve a specific sequence of functions for rationally engineering synthetic constructs. Our strategy, applied to the 16 designs, reproduced previous results and identified two additional designs capable of sustained oscillations that were previously missed. Starting with a system's relatively fixed aspects, its architectural features, our method enables automated analysis of nonlinear biochemical systems from a global perspective, without first specifying parameter values. The examples presented demonstrate the efficiency and power of this automated strategy.

  7. Regulation of the activity of the dual-function DnaA protein in Caulobacter crescentus.

    Directory of Open Access Journals (Sweden)

    Carmen Fernandez-Fernandez

    Full Text Available DnaA is a conserved essential bacterial protein that acts as the initiator of chromosomal replication as well as a master transcriptional regulator in Caulobacter crescentus. Thus, the intracellular levels of active DnaA need to be tightly regulated during the cell cycle. Our previous work suggested that DnaA may be regulated at the level of its activity by the replisome-associated protein HdaA. Here, we describe the construction of a mutant DnaA protein [DnaA(R357A]. The R357 residue in the AAA+ domain of the C. crescentus DnaA protein is equivalent to the R334 residue of the E. coli DnaA protein, which is required for the Regulatory Inactivation of DnaA (RIDA. We found that the expression of the DnaA(R357A mutant protein in C. crescentus, but not the expression of the wild-type DnaA protein at similar levels, causes a severe phenotype of over-initiation of chromosomal replication and that it blocks cell division. Thus, the mutant DnaA(R357A protein is hyper-active to promote the initiation of DNA replication, compared to the wild-type DnaA protein. DnaA(R357A could not replace DnaA in vivo, indicating that the switch in DnaA activity once chromosomal replication has started may be an essential process in C. crescentus. We propose that the inactivation of DnaA is the main mechanism ensuring that chromosomal replication starts only once per cell cycle. We further observed that the R357A substitution in DnaA does not promote the activity of DnaA as a direct transcriptional activator of four important genes, encoding HdaA, the GcrA master cell cycle regulator, the FtsZ cell division protein and the MipZ spatial regulator of cell division. Thus, the AAA+ domain of DnaA may play a role in temporally regulating the bifunctionality of DnaA by reallocating DnaA molecules from initiating DNA replication to transcribing genes within the unique DnaA regulon of C. crescentus.

  8. Extensive genetic and DNA methylation variation contribute to heterosis in triploid loquat hybrids.

    Science.gov (United States)

    Liu, Chao; Wang, Mingbo; Wang, Lingli; Guo, Qigao; Liang, Guolu

    2018-04-24

    We aim to overcome the unclear origin of the loquat and elucidate the heterosis mechanism of the triploid loquat. Here we investigated the genetic and epigenetic variations between the triploid plant and its parental lines using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplified fragment length polymorphism (MSAP) analyses. We show that in addition to genetic variations, extensive DNA methylation variation occurred during the formation process of triploid loquat, with the triploid hybrid having increased DNA methylation compared to the parents. Furthermore, a correlation existed between genetic variation and DNA methylation remodeling, suggesting that genome instability may lead to DNA methylation variation or vice versa. Sequence analysis of the MSAP bands revealed that over 53% of them overlap with protein-coding genes, which may indicate a functional role of the differential DNA methylation in gene regulation and hence heterosis phenotypes. Consistent with this, the genetic and epigenetic alterations were associated closely to the heterosis phenotypes of triploid loquat, and this association varied for different traits. Our results suggested that the formation of triploid is accompanied by extensive genetic and DNA methylation variation, and these changes contribute to the heterosis phenotypes of the triploid loquats from the two cross lines.

  9. Nanoparticle-mediated rhodopsin cDNA but not intron-containing DNA delivery causes transgene silencing in a rhodopsin knockout model.

    Science.gov (United States)

    Zheng, Min; Mitra, Rajendra N; Filonov, Nazar A; Han, Zongchao

    2016-03-01

    Previously, we compared the efficacy of nanoparticle (NP)-mediated intron-containing rhodopsin (sgRho) vs. intronless cDNA in ameliorating retinal disease phenotypes in a rhodopsin knockout (RKO) mouse model of retinitis pigmentosa. We showed that NP-mediated sgRho delivery achieved long-term expression and phenotypic improvement in RKO mice, but not NP housing cDNA. However, the protein level of the NP-sgRho construct was only 5-10% of wild-type at 8 mo postinjection. To have a better understanding of the reduced levels of long-term expression of the vectors, in the present study, we evaluated the epigenetic changes of subretinal delivering NP-cDNA vs. NP-sgRho in the RKO mouse eyes. Following the administration, DNA methylation and histone status of specific regions (bacteria plasmid backbone, promoter, rhodopsin gene, and scaffold/matrix attachment region) of the vectors were evaluated at various time points. We documented that epigenetic transgene silencing occurred in vector-mediated gene transfer, which were caused by the plasmid backbone and the cDNA of the transgene, but not the intron-containing transgene. No toxicity or inflammation was found in the treated eyes. Our results suggest that cDNA of the rhodopsin transgene and bacteria backbone interfered with the host defense mechanism of DNA methylation-mediated transgene silencing through heterochromatin-associated modifications. © FASEB.

  10. Cardiobacterium valvarum infective endocarditis and phenotypic/molecular characterization of 11 Cardiobacterium species strains

    DEFF Research Database (Denmark)

    Chen, Ming; Kemp, Michael; Bruun, Niels E

    2011-01-01

    Cardiobacterium valvarum is a newly recognized human pathogen related to infective endocarditis. Cardiobacterium species are, however, only rarely the aetiology of infective endocarditis. An infective endocarditis case is presented and, additionally, phenotypic and phylogenetic comparison...... of a further 10 collection strains, representing the two species within the genus, was performed. C. valvarum was isolated from the blood and DNA was present in valvular tissue (partial 16S rRNA gene analysis) from a 64-year-old man with infective endocarditis of the mitral valve, rupture of chordae...

  11. Senescent phenotypes of skin fibroblasts from patients with Tangier disease

    International Nuclear Information System (INIS)

    Matsuura, Fumihiko; Hirano, Ken-ichi; Ikegami, Chiaki; Sandoval, Jose C.; Oku, Hiroyuki; Yuasa-Kawase, Miyako; Tsubakio-Yamamoto, Kazumi; Koseki, Masahiro; Masuda, Daisaku; Tsujii, Ken-ichi; Ishigami, Masato; Nishida, Makoto; Shimomura, Iichiro; Hori, Masatsugu; Yamashita, Shizuya

    2007-01-01

    Tangier disease (TD) is characterized by a deficiency of high density lipoprotein (HDL) in plasma and patients with TD have an increased risk for coronary artery disease (CAD). Recently, we reported that fibroblasts from TD exhibited large and flattened morphology, which is often observed in senescent cells. On the other hand, data have accumulated to show the relationship between cellular senescence and development of atherosclerotic CAD. The aim of the present study was to investigate whether TD fibroblasts exhibited cellular senescence. The proliferation of TD fibroblasts was gradually decreased at population doubling level (PDL) ∼10 compared with control cells. TD cells practically ceased proliferation at PDL ∼30. DNA synthesis was markedly decreased in TD fibroblasts. TD cells exhibited a higher positive rate for senescence-associated β-galactosidase (SA-β-gal), which is one of the biomarkers of cellular senescence in vitro. These data showed that TD cells reached cellular senescence at an earlier PDL compared with controls. Although, there was no difference in the telomere length of fibroblasts between TD and controls at the earlier passage (PDL 6), the telomere length of TD cells was shorter than that of controls at the late passage (PDL 25). Taken together, the current study demonstrates that the late-passaged TD fibroblasts showed senescent phenotype in vitro, which might be related to the increased cardiovascular manifestations in TD patients

  12. [Phenotypic and molecular characterization of a Colombian family with phenylketonuria].

    Science.gov (United States)

    Gélvez, Nancy; Acosta, Johana; López, Greizy; Castro, Derly; Prieto, Juan Carlos; Bermúdez, Martha; Tamayo, Marta L

    2016-09-01

    Phenylketonuria is a metabolic disorder characterized by severe neurological involvement and behavioral disorder, whose early diagnosis enables an effective treatment to avoid disease sequelae, thus changing the prognosis. Objective: To characterize a family with phenylketonuria in Colombia at clinical, biochemical and molecular levels. Materials and methods: The population consisted of seven individuals of a consanguineous family with four children with suggestive symptoms of phenylketonuria. After signing an informed consent, blood and urine samples were taken for colorimetric tests and high performance liquid and thin layer chromatographies. DNA extraction and sequencing of the 13 exons of the PAH gene were performed in all subjects. We designed primers for each exon with the Primer 3 software using automatic sequencing equipment Abiprism 3100 Avant. Sequences were analyzed using the SeqScape, v2.0, software. Results: We described the clinical and molecular characteristics of a Colombian family with phenylketonuria and confirmed the presence of the mutation c.398_401delATCA. We established a genotype-phenotype correlation, highlighting the interesting clinical variability found among the affected patients despite having the same mutation in all of them. Conclusions: Early recognition of this disease is very important to prevent its neurological and psychological sequelae, given that patients reach old age without diagnosis or proper management.

  13. A Marfan syndrome gene expression phenotype in cultured skin fibroblasts

    Directory of Open Access Journals (Sweden)

    Emond Mary

    2007-09-01

    Full Text Available Abstract Background Marfan syndrome (MFS is a heritable connective tissue disorder caused by mutations in the fibrillin-1 gene. This syndrome constitutes a significant identifiable subtype of aortic aneurysmal disease, accounting for over 5% of ascending and thoracic aortic aneurysms. Results We used spotted membrane DNA macroarrays to identify genes whose altered expression levels may contribute to the phenotype of the disease. Our analysis of 4132 genes identified a subset with significant expression differences between skin fibroblast cultures from unaffected controls versus cultures from affected individuals with known fibrillin-1 mutations. Subsequently, 10 genes were chosen for validation by quantitative RT-PCR. Conclusion Differential expression of many of the validated genes was associated with MFS samples when an additional group of unaffected and MFS affected subjects were analyzed (p-value -6 under the null hypothesis that expression levels in cultured fibroblasts are unaffected by MFS status. An unexpected observation was the range of individual gene expression. In unaffected control subjects, expression ranges exceeding 10 fold were seen in many of the genes selected for qRT-PCR validation. The variation in expression in the MFS affected subjects was even greater.

  14. CHARGE and Kabuki syndromes: a phenotypic and molecular link.

    Science.gov (United States)

    Schulz, Yvonne; Freese, Luisa; Mänz, Johanna; Zoll, Barbara; Völter, Christiane; Brockmann, Knut; Bögershausen, Nina; Becker, Jutta; Wollnik, Bernd; Pauli, Silke

    2014-08-15

    CHARGE syndrome is a complex developmental disorder caused by mutations in the chromodomain helicase DNA-binding gene CHD7. Kabuki syndrome, another developmental disorder, is characterized by typical facial features in combination with developmental delay, short stature, prominent digit pads and visceral abnormalities. Mutations in the KMT2D gene, which encodes a H3K4 histone methyltransferase, are the major cause of Kabuki syndrome. Here, we report a patient, who was initially diagnosed with CHARGE syndrome based on the spectrum of inner organ malformations like choanal hypoplasia, heart defect, anal atresia, vision problems and conductive hearing impairment. While sequencing and MLPA analysis of all coding exons of CHD7 revealed no pathogenic mutation, sequence analysis of the KMT2D gene identified the heterozygous de novo nonsense mutation c.5263C > T (p.Gln1755*). Thus, our patient was diagnosed with Kabuki syndrome. By using co-immunoprecipitation, immunohistochemistry and direct yeast two hybrid assays, we could show that, like KMT2D, CHD7 interacts with members of the WAR complex, namely WDR5, ASH2L and RbBP5. We therefore propose that CHD7 and KMT2D function in the same chromatin modification machinery, thus pointing out a mechanistic connection, and presenting a probable explanation for the phenotypic overlap between Kabuki and CHARGE syndromes. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  15. Senescent intervertebral disc cells exhibit perturbed matrix homeostasis phenotype.

    Science.gov (United States)

    Ngo, Kevin; Patil, Prashanti; McGowan, Sara J; Niedernhofer, Laura J; Robbins, Paul D; Kang, James; Sowa, Gwendolyn; Vo, Nam

    2017-09-01

    Aging greatly increases the risk for intervertebral disc degeneration (IDD) as a result of proteoglycan loss due to reduced synthesis and enhanced degradation of the disc matrix proteoglycan (PG). How disc matrix PG homeostasis becomes perturbed with age is not known. The goal of this study is to determine whether cellular senescence is a source of this perturbation. We demonstrated that disc cellular senescence is dramatically increased in the DNA repair-deficient Ercc1 -/Δ mouse model of human progeria. In these accelerated aging mice, increased disc cellular senescence is closely associated with the rapid loss of disc PG. We also directly examine PG homeostasis in oxidative damage-induced senescent human cells using an in vitro cell culture model system. Senescence of human disc cells treated with hydrogen peroxide was confirmed by growth arrest, senescence-associated β-galactosidase activity, γH2AX foci, and acquisition of senescence-associated secretory phenotype. Senescent human disc cells also exhibited perturbed matrix PG homeostasis as evidenced by their decreased capacity to synthesize new matrix PG and enhanced degradation of aggrecan, a major matrix PG. of the disc. Our in vivo and in vitro findings altogether suggest that disc cellular senescence is an important driver of PG matrix homeostatic perturbation and PG loss. Published by Elsevier B.V.

  16. Fe65 is required for Tip60-directed histone H4 acetylation at DNA strand breaks

    Science.gov (United States)

    Stante, Maria; Minopoli, Giuseppina; Passaro, Fabiana; Raia, Maddalena; Vecchio, Luigi Del; Russo, Tommaso

    2009-01-01

    Fe65 is a binding partner of the Alzheimer's β-amyloid precursor protein APP. The possible involvement of this protein in the cellular response to DNA damage was suggested by the observation that Fe65 null mice are more sensitive to genotoxic stress than WT counterpart. Fe65 associated with chromatin under basal conditions and its involvement in DNA damage repair requires this association. A known partner of Fe65 is the histone acetyltransferase Tip60. Considering the crucial role of Tip60 in DNA repair, we explored the hypothesis that the phenotype of Fe65 null cells depended on its interaction with Tip60. We demonstrated that Fe65 knockdown impaired recruitment of Tip60-TRRAP complex to DNA double strand breaks and decreased histone H4 acetylation. Accordingly, the efficiency of DNA repair was decreased upon Fe65 suppression. To explore whether APP has a role in this mechanism, we analyzed a Fe65 mutant unable to bind to APP. This mutant failed to rescue the phenotypes of Fe65 null cells; furthermore, APP/APLP2 suppression results in the impairment of recruitment of Tip60-TRRAP complex to DNA double strand breaks, decreased histone H4 acetylation and repair efficiency. On these bases, we propose that Fe65 and its interaction with APP play an important role in the response to DNA damage by assisting the recruitment of Tip60-TRRAP to DNA damage sites. PMID:19282473

  17. The expanding phenotype of mitochondrial myopathy.

    Science.gov (United States)

    DiMauro, Salvatore; Gurgel-Giannetti, Juliana

    2005-10-01

    Our understanding of mitochondrial diseases (defined restrictively as defects in the mitochondrial respiratory chain) continues to progress apace. In this review we provide an update of information regarding disorders that predominantly or exclusively affect skeletal muscle. Most recently described mitochondrial myopathies are due to defects in nuclear DNA, including coenzyme Q10 deficiency, and mutations in genes that control mitochondrial DNA (mtDNA) abundance and structure such as POLG and TK2. Barth syndrome, an X-linked recessive mitochondrial myopathy/cardiopathy, is associated with altered lipid composition of the inner mitochondrial membrane, but a putative secondary impairment of the respiratory chain remains to be documented. Concerning the 'other genome', the role played by mutations in protein encoding genes of mtDNA in causing isolated myopathies has been confirmed. It has also been confirmed that mutations in tRNA genes of mtDNA can cause predominantly myopathic syndromes and - contrary to conventional wisdom - these mutations can be homoplasmic. Defects in the mitochondrial respiratory chain impair energy production and almost invariably involve skeletal muscle, causing exercise intolerance, myalgia, cramps, or fixed weakness, which often affects extraocular muscles and results in droopy eyelids (ptosis) and progressive external ophthalmoplegia.

  18. rDNA Copy Number Variants Are Frequent Passenger Mutations in Saccharomyces cerevisiae Deletion Collections and de Novo Transformants

    Directory of Open Access Journals (Sweden)

    Elizabeth X. Kwan

    2016-09-01

    Full Text Available The Saccharomyces cerevisiae ribosomal DNA (rDNA locus is known to exhibit greater instability relative to the rest of the genome. However, wild-type cells preferentially maintain a stable number of rDNA copies, suggesting underlying genetic control of the size of this locus. We performed a screen of a subset of the Yeast Knock-Out (YKO single gene deletion collection to identify genetic regulators of this locus and to determine if rDNA copy number correlates with yeast replicative lifespan. While we found no correlation between replicative lifespan and rDNA size, we identified 64 candidate strains with significant rDNA copy number differences. However, in the process of validating candidate rDNA variants, we observed that independent isolates of our de novo gene deletion strains had unsolicited but significant changes in rDNA copy number. Moreover, we were not able to recapitulate rDNA phenotypes from the YKO yeast deletion collection. Instead, we found that the standard lithium acetate transformation protocol is a significant source of rDNA copy number variation, with lithium acetate exposure being the treatment causing variable rDNA copy number events after transformation. As the effects of variable rDNA copy number are being increasingly reported, our finding that rDNA is affected by lithium acetate exposure suggested that rDNA copy number variants may be influential passenger mutations in standard strain construction in S. cerevisiae.

  19. rDNA Copy Number Variants Are Frequent Passenger Mutations in Saccharomyces cerevisiae Deletion Collections and de Novo Transformants.

    Science.gov (United States)

    Kwan, Elizabeth X; Wang, Xiaobin S; Amemiya, Haley M; Brewer, Bonita J; Raghuraman, M K

    2016-09-08

    The Saccharomyces cerevisiae ribosomal DNA (rDNA) locus is known to exhibit greater instability relative to the rest of the genome. However, wild-type cells preferentially maintain a stable number of rDNA copies, suggesting underlying genetic control of the size of this locus. We performed a screen of a subset of the Yeast Knock-Out (YKO) single gene deletion collection to identify genetic regulators of this locus and to determine if rDNA copy number correlates with yeast replicative lifespan. While we found no correlation between replicative lifespan and rDNA size, we identified 64 candidate strains with significant rDNA copy number differences. However, in the process of validating candidate rDNA variants, we observed that independent isolates of our de novo gene deletion strains had unsolicited but significant changes in rDNA copy number. Moreover, we were not able to recapitulate rDNA phenotypes from the YKO yeast deletion collection. Instead, we found that the standard lithium acetate transformation protocol is a significant source of rDNA copy number variation, with lithium acetate exposure being the treatment causing variable rDNA copy number events after transformation. As the effects of variable rDNA copy number are being increasingly reported, our finding that rDNA is affected by lithium acetate exposure suggested that rDNA copy number variants may be influential passenger mutations in standard strain construction in S. cerevisiae. Copyright © 2016 Kwan et al.

  20. Nanostructures via DNA scaffold metallization

    OpenAIRE

    Ning, C.; Zinchenko, A.; Baigl, D.; Pyshkina, O.; Sergeyev, V.; Endo, Kazunaka; Yoshikawa, K.

    2005-01-01

    The critical role of polymers in process of noble metals nanostructures formation is well known, however, the use of DNA chain template in this process is yet largely unknown. In this study we demonstrate different ways of silver deposition on DNA template and report the influence of silver nanostructures formation on DNA conformational state. Metallization of DNA chain proceeds by two different scenarios depending on DNA conformation. If DNA chain is unfolded (elongated) chain, silver reduct...

  1. New genes as drivers of phenotypic evolution

    Science.gov (United States)

    Chen, Sidi; Krinsky, Benjamin H.; Long, Manyuan

    2014-01-01

    During the course of evolution, genomes acquire novel genetic elements as sources of functional and phenotypic diversity, including new genes that originated in recent evolution. In the past few years, substantial progress has been made in understanding the evolution and phenotypic effects of new genes. In particular, an emerging picture is that new genes, despite being present in the genomes of only a subset of species, can rapidly evolve indispensable roles in fundamental biological processes, including development, reproduction, brain function and behaviour. The molecular underpinnings of how new genes can develop these roles are starting to be characterized. These recent discoveries yield fresh insights into our broad understanding of biological diversity at refined resolution. PMID:23949544

  2. Animal biometrics: quantifying and detecting phenotypic appearance.

    Science.gov (United States)

    Kühl, Hjalmar S; Burghardt, Tilo

    2013-07-01

    Animal biometrics is an emerging field that develops quantified approaches for representing and detecting the phenotypic appearance of species, individuals, behaviors, and morphological traits. It operates at the intersection between pattern recognition, ecology, and information sciences, producing computerized systems for phenotypic measurement and interpretation. Animal biometrics can benefit a wide range of disciplines, including biogeography, population ecology, and behavioral research. Currently, real-world applications are gaining momentum, augmenting the quantity and quality of ecological data collection and processing. However, to advance animal biometrics will require integration of methodologies among the scientific disciplines involved. Such efforts will be worthwhile because the great potential of this approach rests with the formal abstraction of phenomics, to create tractable interfaces between different organizational levels of life. Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Smooth muscle cell phenotypic switching in stroke.

    Science.gov (United States)

    Poittevin, Marine; Lozeron, Pierre; Hilal, Rose; Levy, Bernard I; Merkulova-Rainon, Tatiana; Kubis, Nathalie

    2014-06-01

    Disruption of cerebral blood flow after stroke induces cerebral tissue injury through multiple mechanisms that are not yet fully understood. Smooth muscle cells (SMCs) in blood vessel walls play a key role in cerebral blood flow control. Cerebral ischemia triggers these cells to switch to a phenotype that will be either detrimental or beneficial to brain repair. Moreover, SMC can be primarily affected genetically or by toxic metabolic molecules. After stroke, this pathological phenotype has an impact on the incidence, pattern, severity, and outcome of the cerebral ischemic disease. Although little research has been conducted on the pathological role and molecular mechanisms of SMC in cerebrovascular ischemic diseases, some therapeutic targets have already been identified and could be considered for further pharmacological development. We examine these different aspects in this review.

  4. DNA damage and polyploidization.

    Science.gov (United States)

    Chow, Jeremy; Poon, Randy Y C

    2010-01-01

    A growing body of evidence indicates that polyploidization triggers chromosomal instability and contributes to tumorigenesis. DNA damage is increasingly being recognized for its roles in promoting polyploidization. Although elegant mechanisms known as the DNA damage checkpoints are responsible for halting the cell cycle after DNA damage, agents that uncouple the checkpoints can induce unscheduled entry into mitosis. Likewise, defects of the checkpoints in several disorders permit mitotic entry even in the presence of DNA damage. Forcing cells with damaged DNA into mitosis causes severe chromosome segregation defects, including lagging chromosomes, chromosomal fragments and chromosomal bridges. The presence of these lesions in the cleavage plane is believed to abort cytokinesis. It is postulated that if cytokinesis failure is coupled with defects of the p53-dependent postmitotic checkpoint pathway, cells can enter S phase and become polyploids. Progress in the past several years has unraveled some of the underlying principles of these pathways and underscored the important role of DNA damage in polyploidization. Furthermore, polyploidization per se may also be an important determinant of sensitivity to DNA damage, thereby may offer an opportunity for novel therapies.

  5. Quantitive DNA Fiber Mapping

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Chun-Mei; Wang, Mei; Greulich-Bode, Karin M.; Weier, Jingly F.; Weier, Heinz-Ulli G.

    2008-01-28

    Several hybridization-based methods used to delineate single copy or repeated DNA sequences in larger genomic intervals take advantage of the increased resolution and sensitivity of free chromatin, i.e., chromatin released from interphase cell nuclei. Quantitative DNA fiber mapping (QDFM) differs from the majority of these methods in that it applies FISH to purified, clonal DNA molecules which have been bound with at least one end to a solid substrate. The DNA molecules are then stretched by the action of a receding meniscus at the water-air interface resulting in DNA molecules stretched homogeneously to about 2.3 kb/{micro}m. When non-isotopically, multicolor-labeled probes are hybridized to these stretched DNA fibers, their respective binding sites are visualized in the fluorescence microscope, their relative distance can be measured and converted into kilobase pairs (kb). The QDFM technique has found useful applications ranging from the detection and delineation of deletions or overlap between linked clones to the construction of high-resolution physical maps to studies of stalled DNA replication and transcription.

  6. Regulating DNA Self-assembly by DNA-Surface Interactions.

    Science.gov (United States)

    Liu, Longfei; Li, Yulin; Wang, Yong; Zheng, Jianwei; Mao, Chengde

    2017-12-14

    DNA self-assembly provides a powerful approach for preparation of nanostructures. It is often studied in bulk solution and involves only DNA-DNA interactions. When confined to surfaces, DNA-surface interactions become an additional, important factor to DNA self-assembly. However, the way in which DNA-surface interactions influence DNA self-assembly is not well studied. In this study, we showed that weak DNA-DNA interactions could be stabilized by DNA-surface interactions to allow large DNA nanostructures to form. In addition, the assembly can be conducted isothermally at room temperature in as little as 5 seconds. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Phenotype development in TgHD minipigs

    Czech Academy of Sciences Publication Activity Database

    Ellederová, Zdeňka; Vidinská, Daniela; Mačáková, Monika; Kučerová, S.; Bohuslavová, Božena; Sedláčková, M.; Lišková, Irena; Valeková, Ivona; Baxa, Monika; Ardan, Taras; Juhás, Štefan; Motlík, Jan

    2015-01-01

    Roč. 78, Suppl 2 (2015), s. 11-11 ISSN 1210-7859. [Conference on Animal Models for neurodegenerative Diseases /3./. 08.11.2015-10.11.2015, Liblice] R&D Projects: GA MŠk ED2.1.00/03.0124; GA MŠk(CZ) 7F14308 Institutional support: RVO:67985904 Keywords : phenotype * minipig model of Huntington ´s disease * reproductive failure Subject RIV: EB - Genetics ; Molecular Biology

  8. Connectomic intermediate phenotypes for psychiatric disorders

    Directory of Open Access Journals (Sweden)

    Alex eFornito

    2012-04-01

    Full Text Available Psychiatric disorders are phenotypically heterogeneous entities with a complex genetic basis. To mitigate this complexity, many investigators study so-called intermediate phenotypes that putatively provide a more direct index of the physiological effects of candidate genetic risk variants than overt psychiatric syndromes. Magnetic resonance imaging (MRI is a particularly popular technique for measuring such phenotypes because it allows interrogation of diverse aspects of brain structure and function in vivo. Much of this work however, has focused on relatively simple measures that quantify variations in the physiology or tissue integrity of specific brain regions in isolation, contradicting an emerging consensus that most major psychiatric disorders do not arise from isolated dysfunction in one or a few brain regions, but rather from disturbed interactions within and between distributed neural circuits; i.e., they are disorders of brain connectivity. The recent proliferation of new MRI techniques for comprehensively mapping the entire connectivity architecture of the brain, termed the human connectome, has provided a rich repertoire of tools for understanding how genetic variants implicated in mental disorder impact distinct neural circuits. In this article, we review research using these connectomic techniques to understand how genetic variation influences the connectivity and topology of human brain networks. We highlight recent evidence from twin and imaging genetics studies suggesting that the penetrance of candidate risk variants for mental illness, such as those in SLC6A4, MAOA, ZNF804A and APOE, may be higher for intermediate phenotypes characterised at the level of distributed neural systems than at the level of spatially localised brain regions. The findings indicate that imaging connectomics provides a powerful framework for understanding how genetic risk for psychiatric disease is expressed through altered structure and function of

  9. NF1 Neuronal Genotype Phenotype Relationships

    Science.gov (United States)

    2017-06-01

    interesting results from the Drosophila functional assays, at present we have decided to focus our attention on selected NF1 patient missense mutations...complexity of NF1 disease phenotypes in different tissues, age and sex dependency of symptoms, impact of environmental factors and genetic heterogeneity...suggesting the role of modifier genes [12]. This work aims to shed light on this issue by studying the functional consequences of selected NF1

  10. HIV coreceptor phenotyping in the clinical setting.

    Science.gov (United States)

    Low, Andrew J; Swenson, Luke C; Harrigan, P Richard

    2008-01-01

    The introduction of CCR5 antagonists increases the options available for constructing antiretroviral regimens. However, this option is coupled with the caveat that patients should be tested for HIV coreceptor tropism prior to initiating CCR5 antagonist-based therapy. Failure to screen for CXCR4 usage increases the risk of using an ineffective drug, thus reducing the likelihood of viral suppression and increasing their risk for developing antiretroviral resistance. This review discusses current and future methods of determining HIV tropism, with a focus on their utility in the clinical setting for screening purposes. Some of these methods include recombinant phenotypic tests, such as the Monogram Trofile assay, as well as genotype-based predictors, heteroduplex tracking assays, and flow cytometry based methods. Currently, the best evidence supports the use of phenotypic methods, although other methods of screening for HIV coreceptor usage prior to the administration of CCR5 antagonists may reduce costs and increase turnaround time over phenotypic methods. The presence of low levels of X4 virus is a challenge to all assay methods, resulting in reduced sensitivity in clinical, patient-derived samples when compared to clonally derived samples. Gaining a better understanding of the output of these assays and correlating them with clinical progression and therapy response will provide some indication on how both genotype-based, and phenotypic assays for determining HIV coreceptor usage can be improved. In addition, leveraging new technologies capable of detecting low-level minority species may provide the most significant advances in ensuring that individuals with low levels of dual/mixed tropic virus are not inadvertently prescribed CCR5 antagonists.

  11. One gene, many phenotypes | Shawky | Egyptian Journal of Medical ...

    African Journals Online (AJOL)

    ... mechanisms underlying genotype-phenotype discrepancies is important, as it will move clinical genetics towards predictive medicine, allowing better selection of therapeutic strategies and individualized counseling of persons affected with genetic disorders. Keywords: Gene, phenotype, mosaicism, epigenetics, pleiotropy ...

  12. Chimeric proteins for detection and quantitation of DNA mutations, DNA sequence variations, DNA damage and DNA mismatches

    Science.gov (United States)

    McCutchen-Maloney, Sandra L.

    2002-01-01

    Chimeric proteins having both DNA mutation binding activity and nuclease activity are synthesized by recombinant technology. The proteins are of the general formula A-L-B and B-L-A where A is a peptide having DNA mutation binding activity, L is a linker and B is a peptide having nuclease activity. The chimeric proteins are useful for detection and identification of DNA sequence variations including DNA mutations (including DNA damage and mismatches) by binding to the DNA mutation and cutting the DNA once the DNA mutation is detected.

  13. Phenotypic Variability in the Coccolithophore Emiliania huxleyi.

    Science.gov (United States)

    Blanco-Ameijeiras, Sonia; Lebrato, Mario; Stoll, Heather M; Iglesias-Rodriguez, Debora; Müller, Marius N; Méndez-Vicente, Ana; Oschlies, Andreas

    2016-01-01

    Coccolithophores are a vital part of oceanic phytoplankton assemblages that produce organic matter and calcium carbonate (CaCO3) containing traces of other elements (i.e. Sr and Mg). Their associated carbon export from the euphotic zone to the oceans' interior plays a crucial role in CO2 feedback mechanisms and biogeochemical cycles. The coccolithophore Emiliania huxleyi has been widely studied as a model organism to understand physiological, biogeochemical, and ecological processes in marine sciences. Here, we show the inter-strain variability in physiological and biogeochemical traits in 13 strains of E. huxleyi from various biogeographical provinces obtained from culture collections commonly used in the literature. Our results demonstrate that inter-strain genetic variability has greater potential to induce larger phenotypic differences than the phenotypic plasticity of single strains cultured under a broad range of variable environmental conditions. The range of variation found in physiological parameters and calcite Sr:Ca highlights the need to reconsider phenotypic variability in paleoproxy calibrations and model parameterizations to adequately translate findings from single strain laboratory experiments to the real ocean.

  14. Phenotypic Variability in the Coccolithophore Emiliania huxleyi.

    Directory of Open Access Journals (Sweden)

    Sonia Blanco-Ameijeiras

    Full Text Available Coccolithophores are a vital part of oceanic phytoplankton assemblages that produce organic matter and calcium carbonate (CaCO3 containing traces of other elements (i.e. Sr and Mg. Their associated carbon export from the euphotic zone to the oceans' interior plays a crucial role in CO2 feedback mechanisms and biogeochemical cycles. The coccolithophore Emiliania huxleyi has been widely studied as a model organism to understand physiological, biogeochemical, and ecological processes in marine sciences. Here, we show the inter-strain variability in physiological and biogeochemical traits in 13 strains of E. huxleyi from various biogeographical provinces obtained from culture collections commonly used in the literature. Our results demonstrate that inter-strain genetic variability has greater potential to induce larger phenotypic differences than the phenotypic plasticity of single strains cultured under a broad range of variable environmental conditions. The range of variation found in physiological parameters and calcite Sr:Ca highlights the need to reconsider phenotypic variability in paleoproxy calibrations and model parameterizations to adequately translate findings from single strain laboratory experiments to the real ocean.

  15. A simple phenotypic classification for celiac disease

    Directory of Open Access Journals (Sweden)

    Ajit Sood

    2018-04-01

    Full Text Available Background/Aims : Celiac disease is a global health problem. The presentation of celiac disease has unfolded over years and it is now known that it can manifest at different ages, has varied presentations, and is prone to develop complications, if not managed properly. Although the Oslo definitions provide consensus on the various terminologies used in literature, there is no phenotypic classification providing a composite diagnosis for the disease. Methods : Various variables identified for phenotypic classification included age at diagnosis, age at onset of symptoms, clinical presentation, family history and complications. These were applied to the existing registry of 1,664 patients at Dayanand Medical College and Hospital, Ludhiana, India. In addition, age was evaluated as below 15 and below 18 years. Cross tabulations were used for the verification of the classification using the existing data. Expert opinion was sought from both international and national experts of varying fields. Results : After empirical verification, age at diagnosis was considered appropriate in between A1 (<18 and A2 (≧18. The disease presentation has been classified into 3 types–P1 (classical, P2 (non-classical and P3 (asymptomatic. Complications were considered as absent (C0 or present (C1. A single phenotypic classification based on these 3 characteristics, namely age at the diagnosis, clinical presentation, and intestinal complications (APC classification was derived. Conclusions : APC classification (age at diagnosis, presentation, complications is a simple disease explanatory classification for patients with celiac disease aimed at providing a composite diagnosis.

  16. Dissecting phenotypic variation among AIS patients

    International Nuclear Information System (INIS)

    Wang Minghua; Wang Jiucun; Zhang Zhen; Zhao Zhimin; Zhang Rongmei; Hu Xiaohua; Tan Tao; Luo Shijing; Luo Zewei

    2005-01-01

    We have created genital skin fibroblast cell lines directly from three patients in a Chinese family affected by androgen insensitivity syndrome (AIS). All patients in the family share an identical AR Arg 840 Cys mutant but show different disease phenotypes. By using the cell lines, we find that the mutation has not influenced a normal androgen-binding capacity at 37 deg C but has reduced the affinity for androgens and may cause thermolability of the androgen-receptor complex. The impaired nuclear trafficking of the androgen receptor in the cell lines is highly correlated with the severity of donors' disease phenotype. The transactivity of the mutant is substantially weakened and the extent of the reduced transactivity reflects severity of the donors' disease symptom. Our data reveal that although etiology of AIS is monogenic and the mutant may alter the major biological functions of its wild allele, the function of the mutant AR can also be influenced by the different genetic backgrounds and thus explains the divergent disease phenotypes

  17. The phenotypic spectrum of congenital Zika syndrome.

    Science.gov (United States)

    Del Campo, Miguel; Feitosa, Ian M L; Ribeiro, Erlane M; Horovitz, Dafne D G; Pessoa, André L S; França, Giovanny V A; García-Alix, Alfredo; Doriqui, Maria J R; Wanderley, Hector Y C; Sanseverino, Maria V T; Neri, João I C F; Pina-Neto, João M; Santos, Emerson S; Verçosa, Islane; Cernach, Mirlene C S P; Medeiros, Paula F V; Kerbage, Saile C; Silva, André A; van der Linden, Vanessa; Martelli, Celina M T; Cordeiro, Marli T; Dhalia, Rafael; Vianna, Fernanda S L; Victora, Cesar G; Cavalcanti, Denise P; Schuler-Faccini, Lavinia

    2017-04-01

    In October 2015, Zika virus (ZIKV) outbreak the Brazilian Ministry of Health (MoH). In response, the Brazilian Society of Medical Genetics established a task force (SBGM-ZETF) to study the phenotype of infants born with microcephaly due to ZIKV congenital infection and delineate the phenotypic spectrum of this newly recognized teratogen. This study was based on the clinical evaluation and neuroimaging of 83 infants born during the period from July, 2015 to March, 2016 and registered by the SBGM-ZETF. All 83 infants had significant findings on neuroimaging consistent with ZIKV congenital infection and 12 had confirmed ZIKV IgM in CSF. A recognizable phenotype of microcephaly, anomalies of the shape of skull and redundancy of the scalp consistent with the Fetal Brain Disruption Sequence (FBDS) was present in 70% of infants, but was most often subtle. In addition, features consistent with fetal immobility, ranging from dimples (30.1%), distal hand/finger contractures (20.5%), and feet malpositions (15.7%), to generalized arthrogryposis (9.6%), were present in these infants. Some cases had milder microcephaly or even a normal head circumference (HC), and other less distinctive findings. The detailed observation of the dysmorphic and neurologic features in these infants provides insight into the mechanisms and timings of the brain disruption and the sequence of developmental anomalies that may occur after prenatal infection by the ZIKV. © 2017 Wiley Periodicals, Inc.

  18. Associations between phenotypes of preeclampsia and thrombophilia.

    Science.gov (United States)

    Berks, Durk; Duvekot, Johannes J; Basalan, Hillal; De Maat, Moniek P M; Steegers, Eric A P; Visser, Willy

    2015-11-01

    Preeclampsia complicates 2-8% of all pregnancies. Studies on the association of preeclampsia with thrombophilia are conflicting. Clinical heterogeneity of the disease may be one of the explanations. The present study addresses the question whether different phenotypes of preeclampsia are associated with thrombophilia factors. Study design We planned a retrospective cohort study. From 1985 until 2010 women with preeclampsia were offered postpartum screening for the following thrombophilia factors: anti-phospholipid antibodies, APC-resistance, protein C deficiency and protein S deficiency, hyperhomocysteineamia, factor V Leiden and Prothrombin gene mutation. Hospital records were used to obtain information on phenotypes of the preeclampsia and placental histology. We identified 844 women with singleton pregnancies who were screened for thrombophilia factors. HELLP complicated 49% of pregnancies; Fetal growth restriction complicated 61% of pregnancies. Early delivery (preeclampsia was associated with protein S deficiency (p=0.01). Fetal growth restriction was associated with anti-phospholipid antibodies (ppreeclampsia was associated with anti-phospholipid antibodies (p=0.01). Extensive placental infarction (>10%) was associated with anti-phospholipid antibodies (ppreeclampsia, especially if complicated by fetal growth restriction, are associated with anti-phospholipid antibodies. Other phenotypes of preeclampsia, especially HELLP syndrome, were not associated with thrombophilia. We advise only to test for anti-phospholipid antibodies after early onset preeclampsia, especially if complicated by fetal growth restriction. We suggest enough evidence is presented to justify no further studies are needed. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  19. Aberrant phenotypes in peripheral T cell lymphomas.

    Science.gov (United States)

    Hastrup, N; Ralfkiaer, E; Pallesen, G

    1989-01-01

    Seventy six peripheral T cell lymphomas were examined immunohistologically to test their reactivity with a panel of monoclonal antibodies against 11 T cell associated antigens (CD1-8, CD27, UCHL1, and the T cell antigen receptor). Sixty two (82%) lymphomas showed aberrant phenotypes, and four main categories were distinguished as follows: (i) lack of one or several pan-T cell antigens (49, 64% of the cases); (ii) loss of both the CD4 and CD8 antigens (11, 15% of the cases); (iii) coexpression of the CD4 and CD8 antigens (13, 17% of the cases); and (iv) expression of the CD1 antigen (eight, 11% of the cases). No correlation was seen between the occurrence of aberrant phenotypes and the histological subtype. It is concluded that the demonstration of an aberrant phenotype is a valuable supplement to histological assessment in the diagnosis of peripheral T cell lymphomas. It is recommended that the panel of monoclonal antibodies against T cell differentiation antigens should be fairly large, as apparently any antigen may be lost in the process of malignant transformation. Images Figure PMID:2469701

  20. Cluster analysis in phenotyping a Portuguese population.

    Science.gov (United States)

    Loureiro, C C; Sa-Couto, P; Todo-Bom, A; Bousquet, J

    2015-09-03

    Unbiased cluster analysis using clinical parameters has identified asthma phenotypes. Adding inflammatory biomarkers to this analysis provided a better insight into the disease mechanisms. This approach has not yet been applied to asthmatic Portuguese patients. To identify phenotypes of asthma using cluster analysis in a Portuguese asthmatic population treated in secondary medical care. Consecutive patients with asthma were recruited from the outpatient clinic. Patients were optimally treated according to GINA guidelines and enrolled in the study. Procedures were performed according to a standard evaluation of asthma. Phenotypes were identified by cluster analysis using Ward's clustering method. Of the 72 patients enrolled, 57 had full data and were included for cluster analysis. Distribution was set in 5 clusters described as follows: cluster (C) 1, early onset mild allergic asthma; C2, moderate allergic asthma, with long evolution, female prevalence and mixed inflammation; C3, allergic brittle asthma in young females with early disease onset and no evidence of inflammation; C4, severe asthma in obese females with late disease onset, highly symptomatic despite low Th2 inflammation; C5, severe asthma with chronic airflow obstruction, late disease onset and eosinophilic inflammation. In our study population, the identified clusters were mainly coincident with other larger-scale cluster analysis. Variables such as age at disease onset, obesity, lung function, FeNO (Th2 biomarker) and disease severity were important for cluster distinction. Copyright © 2015. Published by Elsevier España, S.L.U.

  1. "Artifactual" arsenate DNA

    DEFF Research Database (Denmark)

    Nielsen, Peter E

    2012-01-01

    The recent claim by Wolfe-Simon et al. that the Halomonas bacterial strain GFAJ-1 when grown in arsenate-containing medium with limiting phosphate is able to substitute phosphate with arsenate in biomolecules including nucleic acids and in particular DNA(1) arose much skepticism, primarily due...... to the very limited chemical stability of arsenate esters (see ref. 2 and references therein). A major part of the criticisms was concerned with the insufficient (bio)chemical evidence in the Wolfe-Simon study for the actual chemical incorporation of arsenate in DNA (and/or RNA). Redfield et al. now present...... evidence that the identification of arsenate DNA was artifactual....

  2. The genotype-phenotype map of an evolving digital organism

    OpenAIRE

    Fortuna, Miguel A.; Zaman, Luis; Ofria, Charles; Wagner, Andreas

    2017-01-01

    To understand how evolving systems bring forth novel and useful phenotypes, it is essential to understand the relationship between genotypic and phenotypic change. Artificial evolving systems can help us understand whether the genotype-phenotype maps of natural evolving systems are highly unusual, and it may help create evolvable artificial systems. Here we characterize the genotype-phenotype map of digital organisms in Avida, a platform for digital evolution. We consider digital organisms fr...

  3. EMPReSS: European mouse phenotyping resource for standardized screens.

    Science.gov (United States)

    Green, Eain C J; Gkoutos, Georgios V; Lad, Heena V; Blake, Andrew; Weekes, Joseph; Hancock, John M

    2005-06-15

    Standardized phenotyping protocols are essential for the characterization of phenotypes so that results are comparable between different laboratories and phenotypic data can be related to ontological descriptions in an automated manner. We describe a web-based resource for the visualization, searching and downloading of standard operating procedures and other documents, the European Mouse Phenotyping Resource for Standardized Screens-EMPReSS. Direct access: http://www.empress.har.mrc.ac.uk e.green@har.mrc.ac.uk.

  4. RNA-Mediated cis Regulation in Acinetobacter baumannii Modulates Stress-Induced Phenotypic Variation.

    Science.gov (United States)

    Ching, Carly; Gozzi, Kevin; Heinemann, Björn; Chai, Yunrong; Godoy, Veronica G

    2017-06-01

    In the nosocomial opportunistic pathogen Acinetobacter baumannii , RecA-dependent mutagenesis, which causes antibiotic resistance acquisition, is linked to the DNA damage response (DDR). Notably, unlike the Escherichia coli paradigm, recA and DDR gene expression in A. baumannii is bimodal. Namely, there is phenotypic variation upon DNA damage, which may provide a bet-hedging strategy for survival. Thus, understanding recA gene regulation is key to elucidate the yet unknown DDR regulation in A. baumannii Here, we identify a structured 5' untranslated region (UTR) in the recA transcript which serves as a cis -regulatory element. We show that a predicted stem-loop structure in this 5' UTR affects mRNA half-life and underlies bimodal gene expression and thus phenotypic variation in response to ciprofloxacin treatment. We furthermore show that the stem-loop structure of the recA 5' UTR influences intracellular RecA protein levels and, in vivo , impairing the formation of the stem-loop structure of the recA 5' UTR lowers cell survival of UV treatment and decreases rifampin resistance acquisition from DNA damage-induced mutagenesis. We hypothesize that the 5' UTR allows for stable recA transcripts during stress, including antibiotic treatment, enabling cells to maintain suitable RecA levels for survival. This innovative strategy to regulate the DDR in A. baumannii may contribute to its success as a pathogen. IMPORTANCE Acinetobacter baumannii is an opportunistic pathogen quickly gaining antibiotic resistances. Mutagenesis and antibiotic resistance acquisition are linked to the DNA damage response (DDR). However, how the DDR is regulated in A. baumannii remains unknown, since unlike most bacteria, A. baumannii does not follow the regulation of the Escherichia coli paradigm. In this study, we have started to uncover the mechanisms regulating the novel A. baumannii DDR. We have found that a cis -acting 5' UTR regulates recA transcript stability, RecA protein levels, and DNA

  5. Syndromic (phenotypic diarrhea in early infancy

    Directory of Open Access Journals (Sweden)

    Bodemer Christine

    2008-02-01

    Full Text Available Abstract Syndromic diarrhea (SD, also known as phenotypic diarrhea (PD or tricho-hepato-enteric syndrome (THE, is a congenital enteropathy presenting with early-onset of severe diarrhea requiring parenteral nutrition (PN. To date, no epidemiological data are available. The estimated prevalence is approximately 1/300,000–400,000 live births in Western Europe. Ethnic origin does not appear to be associated with SD. Infants are born small for gestational age and present with facial dysmorphism including prominent forehead and cheeks, broad nasal root and hypertelorism. Hairs are woolly, easily removed and poorly pigmented. Severe and persistent diarrhea starts within the first 6 months of life (≤ 1 month in most cases and is accompanied by severe malabsorption leading to early and relentless protein energy malnutrition with failure to thrive. Liver disease affects about half of patients with extensive fibrosis or cirrhosis. There is currently no specific biochemical profile, though a functional T-cell immune deficiency with defective antibody production was reported. Microscopic analysis of the hair show twisted hair (pili torti, aniso- and poilkilotrichosis, and trichorrhexis nodosa. Histopathological analysis of small intestine biopsy shows non-specific villous atrophy with low or no mononuclear cell infiltration of the lamina propria, and no specific histological abnormalities involving the epithelium. The etiology remains unknown. The frequent association of the disorder with parental consanguinity and/or affected siblings suggests a genetic origin with an autosomal recessive mode of transmission. Early management consists of total PN. Some infants have a rather milder phenotype with partial PN dependency or require only enteral feeding. Prognosis of this syndrome is poor, but most patients now survive, and about half of the patients may be weaned from PN at adolescence, but experience failure to thrive and final short stature. Disease name

  6. Predictable Phenotypes of Antibiotic Resistance Mutations.

    Science.gov (United States)

    Knopp, M; Andersson, D I

    2018-05-15

    Antibiotic-resistant bacteria represent a major threat to our ability to treat bacterial infections. Two factors that determine the evolutionary success of antibiotic resistance mutations are their impact on resistance level and the fitness cost. Recent studies suggest that resistance mutations commonly show epistatic interactions, which would complicate predictions of their stability in bacterial populations. We analyzed 13 different chromosomal resistance mutations and 10 host strains of Salmonella enterica and Escherichia coli to address two main questions. (i) Are there epistatic interactions between different chromosomal resistance mutations? (ii) How does the strain background and genetic distance influence the effect of chromosomal resistance mutations on resistance and fitness? Our results show that the effects of combined resistance mutations on resistance and fitness are largely predictable and that epistasis remains rare even when up to four mutations were combined. Furthermore, a majority of the mutations, especially target alteration mutations, demonstrate strain-independent phenotypes across different species. This study extends our understanding of epistasis among resistance mutations and shows that interactions between different resistance mutations are often predictable from the characteristics of the individual mutations. IMPORTANCE The spread of antibiotic-resistant bacteria imposes an urgent threat to public health. The ability to forecast the evolutionary success of resistant mutants would help to combat dissemination of antibiotic resistance. Previous studies have shown that the phenotypic effects (fitness and resistance level) of resistance mutations can vary substantially depending on the genetic context in which they occur. We conducted a broad screen using many different resistance mutations and host strains to identify potential epistatic interactions between various types of resistance mutations and to determine the effect of strain

  7. DNA from keratinous tissue

    DEFF Research Database (Denmark)

    Bengtsson, Camilla F.; Olsen, Maja E.; Brandt, Luise Ørsted

    2011-01-01

    Keratinous tissues such as nail, hair, horn, scales and feather have been used as a source of DNA for over 20 years. Particular benefits of such tissues include the ease with which they can be sampled, the relative stability of DNA in such tissues once sampled, and, in the context of ancient...... genetic analyses, the fact that sampling generally causes minimal visual damage to valuable specimens. Even when freshly sampled, however, the DNA quantity and quality in the fully keratinized parts of such tissues is extremely poor in comparison to other tissues such as blood and muscle – although little...... systematic research has been undertaken to characterize how such degradation may relate to sample source. In this review paper we present the current understanding of the quality and limitations of DNA in two key keratinous tissues, nail and hair. The findings indicate that although some fragments of nuclear...

  8. DNA fusion gene vaccines

    DEFF Research Database (Denmark)

    Holst, Peter Johannes; Bassi, Maria Rosaria; Thomsen, Allan Randrup

    2010-01-01

    DNA vaccines are versatile and safe, but limited immunogenicity has prevented their use in the clinical setting. Experimentally, immunogenicity may be enhanced by the use of new delivery technologies, by coadministration of cytokines and pathogen-associated molecular patterns, or by fusion...... of antigens into molecular domains that enhance antigen presentation. More specifically, the immunogenicity of DNA vaccines may benefit from increased protein synthesis, increased T-cell help and MHC class I presentation, and the addition of a range of specific cytokines and pathogen-associated molecular...... with viral-vectored vaccines, various synergistic components may need to be incorporated into DNA vaccines. From the perspective of the future clinical use of DNA vaccines, it has been suggested that antigen presentation should be improved and cytokine coadministration attempted. However, even...

  9. DNA Sampling Hook

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The DNA Sampling Hook is a significant improvement on a method of obtaining a tissue sample from a live fish in situ from an aquatic environment. A tissue sample...

  10. Retroviral DNA Integration

    Science.gov (United States)

    2016-01-01

    The integration of a DNA copy of the viral RNA genome into host chromatin is the defining step of retroviral replication. This enzymatic process is catalyzed by the virus-encoded integrase protein, which is conserved among retroviruses and LTR-retrotransposons. Retroviral integration proceeds via two integrase activities: 3′-processing of the viral DNA ends, followed by the strand transfer of the processed ends into host cell chromosomal DNA. Herein we review the molecular mechanism of retroviral DNA integration, with an emphasis on reaction chemistries and architectures of the nucleoprotein complexes involved. We additionally discuss the latest advances on anti-integrase drug development for the treatment of AIDS and the utility of integrating retroviral vectors in gene therapy applications. PMID:27198982

  11. DNA damage and carcinogenesis

    International Nuclear Information System (INIS)

    Stelow, R.B.

    1980-01-01

    Although cancer may arise as a result of many different types of molecular changes, there is little reason to doubt that changes to DNA are one of the more important ones in cancer initiation. Although DNA repair mechanisms seem able to eliminate a very large fraction of deleterious changes to DNA, we not only have little insight into the molecular mechanisms involved in such repair, but have a negligible amount of information to permit us to estimate the shape of dose response relations at low doses. The case of skin cancer is a special one, in that the average population is exposed to sufficient solar uv so that the effects of small increments in uv dose may be estimated. An approximate 85% reduction in DNA repair increases skin cancer incidence 10 4 fold

  12. DNA-Origami

    DEFF Research Database (Denmark)

    Voigt, Niels Vinther; Tørring, Thomas; Gothelf, Kurt Vesterager

    2010-01-01

    DNA-nanostrukturer giver nye muligheder for studier af individuelle molekyler. Ved at udnytte DNAs unikke selvsamlende egenskaber kan man designe systemer, hvorpå der kan studeres kemiske reaktioner, fluoroforer og biiomolekyler på enkeltmolekyle-niveau....

  13. DNA Microarray Technology

    Science.gov (United States)

    Skip to main content DNA Microarray Technology Enter Search Term(s): Español Research Funding An Overview Bioinformatics Current Grants Education and Training Funding Extramural Research News Features Funding Divisions Funding ...

  14. DNA sequencing conference, 2

    Energy Technology Data Exchange (ETDEWEB)

    Cook-Deegan, R.M. [Georgetown Univ., Kennedy Inst. of Ethics, Washington, DC (United States); Venter, J.C. [National Inst. of Neurological Disorders and Strokes, Bethesda, MD (United States); Gilbert, W. [Harvard Univ., Cambridge, MA (United States); Mulligan, J. [Stanford Univ., CA (United States); Mansfield, B.K. [Oak Ridge National Lab., TN (United States)

    1991-06-19

    This conference focused on DNA sequencing, genetic linkage mapping, physical mapping, informatics and bioethics. Several were used to study this sequencing and mapping. This article also discusses computer hardware and software aiding in the mapping of genes.

  15. Close encounters with DNA

    Science.gov (United States)

    Maffeo, C.; Yoo, J.; Comer, J.; Wells, D. B.; Luan, B.; Aksimentiev, A.

    2014-01-01

    Over the past ten years, the all-atom molecular dynamics method has grown in the scale of both systems and processes amenable to it and in its ability to make quantitative predictions about the behavior of experimental systems. The field of computational DNA research is no exception, witnessing a dramatic increase in the size of systems simulated with atomic resolution, the duration of individual simulations and the realism of the simulation outcomes. In this topical review, we describe the hallmark physical properties of DNA from the perspective of all-atom simulations. We demonstrate the amazing ability of such simulations to reveal the microscopic physical origins of experimentally observed phenomena and we review the frustrating limitations associated with imperfections of present atomic force fields and inadequate sampling. The review is focused on the following four physical properties of DNA: effective electric charge, response to an external mechanical force, interaction with other DNA molecules and behavior in an external electric field. PMID:25238560

  16. Gomphid DNA sequence data

    Data.gov (United States)

    U.S. Environmental Protection Agency — DNA sequence data for several genetic loci. This dataset is not publicly accessible because: It's already publicly available on GenBank. It can be accessed through...

  17. HPV DNA test

    Science.gov (United States)

    ... test; Cancer of cervix - HPV DNA test References Hacker NF. Cervical dysplasia and cancer. In: Hacker NF, Gambone JC, Hobel CJ, eds. Hacker and Moore's Essentials of Obstetrics and Gynecology . 6th ...

  18. Close encounters with DNA.

    Science.gov (United States)

    Maffeo, C; Yoo, J; Comer, J; Wells, D B; Luan, B; Aksimentiev, A

    2014-10-15

    Over the past ten years, the all-atom molecular dynamics method has grown in the scale of both systems and processes amenable to it and in its ability to make quantitative predictions about the behavior of experimental systems. The field of computational DNA research is no exception, witnessing a dramatic increase in the size of systems simulated with atomic resolution, the duration of individual simulations and the realism of the simulation outcomes. In this topical review, we describe the hallmark physical properties of DNA from the perspective of all-atom simulations. We demonstrate the amazing ability of such simulations to reveal the microscopic physical origins of experimentally observed phenomena. We also discuss the frustrating limitations associated with imperfections of present atomic force fields and inadequate sampling. The review is focused on the following four physical properties of DNA: effective electric charge, response to an external mechanical force, interaction with other DNA molecules and behavior in an external electric field.

  19. FBI's DNA analysis program

    Science.gov (United States)

    Brown, John R.

    1994-03-01

    Forensic DNA profiling technology is a significant law enforcement tool due to its superior discriminating power. Applying the principles of population genetics to the DNA profile obtained in violent crime investigations results in low frequency of occurrence estimates for the DNA profile. These estimates often range from a frequency of occurrence of 1 in 50 unrelated individuals to 1 in a million unrelated individuals or even smaller. It is this power to discriminate among individuals in the population that has propelled forensic DNA technology to the forefront of forensic testing in violent crime cases. Not only is the technology extremely powerful in including or excluding a criminal suspect as the perpetrator, but it also gives rise to the potential of identifying criminal suspects in cases where the investigators of unknown suspect cases have exhausted all other available leads.

  20. Making DNA Fingerprints.

    Science.gov (United States)

    Nunley, Kathie F.

    1996-01-01

    Presents an activity to simulate electrophoresis using everyday items. Uses adding machine paper to construct a set of DNA fingerprints that can be used to solve crime cases designed by students in any biology class. (JRH)

  1. Radiation damage in DNA

    International Nuclear Information System (INIS)

    Lafleur, V.

    1978-01-01

    A number of experiments are described with the purpose to obtain a better insight in the chemical nature and the biological significance of radiation-induced damage in DNA, with some emphasis on the significance of alkali-labile sites. It is shown that not only reactions of OH radicals but also of H radicals introduce breaks and other inactivating damage in single-standed phiX174 DNA. It is found that phosphate buffer is very suitable for the study of the reactions of H radicals with DNA, as the H 2 PO 4 - ions convert the hydrated electrons into H radicals. The hydrated electron, which does react with DNA, does not cause a detectable inactivation. (Auth.)

  2. Diagnosis of becker muscular dystrophy: Results of Re-analysis of DNA samples

    NARCIS (Netherlands)

    Straathof, Chiara S. M.; van Heusden, Dave; Ippel, Pieternella F.; Post, Jan G.; Voermans, Nicol C.; de Visser, Marianne; Brusse, Esther; van den Bergen, Janneke C.; van der Kooi, Anneke J.; Verschuuren, Jan J. G. M.; Ginjaar, Hendrika B.

    2016-01-01

    The phenotype of Becker muscular dystrophy (BMD) is highly variable, and the disease may be underdiagnosed. We searched for new mutations in the DMD gene in a cohort of previously undiagnosed patients who had been referred in the period 1985-1995. All requests for DNA analysis of the DMD gene in

  3. Histone H3 lysine 56 acetylation and the response to DNA replication fork damage

    DEFF Research Database (Denmark)

    Wurtele, Hugo; Kaiser, Gitte Schalck; Bacal, Julien

    2012-01-01

    but are only mildly affected by hydroxyurea. We demonstrate that, after exposure to MMS, H3K56ac-deficient cells cannot complete DNA replication and eventually segregate chromosomes with intranuclear foci containing the recombination protein Rad52. In addition, we provide evidence that these phenotypes...

  4. An Internet-Accessible DNA Sequence Database for Identifying Fusaria from Human and Animal Infections

    Science.gov (United States)

    Because less than one-third of clinically relevant fusaria can be accurately identified to species level using phenotypic data (i.e., morphological species recognition), we constructed a three-locus DNA sequence database to facilitate molecular identification of the 69 Fusarium species associated wi...

  5. DNA to DNA transcription might exist in eukaryotic cells

    OpenAIRE

    Li, Gao-De

    2016-01-01

    Till now, in biological sciences, the term, transcription, mainly refers to DNA to RNA transcription. But our recently published experimental findings obtained from Plasmodium falciparum strongly suggest the existence of DNA to DNA transcription in the genome of eukaryotic cells, which could shed some light on the functions of certain noncoding DNA in the human and other eukaryotic genomes.

  6. Patterning nanocrystals using DNA

    Energy Technology Data Exchange (ETDEWEB)

    Williams, Shara Carol [Univ. of California, Berkeley, CA (United States)

    2003-01-01

    One of the goals of nanotechnology is to enable programmed self-assembly of patterns made of various materials with nanometer-sized control. This dissertation describes the results of experiments templating arrangements of gold and semiconductor nanocrystals using 2'-deoxyribonucleic acid (DNA). Previously, simple DNA-templated linear arrangements of two and three nanocrystals structures have been made.[1] Here, we have sought to assemble larger and more complex nanostructures. Gold-DNA conjugates with 50 to 100 bases self-assembled into planned arrangements using strands of DNA containing complementary base sequences. We used two methods to increase the complexity of the arrangements: using branched synthetic doublers within the DNA covalent backbone to create discrete nanocrystal groupings, and incorporating the nanocrystals into a previously developed DNA lattice structure [2][3] that self-assembles from tiles made of DNA double-crossover molecules to create ordered nanoparticle arrays. In the first project, the introduction of a covalently-branched synthetic doubler reagent into the backbone of DNA strands created a branched DNA ''trimer.'' This DNA trimer templated various structures that contained groupings of three and four gold nanoparticles, giving promising, but inconclusive transmission electron microscopy (TEM) results. Due to the presence of a variety of possible structures in the reaction mixtures, and due to the difficulty of isolating the desired structures, the TEM and gel electrophoresis results for larger structures having four particles, and for structures containing both 5 and 10 nm gold nanoparticles were inconclusive. Better results may come from using optical detection methods, or from improved sample preparation. In the second project, we worked toward making two-dimensional ordered arrays of nanocrystals. We replicated and improved upon previous results for making DNA lattices, increasing the size of the lattices

  7. Das DNA-Puzzle

    Science.gov (United States)

    Kirchner, Stefan

    Im Jahre 1953 wurde von James Watson und Francis Crick erstmalig der strukturelle Aufbau der sogenannten DNA (Desoxyribonukleinsäure) beschrieben, welche das Erbgut jedes Lebewesens enthält. Der wesentliche Teil des Erbguts wird dabei durch eine sehr lange Folge der vier Basen Adenin (A), Cytosin (C), Guanin (G) und Thymin (T) codiert. Seit einigen Jahren ist es möglich, die Folge der vier Basen zu einer gegebenen DNA zu bestimmen. Biologen bezeichnen diesen Vorgang als Sequenzierung.

  8. PDA: Pooled DNA analyzer

    Directory of Open Access Journals (Sweden)

    Lin Chin-Yu

    2006-04-01

    Full Text Available Abstract Background Association mapping using abundant single nucleotide polymorphisms is a powerful tool for identifying disease susceptibility genes for complex traits and exploring possible genetic diversity. Genotyping large numbers of SNPs individually is performed routinely but is cost prohibitive for large-scale genetic studies. DNA pooling is a reliable and cost-saving alternative genotyping method. However, no software has been developed for complete pooled-DNA analyses, including data standardization, allele frequency estimation, and single/multipoint DNA pooling association tests. This motivated the development of the software, 'PDA' (Pooled DNA Analyzer, to analyze pooled DNA data. Results We develop the software, PDA, for the analysis of pooled-DNA data. PDA is originally implemented with the MATLAB® language, but it can also be executed on a Windows system without installing the MATLAB®. PDA provides estimates of the coefficient of preferential amplification and allele frequency. PDA considers an extended single-point association test, which can compare allele frequencies between two DNA pools constructed under different experimental conditions. Moreover, PDA also provides novel chromosome-wide multipoint association tests based on p-value combinations and a sliding-window concept. This new multipoint testing procedure overcomes a computational bottleneck of conventional haplotype-oriented multipoint methods in DNA pooling analyses and can handle data sets having a large pool size and/or large numbers of polymorphic markers. All of the PDA functions are illustrated in the four bona fide examples. Conclusion PDA is simple to operate and does not require that users have a strong statistical background. The software is available at http://www.ibms.sinica.edu.tw/%7Ecsjfann/first%20flow/pda.htm.

  9. Racemic DNA Crystallography

    OpenAIRE

    Mandal , Pradeep K.; Collie , Gavin W.; Kauffmann , Brice; Huc , Ivan

    2014-01-01

    International audience; Racemates increase the chances of crystallization by allowing molecular contacts to be formed in a greater number of ways. With the advent of protein synthesis, the production of protein racemates and racemic-protein crystallography are now possible. Curiously, racemic DNA crystallography had not been investigated despite the commercial availability of Land D-deoxyribo-oligonucleotides. Here, we report a study into racemic DNA crystallography showing the strong propens...

  10. Celebrating DNA's Repair Crew.

    Science.gov (United States)

    Kunkel, Thomas A

    2015-12-03

    This year, the Nobel Prize in Chemistry has been awarded to Tomas Lindahl, Aziz Sancar, and Paul Modrich for their seminal studies of the mechanisms by which cells from bacteria to man repair DNA damage that is generated by normal cellular metabolism and stress from the environment. These studies beautifully illustrate the remarkable power of DNA repair to influence life from evolution through disease susceptibility. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. GGCX-Associated Phenotypes: An Overview in Search of Genotype-Phenotype Correlations

    Directory of Open Access Journals (Sweden)

    Eva Y. G. De Vilder

    2017-01-01

    Full Text Available Gamma-carboxylation, performed by gamma-glutamyl carboxylase (GGCX, is an enzymatic process essential for activating vitamin K-dependent proteins (VKDP with important functions in various biological processes. Mutations in the encoding GGCX gene are associated with multiple phenotypes, amongst which vitamin K-dependent coagulation factor deficiency (VKCFD1 is best known. Other patients have skin, eye, heart or bone manifestations. As genotype–phenotype correlations were never described, literature was systematically reviewed in search of patients with at least one GGCX mutation with a phenotypic description, resulting in a case series of 47 patients. Though this number was too low for statistically valid correlations—a frequent problem in orphan diseases—we demonstrate the crucial role of the horizontally transferred transmembrane domain in developing cardiac and bone manifestations. Moreover, natural history suggests ageing as the principal determinant to develop skin and eye symptoms. VKCFD1 symptoms seemed more severe in patients with both mutations in the same protein domain, though this could not be linked to a more perturbed coagulation factor function. Finally, distinct GGCX functional domains might be dedicated to carboxylation of very specific VKDP. In conclusion, this systematic review suggests that there indeed may be genotype–phenotype correlations for GGCX-related phenotypes, which can guide patient counseling and management.

  12. Rv0004 is a new essential member of the mycobacterial DNA replication machinery.

    Science.gov (United States)

    Mann, Katherine M; Huang, Deborah L; Hooppaw, Anna J; Logsdon, Michelle M; Richardson, Kirill; Lee, Hark Joon; Kimmey, Jacqueline M; Aldridge, Bree B; Stallings, Christina L

    2017-11-01

    DNA replication is fundamental for life, yet a detailed understanding of bacterial DNA replication is limited outside the organisms Escherichia coli and Bacillus subtilis. Many bacteria, including mycobacteria, encode no identified homologs of helicase loaders or regulators of the initiator protein DnaA, despite these factors being essential for DNA replication in E. coli and B. subtilis. In this study we discover that a previously uncharacterized protein, Rv0004, from the human pathogen Mycobacterium tuberculosis is essential for bacterial viability and that depletion of Rv0004 leads to a block in cell cycle progression. Using a combination of genetic and biochemical approaches, we found that Rv0004 has a role in DNA replication, interacts with DNA and the replicative helicase DnaB, and affects DnaB-DnaA complex formation. We also identify a conserved domain in Rv0004 that is predicted to structurally resemble the N-terminal protein-protein interaction domain of DnaA. Mutation of a single conserved tryptophan within Rv0004's DnaA N-terminal-like domain leads to phenotypes similar to those observed upon Rv0004 depletion and can affect the association of Rv0004 with DnaB. In addition, using live cell imaging during depletion of Rv0004, we have uncovered a previously unappreciated role for DNA replication in coordinating mycobacterial cell division and cell size. Together, our data support that Rv0004 encodes a homolog of the recently identified DciA family of proteins found in most bacteria that lack the DnaC-DnaI helicase loaders in E. coli and B. subtilis. Therefore, the mechanisms of Rv0004 elucidated here likely apply to other DciA homologs and reveal insight into the diversity of bacterial strategies in even the most conserved biological processes.

  13. Introduction to DNA methods

    International Nuclear Information System (INIS)

    Delincee, H.

    1991-01-01

    The purpose of this session is to discuss the various possibilities for detecting modifications in DNA after irradiation and whether these changes can be utilized as an indicator for the irradiation treatment of foods. The requirement to be fulfilled is that the method be able to distinguish irradiated food without the presence of a control sample, thus the measured response after irradiation must be large enough to supersede background levels from other treatments. Much work has been performed on the effects of radiation on DNA, particularly due to its importance in radiation biology. The main lesions of DNA as a result of irradiation are base damage, damage of the sugar moiety, single strand and double strand breaks. Crosslinking between bases also occurs, e.g. production of thymine dimers, or between DNA and protein. A valuable review on how to utilize these DNA changes for detection purposes has already appeared. Tables 1, 2 and 3 list the proposed methods of detecting changes in irradiated DNA, some identified products as examples for a possible irradiation indicator, in the case of immunoassay the substance used as antigen, and some selected literature references. In this short review, it is not intended to provide a complete literature survey

  14. Variations in brain DNA

    Directory of Open Access Journals (Sweden)

    Jesus eAvila

    2014-11-01

    Full Text Available It is assumed that DNA sequences are conserved in the diverse cell types present in a multicellular organism like the human being. Thus, in order to compare the sequences in the genome of DNA from different individuals, nucleic acid is commonly isolated from a single tissue. In this regard, blood cells are widely used for this purpose because of their availability. Thus blood DNA has been used to study genetic familiar diseases that affect other tissues and organs, such as the liver, heart, and brain. While this approach is valid for the identification of familial diseases in which mutations are present in parental germinal cells and, therefore, in all the cells of a given organism, it is not suitable to identify sporadic diseases in which mutations might occur in specific somatic cells. This review addresses somatic DNA variations in different tissues or cells (mainly in the brain of single individuals and discusses whether the dogma of DNA invariance between cell types is indeed correct. We will also discuss how single nucleotide somatic variations arise, focusing on the presence of specific DNA mutations in the brain.

  15. Fanconi anaemia and the repair of Watson and Crick DNA crosslinks.

    Science.gov (United States)

    Kottemann, Molly C; Smogorzewska, Agata

    2013-01-17

    The function of Fanconi anaemia proteins is to maintain genomic stability. Their main role is in the repair of DNA interstrand crosslinks, which, by covalently binding the Watson and the Crick strands of DNA, impede replication and transcription. Inappropriate repair of interstrand crosslinks causes genomic instability, leading to cancer; conversely, the toxicity of crosslinking agents makes them a powerful chemotherapeutic. Fanconi anaemia proteins can promote stem-cell function, prevent tumorigenesis, stabilize replication forks and inhibit inaccurate repair. Recent advances have identified endogenous aldehydes as possible culprits of DNA damage that may induce the phenotypes seen in patients with Fanconi anaemia.

  16. Postreplication repair gap filling in an Escherichia coli strain deficient in dnaB gene product

    International Nuclear Information System (INIS)

    Johnson, R.C.

    1975-01-01

    Gaps in daughter-strand DNA synthesized after exposure of Escherichia coli E279 to ultraviolet light are filled during reincubation at 30 0 C for 20 min. Escherichia coli E279 is phenotypically DnaB - when incubated at 43 0 C. Cells incubated at 43 0 C were tested for their ability to complete postreplication repair gap filling. It is concluded that the dnaB gene product is essential for postreplication repair gap filling and that the inhibition seen is not initially the result of degradation

  17. MELAS syndrome, cardiomyopathy, rhabdomyolysis, and autism associated with the A3260G mitochondrial DNA mutation.

    Science.gov (United States)

    Connolly, Barbara S; Feigenbaum, Annette S J; Robinson, Brian H; Dipchand, Anne I; Simon, David K; Tarnopolsky, Mark A

    2010-11-12

    The A to G transition mutation at position 3260 of the mitochondrial genome is usually associated with cardiomyopathy and myopathy. One Japanese kindred reported the phenotype of mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS syndrome) in association with the A3260G mtDNA mutation. We describe the first Caucasian cases of MELAS syndrome associated with the A3260G mutation. Furthermore, this mutation was associated with exercise-induced rhabdomyolysis, hearing loss, seizures, cardiomyopathy, and autism in the large kindred. We conclude that the A3260G mtDNA mutation is associated with wide phenotypic heterogeneity with MELAS and other "classical" mitochondrial phenotypes being manifestations. Copyright © 2010 Elsevier Inc. All rights reserved.

  18. Genetic and Computational Approaches for Studying Plant Development and Abiotic Stress Responses Using Image-Based Phenotyping

    Science.gov (United States)

    Campbell, M. T.; Walia, H.; Grondin, A.; Knecht, A.

    2017-12-01

    The development of abiotic stress tolerant crops (i.e. drought, salinity, or heat stress) requires the discovery of DNA sequence variants associated with stress tolerance-related traits. However, many traits underlying adaptation to abiotic stress involve a suite of physiological pathways that may be induced at different times throughout the duration of stress. Conventional single-point phenotyping approaches fail to fully capture these temporal responses, and thus downstream genetic analysis may only identify a subset of the genetic variants that are important for adaptation to sub-optimal environments. Although genomic resources for crops have advanced tremendously, the collection of phenotypic data for morphological and physiological traits is laborious and remains a significant bottleneck in bridging the phenotype-genotype gap. In recent years, the availability of automated, image-based phenotyping platforms has provided researchers with an opportunity to collect morphological and physiological traits non-destructively in a highly controlled environment. Moreover, these platforms allow abiotic stress responses to be recorded throughout the duration of the experiment, and have facilitated the use of function-valued traits for genetic analyses in major crops. We will present our approaches for addressing abiotic stress tolerance in cereals. This talk will focus on novel open-source software to process and extract biological meaningful data from images generated from these phenomics platforms. In addition, we will discuss the statistical approaches to model longitudinal phenotypes and dissect the genetic basis of dynamic responses to these abiotic stresses throughout development.

  19. No Association between Variation in Longevity Candidate Genes and Aging-related Phenotypes in Oldest-old Danes.

    Science.gov (United States)

    Soerensen, Mette; Nygaard, Marianne; Debrabant, Birgit; Mengel-From, Jonas; Dato, Serena; Thinggaard, Mikael; Christensen, Kaare; Christiansen, Lene

    2016-06-01

    In this study we explored the association between aging-related phenotypes previously reported to predict survival in old age and variation in 77 genes from the DNA repair pathway, 32 genes from the growth hormone 1/ insulin-like growth factor 1/insulin (GH/IGF-1/INS) signalling pathway and 16 additional genes repeatedly considered as candidates for human longevity: APOE, APOA4, APOC3, ACE, CETP, HFE, IL6, IL6R, MTHFR, TGFB1, SIRTs 1, 3, 6; and HSPAs 1A, 1L, 14. Altogether, 1,049 single nucleotide polymorphisms (SNPs) were genotyped in 1,088 oldest-old (age 92-93 years) Danes and analysed with phenotype data on physical functioning (hand grip strength), cognitive functioning (mini mental state examination and a cognitive composite score), activity of daily living and self-rated health. Five SNPs showed association to one of the phenotypes; however, none of these SNPs were associated with a change in the relevant phenotype over time (7 years of follow-up) and none of the SNPs could be confirmed in a replication sample of 1,281 oldest-old Danes (age 94-100). Hence, our study does not support association between common variation in the investigated longevity candidate genes and aging-related phenotypes consistently shown to predict survival. It is possible that larger sample sizes are needed to robustly reveal associations with small effect sizes. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. Plant phenomics and the need for physiological phenotyping across scales to narrow the genotype-to-phenotype knowledge gap

    Czech Academy of Sciences Publication Activity Database

    Grosskinsky, D. K.; Svensgaard, J.; Christensen, S.; Roitsch, Thomas

    2015-01-01

    Roč. 66, č. 18 (2015), s. 5429-5440 ISSN 0022-0957 Institutional support: RVO:67179843 Keywords : External phenotype * genome–environment–management interaction * genome–phenome map * internal phenotype * phenomics * physiological traits * physiology * plant phenotyping * predictors Subject RIV: EH - Ecology, Behaviour Impact factor: 5.677, year: 2015

  1. Phenotypic plasticity and morphological integration in a marine modular invertebrate

    Directory of Open Access Journals (Sweden)

    Manrique Nelson

    2007-07-01

    Full Text Available Abstract Background Colonial invertebrates such as corals exhibit nested levels of modularity, imposing a challenge to the depiction of their morphological evolution. Comparisons among diverse Caribbean gorgonian corals suggest decoupling of evolution at the polyp vs. branch/internode levels. Thus, evolutionary change in polyp form or size (the colonial module sensu stricto does not imply a change in colony form (constructed of modular branches and other emergent features. This study examined the patterns of morphological integration at the intraspecific level. Pseudopterogorgia bipinnata (Verrill (Octocorallia: Gorgoniidae is a Caribbean shallow water gorgonian that can colonize most reef habitats (shallow/exposed vs. deep/protected; 1–45 m and shows great morphological variation. Results To characterize the genotype/environment relationship and phenotypic plasticity in P. bipinnata, two microsatellite loci, mitochondrial (MSH1 and nuclear (ITS DNA sequences, and (ITS2 DGGE banding patterns were initially compared among the populations present in the coral reefs of Belize (Carrie Bow Cay, Panama (Bocas del Toro, Colombia (Cartagena and the Bahamas (San Salvador. Despite the large and discrete differentiation of morphotypes, there was no concordant genetic variation (DGGE banding patterns in the ITS2 genotypes from Belize, Panama and Colombia. ITS1–5.8S-ITS2 phylogenetic analysis afforded evidence for considering the species P. kallos (Bielschowsky as the shallow-most morphotype of P. bipinnata from exposed environments. The population from Carrie Bow Cay, Belize (1–45 m was examined to determine the phenotypic integration of modular features such as branch thickness, polyp aperture, inter-polyp distance, internode length and branch length. Third-order partial correlation coefficients suggested significant integration between polypar and colonial traits. Some features did not change at all despite 10-fold differences in other integrated

  2. Gaucher disease: molecular heterogeneity and phenotype-genotype correlations.

    Science.gov (United States)

    Theophilus, B; Latham, T; Grabowski, G A; Smith, F I

    1989-08-01

    Gaucher disease (GD) is the most prevalent lysosomal storage disease. This autosomal recessive trait results from the defective activity of acid beta-glucosidase (beta-Glc). Four different exonic point mutations have been identified as causal alleles for GD. To facilitate screening for these alleles, assays were developed using allele-specific oligonucleotide hybridization to amplified genomic DNA sequences. Specifically, intron bases flanking exons 5, 9, and 10 were determined, and conditions for PCR amplification of these exons were obtained. Two different procedures were developed to distinguish signals obtained from the structural beta-Glc gene exons and those from the pseudogene. These procedures were used to determine the distribution of all known GD alleles in a population of 44 affected patients of varying phenotypes and ethnicity. The high frequency of one of the exon 9 mutations in Ashkenazi Jewish GD type 1 patients was confirmed, and, in addition, this mutation was present in ethnically diverse non-Jewish type 1 GD patients. Homozygotes (N = 5) for this allele were midly affected older individuals, and this mutant allele was not found in any patient with neuronopathic disease. The exon 10 mutation was confirmed as the predominant allele in types 2 and 3 GD. However, several type 1 GD patients, including one of Ashkenazi-Jewish heritage, also were heterozygous for this allele. The presence of this allele in type 1 patients did not correlate with the severity of clinical symptoms. The second exon 9 mutation and the exon 5 mutation were rare, since they occurred only heterozygously either in one type 2 GD patient or in two related Ashkenazi-Jewish GD patients, respectively. Although most GD patients (38 of 44) had at least one of the known mutant alleles, 57% were heterozygotes for only one of these mutations. Fourteen percent of patients were negative for all mutations. A total of 73% of GD patients had at least one unknown allele. The varying clinical

  3. Invasion strategies in clonal aquatic plants: are phenotypic differences caused by phenotypic plasticity or local adaptation?

    Science.gov (United States)

    Riis, Tenna; Lambertini, Carla; Olesen, Birgit; Clayton, John S.; Brix, Hans; Sorrell, Brian K.

    2010-01-01

    Background and Aims The successful spread of invasive plants in new environments is often linked to multiple introductions and a diverse gene pool that facilitates local adaptation to variable environmental conditions. For clonal plants, however, phenotypic plasticity may be equally important. Here the primary adaptive strategy in three non-native, clonally reproducing macrophytes (Egeria densa, Elodea canadensis and Lagarosiphon major) in New Zealand freshwaters were examined and an attempt was made to link observed differences in plant morphology to local variation in habitat conditions. Methods Field populations with a large phenotypic variety were sampled in a range of lakes and streams with different chemical and physical properties. The phenotypic plasticity of the species before and after cultivation was studied in a common garden growth experiment, and the genetic diversity of these same populations was also quantified. Key Results For all three species, greater variation in plant characteristics was found before they were grown in standardized conditions. Moreover, field populations displayed remarkably little genetic variation and there was little interaction between habitat conditions and plant morphological characteristics. Conclusions The results indicate that at the current stage of spread into New Zealand, the primary adaptive strategy of these three invasive macrophytes is phenotypic plasticity. However, while limited, the possibility that genetic diversity between populations may facilitate ecotypic differentiation in the future cannot be excluded. These results thus indicate that invasive clonal aquatic plants adapt to new introduced areas by phenotypic plasticity. Inorganic carbon, nitrogen and phosphorous were important in controlling plant size of E. canadensis and L. major, but no other relationships between plant characteristics and habitat conditions were apparent. This implies that within-species differences in plant size can be explained

  4. Genotype-phenotype associations in obesity dependent on definition of the obesity phenotype.

    Science.gov (United States)

    Kring, Sofia Inez Iqbal; Larsen, Lesli Hingstrup; Holst, Claus; Toubro, Søren; Hansen, Torben; Astrup, Arne; Pedersen, Oluf; Sørensen, Thorkild I A

    2008-01-01

    In previous studies of associations of variants in the genes UCP2, UCP3, PPARG2, CART, GRL, MC4R, MKKS, SHP, GHRL, and MCHR1 with obesity, we have used a case-control approach with cases defined by a threshold for BMI. In the present study, we assess the association of seven abdominal, peripheral, and overall obesity phenotypes, which were analyzed quantitatively, and thirteen candidate gene polymorphisms in these ten genes in the same cohort. Obese Caucasian men (n = 234, BMI >or= 31.0 kg/m(2)) and a randomly sampled non-obese group (n = 323), originally identified at the draft board examinations, were re-examined at median ages of 47.0 or 49.0 years by anthropometry and DEXA scanning. Obesity phenotypes included BMI, fat body mass index, waist circumference, waist for given BMI, intra-abdominal adipose tissue, hip circumference and lower body fat mass (%). Using logistic regression models, we estimated the odds for defined genotypes (dominant or recessive genetic transmission) in relation to z-scores of the phenotypes. The minor (rare) allele for SHP 512G>C (rs6659176) was associated with increased hip circumference. The minor allele for UCP2 Ins45bp was associated with increased BMI, increased abdominal obesity, and increased hip circumference. The minor allele for UCP2 -866G>A (rs6593669) was associated with borderline increased fat body mass index. The minor allele for MCHR1 100213G>A (rs133072) was associated with reduced abdominal obesity. None of the other genotype-phenotype combinations showed appreciable associations. If replicated in independent studies with focus on the specific phenotypes, our explorative studies suggest significant associations between some candidate gene polymorphisms and distinct obesity phenotypes, predicting beneficial and detrimental effects, depending on compartments for body fat accumulation. Copyright 2008 S. Karger AG, Basel.

  5. The genotype-phenotype map of an evolving digital organism.

    Directory of Open Access Journals (Sweden)

    Miguel A Fortuna

    2017-02-01

    Full Text Available To understand how evolving systems bring forth novel and useful phenotypes, it is essential to understand the relationship between genotypic and phenotypic change. Artificial evolving systems can help us understand whether the genotype-phenotype maps of natural evolving systems are highly unusual, and it may help create evolvable artificial systems. Here we characterize the genotype-phenotype map of digital organisms in Avida, a platform for digital evolution. We consider digital organisms from a vast space of 10141 genotypes (instruction sequences, which can form 512 different phenotypes. These phenotypes are distinguished by different Boolean logic functions they can compute, as well as by the complexity of these functions. We observe several properties with parallels in natural systems, such as connected genotype networks and asymmetric phenotypic transitions. The likely common cause is robustness to genotypic change. We describe an intriguing tension between phenotypic complexity and evolvability that may have implications for biological evolution. On the one hand, genotypic change is more likely to yield novel phenotypes in more complex organisms. On the other hand, the total number of novel phenotypes reachable through genotypic change is highest for organisms with simple phenotypes. Artificial evolving systems can help us study aspects of biological evolvability that are not accessible in vastly more complex natural systems. They can also help identify properties, such as robustness, that are required for both human-designed artificial systems and synthetic biological systems to be evolvable.

  6. The genotype-phenotype map of an evolving digital organism.

    Science.gov (United States)

    Fortuna, Miguel A; Zaman, Luis; Ofria, Charles; Wagner, Andreas

    2017-02-01

    To understand how evolving systems bring forth novel and useful phenotypes, it is essential to understand the relationship between genotypic and phenotypic change. Artificial evolving systems can help us understand whether the genotype-phenotype maps of natural evolving systems are highly unusual, and it may help create evolvable artificial systems. Here we characterize the genotype-phenotype map of digital organisms in Avida, a platform for digital evolution. We consider digital organisms from a vast space of 10141 genotypes (instruction sequences), which can form 512 different phenotypes. These phenotypes are distinguished by different Boolean logic functions they can compute, as well as by the complexity of these functions. We observe several properties with parallels in natural systems, such as connected genotype networks and asymmetric phenotypic transitions. The likely common cause is robustness to genotypic change. We describe an intriguing tension between phenotypic complexity and evolvability that may have implications for biological evolution. On the one hand, genotypic change is more likely to yield novel phenotypes in more complex organisms. On the other hand, the total number of novel phenotypes reachable through genotypic change is highest for organisms with simple phenotypes. Artificial evolving systems can help us study aspects of biological evolvability that are not accessible in vastly more complex natural systems. They can also help identify properties, such as robustness, that are required for both human-designed artificial systems and synthetic biological systems to be evolvable.

  7. Blood extracellular DNA after irradiation

    International Nuclear Information System (INIS)

    Vladimirov, V.G.; Tishchenko, L.I.; Surkova, E.A.; Vasil'eva, I.N.

    1993-01-01

    It has been shown that blood extracellular DNA of irradiated rats largely consists of the low-molecular DNA and its oligomers. Molecular masses of oligomers are multiple to molecular mass of monomer fragment with nucleosome size. The low-molecular DNA has linear form. The average content of GC-pairs in low-molecular DNA is higher than in total rat's DNA (48.5% against 41.5%). The low-molecular DNA is a part of complex containing RNA, acidic proteins and lipids. It is assumed that the formation of low-molecular DNA is a result of Ca/Mg - dependent nuclear endonuclease action

  8. The phenotypic plasticity of developmental modules

    Directory of Open Access Journals (Sweden)

    Aabha I. Sharma

    2016-08-01

    Full Text Available Abstract Background Organisms develop and evolve in a modular fashion, but how individual modules interact with the environment remains poorly understood. Phenotypically plastic traits are often under selection, and studies are needed to address how traits respond to the environment in a modular fashion. In this study, tissue-specific plasticity of melanic spots was examined in the large milkweed bug, Oncopeltus fasciatus. Results Although the size of the abdominal melanic bands varied according to rearing temperatures, wing melanic bands were more robust. To explore the regulation of abdominal pigmentation plasticity, candidate genes involved in abdominal melanic spot patterning and biosynthesis of melanin were analyzed. While the knockdown of dopa decarboxylase (Ddc led to lighter pigmentation in both the wings and the abdomen, the shape of the melanic elements remained unaffected. Although the knockdown of Abdominal-B (Abd-B partially phenocopied the low-temperature phenotype, the abdominal bands were still sensitive to temperature shifts. These observations suggest that regulators downstream of Abd-B but upstream of DDC are responsible for the temperature response of the abdomen. Ablation of wings led to the regeneration of a smaller wing with reduced melanic bands that were shifted proximally. In addition, the knockdown of the Wnt signaling nuclear effector genes, armadillo 1 and armadillo 2, altered both the melanic bands and the wing shape. Thus, the pleiotropic effects of Wnt signaling may constrain the amount of plasticity in wing melanic bands. Conclusions We propose that when traits are regulated by distinct pre-patterning mechanisms, they can respond to the environment in a modular fashion, whereas when the environment impacts developmental regulators that are shared between different modules, phenotypic plasticity can manifest as a developmentally integrated system.

  9. Cardiac Phenotype of Prehypertrophic Fabry Disease.

    Science.gov (United States)

    Nordin, Sabrina; Kozor, Rebecca; Baig, Shanat; Abdel-Gadir, Amna; Medina-Menacho, Katia; Rosmini, Stefania; Captur, Gabriella; Tchan, Michel; Geberhiwot, Tarekegn; Murphy, Elaine; Lachmann, Robin; Ramaswami, Uma; Edwards, Nicola C; Hughes, Derralynn; Steeds, Richard P; Moon, James C

    2018-06-01

    Fabry disease (FD) is a rare and treatable X-linked lysosomal storage disorder. Cardiac involvement determines outcomes; therefore, detecting early changes is important. Native T1 by cardiovascular magnetic resonance is low, reflecting sphingolipid storage. Early phenotype development is familiar in hypertrophic cardiomyopathy but unexplored in FD. We explored the prehypertrophic cardiac phenotype of FD and the role of storage. A prospective, international multicenter observational study of 100 left ventricular hypertrophy-negative FD patients (mean age: 39±15 years; 19% male) and 35 age- and sex-matched healthy volunteers (mean age: 40±14 years; 25% male) who underwent cardiovascular magnetic resonance, including native T1 and late gadolinium enhancement, and 12-lead ECG. In FD, 41% had a low native T1 using a single septal region of interest, but this increased to 59% using a second slice because early native T1 lowering was patchy. ECG abnormalities were present in 41% and twice as common with low native T1 (53% versus 24%; P =0.005). When native T1 was low, left ventricular maximum wall thickness, indexed mass, and ejection fraction were higher (maximum wall thickness 9±1.5 versus 8±1.4 mm, P gadolinium enhancement was more likely when native T1 was low (27% versus 6%; P =0.01). FD had higher maximal apical fractal dimensions compared with healthy volunteers (1.27±0.06 versus 1.24±0.04; P <0.005) and longer anterior mitral valve leaflets (23±2 mm versus 21±3 mm; P <0.005). There is a detectable prehypertrophic phenotype in FD consisting of storage (low native T1), structural, functional, and ECG changes. © 2018 The Authors.

  10. Sample size calculation in metabolic phenotyping studies.

    Science.gov (United States)

    Billoir, Elise; Navratil, Vincent; Blaise, Benjamin J

    2015-09-01

    The number of samples needed to identify significant effects is a key question in biomedical studies, with consequences on experimental designs, costs and potential discoveries. In metabolic phenotyping studies, sample size determination remains a complex step. This is due particularly to the multiple hypothesis-testing framework and the top-down hypothesis-free approach, with no a priori known metabolic target. Until now, there was no standard procedure available to address this purpose. In this review, we discuss sample size estimation procedures for metabolic phenotyping studies. We release an automated implementation of the Data-driven Sample size Determination (DSD) algorithm for MATLAB and GNU Octave. Original research concerning DSD was published elsewhere. DSD allows the determination of an optimized sample size in metabolic phenotyping studies. The procedure uses analytical data only from a small pilot cohort to generate an expanded data set. The statistical recoupling of variables procedure is used to identify metabolic variables, and their intensity distributions are estimated by Kernel smoothing or log-normal density fitting. Statistically significant metabolic variations are evaluated using the Benjamini-Yekutieli correction and processed for data sets of various sizes. Optimal sample size determination is achieved in a context of biomarker discovery (at least one statistically significant variation) or metabolic exploration (a maximum of statistically significant variations). DSD toolbox is encoded in MATLAB R2008A (Mathworks, Natick, MA) for Kernel and log-normal estimates, and in GNU Octave for log-normal estimates (Kernel density estimates are not robust enough in GNU octave). It is available at http://www.prabi.fr/redmine/projects/dsd/repository, with a tutorial at http://www.prabi.fr/redmine/projects/dsd/wiki. © The Author 2015. Published by Oxford University Press. For Permissions, please email: journals.permissions@oup.com.

  11. Two Clinical Phenotypes in Polycythemia Vera

    Science.gov (United States)

    Spivak, Jerry L.; Considine, Michael; Williams, Donna M.; Talbot, Conover C.; Rogers, Ophelia; Moliterno, Alison R.; Jie, Chunfa; Ochs, Michael F.

    2014-01-01

    BACKGROUND Polycythemia vera is the ultimate phenotypic consequence of the V617F mutation in Janus kinase 2 (encoded by JAK2), but the extent to which this mutation influences the behavior of the involved CD34+ hematopoietic stem cells is unknown. METHODS We analyzed gene expression in CD34+ peripheral-blood cells from 19 patients with polycythemia vera, using oligonucleotide microarray technology after correcting for potential confounding by sex, since the phenotypic features of the disease differ between men and women. RESULTS Men with polycythemia vera had twice as many up-regulated or down-regulated genes as women with polycythemia vera, in a comparison of gene expression in the patients and in healthy persons of the same sex, but there were 102 genes with differential regulation that was concordant in men and women. When these genes were used for class discovery by means of unsupervised hierarchical clustering, the 19 patients could be divided into two groups that did not differ significantly with respect to age, neutrophil JAK2 V617F allele burden, white-cell count, platelet count, or clonal dominance. However, they did differ significantly with respect to disease duration; hemoglobin level; frequency of thromboembolic events, palpable splenomegaly, and splenectomy; chemotherapy exposure; leukemic transformation; and survival. The unsupervised clustering was confirmed by a supervised approach with the use of a top-scoring-pair classifier that segregated the 19 patients into the same two phenotypic groups with 100% accuracy. CONCLUSIONS Removing sex as a potential confounder, we identified an accurate molecular method for classifying patients with polycythemia vera according to disease behavior, independently of their JAK2 V617F allele burden, and identified previously unrecognized molecular pathways in polycythemia vera outside the canonical JAK2 pathway that may be amenable to targeted therapy. PMID:25162887

  12. Wolfram syndrome: new mutations, different phenotype.

    Directory of Open Access Journals (Sweden)

    Concetta Aloi

    Full Text Available BACKGROUND: Wolfram Syndrome (WS is an autosomal recessive neurodegenerative disorder characterized by Diabetes Insipidus, Diabetes Mellitus, Optic Atrophy, and Deafness identified by the acronym "DIDMOAD". The WS gene, WFS1, encodes a transmembrane protein called Wolframin, which recent evidence suggests may serve as a novel endoplasmic reticulum calcium channel in pancreatic β-cells and neurons. WS is a rare disease, with an estimated prevalence of 1/550.000 children, with a carrier frequency of 1/354. The aim of our study was to determine the genotype of WS patients in order to establish a genotype/phenotype correlation. METHODOLOGY/PRINCIPAL FINDINGS: We clinically evaluated 9 young patients from 9 unrelated families (6 males, 3 females. Basic criteria for WS clinical diagnosis were coexistence of insulin-treated diabetes mellitus and optic atrophy occurring before 15 years of age. Genetic analysis for WFS1 was performed by direct sequencing. Molecular sequencing revealed 5 heterozygous compound and 3 homozygous mutations. All of them were located in exon 8, except one in exon 4. In one proband only an heterozygous mutation (A684V was found. Two new variants c.2663 C>A and c.1381 A>C were detected. CONCLUSIONS/SIGNIFICANCE: Our study increases the spectrum of WFS1 mutations with two novel variants. The male patient carrying the compound mutation [c.1060_1062delTTC]+[c.2663 C>A] showed the most severe phenotype: diabetes mellitus, optic atrophy (visual acuity 5/10, deafness with deep auditory bilaterally 8000 Hz, diabetes insipidus associated to reduced volume of posterior pituitary and pons. He died in bed at the age of 13 years. The other patient carrying the compound mutation [c.409_424dup16]+[c.1381 A>C] showed a less severe phenotype (DM, OA.

  13. Methods for Analyzing Multivariate Phenotypes in Genetic Association Studies

    Directory of Open Access Journals (Sweden)

    Qiong Yang

    2012-01-01

    Full Text Available Multivariate phenotypes are frequently encountered in genetic association studies. The purpose of analyzing multivariate phenotypes usually includes discovery of novel genetic variants of pleiotropy effects, that is, affecting multiple phenotypes, and the ultimate goal of uncovering the underlying genetic mechanism. In recent years, there have been new method development and application of existing statistical methods to such phenotypes. In this paper, we provide a review of the available methods for analyzing association between a single marker and a multivariate phenotype consisting of the same type of components (e.g., all continuous or all categorical or different types of components (e.g., some are continuous and others are categorical. We also reviewed causal inference methods designed to test whether the detected association with the multivariate phenotype is truly pleiotropy or the genetic marker exerts its effects on some phenotypes through affecting the others.

  14. Phenotype specific analyses reveal distinct regulatory mechanism for chronically activated p53.

    Directory of Open Access Journals (Sweden)

    Kristina Kirschner

    2015-03-01

    Full Text Available The downstream functions of the DNA binding tumor suppressor p53 vary depending on the cellular context, and persistent p53 activation has recently been implicated in tumor suppression and senescence. However, genome-wide information about p53-target gene regulation has been derived mostly from acute genotoxic conditions. Using ChIP-seq and expression data, we have found distinct p53 binding profiles between acutely activated (through DNA damage and chronically activated (in senescent or pro-apoptotic conditions p53. Compared to the classical 'acute' p53 binding profile, 'chronic' p53 peaks were closely associated with CpG-islands. Furthermore, the chronic CpG-island binding of p53 conferred distinct expression patterns between senescent and pro-apoptotic conditions. Using the p53 targets seen in the chronic conditions together with external high-throughput datasets, we have built p53 networks that revealed extensive self-regulatory 'p53 hubs' where p53 and many p53 targets can physically interact with each other. Integrating these results with public clinical datasets identified the cancer-associated lipogenic enzyme, SCD, which we found to be directly repressed by p53 through the CpG-island promoter, providing a mechanistic link between p53 and the 'lipogenic phenotype', a hallmark of cancer. Our data reveal distinct phenotype associations of chronic p53 targets that underlie specific gene regulatory mechanisms.

  15. Phenotypic and genetic diversity of chlorine-resistant Methylobacterium strains isolated from various environments.

    Science.gov (United States)

    Hiraishi, A; Furuhata, K; Matsumoto, A; Koike, K A; Fukuyama, M; Tabuchi, K

    1995-01-01

    Strains of pink-pigmented facultative methylotrophs which were isolated previously from various environments and assigned tentatively to the genus Methylobacterium were characterized in comparison with authentic strains of previously known species of this genus. Most of the isolates derived from chlorinated water supplies exhibited resistance to chlorine, whereas 29 to 40% of the isolates from air, natural aquatic environments, and clinical materials were chlorine resistant. None of the tested authentic strains of Methylobacterium species obtained from culture collections exhibited chlorine resistance. Numerical analysis of phenotypic profiles showed that the test organisms tested were separated from each other except M. organophilum and M. rhodesianum. The chlorine-resistant isolates were randomly distributed among all clusters. The 16S ribosomal DNA (rDNA) sequence-based phylogenetic analyses showed that representatives of the isolates together with known Methylobacterium species formed a line of descent distinct from that of members of related genera in the alpha-2 subclass of the Proteobacteria and were divided into three subclusters within the Methylobacterium group. These results demonstrate that there is phenotypic and genetic diversity among chlorine-resistant Methylobacterium strains within the genus. PMID:7793931

  16. Phenotypic and genetic diversity of chlorine-resistant Methylobacterium strains isolated from various environments.

    Science.gov (United States)

    Hiraishi, A; Furuhata, K; Matsumoto, A; Koike, K A; Fukuyama, M; Tabuchi, K

    1995-06-01

    Strains of pink-pigmented facultative methylotrophs which were isolated previously from various environments and assigned tentatively to the genus Methylobacterium were characterized in comparison with authentic strains of previously known species of this genus. Most of the isolates derived from chlorinated water supplies exhibited resistance to chlorine, whereas 29 to 40% of the isolates from air, natural aquatic environments, and clinical materials were chlorine resistant. None of the tested authentic strains of Methylobacterium species obtained from culture collections exhibited chlorine resistance. Numerical analysis of phenotypic profiles showed that the test organisms tested were separated from each other except M. organophilum and M. rhodesianum. The chlorine-resistant isolates were randomly distributed among all clusters. The 16S ribosomal DNA (rDNA) sequence-based phylogenetic analyses showed that representatives of the isolates together with known Methylobacterium species formed a line of descent distinct from that of members of related genera in the alpha-2 subclass of the Proteobacteria and were divided into three subclusters within the Methylobacterium group. These results demonstrate that there is phenotypic and genetic diversity among chlorine-resistant Methylobacterium strains within the genus.

  17. DNA Knots: Theory and Experiments

    Science.gov (United States)

    Sumners, D. W.

    Cellular DNA is a long, thread-like molecule with remarkably complex topology. Enzymes that manipulate the geometry and topology of cellular DNA perform many vital cellular processes (including segregation of daughter chromosomes, gene regulation, DNA repair, and generation of antibody diversity). Some enzymes pass DNA through itself via enzyme-bridged transient breaks in the DNA; other enzymes break the DNA apart and reconnect it to different ends. In the topological approach to enzymology, circular DNA is incubated with an enzyme, producing an enzyme signature in the form of DNA knots and links. By observing the changes in DNA geometry (supercoiling) and topology (knotting and linking) due to enzyme action, the enzyme binding and mechanism can often be characterized. This paper will discuss some personal research history, and the tangle model for the analysis of site-specific recombination experiments on circular DNA.

  18. Endangered species: mitochondrial DNA loss as a mechanism of human disease.

    Science.gov (United States)

    Herrera, Alan; Garcia, Iraselia; Gaytan, Norma; Jones, Edith; Maldonado, Alicia; Gilkerson, Robert

    2015-06-01

    Human mitochondrial DNA (mtDNA) is a small maternally inherited DNA, typically present in hundreds of copies in a single human cell. Thus, despite its small size, the mitochondrial genome plays a crucial role in the metabolic homeostasis of the cell. Our understanding of mtDNA genotype-phenotype relationships is derived largely from studies of the classical mitochondrial neuromuscular diseases, in which mutations of mtDNA lead to compromised mitochondrial bioenergetic function, with devastating pathological consequences. Emerging research suggests that loss, rather than mutation, of mtDNA plays a major role across a range of prevalent human diseases, including diabetes mellitus, cardiovascular disease, and aging. Here, we examine the 'rules' of mitochondrial genetics and function, the clinical settings in which loss of mtDNA is an emerging pathogenic mechanism, and explore mtDNA damage and its consequences for the organellar network and cell at large. As extranuclear genetic material arrayed throughout the cell to support metabolism, mtDNA is increasingly implicated in a host of disease conditions, opening a range of exciting questions regarding mtDNA and its role in cellular homeostasis.

  19. Genome-wide DNA methylation sequencing reveals miR-663a is a novel epimutation candidate in CIMP-high endometrial cancer

    OpenAIRE

    Yanokura, Megumi; Banno, Kouji; Adachi, Masataka; Aoki, Daisuke; Abe, Kuniya

    2017-01-01

    Aberrant DNA methylation is widely observed in many cancers. Concurrent DNA methylation of multiple genes occurs in endometrial cancer and is referred to as the CpG island methylator phenotype (CIMP). However, the features and causes of CIMP-positive endometrial cancer are not well understood. To investigate DNA methylation features characteristic to CIMP-positive endometrial cancer, we first classified samples from 25 patients with endometrial cancer based on the methylation status of three ...

  20. Overexpression of the DNA mismatch repair factor, PMS2, confers hypermutability and DNA damage tolerance.

    Science.gov (United States)

    Gibson, Shannon L; Narayanan, Latha; Hegan, Denise Campisi; Buermeyer, Andrew B; Liskay, R Michael; Glazer, Peter M

    2006-12-08

    Inherited defects in genes associated with DNA mismatch repair (MMR) have been linked to familial colorectal cancer. Cells deficient in MMR are genetically unstable and demonstrate a tolerance phenotype in response to certain classes of DNA damage. Some sporadic human cancers also show abnormalities in MMR gene function, typically due to diminished expression of one of the MutL homologs, MLH1. Here, we report that overexpression of the MutL homolog, human PMS2, can also cause a disruption of the MMR pathway in mammalian cells, resulting in hypermutability and DNA damage tolerance. A mouse fibroblast cell line carrying a recoverable lambda phage shuttle vector for mutation detection was transfected with either a vector designed to express hPMS2 or with an empty vector control. Cells overexpressing hPMS2 were found to have elevated spontaneous mutation frequencies at the cII reporter gene locus. They also showed an increase in the level of mutations induced by the alkylating agent, methynitrosourea (MNU). Clonogenic survival assays demonstrated increased survival of the PMS2-overexpressing cells following exposure to MNU, consistent with the induction of a damage tolerance phenotype. Similar results were seen in cells expressing a mutant PMS2 gene, containing a premature stop codon at position 134 and representing a variant found in an individual with familial colon cancer. These results show that dysregulation of PMS2 gene expression can disrupt MMR function in mammalian cells and establish an additional carcinogenic mechanism by which cells can develop genetic instability and acquire resistance to cytotoxic cancer therapies.

  1. Mitochondrial DNA sequence data reveals association of haplogroup U with psychosis in bipolar disorder.

    Science.gov (United States)

    Frye, Mark A; Ryu, Euijung; Nassan, Malik; Jenkins, Gregory D; Andreazza, Ana C; Evans, Jared M; McElroy, Susan L; Oglesbee, Devin; Highsmith, W Edward; Biernacka, Joanna M

    2017-01-01

    Converging genetic, postmortem gene-expression, cellular, and neuroimaging data implicate mitochondrial dysfunction in bipolar disorder. This study was conducted to investigate whether mitochondrial DNA (mtDNA) haplogroups and single nucleotide variants (SNVs) are associated with sub-phenotypes of bipolar disorder. MtDNA from 224 patients with Bipolar I disorder (BPI) was sequenced, and association of sequence variations with 3 sub-phenotypes (psychosis, rapid cycling, and adolescent illness onset) was evaluated. Gene-level tests were performed to evaluate overall burden of minor alleles for each phenotype. The haplogroup U was associated with a higher risk of psychosis. Secondary analyses of SNVs provided nominal evidence for association of psychosis with variants in the tRNA, ND4 and ND5 genes. The association of psychosis with ND4 (gene that encodes NADH dehydrogenase 4) was further supported by gene-level analysis. Preliminary analysis of mtDNA sequence data suggests a higher risk of psychosis with the U haplogroup and variation in the ND4 gene implicated in electron transport chain energy regulation. Further investigation of the functional consequences of this mtDNA variation is encouraged. Copyright © 2016. Published by Elsevier Ltd.

  2. Epigenetic marks: regulators of livestock phenotypes and conceivable sources of missing variation in livestock improvement programs

    Directory of Open Access Journals (Sweden)

    Eveline M Ibeagha-Awemu

    2015-09-01

    Full Text Available Improvement in animal productivity has been achieved over the years through careful breeding and selection programs. Today, variations in the genome are gaining increasing importance in livestock improvement strategies. Genomic information alone however explains only a part of the phenotypic variance in traits. It is likely that a portion of the unaccounted variance is embedded in the epigenome. The epigenome encompasses epigenetic marks such as DNA methylation, histone tail modifications, chromatin remodeling and other molecules that can transmit epigenetic information such as non-coding RNA species. Epigenetic factors respond to external or internal environmental cues such as nutrition, pathogens and climate, and have the ability to change gene expression leading to emergence of specific phenotypes. Accumulating evidence shows that epigenetic marks influence gene expression and phenotypic outcome in livestock species. This review examines available evidence of the influence of epigenetic marks on livestock (cattle, sheep, goat and pig traits and discusses the potential for consideration of epigenetic markers in livestock improvement programs. However, epigenetic research activities on farm animal species are currently limited partly due to lack of recognition, funding and a global network of researchers. Therefore, considerable less attention has been given to epigenetic research in livestock species in comparison to extensive work in humans and model organisms. Elucidating therefore the epigenetic determinants of animal diseases and complex traits may represent one of the principal challenges to use epigenetic markers for further improvement of animal productivity.

  3. Phenotypic and molecular identification of Fonsecaea pedrosoi strains isolated from chromoblastomycosis patients in Mexico and Venezuela.

    Science.gov (United States)

    Carolina Rojas, O; León-Cachón, Rafael B R; Pérez-Maya, Antonio Alí; Aguirre-Garza, Marcelino; Moreno-Treviño, María G; González, Gloria M

    2015-05-01

    Chromoblastomycosis is a chronic granulomatous disease caused frequently by fungi of the Fonsecaea genus. The objective of this study was the phenotypic and molecular identification of F. pedrosoi strains isolated from chromoblastomycosis patients in Mexico and Venezuela. Ten strains were included in this study. For phenotypic identification, we used macroscopic and microscopic morphologies, carbohydrate assimilation test, urea hydrolysis, cixcloheximide tolerance, proteolitic activity and the thermotolerance test. The antifungal activity of five drugs was evaluated against the isolates. Molecular identification was performed by sequencing the internal transcribed spacer (ITS) ribosomal DNA regions of the isolated strains. The physiological analysis and morphological features were variable and the precise identification was not possible. All isolates were susceptible to itraconazole, terbinafine, voriconazole and posaconazole. Amphotericin B was the least effective drug. The alignment of the 559-nucleotide ITS sequences from our strains compared with sequences of GenBank revealed high homology with F. pedrosoi (EU285266.1). In this study, all patients were from rural areas, six from Mexico and four from Venezuela. Ten isolates were identified by phenotypic and molecular analysis, using ITS sequence and demonstrated that nine isolates from Mexico and Venezuela were 100% homologous and one isolate showed a small genetic distance. © 2015 Blackwell Verlag GmbH.

  4. Versican V1 Overexpression Induces a Myofibroblast-Like Phenotype in Cultured Fibroblasts.

    Directory of Open Access Journals (Sweden)

    Jon M Carthy

    Full Text Available Versican, a chondroitin sulphate proteoglycan, is one of the key components of the provisional extracellular matrix expressed after injury. The current study evaluated the hypothesis that a versican-rich matrix alters the phenotype of cultured fibroblasts.The full-length cDNA for the V1 isoform of human versican was cloned and the recombinant proteoglycan was expressed in murine fibroblasts. Versican expression induced a marked change in fibroblast phenotype. Functionally, the versican-expressing fibroblasts proliferated faster and displayed enhanced cell adhesion, but migrated slower than control cells. These changes in cell function were associated with greater N-cadherin and integrin β1 expression, along with increased FAK phosphorylation. The versican-expressing fibroblasts also displayed expression of smooth muscle α-actin, a marker of myofibroblast differentiation. Consistent with this observation, the versican fibroblasts displayed increased synthetic activity, as measured by collagen III mRNA expression, as well as a greater capacity to contract a collagen lattice. These changes appear to be mediated, at least in part, by an increase in active TGF-β signaling in the versican expressing fibroblasts, and this was measured by phosphorylation and nuclear accumulation of SMAD2.Collectively, these data indicate versican expression induces a myofibroblast-like phenotype in cultured fibroblasts.

  5. Mosaicism for dominant collagen 6 mutations as a cause for intrafamilial phenotypic variability.

    Science.gov (United States)

    Donkervoort, Sandra; Hu, Ying; Stojkovic, Tanya; Voermans, Nicol C; Foley, A Reghan; Leach, Meganne E; Dastgir, Jahannaz; Bolduc, Véronique; Cullup, Thomas; de Becdelièvre, Alix; Yang, Lin; Su, Hai; Meilleur, Katherine; Schindler, Alice B; Kamsteeg, Erik-Jan; Richard, Pascale; Butterfield, Russell J; Winder, Thomas L; Crawford, Thomas O; Weiss, Robert B; Muntoni, Francesco; Allamand, Valérie; Bönnemann, Carsten G

    2015-01-01

    Collagen 6-related dystrophies and myopathies (COL6-RD) are a group of disorders that form a wide phenotypic spectrum, ranging from severe Ullrich congenital muscular dystrophy, intermediate phenotypes, to the milder Bethlem myopathy. Both inter- and intrafamilial variable expressivity are commonly observed. We present clinical, immunohistochemical, and genetic data on four COL6-RD families with marked intergenerational phenotypic heterogeneity. This variable expression seemingly masquerades as anticipation is due to parental mosaicism for a dominant mutation, with subsequent full inheritance and penetrance of the mutation in the heterozygous offspring. We also present an additional fifth simplex patient identified as a mosaic carrier. Parental mosaicism was confirmed in the four families through quantitative analysis of the ratio of mutant versus wild-type allele (COL6A1, COL6A2, and COL6A3) in genomic DNA from various tissues, including blood, dermal fibroblasts, and saliva. Consistent with somatic mosaicism, parental samples had lower ratios of mutant versus wild-type allele compared with the fully heterozygote offspring. However, there was notable variability of the mutant allele levels between tissues tested, ranging from 16% (saliva) to 43% (fibroblasts) in one mosaic father. This is the first report demonstrating mosaicism as a cause of intrafamilial/intergenerational variability of COL6-RD, and suggests that sporadic and parental mosaicism may be more common than previously suspected. © 2014 WILEY PERIODICALS, INC.

  6. Allelic Frequencies of 20 Visible Phenotype Variants in the Korean Population

    Directory of Open Access Journals (Sweden)

    Ji Eun Lim

    2013-06-01

    Full Text Available The prediction of externally visible characteristics from DNA has been studied for forensic genetics over the last few years. Externally visible characteristics include hair, skin, and eye color, height, and facial morphology, which have high heritability. Recent studies using genome-wide association analysis have identified genes and variations that correlate with human visible phenotypes and developed phenotype prediction programs. However, most prediction models were constructed and validated based on genotype and phenotype information on Europeans. Therefore, we need to validate prediction models in diverse ethnic populations. In this study, we selected potentially useful variations for forensic science that are associated with hair and eye color, iris pattern, and facial morphology, based on previous studies, and analyzed their frequencies in 1,920 Koreans. Among 20 single nucleotide polymorphisms (SNPs, 10 SNPs were polymorphic, 6 SNPs were very rare (minor allele frequency < 0.005, and 4 SNPs were monomorphic in the Korean population. Even though the usability of these SNPs should be verified by an association study in Koreans, this study provides 10 potential SNP markers for forensic science for externally visible characteristics in the Korean population.

  7. Chronic exposure of zinc oxide nanoparticles causes deviant phenotype in Drosophila melanogaster

    Energy Technology Data Exchange (ETDEWEB)

    Anand, Avnika Singh; Prasad, Dipti N.; Singh, Shashi Bala; Kohli, Ekta, E-mail: ektakohli@hotmail.com

    2017-04-05

    Zinc oxide nanoparticles (ZnO NPs) are commonly used nanomaterials (NMs) with versatile applications from high-end technologies to household products. This pervasive utilisation has brought human in the close interface with nanoparticles (NPs), hence questioning their safety prior to usage is a must. In this study, we have assessed the effects of chronic exposure to ZnO NPs (<50 nm) on the model organism Drosophila melanogaster. Potential toxic effects were studied by evaluating longevity, climbing ability, oxidative stress and DNA fragmentation. Ensuing exposure, the F0 (parent), F1, F2, F3 and F4 generation flies were screened for the aberrant phenotype. Flies exposed to ZnO NPs showed distinctive phenotypic changes, like deformed segmented thorax and single or deformed wing, which were transmitted to the offspring’s in subsequent generations. The unique abnormal phenotype is evident of chronic toxicity induced by ZnO NPs, although appalling, it strongly emphasize the importance to understand NPs toxicity for safer use.

  8. SNORD116 deletions cause Prader-Willi syndrome with a mild phenotype and macrocephaly.

    Science.gov (United States)

    Fontana, P; Grasso, M; Acquaviva, F; Gennaro, E; Galli, M L; Falco, M; Scarano, F; Scarano, G; Lonardo, F

    2017-10-01

    Prader-Willi syndrome is a complex condition caused by lack of expression of imprinted genes in the paternally derived region of chromosome 15 (15q11q13). A small number of patients with Prader-Willi phenotype have been discovered to have narrow deletions, not encompassing the whole critical region, but only the SNORD116 cluster, which includes genes codifying for small nucleolar RNAs. This kind of deletion usually is not detected by the classic DNA methylation analysis test. We present the case of a male patient with a mild Prader-Willi phenotype and a small deletion including SNORD116, diagnosed by methylation-sensitive multiplex ligation-dependent probe amplification (MLPA. The patient showed neonatal hypotonia, hyperphagia, obesity, central hypogonadism, hypothyroidism, strabismus. Stature and intellectual development are within the normal range. The presence of macrocephaly, observed in other cases of SNORD116 deletions as well, is uncommon for the classic phenotype of the syndrome. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  9. Transcription-induced DNA supercoiling: New roles of intranucleosomal DNA loops in DNA repair and transcription.

    Science.gov (United States)

    Gerasimova, N S; Pestov, N A; Kulaeva, O I; Clark, D J; Studitsky, V M

    2016-05-26

    RNA polymerase II (Pol II) transcription through chromatin is accompanied by formation of small intranucleosomal DNA loops. Pol II captured within a small loop drives accumulation of DNA supercoiling, facilitating further transcription. DNA breaks relieve supercoiling and induce Pol II arrest, allowing detection of DNA damage hidden in chromatin structure.

  10. DNA Methylation in Peripheral Blood Cells of Pigs Cloned by Somatic Cell Nuclear Transfer

    DEFF Research Database (Denmark)

    Gao, Fei; Li, Shengting; Lin, Lin

    2011-01-01

    To date, the genome-wide DNA methylation status of cloned pigs has not been investigated. Due to the relatively low success rate of pig cloning by somatic cell nuclear transfer, a better understanding of the epigenetic reprogramming and the global methylation patterns associated with development...... in cloned pigs is required. In this study we applied methylation-specific digital karyotyping tag sequencing by Solexa technology and investigated the genome-wide DNA methylation profiles of peripheral blood cells in cloned pigs with normal phenotypes in comparison with their naturally bred controls....... In the result, we found that globally there was no significant difference of DNA methylation patterns between the two groups. Locus-specifically, some genes involved in embryonic development presented a generally increased level of methylation. Our findings suggest that in cloned pigs with normal phenotypes...

  11. Eukaryotic DNA Replicases

    KAUST Repository

    Zaher, Manal S.; Oke, Muse; Hamdan, Samir

    2014-01-01

    The current model of the eukaryotic DNA replication fork includes three replicative DNA polymerases, polymerase α/primase complex (Pol α), polymerase δ (Pol δ), and polymerase ε (Pol ε). The primase synthesizes 8–12 nucleotide RNA primers that are extended by the DNA polymerization activity of Pol α into 30–35 nucleotide RNA-DNA primers. Replication factor C (RFC) opens the polymerase clamp-like processivity factor, proliferating cell nuclear antigen (PCNA), and loads it onto the primer-template. Pol δ utilizes PCNA to mediate highly processive DNA synthesis, while Pol ε has intrinsic high processivity that is modestly stimulated by PCNA. Pol ε replicates the leading strand and Pol δ replicates the lagging strand in a division of labor that is not strict. The three polymerases are comprised of multiple subunits and share unifying features in their large catalytic and B subunits. The remaining subunits are evolutionarily not related and perform diverse functions. The catalytic subunits are members of family B, which are distinguished by their larger sizes due to inserts in their N- and C-terminal regions. The sizes of these inserts vary among the three polymerases, and their functions remain largely unknown. Strikingly, the quaternary structures of Pol α, Pol δ, and Pol ε are arranged similarly. The catalytic subunits adopt a globular structure that is linked via its conserved C-terminal region to the B subunit. The remaining subunits are linked to the catalytic and B subunits in a highly flexible manner.

  12. Eukaryotic DNA Replicases

    KAUST Repository

    Zaher, Manal S.

    2014-11-21

    The current model of the eukaryotic DNA replication fork includes three replicative DNA polymerases, polymerase α/primase complex (Pol α), polymerase δ (Pol δ), and polymerase ε (Pol ε). The primase synthesizes 8–12 nucleotide RNA primers that are extended by the DNA polymerization activity of Pol α into 30–35 nucleotide RNA-DNA primers. Replication factor C (RFC) opens the polymerase clamp-like processivity factor, proliferating cell nuclear antigen (PCNA), and loads it onto the primer-template. Pol δ utilizes PCNA to mediate highly processive DNA synthesis, while Pol ε has intrinsic high processivity that is modestly stimulated by PCNA. Pol ε replicates the leading strand and Pol δ replicates the lagging strand in a division of labor that is not strict. The three polymerases are comprised of multiple subunits and share unifying features in their large catalytic and B subunits. The remaining subunits are evolutionarily not related and perform diverse functions. The catalytic subunits are members of family B, which are distinguished by their larger sizes due to inserts in their N- and C-terminal regions. The sizes of these inserts vary among the three polymerases, and their functions remain largely unknown. Strikingly, the quaternary structures of Pol α, Pol δ, and Pol ε are arranged similarly. The catalytic subunits adopt a globular structure that is linked via its conserved C-terminal region to the B subunit. The remaining subunits are linked to the catalytic and B subunits in a highly flexible manner.

  13. Genotypic and phenotypic diversity of Bacillus spp. isolated from steel plant waste

    Directory of Open Access Journals (Sweden)

    Chartone-Souza Edmar

    2008-10-01

    Full Text Available Abstract Background Molecular studies of Bacillus diversity in various environments have been reported. However, there have been few investigations concerning Bacillus in steel plant environments. In this study, genotypic and phenotypic diversity and phylogenetic relationships among 40 bacterial isolates recovered from steel plant waste were investigated using classical and molecular methods. Results 16S rDNA partial sequencing assigned all the isolates to the Bacillus genus, with close genetic relatedness to the Bacillus subtilis and Bacillus cereus groups, and to the species Bacillus sphaericus. tDNA-intergenic spacer length polymorphisms and the 16S–23S intergenic transcribed spacer region failed to identify the isolates at the species level. Genomic diversity was investigated by molecular typing with rep (repetitive sequence based PCR using the primer sets ERIC2 (enterobacterial repetitive intergenic consensus, (GTG5, and BOXAIR. Genotypic fingerprinting of the isolates reflected high intraspecies and interspecies diversity. Clustering of the isolates using ERIC-PCR fingerprinting was similar to that obtained from the 16S rRNA gene phylogenetic tree, indicating the potential of the former technique as a simple and useful tool for examining relationships among unknown Bacillus spp. Physiological, biochemical and heavy metal susceptibility profiles also indicated considerable phenotypic diversity. Among the heavy metal compounds tested Zn, Pb and Cu were least toxic to the bacterial isolates, whereas Ag inhibited all isolates at 0.001 mM. Conclusion Isolates with identical 16S rRNA gene sequences had different genomic fingerprints and differed considerably in their physiological capabilities, so the high levels of phenotypic diversity found in this study are likely to have ecological relevance.

  14. TRAIP promotes DNA damage response during genome replication and is mutated in primordial dwarfism.

    Science.gov (United States)

    Harley, Margaret E; Murina, Olga; Leitch, Andrea; Higgs, Martin R; Bicknell, Louise S; Yigit, Gökhan; Blackford, Andrew N; Zlatanou, Anastasia; Mackenzie, Karen J; Reddy, Kaalak; Halachev, Mihail; McGlasson, Sarah; Reijns, Martin A M; Fluteau, Adeline; Martin, Carol-Anne; Sabbioneda, Simone; Elcioglu, Nursel H; Altmüller, Janine; Thiele, Holger; Greenhalgh, Lynn; Chessa, Luciana; Maghnie, Mohamad; Salim, Mahmoud; Bober, Michael B; Nürnberg, Peter; Jackson, Stephen P; Hurles, Matthew E; Wollnik, Bernd; Stewart, Grant S; Jackson, Andrew P

    2016-01-01

    DNA lesions encountered by replicative polymerases threaten genome stability and cell cycle progression. Here we report the identification of mutations in TRAIP, encoding an E3 RING ubiquitin ligase, in patients with microcephalic primordial dwarfism. We establish that TRAIP relocalizes to sites of DNA damage, where it is required for optimal phosphorylation of H2AX and RPA2 during S-phase in response to ultraviolet (UV) irradiation, as well as fork progression through UV-induced DNA lesions. TRAIP is necessary for efficient cell cycle progression and mutations in TRAIP therefore limit cellular proliferation, providing a potential mechanism for microcephaly and dwarfism phenotypes. Human genetics thus identifies TRAIP as a component of the DNA damage response to replication-blocking DNA lesions.

  15. Developmental sculpting of social phenotype and plasticity.

    Science.gov (United States)

    Sakata, Jon T; Crews, David

    2004-04-01

    Early developmental variables engender behavioral and neural variation, especially in species in which embryonic environment determines gonadal sex. In the leopard gecko, Eublepharis macularius, the incubation temperature of the egg (IncT) determines gonadal sex. Moreover, IncT affects the sexual differentiation of the individual and, consequently, within-sex variation. Individuals hatched from eggs incubated at an IncT that produces predominantly males are more masculinized than same-sex counterparts from IncTs that produce predominantly females. Here we review how gonadal sex and IncT interact to affect behavioral, endocrinological, and neural phenotype in the leopard gecko and influence phenotypic plasticity following hormone administration or social experience. We discuss the hormonal dependence of sex- and IncT-dependent behavioral and neural morphological and metabolic differences and highlight the parallels between IncT effects in geckos and intrauterine position effects in rodents. We argue that the leopard gecko is an important model of how the process of sex determination can affect sexual differentiation and of selection forces underlying the evolution of sex ratios. Copyright 2004 Elsevier Ltd.

  16. Refractory versus resistant hypertension: Novel distinctive phenotypes

    Science.gov (United States)

    Dudenbostel, Tanja; Siddiqui, Mohammed; Gharpure, Nitin; Calhoun, David A.

    2017-01-01

    Resistant hypertension (RHTN) is relatively common with an estimated prevalence of 10-20% of treated hypertensive patients. It is defined as blood pressure (BP) >140/90 mmHg treated with ≥3 antihypertensive medications, including a diuretic, if tolerated. Refractory hypertension is a novel phenotype of severe antihypertensive treatment failure. The